Identification of the metabolites of methylophiopogonanone A by ultra‐high‐performance liquid chromatography combined with high‐resolution mass spectrometry

Abstract

Methylophiopogonanone A (MOA) is a naturally occurring homoisoflavonoid from the Chinese herb Ophiopogon japonicus, which has been demonstrated to attenuate myocardial apoptosis. However, the metabolism of MOA remains unknown. The goal of the present work was to investigate the in vitro metabolism of MOA using liver microsomes and hepatocytes. Methods:The metabolites were generated by incubating MOA with rat, monkey and human liver microsomes or hepatocytes. The resulting samples were analyzed by using a quadrupole-orbitrap high-resolution mass spectrometer. The metabolites were identified through the measurements of the exact mass, elemental composition and product ions.Results: A total of 15 metabolites were detected and identified. Among these metabolites, M7 (demethylenation) was the most abundant metabolite in liver microsomes, while M6 (hydroxylation) was the predominant metabolite in hepatocytes, and glucuronidation metabolites (M9 and M10) were also the main metabolites in hepatocytes. The metabolic pathways of MOA included hydroxylation, demethylenation, glucuronidation, methylation, sulfation and glutathione conjugation.Conclusions: This study for the first time provides valuable data on the metabolites of MOA, which will be of great importance for a better understanding of its disposition and to predict human pharmacokinetics.

A high‐resolution accurate mass approach to identification of graveoline metabolites using ultra‐high‐performance liquid chromatography combined with a photo diode array detector and quadrupole/time‐of‐flight tandem mass spectrometry

“…The major soft spots appear to be the methylenedioxyphenyl moiety of the parent compound, which is susceptible to demethylenation, forming the catechol derivative. The catechol derivative undergoes further oxidation producing the reactive ortho ‐quinone intermediate (Wu et al, 2022), resulting in some safety concerns with regard to the usage of graveoline and graveoline‐containing medicinal herb. It is well known that methylenedioxyphenyl can be metabolized into carbene intermediate, which can interact with P450 enzymes and inactivate the enzymes, leading to mechanism‐based inactivation.…”

“…This product ion further produced the product ion at m/z 369.0910 (elemental composition: C 19 H 17 N 2 O 4 S, 1.6 ppm) by the cleavage of the glycinyl moiety (−75.0319 Da). The neutral loss of glycinyl and glutamyl moieties was the characteristic fragmentation of GSH‐associated metabolites (Wen & Zhu, 2015; Wu et al, 2022).…”

“…It fragmented to yield the main product ion at m/z 296.0923 (elemental composition: C 17 H 13 NO 4 , 2 ppm), which coincides with the cleavage of glucuronyl moiety (−176.0322 Da) (Figure 4b). This is an indicative fragmentation of glucuronidated metabolite (Wu et al, 2022).…”

“…Table 1 presents (Figure 4b). This is an indicative fragmentation of glucuronidated metabolite (Wu et al, 2022). F I G U R E 5 LC-UV chromatograms (λ = 300-360 nm) of graveoline and its metabolites from recombinant human P450 enzymes M12 was a hydroxylated metabolite, which was found in both liver microsomes and hepatocytes.…”

“…The major soft spots appear to be the methylenedioxyphenyl moiety of the parent compound, which is susceptible to demethylenation, forming the catechol derivative. The catechol derivative undergoes further oxidation producing the reactive ortho-quinone intermediate (Wu et al, 2022), resulting in some safety concerns with regard to the usage of graveoline and graveoline-containing…”