Grape seeds accumulate in huge quantities as byproduct during wine production and are therefore a cheap source for pharmacologically active agents. However, studies prove poor antibacterial activity, and results of analyses are sometimes contradictory. The aim of this study was, thus, to determine the antibacterial activity of grape seed extracts with special focus on the chromatographic characterization of active fractions. In the course of these investigations, extraction protocols were optimized so that microwave-assisted extraction (MAE) guaranteed highest preconcentration efficiency. Proanthocyanidins, monomeric flavonoid aglycones, as well as some of their glycosides could be identified within yielded extracts via high-performance liquid chromatography-mass spectrometry (HPLC-MS). By that means the coherence number of possible isomers of procyanidins was approximated by a newly developed equation. As far as antibacterial activity determined via screening tests is concerned, the extracts generally have been found to be positively responsive toward 10 different gram-positive and gram-negative bacteria strains. After fractionation of the raw extracts, proanthocyanidins P2, P3, P4 and gallate esters P2G and P3G (P = proanthocyanidin consisting of catechin and epicatechin units, n = oligomerization degree, G = gallate ester) were determined as active antibacterial agents toward 10 different pathogens. Only moderate activity was found for monomeric flavonoid fractions.

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Epidemologic studies have shown inverse correlation between the consumption of carotenoid-rich vegetables and the incidence of cancer. Therefore, analytical techniques for the quantitative determination of carotenoids in complex sample matrices are important. The most used method is reversed-phase (RP)-high-performance liquid chromatography (HPLC). In this study, seventeen mobile-phase systems described in the literature and six RP-HPLC columns with differences in particle size and porosity are evaluated. Derived from these results, a new mobile-phase (acetonitrile, methanol, chloroform, and n-heptane) including solvent modifiers is presented, which allows an improved and more efficient separation of carotenoids. From all columns tested, the best chromatographic parameters are found using a silica C18 column (250 x 2 mm, 5 microm, 100 A). As was found, absorbance detection at 450 nm allows the determination of the carotenoids down to the picogram range with good linearity (R2 > 0.98). For the identification and quantitation of carotenoids in complex sample matrices (containing additionally other ultraviolet-absorbing compounds), the optimized RP chromatographic system is coupled to a mass spectrometer (MS) using an atmospheric pressure ionization interface. The calibration plots show high linearity (R2 > 0.99), and the detection limit is found in the lower nanogram range. Furthermore, collision-induced dissociation in the ion source allows for the identification of carotenoids by their characteristic fragmentation pathways. In this study, a total of nine species of vegetables commonly consumed in Central Europe are analyzed for their contents of carotenoids (namely lutein, zeaxanthin, beta-cryptoxanthin, and beta-carotene) by RP-HPLC and RP-HPLC-MS-MS. It is found that good sources for lutein are spinach, kale, and broccoli, and sources for beta-carotene are broccoli, spinach, kale, carrots, and tomatoes. This new method is an improvement for the identification and quantitation of carotenoids in complex biological tissues.

Components of green tea ( Camellia sinensis) have been of considerable interest in recent years because of their potential utility as pharmaceutical agents, particularly for their antioxidant and anticarcinogenic activity. Responding to the increasing scientific validation of numerous health benefits of tea, a comprehensive approach was adopted to carry out analysis for the quality assessment of flavonoids in tea samples of different origins. For this purpose, extraction, separation, and mass spectrometric parameters were optimized. Extraction methods evaluated include reflux extraction, a modified accelerated solvent extraction (ASE), namely, Aquasolv extraction, and microwave-assisted extraction (MAE) using different percentages of solvents. Separation was performed by a specifically developed reversed phase high-performance liquid chromatography (RP-HPLC) method using different C18 and C8 stationary phases. Optimization of extraction techniques clearly proved the performance of MAE, which delivered highest yields in a very short time. Additionally, the comparison with Aquasolv extraction provided new insights, as variations in quantified amounts of target compounds between the extracts could be explained on the basis of thermal degradation and epimerization phenomena. Especially the epimerization phenomenon for catechin/epicatechin oligomers, that is, of procyanidins P 2 and P 3, was observed for the first time. Finally, an optimized extraction and separation system was used for qualitative and quantitative investigations of compounds from different green tea samples from Ceylon (cultivated under biologically controlled conditions), Japan, India, and China as well as from one black tea sample from India.