Monitoring of anti-tuberculosis drug concentrations and dose adjustment can be helpful in cases that show poor response to treatment. Here, we describe a method that can rapidly and simultaneously measure the blood concentrations of four anti-tuberculosis drugs (isoniazid, rifampicin, pyrazinamide, and ethambutol) and two major metabolic ratios (acetylisoniazid/isoniazid and 25-desacetylrifampicin/rifampicin) using high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS). A C18 reversed-phase column and gradients of methanol in 0.3% formic acid and water were used for HPLC separation. The drug concentrations were determined by multiple reaction monitoring in positive ion mode and the assay performance was evaluated. We determined peak concentration ranges for each drug and acetylisoniazid/isoniazid and 25-desacetylrifampicin/rifampicin ratios by analyzing 2-h post-dose samples in patients treated with standard dosing as a first-line treatment. The preparation of 20 samples including two steps of deproteinization with 50% and 100% methanol was performed within 20 min and chromatographic separation was achieved within 4 min/sample. Interassay calibration variability data obtained over concentrations of 0-8 microg/mL for isoniazid and ethambutol and 0-80 microg/mL for rifampicin and pyrazinamide showed a linear and reproducible curve. Within-run and between-run imprecision (CVs) were 1.9-5.5% and 3.5-10.5% and the lower limits of detection and quantification were 0.01-0.5 microg/mL and 0.05-1.0 microg/mL, respectively. The isoniazid concentration was found to be inversely correlated to the acetylisoniazid/isoniazid ratio (R=-0.739, P<0.001). The devised method allows for the simple, rapid, sensitive and reproducible quantification of isoniazid, rifampicin, pyrazinamide, ethambutol and their two metabolic ratios and should be helpful for therapeutic drug monitoring in tuberculosis patients.

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The OPTIGEN test performed better than the MAST-CLA test for diagnosing allergic rhinitis.

We confirm that hydroxylation at the 8 position of theophylline (1,3-DMU) is significantly affected by genetic polymorphism in CYP2E1 in addition to CYP1A2.

3 Department of Laboratory Medicine, Cheju National University College of Medicine, Jeju, Korea; * address correspondence to this author at: Department of Laboratory Medicine, Seoul National University College of Medicine, 28 Yongon-dong, Chongno-gu, Seoul 110-799, Korea; fax 82-2-745-6653, e-mail jqkim@plaza. snu.ac.kr)Theophylline is metabolized to 1,3-dimethyluric acid (1,3-DMU), 3-methylxanthine (3MX), and 1-methylxanthine (1MX) mainly by CYP1A2, and 1MX is rapidly converted to 1-methyluric acid (1MU) by xanthine oxidase (1-3 ). Individuals differ in terms of their rates of theophylline metabolism and the resulting serum concentrations (1 ); moreover, some theophylline metabolites, such as 3MX, are known to have bronchodilator activity. Thus, we need to determine the plasma concentrations of theophylline and its metabolites simultaneously to ensure the safe use of theophylline, especially in patients with renal insufficiency, in whom serious side effects can occur if metabolites are allowed to accumulate.Many HPLC methods have been used to measure theophylline and its metabolites. However, most of these techniques require the avoidance of caffeine or require a longer separation time because of the interfering effect of caffeine metabolites such as 1,7-dimethylxanthine (1,7-DMX), 3,7-dimethylxanthine, and theophylline (1,3-dimethylxanthine) (4 ). Recently, liquid chromatographymass spectrometry and liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods to determine theophylline and caffeine metabolites have been developed (5, 6 ). However, these methods require two runs per sample, with negative and positive ionization, and the total HPLC run time is ϳ60 min. These methods therefore are not suitable for the high-throughput determinations of theophylline and its metabolites required for routine therapeutic drug monitoring. In the present study, we developed a simple and rapid LC-MS/MS method for the simultaneous determination of plasma theophylline and 2176 Technical Briefs