WO2020024468A1 - 一种具有高sod酶活力的口服液及其制备方法 - Google Patents

一种具有高sod酶活力的口服液及其制备方法 Download PDF

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WO2020024468A1
WO2020024468A1 PCT/CN2018/113302 CN2018113302W WO2020024468A1 WO 2020024468 A1 WO2020024468 A1 WO 2020024468A1 CN 2018113302 W CN2018113302 W CN 2018113302W WO 2020024468 A1 WO2020024468 A1 WO 2020024468A1
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enzyme activity
juice
treatment
sod enzyme
ultra
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French (fr)
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廖小军
侯志强
赵靓
王永涛
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中国农业大学
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Priority to US17/264,841 priority Critical patent/US20210298330A1/en
Publication of WO2020024468A1 publication Critical patent/WO2020024468A1/zh

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    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23LFOODS, 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
    • A23L2/00Non-alcoholic beverages; Dry compositions or concentrates therefor; Their preparation
    • A23L2/02Non-alcoholic beverages; Dry compositions or concentrates therefor; Their preparation containing fruit or vegetable juices
    • A23L2/04Extraction of juices
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23LFOODS, 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
    • A23L33/00Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23LFOODS, 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
    • A23L29/00Foods or foodstuffs containing additives; Preparation or treatment thereof
    • A23L29/06Enzymes
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23LFOODS, 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
    • A23L33/00Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/105Plant extracts, their artificial duplicates or their derivatives
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/0004Oxidoreductases (1.)
    • C12N9/0089Oxidoreductases (1.) acting on superoxide as acceptor (1.15)
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12YENZYMES
    • C12Y115/00Oxidoreductases acting on superoxide as acceptor (1.15)
    • C12Y115/01Oxidoreductases acting on superoxide as acceptor (1.15) with NAD or NADP as acceptor (1.15.1)
    • C12Y115/01001Superoxide dismutase (1.15.1.1)
    • AHUMAN NECESSITIES
    • A23FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23VINDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00Food compositions, function of food ingredients or processes for food or foodstuffs

Definitions

  • the invention belongs to the biological field, and particularly relates to an oral liquid with high SOD enzyme activity and a preparation method thereof.
  • SOD Superoxide dismutase
  • SOD is a type of oxidoreductase that is widely present in various organisms. It is a metal enzyme that can remove superoxide radical anions in the body and can effectively prevent superoxide freedom. The toxic effect of base anions on the body is a widely used medicinal enzyme.
  • SOD has been isolated from organisms such as bacteria, fungi, algae, insects, fish, plants and mammals. Due to the different metal prosthetic groups, these SODs can currently be divided into four types: Cu / Zn-SOD, Mn-SOD, Fe-SOD, and Ni-SOD. Plant-derived SOD has been shown to have a positive effect on human health. Common plant materials for SOD enzymes include sea buckthorn, kiwi, mulberry, blueberry, prickly pear, etc., but SOD enzyme activity per unit weight or volume is low. In previous studies, DEAE ion exchange separation technology has been used to purify superoxide dismutase from plant fruits, with a molecular weight of 30-35 kDa. However, the existing DEAE ion exchange separation technology has the disadvantages of high cost, long separation time, and low product yield, which makes it difficult to apply to large-scale production practice.
  • the present invention provides an oral solution with high SOD enzyme activity and a preparation method thereof. After treatment with the method of the present invention, the SOD enzyme activity in the product is high and stable, which improves the health function of the product and effectively solves the problem Problems of enzyme activity loss and instability during processing. Therefore, the product prepared by the method of the present invention is more beneficial to human health.
  • the method for preparing an oral liquid with high SOD enzyme activity provided by the present invention includes the following steps:
  • the edible plant raw materials may be edible plant roots, stems, leaves, fruits, etc., specifically, one or more of blueberry, kiwi, mulberry, sea buckthorn, prickly pear and other fruits.
  • the process of the juice-making treatment may specifically be crushing juice.
  • step 2) the process conditions of the centrifugal treatment are: 1000-4000 rpm, 0-30 min (the endpoint 0 is not taken); specifically, 3000 rpm, 10 min.
  • the pore diameter of the ultrafiltration membrane is: tubular membrane 60-120kD, and the roll membrane is 8-30kD; specifically, the pore diameter of the ultrafiltration membrane is: tubular membrane 100kD, Roll film 10kD.
  • the pressure of the ultra-high pressure treatment may be: 200-600 MPa, and the time may be: 1-20 min. Specifically, the pressure of the ultra-high pressure treatment may be 550 MPa, and the time may be 5 min.
  • the temperature of the ultra-high pressure treatment is in the range of 0-40 ° C.
  • the ultra-high pressure treated pressure transmitting medium is preferably water.
  • step 5 a liquid having high SOD enzyme activity is made into an oral liquid by a conventional method in the art.
  • the oral solution with high SOD enzyme activity prepared by the above method also belongs to the protection scope of the present invention.
  • the soluble solids of the oral solution with high SOD enzyme activity is 2-15 ° Brix, and the pH is 2.0-6.0.
  • the above process can activate the SOD enzyme, increase the activity of the SOD enzyme in the oral solution, and ensure the health function of the product.
  • the present invention uses centrifugation and dual ultrafiltration with different cut-off molecular weights to separate the oral solution, which can effectively improve its enzyme activity and greatly improve the health function of the product;
  • the present invention uses an ultra-high pressure treatment method to activate the SOD enzyme and increase its enzyme activity.
  • the technology adopted by the present invention has the advantages of low cost, high efficiency, strong practicality and the like.
  • the invention discloses a method for processing an oral liquid with high SOD enzyme activity, which comprises crushing blueberries, mulberries, sea buckthorn, and prickly pear to make juice, centrifugation, ultrafiltration, and ultrahigh pressure.
  • the specific method comprises the following steps: packaging the fruit of a plant into suitable containers, performing centrifugal treatment under the following conditions, taking a supernatant for two-way ultrafiltration separation, and then performing ultra-high pressure treatment, and the pressure transmitting medium is water.
  • the method for processing the oral liquid includes the following steps:
  • Ultra-high pressure treatment the conditions are a pressure of 550 MPa and a time of 5 min.
  • the oral solid solution with high SOD enzyme activity prepared according to the method has a soluble solid content of about 8 ° Brix and a pH of 3.7.
  • a method for processing an oral solution with high SOD enzyme activity including juice making, centrifugation, ultrafiltration, and ultrahigh pressure treatment of blueberry, kiwi, sea buckthorn, and prickly pear.
  • the specific methods include: preparing the raw materials into appropriate portions The container was subjected to centrifugation under the following conditions, and the supernatant was taken for ultrafiltration and separation; thereafter, ultrahigh pressure treatment was performed.
  • the method for processing the oral liquid includes the following steps:
  • Ultra-high pressure treatment the conditions are a pressure of 550 MPa and a time of 5 min.
  • the soluble solids produced by the method with high SOD enzyme activity have a soluble solid content of about 10 ° Brix and a pH of 2.7.
  • a method for processing fruit juice differs from Example 1 only in that ultrafiltration concentration and ultrahigh pressure treatment are not used.
  • a method for processing fruit juice which is different from Example 1 in that only ultrafiltration concentration treatment is used, and ultrahigh pressure treatment is not used.
  • a method for processing fruit juice is different from Example 1 in that ultrafiltration concentration is not used, and other steps are the same.
  • a method for processing fruit juice is different from Example 1 in that the ultrafiltration separation condition is: only a tubular membrane 100 kD is used, and other steps are the same.
  • a method for processing fruit juice differs from Example 1 only in that ultrafiltration concentration and ultrahigh pressure treatment are not used.
  • a method for processing fruit juice which is different from Example 1 in that only ultrafiltration concentration treatment is used, and ultrahigh pressure treatment is not used.
  • the SOD enzyme activities of the fruit and vegetable juices prepared in Example 1 and Comparative Example 1 were tested separately. The results are shown in Table 1 below.
  • the SOD activity detection method is the WST-1 method (references Peskin, AV, & Winterbourn, CC (2000).
  • WST-1 is 2- (4-Iodophenyl) -3- (4-nitrophenyl) -5- (2,4-disulfophenyl) -2H-tetrazolium, monosodium salt.
  • WST-1 can react with the superoxide anion produced by xanthine oxidase to produce water-soluble formazan dye. This reaction step can be inhibited by SOD.
  • the enzyme activity of SOD can be calculated by colorimetric analysis of WST-1 products.
  • Example 2 The SOD enzyme activities of the fruit juice prepared in Example 2 and Comparative Example 2-1 and Comparative Example 2-2 were examined.
  • the SOD activity detection method experimental example 1 is the same. The results are shown in Table 2 below.
  • the process of the invention can activate the SOD enzyme, improve the activity of the SOD enzyme in the oral solution, and ensure the health function of the product.
  • the invention uses centrifugation and dual ultrafiltration with different cut-off molecular weights to separate the oral solution, which can effectively improve its enzyme activity and greatly improve the health function of the product.
  • the invention uses ultra-high pressure treatment to activate the SOD enzyme and increase its enzyme activity. ; Compared with the traditional DEAE ion exchange separation technology, the technology used in the present invention has the advantages of low cost, high efficiency, strong practicality and the like.

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Abstract

一种具有高SOD酶活力的口服液及其制备方法,所述方法包括将可食植物原料进行榨汁,然后离心去除大颗粒,使用两道超滤膜进行分离,再采用超高压处理。

Description

一种具有高SOD酶活力的口服液及其制备方法 技术领域
本发明属于生物领域,具体涉及一种具有高SOD酶活力的口服液及其制备方法。
背景技术
超氧化物歧化酶(Superoxide dismutase,缩写SOD)是一类广泛存在于各类生物体内的氧化还原酶,它是一种能清除体内超氧自由基阴离子的金属酶,能有效地预防超氧自由基阴离子对机体的毒害作用,是一种疗效广泛的药用酶。
迄今为止,已从细菌、真菌、藻类、昆虫、鱼类、植物和哺乳动物等生物体内分离得到了SOD。由于金属辅基的不同,这些SOD目前可以分为四种类型:Cu/Zn-SOD、Mn-SOD、Fe-SOD、Ni-SOD。植物源的SOD被证明对人体健康有积极的作用,SOD酶的常见植物材料有沙棘、猕猴桃、桑葚、蓝莓、刺梨等,但是单位重量或体积SOD酶活性较低。前期研究已经利用DEAE离子交换分离技术从植物果实中纯化出了超氧化物歧化酶,其分子量为30~35kDa。但现有DEAE离子交换分离技术存在成本高、分离时间长、产物得率低等缺点,应用到大规模生产实践难度大。
发明公开
为解决上述技术问题,本发明提供一种具有高SOD酶活力的口服液及其制备方法,经过本发明方法处理后产品中SOD酶活性高且稳定,提升了产品的保健功能,有效解决了SOD酶在加工过程中的酶活力损失和不稳定的问题。因此,经本发明方法制得的产品更有益于人体健康。
本发明所提供的具有高SOD酶活力的口服液的制备方法,包括下述步骤:
1)对可食植物原料进行制汁处理,得到植物汁液;
2)对植物汁液进行离心处理,收集上清液;
3)对上清液进行双道超滤分离,收集滤液;
4)对所得滤液进行超高压处理,得到具有高SOD酶活力的液体;
5)将所述具有高SOD酶活力的液体制成口服液。
上述方法步骤1)中,所述可食植物原料可为可食植物根、茎、叶、果实等,具体可为蓝莓、猕猴桃、桑葚、沙棘、刺梨等果实中的一种或几种。
所述制汁处理的工艺具体可为破碎榨汁。
步骤2)中,所述离心处理的工艺条件为:1000-4000rpm、0-30min(端点0不取);具体可为3000rpm、10min。
步骤3)中,所述双道超滤分离中,超滤膜孔径为:管式膜60-120kD,卷式膜8-30kD;具体地,所述超滤膜孔径为:管式膜100kD,卷式膜10kD。
步骤4)中,所述超高压处理的压力可为:200-600MPa,时间可为:1-20min,具体地,所述超高压处理的压力可为550MPa,时间可为5min。
所述超高压处理的温度在0-40℃范围内。
所述超高压处理的传压介质优选为水。
步骤5)中,采用本领域常规方法将具有高SOD酶活力的液体制成口服液。
通过上述方法制备得到的具有高SOD酶活力的口服液也属于本发明的保护范围。
所述具有高SOD酶活力的口服液的可溶性固形物为2-15°Brix,pH为2.0-6.0。
上述工艺处理可以激活SOD酶,提高口服液中SOD酶的活性,保障产品的保健功能。
本发明的有益效果在于:
1)本发明采用离心和不同截留分子量双超滤的方式对口服液进行了分离,可以有效提升其酶活力,大大提升了产品的保健功能;
2)本发明采用超高压处理方法激活SOD酶,提高其酶活力。
3)本发明采用的技术相比于传统DEAE离子交换分离技术,具有成本低、效率高、实践性强等优点。
实施发明的最佳方式
下述实施例中的实验方法,如无特别说明,均为常规方法
实施例1
一种具有高SOD酶活力的口服液的加工方法,包括将蓝莓、桑葚、沙棘、刺梨进行破碎制汁、离心、超滤和超高压处理。具体方法包括:将植物果实制汁后分装到适宜容器中,在下述条件下进行离心处理,取上清液进行双道超滤分离;之后进行超高压处理,传压介质为水。
其中,所述口服液的加工方法包括以下步骤:
(1)选用新鲜或冷冻无腐烂变质的蓝莓、桑葚、沙棘、刺梨的果实原料;
(2)清洗干净;
(3)将果实进行破碎制汁;
(4)用四层纱布过滤,将皮渣与汁液分离,得到汁液。
(5)离心处理,所述条件为:3000rpm、10min。
(6)超滤分离,所述条件为:管式膜100kD,卷式膜10kD。
(7)超高压处理,所述条件为压力550MPa、时间为5min。
按所述方法制得的具有高SOD酶活力的口服液,其可溶性固形物约为8°Brix,pH为3.7。
实施例2
一种具有高SOD酶活力的口服液的加工方法,包括将蓝莓、猕猴桃、沙棘、刺梨进行制汁、离心、超滤和超高压处理,具体方法包括:将原料制汁后分装到适宜容器中,在下述条件下进行离心处理,取上清液进行超滤分离;之后进行超高压处理。
其中,所述口服液的加工方法包括以下步骤:
(1)选用新鲜或冷冻无腐烂变质的蓝莓、猕猴桃、沙棘、刺梨的果实原料;
(2)清洗干净;
(3)将果实输送至螺旋榨汁机,进行制汁;
(4)用四层纱布过滤,将果核、皮渣与汁液分离,得到原汁;
(5)离心处理,所述条件为:3000rpm、10min。
(6)超滤分离,所述条件为:管式膜100kD,卷式膜10kD。
(7)超高压处理,所述条件为压力550MPa,时间为5min。
按所述方法制得的具有高SOD酶活力的口服液,其可溶性固形物约为10°Brix左右,pH为2.7。
对比例1-1
一种果汁的加工方法,与实施例1的区别仅在于:未采用超滤浓缩和超高压处理。
对比例1-2
一种果汁的加工方法,与实施例1的区别在于:仅采用超滤浓缩处理,未采用超高压处理。
对比例1-3
一种果汁的加工方法,与实施例1的区别在于:未采用超滤浓缩,其他步骤相同。
对比例1-4
一种果汁的加工方法,与实施例1的区别在于:超滤分离条件为:仅采用管式膜100kD,其他步骤相同。
对比例2-1
一种果汁的加工方法,与实施例1的区别仅在于:未采用超滤浓缩和超高压处理。
对比例2-2
一种果汁的加工方法,与实施例1的区别在于:仅采用超滤浓缩处理,未采用超高压处理。
结论1:
分别检验实施例1和对比例1制备的果蔬汁的SOD酶活性。结果见下表1。其中SOD活性检测方法为WST-1法(参考文献Peskin,A.V.,& Winterbourn,C.C.(2000).A microtiter plate assay for superoxide dismutase using a water-soluble tetrazolium salt(WST-1).Clinica Chimica Acta,293(1–2),157-166.)。WST-1即2-(4-Iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium,monosodium salt。WST-1可以和黄嘌呤氧化酶催化产生的超氧化物阴离子反应产生水溶性的甲臜染料(formazan dye),该反应步骤可以被SOD所抑制。通过对WST-1产物的比色分析即可计算SOD的酶活力。
表1
Figure PCTCN2018113302-appb-000001
表1结果表明:超滤浓缩使得SOD酶活力相比对比例1-1提升34%左右,达到9257.4U/mL;超滤处理之后继续使用超高压处理可以激活SOD,使得酶活力相比对比例1-2提升20%左右,超过10000U/mL。说明该制备方法技能显著激活果蔬汁SOD酶活力。
结论2:
分别检验实施例2和对比例2-1、对比例2-2制备的果汁的SOD酶活性。其中SOD活性检测方法实验例1相同。结果见下表2。
表2
Figure PCTCN2018113302-appb-000002
表2结果表明:超滤浓缩处理可以使得果汁SOD酶活力相比对比例2-1提升37%左右,达到8072.9U/mL;超滤浓缩+超高压可以继续激活SOD,使得酶活力相比对比例2-2提升25%左右,达到10091.13U/mL。说明该制备方法能显著激活果蔬汁SOD酶活力。
工业应用
本发明工艺处理可以激活SOD酶,提高口服液中SOD酶的活性,保障产品的保健功能。
本发明采用离心和不同截留分子量双超滤的方式对口服液进行了分离,可以有效提升其酶活力,大大提升了产品的保健功能;本发明采用超高压处理方法激活SOD酶,提高其酶活力;本发明采用的技术相比于传统DEAE离子交换分离技术,具有成本低、效率高、实践性强等优点。

Claims (7)

  1. 一种具有高SOD酶活力的口服液的制备方法,包括下述步骤:
    1)对可食植物原料进行制汁处理,得到植物汁液;
    2)对植物汁液进行离心处理,收集上清液;
    3)对上清液进行双道超滤分离,收集滤液;
    4)对所得滤液进行超高压处理,得到具有高SOD酶活力的液体;
    5)将所述具有高SOD酶活力的液体制成口服液。
  2. 根据权利要求1所述的方法,其特征在于:步骤1)中,所述可食植物原料为可食植物根、茎、叶、果实;
    所述制汁处理的工艺为破碎榨汁。
  3. 根据权利要求1或2所述的方法,其特征在于:步骤2)中,所述离心处理的工艺条件为:1000-4000rpm、0-30min。
  4. 根据权利要求1-3中任一项所述的方法,其特征在于:步骤3)中,所述双道超滤分离中,超滤膜孔径为:管式膜60-120kD,卷式膜8-30kD。
  5. 根据权利要求1-4中任一项所述的方法,其特征在于:步骤4)中,所述超高压处理的压力为:200-600MPa,时间为:1-20min。
  6. 根据权利要求1-5中任一项所述的方法,其特征在于:步骤4)中,所述超高压处理的温度在0-40℃范围内;
    所述超高压处理的传压介质为水。
  7. 通过权利要求1-6中任一项所述的方法制备得到的具有高SOD酶活力的口服液,所述口服液的可溶性固形物为2-15°Brix,pH为2.0-6.0。
PCT/CN2018/113302 2018-08-02 2018-11-01 一种具有高sod酶活力的口服液及其制备方法 WO2020024468A1 (zh)

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