Photoinduced covalent binding of frusemide and frusemide glucuronide to human serum albumin

Aims To study reaction of photoactivated frusemide (F) and F glucuronide (F gnd metabolite) with human serum albumin in order to find a clue to clarify a mechanism of phototoxic blisters from high frusemide dosage. Methods F was exposed to light in the presence of human serum albumin (HSA) . HSA treated with this method (TR-HSA) was characterized by fluorescence s pectroscopic experiment, alkali treatment and reversible binding experiment . Results Less 4-hydroxyl-N-furfuryl-5-sulphamoylanthranilic acid (4HFSA, a p hotodegradation product of F) was formed in the presence of HSA than in the absence of HSA. A new fluorescence spectrum excited at 320 nm was observed for TR-HSA. Alkali treatment of TR-HSA released 4HFSA. Quenching of the fl uorescence due to the lone tryptophan near the warfarin-binding site of HSA was observed in TR-HSA. The reversible binding of F or naproxen to the war farin-binding site of TR-HSA was less than to that of native HSA. These res ults indicate the photoactivated F was covalently bound to the warfarin-bin ding site of HSA. The covalent binding of Fgnd, which is also reversibly bo und to the wafarin-binding site of HSA, was also induced by exposure to sun light. Fgnd was more photoactive than F, indicating that F could be activat ed by glucuronidation to become a more photoactive compound. Conclusions The reactivity of photoactivated F and Fgnd to HSA and/or to ot her endogenous compounds may cause the phototoxic blisters that result at h igh F dosage.