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THE EXCITATION OF 8‐METHOXYPSORALEN WITH VISIBLE LIGHT: REVERSED PHASE HPLC QUANTITATION OF MONOADDUCTS and CROSS‐LINKS

Abstract— The formation of 8‐methoxypsoralen‐DNA monoadducts and cross‐links is presumed to be responsible for the efficacy of photochemotherapies that employ 8‐methoxypsoralen activated with long‐wavelength ultraviolet radiation (UVA,320–400 nm). In this report it is shown that 8‐methoxypsoralen can also be activated with visible light (419 nm). Bovine aorta smooth muscle cells were treated with 8‐methoxypsoralen (1000 ng/mL) and 419 nm light (up to 12 J/cm2). Cellular DNA was isolated, hydrolyzed using nucleolytic enzymes and then analyzed by reversed‐phase high‐performance liquid chromatography. The primary effect of using visible light instead of long‐wavelength ultraviolet radiation is a more than 10‐fold reduction in the extent of cross‐link formation. Because the extent of monoadduct and cross‐link formation has not been routinely measured in experiments in which cellular assays have been performed, it is difficult to correlate cell response to the presence of a particular type of 8‐methoxypsoralen photoadduct (monoadduct or cross‐link). Thus, the use of visible light allows the study of cells containing nearly 100% monoadducts. In addition, the reduction in cross‐link formation when visible light is used to activate the compound may also reduce the mutagenicity of 8‐methoxypsoralen and hence enhance its therapeutic efficacy.