Cordyceps militaris fermentation extract and application thereof in preservation of agaricus bisporus

Abstract

The invention provides a cordyceps militaris fermentation extract and application thereof in preservation of agaricus bisporus, and belongs to the technical field of deep processing of edible mushrooms and preservation of agricultural products. The preparation method of the cordyceps militaris fermentation extract comprises the following steps: (1) taking cordyceps militaris sporocarp, adding water, grinding, and performing enzymolysis by using cellulase and papain in sequence to obtain cordyceps militaris pulp; (2) adding glucose and waste beer yeast powder into the cordyceps militaris pulp, then inoculating lactobacillus paracasei and bifidobacterium bifidum, and fermenting; and (3) after fermentation is completed, carrying out ultrasonic crushing, centrifuging to remove thalli, and carrying out spray drying to obtain the cordyceps militaris fermentation extract. The method can effectively reduce the browning index and weight loss ratio of agaricus bisporus, can slow down the increase of malonaldehyde, reduces the activity of PPO enzyme, and prevents enzymatic browning caused by the synergistic effect of the PPO enzyme and peroxidase. The SOD enzyme activity in the agaricus bisporus can be kept, active oxygen in tissues can be removed, and the metabolic balance of the active oxygen can be maintained.

Description

Technical Field

[0001] The present invention belongs to the technical field of deep processing of edible fungi and preservation of agricultural products, and particularly relates to a fermented extract of Cordyceps militaris and its application in the preservation of Agaricus bisporus. Background Art

[0002] Agaricus bisporus (scientific name: Agaricus bisporus (Large) Sing.) is a fungus of the family Agaricaceae and the genus Agaricus. Due to its high post-harvest respiratory intensity and strong polyphenol oxidase (PPO) activity, it will turn brown, open its umbrella, and the stipe will elongate within 24 hours at room temperature, deteriorate quickly, and is difficult to preserve. Currently, the commonly used preservation methods include chemical preservation, physical preservation, and biological preservatives. Among them, chemical preservation usually uses sulfite soaking, which is likely to cause SO 2 residue. The cost of physical preservation (such as 1-MCP treatment) is relatively high. Existing biological preservatives such as tea polyphenols require a high concentration, which will affect the flavor of Agaricus bisporus. Therefore, there is a need to obtain a preservation method without residue and without affecting the flavor of Agaricus bisporus.

[0003] Cordyceps militaris (scientific name: Cordyceps militaris (L.ex Fr.) Link.) is a fungus of the family Clavicipitaceae and the genus Cordyceps, also known as Cordyceps sinensis var. militaris, Cordyceps militaris, simply called Cordyceps. Cordyceps militaris is a complex composed of a stroma (i.e., the grass part) and a sclerotium (i.e., the body part of the insect). Because it contains rich cordycepic acid, cordycepin, ergosterol and other substances, it has the effects of expanding the trachea, sedation, resisting various bacteria, and lowering blood pressure. It is widely used in the fields of medicine, health products, cosmetics, etc. Summary of the Invention

[0004] The purpose of the present invention is to provide a fermented extract of Cordyceps militaris that can be used for the preservation of Agaricus bisporus.

[0005] In order to achieve the above-mentioned invention purpose, the present invention provides the following technical solutions:

[0006] The present invention provides a fermented extract of Cordyceps militaris, and the preparation method is as follows:

[0007] (1) Take the fruiting body of Cordyceps militaris, grind it, add water, and then enzymatically hydrolyze it with cellulase and papain in sequence to obtain Cordyceps militaris pulp;

[0008] (2) Add glucose and waste beer yeast powder to the Cordyceps militaris pulp, then inoculate Lactobacillus paracasei and Bifidobacterium bifidum, and ferment;

[0009] (3) After fermentation is completed, ultrasonically disrupt, centrifuge to remove the thalli, and spray dry to obtain the fermented extract of Cordyceps militaris.

[0010] Preferably, in step (1), the mass-to-volume ratio of Cordyceps militaris fruiting bodies to water is 1 g: 3-5 mL;

[0011] The addition amount of the cellulase is (3-5)×10 6 U / L, the temperature of enzymatic hydrolysis is: 50-55 °C, the time is: 3-4 h, and the pH is 4.5-5.5.

[0012] Preferably, the addition amount of the papain is (5-10)×10 6 U / L, the temperature of enzymatic hydrolysis is: 50-55 °C, the time is: 3-4 h, and the pH is 6.5-7.5.

[0013] Preferably, in step (2), the addition amount of glucose is 50-100 g / L, and the addition amount of waste beer yeast powder is 400-500 g / L.

[0014] Preferably, the inoculation amount of Lactobacillus paracasei is (3-5)×10 6 CFU / L, and the inoculation amount of Bifidobacterium bifidum is (3-5)×10 6 CFU / L;

[0015] The fermentation conditions are: 25-30 °C, fermenting until the pH is 4.0-4.2.

[0016] Preferably, in step (3), the conditions of ultrasonic treatment are 150-250 w, the ultrasonic time is 3-5 s, the interval is 10-15 s, and the total duration is 10-15 min;

[0017] The conditions of centrifugation are 8000-12000×g, 10-15 min, 4-6 °C.

[0018] The present invention also provides the application of the fermentation extract in the preservation of Agaricus bisporus.

[0019] Preferably, when using the fermentation extract for preservation, the usage method of the fermentation extract is: dissolving the fermentation extract in water according to the mass-to-volume ratio of 1 g: 10-15 mL to obtain a preservation solution, immersing the Agaricus bisporus in the preservation solution for 5-10 min, and after air-drying until there is no obvious water stain on the surface of the Agaricus bisporus, packing it into a preservation bag and storing it sealed.

[0020] In the present invention, small molecules such as cordycepin, cordycepic acid, and small molecule polypeptides of Cordyceps militaris in the Cordyceps militaris fermentation extract have high antioxidant activity. At the same time, it can also produce beneficial substances, which are beneficial to the fermentation of lactic acid bacteria such as Lactobacillus paracasei and Bifidobacterium bifidum, and produce various antibacterial substances such as organic acids, hydrogen peroxide, or bacteriocins. Through experiments, it is found that the present invention can effectively reduce the browning index and weight loss rate of Agaricus bisporus, can slow down the increase of malondialdehyde, reduce the activity of PPO enzyme, and prevent its synergistic effect with peroxidase to cause enzymatic browning. The present invention can maintain the activity of SOD enzyme in Agaricus bisporus, can scavenge reactive oxygen species in tissues, maintain the balance of reactive oxygen species metabolism, and thus play a role in delaying the post-harvest quality deterioration and senescence process of Agaricus bisporus. Detailed implementation manners

[0021] In the embodiments of the present invention, Lactobacillus paracasei LPC-37 was purchased from Shanghai Ruiyingying Biotechnology Co., Ltd.; Bifidobacterium bifidum HC2273 was purchased from Wellcare Health (Shandong) Biotechnology Co., Ltd.; the cellulase was (Novozymes).

[0022] The technical solutions provided by the present invention will be described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0023] Example 1

[0024] A Cordyceps militaris fermentation extract, the preparation method is as follows:

[0025] (1) Take Cordyceps militaris fruiting bodies, grind them with water according to the mass-volume ratio of 1 g:5 mL, then add cellulase (5×10 6 U / L), enzymolysis at 55°C, the time is: 3.5 h, pH is 4.5, and then add papain (5×10 6 U / L) for enzymolysis (50°C, the time is: 3.5 h, pH is 7.0) to obtain Cordyceps militaris slurry;

[0026] (2) Add 100 g / L glucose and 400 g / L waste beer yeast powder to the Cordyceps militaris slurry, and then inoculate Lactobacillus paracasei and Bifidobacterium bifidum respectively with an inoculation amount of 3×10 6 CFU / L, and ferment at 28°C until the pH is 4.2;

[0027] (3) After fermentation, ultrasonicate at 200 w for 5 s, with an interval of 10 s, and ultrasonicate intermittently for a total of 10 min. Centrifuge at 10000×g for 10 min at 4°C to remove the bacteria, and then spray dry to obtain the Cordyceps militaris fermentation extract.

[0028] When in use, dissolve the fermentation extract in water according to the mass-volume ratio of 1 g:10 mL to obtain a fresh-keeping solution.

[0029] Example 2

[0030] A Cordyceps militaris fermentation extract, the preparation method is as follows:

[0031] (1) Take Cordyceps militaris fruiting bodies, grind them with water according to the mass-volume ratio of 1 g:5 mL, then add cellulase (3×10 6 U / L), at 55 °C, enzymolysis for 4 h, pH is 5, then add papain (7×10 6 U / L) for enzymolysis (at 55 °C, the time is: 3 h, pH is 6.5) to obtain Cordyceps militaris pulp;

[0032] (2) Add 50 g / L glucose and 450 g / L waste beer yeast powder to the Cordyceps militaris pulp, then inoculate Lactobacillus paracasei and Bifidobacterium bifidum respectively with an inoculum size of 4×10 6 CFU / L, ferment at 28 °C until the pH is 4.2;

[0033] (3) After fermentation, use ultrasound at 200 w for 3 s, intermittent for 15 s, with a total intermittent ultrasound of 15 min, centrifuge at 10000×g for 10 min at 4 °C to remove the bacteria, and spray dry to obtain the Cordyceps militaris fermentation extract.

[0034] When in use, dissolve the fermentation extract in water according to the mass-volume ratio of 1 g:15 mL to obtain a fresh-keeping solution.

[0035] Example 3

[0036] A Cordyceps militaris fermentation extract, the preparation method is as follows:

[0037] (1) Take Cordyceps militaris fruiting bodies, grind them with water according to the mass-volume ratio of 1 g:5 mL, then add cellulase (4×10 6 U / L), enzymolysis at 55 °C, the time is: 3.5 h, pH is 4.5, then add papain (6×10 6 U / L) for enzymolysis (at 50 °C, the time is: 4 h, pH is 6.5) to obtain Cordyceps militaris pulp;

[0038] (2) Add 100 g / L glucose and 500 g / L waste beer yeast powder to the Cordyceps militaris pulp, then inoculate Lactobacillus paracasei and Bifidobacterium bifidum respectively with an inoculum size of 5×10 6 CFU / L, ferment at 28 °C until the pH is 4.2;

[0039] (3) After fermentation, use ultrasound at 200 w for 5 s, intermittent for 10 s, with a total intermittent ultrasound of 10 min, centrifuge at 10000×g for 10 min at 4 °C to remove the bacteria, and spray dry to obtain the Cordyceps militaris fermentation extract.

[0040] When in use, dissolve the fermentation extract in water according to a mass-volume ratio of 1 g: 10 mL to obtain a fresh-keeping solution.

[0041] Comparative Example 1

[0042] A fermented extract of Cordyceps militaris, different from Example 3, was fermented without adding Lactobacillus paracasei and Bifidobacterium bifidum.

[0043] Comparative Example 2

[0044] A fermented extract of Cordyceps militaris, different from Example 3, used Lactobacillus plantarum to equivalently replace the combination of Lactobacillus paracasei and Bifidobacterium bifidum.

[0045] Test Example

[0046] Select post-harvest fruiting bodies of Agaricus bisporus with regular shapes, 3-5 cm in mushroom caps, uniform sizes, white colors, and no mechanical damage. Divide them into 5 groups, with 5 kg in each group. After treating each group of Agaricus bisporus as follows, take them out and naturally dry them until there is no obvious water stain on the surface of the Agaricus bisporus, then put them into fresh-keeping bags and place them in an environmental mushroom house at 17 ± 1 °C and a relative humidity of 85%-90%. Samples were taken on the 0th day, the 2nd day, and the 6th day to measure the browning index of Agaricus bisporus (measured by the spectrophotometric colorimeter method), weight loss rate (weight loss rate = (initial weight - final weight) / initial weight × 100%), malondialdehyde content, and the activities of metabolic enzymes such as PPO (polyphenol oxidase) and SOD (superoxide dismutase) (the detection kits were purchased from Shanghai Jiwei Biotechnology Co., Ltd.). The results are shown in Table 1 and Table 2:

[0047] Blank treatment group: Soak in distilled water for 8 min;

[0048] Positive control group: Prepare a 1% Na 2 SO 3 (w / v) solution and soak for 8 min;

[0049] Example 3 group: Soak in the fresh-keeping solution prepared in Example 3 for 8 min;

[0050] Comparative Example 1 group: Soak in the fresh-keeping solution prepared in Comparative Example 1 for 8 min;

[0051] Comparative Example 2 group: Soak in the fresh-keeping solution prepared in Comparative Example 2 for 8 min.

[0052] Table 1

[0053]

[0054] Table 2 Effects of different fresh-keeping solutions on the metabolic enzyme activities of Agaricus bisporus

[0055]

[0056] As can be seen from the above embodiments, the present invention uses Cordyceps militaris fermentation extract as a fresh-keeping solution for Agaricus bisporus, which can effectively reduce the browning index and weight loss rate of Agaricus bisporus, slow down the increase of malondialdehyde, reduce the activity of PPO enzyme, and prevent the synergistic effect with peroxidase to cause enzymatic browning. The present invention can maintain the activity of SOD enzyme in Agaricus bisporus, scavenge reactive oxygen species in tissues, maintain the balance of reactive oxygen species metabolism, and thus play a role in delaying aging.

[0057] The above description is only a preferred embodiment of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. A Cordyceps militaris fermentation extract, characterized in that: The preparation method of the Cordyceps militaris fermentation extract is: (1) taking Cordyceps militaris fruiting bodies, grinding them with water, and then enzymolyzing them with cellulase and papain in sequence to obtain Cordyceps militaris slurry; (2) adding glucose and waste brewer's yeast powder to Cordyceps militaris slurry, and then inoculating Lactobacillus paracasei and Bifidobacterium bifidum for fermentation; (3) After the fermentation is completed, the cells are ultrasonically disrupted, centrifuged to remove the cells, and spray-dried to obtain the Cordyceps militaris fermentation extract.

2. The fermentation extract according to claim 1, characterized in that In step (1), the mass volume ratio of Cordyceps militaris fruiting body and water is 1 g: 3-5 mL; The amount of cellulase added is (3-5)×10 6 U / L, the enzymatic hydrolysis temperature is 50-55°C, the time is 3-4h, and the pH is 4.5-5.

5.

3. The fermentation extract according to claim 2, characterized in that The amount of papain added is (5-10)×10 6 U / L, the enzymatic hydrolysis temperature is 50-55°C, the time is 3-4h, and the pH is 6.5-7.

5.

4. The fermentation extract according to claim 1, characterized in that In step (2), the amount of glucose added is 50-100 g / L, and the amount of waste beer yeast powder added is 400-500 g / L.

5. The fermentation extract according to claim 4, characterized in that The inoculum size of Lactobacillus paracasei is (3-5)×10 6 CFU / L, the inoculum amount of Bifidobacterium bifidum is (3-5)×10 6 CFU / L; The fermentation conditions are: 25-30°C, fermentation to pH 4.0-4.

2.

6. The fermentation extract according to claim 1, characterized in that The ultrasonic conditions in step (3) are 150-250W, the ultrasonic time is 3-5s, the interval is 10-15s, and the total time is 10-15min; The centrifugation conditions are 8000-12000×g, 10-15 min, 4-6°C.

7. Use of the fermentation extract according to any one of claims 1 to 6 in the preservation of Agaricus bisporus.

8. The use according to claim 7, characterized in that When the fermentation extract is used for preservation, the method for using the fermentation extract is as follows: the fermentation extract is dissolved in water according to a mass volume ratio of 1g:10-15mL to obtain a preservation solution, the Agaricus bisporus is immersed in the preservation solution for 5-10min, and the Agaricus bisporus is dried until there is no obvious water stain on the surface of the Agaricus bisporus, and then the Agaricus bisporus is packed into a preservation bag for sealed storage.

Images