Nevirapine quantification in human plasma by high‐performance liquid chromatography coupled to electrospray tandem mass spectrometry. Application to bioequivalence study

Abstract

A rapid, sensitive and specific method to quantify nevirapine in human plasma using dibenzepine as the internal standard (IS) was developed and validated. The method employed a liquid-liquid extraction. The analyte and the IS were chromatographed on a C(18) analytical column, (150 x 4.6 mm i.d. 4 microm) and analyzed by tandem mass spectrometry in the multiple reaction monitoring mode. The method had a chromatographic run time of 5.0 min and a linear calibration curve over the range 10-5000 ng ml(-1) (r(2) > 0.9970). The between-run precision, based on the relative standard deviation for replicate quality controls was 1.3% (30 ng ml(-1)), 2.8% (300 ng ml(-1)) and 3.6% (3000 ng ml(-1)). The between-run accuracy was 4.0, 7.0 and 6.2% for the above-mentioned concentrations, respectively. This method was employed in a bioequivalence study of two nevirapine tablet formulations (Nevirapina from Far-Manguinhos, Brazil, as a test formulation, and Viramune from Boehringer Ingelheim do Brasil Química e Farmacêutica, as a reference formulation) in 25 healthy volunteers of both sexes who received a single 200 mg dose of each formulation. The study was conducted using an open, randomized, two-period crossover design with a 3 week washout interval. The 90% confidence interval (CI) of the individual ratio geometric mean for Nevirapina/Viramune was 96.4-104.5% for AUC((0-last)), 91.4-105.1% for AUC((0-infinity)) and 95.3-111.6% for C(max) (AUC = area under the curve; C(max) = peak plasma concentration). Since both 90% CI for AUC((0-last)) and AUC((0-infinity)) and C(max) were included in the 80-125% interval proposed by the US Food and Drug Administration, Nevirapina was considered bioequivalent to Viramune according to both the rate and extent of absorption.

Coverage of Nevirapine-Based Services to Prevent Mother-to-Child HIV Transmission in 4 African Countries

“…The limit of detection for nevirapine was set at 100 ng/mL. 16 Values detected above this limit were reflected as positive and those below as negative. Interday and intraday coefficients of variation were less than 10% for all controls.…”

Sensitive Enzyme Immunoassay for Measuring Plasma and Intracellular Nevirapine Levels in Human Immunodeficiency Virus-Infected Patients

“…Several high-performance liquid chromatographic (HPLC) assays combined with UV detection (6, 10, 22) or tandem mass spectrometry (14,24) for the quantitative determination of NVP in plasma have been described. However, these methods are characterized by a relatively high limit of quantification (10 ng ml Ϫ1 ) and by fastidious workup, thus excluding their use in the ex vivo monitoring of intracellular levels of the drug.…”

“…As a consequence, the intracellular levels may be very different from those recorded in plasma. Knowledge of the intracellular distribution may help in understanding the mechanisms that are involved in the evolution of drug resistance and the development of sanctuary sites.Several high-performance liquid chromatographic (HPLC) assays combined with UV detection (6, 10, 22) or tandem mass spectrometry (14,24) for the quantitative determination of NVP in plasma have been described. However, these methods are characterized by a relatively high limit of quantification (10 ng ml Ϫ1 ) and by fastidious workup, thus excluding their use in the ex vivo monitoring of intracellular levels of the drug.…”

High throughput LC–MS/MS method for simultaneous quantification of lamivudine, stavudine and nevirapine in human plasma

“…Several bioanalytical methods are reported to determine 3TC, d4T and NVP individually in different biological matrices like plasma [16][17][18][19][20][21][22][23][24], tissues and amniotic fluids [16], saliva [17], urine [18,20], cerebrospinal fluids [17,21] and serum [21,25]. Sensitive and selective methods [26][27][28][29][30][31][32][33][34][35][36] based on HPLC-UV and LC-MS/MS methodologies have also been developed for simultaneous analysis of these three drugs in combination with other NRTI, NNRTI or protease inhibitors as they have higher efficacy, acceptable toxicity and broader action range.…”