KR20220125966A - Composition for Preventing or Aemliorating Muscular Diseases Comprising Lactobacillus Plantarum HY7715 - Google Patents
Composition for Preventing or Aemliorating Muscular Diseases Comprising Lactobacillus Plantarum HY7715 Download PDFInfo
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- KR20220125966A KR20220125966A KR1020210029942A KR20210029942A KR20220125966A KR 20220125966 A KR20220125966 A KR 20220125966A KR 1020210029942 A KR1020210029942 A KR 1020210029942A KR 20210029942 A KR20210029942 A KR 20210029942A KR 20220125966 A KR20220125966 A KR 20220125966A
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- A23C9/00—Milk preparations; Milk powder or milk powder preparations
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- A23C9/1234—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt characterised by using a Lactobacillus sp. other than Lactobacillus Bulgaricus, including Bificlobacterium sp.
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Abstract
Description
본 발명은 락토바실러스 플란타럼 HY7715를 포함하는 근육질환 예방 또는 개선용 조성물에 관한 것이다.The present invention relates to a composition for preventing or improving muscle disease comprising Lactobacillus plantarum HY7715.
한국인의 기대수명은 경제 성장과 함께 다른 나라 대비 빠르게 증가하였다. 1960년 52.4세에 불과했던 기대수명이 1990년에는 71.4세, 2016년에는 82.4세로 증가하였다. 하지만 건강수명은 2016년 평균 64.9세로, 생애 마지막 17.5년은 건강 문제로 활동에 제약을 받으며 산다.Life expectancy of Koreans increased faster than other countries along with economic growth. Life expectancy increased from 52.4 years in 1960 to 71.4 years in 1990 and 82.4 years in 2016. However, the average healthy life expectancy in 2016 was 64.9 years, and the last 17.5 years of life are limited by health problems.
기대수명 대비 건강수명이 낮은 이유 중 한가지는 근육이다. 한국인은 시대 발전에 따라 기대 수명이 연장되었으나, 운동량이 적고, 노화로 인해 노년기에 접어들수록 급격히 근육의 양이 감소한다. 일반적으로 50세 이후 근육은 매년 1~2% 감소하고, 10년이면 평균 4kg 정도 감소하는 것으로 알려져 있다. 또한, 65세엔 중년 때 근육의 평균 약 25~35% 정도가 감소하고, 80세엔 40%이상 감소하면서 기력이 떨어진다. 나이가 들면서 겪게 되는 근감소증은 단순히 근력이 감소하는 것을 넘어 면역력을 저하시키며, 고지혈증, 당뇨, 관절질환 등 각종 질환 발병률을 높인다.One of the reasons for the low health life expectancy compared to life expectancy is muscle. The life expectancy of Koreans has been extended according to the development of the times, but the amount of exercise is low and the amount of muscle rapidly decreases as they enter old age due to aging. In general, it is known that after the age of 50, muscle decreases by 1 to 2% every year, and an average of 4 kg in 10 years. In addition, at the age of 65, an average of about 25-35% of the muscle in middle age decreases, and at the age of 80, the energy decreases by more than 40%. Sarcopenia experienced with aging not only decreases muscle strength, but also lowers immunity and increases the incidence of various diseases such as hyperlipidemia, diabetes, and joint diseases.
따라서 앞으로 더 심해질 고령화 시대에 노화로 인해 줄어드는 근육이 감소하는 것을 억제해주거나, 개선할 수 있는 프로바이오틱스, 음료, 건강식품과 같은 제품 개발이 필요하다. Therefore, it is necessary to develop products such as probiotics, beverages, and health foods that can suppress or improve muscle loss due to aging in the aging population, which will become more severe in the future.
일 양상은 락토바실러스 플란타럼(Lactobacillus plantarum) HY7715를 포함하는 근육질환 예방 또는 개선용 식품 조성물을 제공하는 것이다. One aspect is to provide a food composition for preventing or improving muscle disease comprising Lactobacillus plantarum HY7715.
다른 양상은 락토바실러스 플란타럼(Lactobacillus plantarum) HY7715를 포함하는 근기능 강화용 식품 조성물을 제공하는 것이다. Another aspect is to provide a food composition for strengthening muscle function comprising Lactobacillus plantarum HY7715.
일 양상은 락토바실러스 플란타럼(Lactobacillus plantarum) HY7715를 포함하는 근육질환 예방 또는 개선용 식품 조성물을 제공하는 것이다. One aspect is to provide a food composition for preventing or improving muscle disease comprising Lactobacillus plantarum HY7715.
상기 락토바실러스 플란타럼 HY7715는 한국 생명공학 연구원에 2016년 9월 8일자로 수탁하여, 수탁번호 KCTC13101BP를 부여 받은 것이다.The Lactobacillus plantarum HY7715 was entrusted to the Korea Institute of Biotechnology and Biotechnology on September 8, 2016, and was given an accession number KCTC13101BP.
상기 락토바실러스 플란타럼 HY7715는 생균체일 수 있고, 상기 락토바실러스 플란타럼 HY7715를 배양한 배양액, 상기 배양액의 농축액, 상기 농축액을 동결건조한 동결건조분말 일 수 있다. The Lactobacillus plantarum HY7715 may be a live cell, a culture solution of the Lactobacillus plantarum HY7715, a concentrate of the culture solution, and a freeze-dried powder obtained by freeze-drying the concentrate.
일 구체예에 따르면, 상기 근육질환은 노화로 인한 근기능 저하, 근육 소모, 근육 퇴화일 수 있다. 상기 노화로 인한 근기능 저하는 근감소증을 의미하는 사코페니아(Sarcopenia)를 포함할 수 있다.According to one embodiment, the muscle disease may be a decrease in muscle function due to aging, muscle wasting, or muscle degeneration. The decrease in muscle function due to aging may include Sarcopenia, which means sarcopenia.
일 실시예에 따르면, 어린 마우스 대비 노령의 마우스에서 차이를 보였던 근피로도 증가, 근감소증 유발 전사인자 감소, 근섬유 감소, 근육분화 전사인자 감소, 및 미토콘드리아 활성이 감소되었다. 반면, 락토바실러스 플란타럼 HY7715를 섭취한 노령의 마우스는 근피로도가 감소되었으며, 근감소증 유발 전사인자 및 근육분화 전사인자가 증가되었으며, 근섬유 및 미토콘드리아 활성이 증가되는 것을 확인하였다.According to one embodiment, increased muscle fatigue, decreased sarcopenia-induced transcription factors, decreased muscle fibers, decreased muscle differentiation transcription factors, and mitochondrial activity were decreased in the aged mice compared to the young mice. On the other hand, it was confirmed that the aged mice that received Lactobacillus plantarum HY7715 had reduced muscle fatigue, increased sarcopenia-induced transcription factors and muscle differentiation transcription factors, and increased muscle fiber and mitochondrial activity.
일 구체예에 따르면, 상기 근육질환은 가자미근과 비복근에 해당하는 근육에서 나타나는 것일 수 있다. According to one embodiment, the muscle disease may appear in muscles corresponding to the soleus and gastrocnemius muscles.
본 명세서의 용어 '예방'은 락토바실러스 플란타럼 HY7715를 포함하는 조성물을 이용하여 근육질환의 진행을 지연시키거나, 발병을 억제하는 모든 행위를 의미한다.As used herein, the term 'prevention' refers to any action that delays the progression of muscle disease or inhibits the onset of a muscle disease by using a composition containing Lactobacillus plantarum HY7715.
본 명세서의 용어 '개선'은 락토바실러스 플란타럼 HY7715를 포함하는 조성물을 이용하여 근육질환의 증상을 호전시키는 모든 행위를 의미한다. As used herein, the term 'improvement' refers to any action for improving symptoms of muscle disease by using a composition containing Lactobacillus plantarum HY7715.
상기 식품 조성물은 식품을 섭취하는 개체의 무게(kg) 당 락토바실러스 플란타럼 HY7715를 1X109 CFU를 섭취하도록 제조될 수 있으나, 이에 제한되는 것은 아니다. The food composition may be prepared to consume 1X10 9 CFU of Lactobacillus plantarum HY7715 per weight (kg) of an individual ingesting food, but is not limited thereto.
본 발명의 조성물이 식품 조성물로 제조되는 경우, 락토바실러스 플란타럼 HY7715 외에, 식품 제조 시에 통상적으로 첨가되는 성분을 포함할 수 있으며, 예를 들어, 단백질, 탄수화물, 지방, 영양소, 조미제 및 향미제를 포함할 수 있다. 탄수화물의 예는 모노사카라이드, 예를 들어, 포도당, 과당 등; 디사카라이드, 예를 들어, 말토오스, 수크로오스, 올리고당 등; 및 폴리사카라이드, 예를 들어, 덱스트린, 사이클로덱스트린 등과 같은 통상적인 당 및 자일리톨, 소르비톨, 에리트리톨 등의 당알콜일 수 있다. 향미제로서 천연 향미제[타우마틴, 스테비아 추출물(예를 들어, 레바우디오시드 A, 글리시르히진 등)] 및 합성 향미제(사카린, 아스파르탐 등)를 사용할 수 있다.When the composition of the present invention is prepared as a food composition, in addition to Lactobacillus plantarum HY7715, it may include ingredients commonly added during food production, for example, proteins, carbohydrates, fats, nutrients, seasonings and Flavoring agents may be included. Examples of carbohydrates include monosaccharides such as glucose, fructose, and the like; disaccharides such as maltose, sucrose, oligosaccharides and the like; and polysaccharides, for example, conventional sugars such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As the flavoring agent, natural flavoring agents [taumatin, stevia extract (eg, rebaudioside A, glycyrrhizin, etc.)] and synthetic flavoring agents (saccharin, aspartame, etc.) can be used.
예를 들어, 본 발명의 식품 조성물이 드링크제로 제조되는 경우에는 본 발명의 녹용 추출물의 유산균 발효물 외에 구연산, 액상과당, 설탕, 포도당, 초산, 사과산, 과즙, 두충 추출액, 대추 추출액 및/또는 감초 추출액 등이 추가로 포함될 수 있다.For example, when the food composition of the present invention is prepared as a drink, in addition to the lactic acid bacteria fermented product of the antler extract of the present invention, citric acid, high fructose, sugar, glucose, acetic acid, malic acid, fruit juice, cephalopod extract, jujube extract and/or licorice Extracts and the like may be additionally included.
또한, 본 발명의 식품 조성물은 여러 가지 영양제, 비타민, 광물(전해질), 합성 풍미제 및 천연 풍미제 등의 풍미제, 착색제 및 중진제(치즈, 초콜릿 등), 펙트산 및 그의 염, 알긴산 및 그의 염, 유기산, 보호성 콜로이드 증점제, pH 조절제, 안정화제, 방부제, 글리세린, 알콜, 탄산음료에 사용되는 탄산화제 등을 함유할 수 있다. In addition, the food composition of the present invention includes various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and thickening agents (cheese, chocolate, etc.), pectic acid and its salts, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated beverages, and the like.
이러한 성분은 독립적으로 또는 조합하여 사용할 수 있으며, 이러한 첨가제의 비율은 본 발명의 식품 조성물 100 중량부 당 0 내지 약 20 중량부의 범위에서 선택될 수 있으나, 이에 한정되는 것은 아니다.These components may be used independently or in combination, and the ratio of these additives may be selected from 0 to about 20 parts by weight per 100 parts by weight of the food composition of the present invention, but is not limited thereto.
일 구체예에 따르면, 상기 식품 조성물은 발효유, 기능성음료, 건강기능식품 중에서 어느 하나일 수 있으나, 이에 제한되는 것은 아니다. According to one embodiment, the food composition may be any one of fermented milk, functional beverage, and health functional food, but is not limited thereto.
다른 양상은 락토바실러스 플란타럼 HY7715를 포함하는 근기능 강화용 식품 조성물을 제공하는 것이다. Another aspect is to provide a food composition for strengthening muscle function comprising Lactobacillus plantarum HY7715.
상기 '락토바실러스 플란타럼 HY7715' 및 '식품 조성물'에 관한 내용은 상기에 기재한 내용과 동일하다. The contents of the 'Lactobacillus plantarum HY7715' and the 'food composition' are the same as those described above.
일 구체예에 따르면, 상기 근기능 강화는 근섬유 합성, 근육 분화, 근육 피로도 감소, 근육량 증가를 나타내는 것일 수 있고, 근육량의 증가로 인해 운동 능력을 증가시키는 것일 수 있다. According to one embodiment, the strengthening of muscle function may indicate muscle fiber synthesis, muscle differentiation, reduction in muscle fatigue, increase in muscle mass, and increase in exercise capacity due to increase in muscle mass.
일 실시예에 따르면, 노화로 감소된 악력 및 트레이드밀 운동량이 락토바실러스 플란타럼 HY7715를 섭취하였을 때 유지 또는 증가되는 것을 확인하였으며, 근육의 피로도를 감소시키는 것을 확인하였다. 또한, 근섬유가 합성되는 것을 확인하였으며, 근육 분화 전사인자의 발현 증가 및 근감소증 원인인자의 발현 감소를 확인하였다. According to one embodiment, it was confirmed that the grip strength and trade mill exercise reduced with aging were maintained or increased when Lactobacillus plantarum HY7715 was ingested, and it was confirmed that the muscle fatigue was reduced. In addition, it was confirmed that the muscle fibers were synthesized, and it was confirmed that the expression of the transcription factor for muscle differentiation was increased and the expression of the factor causing sarcopenia was decreased.
본 명세서에서 용어 '강화'는 힘을 더 강하고 튼튼하게 하는 것을 일컫는 것으로, 상기 근기능 강화는 근육의 합성을 증가시키거나, 근육의 감소를 억제하는 것을 의미하는 것일 수 있다. As used herein, the term 'strengthening' refers to making the force stronger and stronger, and the strengthening of the muscle function may mean increasing the synthesis of a muscle or inhibiting a decrease in the muscle.
일 구체예에 따르면, 상기 식품 조성물은 발효유, 기능성음료, 건강기능식품 중에서 어느 하나일 수 있으나, 이에 제한되는 것은 아니다.According to one embodiment, the food composition may be any one of fermented milk, functional beverage, and health functional food, but is not limited thereto.
본 발명에 따른 락토바실러스 플란타럼(Lactobacillus plantarum) HY7715을 포함하는 조성물은 근기능을 강화하는 효과가 있어 근육질환 예방 또는 개선용 조성물 또는 근기능 강화용 조성물로 활용할 수 있다.The composition comprising Lactobacillus plantarum HY7715 according to the present invention has an effect of strengthening muscle function, so it can be used as a composition for preventing or improving muscle disease or a composition for strengthening muscle function.
도 1는 각 그룹의 평균값으로 A 평균 몸무게 변화를, B는 평균 식이섭취량을, C는 평균 음수섭취량을 측정한 데이터이다.
도 2는 각 그룹의 조직 무게를 측정한 데이터로, A는 가자미근의 무게를, B는 비복근의 무게를 측정하여 비교한 데이터이다.
도 3은 1, 3, 5 주차에 각 그룹의 악력 및 근력을 테스트한 데이터로, A는 악력을 측정한 데이터이고, B는 트레이드밀 테스트로 근력을 측정한 데이터이다.
도 4는 근피로도를 측정한 데이터로, A는 1,3,5 주차의 혈중 젖산염 농도를 측정하여 변화를 본 데이터이고, B는 5주뒤 혈장 내 젖산염 농도를, C는 5주뒤 혈장 내 BUN 농도를, D는 5주뒤 혈장 내 크레아틴의 농도를 측정하여 비교한 데이터이다.
도 5는 근감소증 유발 전사인자를 측정한 데이터로, A는 가자미근에서 Atrogin-1, B는 가자미근에서 MuRF1, C는 비복근에서 Atrogin-1, D는 비복근에서 MuRF1 발현 수치를 측정한 데이터이다.
도 6은 각 그룹의 근섬유 합성 및 재생 효과를 현미경으로 확인한 것을 나타내는 데이터이다.
도 7은 각 그룹의 근육분화 전사인자를 측정한 것으로, A는 가자미근에서, B는 비복근에서 MyoD 발현 수치를 측정한 데이터이다.1 is the average value of each group, A is the average weight change, B is the average dietary intake, C is the data of measuring the average negative intake.
2 is data obtained by measuring the tissue weight of each group. A is the weight of the soleus muscle, and B is the data comparing the weight of the gastrocnemius muscle.
3 is data obtained by testing the grip strength and muscle strength of each group at
Figure 4 is data measuring muscle fatigue, where A is the change in blood lactate concentration measured at
FIG. 5 is data obtained by measuring sarcopenia-induced transcription factors. A is Atrogin-1 in the soleus muscle, B is MuRF1 in the soleus muscle, C is Atrogin-1 in the gastrocnemius muscle, and D is the measurement data of the expression level of MuRF1 in the gastrocnemius muscle.
6 is data showing the microscopic confirmation of the muscle fiber synthesis and regeneration effects of each group.
7 is a graph showing the measurement of the transcription factors for muscle differentiation in each group. A is the measurement data of the expression level of MyoD in the soleus muscle and B is the gastrocnemius muscle.
이하 하나 이상의 구체예를 실시예를 통해 보다 상세하게 설명한다. 그러나, 이들 실시예는 하나 이상의 구체예를 예시적으로 설명하기 위한 것으로 본 발명의 범위가 이들 실시예에 한정되는 것은 아니다. Hereinafter, one or more specific embodiments will be described in more detail through examples. However, these examples are for illustrative purposes of one or more embodiments, and the scope of the present invention is not limited to these examples.
실시예 1. 락토바실러스 플란타룸(Example 1. Lactobacillus plantarum ( Lactobacillus plantarumLactobacillus plantarum ) HY7715 동결건조분말 제조) HY7715 freeze-dried powder manufacturing
락토바실러스 플란타룸(Lactobacillus plantarum) HY7715를 식용 배지에서 배양하여 얻은 배양액을 8,000rpm에서 15분동안 원심분리하여 프로바이오틱스 농축액을 얻었다. 프로바이오틱스 농축액과 코팅제 및 동결보호제로써 가압 살균된 10 중량%의 탈지분유가 함유된 수용액을 1:1의 중량비율로 혼합한 다음, 영하 70℃에서 6시간동안 동결하여 동결 건조분말을 제조하였다. Lactobacillus plantarum ( Lactobacillus plantarum ) The culture solution obtained by culturing HY7715 in an edible medium was centrifuged at 8,000 rpm for 15 minutes to obtain a probiotic concentrate. A freeze-dried powder was prepared by mixing a probiotic concentrate and an aqueous solution containing 10 wt% of skim milk powder sterilized under pressure as a coating agent and cryoprotectant in a weight ratio of 1:1, and then freezing at −70° C. for 6 hours.
실시예 2.Example 2. 락토바실러스 플란타룸(Lactobacillus plantarum ( Lactobacillus plantarumLactobacillus plantarum ) HY7715를 포함하는 발효유 제조) manufacture of fermented milk containing HY7715
유산균 배양액은 원유 95.36중량%와 탈지분유 4.6중량%를 교반하여 15℃에서 비중은 1.0473~1.0475, 적정산도는 0.200~0.220%, pH는 6.55~6.70, 20℃에서의 브릭스(BrixO)는 16.3~16.5% 정도가 되도록 혼합하였다. 혼합 후에 이를 UHT 열처리(135℃에서 2초간 살균)하고 적정온도로 냉각한 뒤, 스트렙토코커스 써모필러스균과 유당분해효소(Valley laboratory, USA)를 각기 0.02 중량%씩 첨가하고 6시간 동안 배양하여 BCP 배지에서의 총 유산균 수가 1.0 X 109 cfu/ml이상, 적정산도가 0.89~0.91%, pH는 4.55~4.65가 되도록 하여 제조하였다. The lactic acid bacteria culture medium is stirred with 95.36% by weight of crude oil and 4.6% by weight of skim milk powder. At 15℃, the specific gravity is 1.0473~1.0475, the titratable acidity is 0.200~0.220%, the pH is 6.55~6.70, and the BrixO at 20℃ is 16.3~ It was mixed so that it might become about 16.5%. After mixing, UHT heat treatment (sterilization at 135°C for 2 seconds), cooling to an appropriate temperature, Streptococcus thermophilus and lactose degrading enzyme (Valley laboratory, USA) were added in an amount of 0.02 wt% each, and cultured for 6 hours for BCP It was prepared so that the total number of lactic acid bacteria in the medium was 1.0
혼합과즙시럽은 액상과당 13중량%, 백설탕 5중량%, 혼합과즙농축액 56Brix0 10.9중량%, 펙틴 1.0 중량%, 후레쉬 후르츠 믹스 에센스 0.1 중량% 및 정제수 70 중량%를 35℃에서 교반하여 혼합한 후 UHT 열처리(135℃에서 2초간 살균)한 후 냉각하여 제조하였다. Mixed juice syrup is UHT after mixing 13% by weight of fructose liquid, 5% by weight of white sugar, 10.9% by weight of mixed juice concentrate 56Brix0, 1.0% by weight of pectin, 0.1% by weight of fresh fruit mix essence and 70% by weight of purified water at 35°C. After heat treatment (sterilization at 135° C. for 2 seconds), it was prepared by cooling.
상기 유산균 배양액 69.5 중량%와 상기 실시예 1의 락토바실러스 플란타룸 HY7715 동결건조분말 0.1 중량% 및 상기 혼합과즙시럽 30.4 중량%를 조합하여 150bar에서 균질한 후 10℃ 이하로 냉각하여 본 발명의 락토바실러스 플란타룸(Lactobacillus plantarum) HY7715를 유효성분으로 함유하는 발효유를 제조하였다. 69.5 wt% of the lactic acid bacteria culture medium, 0.1 wt% of the Lactobacillus plantarum HY7715 freeze-dried powder of Example 1, and 30.4 wt% of the mixed fruit syrup were combined, homogenized at 150 bar, and cooled to 10 ° C. or less to the lactobacilli of the present invention Fermented milk containing Bacillus plantarum HY7715 as an active ingredient was prepared.
실시예 3. 락토바실러스 플란타룸(Example 3. Lactobacillus plantarum ( Lactobacillus plantarumLactobacillus plantarum ) HY7715를 유효성분으로 함유하는 기능성 음료의 제조) Manufacture of functional beverage containing HY7715 as an active ingredient
혼합과즙시럽은 액상과당 13중량%, 백설탕 2.5중량%, 갈색설탕 2.5중량%, 혼합과즙농축액 56Brix0 10.9중량%, 펙틴 1.0중량%, 후레쉬후르츠 믹스 에센스 0.1중량% 및 정제수 70중량%를 35℃에서 교반하여 혼합한 후 UHT열처리(135℃에서 2초간 살균)한 후 냉각하여 제조하였다. 그리고, 상기의 방법으로 제조된 혼합과즙시럽 30.4중량%와 상기 실시예 1의 락토바실러스 플란타룸(Lactobacillus plantarum) HY7715 동결건조분말 0.1중량% 및 나머지 정제수 69.5중량% 조합하여 150bar에서 균질한 후 10℃이하로 냉각한 후 이를 유리병, 페트병 등 소포장 용기에 포장하여 본 발명의 락토바실러스 플란타룸(Lactobacillus plantarum) HY7715를 유효성분으로 함유하는 기능성 음료를 제조하였다.Mixed juice syrup is prepared by mixing 13% by weight of fructose liquid, 2.5% by weight of white sugar, 2.5% by weight of brown sugar, 10.9% by weight of mixed juice concentrate 56Brix0, 1.0% by weight of pectin, 0.1% by weight of fresh fruit mix essence, and 70% by weight of purified water at 35°C. After mixing by stirring, it was prepared by UHT heat treatment (sterilization at 135° C. for 2 seconds) and then cooling. Then, 30.4% by weight of the mixed juice syrup prepared by the above method, 0.1% by weight of the Lactobacillus plantarum HY7715 freeze-dried powder of Example 1, and 69.5% by weight of the remaining purified water were combined and homogenized at 150
실시예 4. 락토바실러스 플란타룸(Lactobacillus plantarum) HY7715를 유효성분으로 함유하는 건강기능식품의 제조Example 4. Preparation of health functional food containing Lactobacillus plantarum HY7715 as an active ingredient
상기 실시예 1의 락토바실러스 플란타룸(Lactobacillus plantarum) HY7715 동결건조분말 0.1중량%에 영양보조성분(비타민 B1, B2, B5, B6, E 및 초산에스테르, 니코틴산 아미드) 및 올리고당을 상기의 실시예 1의 락토바실러스 플란타룸(Lactobacillus plantarum) HY7715 동결건조분말 100중량부에 대하여 10중량부가 되도록 첨가하여 고속회전 혼합기에서 혼합하였다. 상기 혼합물에 멸균 정제수 10중량부를 첨가, 혼합하고 직경 1~2mm의 과립상으로 성형하였다. 상기 성형된 과립은 50
실험예 1. 실험 마우스 준비 Experimental Example 1. Preparation of experimental mice
동물실험윤리위원회(IACUC) 승인(승인번호: AEC-2019-00012-Y)을 받아 Balb/c 수컷 마우스 6주령 8마리와 80주령 24마리를 가지고 실험하였다. 마우스는 일주일간 21~25℃, 습도 30~50%의 조건에서 순화시켰다. 그 후, 마우스는 8마리씩 하기 표 1과 같이 그룹으로 나누어 총 5주 동안 같은 양의 시료를 경구투여 하였다. 모든 그룹은 일반식이를 하였다. With the approval of the Animal Experimental Ethics Committee (IACUC) (approval number: AEC-2019-00012-Y), the experiment was conducted with 8 6-week-old Balb/c male mice and 24 80-week-old mice. Mice were acclimatized in conditions of 21 to 25 ° C and 30 to 50% humidity for one week. Thereafter, the mice were divided into groups of 8 mice each as shown in Table 1 below, and the same amount of the sample was orally administered for a total of 5 weeks. All groups ate a regular diet.
노화에 따른 변화를 확인하기 위하여, YM 그룹 및 OM 그룹은 일반식이 외에 따로 시료를 투여하지 않았다. 양성 대조군인 OM+C 그룹은 크레아틴을 75mg/kg/day를 기준으로, OM+HY 그룹은 락토바실러스 플란타럼(Lactobacillus plantarum) HY7715를 1x109 CFU/kg/day를 기준으로 정수에 녹여 24시간 간격으로 경구투여하였다. In order to confirm the change according to aging, the YM group and the OM group did not administer the sample other than the regular diet. The positive control group, OM+C group, dissolved creatine at 75mg/kg/day, and OM+HY group dissolved Lactobacillus plantarum HY7715 in purified water based on 1x10 9 CFU/kg/day for 24 hours. It was administered orally at intervals.
실험예 2. 체중 및 근육량 변화Experimental Example 2. Changes in body weight and muscle mass
시료 투여에 따라 몸무게 및 근육량의 변화가 있는지 확인하기 위하여, 식이섭취량, 음수섭취량, 체중 및 근육량을 확인하였다. 식이섭취량 및 음수섭취량은 각 그룹별로 측정하여 평균값을 나타내었다. 체중은 1주일 간격으로 동일한 시간에 측정하였다. 근육량은 5주간의 시료 투여를 마치고 근력 및 운동 능력 테스트를 마친 뒤, 각 그룹 마우스의 가자미근(soleus)과 비복근(gastrocnemius)를 적출하여 무게를 측정하였다. In order to check whether there is a change in body weight and muscle mass according to sample administration, food intake, drinking water intake, body weight and muscle mass were checked. Dietary intake and drinking water intake were measured for each group, and average values were shown. Body weights were measured at the same time at intervals of one week. For muscle mass, after 5 weeks of sample administration and muscle strength and exercise ability tests, the soleus and gastrocnemius of each group of mice were excised and their weight was measured.
그 결과, 도 1A에서 보이는 바와 같이, YM 그룹은 23.54g으로 시작하여 5주차에 26.34g으로 측정되어 성장하면서 무게가 증가한 것을 확인하였다. OM 그룹, OM+C 그룹 및 OM+HY 그룹은 1주차부터 5주차까지 무게가 유의한 차이 없이 유지되는 것을 확인하였다.As a result, as shown in FIG. 1A, the YM group started with 23.54 g and was measured to be 26.34 g at the 5th week, confirming that the weight increased while growing. It was confirmed that the OM group, OM+C group and OM+HY group maintained their weight without any significant difference from
도 1B 및 도 1C에서 보이는 바와 같이, 모든 그룹이 식이섭취량은 차이가 없었으며, 노령인 OM, OM+C 및 OM+HY 그룹에서 어린 YM 그룹보다 음수섭취량은 높았으나, 노령의 마우스들이 속한 그룹끼리는 유의한 차이가 없었다.As shown in FIGS. 1B and 1C , there was no difference in dietary intake for all groups, and the negative intake was higher in the elderly OM, OM+C and OM+HY groups than in the young YM group, but the group to which the elderly mice belonged. There was no significant difference between the two.
도 2A에서 보이는 바와 같이, OM 그룹의 마우스는 노화에 의하여 가자미근의 근육량이 YM 그룹 대비 현저히 감소한 것을 확인하였다. 반면, 크레아틴을 섭취한 OM+C 그룹과 락토바실러스 플란타럼 HY7715를 섭취한 OM+HY 그룹의 가자미근은 OM 그룹 대비 현저히 증가된 상태였으며, OM+HY 그룹의 가자미근의 무게는 YM 그룹의 가자미근 무게와 유사하였다. As shown in FIG. 2A , it was confirmed that the muscle mass of the soleus muscle significantly decreased in the mice of the OM group compared to the YM group due to aging. On the other hand, the soleus muscle of the OM+C group taking creatine and the OM+HY group taking Lactobacillus plantarum HY7715 was significantly increased compared to the OM group. was similar to
또한 도 2B에서 보이는 바와 같이, 마우스의 비복근 조직의 무게의 경우, OM 그룹에서만 근육량이 감소되었고, OM+C 그룹 및 OM+HY 그룹은 YM 그룹과 유사할 정도로 비복근 조직량이 유지되는 것을 확인하였다.Also, as shown in FIG. 2B , in the case of the mouse gastrocnemius tissue weight, it was confirmed that the muscle mass was reduced only in the OM group, and the gastrocnemius tissue mass was maintained similar to that of the YM group in the OM+C group and the OM+HY group.
실험예 3. 근력 및 운동 능력 테스트 Experimental Example 3. Strength and exercise ability test
근력을 측정하기 위하여, 시료 투여기간 동안 1, 3, 5주차 마지막 날에 마우스 앞발과 뒷발 모두를 이용하여 all limb grip strength test를 측정하였다. 마우스가 악력 시험 장치를 잡게 한 후 마우스의 꼬리를 잡고 뒤로 잡아당길 때 나타나는 앞발과 뒷발의 최대 악력을 측정하였다. 악력은 마우스 당 3회 반복 측정한 후 최대값의 평균값을 사용하였고, 측정 힘의 단위는 kgf이며, 체중 대비 악력(kgf/kg)으로 계산하여 표기하였다.To measure muscle strength, the all-limb grip strength test was measured using both the front and hind paws of mice on the last day of
그 결과 도 3A에서 보이는 바와 같이, 1주차에서는 OM 그룹과 OM+HY 그룹이 큰 차이가 나지 않았으나, 5주차에는 OM 그룹은 5.27kgf/kg인 반면, OM+C 그룹은 5.97kgf/kg, OM+HY 그룹은 6.42kgf/kg으로 유의적으로 증가한 것을 확인할 수 있었다.As a result, as shown in FIG. 3A , there was no significant difference between the OM group and the OM+HY group in the 1st week, but the OM group was 5.27kgf/kg at the 5th week, whereas the OM+C group was 5.97kgf/kg, the OM It was confirmed that the +HY group significantly increased to 6.42 kgf/kg.
근력 강화로 인한 운동 능력을 테스트하기 위하여, 식이 기간 동안 1, 3, 5주차 마지막 날에 마우스에서 탈진 트레이드밀 운동을 실시하였다. 트레이드밀(정도비엔피, JD-A-09)을 이용하여, 0°의 경사도에서 5m/min의 속도로 10분, 10m/min 속도로 10분, 15m/min 속도로 10분, 20m/min 속도로 10분, 25m/min 속도로 10분 운동을 시켰다. In order to test the exercise ability due to muscle strengthening, exhaustion trademill exercise was performed in mice on the last day of 1st, 3rd, and 5th weeks during the feeding period. Using a trade mill (Jodo BNP, JD-A-09), at 0° inclination, at a speed of 5 m/min for 10 minutes, at a speed of 10 m/min for 10 minutes, at a speed of 15 m/min for 10 minutes, at a speed of 20 m/min. The exercise was performed for 10 minutes at a speed of 25 m/min and for 10 minutes at a speed of 25 m/min.
도 3B에서 보이는 바와 같이, YM 그룹의 경우, 모든 마우스가 50분의 운동을 종료하고 최대 운동거리인 750m를 기록하였다. 반면, OM 그룹은 YM 그룹 대비 현저히 감소한 거리를 기록한 것을 확인하였다. OM+HY 그룹은 시료 투여 후 3주차가 지나면서 OM 그룹 대비 현저히 증가된 거리를 기록하였으며, 5주차엔 OM 그룹 및 OM+C 그룹보다 현저히 증가된 거리를 기록하는 것을 확인하였다. As shown in FIG. 3B , in the YM group, all mice completed 50 minutes of exercise and recorded the maximum exercise distance of 750 m. On the other hand, it was confirmed that the OM group recorded a significantly reduced distance compared to the YM group. It was confirmed that the OM+HY group recorded a significantly increased distance compared to the OM group after 3 weeks after sample administration, and recorded a significantly increased distance compared to the OM group and OM+C group at the 5th week.
따라서 악력 및 트레이드밀 테스트를 통해 락토바실러스 플란타럼 HY7715를 섭취한 마우스 그룹이 다른 노령 마우스 그룹 대비 근력 및 운동능력이 향상됨을 확인하였다.Therefore, it was confirmed that the mouse group ingesting Lactobacillus plantarum HY7715 improved muscle strength and athletic ability compared to other elderly mouse groups through grip strength and trademill tests.
실험예 4. 근육 피로도 확인Experimental Example 4. Confirmation of muscle fatigue
실험예 4-1. 혈중 젖산염(Lactate) 농도 측정Experimental Example 4-1. Measurement of blood lactate concentration
상기 실험예 3의 트레이드밀 운동 직후, 근육의 피로도를 확인하기 위하여 꼬리 정맥에서 혈액을 소량 채취하였다. 각 마우스의 혈액은 lactate meter(Lactate Pro 2, Cosmed)를 이용하여 혈중 젖산염(lactate) 농도를 측정하여 변화를 확인하였다. Immediately after the trademill exercise of Experimental Example 3, a small amount of blood was collected from the tail vein in order to check the degree of muscle fatigue. Changes were confirmed by measuring the blood lactate concentration in the blood of each mouse using a lactate meter (
도 4A에서 보이는 바와 같이, YM 그룹은 노화 마우스를 사용한 다른 그룹에 비하여 운동 직후 젖산염의 농도가 낮았다. 반면, 노령의 마우스를 사용한 나머지 OM 그룹, OM+C 그룹, OM+HY 그룹은 1주차에는 비슷한 정도의 젖산염이 축적되었다. 그러나 OM 그룹은 5주차가 지났을 때 젖산염이 더 축적되었으나, OM+C 그룹 및 OM+HY 그룹은 3주차까지 증가하다가 다시 감소하여, OM 그룹 대비 젖산염 농도가 낮은 것을 확인하였다. As shown in FIG. 4A , the YM group had a lower concentration of lactate immediately after exercise than the other groups using aged mice. On the other hand, the remaining OM group, OM+C group, and OM+HY group using aged mice accumulated lactate in a similar degree at the 1st week. However, the OM group accumulated more lactate after 5 weeks, but the OM+C group and OM+HY group increased until the 3rd week and then decreased again, confirming that the lactate concentration was lower than that of the OM group.
실험예 4-2. 혈장 내 젖산염, BUN(Blood urea nitrogen) 및 크레아틴을 측정Experimental Example 4-2. Measuring lactate, blood urea nitrogen (BUN) and creatine in plasma
상기 실험예 3의 5주차 트레이드밀 운동 직후, 각 실험 그룹의 복대동맥에서 혈액을 채취하고, 혈액을 4℃, 4000rpm에서 10분간 원심분리하여 혈장(plasma)를 분리하였다. 혈액생화학분석기(7072, Hitachi, Japan)을 이용하여 젖산염, BUN(Blood urea nitrogen) 및 크레아틴을 측정하였다.Immediately after the 5th week trade mill exercise of Experimental Example 3, blood was collected from the abdominal aorta of each experimental group, and the blood was centrifuged at 4° C. and 4000 rpm for 10 minutes to separate plasma. Lactate, blood urea nitrogen (BUN) and creatine were measured using a blood biochemical analyzer (7072, Hitachi, Japan).
도 4B에서 보이는 바와 같이, OM 그룹 대비 OM+C 및 OM+HY의 혈장 내 젖산염의 농도가 감소된 것을 확인하였다. 이를 통해 OM+C 그룹 및 OM+HY 그룹이 탈진의 정도가 일반 정수를 시료로 섭취한 OM 그룹보다 적다는 것을 확인하였다.As shown in FIG. 4B , it was confirmed that the concentration of lactate in plasma of OM+C and OM+HY was reduced compared to the OM group. Through this, it was confirmed that the degree of exhaustion in the OM+C group and the OM+HY group was lower than that in the OM group, which consumed normal purified water as a sample.
BUN(Blood urea nitrogen)은 체내 단백질대사의 최종 생산물로, 단백질이나 아미노산 이화작용 및 피로가 유발되었음을 알려주는 지표이다. 도 4C에서 보이는 바와 같이, 혈장 내 BUN 농도는 OM 그룹을 제외하고는 모두 YM 그룹만큼 감소된 결과를 확인할 수 있었다. BUN (blood urea nitrogen) is the final product of protein metabolism in the body, and is an indicator indicating that protein or amino acid catabolism and fatigue are induced. As shown in FIG. 4C , it was confirmed that the plasma BUN concentration was reduced by the YM group except for the OM group.
도 4D에서 보이는 바와 같이, 혈장 내 크레아틴은 노령의 마우스를 사용한 OM 그룹대비 어린 마우스를 사용한 YM 그룹이 낮은 것을 확인하였으며, OM+C 그룹 및 OM+HY 그룹 모두 OM 그룹 대비 혈장 내 크레아틴이 감소한 것을 확인하였다. As shown in FIG. 4D , it was confirmed that the YM group using young mice was lower in plasma creatine compared to the OM group using old mice, and both the OM+C group and the OM+HY group showed a decrease in plasma creatine compared to the OM group. Confirmed.
이를 통해 락토바실러스 플란타럼 HY7715 섭취가 단백질 이화작용 억제하고, 운동 후 탈진의 정도를 감소시켜 근육 피로도를 감소시키는 것을 확인하였다. Through this, it was confirmed that Lactobacillus plantarum HY7715 intake suppressed protein catabolism and decreased muscle fatigue by reducing the degree of exhaustion after exercise.
실험예 5. 근감소증 유발 전사인자 유전자 수준 측정Experimental Example 5. Sarcopenia-induced transcription factor gene level measurement
5주간 투여 및 실험 후, 각 실험 그룹 마우스의 가자미근(soleus)과 비복근(gastrocnemius)의 근감소증 유발 전사인자 유전자 수준을 q-PCR을 통해 비교하였다. 근감소증 유발 전사인자로는 단백질 분해 인자인 muscle-specific F-box protein(Atrogin-1)과 muscle RING-finger protein-1(MuRF1)을 확인하였다. After 5 weeks of administration and experiment, the gene levels of sarcopenia-induced transcription factors in the soleus and gastrocnemius of mice of each experimental group were compared through q-PCR. As transcription factors for sarcopenia, muscle-specific F-box protein (Atrogin-1) and muscle RING-finger protein-1 (MuRF1), which are proteolytic factors, were identified.
그 결과 도 5A 및 도 5B에서 보이는 바와 같이, 가자미근에서는 OM+HY 그룹이 YM 그룹과 유사할 정도로 Atrogin-1의 mRNA 발현 수준이 감소하였으며, MuRF1은 YM 그룹 및 OM+C 그룹보다 더 낮은 수준으로 발현된다는 것을 확인하였다. As a result, as shown in FIGS. 5A and 5B, in the soleus muscle, the mRNA expression level of Atrogin-1 was reduced in the OM+HY group to a degree similar to that of the YM group, and MuRF1 was lower than that of the YM group and OM+C group. was confirmed to be expressed.
또한 도 5C 및 도 5D에서 보이는 바와 같이, 비복근에서는 OM+HY 그룹이 YM 그룹을 포함한 다른 그룹대비 Atrogin-1 및 MuRF1 발현이 현저히 감소하였다는 것을 확인하였다. In addition, as shown in FIGS. 5C and 5D , it was confirmed that the expression of Atrogin-1 and MuRF1 was significantly reduced in the OM+HY group compared to other groups including the YM group in the gastrocnemius muscle.
이를 통해, 락토바실러스 플란타럼의 섭취가 근감소증 유발 인자의 유전자 발현을 감소 시키므로 근감소증을 억제한다는 것을 확인하였다. Through this, it was confirmed that the ingestion of Lactobacillus plantarum suppressed sarcopenia because it reduced gene expression of sarcopenia-inducing factors.
실험예 6. 근섬유 합성 및 재생 효과 확인Experimental Example 6. Confirmation of muscle fiber synthesis and regeneration effect
5주간의 시료투여 및 실험 후, 비복근(Gastrocnemius)을 절취하고 헤마토실린과 이오신(Hematoxylin & Erosin)으로 염색하여 근육 내 근섬유 크기를 광학 현미경으로 관찰하였다. After 5 weeks of sample administration and experiment, the gastrocnemius was cut and stained with hematoxylin & eosin to observe the size of the muscle fibers in the muscle with an optical microscope.
도 6에서 보이는 바와 같이, OM 그룹은 YM 그룹대비 노화가 진행되어 비복근의 근섬유 크기가 현저히 감소하였다. 반면, OM+HY 그룹 및 OM+C 그룹은 OM 그룹 대비 근섬유가 증가된 것을 확인하였다. As shown in FIG. 6 , in the OM group compared to the YM group, aging progressed and the muscle fiber size of gastrocnemius significantly decreased. On the other hand, the OM+HY group and the OM+C group showed increased muscle fibers compared to the OM group.
따라서 락토바실러스 플란타룸 HY7715의 섭취가 근섬유를 합성하는 것을 확인하였다. Therefore, it was confirmed that the intake of Lactobacillus plantarum HY7715 synthesized muscle fibers.
실험예 7. 근육분화(myogenesis) 전사인자 유전자 수준 측정Experimental Example 7. Measurement of gene levels of myogenesis transcription factors
5주간 투여 및 실험 후, 각 실험 그룹 마우스의 가자미근(soleus)과 비복근(gastrocnemius)의 근육분화 전사인자 유전자 수준을 q-PCR을 통해 비교하였다. 근육분화 전사인자로는 일차 근육 생성 전사 인자 중 하나인 Myogenic Differentiation 1(MyoD)를 측정하였다.After administration and experimentation for 5 weeks, the gene levels of the transcription factors for muscle differentiation in the soleus and gastrocnemius of mice of each experimental group were compared through q-PCR. As the myogenic transcription factor, Myogenic Differentiation 1 (MyoD), one of the primary myogenic transcription factors, was measured.
도 7A 및 도 7B에서 보이는 바와 같이, 노화로 인해 가자미근과 비복근에서 YM 그룹 대비 OM 그룹의 MyoD 유전자 발현이 현저히 감소한 것을 확인하였다. 반면, OM+HY 그룹에서는 가자미근에서는 YM 그룹과 유사할 정도로 MyoD 발현이 증가하였으며, 비복근에서도 OM 그룹 및 OM+C 그룹 대비 유의하게 증가된 것을 확인하였다. As shown in FIGS. 7A and 7B , it was confirmed that the MyoD gene expression of the OM group compared to the YM group was significantly reduced in the soleus and gastrocnemius muscles due to aging. On the other hand, in the OM+HY group, the expression of MyoD was increased in the soleus muscle to a degree similar to that of the YM group, and it was confirmed that the gastrocnemius muscle was significantly increased compared to the OM group and the OM+C group.
따라서, 락토바실러스 플란타럼 HY7715의 섭취가 노화로 인한 근육 생성 전사 인자의 감소를 억제하여 근육 생성에 도움이 된다는 것을 확인하였다. Therefore, it was confirmed that the ingestion of Lactobacillus plantarum HY7715 helps to build muscle by suppressing the decrease in the myogenic transcription factor due to aging.
실험예 8. 통계분석Experimental Example 8. Statistical Analysis
모든 생화확적 데이터는 평균 ± 표준 편차로 나타내었다. 각 실험 군 간의 차이는 단일 요인 분석(ANOVA, single-factor analysis of variance)으로 분석하였다. 각 실험 군 간 의 평균값의 차이에 대한 유의성은 Tukey의 다중 비교 검정(Tukey's multiple comparison tests)을 이용하여 p<0.05 수준에서 평가하였다. (a>b>c)All biochemical data are presented as mean ± standard deviation. Differences between each experimental group were analyzed by single-factor analysis of variance (ANOVA). The significance of the difference in the mean value between each experimental group was evaluated at p<0.05 level using Tukey's multiple comparison tests. (a>b>c)
Claims (5)
Lactobacillus plantarum ( Lactobacillus plantarum ) Food composition for preventing or improving muscle disease comprising HY7715.
The food composition according to claim 1, wherein the muscle disease is decreased muscle function due to aging, muscle wasting, and muscle degeneration.
The food composition according to claim 1, wherein the food composition is any one of fermented milk, functional beverage, and health functional food.
Lactobacillus plantarum ( Lactobacillus plantarum ) Food composition for strengthening muscle function comprising HY7715.
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