Kinetic instability of a Chitosan – Aspartic Acid – Water System as a Method for Obtaining Nano - and Microparticles Chimica Techno Acta ARTICLE published by Ural Federal University 2021, vol. 8(4), № 20218405 eISSN 2411-1414; chimicatechnoacta.ru DOI: 10.15826/chimtech.2021.8.4.05 1 of 8 Kinetic instability of a chitosan – aspartic acid – water system as a method for obtaining nano- and microparticles T.N. Lugovitskaya a* , A.B. Shipovskaya b, X.M. Shipenok b a: Ural Federal State University named after the first President of Russia B. N. Yeltsin, 620002 Mira st., 19, Yekaterinburg, Russia b: Saratov National Research State University named after N.G. Chernyshevsky, 410012 Astrakhanskaya st., 83, Saratov, Russia * Corresponding author: tlugovitskaja@mail.ru This article belongs to the regular issue. © 2021, The Authors. This article is published in open access form under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). Abstract The specific electrical conductivity and dielectric constant of aqueous solutions of ionic aminopolysaccharide chitosan in L-aspartic acid were investigated. An increase of the mobility of charge carriers in these solutions was found in comparison with solutions of an ind i- vidual acid. The evaluation of the kinetic stability revealed that the viscosity, electrical conductivity and dielectric constant of the ch i- tosan – L-aspartic acid – water system decrease, while the pH value increases. It was shown that the time variation of physicochemical and electrochemical parameters is due to the effects of counterionic association with the transition of macromolecules to the ionomeric state and is accompanied by phase segregation of the polymer phase in the form of nano- and microparticles. The conducted studies car- ried out have shown the fundamental possibility of controlling the metastable state of this system in order to obtain nano- and micro- particles. Keywords chitosan L-aspartic acid counterionic association nanoparticles Received: 06.10.2021 Revised: 20.11.2021 Accepted: 23.11.2021 Available online: 26.11.2021 1. Introduction Chitosan (CTS), a product of partial or complete deacetyla- tion of chitin, is an ionic copolymer of aminopolysaccharide. Since the temperature of thermal decomposition of CTS is lower than its melting point, the processing of this polymer into the final products involves the step of dissolving in or- ganic or inorganic acids. In an aqueous acidic medium at pH below pKa, the amino groups of CTS are protonated, resulting in the formation of a salt form of the polymer – a positively charged polyelectrolyte (~ − NH3 + ) with ionic conductivity. The main contribution to the transfer of electricity in an aqueous acidic CTS solution is made by free counterions, as well as excess of hydronium ions [1–3]. In this case, the na- ture of the acid-solvent of CTS significantly affects the quan- titative characteristics of electrical conductivity, which de- termines the conformation of macromolecules in solution and the phase state of the polymer system after removal of the liquid medium [1]. Thus, in strong HCl, a linear decrease in the specific electrical conductivity (æsp) with an increase in the CTS concentration (СCTS) is observed. The conformation of the expanded helix is realized due to the repulsion of charged macrochains. In weak acids (propionic, acetic and formic), the concentration dependence of the specific electri- cal conductivity is nonlinear, the æsp values grow increase with an increase in CCTS value. There is an attraction and “adhesion” of local sections of macrochains through ionic crosslinking ~ − NH3 + with R − COО− and H-bonds of OH- groups of CTS with the carbonyl oxygen of the acid, as a re- sult of which the macromolecules assume the coiled confor- mation. CTS solutions in HCl have a weak film-forming abil- ity and give a loose structure with a pore diameter of more than 10 mm when dried on an inert surface. CTS films cast from aqueous solutions of carboxylic acids are characterized by a dense ordered structure. When an electric field is applied in an aqueous acidic CTS solution, internal polarization phenomena can occur, i.e., displacement of charges that create a volume- distributed dipole moment. The relaxation processes of the electric polarization of the CTS salt form are manifested in dielectric spectra in the frequency range below 1010 Hz [4]. Like conductometry, dielectric relaxation spectroscopy is highly sensitive to changes in the electrical conductive properties of aqueous acidic CTS solutions and makes it possible to reliably control the state of counteri- ons (free, bound) in them. http://chimicatechnoacta.ru/ https://doi.org/10.15826/chimtech.2021.8.4.05 http://creativecommons.org/licenses/by/4.0/ https://orcid.org/0000-0002-8286-0711 Chimica Techno Acta 2021, vol. 8(4), № 20218405 ARTICLE 2 of 8 CTS macromolecules are characterized by highly co m- plex behavior in solutions. This is primarily due to the polyelectrolyte swelling of charged macromolecular coils and increased chain rigidity. CTS solutions are also char- acterized by instability of viscosity properties over time, which is usually explained by the destruction of macro- molecules as a result of acid hydrolysis of glycosidic bonds [5–7]. However, there are studies that make it possible to exclude destruction as the main reason for the decrease in the viscosity of chitosan solutions in time [8, 9] and to explain this effect by conformational rearrangements o f macromolecules [10], changes in the supramolecular structure, including aggregation [9, 11], and also by phase separation of the system as it is stored [11, 12]. CTS is widely used due to a complex of valuable proper- ties, as well as an annually renewable resource base [13–15]. Chitosan-containing nanomaterials, in particular nanoparti- cles, deserve special attention [16, 17]. Compared to the orig- inal CTS, they acquire additional valuable properties due to their small size, increased surface area, and quant um size effects. Like CTS, chitosan-containing nanoparticles are non- toxic, biocompatible, and biodegradable, which predeter- mines their use in medicine, as well as in the food, textile, and cosmetic industries [18–22]. For obtaining nanoparticles, the most common used are spray drying under special condi- tions, or methods of ionotropic gelation and emulsion cros s- linking with tripolyphosphate, glutaraldehyde, genipin [16]. In some cases, dibasic and tribasic pharmacopoeial acids (malic, tartaric, citric, and succinic) were used, which simul- taneously act as a solvent for the polymer and as a cross - linking reagent [23–25]. It should be noted that the use of biologically active acids for the formation of CTS nanoparti- cles significantly expands the directions of t heir application in biomedical and pharmacological applications [26–28], vet- erinary medicine [29], and plant growing [30–33]. The use of L-aspartic acid (AspA), a proteinogenic ami- no acid that plays an important role in a living organism, seems to be promising [34, 35]. Unlike traditional water- acid CTS dissolving media based on monobasic carboxylic acids (HCOOH, AcOH), AspA contains two carbo xylic and one amino groups. Therefore, in an aqueous solution, de- pending on the pH of the medium, its molecules can exist in several ionic forms. At pH = 3.0, the isoelectric point is located, at pH = 4, almost equal proportions of zwitter ions (Н2Asp) and aspartate anions (НAsp−) are realized, whereas in the range of pH = 4.7–8.0, НAsp− is the pre- dominant ion [36]. The resulting high concentration of hydrogen ions (hydronium), which are the main carriers of electricity, provides prototropic conductivity of an aqueous solution of AspA [37, 38]. Previously, we have found that the solubility of AspA in Н2О increases in the presence of CTS [39]. The conditions for the preparation of CTS solutions in AspА – Н2О mixture and some of their properties have also been dis- cussed [40]. Thus, in an aqueous medium, the macromole- cules of the salt CTS form exhibit the properties of a weak- ly dissociating polyelectrolyte with a partially compen- sated charge. The calculated Huggins constants and tem- perature viscosity coefficients showed an increased rigidi- ty of their macrochains. However, kinetic stability was not evaluated, although the CTS – AspA – H2O system showed all the characteristic features of phase separation. As in the case of synthetic polyelectrolytes [41, 42], not only the dissociation of ionogenic groups of CTS aspartate can oc- cur in an aqueous medium, but also the interaction of the polycation with HAsp– with the formation of ion pairs, multiplet structures (combination of ion pairs) and the transition of macromolecules to the ionomeric state. At the initial stage of phase separation, these processes should be accompanied by phase segregation of the polymer phase in the form of nano- and microparticles. This assumption served as the basis for this study. The aim of this work is to study the kinetic stability of the CTS – AspA – H2O system during storage to obtain nano- and microparticles using viscometry, potentiometry, con- ductometry, and dielectric relaxation spectroscopy as the most informative methods for studying the counterions state (free, bound) in polyelectrolyte solutions [4, 40–42]. 2. Materials, objects and methods 2.1. M aterials The starting reagents were powdered CTS with a viscosity - average molecular weight M̅η = 200 kDa, a degree of deacetylation of 82 mol%, and a moisture content of W = 8±1 wt% (ZAO Bioprogress, RF); powdered L-AspА of pharmacopoeial purity, obtained by biocatalytic synthesis using E. coli strain VKPM 7188 (JSC Bioamid, RF); distilled water (pH = 6.0); ethyl alcohol (95.6%), and acetone (99.8%). CTS and AspA were used without additional puri- fication; all other reagents were of analytical grade. 2. 2. P reparation of objects of study A working AspA solution with a concentration of СAspА = 0.06 mol/L was prepared by dissolving a weighed portion of AspA powder in distilled water at 80 °C, followed by cooling down to room temperature at a rate of 20 °C/h. A series of solutions of lower concentrations were obtained by successive dilution of the working solution with distilled wa- ter. The pH values of AspA solutions in Н2О with СAspА = 0.003–0.060 mol/L were in the range of 3.1–4.2. Freshly prepared acid solutions were used in all experiments. To obtain system CTS + AspA + Н2О, calculated weighed portions of CTS (considering the moisture sample) and AspA were placed into some volume of Н2О and stirred on a magnetic stirrer at a rotation speed of 400–500 rpm at 25 °C until visual dissolution of the powders (for 2–3 h). The concentrations of the polymer and acid in the solution were varied in the ranges СCTS = 0.002–0.035 monomol/L and СAspА = 0.03–0.06 mol/L, respectively. Before experi- ments, all systems were filtered through a Millipore filter with a pore diameter ≤0.45 μm. Chimica Techno Acta 2021, vol. 8(4), № 20218405 ARTICLE 3 of 8 The CTS + AspA + Н2О system were stored in a room atmosphere (760 mm Hg, 22±2 °C) for 96 h, regularly tak- ing samples to assess their physicochemical parameters. In all experiments, the conditions for the preparation of the system and the time at which the physicochemical charac- teristics began to be measured were the same. The stability of the systems was assessed by measuring their conductivi- ty, pH, viscosity and permittivity, as well as by SEM. 2. 3. Obtaining p articles To obtain nanoparticles and microparticles, 0.5 μL of a freshly prepared or stored CTS + AspА + Н2О system was sprayed onto a degreased (ethyl alcohol, acetone) glass surface using a mechanical syringe-shaped injector and dried for 1–2 h (the technique is similar [43]). The ob- tained particles obtained were dried at 22±2 °C under at- mospheric pressure to an air-dry state, and analyzed using SEM. 2.4. М e thods Gravimetric measurements were carried out on an Ohaus Discovery analytical balance (USA), weighing accuracy being ±0.0001 g. pH was measured on a Mettler Toledo Five Easy FE20 pH-meter (MTD, Singapore). Conductivity was measured on a WTW inoLab Cond 7110 conductometer (Germany) at 25 °C. A thermostated cell with a volume of 25 mL was used. Specific conductiv i- ty (æsp, S·m−1) was calculated using the specific conductiv- ity of water by the formula: æsp = 𝐾 𝑅⁄ − æH2O = æCTS − æH2O, (1) where К = 0.1213 m−1 is the cell constant evaluated from 0.01 N KCl solution; R is the resistance of the system, S; æCTS and æН2О are the specific conductivity of the chitosan- containing system and water, respectively, S·m−1. The viscometric measurements were carried out in an Ubbelohde viscometer (RF) with a capillary diameter of 0.56 mm at 25 °C. The relative viscosity (rel) was calcu- lated from: rel = 𝑡 𝑡0 ⁄ , (2) where t is the flow time of the CTS + AspА + Н2О sistems, s; t0 is that of the AspA aqueous solution, s; СCTS is ex- pressed in g/dL; the accuracy of measuring t and t0 was ±0.1 s. The frequency dependences of the real (εʹ) and imagi- nary (εʹʹ) parts of the complex permittivity were measured on an Agilent Microwave Network Analyzer PNA-X N5242A vector network analyzer (USA) using an Agilent 85070E coaxial probe in the frequency range f = 107–1011 Hz at 25 °С. The dielectric loss tangent was calculated from the ratio tgδ = εʹʹ/εʹ. SEM images were taken on a MIRA\\LMU scanning elec- tron microscope (Tescan, CZ) equipped with an energy dis- persive detector (EDX) at a voltage of 30 kV and a conduct- ing current of 400 pA. A 5 nm thick gold layer was deposit- ed on each sample using a K450X Carbon Coater (DE). 2. 5. Statistical analysis In each experiment on the study of physicochemical proper- ties, at least 3 parallel experiments were carried out; the arithmetic mean and standard deviation were calculated. 3. R esults and discussion We have previously shown that the concentration depend- ence of the reduced specific viscosity of a freshly prepared aqueous solution of CTS aspartate is not linear, has a max or a plateau and a descending branch with a decrease in CTS [40]. This hydrodynamic behavior indicates the im- plementation of a mixed polyelectrolyte-ionomer mode, when some of the HAsp– counterions are in an associated (bound) state with −NH3 + groups of the macrochain with the formation of ion pairs (Fig. 1a). The revealed features of the counterions state in mac- romolecular coils of the CTS salt form were proved by studying the electrochemical properties of the polymer system. Thus, the specific electrical conductivity of a CTS solu- tion in AspА – Н2О at СCTS = const increases with an in- crease in the acid concentration (Fig. 2a, curve 1). A simi- lar dependence is observed for aqueous solutions of indi- vidual AspA (curve 2). (a ) (b) (c ) (d) Fi g. 1 Distribution of free and bound counterions in the CTS – AspА – Н2О system during storage: (a) polycation with a partially com- pensated charge, (b) ion pairs, (c) multiplets, (d) phase segregation of the polymer phase in the form of a nanoparticle Chimica Techno Acta 2021, vol. 8(4), № 20218405 ARTICLE 4 of 8 The revealed features of the counterions state in mac- romolecular coils of the CTS salt form were proved by stud- ying the electrochemical properties of the polymer system. Thus, the specific electrical conductivity of a CTS solution in AspА – Н2О at СCTS = const increases with an increase in the acid concentration (Fig. 2a, curve 1). A similar dependence is observed for aqueous solutions of individual AspA (curve 2). It is noteworthy that at СAspA < 0.01 mol/L, the increase in æsp values of both AspА – Н2О and CTS solutions in AspА – Н2О is practically proportional to the increase in СAspA. In this case, despite the protonation of the amino groups of the macrochains in an aqueous acidic medium and, as a consequence, the binding of the labile H+ of the carboxyl group to the NH2 moiety of the polymer, the æsp values of aqueous solutions of CTS aspartate and AspA are the same. Taking into account that the electrical conductivi- ty is determined by the number of charged particles and their mobility [1–3], this character of the dependence æsp = f(СAspA) for a polymer solution can be explained by shifting acid-base equilibrium towards the acid in the pres- ence of CTS (base). As a result of ionization of previously undissociated AspA molecules, the concentration of H+ and HAsp– ions increases, and the total amount of free ions (electricity carriers) in solutions of CTS in AspA – H2O and AspA in H2O remains constant. At СAspA > 0.01 mol/L, the rate of increase in the specific conductivity of the CTS solu- tion slightly decreases and the æsp values become lower than that of individual AspA, which may be due to a de- crease in the degree of acid dissociation as it is concentrat- ed in the solution. However, an increase in æ sp value with an increase in the acid concentration in this CAspA range also indicates the intensification of AspA protolysis in an aque- ous acid solution of CTS, although to a lesser extent com- pared to the range of CAspA < 0.01 mol/L. The proposed ex- planation of the associative-dissociative processes is also confirmed by the data in Fig. 2b, if we consider the depend- ence æsp = f(СCTS) at СCTS = const. In addition, a significant increase in the specific electrical conductivity with an in- crease in СCTS at СAspA = const indicates an increase in the mobility of conductive particles not only in comparison with an aqueous solution of AspA, but also in comparison with Н2О. Note that a nonlinear increasing dependence æsp = f(CCTS) has also been observed in CTS solutions in weak propionic, acetic, and formic acids [1]. The evaluation of the dielectric properties also showed an increase in the mobility of the electrically transporting parti- cles in the CTS aspartate solution. It is noteworthy that in the frequency range of f < 109 Hz, the real part of the complex dielectric constant of CTS solutions in AspA–H2O mixture is less than εʹ of an aqueous solution of AspA and water (Fig. 3a), and the dielectric loss factor and dielectric loss tan- gent are larger (Fig. 3b, c). The εʹ values decrease, εʹʹ and tgδ values increase with an increase in СCTS. Consequently, in CTS solutions in AspA–Н2О mixture there is a larger amount of mobile charge carriers providing ohmic current than in AspA–Н2О and aqueous solutions. At f > ~1.6–1.8·109 Hz, the εʹ, εʹʹ, and tgδ values of aqueous solutions of CTS aspartate, AspА and water are comparable. This character of the disper- sion of εʹ, εʹʹ, and tgδ values of a solution of CTS aspartate is explained by the fact that at high frequencies a “fast” mecha- nism of ion polarization is realized, the large sizes of macro- coils prevent the movement of macrodipoles (associations of polar groups of a polymer molecule through dipole-dipole interaction) in an alternating electric field and the electric the induction of the system is determined only by the mobili- ty of low molecular weight ions [4]. As f value decreases, the “slow” mechanism of orientational (dipole) polarization oc- curs, and the possibility of the macrodipoles motion, accom- panied by dissipative losses, appears. The smaller f values, the greater are the dielectric losses (εʹʹ, tgδ), since the macrodipoles have more time to orient themselves along the electric field. Fi g. 2 Dependence of the electrical conductivity of CTS solutions in AspА–Н2О mixture with СCTS = 0.015 monomol/L (1а) and AspА aqueous solution (2а) on AspА concentration, as well as CTS solu- tions in AspА–Н2О mixture with СAspA = 0.030 (1 b), 0.045 (2b) and 0.060 mol/L (3b) on CTS concentration; the dotted line shows æsp value for water (b) (a) Chimica Techno Acta 2021, vol. 8(4), № 20218405 ARTICLE 5 of 8 Fi g . 3 Dispersion of the real part of the complex dielectric con- stant (a), dielectric loss factor (b) and dielectric loss tangent (c) of CTS solutions in AspA – Н2О mixture with СAspA = 0.030 mol/L and СCTS = 0.010 (1), 0.018 (2) and 0.035 monomol/L (3), as well as AspA – Н2О mixture with СAspA = 0.030 (4) and water (5) The high mobility of charged particles in a solution of CTS aspartate was reflected in the kinetic stability of the polymer system. It turned out that exposure at room at- mosphere is accompanied by a significant decrease in the relative viscosity of CTS solutions in AspA–Н2О mixture and an increase in the medium acidity (Fig. 4a, curves 1 and 2). These changes are most intense in the first ~ 16–18 hours after preparation of the solution and increase with increasing CTS concentration. So, after 24 hours of storage of solutions with CCTS = 0.002–0.006 monomol/L, the effect of reducing ηrel was 7–10%, and 23% (Fig. 4а, curve 1) for more concentrated solutions, for example, with CCTS = 0.035 monomol/L. In addition, during storage for ~16 – 18 hours in a vis- ually homogeneous solution of CTS in AspA – Н2О mixture, the specific conductivity increases (Fig. 4b, curve 3). The dielectric constant in the frequency range of f < 109 Hz decreases, and in the range of f > 1.8·109 Hz it increases with a local maximum (curves 4 and 5). At t > 24 hours, opalescence appears æsp value begins to decrease with the realization of lower values than in a freshly prepared solu- tion; εʹ value decreases with time over the entire range of f. Like ηrel value, in this case, the æsp and εʹ values change to the greater extent, the higher the polymer concentra- tion in the system. After ~66–74 hours the phase separa- tion occurs, which is observed visually as the precipitation of a water-insoluble fine sediment. Like relative viscosity, specific electrical conductivity and dielectric constant change with time to the greater extent, the higher the polymer concentration, and pH – vice versa. For example, for a system with СCTS = 0.035 monomol/dL, during ~50–60 days of storage the values of these parame- ters change in the range of æsp = (92–88)·10−5 S·m−1, εʹ = 38–22 (f = 1.2·108 Hz), рН = 4.9–5.0, and for a system with СCTS = 0.002 monomol/dL − æsp = (60–58)·10−5 S·m−1, εʹ = 72–64 and рН = 3.1–3.2. The shortest time interval be- fore the precipitation of the polymer phase is also observed at a higher СCTS. At first glance, a decrease in the viscosity of the system under study with time may be associated with the macro- molecules destruction as a result of acid hydrolysis of gly- cosidic bonds [5–7], and an increase in pH is due to depro- tonation of AspA with the transition of some of the acid molecules into the zwitterionic H2Asp form. However, up- on destruction with a decrease in the molecular weight of CTS, the amount of free counterions and hydronium ions should increase, and the pH value should decrease. As a result, the destructive processes should lead to an increase in ionic conductivity, i.e., electrical conductiv ity and dielec- tric constant should increase monotonically, but not very significantly. Since for the CTS – AspA – Н2О polyelectrolyte system under study a different character of the æsp and εʹ kinetics is observed, it seems that its instability in time is associated with the effects of the association of counterions with protonated amino groups of the polycation and their further transformations. Probably, it is thermodynamically not favorable for counterions to be in a free state, and they continue to form ion pairs with –NH3+ groups of polymer chains (ionomers), while losing in entropy, but gaining in electrostatic energy (Fig. 1b). (a) (b) (c) Chimica Techno Acta 2021, vol. 8(4), № 20218405 ARTICLE 6 of 8 Fi g . 4 Kinetic dependences of the relative viscosity (1a), pH (2a), specific electrical conductivity (3b) and dielectric constant at f = 1.2·108 (4b) and 1010 Hz (5b) of the CTS – AspA – Н2О system with СCTS = 0.035 monomol/L, СAspА = 0.030 mol/L The transition of counterions from the aqueous medi- um to the macrocoil volume leads to screening of electro- static repulsion between the like-charged links of the macrochain, a decrease in æsp, ηrel, and εʹ values of the system and, accordingly, its transition to the ionomeric mode. The resulting effective attraction between mono- mer units contributes to the dipole-dipole interaction of ion pairs and their combination into multiplet structures (Fig. 1c). Most likely, the stabilization of the latter occurs through complex ion-ion-hydrogen contacts of ionogenic groups of the CTS aspartate macromolecule and the AspA molecule (the possibility of the formation of such CTS– acid contacts has been reported in [1]). Multiplets func- tion as physical crosslinks between different polymer chains, which contributes to the compaction of macro- coils and the formation of nanosized nuclei of a new phase (Fig. 1d). Upon aggregation of the latter to micro- and macroparticles, the metastable polymer system is divided into two equilibrium phases: the polymer-rich phase precipitates, the polymer-depleted phase is repre- sented by the supernatant liquid. According to [41], phase separation in a polyelectrolyte solution under con- ditions of ionic association can occur even in a thermo- dynamically good solvent. Fi g. 5 SEM image of particles isolated from the CTS–AspA–Н2О system with СCTS = 0.035 monomol/L, СAspА = 0.030 mol/L after 24 (a) and 36 hours (b) (b) (a) (b) (a) Chimica Techno Acta 2021, vol. 8(4), № 20218405 ARTICLE 7 of 8 The described associative processes were confirmed and visualized using scanning electron microscopy. For example, in a CTS – AspA – Н2О sample, spherical nanoparticles with a size of d = 40−90 nm (Fig.5a) and ellipsoidal ones with a size of d = 70−260 nm (Fig. 5b) were revealed after 24 and 36 hours, respectively. After ~48–70 hours, the average size of chitosan-containing particles was ~1.4–2.0 μm, width was ~0.6–0.8 μm; their aggregation was also observed. Elemental analysis data showed that the nanosized polymer phase contains 42.95±1.2% carbon, 21.09±0.9% nitrogen, and 35.96±1.0% oxygen. This suggests that the nanoparticles are represented by the polymer salt form, CTS aspartate. 4. Conclusions In conclusion, cooperative interactions between fixed charges of the polycation and low-molecular counterions in the CTS – AspA – Н2О system are accompanied by the formation of ion pairs, multiplet structures, and phase segregation of the polymer substance to form nano - and microparticles. The "fixation" of the nuclei of a new phase at the initial stage of phase separation by injection spray- ing onto a glass support makes it poss ible to visualize the formation of particles by the SEM method. Further search for ways to stabilize particles in solution, for example, by creating a “shell” that prevents aggregation, will make it possible to develop methods for their preparation in a preparative amount. Due to the high biological activity of CTS and AspA, chitosan-containing nano- and microparti- cles can be promising as independent pharmaceutical and veterinary drugs, plant protection products, providing synergy of useful properties of their constituent compo- nents, as well as in drug storage and delivery sy stems, and theranostics. Acknowledgements This work was financially supported by the Foundation for Assistance to Innovations of the Russian Federation (Grant No. 16317GU/2021). R eferences 1. Li Q, Song B, Yang Z, Fan H. Electrolytic conductivity behav- iors and solution conformations of chitosan in different acid solutions. Carbohydr Polym. 2006;63(2):272–282. doi:10.1016/j.carbpol.2005.09.024 2. Bobreshova OV, Bobylkina OV., Kulintsov PI, Bobrinskaya GA, Varlamov VP, Nemtsev SV. Conductivity of aqueous solutions of low-molecular chitosan. Russ J Electrochem. 2004;40(7):694–697. doi:10.1023/B:RUEL.0000035250.54523.e8 3. Osman Z, Ibrahim Z, Arof A. 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