On-line solid-phase extraction-HPLC-fluorescence detection for simultaneous determination of puerarin and daidzein in human serum.
Response surface methodology (RSM) was applied to the optimization of on-line solid-phase extraction (SPE) parameters, and an automated system of on-line SPE coupled with high-performance liquid chromatography (HPLC) with fluorescence detection was developed for the determination of puerarin and daidzein in human serum. The human serum sample of 50 microL was injected into a conditioned C18 SPE cartridge, and the matrix was washed out with acetonitrile-KH(2)PO(4)-triethylamine buffer (0.01 M, pH 7.4) (3:97, v/v) for 3 min at a flow rate of 0.25 mL/min. Then the target analytes were eluted and transferred to the analytical column. A chromatographic gradient elution was programmed with the mobile phase consisting of acetonitrile and KH(2)PO(4)-triethylamine buffer, and the analytes were determined with a fluorescence detector at excitation wavelength of 350 nm and emission wavelength of 472 nm, respectively. The proposed method presented good linear relations (0.85-170 microg/mL for puerarin and 0.2-40 microg/mL for daidzein), satisfactory precision (RSD<8%), and accredited recovery (92.5-107.8%).
Liu
Ying-Kun
YK
Key Laboratory of Analytical Chemistry for Life Science of Shaanxi Province, School of Chemistry and Material Science, Shaanxi Normal University, Xi'an 710062, China.
Jia
Xiao-Yan
XY
Liu
Xiao
X
Zhang
Zhi-Qi
ZQ
D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D007529:Isoflavones / Q000097:blood*; D012015:Reference Standards; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence / Q000379:methods*
D007529:Isoflavones; C004742:daidzein; C033607:puerarin