SHORT COMMUNICATIONAntivectorial Activities of Cashew Nut ShellExtracts from Anacardium occidentale L.

Alain Laurens1w, Christophe Fourneau1, Reynald Hocquemiller1, Andre Cave1, Christian Bories2 andPhilippe M. Loiseau21 Laboratoire de Pharmacognosie URA 1843 CNRS2 Laboratoire de Parasitologie, Faculte de Pharmacie 92296 Chatenay-Malabry, France

Sodium salts of cashew nut shell extracts (CNSL) and anacardic acids isolated from Anacardium occidentaledemonstrate a potent antivectorial activity against Aedes aegypti larvae and Biomphalaria glabrata snails. Thestructureactivity relationship is discussed, particularly the hydrogenation of CNSL and anacardic acids whichlower dramatically these properties, showing the importance of the double bonds on the side chains of anacardicacids. 1997 by John Wiley & Sons, Ltd. Phytother. Res. 11, 145146, 1997

(No. of Figures: 0. No. of Tables: 1. No. of Refs: 10.)

Keywords: Anacardium occidentale; Anacardiaceae; CNSL; anacardic acids; larvicides; molluscicides.

INTRODUCTION

In the tropics, numerous human diseases are transmitted bymeans of blood-sucking mosquitoes (e.g. malaria, yellowfever, filariasis) or aquatic snails (e.g. schistosomiasis,fasciolasis). The control of the mosquito by destruction ofthe aquatic stages is a rapid and efficient means of reducingand eliminating the transmission of the disease. In the sameway, snail vector control is possible (WHO, 1985; Marstonet al., 1985). Larvicides of plant origin are currentlyreceiving considerable attention because of their relativelyharmless biodegradable properties. Since the 1920s morethan 2000 plants have been tested for insecticidal activity(Klocke, 1989) but bioassays with plant extracts againstmosquito larvae are rare. Moreover the use of tropical plantscultivated near the endemic areas should be of great interestand cheap for the antivectorial struggle in developingcountries.

In this field, Anacardium occidentale (Anacardiaceae), afruit tree growing widely in tropical and subtropical area iscultivated in Africa, South America and India for its kernel(cashew nut). Cashew nut shell liquid (CNSL) showed apotent molluscicidal activity (Sullivan et al., 1982; Kubo etal., 1986; Laurens et al., 1987) and various pharmacologicalactivities (Bhattacharya et al., 1987). The aim of this workis to study the larvicidal activity of the extract and todevelop a tensioactive extract of Anacardium occidentale.These antivectorial properties against aquatic pests, i.e. thelarvae of the mosquito Aedes aegypti, a yellow fever andfilariasis vector, are studied compared with the well knownmolluscicidal activity against the snail Biom-phalaria glabrata the intermediate host of intestinal schisto-somiasis. The presporal toxin of Bacillus thuringiensisisraeli was the reference compound against mosquito larvae(Nathan, 1993); Phytolacca dodecandra (Endod) spraydried extract and niclosamide were used as referencecompounds against snails (Goldsmith, 1991).

MATERIALS AND METHODS

Plant material. Cashew nuts of Anacardium occidentalewere collected in MBao area near Dakar (Senegal) in 1993.A voucher specimen was deposited in Jardin des PlantesUtiles herbarium, Faculte de Medecine et de Pharmacie,Dakar, Sngal.

Extracts. CNSL: Cashew nut shells were extracted with n-hexane in a Soxhlet apparatus. Solvent was removed underreduced pressure with a yield of 26%.

HPLC. Analysis of CNSL extract was performed using anODS Novapack 15 cm column. The mobile phase wasmethanol with acetic acid 4% (88:15) with detection at280 nm. Separation of anacardic acids: 15 g of extract waschromatographed on a silica gel column (450 g of silica) toisolate anacardic acids. Compounds are eluted by hexaneethyl acetateacetic acid (90:10:1).

Hydrogenation of anacardic acids and CNSL. 80 mg ofanacardic acids was dissolved in toluene and hydrogenatedby H2. The catalyst was Pd-carbon (8 mg). The solvent wasevaporated to dryness in vacuum and the residue obtainedwas constituted by saturated anacardic acid. CNSL wastreated in the same way.

Reference standards. Endod S type 44 spray dried extract ofberries of Phytolacca dodecandra from Ethiopia waspurchased from Dr Lugt. Support Group Endod, Borneo-straat 124, The Netherland 2585, La Hague. Niclosamidewas a gift from Rhone-Poulenc/Rorer; the bacterial toxinfrom Bacillus thuringiensis israeli IPS82 was kindlyprovided by Professor H. de Barjac, Institut Pasteur, Paris,France.

Biological assays. Larvicidal activities: The biological hometest consists of GKep Aedes aegypti laboratory reared strain.w Correspondence to: A. Laurens.

CCC 0951418X/97/02014502 1997 by John Wiley & Sons, Ltd. Accepted 15 July 1996

PHYTOTHERAPY RESEARCH, VOL. 11, 145146 (1997)

24 h old mosquito larvae were exposed to concentrations ofeach tested compound for 24 h at 26C. Twenty larvae wereexposed using a small glass cup in 4.5 mL of non-chlorinated water. Three test cups for each testedconcentration and control were used and three replicationswere run over a period of time to ensure sufficient geneticvariability in the colony larvae that were used. Aftercorrection for control mortality with Abbotts formula, theconcentration that produced 50% mortality was determinedby plotting the mortality points on log-probit paper.

Molluscicidal activity was monitored according to theWHO memorandum Molluscicide screening and Evalua-tion, 1965. Laboratory reared snails Biomphalariaglabrata, Puerto Rico strain, were used. Two 100 mLcontainers at each concentration with ten young maturesnails were used. The plant extracts or chemical compoundswere dissolved or suspended in dimethylsulphoxide anddiluted with water to solutions of known concentrations.Assays were carried out in aerated standard reference water(10% hardness). Third fold series were performed at 26C,12 h day-night periods. Death was measured after a 24 hincubation time period by the lack of heart rate. The lethalconcentration 50% was determined using probit/log10 dosepaper.

RESULTS AND DISCUSSION

The analytical study of CNSL by HPLC showed thatanacardic acids are the main components (73.7%) of CNSLwith 39.5% for the trien, 9.3% for the dien and 24.9% forthe monoen. In addition cardanols, 2-methylcardols andcardols were identified according to previous work (Shobhaet al., 1992).

Biological activities are shown in Table 1. CNSL andEndod molluscicidal activities were potent and quite similar(respectively 5.4 and 5.9 ppm), nevertheless CNSL demon-

strated a more potent larvicidal activity at 2.6 ppm.Moerover, chemical transformation of CNSL using sodiumhydroxide, into a tensioactive derivative enhanced theantivectorial properties against mosquito larvae and snailsby 20% (respectively 2.3 and 4.0 ppm).

After the hydrogenation of CNSL, these propertiesdropped dramatically for both vectors (18 ppm for larvaeand 34 ppm for snails). Saturated anacardic acids withLC50 >100 ppm demonstrating the importance of the dou-ble-bonds on the side chain for snails. Nevertheless, thehydrogenated CNSL remained toxic, but at a higher level(LC50 =34 ppm) probably because phenolic componentsother than anacardic acids are involved in the lethal process.These chemical products are mainly cardanols and cardols.

For mosquito larvae saturated anacardic acids at12.5 ppm were more active than the whole extract. Compar-atively niclosamide was only efficient against snails and theIPS 82 toxin was only efficient against larvae.

Therefore the CNSL sodium salt is a compound ofinterest for controlling mosquito larvae and aquatic snails intropical countries. This low-cost natural product could be aby-product of the agroalimentary industry. The CNSLsodium salt could be incorporated easily in washing powderor soap manufactured in developing countries. It coulddestroy and control mosquito larvae or snail pollution inwaste water, sewage or open main sewers.

REFERENCES

Bhattacharya, S. K., Mukhopadhyay, M., Mohan Rao, J. R.,Bagchi, A., and Ray, A. B. (1987). Pharmacological investiga-tion on sodium salt and acetyl derivative of anacardic acid.Phytother. Res. 1, 127134.

Goldsmith, M. F. (1991). Out of Africa comes the fruit of longresearch: possible self-reliant control of schistosomiasis.JAMA 265, 26502651.

Klocke, J. A. (1989). Plant compounds as sources and models ofinsect-control agents. In, Economic and Medicinal PlantResearch, ed. by H. Wagner, H. Hikino and N. R. Farnsworth),3, 103144. Academic Press, London.

Kubo, I., Komatsu, S., and Ochi, M. (1986). Molluscicides fromthe Cashew Anacardium occidentale and their large-scaleisolation. J. Agric. Food Chem. 34, 970973.

Laurens, A., Belot, J. and Delorme, C. (1987). Activite mollusci-

cide de lAnacardium occidentale L. (Anacardiaces). Ann.Pharm. Fr. 45, 471473.

Marston, A., and Hostettmann, K. (1985). Plant molluscicides.Phytochemistry 24, 639652.

Nathan, M. B. (1993). Critical review of Aedes aegypti controlprograms in the Caribbean and selected neighboring coun-tries. J. Am. Mosq. Control Assoc. 7, 6972.

Shobha, S. V., Krishnaswamy, P. R. and Ravindranath, B. (1992).Phenolic lipid composition during development of cashew.Phytochemistry 31, 22952297.

Sullivan, J. T., Richards, C. S., Lloyd, H. A., and Khrisna, G.(1982). Anacardic acid: mollusicide in cashew nut shellliquid. Planta Med. 44, 175177.

WHO (1985). Tech. Rep. Ser. 728.

Table 1. Results of biological assaysSnails Larvae

24 h LC50 (ppm) 24 h LC50 (ppm)

CNSL 5.4 2.6CNSL Na (expressed in free CNSL) 4.0 2.3Endod 5.9 >100Niclosamide 0.15 >100IPS 82 >10 0.03Unsaturated anacardic acids 3.7 1.5Saturated anacardic acid >100 12.5CNSL hydrogenated 34 18

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