KR20220092204A - Composition for promoting synthesis of milk protein comprising L-phenylalanine, L-tryptophan and acetate - Google Patents

Abstract

The present invention relates to a composition for promoting synthesis of milk protein comprising L-phenylalanine, L-tryptophan, and acetic acid. According to the present invention, there is an effect of increasing the amount of milk protein without changing the feed intake and flow rate of cows.

Description

Composition for promoting synthesis of milk protein comprising L-phenylalanine, L-tryptophan and acetic acid {Composition for promoting synthesis of milk protein comprising L-phenylalanine, L-tryptophan and acetate}

The present invention relates to a composition for promoting milk protein synthesis comprising L-phenylalanine, L-tryptophan and acetic acid.

As consumers' interests change, the demand for high-protein, low-fat milk, cosmetics and cheese containing milk protein is increasing, and interest in increasing milk protein is also increasing. Accordingly, the crude oil price calculation system in Korea is changing to include milk protein content from 2014. In order to maximize the amount of milk protein, the crude protein (CP) content in the feed should be around 22-23% (NRC, 2001). It causes environmental pollution and reduced nitrogen utilization efficiency. Therefore, it is necessary to find a way to increase the amount of milk protein while limiting the nutrients. It is generally known that amino acids, energy sources, and growth hormones are required for the mechanism of milk protein synthesis (Kim et al., 2013). Since Korea currently prohibits the use of growth hormone in cows, attention has been paid to amino acids and energy sources.

Amino acids are not only basic units of proteins, but also physiologically active substances (Kim et al., 2013). Among amino acids that are not synthesized in the body of animals or synthesized in a small amount even if they are synthesized, they are called essential amino acids, which are called essential amino acids. .

Many studies have been conducted to increase cow's milk protein by adding essential amino acids, but these are concentrated on a few amino acids. While the present inventors were studying a method for increasing the content of milk protein in cow's milk, it was confirmed that the production of milk protein was increased when cow-derived mammary epithelial cells (MAC-T) were treated with tryptophan and phenylalin, or tryptophan, phenylalanine and acetic acid. Also, as a result of feeding tryptophan and phenylalin, or tryptophan, phenylalanine and acetic acid to cows, it was confirmed that the amount of milk protein increased and the number of somatic cells was decreased without changing the feed intake and flow rate of cows, thereby completing the present invention. .

Many studies have been conducted to increase cow's milk protein by adding essential amino acids, but these are concentrated on a few amino acids. While the present inventors were studying a method for increasing the content of milk protein in cow's milk, it was confirmed that the production of milk protein was increased when cow-derived mammary epithelial cells (MAC-T) were treated with tryptophan and phenylalin, or tryptophan, phenylalanine and acetic acid. Also, as a result of feeding tryptophan and phenylalin, or tryptophan, phenylalanine and acetic acid to cows, it was confirmed that the amount of milk protein increased and the number of somatic cells was decreased without changing the feed intake and flow rate of cows, thereby completing the present invention. .

It is an object of the present invention to provide a composition for promoting protein-in-oil synthesis in milking ruminants comprising phenylalanine, tryptophan and acetic acid.

Another object of the present invention is to provide a feed additive for promoting protein-in-oil protein synthesis in milking ruminants comprising the above-described composition.

Another object of the present invention is to provide a feed composition for promoting protein-in-oil protein synthesis in milking ruminants comprising the composition described above.

Another object of the present invention is to provide a method for increasing the protein-in-oil content of cows, comprising the step of feeding the above-described composition to the cows.

The present invention may provide a composition for promoting protein-in-oil protein synthesis in milking ruminants comprising phenylalanine, tryptophan and acetic acid.

The milk protein may be beta-casein (beta-casein).

The present invention may also provide a feed additive for promoting protein-in-oil protein synthesis in milking ruminants comprising the above-described composition.

The present invention may also provide a feed composition for promoting protein-in-oil protein synthesis in cows comprising the above-described composition.

The present invention may also provide a method for increasing the protein-in-oil content of cows, comprising the step of feeding the above-described composition to the cows.

The combination of phenylalanine, tryptophan and acetic acid of the present invention has the effect of increasing the amount of milk protein without changing the feed intake and flow rate of cows. In addition, there is an effect of reducing the number of somatic cells due to the anti-inflammatory effect in all treatment groups compared to the control group.

1 is a result of confirming the relative extracellular protein concentration in MAC-T cells when treated with metabolites and various amino acids for 72 hours.
Figure 2 is the result of confirming the relative β- casein mRNA expression in MAC-T cells when the metabolites and various amino acids are treated for 72 hours.

Hereinafter, the present invention will be described in detail.

Many studies have been conducted to increase cow's milk protein by adding essential amino acids, but these are concentrated on a few amino acids. While the present inventors were studying a method for increasing the content of milk protein in cow's milk, it was confirmed that the production of milk protein was increased when cow-derived mammary epithelial cells (MAC-T) were treated with tryptophan and phenylalin, or tryptophan, phenylalanine and acetic acid. Also, as a result of feeding tryptophan and phenylalin, or tryptophan, phenylalanine and acetic acid to cows, it was confirmed that the amount of milk protein increased and the number of somatic cells was decreased without changing the feed intake and flow rate of cows, thereby completing the present invention. .

The present invention may provide a composition for promoting protein-in-oil protein synthesis in milking ruminants comprising phenylalanine, tryptophan and acetic acid.

The phenylalanine and tryptophan may be L-phenylalanine and L-tryptophan, and phenylalanine (Phe) and tryptophan (Trp) are essential amino acids and cannot be synthesized by animal tissues, and even if synthesized, they are sufficiently synthesized as required. can't be As a result of treatment with tryptophan and phenylalanine in mammary epithelial cells, it was confirmed that the concentration of extra cellular protein was significantly increased (FIG. 1). When phenylalanine, tryptophan and acetic acid were treated, the expression level of β-casein was increased ( FIG. 2 ). Therefore, it was confirmed that the combinations that produce milk protein most efficiently in MAC-T cells are tryptophan and phenylalanine and tryptophan, phenylalani and acetic acid.

The tryptophan may be included in a concentration of 0.9 mM.

The phenylalanine may be included in a concentration of 0.6 mM.

The acetic acid may be included in a concentration of 0.3 mM.

The milk protein may be beta-casein (beta-casein).

The ruminant may be cattle, dairy cows, sheep, deer, giraffes or camels, preferably cows, but is not limited thereto.

In general, the nutrient digestion process of ruminants with a rumen consists of a first-stage fermentation process by microorganisms in the rumen and a second-stage digestion process by digestive enzymes in the lower digestive organs (fourth stomach, small intestine and large intestine, etc.). Ruminants need to be supplied through feed because amino acids are consumed as a raw material for microorganisms present in the rumen and, in the case of high-capacity or milking cattle, there is a lack of necessary nutrients. In addition, ruminants are decomposed by ruminant microorganisms in the rumen when synthetic amino acids are fed in the structure of the rumen, and almost all are used as nutrient sources for ruminant microorganisms. In the end, only up to 20% of the fed synthetic amino acids are transferred to the small intestine. Therefore, in ruminant animals, it is necessary to supply feed considering not only the digestibility and absorption rate in the rumen, but also the amount of nutrients that bypass the rumen and reach the small intestine and the absorption rate absorbed by enzymes in the small intestine. . Therefore, in order to maximize the production capacity of ruminants, it is necessary to create an environment that can produce as much microbial protein as possible in the first place, and at the same time to feed the protein protected from the decomposition action of ruminant microorganisms appropriately. However, since the microbial protein cannot meet the amount of amino acids required for maximizing the ability of ruminants, it is necessary to feed a feed protein that can be absorbed and used by going down to the small intestine without being decomposed by the ruminant microorganisms.

The present invention may also provide a feed additive for promoting protein-in-oil protein synthesis in milking ruminants comprising the above-described composition.

The present invention may also provide a feed composition for promoting protein-in-oil protein synthesis in cows comprising the above-described composition.

"Feed additive" means a material added to the feed, and the feed additive may be for improving productivity or health of the subject, but is not limited thereto. In addition, the feed additive may correspond to an auxiliary feed under the Feed Management Act.

In addition to phenylalanine, tryptophan and acetic acid, the feed additive form of the feed composition for promoting protein-in-oil protein synthesis in cows of the present invention may further include nutrients such as nucleotides, amino acids, calcium, phosphoric acid, organic acids, etc. can, but is not limited thereto.

The content of the phenylalani, tryptophan and acetic acid of the present invention can be determined by those skilled in the art in consideration of the feeding target, the feeding target species, body weight, feeding time, feeding feed type, feeding purpose, and the like.

As used herein, the term “feed” refers to any natural or artificial diet, meal meal, etc., or a component of the meal meal, for or suitable for being eaten, ingested, and digested by an individual. The type of feed is not particularly limited, and feed commonly used in the art may be used. Non-limiting examples of the feed include plant feeds such as grains, root fruits, food processing by-products, algae, fibers, pharmaceutical by-products, oils and fats, starches, gourds or grain by-products; and animal feeds such as proteins, inorganic materials, oils and fats, minerals, oils and fats, single cell proteins, zooplankton, or food. These may be used alone or in mixture of two or more.

The present invention may also provide a method for increasing the protein-in-oil content of cows, comprising the step of feeding the above-described composition to the cows.

Hereinafter, the present invention will be described in more detail through examples.

Example 1. Identification of the effect of a combination of a metabolite and essential amino acids on milk protein synthesis in mammary gland epithelial cells

Example 1-1. Confirmation of the effect of combination of metabolites and essential amino acids on extra cellular protein concentration in mammary gland epithelial cells

In order to confirm the effect of the combination of metabolites and essential amino acids on milk protein synthesis in mammary epithelial cells, various essential amino acids, metabolites and combinations thereof were used in mammary epithelial cells (immortalized bovine mammary epithelial cell line; MAC-T ( University of Vermont, Burlington, VT, USA))) to check the extra cellular protein concentration. Mammary gland epithelial cells (MAC-T) were cultured in growth medium (DMEM/F12 medium containing 10% FBS, 100 units/mL penicillin/streptomycin, 50 μg/mL gentamycin, 5 μg/mL insulin and 1 μg/mL hydrocortisone). After incubation, when about 80% of the plate was grown, it was subcultured to another plate. To test the change of extra cellular protein concentration, after subculture in 6 wells, when MAC-T cells were grown to 100% confluence, as a control, differentiation media (DMEM/F12 (Gibco) + 10% Fetal bovine serum) (Gibco) + 1% Penicillin/streptomycin (Hyclone) + 0.1% Gentamycin (Sigma-aldrich) + 0.1% Hydrocortisone (Sigma-aldrich) + 0.1% Insulin (Sigma-aldrich) + 0.1% Prolactin (Sigma-aldrich)) As a treatment group, differentiation media in which each essential amino acid was added for each concentration was cultured for 72 hours after dispensing. The treatment groups were as follows: 0.3 mM (acetate); 0.6 mM (Met, Lys, Phe), 0.9 mM (Trp, glucose), 1.5 mM (Ile, t-10, c-12 CLA). After culturing for 72 hours, extracellular proteins were extracted and analyzed using a BCA protein assay kit (Thermo Scientific, South Logan, UT, USA). Statistical significant differences according to the data mean values were performed using Tucky's HSD test of SPSS statistical software (SPSS Inc., Chicago, IL, USA). All experiments were performed in 3 repetitions, and it was judged to be significant at p < 0.05. As shown in Figure 1, the extracellular protein concentration was found to increase the most in the Trp-Phe treatment group among all the treatment groups.

Example 1-2. Effect of β-casein expression level confirmed by combination of metabolic stimulant and essential amino acid in mammary gland epithelial cells

In order to confirm the effect of the combination of metabolites and essential amino acids on milk protein synthesis in mammary epithelial cells, various essential amino acids, metabolites and combinations thereof were used in mammary epithelial cells (immortalized bovine mammary epithelial cell line; MAC-T ( University of Vermont, Burlington, VT, USA))) to confirm the expression level of β-casein. Mammary gland epithelial cells (MAC-T) were cultured in growth medium (DMEM/F12 medium containing 10% FBS, 100 units/mL penicillin/streptomycin, 50 μg/mL gentamycin, 5 μg/mL insulin and 1 μg/mL hydrocortisone). After incubation, when about 80% of the plate was grown, it was subcultured to another plate. After subculture in 6 wells to test the change in extracellular protein concentration, when MAC-T cells were grown to 100% confluence, as a control, differentiation media (DMEM/F12 (Gibco) + 10% Fetal bovine serum ( Gibco) + 1% Penicillin/streptomycin (Hyclone) + 0.1% Gentamycin (Sigma-aldrich) + 0.1% Hydrocortisone (Sigma-aldrich) + 0.1% Insulin (Sigma-aldrich) + 0.1% Prolactin (Sigma-aldrich)) with treatment As a result, the differentiation media in which each essential amino acid was added by concentration was cultured for 72 hours after dispensing. The treatment groups were as follows: 0.3 mM (acetate); 0.6 mM (Met, Lys, Phe), 0.9 mM (Trp, glucose), 1.5 mM (Ile, t-10, c-12 CLA). After culturing for 72 hours, MAC-T cells were sequentially treated with TRI reagent (MRC), chloroform, isopropanol, 80% ethanol, and 100% ethanol to extract RNA, and cDNA was synthesized from the extracted RNA. RT-PCR was performed with AccuPower 2X GreenStar qPCR MasterMix (Bioneer) to confirm the expression of bCSNB, a beta-casein synthesis gene, using cDNA. The bCSNB primer used at this time was forward(5'-GAGCCTGACTCTCACTGATGTTGAA-3'), reverse(5'-GACAGCACGGACTGAGGAGGAA-3'), and the bBAactin primer was forward(5'-GCATGGAATCCTGCGGC-3'), reverse(5'-GTAGAGGTCCTTGCGGATGT-3 '), and bUXT primers are forward(5'-GCGCTACGAGGCTTTCATCT-3'), reverse(3'-CCAAGGGCCACATAGATCCG-5'). RT-PCR was performed 40 times under the conditions of denaturing at 95° C. for 3 minutes and then reacting at 95° C. for 10 seconds, and reacted at 55° C. to 65° C. for 30 seconds and at 72° C. for 30 seconds. The expression level of β-casein was highest in Acetate-Trp-Phe treatment group among all treatment groups ( FIG. 2 ). Therefore, the combination that produces milk protein most efficiently in MAC-T cells is Trp-Phe and Acetate-Trp-Phe, and these combinations were confirmed through in vivo experiments.

Example 2. Confirmation of the milk protein synthesis effect of the combination of Tryptophan (Trp), Phenylalanine (Phe) and Acetate injected into the jugular vein of cows

As a result of in vitro results, the combination of Trp-Phe and Acetate-Trp-Phe was determined to be the most effective combination for milk protein synthesis. This study was conducted under the approval of the Animal Experimental Ethics Committee of Konkuk University. Five of Holstein-Friesian milking cows were used in the breeding experiments. Total mixed ration (TMR), concentrate, and hay were fed twice a day (0830 and 1700 h) according to the energy and protein requirements of NRC (2001) [Table 1].

chemical composition of feed Chemical compositions, g/d of DM TMR Concentrates Roughage DM (%) 61.91 88.84 85.88 — % of DM — Crude protein 9.79 20.33 9.61 Ether extract 1.79 3.10 3.71 Crude fiber 13.02 7.25 15.68 Crude ash 5.09 6.94 4.86 NDF 34.78 24.33 29.52 ADF 18.25 12.05 18.08 Ca 0.59 0.69 0.52 P 0.28 0.56 0.27 NE _L (Mcal/kg of DM) ³ 2.15 2.10 2.15 amino acids Tryptophan 0.10 0.21 0.15 ^* Threonine 0.36 0.67 0.24 ^* Serine 0.42 0.87 - Proline 0.53 1.06 - Valine 0.43 0.79 0.26 ^* Isoleucine 0.27 0.57 0.30 ^* Leucine 0.60 1.42 0.43 ^* Tyrosine 0.16 0.44 - Methionine 0.09 0.20 0.25 ^* Cysteine 0.21 0.46 0.06 ^* Lysine 0.40 0.59 0.34 ^* Glycine 0.39 0.80 - Alanine 0.48 0.94 - Arginine 0.53 1.14 0.28 ^* Glutamic acid 1.35 3.19 - Aspartic acid 0.79 1.51 - Histidine 0.16 0.37 0.21 ^* Phenylalanine 0.36 0.76 0.46 ^* *Book Value (from NRC, 2001)

Using a 5X5 Latin square design, 5 days of resting period and 6 days of the experiment were repeated. The test was performed with a total of 5 test groups: control (saline), Trp, Phe, Trp-Phe, and Acetate-Trp-Phe. 0.9 mM Trp, 0.6 mM Phe, and 0.3 mM acetate determined in the in vitro study were calculated by converting the molecular weight into grams. Amino acids and acetate powder were added to physiological saline after adjusting so as not to be affected by osmotic pressure and pH. The treatment solution was slowly injected using a jugular vein catheter at 10:00 h. The milking flow rate of each test animal (5 heads in total) in the morning (03:00 h) and in the afternoon (15:00 h) was recorded daily. During the experiment period of the test animals (total of 5), milk milked from milk was pooled every morning (0300 h) and afternoon (1500 h), and the general ingredients [milk fat, milk protein, lactose, solid-not fat (SnF), somatic cell count (SCC), milk urea nitrogen (MUN), acetone, beta-hydroxybutyrate (BHB), beta-casein, mono- and poly-unsaturated fatty acids, saturated fatty acid] were analyzed. In the analysis of feed intake, the daily intake was analyzed by recording the amount of feed remaining before feeding every morning. Blood was collected before injection of the treatment solution on days 1, 3, and 5 of the experiment through a jugular vein catheter. The daily grades obtained through each analysis were averaged for each period. The statistically significant difference according to the mean value of the data was determined by SAS software v. One-way ANOVA of 9.4 (SAS Institute, Cary, NC, USA) was used. Table 2 shows the effect of milk production and milk composition of dairy cows by the feeding of metabolites and amino acids. As a result of treatment with amino acids and acetic acid using a jugular vein catheter of milking cows, there was no change in feed intake and flow rate [Table 2]. Milk protein yield was significantly increased in the treatment group. The Trp-only treatment group increased the most among all the test groups, and thereafter, Acetate-Trp-Phe, Trp-Phe, Control, and Phe increased in the order. The number of somatic cells in oil was decreased compared to the control in all treatment groups, so it was judged that the additives suppressed the inflammatory response in the animal body.

Table 3 shows the effect of the feeding of amino acids on the CBC of cows on days 1, 3 and 5. The addition of amino acids and acetic acid via jugular vein had no effect on blood properties [Table 3]. It was confirmed that there was no problem in the health condition of the test axes.

The amino acid content in blood after 24 hours of injection of amino acids and acetic acid through the jugular vein is shown in Tables 4 and 5. There was no significant change in amino acid composition between treatments during the test period (Table 4). Since this is the result after 24 hours of jugular vein injection, it is considered that the injected amino acids have already been used for protein synthesis and body metabolism. There was no significant difference in the amino acid content on days 1, 3, and 5 of each treatment group (Table 5). In the control group, there was a significant difference in Val in the Ser and Trp-Phe treatment groups, and there was a tendency to change in the Trp in the Asp and Acetate-Trp-Phe treatment groups in the Phe treatment group. In the Phe treatment group, Asp increased on day 3 and then decreased again on day 5. It is thought that this is because Phe increases intestinal transport of amino acids including amide. Val decreased on day 3 by the Trp-Phe treatment group and then increased on day 5 (p = 0.02), which is partly due to competition because Trp, Phe, and Val pass through the same amino acid transporter (LAT1). could be an effect. Blood Trp showed a tendency to increase on day 3 and decrease again on day 5. The transport of amino acids requires energy, and it is thought that this change was caused by the time problem of making and using energy from acetate added to the blood.

Analysis of blood amino acid composition of dairy cows according to the feeding of metabolites and amino acids amino acids Control Trp Phe Trp-Phe Acetate
+
Trp-Phe SEM p -value Aspartic Acid 12.46 15.14 14.73 13.65 18.15 7.92 0.990 Threonine 102.48 77.32 85.18 71.43 95.99 19.75 0.791 Serine 105.72 61.00 100.46 109.80 91.83 14.99 0.191 Asparagine 105.52 132.08 67.52 134.88 104.05 30.04 0.528 Glutamic Acid 289.54 258.28 270.92 267.23 259.85 24.85 0.904 Glycine 345.54 350.85 353.29 322.84 352.12 21.79 0.848 Alanine 289.58 279.12 277.24 266.20 280.07 19.76 0.947 Valine 14.51 14.30 17.53 22.91 13.96 3.39 0.322 Cystine 41.46 37.59 41.84 42.48 40.19 7.85 0.993 Methionine 99.20 100.70 98.21 75.32 102.24 22.66 0.910 Phenylalanine 95.79 91.08 99.35 95.67 93.65 22.23 0.999 Isoleucine 25.57 28.34 30.76 26.92 27.28 8.19 0.994 Leucine 20.64 30.73 28.12 23.60 20.08 6.43 0.718 Tyrosine 16.30 21.22 21.21 24.71 21.93 5.59 0.878 Histidine 55.65 57.74 56.35 48.17 61.93 6.34 0.653 Tryptophan 10.37 18.55 19.66 17.59 14.04 4.83 0.652 Lysine 83.12 81.35 83.81 77.47 74.78 4.11 0.491 Arginine 105.36 99.36 104.95 127.21 105.02 14.76 0.710

Identification of the blood amino acid composition of cows fed with metabolites and aninoic acid for 1 to 5 days amino acids Control Trp Phe Trp-Phe Acetate+Trp-Phe Aspartic Acid D1 5.70 2.60 7.55 30.13 15.13 D3 26.23 36.50 28.72 4.19 43.10 D5 5.44 6.33 7.92 8.24 5.21 SEM 7.13 8.20 7.36 8.78 8.09 p -value 0.42 0.19 0.44 0.45 0.37 Threonine D1 83.17 63.55 84.05 65.92 80.27 D3 108.82 93.05 96.65 51.49 117.98 D5 115.46 75.35 74.84 58.58 118.47 SEM 16.24 16.11 16.57 14.28 16.61 p -Value 0.72 0.78 0.88 0.93 0.54 Serine D1 151.90 ^a 101.23 112.17 77.37 135.91 D3 61.33 ^b 25.47 120.32 161.99 43.37 D5 103.93 ^ab 56.29 68.87 75.25 106.04 SEM 15.32 13.87 17.28 21.21 15.55 p -Value 0.04 0.69 0.46 0.18 0.19 Asparagine D1 90.50 121.60 63.83 216.74 101.20 D3 136.33 64.93 105.24 74.75 80.71 D5 89.74 209.70 33.48 129.11 81.13 SEM 27.53 30.07 13.68 43.12 23.36 p -Value 0.76 0.14 0.09 0.42 0.39 Glutamic Acid D1 311.39 239.85 259.07 265.13 286.44 D3 259.50 307.62 331.16 266.91 216.60 D5 297.74 227.38 222.54 253.36 303.02 SEM 12.27 20.58 28.07 31.90 23.41 p -Value 0.21 0.24 0.29 0.99 0.54 Glycine D1 356.13 375.76 402.90 382.07 362.08 D3 341.44 355.66 392.27 287.92 384.06 D5 339.05 321.13 264.71 264.96 328.79 SEM 16.75 20.19 30.08 36.35 19.80 p -Value 0.92 0.57 0.11 0.39 0.11 Alanine D1 317.49 295.04 303.87 304.35 299.61 D3 278.16 266.32 302.66 243.53 291.13 D5 273.09 275.98 225.19 231.03 264.11 SEM 12.63 14.07 18.97 29.50 15.54 p -Value 0.31 0.73 0.15 0.58 0.13 Valine D1 12.41 13.20 7.77 22.86 ^ab 8.41 D3 13.90 11.16 19.35 8.02 ^b 15.34 D5 17.22 18.52 25.45 43.04 ^a 12.70 SEM 3.17 2.48 3.62 5.50 2.28 p -Value 0.84 0.49 0.13 0.02 0.41 Cystine D1 41.57 33.97 28.41 41.97 34.40 D3 34.99 36.60 51.59 43.16 40.60 D5 47.82 42.20 45.52 55.78 34.91 SEM 5.98 5.28 5.15 5.77 4.74 p -Value 0.71 0.83 0.17 0.61 0.96 Methionine D1 92.14 81.03 95.65 64.68 107.12 D3 117.37 111.96 125.20 78.66 148.86 D5 88.09 109.12 73.79 60.92 99.86 SEM 14.19 16.46 15.00 14.40 15.09 p -Value 0.69 0.73 0.40 0.89 0.65 Phenylalanine D1 81.86 86.61 86.00 88.45 57.22 D3 91.93 96.14 101.19 87.00 78.15 D5 113.59 90.50 110.86 135.83 95.98 SEM 14.15 17.30 11.15 16.72 12.14 p -Value 0.68 0.98 0.79 0.45 0.78 Isoleucine D1 28.22 22.37 14.32 26.27 10.43 D3 21.36 29.28 33.96 18.81 26.02 D5 27.12 33.39 43.99 47.84 29.10 SEM 6.17 5.85 20.81 7.58 5.43 p -Value 0.90 0.77 0.59 0.32 0.12 Leucine D1 26.34 31.38 17.05 25.40 12.37 D3 13.19 21.28 37.70 15.02 34.49 D5 22.40 39.52 29.62 40.07 15.07 SEM 5.37 6.30 5.27 8.21 4.89 p -Value 0.11 0.53 0.29 0.53 0.21 Tyrosine D1 20.63 24.08 13.88 25.08 24.46 D3 15.93 17.96 19.33 16.60 26.35 D5 12.33 21.63 30.42 31.41 21.21 SEM 3.20 4.35 4.85 5.60 3.32 p -Value 0.61 0.87 0.39 0.62 0.79 Histidine D1 62.76 61.64 66.45 55.22 62.06 D3 49.21 55.68 50.11 51.17 60.48 D5 54.99 55.90 52.51 51.84 59.09 SEM 4.31 2.65 5.57 7.68 6.32 p -Value 0.47 0.62 0.47 0.98 0.26 Tryptophan D1 11.16 10.93 14.41 13.83 13.87 D3 10.27 15.77 18.49 12.75 21.93 D5 9.67 28.96 26.08 31.12 10.48 SEM 1.11 5.19 5.26 5.99 2.08 p -Value 0.88 0.37 0.69 0.43 0.08 Lysine D1 85.47 82.95 85.03 83.19 74.80 D3 81.98 85.72 88.01 61.18 76.10 D5 81.89 75.37 78.40 83.88 76.84 SEM 4.04 6.00 4.18 5.64 4.81 p -Value 0.93 0.79 0.66 0.20 0.56 Arginine D1 114.88 97.15 101.01 115.58 108.10 D3 92.75 98.70 111.37 110.59 99.40 D5 108.46 102.22 102.48 105.38 123.90 SEM 6.16 6.41 6.87 13.95 7.74 p -Value 0.34 0.95 0.82 0.96 0.94

Through an in vitro study, the combination that produced the most efficient milk protein was identified as Trp-Phe and Acetate-Trp-Phe. Based on the above results, an in vivo experiment was performed. Amino acids and acetic acid injected into milking cows through a jugular vein catheter in vivo increased milk protein content without changing feed intake and flow rate of cows. In addition, the number of somatic cells was reduced in all treatment groups compared to the control group, which is considered to have an anti-inflammatory effect. The composition of amino acids in the blood did not show significant changes according to the addition of amino acids. The increase in milk protein was confirmed by applying the results of in vitro cell experiments to in vivo.

Claims (5)

A composition for promoting protein-in-oil protein synthesis in milking ruminants, comprising phenylalanine, tryptophan and acetic acid The method of claim 1,
The milk protein is beta-casein (beta-casein), a composition for promoting protein synthesis in milking ruminants. A feed additive for promoting protein-in-oil protein synthesis in milking ruminants, comprising the composition of claim 1 or 2. A feed composition for promoting protein-in-oil protein synthesis in milking ruminants, comprising the composition of claim 1 or 2. The step of feeding the composition of claim 1 or 2 to cows; A method for increasing the protein-in-oil content of cows, comprising a.

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