pH-Dependence of photohemolysis sensitized by psoralen
A.Ya. Potapenkoa*, I. V. Belichenkoa, I. S. Mamedova, N. N. Zhuravela, A. A. Kyagovaa, G. M. J. Beijersbergen van Henegouwenb and H. de Vriesb.
aDepartment of Medical and Biological Physics, Russian State Medical University, Ostrovityanova street 1, 117869 Moscow, Russia. E-mail: firstname.lastname@example.org
bLeiden Amsterdam Center for drug Research, Department of Medicinal Photochemistry, State University of Leiden, P.O.Box 9502, 2300 RA Leiden, The Netherlands.
*To whom correspondence should be addressed.
The effect of pH on psoralen photosensitized (366 nm, 23 W/m2) hemolysis was investigated. Dose (D) dependence of the rate of photohemolysis (V) fitted Eq. V=Vo+kDx, where Vo is the rate of dark hemolysis, and k is a coefficient. Variation of pH did not influence the exponent x, which was equal to ca. two. The pH had a strong influence on the value of k. The lowest V was observed in the pH interval from 8.0 to 8.4. It increased nearly two fold while changing the pH from 8.4 to 9.0 as well as from 8.0 to 7.4. As the pH passed beyond 7.4 to lower values the rate sharply increased (ca. four times) with a pK value of 7.3. So as psoralen does not contain acid/base groups the effect of the pH scarcely could be the result of changing of photophysical properties of the sensitizer. Increase of V in the alkaline region could be attributed to more pronounced photooxidation of reduced glutathione as a substrate, while the jump of the rate at pH around 7.3 would be ascribed to titration of psoralen photoproduct. The letter hypothesis is supported by the data of HPLC analysis. Psoralen was preirradiated in ethanol and mixed in the dark with phosphate buffer at different pH, after which qualitative HPLC analysis was performed. Several pH dependent photoproducts were found and the pH titration curve of one of them resembled closely pH dependence of psoralen photosensitized hemolysis.