Biotransformation of nodakenin and simultaneous quantification of nodakenin and its aglycone in incubated system of human intestinal bacteria by HPLC method

Abstract

When nodakenin (1) was anaerobically incubated with human intestinal bacteria, nodakenetin (2) was found as a main biotransformed product. We developed a simple and selective reversed-phase high-performance liquid chromatographic method for simultaneous quantification of 1 and 2 in incubated system of human intestinal bacteria with 1. Chromatographic separation of 1 and 2 was performed on an analytical C₁₈ column, with a mobile phase of MeOH–H₂O (4:6, v/v) at a flow rate of 1.0 ml/min and the UV detection was at 330 nm. The calibration curves were linear over the range of 0.15–24.0 μg/ml for 1 and 0.7–13.2 μg/ml for 2. The lower limits of detection and quantification were 0.01 and 0.1 μg/ml for 1, and 0.005 and 0.05 μg/ml for 2. The recoveries were (87.66 ± 1.66), (79.89 ± 2.53), and (82.96 ± 5.61)% at 1.0, 2.0, and 8.0 μg/ml, respectively, for 1 and (88.32 ± 4.12), (78.15 ± 4.39), and (76.22 ± 3.29)% at 1.0, 4.0, and 16.0 μg/ml, respectively, for 2. The intra- and interday precision and accuracy were validated by relative standard deviation, which were in the ranges of 1.25–4.16 and 2.16–6.12% for 1, and 1.98–6.45 and 2.56–4.57% for 2, respectively. This method has been applied to the simultaneous quantitation of 1 and 2 in incubated system of human intestinal bacteria with 1.

Keywords:

• nodakenin
• nodakenetin
• biotransformation
• human intestinal bacteria
• column liquid chromatography

Acknowledgements

This research was partly supported by the National Natural Science Foundation of China (30672609), National High Technology Research and Development Program of China (2002AA2Z343C and 2004AA2Z3783), National Sciences and Technology Program of China (2006BAI06A01-02), and Beijing Municipal Special-Purpose Science Foundation of China (Z0004105040311).