阅读说明：本技术 一种鲨鱼软骨粉替代品及其制备方法 (Shark cartilage powder substitute and preparation method thereof ) 是由 付鹏磊 王冠丹 孙倩 王晓瞳 于 2020-05-26 设计创作，主要内容包括：本发明公开了一种鲨鱼软骨粉替代品及其制备方法,属于海洋生物技术领域。本申请以鲨鱼软骨粉和鲟鱼骨为原料,集成生物酶解技术、混菌微生物发酵技术、现代生物分离技术,研制开发出具有突出免疫增强的鲨鱼软骨粉替代品,其中生物酶解技术采用的复合酶系为木瓜蛋白酶、胰蛋白酶、中性蛋白酶和胃蛋白酶的中的一种或几种的混合物,混菌微生物发酵的发酵菌种主要包括地衣芽孢杆菌和枯草芽孢杆菌。利用本发明工艺制备的鲨鱼软骨粉替代品硫酸软骨素含量高,且具有较高的抗氧化能力和显著的抗肿瘤功效,可广泛应用于补钙和预防肿瘤等医用食品中。(The invention discloses a shark cartilage powder substitute and a preparation method thereof, belonging to the technical field of marine organisms. According to the method, shark cartilage powder and sturgeon bone are used as raw materials, a biological enzymolysis technology, a mixed-bacteria microbial fermentation technology and a modern biological separation technology are integrated, and a shark cartilage powder substitute with outstanding immunity enhancement is developed and developed, wherein a compound enzyme system adopted by the biological enzymolysis technology is one or a mixture of papain, trypsin, neutral protease and pepsin, and a fermentation strain fermented by the mixed-bacteria microbial mainly comprises bacillus licheniformis and bacillus subtilis. The shark cartilage powder substitute prepared by the process has high chondroitin sulfate content, high oxidation resistance and obvious anti-tumor effect, and can be widely applied to medical foods for calcium supplement, tumor prevention and the like.)

1. The preparation method of the shark cartilage powder substitute is characterized by comprising the following steps:

(1) pretreatment: crushing shark cartilage powder and sturgeon bone powder, sieving with a 100-mesh sieve, adding a sodium hydroxide solution and a sodium oxide solution, extracting at 40-50 ℃ for 12-13 h, filtering to obtain a filtrate, and adjusting the pH value of the filtrate to 7.6-7.9;

(2) compound enzymolysis: adding a complex enzyme preparation with the mass ratio of 1-2% of shark cartilage powder into the filtrate obtained in the step (1), and carrying out enzymolysis for 4-6 h at 40-60 ℃;

(3) preparing fermentation liquor: adding 2 percent of glucose and 0.15 to 0.20 percent of (NH) into the enzymolysis liquid by mass ratio₄)₂SO₄、0.10～0.15% KH₂PO₄、0.1%MgSO₄Preparing a liquid fermentation culture medium;

(4) and (3) sterilization: sterilizing at 121 deg.C for 15 min;

(5) aerobic fermentation: adding a compound microbial agent with the mass ratio of 2-3% of the fermentation liquor into the fermentation liquor, and carrying out aerobic fermentation for 12-24 h at the temperature of 28-35 ℃;

(6) and (3) filtering: filtering to remove thallus and insoluble solid after fermentation;

(7) concentrating and drying: and (4) drying the fermentation liquor filtered in the step (6) to obtain the shark cartilage powder substitute.

2. The preparation method of the shark cartilage powder substitute according to claim 1, wherein the mass ratio of the shark cartilage powder to the sturgeon bone is 1: 3-5.

3. The preparation method of the shark cartilage powder substitute according to claim 1, wherein the concentration of the sodium hydroxide solution in step (1) is 1-1.5%, the concentration of the sodium oxide solution is 5-6%, and the addition amount of the sodium hydroxide solution and the addition amount of the sodium oxide solution are both 2.5-3.5 times of the sum of the weight of the shark cartilage and the weight of the sturgeon bone.

4. The method for preparing a shark cartilage powder substitute according to claim 1, wherein the complex enzyme preparation in step (2) is one or more of papain, trypsin, neutral protease and pepsin.

5. The method for preparing a shark cartilage powder substitute according to claim 1, wherein the complex enzyme preparation in step (2) is a mixture of papain and pepsin in a mass ratio of 2: 1.

6. The method for preparing a shark cartilage powder substitute according to claim 1, wherein 0.5-1% chitosan is added during the preparation of the fermentation broth in step (3).

7. The method for preparing a shark cartilage powder substitute according to claim 1, wherein the complex microorganism in step (5) is a mixture of Bacillus licheniformis and Bacillus subtilis at an inoculation ratio of 2: 3.

8. The method for preparing shark cartilage powder substitute according to claim 5, wherein the method for preparing the composite microbial inoculum seed liquid comprises: under the aseptic condition, LB culture medium is adopted to culture for 24 hours at the temperature of 28-32 ℃, liquid bacillus licheniformis seed liquid and bacillus subtilis seed liquid are respectively prepared, and the effective viable count of the seed liquid is required to be more than or equal to 10⁹cfu/mL; then mixing the seed solutions of the two kinds of bacillus proportionally.

9. A shark cartilage powder substitute as claimed in claim 1.

10. Use of a shark cartilage powder substitute according to claim 9 in food products.

Technical Field

The invention relates to a shark cartilage powder substitute and a preparation method thereof, in particular to a novel shark cartilage powder substitute product with an immune enhancement effect prepared by the synergy of biological enzymolysis and microbial fermentation, belonging to the technical field of marine organisms.

Background

Shark is one of the oldest and longest-lived deep sea marine organisms in the world, has survived for four hundred million years on earth, has a perfect immune system, is extremely strong in vitality, is hardly infected by diseases, can recover wounds quickly even after being subjected to trauma, and has a cancer probability of about 1/million. The shark medicine in China is originally seen in the book Ben Cao Jing Ji Zhu, the cartilage of the shark medicine mainly comprises cartilage cells, collagen fibers, a large amount of proteoglycan and extracellular matrix, and the polysaccharide extracted from the shark cartilage is mainly used for clinically treating neuralgia, migraine, pain after abdominal cavity operation and the like. In developed countries such as Europe, America, Japan, etc., the shark cartilage polysaccharide is used as health food or health medicine for preventing and treating coronary heart disease, angina pectoris, myocardial infarction, etc. for a long time, and has no obvious toxic and side effects. In recent years, researches show that the shark cartilage contains various mucopolysaccharides and chondroitin sulfate besides minerals such as calcium, phosphorus, magnesium and the like, has remarkable anticoagulation, blood fat reduction, antiviral, anti-tumor and other functions, and is an unavailable natural food resource. Because the shark cartilage powder has the natural effects of promoting the regeneration of human cartilage and inhibiting new vessels, the shark cartilage powder is an excellent immunopotentiating raw material. In recent years, development of shark fin bone meal health food has been rapidly advanced, but due to the restriction of shark resource amount, its wide application in the fields of medicine, food and the like is restricted. The invention provides a shark cartilage powder substitute and a preparation method thereof, aiming at solving the technical problems.

Disclosure of Invention

In order to overcome the defects of the prior art and realize the efficient extraction and the wide application of functional active substances of marine biological resources such as shark cartilage powder and the like, the application provides a preparation method of a shark cartilage powder substitute, integrates the modern bioseparation technology, the biological enzymolysis technology, the mixed microbial fermentation technology and the marine biological resource nutrition allocation technology, develops and develops the shark cartilage powder substitute with high immunity enhancement capability, meets the application requirements of the fields of health care, medical treatment, food and the like on the shark cartilage powder substitute, has high yield of chondroitin sulfate products prepared by the process, has higher oxidation resistance and obvious anti-tumor effect, and can be widely applied to medical food.

The invention realizes the technical effects through the following technical scheme:

a preparation method of shark cartilage powder substitute comprises the following steps:

(1) pretreatment: crushing shark cartilage powder and sturgeon bone powder, sieving with a 100-mesh sieve, adding a sodium hydroxide solution and a sodium oxide solution, extracting at 40-50 ℃ for 12-13 h, filtering to obtain a filtrate, and adjusting the pH value of the filtrate to 7.6-7.9;

(2) compound enzymolysis: adding a complex enzyme preparation with the mass ratio of 1-2% of shark cartilage powder into the filtrate obtained in the step (1), and carrying out enzymolysis for 4-6 h at 40-60 ℃;

(3) preparing fermentation liquor: adding 2 percent of glucose and 0.15 to 0.20 percent of (NH) into the enzymolysis liquid by mass ratio₄)₂SO₄、0.10～0.15% KH₂PO₄、0.1%MgSO₄Preparing a liquid fermentation culture medium;

(4) and (3) sterilization: sterilizing at 121 deg.C for 15 min;

(5) aerobic fermentation: adding a compound microbial agent with the mass ratio of 2-3% of the fermentation liquor into the fermentation liquor, and carrying out aerobic fermentation for 12-24 h at the temperature of 28-35 ℃;

(6) and (3) filtering: filtering to remove thallus and insoluble solid after fermentation;

(7) concentrating and drying: and (4) drying the fermentation liquor filtered in the step (6) to obtain the shark cartilage powder substitute.

The mass ratio of the shark cartilage powder to the sturgeon bone in the step (1) is 1: 3-5.

In the step (1), the concentration of the sodium hydroxide solution is 1-1.5%, the concentration of the sodium oxide solution is 5-6%, and the adding amount of the sodium hydroxide solution and the sodium oxide solution is 2.5-3.5 times of the sum of the weight of the shark cartilage and the weight of the sturgeon bone.

The complex enzyme preparation in the step (2) is one or a mixture of more of papain, trypsin, neutral protease and pepsin.

Preferably, the complex enzyme preparation in the step (2) is a mixture of papain and pepsin, and the mass ratio of the papain to the pepsin is 2: 1.

Preferably, 0.5-1% of chitosan is added in the preparation process of the fermentation liquid in the step (3).

The compound microorganism in the step (5) is a mixed bacterium of bacillus licheniformis and bacillus subtilis, and the inoculation amount ratio of the compound microorganism is 2: 3.

The preparation method of the composite microbial inoculum seed liquid comprises the following steps: under the aseptic condition, LB culture medium is adopted to culture for 24 hours at the temperature of 28-32 ℃, liquid bacillus licheniformis seed liquid and bacillus subtilis seed liquid are respectively prepared, and the effective viable count of the seed liquid is required to be more than or equal to 10⁹cfu/mL; then mixing the seed solutions of the two kinds of bacillus proportionally.

The invention provides a preparation method of shark cartilage powder substitute, the obtained shark cartilage powder substitute has better immunity enhancing effect than shark cartilage powder, and can be widely applied to medical treatment, food and health products.

The invention provides a preparation method of a shark cartilage powder substitute, which has the following remarkable advantages compared with the prior art:

(1) the method utilizes aerobic bacteria (bacillus licheniformis and bacillus subtilis) to jointly ferment the enzymatic hydrolysate, fully utilizes the synergistic effect among microorganisms, and improves the immunity enhancing effect of the shark cartilage powder substitute; the multi-strain mixed fermentation not only improves the yield of the shark cartilage powder substitute, but also improves the immunity enhancing capability of the shark cartilage powder substitute through the fermentation effect among microorganisms;

(2) the experimental result shows that the addition of a certain amount of chitosan in the fermentation liquor can obviously improve the nutrition and health-care activity of the shark cartilage powder substitute;

(3) the invention adopts enzymolysis and mixed bacteria microbial fermentation to prepare the shark cartilage powder substitute, which obviously improves the extraction rate and the product nutrition and health care effect of the shark cartilage powder substitute.

Detailed Description

The activation method of the compound microbial agent in the following examples is prepared by adopting the following processes: under the aseptic condition, LB culture medium is adopted to culture for 24 hours at the temperature of 28-32 ℃, liquid bacillus licheniformis seed liquid and bacillus subtilis seed liquid are respectively prepared, and the effective viable count of the seed liquid is required to be more than or equal to 10⁹cfu/mL; then, the seed solutions of the two kinds of bacillus are mixed according to the proportion of 2: 3.