Abalone peptide for assisting lactation and preparation method thereof

Abstract

The invention discloses an abalone peptide for assisting lactation and a preparation method thereof, and relates to the technical field of biological peptides, and the preparation method comprises the following steps: step 1, taking fresh abalone meat, and sequentially carrying out cleaning, acid-base soaking for deashing and pulping treatment to obtain abalone pulp; step 2, boiling the abalone ground pulp at constant temperature and constant pressure, cooling to 55 DEG C, and performing enzymolysis by multiple compound enzymes to obtain enzymatic hydrolysate; step 3, heating the enzymatic hydrolysate to 85 DEG C and carrying out enzyme deactivation treatment for 5 minutes to obtain a protein peptide mixed solution; and 4, separating the protein peptide, and drying to obtain the abalone peptide particles. The corresponding abalone peptide for assisting lactation is obtained, and the abalone peptide has the effect of effectively promoting mild filling of mammary gland tissue glandular vesicles, lumen expansion and active milk secretion, and further has the effect of being applied to assisting lactation.

Description

Technical Field

[0001] The present application relates to the field of biological peptide technology, and in particular to an abalone peptide used for assisting lactation and a preparation method thereof. Background Art

[0002] Abalone is a common shellfish in the ocean. It is extremely nutritious and has been regarded as the "crown of seafood delicacies" since ancient times. Abalone peptide is a functional fragment of abalone protein and a type of biologically active substance involved in various cell functions in organisms. It has a simpler structure and smaller molecular weight than protein, making it easier for the human body to digest and absorb. At the same time, peptides can regulate, adjust and control various functions of the human body. Abalone peptide has the effect of balancing the human body's immunity, thereby ensuring the intake and supplementation of various energies in the body. It has higher biological efficacy and more important physiological functions than ordinary abalone products.

[0003] The Chinese patent with the announcement number CN107556364B discloses a method and product for extracting abalone protein peptides by subcritical water-assisted enzymolysis, which comprises the following steps: (1) subcritical water treatment: adding purified water to abalone viscera at a material-liquid ratio of 1:3 to 1:5 (w / v), extracting for 15 min to 60 min under subcritical water conditions, cooling to room temperature, removing the filter residue with a stainless steel filter to obtain a protein extract, and the filter residue can be used as seaweed fertilizer after being dried in the sun; (2) enzymolysis: adding a complex enzyme to the protein extract obtained in step (1), wherein the amount of the complex enzyme added is 0.1% of the protein extract. ~0.5% (w / v), enzymolysis reaction 1h~3h; (3) separation, concentration and spray drying: the protein hydrolyzate obtained in step (2) is purified by using activated carbon and resin for decolorization, deodorization, removal of heavy metals, and removal of arsenic, and then concentrated by nanofiltration membrane, vacuum concentration and spray drying to prepare abalone protein peptide powder; (4) product preparation: β-cyclodextrin, collagen peptide, soy lecithin, green olive pulp extract and honey are added to the extracted abalone protein peptide powder, mixed evenly, and made into product particles, and then 1%~5% of the mass of the product particles of magnesium stearate are added, mixed evenly, and pressed into chewable tablets.

[0004] However, the product obtained based on this method only has good antioxidant and anti-cancer functions and cannot be used to assist lactation, and needs to be improved. Summary of the invention

[0005] In view of this, the first purpose of the present application is to provide a method for preparing abalone peptides for assisting lactation, so as to achieve the purpose of assisting lactation. The specific scheme is as follows:

[0006] A method for preparing abalone peptide for assisting lactation comprises the following steps:

[0007] Step 1, taking fresh abalone meat, washing, acid and alkali soaking deashing and grinding treatment in sequence, to obtain abalone grinding liquid;

[0008] Step 2, boiling the abalone grinding liquid at a constant temperature and pressure, cooling it to 55° C., and performing enzymolysis with a multiple complex enzyme to obtain an enzymolysis liquid;

[0009] Step 3, heating the enzymatic hydrolyzate to 85°C and performing enzyme inactivation treatment for 5 minutes to obtain a protein peptide mixture;

[0010] Step 4: Separate the protein peptides and obtain abalone peptide particles by drying.

[0011] Preferably: the cleaning is performed by washing with clean water, the acid-base deashing is performed by an activated carbon acid-base deashing process, and the refining is performed by grinding with a grinder.

[0012] Preferably: the multiple complex enzyme includes papain, lipase and at least one of pepsin, carboxypeptidase, pancreatin and alkaline protease; and the mass ratio of the papain is greater than 22wt%, and the mass ratio of the lipase is greater than 12wt%.

[0013] Preferably: the separation of proteases comprises separating protein peptides using a high-speed bedroom centrifuge.

[0014] Preferably, the separation of protease also includes using a ceramic primary filtration membrane to coarsely filter visible solids and using an ultrafiltration membrane to separate macromolecular protein peptides.

[0015] Preferably, the separation of protease and drying also include fermentation to remove fishy smell, heavy metal removal, desalination, small molecule peptide separation, decolorization and concentration treatment in sequence.

[0016] Preferably: the fermentation to remove fishy smell is carried out by adopting a flavor fermentation process; the heavy metal removal and desalination are carried out by adopting an electrodialysis device to separate heavy metals and salts based on the electric field force or by adopting an ion exchange resin to remove heavy metals; the small molecule peptide separation is carried out by adopting an inorganic nanofiltration membrane to separate small molecule peptides below 1000DA; the decolorization is carried out by physical decolorization using a nanofiltration membrane; the concentration is carried out by adopting a double-effect tank concentration, and the drying is carried out by adopting a centrifugal pressure spray drying tower to carry out low-temperature spray drying.

[0017] The second object of the present invention is to provide an abalone peptide for assisting lactation, which is prepared by the above-mentioned preparation method of the abalone peptide for assisting lactation and is used for assisting lactation.

[0018] It can be seen from the above scheme that the present application provides an abalone peptide for assisting lactation and a preparation method thereof. The preparation method of the abalone peptide for assisting lactation obtains the corresponding abalone peptide for assisting lactation, and has the effect of effectively promoting mild filling of the alveoli of the mammary tissue, expansion of the ductus, and active milk secretion, thereby having the effect of assisting lactation. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] In order to more clearly illustrate the embodiments of the present application or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present application. For ordinary technicians in this field, other drawings can be obtained based on the provided drawings without paying any creative work.

[0020] Figure 1 This is a schematic diagram of the effect of abalone peptide disclosed in the present application on the lactation of pregnant mice;

[0021] Figure 2 This is a schematic diagram of the mammary tissue section results of each group of female mice disclosed in this application. DETAILED DESCRIPTION

[0022] The following will be combined with the drawings in the embodiments of the present application to clearly and completely describe the technical solutions in the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, not all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of this application.

[0023] It should be mentioned that the experimental animals used in the examples of the present application are SPF female KM mice, provided by Sibeifu (Suzhou) Biotechnology Co., Ltd. (license number is SCXK (Beijing) 2022-0006). The abalone peptide obtained for assisting lactation is a white powder and is stored in a cool, dark and dry place.

[0024] In order to evaluate the functional application of abalone peptides for assisting lactation, healthy pregnant mice were used as subjects to measure the milk production of maternal mice, the average weight gain of pups, and the biochemical indicators in the serum of maternal mice, and the pathological diagnosis of the maternal mouse mammary tissue was performed.

[0025] The following is a detailed description of an abalone peptide for assisting lactation and a preparation method thereof according to the present application.

[0026] A method for preparing abalone peptide for assisting lactation comprises the following steps:

[0027] Step 1, taking fresh abalone meat, washing, acid and alkali soaking deashing and grinding treatment in sequence, to obtain abalone grinding liquid;

[0028] Step 2, boiling the abalone grinding liquid at a constant temperature and pressure, cooling it to 55° C., and performing enzymolysis with a multiple complex enzyme to obtain an enzymolysis liquid;

[0029] Step 3, heating the enzymatic hydrolyzate to 85°C and performing enzyme inactivation treatment for 5 minutes to obtain a protein peptide mixture;

[0030] Step 4: Separate the protein peptides and obtain abalone peptide particles by drying.

[0031] Among them, the cleaning is done by washing with clean water, the acid-base soaking and deashing is done by using an activated carbon acid-base deashing process, and the refining is done by grinding with a grinder. The above process steps are all conventional processes and will not be described in detail here.

[0032] It should be noted that the multiple complex enzyme includes at least one of papain, lipase and pepsin, carboxypeptidase, pancreatin and alkaline protease. The mass ratio of papain is greater than 22wt%, and the mass ratio of lipase is greater than 12wt%. The multiple complex enzyme used in the embodiment of the present application is preferably papain, lipase, pepsin, pancreatin and alkaline protease in a mass ratio of 28:17:15:20:20.

[0033] At the same time, the separation of protease includes the use of a high-speed bedroom centrifuge to separate protein peptides. Of course, in order to further improve the purity of the separation, it also includes the use of a ceramic primary filtration membrane to coarsely filter visible solids and an ultrafiltration membrane to separate large molecular protein peptides. And by using fermentation to remove fishy smell, remove heavy metals, desalt, separate small molecule peptides, decolorize and concentrate in sequence between the separation of protease and drying, a high-purity abalone peptide used to assist lactation is obtained.

[0034] Among them, fermentation to remove fishy smell adopts flavor fermentation process. Heavy metal removal and desalination adopt electrodialysis equipment to separate heavy metals and salts based on electric field force or use ion exchange resin to remove heavy metals. Small molecule peptide separation adopts inorganic nanofiltration membrane to separate small molecule peptides below 1000DA. Decolorization adopts nanofiltration membrane for physical decolorization. Concentration adopts double-effect tank concentration, and drying adopts centrifugal pressure spray drying tower for low-temperature spray drying.

[0035] An abalone peptide for assisting lactation is prepared by the above-mentioned preparation method of the abalone peptide for assisting lactation, and is used for assisting lactation.

[0036] Functional application testing:

[0037] Rearing conditions: Experimental animals were reared in the barrier facility SYXK(Su)2021-0056 of the Experimental Animal Center of Jiangnan University at a temperature of 20-26°C and a relative humidity of 40-70%. Irradiated sterilized feed and bedding were provided by Jiangsu Collaborative Pharmaceutical Bioengineering Co., Ltd., Su Feed Certificate (2019)01008.

[0038] Experimental Animals-Female mice were randomly divided into 4 groups according to body weight: a blank control group and 3 sample groups with low, medium and high doses.

[0039] Testing items: milk production, average weight gain of offspring, serum biochemical index testing, and breast tissue pathology diagnosis.

[0040] Serum biochemical indicators - detection of serum prolactin (RPL), estradiol (E2), progesterone (P) and growth hormone (GH) levels.

[0041] The specific experimental steps are as follows:

[0042] (1) From the second day after delivery, except for the blank control group mice which were given distilled water, each dose group was given a corresponding amount of abalone peptide aqueous solution (low dose 0.5g / kg BW, medium dose 1.0g / kg BW, high dose 2.0g / kg BW), and the gavage volume was calculated according to 10mL / kg·BW, once a day for 14 days. The weights of the mother mice and the offspring were recorded before and after lactation. After 14 days, the eyeballs of the mother mice were removed to collect blood, and the serum was centrifuged and stored at -80℃ for testing. At the same time, the third pair of mammary glands were dissected and fixed for staining and microscopic examination;

[0043] (2) Determination of milk production: Starting from the second day after the mother mouse gave birth, the pups were separated from the mother mouse at 08:00 every day. After the pups were starved for 4 hours, they were weighed before lactation at 12:00. The pups were then returned to the mouse nest to lactate for 1 hour. The pups were then separated from the mother mouse again and weighed after lactation for 1 hour. The difference in body weight before and after lactation for 1 hour was the milk production for that hour.

[0044] (3) Determination of average weight gain of offspring: Record the initial weight of offspring and the weight 14 days after gavage, and calculate the difference in body weight before and after gavage. Average weight gain of offspring = difference in body weight before and after gavage / 14;

[0045] (4) Determination of serum biochemical indicators: After the serum was thawed overnight in a 4°C refrigerator, samples were taken evenly to determine the levels of prolactin (RPL), estradiol (E2), progesterone (P), and growth hormone (GH) in the serum. The determination method was carried out according to the requirements of the kit;

[0046] (5) Pathological diagnosis of mammary gland tissue: After the female mice were sacrificed, the third pair of mammary glands were cut out and fixed with 4% paraformaldehyde. After routine dehydration, paraffin embedding, sectioning and HE staining, the expansion of the alveoli and ducts of the mammary gland tissue was observed under a microscope.

[0047] The test results are shown in Tables 1 and 2 below.

[0048] Table 1 Effects of the test substances on the average body weight gain of offspring mice

[0049]

[0050] It can be seen from Table 1 above that, compared with the blank group, there was no significant difference in the average body weight gain of the offspring mice in each dose group (low, medium and high) of the test substance (P>0.05), indicating that the test sample intervention for 14 days could not increase the average body weight gain of the offspring mice, and the result of this indicator was negative.

[0051] Table 2 Effects of the test substances on serum lactation-related hormone levels in mice

[0052]

[0053] From Table 2 above, we can know that the content of prolactin (RPL) and progesterone (P) in the medium and high dose groups of the test substance was significantly higher than that in the blank group (P<0.05). In addition, compared with the blank group, there was no significant difference in estradiol (E2) and growth hormone (GH) in each test group. This shows that abalone peptide has the effect of increasing prolactin (RPL) and progesterone (P), and the results of these two indicators are positive, while the results of estradiol (E2) and growth hormone (GH) are negative.

[0054] like Figure 1 As shown, compared with the blank group, in the first 4 days of lactation, there was no significant difference in the milk secretion of pregnant mice in the test group. As the intervention time prolonged, the milk secretion of pregnant mice in the test group increased significantly (P<0.05). Among them, the high dose had no significant difference only on the 13th day (P>0.05), the medium dose group had no significant difference only on the 12th and 13th days (P>0.05), and the low dose group had significant differences only on the 8th and 14th days (P<0.05). This shows that the abalone peptide can increase the milk secretion of pregnant mice, and the result of this indicator is positive.

[0055] The structure of the mammary gland tissue of each group of rats was observed 14 days after intragastric administration. Figure 2As shown. The mammary gland showed typical secretory mammary gland performance, with well-developed glands, large alveolar cavities, thick duct cavities, pink milk visible in some glandular cavities, and a small amount of connective tissue distribution in the interstitium. Compared with the blank control group, there was no significant difference between the 4 cases in the low-dose group and the blank group. In 4 cases, the alveoli were slightly to slightly filled, the ducts were dilated, and pink milk was seen in the cavities. There was no significant difference between the 4 cases in the medium-dose group and the blank group. In 4 cases, the alveoli were slightly to slightly filled, the ducts were dilated, and pink milk was seen in the cavities. There was no significant difference between the 2 cases in the high-dose group and the blank group. In 6 cases, the alveoli were slightly to slightly filled, the ducts were dilated, and pink milk was seen in the cavities. It can be seen that abalone peptide can make the milk secretion of maternal mice more active, and the higher the dose, the higher the incidence and the more significant the effect.

[0056] According to the above results, the research results of abalone peptides used to assist lactation prepared in this application show that: compared with the blank control group, the milk secretion of the test group increased significantly (P<0.05), and the prolactin (RPL) and progesterone (P) in the serum increased significantly (P<0.05).

[0057] In summary, the present application provides an abalone peptide for assisting lactation and a preparation method thereof. The preparation method of the abalone peptide for assisting lactation obtains the corresponding abalone peptide for assisting lactation, and has the effect of effectively promoting mild filling of the alveoli of the mammary tissue, expansion of the ductus, and active milk secretion, thereby having the effect of assisting lactation.

[0058] The "first", "second", "third", "fourth", etc. (if any) referred to in this application are used to distinguish similar objects, and are not necessarily used to describe a specific order or sequence. It should be understood that the data used in this way can be interchangeable where appropriate, so that the embodiments described herein can be implemented in an order other than that illustrated or described herein. In addition, the terms "including" and "having" and any variations thereof are intended to cover non-exclusive inclusions, for example, a process, method, or device that includes a series of steps or units is not necessarily limited to those steps or units that are clearly listed, but may include other steps or units that are not clearly listed or inherent to these processes, methods, or devices.

[0059] It should be noted that the descriptions involving "first", "second", etc. in this application are only for descriptive purposes and cannot be understood as indicating or implying their relative importance or implicitly indicating the number of technical features indicated. Therefore, the features defined as "first" and "second" may explicitly or implicitly include at least one of the features. In addition, the technical solutions between the various embodiments can be combined with each other, but they must be based on the ability of ordinary technicians in this field to implement them. When the combination of technical solutions is contradictory or cannot be implemented, it should be deemed that such combination of technical solutions does not exist and is not within the scope of protection required by this application.

[0060] Specific examples are used herein to illustrate the principles and implementation methods of the present application. The description of the above embodiments is only used to help understand the method of the present application and its core idea. At the same time, for those skilled in the art, according to the idea of ​​the present application, there will be changes in the specific implementation methods and application scope. In summary, the content of this specification should not be understood as a limitation on the present application.

Claims

1. A method for preparing an abalone peptide for assisting lactation, characterized in that: The steps include: Step 1, taking fresh abalone meat, washing, acid and alkali soaking deashing and grinding treatment in sequence, to obtain abalone grinding liquid; Step 2, boiling the abalone grinding liquid at a constant temperature and pressure, cooling it to 55° C., and performing enzymolysis with a multiple complex enzyme to obtain an enzymolysis liquid; Step 3, heating the enzymatic hydrolyzate to 85°C and performing enzyme inactivation treatment for 5 minutes to obtain a protein peptide mixture; Step 4: Separate the protein peptides and obtain abalone peptide particles by drying.

2. A method for preparing an abalone peptide for assisting lactation according to claim 1, characterized in that: The cleaning is performed by washing with clean water, the acid-base deashing is performed by using an activated carbon acid-base deashing process, and the refining is performed by using a grinder.

3. A method for preparing an abalone peptide for assisting lactation according to claim 1, characterized in that: The multiple complex enzymes include papain, lipase and at least one of pepsin, carboxypeptidase, pancreatin and alkaline protease; and the mass ratio of the papain is greater than 22wt%, and the mass ratio of the lipase is greater than 12wt%.

4. A method for preparing an abalone peptide for assisting lactation according to claim 1, characterized in that: The separation of protease comprises separating protein peptides by using a high-speed bedroom centrifuge.

5. A method for preparing an abalone peptide for assisting lactation according to claim 4, characterized in that: The separation of protease also includes using a ceramic primary filtration membrane to coarsely filter visible solids and using an ultrafiltration membrane to separate macromolecular protein peptides.

6. A method for preparing an abalone peptide for assisting lactation according to claim 1, characterized in that: The separation of protease and drying also include fermentation to remove fishy smell, heavy metal removal, desalination, small molecule peptide separation, decolorization and concentration treatment in sequence.

7. A method for preparing an abalone peptide for assisting lactation according to claim 6, characterized in that: The fermentation to remove fishy smell adopts a flavor fermentation process; the heavy metal removal and desalination adopt electrodialysis equipment to separate heavy metals and salts based on the electric field force or use ion exchange resin to remove heavy metals; the small molecule peptide separation adopts inorganic nanofiltration membrane to separate small molecule peptides below 1000DA; the decolorization adopts nanofiltration membrane for physical decolorization; the concentration adopts double-effect tank concentration, and the drying adopts centrifugal pressure spray drying tower for low-temperature spray drying.

8. An abalone peptide for assisting lactation, characterized in that: The abalone peptide for assisting lactation is prepared by the preparation method for the abalone peptide for assisting lactation as described in any one of claims 1 to 7, and is used for assisting lactation.

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