Conventional HPLC Method Used for Simultaneous Determination of the Seven HIV Protease Inhibitors and Nonnucleoside Reverse Transcription Inhibitor Efavirenz in Human Plasma

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  • Takahashi Masaaki
    Department of Pharmacy, National Hospital Organization Nagoya Medical Center (Tokai Area Central Hospital for AIDS Treatment and Research) Clinical Research Center, National Hospital Organization Nagoya Medical Center (Tokai Area Central Hospital for AIDS Treatment and Research)
  • Yoshida Masao
    Department of Pharmacy, National Hospital Organization Nagoya Medical Center (Tokai Area Central Hospital for AIDS Treatment and Research)
  • Oki Tsuyoshi
    Department of Pharmacy, National Hospital Organization Nagoya Medical Center (Tokai Area Central Hospital for AIDS Treatment and Research)
  • Okumura Naoya
    Department of Pharmacy, National Hospital Organization Nagoya Medical Center (Tokai Area Central Hospital for AIDS Treatment and Research)
  • Suzuki Tatsuo
    Department of Pharmacy, National Hospital Organization Nagoya Medical Center (Tokai Area Central Hospital for AIDS Treatment and Research)
  • Kaneda Tsuguhiro
    Clinical Research Center, National Hospital Organization Nagoya Medical Center (Tokai Area Central Hospital for AIDS Treatment and Research)

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We developed a simple HPLC method for the simultaneous quantitative determination of seven HIV protease inhibitors: amprenavir (APV), atazanavir (ATV), indinavir (IDV), lopinavir (LPV), nelfinavir (NFV), ritonavir (RTV), saquinavir (SQV), and a nonnucleoside reverse transcription inhibitor, efavirenz (EFV). This method involves a rapid liquid–liquid drug extraction from plasma, the use of an isocratic elution on a reversed-phase C18 column, and an ultraviolet detection at a single wavelength (205 nm). The mobile phase consisted of 39% 50 mM phosphate buffer (pH 5.9), 22% methanol and 39% acetonitrile. Forty-eight samples could be measured in one day since the runtime of one sample is 30 min. The assay has been validated over a concentration range of 0.05 to 12.20 μg/ml for APV, 0.09 to 12.05 μg/ml for ATV, 0.05 to 12.01 μg/ml for IDV, 0.12 to 12.36 μg/ml for LPV, 0.18 to 12.20 μg/ml for NFV, 0.12 to 12.33 μg/ml for RTV, 0.12 to 12.06 μg/ml for SQV, and 0.05 to 12.17 μg/ml for EFV. Calibration curves were linear in the described concentration ranges. The average accuracy ranged from 97.2 to 106.8%. Both the interday and intraday coefficients of variation for all drugs tested were less than 8.5%. This method provides a simple, accurate, and precise method for the therapeutic drug monitoring of the seven protease inhibitors and EFV in clinical routine use.

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