A comparative study of the effect of exogenous and endogenous photostabilizers on lens crystallin photodegradation

The purpose of the present study was to determine in vitro the effect of so dium azide, ethanol, trans-beta-carotene, and the reduced form of glutathio ne on phototransformations in the lens crystallin. These photostabilizers s how a specific affinity for different kinds of free radicals. The water-sol uble protein from the cortical part of the eye was irradiated with doses of UV C ranging from 0 to 4.07 J/cm(2). Changes in the structure of the cryst allins have been monitored by steady-state absorption and fluorescence spec troscopy. Irradiation of dialyzed samples of these proteins at a wavelength of 254 nm (1.13 +/- 0.02 mW/cm(2)) caused photooxidation of aromatic resid ues; the crystallin solutions became opaque and turbid. The samples display ed increasing attenuance at a wavelength of 280 nm as photodamage proceeded . The fluorescence of tryptophan at 333 nm systematically decreased and a n ew band between 400 and 500 nm appeared during the UV C irradiation. Our re sults show that the antioxidants can protect proteins from UV C-induced pho todegradation and the protective effect is significantly dependent on their concentration in the protein solution. There are no dramatic differences i n the rates of exogenous and endogenous scavenging of generated free radica ls for all concentrations used, with rate constants varying by a factor no greater than 2. The mechanism and the rate of scavenging or quenching are d ependent on the nature of the radical species and the photostabilizer struc ture. Although this study provides evidence for free radical scavenging and protein protection, extrapolations to possible antioxidant effects in vivo must be made cautiously.