Influence of Laccase and Tyrosinase on the Antioxidant Capacity of Selected Phenolic Compounds on Human Cell Lines.

Polyphenolic compounds affect the color, odor and taste of numerous food products of plant origin. In addition to the visual and gustatory properties, they serve as radical scavengers and have antioxidant effects. Polyphenols, especially resveratrol in red wine, have gained increasing scientific and public interest due to their presumptive beneficial impact on human health. Enzymatic oxidation of phenolic compounds takes place under the influence of polyphenol oxidases (PPO), including tyrosinase and laccase. Several studies have demonstrated the radical scavenger effect of plants, food products and individual polyphenols in vitro, but, apart from resveratrol, such impact has not been proved in physiological test systems. Furthermore, only a few data exist on the antioxidant capacities of the enzymatic oxidation products of phenolic compounds generated by PPO. We report here first results about the antioxidant effects of phenolic substances, before and after oxidation by fungal model tyrosinase and laccase. In general, the common chemical 2,2-diphenyl-1-picrylhydrazyl assay and the biological tests using two different types of cell cultures (monocytes and endothelial cells) delivered similar results. The phenols tested showed significant differences with respect to their antioxidant activity in all test systems. Their antioxidant capacities after enzymatic conversion decreased or increased depending on the individual PPO used.

Allosteric models for multimeric proteins: oxygen-linked effector binding in hemocyanin.

In many crustaceans, changing concentrations of several low molecular weight compounds modulates hemocyanin oxygen binding, resulting in lower or higher oxygen affinities of the pigment. The nonphysiological effector caffeine and the physiological modulator urate, the latter accumulating in the hemolymph of the lobster Homarus vulgaris during hypoxia, increase hemocyanin oxygen affinity and decrease cooperativity of oxygen binding. To derive a model that describes the mechanism of allosteric interaction between hemocyanin and oxygen in the presence of urate or caffeine, studies of oxygen, urate, and caffeine binding to hemocyanin were performed. Exposure of lobster hemocyanin to various pH values between 7.25 and 8.15 resulted in a decrease of p50. In this pH interval, p50 decreases from 95 to 11 Torr without effectors and from 49 to 6 Torr and from 34 to 5 Torr in the presence of 1 mM urate or caffeine, respectively. Thus, the allosteric effects induced by protons and urate or caffeine are coupled. In contrast, isothermal titration calorimetry did not reveal any differences in binding enthalpy (DeltaH degrees ) for urate or caffeine under either normoxic or hypoxic conditions at different pH values. Despite these apparently conflicting results, they can be explained by the nested MWC model if two different types of modulator binding sites are assumed, an allosteric and a nonallosteric type of site. Simulations of in vivo conditions with this model indicate that the naturally occurring modulator urate is physiologically relevant in H. vulgaris only during hypoxic conditions, i.e., either during environmental oxygen limitation or extensive exercise.

Effects of ultrahigh dilutions of 3,5-dichlorophenol on the luminescence of the bacterium Vibrio fischeri.

There is a great need for research in the field of homeopathy for laboratory test systems to investigate the actions of ultrahighly diluted biological effectors. With this in mind, we used the luminescent bacterium Vibrio fischeri, which is used throughout the world in testing water quality. Luminescence inhibition is utilized as a test parameter for the toxicity of a sample. We used ultrahigh dilutions (UHD) of 3,5-dichlorophenol as effector and adapted the standard test procedure for water toxicity in a way that let us evaluate very minute effects. Three groups of samples were prepared and then blinded: 45 dilutions of 3,5-dichlorophenol in steps of 10, starting with 4.2 x 10(-2) M, with vigorous shaking between dilution steps; 45 identical dilutions of 3,5-dichlorophenol without vigorous shaking; and 49 control samples of the diluent. The results of, and the discussion based on, a thorough statistical analysis led to the conclusion that an effect based on UHD, which results in an inhibition of luminescence of less than 1.5%, can be confirmed for some of the potency samples. There were both effective and ineffective samples in the three sample groups. The size of the effect was very small (ca. 1.5%), though statistically significant. The number of effective samples was significantly higher among the vigorously shaken samples than among the controls and the unshaken samples (14, 6 and 7 effective samples, respectively).