The effects of ascorbic acid, iron and ADP on hyaluronic acid, a compound present in inflamed joints, were investigated in an in vitro system. Ascorbic acid induces degradation of hyaluronic acid which increased in the presence of FeCl3 and which is additionally stimulated by ADP chelated ferric ions. The hyaluronic acid degrading reactions induced by the Fe-III/ADP/ascorbic acid system were inhibited by catalase and formate to various extents whereas the presence of superoxide dismutase did not exert any inhibitory effect. Desferrioxamine, a specific iron chelator, completely inhibited hyaluronic acid depolymerisation by ascorbic acid as well as in combination with FeCl3 or FeCl3/ADP, respectively. We suggest that the ultimate hyaluronic acid degrading species is OH, generated via the Fe-III/ADP catalysed Haber Weiss reaction. There is also an indication for the involvement of perferryl or/and ferryl species in the degradation process.

Adenosine Diphosphate↗

PubMed1986

Effects of hyperthermic conditions on the reactivity of oxygen radicals.

Generation and reactivity of superoxide (O2.-) and hydroxyl (OH.) radicals in enzymatic and radiolytic systems were investigated over the temperature range from 20 degrees-50 degrees C. The generation rate and reaction kinetics of both enzymatically and radiolytically produced superoxide radicals were determined by a cytochrome c reduction assay. For OH. radical reaction studies the degradation of hyaluronic acid was assayed. An increase in temperature leads to a greater reactivity of both radicals, but in the case of an enzymatic source a disproportionate increase in the rate of generation is observed. In the pulse radiolysis system, the reactivity of superoxide radicals was found to be stimulated 15-fold over the temperature range from 20 degrees C to 60 degrees C, although the activity of superoxide dismutase was only minimally increased (about 1.6-fold). The results are discussed with respect to the possible importance of active oxygen species to the biological effects of hyperthermia.

Cytochrome c Group↗

PubMedDate not supplied

Comparison of different methods for the determination of phenylalanine hydroxylase activity in rat liver and Euglena gracilis.

Three different methods for the determination of phenylalanine hydroxylase activity have been compared: a) Differential photometric assay of the increase in tyrosine concentration in the presence of phenylalanine; b) Product separation by thin layer chromatography and scintillation counting of the [14C]tyrosine formed; c) HPLC separation and spectrofluorometric quantification of derivatized amino acids. A comparison of the activities of phenylalanine hydroxylase in rat liver and Euglena gracilis clearly showed that only rat liver contains this enzymic activity as shown by methods b) and c) although pseudo-activity of Euglena gracilis preparations was found during the spectrophotometric test a). The HPLC method proved to be the fastest, most reliable and convenient method for direct tyrosine determination and thus for measuring phenylalanine hydroxylase activity.

Animals↗

PubMedDate not supplied

Studies on the possible mechanism of inactivation of phenylalanine hydroxylase by destructive oxygen species.

The enzymic hydroxylation of phenylalanine by phenylalanine hydroxylase (E.C. 1.14.16.1.) in vitro is dependent on the presence of hydrogen peroxide removing processes. The loss of phenylalanine hydroxylase activity can be prevented to the same extent by catalase as well as the presence of optimized amounts of both peroxidase and superoxide dismutase. Peroxidase alone exhibited only two third of the maximal protective effect of catalase whereas superoxide dismutase alone was not able to exert any protective influence on phenylalanine hydroxylase. These findings suggest that the termination of phenylalanine hydroxylation in the absence of hydrogen peroxide removing reactions is probably due to destructive oxygen species generated at the active site iron of phenylalanine hydroxylase in the presence of H2O2 and the tetrahydropterin cofactor.

Animals↗