Analytical Department, CSIR-Indian Institute of Chemical Technology, Hyderabad, 500 007, India.
Analytical Department, CSIR-Indian Institute of Chemical Technology, Hyderabad, 500 007, India.
J Pharm Biomed Anal. 2018 Sep 10;159:200-211. doi: 10.1016/j.jpba.2018.06.050. Epub 2018 Jun 28.
Balofloxacin is a fluroquinolone antibiotic drug which has been used for the treatment of urinary tract infections (UTIs). Identification and structural characterization of metabolites is a critical component of both drug discovery and drug development research. In vivo metabolites of balofloxacin have been identified and characterized by using liquid chromatography positive ion electrospray ionization high resolution tandem mass spectrometry (LC/ESI-HR-MS/MS) experiments. To identify in vivo metabolites, blood, urine and feces samples were collected after oral administration of the drug to the female Sprague-Dawley rats (n = 3 per group). Protein precipitation, freeze liquid separation followed by solid-phase extraction methods were used for sample preparation. The extracted samples were subjected to LC-ESI/HRMS/MS analysis. The chromatographic separation of the drug and its metabolites were achieved on a XDB, C18 (50, 4.6 mm, 5 mm) column using gradient elution method in combination with 0.1% formic acid and acetonitrile at a flow rate of 0.4 mL/min. A total of 13 phase I and phase II metabolites of balofloxacin have been identified in plasma, urine and feces samples. Most of metabolites were observed in plasma and urine samples including dealkylated, desmethylated, decarbonylated, decarboxylated, hydroxylated, methylated, carboxylated, cysteine conjugated metabolites and high abundance glucuronidated metabolite. The structures of metabolites have been elucidated based on fragmentation patterns, accurate mass measurements and LC/MS/MS experiments. The main phase I metabolites of balofloxacin, decarbonylated, decarboxylated and desmethylated metabolites and phase II methylated metabolite undergo subsequent phase II glucuronidation pathways. In silico toxicity of the drug and its metabolites was determined using ProTox-II. Metabolites B-1, B-2, B-5, B-6, B-7, and B-8 to B-13 were predicted to possess immunotoxicity with high probability score. Additionally, Amine Oxidase A and Prostaglandin G/H Synthase 1 are predicted for metabolites B-1, B-3 to B-6 as toxicity targets with binding probability.
巴洛沙星是一种氟喹诺酮类抗生素药物，用于治疗尿路感染（UTIs）。鉴定和结构特征化代谢物是药物发现和药物开发研究的关键组成部分。使用液相色谱-正离子电喷雾电离高分辨串联质谱法（LC/ESI-HR-MS/MS）实验鉴定和描述了巴洛沙星的体内代谢物。为了鉴定体内代谢物，在雌性 Sprague-Dawley 大鼠（每组 3 只）口服药物后收集血液、尿液和粪便样本。采用蛋白沉淀、冷冻液分离后固相萃取方法进行样品制备。提取的样品进行 LC-ESI/HRMS/MS 分析。药物及其代谢物的色谱分离在 XDB、C18（50、4.6mm、5mm）柱上采用梯度洗脱法，结合 0.1%甲酸和乙腈，流速为 0.4mL/min。在血浆、尿液和粪便样品中鉴定出 13 种巴洛沙星的 I 相和 II 相代谢物。大多数代谢物在血浆和尿液样品中被观察到，包括脱烷基化、去甲基化、脱羰化、脱羧化、羟化、甲基化、羧化、半胱氨酸结合代谢物和高丰度的葡萄糖醛酸化代谢物。基于碎片模式、准确质量测量和 LC/MS/MS 实验，阐明了代谢物的结构。巴洛沙星的主要 I 相代谢物、脱羰化、脱羧化和去甲基化代谢物以及 II 相甲基化代谢物随后经历 II 相葡萄糖醛酸化途径。使用 ProTox-II 确定了药物及其代谢物的体内毒性。药物 B-1、B-2、B-5、B-6、B-7 和 B-8 至 B-13 的代谢物被预测具有高概率评分的免疫毒性。此外，胺氧化酶 A 和前列腺素 G/H 合酶 1 被预测为代谢物 B-1、B-3 至 B-6 的毒性靶标，具有结合概率。
