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DEVELOPMENT AND VALIDATION OF HPLC-MS/MS METHOD OF DETERMINATION OF A NEW VALPROIC ACID DERIVATIVE AND 1,3,4-THIADIAZOLE IN RAT BRAIN
DOI: https://doi.org/10.29296/24999490-2020-04-06
Introduction. To assess the pharmacokinetic indices of new drugs, it is necessary to develop analytical methods for determining their concentration in biological objects. Purpose of the study. Development and validation of HPLC-mass spectrometric methods for the determination of a new valproic acid derivative and 1,3,4-thiadiazole in rat brain. Methods. The object of the study was a new anticonvulsant N- (5-ethyl-1,3,4-thiadiazol-2-yl)-2-propylpentanamide (valprazolamide). Determination of valprazolamide was carried out by the HPLC-mass spectrometric method. Selectivity, sample transfer, linearity, accuracy, precision, matrix effect, the degree of extraction of valprazolamide were evaluated. Results. A method of HPLC-mass spectrometric determination of valprazolamide in rat brain homogenates was developed (Zorbax Eclipsi plus C18 analytical column – 4 μm 2,0×150 mm, temperature – 30°C; the mobile phase is acetonitrile and deionized water with the following gradient profile: 0–1 minute 10% acetonitrile; 1–5 minutes a linear increase in the concentration of acetonitrile to 90%; 5–6 minutes isocratic plot with a concentration of acetonitrile 90%; 6–10 minutes column conditioning with 10% acetonitrile; the flow rate of the mobile phase is of 0.6 ml/min; the volume of the injected sample accounts for 10 μl; total elution time – 10 minutes, mass detection with negative polarization). The MRM transition of valprazolamide amounted to m/z 254,0 → m/z 167,1. Conclusion. The analytical limits of the method are in the range from 1 to 1000 ng/ml. The developed method is selective, accurate, precise and linear, it meets the requirements for the validation of bioanalytical methods in all respects.
Keywords:
HPLC-MS/MS, valproic acid, 1, 3, 4-thiadiazole, antiepileptic drugs, mass spectrometry
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