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1 실험적간장해에대한강활의보호효과 윤수홍, 하헌 1, J. ENVIRON. TOXICOL. Vol. 20, No. 2, 161~165 (2005) 대구가톨릭대약학과, 1 상록병원약제과 Protective Effects of Angelica koreana on Experimentally Induced Liver Injury Soo Hong Yoon and Hun Ha 1, Department of Pharmacy, Catholic University of Taegu, Kyongsan , Korea 1 Department of Pharmacy, Sangrok Hospital, Kyongju , Korea ABSTRACT The present study was carried out to find the possible protective effects of Angelica koreana water extract on biochemical parameters in benzo(a)pyrene (B(a)P)-induced liver injury in rats. B(a)P treatment (0.1 mg/kg, i.p.) caused a liver damages, which led to biochemical alterations in serum and liver enzyme activities and serum lipid levels. The activities of liver marker enzymes, especially, aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP) and lactate dehydrogenase (LDH) were markedly changed in B(a)P treatment. Oral administration of Angelica koreana (50 mg/kg) recovered these biochemical parameters to near normal levels. Therefore, the present results have revealed that Angelica koreana water extract might have the antihepatotoxic effect and consequently ameliorate liver damage associated with B(a)P in rats. Key words : Angelica koreana, benzo(a)pyrene, antihepatotoxic effect 서 론 Benzo(a)pyrene (B(a)P) 은탄소화합물의불완전연소에의해생성되는다환방향족탄화수소 (polycyclic aromatic hydrocarbons) 로서식품, 매연, 담배등의실생활전반에걸쳐다양하게분포되어있는강력한발암물질이다 (Fischman et al., 1990). 체내에흡수될경우간의실질세포에서주로대사되며, 혈청 albumin 과결합하여이동하는데, cytochrome To whom correspondence should be addressed. Tel: , hunha@hotmail.com P448 에의해작용하는 arylhydrocarbon hydroxylase 에의해활성화되어 non-k region 에 epoxide 를형성한후, epoxide hydroxylase 에의해 mixed function oxidase 의작용에의해 B(a)P-diolepoxide 로대사된후간을주된 target organ 으로발암작용이개시되는것으로알려져있다 (Osborne et al., 1976; Weinstein et al., 1976). 한편강활은 Umbelliferae 에속하는 Angelica koreana 의뿌리혹은뿌리줄기를말린것을말하는데, 진통, 소염등의효능을나타내므로민간에서주로오한해열, 관절염, 두통, 근육통등에사용되어왔다 (Ryu and Yook, 1968; Moon et al., 1999; Sar- 161

2 162 J. ENVIRON. TOXICOL. Vol. 20, No. 2 kers and Nahar, 2004). 현재까지주성분으로는 imperatorin, isoimperatorin, oxypeucedanin, oxy-peucedanin hydrate 등의 coumarine 계물질 (Yook and Ryu, 1970; Suk, 1976; Lee and Woo, 1982; Kwon et al., 1991; Cho et al., 1998; Ryu et al., 2001; Seo et al., 2002) 과정유성분 (Chi et al., 1993) 등과항간디스토마작용 (Rhee et al., 1981) 을가진다고알려져있을뿐, 기타생체에서의약효및그기전에관한연구의보고는극히미약한실정이다. 따라서본실험에서는강활수침액의간보호능력을알아보고자실험동물에 B(a)P 을투여하여간독성을유발하는한편간기능의지표로사용되는효소활성과지질함량의변화를측정하여강활의간보호효과에대한기초자료를제시하고자한다. 1. 실험재료 재료및방법 본실험에사용된강활 (Angelica koreana) 은 2004 년대구시소재한약건재상에서구입하여세정, 음건하여강활의뿌리 (3 kg) 에물을가하여 10 시간씩 3 회추출한후감압농축시켜 250 g 을얻어적절한농도로희석하여수침액으로사용하였다. 2. 동물실험 외견상건강한 4 주령 SD 계웅성흰쥐를일반고형사료 ( 삼양사, 서울 ) 와물을충분히공급하며환경에적응시킨후, 평균체중이 180±20 g 인것을 8 마리씩 5 군으로난괴법에따라분리하여실험에사용하였고, 사육실의온도는 25±2 C 로유지하였으며, 12 시간주기로명암을조절하였다. 실험군은정상군 (normal), 강활 (Angelica koreana) 투여군, benzo(a)pyrene 투여군, 강활전처리군 (pre-treatment) 과강활후처리군 (post-treatment) 으로분리하였고, 강활전처리군은강활수침액을 1 일 1 회일정시간에 2 주일간투여시킨후, 마지막투여시 B (a)p 를복강투여하였으며, 24 시간이후도살하였다. 강활후처리군은 B(a)P 을 1 회복강투여직후부터강활수침액일정량을각각 1 일 1 회 2 주일간투여하였다. 강활수침액은체중 1kg 당 50 mg 을경구투여하였으며, B(a)P 는체중 1kg 당 0.1 mg 을 복강투여하였다. 3. 시료조제 대구가톨릭대학에서사육한흰쥐를 24 시간절식시킨다음, 에테르로마취시켜개복하여복부대동맥으로부터채혈한뒤실온에서 30 분간방치한후, 3,000 rpm 에서 15 분간원심분리하여혈청을분리하여효소활성및지질함량측정시료로사용하였다. 간조직을생리식염수으로관류시켜조직내혈액을제거하고적출하여, 간조직일정량에 4 배량의 0.25 M sucrose 용액을가하고, glass teflon homogenizer 로마쇄하여 20% (w/v) 균질액을얻고, 600 g 에서 10 분간원심분리하여핵및미마쇄부분을제거한상징액을얻고, 이것을다시 10,000 g 에서 20 분간원심분리하여얻은분획을측정의시료로사용하였다. 4. 혈청및조직에서의효소활성과지질함량측정 조제한시료에대하여혈청과간조직중의 aspartate aminotransferase (AST) 와 alanine aminotransferase (ALT) 활성도는 Reitman-Frankel 법 (1957) 으로, alkaline phosphatase (ALP) 활성도는 Kind-King 법 (1954) 에준하였으며, lactate dehydrogenase (LDH) 활성도와 total cholesterol, HDL-cholesterol, triglyceride, phospholipid 함량은효소법 (Takayama et al., 1977) 으로조제된아산제약 ( 서울, 한국 ) 시약을사용하였다. 5. 통계처리 실험결과의통계처리는 Student s t-test 에준하여평균 ± 표준편차로표시하였고, 통계적유의성은 p 0.05 수준에서검증하였다. 결과및고찰 1. 혈청의효소활성에미치는영향 다양한간질환을진단하고관리하는데혈청중의효소활성을검토하는것이매우유용하다. 가장흔히적용되는효소는 AST, ALT, ALP 등이며,

3 June 2005 Yoon and Ha : 실험적간장해에대한강활의보호효과 163 Table 1. The effects of Angelica koreana water extract on serum enzyme activity in benzo(a)pyrene treated rats AST 1) ALT 1) LDH 2) ALP 3) Normal 21.2± ± ± ±2.85 Angelica koreana 19.6± ± ± ±3.61 Benzo(a)pyrene 80.8±8.53* 57.7±6.07* 53.9± ±6.18 Pretreatment 35.5± ± ± ±2.00 Posttreatment 30.0± ±3.40* 27.0± ±3.61 Values are mean±s.d. of 8 rats. 1) karmen unit, 2) IU/L, 3) king-armstrong unit. Angelica koreana extract (50 mg/kg/day) was given orally for 2 weeks before/after benzo(a)pyrene treatment (0.1 mg/kg, i.p.). *p AST, aspartate aminotransferase; ALT, alanine aminotransferase; LDH, lactate dehydrogenase; ALP, alkaline phosphatase Table 2. The effects of Angelica koreana water extract on liver enzyme activity in benzo(a)pyrene treated rats AST 1) ALT 1) LDH 2) ALP 3) Normal 64.1± ± ± ±1.9 Angelica koreana 72.3± ± ± ±3.5 Benzo(a)pyrene 252.4±19.8* 466.5±26.6* 425.0±5.8* 133.1±4.0* Pretreatment 99.4±6.3* 223.1± ± ±39.2 Posttreatment 96.5± ±1.1* 285.3±23.4* 106.4±20.9 Values are mean±s.d. of 8 rats. 1) karmen unit, 2) IU/L, 3) king-armstrong unit. Angelica koreana extract (50 mg/kg/day) was given orally for 2 weeks before/after benzo(a)pyrene treatment (0.1 mg/kg, i.p.). *p AST, aspartate aminotransferase; ALT, alanine aminotransferase; LDH, lactate dehydrogenase; ALP, alkaline phosphatase gamma-glutamyl transferase (GGT) 도가끔적용된다 (Wolf, 1999). B(a)P 을투여하여간장해를유발한흰쥐혈청중의효소활성도변화에미치는강활수침액의영향은 Table 1 과같다. 강활수침액만의투여는 aminotransferase 활성도에는별다른영향을미치지못하여정상군과거의비슷하였지만, B(a)P 의투여로 AST 는약 4 배, ALT 는약 2 배증가시켜간의손상정도가현저하였다. 반면증가한효소활성은강활수침액투여로현저히감소하였는데, 전처리에의해서는약 56~47%, 후처리에의해서는약 63~49% 정도감소하여강활수침액이 B(a)P 에의한간독성으로증가한활성을감소시키는역할을하였다. 또한간세포손상에기인하는 ALT 활성의증가는보통 AST 활성증가와동반된다 (Mohan et al., 1989). 간에존재하는 aminotransferase 는간세포의괴사와간조직의파괴가진행되고있다는사실을나타내며, 결과적으로혈중으로이효소를방출케되어혈청효소의활성증가를야기시키는데, 이는간세포의변성및괴사의지표가된다 (Zimmerman, 1981). LDH 활성도는강활수침액투여로정상군보다약 1.4 배, B(a)P 투여는약 2.8 배증가를나타내었으며, 증가한효소활성은 수침액전처리로는약 29%, 후처리에서는약 50% 정도현저한감소작용을보였다. 한편 ALP 활성도는수침액단독투여군은정상군과거의비슷하였으며 B(a)P 투여는약 2 배정도활성증가를보였다. 그리고강활수침액전처리와후처리는약 40% 와 42% 정도의감소를보여정상군과거의비슷한활성도를나타내었다. 2. 간조직에서의효소활성에미치는영향 실험적간손상을일으킨흰쥐간조직중의효소활성도변화에미치는강활수침액의영향은 Table 2 와같다. 강활수침액단독투여는간조직의 aminotransferase 활성도에별다른영향을미치지못하였으나, B(a)P 투여는 AST 활성을약 4 배, ALT 활성은약 3 배증가시켜간의손상정도가아주심한것으로나타났다. 반면강활수침액의전처리는약 61~52%, 후처리로는약 62~54% 정도로감소시켜전 후처리의결과가거의비슷하였다. LDH 활성도는강활수침액단독투여군은약 1.3 배, B(a)P 투여는약 2.5 배증가하였으며, 증가한효소활성도는강활수침액전처리로약 20% 후처리는약 33% 의활성감소를보였다. 한편 ALP 활성도는

4 164 J. ENVIRON. TOXICOL. Vol. 20, No. 2 Table 3. The effects of Angelica koreana water extract on serum lipid contents in benzo(a)pyrene treated rats Total cholesterol HDL-cholesterol Triglyceride Phospholipid Normal 35.2± ± ± ±4.38 Angelica koreana 41.8± ± ± ±6.18 Benzo(a)pyrene 57.2±8.77* 21.2± ± ±10.61* Pretreatment 53.0±6.17* 25.2± ± ±9.29 Posttreatmnet 52.8±1.44* 18.1± ± ±7.05 Values are mean±s.d. of 8 rats. All units: mg/dl. Angelica koreana extract (50 mg/kg/day) was given orally for 2 weeks before/after benzo(a)pyrene treatment (0.1 mg/kg, i.p.). *p AST, aspartate aminotransferase; ALT, alanine aminotransferase; LDH, lactate dehydrogenase; ALP, alkaline phosphatase B(a)P 투여로약 5.3 배의높은활성증가를보였으며전처리는약 14%, 후처리는약 20% 의근소한감소를보였다. Cholestasis 는 ALP 의합성과방출을촉진시키며, bile salt 의축적이세포표면으로부터 ALP 방출을증가시킨다 (Schlager et al., 1982). 간손상에대한강활수침액의효과는혈청효소와간조직에서거의유사한결과를나타내었다. 3. 혈청의지질함량에미치는영향 혈청총지질은주로 cholesterol 과 cholesterolester, phospholipid 그리고 triglyceride 로구성된다. 혈청 cholesterol 의약 70% 가 ester 화된것인데, 이 ester 화가주로간에서일어나서, 혈청 cholesterol 함량측정이간기능시험에자주적용된다. B(a)P 을투여하여간장해를유발한흰쥐혈청중의지질함량변화에미치는강활수침액의영향을검토한결과는 Table 3 과같다. Total cholesterol 함량은 B (a)p 투여로정상군과비교할때, 약 62% 정도증가하였다. 반면증가한함량은강활수침액전처리와후처리에서약간의감소를보였다. 한편, HDLcholesterol 은수침액후처리군만이정상군보다약 60% 정도의감소를보였으며별다른유의성이없었다. Triglyceride 함량은강활수침액투여로정상군과비슷하였으나, B(a)P 투여로오히려약 25% 의감소를보였다. 그리고강활수침액전처리와후처리군도 B(a)P 투여군과거의비슷한함량변화를보였다. Phospholipid 함량은수침액투여로정상군에비하여변화가없었지만, B(a)P 투여로약 31% 정도증가하였다. 이는세포막의구성, 구조와안정성의변화를일으키고, 결과적으로세포막기능이상을초래케된다 (Kehrer et al., 1990). 증가한 phospholipid 함량은강활수침액전처리와후처리에 의하여약간감소하였지만유의성은없었다. 결 론 본연구는민간에서소염진통제로널리쓰이는강활의생리활성물질검색및간손상에미치는영향을관찰할목적으로실행되었다. 실험동물에다환방향족탄화수소이며환경공해물질인 B(a)P 를복강투여하여유도한간손상에미치는강활수침액의효과를알아보고자혈청과간조직에서효소활성도및혈청에서지질함량의변화를측정하였다. B(a)P 의투여에의하여혈청과간조직의 AST, ALT, LDH 및 ALP 활성은유의성있게증가하였으며, 강활수침액은증가한효소활성을회복시켰다. 한편 B(a)P 의투여에의하여혈청 total cholesterol 과 phospholipid 함량은증가하였지만 triglyceride 함량은오히려감소하였다. 따라서강활수침액은 B(a)P 의투여로야기된간실질세포의손상과혈청 AST, ALT, ALP 활성증가로암시된담즙분비기능이상등의간독성 (Baraona et al., 1980) 에대해긍정적인보호효과를가지는것으로사료된다. 참고문헌 Baraona E, Pikkarainen P, Salaspuro M, Finkelman F and Lieber CS. Acute effects of ethanol on hepatic protein synthesis and secretion in the rat, Gastroenterol. 1980; 79: 104. Chi HJ and Kim HS. Studies on essential oils of plants of Angelica genus in Korea (IV)-Essential oils of Angelica

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<30382EC0C7C7D0B0ADC1C22E687770> 대한내과학회지: 제 76 권 제 2 호 2009 의학강좌-개원의를 위한 모범처방(Current Clinical Practice) 간기능검사의 이해와 적용 인제대학교 의과대학 일산백병원 내과학교실 김 경 아 Understanding and application of liver function tests Kyung-Ah Kim, M.D. Department of