CN101607982B - Fish skin collagen peptide with whitening effect and preparation process and application thereof - Google Patents

Fish skin collagen peptide with whitening effect and preparation process and application thereof Download PDF

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CN101607982B
CN101607982B CN2008100480836A CN200810048083A CN101607982B CN 101607982 B CN101607982 B CN 101607982B CN 2008100480836 A CN2008100480836 A CN 2008100480836A CN 200810048083 A CN200810048083 A CN 200810048083A CN 101607982 B CN101607982 B CN 101607982B
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collagen peptide
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陈栋梁
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TIANTIANHAO BIOLOGICAL PRODUCTS CO Ltd WUHAN CITY
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Abstract

The invention discloses fish skin collagen peptide with the whitening effect and a preparation process and application thereof, and in particular relates to fish skin collagen peptide obtained by using alkali solution to process fish skin, using a protease hydrolysis method to process the fish skin and performing a series of technical treatment of decoloration, deodorization and impurity removal on hydrolysis liquid and a preparation process thereof and application of the fish skin collagen peptide as a cosmetic or an additive of skin-care cosmetics. Compared with the common collagen peptide, the fish skin collagen peptide has high content of small molecular peptides, lower ash content, high protein content, and greatest whitening effect among like products in the market; and the fish skin collagen peptide as the cosmetic or the additive of the skin-care cosmetics without toxic and side effect has wide development prospect.

Description

A kind of fish skin collagen peptide of Pear Power effect and preparation technology thereof and purposes
Technical field
The present invention relates to a kind of fish skin collagen peptide and preparation technology and purposes of Pear Power effect.Specifically; The present invention relates to a kind of alkaline solution that utilizes and handle fish-skin; Utilize the mode of protease hydrolysis to handle above-mentioned fish-skin again; The fish skin collagen peptide of the Pear Power effect that obtains through the technical finesse of hydrolyzed solution being carried out a series of decolourings, take off flavor, removing impurity, and preparation technology and as the purposes of makeup or skin-care product additive.
Background technology
Collagen is a kind of scleroproein in the organism, mainly is present in the tissues such as skin, bone, cartilage and tendon, accounts for 25%~33% of human body or other animal body total protein contents.Collagen protein is widely used in biomedical materials field because of its good biological characteristics.Therefore, the extraction of collagen how to carry out the biologically active of high purity, high yield is the problem that numerous investigators pay close attention to always.The history of from pigskin and ox-hide, extracting collagen is longer, and because its raw material is sufficient, remains the main source of collagen so far; Collagen mainly extracts from animal tissuess such as ox and pig for a long time.The people that make of mad cow disease and foot and mouth disease begin to consider to use the substitute of ox, pig collagen in recent years, and the extraction of fish skin collagen becomes the new focus of research in recent years.The minimum unit of collagen is that collagen tripeptide (Collagen tripeptide) is called for short CTP.Its structure can be expressed as (Gly-X-Y), is a kind of highly purified collagen tripeptide that has glycocoll at the N end.The research of trace ray auto photographing method shows that CTP has the molecular-weight average of about 280Da.And most collagen peptide in the market mainly constitute polyamino acid by 30 to 100 amino acid. molecular-weight average reaches 3000-10000Da, and its bioavailability is limited.
Summary of the invention
The purpose of this invention is to provide a kind of is raw material with the fish-skin, is rich in the fish skin collagen peptide of the Magnesium Ascorbylphosphate of small-molecular peptides (molecular weight 280Da).
Another object of the present invention provides the preparation technology of the fish skin collagen peptide of said Magnesium Ascorbylphosphate.
Another object of the present invention provides the purposes of the fish skin collagen peptide of said Magnesium Ascorbylphosphate.
Technical scheme of the present invention is: a kind of fish skin collagen peptide of Magnesium Ascorbylphosphate, form by following prepared, and said technology comprises the steps: successively
1) takes by weighing the fish-skin raw material, clean, drain away the water, weigh;
2) fish-skin of above-mentioned processing is added the water of 5~6 times of its quality, add the calcium hydroxide of its quality 3.0~5.0% again, stir and soaked 5~7 days, in the NCP in removing fish-skin, also played certain raw meat effect of going;
3) fish-skin of above-mentioned processing is cleaned to neutral, drained away the water, weigh, the gained fish-skin is the raw material after the pre-treatment;
4) with above-mentioned steps 3) fish-skin raw material after the pre-treatment that obtains is according to its quality 5~6 water extraordinarily; 121 ℃ of high pressure 60~90min are cooled to 50~60 ℃, double gauze elimination residue; Adjust pH is 6.0~8.0; 50 ℃ of waters bath with thermostatic control at first add the bacillus alkaline protease of fish-skin raw materials quality 0.5~1.0%, add the bromeline of fish-skin raw materials quality 0.5~1.0% and the neutral protease of fish-skin raw materials quality 0.5~1.0% more simultaneously; Pick up counting from adding bacillus alkaline protease, enzymolysis is 5~6 hours altogether.After enzymolysis finishes, more than 95 ℃, 10 minutes enzymes that go out, cooling, filter cloth filters, and filtrating is the fish skin collagen peptide hydrolysate;
5) above-mentioned filtrating is taken off flavor with the K15 activity carbon column decoloring;
The filtrating of 6) above-mentioned decolouring being taken off behind the flavor is concentrated into suitable volume, and spraying drying makes finished product.
Another technical scheme of the present invention is: a kind of technology for preparing the fish skin collagen peptide of Magnesium Ascorbylphosphate is characterized in that: comprise the steps: successively
1) takes by weighing the fish-skin raw material, clean, drain away the water, weigh;
2) fish-skin of above-mentioned processing is added the water of 5~6 times of its quality, add the calcium hydroxide of its quality 3.0~5.0% again, stir and soaked 5~7 days, in the NCP in removing fish-skin, also played certain raw meat effect of going;
3) fish-skin of above-mentioned processing is cleaned to neutral, drained away the water, weigh, the gained fish-skin is the raw material after the pre-treatment;
4) with above-mentioned steps 3) fish-skin raw material after the pre-treatment that obtains is according to its quality 5~6 water extraordinarily; 121 ℃ of high pressure 60~90min are cooled to 50~60 ℃, double gauze elimination residue; Adjust pH is 6.0~8.0; 50 ℃ of waters bath with thermostatic control at first add the bacillus alkaline protease of fish-skin raw materials quality 0.5~1.0%, add the bromeline of fish-skin raw materials quality 0.5~1.0% and the neutral protease of fish-skin raw materials quality 0.5~1.0% more simultaneously; Pick up counting from adding bacillus alkaline protease, enzymolysis is 5~6 hours altogether.After enzymolysis finishes, more than 95 ℃, 10 minutes enzymes that go out, cooling, filter cloth filters, and filtrating is the fish skin collagen peptide hydrolysate;
5) above-mentioned filtrating is taken off flavor with the K15 activity carbon column decoloring;
The filtrating of 6) above-mentioned decolouring being taken off behind the flavor is concentrated into suitable volume, and spraying drying makes finished product.
Another technical scheme of the present invention is: the fish skin collagen peptide of the described Magnesium Ascorbylphosphate of a kind of claim 1 is as the purposes of makeup or skin-care product additive.
Fish skin collagen peptide of the present invention; Improve small-molecular peptides in the fish skin collagen peptide (about molecular weight 280Da) components contents to greatest extent; Experiment proof: because the CTP molecular weight is very little; Thereby can effectively infiltrate stratum corneum, in skin corium and the hair root cell, can promote that the synthetic and skin corium of collagen is fibrocellular hyaluronic synthetic.The CTP component is made an addition in the daily necessities of skin cleaning liquid class; Will be to the elasticity of skin; Wettability demonstrates very excellent effect with the coarse state that improves skin; And have safe, the cell agglutinating value(of coal) is good, transparent, odorless and characteristics such as have no stimulation, and can be used as makeup or skin-care product additive, and vast market prospect is arranged.
The fish skin collagen peptide of the Pear Power effect of producing by preparation technology of the present invention; Its small-molecular peptides that is rich in (molecular weight 280Da) can be effectively and the tyrosine oxidase effect; Make its inactivation or blocking-up tyrosine generate melanic oxidative pathway; Except that nutritive ingredient, also have on the market white-skinned face function the strongest in the like product with general collagen product.
Description of drawings
Accompanying drawing 1 prepares the process flow diagram of the collagen peptide of Magnesium Ascorbylphosphate for the present invention.
Embodiment
Understand the technology that the present invention prepares the fish skin collagen peptide of Magnesium Ascorbylphosphate through embodiment below, but following embodiment does not limit content of the present invention.
The technology of the fish skin collagen peptide of preparation Magnesium Ascorbylphosphate:
1) raw material pre-treatment.Clean fish-skin, control water (weighing), add the water of 5~6 times of its quality, add the calcium hydroxide of its quality 3.0~5.0% again, stir and soaked 5~7 days, with the fish-skin after the abundant cleaning and dipping of tap water, till about pH value to 7.0, control water (weighing) is subsequent use.Through alkaline solution treatment process of the present invention, when removing the NCP in the raw material, also played certain raw meat effect of going.
2) sex change.According to 5~6 zero(ppm) water extraordinarily of above-mentioned wet basis raw materials quality, 121 ℃ of high pressure sex change 60~90min.
3) filter.Hydrolyzed solution after the above-mentioned sex change is cooled to 50~60 ℃, and with double gauze elimination fish scale residue and grease, it is inferior that visual grease removal situation repeats to filter youngster.Moisture evaporated when adding distil water is supplied sex change.
4) protease hydrolysis.Adjust pH to 6.0~8.0; Constant temperature is at 50 ℃; In above-mentioned filtrating, add the bacillus alkaline protease of 1 said wet basis raw materials quality 0.5~1.0%, add the bromeline of 1 said wet basis raw materials quality 0.5~1.0% and the neutral protease of 1 said wet basis raw materials quality 0.5~1.0% more simultaneously, pick up counting from adding bacillus alkaline protease; Stirred enzymolysis altogether 5~6 hours (during moisturizing in good time, keep the material liquid mixture constancy of volume).
5) deactivation.With said hydrolyzed liquid more than 95 ℃, deactivation 10min.
6) filter.Hydrolyzed solution after the above-mentioned deactivation is cooled to 50~60 ℃, and filter cloth filters.
7) the active carbon column adsorption bleaching takes off flavor.Above-mentioned filtrating is taken off flavor with the K15 activity carbon column decoloring, note liquid running balance in the control post, in order to avoid leak carbon.
8) drying.Above-mentioned filtrating is concentrated into suitable volume, and spraying drying makes finished product.
Embodiment 1 according to the collagen peptide of above-mentioned prepared Magnesium Ascorbylphosphate:
1, the fish-skin that thaws freezes piece 10kg, cleans residual meat and bloodstain, drain weigh about 9kg fish-skin; Add 54L nature water and 270g calcium hydroxide; Mixing soaks 5 days (stirring every day 2 times), after 6 days, with nature water (200L; 2h consuming time) be washed till neutrality, draining weighs must refrigerate subsequent use by 6 kilograms of fish-skins (fish scale of washing off discards) approximately.
2, platform balance takes by weighing the subsequent use fish-skin of 1kg refrigeration, presses 1: 5 adding distil water 5L of solid-liquid ratio, 121 ℃ of high pressure sex change 60min.
3, double gauze filters and removes the fish-skin residue, 50 ℃ of waters bath with thermostatic control, and adjust pH is 7.0; Add the 7g bacillus alkaline protease; Add 7g bromeline and 7g neutral protease again, pick up counting, stir enzymolysis 5h altogether automatically from adding bacillus alkaline protease.
4, water-bath deactivation (95 ℃ 10min), are cooled to 50 ℃~60 ℃.
5,300 order filter clothes filter, and get the about 5000ml of filtrate volume.
6, active carbon column (K15 granulated carbon, blade diameter length ratio 1: 4) takes off flavor, adds 2000ml distillation washing carbon post, is eluted to total liquid volume 7000ml
7, filtrating spraying drying, 105 the gram Magnesium Ascorbylphosphates fish skin collagen peptide.
Embodiment 2 according to the collagen peptide of above-mentioned prepared Magnesium Ascorbylphosphate:
1, the fish-skin that thaws freezes piece 9kg, cleans residual meat and bloodstain, drain weigh about 8kg fish-skin; Add 40L nature water and 240g calcium hydroxide; Mixing soaks 7 days (stirring every day 2 times), after 7 days, with nature water (200L; 2h consuming time) be washed till neutrality, draining weighs must refrigerate subsequent use by 5.5 kilograms of fish-skins (fish scale of washing off discards) approximately.
2, platform balance takes by weighing the subsequent use fish-skin of 1kg refrigeration, presses 1: 5.5 adding distil water 5.5L of solid-liquid ratio, 121 ℃ of high pressure sex change 80min.
3, double gauze filters and removes the fish-skin residue, 50 ℃ of waters bath with thermostatic control, and adjust pH is 8.0; Add the 9g bacillus alkaline protease; Add 9g bromeline and 9g neutral protease again, pick up counting, stir enzymolysis 5.5h altogether automatically from adding bacillus alkaline protease.
4, water-bath deactivation (95 ℃ 10min), are cooled to 50 ℃~60 ℃.
5,300 order filter clothes filter, and get the about 5500ml of filtrate volume.
6, active carbon column (K15 granulated carbon, blade diameter length ratio 1: 4) takes off flavor, adds 1800ml distillation washing carbon post, is eluted to total liquid volume 7300ml
7, filtrating spraying drying, 110 the gram Magnesium Ascorbylphosphates fish skin collagen peptide.
Product index by the fish skin collagen peptide of the Magnesium Ascorbylphosphate of prepared in this embodiment is:
Total nitrogen >=15.5%
Moisture≤5.0% ash content≤2%
Fat≤0.5% oxyproline accounts for protein contnt 8.5~10.0%
Polypeptide>=90% molecular-weight average, 90%≤1000D
Its component 90% left and right sides molecular weight is below 1000Da, and wherein small-molecular peptides (molecular weight 280Da) content is about 50%, and its physical and chemical index is as shown in table 1:
Table 1 fish skin collagen peptide physical and chemical index table
White-skinned face function research work of the present invention is summarized as follows:
1, experimental principle
The kind and the quantity of the pigment that exists in the color of human skin and the skin are relevant, and melanochrome has the greatest impact to skin color.Melanochrome forms in the melanocyte of epidermis basis pontis.Its process is that the tyrosine in the melanocyte generates DOPA under the effect of tyrosine oxidase, the DOPA quinone finally forms melanochrome.Tyrosine oxidase is the key enzyme that melanochrome generates, and the active or blocking-up tyrosine of restraint of tyrosinase generates melanic oxidative pathway and can reach the effect of whitening.
2 materials and equipment
2.1 tried thing:
Fish skin collagen peptide: be called for short fish, self-control;
Other samples: the collagen product of the same type on the market, amount to 7 kinds, count sample 1-sample 7.
2.2 major equipment and instrument
2.2.1 reagent:
Tyrosine oxidase (Sigma produces, and Beijing chemical reagents corporation buys): 7mg/100ml;
L-DOPA solution (Shanghai Hanhong Chemical Industry Co., Ltd. provides): 5g/L;
Kojic acid (Beijing Brilliance Biochemical Co., Ltd. provides): 20g/L;
PH value 6.8 phosphoric acid buffers: potassium primary phosphate (AR Henan Jiaozuo City chemical industry three factories) 6.8g, sodium hydroxide (AR Shantou Xilong Chemical Factory) 0.944g are settled to 1000ml.
2.2.2 instrument:
Thermostat water bath; The HHW21600 of Qin Tai medical apparatus and instruments factory of Wuhan City type constant temperature water tank;
Ultraviolet spectrophotometer: Shanghai Precision Scientific Apparatus Co., Ltd's 722 type visible spectrophotometers;
3 TPs
3.1 sample preparation: it is subsequent use to be tried thing wiring solution-forming (tyrosine oxidase is a cuproprotein, should not contain cupric ion in the solution) with ultrapure water.
3.2 experimental implementation: in glass test tube, carry out biochemical reaction, each pipe adds required damping fluid (pH6.8 phosphoric acid buffer), sample liquid, kojic acid solution and enzyme solution.In 37 ℃ of water-baths, react 10min, add DOPA solution successively, in 37 ℃ of water-baths, react 10min again, survey the 475nmOD value.Table 2 is seen in the experimental system design.
Table 2 experimental system design table
Reagent Sample hose (ml) Standard control (ml) Positive control (ml) Negative control (ml)
Damping fluid 0.8 0.8 1.8 2.0
Sample liquid 1.0 - - 1.0
Enzyme solution 0.2 0.2 0.2 -
Kojic acid solution - 1.0 - -
DOPA solution 1.0 1.0 1.0 -
Annotate: blank adopts the 3ml ultrapure water
3.3 experiment Quality Control condition
3.3.1 kojic acid standard control result: inhibiting rate is lower than positive control.Kojic acid is the whitening additive of using always, and its effect site and the mechanism of action are clear, and enzyme dynamic curve correlationship is close, and the tyrosinase activity inhibition strength is moderate, so it is used for the accuracy of test experiment operation and result's safety as the standard control thing.
3.3.2OD the value measured result should be: positive control>sample (containing standard control)>negative control.
3.4 effect is estimated method for expressing
3.4.1 inhibiting rate method of calculation: sample is represented with inhibiting rate the inhibition strength of tyrosinase activity.With reference to Li Li introduce than short-cut method; With the positive control result is contrast; Calculate inhibiting rate by following formula: inhibiting rate=[positive control light absorption value-(sample hose light absorption value-negative control light absorption value)]/positive control light absorption value * 100%, inhibiting rate height show that it is high to the enzymic activity inhibition strength.
3.4.2 standard substance reference:, do in the laboratory when detecting and all should be provided with standard control for guaranteeing the accuracy of experimental result.We select for use kojic acid as standard control, and the kojic acid inhibiting rate that will under positive control OD value is about 0.3 condition, record is the reference control value.In different batches when test,, the inhibiting rate of kojic acid standard control can be different, can the inhibiting rate of standard control be converted to 100%, makes comparisons after the inhibiting rate equal proportion of being tried thing changes again.
4 results and analysis
4.1 tried the detection and the comparison of thing tyrosinase inhibition rate
Tried thing according to 3.2 experimental implementation and be made into 2% concentration and make an experiment, and convert through the method for 3.4.2, the result is as shown in table 3:
Table 3 respectively tried the active effect table of thing restraint of tyrosinase (X ± s, n=8)
? Standard control Fish Sample 1 Sample 2 Sample 3
Inhibiting rate % 9208±1.79 33.66±2.35 4.41±0.56 11.66±0.51 6.63±0.35
Variation coefficient % (CV) 194 699 12.59 438 5.24
? Sample 4 Sample 5 Sample 6 Sample 7 ?
Inhibiting rate % 10.45±0.69 9.32±0.39 12.26±0.47 9.14±0.60 ?
Variation coefficient % (CV) 6.63 414 3.85 6.58 ?
Can find out by table 3; The ability that fish skin collagen peptide restraint of tyrosinase activity or blocking-up tyrosine generate melanic oxidative pathway is better than the like product on the market; Its small-molecular peptides that is rich in (molecular weight 280Da) can be effectively and the tyrosine oxidase effect, makes its inactivation or blocking-up tyrosine generate melanic oxidative pathway.With regard to precision, respectively being tried the thing variation coefficient of (comprising standard control) (CV) is: 1.94~12.59%, all meet normal condition variation (RCV) less than 15% requirement, and major part meets top condition variation (OCV) less than 7% requirement.
Can know by above analysis; Use the fish skin collagen peptide of the Magnesium Ascorbylphosphate of prepared in this embodiment; Except that nutritive ingredient with general collagen product; Also have on the market white-skinned face function the strongest in the like product, can be used as the makeup or the skin-care product additive that have no side effect, have extensive market prospects.

Claims (6)

1. the fish skin collagen peptide of a Magnesium Ascorbylphosphate is formed by following prepared, and said technology comprises the steps: successively
1) takes by weighing the fish-skin raw material, clean, drain away the water, weigh;
2) fish-skin of above-mentioned processing is added the water of 5~6 times of its quality, add the calcium hydroxide of its quality 3.0~5.0% again, stir and soaked 5~7 days, in the NCP in removing fish-skin, also played certain raw meat effect of going;
3) fish-skin of above-mentioned processing is cleaned to neutral, drained away the water, weigh, the gained fish-skin is the raw material after the pre-treatment;
4) with above-mentioned steps 3) fish-skin raw material after the pre-treatment that obtains is according to its quality 5~6 water extraordinarily, and 121 ℃ of high pressure 60~90min are cooled to 50~60 ℃, double gauze elimination residue; Adjust pH is 6.0~8.0,50 ℃ of waters bath with thermostatic control, at first adds the bacillus alkaline protease of fish-skin raw materials quality 0.5~1.0%; Add the bromeline of fish-skin raw materials quality 0.5~1.0% and the neutral protease of fish-skin raw materials quality 0.5~1.0% more simultaneously, pick up counting from adding bacillus alkaline protease, enzymolysis is 5~6 hours altogether; After enzymolysis finishes, more than 95 ℃, 10 minutes enzymes that go out; Cooling, filter cloth filters, and filtrating is the fish skin collagen peptide hydrolysate;
5) above-mentioned filtrating is taken off flavor with the K15 activity carbon column decoloring;
The filtrating of 6) above-mentioned decolouring being taken off behind the flavor is concentrated into suitable volume, and spraying drying makes finished product.
2. by the described fish skin collagen peptide of claim 1, it is characterized in that: the molecular weight of the component 90% of said fish skin collagen peptide is below 1000Da, and wherein the small-molecular peptides content of molecular weight 280Da is 50%.
3. by the described fish skin collagen peptide of claim 1, it is characterized in that: the product index of said fish skin collagen peptide is:
Figure FSB00000636816700011
4. a technology for preparing the fish skin collagen peptide of Magnesium Ascorbylphosphate is characterized in that: comprise the steps: successively
1) takes by weighing the fish-skin raw material, clean, drain away the water, weigh;
2) fish-skin of above-mentioned processing is added the water of 5~6 times of its quality, add the calcium hydroxide of its quality 3.0~5.0% again, stir and soaked 5~7 days, in the NCP in removing fish-skin, also played certain raw meat effect of going;
3) fish-skin of above-mentioned processing is cleaned to neutral, drained away the water, weigh, the gained fish-skin is the raw material after the pre-treatment;
4) with above-mentioned steps 3) fish-skin raw material after the pre-treatment that obtains is according to its quality 5~6 water extraordinarily, and 121 ℃ of high pressure 60~90min are cooled to 50~60 ℃, double gauze elimination residue; Adjust pH is 6.0~8.0,50 ℃ of waters bath with thermostatic control, at first adds the bacillus alkaline protease of fish-skin raw materials quality 0.5~1.0%; Add the bromeline of fish-skin raw materials quality 0.5~1.0% and the neutral protease of fish-skin raw materials quality 0.5~1.0% more simultaneously, pick up counting from adding bacillus alkaline protease, enzymolysis is 5~6 hours altogether; After enzymolysis finishes, more than 95 ℃, 10 minutes enzymes that go out; Cooling, filter cloth filters, and filtrating is the fish skin collagen peptide hydrolysate;
5) above-mentioned filtrating is taken off flavor with the K15 activity carbon column decoloring;
The filtrating of 6) above-mentioned decolouring being taken off behind the flavor is concentrated into suitable volume, and spraying drying makes finished product.
5. by the preparation technology of the described fish skin collagen peptide of claim 4, it is characterized in that: the product index of prepared fish skin collagen peptide is:
Figure FSB00000636816700021
6. the fish skin collagen peptide of the described Magnesium Ascorbylphosphate of claim 1 is as the purposes of makeup or skin-care product additive.
CN2008100480836A 2008-06-19 2008-06-19 Fish skin collagen peptide with whitening effect and preparation process and application thereof Active CN101607982B (en)

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