Contribution of aldehyde oxidase, xanthine oxidase and aldehyde dehydro-genase on the oxidation of aromatic aldehydes
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AbstractAliphatic aldehydes have a high affinity toward aldehyde dehydrogenase activity but are relatively poor substrates of aldehyde oxidase and xanthine oxidase. In addition, the oxidation of xenobiotic-derived aromatic aldehydes by the latter enzymes has not been studied to any great extent. The present investigation compares the relative contribution of aldehyde dehydrogenase, aldehyde oxidase, and xanthine oxidase activities in the oxidation of substituted benzaldehydes in separate preparations. The incubation of vanillin, isovanillin, and protocatechuic aldehyde with either guinea pig liver aldehyde oxidase, bovine milk xanthine oxidase, or guinea pig liver aldehyde dehydrogenase demonstrated that the three aldehyde oxidizing enzymes had a complementary substrate specificity. Incubations were also performed with specific inhibitors of each enzyme (isovanillin for aldehyde oxidase, allopurinol for xanthine oxidase, and disulfiram for aldehyde dehydrogenase) to determine the relative contribution of each enzyme in the oxidation of these aldehydes. Under these conditions, vanillin was rapidly oxidized by aldehyde oxidase, isovanillin was predominantly metabolized by aldehyde dehydrogenase activity, and protocatechuic aldehyde was slowly oxidized, possibly by all three enzymes. Thus, aldehyde oxidase activity may be a significant factor in the oxidation of aromatic aldehydes generated from amines and alkyl benzenes during drug metabolism. In addition, this enzyme may also have a role in the catabolism of biogenic amines such as dopamine and noradrenaline where 3-methoxyphenylacetic acids are major metabolites.
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CitationBeedham, C., Panoutsopoulos, G.I. and Kouretas, D. (2004). Contribution of aldehyde oxidase, xanthine oxidase and aldehyde dehydro-genase on the oxidation of aromatic aldehydes. Chemical Research in Toxicology. Vol. 17, No. 10, pp. 1368-1376.
Link to publisher’s versionhttp://dx.doi.org/10.1021/tx030059u
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Enzymatic oxidation of vanillin, isovanillin and protocatechuic aldehyde with freshly prepared Guinea pig liver slices.Panoutsopoulos, Georgios I.; Beedham, Christine (2005)Background/Aims: The oxidation of xenobiotic-derived aromatic aldehydes with freshly prepared liver slices has not been previously reported. The present investigation compares the relative contribution of aldehyde oxidase, xanthine oxidase and aldehyde dehydrogenase activities in the oxidation of vanillin, isovanillin and protocatechuic aldehyde with freshly prepared liver slices. Methods: Vanillin, isovanillin or protocatechuic aldehyde was incubated with liver slices in the presence/absence of specific inhibitors of each enzyme, followed by HPLC. Results: Vanillin was rapidly converted to vanillic acid. Vanillic acid formation was completely inhibited by isovanillin (aldehyde oxidase inhibitor), whereas disulfiram (aldehyde dehydrogenase inhibitor) inhibited acid formation by 16% and allopurinol (xanthine oxidase inhibitor) had no effect. Isovanillin was rapidly converted to isovanillic acid. The formation of isovanillic acid was not altered by allopurinol, but considerably inhibited by disulfiram. Protocatechuic aldehyde was converted to protocatechuic acid at a lower rate than that of vanillin or isovanillin. Allopurinol only slightly inhibited protocatechuic aldehyde oxidation, isovanillin had little effect, whereas disulfiram inhibited protocatechuic acid formation by 50%. Conclusions: In freshly prepared liver slices, vanillin is rapidly oxidized by aldehyde oxidase with little contribution from xanthine oxidase or aldehyde dehydrogenase. Isovanillin is not a substrate for aldehyde oxidase and therefore it is metabolized to isovanillic acid predominantly by aldehyde dehydrogenase. All three enzymes contribute to the oxidation of protocatechuic aldehyde to its acid.
Interaction of phthalazines with molybdenum hydroxylases. Phthalazine and its 1-substituted derivatives as substrates, inhibitors and inducers of aldehyde oxidase and xanthine oxidase, both in vitro and in vivo.Beedham, Christine; Stell, J. Godfrey P.; Johnson, Christine (University of BradfordPostgraduate School of Studies in Pharmaceutical Chemistry., 2009-10-02)The interaction of the 2,3-diazanaphthalene, phthalazine and its 1-substituted derivatives with the molybdenum hydroxylases, aldehyde oxidase and xanthine oxidase, has been investigated both in vivo and /Ok in vitro. Metabolic studies, carried out by treating rabbits with both cold and 14C-labelled phthalazine, have shown that this compound is extensively metabolised in vivo, the major metabolite being a glucuronide conjugate. Very little unchanged phthalazine or its molybdenum hydroxylase mediated oxidation product 1-hydroxyphthalazine were excreted in the urine. Pretreatment of rabbits with phthalazine or 1-hydroxyphthalazine had no effect upon the activity of the microsomal monooxygenases but caused a significant increase in the specific activities of both aldehyde oxidase and xanthine oxidase. Determination of the molybdenum content of purified aldehyde oxidase fractions using electrothermal atomic absorption spectroscopy has confirmed that an increase in the molybdenum content of the enzyme fraction accompanies the increase in activity. A qualitative assessment of purified aldehyde oxidase fractions using iso-electric focusing has indicated that this enzyme may be composed of 2 or 3 active variants and following pretreatment with either phthalazine or 1-hydroxyphthalazine a further band of enzyme activity is apparent on the electropherogram. The Km value for phthalazine is significantly reduced with enzyme prepared from phthalazine treated rabbits, indicating that a form of the enzyme with a high affinity for phthalazine may have been induced. 1-Hydrazinophthalazine (Hydralazine) and two other hydrazine substituted N-heterocycles, endralazine and 1-hydrazinoisoquinoline have been shown to exert a potent progressive inhibition of aldehyde oxidase in vitro, effective only in the presence of substrate, but are inactive towards xanthine oxidase. In addition, administration of hydralazine to rabbits results in a significant reduction in liver aldehyde oxidase activity. Investigations into the interaction of some of the metabolites of hydralazine with aldehyde oxidase in vitro suggest that hydralazine is also the inhibiting species in vivo.
Specificity of aldehyde oxidase towards N-heterocyclic cations. Oxidation of quinolinium and related cations by aldehyde oxidase in vitro; the isolation of two products formed simultaneously from a single substrate.Stell, J. Godfrey P.; Beedham, Christine; Taylor, Susan M. (University of BradfordPostgraduate School of Studies in Pharmaceutical Chemistry, 2009-10-02)Aldehyde oxidase catalysed oxidation of various quinolinium and related cations has been studied in vitro. Oxidation products were identified by comparison of their spectral and chromatographic characteristics with those of authentic compounds. The N-heterocyclic cations and quinolones used required synthesis. Incubation of N-methylquinolinium, N-methyl-7,8-benzoquinolinium and N-phenylquinolinium yielded the corresponding 2- and 4-quinolones simultaneously. The ratio of 2- to 4-quinolone formation was found to be species dependent; the proportion of 4-quinolone was greater with guinea pig enzyme than with rabbit enzyme. Incubation of N-methyl-4-methylquinolinium, N-methyl-4-phenylquinolinium and N-methylphenanthridinium produced the expected 2-quinolones. Cations substituted adjacent to the ring nitrogen, i. e. N-methyl-2- methylquinolinium, N-methyl-2-phenylquinolinium and N-phenyl-2-phenylquinolinium, were oxidised to the corresponding 4-quinolones. Kinetic constants were determined spectrophotometrically. The Km values obtained with rabbit enzyme ranged from 1.6 x 10-3 M for N-methylquinolinium to <10-5 M for N-phenyl-2-phenylquinolinium. Quaternary compounds were found to be better substrates than their non-quaternary counterparts, except for N-methylisoquinolinium and N-methylphenanthridinium. In general, guinea pig aldehyde oxidase was shown to have a greater affinity for N-heterocyclic cations than rabbit enzyme. The substrate binding site has been discussed in the light of the results outlined below. Oxidation of N-methyl-4-phenylquinolinium (to the 2-quinolone) was competitively inhibited by N-methyl-2-phenylquinolinium (which yields the 4-quinolone), indicating that both these cations interact at the same active site. The ratio of 2- to 4-quinolone production from N-methylquinolinium was constant under various conditions, including purification of the enzyme but changed at high pH or in the presence of N-methylphenanthridinium. Inhibition studies indicated that both quaternary and non-quaternary compounds act at the same site on the enzyme. Km and Vmax values for phthalazine, N-methyl-2-phenylquinolinium and N-methylquinolinium were determined over the pH range 5.4 to 10.2. In each case, results indicated that the enzyme has an ionisable group at the active site with a pK ca. 8. Aldehyde oxidase was shown to catalyse the dehydrogenation of the pseudobases 3,4-dihydro-4-hydroxy-3-methyl-2-quinazolinone and 3,4-dihydro- 4-hydroxy-3-methylquinazoline.