Nowadays, measurement uncertainty is considered an essential parameter for method evaluation. Measurement uncertainty estimation requires a detailed study of all possible sources of uncertainty. The aim of this work was to develop and validate an isocratic reversed-phase high-performance liquid chromatographic method for the simultaneous separation and determination of five antihistamines in pharmaceutical formulations and to establish a procedure to evaluate the measurement uncertainty of values generated by the validated method. A relatively short chromatographic separation time, less than 9 min, was achieved by means of a Waters XBridge C18 (4.6 mm×250 mm, 5 µm) analytical column under isocratic conditions using a mobile phase consisting of sodium perchlorate buffer (pH 3.5; 0.1 mol L −1 )–acetonitrile (55:45, v/v) at a flow rate of 1 mL min −1 and UV detection at 235 nm. The selectivity, method linearity, accuracy, precision, limits of detection (LOD) and limits of quantification were examined as parts of the method validation. The described method shows excellent linearity over a range of 30 μg mL −1 to 70 μg mL −1 for all compounds with correlation coefficients higher than 0.995. The standard deviations of the intraday and inter-day precision were all less than 2 %. The LOD of all studied compounds ranged from 0.029 μg mL −1 to 0.03 µg mL −1 . Using EURACHEM/CITAC guide and Valid Analytical Measurement (LGC/VAM) protocol, measurement uncertainty associated with precision and standard preparation were the most important contributing for the five antihistamines, accounting for 87.72 % to 91.03 % and 4.05 % to 10.24 % of total uncertainty, respectively, and overall uncertainties obtained were from 0.05 mg to 0.58 mg. Therefore, the proposed method was suitable for the routine analysis of antihistamines in pharmaceuticals formulations with the required accuracy, precision and measurement uncertainty.

中文翻译：

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测量不确定度的估计和 RP-HPLC 的验证，用于同时测定药物制剂中的五种抗组胺药

如今，测量不确定度被认为是方法评估的重要参数。测量不确定性估计需要对所有可能的不确定性来源进行详细研究。这项工作的目的是开发和验证一种等度反相高效液相色谱方法，用于同时分离和测定药物制剂中的五种抗组胺药，并建立一个程序来评估由验证方法产生的值的测量不确定度. 使用 Waters XBridge C18（4.6 毫米×250 毫米，5 微米）分析柱在等度条件下使用由高氯酸钠缓冲液（pH 3.5；0.1）组成的流动相实现了相对较短的色谱分离时间，不到 9 分钟mol L -1 )-乙腈 (55:45, v/v) 流速为 1 mL min -1，UV 检测波长为 235 nm。作为方法验证的一部分，对选择性、方法线性、准确度、精密度、检测限 (LOD) 和定量限进行了检查。对于相关系数高于 0.995 的所有化合物，所描述的方法在 30 μg mL -1 至 70 μg mL -1 的范围内显示出优异的线性。日内和日间精密度的标准偏差均小于 2%。所有研究化合物的 LOD 范围为 0.029 µg mL -1 至 0.03 µg mL -1 。使用 EURACHEM/CITAC 指南和有效分析测量 (LGC/VAM) 协议，与精密度和标准制备相关的测量不确定度是五种抗组胺药的最重要贡献因素，分别占总不确定度的 87.72 % 至 91.03 % 和 4.05 % 至 10.24 % ， 分别，获得的总体不确定度为 0.05 mg 至 0.58 mg。因此，所提出的方法适用于具有所需准确度、精密度和测量不确定度的药物制剂中抗组胺药的常规分析。