Development and validation of bioanalytical technique of imatinib quantitative determination in human plasma by HPLC
Abstract
Imatinib is one of the best anticancer drugs used in the treatment of melanoma, and myeloleukemia.
According to federal program "PHARMA 2020", involving the import substitution of drugs, imatinib is of
great interest to pharmaceutical manufacturers. For state registration of generics in accordance with the Russian
legislation is necessary to confirm its efficacy and safety of the original drug through a clinical bioequivalence
studies. A mandatory step in this research is the development of analytical methods for quantitative
determination of the drug in human plasma with a preliminary selection of sample preparation conditions.
Currently, according to literature data, the determination of imatinib in human blood plasma by HPLC with
UV detection has at least 50 ng/ml limit of detection. In this context, the aim of this work was to develop a
more sensitive bioanalytical method of imatinib quantitative determination in human plasma by HPLC with
UV detection and its validation.
Quantitative determination of imatinib in plasma was carried out on a chromatograph MilichromA02
(CJSC "EcoNova", Novosibirsk) column size 75.0x2.0 mm filled with 5 micron sorbent ProntoSIL C18
and spectrophotometric detection at 260 nm. To reduce the limit of quantification of imatinib in plasma and,
hence, improve sensitivity of the test, compared with the already known data, applied dynamic modification
of the mobile phase. A mixture of 0.5% potassium dihydrogen phosphate, methanol and triethylamine solution
was used as mobile phase in the ratio 74:25:1 at a pH of 3.3, adjusted with orthophosphoric acid, as mobile
phase B used methanol with a pH of 3.3. Eluent flow rate was 0.1 ml/min, column temperature - 35 ° C,
sample injection volume - 20 microliters. Method liquid-liquid extraction principle acetonitrile QuEChERS
was used for extraction from plasma. To confirm the suitability of the developed method was performed a validation on the basis of its
«Guidelines for the examination of drugs. Volume 1» (Russia, 2013) and in accordance with the validation
guidelines requirements« Guidance for Industry bioanalytical methods: Bioanalytical method validation»
(FDA, USA, 2001) and «Guideline on validation of bioanalytical methods» (EMA, England, 2009). Selectivity
of techniques and its linearity in the concentration range from 42 to 4200 ng/ml were proved. Accuracy of
the procedure ranges is from 92.1 to 104.7%, and its precision - from 92.6 to 97.9%. The absence of the
transfer of an analyte in a sample after the sample single analysis with a high concentration of analyte is
demonstrated, stability of analyzed samples is elicited. Developed and validated technique can be applied to the analysis of human plasma samples containing
imatinib in various pharmacokinetic studies and clinical bioequivalence study.
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