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하선 함
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1 生活科學硏究第 21 輯 Journal of Human Ecology Vol. 21, 여주 (Momordica charantia.) 추출물이고지방식이를급여한 C57/BL6J 생쥐의체중변화및혈중지질에미치는영향 박지영 허영란 * 전남대학교대학원식품영양학과전남대학교생활과학대학식품영양학과 전남대학교생활과학연구소연구원 * Effects of Bitter melon (Momordica charantia.) extracts on Body weight change and Lipid composition in C57/BL6J Mice Fed High Fat Diet Ji-Young Park Young-Ran Heo* Department. of Food and Nutrition, Graduate School, Chonnam National University, Gwangju , Korea *Department. of Food and Nutrition, Colleg of Human Ecology, Human Ecology Research Institute, Chonnam National University, Gwangju , Korea < 목차 > Ⅰ. 서론 Ⅳ. 고찰 Ⅱ. 재료및방법 Ⅴ. 요약및결론 Ⅲ. 결과및고찰 참고문헌 Abstract This study was conducted to investigate the effects of Bitter melon(momorica charantia L.) extracts on anti-obesity and lipid profiles in C57/BL6J mice fed high fat diet. Mice were fed normal diet (NFD, normal control), and high-fat diet only (HFD, high fat control) and high-fat with bitter melon extract 5% (5% BHD), high-fat with bitter melon extract 10% (10% BHD) for 6 weeks. Body weight gains were significantly lower in the experimental groups, and FER was significantly lower in the experimental groups in a dose-dependent manner than HFD (p<0.05). Liver weight in 5% BHD was significantly lower and kidney weight is significantly lower in the experimental groups. Not only Adipose tissue weight - 1 -

2 but also body fat percentage was significantly lower in the experimental groups. The serum TG levels were decreased in the experimental groups and TC levels were significantly lower in the experimental groups in a dose-dependent manner than HFD (p<0.05). LDL-C levels were decreased in the 10% BMD. However, blood glucose levels were not significantly different among the groups. From the findings, it had shown that bitter melon extract have the anti-obesity effect and could improve lipid profiles. Kew words: bitter melon, momordica charantia L, anti-obesity, serum lipids, high fat diet Ⅰ. 서론 최근경제적풍요와함께서구화된식생활및즉석편의식품의섭취증가로인한성인병이증가하고있으며, 특히심혈관질환과암, 당뇨병등으로인한사망률의증가는사회적으로심각한문제이다 (Ministry of Health & Welfare 2003). 이러한질병증가의원인중가장큰비중을차지하는것은비만이다 국민건강통계에의하면, 체질량지수 (kg/m 2 ) 가 25 이상인성인비만인구가 31.9% 로 1998년도 25.8% 에비하여지속적으로늘어나고있음을보고하였다 (Ministry of Health & Welfare 2009). 비만은동물성지방의과다한섭취로인해혈액내콜레스테롤과중성지방을증가시켜성인병유발에중요하게작용한다. 따라서이러한혈중지질환경을개선시키는것이성인병을예방할수있는측면에서매우중요하며, 최근에는식물에함유된일부영양소, 식이섬유및식물성화합물 (phytochemicals) 이체지방축적을억제하는등의작용이있음이밝혀졌다 (Ahn 등 2005). 때문에이러한생리활성기능을갖는여러가지천연물에대한수요도가점차증가하고있는추세이며, 기능성식품소재를이용하여비만을비롯한성인병의예방및치료제의개발이점차가속화되고있다 (Najm & Lie 2010). 여주 (Momorica charantia L.) 는 1년생덩굴성박과식물로, 주로열대및아열대지방에서널리재배되고있으며원산지는인도동부또는동남아시아라고도알려져있지만확실치는않다 (Li 등 2007). 예로부터열매는식용으로, 종자는약용으로사용하여왔고열매의쓴맛때문에 bitter melon, bitter gourd, bitter cucumber라고부르며그모양으로인해 balsam pear, balsam apple 이라고도부른다 (Lee 2011). 6~8월에 열매를맺는여름야채로녹색이짙고혹같은돌기가총총하게많을수록쓴맛이강한데, 여주의쓴맛에는식물스테롤배당체들과많은종류의 amio acids, galacturonic acid, citurulin, pectin 등의성분이들어있으며, charantin, vicine, polypeptide-p 및 antioxidnants 등과같은생리활성물질이혈당강하기능을매개로항당뇨효과가탁월한것으로알려져있다 (Bzsch 등 2003; Zhong 등 2011). 특히여주의열매와종자에주로함유되어있는지용성물질인 charantin은췌장의 β세포에작용하여인슐린의분비를촉진함으로써혈당을낮춰주는역할을하는것으로보고되고있다 (Ali 등 1993; Reyes 등 2006; Park 등 2007; Ha & Park 2008). 뿐만아니라여주에비타민 C, 비타민 E 및카로틴등이풍부해항산화작용이있음은물론이고체내에서항암, 백내장발생방지, 관상동맥질환예방등의기능성이확인된이후항산화, 항균및항당뇨효과등에관한연구가지속적으로이루어져왔다 (Park 1989; Jung 등 2007; Pitchaon 등 2008). 이와같이여주가다양한약리적효능을지니고있는것으로알려졌음에도불구하고우리나라에서는주로정원에서관상용으로재배하여왔다. 한편지금까지보고된여주의약리적효과에대한연구결과들은실험동물, 추출용매, 식이종류, 첨가량및섭취기간등실험조건과여주의사용부위, 재배지, 숙성정도에따라서로상반된다양한결과가보고되어있다 (Grover 등 2004; Najm & Lie 2010). 이에본연구에서는우리나라에서자생하고있는여주를열수추출하여고지방식이와함께마우스에게급여하였을때체중증가와체지방축적억제, 혈중지질대사개선등에미치는영향을조사하여여주의기능성이강화된건강식품의개발을위한자료로활용하고자수행하였다

3 Ⅱ. 재료및방법 1. 실험재료및열수추출물의제조본실험에서사용한여주 (bitter melon) 는전라북도정읍시산야에서재배한 2010년산제품을이용하였으며, 건조오븐으로 30 에서 24시간에걸쳐건조시킨후분쇄한후 4 에서냉장보관하면서사용하였다. 열수추출물제조는분말상태인시료중량의 15배인용매 ( 열수 ) 를첨가하여 12시간동안추출한후, Whatman No.1 여과지를사용하여여과한뒤추출용매를농축한후동결건조하여실험에사용하였다. 2. 실험동물의사육및식이실험동물은 4주령의 C57BL/6J Mice를 ( 주 ) 중앙실험동물 (Seoul, Korea) 에서구입하여 2주일간일반식이로적응시킨후난괴법 (randomized complete block design) 에의해실험군당 7마리씩배치하여 4군으로분리하였다. 즉, 실험군은정상식이만섭취시킨정상식이군 (NFD), 고지방식이 (45%fat) 만섭취시킨고지방식이군 (HFD), 고지방식이에여주열수추출물을각각 5%, 10% 씩중량비 (weight basis) 로첨가해섭취시킨 5% BHD, 10% BHD군이다 (Table 1). 여주열수추출물의첨가량은선행연구결과 (Kar 등 12003; Grover & Yadav 2004) 를토대로설정하였다. 실험동물의식이는펠렛형태로제조하여 6주간공급하였고, 식이와식수는자유롭게섭취할수있도록공급하였다. 사육실의환경은항온 (23±2 ) 및항습 (55±5%) 조건을유지하였고, 조명은 12시간 light/ dark cycle로일정하게조절하였다. 3. 체중, 식이섭취량및식이효율 체중은 1주일에한번씩일정한시간에측정하였으며, 식이섭취량은매일일정한시간에공급하고남은식이를 4oC에서냉장보관한후 1주일에한번측정하였다. 식이효율 (food efficiency ratio, FER) 은실험기간동안의체중증가량을동일기간동안의식이섭취량으로나누어계산하였다. Body weight gain for experimental period (g) FER= Food intake for experimental period (g) 4. 실험동물의처리및시료수집 실험이종료된실험동물은희생전 24시간절식시키고, Diethyl ether를이용해마취시켜심장과정맥에서직접혈액을채취하였다. 채취한혈액은원심분리관에넣어 ice box에정치시킨다음 3000 rpm에서 10분간원심분리시킨후혈청을분리하였다. 혈액채취후, 장기위치를파악하여각장기조직 ( 간, 신장, 췌장 ) 및지방조직 ( 신장주변지방, 정소상체지방 ) 을적출하였다. 생리식염수에세척한다음여과지로물기를제거하고전자저울로무게를측정하였다. 무게를측정한즉시각장기와지방조직및혈청은 -70oC에서냉동보관하였다. 5. 혈청의지질및혈당농도분석 혈청중성지방 (triglyceride, TG) 과총콜레스테롤 (total cholesterol, TC) 및고밀도지단백- 콜레스테롤 (high density lipoprotein-cholesterol, HDL-C) 농도는각각의 Fuji-Chem slide를이용해혈액생화학 Table 1. Experimental design for animal study Group NFD HFD Treatment Normal fat diet (100%, Normal Control) High fat diet (100%, High Fat Control) 5% BHD High fat diet (95%) + Bitter melon extract (5%) 10% BHD High fat diet (90%) + Bitter melon extract (10%) NFD: AIN 76, HFD: 45% fat - 3 -

4 자동분석기 (Fuji Dri-Chem 3,500, Fujifilm, Tokyo, Japan) 로측정하였다. 저밀도지단백- 콜레스테롤 (Low density lipoprotein-cholesterol, LDL-C) 농도는 Freidewald식 {TC-HDL-C-(TG/5)} 에의하여계산하였다. 혈당농도는심장에서혈액을채취후바로전혈을사용하여혈당측정기 (Accu-Check, Roche, Germany) 로측정하였다. 6. 통계처리본실험의모든결과는 SPSS package program (version 18.0) 을이용하여통계분석하였으며, 분석결과는평균 ± 표준편차로나타내었다. 각실험군간의통계적유의성검정은일원배치분산분석 (one-way analysis of variance, ANOVA) 를실시하고, Duncan's multiple range test에의해 p<0.05 수준에서사후검정을실시하였다. Ⅲ. 결과및고찰 1. 증체량, 식이섭취량및식이효율고지방식이에여주열수추출물을첨가한식이가마 우스의체중, 증체량, 식이섭취량및식이효율에미치는영향은 Table 2와같다. 실험초기평균체중은 23.01±1.72 ~ 23.73±0.98으로실험군간에유의한차이가없었으나, 실험종료시체중은여주열수추출물첨가군이각각 5% BHD군 28.47±1.30g과 10% BHD 군 30.70±1.13g으로고지방식이대조군인 HFD군의 35.37±4.35g 보다유의적으로낮았다 (p<0.05). 식이섭취량은 HFD군에비하여 5% BHD군과 10% BHD군이유의적으로낮았으며 (p<0.05), HFD군에비하여 5% BHD군과 10% BHD군의식이효율이유의적으로낮았다 (p<0.05). 2. 장기조직무게실험동물을희생시킨후적출한장기조직의무게는 Table 3과같다. 장기중간의무게는고지방식이대조군에비해여주열수추출물 5% 첨가군이유의적으로낮았다. 신장의무게는고지방식이대조군에비해여주열수추출물첨가군모두에서유의적으로낮았다 (p<0.05). 정소의무게는정상식이대조군에비해고지방식이대조군에서유의적으로높았으며 (p<0.05), 여주열수추출물첨가군과는유의적인차이가없었다. Table 2. Effect of Bitter melon extract on weight gains, food intake and food efficiency ratio in mice (C57/BL6J) fed high fat diet Body weight(g) Group 1) Total food Initial Final Gains intakes(g) FER 2) NFD 23.01±1.72 NS3) 28.19±2.59 b4) 5.17±0.98 b ±0.34 b 0.03±0.01 c HFD 23.73± ±4.35 a 11.64±3.58 a ±0.60 a 0.07±0.02 a 5% BHD 23.71± ±1.13 b 6.99±1.40 b ±0.23 d 0.05±0.01 b 10% BHD 23.06± ±1.30 b 5.41±1.16 b ±0.19 c 0.03±0.01 c 1) See Table 1. 2) FER (food efficiency ratio)=weight gain(g/6week) / food intake(g/6week). 3) NS: not significant. 4) Mean±SD (n=7). Values within a column with different superscript letters are significantly different at p<0.05 by Duncan's - 4 -

Table 3. Effect of Bitter melon extract on organ weight in mice(c57/bl6j) fed high fat diet Group 1) Organ weight (g) Liver Kidney Testicle 4. 혈청지질과혈당농도 혈청지질농도는 Figure 1과같다.

02 ab 1) See Table 1. 2) Mean±SD (n=7). Values within a column with different superscript letters are significantly different at p<0.05 by Duncan's 3. 지방조직무게실험동물을희생시킨후적출한지방조직의무게는 Table 4와같다.

5 Table 3. Effect of Bitter melon extract on organ weight in mice(c57/bl6j) fed high fat diet Group 1) Organ weight (g) Liver Kidney Testicle 4. 혈청지질과혈당농도 혈청지질농도는 Figure 1과같다. 혈청 TG 농도는여주열수추추물첨가군이고지방식이대조군보다 NFD 1.01±0.14 ab2) 0.36±0.02 b 0.20±0.02 b HFD 1.16±0.30 a 0.42±0.06 a 0.24±0.03 a 5% BHD 0.92±0.13 b 0.35±0.04 b 0.23±0.03 ab 10% BHD 0.97±0.07 ab 0.36±0.03 b 0.21±0.02 ab 1) See Table 1. 2) Mean±SD (n=7). Values within a column with different superscript letters are significantly different at p<0.05 by Duncan's 3. 지방조직무게실험동물을희생시킨후적출한지방조직의무게는 Table 4와같다. 신장주변지방 (perirenal fat pad), 정소상체지방 (epididymal fat pad) 및총지방의무게는여주열수추추물첨가군이고지방식이대조군보다유의하게낮았다 (p<0.05). 총지방무게를체중으로나눈체지방률역시여주열수추추물첨가군이고지방식이대조군보다유의하게낮았다 (p<0.05). Fig. 1. Effect of Bitter melon extract on serum lipid in mice(c57/bl6j) fed high fat diet 1) See Table 1. 2) TG: Triglyceride, TC: total cholesterol, HDL-C: high-density lipoprotein-cholesterol, LDL-C: lowdensity lipoprotein-cholesterol, 3) Mean±SD (n=7). Values within a column with different superscript letters are significantly different at p<0.05 by Duncan's Table 4. Effect of Bitter melon extract on adipose tissue weight and body fat percentage in mice (C57/BL6J) fed high fat diet Group 1) Adipose tissue weight (g) Body fat percentage 2) Perirenal fat pad Epididymal fat pad Total fat pad NFD 0.13±0.11 b3) 0.54±0.30 c 0.67±0.34 c 0.02±0.96 c HFD 0.58±0.28 a 2.05±0.47 a 2.63±0.51 a 0.07±0.62 a 5% BHD 0.29±0.10 b 0.91±0.15 b 1.20±0.23 b 0.04±0.70 b 10% BHD 0.25±0.09 b 0.99±0.16 b 1.24±0.24 b 0.04±0.72 b 1) See Table 1. 2) Body fat percentage : Total fat pad weight / body weight 3) Mean±SD (n=7). Values within a column with different superscript letters are significantly different at p<0.05 by Duncan's - 5 -

6 Table 5. Effect of Bitter melon extract on serum lipid and glucose levels in mice(c57/ BL6J) fed high fat diet Group 1) Glucose (mg/dl) NFD 173.1±83.7 b2) HFD 351.6±38.5 a 5% BHD 300.5±59.8 a 10% BHD 288.7±90.2 a 1) See Table 1. 2) Mean±SD (n=7). Values within a column with different superscript letters are significantly different at p<0.05 by Duncan's 유의하게낮았다 (p<0.05). 혈청 TC 농도는고지방식이대조군보다여주열수추추물첨가군에서농도의존적으로유의하게낮았다 (p<0.05). 혈청 HDL-C 농도는고지방식이대조군보다여주열수추출물첨가군이유의하게낮았고, 혈청 LDL-C 농도는고지방식이대조군에비해여주열수추출물 10% 첨가군이유의적으로낮았다. 혈당농도는 Table 5와같다. 혈당농도는고지방식이군과여주열수추출물간에유의적인차이가없었다. Ⅳ. 고찰 본연구는우리나라에서자생하고있는여주의건강기능성식품소재로의활용가능성에대한기초자료를제시하고자시도되었다. 본연구에서는일차적으로과숙여주를열수추출하여고지방식이와함께각각 5% 와 10% 첨가하여마우스에게급여하였을때, 체중증가와체지방축적및혈중지질대사에미치는영향을조사하였다. 본연구결과에서는여주열수추출물의첨가로증체량은대조군에비하여유의하게감소하였으며, 열수추출물첨가농도에비례하여증체량이감소하는경향이었으나통계적인유의성을보이지않았다. 식이섭취량을비교한결과여주첨가군의식이섭취량은대조군에비하여유의하게감소하였으나, 5% BHD 군이 10% BHD군보다더적었으며, 결과적으로식이 효율이여주첨가농도에비례하여농도의존적으로감소하는결과를나타내었다. 그동안보고된여주의약리적효과에대한결과들은실험조건즉추출용매, 섭취량과섭취기간, 여주의숙성정도, 실험동물, 식이종류등에따라상반된다양한결과가보고되어있어본연구결과와직접비교하기는어렵다. 예를들어 Zhong 등 (2011) 은수컷 C57BL/6J 마우스에게 1.2% 여주수용액추출물을 58% 고지방식이에첨가하여 12주동안급여한결과, 식이섭취량은양군에사이에차이가없었으나, 증체량은대조군에비하여유의하게적어체중감량효과가있다고보고하였다. 반면 Gamarallage 등 (2004) 은 Sprague-Dawley rat를대상으로정상식이 (AIN 76) 에여주메탄올추출물 3% 첨가하여 2주동안섭취시킨결과, 대조군에비하여식이섭취량및증체량에유의한차이가없다고보고하였다. 따라서본연구에서 5% BHD군의식이섭취량이현저하게낮은점, 그럼에도불구하고체중증가량은높았던점등의원인과관련기작에대한추가연구가필요할것으로생각된다. 여주열수추출물섭취에다른장기무게는체중변화결과와유사하였으며, 체지방량및체지방률변화또한체중의변화와유사하였다. 즉대조군에비하여여주열수추출물급여한군이유의하게감소하였으며, 5% BHD군과 10% BHD군사이에는유의한차이가없었다. Chan 등 (2005) 과 Chen 등 (2003) 은정상식이및고지방식이 (30% fat) 와함께동결건조한여주쥬스가루를각각 0.35%~1.5% 의다양한수준에서첨가하여 7주간 Sprague-Dawley rat에게섭취시킨결과, 농도의존적으로섭취식이의에너지효율과내장지방축적도를감소시킴을보고하였으며, 관련기작으로지방산화효소활성과 uncoupling 단백질발현증가와연관되어있음을제시한바있다. 따라서본연구결과로볼때여주열수추출물섭취는체지방축적을억제하는데도움을줄수있을것으로생각된다. 그러나상기연구와비교해볼때실험에사용된여주의첨가량및처리방법에크게차이가있으므로, 본연구에서사용한 5% 보다낮은농도의열수추출물첨가시의효과및추출방법을달리했을때의결과에대한추가연구비교가필요할것으로생각된다

7 본연구에서의혈액에서의지질농도들을볼때, 고지방식이대조군의지질농도들과비교하여여주열수추출물을첨가한군들에서유의하게감소하였다. 이로써고지방식이로인하여증가된중성지질함량이여주열수추출물을병행투여함으로써혈청지질환경이개선되어짐을확인할수있었다. Jeong 등 (2008) 의연구에서혈당이 350mg/dl 이상인 C57BL/6J db/db 마우스에게여주추출물을경구투여하였을때혈청 TG 농도는여주열수추출물을 10mg/kg 및 50mg/kg을투여한그룹에서대조군에비해유의적인감소를보였다. 또한혈청 TC 농도와혈청 HDL-C 농도는 50mg/kg을투여한그룹에서유의적인감소를보였으며, 혈청 LDL 농도는여주열수추출물을투여한모든그룹에서유의적인감소를보였다고보고하였다. Popovich 등 (2010) 의연구에서도여주추출물이 3T3-L1 cell에서지질축적, adiponectin의발현및지방세포의감소효과가있었음을보고하였다. 따라서여주추출물은 in vitro 뿐만아니라고지방식이를공급받는정상체중을가진동물이나당뇨질병동물모두에서고지혈증을개선하는효과가있는것으로사료된다. 한편여주의성분들중에서특히주목받고있는것은당뇨병에효과가있는것으로밝혀진식물 insulin (P-insulin) 과카란틴이란성분이다. 식물 insulin은간에서포도당이연소되도록도우며포도당이체내에서재합성되지않도록하고, 카란틴은 insulin을분비하는췌장의기능을활발하게하는지용성성분으로혈당을낮추는작용을하는것으로보고하고있으며 (Jeong 등 2008), 다양한동물실험에서여주의종자, 과일, 잎이혈당을낮추는효과가있음을계속적으로보고하였다 (Day 등 1990; Ali 등 1993; Shibib & Rahman 1993; Sarkar & Marita 1996; Jayasooriya 등 2010). 특히여주는흰쥐에서내당능을개선시키고 (Leatherdale 등 1981) 식후과혈당을낮추며 (Uebanso 등 2007), 여주추출물은인슐린감수성을강화시킨다고보고하였다 (Chen 등 2003; Chen & Li 2005). 그러나본연구에서는여주열수추출물의섭취가혈당농도에는유의한효과를나타내지않았으며, 이러한결과에대해서는당질대사와관련된추후상세한추가연구가요구된다. Ⅴ. 요약및결론 본연구는여주열수추출물이고지방식이를섭취한마우스의체중증가와체지방축적억제, 혈중지질대사개선등에미치는영향을조사하고자 4주령 C57BL/6J 마우스를대상으로고지방식이에여주열수추출물을각각 5%, 10% 중량비로첨가하여 6주간급여하였다. 증체량은고지방식이대조군에비해여주열수추출물첨가군에서유의적으로낮았으며 (p<0.05) 식이효율은여주열수추출물농도의존적으로낮았다 (p<0.05). 장기중간의무게는고지방식이대조군에비해여주열수추출물 5% 첨가군이유의적으로낮았고 (p<0.05), 신장의무게는여주열수추출물첨가군에서모두유의하게낮았다 (p<0.05). 신장주변지방, 정소상체지방및총지방무게뿐만아니라체지방률은고지방식이대조군에비해여주열수추출물첨가군에서모두유의적으로낮았다 (p<0.05). 혈청중성지방농도는고지방식이대조군에비해여주열수추출물에서유의하게낮았으며, 혈청콜레스테롤농도는여주열수추출물농도의존적으로낮았다 (p<0.05). LDL-콜레스테롤농도는고지방식이대조군에비해여주열수추출물 10% 첨가군이유의적으로낮았다. 한편, 혈당농도는실험군간유의적인차이가없는것으로드러났다. 결과적으로 C57BL/6J 마우스에여주열수추출물의 6주간의장기급여는체중감소, 식이효율의감소그리고체지방축적억제뿐만아니라혈중지질대사를개선할수있음을의미한다. 더나아가여주열수추출물이어떤성분에의해비만억제및고지혈증개선효과를나타내는지와어떤기전에의해효과를나타내는지보다자세한연구가요구된다. 참고문헌 Ahn, I.S., Park, K.Y., Do, M.S.(2007). Weight Control Mechanisms and Antiobesity Functional Agents. J Korean Soc Food Sci Nutr, 36, Ali, L., Khan, A.K., Mamun, M.I., Mosihuzzaman., Nahar., Muhammadl.(1993). Studies on hypogly

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