Applications of UV Spectroscopy

Before NMR spectroscopy and mass spectrometry revolutionized the structural elucidation of organic molecules, UV spectroscopy was an important technique and was used to identify the key chromophore of an unknown molecule. The importance of UV is much diminished nowadays, but it still retains its place in certain applications, such as the determination of kinetic parameters, (the Michaelis constant) and A cat (the turnover rate of an enzyme, in molecules per second), for a number of enzymic reactions and in the analysis of pharmaceuticals. 

The molar absorptivity (e) of a known molecule is constant under identical conditions of solvent, concentration and path length, and can be used to quantify the amount of a particular pharmaceutical in a tablet. Such assays form the basis of many quality assurance procedures in the pharmaceutical industry, and have been extensively used by the British Pharmacopoeia (B.P.). More recently, however, high-performance liquid chromatography (HPLC) has replaced UV analysis in many B.P. assays, as most industrial analyses routinely use HPLC. 

Derivatizaiion lo prepare a derivative ol a compound e.g. an ester derivative of a carboxylic acid). 

In the pharmaceutical industry, medicines are standardized to a particular weight of active pharmaceutical per unit (for example, mg of substance per tablet), so the weight of a substance in a tablet, or volume of medicine, is of more interest than the number of moles. For most pharmaceutical preparations, then, we use the specific absorbance, A(l%, 1 cm), which is the absorbance (logio fo/f) of a 1% w/v solution (i.e. 1 g of substance in 100 cm solvent) in a 1 cm path length cell, in place of the molar absorptivity, e. This is the absolute method of substance identification, and the absorbance. A, can be related to T(l%, 1 cm) by equation (2.4)  

The absolute method relies on the UV spectrophotometer being accurate in the measurement of wavelength and intensity, and the B.P. specifies methods for the calibration of both. However, UV-Vis spectra are not specific for any one particular substance, as many UV absorbing molecules have similar UV spectra, and this method is used in conjunction with several identification tests, as described in the B.P. 

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There is very little published information on the UV spectra of 1,2-benzisothiazoles, though more data are available on the 2,1-isomers. The spectra are complex with as many as six maxima above 200 nm. Representative wavelengths of maxima are collected in Table 12. In all cases the most intense bands (e > 15 000) are those at short wavelengths, but all the bands indicated in the table have molar absorptivities greater than 4000, except those of 3-amino-2,l-benzisothiazole. Saccharin absorbs weakly at 350 nm and 277 nm, with intense bands below 230 nm (ethanol solvent) (82UP41700>. It exists as the anion except in acid solutions. The UV spectra of cations formed from 3-amino-2,l-benzisothiazole are discussed in (69CB1961>. Further applications of UV spectroscopy in studying tautomeric... 

UV spectroscopy is a sensitive tool for the elucidation of structures of molecules only if the different tautomers have distinct spectra. In some cases, cytosine or cytidine and their alkylated analogs have similar spectra with seriously overlapping bands, and thus the application of UV spectroscopy to the elucidation of the tautomeric structures of the molecules is inconclusive. In particular, the method is unreliable for the determination of tautomeric equilibria of cytosine (see below). Nevertheless the UV absorption spectra confirm the conclusions drawn from other studies as to the main form of cytosine and cytidine in aqueous solution at room temperature. 

The chromophoric properties of triazolines permit the application of UV spectroscopy for kinetic measurements of formation and decomposition of the triazoline adducts.24,79,113,205 However, triazolines, when irradiated in the region of their UV absorption, undergo facile decomposition with nitrogen expulsion (Section IV,D,1).79,113... 

Some of the useful applications of UV spectroscopy are listed below. 

As to the problem of whether or not Cu, which is inside the frame of Table III, is a catalyst for dehydrogenation of cyclohexane and for hydrogenation of benzene, a discussion has been carried on. A recent study of Erofeev and Nikiforova (203), with the application of UV spectroscopy showud that copper is indeed a catalyst in this reaction, as well as in the direct dehydrogenation of cyclohexane to benzene. This... 

H. H. Jaffe and M. Orchin, Theory and Application of UV Spectroscopy. John Wiley Sons, New York, 1962. 

The application of UV spectroscopy to the analysis of nonionic hydrocarbon-type surfactants is limited to nonionics, which contain functional groups which absorb in the UV region, such as aromatic nuclei [65]. The main functional group of nonionics, the oxyethylene ether linkage, does not absorb in the UV region. In spite of this limitation, UV spectroscopy can be useful for determining impurities in nonionic fluorinated surfactants. 

The main application of UV VIS spectroscopy which depends on transitions between electronic energy levels is in identifying conjugated tt electron systems... 

Applications Applications of UV/VIS spectrophotometry can be found in the areas of extraction monitoring and control, migration and blooming, polymer impregnation, in-polymer analysis, polymer melts, polymer-bound additives, purity determinations, colour body analysis and microscopy. Most samples measured with UV/VIS spectroscopy are in solution. However, in comparison to IR spectroscopy additive analysis in the UV/VIS range plays only a minor role as only a limited class of compounds exhibits specific absorption bands in the UV range with an intensity proportional to the additive concentration. Characteristic UV absorption bands of various common polymer additives are given in Scheirs [24],... 

Stabilisers are usually determined by a time-consuming extraction from the polymer, followed by an IR or UV spectrophotometric measurement on the extract. Most stabilisers are complex aromatic compounds which exhibit intense UV absorption and therefore should show luminescence in many cases. The fluorescence emission spectra of Irgafos 168 and its phosphate degradation product, recorded in hexane at an excitation wavelength of 270 nm, are not spectrally distinct. However, the fluorescence quantum yield of the phosphate greatly exceeds that of the phosphite and this difference may enable quantitation of the phosphate concentration [150]. The application of emission spectroscopy to additive analysis was illustrated for Nonox Cl (/V./V -di-/i-naphthyl-p-phcnylene-diamine) [149] with fluorescence ex/em peaks at 392/490 nm and phosphorescence ex/em at 382/516 nm. Parker and Barnes [151] have reported the use of fluorescence for the determination of V-phenyl-l-naphthylamine and N-phenyl-2-naphthylamine in extracted vulcanised rubber. While pine tar and other additives in the rubber seriously interfered with the absorption spectrophotometric method this was not the case with the fluoromet-ric method. 

An interesting and powerful new development in Raman spectroscopy of catalysts is the use of a UV laser to excite the sample. This has two major advantages. First, the scattering cross section, which varies with the fourth power of the frequency, is substantially increased. Second, the Raman peaks shift out of the visible region of the spectrum where fluorescence occurs. The reader is referred to Li and Stair for applications of UV Raman spectroscopy on catalysts [40]. 

Applications of UV/visible spectroscopy to pharmaceutical quantitative analysis... 

MS. This section will rather briefly concentrate on some of the more recent applications of UV absorption spectroscopy in the flavonoid field. It will mainly cover online UV absorption spectroscopy in chromatography (Section 2.5.1). Because of the current importance of UV Vis in the study of anthocyanins, some more UV Vis spectral details have been included related to this pigment group (Section 2.5.2). Section 2.5.3 indicates the recent use of this technique in studies of flavonoids interacting with other compounds. 

Similarly, fused silica windows are not required except for the most demanding applications in UV spectroscopy, so fitting the vacuum shroud with a pair of well-polished KBr windows is usually appropriate, unless one needs IR transparency at 500-200 cm when one has to use Csl windows. If one needs to probe the far-lR range (<200 cm ) one has to use polyethylene windows (the covers that come with KF flanges have proven adequate), but this makes it of course impossible to probe the same samples in the mid-IR or UV-vis range. 

An authoritative review covering application of UV photoelectron spectroscopy to heterocycles appeared in 1974 <74PMH(6)l) and important work on saturated multi oxygen and sulfur compounds is discussed therein. However, the studies of 1,4-dioxin, its dibenzo analogue, 1,4-dithiin and phenoxathiin, which are referred to in the first part of this section, appeared after the publication of that review. 

The initiation events involving dienes and styrene in hydrocarbon solvents have been thoroughly and accurately studied by the application of UV and visible spectroscopy. The archetype of such studies is the now classic 1960 study of Worsfold and Bywater 94) on the n-butyllithium-styrene system in benzene. The reaction was found to follow the relationship ... 

To avoid these two difficulties for our analysis, we used reflectivity spectra of thin, high-quality crystals, with surface structures subtracted out to recover the bulk reflectivity of a perfect semiinfinite crystal. Since this problem and its inherent difficulties are often ignored, we describe below our approach, up to the final application to UV spectroscopy, in order to point out the source of discrepancies originating either from the sample spectra or from the theoretical approximations. 

Nevertheless, the mere knowledge of r(co) 2 is not sufficient to provide the two parameters v(co) and k(u)) both reflection and transmission spectra are needed. This is the main reason that the KK transformation is used for reflectivity spectra, as we summarize below in application to our problem of UV spectroscopy. 

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