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1 J. Biomed. Lab. Sci. 10 (2004) Effects of Liquid Culture of Agaricus blazei Murill on Protein, Mineral Levels and Enzyme Activities in Rats Jin-Bog Koh 1 and Jai-Young Kim 2 1 Department of Life Science, Silla University, Busan , Korea 2 Department of Clinical Laboratory Science, Catholic University of Pusan, Busan , Korea This study was performed to investigate the effects of liquid culture of Agaricus blazei Murill on the weight gains, food efficiency ratios, serum protein and mineral levels, and serum enzyme activities in growing male rats. Sprague- Dawley rats (7 weeks old) were given four different types of diets for a succeeding period of 6 weeks, respectively: a normal diet group, a control diet group (normal diet + 15% lard + 0.5% cholesterol), a 30% or 40% A. blazei diet groups (control diet + 30% or 40% A. blazei in water) according to the levels of A. blazei supplementation. The body weight gains, food efficiency ratios, and the liver of the rats fed control diet, 30% or 40% A. blazei diets were significantly increased compared to rats fed the normal diet, but those of rats fed the 30% and 40% A. blazei diets were similar to those of rats fed the control diet. The concentrations of total protein, albumin, glucose, and hematocrit value in serum of rats fed the control diet, 30% or 40% A. blazei diets were similar to those of rats fed the normal diet. The concentrations of urea and creatinine in serum of rats fed the 30% or 40% A. blazei diets were similar to those of rats fed the control diet. but the urea of rats fed the 30% and 40% A. blazei diets were significantly decreased compared to rats fed the normal diet. The concentrations of Ca, P, Fe and Mg, and Ca ++, K +, Na + and Cl - of rats fed the control diet, 30% or 40% A. blazei diets were similar to those of rats fed the normal diet. There were no differences in the activities of aspartate aminotransferase, alanine aminotransferase, γ-glutamyltranspeptidase and alkaline phosphatase in the serum among the experimental groups. In conclusion, the rats fed the A. blazei maintained normal protein and mineral levels, and enzyme activities of serum. But the A. blazei feeding could not decrease the body and liver weights in the rats fed high cholesterol diets. Key Words: Mushroom, Liquid cultures, Agaricus blazei, Mineral, Protein, Enzyme activity 서 약용버섯이나식용버섯은다당류, 단백질, 비타민, 무기질, 섬유소, 스테롤등의영양소가풍부할뿐만아니라저열량식품으로성인병의예방이나치료효능이있는것으로알려지면서버섯의이용이증가되고있다. 담자균류의단백다당체들은항암활성을가지고있으며현재가장높은사망원인인암뿐만아니라순환기계질환에도치료효과가있는것으로보고 15,17) 되어있는등생리활성물질에관한연구가진행되고있다. * 논문접수 : 2004 년 4 월 12 일수정재접수 : 2004 년 6 월 18 일 별책요청저자 : Jai-Young Kim, Department of Clinical Laboratory Science, Catholic University of Pusan, Busan, Korea Tel: , Fax: jykim@cup.ac.kr 론 담자균류에속하는신령버섯 (Agaricus blazei Murill) 은식용버섯의한종류로브라질이원산지이다. 신령버섯은국내에서는아가리쿠스버섯또는흰들버섯으로불리며, 전체적인외형은양송이와유사하지만양송이보다향이강하고버섯대가두껍고길며육질의맛이좋은것이특징이다 28). 신령버섯의자실체는 85~87% 가수분이고건조한버섯은단백질 40~45%, 탄수화물 38~45%, 섬유질 6~8%, 회분 5~7%, 지방질 3~4% 를함유하고있고 20), 균사체는수분 10.7%, 단백질 15.6%, 탄수화물 42.4%, 섬유질 26.4%, 회분 5.9%, 지방질 9.6% 로당질과단백질이풍부한버섯이다 2). 신령버섯은혈당과혈압강하효과와콜레스테롤저하, 항종양, 항암및항돌연변이효과등의약리작용이있는것으로알려져있고 19,27,29), 신령버섯의다당체는인터페론을활성화해서암세포를소멸또는억제하는간접적인효과가높으며, 특히다른버섯류와는달리고형암뿐만아니라복수암, 결장암, 난소암, 유방암, 폐암, 간암등에도효과가있다. 이

2 러한항암효과를나타내는다당체는 β-glucan으로그구조는 β-(1-6)-glucosyl의가지를가진 β-(1-3)-glucan이다 4,5,7). 신령버섯의자실체에서분리한단백다당체는당질이 50.2% 이고단백질이 43.3% 로이들복합체가항종양과면역조절작용이있다고하였고 8,9,21), 다당체는생쥐의림프구의 T 세포를자극시킬수있다 22). 최와구 3) 는신령버섯의 β-glucan은비만인당뇨쥐의식후혈당의상승을억제하는효과가있고중성지방농도를낮추어혈액의지질조성을개선시켜심혈관질환을감소시키는것을보고하였다. 이상의연구내용에서신령버섯의다양한약리작용이알려지고또한만성적인성인병의예방이나치료제로이용되고있으나, 고콜레스테롤혈증에미치는신령버섯의효과에대하여는혈청콜레스테롤농도를감소시킨다고 27) 하였으나체내영양상태및생리활성에대한체계적인연구는드문실정이다. 따라서본연구는신령버섯의균사체액체배양액이고콜레스테롤식이를섭취한흰쥐의단백질영양상태, 효소활성및무기질농도에미치는영향을관찰하고자생후 7주령의수컷흰쥐를대상으로하여콜레스테롤식이에신령버섯의균사체액체배양액을 30% 와 40% 로음료수에혼합하여 6주간급여하고, 체중변화, 식이효율, 혈청의단백질농도, 무기질및간질환과관련이있는효소활성등을조사하여영양 생리활성에미치는영향을알아보는것이본연구의목표이다. 재료및방법 1. 신령버섯균사체액체배양액제조방법시료로사용된신령버섯 (Agaricus blazei Murill) 의균사체음료는케이비에프 ( 주 ) 에서제조한것으로신령버섯의균사체 200 ml를 PDB (potato dextrose broth) 에서 25 로조절하여 4일간액체배양하고, 전체배양액을본배양배지에서 3 일간액체배양하였다. 배양된균사체배양액을거즈로여과하여, 여과액을 105 에서 90분간감압멸균하여시료로사용하였다. 시료보관은 -20 에서냉동보관하였다. 2. 실험동물의식이및사육실험동물은본대학실험실에서번식시켜고형사료 ( 삼양유지 ) 로사육한생후 7주령의 Sprague-Dawley계수컷흰쥐를표준사료로 1주일동안적응시킨후, 평균체중이 g 의동물을한군에 8마리씩 4군으로나누어실험에사용하였다. 실험식이의조성은 Table 1과같으며, 실험식이군은정상식이군 (AIN-9326)), 대조식이군 ( 정상식이에 20% 돈지와 0.5% 콜레스테롤을첨가 ), 30% 및 40% 신령버섯군 ( 대조식이에신령버섯균사체액체배양액을 30% 및 40% 수준으로음료수에혼합하여급여한군 ) 등 4군으로나누어해당식이 Table 1. Composition of experimental diets (g % ) Ingredients Normal Control Casein Corn oil Lard Corn starch Sucrose DL-methionine Choline bitartarate Cellulose Mineral mix. 1) Vitamin mix. 2) Cholesterol Sodium cholate ,2) AIN-93-MX mineral and AIN-93-VX vitamin mixture 26) 로 6주간사육하였다. 동물실험실의사육조건은온도 22± 2, 습도 40~50% 로유지시키고, 명암은 12시간을주기로 자동조절되었으며, 실험음료와식이는자유급식하였다. 3. 식이섭취량, 식이효율및체중측정 체중은 1주에한번씩일정한시간에측정하였고, 실험기간 동안의식이는매일오후 4시에일괄적으로급여하였다. 식 이섭취량의오차를최소화하고자손실량을측정하여보정 하였으며급여량을기록하여식이섭취량을산출하였다. 식이 효율은실험전기간의체중증가량을같은기간동안에섭취 한식이량으로나누어산출하였다. 즉식이효율 (food efficiency ratio; FER) = 체중증가량 (g)/ 식이섭취량 (g) 100으로계 산하였다. 4. 시료채취및분석 6주간실험종료일에 16시간절식시킨실험동물을 ethyl ether로마취하고심장에서채혈하고, 채혈된혈액의일부는 EDTA병에넣어전혈로사용하였고, 나머지는 SST관에넣 어실온에서 30분간응고시킨후 3,000 rpm에서 20분간원심 분리하여혈청을분석시료로사용하였다. 각장기는채혈후 즉시떼어생리식염수로혈액을씻은다음무게를측정하였 다. Hematocrit치는 microhema tocrit법 18) 으로측정하였고, 혈 당, 단백질, 알부민농도및 aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), γ-glutamyl transpeptise (γ-gtp) 의활성은자동생화학분석기 (Autohumalyzer 900S, Germany) 로측정하였다

3 Table 2. The body weight gain, food intake, food efficiency ratio and hematocrit value of male rats fed Agaricus blazei (AB) diets for 6 weeks Body weight gains (g) Groups 1) Initial Final Gains Food intake (g/day) FER (%) Glucose (mg/dl) Normal 288.1±19.5 2) 424.0± ±20.9 a 24.96±2.11 NS3) 16.70±1.81 a 188.5±24.2 NS Control 289.5± ± ±30.5 b 22.55± ±2.31 b 170.1± % AB 289.1± ± ±17.6 b 21.85± ±2.54 b 189.9± % AB 282.1± ± ±27.6 b 21.04± ±2.43 b 181.3±22.9 1) Group abbreviations: Normal = normal diet group, Control = normal diet + 15% lard + 0.5% cholesterol diet group, 30% or 40% AB = control diet + 30% or 40% liquid culture of Agaricus blazei mixed with water, 2) All values are mean ± SD (n = 8). 3) NS: not significant among the groups, a-b Values within a column with different superscripts letters are significantly different each other groups at P<0.05 Table 3. The organ weight of male rats fed Agaricus blazei (AB) diets for 6 weeks 1) Liver (g) Kidney (g) Pancreas (mg) Groups Weight L/BW 4) Weight K/BW Weight F/BW Normal 12.41±1.52 2)a 2.92±0.21 2)a 2.73±0.26 NS3) 0.64±0.05 NS 674±102 NS 159±25 NS Control 16.40±3.01 b 3.69±0.36 b 2.98± ± ± ±29 30% AB 17.92±2.50 b 3.76±0.32 b 3.03± ± ± ±31 40% AB 17.50±2.21 b 3.85±0.35 b 3.00± ± ± ±30 1~3) See the legends in Table 1, 4) Liver, Kidney and Pancreas weight for 100 g body weight 5. 통계처리본연구의실험결과는평균치와표준편차로나타내었고, SPSS를이용하여실험군간의유의성은 ANOVA 로검증한후 P<0.05 수준에서 Duncan's multiple range test로상호비교분석을하였다. 결과및고찰 1. 체중증가량, 식이섭취량및식이효율 6주간실험식이를급여한결과실험동물의체중증가량, 식이섭취량및식이효율의변화는 Table 2와같다. 체중증가량은정상군에비하여대조군 (20% 돈지와 0.5% 콜레스테롤첨가군 ), 대조식이와 30% 및 40% 의신령버섯을급여한군 ( 신령버섯군 ) 들이각각 22.0%, 26.6% 및 17.9% 로유의하게증가되었고, 대조군과신령버섯급여군간에는유의적인차이는없었다. Anderson 1) 등은 10주령된쥐에 3주간실험식이로사육한바정상식이군에비해 1% 콜레스테롤첨가군이 12% 증가되었다고하였고, 김 11) 등, 고와최 14) 는 1% 콜레스테롤첨가한식이로성장기쥐나성숙한 (13주령) 쥐를사육하였을때에정상군보다유의하게체중과간의무게가증가되었다고하 였다. 본실험의결과도위의보고와일치하였고, 대조군과신령버섯군들이체중증가가비슷하여신령버섯균사체배양액이콜레스테롤섭취로증가된체중을낮추는효과가나타나지않았다. 식이섭취량은정상군에비해각실험군이다소낮은경향이나유의한차이는아니었고, 식이효율은정상군에비해대조군, 30% 및 40% 신령버섯군이각각 34.7%, 44.3% 및 39.5% 로유의하게증가되었고, 대조군과신령버섯급여군간에는유의적인차이는없었다. 이는콜레스테롤섭취로체중증가와더불어식이효율이증가된것으로나타났다. 2. 장기무게변화 6주간의신령버섯균사체배양액을급여한실험동물의각장기무게는 Table 3에표시하였다. 간의무게는정상군에비하여콜레스테롤첨가군인대조군, 30% 및 40% 신령버섯군이각각 26.3%, 28.7% 및 31.8% 로유의하게증가되었다. 선행연구 11~14,16,23,24) 에서고지방식이나콜레스테롤식이를급여한쥐의간이비대되었다고하였다. 이는식이중과량의콜레스테롤이간으로유입되었을때콜레스테롤과잘결합할수있는 apolipoprotein과결합하여 lipoprotein으로배출되지못하면지방간으로되거나간세포손상을일으킬수있는것으로알려져있다 25). 본실험결과도상기보고와일치되며실험식이에콜레스테롤첨가로간에지방의축적으로간이비

4 대된것으로신령버섯균사체액체배양액이간의지방축적을억제하지는못하는것으로나타났다. 신장및췌장의무게는정상군과각실험군이비슷한경향으로나타났다. 3. 혈청의단백질, 요소질소, 크리아티닌및혈당농도콜레스테롤식이에신령버섯균사체배양액급여시단백질영양상태및혈당농도에미치는영향을조사한결과는 Table 4와같다. 혈청의총단백질과알부민농도및알부민 / 글로불린비율은정상군및대조군과각수준별신령버섯군들이비슷한경향으로단백질농도는정상수준으로나타났다 6,10). 혈청알부민은혈청총단백질의 50~70% 를차지하며혈장교질삼투압의유지와각종영양소등의운반에중요한기능을담당하는데일반적으로총단백질농도와상관관계를가지며체내단백질대사이상의좋은지표가된다. 본실험의결과총단백질과알부민농도및알부민 / 글로불린비율은신령버섯균사체배양액의섭취에따른영향은없었다. 요소질소는단백질대사의최종산물로요로배설된다. 혈 청에서요소질소의농도는요소질소의배설장애, 신부전, 간경변증, 탈수, 조직붕괴, 단백질의이화작용을촉진하는항생제사용등에서상승하고, 성장호르몬이나단백질동화호르몬은요소의농도를감소시키는작용이있다. 본실험결과혈청의요소질소농도는정상군에비하여대조군및각수준별신령버섯군들이유의하게감소되었고, 대조군과신령버섯군들은비슷한농도를보였다. 이는본실험에서정상군에비해대조군과신령버섯군들의체중이유의하게증가되었고신장의무게가다소증가된것으로보아단백질동화작용의증가로혈액의요소가감소된것으로추측되나더연구가필요하다. 크리아티닌은근육에서크리아틴의대사최종산물로신세뇨관에서재흡수없이대부분요로배설된다. 혈청에서크리아티닌농도는요독증, 만성신장염등에서증가하고사구체여과율과밀접한상관성이있으므로신장기능장애의중요한지표가된다. 본실험결과혈청의크리아티닌농도는정상군에비해대조군이유의하게증가되었으나각수준별신 Table 4. The serum protein, urea nitrogen (U-nitrogn), creatinine and glucose concentrations of male rats fed Agaricus blazei (AB) diets for 6 weeks Groups 1) Total protein g/dl Albumin g/dl A/G 4) ratio U-nitrogen mg/dl Creatinine mg/dl Hematocrit (%) Normal 6.66±0.29 2)NS 3.68±0.23 NS3) 1.24±0.10 NS 13.91±1.73 b 0.46±0.02 a 49.58±1.81 NS Control 6.96± ± ± ±2.10 a 0.49±0.02 b 49.61± % AB 6.75± ± ± ±1.27 a 0.48±0.03 ab 50.41± % A 6.78± ± ± ±1.20 a 0.48±0.03 ab 50.36±0.76 1~3) See the legend of Table 2, 4) A/G ratio: albumin/globulin ratio Table 5. The serum mineral concentrations of male rats fed Agaricus blazei (AB) diets for 6 weeks Groups 1) Phosphorus (mg/dl) Magnesium (mg/dl) Iron (µg/dl) Normal 7.75±0.77 2)NS 2.90±0.62 NS3) 114.3±21.6 NS Control 7.16± ± ± % AB 7.83± ± ± % AB 7.46± ± ±23.7 1~3) See the legend of Table 2 Table 6. The serum mineral ion and electrolyte concentrations of male rats fed Agaricus blazei (AB) diets for 6 weeks (mmol/l) Groups 1) Total Ca Ca ++ Ca ++ /T-Ca(%) K + Na + Cl - Normal 2.83±0.12 2)NS 1.19±0.10 NS3) 42.26±3.78 NS 4.26±0.65 NS 144.0±3.31 NS 97.88±3.80 NS Control 2.83± ± ± ± ± ± % AB 2.96± ± ± ± ± ± % A 2.75± ± ± ± ± ±3.32 1~3) See the legend of Table 1, T-Ca: total Ca

5 Table 7. The alanine aminotransferase (ALT), aspartate aminotransferase (AST), γ-glutamyltranspeptidase (γ-gtp) and alkaline phosphatase (ALP) activities in serum of male rats fed Agaricus blazei (AB) diets for 6 weeks (IU/L) Groups 1) ALT AST γ-gtp ALP Normal 42.39±6.95 2)NS 132.1±14.9 NS3) 8.60±1.29 NS 223.1±30.7 a Control 63.61± ± ± ±35.0 b 30% AB 56.52± ± ± ±29.7 ab 40% AB 60.22± ± ± ±28.9 ab 1~3) See the legend of Table 2, a-b Values within a column with different superscripts letters are significantly different at P<0.05 령버섯군들은정상군보다다소증가하였으나유의한차이는나타나지않았다. 30% 나 40% 수준의신령버섯균사체배양액섭취로크리아티닌대사에는영향을주지않는것으로나타났다. 혈당농도는정상군과각실험군이비슷하였다. 이상의결과로보아고콜레스테롤식이와신령버섯균사체배양액을 30% 및 40% 수준의급여시단백질, 요소질소및크리아티닌및혈당의농도에는영향을주지않았다. 4. 혈청의무기질농도신령버섯에함유된다당류나섬유소가혈청의무기질농도에미치는영향을조사한결과는 Table 5와 6과같다. 혈청의칼슘, 무기인, 마그네슘및철의농도는정상군과대조군및각수준별신령버섯군이비슷하였다. 그리고혈청의칼슘이온, 칼슘이온 / 총칼슘의비율, 칼륨이온, 나트륨이온및염소이온의농도도정상군과대조군및각수준별신령버섯군이비슷한경향으로신령버섯균사체배양액섭취에따른영향은나타나지않았다. 5. 혈청의효소활성콜레스테롤식이와신령버섯균사체배양액급여시흰쥐의간질환과관련이있는효소활성을측정한결과는 Table 7과같다. 혈청의 aspartate aminotransferase (AST), alanine aminotransferase (ALT) 및 γ-glutamyltranspeptidase (γ-gtp) 의활성은정상군과각실험군들이유사한경향을보였다. Alkaline phosphatase (ALP) 의활성은정상군에비해대조군이유의하게증가되었으나, 대조군에비해신령버섯군들은유의한차이는아니나감소되어정상군과비슷한수준으로나타났다. 임상에서 AST 및 ALT는간세포에다량존재하는효소로간손상시세포외로다량유출되어혈액에증가됨으로서간손상의지표로이용되는효소이다. 체내에서 ALP는여러가지인산에스터를분리시키고, 골격내에서석회화를촉진시키며장내에서는인흡수등에관여하는효소로특히골질환, 간이나담도질환, 임신및악성종양등에서활성치가상승한다. γ-gtp는 γ-glutamylpeptide의 γ-glutamyl기를아미노 산또는 peptide에전이시키는효소로신장, 췌장, 간, 담도를비롯한여러장기에분포하고특히담즙울체성질환, 알코올성이나약물성간장해등에서높은활성치를보이는효소이다. 상기효소들은간질환이나여러종류의질환에임상적지표로이용된다. 본실험결과신령버섯균사체배양액섭취시 AST, ALT 및 γ-gtp 활성치는정상군과유사한경향으로나타났음은 30~ 40% 수준의신령버섯균사체배양액섭취시이러한효소활성에특별한영향을주지않음을알수있다. 결론신령버섯균사체배양액이콜레스테롤식이를섭취한숫쥐의성장률, 장기무게, 단백질농도, 무기질농도및효소활성에미치는영향을조사하고자, 생후 7주령의흰쥐에표준식이를급여한정상군, 표준식이에 15% 돈지와 0.5% 콜레스테롤을첨가한식이를급여한대조군, 대조식이에신령버섯의균사체배양액을음료수에 30% 및 40% 로혼합급여한군 (30% 및 40% 신령버것군 ) 등 4군으로나누어 6 주간사육한결과는다음과같다. 실험동물의체중증가량, 식이효율및간의무게는정상군에비해대조군과 30% 및 40% 신령버섯군이유의하게증가되어신령버섯섭취에따른체중감소효과는나타나지않았다. 신장및췌장의무게는실험군간에비슷하였고, 총단백질, 알부민, 알부민 / 글로불린비율및혈당농도는실험군간에비슷한농도를유지하였다. 혈청의 Ca, P, Mg 및 Fe 농도, 그리고혈청의 Ca ++, K +, Na + 및 Cl - 농도는처치구간에비슷한경향으로신령버섯균사체배양액섭취에따른무기물과전해질농도에미치는영향은나타나지않았다. 간기능지표로이용되는혈청의 AST, ALT, γ-gtp 및 ALP 의활성도처치구간비슷한수준으로나타난것으로보아신령버섯균사체배양액섭취에따른간기능저하에미치는영향은없는것으로보인다

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경락경혈학회지 Vol.27, No.1, pp.87 106, 2010 Journal of Meridian & Acupoint Dept. of 1 Meridian & Acupoint, 3 Acupuncture & Moxibustion, College of Oriental Medicine, Daejeon University 2 Division of Clinical