CN104673866A - Preparation method of whey protein peptide with antibacterial activity - Google Patents
Preparation method of whey protein peptide with antibacterial activity Download PDFInfo
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- CN104673866A CN104673866A CN201510103796.8A CN201510103796A CN104673866A CN 104673866 A CN104673866 A CN 104673866A CN 201510103796 A CN201510103796 A CN 201510103796A CN 104673866 A CN104673866 A CN 104673866A
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Abstract
The invention discloses a preparation method of a whey protein peptide with antibacterial activity, and relates to the technical field of biology. The preparation method comprises steps of preprocessing, enzymolysis, separation and purification, wherein the purification step adopts a simulated mobile chromatography separation method for industrial production of the coarse whey protein peptide after separation, and the whey protein peptide with antibacterial activity is obtained. According to the preparation method, trypsin is adopted to hydrolyze the whey protein to prepare the peptide with antibacterial activity, so that deep processing of the whey protein is realized; the process of the preparation method is simple, the antibacterial activity of the hydrolysis product is high, and the preparation method is suitable for industrial popularization and production; the method is high in automation degree, low in production cost and high in extraction efficiency, and the antibacterial peptide prepared with the method not only has more remarkable bacteriostatic effect, but also is safe and nutritional, provides a reliable base material or ingredient for development of a natural preservative agent, can increase the weight of preservative products of China and improve the safety of the preservative products of China, and meets the requirement of market development of natural food additives.
Description
Technical field
The present invention relates to biological technical field, particularly relate to a kind of making method with the whey protein peptide of anti-microbial activity.
Background technology
Whey is the byproduct of suitability for industrialized production cheese and casein food grade, is the translucent liquid of discharging when curdled milk.Contain in whey-protein and multiple there is different bioactive peptide sequence, under exogenous protease effect, the precursor substance of inactive is released, these little peptide molecules can not only provide growth in humans, grow needed for nutritive substance and energy, what is more important its also have potential antibacterial, reduce cholesterol, hypotensive, anti-oxidant, growth promoting effects, enhancing body immunological competence, calmness, improve the functions such as substance metabolism.Carry out the exploitation with antimicrobial active protein peptide using whey-protein as raw material, the comprehensive utilization and the intensive processing that can be whey provide basic basis, aid in the market development of anticorrosion based food additive and quality is upgraded.
Be applied to the antibacterial peptide of field of food preservation at present mainly based on nisin, Japan, that poly-lysine is used for Food storage as foodstuff additive by Korea S is fresh-keeping.The development & application of antibacterial peptide has obtained the common concern of numerous scholars, adopt the preparation of chemosynthesis and engineered method enforcement antibacterial peptide the most common, but due to the existence of the separating and extracting method small scale complicated operation of food protein derived antimicrobial peptide, the problem such as chemosynthesis high cost is inefficient, gene engineering expression system is uncertain, limit the application of foodstuff additive class antibacterial peptide in food-processing.
Summary of the invention
For above-mentioned the deficiencies in the prior art, provide a kind of making method with the whey protein peptide of anti-microbial activity.
The making method with the whey protein peptide of anti-microbial activity of the present invention, its operation steps is as follows:
A, pre-treatment: mixed with 3 ~ 20 parts of water by 1 part of lactalbumin powder, at 80 ~ 90 DEG C after constant temperature 5min, be cooled to 45 DEG C ~ 60 DEG C, with NaOH solution regulator solution pH to 7.0 ~ 8.5, make lactoalbumin soln;
B, enzymolysis: in the lactoalbumin soln that a step is obtained, add pancreas proteinase-10 .001 ~ 0.005 part, maintain the pH of system by NaOH solution is 7.0 ~ 8.5 simultaneously, stir 2.0 ~ 8.0h under the condition of 45 DEG C ~ 60 DEG C after, then 85 DEG C ~ 100 DEG C are warming up to, constant temperature 10min ~ 15min, obtain whey protein enzymolysis solution;
C, separation: the solid matter in the whey-protein enzymolysis solution that removal b step obtains, collect the filtrate of enzymolysis solution middle-molecular-weihydroxyethyl between 3000Da ~ 5000Da, obtained thick whey protein peptide;
D, purifying: thick whey protein peptide water is modulated into mixed solution, its mass percent concentration is 5% ~ 25%, carry out separation and purification by simulation moving-bed continuous adsorption, then adsorption liquid is carried out spraying dry, the i.e. obtained whey protein peptide with anti-microbial activity;
More than be mass fraction.
As further optimization of the present invention, its operation steps is as follows:
A, pre-treatment: mixed with 3 ~ 10 parts of water by 1 part of lactalbumin powder, at 80 ~ 90 DEG C after constant temperature 5min, be cooled to 45 DEG C ~ 60 DEG C, with NaOH solution regulator solution pH to 7.0 ~ 8.5, make lactoalbumin soln;
B, enzymolysis: in the lactoalbumin soln that a step is obtained, add pancreas proteinase-10 .002 ~ 0.003 part, maintain the pH of system by NaOH solution is 7.0 ~ 8.5 simultaneously, stir 4.0 ~ 6.0h under the condition of 45 DEG C ~ 60 DEG C after, then 85 DEG C ~ 100 DEG C are warming up to, constant temperature 10min ~ 15min, obtain whey protein enzymolysis solution;
C, separation: the solid matter in the whey-protein enzymolysis solution that removal b step obtains, collect the filtrate of enzymolysis solution middle-molecular-weihydroxyethyl between 3000Da ~ 5000Da, obtained thick whey protein peptide;
D, purifying: thick whey protein peptide water is modulated into mixed solution, its mass percent concentration is 5% ~ 25%, carry out separation and purification by simulation moving-bed continuous adsorption, then adsorption liquid is carried out spraying dry, the i.e. obtained whey protein peptide with anti-microbial activity;
More than be mass fraction.
As a further improvement on the present invention, described simulation moving-bed continuous adsorption is separated, and its sorbent material is macroporous adsorbent resin, absorption raw material is that thick whey protein peptide water is modulated into mixed solution, its control flow is 5 ~ 15BV/h; Parsing agent is ethanolic soln, its mass percentage concentration is 70 ~ 90%, its control flow is 15 ~ 25BV/h; Irrigation is deionized water, its control flow is 10 ~ 20BV/h; The regenerator of polymeric adsorbent is NaOH solution, its mass percentage concentration is 2% ~ 4%, its control flow is 5 ~ 10BV/h; Omnidistance switching time is 300 ~ 600s, temperature control 25 ~ 50 DEG C, pressure-controlling is 0.5 ~ 1.0MPa.
As a further improvement on the present invention, described whey-protein is food grade.
As a further improvement on the present invention, described macroporous adsorbent resin is XAD16N polymeric adsorbent, XAD18 polymeric adsorbent or XAD1600N polymeric adsorbent.
The making method with the whey protein peptide of anti-microbial activity of the present invention, be utilize trypsin hydrolyzing whey-protein to make the peptide with anti-microbial activity, achieve the deep processing of whey-protein, its technique is simple, hydrolysate anti-microbial activity is high, is applicable to industrialization promotion and produces; Utilize the whey protein peptide with anti-microbial activity of gained of the present invention, its mass percent concentration is 10 ~ 20%, is detected by high performance liquid chromatography, and its purity is 50 ~ 70%, has obvious restraining effect to intestinal bacteria, streptococcus aureus; Drawn divided by sample size by desorb sample size, its product yield can reach 95%; This method level of automation is high, reduce the wastes and pollution of sorbent material and water, extraction efficiency is high, the content of anti-microbial activity whey protein peptide in enzymolysis product can be made higher simultaneously; The antibacterial peptide obtained by the present invention not only has comparatively significant fungistatic effect but also safe nutrition, exploitation for crude antistaling agent provides reliable base-material or batching, weight and the security of China's sanitas product can be improved, adapt to the requirement of natural additive for foodstuff market development.
Embodiment
Embodiment 1
Have the making method of the whey protein peptide of anti-microbial activity, its operation steps is as follows:
A, pre-treatment: mixed with 3 parts of water by 1 part of food grade lactalbumin powder, at 80 DEG C after constant temperature 5min, be cooled to 45 DEG C, with NaOH solution regulator solution pH to 7.0, makes lactoalbumin soln;
B, enzymolysis: in the lactoalbumin soln that a step is obtained, add pancreas proteinase-10 .001 part, maintain the pH of system by NaOH solution is 7.0 simultaneously, after stirring 2.0h, is then warming up to 85 DEG C under the condition of 45 DEG C, constant temperature 10min, obtain whey protein enzymolysis solution;
C, separation: the solid matter in the whey-protein enzymolysis solution that removal b step obtains, collect the filtrate of enzymolysis solution middle-molecular-weihydroxyethyl between 3000Da ~ 5000Da, obtained thick whey protein peptide;
D, purifying: thick whey protein peptide water is modulated into mixed solution, its mass percent concentration is 5%, carry out separation and purification by simulation moving-bed continuous adsorption, then adsorption liquid is carried out spraying dry, the i.e. obtained whey protein peptide with anti-microbial activity; Described simulation moving-bed continuous adsorption is separated, and its sorbent material is XAD16N polymeric adsorbent, absorption raw material is that thick whey protein peptide water is modulated into mixed solution, its control flow is 5BV/h; Parsing agent is ethanolic soln, its mass percentage concentration is 70%, its control flow is 15BV/h; Irrigation is deionized water, its control flow is 10BV/h; The regenerator of polymeric adsorbent is NaOH solution, its mass percentage concentration is 2%, its control flow is 5BV/h; Omnidistance switching time is 300s, temperature control 25 DEG C, pressure-controlling is 0.5MPa;
More than be mass fraction.
Embodiment 2
Have the making method of the whey protein peptide of anti-microbial activity, its operation steps is as follows:
A, pre-treatment: mixed with 20 parts of water by 1 part of food grade lactalbumin powder, at 90 DEG C after constant temperature 5min, be cooled to 60 DEG C, with NaOH solution regulator solution pH to 8.5, makes lactoalbumin soln;
B, enzymolysis: in the lactoalbumin soln that a step is obtained, add pancreas proteinase-10 .005 part, maintain the pH of system by NaOH solution is 8.5 simultaneously, after stirring 8.0h, is then warming up to 100 DEG C under the condition of 60 DEG C, constant temperature 15min, obtain whey protein enzymolysis solution;
C, separation: the solid matter in the whey-protein enzymolysis solution that removal b step obtains, collect the filtrate of enzymolysis solution middle-molecular-weihydroxyethyl between 3000Da ~ 5000Da, obtained thick whey protein peptide;
D, purifying: thick whey protein peptide water is modulated into mixed solution, its mass percent concentration is 25%, separation and purification is carried out by simulation moving-bed continuous adsorption, then adsorption liquid is carried out spraying dry, the i.e. obtained whey protein peptide with anti-microbial activity, described simulation moving-bed continuous adsorption is separated, and its sorbent material is XAD18 polymeric adsorbent, absorption raw material is that thick whey protein peptide water is modulated into mixed solution, its control flow is 15BV/h; Parsing agent is ethanolic soln, its mass percentage concentration is 90%, its control flow is 25BV/h; Irrigation is deionized water, its control flow is 20BV/h; The regenerator of polymeric adsorbent is NaOH solution, its mass percentage concentration is 4%, its control flow is 10BV/h; Omnidistance switching time is 600s, temperature control 50 DEG C, pressure-controlling is 1.0MPa;
More than be mass fraction.
Embodiment 3
Have the making method of the whey protein peptide of anti-microbial activity, its operation steps is as follows:
A, pre-treatment: mixed with 3 parts of water by 1 part of food grade lactalbumin powder, at 80 DEG C after constant temperature 5min, be cooled to 45 DEG C, with NaOH solution regulator solution pH to 7.0, makes lactoalbumin soln;
B, enzymolysis: in the lactoalbumin soln that a step is obtained, add pancreas proteinase-10 .002 part, maintain the pH of system by NaOH solution is 7.0 simultaneously, after stirring 4.0h, is then warming up to 85 DEG C under the condition of 45 DEG C, constant temperature 10min, obtain whey protein enzymolysis solution;
C, separation: the solid matter in the whey-protein enzymolysis solution that removal b step obtains, collect the filtrate of enzymolysis solution middle-molecular-weihydroxyethyl between 3000Da ~ 5000Da, obtained thick whey protein peptide;
D, purifying: thick whey protein peptide water is modulated into mixed solution, its mass percent concentration is 5%, separation and purification is carried out by simulation moving-bed continuous adsorption, then adsorption liquid is carried out spraying dry, the i.e. obtained whey protein peptide with anti-microbial activity, described simulation moving-bed continuous adsorption is separated, and its sorbent material is XAD1600N polymeric adsorbent, absorption raw material is that thick whey protein peptide water is modulated into mixed solution, its control flow is 5BV/h; Parsing agent is ethanolic soln, its mass percentage concentration is 70%, its control flow is 15BV/h; Irrigation is deionized water, its control flow is 10BV/h; The regenerator of polymeric adsorbent is NaOH solution, its mass percentage concentration is 2%, its control flow is 5BV/h; Omnidistance switching time is 300s, temperature control 25 DEG C, pressure-controlling is 0.5MPa;
More than be mass fraction.
Embodiment 4
Have the making method of the whey protein peptide of anti-microbial activity, its operation steps is as follows:
A, pre-treatment: mixed with 10 parts of water by 1 part of food grade lactalbumin powder, at 90 DEG C after constant temperature 5min, be cooled to 60 DEG C, with NaOH solution regulator solution pH to 7.5, makes lactoalbumin soln;
B, enzymolysis: in the lactoalbumin soln that a step is obtained, add pancreas proteinase-10 .003 part, maintain the pH of system by NaOH solution is 7.5 simultaneously, after stirring 6.0h, is then warming up to 100 DEG C under the condition of 60 DEG C, constant temperature 15min, obtain whey protein enzymolysis solution;
C, separation: the solid matter in the whey-protein enzymolysis solution that removal b step obtains, collect the filtrate of enzymolysis solution middle-molecular-weihydroxyethyl between 3000Da ~ 5000Da, obtained thick whey protein peptide;
D, purifying: thick whey protein peptide water is modulated into mixed solution, its mass percent concentration is 25%, separation and purification is carried out by simulation moving-bed continuous adsorption, then adsorption liquid is carried out spraying dry, the i.e. obtained whey protein peptide with anti-microbial activity, described simulation moving-bed continuous adsorption is separated, and its sorbent material is XAD16N polymeric adsorbent, absorption raw material is that thick whey protein peptide water is modulated into mixed solution, its control flow is 15BV/h; Parsing agent is ethanolic soln, its mass percentage concentration is 90%, its control flow is 25BV/h; Irrigation is deionized water, its control flow is 20BV/h; The regenerator of polymeric adsorbent is NaOH solution, its mass percentage concentration is 4%, its control flow is 10BV/h; Omnidistance switching time is 600s, temperature control 50 DEG C, pressure-controlling is 1.0MPa;
More than be mass fraction.
Embodiment 5
Have the making method of the whey protein peptide of anti-microbial activity, its operation steps is as follows:
A, pre-treatment: mixed with 8 parts of water by 1 part of food grade lactalbumin powder, at 85 DEG C after constant temperature 5min, be cooled to 55 DEG C, with NaOH solution regulator solution pH to 8.0, makes lactoalbumin soln;
B, enzymolysis: in the lactoalbumin soln that a step is obtained, add pancreas proteinase-10 .003 part, maintain the pH of system by NaOH solution is 8.0 simultaneously, after stirring 5.0h, is then warming up to 95 DEG C under the condition of 55 DEG C, constant temperature 15min, obtain whey protein enzymolysis solution;
C, separation: the solid matter in the whey-protein enzymolysis solution that removal b step obtains, collect the filtrate of enzymolysis solution middle-molecular-weihydroxyethyl between 3000Da ~ 5000Da, obtained thick whey protein peptide;
D, purifying: thick whey protein peptide water is modulated into mixed solution, its mass percent concentration is 20%, carry out separation and purification by simulation moving-bed continuous adsorption, then adsorption liquid is carried out spraying dry, the i.e. obtained whey protein peptide with anti-microbial activity;
More than be mass fraction.
Described simulation moving-bed continuous adsorption is separated, and its sorbent material is XAD1600N polymeric adsorbent, absorption raw material is that thick whey protein peptide water is modulated into mixed solution, its control flow is 10BV/h; Parsing agent is ethanolic soln, its mass percentage concentration is 90%, its control flow is 20BV/h; Irrigation is deionized water, its control flow is 20BV/h; The regenerator of polymeric adsorbent is NaOH solution, its mass percentage concentration is 4%, its control flow is 10BV/h; Omnidistance switching time is 600s, temperature control 50 DEG C, pressure-controlling is 0.8MPa.
Food grade whey-protein used in above embodiment is that Australian Mai Gao company produces, and its protein content is 82%; Trypsinase is a kind of endopeptidase being extracted by ox, sheep, Pancreas Sus domestica and obtained, the peptide chain that can selectively be made up of Methionin or arginic carboxyl in protein hydrolysate, only fracture Methionin or arginic carboxyl participate in the peptide bond of formation, trypsinase used in the application is produced by Ya Xin bio tech ltd, Shanghai, and its enzyme activity is 2,000,000 U/g; The XAD16N polymeric adsorbent used, XAD18 polymeric adsorbent or XAD1600N polymeric adsorbent, for Beijing Sai Fulaibo Science and Technology Ltd. produces.
Claims (5)
1. have the making method of the whey protein peptide of anti-microbial activity, its operation steps is as follows:
A, pre-treatment: mixed with 3 ~ 20 parts of water by 1 part of lactalbumin powder, at 80 ~ 90 DEG C after constant temperature 5min, be cooled to 45 DEG C ~ 60 DEG C, with NaOH solution regulator solution pH to 7.0 ~ 8.5, make lactoalbumin soln;
B, enzymolysis: in the lactoalbumin soln that a step is obtained, add pancreas proteinase-10 .001 ~ 0.005 part, maintain the pH of system by NaOH solution is 7.0 ~ 8.5 simultaneously, stir 2.0 ~ 8.0h under the condition of 45 DEG C ~ 60 DEG C after, then 85 DEG C ~ 100 DEG C are warming up to, constant temperature 10min ~ 15min, obtain whey protein enzymolysis solution;
C, separation: the solid matter in the whey-protein enzymolysis solution that removal b step obtains, collect the filtrate of enzymolysis solution middle-molecular-weihydroxyethyl between 3000Da ~ 5000Da, obtained thick whey protein peptide;
D, purifying: thick whey protein peptide water is modulated into mixed solution, its mass percent concentration is 5% ~ 25%, carry out separation and purification by simulation moving-bed continuous adsorption, then adsorption liquid is carried out spraying dry, the i.e. obtained whey protein peptide with anti-microbial activity;
More than be mass fraction.
2. have the making method of the whey protein peptide of anti-microbial activity as claimed in claim 1, as further optimization, its operation steps is as follows:
A, pre-treatment: mixed with 3 ~ 10 parts of water by 1 part of lactalbumin powder, at 80 ~ 90 DEG C after constant temperature 5min, be cooled to 45 DEG C ~ 60 DEG C, with NaOH solution regulator solution pH to 7.0 ~ 8.5, make lactoalbumin soln;
B, enzymolysis: in the lactoalbumin soln that a step is obtained, add pancreas proteinase-10 .002 ~ 0.003 part, maintain the pH of system by NaOH solution is 7.0 ~ 8.5 simultaneously, stir 4.0 ~ 6.0h under the condition of 45 DEG C ~ 60 DEG C after, then 85 DEG C ~ 100 DEG C are warming up to, constant temperature 10min ~ 15min, obtain whey protein enzymolysis solution;
C, separation: the solid matter in the whey-protein enzymolysis solution that removal b step obtains, collect the filtrate of enzymolysis solution middle-molecular-weihydroxyethyl between 3000Da ~ 5000Da, obtained thick whey protein peptide;
D, purifying: thick whey protein peptide water is modulated into mixed solution, its mass percent concentration is 5% ~ 25%, carry out separation and purification by simulation moving-bed continuous adsorption, then adsorption liquid is carried out spraying dry, the i.e. obtained whey protein peptide with anti-microbial activity;
More than be mass fraction.
3. there is the making method of the whey protein peptide of anti-microbial activity as claimed in claim 1, it is characterized in that described simulation moving-bed continuous adsorption is separated, its sorbent material is macroporous adsorbent resin, absorption raw material is that thick whey protein peptide water is modulated into mixed solution, to control flow be 5 ~ 15BV/h for it; Parsing agent is ethanolic soln, its mass percentage concentration is 70 ~ 90%, its control flow is 15 ~ 25BV/h; Irrigation is deionized water, its control flow is 10 ~ 20BV/h; The regenerator of polymeric adsorbent is NaOH solution, its mass percentage concentration is 2% ~ 4%, its control flow is 5 ~ 10BV/h; Omnidistance switching time is 300 ~ 600s, temperature control 25 ~ 50 DEG C, pressure-controlling is 0.5 ~ 1.0MPa.
4. there is the making method of the whey protein peptide of anti-microbial activity as claimed in claim 1, it is characterized in that described whey-protein is food grade.
5. there is the making method of the whey protein peptide of anti-microbial activity as claimed in claim 1, it is characterized in that described macroporous adsorbent resin is XAD16N polymeric adsorbent, XAD18 polymeric adsorbent or XAD1600N polymeric adsorbent.
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CN106148469A (en) * | 2016-09-19 | 2016-11-23 | 无锡昌穗健康科技有限公司 | A kind of preparation method and applications of crow rice gluten peptide |
CN111227025A (en) * | 2020-01-20 | 2020-06-05 | 安徽鲜森绿色食品有限公司 | Poultry cold chain transportation fresh-keeping method |
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CN101928742A (en) * | 2009-06-23 | 2010-12-29 | 光明乳业股份有限公司 | Whey protein active peptide with antioxidant activity and preparation method thereof |
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Publication number | Priority date | Publication date | Assignee | Title |
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CN106148469A (en) * | 2016-09-19 | 2016-11-23 | 无锡昌穗健康科技有限公司 | A kind of preparation method and applications of crow rice gluten peptide |
CN111227025A (en) * | 2020-01-20 | 2020-06-05 | 安徽鲜森绿色食品有限公司 | Poultry cold chain transportation fresh-keeping method |
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