ASSESSMENT OF BIOLOGY ACTIVITY OF THE PEELING SUBSTANCES BY THE PHYSICOCHEMICAL APPROACHES ON THE SPIROSTOMUM AMBIGUUM CELL MODEL Original Article USPENSKAYA E. V., * Peoples Friendship University of Russia (RUDN University), Department of Pharmaceutical and Toxicological Chemistry, 6, Miklukho- Maklaya St, Moscow, 117198, Russian Federation *Email: PLETENEVA T. V., PHAM MY HANH, KAZIMOVA I. V. uspenskaya75@mail.ru Received: 28 Apr 2021, Revised and Accepted: 01 Jun 2021 ABSTRACT Objective: To evaluate the biological activity of chemical peeling substances based on enzymatic and Arrhenius kinetics using Spirostomum ambiguum as an alternative approach to animal experiments. Methods: The Spirotox method was used to analyze the mechanism of «xenobiotic-cell» interaction, similar to the Michaelis-Menten enzymatic kinetics. The Hill-Langmuir equation was used to determine the degree of cooperativity in the binding of xenobiotics to cellular receptors. Using the Arrhenius kinetics, the observed activation energy obsEa of cell death in the model solutions of glycolic and carbolic acids was determed, which will allow predicting the toxicity parameters of any peeling substances. Results: The relationship Spirostomum ambiguum lifetime t L- lgC concentration of peeling compound solution made it possible to characterize the moment of cellular transition from the intermediate state C•L n to the dead state DC, characterized by irreversible structural and functional changes in the cell/death. The values were 5.3 mmol•l -1 for glycolic acid solutions and 2.8 mmol•l -1 for carbolic acid solutions. Equilibrium constants Keq of complexation, the rate of infusoria death f m, and the degree of ligand cooperativity n were calculated. The activation energy ° bs E a of cell death was determined in Arrhenius coordinates, which were 210±0.39 kJ·mol -1 and 108±0.09 kJ·mol -1 for glycolic and carbolic acids respectively. The correlation between the values of activation energy and DL 50 Conclusion: The obtained kinetic parameters made it possible, without animals and humans testing, to characterize the mechanisms of interaction of peeling substances with the living cell. of mammals (rats) was discovered. Keywords: Peeling, Glycolic acid, Phenol, Spirotox method, Biological activity, Animal-free test © 2021 The Authors. Published by Innovare Academic Sciences Pvt Ltd. This is an open access article under the CC BY license (https://creativecommons.org/licenses/by/4.0/) DOI: https://dx.doi.org/10.22159/ijpps.2021v13i7.41927. Journal homepage: https://innovareacademics.in/journals/index.php/ijpps. INTRODUCTION Chemical peeling (сhemexfoliation) stimulates the appearance of controlled keratocoagulation and denaturing proteins in the epidermis and dermis, the release of pro-inflammatory cytokines and chemokines, and the production of new skin collagen and elastin [1-5]. The clinical effectiveness of chemical peeling, as well as the risk of various complications, depends on many factors: the properties of chemicals, concentration, regimen, and the number of uses, skin type, dermatological condition, and cumulative dose [6, 7]. Despite available reviews about the traumatic nature of this procedure, chemical peeling ranks third in the frequency of non-invasive cosmetic procedures in the USA [8]. Since the use of cosmetic products is not associated with an urgent need for the life and health of consumers, there is the number of advocates of alternative in vitro approaches to animal experiments, for example, when checking the safety of cosmetics [9- 14]. Cosmetic animal testing is banned in a few countries [15]. For the testing of cosmetics, programs are being developed to enable the cosmetics industry to conduct safety assessments [16, 17]. To date, testing methods without the use of animals have been selected, including cell lines [18]. Alternative models use various cellular cultures, specialized moving cells, bacteria, single-celled organisms, primitive crustaceans and other hydrobionts. Methods based on the reactions of the simplest animals are of paramount importance in today's environment, as they allow determining the possible range of properties in a short period the substance under study and making recommendations on whether to investigate it further. For example, there are approaches to describing the kinetics of the development of cellular population under the influence of inhibitors and promoters [19]. It is also important that biological research methods using protozoa significantly reduce the time required to study the properties of substances and are characterized by a relatively low cost. Replacing warm-blooded animals with unicellular ones reduces the time of toxicological and pharmaceutical studies from one year to a month, and the cost of their implementation decreases in ten times [20]. The goal is to directly replace animal testing with non-animal methods. Advances in cell and molecular biology and informatics need to be leveraged to develop new specific preclinical testing strategies that are applicable to specific human situations [21]. This work aims to evaluate the biological activity of chemical peeling substances using enzymatic and Arrhenius kinetics for a single-cell model Spirostomum ambiguum as an alternative approach to animal experiments. MATERIALS AND METHODS Cell model The test culture Spirostomum ambiguum (Sp. ambiguum) has been cultured in the laboratory to carry out studies of individual and combined biological activity of medicines. The protozoan ciliate Sp. ambiguum is characterized by tape-shaped, dorsal body shape (1-3 mm long), has a macronucleus clear-shaped and mouth apparatus up to the back third of the body (fig. 1). Compared to other objects of biological testing, Sp. ambiguum have several advantages, since they are eukaryotic organisms. The statistically reliable sensitivity to toxicants makes it possible to compare the response of protozoa with that of humans [23, 24]. Under favorable conditions in a low-mineralized environment, cells do not die for a period exceeding their cell cycle (about 20 h). When incubated into a toxicant solution, cells die over time, which is a function of both concentration and temperature. Kinetic scheme of ligand-induced death of Sp. ambiguum includes the reversible reaction with the formation of an intermediate product (C • L n ) and the state of transition to cell death (DC) (fig. 2). International Journal of Pharmacy and Pharmaceutical Sciences Print ISSN: 2656-0097 | Online ISSN: 0975-1491 Vol 13, Issue 7, 2021