HPLC fingerprint of Cnidii Fructus formula granule and determination of six main coumarin components / 中草药

ABSTRACT

Objective: To establish an HPLC fingerprint of Cnidii Fructus formula granule analysis method for simultaneous determination of six main coumarin components, including osthol, xanthotoxin, xanthotol, bergapten, imperatorin and isopimpinellin, in order to provide reference for the study of the material basis of Cnidii Fructus formula granule. Methods: The method was performed by high performance liquid chromatography with a Waters XBridge C18 (250 mm × 4.6 mm, 5 μm) column and methanol (A)-0.1% acetic acid (B) as the mobile phase for gradient elution. The flow rate was 0.5 mL/min, the injection volume was 10 μL and the column temperature was 40 ℃. The detection wavelength was set at 320 nm. The chromatographic fingerprint evaluation system published by the State Pharmacopoeia Commission (2012 Edition) was used to establish the fingerprint of Cnidii Fructus formula granule, and the content of six main coumarin components was simultaneously determined. Results: The research on the 18 batches of Cnidii Fructus formula granule showed that the fingerprint similarity was greater than 0.992 and 19 common peaks were calibrated with satisfied peak resolution. The content determination results showed that the content of both xanthotoxin and osthol were the main coumarin components in Cnidii Fructus formula granule. According to the methodological investigation, the precision RSD values were all less than 1.6%. The sample was stable within 48 h and this method had good repeatability. The average recovery rates of xanthotol, xanthotoxin, imperatorin, isopimpinellin, bergapten and osthol were 100.69%, 101.03%, 99.48%, 100.88%, 101.27% and 100.35%, respectively. All of these coumarin components’ RSD were less than 2.5%. The six components showed a good linear relationship within a certain concentration range. The results of the content determination of xanthotol, xanthotoxin, isopimpinellin, bergapten, imperatorin and osthol respectively were 8.01-8.29, 2.37-2.63, 4.30-4.61, 4.04-4.40, 3.45-3.90 and 6.02-6.80 mg/g among the 18 batches of the Cnidii Fructus formula granule. Conclusion: The fingerprint method and the determination method of six main coumarin components in the Cnidii Fructus formula granule established in this study are simple, stable, accurate and reliable. This method can be used for the quality control of the Cnidii Fructus formula granule.