Purpose: To determine the ability of the aqueous humor to generate and degrade H2O2, the effect of environmental factors such as oxygen tension and temperature, and the constituents contributing to the observed results.
Methods: Aqueous humor was carefully removed from bovine eyes within 3 hours of death. Standard and new techniques were used to follow H2O2 degradation and formation. Catalase activity was measured by H2O2 decomposition, usually at 100 microM and 10 mM H2O2, and in some cases by O2 generation.
Results: Bovine aqueous humor generated and degraded H2O2. The generation of H2O2 was minimal at 0 degrees C but increased as temperature increased, so that at 37 degrees C at 3 hours, approximately 90 microM H2O2 was observed. Degradation was more complex. At 0 degrees C, only a slow rate of degradation was observed. At 25 degrees C, it was more rapid, and a steady state between generation and degradation was observed at approximately 30 microM. However, at 37 degrees C, starting with 100 microM H2O2, degradation was initially rapid, but then generation became predominant and H2O2, concentration increased to more than 100 microM in 3 hours. No H2O2 was generated in the absence of O2, and H2O2 production increased with increasing O2 tension. Both low and high molecular weight components contributed to the degradation, but synthesis was completely dependent on low molecular weight constituents. Ascorbic acid and metal ions such as Cu+ made a major contribution to H2O2 production. Catalase may be the macromolecular component responsible for aqueous H2O2 decay, as evidenced by H2O2 degradation, inhibition by boiling or 3-aminotriazole, and the approximate correspondence between oxygen generation and H2O2 degradation.
Conclusions: The results indicate that the aqueous humor is capable of producing levels of H2O2 that have been shown previously to cause cataract in organ culture. Changes in aqueous humor metal ion content and concentration of oxygen level have profound effects on H2O2 concentration and may effect lens viability. The variation in published H2O2 levels may in part be the result of the conditions under which the aqueous humor was obtained, stored, and used for assay. The observed steady state H2O2 concentration of 1 microM in fresh aqueous from bovine eyes must be maintained by the metabolism of surrounding tissues as well as intrinsic components capable of degrading H2O2.