阅读说明：本技术 一种多肽类复合生物保鲜剂及其应用 (Polypeptide composite biological preservative and application thereof ) 是由 谢海伟 吴琳芝 黄欲菲 吴思婷 莫小燕 区嘉朗 于 2019-09-26 设计创作，主要内容包括：本发明公开一种多肽类复合生物保鲜剂及其应用,包括合成鲎素抗菌肽Tac Ⅰ和抗菌组分；所述抗菌组分为壳聚糖和Nisin中的一种或两种。采用本发明提供的技术方案解决了现有缺少对低温环境下的冷鲜肉进行保鲜的应用技术问题。(The invention discloses a polypeptide composite biological preservative and application thereof, comprising synthetic tachyplesin antimicrobial peptide Tac I and an antimicrobial component; the antibacterial component is one or two of chitosan and Nisin. The technical scheme provided by the invention solves the technical problem that the application technology for keeping the chilled meat fresh in the low-temperature environment is lacked.)

1. A polypeptide composite biological preservative is characterized in that: comprises synthesizing tachyplesin antibacterial peptide Tac I and an antibacterial component; the antibacterial component is one or two of chitosan and Nisin.

2. The polypeptide composite biological preservative according to claim 1, which is characterized in that: comprises synthesizing tachyplesin antibacterial peptide Tac I and chitosan; the mass ratio of the synthetic tachyplesin antimicrobial peptide Tac I to the chitosan is 1: 333 to 1000.

3. The polypeptide composite biological preservative according to claim 1, which is characterized in that: comprises synthesizing tachyplesin antibacterial peptides Tac I and Nisin; the mass ratio of the synthetic tachyplesin antimicrobial peptide Tac I to Nisin is 1: 3-10.

4. The polypeptide composite biological preservative according to claim 1, which is characterized in that: comprises synthesizing tachyplesin antibacterial peptide Tac I, chitosan and Nisin; the mass ratio of the synthetic tachyplesin antimicrobial peptide Tac I to the synthetic tachyplesin antimicrobial peptide Nisin is 1: 333-1000: 3 to 10.

5. The application of the polypeptide composite biological preservative is characterized in that: the application of the polypeptide composite biological preservative in chilled meat according to any one of claims 1 to 4.

6. The application of the polypeptide composite biological preservative as claimed in claim 5, wherein the polypeptide composite biological preservative is characterized in that: and forming a protective layer containing the polypeptide composite biological preservative on the surface of the chilled meat.

7. The application of the polypeptide composite biological preservative as claimed in claim 6, wherein the polypeptide composite biological preservative comprises the following components in percentage by weight: the refrigeration temperature of the chilled fresh meat is 0-4 ℃.

8. The application of the polypeptide composite biological preservative according to claim 7, which is characterized in that: and (3) smearing the polypeptide composite biological preservative on a preservative film, and covering the preservative film on the surface of the chilled fresh meat.

9. The application of the polypeptide composite biological preservative according to claim 7, which is characterized in that: the polypeptide composite biological preservative is directly smeared on the outer surface of the chilled fresh meat.

10. The application of the polypeptide composite biological preservative according to claim 7, which is characterized in that: and (3) soaking the polypeptide composite biological preservative into a solution containing the polypeptide composite biological preservative for 1-3 min.

Technical Field

The invention relates to the technical field of food preservation, in particular to a polypeptide composite biological preservative and application thereof.

Background

The preservation of food has become a focus of current research in order to prevent the food from rotting and deteriorating, and to maintain the color, the taste and the nutritional ingredients for a long time.

At present, the food in domestic market is kept fresh mainly by various preservatives or physical modes such as drying, pickling, heating and the like, but the methods can influence the reduction of the sensory flavor and the nutritional value of the food to a certain extent, and the use of chemical preservatives can bring negative effects to health. Various chemical antistaling agents have been shown to be harmful to human body, such as potassium sorbate, sodium benzoate, nitrates, etc. For example, nitrate, while inhibiting microbial growth during use, can also produce nitrite, which can pose a safety hazard. The antibacterial peptide has the characteristics of acid resistance, high temperature resistance, good solubility, strong thermal stability and the like, has obvious inhibiting and eliminating effects on most gram-negative and gram-positive bacteria in food, is easy to digest and hydrolyze by protease in human bodies and livestock bodies without toxic and side effects, and can be developed into a novel green food preservative with application value.

The research and development and application of novel food preservatives are important research directions in the fields of food preservation and preservation, and the polypeptide antibacterial preservatives are more and more emphasized due to the characteristics of high efficiency, low degree, no residue and the like. At present, the nisin is a microbial antibacterial peptide, can obviously inhibit the growth and the propagation of bacteria, and can control the growth of bacterial spores to further prolong the shelf life of food. However, the polypeptide antibacterial peptide is used as the preservative independently, has restriction, and is mainly characterized by poor stability, high price, difficult obtainment and single preservation mechanism when being used independently, so that the composite biological preservative prepared by compounding the polypeptide antibacterial agent and various biological preservatives can play a role in synergistic preservation, has more obvious effect and lower side effect in food preservation, and is an important direction for developing novel preservatives at present.

In the field of food preservation, temperature is also an important factor influencing food preservation. In most cases, the low temperature can effectively inhibit the growth and propagation of most microorganisms, but some low-temperature putrefying bacteria can effectively survive in a low-temperature environment, such as Listeria monocytogenes, which can survive and propagate in an environment at 4 ℃, a common refrigerator cannot prevent the growth and propagation of the low-temperature putrefying bacteria, and if the cold fresh meat is polluted by the Listeria monocytogenes, food poisoning can be caused, so that food safety incidents are caused, and the low-temperature putrefying bacteria are one of the main pathogenic bacteria which threaten human health for refrigerated foods. China is a meat producing country and a consuming country, and the main consumption raw meat in China is supplied to markets in three forms of hot fresh meat, cold fresh meat and frozen meat at present. Among them, chilled fresh meat is more and more favored by consumers in the meat consumption market because of its unique advantages of high nutritional value, bright red color, low juice loss rate, etc.

Therefore, an antistaling agent which can effectively keep fresh of fresh meat in a low-temperature environment is needed. To ensure the effective fresh keeping of the chilled meat. To this end, the present invention has been made to aim at a substance having a broad antimicrobial spectrum and being useful for the preservation of foods, such as tachyplesin.

The tachypleus amebocyte lysate is composed of 17 amino acids, has biological activities of broad-spectrum antibacterial, antiviral, antifungal, tumor cell proliferation and differentiation inhibition and the like, and has huge potential research and application values. Besides being directly applied to food, the antibacterial peptide can also be applied to a fresh-keeping packaging material as an additive, and the antibacterial peptide is applied to the middle of a fresh-keeping film to play a role in bacteriostasis, and the method is safer and more convenient. Therefore, in recent years, research, development and utilization of polypeptide type preservatives become a hot spot in food industry, and the application of the polypeptide type preservatives in the field of food preservation and preservation has wide development prospect.

Disclosure of Invention

In order to solve the problems, the invention provides a polypeptide composite biological preservative and application thereof, and the technical scheme provided by the invention solves the technical problem that the application for preserving cold fresh meat in a low-temperature environment is lacked.

In order to solve the technical problems, the invention provides a polypeptide composite biological preservative and application thereof, which comprises synthesizing a tachyplesin antimicrobial peptide Tac I and an antimicrobial component; the antibacterial component is one or two of chitosan and Nisin.

Preferably, the synthetic tachyplesin antimicrobial peptide Tac I and chitosan are included; the mass ratio of the synthetic tachyplesin antimicrobial peptide Tac I to the chitosan is 1: 333 to 1000.

Preferably, the method comprises synthesizing tachyplesin antimicrobial peptides TacI and Nisin; the mass ratio of the synthetic tachyplesin antimicrobial peptide Tac I to Nisin is 1: 3-10.

Preferably, the method comprises synthesizing tachyplesin antimicrobial peptide Tac I, chitosan and Nisin; the mass ratio of the synthetic tachyplesin antimicrobial peptide Tac I to the synthetic tachyplesin antimicrobial peptide Nisin is 1: 333-1000: 3 to 10.

An application of the polypeptide composite biological preservative comprises the application of the polypeptide composite biological preservative in chilled meat.

Preferably, a protective layer containing the polypeptide composite biological preservative is formed on the surface of the cold fresh meat.

Preferably, the refrigeration temperature of the cold fresh meat is 0-4 ℃.

Preferably, the polypeptide composite biological preservative is smeared on a pretreated preservative film, and then the preservative film is covered on the surface of the chilled fresh meat.

Preferably, the polypeptide composite biological preservative is directly smeared on the outer surface of the chilled meat.

Preferably, the polypeptide composite biological preservative is used for soaking the chilled fresh meat in a solution containing the polypeptide composite biological preservative for 1-3 min.

Compared with the prior art, the invention has the advantages that:

(1) the invention provides a polypeptide composite biological preservative, which comprises a synthetic tachyplesin antibacterial peptide Tac I and an antibacterial component; the antibacterial component is one or two of chitosan and Nisin. The tachypleus amebocyte lysate has the biological activities of broad-spectrum antibacterium, antivirus, antifungal, tumor cell proliferation and differentiation inhibition and the like, can effectively inhibit the growth and the propagation of various microorganisms, and prolongs the fresh-keeping time of food.

(2) The invention provides a polypeptide compound biological preservative, wherein nisin is a microbial antibacterial peptide, can obviously inhibit the growth and the propagation of bacteria, and can control the growth of bacterial spores so as to prolong the shelf life of food.

(3) The invention provides a polypeptide composite biological preservative, wherein chitosan is derived from chitin in shrimp and crab shells and is a polysaccharide substance obtained by deacetylation in a concentrated alkaline solution. A main chain of a chitosan molecule is connected with more-NH 2 and-OH groups which can chelate metal ions initiating grease oxidation, so that a reaction chain of grease oxidation is broken, and fat oxidation is inhibited; the polycation nature of chitosan causes it to interact with the negative charge on the surface of many microorganisms, causing a large change in the cell surface, which in turn causes leakage of intracellular material and cell death. Therefore, the antibacterial preservative has the characteristics of degradability, no toxicity, oxidation inhibition and broad-spectrum bacteriostasis, and is very suitable for the field of food preservation.

(4) If the polypeptide antibacterial peptide is used alone as a preservative, the problems of poor self-stability, high price, difficult obtainment and single preservation mechanism caused by single use can be caused. The invention provides a polypeptide composite biological preservative, which is prepared by compounding synthetic tachyplesin antimicrobial peptide Tac I, chitosan and nisin according to a proper proportion.

(5) The invention provides an application of a polypeptide composite biological preservative, in particular to an application in chilled meat. In the prior art, fresh meat at normal temperature is mostly preserved, and most people normally think that the cold storage of a refrigerator can effectively inhibit the growth of microorganisms, so that the fresh meat does not need to be treated. But some low-temperature putrefying bacteria still can grow and reproduce at low temperature, and the food quality and safety are influenced. The invention fully utilizes the safety, oxidation resistance and broad-spectrum bacteriostasis of the composite biological preservative, screens out proper mixing proportion through a combined formula to effectively inhibit and kill microorganisms causing the decay of the chilled fresh meat products, thereby reducing the storage cost in the production, storage and transportation commercial circulation of the chilled fresh meat products, and prolonging the shelf life of the meat products

Drawings

FIG. 1 is a line graph of the effect of different preservative solutions on pH change during cold storage of chilled fresh pork;

FIG. 2 is a line graph of the effect of different preservation solutions on the change in the total number of colonies during cold storage of chilled fresh pork;

fig. 3 is a line graph of the effect of different preservative solutions on the change in the rate of juice loss during the cold storage of chilled fresh pork.

FIG. 4 is a schematic of the experimental technique of the present invention.

Detailed Description

The invention will be further described with reference to the drawings and specific examples, which are not to be construed as limiting the invention. Modifications or substitutions to the methods, procedures, and conditions of the present invention may be made without departing from the spirit and scope of the invention. Unless otherwise indicated, the experimental procedures used in the examples are all conventional procedures and techniques well known to those skilled in the art, and all reagents or materials used are commercially available.

Aiming at the lack of an application technology for keeping the chilled meat fresh in a low-temperature environment at present, the embodiment of the invention provides a polypeptide composite biological preservative which mainly comprises the following materials:

synthesizing tachyplesin antibacterial peptide Tac I (synthetic sequence: NH2-K-W-C-F-R-V-C-Y-R-G-I-C-Y-R-R-C-R-CONH 2); lactein (nisin), chitosan.

Wherein, the synthetic tachyplesin antimicrobial peptide Tac I is an essential component, and the lactein (nisin) and the chitosan are alternative components, which can be selected to complete the matching, or can be selected to complete the matching. When chitosan is selected to be matched with the synthetic tachyplesin antibacterial peptide Tac I, the mass ratio of the synthetic tachyplesin antibacterial peptide Tac I to the chitosan is 1: 333-1000; when Nisin (Nisin) is selected to be matched with the synthetic tachyplesin antibacterial peptide Tac I, the mass ratio of the synthetic tachyplesin antibacterial peptide Tac I to Nisin is 1: 3-10; if the Nisin (Nisin) and the chitosan are simultaneously selected and matched with the synthetic tachyplesin antibacterial peptide TacI, the mass ratio of the synthetic tachyplesin antibacterial peptide TacI to the chitosan to the Nisin is 1: 333-1000: 3 to 10.

Aiming at the polypeptide composite biological preservative, the fresh longan pulp is used as a detection sample, and the listeria monocytogenes is used as an experimental strain to explain the preservation effect of the polypeptide composite biological preservative provided by the embodiment in detail.

The experimental procedure is as follows:

fresh Longqian meat → 10 g/part cut under aseptic condition → put into different fresh-keeping solution and soak for 2min → refrigeration in refrigerator at 4 deg.C around → test sample. In the detection process, time gradients of 1, 4, 5, 7, 9 and 11 (days) are set according to unit days, changes of freshness index such as sensory quality index, pH value, total bacterial colony number and juice loss rate of each processed sample at one time are measured, and the optimal combination of freshness effect is screened.

Wherein, the sensory quality index in the freshness index is measured

According to the GB/T22210-2008 standard, a sensory evaluation panel consisting of 5 professionals without physiological defects was asked to perform sensory evaluation according to the color, gloss, smell, elasticity, viscosity, water content and the like of meat. And scoring by adopting a ten-point system. The meat is comprehensively evaluated in four aspects of color, viscosity, elasticity and smell. The evaluation criteria are shown in Table 1.

TABLE 1 pork sensory evaluation criteria

Wherein the pH value in the freshness index is measured

The measurement was carried out according to GB/T9695.5-2008 "pH measurement of meat and meat products". The same amount of meat samples is taken from the refrigerating chamber of the refrigerator, cut into pieces, added with a certain amount of sterile water, and measured by a pH acidimeter. The freshness of the meat is evaluated by the pH value, namely deteriorated meat is more than or equal to 6.7; the second-level fresh meat is not less than 6.3 and not more than 6.6; the first-grade fresh meat is not less than 5.8 and not more than 6.2, and the pH value of the cold fresh meat can be increased along with the deepening of the putrefaction degree.

Wherein, the determination of the total number of colonies in the freshness index:

respectively sampling 5g of samples from a refrigerating chamber of a refrigerator, shearing the samples in a superclean workbench, placing the samples in a conical flask filled with 45mL of sterile water, and shaking up to prepare a sample homogenizing solution with the ratio of 1: 10; then, the measurement was carried out with reference to GB 4789.2-2016 national Standard for food safety-Total colony count measurement. The freshness of the meat is divided into three grades according to the total number of bacterial colonies, wherein the freshness is less than 1 multiplied by 104cfu/g is fresh meat, the freshness is inferior meat between 1 multiplied by 104cfu/g and 1 multiplied by 106cfu/g, and the freshness is more than 1 multiplied by 106cfu/g is deteriorated meat.

Wherein, the determination of the juice loss rate in the freshness index:

weighing the prepared meat sample (W)₀) Fresh-keeping and cold-storing in a refrigerator at 0-4 deg.C, taking out the meat sample every three days, wiping the juice on the surface layer of the meat sample with clean filter paper, and weighing (Wt)

Meat-like juice loss rate (DL):

aiming at the determination of the various preservation indexes, the embodiment adopts different preservation solutions to complete the preservation test of the fresh longan pulp, the different preservation solutions comprise experimental groups and the polypeptide composite biological preservative of the scheme, and the experimental groups are explained in detail as follows:

the experimental groups are mainly divided into blank control groups, single fresh-keeping experimental groups and multi-factor control groups with different component ratios under multiple factors. The method comprises the following specific steps:

group A: PBS buffer solution as blank control group;

group B: nisin solution of 0.2g/L, which is a single preservation experimental group;

group C: 2% chitosan solution, a single preservation experimental group.

TABLE 2 influence of different preservative solutions on sensory evaluation changes during chilled meat refrigeration

TABLE 3 Effect of different preservative solutions on the pH of chilled fresh pork

TABLE 4 Effect of different preservative solutions on the total number of colonies in chilled fresh pork

Note: the above data are log of the total number of colonies/(cfu/g)

TABLE 5 Effect of different preservative solutions on the loss of chilled fresh pork juice

The following will illustrate the multi-factor control group with different ratios of ingredients under multiple factors from specific examples: