Development and validation of a liquid chromatographic/tandem mass spectrometric method for determination of tetracycline in human plasma: application to bioequivalence study.

Author(s): Brum Junior L, Pugens AM, Pritsch MC, Mantovani PB, dos Santos MB, Manfio JL

Affiliation(s): BIOCINESE, Centro de Estudos Biofarmaceuticos, Av. Ministro Cirne Lima, 1541, 85903-590, Toledo-PR, Brazil. liberatojunior@mail.ufsm.br

Publication date & source: 2008-07, J AOAC Int., 91(4):731-8.

Publication type: Randomized Controlled Trial; Research Support, Non-U.S. Gov't

A fast, sensitive, and specific liquid chromatographic/tandem mass spectrometric method was developed and validated for determination of tetracycline in human plasma. Tetracycline and oxytetracycline [internal standard (IS)] were extracted from the plasma by protein precipitation. The mobile phase consisted of acetonitrile-formic acid 0.1% (48 + 52, v/v), run at a flow rate of 1 ml/min (split 1:5). Detection was performed by positive electrospray ionization in multiple reaction monitoring mode, monitoring the transitions 444.8 > 410.0 and 461.0 > 426.0 for tetracycline and IS, respectively. The analysis was performed in 3.5 min and the method was linear in the plasma concentration range of 50-6000 ng/mL. The mean extraction recoveries for tetracycline and IS from plasma were 92.14 and 94.04%, respectively. Method validation investigated parameters such as the linearity, precision, accuracy, specificity, and stability, giving results within the acceptable range. The proposed method was successfully applied for determination of tetracycline in human plasma samples to support bioequivalence studies.