Proceeduiqs of the 7th International Workmg Conference on Stored-product Protectzon - Volume 1

The preliminarypolyacetylenespests *

study oncompounds

activity of syntheticstored-product

biologicalagainst several

Xu Hanhong! Wan Shuqmg! Cluu Shm-Foon1 Shang Zhrzheng/ LlU Zhun2

Abstmct

The biological actrvities of twelve synthetic polyacetylenescompounds agamst several stored-product insects werereported m this paper' Tnals under laboratory conditionindicated a very high level of activity on poisorung , repellmgand mhibrtmg reproduction The LC50 of compound 2, 5 and10 on Sttophllus oryzae were 0.0128, 0.0295 and0.0069% respectively, they all could mhibrt ItSreproduction completely The majonty of tested compoundscould also mlubit the reproduction of both Oryzaeplulussuruuuneneis and Tribolium castaneum and the rate ofmhibrtion reached over 80 % at a dose of 0.05 % Compound6 and 7 exhibited repellmg actrvities agamst Triboliurncastaneum adults at the concentration of 100 f-Lg/cm2 andthe repellmg activities were 97.44% and 91.89%respectively after 96 hours The results of the inhibitingrate of enzyme m VItro expenments showed that compound10 was an effective mhibitor of acetylcholmesterase(AChE).

Introduction

Polyacetylenes compounds were produced in plants toprotect them from bemg attacked by other creatures. It wasfound that 19 kmds of plants could produce polyacetylenescompounds, and the major kmds of plants mcludeArahaceae, Campanlaceae, Composi teae, Santalacene,Prttosporaceae and Umbelhfereae (Sorensen, 1977) Mostpolyacetylenenes compounds have been Isolated fromCompositeae plants, and the chemical structures of 650kmds have been known, amoung which the typical compoundIS phenylheptatnyne (PHT) Polyacetylenes compoundscould kIll the larvae of mosqUitoes, the eggs of housefhes

* ThIS IS a project supported by Guangdong Natural ScIence Fund

1Dept of Plant Protection, South Chma Agncultural Umverslty,Guangzhou 510642, Chma

2 InstItute of Elemento-Orgamcal Chermstry, Nankm Umverslty,TlanJm 300071, Chma

and fruit flies, and they could also kill some herbivorouspests under the near-violet light (Macrae, 1980) Theyhave shown antlfeedmg and repellent properties on severalmsect species Xu (1992) found that the essential 011 ofArtemisia scoparux had very high biological activity afterhe studied the biological actrvities of about 100 kmds of plantessential Oils, he also found that the active ingredient wascapillene The results of biologtcal activity expenmentsindicated that capillene had rather high biological activrty tosome stored-product pests and some house pests Twelvepolyacetylenes compounds were synthesized by usmgcapillene as the model Expenments were done to evaluatethe biological activities of the synthesized compounds.

Materials and Methods

Insect materials

Sitoplul us oryzae, Tribolium castaneum andOryzaephllus suruuimeneis were first provided byGuangdong Cereals Research Institute, then they wereraised m the Lab of Insect Toxicology, South ChmaAgncultural University.

Compounds

code chemIcal name

1 1,4-dl-t-butyl-dlacetylene2 l-t-butyl-e-hydroxy methyl-diacetylene3 1, 4-dlhydroxymethyl-dtacetylene4 1-phenyl-t-p-methoxyphenyl-diacetylene5 1-phenyl-4-(3,4-methylene dIOXY)-

-phenyl-diacetylene6 1-phenyl-4- m-mtrophenyl-illacetylene7 1-phenyl-4-o- nitrophenyl-illacetylene8 1-phenyl-n-pent-1, 3-dlynyl benzoate9 I-phenyl-4-methyl-diacetylene10 dl-ehtyl-2-propynyl-phosphorothIOate11 3-bromo-2-propyn-l-0112 N-hydrooxymethylacetylenyl- 7-oxablcyclo

[2,2 , 1] hept-5-ene-2 ,3-dlcarboxlmlde

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Proceedmqe of the 7th International WorkLng Conference on Stored-product Protection - Volume 1

Methods

Method of nnxinq food unih. compoundAccordmg to Xu (1992), firstly dissolve compound with

acetone to different concentrations, then mIX the wheatseeds with them respectively After acetone was vaporized,put the treated seeds to finger-shape tubes with height oflOcm and diameter of 2. 5cm In each treatment 50g ofseeds were used and 30 adult msects were put mto eachtube. The control seeds were only nuxed with acetone Alltreatments were replicated three times. Then put all thetubes mto room with stable temperature(25 ± 2°C) Recordthe number of dead and compute the death rate accordmg tothe followmg formula:

Mortality of Treatment -

Amended Mortahty( %) == Mortahty of Control x 100100 - Mortality of Control

Record agam when the first generation (F1) were produced,the mhibitmg rates of reproduction were computed accordmgto the fol1owmg formula:

The number of F1 of control-

Inhibitmg Rate of The number of F1 of treatmentReproducuon/ %) == The number of F1 of the control x 100

Method of studyzng repellinq effectAccordmg to Xu (1992), firstly drvide the filter paper

(FP) WIth diameter of 6cm mto two halves, one half fortreatment, one half for control, then dissolve compound byacetone, then use 1ml of It on the FP so that 100/-1g/cm2 ofthe compound were on the FP, treat the control FP Withonly acetone, then adhere the two halves together Withgallotape on the back of the FP, then put a glass nng Withheight of 5cm and diameter of 6cm on the 'whole' filter,put 40 Tribolium castaneum adults onto each FP,replicate all treatment 2 times, then cover each glass nng

With nylon net cover, then put them mto msectary WIthtable temperature Cf ± 2°C) Record the number of msectpests on the treated FP and the control respectively atdifferent time, then compute the repellmg rate accordmg tothe followmg formula:

'The number of insects on the control FP-

Repelhng == 'The number of 1ffl2Cts on the treated FP x 100Rate( % ) 'The number of insects on the control FP

Method for studymg the mhslntmq effect on AChE inmtroProcedure for preparmg AChE: FIrst put 100

Tribcliuni castaneum larvae to 2 ml phosphate buffer (pH== 7.4) and then homogemzed them The homogenate wascentnfuged at 4500rpm, - 4°C for 15 mmutes. Thesupernatant was used as the source of AChE.Method for evaluating the znhibitory actwzty to AChE:

FIrst dissolve compounds WIth acetone so that theconcentration of acetone IS below 1%, then dilute the aboveto 100/-1g/ml WIth phosphate buffer (pll == 7.4), then mIX50/-11of the AChE source WIth 50/-11of diluted solution. Themixed solution were put m the environment WIthtemperature of 37°C for 5 mmutes. The tubes of 50/-11senne( 4 x 10 - 5 M) mixed WIth AChE m 37°C environment wereused as zero-adjusting tube, and the tubes of 50/-11phosphatebuffer mixed WIth AChE source were used as control tube,then add 100/-11ATCh (1.0 M), VIbrate the mixture m 37°Cenvironment for 15 mmutes, stop the reaction by DTNB-alcohol - phosphate buffer, Ol) values were gamed byputtmg them m 722-type spectrophotometer(412nm). Theinhibitmg rates of AChE were computed by usmg thefollowmg formula:

Ol) value of the control tube-Inhibitmg Rate == aD value of the treatment tube x 100of AChE (%) OD value of the control tube

Table 1. The results of kilhng rate (KR) and mhibitmg reproduction rates ORR) of the 12 compounds to several stored-product msect pests at concentration of 0.05 % .

compoundSitophalu» oruzae Oryzaephzlus surinarnensts Triboli urn. castaneumKR IRR KR IRR KR IRR

1 -4.15 52.63 29.25 63.85 0 25.032 75.72 100.00 47.57 89.99 27 3 85.003 -3.56 42.11 11.44 19.69 0 100.004 10.07 63.16 68.91 74.09 2.22 85.005 71.91 100.00 29.29 84.33 12.77 89.966 1. 79 94.74 5.42 79.94 4.45 10.047 -4.14 31.67 1.67 49.82 0 59.978 0 36.84 3.96 89.93 0 25.039 0.58 100.00 36.96 89.95 2.22 74.9610 100.00 100.00 97 14 * 92.16 * 55.53** 80.05**11 0 1. 74 33.17 53.79 0 29.9812 0 -15.79 1.48 21.68 0 74.96CK 0 0 0 0 0 0

* The concentration was 0 00625% ** the concentration was 0 0125%Treatments were replicated three tunes

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Proceedings of the 7th Internatumal Worki'ng Conference an Stored-product Protection. - Volurne 1

Results and Discussion

The effects of killing and inhibiting reproduction ofpolyacetylenoid compounds to several stored-productpests

The results of Table 1 showed that most compounds had acertain killmg and mhibitmg effects on Oruzaeplulus:surnuimensis , Sttophtlus oryzae and Tribocuun.castimeum The mortahty of compound 2, 5 and 10 toSiiophiiu» oryzae adults were 70% or higher when theconcentration was 0.05% and the LC50 values were 0.0128,0.029 and 0.0069% respectively (Table 2). Compound 2,5 , 6 ,9 and 10 could mhibi t the reproducnon of Suoplu: usoryzae and the mlubitmg rate were 90% or higher (Table1 ). Only compound 10 could kill OryzaephLlussuruuuneneis With a mortahty more than 70% (Table 1),and compound 2, 5, 8, 9 and 10 could mlubit Itsreproduction with inhibitmg rate more than 80% Theresults showed that only compound 10 had killmg effect andthe mam mode of action of most compounds was to mhibi tthe reproduction. The other modes of action of thesecompounds Will be studied m the future.

Table 2. The toxicity evaluation results of several compoundsto Sitophilus oryzae .

Compound LD- P hneLC50( %)

r(WIW)

2 Y=15947+1.6158X 0.9232 0.0128

6 Y = - 0.3689+ 2.1737 X 0.9924 0.0295

10 Y= -2 4751 +4 0576X 0.9998 0.0069

The effects of repelling Tribolium castaneum

The results of Table 3 showed that most of the twelvecompounds could repel Trtbolium castaneurn adults,especially the compound 6,7 and 9 The repellmg rates ofthem were 97.44, 91. 44 and 40 % respectively after 96hours After cuttmg the antenna of the adults and then didsome repelhng expenments, It was found that adults weredistributed very evenly on the FP treated by compound 6, 7and 9. Tlus mdicated that the repellmg target of thesecompounds were the sensitive cells on the antennas of theseadult msect pests. The relationship between the repellmgbehavior and the electrophysiology of the antenna WIll bestudied in the future.

Table 3. The repelhng rate (%) of the 12 compounds (100p.g/cm2) to Triboluum castaneum *

Compound 12h 24h 48h 72h 96h

1 26.88c 18.18d -22.22d - 85. 71a -50.00e

2 78.79d 88.89a 91.43a O.OOe - 37 .1Oe

3 91.43a 97.74a -60 OOd o OOe 16.67d

4 57.14b 66 67b 66.67bc 33 33d - 66.67e

5 33.33c 61 89b 62.07c 70.97bc O.OOe

6 94.59a 97.74a 100.00a lOO.OOa 97.74a

7 94.59a 82.35ab 70. 97bc 78 79ab 91.89b

8 80.00a 40.00c 51.85c 51.85cd O.OOe

9 97.44a 94.74a 88.89ab 82.35ab 40.00c

10 18.18c -1.33d -16.40d - 3. 71a -6.67e

11 85.71a 88.89a 88.89ab 78.99ab O.OOe

12 66.67b 88 89a O.OOd -20.00e O.OOe

* Treatments were rephcated three times, means wrthm a column followed by the same letter are not significantly different (p >0 05 [SASInstrtute 1985J)

The inhibiting effect to the AChE in vitro

Table 4 showed that only compound 8 and 10 could mlnbitthe AChE of Tribolium castaneuni larvae Compound 10 IS

di-ethyl- 2- propynyl-phosphorothioate, It had both killmgeffect and mhibitmg AChE effect The mode of action ofother compounds needs to be studied m the future

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Proceeduujs of the 7th International WorkLng Conference an Stored-product Proteciuni - Volume 1

Table 4 The mhibitmg results of the compounds to theAChEof Tribolium castaneum. larvae m vitro"

Compound X± SD mhibiting rate( %)

1 0.272 ± 0.003abc -18 262 0 .220 ± 0 .005c 4.353 0.2673 ±O .0025abc -16.224 0.274±0.023ab -19.135 0.2683±0.0355bcd -16.656 0.2253 ± 0.0025e 2.47 0 .235 ± 0 .0055de - 2. 178 0.1403 ± 0.0055f 39.009 0.2779 ± 0.0035ab - 20.8310 0.107 ±O.006g 53.4911 0.284±0.0195a -23.4712 0.245 ± O.001cde - 6 52CK 0 .230 ± 0 .230de 0

.. Treatments were replicated three times; means wrtlun a columnfollowed by the same letter are not sigmficantly different (p >0 05[SAS mstitute 1985J)

References

Macrae, W D , Chan, G. F. Q., wat, C K, 1980Examination of naturally occurmg polyacetylenes and (l-

terthienyl for their ability to induce cytogenetic damageExperien tia 36, 1096- 1097.

SAS Institute, 1985. SASuser's guide: statistics, version 5ed SAS Institute, Cary, NC.

Sorensen, N A 1977 Polyacetylenes and conservatism ofchemical characters in the Compositae In: Heywood, V.H. .Harbome.J. B .Turner.B 1. (eds). The biology andchemistry of the Compositae. Academic press, 385 - 409

Xu Hanhong, 1992. The study of biological activities andefficient constituents of essential oils to stored-productpests. Ph D thesis of South Chma AgnculturalUniversity

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