阅读说明：本技术 乳酸乳球菌kdll2016-01在发酵羽毛制备蛋白饲料中的应用 (Application of lactococcus lactis KDLL2016-01 in preparation of protein feed by fermenting feathers ) 是由 徐良梅 陈志辉 刘洋 李岚雪 郭建新 于 2018-11-29 设计创作，主要内容包括：本发明公开了本发明涉及乳酸乳球菌KDLL2016-01在发酵羽毛制备蛋白饲料中的应用,属于动物营养与饲料技术领域。该应用是将羽毛粉碎成羽毛粉,灭菌后加入发酵培养基中；将乳酸乳球菌KDLL2016-01接种至发酵培养基中进行发酵培养；发酵完成后将发酵产物进行干燥,获得蛋白饲料。该应用所采用的乳酸乳球菌KDLL2016-01在培养过程中不产生有害代谢产物,产物可以直接饲喂畜禽,无需对发酵产品进行脱毒等处理,而且发酵过程中产生的乳球菌肽也可以对饲料中可能存在的致病菌也有杀伤作用,使饲料品质更加安全。本发明方法制备的蛋白饲料适用于饲喂畜禽。(The invention discloses application of lactococcus lactis KDLL2016-01 in preparation of protein feed by fermenting feathers, and belongs to the technical field of animal nutrition and feed. The application comprises pulverizing feather into feather powder, sterilizing, and adding into fermentation culture medium; inoculating lactococcus lactis KDLL2016-01 into a fermentation culture medium for fermentation culture; and drying the fermentation product after the fermentation is finished to obtain the protein feed. The lactococcus lactis KDLL2016-01 adopted in the application does not produce harmful metabolites in the culture process, products can be directly fed to livestock and poultry, detoxification and other treatment of fermented products are not needed, lactococcus peptides produced in the fermentation process can also have a killing effect on pathogenic bacteria possibly existing in the feed, and the feed quality is safer. The protein feed prepared by the method is suitable for feeding livestock and poultry.)

1. The application of Lactococcus lactis KDLL2016-01 in preparing protein feed by fermenting feathers is disclosed, wherein the preservation number of the Lactococcus lactis KDLL2016-01 is CGMCC No. 15450.

2. Use according to claim 1, characterized in that it comprises the following steps:

the method comprises the following steps: pulverizing feather into feather powder, sterilizing, and adding into fermentation culture medium;

step two: inoculating lactococcus lactis KDLL2016-01 into a fermentation culture medium for fermentation culture;

step three: and drying the fermentation product after the fermentation is finished to obtain the protein feed.

3. The use of claim 2, wherein the feather pulverization of the feathers into feather meal in step one is carried out according to the following method: cleaning feather, mixing with water and agar, stirring uniformly, drying, pulverizing, placing on a nylon bag of 400 meshes, cleaning with flowing water to remove agar, drying, and sieving with a 60-mesh sieve; wherein: the weight ratio of the feather to the water is 10: 1; the weight ratio of the agar to the feather is 1: 200.

4. The use of claim 2, wherein the feather meal of step one is added in an amount of: the ratio of the weight of feather meal to the weight of fermentation medium was 1: 4.

5. The application of claim 2, wherein in the second step, the lactococcus lactis KDLL2016-01 is subjected to recovery culture before inoculation, then inoculated into a liquid seed culture medium for seed culture for 48 hours under the conditions of 30 ℃ temperature, pH value of 6.0 and rotation speed of 200rpm to obtain a seed culture solution of the lactococcus lactis KDLL2016-01, and the obtained seed culture solution of the lactococcus lactis KDLL2016-01 is inoculated into a fermentation culture medium.

6. The use according to claim 6, wherein the liquid seed culture medium has a formulation of: 8.0g of beef extract, 10.0g of peptone, 4.0g of yeast extract, 20.0g of glucose, 2.0g of diammonium hydrogen citrate, 2.0g of dipotassium hydrogen phosphate, 5.0g of sodium acetate, 0.2g of magnesium sulfate, 801.0g of tween, 0.05g of manganese sulfate and 1000mL of deionized water.

7. The use of claim 2, wherein the fermentation medium formulation of step two is: 10g of beef extract, 10g of yeast extract, 100g of cane sugar, 2g of sodium chloride, 10g of dipotassium hydrogen phosphate and 0.5g of magnesium sulfate, and the pH value is 6.0.

8. The use according to claim 2, wherein the conditions of the fermentation culture in step two are: the inoculation amount is 5.7% (v/v), the fermentation temperature is 32 ℃, the rotation speed is 220rpm, and the fermentation time is 36h, and the fermentation is aerobic, and the initial pH value is 6.3.

9. The use of claim 2, wherein the drying of the fermentation product in step three comprises filtering the fermentation product to obtain a fermentation broth and a solid, spray-drying the fermentation broth, drying the solid, and combining the dried solid powder with the spray-dried solid powder to obtain the protein feed.

10. A protein feed produced by the use of any one of claims 1 to 9.

Technical Field

The invention relates to application of lactococcus lactis KDLL2016-01 in preparation of protein feed by fermenting feathers, and belongs to the technical field of animal nutrition and feed.

Background

At present, the lack of protein feed is a worldwide problem, and the lack of good quality and sufficient protein feed and the lack of good quality and sufficient meat products greatly limit the rapid and healthy development of livestock and poultry breeding industry. Feather resources in China are very abundant, but feather resources are not well utilized for a long time, which is mainly related to the structural properties of feathers. The feather structure is very stable, so that the feather is stable in property and insoluble in water, dilute acid and dilute alkali, and before processing, the protease cannot hydrolyze the feather, so that animals cannot digest the feather and cannot utilize protein in the feather, and the feather contains high crude protein, mainly keratin which is difficult to digest by livestock and poultry. Therefore, the method is of great importance for destroying the stable structure of the feather, improving the water solubility of the feather and improving the digestibility of the feather so that the feather becomes easy to digest and absorb. The current methods commonly used to hydrolyze feathers are as follows:

the high-temperature high-pressure hydrolysis method is the most widely used treatment method at present, the color of the finished product can change along with the change of the raw materials, when the color of the raw materials is white, the finished product is dark yellow generally, when the color of the raw materials is black, the finished product is dark brown generally, and under general conditions, the finished product of the high-pressure hydrolyzed feather meal is light brown. The high-temperature high-pressure hydrolysis method has the disadvantages that the temperature and pressure are difficult to control, the quality of the produced product is not very stable, and the protein denaturation is caused by overhigh temperature, so that the digestibility of the feather meal is reduced.

The chemical hydrolysis method is a treatment method which is invented by the characteristic that the sensitivity of the feather to acid and alkali is greatly improved under the heating condition. However, the chemical hydrolysis method also has many defects, the production process of the chemical hydrolysis method is relatively complex, the produced hydrolyzed feather meal contains more salt, which can affect the palatability of the hydrolyzed feather meal, and the more salt can easily absorb moisture, thus causing the deterioration of the hydrolyzed feather meal. The chemical hydrolysis method can produce a large amount of sewage in the production process and can cause serious pollution to the environment.

The principle of the puffing method is that feather powder is subjected to a great shearing effect when passing through the puffing channel, the external pressure disappears rapidly at the moment when the feathers are flushed out of the puffing outlet, the interior of the feather powder still keeps a high-pressure state, and the feathers expand rapidly due to the internal and external pressure difference. The firm space structure of the feather can be damaged during puffing, the structure of keratin becomes loose, the hydrophobicity is reduced to some extent, the hydrophilicity is improved, the feather is hydrolyzed more easily by protease, and the feather is digested more easily by animals and absorbs the required nutrition from the feather.

Microbial degradation means that certain microorganisms in nature can produce enzymes capable of hydrolyzing keratin during growth to degrade keratin into small peptides or amino acids. Compared with other treatment methods, the microbial degradation method has great advantages, mainly manifested in low cost, simple process, and less waste water and gas. Microorganisms having keratin degrading ability are mainly bacteria, fungi and actinomycetes. However, the secondary metabolites of conventional microorganisms may contain substances harmful to livestock and poultry, and thus usually require detoxification treatment.

The traditional acid-base treatment and puffing method can cause environmental pollution on one hand and damage to the nutrient components of feather keratin on the other hand.

In addition, the feather is used as protein feed, the cystine content of the feather is too high, the protein quality of the feather is poor compared with that of common fish meal, and amino acid imbalance is easily caused when the feed is prepared.

At present, domestic articles about degrading feather keratin by probiotics are rare, and the research on degrading feather keratin by lactococcus has not been reported yet on the method for feeding livestock and poultry by fermentation products and the feeding effect.

Disclosure of Invention

Aiming at the problems that the feather is greatly damaged by nutritional ingredients and pollutes the surrounding environment when being processed by a chemical hydrolysis method and an expansion method to be used as a protein feed, and the problems that secondary metabolites possibly generate toxicity to livestock and poultry and the like in the microbial fermentation of the feather, lactococcus which is one of probiotics is adopted to ferment the feather to produce the feather protein feed. The method has less pollution to surrounding environment, the amino acids of fermented feather powder are more balanced, the nutritive value is improved to a certain extent, and no harmful secondary pollution is caused to livestock and poultryThe level metabolite can be directly used for feeding livestock and poultry. The invention provides application of Lactococcus lactis KDLL2016-01 in preparation of protein feed by fermenting feathers, and discovers Lactococcus lactis KDLL2016-01 capable of degrading feather keratin and new application thereof for the first time, wherein the Lactococcus lactis can be used for preparing protein feed by fermenting and degrading feathersFermented protein feedThe amino acid of the protein feed product obtained by fermentation is more balanced, the content of cystine is reduced to a certain extent, and the protein feed product is rich in probiotics beneficial to the growth of livestock and poultry, and the specific scheme is as follows:

the invention provides an application of Lactococcus lactis (Lactococcus lactis) KDLL2016-01 in preparation of a protein feed by fermenting feathers, namely a method for preparing the protein feed by fermenting feathers by using Lactococcus lactis (Lactococcus lactis) KDLL2016-01, wherein the preservation number of the Lactococcus lactis (Lactococcus lactis) KDLL2016-01 is CGMCC No. 15450.

Preferably, said application comprises the steps of:

the method comprises the following steps: pulverizing feather into feather powder, sterilizing, and adding into fermentation culture medium;

step two: inoculating lactococcus lactis KDLL2016-01 into a fermentation culture medium for fermentation culture;

step three: and drying the fermentation product after the fermentation is finished to obtain the protein feed.

Preferably, the feather is crushed into feather powder in the first step according to the following method: cleaning feather, mixing with water and agar, stirring uniformly, drying, pulverizing, placing on a nylon bag of 400 meshes, cleaning with flowing water to remove agar, drying, and sieving with a 60-mesh sieve; wherein: the weight ratio of the feather to the water is 10: 1; the weight ratio of the agar to the feather is 1: 200.

Preferably, the addition amount of the feather powder in the first step is as follows: the ratio of the weight of feather meal to the weight of fermentation medium was 1: 4.

Preferably, in the second step, the lactococcus lactis KDLL2016-01 is subjected to recovery culture before inoculation, then inoculated into a liquid seed culture medium for seed culture for 48 hours under the conditions of the temperature of 30 ℃, the pH value of 6.0 and the rotation speed of 200rpm to obtain a seed culture solution of the lactococcus lactis KDLL2016-01, and the obtained seed culture solution of the lactococcus lactis KDLL2016-01 is inoculated into a fermentation culture medium.

More preferably, the formula of the liquid seed culture medium is as follows: 8.0g of beef extract, 10.0g of peptone, 4.0g of yeast extract, 20.0g of glucose, 2.0g of diammonium hydrogen citrate, 2.0g of dipotassium hydrogen phosphate, 5.0g of sodium acetate, 0.2g of magnesium sulfate, 801.0g of tween, 0.05g of manganese sulfate and 1000mL of deionized water.

Preferably, the inoculation amount of the inoculation in the second step is 5.7% (v/v).

Preferably, the formula of the fermentation medium in the second step is as follows: 10g of beef extract, 10g of yeast extract, 100g of cane sugar, 2g of sodium chloride, 10g of dipotassium hydrogen phosphate and 0.5g of magnesium sulfate, and the pH value is 6.0.

Preferably, the conditions of the fermentation culture in the second step are as follows: aerobic, the initial pH value is 6.3, the fermentation temperature is 32 ℃, the rotating speed is 220rpm, and the fermentation time is 36 h.

Preferably, the drying of the fermentation product in the third step is to filter the fermentation product to obtain a fermentation liquid and a solid, spray-dry the obtained fermentation liquid, dry the obtained solid, and then combine the dried solid powder with the spray-dried solid powder to obtain the protein feed.

The invention also provides a protein feed prepared by any one of the applications.

The lactobacillus provided by the invention is a probiotic strain capable of degrading keratin, which is separated and screened from waste feathers accumulated in a chicken farm by using feather meal as a single nitrogen source, and the strain is preserved in China general microbiological culture Collection center (CGMCC) in 2018, 3 months and 14 days, and the preservation number is CGMCC No. 15450.

The invention has the beneficial effects that:

1. the invention discovers a lactococcus lactis KDLL2016-01 strain capable of degrading feather keratin for the first time, wherein the preservation number of the lactococcus lactis KDLL2016-01 strain is CGMCC No.15450, the lactococcus lactis strain is a probiotic strain, harmful metabolites are not generated in the culture process, the product can be directly fed to livestock and poultry, detoxification and other treatment on fermented products are not needed, lactococcus peptide generated in the fermentation process can also kill pathogenic bacteria possibly existing in the feed, and the feed quality is safer.

2. The protein content in feather meal can reach more than 80%, but basically keratin with compact structure and difficult digestion is contained. After the feather is fermented by the method, the compact keratin structure contained in the feather is decomposed or destroyed by lactococcus and metabolites thereof, so that the digestion rate of livestock and poultry on feather protein feed is higher, the pH value of the feather fermented product is properly reduced, the palatability is better, and the feed intake of livestock and poultry can be improved.

3. The feather is used as protein feed, the cystine content of the feather is too high, the protein quality of the feather is poor compared with that of common fish meal, and the amino acid imbalance is easily caused when the feather is prepared into the feed. The content of the cystine in the fermented feather powder obtained by the method is reduced to a certain extent, and the proportion of the amino acid is more balanced.

3. The feather degradation rate can reach 35% by fermenting feathers with KDLL 2016-01.

4. The combined fermentation product obtained by the invention is used as a protein feed to replace fish meal to prepare a feed for feeding laying hens.

5. The invention adopts the feathers as the animal source protein feed, has low cost and is easy to obtain the feathers.

Drawings

FIG. 1 shows the degradation circle of the lactococcus strain.

FIG. 2 is a 16S rDNA phylogenetic tree.

FIG. 3 shows the apparent morphology of feather meal (left) and fermented feather meal (right).

Fig. 4 shows feather powder (left) and fermented feather powder (right) under a normal electron microscope.

FIG. 5 feather meal (left) and fermented feather meal (right) under scanning electron microscope.

Detailed Description

The present invention will be further described with reference to the following specific examples, but the present invention is not limited to these examples.

The "fermented feather meal" in the following examples is the "protein feed".