Application of high-performance liquid chromatography on-line with ultraviolet/visible spectroscopy in food science
2006
Minkiewicz, P. | Dziuba, J., E-mail: [email protected] | Darewicz, M. | Nalecz, D. (University of Warmia and Mazury, Olsztyn (Poland). Dept. of Food Biochemistry)
The aim of the present review was to describe the application of diode-array detectors for the identification of HPLC-separated chemical compounds on the basis of their UV/Vis spectra. Diode-array detectors serve to identify compounds containing conjugated double bonds, especially aromatic compounds, and to evaluate their purity. This group of compounds includes: peptides and proteins (in that case aromatic amino acid residues are chromophores), anthocyanins and other flavonoids, other phenolic compounds, carotenoids and other dyes. The most simple strategy of interpreting UV/Vis spectra is to determine the location of absorbance maximum in zero order spectra (such a strategy is usually applied in dye analysis). The absorbance ratio at different wavelengths was used to identify and investigate structural changes in proteins and peptides. Chemometrical analysis of zero order spectra provides a basis for evaluating peak purity and choosing the background spectrum. Derivatives of spectra may be characterized via determination of convexity intervals, location of minima of second derivative, calculation of amplitudes of second and third derivatives or similarity indices between derivatives of individual spectra. The possibilities offered by UV/Vis spectroscopy cover, among others, discrimination between spectra of proteins with different contents of aromatic amino acids (mainly tyrosine and tryptophan), as well as discrimination between spectra of anthocyanins containing the same chromophore but different glycosidic moieties
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