School of Pharmacy, Utrecht University, Leuvenlaan 4, 3584 CE, Utrecht, The Netherlands.
Department of Pharmacy, Radboud Institute for Medical Innovation, Radboud University Medical Centre, Geert Grooteplein Zuid 10, 6525 GA Nijmegen, The Netherlands.
J Antimicrob Chemother. 2024 Apr 2;79(4):883-890. doi: 10.1093/jac/dkae047.
To develop and validate an UPLC-MS/MS assay for simultaneous determination of the total concentration of ceftazidime, ciprofloxacin, flucloxacillin, piperacillin, tazobactam, sulfamethoxazole, N-acetyl sulfamethoxazole and trimethoprim, and the protein-unbound concentration of flucloxacillin, in human plasma to be used for research and clinical practice.
Sample pretreatment included protein precipitation with methanol. For the measurement of protein-unbound flucloxacillin, ultrafiltration was performed at physiological temperature. For all compounds, a stable isotopically labelled internal standard was used. Reliability of the results was assessed by participation in an international quality control programme.
The assay was successfully validated according to the EMA guidelines over a concentration range of 0.5-100 mg/L for ceftazidime, 0.05-10 mg/L for ciprofloxacin, 0.4-125 mg/L for flucloxacillin, 0.2-60 mg/L for piperacillin, 0.15-30 mg/L for tazobactam, 1-200 mg/L for sulfamethoxazole and N-acetyl sulfamethoxazole, 0.05-10 mg/L for trimethoprim and 0.10-50 mg/L for unbound flucloxacillin. For measurement of total concentrations, the within- and between-day accuracy ranged from 90.0% to 109%, and 93.4% to 108%, respectively. Within- and between-day precision (variation coefficients, CVs) ranged from 1.70% to 11.2%, and 0.290% to 5.30%, respectively. For unbound flucloxacillin, within-day accuracy ranged from 103% to 106% and between-day accuracy from 102% to 105%. The within- and between-day CVs ranged from 1.92% to 7.11%. Results of the international quality control programme showed that the assay is reliable.
The method provided reliable, precise and accurate measurement of seven commonly prescribed antibiotics, including the unbound concentration of flucloxacillin. This method is now routinely applied in research and clinical practice.
建立并验证一种可同时测定人血浆中头孢他啶、环丙沙星、氟氯西林、哌拉西林、他唑巴坦、磺胺甲噁唑、N-乙酰磺胺甲噁唑和甲氧苄啶的总浓度和游离氟氯西林浓度的 UPLC-MS/MS 检测方法，以供研究和临床实践使用。
样品预处理采用甲醇沉淀蛋白。对于游离氟氯西林的测定，采用超滤法在生理温度下进行。所有化合物均使用稳定同位素标记的内标。通过参加国际质量控制计划来评估结果的可靠性。
该方法按照欧洲药品管理局（EMA）指南进行验证，头孢他啶、环丙沙星、氟氯西林、哌拉西林、他唑巴坦、磺胺甲噁唑和 N-乙酰磺胺甲噁唑的浓度范围为 0.5-100mg/L， 0.05-10mg/L，0.4-125mg/L，0.2-60mg/L，0.15-30mg/L，1-200mg/L，0.05-10mg/L，甲氧苄啶的浓度范围为 0.05-10mg/L，游离氟氯西林的浓度范围为 0.10-50mg/L。对于总浓度的测定，日内和日间准确度分别为 90.0%-109%和 93.4%-108%，日内和日间精密度（变异系数，CV）分别为 1.70%-11.2%和 0.290%-5.30%。游离氟氯西林的日内准确度为 103%-106%，日间准确度为 102%-105%。日内和日间 CV 分别为 1.92%-7.11%。国际质量控制计划的结果表明，该方法可靠。
该方法可可靠、精确和准确地测定七种常用抗生素，包括游离氟氯西林的浓度。该方法现已常规应用于研究和临床实践。
