Evaluation of Acute and Subacute Toxicity, Teratogenicity, Reproductive Toxicity and Extended First Generation Developmental Toxicity of Ethanol Leaf Extract of Urtica simensis Hochst. Ex A. Rich in Wistar Albino Rats
| dc.contributor.advisor | Girma Seyoum | |
| dc.contributor.author | Bickes Wube | |
| dc.date.accessioned | 2026-06-22T14:36:59Z | |
| dc.date.available | 2026-06-22T14:36:59Z | |
| dc.date.issued | 2025-10-11 | |
| dc.description.abstract | Introduction: Urtica simensis is a native nutraceutical herb in Ethiopia. It has been used to treat various ailments such as malaria, hypertension, diabetes, gonorrhea, gastritis, body swelling, and wound infections. However, the safety of its repeated oral intake has not been evaluated yet. Therefore, this in-Vivo experimental study was conducted to evaluate the acute and subacute toxicity, teratogenicity, reproductive toxicity, and extended first-generation developmental toxicity in Wistar albino rats. Methods: For the acute and subacute toxicity study, sixty rats (30 males and 30 females) were randomly assigned to six groups. The median lethal dose (LD₅₀) was determined in the acute toxicity study. Groups’ I to III received 250, 500 and 1000 mg/kg of U. simensis leaf extract daily for 4 weeks, while Group IV received distilled water. The satellite 1000 mg/kg and satellite control groups were monitored for an additional 2 weeks. After 4 weeks, rats were euthanized for organ weight, blood chemistry and histopathological evaluations. For teratogenic studies of the leaf extract and essential oil, fifty pregnant rats were randomly assigned to five groups of ten gravid rats for each embryo and fetal experiments. Groups I to III were given the leaf extract and the essential oil at doses of 250, 500 and 1000 mg/kg daily from the 6th to the 12th days of gestation, respectively. Groups IV and V were pair fed and ad libitum controls. The developing embryos and fetuses were retrieved on the 12th and 20th days of gestation, respectively. Embryos were evaluated for growth and developmental delays. Fetuses were assessed for growth retardation, external and visceral anomalies. The skeletal ossifications and histopathology of fetal placentae were also evaluated. For the reproductive toxicity study, 20 rats per sex and group were randomly assigned into four groups. Groups’ I to III were treated with 250, 500 and 1000 mg/kg leaf extract daily for 10 weeks, respectively: two premating, two mating, three pregnancy, and three lactational weeks. The effects of leaf extract on the daily food intake, body weight changes, and histology of primary reproductive organs, liver and kidneys were assessed and evaluated. Biochemical tests including thyroid function were measured and analyzed. Reproductive indices and pregnancy outcomes were also assessed and evaluated. The pups were assessed for gross anomalies at birth. The weights of pups were measured on the postnatal days zero, four, seven, fourteen and twenty one. In addition, the pups' anogenital distances were measured on postnatal day zero. For the extended first generation developmental toxicity studies, the pups were randomly assigned into three cohorts at weaning: F1 cohort-1 for developmental reproductive toxicity, F1 cohort-2 for developmental neurotoxicity and F1 cohort-3 for developmental immunotoxicity. All pups assigned to the three cohorts were orally treated on a daily basis with similar doses of leaf extract as the parental rats. F1 cohort-1 rats, 20 rats per sex and group, were administered up to postnatal day 90. The tests conducted on the parental rats were repeated on F1 cohort-1 pups. F1 cohort-2 rats, 10 rats per sex and group, were treated until postnatal day 70, and the effects of the extract on the body weight and histopathology of brain and spinal cord were evaluated. F1 cohort 3 rats, 10 rats per sex and group, received the leaf extract daily until postnatal day 60, and the effects of the extract on body weight and histopathology of the spleen, thymus, and mandibular lymph nodes were evaluated. Results: The LD₅₀ exceeded 5 g/kg animal body weight. No adverse effects were observed at 250 mg/kg. However, 1000 mg/kg subacute dose caused significant weight gain in both male and female rats. Doses of 500 and 1000 mg/kg significantly elevated ALT in both sexes of rats. Male rats given 1000 mg/kg showed significantly increased serum creatinine, while females exhibited reduced hemoglobin. Some female rats treated with 1000 mg/kg had liver parenchymal necrosis, kidney glomerular distortion and spleen white pulp depletion. In the embryonic experiment, somite numbers and morphological scores were significantly decreased in gravid rats given 1000 mg/kg of leaf extract. Embryonic developments of the caudal neural tube (CNT), otic system, olfactory system and limb buds were significantly delayed in gravid rats given 1000 mg/kg of extract. The 500 mg/kg dose also caused significant developmental delays in the CNT and olfactory system. In the fetal experiment, fetal resorption was significantly increased; whereas crown rump length and fetal weight were significantly decreased in pregnant rats given 1000 mg/kg of leaf extract. Oral doses of 250 and 500 mg/kg of essential oil did not reveal adverse observed effects in all embryos and fetuses. However, somite numbers and morphological scores were significantly decreased in pregnant rats treated with 1000 mg/kg of essential oil. Embryonic developments of the caudal neural tube and forebrain were significantly delayed in pregnant dams administered 1000 mg/kg of essential oil. The crown rump length and fetal weight were significantly decreased in gravid rats given 1000 mg/kg of essential oil. The gravid rats received 1000 mg/kg of essential oil also revealed a significant increase in fetal resorption. In the reproductive toxicity study, female parental rats treated with 500 and 1000 mg/kg of leaf extract showed a significant pre-gestational weight gains. However, during gestation, the 1000 mg/kg treated female parental rats exhibited significantly lower weight gains. Lactational weight changes remained similar across all groups. Similarly, male parental rats treated with 1000 mg/kg of leaf extract revealed a significant weight gains. On the other hand, female parental rats treated with 1000 mg/kg of leaf extract had significantly lengthened estrous cycles. Both parental rats treated with U. simensis leaf extract did not have any adverse effects on mating or fertility indexes. However, the rats treated with 500 and 1000 mg/kg doses showed significant prolonged mating latency. Female parental rats administered the leaf extract at 500 and 1000 mg/kg doses revealed a significant decrease in litter size. Both parental male and female rats treated with 500 and 1000 mg/kg of leaf extract exhibited liver parenchymal necrosis. The biochemical analysis of rats treated with highest dose of leaf extract revealed a significant rise in liver enzyme markers. In the first offspring (F1) cohort-1 study, male rats received the 1000 mg/kg dose showed significant weight gains. Female cohort-1 rats treated with 500 and 1000 mg/kg doses also revealed a significant weight gains. In addition, F1 cohort-1 rats treated with 1000 mg/kg leaf extract showed significant prolongation of the estrous cycle and vaginal cornification. Both male and female offspring exhibited statistically significant elevations in liver enzyme tests at the highest dose, supporting findings in the parental rats. Furthermore, F1 cohort-1 rats that received 500 and 1000 mg/kg of extract also exhibited liver parenchymal necrosis, similar to the observations made in their parental rats. The F1 cohort-2 rats treated with U. simensis leaf extract did not show any signs of behavioral aberrations. The histopathological assessment of F1 cohort-2 rats treated with leaf extract did not reveal any structural changes of tissue sections taken from cerebral cortex, cerebellum, spinal cord, sympathetic ganglia and peripheral nerve fibers. The F1 cohort-3 rats treated with U. simensis leaf extract did not have any signs and symptoms of toxicity during the daily cage side clinical observations. The histological examination of key immunological organs (spleen, thymus, and mandibular lymphoid) of F1 cohort-3 rats treated with the leaf extract revealed no histological abnormalities. Conclusions: In this study, the no observable adverse effect level (NOAEL) for U. simensis leaf extract was 250 mg/kg, while the lowest observable adverse effect level (LOAEL) was 500 mg/kg. The adverse effects observed at the LOAEL were elevated liver enzymes with parenchymal necrosis, embryo-fetal developmental delays, and prolonging of some reproductive functions particularly lengthening of estrous cycle and mating latency. These findings suggest that the repeated oral intakes of high-dose U. simensis derivatives for purported nutraceutical benefits are not without risks and should be approached with caution, particularly during pregnancy and reproductive ages. | |
| dc.identifier.uri | https://etd.aau.edu.et/handle/123456789/8284 | |
| dc.language.iso | en | |
| dc.publisher | Addis Ababa University | |
| dc.subject | Urtica simensis | |
| dc.subject | Teratogenicity | |
| dc.subject | Developmental Toxicity | |
| dc.subject | Reproductive Toxicity | |
| dc.title | Evaluation of Acute and Subacute Toxicity, Teratogenicity, Reproductive Toxicity and Extended First Generation Developmental Toxicity of Ethanol Leaf Extract of Urtica simensis Hochst. Ex A. Rich in Wistar Albino Rats | |
| dc.type | Dissertation |