KR102036864B1 - Hydrolysate of silkworm raised with cudrania tricuspidata leaf, having improved gaba and rutin-enhancing activity, antioxidant activity or inhibitory activity, and preparation method thereof - Google Patents
Hydrolysate of silkworm raised with cudrania tricuspidata leaf, having improved gaba and rutin-enhancing activity, antioxidant activity or inhibitory activity, and preparation method thereof Download PDFInfo
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- KR102036864B1 KR102036864B1 KR1020180004393A KR20180004393A KR102036864B1 KR 102036864 B1 KR102036864 B1 KR 102036864B1 KR 1020180004393 A KR1020180004393 A KR 1020180004393A KR 20180004393 A KR20180004393 A KR 20180004393A KR 102036864 B1 KR102036864 B1 KR 102036864B1
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- silkworm
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- mulberry leaves
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- hydrolyzate
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- 238000001694 spray drying Methods 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- HXJUTPCZVOIRIF-UHFFFAOYSA-N sulfolane Chemical compound O=S1(=O)CCCC1 HXJUTPCZVOIRIF-UHFFFAOYSA-N 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 230000005062 synaptic transmission Effects 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 239000003760 tallow Substances 0.000 description 1
- 235000021195 test diet Nutrition 0.000 description 1
- JZRWCGZRTZMZEH-UHFFFAOYSA-N thiamine Chemical compound CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N JZRWCGZRTZMZEH-UHFFFAOYSA-N 0.000 description 1
- UFTFJSFQGQCHQW-UHFFFAOYSA-N triformin Chemical compound O=COCC(OC=O)COC=O UFTFJSFQGQCHQW-UHFFFAOYSA-N 0.000 description 1
- 235000019154 vitamin C Nutrition 0.000 description 1
- 239000011718 vitamin C Substances 0.000 description 1
- 230000002087 whitening effect Effects 0.000 description 1
- 230000037330 wrinkle prevention Effects 0.000 description 1
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Abstract
본 발명은 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성이 증대된, 꾸지뽕잎으로 사육한 숙잠누에 가수분해물, 및 이의 제조방법에 관한 것으로, 보다 상세하게는 씨누에에서 3 내지 4령까지는 뽕잎으로 사육된 후, 숙잠기까지 꾸지뽕잎으로 사육된 누에를 가수분해함으로써, 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성을 증대시키는 효과는 갖는다.The present invention relates to a hydrolyzate of Suksam silkworms bred with Cudrania leaf, which has increased Gaba and rutin enhancement activity, antioxidant activity or acetylcholinesterase inhibitory activity, and more specifically, 3 to 3 in seedlings. Up to 4 years of age, after raising the mulberry leaves, and by the hydrolysis of silkworms bred with kkujippong leaves until the slumber, has the effect of increasing the Gaba and rutin enhancement activity, antioxidant activity or acetylcholinesterase inhibitory activity.
Description
본 발명은 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성이 증대된, 꾸지뽕잎으로 사육한 누에 가수분해물, 및 이의 제조방법에 관한 것으로, 보다 상세하게는 씨누에에서 3 내지 4령까지는 뽕잎으로 사육된 후, 숙잠기까지 꾸지뽕잎으로 사육된 숙잠누에를 감압진공분해 장치를 통해 가수분해하여, 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성을 증대시키는 기술에 관한 것이다.
The present invention relates to a silkworm hydrolyzate bred with Cudrania leaf, which has increased Gaba and rutin enhancement activity, antioxidant activity or acetylcholinesterase inhibitory activity, and a method for preparing the same, more specifically, in silkworms 3 to 4 Sukjam silkworms were bred with mulberry leaves until the age of aging, and then hydrolyzed through the vacuum decompression device to squeeze the mulberry leaves to increase their gaba and rutin enhancement activity, antioxidant activity or acetylcholinesterase inhibitory activity. It is about.
누에는 단백질, 생리활성 펩타이드, 아미노산, 미네랄, deoxynojirimycin, 섬유질과 같은 생리활성 영양적 가치가 풍부한 성분을 함유하며, 혈당개선, 혈압개선, 간기능 개선, 뇌기능 개선, 혈액순환 촉진 및 미백기능 등의 생리적 활성 작용으로 인행 건강기능식품소재로 주목 받고 있다. 또한 일명 누에 혈림프라고도 부르는 체액은 세포 수명을 연장시키는 효과가 있음이 보고되었다. 또한 누에 고치(Bombyx mori)에서 나온 실크단백질은 성질이 상이한 섬유상 단백질인 세리신(sericin)과 피브로인(fibroin)이 각각 25 %와 75 %로 구성되어 있다. 실크단백질은 항산화, tyrosinase 활성 저해, 항당뇨, 활성산소 생성 억제 및 산화적 스트레스 예방, 피부 보습, 주름방지, 기억력 개선 등의 여러 활성에 대한 연구가 있었으며, 실크단백질을 펩타이드화 하여 다양한 분야에서 기능성식품, 화장품, 미용 비누 및 의료용 소재 등에 응용되고 있다.Silkworm contains ingredients rich in physiologically active nutritional value, such as proteins, bioactive peptides, amino acids, minerals, deoxynojirimycin, and fiber, improving blood sugar, improving blood pressure, improving liver function, improving brain function, promoting blood circulation and whitening Due to its physiological activity, it is attracting attention as a nutritional supplement. Also known as silkworm hemolymph has been reported to have an effect of prolonging cell life. Silk proteins from the cocoon (Bombyx mori) are composed of 25% and 75% of sericin and fibroin, respectively, of different fibrous proteins. Silk proteins have been studied for various activities such as antioxidant, inhibition of tyrosinase activity, anti-diabetic, free radical production and prevention of oxidative stress, skin moisturization, wrinkle prevention, memory improvement, etc. It is applied to food, cosmetics, beauty soap and medical materials.
한편, 누에 체내의 견사선(실샘)은 5령4일이 되면 누에 체내에서 급속하게 비대해져 5령5일에 이르러 최대로 발달하는 것으로 알려져 있다. 최대의 견사선(실샘)을 가지는 누에를 생산하기 위해서는 정확한 날짜에 한번에 대량의 누에를 수거해서 처리해야 한다. 그러나 사육량이 많은 대규모 농가의 경우에는 순간적으로 일시에 노동력이 집중적으로 투여돼야 하기 때문에 정확한 누에 수거시기를 놓치는 경우가 많고, 일부 농가에서는 생산량을 늘리기 위하여 뽕밭에 뽕잎이 남을 경우 추가로 급상하여 제품의 품질을 떨어뜨리는 등 현실적으로 많은 어려움이 발생되고 있다.On the other hand, silk glands (silual glands) in the silkworm body is 5 years old, it is known that it rapidly enlarges in the silkworm body and reaches the maximum of 5 age 5 days. To produce silkworms with the largest silk glands, a large amount of silkworms must be collected and processed at once on the exact date. However, in the case of large farms with a large number of farming farms, the labor force must be concentrated at the moment, so it is often missed the exact time for collecting silkworms.In some farms, the mulberry leaves remain in the mulberry field in order to increase the yield of products. There are many difficulties in reality, such as deterioration in quality.
또한, 누에는 급여하는 잎에 따라 그 성분과 기능성이 상이한데 일반 뽕잎에 비해 꾸지뽕잎은 단백질, 식이섬유, 비타민 B1, 비타민 B2, 가바(GABA; gamma-aminobutyric acid), 루틴(rutin), 플라보노이드 등의 기능성 성분과 항산화 성분 등이 풍부하게 함유되어 있다. 이 중 특히 뇌 혈류개선, 산소공급증가, 뇌세포의 대사기능을 촉진시켜 기억력 증진, 신경안정 및 불안장애 개선, 우울증 완화, 수면유도 및 조절, 혈압강하, 중풍과 치매 예방, 비만 개선, 갱년기 장애 개선 등에 효과적으로 뇌건강 식품소재로써의 잠재성이 매우 기대되는 가바가 일반뽕에 비하여 약 5 배 이상 함유되어 있다. 또한 플라본(flavon) 유도체의 하나인 루틴은 혈관을 강화하고 혈압 강하하여 출혈성 질병을 예방하고, 이외에도 항종양, 항세균, 항암 활성 등의 효과가 보고되어 있으며 일반뽕의 18 배 이상 함유되어 있는 것으로 보고되어 있다. In addition, the components and functionalities of silkworms are different depending on the leaves to be fed. Compared with the common mulberry leaves, Cudrania leaves are composed of protein, dietary fiber, vitamin B 1 , vitamin B 2 , gaba- gamma-aminobutyric acid (GABA), and rutin. And functional ingredients such as flavonoids and antioxidant components. In particular, it improves brain blood flow, increases oxygen supply, promotes metabolic function of brain cells, improves memory, improves nervous and anxiety disorders, reduces depression, induces and controls sleep, lowers blood pressure, prevents stroke and dementia, improves obesity, and menopausal disorders. Gaba, which is expected to be effective as a brain health food material, contains about five times more than ordinary mulberry. In addition, rutin, one of the flavon derivatives, strengthens blood vessels and lowers blood pressure to prevent hemorrhagic diseases. In addition, antitumor, antibacterial and anticancer activity has been reported, and it contains 18 times more than regular mulberry. Reported.
이와 관련하여, 한국 공개특허 공보 제10-2011-0044085호에는 누에를 5령기 때까지 사육하여 익은누에를 급속동결한 후 동결건조 및 실샘 분리하는 단계를 수행하여 실샘 분리 및 실샘 분말을 제조하는 방법이 개시되어 있고, 대한잠사학회지(2015)에는 식용섭취를 위한 숙잠의 가공기술이 개시되어 있으나, 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성 증진에는 한계가 있었으며, 급속동결, 동결건조 및 실샘분리 등이 수행되어야 하므로 공정이 복잡한 단점이 존재하였다.In this regard, Korean Patent Laid-Open Publication No. 10-2011-0044085 discloses a method for producing silk gland separation and silk gland powder by breeding silkworms until the 5th age and then rapidly freezing the ripe silkworms, followed by lyophilization and separating the gland. It is disclosed, the Journal of the Korean Society of Potassium (2015) discloses the processing technology of slumber for edible intake, but there was a limit to the enhancement of Gaba and rutin enhancement activity, antioxidant activity or acetylcholinesterase inhibitory activity, rapid freezing, Freeze-drying and siliceous separation have to be carried out, so there was a complicated process.
따라서, 본 발명자는 씨누에에서 3 내지 4령까지는 뽕잎으로 사육된 후, 숙잠기까지 꾸지뽕잎으로 사육된 숙잠누에를 감압진공분해 장치를 통해 가수분해하여, 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성을 증대시킬 수 있음에 착안하여 본 발명을 완성하기에 이르렀다.
Accordingly, the present inventors hydrolyzed ssam silkworms, which were bred with mulberry leaves from 3 to 4 years old, and then cultivated with courageous mulberry leaves to slumber, using a vacuum decompression vacuum device, to enhance gaba and rutin activity, antioxidant activity or acetyl The present invention has been completed by focusing on the ability to increase cholinesterase inhibitory activity.
본 발명은 상기와 같은 문제점을 고려하여 안출된 것으로, 본 발명의 목적은 씨누에에서 3 내지 4령까지는 뽕잎으로 사육된 후, 숙잠기까지 꾸지뽕잎으로 사육된 숙잠누에를 감압진공분해 장치를 통해 가수분해하여, 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성을 증대시키고자 하는 것이다.
The present invention was conceived in view of the above problems, the object of the present invention was bred in mulberry leaves from 3 to 4 years old in silkworms, sukjam silkworms were bred with cuji mulberry leaves until slumbering through a vacuum decompression device It is intended to increase the Gaba and rutin enhancement activity, antioxidant activity or acetylcholinesterase inhibitory activity by hydrolysis.
상기한 바와 같은 목적을 달성하기 위한 본 발명은, 꾸지뽕잎 및 뽕잎으로 사육한 누에를 가수분해하는 단계를 포함하는 누에 가수분해물의 제조방법을 제공한다.The present invention for achieving the object as described above, provides a method for producing a silkworm hydrolyzate comprising the step of hydrolyzing silkworms bred with kkujippong leaves and mulberry leaves.
상기 누에는 씨누에에서 3 내지 4령까지는 뽕잎으로 사육된 후, 숙잠기까지 꾸지뽕잎으로 사육된 것일 수 있다.The silkworms are bred in mulberry leaves from 3 to 4 years old, and may be bred into cuddle mulberry leaves until sleep.
상기 가수분해는 브로멜린, 파파인, 프로타멕스, 트립신, 알카라아제, 뉴트라아제 및 플라보자임 중에서 선택되는 1종 이상의 단백질 가수분해효소로 효소가수분해 하는 것일 수 있다.The hydrolysis may be to hydrolyze the enzyme with one or more proteolytic enzymes selected from bromelain, papain, protamex, trypsin, alkalase, neutrase and flavozyme.
상기 가수분해효소는 브로멜린, 파파인 및 트립신의 복합효소일 수 있다.The hydrolase may be a complex enzyme of bromelain, papain and trypsin.
상기 가수분해는 30 내지 65 ℃에서 수행될 수 있다.The hydrolysis can be carried out at 30 to 65 ℃.
상기 가수분해는 0.5 내지 10 시간 동안 수행될 수 있다.The hydrolysis may be performed for 0.5 to 10 hours.
상기 가수분해는 감압 조건에서 수행될 수 있다.The hydrolysis can be carried out under reduced pressure conditions.
상기 가수분해는, 상기 가수분해효소를 함유하는 효소액 및 상기 누에을 감압 가수분해 장치에 투입한 후 상기 장치의 상·하측에 연결되는 순환라인을 통하여 상기 효소액을 장치 내부에서 순환시킴으로써 수행될 수 있다.The hydrolysis may be carried out by circulating the enzyme solution containing the hydrolase and the silkworm in a reduced pressure hydrolysis device and then circulating the enzyme solution inside the device through a circulation line connected to the upper and lower sides of the device.
상기 감암 가수분해 장치 내부 효소액의 상기 순환라인에 의한 1 회 순환시간은 30 초 내지 5 분일 수 있다.One cycle time of the circulating line of the enzyme solution inside the gamyeo hydrolysis device may be 30 seconds to 5 minutes.
상기 가수분해 시 산 또는 염을 사용하지 않을 수 있다.
The hydrolysis may not use an acid or a salt.
본 발명에 따르면, 씨누에에서 3 내지 4령까지는 뽕잎으로 사육된 후, 숙잠기까지 꾸지뽕잎으로 사육된 숙잠누에를 감압진공분해 장치를 통해 가수분해하여, 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성을 증대시킬 수 있다.
According to the present invention, the silkworms are bred with mulberry leaves from 3 to 4 years of age, and then hydrolyzed snooze silkworms bred with courageous mulberry leaves until slumbering, using a vacuum decompression device, so that GABA and rutin enhancement activity, antioxidant activity or acetyl It can increase the cholinesterase inhibitory activity.
도 1은 본 발명에 따른 누에 가수분해물의 제조방법을 나타낸 공정도이다.1 is a process chart showing a method for producing a silkworm hydrolyzate according to the present invention.
이하에서, 본 발명의 여러 측면 및 다양하게 구현한 실례에 대해 더욱 구체적으로 설명한다.Hereinafter, various aspects and various embodiments of the present invention will be described in more detail.
본 발명에 따른 누에 가수분해물의 제조방법은, 꾸지뽕잎 및 뽕잎으로 사육한 누에를 가수분해하는 단계를 포함할 수 있다.The method for producing silkworm hydrolyzate according to the present invention may include the step of hydrolyzing silkworms bred with courageous leaves and mulberry leaves.
본 발명에 따른 누에 가수분해물의 제조방법을 통하여 가수분해 전과 비교하여 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성을 증대시킬 수 있다.The method for producing silkworm hydrolyzate according to the present invention can increase Gaba and rutin enhancement activity, antioxidant activity or acetylcholinesterase inhibitory activity as compared to before hydrolysis.
본 발명에서 "가바(GABA)"는 감마 아미노낙산 혹은 감마 아미노부트릭산(gamma-aminobutyric acid의 약자)이라고도 하며, 주로 포유류의 뇌 속에만 존재하는 특이한 아미노산으로 중추신경계에 작용하는 억제 신경전달물질로 여겨지며, 신경계에서 신경흥분을 조정하는 역할을 하는 것으로 알려져 있다. 또한, 가바는 혈압강하, 이뇨작용, 뇌의 산소공급량을 증가시켜 뇌세포의 대사기능을 촉진시키고 신경을 안정시키며 불안감을 해소하기도 한다. 이외에도 인간의 뇌하수체 전엽에서 분비되는 성장호르몬을 증가시키고, 중성지질 및 콜레스테롤의 수치를 낮추는 것으로도 알려져 있다.In the present invention, "GABA" is also referred to as gamma aminobutyric acid or gamma aminobutyric acid (gamma-aminobutyric acid), an inhibitory neurotransmitter that acts on the central nervous system as a specific amino acid mainly present only in the mammalian brain. It is believed to play a role in controlling nervous excitement in the nervous system. In addition, Gaba lowers blood pressure, diuretic, and increases the oxygen supply of the brain to promote metabolism of brain cells, stabilize nerves and relieve anxiety. In addition, it is known to increase the growth hormone secreted by the anterior lobe of the human pituitary gland, and lower the level of neutral lipid and cholesterol.
본 발명에서 "루틴(Rutin)"은 콩과(Leguminosae) 식물인 회화나무(Sophora japonica)의 꽃봉오리 또는 마디풀과(Polygonaceae)의 식물인 메밀(Fagopyrum esculentum) 등에서 추출되는 물질로, 연한 노란색의 바늘 모양 결정 형태이다. 이러한 루틴은 모세혈관을 강화시키는 효능이 있어 동맥경화, 고혈압, 뇌출혈 등과 같은 심장계 질환을 예방하고, 당뇨병이나 비만 등을 치료하는 효능이 있는 것으로 알려져 있다.In the present invention, "Rutin" is a material extracted from a bud of Sophora japonica, a legume plant, or a buckwheat (Fagopyrum esculentum), a plant of the Polygonaceae, and a light yellow needle. It is a crystalline form. These routines are known to have the effect of strengthening capillaries, preventing cardiovascular diseases such as arteriosclerosis, hypertension, and cerebral hemorrhage, and treating diabetes and obesity.
본 발명에서 "항산화 활성"이란 생체 내 활성산소의 생성을 방지하고 세포에 회복 불가능한 손상을 야기하는 산화현상을 방지하는 활성을 말한다. 안정한 상태의 산소 (triplet oxygen)는 효소계, 환원대사, 화학약품, 공해물질, 광화학 반응과 같은 환경적 및 생화학적 요인 등에 의해 수퍼옥사이드 라디칼, 하이드록시 라디칼, 과산화수소와 같은 반응성이 큰 활성산소(Reactive Oxygen Species: ROS)로 전환되어 세포구성성분을 비가역적으로 파괴한다. 이들 활성산소의 작용은 체내 방어기구인 수퍼옥사이드디스뮤타제(SOD), 카탈라아제(catalase), 퍼옥시다아제(peroxidase) 등의 항산화성 효소 및 비타민 C(vitamin C), 비타민 E(vitamin E), 글루타티온 (glutathione) 등의 항산화 물질의 작용에 의하여 최소화 될 수 있다. 그러나, 이러한 생체 방어력에 이상이 생기거나 과도한 활성산소에 노출될 경우, 이 균형이 깨어져서 활성산소가 지질, 단백질, DNA 등을 비가역적으로 파괴하게 된다. 그 결과, 노화(aging), 암(cancer), 복합성 동맥경화(multiple atherosclerosis), 관절염 및 파킨슨병(Parkinson's disease)과 같은 각종 질병이 유발된다.As used herein, the term "antioxidant activity" refers to an activity that prevents the production of free radicals in vivo and prevents oxidative phenomena that cause irreparable damage to cells. The stable oxygen (triplet oxygen) is a highly reactive reactive oxygen such as superoxide radicals, hydroxy radicals and hydrogen peroxide due to enzymatic, reducing metabolism, chemicals, pollutants, environmental and biochemical factors such as photochemical reactions. Oxygen Species (ROS), which irreversibly destroy cellular components. The action of these free radicals is the body's defense mechanisms such as superoxide dismutase (SOD), catalase (catalase), peroxidase (peroxidase) and other antioxidant enzymes (vitamin C), vitamin E (vitamin E), glutathione ( It can be minimized by the action of antioxidants such as glutathione). However, if the biological defenses are abnormal or exposed to excessive free radicals, the balance is broken and the free radicals irreversibly destroy lipids, proteins, DNA, and the like. As a result, various diseases such as aging, cancer, multiple atherosclerosis, arthritis and Parkinson's disease are caused.
본 발명에서 "아세틸콜린에스테라제 저해 활성"는 아세틸콜린에스테라제를 억제함으로써 시냅스(synapse) 내의 아세틸콜린(acetylcholine) 농도를 증가시키고, 신경보호작용(neuroprotective effect) 및 신경손상을 회복시키는 작용을 의미한다. 예를 들어, 알츠하이머성 질환의 경우 뇌 안에서의 아밀로이드 베타 단백질의 축적에 의한 아세틸콜린(acetylcholine, ACh; neurotransmitter)의 레벨(level) 감소가 그 원인의 하나로 지목되고 있는 바 아세틸콜린에스테라제 저해 활성이 알츠하이머성 질환의 치료를 위한 핵심요소가 될 수 있다.In the present invention, "acetylcholinesterase inhibitory activity" is an action to increase the concentration of acetylcholine in the synapse (synapse) by restoring acetylcholinesterase, and to restore the neuroprotective effect and nerve damage Means. For example, in the case of Alzheimer's disease, a decrease in the level of acetylcholine (ACh; neurotransmitter) caused by the accumulation of amyloid beta protein in the brain is considered as one of the causes of acetylcholinesterase inhibitory activity It may be a key factor for the treatment of Alzheimer's disease.
또한, 상기 "아세틸콜린에스테라제"란 아세틸콜린 가수 분해 효소를 말한다. 뇌조직의 모든 신경세포에서 발견되는 신경전달물질로서 아세틸콜린(Acetylcholine ;ACh)은 시냅스(synapse)와 시냅스 사이의 신경전달에 관계하는 중요한 신경전달물질로 알려져 있다. 뇌신경계의 특정부위에서 아세틸콜린(ACh)이 시냅스 전 말단에서 분비되면 그것이 시냅스 후 수용체와 결합하여 신경세포 사이의 자극을 전달한다. 그러나 제2의 자극이 시냅스를 통해 전달하기 전에 제1의 자극 시에 분비된 아세틸콜린(ACh)은 아세틸콜린에스테라제(acetylcholinesterase ;AChE)에 의하여 가수분해 되어야한다. 그런데 아세틸콜린(ACh)의 함량과 합성효소로서 콜린아세틸트랜스퍼라제(choline acetyltransferase ;ChAT)의 활성이 대부분의 사람 및 설치동물에서 연령과 함께 감소하는 것으로 알려져 있다. 또한 아세틸콜린에스테라제의 활성도 아세틸콜린(ACh)과 마찬가지로 감소한다는 사실도 밝혀지고 있다.In addition, the said "acetylcholinesterase" means an acetylcholine hydrolase. Acetylcholine (ACh), a neurotransmitter found in all nerve cells of brain tissue, is known as an important neurotransmitter involved in neurotransmission between synapses and synapses. When acetylcholine (ACh) is secreted at the presynaptic end in certain parts of the cranial nervous system, it binds to postsynaptic receptors and transmits stimuli between nerve cells. However, acetylcholine (ACh) secreted at the first stimulus must be hydrolyzed by acetylcholinesterase (AChE) before the second stimulus transfers through the synapse. However, the content of acetylcholine (ACh) and the activity of choline acetyltransferase (ChAT) as a synthetase are known to decrease with age in most humans and rodents. It has also been found that the activity of acetylcholinesterase decreases like acetylcholine (ACh).
상기 누에는 씨누에에서 3 내지 4령까지는 뽕잎으로 사육된 후, 숙잠기까지 꾸지뽕잎으로 사육된 것일 수 있다. 특히, 누에를 처음부터 꾸지뽕 잎으로만 사육할 경우에는 뽕잎으로만 사육한 누에에 비하여 생체중 및 생존율이 감소되는 데 반해, 상기와 같이 3 내지 4령까지는 뽕잎으로 사육 후, 숙잠기까지 꾸지뽕 잎으로 사육할 경우에는 뽕잎으로만 사육한 누에의 생체중 및 생존율과 유의적인 차이가 나타나지 않을 뿐만 아니라, 꾸지뽕잎의 영양성분이 함유되므로 사육의 효율을 극대화됨을 확인하였다. 상기 숙잠기는 5령 5일 정도의 시기로 누에 단백질을 최대로 수확 가능한 시기이다.The silkworms are bred in mulberry leaves from 3 to 4 years old, and may be bred into cuddle mulberry leaves until sleep. In particular, when the silkworms are bred only from the first time, the live weight and survival rate are reduced compared to the silkworms raised only with the mulberry leaves. When breeding not only did not show a significant difference with the live weight and survival rate of silkworms bred only in mulberry leaves, it was confirmed that the efficiency of the breeding is maximized because it contains the nutrients of custard mulberry leaves. The snoozing period is the time to harvest the silkworm protein to the maximum of about 5 days and 5 days.
다음으로, 상기 가수분해는 브로멜린, 파파인, 프로타멕스, 트립신, 알카라아제, 뉴트라아제 및 플라보자임 중에서 선택되는 1종 이상의 단백질 가수분해효소로 효소가수분해 하는 것일 수 있으며, 바람직하게는 브로멜린, 파파인 및 트립신의 복합효소일 수 있다.Next, the hydrolysis may be to hydrolyze the enzyme with one or more proteolytic enzymes selected from bromelain, papain, protamex, trypsin, alkalase, neutrase and flavozyme, preferably Complex enzymes of bromelain, papain and trypsin.
상기 복합효소를 사용하여 가수분해할 경우 단일 효소를 사용한 경우에 비하여 가바 및 루틴 함량이 증진되고, 항산화 활성 및 아세틸콜린에스테라제 저해 활성이 증진될 뿐만 아니라, 이를 급여한 마우스의 혈압강하 및 지질 함량을 현저히 감소시키는 효과가 있음을 확인하였다.When hydrolyzed using the complex enzyme, Gaba and rutin contents are enhanced, antioxidant activity and acetylcholinesterase inhibitory activity are enhanced as compared to the case of using a single enzyme, and blood pressure drop and lipid of mice fed the same It was confirmed that there is an effect to significantly reduce the content.
상기 가수분해는 30 내지 65 ℃, 바람직하게는 35 내지 65 ℃, 더욱 바람직하게는 45 내지 65 ℃에서, 0.5 내지 10 시간, 바람직하게는 0.3 내지 5 시간, 더욱 바람직하게는 0.5 내지 2 시간 동안 수행될 수 있으며, 효소 종류에 따른 활성 온도 범위에서 수행되는 것이 바람직하다.The hydrolysis is carried out at 30 to 65 ° C, preferably at 35 to 65 ° C, more preferably at 45 to 65 ° C, for 0.5 to 10 hours, preferably for 0.3 to 5 hours, more preferably for 0.5 to 2 hours. It is preferable to be carried out in the active temperature range according to the type of enzyme.
또한, 상기 가수분해는 0.01 내지 0.5 MPa, 바람직하게는 0.05 내지 0.3 MPa, 더욱 바람직하게는 0.08 내지 0.1 MPa의 감압 조건에서 수행될 수 있다.In addition, the hydrolysis may be carried out under reduced pressure of 0.01 to 0.5 MPa, preferably 0.05 to 0.3 MPa, more preferably 0.08 to 0.1 MPa.
특히, 상기 가수분해는, 상기 가수분해효소를 함유하는 효소액 및 상기 누에를 감압 가수분해 장치에 투입한 후 상기 장치의 상·하측에 연결되는 순환라인을 통하여 상기 효소액을 장치 내부에서 순환시킴으로써 수행될 수 있다.In particular, the hydrolysis may be performed by circulating the enzyme solution in the apparatus through a circulating line connected to the upper and lower sides of the enzyme after the enzyme solution containing the hydrolase and the silkworm are put into a reduced pressure hydrolysis apparatus. Can be.
상기 연화탱크 내부 효소액의 상기 순환라인에 의한 1 회 순환시간은 30 초 내지 5 분, 바람직하게는 50 초 내지 4 분, 더욱 바람직하게는 1 분 내지 3 분일 수 있다. One cycle time of the softening tank by the circulation line by the circulation line may be 30 seconds to 5 minutes, preferably 50 seconds to 4 minutes, more preferably 1 minute to 3 minutes.
상기한 바와 같이, 본 발명에 따른 누에 가수분해물의 제조방법은 감압효소처리와 동시에 빠른 속도로 효소액이 순환되기 때문에 종래에 비하여 현저히 적은 효소를 사용함에도 불구하고 상기 탱크 내 효소액 전체의 균일한 혼합이 가능하고, 상기 누에에 균일하게 효소가 침투되어 현저히 향상된 가수분해 효과를 가질 수 있다.As described above, the method for producing silkworm hydrolyzate according to the present invention, because the enzyme solution is circulated at a high speed at the same time as the decompression enzyme treatment, despite the use of significantly less enzyme than the conventional uniform mixing of the entire enzyme solution in the tank If possible, the silkworm may be uniformly infiltrated with an enzyme to have a significantly improved hydrolysis effect.
또한, 상기 가수분해 시 산 또는 염을 사용하지 않아, 저비용 및 환경 친화적인 효과가 있다.In addition, since the acid or salt is not used during the hydrolysis, there is a low cost and environmentally friendly effect.
본 발명에 따른 누에 가수분해물의 제조방법의 구체적인 예로는, (a) 씨누에에서 3 내지 4령까지는 뽕잎으로 사육된 후, 숙잠기까지 꾸지뽕잎으로 사육된 누에를 동결 처리 및 조분쇄하는 단계; (b) 조분쇄된 누에를 감압진공분해 장치에 투입하는 단계; (c) 상기 조분쇄된 누에가 투입된 장치 내부에 효소액을 투입 및 밀폐하는 단계; (d) 상기 조분쇄된 누에 및 효소액이 투입된 장치 내부를 30 내지 65 ℃, 바람직하게는 35 내지 65 ℃, 더욱 바람직하게는 45 내지 65 ℃로 유지하는 단계; (e) 상기 장치 내부를 0.01 내지 0.5 MPa, 바람직하게는 0.05 내지 0.3 MPa, 더욱 바람직하게는 0.08 내지 0.1 MPa로 유지하면서, 장치의 상·하측에 연결되는 순환라인을 통하여 상기 효소액을 장치 내부에서 순환시켜 상기 조분쇄된 누에를 가수분해시키는 단계; 및 (f) 70 내지 120 ℃, 바람직하게는 75 내지 110 ℃, 더욱 바람직하게는 80 내지 100 ℃에서 1 내지 60 분, 바람직하게는 3 내지 30 분, 더욱 바람직하게는 5 내지 15 분 동안 상기 효소를 불활성화시키는 단계를 포함할 수 있다.Specific examples of the method for producing a silkworm hydrolyzate according to the present invention include: (a) freezing and coarsely pulverizing silkworms bred with Cuji mulberry leaves after breeding in mulberry leaves from 3 to 4 years old; (b) injecting the coarsely ground silkworm into the vacuum decompression apparatus; (c) injecting and sealing an enzyme solution in the apparatus into which the coarsely ground silkworm is put; (d) maintaining the inside of the apparatus into which the coarsely ground silkworm and the enzyme solution are added at 30 to 65 ° C, preferably 35 to 65 ° C, more preferably 45 to 65 ° C; (e) While maintaining the inside of the apparatus at 0.01 to 0.5 MPa, preferably 0.05 to 0.3 MPa, more preferably 0.08 to 0.1 MPa, the enzyme liquid is introduced into the apparatus through a circulation line connected to the upper and lower sides of the apparatus. Circulating to hydrolyze the coarsely ground silkworm; And (f) the enzyme for 1 to 60 minutes, preferably 3 to 30 minutes, more preferably 5 to 15 minutes at 70 to 120 ° C, preferably 75 to 110 ° C, more preferably 80 to 100 ° C. May comprise the step of inactivating.
상기 효소액은 상기 조분쇄된 누에 대비 2 내지 20 배, 바람직하게는 4 내지 15 배, 더욱 바람직하게는 6 내지 10배 중량 사용할 수 있고, 상기 효소는 상기 조분쇄된 누에 대비 0.1 내지 20 중량%, 바람직하게는 0.3 내지 10 중량%, 더욱 바람직하게는 0.5 내지 2 중량 사용할 수 있다.
The enzyme solution may be used 2 to 20 times, preferably 4 to 15 times, more preferably 6 to 10 times the weight of the coarsely ground silkworm, the enzyme is 0.1 to 20% by weight relative to the coarsely ground silkworm, Preferably from 0.3 to 10% by weight, more preferably from 0.5 to 2% by weight can be used.
이하에서는 본 발명에 따른 제조예 및 실시예를 첨부된 도면과 함께 구체적으로 설명한다.
Hereinafter, the production examples and embodiments according to the present invention will be described in detail with the accompanying drawings.
<제조예><Production example>
제조예 1 내지 3: 누에의 사육Production Examples 1 to 3: breeding of silkworms
1령에서 5령까지 뽕잎 및 꾸지뽕잎으로 사육한 숙잠누에를 각각 제조예 1 및 2로 하였고, 1령에서 4령까지는 뽕잎으로 사육한 후 5령까지는 꾸지뽕잎으로 사육한 숙잠누에(뽕잎/꾸지뽕)를 제조예 3으로 하였다.Sukjam silkworms bred with mulberry leaves and cuddle mulberry leaves from 1 to 5 were prepared in Examples 1 and 2, respectively. ) Was prepared as Preparation Example 3.
이 후, 상기 제조예 1 내지 3의 숙잠누에의 생체중을 측정하였고, 수확 시기까지의 생존률을 계산하여 하기 표 1에 나타내었다.
Thereafter, the live weight of the silkworm silkworms of Preparation Examples 1 to 3 was measured, and survival rates until harvesting time were calculated and shown in Table 1 below.
하기 표 1을 참조하면, 꾸지뽕잎으로 사육한 숙잠누에의 생체중은 뽕잎으로 사육한 누에의 약 65% 수준이었으나, 뽕잎/꾸지뽕잎으로 사육한 숙잠누에의 령별 생체중은 약 93% 수준이었고, 생존율도 뽕잎으로 사육한 숙잠누에와 유의적 차이는 나타나지 않았다.
Referring to Table 1 below, the live weight of sukjam silkworms bred with cooji mulberry leaves was about 65% of the silkworms bred with mulberry leaves, but the live weight of sukjam silkworms bred with mulberry leaves / kuji mulberry leaves was about 93%, and the survival rate was also high. No significant difference was found between Suksam silkworms grown with mulberry leaves.
(%)Survival rate
(%)
<실시예><Example>
실시예 1 내지 4: 효소 종류별 뽕잎/꾸지뽕잎 누에의 가수분해물 제조Examples 1 to 4: Preparation of hydrolyzate of mulberry leaf / custard mulberry silkworm by enzyme type
상기 제조예 3의 뽕잎/꾸지뽕잎으로 사육한 숙잠누에를 동결 처리 및 조분쇄한 후, 감압진공분해 장치에 상기 조분쇄된 숙잠누에 대비 8 배 중량의 효소액과 함께 투입 및 밀폐하였다. 상기 밀폐된 장치 내부를 50 ℃ 및 0.09 MPa로 유지하면서, 장치의 상·하측에 연결되는 순환라인을 통하여 상기 효소액을 장치 내부에서 1 회 순환시간이 2 분이 되도록 순환시켜 상기 조분쇄된 숙잠누에를 1 시간 동안 가수분해시켰다. 이 후, 장치 내부를 90 ℃로 10 분 동안 유지하면서 상기 효소를 불활성화시킴으로써 효소 종류별 뽕잎/꾸지뽕잎 누에 가수분해물을 제조하였다.After freezing treatment and coarsely pulverized sukjam silkworms bred with the mulberry leaf / Cucumber mulberry leaves of Preparation Example 3, the decompression vacuum decomposition device was added and sealed with 8 times the weight of the enzyme solution compared to the coarsely crushed silkworm silk. While maintaining the sealed apparatus at 50 ° C. and 0.09 MPa, the enzyme liquid was circulated in the apparatus for one minute of circulation time through a circulation line connected to the upper and lower sides of the apparatus so that the crude sieved silkworm silkworm Hydrolysis for 1 hour. Thereafter, the enzyme was inactivated while maintaining the inside of the apparatus at 90 ° C. for 10 minutes, thereby preparing a hydrolyzate of mulberry leaf / cuji mulberry leaf by enzyme type.
상기 효소액에 브로멜린, 파파인, 프로타멕스 및 트립신을 각각 상기 조분쇄된 숙잠누에 대비 1 중량%를 함유시켰고, 상기 효소 종류별로 각각 실시예 1 내지 4로 하였다.
The enzyme solution contained 1% by weight of bromelain, papain, protamex and trypsin, respectively, compared to the coarsely sieved snooze, and each of the enzyme types was 1 to 4, respectively.
실시예 5: 복합효소를 이용한 뽕잎/꾸지뽕잎 누에의 가수분해물 제조Example 5 Preparation of Hydrolysates of Mulberry Leaf / Cucja Leaf Leaf Silkworm Using Complex Enzyme
상기 실시예 1과 동일하게 실시하되, 상기 효소액에 브로멜린, 파파인 및 트립신의 복합효소를 상기 조분쇄된 누에 대비 2 중량%(각각 0.5중량%) 함유시켜 1 시간 동안 가수분해시킨 후, 효소 불활성화시킴으로써 복합효소를 이용한 뽕잎/꾸지뽕잎 누에의 가수분해물을 제조하였다.
The same procedure as in Example 1, except that the enzyme solution containing 2% by weight (0.5% by weight each) of the complex enzyme of bromelain, papain and trypsin compared to the coarsely pulverized silkworm and hydrolyzed for 1 hour, By activating the hydrolyzate of mulberry leaf / Cucumber leaf silkworm using a complex enzyme was prepared.
<비교예>Comparative Example
비교예 1: 상압 조건에서 복합효소를 이용한 뽕잎/꾸지뽕잎 누에의 가수분해물 제조Comparative Example 1 Preparation of Hydrolysates of Mulberry Leaf / Cucja Mulberry Leaf Silkworm Using Complex Enzyme at Normal Pressure
상기 제조예 3의 뽕잎/꾸지뽕잎으로 사육한 숙잠누에를 동결 처리 및 조분쇄한 후, 브로멜린, 파파인 및 트립신의 복합효소를 함유하는 효소액과 함께 진탕 배양기에서 50 ℃ 및 1 시간 조건으로 반응시킨 후, 90 ℃로 10 분 동안 유지하면서 상기 효소를 불활성화시킴으로써, 상업 조건에서 복합효소를 이용한 뽕잎/꾸지뽕잎 누에 가수분해물을 제조하였다.
After freeze treatment and coarsely pulverized simmered silkworms bred with the mulberry leaf / Cucumber leaf of Preparation Example 3, and reacted at 50 ℃ and 1 hour conditions in a shaker incubator with an enzyme solution containing complex enzyme of bromelain, papain and trypsin Thereafter, the enzyme was inactivated while maintaining the mixture at 90 ° C. for 10 minutes, thereby preparing a hydrolyzate of mulberry leaf / cusnip leaf silkworm using the complex enzyme under commercial conditions.
비교예 2: 복합효소를 이용한 뽕잎 누에의 가수분해물 제조Comparative Example 2: Preparation of Hydrolysates of Mulberry Leaf Silkworm Using Complex Enzyme
상기 실시예 5와 동일하게 실시하되, 제조예 3이 아닌 제조예 1의 뽕잎으로 사육한 숙잠누에를 가수분해하여 복합효소를 이용한 뽕잎 누에 가수분해물을 제조하였다.
In the same manner as in Example 5, but not hydrolyzed sukjam silkworms bred with the mulberry leaves of Preparation Example 1 but not Preparation Example 3 to prepare a mulberry leaf silkworm hydrolyzate using a complex enzyme.
<실험예>Experimental Example
실험예 1: 일반성분 분석Experimental Example 1: General Component Analysis
상기 제조예 1 및 3의 뽕잎 및 뽕잎/꾸지뽕잎으로 사육한 누에, 상기 실시예 1 내지 5 및 상기 비교예 1 내지 2의 누에 가수물해물의 일반성분을 AOAC법에 의하여 분석하고, 이를 하기 표 2에 나타내었다.
Silkworms bred with mulberry leaves and mulberry leaves / mulberry leaves of Preparation Examples 1 and 3, the general components of the silkworm hydrolyzate of Examples 1 to 5 and Comparative Examples 1 to 2 were analyzed by the AOAC method, and the following table 2 is shown.
하기 표 2를 참조하면, 뽕잎으로 사육한 누에에 비하여 뽕잎/꾸지뽕잎으로 사육한 누에의 조회분 및 조단백질의 함량이 다소 높았다.
Referring to Table 2 below, the content of crude ash and crude protein of silkworms bred with mulberry leaves / cuji mulberry leaves was higher than that of silkworms bred with mulberry leaves.
(mg/100g, D.B)View minutes
(mg / 100g, DB)
(mg/100g, D.B)Crude protein
(mg / 100g, DB)
(mg/100g, D.B)Crude fat
(mg / 100g, DB)
(mg/100g, D.B)carbohydrate
(mg / 100g, DB)
실험예Experimental Example 2: 2: 가바Gava 및 루틴 함량 분석 And routine content analysis
상기 제조예 1 및 3의 뽕잎 및 뽕잎/꾸지뽕잎으로 사육한 숙잠누에, 상기 실시예 1 내지 5 및 상기 비교예 1 내지 2의 누에 가수물해물의 가바 및 루틴 성분을 분석하여, 하기 표 5에 나타내었다.Sugjam silkworms bred with the mulberry leaves and mulberry leaves / mulberry leaves of Preparation Examples 1 and 3, the Gaba and rutin components of silkworm hydrolyzate of Examples 1 to 5 and Comparative Examples 1 to 2 were analyzed, Indicated.
루틴 분석은 건조시료 1 g을 85% methanol 100 mL에 혼합하여 80 ℃에서 2 시간 2 회 반복하여 추출하고 그 액을 0.45 um syringe filter로 여과하여 HPLC로 분석하였다. 가바는 시료 5 g에 0.02N HCl로 정용한 다음 0.45 μm syringe filter로 여과하여 측정하였다. 루틴과 가바의 HPLC 분석 조건은 하기 표 3에 나타내었고 가바 분석시 이동상 조성은 하기 표 4와 같다.The routine analysis was performed by mixing 1 g of a dry sample in 100 mL of 85% methanol and extracting twice at 80 ° C. for 2 hours. The solution was filtered with a 0.45 um syringe filter and analyzed by HPLC. Gabba was measured by applying 0.02N HCl to 5 g of the sample and then filtration with a 0.45 μm syringe filter. HPLC analysis conditions of Rutin and Gabba are shown in Table 3 below, and the mobile phase composition in Gabba analysis is shown in Table 4 below.
하기 표 5를 참조하면, 뽕잎으로 사육한 누에에 비하여 뽕잎/꾸지뽕잎으로 사육한 누에가 가바의 경우 4 배, 루틴의 경우 4.5 배 이상으로 높게 측정되었고, 가수분해물들(실시예 1 내지 5 및 비교예 1 내지 2)은 가수분해 전(제조예 1 및 3)에 비하여 가바 및 루틴 함량이 증가하였음을 확인할 수 있다.Referring to Table 5 below, silkworms bred with mulberry leaves / cucumber mulberry compared to silkworms bred with mulberry leaves were measured 4 times higher for Gaba, 4.5 times higher for rutin, and hydrolyzates (Examples 1 to 5 and Comparative Examples 1 to 2) can be seen that the content of Gava and rutin increased compared to before hydrolysis (Preparation Examples 1 and 3).
특히, 복합효소를 사용한 실시예 5의 경우에는 단일 효소를 사용한 경우에 비하여 현저히 높은 가바 및 루틴 함량을 나타내었고, 상압 조건에서 복합효소를 사용한 비교예 1의 경우에는 감압을 수행한 경우에 비하여 가바 및 루틴의 함량이 저하되었으며, 뽕잎만을 사육한 누에에 복합효소를 사용한 비교예 2의 경우에는 뽕잎/꾸지뽕잎을 사육한 누에를 사용한 경우에 비하여 가바 및 루틴의 함량이 저하됨을 확인할 수 있다.
In particular, in the case of Example 5 using a complex enzyme showed a significantly higher Gab and rutin content than a single enzyme was used, in the case of Comparative Example 1 using a complex enzyme at atmospheric pressure compared to the case of performing a reduced pressure And the content of rutin was lowered, in the case of Comparative Example 2 using a silkworm complex enzyme breeding mulberry leaves only can be confirmed that the content of Gaba and rutin is reduced compared to the case of using silkworms bred mulberry leaves / cuji mulberry leaves.
(min)Time
(min)
40 mM NaH2PO4(pH 7.8)Solvent A (%):
40 mM NaH 2 PO 4 (pH 7.8)
ACN:MeOH: D.W(45:45:10)Solvnet B (%):
ACN: MeOH: DW (45:45:10)
실험예Experimental Example 3: 총 폴리페놀 및 플라보노이드 함량 분석 3: Total Polyphenol and Flavonoid Content Analysis
상기 제조예 1 및 3의 뽕잎 및 뽕잎/꾸지뽕잎으로 사육한 숙잠누에, 상기 실시예 1 내지 5 및 상기 비교예 1 내지 2의 누에 가수물해물의 총 폴리페놀 및 플라보노이드 함량을 분석하여, 하기 표 6에 나타내었다.Sukjam silkworms bred with mulberry leaves and mulberry leaves / mulberry leaves of Preparation Examples 1 and 3, the total polyphenol and flavonoid content of silkworm hydrolyzate of Examples 1 to 5 and Comparative Examples 1 to 2 were analyzed, 6 is shown.
총 폴리페놀 함량은 알칼리 조건에서 Folin Ciocalteu 시약이 폴리페놀성 화합물에 의해 환원되어 몰리브덴 청색을 나타내는 방법을 이용하였다(Kim, 2009). 즉, 0.2 mL 의 추출물에 0.5 mL의 1 M Folin Ciocalteu를 가한 뒤 상온에서 3 분간 방치하였다. 10% Na2CO3 1.0mL를 가한 후 암소에서 한 시간 방치한 뒤 710 nm에서 흡광도(Elisa reader, Biotex unstrument powerwave XS, Pascalbio, Gyeonggido, Korea)를 측정하였다. 표준물질로는 0-0.1 mg/mL의 garlic acid를 사용하였으며 총 폴리페놀 함량은 시료 1 g당 mg garlic acid로 나타내었다. 플라보노이드 함량은 Jung 등(2004)의 방법을 변형하여 측정하였다. 즉 시료액 5 mL에 5% NaNO2 100 μL를 첨가하여 상온에서 5 분간 반응시킨 후에 10% AlCl3 200μL를 첨가하였다. 이 용액에 1 M NaOH 1 mL를 첨가한 후에 510 nm에서 흡광도를 측정하였으며, catechin을 표준물질로 사용하여 작성한 검량곡선으로 함량을 계산한 후에 추출물의 총 페놀성 화합물 함량을 추출물 mg당 μg catechin으로 나타내었다.
The total polyphenol content was used in a method in which the Folin Ciocalteu reagent was reduced by the polyphenolic compound to exhibit molybdenum blue under alkaline conditions (Kim, 2009). That is, 0.5 mL of 1 M Folin Ciocalteu was added to 0.2 mL of the extract, followed by standing at room temperature for 3 minutes. 1.0mL of 10% Na 2 CO 3 was added and left for 1 hour in the dark, and then absorbance was measured at 710 nm (Elisa reader, Biotex unstrument powerwave XS, Pascalbio, Gyeonggido, Korea). Garlic acid of 0-0.1 mg / mL was used as standard and total polyphenol content was expressed as mg garlic acid per gram of sample. Flavonoid content was measured by modifying the method of Jung et al. (2004). 5 mL of sample solution in 5% NaNO 2 After 100 μL was added and reacted at room temperature for 5 minutes, 200 μL of 10% AlCl 3 was added. After adding 1 mL of 1 M NaOH to this solution, absorbance was measured at 510 nm, and the content was calculated using a calibration curve prepared using catechin as a standard, and the total phenolic compound content of the extract was expressed in μg catechin per mg of extract. Indicated.
하기 표 6를 참조하면, 뽕잎으로 사육한 누에에 비하여 뽕잎/꾸지뽕잎으로 사육한 누에의 총 폴리페놀은 1.4 배 이상, 플라보노이드는 1.5 배 이상 높게 함유되어 있는 것으로 측정되었고, 가수분해물들(실시예 1 내지 5 및 비교예 1 내지 2)은 가수분해 전(제조예 1 및 3)에 비하여 총 폴리페놀 및 플라보노이드 함량이 증가하였음을 확인할 수 있다.Referring to Table 6 below, the total polyphenols of silkworms bred with mulberry leaves / cuji mulberry leaves compared to silkworms bred with mulberry leaves were measured to contain 1.4 times or more and 1.5 times or more flavonoids, respectively. 1 to 5 and Comparative Examples 1 to 2) showed that the total polyphenol and flavonoid content was increased compared to before hydrolysis (Preparation Examples 1 and 3).
특히, 복합효소를 사용한 실시예 5의 경우에는 단일 효소를 사용한 경우에 비하여 현저히 높은 총 폴리페놀 및 플라보노이드 함량을 나타내었고, 상압 조건에서 복합효소를 사용한 비교예 1의 경우에는 감압을 수행한 경우에 비하여 총 폴리페놀 및 플라보노이드의 함량이 저하되었으며, 뽕잎만을 사육한 누에에 복합효소를 사용한 비교예 2의 경우에는 뽕잎/꾸지뽕잎을 사육한 누에를 사용한 경우에 비하여 총 폴리페놀 및 플라보노이드의 함량이 저하됨을 확인할 수 있다.
In particular, Example 5 using the complex enzyme showed significantly higher total polyphenol and flavonoid content than the case of using a single enzyme, and in the case of Comparative Example 1 using the complex enzyme under normal pressure under reduced pressure The total polyphenol and flavonoid contents were lowered, and in the case of Comparative Example 2 using the silkworm complex enzyme in which only the mulberry leaves were raised, the total polyphenol and flavonoid contents were lower than in the case of using the silkworms in which the mulberry leaves / cuji mulberry leaves were raised. can confirm.
*GAE; garlic acid, CE; catechin
* GAE; garlic acid, CE; catechin
실험예Experimental Example 7: 7: DPPHDPPH 라디칼Radical 소거능Scavenging power 분석 analysis
상기 제조예 1 및 3의 뽕잎 및 뽕잎/꾸지뽕잎으로 사육한 숙잠누에, 상기 실시예 1 내지 5 및 상기 비교예 1 내지 2의 누에 가수물해물의 DPPH 라디칼 소거능을 분석하여, 하기 표 7에 나타내었다.Suksam silkworms bred with mulberry leaves and mulberry leaves / mulberry leaves of Preparation Examples 1 and 3, the DPPH radical scavenging ability of the silkworm hydrolyzate of Examples 1 to 5 and Comparative Examples 1 to 2 were analyzed, and is shown in Table 7 below. It was.
시료에 0.4 mM DPPH 용액 0.8 mL과 시료를 각각 0.2 mL 첨가하여 진탕하고 암실에서 10 분 동안 방치하였다. 그 후, spectrophotometer를 이용하여 517 nm에서 흡광도를 측정하였으며 이때 소거효과의 비교를 위한 대조군으로 ascorbic acid(ASA)를 사용하였다. 시료의 DPPH radical 소거능은 하기 수학식 1에 의해 나타내었다.
0.8 mL of 0.4 mM DPPH solution and 0.2 mL of the sample were added to the sample, shaken, and left in the dark for 10 minutes. Afterwards, the absorbance at 517 nm was measured using a spectrophotometer. Ascorbic acid (ASA) was used as a control for the comparison of the scavenging effect. DPPH radical scavenging ability of the sample is represented by the following equation (1).
[수학식 1][Equation 1]
하기 표 7을 참조하면, 뽕잎으로 사육한 누에에 비하여 뽕잎/꾸지뽕잎으로 사육한 누에의 DPPH 라디칼 소거능이 2 배 이상 향상되었고, 가수분해물들(실시예 1 내지 5 및 비교예 1 내지 2)은 가수분해 전(제조예 1 및 3)에 비하여 DPPH 라디칼 소거능이 3 내지 6.3 배 이상으로 증가하였음을 확인할 수 있다.Referring to Table 7 below, the DPPH radical scavenging ability of silkworms bred with mulberry leaves / cuji mulberry leaves was more than doubled, compared to the silkworms bred with mulberry leaves. It can be seen that the DPPH radical scavenging ability increased by 3 to 6.3 times or more as compared to before hydrolysis (Preparation Examples 1 and 3).
특히, 복합효소를 사용한 실시예 5의 경우에는 단일 효소를 사용한 경우에 비하여 현저히 높은 DPPH 라디칼 소거능을 나타내었고, 상압 조건에서 복합효소를 사용한 비교예 1의 경우에는 감압을 수행한 경우에 비하여 DPPH 라디칼 소거능이 저하되었으며, 뽕잎만을 사육한 누에에 복합효소를 사용한 비교예 2의 경우에는 뽕잎/꾸지뽕잎을 사육한 누에를 사용한 경우에 비하여 DPPH 라디칼 소거능이 저하됨을 확인할 수 있다.
In particular, in Example 5 using the complex enzyme showed significantly higher DPPH radical scavenging ability than when using a single enzyme, in the case of Comparative Example 1 using a complex enzyme at atmospheric pressure compared to the case of performing a reduced pressure DPPH radical The scavenging activity was decreased, and in the case of Comparative Example 2 using the silkworm complex enzyme breeding mulberry leaves, the DPPH radical scavenging activity was lowered compared to the case of using the silkworms growing mulberry leaves / cuji mulberry leaves.
실험예Experimental Example 8: 마우스 뇌조직의 아세틸콜린 함량 및 8: Acetylcholine Content in Mouse Brain Tissue and 아세틸콜린에스테라아제Acetylcholinesterase 활성 측정 Active measurement
상기 제조예 1 및 3의 뽕잎 및 뽕잎/꾸지뽕잎으로 사육한 누에, 상기 실시예 1 내지 5 및 상기 비교예 1 내지 2의 누에 가수물해물을 섭취한 마우스 각각의 뇌조직의 아세틸콜린 함량 및 이에 관련된 효소인 아세틸콜린에스테라아제 활성을 측정하여, 하기 표 8에 나타내었다.Acetylcholine content of brain tissues of mice ingested silkworms bred with mulberry leaves and mulberry leaves and mulberry leaves of Preparation Examples 1 and 3, Examples 1 to 5 and Comparative Examples 1 to 2, respectively The acetylcholinesterase activity, a related enzyme, was measured and shown in Table 8 below.
웅성 ICR 마우스 30 마리를 2 주간 예비 사육 후 실험하였다. 투여한 식이는 상기 제조예 1 및 3의 뽕잎 및 뽕잎/꾸지뽕잎으로 사육한 누에, 상기 실시예 1 내지 5 및 상기 비교예 1 내지 2의 누에 가수물해물을 각각 분무건조한 분말을 종류별 5% 첨가하여 8 주 동안 실험식이를 급여하고, 그 뇌조직의 아세틸콜린 및 관련효소에 미치는 식이의 영향을 평가하였다. 기억학습장애모델은 마우스 복강에 scopolamine(30 mg/kg BW)에 주사하여 유발시켰다.Thirty male ICR mice were tested after two weeks of preliminary breeding. The administered diet was the silkworms bred in the mulberry leaves and mulberry leaves / mulberry leaves of Preparation Examples 1 and 3, respectively, 5% by spray drying the silkworm hydrolyzate of Examples 1 to 5 and Comparative Examples 1 to 2 The diet was fed for 8 weeks and the effects of diet on acetylcholine and related enzymes in the brain tissues were evaluated. The memory learning disorder model was induced by injection of scopolamine (30 mg / kg BW) into the mouse abdominal cavity.
아세틸콜린 함량 측정은 Ganggani 등의 방법에 의하였다. 시료 50 ul에 1% hydroxylamine 50 ul를 첨가, 혼합 후 HCl로 pH로 조정하였다. 이후 FeCl3 500 ul를 혼합하여 530 nm에서 흡광도를 측정하였다. 아세틸콜린에스테라아제 효소는 각 microplate well에 0.1 M Tris buffer, pH 8.0(Trizma HCl＋Trizma base)을 300 μL, 0.01 M dithionitrobenzoic acid(DTNB; Sigma, USA) 20 μL, enzyme suspension(뇌조직 균질물) 10 μL을 연속적으로 첨가하고 흡광도 측정 직전에 기질인 0.1 M acetylthiocholine chloride(Sigma, USA) 10 μL을 첨가하였다. 96 well microplate reader(ELISA reader)를 이용하여 405 nm에서 흡광도 변화를 5 분 동안 관찰하여 아세틸콜린에스테라아제의 활성(unit/min/mg protein)을 측정하였다. Acetylcholine content was measured by the method of Ganggani et al. 50 ul of sample was added to 50 ul of 1% hydroxylamine and mixed with HCl to adjust pH. Thereafter, 500 ul of FeCl 3 was mixed to measure absorbance at 530 nm. Acetylcholinesterase enzyme contains 300 μL of 0.1 M Tris buffer, pH 8.0 (Trizma HCl + Trizma base), 20 μL of 0.01 M dithionitrobenzoic acid (DTNB; Sigma, USA) and 10 μL of enzyme suspension (brain tissue homogenate) in each microplate well. 10 μL of substrate 0.1 M acetylthiocholine chloride (Sigma, USA) was added immediately before the absorbance measurement. The acetylcholinesterase activity (unit / min / mg protein) was measured by observing absorbance change at 405 nm for 5 minutes using a 96 well microplate reader (ELISA reader).
아세틸콜린 Acetylcholine(ACh)은 뇌조직 중에 존재하는 가장 중요한 신경전달물질로 acetyl Co A와 choline acetyltransferase(ChAT) 효소의 작용을 받아 합성되며 이것은 다시 acetylcholinesterase(AChE) 효소의 작용을 받아 acetate와 choline으로 분해된다. 또한, Acetylcholine의 합성과 분해에 관련된 효소로는 cholineacetyltransferase(ChAT)와 acetylcholinesterase(AChE)가 있다. 치매의 50% 이상을 차지하고 있는 노인성 치매 환자의 경우 ChAT가 감소하는 경향을 나타내고 있으며 치매 환자의 치료에 있어 AChE 효소를 억제해 체내의 ACh의 농도를 증가시키는 방법이 이용되고 있다. Acetylcholine (ACh) is the most important neurotransmitter in the brain tissue and is synthesized under the action of acetyl Co A and choline acetyltransferase (ChAT) enzymes, which are further broken down into acetate and choline by the action of acetylcholinesterase (AChE) enzymes. do. In addition, the enzymes involved in the synthesis and degradation of acetylcholine include cholineacetyltransferase (ChAT) and acetylcholinesterase (AChE). Chronic dementia patients, which account for more than 50% of dementia, have a tendency to decrease ChAT. In the treatment of dementia patients, the method of increasing ACh concentration in the body by inhibiting AChE enzyme is used.
하기 표 8을 참조하면, 뽕잎으로 사육한 누에에 비하여 뽕잎/꾸지뽕잎으로 사육한 누에의 아세틸콜린함량이 2 배 이상 향상되고, 아세틸콜린아스테라제 활성 또한 저하됨을 확인할 수 있으며, 가수분해물들(실시예 1 내지 5 및 비교예 1 내지 2)은 가수분해 전(제조예 1 및 3)에 비하여 아세틸콜린의 함량이 증가하였고 아세틸콜린에스테라제의 활성은 감소된 것을 확인할 수 있으며, 이를 통하여 기억학습장애와 뇌건강기능 개선에 도움이 될 것으로 판단되었다. Referring to Table 8 below, compared to the silkworms bred with mulberry leaves, the acetylcholine content of silkworms bred with mulberry leaves / cuji mulberry leaves is more than doubled, and acetylcholinesterase activity is also lowered, and the hydrolyzate ( In Examples 1 to 5 and Comparative Examples 1 to 2), the acetylcholine content was increased and the acetylcholine esterase activity was decreased compared to before hydrolysis (Preparation Examples 1 and 3). It was found to be helpful for improving learning disability and brain health function.
특히, 복합효소를 사용한 실시예 5의 경우에는 단일 효소를 사용한 경우에 비하여 현저히 높은 아세틸콜린 함량 및 낮은 아세틸콜린에스테라제 활성을 나타내었고, 상압 조건에서 복합효소를 사용한 비교예 1의 경우에는 감압을 수행한 경우에 비하여 아세틸콜린 함량은 저하되고 아세틸콜린에스테라제 활성은 증가하였으며, 뽕잎만을 사육한 누에에 복합효소를 사용한 비교예 2의 경우에는 뽕잎/꾸지뽕잎을 사육한 누에를 사용한 경우에 비하여 아세틸콜린 함량은 저하되고 아세틸콜린에스테라제 활성은 증가하였음을 확인할 수 있다.In particular, Example 5 using a complex enzyme showed significantly higher acetylcholine content and lower acetylcholinesterase activity than a single enzyme, and in Comparative Example 1 using a complex enzyme under normal pressure under reduced pressure Acetylcholine content was lowered and acetylcholinesterase activity was increased as compared to the case of Comparative Example 2 using silkworm complex enzymes in which only mulberry leaves were bred. In comparison, the acetylcholine content was lowered and the acetylcholinesterase activity was increased.
(μg/mg protein)Acetylcholine Content
(μg / mg protein)
활성(unit/mg protein/min)Acetylcholinesterase
Activity (unit / mg protein / min)
실험예Experimental Example 9: 마우스 혈압강하 및 지질 함량 측정 9: Determination of Mouse Blood Pressure and Lipid Content
상기 제조예 1 및 3의 뽕잎 및 뽕잎/꾸지뽕잎으로 사육한 누에, 상기 실시예 1 내지 5 및 상기 비교예 1 내지 2의 누에 가수물해물을 섭취한 마우스의 혈압강하 및 지질 함량을 측정하여 하기 표 9에 나타내었다.By measuring the blood pressure drop and the lipid content of the mouse ingested silkworms bred in the mulberry leaves and mulberry leaves / mulberry leaves of Preparation Examples 1 and 3, Examples 1 to 5 and Comparative Examples 1 to 2 Table 9 shows.
대조군으로는 기본식이를 급여한 무첨가군, 고지방식이군으로 진행하였다. 이 때 기본식이는 5L79(LabDiet, Charles River, Wilmington, MA, USA), 고지방사료로는 AIN-76A(TestDiet®, Dyets, Bethlehem,PA, USA)에 우지를 40 중량%가 되게 혼합한 Dyets#101556를 사용하였고, 상기 제조예 1 및 3의 뽕잎 및 뽕잎/꾸지뽕잎으로 사육한 누에, 상기 실시예 1 내지 5 및 상기 비교예 1 내지 2의 누에 가수물해물은 각각 상기 기본식이에 분말 형태로 5 중량%를 혼합하여 급여하였으며, 8 주 동안 사육한 후 수축기혈압과 혈청내 지질 함량을 측정하였다. 수축기 혈압을 power lab혈압 측정기를 이용하여 tail cuff method(Cuevas et al., 1996)로 측정하였다. 혈청내 중성지질(TG, tryglyceride), 총콜레스테롤(TC, total cholesterol), LDL-콜레스테롤(LDL-C, LDL-cholesterol), HDL-콜레스테롤(HDL-C, HDL-cholesterol)은 혈청생화학자동분석기를 사용하여 측정하였다.As a control group, the diet-free, high-fat diet group was fed. At this time, the basic diet is 5L79 (LabDiet, Charles River, Wilmington, MA, USA), and for the high-fat diet, Dyets # mixed 40% by weight of tallow in AIN-76A (TestDiet®, Dyets, Bethlehem, PA, USA). 101556 was used, silkworms bred in the mulberry leaves and mulberry leaves / mulberry leaves of Preparation Examples 1 and 3, silkworm hydrolyzate of Examples 1 to 5 and Comparative Examples 1 to 2, respectively, in the form of powder on the basic diet Five percent by weight of the mixture was fed and fed for 8 weeks, after which systolic blood pressure and serum lipid content were measured. Systolic blood pressure was measured by the tail cuff method (Cuevas et al., 1996) using a power lab blood pressure meter. Serum triglyceride (TG, tryglyceride), total cholesterol (TC), LDL-cholesterol (LDL-C, LDL-cholesterol), HDL-cholesterol (HDL-C, HDL-cholesterol) Measured using.
하기 표 9를 참조하면, 뽕잎으로 사육한 누에를 급여한 경우에 비하여 뽕잎/꾸지뽕잎으로 사육한 누에를 급여한 경우 수축기혈압 및 혈청내 TG, TC, LDL-C는 감소하였고, HDL-C은 증가하는 경향을 나타내었고, 아세틸콜린아스테라제 활성 또한 저하됨을 확인할 수 있다. Referring to Table 9 below, systolic blood pressure and serum TG, TC, LDL-C were decreased when feeding silkworms bred with mulberry leaves / cuji mulberry leaves compared to those fed mulberry leaves. It showed a tendency to increase, it can be seen that acetylcholinesterase activity is also reduced.
또한, 가수분해물들(실시예 1 내지 5 및 비교예 1 내지 2)을 급여한 경우에는 가수분해 전(제조예 1 및 3)의 누에 원물을 급여한 경우에 비하여 수축기혈압 및 혈청내 TG, TC, LDL-C는 감소하였고, HDL-C은 증가하는 경향을 보였다. In addition, when the hydrolyzates (Examples 1 to 5 and Comparative Examples 1 and 2) were fed, systolic blood pressure and serum TG and TC were lower than those when the raw silk was fed before hydrolysis (Preparation Examples 1 and 3). , LDL-C decreased and HDL-C increased.
특히, 복합효소를 사용한 실시예 5를 급여한 경우에는 단일 효소를 사용한 실시예 1 내지 4를 급여한 경우에 비하여 수축기혈압 및 혈청내 TG, TC, LDL-C는 감소하였고, HDL-C는 증가하였고, 상압 조건에서 복합효소를 사용한 비교예 1을 급여한 경우에는 감압을 수행한 경우에 비하여 수축기혈압 및 혈청내 TG, TC, LDL-C는 증가하였고, HDL-C은 감소하였으며, 뽕잎만을 사육한 누에에 복합효소를 사용한 비교예 2를 급여한 경우에는 뽕잎/꾸지뽕잎을 사육한 누에를 사용한 경우에 비하여 수축기혈압 및 혈청내 TG, TC, LDL-C는 증가하였고, HDL-C는 감소하였음을 확인할 수 있다.
In particular, when fed Example 5 with a complex enzyme, systolic blood pressure and serum TG, TC, LDL-C were decreased, and HDL-C was increased as compared with Examples 1 to 4 using a single enzyme. In the case of feeding Comparative Example 1 using the complex enzyme under normal-pressure conditions, systolic blood pressure and serum TG, TC, and LDL-C were increased, HDL-C was decreased, and mulberry leaves were raised as compared with the case of decompression. In case of feeding Comparative Example 2 using complex enzyme in one silkworm, systolic blood pressure and serum TG, TC, and LDL-C were increased and HDL-C were decreased compared to the silkworms in which mulberry leaves / cuji mulberry leaves were used. can confirm.
(mmHg)Systolic blood pressure
(mmHg)
실험예Experimental Example 10: 10: 효소액Enzyme solution 순환시간에 따른 성분 함량 측정 Component Content Measurement According to Circulation Time
상기 실시예 5의 복합효소를 이용한 뽕잎/꾸지뽕잎 누에의 가수분해물을 제조시, 장치 내부 효소액의 순환라인에 의한 1 회 순환시간에 따라 가바 및 루틴의 함량과 총 폴리페놀 및 플라보노이드 함량을 측정하여, 하기 표 10에 나타내었다.When preparing the hydrolyzate of mulberry leaf / custard leaf silkworm using the complex enzyme of Example 5, by measuring the content of Gaba and rutin and the total polyphenol and flavonoid content according to one circulation time by the circulation line of the enzyme solution inside the device It is shown in Table 10 below.
하기 표 10을 참조하면, 장치 내부 효소액의 순환라인에 의한 1 회 순환시간이 10 분으로, 본 발명에 따른 순환시간 범위에 비하여 느릴 경우에는 가바, 루틴, 총 폴리페놀 및 플라보노이드 함량이 모두 현저히 낮은 수치를 보였으며, 순환시간이 20 초인 경우에는 10 분인 경우에 모든 함량의 수치가 높으나, 빠른 속도로 순환됨에 따라 효소액의 거품이 다량 발생되어 소포제를 사용해야 하는 문제가 생기는 것을 확인하였다.
Referring to Table 10 below, when one circulation time of the circulation line of the enzyme liquid inside the apparatus is 10 minutes, and when compared to the circulation time range according to the present invention, all of Gaba, rutin, total polyphenol and flavonoid contents are significantly low. When the circulation time was 20 seconds, all the values were high when 10 minutes, but it was confirmed that the problem of having to use the antifoaming agent was caused by the large amount of bubbles generated in the enzyme solution due to rapid circulation.
(mg/100g)Gava
(mg / 100g)
(mg/100g)Routine
(mg / 100g)
(mg GAE/100g)Total polyphenols
(mg GAE / 100g)
(mg CE/100g)Flavonoids
(mg CE / 100g)
*GAE; garlic acid, CE; catechin
* GAE; garlic acid, CE; catechin
그러므로 본 발명에 따르면, 씨누에에서 3 내지 4령까지는 뽕잎으로 사육된 후, 숙잠기까지 꾸지뽕잎으로 사육된 숙잠누에를 감압진공분해 장치를 통해 가수분해하여, 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성을 증대시킬 수 있다.Therefore, according to the present invention, three to four years old in silkworms were bred with mulberry leaves, and then simmered silkworms were bred with courageous mulberry leaves until slumbering, using a vacuum decompression device, so that GABA and rutin enhancement activity, antioxidant activity or It can increase the acetylcholinesterase inhibitory activity.
Claims (11)
상기 누에는 씨누에에서 3 내지 4령까지는 뽕잎으로 사육된 후, 숙잠기까지 꾸지뽕잎으로 사육된 것이고,
상기 가수분해는 브로멜린, 파파인 및 트립신의 복합효소로 효소가수분해 하는 것이며,
상기 가수분해는, 상기 누에 및 상기 가수분해효소를 함유하는 효소액을 감압 가수분해 장치에 투입한 후 상기 장치의 상·하측에 연결되는 순환라인을 통하여 상기 효소액을 장치 내부에서 순환시킴으로써 수행되는 것이며,
상기 감암 가수분해 장치 내부 효소액의 상기 순환라인에 의한 1 회 순환시간은 30 초 내지 5 분인 것을 특징으로 하는 누에 가수분해물의 제조방법.As a method for producing a silkworm hydrolyzate comprising the step of hydrolyzing silkworms bred with courageous leaves and mulberry leaves
The silkworms were bred with mulberry leaves from 3 to 4 years old, and then were bred with courageous leaves until sleep.
The hydrolysis is enzymatic hydrolysis with a complex enzyme of bromelain, papain and trypsin,
The hydrolysis is carried out by introducing the enzyme solution containing the silkworm and the hydrolase into a reduced pressure hydrolysis device and then circulating the enzyme liquid inside the device through a circulation line connected to the upper and lower sides of the device.
Method for producing a silkworm hydrolyzate, characterized in that the circulation time of the enzyme solution inside the gamyeo hydrolysis device by the circulation line is 30 seconds to 5 minutes.
상기 누에 가수분해물은 증대된 가바 및 루틴 증진 활성, 항산화 활성 또는 아세틸콜린에스테라제 저해 활성을 갖는 것을 특징으로 하는 누에 가수분해물의 제조방법.The method of claim 1,
The silkworm hydrolyzate is a method for producing silkworm hydrolyzate, characterized in that it has enhanced Gaba and rutin enhancement activity, antioxidant activity or acetylcholinesterase inhibitory activity.
상기 가수분해는 30 내지 65 ℃에서 수행되는 것을 특징으로 하는 누에 가수분해물의 제조방법.The method of claim 1,
The hydrolysis is a method for producing silkworm hydrolyzate, characterized in that carried out at 30 to 65 ℃.
상기 가수분해는 0.5 내지 10 시간 동안 수행되는 것을 특징으로 하는 누에 가수분해물의 제조방법.The method of claim 6,
The hydrolysis is a method for producing silkworm hydrolyzate, characterized in that performed for 0.5 to 10 hours.
상기 가수분해 시 산 또는 염을 사용하지 않는 것을 특징으로 하는 누에 가수분해물의 제조방법.The method of claim 1,
Method for producing a silkworm hydrolyzate, characterized in that no acid or salt is used during the hydrolysis.
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