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Evaluation of Anticancer and Antimicrobial Activities of Selected Medicinal Plants of Kashmir Himalayas, India

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Evaluation of Anticancer and Antimicrobial Activities of Selected Medicinal Plants of Kashmir Himalayas, India Indian Journal of Traditional Knowledge Vol. 16 (1), January 2017, pp. 141-145 Evaluation of anticancer and antimicrobial activities of selected medicinal plants of Kashmir Himalayas, India Saleem Mushtaqa,b, Qazi Parvaiz Hassana,*, Rashmi Sharmac, Rabiya Majeedd, Abid Hamid Dard, Phalisteen Sultana, Inshad Ali Khanc, Sheikh Abid Alia & Md Niamat Alib aBiotechnology Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar- 190005, India; bCentre of Research for Development (CORD), University of Kashmir, Srinagar- 190006, India; cClinical Microbiology Division, CSIR- Indian Institute of Integrative Medicine, Canal Road, Jammu-180001, India; dCancer Pharmacology Division, CSIR - Indian Institute of Integrative Medicine, Canal Road, Jammu-180001, India E-mail: [email protected]. Received 03 February 2016, revised 09 August 2016 Medicinal plants are known for their multifarious use in folk medicine of Kashmir Himalayas, India. The present study was designed to evaluate the anticancer and antimicrobial activities of extracts of some medicinal plants. Three different types of extracts were prepared by cold maceration technique using methanol, hexane and water. Anticancer activity of extracts was determined by using Sulpharhodamine B assay on four human cancer cell lines. Antimicrobial activity was done by using microdilution method. The methanol extract of Euonymus hamiltonianus exhibited broad spectrum anticancer activity with IC50 value of 20, 14, 27 and 55µg/ml against Lung A-549, Colon HCT-116, Pancreatic MiaPaca and Breast MCF-7 cell lines, respectively. The lowest IC50 value of 14µg/ml was observed against Colon HCT-116 cancer cell line. Colorectal cancer is the second leading cause of cancer deaths worldwide. The methanol and aqueous extracts of Euphorbia wallichii specifically inhibited the growth of Staphylococcus aureus which is one of the foremost pathogens in the list of antibiotic resistant strains. The MIC values were found to be 128-256µg/ml. Euonymus hamiltonianus and Euphorbia wallichii with promising activities could act as important source of biologically active compounds. Keywords: Euonymus hamiltonianus, Euphorbia wallichii, SRB assay, Microdilution method IPC Int. Cl.8: A61K 36/00, A01D 4/04, C12P, C12M, C12N Medicinal plants have been used since time use can be categorized into four main classes of immemorial for the treatment of different ailments compounds: vinca alkaloids, epipodophyllotoxins, and continue to provide front-line pharmacotherapy taxanes, and camptothecins7. Natural products also for millions of people worldwide. About 80% of the continue to be the primary source for antibacterial population in developing countries depends on drug discovery as 66% of all the drugs currently medicinal plants for their primary health care needs1. approved as antibacterial agents are natural products Medicinal plants are an essential part of alternative or natural product derivatives8. 2 and complementary medicine . They have well- The high altitude areas of Kashmir Himalayas, established history, unique chemical diversity and 3 India with their rich biodiversity harbor about 2000 appealing biological properties . Drugs derived from plant species9. Many of these plant species are medicinal plants have an important advantage as they 4 still used traditionally by rural people in the treatment offer a stable market throughout the world . With the of various diseases. These medicaments are used rapid increase in drug resistance, there is an urgent in different forms such as decoctions, pastes need to discover new compounds with diverse and powders. However, it is important to obtain chemical structures and novel mechanisms of 5,6 a scientific validation for the possible use of action . Drug discovery from medicinal plants has these herbal drugs in the treatment of various played a crucial role in the treatment of cancer. diseases. Here, the anticancer and antimicrobial Anticancer agents from plants currently in clinical activities of ten selected medicinal plants of Kashmir —————— Himalayas, India were evaluated. These belong to *Corresponding author nine different families. 142 INDIAN J TRADIT KNOWLE, VOL. 16, NO. 1, JANUARY 2017 Materials and methods Anticancer activity Plant materials Chemicals Different medicinal plants were collected from RPMI-1640 medium, Streptomycin, Fetal bovine various high altitude regions of Kashmir Himalayas, serum, Mitomycin, Sodium bicarbonate, Phosphate India like Gulmarg, Gurez, Budgam, etc., situated at buffer saline (PBS), Trypsin and 5-Fluorouracil an altitude of around 5000-8000 feet. A summary of (5-FU) were purchased from Sigma chemical Co the ethno-botanical data of these plants is given in (MO, USA). Glacial acetic acid was purchased from Table 1. Plant specimens were deposited in the Fisher Scientific (Mumbai, India) and Trichloroacetic acid (TCA) from Merck specialties Private Ltd. Departmental herbarium. The plant names have been verified with the www.plantlist.org. Cell culture, growth conditions and treatment Different human cancer cell lines: breast cancer Preparation of extracts cell line (MCF-7), colon cancer cell line (HCT-116), Different plant parts such as roots, shoots and pancreatic cancer cell line (MiaPaCa) and lung leaves were shade dried and pulverized into powder cancer line (A-549) were obtained from American using a mechanical grinder. Three different types of Type Culture Collection (ATCC, Sigma). All the cell extracts, hexane, methanol and water were prepared lines were grown in RPMI-1640/MEM medium using the cold maceration technique. The plant containing 10% FCS, 100 U/ml Penicillin, 100 µg/ml material was submerged in the respective solvents Streptomycin and sterilized by filtering through 0.2 for 72 hrs at room temperature with regular shaking µm filter in laminar air flow hood. Cells were allowed and filtered through Whatman No 1 filter paper. to grow in CO2 incubator (Thermo Scientific USA) The residue was macerated twice with the same at 37 ºC with 98% relative humidity and 5% CO2 solvent overnight and filtered. The filtrates obtained gas environment. from each extraction were mixed and concentrated under reduced pressure at 45–50 °C by using Sulpharhodamine B assay for % growth inhibition rotavapor. The extracts obtained were kept at 4 °C for The cytotoxic property of different extracts of eight further use. medicinal plants was determined by Sulpharhodamine Table 1—Ethnobotanical data of plant species screened for anticancer and antimicrobial activities Name of the plant species; Plant parts Site of Solvent used Traditional uses Family; Local name tested collection Aconitum heterophyllum Root Gulmarg Methanol Water Roots stored by nomadic tribes and used for different Wall. ex Royle; Ranunculaceae; medicinal purposes17. Patis, Paewakh Artemisia maritima L.; Root Gurez Hexane The water extract is used against skin eruptions as Compositae; Murin Methanol Water external use17. Berberis lycium Royle; Leaves Budgam Hexane Paste prepared from fresh fruits is used to heal Berberidaceae; Kawdach MethanolWater wounds18. Euonymus hamiltonianus Berries Budgam Methanol Water The plant species of this family have cytotoxic, Wall.; Celastraceae; antitumor and immunosuppressive activity19. Shoorkul, Sheelkul Euphorbia wallichii Hook.; Root Gurez Hexane Used as a folk medicine against skin problems17. Euphorbiaceae; Heerab Aerial Methanol Water Ficus carica L.; Fruit Budgam Methanol Dried fruit decoction is given against abdominal pain, Moraceae; Anjeer cold, asthma, constipation, jaundice, kidney and gallbladder stones, indigestion, liver enlargement and as a blood purifier20. Ligustrum lucidum L.; Oleaceae; Berries Srinagar Hexane Used against various infectious diseases in Chinese Privat kul MethanolWater traditional medicine21. Saussurea costus (Falc.) Lipsch.; Root Gurez Water Bitter rhizome powder is used to treat joint pains, Compositae; Kuth back pain, sole ulcers, dysentery and ever18. Viburnum grandiflorum Wall. ex Berries Gurez Hexane Used in the treatment of typhoid and whooping DC.; Adoxaceae; Kulmanh Methanol Water cough18. Viscum album L.; Santalaceae; Shoot Budgam Hexane Used against hypertension and cancer17. Ahul Methanol Water MUSHTAQ et al.: ANTICANCER AND ANTIMICROBIAL ACTIVITIES OF SELECTED MEDICINAL PLANTS 143 B (SRB) assay10. In the present study, all the cell lines fungal cultures 1 x 103 CFU/ml was used. The plates seeded in flat bottomed 96-well plates were allowed were incubated at 37 oC for 24 hrs for bacterial o to adhere overnight in a CO2 incubator set at 37 ºC. cultures and 28 C for 48 hrs for fungal cultures. Different plant extracts were added to the wells at The plates were read visually and the minimum 50 µg/ml concentration followed by incubation for concentration of the plant extracts showing no 48 hrs. After incubation period of 48 hrs, the cell turbidity was recorded as MIC. growth was stopped by adding 50 µl of chilled 50% TCA to each well and plates were further incubated Results and discussion at 4 ºC for an hour to fix the cells attached to bottom Anticancer activity of the wells. The plates were washed five times Different extracts of eight medicinal plants with distilled water and air dried. The cells were evaluated for anticancer activity varied in their stained with 100 µl per well SRB dye (0.4% w/v SRB efficiency and selectivity against the four human in 1% acetic acid) and the plates left at room cancer cell lines tested, viz. Lung A-549, Breast temperature for 30 min. Then plates
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