Qualitative and quantitative analysis of grape seeds by HPLC-MS

Seeds mainly consist of 35% crude fibres, 29% nitrogen free compounds (e.g. polyphenols), 15% crude lipids, 11% crude proteins, 3% ash and 7% water [1]. Especially the class of polyphenolic compounds presents an interesting source for pharmacologically active plant ingredients. The heterogeneity of phenolic compounds in the sample as well as the similarity of some compounds poses a challenge to every analyst. In literature catechin and epicatechin as well as their oligomers already are described in grape seeds [2, 3].

The aim of this work was extraction, separation and qualitative as well as quantitative analysis of phenolic compounds of grape seeds. Extraction was optimized on basis of different extraction methodologies as well as on the basis of diverse solvents. Variable forms of stationary phases were tested for their separation efficiency. Therefore not only conventional carrier material but also monolithic systems in capillaries and columns were evaluated.

The results of these experiments clearly show that microwave extraction results in highest yields of phenolic compounds in connection with short duration time possible. A mixture of methanol water (50/50 v/v) was chosen as extraction solvent. Monolithic carrier systems show higher separation efficiency in comparison to other stationary phases. The separation efficiency of a new carrier material on basis of monolithic poly(p-methylstyrene-co-1,2-bis(p-vinylphenyl)ethane [4] in capillaries can be impressively demonstrated by separation of small molecules. Qualitative analysis of grape seeds allows the identification of monomers, oligomers (e.g., stilbene-oligomers) and of glycosidic flavonoids. Finally due to the lack of commercially available standards the importance of mass spectrometry is pointed out.

References: 1. Dietrich, H. (2005), Der Deutsche Weinbau, pp. 34–39. 2. Gabetta, B., Fuzzati, N. (2000), Fitoterapia, 71: 162–175. 3. Peng, Z., Hayasaka, Y. (2001), J. Agric. Food Chem.) 49: 26–31. 4. Bonn, G. K., Lubbad, S., Trojer, L. (2005), patent pending.