Anastasiya I. Varlamova’s research while affiliated with All-Russian Scientific Research Institute of Fundamental and Applied Parasitology of Animals and Plants named after K.I. Skryabin and other places

Background Due to the fact that animal parasitic diseases often occur in the form of mixed infections, it is necessary to use complex drugs with a combination of active substances that act against different classes of helminths. Fenbendazole has a wide spectrum of nematocidal action and is less effective against cestodes. Considering that sheep are often infected with both gastro-intestinal nematodes and cestodes, a complex drug based on fenbendazole, niclosamide with polyvinylpyrrolidone polymer, was developed using mechanochemical technology in order to increase its solubility and bioavailability and reduce the dosages and side effects. Objective This study aimed to evaluate the influence of mechanochemical technology on the efficacy of complex solid dispersions of anthelmintics against cestodoses and nematodoses. Methods The influence of different types of mills (roller and planetary mills), levels of energy intensity (20, 40, 60 g), and component ratio (2:20:78; 3:30:67; 4:40:56) during mechanochemical processing was studied on the anthelmintic efficacy of complex solid dispersions of fenbendazole, niclosamide, and polyvinylpyrrolidone in laboratory models of trichinellosis and hymenolepiosis of white mice, and on sheep naturally infected with helminths. Results The efficacy of complex solid dispersions obtained in the planetary mill was higher than the activity of dispersions obtained in a roller mill. It showed 87.5, 96.36, 83.77, and 99.39% efficacy, respectively, against experimental trichinellosis, hymenolepiosis of white mice, gastrointestinal nematode infection and monieziosis of sheep at a ratio of 2:20:78. The basic drug – fenbendazole demonstrated 31.37 and 22.77% activity against experimental trichinellosis and gastro-intestinal nematode infection of sheep. The basic drug, niclosamide, showed 36.37% efficacy against hymenolepiosis in mice and 28.31% against monieziosis in sheep. It was established that with an increase in the energy intensity level of the planetary mill (20, 40, and 60 g), the decrease in the efficacy of solid dispersion occurred from 73.90 to 59.12% against T. spiralis and from 92.73% to 79.81% against H. nana. Conclusion The use of mechanochemical technology for the production of solid dispersions of fenbendazole and niclosamide with polyvinylpyrrolidone makes it possible to increase the anthelmintic efficacy by 2.7–3 times compared to the activity of basic substances. It was noted that the complex solid dispersions of anthelmintics obtained in a planetary mill had 7.51–10.17% greater activity in comparison with the samples obtained in a roller mill. The most optimal ratio of active substances was 2:20:78 at a 20 g level of energy intensity of the planetary mill.

The substance fenbendazole is included in the composition of many anthelmintic drugs, in which the “chemical stability” parameter is one of the main characteristics when obtaining permission for the use of drugs in veterinary practice. Fenbendazole is characterized by low solubility in water and therefore the content of the substance is overestimated in its preparations, which increases the cost of the drug as well as the safety risks of pharmacotherapy. The possibilities of mechanochemical modification of fenbendazole were evaluated in order to improve the solubility index. During the mechanical processing treatment of the substance in the presence of polymeric substances, solid dispersions are formed, which have increased solubility and high anthelmintic activity. The inclusion in these dispersions of the third component, which is succinic acid, did not significantly change the solubility of fenbendazole. In all these dispersions, the substance remained unchanged both during the preparation of its solid dispersions and during their storage. When fenbendazole is modified in an organic solvent medium, the substance is partially converted into oxfendazole, which is one of its metabolites. The chemical stability of fenbendazole was confirmed via HPLC/MS and NMR spectroscopy. The anthelmintic activity of these compositions was evaluated and it was found that they have a high nematicidal activity.

Two new multicomponent crystalline phases of fenbendazole (FNB), a benzimidazole anthelmintic agent, with maleic and oxalic acids have been prepared, and their structural and physicochemical properties carefully investigated. The crystal structures of the solid forms have been determined from powder X-ray diffraction data. The positions of dynamic hydrogen atoms have been further refined via dispersion-corrected density functional theory calculations, which validated the salt nature of the resulting solid forms by demonstrating proton transport from the corresponding acids to the FNB molecule. The in vitro dissolution performance of the solid forms in aqueous media at different pH values, as well as the in vivo anthelmintic efficacy of fenbendazole on the laboratory model of Trichinella spiralis infection in mice have been evaluated and compared to that of the previously reported salt of FNB with p-toluenesulfonic acid. A relationship between the in vitro dissolution characteristics and the in vivo therapeutic action has been revealed and discussed

A supramolecular complex of fenbendazole (SFBZ) with polyvinylpyrrolidone (PVP) was created by mechanochemical processing to increase its anthelmintic efficacy and to reduce the dose of applied drugs. The aim of our research was to study the pharmacokinetic profile and tissue residue depletion of fenbendazole (FBZ) and its metabolites: sulfoxide and sulfone in sheep after SFBZ treatment by high-performance liquid chromatography with tandem mass spectrometric detection and to evaluate its efficacy against gastrointestinal strongylatosis of sheep in field trials. The results revealed that FBZ and its metabolites were detected in blood serum in 2 hours after SFBZ administration and in 4-6 hours after the administration of the basic – FBZ. Pharmacokinetic parameters of SFBZ and its metabolites were characterized by higher rate of absorption, concentration of the drug and longer retention times in the blood serum. The maximum concentration of FBZ and its metabolites was detected on the 3rd day in the organs and tissues of sheep that received SFBZ. Thus, in the liver, the content of FBZ was 4878.0 ng/g, sulfoxide and sulfone - 18682.4 and 2483.6 ng/g respectively while the indicators of the basic FBZ and its metabolites were tenfold lower. FBZ and its metabolites were not detected in the organs and tissues of sheep on the 16th day in animals treated with the basic drug and on the 21st day after SFBZ administration. In field trials SFBZ demonstrated a high anthelmintic activity against nematodosis of sheep. It showed 98.2% efficacy against nematodirosis and 99.0 % against other types of gastrointestinal strongylatosis at a dose of 2 mg/kg of active substance (a.s.). Efficacy of mechanical mixture and efficacy of FBZ substance was in 3.1-3.4 times lower in the same dose.

Background: A supramolecular complex of praziquantel (PZQ) with disodium salt of glycyrrhizic acid (Na2GA) was obtained by mechanochemical technology to increase solubility, absorption rate and hence bioavailability of the drug and reduction its therapeutic doses. The aim of our study was evaluation of anthelmintic efficacy of supramolecular complex of PZQ. Methods: Different samples of PZQ with Na2GA were obtained by mechanochemical processing and examined for some physico-chemical properties. The anthelmintic activity of the most perspective samples was studied on the laboratory model of Hymenolepis nana infection of mice and Moniezia expansa infection of sheep by the results of helminthological necropsy of the small intestines (the controlled test). Results: A high efficacy (> 98%) of supramolecular complex of PZQ with Na2GA (1/10) was shown at doses of 3; 2 and 1 mg/kg of body weight at single oral administration against H. nana in mice and M. expansa in sheep. While the basic PZQ had 27.19% and 36.64% efficacy respectively at the dose of 1 mg/kg. The PZQ:Na2GA 1/10 physical mixture (without mechanochemical processing) revealed no anthelmintic efficacy. Conclusion: Joint mechanochemical treatment the PZQ substance and Na2GA led to increased solubility, reduction of particle sizes, amorphization of substance, incorporating it with micelles of glycyrrhizic acid and high anthelmintic efficacy in reduced dose. The supramolecular complex of praziquantel was found to be a perspective anthelminthic with enhanced pharmacological activity that needs further research.

The purpose of the research is to study the immunotoxic properties of the supramolecular complex of fenbendazole (CMF).Materials and methods. The study of the immunotoxic properties of SMF was performed using the agglutination reaction and delayed-type hypersensitivity. In each experimental group, there were 7–8 CBA × C57BL / 6 mice weighing 16–18 g. The CMF was administered once intragastrically at a therapeutic dose of 20 mg/kg for the active substance, in a tenfold dose – 200 mg/kg, the basic drug – fenbendazol asked in a tenfold dose of 20 mg/kg. Control groups of animals received starch paste. Then the mice were immunized intraperitoneally with a 3 % suspension of sheep erythrocytes (test antigen) and distributed into 8 groups. The effect of drugs on antibody production was determined in the hemagglutination reaction, the antibody titer in the serum in the microvariant of the direct hemagglutination reaction with the calculation of the reaction index. The effect of the drug on cellular immunity was established in a delayed-type hypersensitivity reaction (DTH) to sheep erythrocytes.Results and discussion. The introduction of the tested drugs had no inhibitory effect on antibody production in mice of the experimental groups: 6.5±0.22; 6.17±0.47 and 7.75±0.25 (log2) compared with the control – 7.67±0.21 (log2). Reaction indices were respectively 0.85; 0,80 and 1,01. Oral administration of a single CMF in therapeutic (20 mg/kg) and in a tenfold therapeutic dose (200 mg/kg) did not affect the cellular level of immunity. The shifts in the inflammatory response index in animals were 9.05±3.47 and 10.05±2.25% respectively. The introduction of the basic drug stimulated the reaction of HRT, which was manifested by an increase in the inflammation index (21.06±3.32%) compared with the control group (9.27±3.91%).

The purpose of the research is to characterize the methods for testing and assessment of efficiency of anthelmintics against trichinellosis and hymenolepidosis in laboratory model.Materials and methods. The literature and own research results have been analyzed as regards testing and efficiency assessment of anthelmintics against trichinellosis and hymenolepidosis in white mouse as a laboratory model weighting 14–16 g that was infected experimentally with Trichinella spiralis larvae in a dose of 200 larvae per specimen and with Hymenolepis nana eggs in a dose of 200 infective eggs per specimen. The medication efficiency was considered subject to the results of a small intestine autopsy on the 2nd and 4th days after the medications were injected to treat trichinellosis and hymenolepidosis in white mice accordingly.Results and discussion. A detailed description has been given for progressive methods to test medication and new chemical compound effect on nematode activity in white mice infected experimentally with T. spiralis, and on cestode activity in white mice infected experimentally with H. nana in a dose of 200 larvae and 100–200 eggs per specimen accordingly. In order to study effectiveness, the test medications were introduced into animals on the 3rd day in the trichinellosis case and on the 13th day in the hymenolepidosis case. Anthelmintic effectiveness was taken into consideration based on helminthologic autopsy of small intestines in animals on the 2nd and 4th days after the medications were given against T. spiralis и H. nana accordingly. The helminthes found in the experimental and control (uncured) animals were counted by a checkup analysis, namely, in comparison with the control animals.