阅读说明：本技术 一种肉制品蛋白保鲜防腐剂及其制备方法 (Meat product protein fresh-keeping preservative and preparation method thereof ) 是由 汤佳鹏 陈忠平 朱俐 陈肖 于 2020-05-21 设计创作，主要内容包括：本发明涉及食品保鲜技术领域,公开了一种肉制品蛋白保鲜防腐剂及其制备方法,制备方法包括：(1)蛋白酶抑制剂的提取；(2)将乳铁蛋白、乳酸链球菌素、溶菌酶和蛋白酶抑制剂与水混合,加入乙二胺四乙酸二钠,纳滤脱盐后浓缩液掺入羧甲基纤维素钠溶液制得肉制品蛋白保鲜防腐剂。本发明食品保鲜剂主要由蛋白质构成,喷洒到肉类表面后能够长期抑制微生物的生长,且用量少不改变食材的色泽和风味,是一种较安全的食品保鲜剂。(The invention relates to the technical field of food preservation, and discloses a meat product protein preservative and a preparation method thereof, wherein the preparation method comprises the following steps: (1) extracting a protease inhibitor; (2) mixing lactoferrin, nisin, lysozyme, a protease inhibitor and water, adding ethylene diamine tetraacetic acid, performing nanofiltration desalination, and mixing the concentrated solution with a sodium carboxymethylcellulose solution to obtain the meat product protein fresh-keeping preservative. The food preservative mainly comprises protein, can inhibit the growth of microorganisms for a long time after being sprayed on the surface of meat, does not change the color and flavor of food materials with small dosage, and is a safer food preservative.)

1. A preparation method of a fresh-keeping preservative for protein of meat products is characterized by comprising the following steps:

s1, extracting a protease inhibitor, namely α₂Concentrating the M crude extract, passing through Sephacryl S-300 gel chromatography column, eluting with 100mmol/L Tris-HCl buffer solution containing 0.5mol/L NaCl and having pH of 8.0, detecting the eluate at absorption peak of 280nm, collecting the eluate of the first peak, ultrafiltering, desalting, and concentrating to obtain protease inhibitor;

s2, mixing lactoferrin, nisin, lysozyme, the protease inhibitor and water, adding ethylene diamine tetraacetic acid to obtain a mixed solution, performing nanofiltration and desalination on the mixed solution to obtain a concentrated solution, and doping the concentrated solution into a sodium carboxymethylcellulose solution to obtain the meat product protein preservative.

2. The method for preparing the preservative for fresh-keeping of meat product proteins as claimed in claim 1, wherein in step S1, the ultrafiltration membrane has a molecular weight cut-off of 300-500 kDa.

3. The method for preparing the preservative for fresh-keeping of protein in meat products as claimed in claim 1, wherein the protein concentration of the protease inhibitor in step S1 is 80-100 μ g/L.

4. The method for preparing the fresh-keeping preservative for the protein of the meat product as claimed in claim 1, wherein in the step S2, the dosage ratio of the lactoferrin, the nisin, the lysozyme, the protease inhibitor, the disodium edetate and the water is (8-10) g, (3.5-4) g, (15-20) g, (3-5) mL, (8.4-16.8) g and 1000 mL.

5. The method for preparing the preservative for preserving the freshness of the protein of the meat products as claimed in claim 1, wherein in the step S2, the iron saturation degree of the lactoferrin is 10% -15%, and the specific activity of nisin is 1.0 × 10⁵-1.0×10⁶IU/mg, and the specific activity of lysozyme is 10000-20000 IU/mg.

6. The method of claim 1, wherein in step S2, the volume of the concentrated solution is 1/8-1/5 of the volume of the mixed solution.

7. The method of claim 1, wherein the concentrated solution is added in an amount of 5-9mL per 100mL of the sodium carboxymethyl cellulose solution in step S2.

8. The method for preparing the fresh-keeping preservative for protein of meat products as claimed in claim 1, wherein in step S2, the concentration of the sodium carboxymethyl cellulose solution is 10-30g/L, and the molecular weight of the sodium carboxymethyl cellulose in the sodium carboxymethyl cellulose solution is 30-50 kDa.

9. The protein preservative for meat products prepared by the preparation method according to any one of claims 1 to 8.

Technical Field

The invention relates to the technical field of food preservation, in particular to a protein preservative for meat products and a preparation method thereof.

Background

The traditional food fresh-keeping method is divided into a physical fresh-keeping method and a chemical fresh-keeping method. Physical preservation methods, such as low-temperature freezing preservation, salting preservation and the like, can only be used for short-term preservation, and inevitably change the original flavor of the aquatic products. However, the commonly used chemical fresh-keeping method, such as adding chemical antistaling agent and antibiotic, is more and more rejected by consumers because of adding chemical components to different degrees. In contrast, the biological preservative is not only harmless to the health of human bodies, but also does not change the flavor of food. The effect of some biological antistaling agents even exceeds that of chemical antistaling agents. Therefore, the development of novel, safe and efficient biological antistaling agent becomes the focus of attention of food researchers. However, the biological preservative has the problems of poor preservation effect and short time effect. Therefore, there is a need for a preservative which has a good preservative effect, a long preservative life and is harmless to the human body.

Disclosure of Invention

In view of the above, the invention aims to provide a meat product protein preservative and a preparation method thereof, wherein the meat product protein preservative can inhibit activities of microbial protease and amylase, is low in consumption, does not change color and flavor of food materials, and is a safer food preservative.

In order to solve the technical problems, the invention provides a preparation method of a protein fresh-keeping preservative for meat products, which comprises the following steps:

s1, extracting a protease inhibitor, namely α₂Concentrating the M crude extract, passing through Sephacryl S-300 gel chromatography column, eluting with 100mmol/L Tris-HCl buffer solution containing 0.5mol/L NaCl and having pH of 8.0, detecting the eluate at absorption peak of 280nm, collecting the eluate of the first peak, ultrafiltering, desalting, and concentrating to obtain protease inhibitor;

s2, mixing lactoferrin, nisin, lysozyme, the protease inhibitor and water, adding ethylene diamine tetraacetic acid to obtain a mixed solution, performing nanofiltration and desalination on the mixed solution to obtain a concentrated solution, and doping the concentrated solution into a sodium carboxymethylcellulose solution to obtain the meat product protein preservative.

Preferably, in step S1, the ultrafiltration membrane has a molecular weight cut-off of 300-500 kDa.

Preferably, in step S1, the protein concentration of the protease inhibitor is 80-100. mu.g/L.

Preferably, in step S2, the dosage ratio of the lactoferrin, the nisin, the lysozyme, the protease inhibitor, the disodium edetate and the water is (8-10) g, (3.5-4) g, (3-20) g, (3-5) mL, (8.4-16.8) g and 1000 mL.

Preferably, in step S2, the iron saturation of the lactoferrin is 10% to 15%, and the specific activity of nisin is 1.0 × 10⁵-1.0×10⁶IU/mg, and the specific activity of lysozyme is 10000-20000 IU/mg.

Preferably, in step S2, the volume of the concentrated solution is 1/8-1/5 of the volume of the mixed solution.

Preferably, in step S2, the concentrated solution is added in an amount of 5-9mL per 100mL of the sodium carboxymethyl cellulose solution.

Preferably, in step S2, the concentration of the sodium carboxymethyl cellulose solution is 10-30g/L, and the molecular weight of the sodium carboxymethyl cellulose in the sodium carboxymethyl cellulose solution is 30-50 kDa.

The invention also provides the meat product protein fresh-keeping preservative prepared by the preparation method.

Compared with the prior art, the invention has the following beneficial effects:

1) the fresh-keeping preservative for the protein of the meat product comprises Lactoferrin (LF), Nisin (Nisin), Lysozyme (LZ) and a protease inhibitor extracted by the method. The protease inhibitor extracted by the method can inhibit hydrolysis of LF, LZ and Nisin by protease, so that the LF, LZ and Nisin can exert long-acting antibacterial action, LZ can destroy bacterial cell walls, LF and Nisin can directly act on protoplasts, and the aim of keeping food fresh is fulfilled through synergistic antibacterial action among a plurality of components.

2) Disodium ethylene diamine tetraacetate is added in the preparation process of the fresh-keeping preservative for meat product protein, the disodium ethylene diamine tetraacetate can complex divalent iron, and the divalent iron is removed by nanofiltration, so that the purpose of LF deferrization can be achieved, and the antibacterial efficiency of LF is further improved.

3) The fresh meat contains rich iron, and the concentrated solution after nanofiltration desalination is doped into the sodium carboxymethyl cellulose solution, so that the sodium carboxymethyl cellulose film can effectively isolate iron and LF, and the failure of antibacterial action caused by the contact of the LF with iron is prevented.

Detailed Description

The invention will be better understood from the following examples. However, those skilled in the art will readily appreciate that the description of the embodiments is only for illustrating the present invention and should not be taken as limiting the invention as detailed in the claims.

The invention provides a preparation method of a fresh-keeping preservative for protein of meat products, which comprises the following steps:

s1, extracting a protease inhibitor, namely α₂Concentrating the M crude extract, passing through Sephacryl S-300 gel chromatography column, eluting with 100mmol/L Tris-HCl buffer solution containing 0.5mol/L NaCl and having pH of 8.0, detecting the eluate at absorption peak of 280nm, collecting the eluate of the first peak, ultrafiltering, desalting, and concentrating to obtain protease inhibitor;

s2, mixing lactoferrin, nisin, lysozyme, the protease inhibitor and water, adding ethylene diamine tetraacetic acid to obtain a mixed solution, performing nanofiltration and desalination on the mixed solution to obtain a concentrated solution, and doping the concentrated solution into a sodium carboxymethylcellulose solution to obtain the meat product protein preservative.

Specifically, firstly, α is taken₂Concentrating the M crude extract, passing through Sephacryl S-300 gel chromatography column, eluting with 100mmol/L Tris-HCl buffer solution containing 0.5mol/L NaCl and having pH of 8.0, detecting the eluate at absorption peak of 280nm, collecting the eluate of the first peak, ultrafiltering, desalting, and concentrating to obtain protease inhibitor; mixing lactoferrin, nisin, lysozyme and a protease inhibitor with water, adding Ethylene Diamine Tetraacetic Acid (EDTA) to obtain a mixed solution, performing nanofiltration and desalination on the mixed solution to obtain a concentrated solution, wherein the volume of the obtained concentrated solution is preferably 1/8-1/5 of the volume of the mixed solution; and finally, doping the concentrated solution into a sodium carboxymethyl cellulose solution to prepare the meat product protein fresh-keeping preservative.

The fresh meat contains rich iron, the concentrated solution after nanofiltration desalination is doped into the sodium carboxymethyl cellulose solution, the sodium carboxymethyl cellulose membrane can effectively isolate iron and LF, and the antibacterial effect failure caused by the contact of LF with iron is prevented, wherein the doping amount of the concentrated solution is preferably 5-9mL per 100mL of the sodium carboxymethyl cellulose solution, the molecular weight of the sodium carboxymethyl cellulose is preferably 30-50kDa, and the concentration of the sodium carboxymethyl cellulose is preferably 10-30 g/L.

In the present invention, the cut-off molecular weight of the ultrafiltration membrane used for ultrafiltration desalination is preferably 300-500kDa, and the protein concentration of the protease inhibitor after ultrafiltration desalination and concentration is preferably 80-100. mu.g/L. The meat product of the inventionIn the preparation process of the fresh-keeping preservative for the protein, the dosage proportion of lactoferrin, nisin, lysozyme, protease inhibitor, ethylene diamine tetraacetic acid disodium and water is preferably (8-10) g, (3.5-4) g, (15-20) g, (3-5) mL, (8.4-16.8) g and 1000mL, wherein the iron saturation of the lactoferrin is preferably 10-15%, and the specific activity of the nisin is preferably 1.0 × 10⁵-1.0×10⁶IU/mg, the specific activity of lysozyme is preferably 10000-20000 IU/mg.

The invention also provides the meat product protein fresh-keeping preservative prepared by the preparation method.

The Lactoferrin (LF) adopted in the fresh-keeping preservative for the protein of the meat product is an important non-heme iron-binding glycoprotein in milk, and monomer glycoprotein with bactericidal activity is in neutrophilic granulocyte particles. The bacteriostatic effect of LF is influenced by a number of factors, with the iron content of LF determining its bacteriostatic effect, and when LF is saturated with iron, its bacteriostatic properties will disappear.

Nisin (Nisin) also known as Nisin or transliteration as nixin, which is adopted in the fresh-keeping preservative for meat product protein of the invention, is a polypeptide substance generated by streptococcus lactis, can inhibit most gram-positive bacteria, and has strong inhibition effect on spores of bacillus, so that Nisin is widely applied to food industry as a food preservative. After being eaten, the natural food preservative can be quickly hydrolyzed into amino acid under the physiological pH condition of a human body and the action of alpha-chymotrypsin, does not change the normal flora in the intestinal tract of the human body and generate resistance problems such as other antibiotics, does not generate cross resistance with other antibiotics, and is efficient, nontoxic, safe and free of side effect.

The Lysozyme (LZ) adopted in the meat product protein fresh-keeping preservative can effectively hydrolyze peptidoglycan of bacterial cell walls. Against gram-positive bacteria (G)⁺) Such as micrococcus luteus, Bacillus subtilis, or Micrococcus muralis, the cell wall is almost entirely composed of peptidoglycan; for gram negativeSex bacteria (G)^-) Such as colibacillus, proteus, dysentery bacillus, pneumonia bacillus, etc., and the inner wall layer of the cell wall is peptidoglycan. Therefore, lysozyme can effectively destroy bacterial cell walls, and Lactoferrin (LF) and Nisin (Nisin) can directly act on protoplasts.

Through the synergistic antibacterial action among the components, the fresh-keeping preservative for the protein of the meat product achieves the aim of safe and long-acting fresh keeping of food.

In order to further illustrate the invention, the meat product protein preservative and the preparation method thereof provided by the invention are described in detail below with reference to the examples.

α₂The preparation method of the M crude extract comprises the following steps:

adding 0.1% (w/v) soybean trypsin inhibitor into fresh pig plasma at-5 deg.C, adding EDTA (ethylene diamine tetraacetic acid) solution to make its final concentration to 10mmol/L, performing fractional precipitation with 5% -18% polyethylene glycol 6000, stirring for 60min, centrifuging at 10000r/min at 4 deg.C for 30min, dissolving and precipitating with 5 times mass of 50mmol/L pH7.4 Tris-HCl solution, and dialyzing with 50mmol/L pH7.4 Tris-HCl solution to obtain α₂M crude extract.