Journal of Medicinal Plants Research Vol. 6(16), pp. 3213-3216, 30 April, 2012 Available online at http://www.academicjournals.org/JMPR DOI: 10.5897/JMPR12.028 ISSN 1996-0875 ©2012 Academic Journals
Full Length Research Paper
Antimalarial, antiemetic and antidiabetic potential of Grewia aslatica L. leaves M. Zia-Ul-Haq1*, Shakir Ahmad Shahid2, Shafi Muhammed3, Mughal Qayum4, Inamullah Khan5 and Shakeel Ahmad6 1
Research Institute of Pharmaceutical Sciences, Department of Pharmacognosy, University of Karachi, Karachi-75270, Pakistan. 2 Department of Chemistry, University of Sargodha, Sargodha-40100, Pakistan. 3 Department of Pharmacy, University of Balochistan, Quetta, Pakistan. 4 Department of Pharmacy, Abdul Wali Khan University, Mardan 23200, Pakistan. 5 Department of Pharmacy, University of Peshawar, Peshawar-25120, Pakistan. 6 Department of Agronomy, Bahauddin Zakariya University Multan-60800, Pakistan. Accepted 14 February, 2012
Ethnopharmacological knowledge has firm traditional and conceptual base built on a tremendous observational data and theoretical structure but lack experimental base, and scientific evidence is needed to prove rationale of using this indigenous flora to preserve and prosper the cultural legacy. Researchers are blending traditional knowledge with experimental methodology for testing efficacy and safety of these herbal remedies. The current study has been designed to authenticate antimalarial, antiemetic and antidiabetic activities of methanolic extracts of Grewia asiatica L. leaves. The extract revealed significant antimalarial, antiemetic and antidiabetic activity. These results indicated the potential of G. asiatica leaves to further explore as natural source for new lead compounds against malaria, diabetes and emesis. Key words: Antimalarial, antiemetic, antidiabetic, Grewia asiatica L. leaves.
INTRODUCTION Grewia asiatica L. locally known as phalsa and dhaman is a large bushy shrub or small tree, and has height of 4 m or more. It is found throughout Punjab province of Pakistan and has medicinal as well as food uses. Its fruits are eaten fresh as dessert, are made into syrup, and extensively employed in the manufacture of soft drinks. The fruit is used in making jams, pies, squashes, chutneys etc. However, phalsa fruit has a short shelf life and is considered suitable only for local marketing (Anand, 1960; Panda, 2002). The fresh leaves are valued as fodder and are used by indigenous communities as antimalarial aid. All parts of plant are believed to possess anti-emetic properties. Its fruit is mordant, stomachic, laxative, and an aphrodisiac. Unripe fruits are used for treating fever, diarrhea, cardiac, respiratory and blood disorders, and relieves from inflammation. The fruits
*Corresponding author. E-mail:
[email protected].
assuage thirst and burning sensation, and help to remove dead foetus. Fruit extract possesses radioprotective potential. The leaves are applied on skin rashes and pustular eruptions due to their antibiotic potential, while the root is used for treating strangury, pus and clap (Morton, 1987; Pankhaj, 2004; Sharma and Sisoda, 2009; Ahmad, 2007). The plant is believed to possess antipyretic, antidiabetic, analgesic, antifertility, antibiotic and antimicrobial properties (Tripathi et al., 2010). The bark is used to treat rheumatism, while infusion of bark is used as demulcent, febrifuge and anti-diarrhoea. The bark also cures urinary problems and alleviates burning in vagina (Sisoda and Singh, 2009; Muhammad et al., 2006). Besides this, baskets are made from its shoots which are used to carry fruit and vegetables. Small tough articles like spades; golf sticks shafts, shoulder poles for carrying small loads, pestles, bows, billiards cues and shingles are made from its wood. The mucilaginous extract of the bark obtained after pounding in water is used to purify sugar cane juice during the preparation of
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Table 1. Enoyl-ACP reductase inhibitory (antimalaria) potential of Grewia asiatica.
Sample G. asiatica Standard
% Inhibition 69.12 ± 0.01 93.17 ± 0.02
‘gur’ or brown sugar. In Burma, the bark is used as a soap substitute while fiber extracted from the bark is made into rope (Sosef et al., 1998). Despite its so much medicinal and nutritional importance, very little work is reported on this plant. In this regard as part of our continuous studies on exploring the hidden potential of the indigenous flora of Pakistan (Nisar et al., 2010a, b, c, 2011; Zia-Ul-Haq et al., 2007a, b, 2008a, b, 2009, 2010a, b, 2011a, b, c, d, e) and evaluated anti-emetic, antidiabetic and antimalarial potential of methanolic extracts of G. asiatica L. leaves.
MATERIALS AND METHODS Plant material and preparation of crude extract G. asiatica L. leaves were obtained from Department of Agronomy, Bahauddin Zakariya University Multan, Pakistan and authenticated by Dr. Shakeel Ahmad, Assistant Professor of same department. Plant material was cleaned and soaked in methanol for 15 days with occasional shaking. It was filtered through a muslin cloth and then through a filter paper. The extracts so obtained were combined, filtered through filter paper under vacuum and concentrated under reduced pressure in a rotary evaporator (model Q-344B – Quimis, Brazil) using a warm water bath (model Q-214M2 - Quimis, Brazil) to obtain a thick gummy mass, which was further dried in a desiccator and stored in air-tight vial till further use. Antimalarial assay
Inhibition (%) = Where A is the control frequency of retching and B is the frequency of retching of the treated group. The results are given in Table 2. α-Amylase inhibitory activity The α-amylase inhibitory activity was determined by a slightly modified method proposed by Bernfeld (Bernfeld, 1955). α-amylase (20 µl; 0.05 U/μl) was premixed with sample (20 μl) and starch solution (250 μl; 2%) in sodium phosphate buffer (0.1 M; pH 6.9) was added as a substrate to start the reaction. The reaction was carried out at 37°C for 10 min and terminated by the addition of 200 μl of DNS reagent (1% 3,5-dinitrosalicylic acid; 12% sodium potassium tartrate in 0.4 M NaOH). The reaction mixture was heated for 15 min at 100°C, and then diluted with 5 ml of distilled water. α-Amylase activity was determined by measuring absorbance at 540 nm using the following equation:
Inhibition % 1
( AI AIB ) 100 ( AO AOB )
Ai is the A540 of sample reactive solution, A0 is the A540 of control reactive solution, AiB is the blank of sample and A0B is the blank of control. The results are given in Figure 1. α-Glucosidase inhibitory assay
Enoyl-ACP reductase was used as described earlier (Surolia and Surolia, 2001) utilizing crotonyl CoA as a substrate and measuring the decrease in absorbance at 340 nm using a UV-VIS spectrophotometer at 25°C in sodium phosphate buffer (50 mM; pH 7.5) containing NaCl (100 mM). 100 µl assay solution was prepared (Crotonyl CoA 150 µM, NADH 100 µM; PfENR 170 nM; 5% DMSO), that served as the positive control The sample was pipetted in 96well UV-plates, then a mixture containing (buffer + Enzyme + NADH) was added. This mixture was preincubated for 5 min before initiating the reaction by adding the substrate, Crotonyl CoA. The rate of decrease in the amount of NADH in each reaction well was converted to percentage of inhibition using SOFTmax PRO software (Molecular Devices, Sunnyvale, Calif.) and following formula:
Inhibition (%)
al., 2008). Four day old male chicks were divided into four groups of five chicks each and each chick was kept in a large beaker at 25°C for 20 min to stabilize. Group 1 served as the control and was treated with normal saline (5 ml/kg). Groups 2 and 3 were treated with extract (50 and 100 mg/kg) orally, while group 4 was treated with metoclopramide (50 mg/kg) intraperitoneally. After one hour, anhydrous copper sulphate (50 mg/kg) was administered orally to each chick and then number of retches (an emetic action without vomiting gastric material) was counted for 10 min. The antiemetic effect was assessed as the decrease in number of retches in the treated group. The inhibition (%) was calculated as follows:
Rate in the presence of sample 100 Rate of positive control
α-glucosidase was assessed according to a previously reported method (Matsui et al., 1996) with minor modifications. p-nitrophenyl α-D-lucopyranoside (1 ml; 3 mM) in sodium phosphate buffer (0.2 M; pH 6.8) was added as a substrate to the mixture of αglucosidase (50 μl; 0.15U/µl) and sample (50 μl) to start the reaction. The reaction was conducted at 37°C for 15 min and stopped by the addition of Na2CO3 (750 μl; 0.1 M). The αglucosidase activity was assessed by measuring the release of pnitrophenol from pNPG at 410 nm. Glucosidase activity inhibition was determined as percentage of control, as follows:
Where, Ai is absorbance of sample solution, and Ao is absorbance of control solution. Acarbose was used as a positive control. The results are given in Figure 1.
The results are given in Table 1. Antiemetic assay Antiemetic assay was performed as reported previously (Tijani et
RESULTS AND DISCUSSION Chemical analyses and biological and pharmacological
Zia-Ul-Haq et al.
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Table 2. Antiemetic activities of Grewia asiatica.
Drug/dose Control Metoclopramide (50 mg/kg) G. asiatica (50 mg/kg) G. asiatica (100 mg/kg)
No. of retches 50.04 ± 2.31 8.09b ± 1.65 30.45a ± 1.11 20.17b ± 2.76
% inhibition a 83.9 a 39.14 a 59.69
100
Inhibition %
80
60
40
20
0 alpha-glucosidase
100 µg/ml
alpha-amylase
250 µg/ml
500 µg/ml
Acarbose 0.1 µg/ml
Figure 1. α-glucosidase and α-amylase inhibitory (%) activities of Grewia asiatica.
assays provide objective evidences to validate traditional uses by indigenous communities. Research in drug discovery involves multifaceted approach combining botanical, phytochemical, biological and molecular techniques. So, current study is designed to screen a locally used plant for various tagged pharmacological bioactivities. Malaria is a major cause of childhood and adult illness around the globe. Resistance to the most common antimalarial drugs has spread to almost every part of the world. This prompted the urgent development of new compounds to treat malaria. Efforts are now directed towards obtaining drugs with different structural features, along with new strategies in malaria control and the recognition and validation of traditional medical practices (VanDyk et al., 2009). Our results indicate that G. asiatica L. leaves are a potential source of antimalarial drugs. The crude extract had maximum inhibition, that is, 69% inhibition (Table 1). The results are very promising and G.
asiatica L. leaves should be screened to identify the bioactive molecule that is responsible for such a higher antimalarial activity. Plant extracts have long been used for the ethnomedicinal treatment of diabetes in various systems of medicine and are accepted as an alternative for diabetic therapy because natural α-glucosidase inhibitors from plant sources offer an attractive strategy for the control of postprandial hyperglycemia (Subramanian et al., 2008). The results (Figure 1) indicated that G. asiatica L. leaves exhibited antidiabetic potential and this potential is dose dependent, that is, higher antidiabetic potential at higher concentrations and lower antidiabetic potential at low concentrations. Emesis and puking may be sign of many conditions like gestation, peptic ulcer, drug noxiousness, renal failure, and hepatitis, which if uncontrolled, may disturb the quality of life by dehydration, intense metabolic imbalances and nutrient diminution. Retching may occur after administration of cancer chemotherapeutic agents
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which may lead to rejection of curative anti-neoplastic therapy. G. asiatica leaves extract decreased significantly vomiting frequency. Metoclopramide, which provokes antiemetic activity by acceleration of GIT movement (Akita et al., 1998), however was more effective than extract. This antiemetic effect of extract may in part be due to its effect on the visceral afferent sympathetic fibre in the GIT and probably through agonistic effect at the inhibitory receptor sites such as dopamine and GABA in central nervous system (Tijani et al., 2008). Previously antiemetic effect of G. asiatica fruit extract has been observed on dog models (Yaqeen et al., 2008). Development of industries based on medicinal plants should be included as a priority area for translating lab findings to industrial scale to break confinement from academic pursuits. There is dire need to build up more and better organized inter-disciplinary studies among farmers, agricultural scientists, scientific research institutes and herbal industries of Pakistan to add value to medicinal plants from cultivation to marketing by processing, chemical analysis and bioactivity-guided fractionation to rationalize their folk uses. There is need to conserve medicinal germplasm and catalogue of all genetic material indigenous to Pakistan, and should be created and maintained to hold effective intellectual property rights (IPRs). REFERENCES Ahmad SS (2007). Medicinal wild plants from Lahore-Islamabad motorway (M-2). Pak. J. Bot., 39(2): 355-375. Akita Y, Yang Y, Kawai T, Kinoshita K, Koyama K, Takahashi K (1998). New assay method for surveying antiemetic compounds from natural sources. Nat. Prod. Sci., 4: 72-77. Anand JC (1960). Efficacy of sodium benzoate to control yeast fermentation in phalsa (Grewia asiatica L.) juice. Indian J. Hort., 17: 138-141. Bernfeld P (1955). Amylases, alpha and beta; in: Methods in Enzymology (Eds) S.P. Colowick and N.O. Kaplan. New York Acad. Press, 1: 149-158. Matsui T, Yoshimoto C, Osajima K, Oki T, Osajima Y (1996). In vitro survey of α-glucosidase inhibitory food components. Biosci. Biotechnol. Biochem., 60: 2019-2022. Morton J (1987). Phalsa. In: Fruits of warm climates. Miami, FL, USA. pp. 276-277. Muhammad IC, Khan MA, Hanif W (2006). Ethnoveterinary medicinal uses of plants from Samahni valley Distt. Bhimber, (Azad Kashmir) Pakistan. Asian J. Plant Sci., 5(2): 390-396. Nisar M, Qayum M, Shah MR, Kaleem WA, Ali I, Zia-Ul-Haq M (2010a). Antimicrobial screening of Impatiens bicolor Royle. Pak. J. Bot., 42(1): 523-526. Nisar M, Qayum M, Shah MR, Siddiqui HL, Kaleem WA, Zia-Ul-Haq M (2010b). Biological screening of Impatiens bicolor Royle. Pak. J. Bot., 42(3): 1903-1907. Nisar M, Kaleem WA, Qayum M, Hussain A, Zia-Ul-Haq M, Ali I, Choudhary MI (2010c). Biological screening of Zizyphus oxyphylla Edgew leaves. Pak. J. Bot., 42(6): 4063-4069. Nisar M, Kaleem WA, Qayum M, Marwat IK, Zia-Ul-Haq M, Ali I, Choudhary MI (2011). Biological screening of Zizyphus oxyphylla Edgew stem. Pak. J. Bot., 43: 311-317. Panda H (2002). Handbook on Ayurvedic Medicines with Formulae, Processes and Their Uses. National Institute of Industrial Research, Asia Pacific Business Press Inc., India.
Sharma KV, Sisodia R (2009).Evaluation of free radical scavenging activity and radioprotective efficacy of Grewia asiatica fruit. J. Rad. Prot., 29: 429-443. Sisodia R, Singh S (2009). Biochemical, behavioral and quantitative alterations in cerebellum of Swiss albino mice following irradiation and its modulation by Grewia asiatica. Int. J. Rad. Biol., 85(9): 785791. Sosef MSM, Hong LT, Prawirohatmodjo S (1998). PROSEA 5(3) Timber trees: lesser known species. Backhuys Publishers, Leiden, Netherlands. Subramanian R, Asmawi MZ, Amirin S (2008). In vitro α-glucosidase and α-amylase enzyme inhibitory effects of Andrographis paniculata extract and andrographolide. Acta Biochem. Pol., 55(2): 391-398. (Clarify). Surolia N, Surolia A (2001). Triclosan offers protection against blood stages of malaria by inhibiting enoyl-ACP reductase of Plasmodium falciparum. Nat. Med., 7: 167-173. Tijani AY, Okhale SE, Oga FE, Tags SZ, Salawu OA, Chindo BA (2008). Anti-emetic activity of Grewia lasiodiscus root extract and fractions. Afr. J. Biotechnol., 7(17): 3011-3016. VanDyk S, Griffiths S, VanZyl RL, Malan SF (2009). The importance of including toxicity assays when screening plant extracts for antimalarial activity. Afr. J. Biotechnol., 8: 5595-5601. Yaqeen Z, Sohail T, Rahman A, Saleem M, Rehman Z (2008). Evaluation of antiemetic activities of alcoholic extract of Grewia asiatica in experimental model dog. Pak. J. Sci. Ind. Res., 51: 212215. Zia-Ul-Haq M, Iqbal S, Ahmad S, Imran M, Niaz A, Bhanger MI (2007a). Nutritional and compositional study of desi chickpea (Cicer arietinum L.) cultivars grown in Punjab, Pakistan. Food Chem., 105: 13571363. Zia-Ul-Haq M, Ahmad M, Iqbal S, Ahmad S, Ali H (2007b). Characterization and compositional studies of oil from seeds of desi chickpea (Cicer arietinum L.) cultivars grown in Pakistan. J. Am. Oil Chem. Soc., 84: 1143-1148. Zia-Ul-Haq M, Iqbal S, Ahmad M (2008a). Characteristics of oil from seeds of 4 mungbean (Vigna radiate L. wilczek) cultivars grown in Pakistan. J. Am. Oil Chem. Soc., 85: 851-856. Zia-Ul-Haq M, Iqbal S, Ahmad S, Bhanger MI, Wiczkowski W, Amarowicz R (2008b). Antioxidant Potential of Desi Chickpea varieties commonly consumed in Pakistan. J. Food Lipid, 15: 26-342. Zia-Ul-Haq M, Ahmad S, Ahmad M, Iqbal S, Khawar KM (2009). Effects of cultivar and row spacing on tocopherol and sterol composition of chickpea (Cicer arietinum L) seed oil. Tarim Bilimleri Dergisi, 15: 2530. Zia-Ul-Haq M, Ahmad S, Chiavaro E, Mehjabeen, Ahmed S (2010a). Studies of oil from cowpea (Vigna unguiculata (l) walp.) cultivars commonly grown in Pakistan. Pak. J. Bot., 42(2): 214-220. Zia-Ul-Haq M, Ahmad M, Akhter M (2010b). Nematicidal activity of indigenous flora of Pakistan. Pak. J. Bot., 43: 1463-1466. Zia-Ul-Haq M, Ahmad S, Shad MA, Iqbal S, Qayum M, Ahmad A, Luthria DL, Amarowicz R (2011a). Compositional studies of some of lentil cultivars commonly consumed in Pakistan. Pak. J. Bot., 43(3): 15631567. Zia-Ul-Haq M, Ahmad M, Mehjabeen, Jehan N, Ahmad S, Qayum M, Marwat IK (2011b). Antimicrobial screening of selected flora of Pakistan. Arch. Biol. Sci., 63(3): 691-695. Zia-Ul-Haq M, Nisar M, Shah MR, Akhter M, Qayum M, Ahmad S, Shahid SA, Hasanuzzaman M (2011c).Toxicological screening of some selected legumes seed extracts. Leg. Res., 34(4): 242-250. Zia-Ul-Haq M, Ćavar S, Qayum M, Imran I, Feo V (2011d). Compositional studies: antioxidant and antidiabetic activities of Capparis decidua (Forsk.) Edgew. Int. J. Mol. Sci., 12(12): 88468861. Zia-Ul-Haq M, Ahmad S, Iqbal S, Luthria DL, Amarowicz R (2011e). Antioxidant potential of lentil cultivars. Oxid. Comm., 34: 819-831.
