Ammari Wesam G, Al-Qadhi Zainab, Khalil Mohammad, Tayyem Rabab, Qammaz Samir, Oriquat Ghaleb, Basheti Iman A, Chrystyn Henry
1School of Pharmacy and Medical Sciences, Al-Ahliyya Amman University, Amman, Jordan.
2ACDIMA Centre for Bioequivalence and Pharmaceutical Studies, Amman, Jordan.
J Aerosol Med Pulm Drug Deliv. 2015 Jun;28(3):202-10. doi: 10.1089/jamp.2014.1153. Epub 2014 Sep 17.
Indacaterol is a novel once-a-day inhaled ultra-long-acting β2-agonist. Quantitative bioanalysis supports pharmacokinetic and clinical research. The aim of the current work was to validate an in-house developed high performance liquid chromatography (HPLC)-tandem mass spectrometry (MS/MS) analytical method for indacaterol determination in human urine samples.
A liquid-liquid extraction method has been developed to extract indacaterol from human urine samples using ethyl acetate. Indacaterol dry extract was reconstituted with 200 μL of the mobile phase (acidified water:methanol (30:70, v/v)) of which 5 μL was needed for the HPLC-MS/MS analysis. Indacaterol was eluted on a reversed C18 stationary phase with an isocratic mobile phase at a flow of 1 mL/min. Formoterol was the internal standard (IS). The MS/MS detection was employed with a turbo-ion spray ionization in the positive ion mode. A consensus of the international Guidelines for Bioanalytical Method Validation was followed.
Indacaterol was detected at a mass to charge ratio (m/z) of 393.3 and its MS/MS daughter at 173.2. The retention times of indacaterol and IS were 1.60 and 1.20 min, respectively. Validated calibration curves were linear over a range of 0.075-100 ng/mL with correlation coefficients (r)≥0.990. The curves' regression weighting factor was 1/x. Method specificity was established in six different human urine batches. No matrix interference was observed. The intra- and inter-batch precision and accuracy within±20% (at lower limit) and±15% (other quality control (QC) levels) were confirmed. The indacaterol mean recovery (precision) percentages at Low, Mid, and High QC levels were 93.5 (3.84), 89.8 (2.15), and 92.2 (2.17), respectively. Short-term, long-term, freeze-thaw, and auto-sampler stability results were accepted.
A specific, accurate and precise HPLC-MS/MS method has been validated for indacaterol quantification in human urine. This simple method is reproducible and robust to support future, indacaterol-related pharmacokinetic, bioequivalence and clinical studies.
茚达特罗是一种新型的每日一次吸入用超长效β2受体激动剂。定量生物分析支持药代动力学和临床研究。当前工作的目的是验证一种内部开发的高效液相色谱（HPLC）-串联质谱（MS/MS）分析方法，用于测定人尿液样本中的茚达特罗。
已开发出一种液-液萃取方法，使用乙酸乙酯从人尿液样本中萃取茚达特罗。茚达特罗干提取物用200μL流动相（酸化水：甲醇（30:70，v/v））复溶，其中5μL用于HPLC-MS/MS分析。茚达特罗在反相C18固定相上，以等度流动相在1mL/min的流速下洗脱。福莫特罗为内标（IS）。MS/MS检测采用涡轮离子喷雾电离，在正离子模式下进行。遵循国际生物分析方法验证指南的共识。
茚达特罗在质荷比（m/z）为393.3处被检测到，其MS/MS子离子在173.2处。茚达特罗和内标的保留时间分别为1.60和1.20分钟。验证的校准曲线在0.075 - 100 ng/mL范围内呈线性，相关系数（r）≥0.990。曲线的回归加权因子为1/x。在六个不同的人尿液批次中确定了方法特异性。未观察到基质干扰。确认了批内和批间精密度以及准确度在±20%（下限）和±15%（其他质量控制（QC）水平）范围内。低、中、高QC水平下茚达特罗的平均回收率（精密度）百分比分别为93.5（3.84）、89.8（2.15）和92.2（2.17）。短期、长期、冻融和自动进样器稳定性结果均符合要求。
已验证一种用于人尿液中茚达特罗定量的特异性、准确且精密的HPLC-MS/MS方法。这种简单方法具有可重复性且稳健，可为未来与茚达特罗相关的药代动力学、生物等效性和临床研究提供支持。
