A validated HPLC method with dual wavelength detection for chlorogenic acid with an internal standard in plasma and its application in pharmacokinetic studies in rats

Abstract

A validated high performance liquid chromatographic (HPLC) method with dual wavelength detection was developed and applied to the determination of chlorogenic acid in rat plasma. Elution was performed on C[sub 18] column at 27°C with acetronitrile-water (0.2% phosphoric acid, pH = 2.0) at different proportions according to a time scheduled programme and pumped at a flow rate of 1.0 mL min⁻¹. The eluent was detected at 325 nm for chlorogenic acid and 370 nm for the internal standard, kaempferol. The intra-day and inter-day precisions were better than 3.58% and 3.66%, respectively. The standard curve for chlorogenic acid was linear (r² = 0.9993) in the concentration range of 0.1-15 μg mL⁻¹. Accuracy in the measurement of samples ranged from 91.97 to 101.81%. The limit of detection and the limit of quantification for chlorogenic acid in plasma were 30 and 90 ng mL⁻¹, respectively. This assay has been successfully applied in the pharmacokinetic study of chlorogenic acid through oral and intravenous administration in rats. It showed that the low bioavailability of chlorogenic acid and an intravenous administration route is a better dosage regimen. It is a simple and sensitive analytical method with good accuracy and reproducibility. The current method also demonstrated that an internal standard with different optimal UV absorbance from that of the analyte can also be used in an analytical method.