insecticidal properties of Jatropha species and wild Ricinus communis L. found in Mauritius

Transcription

1 S384 Asin Pc J Trop Med 214; 7(Suppl 1): S384-S39 Contents lists vilble t ScienceDirect Asin Pcific Journl of Tropicl Medicine journl homepge: Document heding doi: 1.116/S (14) A comprison of polyphenolic content, ntioxidnt ctivity nd insecticidl properties of Jtroph species nd wild Ricinus communis L. found in Muritius Sillm Rmpdrth, Dneshwr Puchoo *, Vijynti Ml Rnghoo-Snmukhiy Deprtment of Agriculture & Food Science Fculty of Agriculture University of Muritius, Réduit, Muritius PEER REVIEW Article history: Received 6 Aug 214 Received in revised form 18 Aug 214 Accepted 2 Sep 214 Avilble online 28 Sep 214 Keywords: Jtroph curcs Ricinus communis Antimicrobil Phytochemicl Insecticidl ABSTRACT Objective: To evlute the phytochemicl, ntimicrobil nd insecticidl properties of different Jtroph species nd Ricinus communis (cstor) (R. communis) growing in Muritius. Methods: Qulittive nd quntittive methods were used for the determintion of the presence of phytochemicls in the crude plnts solvent extrcts. The ntimicrobil sensitivity (disc diffusion method) nd Antibcteril ctivity (MIC-microdilution method) of 13 microorgnisms were studied. The insecticidl properties of the crude solvent extrcts ginst lrve of two insects, Bctrocer zont nd Bctrocer cucurbite (Dipter, Tephritide), which cuse importnt economic losses to locl fruits were lso demonstrted. Results: Ethyl cette ws proved to be good solvent for extrction. Jtroph multifid showed very interesting ctivity ginst Bcillus lgicol nd Stphylococcus epidermis. Both R. communis nd Jtroph multifid were proved to be very effective s biopesticide ginst different types of insect lrve. Conclusions: This study ws ble to demonstrte tht Jtroph species s well s R. communis growing in Muritius possess ntimicrobil, phytochemicl nd insecticidl properties. 1. Introduction Plnts nd their derivtives hve long been used s both drugs nd dietry supplements by mn. According to Sbndr[1], up to 5% of current phrmceuticl products were derived from plnts but none were used s ntimicrobils. Plnts re rich sources of secondry metbolites, such s tnnins, terpenoids, lkloids, nd flvonoids, which hve been found to hve in vitro ntimicrobil properties[2]. The development of microbil ntibiotic resistnce hs required globl serch for new ntimicrobils of preferbly plnt origin. Euphorbicee, the spurge fmily, is lrge fmily of flowering plnts with bout 3 gener nd round 7 5 species[3]. The spurges re herbs, shrubs or trees[4]. The genus Jtroph belongs to the Euphorbicee fmily, tribe Jonnesiee, which hs bout 17 species. The nme Jtroph is derived from the Greek word jtros (doctor) nd trophe (food)[5]. This fmily occurs *Corresponding uthor: Dneshwr Puchoo, Deprtment of Agriculture & Food Science Fculty of Agriculture University of Muritius, Reduit, Muritius. E-mil: sudeshp@uom.mu Tel: ; Fx: Foundtion Project: Supported by the University of Muritius (Grnt No. R 13) nd Sounit R nd Lockdev R. minly in the tropics. The species re from the Indo-Mlyn region, tropicl Americ, tropicl Afric nd some subtropicl countries like South Afric, Mdgscr nd Muritius[6]. Jtroph species re used in trditionl folklore medicine to cure vrious ilments in Afric, Asi nd Ltin Americ[7]. The usge of plnts s trditionl helth remedies is populr in Asi, Ltin Americ nd Afric. The islnd of Muritius is situted in the Indin Ocen 2 km (1 242 miles) off the southest cost of the Africn continent (2 17 S nd E). Due to its volcnic origin, ge, loction nd soil composition, Muritius is home to diversity of flor nd fun. Plnts of the Euphorbicee fmily re widely cultivted in Muritius nd hve been reported to possess trditionl ntiseptic properties ll round the world[5]. They contins wide rnge of phytochemicls to which their ntimicrobil ctivities re often ttributed[8]. Muritius, being multi-ethnic nd multi-culturl nd with ncestors of Africn, Europen nd Indin origin, trditionl medicine, hs profound root in the Muritin society. Jtroph species nd Ricinus communis L. (R. communis) re non-edible multipurpose shrubs belonging to the fmily of Euphorbicee. They re n uncultivted, non-food wildspecies which grow round the islnd in bckyrds nd in

2 Sillm Rmpdrth et l./asin Pc J Trop Med 214; 7(Suppl 1): S384-S39 S385 the wild. Different prts of the plnt re used loclly s ethno medicine. Mny studies hve been done to demonstrte the efficcy of Jtroph species ginst wide rry of bcteri nd fungi[9]. Severl studies hve lso reveled tht Jtroph curcs (J. curcs) hs nticncer nd ntitumor properties. Moreover, phytochemicl studies crried out on Jtroph hve reveled the presence of secondry metbolites. However, none of the studies hve compred leves of four different species of Jtroph using two different solvents for extrction nd testing their bility to hinder growth of bcteri nd fungus, their phytochemicl constituent nd their insecticidl properties on lrve of Bctrocer zont (B. zont) nd Bctrocer cucurbite (B. cucurbite) The lrve of these two fruit fly species re very dptble to different climtes nd hve the cpcity to survive on different host plnts such s pech, guv, mngo, citrus nd cucurbits. They cuse serious economic losses, either by direct dmge to fruit or by insecticide tretments[1]. 2. Mteril nd methods 2.1. Plnt mterils Mture lef smples (Figure 1) were collected from different loclities of Muritius in Curepipe (Centrl Plteu of Muritius), Nouvelle Frnce (south of Muritius) nd Grnd Bie (north of Muritius). A smll smple of ech plnt specimen ws sent to Muritius Herbrium, Réduit, for identifiction. The fresh plnt mterils were clened, wshed under running tp wter, ir-dried for 3-4 d, then were corsely pounded nd stored in tightly closed plstic continers t 4 C. b c d e Figure 1. Mture lef smples of the selective medicinl plnts. : Jtroph multifid L.; b: R. communis L.; c: J. curcs L.; d: Jtroph podgric Hook.; e: Jtroph integerrim Jck Chemicls nd mterils All the chemicls nd pre-coted silic gel 6 F 254 [used for thin lyer chromtogrphy (TLC)], were purchsed from Sigm Chemicls, the medi, stndrd ntibiotics from Hi-Medi ; nd Oxoid nd micropltes from Merck. The solvents used for extrction were of nlyticl regent grde Metbolite extrction The decoction method ws used to obtin the plnt crude extrcts. A totl of 2 g fresh leves were llowed to be mcerted in 4 ml of two different solvent systems: methnol nd ethyl cette for 48 h. The crude extrcts were filtered using Whtmn filter pper, pore size µm, llowed to concentrte in ventilted fume cupbord t room temperture nd then stored t 4 C in the drk bottles for further use Phytochemicl screening Qulittive tests The tests to detect the presence of flvonoids, lkloids, sponins, steroids, tnnins, coumrins nd phenols were crried out ccording to the method described by Seth nd Srin[11]. All screenings were performed bsed on series of test tube tests. The crude solvent extrcts were used to test phenols, nd lkloids were lso determined by TLC nd UV techniques Quntittive determintion of totl flvonoid nd phenol content The totl flvonoid content ws mesured by AlCl 3 (luminium chloride) colorimetric ssy. To 5 µl of extrct nd 2 µl of distilled wter, 5% of sodium nitrte (w/v) ws dded. After 5 min, 15 µl of 1% AlCl 3 (w/v) ws dded. A volume of 2 sodium hydroxide (1 mol/l) ws dded fter 1 min followed by 1 2 µl of distilled wter. The mixtures were vortexed nd incubted for 3 min nd the bsorbnce ws mesured t 51 nm. A quercetin stndrd curve ws mde (-2 µg/ml). Totl flvonoid content (TFC) ws determined s µg quercetin equivlents (QE)/g of fresh smple weight using formul (1). The totl phenolic content of plnt extrct ws determined using Folin-Cioclteu regent method. To 25 µl of Folin- Cioclteu regent, n liquot of 1 µl of extrct smple ws dded, followed by 3.5 ml of deionised wter. After 3 min, 1 ml of 2% sodium crbonte ws dded. The mixture ws vortexed nd incubted t 4 C for 4 min. It ws llowed to cool in drk cupbord nd bsorbnce of the rection mixtures ws mesured t 685 nm. A gllic cid stndrd curve ws mde (-3 µg/ml). Totl phenolic content (TPC) ws determined s µg gllic cid equivlents (GAE)/g of fresh weight (FW) using formul (1). The bsorbnce for both rection mixes ws mesured by Jenwy spectrophotometer 735-UV-Visible. Triplicte redings were tken for ech smple. TPC (TFC)= R 伊 DF 伊 V 1 W 伊 V 2 (1) Where R=result obtined from the stndrd curve; DF=dilution fctor; V 1 =volume of stock solution; V 2 =volume of extrct used; W=weight of plnt mteril used Antimicrobil ssy Antimicrobil susceptibility test (disc diffusion method) The test microorgnisms included in this study were six

3 S386 Sillm Rmpdrth et l./asin Pc J Trop Med 214; 7(Suppl 1): S384-S39 Grm-positive [Bcillus lgicol Acc.13/5 (B. lgicol), Bcillus cereus ATCC (B. cereus), Listeri innocu ATCC 339 (L. innocu), Stphylococcus ureus ATCC (S. ureus), Stphylococcus epidermis ATCC (S. epidermis), Viridibcillus renosi strin LMG (V. renosi)], six Grm-negtive [(Escherichi coli ATCC (E. coli), E. coli 145:H28 Acc. No.CP627.1, Klebsiell oxycot ATCC 4386, Proteus mirbilis strin NCTC (P. mirbilis), Pseudomons eruginos ATCC (P. eruginos), Slmonell typhimurium ATCC 1428 (S. typhimurium)] nd one fungus [Cndid lbicns ATCC 123 (C. lbicns)]. Regulr sub-cultures of the test microorgnisms were crried out on sterile nutrient gr pltes. The bcteri were grown in nutrient broth t 37 C for h. All the inocul were stndrdized by dilution with sterile nutrient broth (.1 ml inocul in 9.9 ml nutrient broth) to n bsorbnce of.4-.6 t 6 nm. Avilble dehydrted Muller Hinton gr (MHA) medi [19 g/l (w/v)] ws prepred nd sterilized in n utoclve t 121 C nd 15 lbs pressure for 15 min nd poured into Petri dishes for the disc diffusion test. Circulr pper discs (6 mm dimeter,.9 mm thick) were lso sterilized. The sterile discs were septiclly plced over pltes of MHA (5 discs/plte) lredy seeded/spred with ech of test pthogens (1 µl/ plte) nd were impregnted with equl volume (5 µl) of ech crude plnt extrcts t fixed concentrtion of.2 g/ml (w/v). The pltes were incubted in n upright position t 37 C for h nd zone of inhibition ws mesured (in mm dimeter). The clinicl strins were lso tested for their sensitivity ginst positive control (stndrd ntibiotic tetrcycline, 3 µg) nd two negtive controls (solvents ethyl cette nd methnol). The sterile discs were lso tested on sterile plte of MHA without extrcts nd inoculum. Inhibition zones with dimeter greter thn 12 mm were considered s hving low ntimicrobil sensitivity, nd those lying between 12 mm nd 16 mm s modertely ctive nd bove 16 mm s highly ctive Antibcteril ctivity by microdilution-[(minimum inhibitory concentrtion (MIC) method] The seril microdilution method ws used to determine the MIC of frctionted plnt crude extrcts. Ech bcterium ws incubted in 1 ml of Mueller Hinton broth (MHB) overnight t 37 C. The overnight cultures were stndrdized by dilution with sterile MHB (.1 ml inoculum in 9.9 ml MHB) to n bsorbnce of.4-.6 t 6 nm. A totl of 1 µl tested smple for ech bcterium ws two-fold serilly diluted with 1 µl sterile distilled wter in sterile 96-well microplte. A two-fold of tetrcycline (3 µg/ml) ws lso used s positive control ginst ech bcterium. Methnol nd ethyl cette were used s negtive control nd 1 µl of bcteril suspension ws dded to ech well. The pltes were covered nd incubted t 37 C for 24 h, nd the lowest concentrtion of the extrct cusing complete inhibition of the bcteril growth ws tken s MIC. The results were compred with tht of control using ether methnol, ethyl cette nd tetrcycline (ntibiotic). The experiment ws performed in triplicte. Bcteril growth ws ssyed with the ddition of 4 µl of.2 mg/ml iodonitrotetrzolium violet to ech well fter incubtion t 37 C for 3 min. Bcteril growth in the wells ws indicted by the development of red colour nd the colourless wells indicted inhibition by tested extrcts Toxicity ssy Three-dy old lrve of B. zont nd B. cucurbite were collected from the Entomology Division (Ministry of Agro Industry nd Food Security, Réduit). The crude methnol nd ethyl cette extrcts of J. curcs, Jtroph integerrim (J. integerrim), Jtroph multifid (J. multifid) Jtroph podgric (J. podgric) nd R. communis were tested for their lrvicidl effect on two types of fly lrve t three different doses:.2,.4 nd.8 mg/l (w/v). The nturl food diet ws formulted using 6.% (w/v) of mize powder, sugr cne bgsse, wste brewer s yest nd whet brn, respectively, 11.% (w/v) sugr,.1% (w/v) sodium benzote,.1% (w/v) Nipgen,.8 % (w/v) hydrochloric cid nd 64.8% wter. A set of insect lrve ws llowed to grow nd recover in the fixed weight of food diet for two dys, eliminting the risk of nturl deth in Petri-dishes (1 mm 伊 15 mm). Then, the food diet ws spryed with 1 ml of the different concentrtion of the plnt extrcts. The experiment ws set in completely rndom mnner with three replictes per tretments (extrcts) nd controls (with nd without the solvent). The percentge of mortlity ws recorded t 24, 48 nd 72 h Sttisticl nlysis For the phytochemicl nd ntimicrobil ssys, ll dt were expressed s men 依 SD with One-wy ANOVA t 5% nd the lest significnt difference (LSD) test ws used to compre the differences between the mens. The mortlity ssy dt collected were subjected to logrithmic trnsformtion prior to nlysis nd probit nlysis ws used for clculting the lethl doses (LD). Minitb nd Microsoft Excel 21 were used for the sttisticl nlyses nd tbles nd grphs output. 3. Results 3.1. Phytochemicl screening Qulittive tests Phytochemicl nd frctions (TLC) screening initilly reveled the presence of lkloids, coumrins, flvonoids, steroids tnnins, nd phenols in most of the crude extrcts. Alkloids ppered in both ethyl cette nd crude methnol extrcts. Coumrins were observed only in J. curcs, J. podgric nd R. communis. Vrition in the Retention fctor (R f ) vlues ( ) obtined indicted the presence of different phenol nd flvonoid compounds nd for methnolic extrct of J. multifid (R f =.42) the presence of flvonoidglycoside compound ws confirmed (Tble 1).

4 Sillm Rmpdrth et l./asin Pc J Trop Med 214; 7(Suppl 1): S384-S39 S387 Tble 1 Preliminry screening of phytochemicls in the two solvent crude extrcts of Jtroph species nd R. communis using test tubes nd TLC methods. Plnts Solvents Phytochemicl Alkloids Coumrins Flvnoids (R f vlue) Steroids Tnnins Phenols (R f vlue) J. curcs Ethyl cette (.86) Methnol (.87) (.7) J. integerrim Ethyl cette (.73) (.62) Methnol (.74) J. podgric Ethyl cette (.51) Methnol (.55) (.75) J. multifid Ethyl cette (.74) Methnol (.42) (.72) R. communis Ethyl cette (.68) (.69) Methnol (.69) : Presence; --: Absence Quntifiction of TPC nd TFC The TPC nd TFC of the two solvent crude extrcts showed significnt differences (P<.5) for the different medicinl plnts (Figures 2 nd 3). The quntittive nlysis of the phytochemicls of vrious crude extrcts reveled tht the TPC ws higher compred to the flvonoid content nd the ethyl cette crude extrcts of mture leves gve better yield of phenol nd flvonoid. The ethyl extrct of R. communis leves contined the highest mount of TPC [( 依 19.2) µg GAE/g FW] followed by J. integerrim [(48. 依 34.) µg GAE/g]. Phenol ws obtined from both solvents extrcts of J. multifid only. The TFC for the ethyl extrct of leves of J. integerrim, J. podgric nd R. communis were (1.14 依.59) µg QE/g, (7.8 依.89) µg QE/g nd (7.61 依 1.88) µg QE/g, respectively, wheres the methnolic extrcts of J. integerrim nd J. multifid hd vlues of (11.31 依.89) µg QE/g nd (7.7 依.19) µg QE/g respectively. Men TPC (ug GAE/g FW) b b bc d de 3.2. Antimicrobil ssy Antibcteril nd ntifungl susceptibility test Most of the ten crude extrcts of the five plnt species hd significnt vrying degrees of ntibcteril nd ntifungl potentil (P<.5) (Tble 2 nd Figure 4). Tble 2 Summry of the ntimicrobil sensitivity of the different crude lef extrcts ginst bcteri. Antimicrobil sensitivity Micrognisms Respective extrct Ethyl Acette Methnol Moderte (12 mm<inhibition zone B. lgicol J. curcs J. integerrim R. communis E. coli 145:H28 R. communis J. curcs <16 mm) J. podgric R. communis E. coli J. podgric R. communis R. communis L. innocu R. communis P. mirbilis R. communis S. typhimurium J. curcs S. ureus J. curcs J. integerrim S. epidermis J. podgric High (>16 mm) B. lgicol J. multifid S. epidermis J. multifid J. curs J. integerrim J. podgric J. multifid R. communis Medicinl plnts Ethyl cete Methnol Figure 2. Quntifiction of the TPC in the different crude solvent plnt extrcts (n=5). Men TFC (ug QE/g FW) e f b Figure 3. Quntifiction of the TFC in the different crude solvent plnt extrcts (n=5). cd J. curs J. integerrim J. podgric J. multifid R. communis Medicinl plnts Ethyl cete Methnol d cd cd g b c d Figure 4. Zone of inhibitions (mm) of some test microorgnisms. : E. coli 145:H28 Acc. No. CP627.1; b: V. renosi strin LMG 22166; c: S. ureus; d: C. lbicns. The ntibcteril ctivity of ll extrcts depends lrgely upon the type of solvent used for extrction nd the bcteril strins tested in the susceptibility ssy. The ethyl cette extrcts exhibited promising ntibcteril ctivities ginst Grmpositive test microorgnisms compred to the methnolic extrcts. The extrct from leves of J. multifid showed the highest ntibcteril ctivity with men zone of inhibition of (17.2 依 1.79) mm dimeter ginst S. epidermis ATCC 12228

5 S388 Sillm Rmpdrth et l./asin Pc J Trop Med 214; 7(Suppl 1): S384-S39 Tble 3 Men zone of inhibition of solvent crude extrcts from selected medicinl locl plnts ginst Grm positive nd negtive microorgnisms. Microorgnisms J. curcs J. integerrim J. podgric Hook. J. multifid L. R.communis L. Negtive control Positive Ethyl celtte Methnol Ethyl celtte Methnol Ethyl celtte Methnol Ethyl celtte Methnol Ethyl celtte Methnol Ethyl celtte Methnol Grm-positive B. lgicol Acc.13/ 依 依 依 1.82 b 1.2 依 依 依. d 16. 依 1. b 9.2 依.84 b 1.4 依.55 b 12.8 依 依 依 依 1.48 B. cereus ATCC 依. d 6.. d 12. 依. c 8. 依. 1. 依. b 9. 依. c 6. 依. d 6. 依. c 9. 依. c 8. 依. b 6. 依. 6. 依. 2. 依. L. innocu ATCC 依.55 b 8.6 依.55 b 11. 依.71 c 1.6 依 依 2.7 c 1.8 依.85 c 12. 依 3.81 c 1.8 依 依 依 依. 1. 依. 19. 依. S. ureus ATCC 依 依 依 依.89 b 6. 依. e 6. 依. e 11.4 依 1.14 c 9.4 依 1.34 b 6. 依. d 6. 依. e 12. 依 依 依 3.78 S. epidermis ATCC 依.84 c 6.4 依.89 c 6. 依. d 6. 依. c 6.2 依.84 d 15.4 依 依 依 依. c 8.4 依.89 c 11. 依 依 依 2.41 V. renosi strin LMG 依.89 b 1.4 依 1.34 b 11.6 依.89 c 9.8 依.84 c 11.8 依 依.45 b 1.4 依.55 c 8.8 依.45 b 9. 依.71 c 11.4 依.55 b 1. 依. 9. 依 依.45 Grm-negtive E. coli ATCC 依 依 1.41 b 9.6 依.55 b 6.8 依.45 c 12.2 依 依 依.89 b 9.8 依.84 b 13. 依 依 依 依 依 2.39 E. coli 145:H28 Acc. 11. 依 1. b 14.8 依 依.84 b 6. 依. d 11. 依 依 依. c 11.2 依 依 依 依 依 依. No.CP627.1 K. oxytoc ATCC 依. b 9. 依. c 11. 依. 8. 依. b 1. 依. b 8. 依. c 12. 依. 6. 依. e 11. 依. b 1. 依. c 6. 依. 6. 依. 19. 依. P. mirbilis strin NCTC 依. b 8. 依. d 6. 依. c 12. 依. 11. 依. 6. 依. d 12. 依. 9. 依. c 13. 依. 11. 依. b 6. 依. 9. 依. 3. 依. P. eruginos ATCC 依. b 12. 依. b 11. 依. 9. 依. b 8. 依. c 1. 依. b 12. 依. 8. 依. d 6. 依. c 11. 依. b 6. 依. 6. 依. 24. 依. S. typhimurium ATCC 依 依. b 9.8 依.84 b 8.6 依.55 b 8.4 依.55 c 8. 依. c 6. 依. c 9.6 依.55 b 6. 依. c 6. 依. d 9. 依. 1.2 依 依. Dts re expressed s men 依 SD in mm dimeter; Mens tht do not shre letter re significntly different (column). control nd (16. 依 1.) mm dimeter for B. lgicol Acc.13/5 followed by J. integerrim nd J. curcs with men zone of inhibition of (15.6 依 1.52) mm for S. ureus ATCC nd (15.2 依.84) mm for B. lgicol Acc.13/5, respectively. Moreover, methnolic extrct of J. curcs leves hd men inhibition zone of (14.8 依.45) mm for Grm-negtive E. coli 145:H28 Acc. No.CP627.1 (Figure 4 nd Tble 3). P. mirbilis nd E. coli ATCC were both eqully susceptible to methnol extrct of R. communis. The highest mens for the ntifungl effect of C. lbicns (Figures 4 nd 5) were obtined for both extrcts of J. multifid [(14.4 依.1) mm nd (11.8 依.45) mm dimeter], followed by the ethyl cette extrcts of R. communis [(1.4 依 1.34) mm dimeter] nd J. curcs. The susceptibility test lso showed tht the test microorgnisms were more sensitive to the crude plnt extrcts thn the stndrd ntibiotic, tetrcycline Antimicrobil ctivity (MIC) The ntimicrobil ctivity of the ethyl cette crude extrcts seemed to be more effective thn the methnolic crude extrcts for inhibiting microorgnism ctivity of both Grm-positive nd Grm-negtive strins (Tble 4). The most significnt ntimicrobil ctivity which ws lower thn the stndrd ntibiotic tetrcycline, ws noted for methnolic extrcts of J. curcs (2.7 µg/l) ginst S. ureus ATCC nd the minimum concentrtion of 3.6 µg/l for R. communis ethyl cette crude extrcts ws found ginst L. innocu ATCC 339, E. coli ATCC nd Klebsiell oxytoc ATCC 4386 (K. oxytoc). Most of the crude solvent extrcts hd n ntifungl potentil for C. lbicns ATCC 123 (Tble 4). The crude solvent extrcts hd significnt the ntimicrobil ctivity ginst the following strins: L. innocu ATCC 339, S. ureus ATCC 29213, S. epidermis ATCC 12228, V. renosi strin LMG 22166, K. oxytoc ATCC 4386, S. typhimurium ATCC 1428 nd C. lbicns ATCC 123 compred to the positive control, tetrcycline Crude solvent extrcts: toxicity biossy ginst Bctrocer species The effect of the different concentrtions (LD) of the crude extrcts vried significntly on the two Dipter lrve. The crude ethyl cette extrct of J. multifid L. demonstrted greter efficiency for lrvl control of B. zont with LD 9 less thn.1 g/l (Tble 5). B. cucurbite lrve were more susceptible to the methnolic extrcts of J. curcs L. (LD 5 =.18 g/l nd LD 9 =.97 g/l) nd R. communis L. (LD 5 =.22 g/l nd LD 9 =1.12 g/l) (Tble 5). Tble 4 MIC (µg/l) of the solvent crude extrcts from selected medicinl locl plnts ginst Grm-positive nd Grn-negtive microorgnisms. Plnts Solvents Grm-positive Grm-negtive Fungus J. curcs L. Methnol Ethyl cette J. integerrim Jck. Methnol Ethyl cette J. multifid L. Methnol Ethyl cette J. podgric Hook. Methnol Ethyl cette R. communis L. Methnol Ethyl cette Tetrcycline (3 µg) : B. lgicol Acc. No. 13/5; 2: B. cereus ATCC 11778; 3: L. innocu ATCC 339; 4: S. ureus ATCC 29213; 5: S. epidermis ATCC 12228; 6: V. renosi strin LMG 22166; 7: E. coli ATCC 25922; 8: E. coli 145:H28 Acc. No.CP627.1; 9: K. oxytoc ATCC 4386; 1: P. mirbilis strin NCTC 11938; 11: P. eruginos ATCC 27853; 12: S. typhimurium ATCC 1428; 13: C. lbicns ATCC 123.

6 Sillm Rmpdrth et l./asin Pc J Trop Med 214; 7(Suppl 1): S384-S39 S389 Men zone of inhibition (mm) bc de ef f b c d e e e A B C D E F G H I G K L M Extrcts Figure 5. Men inhibition ctivity (mm dimeter) of loclly plnt extrcts ginst C. lbicns ATCC 123. EA: Ethyl cette; ME: Methnol; A: J. curcs EA; B: J. curcs ME; C: J. integerrim EA; D: J. integerrim ME; E: J. podgric EA; F: J. podgric ME; G: J. multifid EA; H: J. multifid ME; I: R. communis EA; J: R. communis ME; K: Negtive control-ea; L: Negtive control-me; M: Positive control (tetrcycline). One-wy ANOVA with LSD t 5%, n=5; Mens tht do not shre letter re significntly different. Tble 5 Lethl dose (LD) of the different crude solvent extrcts (g/l) ginst Dipter species. Plnts Solvents B. cucurbite B. zont LD 5 LD 9 LD 5 LD 9 J. curcs Methnol Ethyl cette J. integerrim Methnol Ethyl cette J. multifid Methnol Ethyl cette <.1 <.1 J. podgric Methnol Ethyl cette R. communis Methnol Ethyl cette No. of lrve nd tretment ws 1, nd experiments were repeted 3 times (Probit nlysis); --: No deth observed; LD 5 nd LD 9 : Dose level t which 5% nd 9% lrve die. The percentge of mortlity hs incresed with time for the different concentrtion of the crude extrcts (Figures 6-8). About 3.3% to 1.% lrve were killed fter 24 h. The insecticidl effect on B. zont ws 1.% fter 24 h t the dose of.8 g/l of crude solvent extrcts of J. curcs, J. multifid nd J. podgric. One hundred percent mortlity ws lso noted on B. cucurbite of J. multifid crude methnolic extrct t dose of.4 nd.8 g/l. At 48 nd 72 h, most of the Dipter lrve hd died. The percentge of mortlity ws rnged from 13.3% to 1.% for Dipter lrve both fter 48 nd 72 h (Figures 7 nd 8). Mortlity (%) B. cucurbite Dose (g/l) B. zont J. curcs EA J. curcs ME J. integerrim EA J. integerrim ME J. multifid EA J. multifid ME J. podgric EA J. podgric ME R. communis EA R. communis ME Figure 6. Mortlity (%) of both lrves for the different concentrtions of crude solvent extrcts fter 24 h. Men 依 SD with LSD t 5%, n=1. For ll tretments with controls with nd without solvents, deth of the lrve were not observed. f g Mortlity (%) B. cucurbite Dose (g/l) B. zont J. curcs EA J. curcs ME J. integerrim EA J. integerrim ME J. multifid EA J. multifid ME J. podgric EA J. podgric ME R. communis EA R. communis ME Figure 7. Mortlity (%) of both lrves for the different concentrtions of crude solvent extrcts fter 48 h. Men 依 SD with LSD t 5%, n=1. For tretments with controls with nd without solvents, deth of the lrve were not observed. Mortlity (%) J. curcs EA J. curcs ME J. integerrim EA J. integerrim ME J. multifid EA J. multifid ME J. podgric EA J. podgric ME R. communis EA R. communis ME Figure 8. Mortlity (%) of both lrves for the different concentrtions of crude solvent extrcts fter 72 h. B. cucurbite Dose (g/l) B. zont Men 依 SD with LSD t 5%, n=1. For tretments with controls with nd without solvents, deth of the lrve were not observed. 4. Discussion Both R. communis nd Jtroph species re used in locl trditionl medicine, for exmple, cstor oil is used s purgtive. Crude extrcts of the leves from the different species of Jtroph present on the islnd of Muritius were never studied previously for the presence of phytochemicl compounds, ntimicrobil/ntifungl properties or their bility to ct s nturl insecticides. The phytochemicl screenings were very dependent on the type of solvent used. In J. curcs nd R. communis, coumrins were present only in the ethyl cette extrct nd in the methnolic extrct of J. podgric. Flvonoids, on the other hnd, present in both ethyl cette extrct nd methnol. R f t different distnces on the TLC pltes nd the computed R f vlues indicted the presence of different flvonoids nd phenolic compounds in the leves crude extrcts. Although some phytochemicl studies conducted before hd reported the presence of sponins in R. communis nd J. curcs, sponins were not detected in this study. Although the bsence of lkloids in J. curcs lef extrcts hd been reported[5], the present study confirmed the presence of lkloid in the lef extrcts. Plnts occurring in vrying hbitts,

7 S39 Sillm Rmpdrth et l./asin Pc J Trop Med 214; 7(Suppl 1): S384-S39 the gret mgnitude of vrition in the concentrtion nd composition of phytochemicl ingredients in different prts of these plnts cn be explined[1]. These compounds hve long been ssocited with medicine nd were reported s efficient therpeuticl substnces. They exert considerble ntimicrobil ctivity through different mechnisms[8]. Antimicrobil ctivity of the Jtroph species hve been widely reported, but not ll the lef mterils hve been studied. In J. multifid, lthough the root does not inhibit the growth of C. lbicns, this study found tht the leves of J. multifid do inhibit the growth of the fungus t n MIC of 25. µg/l when using methnol solvent nd 14.5 µg/l for ethyl cette solvent[4]. The crude methnolic extrct of the leves of J. curcs inhibited the growth of S. ureus, S. epidermidis, P. eruginos, E. coli, B. cereus ATCC 11778, B. subtilis nd Proteus vulgris s observed by Igbinos for the crude root extrcts. Therefore, it cn be sid tht the J. curcs leves hs the sme ntimicrobil properties s the stem brk[8]. Severl phytochemicls, in prticulr, the flvones hve the potentil to interct with the vertebrte oestrogen receptor[3]. The flvones, pigenin, orientin, vitexin, vicenin-2 nd biflvone were isolted from the leves of J. curcs. In this study, these flvones hve lso been proved to be toxic ginst the Dipter species used, hence inducing high mortlity rte. Interesting results were obtined during this study, showing the effect of solvent on phytochemicl extrction. Crude solvent extrcts ffected bcteril growth. Finlly the insecticidl properties of crude leves extrcts were tested on two economiclly importnt plnt pests in Muritius, thereby pving wy for more reserch on J. multifid nd R. communis. Conflict of interest sttement We declre tht we hve no conflict of interest. Acknowledgements The uthors wish to cknowledge the University of Muritius for funding this project (Grnt No. R 13) nd the Entomology Division of the Ministry of Agro Industry nd Food Security for providing ll the fcilities required. The support of Mrs. Bhookhun-Seeruttun D in the sttisticl nlyses s well s the finncil nd morl support of Sounit R nd Lockdev R is grtefully cknowledged. References [1] Sbndr CW, Ahmt N, Jfr FM, Shidin I. Medicinl property, phytochemistry nd phrmcology of severl Jtroph species (Euphorbicee): review. Phytochemistry 213; 85: [2] Jitendr J, Ashish K G. Ricinus communis Linn: phytophrmcologicl review. Int J Phrm Phrm Sci 212; 4(4): [3] Arekemse MO, Kyode RMO, Ajiboye AE. Antimicrobil ctivity nd phytochemicl nlysis of Jtroph curcs plnt ginst some selected microorgnisms. Int J Biol 211; 3(3): [4] Sete D, Suntornsuk W. Vrition of phorbol ester contents in Jtroph curcs from different provinces in Thilnd nd the ppliction of its seed cke for strter broiler diets. Am Eursin J Agric Environ Sci 21; 8(5): [5] Ahmed WA, Slimon J. Phorbol ester s toxic constituents of tropicl Jtroph curcs seed oil. Eur J Sci Res 29; 31(3): [6] Akinpelu DA, Aiyegoro OA, Anthony IO. The bioctive potentils of two medicinl plnts commonly used s folklore remedies mong some tribes in West Afric. Afr J Biotechnl 29; 8(8): [7] El Diwni G, El Rfie Sh, Hwsh S. Antioxidnt ctivity of extrcts obtined from residues of nodes leves stem nd root of Egyptin Jtroph curcs. Afr J Phrm Phrmcol 29; 3(11): [8] Igbinos OO, Igbinos EO, Aiyegoro OA. Antimicrobil ctivity nd phytochemicl screening of stem brk extrcts from Jtroph curcs Linn. Afr J Phrm Phrmcol 29; 3(2): [9] Ds B, Lxminryn K, Krishnih M, Srinivs Y, Rju TV. Multidione, novel diterpenoid from Jtroph multifid. Tetrhedron Lett 29; 5(34): [1] Kumr A, Shrm S. An evlution of multipurpose oil seed crop for industril uses (Jtroph curcs L.): review. Ind Crops Prod 28; 28(1): 1-1. [11] Seth R, Srin R. Anlysis of the phytochemicl content nd nti-microbil ctivity of Jtroph gossypifoli L. Arch Appl Sci Res 21; 2(5):