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Title: Biological Activities and Phytochemical Analysis of Selected Medicinal Plants of Khyber Pakhtunkhwa, Pakistan.
Authors: Begum, Hussan Ara
Keywords: Botany
Issue Date: 2017
Publisher: Abdul Wali Khan University, Mardan
Abstract: Secondary metabolites of medicinal plants are a reservoir of naturally active compounds which contributing to fulfill the needs of effective treatment of diseases. To accomplish these demands new analytical techniques have been introduced in the past to determining the current drug. Biological activities are used to provide evidences about presence of active compounds. At present plant-derived drugs are used to be more operative than modern drugs as they are less toxic to human health but effective in cure. Cucumis sativus, Portulaca oleracea, Malus baccata, Saxifraga flagillaris, Geranium wallichianum and Monotheca buxifolia were collected from local flora based on ethno botanical significance. The extracts of these plants were obtained in 70% ethanol and their subsequent fractions were made on polarity based with n-hexane, dichloromethane and ethyl acetate. The extracts were screened through various biological activities like antifungal, antimicrobial, phytotoxic, anticancer and anti-oxidant. These plants were also subjected to heavy metal, nutrients and phytochemical analysis with compound isolation and its application. Agar well diffusion method was used to evaluate the antifungal activity. All the tested fungal strains Alternaria, Acremonium, Verticellium, Pythium and Tricoderma were susceptible to the crude extracts and their fractions of selected medicinal plants. The crude extract of Geranium wallichianum was most active against all selected fungal strains with zone of inhibition from 12.83± 0.95 to 16.06± 0.97, followed by Portulaca oleracae with zone of inhibition ranging from 10.33± 0.88 to 17.00± 0.58. Saxifraga flagillaris showed zone of inhibition ranging from 7.90± 0.66 to 15.33± 1.129. Antibacterial activity was carried out using agar well diffusion method against Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Shigella flexneri and Salmonella typhi. Crude extract of Saxifraga flagillaris was most active against all selected bacterial strains showing zone of inhibition ranging from 11.67± 0.67 against Shigella flexneri to 17.67± 0.33 against Escherichia coli. Crude extract of Portulaca oleracae was very potent against Staphylococcus aureus with 26.17± 0.73 while showed mild activity against rest of the bacterial strains. Malus baccata crude extract also showed mild activity ranging from 11.00± 0.58 against Staphylococcus aureus to 13.41± 0.46 against Pseudomonas aeruginosa. Selected bacterial strains were resistant to Cucumis sativus crude extract showing least activity of 2.12± 0.12 against Salmonella typhi and 4.89± 2.47 against Shigella flexneri. Among the fractions, Saxifraga flagillaris n-hexane and DCM fractions were most potent against all selected bacterial strains. Zone of inhibition ranging with least value from 21.67 ±0.882 against Salmonella typhi by n-hexane fraction and high value of 25.33 ±1.4534 against Salmonella typhi by DCM fraction. It showed that both fractions have different secondary metabolites which affect the bacteria having different zones of inhibition. Fractions of rest of the medicinal plants have potent to mild antibacterial activity. Antioxidant activity was carried out with DPPH and ABTS assays. The ethyl acetate fractions of Saxifraga flagillaris, Malus baccata, Geranium wallichianum and hexane fraction of leaves of Monotheca buxifolia showed strongest activity with IC50 of 18.44, 15.11, 15.52 and 20.89 respectively. It suggested that the plants possess active antioxidant complexes for curing diseases. The results of preliminary phytochemical analysis of the crude extract of selected medicinal plants showed that the potentials of curing diseases are due to the presence of alkaloids, saponins, glycosides, flavonoids and phenols from low to moderate amounts. The heavy metals determination showed that very small amounts of Pb, As, Cd, Hg and Zn were present in these plants. The proximate analysis showed that these plants have considerable amounts of basic nutrients like carbohydrate, proteins, fat, ash and moisture. Sandwich method was used for determining phytotoxicity of these plants in terms of radicle and plumule length of Lactuca sativa seeds with different concentrations, i.e. 10, 20, and 40mg of powdered plant material. The 20mg concentration was more effective then 10 and 40mg. Anticancer activity was determined with different concentrations against HepG2 cancer cell line using MTT bioassay. All fractions of these plants were active against the HepG2 cancer cell line which showed varying degrees of cell viability due to the differences in secondary metabolites present in fractions of plants. The n-hexane, DCM and water fraction of Cucumis sativus were active anticancer fractions with 26.84%, 56.35% and 60.76% effectiveness. n-hexane fraction of Portulaca oleracae with 23.68 % effective. Saxifraga flagillaris n-hexane and DCM frictions were the most active with 8.31% and 10.06% cell viability having 92 and 90 % effectiveness respectively. N-hexane fraction of Geranium wallichianum showed 21.5% activity and leaf n-hexane fraction of Monotheca buxifolia showed 40.9% activity. One compound was isolated from extract of Saxifraga flagillaris named adlumine with molecular formula C21H21NO6 and molecular weight 383.400. This compound was for the first time isolated from Saxifraga flagillaris and was screened for cell viability activity, Apoptosis, ROS generation, Caspase-3 and western immunoblotting assays against cancer cell lines. The isolated compound was tested for anticancer activity. The compound adlumine reduce the cell viability of HCT 116 and PC3 cells. 20 μg/ml of adlumine individually killed 19.58% HCT 116 cells. In case of PC3 cells, adlumine caused 35.19% cell death at 10 μg/ml concentration. This compound proved toxic for HCT 116 and PC3 cell lines at ≥ 20 μg/ml and ≥ 10 μg/ml concentrations respectively. Adlumine increased early apoptosis to 16.8%, while boosted late apoptosis to 11.3%. Adlumine induced ROS generation in HCT 116 which may contribute in apoptosis induction. Adlumine induced caspase-3- dependent apoptosis in HCT 116 and PC3 cell lines. It is sured from the present study that this compound possess potent anticancer activity. The present findings suggested that Saxifraga flagillaris plant has potential secondary metabolites for curing cancer and other microbial and infectious diseases.
Gov't Doc #: 16809
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