Interaction of Sanguinarine Alkaloid, Isolated From Argemone Oil, With Hepatic Cytochrome P450 in Rats

ABSTRACT

Prior studies have shown that argemone oil (AO), responsible for ‘Epidemic dropsy’, causes inhibition of catalytic activities of Cytochrome P450 (P450). In this study interaction of sanguinarine (SAN) alkaloid, isolated from AO, with rat hepatic P450 was investigated. Hepatic microsomes prepared from 3-methylcholantherene (3MC) treated rats when incubated with SAN (1–3 μM) resulted in a spectral peak at 385 nm and a trough at 415 nm, indicative of Type I binding. Incubation of SAN (50–200 μM) with hepatic microsomes prepared from phenobarbitone (PB) treated rats also showed a Type I spectra with a peak at 395 nm and a trough at 420 nm. Relative binding efficiency (ΔA_max/K_s^app factor) of SAN with P450 was found to be 1540 and 1030 absorbance units/nmol CYP/M for 3MC and PB induced microsomes, respectively. In a P450 spectral inhibition study SAN showed higher affinity towards 3MC eliciting inhibition at much lesser concentrations (0.25–5 μM) as compared to PB (100–300 μM). The IC50s of SAN with different catalytic markers of P450 isoforms, i.e. ethoxyresorufin-O-deethylase (EROD) for CYP1A1, was 2.8 μM and for methoxyresorufin-O-deethylase (MROD) for CYP1A2 was 2.2 μM in 3MC induced microsomes, while benzoyloxyresorufin-O-deethylase (BROD) for CYP 2B1/1A1 showed an IC50 of 50 μM but pentoxyresorufin-O-deethylase (PROD) for CYP2B1 showed no inhibition even at higher concentrations of SAN (> 60 μM) in PB-induced microsomes. These results indicate that higher affinity of SAN binding towards the CYP1A family may have a role in SAN toxicity.

KEYWORDS:

• Sanguinarine
• Cytochrome P450
• Binding spectra
• Relative binding efficiency
• Inhibition spectra and IC50

The authors are thankful to Dr C. M. Gupta, Director, ITRC for his keen interest and assistance offered in carrying out this study. One of the authors (NPR) is thankful to the Council for Scientific and Industrial Research (CSIR), New Delhi, for the award of Senior Research Fellowship. We gratefully acknowledge Dr A. K. Jain, Scientist & Head, Pharmacokinetics Division, Central Drug Research Institute, Lucknow, for providing LC-MS/MS facility.

This research work has been supported by Technology Mission on Oil seed Pulses and Maize (TMOP&M), New Delhi.

The manuscript is IITR Communication #2606.

A part of this manuscript has been presented at ‘International symposium on molecular toxicology and environmental health’ held at ITRC, Lucknow from November 5–8, 2003, and has been awarded the best oral presentation award of the symposium.