Structure−Activity Study on the Quinone/Quinone Methide Chemistry of Flavonoids

Abstract

A structure-activity study on the quinone/quinone methide chemistry of a series of 3',4'-dihydroxyflavonoids was performed. Using the glutathione trapping method followed by HPLC, (1)H NMR, MALDI-TOF, and LC/MS analysis to identify the glutathionyl adducts, the chemical behavior of the quinones/quinone methides of the different flavonoids could be deduced. The nature and type of mono- and diglutathionyl adducts formed from quercetin, taxifolin, luteolin, fisetin, and 3,3',4'-trihydroxyflavone show how several structural elements influence the quinone/quinone methide chemistry of flavonoids. In line with previous findings, glutathionyl adduct formation for quercetin occurs at positions C6 and C8 of the A ring, due to the involvement of quinone methide-type intermediates. Elimination of the possibilities for efficient quinone methide formation by (i) the absence of the C3-OH group (luteolin), (ii) the absence of the C2=C3 double bond (taxifolin), or (iii) the absence of the C5-OH group (3,3',4'-trihydroxyflavone) results in glutathionyl adduct formation at the B ring due to involvement of the o-quinone isomer of the oxidized flavonoid. The extent of di- versus monoglutathionyl adduct formation was shown to depend on the ease of oxidation of the monoadduct as compared to the parent flavonoid. Finally, unexpected results obtained with fisetin provide new insight into the quinone/quinone methide chemistry of flavonoids. The regioselectivity and nature of the quinone adducts that formed appear to be dependent on pH. At pH values above the pK(a) for quinone protonation, glutathionyl adduct formation proceeds at the A or B ring following expected quinone/quinone methide isomerization patterns. However, decreasing the pH below this pK(a) results in a competing pathway in which glutathionyl adduct formation occurs in the C ring of the flavonoid, which is preceded by protonation of the quinone and accompanied by H(2)O adduct formation, also in the C ring of the flavonoid. All together, the data presented in this study confirm that quinone/quinone methide chemistry can be far from straightforward, but the study provides significant new data revealing an important pH dependence for the chemical behavior of this important class of electrophiles.

Reactions of Plant Phenolics with Food Proteins and Enzymes under Special Consideration of Covalent Bonds

“…As recently reported, o-quinone trapping method by glutathione appeared to be an accurate method for quantification of o-quinone metabolites (Awad et al, 2000;Boersma et al, 2000;Awad et al, 2001;Awad et al, 2002a,b). In these studies, the method proved useful for analysing the nature and biochemical behavior of quercetin o-quinone, and characterization of its p-quinone methide isomers (Awad et al, 2000;Boersma et al, 2000;Awad et al, 2001;Awad et al, 2002a,b). According to these authors, for quercetin o-quinone and p-quinone methide III, glutathione adducts can be formed only at C2´, C5´, and C6´, whereas for pquinone methides I and II, adduct formation at C2´, C5´, C6´, C6, and C8 is possible Awad et al, 2001).…”

“…In these studies, the method proved useful for analysing the nature and biochemical behavior of quercetin o-quinone, and characterization of its p-quinone methide isomers (Awad et al, 2000;Boersma et al, 2000;Awad et al, 2001;Awad et al, 2002a,b). According to these authors, for quercetin o-quinone and p-quinone methide III, glutathione adducts can be formed only at C2´, C5´, and C6´, whereas for pquinone methides I and II, adduct formation at C2´, C5´, C6´, C6, and C8 is possible Awad et al, 2001). Translating this information to our case and consequently in accordance to the observed decrease in the nucleophilic protein side chains, similar covalent adduct formation at the proposed C atoms for the flavonoids is possible.…”

“…The general picture emerging from these studies is that the oxidation of fla- vonoids is complex, resulting in a wide variety of, in many cases, unidentified reaction products. In the presence of glutathione, however, these products were no longer observed, formation and dimerization of glutathione adducts originating from glutathione conjugation with quercetin were observed and identified by 1 H NMR (Awad et al, 2000;Boersma et al, 2000;Awad et al, 2001;Awad et al, 2002a,b). According to these studies, the adduct formation at following sites-C2¢, C5¢, C6¢, C6, and C8 of quercetin is possible.…”

“…The different configurations of hydroxyl groups, i.e their amount and their position in the phenolic compounds, also determine the degree of protein derivatization Rawel et al, 2001a,b;Rawel et al, 2002a;Rawel et al, 2003). The dependence of the reactivity on the different configurations of hydroxyl groups in flavonoids is also demonstrated by studies using the glutathione trapping method followed by HPLC, NMR, MALDI-TOF, and LC/MS analysis to identify the different glutathionyl adducts formed (Awad et al, 2000;Awad et al, 2001). …”

Characterisation of metabolites of the putative cancer chemopreventive agent quercetin and their effect on cyclo-oxygenase activity

“…In the light of the indications of its potential nephrotoxicity demonstrated in that trial, we tested the potential of quercetin to undergo biotransformation via conjugation with glutathione (GSH) to a proximate nephrotoxicant, analogous to the nephrotoxicity associated with hydroquinone (Peters et al, 1997), bromohydroquinone (Monks et al, 1985), 17b-estradiol (Butterworth et al, 1997) or haloalkenes (Iverson et al, 1996). Previous reports have indicated that GSH conjugates are formed in vitro (Awad et al, 2000(Awad et al, , 2001Boersma et al, 2000;Galati et al, 2001). Furthermore, mindful of the fact that COX activity is a potential mechanistic target of quercetin, we compared its effect on cellular prostaglandin E-2 (PGE-2) production with that of representative quercetin metabolites.…”