22(4)-11(오덕환).fm

J. Fd Hyg. Safety Vol. 22, No. 4, pp. 359~364 (2007) Journal of Food Hygiene and Safety Available online at http://www.foodhygiene.or.kr Icariin 함량에따른삼지구엽초추출물의추출조건및항산화효과 김서진 오덕환 * 강원대학교생명공학부식품생명공학전공 Free Radical Scavenging Effect and Extraction of Ethanol Extracts of Epimedium koreanum Nakai Containing Different Icariin Quantity Seo-Jin Kim and Deog-Hwan Oh* Department of Food Science and Biotechnology, College of Bioscience and Biotechnology, Kangwon National University, Chunchon 200-701, Korea (Received November 21, 2007/Accepted December 19, 2007) ABSTRACT This study was conducted to determine the optimum condition of extraction and antioxidant activity of ethanol extracts of Epimedium koreanum Nakai based on the icariin quantity. Also, further organic solvent fractions of hexane, chloroform, ethyl acetate and butanol were obtained from the ethanol extract of Epimedium koreanum Nakai at different temperatures. Total ethanol extraction yield of wild grape seed ranged from 11.8% to 39.3% depending on the concentration of icariin as well as different ethanol concentration, extraction temperature and time condition. Among different extraction temperatures and time, the highest extraction yield of 39.3% was obtained at 70% ethanol for 3 hour at 50 o C in the sample containing the 0.596% icariin (KE9412). In the meantime, the strongest free radical scavenging effect (RC 50 ) with 18.9 µg/ml in the KE9412 sample was observed in 70% ethanol extract of Epimedium koreanum Nakai extracted for 7 hour at 70 o C, while RC 50 with 35.2 µg/ml was observed in the KE9405 (0.20% icariin content) sample at same condition. Also, antioxidant activity of ethanol extracts of Epimedium koreanum Nakai increased as icariin concentration increased. Among each fraction obtained from organic solvents, butanol fraction was found to have the strongest RC 50 (39.2 µg/ml) and followed by ethylacetate (49.0 µg/ml), water (118.8 µg/ml), ethylacetate (119.8 µg/ml), and chloroform fraction (138.5 µg/ml) respectively. Key words: Epimedium koreanum Nakai, free radical scavenging, icariin 삼지구엽초 (Epimedium koreanium Nakai) 는매자나무과에속하는다년생초본식물로서주로경기, 강원중북부지역을중심으로자생하고있으며초장은 30 cm 내외로줄기의가지가세개로 1가지에 3개의잎이달려삼지 ( 三枝 ) 구엽 ( 九葉 ) 초라한다. 잎은잎자루가길고난형이고끝이뾰족하며밑부분이심장형으로서가장자리에털같은잔톱니가있다. 2) 삼지구엽초의주성분은 flavonoid인 icariin, 3,4) epimedoside A등으로밝혀졌으며, 5) 이중 icariin은삼지구엽초의주약효성분으로고혈압에효과를나타내는보고와 6) 간독성을억제한다는보고가있으며, 7) flavonoid glycoside 인 epimedin A, B 및 C가추가로보고되었다. 8) 또한삼 *Correspondence to: Deog-Hwan Oh, Department of Food Science and Biotechnology, College of Bioscience and Biotechnology, Kangwon National University, Chunchon, 200-701, Korea. Tel: 82-33-250-250-6457, Fax: 82-33-250-250-6457, E-mail: deoghwa @kangwon.ac.kr 지구엽초지상부를건조한것을음양곽 ( 淫羊 ) 이라하는데한방에서는강정, 항암, 항바이러스, 이뇨등에효능이있는것으로알려지고있다. 9,10) 식품의가공또는저장중에있어서품질을저하시키는화학적원인중의하나는지질의산화이며이를방지하기위한항산화제의첨가방법이널리이용되어왔다. 그러나그간가장널리이용되어온 tocopherol류는효과가비교적낮은편이고 BHA와 BHT는효과는뛰어나지만최근변이원성및독성이지적되면서보다안전하고효력이강한천연항산화제의개발이요청되기에이르렀다. 천연물의추출조건에대한연구로는추출조건에따른삼백초추출물의항산화활성및성분변화, 1 추출조건에따른우롱차의이화학적품질특성에관한연구, 12) 쌍화추출물의추출조건에따른품질특성에관한연구, 13) 용매와추출조건이계피추출액의항산화성에미치는영향등다양한천연물의추출조건에대한연구들이보고되었다. 14) 한편, 싸리, 15) 359

360 S.-J. Kim and D.-H. Oh 천마, 16) 오미자, 17) 산수유, 18) 국내산생약추출물 19) 및한약재 20) 등의추출물에대한항산화활성에대한연구가보고되었지만삼지구엽초에대한추출조건및삼지구엽초주성분인 icariin 함량별추출조건에대한연구는전혀보고되지않았다. 따라서, 본연구에서는 icariin 함량별삼지구엽초를이용하여물과에탄올로추출시간, 추출농도및추출온도를달리하여 icariin 함량별최적의추출조건및항산화활성을검토하였다. 재료및방법 시료본실험에사용한삼지구엽초 (Epimedium koreanium Nakai) 는경기도농업기술원에서재배되고있는삼지구엽초를 icariin 함량의차이에따라 KE9405(icariin 함량 0.20%) 와 KE9412(icariin 함량 0.59%) 품종을제공받아본실험에사용하였다 2. 추출, 수율및분획건조된삼지구엽초를마쇄하여분말화한후시료중량 10배의증류수를사용하여열수추출에서는 70 o C 90 o C 에서 3, 7시간동안 2회환류추출하였고, 에탄올추출은 (50%, 70%, 95%) 각각 50 o C 70 o C에서 3과 7시간동안 2회환류추출하여감압여과장치에서뜨거운상태로여과한다음감압농축기를사용하여농축후동결건조기로건조시킨다음본실험에사용하였다. 각조건에서얻어진삼지구엽초추출물의총추출수율은초기시료량에대한고형분의양을백분율로계산하였다. 한편, 70% 에탄올로 70 o C에서 7시간추출하여얻은각시료의농축물을용매의극성에따라분별분리를행하여 hexane, chloroform, ethyl acetate, butanol 및물층으로극성의차이에의해순차적으로용매분획한다음각분획물을농축하여동결건조하였다. 또한, 각분획물의수율은 ethanol 추출물의동결건조중량에대한각유기용매추출물의동결건조량의조성비로각각나타내었다. DPPH free radical 소거법을이용한항산화효과 1,1-diphenyl-2-picrylhydrazyl(DPPH) 를이용한항산화활성검정법으로여러농도의시료를 4 ml의메탄올에녹여 1.5 10-4 M DPPH methanol 용액 1 ml를첨가한후 30분간실온에방치하여 517 nm에서흡광도를 3회반복하여측정한뒤평균을내어시료를첨가하지않은대조군의흡광도를 1/2로감소시키는데필요한시료의양 (µg) 을 reduction concentration(rc 50 ) 으로나타냈으며, 기존의항산화제인 α-tocopherol과 ascorbic acid를비교하였다. 통계처리본실험은 3회반복실시하였고그값을 SPSS Ver. 10.0 package program 22) 을이용하여각시험구의평균과표준편차를산출한후각시험구간의차이유무를 ANOVA 로분석한뒤 a=0.05에서유의적차이를 Turkey법으로검증하였다 23). 결과및고찰 추출조건별수율측정삼지구엽초의 icariin 함량에대하여추출용매, 추출온도, 추출시간등의추출조건에따른수율을측정한결과는 Table 1 및 2와같다. 열수추출의경우 (Table, icariin 함량별로나타난결과 KE9412에서추출수율이가장높았으며, 추출온도는 70 o C보다 90 o C에서수율이높게측정되었고, 추출시간은 3시간보다 7시간추출했을때더높게나타났다. 삼지구엽초의 icariin 함량에따른추출조건은 KE9405와 KE9412 모두 90 o C에서 7시간추출시 22.0% 및 27.5% 로추출온도가높아질수록, 추출시간이길어질수록수율이높게나타났다. 에탄올추출 (Table 2) 도열수추출과마찬가지로 KE9412에서가장높은추출수율을나타났다. 에탄올농도별로는 70% 에탄올에서 KE9405와 KE9412 모두 30.1% 와 39.3% 로가장높은함량을나타내었으며, 100% 에서는 11.8% 와 18.5% 로가장낮은수율을나타내었다. 이러한결과는삼백초, 결명자를이용한추출수율조건에대한연구에서 50~70% 에탄올농도에서높은함량을나타내었다는결과와유사하였다. 11,24) 에탄올추출물은열수추출물과달리온도와시간은고형분의함량에크게영향을주지않았으나에탄올농도에는많은영향을받는것으로나타났다. 100% 에탄올에서는 50% 와 70% 에탄올보다현저하게수율이감소된것으로나타났는데이는 Kim 등 25) 의계피추출액의유효성분과가용성고형분의용출량에미치는추출용매및추출시간등의추출조건을연구한결과에서 100% 에탄올로추출한것보다는 70% 에탄올로추출한추출액의가용성고형분함량이더높은수치를나타냈다는결과와일치하였다. 열수추출보다에탄올추출에서더많은수율이측정된것은삼지구 Table 1. The water extraction yields of Epimedium koreanum Nakai Yields(%) Tem. ( o C) 70 90 Time (h) 3 7 3 7 L * 18.0±0.17 b1,2) 19.8±0.11 a 18.4±0.07 a 22.0±0.21 a H * 16.9±0.08 a 23.4±0.39 b 26.9±0.14 b 27.5±0.09 b Means with different letters in the same column are significantly different (p<0.05). 2) Value=mean±S.D.

Icariin Epimedium koreanum Nakai Antioxidant 361 Table 2. The ethanol extraction yields of Epimedium koreanum Nakai Yields(%) EtOH (%) 50 70 100 Tem. ( o C) 50 70 50 70 50 70 Time (h) 3 7 3 7 3 7 3 7 3 7 3 7 KE9405 29.1±0.09 a1,2) 22.2±0.21 a 25.4±0.26 a 28.3±0.17 b 28.4±0.06 a 23.5±0.17 a 22.1±0.21 a 30.1±0.12 a 12.8±0.08 a 18.5±0.06 a 17.4±0.17 b 11.8±0.10 a KE9412 35.5±0.26 b 27.5±0.14 b 37.3±0.09 b 26.3±0.33 a 39.3±0.08 b 28.1±0.17 b 35.2±0.07 b 33.5±0.15 b 18.5±0.09 b 22.7±0.14 b 16.6±0.21 a 19.5±0.09 b Means with different letters in the same column are significantly different (p<0.05). 2) Value=mean±S.D. 엽초추출물중친수성물질보다소수성물질이더많이함유되어있는것으로사료되어진다. DPPH free radical 소거법을이용한항산화활성일반적으로특정물질에대한항산화활성을측정하는것은여러방법이있으나 spectrophotometer를이용한 DPPH radical 소거활성법이간단하며대량으로측정이가능하여널리이용되고있다. 이물질은 radical을갖는물질중에서비교적안정한화합물로 MeOH 용액에서는보라색으로발색된다. 그러나항산화활성을갖는물질을만나면항산화활성물질이 DPPH radical을포획하기때문에보라색이소실된다. 따라서미지의식물추출물의항산화활성을쉽게측정할수있으며실제항산화활성과도연관성이매우높은장점이있다. Icarrin 함량별삼지구엽초의각추출물의 free radical 소 거능 RC 50 ( 산화를 50% 억제시키는데요구되는시료의양 ) 은그수치가낮을수록 free radical 소거능 ( 항산화활성 ) 이좋으며그결과는 Table 3 및 4와같다. Icariin 함량별열수추출물의항산화활성은온도에의한영향은대체로 70 o C보다 90 o C에서더높게나타났으며, 시간에의한영향은온도에따라다르게나타났는데 70 o C에서는 7시간추출물에서항산화활성이좋았고 90 o C에서는 3시간추출물에서항산화활성이좋게나타나장시간추출할때는낮은온도에서, 단시간추출할때는높은온도에서추출하는것이더바람직한것으로나타났다 (Table 3). 한편, Icariin 함량에따른항산화활성을비교해보면 KE9412 가 70 o C7시간에서 34.9(µg/mL), 90 o C3시간에서 28.4(µg/mL) 로같은추출조건에서 KE9405의 114.3(µg/mL) 및 83.1(µg/ ml) 에비하여각각 3.2배와 2.3배높은항산화활성을나타내었다. Table 3. Free radical scavenging effect of water extracts of Epimedium koreanum Nakai at various temperatures and hours RC 50 (µg/ml) Temperature( o C) 70 90 Time (h) 3 7 3 7 KE9405 54.3±0.55 a1,2) 114.3±0.33 b 83.1±0.14 c 561.2±0.21 c KE9412 57.2±0.39 b 34.9±0.26 a 28.4±0.10 b 39.1±0.17 b Ascorbic acid 19.5±0.09 a 19.5±0.09 a α-tocopherol 18.2±0.08 a 18.2±0.08 a Means with different letters in the same column are significantly different (p<0.05). 2) Value=mean±S.D. Table 4. Free radical scavenging effect of ethanol extracts of Epimedium koreanum Nakai at various temperatures and hours RC 50 (µg/ml) EtoH(%) 50 70 100 Tem. ( o C) 50 70 50 70 50 70 Time (h) 3 7 3 7 3 7 3 7 3 7 3 7 KE9405 43.2±0.13 c1,2) 44.1±0.08 c 70.1±0.10 c 36.3±0.25 c 93.4±0.17 c 32.1±0.05 c 35.3±0.19 c 36.2±0.36 b 87.4±0.28 c 42.1±0.55 c 168.1±0.25 c 58.1±0.20 c KE9412 22.3±0.16 b 22.2±0.05 b 29.2±0.09 b 22.1±0.18 b 23.2±0.19 b 22.0±0.10 b 27.1±0.28 b 18.9±0.12 a 30.2±0.13 b 23.0±0.23 b 26.2±0.21 b 26.4±0.10 b 1Ascorbic acid 19.5±0.09 a α-tocopherol 18.2±0.08 a Means with different letters in the same column are significantly different (p<0.05). 2) Value=mean±S.D.

362 S.-J. Kim and D.-H. Oh 에탄올추출물의항산화활성은에탄올농도가추출온도및추출시간보다더큰영향을미치는것으로나타났다 (Table 4). KE9405와 KE9412 모두 70% 에탄올농도에서각각 32.1(µg/mL) 와 18.9(µg/mL) 로강한항산화활성을나타냈다. 이러한결과는 Lee 등 26) 의천년초선인장추출물의항산화효과연구에서 70% 에탄올추출물에서의항산화활성 (49.5µg/mL) 보다높은결과이며, Lee 등 27) 의시판한약재애대한항산화활성연구에서항산화활성이높게측정된복분자 (15.26µg/mL), 초두구 (16.15µg/mL) 와비슷한결과를보이고있다. Icariin 함량과에탄올농도와의항산화활성과의관계를보면 KE9412가에탄올농도 50%, 70%, 100% 모두 KE9405보다높은항산화활성을나타낸결과를근거로볼때 icariin의항산화활성은농도의존성인것으로보인다. 또한, 기존천연항산화제로알려진 ascorbic acid(19.5 µg/ml) 나 α-tocopherol(18.2 µg/ ml) 와비교해도항산화활성이큰차이를나타내지않아삼지구엽초추출물을이용한천연항산화제의개발에도많은도움이될것이라사료된다. 한편, 삼지구엽초의항산화활성에관여하는물질을탐색하기위한기초자료를얻기위하여최적조건을나타낸 70% 에탄올 70 o C 7시간추출물로부터유기용매별로순차적으로분획한결과는 Table 5와같다. 추출수율은 KE9405와 KE9412 모두 butanol 분획층에서각각 36.2% 와 42% 로가장높았으며, 물분획층, ethylacetate 분획층의순으로높게나타났으며, hexane과 chloroform 분획층이각 9.3% 와 5.8% 로지용성물질이많이추출되는비극성용매계에서의수율은상대적으로적은편이었다. 한편, 삼지구엽초분획물의항산화활성은 butanol 분획층에서 39.2 µg으로가장높았으며, ethylacetate(49.0 µg), 물 (118.8 µg), heaxane(119.8 µg), chloroform(138.5 µg) 층으로나타났다. 반면에, 에탄올추출물은 KE9405와 KE9412에서각각 34.9 µg과 18.8 µg을나타내어추출물을더정제할수록오히려항산화활성은저하되는것으로나타나삼지구엽초의경우기능성식품 을개발할때정제물질보다는추출물을이용하는것이여러가지측면에서더바람직한것으로나타났다. 한편, Noh 등 28) 은삼지구엽초의경우증류수나 100% 에탄올보다 70% 에탄올에서더높은항산화활성을나타내었다고보고하였는데이는본연구결과와일치하며삼지구엽초의항산화활성을나타내는물질로 Kim 29) 은에탄올추출물에 DPPH와반응하기적합한입체구조를가지는 flavonoid 물질이존재하기때문으로사료된다고보고하였다. 또한, Iinuma 등 30) 은삼지구엽초의주성분인 icariin과 epimedin 이생쥐에서간의 Kupffer cell에의한탐식작용을활성화시킨다고하였으며, Liu 등 3 은삼지구엽초의메탄올추출물이생명력이결핍된환자에있어서백혈구수를유의하게증가시켜면역능을촉진시킨다고보고하였다. 삼지구엽초추출물의성분이대부분 flavonoid계물질로분석된것으로보아추후이와관련된연구가더심도있게검토되어야하며삼지구엽초추출물을이용한새로운기능성식품의개발이매우필요한것으로사료된다. 요 본연구는 Icariin 함량별로재배된삼지구엽초의추출조건및항산화활성을비교하기위하여수행하였다. 추출수율은열수추출에서는 90 o C에서 7시간, 에탄올추출에서는 70% 와 50 o C에서 3시간추출한경우가장높은수율을나타내었다. 열수추출보다에탄올추출이높은수율을나타내었으며에탄올추출농도는 100%( 추출수율 22.7%) 보다는 50% 와 70% 농도에서각각추출수율이각각 37.3% 와 39.3% 로높게나타났다. 또한, 에탄올추출시추출시간과추출온도보다는추출용매농도가추출수율에더많은영향을미치는것으로나타났다. Icariin 함량별추출수율은열수추출과에탄올추출모두 icariin 함량 0.5~0.6% 인대에서우수하게나타났으며 icariin 함량에따른항산화활성은 KE9412가 70 o C 7 시간에서 34.9(µg/mL), 90 o C 약 Table 5. DPPH free radical scavenging activities of each fraction Epimedium koreanum Nakai Extraction Yield(%) RC 50 ( μg /ml) solvant KE9405 KE9412 KE9405 KE9412 Hexane 29.3±0.08 b1,2) 10.8±0.19 b 228.3±0.09 119.8±0.19 Chloroform 25.8±0.26 a 26.6±0.10 a 470.3±0.20 138.5±0.16 Ethyl acetate 13.2±0.20 c 15.5±0.05 260.4±0.21 b 249.0±0.07 Butanol 36.2±0.19 e 42.0±0.34 288.0±0.21 239.2±0.11 b Water 18.4±0.09 d 23.0±0.08 270.7±0.12 122.0±0.15 Ascorbic acid 219.5±0.09 a 219.5±0.09 a α-tocopherol 218.2±0.08 a 218.2±0.08 a Means with different letters in the same column are significantly different (p<0.05) 2) Value=mean±S.D.

Icariin Epimedium koreanum Nakai Antioxidant 363 3시간에서 28.4(µg/mL) 로같은추출조건에서 KE9405의 114.3(µg/mL) 및 83.1(µg/mL) 에비하여각각 3.2배와 2.3 배높은항산화활성을나타내었다. 한편, 70% 에탄올 70 o C 7시간추출물로부터유기용매별로순차적으로분획한결과, 추출수율은 KE9405와 KE9412 모두 butanol 분획층에서각각 36.2% 와 42% 로가장높았고항산화활성도 butanol 분획층에서 39.2 µg으로가장높았으며, ethylacetate(49.0 µg), 물 (118.8 µg), heaxane(119.8 µg), chloroform(138.5 µg) 층으로나타났다. 감사의글 본연구는경기도농업기술원의연구비지원에의하여수행되었으며이에감사드립니다. 참고문헌 1. Lee, Y.G., Sohn, H.O., Lee, D.W., and Lim, H.B. : The effect of water-extract of Epimedium koreanum Nakai on ageralated change of the xenobiotic matabolizing enzyme system in the liver of rats. Korean J. Medicinal Crop Sci. 10, 29-36 (2002) 2. Noh, J.H., Kim, Y.J., Choi, K.J., Kim, S.W., Kim, S.K., and Kim, J.H. : Characteristics of seeding and rhizome propagation in Epimedium koreanum NAKAI. Korean J. Medicinal Crop Sci. 11(2), 155-160 (2003) 3. Kang, S.S., Shin, K.H., Chung, S.G., and Cho, E.H. : Flavonoids from Epimedium koreanum. Kor. J. Pharmacogn. 19, 93-98 (1998) 4. Xu, G.W., Xu, B.J., and Wang, M.T. : Isolation and identification of icariin and icariside I. Chin. Pharm. Bull. 22, 129-132 (1987) 5. Xu, S.C., Wang, Z.X., Wu, L.J.. Wang, N.B., and Chen, Y.J. : Isolation and identification do icariin and epimedoside A. Chin. Tard. Herb Drugs. 13, 9-15 (1982) 6. Hong, H. : Oriental materica medica. Oriental Healing Arts Institute, Taiwan, pp. 563 (1986) 7. Lee, M.K., Choi, Y.J., Sung, S.H., Shin, D.I., Kim, J.W., and Kim, Y.C. : Antihepatotoxic activity of icariin, a major constituent of Epimedium koreanum. Planta Med. 61, 523-526 (1995) 8. Miyase, T., Ueno, A., Takizawa, N., Kobayashi, H., and Oguchi, H. : Studies on the glycosides of Epimedium grandiflorum (II). Chem. Pharm. Bull. 35, 3713-3716 (1987) 9. Shin, K.H., Lim, S.S., Ahn, S.D., Kim, S.K., and Park, K.Y. : Difference in components of Epimedium koreanum in compliance with seasons and places of collection. Korean J. Medicinal Crop Sci. 4, 321-328 (1996) 10. Han, Y.H., Choi, B.R., Soh, H.S., Lee, S.J., Choi, Y.J., and Kim, S.Y. : In vitro plant regeneration for mass propagation of Epimedium koreanum Nakai. Korean J. Hort. Sci. & Technol. 18, 834-838 (2000) 11. Kim, S.K., Ban, S.Y., Kim, J.S., and Chung, S.K. : Change of antioxidant activity and antioxidant compounds in Saururus chinensis by extraction conditions. J. Korean Soc. 48, 89-22 (2005) 12. Park, S.K., Kim, J.K., Kim, J.H., Moon, K.D., and Oh, S.L. : Study on the characteristic of physicochemical quality of oolong herbs tea by extraction conditions. Korean J. Dietary Culture 9, 411-417 (1994) 13. Park, S.S., Lee, K.B., and Han, M.K. : Quality Characteristics of ssangwha extraction according to extraction conditions. Korean J. Food & Nutr. 17, 368-373 (2004) 14. Kim, N.M., Soon, H.S., and Kim, W.J. : Effect of Solvents and some extraction conditions on antioxidant activity in cinnamon extracts. Korean J. Food Sci. Technolol. 25, 204-209 (1993) 15. Lee, Y.S., Joo, E.Y., and Kim, N.W. : Antioxidant activity of extracts from Lespedeza bicolor. Korean J. Food Preserv. 12, 75-79 (2005) 16. Heo, J.C., Park, J.Y., An, S.M., Lee, J.M., Yun, C.Y., Shin, H.M., Kwon, T.K., and Lee, S.H. : Anti-oxidant and antitumor activities of crude extracts by Gastrodia elata Blume. Korean J. Food Preserv. 13, 83-87 (2006) 17. Jang, E.H., Pyo, Y.H., and Ahn, M.S. : Antioxidant effect of Omija (Schizandra Chinesis Baillon) extracts. Korean J. Food Sci. 12, 372-376 (1996) 18. Seo, K.I., Lee, S.W., and Yang, K.H. : Antimicrobial and antioxidative activities of Corni Fructus extracts. Korean J. Postharvest Sci. Technol. 6, 99-103 (1999) 19. Kim, H.K., Kim, Y.E., Do, J.R., Lee, Y.C., and Lee, B.Y. : Antioxidative activity and physiological activity of some korean medicinal plants. Korean J. Food Sci. Technol. 27, 80-85 (1995) 20. Nam, S.H. and Kang, M.Y. : Screening of antioxidative activity of hot-water extracts from medicinal plants. J. Korean Agric. Chem. Biotechnol. 43, 141-147 (2000) 21. 강승원, 최병렬, 박경열, 강창성, 안상득 : 삼지구엽초 ( 음양곽 ) 양질다수성품종육성. 시험연구보고서. 378-381 (1997) 22. SPSS. Statistical PaCkage for Social Sciences for Windows. Rel. 10.0. SPSS Inc., Chicago, IL, USA (1999) 23. Jung, C.Y. and Choi, L.G.: SPSSWIN for Statistics Analysis, Ver. 10.0, 4th ed. Muyok Publishing Co., Seoul, Korea, 276-283 (2002) 24. Kim, I.T., Kim, H.T., and Hwang, S.M. : Instant tea Preparation from Cassia tora seeds. Korean J. Food Sci. Technol. 22, 241-247 (1990) 25. Kim, N.M., Ko, S.R., Choi, K.J., and Kim, W.J. :Effect of some factors on extraction of effectual components in cinnamon extracts. J. Korean Agric. Chem. Soc. 36,17-22 (1993) 26. Lee, K.S., Oh, C.S., and Lee, K.Y. : Antioxidative effect of the fraction extracted from a cactus Cheonnyuncho (Opuntia humifusa). Korean J. Food Sci. Technol. 37, 474-478 (2005) 27. Lee, S.E., Seong, N.S., Park, C.G., and Seong, J.S. : Screening for antioxidative of oriental medicinal plant materials. Korean J. Medicinal Crop Sci. 10, 171-176 (2002)

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