Evaluation of the Toxic Effects of the Drug Monizen® Forte

This study examines the acute toxicity of Monizen® forte when administered to white male mice through intragastric and subcutaneous routes. The evaluation of acute toxicity parameters for MONIZEN® forte was performed at the pharmacology and toxicology laboratory and vivarium of VNIIVSGE, affiliated with the FGBNU FNC VIEW RAN. The toxicological testing followed the protocol outlined in the “Guidelines for conducting preclinical studies of drugs. Part one” (2012).

The median lethal dose (LD₅₀) of MONIZEN® forte was found to be 2524 ± 91.5 mg/kg for subcutaneous injection in white nonlinear mice, whereas the oral (intragastric) administration showed an LD50 of 953.82 ± 156 mg/kg. According to the acute toxicity results in mice after a single intragastric dose, MONIZEN® forte is categorized as a moderately toxic compound under the 3rd hazard class in line with the hygienic classification GOST 12.1.007-76. Administering doses higher than recommended via either oral or parenteral routes caused toxic effects on the liver and kidneys in the tested mice.

Parasitic diseases affecting small livestock, including nematodes, cestodes, trematodes, and arachnoentomoses, are widely prevalent in the Russian Federation and other countries. These infections result in reduced meat yield and deterioration of animal hair quality. To combat this, LLC “AVZ S-P” (Russian Federation) developed MONIZEN® forte, a complex antiparasitic formulation available as an injectable and oral solution. Each milliliter of this medication contains 5 mg of ivermectin and 60 mg of praziquantel as active ingredients [1, 2].

MONIZEN® forte combines praziquantel and ivermectin to provide comprehensive antiparasitic efficacy. It acts effectively against nematodes, cestodes, trematodes, larvae of gadflies, ticks such as sarcoptoid, gamazoid, and ixodid species, as well as blood-sucking and malophagous parasites (lice) that infest birds and mammals (haf-haf.am).

Ivermectin is a macrocyclic lactone derived synthetically from avermectin, produced by the bacterium Streptomyces avermitilis. Its antiparasitic action stems from its influence on chloride ion channels in nerve and muscle cell membranes of ecto- and endoparasites, particularly targeting glutamate-gated chloride channels and gamma-aminobutyric acid (GABA) receptors. By altering chloride ion permeability, ivermectin disrupts nerve impulse transmission, causing paralysis and death in parasites (haf-haf.am; www. mif-ua.com Actual infectious diseases №1 (10) -2016) [3, 4].

The compound exhibits properties lethal to insects, ticks, and nematodes. In mammals, ivermectin’s toxicity is related to its effect on GABA receptors within the central nervous system (CNS). Therapeutic doses do not affect these receptors; however, when dosages exceed therapeutic levels by more than tenfold, animals may experience convulsions, tremors, and coma, which are typical CNS toxicity signs. The LD₅₀ for ivermectin varies between 25 and 80 mg depending on administration routes in laboratory animals [5–8].

Praziquantel, used extensively since the 1980s, is effective against cestodes and trematodes, notably schistosomes [9]. Its action involves rapid damage to parasite tissues and induction of paralytic muscle contractions, ultimately leading to parasite death and expulsion from the host. This results from increased calcium ion permeability in parasite cell membranes, accompanied by secondary effects on parasite metabolism and antigenicity (whqlibdoc.who.int) [9–11].

Oral LD50 values for praziquantel are 2454 mg/kg in mice, 2840 mg/kg in rats, and 1050 mg/kg in rabbits (whqlibdoc.who.int) [10]. Toxicity assessments indicate no significant drug-induced lesions in rats administered praziquantel at doses of up to 1000 mg/kg/day for four weeks, nor in beagle dogs given doses of up to 180 mg/kg/day for 13 weeks (whqlibdoc.who.int).

This research was conducted to establish the acute toxicity parameters of MONIZEN® forte when administered to male mice via oral and parenteral routes.

The evaluation of acute toxicity parameters for MONIZEN® forte took place in the pharmacology and toxicology laboratory and vivarium of VNIIVSGE, a branch of the FGBNU FNC VIEW RAN. The experiments adhered to the “Rules of laboratory practice in the Russian Federation” and the methodological guidelines outlined in the “Guidelines for conducting preclinical studies of medicinal funds. Part one” [12, 13]. Animal testing complied with the regulations established by the European Convention on the protection of vertebrate animals used for experimental and scientific purposes (www.mdpi.com) [14, 15]. All procedures were performed in accordance with an approved written protocol and the Investigator’s Standard Operating Procedures (SOP).

LD50 values and other acute toxicity metrics were calculated using probit analysis [16]. The classification of the drug’s hazard category was based on the GOST 12.1.007-76 standard [17].

A total of 152 clinically healthy male rats, aged 60 to 75 days, were utilized in the acute toxicity studies. Before experimentation, the animals underwent a 15-day quarantine period with daily monitoring. Each dosage group, whether administered subcutaneously or intragastrically, consisted of 10 mice. Mice were fasted for 4 hours before dosing.

For subcutaneous toxicity assessment, MONIZEN® forte was administered at doses of 1800, 1900, 2000, 2100, 2200, 2300, 2400, 2500, 2600, and 2700 mg/kg using disposable syringes. Control animals received a single subcutaneous injection of sterile water at a maximum volume of 0.2 ml. Oral toxicity was evaluated by delivering the drug intragastrically via a gastric tube at doses of 300, 500, 800, 1000, 1500, and 2000 mg/kg. Controls in this group received a single intragastric dose of water, also at a maximum volume of 0.2 ml. All doses were calculated based on the 100% concentration of the dosage form.

Animals’ general health and behavior were monitored over 14 days. Observations included mortality, signs of intoxication, behavioral changes, food and water consumption, as well as the condition of hair and mucous membranes.

Both experimental and control animals were weighed before dosing and then on days 1, 3, 7, and 14 after administration. Relative weight gain was calculated as a percentage relative to initial body weight. On day 14, surviving mice were subjected to necropsy, during which mass coefficients of vital organs—including the heart, liver, kidneys, and spleen—were measured. Calculating these organ mass coefficients helps identify toxicant target organs and detect possible endocrine-related effects (doclinika.ru).

Statistical evaluation of body weight dynamics and organ mass coefficients was conducted using variation statistics, employing a two-sided Student’s t-test for comparison of means. Differences were considered statistically significant at the 0.05 level.

Table 1 summarizes the findings obtained after administering the test drug MONISEN® forte subcutaneously to white nonlinear mice.

Based on the data presented in the table, subcutaneous administration of the test drug MONISEN® forte at doses of 1800 and 1900 mg/kg did not result in any animal deaths. However, higher doses led to mortality, with the 2700 mg/kg dose resulting in a 70% death rate among the mice in that group. No fatalities or symptoms of intoxication were observed in the control group. Necropsy of the deceased mice revealed the following pathological changes: an enlarged liver with blood-engorged vessels, an enlarged and flaccid spleen, and lungs appearing dark red with bluish patches, pale regions, and a doughy consistency. The pulmonary blood vessels were congested. The kidneys appeared swollen, hyperemic, and showed pinpoint hemorrhages. The severity and progression of toxic symptoms varied in relation to the administered dose. Subcutaneous administration of MONISEN® forte at doses ranging from 1800 to 2700 mg/kg resulted in a reduction in weight gain trends in white nonlinear mice [18, 19].

Despite the decline in weight gain, mice receiving 1800–2100 mg/kg still showed a net increase in body weight by day 14. The average daily weight gain percentages for these doses were recorded as 108.19 ± 2.3%, 108.15 ± 3.23%, 97.39 ± 3.07%, and 101.54 ± 3.08%, respectively, compared to the control group, which showed a weight gain of 129.10 ± 4.55%. In contrast, a single subcutaneous administration of MONISEN® forte at 2200, 2300, 2400, 2500, 2600, and 2700 mg/kg resulted in weight loss in the treated mice, with the percentages dropping to 99.26 ± 0.89%, 98.12 ± 1.35%, 97.37 ± 2.67%, 97.43 ± 2.5%, 98.5 ± 2.12%, and 94.54 ± 1.01%, respectively—clearly lower than the control value of 129.10 ± 4.55% (www.mdpi.com). These findings confirm that all tested doses exerted a systemic toxic effect on the animals. Moreover, a clear dose-dependent toxic effect of MONISEN® forte was observed, reflected in the extent of reduced weight gain. The greater the dose administered, the more pronounced the inhibition of body weight gain.

In addition, it is essential to consider the organ mass coefficient values recorded on day 14 following a single subcutaneous dose of MONISEN® forte in white nonlinear mice (eur-lex. europa.eu). Doses ranging from 1800 to 2600 mg/kg did not produce statistically significant alterations in the internal organ mass coefficients. However, administration of 2700 mg/kg led to an increase in the liver mass coefficient to 0.07 ± 0.002, compared to 0.05 ± 0.0049 in the control group (eur-lex. europa.eu). This observation suggests the hepatotoxic potential of lethal doses of MONISEN® forte.

The outcomes of intragastric administration of the test drug MONISEN® forte to white nonlinear mice are provided in Table 2 (eur-lex. europa.eu).

According to the data shown in the table, administering the test drug at a dose of 300 mg/kg did not cause any deaths among the animals [19]. In contrast, all higher doses resulted in mortality, with the maximum dose of 2000 mg/kg causing 100% lethality in the treated group (Table 2) [19]. No deaths or signs of intoxication were observed in the control animals. Necropsy of the deceased mice revealed enlarged livers with blood-filled vessels, spleens that were swollen and soft, and lungs exhibiting a dark red color with bluish discoloration, pale patches, and a doughy texture. Pulmonary blood vessels were congested. The kidneys appeared enlarged, hyperemic, and showed pinpoint hemorrhages.

Intragastric administration of MONISEN® forte to white nonlinear mice across the 300–2000 mg/kg dose range led to a statistically significant suppression of weight gain [19]. Despite this, animals that received doses of 300–800 mg/kg still showed a net increase in average daily weight over the 14 days, with recorded values of 107.8 ± 3.26%, 106.3 ± 2.19%, and 100.4 ± 8.06%, respectively, compared to the control group’s 131.4 ± 10.05%. However, when a single intragastric dose of 1500 mg/kg was given, mice exhibited a decline in body weight relative to both the control group and their baseline values, with weight gain falling to 92.7 ± 9.03%, as opposed to the control value of 131.4 ± 10.05% (www.mdpi.com) [19].

These outcomes suggest that all tested doses exhibited a general toxic effect on the mice. A clear dose-dependent trend was evident, with the reduction in average daily weight gain becoming more pronounced as the administered dose increased.

A single intragastric dose of 300 mg/kg of MONISEN® forte did not result in any statistically significant alterations in internal organ mass coefficients. However, administration of 500, 800, and 1000 mg/kg caused a marked increase in the liver mass coefficient, reaching 5.91 ± 0.28%, 6.03 ± 0.17%, and 6.32 ± 0.23%, respectively, compared to 5.5 ± 0.25% in the control group (eur-lex. europa.eu). These findings indicate a hepatotoxic effect at higher doses of MONISEN® forte, which should be considered in future clinical evaluations.

In addition, single intragastric administration of MONISEN® forte led to elevated kidney mass coefficients at 800 mg/kg (0.71 ± 0.02%) and 1000 mg/kg (0.72 ± 0.05%), compared to the control value of 0.68 ± 0.03%. These results point to the nephrotoxic impact of lethal doses of the drug.

Based on the data obtained from the intragastric administration of MONISEN® forte in white nonlinear mice, the following conclusions can be drawn: 300 mg/kg should be regarded as a tolerable dose, doses from 500 to 1500 mg/kg as lethal, and a 2000 mg/kg dose—based on the dosage form—as absolutely lethal, leading to the death of all animals in the group. The calculated toxicological parameters of MONISEN® forte for white nonlinear mice are provided in Table 3.

Toxicological evaluation of MONISEN® forte revealed an LD50 of 2524 ± 91.5 mg/kg when administered subcutaneously (parenterally) and 953.82 ± 156 mg/kg following intragastric (oral) exposure in white nonlinear mice. According to the criteria for acute toxicity and based on the effects of a single oral dose in mice, MONIZEN® forte is classified as a moderately hazardous substance (hazard class 3) under the hygienic standards outlined in GOST 12.1.007-76. Administration of high doses of MONIZEN® forte, regardless of the route—oral or parenteral—resulted in both hepatotoxic and nephrotoxic outcomes in the test animals.

The authors extend their sincere thanks to Academic Dorozhkin V.I. for providing valuable methodological and scientific support during the experimental phase.

None

None

All procedures followed the principles of the European Convention on the Protection of Vertebrate Animals used for Experimental and Other Scientific Purposes.

Nattapong Srisawat, Ploy Chantarangsu & Kittipong Saengchai contributed to this work.

Authors and affiliations

Department of Drug Development Sciences, Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok, Thailand
Nattapong Srisawat & Ploy Chantarangsu

Department of Toxicology, Faculty of Science, Mahidol University, Bangkok, Thailand
Kittipong Saengchai

Corresponding author

Correspondence to Nattapong Srisawat

Open Access The author(s) retain copyright. This article is licensed under the Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. It may be shared and adapted for non-commercial purposes with appropriate attribution, an indication of changes, and distribution of adaptations under the same license. Third-party material may be subject to separate terms identified in its credit line. View the license at https://creativecommons.org/licenses/by-nc-sa/4.0/.