Preparation of aflatoxin B1 8,9-epoxide using m-chloroperbenzoic acid

Abstract

Aflatoxin B1 is a potent carcinogen which requires activation in order to react with DNA. The exo-8,9-epoxide is putatively the active form although it has thus far eluded direct detection in biological systems. This laboratory has reported a synthesis of the epoxide using dimethyldioxirane as the oxidant [(1988) J. Am. Chem. Soc. 110, 7929-7931]. A new, very convenient synthesis is described herein in which m-chloroperbenzoic acid is used as the oxidant in a two-phase system. The reaction is carried out in CH2Cl2 in contact with an aqueous phosphate buffer (pH 7.2) to remove the byproduct m-chlorobenzoic acid, which would otherwise react with the epoxide. Excess m-chloroperbenzoic acid is removed with aqueous sodium thiosulfate.

Direct synthesis of aflatoxin B1‐N7guanine adduct: A reference standard for biological monitoring of dietary aflatoxin exposure in molecular epidemiological studies

“…The resulting MCPBA solution was passed through anhydrous sodium sulphate. Aflatoxin Bi (0-64 umoles) was dissolved in 250 ul of methylene dichloride and was converted to AFBi-8,9-epoxide by addition of 250 ul of the above MCPBA solution (4 umoles) and 500 ul of 100 mM phosphate buffer, pH 7-2 (Iyer and Harris 1993). The reaction was carried out at 5°C for lOOmin with continuous vigorous stirring in an all glass mini-reactor, connected to a refrigerated circulating bath (RCB 300, Hoeffer Scientific Company, San Francisco, USA).…”

“…Briefly, the antigen BSA-guanine-AFBi was synthesized in two steps: (i) synthesis of BSA-guanine conjugate using homobifunctional reagent, dimethyl suberimidate (Van Regen Mortal et al 1988), (ii)the BSA-guanine conjugate synthesized was further reacted with the AFBi-8,9-epoxide generated using mchloroperbenzoic acid on pure AFBi (Iyer and Harris 1993). The mole to mole ratio of BSA to guanineAFBi was determined by trinitrobenzene sulphonic acid (TNBS) assay (Sashidhar et al 1994).…”

“…However, this putative electrophile, AFBi-8,9-epoxide, has not been detected in biological systems and its role as an active intermediate has been inferred from the structure of the adducts in vivo (Essigmann et al 1977, Martin and Garner 1977, Coles et al 1980, Iyer and Harris 1993. In the recent past, the AFBi-8,9-epoxide has been successfully synthesized in vitro by chemical methods.…”

“…Two chemical oxidants, namely m-chloroperbenzoic acid. (MCPBA) and dimethyl dioxirane have been used to generate AFBi-8,9-epoxide, which was found to be stable in in vitro conditions (Baertschi et al 1988, Iyer andHarris 1993). However, it was observed that dimethyl dioxirane was highly unstable and had a low shelf-life, while MCPBA was found to be a stable oxidant to generate AFB]-8,9-epoxide (Iyer and Harris 1993).…”

“…(MCPBA) and dimethyl dioxirane have been used to generate AFBi-8,9-epoxide, which was found to be stable in in vitro conditions (Baertschi et al 1988, Iyer andHarris 1993). However, it was observed that dimethyl dioxirane was highly unstable and had a low shelf-life, while MCPBA was found to be a stable oxidant to generate AFB]-8,9-epoxide (Iyer and Harris 1993). Earlier, these oxidants had been used to synthesize CT-DNA (calf thymus DNA) and oligodeoxynucleotide adducts of aflatoxin Bi (Martin and Garner 1977, Iyer et al 1994).…”