Determination of relevant substances in Clofazimine active pharmaceutical ingredients by HPLC method
Objective To establish a method for determining the contents of substances in Clofazimine active pharmaceutical ingredient(API).Methods High performance liquid chromatography(HPLC)was performed on a Caprisil C8-P(250 mm×4.6 mm,5 μm)column.And 4.5 g of sodium dodecyl sulfate,1.7 g of tetrabutylammonium hydrogensulfate and 1.77 g of disodium hydrogen phosphate dodecahydrate were dissolved in 1 000 mL of water,then adjusted to pH=3.0 with dilute phosphoric acid,and then the above solution was mixed with acetonitrile in the ratio of 35∶65 as the mobile phase.The flow rate was 1.0 mL/min,the detection wavelength was 280 nm,the column temperature was 30℃,and the injection volume was 20 μL.Methodological validation of this HPLC method was carried out.The contents of relevant substances in three batches of Chlorpheniramine API were determined,and the results measured by the external standard method were compared with the principal component self-control method with correction factor.Results There was a good linear relationship for Clofazimine,impurity A and impurity B in the range of 0.019-0.959 μg/mL,0.017-0.858 μg/mL,and 0.060-2.995 μg/mL,respectively(all R2＞0.999),the limits of detection were 0.22 ng,0.27 ng,and 0.23 ng,and the limits of quantification were 0.72 ng,0.91 ng,and 0.76 ng,respectively.The RSDs of precision,repeatability,stability and durability experi-ments were all less than 5%,and the average recoveries were 93.37%-98.17%.Impurity A and impurity B were detected in all three batches of Clofazimine raw material,with the content range of 0.013%-0.014%for impurity A,and 0.025%-0.026%for impurity B.There was no significant difference in the contents of related substances determined by the principal components self-control method with correction factor and the external standard method.Conclusion The established HPLC method is convenient,efficient and accurate,and can be used for determining the contents of the relevant substances in Clofazimine API.The principal component self-control method with correction factor is low cost and practical.
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