curcuma j j econ entomol 95(1) 183

8/14/2019 Curcuma J J Econ Entomol 95(1) 183

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STORED-PRODUCT AND QUARANTINE ENTOMOLOGY

Bioactivities of the Leaf Essential Oil of Curcuma Longa (Var. Ch-66)On Three Species of Stored-Product Beetles (Coleoptera)

A. K. TRIPATHI, V. PRAJAPATI, N. VERMA, J. R. BAHL, R. P. BANSAL, S.P.S. KHANUJA,AND S. KUMAR

Central Institute of Medicinal and Aromatic Plants, P.O. CIMAP, Lucknow-15, India

J. Econ. Entomol. 95(1): 183189 (2002)

ABSTRACT Essential oil extracted from the leaves of turmeric, Curcuma longa L., was investigatedfor contact and fumigant toxicity and itseffect on progeny production in three stored-product beetles,Rhyzopertha dominica F. (lesser grain borer), Sitophilus oryzae L. (rice weevil), and Triboliumcastaneum Herbst (redour beetle). Oviposition-deterrentandovicidal actionsofC. longa leaf oil were

also evaluated against T. castaneum. The oil was insecticidal in both contact and fumigant toxicityassays. The adults of R. dominica were highly susceptible to contact action of C. longa leaf oil, withLD50 value of 36.71 g/mg weight of insect, whereas in the fumigant assay, adults of S. oryzae werehighly susceptible with LC50 value of 11.36 mg/liter air. Further, in T. castaneum, the C. longa oilreduced oviposition and egg hatching by 72 and 80%, respectively at the concentration of 5.2 mg/cm2.At the concentration of 40.5 mg/g food, the oil totally suppressed progeny production of all the threetest insects. Nutritional indices indicate 81% antifeedant action of the oil against R. dominica, S.oryzae and T. castaneum at the highest concentration tested.

KEY WORDS Curcuma, Rhyzopertha, Sitophilus, leaf oil, oviposition-deterrent, antifeedant

TURMERIC, CurcumalongaL. (Zingiberaceae) rhizome,commonlyused asa spice,iswell known for itsmedicinalvalues in the Indian traditional system of medicine andhas been a recipe for several ailments (Sivananda 1958,Raghunath and Mitra 1982, Srimal 1997). Turmeric con-tains pungent, odoriferous oils and oleoresins; the rhi-zomes have been reported to possess many kinds ofbiological activities (Rath et al. 1998).

A number of anti-insect properties of turmeric havebeen documented in the literature. The insect repel-lent components in turmeric are turmerones and ar-turmerone (Su et al. 1982). Petroleum ether extract ofC. longa rhizome has been reported as repellent toTribolium castaneum (Jilani and Su 1983) and insec-ticidal to Plutella xylostella (Morallo-Rejesus et al.1992). Hexane extract of rhizome ofC. longa reducedprogeny production in T. castaneum at 200 ppm con-centration (Jilani et al. 1988). Aqueous extract of C.longa rhizome acted as repellent against Callosobru-chus chinensis (Pati et al. 1996, Patro and Pati 1997),insecticidal to Myzus persicae (Parihar et al. 1999) andreduced larval populations ofAnopheles stephensi Lis-ton(Murugan andJeyabalan1998). Acetone extract ofC. longa rhizome acted as repellent to T. castaneum

(Chander et al. 1994).Turmeric powder has been documented as a stored

grain protectant against T. castaneum (Chander et al.1992 and Parveen and Mondal 1992), S. oryzae and R.dominica (Jilani and Su 1983, Islam et al. 1989), C.chinensis (Shivanna et al. 1994), and also suppressed

populations ofS. oryzae (Chander et al. 1991). Essen-tial oil of turmeric is reported to possess antifungal,anti-bacterial andinsect repellent activities(Saju et al.1998 and Rath et al. 1999).It has been shown to possessfeeding-deterrence against the lesser grain borer,Rhyzopertha dominica (Jilani and Saxena 1990) andrepellent to T. castaneum and R. dominica (Mohiuddinet al. 1993).

Essential oil of turmeric extracted from rhizomecontains-turmerone,-turmeroneand ar-turmeronewhereas leaf oil contains-phellandrene (18.2%), 1,8-cineole (14.2%) and p-cymene (13.3%) as major con-stituents (Sharma et al. 1997). The fresh leaf oil of C.longa contains 82.9% monoterpenes whereas fresh rhi-zome oil has only 16.3% monoterpenes (McCarron etal. 1995). Monoterpenes in general have been welldocumented to be active as fumigants, repellents orinsecticides toward stored grain insects (Tsao et al.1995, Obeng-Ofori and Reichmuth 1997). However,work on the anti-insect potential of essential oil of C.longa leaf is negligible. In the present paper, we reporton the toxic, oviposition-deterrent and populationsuppressant activities ofC. longa leaf oil against threestored-product beetles, R. dominica, S. oryzae and T.

castaneum.

Materials and Methods

Insect Culture. Eggs, larvae and adults of our bee-tle, T. castaneum, were obtained from laboratory- bred

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cultures maintained on whole-wheat our mixed withyeast (17:1, wt:wt) in a dark BOD(Biological OxygenDemand) incubator set at 30 1C and 6570% RH.The T. castaneum larvae used in the experiments were14, 16, and 18 d old, and adults used were approxi-mately 1 wk old. Adults of R. dominica and S. oryzaewere also obtained from laboratory-bred culturemaintained on wheat grains at above rearing condi-tions. All the experiments were conducted in the darkat rearing condition described above.

Extraction of Essential Oil ofC. longa. Fresh leavesof C. longa grown at the farm of this institute werecollected (500 g) and placed in a round-bottom askof the Clevenger type apparatus with water. The oilwas hydrodistilled 34 h in this apparatus (Guenther1955). The essential oil was extracted with acetone,which was then evaporated in vacuum in a rotatoryevaporator (Buchi). Analysis of the essential oil by gaschromatography was conducted using a model CX-

3400, Varian Gas Chromatogram under the followingconditions: carrier gas hydrogen, injector tempera-tures, 220 and 225C, respectively, using a capillarycolumn (Supelcowax 10, 30 m by 0.32 mm, lm thick-ness 0.20 m). The oven was programmed from 80 to150C at 5C/min, then 215C at 7C/min. The areapercentage was obtained on a Varian 440 integrator.The identity of the component was assigned by com-paring their retention time with those of authenticsamples. The density of the oil was also determinedand expressed as g/ml.

Contact Toxicity. A series of dilutions of essential oil

ofC. longa were prepared in acetone. Aliquots of 1 lof the dilutions were topically applied on the thorax ofthe adults of R. dominica, S. oryzae, T. castaneum anddorsal surface of the larvae of T. castaneum using aHamilton Microsyringe (Burkard). Control insectswere treated with 1 l of alone acetone. One hundredadults/larvae in replicates with 10 insects/replicationwere treated for the control and for each concentra-tion of C. longa oil. The treated insects were trans-ferred to glass vials after 1 h of the treatment appli-cation. Culture media were added to the respectivevials after 24 h. Mortality was observed at 24-h periodicintervals up to 7 d. The LD50 and LD95 values weredetermined based on mortality data recorded after 7 d.

Fumigant Toxicity. The fumigant toxicities (Tri-pathi et al. 2000) ofessential oil ofC. longa were tested.Filter paper strips (6 x 8 cm) were impregnated with 10l of one dilution of C. longa oil or acetone (control).

After evaporating the solvent for 5 min in air, the treatedlterpaperstripwas placedatthebottom ofa 1-liter glass

bottle. Twenty adults of three test insect species and/orlarvae of T. castaneum along with culture media werekept separately in small vials that had both open endscovered with 40-mesh copper wire net. Each small vialwas hung with thin metal wire at the geometrical centerof the glass bottle and the bottle tightly closed with thelid. Each set was considered as one replication. Therewere 10 replicates for each dilution and control. After24 h, insects were transferred to fresh vials containingculture media. Mortality data were recorded at 24-hintervals up to 7 d. The LC50 and LC95 values weredetermined after 7 d.

Effect of Essential Oil ofC. longa on Oviposition ofT. castaneum. Aliquots of 0.5 ml of various concen-trations (0.092, 0.92, 2.75, 5.20, 6.42, and 8.26 mg/cm2)ofC. longa leaf oil were applied to pleated black lterpapers (7.0 cm), dried for 30 min, and placed in a glassbowl (7.0 cm diameter by 5 cm high). Twenty adultinsects were introduced on each treated lter paper.Some wheat our was added to the lter paper toprovide food for the insects. Acetone-treated lterpapers were used as controls. Five replicates wereprepared for each concentration and control. Thenumber of eggs laid and adult insects was recordedafter 24 h.

Effect of Essential Oil ofC. longa On Egg Hatchingand subsequent Larval and Adult Survival of T. cas-taneum. Eggs (024 h old) were collected on thepleated black lter papers by placing each lter paperwith 50 adult insects in a bowl (7 cm diameter by 5 cm

Table 1. Main constituents of the leaf essential oil ofC. longa

var. CH-66

Compound name % content in oil

-Pinene 1.83-Pinene 3.42Myrcene 40.191,8-Cineole 13.16-Terpinene 2.03p-Cymene 23.05Terpenolene 0.11Linalool 1.37p-Cymene-8-ol 0.17Myrtenal 0.53ar-Turmerone 0.71-Turmerone 3.0-Turmerone 1.06

Table 2. Contact toxicity ofC. longaleaf essentialoil towardstoredgrain insects determined by topical application to adults andlarvae

of T. castaneum and adults of R. dominica and S. oryzae

Beetle Life stage nLD50

(g/mg wt. of insect)95% ducial

limitsLD95

(g/mg wt. of insect)95% ducial

limitsSlope SE 2a df

R. dominica Adult 300 36.71 11.18120.55 153.84 140.39178.10 6.1 0.02 20.2 NS 16S. oryzae Adult 300 95.50 33.72270.45 640.13 590.12685.18 8.4 0.01 7.9 NS 16

T. castaneum Adult 300 51.49 15.43171.80 259.30 225.78279.82 7.6 0.03 13.1 NS 16Larvae14 d old 300 69.85 23.08211.41 351.56 301.32372.69 6.4 0.01 11.7 NS 1616 d old 300 89.38 62.23128.40 402.25 388.46450.63 7.9 0.05 18.3 NS 1618 d old 300 95.07 65.80137.35 413.26 493.21455.13 8.3 0.03 15.5 NS 16

a NS, 2 is not signicant (P 0.05).

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high) half-lled with rice. There were 4280 eggs perlter paper. These papers were treated with 1.0-mlaliquots of essential oil of C. longa (0.18, 1.8, 5.4, 9.0,12.6, and 16.2 mg/cm2) and allowed to air-dry for 1 h.The treated lter papers were transferred to dishes (7cm diameter, Tarson, India) having 200 g of wheat

our and the mouth of dishes were sealed with blacklter papers using the wax method to prevent theinsects crawling out. The number of eggs that hatchedwas recorded and the number of live larvae wascounted after 7 d. The number of adults that emergedfrom the live larvae was recorded until no furtheradults had emerged. There were 10 replicates for eachtreatment concentration and control.

Progeny Production of T. castaneum in TreatedMedium. One hundred grams of rice were sprayedwith 3.0 ml of various concentrations (0.18,1.8, 5.4, 9.0,12.6, and 16.2 mg/cm2) ofC. longa leaf oil. After thor-ough mixing and evaporation of the solvent, 20 adultsofT. castaneum were added to 20 g treated wheat ourin crystallizing dishes. In the trials with R. dominicaand S. oryzae, wheat grains (20 g) were treated withthe oil. Controls were treated with acetone only. Thedishes were sealed with wax and kept in an incubatorat 30 1C and 6570% RH without light. After 48 h,the adults were removed and the number of deadadults was recorded. The dishes were placed back inthe incubator until F1 adults emerged. The F1 adultswere counted and weighed. Ten replicates were set upfor each concentration and control.

Flour Disc Bioassay. Flour disks were prepared ac-

cording to the method of Huang et al. (1997) withsome modications. Aliquots of 200l of a suspensionof wheat our in water (10 g in 50 ml) were poured into a petri dish to form the disks. The disks were left inthe room to dry, after which they were equilibrated at

30 1C and 6570% RH for 24 h. The weight of theour disks ranged from 35 to 39 mg and the moisturecontent was12.8 0.1%. Flour disks were treated withacetone solutions (10 l) containing various concen-trations of essential oil of C. longa (2.36, 23.66, 71.05,165.75, and 213.15mg/g food) or acetone alone for the

controls. After evaporation of solvent, two disks wereplaced in each preweighed glass vial (2.5 cm diameterby 5.5 cm high). Ten unsexed adults or larvae (1416d old) of T. castaneum were weighed as a group andadded to each vial. Insectswere starved for 24 h beforethe tests. There were ve replicates of each treatmentconcentration and control. The glass vials containingour disks and insects were weighed again after 72 hand the number of dead insects was recorded. Deadinsects were removed from the vials before weighing.

Nutritionalindices werecalculated (ManuwotoandScriber 1982, Farrar et al. 1989) with some modica-tions: Relative Growth Rate (RGR) (A B)/B day, where A weight of live insects (mg) on thethird day/number of live insects on the third day andB original weight of insects (mg)/original numberof insects. Relative consumption rate (RCR)D/(Bday),where Dbiomass ingested (mg)/number oflive insects on the third day. Biomass ingested (orig-inal weight of our disks weight of our disks on thethird day). Percentage efcacy of conversion of in-gested Food (ECI) (RGR/RCR) 100.

The feeding deterrent action was calculated asFeeding-Deterrent Index (Isman et al. 1990)(% FDI)[(CT)/C]100, whereC is the relative

consumption rate of control disks and T is the relativeconsumption rate of treated disks, as the control andtreated disks were placed in separate vials under no-choice tests.

Data Analysis. Probit analysis (Finney 1971) wasused to analyze the dosage-mortality data, whereasmaximum likehood program software (Ross 1987) wasused for the chi-square test. Linear regression analysisusing SPSS (1999) was carried out.

Results

Composition of the C. longa Leaf Essential Oil. Themain constituent prole of the oil is given in Table 1.The oil contains myrcene, p-cymene and 1,8-cineoleas the major compounds. The minor compounds likear-turmerone and -and -turmerone which are re-ported in the literature as insect repellent or insecti-

Table 3. Fumigant toxicity ofC. longa leaf essential oil to adults and larvae of T. castaneumand adults ofR. dominica and S. oryzae

Insect Life stage nLC50

(mg/liter air)95% ducial

limitsLC95

(mg/liter air)95% ducial

limitSlope SE 2a df

R. dominica Adult 600 13.65 10.4717.81 60.20 40.3973.71 6.8 0.04 16.8 NS 20S. oryzae Adult 600 11.36 8.7014.83 51.10 45.7863.90 5.1 0.01 9.7 NS 20T. castaneum Adult 600 19.62 14.0427.41 75.78 68.8093.31 8.1 0.03 11.6 NS 20

Larvae14 d old 600 30.79 17.0555.59 79.50 64.4095.50 9.8 0.01 13.9 NS 2016 d old 600 34.47 29.5040.17 85.40 72.40101.50 7.9 0.01 8.6 NS 2018 d old 600 34.23 30.2338.75 82.18 69.9099.10 5.6 0.04 14.6 NS 20

a NS, 2 is not signicant (P 0.05).

Table 4. Effect of 24-h exposure to C. longa leaf oil impreg-

nated filter papers on oviposition by T. castaneum

Conc,mg/cm2

Mortality of adults at 24 h(mean SE)

No. eggs laid per &(mean SE)

0.0 0.0 0.0 17.5 0.4

0.092 0.0

0.0 13.3

0.50.92 0.0 0.0 11.4 0.42.75 4.4 0.0 7.3 0.35.20 5.9 0.3 4.9 0.66.42 14.3 0.5 1.8 0.48.26 18.0 0.5 1.1 0.3F 50.56

February 2002 TRIPATHI ET AL.: BIOACTIVITIES OF Curcuma longa LEAF OIL 185


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cidal and are also present in higher concentrationcompared with other cultivars ofC. longa grown at thisinstitute. The density of the oil was 0.928 0.005 g/ml.

Contact and Fumigant Toxicity. The essential oil ofC. longa showed both contact and fumigant toxicity toall the insect species tested (Tables 2 and 3). Adults ofR. dominica were highly susceptible to the contacttoxicity ofC. longa leaf oil, but adults ofS. oryzae werehighly tolerant toward leaf oil. In the case of T. cas-taneum, adults were more susceptiblethan larvae.Fur-thermore, the larvae (1418 d old) of T. castaneumbecame progressively more tolerant with age (Table2). However, adults ofS. oryzae were more susceptible

to the fumigant toxicity of C. longa leaf oil than R.dominica and T. castaneum adults. Adults of T. casta-neum were1.5 times more susceptible than its larvae(Table 3). In addition, susceptibilities of the variouslarval stages (1418 d old) to the fumigant toxicity ofC. longa leaf oil were almost equal (Table 3).

Effect of C. longa Leaf Oil on Oviposition, EggHatching and Larval Survival of T. castaneum. Ovi-position was signicantly (F 50; df 5, 56; P 0.05)reduced by C. longa leaf oil at the concentrations of6.42 and 8.26 mg/cm2 (Table 4). However, as therewas14.3 and18.0% mortality, respectively, observed inthese two concentrations after 24 h, the decrease in

Fig. 1. Percentage of egg hatch and survival of larvae (mean SE) ofTribolium castaneum exposed to Curcuma longaleaf oil

Fig. 2. Percentage of adult emergence (mean SE) ofTribolium castaneum from eggs exposed to Curcuma longa leafoil.

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egg production was probably due to fewer survivinginsects laying eggs or the weakened physical stage ofthe insects.Since the oviposition dropped to 28%at 5.2mg/cm2 concentration of treatment with C. longa leafoil, the oil probably acted as an oviposition deterrentat high concentrations.

The hatching of T. castaneum eggs and emergenceof larvae were adversely affected by C. longa leaf oil(Fig. 1). At 5.4 mg/cm2, egg hatching was reduced by80% and no larvae were observed the 9th day afterhatching at the concentration of 12.6 mg/cm2 andabove. Thus, C. longa leaf oil was ovicidal also to theeggsofT. castaneum. When theT. castaneum eggs wereexposed to C. longa leaf oil, the number of adults thatemerged from the treated eggs was signicantly (F89; df 5, 42; P 0.05) decreased (Fig. 2). At 16.2mg/cm2 concentration of C. longa leaf oil, no adultsemerged, since no larvae survived at this concentra-tion in the previous experiment (Fig. 1). Therefore, C.

longa leaf oilprevented theoviposition ofT. castaneumand it suppressed the egg hatching along with thesubsequent development of eggs to larvae and adults.

Effect of C. longa Leaf Oil on F1

Progeny Produc-tion. The number of F1 adults of three species ofbeetles that emerged from treated media decreasedwith increasing concentrations of C. longa leaf oil

Table 5. Emergence and weight of F1 adults of R. dominica,S. oryzae and T. castaneum from eggs placed on culture mediatreated with Curcuma longa leaf oil

Conc.,(mg/g of

culture media)

% parent insectsknocked down

after 48 h(mean SE)

No. F1 adults(mean SE)

Wt. F1 adults, mg(mean SE)

R. dominica

0.0 0.0 0.0 26.0 2.1 0.84 0.034.50 20.40 1.7 3.0 0.3 0.61 0.19

13.50 36.30 2.4 0.0 0.0 0.00 0.0022.50 68.30 4.3 0.0 0.0 0.00 0.0040.50 98.40 5.6 0.0 0.0 0.00 0.00

Fvalue 1.91 5.14

S. oryzae

0.0 0.0 0.0 29.0 8.0 2.81 0.244.50 19.60 2.4 6.0 0.1 2.65 0.63

13.50 35.20 0.9 1.0 0.0 2.74 0.8922.50 67.30 6.9 0.0 0.0 0.00 0.0040.50 100.0 0.0 0.0 0.0 0.00 0.00

Fvalue 2.54 9.25

T. castaneum

0.0 0.0 0.0 22.0 2.4 2.14 0.264.50 11.60 1.9 8.0 1.0 2.00 1.10

13.50 29.80 3.2 6.0 0.9 1.87 0.6722.50 62.70 8.9 2.0 0.2 1.89 0.5640.50 97.30 9.0 0.0 0.0 0.00 0.00

F value 5.90 11.90

Table 6. Nutritional and feeding-deterrence indices ofR. dominica, S. oryzae adults and T. castaneumadults and larvae treated withessential of leaf of Curcuma longa

Life stageOil

(mg/g food)

RGR

(mg/mg/d)

RCR

(mg/mg/d)

% ECI % FDI%

MortalityR. dominica

Adult 0 0.040 0.003 0.379 0.02 10.8 1.2 02.36 0.039 0.006 0.305 0.04 12.6 1.4 32.0 5.83 0

23.66 0.035 0.005 0.257 0.02 13.9 2.5 32.1 2.84 071.05 0.024 0.004 0.218 0.02 11.5 2.5 42.8 2.61 4 7.0

165.75 0.010 0.003 0.109 0.01 17.9 8.3 71.4 2.81 7 2.3213.15 0.030 0.004 0.067 0.01 52.3 3.7 82.4 2.73 8 3.5

S. oryzae

Adult 0 0.017 0.005 0.176 0.01 10.9 0.9 02.36 0.018 0.001 0.167 0.02 10.9 0.6 6.90 2.66 0

23.66 0.014 0.001 0.145 0.02 10.1 1.3 19.5 5.05 071.05 0.012 0.001 0.123 0.03 10.1 0.8 29.9 2.27 0

165.75 0.004 0.001 0.044 0.01 11.6 4.6 74.9 5.32 3 1.5

213.15 0.013 0.001 0.026 0.02 64.9 8.9 84.8 3.58 3 3.8T. castaneum

Adult 0 0.020 0.002 0.181 0.02 11.3 1.3 02.36 0.015 0.001 0.151 0.01 10.2 1.3 15.9 3.64 0

23.66 0.010 0.002 0.103 0.02 11.2 3.6 42.4 5.17 071.05 0.008 0.001 0.107 0.01 8.4 1.5 40.3 4.71 0

165.75 0.007 0.001 0.055 0.00 12.9 2.2 69.2 2.98 1 1.0213.15 0.016 0.002 0.028 0.00 59.1 9.9 84.3 1.76 2 1.1

Larva 0 0.028 0.005 0.211 0.01 13.5 2.3 02.36 0.018 0.007 0.193 0.01 9.50 3.2 13.1 4.38 0

23.66 0.016 0.003 0.161 0.01 5.50 2.5 23.3 7.45 071.05 0.009 0.002 0.134 0.02 10.0 2.3 35.4 8.15 7 5.1

165.75 0.008 0.001 0.070 0.01 10.1 4.0 65.9 6.41 6 2.2213.15 0.016 0.002 0.038 0.01 38.7 5.7 81.7 4.62 8 7.7

Regression data of RGR, relative growth rate; RCR, relative consumption rate; ECI, efciency of conversion of ingested food; and FDI,feeding-deterrence index. R. dominica adults. RGR: Y 0.153 0.0009x, n 9, r2 0.32, P 0.05. RCR: Y 0.322 0.0012x, n 9, r2 0.92, P 0.05. ECI: Y 18.31 0.2750x, n 9, r2 0.86, P 0.05. S. oryzae adult. RGR: Y 0.018 0.0001x, n 9, r2 0.97, P 0.05. RCR:Y 0.17 0.0007x, n 9, r2 0.98, P 0.05. ECI: Y 17.67 0.0295x, n 9, r2 0.77, P 0.05, T. castaneum adults. RGR: Y 0.017 0.0001x, n 9, r2 0.97, P 0.05. RCR: Y 0.151 0.0005x, n 9, r2 0.87, P 0.05. ECI: Y 14.11 0.133x, n 9, r2 0.75, P 0.05.T. castaneum larvae. RGR: Y 0.021 0.0001x, n 9, r2 0.71, P 0.05. RCR: Y 0.19 0.0007x, n 9, r2 0.97, P 0.05. ECI: Y 14.29 0.159x, n 9, r2 0.52, P 0.05. No feeding deterrence was observed.



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Received for publication 3 October 2000; accepted 24 July2001.


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