KR20160083781A - Method for preparing fermented citrus composition with improved bioactivity - Google Patents
Method for preparing fermented citrus composition with improved bioactivity Download PDFInfo
- Publication number
- KR20160083781A KR20160083781A KR1020150035746A KR20150035746A KR20160083781A KR 20160083781 A KR20160083781 A KR 20160083781A KR 1020150035746 A KR1020150035746 A KR 1020150035746A KR 20150035746 A KR20150035746 A KR 20150035746A KR 20160083781 A KR20160083781 A KR 20160083781A
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- South Korea
- Prior art keywords
- citrus
- fermented
- activity
- present
- converting enzyme
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Abstract
Description
본 발명은 생리활성 기능성이 향상된 감귤 발효물 제조방법에 관한 것으로, 구체적으로 락토바실러스 사케이(Lactobacillus sakei) KACC92032P, 페디오코커스 아시딜락티시(Pediococcus acidilactici) KACC92031P 및 류코노스톡 메센테로이데스(Leuconostoc mesenteroides) KCTC3505로 이루어진 군에서 선택된 젖산균으로 감귤을 발효하여 아세틸콜린에스터레이즈 저해활성, 안지오텐신 전환 효소 저해활성 또는 항균활성을 갖는 감귤 발효물을 제조하는 방법, 이 방법으로 제조된 발효물 및 이 발효물을 포함하는 식품 조성물에 관한 것이다.
The present invention relates to a method for preparing a citrus fermented product having improved physiological activity, and more particularly, to a method for producing a citrus fermented product having Lactobacillus sakei ) KACC92032P, Pediococcus, Pediococcus acidilactici KACC92031P and Leuconostoc < RTI ID = 0.0 > mesenteroides KCTC3505 to produce a citrus fermented product having an inhibitory activity on acetylcholine esterase inhibition, an angiotensin converting enzyme inhibitory activity or an antimicrobial activity, a fermented product produced by the method and a fermented product ≪ / RTI >
현대인들은 서구화된 식생활과 영양불균형으로 인해 각종 만성질환이 증가하면서 건강에 대한 관심이 높아지고 있다. 또한 노화억제 및 질병예방 등의 의학적 접근뿐 아니라 식품 영양학적 접근을 통해 예방하려는 노력이 증대되고 있다. 식품으로 사용되는 각종 재료들 중에는 인체에 유익한 성분들을 함유하고 있는 것들이 많이 존재한다. 특히, 식물성 재료들에는 매우 다양한 생리활성물질들이 존재하며 이들의 생리활성 및 분리 정제방법 등을 연구하여 다양한 질병의 치료용도로도 활용하고 있다.Modern people are increasingly interested in health due to the increasing number of chronic diseases due to Westernized diet and nutritional imbalance. In addition, there is an increasing effort to prevent through aging and disease prevention as well as medical approaches such as food nutritional approach. Among the various materials used as food, there are many things that contain ingredients that are beneficial to the human body. In particular, there are a wide variety of physiologically active substances in plant materials, and their physiological activities, separation and purification methods, and the like have been studied and utilized for treatment of various diseases.
감귤은 운항과(Rutaceae) 감귤속(Citrus)의 식용 식물로 6대륙 100개국 이상에서 재배되는 전 세계적으로 즐겨먹는 과일 중 하나이다. 감귤에는 carotenoid, flavonoid, cellulose, pectin, limonoid, 비타민류 등의 다양한 생리활성물질들이 다량 함유되어 있어 항산화, 항염, 항암 등 다양한 생리적 작용을 나타낸다고 보고되어 있다. 감귤은 생과 형태로 주로 소비가 되고, 일부는 착즙하여 주스가공으로 이용되어져 왔다. 또한 근래에는 초콜릿, 크런치, 말랭이 등의 다양한 감귤 가공 제품으로 확산되고 있다.Citrus is an edible plant of the Rutaceae Citrus which is one of the world's favorite fruits grown in more than 100 countries on six continents. It is reported that citrus fruits contain various physiologically active substances such as carotenoids, flavonoids, cellulose, pectin, limonoids and vitamins and exhibit various physiological functions such as antioxidant, anti-inflammation and anti-cancer. Citrus fruits are mainly consumed in raw and dried forms, and some have been used for juice processing. In recent years, various citrus processed products such as chocolate, crunch, and malt liquor have been spread.
젖산균(lactic acid bacteria)은 당류를 발효하여 젖산 및 다양한 대사산물을 생산하는 미생물로 최근에는 건강 증진과 질병을 예방하고자 하는 probiotics로서의 연구가 폭넓게 진행되고 있으며, 각종 발효식품, 사료첨가제 등의 제조에 널리 이용되고 있다. 또한 발효식품을 통하여 인체 내로 섭취한 젖산균은 장 내로 유입되어 유익균주의 생육을 부양하고 유해세균을 억제하여 장내 유용미생물의 균형을 유지시킨다. 그리고 항산화작용, 항암작용, 면역 활성화 작용, 혈중 콜레스테롤 저하 등의 작용이 있어 질병을 예방하는데 도움이 된다고 보고되어 있다.Lactic acid bacteria are microorganisms that produce lactic acid and various metabolites by fermenting saccharides. Recently, researches as probiotics for prevention of health promotion and diseases have been extensively carried out and various fermented foods, feed additives and the like It is widely used. In addition, lactic acid bacteria taken into the human body through fermented foods are introduced into the intestines to support the growth of beneficial bacteria and inhibit harmful bacteria to maintain the balance of useful microorganisms in the intestines. And antioxidant, anticancer, immune activation, and blood cholesterol lowering effects are known to help prevent disease.
본 발명자들은 감귤을 사용하여 보다 유익하고 우수한 식품을 개발하고자 하였으며, 특히 질병의 예방 또는 개선에 도움을 줄 수 있는 식품을 개발하기 위하여 연구하였다. 이에 감귤에 포함되어 있는 유용성분들과 젖산균이 보유하는 효소의 상호작용을 통해 새로운 생리활성물질이 생성되거나 감귤 고유의 유용 생리활성물질의 농도를 높일 수 있을 것이라 기대하고, 다양한 젖산균으로 감귤을 발효하는 연구를 수행하였다.The present inventors tried to develop a more beneficial and excellent food by using citrus fruits, and in particular, to develop foods that can prevent or improve disease. Therefore, it is expected that the interaction of the enzymes contained in the citrus fruits with the enzymes possessed by the lactic acid bacteria will produce new physiologically active substances or increase the concentration of useful physiologically active substances inherent in citrus, and ferment the citrus fruits with various lactic acid bacteria Study.
이의 결과, 감귤을락토바실러스 사케이(Lactobacillus sakei) KACC92032P, 페디오코커스 아시딜락티시(Pediococcus acidilactici) KACC92031P 및 류코노스톡 메센테로이데스(Leuconostoc mesenteroides) KCTC3505로 발효하면, 아세틸콜린에스터레이즈 저해활성, 안지오텐신 전환 효소 저해활성 또는 항균활성이 우수한 발효물을 생성할 수 있음을 확인하였으며, 이러한 효과를 바탕으로 알츠하이머 질환과 고혈압을 예방하거나 개선할 수 있고 항균효과가 우수한 기능성 식품을 제조할 수 있음을 확인하고 본 발명을 완성하게 되었다.
As a result, the citrus fruits were treated with Lactobacillus sakei KACC92032P, Pediococcus acidilactici KACC92031P and Leuconostoc mesenteroides KCTC3505 can produce a fermented product having excellent acetylcholine esterase inhibitory activity, angiotensin converting enzyme inhibitory activity or antimicrobial activity. Based on these effects, it is possible to prevent or ameliorate Alzheimer's disease and hypertension And can produce a functional food excellent in antimicrobial effect. Thus, the present invention has been completed.
따라서 본 발명의 주된 목적은 감귤을 재료로 하여 질병의 예방 또는 개선에 효과적인 식품을 제조하는 방법을 제공하는데 있다.Accordingly, a main object of the present invention is to provide a method for producing a foodstuff effective for prevention or improvement of disease by using citrus as a material.
본 발명의 다른 목적은 상기 제조방법으로 제조된 기능성 식품을 제공하는데 있다.
Another object of the present invention is to provide a functional food prepared by the above production method.
본 발명의 한 양태에 따르면, 본 발명은 락토바실러스 사케이(Lactobacillus sakei) KACC92032P, 페디오코커스 아시딜락티시(Pediococcus acidilactici) KACC92031P 및 류코노스톡 메센테로이데스(Leuconostoc mesenteroides) KCTC3505로 이루어진 군에서 선택된 젖산균으로 감귤을 발효하는 것을 특징으로 하는 아세틸콜린에스터레이즈(Acetylcholinesterase, AChE) 저해활성, 안지오텐신 전환 효소(Angiotensin converting enzyme, ACE) 저해활성 또는 항균활성을 갖는 감귤 발효물 제조방법을 제공한다.According to one aspect of the present invention, the present invention provides a pharmaceutical composition comprising Lactobacillus sakei KACC92032P, Pediococcus acidilactici KACC92031P and Leuconostoc (ACE) inhibitory activity, an angiotensin converting enzyme (ACE) inhibitory activity, or an antimicrobial activity of citrus fruits, characterized in that citrus fruits are fermented with lactic acid bacteria selected from the group consisting of mesenteroides KCTC3505 And a manufacturing method thereof.
본 발명의 감귤 발효물 제조방법에 있어서, 상기 발효는 20 내지 40℃에서 하루 이상 이루어지는 것이 바람직하다.In the method for preparing a citrus fermented product according to the present invention, it is preferable that the fermentation is performed at 20 to 40 DEG C or more per day.
본 발명에 따르면, 젖산균으로 락토바실러스 사케이(Lactobacillus sakei) KACC92032P를 사용할 경우, 아세틸콜린에스터레이즈 저해활성, 안지오텐신 전환 효소 저해활성 또는 항균활성을 높이기 위해 28 내지 32℃에서 7일 이상 발효하는 것이 바람직하다. 보다 바람직하게는 7 ~ 10일간 발효하는 것이 좋다.According to the present invention, Lactobacillus < RTI ID = 0.0 > sakei ) When KACC92032P is used, it is preferable that fermentation is carried out at 28 to 32 DEG C for 7 days or more in order to increase acetylcholine esterase inhibitory activity, angiotensin converting enzyme inhibitory activity or antimicrobial activity. More preferably 7 to 10 days.
본 발명에 따르면, 젖산균으로 페디오코커스 아시딜락티시(Pediococcus acidilactici) KACC92031P를 사용할 경우, 아세틸콜린에스터레이즈 저해활성, 안지오텐신 전환 효소 저해활성 또는 항균활성을 높이기 위해 28 내지 32℃에서 7일 이상 발효하는 것이 바람직하다. 보다 바람직하게는 7 ~ 10일간 발효하는 것이 좋다.According to the present invention, when Pediococcus acidilactici KACC92031P is used as a lactic acid bacterium, fermentation for more than 7 days at 28 to 32 ° C in order to enhance acetylcholine esterase inhibitory activity, angiotensin converting enzyme inhibitory activity or antimicrobial activity . More preferably 7 to 10 days.
본 발명에 따르면, 젖산균으로 류코노스톡 메센테로이데스(Leuconostoc mesenteroides) KCTC3505를 사용할 경우, 안지오텐신 전환 효소 저해활성 또는 항균활성을 높이기 위해 28 내지 32℃에서 3 ~ 7일간 발효하는 것이 바람직하다.According to the present invention, when Leuconostoc mesenteroides KCTC3505 is used as lactic acid bacteria, fermentation is preferably performed at 28 to 32 ° C for 3 to 7 days to enhance angiotensin converting enzyme inhibitory activity or antimicrobial activity.
본 발명의 다른 양태에 따르면, 본 발명은 감귤을 분쇄하는 단계; 및 감귤 분쇄물에 락토바실러스 사케이(Lactobacillus sakei) KACC92032P, 페디오코커스 아시딜락티시(Pediococcus acidilactici) KACC92031P 및 류코노스톡 메센테로이데스(Leuconostoc mesenteroides) KCTC3505로 이루어진 군에서 선택된 젖산균을 접종하여 20 내지 40℃에서 발효하는 단계;를 포함하는 아세틸콜린에스터레이즈 저해활성, 안지오텐신 전환 효소 저해활성 또는 항균활성을 갖는 감귤 발효물 제조방법을 제공한다.According to another aspect of the present invention, Lactobacillus sakei KACC92032P, Pediococcus acidilactici KACC92031P, and Leuconostoc ( Lactobacillus sisteri) were added to citrus pulverized product, mesenteroides KCTC3505 and fermenting at 20 to 40 占 폚. The present invention also provides a method for producing a citrus fermented product having an inhibitory activity against angiotensin converting enzyme or an antimicrobial activity.
본 발명의 또 다른 양태에 따르면, 본 발명은 상기 제조방법으로 제조된 아세틸콜린에스터레이즈 저해활성, 안지오텐신 전환 효소 저해활성 또는 항균활성을 갖는 감귤 발효물을 제공한다.According to another aspect of the present invention, there is provided a citrus fermented product having the acetylcholine esterase inhibiting activity, angiotensin converting enzyme inhibiting activity or antimicrobial activity produced by the above production method.
본 발명의 또 다른 양태에 따르면, 본 발명은 상기 감귤 발효물을 포함하는 식품 조성물을 제공한다.According to another aspect of the present invention, the present invention provides a food composition comprising the citrus fermented product.
본 발명의 식품 조성물은 알츠하이머 질환의 예방 또는 개선용 기능성 식품, 고혈압의 예방 또는 개선용 기능성 식품, 또는 항균용 기능성 식품일 수 있다.The food composition of the present invention may be a functional food for preventing or improving Alzheimer's disease, a functional food for preventing or improving hypertension, or a functional food for antibacterial activity.
본 발명에서 아세틸콜린에스터레이즈 저해활성은 아세틸콜린에스터레이즈가 아세틸콜린(acetylcholine, ACh)을 아세테이트(acetate)와 콜린(choline)로 분해하는 작용을 저해하는 활성을 의미한다.In the present invention, the acetylcholine esterase inhibitory activity means an activity of inhibiting the action of acetylcholine esterase to decompose acetylcholine (ACh) into acetate and choline.
본 발명에서 안지오텐신 전환 효소 저해활성은 안지오텐신 전환 효소가 안지오텐신 I(angiotensin I)을 안지오텐신 II(angiotensin II)로 전환하는 작용을 저해하는 활성을 의미한다.In the present invention, the angiotensin converting enzyme inhibitory activity refers to an activity of inhibiting the action of angiotensin converting enzyme to convert angiotensin I to angiotensin II.
본 발명에서 항균활성은 균의 생장을 억제 또는 지연시키거나 균을 사멸시키는 활성을 의미한다.In the present invention, the antimicrobial activity refers to an activity of inhibiting or delaying the growth of bacteria or killing bacteria.
본 발명은 본 발명에 따른 감귤 발효물 그 자체 또는 식품학적으로 허용된 담체를 혼합한 조성물을 식이소재로 포함하는 식품을 제공하는데, 상기 식품은 식육가공품, 어육제품, 두부, 묵, 죽, 라면이나 국수 등의 면류, 간장, 된장, 고추장, 혼합장 등의 조미식품, 소스, 과자, 발효유나 치즈 등의 유가공품, 김치나 장아찌 등의 절임식품, 과실, 채소, 두유, 발효음료 등의 음료수의 식품에 포함하여 사용할 수 있다. 이는 본 발명의 분야에서 통상의 지식을 가진 자라면 자명한 사항으로 이의 구체적인 조리 방법이나 생산 방법의 기재는 생략하기로 한다. 또한 식품학적으로 허용된 담체는 약제학적으로 허용된 담체도 사용할 수 있다.
The present invention provides a food containing the citrus fermented product itself or a composition comprising a pharmaceutically acceptable carrier according to the present invention as a dietary material, wherein the food is selected from the group consisting of meat products, fish meat products, tofu, Such as noodles such as soy sauce, soy sauce, soy sauce, soy sauce, soybean paste, kochujang, mixed potatoes, sauces such as sauces, confectionaries, fermented milk and cheese, pickles such as kimchi and pickles, fruits, vegetables, soy milk, fermented beverages It can be used in food. It will be apparent to those skilled in the art that the specific cooking methods and production methods are not described herein. The pharmaceutically acceptable carrier may also be a pharmaceutically acceptable carrier.
본 발명에 따르면 감귤과 젖산균을 사용하여 아세틸콜린에스터레이즈 저해활성, 안지오텐신 전환 효소 저해활성 또는 항균활성이 우수한 감귤 발효물을 제조할 수 있다. 또한 이 감귤 발효물의 우수한 생리활성을 바탕으로 알츠하이머 질환과 고혈압을 예방하거나 개선할 수 있고 항균효과가 우수한 기능성 식품을 제조할 수 있다.
According to the present invention, citrus and lactic acid bacteria can be used to produce a citrus fermented product having an acetylcholine esterase inhibitory activity, angiotensin converting enzyme inhibiting activity or antimicrobial activity. In addition, based on the excellent physiological activity of the citrus fermented product, it is possible to prevent or ameliorate Alzheimer's disease and hypertension, and produce a functional food having an excellent antibacterial effect.
도 1은 각각 7가지 젖산균으로 발효한 감귤 발효물 시료(×10 희석)의 아세틸콜린에스터레이즈(Acetylcholinesterase, AChE) 저해활성 분석결과를 나타낸 그래프이다. a~g, w~z: 같은 그룹 bar 또는 같은 컬러 bar에서 서로 다른 알파벳은 유의적인 차이(P<0.05)가 있다는 것을 의미한다. 저해활성 수치(%)는 3반복 실험결과의 평균±표준편차로 나타내었다. UC: 발효하지 않은 감귤 시료, CWS: 균주를 접종하지 않고 발효한 감귤 시료, EL ~ No. 2: 각 균주를 접종하여 발효한 감귤 발효 추출물 시료, EL: Enterococcus lactis, EF: Enterococcus faecalis, LM: Leuconostoc mesenteroides(KCTC3505), LA: Lactobacillus acidophilus, LP: Lactobacillus plantarum, No. 1: Pediococcus sp .(KACC92031P), No. 2: Lactobacillus sp(KACC92032P).
도 2는 각각 7가지 젖산균으로 발효한 감귤 발효물 시료(×20 희석)의 안지오텐신 전환 효소(Angiotensin converting enzyme, ACE) 저해활성 분석결과를 나타낸 그래프이다. a-g, v-z: 같은 그룹 bar 또는 같은 컬러 bar에서 서로 다른 알파벳은 유의적인 차이(P<0.05)가 있다는 것을 의미한다. 저해활성 수치(%)는 3반복 실험결과의 평균±표준편차로 나타내었다. UC: 발효하지 않은 감귤 시료, CWS: 균주를 접종하지 않고 발효한 감귤 시료, EL ~ No. 2: 각 균주를 접종하여 발효한 감귤 발효 추출물 시료, EL: Enterococcus lactis, EF: Enterococcus faecalis, LM: Leuconostoc mesenteroides(KCTC3505), LA: Lactobacillus acidophilus, LP: Lactobacillus plantarum, No. 1: Pediococcus sp .(KACC92031P), No. 2: Lactobacillus sp(KACC92032P).
도 3은 본 발명의 실시예에서 사용한 젖산균 중 김치에서 분리한 Pediococcus sp .(No. 2)(KACC92031P)의 16S rRNA 서열분석 결과를 통해 작성한 계통수(Phylogenetic tree)이다.
도 4는 본 발명의 실시예에서 사용한 젖산균 중 김치에서 분리한 Lactobacillus sp .(No. 2)(KACC92032P)의 16S rRNA 서열분석 결과를 통해 작성한 계통수(Phylogenetic tree)이다.FIG. 1 is a graph showing the results of analysis of acetylcholinesterase (AChE) inhibitory activity of a citrus fermented product (× 10 dilution) fermented with seven kinds of lactic acid bacteria. a ~ g, w ~ z: Different alphabets in the same group bar or the same color bar mean that there is a significant difference (P <0.05). The inhibitory activity (%) was expressed as the mean ± standard deviation of the results of three replicates. UC: citrus sample not fermented, CWS: citrus sample fermented without inoculation, EL ~ No. 2: ELA: Enterococcus lactis , EF: Enterococcus faecalis , LM: Leuconostoc mesenteroides (KCTC3505), LA: Lactobacillus acidophilus , LP: Lactobacillus plantarum , No. 1: Pediococcus sp . (KACC92031P), No. 2: Lactobacillus sp (KACC92032P) .
FIG. 2 is a graph showing the results of assay of an angiotensin converting enzyme (ACE) inhibitory activity of a citrus fermented product (× 20 diluted) fermented with seven kinds of lactic acid bacteria, respectively. ag, vz: different alphabets in the same group bar or same color bar mean that there is a significant difference (P <0.05). The inhibitory activity (%) was expressed as the mean ± standard deviation of the results of three replicates. UC: citrus sample not fermented, CWS: citrus sample fermented without inoculation, EL ~ No. 2: A sample of fermented citrus fruit fermented by inoculation with each strain, EL: Enterococcus lactis , EF: Enterococcus faecalis , LM: Leuconostoc mesenteroides (KCTC3505), LA: Lactobacillus acidophilus , LP: Lactobacillus plantarum , No. 1: Pediococcus sp . (KACC92031P), No. 2: Lactobacillus sp (KACC92032P) .
Fig. 3 is a graph showing the results of Pediococcus sp . Lactic acid bacteria isolated from kimchi among the lactic acid bacteria used in the examples of the present invention . (SEQ ID NO: 2) (KACC92031P), which is a phylogenetic tree.
Fig. 4 is a graph showing the effect of Lactobacillus sp . (SEQ ID NO: 2) (KACC92032P), which is a phylogenetic tree.
이하, 실시예를 통하여 본 발명을 더욱 상세히 설명하기로 한다. 이들 실시예는 단지 본 발명을 예시하기 위한 것이므로, 본 발명의 범위가 이들 실시예에 의해 제한되는 것으로 해석되지는 않는다.
Hereinafter, the present invention will be described in more detail with reference to Examples. These embodiments are only for illustrating the present invention, and thus the scope of the present invention is not construed as being limited by these embodiments.
실시예Example 1. 감귤의 젖산균 발효 1. Fermentation of lactic acid bacteria of citrus fruit
1-1. 재료1-1. material
감귤은 2014년 7월에 제주도에서 시판되는 온주밀감(Citrus unshiu)을 구입하여 사용하였다. 또한 acetylthiocholin esterase, acetylthiocholine iodide, rabbit lung acetone powder, hipouril histidine leucicne(HHL), eserine은 Sigma-Aldrich(St. Louis, MO, USA)에서 구입하였다. Sodium carbonate anhydrous, sodium bicarbonate는 Daejung Chemical Co.(Siheung, Korea)의 제품을 사용하였다. Folin-Ciocalteu reagent은 Wako Pure Chemical Industries, Ltd.(Osaka, Japan)에서 구입하였다.
Tangerine was purchased in July 2014 from Citrus unshiu (Citrus unshiu), which is commercially available from Jeju Island. Acetylthiocholin esterase, acetylthiocholine iodide, rabbit lung acetone powder, hypouril histidine leucicne (HHL) and eserine were purchased from Sigma-Aldrich (St. Louis, Mo., USA). Sodium carbonate anhydrous and sodium bicarbonate were purchased from Daejung Chemical Co. (Siheung, Korea). Folin-Ciocalteu reagent was purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan).
1-2. 사용균주 및 배양조건1-2. The strains used and culture conditions
감귤 발효를 위해 미생물자원센터(Korea Collection for Type Cultures, Daejeon, Korea)에서 5종의 균주 Lactobacillus acidophilus KCTC 3140(LA), Lactobacillus plantarum KCTC 3104 (LP), Enterococcus lactis KCTC 21015 (EL), Enterococcus faecalis KCTC 2011 (EF), Leuconostoc mesenteroides KCTC 3505 (LM)를 분양받아 이용하였다. 또한, 김치에서 분리한 젖산균 Pediococcus sp . (No. 1)(KACC92031P) 및 Lactobacillus sp . (No. 2)(KACC92032P)도 활용하였다. 모든 균주의 전배양은 MRS 액체배지(Difco, Detroit, MI, USA)에서 30℃, 100rpm, 2일간 진탕배양하여 활성화시키고 감귤퓨레에 접종하였다.
For the citrus fermentation, five strains of Lactobacillus acidophilus KCTC 3140 (LA), Lactobacillus plantarum KCTC 3104 (LP), Enterococcus lactis KCTC 21015 (EL), Enterococcus faecalis KCTC 2011 (EF) and Leuconostoc mesenteroides KCTC 3505 (LM) were purchased and used. In addition, lactic acid bacteria Pediococcus sp . (No. 1) (KACC92031P) and Lactobacillus sp . (No. 2) (KACC92032P) was also utilized. All cultures of all strains were activated by shaking culture in MRS liquid medium (Difco, Detroit, MI, USA) at 30 ° C, 100 rpm for 2 days and inoculated into citrus fruit purée.
1-3. 감귤 발효물 제조1-3. Manufacture of citrus fermented products
감귤을 수돗물에 깨끗이 세척한 후 증류수로 다시 세척하였다. 발효 과정 중 오염을 방지하기 위하여 세척된 감귤 표면에 UV를 15분간 조사하여 멸균하였다. 믹서기로 감귤을 고르게 분쇄하고 미리 멸균된 플라스크에 담아 270㎖의 감귤퓨레를 준비하였다.The citrus fruits were cleanly washed in tap water and then washed again with distilled water. To prevent contamination during the fermentation process, the washed citrus surface was irradiated with UV for 15 minutes and sterilized. The citrus fruits were evenly ground with a blender and placed in a pre-sterilized flask to prepare 270 ml of citrus puree.
Lactobacillus acidophilus KCTC 3140(LA), Lactobacillus plantarum KCTC 3104(LP), Enterococcus lactis KCTC 21015(EL), Enterococcus faecalis KCTC 2011(EF), Leuconostoc mesenteroides KCTC 3505(LM), Pediococcus sp.(No. 1), Lactobacillus sp.(No. 2)의 총 7종의 균주를 각각 MRS 액체배지에서 전배양한 다음 전배양액을 각각 30㎖씩 준비된 감귤퓨레에 첨가하여 접종하였다. 대조군으로는 균주 전배양액 대신 멸균 증류수를 사용하였다. 각 균주를 감귤퓨레에 접종한 후 30℃, 100rpm, 진탕 배양기에서 3, 5, 7, 그리고 10일간 발효시킨 다음 발효물을 회수하였다. Lactobacillus acidophilus KCTC 3140 (LA), Lactobacillus plantarum KCTC 3104 (LP), Enterococcus lactis KCTC 21015 (EL), Enterococcus faecalis KCTC 2011 (EF), Leuconostoc mesenteroides KCTC 3505 (LM), Pediococcus sp. (No. 1), Lactobacillus sp. (No. 2) before the culture was then inoculated in addition to the citrus puree prepared by each 30㎖ the pre-culture in a MRS liquid medium, each of the strains of 7 species. As a control group, sterilized distilled water was used instead of the strain pre-culture solution. Each strain was inoculated on citrus puree and fermented at 30 ° C, 100 rpm, shaking incubator for 3, 5, 7, and 10 days, and then the fermented product was recovered.
회수한 발효물을 원심분리(4℃, 10분, 5,000×g)하고 상징액을 Whatman No.1 여과지(Advantec, Tokyo, Japan)로 통과시킨 여과액을 다음 실험의 시료로 사용하였다.
The recovered fermented product was centrifuged (4 ° C, 10 min, 5,000 × g) and the supernatant was passed through Whatman No.1 filter paper (Advantec, Tokyo, Japan).
실시예 2. 감귤 발효물의 생리활성 분석Example 2. Analysis of physiological activities of citrus fermented products
2-1. 실험방법2-1. Experimental Method
2-1-1. Acetylcholinesterase(AChE) 저해능 분석2-1-1. Acetylcholinesterase (AChE) Incompatibility Analysis
AChE 저해능은 acetylcholine iodide를 기질로 사용하는 Prashanth의 방법을 변형하여 측정하였다. 시료 90㎕에 AChE(2U/㎖) 30㎕와 DTNB 용액(3.96㎎의 DTNB와 1.5㎎의 sodium bicarbonate가 10㎖의 phosphate buffer(pH 7.7)에 용해된 용액) 160㎕를 혼합한 후, 25℃에서 5분간 반응시켰다. 반응물에 acetylcholine iodide 용액(10.85㎎의 acetylthiocholine iodide가 5㎖의 phosphate buffer(pH 7.7)에 용해된 용액) 30㎕를 가하고 25℃에서 30분간 반응시킨 다음 Multiplate Reader(Sunrise RC/TS/TS Color-TC/TW/BC/6Filter, Tecan Austria GmbH, Gradig, Austria)를 이용하여 405nm에서 흡광도를 측정하고, 다음 계산식을 사용하여 AChE 저해능을 구하였다.AChE inhibition was measured by using Prashanth 's method using acetylcholine iodide as a substrate. To 90 μl of the sample was added 160 μl of AChE (2 U / ml) and 30 μl of DTNB solution (3.96 mg of DTNB and 1.5 mg of sodium bicarbonate in 10 ml of phosphate buffer (pH 7.7)), For 5 minutes. 30 μl of acetylcholine iodide solution (10.85 mg of acetylthiocholine iodide dissolved in 5 ml of phosphate buffer (pH 7.7)) was added to the reaction mixture, followed by reaction at 25 ° C for 30 minutes. / TW / BC / 6Filter, Tecan Austria GmbH, Gradig, Austria), and the AChE inhibition was determined using the following equation.
AChE 저해능(%) = 1 - (시료 첨가구의 흡광도 / 무처리구의 흡광도)
AChE inhibition (%) = 1 - (absorbance of sample added / absorbance of untreated)
2-1-2. Angiotensin converting enzyme(ACE) 저해능 분석2-1-2. Angiotensin converting enzyme (ACE) inhibitory assay
ACE 저해능은 Cushman과 Cheung의 방법에 따라 측정하였다. 50mM sodium borate buffer(pH 8.3) 20㎖에 1g의 rabbit lung acetone powder를 4℃에서 24시간 동안 교반한 후, 10분간 원심분리(4℃, 5,000×g)하여 ACE 조효소액을 얻었다. 희석한 시료 50㎕에 ACE 조효소액 50㎕를 가하여 37℃에서 10분간 방치한 후, 25mM HHL 100㎕를 첨가하여 37℃에서 60분간 반응시켰다. 1M HCI 250㎕를 가하여 30초간 교반하고 원심분리(4℃, 10분, 5,000×g)하여 상징액 200㎕를 얻었다. 이 상징액을 80℃에서 30분간 완전히 건조시키고 증류수 1㎖를 넣은 후에 228nm에서 흡광도를 측정하였다. 대조군은 시료 대신 증류수를 가해 실험하였으며, 양성 대조군으로 captopril을 사용하였다. ACE 저해능은 다음 계산식을 이용하여 구하였다.ACE inhibition was measured by the method of Cushman and Cheung. 1 g of rabbit lung acetone powder was added to 20 ml of 50 mM sodium borate buffer (pH 8.3) at 4 ° C for 24 hours, and centrifuged at 4 ° C for 5 minutes to obtain ACE enzyme solution. 50 AC of the diluted sample was added with 50 AC of ACE enzyme solution, and the mixture was allowed to stand at 37 캜 for 10 minutes. Then, 100 25 of 25 mM HHL was added and reacted at 37 캜 for 60 minutes. 250 μl of 1M HCI was added and the mixture was stirred for 30 seconds and centrifuged (4 ° C, 10 minutes, 5,000 × g) to obtain 200 μl of a supernatant. The supernatant was completely dried at 80 ° C for 30 minutes, and after adding 1 ml of distilled water, the absorbance was measured at 228 nm. In the control group, distilled water was used instead of the sample, and captopril was used as a positive control. The ACE inhibition was calculated using the following equation.
ACE 저해능(%) = 1 - (시료 첨가구의 흡광도 / 무처리구의 흡광도)
ACE inhibition (%) = 1 - (absorbance of sample added / untreated)
2-1-3. 항균 활성 분석2-1-3. Antimicrobial activity analysis
각 젖산균으로 발효한 감귤 발효물을 3, 5, 7 그리고 10일째의 발효 기간에 분취하여 원심분리한 후 여과하여 상징액을 얻었다. 0.5% yeast extract agar plate 위에 지시균주배양액 1%와 0.1M phosphate buffer(pH 7.0)를 포함한 0.8% NB soft agar 10㎖를 중층하여 test plate를 제조하였다. 감귤 발효 추출물 상징액 50㎕를 test plate에 spotting하고 상징액을 건조한 후, 37℃ 항온 배양기에서 12시간 이상 배양하여 시험균주에 대한 투명환 형성 유무를 조사하여 항균 활성을 측정하였다.
The fermented citrus fruits fermented with each lactic acid bacterium were collected during fermentation on
2-1-4. 통계처리2-1-4. Statistical processing
모든 실험은 3회 반복으로 이루어졌으며, 그 결과를 SPSS software(Ver. 12.0K, SPSS Inc., Chicago, IL, USA)로 처리하였다. 각 측정군의 평균과 표준편차를 산출하고 Duncan의 다중범위 검정(Duncan's multiple range test)으로 유의성을 검정하였다. 모든 처리값의 차이는 신뢰 수준 95%(P<0.05)로 비교 분석하였다.
All experiments were repeated 3 times and the results were processed with SPSS software (Ver. 12.0K, SPSS Inc., Chicago, IL, USA). The mean and standard deviation of each measurement group were calculated and the significance was tested by Duncan's multiple range test. The difference of all treatment values was compared with 95% confidence level (P <0.05).
2-2. 실험결과2-2. Experiment result
2-2-1. Acetylcholinesterase(AChE) 저해능 분석결과2-2-1. Results of Acetylcholinesterase (AChE)
체내의 신경전달물질 중 하나인 acetylcholine(ACh)은 acetyl Co A와 choline이 choline acetyltransferase(ChAT) 효소의 작용을 받아 합성되며 이것은 다시 AChE 효소의 작용을 받아 acetate와 choline으로 분해된다. 따라서 체내의 신경전달이 원활하게 이루어지려면 ACh에 영향을 미치는 ChAT와 AChE 효소의 활성이 중요하다. 치매의 50%를 차지하고 있는 Alzheimer disease(AD) 환자의 경우는 ChAT가 감소하는 경향을 나타내고 있으며 또한 AD 환자의 치료에 있어 AChE를 억제해 체내의 ACh의 농도를 증가시키는 방법이 이용되고 있다.Acetylcholine (ACh), one of neurotransmitters in the body, is synthesized by acetyl CoA and choline under the action of choline acetyltransferase (ChAT) enzyme, which is decomposed into acetate and choline under the action of AChE enzyme. Therefore, the activation of ChAT and AChE enzymes, which affect ACh, is important for smooth neurotransmission in the body. In patients with Alzheimer disease (AD), which accounts for 50% of dementia, ChAT tends to decrease and AChE is inhibited in patients with AD to increase the concentration of ACh in the body.
감귤 발효 추출물을 10배 희석한 시료의 acetylcholinesterase(AChE) 저해 활성을 분석한 결과를 도 1에 나타내었다. 양성 대조군인 eserine은 100㎍/㎖ 농도에서 87%의 저해활성을 보였으며 No. 1(Pediococcus sp.) 및 No. 2(Lactobacillus sp.) 균주 발효 추출물이 다른 추출물에 비해 발효가 진행될수록 높은 저해 활성이 나타났다. 보통 AChE 저해제로 donepezil, tacrine, rivastigmine, galactamine 등이 FDA에 허용되어 있으며 coumarine 계의 페놀 물질이 galactamine과 유사한 방식으로 AChE와 결합하여 저해한다고 보고되어 있다. 따라서 DPPH와 ABTS 라디칼 소거 활성의 결과와는 다른 양상을 보이는데 이는 항산화능과 AChE 저해 활성에 영향을 미치는 물질에 차이가 있음을 의미한다.
The results of assaying acetylcholinesterase (AChE) inhibitory activity of a 10-fold diluted citrus fermented extract are shown in FIG. The positive control group, eserine, showed 87% inhibition activity at 100 μg / ml concentration. 1 ( Pediococcus sp. ) And No. 2 ( Lactobacillus sp. ) Fermented extract showed higher inhibitory activity as fermentation progressed than other extracts. It has been reported that donepezil, tacrine, rivastigmine, galactamine, etc., as AChE inhibitors, are allowed in the FDA and that coumarine phenol substances inhibit AChE in a manner similar to galactamine. Therefore, the results of DPPH and ABTS radical scavenging activities are different from those of DPPH and ABTS radical scavenging activity, which means that there are differences in the substances that affect antioxidant activity and AChE inhibitory activity.
2-2-2. Angiotensin converting enzyme(ACE) 저해능 분석결과2-2-2. Angiotensin converting enzyme (ACE) inhibitory assay results
인체 내에 존재하는 renin은 angiotensinogen을 angiotensin Ⅰ으로 분해하고, 비활성의 angiotensin Ⅰ은 angiotensin-converting enzyme(ACE)에 의해 혈관 수축작용이 있는 angiotensin Ⅱ가 된다. 이 angiotensin Ⅱ는 동맥 혈관을 수축하여 혈압을 상승시키고, 부신에서 aldosterone의 분비를 촉진하여 신장의 나트륨 및 수분의 재흡수를 증가시킴으로써 고혈압 발병에 관여한다.Renin in the body decomposes angiotensinogen into angiotensin Ⅰ and inactive angiotensin Ⅰ becomes angiotensin Ⅱ with angiotensin-converting enzyme (ACE). This angiotensin Ⅱ is involved in the onset of hypertension by contracting the arterial blood vessels, raising blood pressure, and promoting the secretion of aldosterone from the adrenal gland, thereby increasing renal sodium and water reabsorption.
20배 희석한 감귤 발효 추출물 시료에 대한 ACE 저해 활성을 도 2에 나타내었다. ACE 저해능은 4.62±2.71 ~ 86.22±1.57%의 범위를 보이며 20배 희석한 추출물임에도 높은 저해능을 보였고 양성 대조군인 captopril은 2㎍/㎖ 농도에서 91%의 저해능을 나타내었다. LA(L. acidophilus) 발효 추출물은 오히려 발효기간이 길어질수록 38.17%에서 14.19%로 감소하였으나, No. 1(Pediococcus sp.), No. 2(Lactobacillus sp.) 발효 추출물은 발효가 진행될수록 42.90%에서 85.97%, 41.94%에서 83.88%로 ACE 저해능이 증가하였다. 이전 Lee 등이 발표한 부추 당침액의 젖산균 발효에 따른 항산화 및 ACE 저해능 연구 결과에 따르면, 젖산균 종류에 따라 50.4 ~ 67%의 ACE 저해능을 보였는데, 사용된 젖산균의 종류가 다르지만 본 발명에서 더 높은 저해능을 보이는 것을 확인할 수 있었다. 또한 ACE 저해능은 주로 단백질이나 펩티드에 의해 유발되는데 젖산균을 이용하여 발효한 감귤 추출물에서 생성된 산물이 단백질, 펩티드로 이루어졌을 가능성이 커 보이며 이로 인해 ACE 저해능이 우수하게 나타난 것으로 사료된다.
The ACE inhibitory activity of 20-fold diluted citrus fermented extract samples is shown in FIG. The ACE inhibitory effect was 4.62 ± 2.71 ~ 86.22 ± 1.57%. The inhibitory effect of captopril was 91% at 2 μg / ㎖. LA ( L. acidophilus ) fermented extract decreased from 38.17% to 14.19% as the fermentation period became longer. 1 ( Pediococcus sp. ), No. 2 ( Lactobacillus sp. ) Fermented extracts increased from 42.90% to 85.97% and from 41.94% to 83.88%, respectively. According to Lee et al.'S results of antioxidative and ACE inhibition studies on the fermentation of lactic acid bacteria in leek slurries, leucine fermentation showed 50.4 ~ 67% ACE inhibition depending on the type of lactic acid bacteria. Although the kinds of lactic acid bacteria used are different, And it was confirmed that it showed an inferiority. In addition, ACE inhibition is mainly caused by proteins or peptides. It is likely that the products produced from fermented citrus extracts using lactic acid bacteria are composed of proteins and peptides, and as a result, ACE inhibition is excellent.
2-2-3. 항균 활성 분석결과2-2-3. Antimicrobial activity analysis result
병원성 미생물 L. delbrueckii, B. subtilis, E. coli, S. aureus, E. faecium, X. citri, B. cereus, S. typhimurium, M. luteus, P. acidilactici, B. sphaericus, B. megaterium에서 항균성을 조사했고 결과는 표 1 내지 3과 같다. 대조군의 경우 실험에 사용된 모든 균주에 대해 항균 활성이 나타나지 않았으며 LP(L. plantarum), LM(L. mesenteroides), No. 1(Pediococcus sp.), No. 2(Lactobacillus sp.) 발효 추출물을 제외하고는 항균 활성이 확인되지 않았다. 이 4종의 균주 발효 추출물은 발효 기간이 증가될수록 항균 활성을 보였으며, LP(L. plantarum) 발효 추출물은 B. subtilis, E. coli, S. aureus, E. faecium, X. citri, B. cereus, S. typhimurium, M.luteus, P. acidilactici, B. sphaericus, B. megaterium에서 LM(L. mesenteroides) 발효 추출물은 E. coli, X. citri, B. cereus, S. typhimurium, P. acidilactici, B. sphaericus, B. megaterium에서 활성을 확인할 수 있었고, No. 1(Pediococcus sp.) 발효 추출물은 L. delbrueckii, M. luteus, No. 2(Lactobacillus sp.) 발효 추출물은 L. delbrueckii을 제외한 병원성 미생물에 대해 항균 활성을 보였다. 특히 No. 2(Lactobacillus sp.) 발효 추출물에서 강한 항균 활성을 나타내었다. No. 2(Lactobacillus sp.)에 감귤 발효 시 항균 활성은 대사산물로 항균물질을 생산하는 것으로 여겨지며 이때 생산되어지는 물질에 대한 연구가 더 진행되어야 할 것이다. Cho 등은 자연 발효한 오미자 발효액에서 원래 항균 활성이 없었으나 발효에 의해 오미자액이 E. coli 균주에 대해 항균 활성을 나타내었다고 보고한 바 있다.The pathogenic microorganisms L. delbrueckii , B. subtilis , E. coli , S. aureus , E. faecium , X. citri , B. cereus , S. typhimurium , M. luteus , P. acidilactici , B. sphaericus , B. megaterium The antibacterial properties were examined and the results are shown in Tables 1 to 3. In the control group, no antimicrobial activity was observed for all the strains used in the experiment, and LP ( L. plantarum ), LM ( L. mesenteroides ), and No. 1 ( Pediococcus sp. ), No. 2 ( Lactobacillus sp. ) Fermented extract. E. coli , S. aureus , E. faecium , X. citri , and B. subtilis extracts of fermented LP ( L. plantarum ) showed higher antimicrobial activity with increasing fermentation period . cereus, S. typhimurium, M.luteus, P. acidilactici, B. sphaericus, B. in megaterium LM (L. mesenteroides) fermented extract of E. coli, X. citri, B. cereus , S. typhimurium, P. acidilactici , B. sphaericus , and B. megaterium . 1 ( Pediococcus sp. ) Fermented extracts were identified as L. delbrueckii , M. luteus , N. 2 ( Lactobacillus sp. ) Fermented extract showed antimicrobial activity against pathogenic microorganisms except L. delbrueckii . Especially No. 2 ( Lactobacillus sp. ) Fermented extract showed strong antimicrobial activity. No. 2 ( Lactobacillus sp. ), The antimicrobial activity of citrus fruit is considered to produce antimicrobial substance as a metabolite. Cho et al. Reported that the natural fermented Omija fermentation broth had no antimicrobial activity, but the fermentation showed an antimicrobial activity against the E. coli strain.
L. delbrueckii
L. delbrueckii
B. subtilis
B. subtilis
E. coli
E. coli
S. aureus
S. aureus
1) CWS: 균주를 접종하지 않고 발효한 감귤 시료, EL ~ No. 2: 각 균주를 접종하여 발효한 감귤 발효 추출물 시료, EL: Enterococcus lactis, EF: Enterococcus faecalis, LM: Leuconostoc mesenteroides, LA: Lactobacillus acidophilus, LP: Lactobacillus plantarum, No. 1: Pediococcus sp., No. 2: Lactobacillus sp. 1) CWS: citrus sample fermented without inoculation of strains, EL ~ No. 2: A sample of fermented citrus fruit fermented by inoculation with each strain, EL: Enterococcus lactis , EF: Enterococcus faecalis , LM: Leuconostoc mesenteroides , LA: Lactobacillus acidophilus , LP: Lactobacillus plantarum , No. 1: Pediococcus sp. , No. 2: Lactobacillus sp.
2) -: 항균활성 검출되지 않음, 항균활성: +, < 1.0mm; ++, 1.0 ~ 1.5mm; +++, > 1.5mm.
2) -: Antibacterial activity not detected, Antimicrobial activity: +, <1.0 mm; ++, 1.0 to 1.5 mm; +++, > 1.5mm.
E. faecium
E. faecium
X. citri
X. citri
B. cereus
B. cereus
S. typhimurium
S. typhimurium
1) CWS: 균주를 접종하지 않고 발효한 감귤 시료, EL ~ No. 2: 각 균주를 접종하여 발효한 감귤 발효 추출물 시료, EL: Enterococcus lactis, EF: Enterococcus faecalis, LM: Leuconostoc mesenteroides, LA: Lactobacillus acidophilus, LP: Lactobacillus plantarum, No. 1: Pediococcus sp., No. 2: Lactobacillus sp. 1) CWS: citrus sample fermented without inoculation of strains, EL ~ No. EL: Enterococcus lactis , EF: Enterococcus faecalis , LM: Leuconostoc mesenteroides , LA: Lactobacillus acidophilus , LP: Lactobacillus plantarum . 1: Pediococcus sp. , No. 2: Lactobacillus sp.
2) -: 항균활성 검출되지 않음, 항균활성: +, < 1.0mm; ++, 1.0 ~ 1.5mm; +++, > 1.5mm.
2) -: Antibacterial activity not detected, Antimicrobial activity: +, <1.0 mm; ++, 1.0 to 1.5 mm; +++, > 1.5mm.
M. luteus
M. luteus
P. acidilactici
P. acidilactici
B. sphaericus
B. sphaericus
B. megaterium
B. megaterium
1) CWS: 균주를 접종하지 않고 발효한 감귤 시료, EL ~ No. 2: 각 균주를 접종하여 발효한 감귤 발효 추출물 시료, EL: Enterococcus lactis, EF: Enterococcus faecalis, LM: Leuconostoc mesenteroides, LA: Lactobacillus acidophilus, LP: Lactobacillus plantarum, No. 1: Pediococcus sp., No. 2: Lactobacillus sp. 1) CWS: citrus sample fermented without inoculation of strains, EL ~ No. EL: Enterococcus lactis , EF: Enterococcus faecalis , LM: Leuconostoc mesenteroides , LA: Lactobacillus acidophilus , LP: Lactobacillus plantarum . 1: Pediococcus sp. , No. 2: Lactobacillus sp.
2) -: 항균활성 검출되지 않음, 항균활성: +, < 1.0mm; ++, 1.0 ~ 1.5mm; +++, > 1.5mm.
2) -: Antibacterial activity not detected, Antimicrobial activity: +, <1.0 mm; ++, 1.0 to 1.5 mm; +++, > 1.5mm.
실시예 3. 젖산균 동정Example 3 Identification of lactic acid bacteria
기능성 분석 결과 높은 활성을 보인 균주들(No. 1 및 2)을 16S rRNA 유전자 분석으로 동정하였다. Colony 형태로 균주를 솔젠트사(대전)에 송부하여 유전자 분석을 의뢰하였다. 16S rRNA sequencing에 사용되는 universal primer인 27F primer(5'-AGAGTTTGATCCTGGCTCAG-3')와 1492R primer(5'-GGTTACCTTGTTACGACTT-3')를 이용하여 PCR을 실시하였다. Applied Biosystems사의 3730XL Capillary RNA Sequencer를 이용하여 염기서열을 분석하였고, 분석된 염기서열을 이용하여 NCBI(National Center for Biotechnology Information, http://www.ncbi.nlm.nih.gov/)의 blast search를 통해 상동성을 조사하여 동정하였다.As a result of functional analysis, highly active strains (No. 1 and No. 2) were identified by 16S rRNA gene analysis. Colony type strain was sent to Solent (Daejeon) for genetic analysis. PCR was performed using the universal primer 27F primer (5'-AGAGTTTGATCCTGGCTCAG-3 ') and 1492R primer (5'-GGTTACCTTGTTACGACTT-3') used for 16S rRNA sequencing. The nucleotide sequence was analyzed using a 3730XL Capillary RNA Sequencer from Applied Biosystems, and a blast search of NCBI (National Center for Biotechnology Information, http://www.ncbi.nlm.nih.gov/) was performed using the analyzed nucleotide sequence Were identified and identified.
이의 결과 No. 1 균주의 16S rRNA는 Pediococcus acidilactici)와 No. 2 균주의 16S rRNA는 Lactobacillus sakei와 99% 이상의 상동성을 보였다. 그리고 이들 균주를 국립농업과학원 농업유전자원센터에 기탁하여 각각 KACC92031P와 KACC92032P의 수탁번호를 부여받았다.Result No. One strain of 16S rRNA was identified as Pediococcus lt ; / RTI > 2 strains of 16S rRNA were found in Lactobacillus and 99% or more homology with sakei . These strains were deposited with the National Institute of Agricultural Science and Technology, National Institute of Agricultural Science and Technology, and were assigned accession numbers of KACC92031P and KACC92032P, respectively.
L. sakei는 당으로부터 알코올 또는 유산균을 생성하는 젖산균으로 sakacin이라는 박테리오신을 생성한다. 박테리오신은 펩타이드, 단백질로 구성된 항균성 물질로 같은 환경에서 자라는 다른 미생물에 대한 경쟁적인 저해제로 다양한 미생물에서 분리된다. 특히 유산균은 김치 발효에 중요한 역할을 하며 다양한 항균물질을 생산하는 것으로 알려졌으며 박테리오신의 항균 범위는 매우 광범위하여 부패균, 식중독균, 전염병균이나 포자형성균 등의 증식을 억제하거나 사멸시키는데 효과적이다. 박테리오신은 항생제와 달리 단백질로 구성되어 인체에 들어오면 소화 기관 내에서 단백질 가수분해효소에 의해 쉽게 분해되어 인체에 무독성이고 잔류성이 없다. 항생제는 2차 대사산물인데 비해 박테리오신은 자신의 유전자로부터 직접 생합성 된다. L. sakei가 생산하는 sakacin은 A, G, K, P, Q 형이 보고되어 있는데, 그중에서 A형과 P 형이 잘 알려져 있다. 본 실시예에서 확인된 L. sakei가 생산하는 sakacin은 저해 범위가 좁은 박테리오신이 아니라 Streptococcus lactis가 생성하는 박테리오신인 nisin과 같이 저해범위가 매우 넓은 것을 알 수 있었다.
L. sakei is a lactic acid bacterium that produces alcohol or lactic acid bacteria from sugars and produces a bacteriocin called sakacin. Bacteriocin is an antimicrobial substance composed of peptides and proteins. It is a competitive inhibitor against other microorganisms that grow in the same environment and is isolated from various microorganisms. Especially, lactic acid bacteria plays an important role in kimchi fermentation and is known to produce various antimicrobial substances. Bacteriocin has a very broad antibacterial range and is effective in suppressing or killing the growth of spoilage bacteria, food poisoning bacteria, infectious disease bacteria and spore forming bacteria. Unlike antibiotics, bacteriocin is made up of proteins and is easily degraded by protein hydrolytic enzymes in the digestive tract when it enters the body, making it non-toxic and non-persistent to the human body. Antibiotics are secondary metabolites, whereas bacteriocins are biosynthesized directly from their genes. Sakacin, produced by L. sakei, has been reported as A, G, K, P and Q type, of which A and P type are well known. Sakacin produced by L. sakei identified in this Example is not a bacteriocin with a narrow inhibition range but a broad inhibition range such as nisin, a bacteriocin produced by Streptococcus lactis .
Claims (8)
상기 발효는 20 내지 40℃에서 하루 이상 이루어지는 것을 특징으로 하는 감귤 발효물 제조방법.The method according to claim 1,
Wherein the fermentation is performed at 20 to 40 占 폚 for one day or more.
감귤 분쇄물에 락토바실러스 사케이(Lactobacillus sakei) KACC92032P, 페디오코커스 아시딜락티시(Pediococcus acidilactici) KACC92031P 및 류코노스톡 메센테로이데스(Leuconostoc mesenteroides) KCTC3505로 이루어진 군에서 선택된 젖산균을 접종하여 20 내지 40℃에서 발효하는 단계;를 포함하는 아세틸콜린에스터레이즈 저해활성, 안지오텐신 전환 효소 저해활성 또는 항균활성을 갖는 감귤 발효물 제조방법.Crushing citrus fruits; And
The tangerine crushed material was mixed with Lactobacillus sakei ) KACC92032P, Pediococcus, Pediococcus acidilactici KACC92031P and Leuconostoc < RTI ID = 0.0 > mesenteroides KCTC3505 and fermenting at 20 to 40 占 폚. The method for producing a citrus fermented product according to claim 1, wherein the fermentation product has an inhibitory activity against angiotensin converting enzyme or an antimicrobial activity.
상기 식품 조성물은 알츠하이머 질환의 예방 또는 개선용 기능성 식품인 것을 특징으로 하는 식품 조성물.6. The method of claim 5,
Wherein the food composition is a functional food for preventing or ameliorating Alzheimer's disease.
상기 식품 조성물은 고혈압의 예방 또는 개선용 기능성 식품인 것을 특징으로 하는 식품 조성물.6. The method of claim 5,
Wherein the food composition is a functional food for preventing or improving hypertension.
상기 식품 조성물은 항균용 기능성 식품인 것을 특징으로 하는 식품 조성물.
6. The method of claim 5,
Wherein the food composition is a functional food for antibacterial activity.
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KR102224926B1 (en) * | 2019-11-26 | 2021-03-08 | 전남대학교산학협력단 | Lactobacillus sakei NY518 strain, culture thereof and enteric health improving functional food composition comprising the same |
KR20210089354A (en) * | 2020-01-08 | 2021-07-16 | 전남대학교산학협력단 | Method for preparing oligosaccharide and method for preparing composition for improving bowel health comprising the same |
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Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
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KR102224926B1 (en) * | 2019-11-26 | 2021-03-08 | 전남대학교산학협력단 | Lactobacillus sakei NY518 strain, culture thereof and enteric health improving functional food composition comprising the same |
KR20210089354A (en) * | 2020-01-08 | 2021-07-16 | 전남대학교산학협력단 | Method for preparing oligosaccharide and method for preparing composition for improving bowel health comprising the same |
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