A novel kinetic cholinesterase-inhibition based method for quantification of biperiden in pharmaceutical preparations

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Vladan R. Djurić
Violeta D. Mitić
Nebojša R Deletić
Marija D. Ilić
Bratislav M. Ćirković
Vesna P. Stankov-Jovanović


Biperiden, an antiparkinsonian anticholinergic drug, has been found to inhibit enzymatic hydrolysis of butyrylthiocholine iodide, which is catalyzed by serum cholinesterase. By measuring the difference in the basic and inhibitory hydrolysis reaction rates, in the presence of biperiden as an inhibitor, it is possible to develop a kinetic method for its determination. Both systems, enzyme-substrate-chromogen and enzyme-substrate-chromogen-inhibitor, were characterized by biochemical parameters (KM = 0.326 – 0.330 mmol dm-3; Vmax = 40 ‑ 99 μmol dm-3 min-1), while inhibition was defined as non-competitive with the constant of inhibition Ki = 6.142 μmol dm-3. The reaction conditions have been optimized followed by determination of the calibration curve, the corresponding equation and the limits of detection and quantification yielding 3.84 and 12.80 nmol dm-3, respectively. Using the calibration chart, it is possible to determine biperiden in different samples in the concentration range of 0.035–35.940 µmol dm-3. Influence of a number of substances, found in the sample, on the reaction rate was also examined. The optimized method was applied for determination of biperiden in pharmaceutical preparations. Accuracy of the method was tested using the standard addition method. The proposed method has good sensitivity, selectivity, it is simple and fast, and above all easily accessible, and thus applicable in biochemical and pharmaceutical laboratories.


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Djurić, V. R., Mitić, V. D., Deletić, N. R., Ilić, M. D., Ćirković, B. M., & Stankov-Jovanović, V. P. (2020). A novel kinetic cholinesterase-inhibition based method for quantification of biperiden in pharmaceutical preparations. HEMIJSKA INDUSTRIJA (Chemical Industry), 74(4), 265–271. https://doi.org/10.2298/HEMIND200413021D
Biochemical Engineering - Kinetics of Bioprocesses


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