Livre Blanc Volume 1 Spectrophotomètrie UV-Visible et ... · Livre Blanc Volume 1...

Livre Blanc Volume 1

SpectrophotomètrieUV-Visible et Infrarouge

« Plus de 70 pages d’applications UV-Vis et FTIR..»

©Shimadzu Corporation 2013

Fondé en 1875, Shimadzu est un groupe multinational japonais de 3 milliards de dollars côté à la bourse de Tokyo. Avec près de 10000 employés dans le monde Shimadzu Corporation regroupe trois acti-vités principales : l’instrumentation analytique et physique, le diagnostic médical et l’aéronautique. Présent dans plus de 100 pays, Shimadzu est un fabricant d’instrumentation analytique et d’équi-pement de contrôle environnemental. En effet Shimadzu dispose d’une large gamme d’instru-ments analytiques : Chromatographie liquide et gazeuse, spectrométrie de masse, une gamme complète de Maldi, des robots de préparation, spectrométrie UV-Vis, FTIR, analyse élémentaire.

Shimadzu est aussi un fabricant mondial de machines de caractérisation de matériaux et d’es-sais mécaniques. En effet Shimadzu offre une large gamme de machines électromécaniques ou hydrauliques, statiques ou dynamiques, de traction, compression, flexion, pelage, cisaillement fatigue…

Fondé en 1968 en Allemagne, Shimadzu Europe fourni des solutions analytiques aux scientifiques européens. Aujourd’hui Shimadzu Europe représente plus de 9 filiales et 13 distributeurs répartis à travers l’Europe dont Shimadzu France, filiale de plus de 65 personnes en forte croissance depuis sa création en 2002.

A propos de Shimadzu

Shimadzu France possède en ses locaux à Noisiel (77) plusieurs showrooms représentant chaque gamme, d’un laboratoire de préparations d’échantillons et de salles de formation et de réunions.Notre laboratoire vient en complément des 1500m² de laboratoire disponibles à notre siège eurôpéen basé à Duisburg (Alle-magne) récemment inuagurés.

Laboratoires de démonstration

UV-1800Allant bien au-delà des normes de la pharmacopée, l’UV-1800 offre un éventail de fonctionnalités complètes et conviviales. Il peut être configuré en tant qu’instrument autonome ou contrôlé par PC.

• Avec la résolution la plus élevée de sa catégorie (1 nm), il répond aisément à la résolution de longueur d’onde requise.

• Montage optique Czerny-Turner dans un système compact atteignant une lumière parasite faible, une excellente répétabilité de la longueur d’onde et une grande stabilité de la ligne de base.

• Mesurant seulement 450 mm de large, c’est l’un des instruments les plus compacts de sa catégorie.

• Les clés USB permettent aux utilisateurs d’analyser les données sur un PC séparé en utilisant le logiciel UVProbe standard qui peut également permettre le contrôle total de l’instrument.

UVmini-1240L’UVmini-1240 est un instrument d’analyse à faible coût et très peu encombrant.

• Plage de mesure de 190 nm à 1100 nm.

• Largeur de bande spectrale de 5 nm.

• Logiciel de contrôle embarqué avec grand écran LCD.

• Les opérations incluent la photométrie, l’acquisition spectrale et la quantification.

UV-3600 / SolidSpec-3700Les spectrophotomètres UV-3600 et SolidSpec UV-VIS-NIR ont été conçus pour les mesures nécessitant la sensibilité la plus élevée possible. Ils incluent tous les deux une technologie à trois détecteurs en transmission et en réflexion, permettant un niveau de performance maximale de l’UV au proche Infra Rouge.

• Plage de mesure possible de 185 nm à 3300 nm (UV-3600) ou 165-3300 nm (SolidSpec-3700).

• Trois détecteurs : PMT, InGaAs et PbS.

• Grand compartiment d’échantillons 900x700x350 mm (SolidSpec-3700)

• Le chemin optique vertical permet un positionnement horizontal de l’échantillon. (SolidSpec-3700).

RF-5301Instrument de fluorescence de pointe pour la recherche et le développement, les analyses de routine et l’enseignement.

• Gamme dynamique de mesure de fluorescence basée sur la sélection du détecteur en deux étapes.

• Optique double pour les mesures du spectre d’excitation et des émissions, notamment l’obturation et le choix de deux taux de largeur de bande et de mesure.

Gamme UV-VIS/NIRFL

UORE

SCEN

CE

UV-2600/2700Gâce à une toute nouvelle génération de réseaux Low-Ray-Ligh développé par Shimadzu, les spectrophotomètres UV-2600/UV-2700 vous permettront d’atteindre des niveaux de performance inégalés en UV-Visible. Ils regroupent tout le savoir-faire de Shimadzu depuis plus de 65 ans avec un minimum d’encombrement.

• L’UV-2600 est le système universel quels que soient vos échantillons. Avec une linéarité de 0 à 6 Abs et un bruit de fond inférieur à 0,00003Abs, il vous permettra la mesure de tout produit, de la plus infime quantité au produit les plus opaques. Couplé à notre nouvelle sphère d’intégration ISR-2600Plus, il vous ouvrira les portes du Proche Infra Rouge grâce à une plage de mesure exceptionnelle allant de 185 à 1400 nm.

• L’UV-2700 permet d’analyser vos échantillons les plus opaques et les plus absorbants. Avec son double monochromateur, il atteint une linéarité maximale jusqu’à 8 Abs entre 190 et 900 nm, permettant ainsi de travailler sur vos échantillons sans dilutions supplémentaires.

http://www.shimadzu.fr/spectroscopie-uv-visible-nir

IRAffinityL’IRAffinity est muni d’un système innovant et unique de protection de l’optique. Grâce à un compartiment étanche muni d’une membrane hydrolytique, l’IRAffinity est protégé de l’humidité en permanence, éteint comme allumé, sans aucune intervention manuelle tout au long de sa durée de vie. Ajouté à une optique de haute qualité, il est le spectrophotomètre FTIR le plus performant de sa catégorie.

• Résolution 0,5 cm-1 et dimensions compactes.

• Rapport signal/bruit 30000:1 en utilisant une source de source céramique haute densité, et le détecteur DLATGS à température contrôlée.

AIM-8800 MicroscopeLe microscope infrarouge AIM-8800 permet un contrôle aisé des étapes et un réglage de l’ouverture à partir de l’écran du PC.

• Transmission, réflexion et réflexion totale atténuée.

• L’ouverture automatique optimise l’éclairage infrarouge.

• L’étape X-Y automatique simplifie le positionnement de l’échantillon.

• Mise au point automatique et centrage automatique.

IRPrestige-21• L’IRPrestige-21 offre des possibilités d’extension rapides allant de l’infrarouge proche, à

l’infrarouge moyen jusqu’à l’infrarouge lointain.

• Le système optique unique avec des miroirs en or permet d’atteindre des performances de recherche comme un rapport signal/bruit de 40000:1 et une résolution de 0,5 cm-1.

• Optimisation en temps réel de l’interféromètre, avec un alignement dynamique avancé.

GladiATR 10Echantillons durs

(Diamant Massif Naturel)

HATR 10Liquides, Polymères, Pâtes

MiRacle 10ATR Universel

ATR

ATR Shimadzu vous propose sa gamme d’ATR série 10 dernière génération, optimisée pour l’optique de l’IRAffinity-1. Chaque ATR est équipé d’une presse avec dé-brayage automatique ainsi qu’un positionnement Plug and Play. Un capteur de pression est aussi disponible en option.

Gamme FTIR

http://www.shimadzu.fr/spectroscopie-ftir-ft-nir

Shimadzu News Article Collection

Volume 1

UV Ultra violet Visible and Near Infrared

Spectroscopy

Blank page

APPLICATION Shimadzu News 1/2011

6

Windows in the spotlight

In the Hitchcock movie ‘RearWindow’, the window open-ing onto the courtyard was

open most of the time. In thisway, James Stewart could toleratethe summer heat of 1954 and alsosee the murder happening in hisneighbor’s house. But how, inthese times of global warming,can a comfortable indoor climatebe provided during the hot sum-mer months? Climate changeplaces increasingly higherdemands not only on environ-mental behavior, but also on ourway of life.

In addition to the use of air con-ditioning systems, efficient use ofbuilding materials is also helpful,and windows play a significantrole. Use of selected flat glasshelps to prevent as much heat aspossible from entering the room,while still allowing enough lightto pass.

Light transmittance in flat glass

The quality of flat glass in thebuilding industry is testedaccording to DIN EN 410: ‘Glassin building – determination ofluminous and solar characteristicsof glazing.’ In Japan JIS R 3106 isapplied: ‘Testing method onTransmittance, Reflectance andEmittance of Float Glasses andEvaluation of Solar Gain Coeffi-cient.’ Testing includes lighttransmittance and reflectance inthe visible spectral range of 380to 780 nm, solar transmittanceand reflectance between 300 and2,500 nm as well as normal emit-tance in the 2,000 - 400 cm-1

range (5,000 - 25,000 nm as IRmeasurement).

In the examples shown here,measurements were carried outaccording to the Japanese indus-trial standard JIS R 3106. Formeasurements in the visible

Determination of the transmittance and reflectance of

Figure 1: Display of the ‘Daylight’ software with calculation of results

Table 1: Instrument measurement parameters according to JIS R3106

Close to parallel

beam of light

incident from the

normal direction

The air layer is

used as the

standard sample,

and its spectral

transmittance is

taken to be 1

300 - 2,500 nm

< 300 nm, 5 nm max. 380 - 780 nm, 10 nm

max. 780 nm and higher, < = 50 nm max.

380 - 780 nm

10 nm max.

Close to parallel

beam of light

incident from the

radiation slit at

an angle not

exceeding 15

degrees

Specular reflector

of reflectance

specified by the

absolute reflec-

tance measurement

method (sample 1),

or specular reflec-

tor of reflectance

specified by

comparison with

sample 1

Close to parallel

beam of light inci-

dent from the radia-

tion slit at an angle

not exceeding 15

degrees.

Close to parallel

bundle of rays

incident from nor-

mal direction

Specular reflector

of reflectance

specified by the

absolute reflec-

tance measurement

method (sample 1),

or specular reflec-

tor of reflectance

specified by

comparison with

sample 1

IRAffinity-1 FTIR

spectrophotometer

and SRM-8000, an

accessory for spec-

ular reflection

5 - 25 μm

4 cm-1

Light radiation

at an angle not

exceeding 15

degree

Aluminum-coated

mirror with

certified absolute

reflectance (float

flat glass with

vacuumdeposited

aluminum film)

Visible range Solar range

Transmittance Reflectance Transmittance Reflectance

NormalEmittance

UV-3600 UV-VIS-NIR spectrophotometer ISR-3100 Integration sphere

(Ulbricht sphere) with three detectors

Analytical

instrument

Measurement

wavelength

range

Resolution

Incident light

conditions

Standard sample

for comparison

Close to parallel

beam of light

incident from the

normal direction

The air layer is

used as the stan-

dard sample and

its spectral trans-

mittance is taken

to be 1

7

APPLICATIONShimadzu News 1/2011

– UV-VIS spectroscopy

as well as in the solar range, Shimadzu’s UV-3600 with theISR-3100 integration sphere wasused. The company’s IRAffinity-1with a directed SRM-8000 reflec-tance unit was applied for thedetermination of the normalemittance. Both instruments canbe used in the infrared range(thermal radiation). The UV-VIS-NIR UV-3600 system covers thenear infrared (NIR) range, andthe FTIR system covers the mid-infrared range (MIR).

According to JIS R 3106, themeasurement parameters can belisted as shown in table 1.

Evaluation of results is carriedout in accordance with JIS R3106. A spreadsheet program oroptimized software can be usedfor calculation. Parameters forthe visible (te) and solar (tv)transmittance as well as re-flectance determined as pe andpv, can be listed using Shimadzu’s‘Daylight’ software.

The following standard valuescan be determined optionally (see table 3).

As an example, the visible andsolar transmittance and the chro-maticity values for five types ofglass were calculated according to

flat glass in the building industry according to DIN EN 410/JIS R 3106

Table 3: Tristimulus values which can be determined using the

‘Daylight’ software

Figure 2: UV-VIS-NIR spectra of glasses in transmittance: black corresponds to

Glass-0, red to Glass-1, blue to Glass-2, green to Glass-3 and violet to Glass-4

Table 2: Visible and solar transmittance for five flat glass samples with Standard Illuminant D65 calculated under an observation

angle of 2°

JIS R 3106. The values are listedin table 2 and the UV-VIS-NIRspectra are shown in figure 2. TheStandard Illuminant D65 wasselected for the calculation as thisrepresents daylight according tothe CIE (International Commis-sion on Illumination). An angleof observation of 2° was selected.The results presented show just asmall extract of the determinationrange. [1][2]

[1] Shimadzu Application News No.

A396, ‘Daylight Transmittance

Application Data of Glass Plate’

[2] Shimadzu Application News No.

A404, ‘Glass Plate Analysis in

Accordance with JIS R 3106’

1/nm

T%

-9,112

0

50,000

100,123

300 500 1,000 1,500 2,000 2,100

x

1

2

3

4

5

86.756

36.167

31.559

9.164

8.108

90.672

29.913

27.223

1.527

1.389

85.830

28.586

25.912

9.124

8.267

90.672

29.912

27.223

1.525

1.387

98.478

60.079

54.484

48.573

44.027

GLASS-0

GLASS-1

GLASS-2

GLASS-3

GLASS-4

Sample number te tv y z File name

te, pe

dte, tpe

tv, pv

dtv, dpv

User

X, Y, Z

dX, dY, dX

x, y

dx, dy

dWL

Daylight transmittance, reflectance (JIS R 3106)

Difference of daylight transmittances, reflectances

(JIS R 3106)

Visible light transmittance, reflectance

(JIS R 3106, JIS Z 8722)

Difference of visible light transmittances, reflectances

(JIS R 3106, JIS Z 8722)

Calculated by user defined weighted factor file

Tristimulus Value X, Y, Z (JIS Z 8722)

Difference of X, Y, Z

Chromaticity Coordinates x, y (JIS Z 8722)

Difference of x, y

Dominant Wavelength dWL (JIS Z 8701)

We will gladly send you further information. Please note

the appropriate number on your reply card or order via the

News App resp. News WebApp. Info 392


4

‘Photons’ and ‘holes’ are termscommonly used in photovoltaics.Rays of sunlight (photo-effect)hitting a solid-state body such asa solar cell, will release positiveand negative charge carriers. Asolar cell contains p-conductingand n-conducting semiconductorlayers, resulting in p-n transitionswhere these two layers meet.

Free electrons and holes becomeactivated based on their chargedifferences. When p-n-transitionsoccur, electrons and holes formpairs. As a result, the n-dopedarea acquires a positive chargedue to lack of electrons. At the

of solar energy due to back-reflection from the surface (bycomparison: a simple glass sur-face already reflects 4 % of theincident energy).

The finite supply of fossilfuels and high costs ofenergy as well as CO2

emissions have recently brought awell-known alternative technolo-gy back into prominence: photo-voltaics, which harvests energyfrom the sun. Photovoltaics con-verts solar energy instantly intoelectrical energy.

An important component of solarenergy technology is silicon,which is one of the most abun-dant chemical elements. Underthe influence of solar radiation,silicon exhibits electrical proper-ties and acts as a semiconductor.

same time, the p-doped area islacking positively charged holesand acquires a negative charge.Between these oppositely chargedareas an electrical field is generat-ed which becomes stronger asmore electron-hole pairs areformed.

Sunlight activates the flow ofelectrons, which is the first steptowards generation of an electri-cal current. A voltage can betapped off the electrodes. Figure 1shows a schematic representationof a solar cell.

Anti-reflection film increasesefficiency

Although the efficiency of a solarcell has often been the subject offierce criticism, the performanceof solar cells has been optimizedcontinuously. One step in theoptimization of solar cells is toensure that they can accommo-date all incident solar irradiation.High conversion yields can beobtained by coating the solar cellsurface with an anti-reflectionfilm. This film prevents any loss

Figure 1: Configuration of a solar cell

Figure 2: Schematic representation of the polarization levels of non-polarized and

polarized light

Figure 3: Schematic representation of the planes of incidence of p-polarized and

s-polarized light

Figure 4: Shimadzu’s UV-3700 spectro-

photometer, a UV-VIS-NIR system with

three-detector configuration and large

sample compartment

This application note discussesthe reflectance measurement ofthe anti-reflection coating of aSi3Ni4 (silicon nitrate) solar cell.In this study, the spectrum in theUV-VIS-NIR range (300 – 2200nm) was investigated using themethod of absolute specularreflectance.

5


0.0

20.0

40.0

60.0

80.0

100.0

300.0 500.0 1000.0 1500.0 2000.0

nm

R %

0.0

20.0

40.0

60.0

80.0

100.0

300.0 500.0 1000.0 1500.0 2000.0

nm

R %

Figure 7: UV-VIS-NIR spectra recorded using s-polarized light and specular

total reflectance

Figure 8: UV-VIS-NIR spectra recorded using p-polarized light and absolute

specular total reflectance

Figure 5: Absolute specular reflec-

tance attachment with variable angle

of incidence

Figure 6: The inner compartment of

the UV-3700 with built-in variable abso-

lute reflectance attachment. The solar

cell sample is placed in the holder.

Absolute reflectance is that partof specular reflectance withoutthe contribution of diffuse orstray-light phenomena. In thisseries of measurement, incidentangles of 5 – 60° were used.When carrying out this measure-ment technique under higherangles of incidence, polarizationeffects can occur. A polarizer wastherefore used to generate p- ands-polarized light. Depending onthe angle of incidence and therefractive index, non-linear influ-ences on the reflectance resultswere observed.

The component of light on theplane of incidence is called p-polarized light (parallel-polar-ized); s-polarized light (vertical-ly-polarized) strikes the plane ofincidence vertically. The plane ofincidence is the direction of theincident light and the line per-pendicular to the reflective sur-face.

For small angles of incidence,hardly any difference in reflec-tion with respect to polarizationcould be observed. For largeangles of incidence the influenceof polarization increased and apolarizer should therefore beused during measurement.

Highest sensitivity using the UV-3700

Using this reflectance technique,it is possible to find the optimumangle of solar radiation incidenceon a silicon surface in order toobtain the highest conversionyield for the solar cell. The fol-lowing measuring results wereobtained using Shimadzu’s UV-3700. Its three-detector-configu-ration enables highly accurateand sensitive measurement in the

300 – 2200 nm measuring range.The large sample compartment ofthe UV-3700 facilitates the inte-gration of a variable reflectanceattachment with integrationsphere.

Measurements were carried outusing s- and p-polarized light.The measured results which weredependent on the angles of inci-dence, are shown in Figures 7 and 8.

The following angles of incidencehave been selected for measure-ments and the associated spectraare indicated according to linecolour: green 5°, red 15°, blue30°, black 45° and purple 60°.

The results for the s-polarizedlight and the p-polarized light areshown in the above figures. Theleast amount of reflection occursin the range of 500 to 600 nm.From the measured results it isapparent that the anti-reflectionfilm minimizes reflection mainlyin the visible wavelength range.

We will gladly send you further informa-

tion. Please note the appropriate number

on your reply card. Info 347

CNT

1.3 - 1.4

45

Looking into tubes with spectroscopy – The world of “carbon nanotubes” »2

What is purging? – POC measuring process »5

Low flows – high potential –prominence Nano-pump »8

Analysis of primary amino acids »12

Highly efficient – Determination of environmental contaminants »14

A global challenge – State-of-the-art

water analysis »16

How paper tells medieval stories – Antiquarian books as a source of

environment historical data »18

FTIR spectrometry in traditional Chinese medicine »19

Antarctic expedition »10

Frost & Sullivan awards Shimadzu Europa »9

Silver Jubilee Medal 2008 for Luigi Mondello »17

Installation kit for HPLC systems »27

Teddy bears and Jelly bears – Universal testing machines »20

One box for all purposes – New valve techniques for GC »22

More is better – MOC-120H »24

prominence UFLC-XR in MS software »27

A matter of taste – Determination of

dimethyl sulfide in wort »25

European Symposium on Atomic Spectroscopy »26

Dinner for all – 40 years Shimadzu Europe »28

APPLICATION

INTERNAL

ANNIVERSARY

CONFERENCE

READ FOR YOU

PRODUCTS

TELEGRAM

The “best in class”

FTIR system –

IRAffinity-1: Multiple

Choice.

Looking into tub


2

What are nanotubes?

A nanotube is a cylindrical struc-ture with a diameter within thenanometer-range and several millimeters in length. There aretwo main types: the SWNT (sin-gle-walled nanotube) and theMWNT (multi-walled nanotube).Nanotubes belong to the familyof fullerenes; they are cylindricaland typically capped with a half-sphere at one end of the cylinder.The known and various manifes-

The world of “carbon nanotubes”

Carbon nanotubes (CNTs)are tube-like structures thatconsist entirely of carbon(Figure 1). They were dis-covered in 1991 by SumioIijima.

Depending on the struc-ture of the tube, carbonnanotubes are either

metallic or semiconducting. Dueto these two important character-istics, CNTs are of great interestto the industry.

Figure 1: Picture of a nanotube

Table 1: Comparison of CNT with steel properties

Property

Density [g/cm3]

Tensile strength [GPa]

Steel

7.8

2

IMPRINTShimadzu NEWS, Customer Magazine of

Shimadzu Europa GmbH, Duisburg

Publisher:

Shimadzu Europa GmbHAlbert-Hahn-Str. 6 -10 · D - 47269 DuisburgPhone: +49 - 203 - 7687- 0 Telefax: +49 - 203 - 766625Email: [email protected]: www.shimadzu.eu

Editorial Team:

Uta Steeger · Phone: +49 - 203 - 7687- 410 Ralf Weber, Angela Bähren

Design and Production:

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Circulation:

German: 7,600 · English: 22,000

©Copyright:

Shimadzu Europa GmbH, Duisburg, GermanyNovember 2008

Windows is a Trademark of Microsoft

Corporation

es with spectroscopy

3


of the excellent mechanical prop-erties of CNTs is the density ofapproximately 1.3 – 1.4 g/cm3 incombination with an extraordi-nary tensile strength that exhibitsa 135-fold better density/tensilestrength ratio than steel (Table 1).

For the electronics industry, theelectrical charge density as well asthe thermal conductivity, whichat room temperature amounts tovirtually twice that of diamond(CNT 6000 W/m*K and diamond3320 W/m*K), are of interest.

The CNT optical transitionsoccur in the following sequence,as seen from the low energeticside of the spectrum: semicon-ductor – semiconductor – metal.On decreasing nanotube diame-ter, the absorptions will shift tothe higher energetic side of thespectrum. At increasing diameter,

Figure 2: Various manifestations of carbon (carbon allotropes): (a) diamond,

(b) graphite, (c) lonsdaelite, (d) C60, (e) C540, (f) C70, (g) amorphous carbon and

(h) carbon nanotube

Figure 3: Depending on the rolling mechanism, nanotubes with either metallic pro-

perties (armchair) or semiconductor properties are formed. During rolling the point

labelled (0,0) will join with another point (10.0, 10.5 or 10.10). The names “armchair”

and “zigzag” originate from the way the C-atoms are rolled along the axis connecting

the two points (emphasized atoms).

the absorption will result inbroader absorption bands.

Spectroscopy visualizesenergy states

Energy states can be visualizedwith the aid of UV-VIS/NIRspectroscopy. Several examples ofdifferent CNT-states are shownbelow:

spectroscopy is the method ofchoice for the analysis of thethree signals, as these will be displayed very sharply. This isattributed to the nature of theCNTs and is also described as“Van Hove singularities”.

A typical SWCNT spectrum isshown in Figure 4 (Page 4). M1 isthe metallic property and S1 andS2 represent the semiconductorproperties. Figure 5 shows thespectrum in the nanometer scale,whereas Figure 4 is expressed inelectron Volts (eV). Typically, the

signals are found between 0.5 and1.2 eV for SWCNTs. This corre-sponds to a tube diameter of 0.8up to 1.5 nm. �

tations of carbon, the origin ofCNTs, are shown in Figure 2.

A carbon nanotube is formedwhen a graphene layer joinstogether. Depending on the waythe hexagonal graphene networkis “rolled up”, CNTs with eithermetallic or semiconductor char-acteristics are formed. The threedifferent variants are clearlydefined. They are called zigzag,chiral or armchair and displaydifferent characteristics. The arm-chair type, for instance, is alwaysmetallic.

Of interest to the electronics industry

In theory, metallic nanotubespossess an approximately 1000-fold better “electrical currentdensity” when compared withmetals like silver or copper. One


4

The CNTs are assembled accord-ing to various procedures. Twoprocedures for the assembly HiP-Co (high-pressure CO conver-sion) (Figure 6) and CoMoCAT(Figure 7) result in different spec-tra.

At the University of Oklahoma,USA, the catalytic CoMoCATprocedure was developed inorder to manufacture SWCNTsin high yields (Figure 7). Thespectra shown were recordedusing Shimadzu’s SolidSpec-3700,whereas the instrument wasequipped with an integratingsphere.

Another example is transmissionspectroscopy carried out in solu-tion using the UV-3600. Thespectrum of the liquid sample –CNT diluted in N-methyl-2-pyrrolidon (NMP) – is shown inFigure 8. The fluid matrix exhib-its better stray light behavior.This leads to improved quality ofthe spectra and higher detectionlimits in the UV-VIS/NIR meas-uring range.

The “Van Hove singularities” canbe identified clearly in the spec-trum. The signals and the conver-sion to electron volts are listed inTable 2.

The suspension consisting ofCNT and NMP was filled into a10 mm quadratic quartz cell. The pure NMP solvent was usedas reference and was filled intothe reference position of the Shimadzu UV-3600.

Literature and references

1. Recommended practice guide for

Nano Carbon tubes characterization,

Michael E. Itkis, Robert C. Haddon,

University of California

2. Mr. Iijima is Professor in Meijo

University and belongs to AIST and

NEC Corporation

Instrument and sample preparation

Instrument: UV-3600 with threedetectors

Slit width: 1 nmScan speed: mediumCell: two 10 mm quartz

cells

The measurement took place inthe range of 350 to 1700 nm asthe dilution agent exhibits astrong absorption in the wave-length range > 1700 nm.

Conversion into electron volts isnecessary for the determinationof the CNT diameter. The DOS(electronic density of states) canbe calculated according to fixedparameters:

S1 = 2a�/dS2 = 4a�/dM1 = 6a�/d

Where a = carbon-carbon bondlength (nm), � = energy in the p�

orbitals (~2.9 eV) and d = SWNTdiameter.

0

0.05

0.1

0.15

0.2

0.25

0.3

0.35

0.5 1.0 1.5 2.0 2.5 3.0 3.5

Photon Energy [eV]

Abs

0.000

0.100

0.200

0.300

0.400

0.500

190.0 500.0 1000.0 1500.0 2000.0 2500.0

Wavelength [nm]

Abs

Figure 4: Typical spectrum of an SWCNT (single-wall carbon nanotube) wherein the

absorption is plotted against the energy scale

Figure 5: UV-VIS/NIR spectrum of an SWCNT dispersed onto a quartz substrate.

Measurement was carried out using an integrating sphere.

[eV]

0.8613

0.9493

1.0621

1.1506

1.2486

[nm]

1439.5

1306

1167.4

1077.6

993

CNT 1-D

Figure 6: Representation of the

UV-VIS/NIR spectrum of a HiPCo (high-

pressure carbon monoxide) measured

using Shimadzu’s UV-3700

Figure 7: Representation of the

UV-VIS/NIR spectrum of a CoMoCAT

measured using Shimadzu’s UV-3700

Table 2: Analytical wavelength of the

carbon nanotube (CNT) in the UV-VIS/NIR

range

Figure 8: Representation of the CNT

spectrum in the NMP matrix measured

using Shimadzu’s UV-3600

0.00

1.50

0.75

350.0 1075.0 1800.0

nm

Abs

0.00

0.03

0.01

350.0 1075.0 1800.0

nm

Abs

0.2278

0.5000

1.0000

1.5381

305.19 1000.00 1700.00

nm

Abs

Blank page


4

Mineral water in a diffe

With 130 liters consump-tion per capita and peryear in Germany, they

are an important food source inour daily nutritional intake, sup-plying important minerals andtrace elements, both sparklingand still – mineral waters. Theirlabels are their trademarks andbusiness cards as they containspecific information on the con-tents. These labels testify to thequality of the mineral waterspring where the water comesfrom. Interestingly, the labels donot mention the actual test dateof the water, as the informationcannot be printed fast enough onthe labels. When checking theimprints on the bottles, one findsa date that differs by severalmonths from the mineral analysisof the well itself.

Drinking water is one of themost tested food sources. It issubject to continuous monitoringaccording to national and Euro-pean drinking water ordinances.Monitoring includes, among oth-ers, the analysis of minerals andtrace elements. These analyses are

usually carried out using atomicabsorption spectroscopy or ICP-OES methods. The total organiccarbon content is determinedusing TOC methods; nitrate andnitrite content can be determinedusing UV spectroscopy. Manyfoods are also characterized usingNIR spectroscopy, a measuringtechnique that is not quite assuitable for the identification ofmineral water.

UV spectroscopy also for the identification ofspring waters

The following application showsthat UV spectroscopy is suitablenot only for the determination ofelements and their salts, but alsofor straightforward identificationof spring waters. Waters fromvarious sources exhibit differ-ences and these differences can berevealed using absorption spec-troscopy and UV spectra, as wellas NIR measurements. As miner-al waters are all transparent in thevisible range, special attentionwas focused on the ultravioletand near-infrared ranges.

Quality control using UV spectroscopy

0.0

0.5

1.0

1.5

1.8

190.0 200.0 220.0 240.0 250.0 nm

Abs

Figure 1: Ultraviolet spectra of four different still mineral waters

5


Sample (theoretical) A Water (%) B Water (%) C Water (%)

10

40

20

C Water (%)

9.72

39.92

19.91

1

2

3

Sample (calculated)

1

2

3

10

30

20

A Water (%)

10.36

30.12

20.41

80

30

60

B Water (%)

79.90

29.96

59.68

Standard A Water (%) B Water (%) C Water (%)

1

2

3

4

5

20

50

30

0

50

30

20

50

50

0

50

30

20

50

50

rent light

Analysis in the ultra-violet range

Spectra of four store-bought stillmineral waters were measured inthe ultraviolet range of 190 - 250nm using Shimadzu’s UV-3600spectrophotometer. The result isshown in figure 1. A 10 mm thickquartz cuvette was used for themeasurements, and deionizedwater was used and measured asreference sample. When compar-ing the spectra, one can see aclear difference between thecurves, which can be attributed tothe individual characteristics ofthe mineral waters.

Water in the near infrared range

In the infrared range of 100 -1800 nm using a layer thicknessof 2 mm, the same waters did notexhibit any individual character-istics for the water absorptionsignal at 1440 nm (Figure 2). Thisis due to the strong absorptionby water in the NIR range, as canbe seen in figure 2. This value lies within the linear measuring

Table 1: Mixing ratios for measurements using the UV-3600 of three different still

mineral waters (see Figure 3).

Table 2: The result of a model calculation with respect to four analytical wavelengths

and three samples

0.0

1.0

2.0

3.0

3.3

1000.0 1200.0 1400.0 1600.0 1800.0nm

Abs

Figure 2: Near-infrared spectra of 4 still mineral waters, measured using a 2 mm

thick cuvette

0.0

0.5

1.0

1.5

1.7

190.0 200.0 220.0 240.0 250.0

nm

Abs

Figure 3: Five UV absorption spectra of mixtures of three mineral waters.

The different mixing ratios are listed in Table 1; red = standard 1, blue = standard 2,

black = standard 3, green = standard 4, brown = standard 5

range of the instrument. The dif-ferences among the water samplescannot be determined using NIR,as the NIR range is representedby -CH, -OH and -NH vibra-tions. Minerals are inorganic innature.

Multiple linear regression

To verify the assumption that theultraviolet range reveals differ-ences, multiple linear regressionwas carried out at four analyticalwavelengths in the UV range.The wavelengths 200, 205, 210and 215 were selected and a cali-bration was carried out. Calibra-tion standards were prepared bymixing 3 mineral waters (Tab. 1).Figure 3 shows five UV spectraof these standards.

The result from this straightfor-ward multiple-component analy-sis model (Table 2) shows that themineral waters are distinguish-able. It should be noted that mul-tiple-component analysis can alsobe applied in the UV spectros-copy range of 190 to 250 nm.

This evidently requires an instru-ment technology which stilldelivers reliable results in thisrange. Shimadzu’s UV-VIS-NIRUV-3600 spectrophotometer isparticularly suitable for thesetypes of analysis.

We will gladly send you additional

information. Please enter the cor-

responding number on the reply

card or order via Shimadzu’s News

App or News WebApp. Info 396

The purple colored UV-VIS-NIRspectrum shows filtering at 800nm and an inflection point at awavelength of 737 nm, as well asthe highest reflectance in the NIRrange of 2,500 up to 750 nm com-pared to other textiles. The textilesample shown in figure 2 is espe-cially UV reflecting (orange-redspectrum). Both surfaces (silverand cream colored surface) showweaker reflectance in the visiblerange. These surfaces show goodheat-reflecting properties in theNIR range.

The spectrum of the grey sample(light brown spectrum) exhibitseven lower thermal radiation aswell as reduced light transmissi-bility in the visible range. This iscertainly a suitable textile fordarkening a room. Furthermorethe various organometallic com-pounds in combination with thepolymer generate a NIR spec-trum that enables identificationof the polymer. The fibers consistof polyester plus an ‘intelligent’metallic dye.

With the Shimadzu UV-3600triple-detector system and theassociated ISR-3100 integratingsphere with three detectors, the


6

light, respectively sunlight. Forthis purpose, materials with twodifferent surfaces have been in-vestigated. An example of a silverand a cream colored surface isshown in figure 2.

The behavior of the material,when light, heat and strong UVradiation hits the textile, is ofspecial interest.

Instrument and measuringconditions

For analysis, a UV-VIS-NIRinstrument was used. The UV-3600 spectrophotometer con-tains an integrating sphere (Ul-bricht sphere). It is equipped withthree detectors in order to meas-ure the entire range from UV toNIR with higher sensitivity.• Instrument: UV-3600• Integrating sphere: ISR-3100

with three detectors.

Measurements were carried outunder 0° reflectance conditions.This method leads to the determi-nation of diffuse reflectance ofthe sample surface. Figure 3 pro-vides a short description of thereflectance of light at a surfacewith specularly reflected light and

I n Europe the previousdecade was the warmestsince recording of tempera-

ture in Germany which approxi-mately began 130 years ago. Tem-peratures have also been risingaround the rest of the globe. Thisdoes not lead to energy-savinghowever, as air conditioning sys-tems are used during the summerto cool living spaces and workplaces. These require much elec-tricity, which also increases costs.Ideas are welcome on how toreduce warming of living andworkspaces resulting from solarirradiation.

Suitable textiles already exist forcars or commercial buildings andoffices. Glass panes coated withlight-reflecting layers are alsoavailable to reduce heat or toprovide anti-glare shields forwindows. At the same time,roller blinds or sun blinds canserve as screens.

The surfaces of these types oftextiles, as used for sun blinds,are shown in figure 2.

This article shows how textilescan be used to allow or to blockdifferent wavelength ranges of

diffusely transmitted light, at anincident light angle of 45°. Inorder to block specularly reflect-ed light, an incident light angle of0° is applied. Figure 4 schemati-cally shows the alignment of thesample to the integrating spherewhen only the diffusely transmit-ted light is measured.

Measuring results

Under these measuring condi-tions, several textiles were ana-lyzed in order to determine thebest properties of the materialunder the aspects of heat trans-missibility and UV transmissibi-lity.

Intelligent textiles reduce warmiworkspaces Advantages of the triple-detector integratin

Figure 1: A typical window blind – a vertical blind Figure 2: Textile with two surfaces, a silver and a cream colored layer of a roller

blind material

Figure 3: Reflectance of light at a

surface

7

TELEGRAMShimadzu News 1/2010

entire UV-VIS-NIR range can beinvestigated with high measuringsensitivity in one single measure-ment.

Legend

UV = Ultraviolet – too much UV

light can cause skin cancer

and eye damage

ing of ng sphere

Figure 4: Alignment of the experiment

VIS = Visible light – this is the work-

ing range of the human eye

NIR = IR is the thermal radiation;

NIR is the near-infrared spec-

tral range of the radiation.

-5.004

0.000

50.000

98.274

190.00 500.00 1000.00 1500.00 2000.00 2500.00

nm.

R %

orange-red = sample with silver and cream

colored surfaces

purple = sample with green surface

blue = sample with brown surface

green = sample with grey surface

light brown = low-density grey sample

Figure 5: UV-VIS-NIR spectra of various textiles composed of polymer and metal

layering

New integrated FTIR accessoriesIRAffinity-1

FTIR instruments with attenuated total reflectance (ATR) for surface meas-

urements have revolutionized infrared spectroscopy. The ATR measuring

technique requires an easy sample preparation and post-processing. Identi-

fication or quality control can be carried out simply, effectively and quickly.

The ATR technique has been further advanced – through its single reflec-

tance accessories. Now, even less sample quantity and time are needed for

sample preparation and post-processing than ever before. In addition, due to

their small sample plate surface area, these accessories are extremely

robust. Sample plates manufactured out of diamond are almost indestructi-

ble. This leads to higher sample throughput and to any possible contamina-

tion of the sample compartment.

In order to meet the demands of high sample throughput and fast measure-

ment, a new accessory concept has been developed for the IRAffinity-1.

In 2010, Shimadzu will introduce its new ‘integrated accessories’ for the

IRAffinity-1. The accessories originate from the single reflectance and the

HATR-techniques. A single reflectance unit, for instance, enables the pro-

cessing of large samples as the measuring position is situated on top of the

instrument housing. These accessories expand the already wide range of

products. More news will be available at analytica 2010.

15

APPLICATIONShimadzu News 2/2001INTERNAL Shimadzu News 2/2001

14

Highlighted Piece by Piece Determination of hydroxyproline content in meat and meat products according to § 35 LMBG

Shimadzu Biotech has beencreated to bring together astrong solutions-based offer-

ing to accelerate the progress ofbiotechnology research and devel-opment.

Shimadzu Biotech captures, with-in one dedicated organisation, thebest expertise and technologyfrom within Shimadzu Corpora-tion and its subsidiary, KratosAnalytical.

The group’s initial offering will bebased on delivering a wide rangeof key products covering tech-nologies from DNA sequencingto high performance mass spec-trometry to provide an integratedapproach to the fast growing pro-teomics and genomics markets.Proteomics is rapidly becomingthe focal point of biopharmaceu-tical research and is predicted togrow by 20 % to a total of $ 10billion over the next 5 years.

Kratos Analytical’s new MALDI-TOF (Matrix-Assisted Laser Des-orption Ionisation Time-of-Flight) and MALDI IT-TOF(Quadrupole Ion Trap Mass Spec-trometer) represent the cuttingedge of mass spectrometry – akey technology ideally suited tofuture developments in pro-teomics.

Other applications that the newunit will be supporting includecombinatorial chemistry, micro-satellite DNA and SNP analysis.

Shimadzu Biotech will be head-quartered in Kyoto and will oper-ate in the USA, Japan and Centraland Northern Europe under thestrategic direction of an interna-tional management team, headedby Dr Ichikawa who joins Shimadzu Biotech from his previ-ous position as head of life sci-ence at Shimadzu Corporation.

Dr Ichikawa commented, „We seean excellent strategic fit betweenour know-how and the criticalsuccess factors that will be essen-tial to the future of the biotechindustry. Shimadzu Biotech is oursolution to guaranteeing that we

can deliver this expertise to thesector“.

The new business unit will pro-vide a full service offering usingdedicated sales teams, supportedby full applications and serviceback-up.

In addition to bringing togetherproducts from both ShimadzuCorporation and Kratos Analyti-cal to provide scientists with aunified solution to the drug dis-covery process, Shimadzu Biotechwill also be a vehicle for evaluat-ing and developing exciting newtechnologies to advance researchin this field. A key part of thiswill be the creation of strategicpartnerships, such as the Pro-teomics Alliance. This combinesShimadzu Biotech, Proteome Sys-tems and Sigma-Aldrich into asingle force focused on providingintegrated practical solutions forproteomics.

The three companies will worktogether to provide quality coreproducts that meet the demandsof protein scientists – from front-end chemicals to protein identifi-cation and informatics.

Other key relationships that havebeen formed include the multi-million dollar agreement betweenLumiCyte, Inc. of Fremont Cali-fornia and Kratos (part of thenewly formed Shimadzu Biotech)to supply mass spectrometers tosupport LumiCyte’s ground-breaking proteomics work. Underthis arrangement, LumiCyte willpurchase a minimum of 105 AXIMA-CFR MALDI-TOF sys-tems over the next thirty monthsto incorporate into its ProteinBioChip data aggregation facili-ties worldwide.

Customer defined equation Setting of Pass/Fail criteria

Common experience showsthat meat is the most nutritious of all foods

according to Justus von Liebig,one of the best-known chemistsin Germany. As a main food itemon our plates, meats and meatproducts must be constantly sub-jected to stringent quality control.

One of the analytical methodsused is UV-VIS spectrophotome-try. This article describes the determination of hydroxyprolineaccording to the German standard§ 35 LMBG. Hydroxyproline is aparameter for the metabolism ofcollagen. Via the hydroxyprolinecontent, the collagen content canbe calculated, which in turn is anindication of the meat content inmeats and meat products.

For the determination of hydroxy-proline a double-beam UV-1700UV-VIS spectrophotometer withUV-Probe software is used. In addition to the quantification ofhydroxyproline via a calibrationcurve, it is also possible to usevarious arithmetic calculationfunctions and to define pass/failcriteria. A report generator facili-tates printing of the results. Thephotometric determination is car-

ried out via solutions of red 4-dimethyl aminobenzaldehyde ata wavelength of 560 nm. Themethod can be automated using aperistaltic sipper in combinationwith a flow cell. The data is eval-uated via a 6-point calibrationcurve.

During measurement it is possibleto switch between standard andsample measurements. The stan-dards therefore, do not have to bedetermined before the actual sam-ples but can be included in thesample analysis.

With the use of user definablearithmetic calculation functions, it is possible, for instance, torecalculate the hydroxyprolinecontent from units in µg/0.1 mL

to g/100 g. This way the collagencontent can be calculated directlyfrom the hydroxyproline content.

The pass/fail function of the soft-ware allows the definition of vari-ous control criteria. The sampletables show whether a measuredvalue meets these criteria. Thepass/fail function is employedduring method development inorder to test whether the sampleconcentration lies within the cali-bration range. In addition to thehydroxyproline method, the UV-1700 spectrophotometer issuitable for other meat qualitycontrol methods: the determina-tion of nitrite/nitrate in cold cutsor the determination of totalphosphate in meats and meatproducts.

We will gladly send you further information. Please note the appropriate number on your reader reply card. Info 245

µg/0.1 mL

Fail: Concentration above calibration area• Probe 3: Conc > 3.8 µg/0.1 mL (highest Standard)• Probe 4: Conc > 0.3 µg/0.1 mL (lowest Standard)

g/100 g

New Biotech Unit

w w w . s h i m a d z u - b i o t e c h . n e t

Blank page

Extinction using the fibre-optical immersion probe

UV-1700

Extinction using the 10 mm quartz cuvette

260 nm

0.210

0.430

0.856

1.744

2.599

0.205

0.425

0.851

1.739

2.648

350 nm

0.161

0.326

0.644

1.281

1.938

0.157

0.320

0.635

1.289

1.928

15


It is often necessary to moni-tor the quality of products orraw materials located remote-

ly from the UV-VIS spectropho-tometer. Fibre-optics technologymakes this possible. In this arti-cle the performance of a fibre-optical probe is demonstrated bycomparing the transmissionmeasurements of a stronglyabsorbing solution, successfullyachieved with a routine system.

Shimadzu’s UV-1700 is a high-performance, application-orient-ed and compact spectrophotome-ter and in combination with theHELLMA fibre-optical probeand fibre-optic enables fast andprecise measurements of UV-VISspectra. The UV-1700 (190 - 1100 nm) can be operated as astand-alone instrument or via theuser-friendly UVProbe software.Using the standard SUPRASIL®

300 dip probe and the quartz fibre-optic (monofibre) for theUV-VIS, a spectral range of 220 nm up to 1100 nm is attain-able. HELLMA offers a fibre-optic adapter that can be attached

via the standard cuvette holder(Figures 1 and 2) connecting the fibre-optic probe to the UV-1700.

Performance

In order to verify the perform-ance of the system, potassiumdichromate (K2Cr2O7) solutionsof differing concentrations wereexamined. As a reference theequivalent spectra of the 10 mmquartz cell measurements were

used. The absorption path lengthof the dip probe used was also 10 mm. Due to the constructionof the dip probe, where the lightbeam is passed through the sam-ple only once, the measuringprinciple applied is the same as incuvette measurements. A total offive different concentratedK2Cr2O7 solutions in 0.01 n(0.005 M) H2SO4 were used.

The complete UV-VIS spectrumwas acquired for each solution


14

l within reachFar away, but stil

and the absorbances at the char-acteristic wavelengths at � 1 = 260 nm and � 2 = 350 nm weremeasured. Figure 3 shows a com-parison of the spectra obtainedfor the cuvettes and the dipprobe with a 2 m fibre-opticprobe for the dichromate meas-urement.

It is clear that above 270 nmthere is very good correspon-dence between the spectraobtained via both techniques.Only below 270 nm is a decreasein transparency of the fibre-opticobserved for the highest concen-trations, with a correspondingslight decrease in the signal tonoise ratio of the absorption sig-nal. To get a complete overview,the absorbances at 260 nm and350 nm are shown in Table 1.

Application area

The spectra obtained using thestandard dip probe and the 2 mUV-VIS fibre-optic with fibre-optic adapter correspond verywell with the 10 mm quartzcuvette measurements over theentire spectral range of 220 up to1100 nm. Shimadzu’s UV-1700with fibre-optic and dip probe istherefore very well suited forspectroscopic measurements thatneed to be carried out at a dis-tance from the sample.

This includes measurements ofraw materials for manufacturingprocesses at production site aswell as measurements that due totheir hazardous nature cannot becarried out directly in the spec-trophotometer. Furthermore,measurements in hazardous explosive environments are included, measurements of high-

ly toxic and radioactive com-pounds as well as measurementsin highly contaminated areas.

The fibre-optic system presentedhere can also be used in unal-tered form for the UVmini-1240,MultiSpec-1501, UV-1650PC,UV2401PC and UV2501PC.

We will gladly send you further infor-

mation. Please note the appropriate

number on your reader reply card.

Info 292

Figure 1: UV-1700 with UV-VIS fibre-optic and

fibre-optic adapter

Figure 2: Fibre-optic adapter for standard cuvette holder

Table 1: Absorbances at 260 nm and 350 nm of the five dichromate solutions

of different concentrations.

Figure 3: Comparison of UV-VIS

measurements of a K2Cr2O7 solution

in 0.01 n (0.005 M) H2SO4.10 mm

quartz cuvettes and dip probe measu-

rements are shown together

Figure 4: Comparison of the 10 mm cuvette- and the dip probe measurements

at 260 nm (black graph) and at 350 nm (blue graph). Equal wavelengths have the

same colour.

The deviations from the cuvette measurement, even at the highest concentrations

are less than 2 % (260 nm), and less than 1 % (350 nm)

optics – Successful analyses using routine systemsUV-VIS measurements with fibre-

Concentration (mg/L)

K2Cr2O7

15

30

60

120

180

15

30

60

120

180

nm.

220.00 300.00 400.00 500.00 550.000.000

3,000

2,000

1,000

Ext

inct

ion

(K2Cr2O7)/mg/L

0 25 50 75 100 125 150 175 2000.0

0.5

1.0

1.5

2.0

2.5

3.0

Ext

inct

ion

Extinction using the fibre-optical immersion probe

UV-1700

Extinction using the 10 mm quartz cuvette

260 nm

0.210

0.430

0.856

1.744

2.599

0.205

0.425

0.851

1.739

2.648

350 nm

0.161

0.326

0.644

1.281

1.938

0.157

0.320

0.635

1.289

1.928

15


It is often necessary to moni-tor the quality of products orraw materials located remote-

ly from the UV-VIS spectropho-tometer. Fibre-optics technologymakes this possible. In this arti-cle the performance of a fibre-optical probe is demonstrated bycomparing the transmissionmeasurements of a stronglyabsorbing solution, successfullyachieved with a routine system.

Shimadzu’s UV-1700 is a high-performance, application-orient-ed and compact spectrophotome-ter and in combination with theHELLMA fibre-optical probeand fibre-optic enables fast andprecise measurements of UV-VISspectra. The UV-1700 (190 - 1100 nm) can be operated as astand-alone instrument or via theuser-friendly UVProbe software.Using the standard SUPRASIL®

300 dip probe and the quartz fibre-optic (monofibre) for theUV-VIS, a spectral range of 220 nm up to 1100 nm is attain-able. HELLMA offers a fibre-optic adapter that can be attached

via the standard cuvette holder(Figures 1 and 2) connecting the fibre-optic probe to the UV-1700.

Performance

In order to verify the perform-ance of the system, potassiumdichromate (K2Cr2O7) solutionsof differing concentrations wereexamined. As a reference theequivalent spectra of the 10 mmquartz cell measurements were

used. The absorption path lengthof the dip probe used was also 10 mm. Due to the constructionof the dip probe, where the lightbeam is passed through the sam-ple only once, the measuringprinciple applied is the same as incuvette measurements. A total offive different concentratedK2Cr2O7 solutions in 0.01 n(0.005 M) H2SO4 were used.

The complete UV-VIS spectrumwas acquired for each solution


14

l within reachFar away, but stil

and the absorbances at the char-acteristic wavelengths at � 1 = 260 nm and � 2 = 350 nm weremeasured. Figure 3 shows a com-parison of the spectra obtainedfor the cuvettes and the dipprobe with a 2 m fibre-opticprobe for the dichromate meas-urement.

It is clear that above 270 nmthere is very good correspon-dence between the spectraobtained via both techniques.Only below 270 nm is a decreasein transparency of the fibre-opticobserved for the highest concen-trations, with a correspondingslight decrease in the signal tonoise ratio of the absorption sig-nal. To get a complete overview,the absorbances at 260 nm and350 nm are shown in Table 1.

Application area

The spectra obtained using thestandard dip probe and the 2 mUV-VIS fibre-optic with fibre-optic adapter correspond verywell with the 10 mm quartzcuvette measurements over theentire spectral range of 220 up to1100 nm. Shimadzu’s UV-1700with fibre-optic and dip probe istherefore very well suited forspectroscopic measurements thatneed to be carried out at a dis-tance from the sample.

This includes measurements ofraw materials for manufacturingprocesses at production site aswell as measurements that due totheir hazardous nature cannot becarried out directly in the spec-trophotometer. Furthermore,measurements in hazardous explosive environments are included, measurements of high-

ly toxic and radioactive com-pounds as well as measurementsin highly contaminated areas.

The fibre-optic system presentedhere can also be used in unal-tered form for the UVmini-1240,MultiSpec-1501, UV-1650PC,UV2401PC and UV2501PC.




Info 292

Figure 1: UV-1700 with UV-VIS fibre-optic and

fibre-optic adapter

Figure 2: Fibre-optic adapter for standard cuvette holder

Table 1: Absorbances at 260 nm and 350 nm of the five dichromate solutions

of different concentrations.

Figure 3: Comparison of UV-VIS

measurements of a K2Cr2O7 solution

in 0.01 n (0.005 M) H2SO4.10 mm

quartz cuvettes and dip probe measu-

rements are shown together

Figure 4: Comparison of the 10 mm cuvette- and the dip probe measurements

at 260 nm (black graph) and at 350 nm (blue graph). Equal wavelengths have the

same colour.

The deviations from the cuvette measurement, even at the highest concentrations

are less than 2 % (260 nm), and less than 1 % (350 nm)

optics – Successful analyses using routine systemsUV-VIS measurements with fibre-

Concentration (mg/L)

K2Cr2O7

15

30

60

120

180

15

30

60

120

180

nm.

220.00 300.00 400.00 500.00 550.000.000

3,000

2,000

1,000Ext

inct

ion

(K2Cr2O7)/mg/L

0 25 50 75 100 125 150 175 2000.0

0.5

1.0

1.5

2.0

2.5

3.0

Ext

inct

ion

d = x1�2

1�2

–

1

� ��m

2√n2 – sin2 �


20

olProduction quality contr

The UV spectroscopy canbe applied for the non-destructive determination

of layer thicknesses of thin films.This determination is based onthe simple physical phenomenonof interference, resulting fromstanding waves between twophase boundaries. In order tomake this possible in UV spec-troscopy, the materials under in-vestigation must exhibit layerthicknesses of approximately 0.3to 60 µm. The refractive index of

where d is the film thickness, �mis the number of peaks in thefixed wavelength range, n is theindex of refraction, � is the angleof incidence of the light on thesample surface, and �1 and �2 arethe initial and final wavelength ofthe respective range.

The calculation for two samples,a 10 µm polyvinyl chloride filmand a 46 µm polycarbonate film,with different film thickness is

production, where process con-trol of the thicknesses of theapplied SiO2 films is critical.Wide variations in film thicknessresult in high failure rates duringfurther processing.

The transmission mode is appliedfor thin films on a transparentsubstrate, for instance in the man-ufacture of foils. In the transmis-sion mode it is also possible toanalyze films consisting of onlyone material.

Theory

An example is presented toexplain the physical principle ofreflectance spectroscopy. Lightstriking a film at an angle of inci-dence � invokes an interaction/interference of the incident sur-face A with an angle of reflec-tance of the opposite surface B(bottom), as shown in Figure 1,which results in a spectrum withinterference patterns (Figure 2).Counting the number of peaks(or valleys) in this pattern in afixed wavelength range, the filmthickness can be calculatedaccording to the following equa-tion:

the material must be known to beable to carry out this determina-tion.

Reflectance spectroscopyand transmission measure-ment

There are basically two UVmeasuring modes – reflectanceand transmittance spectroscopy –which enable interference meas-urement in application areas suchas thin films deposited ontomaterials or transparent filmsconsisting of a single material.

Thin films can be present on non-transparent carrier materials. Theuniformity of the surface film canbe determined using reflectancespectroscopy. The sample surfacemust be smooth and mirror-like.Rough surfaces are not suitable,as the diffuse reflectance takingplace at the surface prevents theformation of an interference pat-tern.

A typical example of this appli-cation is film thickness determi-nation of wafers during chip

Film thickness determination using UV spectroscopy

Figure 1: Principle of interference

Figure 2: Screenshot of the film thickness software and an example of the selection

criteria for film thickness determination

NEWS 01.2009_01_GB 11.03.2009 12:55 Uhr Seite 21

Now also Vista compatible

Most valued features of the

TOC-Control V software

21

TELEGRAMShimadzu News 1/2009

olntrcopy

Figure 3: Reflectance measurement of a polyvinyl chloride film with a

film thickness of 10 µm

TOC-Control V software 2.1

Figure 4: UV reflectance spectrum of a polycarbonate film with a

film thickness of 46 µm

0.00

5.00

10.00

15.00

600.0 700.0 800.0 900.0

Wavelength (nm)

Ref

lect

ance

(% R

)

5.00

6.00

8.00

10.00

12.00

14.00

15.00

600.0 700.0 800.0 900.0

Wavelength (nm)

Ref

lect

ance

(% R

)

presented below. Figure 3 showsthe interference pattern of the 10µm PVC film in the wavelengthrange of 600 to 900 nm. This iscompared with the higher filmthickness sample of 46 µm. Theresult is an excellent exampledemonstrating that thin films leadto wide amplitudes and thickerfilms to narrower amplitudes.

The calculation can be easily car-ried out using the film thickness

determination tool featured in the software. When all physicalparameters such as refractiveindex, angle of incidence andwavelength range are known, thefilm thicknesses are calculatedautomatically (Figure 2).

Most valued features of the

TOC-Control V software

Now also Vista compatible

An update (version 2.1) for Windows Vista is now available for the TOC-

Control V software successfully introduced last year. Among the features

most valued by our users are:

Adding additional samples during ongoing operation

Any number of samples, calibration curves or control samples may be

added in the edit mode. Previously measured samples can be recalculated

and reports can be printed. During activation of the edit mode the ongoing

measurement cycle continues running.

Other users can take over during ongoing operation

In access-controlled systems, it is important that change of personnel dur-

ing shift operation allows users to log in and out without interrupting the

ongoing measuring cycle. This can be configured in the user authorization

rights for each user.

Simplified recalculation in a single step

In the past, recalculation of previously measured samples using another

calibration curve had to be carried out individually for each sample. In the

TOC-Control V version 2.00 software, however, the respective samples are

tagged in the sample table before a new calibration curve is selected and

the recalculation is carried out.

Simple operation

Sample tables can be established via a simple ”drag & drop“ operation.

Using the mouse, the desired elements are dragged into the sample table

from a list of the established calibration curves, measuring methods, con-

trol samples and measuring sequences. The right mouse button contains

many functions which greatly simplify operation.

The use of shorter ”Wizards“ enables even faster establishment of cali-

bration curves and measuring methods as well as improved templates for

sample tables in routine analyses.

NEWS 01.2009_01_GB 11.03.2009 12:55 Uhr Seite 22

9


Figure 1: Two camera objectives from

different manufacturers, featuring a stan-

dard lens and a macro lens

Figure 2: UV-VIS spectra of a zoom

objective at a setting of 25 mm (green)

and 300 mm (blue)

Figure 3: UV-VIS spectra and compari-

son of the light transmittance of a fixed

objective (black) of 50 mm with a zoom

objective (green) at a setting of 25 mm

Everything within viewQuality control of camera objectives with UV-2600and MPC-2600

Testing two objectives

In the experiment shown here,two objectives from different man-ufacturers were tested. Since theobjectives have been designed fordifferent uses, they have differentcharacteristics:1. Fixed lens system, 50 mm F/1.22. Macro lens, 28-300 mm F3.5 -

6.3 DG Macro (F = focal length,1.2 or 3.5 -6.3 are aperture val-ues.)

The objectives were measuredusing the UV-2600 and MCP-2600sample compartment. The objec-tives were placed on the V-table ofthe MPC-2600 and were broughtto the measuring position. Themeasuring assembly simulates theincident light for imaging withinthe camera. The objectives weremeasured with the incident lightfrom outward to inward.

The detector (photomultiplier) islocated in an integrating sphere(Ulbricht sphere) and displays thetransmittance, i.e. the light of thetransmittance of the object (objec-tive). Furthermore, absorptions bysealings can be expected, as well aseffects of reflections and antire-flective coatings.

Discussion of the spectra

When the focal length is increased,the light transmittance decreasesdue to the reduction of the field ofview, as can be seen in Figure 2.The spectra displayed are the lighttransmittance at 28 mm (70.6 %)and 300 mm (37 %). Due to thereduced light transmittance, takinga photograph using the zoom set-ting needs a longer exposure time,a larger aperture or additionallighting in order to obtain morelight intensity.

The quality of the objectives variesaccording to the function of the

In addition, it is also possible tocheck the specifications that char-acterize the objective. The goal is,after all, to manufacture high-quality objectives. There are sever-al criteria in photography that canbe met by implementing suchquality control:• the visible range of an objective

for the image quality of the colors, or depth of field in thephotographs

• the quality of the coatings onglasses and lenses

• sensitivity in the red or blueranges.

In addition, a UV-VIS spectro-photometer can be used to qualifycamera accessories such as polariz-ing filters or UV filters.

With the introduction of the new generationof LO-RAY-LIGH®

gratings in Shimadzu’s new UV-2600/UV-2700 UV-VIS spec-trophotometer series, many appli-cation areas in the field of opticscan now be covered. In addition to single optical elements such aslenses and glasses, composite ma-terials or single-coated systemsand even entire assemblies cannow be analyzed. Camera objec-tives are small optical benches,equipped with different lenses andglasses with protective coatings orsurface finishes. The quality of anobjective is determined by its lightintensity and low optical aberra-tions.

The light intensity can be deter-mined spectroscopically. By com-bining the UV-2600 or UV-2700with a MPC-2600 extra large sam-ple compartment, non-destructiveanalysis of entire camera objectivesis possible. This sample compart-ment is equipped with a V-shapedholder, ensuring stable positioningof the objective. The holder can bepositioned in all three dimensions,so that the analytical irradiationhits the center of the objective op-tics and passes through the opticalbench, while the spectrophotome-ter measures the incident lightintensity, which is displayed in theform of transmittance spectra.These spectra show not only thelight throughput in percentagesbut also the transmission range asa function of wavelength for visi-ble and ultraviolet light.

Quality determination of objectives and cameraaccessories

Using this combination of instru-ments, it becomes possible to es-tablish a quality determinationprocess that allows productioncontrol of a production series.

50,000

0

-8,873

97,530

nm.

T %

190.0 400.0 600.0 800.0 900.0

50,000

0

-8,873

97,530

nm.

T %

190.0 400.0 600.0 800.0 900.0

objective. An objective with fixedfocal length can result in goodlight transmittance with few com-ponents. Figure 3 shows two verydifferent representative examples.

The light transmittance of thefixed focal length results in a valueof 86.7 % while the variable focallength results in a transmittance ofup to 70.6 %. Assuming the lossof 4 % of the original energy at allsurfaces based on the physics offlat glass, it might be concludedthat the fixed objective consists of4 glass components.

With extrapolation, four glasscomponents should result in a lossof approx. 15 % of transmittance.This corresponds approximately tothe measured value of 86.7 %. Butthis is, of course, a rough estimatefor an unknown object in whichother aspects, e.g. filtering surfacecoatings, can have an influence.

Both objectives are distinguishedby their wavelength range. Theobjective with fixed focal length

features a high light transmittance.In addition, a profile maximum atapprox. 520 nm corresponding tothe green wavelength range is ap-parent. In comparison, the zoomobjective is optimized for wave-lengths in the red range (approxi-mately 620 nm).

Blank page

Cap

Cell

Mirror

Pipette

Window

Samplesolution


4

orAdvances in UV-VIS absShimEvaluation of low DNA/RNA sample volumes on the

spectrophotometer using a Hellma TrayCell

In the last years, new tech-nologies have emerged foraccurate UV-VIS spec-

trophotometric measurements ofsample volumes in the microliterrange. In this respect, the HellmaTrayCell is a more recent opticaldevice that provides, in combina-tion with a standard single beamspectrophotometer, a cost-effec-tive option for assessing sampleconcentration with undiluted,minimal volumes. Hellma Tray-Cell is a highly innovative toolbased on fibre-optics and inte-grated beam deflection technolo-gy (Figure 1).

Nucleic acid measurements

The study focused on nucleicacid measurements at 260 nm,using a plasmid DNA sample. Itsconcentration was arbitrarilyconsidered as ‘reference’ whenmeasured with the classical 10mm-path ultra micro cuvette.Readings were performed at fourdilutions factors (1 : 50, 1 :100,1 : 200 and 1 : 400) and an averagesample concentration of 1.725µg/µL was obtained (CV = 4.3 %)(Table 1). The concentration wasfurther estimated on the HellmaTrayCell with 4 µL of undilutedsample. With the 0.2 mm-pathcap an average absorbance of0.664 was obtained (n = 4, CV =3.7 %), corresponding to an aver-age concentration of 1.661 µg/µL(Table 2). The linearity of theabsorbance readings with theHellma TrayCell/0.2 mm cap set-ting was next assessed, making

Most importantly, its dimensionsare equivalent to a standardcuvette allowing its use in con-ventional spectrophotometers.However, two exchangeable mir-ror caps create defined opticallight path of only 0.2 or 1 mm,which at the sample level trans-lates into virtual dilution factorsof 1 : 50 and 1 : 10 respectively.Higher concentrated sampleswhich cannot be estimated direct-ly with the more classical cuvettes(5 or 10 mm optical path) can bemeasured with the Hellma Tray-Cell without prior dilution.

The sample is placed directly onthe surface of the optical window,and recommended volumes areonly 0.7- 4 µL for the 0.2 mm capor 3- 5 µL for the 1 mm cap.Another valuable feature is thatthe Hellma TrayCell remains inthe photometer during sampleloading, retrieval or cleaning,ensuring not only optical stabilitybut also higher turn around timeof the measurements. Theretrieved sample can be used forfurther analysis if desired.

The aim of this preliminary studywas to compare absorbance read-ings with the Hellma TrayCell onthe Shimadzu UVmini-1240 spec-trophotometer against readingswith a classical 10 mm ultra microcuvette. The latter is of blackbody style with a 10-mm opticalpath, a 2.5 x 2 mm measurementwindow and a minimum requiredsample volume of 70 µL. TheUVmini-1240 spectrophotometeris a single beam optics systemwith concave holographic gratingas monochromator, a 5 nm spec-tral band width and silicon pho-todiode detector.

Figure1: The Shimadzu UVmini-1240 spectrophotometer and the Hellma TrayCell.

The UVmini-1240 is a stand-alone UV-VIS single beam spectrophotometer with an optical

band width of 5 nm. The system uses a concave holographic grating as monochromator and

has a silicon photodiode detector. The measurement wavelength range is 190 to 1100 nm.

On-board software contains basic menus including photometric mode for fixed wavelengths,

spectrum mode for wavelength scanning and quantitation mode for single component analy-

sis. In addition, several optional program packs are available with dedicated research applica-

tions such as multiwavelength measurements, kinetics and protein analysis.

The Hellma TrayCell is a micro cell with fibre optic cables and integrated beam deflection.

Due to these features the sample is placed directly on the surface of the optical window and

readings can be performed either with the 0.2 mm or the 1 mm light path mirror cap,

depending on sample concentration.

Cristiana Stefan, Clin. Chem., Ph.D, Cath. University of Leuven, Faculty of Medicine, Department of Oncology, Belgium; Pascal Mannaert, Shimadzu Benelux, BB´s Hertogenbosch, The Netherlands

*NEWS 01.2007 GB 21.02.2007 15:39 Uhr Seite 5

0.641

0.690

0.646

0.68

Average

SD

CV

9

8

6

4

3

2

1

1

2

3

4

No.

Dilution factor

Dilution factor

0 0.2 0.4 0.6 0.8 1.0 1.20

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

A 2

60 n

m

1/ Dilution factor

y = 0.6951x – 0.0023R2 = 0.9992

Window

Sampleolution

5


orbance measurementsbsShimadzu UVmini-1240the

Figure 2a: Evaluation of absorbance linearity with the UVmini-1240 and the Hellma TrayCell/

0.2 mm cap setting – Absorbance readings and calculated concentrations

Figure 2b: Linearity curve

Table 1: Evaluation of the DNA sample concentration with the UVmini-1240 and the classical

10 mm-path ultra micro cuvette setting

A 260 nm = absorbance at 260 nm · SD = standard deviation · CV = coefficient of variation

µg/µL = [A260 nm (10 mm) x dilution factor x 50 µg/mL] /1000

Table 2: Evaluation of the DNA sample concentration with the UVmini-1240 and the Hellma

TrayCell/0.2 mm cap setting

A 260 nm = absorbance at 260 nm · SD = standard deviation · CV = = coefficient of variation

µg/µL = [A260 nm (0.2 mm) (correction for 10 mm path) x dilution factor x 50 µg/mL] /1000

use of various sample dilutions(n = 7) (Figure 2). In this analysis,the absorbance values covered the0.069 to 0.69 unit interval andaverage sample concentration was1.694 µg/µL (n = 7, CV = 6.4 %).Correlation analysis showedexcellent linearity parameters(r2 = 0.9992, y = 0.6951 x -0.0023),indicating that readings with theundiluted sample were within alinear range. Measurements werealso taken with the 1 mm cap insimilar studies, and an averagesample concentration of 1.614µg/µL (n = 5, CV = 2.7 %) wasfound.

In conclusion, the Hellma Tray-Cell delivers its promises whenused with the Shimadzu UVmini-1240 spectrophotometer: accurateanalysis of small volumes withremarkable reproducibility. Thisstudy was selected as representa-tive and follows a complex sys-tem check procedure.

Accurate measurement of smallvolumes of undiluted nucleic acidsamples is of particular value fortranslational clinical researchfocussing on gene expressionstudies, where small amounts ofpatient specimen, such as biop-

ment rlands

sies, allow only for limited vol-umes of DNA/RNA-containingsamples to be measured. The useof a standard spectrophotometer-Hellma TrayCell setting forabsorbance measurement is alsojustified even when larger vol-umes of DNA/RNA samples areavailable due to elimination ofdilution-related errors and fasterresults through a higher turnaround time.

We will gladly send you further information.

Please note the appropriate number on your

reply card. Info 319

0.069

0.077

0.123

0.173

0.229

0.35

0.69

Average

SD

CV

1.552

1.540

1.845

1.730

1.717

1.750

1.725

1.694

0.109

6.4

A 260 nm (0.2 mm) µg /µL

6.7

8.6

3.4

6.0

0.085

0.180

0.353

0.652

400

200

100

50

Average

SD

CV

0.006

0.016

0.012

0.039

1.700

1.800

1.768

1.631

1.725

0.075

4.3

A 260 nm (10 mm)

(Average of two readings) SD CV (%) µg/µL

1.602

1.725

1.615

1.7

1.661

0.024

3.7

A 260 nm (0.2 mm) µg/µL

*NEWS 01.2007 GB 21.02.2007 15:39 Uhr Seite 6

Shimadzu NEWS, Customer Magazine of Shimadzu Europa GmbH, Duisburg

Publisher:Shimadzu Europa GmbHAlbert-Hahn-Str. 6 -10 · D - 47269 DuisburgPhone: +49 - 203 - 76 87- 0 Telefax: +49 - 203 - 76 66 [email protected]

Editorial Team:Uta Steeger · Phone: +49 - 203 - 76 87- 410 Ralf Weber, Tobias Ohme

Design and Production:m/e brand communication GmbH GWADüsseldorf

Circulation:German: 9,175 · English: 22,920

©Copyright:Shimadzu Europa GmbH, Duisburg, GermanyNovember 2009

Windows is a Trademark of Microsoft Corporation

IMPRINT

The color sells the tea – Color testing using UV-VIS spectroscopy »2

Fast, easy, accurate – Three-step TOC determination in solid materials »4

White Gold – Salt in foods »6

FTIR lemon squeezer extracts citrus fragrance – Single-reflection unit and FTIR spectrophotometer »8

Inside Polymers – GPC for monitoring polymerization kinetics »10

TOC analysis traces efficiency of photocatalysts »12

Drinking water at risk – World Water Forum 2009 »14

Composite materials – how do they react? 17th ICCM in Edinburgh »5

White wine and coffee – Flavour profiling of food samples with SPME-GCMS and On-Column injection »15

Excellence Plus – The new GC-2010 Plus –Even excellence can be better »18

Three turbos for GC – Advanced Flow Technology »20

Recognizing harmful substances in the body – Using graphite furnace AAS in biomonitoring »22

AGS-X – the popular tester – Shimadzu’s new universal testing machine »24

Pilot study – Online TOC for ozonation of treated wastewaters »23

APPLICATION

TELEGRAM

PRODUCTS

CONFERENCE

New TOC draft normDIN EN 15936


2

The color sells t

“Green camomile tea doesn’ttaste.” This might be thefirst reaction of a tea con-noisseur who discoversgreen-colored instead of yel-low tea in his cup.

Although the tea tastes likecamomile, the consumer may notbe able to associate the taste ofhis chosen drink with camomiletea. Not only the taste but alsothe color must be right, since thehuman brain automatically asso-ciates colors with certain fra-grances or aromas. This is whymanufacturers should stick to thecolor of a product when aimingfor a positive taste association.

Perception of colors is a difficulttask for the human eye and, inconsequence, for manufacturers.Their classification dependsstrongly on experience, and it issubjective and hardly applicablein an industrial production pro-cess. This is why a standardizedcolor system was established fornatural minerals. The color tablesenable consistent reproduction ofa required color and support

industrial processes. In naturalproducts, free of artificial color-ing, these color scales are suitablefor the control of technically created color blends, for instance. In the case described below, acamomile tea and its color arecorrelated to the brewing timerecommended by the manufac-turer.

The determination of color is oneof the main applications in UV-VIS instrumental measuring tech-niques. This method, indepen-dently of the human eye, enablesthe use of effective classificationmethods instead of subjectiveassessments.

In this application, a tea bag wasextracted with distilled water atroom temperature. The tea bag

was placed in a 50 mL beakerwithout shaking or stirring.Approximately 3 mL of theextract was transferred to aquartz cuvette suitable for UV-VIS spectroscopy measurementand a UV-VIS spectrum was sub-sequently obtained. Additionalmeasurements were carried outafter 5, 10 and 15 minutes. Thetest was carried out under totaltransmission conditions with thehelp of an integrating sphere of150 mm diameter.

Instrumentation:UV-3600, LISR-3100, transmission integrating sphereattachment, 10 mm quartz cells, 50 mL beaker

Total transmission using anintegrating sphere

In direct transmission measure-ments, the energy of the lightsource is measured before andafter it has been transmittedthrough the sample.

In standard measurement of clearsamples, the detection elementrecords directly transmitted lightleaving the sample. The samplesare transparent. In the case ofnatural products and extractsfrom tea bags, a certain turbidityarising from very small particlesis assumed to occur. An integrat-ing sphere was used to measurescattered light effects as well asdirectly transmitted light. Thissphere captures scattered opticalphenomena and focusses themback towards the detector. Byplacing a measuring cell directlyin front of an integrating sphere,it is possible to measure the totaltransmission, which is the sum of

Figure 1: Dried and crushed camomile from a tea bag

Figure 2: Representation of directly transmitted light

Extraction time [min]

0

5

10

20

3


he tea Color testing using UV-VIS spectroscopy

directly transmitted and scatteredlight.

Visual perception of color ischaracterized via standardizedcolor tables. The yellownessindex and the dominant wave-length in a UV-VIS spectrum canbe determined using the CIEstandard color space (since 1931)and the XYZ tristimulus values,specifically in the visible spectralrange. Table 1 shows some of theproperties and time-dependenttrends. Special attention shouldbe paid to the yellowness indexYI and the dominant wavelengthdWL. Figures 5 and 6 show bothvalues as a function of time. Thecolor values were calculated forthe CIE standard illuminant C,which represents daylight with-out UV radiation under an obser-vation angle of 2°.

Results

The values measured show a defi-nite trend: the tea is ready todrink after approximately 6 min-utes brewing time. This experi-

ment confirms the manufacturer’srecommendation of 6 minutesbrewing time, since in this periodthe dWL as well as the YI (Fig-ures 5 and 6) reach their mathe-matical inflection point in thecurve, after which saturation setsin. This is the point where thegraph reaches a plateau.

The characteristics with respectto the dominant wavelengthchange in time, as the absorptionof the soluble and less solublecomponents in tea are added sub-sequently in time. The observa-tion that the tea extract looksgreen rather than yellow is con-firmed by the analytical wave-length of 570 nm.


mation. Please note the corresponding

number on your reply card. Info 359

Table 1: Calculation result from the color tables, using standard illuminant C and a

standard observation angle of 2°

Figure 3: Placement of the sample in the optical path for measurement of total

transmission in an integrating sphere

Figure 4: UV-VIS spectrum of camomile tea

Figure 5: Representation of the time dependence of the dominant wavelength dWL

for the extraction of camomile tea

Figure 6: Representation of the extraction time dependence of the color yellow for

a camomile tea

-10.048

0.000

50.000

100.000

110.275

300.00 400.00 600.00 800.00 900.00

Wavelength [nm]

R %

Black represents the start time

red = 5 minutes

green = 10 minutes

olive = 20 minutes

In order to avoid subjective analyses, color

scales were applied to indicate the chroma-

ticity of the tea. The represented measuring

range is 300 nm to 900 nm. This corre-

sponds to the visible spectral range.

566

568

570

572

574

576

0 5 10 15 20 25

Time [min]

Dominat wavelength for tea over a time period

0

10

20

30

40

50

60

0 5 10 15 20 25

Time [min]

Development of Yellowness Index for tea

X Y Z dWL Yl

96.71

84.12

72.86

67.48

98.79

87.84

76.87

71.28

115.97

81.75

55.47

44.01

566.8

573.1

574.0

574.6

0.88

23.93

44.82

55.74

Wav

elen

gth

[nm

]Y

I Yel

low

ness

Inde

x

11


• After measurements are com-pleted, the concentrations aredisplayed immediately.

The application example descri-bed above offers an overview ofthe current state-of-the art in thedetermination of hazardous com-pounds in electrical and electron-ic equipment as well as electricalscrap materials according toWEEE and RoHS. The thresholdvalues for lead, mercury, chro-mium, polybrominated biphenylsand polybrominated diphenylethers are determined as 1,000 mg/kg and for cadmium as100 mg/kg, corresponding toconcentrations already used inthe earlier ELV directive.

Hardware and software for accurate determination of hazardous substances

Shimadzu offers an extensiveproduct range featuring a com-plete hardware and software solution for accurate determina-tion of hazardous substances, as well as the competence andknow-how of a market leader inanalytical instrumentation.

Please have a look at the European

seminar tour dates on page 17.




Info 302


10

ccording to the RoHS guidelinesChromium (VI) analysis a

The European Union hasofficially announced in its official bulletin of 13

February 2003, new directivesregulating electrical and electron-ic used equipment (WEEE, WasteElectrical and Electronic Equip-ment) as well as the Restrictionof the use of certain HazardousSubstances in electrical and elec-tronic equipment (RoHS). Thismarks the ratification of bothdirectives in the EU (2002/95/ECand 2002/96/EC) which wereadopted into national law in Jan-uary 2005.

According to the RoHS guide-lines, as of 1 July 2006, a thresh-old value will apply for lead,mercury, cadmium, hexavalentchromium, polybrominatedbiphenyls (PBB) and polybromi-nated diphenyl ethers (PBDE).However, RoHS does not applyto all ten equipment categorieslisted in WEEE. According toArticle 2 of the RoHS directivethe equipment categories 8 (med-ical devices) and 9 (monitoringand control instruments) as wellas certain other equipment classes(computer servers, memory sys-tems and network infrastructurein telecommunications) are cur-rently exempted from the direc-tives under RoHS guidelines.

um (VI) oxidizes 1.5-diphenylcarbazide to 1.5-diphenyl carba-zone, forming a red/violet-col-ored complex with chromium.The extinction of the dye is linearwith respect to the chromium(VI) concentration (Figures 3and 4).

Pentavalent vanadium (V5+),trivalent iron (Fe3+) and tetra-valent molybdenum (Mo4+) alsoreact with 1.5-diphenyl carbazideand, in the presence of the reac-tion solution, result in an appar-ently higher Cr6+ concentration.The concentrations of these ionsmust therefore be determined in a preliminary experiment.

Summary

UV-VIS spectrometry is highlysuitable for fast and straightfor-ward determination of hexavalentchromium and can be applied in routine analysis using theUVmini-1240. The UVmini offersthe following advantages for theanalysis of ions in aqueous solu-tions using the specially devel-oped water analysis programpack:

• Fast and accurate analysis of selected ions in aqueoussolutions.

• 55 measuring programs(optional water analysis pro-gram pack) for 34 different ionsincluding hexavalent chromium.

• All analysis parameters includ-ing the analytical wavelength,calibration curve, measuringtime etc. are adjusted automati-cally by the selected measuringprogram.

and using electrothermal atomisa-tion mode in a concentrationrange of 0.1 up to 20 µg/L.

Photometric quantification ofhexavalent chromium (Cr 6+) can be carried out with theUVmini-1240 (Figure 1) using 1.5-diphenyl carbazide (Figure 2)as reactant to form coloredchromium complexes.

This procedure is suitable for thedetermination of chromium (VI),primarily used as a corrosion-protective layer on metallic sur-faces, but also found in smallparts such as screws, washers andspring rings.

The sample material is eluted intoa reaction vessel during a prede-fined timeframe. A blank value aswell as the extinction at 540 nm issubsequently measured. Chromi-

Where the equipment has beenon the market prior to July 2006,use of these prohibited substan-ces is allowed in replacementparts for repair purposes. Theyare, however, banned in newequipment.

Determination of hexavalentchromium

X-ray fluorescence or atomicabsorption spectrometry onlyenables determination of totalchromium content. X-ray fluo-rescence is a fast technique forthe determination of total chro-mium content and requires virtu-ally no sample pretreatment.After dissolution of the sample,atomic absorption spectrometryin the flame atomisation modecan detect the elements cadmium,lead and chromium in a concen-tration range of 0.1 up to 5 mg/L

Figure 2: 1.5-diphenyl carbazide

Figure 1: UVmini-1240 with cuvette

Figure 3: Chromium (VI) calibration curve

Figure 4: UV-VIS spectra of different concentrations of Cr6+-solutions

hazardous substancesHardware and software for accurate determination of

Cr (V

I) [m

g/L]

00.0

0.2

0.4

0.6

0.8

1.0

1.2

0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9

Calibration Curve

Extinction

Cr (VI) = 1.2302 AU - 0.0021R2 = 0.9993

308.72 400.00 500.00 600.00 676.090.000

0.200

0.400

0.600

0.800

0.881

nm.

Abs

.

Shimadzu News Article Collection

Volume 1

FTIR Fourier Transform Infrared

Spectroscopy

B

3

0.99492

0.00949

0.09743

PLS Calibration ReportPLS I

2

16

4,000.00 - 8,700.00

PLS Calibration

Base line: Auto Zero

Yes

A

3

0.99427

0.01071

0.10347

Algorithm

Number of components

Number of references

Range [cm-1]

Pretreatment

Centered data

Components

Number of factors

Correlation coefficient

MSEP*

SEP*

1/cmFT-NIR IRPrestige-21/Shimadzu

10.000 9.500 9.000 8.500 8.000 7.500 7.000 6.500 6.000 5.500 5.000 4.500 4.000

% T

11


10


When it comes to mod-ern fibres, this sayingseems truly outdated,

even if the requirements withrespect to comfortable wear anddurability of yarns and fibres arecontinually increasing. But thisgoes hand in hand with the con-tinuous developments of newfibre coatings which enable fastand reliable production processesin mechanical looms. By coatinga fibre with polymers, its surfacebecome smoother and its resist-ance to tearing is increased.

In order to guarantee a constantquality during the fibre produc-tion process, a fast and preciseanalytical method is needed and

this is offered by the FT-NIRtechnique. Shimadzu’s IRPres-tige-21, a powerful FTIR spec-trophotometer, covers the entireinfrared wavelength range (MIR[standard], NIR and FIR[optional]).

Using the NIR integrationsphere (Pike Technologies, seeFigure 1) with integrated Indi-um-Gallium-Arsenic detector,which is available as an accesso-ry, the time- and materials-consuming routine analyticalmethod is simplified resulting inincreased productivity. A com-plex sample preparation processis generally not required.

In NIR spectroscopy, parameterssuch as particle size, particle sizedistribution, phase (solid or liq-uid), compression, temperatureand other physical and chemicalcharacteristics have a significantinfluence on the sample spectrumand therefore must be taken intoaccount during evaluation of thespectra and sample preparation.

In all cases, the IRPrestige-21with the NIR integration sphere

Table 1: Summary from the calibration report of the IRsolution software 1.10

*MSEP: Mean Square of Prediction

**SEP: Standard Error of Prediction

Production control of fibres using FT-NIR

1/cmFT-NIR IRPrestige-21/Shimadzu

10.000 9.500 9.000 8.500 8.000 7.500 7.000 6.500 6.000 5.500 5.000 4.500 4.00030

37.5

45

52.5

60

67.5

75

82.5

90

97.5

% T

Figure 1: IRPrestige-21 with NIR integration sphere and sample holder

Figure 2: NIR spectrum of a coated fibre sample, measured in diffuse reflection

mode. 40 spectra were averaged, 8.0 cm-1 resolution

Figure 3: NIR spectra of 16 differently treated fibre samples, each resulting

from averaging 40 spectra, 8.0 cm-1 resolution

Figure 4: Screen shot of the option Calc vs. Input in the IRsolution software,

the red cross indicates outliers. A data name is assigned to each data point

Double stitched provides more strength

measures the back-reflected light.Transparent materials or filmsonly generate very weak absorp-tion signals that can be stronglyenhanced via a reflecting mirrororiented towards the sample sur-face as the NIR light is directedback into the integration sphere.This double transmission (via thesample surface, reflected by themirror back to the sample sur-face and subsequently into theintegration sphere) is an elegantway of analysing the layer thick-ness of transparent samples. Thisstate-of-the-art technique is alsoreferred to as transflectance.

The NIR spectra obtained gener-ally show broad bands that canseldom be individually assigned.The sample is therefore bettercharacterised by the totality ofthe spectrum. Using calibrationmodels, the required selectivitycan be attained. Figure 2 shows a spectrum of a coated fibre sam-ple obtained using the IRPres-tige-21 with integration sphere.The fibre sample was positionedin the desired orientation on thesample window of the integra-tion sphere and placed inside the

sample holder (the fibre is posi-tioned on a card).

The overlapping NIR absorptionbands in the spectrum require amultivariate mathematical proce-dure such as partial least squares(PLS) in order to take full advan-tage of the benefits of the highsignal to noise ratios, irrespectiveof the overlapping bands. Thisprocedure does not require theselection of suitable wavelengthsas this concerns a full-spectrumanalysis. To increase the robust-ness of the calibration it mayonly be necessary to exclude sev-eral non-variable ranges in thespectrum.

The PLS method is especiallysuitable in cases where a largenumber of standard spectra (ref-erence spectra) are available, butvery little information on theNIR absorption behaviour of the compounds.

In order to create a suitable cali-bration model, standards arerequired that reflect the expectedvariance of the analytical sam-ples.

Chemometrics of a fibresample series

The NIR spectra obtained werecalibrated on two productionparameters A and B. Parameter Adescribes the production rate and

B the percentage uptake of thepolymer. Both parameters pro-vide an indirect indication of thecoating process of the fibres withthe polymers or polymer blends.For calibration, a wavenumberrange between 8700 and 4000 cm-1

was selected, as this range corre-lates strongly with the two pre-determined parameters A and B.

To improve the PLS calibration,all spectra were centered andbaseline corrected using theIRsolution software. Figure 4shows the option Calc vs. Inputthat provides a fast overview onthe quality of the calibration.The IRsolution software pro-vides the reference parametersversus the spectroscopicallyobtained parameters. Outliers areindicated with a cross.

In addition to the various graphi-cal representations, as shown inFigure 4, the software also dis-plays analysis results in tabularform. A summary is shown inTable 1. For the 16 fibre samplesanalysed, a correlation coefficientof 0.99427 for A and 0.99492 forB was obtained, which reflectsthe high precision of NIR meas-urement.

Advantages

In comparison with routine ana-lytical methods, FT-NIR spec-troscopy delivers accurate and

reproducible results with lesseffort. Using the IRPrestige-21with the NIR integration sphereand integrated InGaAs detector,meaningful NIR spectra can beobtained easily and quickly with-out the need for sample pretreat-ment.

In addition to the quantitativefibre measurements presentedhere, it is also possible to measuremany types of compounds direct-ly in their glass containers, pro-vided that the container walls arenot too thick. For quantitative

evaluations, however, a very thintype of glass with a reproduciblelayer thickness can be used.

Non-homogeneous samples (forinstance foods such as flour) canbe spectroscopically analysedand quantitatively evaluated withvery high precision using theoptionally available rotating sam-ple- and container holder.




Info 291

19


Tiger bones, insects and herbs

FTIR spectrometry in traditional Chinese medicine

The roots of traditionalChinese medicine (TCM)date back to more than

two thousand years. With its ori-gin in Eastern Asia and separateroots in Korea and Japan, TCMis today widely practised inEurope – and also within theGerman health care system. Froma health policy point of view,however, TCM is still acceptedonly to a limited extent.

TCM is represented by numerousmedical societies, such as theDeutsche Ärztegesellschaft fürAkupunktur (DÄGfA), one ofthe largest natural medicine pro-fessional societies in Germany. In addition, there are a number of scientific organizations forquality assurance and implemen-tation of medical studies in thearea of TCM. This includes thenon-profit TCM-initiative(www.tcm-initiative.de) offeringthe highest possible transparencywith respect to TCM herbal qual-ity available in Germany. Thisassociation also organizes collab-oration among research institutes,clinics and conventional physi-cians as well as TCM specialists,and ensures the future implemen-tation of TCM studies. TheTCM-initiative also providesinformation on possible healthinsurance reimbursements of var-ious forms of TCM therapy.

TCM usually combines variousmethods. The five most impor-tant ones are acupuncture, mas-sage, dietary therapy, exercise andalternative medical therapy.

Alternative medical therapy,the most significant methodof treatment

Alternative therapy is based onthe administration of formula-tions derived from natural prod-ucts and offers the most signifi-cant therapeutic range of all five

methods. The chemical composi-tion of TCMs is of special impor-tance with respect to the Arznei-mittelgesetz (German Drug Reg-istration and Administration Act)

The professional literature, forinstance the journal “ClinicalChinese Pharmacology” [1],names 515 individual formula-tions. Approximately five ofthese are derived from vertebratepreparations or parts such as tiger

bones, as well as from fossilbones of pre-glacial animals. Fiveother formulations are of mineralorigin or consist of excrements,secretions, worms, insects andmollusc parts and 85 originatefrom plants.

In Europe, alternative medicaltherapy is usually limited to phy-totherapy, i.e. the use of activecompounds originating fromplants under controlled cultiva-tion. In Europe, the only animalingredients officially used inalternative medicine are seashells(for instance the Chinese oysteror the arca shell).

FTIR spectrometers forquality control of raw materials, finished productsand packaging

FTIR spectrometers such as Shi-madzu’s IRAffinity-1 (Figure 1)

are routinely used in quality con-trol for the unequivocal identifi-cation of compounds. This relatesto raw materials as well as to finished products and packagingmaterials.

Individual compounds exhibit acharacteristic infrared spectrumso FTIR technologies thereforeoffer fast and straightforward,unequivocal analytical results ofthe most diverse samples.

IRAffinity -1 with microscope AIM-8800

FTIR analysis is carried out usingthe IRAffinity-1 in combinationwith a single reflection ATR(attenuated total reflection) unitwith a diamond or KRS-5 crystal.This measurement setup providesunequivocal infrared spectra oftypical packaging materials suchas polypropylene and polyure-thane.

In this way, it is also possible toanalyze natural compounds suchas ginseng, whose compositioncan vary widely depending on theharvest and soil conditions. FTIRalso provides unequivocal spectrafor these complex samples andenables reliable quality control intraditional Chinese medicine.

Currently, more than 1100 infra-red spectra are registered in theChinese Pharmacopoeia, signi-ficantly more than in the Britishor Japanese Pharmacopoeias, as

all raw materials, mixtures andpharmaceutical products such asTCMs are controlled using infra-red spectrometry.

The IRAffinity-1 enables fast andreliable routine analysis andensures a uniform quality of theactive drug compounds in orderto protect consumers.

[1] Manfred Porkert:

Klinische Chinesische Pharmakologie.

Fischer, Heidelberg 1978

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PolymerPolyimide

Polymethyl methacrylate

PMMA containing PBDE

Polyoxymethylene (acetal)

POM containing PBDE

Polypropylene

PP containing PBDE

Poly(phenylen sulfide)

PPS containing PBDE

Polystyrene

PS containing PBDE

PSPS containing TBBPA

PS (containing brominated aromatic triazine)

Polyurethane

Poly(vinyl alcohol)

Poly(vinyl chloride)

Poly(vinylidene fluoride)

PVDF containing PBDE

Styrene-butadiene-styrene copolymer

SBS containing PBDE

Acrylonitrile-butadiene-styrene

ABS containing PBDE

Ethylene-vinylacetate copolymer

Nylon

Nylon containing PBDE

Poly(butylene terephthalate)

PBT containing PBDE

Polycarbonate

PC containing PBDF

PC containing TBBPA (Tetrabromobisphenol A)

Polychlorotrifluoroethylene

PCTFE containing PBDE

Polyethylene

Polyethylene containing PBDE

Polyether-ether-ketone

PEEK containing PBDE

Polyetherimide

Polyester

PES containing PBDE

Poly(ethylene terephthalate)

PET containing PBDE

Polymer

Tetrabromobisphenol A

Decabromodiphenyl Ether

Octabromodiphenyl Ether

Brominated Aromatic Triazine

Brominated Polystyrene

Tetrabromobisphenol A

Decabromodiphenyl Ether

Octabromodiphenyl Ether

Brominated Aromatic Triazine

Brominated Polystyrene

ABS

ABS+PBDE

EVA

NYLON

NYLON+PBDE

PBT

PBT+PBDE

PC

PC+PBDE

PC+TBBPA

PCTFE

PCTFE+PBDE

PE

PE+PBDE

PEEK

PEEK+PBDE

PEI

PES

PES+PBDE

PET

PET+PBDE

Abbreviation

Brominated Flame Retardants WoodyOthers

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Thermoplastic Polymers

4000 3500 3000 2500 2000 1750 1500 1250 1000 750

0.0

0.2

0.4

0.6

0.8

1.0

AB

S

1/cm

Polystyrene with TBBPA(Tetrabromobisphenol A)

Polystyrene with brominated aromatic Triazine

Polystyrene with DBDPE

pure Polystyrene

AB

S

brominated biphenyls exhibit theirown very characteristic infraredspectra. The polystyrene exampleexhibits three spectra: DBDPE, PS with DBDPE and pure PS. Therange in the IR fingerprint, whereDBDPE in PS identification is pos-sible, is clearly discernible (Figure2). Figure 3 shows polystyrenewith other brominated flame-retardant additives.

Fast identification of brominated flame-retardants

The fingerprint region of 1500 cm-1

- 1000 cm-1 is important for the

state. The diamond sample surfaceenables the application of highpressures to ensure that the sampleis positioned tightly on the crystalso that optimum penetration of thesample by the IR beam is guaran-teed. The beam penetrates the sam-ple surface to a depth of approxi-mately 2 µm.

As RoHS specifies a homogeneoussample material, this depth of penetration is sufficient in order tocompletely characterise the sample.Using this measuring configura-tion, the spectrum is acquiredwithin a very short time interval


10

(approximately 1 min.) and is eval-uated automatically according toRoHS guidelines.

To validate analysis results, thespectrum is compared subsequentlywith a library of polymer spectra.For polymer identification, adatabase already containing 41polymers (see table 2) is used. Thisdatabase contains logical associa-tions and the Distinction Softwaretests for plausibility, for instanceby evaluation of signal ratios.

The decision criterion includeswarning messages that range from“Identification of the polymer notpossible” to “Applied pressure notsufficient” and finally to the con-clusion “O.K.” or “Not O.K.”.These FTIR analysis results can beconsidered as unambiguous whencombined with the pre-analysisfrom the EDX System.

Infrared spectrometry can there-fore be regarded as a fast and sim-ple alternative solution to the pre-selection of polymers. Minimalsample pretreatment is necessaryand fast results are obtained viapredefined methods.

* Directive 2002/95/EC of the European

Parliament and of the council of 27 January

2003 on the restriction of the use of certain

hazardous substances in electrical and

electronic equipment

** Flame retardants chemicals association

of Japan (FRCJ)

Figure 3: IR spectra of polystyrene with several flame-retardants

400 350 300 250 200 175 150 125 100 75 501/cm

Figure 2: Polystyrene spectra with and without flame-retardant as well as the

IR spectrum of the flame-retardant decabrominated diphenyl ether.

DBDPE (Decabromodiphenyl ether)

DBDPE

PS with 5 % DBDPE

pure PS

DBDPE Bands

DBDPE measured with the single-reflection ATR method (diamond/KRS-5)

identification of brominated flame-retardants, where clear differencesbetween the spectra can be seen.Based on this information, an ana-lytical method for fast identifica-tion of brominated flame-retar-dants and polymers has been developed using Shimadzu’s FTIR-8400S in combination with a single-reflection accessory.

In the present example, a diamondATR unit with KRS-5 crystal wasused as single-reflection accessory.A diamond as sample surface isrecommended as the polymer canbe present in a flexible or solid

Table 2: Overview of 41 polymers and polymer mixtures in an expandable database

PI

PMMA

PMMA+PBDE

POM

POM+PBDE

PP

PP+PBDE

PPS

PPS+PBDE

PS

PS+PBDE

PS+TBBPA

PS+Triazine

PU

PVA

PVC

PVDF

PVDF+PBDE

SBS

SBS+PBDE

Abbreviation

11


Figure 1: Structural formulae of brominated biphenyls

The RoHS* directive (Restriction of the use ofcertain Hazardous Sub-

stances in electrical and electronicequipment) regulates the restric-tion of the use of brominatedflame-retardants in electrical andelectronic devices as of July 2006.Consequently, polybrominatedbiphenyls (PBB) and polybromi-nated diphenylethers (PBDE) canno longer be used as flame-retar-dants in polymers unless their con-centrations are lower than 1,000ppm. The objective of the RoHSdirective is the protection of hu-man health and the environmentfrom hazardous effects. During recycling of electronics waste, contamination by brominatedcompounds should therefore be reduced.

Polybrominated biphenyls areclassified as health hazards. PBBand PBDE are chemicals that have,in the past, been used as flame-retardants in polymers in concen-

trations of 5 % up to 10 %. RoHSrestricts the use of compoundssuch as tetrabrominated biphenylA (TBBA), brominated poly-styrene and brominated aromatictriazine.

Figure 1 presents the structuralformulae of brominated biphenyls.

According to RoHS, the followingcompounds are considered haz-ardous: pentabrominated diphenylether (PentaBDE) and octabromi-nated diphenyl ether (OctaBDE).OctaBDE has been used in poly-mers such as ABS and PS. Current-ly, decaBDE is largely being usedas a flame retardant in PS, PE, ABSand polyester. DecaBDE has notyet been included in the RoHS di-rective. Commercial decaBDEhowever consists of a mixture ofapproximately 97 % - 98 % decaBDE and 0.3 % up to 3 % ofother BDE’s. Therefore, when apolymer contains 10 % decaBDE(containing 1 % contamination of

other brominated BDE’s), thePBDE content will exceed theRoHS threshold value of 1,000 ppm.

FTIR spectroscopy – fast, non-destructive, simple

In order to comply with the requi-rements of the RoHS directive,first the total bromine content of a sample is determined. If this exceeds 5 % after the preliminaryexamination using the EDX sys-tems, infrared spectroscopy is recommended as this will enableidentification of compounds. This simple and non-destructivemethod quickly leads to useful results. Compound identificationis possible as the flame-retardants,were present up to now in poly-mers in concentrations of higherthan 5 %. This level is still detect-able in polymer mixtures usingFTIR. Concentrations that appro-ach the detection limit, however,must be measured using other ana-lytical methods. In this case,

GCMS is highly suitable as allbrominated compounds can beseparated and detected down to thetrace level. GCMS, on the otherhand, is more time consuming withrespect to sample preparation anddata analysis.

In general, it is recommended tocarry out an overall pre-screeningvia energy-dispersive X-ray fluorescence (EDX). Using thisanalytical method the total concen-tration of elemental bromine in thesample is detected, although it isnot possible to distinguish whichcompound actually containsbromine. When more than 5 % oftotal bromine is detected, FTIRcan be used for further identifica-tion of bromine compounds. Whenless than 5 % bromine is detected,GCMS analysis can be implement-ed for separation and identifica-tion.

Fast and straightforward IR-analy-sis of polymers is possible since

FTIR-Spectroscopy – Method for compliance with the RoHS directive

retardants in polymersIdentification of brominated flame-

Brominated Flame Retardants ABS PS PP PE PC PC/ABS Poly amide Polyester PVC S-PS S-PU

Table 1**: Typical polymers and their flame-retardants. Listing of brominated flame-retardants and their use in polymers. (ABS = acrylonitrile-butadiene-styrene,

PS = polystyrene, PP = polypropylene, PE = polyethylene, PC = polycarbonate, PVC = polyvinylchloride, PU = polyurethane)

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Thermoplastic PolymersEpoxy Unsaturated Polyester Phenol Elastomer Adhesive · Paint Fiber

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6

Plastics analysis in pracFTIR measuring technology – untreated and recycled

Raw materials are becom-ing increasingly scarceand more expensive. In its

IP/11/122 press release entitled“The Commission calls for actionon commodities and raw materi-als”, the EU Commission notonly discussed the supply of rawmaterials but also specificallymentioned resource efficiency inorder to promote recycling.

Recycling of polymers is animportant environmental issue.But this is a difficult road to trav-el, as new legislation specifiesclear threshold values (RoHS*),thereby limiting recyclingoptions. Regranulates designatedfor subsequent processing mustbe tested according to these regu-lations.

Fast analysis of recycled poly-mers is an advantage for incom-ing goods inspection. In theshortest possible time, the basicpolymer as well as problems canbe identified. The increasing

interest in recycling of materialsis of course related to costs. Nat-ural resources can be saved whenrecycling-based materials areused instead.

In comparison: untreated and recycled polymers

What happens during the recy-cling process? Polymers such asABS are mixed from varioussources and returned to the mar-ket as pure-grade material. Thesematerials are, for instance, avail-able as granules and are termed asregranulates. Quality assuranceshould highlight the differencesbetween such mixtures. The pres-ent application compares anuntreated polymer with a treatedone, using an example of pureand recycled ABS.

ABS, a terpolymer, is often usedin the automotive industry as amaterial for casings for lightsources, cooler grilles or hubcaps.The advantage of this material isthat it can easily be galvanized.Since years, mirrors and reflec-tors have no longer been manu-factured from metal.

The terpolymer ABS consists ofacryl nitrile, 1,3-butadiene and

Figure 1: Infrared spectrum of pure ABS, measured using a diamond-based single-reflectance unit

IRAffinity -1

-0.005

4600

1/cmABS, DuraSamplIR

Abs

.

4400 4200 4000 3800 3600 3400 3200 3000 2800 2600 2400 2200 2000 1900 1800 1700 1600 1500 1400 1300 1200 1100 1000 900 800 700 600 500

00.005

0.010.015

0.020.025

0.030.035

0.040.045

0.050.055

0.060.065

0.070.075

0.080.085

0.090.095

0.10.105

0.110.115

0.120.125

0.130.135

0.140.145

0.150.155

0.160.165

0.170.175

0.180.185

0.190.195

0.20.205

7


ticepolymers compared

Figure 2: Infrared spectra of two ABS polymers. The black spectrum represents untreated ABS; the red spectrum recycled ABS

-0.015

4600

1/cmABS, DuraSamplIR

Abs

.

4400 4200 4000 3800 3600 3400 3200 3000 2800 2600 2400 2200 2000 1900 1800 1700 1600 1500 1400 1300 1200 1100 1000 900 800 700 600 500

0

0.015

0.03

0.045

0.06

0.075

0.09

0.105

0.12

0.135

0.15

0.165

0.18

0.195

0.21

0.225

0.24

0.255

0.27

0.285

0.3

0.315

0.33

0.345

0.36

0.375

0.39

0.405

0.42

0.435

0.45

0.465

0.48

0.495

0.51

0.525

0.54

0.555

0.57

0.585styrene. FTIR spectrometry com-bined with single-reflectancemeasurements offers a fast, reli-able and non-destructive analysisof this material. In the presentexample, a diamond-based ATRaccessory was used for single-reflectance. The polymer sample,in the form of a granulate, waspressed with reproducible pres-sure onto the measuring window.Single-reflectance from the IR-beam was already sufficient toobtain a result.

A look at the spectra

To interpret the ABS spectrum,the spectra of three individualpolymers can be used to revealdifferences. The acryl nitrilespectrum, for instance, exhibits acharacteristic band at 2237 cm-1

(nitrile band).

The ABS spectrum exhibits alltypical characteristics of styrene,acryl nitrile and butadiene. At975 cm-1, the spectrum features apeak which can be attributed tothe butadiene group. The basicstructure of the spectrum can betraced to styrene and the nitrilebands are exhibited at 2237 cm-1.

The ABS-typical spectrum isshown in figure 1.

In the spectrum of recycled ABSthe following structures could beidentified: polybromated di-phenyl ether at 1725 cm-1, bromi-nated flame retardant, polycar-bonate with its typical triplebands at 1200 cm-1, 1,3-butadieneat 975 cm-1 and the nitrile struc-ture of acryl nitrile at 2237 cm-1.

The spectrum of the recycledpolymer exhibits the pattern of amixed spectrum: it is not pure.However, this is not necessarily adisadvantage. In the present

example, the polymer was anadmixture with polycarbonate(PC), used to harden polymers.Each of the functional groupsinfluences the physics of thepolymer and this is specificallyused in the manufacturing ofcomponent parts. Additional dis-tinctive features in the spectrumstill need to be clarified. Thisrequires further analysis stepsusing pyrolysis-GCMS and ele-ment analysis using EDX, AASor ICP as already reported in theShimadzu News and variousapplication notes.

*RoHS: Recycling of HazardousSubstances

Literature:

1. “The Commission calls for

action on commodities and

raw materials”, IP/11/122,

Brussels, 2 February 2011

2. “Directive on the restriction

of the use of certain hazardous

substances in electrical and

electronic equipment”, RoHS

2008/385/EC

3. Shimadzu News 2005 and

2006, www.shimadzu.eu,

Application Information

Albert van Oyen of CARAT GmbH in

Bocholt, Germany is gratefully

acknowledged for his kind support

of this topic.

Instruments used:

• Instrument: Shimadzu IRAffinity-1

FTIR spectrophotometer

• Single-reflectance unit:

DuraSamplIR

• Library: Shimadzu RoHS,

Polymer

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Resid. 3

0.0698625

0.0553206

0.418387

0.206929

Sample name

test_sample_1_1

test_sample_1_1_1

test_sample_3_1

test_sample_7_2

Resid. 2

0.0698724

0.0553382

0.418396

0.206971

Results optained with the PLS-I methodParacetamol

36.7629

36.8132

44.5692

25.0618

Paracetamol

35.93

35.93

43.10

31.10

Used composition in %

A total of 15 powder samples ofdifferent concentrations wereused for the calibration in con-centration ranges of: aspirin(acetyl salicylic acid) 40 % - 60 %, caffeine (1,3,7-trimethyl-xanthine) 4.95 % - 25.95 % andparacetamol (4-acetylamidophe-nol) 18.90 % - 55 %. In order toobtain optimal conditions (equalparticle sizes) for diffuse reflec-tion, all samples were ground to a fine powder and mixed. Theresulting powder was placed insmall aluminium holders (Al pel-let holder, diameter 6 mm) andlightly compacted with a pestle inorder to obtain a smooth surface.Figure 2 shows a typical NIRspectrum that was obtained usingthis method.

PLS-I calibration

In order to improve the PLS-Icalibration, all spectra were firstcentred using the IRsolution soft-ware. For this purpose, the aver-age value spectra, obtained from

all calibration spectra, were sub-tracted from individual spectra. Itis commonly recommended indaily laboratory practice, to carryout a baseline correction in orderto correct for possible drift of thebaseline. Table 1 shows an excerptof the PLS calibration report.

As can be concluded from Table 1,all three compounds (aspirin, caf-feine, paracetamol) can be deter-mined with a correlation coeffi-cient of r > 0.998 and a standarderror of prediction of SEP < 0.06.Here the excellent suitability ofthe PLS method for multi-com-ponent systems is evident.

Based on the diagnosis results ofthe IRsolution software, it is pos-sible to make more detaileddeductions from the NIR spec-tra. For example, under the titleVariance, Correlation vs. Sqr.Correlation, information on thevarious spectral ranges and theirsignificance for the correlationcan be obtained. Figure 3 shows

15

PRODUCTSShimadzu News 2/2004PRODUCTS Shimadzu News 2/2004

14

the obtained square correlationspectrum for aspirin.

The square correlation graphshows which ranges of the NIRspectra correlate with the concen-tration of the three compounds.The correlation values lie between+1 and –1. As can be easily seen,the wavelength range between7000 and 3800 cm-1 is especiallysuitable for PLS factor analysisdue to its strong concentrationcorrelation. In this range thegreatest variations in the spec-trum arise from the dependenceon concentration.

Using the option Calc vs. Input(Figure 4), a quick overview onthe quality of the calibration canbe obtained. The outliers areautomatically marked with a redcross (this example does notshow any outliers). In the graph,the measured concentrations areplotted against (depending on theselected unit) the calculated (pre-dicted) values.

For a better overview, the IRso-lution software automaticallywrites the data file name next toeach measuring point. In addi-tion, the software addresses thefollowing diagnosis points:

• Influence: Influence of theindividual standard spectra onthe calibration

• Spectral Residual and Predict-ed Residual: Shows the devia-tions from the predicted con-centrations, for instance expect-ed versus the measured spec-trum

• Scores: Score-score plots foreach component. This enablesthe recognition of informationon the compound as well as thepersonal style of preparation

• P Loadings and weights:Generates the loading, for in-stance ‘Weight Plot’ for indivi-dual factor (four factors in thisexample, Table 1). Based on theindividual plots, the significanceof the calibration can be estima-ted for each factor. The strongerthe noise and the smaller thevariance in the spectrum, thelower is its significance for thequality of the calibration. Ideal-ly, the number of factors shouldcorrespond with the number ofchemical components (aspirin,caffeine, paracetamol)

• PRESS values: This functionshows a graphical representa-tion of the ‘Prediction ResidualError Sum of Squares’ for eachfactor

• PLS Reconstruct: This func-tion is used to analyse the errorbetween the actual and the sim-ulated spectrum and to be ableto adjust the PLS factors

• PLS-Analysis: Using PLS anal-ysis, the determination of con-centrations of unknown sam-ples can be easily carried out.

In this example, a total of foursamples of known compositionwas used as testing criterion forthe robustness of the calibration.Tables 2 and 3 summarise theconcentrations used (composition

Figure 3: ‘Square Correlation’ spectrum for aspirin

Table 3: Results optained with the PLS-I methodeTable 2: Used composition in %

Figure 4: graphical representation of the predicted (predicted) versus

the measured concentrations (actual) for aspirin

New autosampler for UV-VIS and RF

Shimadzu offers, together with CETAC Technologies, two newautosamplers for UV-VIS- and RF spectrometers.

The ASX-520 and the ASX-260 are two autosamplers thatdistinguish themselves by high sample throughput and highprecision. Despite its small size, the ASX-520 can handle upto 360 samples (180 samples in the ASX-260) and guaran-tees fast and simple analyses of large sample numbers evenwhen bench space is limited. Due to its easy installation, theinstrument can be build up and running fully functional withinthe day of delivery.

Five different sample racks can be selected, holding 90 samples (7 mL), 60 samples (14 mL, standard), 40 samples(20 mL), 24 samples (30 mL) or 21 samples (50 mL).

The autosamplers are available for the following spectrome-ters: UV-2401PC / 2501PC, UV-1700 Pharmaspec, UV-1650PC, UVmini-1240, RF-1501 and RF-5301PC.

Fully automatic and precise

TELEGRAMin %) and the results that wereobtained via the PLS-I method.

The test samples 1_1 and 1_1_1are samples from the same pow-der mixture that have been placedonto two different sample hold-ers. They deviate by less than 2 % from the used concentra-tions. The deviations between thesamples 1_1 and 1_1_1 are alsovery small (> 0.1 %), due to theexcellent reproducibility of Shi-madzu’s IRPrestige-21 system.Test sample 3_1 shows a devia-tion of 1.8 % for caffeine up toapproximately 3.3 % for parac-etamol, which is very close to theexpected values. Of interest istest sample 7_2, which showshigher than average deviations of6 % for caffeine and 24.1 % forparacetamol. For this test sample,aspirin and caffeine from a differ-ent batch than the standard wasused. In spite of the same quality,very clear differences in the PLS-I analysis can be seen.

Conclusion

These few examples show veryclearly the possibilities, as well as the limitations of the PLSmethod. For a robust (excellentprediction of the concentration)calibration, it is absolutely neces-sary to take into account the pos-sible variations in the sourceproducts. Particularly in the anal-ysis of products of biological origin, a seasonal update of thecalibration procedure is recom-mended in order to maintainrobustness.

Once this hurdle in the calibra-tion is overcome, NIR-FTIRspectroscopy in combinationwith the DRS-8010ASC diffusereflection unit offers the possibil-

ity for fast and cost effectivequalitative and quantitative anal-ysis of NIR-active compounds.Especially for powder samples,diffuse reflection enables fastanalysis without the need for fur-ther sample preparation. NIR-FTIR spectroscopy also offersthe possibility of non-destructiveanalysis of NIR transparentpackaged products right throughthe packaging material for onlinequality control.

Sample name

test_sample_1_1

test_sample_1_1_1

test_sample_3_1

test_sample_7_2

Aspirin

49.2041

49.1671

45.3284

55.237

Resid. 1

0.0698462

0.0553044

0.418377

0.206874

Caffeine

14.1642

14.1315

10.2883

20.1144

Aspirin

50.11

50.11

46.43

51.90

Caffeine

14.96

14.96

10.47

17.00

Must content (% Bx)

1

2

3

4

5

Standard


12

Truth lies in the mustscopyMust content as wine quality indicator – FTIR-Spectro

Wine is one of the oldestcultural products inhuman history. Vines

have been cultivated for over8000 years. The oldest knownarchaeological evidence of wine-making is an 8000-year old wine- and fruit press found nearDamascus. Awareness of themedicinal effects of wine alsodate back to this time. Hippo-crates (460 – 377 B.C.) recom-mended wine diluted with wateras a remedy against headachesand digestive disorders.

Winemaking is a rather simpleprocess: freshly harvested grapesare crushed and the resultingjuice (must) is collected. Themust contains fermentable sugarsand natural yeasts which, eitherby themselves or with the help ofadditional yeast cultures, start thefermentation process in whichmainly ethyl alcohol and carbondioxide are formed. The latter is agas and escapes. The fermentationprocess comes to a halt when allof the sugars are fermented or thealcohol concentration becomestoo high and kills off the yeasts.

content based on acquisition ofinfrared spectra.

FTIR – fast analysis of IR-active compounds

FTIR spectroscopy is a fast ana-lytical technique for the identifi-cation and quantification of IR-active compounds, for instancethose present in must and wine.Both samples consist mainly ofwater, sugar, acids and otheringredients whereby the crucialdistinction is the alcohol presentin the wine.

In IR spectroscopy it is wellknown that water present in thesamples strongly influences theinfrared spectra. Nevertheless, ameasurable region was found inthe fingerprint where the IR-bands of the must ingredients andof the wine are clearly visible(Figure 1).

As mentioned before, both themust and the wine differ withrespect to their sugar (must) andalcohol (wine) content as indi-cated in the spectral range of1200 up to 1000 cm-1. This is alsothe case in other wavelengthranges. As both substances can beidentified in the water back-ground, these wavelength rangescan also be used for quantifi-cation.

Measurement

Measurement was carried outusing a Shimadzu IRPrestige-21FTIR spectrophotometer in the

At this point the must has turnedinto wine.

Spectroscopic methods forquality assurance

A meticulous quality controlprocedure is essential, and duringeach stage of the production pro-cess spectroscopic methods suchas FTIR spectroscopy are appliedfor quality assurance or for prod-uct characterization, for examplethe determination of must con-tent in grapes. Must is the juicecollected from grapes after crush-ing or pressing, before the pro-cess of fermentation has begun.

This application note describesthe determination of must con-tent of different types of grapesfrom the Trieste region in Italyaccording to the Brix method.The Brix value [% Brix] is meas-ured using a refractometer and isa percentage indication of thegeneral must value or sugar con-tent of the grapes. This value, inturn, indicates the potential alco-hol content of the future wine. InGermany this determination isusually measured in degreesOechsle (°OE ) and in otherEuropean regions in KMW(Klosterneuburger Mostwaage,where 1 KMW = 5 °OE), Babo,KMN and Baumé. The mustweight is an important classifica-tion criterion for the quality of awine.

This article will present FTIR asa suitable analytical technique forvery rapid measurement of must

1,5751,650 1,500 1,425 1,350 1,275 1,200 1,125 1,050 975

-0.5

-1.0

0.0

0.5

1.0

Abs

1/cm

Figure 1: Infrared spectra of liquids after subtraction of the reference (water) where green =

water difference, blue = wine and red = must. The fingerprint region is shown.

Table 1: List of the standards used

32.0

27.2

16.9

26.0

16.5

most_sample4

most_sample2

most_sample3F

most_sample2_1

most_sample1F

Spectrum

NEWS 03.2006 GB 20.09.2006 10:38 Uhr Seite 13

22.3893

22.4411

22.4413

22.3474

22.3119

22.3862

0.0138

PLS I

1

48

2432.60 – 3000.60

1122.59 – 1558.41

Yes

Must content

3

0.99905

0.00186

0.04319

Must content (% Bx)

Report of PLS Calibration

most_sampleF1and2__003





Mean value

Difference in terms of the mean value

Sample

13


scopyectro

conventional transmission modeusing a flow cell with a layerthickness of 50 µm. The cell isequipped with water-resistantCaF2 windows. As infrared radia-tion is also a form of heat radia-tion, the cell was thermostattedin order to maintain a constanttemperature in the sample. Varia-tion of temperature of the analy-tical system during spectralacquisition can lead to high meas-uring errors in the quantificationof liquid samples and errors ashigh as 0.0125 Abs per 1 °C havebeen reported.

The accuracy of the FTIR systemwith respect to the baseline canbe specified to within 0.005 Abswith water serving as reference aswell as sample. In comparison,for refractometers an error rangeof 0.1 up to 0.5 % Bx is reported,depending on the measuringrange. The temperature stabilityof the cell is achieved with anerror of ± 1 °C. In this measure-ment set-up, the water spectrumwas used as reference and meas-ured against the sample.

Quantification

In this case, as an indicator of theearly stages during wine produc-tion, the must content and itsdetermination according to Brixis considered to be a one-compo-nent system. Using mathematicalquantification models, the Brixparameter can be correlated tothe absorption of a spectrum.

Although the “must content”characteristic property is consid-ered here, the conventional cali-bration method – plotting theconcentration of one componentagainst a discrete analytical wave-length – does not lead to a suit-able quantification model. Whentaking into account the character-

istics reflected in the must con-tent, a multivariate model span-ning the entire spectrum or partsof it will be a more suitable toolfor tackling this quantificationissue.

The infrared spectrum representsall ingredients and their corre-sponding characteristics. The PLS(Partial Least Square) methodwas therefore used for this appli-cation. In PLS, a factor set issought after which represents thestandard spectral set and its com-ponents. Based on the establishedfactors, the content of a certaincomponent (in this case must)can be determined in a sample.

For this model, 5 must samples(Table 1) were determined in 10-fold using an autosampler. 48 measuring values wereacquired in the calibration (Table2). In order to test the calibrationmodel, a sample was created frommust standards 2 and 1. Theresult is listed in Table 3.

The deviation of the Brix meanvalue of five measurements with0.0138 % Bx lies below the errorthat can result from a false read-ing of 0.2 % Bx on the refracto-meter scale. Each single measure-ment lies in turn below the read-ing error.

Evaluation

This limited standard model wasmeant to illustrate that a calibra-tion of must with the aid ofinfrared spectroscopy is possible.The quality of calibration modelsis obviously dependent on thequality of the standards and ref-erence methods used. This modelcan be further improved by usingmore standards, enabling in turnmore accurate calibration. It isalso important to use a reliable

reference method, fast processingof the standards – as these are notstable over time – and tempera-ture control as the measurementsare strongly influenced by tem-perature variations.

We acknowledge with thanks the friendly

support and use of the standards provided by

the laboratory of Dr. Dust in Corni di Rosazzo

and Emanuele Canu of Shimadzu in Milan, Italy.




Table 2: Calibration model of the must application

Table 3: Testing the model using a sample representing a must content of 22,4 % Bx

No.

1

2

3

4

5

Algorithm

Number of components

Number of references

Range [1]

Range [2]

Centered data

Components

Number of factors

Correlation coefficient

MSEP

SEP

NEWS 03.2006 GB 20.09.2006 10:38 Uhr Seite 14


4

The determination of mois-ture in the quality controlof meat is important in

assessing impurities such as extrawater or salts for preservation ofthe natural product. Too muchsalt is unhealthy and may have anegative impact on the humanblood pressure system. A highwater content is problematicwhen cooking or grilling and themeat is dry and tough afterwards.

Various analysis techniques canbe used in salt determination, e.g.free energy dispersive X-ray anal-ysis. UV-VIS instrumentationanalyses the quality of meat andmeat products from a differentperspective by determination ofhydroxyproline (connective tis-sue). The FT-NIR measurementtechnique can determine severalcharacteristics from meat (such asfat and moisture) in one measure-ment without time-consumingsample preparation.

Meat mattersDetermination of moisture and fat in pork meat with measurement technique

Figure 1: FT-NIR spectra from pork meat (black line) and dried meat (red line).

The transparent boxes in blue mark the change of signals due to moisture, the blue

vertical lines mark the approx. position of the main sample signal, predominantly

-OH from water

Table 1: Meat products and typical values for fat, moisture and protein (g/100g)

as found in literature (Source: 1. FNB+FSB, Heinzig, 2002, 2. Wikipedia- Liste der

Inhaltsstoffe von Fleisch, 2010 )

Quality control

Conventional analysis in thequality control of meat needsseveral process steps and is time-consuming. This application willshow an easy and simple non-destructive method for the deter-mination of total fat and moisturein meat products with FT-NIRanalysis.

The classical applications of qual-ity control target at farm prod-ucts such as grain, milk and oily

fruits. With animal food, themoisture (OH-Bonding) is ana-lyzed as well as protein- (eggwhite, NH-Bonding), raw fibers-(fibers, CH-Bonding and other),carboxylic groups in polymers(COOH) and fat content (CH-Bonding).

A typical NIR spectrum is shownin figure 1. The NIR spectrumcontains information on combi-nation bands and overtones ofvibrations in the infrared-lightregion. In comparison to the

-0.20-0.15-0.10-0.050.000.050.100.150.200.250.30

0.40

0.50

0.60

0.70

0.80

0.90

1.00

1.10

0.35

0.45

0.55

0.65

0.75

0.85

0.95

1.05

1.151.201.25

9500 9200 8900 8600 8300 8000 7700 7400 7100 6800 6500 6200 5900 5600 5300 5000 4700 4400 4000

1/cm

Abs

.

Item Fat Moisture(water) Protein

Boiled sausage

Liver sausage

Raw bacon (ham)

Pork meat

Bacon

Cooked bacon (ham)

60.0

55.2

69.5

60.7

54.4

73.8

12.0

16.7

18.3

28.7

16.0

18.4

25.0

22.8

4.4

9.6

28.9

3.9

5


FT-NIR

Figure 2: PLS reconstruction of the measured spectrum and the difference spectrum

from measured minus calculated spectrum

Figure 3: The spread of measurement points in the range from 4 to 50 % fat in meat

and meat products. Scale is actual vs. predicted value of fat in percentage values.

Table 2: Calibration results for the fat and moisture determination in meat products

highly resolved signal structuresof the mid-infrared spectrum theNIR spectrum shows broad,smooth signal structures, makingthe whole spectrum sensitive forquantitative analysis.

Fat in pork meat

In order to establish a calibrationmodel, 104 pork-based meatproducts were analyzed. Thesources of the meat were bothsupermarkets and butchers. Theitems included low-fat pork (filetwith 5.9 % fat), a pork tongue

sausage with 20 % fat, cookedsausage with approx. 30 %, andbacon with 45 % of fat.

Reference Methods

The reference values of fat weredetermined according toCHE004W (NEN-ISO 1443/NEN-ISO 1444, Meat and meatproducts – Determination of totalfat content / Meat and meat prod-ucts – Determination of free fatcontent). The fat is extractedfrom the samples after acid hy-drolysis. The extraction was car-

ried out with petroleum etherafter which the ether was evapo-rated and the remainder wasdried and weighed (Soxhletextraction).

Sample preparation for FT-NIR measurement

The samples were homogenizedwith a mixer. A portion of themeat was transferred into a dis-posal Petri dish. Tests showedthat error due to the polymericmaterial is in an acceptable range,and even better than when usinga glass Petri dish. The StandardError of Prediction (SEP) of thecalculation was 0.157 % for mois-ture and 0.166 % for fat. Packingthe sample to the bottom of thePetri dish was beneficial. Thiscorrelated to the rule for diffusereflectance for powders, where amaterial thickness of �

0.00

0.05

0.10

0.15

0.20

0.25

0.30

0.35

0.40

0.45

0.50

0.55

0.60

0.65

0.70

0.75

0.80

0.85

0.90

0.95

1.00

1.15

1.20

1.25

10000 90009500 80008500 70007500 60006500 50005500 40004500

1/cm

Abs

.

Component Fat Moisture

Number of factors

Correlation coeff.

MSEP*

SEP**

*Mean Squared Error of Prediction, **Standard Error of Prediction

6.00000

0.98817

0.02326

0.15251

6.00000

0.97759

0.04383

0.20936

0

5

10

15

20

25

30

35

40

45

50

0 5 10 15 20 25 30 35 40 45 50

Calibration of 92 meat and meat products – fat content

Actual [%]

Pre

dict

ed [%

]

forms of meat and meat productswere analyzed – from filet tosmoked sausage and bacon. Forthe calibration model a PartialLeast Square method (PLS) wasinvestigated.

Calibration result

Report of PLS CalibrationCalibration table:Algorithm: PLS INumber of components: 1Number of references: 92Range: 4,000 - 10,000Centered data: Yes

Reconstruction

A good calibration based on PLSmodelling is indicated by theMean Squared Error of Predic-tion (MSEP), Standard Error ofPrediction (SEP) and the recon-struction of the spectra with thehelp of the factors analyzed. Infigure 2 the result of the recon-


6

minimum 3 mm is necessary, withhomogeneous particles and pack-ing of the sample.

Instrument and accessory

• IRPrestige-21 with NIR Kit• IntegratIR – Integrating sphere

for diffuse reflectance measure-ments with rough golden sur-face; the accessory was equip-ped with a rotating Petri dishholder for signal averaging overa wide surface of the meat sam-ple.

Discussion of NIR spectra

Figure 1 shows two spectra withdifferent moisture content. Thetop one is the natural meat andthe lower one is a meat powder.The diffuse reflectance FT-NIRspectra highlight the differences.The major changes are visible at:8694, 6890, 6091, 5950 and 4616 cm-1.

The first overtone from the OH-bonding is by definition around6900 cm-1 (approx. 1450 nm) forfree water. Water (in this casemoisture) can be free or bonded-style of water and other sub-stances, which will disappearunder drying conditions.

The meat sample shown wasdefined as having a water contentof 72.3 %, 19.4 % protein and 5.1 % fat (black in figure 1). Thedried sample contained 39.4 %fat (red in figure 1).

For the analysis, many validationchecks were carried out to estab-lish a robust calibration.1. The reproducibility, repeatabil-

ity and accuracy of thea. polymer-based Petri dish b. packing of meat samplec. mixed meat samples.

2. The reference methods for thedetermination of the fat, pro-tein and moisture contentswere analyzed following thespecified norms.

Based on this knowledge a cali-bration was established. Different

Figure 4: IRPrestige-21 with the NIR Kit and Pike IntegratIR –

diffuse reflection accessory

Table 3: Quality check of the FT-NIR calibration model using a certified meat sample

struction is shown. On top, themeasured spectrum is overlaidwith the recalculated spectrum.The difference between the spec-tra is visible at the bottom.

Reconstruction of the spectrumwith the help of the factorsshowed a good fit. The differencewas almost zero (violet in graphat bottom). Some noise wasapparent only in the water ranges.

A certified pork material wasused to prove the quality of thecalibration.

The reference value did not definethe reference method. Literaturementions that the Soxhlet methodis used for the determination offat, but it shows standard errorsranging from 0.41 to 1.14 %. Thisis a wide error range for onemethod and will influence thequality of the FT-NIR calibra-tion.

The calibration shown here isbased on 92 samples with fat con-tents in a range from 4 to 50 %fat in pork meat and meat prod-ucts.

Benefits

In comparison to wet chemicaltreatments of the meat, the analy-sis is simple and quick. The shortmeasurement time reduces costs,while the material is economicwith chemicals. The method saves95 % of the standard costs involved with conventional FT-NIR analysis.

The analysis results are good fora technique using diffuse reflec-tance method without tempera-ture control and a disposal sam-ple Petri dish. Compared with theSoxhlet method which is funda-mental for fat calibration theFTIR method is more sensitivebecause the SEP value of the fatcalibration is in a range of 0.15 %and for moisture 0.21 %. Com-paring the error from the disposalPetri dish which was calculated at approx. 0.166 % for fat and0.156 % for moisture, and theerror of the Soxhlet method, theNIR technique offers an alterna-tive method to the classical wetchemical treatments. The calibra-tion result is as good as the refer-ence method. Where the referencemethod is improved, the calibra-tion result will be even better.The same applies to the sample ofstandards, which can be evenmore than the approx. 100 usedin this application.



number on your reply card. Info 386

Item Sample Fat (%) Moisture (%)

Measurement NIR

Reference

Difference

Certified material

Pork

14.0299

14.30

0.2701

68.4339

68.8

0.3661

Blank page

Classification

Wax

Wax

Wax

Citronellyl ester

Citronellyl ester

Citronellyl ester

Citronellyl ester

Citronellyl ester

Inorganic (X) e.g.

phosphorus-based salts,

X-OH, X-O


8

technique allows the infraredradiation to penetrate to a depthof approximately 2 µm into thesample surface.

In the ATR technique, thermalradiation within the infraredrange is transmitted to the opticalelement from where it is directedalong a defined single-reflectionangle onto the sample, whichabsorbs and reflects the radiationand then directs it back to thedetector.

Polymers, powders and oth-er materials have been suc-cessfully analyzed using

single-reflection FTIR units. Butwhat about surface analysis of afruit? This is demonstrated in thefollowing experiment with alemon. When touching a lemonwith the fingertips, one clearlyfeels the waxy layer on the

lemon’s surface which is a naturalprotection against evaporation.

Investigation of the outermostsurface (exocarp) layer of thelemon is quite difficult as thelemon releases its fragrances uponstronger touch, while forming anoily film at the surface. The entireyellow surface, the flavedo, con-sists mainly of cellulose, essentialoils, wax, pigments, limonenesand other components.

FTIR enables straightforward,effortless analysis of a slice oflemon peel. By applying varyingcontact pressures, it was attempt-ed to capture the surface effects.

Surface analysis was carried outwith the Specac Silver GateTM

Evolution, equipped with a ZnSecrystal as optical reflection ele-ment. The measuring surface of the sample is approximately 7 mm. The applied measuring

FTIR lemon squeezer extractscitrus fragranceSingle-reflection unit and FTIR spectrophotometer

Figure 1: Slice of lemon peel: the yellow and white surface of the lemon peel was inspected via infrared spectroscopy; amplification of the image shows part of the cell structure

whereby some fluid can be observed on the cut edge between the yellow and white layer – Figure 2: A slice of lemon peel clamped into a Silver GateTM unit

Table 1: Classification of IR bands

Specac Silver GateTM Evolution,

a single-reflection unit

Figure 3 shows the result of themeasurement on the yellow sur-face of a lemon peel. The infraredspectrum features signals arisingfrom oils, wax, water and cellu-lose. After removing the lemonpeel, an oily phase remained onthe surface of the optical element.This was measured in the mid-infrared range as well. Spectraldatabase comparison of theobtained spectrum shown in Figure 4 provided a match forcitronellyl ester.

Spectrum

720

1462

~1740

~1740

1438

1370

1235

1154

881

Lemon peel

Oily phase

In all spectra

Wave number [cm -1]

Figure 1 Figure 2

9


Figure 3: Infrared spectrum of a lemon peel slice measured via an ATR

single-reflection unit

Figure 4: Mid-infrared spectrum of an oily phase originating from a lemon

peel slice

Figure 5: Infrared spectra measured on a lemon peel surface using various contact pressures of the ATR single-reflection unit

IRAffinity -1

52.5

60

67.5

75

82.5

90

97.5

4500 4000 3500 3000 2500 2000 1750 1500 1250 1000 750 500

1/cmFood citron / lemon wax layer, Silver GateTM

% T

94

95

96

97

98

99

100

101

4500 4000 3500 3000 2500 2000 1750 1500 1250 1000 750 500

1/cmFood citron / lemon wax layer, Silver GateTM

% T

4500 4000 3500 3000 2500 2000 1750 1500 1250 1000 750 500

Lemon yellow surface, Silver GateTM, highest pressure

% T

This experiment can be repeatedby applying different contactpressures. Depending on thepressure applied onto the lemonpeel surface, the infrared spec-trum will exhibit additional sam-ple component signals. In thisway it is possible to determinethe flavedo structure.

Figure 5 shows the effect of theapplied pressure on the increaseand decrease of signals in theinfrared spectrum. Low contactpressure results in the formationof a thin oily film between theoptical element and the lemonpeel. When the pressure isincreased, the oily film is pushedaside and cellulose together withother components present in thepeel become visible. The signal at880 cm-1 greatly increases in thisprocedure. A library search forthis signal strongly points towardphosphorus-containing salts. Thisis confirmed in the literature.

We will gladly send you additional infor-

mation. Please enter the corresponding

reference number on the reply card.

Info 362

1/cm

1 1 Stone 2 - 11% Whewellite, 47% Apatite, 42% Struvite

2 2 Stone 19 - 62% Whewellite, 13% Weddelite, 25% Whitlockite

3 3 Stone 17 - 56% Whewellite, 39% Weddelite, 5% Apatite

4 5 Stone 10 - 43% Whewellite, 57% Apatite




8 9 Stone 8 - 39% Whewellite, 61% Weddelite

9

APPLICATIONShimadzu News 2/2003APPLICATION Shimadzu News 2/2003

8

lysis by FTIRKidneystones

Apatite, Stru-vite, Weddelite – thesenames sound nice, but as

kidneystones they are very harm-ful to the patients. In Germany, 5 percent of the people are affect-ed at least once in their lifetime(each year 1.2 million people suf-fer from kidneystones).

The com-pounds of the stonesgive hint about the reason of theillness and appropriate stepsagainst a relapse in the future canbe taken.

At the Bosch Medicentrum, ahospital in the center of ‘s-Herto-genbosch, the Netherlands, anFTIR spectral library has been

developed consisting of lots ofkidneystones with a known

composition. Since mostof the stones are also

analysed by XRF theconcentration of the

components isknown too.

The FTIRlibrary is ahelpful tool toanalyse thepatients kid-neystones. Atall the infraredspectroscopy isa very wellknown tech-

nique doing thisstyle of analysis

[1-3]. A more newaspect using it, is the

reduction of samplepreparation and analysis time

due to high speed FTIR technolo-gy and accessory selection.

In the past the KBr pellet tech-nique taking 15 to 20 minutes was

investigated. Nowadays, it is pos-sible to use the more comfortableand easy to handle ATR tech-nique, and the sample preparationis reduced to nearly zero. Theinstrument used at the BoschMedicentrum is an FTIR-8400with Hyper IR software Version1.57 and a Golden Gate withKRS-5 lenses. With those opticsthe range is extended down to350 cm-1.

Comparison measurements havebeen done on the new FTIR-8400S (Figure 1) and the IRSolu-tion software .

The sample preparation itself isreduced to the following activi-ties. At first, the stone is grindedin a mortar to obtain a fine pow-der. This is necessary because thedifferences in particle size mayeffect the IR spectrum. Then, avery small part of this powder(only a few milligrams) is put onthe diamond-crystal of the Gold-en Gate and the following spec-trum is measured (Figure 2).

When this spectrum is searched inthe library the result will be asshown in figure 3.

The result is, that the determinedstone material consists of 11 %Whewellite, 47 % Apatite, and 42 % Struvite. In this way thechemical content and the concen-tration of the components can bedetermined quickly and easy.

Literatur:

[1] Analysis of urinary calculi by infrared spectrophotometry, Menendez Gutierrez,

R., Med. Segur. Trab., 36 (146), 53-62, 1989.

[2] Technique for the analysis of urinary calculi by infrared spectroscopy, Hesse,

A., Molt, K., J. Clin. Chem., Biochem., 20 (12), 861-73, 1982.

[3] Analysis of urinary calculiby infrared spectroscopy, Laurence, C., Dubreil, D.,

Lustenberger,P., Ann. Chim. (Paris), 1 (1), 55-63.

The photographs on page 8 and 9 are printed with permission of Dr. Gerd von

Unruh, Medizinische Universitätsklinik Bonn, Germany

Figure 4: FTIR kidneystones library created with the FTIR-8400 and Golden Gate

(KRS-5 lenses)

Fig.1: FTIR-8400S: Single-beam instru-

ment with completely encapsulated optics

Figure 3: Typical Search result using a homemade libraryFigure 2: Spectrum of a kidneystone with unknown

composition

Nuisance for millions

of people

analysis by FTIR

1 1 Stone 2 - 11% Whewellite, 47% Apatite, 42% Struvite

2 2 Stone 19 - 62% Whewellite, 13% Weddelite, 25% Whitlockite






8 9 Stone 8 - 39% Whewellite, 61% Weddelite

9


8

KidneystonesKidneystones ana

Apatite, Stru-vite, Weddelite – thesenames sound nice, but as

kidneystones they are very harm-ful to the patients. In Germany, 5 percent of the people are affect-ed at least once in their lifetime(each year 1.2 million people suf-fer from kidneystones).

The com-pounds of the stonesgive hint about the reason of theillness and appropriate stepsagainst a relapse in the future canbe taken.

At the Bosch Medicentrum, ahospital in the center of ‘s-Herto-genbosch, the Netherlands, anFTIR spectral library has been

developed consisting of lots ofkidneystones with a known

composition. Since mostof the stones are also

analysed by XRF theconcentration of the

components isknown too.

The FTIRlibrary is ahelpful tool toanalyse thepatients kid-neystones. Atall the infraredspectroscopy isa very wellknown tech-

nique doing thisstyle of analysis

[1-3]. A more newaspect using it, is the

reduction of samplepreparation and analysis time

due to high speed FTIR technolo-gy and accessory selection.

In the past the KBr pellet tech-nique taking 15 to 20 minutes was

investigated. Nowadays, it is pos-sible to use the more comfortableand easy to handle ATR tech-nique, and the sample preparationis reduced to nearly zero. Theinstrument used at the BoschMedicentrum is an FTIR-8400with Hyper IR software Version1.57 and a Golden Gate withKRS-5 lenses. With those opticsthe range is extended down to350 cm-1.

Comparison measurements havebeen done on the new FTIR-8400S (Figure 1) and the IRSolu-tion software .

The sample preparation itself isreduced to the following activi-ties. At first, the stone is grindedin a mortar to obtain a fine pow-der. This is necessary because thedifferences in particle size mayeffect the IR spectrum. Then, avery small part of this powder(only a few milligrams) is put onthe diamond-crystal of the Gold-en Gate and the following spec-trum is measured (Figure 2).

When this spectrum is searched inthe library the result will be asshown in figure 3.

The result is, that the determinedstone material consists of 11 %Whewellite, 47 % Apatite, and 42 % Struvite. In this way thechemical content and the concen-tration of the components can bedetermined quickly and easy.

Literatur:

[1] Analysis of urinary calculi by infrared spectrophotometry, Menendez Gutierrez,

R., Med. Segur. Trab., 36 (146), 53-62, 1989.

[2] Technique for the analysis of urinary calculi by infrared spectroscopy, Hesse,

A., Molt, K., J. Clin. Chem., Biochem., 20 (12), 861-73, 1982.

[3] Analysis of urinary calculiby infrared spectroscopy, Laurence, C., Dubreil, D.,

Lustenberger,P., Ann. Chim. (Paris), 1 (1), 55-63.

The photographs on page 8 and 9 are printed with permission of Dr. Gerd von

Unruh, Medizinische Universitätsklinik Bonn, Germany

Figure 4: FTIR kidneystones library created with the FTIR-8400 and Golden Gate

(KRS-5 lenses)

Fig.1: FTIR-8400S: Single-beam instru-

ment with completely encapsulated optics

Figure 3: Typical Search result using a homemade libraryFigure 2: Spectrum of a kidneystone with unknown

composition

Nuisance for millions

of people

analysis by FTIR

5


The apple is one of the most widely cultivated tree fruits. In2008, 64 million tons of apples were grown worldwide. Chinaproduced 27.5 million tons, over 40 % of this total. The US is

the second leading producer, with approx. 7 % of world production.

Research suggests that apples may reduce the risk of colon cancer,prostate cancer and lung cancer. They are a rich source of antioxidantcompounds. They may also help to combat heart diseases.

Apples are grown and consumed worldwide. They are global trav-ellers. In order to get them in good shape and best condition for mar-kets and consumers they have to be treated well and need to enjoy asafe trip.

Preparation and packing processes are automated. Modern machinessort the fruits for size and quality and pack them into a carton. Impor-tant criteria for the packing are:

1. the contents must be of the same appearance – the apples must have the same size and color

2. the packaging must protect the apples from impact3. the packaging must be new, clean and with printed marks

and information

The carton is regarded as being the best transportation device forapples. The insert has been designed particularly to shelter and alsostabilize the fruits. The insert shown here has two areas: a stiff oneframing the fruits (Figure 1 and 2), and a soft and smooth area (Fig-ure 1 and 3). In the following text the polymer foil insert of a cartontransport packaging is analyzed.

The following application shows how FTIR, EDX and ICP techniquescomplement each other. Whereas FTIR and EDX do not destroy thesamples, ICP needs sample preparation allowing better detection lim-its regarding the elements in the sample.

Modern analysis techniques

The material was analyzed with molecular spectroscopy representedby FTIR and elemental analysis through EDX and ICP technique. A benefit of FTIR and EDX is non-destruction of the sample duringthe analysis process. The advantage of the ICP is the multi-elementaldetection in one measurement, independent of the weight of the ele-ments of interest.

Analysis with FTIR IRAffinity-1

The insert sample was analyzed without chemical treatment. Since thematerial in figure 3 was very thin, transmission mode measurementwas possible. Figure 4 shows the results. The infrared spectrum showsa polypropylene spectrum as expected based on the PP sign printed onthe insert. Of interest is the view of the baseline from this measure-ment showing a strong slope.

A global travelerSpectroscopic methods in polymer packaging analysis

Figure 1: An apple in a polymer foil/insert with specific dimensions for the transport of

fruits. The red circles mark the two analysis areas shown in figure 2 and 3.

Figure 2: Surface of the black colored foil part, a polymer which is rough and stiff.

It is from the edge of the insert shown in figure 1.

Figure 3: Surface of the silver-gray colored, very thin soft foil part

3 2

The slope in the baseline of the insert is comparable with the Chris-tiansen effect of particles in a KBr pellet (potassium bromide) knownfor the transmission measurement. Knowing this, it is to be expectedthat some solid particles are part of the foil construction.

The single reflection “Silver Gate” unit equipped with a ZnSe crystal(zinc selenide) was used for surface measurements of area 2 in figure 1.The material is too thick for transmission mode analysis. �

Analysis with EDX-720

As some polysilicate and filler were found, it was interesting to research the type of filler used for the foil. Based on the match thelibrary first suggested an asbestos mineral based on magnesium (Mg), iron (Fe) and silicon (Si). However, fillers are not based on these materials. EDX quickly helped to identify the main elementswhich were bases of the mineral/filler. Calcium (Ca), silicon (Si) and iron (Fe) were found in high concentrations. The Ca signal wasextraordinary high. The light elements were not analyzed.

Analysis with ICP-9000

The presence of silicon in the samples was found by FTIR and EDX,so the samples were decomposed by a solution of 5 ml HF and 10 mlHNO3 for the ICP-analysis.


6

The measurement technique allows a penetration of 2 μm of the beaminto the sample surface. The spectrum is shown in figure 5. For theanalysis of the unknown spectra, the Shimadzu and commerciallibraries such as Sadtler were used to identify the infrared spectra.

In these libraries the base material polypropylene (PP) was found.Next major deviations were related to supra plast material and a poly-silicate (reinforcing filler silver white fibres with black inclusions)being found in the Sadtler Hummel library. The presence of this filleris correct because the silicate contained can be the cause of the tre-mendous slope of the baseline of the transmission mode. The silicateparticles generate stray light phenomena.

Instrumentation:Instrument: IRAffinity-1, ShimadzuAccessory: Silver Gate, ZnSe crystal, SpecacLibraries: Shimadzu, Sadtler – BioRad Devision

Figure 4: Transmission mode spectrum from the sample area three of the insert

(black line), green is the typical polypropylene peak from the spectral library

Figure 5: Reflection mode spectrum from the insert part surface, marked with two in

figure 1, consisting mainly of polypropylene

Figure 6: Search result for the signals differing from PP, Hummel/Sadtler Polymers,

SUPRAPLAST-PRESSMASSE TYPE 31, HARDENED (blue line)

Figure 7: Last signals search in the inorganics library showed the following:

SILVER-WHITE FIBERS WITH BLACK INCLUSIONS, Chemical Description = POLY-

SILICATE, Comments, Use: REINFORCING FILLER

0

0,15

0,30

0,45

0,60

0,75

0,90

1,05

4500 4000 3500 3000 2500 2000 1750 1500 1250 1000 750 500

1/cm

Abs

.

0,015

0,030

0,045

0,075

0,060

0,090

4500 4000 3500 3000 2500 2000 1750 1500 1250 1000 750 500

1/cm

Abs

.

0

0,15

0,30

0,45

0,60

0,75

0,90

1,05

4500 4000 3500 3000 2500 2000 1750 1500 1250 1000 750 500

1/cm

Abs

.

0

0,15

0,30

0,45

0,60

0,75

0,90

1,05

4000 3500 3000 2500 2000 1750 1500 1250 1000 750 500

1/cm

Abs

.

7


Table 1: Measurement results of polypropylene after microwave digestion [all values in mg/L]

blank 1

blank 2

p1a

p1b

p2a

p2b

Control 10 ppm

Sample name

all data in mg/L

Ca

4.71

6.01

14,760

14,580

14,750

14,600

9.94

Cd

0.050

0.030

2.63

2.35

2.93

2.83

10.3

Fe

1.35

1.17

359

350

589

534

10.4

K

1.75

0.535

62.3

62.2

98.3

98.7

9.89

Mg

1.35

0.25

3,650

3,620

5,840

5,680

10.2

Na

2.12

1.46

44.3

40.2

103

105

10.2

Ni

0.135

0.235

12.1

10.7

6.20

7.50

10.3

Pb Si Zn

0.134

0.145

27.0

21.0

39.9

34.9

10.1

30.1

11.9

6,190

6,400

2,300

2,400

9.97

0.450

0.257

14.9

14.3

18.6

18.3

10.2

Using this decomposition medium the silicon was completely dis-solved. The sample was measured with the Shimadzu ICPE-9000. Theresult is shown in the following table. A double measurement fromeach sample was performed as well as a blank and a control standardto proof the accuracy of the method.

Conclusion

The polymer foil contains components made of materials other thanpolypropylene. Based on the characteristics of the material and thetarget usage, the PP has been treated with some fillers. The fillers are

0.000

0.100

0.075

0.050

0.025

0.000

0.100

0.200

0.300

0.400

Ti-U

0.00

0.00 5.00 10.00

10.00 20.00 30.00

[keV]

[keV]

[cps

/uA

]

Na-Sc

[cps

/uA

]

Figure 8: EDX-Analysis of two Polymer samples (apple film grey and apple film black), measured from 0 - 40 keV and 0 -15 keV, the peaks of Calcium (Ca), Silicon (Si), Iron (Fe)

and Potassium (K) are clearly visible

usually low-priced minerals such as talc, sand, calcium carbonate andhematite. The analysis time with all three instruments was less than 15minutes.

Talc: Mg3Si4O10(OH)2 · Sand/quartz: SiO2 · Feldspar: KAl Si3O8

Calcium carbonate: CaCO3 · Hematite: Fe2O3

We will gladly send you further information. Please note the appropriate number on your


Blank page

9


8

The determination of pro-teins in quality control is awell-known technique in

UV-VIS spectroscopy. AlthoughUV-VIS spectroscopy focusesmainly on the simple, fast deter-mination of concentration, FTIRspectroscopy is the method ofchoice for identification andstructure recognition.

„Golden Gate“ in combinationwith Shimadzu FTIR instrument.

The instrument parameters wereoptimised using the HyperIRsoftware, which can handleinstrument control as well as allrequired data handling tasks. The „Golden Gate“ is a single-reflectance ATR measuring unit,which uses a diamond as refract-ing medium. Liquid sample issimple dropped to the diamondsurface and measured spectro-scopical. Solide samples can bepressed against the diamondusing a sapphire.

For sample pre-treatment theprotein bovine serum albuminwas first degraded using trehaloseand subsequently lyophilised.Trehalose was used to maintainthe secondary structure of theprotein in the freeze-dried state.According to Pretrelski (1) thepercentage of the intact second-ary structure can be correlateddirectly with the biological activi-ty of the protein. For optimisa-tion, spectra of native albumin inaqueous and freeze-dried form as

well as samples of three differenttrehalose preparations weremeasured.

Comparison of the spectra shows that trehalose is a suitablereplacement for water withrespect to maintaining the sec-ondary protein structure. Theinvestigated IR amide bandsclearly show the differencesbetween the individual prepara-tions.

Figures 1-3 show a step-by-stepoverview of the data handlingprocedure. The second-orderderivative spectra in the amide Iregion are shown in Figure 4.

Reference

Prestrelski, S.J., T. Arakawa, et al.

(1993). „Separation of freezing- and

drying induced denaturation of

lyophilized proteins using stress-specif-

ic stabilization. II Structural studies

using infrared spectroscopy.“

Archives of Biochemistry and

Biophysics 303(2): 465-73.

Flawless ident ificationDetermination of the secondary structure of a protein using

This application presents thedevelopment of a new qualitycontrol method via the secondarystructure of a protein. Themethod called for a fast, simpleand non-destructive sample pre-treatment step and therefore theprotein sample was freeze-dried.The analyses was carried outusing single-reflectance accessory

Figure 1: Raw data spectrum of a lyophilised bovine serum albumin sample with

trehalose pre-treatment

Figure 2: Spectra of BSA smoothed according to Savitsky-Golay in 9 data points

Figure 4: Second order derivative spectrum of BSA in the amide I region. A = BSA in aqueous solution, B = freeze-dried from

an aqueous solution, C = freeze-dried from a trehalose solution (low), D = freeze-dried from a trehalose solution (medium),

E = freeze-dried from a trehalose solution (high)

Figure 3: Second order derivative spectrum (Savitsky-Golay, 7 data points) of

lyophilised BSA with trehalose. Arrow: range of amide I IR bands




Info 262

A more detailed description

of this publication is available in

Labo 7/2002.

0,05

0,10

0,15

0,20

0,25

0,30

0,35

4000,0 3000,03500,0 2500,0 2000,0 1750,0 1500,0 1250,0 1000,0 750,0

AB

S

1/cm

0,05

0,10

0,15

0,20

0,25

0,30

0,35

4000,0 3000,03500,0 2500,0 2000,0 1750,0 1500,0 1250,0 1000,0 750,0

AB

S

1/cm

0,05

0,10

0,15

0,20

0,25

0,30

0,35

4000,0 3000,03500,0 2500,0 2000,0 1750,0 1500,0 1250,0 1000,0 750,0

AB

S

1/cm

1700,0 1680,0 1660,0 1640,0 1620,0 1600,0

1/cmA B C D E

1700,0 1680,0 1660,0 1640,0 1620,0 1600,0

1/cm1700,0 1680,0 1660,0 1640,0 1620,0 1600,0

1/cm1700.0 1680.0 1660.0 1640.0 1620.0 1600.0

1/cm1700.0 1680.0 1660.0 1640.0 1620.0 1600.0

1/cm

J.D.H. Leebeek, Rephartox Laboratories Amsterdam and M. Egelkraut-Holtus, Shimadzu Germany

IMPRINTShimadzu NEWS, Customer Magazin of Shimadzu Deutschland GmbH, Duisburg

Publisher:

Shimadzu Deutschland GmbH

Albert-Hahn-Straße 6 -10

47269 Duisburg

Tel.: (0203) 7687-0

Fax: (0203) 766625

Email: [email protected]

http://www.shimadzu.de

Editorial Team:

Uta Steeger · Tel.: (0203) 7687-410

Susanne Bieber · Ralf Weber


ME Werbeagentur GWA · Düsseldorf

Circulation: 22.000 Copies

©Copyright:

Shimadzu Deutschland GmbH, Duisburg,

September 2002.


Corporation.

FTIR

9


8

The determination of pro-teins in quality control is awell-known technique in

UV-VIS spectroscopy. AlthoughUV-VIS spectroscopy focusesmainly on the simple, fast deter-mination of concentration, FTIRspectroscopy is the method ofchoice for identification andstructure recognition.

„Golden Gate“ in combinationwith Shimadzu FTIR instrument.

The instrument parameters wereoptimised using the HyperIRsoftware, which can handleinstrument control as well as allrequired data handling tasks. The „Golden Gate“ is a single-reflectance ATR measuring unit,which uses a diamond as refract-ing medium. Liquid sample issimple dropped to the diamondsurface and measured spectro-scopical. Solide samples can bepressed against the diamondusing a sapphire.

For sample pre-treatment theprotein bovine serum albuminwas first degraded using trehaloseand subsequently lyophilised.Trehalose was used to maintainthe secondary structure of theprotein in the freeze-dried state.According to Pretrelski (1) thepercentage of the intact second-ary structure can be correlateddirectly with the biological activi-ty of the protein. For optimisa-tion, spectra of native albumin inaqueous and freeze-dried form as

well as samples of three differenttrehalose preparations weremeasured.

Comparison of the spectra shows that trehalose is a suitablereplacement for water withrespect to maintaining the sec-ondary protein structure. Theinvestigated IR amide bandsclearly show the differencesbetween the individual prepara-tions.

Figures 1-3 show a step-by-stepoverview of the data handlingprocedure. The second-orderderivative spectra in the amide Iregion are shown in Figure 4.

Reference

Prestrelski, S.J., T. Arakawa, et al.

(1993). „Separation of freezing- and

drying induced denaturation of

lyophilized proteins using stress-specif-

ic stabilization. II Structural studies

using infrared spectroscopy.“

Archives of Biochemistry and

Biophysics 303(2): 465-73.

Flawless identDetermination of the secondary structure of a

This application presents thedevelopment of a new qualitycontrol method via the secondarystructure of a protein. Themethod called for a fast, simpleand non-destructive sample pre-treatment step and therefore theprotein sample was freeze-dried.The analyses was carried outusing single-reflectance accessory

Figure 1: Raw data spectrum of a lyophilised bovine serum albumin sample with

trehalose pre-treatment

Figure 2: Spectra of BSA smoothed according to Savitsky-Golay in 9 data points

Figure 4: Second order derivative spectrum of BSA in the amide I region. A = BSA in aqueous solution, B = freeze-dried from

an aqueous solution, C = freeze-dried from a trehalose solution (low), D = freeze-dried from a trehalose solution (medium),

E = freeze-dried from a trehalose solution (high)

Figure 3: Second order derivative spectrum (Savitsky-Golay, 7 data points) of

lyophilised BSA with trehalose. Arrow: range of amide I IR bands




Info 262

A more detailed description

of this publication is available in

Labo 7/2002.

0,05

0,10

0,15

0,20

0,25

0,30

0,35

4000,0 3000,03500,0 2500,0 2000,0 1750,0 1500,0 1250,0 1000,0 750,0

AB

S

1/cm

0,05

0,10

0,15

0,20

0,25

0,30

0,35

4000,0 3000,03500,0 2500,0 2000,0 1750,0 1500,0 1250,0 1000,0 750,0

AB

S

1/cm

0,05

0,10

0,15

0,20

0,25

0,30

0,35

4000,0 3000,03500,0 2500,0 2000,0 1750,0 1500,0 1250,0 1000,0 750,0

AB

S

1/cm

1700,0 1680,0 1660,0 1640,0 1620,0 1600,0

1/cmA B C D E

1700,0 1680,0 1660,0 1640,0 1620,0 1600,0

1/cm1700,0 1680,0 1660,0 1640,0 1620,0 1600,0

1/cm1700.0 1680.0 1660.0 1640.0 1620.0 1600.0

1/cm1700.0 1680.0 1660.0 1640.0 1620.0 1600.0

1/cm

J.D.H. Leebeek, Rephartox Laboratories Amsterdam and M. Egelkraut-Holtus, Shimadzu Germany

IMPRINTShimadzu NEWS, Customer Magazin of Shimadzu Deutschland GmbH, Duisburg

Publisher:

Shimadzu Deutschland GmbH

Albert-Hahn-Straße 6 -10

47269 Duisburg

Tel.: (0203) 7687-0

Fax: (0203) 766625

Email: [email protected]

http://www.shimadzu.de

Editorial Team:

Uta Steeger · Tel.: (0203) 7687-410

Susanne Bieber · Ralf Weber


ME Werbeagentur GWA · Düsseldorf

Circulation: 22.000 Copies

©Copyright:

Shimadzu Deutschland GmbH, Duisburg,

September 2002.


Corporation.

Figure 2: Representation of the theoretical position of the –OH (red) und –OD (blue)

valence vibrational bands in an infrared spectrum

Figure 3: Spectrum of deuterium-labelled water 1 % D2O (L = 0.1 mm),

� (HDO) = 2500 cm-1

4000.00 3000.00 2000.00 1000.00 wavenumber

= H

= DO

2800.0 2000.0 1800.0 1600.0 1/cm2400.0

1.0

3.0

2.0

5.0

4.0

Abs

FTIR spectrometry is particularly well suited to the identificationof compounds in all kinds of aggregation states. A classic infraredspectrometric analysis is that of solids, which are embedded in

potassium bromide (KBr) and pressed into pellets. Another applicationarea is the measurement of solvents that are preferably water-free, usingcells.

Water has always been considered to interfere with infrared measure-ments and was removed via suitable methods, for instance drying. Waterwith its –OH vibrational groups generates a strong, completely overlap-ping spectrum (Figure 1).

The purpose of this application is to demonstrate that not only waterwith its –OH group can be analysed, but also ‘light’ isotopically labelledwater with its –OD groups. The analysis is not limited to the identifica-tion of the molecular vibrations but also attempts the quantitative deter-mination in simple matrices such as water and even in complex matricessuch as body fluids.

Sensitive, accurate and fast – the FTIR technique

In addition to high sensitivity, accuracy and reproducibility, FTIR alsooffers another important analytical criterion: speed of analysis. We willshow that fast FTIR measurement generates results that are comparablewith the more complex and time-consuming mass spectrometric deter-minations commonly used for these types of analyses*.

To illustrate the features of the two types of analysis, very differentapplication areas are selected. In natural water – in this study the Russianriver Neva – the percentage of deuterated water is of interest from anecological point of view. In the analysis of body fluids, deuterated wateris often used to study metabolic processes. One such application is thestudy of the exchange of substances between mother and child during


10

Abs

4000.0 3000.0 1500.0 1000.0 500.0 1/cm2000.0

0.25

0.5

0.75

Figure 1: IR spectrum of water (L = 0.02 mm), where L = pathlength of the cell

breastfeeding. A control group of ten mothers were given a known con-centration of deuterated water to drink. After a certain period of time,samples of breast milk and saliva were collected from mother and child.These samples were analysed via FTIR.**

Theory

The effect of substitution with deuterium against hydrogen atoms causesa shift in valence vibrational bands. The literature reports the vibrationalbands �–OH = 3643 and �–OD = 2651 cm-1 in the gas phase, and themethod of calculation. Considering*** the literature data and looking atthe bands at approximately 3330 cm-1, which corresponds to the –OHvibration of the liquid aggregational state of water, the position of thecorresponding –OD vibration is calculated to be approximately 2440 cm-1. Figure 2 shows the theoretical positions of the valence vibra-tional bands.

In practice, both applications discussed here show a wavenumber rangeof 2600 up to 2400 cm-1 for the –OD vibrations (Figure 3). When thedeuterium concentration in water is low, only the HDO molecule isformed because the equilibrium is shifted towards the right-hand side ofthe equation: H2O + D2O � 2HDO k = 4.0 (300 °K)*.

From the above, it is concluded that the concentrations are interrelatedas follows: CHDO = 1-CHDO, i.e. the concentration CHDO is proportionalto D. In addition, during the analysis one has to take into account thatwater exhibits a high absorption band in this range and that even theCO2 doublet signal can interfere with the determination. This imposesextra requirements on the measuring technique and the detectionmethod.

Accessories

Classical measuring cells with water-resistant CaF2 windows are used inboth applications. In the case of natural water, cells with pathlengths of0.1 mm and 0.2 mm are used. For the acquisition of water spectra (Figures 1 and 3) pathlenghts of 0.1 mm and 0.02 mm were used and a0.2 mm pathlength was used for the –OD determination within the rangeof the detection limits. The measurements were carried out on ShimadzuFTIR-8000 series instruments.

Sample preparation

Saliva and breast milk samples were centrifuged and the samples weresubsequently transferred to the IR measuring cell. After IR measure-

Fast determination of deuterated

11


ment, the samples were analysed via MS. The natural water samples weretransferred directly into the cells. These minimal sample preparationsteps are sufficient to meet the requirements for fast and relatively inter-ference-free sample preparation and measuring techniques.

Evaluation of the measuring results

The analysis of deuterated water is based on the accuracy provided viaFTIR techniques and consequently the accuracy in the differencebetween absorption of a sample with respect to a standard. Water with aknown percentage of deuteration is used. Water with higher or lowerdeuterium content can be determined against a known reference. In thecase of natural water analysis, the standard was a water sample spikedwith D2O at a deuterium content of 0.0018 %, which had been deter-mined via comparison measurement using MS. The sample was unalteredwater from natural origin that usually exhibits a deuterium content of0.014 – 0.016 %. In order to confirm these values, water from the riverNeva was used for analysis.

For representation of the –OD bands, the ‘light’ water spectrum of thestandard (Figure 5) was subtracted from the Neva riverwater spectrum(Figure 4). The result is shown in figure 6 with the maximum of the–OD bands at approximately 2500 cm-1. The analysis of the sampleshows a value of 0.0146 % deuterium. The MS analysis results in a valueof 0.0143 % deuterium.

In the case of body fluids, the interpretation of the spectra was carriedout in the same way. Standard spectra were acquired and these were sub-tracted from the sample spectra. The standard spectra were generatedthrough addition of D2O to water. The analysis was carried out as atime-dependent experiment. Therefore a reference spectrum wasobtained for the body fluids at a time t = 0, without the influence of thedeuterated water. The results of the body water kinetics, when compar-ing IR with MS analyses, are shown in the following table. These Fbmvalues, in relation to the total composition of breast milk consisting ofapproximately 87 % water, result in a calculated volume of 850 mL

breast milk, and this corresponds to the average milk-intake of infantsper day (Table 1).

Conclusions

The high sensitivity of this method is based on high baseline stability,high signal-to-noise ratio and data accuracy of the photometric method.For the natural water sample, a detection limit of 0.003 % deuteriumcould be obtained. Furthermore, FTIR was proven to be a fast tech-nique, as only 10 minutes were needed from sample preparation to dataevaluation. For body fluids, representing a more complex sample matrix,concentrations less than 200 ppm could be detected via FTIR. FTIRmeasurements with respect to water kinetics are all comparable with thereference MS method.

* Yu. Predtechenskii and N. Kholodova, „Rapid Determination of Deuterium

Content of Water by FTIR-8400“, Russian Scientific Center Applied

Chemistry, St. Petersburg, Russia

** Zewditu Getahun, Rachel Elsom, Hailemichael G/sellasie, Leslie John Charles

Buck, Yonas Taffese, Anthony Wright, Graham Jennings, „Breast milk intake

measured by deuterium kinetics in mother-infant pairs in Adis Ababa”, Ethiopia.

J. Health Dev. 1999; 13(3): 271 - 280

*** Günzler, Böck,“IR-Spektroskopie“, Taschentext 43/44, 1975

We will gladly send you further information. Please note the appropriate number on

your reader reply card. Info 275

Thanks to the Shimadzu UK office for providing the paper „The measurement of

Deuterium in Body Water using an FTIR“, prepared by Leslie J.C. Buck, Medical

Research Council, Resource Centre for Human Nutrition Research, Cambridge, and

Russ J. Gill, Shimadzu (UK), 1999 and Dr. Ilia Grinshtein, Russian Scientific Center

for Applied Chemistry, St. Petersburg, Russia, 2003, for the natural waters topic.

ter in body fluids and natural waters

Figure 4: Spectrum of Neva riverwater (L= 0.2 mm)

2800.0 2600.0 2550.0 2500.0 2450.0 2400.0 1/cm2750.0 2700.0 2650.0

1.25

1.75

1.5

Abs

Figure 5: Spectrum of deuterated water with deuterium concentration of

0.0018 % (L= 0.2 mm)

2800.0 2600.0 2550.0 2500.0 2450.0 2400.0 1/cm2750.0 2700.0 2650.0

1.25

1.75

1.5Abs

Figure 6: Difference spectrum of Neva riverwater and deuterated water

Subtracted Data: NEVA02.IRS -1 * LIGHT02.IRS

2800.0 2600.0 2550.0 2500.0 2450.0 2400.0 1/cm2750.0 2700.0 2650.0

0.0

0.015

0.01

0.005

-0.005

Abs

M = Fbm/0.87 (mL/day)

880 MS

850 FTIR

Water from mother to child (Fbm)

0.760 + -0.10 MS

0.74 + -0.10 FTIR

Table 1: Results after acquisition and evaluation of the deuterated samples,

where Fbm represents the average continuous transport of water from mother to

infant. M is the conversion factor for the total volume of breast milk exchanged.

MS = mass spectrometry, FTIR = Fourier transform infrared spectrometry

water in body fluids and natural water

Figure 2: Representation of the theoretical position of the –OH (red) und –OD (blue)

valence vibrational bands in an infrared spectrum

Figure 3: Spectrum of deuterium-labelled water 1 % D2O (L = 0.1 mm),

� (HDO) = 2500 cm-1

4000.00 3000.00 2000.00 1000.00 wavenumber

= H

= DO

2800.0 2000.0 1800.0 1600.0 1/cm2400.0

1.0

3.0

2.0

5.0

4.0

Abs

FTIR spectrometry is particularly well suited to the identificationof compounds in all kinds of aggregation states. A classic infraredspectrometric analysis is that of solids, which are embedded in

potassium bromide (KBr) and pressed into pellets. Another applicationarea is the measurement of solvents that are preferably water-free, usingcells.

Water has always been considered to interfere with infrared measure-ments and was removed via suitable methods, for instance drying. Waterwith its –OH vibrational groups generates a strong, completely overlap-ping spectrum (Figure 1).

The purpose of this application is to demonstrate that not only waterwith its –OH group can be analysed, but also ‘light’ isotopically labelledwater with its –OD groups. The analysis is not limited to the identifica-tion of the molecular vibrations but also attempts the quantitative deter-mination in simple matrices such as water and even in complex matricessuch as body fluids.

Sensitive, accurate and fast – the FTIR technique

In addition to high sensitivity, accuracy and reproducibility, FTIR alsooffers another important analytical criterion: speed of analysis. We willshow that fast FTIR measurement generates results that are comparablewith the more complex and time-consuming mass spectrometric deter-minations commonly used for these types of analyses*.

To illustrate the features of the two types of analysis, very differentapplication areas are selected. In natural water – in this study the Russianriver Neva – the percentage of deuterated water is of interest from anecological point of view. In the analysis of body fluids, deuterated wateris often used to study metabolic processes. One such application is thestudy of the exchange of substances between mother and child during


10

Abs

4000.0 3000.0 1500.0 1000.0 500.0 1/cm2000.0

0.25

0.5

0.75

Figure 1: IR spectrum of water (L = 0.02 mm), where L = pathlength of the cell

breastfeeding. A control group of ten mothers were given a known con-centration of deuterated water to drink. After a certain period of time,samples of breast milk and saliva were collected from mother and child.These samples were analysed via FTIR.**

Theory

The effect of substitution with deuterium against hydrogen atoms causesa shift in valence vibrational bands. The literature reports the vibrationalbands �–OH = 3643 and �–OD = 2651 cm-1 in the gas phase, and themethod of calculation. Considering*** the literature data and looking atthe bands at approximately 3330 cm-1, which corresponds to the –OHvibration of the liquid aggregational state of water, the position of thecorresponding –OD vibration is calculated to be approximately 2440 cm-1. Figure 2 shows the theoretical positions of the valence vibra-tional bands.

In practice, both applications discussed here show a wavenumber rangeof 2600 up to 2400 cm-1 for the –OD vibrations (Figure 3). When thedeuterium concentration in water is low, only the HDO molecule isformed because the equilibrium is shifted towards the right-hand side ofthe equation: H2O + D2O � 2HDO k = 4.0 (300 °K)*.

From the above, it is concluded that the concentrations are interrelatedas follows: CHDO = 1-CHDO, i.e. the concentration CHDO is proportionalto D. In addition, during the analysis one has to take into account thatwater exhibits a high absorption band in this range and that even theCO2 doublet signal can interfere with the determination. This imposesextra requirements on the measuring technique and the detectionmethod.

Accessories

Classical measuring cells with water-resistant CaF2 windows are used inboth applications. In the case of natural water, cells with pathlengths of0.1 mm and 0.2 mm are used. For the acquisition of water spectra (Figures 1 and 3) pathlenghts of 0.1 mm and 0.02 mm were used and a0.2 mm pathlength was used for the –OD determination within the rangeof the detection limits. The measurements were carried out on ShimadzuFTIR-8000 series instruments.

Sample preparation

Saliva and breast milk samples were centrifuged and the samples weresubsequently transferred to the IR measuring cell. After IR measure-

Fast determination of deuterated wa

11


ment, the samples were analysed via MS. The natural water samples weretransferred directly into the cells. These minimal sample preparationsteps are sufficient to meet the requirements for fast and relatively inter-ference-free sample preparation and measuring techniques.

Evaluation of the measuring results

The analysis of deuterated water is based on the accuracy provided viaFTIR techniques and consequently the accuracy in the differencebetween absorption of a sample with respect to a standard. Water with aknown percentage of deuteration is used. Water with higher or lowerdeuterium content can be determined against a known reference. In thecase of natural water analysis, the standard was a water sample spikedwith D2O at a deuterium content of 0.0018 %, which had been deter-mined via comparison measurement using MS. The sample was unalteredwater from natural origin that usually exhibits a deuterium content of0.014 – 0.016 %. In order to confirm these values, water from the riverNeva was used for analysis.

For representation of the –OD bands, the ‘light’ water spectrum of thestandard (Figure 5) was subtracted from the Neva riverwater spectrum(Figure 4). The result is shown in figure 6 with the maximum of the–OD bands at approximately 2500 cm-1. The analysis of the sampleshows a value of 0.0146 % deuterium. The MS analysis results in a valueof 0.0143 % deuterium.

In the case of body fluids, the interpretation of the spectra was carriedout in the same way. Standard spectra were acquired and these were sub-tracted from the sample spectra. The standard spectra were generatedthrough addition of D2O to water. The analysis was carried out as atime-dependent experiment. Therefore a reference spectrum wasobtained for the body fluids at a time t = 0, without the influence of thedeuterated water. The results of the body water kinetics, when compar-ing IR with MS analyses, are shown in the following table. These Fbmvalues, in relation to the total composition of breast milk consisting ofapproximately 87 % water, result in a calculated volume of 850 mL

breast milk, and this corresponds to the average milk-intake of infantsper day (Table 1).

Conclusions

The high sensitivity of this method is based on high baseline stability,high signal-to-noise ratio and data accuracy of the photometric method.For the natural water sample, a detection limit of 0.003 % deuteriumcould be obtained. Furthermore, FTIR was proven to be a fast tech-nique, as only 10 minutes were needed from sample preparation to dataevaluation. For body fluids, representing a more complex sample matrix,concentrations less than 200 ppm could be detected via FTIR. FTIRmeasurements with respect to water kinetics are all comparable with thereference MS method.

* Yu. Predtechenskii and N. Kholodova, „Rapid Determination of Deuterium

Content of Water by FTIR-8400“, Russian Scientific Center Applied

Chemistry, St. Petersburg, Russia

** Zewditu Getahun, Rachel Elsom, Hailemichael G/sellasie, Leslie John Charles

Buck, Yonas Taffese, Anthony Wright, Graham Jennings, „Breast milk intake

measured by deuterium kinetics in mother-infant pairs in Adis Ababa”, Ethiopia.

J. Health Dev. 1999; 13(3): 271 - 280

*** Günzler, Böck,“IR-Spektroskopie“, Taschentext 43/44, 1975

We will gladly send you further information. Please note the appropriate number on

your reader reply card. Info 275

Thanks to the Shimadzu UK office for providing the paper „The measurement of

Deuterium in Body Water using an FTIR“, prepared by Leslie J.C. Buck, Medical

Research Council, Resource Centre for Human Nutrition Research, Cambridge, and

Russ J. Gill, Shimadzu (UK), 1999 and Dr. Ilia Grinshtein, Russian Scientific Center

for Applied Chemistry, St. Petersburg, Russia, 2003, for the natural waters topic.

in body fluids and natural waters

Figure 4: Spectrum of Neva riverwater (L= 0.2 mm)

2800.0 2600.0 2550.0 2500.0 2450.0 2400.0 1/cm2750.0 2700.0 2650.0

1.25

1.75

1.5

Abs

Figure 5: Spectrum of deuterated water with deuterium concentration of

0.0018 % (L= 0.2 mm)

2800.0 2600.0 2550.0 2500.0 2450.0 2400.0 1/cm2750.0 2700.0 2650.0

1.25

1.75

1.5Abs

Figure 6: Difference spectrum of Neva riverwater and deuterated water

Subtracted Data: NEVA02.IRS -1 * LIGHT02.IRS

2800.0 2600.0 2550.0 2500.0 2450.0 2400.0 1/cm2750.0 2700.0 2650.0

0.0

0.015

0.01

0.005

-0.005

Abs

M = Fbm/0.87 (mL/day)

880 MS

850 FTIR

Water from mother to child (Fbm)

0.760 + -0.10 MS

0.74 + -0.10 FTIR

Table 1: Results after acquisition and evaluation of the deuterated samples,

where Fbm represents the average continuous transport of water from mother to

infant. M is the conversion factor for the total volume of breast milk exchanged.

MS = mass spectrometry, FTIR = Fourier transform infrared spectrometry

Fast determination of deuterated water

CH3OH

H2C – O – CO – R1

HC – O – CO – R2

H2C – O – CO – R3

Triglyceride


14

fuels. This application noteshowcases the ability of ATR incombination with FTIR spec-troscopy to identify commercial-ly available biodiesel blends.While FTIR does not possess thedetection limits or industrialacceptance of GC-MS, it doesprovide the advantages of quick,non-destructive and inexpensiveanalysis. The results indicate thatit is an excellent choice as quality

Identification and quantification obased biodiesel and blends

Biodiesel is a clean-burningalternative fuel, producedfrom domestic, renewable

create a biodiesel blend. It can beused in compression-ignition(diesel) engines with little or nomodifications. Biodiesel is simpleto use, biodegradable, nontoxicand essentially free of sulfur andaromatics.*

Soy bean and rapeseed diesel arethe most promising of severalvarieties of agriculturally pro-duced alternatives to petroleum

Time-saving

resources. It contains no petro-leum, but can be mixed at anylevel with petroleum diesel to

H2C – O – H

HC – O – H

H2C – O – H

Glycerine

H3CO – CO – R1

H3CO – CO – R2

H3CO – CO – R3

FAME

Figure 1: Transesterification of triglycerides to FAMEs

Figure 2A: Typical infrared absorbance spectra of pure rape oil (blue) and a pure rape biodiesel (black).

Figure 2B: Visible are the differences in the area of 1600 to 400 cm-1

A

A

B

B

Vibration [cm-1]584.43721.38

1095.57

1458.18

2852.722922.163007.02

15


f rapeseed oiloATR-FTIR Spectroscopy

control technique for large batch-es of biodiesel blends.

Biodiesel is produced through thechemical process of transesterifi-cation. The process leaves behindethyl esters, the chemical namefor biodiesel and glycerine. Fuel-grade biodiesel must be producedin accordance with strict industryspecifications such as ASTMD6751 or EN 14214 in order toensure proper performance. Allnewer diesel powered vehiclescan run on this fuel without fur-ther modification.

The FAMEs (Fatty Acid MethylEster) resulting from transesteri-fication are regulated via DINEN 14078, specifying infraredspectroscopic analysis of bio-diesel blends in a solution ofcyclohexane.

Transesterification

Biodiesel, just like rapeseed oil, isrich on fat molecules which aretreated in a transesterificationprocess to contain a variety ofesters. The transesterification isnecessary since engines cannotrun with the natural rapeseed oilwhich is too viscous.

Figure 1 shows the graph of thechemical treatment. The largemolecule of a triglycerol is sepa-rated into the glycerine moleculeand several FAMEs.

In order to visualize the processof changing the natural productinto a technical substance, FTIRspectroscopy was used. Itdemonstrates the effect of thetransesterification on infrared

spectra using edible oil andbiodiesel prepared from rapeseed.The differences are demonstratedin Figures 2A and 2B.

The peak analysis of both spectraindicates significant differences.The change from triglycerideester groups to methyl estershows the strongest impact in theinfrared spectrum. The estergroups are generally described asR1-C(OR) = O in edible oils andas R1-C(OCH3) = O in biodiesel.R1 represents long chains ofhydrocarbons.

Additional chains representingpalmitic, stearic, oleic and linoleicacid are present in both spectra,mainly with the -CH2 hydro car-bon bands. Signal groups belong-ing to these vibrations are speci-fied as “basic oil” in Table 1. The

strongest influence of transesteri-fication can be seen in the newsignal at 1435 cm-1 which is defi-nitely the deformation vibrationof the methyl ester group.

FTIR technique is a suitablemethod for showing such differ-ences in molecular structures.

Sample preparation

Instead of the classical infraredtransmission cell the single reflec-tion technique was used formeasurements of the infraredspectra. One drop of oil wasplaced on the measurement win-dow of the single reflectionaccessory, measured and laterremoved simply with a tissue.Remaining parts were cleanedwith a drop of solvent. Using theIRPrestige-21, the measurement

result was quickly available. Theaccessory used was a singlereflection ATR equipped withKRS-5 optical element. D

Table 1: Discussion of infrared spectra based on rapeseed oil – comparison of methyl ester and triglycerol ester

Vibration [cm-1]584.43721.38 914.26964.41

1029.99-

1095.571118.711159.22

-1236.371415.751458.181743.65 2852.72 2922.163007.02

Remark

-CH2 rocking-OH out of plane-CH2 wagging in RCOCO-

(1137 shoulder) average of signals

C-CO-O-

C=O Ester-CH2

-CH2

-CH

Vibration [cm-1]584.43721.38 912.33

-

1016.491060.851095.571120.641168.86

1195.871244.09 1435.041458.181741.722852.722922.163007.02

Remark

-CH2 rocking-OH out of plane

-O-CH2-C

C-OC-CO-O-(CO)-O-CH3

C=O Ester-CH2

-CH2

-CH

Edible rape oil Biodiesel on rapeseed Basic oil

Algorithm: PLS INumber of components: 1Number of references: 6Centered data: YesComponent: FAMENumber of factors: 1Correlation coeff.: 0.99832MSEP: 0.00280SEP: 0.05288

Calibration Table


16

FAME determination withsingle reflection ATR as analternative to DIN EN 14078

This application can be adaptedto any form of biodiesel andrapeseed, soy bean or coconutbased oil. Mixtures such asbiodiesel blends can be analyzedwith infrared spectroscopy.Biodiesel is commonly mixedwith ordinary petroleum dieselfor B5 or B20 blends. B5 consistsof 5 % biodiesel and 95 %petroleum diesel. The differencesbetween biodiesel and mineral oilor petrodiesel respectively areshown in Figure 3. The sampleset for this experiment containedrapeseed biodiesel blends of B0to B100.

It is readily apparent that the twopure compounds possess distinctspectral signatures. The strongcarbonyl peak at 1750 cm-1 in thebiodiesel spectrum is representa-tive of ester functionality whichis absent in the petrodiesel spec-trum. The additional ester bandsin the fingerprint range between1200 and 1000 cm-1 in thebiodiesel spectrum provide anadditional area to distinguishbetween the two components ofinterest. The overlay spectra

(Figure 4) of the blends B0 toB100 illustrate that both the car-bonyl and the ester bands can beused to differentiate between theblends quickly and easily.

This is a very successfulapproach, so it is reasonable toalso use IR spectroscopy forquantification of FAME as statedin DIN EN 14078.

Biodiesel blends can be used forstandard diesel engines. Mostcommercial engines cannot runon 100 % biodiesel. Biodieselblends are therefore necessaryand the quality of such biofuelsand blends is regulated. Oneaspect of the control is theFAME content, which can becorrelated directly with the blendlevel.

Figure 3: Typical infrared absorbance spectra of pure mineral oil (black) and pure biodiesel

(red), visible are the differences in the areas of 3007, 1750 and 1000 to 1500 cm-1

Figure 4: IR spectra from petrodiesel and biodiesel blends

Table 2: PLS calibration result for

rapeseed biodiesel blends

DIN EN 14078 describes theprocedures to obtain best trans-mission mode spectra of standardsolutions. The biodiesel blend isdissolved – in this case in cyclo-hexane – and this mixture istransferred to a liquid cell andmeasured using FTIR. Target ofthis application note is to showthat the ATR technique is a sim-ple method for the quantitativedetermination of FAME withoutany chemical treatment.

To determine the contents of theFAME, the PLS (Partial LeastSquare) quantitative modellingwas used. Figure 4 shows a dis-tinct signal development of theFAME in the infrared spectrum.Six blends of rapeseed biodieselwere prepared for this calcula-tion: B33.33, B25, B20, B16.67,B14.28 and B10.

Summary: time-saving ATR technique

The best quality of infrared spec-tra can be achieved with trans-mission mode spectra preparedfrom a liquid in a liquid cell.However, the preparation is time-consuming: Standards have to beprepared in solvent, a calibrationhas to be carried out for the

selected liquid cell and the cellhas to be cleaned in order toavoid contamination. The treat-ment includes some possibleproblems with regard to cellthickness and dilution of the sam-ples. The necessary time frame isabout 10 minutes per sample. TheATR technique can be handledwithin two minutes without anysample preparation and is favor-able since there is no need forpreparation of a standard solutionof cyclohexane and sample solu-tions for the analysis.

This simple test of six samplesalready correlates with correla-tion coefficient of 0.99832. Thisrough model demonstrates that itwill work properly. Of course,the results will become muchmore robust with the inclusion ofmore standards. Materials usedwere rapeseed biodiesel and min-eral oil from the market insteadof standardized solutions.

*Source: www.biodiesel.org


It is said that the truth is foundin wine. This is however onlypartially true, at least when it

applies to pricing. Not only thequality of the wine, but other fac-tors contribute to the final price -the bottle for instance, distribu-tion and even the type of corkused. This small piece of nature atthe end of the bottleneck givesthe wine an emotional added val-ue and underlines the authenticityof wines and champagnes.

For the beverage industry there isa difference in price between aone piece cork and a cork pressedfrom granulate material. Whatapplies to both is, that theyshould be easily pulled from thebottle. Both, paraffin and modernpolysiloxane are applied to thecork surface as lubricants. Thesecompounds also contribute to theoveral price of the wine.

In times of heavy competition formarket share, the beverage indus-try pays close attention to whattheir competitors are doing and totheir own market research. Theyhave a keen eye for their competi-tors, their marketing strategiesand pricing policies. An impor-tant factor is product quality con-trol, which included the cork andthe paraffin or polysiloxane lubri-cants used.

Infrared spectroscopy can behelpful in the task of identifyingthese compounds. The ShimadzuFTIR-8400 was equipped with asingle reflectance unit whichallows surface analysis wherebythe infrared beam can penetratethe sample surface to a depth ofapproximately 2 µm. The corksare first cut into slices in order toallow the surface to be measured.The infrared spectrum of the sur-

face measurement is carried outfor polydimethylsiloxane andsuberine (suberine acid diethyl-ester). The results have shownthat depending on the function ofthe cork, the polymethylsiloxanelayer has a varying thickness.Research on a small sample ofEuropean wine bottle corks showthat paraffin is used less frequent-ly than siloxanes.

The conclusions that the beveragemanufacturers draw from thesetypes of analyses are, of course,proprietary. What is important isthat the product pays off. Andthat is indeed the case, when theconsumer is satisfied with respectto quality, price and what theproduct promises. And vice versa,who does not recognise this: acork that is stuck in a wine bottle?Or the champagne bottle wherethe cork does not want to pop?


16

What makes a Cork pop?FTIR identifies lubricants

IR spectra: Left: top suberine acid diethylester; bottom: polydimethylsiloxane, Right: top: cork of a wine bottle;

bottom: cork of a champagne bottle

Separation of a standard test mixture on a narrow-bore C18 column

Reproducibility of retention times and peak areas

RT Detector A (254 nm) Area

The efficient use of materi-als and capital, the care forthe environment and the

use of resources is not only animportant economic necessity foreach enterprise; it also stronglyaffects the company’s perceptionand reputation in general. Andthis is not only true for largequantities, also the repeated useof small amounts of solvents can

add up and pollute the environ-ment.

Time has already been mentionedas an important economic factorwhen considering the use of thecompact LC-2010 HPLC system.Based on its fast injection routine(15 seconds/standard injection)the instrument is particularly wellsuited for high sample throughput

and offers a signifi-cant reduction inanalysis time forroutine analyses,compared with con-ventional instru-ments. But whatabout the reductionof solvent con-sumption? Thisaspect becomesmore and moreimportant in theassessment of ana-lytical methods dueto increased needfor cost savings inlarge laboratories.

In HPLC, using optimised ana-lytical methods, short analysistimes and small column diameterscan generally reduce solvent con-sumption. For example, bydecreasing the column diameterby a factor of 2, a saving of 70 to80 % of solvents used can beachieved. Cost savings are partic-ularly effective in those caseswhere highly toxic solvents areemployed that require expensivedisposal measures. Next to thecost saving when purchasing thesesolvents, the lower disposal costscontribute to further cost reduc-tions. The reduction of the use ofhazardous solvents is also desir-able from an ecological point ofview.

In order to meet these require-ments for modern analyticalinstruments, Shimadzu has devel-oped all HPLC systems toinclude narrow-bore applicationsin their specifications. In general,all standard systems allow the useof analytical columns with aninner diameter of 2 mm – withoutany further instrument modifica-tions. To further improve the

Quick, Reliable, Economical:The new Compact ClassNarrow-bore HPLC with the LC-2010

0.0

0.1

0.2

0.3

0.4

0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0

Volt

Minutes

Pea

k 1

Pea

k 2

Pea

k 3

Pea

k 4

resolution of two closely elutingpeaks, the standard flow cell

of most detectors can beexchanged with a semi-

micro cell.

An example of the performance ofthe compact LC-2010 HPLC sys-tem is shown for asemi-micro appli-cation. In order toimprove the sepa-ration of the two

peaks of the testmixture in the middle

of the chromatogram,the standard flow cell of

the detector (volume: 8 µL) wasreplaced by a semi-micro flowcell (volume: 2 µL). The repro-ducibility of a 5 µL injection wasdetermined for 10 repeated injec-tions of a standard mixture. A 2 x100 mm Hypersil C18 columnwas used as analytical column.The chromatograms are presentedin Figure 1. The analytical resultswith respect to the reproducibili-ty of retention times and peakareas (Table 1) demonstrate theexcellent performance of the sys-tem. An injection reproducibilitywith a relative standard deviationof less than 0.1 % for 10 injec-tions indicates that the LC-2010can also be used for micro-HPLCapplications with column diame-ters of 1 mm.

Due to fast injection routines andlow solvent consumption withproven narrow-bore compatibili-ty, the LC-2010 is an ideal instru-ment for routine analysis usingHPLC and UV detection.

Info 247

Run#

1

2

3

4

5

6

7

8

9

10

Mean

Std Dev

% RSDCork – its properties

Cork contains suberine, lignine and

cellulose. These keep cork afloat.

Cork cells consist of a cellulose

skeleton which is cross-linked with

lignine. Together this forms the

backbone for suberine, a natural

polymer ester.

The physical properties of cork are

also interesting. As a natural raw

material, cork has a limited perme-

ability to liquids and gases, is heat-

and sound resistant, elastic and

chemically neutral. This makes cork

a very attractive material for home

decorating, finds use in life jackets

and seals wine and champagne

bottles.

Peak 1

0.753

0.753

0.753

0.753

0.753

0.753

0.753

0.753

0.753

0.753

0.753

0.0000

0.000

Peak 2

1.510

1.510

1.510

1.510

1.510

1.510

1.510

1.510

1.510

1.510

1.510

0.0000

0.000

Peak 3

1.887

1.887

1.887

1.883

1.883

1.883

1.883

1.883

1.883

1.883

1.884

0.0016

0.085

Peak 4

3.333

3.330

3.330

3.330

3.330

3.330

3.330

3.330

3.330

3.330

3.330

0.0011

0.032

Peak 1

206280

206481

206463

206373

206292

206528

206269

206311

206220

206302

206352

104.2

0.050

Peak 2

728003

728855

728632

728547

728424

728683

728119

728421

728174

728210

728407

276.3

0.038

Peak 3

384818

385342

384991

384923

384834

385185

384792

384998

384739

384879

384951

189.4

0.049

Peak 4

568964

569494

569153

569200

568965

568843

568425

568903

568410

568191

568855

403.6

0.071




Info 246

17

LC-2010 HPLC System


4

According to estimations(European Commissionstandardization mandate,

M/425), carelessly unattendedcigarettes cause some 14.000 firesevery year in the EU, with 7,000fatalities, 2,500 injuries andaround 50 million euro of materi-al damage. How can moleculargastronomy decrease the numberof these accidents? Spherificationis one of many applications ofmolecular gastronomy whichcombine unconventional texturesand flavors. It’s a process of turn-ing liquid juice into juice-filledpearls.

To produce these pearls somealginate – a substance derivedfrom algae – is simply dissolvedin a juice. Droplets of the juiceare then dropped into a calcium

Caviar and cigarFTIR-ATR analysis of alginate on cig

water bath. The calcium from thewater bath reacts immediatelywith the alginate in the juice toform a film around the droplet.Spheres of juice are thus createdwhich look just like caviar (Fig-ure 1). How is fake caviar relatedto cigarettes?

Fake caviar and its relation-ship to cigarettes

Cigarettes are a source of heatand therefore represent a firehazard. They can ignite materialssuch as furniture or textiles. Self-extinguishing cigarettes can re-duce the number of accidentscompared to unattended ciga-rettes. These cigarettes are pro-duced by adding two specialretardant bands to the cigarettepaper during manufacturing.

Figure 1: Alginate based Caviar prepared with fluoresceine excited with analytical

wavelength of 460 nm in a Shimadzu fluorescence spectrophotometer RF-5301PC:

it is fluorescent caviar filled into a 1 cm quartz cell

Figure 2: View of a cigarette and zoom onto one form of the cigarette paper. In this

case the rings which help to burn the cigarette continuously are apparent.

Figure 3: Structure of alginate acid base

of the sodium alginate salt

Figure 4: Structure of a common cellu-

lose molecule

We will gladly send you additional information. Please enter the

corresponding number on the reply card or order via Shimadzu’s

News App or News WebApp.

Info 401

5


These bands act as speed bumpsby decreasing the flow of oxygenthrough the paper to the burningtobacco. They slow down therate at which the cigarette burnsas the lit end crosses over. Theyare more likely to self-extinguish.These speed bumps are made of alayer of alginate on the cigarettepaper – the same alginate used forthe molecular caviar.

A simple application of the FTIRspectrophotometer in combina-tion with the single reflectionunit demonstrates the differencesof the materials. The single mate-rial spectra and the spectrum ofthe final paper layer are shownstep by step. All significant com-ponents such as cellulose in thepaper, sodium carbonate and algi-nate have broad signals in theinfrared spectrum. All have incommon the polysaccharide char-acteristics of the 6 ring structurein conjunction with -C-O-C- andthe -OH bonded groups.

The paper spectrum shows signif-icant signals for the whitener of

the sodium carbonate. These are a sharp signal at 700, at 900 and a broad signal at 1400 cm-1. This is reasonable based on the sub-structure of the carbonate groupwhich generates two -CO- andone -C = O bonding constellationand also distribution of electronsover this bonding system, re-sulting in the broad signal at 1400 cm-1. In the literature, thesymmetric valence vibration �sy(-COO-) is calculated as 1440 -1360 cm-1.

The spectrum of alginate onpaper shows a mixture of alginateand paper. It is possible to differ-entiate between both spectra.Even though both materials arebased on polysaccharide, theyhave differences in their molecu-lar structure which are visible inthe infrared spectrum. The signalat 1600 cm-1 is the -OH bondingin the huge molecules. Whencomparing the structure of thepolysaccharides the differentpositioning at the ring systemsgenerates the additional signal incomparison to the cellulose.

rettesgarette paper

Figure 5: Infrared spectrum of a white cigarette paper measured with single reflection

technique

Figure 6: Infrared spectrum of cigarette paper with a thin layer of Sodium-Alginate,

surface analysis with single reflection ATR technique

Figure 7: Infrared Spectrum of sodium alginate powder measured with single reflection

technique

-0.01

0

0.01

0.02

0.03

0.04

0.05

0.06

0.07

0.08

0.09

0.10

0.11

0.12

0.13

0.14

0.15

0.16

0.17

0.18

0.19

0.20

0.21

0.22

0.25

Cigarette paper, white DuraSampleIIR

4400 4000 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 400

1/cm

Abs

.

-0.0225

-0.0075

0.0075

0.0225

0.0375

0.0525

0.0675

0.0825

0.0975

0.1125

0.1275

0.1425

0.1575

0.1725

0.1875

0.2025

0.2175

0.2325

0.2475

0.2625

0.2775

0.2925

0.3075

0.3225

0.3375

0.3525

Cigarette paper, white plus algenate Na DuraSampleIIR

4400 4000 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 400

1/cm

Abs

.

-0.01

0

0.01

0.02

0.03

0.04

0.05

0.06

0.07

0.08

0.09

0.10

0.11

0.12

0.13

0.14

0.15

0.16

0.17

0.18

0.19

0.20

0.21

0.22

0.23

Sodium alginate, Sosa DuraSampleIIR

4400 4000 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 400

1/cm

Abs

.


4

Bread packagingFTIR measurement of polymers in the food industry

Renewable energy andrenewable raw materialsare the keywords most

often used in relation to fuels.Renewable raw materials are,however, also an important issuefor packaging materials. Instead ofpackaging chips made ofpolyesteramide or polycaprolac-tone (which are derived from fos-sil raw materials), alternativepolymer chips from renewableraw materials can be used, forinstance of vegetable origin.Examples of target polymers arestarch, cellulose and lignin [1].

Packaging chips based on renew-able materials have more in com-mon with bread and baked goodsthan with plastics. In many adelivered package, one can findleaflets pointing out that the pack-aging chips are edible. Thesepackaging chips consist of starch,or in the case of 'flupis®', ofpaper foam manufactured fromwaste paper and starch. Bothtypes of chips can be disposed of

used such as flour, sugar, yeast,water and other components eachexhibit their own distinct infraredspectrum. When these spectra areoverlaid, assignment of individualsignals is difficult. Starch, cellu-lose and sugar have similar spec-tra as they are all polysaccha-rides. Water is also a difficultmaterial, as it exhibits a highlyintense spectrum. For compari-son, dry baked goods such ascrisp bread and baking wafers(wheat flour and starch) requiringvery little water, were usedaccordingly.

Correlation between spec-trum and material hardness

As seen in figure 6, the spectra ofthe filling materials in the rangeof 1540 cm-1 show more similari-ty to the starch spectra than thetwo baked goods. Furthermore,differences can be observed in therange of the carbonyl bands at1750 cm-1. In this range, thebaked goods can be distinguished

easily by composting or in organicwaste containers. In the presenceof water, the chips immediatelydisintegrate and form a pulp, simi-lar to that perceived in the mouthwhen biting into baking wafers oredible paper.

Packaging chip or bread

Can these chips be distinguishedfrom bread or baked goods? Forcomparison, a piece of crispbread and a baking wafer havebeen analyzed using FTIR spec-troscopy in combination withsingle reflectance measurements,allowing fast non-destructiveanalysis of these types of materi-als. The infrared radiation appliedpenetrates approx. 2 μm into thesample surface. The interactionbetween the radiation and thematerial provides information onits composition.

The bread mixture is a highlycomplex composition for infraredspectroscopy, since all materials

Figure 2: Infrared spectrum of a baking wafer, measured using a single reflectance unit

Figure 1: Edible packaging material:

a packaging chip made out of starch

-0.0075

Abs

.

1 /cm

4600

00.0075

0.0150.0225

0.0030.0375

0.0450.0525

0.060.0675

0.0750.0825

0.090.0975

0.1050.1125

0.120.1275

0.135

0.15

0.165

0.18

0.195

0.21

0.225

0.1425

0.1575

0.1725

0.1875

0.2025

0.2175

0.23250.24

0.24750.255

0.26250.27

0.27750.285

0.29250.3

4400 4200 4000 3800 3600 3400 3200 3000 2800 2600 2400 2200 2000 1900 1800 1700 1600 1500 1400 1300 1200 1100 1000 900 800 700 500 600 400

5


products are exhibited in the IRspectrum and enable an unequiv-ocal signal assignment.

Conclusion

Using infrared spectroscopy,complex materials can be ana-lyzed directly and non-destruc-tively. Within the shortest possi-ble time, edible filling materialscan be distinguished from con-ventional baked goods or starches(corn starch in this example)using infrared spectroscopy.

from the filling materials as wellas from the starch.

The filling materials containaggregates exhibiting strong sig-nals at 1734 and 1713 cm-1. Thevarious intense spectral signalscan be correlated to the hardnessof the material. Crisp breads, aswell as baking wafers are quitehard compared to the fillingmaterials. Hard materials do notmake good contact with themeasuring window. As expected,the various compositions of the

Literature:

[1] Nachwachsende Biopolymere

als Substitution für Massen-

kunststoffe; K. Wilhelm, K.

Reitinger; Berichte aus Energie-

und Umweltforschung

14/2006; Federal Ministry for

Transport, Innovation and Tech-

nology, Vienna, Austria

Figure 3: Infrared spectrum of a crisp bread Figure 4: Infrared spectrum of flupis®, filling material made of waste paper and starch

Figure 5: Infrared spectrum of an edible filling material, see Figure 1 Figure 6: Zoom in the range of 1,900 to 1,150 cm-1. An additional substance

(cornstarch spectrum, green line) was used for comparison

-0.01

Abs

.1 /cm

4600

00.010.020.030.040.050.060.070.080.09

0.10.110.120.130.140.150.160.170.180.19

0.20.210.220.230.240.250.260.270.280.29

0.3

0.32

0.34

0.36

0.38

0.4

0.42

0.44

0.46

0.48

0.31

0.33

0.35

0.37

0.39

0.41

0.43

0.45

0.47

0.490.5

0.510.520.53

4400 4200 4000 3800 3600 3400 3200 3000 2800 2600 2400 2200 2000 1900 1800 1700 1600 1500 1400 1300 1200 1100 1000 900 800 700 600 500 400

-0.005

Abs

.

1 /cm

4500

00.005

0.010.015

0.020.025

0.030.035

0.040.045

0.050.055

0.060.065

0.070.075

0.080.085

0.090.095

0.10.105

0.110.115

0.120.125

0.130.135

0.140.145

0.150.155

0.160.165

0.170.175

0.180.185

0.190.195

0.20.205

0.21

4300 4100 3900 3700 3500 3300 3100 2900 2700 2500 2300 2100 1950 1850 1750 1650 1550 1450 1350 1250 1150 1050900 700

950 850 750 650

0

Abs

.

1 /cm

4600

0.00750.015

0.02250.03

0.03750.045

0.05250.06

0.06750.075

0.08250.09

0.09750.105

0.11250.12

0.12750.135

0.14250.15

0.15750.165

0.17250.18

0.18750.195

0.20250.21

0.21750.225

0.23250.24

0.24750.255

0.26250.27

0.27750.285

0.29250.3

0.30750.315

0.3225

4400 4200 4000 3800 3600 3400 3200 3000 2800 2600 2400 2200 2000 1900 1800 1700 1600 1500 1400 1300 1200 1100 1000 900 800 700 500 600 400

0

Abs

.

1 /cm

1750

0.02

0.04

0.06

0.08

0.1

0.12

0.14

0.16

1700 1650 1600 1550 1500 1450 1400 1350 1300 1250 1200

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14

serpentine and amphiboles groups(Table 1).

Figure 1 shows the asbestos fibreof Chrysotile in 100 x zoomedoptical microscopy image. Thesilicate microscopic needle struc-ture indicates that pretreatmentshould be carried out with greatcare.

Analysis techniques

Various asbestos analysis tech-niques are available, such asmicroscopic methods, with opti-cal or electronic techniquesincluding PCM (Phase ContrastMicroscopy), PLM (polarizedlight microscopy) and TEM(transmission electron micros-copy). Another common tech-nique is the X-ray diffractometrymethod (XRD) which is able todetect elements other than crys-talline structures.

Another known method is IRspectrometry represented byFTIR, Fourier Transformed InfraRed spectroscopy. This techniquedetects and identifies asbestosfibres independently of subjectiveevaluations such as the visualmethod.

Solid analysis

aerosols. Asbestos may also befound in natural water (springwater flowing through asbestosrocks) or in drinking water col-lected in asbestos cement ducts.

Former uses of asbestos were:• building materials (in cement

and concrete production)• profile sheeting (Eternit)• linoleum tile flooring• heating pipe insulation coatings• oven door rope seals and

protective clothing for fire-fighters

• filtration purposes• automotive brakes.

Controls must be establishedwhen natural asbestos is present.Part of this application is a proce-dure describing a successful sam-ple treatment in combination withFTIR analysis.

Asbestos minerals and theirchemical formulas

A wide range of naturally occur-ring asbestos minerals have beenused for many technical and com-mercial applications. Asbestosfibres derive essentially from twogroups of silicates which can gen-erate a fibrous crystalline form:

Figure 2a: Untreated sample Figure 2b: Sample after solvent purification

Analysis of mineral fibres (i.e. asbestos) in water,soil and waste material using FTIR spectroscopyEmanuele Canu, Shimadzu Italy, Milano, ItalyZeno Marco Sedin, ARC, Italy

Asbestos is a highly heat-resistant and non-flamm-able natural fibre. It has

been used as a construction material and for many specificapplications. However, since the1970s, it has been proven to be ahuman carcinogen known tocause lung cancer. As a result, theuse of asbestos was restrictedbefore being banned in Austria(1990) and Germany (1993). Since2005 the use of asbestos has beenbanned in the EU.

Asbestos occurs in natural rocksas well as technical materials suchas raw materials used in the man-ufacture of cement. Fine fibrebased asbestos particles can befound in the form of dusts and

Figure 1: 100 x zoomed optical microscope image of Chrysotile

Formula

Mg3Si2O5(OH)4

(Fe,Mg)7Si8O22(OH)2

Ca2Mg5Si8O22(OH)2

Mg7Si8O22(OH)2

Ca2(Mg,Fe)5Si8O22(OH)2

Na2(Fe++3Fe+++2)Si8O22(OH)2

SerpentineAmphiboles

15


Figure 3: Typical band patterns confirm the asbestos type (Canadian Norm, NEN-Norm)

Chrysotile (white asbestos) Amosyte (brown asbestos)TremoliteAnthophylliteAktinoliteCrokydolite (blue asbestos)

Groups Name

Figure 2c: Sample after acid attack

Table 1: Asbestos materials

In comparison with the micro-scope sample preparation tech-nique, a procedure is shownwhich isolates the fibres andresults in a precise identification.

The suggested procedure forFTIR is:1. dissolve organic solvent

(acetone suitable for mixed

esters membranes) in a glasstest tube;

2. add KBr in the test tube, stirand centrifuge it and removethe supernatant

3. repeat item 2 to eliminate thedissolved polymer

4. dry and grind the solid residue and prepare a 13 mmKBr pellet.

Sample preparation

When approaching samples analy-sis, it is necessary to consider thefollowing:

Elimination of organicinterference via:

1. solvent dissolution;2. chemical degradation (humid-

nitric acid or sulphuric/nitricacid mixture or dry treatment),having a clear possibility ofinteraction of the degradingagent or temperature with thesilicates.

Elimination of inorganic interference via:1. acid dissolution (from acetic to

hydrochloric acid);2. specific weight separation

(by means of heavy-liquid).

This last point relates to samplescontaining components insolublein acids: grinding, sifting, sus-pending the mixture and separat-ing insoluble components bymeans of centrifuging with heavy-liquid. The separation will occurbased on difference of the specificweight among phases (suggested:bromoform CHBr3-D≈2,85-,diiodmethane CH2I2-D≈3,31-; itis possible to obtain differentdensities while diluting the heavy-liquid with 1,1,2-trichloroetheneC2HCl3 or 1,1,2,2-tetrachlor-ethene C2Cl4.

Once the fibres have been sepa-rated, ground in an agate capsulewith a vibrating mill and mixedwith 200 mg of IR grade KBr(around 1 % of pellet weight), aKBr pellet is formed.

in the sample may no longer beapparent as it was found whenusing dry ashing.

Separation with heavy-liquidshows the benefit of the treat-ment: see result of the top rightspectra in comparison with theSRM Chrysotile (lower right).Chrysotile fibre has been isolatedand identified. The related peaksat 3690, 1080, 605 and 440 cm-1

are detected.

Future developments

Whereas inhalation of asbestosfibres is considered to beextremely dangerous for the res-piratory system, medical scienceis currently studying the dangerof ingestion of fibres in the diges-tive tract. D




Analytical conditions

The Shimadzu IRPrestige-21spectrophotometer and KBr-Pel-let holder are required for theanalysis in transmission mode.Identification is carried out usingthe decision table (Figure 3) or areference spectrum as in Figures4A to 4D on Page 16. The refer-ence spectrum is from Chrysotileand prepared from certified refer-ence material. Infrared spectra arepresented in absorbance scale.

Interpreting asbestos IR spectra

A sample, an Eternit based profilesheeting was analysed using FTIRmeasurement technique. Themeasured infrared spectra showthe separation steps between orig-inal material, solvent and acidtreatment and the isolation of thefibres. Chrysotile was positivelyidentified in parts of this sample. The influence of the treatmentmethod used is significant. Whenstronger purification techniquesare used, the presence of asbestos


PRODUCTS

16

“Everything flows” – literally Method development and optimization using the prominence HPLC

The times when nothingchanged in HPLC as anestablished method are

over. After a relatively long phasewithout large change, methodsare now being challenged; exist-ing HPLC methods are beingoptimized while others are beingredeveloped from scratch. Inaddition, faster and simpler sepa-rations are being considered. Or, depending on the particularapplication area, even more com-plex solutions are called for.

All of this within laboratoryenvironments which for manyyears have been working withstandard validated methodsallowing no alteration.

New developments in columntechnology and HPLC hardwarehave stimulated new demands.The same applies to environmen-tal aspects and adaptations ofpossible MS detection in order to attain improved detection sensitivities. With the famous-

Figure 1: Solvent selection or quaternary

gradient switching valve integrated in the pump

4B: Organic acid purification and separation with heavy-liquid

4D: Standard reference material Chrysotile SRM1866a

Figure 4A: Untreated Eternit sample

4C: Mineral acid purification (sulphuric-nitric acid mixture)

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A B

C D

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4

Tracing an allergy with single reflection

Contact allergies are well-known phenomena of thehuman skin surface. Very

often, the source of the allergy isunclear. Most allergy tests in useare for the exclusion of pollens,animal fibres or colors. Colorsare mainly used for daily textiles,e.g. leathers. Tests are also neces-sary on special metals in theworld of jewellery, such as Nick-el with known allergy connec-tions.

What is a contact allergy?

The allergy symptom startsthrough first contact of the aller-gen with the skin or mucous

Allergenic mouse pad anFTIR spectroscopy

membrane. This first contact does not cause any skin irritation,but it causes a sensitivity in thebody. With the next contact tothe material the skin activates itsimmune system an allergic reac-tion can soon be observed. As aninflammation it fights the aller-gen, but only in the area wherethe allergen contacted the skin.

In this application a contact aller-gy is described which was causedby two objects. Standard allergytests had not been helpful. Thetwo materials of interest were amouse pad and a leather wrist-band. What do both have in com-mon causing the skin reaction?

For the analysis, an instrumentdedicated to surface analysis isneeded, e.g. a single reflection

diamond based ATR (AttenuatedTotal Reflectance) unit integratedinto a FTIR sample compartment.

The test person showed differentreactions when applying twoleather wristbands. In one case,the skin appeared normal but inthe other case, an eczema wassoon apparent (Figure 1).

Single reflection ATR enabled theanalysis of the surface of differentleather-based materials as well as the mouse pad. Both of theleather samples were in contactwith the skin – one was a colored,smooth leather, and the second anundyed leather with the usualrough surface. The ATR measure-ment technique with a KRS-5crystal allows beam penetrationof approx. 2 μm into the samplesurface.

The analysis was done with FTIR spectroscopy and an ATRaccessory enabling a non-destruc-tive sample analysis. The sampleis placed on a measurement window and does not need anychemical pretreatment. Figure 2shows both spectra.

The main difference in figure 2 isthe signal at 1,727 cm-1. The sig-nal is a –C = O signal belongingto the acid based –COOH groupand may be present from differ-ent sources of molecular structureor substances of this class.

The spectrum was compared to a skin spectrum to prove thathuman skin traces at the surfacedid not influence the analysis ofthe leather band.

The skin spectrum shows no sig-nal peak at around 1,727 cm-1.The same infrared analysis wasdone with the surface of themouse pad that caused the skintrouble.

Figure 1: View of a contact allergy seen on an arm – a smooth skin reaction caused by a brown coloured leather wristband.

The zoom shows details of a typical eczema resulting from a contact allergy.

5


d leather wristlet

A search of the infrared spectrafrom the mouse pad within an IRlibrary resulted in positive identi-fication as Polymethylmethacry-late (PMMA). It has a good over-lapping for the signal at 1,727 cm-1.Acrylate is known as a materialwhich can cause contact allergy.

Conclusion

It can be helpful to do a surfaceanalysis of the materials suspect-ed of being the source of the con-

tact allergy. The FTIR techniquecombined with ATR, particularlythe single reflection methodenables control of surface layersfrom thicknesses between 0.6 to 2 μm. These measurements helpto point the direction of the nextsearch for the allergy. Whether itis the polymer or a polymer com-ponent needs to be researched inmore depth with additional spe-cific allergy tests.

Instrumentation• IRAffinity-1

• DuraSamplIR, diamond based

single reflection unit

• IRsolution and libraries



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Figure 2: FTIR leather spectra. Blue is untreated leather while red is the treated

leather surface

Figure 3: FTIR spectrum from the skin of a finger with single reflection accessory

Figure 4: FTIR single reflectance spectrum from the surface of a mouse pad Figure 5: Comparison of Library spectrum (blue line) and the mouse pad surface

(black line)

0.010.020.030.040.050.060.070.080.090.1

0.110.120.130.140.150.160.170.180.19

0.20.210.220.230.240.250.260.270.280.29

0.30.310.320.330.340.350.360.370.380.39

4600 4400 4200 4000 3800 3600 3400 3200 3000 2800 2600 2400 2200 2000 1900 1800 1700 1600 1500 1400 1300 1200 1100 1000 900 800 700 600 500 400

1/cm

Abs

0.010

0.020.030.040.050.060.070.080.09

0.10.110.120.130.140.150.160.170.180.19

0.20.210.220.230.240.250.260.270.280.29

0.30.310.320.330.340.350.360.370.380.39

0.40.410.42

4600 4400 4200 4000 3800 3600 3400 3200 3000 2800 2600 2400 2200 2000 1900 1800 1700 1600 1500 1400 1300 1200 1100 1000 900 800 700 600 500 400

1/cm

Abs

0.010

0.020.030.040.050.060.070.080.090.1

0.110.120.130.140.150.160.170.180.19

0.20.210.220.230.240.250.260.270.280.29

0.30.310.320.330.34

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1/cm

Abs

-0.05-0.025

00.025

0.050.075

0.10.125

0.150.175

0.20.225

0.250.275

0.30.325

0.350.375

0.40.425

0.450.475

0.50.525

0.55

0.6

0.65

0.7

0.75

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0.95

1.0

1.05

0.575

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0.675

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1/cm

Abs

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