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290 J. Agric. Food Chem. 1982, 30, 90-292Isolation, Purification, and Characterization of Insect Repellents from Curcumalonga L.

Helen C. F. Su,* Robert H orvat, and G hulam Jilani'

Two compounds were isolated from C urcuma longa L. a nd identified from their sp ectral characteristicsas 2-methyl-6-(4-methylphenyl)-2-hepten-4-onear-turmerone) and 2-methyl-6-(4-methyl-l,4-cyclo-hexadien-l-yl)-2-hepten-4-oneturmerone). ar-Turme rone and turm erone gave an average 62.9% (classIV) nd 43.1% (class 111) repellency, respectively, to TriboZium castaneum (Hbst.)after 8w ee b of study.Turmerone was unstable thermally an d also a t amb ient temperature in th e presence of air, yielding itsdimer or the more stable ar-turmerone.

Tumeric, Curcuma longa L., is a tropical herb of theZingiberaceae family indigenous to south ern Asia. Th earomatic yellow powder from its mature rhizomes wa s usedin Asian coun tries for many centuries as a yellow vegetabledye for silks and cottons. I t is still used in foods as acondiment, particularly as an e ssential ingred ient of currypowder, in m edicine as a stomachic, carm inative, anthe l-mintic, laxative, and cure for liver ailment, and also asanant repellent in India (Sreenivasamurthy and Krishna-murthy, 1959; Wa tt a nd Breyer-Brandwijk, 1962).Th e strong coloring pigment in turm eric contains cur-cumin, 1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadi-ene-3,5-dione, and two other curcuminoids (Srinivasan,1953; Lubis, 1968; Sha nka racha rya and N atara jan, 1974;Krishnamurthy et al., 1976). The red-yellow essential oilof turmeric, either from steam distillation or from anoil-selecting solvent extraction, contains d-a-p helland rene,d-sabinene , zingiberene, borneol, l,&cineole, turmeron e,ar-turmeron e, sesqu iterpene alcohols, a- nd y-atlantone,and bisabolene (Gildemeister and Hoffmann, 1956; Mima,1959; Shanka racharya a nd Natarajan, 1974; Salzer, 1975Govindarajan, 1980).

Shankar et al. (1980) conducted feeding toxicity studiesof turmeric and its alcohol extra ct in rats, guinea pigs, an dmonkeys. Th e results indicated that turmeric is nontoxiceven at th e very high level tested. The re was absolutelyno mortality or any morphological and histological ab-normalities in the experimental animals at 2.5 g of tur-meric/kg of body weight which corresponds to a maximumconsum ption of 5 g/day for an adult hum an with an av-erage body weight of 70 kg. It is th e practice in somesouth ern Asian countries; i.e., India an d Paki stan, to storerice or wheat by mixing it with 2% of turmeric powder(Chatterjee, 1980). In th is paper, we report the isolation,purification, and identification of th e rep ellent componentsof turmeric to T ribolium ca staneu m (Hbst.).MATERIALS AND M ETHODS

Extraction of Turmeric. Powdered turmeric (im-ported from India) was purchased from Lex Enterprise,Stored-Product Insects Research and DevelopmentLabo ratory, Agricultural Research, Science and EducationAdministration, US. epartm ent of Agriculture, Savan -na h, Georgia 31403 (H.C.F.S. an d G.J.), and Rich ard B.Russell Agricultural Research Center, Agricultural Re-search, Science and Educa tion Adminstration, U S . De-par tment of Agriculture, Athens, Georgia 30604 (R.H.).lFulbright-Hays Predoctoral Research Associate,1978-1979. Pre sen t address: Honeybee Man agem ent,Pak istan A gricultural Research Coun cil, Islamabad, Pak-istan.

Atla nta, GA. One kilogram of the powder was extractedin a Soxhlet extractor with petroleum e ther ( bp 30-60 "C)for 24 h. Th e petroleum ether solution was concen tratedunder reduced pressure in a rotary evaporator to obtainth e crude turmeric extract.Liquid Column Chromatographic (LCC) Fraction-ation of Crude Turmeric Extract. S t ep 1. Each 1.4-1.6-g portion of th e crud e extrac t was placed on a colum n(40 X 2.0 cm i.d.) of silica gel (70-230 mesh; EM R eagent;extra pure) and eluted with chloroform. A total of 1202-mL fractions was collected. After th e solvent was re-moved, every third fraction was spot teste d by TL C (seebelow for procedure). Th e fractions possessing the sa meRr values were combined, and each combined fraction wasbioassayed for repellency to T. castaneum. The insectactive fraction was further fractionated.S tep 2. Eac h 150-160-mg sample of the ac tive fractionfrom step 1wa s furthe r separated by column chromatog-raphy in a 40 X 1.2 cm i.d. column of silica gel and elute dwith benzene. A totalof 90 2-mL fractions wa s collected.TL C analysis was made on every third fraction. Th efractions with only Rf0.25-0.28, the mixture of R 0.25-0.28and 0.30-0.33, and only R 0.30-0.33 were com6ined sep-arately. Th e two pure combined fractions were bioassayed.Thin-Layer Chromatographic Analysis of LCCFractions. Brinkm an EM reagent, precoated silica gelG FZu, 0 X 20 cm a nalytical chroma toplates were used.A small samp le of each selected fraction was applied onspots 2.5 cm above th e lower edge and 1.5 cm from eachother. Eac h plate was developed in benzene and thenexamined under UV at 254 nm.High-Performance Liquid Chromatographic Puri-fication. A Waters Associates Model ALC/GPC 244high-pressure liquid chromatograph with a Model 6000Apum p, a U6K injector, an R401 differential refractometer,and a Model 440 UV detector with a 300 X 7.8 mm i.d.pBondap ak C18 column (octadecyltrichlorsilanecovalentlybonded to lo-" pPorasil packing) was used. Me tha-nol-water ( 8 2 by volume, degassed) was used as the elutingsystem. Th e column effluent was mon itored at 254 nm(2.0 AUFS), and th e response wa s recorded on a HoustonOmniscribe Model B-5217-1 recorder.Each of the two LCC separated components was dis-solved individually in methanol to obtain the con centrationof approximately 20 Mg/pL an d filtered throu gh a W atersAssociates sample clarification kit with 0.5-pm Milliporeorganic filter system. For each run, a 5-pL aliquot of th estock solution was injected onto the column with th e flowrate of th e eluting solvent at 1.5 mL/m in. Th e desiredpeak of the compound as ndicated from th e recorder wasrecycled through t he column once, and the effluent of th erecycled peak was collected. Th e process was repeated

This article not subject to US . Copyright. Published 1982 by th e American Chemical Society