CN103194518A - Preparation method of fish collagen peptides with narrow molecular weight ranges - Google Patents
Preparation method of fish collagen peptides with narrow molecular weight ranges Download PDFInfo
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Abstract
The invention discloses a preparation method of fish collagen peptides with narrow molecular weight ranges. The preparation method comprises the following steps of: (1) raw material enzymolysis: carrying out enzymolysis on an aqueous solution of macromolecular raw materials such as fish skins, fish scales and the like by using a biological enzyme to obtain the fish collagen peptides with the molecular weights of 200-2000 daltons, and decoloring by using activated carbon; and (2) carrying out secondary nanofiltration on the fish collagen peptides with the molecular weights of 200-2000 daltons, and removing the fish collagen peptides with the molecular weights higher than 1000 daltons and lower than 500 daltons through filtration to obtain the fish collagen peptides with the molecular weights of 500-1000 daltons. The molecular weights of the fish collagen peptides are graded by using a special biological enzyme and a nanofiltraiton technology in the preparation method disclosed by the invention, so that the collagen peptides which are more concentrative in molecular weight distribution range, higher in physiological activity and suitable for being used as raw materials of cosmetics are obtained.
Description
Technical field
The present invention relates to a kind of processing treatment technology of Isin glue collagen raw material, belong to chemical field.
Background technology
At present, at home and abroad the collagen peptide produced of fish-skin or the fish scale as the cosmetic material that generally use, its range of molecular weight distributions is very wide, 200-2000 dalton.And the molecular weight of the used collagen peptide of cosmetics production requires 500-1000 dalton, is not suitable for the cosmetics industry without the collagen peptide of crossing molecular-weight gradation.In the course of processing, generally use heating under vacuum to concentrate in addition, reduced the physiologically active of product.
Traditionally, in order to obtain the narrower Isin glue collagen peptide of molecular weight ranges, this area has been developed multiple modes such as alkaline process, salt method, acid system, enzyme process raw material has been carried out method for processing, though the applicant finds that these methods can effectively decompose macromolecular collagen protein raw material in industrial practice, but there are many shortcomings in it, especially the Collagen Hydrolysate process is consuming time too of a specified duration, and hydrolysis efficiency is too poor, causes follow-up production technique efficient to be subjected to very big influence.Accelerated hydrolysis rate though adopt enzyme process at certain-length, but adopt biological enzymes such as existing stomach en-, papoid raw material to be hydrolyzed harsh to parameter requests such as temperature, acid-basicity, wayward, even adopt these biological enzymes simultaneously, hydrolytic process also needed about 5 hours.
Summary of the invention
Defective at prior art, the invention discloses a kind of preparation method of Isin glue collagen peptide, can effectively prepare the Isin glue collagen peptide of molecular weight between 500-1000 dalton, compare with existing complete processing, preparation method of the present invention can provide the Isin glue collagen peptide of narrow molecular weight range, molecular weight ranges distributes more concentrated, has more physiologically active, is suitable for to do cosmetic material; And compare with existing scheme, preparation method of the present invention has effectively accelerated process velocity, has reduced preparation parameter requirements such as temperature, acid-basicity, greatly reduces the production difficulty, has improved production efficiency.
For achieving the above object, the present invention is achieved through the following technical solutions:
A kind of preparation method of Isin glue collagen peptide of narrow molecular weight range, comprise the steps: (1) enzymolysis raw material: macromole aqueous solution of raw material such as fish-skin, fish scale are carried out enzymolysis with biological enzyme, obtain molecular weight at the daltonian Isin glue collagen peptide of 200-2000, and through activated carbon decolorizing; (2) be that the daltonian Isin glue collagen peptide of 200-2000 carries out the secondary nanofiltration to molecular weight, the Isin glue collagen peptide that filtering molecular weight 1000 dalton are above and 500 dalton are following, thus obtain the Isin glue collagen peptide of molecular weight between 500-1000.
In above-mentioned steps, step (1) is used for macromolecular raw material feed liquid is efficiently decomposed, and makes its Isin glue collagen peptide that becomes lower molecular weight, thereby supplies raw materials for follow-up nano-filtration step; Step (2) thus the secondary nanofiltration process be used for the too high and low excessively Isin glue collagen peptide of filtering molecular weight and obtain being applicable to Isin glue collagen peptide as cosmetic material.
Wherein, the biological enzyme selection of step (1), temperature, potential of hydrogen control meeting produce a very large impact the whole process of preparation required time.The applicant studies various consumptions and physical parameter by a large amount of experiments, finds to have preparation efficiency preferably under the following conditions: described step (1) detailed process is: with macromole raw material such as fish-skin, fish scale and water with 1: the mass ratio of 1-5 stirs into feed liquid in stirred pot; At 30-45 degree centigrade, the pH value of feed liquid is adjusted to 6.5-8.8 with feed temperature control; Biological enzyme is added in the feed liquid according to the mass ratio with raw material 1: 30-70, stir; Enzymolysis fully after, be warmed up to the 95-100 degree enzyme that goes out.
Preferably, described feed temperature is 40-42 degree centigrade, the pH value of feed liquid is 7.8-8.5, though other temperature range or other pH scope might be the environment that meets bioenzyme activity most, but in test pleasantly surprised discovery of applicant, at 40-42 degree centigrade, the pH value is under this scope of 7.8-8.5, is the hydrolysis that combines biological enzyme activity, macromole stock liquid, the stability of enzymolysis process, the best of breed scope of enzymolysis time.
In the present invention, used biological enzyme can be the arbitrarily available proteolytic enzyme that is used for hydrolysis macromole biological raw material of this area, include but not limited to trypsinase, hydrolysising protease, Sumizyme MP, papoid, Sumizyme MP etc., preferably, described biological enzyme is trypsinase, preferred, described biological enzyme is trypsinase, pepsic mixture, and the mass ratio of the two is 1: 0.1-0.3.
Adopt trypsinase, in conjunction with above-mentioned preferred reaction conditions, the present invention is able to enzymolysis process was accelerated at 3.5-3.8 hour, adopt trypsinase, pepsic mixture, enzymolysis process of the present invention can be finished in 3.0-3.5 hour, with respect to required 5-9 hour of traditional enzymolysis, improved efficient greatly, reduced the time that preparation consumes.
In the present invention, by adopting the nanofiltration membrane technique to high-efficiency to separate the Isin glue collagen peptide of desired molecule weight range, nanofiltration is the technique means of separation differing molecular material emerging in the biotech development process, be a kind of be impellent with the low pressure, film between reverse osmosis and ultrafiltration, aperture, nanofiltration membrane top layer is in nano level scope (10
-9M), divalent ion is had higher rejection, and monovalent ion is had transmitance.
In the present invention, the secondary nanofiltration of described step (2) is respectively: the one-level nanofiltration, the feed liquid of decolouring is passed through the one-level nanofiltration system, and utilize the nanofiltration membrane molecular weight cut-off to be not less than 1000 daltonian Isin glue collagen peptides; The secondary nanofiltration will be passed through the secondary nanofiltration system through the filtrate of one-level nanofiltration system, and molecular weight cut-off is not less than 500 daltonian Isin glue collagen peptides, and the gained concentrated solution is the daltonian collagen peptide of molecular weight 500-1000.
It is as follows that it filters principle: it is 1000 daltonian nanofiltration membrane that molecular weight cut-off is adopted in the one-level nanofiltration, can hold back the above collagen peptide of most molecular weight 1000 dalton, and allow molecular weight see through less than 1000 daltonian collagen peptides, amino acid and water.The concentrated solution of nanofiltration is got back to the feed liquid case and is circulated concentratedly, sees through liquid and enters one-level and see through the liquid hopper.
The secondary nanofiltration: the secondary nanofiltration system adopts molecular weight cut-off 500 daltonian nanofiltration membrane, holds back the above collagen peptide of most molecular weight 500 dalton, and allows molecular weight see through less than 500 daltonian collagen peptides, amino sour water.
Seeing through liquid with first step nanofiltration is charging, enters the secondary nanofiltration system after high-pressure pump, and the concentrated solution of nanofiltration is got back to the one-level nanofiltration and seen through the liquid hopper and circulate concentratedly, sees through liquid and enters the secondary nanofiltration and see through the liquid case.After two-stage nanofiltration classification, the molecular weight of resulting collagen peptide concentrated solution is 500-1000 dalton.
Pass through said process, the invention provides a kind of can to macromole raw materials such as fish-skin, fish scale efficiently resolve into small-molecule substance and according to molecular weight ranges to its preparation method who effectively separates, not only can be with the molecular weight control of product in 500-1000 dalton scope, and in classification, also product has been carried out concentrating and desalination, saved the expense that concentrates with this two procedures of desalination.Not only avoid traditional method Yin Gaowen to make the shortcoming of product sex change, and improved preparation efficiency to greatest extent, reduced the whole required time of preparation process.
Embodiment
In the following embodiments, the present invention has provided preparation method's of the present invention some preferred realization, so that effect of the present invention to be described.Those skilled in the art all belong to protection domain of the present invention to adjustment such as various composition consumptions on this basis.
Embodiment 1
Stir into feed liquid during fresh fish skin and water added jar according to 1: 2 ratio, feed temperature is heated to 45 degree, the pH value of feed liquid is adjusted to 6.5, according in feed liquid, adding trypsinase with 1: 50 ratio of fish-skin raw material, stir, 4.5 enzymolysis finishes after hour, is warmed up to 95 degree again, keeps 30 minutes enzymes that go out.
Decolouring: reduce the temperature to 75 degree after the enzyme that goes out finishes, add the gac of fish-skin raw materials quality 9%, decoloured 30 minutes.
Secondary filtration: at first the feed liquid of decolouring is carried out pre-treatment, enzymolysis feed liquid through activated carbon decolorizing enters the feed liquid case, be pressurized to cloth envelop collector (aperture of preferred cloth bag type filter is 2 μ m) through topping-up pump, remove oarse-grained suspended substance and partial organic substances in the waste water, colloid, to improve film feeding liquid quality, enter first step nanofiltration concentration systems then.
The one-level nanofiltration: adopting molecular weight cut-off is 1000 daltonian nanofiltration membrane, holds back the above collagen peptide of most molecular weight 1000 dalton, allows molecular weight see through less than 1000 daltonian collagen peptides, amino acid and water.The concentrated solution of nanofiltration is got back to the feed liquid case and is circulated concentratedly, sees through liquid and enters one-level and see through the liquid hopper.
The secondary nanofiltration: seeing through liquid with the one-level nanofiltration is raw material, enters the secondary nanofiltration device after high-pressure pump, and the concentrated solution of nanofiltration is got back to one-level and seen through the liquid hopper and circulate concentratedly, sees through liquid and enters secondary and see through the liquid case.Adopt molecular weight cut-off 500 daltonian nanofiltration membrane, hold back the above collagen peptide of most molecular weight 500 dalton, allow molecular weight see through less than 500 daltonian collagen peptides, amino sour water.
After two-stage nanofiltration classification, the molecular weight of resulting collagen peptide concentrated solution is 500-1000 dalton, and this concentrated solution gets product through after the spraying drying.
Embodiment 2
Stir into feed liquid during fresh fish skin and water added jar according to 1: 2 ratio, be heated to 40 degree, the pH value of feed liquid is adjusted to 8.0,1: 50 consumption of quality adding trypsinase according to fish-skin stirs, 3.8 enzymolysis finishes after hour, and feed liquid is warmed up to 95 degree, keeps 30 minutes enzymes that go out.
Decolouring: reduce the temperature to 80 degree after the enzyme that goes out finishes, add 15% gac of fish-skin weight, decoloured 25 minutes.
The secondary nanofiltration:
Pre-treatment: filtrate enters the feed liquid case, is pressurized to cloth envelop collector through topping-up pump, enters first step nanofiltration system.
The one-level nanofiltration: it is 1000 daltonian nanofiltration membrane that molecular weight cut-off is adopted in the one-level nanofiltration, can hold back the above collagen peptide of most molecular weight 1000 dalton, allows molecular weight see through less than 1000 daltonian collagen peptides, amino acid and water.The concentrated solution of nanofiltration is got back to the feed liquid case and is circulated concentratedly, sees through liquid and enters one-level and see through the liquid hopper.
The secondary nanofiltration: seeing through liquid with the one-level nanofiltration is raw material, enters the secondary nanofiltration device after high-pressure pump, and the concentrated solution of nanofiltration is got back to one-level and seen through the liquid hopper and circulate concentratedly, sees through liquid and enters secondary and see through the liquid case.Adopt molecular weight cut-off 500 daltonian nanofiltration membrane, hold back the above collagen peptide of most molecular weight 500 dalton, allow molecular weight see through less than 500 daltonian collagen peptides, amino sour water.
After two-stage nanofiltration classification, the molecular weight of resulting collagen peptide concentrated solution is 500-1000 dalton, and this concentrated solution gets product through after the spraying drying.
Embodiment 3
Stir into feed liquid during fresh fish skin and water added jar according to 1: 2 ratio, be heated to 40 degree, the pH value of feed liquid is adjusted to 8.2, according to the quality of fish-skin, add trypsinase according to 1: 50 ratio, enzymolysis finishes after 3.7 hours, be warmed up to 95 degree, keep 20 minutes enzymes that go out.
Decolouring: reduce the temperature to 80 degree after the enzyme that goes out finishes, add 10% gac of fish-skin weight, decoloured 30 minutes.
The secondary nanofiltration:
Pre-treatment: filtrate enters the feed liquid case, is pressurized to cloth envelop collector through topping-up pump, enters first step nanofiltration system.
The one-level nanofiltration: it is 1000 daltonian nanofiltration membrane that molecular weight cut-off is adopted in the one-level nanofiltration, can hold back the above collagen peptide of most molecular weight 1000 dalton, allows molecular weight see through less than 1000 daltonian collagen peptides, amino acid and water.The concentrated solution of nanofiltration is got back to the feed liquid case and is circulated concentratedly, sees through liquid and enters one-level and see through the liquid hopper.
The secondary nanofiltration: seeing through liquid with the one-level nanofiltration is raw material, enters the secondary nanofiltration device after high-pressure pump, and the concentrated solution of nanofiltration is got back to one-level and seen through the liquid hopper and circulate concentratedly, sees through liquid and enters secondary and see through the liquid case.Adopt molecular weight cut-off 500 daltonian nanofiltration membrane, hold back the above collagen peptide of most molecular weight 500 dalton, allow molecular weight see through less than 500 daltonian collagen peptides, amino sour water.
After two-stage nanofiltration classification, the molecular weight of resulting collagen peptide concentrated solution is 500-1000 dalton, and this concentrated solution gets product through after the spraying drying.
Embodiment 4
Stir into feed liquid during fresh fish skin and water added jar according to 1: 2 ratio, be heated to 40 degree, the pH value of feed liquid is adjusted to 7.9, quality according to fish-skin, add trypsinase according to 1: 50 ratio: 1: 0.2 mixture of stomach en-, 3.4 enzymolysis finishes after hour, is warmed up to 95 degree, keeps 20 minutes enzymes that go out.
Decolouring: reduce the temperature to 80 degree after the enzyme that goes out finishes, add 10% gac of fish-skin weight, decoloured 30 minutes.
The secondary nanofiltration:
Pre-treatment: filtrate enters the feed liquid case, is pressurized to cloth envelop collector through topping-up pump, enters first step nanofiltration system.
The one-level nanofiltration: it is 1000 daltonian nanofiltration membrane that molecular weight cut-off is adopted in the one-level nanofiltration, can hold back the above collagen peptide of most molecular weight 1000 dalton, allows molecular weight see through less than 1000 daltonian collagen peptides, amino acid and water.The concentrated solution of nanofiltration is got back to the feed liquid case and is circulated concentratedly, sees through liquid and enters one-level and see through the liquid hopper.
The secondary nanofiltration: seeing through liquid with the one-level nanofiltration is raw material, enters the secondary nanofiltration device after high-pressure pump, and the concentrated solution of nanofiltration is got back to one-level and seen through the liquid hopper and circulate concentratedly, sees through liquid and enters secondary and see through the liquid case.Adopt molecular weight cut-off 500 daltonian nanofiltration membrane, hold back the above collagen peptide of most molecular weight 500 dalton, allow molecular weight see through less than 500 daltonian collagen peptides, amino sour water.
After two-stage nanofiltration classification, the molecular weight of resulting collagen peptide concentrated solution is 500-1000 dalton, and this concentrated solution gets product through after the spraying drying.
Embodiment 5
Stir into feed liquid during fresh fish skin and water added jar according to 1: 2 ratio, be heated to 42 degree, the pH value of feed liquid is adjusted to 8.1, quality according to fish-skin, add trypsinase according to 1: 50 ratio: 1: 0.3 mixture of stomach en-, 3.3 enzymolysis finishes after hour, is warmed up to 95 degree, keeps 20 minutes enzymes that go out.
Decolouring: reduce the temperature to 75 degree after the enzyme that goes out finishes, add 12% gac of fish-skin weight, decoloured 30 minutes.
The secondary nanofiltration:
Pre-treatment: filtrate enters the feed liquid case, is pressurized to cloth envelop collector through topping-up pump, enters first step nanofiltration system.
The one-level nanofiltration: it is 1000 daltonian nanofiltration membrane that molecular weight cut-off is adopted in the one-level nanofiltration, can hold back the above collagen peptide of most molecular weight 1000 dalton, allows molecular weight see through less than 1000 daltonian collagen peptides, amino acid and water.The concentrated solution of nanofiltration is got back to the feed liquid case and is circulated concentratedly, sees through liquid and enters one-level and see through the liquid hopper.
The secondary nanofiltration: seeing through liquid with the one-level nanofiltration is raw material, enters the secondary nanofiltration device after high-pressure pump, and the concentrated solution of nanofiltration is got back to one-level and seen through the liquid hopper and circulate concentratedly, sees through liquid and enters secondary and see through the liquid case.Adopt molecular weight cut-off 500 daltonian nanofiltration membrane, hold back the above collagen peptide of most molecular weight 500 dalton, allow molecular weight see through less than 500 daltonian collagen peptides, amino sour water.
After two-stage nanofiltration classification, the molecular weight of resulting collagen peptide concentrated solution is 500-1000 dalton, and this concentrated solution gets product through after the spraying drying.
Embodiment 6
Stir into feed liquid during fresh fish skin and water added jar according to 1: 2 ratio, be heated to 40 degree, the pH value of feed liquid is adjusted to 8.0, quality according to fish-skin, add trypsinase according to 1: 45 ratio: 1: 0.3 mixture of stomach en-, 3.1 enzymolysis finishes after hour, is warmed up to 90 degree, keeps 30 minutes enzymes that go out.
Decolouring: reduce the temperature to 75 degree after the enzyme that goes out finishes, add 12% gac of fish-skin weight, decoloured 30 minutes.
The secondary nanofiltration:
Pre-treatment: filtrate enters the feed liquid case, is pressurized to cloth envelop collector through topping-up pump, enters first step nanofiltration system.
The one-level nanofiltration: it is 1000 daltonian nanofiltration membrane that molecular weight cut-off is adopted in the one-level nanofiltration, can hold back the above collagen peptide of most molecular weight 1000 dalton, allows molecular weight see through less than 1000 daltonian collagen peptides, amino acid and water.The concentrated solution of nanofiltration is got back to the feed liquid case and is circulated concentratedly, sees through liquid and enters one-level and see through the liquid hopper.
The secondary nanofiltration: seeing through liquid with the one-level nanofiltration is raw material, enters the secondary nanofiltration device after high-pressure pump, and the concentrated solution of nanofiltration is got back to one-level and seen through the liquid hopper and circulate concentratedly, sees through liquid and enters secondary and see through the liquid case.Adopt molecular weight cut-off 500 daltonian nanofiltration membrane, hold back the above collagen peptide of most molecular weight 500 dalton, allow molecular weight see through less than 500 daltonian collagen peptides, amino sour water.
After two-stage nanofiltration classification, the molecular weight of resulting collagen peptide concentrated solution is 500-1000 dalton, and this concentrated solution gets product through after the spraying drying.
With following method the molecular weight of the Isin glue collagen peptide of above-mentioned preparation is measured, the molecular weight distribution of products therefrom is as shown in table 1.
Measuring method: come the distribution of detection molecules amount by high performance liquid chromatography, the condition of molecular weight detection is as follows:
Chromatographic column: TSKgel G2000SWXL300mmx7.8mm
Moving phase: acetonitrile: water: trifluoroacetic acid is 45: 55: 0.1 (volume ratio)
Detect wavelength: UV220nm
Flow velocity: 0.5mL/min
Column temperature: 30 ℃
Sampling volume: 10 μ L.
Detected result is as shown in the table:
Table 1: Isin glue collagen peptide detected result
From as can be seen above-mentioned, preparation method of the present invention effectively sub-argument goes out the Isin glue collagen peptide of molecular weight ranges 500-1000, and preparation method of the present invention can isolate the Isin glue collagen peptide that the molecular weight ranges distribution uniformity is good, content of peptides is high under most preferred biological enzyme, enzymatic hydrolysis condition.
Claims (6)
1. the preparation method of the Isin glue collagen peptide of a narrow molecular weight range is characterized in that comprising the steps:
(1) enzymolysis raw material: macromole aqueous solution of raw material such as fish-skin, fish scale are carried out enzymolysis with biological enzyme, obtain molecular weight at the daltonian Isin glue collagen peptide of 200-2000, and through activated carbon decolorizing; (2) be that the daltonian Isin glue collagen peptide of 200-2000 carries out the secondary nanofiltration to molecular weight, the Isin glue collagen peptide that filtering molecular weight 1000 dalton are above and 500 dalton are following, thus obtain the Isin glue collagen peptide of molecular weight between 500-1000.
2. preparation method according to claim 1 is characterized in that described step (1) detailed process is: with macromole raw materials such as fish-skin, fish scale and water with 1: the mass ratio of 1-5 stirs into feed liquid in stirred pot; At 30-45 degree centigrade, the pH value of feed liquid is adjusted to 6.5-8.8 with feed temperature control; Biological enzyme is added in the feed liquid according to the mass ratio with feed liquid 1: 30-70, stir; Enzymolysis fully after, be warmed up to the 95-100 degree enzyme that goes out.
3. preparation method according to claim 2 is characterized in that described feed temperature is 40-42 degree centigrade, and the pH value of feed liquid is 7.8-8.5.
4. preparation method according to claim 2 is characterized in that described biological enzyme is trypsinase.
5. preparation method according to claim 2 is characterized in that described biological enzyme is trypsinase, pepsic mixture, and the mass ratio of the two is 1: 0.1-0.3.
6. preparation method according to claim 1, the secondary nanofiltration that it is characterized in that described step (2) is respectively: the one-level nanofiltration, the feed liquid of decolouring is passed through the one-level nanofiltration system, utilize the nanofiltration membrane molecular weight cut-off to be not less than 1000 daltonian Isin glue collagen peptides; The secondary nanofiltration will be passed through the secondary nanofiltration system through the filtrate of one-level nanofiltration system, and molecular weight cut-off is not less than 500 daltonian Isin glue collagen peptides, and the gained concentrated solution is the daltonian collagen peptide of molecular weight 500-1000.
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| CN104140992A (en) * | 2013-11-26 | 2014-11-12 | 国家海洋局第三海洋研究所 | Large-scale preparation method of fish scale type I collagen peptides |
| CN109971812A (en) * | 2019-05-09 | 2019-07-05 | 王明 | A kind of processing technology hydrolyzing Isin glue collagen |
| CN110144376A (en) * | 2019-06-05 | 2019-08-20 | 杭州柏淼生物科技有限公司 | Nanoscale collagen peptide and preparation method thereof |
| CN111118094A (en) * | 2020-02-24 | 2020-05-08 | 长春大学 | Preparation method of cod skin collagen peptide |
| CN114129712A (en) * | 2021-12-16 | 2022-03-04 | 天津知了大健康产业发展有限公司 | Whitening collagen peptide freeze-dried powder and preparation method thereof |
| CN117126907A (en) * | 2023-04-14 | 2023-11-28 | 海南华研胶原科技股份有限公司 | III type collagen and preparation method thereof |
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| CN114129712A (en) * | 2021-12-16 | 2022-03-04 | 天津知了大健康产业发展有限公司 | Whitening collagen peptide freeze-dried powder and preparation method thereof |
| CN117126907A (en) * | 2023-04-14 | 2023-11-28 | 海南华研胶原科技股份有限公司 | III type collagen and preparation method thereof |
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