CN117179146A - Methane inhibitor and application thereof in feed preparation - Google Patents

Abstract

The invention discloses a methane inhibitor, which comprises isosorbide dinitrate. The invention also discloses an application of the methane inhibitor in feed preparation, wherein the methane inhibitor comprises isosorbide dinitrate. The methane inhibitor can continuously inhibit the generation of methane, and simultaneously, the hydrogen is effectively solved and utilized.

Description

A methane inhibitor and its application in feed preparation

Technical field

The present invention relates to the technical field of animal husbandry, and more specifically to a methane inhibitor and its application in feed preparation.

Background technique

Because the rumen of ruminants cannot secrete gastric juice and is insulated from air, which provides a comfortable growth environment for methanogens, ruminants will produce a large amount of methane during repeated chewing. In addition, some methane emissions are also produced when ruminants experience flatulence.

The study found that from 1961 to 2019, the global methane emission reduction from animal husbandry showed an overall upward trend. Among them, methane emissions from ruminants increased from 68.0479 million tons to 103.5291 million tons, accounting for 97.5% of the total methane emissions from animal husbandry. Therefore, the key to reducing methane emissions from animal husbandry lies in ruminants, and the key to reducing ruminant emissions is to reduce methane emissions.

At present, domestic measures to reduce ruminant methane emissions include: First, changing the nutritional composition of ruminant feed, controlling the proportion of acetic acid and propionic acid, and increasing the number of feedings (such as feeding a small amount once, adding multiple supplements ), the feed is crushed or granulated; the second is to reduce methane emissions through anthelmintic technology, such as adding lipoic acid or rumen protective fat containing saturated fatty acids to the diet to expel protozoa or reduce the number of protozoa and provide electron acceptors (such as succinic acid, fumaric acid and other intermediate metabolites in the tricarboxylic acid cycle), adding plant extracts (such as tannins, saponins and plant essential oils) to promote the production of propionic acid and thereby reducing methane production, adding halogen compounds and derivatives (such as bromochloromethane, bromoethane sulfonic acid, chlorinated fatty acids, nitrites, etc.) and adding ionophores (such as monensin, lasalocid, etc.) to regulate the generation of methane.

However, these methods of regulating methane production have problems such as short duration, microbial adaptability and tolerance, high host specificity, reduced fiber digestibility, and toxicity. Therefore, their application in ruminant production is limited. sex.

Therefore, how to regulate methane production in ruminants is an urgent problem that those skilled in the art need to solve.

Contents of the invention

In view of this, the object of the present invention is to provide a methane inhibitor and its application in feed preparation to solve the deficiencies in the prior art.

In order to achieve the above objects, the present invention adopts the following technical solutions:

A methane inhibitor including isosorbide dinitrate.

Furthermore, the above-mentioned methane inhibitor also includes L-malic acid.

Furthermore, the mass ratio of the above-mentioned isosorbide dinitrate and L-malic acid is 1:50.

The present invention also claims the use of a methane inhibitor in feed preparation, wherein the methane inhibitor includes isosorbide dinitrate.

Furthermore, the addition amount of the above-mentioned isosorbide dinitrate is: 200 mg of isosorbide dinitrate is added per 1kg of feed dry matter.

Furthermore, the above-mentioned methane inhibitor also includes L-malic acid.

Furthermore, the addition amount of the above-mentioned L-malic acid is: 10g L-malic acid per 1kg of feed dry matter.

It can be seen from the above technical solutions that compared with the prior art, the beneficial effects of the present invention are as follows:

1. The applicant’s research found that the nitrooxy group in isosorbide dinitrate can selectively bind to methyl-coenzyme M reductase and temporarily oxidize nickel ions from +1 to +2 at the active site. The valence is used to inactivate its enzyme, thereby inhibiting the last step in the methane production process, thereby continuously inhibiting the production of methane, and at the same time producing a large amount of hydrogen. L-malic acid is the electron carrier of hydrogen, and it can be used by strengthening the malic acid shuttle system, α-glycerol phosphate shuttle system and tricarboxylic acid cycle pathway in the process of biological oxidation (the pathway in which reducing equivalents are oxidized during rumen fermentation). Hydrogen is oxidized by the animal body to generate ATP, and generates more additional energy for the animal body to be used in animal production, so that hydrogen gas can be effectively solved and utilized.

2. Based on the background that methane can cause global warming and hydrogen is a high-energy gas, the present invention selects isosorbide dinitrate (ISD) as a methane emission inhibitor and observes whether it has the effect of reducing methane emissions. At the same time, ISD plus L-malic acid (ISD×MAL) was selected as another treatment group to observe whether H ⁺ was oxidized by cell mitochondria into adenosine triphosphate (ATP) to provide energy for the body. Therefore, the present invention aimed to determine the effects of ISD and ISD×MAL on methane emissions, milk production, ruminal fermentation and milk production performance of mid-lactation dairy cows.

Detailed ways

The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only some of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts fall within the scope of protection of the present invention.

Example 1

Methane inhibitors, including isosorbide dinitrate.

Example 2

Methane inhibitors include isosorbide dinitrate and L-malic acid; the mass ratio of isosorbide dinitrate and L-malic acid is 1:50.

Performance Testing

The experiment was completed at the Teaching Experimental Base of Henan Agricultural University from April 20, 2022 to August 12, 2022, using 9 animals with the same age, weight (659±20kg), lactation period (115±10 days) and milk production ( A repeated 3×3 Latin square design study was conducted on multiparous Holstein cows (3 parities) of similar size (24.4±1.6kg), with 3 phases including: each phase contained a 21-day adaptation period and a 7-day sample collection period. All cows were housed in a shady open barn and assigned to one of three blocks. Since there were not enough cows to support the study, the entire research protocol was conducted with a 7-day interval between each trial cycle before the next cycle.

Before the start of the formal trial, a 10-day pre-test period was conducted for all cows to make the physiological status of the cows basically the same before the start of the formal trial. In this study, all cows were milked twice a day at 6:00 and 18:00 and fed total mixed rations (TMR) twice a day at 7:00 and 19:00. During the entire trial, the cows could Have free access to food and water. Cows were fed TMR without isosorbide dinitrate (ISD) and L-malic acid (MAL) as the control group (CON), cows were fed ISD with 200 mg/kg feed dry matter as the ISD treatment group, and cows were fed 200 mg/kg feed dry matter The ISD of kg feed dry matter plus the MAL of 10 g/kg feed dry matter was used as the ISD×MAL treatment group, in which powdered ISD and MAL feed additives were added to the animals’ diet in the form of premixes mixed with TMR to support dairy cows Consumption of additives throughout the day. Cows have free access to food and water.

Milk production was continuously recorded from days 22 to 28 of each cycle, and milk samples were collected in the morning and evening from days 27 to 28 (6:4) and divided into two portions through two 50 mL sterile test tubes. . The first milk sample was mixed with preservatives and stored at 4°C; the other milk sample was stored at -20°C until used for analysis of milk composition and fatty acids. In addition, the index of enzyme activity in the serum of dairy cows is to collect blood from the tail vein of the cows with a disposable venous blood collection needle and a negative pressure blood collection tube before feeding the cows at 7 a.m. on the 26th and 27th days, and then incubate at 4°C Centrifuge at 2000 × g for 20 min, remove the serum with a plastic pipette and store it at -30°C for enzyme activity analysis (for example: MDH = malate dehydrogenase, PCK = phosphoenolpyruvate carboxykinase, PK = acetone Acid kinase, CS = citrate synthase). One milk sample was sent to the Henan Provincial DHI Measurement Center for determination of milk composition and milk urea nitrogen, and the other milk sample was sent to Qingdao Yixin Testing Technology Service Co., Ltd. to detect the residue and metabolism of fatty acids and isosorbide dinitrate in the milk. Objects, in which the concentration of fatty acids was detected using a Shimadzu GC-2010 gas chromatograph. Ruminal digestive juices were collected through ruminal cannula before feeding on days 22 and 23. Immediately after that, the collected digestive juice was thoroughly mixed, and the mixed rumen digestive juice was injected into a 10mL cryovial via a 10mL sterile syringe, and quickly stored in -80°C liquid nitrogen to facilitate subsequent analysis of rumen microorganisms. At 7:00 on the 22nd and 23rd days after feeding, rumen digestive juices were collected through rumen intubation and mixed, and then the mixed rumen digestive juices were filtered with a 2-layer filter to obtain filtered rumen fluid samples. Filtered rumen fluid samples were placed into 50 mL centrifuge tubes, centrifuged at 4000 × g for 20 min at 4°C, and then immediately stored at -20°C until analysis of volatile fatty acids (VFA). The method for measuring intestinal methane emissions is to use sulfur hexafluoride tracer gas technology. The daily intestinal methane emissions of dairy cows can be obtained through the following formula: QCH ₄ =QSF ₆ ×([CH ₄ ] _y -[CH ₄ ] _b )/[SF ₆ ]. The methane emission rate ( _QCH4 ) can be calculated from the measured [ _CH4 ] _y and _SF6 concentrations in the yoke, the background [ _CH4 ] _b concentration in the yoke, and the known _QSF6 release rate.

Effects of adding isosorbide dinitrate (ISD) and isosorbide dinitrate plus L-malate (ISD×MAL) to the diet on dry matter intake (DMI), methane emissions, milk production, feed efficiency, and milk production of dairy cows The effects of ingredients and serum enzyme activity are shown in Table 1.

The effects of adding isosorbide (ISD) and isosorbide plus malic acid (ISD×MAL) in mid-lactation on rumen fluid volatile fatty acids (VFA) and rumen microbial profile counts are shown in Table 2.

The effects of adding isosorbide dinitrate (ISD) and isosorbide dinitrate plus L-malic acid (ISD×MAL) to the diet on fatty acids in the milk of mid-lactation cows (g/mL of total fatty acids) are shown in Table 3.

Table 1

In Table 1, ¹ CON=Control.

² C vs I, CON vs ISD; I vs I×M, ISD vs ISD×MAL.

³ ECM (kg/d) energy-corrected milk = kg of milk production milk production × (383 × fat% milk fat + 242 × protein% milk protein + 165.4 × lactose% lactose + 20.7) ÷ 3,140 (Sjaunja et al., 1990 ).

⁴ Milk NE _L (Mcal/d) Net milk production energy = kg of milk production milk production × (0.0929 × fat% milk fat + 0.0563 × protein% milk protein + 0.0395 × lactose% lactose) (NRC, 2001).

⁵ ECM yield energy-corrected milk yield ÷ dry matter intake (DMI) dry matter intake.

⁶ Milk yield ÷DMI dry matter intake.

⁷ MDH=malate dehydrogenase; PCK=phosphoenolpyruvatecarboxykinase; PK=pyruvate kinase; CS=citratesynthase.

Table 2

In Table 2, ¹ CON=Control.

² C vs I, CON vs ISD; I vs I×M, ISD vs ISD×MAL.

table 3

In Table 3, ¹ CON=Control.

² C vs I, CON vs ISD; I vs I×M, ISD vs ISD×MAL.

³ ΣSFAs = Total saturated fatty acids; ⁴ ΣMUFAs = Total monounsaturated fatty acids; ⁵ ΣPUFAs = Total polyunsaturatedfatty acids.

It can be seen from Table 1-3:

1. The methane emission reduction effect of the existing technology is instantaneous, while the ISD treatment (200mg/kg feed dry matter) and ISD×MAL treatment (ISD: 200mg/kg feed dry matter plus L-malic acid: 10g/kg feed dry matter Substances) can continuously reduce methane emissions in the intestines of dairy cows. Moreover, ISD treatment did not affect dry matter intake and milk production, and ISD×MAL treatment reduced dry matter intake but did not affect milk production and increased milk protein and milk fat concentrations.

2. Both ISD treatment and ISD×MAL treatment reduced the ratio of acetic acid to propionic acid and tended to reduce the copy number of methanogens, which was also the result of reduced methane emission.

3. ISD treatment increased the concentration of short-chain fatty acids and total SFA, and reduced the total amount of monounsaturated fatty acids (MUFA) due to a small part of biohydrogenation in the rumen. Compared with ISD treatment, ISD×MAL treatment increased the concentrations of short-chain fatty acids, total saturated fatty acids, monounsaturated fatty acids and polyunsaturated fatty acids, indicating that ISD×MAL can improve the energy utilization of animals more than ISD treatment. .

The above description of the disclosed embodiments enables those skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be practiced in other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims (7)

1. A methane inhibitor, characterized by comprising isosorbide dinitrate. 2. A methane inhibitor according to claim 1, further comprising L-malic acid. 3. A methane inhibitor according to claim 2, characterized in that the mass ratio of the isosorbide dinitrate and the L-malic acid is 1:50. 4. The application of a methane inhibitor in feed preparation, characterized in that the methane inhibitor includes isosorbide dinitrate. 5. The application of a methane inhibitor in feed preparation according to claim 4, characterized in that the addition amount of the isosorbide dinitrate is: 200 mg of isosorbide dinitrate is added per 1kg of feed dry matter. 6. Application of a methane inhibitor in feed preparation according to claim 4 or 5, characterized in that the methane inhibitor further includes L-malic acid. 7. The application of a methane inhibitor in feed preparation according to claim 6, characterized in that the amount of L-malic acid added is: 10g L-malic acid is added to every 1kg of feed dry matter.