Analytical Chemistry

Qualitative and Quantitative aspects of IR Spectroscopy

Sandhya Tiwari
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Introduction
Infrared spectroscopy can be used for qualitative and quantitative analysis. For qualitative analysis, it can be used to identify organic compounds. The IR spectra of an organic compound are a unique fingerprint with many peaks. IR spectra can also be used for quantitative measurements as well. For instance, it can be used to determine the type and quantity of atmospheric pollutants from industrial processes without purifying and separating the sample. Infrared radiation spans a section of the electromagnetic spectrum having wave-numbers from roughly 13,000 to 10 cm1, or wavelengths from 0.78 to 1000 m. It is bound by the red end of the visible region at high frequencies and the microwave region at low frequencies. IR absorption positions are generally presented as either wave numbers or wavelengths. Wave-number defines the number of waves per unit length. Thus, wavenumbers are directly proportional to frequency, as well as the energy of the IR absorption. The wave-number unit (cm1, reciprocal centimeter) is more commonly used in modern IR instruments that are linear in the cm1 scale. In the contrast, wavelengths are inversely proportional to frequencies and their associated energy. The transmittance spectra provide better contrast between intensities of strong and weak bands because transmittance ranges from 0 to 100% T whereas absorbance ranges from infinity to zero. The IR region is commonly divided into three smaller areas: near IR, mid IR, and far IR. Region of Infrared Radiation Near Mid Far Wavelength (in cm) (0.7-1) to 5 5 to (25-40) (25-40) to (200-350)

Qualitative Analysis
The appearance of I.R. instrument revolutionized the way chemists went about identifying organic, inorganic, and biological species. The time required to perform a structural determination was reduced by a factor of ten, one hundred, or even one thousand. Identification of an organic compound is a two-step process. The first step involves determining what functional groups are most likely present by examining the group frequency region. The second step then involves a detailed comparison of the spectrum of the unknown with the spectra of pure compounds that contain all of the functional groups found in the first step. The fingerprint region, from 1200 to 600 cm-1 is particularly useful because small differences in the structure and constitution of a molecule result in significant changes in the appearance and distribution of absorption peaks in this region. Group Frequencies The approximate frequency (or wave-number) at which an organic functional group, such as C=O, C=C, CH, CC, or OH absorbs infrared radiation can be calculated from the masses of the atoms and the force constant of the bond between them. These frequencies, called group frequencies, are seldom totally invariant because of interactions with other vibrations associated with one or both of the atoms composing the group. A range of frequencies can be assigned within which it is highly probable that the absorption peak for a given functional group will be found.

The Fingerprint Region Small differences in the structure and constitution of a molecule result in significant changes in the distribution of absorption peaks in this region of the spectrum that extends from about 1200 to 700 cm-1. As a consequence, a close match between two spectra in this fingerprint region constitutes strong evidence for the identity of compounds yielding the spectra.
Although many of the bands in the IR spectrum can be used to identify particular functional groups, the region from about 500 - 1500 cm-1 usually contains a complicated series of bands, many of which overlap. This region contains absorptions which correspond to bending as well as stretching vibrations. It is difficult to pick out individual bands and assign them reliably to a particular functional group or bond in a molecule. This part of the spectrum is called the fingerprint region. Although this region is not very useful for detecting which functional groups are present, it can still be useful for identifying the molecule provided its IR spectrum is already known. Each compound's IR spectrum is unique and so, by matching a spectrum to a library or database either electronically or by eye, it can be positively identified. Such methods are used in forensic, analytical and environmental applications as well as in research chemistry.

The Fingerprint Region, 1300 to 910 cm-1, include the contributions from complex interacting vibrations, giving rise to the generally unique fingerprint for each compound. A good match between the IR spectra of two compounds in all frequency ranges, particularly in the fingerprint region, strongly indicates that they have the same molecular structures. Detailed interpretation of IR bands in this region is difficult. However, some assignments of bands in the fingerprint region to a few important vibrational frequencies of functional groups can be done when IR absorptions in other regions are correlated together .

Overtones and Combination bands

The sound we hear is a mixture of harmonics, that is, a fundamental frequency mixed with multiples of that frequency. Overtone bands in an infrared spectrum are analogous and are multiples of the fundamental absorption frequency. The energy required for the first overtone is twice the fundamental, assuming evenly spaced energy levels. Since the energy is proportional to the frequency absorbed and this is proportional to the wave number, the first overtone will appear in the spectrum at twice the wave number of the fundamental. Combination bands arise when two fundamental bands absorbing at 1 and 2 absorb energy simultaneously. The resulting band will appear at (1 + 2) wave numbers.

Once an infrared spectrum has been recorded, the next stage of this experimental technique is interpretation. Fortunately, spectrum interpretation is simplified by the fact that the bands that appear can usually be assigned to particular parts of a molecule, producing what are known as group frequencies. The characteristic group frequencies observed in the mid-infrared region have already been discussed. The types of molecular motions responsible for infrared bands in the near-infrared and far-infrared regions are also introduced.

In addition to the fundamental frequency band, some other bands which are multiple of the fundamental frequency band also occur but their intensity is very low and so generally they do not hamper the process of spectral manipulation.

Fermi resonance
Two conditions must be satisfied for the occurrence of Fermi Resonance:

The two vibrational states of a molecule transform according to the same irreducible representation of the molecular point group. In other words the two vibrations must have the same symmetries (Mullikan symbols). The transitions (accidentally) have almost the same energy.

Fermi resonance most often occurs between normal and overtone modes, if they are nearly coincident in energy. Fermi resonance leads to two effects. First, the high energy mode shifts to high energy and the low energy mode shifts to still lower energy. Second, the weaker mode gains intensity (becomes more allowed) and the more intense band decreases in intensity. The two transitions are describable as a linear combination of the parent modes. Fermi resonance does not really lead to additional bands in the spectrum.
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Quantitative Analysis
In quantitative analysis of a compound, we actually try to determine the amount of a particular component present in the sample. There are various methods for this and they are discussed below.

Computer Search Systems

Virtually all infrared instruments manufactures now offer computer search systems to assist chemist in identifying compounds from stored infrared spectral data. The position and relative magnitudes of peaks in the spectrum of the analyte are determined and stored in memory to give a peak profile, which can then be compared with profiles of pure compounds stored. The computer then matches profiles and prints a list of compounds having spectra similar to that of the analyte. Usually the spectrum of the analyte and that of each potential match can then be shown simultaneously on the computer display for comparison. Quantitative infrared spectroscopy can provide certain advantages over other analytical techniques. This approach may be used for the analysis of one component of a mixture, especially when the compounds in the mixture are alike chemically or have very similar physical properties (for example, structural isomers). In these instances, analysis using ultraviolet/visible spectroscopy, for instance, is difficult because the spectra of the components will be nearly identical. Chromatographic analysis may be of limited use because separation, of say isomers, is difficult to achieve. The infrared spectra of isomers are usually quite different in the fingerprint region. Another advantage of the infrared technique is that it can be non-destructive and requires a relatively small amount of sample.

Spectrum Manipulation
Quantitative infrared absorption methods differ somewhat from ultraviolet/visible molecular spectroscopic methods because of the greater complexity of the spectra, the narrowness of the absorption bands, and the instrumental limitations of infrared instruments. Quantitative data obtained with infrared instruments are generally significantly inferior in quality to data obtained with ultraviolet/visible spectrophotometers.

There are a number of techniques available to users of infrared spectrometers that help with both the qualitative and quantitative interpretation of spectra.
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Baseline Correction It is usual in quantitative infrared spectroscopy to use a baseline joining the points of lowest absorbance on a peak, preferably in reproducibly flat parts of the absorption line. The absorbance difference between bottom and the top of the band is then used.

Smoothing Noise in a spectrum can be diminished by a smoothing process. After a spectrum is smoothed, it becomes similar to the result of an experiment at a lower resolution. The features are blended into each other and the noise level decreases. A smoothing function is basically a convolution between the spectrum and a vector whose points are determined by the degree of smoothing applied. Generally, a degradation factor is required, which will be some positive integer. A value, says one, will produce only subtle changes, while a height value has a more pronounced effect on the spectrum.

Difference Spectra The most straightforward method of analysis for complex spectra is difference spectroscopy. These techniques may be carried out by simply subtracting the infrared spectrum of one component of the system from the combined spectrum to leave the spectrum of the other component. If the interaction between components results in a change in the spectral properties of either one or both of the components, the changes will be observed in the difference spectra. Such changes may manifest themselves via the appearance of positive or negative peaks in the spectrum. Spectral subtraction may be applied to numerous applications and can be used for the data collected for solutions. In order to obtain the spectrum of a solution it is necessary to record spectra of both the solution and solvent alone. The solvent spectrum may then be subtracted from the solution spectrum. Under certain circumstances, the spectrum due to the regions of solvent may be very intense, making simple subtraction impossible. This situation can make it difficult to investigate the sample spectrum, as solvent bands may overlap with the region under investigation. In such experiments, attenuated total reflectance spectroscopy provides a suitable approach. Subtraction may be applied to solid samples and is especially useful for mulls. The spectrum of the mulling agent can be subtracted, hence giving the spectrum of the solid only.
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Derivatives Spectra may also be differentiated. The benefits of derivative techniques are two-fold. Resolution is enhanced in the first derivative since changes in the gradient are examined. The second derivative gives a negative peak for each band and shoulder in the absorption spectrum. The advantage of derivatization is more readily appreciated for more complex spectra. With FTIR, it is possible to apply what is known as Fourier Derivation. During the process, the spectrum is first transformed into an interferogram. It is then multiplied by appropriate weighing function and finally it is re-transformed to give the derivative. This technique provides more sensitivity than conventional derivatization.

Deconvolution Deconvolution is a process of compensation for the intrinsic line widths of bands in order to resolve overlapping bands. This technique yields spectra that have much narrower bands and is able to distinguish closely spaced features. The instrumental resolution is not increased, but the ability to differentiate spectral features can be significantly improved. The peaks at quite close wave-numbers are now easily distinguished. The deconvolution technique generally involves several steps: computation of an interferogram of sample by computing the inverse Fourier transform of the spectrum, multiplication of the interferogram by a smoothing function and by a function consisting of a Gaussian Lorentzian band shape, and Fourier transformation of modified interferogram. The deconvolution procedure is typically repeated iteratively for best results. At iteration, the line-shape is adjusted in an attempt to provide narrower bands without excessive distortion. There are three parameters that can tune the lineshape, as follows. First, the proportion of Gaussian and Lorentzian origins of band-shape. For instance, it will vary depending on whether a solid, liquid or gas is being investigated. The second factor is the half width: this is the width of line-shape. The half width is normally the same as or larger than the intrinsic line-width of the band. If the value specified is too large, distinctive negative side-lobes are produced. The narrowing attempted on a scale from 0 to1. If the narrowing function is too small, then the resulting spectrum will not be significantly different from theoriginal, while if too large, the spectrum may produce false peaks as noise starts to bedoconvoluted. In the figure shown below, the first figure represents the original peak obtained whereas the the second figure gives the first derivative of the original peak from which a lot of information can be decoded.

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Curve fitting Quantitative values for band areas of heavily overlapped bands can be achieved by using curve fitting procedures. Many of these are based on a least-squares minimization procedure. Least squares curve fitting covers the general class of techniques whereby one attempts to minimize the sum of the squares of the difference between an experimental spectrum and computed spectrum generated by summing the component curves. Generally, the procedure involves entering the values of the wave-numbers of the component bands and then the program determines the best estimate of the parameters of component curves. Apart from the obvious variables of peak height and width, the type of band-shape needs to be considered. The class of bandshape of an infrared spectrum depends on the type of sample.

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Analysis of samples
Both solid and liquid substances can be estimated with the help of ir spectroscopy. In liquid state, generally the sample is mixed with nujol and for the solid state analysis, KBr pellets are used. Liquid Samples The quantitative analysis of a component in solution can be successfully carried out given that there is a suitable band in the spectrum of the component of interest. The band chosen for analysis should have a higher molar absorptive, not overlap with other peaks from other components in the mixture or the solvent, be symmetrical, and give a linear calibration plot of absorbance versus concentration. Most simple quantitative infrared methods of analysis use the intensities of the carbonyl or amine or hydroxyl groups. The carbonyl stretching band is the most used band because it is a strong band in a spectral region relatively free of absorption by other functional groups. In addition, the carbonyl band is not as susceptible to distortion as are hydroxy or amine bands. Let us take the example of aspirin dissolved in chloroform. The best peak to choose in this example is the C=O stretching band of aspirin, observed at 1764 cm-1, because it is an intense peak and lies in a region where there is no interference from the chloroform spectrum. The next step is to draw a calibration plot of absorbance versus aspirin concentration. A series of chloroform solutions of known aspirin concentration is prepared and the infrared spectrum of each solution recorded by using a 0.1 mm NaCl transmission cell. The absorbance of the 1764 cm-1 band is measured for each solution, with the results obtained being listed in the table below. The data can be graphed to give a linear plot. The next stage in the analysis is to determine the amount of aspirin present in a chloroform solution of unknown concentration. Using the calibration graph, the concentration of aspirin comes out to be 67 mg/mL.

Concentration (mg / mL )

Absorbance at 1656 cm-1

0 5 10 15 20

0.000 0.105 0.190 0.265 0.333

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Solid samples Solid mixtures can also be quantitatively analysed. These are more susceptible to errors because of the scattering of radiation. Such analyses are usually carried out with KBr discs or in mulls. The problem here is the difficulty in measuring the path length. However, this measurement becomes unnecessary when an internal standard is used. When using this approach, addition of a constant known amount of an internal standard is made to all samples and calibration standards. The calibration curve is then obtained by plotting the ratio of the absorbance of the analyte to that of the internal standard, against the concentration of the analyte. The absorbance of internal standard varies linearly with the sample thickness and thus compensates for this parameter. The discs or mulls must be prepared under exactly the same conditions to avoid intensity changes or shifts in band position. The standard must be carefully chosen and it should ideally possess the following properties, have a simple spectrum with very few bands, be stable to heat and not absorb moisture, be easily reduced to particle size less than the incident radiation without lattice deformations, be non toxic, should give clear disc in short time, and be readily available in pure state. e.g : calcium carbonate, sodium azide and lead thiocyanate.

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Bibliography

1. Quantitative Analysis from Analytical Chemistry by Barbara Stuart (4th Editon) -- Page No : 45-65 2. Qualitative Aspects from Fundamentals Of Analytical Chemistry by Skoog and Niemann.(4th edition ) 3. Online copy of Mallinckrodt Baker Division. 4. Spectroscopy of organic compounds by P.S.Kalsi New Age International Publishers Fifth edition.

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