KR102559904B1 - Manufacturing method of oxidized saccharide with antioxidant activity - Google Patents
Manufacturing method of oxidized saccharide with antioxidant activity Download PDFInfo
- Publication number
- KR102559904B1 KR102559904B1 KR1020200178108A KR20200178108A KR102559904B1 KR 102559904 B1 KR102559904 B1 KR 102559904B1 KR 1020200178108 A KR1020200178108 A KR 1020200178108A KR 20200178108 A KR20200178108 A KR 20200178108A KR 102559904 B1 KR102559904 B1 KR 102559904B1
- Authority
- KR
- South Korea
- Prior art keywords
- saccharide
- glucose
- weight
- saccharide composition
- oxidized
- Prior art date
Links
- 150000001720 carbohydrates Chemical class 0.000 title claims abstract description 264
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 9
- 230000003078 antioxidant effect Effects 0.000 title abstract description 42
- 239000000203 mixture Substances 0.000 claims abstract description 162
- 238000007254 oxidation reaction Methods 0.000 claims abstract description 78
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims abstract description 66
- 239000008103 glucose Substances 0.000 claims abstract description 65
- 102000004316 Oxidoreductases Human genes 0.000 claims abstract description 31
- 108090000854 Oxidoreductases Proteins 0.000 claims abstract description 31
- DLRVVLDZNNYCBX-RTPHMHGBSA-N isomaltose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1OC[C@@H]1[C@@H](O)[C@H](O)[C@@H](O)C(O)O1 DLRVVLDZNNYCBX-RTPHMHGBSA-N 0.000 claims abstract description 23
- 238000000034 method Methods 0.000 claims abstract description 22
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- 239000007787 solid Substances 0.000 claims description 39
- 230000003647 oxidation Effects 0.000 claims description 30
- 230000002292 Radical scavenging effect Effects 0.000 claims description 28
- MGJZITXUQXWAKY-UHFFFAOYSA-N diphenyl-(2,4,6-trinitrophenyl)iminoazanium Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1N=[N+](C=1C=CC=CC=1)C1=CC=CC=C1 MGJZITXUQXWAKY-UHFFFAOYSA-N 0.000 claims description 28
- 238000006243 chemical reaction Methods 0.000 claims description 25
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- 108010015776 Glucose oxidase Proteins 0.000 claims description 22
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- 235000019420 glucose oxidase Nutrition 0.000 claims description 22
- 102000016938 Catalase Human genes 0.000 claims description 20
- 108010053835 Catalase Proteins 0.000 claims description 20
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- 238000007865 diluting Methods 0.000 description 3
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- 125000003172 aldehyde group Chemical group 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 239000003963 antioxidant agent Substances 0.000 description 2
- 235000019606 astringent taste Nutrition 0.000 description 2
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 2
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/16—Preparation of compounds containing saccharide radicals produced by the action of an alpha-1, 6-glucosidase, e.g. amylose, debranched amylopectin
-
- 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
- A23L33/125—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives containing carbohydrate syrups; containing sugars; containing sugar alcohols; containing starch hydrolysates
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/60—Sugars; Derivatives thereof
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q17/00—Barrier preparations; Preparations brought into direct contact with the skin for affording protection against external influences, e.g. sunlight, X-rays or other harmful rays, corrosive materials, bacteria or insect stings
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
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- 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
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- 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
- A23V2200/00—Function of food ingredients
- A23V2200/30—Foods, ingredients or supplements having a functional effect on health
- A23V2200/302—Foods, ingredients or supplements having a functional effect on health having a modulating effect on age
Abstract
본 발명의 일 예는 당류 조성물 용액에 당류 산화효소를 첨가하고 산화반응시켜 당류 조성물을 구성하는 적어도 일부 당류를 산화당류로 전환시키는 단계를 포함하는 산화당류 조성물의 제조방법을 제공한다. 상기 당류 조성물은 당류 조성물 내의 당류 전체 중량을 기준으로 포도당 15~35 중량% 및 이소말토올리고당 20~60 중량%를 포함한다. 본 발명의 제조방법에 의해 제조되는 산화당류 조성물은 소정의 산미를 가지며 우수한 항산화 활성을 보인다. 본 발명에 따른 산화당류 조성물은 식품 소재로 사용되는 경우 식품에 산미와 항산화와 같은 다기능을 부여할 수 있고 화장품 소재로 사용되는 경우 화장품에 보습과 항산화와 같은 다기능을 부여할 수 있다. 따라서, 본 발명의 제조방법에 의해 제조되는 산화당류 조성물은 식품 산업 및 화장품 산업에서 다기능성을 부여할 수 있는 고부가가치 물질로 이용될 수 있다.One embodiment of the present invention provides a method for preparing an oxidized saccharide composition comprising the step of adding a saccharide oxidase to a solution of the saccharide composition and performing an oxidation reaction to convert at least some of the saccharides constituting the saccharide composition into oxidized saccharides. The saccharide composition includes 15 to 35% by weight of glucose and 20 to 60% by weight of isomaltooligosaccharide based on the total weight of sugars in the saccharide composition. The oxidized saccharide composition prepared by the production method of the present invention has a certain acidity and exhibits excellent antioxidant activity. When the oxidized saccharide composition according to the present invention is used as a food material, it can impart multi-functions such as acidity and anti-oxidation to foods, and when used as a cosmetic material, it can impart multi-functions such as moisturizing and anti-oxidation to cosmetics. Therefore, the oxidized saccharide composition prepared by the production method of the present invention can be used as a high value-added material capable of imparting multifunctionality in the food and cosmetic industries.
Description
본 발명은 산화당류 조성물의 제조방법에 관한 것으로서, 더 상세하게는 원료 당류 및 산화효소의 조합에 의해 항산화 활성이 크게 증가된 산화당류 조성물의 제조방법에 관한 것이다.The present invention relates to a method for preparing an oxidized saccharide composition, and more particularly, to a method for preparing an oxidized saccharide composition whose antioxidant activity is greatly increased by a combination of a raw material saccharide and an oxidase.
당류는 일반적으로 항산화 활성이 낮은 물질이다. Hu 등[Hu et al., In vitro evaluation of the antioxidant activities of carbohydrates, Bioactive Carbohydratesand Dietary Fibre 7 (2016) 19-27]에 따르면 단당 및 올리고당은 약 2,000 ppm의 농도에서도 약 10% 미만의 항산화 활성을 보인다.Sugars are generally substances with low antioxidant activity. According to Hu et al. [Hu et al., In vitro evaluation of the antioxidant activities of carbohydrates, Bioactive Carbohydrates and Dietary Fiber 7 (2016) 19-27], monosaccharides and oligosaccharides show less than about 10% antioxidant activity even at a concentration of about 2,000 ppm.
당류는 식품 소재나 화장품 소재로 널리 사용되는데, 당류에 높은 항산화 활성이 부여되는 경우 이의 적용 분야는 훨씬 더 다양해질 수 있다. 항산화 활성을 가지는 대표적인 산화 당류로는 말토비온산(Maltobionic acid, MBA)가 있다. 말토비온산은 말토스(맥아당)를 브롬수로 산화시켜 얻는 물질로서 다음과 같은 화학 구조를 갖는다. 미국 공개특허공보 제2016-0081387호에는 전분을 효소 반응을 통해 말토스로 전환하는 단계; 및 말토스를 효소 반응을 통해 말토비오산으로 전환하는 단계를 포함하는 말토비온산 제조방법과 함께 말토비온산이 항산화 효과를 가지는 식품 첨가제로 사용될 수 있다는 점이 개시되어 있다.Sugars are widely used as food materials or cosmetic materials, and when high antioxidant activity is given to sugars, their application fields can be much more diverse. A typical oxidized saccharide having antioxidant activity is maltobionic acid (MBA). Maltobionic acid is a substance obtained by oxidizing maltose (malt sugar) with bromine water and has the following chemical structure. U.S. Patent Publication No. 2016-0081387 includes converting starch into maltose through an enzymatic reaction; and a method for producing maltobionic acid including converting maltose into maltobionic acid through an enzymatic reaction, and that maltobionic acid can be used as a food additive having an antioxidant effect.
[말토비온산의 화학 구조][Chemical structure of maltobionic acid]
일반적으로 대부분의 당류는 산화되면 말단의 알데히드기가 카르복실기로 되어 카르복실산 형태의 산화당류로 전환된다. 다만, 다양하게 존재하는 산화당류의 항산화 활성에 대해서는 알려진바가 거의 없다.In general, when most saccharides are oxidized, the terminal aldehyde group becomes a carboxyl group and is converted into oxidized saccharide in the form of carboxylic acid. However, little is known about the antioxidant activity of various oxidized saccharides.
본 발명은 종래의 기술적 배경하에서 도출된 것으로서, 본 발명의 목적은 항산화 활성이 우수한 산화당류 조성물의 제조방법을 제공하는데에 있다.The present invention was derived under the conventional technical background, and an object of the present invention is to provide a method for preparing an oxidized saccharide composition having excellent antioxidant activity.
본 발명의 발명자들은 다양한 당류 제품을 조합하여 당류 조성물을 제조하고, 이를 당류 산화효소로 산화반응시켜 다양한 산화당류를 제조한 후, 산화당류의 항산화 활성을 DPPH 라디칼 소거 활성으로 평가한 결과, 산화당류가 높은 항산화 활성을 가지기 위해서는 산화반응의 기질이 되는 당류 조성물 내 포도당 함량 및 이소말토올리고당 함량이 가장 중요한 요소이고, 당류 조성물 내 중합도가 1 내지 4인 당류 함량이 그 다음으로 중요한 요소라는 점을 확인하고 본 발명을 완성하였다.The inventors of the present invention prepared saccharide compositions by combining various saccharide products, produced various oxidized saccharides by oxidizing them with saccharide oxidase, and evaluated the antioxidant activity of oxidized saccharides by DPPH radical scavenging activity. confirmed and completed the present invention.
상기 목적을 해결하기 위하여, 본 발명의 일 예는 당류 조성물 용액에 당류 산화효소를 첨가하고 산화반응시켜 당류 조성물을 구성하는 적어도 일부 당류를 산화당류로 전환시키는 단계를 포함하는 산화당류 조성물의 제조방법을 제공한다. 상기 당류 조성물은 당류 조성물 내의 당류 전체 중량을 기준으로 포도당 15~35 중량% 및 이소말토올리고당 20~60 중량%를 포함한다.In order to solve the above object, one embodiment of the present invention provides a method for preparing an oxidized saccharide composition comprising the step of converting at least some of the saccharides constituting the saccharide composition into oxidized saccharides by adding a saccharide oxidase to a solution of the saccharide composition and performing an oxidation reaction. The saccharide composition includes 15 to 35% by weight of glucose and 20 to 60% by weight of isomaltooligosaccharide based on the total weight of sugars in the saccharide composition.
본 발명의 제조방법에 의해 제조되는 산화당류 조성물은 소정의 산미를 가지며 우수한 항산화 활성을 보인다. 본 발명에 따른 산화당류 조성물은 식품 소재로 사용되는 경우 식품에 산미와 항산화와 같은 다기능을 부여할 수 있고 화장품 소재로 사용되는 경우 화장품에 보습과 항산화와 같은 다기능을 부여할 수 있다. 따라서, 본 발명의 제조방법에 의해 제조되는 산화당류 조성물은 식품 산업 및 화장품 산업에서 다기능성을 부여할 수 있는 고부가가치 물질로 이용될 수 있다.The oxidized saccharide composition prepared by the production method of the present invention has a certain acidity and exhibits excellent antioxidant activity. When the oxidized saccharide composition according to the present invention is used as a food material, it can impart multi-functions such as acidity and anti-oxidation to foods, and when used as a cosmetic material, it can impart multi-functions such as moisturizing and anti-oxidation to cosmetics. Therefore, the oxidized saccharide composition prepared by the production method of the present invention can be used as a high value-added material capable of imparting multifunctionality in the food and cosmetic industries.
도 1은 본 발명의 실시예에서 산화당류6의 항산화 활성에 영향을 미치는 유효성분을 탐색하기 위해 제2 조합에 의해 제조한 다양한 시료들의 DPPH 라디칼 소거 활성을 나타낸 그래프이다.
도 2는 본 발명의 실시예에서 산화당류6의 항산화 활성에 영향을 미치는 유효성분을 탐색하기 위해 제3 조합에 의해 제조한 다양한 시료들의 DPPH 라디칼 소거 활성을 나타낸 그래프이다.
도 3은 본 발명의 실시예에서 산화당류6의 성분 특성을 규명하기 위해 당류 조성물6와 산화당류6의 성분을 LC-TOF/MS를 이용하여 분석한 결과이다.Figure 1 is a graph showing the DPPH radical scavenging activity of various samples prepared by the second combination to search for active ingredients that affect the antioxidant activity of oxidized saccharide 6 in Examples of the present invention.
Figure 2 is a graph showing the DPPH radical scavenging activity of various samples prepared by the third combination to search for active ingredients that affect the antioxidant activity of oxidized saccharide 6 in Examples of the present invention.
3 is a result of analyzing the saccharide composition 6 and the components of oxidized saccharide 6 using LC-TOF/MS to identify the component characteristics of oxidized saccharide 6 in Examples of the present invention.
이하, 본 발명을 구체적으로 설명한다.Hereinafter, the present invention will be described in detail.
본 발명에서 사용되는 용어인 "이소말토올리고당(Isomaltooligosaccharide, IMO)"은 소화 저항성을 가질 수 있는 단사슬 탄수화물의 혼합물로서, 단위구조인 글루코스의 중합도가 2~10 이고 글루코스들이 적어도 1개의 1,6-α-글리코시드 결합으로 연결되어 있다. 이소말토올리고당을 구성하는 당들로는 이소말토스, 이소말토트리오스, 판노스(Panose), 이소말토테트라오스, 이소말토펜타오스, 니게로스(Nigerose), 코지비오스(Kojibiose), 고도 분지 올리고당 등이 있다.As used herein, the term "isomaltooligosaccharide (IMO)" is a mixture of short-chain carbohydrates that can have digestion resistance, and the degree of polymerization of glucose as a unit structure is 2 to 10, and the glucose is linked by at least one 1,6-α-glycosidic bond. Saccharides constituting isomalto-oligosaccharide include isomaltose, isomaltotriose, pannose, isomaltotetraose, isomaltopentaose, nigerose, kojibiose, and highly branched oligosaccharides.
본 발명에서 사용되는 용어인 "말토올리고당(Maltooligosaccharide"은 단위구조인 글루코스의 중합도가 2~10 이고 글루코스들이 모두 1,4-α-글리코시드 결합으로 연결되어 있는 올리고당을 의미한다. 말토올리고당을 구성하는 당들로는 말토스, 말토트리오스, 말토테트라오스, 말토펜타오스, 말토헥사오스, 말토헵타오스 등이 있다.As used herein, the term "maltooligosaccharide" refers to an oligosaccharide in which glucose as a unit structure has a degree of polymerization of 2 to 10 and all glucoses are linked by 1,4-α-glycosidic bonds. Saccharides constituting maltooligosaccharide include maltose, maltotriose, maltotetraose, maltopentaose, maltohexaose, and maltoheptaose.
본 발명에서 사용되는 용어인 "당류 산화효소"는 당의 말단 할데히드기가 산화되어 카르복실기로 전환하는 산화반응에 대해 촉매 작용을 하는 효소로서, 산화 효소 뿐만 아니라 산화효소 및 이의 산화작용을 보조하는 효소의 조합을 포함한다. 공지의 당류 산화효소로는 글루코스옥시다제(Glucose oxidase; CAS No. 9001-37-0), 카보하이드레이트옥시다제(Carbohydrate oxidase; CAS No. 372580-9), 헥소스옥시다제(Hexose oxidase; CAS No. 9028-75-5) 등이 있다. 상기 글루코스옥시다제(Glucose oxidase; CAS No. 9001-37-0), 카보하이드레이트옥시다제(Carbohydrate oxidase; CAS No. 372580-9) 및 헥소스옥시다제(Hexose oxidase; CAS No. 9028-75-5)는 공통적으로 D-글루코스가 D-글루코노-델타-락톤(D-glucono-δ-lactone, GDL) 및 과산화수소로 산화되는 반응을 촉매하는 산화-환원 효소(oxido-reductase)이다. 상기 카보하이드레이트옥시다제(Carbohydrate oxidase; CAS No. 372580-9)는 Polyporus obtusus에서 분리된 효소이다. 또한, 본 발명에서 당류 산화효소는 글루코스옥시다제(Glucose oxidase; CAS No. 9001-37-0), 카보하이드레이트옥시다제(Carbohydrate oxidase; CAS No. 372580-9) 및 헥소스옥시다제(Hexose oxidase; CAS No. 9028-75-5)에 선택되는 1종 이상의 효소와 카탈라제(Catalase; CAS No. 9001-05-2)의 조합을 포함한다. 상기 카탈라제(Catalase; CAS No. 9001-05-2)는 과산화수소를 물과 산소로 분해하는 반응을 촉매하는 효소이다.As used herein, the term "saccharide oxidase" is an enzyme that catalyzes an oxidation reaction in which a terminal haldehyde group of a sugar is oxidized and converted to a carboxyl group, and includes an oxidase as well as an oxidase and a combination of enzymes assisting in its oxidation. Known saccharide oxidases include glucose oxidase (CAS No. 9001-37-0), carbohydrate oxidase (CAS No. 372580-9), and hexose oxidase (CAS No. 9028-75-5). The glucose oxidase (CAS No. 9001-37-0), carbohydrate oxidase (CAS No. 372580-9) and hexose oxidase (CAS No. 9028-75-5) are commonly D-glucose D-glucono-delta-lactone (D-glucono-δ-lactone, GDL) and It is an oxido-reductase that catalyzes the oxidation reaction to hydrogen oxide. The carbohydrate oxidase (CAS No. 372580-9) is an enzyme isolated from Polyporus obtusus. In addition, in the present invention, the saccharide oxidase is glucose oxidase (CAS No. 9001-37-0), carbohydrate oxidase (CAS No. 372580-9) and hexose oxidase (CAS No. 9028-75-5) and at least one enzyme selected from catalase (Catalase; CAS No. 900). 1-05-2). The catalase (CAS No. 9001-05-2) is an enzyme that catalyzes a reaction of decomposing hydrogen peroxide into water and oxygen.
본 발명은 항산화 활성이 우수한 산화당류 조성물의 제조방법에 관한 것이다.The present invention relates to a method for preparing an oxidized saccharide composition having excellent antioxidant activity.
본 발명의 일 예에 따른 산화당류 조성물의 제조방법은 당류 조성물 용액에 당류 산화효소를 첨가하고 산화반응시켜 당류 조성물을 구성하는 적어도 일부 당류를 산화당류로 전환시키는 단계를 포함한다. 이하, 본 발명의 일 예에 따른 산화당류 조성물의 제조방법을 구성요소별로 나누어 설명한다.A method for preparing an oxidized saccharide composition according to an embodiment of the present invention includes adding a saccharide oxidase to a saccharide composition solution and performing an oxidation reaction to convert at least some of the saccharides constituting the saccharide composition into oxidized saccharides. Hereinafter, a method for preparing an oxidized saccharide composition according to an embodiment of the present invention will be described separately for each component.
당류 조성물sugar composition
본 발명에서 당류 조성물은 효소 산화반응의 기질로 사용된다. 상기 당류 조성물을 구성하는 적어도 일부 당류(예를 들어, 단당류, 이당류, 삼당류, 사당류 등)는 효소 산화반응에 의해 산화당류로 전환된다.In the present invention, a saccharide composition is used as a substrate for an enzymatic oxidation reaction. At least some saccharides (eg, monosaccharides, disaccharides, trisaccharides, tetrasaccharides, etc.) constituting the saccharide composition are converted into oxidized saccharides by an enzymatic oxidation reaction.
상기 당류 조성물은 당류 조성물 내의 당류 전체 중량을 기준으로 포도당 15~35 중량% 및 이소말토올리고당 20~60 중량%를 포함하고, 바람직하게는 포도당 18.5~28.5 중량% 및 이소말토올리고당 22~45 중량%를 포함한다.The saccharide composition includes 15 to 35% by weight of glucose and 20 to 60% by weight of isomaltooligosaccharide, preferably 18.5 to 28.5% by weight of glucose and 22 to 45% by weight of isomaltooligosaccharide based on the total weight of saccharides in the saccharide composition.
또한, 상기 당류 조성물은 당류 조성물 내의 당류 전체 중량을 기준으로 바람직하게는 포도당 15~35 중량%, 이소말토올리고당 20~60 중량% 및 말토올리고당 20~60 중량%를 포함하고, 더 바람직하게는 포도당 18.5~28.5 중량%, 이소말토올리고당 22~45 중량% 및 이소말토올리고당 20~45 중량%를 포함한다.In addition, the saccharide composition preferably includes 15 to 35% by weight of glucose, 20 to 60% by weight of isomaltooligosaccharide, and 20 to 60% by weight of maltooligosaccharide, based on the total weight of saccharides in the saccharide composition.
또한, 상기 당류 조성물 내에서 글루코스를 단위구조로 가지고 중합도가 2~4인 올리고당의 함량은 당류 전체 중량을 기준으로 바람직하게는 55~80 중량%이고, 더 바람직하게는 62~78 중량%이다.In addition, the content of oligosaccharides having glucose as a unit structure and a degree of polymerization of 2 to 4 in the saccharide composition is preferably 55 to 80% by weight, more preferably 62 to 78% by weight, based on the total weight of the saccharide.
또한, 상기 당류 조성물 내에서 포도당 및 글루코스를 단위구조로 가지고 중합도가 2~4인 올리고당을 합한 함량은 당류 전체 중량을 기준으로 바람직하게는 80~98 중량%이고, 더 바람직하게는 85~96 중량%이다.In addition, the content of glucose and oligosaccharides having glucose as a unit structure and a degree of polymerization of 2 to 4 in the saccharide composition is preferably 80 to 98% by weight, more preferably 85 to 96% by weight, based on the total weight of the saccharide.
또한, 상기 당류 조성물 내에서 포도당 및 글루코스를 단위구조로 가지고 중합도가 2~10인 올리고당을 합한 함량은 당류 전체 중량을 기준으로 바람직하게는 95~99.9 중량%이고, 96~99.8 중량%이다.In addition, the content of glucose and oligosaccharides having glucose as a unit structure and a degree of polymerization of 2 to 10 in the saccharide composition is preferably 95 to 99.9% by weight and 96 to 99.8% by weight based on the total weight of the saccharide.
또한, 상기 당류 조성물은 당류 조성물 내의 당류 전체 중량을 기준으로 바람직하게는 포도당 16~30 중량%, 글루코스를 단위구조로 가지는 이당류 18.5~32.5 중량%, 글루코스를 단위구조로 가지는 삼당류 23~37 중량%, 글루코스를 단위구조로 가지는 사당류 8.5~18.5 중량%, 글루코스를 단위구조로 가지는 오당류 1.5~7.5 중량%, 글루코스를 단위구조로 가지는 육당류 0.5~3 중량% 및 글루코스를 단위구조로 가지는 칠당류 0.2~2 중량%를 포함하고, 더 바람직하게는 포도당 18.5~28.5 중량%, 글루코스를 단위구조로 가지는 이당류 20.5~30.5 중량%, 글루코스를 단위구조로 가지는 삼당류 25~35 중량%, 글루코스를 단위구조로 가지는 사당류 10~17 중량%, 글루코스를 단위구조로 가지는 오당류 2.5~6.5 중량%, 글루코스를 단위구조로 가지는 육당류 0.8~2.2 중량% 및 글루코스를 단위구조로 가지는 칠당류 0.3~1.5 중량%를 포함한다.In addition, the saccharide composition preferably contains 16 to 30% by weight of glucose, 18.5 to 32.5% by weight of disaccharides having glucose as a unit structure, 23 to 37% by weight of trisaccharides having glucose as a unit structure, 8.5 to 18.5% by weight of tetrasaccharides having glucose as a unit structure, 1.5 to 7.5% by weight of pentasaccharides having glucose as a unit structure, based on the total weight of saccharides in the saccharide composition. 0.5 to 3% by weight of hexasaccharides having glucose as a unit structure and 0.2 to 2% by weight of hepsaccharides having glucose as a unit structure, more preferably 18.5 to 28.5% by weight of glucose, 20.5 to 30.5% by weight of disaccharides having glucose as a unit structure, 25 to 35% by weight of trisaccharides having glucose as a unit structure, 10 to 17% by weight of tetrasaccharides having glucose as a unit structure, It contains 2.5 to 6.5% by weight of pentasaccharides having glucose as a unit structure, 0.8 to 2.2% by weight of hexasaccharides having glucose as a unit structure, and 0.3 to 1.5% by weight of hepsaccharides having glucose as a unit structure.
상기 당류 조성물은 상업적인 당류 제품들 또는 이들의 조합 등에서 선택될 수 있다.The saccharide composition may be selected from commercial saccharide products or combinations thereof.
당류 산화효소sugar oxidase
본 발명에서 당류 산화효소는 당류 조성물을 구성하는 적어도 일부 당류(예를 들어, 단당류, 이당류, 삼당류, 사당류 등)의 산화 반응을 촉매하는 효소로서, 공지의 다양한 당류 산화효소에서 선택될 수 있다. 상기 당류 산화효소는 바람직하게는 글루코스옥시다제(Glucose oxidase; CAS No. 9001-37-0), 카보하이드레이트옥시다제(Carbohydrate oxidase; CAS No. 372580-9) 또는 헥소스옥시다제(Hexose oxidase; CAS No. 9028-75-5)에서 선택되는 1종 이상으로 구성될 수 있고 산화당류 조성물의 항산화 활성, 효소 반응의 경제성 등을 고려할 때 글루코스옥시다제(Glucose oxidase; CAS No. 9001-37-0)이다.In the present invention, the saccharide oxidase is an enzyme that catalyzes an oxidation reaction of at least some saccharides (eg, monosaccharide, disaccharide, trisaccharide, tetrasaccharide, etc.) constituting the saccharide composition, and may be selected from various known saccharide oxidases. The saccharide oxidase is preferably composed of at least one selected from glucose oxidase (CAS No. 9001-37-0), carbohydrate oxidase (CAS No. 372580-9) or hexose oxidase (CAS No. 9028-75-5), and the antioxidant activity of the oxidized saccharide composition, the economics of the enzyme reaction, etc. When considered, it is glucose oxidase (CAS No. 9001-37-0).
또한, 상기 당류 산화효소는 산화당류 조성물의 항산화 활성, 효소 반응의 경제성 등을 고려할 때 글루코스옥시다제(Glucose oxidase; CAS No. 9001-37-0) 및 카탈라제(Catalase; CAS No. 9001-05-2)의 조합으로 구성되거나, 글루코스옥시다제(Glucose oxidase; CAS No. 9001-37-0), 카보하이드레이트옥시다제(Carbohydrate oxidase; CAS No. 372580-9) 및 카탈라제(Catalase; CAS No. 9001-05-2)의 조합으로 구성될 수 있다.In addition, the saccharide oxidase is composed of a combination of glucose oxidase (CAS No. 9001-37-0) and catalase (CAS No. 9001-05-2), glucose oxidase (CAS No. 9001-37-0), or carbohydrate oxidase (Carbo It can be composed of a combination of hydrate oxidase; CAS No. 372580-9) and catalase (CAS No. 9001-05-2).
효소 산화반응 조건Enzymatic Oxidation Conditions
본 발명에서 효소 산화반응 조건은 당류 조성물 용액의 고형분 농도, 당류 산화효소의 첨가량, 산화반응 온도, 산화반응 pH, 산화반응 시간, 당류 조성물 용액의 교반 속도, 에어레이션(aeration) 속도 등과 같이 다양한 파라미터들로 구성된다.In the present invention, the enzyme oxidation reaction conditions are composed of various parameters such as the solid content concentration of the saccharide composition solution, the amount of saccharide oxidase added, the oxidation reaction temperature, the oxidation reaction pH, the oxidation reaction time, the stirring speed of the saccharide composition solution, and the aeration rate.
상기 당류 조성물 용액의 고형분 농도는 크게 제한되지 않으나 효소 산화반응 효율, 산화반응 속도의 제어, 작업성 등을 고려할 때 바람직하게는 6~50 브릭스(Brix)이고, 더 바람직하게는 10~45 브릭스(Brix)이다.The solid content concentration of the saccharide composition solution is not particularly limited, but is preferably 6 to 50 Brix, more preferably 10 to 45 Brix, in consideration of enzyme oxidation reaction efficiency, oxidation reaction rate control, and workability.
또한, 상기 당류 산화효소의 첨가량은 크게 제한되지 않으나 효소 산화반응 효율, 산화반응 속도의 제어, 경제성 등을 고려할 때 바람직하게는 당류 조성물 고형분 중량 대비 0.05~1.0 %(v/w)이고, 더 바람직하게는 당류 조성물 고형분 중량 대비 0.1~0.6 %(v/w)이다. 예를 들어, 당류 산화효소로 글루코스옥시다제(Glucose oxidase; CAS No. 9001-37-0)를 사용하는 경우 당류 산화효소의 첨가량은 당류 조성물 고형분 중량 대비 0.1~0.4 %(v/w)에서 선택될 수 있다. 또한, 당류 산화효소로 글루코스옥시다제(Glucose oxidase; CAS No. 9001-37-0) 및 카탈라제(Catalase; CAS No. 9001-05-2)의 조합을 사용하는 경우 글루코스옥시다제(Glucose oxidase; CAS No. 9001-37-0) 및 카탈라제(Catalase; CAS No. 9001-05-2)의 첨가량은 각각 당류 조성물 고형분 중량 대비 0.1~0.4 %(v/w) 및 0.05~0.2%(v/w)에서 선택될 수 있다.In addition, the addition amount of the saccharide oxidase is not particularly limited, but is preferably 0.05 to 1.0% (v/w) based on the solid content of the saccharide composition, and more preferably 0.1 to 0.6% (v/w) based on the solid content of the saccharide composition, in consideration of enzyme oxidation reaction efficiency, control of the oxidation reaction rate, economic feasibility, and the like. For example, when glucose oxidase (CAS No. 9001-37-0) is used as the saccharide oxidase, the added amount of the saccharide oxidase may be selected from 0.1 to 0.4% (v/w) based on the solid content of the saccharide composition. In addition, when a combination of glucose oxidase (CAS No. 9001-37-0) and catalase (CAS No. 9001-05-2) is used as a saccharide oxidase, the added amount of glucose oxidase (CAS No. 9001-37-0) and catalase (CAS No. 9001-05-2) It may be selected from 0.1 to 0.4% (v/w) and 0.05 to 0.2% (v/w) based on the solid content of the saccharide composition, respectively.
또한, 상기 산화반응 온도는 사용하는 당류 산화효소의 종류나 최적 온도에 따라 다양한 범위에서 선택될 수 있고, 바람직하게는 25~50℃이고, 더 바람직하게는 28~48℃이다.In addition, the oxidation reaction temperature may be selected from a variety of ranges depending on the type of saccharide oxidase used or the optimum temperature, preferably 25 to 50 ° C, more preferably 28 to 48 ° C.
또한, 상기 산화반응 pH는 당류 산화효소의 종류나 최적 pH에 따라 다양한 범위에서 선택될 수 있고, 바람직하게는 5.0~8.0이고, 더 바람직하게는 5.5~7.5이다.In addition, the pH of the oxidation reaction may be selected from various ranges depending on the type of saccharide oxidase or the optimum pH, preferably 5.0 to 8.0, and more preferably 5.5 to 7.5.
또한, 상기 산화반응 시간은 당류 조성물 용액의 고형분 농도, 당류 산화효소의 종류, 산화반응 온도, 산화반응 pH 에 따라 다양한 범위에서 선택될 수 있고, 산화당류 조성물의 항산화 활성, 경제성 등을 고려할 때 바람직하게는 30분 내지 24 hr이고, 더 바람직하게는 1~6 hr이다.In addition, the oxidation reaction time may be selected from a wide range depending on the solid content concentration of the saccharide composition solution, the type of saccharide oxidase, the oxidation reaction temperature, and the oxidation reaction pH, and is preferably 30 minutes to 24 hr, more preferably 1 to 6 hr, considering the antioxidant activity and economic efficiency of the oxidized saccharide composition.
또한, 상기 당류 조성물 용액의 교반 속도는 균일한 효소 산화반응을 위해 50~450 rpm에서 선택될 수 있고, 바람직하게는 100~350 rpm에서 선택될 수 있다.In addition, the stirring speed of the saccharide composition solution may be selected from 50 to 450 rpm for a uniform enzymatic oxidation reaction, and preferably from 100 to 350 rpm.
또한, 상기 에어레이션(aeration) 속도는 다양한 산화반응 조건들에 의해 요구되는 공기(산소)의 공급량이 달라지기 때문에 다양한 범위에서 선택될 수 있고, 효소 산화반응 효율 내지 속도 제어 등을 고려할 때 바람직하게는 0.5~5 vvm이고, 더 바람직하게는 0.8~2 vvm이다.In addition, the aeration rate can be selected in a wide range because the amount of air (oxygen) supplied varies according to various oxidation reaction conditions, and is preferably 0.5 to 5 vvm, more preferably 0.8 to 2 vvm, considering enzyme oxidation reaction efficiency or rate control.
본 발명의 방법에 의해 제조되는 산화당류 조성물은 기질로 사용되는 당류 조성물의 항산화 활성과 비교할 때 약 2.5배 내지 7배 정도 높은 항산화 활성을 가진다. 상기 항산화 활성은 공지의 다양한 방법에 의해 평가될 수 있으며, 예를 들어 DPPH 라디칼 소거 활성에 의해 평가될 수 있다. 본 발명의 방법에 의해 제조되는 산화당류 조성물의 DPPH 라디칼 소거 활성은 고형분 농도가 40 브릭스(Brix)인 용액으로 측정할 때 바람직하게는 75~92%일 수 있고, 더 바람직하게는 80~90%일 수 있다.The oxidized saccharide composition prepared by the method of the present invention has antioxidant activity about 2.5 to 7 times higher than that of the saccharide composition used as a substrate. The antioxidant activity can be evaluated by various methods known in the art, for example, it can be evaluated by DPPH radical scavenging activity. The DPPH radical scavenging activity of the oxidized saccharide composition prepared by the method of the present invention may be preferably 75 to 92%, more preferably 80 to 90% when measured in a solution having a solid concentration of 40 Brix.
이하, 본 발명을 실시예를 통하여 보다 구체적으로 설명한다. 다만, 하기 실시예는 본 발명의 기술적 특징을 명확하게 예시하기 위한 것일 뿐, 본 발명의 보호범위를 한정하는 것은 아니다.Hereinafter, the present invention will be described in more detail through examples. However, the following examples are only for clearly illustrating the technical features of the present invention, but do not limit the protection scope of the present invention.
1. 다양한 당류 조성물의 산화를 통한 산화당류의 제조 및 항산화 활성 평가1. Preparation of oxidized saccharides through oxidation of various saccharide compositions and evaluation of antioxidant activity
본 발명의 발명자는 본 발명의 출원인인 대상주식회사가 상업적으로 시판하는 당류 제품을 이용하여 다양한 당 성분을 함유하는 당류 조성물을 제조하고, 제조한 당류 조성물을 효소 산화반응시켜 산화당류를 제조하고, 제조된 산화당류의 DPPH 라디칼 소거 활성을 측정하여 항산화 활성을 평가하였다.The inventor of the present invention prepared a saccharide composition containing various sugar components using saccharide products commercially available from Daesang Co., Ltd., the applicant of the present invention, enzymatically oxidized the prepared saccharide composition to prepare oxidized saccharides, and evaluated the antioxidant activity by measuring the DPPH radical scavenging activity of the prepared oxidized saccharides.
산화당류를 제조하기 위해 사용한 당류 조성물의 당 성분 및 함량을 HPLC를 이용하여 분석하였고, 하기 표 1에 그 결과를 정리하였다.The sugar component and content of the saccharide composition used to prepare the oxidized saccharide were analyzed using HPLC, and the results are summarized in Table 1 below.
* DP2 : 글루코스를 단위구조로 하고 중합도가 2인 당* DP2: Glucose as unit structure and polymerization degree is 2 per person
* DP3 : 글루코스를 단위구조로 하고 중합도가 3인 당* DP3: sugar with glucose as unit structure and degree of polymerization of 3
* DP8+ : 글루코스를 단위구조로 하고 중합도가 8 이상인 당* DP8+: A sugar with glucose as a unit structure and a degree of polymerization of 8 or higher
* 상기 표 1에서 당류 조성물 2, 당류 조성물3, 당류 조성물4, 당류 조성물5에 존재하는 당 성분은 단위구조인 글루코스들이 1,4-α-글리코시드 결합으로 연결된 것임* In Table 1, sugar components present in saccharide composition 2, saccharide composition 3, saccharide composition 4, and saccharide composition 5 are units of glucose connected by 1,4-α-glycosidic bonds.
* 당류 조성물 1 : 중합도 3 이상의 말토올리고당 및 중합도 3 이상의 이소말토올리고당을 포함하지 않음* Saccharide composition 1: does not contain maltooligosaccharide having a degree of polymerization of 3 or higher and isomaltooligosaccharide having a degree of polymerization of 3 or higher
* 당류 조성물 2 : DP2에 해당하는 당은 말토스이고, 중합도 3 이상의 말토올리고당 함량은 약 5.7%이며, 중합도 3 이상의 이소말토올리고당을 포함하지 않음* Saccharide composition 2: The sugar corresponding to DP2 is maltose, the content of maltooligosaccharide having a polymerization degree of 3 or higher is about 5.7%, and isomaltooligosaccharide having a polymerization degree of 3 or higher is not included.
* 당류 조성물 3 : DP2에 해당하는 당은 말토스이고, 중합도 3 이상의 말토올리고당 함량은 약 18.3%이며, 중합도 3 이상의 이소말토올리고당을 포함하지 않음* Saccharide composition 3: The sugar corresponding to DP2 is maltose, the content of maltooligosaccharide having a polymerization degree of 3 or higher is about 18.3%, and isomaltooligosaccharide having a polymerization degree of 3 or higher is not included.
* 당류 조성물 4 : DP2에 해당하는 당은 말토스이고, 중합도 3 이상의 말토올리고당 함량은 약 36.7%이며, 중합도 3 이상의 이소말토올리고당을 포함하지 않으며, DE는 약 40~44임* Saccharide composition 4: The saccharide corresponding to DP2 is maltose, the content of maltooligosaccharide having a polymerization degree of 3 or higher is about 36.7%, and isomaltooligosaccharide having a polymerization degree of 3 or higher is not included, and the DE is about 40 to 44
* 당류 조성물 5 : DP2에 해당하는 당은 말토스이고, 중합도 3 이상의 말토올리고당 함량은 약 81.5%이며, 중합도 3 이상의 이소말토올리고당을 포함하지 않으며, DE는 약 20~25임* Saccharide composition 5: The sugar corresponding to DP2 is maltose, the content of maltooligosaccharide having a degree of polymerization of 3 or more is about 81.5%, it does not contain isomaltooligosaccharide having a degree of polymerization of 3 or more, and the DE is about 20-25
* 당류 조성물 6 : 당류 성분으로 포도당 약 23.4%, 중합도 2 이상의 말토올리고당 약 25.3%, 중합도 2 이상의 이소말토올리고당 약 40%, 잔량으로 기타 당류를 포함* Saccharide composition 6: saccharide components including about 23.4% of glucose, about 25.3% of maltooligosaccharide having a degree of polymerization of 2 or more, about 40% of isomaltooligosaccharide having a degree of polymerization of 2 or more, and other sugars as the balance
* 당류 조성물 7 : 당류 성분으로 포도당 약 0.7%, 중합도 2 이상의 말토올리고당 약 9.3%, 중합도 2 이상의 이소말토올리고당 약 90%를 포함* Saccharide Composition 7: Contains about 0.7% of glucose, about 9.3% of maltooligosaccharide having a polymerization degree of 2 or more, and about 90% of isomaltooligosaccharide having a polymerization degree of 2 or more as saccharide components
당류 조성물의 고형분 농도를 약 40 브릭스가 되게 조정한 후, 여기에 산화효소인 글루코스옥시다제(Glucose oxidase; 공급자 : Novozymes)를 당류 고형분 중량 대비 0.2%(v/w)의 양으로 첨가하고 효소 산화반응을 진행하였다. 상기 효소 산화반응은 공기가 공급되고 교반이 가능하며 pH 및 온도가 조절되는 반응기에서 실시하였다. 효소 산화반응 조건은 다음과 같다.After adjusting the solid concentration of the saccharide composition to be about 40 Brix, glucose oxidase (supplier: Novozymes), an oxidizing enzyme, was added thereto in an amount of 0.2% (v/w) based on the weight of the saccharide solid content, and an enzymatic oxidation reaction was performed. The enzymatic oxidation reaction was carried out in a reactor where air was supplied, agitation was possible, and pH and temperature were controlled. Enzymatic oxidation reaction conditions are as follows.
* 반응 온도 : 약 45℃; 반응 pH : 약 7; 에어레이션(aeration) 속도 : 1 vvm; 교반 속도 : 150 rpm; 반응 시간 : 약 24 hr* Reaction temperature: about 45°C; reaction pH: about 7; aeration rate: 1 vvm; Stirring speed: 150 rpm; Reaction time: about 24 hr
반응 완료 후 반응 생성물을 샘플링하고 당류 조성물의 산화 전과 산화 후의 DPPH 라디칼 소거 활성을 측정하였다(Blois, 1958 방법 참조). 먼저 1.5×10-4 mol/L 농도의 DPPH(2,2-diphenyl-1-picrylhydrazyl) 용액을 제조하고, DPPH 용액을 고형분 농도가 40 브릭스인 시료와 1:1의 부피비로 혼합한 후 상온에서 30분 동안 반응시켰다. 이후, 반응 완료액의 517㎚에서의 흡광도를 UV-visible spectrophotometer를 이용하여 측정하고 하기의 식을 이용하여 DPPH 라디칼 소거 활성을 계산하였다.After completion of the reaction, the reaction product was sampled and the DPPH radical scavenging activity before and after oxidation of the saccharide composition was measured (see Blois, 1958 method). First, a 2,2-diphenyl-1-picrylhydrazyl (DPPH) solution at a concentration of 1.5 × 10 -4 mol / L was prepared, and the DPPH solution was mixed with a sample having a solid concentration of 40 Brix at a volume ratio of 1: 1, followed by reaction at room temperature for 30 minutes. Then, the absorbance at 517 nm of the reaction solution was measured using a UV-visible spectrophotometer, and DPPH radical scavenging activity was calculated using the following formula.
또한, 대조군으로 락토스(Lactose), 락토비온산(Lactobionic acid, LBA; 순도 97% 이상), 말토비온산(Maltobionic acid, MBA) 및 글루콘산(gluconic acid, GA)을 사용하여 40 브릭스 농도의 시료를 제조하고 DPPH 라디칼 소거 활성을 측정하였다. 상기 락토비온산(Lactobionic acid, LBA)은 락토스(Lactose)의 산화에 의해 생성되는 카르복실산이고, 말토비온산(Maltobionic acid, MBA)은 말토스(Maltose)의 산화에 의해 생성되는 카르복실산이고, 글루콘산은 글루코스의 말단 알데히드기가 산화되어 카르복시기가 된 모노카르복시산이다. 하기 표 2에 당류 조성물의 산화 전 및 산화 후의 DPPH 라디칼 소거 활성을 정리하였다. In addition, using lactose, lactobionic acid (LBA; purity of 97% or higher), maltobionic acid (MBA), and gluconic acid (GA) as a control, samples having a concentration of 40 brix were prepared and DPPH radical scavenging activity was measured. Lactobionic acid (LBA) is a carboxylic acid produced by oxidation of lactose, maltobionic acid (MBA) is a carboxylic acid produced by oxidation of maltose, and gluconic acid is a monocarboxylic acid obtained by oxidation of a terminal aldehyde group of glucose to form a carboxy group. Table 2 below summarizes the DPPH radical scavenging activity of the saccharide composition before and after oxidation.
상기 표 2에서 보이는 바와 같이 다양한 당류 조성물의 산화당류 중 당류 조성물6의 산화당류가 가장 우수한 DPPH 라디칼 소거 활성을 나타냈다. 당류 조성물6의 산화당류는 단일 당류에 가까운 당류 조성물1 및 당류 조성물2의 산화당류보다 항산화 활성이 현저히 높았다. 본 발명자는 당류 조성물6의 산화당류를 '산화당류6'으로 명명하였다. 산화당류6은 폴리히이드록시산(Polyhydroxy acid, PHA)의 대표물질인 락토비온산(Lactobionic acid, LBA), 말토비온산(Maltobionic acid, MBA) 및 글루콘산(gluconic acid, GA)보다 항산화 활성이 우수하였다. 산화당의 구조를 가지는 GDL, LBA, MBA는 대표적인 PHA(polyhydroxy acid) 물질로서, 현재 화장품 산업에서 항산화, 보습, 각질 제거 효능을 부여하는 성분으로 이용되고 있고, 식품산업에서도 산화방지제로도 이용되고 있다. 따라서, 산화당류6은 항산화 활성 및 당류로서 발휘되는 다양한 기능성을 부여하기 위한 화장품 소재 및 식품 소재로 이용이 가능할 것으로 판단된다.As shown in Table 2, among the oxidized saccharides of various saccharide compositions, the oxidized saccharide of saccharide composition 6 showed the best DPPH radical scavenging activity. The oxidized saccharide of saccharide composition 6 had significantly higher antioxidant activity than the oxidized saccharides of saccharide composition 1 and saccharide composition 2, which are close to single saccharides. The present inventors named the oxidized saccharide of the saccharide composition 6 as 'oxidized saccharide 6'. Oxidized saccharide 6 was superior in antioxidant activity to lactobionic acid (LBA), maltobionic acid (MBA), and gluconic acid (GA), which are representative substances of polyhydroxy acid (PHA). GDL, LBA, and MBA, which have an oxidized sugar structure, are representative PHA (polyhydroxy acid) substances, and are currently used as ingredients that impart antioxidant, moisturizing, and exfoliating effects in the cosmetic industry, and are also used as antioxidants in the food industry. Therefore, it is believed that oxidized saccharide 6 can be used as a cosmetic material and food material to impart antioxidant activity and various functionalities exerted as saccharides.
당류 조성물6 및 당류 조성물7은 주성분으로 이소말토올리고당을 포함하는 혼합 당류이다. 당류 조성물6의 산화당류인 산화당류6과 당류 조성물7의 산화당류를 비교했을 때, 산화당류6(DPPH 라디칼 소거 활성 : 81.09%)은 당류 조성물 7의 산화당류(DPPH 라디칼 소거 활성 : 41.90%)에 비해 약 2배 가량 높은 DPPH 라디칼 소거 활성을 나타내었다. 한편, 당류 조성물2, 당류 조성물3, 당류 조성물4, 당류 조성물5와 같은 당류 조성물은 당 성분 및 함량을 살펴보면 포도당 함량이 낮고 이소말토올리고당이 아닌 말토올리고당을 포함하는데, 이들 당류 조성물의 산화당류는 약 9.71~25.50 % 범위의 낮은 DPPH 라디칼 소거 활성을 보였다. 또한, 당류 조성물6과 당류 조성물7의 당 성분 및 함량을 살펴보면, 당류 조성물6은 포도당 함량이 약 23.4%로서 당류 조성물7의 포도당 함량인 0.7%에 비해 현저하게 높았다. 상기 결과로부터 당 성분으로 소정 수준의 포도당 및 이소말토올리고당을 포함하는 당류 조성물을 산화하였을 때 높은 DPPH 라디칼 소거 활성을 가지는 산화당류를 얻을 수 있을 거라 판단된다.Saccharide composition 6 and saccharide composition 7 are mixed saccharides containing isomaltooligosaccharide as a main component. When comparing the oxidized saccharide 6 of saccharide composition 6 and the oxidized saccharide of saccharide composition 7, oxidized saccharide 6 (DPPH radical scavenging activity: 81.09%) exhibited about twice as high DPPH radical scavenging activity as compared to the oxidized saccharide of saccharide composition 7 (DPPH radical scavenging activity: 41.90%). On the other hand, saccharide compositions such as saccharide composition 2, saccharide composition 3, saccharide composition 4, and saccharide composition 5 contain maltooligosaccharides other than isomaltooligosaccharide with low glucose content and low sugar content. In addition, looking at the sugar component and content of saccharide composition 6 and saccharide composition 7, saccharide composition 6 had a glucose content of about 23.4%, which was significantly higher than that of saccharide composition 7, which was 0.7%. From the above results, it is believed that oxidized saccharides having high DPPH radical scavenging activity can be obtained when a saccharide composition containing a predetermined level of glucose and isomaltooligosaccharide as sugar components is oxidized.
2. 다양한 효소 산화반응 조건을 이용한 산화당류6의 제조 및 항산화 활성 평가2. Preparation of oxidized saccharide 6 using various enzymatic oxidation reaction conditions and evaluation of antioxidant activity
산화반응 기질로 당류 조성물6을 사용하고, 산화반응을 위한 효소로 글루코스옥시다제(Glucose oxidase; 공급자 : Novozymes), 카보하이드레이트옥시다제(Carbohydrate oxidase; 공급자 : Novozymes), 카탈라제(Catalase; 공급자 : Novozymes)의 조합을 사용하여 산화당류6을 제조하고 효소 산화반응 조건에 따른 산화당류6의 항산화 활성을 평가하였다.Oxidized saccharide 6 was prepared using a saccharide composition 6 as an oxidation reaction substrate and a combination of glucose oxidase (Supplier: Novozymes), carbohydrate oxidase (Supplier: Novozymes), and catalase (Supplier: Novozymes) as enzymes for the oxidation reaction, and the antioxidant activity of the oxidized saccharide 6 was evaluated according to the enzyme oxidation reaction conditions.
구체적으로, 당류 조성물6의 고형분 농도를 40 브릭스, 20 브릭스, 10 브릭스 및 5 브릭스가 되게 당류 조성물6 희석 용액을 준비하였다. 또한, 효소 산화반응은 공기가 공급되고 교반이 가능하며 pH 및 온도가 조절되는 반응기에서 실시하였다. 공통되는 효소 산화반응 조건은 다음과 같다.Specifically, diluted solutions of saccharide composition 6 were prepared so that the solid concentration of the saccharide composition 6 was 40 brix, 20 brix, 10 brix, and 5 brix. In addition, the enzymatic oxidation reaction was carried out in a reactor where air was supplied, agitation was possible, and pH and temperature were controlled. Common enzymatic oxidation reaction conditions are as follows.
* 에어레이션(aeration) 속도 : 1 vvm; 교반 속도 : 300 rpm; 총 반응 시간 : 약 24 hr* Aeration rate: 1 vvm; Stirring speed: 300 rpm; Total reaction time: about 24 hr
효소 산화반응 조건에 따른 효소 조합, 효소 첨가량(당류 고형분 중량 대비, v/w) 및 반응 온도는 다음과 같다.Enzyme combination, enzyme addition amount (based on sugar solid content, v/w) and reaction temperature according to enzyme oxidation reaction conditions are as follows.
* 효소 산화반응 1 : 카탈라제(Catalase) 0.1%, 글루코스옥시다제(Glucose oxidase) 0.1% 및 카보하이드레이트옥시다제(Carbohydrate oxidase) 0.1% 첨가; 반응 온도는 30℃이고 반응 pH는 7임* Enzymatic oxidation reaction 1: Catalase 0.1%, Glucose oxidase 0.1% and Carbohydrate oxidase 0.1% added; The reaction temperature is 30 °C and the reaction pH is 7
* 효소 산화반응 2 : 카탈라제(Catalase) 0.1% 및 글루코스옥시다제(Glucose oxidase) 0.2% 첨가; 반응 온도는 45℃이고 반응 pH는 6임* Enzymatic oxidation reaction 2: Catalase 0.1% and glucose oxidase 0.2% added; The reaction temperature is 45 °C and the reaction pH is 6
* 효소 산화반응 3 : 카탈라제(Catalase) 0.1% 및 카보하이드레이트옥시다제(Carbohydrate oxidase) 0.2% 첨가; 반응 온도는 30℃이고 반응 pH는 7임* Enzyme oxidation reaction 3: Catalase (Catalase) 0.1% and Carbohydrate oxidase (Carbohydrate oxidase) 0.2% addition; The reaction temperature is 30 °C and the reaction pH is 7
반응 완료 후 반응 생성물을 샘플링하고 당류 고형분 농도가 5 브릭스, 10 브릭스, 20 브릭스 및 40 브릭스인 시료의 DPPH 라디칼 소거 활성을 측정하였다(Blois, 1958 방법 참조). 하기 표 3에 효소 산화반응 조건에 따른 반응 생성물의 DPPH 라디칼 소거 활성을 정리하였다.After completion of the reaction, the reaction product was sampled and the DPPH radical scavenging activity of samples having saccharide solid concentrations of 5, 10, 20, and 40 Brix was measured (see Blois, 1958 method). Table 3 below summarizes the DPPH radical scavenging activity of the reaction product according to the enzyme oxidation reaction conditions.
상기 표 3에서 보이는 바와 같이 당류 조성물6의 효소 산화반응을 통해 산화당류6을 제조시 다양한 산화효소 조합을 사용하는 경우 반응 시간은 4 hr 이하일 때 높은 항산화 활성을 보였다. 또한, 카탈라제(Catalase) 0.1% 및 글루코스옥시다제(Glucose oxidase) 0.2%를 첨가한 효소 산화반응 조건 2에서는 반응 완료 후 샘플링한 산화당류6 용액의 고형분 농도가 낮은 경우에도 높은 항산화 활성을 나타냈다. 한편, 카탈라제(Catalase) 0.1% 및 카보하이드레이트옥시다제(Carbohydrate oxidase) 0.2%를 첨가한 산화반응 조건 3에서 가장 낮은 항산화 활성을 나타냈다.As shown in Table 3, when preparing oxidized saccharide 6 through enzymatic oxidation of saccharide composition 6, when various combinations of oxidases were used, high antioxidant activity was exhibited when the reaction time was 4 hr or less. In addition, in the enzyme oxidation reaction condition 2 in which 0.1% of catalase and 0.2% of glucose oxidase were added, high antioxidant activity was exhibited even when the solid concentration of the oxidized saccharide 6 solution sampled after completion of the reaction was low. On the other hand, the oxidation reaction condition 3 in which 0.1% of catalase and 0.2% of carbohydrate oxidase were added showed the lowest antioxidant activity.
3. 산화당류6의 항산화 활성에 영향을 미치는 유효성분의 탐색3. Exploration of active ingredients that affect the antioxidant activity of oxidized saccharide 6
산화당류6의 높은 항산화 활성에 영향을 미치는 유효성분을 탐색하기 위해 다양한 당류를 조합하여 임의의 시료를 제조하였다.In order to explore active ingredients that affect the high antioxidant activity of oxidized saccharide 6, random samples were prepared by combining various saccharides.
* 제1 조합 : 포도당, 말토스 등과 같은 DP1, DP2 당류에 GDL, MBA와 같은 표준시료를 당류 고형분 중량을 기준으로 1:4 내지 4:1의 중량비로 혼합하고 희석하여 총 고형분 농도가 20 브릭스인 시료를 제조함* First combination: DP1 and DP2 sugars such as glucose and maltose are mixed with standard samples such as GDL and MBA at a weight ratio of 1:4 to 4:1 based on the weight of the sugar solid content, and then diluted to prepare a sample with a total solid concentration of 20 brix.
* 제2 조합 : 당류 조성물6이나 이미 제조한 산화당류6[제조조건 : 당류 고형분 농도가 40 브릭스인 당류 조성물6 희석 용액에 카탈라제(Catalase) 및 글루코스옥시다제(Glucose oxidase)를 각각 당류 고형분 중량 대비 0.1%(v/w) 및 0.2%(v/w)의 양으로 첨가하고 반응 온도 45℃, 반응 pH 6, 에어레이션(aeration) 속도 1 vvm 및 교반 속도 300 rpm의 조건에서 4 hr 동안 효소 산화반응을 진행시킴]에 GA, MBA와 같은 표준시료를 당류 고형분 중량을 기준으로 1:4 내지 4:1의 중량비로 혼합하고 희석하여 총 고형분 농도가 20 브릭스인 시료를 제조함* Second combination: Saccharide composition 6 or previously prepared oxidized saccharide 6 [Manufacturing conditions: Catalase and glucose oxidase were added in an amount of 0.1% (v/w) and 0.2% (v/w), respectively, based on the weight of the saccharide solid content, to a diluted solution of saccharide composition 6 having a saccharide solid content concentration of 40 brix, reaction temperature 45 ° C, reaction pH 6, aeration rate 1 Enzymatic oxidation reaction proceeds for 4 hr under conditions of vvm and agitation speed of 300 rpm], standard samples such as GA and MBA are mixed and diluted in a weight ratio of 1: 4 to 4: 1 based on the weight of sugar solid content and diluted to prepare a sample with a total solid concentration of 20 brix
* 제3 조합 : 당류 조성물6에 포도당, 말토스 등과 같은 DP1, DP2 당류를 당류 고형분 중량을 기준으로 1:1의 중량비로 혼합하고 희석하여 총 고형분 농도가 20 브릭스인 반응 원액용 당류 용액을 제조하고, 제조한 당류 용액에 카탈라제(Catalase) 및 글루코스옥시다제(Glucose oxidase)를 각각 당류 고형분 중량 대비 0.1%(v/w) 및 0.2%(v/w)의 양으로 첨가하고 반응 온도 45℃, 반응 pH 6, 에어레이션(aeration) 속도 1 vvm 및 교반 속도 300 rpm의 조건에서 4 hr 동안 효소 산화반응을 진행시켜 총 고형분 농도가 20 브릭스인 시료를 제조함* Third combination: A saccharide solution for the reaction stock solution having a total solid concentration of 20 brix was prepared by mixing and diluting DP1 and DP2 saccharides such as glucose and maltose in saccharide composition 6 at a weight ratio of 1: 1 based on the solid weight of the saccharide, and adding catalase and glucose oxidase to the prepared saccharide solution at 0.1% (v/w) and 0.2% (v/w), respectively, based on the solid weight of the saccharide. % (v/w) was added and enzymatic oxidation was carried out for 4 hr under the conditions of a reaction temperature of 45 ° C, a reaction pH of 6, an aeration rate of 1 vvm and an agitation rate of 300 rpm to prepare a sample with a total solid concentration of 20 Brix
상기 제1 조합, 제2 조합 및 제3 조합을 통해 제조한 총 고형분 농도가 20 브릭스인 시료의 DPPH 라디칼 소거 활성을 측정하였다(Blois, 1958 방법 참조). 그 결과, 제1 조합에서는 제조한 모든 시료에서 약 20~30%의 DPPH 라디칼 소거 활성을 나타냈고, 산화당류6보다 우월한 값을 나타내는 시료는 없었다. 도 1은 본 발명의 실시예에서 산화당류6의 항산화 활성에 영향을 미치는 유효성분을 탐색하기 위해 제2 조합에 의해 제조한 다양한 시료들의 DPPH 라디칼 소거 활성을 나타낸 그래프이다. 도 1에서 X축은 당류 고형분 중량을 기준으로 한 혼합 중랑비를 나타낸다. 도 1에서 보이는 바와 같이 MBA와 산화당류6을 혼합한 시료에서 가장 높은 DPPH 라디칼 소거 활성을 나타냈다. 그러나, 고형분 농도가 20 브릭스인 산화당류6 자체의 DPPH 라디칼 소거활성 값이 87.4%인 것을 감안하면 MBA와 산화당류6을 혼합한 시료의 항산화 활성은 산화당류6에 의해 기여되는 것이며, 산화당류6에 MBA를 혼합하면 DPPH 라디칼 소거활성 값이 오히려 낮아지는 결과를 보였다. 한편, 당류 조성물6에 MBA나 GA를 혼합하는 경우 항산화 활성이 높아지는 결과를 보였다. 도 2는 본 발명의 실시예에서 산화당류6의 항산화 활성에 영향을 미치는 유효성분을 탐색하기 위해 제3 조합에 의해 제조한 다양한 시료들의 DPPH 라디칼 소거 활성을 나타낸 그래프이다. 도 2에서 보이는 바와 같이 당류 조성물6에 포도당 또는 말토스를 혼합하고 효소 산화반응에 의해 산화시켰을 때, 반응 생성물의 DPPH 라디칼 소거 활성이 산화당류6보다 낮은 값을 나타냈다. 이러한 결과는 산화당류6에 GA 또는 MBA를 혼합한 제2 조합과 유사한 결과로서, 당류 조성물6에 다른 당을 혼합하여 산화시키거나 산화당류6에 다른 산화당을 혼합하는 경우보다 당류 조성물6 자체를 산화시켜 산화당류6을 제조할 때 가장 우수한 항산화 활성을 가지는 것으로 나타났다. 또한, 도 2의 무처리구와 같이 당류 조성물6에 포도당 또는 말토스를 혼합한 경우보다 당류 조성물6 자체의 항산화 활성이 높은 것으로 나타났는바, 산화당류6의 높은 항산화 활성은 산화당류6을 제조하기 위해 사용된 당류 조성물6의 당 성분 및 함량에 의해 기인하고, 당류 조성물6을 산화시킴에 따라 항산화 활성에 대해 시너지 작용을 갖는 산화당 조성으로 전환되는 것으로 판단된다.The DPPH radical scavenging activity of samples having a total solid concentration of 20 brix prepared through the first, second and third combinations was measured (see Blois, 1958 method). As a result, in the first combination, about 20 to 30% of the DPPH radical scavenging activity was exhibited in all samples prepared, and there was no sample showing a value superior to oxidized saccharide 6. Figure 1 is a graph showing the DPPH radical scavenging activity of various samples prepared by the second combination to search for active ingredients that affect the antioxidant activity of oxidized saccharide 6 in Examples of the present invention. In Figure 1, the X-axis represents the mixing medium ratio based on the weight of the saccharide solid content. As shown in FIG. 1, the highest DPPH radical scavenging activity was shown in the mixture of MBA and oxidized saccharide 6. However, considering that the DPPH radical scavenging activity value of oxidized saccharide 6 itself with a solid concentration of 20 Brix is 87.4%, the antioxidant activity of the sample mixed with MBA and oxidized saccharide 6 is contributed by oxidized saccharide 6, and the DPPH radical scavenging activity value was rather lowered when MBA was mixed with oxidized saccharide 6. On the other hand, when MBA or GA was mixed with the saccharide composition 6, the antioxidant activity was increased. Figure 2 is a graph showing the DPPH radical scavenging activity of various samples prepared by the third combination to search for active ingredients that affect the antioxidant activity of oxidized saccharide 6 in Examples of the present invention. As shown in FIG. 2, when glucose or maltose was mixed with saccharide composition 6 and oxidized by enzymatic oxidation, the DPPH radical scavenging activity of the reaction product was lower than that of oxidized saccharide 6. These results are similar to the second combination of GA or MBA mixed with oxidized saccharide 6, and the oxidized saccharide 6 is produced by oxidizing the saccharide composition 6 itself, rather than oxidizing by mixing other sugars with saccharide composition 6 or mixing other oxidized sugars with oxidized saccharide 6. It was found to have the best antioxidant activity. In addition, it was found that the antioxidant activity of the saccharide composition 6 itself was higher than that of the case where glucose or maltose was mixed with the saccharide composition 6 as in the untreated group of FIG.
4. 산화당류6의 성분 특성4. Component Characteristics of Oxidized Saccharide 6
당류 조성물6을 효소 산화반응시켜 산화당류6을 제조할 때 당류 조성물6을 구성하는 당 성분 및 함량의 변화를 분석하였다.When oxidized saccharide 6 was prepared by enzymatic oxidation of saccharide composition 6, changes in sugar components and contents constituting saccharide composition 6 were analyzed.
먼저, 당류 조성물6의 고형분 농도를 40 브릭스로 희석하여 당류 조성물6 희석 용액을 준비하였다. 이후, 당류 조성물6 희석 용액에 카탈라제(Catalase) 및 글루코스옥시다제(Glucose oxidase)를 각각 당류 고형분 중량 대비 0.1%(v/w) 및 0.2%(v/w)의 양으로 첨가하고 반응 온도 45℃, 반응 pH 6, 에어레이션(aeration) 속도 1 vvm 및 교반 속도 300 rpm의 조건에서 4 hr 동안 효소 산화반응을 진행시켜 총 고형분 농도가 약 40 브릭스인 산화당류6을 제조하였다. 이후, 당류 조성물6과 산화당류6의 성분을 LC-TOF/MS를 이용하여 분석하였다. 당류 조성물6과 산화당류6의 성분 분자량을 150~500 범위와 500~1700 범위로 나누어 분석하였다. 분자량 150~500 범위를 측정하는 경우 Detector[V] : 2300; Orifice[V] : 20; Ring Lens : 5; Orifice[V] : 2; 및 IonGuide radio frequency: 500의 조건으로 분석하였고, 분자량 500~1700 범위를 측정하는 경우 Detector[V] : 2300; Orifice[V] : 80; Ring Lens : 5; Orifice[V] : 2; 및 IonGuide radio frequency : 2000의 조건으로 분석하였다. 시료는 물에 녹인 후 0.1% FA MeOH로 농도를 맞춰가며 ESI+로 측정하였다.First, a diluted solution of saccharide composition 6 was prepared by diluting the solid concentration of saccharide composition 6 to 40 brix. Thereafter, catalase and glucose oxidase were added in an amount of 0.1% (v/w) and 0.2% (v/w) based on the weight of the saccharide solid content, respectively, to the diluted saccharide composition 6 solution, and an enzymatic oxidation reaction was carried out for 4 hr under the conditions of a reaction temperature of 45 ° C, a reaction pH of 6, an aeration rate of 1 vvm and a stirring speed of 300 rpm, so that the total solid concentration was about Oxidized saccharide 6, which is 40 Brix, was prepared. Then, components of saccharide composition 6 and oxidized saccharide 6 were analyzed using LC-TOF/MS. The component molecular weights of saccharide composition 6 and oxidized saccharide 6 were divided into 150 to 500 ranges and 500 to 1700 ranges for analysis. Detector[V]: 2300 when measuring the molecular weight range of 150 to 500; Orifice[V] : 20; Ring Lens: 5; Orifice[V] : 2; and IonGuide radio frequency: analyzed under the condition of 500, and when measuring the molecular weight in the range of 500 to 1700, Detector [V]: 2300; Orifice[V] : 80; Ring Lens: 5; Orifice[V] : 2; and IonGuide radio frequency: analyzed under the conditions of 2000. After the sample was dissolved in water, the concentration was adjusted with 0.1% FA MeOH and measured by ESI+.
도 3은 본 발명의 실시예에서 산화당류6의 성분 특성을 규명하기 위해 당류 조성물6과 산화당류6의 성분을 LC-TOF/MS를 이용하여 분석한 결과이다. 도 3에서 보이는 바와 같이 LC/MS 분석 결과, DP1~DP4까지의 당류와 이들의 산화된 당의 분자량이 확인되었다. 산화된 당은 주 peak인 DP1~DP4의 피크에서 분자량이 약 16 정도 상승되는 것으로 확인되었다. 당류 조성물6은 다른 당류 제품에 비해 DP1~DP4의 함량이 높고, 효소 산화반응에 의해 DP1~DP4에 해당하는 당류가 주로 산화되어 항산화 활성이 크게 상승하는 것으로 판단된다.3 is a result of analyzing the saccharide composition 6 and the components of oxidized saccharide 6 using LC-TOF/MS to identify the component characteristics of oxidized saccharide 6 in Examples of the present invention. As shown in FIG. 3, as a result of LC/MS analysis, the molecular weights of saccharides from DP1 to DP4 and their oxidized sugars were confirmed. It was confirmed that the molecular weight of the oxidized sugar increased by about 16 at the peaks of DP1 to DP4, which are the main peaks. Saccharide composition 6 has a higher content of DP1 to DP4 than other saccharide products, and it is judged that the antioxidant activity is greatly increased because the sugars corresponding to DP1 to DP4 are mainly oxidized by the enzymatic oxidation reaction.
본 발명에서 사용한 당류 산화효소는 주로 글루코스(glucose)를 타겟하여 산화시키는 효소로서, 당류 조성물6을 구성하는 DP1~DP10까지의 당류 중 주로 DP1~D4까지의 당류를 산화시키는 것으로 판단된다. 또한, 효소 산화반응에서 부산물로 생성되는 신규 물질도 약 200~235 사이의 분자량을 가지는바, DP1~DP2에서 유래된 산화 부산물인 것으로 판단된다. 당류 조성물6을 구성하는 DP1~D4까지의 당류는 포도당과 이소말토올리고당에 해당하는바, 포도당 산화물과 이소말토올리고당 산화물의 조합에 의해 항산화 활성이 매우 우수한 것으로 판단된다. 반면, 당류 조성물7은 당류 조성물6에 비해 DP5~DP10까의 당류 함량이 상대적으로 높고 포도당을 거의 함유하고 있지 않기 때문에 당류 산화효소에 의한 산화가 적정 수준으로 이루어지지 않게 되고, 산화에 의한 항산화 활성의 상승도 극적이지 않은 것으로 판단된다.The saccharide oxidase used in the present invention is an enzyme that mainly targets and oxidizes glucose, and is considered to oxidize mainly saccharides from DP1 to D4 among the saccharides from DP1 to DP10 constituting saccharide composition 6. In addition, since the new material produced as a by-product in the enzymatic oxidation reaction has a molecular weight between about 200 and 235, it is considered to be an oxidation by-product derived from DP1 to DP2. The saccharides DP1 to D4 constituting the saccharide composition 6 correspond to glucose and isomaltooligosaccharide, and the combination of glucose oxide and isomaltooligosaccharide oxide is considered to have excellent antioxidant activity. On the other hand, since saccharide composition 7 has a relatively high saccharide content from DP5 to DP10 compared to saccharide composition 6 and contains almost no glucose, oxidation by saccharide oxidase does not occur at an appropriate level, and the increase in antioxidant activity due to oxidation is also not dramatic.
5. 산화당류6의 관능 특성5. Sensory characteristics of oxidized saccharide 6
당류 조성물6의 고형분 농도를 40 브릭스로 희석하여 당류 조성물6 희석 용액을 준비하였다. 이후, 당류 조성물6 희석 용액에 카탈라제(Catalase) 및 글루코스옥시다제(Glucose oxidase)를 각각 당류 고형분 중량 대비 0.1%(v/w) 및 0.2%(v/w)의 양으로 첨가하고 반응 온도 45℃, 반응 pH 6, 에어레이션(aeration) 속도 1 vvm 및 교반 속도 300 rpm의 조건에서 4 hr 동안 효소 산화반응을 진행시켜 총 고형분 농도가 약 40 브릭스인 산화당류6을 제조하였다. 이후, 당류 조성물6과 산화당류6의 신맛, 쓴맛, 떫은 맛, 짠맛, 감칠맛, 풍미를 미각테스터기를 사용하여 측정하였다. 그 결과, 산화당류6은 당류 조성물6에 비해 신맛은 38.7 증가하였고 쓴맛, 떫은맛 및 짠맛은 각각 0.5, 1.9 및 3.8 증가하였으며, 감칠맛 및 풍미는 각각 6.6 및 0.2 감소하였다. 상기 결과로부터 산화당류6은 당류 조성물6와 비교할 때 신맛이 크게 증가하였지만 나머지 맛들의 변화는 크지 않은 것을 알수 있다. 따라서, 산화당류6은 산미 및 다양한 맛을 부여하는 식품 첨가물로 사용될 수 있다.A diluted solution of saccharide composition 6 was prepared by diluting the solid concentration of saccharide composition 6 to 40 brix. Thereafter, catalase and glucose oxidase were added in an amount of 0.1% (v/w) and 0.2% (v/w) based on the weight of the saccharide solid content, respectively, to the diluted saccharide composition 6 solution, and an enzymatic oxidation reaction was carried out for 4 hr under the conditions of a reaction temperature of 45 ° C, a reaction pH of 6, an aeration rate of 1 vvm and a stirring speed of 300 rpm, so that the total solid concentration was about Oxidized saccharide 6, which is 40 Brix, was prepared. Thereafter, sourness, bitterness, astringency, saltiness, umami, and flavor of the saccharide composition 6 and the oxidized saccharide 6 were measured using a taste tester. As a result, compared to saccharide composition 6, oxidized saccharide 6 increased acidity by 38.7, bitterness, astringency, and saltiness by 0.5, 1.9, and 3.8, respectively, and umami and flavor decreased by 6.6 and 0.2, respectively. From the above results, it can be seen that the acidity of oxidized saccharide 6 was greatly increased compared to that of saccharide composition 6, but the other tastes were not changed significantly. Therefore, oxidized saccharide 6 can be used as a food additive that imparts acidity and various tastes.
이상에서와 같이 본 발명을 실시예를 통해 설명하였지만 본 발명이 반드시 여기에만 한정되는 것은 아니며 본 발명의 범주와 사상을 벗어나지 않는 범위 내에서 다양한 변형실시가 가능함은 물론이다. 따라서, 본 발명의 보호범위는 본 발명에 첨부된 특허청구의 범위에 속하는 모든 실시 형태를 포함하는 것으로 해석되어야 한다.As described above, the present invention has been described through examples, but the present invention is not necessarily limited thereto, and various modifications are possible without departing from the scope and spirit of the present invention. Accordingly, the scope of protection of the present invention should be construed to include all embodiments falling within the scope of the claims appended hereto.
Claims (9)
상기 당류 조성물은 당류 조성물 내의 당류 전체 중량을 기준으로 이소말토올리고당 20~60 중량%를 포함하고,
상기 당류 조성물은 당류 조성물 내의 당류 전체 중량을 기준으로 포도당 16~30 중량%, 글루코스를 단위구조로 가지는 이당류 18.5~32.5 중량%, 글루코스를 단위구조로 가지는 삼당류 23~37 중량%, 글루코스를 단위구조로 가지는 사당류 8.5~18.5 중량%, 글루코스를 단위구조로 가지는 오당류 1.5~7.5 중량%, 글루코스를 단위구조로 가지는 육당류 0.5~3 중량% 및 글루코스를 단위구조로 가지는 칠당류 0.2~2 중량%를 포함하는 것을 특징으로 하는 산화당류 조성물의 제조방법.
A method comprising the step of adding a saccharide oxidase to a saccharide composition solution and performing an oxidative reaction to convert at least some of the saccharides constituting the saccharide composition into oxidized saccharides,
The saccharide composition contains 20 to 60% by weight of isomaltooligosaccharide based on the total weight of saccharides in the saccharide composition,
The saccharide composition comprises 16 to 30% by weight of glucose, 18.5 to 32.5% by weight of disaccharides having glucose as a unit structure, 23 to 37% by weight of trisaccharides having glucose as a unit structure, 8.5 to 18.5% by weight of tetrasaccharides having glucose as a unit structure, 1.5 to 7.5% by weight of pentasaccharides having glucose as a unit structure, and glucose as a unit structure, based on the total weight of saccharides in the saccharide composition. A method for producing an oxidized saccharide composition comprising 0.5 to 3% by weight of hexaccharides and 0.2 to 2% by weight of heptasaccharides having glucose as a unit structure.
The method of claim 1, wherein the saccharide composition comprises 18.5 to 28.5% by weight of glucose, 20 to 60% by weight of isomaltooligosaccharide, and 20 to 60% by weight of maltooligosaccharide based on the total weight of saccharides in the saccharide composition.
The method for preparing an oxidized saccharide composition according to claim 1, wherein the content of the oligosaccharide having glucose as a unit structure and a degree of polymerization of 2 to 4 in the saccharide composition is 55 to 80% by weight based on the total weight of the saccharide.
The method of claim 1, wherein the total content of glucose and oligosaccharides having glucose as a unit structure and a degree of polymerization of 2 to 4 in the saccharide composition is 80 to 98% by weight based on the total weight of the saccharide.
The method of claim 1, wherein the content of glucose and oligosaccharides having glucose as a unit structure and a degree of polymerization of 2 to 10 in the saccharide composition is 95 to 99.9% by weight based on the total weight of the saccharide.
The saccharide composition according to claim 1, based on the total weight of saccharides in the saccharide composition, 18.5 to 28.5% by weight of glucose, 20.5 to 30.5% by weight of disaccharides having glucose as a unit structure, 25 to 35% by weight of trisaccharides having glucose as a unit structure, 10 to 17% by weight of tetrasaccharides having glucose as a unit structure, 2.5 to 6.5% by weight of pentasaccharides having glucose as a unit structure, A method for producing an oxidized saccharide composition comprising 0.8 to 2.2% by weight of hexasaccharides having glucose as a unit structure and 0.3 to 1.5% by weight of hepsugars having glucose as a unit structure.
The method of claim 1, wherein the saccharide oxidase is composed of one or more selected from glucose oxidase (CAS No. 9001-37-0), carbohydrate oxidase (CAS No. 372580-9) or hexose oxidase (CAS No. 9028-75-5), or glucose oxidase (Glucose oxidase). ASE; CAS No. 9001-37-0) and catalase (Catalase; CAS No. 9001-05-2).
The method of claim 1, wherein the solid concentration of the saccharide composition solution is 6 to 50 Brix, the amount of saccharide oxidase added is 0.05 to 1.0% (v/w) based on the solid content of the saccharide composition, the oxidation reaction temperature is 25 to 50 ° C, the oxidation pH is 5.0 to 8.0, and the oxidation reaction time is 30 minutes to 24 hr.
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