We discussed critical laboratory steps in the drug registration process, reviewed current regulatory requirements, and shared practical approaches to organizing and conducting research, the results of which form key sections of the registration dossier.

We present a new issue of the "Leadership Opinion" series, dedicated to strategic projects in the field of biotechnology and immunoprophylaxis.

We present the latest issue of the "Leadership Opinion" series. Collaboration with universities and research organizations has always been an important part of the work of the Center of Pharmaceutical Analytics and the journal "Drug Development and Registration". The St. Petersburg Chemical and Pharmaceutical University has been the Center's key partner university for many years.

The article presents a transcript of a report by A. G. Natradze, Deputy Commissar of Health of the USSR, which was read at a meeting of industrial workers on January 31, 1944. The report summarizes the industry's performance, highlighting the increase in drug production by chemical and pharmaceutical enterprises in 1943 compared to 1942 for most products, as well as the failure to meet the plan for restoring the previous range of drugs and introducing new products. During 1942 and 1943, the newly established enterprises of Glavmedfarmprom, which had previously produced only simple galenic preparations, began to produce complex chemical drugs. New enterprises were established in the Union republics (Tashkent, Frunze, and Baku), which began to produce both galenic preparations and complex substances. At the same time, A. G. Natradze pointed out the measures that, during the reporting period, led to an increase in labor productivity and a decrease losses of energy, raw materials, and supplies. The majority of the report focused on the industry's plans for 1944.

Ion or ion-exchange chromatography (IC), although somewhat less popular than traditional HPLC, is also widely used for quality control of drugs and substances in the pharmaceutical industry. Most often, the target compounds are inorganic ions, but IC is also used to separate organic acids, sugars, aminoglycosides, amino acids, and other substances.

From April 22 to 24, 2026, the 24th International Exhibition of Laboratory Equipment and Chemical Reagents "Analitika Expo 2026" was held at the Crocus Expo IEC.

Introduction. Polyphenols are widely distributed in plant world and recognized for their high antioxidant activity. Traditional methods of polyphenolic compounds quantification, such as Folin – Ciocalteu and aluminum chloride assays, measure total phenolic or flavonoid content but fail to identify individual compounds in complex mixtures. Combining the kinetic profiling of antioxidant activity manifestation for a combination of antioxidants has certain potential for the development of inexpensive and simple quantitate analysis approaches.

Aim. To develop an approach for the quantitative determination of individual polyphenols in binary combinations through the measurement of ABTS^•+ radical cations accumulation kinetics based on a lag-time experimental strategy.

Materials and methods. We have studied formulations of flavonoid dihydroquercetin (DHQ) and lignan secoisolariciresinol (SECO) and a combination of DHQ with essential antioxidant alpha-tocopherol (α-TOH). The antioxidants induced the inhibition of ABTS^•+ radical cations accumulation initiated by potassium persulfate, which resulted in a lag-time formation and was monitored spectrophotometrically at 730–750 nm.

Results and discussion. Both combinations demonstrated two lag-time periods pattern typical for additive type interactions between its components. Lag-time period duration depends on concentration of the component. We matched every lag-time period with specific polyphenol which allowed us to determine the content of every component of studied combinations. Developed technique measured the content of polyphenols with value of relative standard deviation (RSD) being not higher than 2 % and value of relative measurement error (δ) not exceeding 3,7 %.

Conclusion. Our study confirmed that an antioxidant capacity measurement approach could be successfully applied for content determination of individual polyphenols in complex mixtures. Developed methodology demonstrated high reproducibility and acceptable accuracy levels for binary combinations constituents’ quantification.

Introduction. Continuous efforts to uncover new antiviral entities targeting HIV RT RNase H stem from the suffering of affected individuals and the relentless increase in death cases. The present study focused on ten molecules based on the pyrimidine-2,4-dione scaffold, which were discovered to have potent antiviral activity against HIV RT RNase H and acceptable bioavailability and pharmacokinetic properties.

Aim. The main goal of this study is to discover and evaluate promising pyrimidine-2,4-dione-derived molecules as potential inhibitors of HIV.

Materials and methods. A docking protocol was executed to investigate the binding mechanisms of the designed compounds within RNase H's active site using the AutoDock Vina program. The molecular dynamics simulation analysis for 100 ns was carried out using Desmond, and the complexes were constructed using the OPLS3-2005 force field. The frontier molecular orbital energies and related reactivity factors of the best antiviral agents were determined using the DFT-B3LYP/6-31G (d,p) calculations performed with the Gaussian program.

Results and discussion. Two molecules, Pyr06 and Pyr07, displayed a higher docking score (–10.7 kcal/mol) than the control compound (MPD: –9.8 kcal/mol) and the other designed compounds (with affinities ranging from –9.6 to –10.4 kcal/mol). On the other hand, it was noted that these two molecules formed significant hydrogen bonds and hydrophobic contacts with active site residues. Furthermore, our analysis through MD simulations for 100 ns validated the binding stability and conformation of molecules Pyr06 and Pyr07 in complexes with the 5J1E receptor. The DFT findings for both molecules, Pyr06 and Pyr07, also confirmed their docking scores and their ability to form stable complexes with RNase H.

Conclusion. The results obtained could be highly beneficial for designing and developing Pyr06 and Pyr07 as potential therapeutic drugs to combat HIV RT RNase H.

Introduction. Matrix tablets have great potential as an oral drug delivery system due to their simplicity, reduced risk of systemic toxicity and minimal likelihood of dose splitting effects. Based on the DoE design of experiments, the response surface methodology involves generating polynomial equations and response in the experimental domain to determine the optimal formulation of a dosage form. Traditionally, developers select optimal formulations by changing one variable at a time, which is a time-consuming method. However, many experiments fail to achieve their goal because they are not thought out and designed properly, and even the best data analysis cannot compensate for the lack of planning. Therefore, it is important to understand the impact of variables in a pharmaceutical product formulation on quality using a minimum number of experimental trials and variables to develop an optimized formulation using established statistical tools.

Aim. Development of a modified release matrix tablet formulation based on 3,6,9-triazatricyclotetradecane derivative using response surface methodology.

Materials and methods. The object of the study is the drug candidate compound QM-25, a derivative of 3,6,9-triazatricyclotetradecane. A 2³ full factorial experiment was used to develop the composition of the matrix tablet based on QM-25. The matrix tablets were obtained by direct compression on a rotary press after proper mixing of suitable ratios of various hydrophilic polymers as release modifiers with other excipients. A tablet hardness tester was used to measure the crushing strength of the prepared matrix tablets. The in vitro release studies were carried out on a dissolution apparatus II "Paddle mixer", at 50 rpm and a temperature of 37 ± 0.5 °C in two stages for drugs of group 2 in accordance with OFS.1.4.2.0014. The completeness of the formulation release after 2 hours 45 minutes in % and the tablet crushing strength in N were selected as the responses of the factorial design system. To select the optimal formulation and to evaluate the influence of the effects of different variables on the measured responses, a mathematical model equation was used, including independent variables and their interactions for the different measured responses generated by the design 2³. Data approximation and regression parameters were calculated using the built-in Data Analysis module of Microsoft Excel. To construct the regression equation in Microsoft Excel, a matrix was constructed using the correlation function.

Results and discussion. The algorithm of the response surface methodology based on the regression model is presented. Each response coefficient of the design parameter space was studied for its statistical significance. Factors with values in the interval p < 0.05 were identified. Using the regression model, regression coefficients of the main factors affecting release and crushing strength were established in the model equation, equations for the dependence of factors on responses, and the paired effect of the amount of sodium alginate and carbomer 940 per unit of the dosage form were derived. The influence of the main effects (factors) on the studied formulations was analyzed using the design parameter space with visualization in three-dimensional space. To select the optimal composition with the desired responses, a numerical optimization method was used based on the approach to obtaining the desired values, which made it possible to select the optimal values of the content of excipients per unit of the model composition of the dosage form.

Conclusion. Using the design method – construction of the design parameter space, the influence of many parameters of the matrix tablet composition was studied using a small number of experiments. The mathematical relationship between the X factors and the Y response indices was established using the calculated regression model for the solid dosage form based on the 3,6,9-triazatricyclotetradecane derivative. The calculated factor dependencies using the regression model allowed us to establish the optimal model composition of the solid dosage form with modified release for further experimental studies, the crushing strength values of which were in the confidence interval acceptable for tablets with the used punch diameter, and the dissolution kinetics results demonstrated the nature of the modified release behavior.

Introduction. Water-soluble polymers that are sensitive to temperature are promising for use in pharmaceutical development to create "smart" drug delivery systems.

Text. This review article discusses three groups of thermosensitive polymers: those with a lower critical solution temperature (LCST) and those with an upper critical solution temperature (UCST), as well as polymers exhibiting both types of behavior. The article presents their classification, development features, application examples and modifications to give the necessary properties.

Conclusion. Polymers with LCST and UCST behavior are promising materials for the development of thermosensitive drug delivery systems that are already used in biomedicine.

Introduction. Maintaining the basic quality parameters of a drug is the key to its successful use in clinical practice. Productive use of the results of risk identification and assessment allows us to guarantee the effectiveness and safety of medicines through preventive measures to identify and eliminate potential threats to quality during their development and production.

Aim. The aim of the study was to identify critical points in the technological process of obtaining a lyophilized dosage form of a compound from the class of indolocarbazoles LCS-1269 using a risk-based approach.

Materials and methods. To analyze the risks in the technology for obtaining a lyophilisate based on LCS-1269, the risk management analysis tools and methods presented in the ICH Q8, Q9, Q10, State Pharmacopoeia of Russia XV ed. and ISO 31000-2019 guidelines were used.

Results and discussion. When identifying risks of the production process, the main critical process parameters and critical control points were identified, among which the most important are compliance with the mode of dissolution of the pharmaceutical substance and excipients, filtration of the resulting solution and its lyophilization, as well as control over the main quality indicators characteristic of lyophilized dosage forms for parenteral use.

Conclusion. The application of the selected approach made it possible to identify and assess possible risks to product quality at different stages of production of the drug "LCS-1269, lyophilisate for preparation of injection solution 25 mg" and to develop a strategy to eliminate or mitigate these risks and achieve the necessary goals, namely, obtaining а high-quality, effective and safe medicinal product.

Introduction. Calcium is a key macroelement that ensures bone mineral density, muscle and cardiovascular function, and is involved in enzymatic and neurotransmitter processes. Calcium deficiency is widespread and associated with the development of osteoporosis and other metabolic disorders. The administration of medicinal preparations with calcium compensates its deficiency in patients and contribute to disease prevention and treatment. In the context of sustainable development, there is growing interest in new technologies for isolating organic calcium salts from marine bioresources for medicinal use.

Aim. To develop and compare technologies for extraction of organic calcium from the shells of Arctic shrimp Pandalus borealis (a waste of food industry), and to evaluate the composition and purity of the obtained products.

Materials and methods. The shells of cooked-frozen Arctic shrimps Pandalus borealis were processed by two calcium extraction technologies: (1) direct demineralization with organic acids (citric, malic and lactic) and (2) preliminary defatting of the raw material with ethanol followed by demineralization. Quantitative analysis of calcium (Ca) and other elements was performed by Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES). A hazard quotient (HQ) was calculated for safety assessments.

Results and discussion. It was found that the maximum calcium yield was achieved after demineralization with citric acid (153.0 ± 10.3 mg Ca/g). Malic and lactic acids showed less efficiency. Preliminary defatting reduced the Ca yield by an average of 2.2 %; however, the lipid fraction is of interest as a source of carotenoids and other natural lipids. Toxic elements (Pb, Cd, Ni, Cr, As) were either not detected or their concentrations were below the limit of quantification. The content of associated elements (Mg, Zn, Fe, Cu, and Mn) was below the toxic level (calculated HQ < 1). Consumption of isolated Ca products will provide enrichment of the dietary intake with certain microelements.

Conclusion. The shells of Arctic shrimps are a valuable source of organic calcium for the environmentally friendly production of physiologically balanced calcium salts – calcium citrate, malate, and lactate. The resulting calcium salts are not polluted with toxic elements and can be used in the pharmaceutical and food industries.

Introduction. Tirzepatide is a molecule capable of controlling blood glucose levels by combining dual agonism of the glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1) receptors. GLP-1 and GIP are incretin hormones involved in the regulation of glycemia and energy metabolism. Their combined agonism enhances the insulin response, reduces glucagon secretion and appetite, making it an effective strategy for the treatment of type 2 diabetes mellitus (T2DM) and obesity. The specific biological activity of such compounds can be evaluated in vitro using cell lines expressing GLP-1R or GIPR.

Aim. To validate a method for determining GLP-1R receptor activation in vitro using a cell line stimulated with tirzepatide in the "Calcium Flux" assay. This approach is necessary for subsequent studies of the comparability of bioequivalent drugs.

Materials and methods. A genetically engineered HTS163L cell line expressing the human GLP-1 receptor was used as the model. Receptor activation was recorded by changes in fluorescence signal caused by an increase in intracellular calcium concentration after incubation of cells with tirzepatide at various concentrations. Method validation was carried out in accordance with the requirements of the State Pharmacopoeia of the Russian Federation, ICH, and EMA. Statistical analysis of the results was performed using MARS software (BMG LABTECH).

Results and discussion. The applied method demonstrated a high level of specificity (placebo signal <2 % of the drug signal), a satisfactory limit of quantification (signal exceeded the blank by 43 %), good linearity (R² = 0.981), accuracy (within 77–119 % of nominal EC₅₀), and precision (CV ≤ 5 % within runs, ≤15 % between runs). All validation parameters met the established acceptance criteria, confirming the correctness of the chosen approach.

Conclusion. The proposed method demonstrates high sensitivity and reproducibility, making it suitable for functional analysis of the biological activity of GLP-1 receptor agonists, including tirzepatide, in in vitro models.

Introduction. In the context of digital transformation and global harmonization of regulatory requirements, the implementation of integrated quality management systems in clinical trials based on ICH E6(R3) has become increasingly relevant.

Aim. To conduct a comparative analysis of ICH E6(R3) against international standards (ISO, GAMP 5, PMBOK/APM-BOK) and assess their adaptability within the Russian regulatory environment.

Materials and methods. A systematic analysis was performed of the Russian regulatory framework – including Federal Law No. 61-FZ of April 12, 2010; Decision No. 79 of the Eurasian Economic Commission Council of November 3, 2016, approving the Good Clinical Practice Rules of the Eurasian Economic Union; and GOST R 52379-2005 "Good Clinical Practice" – as well as international guidelines (ICH, ISO, EMA, PMI, ISPE) and scientific publications from 2020 to 2025 indexed in Scopus and Web of Science.

Results and discussion. It was established that ICH E6(R3) necessitates a shift from reactive oversight to proactive quality management through the integration of risk-based approaches, digital technologies, and interdisciplinary standards. Within the Russian context, gaps in regulatory provisions and insufficient integration of quality management systems were identified.

Conclusion. To align with ICH E6(R3), Russian regulatory authorities and organizations must modernize internal processes, implement Risk-Based Quality Management (RBQM), and harmonize national requirements with international standards.

Introduction. The development of parenteral depot formulations of drugs with modified release is a critical area in the treatment of chronic diseases such as cancer, schizophrenia, and diabetes. These formulations improve treatment adherence, maintain stable plasma concentrations, and reduce the frequency of injections. However, their development is fraught with technological and pharmacokinetic challenges, including nonlinear release of the active ingredient and high interindividual variability.

Aim. To systematically analyze and summarize current methodological approaches to mathematical modeling of the pharmacokinetics of parenteral long-acting drug formulations.

Materials and methods. A targeted literature search was conducted in PubMed, Google Scholar, and Scopus for the period 2015–2025 using Boolean operators and combinations of relevant keywords. The review includes data on clinically used long-acting parenteral formulations as well as on polymeric carriers (PLGA, hydrogels, in situ gels, and hybrid systems). To illustrate instrumental approaches, we considered compartmental PK models, PBPK models, and models based on machine learning methods.

Results and discussion. The characteristic features of pharmacokinetic profiles of parenteral depot formulations are summarized, including the phases of initial burst, lag phase, controlled release, and late decay, as well as their relationship with the physicochemical properties of the carrier and the drug substance. The variability of the profile is shown to be associated with a combination of technological parameters (particle size, polymer composition and architecture), injection site and route of administration, and individual patient characteristics. The capabilities of nonlinear mixed-effects models, PBPK approaches, and machine learning-based models are demonstrated for describing inter- and intraindividual variability, performing simulations of dosing regimens, and supporting in silico optimization of formulation development. A basic system of ordinary differential equations (ODEs) is proposed that reflects sequential release from different depot fractions followed by distribution and elimination of the drug.

Conclusion. Pharmacokinetic modeling of parenteral long-acting formulations is a key tool within the MIDD (Model-Informed Drug Development) concept, enabling integration of mechanistic knowledge on release, absorption, and distribution with clinical data. The proposed basic ODE model structure can serve as a methodological framework for developing and adapting models for specific drug products, thereby supporting optimization of formulation composition and dosing regimens and reducing the extent of costly in vivo studies.

Introduction. Metformin is an oral antidiabetic drug with a known efficacy and safety profile. Currently, there is more data on the effect of metformin on central nervous system functions in small doses, not associated with hypoglycemic effects to influence the aging process and concomitant age-related diseases. However, there is insufficient information about the effect of metformin on the behavioral characteristics and cognitive functions of a healthy body.

Aim. Investigation of the effect of metformin in small doses on behavioral characteristics and some biochemical parameters of blood serum (glucose, cholesterol, and low-density lipoprotein (LDL) levels) in rats.

Materials and methods. The study was performed on 72 male rats weighing 200–250 g of the Wistar line. Behavioral testing was performed after 30 days of metformin administration at doses of 35 and 70 mg/kg. The «T-maze test» installation was used to study the effects on learning and memory processes, and the «Water maze test» was used to study spatial memory. The «Elevated plus-maze test» and «Extrapolation escape task» settings were used to assess the effect of metformin on anxiety in rats. «The Rotarod» complex was used to assess the effect on the physical activity of rats. The biochemical parameters of rat blood serum were measured using an automatic DIRUI CS-300B biochemical analyzer.

Results and discussion. It was found that 30-day administration of metformin in both doses (35 and 70 mg/kg) improved learning and memory processes in «T-maze test» test (p < 0.05). In addition, administration of metformin in small doses (35 and 70 mg/kg) It contributed to the development of the anxiolytic effect in «Elevated plus-maze» test and «Extrapolation escape task» tests (p < 0.05). It was shown that the administration of metformin (70 mg/kg) improved muscle endurance in «Rotarod» test (p < 0.05) against the background of unchanged glucose levels.

Conclusion. The results of the conducted studies determine the direction of further research on the possibility of using metformin in small doses that do not have a hypoglycemic effect for the correction of anxiety and cognitive disorders in pathological conditions of the CNS, including in the process of physiological aging.

Introduction. Obesity and its complications are a leading medical and social problem in Russia and worldwide. The agouti-mutant A^y/a mice are used as a model of monogenic obesity caused by a suppression of the melanocortin system. The so-called melanocortin syndrome includes not only a variety of metabolic abnormalities but also a number of neurocognitive alterations, which have been relatively little studied.

Aim. Our objective was to assess the short-term recognition memory in male and female A^y/a mice compared to wild-type littermates.

Materials and methods. The experiments were carried out on 8 male and 7 female C57Bl/6- A^y/a mice along with 8 male and 7 female wild-type (C57Bl/6-a/a) littermates, aged 1 year. Recognition memory was assessed via sequential testing in the "Novel object recognition", "Novel object location", "Object-in-Place" (OiP), and "Object recency discrimination" (ORD) tests with a 5 min- and 3 day-long intervals between the testing phases and between the tests themselves, respectively. For each of the tests, we calculated the discrimination index (DI) using a conventional formula.

Results and discussion. Wild-type mice of both sexes demonstrated positive DI values in all of the tests. A^y/a males performed comparably with their wild-type littermates in all of the tests, while A^y/a females had significantly lower mean DI values in the OiP and ORD tests (p < 0.05 for both). These results may reflect an impairment of allocentric spatial memory and associative recognition, which may indicate damage to the medial prefrontal cortex, the medial dorsal nucleus and/or nucleus reuniens of the thalamus. The presence of such deficit in A^y/a females exclusively may be related to greater severity of systemic and central metabolic abnormalities, insulin resistance, and oxidative stress.

Сonclusion. In this study, we found an impairment of higher-level aspects of short-term recognition memory in female agouti-mutant C57Bl/6- A^y/a mice. The state of other types of memory, sex- and age-specific differences and dynamics of memory deficits in A^y/a mice require further elucidation. The distinct features of short-term recognition memory in male and female A^y/a mice need to be taken into consideration when planning and conducting preclinical trials.

Introduction. BBP2023 is a new drug candidate of the sydnonymine group with cerebral vasodilating activity and procognitive properties. The pharmacological effects of BBP2023 are due to the presence of active metabolites. To conduct pharmacokinetic studies at the preclinical stage, it is necessary to develop a bioanalytical method for the determination of BBP2023 and its metabolites in rabbit blood plasma.

Aim. Development and validation of a chromatography-mass spectrometric method for the quantitative determination of a new derivative of sydnonymine and its metabolites in rabbit blood plasma.

Materials and methods. Quantitative determination of BBP2023 in rabbit blood plasma was performed by HPLC-MS/MS. The biomaterial sample preparation was based on the method of protein precipitation with methanol. Chromatographic separation was performed with a mixture of deionized water and acetonitrile in a gradient mode on a ZORBAX Eclipse XDB-C18 analytical column (4.6 × 50 mm, 5 μm) using an Agilent 1260 Infinity II chromatograph. Detection was performed in the MRM mode using an AB Sciex QTRAP 3200 MD mass spectrometer. N-(ethoxycarbonyl)-3-(4-methylpentan-2-yl)sydnonymine was used as the internal standard.

Results and discussion. The method for sample preparation of rabbit blood plasma has been developed, optimal conditions for chromatography and detection of the studied compound and its metabolites have been selected. The bioanalytical method has been validated according to the following parameters: selectivity, matrix effect, recovery rate, carry-over effect, linearity of the calibration range, lower limit of quantitative determination, intra- and inter-run accuracy and precision, as well as stability of analytes and the internal standard at the analysis stages.

Conclusion. A method for the quantitative determination of a new derivative of sydnonymine BBP2023 and its metabolites in rabbit blood plasma using HPLC-MS/MS has been developed and validated. The confirmed analytical ranges for BBP2023, BBP2023 A and BBP2023 C were 5.0–500.0 ng/ml (5.0–1000.0 ng/ml for geranamine). The method was used in the analytical part of the preclinical pharmacokinetic study of BBP2023 in rabbits.

Introduction. Increasing the survival of neurons in the perinecrotic zone of the brain is a promising strategy in the comprehensive treatment of ischemic stroke.

Aim. To study some molecular mechanisms of hydroxypyridine derivative 3-EA to increase survival of cerebral GABAergic neurons under ischemia-like conditions.

Materials and methods. The cell-protective property of the novel hydroxypyridine compound 3-EA was compared with that of nimodipine. A series of experiments were conducted on 7–9 DIV rat hippocampal neuroglial cell culture containing GABAergic neurons expressing the calcium-binding proteins calbindin and parvalbumin under ischemia-like conditions. Neuron survival was determined using a cytochemical reaction with propidium iodide, and calcium signaling was studied by assessment of fluorescence signal emitted by cells loaded with Fura-2.

Results and discussion. 1 mM 3-EA increased the survival rate of rat brain GABAergic neurons expressing calcium-binding proteins by 25 % (p = 0.001), and was comparable to the reference drug nimodipine. The compound induced an average 2.5-fold suppression of intracellular calcium concentrations by 40 minutes after the ischemia-like environment onset.

Conclusion. The 3-hydroxypyridine compound 3-EA exhibited cell-protective properties and preserved the population of GABAergic neurons containing calbindin and parvalbumin by suppressing glutamatergic excitotoxicity and calcium influx into the cell cytosol. This compound may be considered a potential drug candidate.

Introduction. Currently, the search and development of innovative cardiotropic drugs is an urgent task for pharmaceutical science. One such drug is a derivative of malonic acid, etmaben, which was synthesized at the St. Petersburg State Chemical and Pharmaceutical University (SPCPU). To conduct preclinical and clinical studies, it is necessary to develop and validate a bioanalytical method that allows for the quantitative determination of etmaben in complex biological matrices with minimal time and resource requirements.

Aim. The aim of the study is to develop and validate a method for the determination of etmaben in the blood plasma and organs of laboratory animals using high-performance liquid chromatography with ultraviolet detection (HPLC-UV) followed by testing and obtaining a pool of data on the distribution of etmaben in the organs and tissues of experimental animals.

Materials and methods. The development and validation of the bioanalytical method, as well as the quantitative determination of etmaben, were performed on a Flexar liquid chromatograph (PerkinElmer, USA) equipped with a UV detector and a thermostatted autosampler, using a Kromasil 100, 150 × 2.1, C8, 3.5 μm column (AkzoNobel, Netherlands). Detection was performed at a wavelength of 270 nm. A pharmacological experiment to test the method was conducted on 20 outbred male rats (Kurchatov Institute – "Rappolovo", Russia), divided into a control group (intact animals, n = 10) and a group with experimental chronic heart failure (CHF) (n = 10), a model of which was induced by ligation of the left coronary artery. All animals received etmaben intragastrically at a dose of 60 mg/kg for 30 days. Whole blood samples were collected for analysis on day 30, and organs (heart and kidneys) were isolated. Etmaben was quantified in plasma and tissue homogenates using HPLC-UV after sample preparation, including protein precipitation with acetonitrile (HPLC Grade, cat. no. A/0627/17, Thermo Fisher Scientific, USA).

Results and discussion. Optimal conditions for the chromatographic determination of etmaben in biological matrices (blood plasma and tissue homogenates) using HPLC-UV were developed. The developed method was validated using the following validation parameters: selectivity, calibration curve, accuracy, precision, lower limit of quantification (LLOQ), carryover effect, no effect of sample dilution and stability. The degree of etmaben recovery from the matrix was also assessed. The LLOQ for etmaben was 5.0 ng/mL. The developed method was successfully applied to assess etmaben distribution in the organs and tissues of laboratory animals at specific stages of preclinical testing. In CHF, plasma concentrations of etmaben doubled, and in pathologically altered myocardium, they increased by 47 % compared to control animals.

Conclusion. A bioanalytical method for determining etmaben using HPLC-UV was developed and validated. It was tested during preclinical trials to study the drug's distribution in organs and tissues of experimental animals. The observed selective accumulation of etmaben in damaged myocardium confirms its therapeutic relevance for the treatment of CHF.

Introduction. Access to insulin is limited by global barriers, the most crucial of which is the pricing monopoly of multinational originator companies. The expiry of patent on the originator product opens the way for the production of biosimilars being affordable products with comparable quality, effectiveness and safety. The imperative for the development of biosimilars is to conduct comprehensive comparability studies with the originator product, including physicochemical and biological characterization. The development of a comparability study program, specifically the selection of attributes and orthogonal methods for analyzing the structure and the functional integrity of the insulin molecule, should be based on a deep understanding of the structure and properties of the molecule, as well as on the latest scientific knowledge.

Aim. To conduct physicochemical and biological characterization of the medicinal product RinLis® (GP40011) in comparison with the originator medicinal product Humalog® (insulin lispro) to establish their comparability within the framework of the biosimilarity assessment.

Materials and methods. The primary protein structure was confirmed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and peptide mapping. Higher-order protein structures were investigated using capillary isoelectric focusing, Ellman's method, circular dichroism, fluorescence spectrophotometry and dynamic light scattering. The comparability of impurity profiles of the tested batches was studied using size-exclusion chromatography (SEC) and reversed-phase high-performance liquid chromatography (RP HPLC). Insulin receptor (IR-A and IR-B) binding kinetics assay, insulin receptor phosphorylation assay and metabolic cell-based assays (glucose uptake, insulin-induced lipogenesis, inhibition of stimulated lipolysis) were performed to characterize the functional properties and biological activity of the insulin products.

Results and discussion. GP40011 was found to have high similarity to the reference product Humalog® in terms of the primary protein structure, higher-order structures and impurity profile based on the results of the evaluation of the physicochemical properties. Functional characterization studies showed that GP40011 and Humalog® had identical insulin receptor binding kinetics to IR-A, IR-B and comparable biological activity.

Conclusion. The study results demonstrated full analytical similarity of RinLis® to Humalog®. The study made a significant contribution to the scientific substantiation of the equivalence of the biosimilar RinLis® and the reference product Humalog®, thereby providing a reliable basis for successful approval and subsequent clinical use.

Introduction. The neurodegenerative disorder known as Alzheimer’s disease progresses over time as it causes damage to different parts of the brain. Ranolazine, a piperazine derivative, is a treatment that is considered to be of secondary relevance for persistent aortic stenosis in individuals with stable angina who do not react to other drugs. Additionally, famotidine is a competitive H2-receptor antagonist that reduces stomach acid secretion and treats conditions such as acid reflux and ulcers. In mouse models, ranolazine may help protect the brain against Alzheimer’s disease-like characteristics induced by scopolamine.

Aim. To examine the potential neuroprotective properties of the combined administration of ranolazine and famotidine in mitigating symptoms of Alzheimer’s disease in a mouse model induced by scopolamine.

Materials and methods. The experiment included four groups of ten mice each: a control group, an induction group that received 1 mg/kg of scopolamine intraperitoneally once a day for seven days to mimic Alzheimer’s disease symptoms, and a memory loss group that received no such medication. For the remaining two groups of mice, the following medications were given orally once daily: donepezil (5 mg/kg/d) and a combination of ranolazine (40 mg/kg/d) and famotidine (40 mg/kg/d). After 14 days of prophylactic medication, the induction was performed with scopolamine (1 mg/kg i.p. once daily), and the medication was continued for an additional week. Research on the brain tissue sample included histopathological examinations, evaluation of inflammatory cytokines and oxidative stress parameters (such as acetylcholinesterase concentration), and assessment of behavioral parameters (such as novel object recognition and Y-maze tests).

Results and discussion. Scopolamine significantly impaired behavior (Y-maze 53.72, p ≤ 0.001), which was reversed by treatments (donepezil: 67.58, p ≤ 0.001). Treatments significantly reduced oxidative stress (MDA 1.65, p < 0.01) and inflammation (TNF-α 125.91, p ≤ 0.001) compared to the induction group. All treated groups showed no significant difference compared to controls (p > 0.05). In comparison to the scopolamine group, ranolazine and famotidine combination significantly improved behavioral and memory performance, oxidative stress parameters, and inflammatory cytokine levels. However, there was no significant reduction in the concentration of acetylcholinesterase in brain homogenate.

Conclusion. Ranolazine and famotidine protected mice from scopolamine-induced AD-like symptoms in this study. The recent investigation showed that ranolazine and famotidine's antioxidant and anti-inflammatory actions may explain this benefit.