Доклиническое тестирование нового отечественного супрамолекулярного комплекса триклабендазола «триклафасцид» на эмбриотропную активность

The purpose of the study: embryotoxic and teratogenic effects of a new domestic supramolecular complex of triclabendazole “Triclafascid”. Materials and methods. Embryotoxic and teratogenic effects of new domestic formulations studied Triclafascid accordance with the “Manual on experimental (preclinical) study of new pharmacological substances”. The study embryotrophic actions supramolecular complex preparation on the basis of the substance of triclabendazole was performed on 40 white mongrel female rats weighing 220-260g and 10 males, in accordance with the guidelines on the assessment of the impact of drugs on generative function of animals. To rats-females were placed overnight male ratio of 1:4. Detection of sperm in the vaginal smear, the females, on the morning after the infusion of the male is pointed at fertilization -the first day of pregnancy. Since the sensitivity of the embryo to chemical and depends on the various stages of fetal development, the animals were divided into 4 groups of 10 animals each. Triclafascid was administered orally to pregnant females three times increased therapeutic dose of 6.0 mg/kg (60 mg/kg of the drug), the first group 1 on day 6 of pregnancy, the second from 7 - 14, third 15 and 19, the fourth group served as control and received 1% starch gel. On the 20th pregnancy day, the rats were euthanized with carbon dioxide. After slaughter and opening of the abdominal cavity have been removed the uterus with the fruit. Counted the number of yellow bodies of pregnancy, places of implantation, resorption, live and dead embryos. To assess the embryotoxic effect of the fruits were viewed under binocular magnifying glass to detect external anomalies, weighed, measured the cranio-caudal size, weight and diameter of placenta. Was determined pre - and postimplantation loss and total embryonic mortality of embryos. After inspection, the embryos from each rat was divided into two equal groups: the first were fixed in solution of Bouin for 14 days to study the internal organs of fetuses, and anomalies in developing fetuses, which are indications of teratogenic effect is determined by the method of J. G. Wilson (1965) in modification of the Department of embryology held the Academy of medical Sciences of the USSR (the scheme of transects made through the fetus); the second was fixed in 96 alcohol for study of the bone system after its dyeing by the method of Dawson (A. B. Dawson, 1926). The parameters obtained were processed by variation statistics with the help of simple comparisons of average according to the bilateral student’s t-test. The difference was determined at 0.05 level of significance. The calculations were performed using the “Student-200”. The results and discussion. As shown by the results of studies Triclafascid showed no embryotoxic activity when exposed to 3-fold increased therapeutic dose of 6.0 mg/kg po DV. So, overall, pre-and postimplantation mortality of fetuses in the experimental and control groups did not differ significantly, as with the introduction of the drug for 1-6 days of pregnancy and 7 -14 and 15 - 19 days. Based on these data it can be concluded that the drug Triclafascid has no negative influence on embryonic development. Values pre - and postimplantation and total embryonic mortality of experimental animals in comparison with control values, we can say that the drug did not induce the death of embryos in different periods of embryogenesis. The mass and size of the fruit also did not differ from the control, which indicates the absence of embryotoxic effect. A careful visual inspection of fruits in all experimental groups was not detected for any external malformations compared with controls. By the execution of nine sagittal sections of internal abnormalities, malformations of the internal organs, disorders of the topography was found. A teratogenic effect characterized by different anomalies of the internal organs of fetuses (Wilson’s method) and external defects were also not observed. When studying the skeletal system: the sizes of the rudiments of the shoulder; brachial; ulnar; radial; femoral; large and small tibial bones from experimental and control embryos were similar in metrics (length, mm). The condition of the bone system was unchanged (P>0,05). Therefore, Triclafascid showed no teratogenic activity when exposed at critical periods of embryogenesis of rats.Цель исследования: эмбриотоксическое и тератогенное действие нового отечественного супрамолекулярного комплекса триклабендазола «триклафасцид». Материалы и методы. Эмбриотоксическое и тератогенное действие новой отечественной лекарственной формы триклафасцида изучали соответствии с «Руководством по экспериментальному (доклиническому) изучению новых фармакологических веществ». Изучение эмбриотропного действия супрамолекулярного комплексного препарата на основе субстанции триклабендазола проводили на 40 белых беспородных крысах - самках массой 220-260г и 10 самцах, в соответствии с Методическими рекомендациями по оценке влияния препарата на генеративную функцию животных. К крысам-самкам подсаживали на ночь самцов из соотношения 1:4. Обнаружение спермиев во влагалищном мазке самки, на утро, после подсадки самца указывало на оплодотворение -первый день беременности. Так как чувствительность эмбриона к химическому веществу различна и зависит от стадий развития плода, животных разделили на 4 группы, по 10 особей в каждой. Триклафасцид вводили беременным самкам перорально в трехкратно увеличенной терапевтической дозе 6,0 мг/кг (60 мг/кг по препарату), первой группе с 1 по 6 день беременности, второй с 7 - 14, третьей с 15 - 19, четвертая группа служила контролем и получала 1% крахмальный гель. На 20-й день беременности крыс усыпляли углекислым газом. После убоя и вскрытия брюшной полости извлекали матку с плодами. Подсчитывали количество желтых тел беременности, мест имплантации, резорбции, живых и мертвых эмбрионов. Для оценки эмбриотоксического эффекта плоды просматривали под бинокулярной лупой для обнаружения внешних аномалий развития, взвешивали, измеряли кранио-каудальный размер, массу и диаметр плаценты. Определяли пред- и постимплантационную гибель и общую эмбриональную смертность эмбрионов. После осмотра эмбрионы от каждой крысы делили на две равные группы: первую фиксировали в растворе Буэна на 14 суток для изучения внутренних органов плодов и аномалий в развитии плодов, которые являются показателями тератогенного эффекта определяемые по методу J.G. Wilson (1965) в модификации отдела эмбриологии НИИЭМ АМН СССР (схема разрезов, сделанных через плод); вторую-фиксировали в 96° спирте для изучения костной системы после ее окрашивания по методу Даусона (A.B. Dawson, 1926). Полученные параметры обрабатывали методом вариационной статистики с помощью простого сравнения средних по двустороннему t-критерию Стьюдента. Различие определяли при 0,05 уровне значимости. Расчет выполнен с помощью программы «Student-200». Результаты и обсуждение. Как показали результаты исследований, триклафасцид не проявил эмбриотоксической активности при воздействии в 3-кратно увеличенной терапевтической дозе 6,0 мг/кг по ДВ. Так, уровень общей, пред-и постимплантационной смертности плодов в подопытных и контрольной группах достоверно не отличались, как при введении препарата на 1-6 дни беременности, так и на 7 - 14 и 15 - 19 дни. На основании полученных данных можно сделать вывод о том, что препарат Триклафасцид не оказывает отрицательного влияния на эмбриональное развитие плода. По значениям пред- и постимплатационной и общей эмбриональной смертности опытных животных, в сравнении с контрольными значениями, можно сказать, что препарат не индуцировал гибель эмбрионов в различные периоды эмбриогенеза. Масса и размеры плодов также не отличались от контроля, что говорит об отсутствии эмбриотоксического эффекта. При тщательном визуальном осмотре плодов во всех опытных группах не было обнаружено каких-либо внешних аномалий развития, в сравнении с контролем. По результатам выполнения девяти сагиттальных разрезов внутренних уродств, аномалий развития внутренних органов, нарушения топографии не было обнаружено. Тератогенный эффект, характеризующийся различными аномалиями со стороны внутренних органов плодов (метод Вильсона) и внешними дефектами развития, также не был отмечен. При изучении костной системы: размеры зачатков лопаточной; плечевой; локтевой; лучевой; бедренной; большой и малой берцовых костей у опытных и контрольных эмбрионов были близки по показателям (длина, мм). Состояние костной системы было без изменений (P>0,05). Следовательно, триклафасцид не проявил тератогенной активности при воздействии в “критические” периоды эмбриогенеза крыс

ИЗУЧЕНИЕ ИММУНОТРОПНОЙ АКТИВНОСТИ СУПРАМОЛЕКУЛЯРНОГО КОМПЛЕКСА ТРИКЛАБЕНДАЗОЛА

The supramolecular complex of triclabendazole is a complex preparation based on triclabendazole with the water-soluble polysaccharide - arabinogalactan produced in impact grinders with the use of mechanochemical nanotechnology. Objective of research: To provide a preclinical assessment of immunotoxic properties of the supramolecular complex of triclabendazole used on laboratory animals. Materials and methods: Two experiments were conducted on 60 male mice with the mass 18-20 g. to determine effects of the supramolecular complex of triclabendazole on humoral and cell-mediated immune responses. 20 mice received intragastric injection of preparation once at a therapeutic dose 30 mg/kg in 1% of starch gel; 20 mice at a tenfold dose - 300 mg/kg, and 20 mice served as controls and did not receive the preparation. Then, all animals (60 ind.) were immunized once intraperitoneally with 0,5 ml of 3% suspension of sheep erythrocytes (antigen test) and divided into 6 groups (10 ind. in each). The effect of the drug on antibody formation was estimated by agglutination test in 30 mice. The antibody titre in blood serum was determined on the seventh day after immunization by a direct microhemagglutination assay. To compare the immune response in experimental and control groups, the index of drug effects was established. Effects of the drug on cell immunity were determined by the delayed-type hypersensitivity reaction. Experiment was carried out on 30 mice divided into three groups (10 ind. in each). Research was conducted according to the «Manual on experimental (preclinical) study of new pharmacological substances (2005)». Results and discussion: Antibody titres in blood serum of control animals were 7,11±0,31 (log2). Peroral single administration of the tested drug at a therapeutic and tenfold dose did not cause any changes in agglutinin titres in blood serum of animals. Index of drug effects in the 1st and 2nd group was 1,04 and 1,12 respectively, which confirms the absence of negative effects of the humoral immune response. Peroral single administration of the drug at a therapeutic dose 30 mg/kg and at a tenfold dose - 300 mg/kg inhibits the delayed-type hypersensitivity reaction in comparison to controls. Inflammatory factors in animals from 1st and 2nd groups were 6,12±0,87 and 6,64±1,37 %, respectively; in control group - 8,11±0,93 %, but this difference was not statistically significant (Р ≥ 0,05).Супрамолекулярный комплекс триклабендазола - это комплексный препарат на основе триклабендазола с водорастворимым полисахаридом арабиногалактаном, полученный с применением механохимической нанотехнологии в измельчителях ударно-истирающего типа. Цель исследования - доклиническая оценка иммунотоксических свойств супрамолекулярного комплекса триклабендазола (СМКТ) на лабораторных животных. Материалы и методы. Проведено два опыта на 60 мышах-самцах массой 18-20 г с целью определения влияния СМКТ на гуморальный и клеточный иммунный ответ. 20 мышам препарат вводили однократно внутрижелудочно в терапевтической дозе 30 мг/кг в 1%-ном крахмальном геле, 20 - в десятикратно увеличенной дозе - 300 мг/кг и 20 мышей служили контролем и препарат не получали. Затем всех животных (60 гол.) иммунизировали интраперитонеально в объеме 0,5 мл 3%-ной взвеси эритроцитов барана (тест-антиген) и распределили на 6 групп по 10 голов в каждой. Влияние препарата на антителообразование определяли в реакции агглютинации на 30 мышах. Титр антител в сыворотке крови определяли на 7-е сутки после иммунизации в микроварианте прямой реакции гемагглютинации. Для сравнения выраженности иммунного ответа в опыте и контроле определяли индекс действия препарата (ИД). Влияние препарата на клеточный иммунитет устанавливали в реакции гиперчувствительности замедленного типа (ГЗТ). Опыт проводили на 30 мышах, которых разделили на три группы по 10 голов в каждой. Исследования проводили согласно Руководству по экспериментальному (доклиническому) изучению новых фармакологических веществ (2005). Результаты и обсуждение. При исследовании сывороток крови контрольных животных титры антител зарегистрированы на уровне 7,11±0,31 (log2). Пероральное однократное введение испытуемого препарата в терапевтической и десятикратно увеличенной дозах не привело к изменению титров агглютининов в сыворотке крови животных. ИД для 1 и 2-й групп составил соответственно 1,04 и 1,12, что оценивается как отсутствие отрицательного влияния на гуморальный иммунный ответ. Пероральное однократное введение препарата в терапевтической (30 мг/кг) и десятикратно увеличенной терапевтической дозе (300 мг/кг) тормозило индукцию ГЗТ по сравнению с контрольным значением. Сдвиг индекса воспаления у животных 1 и 2-й опытных групп составил соответственно 6,12±0,87 и 6,64±1,37 %, в контрольной группе - 8,11±0,93 %, но эта разница была статистически недостоверна (Р ≥ 0,05)

Субхроническая токсичность супрамолекулярного комплекса триклабендазола

Objective of research: Preclinical assessment of subchronic toxicity of the supramolecular complex of Triclabendazole applied on laboratory animals. Materials and methods: Investigations were conducted on 40 male rats with the body mass of 200-220 g. Animals were divided into 4 equal groups. The drug was given to rats of 1, 2 and 3 groups at the doses of 1/5 (2600 mg/kg), 1/10 (1300 mg/ kg) and 1/20 (650 mg/kg) of LD50 (13000 mg/kg), respectively, during 7 days daily orally into the stomach using the gastric tube. Animas from the 4th group received starch paste 1% and served as controls. During the experiment, we observed the general condition of animals, visible physiological functions (food and water intake, etc.), possible signs of intoxication; animals were weighted on the 1st, 3rd, 5th and 7th day of the experiment. On the 8th day of the experiment, animals were killed by decapitation. After killing rats and blood taking, laparotomy was conducted, mass of the main organs (heart, lungs, liver, spleen, brain, seminal glands, thymus, pancreas, and adrenal glands) was determined, their mass coefficients calculated, visible changes detected. Hematological and biochemical indices of rats from experimental and control groups were investigated using the automatic analyzer. Results and discussion: When using the drug in three test doses, general condition and behavior of animals were normal; no signs of intoxication were detected. Triclafascid did not induce an increase in body mass. The investigation of internal organs of experimental animals did not reveal abnormalities. Mass coefficients of internal organs of rats from experimental and control groups did not significantly differ from each other. The application of the drug at the doses of 1/5 and 1/10 of LD50 caused minor decrease in the hemoglobin level related to the controls. The number of erythrocytes, thrombocytes, leucocytes, erythrocyte sedimentation rate (ESR) showed no significant changes. In tested doses, Triclafascid had no significant effect on concentrations of total protein and glucose. Kidney function was estimated by urea and creatinine levels. Both values were equal to the controls. Activities of aspartate and alanine aminotransferase did not show any significant changes after application of the drug in the tested doses, which indicated the normal liver function.Цель исследования - доклиническая оценка субхронической токсичности супрамолекулярного комплекса триклабендазола на лабораторных животных. Материалы и методы. Исследования проводили на 40 крысах-самцах массой тела 200-220 г Животных разделили на 4 равные группы. Препарат вводили крысам 1, 2 и 3-й групп в дозах соответственно 1/5 (2600 мг/кг), 1/10 (1300 мг/кг) и 1/20 (650 мг/кг) от ЛД50 (13000 мг/кг) в течение 7 сут ежедневно перорально в желудок с помощью зонда. Животные 4-й группы получали 1%-ный крахмальный клейстер и служили контролем. В течение опыта проводили наблюдение за общим состоянием животных, видимыми физиологическими функциями (приемом корма, воды и т.д.), наличием признаков интоксикации; взвешивали животных на 1, 3, 5 и 7-е сутки опыта. На 8-е сутки животных всех групп убивали методом декапитации. После убоя крыс и забора крови проводили лапаротомию, определяли массу основных органов (сердца, легких, печени, почек, селезенки, мозга, семенников, тимуса, поджелудочной железы и надпочечников) и рассчитывали массовые коэффициенты, устанавливали наличие видимых изменений. Гематологические и биохимические показатели крови у крыс подопытных и контрольной групп исследовали на автоматическом анализаторе. Результаты и обсуждение. При введении препарата в трех тестируемых дозах общее состояние и поведение животных было в норме; признаки интоксикации отсутствовали. Триклафасцид не влиял на прирост массы тела. При исследовании внутренних органов опытных животных отклонений от нормы выявлено не было. Массовые коэффициенты внутренних органов у крыс опытных и контрольной группы достоверно не отличались друг от друга. При введении препарата в дозах 1/5 и 1/10 от ЛД50 наблюдали незначительное понижение уровня гемоглобина по отношению к контролю. Количество эритроцитов, тромбоцитов, лейкоцитов, СОЭ достоверных изменений не претерпели. В испытанных дозах триклафасцид не оказал значимого влияния на концентрацию общего белка, глюкозы. Функциональное состояние почек оценивали по уровню мочевины и креатинина. Оба показателя были на уровне с контролем. Активность аспартат- и аланинаминотрансфераз не подверглась значительным изменениям после введения препарата в тестируемых дозах, что говорит о том, что функциональное состояние печени остается в норме

Комиссионное и производственное испытание эффективности супрамолекулярного комплекса триклабендазола триклафасцид при фасциолёзе крупного рогатого скота

The purpose of the research: to conduct commission and field testing of efficacy of the supramolecular complex of triclabendazole «Triclafascid» applied at the dose of 2,5-3,0 mg/kg against cattle fasciolosis. Materials and methods. Commission and field testing of Triclafascid were carried out in the private sector of the Chechen Republic in March-April 2017 on cattle naturally infected with Fasciola. Commission testing was performed on 30 animals infected with Fasciola that were divided into two equal groups of 15 animals each. Animals from the first group received triclafascid orally at the single therapeutic dose 2,5 mg a.i./kg (therapeutic dose 25 mg/kg) in water solution. The second group of animals as well as the first group received the substance of triclabendazole at five times lower dose (2,5 mg/kg) in water suspension. In field experiment, triclafascid was given individually to 108 animals at the dose of 3,0 mg a.i./kg (therapeutic dose 30 mg/kg) with compound feed. To determine the grade of animals’ invasion with fascioles, fecal samples were examined by Fuelleborn's method using ammonium nitrate. The average number of Fasciola eggs in 1 g of feces was defined with the use of VIGIS counting chamber. Fecal samples were investigated; the efficacy of the preparation was evaluated 30 days after dehelmintization. The efficacy of triclafascid was estimated by a «critical test» according to the Manual approved by World Association for the Advancement of Veterinary Parasitology (1995). Results and discussion. In commission testing of triclafascid for the treatment of cattle fasciolosis individually orally in water solution at the dose of 2,5 mg /kg and in field experiment at the dose of 3,0 mg /kg with feed compound at five times lower dose, 100 % efficacy was reached (in comparison with triclabendazole).Цель исследований: комиссионное и производственное испытание эффективности отечественного антигельминтика трикла фасцида в дозе 2,5-3,0 мг/кг при фасциолёзе крупного рогатого скота. Материалы и методы. Комиссионное и производственное испытание триклафасцида проводили в частном секторе Чеченской республики в марте-апрель 2017 г. на крупном рогатом скоте, спонтанно инвазированном фасциолами. Комиссионный опыт проводили на 30 спонтанно инвазированных фасциолами животных, которых распределили по принципу аналогов на две равноценные группы по 15 голов в каждой. Животные первой группы получали триклафасцид перорально в терапевтической дозе 2,5 мг/кг по ДВ (по препарату 25 мг/кг) в форме водного раствора однократно. Второй группе животных вводили субстанцию триклабендазола в форме водной взвеси в 5 раз уменьшенной дозе (2,5 мг/кг) в качестве контроля аналогично как и животным первой группы. В производственном опыте триклафасцид задавали 108 животным в дозе 3,0 мг/кг по ДВ (по препарату 30 мг/кг) индивидуально в смеси с комбикормом. Для определения степени инвазированности животных фасциолами пробы фекалий исследовали методом Фюллеборна с использованием аммиачной селитры. Среднее число яиц фасциол в 1 г фекалий определяли с помощью камеры ВИГИС. Эффективность препарата оценивали через 30 сут после дегельминтизации путем исследования проб фекалий. Учёт эффективности триклафасцида проводили по типу «критический тест» согласно Руководству, одобренному Всемирной Ассоциацией за прогресс ветеринарной паразитологии (1995 г). Результаты и обсуждение. При комиссионном испытании триклафасцида при фасциолёзе крупного рогатого скота в дозе 2,5 мг/ кг индивидуально в форме водного раствора перорально и в производственном опыте в дозе 3,0 мг/кг в смеси с кормом в 5 раз уменьшенной дозе по сравнению с субстанцией триклабендазола, получена 100%-ная эффективность

Эффективность супрамолекулярных комплексов триклабендазола с полимерными наполнителями при фасциолёзе

With the usuing of mechanochemical nanotechnology in the mills of shock-attrition type with adjustable energy densities, developed 10 innovative drugs in the form of supramolecular complexes on the basis triclabendazole with various water-soluble polymers without participation of the liquid phase in one stage. They are fine dry water-soluble powders with a particle size of 1-10 microns. All tested supramolecular complexes of triclabendazole against sexually Mature at 5, and immature Fasciala 10 times more effective than the drug substance of Tticlabendazole. For technical and economic reasons the most optimal product for introduction into veterinary practice is the complex Tticlabendazole, with a water-soluble polysaccharide arabinogalactan, extracted from larch, environmentally and safe product that is widely used in medicine. It is known that water-insoluble anthelmintic, applied to animals orally, to 70.0% excreted into the environment unchanged in the faeces, contaminating the environment. Water-soluble supramolecular complexe Tticlabendazole used in a reduced dose, soaked in blood, provide a high biological availability. In the future, they are broken down and eliminated in the urine from the body in miniscule amounts, without polluting the environment. These positive signs supramolecular complex compared with the substance triclabendazole anthelmintic and provide high economic efficiency and safety.С применением механохимической нанотехнологии в измельчителях ударно-истирающего типа с регулируемой энергонапряжённостью, наработаны 10 инновационных препаратов в виде супрамолекулярных комплексов на основе субстанции триклабендазола с различными водорастворимыми полимерами. Синтез проведен без участия жидких фаз в одну стадию. Комплексы представляют собой тонкодисперсные, легко сыпучие и растворимые в воде порошки с размером частиц до 1-10 микрон. Все испытанные супрамолекулярные комплексы триклабендазола против половозрелых в 5, а неполовозрелых фасциол 10 раз эффективнее, чем субстанция базового препарата триклабендазола. По техническим и экономическим причинам наиболее оптимальным препаратом для внедрения в ветеринарную практику выбран комплекс триклабендазола (Триклафасцид) с водорастворимым полисахаридом арабиногалактаном, выделяемым из лиственницы, экологически чистым и безопасным продуктом, который широко применяется в медицине и ветеринарии. Известно, что нерастворимые в воде антигельминтики, применяемые животным перорально, до 70,0% выводятся во внешнюю среду в неизменном виде с фекалиями, загрязняя окружающую среду. Растворимые в воде супрамолекулярные комплексы, применяемые в сниженной дозе, всасываясь в кровь, обеспечивают высокую биологическую доступность. В дальнейшем, они расщепляются и выводятся в основном с мочой из организма в мизерных количествах, не загрязняя окружающую среду Перечисленные положительные признаки супрамолекулярного комплекса по сравнению с субстанцией триклабендазола обеспечивают высокую антигельминтную и экономическую эффективность и безопасность применения

Preclinical testing of new domestic supramolecular complex of triclabendazole «Triclafascid» embryotrophic activity

The purpose of the study: embryotoxic and teratogenic effects of a new domestic supramolecular complex of triclabendazole “Triclafascid”. Materials and methods. Embryotoxic and teratogenic effects of new domestic formulations studied Triclafascid accordance with the “Manual on experimental (preclinical) study of new pharmacological substances”. The study embryotrophic actions supramolecular complex preparation on the basis of the substance of triclabendazole was performed on 40 white mongrel female rats weighing 220-260g and 10 males, in accordance with the guidelines on the assessment of the impact of drugs on generative function of animals. To rats-females were placed overnight male ratio of 1:4. Detection of sperm in the vaginal smear, the females, on the morning after the infusion of the male is pointed at fertilization -the first day of pregnancy. Since the sensitivity of the embryo to chemical and depends on the various stages of fetal development, the animals were divided into 4 groups of 10 animals each. Triclafascid was administered orally to pregnant females three times increased therapeutic dose of 6.0 mg/kg (60 mg/kg of the drug), the first group 1 on day 6 of pregnancy, the second from 7 - 14, third 15 and 19, the fourth group served as control and received 1% starch gel. On the 20th pregnancy day, the rats were euthanized with carbon dioxide. After slaughter and opening of the abdominal cavity have been removed the uterus with the fruit. Counted the number of yellow bodies of pregnancy, places of implantation, resorption, live and dead embryos. To assess the embryotoxic effect of the fruits were viewed under binocular magnifying glass to detect external anomalies, weighed, measured the cranio-caudal size, weight and diameter of placenta. Was determined pre - and postimplantation loss and total embryonic mortality of embryos. After inspection, the embryos from each rat was divided into two equal groups: the first were fixed in solution of Bouin for 14 days to study the internal organs of fetuses, and anomalies in developing fetuses, which are indications of teratogenic effect is determined by the method of J. G. Wilson (1965) in modification of the Department of embryology held the Academy of medical Sciences of the USSR (the scheme of transects made through the fetus); the second was fixed in 96 alcohol for study of the bone system after its dyeing by the method of Dawson (A. B. Dawson, 1926). The parameters obtained were processed by variation statistics with the help of simple comparisons of average according to the bilateral student’s t-test. The difference was determined at 0.05 level of significance. The calculations were performed using the “Student-200”. The results and discussion. As shown by the results of studies Triclafascid showed no embryotoxic activity when exposed to 3-fold increased therapeutic dose of 6.0 mg/kg po DV. So, overall, pre-and postimplantation mortality of fetuses in the experimental and control groups did not differ significantly, as with the introduction of the drug for 1-6 days of pregnancy and 7 -14 and 15 - 19 days. Based on these data it can be concluded that the drug Triclafascid has no negative influence on embryonic development. Values pre - and postimplantation and total embryonic mortality of experimental animals in comparison with control values, we can say that the drug did not induce the death of embryos in different periods of embryogenesis. The mass and size of the fruit also did not differ from the control, which indicates the absence of embryotoxic effect. A careful visual inspection of fruits in all experimental groups was not detected for any external malformations compared with controls. By the execution of nine sagittal sections of internal abnormalities, malformations of the internal organs, disorders of the topography was found. A teratogenic effect characterized by different anomalies of the internal organs of fetuses (Wilson’s method) and external defects were also not observed. When studying the skeletal system: the sizes of the rudiments of the shoulder; brachial; ulnar; radial; femoral; large and small tibial bones from experimental and control embryos were similar in metrics (length, mm). The condition of the bone system was unchanged (P>0,05). Therefore, Triclafascid showed no teratogenic activity when exposed at critical periods of embryogenesis of rats

STUDIES ON IMMUNOTROPIC ACTIVITY OF THE SUPRAMOLECULAR COMPLEX OF TRICLABENDAZOLE

The supramolecular complex of triclabendazole is a complex preparation based on triclabendazole with the water-soluble polysaccharide - arabinogalactan produced in impact grinders with the use of mechanochemical nanotechnology. Objective of research: To provide a preclinical assessment of immunotoxic properties of the supramolecular complex of triclabendazole used on laboratory animals. Materials and methods: Two experiments were conducted on 60 male mice with the mass 18-20 g. to determine effects of the supramolecular complex of triclabendazole on humoral and cell-mediated immune responses. 20 mice received intragastric injection of preparation once at a therapeutic dose 30 mg/kg in 1% of starch gel; 20 mice at a tenfold dose - 300 mg/kg, and 20 mice served as controls and did not receive the preparation. Then, all animals (60 ind.) were immunized once intraperitoneally with 0,5 ml of 3% suspension of sheep erythrocytes (antigen test) and divided into 6 groups (10 ind. in each). The effect of the drug on antibody formation was estimated by agglutination test in 30 mice. The antibody titre in blood serum was determined on the seventh day after immunization by a direct microhemagglutination assay. To compare the immune response in experimental and control groups, the index of drug effects was established. Effects of the drug on cell immunity were determined by the delayed-type hypersensitivity reaction. Experiment was carried out on 30 mice divided into three groups (10 ind. in each). Research was conducted according to the «Manual on experimental (preclinical) study of new pharmacological substances (2005)». Results and discussion: Antibody titres in blood serum of control animals were 7,11±0,31 (log2). Peroral single administration of the tested drug at a therapeutic and tenfold dose did not cause any changes in agglutinin titres in blood serum of animals. Index of drug effects in the 1st and 2nd group was 1,04 and 1,12 respectively, which confirms the absence of negative effects of the humoral immune response. Peroral single administration of the drug at a therapeutic dose 30 mg/kg and at a tenfold dose - 300 mg/kg inhibits the delayed-type hypersensitivity reaction in comparison to controls. Inflammatory factors in animals from 1st and 2nd groups were 6,12±0,87 and 6,64±1,37 %, respectively; in control group - 8,11±0,93 %, but this difference was not statistically significant (Р ≥ 0,05)

Subchronic toxicity of the supramolecular complex of triclabendazole

Objective of research: Preclinical assessment of subchronic toxicity of the supramolecular complex of Triclabendazole applied on laboratory animals. Materials and methods: Investigations were conducted on 40 male rats with the body mass of 200-220 g. Animals were divided into 4 equal groups. The drug was given to rats of 1, 2 and 3 groups at the doses of 1/5 (2600 mg/kg), 1/10 (1300 mg/ kg) and 1/20 (650 mg/kg) of LD50 (13000 mg/kg), respectively, during 7 days daily orally into the stomach using the gastric tube. Animas from the 4th group received starch paste 1% and served as controls. During the experiment, we observed the general condition of animals, visible physiological functions (food and water intake, etc.), possible signs of intoxication; animals were weighted on the 1st, 3rd, 5th and 7th day of the experiment. On the 8th day of the experiment, animals were killed by decapitation. After killing rats and blood taking, laparotomy was conducted, mass of the main organs (heart, lungs, liver, spleen, brain, seminal glands, thymus, pancreas, and adrenal glands) was determined, their mass coefficients calculated, visible changes detected. Hematological and biochemical indices of rats from experimental and control groups were investigated using the automatic analyzer. Results and discussion: When using the drug in three test doses, general condition and behavior of animals were normal; no signs of intoxication were detected. Triclafascid did not induce an increase in body mass. The investigation of internal organs of experimental animals did not reveal abnormalities. Mass coefficients of internal organs of rats from experimental and control groups did not significantly differ from each other. The application of the drug at the doses of 1/5 and 1/10 of LD50 caused minor decrease in the hemoglobin level related to the controls. The number of erythrocytes, thrombocytes, leucocytes, erythrocyte sedimentation rate (ESR) showed no significant changes. In tested doses, Triclafascid had no significant effect on concentrations of total protein and glucose. Kidney function was estimated by urea and creatinine levels. Both values were equal to the controls. Activities of aspartate and alanine aminotransferase did not show any significant changes after application of the drug in the tested doses, which indicated the normal liver function

Commission and field testing of efficacy of the supramolecular complex of triclabendazole «Triclafascid» against cattle fascioliasis

The purpose of the research: to conduct commission and field testing of efficacy of the supramolecular complex of triclabendazole «Triclafascid» applied at the dose of 2,5-3,0 mg/kg against cattle fasciolosis. Materials and methods. Commission and field testing of Triclafascid were carried out in the private sector of the Chechen Republic in March-April 2017 on cattle naturally infected with Fasciola. Commission testing was performed on 30 animals infected with Fasciola that were divided into two equal groups of 15 animals each. Animals from the first group received triclafascid orally at the single therapeutic dose 2,5 mg a.i./kg (therapeutic dose 25 mg/kg) in water solution. The second group of animals as well as the first group received the substance of triclabendazole at five times lower dose (2,5 mg/kg) in water suspension. In field experiment, triclafascid was given individually to 108 animals at the dose of 3,0 mg a.i./kg (therapeutic dose 30 mg/kg) with compound feed. To determine the grade of animals’ invasion with fascioles, fecal samples were examined by Fuelleborn's method using ammonium nitrate. The average number of Fasciola eggs in 1 g of feces was defined with the use of VIGIS counting chamber. Fecal samples were investigated; the efficacy of the preparation was evaluated 30 days after dehelmintization. The efficacy of triclafascid was estimated by a «critical test» according to the Manual approved by World Association for the Advancement of Veterinary Parasitology (1995). Results and discussion. In commission testing of triclafascid for the treatment of cattle fasciolosis individually orally in water solution at the dose of 2,5 mg /kg and in field experiment at the dose of 3,0 mg /kg with feed compound at five times lower dose, 100 % efficacy was reached (in comparison with triclabendazole)

The effectiveness of supramolecular complxes of triclabendazole with polymers against Fasciola

With the usuing of mechanochemical nanotechnology in the mills of shock-attrition type with adjustable energy densities, developed 10 innovative drugs in the form of supramolecular complexes on the basis triclabendazole with various water-soluble polymers without participation of the liquid phase in one stage. They are fine dry water-soluble powders with a particle size of 1-10 microns. All tested supramolecular complexes of triclabendazole against sexually Mature at 5, and immature Fasciala 10 times more effective than the drug substance of Tticlabendazole. For technical and economic reasons the most optimal product for introduction into veterinary practice is the complex Tticlabendazole, with a water-soluble polysaccharide arabinogalactan, extracted from larch, environmentally and safe product that is widely used in medicine. It is known that water-insoluble anthelmintic, applied to animals orally, to 70.0% excreted into the environment unchanged in the faeces, contaminating the environment. Water-soluble supramolecular complexe Tticlabendazole used in a reduced dose, soaked in blood, provide a high biological availability. In the future, they are broken down and eliminated in the urine from the body in miniscule amounts, without polluting the environment. These positive signs supramolecular complex compared with the substance triclabendazole anthelmintic and provide high economic efficiency and safety