Spectral subtraction, FTIR

ATR spectroscopy in the infrared has been used extensively in protein adsorption studies. Transmission IR spectra of a protein contain a wealth of conformational information. ATR-IR spectroscopy has been used to study protein adsorption from whole, flowing blood ex vivo 164). Fourier transform (FT) infrared spectra (ATR-FTIR) can be collected each 5-10 seconds165), thus making kinetic study of protein adsorption by IR possible 166). Interaction of protein with soft contact lens materials has been studied by ATR-FTIR 167). The ATR-IR method suffers from problems similar to TIRF there is no direct quantitation of the amount of protein adsorbed, although a scheme similar to the one used for intrinsic TIRF has been proposed 168) the depth of penetration is usually much larger than in any other evanescent method, i.e. up to 1000 nm water absorbs strongly in the infrared and can overwhelm the protein signal, even with spectral subtraction applied. 

Spectral subtraction and spectral search aid the identification of evolved gases, which are often a mixture of products. Nevertheless, for unambiguous identification of unknown volatiles more powerful methods are required. Jansen and co-workers [86] have incorporated a parallel mass spectrometer onto the FTIR stage of a thermogravimetry-FTIR (TG-FTIR). The sample is thermally decomposed by TGA and the products collected in a Tenax (absorbent charcoal) trap. After desorption, the products are separated by a GC and the sample split, with 99% going to the IR spectrometer and 1% to the mass spectrometer. 

The infrared spectrum of diltiazem hydrochloride dispersed in potassium bromide was recorded at 4 cm 1 resolution on a Nicolet Model 740 FTIR (11). Figure 3 shows the diltiazem HCI vibrational features in the 4000 to 400 cm-1 region after spectral subtraction of the absorptions (3430 and 1629 cm 1) due to adsorbed water. Table I lists characteristic frequencies, relative intensities, and vibrational assignments for the primary diltiazem HCI absorption bands. 

Figure 10 Example of reaction monitoring by FTIR spectroscopy through the use of spectral subtraction. The oxidation of trihnolenin at 40 C as a function of time can be followed by monitoring the increase in the hydroperoxide absorption band (3444 cm ) and the decrease in the cis (H-C=C) absorption band (3011 cm ). These changes are clearly observed by subtracting the spectrum of the unoxidised oil from subsequent spectra recorded as a function of time.

Figure 11 FTIR spectra of a 10% solution of sucrose in water A) and of water B) and the difference spectrum obtained by spectral subtraction (A-B). The features in the difference spectrum are the absorption bands of sucrose. The region between 3500 and 3150 cm-i is obscured by digitisation noise due to the intense water absorption band in this region.

Citrate washed hydrogel coatings were examined by Fourier Transform Infrared (FTIR) spectroscopy with spectral subtraction and were found to consist of a reaction product of polymers 10a and 52 and to be free of alginate. 

The Fourier transform Raman (FT-Raniiin) instrument uses a Michelson interferometer, similar to that used in FTIR spectrometers, and a eontinuoits-wave (CW) Nd-YA(i laser as shown in Figure 18-12. fhe use of a 1064-nm (1.064-pm) source virtually eliminates fluorescence and photodecomposition of samples. Hence, dyes and other fluorescing compounds can be investigated with I T-RarniUi instruments. T he Ff-Raman instrument also provides superior Irequency precision relative to conventional instruments, which enable spectral subtractions and high resolution measurements. 

Enhancement of the accuracy of quantitative infrared determinations by use of mathematical manipulation of the spectral data as performed by Fourier Transform (FTIR) and Coiqruterlzed Dispersive (GDIS) Infrared Spectroscopy has been compared. Cotton-polyester blends and cotton treated with THPOH-NHo and with dimethyloldihydroxy-ethylene urea (DMDHEU) were analyzed by FTIR and GDIS. The mathematical techniques used Included direct spectral subtraction attd spectral subtraction combined with analysis of peak areas. 

The use of FTIR spectrometers is now almost universal. The accuracy, sensitivity and reproducability of the new instrumentation have facilitated the development of a range of new data processing methods such as spectral subtraction, deconvolution, curve resolving etc., and new sampling techniques, but it is essential to remember that the underlying principles of infrared spectroscopy have not changed. The precautions necessary to avoid spectral artefacts are thus an important aspect of the work. 

FTIR spectra of syndiotactic PS semi-crystalline samples were examined by using the spectral subtraction approach. It was shown that FTIR analysis provided a direct method for the evaluation of the amount of alpha and beta form crystalline phases, although both contain chains in the same conformation (trans-planar). 50 refs. 

An example of where FTIR spectroscopy may be used as a tool for the investigation of plant material is in the examination of the quality of an animal food such as alfalfa. There is a relationship between the quality of alfalfa as a food supplement and the plant age. Infrared microscopy can be used to compare an old plant with a young plant [53]. Figure 7.18 shows the spectra of an old and young leaf (both 100 x 100 p,m). At first sight, it is difficult to discern the major differences between these spectra. However, spectral subtraction illustrates some... 

A Fourier transform infrared (FTIR) speetrometer equipped with an attenuated total refleetanee (ATR) aeeessory is used for the study of surfaees and coatings. A microscope attachment is useful for identifying particulate impurities. Through the technique of computerized spectral subtraction, many sample mixtures may be identified by FT-IR without prior physical separation and, thus, the technique lends itself to compositional analysis. Typical applications of FT-IR are the study of surfaces of polymers by ATR, identification of samples isolated by thin-layer chromatography or other preparatory chromatographic techniques, identification of impurities in polymer and polymer blends, product characterization and product formulations by spectral subtraction, as well as routine analysis of solid and liquid materials. 

This internal registration of wavelength provides FTIR instruments with unparalleled wavelength accuracy and repeatability, enabling mathematical manipulations such as spectral subtraction to be performed. Note, however, that if laser and infrared radiation differ in their pathlengths after modulation by the interferometer, accuracy is compromised. 

Normal sample, modern FTIR spectrometer 1-4 minute scan time, with spectral resolution of 2-4 cm providing a signal-to-noise ratio (S/N) of between 1000 1 and 10,000 1. If a spectrum is to be used for qualitative purposes only, then a S/N of approximately 1000 1 to 2000 1 is normally adequate and is typically obtained in well under 1 min often within a few seconds. For quantitative apphcations, or applications involving spectral subtraction or resolution enhancement techniques, higher signal-to-noise performance is required. Note that scanning for more than 256 scans may approach a point of diminishing returns in terms of spectral quality and S/N improvement (limited by the law, and the performance of the... 

Polymer samples can be analysed in all possible textures and excellent spectra can be obtained. FTIR exhibits sensitivity to sample geometry and sample surface. As the additives are heterogeneously distributed in the polymer, measurements at various positions are recommended when necessary. The usefulness for exhaustive IR in-polymer analysis of additive packages containing unknown components (i.e. not contained in any reference library) is limited by the inherent characteristics of the method (essentially only functional group identification). Unique identification of unknown components may also be restricted by interference with co-additives and absorption of the polymeric matrix. Spectral subtraction of an appropriate reference polymer may be used to remove matrix interferences and allows tracing of minor components. However, this is not al-... 

The National Physical Laboratory offers a service for calibrating the transmittance scale of mid-IR spectrophotometers [55]. Excellent wavelength accuracy is an important property of FTIR, making highly accurate spectral subtraction possible. 

Brandolini et al. [119] have pointed out some drawbacks to the use of FTIR spectroscopy for quantitative analysis of the extent of phosphite and phos-phonite additive degradation in PE. Band positions are not as distinctive as P NMR resonances. Consequently, it is difficult to distinguish degradation products of similar additives, such as A and B of Fig. 9.3 of ref. [1]. Quantitation in FTIR is also not as straightforward as in NMR. To accurately assess the extent of degradation, appropriate standards would have to be developed. Most importantly, however, the FTIR spectrum contains absorbances from the polymer background and other additives which may obscure the peaks of interest. Spectral subtraction of an appropiate reference polymer can obviate some of this concern, if available. Use of a similar, but not identical, polymer can result in artifacts. In the specific case at hand, the polymer sample was pressed as a 0.1 mm film. An appropriate, secondary oxidant-free reference polymer was available so that spectral subtraction could be performed to remove matrix interferences. 

The surface of printed paper has been examined through fully automated /xATR-FTIR mapping [464]. Compositional differences attributed to the printed ink, kaolinite, and cellulose distributions were revealed, which are not discemable in the visible. After spectral subtraction of the carbonate also DOP and an aromatic acrylate, both used in paper manufacturing, could be identified. Coles et al. [465] have compared /iFTIR and ATR-FTIR in the quantitative determination of fillers such as kaolin clay in polyethylene/vinyl acetate. Although ATR-FTIR is not as sensitive to kaolin as /xFTIR, the former provides a larger sampling area and more consistent results. ATR-FTIR is sometimes used for in-depth analysis. 

The use of liquid crystals as orienting media in polarized infrared spectroscopy, though first demonstrated (as applied to solutes) in the late 1960s [310, 320], has received considerably less attention due to the fact that solvent absorptions severely limit the spectral range accessible to investigation, particularly with conventional dispersion instruments [1, 304], Infrared linear and circular dichroism (IR-LD and IR-CD, respectively) of liquid crystalline materials themselves have, of course, been studied extensively [2]. With the advent of commercial FTIR spectrometers, whose enhanced sensitivity and superior spectral subtraction capabilities allow for far greater precision in IR spectral measurements than is possible with dispersion instruments, activity in this field has picked up considerably [314,321 - 324] and has been reviewed recently [325]. 

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