CN113558146A

CN113558146A - Feed buffering agent, preparation method and application thereof, and feed using feed buffering agent

Info

Publication number: CN113558146A
Application number: CN202010359860.XA
Authority: CN
Inventors: 张源淑; 叶平生; 李林; 倪迎冬; 庄苏; 沈向真; 陈杰
Original assignee: Nanjing Agricultural University
Current assignee: Nanjing Agricultural University
Priority date: 2020-04-29
Filing date: 2020-04-29
Publication date: 2021-10-29

Abstract

The invention provides a feed buffering agent, a preparation method and application thereof as well as a feed using the same, and relates to the technical field of feed additives, the feed buffering agent provided by the invention mainly comprises sodium bicarbonate, magnesium oxide, sodium acetate and sodium butyrate, and the sodium bicarbonate can play an acid-base buffering effect on rumen and maintain an ideal rumen fermentation environment; the magnesium oxide can maintain beneficial biological flora in the gastrointestinal tract and increase the number of beneficial bacteria in the intestinal tract; sodium acetate can increase acetic acid concentration in rumen, and improve milk fat rate; sodium butyrate can increase ruminant appetite. The feed buffer provided by the invention has certain effects of preventing and relieving rumenic acidosis of the ruminant, maintaining the number of beneficial flora in the intestinal tract of the ruminant, improving the health state of an organism, improving the lactation performance of the lactating animal, improving the milk quality, increasing the milk fat, the milk sugar and the milk yield content, and enhancing the protection effect on mammary gland by matching the specific components.

Description

Feed buffering agent, preparation method and application thereof, and feed using feed buffering agent

Technical Field

The invention relates to the technical field of feed additives, in particular to a feed buffering agent, a preparation method and application thereof, and a feed using the same.

Background

Lactation is a complex process, and the main metabolic feature of lactating animals in the process of producing milk is high energy demand. At present, due to the shortage of cultivated land and grassland, the coarse material is deficient or the quality is not good in China. Therefore, the high concentrate feed is usually added in production to meet the requirement of high energy of the lactating cows or dairy goats, but studies have proved that when the lactating cows or dairy goats are fed with the high concentrate feed for a long time, rumen pH is reduced, which in turn causes the excessive release of Subacute rumen acidosis (SARA) and abnormal products such as Lipopolysaccharides (LPS), which exceed the intestinal detoxification capability and are shifted to blood, thus causing inflammatory reaction of the organism and metabolic disturbance of the liver, affecting the synthesis and content of milk components in the mammary gland, and finally causing the reduction of milk quality. That is to say, when SARA is adopted, the reduction of the pH value of rumen and the excessive release of abnormal products LPS and the like are key links which cause the reduction of milk quality caused by the long-term feeding of high concentrate. Therefore, controlling the SARA of ruminants and improving the pH value of rumen can be one way to relieve the milk quality reduction caused by long-term feeding of high concentrate. SARA is currently a problem of great concern to dairy farmers because it is associated with the development of various diseases such as mastitis, milk fat suppression, laminitis, liver abscesses and even death. The harm brought by high-precision materials seriously influences the healthy development of the milk industry, and how to regulate the negative effect is very important.

Buffering agents are commonly used to prevent ruminal acidosis in ruminants. The buffering agent can not only improve the reduction of the milk fat rate caused by the feeding of high-concentrate daily ration by the ruminant, but also improve the production performance of the ruminant. Therefore, the addition of buffers to the ration has become an important measure to improve animal performance in recent years. At present, buffering agents commonly used for ruminants at home and abroad include sodium bicarbonate, sodium acetate, sodium butyrate, magnesium oxide, calcium carbonate and the like. A single buffer is researched more, and the research on the comprehensive regulation and control of ruminants by two or more than two composite buffers is not reported much.

In view of the above, the present invention is particularly proposed.

Disclosure of Invention

It is a first object of the present invention to provide a feed buffer to alleviate at least one of the technical problems of the prior art.

The second purpose of the invention is to provide the preparation method of the feed buffer, the method has simple process and convenient operation, can save a large amount of manpower and material resources and reduce the cost, and the prepared feed buffer has good effect of improving the rumen acidosis of ruminants.

The third purpose of the invention is to provide the application of the feed buffering agent in the preparation of products for preventing and/or treating rumen acidosis of ruminants.

A fourth object of the present invention is to provide a feed which is effective in improving the lactation performance of lactating animals while preventing and/or treating ruminal acidosis in ruminants.

The invention provides a feed buffering agent, which mainly comprises the following components:

sodium bicarbonate, magnesium oxide, sodium acetate, and sodium butyrate.

Further, the feed buffer mainly comprises the following components in parts by weight:

1-10 parts of sodium bicarbonate, 1-10 parts of magnesium oxide, 5-15 parts of sodium acetate and 5-15 parts of sodium butyrate.

2-8 parts of sodium bicarbonate, 2-8 parts of magnesium oxide, 8-12 parts of sodium acetate and 8-12 parts of sodium butyrate.

6 parts of sodium bicarbonate, 6 parts of magnesium oxide, 10 parts of sodium acetate and 10 parts of sodium butyrate.

The invention also provides a preparation method of the feed buffer, which is characterized in that sodium bicarbonate, magnesium oxide, sodium acetate and sodium butyrate in parts by weight of the formula are uniformly mixed to obtain the feed buffer.

The invention also provides the application of the feed buffer or the feed buffer prepared by the preparation method in preparing a product for preventing and/or treating ruminant rumen acidosis.

Further, the product is a feed.

In addition, the invention also provides a feed which comprises a basal feed and the feed buffering agent or the feed buffering agent prepared by the preparation method.

Further, the basal feed comprises roughage and/or concentrate;

preferably, the basal feed comprises a concentrate.

Further, the mass ratio of the basal feed to the feed buffer is 25-35: 1, preferably 31: 1.

The feed buffering agent provided by the invention mainly comprises sodium bicarbonate, magnesium oxide, sodium acetate and sodium butyrate, wherein the sodium bicarbonate can play an acid-base buffering effect on rumen and maintain an ideal rumen fermentation environment; the magnesium oxide can maintain beneficial biological flora in the gastrointestinal tract and increase the number of beneficial bacteria in the intestinal tract; sodium acetate can increase acetic acid concentration in rumen, and improve milk fat rate; sodium butyrate can increase ruminant appetite. The feed buffer provided by the invention has certain effects of preventing and relieving rumenic acidosis of the ruminant, maintaining the number of beneficial flora in the intestinal tract of the ruminant, improving the health state of an organism, improving the lactation performance of the lactating animal, improving the milk quality, increasing the milk fat, the milk sugar and the milk yield content, and enhancing the protection effect on mammary gland by matching the specific components.

The preparation method of the feed buffering agent provided by the invention is characterized in that the components of the formula are uniformly mixed, the method is simple in process and convenient to operate, a large amount of manpower and material resources can be saved, the cost is reduced, and the prepared feed buffering agent has good effects of improving ruminant rumen acidosis and improving lactation performance.

The feed provided by the invention comprises a basic feed and the feed buffering agent provided by the invention, and the feed can effectively improve the lactation performance of lactating animals on the basis of ensuring the nutrition supply, and simultaneously prevent and/or treat ruminal acidosis of the ruminants. The feed is supplemented with a proper amount of feed buffer, which is a beneficial feeding mode.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.

FIG. 1 is a graph showing the effect of feed buffer on the milk production of lactating goats according to the experimental examples of the present invention;

FIG. 2 is a graph showing the results of the effect of feed buffer on the milk fat content of lactating goats, as provided in the experimental examples of the present invention;

FIG. 3 is a graph showing the results of the effect of feed buffer on the lactose content of lactating goats, as provided by the experimental examples of the present invention;

FIG. 4 is a graph showing the results of the effect of feed buffer on the rumen pH of lactating goats, as provided by the experimental examples of the present invention;

FIG. 5A is a graph showing the effect of feed buffer on the content of LPS in the rumen of a lactating goat in accordance with an exemplary embodiment of the present invention;

FIG. 5B is a graph showing the results of the effect of feed buffer on LPS content in lactating goat blood provided by the experimental example of the present invention;

FIG. 6A is a graph showing the effect of a feed buffer on Caspase-3 in lactating goats in accordance with an illustrative embodiment of the present invention;

FIG. 6B is a graph showing the effect of a feed buffer on Caspase-9 in lactating goats in accordance with an illustrative embodiment of the present invention;

FIG. 6C is a graph showing the effect of feed buffer on apoptosis protein Bax of lactating goats in accordance with the experimental examples of the present invention;

FIG. 6D is a graph showing the effect of the feed buffer on apoptosis protein Bcl-2 of lactating goats in accordance with the experimental examples of the present invention;

FIG. 7A is a graph showing the results of the detection of the blood lactic acid content of lactating goats by the feed buffer according to the experimental examples of the present invention;

FIG. 7B is a graph showing the results of the measurement of histamine content in blood of lactating goats by using the feed buffer according to the experimental example of the present invention;

FIG. 7C is a graph showing the results of the content detection of inflammatory cytokine TNF- α in blood of lactating goats by the feed buffer according to the experimental examples of the present invention;

FIG. 7D is a diagram showing the results of the content detection of IL-1 β in inflammatory cytokines in blood of lactating goats by using the feed buffer provided in the experimental examples of the present invention.

Detailed Description

The technical solutions of the present invention will be described clearly and completely with reference to the accompanying drawings, and it should be understood that the described embodiments are some, but not all embodiments of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

It should be noted that:

in the present invention, all embodiments and preferred 10-preferred embodiments mentioned herein can be combined with each other to form a new technical solution, if not specifically stated.

In the present invention, all the technical features mentioned herein and preferred features may be combined with each other to form a new technical solution, if not specifically stated.

In the present invention, the percentage (%) or parts means the weight percentage or parts by weight with respect to the composition, if not otherwise specified.

In the present invention, the components referred to or the preferred components thereof may be combined with each other to form a novel embodiment, if not specifically stated.

In the present invention, unless otherwise stated, the numerical range "a-b" represents a shorthand representation of any combination of real numbers between a and b, where a and b are both real numbers. For example, a numerical range of "3 to 30" means that all real numbers between "3 to 30" have been listed herein, and "3 to 30" is simply a shorthand representation of the combination of these numbers 20.

The "ranges" disclosed herein may have one or more lower limits and one or more upper limits, respectively, in the form of lower limits and upper limits.

In the present invention, unless otherwise specified, the individual reactions or operation steps may be performed sequentially or may be performed in sequence. Preferably, the reaction processes herein are carried out sequentially.

Unless otherwise defined, technical and scientific terms used herein have the same meaning as is familiar to those skilled in the art. In addition, any methods or materials similar or equivalent to those described herein can also be used in the present invention.

The high concentrate can cause the milk yield, milk fat and lactose content of the lactating ruminants to be reduced after long-term feeding, and can also induce the body health of the lactating ruminants to be damaged, and abnormal metabolic products such as SARA, rumen and blood LPS (LPS) and the like to be increased. Based on this, according to one aspect of the present invention, there is provided a feed buffer consisting essentially of:

sodium bicarbonate, magnesium oxide, sodium acetate, and sodium butyrate.

Wherein, the sodium bicarbonate is an acid salt which is alkalescent when dissolved in water, thus playing a role of acid-base buffering for rumen, keeping the pH value (6-7) in a relatively stable state and maintaining an ideal rumen fermentation environment. Food grade calcium bicarbonate is preferably used in the present invention.

The magnesium oxide can maintain beneficial biological flora in gastrointestinal tract, increase the quantity of beneficial bacteria in intestinal tract, and not only has the function of enhancing immunity, but also can supplement the deficiency of magnesium element in daily ration. Food grade magnesium oxide is preferably used in the present invention.

Sodium acetate can increase the concentration of acetic acid in rumen, and acetic acid as precursor of lipid synthesis can be added into ruminant feed to improve the milk fat rate. Food grade sodium acetate is preferred for use in the present invention.

The sodium butyrate can increase the appetite of ruminants, has a certain protection effect on the health of organisms, can change the content of butyric acid in blood, enhances the antioxidant stress capability of mammary glands, reduces apoptosis and further enhances the protection effect on mammary glands. Preferably, food grade sodium butyrate is used in the present invention.

The feed buffer provided by the invention has certain effects of preventing and relieving rumenic acidosis of the ruminant, maintaining the number of beneficial flora in the intestinal tract of the ruminant, improving the health state of an organism, improving the lactation performance of the lactating animal, improving the milk quality, increasing the milk fat, the milk sugar and the milk yield content, and enhancing the protection effect on mammary gland by matching the specific components.

In some preferred embodiments, the feed buffer consists essentially of the following components in parts by weight:

The sodium bicarbonate can be, but is not limited to, 1 part, 2 parts, 3 parts, 4 parts, 5 parts, 6 parts, 7 parts, 8 parts, 9 parts or 10 parts; the magnesium oxide can be, for example, but is not limited to, 1 part, 2 parts, 3 parts, 4 parts, 5 parts, 6 parts, 7 parts, 8 parts, 9 parts, or 10 parts; sodium acetate can be, for example, but is not limited to, 5 parts, 6 parts, 7 parts, 8 parts, 9 parts, 10 parts, 11 parts, 12 parts, 13 parts, 14 parts, or 15 parts; the sodium butyrate can be, for example, but not limited to, 5 parts, 6 parts, 7 parts, 8 parts, 9 parts, 10 parts, 11 parts, 12 parts, 13 parts, 14 parts, or 15 parts.

The feed buffer agent obtained by limiting the specific mixture ratio of each specific component has better effects of improving ruminant rumen acidosis and improving lactation performance.

In some more preferred embodiments, the feed buffer consists essentially of the following components in parts by weight:

In some further preferred embodiments, the feed buffer consists essentially of the following components in parts by weight:

Through further adjustment and optimization of the proportion of the components of the feed buffering agent, the feed buffering agent provided by the invention has better effects of improving ruminant rumen acidosis and improving lactation performance.

According to the second aspect of the invention, the preparation method of the feed buffer is also provided, wherein the feed buffer is obtained by uniformly mixing the sodium bicarbonate, the magnesium oxide, the sodium acetate and the sodium butyrate according to the weight parts of the formula.

The preparation method of the feed buffer provided by the invention has the advantages of simple process and convenience in operation, a large amount of manpower and material resources can be saved, the cost is reduced, and the prepared feed buffer has good effects of improving ruminant rumen acidosis and improving lactation performance.

According to the third aspect of the invention, the application of the feed buffer or the feed buffer prepared by the preparation method in the preparation of products for preventing and/or treating ruminant rumen acidosis is also provided.

The feed buffer provided by the invention can play a role in acid-base buffering on rumen and maintain an ideal rumen fermentation environment, so that the feed buffer provided by the invention can be used for preparing products for preventing and/or treating ruminant rumen acidosis.

The product can be, but is not limited to, feed, medicine, health product, etc.

It is understood that "prevention and/or treatment" means that the product has a prophylactic effect, or has a therapeutic effect, or has both a prophylactic and a qualitative effect.

In prophylactic applications, relatively low doses may be administered chronically at relatively infrequent intervals. In therapeutic applications, it is sometimes desirable to administer relatively high doses at relatively short intervals until the progression of the disease is delayed or halted, preferably until the individual exhibits a partial or complete improvement in the symptoms of the disease, after which a prophylactic regimen may be administered.

In some preferred embodiments, the product is a feed.

According to a fourth aspect of the present invention, there is also provided a feed comprising a basal feed and the feed buffer described above or prepared by the preparation method described above.

The feed can effectively improve the lactation performance of the lactating animals on the basis of ensuring the nutrition supply, and simultaneously prevent and/or treat ruminant rumen acidosis. The feed is supplemented with a proper amount of feed buffer, which is a beneficial feeding mode.

In some preferred embodiments, the basal feed comprises a roughage and/or a concentrate.

The coarse fodder is fodder with natural water content below 60% and crude fiber content equal to or higher than 18% and fed in air-dried form, such as pasture, crop straw, distiller's grains, etc. Concentrated feed, also called concentrate, is a feed containing rich nutrients, low crude fiber content and high digestible nutrients in unit volume or unit weight, relative to coarse feed, and mainly comprises seeds of crops (grains, beans and seeds of oil crops) and by-products of processing.

Preferably, the basal feed comprises a concentrate.

When the requirement of high energy of the dairy cows or dairy goats in the lactation period is met by adding the concentrated feed, the pH value of the rumen is reduced, and the risk of subacute rumen acidosis is higher, so that the feed buffer is added into the concentrated feed to achieve better prevention and treatment effects.

In some preferred embodiments, the mass ratio of the basic feed to the feed buffer is 25-35: 1, such as, but not limited to, 25: 1, 26: 1, 27: 1, 28: 1, 29: 1, 30: 1, 31: 1, 32: 1, 33: 1, 34: 1 or 35: 1, and when the mass ratio of the basic feed to the feed buffer is within the above range, the mixed feed can have good effects of improving ruminant rumen acidosis and improving lactation performance on the basis of ensuring nutrition supply.

Preferably, the mass ratio of the basal feed to the feed buffer is 31: 1.

By further adjusting and optimizing the mass ratio of the basic feed to the feed buffering agent, the feed obtained by mixing has better effects of improving ruminant rumen acidosis and improving lactation performance on the basis of ensuring nutrition supply.

The present invention will be further described with reference to specific examples and comparative examples.

Example 1

The embodiment provides a feed buffer which mainly comprises the following components in parts by weight:

1 part of sodium bicarbonate, 10 parts of magnesium oxide, 5 parts of sodium acetate and 15 parts of sodium butyrate.

Example 2

10 parts of sodium bicarbonate, 1 part of magnesium oxide, 15 parts of sodium acetate and 5 parts of sodium butyrate.

Example 3

2 parts of sodium bicarbonate, 8 parts of magnesium oxide, 8 parts of sodium acetate and 12 parts of sodium butyrate.

Example 4

8 parts of sodium bicarbonate, 2 parts of magnesium oxide, 12 parts of sodium acetate and 8 parts of sodium butyrate.

Example 5

Example 6

12 parts of sodium bicarbonate, 15 parts of magnesium oxide, 3 parts of sodium acetate and 18 parts of sodium butyrate.

Comparative example 1

The comparative example provides a feed buffer, which mainly comprises the following components in parts by weight:

6 parts of magnesium oxide, 10 parts of sodium acetate and 10 parts of sodium butyrate.

Comparative example 2

6 parts of sodium bicarbonate, 10 parts of sodium acetate and 10 parts of sodium butyrate.

Comparative example 3

6 parts of sodium bicarbonate, 6 parts of magnesium oxide and 10 parts of sodium butyrate.

Comparative example 4

6 parts of sodium bicarbonate, 6 parts of magnesium oxide and 10 parts of sodium acetate.

Examples of the experiments

The 12 Saaner dairy goats (39 +/-7 kg, Mean +/-SEM kg and 2-3 weeks after delivery) with similar body weight and lactation period used in the experimental example have good health conditions, are purchased from test farms of northwest agriculture and forestry science and technology university and are bred in animal physical and chemical key laboratory of agriculture department of Nanjing agriculture university.

After all animals are adapted to one week by high-precision feeding, rumen fistula and liver blood fistula are respectively arranged on the test animals, and the test animals recover for 2 weeks after the operation. In order to keep hepatic vascular fistula open, 0.3mL (500IU/mL) of heparin sodium normal saline solution is injected into each fistula every 8h until the test is finished.

After the recovery period, the lactating goats were randomly divided into 11 groups of 6 High concentrate group (HG) and feed buffer groups 1-10 (BG, 1-10 correspond to inventive examples 1-6 and comparative examples 1-4). The animals in the two groups were fed the same basic ration, the mass ratio of the fed fine and coarse ration was 60: 40, and the formula is shown in table 1. Buffer

The formulation was supplemented with the feed buffers provided in examples 1-6 of the present invention and comparative examples 1-4 on a basal ration basis. The test period is 20 weeks, and the animals are raised in a single cage, during which water is freely drunk.

TABLE 1 test diet and nutritional composition

Note: a. every kilogram of daily ration provides VD 2500 IU, VA 6000 IU, VE 80mg, copper 6.25mg, iron 62.5mg, zinc 62.5mg, manganese 50mg, iodine 0.125mg and cobalt 0.125 mg; b. nutrition levels were calculated according to the NRC (2001) method. Note: rendered per kg of die: vitamin A6,000 IU, vitamin D2,500 IU, vitamin E80 mg, Cu 6.25mg, Fe 62.5mg, Zn 62.5mg, Mn 50mg, I0.125 mg, Co 0.125mg. b Nutrient levels measured and recorded to National Research Council methods (2001).

The duration of the experiment was 2 milkings per day (time: 8: 00 am and 18: 00 am). Milk was discarded before 3, weighed after milking and milk production recorded. Rumen fluid was collected through the rumen fistula once every two weeks for pH determination. The results are shown in table 2 below.

TABLE 2 milk yield and rumen pH measurements

Group of	Milk yield	Rumen pH
			High concentrate group	1.15±0.12	5.70±0.12
Feed buffer group 1	1.22±0.10	6.18±0.13
			Feed buffer group 2	1.21±0.11	6.16±0.16
Feed buffer group 3	1.25±0.13	6.26±0.13
			Feed buffer group 4	1.24±0.21	6.25±0.11
Feed buffer group 5	1.28±0.15	6.40±0.14
			Feed buffer group 6	1.20±0.17	6.13±0.10
Feed buffer group 7	1.18±0.16	6.14±0.12
			Feed buffer group 8	1.17±0.15	6.11±0.17
Feed buffer group 9	1.19±0.13	6.13±0.13
			Feed buffer group 10	1.16±0.10	6.12±0.11

From the above results, it can be seen that the feed buffers provided in examples 1 to 6 of the present invention have good effects of improving rumen acidosis and improving lactation performance in ruminants due to the synergistic combination of the specific components. The feed buffers provided in comparative examples 1 to 4 were not as effective as those of the examples of the present invention in terms of milk production and rumen pH improvement.

The feed buffers provided in examples 1-6 are completely the same in raw material components and only different in proportioning, but the feed buffers provided in examples 1-5 are superior to those in example 6 in both milk yield and rumen pH improvement, which indicates that the feed buffers provided in the present invention have better effects of improving ruminant rumen acidosis and improving lactation performance under the preferable proportioning conditions of the present invention.

Moreover, the effect of example 5 is better than that of examples 1-4, and the effect of examples 3 and 4 is better than that of examples 1 and 2, which shows that the feed buffer provided by the invention has better effects of improving ruminant rumen acidosis and improving lactation performance by further adjusting and optimizing the component ratio.

The feed buffer provided in example 5 and comparative examples 1 to 4 have the same mixture ratio of the same components, but the feed buffer provided in example 5 has better levels than those of comparative examples 1 to 4 in terms of milk yield and rumen pH improvement due to the absence of sodium bicarbonate in comparative example 1, magnesium oxide in comparative example 2, sodium acetate in comparative example 3 and sodium butyrate in comparative example 4, which shows that the feed buffer provided in the invention has better effects of improving ruminant rumen acidosis and improving lactation performance only through synergistic cooperation between specific components of each raw material under the condition that the mixture ratio of the components of the same raw materials is the same.

In order to save costs, example 5, which is superior in the above effects, was selected as the feed buffer group and the high-concentrate group for the test.

Milk sample collection and milk component content determination

The duration of the experiment was 2 milkings per day (time: 8: 00 am and 18: 00 am). Milk was discarded before 3, weighed after milking and milk production recorded.

The milk fat and lactose contents were measured once a week. Sending to milk industry detection center of Nanjing Weigang of Jiangsu province, and detecting milk fat, lactose, etc. content with milk component analyzer.

1. Influence of feed buffer on milk yield of lactating goat fed with high-precision feed

As shown in FIG. 1, the milk production of the two groups of goats at the first 2 weeks of the experiment was substantially similar, and the milk production of the lactating goats in the buffer group was higher than that in the high-formula group from week 3, wherein the milk production was significantly higher than that in the high-formula group from week 8 (1.26. + -. 0.14vs 1.15. + -. 0.12, P < 0.05), and continued until week 15 to 16-19, and the milk production of the lactating goats in the buffer group was also higher than that in the high-formula group, but the difference was not significant (P > 0.05).

2. Effect of feed buffer on the milk fat content of lactating goats

As can be seen from FIG. 2, the difference in the milk fat rates of the two groups of goats from week 1 to week 5 was small, and the trends were that the milk fat rates were increased first, then decreased, and then increased. Beginning at week 6, the milk fat content of the buffer group began to be higher than that of the high-profile group, wherein the milk fat rate of the buffer group reached a maximum of 3.60 ± 0.13% at week 10, and the milk fat rate of the buffer group began to decrease at week 18.

3. Effect of Complex buffer on lactose content in lactating goats

As can be seen from fig. 3, the lactose content was substantially consistent for the first 3 weeks of the experiment. The lactose content in the buffer group was higher than that in the high-concentrate group starting at week 4, the lactose content reached the highest at week 6 and was significantly higher than that in the high-concentrate group (3.42. + -. 0.12 vs. 2.80. + -. 0.11, P < 0.05), and the buffer group was higher than that in the high-concentrate group by week 7 to the end of the experiment.

Second, blood sample collection

Jugular vein blood was collected from the last 1d of the trial to week 19. The collected blood sample is quickly transferred into an anticoagulant tube containing heparin sodium, centrifuged for 12min at 4 ℃ and 2500 Xg, and serum is separated into an EP tube and stored at-20 ℃ to be tested.

1. Blood biochemical index determination

Taking 1mL of the collected jugular vein blood sample, sending to a department of hematology of the Chinese and Western medicine combination hospital in Nanjing, and detecting biochemical indexes of blood by using a full-automatic biochemical analyzer, wherein the biochemical indexes include indexes such as Alanine transaminase (ALT), Glutamic-oxaloacetic transaminase (AST), Total Bilirubin (TB), Alkaline phosphatase (AKP), Triglyceride (Triglyceride, TG), Lactate Dehydrogenase (LDH), Hypersensitive C-reactive protein (HS-CRP) and the like.

As shown in Table 3, the buffer group has lower ALT, AST and AKP content than the high concentrate group in the blood of the lactating goats, and the difference is significant (P is less than 0.05), which indicates that the liver injury state of the lactating goats is relieved. The content of TB in the blood of the buffer group is obviously higher than that of the high-precision group (P is less than 0.05), and no obvious difference exists between the two groups of other indexes.

TABLE 3 Effect of Complex buffers on biochemical indices related to lactation goat blood (n ═ 6)

2. Blood hormone index detection

Collecting jugular vein blood sample, and determining related hormone content by adopting ELISA detection kit of Insulin (INS), glucagon, cortisol and insulin-like growth factor-1, wherein the test steps are carried out according to the kit instruction. The results are shown in Table 4. Generally, the endocrine hormones in the animal body are at a normal physiological level under normal conditions. Mainly secreted by some specific organs and entered into the circulating blood in a dispersed manner to play a role in the target tissue. As can be seen from table 4, the complex buffer resulted in an increase in the amount of INS in the blood. INS can regulate glycolipid metabolism in the body. The increase of INS content in blood suggests that it may play a role in the major organs (liver) of body's glycolipid metabolism.

Table 4 effect of complex buffer on the blood hormone content of lactating goats (n ═ 6)

Note: indicates significant difference (P < 0.05) compared to the high concentrate group.

Note：Compared with the high concentrate group，*indicates a significant difference(P＜0.05).

3. Measurement of blood metabolic abnormality product and inflammatory factor

Measuring histamine in jugular vein blood by adopting an ELISA method, measuring the absorbance content of each hole at the wavelength of 450nm, and calculating the concentration of a sample by a regression equation, wherein the unit is expressed by ng/mL; the lactic acid content is determined by a lactic acid dehydrogenase method, the absorbance content of each hole is determined at the wavelength of 530nm, and the concentration of the sample is calculated by a regression equation, wherein the unit is expressed by mmol/L. The operation steps are respectively carried out according to the relevant specifications. The results are shown in FIGS. 7A and 7B.

Inflammatory cytokines TNF-alpha and IL-1 beta in jugular vein blood are detected by a 125I radioimmunoassay kit. The operation steps are carried out according to the instruction. As shown in fig. 7C and 7D, the study found that high-quality diet can induce SARA in the body of lactating goats, the increased LPS in rumen fluid enters the blood through the rumen wall, and the contents of lactic acid and TNF-alpha in the blood are significantly increased, which causes inflammation in the body. After the compound buffer is added, no SARA state appears, and simultaneously, the contents of LPS released by rumen, metabolic abnormal products and inflammatory factors in blood are obviously reduced. The composite buffering agent can relieve the decrease of rumen pH value caused by high-precision feeding, reduce the release of LPS and the like, and improve the health state of organisms.

Thirdly, collecting rumen fluid and measuring pH

1. Determination of the pH of rumen fluid

Rumen fluid was collected through the rumen fistula once every two weeks for pH determination. The specific method comprises the following steps: rumen contents are collected from 15min before ingestion and 0, 2, 4, 6, 8 and 10h after feeding, and rumen fluid is filtered and collected by sterilized medical gauze. The pH was determined immediately and three consecutive measurements were made per test animal.

By detecting the average pH values of rumen fluid at 0, 2, 4, 6, 8 and 10h after 2-18 weeks of feeding, the result is shown in figure 4, and the total change trend of rumen pH value is firstly decreased and then increased. After the feeding, the pH value is in a descending trend, after 4 hours, the pH value is reduced to the minimum value (5.65 +/-0.03), and then the pH value is gradually increased to the initial level due to the strong buffering capacity of the rumen. However, the pH value of the rumen fluid of the lactating goats in the buffering agent group is higher than that in the high-precision material group (6.0 +/-0.10 vs 5.8 +/-0.08).

When SARA is taken as the time that the pH value of the rumen is less than 5.8 per day and is not less than 4h, SARA phenomenon begins to appear after the high-feed group of lactating goats eat the feed for 2 h. And after the compound buffering agent is added, the decrease of the pH value of the rumen caused by high-precision materials is relieved, so that the pH value of rumen fluid is stabilized.

2. Determination of rumen content and LPS content in blood

Diluting rumen fluid and blood sample by 5 times with sample diluent in the kit, loading according to LPS enzyme-linked immunoassay kit instruction, sequentially determining absorbance value (OD) of each well at 450nm wavelength, and calculating LPS content (activity unit is EU/mL) in rumen fluid and blood by using standard curve equation with blank well zeroing as reference. LPS standard curve equation: 1445.5x-92.211, R²0.9995(y represents concentration and x represents OD value).

By detecting the contents of LPS in gastric juice and blood of two groups of lactating goats, the results of the two groups of rumens and blood show that the content of LPS (26201.41 +/-2398.18 vs 40395.43 +/-4723.14, P < 0.01) in rumens of the buffer group is remarkably lower than that of the high-concentrate group, and the content of LPS (2.01 +/-0.24 vs 3.62 +/-0.50, P < 0.05) in blood is remarkably lower than that of the high-concentrate group in the buffer group. The pH level of the rumen of the lactating goat is stabilized by adding the compound buffering agent, so that the generation of an abnormal product LPS in the rumen is reduced.

3. Determination of Volatile Fatty Acid (VFA) content in rumen fluid and blood

And (3) rumen fluid treatment: adding 1mL rumen fluid into EP tube, adding 25% metaphosphoric acid mixed solution (containing 0.2mL crotonic acid), mixing well, storing at-20 deg.C overnight, standing at room temperature for thawing, centrifuging at 4 deg.C and 12000 Xg for 10min, and filtering supernatant with needle filter (0.22 μm).

Blood treatment: adding 200 μ L of plasma into EP tube, adding 200 μ L of 5% sulfosalicylic acid mixture (precipitated protein) containing crotonic acid, mixing, storing at-20 deg.C overnight, centrifuging at 4 deg.C and 12000 Xg for 30min, collecting supernatant, filtering with needle filter, and testing. As shown in tables 5 and 6, the present study found that the addition of complex buffer not only improved the decrease in rumen pH due to the excessive concentrate ratio, but also changed the content and ratio of VFA in rumen and blood. The main reasons for this are the buffering effect of sodium bicarbonate on the one hand, and the weak acids of acetic acid and butyric acid on the other hand, which combine with hydrogen ions in the rumen to form weak acids after entering the rumen, so as to inhibit the decrease of the pH value. And the content of butyric acid in blood is also obviously increased by the absorption of rumen epithelium. The increase in blood butyrate concentration in this test. Probably due to the addition of sodium butyrate to the feed. Butyric acid, as a main component of sodium butyrate, is increasingly important in the aspects of gastrointestinal health and anti-inflammation at present, and can relieve damage of liver cells caused by high-concentrate feeding. Therefore, elevated butyrate in blood may play a role in protecting the health of the body.

Table 5 effect of complex buffer on rumen VFA of lactating goats (n ═ 6)

Table 6 effect of complex buffer on blood VFA of lactating goats (n ═ 6)

Fourthly, determination of mammary gland oxidative stress index and tissue apoptosis related protein

1. The antioxidant stress index measured in the test comprises glutathione peroxidase (GSH-PX), Superoxide dismutase (SOD), Catalase (CAT), Total antioxidant capacity (T-AOC) and Malondialdehyde (MDA). The measurement of various indexes is carried out by adopting an enzyme-linked immunosorbent assay. The kits were purchased from Nanjing, institute for bioengineering.

As shown in Table 7, the antioxidant stress indexes SOD, CAT and T-AOC of the mammary gland of the lactating goat in the buffer group are all higher than those in the high-precision group, wherein the CAT and the T-AOC are significantly higher than those in the high-precision group (P is less than 0.05). While the oxidative stress index MDA is obviously lower than that of the high concentrate group (P is less than 0.05); the buffer group GSH-PX was lower than the high concentrate group, but there was no significant difference (P > 0.05). The fact that the compound buffering agent is added improves the breast oxidative stress state of the lactating goats induced by high-precision feeding.

TABLE 7 Effect of Complex buffers on oxidative stress indicators of lactating goat mammary glands (n ═ 6)

2. Western Blot detection of mammary tissue apoptosis-related protein

Wherein the Caspase-3, Caspase-9 and Bcl-2 primary antibodies are diluted by 1: 1000 times; diluting the Bax primary antibody by 1: 2000 times; the internal control GAPDH is diluted by 1: 5000 times.

(1) Protein sample extraction

Preparing precooled RIPA lysate (containing PMSF with concentration of 1 mmol/L), taking about 80mg of liver, and homogenizing on ice for about 5 min; standing for 30min, centrifuging at 4 deg.C and 12000 Xg for 20min in a high speed centrifuge, and collecting supernatant. The protein concentration was determined by total protein quantitation, with the steps performed strictly according to the instructions. Adjusting the protein concentration to the same concentration, and subpackaging in EP tubes at-80 deg.C for use.

(2) SDS-PAGE electrophoresis and Wet transfer

Separating gel (10%) and concentrating gel (5%) were prepared. And (3) sampling 60 mu g of denatured protein, carrying out 80V constant-pressure ice bath electrophoresis for 30min by using a protein standard substance marker with the sampling amount of 4 mu L, and then changing to 110V constant pressure until the ice bath electrophoresis is finished.

Intercepting the gel at the corresponding position of the target protein, and cutting the PVDF membrane according to the size of the gel. Activating the cut PVDF membrane by methanol for 10-20s, and then putting the gel, the filter paper and the PVDF membrane into the transfer printing liquid for soaking for 10 min. From the positive electrode to the negative electrode are: filter paper, PVDF membrane, gel, filter paper, wet transfer printed for 1.5h with 100V constant pressure.

(3) Blocking and antibody incubation

And (3) sealing: the transferred PVDF membrane was removed, and the protein was blocked with skimmed milk powder (5%) blocking solution on a shaker at room temperature for 2 h.

Antibody incubation: after the blocking is finished, the blocking solution is discarded. Rabbit anti-primary antibody was diluted 1: 1000 times, GAPDH was diluted 1: 5000 times, and incubated overnight at 4 ℃. After incubation of primary antibody was complete, primary antibody was gently aspirated and membrane washed 5 times with TBST for 10min each. Then, a secondary goat anti-rabbit antibody, which was diluted with TBST at 1: 10000 times, was added and incubated at room temperature for 2 h. After the secondary antibody incubation is finished, the membrane washing process is repeated.

(4) Color development and analysis

After the membrane is sucked dry by filter paper, ECL luminous liquid is quickly added, after the membrane is kept out of the sun for 5min, the membrane is developed by a full-automatic chemiluminescence image analysis system, and the gray value of each group of strips is observed. The band intensity values were analyzed by Image J software to calculate the relative expression levels of the respective target proteins.

Results are shown in fig. 6A, 6B, 6C and 6D, Caspase-3(0.65 ± 0.05 vs. 1.00 ± 0.25) and Bax (0.74 ± 0.08 vs. 1.00 ± 0.22) protein expression was significantly down-regulated (P < 0.05) in mammary glands of lactating goats after the addition of complex buffer; and the expression level of the anti-apoptotic protein Bcl-2(1.52 +/-0.18 vs 1.00 +/-0.11) is obviously up-regulated (P < 0.05). The result of the oxidative stress index is combined to prompt that the high-precision feed feeding induces the oxidative stress of the mammary gland of the lactating goat to cause the apoptosis injury of the mammary gland. The addition of the compound buffer can improve the stress or apoptosis injury of mammary glands induced by high-precision feeding of the lactating goats.

Finally, it should be noted that: the above embodiments are only used to illustrate the technical solution of the present invention, and not to limit the same; while the invention has been described in detail and with reference to the foregoing embodiments, it will be understood by those skilled in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some or all of the technical features may be equivalently replaced; and the modifications or the substitutions do not make the essence of the corresponding technical solutions depart from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A feed buffer, characterized in that said feed buffer consists essentially of:

sodium bicarbonate, magnesium oxide, sodium acetate, and sodium butyrate.

2. The feed buffer of claim 1, wherein the feed buffer consists essentially of the following components in parts by weight:

3. The feed buffer of claim 2, wherein the feed buffer consists essentially of the following components in parts by weight:

4. The feed buffer of claim 3, wherein the feed buffer consists essentially of the following components in parts by weight:

5. The method for preparing a feed buffer according to any one of claims 1 to 4, wherein the feed buffer is obtained by uniformly mixing sodium bicarbonate, magnesium oxide, sodium acetate and sodium butyrate in parts by weight of a formulation.

6. Use of a feed buffer according to any of claims 1-4 or prepared by the preparation method of claim 5 for the preparation of a product for the prevention and/or treatment of ruminal acidosis in a ruminant.

7. Use according to claim 6, wherein the product comprises a feed.

8. A feed comprising a basal feed and a feed buffer according to any one of claims 1 to 4 or prepared by the preparation method according to claim 5.

9. The feed of claim 8, wherein the basal feed comprises a roughage and/or a concentrate;

preferably, the basal feed comprises a concentrate.

10. The feed according to claim 8 or 9, wherein the mass ratio of the basal feed to the feed buffer is 25-35: 1, preferably 31: 1.