阅读说明：本技术 一种从新鲜虾和/或蟹废弃物中提取天然虾青素的方法 (Method for extracting natural astaxanthin from fresh shrimps and/or crab waste ) 是由 林大昌 叶伟 江艾佳 于 2019-09-20 设计创作，主要内容包括：本发明涉及从虾、蟹废弃物生物处理技术领域,尤其涉及一种从新鲜虾和/或蟹废弃物中制备高含天然虾青素干壳粉的方法。该方法包括以下的步骤：1)将新新鲜虾和/或蟹废弃物粉碎制浆,制得新鲜浆料；2)将新鲜浆料抽入装备超声波装置的酶解反应釜中进行采用混合酶制剂进行酶解,所述的混合酶制剂中加入食品级抗氧化剂,食品级抗氧化剂的加入量为100-5000ppm；3)酶解结速后,高速离心,离心渣用去离子水清洗后,加入酸水脱钙；4)脱钙结速后,高速离心,离心渣30-60℃真空烘干,得含天然虾青素的干壳粉。该方法简便,采用新鲜虾和/或蟹废弃物,可以就地处理,通过混合蛋白酶酶解,超声波辅助能较彻底去除残留物,并在酶解过程中加入食品级抗氧化剂,保护了天然虾青素的氧化,提高了产品中天然虾青素含量。(The invention relates to the technical field of biological treatment of shrimp and crab wastes, in particular to a method for preparing high-natural-astaxanthin-content dry shell powder from fresh shrimp and/or crab wastes. The method comprises the following steps: 1) crushing and pulping the waste of the fresh shrimps and/or the crabs to prepare fresh pulp; 2) pumping fresh slurry into an enzymolysis reaction kettle equipped with an ultrasonic device for enzymolysis by adopting a mixed enzyme preparation, wherein a food-grade antioxidant is added into the mixed enzyme preparation, and the adding amount of the food-grade antioxidant is 100-5000 ppm; 3) after enzymolysis, high-speed centrifugation is carried out, centrifugal slag is washed by deionized water, and acid water is added for decalcification; 4) and after decalcification and hardening, high-speed centrifugation is carried out, and centrifugal slag is dried in vacuum at 30-60 ℃ to obtain dry shell powder containing natural astaxanthin. The method is simple, adopts fresh shrimp and/or crab waste, can be treated on site, can remove residue thoroughly by enzymolysis of mixed protease and ultrasonic wave assistance, and adds food-grade antioxidant during enzymolysis process to protect oxidation of natural astaxanthin and improve natural astaxanthin content in product.)

1. A method for preparing a dry shell powder with high natural astaxanthin content from fresh shrimp and/or crab waste comprises the following steps:

1) crushing fresh shrimps and/or crab wastes for pulping to prepare fresh pulp;

2) pumping fresh slurry into an enzymolysis reaction kettle equipped with an ultrasonic device for enzymolysis by adopting a mixed enzyme preparation, wherein a food-grade antioxidant is added into the mixed enzyme preparation, and the adding amount of the food-grade antioxidant is 100-5000 ppm;

3) after enzymolysis, high-speed centrifugation is carried out, centrifugal slag is washed by deionized water, and acid water is added for decalcification;

4) and after decalcification is finished, centrifuging at a high speed, and drying centrifugal slag at 30-60 ℃ in vacuum to obtain dry shell powder containing natural astaxanthin.

2. The method of producing a dry shell meal high in natural astaxanthin from fresh shrimp and/or crab waste according to claim 1, wherein the animal source of the fresh shrimp and/or crab waste comprises one or more of sea shrimp, freshwater shrimp, crayfish and crab; the waste is in the form of one or more of shrimp head, rejected shrimp, shrimp leg, shrimp shell, crab leg, crab shell and rejected crab.

3. The method for preparing the dry shell powder with high natural astaxanthin content from the waste of fresh shrimps and/or crabs according to claim 1, wherein the power of an ultrasonic device is as follows: 50W-250W, frequency: 15kHz-50 kHz; the fresh slurry is 5-10 mesh in sheet shape, and has water content of 50-55%.

4. The method for preparing the dry shell powder with high natural astaxanthin content from the fresh shrimp and/or crab wastes as claimed in claim 1, wherein the solid-to-liquid ratio of enzymolysis is 1: 1.5 ~ 1:5, the pH is adjusted to be 7.0-8.0, the enzyme dosage is 5000-.

5. The method for preparing the dry shell powder with high natural astaxanthin content from the fresh shrimps and/or crabs waste according to the claim 4 or 5, characterized in that the mixed enzyme preparation comprises papain, trypsin and lipase, the papain, the trypsin and the lipase are mixed according to the proportion of U/g: 1-2: 0.2-0.5: 0.1-0.3.

6. The method for preparing the dry shell powder with high natural astaxanthin content from the fresh shrimp and/or crab wastes as claimed in claim 1, wherein the acid water is decalcified and added with acid water while adding food-grade antioxidant 100-2000 ppm; the acid water is low-concentration HCl or organic acid.

7. The method for preparing the dry shell powder with high natural astaxanthin content from the waste of fresh shrimps and/or crabs as claimed in claim 1 or 6, wherein the food grade antioxidant is one or more of vitamin E, vitamin C, L-sodium ascorbate, BHA, PG and BHT.

8. The method for preparing the dry shell powder with high natural astaxanthin content from the fresh shrimps and/or crabs waste according to claim 1, wherein after the high-speed centrifugation in the step 3), the centrifugate is transferred to a protein extraction process, and the protein extraction process comprises the following steps:

stirring the enzymolysis centrifugate at 100 deg.C for 0.5h, controlling temperature at 60 deg.C, adding 5% (w: v) chitosan-cellulose-activated carbon composite adsorption resin, stirring for 1h for decolorizing, removing fishy smell, removing heavy metal ions, and removing pesticide residue and harmful substances;

microfiltering to remove particle impurities while the solution is hot;

vacuum concentrating to solid content not less than 10%, and spray drying to obtain concentrated protein powder.

9. The method for preparing the dry shell powder with high natural astaxanthin content from the waste of fresh shrimps and/or crabs according to claim 1, wherein after the high speed centrifugation in the step 4), the centrifugate containing hydrochloric acid is added to K₂CO₃Adjusting Ph to 6-6.5, introducing CO₂White precipitate is generated, high-speed filtration and centrifugation are carried out, the centrifugal slag is dried in vacuum, the temperature is controlled to be less than or equal to 80 ℃, the water content is less than or equal to 10 percent, the finished product of the bioactive calcium is obtained, and the centrifugate is transferred into liquid fertilizer water;

or, after high-speed centrifugation in the step 4), carrying out microfiltration on the centrifugate containing the organic acid to remove particle impurities, carrying out vacuum concentration to obtain organic bioactive calcium powder with the solid content of more than or equal to 10 percent, and carrying out spray drying to obtain the organic bioactive calcium powder;

further, the method also comprises the preparation steps of the chitosan oligosaccharide liquid organic fertilizer:

A. mixing the cleaning water for cleaning the shrimp heads and the residues with the cleaning water of the reaction kettle; adding chitosan flocculant for precipitation, taking supernatant, passing through anion and cation resins, and recycling as cleaning water;

B. mixing the flocculation solution and the bioactive calcium centrifugate, detecting N, P, K and essential trace element content, adjusting Ph6.5-7.0 according to the national standard of liquid fertilizer, adding N, P, K and other essential trace element organic compounds which are easily soluble in water, stirring thoroughly, adding 2.5-8% (w/v) chitosan oligosaccharide, and stirring thoroughly;

C. adding large-particle chitosan-cellulose-activated carbon composite adsorption resin, stirring, removing heavy metal, harmful metal ions, pesticide residue and harmful substances, adding 0.5-1% (w/v) potassium sorbate, high-speed centrifuging, sieving with 200-250 mesh sieve, and filling into a barrel filled with nitrogen or vacuumizing.

10. Dry shell flour containing natural astaxanthin obtained by the process according to any one of claims 1 ~ 10.

Technical Field

The invention relates to the technical field of biological treatment of shrimp and crab wastes, in particular to a method for extracting natural astaxanthin from fresh shrimp and/or crab wastes.

Background

With the rapid increase of aquaculture and processing enterprises, the leftovers generated in the processing process are more and more, and are also serious pollution sources; take Zhejiang as an example; when manufacturing enterprises taking seawater shrimps or freshwater shrimps and crayfish as main raw materials process the leftovers, one part of the leftovers are sold as waste materials, the leftovers are hundreds of yuan per ton, or the leftovers are conveyed to a farm to be dried and crushed to be used as feed, and a plurality of leftovers which are not processed in time are piled up to be mildewed and rotten, so that the environment is seriously polluted, and finally the leftovers are directly conveyed to a garbage yard. The byproducts contain rich nutrients such as protein, astaxanthin and unsaturated fatty acid, and mineral salts such as calcium, magnesium and phosphorus, and nutrient components such as cephalin, lecithin, carotenoid, carbohydrate, cellulose and vitamin, and the shrimp byproducts mainly refer to leftovers (shrimp heads and rejected shrimps) of products such as shrimp meat and shrimp tails obtained by screening and processing raw material shrimps, and the leftovers contain a lot of residual meat. At present, many reports have been made on mixing a small amount of crushed shrimp heads, shrimp shells, etc. with fish meal as feed and bait, and extracting functional nutrients from the shrimp heads and the shrimp shells. A great amount of shrimp meat, shrimp brain and shrimp yellow are remained on the heads and the feet of the crayfish, the residues have higher utilization value, and the shrimp heads of the shrimp shells after meat collection can be further processed into biological protein calcium powder, astaxanthin, chitin, chitosan and the like.

At present, a large amount of crustacean waste of shrimp and crab processing industry is generated at home and abroad every year, and the extraction and recovery of astaxanthin from crustacean waste become one of the main ways for producing natural astaxanthin.

Astaxanthin is mainly present in the shells of crustaceans (such as shrimps and crabs) in 3 forms: (1) a protein-bound form; (2) in the form of an esterification; (3) in free form.

The free state is the least stable, the esterified form exists the most, and the astaxanthin in the esterified form is mainly effectively separated and purified in the actual extraction process. In recent years, inorganic acid or organic acid is added in the treatment process of countries such as Norway, the combination of astaxanthin and protein or skeleton parts is destroyed, the release amount of astaxanthin is increased, the recovery rate can reach 180 mug/g, the purity is greatly improved, but the market demand cannot be met.

Natural extract from crayfish headThe content of astaxanthin and fresh crayfish head is 25-30 mug/g, the traditional process adopts integral drying and then crushing, the oxidation loss of the astaxanthin reaches 15-20 percent due to the drying and crushing temperature, no matter the alkali extraction method, the organic solvent method and the supercritical CO method₂Extracting with adjuvant (alkali, oil, organic solvent, CO)₂) And the energy consumption is 3 times of that of the method; as the crayfish is a seasonal product (4-9 months), the oxidation loss of the natural astaxanthin reaches 30-40 percent in the conventional storage and transportation.

The invention patent (publication number: CN1715255A, published: 2006.01.04) applied by the applicant discloses a method for producing chitin, astaxanthin, protein, calcium powder and biological fertilizer by using shrimp shells, which comprises the following steps: crushing shrimp shells, adding a complex enzyme for enzymolysis, adding an organic solvent for extraction, separating into a shrimp liquid and a shrimp solid after full reaction, separating the shrimp liquid into a hydrolysate and an organic liquid, and performing vacuum concentration on the organic liquid containing astaxanthin to extract crude astaxanthin.

The Chinese invention patent application (publication No. CN108559765A, published: 2017, 12 and 28) discloses a method for extracting N-acetylglucosamine and astaxanthin from crayfish shells by a biological enzyme method. The method comprises the following steps: pretreating the crayfish shell to obtain crayfish shell powder; taking chitinase-producing microorganisms as test bacteria, inoculating the test bacteria to a seed culture medium for culture to obtain a seed solution, transferring the seed solution to a fermentation culture medium for fermentation, and purifying to obtain a crude pure enzyme solution; putting the crayfish shell powder into a reaction kettle, sequentially adding the crude pure enzyme solution and the organic reagent, stirring while keeping out of the sun, and heating to 20-37 ℃ at constant temperature to obtain an enzymatic hydrolysate; standing and layering the enzymolysis liquid, pouring out the upper layer liquid, and recovering the organic reagent and the shrimp sauce rich in astaxanthin under dark.

The method adopts the shrimp shell powder for enzymolysis and then adopts an organic reagent for extraction to obtain the astaxanthin, but the process steps are relatively complicated, and the oxidation loss of the astaxanthin is relatively large in the process. In addition, the complex process conditions are not beneficial to the on-site treatment of the raw materials, so that the centralized transportation and centralized treatment are needed; this in turn often leads to astaxanthin oxidation losses during transport and intensive handling and also to costs for inventory and logistics and environmental pollution from waste products during production.

Disclosure of Invention

In order to solve the technical problems, the invention aims to provide a method for extracting natural astaxanthin from fresh shrimp and/or crab wastes, which is simple and convenient, adopts the fresh shrimp and/or crab wastes, can be treated on site, can remove residues more thoroughly through mixed protease enzymolysis and ultrasonic wave assistance, and adds a food-grade antioxidant in the enzymolysis process, thereby protecting the oxidation of natural astaxanthin and improving the content of natural astaxanthin in products.

In order to achieve the purpose, the invention adopts the following technical scheme:

a method for extracting natural astaxanthin from fresh shrimp and/or crab waste, comprising the steps of:

1) crushing and pulping the waste of the fresh shrimps and/or the crabs to prepare fresh pulp;

2) pumping fresh slurry into an enzymolysis reaction kettle equipped with an ultrasonic device for enzymolysis by adopting a mixed enzyme preparation, wherein a food-grade antioxidant is added into the mixed enzyme preparation, and the adding amount of the food-grade antioxidant is 100-5000 ppm;

3) after enzymolysis, high-speed centrifugation is carried out, centrifugal slag is washed by deionized water, and acid water is added for decalcification;

4) and after decalcification and hardening, high-speed centrifugation is carried out, and centrifugal slag is dried in vacuum at 30-60 ℃ to obtain dry shell powder containing natural astaxanthin.

Preferably, the animal source of fresh shrimp and/or crab waste comprises one or more of sea shrimp, freshwater shrimp, crayfish and crab; the waste is in the form of one or more of shrimp head, rejected shrimp, shrimp leg, shrimp shell, crab leg, crab shell and rejected crab. Most preferred herein are crawfish heads and crawfish tails.

Preferably, the power of the ultrasonic device: 50W-250W, frequency: 15kHz-50 kHz; the fresh slurry is 5-10 mesh in sheet shape, and has water content of 50-55%.

Preferably, the solid-liquid ratio of enzymolysis is 1: 1.5 ~ 1:5, the pH is adjusted to be 7.0-8.0, the enzyme dosage is 5000-10000U/g, and the enzymolysis is carried out for 4.0-6.0h at 50-60 ℃.

Preferably, the mixed enzyme preparation comprises papain, trypsin and lipase, wherein the papain, the trypsin and the lipase are mixed according to the proportion of U/g: 1-2: 0.2-0.5: 0.1-0.3.

Preferably, the acid water is decalcified and added with acid water, and simultaneously added with food-grade antioxidant 100-2000ppm, and the acid water adopts low-concentration HCl or organic acid.

Preferably, the food grade antioxidant is one or more of vitamin E, vitamin C, L-sodium ascorbate, BHA, PG and BHT.

As a further improvement, after the high-speed centrifugation in the step 3), the centrifugate is transferred to a protein extraction process, and the protein extraction process comprises the following steps:

1.1) stirring the enzymolysis centrifugate at 100 ℃ for 0.5h, controlling the temperature at 60 ℃, adding 5% (w: v) of chitosan-cellulose-activated carbon composite adsorption resin, stirring for 1h, and decoloring, deodorizing, removing heavy metal ions, residual and harmful substances;

1.2) microfiltration is carried out to remove particle impurities while the solution is hot;

1.3) concentrating in vacuum to obtain concentrated protein powder with solid content not less than 10%.

As a further improvement, after high-speed centrifugation in the step 4), adding K2CO3 into the hydrochloric acid-containing centrifugate to adjust Ph6-6.5, introducing CO2 to generate white precipitate, carrying out high-speed filtration and centrifugation, carrying out vacuum drying on the centrifugal slag, controlling the temperature to be less than or equal to 80 ℃, and controlling the moisture to be less than or equal to 10% to obtain finished bioactive calcium, and transferring the centrifugate into liquid fertilizer water;

or, after high-speed centrifugation in the step 4), microfiltration is carried out on the centrifugate containing the organic acid to remove particle impurities, the solid content is more than or equal to 10 percent through vacuum concentration, and the organic bioactive calcium powder is prepared through spray drying.

As a further improvement, the method also comprises the preparation steps of the chitosan oligosaccharide liquid organic fertilizer:

A. mixing the cleaning water for cleaning the shrimp heads and the residues with the cleaning water of the reaction kettle; adding chitosan flocculant for precipitation, taking supernatant, passing through anion and cation resins, and recycling as cleaning water;

B. mixing the flocculation solution and the bioactive calcium centrifugate, detecting N, P, K and essential trace element content, adjusting Ph6.5-7.0 according to the national standard of liquid fertilizer, adding N, P, K and other essential trace element organic compounds which are easily soluble in water, stirring thoroughly, adding 2.5-8% (w/v) chitosan oligosaccharide, and stirring thoroughly;

C. adding large-particle chitosan-cellulose-activated carbon composite adsorption resin, stirring, removing heavy metal, harmful metal ions, pesticide residue and harmful substances, adding 0.5-1% (w/v) potassium sorbate, high-speed centrifuging, sieving with 200-250 mesh sieve, and filling into a barrel filled with nitrogen or vacuumizing.

The application also discloses the method for preparing the dry shell powder containing the natural astaxanthin.

The method is simple and convenient due to the adoption of the technical scheme, the waste of fresh shrimps and/or crabs can be treated on site, residues can be thoroughly removed through enzymolysis of mixed protease and ultrasonic assistance, the food-grade antioxidant is added in the enzymolysis process, the oxidation of natural astaxanthin is protected, the natural astaxanthin content in the product is improved, the natural astaxanthin content of the dry shell powder is 200-250 mu g/g, and the dry shell powder can be directly added and used as feed (the application is disclosed as CN 108208313A) or used as a raw material for astaxanthin extraction.

In addition, according to the method, residual meat contained in leftovers after hydrolysis by mixed protease is prepared into concentrated protein powder, the shrimp shell is prepared into biological calcium powder by organic acidification, process wastewater and washing water are mixed to prepare a chitosan liquid fertilizer (plant nutrient solution), the process conditions are further optimized, and the dry shell powder rich in natural astaxanthin is prepared, so that the waste resource utilization of enterprises is realized, the economic benefit is greatly increased, and the pollution of waste to the environment is effectively reduced; realizing zero discharge of sewage.

Drawings

FIG. 1 is a HPLC check chart of astaxanthin standard control solution.

FIG. 2 is a HPLC check chart of a test solution of astaxanthin from shrimp meal.

Detailed Description