Antibacterial and fresh-keeping agent for post-harvest sugar oranges and preparation and use method thereof

Abstract

The present invention provides an antibacterial fresh-keeping agent for post-harvest sugar oranges and a preparation and use method thereof, and relates to the technical field of citrus fruit preservation. The antibacterial fresh-keeping agent is composed of carboxymethyl chitosan, sodium alginate, cooked rice stem powder, konjac flour, walnut green peel fermentation extract, and Bacillus amyloliquefaciens, wherein the walnut green peel fermentation extract is obtained by fermenting the walnut green peel with red yeast. The present invention overcomes the shortcomings of the prior art, effectively ensures the fresh-keeping and water-retaining effect of sugar oranges through the combination of various components, natural substances and microorganisms, and effectively inhibits the growth of pythium, thereby achieving a good antibacterial effect.

Description

Technical Field

[0001] The invention relates to the technical field of citrus fruit preservation, and in particular to an antibacterial and fresh-keeping bacterial agent for post-harvest sugar oranges and a preparation method and use method thereof. Background Art

[0002] Sugar orange is a low-acid local excellent variety selected from the seedling mutation of ordinary sweet orange. It has the characteristics of beautiful fruit shape, bright color, sweet flavor, crisp and tender flesh. As a kind of citrus, post-harvest storage of sugar orange is one of the important links in production.

[0003] Generally speaking, the storage of sugar oranges after picking is mainly to prevent the sugar oranges from losing water, wilting, rotting and deterioration, and traditional storage methods include film coating, film single fruit packaging and ozone technology, all of which can reduce water loss to a certain extent. However, sugar oranges are easily infected by Fusarium during the planting process and are also easily carried with these Fusarium after picking. Among them, citrus brown rot fungus is the most prominent. The pathogen generally invades from the wound. The lesions start to be light brown and spread rapidly. It soon expands to the whole fruit and becomes water-soaked and soft. The lesions become soft and slightly concave, with a foul odor. After infection, the sugar oranges will quickly deteriorate, seriously affecting the storage quality. Therefore, for the storage of sugar oranges, it is necessary to effectively prevent Fusarium infection while preventing it from losing water to achieve the purpose of preservation and antibacterial. However, most of the existing antibacterial and bactericides are chemical synthetic substances. Although the antibacterial and sterilization effects are good, there are certain risks to food safety. Summary of the invention

[0004] In view of the deficiencies in the prior art, the present invention provides an antibacterial and fresh-keeping bacterial agent for post-harvest sugar oranges and a preparation and use method thereof. The antibacterial and fresh-keeping effect of the sugar oranges can be effectively guaranteed by the combination of various components, natural substances and microorganisms, while the growth of pythium can be effectively inhibited, thereby achieving a good antibacterial effect.

[0005] To achieve the above objectives, the present invention is implemented through the following technical solutions:

[0006] The invention discloses an antibacterial and fresh-keeping bacterial agent for post-harvest sugar oranges. The antibacterial and fresh-keeping bacterial agent is prepared from the following raw materials in parts by weight: 4-8 parts of carboxymethyl chitosan, 2-4 parts of sodium alginate, 2-4 parts of wild rice stem flour, 1-2 parts of konjac flour, 0.1-0.3 parts of walnut green peel fermentation extract, and 0.01-0.03 parts of Bacillus amyloliquefaciens; the walnut green peel fermentation extract is prepared by fermenting the walnut green peel with red yeast.

[0007] Preferably, the cooked rice stem powder is prepared by washing fresh rice stem, steaming it at 100-110° C. for 15-30 minutes, drying it, and then grinding it through a 100-mesh sieve.

[0008] Preferably, the preparation method of the walnut green peel fermentation extract comprises the following steps:

[0009] (1) Add water to the walnut green peel and boil for 3-5 minutes, then take it out and add 1-2 times distilled water to grind it into slurry, and the slurry is set aside;

[0010] (2) Inoculate 0.1%-0.2% of the total mass of walnut peel with red yeast into the above slurry, ferment for 18-24 hours, then press and filter, separate the filtrate and the residue, add 2-4 times of distilled water to the residue, stir evenly, let stand for 2-4 hours, centrifuge again, separate the supernatant, combine the supernatant and the filtrate, and obtain the extract for later use;

[0011] (3) The extract is vacuum concentrated to 1 / 5-1 / 8 of its original volume, then freeze-dried and crushed through a 100-mesh sieve to obtain a walnut green peel fermentation extract.

[0012] Preferably, the viable count of the red yeast is 1×10 7 -2×10 9 Pieces / g.

[0013] Preferably, the viable count of the Bacillus amyloliquefaciens is 1×10 7 -2×10 9 Pieces / g.

[0014] The preparation method of the antibacterial fresh-keeping agent comprises the following steps:

[0015] S1, the wild rice flour mixed with konjac flour was added to 10-15 times the volume of distilled water, and then mixed with Bacillus amyloliquefaciens, and cultured on a shaking table for 8-12h, and then mixed with carboxymethyl chitosan and stirred evenly to obtain a preliminary mixture for standby use;

[0016] S2. Add walnut green peel fermentation extract and sodium alginate to the above preliminary mixture, stir evenly and freeze-dry to obtain an antibacterial and fresh-keeping bacterial agent.

[0017] Preferably, in step S1, the rotation speed of the shaking table culture is 80-120 r / min, and the culture temperature is 30±2°C.

[0018] Preferably, during stirring in step S2, the water bath temperature is maintained at 40-45° C., the stirring time is 10-15 min, and the stirring speed is 120-800 r / min.

[0019] The method of using the antibacterial preservative agent is to add the antibacterial preservative agent into 30-40°C warm water and dilute it 30-50 times to prepare an antibacterial solution, then soak the sugar orange for 3-5 minutes and then take it out and air dry it naturally, or use the antibacterial solution to directly spray on the surface of the sugar orange and then air dry it naturally.

[0020] The present invention provides a post-harvest antibacterial and fresh-keeping bacterial agent for rock sugar oranges and a preparation and use method thereof, which has the following advantages over the prior art:

[0021] The invention adopts cooked rice stem flour, konjac flour and the like as raw materials, adopts red yeast to ferment walnut green peel to prepare walnut green peel fermentation extract, and combines with Bacillus amyloliquefaciens to prepare antibacterial and fresh-keeping bacterial agent for sugar orange after harvest, which can effectively inhibit the growth of citrus brown rot phytophthora, and is used in combination with carboxymethyl chitosan and sodium alginate to effectively prolong the storage period of sugar orange, prevent water loss of sugar orange, reduce the use of chemical substances, and improve the quality of sugar orange after storage. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is a schematic diagram of the growth of lesions of the sugar oranges in Experimental Group 1 after storage for 7 days in Example 2 of the present invention;

[0023] Figure 2 This is a schematic diagram of the growth of lesions of the sugar oranges in Experimental Group 2 after storage for 7 days in Example 2 of the present invention;

[0024] Figure 3 This is a schematic diagram of the growth of lesions of the sugar oranges in Experimental Group 3 after storage for 7 days in Example 2 of the present invention;

[0025] Figure 4 This is a schematic diagram of the growth of lesions of the sugar oranges in Experimental Group 4 after storage for 7 days in Example 2 of the present invention;

[0026] Figure 5 This is a schematic diagram of the growth of lesions of the sugar oranges in Experimental Group 5 after storage for 7 days in Example 2 of the present invention;

[0027] Figure 6 This is a schematic diagram of the growth of lesions of the sugar oranges in Experimental Group 6 after storage for 7 days in Example 2 of the present invention;

[0028] Figure 7 This is a schematic diagram of the growth of lesions on the sugar oranges in Experimental Group 7 in Example 2 of the present invention after storage for 7 days. DETAILED DESCRIPTION

[0029] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention is clearly and completely described below in combination with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0030] The red yeast used below was purchased from China Center for Type Culture Collection, with the collection number CCTCC No.AY2018006 and the viable count of 2×10 9CFU / g; Bacillus amyloliquefaciens was purchased from China General Microbiological Culture Collection Center, with the preservation number of CGMCC No. 1.803 and the viable count of 1×10 9 CFU / g; Saccharomyces boulardii was purchased from Xi'an Minglang Biotechnology Co., Ltd., with the product number of 265987 and the viable count of 2×10 9 CFU / g. Example 1:

[0031] I. Raw material preparation:

[0032] 1. Preparation of water bamboo powder:

[0033] 1.1 Water bamboo powder A: Wash fresh water bamboo, steam it with 105±5°C hot steam for 20 min, then dry it at 50°C and grind it through a 100-mesh sieve to obtain water bamboo powder A.

[0034] 1.2 Water bamboo powder B: Wash fresh water bamboo, dry it at 50°C and grind it through a 100-mesh sieve to obtain water bamboo powder B.

[0035] 2. Preparation of fermented extract of walnut green husk:

[0036] 2.1 Extract A:

[0037] (1) Boil walnut green husk in water until boiling for 5 min, then take it out, add 2 times distilled water and grind it into pulp to obtain the pulp for standby;

[0038] (2) Inoculate 0.15% of Rhodotorula glutinis based on the total mass of walnut green husk into the above pulp, ferment for 20 h, then press and filter to separate the filtrate and the residue. Add 3 times distilled water to the residue, stir evenly and let it stand for 3 h, then centrifuge at 3200 r / min for 10 min to separate the supernatant. Combine the supernatant and the filtrate to obtain the extract for standby;

[0039] (3) Vacuum-concentrate the above extract to 1 / 6 of the original volume, then freeze-dry, pulverize and pass through a 100-mesh sieve to obtain extract A.

[0040] 2.2 Extract B:

[0041] (1) Boil walnut green husk in water until boiling for 5 min, then take it out, add 2 times distilled water and grind it into pulp to obtain the pulp for standby;

[0042] (2)Inoculate 0.15% of Saccharomyces boulardii based on the total mass of walnut green husk into the above pulp, ferment for 20 h, then press and filter to separate the filtrate and the residue. Add 3 times distilled water to the residue, stir evenly and let it stand for 3 h, then centrifuge at 3200 r / min for 10 min to separate the supernatant. Combine the supernatant and the filtrate to obtain the extract for standby;

[0043] (3) The above extract was concentrated under vacuum to 1 / 6 of its original volume, then freeze-dried and pulverized through a 100-mesh sieve to obtain extract B.

[0044] 3. Microorganisms:

[0045] Microorganism A: Bacillus amyloliquefaciens; Microorganism B: Bacillus amyloliquefaciens and red yeast are mixed in equal quantities.

[0046] 2. Preparation of antibacterial and fresh-keeping agents:

[0047] (1) Add 12 times the volume of distilled water to the mixture of wild rice flour and konjac flour, then mix with microorganisms, culture on a shaker at 100 r / min and 30±2°C for 10 h, then mix with carboxymethyl chitosan and stir evenly to obtain a preliminary mixture for use;

[0048] (2) Add the extract and sodium alginate to the preliminary mixture, maintain the water bath temperature at 40°C, stir at 400 r / min for 12 min, and then freeze-dry to obtain an antibacterial preservative agent.

[0049] According to Table 1 below, different weight portions of raw materials are set and different antibacterial fresh-keeping agents are prepared by referring to the above preparation method:

[0050] Table 1

[0051] Embodiment 2:

[0052] Sugar orange preservation experiment:

[0053] 1. Material preparation:

[0054] Sugar oranges were purchased from the sugar orange orchard in Ailao Mountain, Yunnan (same maturity); the same batch of sugar oranges with the same maturity were selected as experimental materials; Phytophthora citrophthora ) was isolated from diseased rock sugar orange.

[0055] 2. Detect the antibacterial effect of each group of bacterial agents on citrus brown rot fungus:

[0056] (1) 35 rock sugar oranges of the same size, color and maturity were selected and randomly divided into 7 groups, namely experimental groups 1-7, with 5 oranges in each group. The rock sugar oranges in each group were washed with distilled water, and then the surface of the fruit was wiped with 75% alcohol for disinfection. The oranges were placed at room temperature for 2 hours before use;

[0057] (2) Activation treatment of citrus brown rot Phytophthora: Inoculate citrus brown rot Phytophthora onto PDA medium, culture at 30°C, then rinse with sterile water to collect spores and prepare 1×10 5 CFU / mL of citrus brown rot Phytophthora spore suspension for later use;

[0058] (3) Use a sterilized punch needle to randomly punch a hole (4 mm in diameter and approximately 4 mm in depth) at the equator of each sugar orange in each group, and inoculate each hole with 15 μL of spore suspension of Phytophthora citri citri;

[0059] (4) Dilution of bacterial agents with warm water: dilute each group of bacterial agents 40 times with 35°C warm water and keep them at 35°C for 80 minutes for later use;

[0060] Dilution of bacterial agent at room temperature: Add bacterial agent 1 to clean water at room temperature and dilute it 40 times for later use;

[0061] (5) The punched rock sugar oranges of each group were soaked in the diluted bacterial agent for 4 minutes, then taken out, naturally air-dried, and then stored at a temperature of 20±2°C. The average lesion diameters after 3d and 7d of storage were recorded. The specific results are shown in Table 2 below:

[0062] Table 2

[0063]

[0064] The growth of lesions in the sugar oranges of experimental groups 1-7 after storage for 7 days is as follows: Figure 1-7 As shown, the lesions in experimental group 1 and experimental group 2 were well inhibited.

[0065] 3. Detect the preservation effect of each group of bacterial agents on rock sugar orange:

[0066] (1) 35 rock sugar oranges of the same size, color and maturity were selected and randomly divided into 7 groups, namely experimental groups 8-14, with 5 oranges in each group. The surface of the fruit was then wiped with 75% alcohol for disinfection and placed at room temperature for 2 hours for later use;

[0067] (2) Dilution of bacterial agents with warm water: dilute each group of bacterial agents 40 times with 35°C warm water and keep them at 35°C for 80 minutes for later use;

[0068] Dilution of bacterial agent at room temperature: Add bacterial agent 1 to clean water at room temperature and dilute it 40 times for later use;

[0069] (3) Each group of rock sugar oranges was placed in the diluted bacterial agent and soaked evenly for 4 minutes, then taken out and naturally air-dried, and then weighed. The initial weight was recorded, and then stored in a cold storage at 10±2℃. The weights were weighed at 7d, 14d, 21d, and 28d of storage, and the weight loss rate was calculated (weight loss rate = (initial weight - current weight) / initial weight × 100%). The specific results are shown in Table 3 below:

[0070] Table 3

[0071]

[0072] From the above tests, it can be seen that experimental groups 8 and 9 have better preservation effects and effectively alleviate the weight loss of sugar oranges.

[0073] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features may be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A post-harvest antibacterial and fresh-keeping agent for sugar oranges, characterized in that: The antibacterial and fresh-keeping agent is prepared from the following raw materials in parts by weight: 4-8 parts of carboxymethyl chitosan, 2-4 parts of sodium alginate, 2-4 parts of wild rice stem flour, 1-2 parts of konjac flour, 0.1-0.3 parts of walnut peel fermentation extract, and 0.01-0.03 parts of Bacillus amyloliquefaciens; The rice stem cooked powder is prepared by washing fresh rice stem, steaming it at 100-110°C for 15-30 minutes, drying it, and then grinding it through a 100-mesh sieve. The walnut green peel fermentation extract is prepared by fermenting the walnut green peel with red yeast; and the preparation method comprises the following steps: (1) Add water to the walnut green peel and boil for 3-5 minutes, then take it out and add 1-2 times distilled water to grind it into slurry, and the slurry is set aside; (2) Inoculate 0.1%-0.2% of the total mass of walnut peel with red yeast into the above slurry, ferment for 18-24 hours, then press and filter, separate the filtrate and the residue, add 2-4 times of distilled water to the residue, stir evenly, let stand for 2-4 hours, centrifuge again, separate the supernatant, combine the supernatant and the filtrate, and obtain the extract for later use; (3) The extract is vacuum concentrated to 1 / 5-1 / 8 of its original volume, then freeze-dried and crushed through a 100-mesh sieve to obtain a walnut green peel fermentation extract.

2. The antibacterial and fresh-keeping agent for postharvest sugar orange according to claim 1, characterized in that: The viable count of the red yeast is 1×10 7 -2×10 9 Pieces / g.

3. The antibacterial and fresh-keeping agent for post-harvest sugar orange according to claim 1, characterized in that: The viable bacterial count of the Bacillus amyloliquefaciens is 1×10 7 -2×10 9 Pieces / g.

4. A method for preparing the antibacterial fresh-keeping agent according to any one of claims 1 to 3, characterized in that: The preparation method comprises the following steps: S1, the wild rice flour mixed with konjac flour was added to 10-15 times the volume of distilled water, and then mixed with Bacillus amyloliquefaciens, and cultured on a shaking table for 8-12h, and then mixed with carboxymethyl chitosan and stirred evenly to obtain a preliminary mixture for standby use; S2. Add walnut green peel fermentation extract and sodium alginate to the above preliminary mixture, stir evenly and freeze-dry to obtain an antibacterial and fresh-keeping bacterial agent.

5. The preparation method according to claim 4, characterized in that: In step S1, the rotation speed of the shaking table culture is 80-120 r / min, and the culture temperature is 30±2°C.

6. The preparation method according to claim 4, characterized in that: During the stirring in step S2, the water bath temperature is maintained at 40-45° C., the stirring time is 10-15 min, and the stirring speed is 120-800 r / min.

7. A method for using the antimicrobial fresh-keeping agent according to any one of claims 1 to 3, characterized in that: The method of use is to add the antibacterial and fresh-keeping agent into warm water at 30-40°C and dilute it 30-50 times to obtain an antibacterial solution, keep the antibacterial solution warm for 1-2 hours, soak the sugar orange for 3-5 minutes, take it out and air dry it naturally, or directly spray the antibacterial solution on the surface of the sugar orange and air dry it naturally.

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