A toxicological evaluation for safety assessment of ruthenium-based diosmetin complex in rats.

The flavonoid-based organometallic complexes have been identified as novel bioactive compounds with enhanced pharmacological and therapeutic activity. In this study, the ruthenium-p-cymene diosmetin complex was synthesized, characterized, and investigated for toxicological profiling through different toxicological and genotoxicological studies which include acute and sub-acute toxicity, chromosomal aberration, and bone marrow micronucleus study. The acute oral toxicity study demonstrated the LD50 dose of the complex at 500 mg/kg body weight which further instigated the sub-acute doses i.e. 50, 100, and 200 mg/kg. The histopathological analysis demonstrated that the 400 mg/kg dose was associated with severe toxicological incidences of the vital organs (liver, kidney, pancreas, testis, and stomach) except the ovary with increased levels of ALP, AST, ALT, and WBC count. However, 50, 100, and 200 mg/kg doses did not show any toxicological alteration and maintained the normal levels of hematological and serum biochemical parameters. The genotoxicological assessment of the complex depicted no such genetic damage or mutagenicity in any complex treated groups. In conclusion, the 50, 100, and 200 mg/kg doses were determined as therapeutic dose of the novel ruthenium-p-cymene diosmetin complex without any genotoxic and mutagenic potential which can be further implemented in the investigation of various pharmacological and therapeutic interventions.

Safety assessment of marigold flavonoids from marigold inflorescence residue.

ETHNOPHARMACOLOGICAL RELEVANCE: Marigold flavonoids, extracted from marigold (Tagetes erecta L.) inflorescence residues, have attracted significant attention with respect to antioxidant, anti-inflammatory and chelating properties. However, the toxicity of marigold flavonoids have not yet been fully investigated. AIM OF THE STUDY: The main purpose of this study was to assess the safety of marigold flavonoids extracted from Marigold (Tagetes erecta L.) in order to provide information on its nonclinical safety. Thus, the acute oral toxicity, in vitro Ames test, sperm aberration study, bone marrow micronucleus test, subchronic oral toxicity test, and teratogenic potential were carried out in rats or mice. MATERIALS AND METHODS: For an acute oral toxicity test, SD rats and ICR mice (male and female, n = 5) orally received a single dose of 5000 mg/kg marigold flavonoids. Evaluation of marigold flavonoids genotoxic potential with a battery of tests, including an in vitro bacterial reverse mutation test using four mutant strains of Salmonella typhimurium (TA97、TA98、TA100、TA102), an sperm aberration test and an in vivo micronucleus test using bone marrow cells ICR mice that were orally administered marigold flavonoids, an subchronic oral toxicity study and teratogenic test employing male and female SD rats that were orally administered marigold flavonoids. All animals tests were completed in accordance with GB 15193 for toxicity tests. RESULTS: In the acute oral toxicity test, marigold flavonoids given at the dose of 5000 mg/kg body weight for 14 days didn't produce any abnormal clinical symptoms or mortality in SD rats and ICR mice (both sex, n = 5). There was no evidence of genotoxicity of marigold flavonoids based on the results of the in vitro bacterial reverse mutation test (up to 1250 µg/plate), the sperm aberration test (up to 5000 mg/kg body weight), the in vivo micronucleus test (up to 5000 mg/kg body weight), the subchronic oral toxicity study (up to 10 g/kg feed dose) and the teratogenic test (up to 1250 mg/kg body weight). CONCLUSIONS: We found that marigold flavonoids are safe with regard to acute toxicity in rats or mice as well as genotoxicity such as mutagenesis or clastogenesis under the present experimental conditions. These results might support the safety of marigold flavonoids as a potential therapeutic material for the traditional use of herbal medicines and for the further development of novel antioxidant.

Standardization, in-silico and in-vivo safety assessment of methanol extract of Ziziphus mauritiana Lam leaves.

Ziziphus mauritana Lam leaves were used to treat asthma, diabetes, pain, and inflammation in the Indian traditional system of medicine. The leaves of the Ziziphus mauritiana Lam were consumed as a vegetable in Indonesia and India. The present study aims to predict the pharmacokinetic properties of flavonoids identified & quantified through U(H)PLC and to evaluate the safety of methanol extract of Ziziphus mauritana Lam leaves (MEZ) in rats. A U(H)PLC-ESI-QTOF-MS/MS was performed to identify flavonoids present in MEZ and quantified using U(H)PLC method. The in-silico ADME properties of the flavonoids were analyzed using Schrodinger Maestro software. The acute oral toxicity study was performed by administering a single dose of MEZ (5000 mg/kg) in female rats and observed for 14 days. The sub-chronic studies were carried out by oral administration of MEZ at 500, 750, and 1000 mg/kg daily for 90 days. The changes in hematological parameters, clinical biochemistry, and histopathology were observed after the treatment period. Eight flavonoids rutin, kaempferol, luteolin, myricetin, catechin, and apigenin were identified from were identified in UPLC-QTOF-MS/MS analysis. These results showed the highest amount of luteolin (5.41 µg/ml) and kaempferol (4.02 µg/ml) present in MEZ. No signs of toxicity or mortality were observed in acute toxicity studies. In the sub-chronic studies, data showed that MEZ does not produce any changes in hematological and clinical biochemical parameters compared to control rats. MEZ (1000 mg/kg) significantly (p < 0.05) reduced total cholesterol, triglycerides, in male rats, which was more prominent on day 90. The histopathological analysis also revealed no changes in the vital organs. These results conclude that MEZ was considered safe and well-tolerated in rats.

Assessment of the risks of copper- and zinc oxide-based nanoparticles used in Vigna radiata L. culture on food quality, human nutrition and health.

The present article aims to assess the phytotoxic effects of copper and zinc oxide nanoparticles (Cu NPs, ZnO NPs) on mung bean (Vigna radiata L.) and their possible risk on food quality and safety. We also study the molecular mechanisms underlying the toxicity of nanosized Cu and ZnO. Seeds of mung bean were germinated under increasing concentrations of Cu NPs and ZnO NPs (10, 100, 1000, 2000 mg/L). We analyzed levels of free amino acids, total soluble sugars, minerals, polyphenols and antioxidant capacity. Our results showed that depending on the concentrations used of Cu NPs and ZnO NPs, the physiology of seed germination and embryo growth were modified. Both free metal ions and nanoparticles themselves may impact plant cellular and physiological processes. At 10 mg/L, an improvement of the nutritive properties, in terms of content in free amino acids, total soluble sugars, essential minerals, antioxidant polyphenols and flavonoids, was shown. However, higher concentrations (100-2000 mg/L) caused an alteration in the nutritional balance, which was revealed by the decrease in contents and quality of phenolic compounds, macronutrients (Na, Mg, Ca) and micronutrients (Cu, Fe, Mn, Zn, K). The overall effects of Cu and ZnO nanoparticles seem to interfere with the bioavailability of mineral and organic nutrients and alter the beneficial properties of the antioxidant phytochemicals, mineral compounds, phenolic acids and flavonoids. This may result in a potential hazard to human food and health, at some critical doses of nanofertilizers. This study may contribute in the guidelines to the safe use of nanofertilizers or nanosafety, for more health benefit and less potential risks.

Preclinical safety evaluation of the aqueous extract from Mangifera indica Linn. (Anacardiaceae): genotoxic, clastogenic and cytotoxic assessment in experimental models of genotoxicity in rats to predict potential human risks.

ETHNOPHARMACOLOGICAL RELEVANCE: Medicinal plants widely used by the population contain significant concentrations of biologically active compounds and, although they have proven pharmacological properties, can cause DNA damage and develop fatal diseases. AIM OF THE STUDY: The present study aimed to evaluate the genotoxic, cytotoxic potential and clastogenic effects of the aqueous extract from Mangifera indica leaves (EAMI) on rats submitted to experimental genotoxicity models and through the SMART test performed in Drosophila melanogaster. MATERIAL AND METHODS: The comet assay and the micronucleus test were performed on peripheral and bone marrow blood, respectively, of Wistar rats, orally treated with EAMI at doses of 125, 250, 500 and 1000 mg/kg/bw for 28 days. In the SMART test, the standard cross between three mutant D. melanogaster strains was used. Larvae were treated with EAMI at different concentrations, and the wings of adult flies were evaluated for the presence/frequency of mutant spots and compared to the negative control group. RESULTS: Phytochemical analysis of EAMI indicated high levels of flavonoids. The tests performed in rats showed that EAMI did not present significant genotoxic or clastogenic effects. The results showed a critical dose-dependent cytoprotective effect exerted by EAMI. This result was attributed to the high content of polyphenols and flavonoids. The biotransformation metabolites of EAMI did not present genotoxic activity, as demonstrated by the SMART test. CONCLUSIONS: These results are relevant since they provide safety information about a plant species of great therapeutic, economical, nutritious and ethnopharmacological value for the population.

Skin and Eye Irritation Assessment of Oil Palm ( Elaeis guineensis) Leaf Extract for Topical Application.

Many types of phytochemicals have been found to be present in oil palm leaf and could potentially be used as functional ingredients for skincare product. However, as of today, there is no published report on hazard identification and safety assessment of oil palm ( Elaeis guineensis) leaf extract (OPLE), particularly on skin and eye irritation. In this study, potential hazard of OPLE on skin and eye irritation was evaluated as an initial step to the safety assessment of OPLE. In vitro cell viability study of OPLE on normal human dermal fibroblasts showed that OPLE was nontoxic to the cells with percentage viability more than 90% after 24 and 48 hours of incubation. Skin irritation potential of OPLE was evaluated using in vitro SkinEthic reconstructed human epidermis (RHE) model (Organization for Economic Cooperation and Development [OECD] Test Guideline 439, 2015), while eye irritation potential was evaluated using in vitro SkinEthic Human corneal epithelium (HCE) model (OECD test guideline 492, 2017). Hazard identification results showed that OPLE at 1%, 5%, and 10% (wt/wt) was classified as nonirritant to the skin and eye where mean tissue viabilities of SkinEthic RHE and SkinEthic HCE were more than 50% and 60%, respectively. Therefore, we recommend a further safety assessment, such as human patch testing, to confirm the nonirritant of OPLE.

Safety assessment and single-dose toxicokinetics of the flavouring agent myricitrin in Sprague-Dawley rats.

Myricitrin, a flavonol rhamnoside of myricetin extracted from the Chinese bayberry (Myrica rubra Siebold) plant, has been used in Japan since 1992 as a flavour modifier in snack foods, dairy products, and beverages. It is affirmed as generally recognised as safe (GRAS) by the US Flavour and Extract Manufacturer Association (FEMA) and is considered safe by the Joint FAO/WHO Expert Committee on Food Additives (JECFA) at current estimated dietary exposures. In anticipation of expanded marketing, 97% pure myricitrin was fed to male and female Sprague-Dawley rats at dietary concentrations of 0.5%, 1.5% and 5.0% in a 90-day toxicity study. There was increased food consumption and decreased body weight gain in males exposed to 5% myricitrin. Blood values were within laboratory reference ranges except for mean increases in basophils in low- and high-dose males and serum phosphorus in high-dose males. In the absence of abnormal clinical or histopathological changes, these changes are not considered adverse. Based on the 90-day rat toxicity study, the no observed adverse effect level (NOAEL) is 2926 mg kg(-1) day(-1) in males and 3197 mg kg(-1) day(-1) in females. Gavage administration of myricitrin resulted in blood levels of myricitrin within 1 h after single oral doses of 250, 500 or 1000 mg kg(-1) body weight, indicating direct absorption of the glycosylated form of this flavonoid. Blood levels of myricetin, a metabolite of myricitrin, were not present in rats dosed orally with 1.6 mg kg(-1) myricetin, but were present only at 12 or 24 h in one of five, in three of five, and in four of five rats dosed with 250, 500 and 1000 mg myricitrin kg(-1) body weight, respectively, possibly a result of hepatic conversion of myricitrin to myricetin and enterohepatic recirculation of the resulting myricetin. The current studies further support prior safety assessments of myricitrin as a food flavouring.

Assessment of DNA damage induced by extracts, fractions and isolated compounds of Davilla nitida and Davilla elliptica (Dilleniaceae).

Davilla nitida and Davilla elliptica (Dilleniaceae) are plants that occur predominantly in the cerrado region of South America. They are used in popular medicine to treat stomach diseases, diarrhea and swelling, particularly of the lymph nodes and testicles. Chemical investigation of these two plant species led to the identification of the compounds myricetin-3-O-α-l-rhamnoside (myricitrin), quercetin-3-O-α-l-rhamnoside (quercitrin), myricetin, quercetin and gallic acid derivatives in the leaves of D. nitida and D. elliptica. Therefore, it was concluded that the two species of Davilla possess qualitatively similar chemical profiles. In the present study, the mutagenic and genotoxic potential of these plants and of their isolated compounds was tested in the Salmonella typhimurium assay (Ames test) with strains TA100, TA98, TA102 and TA97a, in the micronucleus test with peripheral blood cells of mice treated in vivo, and in plasmid DNA to analyze DNA strand-breaks. In the assessment of mutagenic potential by the Ames test, extracts from both plant species and a D. nitida ethyl-acetate fraction induced positive responses. On the other hand, none of the extracts showed genotoxic activity in the mouse cells. In the presence of metal ion, D. nitida and D. elliptica aqueous and ethyl-acetate fractions, as well as their isolated compounds, induced single- and double-strand-breaks in plasmid DNA in a cell-free system.

Phytochemical characterization of Rhododendron ferrugineum and in vitro assessment of an aqueous extract on cell toxicity.

Within a systematic phytochemical investigation of the leaves of RHODODENDRON FERRUGINEUM L. (Ericaceae), the volatile oil was isolated (0.7 %) and its constituents were characterized. Eleven flavonoids were isolated and characterized, with quercetin 3- O-[6''- O-(2-methyl-1-oxobutyl)]-ß- D-glucopyranoside and 2 R,3 R-dihydromyricetin 3- O- ß- L-arabinopyranoside as new natural products. Beside monomeric flavan-3-ols (catechin, epicatechin, gallocatechin, epigallocatechin) from the tannin fraction (about 3.5 % calculated as pyrogallol), the dimeric procyanidins B1 to B7 were identified, as well as the trimeric compounds procyanidin C1, epicatechin-(4 ß → 8)-epicatechin-(4 ß → 8)-catechin and the trimeric A type-linked cinnamtannin B1. Additionally, phloroacetophenon 4- O- ß- D-glucopyranoside and chlorogenic acid were isolated. Water-soluble carbohydrates comprised about 13.5 % of the dried leaves, including fructans (3 %), polysaccharides (1 %) (mainly type II arabinogalactans), glucose, fructose, sucrose, stachyose and raffinose. The IN VITRO effects on cellular vitality (MTT test), proliferation (BrdU incorporation) and necrosis (LDH release) of an aqueous extract were investigated. The extract did not exert any toxic effects, while the vitality and the proliferation rates of epithelial HaCaT keratinocytes were significantly increased at 100 µg/mL, indicating that the aqueous extract does not have negative effects against cellular activity.