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Genotoxic and cytotoxic potential of whole plant extracts of Kalanchoe laciniata by Ames and MTT assay.

ABSTRACT: Lack of data on safety of herbal medicines have endangered human health and life. The present study evaluated the genotoxic and mutagenic effect of Kalanchoe laciniata to access the safety and usefulness of the medicinal plant. Aqua-methanolic and n-hexane extracts of K. laciniata were evaluated for the genotoxic potential using Ames assay and cytotoxicity was evaluated using MTT assay. Ames assay was conducted using two strains of Salmonella typhimurium TA-100 and TA-102 whereas MTT assay was performed on baby hamster kidney cell line BHK-21. Aqua-methanolic extract of K.laciniata exhibited significant mutagenicity when exposed to TA-102 strain with a mutagenic index of 50.66 and 54.74 at maximum dose 150 mg/plate. The extract was also mutagenic to TA-100 strain but to a lesser extent. M.I of n-hexane extract was 12.15 and 15.51 for TA-100 and TA-102 respectively. n-hexane extract was mutagenic but little difference was observed between results of two strains. Both extracts were found to be cytotoxic with an IC₅₀ of 321.9 and 638.5 µg/mL for aqua-methanolic and n-hexane extracts respectively. On the basis of results it was concluded that aqua-methanolic and n-hexane extracts of K.laciniata possess mutagenic and cytotoxic potential. It is suggested to explore the plant further to evaluate its safety in rodents and other species.

SUBMITTER: Sharif A

PROVIDER: S-EPMC5491922 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Genotoxic and cytotoxic potential of whole plant extracts of Kalanchoe laciniata by Ames and MTT assay.

Sharif Ali A Akhtar Muhammad Furqan MF Akhtar Bushra B Saleem Ammara A Manan Maria M Shabbir Maryam M Ashraf Muneeb M Peerzada Sohaib S Ahmed Shoaib S Raza Moosa M

EXCLI journal 20170424

Lack of data on safety of herbal medicines have endangered human health and life. The present study evaluated the genotoxic and mutagenic effect of Kalanchoe laciniata to access the safety and usefulness of the medicinal plant. Aqua-methanolic and n-hexane extracts of K. laciniata were evaluated for the genotoxic potential using Ames assay and cytotoxicity was evaluated using MTT assay. Ames assay was conducted using two strains of Salmonella typhimurium TA-100 and TA-102 wh ...[more]

PMID: 28694760

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Project description:Kalanchoe species are succulents occurring in tropical regions. They have many biological and pharmacological properties. In this study, the cytotoxic and antimicrobial activities of water and dichloromethane Kalanchoe fractions obtained from ethanol extracts of three Kalanchoe species-K. daigremontiana, K. pinnata, and K. blossfeldiana were estimated. The cytotoxic effect was assessed on human cancer cell lines-ovarian SKOV-3, cervical HeLa, breast MCF-7, and melanoma A375-by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. The antimicrobial activity was estimated on selected Gram-positive and Gram-negative bacteria strains and on Candida albicans. The phytochemical analysis of selected Kalanchoe extracts was conducted by LC-QTOF-MS. The obtained results showed that the water fraction of K. blossfeldiana was active both on the tested cancer cells (IC50 values were 28.28 ± 2.76 and 32.51 ± 0.69 µg/mL on HeLa and SKOV-3, respectively) and bacteria strains (MIC values were 16 and 32 µg/mL on S. epidermidis and S. aureus, respectively). The water fraction of K. pinnata also had a significant effect on S. epidermidis and S. aureus, with MIC values of 32 and 64 µg/mL, respectively. The water fraction of K. blossfeldiana triggered a decrease in mitochondrial membrane potential (MMP) and induced cell cycle arrest in the G2/M phase in the SKOV-3 and HeLa cells. This fraction did not significantly increase cellular oxidative stress level. The DPPH and ABTS assays revealed that the water fraction of K. blossfeldiana had a strong antioxidant effect (IC50 was 9.44 ± 0.06 and 3.17 ± 0.1 µg/mL, respectively). The phytochemical analysis of the extracts of K. blossfeldiana and K. pinnata revealed the presence of at least 218 main components. The most frequently occurring were flavonol glycosides (31 metabolites), phenylpropanoids (13 metabolites), gallic acid derivatives (13 compounds), benzoic acid derived compounds (14 metabolites), and acyclic alcohol glycosides (16 compounds). In addition, proanthocyanidins were detected mainly in K. blossfeldiana. The study indicates that the water fraction of K. blossfeldiana has significant biological potential and can be further investigated towards anticancer and antimicrobial application.

Project description:The MTT assay, based on the enzymatic reduction of the water-soluble, yellowish tetrazolium salt 3-(4,5-dimethylthiazol)-2,5-diphenyl-tetrazolium bromide (MTT) to purple formazan, is commonly used for assessment of cell viability and proliferation. Accurate performance by the MTT assay depends on complete solubilization of cells and formazan and stability of the colored solution. Comparison of different solubilization solutions revealed that dimethylformamide (DMF) and dimethyl sulfoxide (DMSO), buffered with ammonia buffer, pH 10, and containing 5% SDS, produced the best results. These two solvents provided rapid and complete solubilization of formazan and cells, with minimal background absorbance at 700 nm, good reproducibility (low interassay coefficient of variation), high sensitivity, and color stability for at least 24 h. A linear relationship between viable-cell number and formazan absorbance was preserved for cell densities up to ∼1 × 109 cells/mL for Gram-negative and Gram-positive microorganisms. Since MTT can be reduced by medium components in the absence of cells, blanks containing all medium components but no cells should be run simultaneously. Measurements at two wavelengths, one corresponding to absorption peak of formazan (570 nm) and a background absorbance far from the peak (700 nm), are necessary to avoid artifacts due to incomplete solubilization and turbidity. IMPORTANCE Reduction of the water-soluble tetrazolium salt 3-(4,5-dimethylthiazol)-2,5 diphenyl-tetrazolium bromide (MTT) to purple, water-insoluble formazan is commonly used for assessment of cell viability and proliferation. Spectrophotometric detection of formazan requires its solubilization. The solubilization solvent has a strong influence on data acquisition and often introduces artifacts, leading to misreading of results. This study offers a choice of solvents that minimize solubilization artifacts when the MTT test is applied to microbiological cultures.

Project description:The species Kalanchoe brasiliensis and Kalanchoe pinnata, both known popularly as "Saião," are used interchangeably in traditional medicine for their antiophidic properties. Studies evaluating the anti-venom activity of these species are scarce. This study aims to characterize the chemical constituents and evaluate the inhibitory effects of hydroethanolic leaf extracts of K. brasiliensis and K. pinnata against local effects induced by Bothrops jararaca snake venom. Thin Layer Chromatography (TLC) and High Performance Liquid Chromatography coupled with Diode Array Detection and Electrospray Mass Spectrometry (HPLC-DAD-MS/MS) were performed for characterization of chemical markers of the extracts from these species. For antiophidic activity evaluation, B. jararaca venom-induced paw edema and skin hemorrhage in mice were evaluated. In both models, hydroethanolic extracts (125-500 mg/kg) were administered intraperitoneally in different protocols. Inhibition of phospholipase enzymatic activity of B. jararaca was evaluated. The HPLC-DAD-MS/MS chromatographic profile of extracts showed some particularities in the chemical profile of the two species. K. brasileinsis exhibited major peaks that have UV spectra similar to flavonoid glycosides derived from patuletin and eupafolin, while K. pinnata showed UV spectra similar to flavonoids glycosides derived from quercetin and kaempferol. Both extracts significantly reduced the hemorrhagic activity of B. jararaca venom in pre-treatment protocol, reaching about 40% of inhibition, while only K. pinnata was active in post-treatment protocol (about 30% of inhibition). In the antiedematogenic activity, only K. pinnata was active, inhibiting about 66% and 30% in pre and post-treatment protocols, respectively. Both extracts inhibited phospholipase activity; however, K. pinnata was more active. In conclusion, the results indicate the potential antiophidic activity of Kalanchoe species against local effects induced by B. jararaca snake venom, suggesting their potential use as a new source of bioactive molecules against bothropic venom.

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Genotoxic and cytotoxic potential of whole plant extracts of Kalanchoe laciniata by Ames and MTT assay.

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Genotoxic and cytotoxic potential of whole plant extracts of Kalanchoe laciniata by Ames and MTT assay.

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