QUANTITATION OF THE SINGLET OXYGEN PRODUCED BY UVA IRRADIATION OF HUMAN LENS PROTEINS

Ultraviolet irradiation of aged human lens proteins in vitro causes ex tensive photolytic damage of His and Trp residues. Protection by sodiu m azide argues for a process mediated by singlet oxygen (O-1(2)) In th e work described here, the synthesis of O-1(2) was measured by the ble aching of N,N-dimethyl-4-nitrosoaniline (RNO), the oxidation of added histidine and the oxidation of furfuryl alcohol, To obtain a more accu rate value for O-1(2) generation, a known quantity of O-1(2) was gener ated by the thermal dissociation of 3-(4-methyl-1-naphthyl)propionic a cid endoperoxide, and the efficiency of each assay method to report on the O-1(2) generated was determined. The values obtained were 0.003 m ol of RNO bleached/mol of O-1(2) generated, 0.55 mol of furfuryl alcoh ol oxidized/mol O-1(2) and 0.5 mol of His oxidized/mol O-1(2) generate d, Irradiation of the human lens proteins,vith UVA light produced from 2.1 to 2.4 mM of O-1(2) by RNO bleaching, 2.6-2.8 mM O-1(2) by furfur yl alcohol oxidation and up to 1.9 mM of O-1(2) by histidine oxidation during a 1 h irradiation period, The average value (2.2 mM of O-1(2)) corresponds to the theoretical production of 30 nmol of singlet oxyge n at UVA light intensities equivalent to a 1 h exposure to sunlight at noon in the northern hemisphere.