高效液相色谱-质谱联用法测定血浆中异烟肼及其代谢物浓度的研究OA北大核心CSTPCD

Objective:To establish a method for simultaneous determination of isoniazid and its metabolites—acetylhydrazine and hydrazine in the plasma of pulmonary tuberculosis patients using high-performance liquid chromatography-mass spectrometry(HPLC-MS/MS).Methods:Plasma samples were collected from 104 confirmed pulmonary tuberculosis patients(all receiving standardized anti-tuberculosis treatment,taking isoniazid at a dose of 300-500 mg once a day,and venous blood was collected 2 hours after medication)from Beijing Chest Hospital,Capital Medical University.After protein precipitation in methanol,plasma samples were subjected to derivatization using isoniazid-D4 as the internal standard and p-methylbenzaldehyde as the derivatization reagent.The analyte was separated using an InfinityLab Poroshell 120 HILIC-Z column(100 mm×2.1 mm,2.7 μm).The binary elution system(A phase was an aqueous solution containing 0.1％formic acid and 5 mmol ammonium formate;B phase was acetonitrile)was used for gradient elution,with a column temperature of 30 ℃ and a flow rate of 0.4 ml/min);electro spray ionization positive ion mode detection and multi-reaction monitoring mode scanning was adopted,and the operation time was 8 min.Results:This determination method was not affected by endogenous substances in plasma.Isoniazid,acetylhydrazine,and hydrazine had good linearity in the range of 50-6000,25-3000,and 1-120 ng/ml,respectively(determination coefficients(R2)＞0.99);intra-day and inter-day precision＜15％;absolute accuracy value＜15％;the extraction recovery rates were all above 85％;the matrix effect ranges from 85％to 115％;the stability was good.The method was applied to detect the concentrations of isoniazid,acetylhydrazine,and hydrazine in 104 patients with pulmonary tuberculosis,which were(2402.33± 1248.57)ng/ml,(1902.51±596.82)ng/ml,and(26.50±17.13)ng/ml,respectively.Conclusion:The HPLC-MS/MS method established in this study was easy to operate,rapid,specific,and sensitive.It could be applied to monitor the therapeutic drug concentration of isoniazid and its metabolites(acetylhydrazine and hydrazine)in patients with pulmonary tuberculosis.