Studies on Photoconjugation of 3, 3', 4', 5-Tetrachlorosalicylanilide and Proteins

Abstract

The characteristics of photoallergy lies in the antigen formation process where light is playing some role. 3, 3', 4', 5-tetrachlorosalicylanilide (T_4CSA) has been known as one of the most potent photocontact allergens. The author studied on the photoconjugation of T_4CSA and proteins in vivo as well as in vitro system to clarify the fundamental mechanisms of the photoantigen formation. The results were as followes; 1) On thin layer chromatography at least 4 new spots of photoproducts of T_4CSA by ultraviolet ray (UV) irradiation (320-400 nm) were recognized, two of which were identified as 3', 4', 5-T_3CSA and 3', 4-DCSA from the Rf values respectively. 2) T_4CSA could combine with prealbumin, albumin and α_1globulin fraction of human serum protein by UV irradiation in vitro. It made the photoconjugate in vivo with the fraction of the guinea pig epidermal soluble protein which corresponded electrophoretically to α_1globulin of human serum. 3) The mole ratio of T_4CSA to HSA composing the photoconjugate was 1.8: 1.0. 4) The binding force of T_4CSA-HSA photoconjugate was so strong that it could not be dissociated by guanidine・HCI treatment. 5) Effects of pH and temperature on the amount of the photoconjugate in vitro suggested that the combination might be influenced not only by the conditions of T_4CSA but by that of protein. 6) To form the photoconjugate of T_4CSA and protein, they should coexist when they were irradiated. 7) After photoconjugation the fluorescence of T_4CSA-HSA photoconjugate decaied without dissociation by UV irradiation. It suggested that T_4CSA bound with HSA could lose halogen molecules by irradiation. 8) Antigen for sensitization was prepared by irradiation to the mixture of guinea pig epidermis homogenate and T_4CSA. Some of the guinea pigs which were sensitized by the injection of the antigen could show positive allergic reaction by the challenge of T_4CSA application followed by UV irradiation.