Molnupiravir is an antiviral drug against viral RNA polymerase activity approved by the FDA for the treatment of COVID-19, which is metabolized to β-D-N4-hydroxycytidine (NHC) in human blood plasma. A novel method was developed and validated for quantifying NHC in human plasma within the analytical range of 10–10,000 ng/mL using high-performance liquid chromatography with tandem mass spectrometry (HPLC-MS/MS) to support pharmacokinetics studies. For sample preparation, the method of protein precipitation by acetonitrile was used, with promethazine as an internal standard. Chromatographic separation was carried out on a Shim-pack GWS C18 (150 mm × 4.6 mm, 5 μm) column in a gradient elution mode. A 0.1% formic acid solution in water with 0.08% ammonia solution (eluent A, v/v) and 0.1% formic acid solution in methanol with 0.08% ammonia solution mixed with acetonitrile in a 4:1 ratio (eluent B, v/v) were used as a mobile phase. Electrospray ionization (ESI) was used as an ionization source. The developed method was validated in accordance with the Eurasian Economic Union (EAEU) rules, based on the European Medicines Agency (EMA) and Food and Drug Administration (FDA) guidelines for the following parameters and used within the analytical part of the clinical study of molnupiravir drugs: selectivity, suitability of standard sample, matrix effect, calibration curve, accuracy, precision, recovery, lower limit of quantification (LLOQ), carryover, and stability.

Изучение фармакокинетики препарата «КОВИД-глобулин» (специфический иммуноглобулин человека против COVID-19) (АО «Нацимбио», Россия) в рамках фазы I клинического исследования

Introduction. Coronavirus disease (COVID-19) is an acute infectious disease caused by SARS-CoV-2 (severe acute respiratory syndrome-related coronavirus 2). Favipiravir is a synthetic prodrug with antiviral activity used for the treatment of COVID-19. There are oral and parenteral dosage forms of favipiravir. Compared with oral administration, parenteral administration has some advantages. Developing a method for the determination of favipiravir in human blood plasma is necessary for performing the analytical part of clinical studies of favipiravir for parenteral administration as an infusion, studying pharmacokinetics, and choosing the optimal dosage of the drug.Aim. The aim of this study is to develop and validate a method for quantitative determination of favipiravir in human plasma by high-performance liquid chromatography with ultraviolet detection (HPLC-UV) for pharmacokinetic studies.Materials and methods. Determination of favipiravir in human plasma by HPLC-UV. The UV detection was set at 323 ± 2 nm. The samples were processed by methanol protein precipitation. Internal standard: raltegravir. Mobile phase: 0.1 % formic acid in water with 0.08 % aqueous ammonia (eluent A), 0.1 % formic acid in acetonitrile with 0.08 % aqueous ammonia (eluent B). Column: Phenomenex Kinetex^®, C18, 150 × 4.6 mm, 5 μm. Analytical range: 0.25–200.00 μg/mL.Results and discussion. This method was validated by selectivity, calibration curve, accuracy, precision, spike recovery, the lower limit of quantification, carry-over effect and stability.Conclusion. We developed and validated the method of quantitative determination of favipiravir in human plasma by HPLC-UV. The analytical range was 0.25–200.00 μg/mL in human plasma. The method could be applied in pharmacokinetics studies of favipiravir.

Introduction.Allaforte® (JSC "Pharmcenter VILAR", Russia) is an antiarrhythmic long-acting drug. The dosage form of the drug Allaforte® provides a decrease in the frequency of taking the drug and also reduces the risk of side effects. It is relevant when taking antiarrhythmic drugs of the IC class. However, the pharmacokinetics of this drug has not been studied on humans. Therefore, it is important to fully study the pharmacokinetics to ensure the maximum efficacy and safety of arrhythmia therapy.Aim.The aim is pharmacokinetics study of long-acting antiarrhythmic drug Allaforte® (JSC "Pharmcenter VILAR", Russia), 25 mg. Materials and methods. Concentration of lappaconitine and its active metabolite N-desacetyllappaconitine in human plasma determinates by high performance liquid chromatography with tandem mass-spectrometry. Pharmacokinetic parameters calculated by R Project 3.5.1 software (package «bear», version 2.8.3-2), originally created by Hsin-ya Lee and Yung-jin Lee, Taiwan.Results and discussion.Pharmacokinetic parameters of lappaconitine and N-desacetyllappaconitine were calculated. Averaged pharmacokinetic profiles (in linear and semi-log scale) of lappaconitine and N-desacetyllappaconitine after single administration under fasting were built. The means of the maximum concentrations (Cmax) determined in the blood plasma of volunteers after single administration Allaforte® are 5.09 ± 4.07 ng/ml for lappaconitine and 11.66 ± 6.21 ng/ml for N-deacetyllappaconitine (Mean ± SD). The peak time of the maximum concentrations (Tmax) is 4.43 ± 3.54 hours for lappaconitine and 4.04 ± 2.18 hours for N-deacetyllappaconitine. The means of the areas under the curve plasma concentration – time from 0 to 48 hours (AUC0-t) and under the curve plasma concentration–time from zero to infinity (AUC0-∞) of Allaforte® is 42.96 ± 34.48 ng ∙ h/ml and 71.24 ± 43.20 ng ∙ h/ml for lappaconitine; 167.42 ± 114.41 ng ∙ h/ml and 189.42 ± 115.20 ng ∙ h/ml for N-deacetyllappaconitine. Allaforte® was eliminated from blood plasma with means of terminal half-life (T1/2) 8.45 ± 5.10 hours for lappaconitine and 9.04 ± 2.57 hours for N-deacetyllappaconitine.Conclusion.Pharmacokinetics study of long-acting antiarrhythmic drug Allaforte® (JSC "Pharmcenter VILAR", Russia) after single administration was researched. Results of the study allows to conduct an effective therapy of arrhythmia by study drug and minimize side effects.

Introduction. Coronavirus disease 2019 (COVID-19) is an infectious disease caused by the SARS-CoV-2 virus (severe acute respiratory syndrome-related coronavirus 2). COVID-19 is now expected to stay with us for many years as a recurring disease. Molnupiravir and favipiravir are oral antiviral drugs with anti-RNA polymerase activity. The Russian Health Ministry has approved molnupiravir and favipiravir for the treatment of COVID-19. The study describes development and validation of high-performance liquid chromatography – tandem mass spectrometry (HPLC-MS/MS) method for the simultaneous determination of β-D-N4-Hydroxycytidine and favipiravir in human blood plasma. The method could be applied in pharmacokinetic study of molnupiravir and favipiravir.Aim. The aim of this study is to develop and validate a HPLC-MS/MS bioanalytical method for the determination of β-D-N4-Hydroxycytidine and favipiravir in human plasma.Materials and methods. The determination of β-D-N4-Hydroxycytidine and favipiravir in human plasma by HPLC-MS/MS. The samples were processed by 0.1 % formic acid in acetonitrile. Internal standard: promethazine. Mobile phase: 0.01 mol/L Ammonium formate buffer solution (Eluent A), 0.1 % formic acid and 0.08 % aqueous ammonia in water/acetonitrile 10 : 90 (Eluent B). Column: Shim-pack GWS C18, 150 × 4.6 mm, 5 μm. Analytical range: 50.00–10000.00 ng/mL for β-D-N4-Hydroxycytidine, 250.00–20000.00 ng/mL for favipiravir in human plasma. Ionization source: electrospray ionization. Detection conditions: 260.00 m/z → 82.10 m/z, 260.00 m/z → 111.00 m/z, 260.00 m/z → 127.95 m/z (β-D-N4-Hydroxycytidine); 156.15 m/z → 65.95 m/z, 156.15 m/z → 85.00 m/z, 156.15 m/z → 113.10 m/z (favipiravir); 285.05 m/z → 198.05 m/z (promethazine).Results and discussion. This method was validated by selectivity, suitability of reference standard, matrix effect, calibration curve, accuracy, precision, spike recovery, the lower limit of quantification, carry-over effect and stability.Conclusion. The HPLC-MS/MS method for quantitative determination of β-D-N4-Hydroxycytidine and favipiravir in human plasma was developed and validated. The analytical range was 50.00–10000.00 ng/mL for β-D-N4-Hydroxycytidine, 250.00–20000.00 ng/mL for favipiravir in human plasma. This method was applied to investigate the pharmacokinetics of molnupiravir and favipiravir.

Introduction. Currently, physicochemical methods of quantification are actively used to determine the content of drugs in biological fluids. High-performance liquid chromatography with various detection methods is particularly widespread. One of the most difficult practical tasks is the chromatographic separation of so-called poorly retained compounds – drug substances poorly retained on the chromatographic column. Valganciclovir and Ganciclovir are among such substances. Aim. The aim of this study is to develop a method for valganciclovir and ganciclovir in human plasma by high performance liquid chromatography with tandem mass-spectrometry (HPLC-MS/MS) for pharmacokinetic studies.Materials and methods. Determination of valganciclovir and ganciclovir in plasma by HPLC-MS/MS. The samples were processed by acetonitrile protein precipitation.Results and discussion. This method was validated by next parameters: selectivity, matrix effect, calibration curve, accuracy, precision, recovery, lower limit of quantification, carry-over and stability.Conclusion. The method of the determination of valganciclovir and ganciclovir in human plasma was developed and validated by HPLC-MS/MS. The linearity in plasma sample was achieved in the concentration range of 5.00–1000.00 ng/ml for valganciclovir and 50.00–10000.00 ng/ml for ganciclovir. Method could be applied to valganciclovir and ganciclovir determination in plasma for PK and BE studies.