치자 (Gardenia jasminoides Ellis fructus) 껍질추출물의항산화활성 진동혁 오다영 김한수 부산대학교식품공학과 (2017 년 2 월 7 일접수 : 2017 년 3 월 27 일수정 : 2017 년 3 월 30 일채택 ) Antioxidant Activity of Peel from Gardenia jasminoides Ellis Fructus Extracted by Various Solvents Dong-Hyeok Jin Da-Young Oh Han-Soo Kim Department of Food Science and Technology, Pusan National University, Miryang 50463, Korea (Received February 7, 2017; Revised March 28, 2017; Accepted March 30, 2017) 요약 : 치자껍질의항산화활성을측정하기위하여 70% methanol, distilled water (DW) 및 ethyl acetate (EA) 의 3 가지용매를사용한추출물의용매별 flavonoid 함량및항산화활성, 금속 chelating 능력측정을통하여치자껍질의기능성식품재료로서의가치를검토한결과, 치자껍질의 proanthocyanidin 함량은 48.165±0.811 mg CE/g DW 로나타났으며, 추출수율은 70% methanol (36.26%), DW (39.87%), EA (2.88%) 로나타났다. 추출용매별항산화활성은농도 (0.2, 0.4, 0.6 mg/ml) 가증가할수록유의적으로증가하였으며, control 로사용된 ascorbic acid, BHA, EDTA 보다는낮은활성이관찰되었다. Flavonoid 함량 (mg QE/g) 은 70% methanol (0.416), DW (0.225), EA (0.212) 순으로확인되었으며다른항산화분석및금속 chelating 능력실험에서도이와유사하게관찰되어모든분석에서 70% methanol 추출물이가장활성이강한것으로나타났다. 이상의결과를미루어볼때치자껍질의용매별 flavonoid 함량에따라항산화능, 금속 chelating 능력이증가하는것으로사료되며, EA 추출물을제외한다른추출물에서의수율과항산화활성은높은수준으로관찰되었다. 따라서치자의껍질은 proanthocyanidin 과 flavonoid 화합물을다량함유하고있으며그로인한높은항산화활성과생리활성을가지고있어기능성식품및천연항산화제로서의가치가매우기대된다. 주제어 : 치자껍질, 항산화활성, 프로안토시아니딘, 플라보노이드 Abstract : The purpose of this study was to measure the bioactivity and antioxidant activity of peel from Gardenia jasminoides Ellis fructus (GJE). GJE have been known to contain functional materials such as crocin, crocetin, geniposide, gardenosid, geniposidic acid, iridoid glycosides etc. We were separated into GJE peel. After that, we determined proanthocyanidin. GJE peel were extracted by 70% methanol, ethyl acetate (EA) and distilled water (DW) of three solvents. To Corresponding author (E-mail: kimhs777@pusan.ac.kr) - 163 -
2 진동혁 오다영 김한수 investigate by the solvent extract of flavonoid content and value as a functional food ingredient of GJE peel through antioxidant activities (DPPH radical scavenging activity, ABTS radical scavenging activity, superoxide dismutase like ability, hydroxyl radical scavenging activity, ferrous ion-chelating capacity) were performed. Solvent extract antioxidant activity of increasing concentrations (0.2, 0.4, 0.6 mg/ml) were significantly increased (p<0.05). GJE peel extracts showed lower activity than positive control (ascorbic acid, BHA, EDTA). These results, by a solvent of peel were found that the relationship with the increase of flavonoid content increased physiological activity. The antioxidant activity of the extract from the other except for the EA extract on peel was observed at a high level. The results suggest that GJE peel can be used as nutraceutical foods and natural antioxidant. Keywords : peel of Gardenia jasminoides Ellis fructus, antioxidant activity, proanthocyanidin, flavonoid 1. 서론 지난 50 년동안농업의발달로생산성이증가하여, 식품의다양성과계절의존성감소를통한공급량증가와그로인한가격안정화의결과식품소비에상당한변화를가져왔다 [1]. 서구화된식생활과스트레스, 환경, 신체활동감소등으로인한이상지질혈증 (dyslipidemia), 고혈압과같은심혈관계질환과비만, 당뇨등생활습관병발병이사회적문제가되고있으며 [2-3], 건강의중요성에따라식품은단순한영양원이아닌기능성을고려한건강기능식품에대한관심과소비가증가하는추세이다 [4]. 여러질병의주된원인으로인체내의호흡과정중발생하는산화생성물의 free radical 은 superoxide radical (O 2 -), singlet oxygen ( 1 O 2 ), hydroxyl radical (OH-), hydrogen peroxide (H 2 O 2 ) 등의 reactive oxygen species (ROS) 로다양하게존재하며, 불안정하고홑전자를가지므로그안정성을확보하기위해반응성이매우높은것으로알려져있다 [5]. 또한세포막파괴, DNA 손상등과함께여러산화에의한생성물들을생성하여암을유발하고노화와관련해생리적장애를일으킨다고알려져있다 [6]. 경남남해군의삼자중하나인치자는꼭두서니과 (Rubiaceae) 에속하는치자나무 (Gardenia jasminoides Ellis) 의열매로국내를비롯한중국, 일본, 대만등의기온이따뜻한지방에자생하고있다. 한방에서는소변을잘나오게하여열을제거하고화를가라앉혀피를맑게하며, 지혈, 진정, 염증완화등의작용이있다고한다. 또한 geniposide, β-carotene, crocin, quercetin, shanzhiside 등여러생리활성물질을함유하고있어이러한천연화합물들은체내에서 ROS 및 free radical 을제거할수있는능력과이에대한상호작용을통해생활습관병을예방하는것으로알려져있다 [7-8]. 그중항산화기능에대한 flavonoid 연구는 1980 년대초부터많은관심을받아왔으며 [9], Steinmetz & Potter (199 의연구 [10] 에서많은 flavonoid 를함유하고있는야채를섭취시만성질환을감소시키는것으로보고하였다. 식물체에서 5,000 개이상의다양한 flavonoid 가존재하는것으로알려져있으며, 국내외에서 flavonoid 가건강에미치는영향에대한연구가진행되고있다 [11-13]. 본연구에서는치자의껍질의 proanthocyanidin 함량을측정하여 70% methanol, distilled water (DW) 및 ethyl acetate (EA) 의용매별로추출한뒤, flavonoid 함량을측정하고, 항산화능력 (DPPH radical scavenging activity, ABTS radical scavenging activity, superoxide dismutase like ability, hydroxyl radical scavenging activity), 금속킬레이팅능력 (ferrous chelating activity) 을측정하여치자껍질의추출용매에따른항산화능력을비교하여생활습관병예방과건강기능식품개발의목적으로이들의소비효율성을높이고이에대한기초자료를제시함으로써고부가가치의천연항산화제의이용가능성을검토하고자하였다. - 164 -
치자 (Gardenia jasminoides Ellis fructus) 껍질추출물의항산화활성 3 2. 재료및방법 2.1. 실험재료본실험에사용된시료인치자 (Gardenia jasminoides Ellis) 는경상남도남해군에서 2014 년 11 월에수확하여반건조상태로되어있는시료를남해소재약재상에서 2015 년 1 월에구입하였다. 구입한치자의껍질을분리한뒤자연건조시켜분쇄기 (HMF-3250S, Han-Il Co., Seoul, Korea) 에각각분쇄하여분말로만든다음 deep freezer (DF-8514, Il-Shin BioBase Co., Daegu, Korea) 에 80 로저장하여실험에사용하였다. 2.2. Proanthocyanidin 함량측정치자껍질의 proanthocynidin 함량은 vanillin 측정법을변형하여측정하였다 [14]. 시료분말 0.03 g 에 methanol 2.0 ml 를넣어혼합한후여과 (filter paper, Advantec, No.2, Tokyo, Japan) 한뒤 1% vanillin/methanol (w/v) 2.0 ml 를넣고 25% sulfuric acid/methanol 2.0 ml 를가하여섞은후 30 water bath 에서 15 분간방치한뒤 methanol 1.0 ml 를넣고 4,000 rpm 에서 10 분간원심분리한후상등액을취하여 500 nm 에서흡광도를측정하였다. 표준물질로는 (+)-catechin (Sigma-Aldrich, St. Louis, USA) 을사용하였으며검량선을작성하여치자껍질에서의 proanthocyanidin 함량을산출하였고시료 g 당 mg CAE (mg of catechin equivalents) 로나타내었다. 2.3. 시료의추출시료의추출은동결저장된치자분말 100 g 씩취하여 70% methanol, ethyl acetate (EA) 용매를각 10배가하여 (1:10, w/v) 24시간씩 2 회추출한뒤여과 (filter paper, Advantec, No.2, Tokyo, Japan) 하였고, distilled water (DW) 추출물은 70 에서 2시간씩 2회추출하여여과 (filter paper, Advantec, No.1, Tokyo, Japan) 하였다. 각추출물은 rotary vacuum evaporator (Hei-VAP Advantage, Heidolph Co., Germany) 를이용하여 40 에서감압농축하여용매를제거한후, 실험에사용하였으며, 시료의수율은추출전시료중량에대한추출후건조중량백분율 (%) 로나타내었다. 2.4. Flavonoid 함량측정 Flavonoid 함량측정은 two complementary colorimetric method 를변형하여사용하였으며 [15], 시료추출액 0.5 ml 에 10% aluminium nitrate 0.5 ml 와 1 M potassium acetate 0.5 ml 를넣은뒤, 80% ethanol 2.0 ml 를첨가하여잘혼합한후 40 분간실온에방치하여반응시킨후 415 nm 에서흡광도값을측정하였다. 이때표준물질인 quercetin (Sigma-Aldrich, St. Louis, USA) 을사용해검량선을작성하여시료 g 당 mg QE (mg of quercetin equivalents) 로계산하였다. 2.5. DPPH radical scavenging activity 측정 DPPH (1,1'-diphenyl-2-picrylhydrazyl) radical 소거활성은각용매별시료추출물 0.2 ml 와 0.2 mm DPPH in 80% methanol 2.8 ml 를혼합하고암실에서 30 분간반응시킨후 517 nm 에서흡광도를측정하여백분율로나타내었으며, 그에따른 IC 50 계산하여표시하였다 [16]. 이때활성비교를위하여 control 로합성항산화제인 BHA (butylated hydroxyanisole, Sigma-Aldrich, St. Louis, USA) 를사용하여같은방법으로흡광도를측정하였다. 2.6. ABTS radical scavenging activity 측정 ABTS cation decolorization assay 에의한방법 [17] 을변형하여 ABTS (2,2-azino-bis-3- ethylbenzothiazoline-6-sulfonic acid) radical 을이용한항산화력측정을시행하였다. 7 mm ABTS 와 2.45 mm potassium persulfate 를 1:1 (v/v) 의비율로섞어 16 시간동안암소에방치하여 ABTS radical 을형성시킨후, 735 nm 에서흡광도값이 0.70(±0.02) 이되도록 ethanol 로희석하였다. 희석된용액 3.9 ml 에시료추출액 0.1 ml 를넣은후 10 분뒤 734 nm 에서흡광도를측정하였다. Control 은 BHA 를사용하였으며 ABTS radical 소거활성은백분율로나타내었다. 2.7. Superoxide dismutase (SOD) like ability 측정 SOD 유사활성으로 McCord & Fridovich (1969) 의방법 [18] 을변형하여시료 0.1 ml 에 Tris-HCl buffer (50 mm tris (hydroxymethyl) aminomethane buffer + 10 mm EDTA (ethylenediaminetetraacetic acid disodium salt - 165 -
4 진동혁 오다영 김한수 dihydrate (ph 8.5)) 3.0 ml 와 7.2 mm pyrogallol 0.2 ml 를넣어 25 water bath 에서 10 분동안반응시킨뒤, 1.0 N HCl 1.0 ml 로반응을정지시켜 420 nm 에서흡광도를측정하였다. SOD 유사활성은나타내어계산하였으며이를통해 IC 50 을구하였다 control 로 BHA 를사용하여나타내었다 2.8. Hydroxyl radical (OH-) scavenging activity 측정 Hydroxyl radical 소거능측정은 deoxyribose degradation assay [19] 와 1,10-phenanthroline/ ferrous iron (II) oxidation assay [20] 를변형하여사용하였다. 각농도의시료 1.0 ml 를 0.75 mm 1,10-phenanthroline 용액과 ethanol 1.0 ml 및 0.75 M PBS (phosphate buffer saline, ph 7.4) 2.0 ml, distilled water 1.0 ml, 0.75 mm ferrous sulfate 1.0 ml, 0.032% hydrogen peroxide 1.0 ml 를순차적으로가하여 37 water bath 에 60 분간반응시켰다. Control 로 BHA 를사용하여표시하였고, 흡광도는 536 nm 에서측정하여백분율로나타내었다. 2.9. Ferrous ion-chelating capacity 측정치자껍질의각농도별추출물에서 ferrous ion chelating activity 는 Robu et al.(2012) 의방법 [21] 을변형하여시료 1.0 ml 에 2 mm iron (II) chloride tetrahydrate 0.1 ml 과 5 mm ferrozine (3-(2-pyridyl)-5,6-diphenyl-1,2,4-triazine-4`,4 ``-disulfonic acid sodium salt) 0.2 ml, ethanol 3.0 ml 을가하여잘섞은후실온에 10 분간반응시켜 Fe2+-ferrozine complex 를형성시킨뒤 562 nm 에서흡광도를측정하였다. Control 로 EDTA (ethylenediaminetetraacetic acid disodium salt dihydrate, Sigma-Aldrich, St. Louis, USA) 를사용하였고치자껍질에서의 ferrous ion-chelating activity 는백분율로환산하여나타내었다. 2.10. 통계처리실험데이터는 3 회반복측정하였으며, mean±sd (n=3) 으로표현하였다. 또한실험결과값간의유의적인차이는 one-way ANOVA (analysis of variance) 로분석한뒤 p<0.05 수준에서 Duncan`s multiple range test 에의하여각농도간의유의성을검증하였다. 통계처리에대한 프로그램은 IBM SPSS statistic ver. 22 를사용하였다. 3. 결과및고찰 3.1. Proanthocyanidin 함량치자껍질의 proanthocyanidin 함량은 Table 1 에나타내었으며 48.165±0.811 mg CE (catechin equivalent)/g DW (dry weight) 로관찰되었다. Proanthocyanidin 은식물에자연적으로존재하는 flavonoid polymer complex 로서식물의씨앗, 포도와같은베리류를포함한일부과일과껍질에풍부하며구조적인특징으로 polyphenol 을가지고있어인체내에서금속이온및단백질과복합체를형성하며 [22-23], 안정적인환원제로작용하여강력한항산화작용을가지고있는것으로알려져있다 [24]. Carpenter et al.(2014) 는슈퍼푸드로알려진크랜베리의 proanthocyanidin 함량이재배지마다다르지만 Mullica Queen 과 Early Black 품종에서 48~82 mg/g 의 proanthocyanidin 이함유되어있다고보고하였다 [25], 이에본실험에서의치자껍질은 proanthocyanidin 의함량이많다고알려진크랜베리와비교하였을때 proanthocyanidin 함량이비슷한수준으로판단되어이로인한생활습관병예방및노화방지등의효과가기대된다. 3.2. 수율치자껍질의 70% methanol, distilled water (DW) 및 ethyl acetate (EA) 에서의추출수율은 Table 1 에나타내었다. 각용매별추출수율은 70% methanol 에서 36.26%, DW 39.87%, EA 2.88% 로 DW 에서추출수율이가장높게나타났다. 3.3. Flavonoid 함량치자껍질의용매별추출물에서의 flavonoid 함량은 Table 1 에나타내었으며, 추출물중 70% methanol 추출물에서 0.416±0.002 mg QE/g 으로유의적으로가장높게나타났으며, DW 추출물 0.225±0.002 mg QE/g, EA 추출물 0.212±0.002 mg QE/g 순으로 EA 추출물에서가장낮은값이관찰되어각추출물에서유의적인차이가나타났다. Chu et al.(2000) 의연구에서는양배추, 시금치, 양파를포함한여러가지 - 166 -
치자 (Gardenia jasminoides Ellis fructus) 껍질추출물의항산화활성 5 Table 1. Contents of proanthocyanidin, flavonoid and IC 50 values in the antioxidant activity evaluation assays of peel from Gardenia jasminoides Ellis fructus Assays Values Proanthocyanidin content (mg CE 2) /g DW 3) ) 48.165±0.811 EA DW 70% methanol Extraction yields (%) 2.88 39.87 36.26 Flavonoid content (mg QE 4) /g) 0.212±0.002 a5) 0.225±0.002 b 0.416±0.002 c DPPH (IC 50, mg/ml) 3.585±0.011 c 0.940±0.001 b 0.610±0.001 a ABTS (IC 50, mg/ml) 1.929±0.027 c 1.640±0.005 b 1.208±0.004 a SOD (IC 50, mg/ml) 2.013±0.026 b 2.122±0.006 c 1.554±0.006 a OH- (IC 50, mg/ml) 3.698±0.037 c 0.949±0.001 b 0.636±0.001 a FIC (IC 50, mg/ml) 1.763±0.018 c 0.405±0.004 b 0.209±0.002 a DPPH radical scavenging activity (DPPH), ABTS radical scavenging activity (ABTS), superoxide dismutase like ability (SOD), hydroxyl radical scavenging activity (OH-), ferrous ion-chelating capacity (FIC). 2) CE: catechin equivalents. 3) DW: dry weight. 4) QE: quercetin equivalents. 5) The values are means±sd (n=3). Values with the different letters in the same row are significantly different (p<0.05) by Duncan`s multiple range tests. 채소류에서의 flavonoid 함량은 185.01~426.82 mg/kg 로나타났다고보고하였으며, 국화 52.19 mg/100 g (Andarwulan et al., 2010), 양파 486 mg QE/kg, 케일 110 mg QE/kg, 브로콜리 30 mg QE/kg 으로보고하였다 [26]. 이에치자껍질의 flavonoid 함량은높은수준으로판단되며그에따른생리활성효과가기대된다. 결과치가관찰되었다. 3.4. DPPH radical scavenging activity 치자껍질의각용매별추출물과 control 인 BHA (butylated hydroxyanisole) 의 DPPH radical 소거능을각농도별로비교한결과는 Fig. 1 에나타내었고, IC 50 값을구하여 Table 1 에나타내었다. 치자껍질의각용매별추출물 0.2, 0.4, 0.6 mg/ml 의농도에서측정한결과, 농도가증가함에따라유의적으로 DPPH radical 소거능이증가하는경향을보였다. 70% methanol 추출물에서농도별로각각 18.85±0.26%, 33.51±0.07%, 49.42±0.33%, IC 50 0.610±0.001 mg/ml 로추출용매중유의적인차이를보이며가장강한소거능이관찰되었으며, DW 추출물에서 12.71±0.07%, 23.13±0.33%, 32.81±0.36%, IC 50 0.940±0.001 mg/ml, EA 추출물 5.69±0.12%, 8.47±0.07%, 10.92±0.25%, IC 50 3.585±0.011 mg/ml 순으로추출물중에서는 EA 용매추출물에서가장낮은활성과높은 IC 50 Fig. 1. DPPH radical scavenging activity of various solvent extracts from peel of Gardenia jasminoides fructus. The values are means±standard deviation (n=3). Bars with the different letters are significantly different (p<0.05) by Duncan s multiple range 2) 3) tests. EA: ethyl acetate. DW: 4) distilled water. BHA: butylated hydroxyanisole. DPPH 는안정된 free radical 을가지며수소공여를통해산화되어소거되며이과정에서진한보라색에서황색으로정색반응하는것으로알려져있다 [27]. 치자용매별추출물의항산화능은 - 167 -
6 진동혁 오다영 김한수 positive control 인 BHA 보다낮았으나 70% methanol 추출물에서비교적으로높은활성이관찰되었으며, 용매별로 DPPH 소거능의명확한차이를보여사용된추출용매의극성에의해생리활성물질이용해되는정도가달라항산화능의차이가나타난다는보고와유사한것으로나타났다 [28]. 3.5. ABTS radical scavenging activity 치자껍질의각용매별추출물과 positive control 인 BHA 의 ABTS radical 소거능을각농도별로비교한결과를 Fig. 2 에표시하였으며, IC 50 값을구하여 Table 1 에나타내었다. 각시료는농도가증가함에따라 ABTS radical 소거능이유의적으로증가하는것으로관찰되었다. 70% methanol 추출물에서 24.20±0.01%, 29.66± 0.17%, 34.39±0.47%, IC 50 1.208±0.004 mg/ml 로추출용매중유의적인차이를보이며가장강한소거능이관찰되었으며, DW 추출물에서 23.76±0.15%, 27.17±0.22%, 31.07±0.15%, IC 50 1.640±0.005 mg/ml, EA 추출물 15.61±0.08%, 19.02±0.08%, 23.61±0.25%, IC 50 1.929±0.027 mg/ml 순으로 EA 용매추출물에서가장낮은 ABTS 소거활성과높은 IC 50 결과치가관찰되었다. 3.6. Superoxide dismutase (SOD) like ability 치자껍질추출물의 superoxide dismutase (SOD) 유사활성은 Fig. 3 에표시하였으며, IC 50 을구하여 Table 1 에나타내었다. 껍질의 70% methanol 추출물 20.51±0.01%, 25.17±0.14%, 29.19±0.40%, IC 50 1.554±0.006 mg/ml, DW 추출물 20.14±0.12%, 23.05±0.19%, 26.37± 0.12%, IC 50 2.122±0.006 mg/ml, EA 추출물 19.44±0.07%, 22.34±0.07%, 26.20±0.07%, IC 50 2.018±0.026 mg/ml 순으로 EA 용매추출물에서가장낮은 SOD 유사활성이나타났으나, DW 추출물에서유의적으로가장높은 IC 50 결과치가관찰되었다. 이는농도가증가함에따라 SOD 유사활성증가폭이 EA 추출물에서 DW 추출물보다크기때문에이러한결과가나타난것으로사료된다. 또한 EA 추출물과 DW 추출물은 0.6 mg/ml 농도에서유의적인차이가없는것으로관찰되었다. Positive control 인 BHA 는각농도에서 28.78±0.72%, 47.34±0.26%, 65.57±0.19% 로농도가증가함에따라 SOD 유사활성이큰폭으로증가하는것을관찰할수있었다. Fig. 2. ABTS radical scavenging activity of various solvent extracts from peel of Gardenia jasminoides fructus. The values are means±standard deviation (n=3). Bars with the different letters are significantly different (p<0.05) by Duncan s multiple range 2) 3) tests. EA: ethyl acetate. DW: 4) distilled water. BHA: butylated hydroxyanisole. Fig. 3. Superoxide dismutase like ability of various solvent extracts from peel of Gardenia jasminoides fructus. The values are means±standard deviation (n=3). Bars with the different letters are significantly different (p<0.05) by Duncan s multiple range 2) 3) tests. EA: ethyl acetate. DW: 4) distilled water. BHA: butylated hydroxyanisole. - 168 -
치자 (Gardenia jasminoides Ellis fructus) 껍질추출물의항산화활성 7 3.7. Hydroxyl radical (OH-) scavenging activity 치자껍질의각용매별추출물과 control 인 ascorbic acid 의 hydroxyl radical (OH-) 소거능을각농도별로비교한결과는 Fig. 4 에나타내었고, IC 50 을구하여 Table 1 에나타내었다. 70% methanol 추출물에서농도별로 20.02±0.53%, 33.12±0.07%, 47.75±0.30%, IC 50 0.636± 0.001 mg/ml 로추출물중가장높은활성이관찰되었으며, DW 추출물에서 13.98±0.07%, 23.57±0.30%, 33.23±0.33%, IC 50 0.949± 0.001 mg/ml, EA 추출물 7.56±0.13%, 10.08±0.07%, 12.41±0.13%, IC 50 3.698± 0.037 mg/ml 순으로 EA 용매추출물에서가장낮은활성과높은 IC 50 결과치가관찰되었다. Control 인 ascorbic acid 는각농도에서 93.58± 0.01%, 93.93±0.20%, 95.03±0.18% 로강한 hydroxyl radical 소거능이관찰되었다. 본실험에서의용매별추출물의 DPPH 소거능을포함한다른항산화능분석실험의결과를비교하였을때, 같은시료에서의항산화활성결과의정도의차이는있지만그결과는매우유사하다는 Elfalleh et al.(2009) 의보고 [29] 와동일하게각실험에서 70% methanol, DW, EA 추출물순으로항산화능이관찰되었다. 3.8. Ferrous ion-chelating capacity 치자껍질의각용매별추출물과 control 인 EDTA (ethylenediaminetetraacetic acid) 의각농도별로비교한 ferrous ion-chelating capacity 결과는 Fig. 5 에나타내었으며, IC 50 을구하여 Table 1 에나타내었다. 70% methanol 추출물 48.37±0.19%, 64.79±0.06%, 75.04±0.15%, IC 50 0.209±0.002 mg/ml 로유의적인차이를보이며강한 chelating 능력이관찰되었으며, DW 추출물에서 40.10±0.23%, 48.45±0.10%, 60.69±0.06%, IC 50 0.405±0.004 mg/ml, EA 추출물 30.17±0.09%, 33.87±0.18%, 35.14± 2.24%, IC 50 1.763±0.018 mg/ml 순으로관찰되었다. 본실험에서 control 로사용된 EDTA 는 6 자리의리간드구조를가지고있기때문에각농도에서 99% 이상의매우강한 ferrous ion-chelating 능력을나타낸것으로사료된다. Flavonoid 화합물의화학구조는각종산화작용의촉매역할을하는금속이온과결합하여산화반응을상당히줄일수있다는것으로알려져있다 [30]. 뿐만아니라 Kostyuk et al.(2004) 과 Mira et al.(2002) 의연구는 lutein, taxifolin, (-)-epicatechin 등 flavonoid 화합물과금속이온이결합한복합체가비복합체 flavonoid 보다더 Fig. 4. Hydroxyl radical scavenging activity of various solvent extracts from peel of Gardenia jasminoides fructus. The values are means±standard deviation (n=3). Bars with the different letters are significantly different (p<0.05) by Duncan s multiple range 2) 3) tests. EA: ethyl acetate. DW: distilled water. 4) AA: ascorbic acid. Fig. 5. Ferrous ion-chelating capacity of various solvent extracts from peel of Gardenia jasminoides fructus. The values are means±standard deviation (n=3). Bars with the different letters are significantly different (p<0.05) by Duncan s multiple range 2) 3) tests. EA: ethyl acetate. DW: 4) distilled water. EDTA: ethylenediaminetetraacetic acid. - 169 -
8 진동혁 오다영 김한수 뛰어난 radical 소거능을가진다고보고하였다 [31-32]. 이에 70% methanol 추출물 0.6 mg/ml 농도의시료는 70% 이상의높은 ferrous chelating 능력을가지는것으로관찰되어, 치자껍질추출물의금속이온결합및그에따른항산화작용에대한시너지효과가기대된다. 4. 결론 치자껍질의 proanthocyanidin 함량을알아보고 70% methanol, distilled water (DW) 및 ethyl acetate (EA) 의 3 가지용매를사용한추출물의용매별 flavonoid 함량및항산화활성, 금속 chelating 능력측정을통하여치자껍질의기능성식품재료로서의가치를검토하기위하여본실험을수행하였다. 치자껍질의 proanthocyanidin 함량은 48.165±0.811 mg CE/g DW 로나타났으며추출수율은 70% methanol (36.26%), DW (39.87%), EA (2.88%) 로나타났다. 추출용매별항산화활성은농도 (0.2, 0.4, 0.6 mg/ml) 가증가할수록유의적으로증가하였으며, control 로사용된 ascorbic acid, BHA, EDTA 보다는낮은활성이관찰되었다. Flavonoid 함량 (mg QE/g) 은 70% methanol (0.416), DW (0.225), EA (0.212) 순으로관찰되었으며다른항산화분석및금속 chelating 능력실험에서도이와마찬가지로관찰되어모든분석에서 70% methanol 추출물이가장활성이강한것으로나타났다. 이상의결과를미루어볼때치자껍질의용매별 flavonoid 함량에따라항산화능, 금속 chelating 능력이증가하는것으로사료되며, EA 추출물을제외한다른추출물에서의수율과항산화활성은높은수준으로관찰되었다. 본실험결과치자의껍질은 proanthocyanidin 과 flavonoid 화합물을다량함유하고있으며그로인한높은항산화활성과생리활성을가지고있어기능성식품및천연항산화제로서의가치가매우기대된다. References 1. J. Kearney, Food consumption trends and drivers, Philosoph. Transact. Royal Soc. B: Biol. Sci., 365, 2793 (2010). 2. S. Klein, D. B. Allison, S. B. Heymsfield, D. E. Kelley, R. L. Leibel, C. Nonas and R. Kahn, Waist circumference and cardiometabolic risk: a consensus statement from shaping America's health: Association for Weight Management and Obesity Prevention; NAASO, the Obesity Society; the American Society for Nutrition; and the American Diabetes Association, Obesity, 15, 1061 (2007). 3. T. A. Pearson, G. A. Mensah, R. W. Alexander, J. L. Anderson, R. O. Cannon, M. Criqui, Y. Y. Fadl, S. P. Fortmann, Y. Hong, G. L. Myers, S. C. Smith, K. Taubert, R. P. Tracy, F. Vinicor and N. Rifai, Markers of inflammation and cardiovascular disease application to clinical and public health practice: a statement for healthcare professionals from the centers for disease control and prevention and the American Heart Association, Circulation, 107, 499 (2003). 4. K. Menrad, Market and marketing of functional food in Europe, J. Food Eng., 56, 181 (2003). 5. B. Halliwell, J. M. Gutteridge and C. E. Cross, Free radicals, antioxidants, and human disease: where are we now?, J. Lab. Clinic. Med., 119, 598 (1992). 6. J. S. Hwang, B. H. Lee, X. An, H. R. Jeong, Y. E. Kim, I. Lee, H. Lee and D. O. Kim, Total phenolics, total flavonoids, and antioxidant capacity in the leaves, bulbs, and roots of Allium hookeri, Korean J. Food Sci. Technol., 47, 261 (2015). 7. H. J. Koo, K. H. Lim, H. J. Jung, and E. H. Park, Anti-inflammatory evaluation of gardenia extract, geniposide and genipin, J. Ethnopharmacol., 103, 496 (2006). 8. M. Yamauchi, K. Tsuruma, S. Imai, T., Nakanishi, N. Umigai, M. Shimazawa, and H. Hara, Crocetin prevents retinal degeneration induced by oxidative and endoplasmic reticulum stresses via - 170 -
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