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Quantitative determination of cyclophosphamide in rat plasma using
an on-line SPE HPLC-DAD

Xiao-Na Li, Ning Yu, Jian-Mei Zhang, Wen-Si Lin, Xiao-Mei Ling*, Ge Fu, Run-Tao Li, Jing-Rong Cui   

  1. 1. Department of Pharmaceutical Analysis, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China
    2. State Key Laboratory of Natural and Biomimetic Drugs, Peking University Health Science Center, Beijing 100191, China
  • Received:2011-12-15 Revised:2012-02-10 Online:2012-03-15 Published:2012-03-15
  • Contact: Xiao-Mei Ling*

Abstract:

A rapid and simple liquid chromatography method with on-line solid phase extraction was developed and validated for the quantitative determination of cyclophosphamide in rat plasma. The plasma sample was first extracted on an Acclaim® Polar Advantage II C18 guard column (PA II C18, 10 mm×4.6 mm, 5 μm), which was also the on-line Extraction Cartridge SPE column, by washing with 100% H2O for 1 min. The extracted sample was then eluted onto a PA II C18 column (150 mm×4.6 mm, 5 μm) and separated by isocratic elution with acetonitrile–water (40:60, v/v). The mobile phase was run at a flow rate of 1.0 mL/min, and the UV detector was set at 195 nm. Retention time of cyclophosphamide was 4.3 min and the total run-time was 6 min. The linear range of the standard curve was from 1.0 to 200 μg/mL (r2 = 0.9999), and the limits of quantification and detection were 1.0 μg/mL (RSD<10%, n = 5) and 0.3 μg/mL (RSD<13%, n = 5), respectively. Both intra- and inter-day variations were less than 5.6%. The developed method can be used for the therapeutic drug monitoring of cyclophosphamide in the clinic.

Key words: On-line SPE, HPLC-DAD, Cyclophosphamide, Rat plasma

CLC Number: 

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