2-[(4"-hydroxy-3′-methoxy)-phenoxy]-4-(4"-hydroxy-3"-methoxyphenyl)-8-hydroxy-6-oxo-3-oxabicylo[3.3.0]-7-octene:: unusual product of the soybean lipoxygenase-catalyzed oxygenation of curcumin

Racemic (1SR,2 SR,4SR,5SR)-2-[(4'-hydroxy-3'-methoxy)-phenoxy]-4-(4 "-3 "-methoxy-phenyl)-8-hydroxyl-6-oxo-3-oxabicyclo[3.3.0]-7-octene (2, C21H20O8) was isolated as major product of soybean lipoxygenase action on curcumin (1, C21H20O6). The structure of 2 was elucidated by HPLC-APCI-MS and tandem MS, H-1, C-13, DEFT, H,H-COSY, H,C-HMQC, H,C-HMBC and phase sensitive 2D NOESY NMR techniques. For kinetic studies the rate of substrate degradation was followed spectrophotometrically at 430 nm, and the rate of oxygen consumption was measured polarographically. As evaluated by both methods, K-m for 1 was about four times higher than that obtained for linoleic acid (as the best substrate for soybean lipoxygenase); V-max was reduced five-fold. Lipoxygenase-mediated oxygenation of 1 was confirmed by the following criteria: (i) curcumin did not react with inactivated lipoxygenase; (ii) the enzymatic reaction was strongly inhibited by inhibitors such as BHA, deferoxamine and HgCl2; (iii) oxygen consumption (measured polarographically) and curcumin degradation (measured photometrically) were shown to occur simultaneously at a ratio of 0.8 to 1, suggesting insertion of oxygen into I by lipoxygenase; (iv) molecular mass estimation by APCI-MS showed a shift of 32 in molecular mass from 1 (M-r 368) to 2 (M-r 400) being equivalent to an insertion of dioxygen. Curcumin meets none of the common features for lipoxygenase substrates and, therefore, may represent a new type of substrates for this enzyme. (C) 1998 Elsevier Science B.V. All rights reserved.