Multiwavelength method

Quantitative techniques are essentially of three kinds single wavelength methods, multiwavelength methods and derivative spectroscopy. 

Multiwavelength methods. Least squares curve fitting techniques may be used in the determination of multicomponent mixtures with overlapping spectral features. Two classical quantitation methods, the Classical Least Squares (CLS) mode and the Inverse Least Squares (ILS) model, are applied when wavelength selection is not a problem. CLS is based on Beer s law and uses large regions of the spec-tram for calibration but cannot cope with mixtures of interacting constituents. ILS (multivariate method) can accurately build models for complex mixtures when only some of the constituent concentrations are known. 

Two additional methods for determining the composition of a mixture deserve mention. In multiwavelength linear regression analysis (MLRA) the absorbance of a mixture is compared with that of standard solutions at several wavelengths. If Asx and Asy are the absorbances of standard solutions of components X and Y at any wavelength, then... 

The intensity differences obtained in the diffraction pattern by illuminating such a crystal by x-rays of different wavelengths can be used in a way similar to the method of multiple isomorphous replacement to obtain the phases of the diffracted beams. This method of phase determination which is called Multiwavelength Anomalous Diffraction, MAD, and which was pioneered by Wayne Hendrickson at Columbia University, US, is now increasingly used by protein cystallographers. 

Hendriksen, B. A., Sanchez-Eelix, M. V., Tam, K. Y. A new multiwavelength spectrophotometric method for the determination of the molar absorption coeffidents of ionizable drugs. Spectrosc. Lett. 2002, 35, 9-19. 

A multiwavelength approach might have been considered as an alternative to chemical derivatisation. Ruddle and Wilson [62] reported UV characterisation of PE extracts of three antioxidants (Topanol OC, Ionox 330 and Binox M), all with identical UV spectra and 7max = 277 nm, after reaction with nickel peroxide in alkaline ethanolic solutions, to induce marked differentiation in different solvents and allow positive identification. Nonionic surfactants of the type R0(CH2CH20) H were determined by UV spectrophotometry after derivatisation with tetrabromophenolphthalein ethyl ester potassium salt [34]. Magill and Becker [63] have described a rapid and sensitive spectrophotometric method to quantitate the peroxides present in the surfactants sorbitan monooleate and monostearate. The method, which relies on the peroxide conversion of iodide to iodine, works also for Polysorbate 60 and other surfactants and is more accurate than a titrimetric assay. 

Lee et al. [30] described a micellar electrokinetic capillary chromatographic method for the determination of some antiepileptics including valproic acid. They used a fused silica capillary column (72 cm x 50 pm) and SDS as the micellar phase and multiwavelength UV detection. Reaction conditions, such as pH and concentration of running buffer were optimized. Solutes were identified by characterizing the sample peak in terms of retention time and absorption spectra. Recoveries were 93-105%. 

La Fortelle, E. D. and Bricogne, G. (1997). Maximum-likelihood heavy-atom parameter refinement for multiple isomorphous replacement and multiwavelength anomalous diffraction methods. Method Enzymol. 276, 472 94. 

Ramakrishnan, V. and Biou, V. (1997). Treatment of multiwavelength anomalous diffraction data as a special case of multiple isomorphous replacement. Method Enzymol. 276, 538-557. 

In this work we describe a method based on modern LC which avoids an extraction step. This technique is rapid and selective and gives, with multiwavelength detection, good qualitative and quantitative information. 

Identification is considerably improved by the use of multiwavelength detection and absorbance ratioing. It was possible to analyse some commercial hair dyes with the method described here. However, several substances were found that could not be identified with the present standard substances. Recent results in this laboratory make a possible improvement of the chromatography probable. Future work will consider this aspect and an increased number of standard substances (17) will be examined. 

Model-based nonlinear least-squares fitting is not the only method for the analysis of multiwavelength kinetics. Such data sets can be analyzed by so-called model-free or soft-modeling methods. These methods do not rely on a chemical model, but only on simple physical restrictions such as positiveness for concentrations and molar absorptivities. Soft-modeling methods are discussed in detail in Chapter 11 of this book. They can be a powerful alternative to hard-modeling methods described in this chapter. In particular, this is the case where there is no functional relationship that can describe the data quantitatively. These methods can also be invaluable aids in the development of the correct kinetic model that should be used to analyze the data by hard-modeling techniques. 

Several unsuccessful attempts were made to analyze the fluorophor produced by the reaction of MIC with diacetylmonoxime using HPLC and a multiwavelength fluoroescence detector. Because of a series of analytical problems, a search was initiated for a method that would lend itself to liquid chromatographic analysis more readily. Since methyl isocyanate probably reacts with water according to the following equation ... 

Today most pKas are determined by classical titration with UV (Albert) or potentiometric (Sirius GLpKa) detection. These methods could be coupled with multiwavelength spectrophotometer detection (Sirius, D-PAS). Tam et al. compared both methodologies and found good correlation between them. 

Ultraviolet spectrophotometers cont.), single-beam, 225 standardisation, 226 Ultraviolet spectrophotometry, 221-232 absorption cells, 226 colorimetry, 228 derivative, 230 difference method, 229 dual-wavelength, 229 identification by, 231 influence of pH, 224 influence of solvent, 224 laws of absorption, 222 quantitative applications, 227 stray-light effects, 224 Ultraviolet-visible detector, 202 multiwavelength, 211 Unicontin, 1011 Unidiarea, 474 Unidone, 356 Uniflu, 557, 893 Unilobin, 709 Unimycin, 846 Uniphyllin, 1011 Uniprofen, 677 Unisom, 576... 

Recently, microconstant measurements have been validated for 25 drug molecules, based primarily on multiwavelength spectroscopy. The apphed methodologies, however, are difficult and limited up to triprotic molecules when no symmetry constraints are present.In more complex cases, the microspecies resolution is still possible by applying some in silico methods for predicting ionization constants. 

X-ray crystallography is a widely used teclmique for the three-dimensional structure analysis of RNA molecules. The extraction of information from X-ray diffraction patterns from RNA crystals requires proper phase determinations using methods such as multiwavelength anomalous dispersion in which placement of specific heavy atoms is required. Seleiuum-modi-fled ribonucleotides have been used for these purposes by Du et al. (36), because the selenium atom does not seem... 

A Laue photograph -is produced by irradiating a stationary crystal with a beam of X rays that has a wide range of wavelengths ( white radiation). It differs from all of the other methods for collecting diffraction data in that the crystal is stationary throughout the experiment. Diffraction is therefore dependent on the multiwavelength feature of the... 

---