Development of spectrophotometric and chromatographic methods for the determination of flavanols in food samples

Abstract

“A-type” proanthocyanidins (PACs) are bioactive substances present in cranberries (Vaccinium Macrocarpon). They are useful to prevent urinary tract infections by inhibiting the adhesion of bacteria to the bladder wall. The bioactive behaviour is attributed to the presence of “A-type” interflavan bonds. A recent increase in cranberry consumption and the development of food supplements which praise to have a high content of “A-type” PACs opens a new field of research which consists of finding a reliable method to determine the concentration of bioactive molecules in a product. In this work, a reaction of flavanols with 4-(dimethylamino)-cinnamaldehyde (DMAC) was initially performed. Afterwards, analyses were completed using spectrophotometry and liquid chromatography. In the spectrophotometric method, advantage was taken of the fact that the condensation products of DMAC with flavanols show a maximum absorbance at 640 nm while the rest of flavonoids do not react. This leads to a selective method to identify the substances of interest and also study their chemical behaviour by modifying several variables which have an effect on the reaction. In the chromatographic method, a Kinetex C18 reversed-phase column with an UV-Vis detector was used in order to achieve a quantitative determination. With the derivatization, interferences were avoided when signals were recorded at 640 nm. Moreover, by selecting other wavelengths it was possible to study secondary peaks of PACs which did not present “A-type” interflavan bonds and peaks corresponding to the reagent as well. A process of optimization and analysis of standards was carried out to select the most adequate working conditions. Afterwards, samples of different types were studied to check if the method was satisfactory to quantify bioactive PACs.