Novel bio-analytical technique for estimation of gamma oryzanol in rat plasma and brain homogenate using HPLC

Gamma oryzanol, a component of rice bran oil is used for its anticancer and antihyperlipidemic properties. Bioanalytical method for rat plasma and brain homogenate was developed by HPLC system with a PDA detector in which drug elution was performed using C-18 column (4.6mm×150cm, 5μ) with 1% acetic acid in methanol: acetonitrile (65/35, v/v) as mobile phase at 1.2ml/min flow rate and detected at 326nm wavelength. Liquid liquid extraction method was chosen for extraction of oryzanol from plasma as well as brain homogenate as it provided highest recovery (95% in plasma, 85% in brain homogenate). Various extraction solvents for each body fluid were analysed, out of which highest recovery for plasma (95%), in acetone: IPA (1/1, v/v) and for brain homogenate (85%) in isopropyl alcohol (IPA) was observed. Observed linearity was between 500ng/mL-5000ng/mL. The interday and intraday precision (i.e. %RSD) was less than 10% and accuracy was±5%. Selectivity and matrix effect was checked and found as per USFDA criteria. Plasma samples were found to be stable over the analysis period, HQC samples were stable up to third cycle in freeze and thaw stability while LQC samples were stable over fourth cycle. The method proved to be simple, useful and is appropriate, for preclinical and experimental research.