Formation, characterization, and toxicity of the glutathione and cysteine conjugates of toxic heptapeptide microcystins

Abstract

Microcystins LR, YR, and RR, cyclic heptapeptide hepatotoxins produced by cyanobacteria, were synthetically converted into glutathione (GSH) and cysteine (Cys) conjugates. Fast atom bombardment mass spectra showed [M + H]+ ions corresponding to GSH and Cys conjugates of microcystins LR, YR, and RR for the obtained compounds. 1H NMR spectral analyses revealed that two singlet signals of olefinic protons of N-methyldehydroalanine (Mdha) in microcystins disappeared in the conjugates, confirming that thiols of GSH and Cys added nucleophilically to the alpha, beta-unsaturated carbonyl of the Mdha moiety. On examination of the 50% lethal dose (LD50) with intravenous injection using mice, both GSH and Cys conjugates showed reduction in toxicity compared with microcystins, but their toxicity still remained. Microcystin LR and its GSH conjugate were separated and identified in a standard mixture by using a frit-fast atom bombardment liquid chromatography/mass spectrometry (Frit-FAB LC/MS) method. Obtained conjugates in the present study would be important compounds as the standard samples for study of metabolism of microcystins, and the Frit-FAB LC/MS method would be applicable to mass spectrometric identification of metabolites of microcystins.

Quantitatively evaluating detoxification of the hepatotoxic microcystin-LR through the glutathione (GSH) pathway in SD rats

“…The glutathione and cysteine conjugates of MCs (MC-GSH and MC-Cys) were firstly synthesized under chemical conditions by Kondo et al (1992) with Frit-FAB liquid chromatographymass spectrometry (LC/MS), and then the resulting conjugates were identified as two metabolites of MCs in vivo in the livers of mouse and rat. Subsequently, MC-LR-GSH was proved to form in various aquatic animals via glutathione Stransferase (GST) in vitro, and it was suggested to be the first step in the detoxification of MCs in aquatic organisms, followed by degradation to the cysteine conjugate (Pflugmacher et al 1998(Pflugmacher et al , 2001).…”

Quantitatively evaluating detoxification of the hepatotoxic microcystins through the glutathione and cysteine pathway in the cyanobacteria-eating bighead carp

“…Many studies have demonstrated that glutathione plays an important role in the detoxification of MCs in both mammals and aquatic organisms (Ito et al, 2002;Kondo et al, 1992Kondo et al, , 1996Pflugmacher et al, 1998;Zhang et al, 2009). For example, the glutathione and cysteine conjugates of MCs were identified by Frit-FAB LC/MS in liver samples of mouse and rat treated with MCs (Kondo et al, 1992(Kondo et al, , 1996.…”

Rapid quantification of total microcystins in cyanobacterial samples by periodate-permanganate oxidation and reversed-phase liquid chromatography

“…Biochemical methods such as enzyme-linked immunosorbent assay (ELISA) [18][19][20] and the protein phosphatase inhibition assay [5,21] are useful as screening methods due to their high sensitivity and high throughput, but they often have poor identification ability and the potential for false-positives. Reversedphase high-performance liquid chromatography (HPLC) with UV detection [20], diode array detection [22][23][24][25][26][27] or electrochemical detection [28], LC-mass spectrometry (LC-MS) [29][30][31], capillary electrophoresis (CE) [30], and CE-MS [30] have also been used for the identification and quantification of MCs. The LC method can be much more definitive than bioassays and biochemical methods 0003 since it can provide information for the identification of individual MCs.…”