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Secondary Metabolites, Antioxidant, and Antiproliferative Activities of Dioscorea Bulbifera Leaf Collected from Endau Rompin, Johor, Malaysia

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Secondary Metabolites, Antioxidant, and Antiproliferative Activities of Dioscorea Bulbifera Leaf Collected from Endau Rompin, Johor, Malaysia Hindawi Evidence-Based Complementary and Alternative Medicine Volume 2021, Article ID 8826986, 10 pages https://doi.org/10.1155/2021/8826986 Research Article Secondary Metabolites, Antioxidant, and Antiproliferative Activities of Dioscorea bulbifera Leaf Collected from Endau Rompin, Johor, Malaysia Muhammad Murtala Mainasara ,1,2 Mohd Fadzelly Abu Bakar ,1 Abdah Md Akim ,3 Alona C Linatoc ,1 Fazleen Izzany Abu Bakar ,1 and Yazan K. H. Ranneh 1 1Faculty of Applied Sciences & Technology, Universiti Tun Hussein Onn Malaysia (UTHM), Hab Pendidikan Tinggi Pagoh, KM1, Jalan Panchor, Muar 84600, Johor, Malaysia 2Faculty of Science, Department of Biological Sciences, Usmanu Danfodiyo University Sokoto, PMB 1046, Sokoto, Nigeria 3Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, Seri Kembangan 43400, Selangor, Malaysia Correspondence should be addressed to Mohd Fadzelly Abu Bakar; [email protected] Received 11 August 2020; Revised 26 October 2020; Accepted 14 December 2020; Published 11 January 2021 Academic Editor: Amin Tamadon Copyright © 2021 Muhammad Murtala Mainasara et al. +is is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Breast cancer is among the most commonly diagnosed cancer and the leading cause of cancer-related death among women globally. Malaysia is a country that is rich in medicinal plant species. Hence, this research aims to explore the secondary metabolites, antioxidant, and antiproliferative activities of Dioscorea bulbifera leaf collected from Endau Rompin, Johor, Malaysia. Antioxidant activity was assessed using 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric reducing antioxidant power (FRAP), and 2,2′-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) assays, while the cytotoxicity of D. bulbifera on MDA-MB-231 and MCF-7 breast cancer cell lines was tested using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Cell cycle analysis and apoptosis were assessed using flow cytometry analysis. Phytochemical profiling was conducted using gas chromatography-mass spectrometry (GC-MS). Results showed that methanol extract had the highest antioxidant activity in DPPH, FRAP, and ABTS assays, followed by ethyl acetate and hexane extracts. D. bulbifera tested against MDA-MB-231 and MCF-7 cell lines showed a pronounced cytotoxic effect with IC50 values of 8.96 μg/mL, 6.88 μg/mL, and 3.27 μg/mL in MCF-7 and 14.29 μg/mL, 11.86 μg/mL, and 7.23 μg/mL in MDA-MB-231, respectively. Cell cycle analysis also indicated that D. bulbifera prompted apoptosis at various stages, and a significant decrease in viable cells was detected within 24 h and substantially improved after 48 h and 72 h of treatment. Phytochemical profiling of methanol extract revealed the presence of 39 metabolites such as acetic acid, n-hexadecanoic acid, acetin, hexadecanoate, 7-tetradecenal, phytol, octadecanoic acid, cholesterol, palmitic acid, and linolenate. Hence, these findings concluded that D. bulbifera extract has promising anticancer and natural antioxidant agents. However, further study is needed to isolate the bioactive compounds and validate the effectiveness of this extract in the In in vivo model. 1. Introduction about 25% of all new identified cancer case is breast cancer, which accounts for 15% of deaths in women each year [2, 3]. Millions of mortalities were reported globally as a result of Breast cancer is a commonly diagnosed cancer in cancer, and the numbers of new cases are expected to in- Malaysia, having a standardized age rate of 47.4 per 100,000 crease in the future [1]. Breast tumour is the second highest [4, 5]. +e National Cancer Registry of Malaysia (NCR) cancer recorded worldwide and the most common cause of records that 21,773 Malaysians who are perceived with cancer death among women. It has been ascertained that cancer and approximations show that unregistered cases 2 Evidence-Based Complementary and Alternative Medicine have reached about 10,000 cases every year. It is likely that by of Research for Sustainable Uses of Natural Resources (CoR- 75 years old, ¼ of Malaysians will be diagnosed with cancer SUNR), Faculty of Applied Sciences and Technology, Uni- [6]. +erefore, discovering new drugs with minimal toxicity versiti Tun Hussein Onn Malaysia. Fresh sample was cleaned is a broad scientific challenge, and there is an immediate using tap water to eliminate impurities or soil debris. +e need to search for a new agent to mitigate the menace by sample was crushed to a fine powder with an electric blender using alternative evidence-based herbal medicines. after shade drying at room temperature and kept in a zip lock Dioscorea bulbifera (Figure 1) is a member of the family bag in a freezer (−20°C) for further analysis [8, 9]. Dioscoreaceae, categorised in the order Dioscoreales and genus Dioscorea. It is usually referred to as air potato, air yam, or bulbil-bearing yam. It is a prolific climbing plant 2.2. Sample Extraction. Methanol, ethyl acetate, and hexane indigenous to Southern Asia and West Africa, mostly solvents were used for the extraction of plant samples by found along forest edges. +e direction of circular twin- successive maceration methods as described by Bhunu et al. ning had been reported in D. bulbifera and can grow to a [10] with some modifications. +e mixture was filtered using height of 12 m. It has shiny green leaves that are alternate a vacuum filter and then evaporated using a rotary evapo- with a long leafstalk. Resembling true yam leaves, rator where the yield of extracts was found; hexane (6.22%), D. bulbifera has an annual vegetative cycle. Hence, this ethyl acetate (8.63%), and methanol (20.6%) were used. +e study was directed toward discovering secondary metab- extract was later used to identify its phytochemicals, anti- olites, antioxidant, and antiproliferative effects of oxidant, and antiproliferative effects. D. bulbifera leaf extract from Kampung Peta, Endau Rompin, Mersing, Johor, Malaysia. 2.3. Determination of Antioxidant Activities 2. Materials and Methods 2.3.1. 2,2-Diphenyl-1-picrylhydrazyl (DPPH) Assay. 2.1. Sample Collection and Preparation. D. bulbifera fresh DPPH scavenging activity of the sample was determined leaves were collected at Kampung Peta, Johor, Malaysia, on according to Bakar et al. [11] and Hassan and Bakar [12]. 5th May 2017 with the permission of Perbadanan Taman Briefly, a methanol solution of DPPH (0.3 mM) was added Negara Johor (PTJN) and with full observation of rules and into the samples (2.5 mL). +e extract was kept in a light regulations on collection practices of medicinal plants as laid protected place at room temperature for 30 min after mixing down by the World Health Organisation (WHO) collection vigorously. +e absorbance was read at 518 nm. +e fol- of plant materials [7]. Identification was made, and the lowing equation was used for calculating the scavenging or voucher specimen was deposited in the Herbarium of Centre antioxidant activity (AA): Absorbance (sample) − absorbance (empty sample) AA(%) � × 100: (1) absorbance (control) 2.3.2. Ferric Reducing Antioxidant Power (FRAP) Assay. preformed radical monocation of ABTS. +e solution was FRAP assay was conducted according to Dusuki et al. [13] kept in the dark and left to stand for 15 h at room tem- with some modifications. +e FRAP mixture was formed by perature. To obtain the units of absorbance at 734 nm, the mixing 300 mM acetate buffer with 3.6 pH, 2.5 mL of 10 mM solution was diluted with either methanol, ethyl acetate, or TPTZ solution in 40 mM HCl, and 0.25 mL of 20 mM FeCl3, hexane; aliquot for every sample (200 µL) was added to and the substances were tested in 0.1 mL methanol, ethyl 2000 µL of ABTS. Using a spectrometer, the absorbance was acetate, and hexane or methanol. Plain reading was recorded monitored for 5 min at 734 nm. Blank solvents were run at 593 nm using a test tube with extra 3 mL of FRAP reagent. appropriately in every assay. +e proportion of inhibition Plant extract (100 µL) and distilled water (300 µL) were was calculated against control and matched to a vitamin C supplemented to the test tube. Using a spectrometer, after standard curve (10–100 mm), and the percentage was used to 30 min of incubation, absorbance was read at 593 nm. Ferric express the radical scavenging activities. reducing capacity in 1 g of dried sample (mM/g) was measured as mM for the final result. All tests were run in triplicate. 2.4. Anticancer Activities 2.4.1. Cell Culture Condition. MCF-7 and MDA-MB-231 2.3.3. 2,2′-Azino-bis-3-ethylbenzthiazoline-6-sulphonic Acid breast cancer cell lines were purchased from American Type (ABTS) Assay. ABTS assay was carried in line with Re et al. Culture Collection (ATCC). +e cells were maintained in [14] with some modifications. Reacting of ABTS solution Roswell Park Memorial Institute (RPMI) 1640 media sup- (7 mM) with 2.45 mM of potassium persulfate produced the plemented with L-glutamine, 1% penicillin-streptomycin, Evidence-Based Complementary and Alternative Medicine 3 Figure 1: D. bulbifera plant (leaves). and 10% fetal bovine serum in a humidified atmosphere of incubated at −20°C for 2 h. +en, the ethanol was expelled ° 5% carbon dioxide (CO2) at 37 C. from the sample after centrifugation. +e cells that have been rinsed twice using cool PBS previously were stained with propidium iodide (500 μL of 10 μg/mL) and RNase (100 μg/ 2.4.2. Extracts Preparation. To solubilise the plant extract, mL) at room temperature for about 30 min. Dispersion of dimethyl sulphoxide (DMSO) at a concentration of 10 mM the cell cycle was analysed using flow cytometry.
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