阅读说明：本技术 过瘤胃保护淀粉酶及其制备方法 (Rumen-protected amylase and preparation method thereof ) 是由 武瑞 刘春海 陶春卫 于 2019-06-11 设计创作，主要内容包括：本发明公开了一种过瘤胃保护淀粉酶及其制备方法。该过瘤胃保护淀粉酶由芯材和囊材两部分组成；囊材的主要成分为饱和脂肪酸、丙烯酸树脂IV或壳聚糖中任意一种或几种,丙烯酸树脂IV和壳聚糖不混合,芯材的主要成分为淀粉酶。制法：将淀粉酶进行干法制粒,低温烘干,流化；将熔融态的饱和脂肪酸、溶解于乙醇中的丙烯酸树脂IV或溶解于乙酸中的壳聚糖喷涂到颗粒表面,进行包被,制得过瘤胃保护淀粉酶。本发明使得淀粉酶在反刍动物的瘤胃中不易被降解,在4小时间里过瘤胃率仍达到90％以上,过瘤胃保护淀粉酶到达反刍动物的肠道中可有效释放,能够有效消化小肠内未被消化的淀粉,减少了酮病、亚临床酮病及脂肪肝病的发生,提高了产奶量。(The invention discloses a rumen protected amylase and a preparation method thereof. The rumen protected amylase consists of a core material and a capsule material; the main component of the capsule wall material is one or more of saturated fatty acid, acrylic resin IV and chitosan, the acrylic resin IV and the chitosan are not mixed, and the main component of the core material is amylase. The preparation method comprises the following steps: dry granulating amylase, drying at low temperature, and fluidizing; and spraying the saturated fatty acid in a molten state, the acrylic resin IV dissolved in ethanol or the chitosan dissolved in acetic acid on the surface of the granules for coating to prepare the rumen protected amylase. The invention ensures that amylase is not easily degraded in the rumen of the ruminant, the rumen bypass rate still reaches more than 90 percent within 4 hours, the amylase can be effectively released when the amylase reaches the intestinal tract of the ruminant after rumen bypass protection, the undigested starch in the small intestine can be effectively digested, the occurrence of ketosis, subclinical ketosis and fatty liver disease is reduced, and the milk yield is improved.)

1. A rumen protected amylase, which is characterized in that: consists of a core material and a capsule material; the main component of the capsule wall material is any one or more of saturated fatty acid, acrylic resin IV and chitosan, wherein the acrylic resin IV and the chitosan are not mixed, and the main component of the core material is amylase.

2. The rumen-protected amylase according to claim 1, wherein: the melting point of the saturated fatty acid is above 52 ℃ and the mass percentage of the C16-C18 saturated fatty acid is above 70%; preferably, the C16-C18 saturated fatty acid is palm oil fat powder.

3. The rumen-protected amylase according to claim 1, wherein: the C16-C18 saturated fatty acid is selected from any one of stearic acid and salt thereof, animal and vegetable source hydrogenated oil and animal and vegetable source fatty alcohol.

4. The rumen-protected amylase according to claim 1, wherein: the rumen-protected amylase per 1g comprises 100U-5000U of amylase.

5. The rumen-protected amylase according to claim 4, wherein: the amylase is selected from any one of alpha-amylase, beta-amylase, isoamylase and saccharifying enzyme.

6. The method for preparing rumen protected amylase according to any one of claims 1-5, comprising the steps of:

(1) dry granulating amylase, and drying at low temperature of 60-75 deg.C;

(2) heating and melting saturated fatty acid to obtain molten saturated fatty acid; dissolving acrylic resin IV in ethanol to obtain acrylic resin IV dissolved in ethanol; dissolving chitosan in acetic acid to obtain chitosan dissolved in acetic acid; then, any one or more of the treated saturated fatty acid, acrylic resin IV or chitosan is used as a capsule wall material, wherein the acrylic resin IV and the chitosan are not mixed;

(3) and (3) adding the dried particles obtained in the step (1) into a fluidized bed for fluidization, spraying the capsule wall material obtained in the step (2) onto the surfaces of the particles, and coating to obtain the rumen protected amylase.

7. The preparation method according to claim 6, wherein in the step (1), other components not containing starch are further added to the amylase to be mixed, and then dry granulation is performed; preferably, the other ingredient not containing starch is an amino acid; further preferably, the amino acid is lysine.

8. The method according to claim 7, wherein the mass ratio of the amylase to the amino acid is 1: 0-9.

9. The production method according to claim 6, wherein in the step (1), low-temperature baking is performed at 70 ℃.

10. The production method according to claim 1, wherein in the step (2), the temperature of the heat-melting is 80 ℃ to 100 ℃.

Technical Field

The invention relates to the field of nutrient substances of ruminants, in particular to rumen-bypass protective amylase and a preparation method thereof.

Background

Starch is a high polymer formed by polycondensation of a plurality of glucose molecules, can be converted into glucose under the action of amylase, is a main energy source of the dairy cows, and is also a main source of the glucose of the dairy cows. Starch is mainly connected by alpha-1, 4 glycosidic bonds, and a small amount of alpha-1, 6 glycosidic bonds, and is divided into amylose and amylopectin by different molecular structures. Ruminants are fundamentally different from monogastric animals in the digestion and utilization of starch due to the fermentation of the rumen. After entering the rumen, starch is firstly decomposed into Volatile Fatty Acid (VFA) by rumen microorganisms, and starch which is not decomposed enters the small intestine to be rumen bypass starch (RES), and is hydrolyzed into glucose under the action of pancreatic alpha-amylase and mucosa oligosaccharidase to be absorbed. Compared with starch which passes through the rumen and reaches the small intestine, the starch degraded in the rumen has higher utilization rate of the energy of the starch digested and absorbed in the small intestine, and the functional efficiency of the starch in the small intestine is 42 percent higher than that of the rumen. Meanwhile, the increase of the rumen-bypass starch amount can increase the starch amount digested by the small intestine, and the glucose amount absorbed by the small intestine can be correspondingly increased, so that the amino acid required in the gluconeogenesis process can be reduced, the amino acid in the body can be saved, the deposition of the body protein can be promoted, and the research result of Cameron et al (1991) shows that the addition of the starch in the daily ration can increase the methionine and arginine amount entering the small intestine. However, excessive rumen-bypass starch supply reduces the small intestine digestibility of starch, resulting in energy loss. Insufficient secretion of pancreatic alpha-amylase is the most critical factor limiting the digestibility of the small intestine. Philippeau et al (1999b) considered that there was some potential for large amounts of starch to escape from small intestine digestion into large intestine fermentation, increasing the amount of starch digested in the small intestine.

However, the addition of too much starch to the ration may result in a change in the type of rumen fermentation, resulting in increased propionic acid production, acetic acid: the lower the propionic acid proportion, thus affecting the normal digestion of the fiber and the reduction of the milk fat rate. The rumen bypass starch is added, so that unbalanced rumen fermentation can be prevented, and higher feed intake can be kept, and the intake of daily ration energy can be improved; however, the rumen bypass starch has a limited ability to digest and absorb glucose in the small intestine. Huntington et al (1997) reported that the maximum daily glucose absorption by the small intestine of a cow was 1300 g. To prevent excessive fermentation of starch in the posterior digestive tract, the maximum amount of rumen bypass starch in the dairy cow ration should also be limited. In production, in order to improve the production performance of dairy cows and dairy goats, the dairy cows and dairy goats are fed with excessive feed rich in fermentable carbohydrates, so that nutritional metabolic diseases such as tumor, gastric acidosis and the like are caused. It is generally believed that subacute rumenic acidosis (SARA) may have occurred when the rumen pH falls between 5.5 and 5.0. Concomitant with SARA are nutritional and metabolic diseases such as laminitis, which all reduce productivity.

Starch is the major source of energy for animals, and 60% -80% of the energy required by animals comes from starch in the feed. Amylases are key enzymes in starch digestion and absorption. The amylase is added into the feed, so that the digestion and absorption of animals to starch can be improved, the production performance is improved, and the utilization rate of the feed is improved. Amylases are a class of enzymes that specifically hydrolyze starch. At present, people make great progress on the research of amylase in all aspects, and besides the animal production industry, the amylase is also commonly applied to the industries of paper making, textile, medicine, food processing and the like. In the animal production industry, amylase is mainly added into animal feed to make up for the shortage of amylase in animals, thereby improving the utilization rate of the feed, reducing the breeding cost and reducing the pollution of the breeding environment. The animal can obtain the exogenous amylase artificially added by the feed, and the endogenous digestive enzyme can be secreted by the digestive system of the animal. Both exogenous and endogenous amylases can help animals digest starch in the feed, enhance the immunity of animal bodies and facilitate the growth of the animals.

Amylases are classified in various ways, most commonly into alpha-amylase, beta-amylase, isoamylase and saccharifying enzyme according to the difference of starch hydrolysis forms, and can be directly named as corresponding amylase according to the source and main products generated by hydrolysis. The four common mechanisms of amylase action are:

alpha-amylase can act on any alpha-1, 4 glycosidic bond in a starch molecule, and when encountering the alpha-1, 6 glycosidic bond, the alpha-amylase crosses the alpha-1, 4 glycosidic bond to break the alpha-1, 4 glycosidic bond;

② beta-amylase can also break alpha-1, 4 glycosidic bonds, however it can not break alpha-1, 6 glycosidic bonds and the reaction is immediately cut off as soon as this bond is touched. The method can start from the non-reducing end of a starch molecule, and sequentially break alpha-1, 4 glycosidic bonds to finally generate maltose;

the isoamylase can specifically hydrolyze alpha-1, 6 glycosidic bond of branch position in the amylopectin and convert the alpha-1, 6 glycosidic bond into amylose;

the glucoamylase can act on various glycosidic bonds, not only can break alpha-1, 4 glycosidic bonds, but also can hydrolyze alpha-1, 3 and alpha-1, 6 glycosidic bonds. The final product, which in turn cleaves the alpha-1, 4 glycosidic bond, is beta-D glucose.

From the view of the physiological characteristics of animal digestion, the amylase which can really play an effective role should have the following characteristics: (1) has higher activity under the condition of animal body temperature (37-42 ℃); (2) the optimum pH value is consistent with the pH value of chyme in the digestive tract; (3) has higher enzymolysis effect on starch (should be endo-amylase);

has good stability, including stability in high temperature granulation process of feed, stability in preservation process, and tolerance to gastric acid, pepsin, trypsin, metal ions, etc. in animal digestive tract.

The use of enzyme preparations is influenced by rumen and rumen microorganisms due to the specific digestive system of ruminants. In addition, the daily ration for the ruminant takes coarse feed as a main material, and the content of coarse fiber is high, so that the digestive function of the ruminant is not obviously affected by adding amylase in the feed for the ruminant. The research of the WangIn wave et al (2000) shows that the higher content of fibers in the ration can cause the reduction of the activity of endogenous digestive enzymes of animals, and the reason is mainly that the crude fibers are complexed with the digestive enzymes, thereby hindering the reaction between the digestive enzymes and the substrate. Therefore, the common amylase is applied to the daily ration of the ruminant, and has no obvious effect of improving the production performance of the ruminant.

The information disclosed in this background section is only for enhancement of understanding of the general background of the invention and should not be taken as an acknowledgement or any form of suggestion that this information forms the prior art already known to a person skilled in the art.

Disclosure of Invention

The invention aims to provide rumen-protected amylase and a preparation method thereof, and the obtained rumen-protected amylase can prevent amylase from being degraded in rumen, and can effectively release in intestinal tract to further improve the digestibility of intestinal starch, provide more energy for ruminants, and improve weight gain or milk yield.

In order to realize the aim, the invention provides rumen-protected amylase, which consists of a core material and a capsule material; the main component of the capsule wall material is any one or more of saturated fatty acid, acrylic resin IV and chitosan, wherein the acrylic resin IV and the chitosan are not mixed, and the main component of the core material is amylase.

In a preferred embodiment, the melting point of the saturated fatty acid is above 52 ℃ and the mass percentage of the C16-C18 saturated fatty acid is above 70%; preferably, the C16-C18 saturated fatty acid is palm oil fat powder.

In a preferred embodiment, the saturated fatty acid having 16 to 18 atoms is selected from stearic acid and salts thereof, hydrogenated oils derived from animals and plants, and fatty alcohols derived from animals and plants.

In a preferred embodiment, the rumen-protected amylase comprises 100U-5000U of amylase per 1g of the rumen-protected amylase;

the definition of U above is: the enzyme activity is specified as an international unit: under specific conditions, the amount of enzyme required to convert 1. mu. mol of substrate, or 1. mu. mol of the relevant group in the substrate, in 1 minute is called an international unit of enzyme activity (IU, also called U).

In a preferred embodiment, the amylase is selected from any one of α -amylase, β -amylase, isoamylase and saccharifying enzyme.

The invention also provides a preparation method of the rumen protected amylase, which comprises the following steps:

(1) dry granulating amylase, and drying at low temperature of 60-75 deg.C;

(2) heating and melting saturated fatty acid to obtain molten saturated fatty acid; dissolving acrylic resin IV in ethanol to obtain acrylic resin IV dissolved in 95% ethanol; dissolving chitosan in acetic acid to obtain chitosan dissolved in acetic acid; then, any one or more of the treated saturated fatty acid, acrylic resin IV or chitosan is used as a capsule wall material, wherein the acrylic resin IV and the chitosan are not mixed;

(3) and (3) adding the dried particles obtained in the step (1) into a fluidized bed for fluidization, spraying the capsule wall material obtained in the step (2) onto the surfaces of the particles, and coating to obtain the rumen protected amylase.

In a preferred embodiment, in step (1), other components not containing starch can be added into amylase for mixing, and then dry granulation is carried out; preferably, the other ingredient not containing starch is an amino acid; further preferably, the amino acid is lysine.

In a preferred embodiment, the mass ratio of the amylase to the amino acid is 1: 0-9.

In a preferred embodiment, in step (1), low temperature drying is performed at 70 ℃.

In a preferred embodiment, in the step (2), the temperature of the heat melting is 80 to 100 ℃.

The rumen protected amylase is granular, and has a diameter of about 1mm, so as to facilitate rapid rumen passing and ensure the mixing uniformity of the product in the feed.

Definition of amylase activity: 1g of enzyme powder was liquefied at pH 6.0 at 60 ℃ for 1 hour to obtain 1g of soluble starch as one unit of enzyme activity, expressed in U/g.

Compared with the prior art, the invention has the following beneficial effects:

(1) the invention relates to a rumen-bypass granule prepared by amylase by a microcapsule coating technology, wherein the rumen-bypass technology protects nutrient substances easily damaged by rumen microorganisms from being decomposed by the rumen microorganisms by a physical or chemical method, and the nutrient substances pass through the rumen completely and reach the abomasum and the intestinal tract to be released and play a role in the small intestine, so that the requirement of organisms on the nutrient substances is met; the rumen-protected amylase obtained by the invention can ensure that the amylase is not easily degraded in the rumen of a ruminant, the rumen-protected rate still reaches more than 90 percent within 4 hours, and the coating effect of the amylase is good.

(2) The rumen protected amylase obtained by the invention can be effectively released when reaching the intestinal tract of a ruminant, the small intestine release rate within 2 hours reaches more than 90%, and after 8 hours, the amylase has been released by 100%, so that the rumen protected amylase can effectively digest undigested starch in the small intestine, further improve the digestion rate of the intestinal starch in the ruminant, and provide more energy for the ruminant, thereby effectively preventing the ketosis of the ruminant in the perinatal period, reducing the occurrence of ketosis or subclinical ketosis and fatty liver disease, reducing the postpartum weightlessness of the ruminant, improving the estrus conception rate of the ruminant and improving the milk yield.

Detailed Description

The following detailed description of specific embodiments of the invention is provided, but it should be understood that the scope of the invention is not limited to the specific embodiments.

The alpha-amylase used in the examples below was purchased from Shandong Longglong bioengineering, Inc.; palm oil fat powder was purchased from Tianjin Nuoist trade, Inc.

The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.

Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.