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HPLC标准曲线法测定氟康唑杂质校正因子的测量不确定度评定

Evaluation on measurement uncertainty of correction factors of fluconazole impurities determined by HPLC standard curve method

  • 摘要: 通过对HPLC标准曲线法测定氟康唑杂质校正因子的实验过程进行不确定度评定,探索影响杂质校正因子测定准确性的主要共性因素,为提高测定结果的准确性提供研究思路。首先分别建立了氟康唑及其杂质A、B、C、D、F、I相应的拟合直线,计算各杂质及其对应主成分的拟合直线斜率之比作为该杂质的校正因子。接着以GUM法为基础,根据已经建立的校正因子测定过程的不确定度评定方案,分别计算得到标准曲线法测定各杂质校正因子的不确定度。氟康唑杂质A、B、C、D、F、I校正因子分别为 1.068 ± 0.046、0.102 ± 0.005、0.0582 ± 0.003、1.382 ± 0.121、0.802 ± 0.067和1.383 ± 0.119,其中包含因子k = 2。最后计算各不确定度分量的贡献率,氟康唑杂质A、B、C、D、F、I校正因子测定的相对合成标准不确定度urelf)中主成分和杂质的线性方程斜率不确定度urelK)贡献率之和占比均大于85%;在线性方程斜率不确定度urelK)中,12组数据中的8组溶液浓度不确定度贡献率占比在80%以上,其中由对照品含量在溶液浓度中引入的不确定度贡献率约为80%。由此可见,标准曲线法测定杂质校正因子的过程中线性溶液浓度的配制是最大影响因素,其次是直线拟合过程;在线性溶液浓度配制过程中对照品的纯度是最大影响因素,其次是称量和移液次数。该结论的得出可以帮助实验人员在从事类似工作时更好的制定实验方案,保障结果的准确性。

     

    Abstract: In this paper, the uncertainties of correction factors of fluconazole impurities determined by HPLC standard curve method were evaluated, and the main common factors affecting the accuracy of standard curve method were found, so as to improve the accuracy of the method.In this study, the corresponding fitting lines of fluconazole and its impurities A, B, C, D, F and I were established respectively, and the ratio of the slope of fitting lines of each impurity and its corresponding principal component was calculated as the correction factor of the impurity.Then on the basis of GUM method, the uncertainty of each impurity correction factor determined by standard curve method was evaluated according to the established uncertainty evaluation scheme of correction factor determination process.The correction factor and uncertainty of fluconazole impurities A, B, C, D, F and I were 1.068 ± 0.046, 0.102 ± 0.005, 0.0582 ± 0.0031, 1.382 ± 0.121, 0.802 ± 0.067 and 1.383 ± 0.119, respectively, and the coverage factor k was 2.Finally, the contribution rate of each uncertainty component was calculated.In the relative combined standard uncertainties urel(f) of fluconazole impurities A, B, C, D, F and I correction factors, the sum of contribution rate of slope uncertainty urel(K) of the linear equation of principal component and its impurity is more than 85%; in the slope uncertainties urel(K) of linear equation, the contribution rates of uncertainties of solution concentration in 8 of 12 data groups are more than 80%, and the contribution rates of uncertainties introduced by reference substance content in solution concentration are about 80%.It can be seen that the preparation of linear solution concentration is the most influential factor in the determination of impurity correction factor by standard curve method, followed by the linear fitting process.In the preparation process of linear solution concentration, the purity of reference substance is the most influential factor, followed by weighing and pipetting times.The conclusion can help the experimenters to better formulate experimental plans and ensure the accuracy of the results when doing similar work.

     

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