KR20230001021A - A process for the preparation of cinnamon punch containing lactobacillus and the cinnamon punch containing lactobacillus prepared therefrom - Google Patents
A process for the preparation of cinnamon punch containing lactobacillus and the cinnamon punch containing lactobacillus prepared therefrom Download PDFInfo
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- KR20230001021A KR20230001021A KR1020210082291A KR20210082291A KR20230001021A KR 20230001021 A KR20230001021 A KR 20230001021A KR 1020210082291 A KR1020210082291 A KR 1020210082291A KR 20210082291 A KR20210082291 A KR 20210082291A KR 20230001021 A KR20230001021 A KR 20230001021A
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- Prior art keywords
- lactic acid
- acid bacteria
- lactobacillus
- extract
- cinnamon
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Abstract
Description
본 발명은 유산균 수정과의 제조방법 및 이에 따라 제조된 유산균 수정과에 관한 것으로, 보다 구체적으로는 유산균 대사산물을 수정과 제조시 첨가함으로써 소화기능을 향상시키고 피로회복에 도움을 주며, 항비만 및 항염증 효능뿐만 아니라 장기기능 강화효과를 가짐으로써 다이어트 및 건강음료로 애용될 수 있는 유산균 수정과의 제조방법 및 이에 따라 제조된 유산균 수정과에 관한 것이다.The present invention relates to a manufacturing method of fertilized lactobacillus and the fertilized lactobacillus produced thereby, and more specifically, by adding lactobacillus metabolites during fertilization and manufacturing, it improves digestive function and helps to recover from fatigue, anti-obesity and It relates to a method for producing lactic acid bacteria fertilized fruit that can be used as a diet and health drink by having an anti-inflammatory effect as well as a long-term function enhancing effect and a lactic acid bacteria fertilized fruit produced thereby.
유산균(lactic acid bacteria)은 사람이나 동물의 장(腸)과 발효식품 등에서도 쉽게 발견되는 안전한 미생물로(Orrhge, K. et al., 2000, Bifidobacteria and lactobacilli in human health, Drugs Exptl. Clin. Res., 26, 95-111) 장내 상피세포에 부착하여 기생하게 되어 장내 균총의 성질을 개선시켜 장내 균총의 안정화, 유해세균의 정착 억제에 따른 부패산물 생성 감소 및 질병 예방, 면역 활성화 작용, 항암작용, 콜레스테롤 저하 등 인체에 많은 도움을 주는 것으로 알려져 있다. 유산균이 여러 부패성 미생물 및 병원성 미생물에 대하여 생육억제 작용을 갖는 것은 몇 가지 대사적인 특성 때문인데 유산균은 대사산물로서 항균활성 인자인 organic acid, hydrogen peroxide, reuterin, diacetyl, acetaldehyde, bacteriocin 등을 생산하기 때문이다(Fuller, K., 1989, Probiotics in man and animals, J. Appl. Bacteriol., 66, 365-378). 그러나 종래의 유산균 제품들은 대부분 우유를 주재료로 하여 발효시킨 유제품이며 우유의 유당(Lactose)을 잘 소화하지 못하는 우리나라 사람들에게는 적합한 식품이라고 볼 수 없다. 또한, 유산균은 종속영양미생물로서 기본적으로 당을 포함한 각종 아미노산, 비타민 등 여러 가지 영양성분을 필요로 하기 때문에 식물체만을 재료로 하여 유산균 발효액을 만들면 영양성분의 부족으로 요구르트처럼 다량의 유산균을 함유한 유산균 발효액을 만들기가 어렵고 제품의 균일화가 힘들다.Lactic acid bacteria are safe microorganisms that are easily found in human or animal intestines and fermented foods (Orrhge, K. et al., 2000, Bifidobacteria and lactobacilli in human health, Drugs Exptl. Clin. Res ., 26, 95-111) Adheres to epithelial cells in the intestine and becomes parasitic, improving the properties of the intestinal flora, stabilizing the intestinal flora, reducing the production of decay products by inhibiting the settlement of harmful bacteria, preventing diseases, activating immunity, and acting as an anticancer It is known to help the human body, such as lowering cholesterol. Lactic acid bacteria have a growth inhibitory effect on various putrefactive microorganisms and pathogenic microorganisms because of several metabolic characteristics. Lactic acid bacteria produce antibacterial active factors such as organic acid, hydrogen peroxide, reuterin, diacetyl, acetaldehyde, and bacteriocin as metabolites. (Fuller, K., 1989, Probiotics in man and animals, J. Appl. Bacteriol., 66, 365-378). However, most of the conventional lactic acid bacteria products are fermented milk products using milk as a main ingredient, and cannot be considered suitable foods for Koreans who do not digest lactose in milk well. In addition, lactic acid bacteria are heterotrophic microorganisms that basically require various nutrients such as sugars, various amino acids, and vitamins, so if you make a lactic acid bacteria fermentation broth using only plants as ingredients, lactic acid bacteria containing a large amount of lactic acid bacteria like yogurt due to lack of nutrients It is difficult to make fermented liquid and difficult to homogenize the product.
한편, 한국 전통 음청류의 한 종류인 수정과는 계피와 생강, 후추, 설탕,넣고 끓인 물에 곶감과 잣을 첨가하고, 여기에 설탕 또는 꿀을 넣어 차갑게 해서 마치는 기호성 음료이다. 수정과 재료로 사용되는 계피, 생강, 후추는 방향, 건위 혈액순환 촉진, 식욕증진, 소화촉진 등의 효과가 있다고 알려져 있다. 계피(Cinnamomum cassia)는 녹나무과에 속하는 계수나무의 껍질로 항균, 항산화, 혈중 지질저하 효과 및 항당뇨 효과가 있음이 보고되고 있다. 계피의 cinnamaldehyde, cinnamic acid의 지질저하 효과는 많은 연구를 통해 보고되어 있으며, 항당뇨 효과로는 인슐린 리셉터의 자동인산화를 촉진시킴으로써 인슐린 민감성을 개선시킨다고 보고되고 있다.On the other hand, Sujeonggwa, a kind of Korean traditional drink, is a tasty beverage made by adding dried persimmons and pine nuts to boiled water with cinnamon, ginger, pepper, sugar, and then adding sugar or honey to cool it. Cinnamon, ginger, and pepper, which are used as crystals and ingredients, are known to have effects such as aroma, promotion of blood circulation in the stomach, appetite, and digestion. Cinnamon (Cinnamomum cassia) is the bark of a cinnamon tree belonging to the Lauraceae family and has been reported to have antibacterial, antioxidant, blood lipid-lowering effects, and antidiabetic effects. The lipid-lowering effects of cinnamaldehyde and cinnamic acid in cinnamon have been reported through many studies, and as an antidiabetic effect, it has been reported that insulin sensitivity is improved by promoting autophosphorylation of insulin receptors.
생강(Zingiber officinale)은 아시아에서 널리 사용되어 온 약용식물로 관절염, 근육통, 후두염, 변비, 구토, 고혈압, 및 감염성 질환 등의 다양한 질병 치유에 사용되어왔다. 생강의 유효성분인 진저롤(gingerol), 진저론(zingerone) 그리고 쇼가올(shogaol)의 항산화 효과는 많은 연구를 통해 잘 알려져 있으며 특히 6-쇼가올(shogaol)의 유리기 소거 효과가 높은 것으로 알려져 있다. 최근에는 항염증, 항궤양 및 항암, 항당뇨, 간독성 보호효과 및 콜레스테롤 저하효과 등이 보고되고 있다.Ginger (Zingiber officinale) is a medicinal plant that has been widely used in Asia and has been used to treat various diseases such as arthritis, muscle pain, laryngitis, constipation, vomiting, high blood pressure, and infectious diseases. The antioxidant effects of gingerol, zingerone, and shogaol, which are active ingredients of ginger, are well known through many studies, and in particular, 6-shogaol is known to have a high free radical scavenging effect. there is. Recently, anti-inflammatory, antiulcer and anticancer, antidiabetic, hepatotoxicity protective effects and cholesterol lowering effects have been reported.
한편, 현대인의 비만은 만성질환을 유발하는 가장 중요한 원인으로 알려져 있으며, 비만의 원인 중 하나로 당 함량이 높은 음료의 섭취가 지적되고 있어 최근 세계 음료시장은 단맛은 느껴지나 저칼로리인 음료 그리고 카페인이 함유되어 있지 않은 기능성 음료 형태로 시장이 확대되고 있다. 전통 수정과 제조법에 의하면 수정과의 단맛을 내는 급원으로 꿀 또는 설탕을 사용하고 있어 수정과 섭취 시 상당량의 칼로리가 얻어지고 이는 콜라나 사이다 등의 청량음료 섭취로부터 얻어지는 칼로리와 유사한 것으로 나타났다(비락 수정과: 44 Kcal/100 mL, 코카콜라 44 Kcal/100mL). 따라서 무가당 음료, 저칼로리 음료에 대한 요구가 커지고 있는 실정에 맞춘 전통 음청류의 개발이 요구되고 있다. On the other hand, obesity in modern people is known as the most important cause of chronic diseases, and consumption of beverages with high sugar content is pointed out as one of the causes of obesity. The market is expanding in the form of functional beverages that are not yet available. According to the traditional Sujeonggwa manufacturing method, honey or sugar is used as a source of sweetness for Sujeonggwa, so a significant amount of calories are obtained when ingesting Sujeonggwa, which is similar to calories obtained from consumption of soft drinks such as cola or cider (Virak Sujeong And: 44 Kcal/100 mL, Coca-Cola 44 Kcal/100 mL). Therefore, there is a demand for the development of traditional beverages tailored to the increasing demand for sugar-free and low-calorie beverages.
이와 관련하여, 국제등록특허 제10-1576398호에는 탄산미가 부가된 수정과가, 단맛 소재의 저칼로리 기능성 수정과는 10-2016-8762호, 소재의 다양화를 추구하기 위해 감귤 또는 파프리카의 유효성분을 함유시킨 수정과 음료는 10-2013-49681호에 각각 공지되어 있다. 소재와 관련된 선행기술로는 산사를 이용한 것, 고로쇠 수용액을 이용한 것, 야자수액이 첨가된 것이 각각 10-1537465호, 10-1260472호, 10-1775783호로 공지되어 있다.In this regard, International Patent Registration No. 10-1576398 has Sujeonggwa with added carbonic acid taste, and Low-calorie functional Sujeonggwa of sweet material No. 10-2016-8762, and active ingredients of citrus or paprika to pursue diversification of materials Crystals and beverages containing are known from 10-2013-49681, respectively. As the prior art related to the material, those using hawthorn sand, using an aqueous solution of gorosoe, and adding palm tree liquid are known as Nos. 10-1537465, 10-1260472, and 10-1775783, respectively.
그러나 아직까지 유산균 대사산물을 함유하는 수정과 및 그 제조방법에 관하여는 개시된 바가 없다.However, no disclosure has yet been made regarding fertilization containing lactic acid bacteria metabolites and a method for producing the same.
이에 본 발명자들은 이러한 다양한 효능을 가지는 유산균 대사산물을 수정과에 첨가함으로써 피로회복, 소화기능 개선, 항비만 및 항염증 효과뿐만 아니라 장기능을 개선하고 맛과 향을 향상시킬 수 있다는 사실을 발견하여 본 발명을 완성하였다. Accordingly, the inventors of the present invention found that by adding lactic acid bacteria metabolites having various effects to fertilized fruit, fatigue recovery, digestive function improvement, anti-obesity and anti-inflammatory effects, as well as intestinal function and taste and aroma can be improved. completed the present invention.
따라서, 본 발명에서 해결하고자 하는 기술적 과제는 피로회복, 소화기능 개선, 항비만 및 항염증 효과뿐만 아니라 장기능을 개선하고 맛과 향을 향상시킬 수 있는 유산균 수정과의 제조방법을 제공하기 위한 것이다.Therefore, the technical problem to be solved in the present invention is to provide a method for producing modified lactic acid bacteria, which can improve intestinal function and improve taste and aroma, as well as fatigue recovery, digestive function improvement, anti-obesity and anti-inflammatory effects. .
또한, 본 발명에서 해결하고자 하는 다른 기술적 과제는 상기 제조방법에 따라 제조된 유산균 수정과를 제공하기 위한 것이다. In addition, another technical problem to be solved in the present invention is to provide a fertilized fruit of lactic acid bacteria prepared according to the above manufacturing method.
상기한 기술적 과제를 해결하기 위하여, 본 발명에서는 하기 단계를 포함하는 것을 특징으로 하는 유산균 수정과의 제조방법을 제공한다:In order to solve the above technical problem, the present invention provides a method for producing lactic acid bacteria fertilized fruit, characterized in that it comprises the following steps:
(S1) 계피, 생강 및 곶감에 각각 정제수를 혼합하여 80 내지 100℃에서 110 내지 130분 동안 가열하여 계피 추출물, 생강 추출물 및 곶감 추출물을 각각 수득하는 단계;(S1) mixing cinnamon, ginger, and dried persimmon with purified water and heating at 80 to 100° C. for 110 to 130 minutes to obtain cinnamon extract, ginger extract, and dried persimmon extract, respectively;
(S2) 상기 단계 (S1)에서 수득된 계피 추출물, 생강 추출물 및 곶감 추출물을 혼합한 다음 유산균 대사산물, 프락토올리고당과 설탕 혼합물 및 곶감을 첨가하는 단계;(S2) mixing the cinnamon extract, ginger extract, and dried persimmon extract obtained in step (S1) and then adding lactic acid bacteria metabolites, fructooligosaccharide and sugar mixture, and dried persimmon;
(S3) 상기 단계 (S2)에서 수득된 혼합물을 95 내지 100℃에서 50 내지 70분 동안 가열하여 농축 및 살균처리 하는 단계;(S3) concentrating and sterilizing the mixture obtained in step (S2) by heating at 95 to 100° C. for 50 to 70 minutes;
(S4) 상기 단계 (S3)에서 수득된 유산균 수정과를 살균 및 여과하는 단계; 및(S4) sterilizing and filtering the fertilized fruit of lactic acid bacteria obtained in step (S3); and
(S5) 상기 단계 (S4)에서 수득된 유산균 수정과를 60 내지 80℃에서 식힌 후 충진기로 이송하여 충진하여 포장하는 단계.(S5) Cooling the fertilized fruit of lactic acid bacteria obtained in step (S4) at 60 to 80 ° C, transferring it to a filling machine, filling and packaging.
바람직하게, 계피 추출물, 생강 추출물 및 곶감 추출물을 1.6:1.5:1.2의 중량비로 혼합하는 것을 특징으로 한다.Preferably, cinnamon extract, ginger extract and dried persimmon extract are mixed in a weight ratio of 1.6: 1.5: 1.2.
바람직하게, 상기 유산균 대사물질은 락토바실러스 플란타룸(L.plantarum), 락토바실러스 카제이(L.casei) 및 비피도박테리움 인펀티스(B. infantis)로 이루어진 군에서 선택된 하나 이상의 유산균을 액체 배지에 접종하고 20 내지 40℃에서 12 내지 48시간 배양하여 수득된 발효액을 동결건조하여 제조될 수 있다.Preferably, the lactic acid bacteria metabolite is Lactobacillus plantarum ( L.plantarum ), Lactobacillus casei ( L.casei ) and Bifidobacterium infantis ( B. infantis ) One or more lactic acid bacteria selected from the group consisting of a liquid It may be prepared by freeze-drying a fermentation broth obtained by inoculating a medium and incubating at 20 to 40° C. for 12 to 48 hours.
바람직하게, 상기 단계 (S2)에서 프락토올리고당과 설탕 혼합물은 프락토올리고당 및 설탕이 2:8의 중량비로 혼합된 것을 특징으로 한다Preferably, the fructooligosaccharide and sugar mixture in step (S2) is characterized in that fructooligosaccharide and sugar are mixed in a weight ratio of 2:8
바람직하게, 상기 단계 (S4)에서 살균은 95 내지 100 ℃에서 20 내지 30분 동안 수행하는 것을 특징으로 한다.Preferably, the sterilization in step (S4) is performed at 95 to 100 ° C. for 20 to 30 minutes.
상기한 다른 기술적 과제를 해결하기 위하여, 본 발명에서는 상기 제조방법에 따라 제조된 유산균 수정과를 제공한다.In order to solve the above other technical problems, the present invention provides a lactic acid bacteria fertilized fruit prepared according to the above manufacturing method.
이와 같이, 본 발명에 따라 제조된 유산균 대사산물을 포함하는 유산균 수정과는 유산균의 다양한 효능이 부가되어 피로회복, 소화기능 개선효과가 있을 뿐만 아니라 장기강화, 항비만 및 항염증 효능을 가진다. 또한, 유산균과 함께 사용함으로써 기호도를 보다 상승시킬 수 있다. 따라서, 본 발명의 유산균 수정과는 피로회복, 항염 및 다이어트 음료로도 사용될 수 있는 바, 공부하는 학생 및 직장인에게 비만의 걱정없는 웰빙 건강음료로 매우 유용하게 이용될 수 있을 것으로 기대된다.As such, fertilized lactic acid bacteria containing the metabolites of lactic acid bacteria prepared according to the present invention have various effects of lactic acid bacteria added to have fatigue recovery and digestive function improvement effects as well as long-term strengthening, anti-obesity and anti-inflammatory effects. In addition, preference can be further increased by using it together with lactic acid bacteria. Therefore, the fertilized fruit of lactic acid bacteria of the present invention can be used as a fatigue recovery, anti-inflammatory and diet drink, and is expected to be very useful as a well-being health drink without worrying about obesity for studying students and office workers.
이하 본 발명을 보다 구체적으로 설명한다.Hereinafter, the present invention will be described in more detail.
본 발명에서는 하기 단계를 포함하는 것을 특징으로 하는 유산균 수정과의 제조방법을 제공한다:The present invention provides a method for producing fertilized lactic acid bacteria, characterized in that it comprises the following steps:
(S1) 계피, 생강 및 곶감에 각각 정제수를 혼합하여 80 내지 100℃에서 110 내지 130분 동안 가열하여 계피 추출물, 생강 추출물 및 곶감 추출물을 각각 수득하는 단계;(S1) mixing cinnamon, ginger, and dried persimmon with purified water and heating at 80 to 100° C. for 110 to 130 minutes to obtain cinnamon extract, ginger extract, and dried persimmon extract, respectively;
(S2) 상기 단계 (S1)에서 수득된 계피 추출물, 생강 추출물 및 곶감 추출물을 혼합한 다음 유산균 대사산물, 프락토올리고당과 설탕 혼합물 및 곶감을 첨가하는 단계;(S2) mixing the cinnamon extract, ginger extract, and dried persimmon extract obtained in step (S1) and then adding lactic acid bacteria metabolites, fructooligosaccharide and sugar mixture, and dried persimmon;
(S3) 상기 단계 (S2)에서 수득된 혼합물을 95 내지 100℃에서 50 내지 70분 동안 가열여 농축 및 살균처리 하는 단계;(S3) concentrating and sterilizing the mixture obtained in step (S2) by heating at 95 to 100° C. for 50 to 70 minutes;
(S4) 상기 단계 (S3)에서 수득된 유산균 수정과를 살균 및 여과하는 단계; 및(S4) sterilizing and filtering the fertilized fruit of lactic acid bacteria obtained in step (S3); and
(S5) 상기 단계 (S4)에서 수득된 유산균 수정과를 60 내지 80℃에서 식힌 후 충진기로 이송하여 충진하여 포장하는 단계.(S5) Cooling the fertilized fruit of lactic acid bacteria obtained in step (S4) at 60 to 80 ° C, transferring it to a filling machine, filling and packaging.
본 발명의 하나의 구현예에 따르면, 곶감, 계피 및 생강의 통상의 수정과의 제조시 첨가되는 양으로 사용되며, 예를 들어 곶감, 계피 및 생강은 1.6:1.5:1.2의 중량비로 사용된다.According to one embodiment of the present invention, dried persimmon, cinnamon, and ginger are used in an amount added during the manufacture of a conventional fertilized fruit, for example, dried persimmon, cinnamon, and ginger are used in a weight ratio of 1.6: 1.5: 1.2.
본 발명의 유산균 수정과는 기존의 완성된 수정과에 기호에 따라 잘게 썬 곶감을 띄워서 식용한 것과는 달리, 수정과의 원물로 곶감 추출액을 사용함으로써 유산균의 당도를 조절하고 곶감 고유의 단맛을 구현할 수 있는 이점이 있다. Unlike the existing finished Sujeonggwa of the present invention, dried persimmons cut according to taste are floated and eaten. By using the dried persimmon extract as the raw material of Sujeonggwa, the sugar content of lactic acid bacteria can be adjusted and the unique sweetness of dried persimmons can be realized. There is an advantage to being
본 발명의 하나의 구현예에 따르면, 본 발명의 유산균 수정과의 제조방법에서 단계 (S2)는 유산균 대사산물 및 프락토올리고당과 설탕의 혼합물과 곶감을 첨가하는 것을 특징으로 한다. 이 때 유산균 대사산물은 유산균 수정과 100 중량부에 대하여 0.01 내지 0.03 중량부의 양으로 첨가하고, 상기 프락토올리고당과 설탕의 혼합물은 유산균 수정과 100 중량부에 대하여 5 내지 7 중량부의 양으로 첨가하고,곶감은 1 내지 3 중량부의 양으로 첨가하는 것을 특징으로 한다. According to one embodiment of the present invention, step (S2) in the manufacturing method of lactobacillus sujeonggwa of the present invention is characterized in that a mixture of lactic acid bacteria metabolites, fructooligosaccharide and sugar and dried persimmon are added. At this time, the metabolite of lactic acid bacteria is added in an amount of 0.01 to 0.03 parts by weight based on 100 parts by weight of modified lactic acid bacteria, and the mixture of fructooligosaccharide and sugar is added in an amount of 5 to 7 parts by weight based on 100 parts by weight of modified lactic acid bacteria. , Dried persimmon is characterized in that it is added in an amount of 1 to 3 parts by weight.
상기 프락토올리고당과 설탕은 2:8의 중량비로 혼합하는 것이 바람직하며, 상기 프락토올리고당은 유산균의 먹이로 첨가된다.It is preferable to mix the fructooligosaccharide and sugar in a weight ratio of 2:8, and the fructooligosaccharide is added as food for lactic acid bacteria.
본 발명에서 유산균 대사산물이란 유산균이 먹이를 먹고 만들어낸 물질로서, 대표적으로 사균체, 박테리오신 등이 있다. 이러한 유산균 대사산물은 유산균과 달리 살아있는 형태가 아니기 때문에 위산과 담즙산에 영향을 받지 않아 장까지 안정적으로 도달하고 유해균을 직접 사멸하는 것이 가능해 장내 빠른 안정화를 돕는다.In the present invention, the metabolites of lactic acid bacteria are substances produced by eating lactic acid bacteria, and representatively include dead cells, bacteriocins, and the like. Since these metabolites of lactic acid bacteria are not in a living form unlike lactic acid bacteria, they are not affected by stomach acid and bile acid, so they stably reach the intestines and can directly kill harmful bacteria, helping to stabilize the intestine quickly.
유산균 대사산물은 그 자체로도 효능을 가지고 있는데, 사균체의 경우 아토피 피부염을 완화하는 데 도움이 된다. 2008년 생물학제약회보(Biological and Pharmaceutical Bulletin)에 실린 연구에 따르면, 아토피를 유발한 실험쥐에 사균체를 투입한 결과 아토피 피부염 등 알레르기를 유발하는 IgE(이뮤노글로불린E)의 생성이 억제된 것으로 나타났다.The metabolite of lactic acid bacteria has an effect by itself, and in the case of dead cells, it helps to alleviate atopic dermatitis. According to a study published in the Biological and Pharmaceutical Bulletin in 2008, the production of IgE (immunoglobulin E), which causes allergies such as atopic dermatitis, was suppressed as a result of injecting dead cells into atopic mice. appear.
본 발명에서 사용하는 유산균은 락토바실러스 플란타룸(L.plantarum), 락토바실러스 카제이(L.casei) 및 비피도박테리움 인펀티스(B. infantis)로 이루어진 군에서 선택된 하나 이상의 유산균, 바람직하게는 락토바실러스 플란타룸(L.plantarum), 락토바실러스 카제이(L.casei) 및 비피도박테리움 인펀티스(B. infantis)를 1:1:1의 비율로 혼합한 혼합 유산균을 접종하는 것을 특징으로 한다.The lactic acid bacteria used in the present invention is one or more lactic acid bacteria selected from the group consisting of Lactobacillus plantarum ( L.plantarum ), Lactobacillus casei ( L.casei ) and Bifidobacterium infantis ( B. infantis ), preferably The Lactobacillus plantarum ( L.plantarum ), Lactobacillus casei ( L.casei ) and Bifidobacterium infantis ( B. infantis ) are inoculated with mixed lactic acid bacteria mixed at a ratio of 1: 1: 1 to be characterized
상기 유산균 중에 하나인 락토바실러스 플란타룸(L.plantarum)은 락토바실러스(Lactobacillus)속 젖산균으로 막대기 모양의 간균이며 그람양성이다. 크기는 0.7 내지 1.0 x 3.0 내지 8.0 μ으로 생육 적정 온도는 29 내지 33 ℃ 이며 약간의 호기성을 띤다. 락토바실러스 플란타룸은 아라비노오스(arabinose), 글루코오스(glucose), 과당(fructose), 갈락토오스(galactose), 엿당(maltose), 수크로오스(sucrose), 덱스트란(dextran), 라피노오스(raffinose)와 트레할로스(trehalose) 등을 발효하여 젖산을 생성한다. 락토바실러스 플란타룸은 주로 우유, 치즈, 버터, 케피어(kefir), 발효된 소시지, 발효된 감자, 곡물, 빵의 산성 반죽(dough), 피클이나 김치와 같은 침채류, 토마토 등에서 분리되며, 자연계에서 가장 분포가 넓은 젖산균 중의 하나이다. 특히 우리나라 김치의 발효에 중요한 역할을 하는 젖산균으로 식염 내성은 약 5.5%이다. One of the lactic acid bacteria, Lactobacillus plantarum ( L. plantarum ) is a lactic acid bacterium of the genus Lactobacillus, and is a rod-shaped bacillus and is Gram-positive. The size is 0.7 to 1.0 x 3.0 to 8.0 μ, the optimum temperature for growth is 29 to 33 ° C, and it is slightly aerobic. Lactobacillus plantarum contains arabinose, glucose, fructose, galactose, maltose, sucrose, dextran, and raffinose and trehalose are fermented to produce lactic acid. Lactobacillus plantarum is mainly isolated from milk, cheese, butter, kefir, fermented sausages, fermented potatoes, grains, acidic bread dough, pickles and kimchi, and tomatoes. It is one of the most widely distributed lactic acid bacteria in nature. In particular, it is a lactic acid bacterium that plays an important role in the fermentation of Korean kimchi, and its salt tolerance is about 5.5%.
또한, 락토바실러스 카제이(Lactobacillus casei)는 인간의 장이나 입에서 발견되는 락토바실러스(Lactobacillus)속의 혐기성 미생물로서 우유와 치즈에서 분리되었다. 프로바이오틱스(probiotics)인 락토바실러스 카제이는 막대기 모양의 간균으로 그람 양성이고, 운동성이 없으며, 산이나 열에 강하다. 또한 락토바실리러스 아시도필루스(Lactobacillus acidophilus)의 성장에 도움을 주며, 탄수화물을 분해시키는 효소인 아밀라아제(amylase)를 생산한다. In addition, Lactobacillus casei is an anaerobic microorganism of the genus Lactobacillus found in the human intestine or mouth and has been isolated from milk and cheese. Lactobacillus casei, a probiotic, is a rod-shaped bacillus that is Gram-positive, non-motile, and resistant to acid or heat. It also helps the growth of Lactobacillus acidophilus and produces amylase, an enzyme that breaks down carbohydrates.
이 균은 인체 내 소화액에 의해 사멸되지 않고 소장까지 가서 소장 내 균총을 정상화시키고, 유당불내증(milk intolerance)을 줄여주며, 정장 작용 및 소화작용을 돕는 인체에 매우 유익한 균이다. 우유는 락토바실러스 카제이가 생산하는 젖산에 의해 3-5일 만에 응고되어 점성을 가지는데, 이때 카제인(casein)을 이용하므로 치즈 숙성에서는 매우 중요한 균이다. 최근에는 락토바실러스 카제이가 위궤양이나 위염을 일으키는 헬리코박터 파일로리(Helicobacter pylori)의 성장을 저해하며, 대장 내의 미생물 균총의 균형에 도움을 주는 것으로 보고되고 있다. This bacterium is not killed by the digestive juices in the human body, but goes to the small intestine, normalizes the flora in the small intestine, reduces milk intolerance, and is a very beneficial bacterium to the human body that helps the intestinal function and digestion. Milk is coagulated in 3-5 days by lactic acid produced by Lactobacillus casei and becomes viscous. At this time, casein is used, which is a very important bacterium in cheese ripening. Recently, it has been reported that Lactobacillus casei inhibits the growth of Helicobacter pylori, which causes gastric ulcer or gastritis, and helps balance the microbial flora in the large intestine.
비피도박테리움 인펀티스(B. infantis)은 음식물 소화를 돕고 과민성 대장증후군을 개선시킨다. 면역을 강화하며 장 내부 유해균을 억제하는 효과가 높은 것으로 알려져 있다. Bifidobacterium infantis ( B. infantis ) helps digestion of food and improves irritable bowel syndrome. It is known to strengthen immunity and suppress harmful bacteria in the intestines.
상기 유산균을 배양하기 위한 식품용 액체배지(포도당 2.0 ~ 4.0 중량%, 펩톤 0.5 ~ 2.0 중량%, 효모추출물 0.2 ~ 1.0 중량%, 무기염류 2.0 ~ 4.0 중량% 및 정제수 89.0 ~ 95.3 중량%로 이루어진 액체배지)에 1x108 cfu/mL 이상의 유산균을 접종하고 20 내지 40℃에서 12 내지 48시간 배양하여 수득된 발효액을 동결건조하여 유산균 대사산물을 제조할 수 있다. Food liquid medium for culturing the lactic acid bacteria (2.0 to 4.0% by weight of glucose, 0.5 to 2.0% by weight of peptone, 0.2 to 1.0% by weight of yeast extract, 2.0 to 4.0% by weight of inorganic salts, and 89.0 to 95.3% by weight of purified water) Medium) is inoculated with 1x10 8 cfu/mL or more of lactic acid bacteria and incubated at 20 to 40° C. for 12 to 48 hours, and freeze-drying the obtained fermentation broth to produce metabolites of lactic acid bacteria.
본 발명의 하나의 구현예에 따르면, 상기 유산균 대사물질은 락토바실러스 플란타룸(L.plantarum), 락토바실러스 카제이(L.casei) 및 비피도박테리움 인펀티스(B. infantis)로 이루어진 군에서 선택된 하나 이상의 유산균을 액체 배지에 접종하고 20 내지 40℃에서 12 내지 48시간 배양하여 수득된 발효액을 동결건조하여 제조될 수 있다.According to one embodiment of the present invention, the lactic acid bacteria metabolite is a group consisting of Lactobacillus plantarum ( L. plantarum ), Lactobacillus casei ( L. casei ) and Bifidobacterium infantis ( B. infantis ). It can be prepared by inoculating one or more lactic acid bacteria selected from in a liquid medium and culturing at 20 to 40 ° C. for 12 to 48 hours and freeze-drying the obtained fermentation broth.
본 발명의 하나의 구현예에 따르면, 본 발명의 유산균 수정과 제조방법에서 단계 (S4)에서 살균은 95 내지 100 ℃에서 20 내지 30분 동안 수행하는 것을 특징으로 한다.According to one embodiment of the present invention, sterilization in step (S4) in the lactic acid bacteria fertilization and manufacturing method of the present invention is characterized in that it is performed at 95 to 100 ° C. for 20 to 30 minutes.
본 발명의 하나의 구현예에 따르면, 본 발명의 하나의 구현예에 따르면, 본 발명의 유산균 수정과의 제조방법에서 단계 (S5)는 유산균 수정과를 60 내지 80℃에서 식힌 후 충진기로 이송하여 충진하여 포장하는 것을 특징으로 한다.According to one embodiment of the present invention, in the manufacturing method of the lactic acid bacteria fertilized fruit of the present invention, step (S5) is to cool the lactic acid bacteria fertilized fruit at 60 to 80 ° C and transfer it to a filling machine. It is characterized by filling and packaging.
한편, 본 발명에서는 상기 제조방법에 따라 제조된 유산균 수정과를 제공한다.On the other hand, the present invention provides a fertilized fruit of lactic acid bacteria prepared according to the above manufacturing method.
이와 같이, 본 발명에 따라 제조된 유산균 대사산물을 포함하는 유산균 수정과는 유산균의 다양한 효능이 부가되어 피로회복, 소화기능 개선효과가 있을 뿐만 아니라 장기강화, 항비만 및 항염증 효능을 가진다. 또한, 유산균과 함께 사용함으로써 기호도를 보다 상승시킬 수 있다. 따라서, 본 발명의 유산균 수정과는 피로회복, 항염 및 다이어트 음료로도 사용될 수 있는 바, 공부하는 학생 및 직장인에게 비만의 걱정없는 웰빙 건강음료로 매우 유용하게 이용될 수 있을 것으로 기대된다As such, fertilized lactic acid bacteria containing the metabolites of lactic acid bacteria prepared according to the present invention have various effects of lactic acid bacteria added to have fatigue recovery and digestive function improvement effects as well as long-term strengthening, anti-obesity and anti-inflammatory effects. In addition, preference can be further increased by using it together with lactic acid bacteria. Therefore, the fermented fruit of lactic acid bacteria of the present invention can be used as a fatigue recovery, anti-inflammatory and diet drink, and it is expected that it can be used very usefully as a well-being health drink without worrying about obesity for studying students and office workers.
이하, 본 발명의 이해를 돕기 위하여 도면과 실시예 등을 들어 상세하게 설명하기로 한다. 그러나, 본 발명에 따른 실시예들은 여러 가지 다른 형태로 변형될 수 있으며, 본 발명의 범위가 하기 실시예들에 한정되는 것으로 해석되어서는 안 된다. 본 발명의 실시예들은 당업계에서 평균적인 지식을 가진 자에게 본 발명을 보다 완전하게 설명하기 위해 제공되는 것이다. Hereinafter, in order to aid understanding of the present invention, it will be described in detail with reference to drawings and embodiments. However, the embodiments according to the present invention can be modified in many different forms, and the scope of the present invention should not be construed as being limited to the following examples. Embodiments of the present invention are provided to more completely explain the present invention to those skilled in the art.
<제조예 1> 유산균 대사물질 제조<Preparation Example 1> Preparation of lactic acid bacteria metabolites
유산균의 배양조건은 배양 온도 20 내지 40℃에서 12 내지 48시간 동안 유산균 수가 1x108 cfu/mL 이상, pH 4.8 이하가 될 때까지 액체배지에서 각각 배양하여 유산균 배양액을 제조한 다음 동결건조하여 유산균 대사물질을 수득하였다.The culture conditions of lactic acid bacteria are cultured in a liquid medium for 12 to 48 hours at a culture temperature of 20 to 40 ° C until the number of lactic acid bacteria reaches 1x10 8 cfu / mL or more and pH 4.8 or less to prepare a culture medium of lactic acid bacteria, and then lyophilize to metabolize lactic acid bacteria. material was obtained.
구체적으로, 상기 유산균을 배양하기 위한 식품용 액체배지로 포도당 2.0 ~ 4.0 중량%, 펩톤 0.5 ~ 2.0 중량%, 효모추출물 0.2 ~ 1.0 중량%, 무기염류 2.0 ~ 4.0 중량% 및 정제수 89.0 ~ 95.3 중량%로 이루어진 액체배지를 121℃에서 30분간 멸균하여 제조하였다.Specifically, as a food liquid medium for culturing the lactic acid bacteria, 2.0 to 4.0% by weight of glucose, 0.5 to 2.0% by weight of peptone, 0.2 to 1.0% by weight of yeast extract, 2.0 to 4.0% by weight of inorganic salts and 89.0 to 95.3% by weight of purified water It was prepared by sterilizing a liquid medium consisting of at 121 ° C. for 30 minutes.
본 발명에 따른 액체배지를 구성하는 성분 조성에서 무기염류는 배지조성에서 통상적으로 사용하는 Mg, Mn, Co 성분의 중량부 합계이다.In the component composition constituting the liquid medium according to the present invention, inorganic salts are the sum of parts by weight of Mg, Mn, and Co components commonly used in the medium composition.
상기 배양한 유산균 배양액을 7,000 ∼ 10,000 rpm으로 원심분리하여 유산균체와 배양상등액을 분리하였다. 상기에서 분리된 배양여액은 다시 살균(100℃, 30분)한 후 90℃까지 냉각하여 유산균 배양여액을 제조한 다음 통상의 방법에 따라 감압 농축하여 2배 이상의 농축된 배양농축액을 제조하였다.The cultured lactic acid bacteria culture medium was centrifuged at 7,000 to 10,000 rpm to separate lactic acid bacteria and culture supernatant. The culture filtrate separated above was sterilized again (100 ° C, 30 minutes), cooled to 90 ° C to prepare a lactic acid bacteria culture filtrate, and then concentrated under reduced pressure according to a conventional method to prepare a culture concentrate twice or more concentrated.
상기 방법으로 제조된 본 발명 유산균 배양농축액에는 유산균의 발효과정 중 생성된 대사산물로서 주로 감마 아미노뷰티르산(GABA), 아미노산, 펩타이드, 비타민, 유기산 및 박테리오신 등이 포함된다.The lactic acid bacteria culture concentrate of the present invention prepared by the above method includes mainly gamma aminobutyric acid (GABA), amino acids, peptides, vitamins, organic acids and bacteriocin as metabolites produced during the fermentation of lactic acid bacteria.
상기 수득된 유산균 배양농축액을 동결건조하여 유산균 대사산물을 제조하였다.Lactic acid bacteria metabolites were prepared by freeze-drying the obtained lactic acid bacteria culture concentrate.
<실시예 1> 유산균 수정과 제조<Example 1> Fertilization and preparation of lactic acid bacteria
곶감 1.6 중량%, 계피 1.5 중량%, 생강 1.2중량% 및 정제수 89.79 중량%를 혼합하여 80 내지 100℃에서 110 내지 130분 동안 가열하여 재료의 원물을 추출한 다음 상기 제조예 1에서 수득된 유산균 대사물질을 건져낸 다음 유산균 대사산물을 유산균 수정과 100 중량부에 대하여 0.01 내지 0.03 중량부의 양으로 첨가하고, 프락토올리고당과 설탕의 혼합물은 유산균 수정과 100 중량부에 대하여 5 내지 7 중량부의 양으로 첨가하고 곶감은 유산균 수정과 100 중량부에 대하여 1 내지 3 중량부의 양으로 첨가하여 95 내지 100℃에서 50 내지 70분 동안 가열하여 농축하였다.Dried persimmon 1.6% by weight, cinnamon 1.5% by weight, ginger 1.2% by weight and purified water 89.79% by weight were mixed and heated at 80 to 100 ° C. for 110 to 130 minutes to extract the raw material of the material, and then Lactic acid bacteria metabolites obtained in Preparation Example 1 Then, lactic acid bacteria metabolites are added in an amount of 0.01 to 0.03 parts by weight based on 100 parts by weight of modified lactic acid bacteria, and a mixture of fructooligosaccharide and sugar is added in an amount of 5 to 7 parts by weight based on 100 parts by weight of modified lactic acid bacteria. Dried persimmon was added in an amount of 1 to 3 parts by weight based on 100 parts by weight of modified lactic acid bacteria, and concentrated by heating at 95 to 100 ° C. for 50 to 70 minutes.
이 후 수득된 유산균 수정과의 살균은 95 내지 100 ℃에서 20 내지 30분 동안 수행하고, 60 내지 80℃에서 식힌 후 충진기로 이송하여 충진하여 유산균 수정과를 포장하였다.Sterilization of the obtained Lactobacillus modified fruit was performed at 95 to 100 ° C. for 20 to 30 minutes, cooled at 60 to 80 ° C., and then transferred to a filling machine to fill the lactic acid bacteria modified fruit.
<비교예 1> <Comparative Example 1>
유산균 대사물질을 첨가하지 않은 일반 수정과를 제조하였다.General fertilized fruit without the addition of lactic acid bacteria metabolites was prepared.
<시험예 1> 당도 측정<Test Example 1> Sugar content measurement
상기 실시예 1에서 제조된 유산균 수정과 당도는 당도계(digital refractometer, pocket PAL-1, Atago co., Japan)로 측정한 뒤 ˚Brix로 표시하였다. 모든 시료는 1g씩 채취하여 3회 반복한 평균값으로 하였다. 비교를 위해 유산균 대사물질을 넣지 않은 비교예 1의 일반 수정과를 사용하였다. 그 결과는 하기 표 1에 나타내었다.The lactobacillus crystals and sugar content prepared in Example 1 were measured with a saccharometer (digital refractometer, pocket PAL-1, Atago co., Japan) and expressed as °Brix. All samples were taken 1g each and made into an average value repeated three times. For comparison, the general fertilized fruit of Comparative Example 1, which did not contain lactic acid bacteria metabolites, was used. The results are shown in Table 1 below.
상기 표 1에서 보듯이, 실시예 1에서 제조된 유산균 수정과 및 비교예 1에서 제조된 일반 수정과의 ˚Brix는 12˚Brix로 유산균의 첨가가 유의적으로 당도가 변화하지 않음을 알 수 있다. As shown in Table 1, the ˚Brix of the modified lactobacillus prepared in Example 1 and the general modified fruit prepared in Comparative Example 1 is 12˚Brix, and it can be seen that the addition of lactic acid bacteria does not significantly change the sugar content. .
<시험예 2> pH 측정<Test Example 2> pH measurement
유산균 수정과의 pH는 pH/ISE Meter(pH/Temp Meter pH-200L)로 측정하였다. 모든 시료는 30ml씩 채취하여 3회 반복한 평균값으로 하였다. 그 결과는 하기 표 2에 나타내었다.The pH of the fertilized lactic acid bacteria was measured with a pH/ISE Meter (pH/Temp Meter pH-200L). All samples were collected in 30 ml each, and the average value was repeated three times. The results are shown in Table 2 below.
상기 표 2에서 보듯이, 본 발명의 유산균 수정과의 pH는 5.21 및 5.24으로, 일반 수정과와 비슷한 pH를 가지는 것을 알 수 있었다.As shown in Table 2, the pH of the fertilized lactobacillus of the present invention was 5.21 and 5.24, which was found to have a pH similar to that of the normal fertilized fruit.
<시험예 3> 유산균 수정과의 대식세포주 (RAW264.7)에서 LPS에 의해 유도된 NO 억제 측정<Test Example 3> Measurement of NO inhibition induced by LPS in macrophage cell line (RAW264.7) of fertilized lactic acid bacteria
대식세포에서는 LPS와 같은 외부 자극 등에 의해 염증반응이 일어나면 NO를 분비하고 염증성 사이토카인과 같은 다양한 물질을 생성하고, 염증반응을 조절하는 다양한 병리적인 반응이 일어난다. LPS로 염증반응을 유도한 RAW264.7 세포에서 고들빼기 추출물을 처리하여 항염증 효능을 조사하였다. In macrophages, when an inflammatory response occurs due to an external stimulus such as LPS, NO is secreted, various substances such as inflammatory cytokines are produced, and various pathological responses that control the inflammatory response occur. In RAW264.7 cells induced with LPS, the anti-inflammatory effect was investigated by treating the extract of P. japonica.
안정화된 NO 산화물인 NO2 (Nitrite)는 Griess 반응을 이용하여 측정하였다. 먼저 마우스의 대식세포주(RAW264.7)를 96웰 플레이트에 well당 5 X 104 cells/100μl씩 분주한 뒤, 24 시간 동안 CO2 배양하였다. LPS (lipopolysaccharide)를 1μg/ml 농도로 처리하고 1시간 CO₂ 배양한 뒤 준비한 고들빼기 추출물을 농도별로 처리하여 24 시간 동안 CO₂배양하였다. 그 다음 각 배양 상층액을 96웰 플레이트에 넣고 여기에 동량의 Griess 시약 (0.1% N-1-naphtyl-ethylendiamine in H2O : 1% sulfanilamide in 5% H3PO4 = 1 : 1)을 첨가하여 10분간 반응시킨 후, 마이크로플레이트 리더로 550nm에서 흡광도를 측정하였다. NO2 (Nitrite)의 농도는 NaNO2 (sodium nitrite)를 100 μM에서부터 0.7 μM 까지 2배씩 희석하여 얻은 표준곡선과 비교하여 계산하였다. LPS는 양성대조군으로 사용하였다.NO 2 (Nitrite), a stabilized NO oxide, was measured using the Griess reaction. First, a mouse macrophage cell line (RAW264.7) was dispensed in a 96-well plate at 5 X 10 4 cells/100 μl per well, followed by CO 2 culture for 24 hours. After treatment with LPS (lipopolysaccharide) at a concentration of 1 μg/ml and incubation with CO2 for 1 hour, the prepared extracts from the prepared extract were treated with concentration and cultured with CO2 for 24 hours. Then, each culture supernatant was placed in a 96-well plate and the same amount of Griess reagent (0.1% N-1-naphtyl-ethylendiamine in H 2 O : 1% sulfanilamide in 5% H 3 PO 4 = 1 : 1) was added thereto. After reacting for 10 minutes, the absorbance was measured at 550 nm using a microplate reader. The concentration of NO 2 (Nitrite) was calculated by comparing with a standard curve obtained by diluting NaNO 2 (sodium nitrite) from 100 μM to 0.7 μM by 2 times. LPS was used as a positive control.
그 결과는 하기 표 3에 나타내었다.The results are shown in Table 3 below.
결과에서 보듯이, 본 발명의 실시예 1의 유산균 수정과는 NO를 억제하는 것을 확인하였다.As shown in the results, it was confirmed that the fertilized lactic acid bacteria of Example 1 of the present invention suppressed NO.
<시험예 4> 유산균 수정과의 대식세포주 (RAW 264.7)에서 LPS에 의해 유도된 염증성 사이토카인 (Cytokine)에 미치는 영향 측정<Test Example 4> Measurement of the effect on inflammatory cytokines induced by LPS in a macrophage cell line (RAW 264.7) of the fertilized lactobacillus family
대식세포주인 RAW 264.7 세포는 한국세포주은행(KTCC)에서 분양받아 10% FBS(fetalbovine serum), 1% 항생제, 2-ME(2-mercaptoethanol)를 함유한 RPMI 1640 배지를 이용하여 37℃, 4% CO2에서 조절된 인큐베이터에서 배양하였다.RAW 264.7 cells, a macrophage cell line, were purchased from the Korea Cell Line Bank (KTCC) and cultured at 37°C and 4% in RPMI 1640 medium containing 10% fetalbovine serum (FBS), 1% antibiotics, and 2-mercaptoethanol (2-ME). Cultivated in a controlled incubator at CO 2 .
RAW264.7 대식세포를 24-웰 플레이트에 well당 5 x 105cells/1ml이 되도록 분주하고 24시간 CO₂배양하였다. 이후 well에 LPS (lipopolysaccharide)를 1μg/ml 농도, 유산균 수정과를 농도별로 처리하여 24시간 CO₂ 배양하였다. 그 다음 세포배양 상층액을 수거하였다. 상층액에 포함된 사이토카인 (cytokine)인 IL-6, TNF-α, GM-CSF, IL-1β를 효소항체법 (enzyme-linked immunosorbent assay : ELISA) 을 이용하여 측정하였다. 즉, plate-bottom micro-well에 1차 항체 (capture antibody)를 코팅액(0.1 M sodium carbonate, pH 9.5)에 희석하여 100 μl/well로 분주하고 4℃에서 밤새 배양한 후, 세척용액 (0.05% Tween 20/PBS)으로 세척하였다. 세척된 마이크로웰은 10% FBS가 첨가된 PBS로 블로킹(blocking)하였으며, 실험에서 채취한 배양 상층액을 적당한 비율로 희석한 후 각 well에 분주하여 상온에서 반응시켰다. 그 다음, 바이오틴이 부착된 2차 항체 100㎕/well와 일정시간 상온에서 반응시킨 후, 아비딘-퍼옥시다제 (enzyme reagent) 100 μl/well을 첨가하였다. 마지막으로 기질 (3,3′,5,5′-Tetramethylbenzidine Liquid Substrate, H2O2) 을 첨가하여 발색시킨 다음 마이크로플레이트 리더를 이용하여 측정하였다. 측정된 IL-6, TNF-α, GM-CSF, IL-1β의 농도는 표준곡선을 이용하여 환산하였다.RAW264.7 macrophages were dispensed so as to be 5 x 10 5 cells/1ml per well in a 24-well plate and cultured with CO2 for 24 hours. Then, LPS (lipopolysaccharide) was treated at 1μg/ml concentration and lactic acid fertilized fruit by concentration, followed by CO2 incubation for 24 hours. Then, the cell culture supernatant was collected. IL-6, TNF-α, GM-CSF, and IL-1β, which are cytokines contained in the supernatant, were measured using an enzyme-linked immunosorbent assay (ELISA). That is, the primary antibody (capture antibody) is diluted in the coating solution (0.1 M sodium carbonate, pH 9.5) and dispensed at 100 μl/well in the plate-bottom micro-well, incubated overnight at 4°C, and washed with a washing solution (0.05% Tween 20/PBS). The washed microwells were blocked with PBS supplemented with 10% FBS, and the culture supernatant collected in the experiment was diluted in an appropriate ratio and then dispensed into each well and reacted at room temperature. Then, after reacting with 100 μl/well of a biotin-attached secondary antibody at room temperature for a certain period of time, 100 μl/well of avidin-peroxidase (enzyme reagent) was added. Finally, a substrate (3,3′,5,5′-Tetramethylbenzidine Liquid Substrate, H 2 O 2 ) was added to develop color, and then measured using a microplate reader. The measured concentrations of IL-6, TNF-α, GM-CSF, and IL-1β were converted using a standard curve.
그 결과는 하기 표 4에 나타내었다.The results are shown in Table 4 below.
(1 μg/ml)LPS
(1 µg/ml)
결과에서 보듯이, 유산균 수정과가 RAW 264.7 세포에서 LPS에 의해 유도된 염증성 사이토카인에 미치는 영향을 확인한 결과 IL-6, IL-1β, TNF-α, GM-CSF의 염증전 사이토카인(pro-inflammatory cytokine)을 모두 통계적으로 유의하게 억제하는 것을 확인하였다. 이 결과를 통해 유산균 수정과가 RAW 264.7 세포에서 염증 매개성 사이토카인을 효과적으로 억제하여 항염증 기능에 관여하는 것을 확인할 수 있다.As shown in the results, the effects of Lactobacillus fernaceae on LPS-induced inflammatory cytokines in RAW 264.7 cells were confirmed, and as a result, IL-6, IL-1β, TNF-α, and pro-inflammatory cytokines of GM-CSF (pro- inflammatory cytokines) were all statistically significantly inhibited. Through this result, it can be confirmed that Lactobacillus fernaceae is involved in anti-inflammatory function by effectively suppressing inflammatory mediating cytokines in RAW 264.7 cells.
<시험예 5> 피로회복 개선 효과 확인<Test Example 5> Confirmation of fatigue recovery improvement effect
실시예 1의 유산균 수정과 및 비교예 1의 일반 수정과를 10명씩 10~40대의 남녀노소 20명에게 30일간, 매일 하루 3번 110㎖씩 점심식사 후 2시간 후에 복용하게 하였으며, 30일간의 복용 후, 소화기능 개선, 무기력감 개선 및 피로감 개선 정도를 하기 표 5에 5점 타점법(5점 : 아주 우수함, 4점 : 우수함, 3점 : 보통, 2점 : 나쁨, 1점 : 아주나쁨)으로 나타내게 하였다. 이를 위해, 소화기능 개선 효과, 무기력감 개선 효과, 전체적인 피로감 개선 효과 등을 확인하였다. Lactobacillus fertilized fruit of Example 1 and general fertilized fruit of Comparative Example 1 were given to 20 men and women in their 10s to 40s by 10 people each for 30 days, 3 times a day, 110㎖ 2 hours after lunch, 30 days After taking, the degree of improvement in digestive function, lethargy, and fatigue was evaluated by the 5-point scoring method in Table 5 below (5 points: very good, 4 points: excellent, 3 points: normal, 2 points: bad, 1 point: very bad) was shown as To this end, the digestive function improvement effect, lethargy improvement effect, overall fatigue improvement effect, etc. were confirmed.
개선 효과digestive function
improvement effect
개선 효과helplessness
improvement effect
개선 효과Fatigue
improvement effect
상기 표 5에서 보듯이, 본 발명의 유산균 수정과는 소화기능 개선 효과가 매우 우수하며, 무기력을 감소시키는 효과가 있었으며, 전체적인 피로감을 개선시켰다. 특히 점심식사 후 섭취하였는데 소화기능이 오히려 향상되었으며, 피로감이 개선되어 오후에 공부 또는 일에 보다 집중할 수 있었다고 평가하였다.As shown in Table 5, the fertilized lactic acid bacteria of the present invention was very effective in improving digestive function, had an effect of reducing lethargy, and improved overall fatigue. In particular, when taken after lunch, it was evaluated that the digestive function was rather improved, and fatigue was improved, so that it was possible to concentrate more on study or work in the afternoon.
<시험예 6> 체중 및 혈중 콜레스테롤 감소 실험<Test Example 6> Body weight and blood cholesterol reduction test
실험동물로 ICR 마우스를 이용하여, 상기 실시예 1로부터 제조된 유산균 수정과 및 비교예 1의 일반 수정과를 재료로 한 시험대조군과 정상군을 식이 급여하며 사육 비교하였다.Using ICR mice as experimental animals, breeding and comparison were performed between a test control group and a normal group using the fertilized lactobacillus prepared in Example 1 and the fertilized fruit of Comparative Example 1 as materials.
본 실험에서 기본 식이는 AIN-76을 기본으로 단백질급원은 카제인(Japan), 탄수화물급원은 옥수수전분(제일제당), 지방급원으로 옥수수유(대상)을 사용한 정상군과 시험대조군을 비교하는 방식으로 진행하였다. 본 실험에 사용한 유산균 수정과는 매일 일정시간에 1 ml씩 경구 급여하였으며 식이와 식수는 임의로 자유롭게 섭취하도록 하였다. 시험동물 사육환경은 항온(20±2℃), 항습(50±5%)하고 12 시간 광주기로 일정 조건을 유지하면서 폴리카보네이트 케이지에서 2 마리씩 분리하여 사육하였다. 동물에서 콜레스테롤은 혈류를 따라 순환하며 몇몇 기관에서 합성되며 혈류 내에서 콜레스테롤 수치가 크면 동맥경화의 주요 원인이 되므로 분석항목은 (1) 체중변화 및 식이섭취량, (2) 혈중 콜레스테롤 및 중성지질을 측정하는 것으로 진행하였다. In this experiment, the basic diet was AIN-76, the protein source was casein (Japan), the carbohydrate source was corn starch (CheilJedang), and the fat source was corn oil (subject). proceeded. Lactic acid fertilized fruit used in this experiment was fed orally by 1 ml every day at a certain time, and food and drinking water were freely consumed. The test animal breeding environment was kept at constant temperature (20 ± 2 ° C), constant humidity (50 ± 5%), and kept at constant conditions with a 12-hour photoperiod, while breeding by separating two animals in a polycarbonate cage. In animals, cholesterol circulates along the bloodstream and is synthesized in several organs, and high cholesterol levels in the bloodstream are the main cause of arteriosclerosis. Therefore, analysis items are (1) weight change and dietary intake, (2) blood cholesterol and triglyceride proceeded to do.
사육이 끝난 실험동물은 12 시간 절식시킨 후 에테르(Ether)를 흡입시켜 마취시킨 다음 복부 하대정맥으로부터 채혈하였으며, 헤파린을 처리하여 원심분리(15 mins, 3,000 RPM)로 혈장을 분리하여 지질함량을 측정하였다.After breeding, the experimental animals were fasted for 12 hours, anesthetized by inhalation of ether, blood was collected from the inferior vena cava, treated with heparin, and plasma was separated by centrifugation (15 mins, 3,000 RPM) to measure lipid content. did
각 실험동물의 내장(간, 부고환, 신장)을 혈액채취 후 즉시 적출하여 PBS(Phosphate Buffered saline) 용액으로 수차례 헹군 후 표면 수분을 제거하여 비교 정량하였다. 혈장 중성지질은 McGowan 등(1983)의 효소법을 이용한 발색방식의 중성지방 측정용 시액(Asan kit)를 사용하여 550 nm에서 흡광도를 측정하였으며, 혈장 총 콜레스테롤은 Allain 등(1974)의 효소법을 응용한 총콜레스테롤 측정용 시액(Asan kit)를 사용하여 발색처리하여 500 nm에서 흡광도를 측정하여 이를 콜레스테롤 표준곡선과 비교 정량하는 방식으로 하였으며, 장기의 지질은 Folch 등(1957)의 방법으로 잘게 자른 후 2 ml, Chloroform: Methyl alcohol 2:1 용액하에서 추출하여 중성지질, 콜레스테롤은 혈장에서와 동일 방법으로 정량하였다. 각 항목은 통계처리하여 평균값으로 하였다. The intestines (liver, epididymis, kidney) of each experimental animal were removed immediately after blood collection, rinsed several times with a PBS (Phosphate Buffered saline) solution, and then surface moisture was removed for comparative quantification. Plasma triglyceride was measured for absorbance at 550 nm using the assay (Asan kit) for triglyceride measurement using the enzymatic method of McGowan et al. (1983), and plasma total cholesterol was measured using the enzymatic method of Allain et al. The total cholesterol measurement solution (Asan kit) was used for color development, and the absorbance was measured at 500 nm, and the result was compared and quantified with the cholesterol standard curve. ml, Chloroform: Extracted under a 2:1 solution of methyl alcohol, and triglyceride and cholesterol were quantified in the same way as in plasma. Each item was statistically processed and averaged.
(1) 체중변화(1) Weight change
실험동물의 체중변화 및 식이섭취량을 하기 표 6 및 7에 각각 나타내었다. Changes in body weight and food intake of the experimental animals are shown in Tables 6 and 7, respectively.
하기 표 6을 참조하면, 실시예 1로부터 제조된 유산균 수정과를 경구 급여한 실험동물의 경우 실험전에 비하여 체중이 유사 또는 저하되었고, 반면 비교예 1로부터 제조된 일반 수정과를 경구 급여한 실험동물 및 정상군의 경우 실험 전에 비하여 체중이 증가했음을 확인할 수 있다.Referring to Table 6 below, in the case of laboratory animals fed orally with the fertilized fruit of lactobacillus prepared in Example 1, the body weight was similar or decreased compared to before the experiment, whereas the experimental animals fed orally with the fertilized fruit prepared in Comparative Example 1 And in the case of the normal group, it can be confirmed that the weight increased compared to before the experiment.
하기 표 7을 참조하면, 실시예 1로부터 제조된 유산균 수정과 및 비교예 1의 일반 수정과를 경구 급여한 실험동물 및 정상군의 식이섭취량은 모두 유사한 것으로 확인하였다.Referring to Table 7 below, it was confirmed that the dietary intake of the fertilized lactobacillus prepared in Example 1 and the fertilized fruit of Comparative Example 1 was orally fed to both the experimental animals and the normal group.
이를 통하여 모든 실험동물의 식이 섭취량이 유사함에도 불구하고, 실시예 1의 유산균 수정과를 섭취한 실험동물의 체중이 모두 증가하지 않고, 실험 전과 유사하거나 실험 전에 비하여 감소하여 다이어트 음료로서 활용 가능함을 확인하였다.Through this, despite the similar dietary intake of all laboratory animals, the body weight of the laboratory animals ingesting the lactobacillus fertilized fruit of Example 1 did not increase, and was similar to before the experiment or decreased compared to before the experiment, confirming that it can be used as a diet drink. did
(2) 혈중 콜레스테롤 및 중성지질(2) Blood cholesterol and triglyceride
실험동물의 혈장 중성지질, 혈장 콜레스테롤, 부고환 중성지질, 신장 조직의 중성지질 및 간조직에서의 콜레스테롤을 하기 표 8 및 9에 나타내었다.Plasma triglyceride, plasma cholesterol, epididymal triglyceride, kidney tissue neutral lipid, and cholesterol in liver tissue of experimental animals are shown in Tables 8 and 9 below.
하기 표 8을 참조하면, 실시예 1의 유산균 수정과를 섭취한 시험대조군은 실험 전에 비해 혈장 중성지질이 현저히 감소한 반면, 비교예 1의 일반 수정과를 섭취한 시험대조군은 유의적인 변화가 나타나지 않았다.Referring to Table 8 below, the test control group ingesting the fertilized fruit of lactic acid bacteria of Example 1 showed a significant decrease in plasma triglyceride compared to before the experiment, whereas the test control group ingesting the normal fertilized fruit of Comparative Example 1 showed no significant change. .
하기 표 9를 참조하면, 실시예 1의 유산균 수정과를 섭취한 시험대조군은 실험 전에 비해 혈장 콜레스테롤이 현저히 감소한 반면, 비교예 1은 유의적인 변화가 나타나지 않았음을 확인할 수 있다.Referring to Table 9 below, it can be seen that the test control group ingesting fertilized fruit of lactic acid bacteria in Example 1 significantly decreased plasma cholesterol compared to before the experiment, whereas in Comparative Example 1, no significant change was observed.
상기 결과를 통하여, 본 발명에 따른 실시예 1에서 제조된 유산균 수정과는 중성지질의 양의 감소현상을 확인할 수 있어 지방과 콜레스테롤을 개선시키는 것을 확인하였다. Through the above results, it was confirmed that the modified lactic acid bacteria prepared in Example 1 according to the present invention improved fat and cholesterol by confirming the decrease in the amount of neutral lipids.
<시험예 7> 장기능 개선 및 변비 개선효과 <Test Example 7> Improvement of intestinal function and constipation
실시예 1에서 제조된 유산균 수정과의 장기능 개선 및 변비 개선에 대한 효과를 알아보기 위하여 하기와 같은 방법을 수행하여 실시하였다.In order to examine the effect on the improvement of intestinal function and constipation improvement of the lactic acid bacteria fertilization prepared in Example 1, the following method was performed and carried out.
배변량을 측정하기 위해, 동물모델로 웅성의 평균 체중 220-240g 정도의 스프라그 도울리(Sprague Dawley) 랫트 (대한바이오링크, 음성)를 1군당 8 내지 10 마리를 사용하여 대사 케이지에서 3일간 순화 적응시키고, 4일째부터 변비를 유발시키기 위해 로페라마이드 (Sigma사, 미국)를 사료 3g당 1mg 함유하는 사료를 공급하였다. 대조군(로페라마이드군 처리군)은 식수만을 공급하였으며, 실시예 1의 유산균 수정과를 각각 3.2㎎/㎖ 농도로 식수에 녹여 공급하여 실험 종료시까지 투여하였다. 변은 매일 채취하여 무게를 측량하였다.To measure the amount of defecation, as an animal model, 8 to 10 Sprague Dawley rats (Daehan Biolink, Negative) with an average male weight of about 220-240g were acclimatized in a metabolic cage for 3 days. After adaptation, a feed containing 1 mg of loperamide (Sigma, USA) per 3 g of feed was supplied to induce constipation from the 4th day. The control group (loperamide group treatment group) was supplied with only drinking water, and the Lactobacillus modified fruit of Example 1 was dissolved in drinking water at a concentration of 3.2 mg/ml, respectively, and administered until the end of the experiment. Feces were collected daily and weighed.
조성물의 변비 개선 효과를 실험한 결과, 대조군에 비하여 변비유발 기간동안에 배변량을 증가시켜 변비개선 효과가 있음을 확인하였다.As a result of testing the constipation improvement effect of the composition, it was confirmed that there is an effect of improving constipation by increasing the amount of defecation during the constipation induction period compared to the control group.
<시험예 8> 관능검사<Test Example 8> Sensory test
본 발명의 유산균 수정과의 관능성을 평가하기 위하여 실시예 1에서 제조된 유산균 수정과 및 비교예 1의 일반 수정과를 비교하는 관능테스트를 실시하였다. In order to evaluate the functionality of the modified lactic acid bacteria of the present invention, a sensory test was conducted to compare the modified lactic acid bacteria prepared in Example 1 and the modified fertilized lactic acid bacteria of Comparative Example 1.
관능검사 요원 30명을 대상으로 향, 맛, 색상, 조직감 및 전체적인 기호도에 대한 관능검사를 3회 반복하여 아래의 표 11과 같은 결과를 얻었다. The sensory test for aroma, taste, color, texture, and overall preference was repeated three times for 30 sensory test agents, and the results shown in Table 11 below were obtained.
검사 방법으로는 5 점 비교 척도법을 사용하였으며, 아주 좋다는 5 점, 좋다 는 4 점, 보통이다는 3 점, 싫다는 2 점, 아주 싫다는 1의 점수를 각각 부여하고, 최종 응답 된 점수를 합하여 산술평균값을 연산하였다. As a test method, a 5-point comparison scale method was used, and 5 points for very good, 4 points for good, 3 points for average, 2 points for dislike, and 1 point for very dislike were given respectively, and the final responses were summed. The arithmetic average value was calculated.
실험결과 본 발명에 따른 실시예 1의 유산균 수정과는 향, 맛, 색상 및 식감이 매우 높아 전체적인 기호도가 5인 한편, 비교예 1에서 제조된 일반 수정과는 전체적인 기호도가 본 발명의 유산균 수정과에 비하여 3.2로 매우 낮았다. 이로부터 유산균 대사물질이 수정과와 매우 융합이 잘되어 관능적 특성에 상승효과를 부여한다는 것을 확인하였다.As a result of the experiment, the modified lactic acid bacteria of Example 1 according to the present invention has a very high aroma, taste, color and texture, and the overall preference is 5, while the general modified fruit produced in Comparative Example 1 has an overall preference compared to the modified lactic acid bacteria of the present invention. compared to 3.2, which was very low. From this, it was confirmed that the lactic acid bacteria metabolites were very well fused with fertilized fruits, giving a synergistic effect to sensory characteristics.
이상으로 본 발명의 특정한 부분을 상세히 기술하였는바, 당업계의 통상의 지식을 가진 자에게 있어서 이러한 구체적인 기술은 단지 바람직한 구현예일 뿐이며, 이에 본 발명의 범위가 제한되는 것이 아닌 점은 명백하다. 따라서, 본 발명의 실질적인 범위는 첨부된 청구항과 그의 등가물에 의하여 정의된다고 할 것이다.Having described specific parts of the present invention in detail above, it is clear that these specific techniques are only preferred embodiments for those skilled in the art, and the scope of the present invention is not limited thereto. Accordingly, the substantial scope of the present invention will be defined by the appended claims and equivalents thereof.
Claims (6)
(S1) 계피, 생강 및 곶감에 각각 정제수를 혼합하여 80 내지 100℃에서 110 내지 130분 동안 가열하여 계피 추출물, 생강 추출물 및 곶감 추출물을 각각 수득하는 단계;
(S2) 상기 단계 (S1)에서 수득된 계피 추출물, 생강 추출물 및 곶감 추출물을 혼합한 다음 유산균 대사산물, 프락토올리고당과 설탕 혼합물 및 곶감을 첨가하는 단계;
(S3) 상기 단계 (S2)에서 수득된 혼합물을 95 내지 100℃에서 50 내지 70분 동안 가열하여 농축하는 단계;
(S4) 상기 단계 (S3)에서 수득된 유산균 수정과를 살균 및 여과하는 단계; 및
(S5) 상기 단계 (S4)에서 수득된 유산균 수정과를 60 내지 80℃에서 식힌 후 충진기로 이송하여 충진하여 포장하는 단계.A method for producing fertilized lactic acid bacteria, characterized in that it comprises the following steps:
(S1) mixing cinnamon, ginger, and dried persimmon with purified water and heating at 80 to 100° C. for 110 to 130 minutes to obtain cinnamon extract, ginger extract, and dried persimmon extract, respectively;
(S2) mixing the cinnamon extract, ginger extract, and dried persimmon extract obtained in step (S1) and then adding lactic acid bacteria metabolites, fructooligosaccharide and sugar mixture, and dried persimmon;
(S3) concentrating the mixture obtained in step (S2) by heating at 95 to 100° C. for 50 to 70 minutes;
(S4) sterilizing and filtering the fertilized fruit of lactic acid bacteria obtained in step (S3); and
(S5) Cooling the fertilized fruit of lactic acid bacteria obtained in step (S4) at 60 to 80 ° C, transferring it to a filling machine, filling and packaging.
상기 계피 추출물, 생강 추출물 및 곶감 추출물을 1.6:1.5:1.2의 중량비로 혼합하는 것을 특징으로 하는 유산균 수정과의 제조방법.According to claim 1,
A method for producing lactic acid bacteria fertilized fruit, characterized in that mixing the cinnamon extract, ginger extract and dried persimmon extract in a weight ratio of 1.6: 1.5: 1.2.
상기 유산균 대사물질이 락토바실러스 플란타룸(L.plantarum), 락토바실러스 카제이(L.casei) 및 비피도박테리움 인펀티스(B. infantis)로 이루어진 군에서 선택된 하나 이상의 유산균을 액체 배지에 접종하고 20 내지 40℃에서 12 내지 48시간 배양하여 수득된 발효액을 동결건조하여 제조되는 것을 특징으로 하는 유산균 수정과의 제조방법.According to claim 1,
The lactic acid bacteria metabolites Lactobacillus plantarum ( L.plantarum ), Lactobacillus casei ( L.casei ) and Bifidobacterium infantis ( B. infantis ) Inoculate one or more lactic acid bacteria selected from the group consisting of liquid medium And a method for producing lactic acid bacteria fertilized fruit, characterized in that produced by freeze-drying the fermentation broth obtained by culturing at 20 to 40 ° C. for 12 to 48 hours.
상기 단계 (S2)에서 프락토올리고당과 설탕은 2:8의 중량비로 혼합된 것을 특징으로 하는 유산균 수정과의 제조방법.According to claim 1,
In the step (S2), fructooligosaccharide and sugar are mixed in a weight ratio of 2:8.
상기 단계 (S3)에서 농축은 95 내지 100 ℃에서 50 내지 70분 동안 수행한 후
상기 단계 (S4)에서 살균은 95 내지 100 ℃에서 20 내지 30분 동안 수행하는 것을 특징으로 하는 유산균 수정과의 제조방법.According to claim 1,
In the step (S3), the concentration is performed at 95 to 100 ° C. for 50 to 70 minutes, and then
In the step (S4), sterilization is performed at 95 to 100 ° C. for 20 to 30 minutes.
A fertilized lactic acid bacteria prepared by the method according to any one of claims 1 to 5.
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| KR1020210082291A KR20230001021A (en) | 2021-06-24 | 2021-06-24 | A process for the preparation of cinnamon punch containing lactobacillus and the cinnamon punch containing lactobacillus prepared therefrom |
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| KR (1) | KR20230001021A (en) |
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2021
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