CN118165100A - Extraction method and application of fish scale collagen - Google Patents
Extraction method and application of fish scale collagen Download PDFInfo
- Publication number
- CN118165100A CN118165100A CN202410473115.6A CN202410473115A CN118165100A CN 118165100 A CN118165100 A CN 118165100A CN 202410473115 A CN202410473115 A CN 202410473115A CN 118165100 A CN118165100 A CN 118165100A
- Authority
- CN
- China
- Prior art keywords
- fish scale
- collagen
- fish
- powder
- scales
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 241000251468 Actinopterygii Species 0.000 title claims abstract description 149
- 102000008186 Collagen Human genes 0.000 title claims abstract description 76
- 108010035532 Collagen Proteins 0.000 title claims abstract description 76
- 229920001436 collagen Polymers 0.000 title claims abstract description 75
- 238000000605 extraction Methods 0.000 title claims abstract description 17
- 239000000843 powder Substances 0.000 claims abstract description 47
- 238000000855 fermentation Methods 0.000 claims abstract description 28
- 230000004151 fermentation Effects 0.000 claims abstract description 28
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 28
- 238000001914 filtration Methods 0.000 claims abstract description 26
- 238000000034 method Methods 0.000 claims abstract description 23
- 244000063299 Bacillus subtilis Species 0.000 claims abstract description 16
- 235000014469 Bacillus subtilis Nutrition 0.000 claims abstract description 16
- 241000186660 Lactobacillus Species 0.000 claims abstract description 16
- 229940039696 lactobacillus Drugs 0.000 claims abstract description 16
- 239000004365 Protease Substances 0.000 claims abstract description 15
- 108090000526 Papain Proteins 0.000 claims abstract description 14
- 102000057297 Pepsin A Human genes 0.000 claims abstract description 14
- 108090000284 Pepsin A Proteins 0.000 claims abstract description 14
- 102000004142 Trypsin Human genes 0.000 claims abstract description 14
- 108090000631 Trypsin Proteins 0.000 claims abstract description 14
- 229940055729 papain Drugs 0.000 claims abstract description 14
- 235000019834 papain Nutrition 0.000 claims abstract description 14
- 229940111202 pepsin Drugs 0.000 claims abstract description 14
- 239000006228 supernatant Substances 0.000 claims abstract description 14
- 229960001322 trypsin Drugs 0.000 claims abstract description 14
- 239000012588 trypsin Substances 0.000 claims abstract description 14
- 238000010025 steaming Methods 0.000 claims abstract description 11
- YWYZEGXAUVWDED-UHFFFAOYSA-N triammonium citrate Chemical compound [NH4+].[NH4+].[NH4+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O YWYZEGXAUVWDED-UHFFFAOYSA-N 0.000 claims abstract description 11
- 102000004190 Enzymes Human genes 0.000 claims abstract description 10
- 108090000790 Enzymes Proteins 0.000 claims abstract description 10
- 229940088598 enzyme Drugs 0.000 claims abstract description 10
- 241000894006 Bacteria Species 0.000 claims abstract description 9
- 238000000926 separation method Methods 0.000 claims abstract description 9
- 238000000265 homogenisation Methods 0.000 claims abstract description 8
- 238000009835 boiling Methods 0.000 claims abstract description 7
- 238000001816 cooling Methods 0.000 claims abstract description 7
- 239000000706 filtrate Substances 0.000 claims abstract description 7
- 238000004108 freeze drying Methods 0.000 claims abstract description 7
- 230000000415 inactivating effect Effects 0.000 claims abstract description 7
- 230000007935 neutral effect Effects 0.000 claims abstract description 7
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical group [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 18
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims description 18
- 239000000243 solution Substances 0.000 claims description 14
- 239000007788 liquid Substances 0.000 claims description 13
- 238000005406 washing Methods 0.000 claims description 13
- 238000002791 soaking Methods 0.000 claims description 12
- 239000007787 solid Substances 0.000 claims description 12
- 239000002245 particle Substances 0.000 claims description 10
- 238000001035 drying Methods 0.000 claims description 8
- 239000002253 acid Substances 0.000 claims description 5
- 235000013305 food Nutrition 0.000 claims description 5
- 238000012545 processing Methods 0.000 claims description 5
- 239000003513 alkali Substances 0.000 claims description 3
- 238000005238 degreasing Methods 0.000 claims description 3
- 230000002328 demineralizing effect Effects 0.000 claims description 3
- 230000036541 health Effects 0.000 claims description 3
- 239000000047 product Substances 0.000 claims description 3
- 239000012670 alkaline solution Substances 0.000 claims description 2
- 230000000694 effects Effects 0.000 abstract description 6
- 230000000052 comparative effect Effects 0.000 description 9
- 238000009826 distribution Methods 0.000 description 5
- 239000012535 impurity Substances 0.000 description 4
- 108090000765 processed proteins & peptides Proteins 0.000 description 3
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 2
- 239000008280 blood Substances 0.000 description 2
- 210000004369 blood Anatomy 0.000 description 2
- 238000004364 calculation method Methods 0.000 description 2
- 230000029087 digestion Effects 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 229910052500 inorganic mineral Inorganic materials 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 239000011707 mineral Substances 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 102000011782 Keratins Human genes 0.000 description 1
- 108010076876 Keratins Proteins 0.000 description 1
- 102000035195 Peptidases Human genes 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 101710172711 Structural protein Proteins 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- 239000002585 base Substances 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 229910000389 calcium phosphate Inorganic materials 0.000 description 1
- 235000011010 calcium phosphates Nutrition 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000010411 cooking Methods 0.000 description 1
- 238000004132 cross linking Methods 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- 238000005115 demineralization Methods 0.000 description 1
- 210000004207 dermis Anatomy 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 230000007071 enzymatic hydrolysis Effects 0.000 description 1
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 description 1
- 235000020776 essential amino acid Nutrition 0.000 description 1
- 239000003797 essential amino acid Substances 0.000 description 1
- 238000010297 mechanical methods and process Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 230000037323 metabolic rate Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000000050 nutritive effect Effects 0.000 description 1
- 238000004806 packaging method and process Methods 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 235000019419 proteases Nutrition 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 210000003491 skin Anatomy 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 description 1
Abstract
The invention discloses an extraction method and application of fish scale collagen, and belongs to the technical field of collagen extraction. Adding 20-30 times of water into the washed, demineralized, defatted and washed neutral fish scale powder for homogenization, steaming at 90-110 ℃ for 20-30 min, cooling to room temperature to obtain fish scale homogenate, adding 0.1-0.3% of ammonium citrate into the fish scale homogenate, adding bacillus subtilis and lactobacillus mixed bacteria, and fermenting at 35-38 ℃ for 24-48 h to obtain fermentation liquor; adding papain, pepsin and trypsin into the fermentation liquor, carrying out enzymolysis for 1-2 hours at 50-60 ℃, inactivating enzyme in a boiling water bath to obtain an enzymolysis liquor, carrying out preliminary filtration by using a filter screen, carrying out high-speed centrifugal separation on the filtrate to obtain a supernatant, and freeze-drying the supernatant to obtain the fish scale collagen. The molecular weight of the fish scale collagen extracted by the method is more than 80% and is concentrated at 500-1000 Da, so that the effect of the collagen can be exerted to the greatest extent.
Description
Technical Field
The invention belongs to the technical field of collagen extraction, and particularly relates to an extraction method and application of fish scale collagen.
Background
The fish resources in China are rich, a large amount of offal is generated in the processing and consuming processes of related products, the offal accounts for about 50% of the mass of fresh fish, the offal comprises fish bones, fish fins, fish scales and the like, and the fish scales account for 6-10%. The fish scale byproduct is processed, so that the utilization value of the fish scale can be improved, the pollution of the fish scale to the environment is reduced, and the development of the fish processing industry can be promoted. The fish scales mainly comprise organic matters and mineral matters, wherein collagen and keratin in the organic matters account for more than 90% of the total content of the organic matters, and the mineral matters mainly comprise calcium phosphate.
Collagen is an important structural protein, and its microstructure contains 1 or several triple-helical regions, and these helical structures are composed of alpha peptide chains. The fish scale collagen has unique molecular structure, good intermolecular crosslinking capability, good film forming property, biodegradability, various essential amino acids of human body, health care function and high nutritive value, and can be added into food and food packaging.
At present, the method for extracting the collagen from the fish scales is mainly divided into an acid method, an alkali method, an enzyme method, a hot water method and a compound method, and adopts a physical-mechanical method or a specific chemical reagent to change the existence environment of the collagen in the fish scales, so that the collagen and other components are finally separated, and the collagen is obtained. However, the extraction method is different, and the extraction rate and the main form of the extract are different. The traditional method is an acid-base extraction method, wherein fish scales are cleaned, degreased and decolored, then soaked in hydrochloric acid (pH 4-5) for decalcification treatment (2-3 weeks), and the fish scales are taken out, cleaned and limed for 15-20 d, so that the raw material tissues are loosened. The method has the advantages that the steps are numerous, the time is long, and the collagen loss is serious in the extraction process.
At present, the molecular weight distribution of the fish scale collagen obtained from the fish scales is relatively wide, and the problems of overlarge molecular weight and overlarge molecular weight exist. Patent CN107723331 a discloses a process for extracting scale collagen from scales, which sequentially carries out demineralization, degreasing, cooking, expansion treatment and fermentation treatment on scales, then carries out protein decomposition by culturing protease-producing strains, obtains low molecular weight collagen polypeptide and improves the extraction rate, but the obtained scale collagen has a high content of molecular weight less than 500Da (daltons), and when the scale collagen is applied to processing of foods, medicines and the like, the scale collagen is particularly easy to penetrate cell membranes in organisms, has high metabolic rate, is extremely easy to digest and discharge outside bodies, and cannot exert the efficacy. Collagen with an excessive molecular weight is difficult to be absorbed by living bodies and cannot exert its efficacy. Therefore, it is extremely important to control the molecular weight of the collagen peptide obtained from fish scales.
Disclosure of Invention
Aiming at the problems that the molecular weight distribution range of the fish scale collagen obtained from the fish scales is large and the collagen effect of too high or too low molecular weight cannot be exerted in the prior art, the invention provides an extraction method and application of the fish scale collagen, wherein the molecular weight of the extracted fish scale collagen is concentrated to 500-1000 Da to the greatest extent.
The invention is realized by the following technical scheme:
The extraction method of the fish scale collagen comprises the following steps:
(1) Adding water with the volume of 20-30 times into the washed, demineralized, defatted and washed neutral fish scale powder for homogenating, steaming at 90-110 ℃ for 20-30 min, and cooling to room temperature to obtain fish scale homogenate;
(2) Fermentation: adding 0.1-0.3% of ammonium citrate into the fish scale homogenate, adding mixed bacteria of bacillus subtilis and lactobacillus, and fermenting at 35-38 ℃ for 24-48 hours to obtain fermentation liquor;
(3) Enzymolysis: adding papain, pepsin and trypsin into the fermentation broth in the step (2), carrying out enzymolysis for 1-2 hours at 50-60 ℃, and inactivating enzyme in a boiling water bath to obtain an enzymolysis liquid;
(4) And (3) filtering: and (3) performing preliminary filtration on the enzymolysis liquid in the step (3) by using a filter screen, performing high-speed centrifugal separation on the filtrate to obtain supernatant, and performing freeze drying on the supernatant to obtain the fish scale collagen.
Preferably, the dosage of the bacillus subtilis in the step (2) is 2-5% of the mass of the fish scale powder; the dosage of the lactobacillus is 0.8-1.5% of the mass of the fish scale powder.
Preferably, 1X 10 4~2×104 U papain, 2X 10 4~5×104 U pepsin and 1X 10 3~5×103 U trypsin are added to each kilogram of fish scale powder in step (3).
Preferably, the high-speed centrifugal separation speed in the step (4) is 10000-18000 rpm.
Preferably, the homogenization conditions in the step (1) are 2000-2500 rpm for 8-10 min.
Preferably, the method for cleaning, demineralizing and degreasing the fish scales in the step (1) comprises the following steps: drying the cleaned fish scales, crushing the fish scales into fish scale powder with the particle size of 100-200 meshes, adding the fish scale powder into acid liquor, carrying out ultrasonic soaking for 30-40 min, filtering, washing the solid with water to pH 6-7, adding the washed fish scale powder into alkaline solution, carrying out ultrasonic soaking for 30-40 min, filtering, and washing the solid with water to pH 6-8.
Preferably, the acid solution is a citric acid solution with the mass concentration of 1-4%; the alkali liquor is sodium hydroxide solution with the mass concentration of 0.05-0.1%.
The invention also discloses the fish scale collagen obtained by the extraction method; further, the mass ratio of the molecular weight of the fish scale collagen in the fish scale collagen is more than 80% between 500 and 1000 Da.
The invention relates to application of fish scale collagen in food, medicine and health care product processing.
Advantageous effects
According to the invention, ammonium citrate is added into the fish scale homogenate, and the molecular weight of the obtained fish scale collagen is concentrated to 500-1000 Da by fermentation and enzyme treatment, so that the collagen with the molecular weight can be directly absorbed by blood without digestion, and is transferred to skin, and can reach dermis quickly within 2-4 hours, and the collagen can not be digested and discharged out of the body easily due to too high metabolism speed, thus the effect of the collagen can not be exerted, and the collagen is suitable for organism absorption and utilization, and the effect of the collagen can be maximally exerted;
The invention adopts the mixed bacteria of bacillus subtilis and lactobacillus for fermentation, and controls the molecular weight range of collagen by matching with the action of ammonium citrate; papain, pepsin and trypsin are added into the fermentation liquid, and the types, the addition amount and the enzymolysis time of the enzymes are controlled, so that the molecular weight of the obtained fish scale collagen is concentrated to 500-1000 Da, and the effect of the collagen is exerted to the greatest extent.
Detailed Description
The present application is further illustrated below with reference to specific examples, which are to be construed as merely illustrative of the application and not limiting of its scope, as various equivalent modifications to the application will fall within the scope of the application as defined in the appended claims after reading the application.
The protein content in the scales used in the following examples and comparative examples was 66.3%, and the collagen content was 37.2%.
Example 1
(1) Pretreatment: drying the cleaned fish scales, crushing the fish scales into fish scale powder with the particle size of 100-200 meshes, adding the fish scale powder into a citric acid solution with the mass concentration of 3%, soaking the fish scales in ultrasonic (ultrasonic frequency of 40kHz, ultrasonic sound intensity of 0.6W/cm 2) for 35min, filtering, washing the solid with water to pH6.2, adding the washed fish scale powder into a sodium hydroxide solution with the mass concentration of 0.08%, soaking the fish scale powder in ultrasonic (ultrasonic frequency of 35kHz, ultrasonic sound intensity of 0.3W/cm 2) for 35min, filtering, and washing the solid with water to pH7.6;
(2) Homogenizing and steaming: adding 25 times of water into the washed, demineralized, defatted and washed neutral fish scale powder for homogenization (10 min under 2000 rpm), steaming at 100deg.C for 25min, and cooling to room temperature to obtain fish scale homogenate;
(3) Fermentation: adding 0.25% ammonium citrate into the fish scale homogenate, adding mixed bacteria of bacillus subtilis and lactobacillus (the dosage of the bacillus subtilis is 4% of the mass of fish scale powder, and the dosage of the lactobacillus is 1.2% of the mass of fish scale), and fermenting at 37 ℃ for 36 hours to obtain fermentation liquor;
(1) Enzymolysis: adding papain, pepsin and trypsin (2X 10 4 U papain, 2X 10 4 U pepsin and 1X 10 3 U trypsin are added into each kilogram of fish scale powder) into the fermentation broth in the step (2), carrying out enzymolysis for 1.5 hours at 60 ℃, and inactivating enzyme in a boiling water bath to obtain an enzymolysis liquid;
(2) And (3) filtering: and (3) performing preliminary filtration on the enzymolysis liquid in the step (3) by using a filter screen to remove large-particle impurities, performing high-speed centrifugal separation (15000 rpm) on the filtrate to obtain supernatant, and performing freeze drying on the supernatant to obtain the fish scale collagen.
Example 2
(1) Pretreatment: drying the cleaned fish scales, crushing the fish scales into fish scale powder with the particle size of 100-200 meshes, adding the fish scale powder into a citric acid solution with the mass concentration of 4%, soaking the fish scales in ultrasonic (ultrasonic frequency of 40kHz, ultrasonic sound intensity of 0.6W/cm 2) for 30min, filtering, washing the solid with water to pH6.3, adding the washed fish scale powder into a 0.06% sodium hydroxide solution, soaking the fish scale powder in ultrasonic (ultrasonic frequency of 35kHz, ultrasonic sound intensity of 0.3W/cm 2) for 30min, filtering, and washing the solid with water to pH7.5;
(2) Homogenizing and steaming: adding 20 times of water into the washed, demineralized, defatted and washed neutral fish scale powder for homogenization (8 min under 2500 rpm), steaming at 100deg.C for 25min, and cooling to room temperature to obtain fish scale homogenate;
(3) Fermentation: adding 0.2% ammonium citrate into the fish scale homogenate, adding mixed bacteria of bacillus subtilis and lactobacillus (the dosage of the bacillus subtilis is 5% of the mass of the fish scale powder, and the dosage of the lactobacillus is 0.8% of the mass of the fish scale powder), and fermenting at 36 ℃ for 42h to obtain fermentation liquor;
(4) Enzymolysis: adding papain, pepsin and trypsin (10 4 U papain, 4X 10 4 U pepsin and 2X 10 3 U trypsin are added into each kilogram of fish scale powder) into the fermentation broth in the step (2), carrying out enzymolysis for 1.5 hours at 60 ℃, and inactivating enzyme in a boiling water bath to obtain an enzymolysis liquid;
(5) And (3) filtering: and (3) performing preliminary filtration on the enzymolysis liquid in the step (3) by using a filter screen to remove large-particle impurities, performing high-speed centrifugal separation (15000 rpm) on the filtrate to obtain supernatant, and performing freeze drying on the supernatant to obtain the fish scale collagen.
Example 3
(1) Pretreatment: drying the cleaned fish scales, crushing the fish scales into fish scale powder with the particle size of 100-200 meshes, adding the fish scale powder into a citric acid solution with the mass concentration of 2.5%, soaking the fish scales in ultrasonic (ultrasonic frequency of 40kHz, ultrasonic sound intensity of 0.6W/cm 2) for 40min, filtering, washing the solid with water to pH6.4, adding the washed fish scale powder into a sodium hydroxide solution with the mass concentration of 0.06%, soaking the fish scale powder in ultrasonic (ultrasonic frequency of 35kHz, ultrasonic sound intensity of 0.3W/cm 2) for 40min, filtering, and washing the solid with water to pH7.6;
(2) Homogenizing and steaming: adding 30 times of water into the washed, demineralized, defatted and washed neutral fish scale powder for homogenization (8 min under 2500 rpm), steaming at 100deg.C for 25min, and cooling to room temperature to obtain fish scale homogenate;
(3) Fermentation: adding 0.3% ammonium citrate into the fish scale homogenate, adding mixed bacteria of bacillus subtilis and lactobacillus (the dosage of the bacillus subtilis is 3% of the mass of the fish scale powder, and the dosage of the lactobacillus is 1.5% of the mass of the fish scale powder), and fermenting at 37 ℃ for 30 hours to obtain fermentation liquor;
(4) Enzymolysis: adding papain, pepsin and trypsin (2X 10 4 U papain, 2X 10 4 pepsin and 2X 10 3 U trypsin are added into each kilogram of fish scale powder) into the fermentation broth in the step (2), and carrying out enzymolysis for 2 hours at 55 ℃, and inactivating enzyme in a boiling water bath to obtain an enzymolysis liquid;
(5) And (3) filtering: and (3) performing preliminary filtration on the enzymolysis liquid in the step (3) by using a filter screen to remove large-particle impurities, performing high-speed centrifugal separation (15000 rpm) on the filtrate to obtain supernatant, and performing freeze drying on the supernatant to obtain the fish scale collagen.
Example 4
(1) Pretreatment: drying the cleaned fish scales, crushing the fish scales into fish scale powder with the particle size of 100-200 meshes, adding the fish scale powder into citric acid solution with the mass concentration of 3%, soaking the fish scales in ultrasonic (ultrasonic frequency of 40kHz, ultrasonic sound intensity of 0.6W/cm 2) for 35min, filtering, washing the solid with water to pH6.2, adding the washed fish scale powder into sodium hydroxide solution with the mass concentration of 0.08%, soaking the fish scale powder in ultrasonic (ultrasonic frequency of 35kHz, ultrasonic sound intensity of 0.3W/cm 2) for 35min, filtering, and washing the solid with water to pH7.4;
(2) Homogenizing and steaming: adding 25 times of water into the washed, demineralized, defatted and washed neutral fish scale powder for homogenization (10 min under 2000 rpm), steaming at 100deg.C for 25min, and cooling to room temperature to obtain fish scale homogenate;
(3) Fermentation: adding 0.25% ammonium citrate into the fish scale homogenate, adding mixed bacteria of bacillus subtilis and lactobacillus (the dosage of the bacillus subtilis is 3% of the mass of the fish scale powder, and the dosage of the lactobacillus is 1.2% of the mass of the fish scale powder), and fermenting at 37 ℃ for 36 hours to obtain fermentation liquor;
(4) Enzymolysis: adding papain, pepsin and trypsin (2X 10 4 U papain, 4X 10 4 U pepsin and 1X 10 3 U trypsin are added into each kilogram of fish scale powder) into the fermentation broth in the step (2), carrying out enzymolysis for 1.5 hours at 60 ℃, and inactivating enzyme in a boiling water bath to obtain an enzymolysis liquid;
(5) And (3) filtering: and (3) performing preliminary filtration on the enzymolysis liquid in the step (3) by using a filter screen to remove large-particle impurities, performing high-speed centrifugal separation (15000 rpm) on the filtrate to obtain supernatant, and performing freeze drying on the supernatant to obtain the fish scale collagen.
Comparative example 1
Comparative example 1 the procedure was the same as in example 1 except for the fermentation in step (3), and fish scale collagen was obtained by pretreatment, homogenization, fermentation, enzymatic hydrolysis, and filtration;
fermentation (no ammonium citrate added): adding mixed bacteria of bacillus subtilis and lactobacillus into the fish scale homogenate (the dosage of the bacillus subtilis is 4% of the mass of the fish scale powder, and the dosage of the lactobacillus is 1.2% of the mass of the fish scale), and fermenting at 37 ℃ for 36 hours to obtain fermentation liquor.
Comparative example 2
The procedure of comparative example 1 was the same as in example 1, except that the procedure of comparative example 2 was followed, and fish scales were pretreated, homogenized, enzymatically hydrolyzed, fermented, and filtered to obtain collagen.
Comparative example 3
Compared with the example 1, the comparative example 3 does not undergo enzymolysis, and fish scales are pretreated, homogenized, fermented and filtered to obtain the fish scale collagen.
Testing
(1) Yield calculation
The calculation method of the fish scale collagen yield comprises the following steps:
Yield (%) = m1/m2;
wherein m1 is the actual mass of the fish scale collagen protein, g;
m2 is the theoretical mass of the fish scale collagen, g.
(2) Molecular weight distribution test
Dissolving fish scale collagen in 20 times of water by mass, separating by using a membrane to obtain fish scale collagen with molecular weight smaller than 500Da, 500-1000 Da and larger than 1000Da respectively, drying, and calculating the mass ratio in each molecular weight range after drying.
(3) The yields and molecular weight distribution of the fish scale collagen obtained by extraction in examples 1 to 4 and comparative examples 1 to 3 were tested, and the results are shown in the following table 1:
TABLE 1 Scale collagen yield and molecular weight distribution Table
As can be seen from Table 1, according to the technical scheme of the invention, the molecular weight of the extracted fish scale collagen is intensively distributed between 500-1000 Da (the ratio of the extracted fish scale collagen is more than 80%), the ratio of low molecular weight (< 500 Da) to high molecular weight (> 1000 Da) is small, and the molecular weight of the obtained collagen is controlled by adding ammonium citrate, fermenting and enzymolysis sequences, and the types and the addition amounts of zymophyte and bacterial strain. The fish scale collagen obtained by the method of the invention can be directly absorbed by blood without digestion, but is not easy to digest and discharge out of the body due to excessively high metabolism speed, is suitable for organism absorption and utilization, and maximally exerts the efficacy of the collagen.
Claims (10)
1. The extraction method of the fish scale collagen is characterized by comprising the following steps of:
(1) Adding water with the volume of 20-30 times into the washed, demineralized, defatted and washed neutral fish scale powder for homogenating, steaming at 90-110 ℃ for 20-30 min, and cooling to room temperature to obtain fish scale homogenate;
(2) Fermentation: adding 0.1-0.3% of ammonium citrate into the fish scale homogenate, adding mixed bacteria of bacillus subtilis and lactobacillus, and fermenting at 35-38 ℃ for 24-48 hours to obtain fermentation liquor;
(3) Enzymolysis: adding papain, pepsin and trypsin into the fermentation broth in the step (2), carrying out enzymolysis for 1-2 hours at 50-60 ℃, and inactivating enzyme in a boiling water bath to obtain an enzymolysis liquid;
(4) And (3) filtering: and (3) performing preliminary filtration on the enzymolysis liquid in the step (3) by using a filter screen, performing high-speed centrifugal separation on the filtrate to obtain supernatant, and performing freeze drying on the supernatant to obtain the fish scale collagen.
2. The method for extracting fish scale collagen according to claim 1, wherein the amount of bacillus subtilis in the step (2) is 2-5% of the mass of the fish scale powder; the dosage of the lactobacillus is 0.8-1.5% of the mass of the fish scale powder.
3. The method for extracting collagen from fish scales according to claim 1, wherein 1 x 10 4~2×104 U papain, 2 x 10 4~5×104 U pepsin and 1 x 10 3~5×103 U trypsin are added to each kilogram of fish scale powder in the step (3).
4. The method for extracting collagen from fish scales according to claim 1, wherein the high-speed centrifugal separation speed in the step (4) is 10000-18000 rpm.
5. The method for extracting collagen from fish scales according to claim 1, wherein the homogenization condition in the step (1) is 2000-2500 rpm for 8-10 min.
6. The method for extracting collagen from fish scales according to claim 1, wherein the method for washing, demineralizing and degreasing the fish scales in step (1) comprises the steps of: drying the cleaned fish scales, crushing the fish scales into fish scale powder with the particle size of 100-200 meshes, adding the fish scale powder into acid liquor, carrying out ultrasonic soaking for 30-40 min, filtering, washing the solid with water to pH 6-7, adding the washed fish scale powder into alkaline solution, carrying out ultrasonic soaking for 30-40 min, filtering, and washing the solid with water to pH 6-8.
7. The method for extracting fish scale collagen according to claim 1, wherein the acid solution is a citric acid solution with a mass concentration of 1-4%; the alkali liquor is sodium hydroxide solution with the mass concentration of 0.05-0.1%.
8. A fish scale collagen obtained by the extraction method of any one of claims 1 to 7.
9. The fish scale collagen according to claim 8, wherein the mass ratio of molecular weight of the fish scale collagen is more than 80% between 500 and 1000 da.
10. Use of the fish scale collagen according to claim 8 in the processing of food, pharmaceutical and health products.
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN118165100A true CN118165100A (en) | 2024-06-11 |
Family
ID=
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN112062834B (en) | Deep sea fish skin collagen peptide and extraction and preparation method thereof | |
| CN109251954B (en) | Production method of sea cucumber polypeptide | |
| CN109180808B (en) | Fish scale collagen and preparation method and application thereof | |
| CN111793145A (en) | Process for improving quality and yield of sodium chondroitin sulfate co-produced collagen peptide | |
| KR101373940B1 (en) | Method of preparing fermented and enzyme treated silkworm segment extract having high bioactive substances, the silkworm extract obtained thereby, and the use of the silkworm extract having antiinflammatory efficacy | |
| CN111084346A (en) | Preparation process of sea cucumber tablet and sea cucumber tablet prepared by preparation process | |
| CN107365824A (en) | A kind of preparation method for hydrolyzing Isin glue collagen Gly-His-Lys | |
| CN104212863A (en) | Preparation method of feed-use animal protein peptide | |
| CN112608968B (en) | Method for producing fish collagen peptide by using tilapia mossambica scale as raw material | |
| AT398436B (en) | Process for the degradation of polysaccharides | |
| CN110592053A (en) | Collagen hydrolysis complex enzyme, protein oligopeptide and preparation method thereof | |
| Suo-Lian et al. | Technology for extracting effective components from fish scale | |
| CN107200781B (en) | A kind of method that collagen is extracted from giant salamander | |
| CN113151385B (en) | Method for preparing livestock and poultry cartilage collagen polypeptide | |
| JP2870871B2 (en) | A method for treating crustacean shells using enzymes | |
| CN118165100A (en) | Extraction method and application of fish scale collagen | |
| CN107488696B (en) | Chicken bone double-enzymolysis method | |
| CN109468356A (en) | A kind of preparation method of rice protein | |
| CN110818791B (en) | Preparation method of fish collagen | |
| CN114592024A (en) | Sheep placenta polypeptide and preparation method and application thereof | |
| CN113907367A (en) | Preparation method of shaddock soluble dietary fiber | |
| CN1552889A (en) | Use and preparation of collagen protein and collagen polypeptide | |
| CN111773169B (en) | Preparation method of sturgeon placenta extract | |
| CN111471738A (en) | Extraction method of shark chondroitin sulfate | |
| CN105063126A (en) | Method for preparing bacterial cellulose from peanut shells |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication |