CN104357206A - Method for preparing phospholipid-rich Antarctic krill oil by water enzyme process - Google Patents
Method for preparing phospholipid-rich Antarctic krill oil by water enzyme process Download PDFInfo
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- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
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- C11B1/025—Pretreatment by enzymes or microorganisms, living or dead
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- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
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- C11B1/10—Production of fats or fatty oils from raw materials by extracting
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- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P21/00—Preparation of peptides or proteins
- C12P21/06—Preparation of peptides or proteins produced by the hydrolysis of a peptide bond, e.g. hydrolysate products
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Abstract
The invention discloses a method for preparing phospholipid-rich Antarctic krill oil by a water enzyme process. The method is implemented by enzymolysis with autoenzyme in combination with proteinase and comprises the following steps: carrying out proteinase enzymolysis to liquefy Antarctic krill flesh, centrifuging to remove shells, carrying out enzymolysis to destroy the composite structure of proteins and phospholipids, isolating the phospholipids, and centrifuging to separate the phospholipid-rich layer; and drying the phospholipid-rich layer, and extracting with ethanol to obtain the Antarctic krill oil with the phospholipid content of greater than 40 w%, Antarctic krill peptides and Antarctic krill proteins. The method is suitable for shipboard operation; and the semifinished product can be prepared firstly, and then transported to the shore for subsequent processing, thereby reducing the proteinase consumption and lowering the transportation cost. Double enzymolysis is utilized to destroy the composite structure of the proteins and phospholipids, so that the phospholipids are isolated, and thus, the extraction rate of the Antarctic krill oil is higher.
Description
Technical field
The invention belongs to manufacturing processing technic technical field, relate to the method that the antarctic krill oil of phosphatide is rich in the preparation of a kind of aqueous enzymatic method.
Background technology
Krill (
euphausia superba) be the crustaceans zooplankton of one way of life at Southern Oceans, take algae as food.Krill grows up to and reaches 6 cm long, 2 grammes per square metres.Biomass energy estimates at 4 ~ 1,500,000,000 tons, year amount of fishing about 1.3 hundred million tons.Krill water content accounts for 77.9 ﹪ ~ 83.1 ﹪, and crude protein accounts for 11.9 ﹪ ~ 15.4 ﹪, and total fat accounts for 0.5 ﹪ ~ 3.6 ﹪.Sex, the maturity and catching season etc. of total lipid content and krill are relevant.
Soevik & Braekkan reported first in 1979 problem of krill Oil repellent exception, Oil repellent average out to 2400ppm/DW, and also fluorine mainly concentrates in shrimp shell, and therefore early stage shelling contributes to reducing the Oil repellent in product.
Because krill is lived in the seawater of about 0 degree, its autolytic enzyme comprises proteolytic enzyme, and Phospholipid hydrolase still keeps very high activity when low temperature, processes not in time if fish for up, and product quality is just difficult to ensure.
Antarctic krill oil contains the Omega-3 polyunsaturated fatty acid (as EPA and DHA) of phosphatide type, has the effects such as reducing blood-fat, hypotensive, hypoglycemic, arthritis, simultaneously also containing astaxanthin and vitamin A, and the anti-oxidation health compositions such as vitamin-E.Human clinical trial shows, EPA and DHA(of phosphatide type comes from antarctic krill oil) bioavailability come from fish oil higher than EPA and DHA(of triglyceride type).
The preparation method of antarctic krill oil roughly has following several:
The invention of application number CN102071101A " a kind of extract the method being rich in the krill oil of phosphatide from krill ", the method take krill as raw material, first utilize the proteases for decomposing release lipid of self, through rapid heating, the enzyme in krill body is lost activity, then cryodrying, take ethanol as solvent, extract the antarctic krill oil being rich in timnodonic acid (EPA) and docosahexenoic acid (DHA) phosphatide.The method is also utilize proteolytic enzyme to discharge lipid, but oneself protease cannot abundant enzymolysis protein-lipid complex, and dissociate phosphatide, so phospholipids content only accounts for 32 ~ 34 ﹪ of antarctic krill oil.
The invention " for obtaining the solvent-free extraction method of the krill oil being rich in phosphatide and neutral grease " of application number CN102638998A, the method comprises: heat boiling krill, meanwhile, avoid stirring and grinding; The krill that decantation separated heating is crossed is to obtain solid and the liquid of part degrease and dehydration; Extrude the solid obtained and squeeze liquid and solid part to obtain; Centrifugation squeezing liquid is to obtain the antarctic krill oil being rich in phosphatide; Centrifugation liquid is rich in the antarctic krill oil of neutral grease and dense thick juice to obtain.The method does not use solvent, but uses milling process to obtain antarctic krill oil, therefore no solvent residue.But the antarctic krill oil that milling process obtains, mainly based on triglyceride level, phospholipids content is not high.
The invention " method extracting shrimp sauce is leached in the expanded granulation of a kind of euphausia superba powder " of application number CN102952625A, it is characterized in that, comprise the steps: 1, get euphausia superba powder and mix with the plant material powder of starch or rich in starch, wherein, the mass content of described euphausia superba powder is 50 ~ 100 ﹪; 2, moisture adjustment is carried out to mixture, to water content 10 ~ 40 ﹪; 3, be input in extruding puffing tablets press by mixture and carry out expanded granulation, prilling temperature is 80 ~ 140 DEG C, die pressure 10 ~ 120Bar, obtains 2 ~ 10mm length puffing material grain; 4, described puffing material grain moisture is regulated, to water content 4 ~ 12 ﹪ with dryer; 5, by described puffing material grain input oil extraction equipment, antarctic krill oil is extracted.The method uses extruding and puffing technology, makes the abundant sex change of protein and forms cell texture, being convenient to oil extraction.But due in high temperature and high pressure environment, the problem of antarctic krill oil oxidation can be there is.
Aqueous enzymatic method just day by day receives publicity as a kind of emerging oil-producing technique and payes attention to, and it is mainly on the basis of Mechanical Crushing, adopts enzymolysis process release grease, utilizes non-oil component different from the avidity difference of water and profit proportion non-oil component and separating of oil to oil; Compare with traditional technology, its mild condition, operational path is simple, and can extract oil and peptide simultaneously.
Summary of the invention
The object of the invention is to overcome above-mentioned Problems existing, provide the method that the antarctic krill oil of phosphatide is rich in the preparation of a kind of aqueous enzymatic method, the method also can obtain krill peptide and krill albumen simultaneously, is particularly suitable for ship operates.
For solving the problems of the technologies described above, the technical solution used in the present invention is as follows:
The present invention in conjunction with added proteins enzyme enzymolysis, first by protease hydrolyzed liquefaction krill meat, centrifugally slough shrimp shell, then enzymolysis destroys the composite structure of albumen and phosphatide, make phosphatide free out, then centrifugation goes out to be rich in phospholipid layer by autolytic enzyme; Be rich in phospholipid layer through super-dry, alcohol extraction obtains phospholipids content and is greater than the antarctic krill oil of 40w ﹪, krill peptide and krill albumen.
The inventive method specifically comprises the following steps:
Step (1), fresh krill or freezing krill are crushed to particle diameter are less than or equal to 10mm;
Step (2), the water krill after step (1) fragmentation and salts contg being less than 1w ﹪ are that 1:0.5 ~ 1.5 mix according to weight ratio, then 1500U/g ~ 4500 U/g protease hydrolyzed is added, be warming up to 50 ~ 65 DEG C, insulation reaction 15 ~ 60 minutes, obtains enzymolysis solution I;
Described proteolytic enzyme is the Sumizyme MP that the neutral protease that produces of subtilis (Bacillus subtilis) or Bacillus licheniformis (Bacillus licheniformis) produce;
The enzymolysis solution I that step (3), step (2) obtain is after 50 ~ 200 order filter-cloth filterings, centrifugal under centrifugal force 350 ~ 500G, and removing solid (i.e. shrimp shell), is cooled to 50 ~ 55 DEG C, then carries out secondary enzymolysis, obtain enzymolysis solution II;
If what step (2) adopted is proteolytic enzyme is Sumizyme MP, step (3) secondary enzymolysis process must by enzymolysis solution I adjust ph after process, specifically regulate enzymolysis solution I pH value to 7.5 ~ 8.5 with sodium hydroxide, be incubated after 3 ~ 6 hours, use salt acid for adjusting pH value to 6.0 ~ 7.0 again, obtain enzymolysis solution II;
If what step (2) adopted is proteolytic enzyme is neutral protease, the enzymolysis solution I in step (3) secondary enzymolysis process after process need not adjust ph, is directly incubated 3 ~ 6 hours, obtains enzymolysis solution II;
The enzymolysis solution II to 70 ~ 90 DEG C that step (4), heating steps (3) obtain, is incubated 5 ~ 30 minutes, produces precipitation; Under being deposited in centrifugal force 500 ~ 700G centrifugal 5 ~ 20 minutes, that isolates upper strata was rich in phospholipid layer PEF(Phospholipid Enriched Fraction) and the peptide liquid layer of lower floor; By peptide liquid layer at 60 ~ 80 DEG C, vacuum tightness is krill peptide under-0.05 ~-0.1MPa after concentrating under reduced pressure;
Step (5), phospholipid layer PEF cryodrying will be rich in or lyophilize to moisture content is less than 10w ﹪;
The requirement of described low temperature is lower than 50 DEG C;
Step (6), obtain driedly being rich in phospholipid layer PEF by alcohol extraction step (5), extraction temperature is 30 ~ 60 DEG C, and extraction time is 1 ~ 3 hour, constantly stirs in extraction process, is extracted liquid;
The solid-liquid ratio being rich in phospholipid layer PEF and ethanol after described drying is 1:5 ~ 15, and unit is kg:L;
Described alcohol concn is more than or equal to 95.5 ﹪ (v/v);
Step (7), under centrifugal force 500 ~ 700G, centrifugation or vacuum tightness carry out suction filtration to the extraction liquid that step (6) obtains under-0.05 ~-0.1MPa, obtain solid residue and filtrate; Described solid residue is krill albumen, and filtrate is scarlet;
The filtrate that step (8), step (7) obtain is at 30 ~ 60 DEG C, and vacuum tightness is rotary evaporation under-0.05 ~-0.1MPa, and removing ethanol, obtains the antarctic krill oil that phospholipids content is greater than 40w ﹪.
Step (1) ~ (5) are particularly suitable for ship operates.Owing to containing abundant low temperature autolytic enzyme in krill body, so krill adopts this technique that raw material can be kept fresh immediately and utilizes its low temperature autolytic enzyme assistance enzymolysis after fishing for, reduce added proteins enzyme dosage; Step (6) ~ (8), owing to relating to inflammable and explosive ethanol, therefore should not aboard ship process.
The invention has the beneficial effects as follows:
1, compared with high-temperature cooking process, adopt secondary enzymolysis processing condition gentle, protect natural unsaturated fatty acids (DHA, EPA etc.) and antioxidant (astaxanthin).
2, because secondary enzymolysis destroys the composite structure of albumen and phosphatide, make phosphatide free out, therefore the extraction yield of antarctic krill oil is higher.
3, due to enzymolysis hulling technology, obtain the dried phospholipid layer that is rich in and do not contain shrimp shell, than the antarctic krill oil of the euphausia superba powder enrichment greater concn after high temperature steaming, Oil repellent is lower.
4, because step (1) ~ (5) are suitable for ship operates, can prepare work in-process, then be transported to by work in-process and carry out step (6) ~ (8) operation on the bank, operation makes both to reduce proteolytic enzyme consumption like this, and transportation cost is also lower simultaneously again.
Embodiment
Below the present invention is further analyzed.
The measurement of following examples phospholipids content all adopts GB/T 5537-2008 method; The fresh water salts contg used is less than 1w ﹪; The Sumizyme MP used produces for Bacillus licheniformis (Bacillus licheniformis), and neutral protease is that subtilis (Bacillus subtilis) produces.
Embodiment 1 ~ 8 step is as shown in Figure 1, specific as follows:
Embodiment 1
(1), get freezing krill 500g, be broken for the particle of 10mm.
(2), add fresh water 500mL, add 4000 U/g Sumizyme MP enzymolysis, be warming up to 65 DEG C, be incubated 15 minutes, obtain enzymolysis solution I.
(3), to choose filter cloth be 50 orders, and centrifugal force 350G is centrifugal, and removing shrimp shell, obtains enzymolysis solution II.
(4), enzymolysis solution II is cooled to 55 DEG C, regulates enzymolysis solution pH to 8.0, be incubated 5 hours with 6mol/L sodium hydroxide solution, then with 6 mol/L hydrochloric acid adjustment enzymolysis solution pH to 6.5.
(5), heat enzymolysis solution to 70 DEG C, be incubated 20 minutes.
(6), centrifugal force 500G, centrifugal 20 minutes.Isolate the peptide liquid layer being rich in phospholipid layer PEF and lower floor on upper strata.Peptide liquid layer is at 60 DEG C, and vacuum tightness is krill peptide under-0.05MPa after concentrating under reduced pressure.
(7), 40 DEG C of cryodrying PEF are 8w ﹪ to moisture content.
(8), with 99.5 ﹪ ethanol 1.5L extract the dried PEF of 0.1kg, extraction temperature is 30 DEG C, and extraction time is 2 hours and constantly stirs.
(9), under-0.07MPa vacuum tightness carry out decompress filter, solid residue is krill albumen, and filtrate is scarlet.
(10), filtrate at 40 DEG C, vacuum tightness is rotary evaporation under-0.1MPa, removing ethanol, obtain the antarctic krill oil that phospholipids content is 40.7w ﹪.
Embodiment 2
(1), get freezing krill 500g, be broken for the particle of 8mm.
(2), add fresh water 450mL, add 3500U/g neutral protease enzymolysis, be warming up to 60 DEG C, be incubated 20 minutes, obtain enzymolysis solution I.
(3), to choose filter cloth be 100 orders, and centrifugal force 400G is centrifugal, and removing shrimp shell, obtains enzymolysis solution II.
(4), enzymolysis solution II is cooled to 50 DEG C, is incubated 4 hours.
(5), heat enzymolysis solution to 75 DEG C, be incubated 15 minutes.
(6), centrifugal force 550G, centrifugal 15 minutes.Isolate the peptide liquid layer being rich in phospholipid layer PEF and lower floor on upper strata.Peptide liquid layer is at 70 DEG C, and vacuum tightness is krill peptide under-0.07MPa after concentrating under reduced pressure.
(7), lyophilize PEF is 6w ﹪ to moisture content.
(8), with 99.5 ﹪ ethanol 1L extract the dried PEF of 0.1kg, extraction temperature is 60 DEG C, and extraction time is 3 hours and constantly stirs.
(9), use 600G centrifugation, solid residue is krill albumen, and filtrate is scarlet.
(10), filtrate at 45 DEG C, vacuum tightness is rotary evaporation under-0.05MPa, removing ethanol, obtain the antarctic krill oil that phospholipids content is 42.1w ﹪.
Embodiment 3
(1), get freezing krill 500g, be broken for the particle of 6mm.
(2), add fresh water 400mL, add 3000 U/g neutral protease enzymolysis, be warming up to 65 DEG C, be incubated 15 minutes, obtain enzymolysis solution I.
(3), to choose filter cloth be 150 orders, and centrifugal force 450G is centrifugal, and removing shrimp shell, obtains enzymolysis solution II.
(4), enzymolysis solution II is cooled to 55 DEG C, is incubated 5 hours.
(5), heat enzymolysis solution to 80 DEG C, be incubated 10 minutes.
(6), centrifugal force 600G, centrifugal 10 minutes.Isolate the peptide liquid layer being rich in phospholipid layer PEF and lower floor on upper strata.Peptide liquid layer is at 75 DEG C, and vacuum tightness is krill peptide under-0.08MPa after concentrating under reduced pressure.
(7), lyophilize PEF is 4w ﹪ to moisture content.
(8), with 99.5 ﹪ ethanol 0.8L extract the dried PEF of 0.1kg, extraction temperature is 50 DEG C, and extraction time is 3 hours and constantly stirs.
(9), under-0.08MPa vacuum tightness carry out decompress filter, solid residue is krill albumen, and filtrate is scarlet.
(10), filtrate at 50 DEG C, vacuum tightness is rotary evaporation under-0.1MPa, removing ethanol, obtain the antarctic krill oil that phospholipids content is 45.8w ﹪.
Embodiment 4
(1), get freezing krill 500g, be broken for the particle of 4mm.
(2), add fresh water 350mL, add 2500U/g Sumizyme MP enzymolysis, be warming up to 60 DEG C, be incubated 15 minutes.
(3), to choose filter cloth be 200 orders, and centrifugal force 500G is centrifugal, and removing shrimp shell, obtains enzymolysis solution II.
(4), enzymolysis solution II is cooled to 50 DEG C, regulates enzymolysis solution pH to 8.0, be incubated 6 hours with 6mol/L sodium hydroxide solution, then with 6 mol/L hydrochloric acid adjustment enzymolysis solution pH to 6.5.
(5), heat enzymolysis solution to 85 DEG C, be incubated 5 minutes.
(6), centrifugal force 650G, centrifugal 5 minutes.Isolate the peptide liquid layer being rich in phospholipid layer PEF and lower floor on upper strata.Peptide liquid layer is at 65 DEG C, and vacuum tightness is krill peptide under-0.1MPa after concentrating under reduced pressure.
(7), 50 DEG C of cryodrying PEF are 2w ﹪ to moisture content.
(8), with 99.5 ﹪ ethanol 1.2L extract the dried PEF of 0.1kg, extraction temperature is 40 DEG C, and extraction time is 2 hours and constantly stirs.
(9), use 700G centrifugation, solid residue is krill albumen, and filtrate is scarlet.
(10), filtrate at 55 DEG C, vacuum tightness is rotary evaporation under-0.08MPa, removing ethanol, obtain the antarctic krill oil that phospholipids content is 43.3w ﹪.
Embodiment 5
(1), get freezing krill 500g, be broken for the particle of 10mm.
(2), add fresh water 250mL, add 4500 U/g neutral protease enzymolysis, be warming up to 50 DEG C, be incubated 60 minutes, obtain enzymolysis solution I.
(3), to choose filter cloth be 150 orders, and centrifugal force 450G is centrifugal, and removing shrimp shell, obtains enzymolysis solution II.
(4), enzymolysis solution II is cooled to 55 DEG C, is incubated 6 hours.
(5), heat enzymolysis solution to 90 DEG C, be incubated 30 minutes.
(6), centrifugal force 700G, centrifugal 10 minutes.Isolate the peptide liquid layer being rich in phospholipid layer PEF and lower floor on upper strata.Peptide liquid layer is at 80 DEG C, and vacuum tightness is krill peptide under-0.08MPa after concentrating under reduced pressure.
(7), lyophilize PEF is 4w ﹪ to moisture content.
(8), with 99.5 ﹪ ethanol 0.5L extract the dried PEF of 0.1kg, extraction temperature is 60 DEG C, and extraction time is 1 hour and constantly stirs.
(9), under-0.1MPa vacuum tightness carry out decompress filter, solid residue is krill albumen, and filtrate is scarlet.
(10), filtrate at 60 DEG C, vacuum tightness is rotary evaporation under-0.06Pa, removing ethanol, obtain the antarctic krill oil that phospholipids content is 45.2w ﹪.
Embodiment 6
(1), get freezing krill 500g, be broken for the particle of 4mm.
(2), add fresh water 750mL, add 1500U/g Sumizyme MP enzymolysis, be warming up to 60 DEG C, be incubated 15 minutes.
(3), to choose filter cloth be 200 orders, and centrifugal force 500G is centrifugal, and removing shrimp shell, obtains enzymolysis solution II.
(4), enzymolysis solution II is cooled to 50 DEG C, regulates enzymolysis solution pH to 8.5, be incubated 3 hours with 6mol/L sodium hydroxide solution, then with 6 mol/L hydrochloric acid adjustment enzymolysis solution pH to 7.0.
(5), heat enzymolysis solution to 70 DEG C, be incubated 30 minutes.
(6), centrifugal force 650G, centrifugal 5 minutes.Isolate the peptide liquid layer being rich in phospholipid layer PEF and lower floor on upper strata.Peptide liquid layer is at 65 DEG C, and vacuum tightness is krill peptide under-0.1MPa after concentrating under reduced pressure.
(7), 50 DEG C of cryodrying PEF are 2w ﹪ to moisture content.
(8), with 99.5 ﹪ ethanol 0.5L extract the dried PEF of 0.1kg, extraction temperature is 40 DEG C, and extraction time is 2 hours and constantly stirs.
(9), use 500G centrifugation, solid residue is krill albumen, and filtrate is scarlet.
(10), filtrate at 30 DEG C, vacuum tightness is rotary evaporation under-0.1MPa, removing ethanol, obtain the antarctic krill oil that phospholipids content is 42.6w ﹪.
Embodiment 7
(1), get freezing krill 500g, be broken for the particle of 10mm.
(2), add fresh water 250mL, add 4500 U/g neutral protease enzymolysis, be warming up to 50 DEG C, be incubated 60 minutes, obtain enzymolysis solution I.
(3), to choose filter cloth be 150 orders, and centrifugal force 450G is centrifugal, and removing shrimp shell, obtains enzymolysis solution II.
(4), enzymolysis solution II is cooled to 55 DEG C, is incubated 3 hours.
(5), heat enzymolysis solution to 90 DEG C, be incubated 30 minutes.
(6), centrifugal force 700G, centrifugal 10 minutes.Isolate the peptide liquid layer being rich in phospholipid layer PEF and lower floor on upper strata.Peptide liquid layer is at 80 DEG C, and vacuum tightness is krill peptide under-0.08MPa after concentrating under reduced pressure.
(7), lyophilize PEF is 4w ﹪ to moisture content.
(8), with 99.5 ﹪ ethanol 0.8L extract the dried PEF of 0.1kg, extraction temperature is 60 DEG C, and extraction time is 1 hour and constantly stirs.
(9), under-0.05MPa vacuum tightness carry out decompress filter, solid residue is krill albumen, and filtrate is scarlet.
(10), filtrate at 60 DEG C, vacuum tightness is rotary evaporation under-0.06Pa, removing ethanol, obtain the antarctic krill oil that phospholipids content is 45.1w ﹪.
Embodiment 8
(1), get freezing krill 500g, be broken for the particle of 4mm.
(2), add fresh water 750mL, add 1500U/g Sumizyme MP enzymolysis, be warming up to 60 DEG C, be incubated 15 minutes.
(3), to choose filter cloth be 200 orders, and centrifugal force 500G is centrifugal, and removing shrimp shell, obtains enzymolysis solution II.
(4), enzymolysis solution II is cooled to 50 DEG C, regulates enzymolysis solution pH to 7.5, be incubated 6 hours with 6mol/L sodium hydroxide solution, then with 6 mol/L hydrochloric acid adjustment enzymolysis solution pH to 6.0.
(5), heat enzymolysis solution to 70 DEG C, be incubated 30 minutes.
(6), centrifugal force 650G, centrifugal 5 minutes.Isolate the peptide liquid layer being rich in phospholipid layer PEF and lower floor on upper strata.Peptide liquid layer is at 65 DEG C, and vacuum tightness is krill peptide under-0.1MPa after concentrating under reduced pressure.
(7), 50 DEG C of cryodrying PEF are 2w ﹪ to moisture content.
(8), with 99.6 ﹪ ethanol 0.5L extract the dried PEF of 0.1kg, extraction temperature is 40 DEG C, and extraction time is 2 hours and constantly stirs.
(9), use 650G centrifugation, solid residue is krill albumen, and filtrate is scarlet.
(10), filtrate at 55 DEG C, vacuum tightness is rotary evaporation under-0.09MPa, removing ethanol, obtain the antarctic krill oil that phospholipids content is 43.3w ﹪.
Comparative example
(1), get freezing krill 500g, be crushed to 15 orders with tissue pulverizer;
(2), by the krill after pulverizing be placed in vacuum drying oven, adjust the temperature to 45 DEG C, be incubated 45 minutes, make krill self-dissolving;
(3), the temperature to 90 DEG C of Raise vacuum loft drier, be incubated 15 minutes, make enzyme deactivation;
(4), the temperature of vacuum drying oven is dropped to 40 DEG C after start insulation, open vacuum compressor simultaneously, dry moisture to 5 below ﹪, obtain dry krill meal 92g;
(5), by dried krill meal be placed in 1500ml flask, then add 920g and measure the ethanol that percentage concentration is greater than 95 ﹪ and carry out water-bath, bath temperature is 45 DEG C, and the reaction times is 4 hours;
(6), be cooled to room temperature, filter, get solvent content and obtain the antarctic krill oil that phospholipids content is 33w ﹪ after Rotary Evaporators evaporation.
Embodiment 1 ~ 8 is compared with comparative example, and the extraction yield of antarctic krill oil is higher, and phospholipids content is also higher, and obtains krill peptide and the krill albumen of high added value simultaneously, provides a kind of new solution for making full use of krill resource.
The above; be only the present invention's preferably embodiment; but protection scope of the present invention is not limited thereto; anyly be familiar with those skilled in the art in the technical scope that the present invention discloses; be equal to according to technical scheme of the present invention and inventive concept thereof and replace or change, all should be encompassed within protection scope of the present invention.
Claims (8)
1. an aqueous enzymatic method prepares the method being rich in the antarctic krill oil of phosphatide, it is characterized in that the method utilizes autolytic enzyme in conjunction with added proteins enzyme enzymolysis, first by protease hydrolyzed liquefaction krill meat, centrifugally slough shrimp shell, enzymolysis destroys the composite structure of albumen and phosphatide again, make phosphatide free out, then centrifugation go out to be rich in phospholipid layer; Be rich in phospholipid layer through super-dry, alcohol extraction obtains phospholipids content and is greater than the antarctic krill oil of 40w ﹪, krill peptide and krill albumen.
2. the method for the antarctic krill oil of phosphatide is rich in a kind of aqueous enzymatic method preparation as claimed in claim 1, it is characterized in that the method specifically comprises the following steps:
Step (1), fresh krill or freezing krill are crushed to particle diameter are less than or equal to 10mm;
Step (2), the water krill after step (1) fragmentation and salts contg being less than 1w ﹪ are that 1:0.5 ~ 1.5 mix according to weight ratio, then 1500U/g ~ 4500 U/g protease hydrolyzed is added, be warming up to 50 ~ 65 DEG C, insulation reaction 15 ~ 60 minutes, obtains enzymolysis solution I;
The enzymolysis solution I that step (3), step (2) obtain is after 50 ~ 200 order filter-cloth filterings, centrifugal under centrifugal force 350 ~ 500G, and removing shrimp shell, is cooled to 50 ~ 55 DEG C, then carries out secondary enzymolysis, obtain enzymolysis solution II;
The enzymolysis solution II to 70 ~ 90 DEG C that step (4), heating steps (3) obtain, is incubated 5 ~ 30 minutes, produces precipitation; Under being deposited in centrifugal force 500 ~ 700G centrifugal 5 ~ 20 minutes, isolate the peptide liquid layer being rich in phospholipid layer PEF and lower floor on upper strata; By peptide liquid layer at 60 ~ 80 DEG C, vacuum tightness is krill peptide under-0.05 ~-0.1MPa after concentrating under reduced pressure;
Step (5), phospholipid layer PEF cryodrying will be rich in or lyophilize to moisture content is less than 10w ﹪;
Step (6), obtain driedly being rich in phospholipid layer PEF by alcohol extraction step (5), extraction temperature is 30 ~ 60 DEG C, and extraction time is 1 ~ 3 hour, constantly stirs in extraction process, is extracted liquid;
The solid-liquid ratio being rich in phospholipid layer PEF and ethanol after described drying is 1:5 ~ 15, and unit is kg:L;
Step (7), under centrifugal force 500 ~ 700G, centrifugation or vacuum tightness carry out suction filtration to the extraction liquid that step (6) obtains under-0.05 ~-0.1MPa, obtain solid residue and filtrate; Described solid residue is krill albumen, and filtrate is scarlet;
The filtrate that step (8), step (7) obtain is at 30 ~ 60 DEG C, and vacuum tightness is rotary evaporation under-0.05 ~-0.1MPa, and removing ethanol, obtains the antarctic krill oil that phospholipids content is greater than 40w ﹪.
3. the method for the antarctic krill oil of phosphatide is rich in a kind of aqueous enzymatic method preparation as claimed in claim 2, it is characterized in that described proteolytic enzyme is neutral protease or Sumizyme MP.
4. the method for the antarctic krill oil of phosphatide is rich in a kind of aqueous enzymatic method preparation as claimed in claim 2, what if it is characterized in that, step (2) adopted is proteolytic enzyme is Sumizyme MP, step (3) secondary enzymolysis process must by enzymolysis solution I adjust ph after process, specifically regulate enzymolysis solution I pH value to 7.5 ~ 8.5 with sodium hydroxide, be incubated after 3 ~ 6 hours, use salt acid for adjusting pH value to 6.0 ~ 7.0 again, obtain enzymolysis solution II.
5. the method for the antarctic krill oil of phosphatide is rich in a kind of aqueous enzymatic method preparation as claimed in claim 2, what if it is characterized in that, step (2) adopted is proteolytic enzyme is neutral protease, enzymolysis solution I in step (3) secondary enzymolysis process after process need not adjust ph, direct insulation 3 ~ 6 hours, obtains enzymolysis solution II.
6. the method for the antarctic krill oil of phosphatide is rich in a kind of aqueous enzymatic method preparation as claimed in claim 2, it is characterized in that the requirement of described low temperature is lower than 50 DEG C.
7. the method for the antarctic krill oil of phosphatide is rich in a kind of aqueous enzymatic method preparation as claimed in claim 2, it is characterized in that described alcohol concn is more than or equal to 95.5 ﹪ (v/v).
8. the method for the antarctic krill oil of phosphatide is rich in a kind of aqueous enzymatic method preparation as claimed in claim 2, it is characterized in that step (1) ~ (5) are suitable for ship operates, owing to containing abundant low temperature autolytic enzyme in krill body, so krill adopts this technique that raw material can be kept fresh immediately and utilizes its low temperature autolytic enzyme assistance enzymolysis after fishing for, reduce added proteins enzyme dosage; Step (6) ~ (8), owing to relating to inflammable and explosive ethanol, therefore should not aboard ship process.
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| CN106588977A (en) * | 2016-12-13 | 2017-04-26 | 山东省科学院生物研究所 | Method for extracting high-purity marine phospholipids from Euphausia superba |
| CN106588977B (en) * | 2016-12-13 | 2018-06-29 | 山东省科学院生物研究所 | A kind of method that high-purity marine phospholipids are extracted from krill |
| CN106588976B (en) * | 2016-12-13 | 2018-06-29 | 山东省科学院生物研究所 | The method that high-purity omega-3 polyunsaturated fatty acids phosphatide is extracted from shrimp head |
| CN111432654A (en) * | 2017-12-04 | 2020-07-17 | 雾凇科技有限公司 | Method for producing protein phospholipid complex from crustacean capture |
| CN108179053A (en) * | 2018-01-26 | 2018-06-19 | 日照职业技术学院 | A kind of preparation method of high quality Antarctic krill oil |
| CN108179053B (en) * | 2018-01-26 | 2021-07-20 | 日照职业技术学院 | Preparation method of high-quality euphausia superba oil |
| CN109349416A (en) * | 2018-12-01 | 2019-02-19 | 浙江海洋大学 | A method of albumen powder is made using South Pole squama shrimp |
| CN109880871A (en) * | 2019-04-18 | 2019-06-14 | 威海市宇王集团海洋生物工程有限公司 | A method of small-molecular peptides are extracted by raw material of euphausia superba powder |
| WO2021194360A1 (en) * | 2020-03-27 | 2021-09-30 | Pharmazen Limited | Extraction method for bio-active fractions |
| CN114717043A (en) * | 2020-12-22 | 2022-07-08 | 上海崇瀚生物科技有限公司 | Method for extracting phospholipid-rich Omega-3 fatty acid oil and/or selenium-rich small molecule peptide substances from Antarctic krill |
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