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 of the ethanolic extract of Siparuna guianensis: Cell viability, molecular risk predictions and toxicity risk for acute and sub-chronic oral ingestion.

The species Siparuna guianensis Aublet (family Siparunaceae) is traditionally used by indigenous peoples and riverine communities in Central and South America to treat migraines, flu, respiratory diseases, fever, pain, edema and inflammatory and infectious diseases. Scientific studies on the species have proven its antiparasitic, insecticidal, antimicrobial, neuroprotective, anti-inflammatory, antioxidant and antinociceptive action. However, the safety profile of the species has not yet been established. To verify the levels of cytotoxicity, evaluate the molecular toxicological prediction in silico, determine the median lethal dose and verify the possible acute and sub-chronic toxicological effects of S. guianensis in rats. After obtaining the aqueous and ethanolic extracts of S. guianensis leaves, the total phenolic content, ABTS radical scavenging activity and cytotoxicity in fibroblasts were evaluated. The ethanolic extract was used in chromatography and in the in silico and in vivo studies. The in silico test evaluated carcinogenicity, mutagenicity and skin irritation. Acute oral toxicity and LD50 were evaluated after the single dose of 2000 mg/kg, with monitoring for 14 days. Sub-chronic toxicity was evaluated at doses of 200, 400 and 800 mg/kg for 30 days. Murinometric parameters, water and food consumption and feed efficiency were evaluated. At the end of the experiments, hematological, biochemical, macroscopic organ and histopathological analyses were performed. Cell viability was greater than 90 %, without cytotoxicity up to 25 μg/mL. In the in silico predictions, the molecules 2-undecanone, decanoic acid, decanoic acid ethyl ester and 2-tetradecanone showed no risk of carcinogenicity, mutagenicity, skin sensitization or eye irritation. The estimated LD50 was greater than 2000 mg/kg and daily oral use for 30 days was safe up to a limit of 800 mg/kg. The SG2000 group showed weight loss (p<0.01). In the hematological parameters, there was no difference between the groups (p>0.05), but in the biochemical findings, the urea rate was higher in the SG800 group (p<0.05), total proteins were higher in the SG400 group (p<0.05), while alkaline phosphatase was higher in the control group compared to the SG200, SG800 (p<0.01) and SG400 (p<0.05) groups. Triglycerides and VLDL-C were higher in the SG400 group, while non-HDL-C was higher in the SG800 group (p<0.05). The SG2000 group showed the lowest relative weights of the liver, spleen (p<0.05) and lungs (p<0.01), and the SG800 group showed increased weights for the liver (p<0.05) and lungs (p<0.01). Only the animals treated with a single dose of 2000 mg/kg showed histopathological changes in the liver, with slight cytoplasmic and tubular vacuolization in the kidneys. The concentration of 25 μg/mL showed no cytotoxicity. Four molecules were detected in silico did not present a risk of carcinogenicity in female mice, nor mutagenicity, skin sensitization or ocular irritation. In rats, the LD50 is greater than 2000 mg/kg. Daily oral use for 30 days at up to 800 mg/kg was considered safe, with no significant hematological or histological alterations. These results may support further studies and pre-clinical trials.

Rodrigues ESR ，Moreira RF ，Ramos RDS ，Souza SA ，Sotero Filho JWM ，Silva BJPD ，Jumbo LOV ，Oliveira EE ，Lima ES ，Aguiar RWS ... -  《-》

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 《-》

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 ... -  《-》

Toxicological analysis and anti-inflammatory and antioxidant evaluations of extract, fractions and secoxyloganin obtained from Guettarda viburnoides Cham. & Schltdl. in mice.

Guettarda viburnoides, "veludinho do campo," is traditionally used for the treatment of pain and inflammatory conditions in humans; however, only one scientific study has reported this effect in an ear inflammatory model. Therefore, it is necessary to explore other in vivo models and the chemical composition of this medicinal plant. A chemical investigation of methanolic extract of G. viburnoides (MEGV) (leaves) led to the isolation of secoxyloganin (GV-1). In addition, the preclinical safety of MEGV (in acute and subacute toxicological models, gavage = p.o.), antioxidants of MEGV, ethyl acetate (EAFGV) and hydromethanolic (HMFGV) fractions were tested using free radical scavenging and lipid peroxidation methodologies, and the anti-inflammatory effects of MEGV, HMFGV and GV-1 (p.o.) were evaluated on carrageenan and complete Freund's adjuvant (CFA) models of inflammation in mice. MEGV was obtained from air-dried leaves by maceration with methanol at room temperature. MEGV was then purified by liquid-liquid partitioning, to obtain the EAFGV and HMFGV fractions. Purification of HMFGV afforded GV-1. The quantification of total phenols, flavonoids, flavonols, and condensed tannins was subsequently performed for MEGV. The antioxidant activity of 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2, 2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid (ABTS) and oxidation of β-carotene were evaluated in MEGV and its fractions. The anti-inflammatory activity of MEGV (3, 30, and 100 mg/kg, p.o.) was assayed in carrageenan-induced models, followed by assessments of MEGV (100 mg/kg, p.o.), HMFGV (3 and 30 mg/kg, p.o.) and GV-1 (3 mg/kg, p.o.) in CFA-induced models in mice (including paw oedema, mechanical allodynia and cold sensitivity). Acute (14 days of MEGV, 2000 mg/kg, p.o.) and subacute (28 days, MEGV 30, 100, and 300 mg/kg, p.o.) toxicity was assessed in female Swiss mice. The major compound was secoxyloganin (GV-1). The oral acute toxicity test of MEGV revealed no evidence of toxicity, indicating low toxicity according to the Organization for Economic Cooperation and Development (OECD) guidelines. In the subacute toxicity group, no clinical signs of toxicity or changes in body weight, water consumption, food consumption, or organ weight or morphology were observed after 28 days of gavage with MEGV (30, 100, and 300 mg/kg) compared with those in the control group. MEGV, EAFGV, and HMFGV showed significant free-radical scavenging and lipid peroxidation activities, with IC50 values ≤ 26.38 ± 4.56 μg/mL. In in vivo anti-inflammatory assays, MEGV (3, 30 and 100 mg/kg) reduced carrageenan-induced oedema (2 and 4 h) and hyperalgesia (3 and 4 h). In the CFA model, MEGV (100 mg/kg), HMFGV (30 mg/kg) and GV-1 (3 mg/kg) reduced inflammation (at 3, 4 and 24 h) in all parameters (oedema, mechanical allodynia and cold sensitivity). This study revealed that G. viburnoides has antioxidant and anti-inflammatory properties, and no toxicity was detected after acute or subacute gavage with MEGV, validating its traditional use in the treatment of inflammatory conditions.

de Oliveira Junior PC ，Sanjinez Argandoña EJ ，Dos Santos SM ，Santos JMD ，de Souza MF ，Faoro JAM ，Kassuya CAL ，Arena AC ，Matos Manoel B ，Silva RMMF ，Formagio ASN ... -  《-》