Over the last few months I've become a great enthusiast for the diamond ATR accessory. Even, I a Raman man can be trusted to get decent mid infrared spectra. I asked David Coombs of Graseby Specac and the folks at Perkin Elmer's Beaconsfield laboratories to write pieces telling us about diamond ATR, what it is, how it works and giving examples of applications. As it turns out the contribution from Sharon Cooke, Cathy Deeley and Mark Billingham puts forward a rather challenging idea - why not use the quantitative approach in mid i.r. and Raman spectroscopy normally associated with the new infrared? Don't bother about the meaning of the spectra or the way that the data is recorded but rather identify spectral trends in series of calibrated samples and use these to quantitatively analyse unknowns. It’s a challenging idea that exploits the convenience of diamond ATR. I sense their approach is far from developed but it is challenging and of great potential value in industrial and Q.C. labs.

Janet Tyas of Kodak gave a wonderful talk at last Autumn's P.E. Users meeting on her experience in running a routine ir/Raman/nir lab in a busy R&D Department. I recommend her paper to you.

Hans Betterman and his colleagues have recently demonstrated how, with careful application of good optical methods, Raman spectra, feint though they may be can be used to make measurements on low concentration solutions. So we asked Hans to produce a paper for us. Their sensitivity is really very impressive and as they point out lends to the possibility if using Raman for the analysis of components from chromatographic separators. One amusing point, when I read their paper I complained that the optical diagram was hard to follow because it was horribly out of scale. After all, one lens was shown as if it was enormous. Checking the manuscript again revealed that said lens was 47cm in diameter - obviously a typo - who had ever heard of lenses half a metre in diameter - the author must have meant 47mm. WRONG! - the figure is indeed to scale and these folks do use enormous optics! Read it - very interesting piece of work and it certainly refutes the idea that Raman will not work on traces.

Contributed articles - delighted to report that these are coming in at a steadily increasing rate and the coverage is very wide indeed. Everyone interested in vibrational spectroscopy should find the papers on offer this time really interesting. David Schmierer and colleagues tells us about polymorphism in drugs - a real problem in the pharmaceutical industry.

Liquid crystals are important to us all (and not only in LC displays) so a description from Jaap Leyte, of how mid infrared can be used to follow changes that occur when you apply an electric field (ah ah - as you do in a LC display) is timely.

Kristine Moore has contributed to Ed I, Vol I in the Dear Readers section, so I asked her for an account of the fascinating work she does on mediaeval paintings and documents.

And finally Derek Gardiner has responded to my request and tells us about the effect of stress on silicon slices. A wonderful varied bunch.

Patrick Hendra
Editor


2. Assistant Editorial

Adding to my seemingly endless list of things to do, Patrick has asked me to write a few words on the production side of IJVS. As he has already mentioned I am now responsible for the production of the Journal as well as administration, so my first request is PLEASE go easy on me. If there are any broken links or problems with this Edition, please do not hesitate to contact me, but take note that this is my first one!

The purpose of my section will be to advise and update readers and would-be authors how to get the best from IJVS, as we endeavour to provide a quality publication for you. Until now we've mentioned that the Journal is becoming increasingly popular, but possibly not clarified what this entails. Software built into the website, tells us how many people have visited the site, who subscribes to our mailing list and also readers comments - which we greatly appreciate. It was a pleasant surprise to find out that our last Edition (Ed I, Vol II) since it was issued on the Web in May had received nearly 900 hits! Our subscription list is around the 500 mark and steadily increasing at a rate of about 10 readers each week. It is all very encouraging.

As everyone will realise - IJVS is in colour. We invested in a colour printer a few months ago here in the IJVS office and were pleasantly surprised when printing out the last edition, to see it in colour. Relatively few readers use colour printers but if you do, the hard copy looks far better. One particular problem solved by the use of colour is that multiple spectra are easier to follow when overlapping occurs. More contributors are suppling some very pretty pictures, this Edition is no exception, so try to get access to a colour printer, its worth it.

I'm not going to say much more, other than a BIG thank you to all the contributors to this Edition and apologies for the lateness of this publication. I realise that we're more than half way through this year, with only two editions to show for it. The remaining four editions planned for Volume II will be produced a lot quicker and closer together.

Louise Martin
Assistant Editor

Feature Article


3. The use of Diamond as an ATR material

Dave Coombs
Graseby  Specac,
River House, 97 Cray Avenue,
St Mary Cray, Orpington,
Kent, BR5 4HE
United Kingdom

Telephone: +44 (0) 1689  873134
Fax: +44 (0) 1689  878527
E-mail: coombsd@compuserve.com

Introduction

 
The development of Diamond as an ATR material has opened up a number of sampling opportunities in the mid infrared. Diamond has a number of properties important for the infrared spectroscopist. The high refractive index ensures that for incidence angles of 45 degrees, a typical infrared penetration of around 2 microns is achieved. Consequently, single reflection measurements are capable of yielding good quality spectra under conditions of good optical contact with the ATR crystal.

The extraordinarily stable covalent bonding structure of diamond accounts for its hardness and physical strength .The principal benefit is that ATR sampling for abrasive or chemically hostile samples is now routine. The large band gap between the valence and conduction electrons in diamond means that radiation of a very broad wavelength range is able to pass through almost unhindered. Diamond therefore is able to transmit from the UV-Visible to the far infrared. There are mid infrared lattice vibrations at around 2000 cm-1 however careful optical design and short path lengths through the diamond ensure that ATR accessories perform well in this region of the spectrum and that the full transmission capabilities of diamond are properly utilised.

Finally, the high frequency vibrations of the carbon atoms in the lattice impart yet another useful property open to exploitation by the optical engineer. Heated ATR experiments are straightforward because of the high thermal conductivity of diamond. In fact diamond is five times more efficient at conducting heat than even copper which ensures the place of this unique material in the infrared hall of fame.

In a simple transmission experiment, the light beam enters the sample normal to its surface and passes through. If we change the incidence angle to one other than zero degrees, the sample would reflect much of the incident radiation depending upon its actual reflectivity. This forms the basis for specular reflectance measurements. If an IR transparent crystal such as diamond is placed behind and in good optical contact with the sample, light passing into the diamond can be made to emerge on the opposite side having made a reflection at the diamond-sample interface. In fact the emerging light contains important absorbance information about the sample. There are certain geometric requirements for this and importantly the refractive index of the crystal must exceed that of the sample.

Figure 1. The ATR Experiment

Regarding the optical and geometrical requirements for this process, we need to consider the differences in the refractive indices between the two materials and the angle of incidence of the radiation. At the first contact between the radiation and the sample, a fraction will be reflected and a fraction transmitted. When light passes through two media in intimate optical contact with each other and having different refractive indices, the path of the light will be distorted depending on the incidence angle. The incident light must therefore be below some critical angle in order to set up the process. In fact the situation is a little more complex than this because the light is polarised perpendicular and parallel to its direction of travel and the reflectivity is different for each of these polarisation's and also varies with incidence angle. # ATR is a surface sensitive technique. Considering the refractive index of diamond and a typical organic polymer to be 2.4 and 1.5 respectively, the top two microns or so of the surface will be sampled at 45 ŗ incidence. Finally, this sampling depth changes in two fundamental ways. It decreases as the refractive index of the crystal increases and it also decreases as the incidence angle is increased. Also explained in IJVS Vol I, 438-444 (1997) - Editor.

# Editor's Note: This property is discussed further in the next article.

Some experimental aspects of choosing
diamond as a good ATR crystal.

A key experimental requirement is that a smooth flat scratch free sampling surface is maintained in order to maximise both contact and signal to noise ratio performance for the analysis. A uniform pressure distribution is also required. This is more easily achieved using a robust, well supported small surface area diamond. Reproducibility is also easier than with solids sampling using the bulkier classical 6 reflection ZnSe type ATR units. These larger ZnSe units also have a more open architecture with a wide ray acceptance angle cone. Diamond has a smaller acceptance angle cone and requires good optical design in order to avoid working close to the critical angle whilst enjoying a compact beam condensing arrangement in the accessory. The high load bearing capacity of diamond allows for (near) perfect optical contact between the sample and diamond and results in good spectra of even the most intractable of sample types.

Many of the ideal design characteristics for a diamond ATR unit are incorporated into the ‘ Golden Gate’ single reflection diamond ATR accessory manufactured by Specac, Orpington, Kent, UK. A type IIa industrial grade single diamond crystal is high temperature metal bonded into a Tungsten carbide support disc at 1000ŗ C (Figure 2). This ensures the most durable robust and permanent mounting arrangement possible and has no dependence on adhesive bonding which could otherwise result in the removal of the diamond when cleaning with certain solvents.

Figure 2. Diamond in it's mount

Samples are forced into almost perfect optical contact with the diamond using a sapphire anvil assembly mounted overhead on a unique swing bridge. Reproducible loads of 3 kbar can be applied using a special torque wrench supplied with the unit. The anvil is fully interchangeable with a series of alternative special anvils. These include a stainless steel anvil for polymer pellets, grooved anvils for analysing polymer coatings on wires, and an anvil for studying air sensitive or reactive compounds.

Figure 3. Golden Gate ATR Diamond accessory

In this application, the top plate of the accessory may be removed to a glove box or controlled atmosphere and the sample loaded into position and sealed using a special anvil in order to protect it from the external environment. The top plate is subsequently replaced ready for the measurement. A micro reaction/flow cell anvil is also fitted in a similar manner and can be used to study low volume (28 microlitres) liquid flows or static injections up to 1000 psi.

The diamond ATR top plate is mounted onto a fully enclosed beam condensing optics box used to condense the IR beam by a factor of 4X. Typically this results in a sensitive sampling area of approximately 1mm in the centre of the diamond crystal. The optics are robust and maintain their stability of alignment and can also be purged with dry air or nitrogen. The beam condensation is achieved using the more optically efficient lens option in combination with mirrors. The lenses are made of ZnSe, which transmits, to around 650cm-1. Alternatively KRS-5 lenses can be used for transmitting to longer wavelengths (300 cm-1). The small sample hot spot makes the accessory ideal for microsampling.

Figure 4. Schematic Diamond ATR

Single reflection diamond ATR has been successfully applied in a number of micro-sampling applications. Particularly, single polymer fibres as small as 20 microns have been analysed with the technique and other forensic type applications such as paint fragments and particulates can yield good quality spectra without the need for cumbersome sample preparation. A benefit of this approach is that qualitative analyses can be quickly performed which can be used as a screening technique before engaging in the more expensive and time consuming use of infrared microscopy. A combination of the two makes a real contribution towards cost effective and efficient analysis of small samples.

Single pellets, powders, coated wires, intractable or opaque solids, corrosive liquids, curing polymers, and air sensitive compounds can now be analysed with one multi purpose accessory.

Many reactions and processes occur at high temperatures. The heated version Diamond ATR top plate allows for all the benefits of the standard unit up to 200 C.

The uniquely high thermal conductivity of diamond in combination with the low thermal mass top plate, ensure that the high power heaters built in to the plate in close proximity to the diamond, leads to both rapid and efficient heating of the diamond. The heated unit is used with an electronic temperature controller with digital readout to 1 C. Some applications for this unit have been polymerisation reactions, cooking processes, degradation/decomposition, and phase transitions. A special reaction cell version of the diamond ATR can be configured for the study of low volume (24ml) reaction studies and can incorporate flow, mixing, options at combinations up to 200 C at 3000psi.This unit has proved ideal for small scale optimization of process parameters, for the studies of highly acidic and caustic solutions and for the study of slurries with abrasive particulates in suspension.

4. Some applications examples for
single reflection Diamond ATR

We have included some typical applications for this technology to illustrate the versatility of using just a single reflection from a diamond ATR crystal. Remember these spectra all correspond to a sample penetration of just two microns. This shows the awesome capability of using Diamond, as you may never have thought possible before!

Liquids have never posed a problem for the ATR technique except when they contain acids that would attack conventional ATR crystals such as ZnSe. The data shown in Figure 5 was recorded at room temperature using a sample volume of less than 30 microlitres.

Understanding the process of cement curing

You may well have seen spectra of paint drying in the past but here is a novel variation on a theme. Cement is mineralogically complex and so too are the reactions involved in the cure process. Even more demanding is understanding how cement minerals decompose under conditions of high temperature and pressure. This is especially important in the oil industry. Spectra shown in Figure 6 were recorded at high temperature and illustrate the decomposition of the mineral Ettringite in the presence of water under sealed conditions. Classically, X-ray diffraction techniques would have been used in crystallographic studies of these systems. Infrared however has an important complementary role in yielding information on how water is involved - a study not possible before.

Figure 6. The FTIR-ATR spectrum of ettringite
showing decomposition at 114°C

What happens when bread stales?

There are a number of schools of thought on this one! Spectra of fresh bread show distinct features attributed to water, fat, starch and protein (Figure 7). When the bread stales there is good evidence for a reduction in the water content and it would appear that a subtle difference in the starch region of the spectrum may account for a change in the crystal structure of the starch with its corresponding effect on texture!(Figure 8)

>

Figure 7. Fresh Bread by Diamond single reflection ATR

Figure 8. ATR Infrared spectra of fresh and stale bread

Surely not…. it cannot be that easy to
obtain an infrared spectrum of oleum?

Sulphuric acid is a strong dibasic acid with ample capability of digesting standard ATR materials. The oleums are sulphuric acid/sulphur trioxide cocktails with enormous industrial importance. This includes the manufacture of important nitrogenous chemicals such as nitro-cellulose and various dyestuffs. They are also used in the purification processes for petroleum products. The spectrum shown was recorded using the most minute pool of liquid imaginable…and why not!(Figure 9).

Figure 9. ATR Infrared spectra of oleum H₂SO₄+SO₃

Are you sure that it was my car that
you claim ran into you…….?

These spectra shown in Figure 10 were recorded in a real forensic crime laboratory. They illustrate the data obtained from a real vehicle interaction in a case where the suspect failed to stop. These fragments were approximately 0.5 mm in diameter and the data was recorded using a DTGS detector. Undoubtedly a big success for three minutes analysis time.

Figure 10.

We will be pleased to show also, other examples of data we have recorded using single reflection diamond ATR .If you have a particular interest not mentioned in this short applications brief, then please contact me via one of the routes below and I will be pleased to help.

Editor's Note:
So, there you have it. Until January I had never used a diamond ATR accessory. I then became involved in a project to examine a very wide range of polymers and I was staggered at the convenience. I found that Davids' claim above is real- amost anything, a powder, lump or film placed on the horizontal diamond surface and pressurised, gave an excellent spectra. The ease and speed of sampling is truly impressive. Of course, most of the comments David makes apply equally to systems from other competing manufacturers.

Feature Article


5. A Technological Approach to
Quantitative Analysis using
Vibrational Spectroscopy
in the Polymer Industry

Sharon Cooke, Catherine Deeley and Mark Billingham
Perkin-Elmer Limited,
Post Office Lane,
Beaconsfield, Bucks.
HP9 1QA
United Kingdom

Telephone: +44 (0) 1494 676161

Mid infrared spectroscopy has a long history in polymer science. At the fundamental level, the spectra are well understood whilst those of unknowns are readily identified using `fingerprinting’. Vibration spectra of polymers are sensitive to polymer morphology (crystallinity and/or orientation) and such information can be obtained using mid infrared spectroscopy. Quantitative analysis is also possible using mid infrared. Several standard procedures are available for these analyses but they are often time consuming and expensive because they invariably require, as part of the analysis, the production of reproducible film samples. In fact, one of the main problems when using mid infrared spectroscopy for polymer analysis has been the difficult and time consuming sample preparation. Traditionally, the standard procedure for MIR transmission sampling is to produce a film ~ 30 mm thick either by microtoming or by the use of a hot press and a film making accessory. Unfortunately, microtoming is highly skilled, time consuming and expensive whereas melting and film making can frequently ruin the sample if, for example, the purpose of the analysis is to measure orientation or crystallinity.

Raman spectroscopy is an alternative method of investigating the vibrational behaviour of polymers but is far less used than mid infrared spectroscopy. The reasons for this are largely historical but with the development of Fourier Transform (FT) Raman spectroscopy spectra can be recorded very rapidly directly on samples with little or no sample preparation. Samples as different as a fibre, a lump or even a screw driver handle can be studied almost instantly. Unfortunately, the main weakness of Raman spectroscopy is its poor repeatability in quantitative analysis and perhaps it is this that restricts its acceptability to industrial analysts. This is unfortunate because the spectra are of very high quality, constitute an excellent fingerprint and are often well understood in the literature. Morphological features of a specimen are clearly indicated in the Raman spectrum as they are in the mid infrared.

In contrast, near infrared spectra are very different from those obtained using mid infrared and Raman methods. The spectra arise from overtones of the fundamental frequencies and so the spectra consist of many overlapping bands. This results in very complex spectra which tend to be unresolved and almost featureless and to the eye, often constitute no useful fingerprint. However, it must be accepted that in the quantitative measurement of complex mixtures (for example in the food or agricultural industries or more recently in the pharmaceutical field) near infrared spectroscopy has proved to be a valuable tool and its application is expanding very rapidly. The NIR analysis of such complex spectra typically relies on a variable which follows the property change of interest and simply relates the two in some simple manner, say by a linear or simple quadratic relationship. The advantage of NIR spectroscopy over other analytical techniques is cost per sample – sample preparation is often non existent and the complete analysis can be performed very rapidly. The data analysis is mathematically sophisticated but the analysis is routine, reliable, rapid and if carefully designed, almost fool-proof. Near infrared spectroscopy is normally carried out in transmission or reflection. Solid samples are presented as films, lumps, sheets or powders in their original containers with little or no sample preparation. Mid infrared and Raman spectroscopy have been seen, perhaps unfairly, to lag behind near infrared with respect to sample preparation and ease of analysis but, in appropriate experimental circumstances, these techniques can rival near infrared in speed and convenience. The aim of this article is to demonstrate how the different technological approaches mentioned above can be applied to quantitative analysis in the polymer industry.

Sampling alternatives for mid infrared spectroscopy do exist; one can make reflection measurements either specularly or by diffuse reflection. In both, the surface condition and presence of fillers are highly significant and hence tend not to be reproducible. Also, the spectra recorded may need mathematical processing to make them recognisable. Attenuated Total Reflectance (ATR) is a possibility but is initially unattractive because polymers are often hard and rigid and hence it is difficult to produce the essential optical contact between the sample and the ATR crystal. Recently, this problem has been reduced thanks to the development of single reflection diamond ATR accessories. Hence, hard or soft specimens are squeezed against one surface of the diamond prism (~1mm sq.). These accessories are very easy to use and sampling takes only seconds.

Samples of ethylene and vinyl acetate copolymers with a wide range of concentrations, were supplied by ICI Wilton Research Centre in an attempt to analyse the vinyl acetate content by mid infrared spectroscopy. Initially the mid infrared spectra were recorded with no sample preparation. All spectra were recorded on a Perkin Elmer Spectrum GX fitted with the Golden Gate diamond ATR accessory. Data was collected at 4 cm^-1 resolution over a period of one minute scanning time. Each polymer was squeezed onto the diamond ATR to give enough pressure to ensure good spectra were obtained. Spectra of the EVA calibration samples are illustrated in Figure 1 where the different levels of vinyl acetate can clearly be seen. The spectra obtained were mathematically analysed using a partial least squares algorithm using Spectrum Quant+ software. Further EVA samples were supplied by the University of Southampton to use as an independent validation set. The data was referenced by NMR spectroscopy. Following the PLS analysis a standard error of prediction of 0.5 was obtained using 2 PLS factors and X number of standards in the calibration. It is believed that the SEP value would improve with the addition of more standards. A regression summary is shown in Table 1 and the Estimated vs Specified plot is shown in Figure 2. The methods built were tested using independent validation samples supplied by the University of Southampton. The validation spectra were recorded and a typical prediction report is shown in Table 2. Although the predicted value of vinyl acetate is very good at 8.74% compared to the referenced value of 9%, the statistics in the report indicate that the residual ratio is quite large. This is most likely due to the difference in sample form, pellets as opposed to slabs, which were not incorporated into the calibration.

Figure 1. Mid infrared spectra of EVA polymers




Figure 2. Estimated vs Specified plot for EVA by mid infrared

Table 1. Regression summary for EVA

Table 2. Prediction Report for EVA

Following the success of the EVA calibration using mid infrared spectroscopy it was decided to repeat the analysis using both Raman and NIR spectroscopy. Raman measurements were recorded on a Spectrum GX Raman and all near infrared spectra were recorded on a Perkin Elmer Spectrum IdentiCheck FT-NIR spectrometer fitted with the IdentiCheck Reflectance accessory (ICRA). Raman measurements were recorded at 4 cm^-1 resolution whilst the NIR spectra were recorded at 8cm^-1 resolution. The regression summaries (Table 3 and Table 4) and the Estimated vs Specified plots for both the Raman and NIR analyses are shown in Figure 3 and Figure 4. Although both techniques produce very good calibrations and would be ideal to use for the analysis of ethylene vinyl acetate polymers it can be seen that mid infrared produced the most superior results based on the samples used.

Figure 3. Estimated vs Specified plot for EVA by Raman

Table 3. Regression summary for EVA by Raman



Figure 4. Estimated vs Specified plot for EVA by NIR

Table 4. Regression summary for EVA by NIR

Composition in natural rubber-gutta purcha mixtures

MIR, Raman and NIR techniques were used to analyse a second set of polymer samples supplied by the University of Southampton. The mixture of naturally occuring cis and trans polyisoprene samples that were supplied was rather unusual. Trans polyisoprene - gutta percha was historically quite important because like natural rubber, it is biologically derived, but these days is rarely encountered. The set of samples are hence laboratory oddities, but they do provide a series similar to ones encountered in the commercial world.  Raman spectroscopy was the most obvious choice for this analysis because the V_c=c Raman band in unsaturated compounds is invariably strong and its frequency varies with isomer type. The main problem arises because both the cis and trans polyisoprene are tertiary olefines and hence the c=c stretching frequency in each case overlaps. It was for this reason that we decided to look at the mid infrared spectra. The spectra were recorded using as light a pressure between the sample and the diamond as possible and examples of the calibration spectra are illustrated in
Figure 5. A quantitative model was built, again using PLS. The Estimated vs Specified plot is shown in Figure 6 and the regression summary can be seen in Table 5. If excessive pressure is used the spectra can change dramatically. For example, the undiluted natural rubber specimen can crystallise. Keeping rubber for extended periods at around 0° C or stretching its crosslinked form to extension ratios beyond ~400% both induce crystallisation and this morphological change has been monitored in the past by both infrared transmission and F-T Raman methods. Figure 7 shows the crystallisation induced by the application of pressure.

Figure 5. Calibration spectra of Natural Rubber samples by MIR

Table 5. Regression summary for Natural rubber by MIR



Figure 6. Estimated vs Specified plot for Natural Rubber
by mid infrared

Table 6. Regression summary for Natural rubber by NIR


Figure 7. Natural rubber in Diamond ATR Pressure Sensitive Morphology



Figure 8. Estimated vs Specified plot for Natural Rubber by NIR

NIR spectra were also recorded on the same rubber samples to obtain a very good correlation between the spectra and the cis-trans ratio. The results from this analysis, shown in Figure 8 and Table 6, can be compared with those obtained using ATR (Figure 6, Table 5). Raman spectra were not recorded on the natural rubber samples.

Crystallinity in Polyether ether ketone (PEEK)

Polyether ether ether ketone is a high temperature thermoplastic which is widely used in the Aero Industry. The material can easily be made in the form of a glass and with appropriate heat treatment will crystallise up to about 50% degree of crystallinity. The polyether ketone samples supplied by ICI were used in an attempt to determine the crystallinity of such samples. The data set comprised of well referenced samples that had previously been analysed by X-Ray diffraction. Secondary measurements of crystallinity have been reported in the literature based on both mir and Raman measurements. However, a serious problem arises when Raman spectroscopy is used to determine crystallinity since degradation often takes place during heat treatment and even with near infrared laser excitation fluorescence is a problem in Raman spectroscopy. However, when these samples were scanned using mir spectroscopy with the diamond ATR accessory the problem is overcome and excellent spectra are produced. Examples of the spectra and the calibration plot are given Figure 9 and Figure 10 and a regression summary is shown in Table 7. It is clear that an excellent correlation can be generated. NIR spectra were generated but unfortunately, an acceptable calibration could not be obtained with the samples supplied and so MIR is the preferred technology in this case.

Figure 9. Extremes of calibration spectra for PEEK samples - MIR



Figure 10. Estimated vs Specified plot for PEEK samples by MIR

Table 7. Regression summary for PEEK samples by MIR

Young's Modulus in oriented polypropylene
 

In principle one can relate any variable to a chosen spectroscopic change from sample to sample. The variable does not have to be composition or molecular. Highly oriented polymers (materials that have been irreversibly extended) show high levels of stiffness, a property exploited in the textile industry in tyre cords and packaging. These industries need to monitor the elongation and often Young's modulus of the materials they produce. This is done by gripping a sample, applying a load and measuring the deflection on a tensile testing machine. Gripping is problematic, the reproducibility is poor, special samples have to be made and it is normal to make several measurements and average - all very expensive and slow.

It is known that when elongated, molecular orientation occurs in a polymer the modulus increases. The correlation between orientation and modulus is complex, as is the elongation vs orientation. The degree of orientation can be measured spectroscopically but obviously the correlation between any spectroscopic effect and the modulus will be pretty obscure. Orientation in polymer films are measured spectroscopically by recording the mir spectrum in polarised light, orienting the sample with its prepared axis parallel and perpendicular to the electric vector of the radiation. Each band intensity in each spectrum is then compared and a set of dichroic ratios is generated. Since the general direction of the movement vectors of the atoms with respect to the molecular axis in at least some of the vibrational modes is known, the dichroic ratios can be used to deduce the perfection of the orientation. Although not strictly a completely relevant measurement, we show in Figure 11 the effect of aligning a sample parallel and perpendicular to the vector direction and the appearance of dichroism.

Figure 11. Effect of sample orientation in Diamond ATR

Figure 12. Diamond ATR Polarisation and sample orientation convention

An experiment was set up as in Figure 12 using the diamond ATR in polarised light. The chosen orientation is clearly identified in the caption. Samples of oriented polypropylene extended to different degrees and of known modulus were supplied by Dr. Shilin Lu of the University of Southampton. Each sample was examined with its molecular axis parallel to the front of the instrument - (perpendicular) (Figure 13) and then arranged normal to the front of the instrument - (parallel) (Figure 14). The set of spectra in each series were submitted to PLS analyses using Spectrum Quant+ and the results are reported below in Table 8a and Table 8b. Clearly the correlation between the spectral variables and Young's Modulus is outstanding and the spectra are very simple indeed to record.

Figure 13. Polypropylene samples - perpendicular


Figure 14. Polypropylene samples - parallel

Table 8a. Tensile modulus by Diamond ATR.
Polypropylene sample Perpendicular

Table 8b. Tensile modulus by Diamond ATR.
Polypropylene sample Parallel

By adopting the logic universally accepted by near infrared users, one can indeed use simple, rapid and easy-to-use sampling methods in the mid infrared or Raman for both qualitative and quantitative analysis of a wide range of polymers. Their use will significantly reduce the cost of analysis since the experimental data can be acquired from an unknown in a few minutes and the mathematical processing is automatic and almost instantaneous.

Editor's Note: We will cover the mathematical analysis of results - so called chemometrics in a future edition.

Feature Article

Janet Tyas
Analytical Laboratories
Kodak European Research and Development
Kodak Limited
Headstone Drive
Harrow, Middlesex
HA1 4TY
United Kingdom

Telephone: +44 (0) 181 424 5346
Fax: +44 (0) 181 424 3750
Email: 923966N@knotes.kodak.com

Background

Our 10 year old infrared microscope needed replacement. After years of heavy use it had poor stability and needed a lot of care and attention, so it was decided to allow it to retire. Our requirements for a replacement microscope were:

• Reasonable price

• Good performance

• Good stability (robust)

• Easy to use.

It was essential to be able to use our existing data; we have been adding spectra to our user generated libraries since 1983. The main uses of the microscope are problem solving and process understanding; it is a resource for all parts of the company.

We make full use of user generated and commercial libraries in various formats. These libraries contain over 45 000 spectra in total. We needed to be able to use all of these libraries on our new system, with minimum inconvenience. With the amount of data involved, going back many years, re-compiling spectral libraries was not an option. At this time we also started to think about the many dispersive reference spectra we had filed in the laboratory, which on paper were of limited use. We sorted through these reference spectra, selecting those that were of good quality and not duplicated in our electronic libraries. The spectra were sent to Mathshop(1) who did an excellent job of digitising them. A library was then compiled. The instrument supplier we selected for the new infrared microscope, supplied a program to convert our user generated libraries to their search format. Commercial libraries cannot be converted for copyright reasons, but can be searched using another supplied program.

Why Library Searching?

As soon as a member of staff has been trained to obtain spectra they can start to solve problems (IR knowledge not essential). A lengthy period of time spent training new members of staff before they can generate information is not a luxury we can afford. A certain amount of initial training is needed, so that the library user makes the correct inferences from the library search result. An experienced team member is still essential, of course, when more complex problems arise.

We have, perhaps, an unusual approach to building spectral libraries. Virtually all data is entered into our libraries, including mixtures and unknowns. Similar spectra can occur several years apart. On more than one occasion a recurring problem has been detected, even if a full identification of the chemical(s) present has not been possible. The background information of both can be used to help solve the current problem. Also, library searching can suggest the class of compound present, even if the exact identity is not obvious. An example of a library search result is below:

Figure 1. A typical library search report

The top spectrum is the sample spectrum and the other spectra are earlier spectra revealed by the library. Referring to the records of these earlier samples can enable identification of the major components present by even a fairly inexperienced person.

Process Understanding

Figure 2. A blocked filter from a production area

Problem Solving Process

Discussion with client... We need to know:

• Background information to the problem

• Possible causes of the problem

• Contaminants that may be in the system

• WHAT do they need to know in order to solve the problem?

• WHEN do they need the information (usually yesterday!)

Other techniques may provide complementary information, such as:

• Optical Microscopy

• Raman Spectroscopy

• Scanning Electron Microscopy

• Mass Spectrometry

• Atomic Absorption Spectroscopy

Infrared microscopy is a very useful problem solving tool, but other techniques may be appropriate instead or as well, depending on the situation. When deciding on the techniques to use, it is important to focus on the information needed to solve the problem. Unexpected results may result in further techniques being used.

Reference Data

It is important to have reference spectra of all substances that may be expected to be present. We ensure that all relevant reference spectra are in our libraries.

We then obtain spectra of the relevant samples concerned with the problem. We search the spectra against the libraries held on the instrument PC. If that does not reveal a good match we extend the search to other data held locally. If necessary, we then transmit spectra to our world-wide colleagues for their input. Consulting world-wide colleagues introduces a delay because of different time zones. We are looking at ways of sharing data more efficiently.

Further discussion with client

Is the information supplied to us enough to solve the problem?

• Has the cause of the problem been identified?

• Is any further analytical work needed?

We then issue a formal written report. Feedback is very important; additional information can then be included with the library entry. Should a similar situation arise in the future, then this information would be invaluable.

Examples of types of samples we look at:

• Surface deposits on photographic film and paper

• Deposits from solutions used in equipment at photo-finishers

• Inclusions in polymer samples

• The ordinary and the extraordinary!

Typically we are asked to identify unknown substances.

For transmission samples, preparation is everything! Samples must be thin and flat. For preparation we use a good stereo zoom optical microscope - it is important to be able to see clearly what you are doing when you prepare a sample. It is not a good idea to try to use the infrared microscope for preparation, as there is the danger of causing damage to the optics. To manipulate samples, you need a fine pointed probe. We find butterfly mounting pins (!) are ideal, because they can be treated as disposable and have a very fine tip (2).

A roller is needed to flatten samples when using salt windows (we use 13x1mm, barium fluoride). Particularly useful for hard or darkly coloured samples are diamond windows (3). The use of hard diamond windows permits the sample in between to be squashed very thin. (Diamond windows are quite expensive, but never need replacing). Where possible one of the windows is removed before analysis to avoid fringing effects.

Samples such as polymers containing inclusions are prepared by obtaining a microtome cross section (5-10 microns thick).

The microscope system we chose is highly automated and allows us to select on-screen several areas to be analysed. We can then analyse automatically all areas marked (with different size apertures for each sample, if required). The image of the particle(s) can then be recorded (as below).

Figure 3. Several points have been selected for analysis
and been analysed automatically

Surface Deposits

Surface deposits (for example on photographic paper) can be analysed by Attenuated Total Reflectance (ATR) microscopy or ATR micro-mapping. The choice between the two depends on the appearance and the background knowledge of the sample (and whether your IR microscope system allows ATR mapping). Mapping is required if the surface deposit is believed to be inhomogenous.

ATR has the advantage of being non-destructive and so is particularly important for deposits on negatives when the consumers want them back afterwards.

The spectra below illustrate an amusing problem we were asked to look at:

A consumer sent some negatives into a photo-finisher for re-prints. When the photo-finisher opened the envelope the negatives had white bits all over them. Alarmed, the photo-finisher telephoned the customer. It emerged that the customer had spilt orange juice over the negatives!! To help the photo-finisher to try to clean the negatives, we were asked to identify the white debris and compare it with the envelope to see if it was the same material. The top spectrum shows the gelatin surface of the negative. The middle one shows the surface deposit and the bottom one shows the lining on the inside of the paper. The spectra are not of paper itself, but of inorganic coatings on the paper.

Figure 4. Example of problem solving using the ATR technique.
The deposit matched the lining of the negative pocket.

Conclusion

Our new FTIR microscope system (4) has proved to be:

• Very stable

• Easy to use

• A means of improving our productivity in analysing small samples

About the author:

I am 26 years old. I have been working at Kodak for nearly six years, of which I spent three years doing synthetic organic chemistry, completing an HNC in Chemistry. Since 1995, I have been working in the vibrational spectroscopy section. As well as infrared microscopy I do "routine" bulk infrared, near infrared and Raman spectroscopy. I am also Safety Officer for R&D ("part time"). I find problem solving work both interesting and enjoyable. If you (the reader) are a young person considering a career in this field, I wish you good luck and hope you enjoy it as much as I do.

Acknowledgements

I would like to thank Alan Strawn, my supervisor, for his encouragement and support.

References

1. Mathshop, 127 Middlesbridge Stret, Romsey, Hants, SO51 8HH.

2. Watkins and Doncaster, PO Box 5, Cranbrook, Kent, TN18 5EZ. They can send samples of probes so that you can choose the size best suited to your needs.

3. We use a Spectra-Tech Sample Plan fitted with diamond windows.

4. Perkin-Elmer AutoImage microscope coupled with a PE1000 spectrometer.

Feature Article

7. Identification of compounds in
HPLC-eluates by Raman Spectroscopy

Introduction

The versatility of Raman spectroscopy in determining molecular properties has been proved by solving innumerable problems over many decades. The success of Raman spectroscopy is based on its simple instrumentation which consists of a powerful light source, a well-dispersing monochromator or interferometer and a sensitive detection device. Furthermore, no special efforts have to be made to prepare samples for Raman analysis.

In addition, modern computers enable the prediction of vibrational spectra very rapidly† so that the analysis of transitions is theoretically assisted and this gives a comprehensive interpretation of Raman spectra.

† Editor’s Note : we plan a special edition devoted to the calculation of vibrational frequencies early in Vol III.

Compared to molecular fluorescence emissions, Raman signals are notourisly weak. As a rule of thumb, one million laser photons generate only one Raman photon! The magnitude of the scattered radiation per unit solid angle depends linearly on the intensity of the excitation laser light source, the number of exposed molecules, the fourth power of the absolute scattering wavenumber, and the sum of the squared elements of the scattering tensor. The smallness of the product determines the small yield of Raman photons. The magnitude of scattering tensor elements can be increased by matching the frequency of the laser to an optically allowed transition. In this case, certain scattering signals of the molecule being investigated can be strongly enhanced by resonance effects*. Beside this property, the increase of the frequency and the power of the excitation source as well as the extension of the solid angle of the scattered light are also tools to enhance the scattering intensity.

* Another subject for a future edition.

In this article we are concerned with the best possible detectability of Raman signals by improving the instrumental components especially the imaging of the scattered light [1]. In addition, the imaging conditions of our setup enables us to record scattering signals from small-sized sample. As a result, it is possible to consider Raman spectroscopy as a detection device for HPLC [2-6].

In HPLC using normal conditions, structural isomers often have the same retention times. Thus, it can sometimes be hard to decide which isomer or indeed how many compounds generate a signal in a chromatogram. Since electronic spectra of compounds in the liquid phase have large halfwidths and show only slight differences between them, fluorescence spectroscopy and absorption spectroscopy are overtaxed as conventional detection methods in HPLC. The answer, we think is Raman spectroscopy.

The setup (Figure. 1) can be divided into five functional sub-units, which will be discussed successively: the excitation, the sample cell, the light collection unit, the light-dispersive system, and finally the light detection assembly.

Figure 1. The experimental set-up

The excitation unit consists of an argon ion laser (Coherent, model Innova 90), a band pass filter (F1; Kaiser optics, model HLBF-488-1.0), a focusing lens (Lf), a pinhole (P), and the feedback mirror (M3). The laser is set to a wavelength of 488 nm at a power of 3 W. After passing the sample cell (S), the laser beam is coupled back into the laser by the feedback mirror (radius 60 cm, reflectivity 99.9%). This enhances the laser power to a maximum value of up to 8 W inside the sample. To achieve this increase in power, the feedback mirror must be adjusted in such a way that its radius of curvature fits the radius of the phase front of the laser beam. Compared with a simple intracavity setup, as it has been used mainly for the measurement of gaseous samples [7], the coupled resonator has the advantage that the alignment of its different optical components can be more easily optimized. Furthermore, even if problems arise, e.g., by the formations of thermal lenses inside the sample, Raman spectra can still be recorded at the lower output power of the resonator formed by the mirrors M1 and M2.†

† Editor's Note: The technique sounds more difficult than it is, so follow Hans’ advice and try it. We used it at Southampton as early as 1970 to record the rotational Raman of Cl₂ see Spectrochim. Acta. 28A 1949 (1991). One day I must write a piece for you on rotational Raman. Trouble is it is not vibrational spectroscopy!!

The bandpass filter F₁, which consists of a holographic diffraction grating between two quartz prisms, reduces the spontaneous emission lines from the argon discharge. This task is achieved by effectively directing the exciting laser beam at an angle of 90 degrees. The spectral lines, that contaminate the monochromatic laser radiation are angularly dispersed and further rejected by a pinhole (P). The hole (diameter 3 mm) is drilled into a reflector of stainless steel ®. The reflector R is also part of the light collecting unit. The lens Lf produces a sharp focus of the laser beam with a diameter of about 10 µm in the sample.

The sample cell S has an optical path length of 1 cm and a width of 4 mm. Filling the cell with 1 ml of liquid leads to a column height of about 2.5 cm. This avoids an unnecessarily large cutoff of the scattered light, since the laser beam is focussed inside the cell at a height of 1 cm.

With regard to HPLC; our sample cell can be replaced by a flow cell. An HPLC column (LichroCART 125-4 with Lichrospher 100 RP 18 (Merck)) was used in combination with the appropriate precolumn - LichroCART 4-4. The selected injection volume was 100 µL for each measurement. The flow velocity of the mobile phase (water/acetonitrile 50:50) was set to the commonly selected value of 1 mL/min.

The light collection unit (magnification <10, depending on its alignment) consists of two aspheric multi-element lenses ("Fresnel lenses" from LOT-Oriel), made of acrylic plastic, and a spherical reflector (R). The first lens (L1, focal length 7.6 cm, diameter 15 cm) has a hole of 3 mm diameter in its centre to let the laser beam pass without focusing. The second lens (L2, focal length 45.7 cm, diameter 45 cm) holds the feedback mirror M3, which is fitted into a bore in its centre. This construction reduces the shadowing of the scattered light by the mirror and its holder to a minimum. Furthermore, that part of the light which is scattered in the reverse direction of the imaging lenses is collected by reflector R (radius of curvature 2.5 cm, diameter 4 cm). The reflector is made of highly polished stainless steel and contains a hole (P) for the laser passage.

The use of multi-element Fresnel lenses has several advantages. They are excellent for imaging a pointlike light source into a pointlike image, i.e. for illuminating the entrance slit of a spectrometer. They give better optical apertures (ratios of focal length to diameter), hence larger solid angles of scattered light can be collected and they are better corrected for spherical aberration than single element lenses. Their large diameter diminishes the amount of scattered light lost by the holes in their centres. Finally, since Fresnel lenses are lighter than conventional ones of comparable optical properties, their adjustable mountings easier to construct.

The light detection unit contains the spectrograph (Kaiser optics, model HoloSpec f/1.8i) and the CCD camera (Photometrics, model SDS 9000 with a head CH 270). This spectrograph has an excellent entrance optical aperture (f/1.8) that ensures that practically all the light passing the slit is imaged onto the focal plane. Inside the housing of the spectrograph, a holographic notch filter (SuperNotch Plus model HSPF-488AR-2.0) is inserted between the entrance plane of the spectrograph and the internal slit (SL, 100µm). This filter suppresses the Rayleigh scattering that would generate a large amount of stray light. The reduction of stray light enhances the signal to background ratio considerably.

The camera is cooled with liquid nitrogen (temperature adjusted to -90° C) and equipped with a 1024x256 pixel CCD chip (Metachrome™ II extended UV). Its sensitivity is high enough to detect signals of only a few photons. The spectral range of the setup encloses about 3000 cm-1 from the excitation line, and the achieved spectral resolution in the central region of the chip is about 10 cm-1. This corresponds to illuminating three pixels.

The scattering zone which contributes principally to the light that passes the entrance slit of the spectrograph is very small. It is concentrated in a cylinder of about 10µm diameter and a length of about 1 mm. Most of the scattered light produced from the volume of the diverging beam and outside this zone does not enter the spectrograph. However calculation of the total detected amount of light from the beam in the sample as a function of this cylinder length suggests that the cylinder contributes about two thirds of the available signal. Hence, the main part of the scattered light originates from a sample volume of less than 1 nL.

HPLC measurements start by recording signals from the UV/VIS detector. Once an interesting peak is visible in the chromatogram, the eluate is purged into the Raman sample cell. Otherwise the eluate is directed through a bypass. HPLC peaks typically have a full width of about one minute (with a FWHM of about 20 to 30 s), and provides the required amount of about 1 mL of liquid to fill the sample cell at typical flow rates. In the case of more than one interesting peak in the HPLC spectrum, the samples are collected and passed into the sample cell successively.

The sensitivity of the measurements of the CCD is limited by the amount of background signal. This depends on the number of pixel columns in the selected pixel area and on the total number of incoming photons in the same area. To obtain best results, the spectral range must be set to a region which is characteristic of the examined molecules and not congested by the Raman signals of the solvent. To avoid saturation effects on the CCD, the total measuring time (typically 150 s) is divided into a sequence of several single exposures (typically 10 to 50 s each).

Near the detection limit, the evaluation of the data consists of several steps. The measured spectrum is corrected for background signals by subtracting a properly scaled spectrum of the pure solvent. The remaining background may contain fluorescence signals, which have a much larger spectral bandwidth than the Raman ones, and may show broad artificial structures that arise from imperfect correction. Both contributions to the background signal are removed to obtain a flat baseline. This is carried out by subtracting a carefully smoothed graph of the background. Finally, there are remaining periodic structures in the spectra caused by the CCD camera. These structures contribute a relatively large standard deviation to the Raman signal and the background and in this way limit the sensitivity of the measurements. The resulting spectrum is Fourier filtered.

We define the detection limit by the ratio of the height of Raman signals to the height of signals originated from adjacent residual structures. The detection limit is reached, when the ratio is unity.

The ability of the setup to detect small sample concentrations was first tested by measuring dilute solutions of benzene. Acetonitrile was chosen as the solvent since it provides suitable spectral ranges in which vibrations of the solute are missing. Figure. 2, Figure 3 and Figure 4 present the most prominent Raman signal of benzene at 992 cm-1 in low-concentrated solutions. The peaks appearing between 1025 and 1250 cm-1 at lower concentrations are artificial and their behaviour is similar to noise. Restricting the measurement time for one spectrum to a value of about 150 s, the detection limit is around 3 10-6 mol/L. As can be seen from the number of counts in the spectra, the relation between the signal height and the selected concentrations is not linear. The non-linearity probably originated from both saturation effects of the CCD-chip and the effect of the mathematical procedure, we use to extract the spectropic data from the recorded spectra.

Figure 2. The Raman transition at 992cm^-1 of benzene,
solvent:acetonitrile, 10^-2mol/L

Figure 3. The Raman transition at 992 cm-1 of benzene,
solvent: acetonitrile, 10-4mol/L

Figure 4. The Raman transition at 992 cm-1 of benzene,
solvent: acetonitrile, 3 10-6 mol/L

As a second test, spectra were recorded from a mixture of m-xylene and p-xylene also diluted with acetonitrile. Figure. 5, 6 and 7 present Raman band transitions of p-/m-xylene mixtures at 10-2, 10-3 and at 10-5 mol/L within the range AV 950 and 1275 cm-1. At 10-2 mol/L, all Raman signals of the xylene isomers within the selected spectral range can be detected
(Table 1). The small signals at about 1070, 1120 and 1130 cm-1 are artificial.

The xylenes could still be identified through their main bands at concentrations down to 10-5 mol/L (1µg/mL or 430 ppb) for each isomer. Here, the ratio of the signal heights to those of the noise is slightly larger than unity. Since the spectral position of the xylene signals remain constant, while the noise varies, an identification is possible at this very low concentration.

Figure 5. The Raman spectrum of p-xylene/m-xylene in
acetonitrile at 10^-2 mol/L between 950 and 1250cm^-1

Figure 6. The Raman spectrum of p-xylene/m-xylene in
acetonitrile at 10^-3 mol/L between 950 and 1250cm^-1

Figure 7. The Raman spectrum of p-xylene/m-xylene in
acetonitrile at 10^-5 mol/L between 950 and 1250cm^-1

The final example demonstrates the efficiency of Raman spectroscopy in analysing multi-component systems. Figure. 8 shows Raman spectra of the dichlorophenols isomers each at 10-4 mol/L dissolved in acetonitrile as well as the spectrum of the mixture including all dichlorophenols. The dichlorophenols differ only in the substitution pattern of the two chloroatoms. Since their chromophores are almost identical, the electronic spectra (both absorption and fluorescence spectra) of the isomers are nearly indistinguishable. On the other hand, their Raman spectra are very different. Without going into details it is quite clear from the bottom spectra of P.8 that all the isomers can easily be identified.

Figure 8. Raman spectra of the dichlorophenols between 950 and 1350 cm-1;
the Raman peaks of all spectra are numbered consecutively;
the graph at the bottom presents the collective spectrum;

spectra of single compounds: green: 3,5-dichloro, red: 3,4-dichloro,
blue: 2,6-dichloro, magenta: 2,5-dichloro,
dark blue: 2,4-dichloro, yellow: 2,3-dichloro