The investigation on an ethnic medicinal plant of Elsholtiza bodinieri Vaniot: Chemical constituents, acute, 28-day subacute and 90-day subchronic toxicity evaluation.

Elsholtiza bodinieri Vaniot, belonging to the family Lamiaceae, has important medicinal value in Yunnan province of China. Traditionally, its aerial parts have been used as an ethnomedicine to treat diaphoresis, headache, fever, cough, pharyngitis, dyspepsia, and hepatitis. However, the safety assessment of E. bodinieri is still unexplored. This study aimed to investigate the phytochemical constituents of the hot water extract from E. bodinieri (HEEB) and evaluate the 14-day acute, 28-day subacute and 90-day subchronic toxicity by oral administration in Sprague-Dawley (SD) rats. The chemical constituents of HEEB were analyzed by UHPLC-ESI-HRMS/MS. Firstly, SD rats were chosen for a single oral administration of the maximum dose of 5000 mg/kg to evaluate toxicity. Subsequently, consecutive 28-day subacute and 90-day subchronic toxicity assessments of HEEB were conducted on Sprague-Dawley (SD) rats through repeated doses of 2500, 1250, 625, and 312.5 mg/kg for the former, and 1500, 1000, and 500 mg/kg for the latter. For toxicity evaluation, hematology and serum biochemical indicators were determined, and major organs of the rats were collected to calculate organ coefficients. Additionally, hematoxylin-eosin (H&E) staining was performed on the collected tissues to assess histopathological changes induced by repeated oral administration of HEEB. A total of 23 compounds were identified by UHPLC-ESI-HRMS/MS analysis. Acute toxicity assessment revealed that oral administration of HEEB did not induce mortality and unnormal behavior changes in female rats over a 14-day period, suggesting that the approximate lethal dose (ALD) was higher than 5000 mg/kg. In consecutive 28-day and 90-day toxicity evaluations, HEEB doses of 2500 mg/kg and 1500 mg/kg resulted in hepatic and kidney tissue damage in both female and male rats, which was verified by the increased levels of AST, ALT, BUN, Na+, and Cl-. After the acute, 28-day subacute and 90-day subchronic toxicity evaluation, the No Observed Adverse Effect Level (NOAEL) was determined as 1000 mg/kg/day. These findings not only provided a safety information for its medicinal and edible application, but also promoted the further comprehensive development of this plant.

Chen T ，Teng S ，Yang H ，Zhao Y ，Zhang J ，Liu J ，Zhou W ，Liu Y ，Cheng G ... -  《-》

Nephroprotective and diuretic effect of Alternanthera brasiliana (L.) Kuntze leaf extract; acute and sub-acute toxicity assessment.

Alternanthera brasiliana (L.) Kuntze of the family Amaranthaceae has been extensively used in traditional medicinal practices in Brazil and India for its reputed efficacy in promoting diuresis, as well as treating wounds, inflammation, postnatal symptoms, diarrhea, and cough. Its selection for this study was driven not only by its ethnomedicinal significance but also by its rich phytochemical composition, including bioactive compounds such as flavonoids, saponins, and alkaloids, which are known to exhibit nephroprotective and diuretic effects. Additionally, while many species from the Amaranthaceae family have demonstrated similar therapeutic properties, A. brasiliana remains underexplored in this context. Therefore, this research aimed to scientifically evaluate its potential nephroprotective and diuretic activities, providing a pharmacological basis for its traditional uses. This experiment was designed to determine nephroprotective effect against cisplatin-induced kidney injury and diuretic effect of Alternanthera brasiliana (L.) in rats. This study also aimed to evaluate the toxicity of plant's extract by performing acute and sub-acute toxicity trials. In current study, the nephroprotective effect of aqueous-ethanol extract of A. brasiliana was evaluated after induction of kidney injury with cisplatin. Extract was given in three doses as 75 mg/kg, 150 mg/kg and 300 mg/kg. A diuretic activity was also performed by comparing results with control and standard (furosemide). Extract was given in three doses as 75 mg/kg, 150 mg/kg and 300 mg/kg. A 14 day trial for acute toxicity assessment was performed at doses 2000 mg/kg and 3000 mg/kg, whereas a 28 day trial for sub-acute toxicity assessment was performed at doses 250 mg/kg, 500 mg/kg and 1000 mg/kg. The biological active ingredients were identified and determined using HPLC technique. The aqueous-ethanol extract of A. brasiliana (ABAE) safeguarded the rats from toxic effects of cisplatin. This extract also enhanced urine output. The protective effect of ABAE increased with increasing dose and produced maximum nephroprotective effect and diuresis at a dose 300 mg/kg. The outcomes from acute toxicity trials suggested that LD50 lied beyond 3000 mg/kg, and no antagonizing effects occurred in sub-acute toxicity trials at doses 250 mg/kg, 500 mg/kg and 1000 mg/kg. ABAE posed no toxicities on kidney, liver, and heart tissues as evident from histopathological, hematological, and serum biochemical analysis. HPLC-DAD analysis of ABAE indicated the presence of betanin, kaempherol, gallic acid, p-coumaric acid and oxalic acid. These results demonstrate an abundant supply of bioactive chemicals found in A. brasiliana (L.) extracts, which should be taken into account to improve renal functions with fewer negative effects.

Bilal MH ，Bibi I 《-》

Toxicological evaluation of Chlamydomonas reinhardtii enriched with protoporphyrin IX.

The Chlamydomonas reinhardtii (red) is a strain rich in protoporphyrin IX, which could be used as algal-derived novel sustainable nutritional sources. However, the safety research on C. reinhardtii (red) remains insufficient and further investigation is needed. In order to assess the safety of C. reinhardtii (red) and to expand its use in food applications, a series of toxicological safety evaluations were carried out to observe acute toxicity, subchronic toxicity and genotoxicity of C. reinhardtii (red). The C. reinhardtii (red) was administered to an acute oral toxicity test, mammalian micronucleus test, mouse chromosomal aberration test in Institute of Cancer Research (ICR) mice, and 90-day repeated-dose oral toxicity test in Sprague-Dawley (SD) rats. In the acute oral toxicological test, no mortality or abnormalities were observed in male and female rats, with an median lethal dose(Lethal Dose, 50%) (LD50) exceeding 10.0 g/kg body weight (BW). Similarly, male and female mice exhibited no mortality or abnormalities, with an LD50 surpassing 20.0 g/kg BW. The results of bacterial reverse mutation test, mammalian micronucleus test, and mouse chromosomal aberration test were all negative, suggesting that the C. reinhardtii (red) did not show mutagenicity under the conditions of this experiment. In a 90-day repeated-dose oral toxicity study, the SD rats were orally administered daily doses of 2.0 g/kg BW, 4.0 g/kg BW, and 8.0 g/kg BW without exhibiting any signs or symptoms of toxicity. Therefore, the no observed adverse effect level (NOAEL) of C. reinhardtii (red) was 8.0 g/kg BW/day. This study contributes to information necessary to form a basis for safety for expanding the application range of C. reinhardtii (red) in food. PRACTICAL APPLICATION: The aim of this study was to assess the safety of C. reinhardtii (red) and to expand its use in food applications. The major finding of this study is that the C. reinhardtii (red) did not show acute toxicity, subchronic toxicity and genotoxicity under the conditions of this experiment. This study contributes to information necessary to form a basis for safety for expanding the application range of C. reinhardtii (red) in food.

Qu X ，Wang Q ，Xie Z ，Wu Y ，Zhang Y ，Liu Z ，Wang Y ... -  《-》

Toxicological evaluation of Artocarpus lacucha ethyl acetate extract: in vitro and in vivo assessment.

Artocarpus lacucha Buch. -Ham. (syn. Artocarpus lakoocha) (A. lacucha), is a tropical fruit tree and a member of the Moraceae family. Fruit, bark, foliage, and roots of A. lacucha are broadly utilized in folklore medicine to treat various metabolic and gastrointestinal disorders. The bark of A. lacucha has demonstrated antimicrobial and antinociceptive effects. The study investigated the toxicity of A. lacucha ethyl acetate bark extract (AL-EAE) employing both in vitro and in vivomodels. AL-EAE was extracted by soxhlation of powdered bark. The phytochemical fingerprint of AL-EAE was established employing LC-MS technique. The presence of heavy metals was determined by atomic absorption spectroscopy (AAS). Cytotoxicity of AL-EAE (3.9-250 μg/mL) was evaluated by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay in human embryonic kidney cells (HEK293). The acute oral toxicity of AL-EAE was ascertained by administering 500, 1000, and 2000 mg/kg B.W. doses in rats of both sexes. Rats were treated with AL-EAE orally at 200, 400, and 600 mg/kg B.W. doses on a daily basis for 28 days in order to evaluate subacute toxicity. Clinical and toxicological parameters were noted after every 24 h. After the course of treatment, the safety of AL-EAE was evaluated by investigating various biochemical, haematological, and histopathological markers. LC-MS analysis indicates the presence of phenols and flavonoids in A. lacucha bark extracts. AAS analysis revealed the safety limits of heavy metals in AL-EAE as recommended by ICH. AL-EAE was not found cytotoxic in HEK293 cells (IC50 more than 250 μg/mL). No adverse clinical syndromes or mortality were observed in the acute toxicity study. The alterations in body weight (B.W.) and relative organ weights (ROWs) were found to be non-significant. In the sub-acute toxicity assessment, changes in food intake and mild diarrhoea were observed at 600 mg/kg/day B.W. Changes in ROWs at 600 mg/kg/day and B.W. at 400 and 600 mg/kg/day dose were observed. However, significant alterations in biochemical (TG, VLDL, AST, and ALP) and haematological (haemoglobin, MCHC, eosinophils, monocyte) parameters were observed at 400 and 600 mg/kg/day B.W. doses. Histopathological studies illustrated significant inflammation in the liver, pancreas, and spleen at 600 mg/kg/day B.W. doses. The findings demonstrated that AL-EAE did not possess cytotoxicity in vitro. In both acute and subacute toxicity studies, AL-EAE was shown to be nontoxic, where 400 mg/kg/day was the no-observed-adverse-effect level (NOAEL) dosage in rodents.

Banerjee T ，Sarkar A ，De A ，Sar S ，Mandal A ，Panda P ，Halder AK ，Karmakar S ，Ghosh N ... -  《-》

Safety assessment and exploration of the mechanism of toxic effects of diallyl trisulfide: Based on acute and subacute toxicity and proteomics experiments.

Garlic (Allium sativum L.) is a widely consumed spice and condiment around the world, applied both as a food and as a traditional medicine, and is a natural strengthening agent for the body's circulatory and nervous systems. Diallyl trisulfide (DATS) is the major volatile organosulfur phytochemical found in garlic, with antithrombotic, anticoagulant, and antiplatelet activities as well as antioxidant, anti-infective, and other pharmacological effects. However, the safe dose and the underlying mechanisms of its toxic effects remain elusive. DATS, an important pharmacologically active compound found in garlic, has garnered attention for its ability to fight cancer, antioxidant, anti-infective, and cardioprotective. The aim of this study was to evaluate the safety of DATS and to elucidate the potential mechanisms of its toxicity. In this study, ICR mice were selected for acute and subacute toxicity experiments according to OECD guidelines. The toxicity profile of DATS was analyzed by computer prediction software. Also, key differential proteins in spleen and serum were analyzed by proteomics. The binding stability of DAST to differential proteins was analyzed by molecular docking. Additionally, the regulatory relationship between DATS and differential proteins was verified by Western blot and ELISA experiments. The results showed that the LD50 value of DATS in acute toxicity was 188.67 mg/kg. In subacute toxicity, water consumption and food intake were reduced in both male and female mice. In addition, the spleen and small intestinal organ coefficients were significantly elevated in male mice at the high dose of DATS; the blood biochemical indices ALB and TP were also significantly elevated. HE staining results showed significant damage to the spleen, liver, small intestine, and kidney of mice at high doses of DATS. Spleen and serum proteomics analyses showed that DATS significantly inhibited ZBP1 expression and upregulated TEC. ADMETlab 2.0 software predictions identified DATS as having potential genotoxicity, dermal sensitization, carcinogenicity, and respiratory and ocular toxicity. Docking results showed that the binding energies between DATS and TEC protein (PDB: 6F3F) and ZBP1 protein (PDB: 4KA4) were -3.7 kcal/mol, -2.4 kcal/mol, respectively. Western blot results showed that DATS-H significantly inhibited the expression of ZBP1 (only in male mice) and Bcl-2 proteins. ELISA results showed that DATS-H significantly increased the level of TEC protein both in male and female mice. Long-term administration of high-dose DATS may carry some risk of toxicity. Based on the amount of DATS in garlic, it is recommended that adults should not take more than 359 mg of DATS and 84.5 g of garlic per day. The mechanism of toxicity may be related to the fact that DATS significantly inhibits ZBP1 expression, upregulates TEC, and promotes apoptosis. This study provides valuable toxicological data for the effective evaluation of the long-term toxicity of DATS and offers an additional experimental basis for developing DATS as a healthy food or drug.

Wu Z ，Tu B ，Li S ，Chen J ，Shen P ，Zhou W ，Ma Z ，Tang X ，Xiao C ，Wang Y ，Gao Y ... -  《-》