Multiwavelength analysis

Sen, M., Thevanat, C., and Prioul, J.L. 1997. Simultaneous spectrophotometric determination of amylose and amylopectin in starch from maize kernel by multiwavelength analysis. J. Cereal Sci. 26 211-221. 

In addition, since many compounds have characteristic UV absorption profiles, multiwavelength analysis helped their identification. Because the instrument used was not capable of peak spectra acquisition, these authors had to perform multiple runs of the same sample at different wavelengths and draw absorbance profiles from the comparison of the peak heights in the individual electropherograms. More modem instrumentation fitted with diode array or fast-scanning UV detectors would facilitate analyses of this type. 

The advent of low priced powerful microcomputers has enabled easy processing of spectrophotometric data. In addition, fast scanning detectors have allowed elimination of errors resulting from poor time stability when a dual wavelength method is used with a single wavelength spectrophotometer. Modem multiwavelength analysis utilizes the reversed matrix representation of Beer-Lambert s law. This can be combined with chemometric techniques such as principal component analysis (PCA) and partial least squares (PLS). 

PDA detection, however, can provide much more information than simple multiwavelength analysis. Despite the lack of distinguishing minima and maxima, protein UV spectra are more information rich than is apparent to the eye. Characteristic spectra in the 250-300 nm range have been useful in numerous studies of protein aromatic content. 

Walters 1986 Multiwavelength analysis of differentiated spectra of urinary porphyrins 303... 

Table 5-41. Other spectroscopic methods Multiwavelength analysis, NMR, raman spectroscopy, and surface photovoltage spectroscopy. ...

Equation (8.8a) and Equation (8.8b) display many characteristics that are important in understanding what is going on because they represent the prototype equations for multiwavelength analysis. 

Fast-scanning PDA detectors and powerful signalprocessing techniques, such as multiwavelength analysis and derivative spectrophotometry, greatly facilitate multicomponent determinations and eliminate or reduce interferences. However, the precision is usually degraded for mixture analysis requiring multivariate techniques when compared to analyses where interferences are not observed. 

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... 

This experiment describes the application of multiwavelength linear regression to the analysis of two-component mixtures. Directions are given for the analysis of permanganate-dichromate mixtures, Ti(IV)-V(V) mixtures and Cu(II)-Zn(II) mixtures. 

Blanco and co-workers" reported several examples of the application of multiwavelength linear regression analysis for the simultaneous determination of mixtures containing two components with overlapping spectra. For each of the following, determine the molar concentration of each analyte in the mixture. 

Tam, K. Y., Takacs-Novak, K. Multiwavelength spectrophotometric determination of add dissociation constants. Part 11. First derivative versus target fador analysis. Pharm. Res. 1999, 16, 374-381. 

H. Gampp, M. Maeder, C.J. Meyer and A.D. Zuberbuhler, Calculation of equilibrium constants from multiwavelength spectroscopic data. Ill Model-free analysis of spectrophotometric and ESR titrations. Talanta, 32 (1985) 1133-1139. 

Thormartn, W., Meier, R, Marcolli, C., and Binder, F., Analysis of barbiturates in human serum and urine by high-performance capillary electrophoresis-micellar electrokinetic capillary chromatography with on-column multiwavelength detection, /. Chromatogr., 545, 445, 1991. 

Hendrickson WA, Horton JR, LeMaster DM. Selenomethionyl proteins produced for analysis by multiwavelength anomalous diffraction (MAD) a vehicle for direct determination of three-dimensional structure. EMBO J... 

Membrane-integrated proteins were always hard to express in cell-based systems in sufficient quantity for structural analysis. In cell-free systems, they can be produced on a milligrams per milliliter scale, which, combined with labeling with stable isotopes, is also very amenable forNMR spectroscopy [157-161]. Possible applications of in vitro expression systems also include incorporation of selenomethionine (Se-Met) into proteins for multiwavelength anomalous diffraction phasing of protein crystal structures [162], Se-Met-containing proteins are usually toxic for cellular systems [163]. Consequently, rational design of more efficient biocatalysts is facilitated by quick access to structural information about the enzyme. 

This is for univariate data what happens in the case of multivariate (multiwavelength) spectroscopic analysis. The same thing, only worse. To calculate the effects rigorously and quantitatively is an extremely difficult exercise for the multivariate case, because not only are the errors themselves are involved, but in addition the correlation stmcture of the data exacerbates the effects. Qualitatively we can note that, just as in the univariate case, the presence of error in the absorbance data will bias the coefficient(s) toward zero , to use the formal statistical description. In the multivariate case, however, each coefficient will be biased by different amounts, reflecting the different amounts of noise (or error, more generally) affecting the data at different wavelengths. As mentioned above, these... 

UV absorption occurs only below 220nm, thereby it is affected by the interference from mobile phase and from artifacts in complex foods. A multiwavelength UV detection has been experimented successfully for unambiguous evaluation of pantothenic acid [609]. However, UV detection presents a low sensitivity, compared to other techniques, like FLD or MS. FLD is applied by using a postcolumn derivatization. Pantothenic acid is converted to 3-alanine by hot alkaline hydrolysis and a reaction with OPA [610]. Also MS is successfully applied to increase the sensitivity of pantothenic acid analysis. 

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