APPLE ENZYME FOR EFFICIENT RECONSTRUCTION OF INTESTINAL MICROECOLOGY AND PROCESSING TECHNOLOGY

Abstract

The present invention relates to an apple enzyme for efficient reconstruction of an intestinal microecology and a processing technology, which comprises the following steps: (1) pretreating raw materials; (2) pulping; (3) detoxifying patulin: firstly biologically detoxifying the pulp: inoculating inoculating plant lactobacillus (ATCC 8014) into the pulp, adjusting the pH to 3-7 at the temperature of 20-30° C., stirring, and detoxifying for 20-24 h; and then repeatedly absorbing and filtering with a carboxylation nano multiwall carbon-neutral aluminum oxide filter screen for three times; and (4) carrying out the division intensified and dynamic fermentation. For the problems of the apple enzymes such as low fermentation efficiency and long time, the division decompression dynamic intensified fermentation method of substrates is used, so that the high-speed and high-efficient fermentation of the apple can be realized, and the fermentation time can be greatly shortened.

Claims

1. A processing method of an apple enzyme for efficient reconstruction of an intestinal microecology, comprising the following steps: (1) pretreating raw materials removing molded parts of post-floral fallen fruits and artificially-thinned young fruits, washing ash and soil on the surfaces of the fruits, reserving original wild yeasts of pericarp, removing cores and dicing; (2) pulping exhausting oxygen with low-temperature liquid nitrogen, pulping, reducing browning of pulp, and reserving the activity of a majority of enzymes; controlling the low temperature at 5-10° C., wherein the dripping amount of the liquid nitrogen is 2%-3.5%; after the pulping, carrying out the enzymolysis by appropriately reducing the pressure, adding 50 mg/L-70 mg/L of pectinase and 85 mg/L-100 mg/L of cellulase, and stirring for 1-2 h at 40-45° C.; (3) detoxifying patulin firstly biologically detoxifying the pulp: inoculating plant lactobacillus (ATCC 8014) into the pulp, adjusting the pH to 3-7 at the temperature of 20-30° C., stirring, and detoxifying for 20-24 h; and then repeatedly absorbing and filtering with a carboxylation nano multiwall carbon-neutral aluminum oxide filter screen for three times; (4) carrying out the division intensification and dynamic fermentation adding an enzyme keep-alive agent into the detoxified apple pulp, wherein the enzyme keep-alive agent is composed of trehalose, mannitol and cysteine in a weight ratio of (1-2):(1-1.5):(1-2); and then equally dividing the apple pulp into five portions for separate intensive fermentation: fermenting with saccharomycetes, Lactobacillus plantarum, Lactobacillus acidophilus, probiotic bacillus, bifidobacterium and Clostridium butyricum; supplying oxygen to the enzyme solution fermented under an anaerobic condition for 1-2 hours every two days, and stopping the oxygen supply to the enzyme solution fermented under aerobic and facultative anaerobic conditions for 2-3 hours every two days; at the same time, carrying out the temperature-varied exercise at a rate of ±2-4° C. every day to improve the stress resistance of the flora and to improve the strength of probiotics; and maintaining appropriate low pressure in the entire fermentation process, introducing the nitrogen to disturb the fermentation broth for 0.5-1 h every day, then returning the pressure, and re-suspending sediments, thereby improving the fermentation efficiency; (5) precisely intensifying and mixing the probiotics: mixing the six fermentation broths in the above steps for use; (6) filtering: carrying out three-level filtration to remove suspended solids and undecomposed residues on the surface: {circle around (1)} fermenting with saccharomycetes: adding fruit Saccharomyces cerevisiae (ATCC 9080) into the fermentation broth, and fermenting for 12 days at 28-30° C.; {circle around (2)}, fermenting with Lactobacillus plantarum: adding 1%-1.5% of albumen powder into the fermentation broth, and inoculating plant lactobacillus (ATCC 8014), followed by anaerobic fermentation for one week at 36-40° C.; {circle around (3)} fermenting with Lactobacillus acidophilus: adding 2-3% of lactose and 1-1.5% of fructo-oligose into the fermentation broth, and inoculating Lactobacillus acidophilus (AS 1.1854), followed by oxygen-free fermentation for one week at 37° C.; {circle around (4)} fermenting with probiotic bacillus: adding 1%-1.5% of albumen powder and 2-2.5% of starch into the fermentation broth, and inoculating probiotic bacillus (CGMCC1.3358), followed by aerobic fermentation for one week at 37° C.; {circle around (5)} fermenting with bifidobacterium: adding 2-3% of oligosaccharide and 1-1.5% of fructo-oligose into the fermentation broth, and inoculating bifidobacterium (ATCC 15700), followed by the anaerobic fermentation for one week at 37° C.; {circle around (6)} fermenting with Clostridium butyricum: adding 1-1.5% of lactose into the fermentation broth, and inoculating Clostridium butyricum (ATCC 19398), followed by the anaerobic fermentation for one week at 37° C.; (7) adding a flavoring, agent, blending and stirring adding 3-4 wt % of sucrose, 1-3 wt % of honey, 1-2 wt % of brown sugar, and 1-1.5 wt % of dark brown sugar, and fully and uniformly stirring; (8) chelating and aging in a dark place sealing the blended apple enzymes, standing in a dark place, chelating and aging for 1 to more than 6 months, aging for one month to obtain the low-quality and medium-quality apple enzymes, and aging for more than six months to obtain the high-quality apple enzymes; (9) exhausting the air regularly: opening a cover for air exhaustion every 15 days during the later aging period; and (10) filtering: filtering again before obtaining the finished product.

2. The processing method of the apple enzyme for efficient reconstruction of the intestinal microecology according to claim 1, wherein a method for precisely intensifying and mixing the probiotics in step (5) comprises the following steps: precisely intensifying and mixing the probiotics for different target groups; mixing the saccharomycetes fermentation broth, the Lactobacillus plantarum fermentation broth, the Lactobacillus acidophilus fermentation broth, the probiotic bacillus fermentation broth, the bifidobacterium fermentation broth and the Clostridium butyricum fermentation broth in a ratio of 1:1:1:1:1:1 for general population, in a ratio of 1:2:2:2:2:1 for people having intestinal microecological disorders caused by antibiotics, in a ratio of 1:1:2:1:2:1 for people having intestinal microecological disorders caused by high-fat diet, in a ratio of 1:1:2:1:2:1 for people having the intestinal microecological disorders caused by the psychological stress and physical stress and in a ratio of 1:2:2:1:2:2 for people having the intestinal microecological disorders caused by advanced ages.

3. The processing method of the apple enzyme for efficient reconstruction of the intestinal microecology according to claim 1, wherein a structure of the carboxylation nano multiwall carbon-neutral aluminum oxide filter screen is that the filter screen has a five-layer structure, wherein a first layer is a sieve plate with an aperture of 5-10 meshes (the diameter is about 2-4 mm); a second layer is carboxylation nano multiwall carbon with a particle size of 3-5 nm; a third layer is macroporous adsorption resin; a fourth layer is neutral aluminum oxide with a particle size of 50-70 μm; a fifth layer is also a sieve plate with an aperture of 5-10 meshes; and the pulp containing solute with a particle size not greater than 3 mm can pass through the adsorption filter screen.

4. The processing method of the apple enzyme for efficient reconstruction of the intestinal microecology according to claim 1, wherein a fermentation process using the saccharomycetes, Lactobacillus plantarum, Lactobacillus acidophilus, probiotic bacillus, bifidobacterium and Clostridium butyricum comprises the following steps: {circle around (1)} fermenting with saccharomycetes: adding fruit Saccharomyces cerevisiae (ATCC 9080) into the fermentation broth, and fermenting for 12 days at 28-30° C.; {circle around (2)} fermenting with Lactobacillus plantarum: adding 1%-1.5% of albumen powder into the fermentation broth, and inoculating plant lactobacillus (ATCC 8014), followed by anaerobic fermentation for one week at 36-40° C.; {circle around (3)} fermenting with Lactobacillus acidophilus: adding 2-3% of lactose and 1-1.5% of fructo-oligose into the fermentation broth, and inoculating Lactobacillus acidophilus (AS 1.1854), followed by oxygen-free fermentation for one week at 37° C.; {circle around (4)} fermenting with probiotic bacillus: adding 1%-1.5% of albumen powder and 2-2.5% of starch into the fermentation broth, and inoculating probiotic bacillus (CGMCC1.3358), followed by the aerobic fermentation for one week at 37° C.; {circle around (5)} fermenting with bifidobacterium: adding 2-3% of oligosaccharide and 1-1.5% of fructo-oligose into the fermentation broth, and inoculating bifidobacterium (ATCC 15700), followed by the anaerobic fermentation for one week at 37° C.; {circle around (6)} fermenting with Clostridium butyricum: adding 1-1.5% of lactose into the fermentation broth, and inoculating Clostridium butyricum (ATCC 19398), followed by the anaerobic fermentation for one week at 37° C.

Description

DESCRIPTION OF THE DRAWINGS

[0058] FIG. 1 shows, an apple enzyme fermentation broth.

DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS

[0059] The present invention is further described below through specific embodiments. The following embodiments are only descriptive rather than limiting, and shall not be used to limit the protection scope of the present invention.

[0060] Apple enzymes for efficient reconstruction of an intestinal microecology and a processing technology, includes the following steps:

[0061] (1) Pretreatment of Raw Materials.

[0062] Molded parts of post-floral fallen fruits and artificially-thinned young fruits are removed, ash and soil on the surfaces of the fruits are washed, original wild yeasts of pericarp are reserved, and the fruits are diced after cores are removed.

[0063] (2) Pulping

[0064] Oxygen is exhausted by low-temperature liquid nitrogen, and pulping is carried out, so that the browning of the pulp is reduced, and the activity of a majority of enzymes can be maintained. The low temperature is controlled at 5-10° C., and the dripping amount of the liquid nitrogen is 2%-3.5%. After the pulping, the enzymolysis is carried out by appropriately reducing the pressure, and 50 mg/L-70 mg/L of pectinase and 85 mg/L-100 mg/L of cellulase are added, followed by stirring for 1-2 h at 40-45° C.;

[0065] (3) Detoxification of Patulin

[0066] Firstly the pulp is biologically detoxified: plant lactobacillus (ATCC 8014) is inoculated into the pulp, followed by stirring and detoxification for 20-24 h under the conditions that the temperature is 20-30° C., and the PH is 3-7. Then a carboxylation nano multiwall carbon-neutral aluminum oxide filter screen is used for repeated adsorption and filtration for three times. The detoxification effect (the detoxification residue is detected) is shown as table 1.

[0067] The structure of the adsorption filter screen is as follows: the filter screen has a five-layer structure, wherein a first layer is a sieve plate with an aperture of 5-10 meshes (the diameter of about 2-4 mm); a second layer is carboxylation nano multiwall carbon with a particle size of 3-5 nm; a third layer is macroporous adsorption resin; a fourth layer is neutral aluminum oxide with a particle size of 50-70 μm; and a fifth layer is also a sieve plate with an aperture of 5-10 meshes. The pulp containing solute with a particle size not greater than 3 mm can pass through the adsorption filter screen.

[0068] (4) Division Intensification and Dynamic Fermentation

[0069] An enzyme keep-alive agent is added into the detoxified apple pulp, wherein the enzyme keep-alive agent is composed of trehalose, mannitol and cysteine in a weight ratio of (1-2):(1-1.5):(1-2). The apple pulp is equally divided into five portions which are subjected to separate intensified fermentation.

[0070] {circle around (1)} Fermentation with saccharomycetes: fruit Saccharomyces cerevisiae (ATCC 9080) is added into the fermentation broth, followed by fermentation for 12 days at 28-30° C.;

[0071] {circle around (2)} Fermentation with Lactobacillus plantarum: 1%-1.5% of albumen powder is added into the fermentation broth, and plant lactobacillus (ATCC 8014) is inoculated, followed by temperature-preservation anaerobic fermentation for one week at 36-40° C.;

[0072] {circle around (3)} Fermentation with Lactobacillus acidophilus: 2-3% of lactose and 1-1.5% of fructo-oligose are added into the fermentation broth, and Lactobacillus acidophilus (AS 1.1854) is inoculated, followed by oxygen-free fermentation for one week at 37° C.;

[0073] {circle around (4)} Fermentation with probiotic bacillus: 1%-1.5% of albumen powder and 2-2.5% of starch is added into the fermentation broth, and probiotic bacillus (CGMCC1.3358) is inoculated, followed by aerobic fermentation for one week at 37° C.;

[0074] {circle around (6)} Fermentation with bifidobacterium: 2-3% of oligosaccharide and 1-1.5% of fructo-oligose are added into the fermentation broth, and bifidobacterium (ATCC 15700) is inoculated, followed by the anaerobic fermentation for one week at 37° C.;

[0075] {circle around (7)} Fermentation with Clostridium butyricum: 1-1.5% of lactose is added into the fermentation broth, and Clostridium butyricum (ATCC 19398) is inoculated, followed by anaerobic fermentation for one week at 37° C.;

[0076] The enzyme solution fermented under an anaerobic condition is oxygenated for 1-2 h every two days, and the oxygen supply to the enzyme solution fermented under aerobic and facultative anaerobic conditions is stopped for 2-3 h every two days. Meanwhile, the temperature-varied exercise of ±2-4° C. is carried out every day, thereby improving the stress resistance of the bacterial flora, and improving the strength of the probiotics. The appropriate low pressure is kept in the entire fermentation process, and the pressure is returned after the fermentation broth is disturbed for 0.5-1 h by introducing the nitrogen every day, so that the sediments are re-suspended, and the fermentation efficiency can be improved.

[0077] (5) Precise Intensification and Mixing of Probiotics

[0078] The concentrated probiotics are precisely intensified and mixed for different target groups. The saccharomycetes fermentation broth, the Lactobacillus plantarum fermentation broth, the Lactobacillus acidophilus fermentation broth, the probiotic bacillus fermentation broth, the bifidobacterium fermentation broth and the Clostridium butyricum fermentation broth are mixed in a ratio of 1:1:1:1:1:1 for general population, in a ratio of 1:2:2:2:2:1 for people having intestinal microecological disorders caused by antibiotics, in a ratio of 1:1:2:1:2:1 for people having intestinal microecological disorders caused by high-fat diet, in a ratio of 1:1:2:1:2:1 for people having the intestinal microecological disorders caused by the psychological stress and physical stress and in a ratio of 1:2:2:1:2:2 for people having the intestinal microecological disorders caused by advanced ages.

[0079] (6) Filtration

[0080] By virtue of three-level filtration, suspended solids and undecomposed residues on the surface are removed.

[0081] (7) Flavoring Agent is Added, Followed by Blending and Stirring

[0082] 3-4 wt % of sucrose, 1-3 wt % of honey, 1-2 wt % of brown sugar, and 1-1.5 wt % of dark brown sugar are added and fully and uniformly stirred.

[0083] (8) Chelating and Aging in a Dark Place

[0084] The blended apple enzymes are sealed, and stand at a dark place for chelating and aging for 1 to more than six months. The apple enzymes are aged for one month to obtain the low-quality and medium-quality apple enzymes, and aged for more than six months to obtain high-quality apple enzymes.

[0085] (9) Regular Air Exhaustion

[0086] A cover is opened for air exhaustion every 15 days during the later aging period.

[0087] (10) Filtration

[0088] The filtration is carried out again before a finished product is obtained to remove the suspended solids and undecomposed residues on the surface as well as dead valueless probiotics flora and excrement deposited on the bottom.

TABLE-US-00001 TABLE 1 Detection of patulin residue Adsorption with carboxylation Detoxification nano multi wall Combined Apple raw with plant carbon and detoxifi- Method stock lactobacillus aluminum oxide cation Detected >75-90 4.6-5.7 24-33 0.9-1.5 amount of μg/kg μg/kg μg/kg μg/kg residues Note: the health standard for the limited quantity of patulin in China is not greater than 50 μg/kg.

TABLE-US-00002 TABLE 2 Contrast of effective components (content of the component in every 100 g of enzymes Antibiotic High-fat Psychological disorder obesity stress and Advanced-age Main Common treatment treatment physical stress people component enzymes type type treatment type healthcare type Energy (KJ) 313 307 177 334 297 Fat (g) 2.3 2.2 0.5 2.2 2.1 Carbohydrate (g) 13.7 14.1 5.9 19.9 11.4 Proteins (g) 0.6 1.1 0.9 3.3 2.9 Dietary fibers (g) 1.2 0.7 2.9 1.1 1.1 Vitamin C (mg) 3.7 4.4 4.2 4.8 4.3 Vitamin E (mg) 0.19 0.19 0.15 0.29 0.19 Vitamin D (mg) 0 0 0.3 0.44 0.23 Vitamin B (mg) 0.05 3.3 1.7 0.03 1.8 Calcium Ca (mg) 3.3 3.4 2.9 5.5 5.4 Folic acid 3.5 3.9 3.5 3.1 3.3 (microgram) Pantothenic acid 0.03 0.05 0.03 0.04 0.97 (microgram)