KR102565191B1 - Method for preparing composite including allomyrina dichotoma larva extract, composite including allomyrina dichotoma larva extract thereby and food including the same - Google Patents
Method for preparing composite including allomyrina dichotoma larva extract, composite including allomyrina dichotoma larva extract thereby and food including the same Download PDFInfo
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- KR102565191B1 KR102565191B1 KR1020170146217A KR20170146217A KR102565191B1 KR 102565191 B1 KR102565191 B1 KR 102565191B1 KR 1020170146217 A KR1020170146217 A KR 1020170146217A KR 20170146217 A KR20170146217 A KR 20170146217A KR 102565191 B1 KR102565191 B1 KR 102565191B1
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- Prior art keywords
- extract
- beetle larvae
- composition
- rhinoceros
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- OYHQOLUKZRVURQ-IXWMQOLASA-N linoleic acid Natural products CCCCC\C=C/C\C=C\CCCCCCCC(O)=O OYHQOLUKZRVURQ-IXWMQOLASA-N 0.000 description 1
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- FDDDEECHVMSUSB-UHFFFAOYSA-N sulfanilamide Chemical compound NC1=CC=C(S(N)(=O)=O)C=C1 FDDDEECHVMSUSB-UHFFFAOYSA-N 0.000 description 1
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Classifications
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L29/00—Foods or foodstuffs containing additives; Preparation or treatment thereof
- A23L29/06—Enzymes
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/17—Amino acids, peptides or proteins
- A23L33/175—Amino acids
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L35/00—Food or foodstuffs not provided for in groups A23L5/00 – A23L33/00; Preparation or treatment thereof
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2250/00—Food ingredients
- A23V2250/02—Acid
- A23V2250/06—Amino acid
- A23V2250/0652—Tyrosine
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2250/00—Food ingredients
- A23V2250/20—Natural extracts
- A23V2250/204—Animal extracts
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Health & Medical Sciences (AREA)
- Nutrition Science (AREA)
- Chemical & Material Sciences (AREA)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
- Mycology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Microbiology (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Coloring Foods And Improving Nutritive Qualities (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
본 출원은 장수풍뎅이 유충 추출물 함유 조성물의 제조방법, 이에 의해 제조된 곤충 추출물 함유 조성물 및 이를 포함하는 식품에 관한 것이다. The present application relates to a method for preparing a composition containing an extract of rhinoceros beetle larvae, a composition containing an insect extract prepared thereby, and a food containing the same.
Description
본 출원은 장수풍뎅이 유충 추출물 함유 조성물의 제조방법, 이에 의해 제조된 곤충 추출물 함유 조성물 및 이를 포함하는 식품에 관한 것이다.The present application relates to a method for preparing a composition containing an extract of rhinoceros beetle larvae, a composition containing an insect extract prepared thereby, and a food containing the same.
2013년 곤충을 식량으로 추천하는 유엔 보고서가 나온 이후 미래 식량 및 환경에 대한 관심이 더욱 높아지면서 식용 곤충에 관한 연구가 활발히 이루어지고 있다. 곤충은 다량의 불포화 지방산 및 아미노산이 고루 들어있으며, 이외에도 다양한 비타민, 무기질, 키토산 등의 영양성분을 함유하고 있다. 특히, 장수풍뎅이 유충은 탄수화물 9.4%, 단백질 11.3%, 지방 6.1%으로 양질의 영양분을 함유하고 있다.Since the publication of the UN report recommending insects as food in 2013, research on edible insects has been actively conducted as interest in future food and the environment has increased. Insects contain a large amount of unsaturated fatty acids and amino acids evenly, and contain nutrients such as various vitamins, minerals, and chitosan. In particular, the rhinoceros beetle larvae contain 9.4% carbohydrate, 11.3% protein, and 6.1% fat, containing high-quality nutrients.
그러나 곤충을 원물 형태로 섭취하기에는 외관 및 곤충 특유의 이미, 이취 때문에 일반 소비자들이 쉽게 섭취하기에는 어려움이 있다. 따라서 곤충 식량의 혐오감을 없애고 유용한 영양성분들을 활용하고자 하는 시도가 있어 왔고, 특히 곤충의 불포화지방산의 조성과 같은 영양성분에 대한 연구 및 풍미개선에 대한 연구가 주로 이루어져 왔다. 다만, 장수풍뎅이 유충에 대해서는 대한민국 등록특허공보 제10-1533600호와 같은 기능성소재로서 사용하려는 시도는 있었었으나, 풍미개선과 관련된 연구는 미흡한 실정 식품 소재용 추출물에 관한 개발은 많이 이루어지지 않았다.However, it is difficult for general consumers to easily consume insects in raw form because of their appearance and insect-specific taste and odor. Therefore, attempts have been made to eliminate the aversion of insect food and to utilize useful nutrients, and in particular, studies on nutritional components such as the composition of unsaturated fatty acids of insects and research on flavor improvement have been mainly conducted. However, there have been attempts to use the beetle larvae as a functional material, such as Korean Patent Registration No. 10-1533600, but research related to flavor improvement is insufficient. Development of extracts for food materials has not been made much.
상기와 같은 문제를 해결하기 위해 본 출원은 곤충, 특히 장수풍뎅이 유충 추출물 함유 조성물의 제조방법에 관한 것으로, 장수풍뎅이 유충의 식이를 조절하는 단계, 상기 장수풍뎅이 유충을 분쇄하는 단계, 및 상기 분쇄물을 원심분리 하는 단계 및 원심 분리된 상등액에 단백질 분해 효소를 처리하는 단계를 포함하는, 식품 소재용 장수풍뎅이 유충 추출물 함유 조성물의 제조방법으로서, 상기 장수풍뎅이 유충 추출물은 헥사날(hexanal) 함량이 0mg/L 초과 0.7mg/L 이하인 것을 특징으로 하는, 제조방법을 제공하는 것을 목적으로 한다.In order to solve the above problems, the present application relates to a method for preparing a composition containing an extract of insects, in particular, rhinoceros beetle larvae, the step of controlling the diet of the rhinoceros beetle larvae, the step of grinding the rhinoceros beetle larvae, and the pulverized product A method for producing a composition containing an extract of rhinoceros beetle larvae for food material, comprising the step of centrifuging and treating the centrifuged supernatant with a proteolytic enzyme, wherein the extract of the rhinoceros beetle larvae has a hexanal content of 0 mg It is an object to provide a manufacturing method, characterized in that / L more than 0.7 mg / L or less.
또한, 본 출원은 상기 제조방법으로 제조된 식품 소재용 장수풍뎅이 유충 추출물 함유 조성물을 제공하는 것을 목적으로 한다.In addition, an object of the present application is to provide a composition containing an extract of beetle larvae for food material prepared by the above manufacturing method.
그리고 본 출원은 상기 조성물을 포함하는 식품을 제공하는 것을 목적으로 한다. And the object of this application is to provide a food containing the composition.
상기의 목적을 달성하기 위한 본 출원의 일 양태는 장수풍뎅이 유충의 식이를 조절하는 단계, 상기 장수풍뎅이 유충을 분쇄하는 단계 및 상기 분쇄물을 원심분리 하는 단계 및 원심 분리된 상등액에 단백질 분해 효소를 처리하는 단계를 포함하는, 식품 소재용 장수풍뎅이 유충 추출물 함유 조성물의 제조방법으로서, 상기 장수풍뎅이 유충 추출물은 헥사날(hexanal) 함량이 0mg/L 초과 0.7mg/L 이하인 것을 특징으로 하는, 제조방법을 제공할 수 있다.One aspect of the present application for achieving the above object is adjusting the diet of the rhinoceros beetle larvae, grinding the rhinoceros beetle larvae and centrifuging the ground material, and adding proteolytic enzymes to the centrifuged supernatant A method for producing a composition containing an extract of rhinoceros beetle larvae for food material, comprising a step of treating, wherein the extract of the rhinoceros beetle larvae has a hexanal content of greater than 0 mg/L and less than or equal to 0.7 mg/L. can provide.
장수풍뎅이 유충(장수애)은 (Trypoxylus dichotomus)는 딱정버레목 풍뎅이과 장수풍뎅이의 유충으로 메뚜기, 식용번데기, 백강잠누에, 갈색거저리 애벌레 (고소애), 흰점박이꽃무지 애벌레 (꽃무지), 쌍별귀뚜라미와 함께 국내에 식품원료로 등록되어 있는 곤충이다. 장수풍뎅이 유충의 영양성분은 탄수화물 9.4%, 단백질 11.3%, 지방 6.1%으로 양질의 영양분을 가지는 곤충으로 장수풍뎅이 유충을 원물 형태 그대로 섭취하기에는 익숙하지 않은 형태로 종래에는 단순히, 건조 분말화하여 사용되었으나, 장수풍뎅이 유충에 포함된 지방으로 인하여 분말의 형태가 곱지 않고, 이미 (곤충 특유의 비린맛)과 이취가 있어 풍미의 기호도가 매우 낮은 문제점이 있었다.The larvae of the beetle ( Trypoxylus ) dichotomus ) is a larva of the coleoptera chafer and rhinoceros beetle, which is registered as a food ingredient in Korea along with locusts, edible pupae, white silkworms, brown mealworm larvae (Kosorae), white-spotted flower caterpillars (flower muji), and twin crickets. It is an insect. The nutritional components of the beetle larvae are 9.4% of carbohydrates, 11.3% of protein, and 6.1% of fat, which are insects with high-quality nutrients. , Due to the fat contained in the rhinoceros beetle larvae, the shape of the powder is not fine, and it already has (insect-specific fishy taste) and off-odor, so the preference of the flavor is very low.
상기 식이를 조절하는 단계는 장수풍뎅이 유충을 분쇄하기 전에 식이조절시키는 단계로서 상기 식이조절은 밀가루 식이로 생육한 뒤 1 내지 72시간 동안 절식하여 이루어질 수 있다. 상기 장수풍뎅이 유충은 적당한 온도와 습도를 유지시켜 생육되는 것으로, 주로 톱밥이나 밀겨 등의 식이로 생육되는데, 장수풍뎅이 유충을 밀가루 식이로 생육 및 절식 없이 가공하였을 때는 유충의 내장에 있던 잔여 사료 및 배설물로 인하여 가공에도 불구하고 강한 이미, 이취가 있어 기호도를 더욱 떨어뜨리는 문제가 있다. 그러나, 장수풍뎅이 유충을 밀가루 식이로 생육한 뒤 절식시킴으로써 유충 내장의 사료 및 배설물의 함량을 낮출 수 있다. 더욱 구체적으로 상기 밀가루 식이로 생육한 뒤 절식은 24 내지 72시간동안 이루어질 수 있다. 절식이 1시간 미만 이루어질 경우 유충의 내장의 사료 및 배설물이 충분히 제거되지 않아 가공 후에도 이미, 이취가 남을 수 있고, 72시간 초과하여 절식이 이루어질 경우 장수풍뎅이 유충의 생육이 저해될 수 있는 문제가 있다. The step of adjusting the diet is a step of adjusting the diet before crushing the beetle larvae, and the diet control may be performed by fasting for 1 to 72 hours after growing on a flour diet. The rhinoceros beetle larvae are grown by maintaining appropriate temperature and humidity, and are mainly grown on a diet such as sawdust or wheat bran. Due to this, despite the processing, there is a problem that there is a strong image and off-flavor, further reducing the degree of preference. However, it is possible to lower the content of feed and excretion in the intestines of the beetle by growing the beetle larvae on a flour diet and then fasting. More specifically, fasting after growing on the flour diet may be performed for 24 to 72 hours. If the fasting is done for less than 1 hour, the feed and excretion of the intestines of the larvae are not sufficiently removed, so that an off-odor may already remain even after processing, and if the fasting is performed for more than 72 hours, the growth of the beetle larvae may be inhibited. .
상기 분쇄하는 단계는 후술하는 단백질 분해 효소의 작용이 용이할 수 있도록 절식시킨 장수풍뎅이 유충을 분쇄하는 단계로서 장수풍뎅이 유충을 부위별로 분해하거나 별도의 절단 공정 없이 장수풍뎅이 유충 모든 부위를 분쇄하여 사용할 수 있다. 분쇄를 용이하게 하며 단백질 분해 쇼소의 작용이 용이하도록 하기 위하여 일정량의 물을 첨가할 수 있다. 구체적으로 장수풍뎅이 유충 100중량부 대비 100 내지 900 중량부, 더욱 구체적으로 200 내지 800중량부의 물을 첨가할 수 있다, 물이 100 중량부 미만으로 첨가될 경우 장수풍뎅이 유충의 분쇄가 용이하지 않을 수 있고, 900 중량부 초과하여 첨가될 경우 고형분이 낮아 추후 농축 과정 등에서 공정 경제성이 떨어질 수 있다.The crushing step is a step of crushing the beetle larvae that have been fasted so that the action of the proteolytic enzyme described later can be easily performed. there is. A certain amount of water may be added to facilitate grinding and to facilitate the action of proteolytic shiso. Specifically, 100 to 900 parts by weight, more specifically, 200 to 800 parts by weight of water may be added, based on 100 parts by weight of the rhinoceros beetle larvae. And, if it is added in excess of 900 parts by weight, the solid content is low, and the economic efficiency of the process may be reduced in the subsequent concentration process.
상기 원심 분리하는 단계는 상기 장수풍뎅이 유충 분쇄물을 원심분리하여 지방과 침전물을 제외한 수용성 상등액만을 분리하는 단계이다. 구체적으로 상등액을 분리하는 단계는 장수풍뎅이 유충 분쇄물을 3000 내지 8000rpm에서 10 내지 30분간 원심분리하여 분리할 수 있다. 상기 범위 외에서는 장수풍뎅이 유충 분쇄물이 충분히 분리되지 않거나 공정경제성이 떨어지는 문제점이 있다.The centrifuging is a step of centrifuging the pulverized rhizome larvae to separate only the aqueous supernatant excluding fat and sediment. Specifically, the step of separating the supernatant may be separated by centrifuging the ground beetle larvae at 3000 to 8000 rpm for 10 to 30 minutes. Outside of the above range, there is a problem that the pulverized beetle larvae are not sufficiently separated or the process economy is poor.
상기 효소처리 단계는 상기 상등액에 단백질 분해효소를 처리하는 단계로서 체내에 흡수가 용이한 유리아미노산의 비율을 높이고 이미, 이취를 줄일 수 있다. 상기 단백질 분해효소는 특별히 제한되지는 않으나, 경제성을 고려하여 상업적으로 널리 사용되는 단백질 분해효소를 사용할 수 있고, 구체적으로는 펩신, 트립신, 플라보르자임(Flavourzyme), 프로타멕스(Protamex), 파파인, 알파키모트립신, 판크레아제 중에 선택된 어느 하나 이상일 수 있고, 더욱 구체적으로는 플라보르자임 및 프로타맥스 중 선택된 어느 하나 이상일 수 있고, 가장 구체적으로는 플라보르자임 및 프로타맥스을 함께 사용할 수 있다. 상기 플라보르자임은 아스퍼질러스 오리재 (Aspergillus oryzae) 유래의 단백질 분해 효소로 엔도 프로테이즈 (endoprotease) 및 엑소 프로테이즈 (exoprotease) 특성을 가지고 있고, 상기 프로타멕스는 바실러스 서브틸러스 (Bacillus sp.) 유래의 단백질 분해효소로 엔도 프로테이즈 (endoprotease)의 특성을 가지고 있다 (도 2 참조). 상기 단백질 분해 효소는 0.0001 내지 5% 첨가하여, 5분 내지 5시간동안 처리할 수 있다. 더욱 구체적으로는 0.001 내지 3% 첨가하여 30분 내지 2시간 동안 처리할 수 있다. 상기 범위외의 조건으로 단백질 분해효소가 첨가되거나 처리될 경우 조성물 내에 유리아미노산의 비율이 낮거나 이미, 이취를 줄일 수 없는 문제점이 있거나, 생산 효율이 떨어져 공정 경제성이 낮아지며, 조성물의 품질이 저하될 수 있는 문제점이 있다.The enzyme treatment step is a step of treating the supernatant with a proteolytic enzyme, which can increase the ratio of free amino acids that are easily absorbed into the body and reduce off-flavors. The proteolytic enzyme is not particularly limited, but commercially widely used proteolytic enzymes may be used in consideration of economic feasibility, specifically pepsin, trypsin, flavourzyme, protamex, papain , alpha chymotrypsin, may be any one or more selected from pancrease, more specifically, may be any one or more selected from flavorzyme and protamax, and most specifically, flavorzyme and protamax may be used together. . The flavorzyme is a proteolytic enzyme derived from Aspergillus oryzae and has endoprotease and exoprotease properties, and the protamex is a proteolytic enzyme derived from Bacillus subtilus ( It is a proteolytic enzyme derived from Bacillus sp.) and has the characteristics of an endoprotease (see FIG. 2). The proteolytic enzyme may be added in an amount of 0.0001 to 5% and treated for 5 minutes to 5 hours. More specifically, it may be treated for 30 minutes to 2 hours by adding 0.001 to 3%. When proteolytic enzyme is added or treated under conditions outside of the above range, the ratio of free amino acids in the composition is low or there is already a problem that off-flavor cannot be reduced, production efficiency is lowered, process economics is lowered, and the quality of the composition is lowered. There is a problem with
상기 장수풍뎅이 유충 추출물 함유 조성물은 0mg/L 초과 0.7mg/L 이하의 헥사날 (hexanal)함량을 가질 수 있다. 전술한 바와 같이 장수풍뎅이 유충은 가공하더라도 이미, 이취가 있을 수 있는데 본 출원에 따른 제조방법은 식이를 조절하는 단계를 통하여 제조된 장수풍뎅이 유충 추출물의 이취를 줄일 수 있어, 이취를 내는 산화 생성물 중 한 성분인 헥사날 (hexanal)이 0.7mg/L이하로 함유된다. (실험예 5 참조)The composition containing the rhinoceros beetle larvae extract may have a hexanal content of greater than 0 mg/L and less than 0.7 mg/L. As described above, even if the rhinoceros beetle larvae are processed, they may already have an off-flavor. One component, hexanal, is contained at less than 0.7mg/L. (See Experimental Example 5)
상기 제조방법은 장수풍뎅이 유충 추출물 함유 조성물을 살균, 냉각, 농축 및 동결 건조로 이루어진 군에서 선택된 1종 이상의 단계를 추가로 더 포함할 수 있다. The manufacturing method may further include at least one step selected from the group consisting of sterilization, cooling, concentration, and freeze-drying of the rhizome extract-containing composition.
상기 살균, 냉각하는 단계를 통하여 첨가된 효소를 불활성화 시키고 미생물을 조절하여 유통안전성을 확보할 수 있다. 구체적으로 상기 살균하는 단계는 90 내지 150℃에서 10 내지 120분, 구체적으로는 100 내지 130℃에서 30 내지 60분 이루어질 수 있다. 살균하는 단계가 90℃ 미만이거나 10분 미만에서 이루어질 경우 효소 실활 및 살균이 충분히 이루어지지 않을 수 있고, 150℃ 이상이거나 120분 이상에서 이루어질 경우 영양소가 파괴될 수 있다. 상기 냉각하는 단계는 단계는 0 내지 10℃에서 30분 내지 2시간, 구체적으로 1 내지 7℃에서 45분 내지 1.5시간 이루어질 수 있다. 냉각하는 단계가 0℃ 미만이거나 2시간 초과하여 이루어질 경우 공정경제성이 떨어질 수 있고, 10℃ 초과하거나 30분 미만으로 이루어질 경우 추출물함유 조성물이 충분히 냉각되지 않을 수 있다.Distribution safety can be secured by inactivating the added enzyme and controlling microorganisms through the sterilization and cooling steps. Specifically, the sterilizing step may be performed at 90 to 150 ° C for 10 to 120 minutes, specifically, at 100 to 130 ° C for 30 to 60 minutes. If the sterilization step is performed at less than 90 ° C. or less than 10 minutes, enzyme inactivation and sterilization may not be sufficiently performed, and nutrients may be destroyed when performed at 150 ° C. or more or 120 minutes or more. The cooling step may be performed at 0 to 10°C for 30 minutes to 2 hours, specifically at 1 to 7°C for 45 minutes to 1.5 hours. If the cooling step is performed at less than 0°C or for more than 2 hours, process economics may deteriorate, and when the temperature exceeds 10°C or is performed for less than 30 minutes, the extract-containing composition may not be sufficiently cooled.
상기 농축하는 단계를 통하여 고형분을 높게 하여 식품 소재로서 사용이 더 용이하도록 하는 단계를 포함한다. 농축 과정을 거치는 경우, 90℃이상 10분 이상 효소 실활 공정만 간단히 거친 후 농축하고, 살균 냉각의 공정은 농축 후에 하도록 한다. 구체적으로는 공지의 농축기를 이용하여 40 내지 60℃, 30 내지 50rpm, 20 내지 40mbar 조건에서 농축할 수 있다. 상기 농축하는 단계는 추출물 함유 조성물의 농도가 5 내지 30brix, 구체적으로는 10 내지 25brix가 되도록 농축할 수 있다. 상기 범위 외로 농축할 경우 후술되는 분말화 단계가 용이하지 않거나, 공정 경제성이 떨어질 수 있고, 식품 소재로서 이용하기 어려울 수 있다.It includes a step of increasing the solid content through the concentrating step to make it easier to use as a food material. In the case of the concentration process, only the enzyme deactivation process at 90 ° C. or more for 10 minutes or more is followed by concentration, and the sterilization cooling process is performed after concentration. Specifically, it can be concentrated at 40 to 60 ° C., 30 to 50 rpm, and 20 to 40 mbar conditions using a known concentrator. The concentrating step may be concentrated such that the concentration of the extract-containing composition is 5 to 30 brix, specifically 10 to 25 brix. If concentrated outside the above range, the powdering step described later may not be easy, the process economy may be reduced, and it may be difficult to use as a food material.
상기 동결건조하는 단계를 통하여 추출물 함유 조성물을 분말화 함으로써 식품 소재로서 저장이 용이하고 다양한 제품에 적용할 수 있다. 구체적으로 동결건조 후 분말화하는 방법을 사용할 수 있다. 추출하여 동결 건조한 분말은 일반적인 열풍건조 등의 방법으로 건조한 후 분말화 한 것에 비하여 지방 함량이 낮아 뭉치지 않고 고운 분말을 얻을 수 있다.By pulverizing the extract-containing composition through the freeze-drying step, it is easy to store as a food material and can be applied to various products. Specifically, a method of powdering after freeze-drying may be used. The extracted and freeze-dried powder has a lower fat content than powdered powder after drying by a method such as general hot air drying, so that a fine powder can be obtained without clumping.
상기 장수풍뎅이 유충 추출물 함유 조성물은 L-티로신(L-tyrosine)을 0.1 내지 1.0mg/ml로 포함할 수 있다. L-티로신은 단백질 가수분해 효소가 단백질을 가수분해 함으로써 생성되는 아미노산 중 하나로써, 본 출원은 단순히 장수풍뎅이 유충에 포함된 아미노산 뿐만 아니라 추가적으로 단백질 분해효소처리를 하였기 때문에 더욱 높은 L-티로신 함량을 나타낼 수 있다 (실험예 1 참조). The rhinoceros beetle larvae extract-containing composition may include 0.1 to 1.0 mg/ml of L-tyrosine. L-tyrosine is one of the amino acids produced by proteolytic enzyme hydrolysis of protein, and the present application shows a higher L-tyrosine content because it is additionally treated with proteolytic enzyme as well as the amino acid contained in rhinoceros beetle larvae. It can (see Experimental Example 1).
또한, 상기 제조방법은 장수풍뎅이 유충을 분쇄하는 단계 이후 수용성 영양성분만을 추출하는 단계를 더 포함할 수 있다. 상기 제조방법은 분쇄물 전체적으로 원심 분리 후 단백질 분해 효소를 처리함으로써 가용성 펩타이드 (L-티로신 등)를 많이 함유하는 추출물을 포함하는 조성물을 제조할 수도 있으나, 수용성 영양성분만을 추출함으로써 항산화력 및 항염증 효과를 높일 수 있다 (실험예 2 내지 4 참조). 구체적으로, 상기 수용성 영양성분을 추출하는 단계는 열수 추출을 통해 이루어 질 수 있고, 더욱 구체적으로는 80 내지 120℃에서 10 내지 30분간 추출, 가장 구체적으로는 100℃에서 20분간 추출하여 이루어질 수 있다. In addition, the manufacturing method may further include a step of extracting only water-soluble nutrients after the step of grinding the beetle larvae. In the above manufacturing method, a composition containing an extract containing a large amount of soluble peptides (L-tyrosine, etc.) can be prepared by treating the pulverized product with a proteolytic enzyme after centrifugation, but by extracting only water-soluble nutrients, antioxidant power and anti-inflammatory The effect can be enhanced (see Experimental Examples 2 to 4). Specifically, the step of extracting the water-soluble nutrients may be performed through hot water extraction, more specifically, extraction at 80 to 120 ° C. for 10 to 30 minutes, and most specifically, extraction at 100 ° C. for 20 minutes. .
또 다른 본 출원의 일 양태는, 상기의 제조방법으로 제조된, 식품 소재용 장수풍뎅이 유충 추출물 함유 조성물을 제공할 수 있다. 구체적으로 상기 조성물은 분말, 과립 또는 액상으로 제조될 수 있으며, 상기의 제조방법으로 제조되었기 때문에 이미, 이취가 낮으면서도 유리아미노산의 함량이 높다. 또한, 상기 조성물은 항산화, 항염증 효과를 가지면서도 항염증 작용에서 세포 사멸에 큰 영향을 주지 않는다 (실험예 2, 3 및 4 참조)Another aspect of the present application may provide a composition containing a beetle larvae extract for food material, prepared by the above manufacturing method. Specifically, the composition may be prepared in powder, granule or liquid form, and since it is prepared by the above preparation method, it already has a low off-flavor and high free amino acid content. In addition, the composition does not significantly affect cell death in anti-inflammatory action while having antioxidant and anti-inflammatory effects (see Experimental Examples 2, 3 and 4)
다른 본 출원의 일 양태는, 상기의 조성물을 포함하는 식품을 제공할 수 있다. 본 출원의 식품은 상기 조성물을 포함항에 따라, 이취가 개선되어 풍미가 향상되고, 항산화, 항염증 효능이 있다. Another aspect of the present application may provide a food containing the above composition. As the food of the present application includes the above composition, off-flavor is improved, flavor is improved, and antioxidant and anti-inflammatory effects are exhibited.
본 출원에 따른 장수풍뎅이 유충 추출물 함유 조성물의 제조방법, 이에 의해 제조된 곤충 추출물 함유 조성물 및 이를 포함하는 식품은 이미, 이취가 적어 풍미가 향상되고 유리아미노산의 함량이 높을 뿐만 아니라 항산화, 항염증 효능을 가지는 효과가 있다.The method for preparing a composition containing rhinoceros beetle larvae extract according to the present application, the insect extract-containing composition prepared thereby, and the food containing the same already have less off-flavor, improved flavor, and high content of free amino acids, as well as antioxidant and anti-inflammatory effects. has the effect of
도 1은 본 출원의 장수풍뎅이 유충 추출물 함유 조성물의 제조방법을 나타낸 흐름도이다.
도 2는 본 출원의 장수풍뎅이 유충 추출물에 처리한 단백질 분해 효소의 적정 처리 조건을 나타내었다.
도 3는 실험예 2에 따른 본 출원의 장수풍뎅이 유충 추출물의 전자 공여능을 DPPH 라디칼 소거능 실험으로 확인한 결과이다.
도 4은 실험예 2에 따른 본 출원의 장수풍뎅이 유충 추출물의 EC50(ppm) 값을 나타낸 표이다.
도 5은 실험예 3에 따른 본 출원의 장수풍뎅이 유충 추출물의 RAW 264.7 대식세포에서 Nitrite 생성 저해 효과를 나타낸 그래프로서 도 5a는 제조방법 (열수 추출 여부, 효소처리 여부)에 따른 Nirite 생성 저해 효과를 나타낸 그리프이고, 도 5b는 효소처리, 농도 차이에 따른 Nirite 생성 저해 효과를 나타낸 그래프이다.이다.
도 6은 실험예 4에 따른 본 출원의 장수풍뎅이 유충 추출물의 RAW 264.7 대식세포에서 세포 생존율 분석 결과를 나타낸 그래프이다.
도 7는 실험예 5에 따른 본 출원의 장수풍뎅이 유충 추출물의 제조 공정 중 헥사날 (Hexanal) 성분의 감소를 나타낸 것이다.1 is a flow chart showing a method for preparing a composition containing an extract of rhinoceros beetle larvae of the present application.
Figure 2 shows the appropriate treatment conditions of the proteolytic enzyme treated with the rhinoceros beetle larvae extract of the present application.
3 is a result of confirming the electron donating ability of the rhinoceros beetle larvae extract of the present application according to Experimental Example 2 through a DPPH radical scavenging activity test.
4 is a table showing the EC50 (ppm) values of the rhinoceros beetle larvae extract of the present application according to Experimental Example 2.
5 is a graph showing the nitrite production inhibitory effect in RAW 264.7 macrophages of the rhinoceros beetle larvae extract of the present application according to Experimental Example 3. FIG. Figure 5b is a graph showing the effect of inhibiting Nirite production according to enzyme treatment and concentration difference.
6 is a graph showing the results of cell viability analysis in RAW 264.7 macrophages of the rhinoceros beetle larvae extract of the present application according to Experimental Example 4.
Figure 7 shows the reduction of hexanal (Hexanal) component during the manufacturing process of the rhinoceros beetle larvae extract of the present application according to Experimental Example 5.
이하에서는 실시예를 통해 본 출원의 내용을 더욱 구체적으로 설명한다. 다만, 실시예는 본 출원의 일 예시일 뿐이고, 본 출원의 범위가 실시예의 범위로 한정되는 것을 의미하는 것은 아니다. Hereinafter, the contents of the present application will be described in more detail through examples. However, the examples are only examples of the present application, and do not mean that the scope of the present application is limited to the scope of the examples.
<실시예><Example>
제조예manufacturing example 1: 장수풍뎅이 유충 추출물의 제조 1: Preparation of rhinoceros beetle larvae extract
(1) 장수풍뎅이 유충의 식이를 조절하는 단계(1) Step of controlling the diet of rhinoceros beetle larvae
톱밥 식이하여 5-6개월 된 장수풍뎅이 유충은 사용 전 밀가루 식이로 변경하고, 절식하면서 배설물을 배출하는 과정을 거친다. 48시간 절식 과정을 거친 장수풍뎅이 유충은 세척한 후 냉동보관한다.Beetle larvae aged 5-6 months on a sawdust diet change to a flour diet before use, and go through the process of excreting excrement while fasting. After 48 hours of fasting, the beetle larvae are washed and stored frozen.
(2) 장수풍뎅이 유충에 가수, 분쇄하는 단계(2) Singing and pulverizing rhinoceros beetle larvae
상기 식이 조절한 장수풍뎅이 유충 100 중량부에 300 중량부 물을 첨가한 후 분쇄하여 추출 및 효소 처리가 용이한 액상으로 만든다.300 parts by weight of water is added to 100 parts by weight of the rhinoceros beetle larvae adjusted to the above diet, and then pulverized to make a liquid phase that is easy to extract and enzymatically treat.
(3) 장수풍뎅이 유충 분쇄물을 원심 분리하는 단계(3) centrifuging the ground beetle larvae
상기 분쇄물을 6000rpm, 20분 원심 분리한 후 상등액만 취한다. After centrifuging the pulverized product at 6000 rpm for 20 minutes, only the supernatant is taken.
(4) 장수풍뎅이 유충 원심 분리 상등액에 효소를 처리하는 단계(4) Enzyme treatment of the supernatant of centrifugation of rhinoceros beetle larvae
상기 원심 분리한 상등액에 단백질 분해 효소인 플라보르자임 (Flavourzyme) 또는 프로타맥스(Protamex)를 0.1%, 0.25%, 0.5%, 0.75%, 1% 첨가하고 플라보르자임 (Flavourzyme) 50℃, 프로타맥스(Protamex) 60℃온도에서 1, 2, 3, 4, 5시간 처리하거나, 플라보르자임 (Flavourzyme) 1% + 프로타맥스(Protamex) 1% 를 첨가하고 55℃에서 1, 2, 3, 4, 5시간 처리한 후 100, 10분 간 효소를 불활성화 시켰다.0.1%, 0.25%, 0.5%, 0.75%, or 1% of a proteolytic enzyme, Flavorzyme or Protamex, was added to the centrifuged supernatant, and Flavorzyme was heated at 50 ° C. Treatment for 1, 2, 3, 4, 5 hours at 60℃ of Protamex or adding 1% of Flavorzyme + 1% of Protamex and 1, 2, 3 at 55℃ , 4, 5 hours of treatment followed by 100, 10 minutes of enzyme inactivation.
(5) 장수풍뎅이 유충 효소 처리액을 살균, 냉각하는 단계(5) sterilizing and cooling the rhinoceros beetle larvae enzyme treatment solution
상기 효소 처리액을 100℃, 30분 간 살균하고, 4℃, 60분간 냉각하여 냉장 조건에서 장기간 보관이 가능하도록 하였다.The enzyme-treated liquid was sterilized at 100° C. for 30 minutes and cooled at 4° C. for 60 minutes to enable long-term storage under refrigerated conditions.
제조예manufacturing example 2: 장수풍뎅이 유충 추출물의 제조 2: Preparation of rhinoceros beetle larvae extract
(1) 장수풍뎅이 유충의 식이를 조절하는 단계(1) Step of controlling the diet of rhinoceros beetle larvae
밀겨 식이하여 5-6개월 된 장수풍뎅이 유충은 사용 전 밀가루 식이로 변경하고, 절식하면서 배설물을 배출하는 과정을 거친다. 48시간 절식 과정을 거친 장수풍뎅이 유충은 세척한 후 냉동보관한다.Beetle larvae aged 5-6 months on a wheat bran diet are changed to a wheat diet before use, and pass through the process of excretion while fasting. After 48 hours of fasting, the beetle larvae are washed and stored frozen.
(2) 장수풍뎅이 유충에 가수, 분쇄하는 단계(2) Singing and pulverizing rhinoceros beetle larvae
상기 식이 조절한 장수풍뎅이 유충 100 중량부에 300 중량부 물을 첨가한 후 분쇄하여 추출 및 효소 처리가 용이한 액상으로 만든다.300 parts by weight of water is added to 100 parts by weight of the rhinoceros beetle larvae adjusted to the above diet, and then pulverized to make a liquid phase that is easy to extract and enzymatically treat.
(3) 분쇄물을 추출하는 단계(3) extracting the pulverized product
상기 분쇄물을 100℃에서 20분간 추출하여 원심분리 전 수용성 영양 성분을 추출한다.The pulverized product is extracted at 100° C. for 20 minutes to extract water-soluble nutrients before centrifugation.
(4) 장수풍뎅이 유충 추출물을 원심 분리하는 단계(4) centrifuging the rhinoceros beetle larvae extract
상기 추출액을 6000rpm, 20분 원심 분리한 후 상등액만 취한다. After centrifuging the extract at 6000 rpm for 20 minutes, only the supernatant is taken.
(5) 장수풍뎅이 유충 원심 분리 상등액에 효소를 처리하는 단계(5) Enzyme treatment of the supernatant of the centrifugation of the rhinoceros beetle larvae
상기 원심 분리한 상등액에 단백질 분해 효소인 플라보르자임 (Flavourzyme) 또는 프로타맥스(Protamex)를 0.1%, 0.25%, 0.5%, 0.75%, 1% 첨가하고 플라보르자임 (Flavourzyme) 50℃, 프로타맥스(Protamex) 60℃에서 1, 2, 3, 4, 5시간 처리하거나, 플라보르자임 (Flavourzyme) 1% + 프로타맥스(Protamex)1% 첨가하고 55℃에서 1, 2, 3, 4, 5시간 처리한 후 100℃, 10분 간 효소를 불활성화 시켰다.0.1%, 0.25%, 0.5%, 0.75%, or 1% of a proteolytic enzyme, Flavorzyme or Protamex, was added to the centrifuged supernatant, and Flavorzyme was heated at 50 ° C. Treatment with Protamex at 60℃ for 1, 2, 3, 4, 5 hours, or 1% of Flavorzyme + 1% of Protamex added and 1, 2, 3, 4 at 55℃ , After treatment for 5 hours, the enzyme was inactivated at 100 ° C for 10 minutes.
(6) 장수풍뎅이 유충 효소 처리액을 살균, 냉각하는 단계(6) sterilizing and cooling the rhinoceros beetle larvae enzyme treatment solution
상기 효소 처리액을 100℃, 30분간 살균하고, 4℃, 60분간 냉각하여 냉장 조건에서 장기간 보관이 가능하도록 하였다.The enzyme-treated liquid was sterilized at 100° C. for 30 minutes and cooled at 4° C. for 60 minutes to enable long-term storage under refrigerated conditions.
실험예 1: 효소 종류 및 처리 조건에 따른 L-티로신 (L-tyrosine) 함량 측정Experimental Example 1: Measurement of L-tyrosine content according to enzyme type and treatment conditions
장수풍뎅이 유충 추출물 함유 조성물의 효소적 가수분해를 위한 적정 효소 농도 및 반응시간을 설정하기 위하여 가수분해 시간에 따른 가수분해물의 함량을 L-티로신 (L-tyrosine)을 측정하여 확인하였다. 플라보르자임 (Flavourzyme)과 프로타맥스(Protamex) 단일 효소 농도를 각각 0.1%, 0.25%, 0.5%, 0.75%, 1%로 하고, 반응 시간을 1, 2, 3, 4, 5시간으로 설정하여 실험하였다. 또한, 수용성 영양성분만의 추출단계 유무 및 가수분해 시간에 따른 가수분해물의 함량을 L-티로신 (L-tyrosine)을 측정하여 확인하였다. 수용성 영양성분만의 추출단계 유무의 차이 외 플라보르자임 (Flavourzyme)과 프로타맥스(Protamex)를 각 1%씩 혼합하고, 반응 시간을 1, 2, 3, 4, 5시간으로 설정하여 실험하였다.In order to set an appropriate enzyme concentration and reaction time for enzymatic hydrolysis of the rhizome extract-containing composition, the content of the hydrolyzate according to the hydrolysis time was confirmed by measuring L-tyrosine. Flavorzyme and Protamex single enzyme concentrations were set to 0.1%, 0.25%, 0.5%, 0.75%, and 1%, respectively, and the reaction time was set to 1, 2, 3, 4, and 5 hours. was experimented with. In addition, the presence or absence of an extraction step of only water-soluble nutrients and the content of the hydrolyzate according to the hydrolysis time were confirmed by measuring L-tyrosine. In addition to the difference in the presence or absence of the extraction step of only water-soluble nutrients, Flavorzyme and Protamex were mixed at 1% each, and the reaction time was set to 1, 2, 3, 4, and 5 hours.
그 결과 플라보르자임 (Flavourzyme)과 프로타맥스 (Protamex) 단일 효소 처리 및 플라보르자임 (Flavourzyme)과 프로타맥스 (Protamex) 혼합처리구 중 가수분해능은 플라보르자임 (Flavourzyme) + 프로타맥스 (Protamex) 혼합처리구의 가수분해능이 가장 우수하였으며, 플라보르자임 (Flavourzyme)과 프로타맥스 (Protamex) 단일 처리구는 유사한 수준의 가수분해능을 나타내었고, 농도와 시간이 증가할수록 우수한 가수분해능을 보였다. 반응시간 측면에서는 효소 처리 후 1시간부터 급격히 가수분해가 이루어지는 것으로 보이며, 이후에는 약간의 증가가 있었지만, 그 차이가 크지 않았다. 따라서, 플라보르자임 (Flavourzyme)과 프로타맥스 (Protamex) 효소의 단독 처리구는 1-5시간의 반응시간이 요구되는 것으로 보이며, 플라보르자임 (Flavourzyme)과 프로타맥스 (Protamex) 혼합 처리구는 1-3시간의 반응 시간이 필요한 것으로 판단되었다. As a result, the hydrolysis ability of Flavorzyme and Protamex single enzyme treatment and Flavorzyme and Protamex mixture treatment was higher than that of Flavorzyme + Protamex ) The hydrolysis ability of the mixed treatment was the best, and the single treatment of Flavourzyme and Protamex showed a similar level of hydrolysis, and as the concentration and time increased, the hydrolysis performance was excellent. In terms of reaction time, hydrolysis seems to occur rapidly from 1 hour after enzyme treatment, and there was a slight increase thereafter, but the difference was not large. Therefore, it seems that the reaction time of 1-5 hours is required for the single treatment of Flavorzyme and Protamex enzymes, and the treatment of Flavorzyme and Protamex mixed treatment is 1-5 hours. A reaction time of -3 hours was determined to be necessary.
플라보르자임 (Flavourzyme) + 프로타맥스 (Protamex) 혼합 처리구에서 장수풍뎅이 유충은 분쇄물에서 수용액을 추출 후 원심 분리하여 효소 처리하는 것이 가수분해 효율이 우수하였다. In the Flavorzyme + Protamex mixed treatment group, the hydrolysis efficiency of the rhizome was excellent when the aqueous solution was extracted from the pulverized material, centrifuged, and enzymatically treated.
이하의 실험예 2 - 5에서는 플라보르자임 (Flavourzyme)과 프로타맥스 (Protamex)를 각 1%씩 55℃, 2시간 혼합 처리한 것을 사용하였다.In Experimental Examples 2 to 5 below, Flavorzyme and Protamex were mixed at 55° C. for 2 hours at 1% each.
실험예 2: 장수풍뎅이 유충 추출물의 전자공여능 측정Experimental Example 2: Measurement of electron donating ability of rhinoceros beetle larvae extract
전자공여능은 DPPH(1,1 diphenyl-2-picrylhydrazyl)의 환원력을 이용하여 측정하였다. 제조방법 별(열수 추출 여부, 효소처리 여부) 98~50000ppm의 장수풍뎅이 유충 추출물 100 ㎕에 0.2mM DPPH 용액 100 ㎕를 가한 후 실온에서 30분간 반응시키고 525 nm에서 흡광도를 측정하여 산화 저해율을 계산하였고, 이를 도 3 및 4에 나타내었다. 그 결과 도 3에서 도시된 바와 같이 장수풍뎅이 유충 (장수애)의 수용성 영양성분 추출만을 한 경우를 제외하고 다른 제조방법으로 제조된 추출물은 항산화력이 있음이 확인되었고, 특히, 도 4에서 도시된 바와 같이 장수풍뎅이 유충 (장수애)의 수용성 영양성분을 추출단계 없이 플라보르자임 (Flavourzyme)과 프로타맥스 (Protamex) 효소 처리를 한 경우 EC50 값이 3953ppm 수준으로 우수한 항산화력이 있음을 확인하였다.The electron donating ability was measured using the reducing power of DPPH (1,1 diphenyl-2-picrylhydrazyl). 100 μl of 0.2 mM DPPH solution was added to 100 μl of 98-50000 ppm rhinoceros beetle larvae extract by manufacturing method (whether hot water extraction or enzyme treatment), followed by reaction at room temperature for 30 minutes. The oxidation inhibition rate was calculated by measuring the absorbance at 525 nm. , which is shown in Figures 3 and 4. As a result, as shown in FIG. 3, it was confirmed that the extracts prepared by other manufacturing methods had antioxidant power, except for the case of extracting only the water-soluble nutrients of the rhinoceros beetle larvae (Jangsuae). In particular, as shown in FIG. Likewise, when the water-soluble nutrients of the beetle larvae (Jangsuae) were treated with Flavorzyme and Protamex enzymes without an extraction step, it was confirmed that the EC50 value was 3953ppm and had excellent antioxidant power.
실험예 3: 장수풍뎅이 유충 추출물의 NO 생성 저해 효과Experimental Example 3: NO production inhibitory effect of rhinoceros beetle larvae extract
장수풍뎅이 유충의 항염증 효능을 분석하기 위해 LPS(Lipopolysaccharide)를 통해 염증반응을 유도한 RAW 264.7 세포에 장수풍뎅이 유충 추출물을 처리하여 생성된 NO의 양을 세포 배양액 중에 존재하는 NO2의 형태로서 측정하였다. In order to analyze the anti-inflammatory efficacy of rhinoceros beetle larvae, RAW 264.7 cells, in which inflammatory response was induced through LPS (Lipopolysaccharide), were treated with rhizome larvae extract, and the amount of NO produced was measured as the form of NO 2 present in the cell culture medium. did
Raw 264.7 세포는 페니실린-스트렙토마이신(penicillin-streptomycin) 100 unit/ml과 10% 소태아혈청(fetal bovine serum/ Gibco, MD, USA)이 함유된 배지(Dublecco's Modified Eagle Medium/ Gibco, MD, USA)를 사용하여 37℃, 5% CO2 인큐베이터(incubator/Thermo Scientific, IL, USA)에서 배양하였으며, 2일 간격으로 계대 배양을 실시하였다.Raw 264.7 cells were cultured in medium (Dublecco's Modified Eagle Medium/ Gibco, MD, USA) containing 100 unit/ml of penicillin-streptomycin and 10% fetal bovine serum (Gibco, MD, USA). 37 ℃, 5% CO 2 It was cultured in an incubator (incubator/Thermo Scientific, IL, USA), and subculture was performed at 2-day intervals.
NO 생성 측정을 위해 RAW 264.7 세포를 2x105 cells/well로 96 웰 플레이트(well plate)에 분주하고 5시간 후에 상기 장수풍뎅이 유충 추출물을 LPS와 함께 제조방법 별 (열수 추출 여부, 효소 처리 여부, 도 5a) 또는 효소처리별,농도별(8, 40, 200, 1000ppm 도 5b)로 처리 후 37℃, 5% CO2incubator에서 24시간 배양하였다. 배양이 완료된 후 배양액 100㎕와 Griess reagent[1%(w/v) sulfanilamide in 5%(v/v) phosphoric acid와 0.1(w/v) naphtylethylenediamine-HCl] 100㎕를 혼합하여 96 웰 플레이트(well plate)에서 10분 동안 반응시킨 후 540nm에서 흡광도를 측정하였다. 표준 물질로는 NaNO2를 사용하였다.To measure NO production, RAW 264.7 cells were dispensed into a 96-well plate at 2x10 5 cells/well, and after 5 hours, the rhesus beetle larvae extract was mixed with LPS by manufacturing method (whether hot water extraction, enzyme treatment, 5a) or by enzyme treatment, by concentration (8, 40, 200, 1000 ppm Fig. 5b), and then cultured for 24 hours at 37 ℃, 5% CO 2 incubator. After incubation was completed, 100 μl of culture medium and 100 μl of Griess reagent [1% (w/v) sulfanilamide in 5% (v/v) phosphoric acid and 0.1 (w/v) naphtylethylenediamine-HCl] were mixed to form a 96-well plate (well After reacting for 10 minutes in the plate), absorbance was measured at 540 nm. NaNO 2 was used as a standard material.
NO는 NOS(NO Synthase)의 효소 촉매작용을 통해 생성되는 자유 라디칼로 혈압 조절과 신경 전달 매개체로 면역 반응에 중추적인 역할을 담당하나, 과량의 NO 생성은 염증반응을 일으킨다고 알려져 있다.NO is a free radical produced through the enzyme catalysis of NOS (NO Synthase) and plays a central role in the immune response as a mediator of blood pressure regulation and neurotransmission, but excessive production of NO is known to cause inflammatory reactions.
실험 결과 도5a 및 5b에 도시한 바와 같이, LPS로 유도한 Raw cell 264.7에서의 NO2생성은 통계적으로 유의성 있게 억제되었으며, 이를 통해 항염증 효과를 확인하였다. 후술되는 실험예 4에서의 세포 생존율 분석을 토대로 세포 독성에도 큰 영향이 없는 것으로 보아 장수풍뎅이 유충 추출물은 천연 기능성 소재로도 활용 가능할 것으로 보인다.Experimental results As shown in FIGS. 5a and 5b, NO 2 production in Raw cell 264.7 induced by LPS was statistically significantly suppressed, thereby confirming the anti-inflammatory effect. Based on the analysis of cell viability in Experimental Example 4 described later, it seems that the beetle larvae extract can be used as a natural functional material as it is seen that there is no significant effect on cytotoxicity.
실험예 4: 세포생존율 분석Experimental Example 4: Cell viability analysis
Raw 264.7 세포는 페니실린-스트렙토마이신(penicillin-streptomycin) 100 unit/ml과 10% 소태아혈청(fetal bovine serum/ Gibco, MD, USA)이 함유된 배지(Dublecco's Modified Eagle Medium/ Gibco, MD, USA)를 사용하여 37℃, 5% CO2 인큐베이터(incubator/Thermo Scientific, IL, USA)에서 배양하였으며, 2일 간격으로 계대 배양을 실시하였다.Raw 264.7 cells were cultured in medium (Dublecco's Modified Eagle Medium/ Gibco, MD, USA) containing 100 unit/ml of penicillin-streptomycin and 10% fetal bovine serum (Gibco, MD, USA). 37 ℃, 5% CO 2 It was cultured in an incubator (incubator/Thermo Scientific, IL, USA), and subculture was performed at 2-day intervals.
세포 생존율 측정을 위해 RAW 264.7 세포를 2x105 cells/well로 96 웰 플레이트(well plate)에 분주하고, 상기 장수풍뎅이 유충 추출물을 농도별(8, 40, 200, 500, 1000ppm)로 처리 후 37℃, 5% incubator에서 24시간 배양하였다. To measure cell viability, RAW 264.7 cells were dispensed into a 96-well plate at 2x10 5 cells/well, and the beetle larvae extract was treated at different concentrations (8, 40, 200, 500, 1000 ppm), followed by treatment at 37°C. , 5% Incubated for 24 hours in an incubator.
CellTiter 96® aqueousnon-radio active cell proliferation assay reagent(Promega,WI,USA)를 첨가한 후, 37℃, 5% CO2incubator에서 90분 반응시켜 마이크로 플레이트 리더기(microplate reader/ Beckman Coulter, CA, USA)를 이용하여 490 nm에서 흡광도를 측정하여 세포 생존율을 측정하였다.After adding CellTiter 96® aqueous non-radio active cell proliferation assay reagent (Promega, WI, USA), react in a 37°C, 5% CO 2 incubator for 90 minutes, using a microplate reader (Beckman Coulter, CA, USA) Cell viability was measured by measuring absorbance at 490 nm using .
그 결과, 도 6에 도시한 바와 같이, 장수풍뎅이 유충 추출물은 88.03±3.74~99.78±5.13%, 장수풍뎅이 유충 추출물은 88.45±2.65~103.71±4.68% 세포 생존율을 나타내어 8~1000 ppm 농도에서 항염증 활성 측정 시 세포 사멸에 큰 영향을 주지 않는 것으로 분석되었다.As a result, as shown in FIG. 6, the rhinoceros beetle larvae extract showed 88.03 ± 3.74 to 99.78 ± 5.13%, and the rhinoceros beetle larvae extract showed a cell viability of 88.45 ± 2.65 to 103.71 ± 4.68%, resulting in anti-inflammatory at a concentration of 8 to 1000 ppm. It was analyzed to have no significant effect on cell death when measuring activity.
실험예 5: 효소 종류 및 처리 조건에 따른 헥사날 (hexanal) 함량 측정Experimental Example 5: Measurement of hexanal content according to enzyme type and treatment conditions
곤충 유래의 이취, 비린취 성분을 저감화하여 식품의 소재로 활용하고자, 산화생성물 중 한 성분인 헥사날(Hexanal)을 가스 크로마토그램으로 분석하여 비린취가 감소하는 것을 확인하였다.In order to reduce insect-derived odor and fishy odor components and use them as food materials, Hexanal, one of the oxidation products, was analyzed with a gas chromatogram to confirm that fishy odor was reduced.
헥사날 (Hexanal)은 올레산, 리놀레산, 아라키돈산 (oleic acid, linoleic acid, arachidonic acid)와 같은 지방산의 산화와 알데하이드류의 분해에 의해 주로 생성되는 물질이며, 산화 생성물 중 하나로 이취, 비린취를 내는 성분으로 알려져 있다. 장수풍뎅이 유충의 지방함량은 생물 기준 4.7% (oleic acid 44%, linoleic acid 4%)이다.Hexanal is a substance mainly produced by the oxidation of fatty acids such as oleic acid, linoleic acid, and arachidonic acid and the decomposition of aldehydes. known as ingredients. The fat content of the rhinoceros beetle larvae was 4.7% (oleic acid 44%, linoleic acid 4%) based on living organisms.
도 7에 나타난 바와 같이 전처리(식이조절, 절식), 추출, 원심분리, 효소 처리 과정을 거치면서 헥사날(Hexanal)이 줄어들어 이취와 비린취가 감소하는 것을 확인하였다. As shown in FIG. 7, it was confirmed that off-flavor and fishy odor were reduced by reducing hexanal through pretreatment (diet control, fasting), extraction, centrifugation, and enzyme treatment.
Claims (12)
상기 장수풍뎅이 유충을 분쇄하는 단계;
상기 분쇄물을 80 내지 120℃에서 10분 내지 30분간 열수 추출하는 단계;
상기 열수 추출물을 원심분리 하는 단계; 및
원심 분리된 상등액에 단백질 분해 효소를 처리하는 단계;
를 포함하는, 식품 소재용 장수풍뎅이 유충 추출물 함유 조성물의 제조방법으로서,
상기 장수풍뎅이 유충 추출물은 헥사날(hexanal) 함량이 0mg/L 초과 0.7mg/L 이하인 것을 특징으로 하고,
상기 식이 조절은 밀가루 식이로 생육한 뒤 1 내지 72시간동안 절식하여 이루어지는 것인, 제조방법.Controlling the diet of rhinoceros beetle larvae;
crushing the rhinoceros beetle larvae;
hot water extraction of the pulverized product at 80 to 120° C. for 10 to 30 minutes;
centrifuging the hot water extract; and
Processing a proteolytic enzyme in the centrifuged supernatant;
A method for producing a composition containing rhizome larvae extract for food material, comprising:
The rhinoceros beetle larvae extract is characterized in that the hexanal content is greater than 0 mg/L and less than 0.7 mg/L,
The dietary control is made by fasting for 1 to 72 hours after growing on a flour diet.
상기 장수풍뎅이 유충 추출물은 헥사날(hexanal) 함량이 0mg/L 초과 0.7mg/L 이하인, 조성물.A composition containing an extract of beetle larvae for food material, prepared by the manufacturing method of any one of claims 1 and 3 to 8,
The rhinoceros beetle larvae extract has a hexanal content of more than 0 mg / L and less than 0.7 mg / L, composition.
A food comprising the composition of claim 10 .
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| KR102445696B1 (en) * | 2019-11-18 | 2022-09-22 | 대한민국 | Processed boiled and steamed mature silkworm with improved absorption rate and the method thereof |
| KR102407946B1 (en) * | 2020-04-23 | 2022-06-13 | 씨제이제일제당 (주) | Composition For Improving Bowel Movement Comprising Mimela splendens Larva or Extract Thereof |
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| NZ563859A (en) * | 2005-06-14 | 2010-03-26 | Meiji Dairies Corp | Off flavor (flavour) controlling method in raw milk and pasteurized milk, and pasteurized milk processed using the method |
| KR101480158B1 (en) * | 2013-01-03 | 2015-01-14 | 대한민국 | Pre-treatment method for food of allomyrina dichotoma having anti-inflammatory effect and food of allomyrina dichotoma extract having anti-inflammatory effect using the same |
| KR101533600B1 (en) | 2013-06-20 | 2015-07-10 | 대한민국 | Manufacturing Method for Allomyrina dichotoma extract having anti-Obesity effect and Anti-Obesity Composition Containing the same |
| KR101702046B1 (en) * | 2015-06-23 | 2017-02-03 | 대한민국(농촌진흥청장) | Composition for protecting liver or for anticancer comprising allomyrina dichotoma larva as effective component |
| KR20170014448A (en) * | 2015-07-30 | 2017-02-08 | (주) 에이티바이오 | Manufacturing method of feed composition using insects and that of using feed composition |
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| 국립농업과학원. ‘귀뚜라미의 식품원료 인증을 위한 기반연구’에 대한 최종보고서(2016.06.25.)* |
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| Publication number | Publication date |
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| WO2019088774A3 (en) | 2019-06-27 |
| WO2019088774A2 (en) | 2019-05-09 |
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