Substrate specificity for the 12beta-hydroxylation of bufadienolides by Alternaria alternata.
Hydroxylation is an important route to synthesize more hydrophilic compounds of pharmaceutical significance. Microbial hydroxylation offers advantages over chemical means for its high specificity. In this study, a fungal strain Alternaria alternata AS 3.4578 was found to be able to catalyze the specific 12beta-hydroxylation of a variety of cytotoxic bufadienolides. Cinobufagin and resibufogenin could be completely metabolized by A. alternata to generate their 12beta-hydroxylated products in high yields (>90%) within 8 h of incubation. A. alternata could also convert 3-epi-desacetylcinobufagin into 3-epi-12beta-hydroxyl desacetylcinobufagin as the major product (70% yield). C-3 dehydrogenated products were detected in these reactions in fair yields, while their accumulation was relatively slow. The 12beta-hydroxylation of bufadienolides could be significantly inhibited by the substitution of 1beta-, 5-, or 16alpha-hydroxyl groups, and the 14beta,15beta-epoxy ring appeared to be a necessary structural requirement for the specificity. For the biotransformation of bufalin, a 14beta-OH bufadienolide, this reaction was not specific, and accompanied by 7beta-hydroxylation as a parallel and competing metabolic route. The biotransformation products were identified by comparison with authentic samples or tentatively characterized by high-performance liquid chromatography-diode array detection-atmospheric pressure chemical ionization-mass spectrometry analyses.
Ye
Min
M
The State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Xueyuan Road 38, Beijing 100083, PR China.
Guo
Dean
D
D000528:Alternaria / Q000378:metabolism*; D001711:Biotransformation; D002018:Bufanolides / Q000737:chemistry / Q000378:metabolism; D002770:Cholenes / Q000737:chemistry / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D006900:Hydroxylation; D007700:Kinetics; D021241:Spectrometry, Mass, Electrospray Ionization; D013379:Substrate Specificity
D002018:Bufanolides; D002770:Cholenes; C087925:bufadienolide; C005734:bufogenin; C002471:cinobufagin; C022777:bufalin