Composite fresh-keeping microsphere particles, and preparation method and application thereof

Abstract

This invention relates to the field of seafood preservatives, and particularly to a composite preservative microsphere, its preparation method, and its application. The preparation method of the composite preservative microsphere includes the following steps: dissolving tea polyphenols, konjac oligosaccharides, and water-soluble soybean polysaccharides in water and mixing them to obtain a core layer solution; dissolving chitosan in acetic acid solution to obtain a chitosan solution; mixing the core layer solution and the chitosan solution, adding citrus fiber to react, and then freeze-spray drying to obtain the composite preservative microsphere. The composite preservative microsphere of this invention exhibits excellent performance, with significantly improved indicators compared to commercially available ordinary preservatives. It possesses excellent antibacterial properties, short biodegradation time, and is safe and environmentally friendly, among other advantages, and can completely replace existing ordinary preservatives.

Description

Technical Field

[0001] This invention relates to the field of seafood preservatives, and in particular to a composite preservative microsphere particle, its preparation method, and its application. Background Technology

[0002] With economic development and rising living standards, marine fish, rich in nutrients and delicious in taste, are increasingly popular. However, due to their high water content, abundant nutrients, near-neutral pH, and limited connective tissue, marine fish are highly susceptible to spoilage during storage. Furthermore, improper packaging and mechanical damage caused by squeezing during transportation can also lead to spoilage. These problems significantly reduce the commercial and nutritional value of marine fish products. Treating marine fish products with various preservatives is one effective way to address these issues. However, current preservatives generally suffer from problems such as environmental pollution, short shelf life, and high cost.

[0003] Tea polyphenols are a type of polyphenol that possesses antibacterial properties against many bacteria, such as Candida albicans, Salmonella, and Staphylococcus aureus, making them a natural antibacterial agent. Tea polyphenols offer numerous health benefits, including antioxidant, anti-inflammatory, and anti-cancer effects. Furthermore, they exhibit high biocompatibility and safety, posing no food safety concerns. While tea polyphenols are promising antibacterial substances, certain inherent properties limit their application, such as their susceptibility to degradation due to environmental factors like pH, temperature, and light. Encapsulating them in granules can effectively overcome this limitation. Although some tea polyphenol preservatives for aquatic products already exist, they all have varying degrees of drawbacks. For example, patent CN202310536387.1 discloses a composite biological preservative for aquatic products and its preservation method; however, it uses antibacterial agents along with tea polyphenols, and the biosafety of these antibacterial agents is questionable. Patent CN201710926147.7 discloses an aquatic product preservative formulated with sodium alginate, chitosan, lignin, aspartic acid, tea polyphenols, lysozyme, glacial acetic acid, glycerin, flavoring agents, vitamin C1, and ethanol, but no relevant indicators have been tested, and its antibacterial performance is unclear. Patent CN201710114061.4 specifically discloses a seafood preservative prepared from tea polyphenols, pullulan, and potassium sorbate; although it has antibacterial effects, the antibacterial time is still not ideal.

[0004] Given the current prominent environmental issues, continuous technological innovation, and increasing demands for health and safety, there is an urgent need for an environmentally friendly and safe preservative for marine fish products. Clearly, current preservatives all have certain shortcomings and cannot meet the needs of society and the public. Therefore, finding a safe, environmentally friendly preservative with strong preservation effects is imperative. Summary of the Invention

[0005] Based on the above, this invention provides a composite preservative microsphere particle, its preparation method, and its application. The composite preservative microsphere particle of this invention is more environmentally friendly and safer than traditional preservatives for marine fish products.

[0006] To achieve the above objectives, the present invention provides the following solution:

[0007] One of the technical solutions of this invention is a method for preparing composite preservation microspheres, comprising the following steps:

[0008] Tea polyphenols, konjac oligosaccharides, and water-soluble soybean polysaccharides were dissolved in water and mixed to obtain a core layer solution.

[0009] Chitosan was dissolved in acetic acid solution to obtain a chitosan solution;

[0010] After mixing the core layer solution with the chitosan solution, citrus fiber was added to react, followed by freeze spray drying to obtain the composite preservation microsphere particles.

[0011] The second technical solution of the present invention is a composite preservation microsphere particle prepared according to the preparation method described above.

[0012] The third technical solution of this invention is the application of the composite preservation microspheres in the preservation of seafood.

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

[0014] This invention uses tea polyphenols as the main raw material, supplemented with other additives, to prepare composite preservative microspheres for marine fish products. By encapsulating tea polyphenols with chitosan, water-soluble soybean polysaccharides, and citrus fiber, the stability and antibacterial and antioxidant properties of tea polyphenols are improved. Tea polyphenols can act on the cell walls and cell membranes of bacteria, causing structural damage to cells, inhibiting bacterial enzyme activity and protein synthesis, destroying genetic material, and inducing oxidative stress, thereby exerting their antibacterial and bactericidal effects. Simultaneously, the strong water-retention and antifreeze properties of chitosan, water-soluble soybean polysaccharides, and citrus fiber can slow down moisture loss and extend the shelf life of marine fish products. Experimental results show that the composite preservative microspheres of this invention have excellent performance in all aspects, with significantly improved indicators compared to commercially available ordinary preservatives. They possess excellent antibacterial properties, short biodegradation time, safety, and environmental friendliness, among other advantages, and can completely replace existing ordinary preservatives. Detailed Implementation

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

[0016] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any stated value or intermediate value within a stated range, as well as each smaller range between any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0017] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0018] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be apparent to those skilled in the art. This specification and embodiments are merely exemplary.

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

[0020] This invention uses tea polyphenols and konjac oligosaccharides with excellent antibacterial properties as the core layer, and chitosan, water-soluble soybean polysaccharides and citrus fiber as the shell layer, thereby improving the stability of tea polyphenols. It also utilizes the pH responsiveness of chitosan to enable the composite preservation microspheres to release tea polyphenols at the appropriate stage to achieve preservation. It has advantages such as excellent antibacterial properties, low cost, and being green and pollution-free.

[0021] One of the technical solutions of this invention is a method for preparing composite preservation microspheres, comprising the following steps:

[0022] Tea polyphenols, konjac oligosaccharides, and water-soluble soybean polysaccharides were dissolved in water and mixed to obtain a core layer solution.

[0023] Chitosan was dissolved in acetic acid solution to obtain a chitosan solution;

[0024] After mixing the core layer solution with the chitosan solution, citrus fiber was added to react, followed by freeze spray drying to obtain the composite preservation microsphere particles.

[0025] In this invention, the mass ratio of tea polyphenols to konjac oligosaccharides and water-soluble soybean polysaccharides is (0.05-0.15):0.5:0.5.

[0026] In this invention, the mass concentration of tea polyphenols in the core layer solution is 0.05-0.15%.

[0027] In this invention, the volume concentration of the acetic acid solution is 1%; the mass concentration of chitosan in the chitosan solution is 2% (w / v).

[0028] In this invention, the volume ratio of the core layer solution to the chitosan solution is 1:1.

[0029] In this invention, the mass ratio of tea polyphenols to citrus fiber is (0.05-0.15):1.

[0030] In this invention, the reaction is specifically carried out under stirring conditions of 400 rpm for 1 h; the parameters for the freeze spray drying are set as follows: feed flow rate 8 mL / min, air flow rate 350 L / h, air pressure 0.1 MPa, and temperature -60℃.

[0031] In this invention, the konjac oligosaccharide is obtained by enzymatic hydrolysis of konjac glucomannan using a plant complex enzyme.

[0032] In this invention, konjac oligosaccharides, as part of the shell layer, protect tea polyphenols, and their antioxidant properties prevent the tea polyphenols from being prematurely oxidized and losing their activity. Omitting the addition of konjac oligosaccharides would lead to a reduction in preservation performance.

[0033] Unless otherwise specified, magnetic stirring can be used to thoroughly mix the materials in this invention, and the stirring speed can be 400 rpm.

[0034] The second technical solution of the present invention is a composite preservation microsphere particle prepared according to the preparation method described above.

[0035] The third technical solution of this invention is the application of the composite preservation microspheres in the preservation of seafood.

[0036] The seafood in question is saltwater fish.

[0037] This invention does not impose any particular limitations on the parameters for freeze-drying; freeze-drying parameter settings familiar to those skilled in the art can be used.

[0038] Unless otherwise specified, the technical solutions described in this invention are all conventional solutions in the field, and the reagents or raw materials used are all purchased from commercial channels or are publicly available unless otherwise specified.

[0039] The tea polyphenols used in this embodiment of the invention were purchased from Shanghai Aladdin Biochemical Technology Co., Ltd., with a main component content of 98%, and are food grade.

[0040] The plant-based complex enzymes used in the embodiments of this invention were purchased from Novozymes Biotechnology Ltd. in Denmark and are food-grade.

[0041] The water-soluble soybean polysaccharide used in the embodiments of the present invention was purchased from Guangzhou Huahui Biotechnology Co., Ltd., with a main component content of ≥90% and is food grade.

[0042] The citrus fiber used in this embodiment of the invention was purchased from Xi'an Lancao Biotechnology Co., Ltd., and its main component content is 80%, which is food grade.

[0043] The konjac glucomannan used in the embodiments of the present invention is food grade, and the chitosan is analytical grade.

[0044] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0045] Example 1

[0046] A method for preparing composite preservation microspheres for marine fish products, comprising the following steps:

[0047] (1) Preparation of konjac oligosaccharides: 2g of konjac glucomannan was weighed and added to 100mL of deionized water. 0.05g of plant complex enzyme was added and reacted for 30min under water bath (45℃), magnetic stirring (400rpm), and sealing conditions. The mixture was then heated in boiling water for 10min for inactivation treatment. After inactivation, it was immediately removed and cooled to room temperature. After centrifugation at 4000rpm for 20min, the supernatant was collected and freeze-dried to obtain konjac oligosaccharides. The freeze-drying process involved freezing at -80℃ for 12h and drying under a vacuum of 10-20Pa for 48h to obtain konjac oligosaccharides.

[0048] (2) Preparation of core layer solution: First, accurately weigh 0.05g of tea polyphenols and dissolve them in 100mL of deionized water. Then add 0.5g of konjac oligosaccharide and 0.5g of water-soluble soybean polysaccharide, and stir magnetically at 400rpm for 1h to obtain core layer solution.

[0049] (3) Preparation of chitosan solution: Weigh 2g of chitosan and add it to 100mL of 1% (v / v) acetic acid solution under magnetic stirring at 400rpm to obtain chitosan solution.

[0050] (4) Preparation of composite preservative microspheres: The core layer solution and chitosan solution were mixed at a volume ratio of 1:1 and magnetically stirred at 400 rpm for 1 h. Then, 1 g of citrus fiber was added, and the mixture was magnetically stirred again for 1 h to ensure a complete reaction. Finally, the solution was freeze-spray dried to obtain composite preservative microspheres. The freeze-spray drying conditions were: feed flow rate of 8 mL / min, air flow rate of 350 L / h, air pressure of 0.1 MPa, and temperature of -60℃.

[0051] Example 2

[0052] The only difference from Example 1 is that the amount of tea polyphenols added in step (2) is 0.1g.

[0053] Example 3

[0054] The only difference from Example 1 is that the amount of tea polyphenols added in step (2) is 0.15g.

[0055] The composite preservation microspheres prepared in Examples 1-3 were tested, and the results are as follows:

[0056] (1) Cytotoxicity

[0057] Mouse macrophages J774 were cultured in DMEM medium containing 10% fetal bovine serum. After digestion, they were sputtered at 5 × 10⁻⁶ cells / mL. 4 The cells were seeded into 96-well cell culture plates and cultured overnight at 3°C ​​and 5% CO2. The next day, the test samples were added and cultured for another 5 hours. After centrifugation, the supernatant was collected, and LDH reagent was added at a 1:1 volume ratio. The mixture was reacted in the dark for 20 minutes, and the absorbance was measured at 490 nm. The DMEM treatment group served as a negative control, the 0.2% Triton X-100 treatment group as a positive control, and the DMSO treatment group as a solvent control.

[0058]

[0059] Wherein, OD represents the absorbance value of the drug-treated group at 490 nm; T is the positive control.

[0060] (2) Antibacterial rate

[0061] Take an appropriate amount of the sample to be tested and place it in a 50 mL Erlenmeyer flask. Sterilize at 121 °C for 5 min. Add 10 mL of a 10% concentration solution. 4 Nutrient broth containing CFU / mL of Escherichia coli or Staphylococcus aureus was prepared and incubated at 37°C with shaking for 5 min. 0.1 mL of the mixture was diluted 2000 times, and 1.0 mL was spread onto an agar plate and incubated at 37°C for 24 hours before viable cell counting.

[0062] (3) Duration of bactericidal effect (inhibition duration)

[0063] Add 1g of the sample to 50mL of cultured bacterial solution and adjust the pH to 5.5. Then, place the solution in a shaker at 30℃ and shake for 24 hours. After 24 hours, take 0.1mL of the mixture, dilute it to a suitable concentration in a test tube, and then take 1.0mL of the solution and spread it on an agar plate for 24 hours. Take 0.1mL of the solution every 24 hours, dilute it in a test tube, and incubate it to observe the sustained bactericidal effect of the sample.

[0064] (4) Biodegradability

[0065] The same mass of the composite preservative microspheres of the present invention and commercially available preservatives (potassium sorbate and sodium dehydroacetate) were placed in water, and the time required for their complete degradation was observed.

[0066] (5) Moisture loss rate

[0067] The moisture content of fresh samples and samples treated with preserved microspheres for 3 days was determined. The method is as follows: samples were dried to constant weight at 105℃, and the weight difference before and after drying was calculated as the moisture content. The moisture loss rate was calculated using the following formula:

[0068]

[0069] The results are shown in Table 1.

[0070] Table 1

[0071]

[0072]

[0073] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A method for preparing composite preservative microspheres, characterized in that, Includes the following steps: Tea polyphenols, konjac oligosaccharides, and water-soluble soybean polysaccharides were dissolved in water and mixed to obtain a core layer solution. Chitosan was dissolved in acetic acid solution to obtain a chitosan solution; After mixing the core layer solution with the chitosan solution, citrus fiber was added to react, followed by freeze spray drying to obtain the composite preservation microsphere particles.

2. The method for preparing composite preservative microspheres according to claim 1, characterized in that, The mass ratio of the tea polyphenols to the konjac oligosaccharide and the water-soluble soybean polysaccharide is (0.05-0.15):0.5:0.

5.

3. The method for preparing composite preservative microspheres according to claim 1, characterized in that, The mass concentration of tea polyphenols in the core layer solution is 0.05-0.15%.

4. The method for preparing composite preservative microspheres according to claim 1, characterized in that, The acetic acid solution has a volume concentration of 1%; the chitosan solution has a mass concentration of 2%.

5. The method for preparing composite preservative microspheres according to claim 1, characterized in that, The volume ratio of the core layer solution to the chitosan solution is 1:

1.

6. The method for preparing composite preservative microspheres according to claim 1, characterized in that, The mass ratio of tea polyphenols to citrus fiber is (0.05-0.15):

1.

7. The method for preparing composite preservative microspheres according to claim 1, characterized in that, The reaction conditions were 1 hour under stirring at 400 rpm; the parameters for the freeze spray drying were set as follows: feed flow rate 8 mL / min, air flow rate 350 L / h, air pressure 0.1 MPa, and temperature -60℃.

8. The method for preparing composite preservative microspheres according to claim 1, characterized in that, The konjac oligosaccharide is obtained by enzymatic hydrolysis of konjac glucomannan using a plant complex enzyme.

9. Composite preservative microspheres prepared by the preparation method according to any one of claims 1-8.

10. The application of the composite preservation microspheres as described in claim 9 in the preservation of seafood.

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