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Phytochemical, Anticancer and Antioxidant Evaluation of Potential Chemical Consituents of Calliandria Surinamensis

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Phytochemical, Anticancer and Antioxidant Evaluation of Potential Chemical Consituents of Calliandria Surinamensis Nig J. Biotech. Vol. 21 (2010) 55 – 59 ISSN: 0189 17131 Available online at http://www.ajol.info/index.php/njb/index and www.biotechsocietynigeria.org. Phytochemical, Anticancer and Antioxidant Evaluation of Potential Chemical Consituents of Calliandria Surinamensis Falodun, A.1, 2Uzoekwe Anayo Stephen, and 3 Shengxiang Qiu, 1Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Benin, Benin City, Nigeria 2Department of Biochemistry, Faculty of Basic and Applied Sciences, Benson Idahosa University, Benin City, Edo State, Nigeria 3Natural Product Medicinal Chemistry and Drug Discovery, South China Botanical Garden, Chinese Academy of Sciences Guangzhou, 510650, China Abstract Some chemical constituents (β-Sitosterol, β-Sitosterol glucoside, β-amyrin, xanthone glycoside , and a flavanol glycoside) previously isolated and characterized from the stem bark of Calliandria surinamensis were subjected to in vitro free radical scavenging and anticancer activities using DPPH free radical scavenging method and lung cancer cell lines. Evaluation of the antioxidant and anticancer activities of the compounds revealed significant in-vitro antioxidant activity of the flavanol and xanthone glycosides. The xantone glycoside was active on the cancer cell line used for the study. The work reports for the first time the in-vitro antioxidant and anticancer profile of the isolated chemical compounds from the medicinal plant. Key words: anticancer, antioxidant, C. surinamensis, phytochemical investigation, plant extract Correspondence: [email protected]; [email protected] Introduction Huisman et al. 2004). Cancer is a dreadful Current research has shown that disease caused by abnormal and food rich in antioxidants plays an vital role in uncontrolled cell division. About 6 million the prevention of several diseases which new incidences of cancer are reported possess serious threat to man’s existence yearly worldwide. Nature has given man a such as cardiovascular diseases (Blois 1958; variety of useful sources of remedies to cure Emmanuel et al. 2007; Butler, 2008). Some a number of diseases. About 1500, natural synthetic antioxidants, such as 2 - and 3 - products have played a significant role in tert-butyl-4-methoxyphenol ( butylated drug discovery and development, especially hydroxyanisole, BHA), 2,6-di-tert-butyl-4- agents active against cancer and infectious methylphnol (i.e.butylated hydroxytoluene, diseases (Ioannis, 2008). More than 70 per BHT) and tert-butylhydroquinone (TBHQ) cent of all cancer deaths occurred in low- are widely used as antioxidants in foodstuffs and middle-income countries. The WHO and drugs but, because of toxicity issues, noted that tobacco use, alcohol use, low their use is being questioned (Falodun and fruit and vegetable intake, and chronic Irabor, 2008). Reactive oxygenic species in infections from hepatitis B (HBV), hepatitis C the form of superoxide anion (O2), hydrogen virus (HCV) and some types of HPV are peroxide (H2O2) and hydroxyl radical (HO) leading risk factors for cancer in low- and are natural by-products of our body’s middle-income countries. Deaths from metabolism. These chemical substances are cancer worldwide are projected to continue dangerous when present in excess, and can rising, however, with an estimated 12 attack biological molecules such as lipids, million deaths in 2030 (Lefer and Grander, proteins, enzymes, DNA and RNA, leading to 2000). The most frequent types of cancer cell or tissue injury associated with worldwide in order of the number of global degenerative diseases(Harborne, 1973; deaths are; among men – lung, stomach, Falodun et al. / Nig J. Biotech Vol. 21 (2010) 55 – 59 liver, colorectal, oesophagus and prostate; presence of some secondary metabolites and among women – breast, lung, stomach, such as alkaloids, tannins, sapnonins, colorectal and cervical. The need to screen flavonoids and anthraquinones. Standard medicinal plants for anticancer and procedures were used in the determination antioxidant properties cannot be of these metabolites (Sofowora, 1984; overemphasized. Trease et al. 1989; Siddhuraju et al. 2002). Calliandra surinamensis is a Extraction and preparation of plant perennial tree distributed in Nigeria, East material: The fresh stem bark of C. and South Africa. The flower pigment is surinamensis was dried at temperature of used as dye pigment for the manufacture of 30oC and reduced to a fine powder using a some drugs of pharmaceutical importance. mechanical mill. The powdered sample The plant is known in the tropics for its (1Kg) was extracted with methanol (4 X 25 ornamental/horticultural purposes. L) by maceration for 48 hours at room Calliandra surinamensis is a large multi- temperature. The combined methanol trunked shrub with detailed descriptive extracts were evaporated to dryness using a properties. The stem bark is used in folk rotary evaporator at reduced pressure. medicine for the treatment of various Cell line and culture: A-549 (human disease and infections such as cough, lung adenocarcinoma epithelial cell line) cell wound healing, inflammations. Scanty culture was obtained from Biomedicine information exists in the literature on the Research and Development Centre of Jinan phytochemical and biological investigation of University (Guangzhou, China). The cell line the plant. We have also reported the was cultured in growth medium (RPMI-1640 antioxidant activity, free radical, and medium, pH 7.4), supplemented with 10 % proximate analysis of the flower pigment fetal bovine serum (FBS) and antibiotics using DPPH assay procedure and in [penicillin (100 units/ml) and streptomycin comparison with standard antioxidants (Liu sulfate (100 µg/ml)]. The cell lines were and Wang 2008). The flower extract have grown in 50 cm2 tissue culture flasks also been shown to possess antibacterial (Corning, NY, USA) and used for cytotoxicity properties (McCune and Johns (2005). The assay. pharmacological importance of this plant Cytotoxicity assay: The cytotoxicity motivated us to explore this species for its assay of the plant samples were determined chemical constituents. using the method Zhao et al. (2007) MTT Therefore, the search for natural (3-(4, 5-dimethyl thiazole-2yl)-2, 5-diphenyl products represents an area of great tetrazolium bromide) assay. In brief, human research interest in which the plant kingdom cancer cells were plated at 2 104 cells per has been documented the most important well in 96 well microtiter plates (Costar source to provide many antioxidant and 3599, Corning, NY, USA ) with 100 µl RPMI- cancer chemopreventive agents with novel 1640 growth medium and incubated for 24 chemical structure (McCune and Johns h at 37C, with 5 % CO2 in a humidified (2005). atmosphere (Incu-Safe, Sanyo, Japan), during which period a partial monolayer was Materials and Methods formed. Later, the medium was removed Plant Material: Fresh stem bark of and fresh growth medium containing Calliandra surinamensis was collected from different concentrations (100, 50, 25, 12.5, the Ugbowo Campus of the University of 6.5, and 3.125 g/ml) of the tested Benin, Benin City, Nigeria in April, 2009. compound was added. After 2 days of Botanical identification and authentification incubation at 37C, with 5 % CO2, the was done by a Taxonomist Prof. M. Idu, growth medium was removed and MTT Faculty of life Sciences, University of Benin, reagent (0.1 mg/ml) was added. After Benin City, Nigeria. incubating at 37 ºC for 4 h, the MTT reagent Phytochemical screening: The crude was removed and DMSO (100 µl) was added plant powdered material was subjected to to each well and then shaken for another 15 phytochemical screening to test for the min. The absorbance was then determined Falodun et al. / Nig J. Biotech Vol. 21 (2010) 55 – 59 by an ELISA reader (Bio-Rad, USA) at a determined by spectroscopic techniques are wavelength of 492 nm. Control wells shown in figure 1. received only the media without the tested In Table 2, the results of the antioxidant compound. The conventional anticancer property of the compounds 1- 6 revealed drug, cisplatin, was used as a positive that compounds 1, 2 and 4 did not show control in this study. The inhibition of antioxidant activity against DPPH radical at cellular growth by the tested sample was concentrations of 10, 25 and 50 µg/ml. The calculated as the percent inhibitory activity compounds therefore lacked DPPH and expressed as the IC50 value antioxidant property. There was high (concentration of the tested sample to absorbance and antioxidant activity of 5 and inhibit 50 % growth of the cells). 6 using DPPH radical as control. Flavonoids Quantitative test: The DPPH radical have been shown to possess antioxidant scavenging of compounds from Calliandra activity (Zhao et al. 2007). The class of surinamensis extract (CSE) activity was compounds called xanthones is potential measured by the method of Blois with minor anti radical agents in medicinal plants. modifications (Wang et al. 2007). The The result of the anticancer activity compounds were each dissolved in 10 ml of presented in Table 3 showed that the distilled water to a final concentration of 100 compounds (except compound 5) were not g/ml. Two milliliters of 0.2 mM DPPH in active against Human lung cancer cell line methanol was added to 1 ml of the test A549 at the concentration investigated. compound solution. The absorbance was Compound 5 gave an IC50 of 106.35. measured at 517 nm after 20 min of Compounds 1 and 2 did not exhibit any incubation at 25oC. Quercetin was
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