Microbial antioxidant nutrient

Abstract

Microbial antioxidant nutrient obtainable by a manufacturing process characterized by the following steps: 1) cultivating a strain composition consisting of Lactobacillus plantarum, Bifidobacterium breve and Saccharomyces cerevisiae in a composite stock medium to obtain a liquid bacterial composition; and 2) Freeze-drying of the liquid bacterial composition to obtain a composite bacterial powder, which can be used as a microbial antioxidant nutrient.

Description

[0001] The present invention relates to a microbial antioxidant nutrient beneficial to the health of companion animals.

[0002] When cats and dogs are kept as pets in households, they are prone to digestive disorders, unbalanced diets, and low disease resistance due to a limited variety of food. When infected with pathogenic microorganisms, they are highly likely to suffer from diarrhea and skin diseases. Infections caused by pathogenic microorganisms can be treated with antibiotics. While antibiotics have the advantage of quickly curing diarrhea in pets, long-term and excessive use leads to environmental pollution with antibiotic residues, the emergence of drug-resistant strains, and thus a disruption of the microbial balance in the host and endogenous infections in pets.

[0003] In recent years, microbial nutrients have become widespread, which are considered safe and directly supply beneficial exogenous bacteria that regulate the microecological balance in the intestine of animals, inhibit harmful bacteria, strengthen immunity and promote the healthy growth of pets, being a desirable approach to solving the problem outlined above.

[0004] The object of the present invention is to provide a nutrient that promotes health in the animal organism. The nutrient should be obtainable by a simple and cost-effective process.

[0005] The present invention provides a microbial antioxidant nutrient obtainable by a process comprising the following steps: Cultivating a strain composition consisting of Lactobacillus plantarum, Bifidobacterium breve and Saccharomyces cerevisiae in a composite stock medium to obtain a liquid bacterial composition; and Freeze-drying the liquid bacterial composition to obtain a composite bacterial powder.

[0006] The compound bacteria powder can be used as a microbial antioxidant nutrient.

[0007] In a preferred embodiment of the microbial antioxidant nutrient, it is obtainable by a process which, following steps 1 and 2, comprises the following step

[0008] 3) Blending the composite bacterial powder with xylanase, neutral protease, and gamma-aminobutyric acid (GABA) produces an enhanced microbial antioxidant nutrient.

[0009] A nutrient is described herein as a composition which is ingested and processed in the metabolism by animals, in particular cats and / or dogs, for their survival, in particular to improve their immune and health status.

[0010] According to the invention, it can be provided that the individual strains in the strain composition have the following parts by weight: 30 to 65 parts by weight, preferably 40 to 60 parts by weight of Lactobacillus plantarum; 30 to 65 parts by weight, preferably 40 to 60 parts by weight of Bifidobacterium breve; 5 to 25 parts by weight, preferably 10 to 20 parts by weight of Saccharomyces cerevisiae.

[0011] It can also be provided that the composite stock medium preferably comprises the following components, particularly preferably consists of these: 0.2 to 5% by weight, preferably 0.5 to 3.5% by weight molasses, 0.2 to 3% by weight, preferably 0.5 to 2% by weight lactose, 0.2 to 2 wt.%, preferably 0.5 to 1 wt.% tryptone, 0.2 to 4% by weight, preferably 0.5 to 2.5% by weight corn steep liquor, 0.05 to 0.5 wt.%, preferably 0.1 to 0.3 wt.% sodium chloride, 0.005 to 0.05 wt.%, preferably 0.01 to 0.03 wt.% magnesium sulfate, 0.005 to 0.1 wt.%, preferably 0.01 to 0.05 wt.% iron sulfate, 0.05 to 1.5% by weight, preferably 0.2 to 0.8% by weight potassium dihydrogen phosphate, 90 to 98% by weight, preferably 90.82 to 97.68% by weight water.

[0012] If the composite stock medium consists of the components mentioned above, the water content adds up to the sum of the components consisting of 0.2 to 5% by weight, preferably 0.5 to 3.5% by weight molasses, 0.2 to 3% by weight, preferably 0.5 to 2% by weight lactose, 0.2 to 2 wt.%, preferably 0.5 to 1 wt.% tryptone, 0.2 to 4% by weight, preferably 0.5 to 2.5% by weight corn steep liquor, 0.05 to 0.5 wt.%, preferably 0.1 to 0.3 wt.% sodium chloride, 0.005 to 0.05 wt.%, preferably 0.01 to 0.03 wt.% magnesium sulfate, 0.005 to 0.1 wt.%, preferably 0.01 to 0.05 wt.% iron sulfate, and 0.05 to 1.5 wt.%, preferably 0.2 to 0.8 wt.% potassium dihydrogen phosphate, to 100 wt.%.

[0013] Furthermore, it can be provided that the composite stock medium has a pH in the range of 5.2 to 6.2, preferably 5.5 to 6.0.

[0014] A particular embodiment of the present invention further comprises that the cultivation of the strain composition in step 1 is carried out by inoculating a strain composition into the composite stock medium, wherein this inoculation is preferably carried out in a weight ratio of 1:100 to 20:100, preferably 2:100 to 10:100. Preferably, the cultivation is carried out at a temperature in the range of 15 to 38°C, preferably 25 to 35°C, for a period of time in the range of 12 to 48 hours, preferably 16 to 24 hours. Preferably, it can be provided that the recovery of the liquid bacterial composition takes place as soon as the total viable bacteria reach a concentration of 12 billion CFU / ml, preferably 2 billion CFU / ml.The measured value "colony forming unit" (CFU) is widely known in the field and can be measured using conventional methods, for example, by plate counting, which can also be done automatically or semi-automatically. The extraction process here refers to the previously described freeze-drying. The composite bacterial powder is preferably sterilized before use for a period of time in the range of 5 to 60 minutes, preferably 15 to 30 minutes, at a temperature in the range of 95 to 145°C, preferably 115 to 130°C.

[0015] Furthermore, in a preferred embodiment, it can be provided that the weight proportions in a microbial antioxidant nutrient are preferably in the following ranges: 35 to 75, preferably 40 to 60 parts by weight of composite bacterial powder, 10 to 25, preferably 10 to 20 parts by weight of xylanase 10 to 25, preferably 10 to 20 parts by weight of neutral protease and 2.5 to 15, preferably 5 to 10 parts by weight of gamma-aminobutyric acid (GABA).

[0016] Furthermore, in a particularly preferred embodiment, it can be provided that the weight proportions in a microbial antioxidant nutrient are preferably in the following ranges: 40 to 60 parts by weight of composite bacterial powder, 10 to 20 parts by weight of xylanase 10 to 20 parts by weight of neutral protease and 5 to 10 parts by weight of gamma-aminobutyric acid (GABA).

[0017] The present invention provides a microbial antioxidant nutrient beneficial to the health of companion animals, obtainable by the above manufacturing process. Due to the complexity of the composition, it can only be described by the manufacturing process.

[0018] Xylanases are widely known in the scientific community and refer to enzymes that can break down xylans. Xylans are plant heteropolysaccharides. Preferred xylanases include enzymes such as β-xylosidase, among others. These enzymes are widely available commercially, including from Qingdao Vland Biotech Inc.

[0019] Neutral proteases are enzymes that can cleave peptides and / or proteins. Neutral proteases preferentially perform this cleavage in the neutral pH range (pH 5 to 8). Preferred neutral proteases include subtilisin, Aspergillus oryzae neutral proteinase, and others. These enzymes are widely available commercially, including from Novozymes A / S and Wuhan Sunhy Biology Co., Ltd.

[0020] The nutrient according to the invention brings about a surprising improvement in the well-being of pets, especially dogs and cats. The following statements represent assumptions that further explain this surprising effect, but are not limited thereto. The xylanase and neutral protease preferably used in the present invention can, under certain circumstances, promote the digestion and absorption of food ingested by pets and maintain all functions of the internal organs of pets, including: cell repair, anti-inflammatory detoxification, metabolism, improved immunity, energy production, and promoting blood circulation.The GABA used can surprisingly improve phospholipase activity, thereby promoting the energy metabolism of the animal brain, activating cerebral blood flow, increasing oxygen supply, and ultimately restoring brain cell function and improving nerve function. It can inhibit the integration of the brainstem respiratory center, slow the respiratory rate, alleviate heat-related panting in animals under heat stress, increase body weight, and improve survival rate. Furthermore, GABA also has a feeding-promoting effect.

[0021] The strains used in the microbial nutrients of the present invention can surprisingly effectively inhibit the invasion of E. coli and Salmonella into the body, establish a biological barrier in the intestine, reduce the penetration of toxic and harmful substances into pets, maintain the ecological balance in the intestine, and stimulate the body to produce antibacterial agents, enhance immune function, improve disease resistance in pets, and significantly improve diarrhea. Beneficial bacteria promote the intestinal development of pets, increase the activity of digestive enzymes, improve animal food intake, and reduce fecal odor. In addition, the beneficial microorganisms can synthesize various nutrients such as vitamins and amino acids during the preparation process, which can lead to a significant improvement in the coat of pets.

[0022] The technical solution to the problem underlying the present invention surprisingly has the following advantageous effects: 1. The microbial nutrients for pets of the present invention can compete with harmful microorganisms for the attachment site of the intestinal epithelium, establish a dominant flora of beneficial bacteria, inhibit the growth of pathogenic bacteria, avoid intestinal pathological reactions, and reduce the risk of bacterial enteritis. 2. The microbial nutrients for pets of the present invention can decompose, transform, and utilize harmful substances generated by metabolism, reduce the accumulation of toxic substances in pets, and protect the liver; reduce the concentration of ammonia and nitrogen in the pet intestines, thereby reducing the odor of excreta. 3. The microbial nutrients for pets of the present invention can stimulate intestinal mucosal cells to produce various immune factors, thereby improving the disease resistance of pets. 4. The organic acids produced by the microbial nutrients for pets of the present invention can create an acid-base environment in the intestine, which is conducive to the absorption of minerals and other nutrients, and can also produce B vitamins to supplement the nutrients needed by pets. 5. The microbial nutrients for pets of the present invention can increase the adaptability of pets to changes in diet, environment and other conditions, reduce or alleviate pathological reactions, and improve the comprehensive health level of the body. Examples of implementation:

[0023] The invention is described in detail below using several working examples. The respective strains of the microbes used in Working Examples 1 and 2, Lactobacillus plantarum (ACCC 11118), Bifidobacterium breve (ACCC 11054), and Saccharomyces cerevisiae (ACCC 21429), were commercially purchased from the Agricultural Culture Collection of China.

[0024] The NBT reduction method was performed as described in Liu Ruiheng, Fu Shiyu, Zhan Huaiyu. Spectrophotom Determination of Superoxide Anion Radical with Nitroblue Tetrazolium [J]. Journal of Instrumental Analysis, 2008, 27(4), 355-359. Example 1 (1) To prepare a composite stock medium, 1.0 wt% molasses, 1.5 wt% lactose, 0.8 wt% tryptone, 0.8 wt% corn steep liquor, 0.1 wt% sodium chloride, 0.01 wt% magnesium sulfate, 0.02 wt% ferrous sulfate and 0.8 wt% potassium dihydrogen phosphate are dissolved in water, the composite stock medium having a pH of 6.0 and sterilized at 121 °C for 15 minutes.

[0025] The resulting composite stock medium is inoculated with a stock composition in a weight ratio of 8:100, the stock composition consisting of 45 parts by weight of Lactobacillus plantarum, 40 parts by weight of Bifidobacterium breve and 15 parts by weight of Saccharomyces cerevisiae.

[0026] The culture is grown for 16 hours at 30°C, after which a composite bacterial fluid is obtained when the total viable bacteria reach a concentration of 2 billion CFU / ml.

[0027] (2) The composite bacterial liquid obtained in step (1) is then freeze-dried to obtain a composite bacterial powder.

[0028] (3) 50 parts by weight of the composite bacterial powder obtained in step (2) is mixed with 20 parts by weight of xylanase, 20 parts by weight of neutral protease, and 10 parts by weight of GABA to obtain a microbial antioxidant nutrient. The microbial antioxidant nutrient can be packaged and has a total viable bacteria content of ≥1.0×10 10 CFU / ml.

[0029] The following experiments were conducted to ensure the effect of the inventive product regarding food safety and antioxidant functionality. 3. Testing of product efficacy3.1 Medical evidence of efficacy

[0030] Experiments and observations were conducted on the ammonia content in animal excreta, daily weight gain, diarrhea rate, mortality rate, immune organ index, and incidence rate of skin diseases in cats and dogs, which were divided into a control group and an experimental group. - The control group was fed cat food or dog food and received no microbial nutrient. - The test group was fed cat food or dog food. In addition, the cat test group was fed 0.5 g of the microbial nutrient prepared in Example 1 daily, and the dog test group was fed 1.0 g of the microbial nutrient prepared in Example 1 daily.

[0031] The specific test results are presented in Table 1, Table 2 and Table 3. Table 1. Effects on ammonia levels in animal houses, daily weight gain and diarrhea rate Object Cats Cats dogs dogs Comparison group experimental group Comparison group experimental group Number of animals [head] 101 99 102 119 Average ammonia content in the 14,4 6,7 25,2 9,1 Excretions [ppm] Average daily weight gain [g] 5,88 6,24 32,1 35,4 Diarrhea rate [%] 23,7 5,5 15,5 3,4

[0032] As shown in Table 1, the ammonia content and diarrhea rate in the cat group were reduced by 53.47% and 76.8%, respectively, compared with the control group; the ammonia content and diarrhea rate in the dog group were reduced by 63.89% and 71.6%, respectively, compared with the control group. The daily weight gain of the cat group was 6.12% higher than that of the control group, and the daily weight gain of the dog group was 10.28% higher than that of the control group. Furthermore, no animals died in any of the experimental groups. Table 2. Effects on the immune organ index of pets Object Cats Cats dogs dogs Comparison group experimental group Comparison group experimental group Spleen Index[%] 0,1144±0,0042a 0,1385±0,027a 0,1983±0,016a 0,2183±0,026a Thymus index [%] 0,1230±0,0045a 0,1534±0,0023a 0,2214±0,0018a 0,2551±0,0015a

[0033] As shown in Table 2, the spleen and thymus indices of cats and dogs in the experimental group were significantly higher than those of the control group (P<0.01). Table 3. Impact on the incidence rate of skin diseases in companion animals Object Cat comparison group Cat experimental group Dog comparison group Dog test group Number of animals [head] 101 99 102 119 Incidence rate of skin diseases [%] 18,8 0 37,3 0,8

[0034] As shown in Table 3, the incidence rate of skin diseases in the cat experimental group was reduced by 100% compared with the control group; the incidence rate of skin diseases in the dog experimental group was reduced by 97.9% compared with the control group. 3.2 Proof of antioxidant capacity:

[0035] Detection method: Superoxide anion radicals were collected in the PMS-NADH system and detected by the NBT reduction method. Vitamin C (VC), vitamin E (VE), lipoic acid, and epigallocatechin gallate (EGCG) at a final concentration of 5 µg / ml were used as controls. Pure water served as the control, and NBT-free water was used as a blank. The scavenging rate of superoxide anion radicals is expressed by the following formula: Superoxidation radical scavenging rate (%) = [(AControl−ALeak)−(ASample−ALeak)] / AControl×100% Example Superoxide anion radical scavenging rate (%) 50% Microbial Antioxidant Nutrient 81,05 Microbial antioxidant nutrient 90,12 Vitamin C (VC) 5,6 Vitamin E (VE) 3,8 Lipoic acid 0 EGCG 54,18 4. Detection data of the individual components of the microbial antioxidant nutrient for pets (Table 4) Nr. Ingredient Salary 1 Lactic acid 16,2% 2 Fructooligosaccharides 240 mg / 100 g 3 Total amino acids 6,89% 4 Betaine 326 mg / kg 5 VB 952 mg / kg 6 VC 300 mg / kg

[0036] Through the above research data, it is known that the microbial antioxidant nutrient for pets of the present invention can increase the protective barrier function of the intestinal mucosa, significantly reduce the ammonia content in the pen, reduce diarrhea of ​​pets, improve the immunity of pets, reduce the mortality of pets, while neutralizing free radicals, reducing the invasion of external harmful substances, and reducing the incidence of skin diseases in pets. QUOTES CONTAINED IN THE DESCRIPTION

[0000] This list of documents submitted by the applicant was generated automatically and is included solely for the convenience of the reader. This list is not part of the German patent or utility model application. The DPMA assumes no liability for any errors or omissions. Cited non-patent literature

[0000] Liu Ruiheng, Fu Shiyu, Zhan Huaiyu. Spectrophotom Determination of Superoxide Anion Radical with Nitroblue Tetrazolium [J]. Journal of Instrumental Analysis, 2008, 27(4), 355-359

[0024]

Claims

[1] Microbial antioxidant nutrient obtainable by a process for preparation characterized by the following steps: 1) cultivating a strain composition consisting of Lactobacillus plantarum, Bifidobacterium breve and Saccharomyces cerevisiae in a composite stock medium to obtain a liquid bacterial composition; and 2) Freeze-drying of the liquid bacterial composition to obtain a composite bacterial powder, which can be used as a microbial antioxidant nutrient. [2] Nutrient according to claim 1, characterized by that the manufacturing process comprises the following step following steps 1 and 2: 3) Mixing the composite bacterial powder with xylanase, neutral protease and gamma-aminobutyric acid (GABA). [3] Nutrient according to claim 1 or 2, characterized bythat the individual strains in the strain composition have the following parts by weight: 40 to 60 parts by weight of Lactobacillus plantarum; 40 to 60 parts by weight of Bifidobacterium breve; 10 to 20 parts by weight of Saccharomyces cerevisiae. [4] Nutrient according to one of claims 1 to 3, characterized by that the composite stock medium comprises the following components: 0.5 to 3.5 wt% molasses, 0.5 to 2% w / w lactose, 0.5 to 1 wt% tryptone, 0.5 to 2.5 wt.% corn steep liquor, 0.1 to 0.3 wt% sodium chloride, 0.01 to 0.03 wt.% magnesium sulfate, 0.01 to 0.05 wt% iron sulfate, 0.2 to 0.8 wt.% potassium dihydrogen phosphate, 90.82 to 97.68 wt% water. [5] Nutrient according to one of claims 1 to 4, characterized by that the composite stock medium has a pH in the range of 5.5 to 6.

0. [6] Nutrient according to one of claims 1 to 5, characterized by that the cultivation of the stock composition in step 1 is carried out by inoculating into the composite stock medium in a weight ratio of 2:100 to 10:

100. [7] Nutrient according to one of claims 1 to 6, characterized by that the cultivation is carried out at a temperature in the range of 25 to 35°C for a period of time in the range of 16 to 24 hours. [8] Nutrient according to one of claims 1 to 7, characterized by that freeze-drying according to step 2 is carried out as soon as the total viable bacteria reach a concentration of 2 billion CFU / ml. [9] Nutrient according to one of claims 1 to 8, characterized by that after step 2, sterilization of the bacterial powder obtained in step 2 is carried out. [10] Nutrient according to claim 9, characterized bythat sterilization is carried out at a temperature of 115 to 130°C for a period of time in the range of 15 to 30 minutes. [11] Nutrient according to one of claims 2 to 10, characterized by that the microbial antioxidant nutrient has the following components: 40 to 60 parts by weight of compound bacteria powder, 10 to 20 parts by weight of xylanase 10 to 20 parts by weight of neutral protease and 5 to 10 parts by weight of gamma-aminobutyric acid (GABA).