Influence of prior complex formation on the photoaddition of chlorpromazine to calf thymus deoxyribonucleic acid

Abstract

Chlorpromazine (CPZ), 2-chloro- N-(3-dimethylaminopropyl) phenothiazine, causes cutaneous photosensitivity in man. The photoaddition of CPZ to deoxyribonucleic acid (DNA) may be an important mechanism for the phototoxicity. We have investigated the complexes formed between CPZ and calf thymus DNA prior to irradiation, related their formation to the photoaddition of CPZ to DNA and initiated studies to identify the photoadducts. In the presence of high concentrations of double-stranded DNA, the CPZ absorption maximum shifted from 305 to 340 nm with an isosbestic point at 323 nm. The CPZ fluorescence at 460 nm was quenched a maximum of 90%. The excitation and emission spectra for the unquenchable fluorescence are the same as those for free CPZ. These results together with those from flow dichroism measurements indicated that CPZ formed two complexes with double-stranded DNA. One complex involves intercalations, is non-fluorescent and absorbs at 345 nm. The second complex absorbs at 310 nm, fluoresces at 460 nm and has the phenothiazine ring parallel to the DNA axis. Non-covalent binding of CPZ to heat-denatured DNA did not shift the CPZ absorption spectrum but quenched 65% of the CPZ fluorescence. One complex between CPZ and denatured DNA will account for these results. CPZ photolysis was inhibited compared with that of free CPZ by binding to double-stranded DNA (more than 98%) or denatured DNA (65%). CPZ photoadded ten times more efficiently to denatured DNA than to double-stranded DNA. These results indicate that CPZ photolysis and photoaddition are quenched in the intercalation complex. The photoaddition to double-stranded DNA does not result from intercalated CPZ because the action spectrum maximized at 310 nm rather than at 340 nm.