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한국환경농학회지제 28 권제 3 호 (2009) Korean Journal of Environmental Agriculture Vol. 28, No. 3, pp. 281-288 연구보문 Strobilurin 계살균제의시설재배참외중잔류양상 박은정 이주희 김태화 김장억 * 경북대학교농업생명과학대학응용생명과학부 (2009 년 9 월 9 일접수, 2009 년 9 월 26 일수리 ) Residual Patterns of Strobilurin Fungicides in Korean Melon under Plastic Film House Condition Eun-jeong Park, Ju-Hee Lee, Tae-Hwa Kim, and Jang-Eok Kim * (School of Applied Biosciences, College of Agricultural Life Science, Kyungpook National University, Daegu 702-701, Korea.) ABSTRACT: The strobilurin fungicides, azoxystrobin and kresoxim-methyl, were investigated to know the biological half-lives and dissipation patterns in Korean melon under plastic film house condition. Used pesticides for field application were 20% azoxystrobin of suspension concentrate and 47% kresoxim-methyl of water dispersible granule. Two pesticides were sprayed at recommended and double dose rate. Pesticide residues in Korean melon were analyzed until 14 days after application. The azoxystrobin was analyzed by HPLC equipped with UV detector after cleanup with florisil glass column. Initial residue concentrations of azoxystrobin in Korean melon at recommended and double dose rate were 0.09 mg/kg and 0.14 mg/kg, respectively. Those were less than 0.2 mg/kg maximum residue limit of Korean melon established by KFDA. The biological half-lives of azoxystrobin in Korean melon were 4.7 days at recommended dose rate and 7.8 days at double dose rate. Initial concentrations of kresoxim-methyl which was analyzed by GLC-ECD in Korean melon at recommended and double dose rate were 0.10 mg/kg and 0.23 mg/kg, respectively. Those were less than 1.0 mg/kg, MRL. The biological half-lives of kresoxim-methyl in Korean melon were 4.1 days at recommended dose rate and 4.8 days at double dose rate. The residue amounts of both pesticide was lower than MRL and biological half-lives were not so long. Because the weight of Korean melon under plastic film house condition was fast increased during cultivation. Key Words: Biological half-life, Strobilurin Fungicides, Azoxystrobin, Kresoxim-methyl, Korean melon 서론 현재국내농식품의잔류농약에대한안전관리는생산단계와유통단계로구분하여국립농산물품질관리원과식품의약품안전청이담당하고있다. 국내에서유통전까지농산물에대한농약의잔류검사는국립농산물품질관리원을중심으로이루어지고있으며농림수산식품부에서는 1999년부터농산물품질관리법에근거하여생산단계농작물에대하여잔류허용기준을설정하고계속적인잔류검사및모니터링연구를통하여부적합한농산물의출하를사전에차단하고있다 1-5). 이기준에따라농산물이유통및소비되기전에수확물의농약잔류량을조사평가하여부적합한농산물에대한출하 * 연락저자 : Tel: +82-53-950-5720 Fax: +82-53-953-7233 E-mail: jekim@knu.ac.kr 연기및폐기등의과정을거쳐생산자의손실을최소화하고소비자의안전성을확보하고있다. 따라서잔류허용기준이설정되어있는농약에대해서작물의재배기간중분해양상을조사하고반감기를산출하여적절한농산물의출하시기를조절하는연구는소비자의안전과생산자의권익을위해꼭필요한연구이다. 참외는박과에속하는 1년생식물로서우리나라여름철의대표적인과채류의하나이다. 그중에서경상북도성주시의참외재배면적은 2008년을기준으로 3,820 ha를차지하여전국대비 71% 에이르고있다 6). 시설재배의특성상참외재배시농약의사용은불가피하며사용되는농약들은참외에주로발생하는노균병, 흰가루병, 총채벌레및응애류등의방제를위해서 strobilurin계, anilide계, triazole계, pyrimidine계와유기인계통의농약들이사용되고있다 7-15). 이렇게사용된여러가지농약들은참외의재배기간중분 281

282 박은정 이주희 김태화 김장억 해, 휘산, 용탈및세척등에의해서참외중잔류농도는상당히감소하지만출하되는시점까지도일정수준은참외에그대로잔류하고있다 16). 또한참외에잔류된농약은국내과채류에등록된농약의독성과토양잔류시험결과를이용한농약에대한환경영향평가에서도다른과채류에비하여농작업자, 소비자및환경에대한영향지수가약간더높게나타났다 17). 따라서참외에사용되는농약에대하여생산단계에서출하단계까지잔류농약의변화정도를정확히파악하여출하시점에서의안전성유무를평가하는연구는생산자에게는작물의재배단계에서합리적인농약의사용을유도하고소비자에게는안전한농산물을공급하는데중요한연구이다. 따라서본연구는 strobilurin계살균제 azoxystrobin 과 kresoximmethyl를참외재배포장에직접살포하여일정기간별로참외중의농약잔류특성을조사하고, kinetic 해석에따른합리적인회귀식과생물학적인반감기를산출하여잔류허용기준치와비교평가함으로서생산단계에서의농약잔류허용기준을설정하기위한기초자료로활용하고자하였다. 재료및방법 약제및시약살균제인 azoxystrobin(99.7%) 과 kresoxim-methyl(99.8%) 의표준품은 Dr. Ehrenstorfer( 독일 ) 로부터구입하여사용하였으며시험농약의구조는 Table 1과같다. 포장시험에사용된 azoxystrobin 은 20% 액상수화제 ( 상표명 : 오티바 ), kresoximmethyl은 47% 입상수화제 ( 상표명 : 해비치 ) 를살포하였다. 잔류농약의분석을위한 acetone, acetonitrile, dichloromethane, ethyl acetate 및 n-hexane은 Burdick & Jackson(U.S.A.) 사로부터농약잔류분석용시약을구입하여사용하였다. 시료의정제를위한 glass column( 지름 16 mm) 의충진제는 florisil이었으며 Sigma-aldrich Chemical Co.(U.S.A.) 사의 F9127(60-100 mesh, 농약잔류분석용 ) 을구입하여사용하였다. Sodium sulfate(gr급 ) 및 sodium chloride (EP급) 은 Junsei Chemical(Japan) 에서구입하여사용하였다. 시험구배치및약제살포시험포장은경상북도성주군대가면옥성리에위치한시 설재배단지의참외경작농가의포장을사용하였다. 시험에사용된참외 (Cucumis melo L.) 의품종은명문으로 2007 년 12월에정식하여이듬해인 2008년 4월부터수확을시작하였으며참외의주산지인성주지역의관행적인참외재배방법에따라서폭 5 m 길이 75 m의단동식비닐하우스내에서시험구는폭 2 m 길이 10 m크기로구획을정리하고, 시험구배치는완전임의배치법 3반복으로하였으며, 처리구간의오염을방지하기위하여폭 2 m 길이 1 m의완충지대를설치하였다. 참외에대한농약의살포는농약사용지침서의안전사용기준에준하여 2008년 6월 13일에 azoxystrobin 20% 액상수화제는 8 ml/20 L(2,500배희석, 기준량 ) 및 16 m/20 L(1,250배희석, 배량 ) 희석하고, kresoxim-methyl 47% 입상수화제는 6.7 g/20 L(3,000배희석, 기준량 ) 및 13.4 g/20 L(1,500배희석, 배량 ) 수준으로희석하여배부식분무기를사용하여시험구전체에골고루살포하였다. 약제살포후 2 시간부터 0, 1, 3, 5, 7, 10 및 14일까지생육상태가균일한시료를 15개이상씩무작위로채취하였다. 이시료는처리구별로포장상자에개별포장하여실험실로운반하였으며즉시균질화한후플라스틱용기에넣고밀봉한후분석할때까지 -20 의냉동고에보관하였다. 농약분석 균질화된참외시료 25 g을칭량하여취하고 acetone 100 ml를가한후 homogenizer를이용하여 12,000 rpm으로 3분동안마쇄추출하였다. 이추출액을 Celite 545가깔린 Büchner funnel에서감압여과한후 1,000 ml의분액여두로옮기고증류수 500 ml, 소금으로포화된증류수 50 ml 와 dichloromethane 50 ml를가하고 3분동안격렬하게흔들어정치시켰다. dichloromethane층을 sodium sulfate anhydrous층을통과시켜탈수시키고감압농축기를이용하여농축건고한후 florisil 10 g이충진된 glass column에서정제하였다. Azoxystrobin과 kresozim-methyl의추출및분배과정은동일하였으며정제과정은서로다른혼합용출용매를이용하여농약을용출시켰다. Azoxystrobin의정제는 florisil 이충진된 column을 n-hexane 100 ml로활성화시킨후농축건고된시료를 n-hexane 10 ml 에재용해하여옮기고 ethyl Table 1. Chemical Structure of azoxystrobin and kresoxim-methyl Structure IUPAC Name CN O N Azoxystrobin N O CH 3 O CO 2 CH 3 methyl (E)-2-{2-[6-(2-cyanophenoxy) pyrimidin-4-yloxy]phenyl}-3-methoxyacrylate CH 3 Kresoxim-methyl O CH N OCH 3 O 3 O methyl (E)-methoxyimino[2-(o-tolyloxymethyl) phenyl]acetate

Strobilurin 계살균제의잔류 283 acetate : n-hexane(5/95, v/v) 혼합용액 100 ml와 acetone : n-hexane(20/80, v/v) 혼합용액 30 ml를차례대로용출시켜서버렸다. 계속해서 acetone : n-hexane(20/80, v/v) 혼합용액 60 ml로다시용출시켜서모은후 40 이하의수욕상에서감압농축하였다. 농축된잔사를 acetonitrile 2.5 ml 에다시용해시켜 0.45 μm membrane filter로여과하여그여과액을 HPLC-UVD(YoungLin ACME-9000, 235 nm) 로분석하였다. Column은 Waters X-Bridge[15 cm(l.) x 4.6 mm(i.d.), particle size: 5 μm] 를사용하였으며 mobile phase로는 acetonitrile : H 2O(50/50, v/v) 를사용하여유속 1.0 ml/min 조건으로분석하였다. Kresoxim-methyl의정제는 florisil이충진된 column 을 n-hexane 100 ml로활성화시킨후농축건고된시료를 n-hexane 10 ml에재용해하여가하고 ethyl acetate : n-hexane(5/95, v/v) 혼합용액으로 30 ml를용출시켜서버렸다. 계속해서 ethyl acetate : n-hexane(5/95, v/v) 혼합용액 70 ml로다시용출시켜서모은시료를 40 이하의수욕상에서감압농축하였다. 이농축된잔사를잔류분석용 acetone 2.5 ml 에재용해한후 GLC-ECD(Varian CP-3800) 로분석하였다. Column 은 DB-1[30 m(l.) 0.53 mm(i.d.), film thickness 0.5 μm] 을사용하였으며기기의분석온도는 injector 260, oven 220, detector 300 에서 gas flow(n 2) 4.0 ml/min 의조건으로분석하였다. 회수율시험무처리참외시료 25 g에 azoxystrobin의경우에는 0.1 mg/kg 및 1.0 mg/kg 수준으로농약표준품을처리하였고, kresoxim-methyl의경우에는 0.1 mg/kg 및 0.4 mg/kg 수준으로농약표준품을처리한후 1시간정도방치하여유기용매를휘발시켰다. 이시료를확립된분석방법에따라서 acetone을가하고추출, 분배및정제과정을거쳐분석을실시하였다. 분석은각처리농도별로 3반복으로실시하였으며처리구의평균과표준편차를구하였다. 결과및고찰 회수율및검출한계참외중회수율시험결과는 Table 2와같았으며 azoxystrobin 및 kresoxim-methyl 표준검량선의상관계수 (r 2 ) 는각각 0.9998 및 0.9995로서정량분석을위한양호한직선성을나타내었다. Azoxystrobin의회수율시험결과는 93.5~104.3% 수준이었으며 HPLC/UVD 상에서의최소검출량은 4.0 ng 이었고검출한계는 0.02 mg/kg 이었다. Kresoxim-methyl의회수율시험의결과는 97.2 ~ 112.4% 이었고최소검출량은 0.1 ng, 검출한계는각각 0.01 mg/kg 이었다. 표준품및회수율시험중의분석 chromatogram은각각 Fig. 1 및 2와같이나타났으며농약의 retention time과중첩되는방해 peak는발생되지않았다. Table 2. Recoveries and limit of detection for azoxystrobin and kresoxim-methyl analysis Pesticide Azoxystrobin Kresoximmethyl Fortification Recovery(%) MDA b) level (ng) (mg/kg) 1 2 3 Mean±SD a) 0.1 104.3 99.0 99.5 100.9±2.9 1.0 96.6 93.5 98.7 96.3±2.6 0.1 105.9 112.4 106.0 108.1±3.7 0.4 97.4 101.2 104.1 100.9±3.4 a) SD, Standard Deviation ; b) MDA, Minimum detectable amount; c) LOD, Limit of detection LOD c) (ng) 4 0.02 0.1 0.01 A B C Azoxystrobin 200 ng Control Recovery 1.0 mg/kg Fig. 1. HPLC chromatograms of azoxystrobin in Korean melon.

284 박은정 이주희 김태화 김장억 A B C Kresoxim-methyl 10 ng Control Recovery 0.4 mg/kg Fig. 2. GLC chromatograms of kresoxim-methyl in Korean melon. 400 100 100 Temperature Humidity 350 80 80 Weight (g) 300 Temperature ( ) 60 40 60 40 Humidity (%) 250 20 20 200 16 0 0 Days after application (days) Days after application (days) Fig. 3. Variation of Korean melon weight during the experimental period. Fig. 4. The climatic conditions of the plastic film house during the experimental period. 참외의증체율및기상조건 포장실험기간중참외의무게변화는 Fig. 3에나타내었다. 경과일수별로채취된참외시료는실험실로이송직후개별로무게를측정하여시험기간중참외의무게변화및증체율을기록하였다. 참외는다른박과류 18) 나엽채류 19-20) 에비하여무게의증체율은상대적으로낮았으며시험기간 (2008년 6 월 13일~6월 27일 ) 중평균 50% 정도의무게가증가하였다. 시험기간중시설내의온도및습도의변화를연속적으로측정한결과는 Fig. 4와같았으며온도의측정및기록은자동온습도기록계 (SATO, SK-L200TH, Japan) 를이용하였다. 시험기간중비닐하우스내의평균온도는 26.6 ± 3.7, 평균습도는 68.5 ± 9.8% 이었다. 참외중잔류농약시설재배참외에살포한농약잔류량은 Fig. 5와같이살포후시간의경과에따라서지속적으로감소하는경향을보였다. Azoxystrobin 살포직후의참외중잔류량은기준량 및배량처리구에서각각 0.09과 0.14 mg/kg으로참외에식품의약품안전청에서설정한 azoxystrobin의잔류허용기준인 0.2 mg/kg보다낮게나타났으며, kresoxim-methyl의최대잔류량도처리직후기준량및배량처리구에서각각 0.10 mg/kg 및 0.23 mg/kg으로서참외에설정된잔류허용기준인 1.0 mg/kg 보다낮은수준이었다. Azoxystrobin 과 kresoxim-methyl의반감기소실곡선식은 Table 3에나타낸바와같았다. Azoxystrobin의반감기소실곡선은기준량처리구에서 y=0.0766e -0.138x, 배량처리구에서 y=0.1143e -0.089x 이었으며잔류반감기는각각 5.0일및 7.8일이었다. Kresoximmethyl 의반감기소실곡선은기준량처리구에서 y=0.0896e -0.1672x, 배량처리구에서 y=0.1504e -0.1446x 이었으며잔류반감기는기준량처리구에서 4.1일, 배량처리구에서 4.8일이었다. Azoxystrobin과 kresoxim-methyl은동일한 strobilurin 계통의약제이며주성분의함량이각각 20% 와 47% 로서함량이높은 kresoxim-methyl 이 azoxystrobin에비하여다소높게나타났다.

Strobilurin 계살균제의잔류 285 e -1 e -1 Residue Conc. of Azoxystrobin (ln) e -2 e -3 e -4 Recommended Double A Residue Conc. of Kresoxim-methyl (ln) e -2 e -3 e -4 Recommended Double B e -5 Days after application(days) e -5 Days after application(days) Fig. 5. Dissipation patterns of azoxystrobin and kresoxim-methyl in Korean melon under plastic film house condition. A: Azoxystrobin, B: Kresoxim-methyl. Table 3. Biological half-lives of pesticides in Korean melon under plastic film house condition Pesticides Application Regression curve a) Equation r 2 Half-lives(days) Azoxystrobin Recommended Y=0.0766e -0.138x 0.9424 5.0 Double Y=0.1143e -0.089x 0.9310 7.8 Kresoxim-methyl Recommended Y=0.0896e -0.1672x 0.9428 4.1 Double Y=0.1504e -0.1446x 0.9040 4.8 a) Regression curve, Based on the first-order kinetics 그러나잔류량의차이는두약제의함량차이가 2.35배인것을감안한다면비교적크게차이가나지않았으며, 이렇게차이가적은이유는 kresoxim-methyl의제형이입상수화제로서액상수화제인 azoxystrobin에비하여부착성이다소떨어지기때문인것으로사료된다. 또한두농약모두살포직후의잔류량이이렇게낮은이유는참외의생육특성에기인할것으로사료된다. 사과나복숭아등과같은과수의경우에는움푹파인과일꼭지부분에농약살포액이상당량집적되어고농도로존재하면서일부만이휘산, 광분해및강우에의한유실등으로제거된다 21). 그러나참외는사과나복숭아처럼움푹파인꼭지부분이없고포도나방울토마토등과비교해서표면적이적으며표면도털이없이매끄럽기때문에농약의부착이적게이루어지는것으로사료된다 22). 또한수정이이루어진꽃은착과가이루어지는데시험이이루어진시기는잎의생육이왕성하게이루어져서참외과일을가리기때문에농약살포시부착성이떨어지는것도살포직후농약의잔류량이낮은원인중한가지일것으로사료된다. 참외중농약의절대잔류량참외포장에살포된농약은시설내의높은온도와습도에의해분해및휘발이이루어지고작물의생육을위한관수처리등에의해서세척되어농약의잔류량은점차감소하게된다. 또한이런순수한분해, 휘발및세척외에도참외의재배 기간중무게가증가함에따라서희석효과도 azoxystrobin 과 kresoxim-methyl의잔류농도감소에큰영향을끼치게된다. 따라서참외의무게증가에의한희석효과를배제한참외중농약의순수한잔류량을다음식 1과같이계산하여참외중농약의절대잔류농도라고정의한다면이절대잔류농도의감소추이는다음 Fig. 6과같이나타낼수있으며, 반감기소실곡선식은 Table 4와같이나타내었다. Absolute Residue Concentrate in Korean melon(mg/kg) - Equation 1 = Residue Concentrate in Korean melon(mg/kg) Increase Rate of Korean melon weight(mg/kg) 또한이절대잔류농도의감소는참외중 azoxystrobin과 kresoxim-methyl의잔류량이실제로분해되는것으로추정할수있다. Azoxystrobin 살포직후의 0일차절대잔류농도는기준량및배량처리구에서각각 0.09 및 0.14 mg/kg으로나타났으며 14일경과한때의절대잔류농도는각각 0.01 및 0.05 mg/kg 이었다. 시험기간중의무게의증가율이약 50% 정도일어났으므로 14일동안의시험기간중분해나휘발및관수처리중세척등에의한순수한농약의감소는기준량의경우 83.6%, 배량의경우에는 67.0% 정도이루어진

286 박은정 이주희 김태화 김장억 Absolute Residue Conc. of Azoxystrobin (ln) e -1 e -2 e -3 e -4 e -5 Days after application(days) Recommended Double A Absolute Residue Conc. of Kresoxim-methyl (ln) e -1 e -2 e -3 e -4 e -5 Days after application(days) Recommended Double Fig. 6. Dissipation patterns of absolute residue amount of azoxystrobin and kresoxim-methyl. A: Azoxystrobin, B: Kresoxim-methyl. B Table 4. Biological half-lives of absolute residue amount pesticides in Korean melon under plastic film house condition Pesticides Application Regression curve a) Equation r 2 Half-lives(days) Azoxystrobin Recommended Y=0.075e -0.114x 0.9687 6.1 Double Y=0.112e -0.0633x 0.8545 11.0 Kresoximmethyl Recommended Y=0.087e -0.1251x 0.8797 5.5 Double Y=0.152e -0.1248x 0.8808 5.6 a) Regression curve, Based on the first-order kinetics 것을알수있다. Kresoxim-methyl의살포직후의절대잔류농도는기준량및배량처리구에서각각 0.10 및 0.23 mg/kg으로나타났으며 14일경과한때의절대잔류농도는각각 0.01 및 0.03 mg/kg 이었다. 시험기간중의무게의증가율은약 50% 이었으며 14일동안의시험기간중분해나휘발및관수처리중세척등에의한순수한농약감소율은기준량의경우 86.2%, 배량의경우에는 87.8% 이었다. 본연구의결과에서알수있듯이 azoxystrobin과 kresoximmethyl의경우에는참외에대한잔류허용기준이각각 0.2 및 1.0 mg/kg으로설정되어있어서농약의살포직후잔류량이허용기준을초과하지않았으며정상적인시설재배환경에서농약이살포된경우에는농약살포직후에도참외의출하가가능할것으로사료된다. 요약 Azoxystrobin과 kresoxim-methyl의참외중반감기와잔류양상을조사하였다. Azoxystrobin의참외중 0일차잔류량은기준량및배량처리구에서각각 0.09 및 0.14 mg/kg 으로나타났으며농약의반감기소실곡선식은 y=0.0766e -0.138x (r 2 =0.9424) 및 y=0.1143e -0.0890x (r 2 =0.9310) 이었으며생물학적반감기는각각 5.0일및 7.8일이었다. Kresoxim-methyl 의참외중 0일차잔류량은기준량및배량처리구에서각각 0.10 및 0.23 mg/kg으로나타났으며농약의소실곡선식은 y=0.0896e -0.1672x (r 2 =0.9428) 및 y=0.1504e -0.1446x (r 2 =0.9040) 이었고생물학적인반감기는각각 4.1일및 4.8일이었다. 재배기간중참외의무게증가에농약희석효과를배제한절대잔류농도는약제살포후 14일경과시 azoxystrobin은기준량및배량에서 0.01 및 0.05 mg/kg으로서각각 83.6 및 67% 의농약이분해되었다. Kresoxim-methyl은각각 0.01 및 0.03 mg/kg 으로서 86.2 및 87.8% 정도의농약분해율을보였다. 감사의글 이연구는 2008년국립농산물품질관리원의 생산단계농산물의잔류농약허용기준설정연구 의연구비지원으로수행한결과의일부이며, 연구비지원에감사드립니다. 참고문헌 1. Park, D. S., Seong, K. Y., Choi, K. I. and Hur, J. H. (2005) Field tolerance of pesticides in the strawberry and comparison of biological half-lives estimated from kinetic models. The Korean Journal of Pesticide Science, 9(3), 231-236.

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