Preparation method of donkey-hide gelatin peptide

Abstract

The invention relates to the technical field of colla corii asini peptide preparation, in particular to a preparation method of colla corii asini peptide. The preparation method comprises the following steps: step 1, unhairing and degreasing fresh donkey hide, dicing, blanching and cleaning to obtain blocky donkey hide; step 2, adding the blocky donkey hide into water for ultrasonic treatment, then cooking, filtering and collecting filtrate 1; and step 3, adding a compound enzyme into the filtrate 1, carrying out ultrasonic treatment, sequentially carrying out enzymolysis, enzyme deactivation and centrifugal filtration, carrying out debitterizing, decolorizing and degreasing treatment on the obtained filtrate 2, carrying out ultrafiltration, and carrying out spray drying to obtain the colla corii asini peptide, the compound enzyme is a mixture of alkaline protease, neutral protease, pepsase and papain in a mass ratio of 3: 1: 2: 1. According to the method, the donkey-hide gelatin peptide is prepared from the fresh donkey hide, the production cost is reduced, and the donkey-hide gelatin peptide with the molecular weight smaller than 1000 Daltons in the donkey-hide gelatin peptide prepared through the method is high in proportion and can be easily absorbed and utilized by the human body.

Description

Technical Field

[0001] The invention relates to the technical field of donkey-hide gelatin preparation, and in particular to a method for preparing donkey-hide gelatin peptide. Background Art

[0002] Donkey hide gelatin is one of the famous traditional Chinese medicines and nutritional supplements. It is a gelatin block made from donkey skin of equine animals through decoction and concentration. It contains animal glue, protein, various amino acids and trace elements such as iron and calcium. Modern pharmacological research has confirmed that donkey hide gelatin has many clinical functions such as blood tonic and hemostasis, anti-aging, and anti-fatigue. At present, the main way to eat donkey hide gelatin is to make it into donkey hide gelatin blocks and eat it directly; however, modern research shows that donkey hide gelatin blocks with too large relative molecular weight are difficult to be digested and absorbed by the body, and the bioavailability is extremely low. For people with poor digestive function, simply taking donkey hide gelatin orally will increase the burden on the stomach and intestines.

[0003] In the prior art, there are products that process donkey-hide gelatin into donkey-hide gelatin small molecule peptides for people to eat, but they are generally made from finished donkey-hide gelatin blocks as raw materials, which greatly increases the production cost and is not suitable for large-scale production. In addition, studies have shown that donkey-hide gelatin peptides with a molecular weight of less than 1000 Daltons can more easily pass through the cell membrane and can be directly absorbed by the human body without going through a complicated digestion process, and the absorption rate is significantly improved. Therefore, a method for preparing donkey-hide gelatin peptides using fresh donkey skin as raw material is provided, which reduces the production cost and increases the percentage of donkey-hide gelatin peptides with a molecular weight of less than 1000 Daltons, which is of great significance to the technical field of donkey-hide gelatin peptide preparation. Summary of the invention

[0004] Based on the above content, the present invention provides a method for preparing donkey-hide gelatin peptide.

[0005] To achieve the above object, the present invention provides the following solutions:

[0006] One of the technical solutions of the present invention is a method for preparing donkey-hide gelatin peptide, comprising the following steps:

[0007] Step 1, after removing hair and fat from fresh donkey skin, cutting into pieces, blanching and washing, to obtain block donkey skin;

[0008] Step 2, adding the block of donkey skin into water for ultrasonic treatment, then steaming, and filtering to collect filtrate 1;

[0009] Step 3, adding a composite enzyme to the filtrate 1 for ultrasonic treatment, followed by enzymatic hydrolysis, enzyme inactivation, and centrifugal filtration, and the obtained filtrate 2 is subjected to debittering, decolorizing, and defatting treatment, followed by ultrafiltration, and then spray drying to obtain donkey-hide gelatin peptide;

[0010] The complex enzyme is a mixture of alkaline protease, neutral protease, pepsin and papain in a mass ratio of 3:1:2:1.

[0011] The second technical solution of the present invention is a donkey-hide gelatin peptide prepared according to the above-mentioned preparation method.

[0012] The third technical solution of the present invention is the use of the above-mentioned donkey-hide gelatin peptide in the preparation of iron or blood tonic drugs.

[0013] The present invention discloses the following technical effects:

[0014] The present invention uses fresh donkey hide as a raw material to prepare donkey-hide gelatin peptides, and does not need to use finished donkey-hide gelatin blocks, thereby reducing production costs. At the same time, the proportion of donkey-hide gelatin peptides with a molecular weight of less than 1000 Daltons in the donkey-hide gelatin peptides prepared by the method of the present invention is high, which is conducive to absorption and utilization by the human body. DETAILED DESCRIPTION

[0015] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0016] It should be understood that the terms described in the present invention are only for describing a particular embodiment and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. The intermediate value in any stated value or stated range, and each smaller range between any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0017] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0018] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to the skilled artisan. The present invention description and examples are exemplary only.

[0019] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0020] The first aspect of the present invention provides a method for preparing donkey-hide gelatin peptide, comprising the following steps:

[0021] Step 1, after removing hair and fat from fresh donkey skin, cutting into pieces, blanching and washing, to obtain block donkey skin;

[0022] Step 2, adding the block of donkey skin into water for ultrasonic treatment, then steaming, and filtering to collect filtrate 1;

[0023] Step 3, adding a composite enzyme to the filtrate 1 for ultrasonic treatment, followed by enzymatic hydrolysis, enzyme inactivation, and centrifugal filtration, and the obtained filtrate 2 is subjected to debittering, decolorizing, and defatting treatment, followed by ultrafiltration, and then spray drying to obtain donkey-hide gelatin peptide;

[0024] The complex enzyme is a mixture of alkaline protease, neutral protease, pepsin and papain in a mass ratio of 3:1:2:1.

[0025] The present invention does not impose any particular limitation on the sources of alkaline protease, neutral protease, pepsin and papain, and commercially available products known to those skilled in the art may be used.

[0026] The present invention has concluded through single factor experiments and orthogonal tests that only when the mass ratio of alkaline protease, neutral protease, pepsin and papain is 3:1:2:1, the proportion of donkey-hide gelatin peptides with a molecular weight of less than 1000 Daltons in the prepared donkey-hide gelatin peptides is the highest; adjusting the dosage of any one of the proteases beyond the above ratio will reduce the proportion of donkey-hide gelatin peptides with a molecular weight of less than 1000 Daltons in the donkey-hide gelatin peptides.

[0027] In some embodiments of the present invention, in step 1, the blanching is specifically to add the donkey skin into a sodium carbonate aqueous solution for heating; the heating temperature is 60 to 80° C., the time is 20 to 30 minutes; the mass concentration of the sodium carbonate aqueous solution is 3 to 5%. The present invention does not specifically limit the amount of the sodium carbonate aqueous solution, and the amount of the sodium carbonate aqueous solution can fully immerse the donkey skin.

[0028] In the present invention, the purpose of blanching with sodium carbonate aqueous solution is: sodium carbonate can cause the protein and collagen fibers in the donkey skin to swell and soften to a certain extent, which is helpful for the subsequent steaming and extraction process; at the same time, the alkaline environment helps to remove fat, impurities and residual hair on the surface of the donkey skin, making the donkey skin cleaner; sodium carbonate can also neutralize the acidic substances in the donkey skin and reduce the fishy smell.

[0029] In some embodiments of the present invention, in step 1, the washing is carried out using water.

[0030] In a preferred embodiment of the present invention, in step 2, the mass ratio of the block donkey skin to the water is 1:10-15; the ultrasonic treatment conditions are set as: ultrasonic power 200-300w, ultrasonic time 1h.

[0031] In a preferred embodiment of the present invention, in step 2, the cooking temperature is 100° C. and the cooking time is 1 to 2 hours.

[0032] In step 2, the filtration is performed using a 300-mesh non-woven fabric.

[0033] In a preferred embodiment of the present invention, in step 3, the concentration of the complex enzyme in the filtrate 1 is 2 wt% to 3 wt%.

[0034] In a preferred embodiment of the present invention, in step 3, the conditions of the ultrasonic treatment are set as: ultrasonic power 200-300w, ultrasonic time 1h.

[0035] In a preferred embodiment of the present invention, in step 3, the temperature of the enzymatic hydrolysis is 45° C. and the time is 3 to 5 hours.

[0036] In some embodiments of the present invention, the enzyme is inactivated by raising the temperature to 95-100° C. and maintaining the temperature for 10 min.

[0037] In a preferred embodiment of the present invention, in step 3, debittering, decolorizing and degreasing are carried out by adding activated carbon.

[0038] In a preferred embodiment of the present invention, the ultrafiltration uses an ultrafiltration membrane with a molecular cutoff of 3000Da.

[0039] The present invention does not impose any particular limitation on the spray drying equipment, and any spray drying equipment commonly used by those skilled in the art may be used.

[0040] The second aspect of the present invention provides donkey-hide gelatin peptide prepared according to the above preparation method.

[0041] The third aspect of the present invention provides the use of the above-mentioned donkey-hide gelatin peptide in the preparation of iron or blood supplement drugs.

[0042] The technical solutions described in the present invention, unless otherwise specified, are all conventional solutions in the art, and the reagents or raw materials used, unless otherwise specified, are purchased from commercial channels or have been disclosed.

[0043] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0044] Example 1

[0045] Step 1, after dehairing and degreasing the fresh donkey hide, cutting into pieces, blanching in a sodium carbonate aqueous solution having a mass concentration of 3-5% at 70-80° C. for 25 min, and then washing with drinking water to obtain a block of donkey hide having a size of (5-10)×(5-10);

[0046] Step 2, adding the block donkey skin to water (the mass ratio of the block donkey skin to water is 1:15) for ultrasonic treatment, with an ultrasonic power of 250w and an ultrasonic time of 1h, then boiling (100°C) for 1h, filtering with a 300-mesh non-woven fabric, and collecting the filtrate to obtain a filtrate 1;

[0047] Step 3, adding a composite enzyme (a mixture of alkaline protease, neutral protease, pepsin and papain in a mass ratio of 3:1:2:1) to the above-mentioned filtrate 1 for ultrasonic treatment, wherein the concentration of the composite enzyme in the filtrate 1 is 2.5wt%, the ultrasonic power is 250w, the ultrasonic time is 1h, and then enzymolysis is carried out at 45°C for 4h. After the enzymolysis is completed, the enzyme is inactivated at 100°C for 10min, and centrifugal filtration is performed. Activated carbon is added to the obtained filtrate 2 for debittering, decolorizing and defatting treatment. After filtering and removing the activated carbon, ultrafiltration is performed using an ultrafiltration membrane with a molecular retention capacity of 3000Da, and then spray drying is performed to obtain donkey-hide gelatin peptide.

[0048] Comparative Example 1

[0049] The only difference from Example 1 is that the ultrasonic treatment step in step 2 is omitted.

[0050] Comparative Example 2

[0051] The only difference from Example 1 is that the ultrasonic treatment step in step 3 is omitted.

[0052] Comparative Example 3

[0053] The only difference from Example 1 is that the ultrasonic treatment steps in step 2 and step 3 are omitted.

[0054] The yields of donkey-hide gelatin peptides prepared in the examples and comparative examples were calculated, and the results are summarized in Table 1.

[0055] Table 1

[0056] Yield (%) Example 1 38.9 Comparative Example 1 32.0 Comparative Example 2 36.1 Comparative Example 3 35.4

[0057] Note: The yield calculation formula in Table 1 is as follows: Yield = mass of donkey-hide gelatin peptide / mass of donkey skin × 100%.

[0058] The molecular weight distribution of donkey-hide gelatin prepared in the examples and comparative examples was tested according to GB / T22492, and the results are shown in Table 2.

[0059] Table 2

[0060]

[0061]

[0062] Note: “ / ” in the table means no test was conducted or no result was detected.

[0063] The iron supplementation performance of the donkey-hide gelatin peptides prepared in the examples and comparative examples was tested as follows:

[0064] Test method: Kunming mice, SPF grade, half male and half female, free drinking water and feeding under suitable temperature and humidity conditions, adaptive feeding for one week, randomly divided into blank group, model control group and experimental group (respectively fed with donkey-hide gelatin peptides in Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 3, corresponding to Example 1, Comparative Example 1, Comparative Example 2, and Comparative Example 3), 10 in each group. Modeling was performed on the model control group mice and the experimental group mice, specifically giving the model control group mice low-iron feed, the blank control group was given normal feed, and continued to feed for 42 days. At the same time, each experimental group of mice was gavaged with donkey-hide gelatin peptides in the corresponding embodiment and comparative example (gavage dose of 30 mg / kg) every day, and the model control group and the blank group were gavaged with equal amounts of normal saline, once a day. After the last administration, blood was collected to measure the average value of mouse serum iron (SI) and serum ferritin (SF), and the results were counted in Table 3.

[0065] Table 3

[0066] SI (μmol / L) SF (ng / mL) Blank group 106.3 56.8 Model control group 36.9 18.7 Example 1 102.4 42.8 Comparative Example 1 79.3 28.1 Comparative Example 2 72.5 26.4 Comparative Example 3 69.2 24.0

[0067] The blood-tonifying performance of the donkey-hide gelatin peptides prepared in the examples and comparative examples was tested as follows:

[0068] Experimental method: Kunming mice, SPF grade, half male and half female, free drinking water and food under suitable temperature and humidity conditions, after one week of adaptive feeding, were randomly divided into blank group, model control group and experimental group, 10 in each group. Each experimental group was gavaged with donkey-hide gelatin peptide prepared in each embodiment and comparative example (gavage dose was 30 mg / kg), and the blank group and model control group were gavaged with an equal amount of normal saline, once a day, for a total of 15 days. From the 5th day of administration, the model control group and each example group mice were intraperitoneally injected with cyclophosphamide (once a day, 5 mg / kg each time) for 4 consecutive days, and the blank group was injected with an equal amount of normal saline. 2h after the last administration, blood was collected to determine the number of red blood cells (RBC) and hemoglobin (HGB) of mice and averaged, and the results were statistically recorded in Table 4.

[0069] Table 4

[0070] <![CDATA[RBC(×10 12 / L)]]> HGB(g / L) Blank group 8.69 148.5 Model control group 6.03 92.4 Example 1 8.07 135.8 Comparative Example 1 7.13 109.3 Comparative Example 2 6.85 102.4 Comparative Example 3 6.51 99.8

[0071] It can be seen from Table 3 and Table 4 that the donkey-hide gelatin peptide prepared in Example 1 of the present invention has good iron and blood supplementing effects.

[0072] The embodiments described above are only descriptions of the preferred modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims

1. A method for preparing donkey-hide gelatin peptide, characterized in that: The following steps are involved: Step 1, after removing hair and fat from fresh donkey skin, cutting into pieces, blanching and washing, to obtain block donkey skin; Step 2, adding the block of donkey skin into water for ultrasonic treatment, then steaming, and filtering to collect filtrate 1; Step 3, adding a composite enzyme to the filtrate 1 for ultrasonic treatment, followed by enzymatic hydrolysis, enzyme inactivation, and centrifugal filtration, and the obtained filtrate 2 is subjected to debittering, decolorizing, and defatting treatment, followed by ultrafiltration, and then spray drying to obtain donkey-hide gelatin peptide; The complex enzyme is a mixture of alkaline protease, neutral protease, pepsin and papain in a mass ratio of 3:1:2:

1.

2. The method for preparing donkey-hide gelatin according to claim 1, characterized in that: In step 2, the mass ratio of the block donkey skin to the water is 1:10-15; the ultrasonic treatment conditions are set as: ultrasonic power 200-300w, ultrasonic time 1h.

3. The method for preparing donkey-hide gelatin according to claim 1, characterized in that: In step 2, the cooking temperature is 100° C. and the cooking time is 1 to 2 hours.

4. The method for preparing donkey-hide gelatin according to claim 1, characterized in that: In step 3, the concentration of the complex enzyme in the filtrate 1 is 2 wt% to 3 wt%.

5. The method for preparing donkey-hide gelatin according to claim 1, characterized in that: In step 3, the conditions of the ultrasonic treatment are set as: ultrasonic power 200-300w, ultrasonic time 1h.

6. The method for preparing donkey-hide gelatin according to claim 1, characterized in that: In step 3, the temperature of the enzymatic hydrolysis is 45° C. and the time is 3 to 5 hours.

7. The method for preparing donkey-hide gelatin peptide according to claim 1, characterized in that: In step 3, activated carbon is added to carry out debittering, decoloring and degreasing.

8. The method for preparing donkey-hide gelatin according to claim 1, characterized in that: The ultrafiltration uses an ultrafiltration membrane with a molecular cutoff of 3000Da.

9. The donkey-hide gelatin peptide prepared according to the preparation method according to any one of claims 1 to 8.

10. Use of the donkey-hide gelatin peptide according to claim 9 in the preparation of iron-supplementing or blood-supplementing drugs.

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