Method for improving slaughter performance of sheep and umami of mutton

Abstract

According to the method for enhancing the slaughter performance of the sheep and the umami substances in the mutton, the optimized feed is obtained by adding 20 g of sodium propionate into basic feed of the sheep per day, daily feeding is carried out, the palatability is high, the method is not limited to regions and feeding environment conditions, and the slaughter rate of the sheep and the umami of the mutton can be effectively improved. Sodium propionate is added to sheep feed to increase the level of umami substances (such as glutamic acid) in mutton. In addition, the supplementation of sodium propionate can also optimize and change the gastrointestinal flora composition, the metabolite content and the muscle metabolite level of the sheep. The method has potential application value in the field of sheep breeding.

Description

Technical Field

[0001] The invention relates to the technical field of breeding, and in particular to a production method for improving the freshness of mutton. Background Art

[0002] Sheep meat is known for its high protein and low cholesterol content, especially compared to pork and beef, and has higher calcium and iron content. However, feeding strategies have a significant impact on sheep's growth performance, metabolic processes in the body, etc.

[0003] Flavor is the feeling produced by food stimulating the senses, including taste and aroma. Among them, flavor mainly comes from substances such as organic acids, inorganic salts, inosinic acid, free amino acids and ribose, which are called flavor components. It is pointed out here that water-soluble and fat-soluble metabolites are related to taste, and it is emphasized that amino acid metabolites play a key role in regulating muscle growth and protein synthesis. The type of feed can affect the flavor of mutton to varying degrees. At present, more research is on the addition of plants with high protein content at different levels, fatty acid substances such as flaxseed rich in α-linolenic acid, etc. in the diet. Most volatile flavor compounds in meat are generated by the Maillard reaction. The Maillard reaction is a non-enzymatic browning reaction between free amino compounds and reducing sugars during heating. At present, there is no research on the use of quantitative probiotics to regulate the production of umami substances. Previous non-targeted metabolomics analysis has found some major metabolic pathways and differential metabolites such as N-acetyl-L-aspartate, N-acetyl-aspartyl-glutamate and acetyl phosphate.

[0004] In the prior art, 5'-inosinic acid disodium is added to feed to improve the flavor of pork and improve the quality of pork, but the amount added is small, which is prone to uneven mixing and the problem that pigs cannot ensure that they can eat. Therefore, the optimization effect can be achieved by adjusting feed additives. Multi-omics technology is used to investigate the changes in muscle metabolites and proteins during the growth of sheep, and a new theoretical basis is provided for improving the flavor characteristics of sheep, and the existence of regulatable flavor proteins and metabolites is determined. Therefore, this technical solution is of great significance in seeking more effective feeding strategies in this field. Summary of the invention

[0005] The purpose of the invention is to provide a method for enhancing the slaughtering performance of sheep and the umami substances in mutton. The method has high palatability, is not limited by regions and feeding environment conditions, and can effectively improve the slaughtering rate of sheep and the umami taste of mutton.

[0006] In order to achieve the above object, the present invention provides an application of sodium propionate in improving sheep slaughtering performance and mutton umami taste. The sheep slaughtering rate and the content of umami substances can be increased by supplementing sodium propionate in the basic diet.

[0007] Furthermore, sodium propionate can be used to increase the level of free amino acids, especially glutamate, in the basal diet.

[0008] By supplementing sodium propionate in the basic diet, the optimal amount of sodium propionate supplementation is 20g / head / day.

[0009] Furthermore, the above-mentioned basic diet, by weight, comprises 44 parts of corn, 5.24 parts of soybean meal, 10.93 parts of wheat bran, 34.96 parts of compound feed, 2.97 parts of premix, 1.05 parts of sodium carbonate, and 1.14 parts of sodium chloride; the compound feed, by weight, comprises 54-55 parts of flaked corn, 3 parts of grass meal, 7-9 parts of soybean meal, 8 parts of cottonseed meal, 20 parts of stone powder, 5.4-6 parts of calcium hydrogen phosphate, 0.25 parts of vitamin A, 0.15 parts of vitamin D3, and 0.3 parts of vitamin E; the premix, by weight, comprises 58 parts of corn, 7 parts of soybean meal, 10 parts of cottonseed meal, 8 parts of rapeseed meal, 12 parts of DOGS, 0.15 parts of vitamin A acetate, 0.03 parts of vitamin D3, and 0.3 parts of vitamin E. 0.8 parts, EDL-α-tocopherol acetate 0.16 parts, copper sulfate 0.15 parts, magnesium sulfate 0.6 parts, zinc sulfate 1.2 parts, manganese sulfate 0.5 parts, calcium phosphate 0.4 parts, sodium selenite 0.01 parts, saccharin sodium 0.1-0.2 parts, ethoxyquinoline 0.5 parts, sodium bicarbonate 1-1.5 parts, pentose 0.2 parts, sodium chloride 0.18 parts, stone powder 1-1.5 parts.

[0010] The present invention also provides a sodium propionate-added feed capable of improving the slaughtering performance of sheep and the umami taste of mutton. The sodium propionate-added feed is prepared by adding 20 g / sheep / day of sodium propionate to the basic daily diet of the sheep.

[0011] The sodium propionate-added feed provided by the present invention can be applied in the field of sheep breeding, including improving sheep slaughtering performance and mutton flavor, and has potential application value.

[0012] The present invention has the following advantages: The invention provides sodium propionate as an additive in feed. The prepared optimized feed has a series of positive effects on sheep, changes the proportion of fiber decomposing bacteria in the gastrointestinal tract of sheep, and significantly improves the slaughter rate.

[0013] The present invention adds 20g / head / day of sodium propionate to the basic feed, and the optimized feed prepared in the sheep breeding has the advantages of short feeding time, easy operation, and disease-free sheep. It provides an effective method for the agricultural field to improve the livestock and poultry breeding business and further optimize product quality and economic benefits. It also provides potential evidence for enhancing the umami taste of mutton to increase the content of umami substances and glutamate levels.

[0014] In addition, sodium propionate supplementation can increase the level of muscle metabolite L-carnitine, identify 5 differential proteins associated with protein degradation pathways, and improve the sensory and nutritional quality of sheep by increasing the concentration of flavor amino acids. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 The PCoA scatter plot cluster analysis results of the intestinal flora of sheep after feeding the optimized feed with sodium propionate added; the horizontal and vertical coordinates represent the two selected principal coordinate components, and the percentage represents the contribution of the principal coordinate components to the sample composition differences.

[0016] Figure 2 Distribution of different bacterial genera in the intestine of sheep fed an optimized diet supplemented with sodium propionate.

[0017] Figure 3 Effect of feeding an optimized diet supplemented with sodium propionate on the meat flavor of sheep.

[0018] Figure 4 These are the OPLS-DA scoring results of metabolites in the two groups of samples before and after feeding the optimized feed with sodium propionate.

[0019] Figure 5 Results of differential metabolite determination in muscle of two groups of sheep before and after feeding an optimized diet supplemented with sodium propionate.

[0020] Figure 6 Results of the effects of feeding an optimized diet supplemented with sodium propionate on differential protein in muscle samples of sheep. DETAILED DESCRIPTION

[0021] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0022] Note: The experimental methods in the following examples are conventional methods unless otherwise specified, and are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials, reagents, etc. used in the following examples are all commercially available unless otherwise specified.

[0023] Experimental Example 1 Preparation and feeding scheme of sodium propionate-added feed 14 healthy Dorper × Suffolk crossbred sheep were randomly selected as test subjects. The sheep had similar body weight (26.51 + 0.49 kg) and were randomly divided into a control group (Group C) and a sodium propionate group (Group P), with 7 sheep in each group, half male and half female. To ensure the uncertainty and universality of sheep, it was used to observe the regulatory effect and results of sodium propionate addition on mutton quality.

[0024] Among them, the sheep in Group C received a basic diet, which was calculated according to the weight of the growth and fattening period of the mutton sheep, including 44 parts of corn, 5.24 parts of soybean meal, 10.93 parts of wheat bran, 34.96 parts of compound feed, 2.97 parts of premix, 1.05 parts of sodium carbonate, and 1.14 parts of sodium chloride. The compound feed was calculated by weight: it contained 54-55 parts of flaked corn, 3 parts of grass meal, 7-9 parts of soybean meal, 8 parts of cottonseed meal, 20 parts of stone powder, 5.4-6 parts of calcium hydrogen phosphate, 0.25 parts of vitamin A, 0.15 parts of vitamin D3, and 0.3 parts of vitamin E; the premix included 58 parts of corn, 7 parts of soybean meal, 10 parts of cottonseed meal, 8 parts of rapeseed meal, 12 parts of DOGS, 0.15 parts of vitamin A acetate, 0.03 parts of vitamin D3, and 0.3 parts of vitamin E. 0.8 parts, EDL-α-tocopherol acetate 0.16 parts, copper sulfate 0.15 parts, magnesium sulfate 0.6 parts, zinc sulfate 1.2 parts, manganese sulfate 0.5 parts, calcium phosphate 0.4 parts, sodium selenite 0.01 parts, saccharin sodium 0.1-0.2 parts, ethoxyquinoline 0.5 parts, sodium bicarbonate 1-1.5 parts, pentose 0.2 parts, sodium chloride 0.18 parts, stone powder 1-1.5 parts.

[0025] In addition to the basic diet, the sheep in group P were also supplemented with 20g / head / day of sodium propionate powder directly mixed into the basic diet. The pre-experimental phase lasted for 7 days. After the test subjects showed adverse reactions during the pre-trial phase, the formal test was conducted. The feeding test lasted for 4 months before slaughter. During the entire trial period, all sheep had free access to water and food, and the feed trough was kept clean every day, and the sheep were supervised to complete their feeding.

[0026] Experimental Example 2: Effect of Sodium Propionate Added Feed on Sheep The sample collection method in this experimental example includes the following: (1) Muscle sample collection: The longissimus dorsi muscle was extracted from the left carcass of each sheep and divided into two parts within 45 min.

[0027] The first part was immediately frozen in liquid nitrogen and stored at −80 °C for subsequent metabolism and analysis.

[0028] The second part was stored at 4°C for analysis of lamb flavor and amino acids.

[0029] (3) Gastrointestinal sample collection: The rumen and colon contents of each sheep were collected and placed into enzyme-free 15 ml centrifuge tubes.

[0030] Store the tubes in a -80°C freezer to facilitate determination of gastrointestinal flora and internal metabolites.

[0031] (4) Sample naming method: The rumen, colon contents, and longissimus dorsi muscle of group C sheep were designated CR, CF, and CL, respectively; The rumen, colon contents and longissimus dorsi muscle of the sheep in group P were designated as PR, PF and PL, respectively.

[0032] 1. Determination of sheep and mutton quality and slaughter performance Live weight refers to the weight of the animal before slaughter after fasting for 24 hours. Carcass weight is defined as the weight after removing the skin, head, hoof, tail, viscera and all fat in the body cavity. Slaughter rate = carcass weight / live weight × 100%. Carcass depth is the length of the inner edge of the connection between the midpoint of the anterior edge of the pubic symphysis and the midpoint of the anterior edge of the first cervical vertebra. Carcass height is determined by the distance from the end of the ribs perpendicular to the spine. Back fat thickness is measured from the 12th to the 13th ribs, using a vernier caliper.

[0033] Determination of muscle pH: The pH meter was calibrated using a pH standard buffer (e.g., phosphate buffer). The carcass PH-STAR instrument was inserted directly into the longissimus dorsi muscle after slaughter, and the pH values ​​were recorded 45 min and 24 h after slaughter, recorded as pH45min and pH24h, respectively. Seven measurements were made on different tissues for each muscle sample, and the average value was taken. Determination of muscle color: The CR-410 colorimeter was used, and the measurement was started after calibration on a standard white tile at a 2° observation angle. One hour after the longissimus dorsi muscle of sheep was cut, the sheep muscle color was measured using a standard D65 light source. Each sample was measured three times on different tissues, and the final average value was obtained. Determination of cooking loss and shear force of mutton (Su, L., et al. Effects of exercise on muscle fiber conversion, muscle development and meat quality of Sunit sheep. Meat science). All meat samples were heated in a water bath at 80°C, and kept heated until the core temperature of the meat reached 70°C, then the meat was cooled at room temperature and measured. The weight before heating (W1) and after heating (W2) was recorded, and the cooking loss rate was calculated using the formula ((W1−W2) / W1 × 100%). The cooked mutton was cut into (4cm×1cm×1cm) cubes, and then the shear force value of the muscle sample was measured using a 500N CLM3B tenderizer. Each sample was repeated seven times and the average value was taken.

[0034] The mutton samples were taken by direct drying method, combustion method, Soxhlet extraction method and Kjeldahl nitrogen determination method to determine the contents of moisture, ash, fat and protein in the longissimus dorsi muscle of sheep.

[0035] Generally, meat quality includes sensory quality and nutritional components, and pH reflects the degree of muscle glycogen degradation in sheep after slaughter (Zhang R, et al. Effects of organic trace minerals chelated with oligosaccharides on growth performance, blood parameters, slaughterperformance and meat quality in sheep. Front Vet Sci). The effects of sodium propionate on sheep slaughter performance and meat quality obtained by the above statistical and measurement methods are shown in Table 1 below. It can be seen that the pH45min and pH24h of the CL group and the PL group were both within the normal range. Cooking loss is closely related to the water retention of meat. Studies have shown that the smaller the cooking loss, the better the water retention of meat (Liang, Yanping, et al. "Effect of dietary Agriophyllumsquarrosum on average daily gain, meat quality and muscle fatty acids ingrowing Tan lambs. " Meat science 201(2023):, 109195). Color directly reflects the state of meat. The yellowness value (b*) of the PL group was significantly higher than that of the CL group, while there was no significant difference in redness value (a*) and brightness value (L*) between the two groups (P>0.05). Studies have found that the degree of protein cross-linking and aggregation after oxidation may affect the light reflection properties of meat, especially the yellowness value (Utrera M, Morcuende D, Estévez, Mario. Fat content has a significant impact on protein oxidation occurred during frozen storage of beef patties[J]. LWT - Food Science and Technology).Unsaturated fatty acids in fat are easily oxidized and cause pigment deposition in adipocytes to form melanin granules (Hoa VB, Song DH, Seol KH, et al. Effect of coating with combined chitosan and gallic acid on shelf-life stability of Jeju black cattle beef[J]. Animal Bioscience, 2024, 37(1)). Shear force is negatively correlated with meat tenderness, but there is no significant difference in shear force between the two groups (P>0.05). Slaughter performance reflects the growth and development status of animals, including carcass weight, slaughter rate, net meat weight, eye area, and backfat thickness. The results of this patent study showed that except for the slaughter rate of sheep (P<0.05), there was no significant difference in other indicators between the two groups (P>0.05). Therefore, it can be seen that the slaughter rate of sheep can be significantly improved by supplementing sodium propionate in the feed.

[0036] Table 1 Effects of sodium propionate on slaughter performance and meat quality of sheep

[0037] By measuring the moisture content in sheep muscle samples, its basic nutritional quality can be directly reflected. The results of the determination of the basic nutritional indicators of the longissimus dorsi muscle of sheep are shown in Table 2. The results show that the moisture content of the longissimus dorsi muscle samples of the two groups of sheep is between 75%. Higher ash and protein content means that the supplementation of sodium propionate makes the minerals, trace elements and nutrients in the sheep samples richer. The results of the study observed that the sheep samples in the PL group had higher ash and protein content than those in the CL group. In addition, in terms of taste and tenderness, it is proved that propionate can increase the intramuscular fat deposition of sheep, which provides a reasonable explanation for improving the tenderness of mutton.

[0038] Table 2 Determination of basic nutritional indexes of sheep longissimus dorsi muscle

[0039] 2. Effects of sodium propionate addition on sheep gastrointestinal flora and its metabolites 1. Determination of ovine gastrointestinal flora: Total microbial genomic DNA was extracted from rumen fluid and fecal samples using a kit for DNA extraction and PCR amplification according to the manufacturer's instructions. DNA quality and concentration were assessed using 1.0% agarose gel electrophoresis and a NanoDrop2000 spectrophotometer (Thermo Scientific, USA). To amplify the hypervariable region V3-V4 of the bacterial 16SrRNA gene, universal primer pairs 338F and 806R were used, and an ABI GeneAmp® 9700 PCR instrument (BIO-RAD, USA) was used. The PCR reaction mixture included 4 μL 5× Fast Pfu buffer, 2 μL 2.5 mM dNTPs, 0.8 μL of each primer (5 μM), 0.4 μL Fast Pfu polymerase, 10 ng template DNA, and ddH2O to a final volume of 20 μL. The PCR amplification cycle conditions were as follows: an initial denaturation step at 95 °C for 3 min, followed by 30 s denaturation at the same temperature in each cycle, 30 s annealing at 55 °C, and an extension of about 45 s at a temperature equal to or higher than that used for denaturation (72 °C), and a single extension at the same higher temperature (72 °C) for ten min before the process was terminated by a decrease to about 10 °C. To obtain the PCR products from the 2% agarose gel electrophoresis analysis and to ensure their purity according to the instructions provided by the kit manufacturer, the quantification was performed using the Qubit™ Fluorometer Model 4.0. The microbiome analysis method based on paired-end sequencing on the Illumina PE300 platform followed the standard protocol provided by Majorbio Bio-PharmTechnology Co. Ltd. (Shanghai, China). The raw sequencing reads have been deposited in the NCBI Sequence Read Archive (SRA) database and obtained the accession numbers of rumen microbiota and colon microbiota: PRJNA1110089 and PRJNA1110070. The obtained sequences were quality filtered by fastp (v0.19.6) and merged using FLASH (v1.2.7). The high-quality sequences were then denoised using the DADA2 plugin with recommended parameters in the Qiime2 pipeline. The plugin achieves single-nucleotide resolution based on the intra-sample error profile. The sequences denoised by DADA2 are called amplicon sequence variants (ASVs). The ASVs were classified and assigned by using the Blast classifier implemented in Qiime2 and the SILVA16S rRNA database (v138). Based on the ASV representative sequences, PICRUSt2 was used to predict the microbial functional potential, and the ASV representative sequences were aligned with the reference sequences using HMMER.

[0040] (1) PCoA principal component analysis: Based on the distance matrix, the binary Curtis distance algorithm was used for PCoA clustering analysis, and the ANOSIM inter-group difference test was used at the ASV level. The scatter plot clustering results of the principal coordinate analysis (PCoA) of the gastrointestinal flora of sheep after the two treatments are as follows Figure 1 shown. The horizontal and vertical coordinates therein represent the two selected principal coordinate components, and the percentage represents the contribution degree of the principal coordinate components to the sample composition difference. Figure A therein is the PCoA graph of the rumen flora; Figure B is the PCoA graph of the intestinal flora. PR is the rumen content of sheep in the sodium propionate group; CR is the rumen content of sheep in the control group; PF is the intestinal feces of sheep in the sodium propionate group; CF is the intestinal feces of sheep in the control group. The distribution of different bacterial genera in the intestinal samples of sheep in the two treatments is shown in Figure 2 shown, where Figure 2 in A and B, different colors represent different groups (or samples), the overlapping part represents the species shared by multiple groups (or samples), and the non-overlapping part represents the species unique to this group (or sample), and the numbers represent the quantity of the corresponding species. Figure 2 In C and D, the abscissa represents the species names at different taxonomic levels, the ordinate represents the percentage value of the abundance of a specific species in this sample, and different colors represent different groups; the P values therein are *0.01 < P ≤ 0.05, **0.001 < P ≤ 0.01, ***P ≤ 0.001; PR therein is the rumen content of sheep in the sodium propionate group; CR is the rumen content of sheep in the control group; PF is the intestinal feces of sheep in the sodium propionate group; CF is the intestinal feces of sheep in the control group. The results show that there are significant differences between the two groups, and they show obvious clustering (R = 0.1973, P = 0.0060). The principal components of PC1 and PC2 account for 21.69% and 17.53% of the total variance respectively, indicating that they are representative in the rumen of ruminants (see Figure 1 A therein). These results indicate that sodium propionate feeding has a regulatory effect on the rumen microbiota of sheep at the ASV level. There are no significant differences in the PCoA of the intestinal flora (see Figure 1 B therein). The VENN diagram shows that there are 142 bacterial genera in common between CR and PR, including 38 specific genera unique to PR, and 45 specific genera are parasitized in CR (see Figure 2 A therein). In the two groups of PF and CF, 216 bacterial genera were identified at the intestinal flora level, including 103 specific genera of PF and 32 specific genera of CF (see Figure 2Other studies have also reported that the rumen microbiota of ruminants has a higher diversity than the lower intestinal microbiota (Huang S, Ji S, Wang F, et al. Correction to: Dynamic changes of the fecal bacterial community in dairycows during early lactation[J]. AMB Express, 2021, 11(1)), which may lead to the observed clustering between rumen microbiota and intestinal microbiota in ruminants.

[0041] (2) Analysis of the composition and structure of the gastrointestinal flora of sheep: At the genus level, Succinivibrionaceae_ UCG-001 and Streptococcus ( Streptococcus ) was significantly higher than that in the CR group (see Figure 2C in the table). Succinivibrionaceae_UCG-001 accounted for more than 10% in both groups. Microorganisms of this genus play a key role in feed digestion, producing succinate (a precursor of propionate) rather than methane by competing with methanogens for hydrogen (Tong J, Zhang H,Yang D, Zhang Y, Xiong B, Jiang L. (2018) Illumina sequencing analysis of theruminal microbiota in high-yield and low-yield lactating dairy cows. PlosOne). Studies have shown that it is associated with reduced methane production in ruminants (McCabe MS, et al. (2015)MiSeq phylogenetic amplicon sequencing shows a large reduction of anuncharacterised Succinivibrionaceae and an increase of the Methanobrevibactergottschalkii clade in feed restricted cattle. Plos One). Streptococci were identified as one of the main bacterial groups present in corn starch, and these rumen microorganisms interact to promote metabolism in the body and promote protein and carbohydrate decomposition (Tett A, Pasolli E, Masetti G, Ercolini D, Segata N. (2021) Prevotella diversity, niches and interactions with the human host. Nat Rev Microbiol). The contents of acetate, propionate and butyrate in the PR group were significantly higher than those in the CR group (P<0.05) (see Table 3). KEGG analysis results showed that rumen microorganisms mainly showed functional enrichment in metabolism, especially amino acid and carbohydrate metabolism. The second was lipid and energy metabolism (see Table 3). In addition to the AMPK signaling pathway and HIF-1 signaling pathway, which were significantly increased in the CR group compared with the PR group, other pathways in signal transduction such as FoxO signaling pathway, MAPK signaling pathway and phosphoinositide signaling system were more obvious in the PR group. KEGG level 3 mainly includes major amino acid metabolic pathways such as arginine and proline metabolism, cysteine ​​and methionine metabolism, and lysine biosynthesis.In summary, it is necessary to further explore the effect of propionate on rumen microbial changes, which will help to better understand its effects on sheep metabolism, slaughter performance and meat quality.

[0042] Among the different intestinal bacteria, the bacteria with significantly higher relative abundance in the PF group than in the CF group included norank_f_ Ruminococcaceae, Acetitomaculum, Aeriscardovia, Colidextribacter, Syntrophococcus, Mogibacterium, Streptococcus, Coriobacteriaceae_UCG-003 and DNF00809 (P<0.05) (see Figure 2 D in ). On the contrary, Eubacterium_siraeum_group, norank_f_ Flavobacteriaceae, Odoribacter, Anaerovibrio and Paeniclostridium The relative abundance of Ruminobacteriaceae was lower than that of the CF group (P<0.05). In addition, the PF group had a higher content of acetic acid in the intestine (P<0.05) (see Table 3). The addition of sodium propionate caused the Ruminobacteriaceae to increase in the gastrointestinal tract of sheep and significantly produced acetic acid (Kiarie, EG, Mohammadigheisar, M., Kakhki, RAM, & Madsen, MH (2021). Impact of feeding modified soyprotein concentrate in the starter phase on growth performance and gastrointestinal responses in broiler chickens through to day 42 of age. Poultry science). Previous results showed two different types of bovine enterotypes: type I and type II. Prevotella Type II is dominated by Ruminococcaceae The presence of key microbial species in different gut types may be associated with specific phenotypic characteristics (Knights, D., Ward, TL, McKinlay, CE, Miller, H., Gonzalez, A., McDonald, D., & Knight, R. (2014). Rethinking "enterotypes". Cell Host Microbe). PrevotellaThere is a positive correlation in amino acid metabolism, especially branched-chain amino acids (BCAA) (Pedersen, HK, Gudmundsdottir, V., Nielsen, HB, Hyotylainen, T., Nielsen, T., Jensen, BA,..., Pedersen, O. (2016). Human gut microbes impact host serum metabolome and insulin sensitivity. Nature). In the rumen of beef cattle fed a high-fiber diet, Acetitomaculum was identified as a significant contributor, resulting in increased production of acetic and butyric acids. Aeriscardovia The treatment increased the average daily weight gain, significantly reduced feed conversion rate, and improved the intestinal health and growth performance of chickens (Farooq MZ, Wang X, Yan X. (2023) Effects of Aeriscardovia aeriphila on growth performance, antioxidant functions, immune responses, and gut microbiota in broiler chickens. JZhejiang Univ Sci B). Species belonging to the phylum Firmicutes Colidextribacter It may play a role in regulating systemic inflammatory responses and maintaining intestinal mucosal integrity (Liu T, et al. (2022) Impact of FeedingProbiotics on Blood Parameters, Tail Fat Metabolites, and Volatile FlavorComponents of Sunit Sheep. Foods). Syntrophococcus There was a significant positive correlation with the production of acetic acid, propionic acid, and butyric acid (Hao Y, et al. (2021) Rumen Fermentation, Digestive Enzyme Activity, and Bacteria Composition between Pre-Weaning and Post-Weaning DairyCalves. Animals (Basel)). The acetic acid content in the PF group was significantly higher than that in the CF group (P<0.05). In the KEGG functional analysis of colonic microorganisms, the PF group showed significantly higher functional activity compared with the CF group.

[0043] 2. Determination of ovine gastrointestinal metabolites: 0.6 g of rumen and colon contents were placed in a 15 mL centrifuge tube, 2 mL of iced saline and 1 mL of 50% sulfuric acid solution were added, vortexed and centrifuged at 10,000 g for 5 min at 4 °C. Then 2 mL of ether was added for thorough mixing, and centrifuged at 10,000 g for 5 min at 4 °C. After chromatography, the supernatant was taken. The supernatant was filtered through a 0.22 μm organic filter membrane into an injection bottle for gas chromatography analysis.

[0044] Gas chromatography conditions: DB-5 column (30 m×0.25 mm, 0.25 μm), helium carrier gas, flow rate of 1.2 μL / min, injection port temperature of 250 ℃, injection volume of 1 μL. The temperature program is as follows: the initial temperature is set to 100 ℃ and maintained for 0.5 min; it is increased to 180 ℃ at a rate of 8 ℃ / min and maintained for 1 min; it is increased to 200 ℃ at a rate of 20 ℃ / min and maintained for 5 min. Mass spectrometry conditions: ion source temperature of 250 ℃, transfer line temperature of 250 ℃, and mass spectrometry scanning range of 40-450 m / z. The metabolites of sheep rumen and colon microorganisms are shown in Table 3.

[0045] Table 3 Metabolites of sheep rumen and colon microorganisms

[0046] The results of this patent experiment showed that the contents of acetic acid, propionic acid and butyric acid in the PR and PF groups were higher than those in the CR and CF groups, indicating that the addition of sodium propionate can improve the flora environment of the gastrointestinal tract of sheep. Succinivibrionaceae _ UCG-001 The abundance of acetic acid in the PR group was significantly higher than that in the CR group, which was also related to the increase in propionic acid content. In addition, the content of isovaleric acid was higher in the PR group than in the CR group. Branched short-chain fatty acids such as isovaleric acid and isocaproic acid are mainly synthesized through the amino acid metabolic pathway using amino acids as substrates, and the degradation of amino acids also affects the production of acetic acid and propionic acid. In short, the supplementation of propionate can enhance the growth and proliferation ability of carbohydrate-degrading microorganisms and regulate the composition and structure of short-chain fatty acids.

[0047] 3. Effect of adding sodium propionate on the flavor of mutton In this experiment, the flavor of mutton was determined by subjecting it to odorization using a PEN3 electronic nose system (AIRSENSE, Germany). This system consists of multiple metal oxide gas sensors, each with different characteristics. First, sheep muscle samples (5 g) were crushed and placed into 15 mL electronic nose sampling bottles. Then, they were heated to 60 °C at room temperature and maintained for 40 min to reach an equilibrium state. To ensure accuracy and reliability, the carrier gas inlet flow rate was set at 200 mL / min, and the self-cleaning time was set at 90 s to remove residues from the previous sample. Finally, a detection time of 120 s was carried out to obtain the odor information released by the mutton samples. Multiple metal oxide sensors were used in this method to identify different types of volatile compounds. For example, the W1C sensor is mainly used to detect umami substances, the W5S sensor is used to detect nitrogen oxides, the W3C sensor is used to detect aromatic components and ammonia, and so on. By using a combination of sensitivities to these specific volatile compounds, a more comprehensive and accurate description of the various volatile components released by the mutton samples can be obtained.

[0048] The measured effects of feeding sheep with feed supplemented with sodium propionate on sheep flavor are shown in Figure 3 as follows. Among them, A is a radar chart of the flavor of muscle samples of two different groups of sheep identified by an electronic nose; B is a comparative analysis of the differences in the sensitivities of each sensor in each group. The differences are statistically significant: * represents 0.01 < P ≤ 0.05, ** represents 0.001 < P ≤ 0.01, *** represents P ≤ 0.001; C is the result of principal component analysis (PCA), which is used to evaluate and compare the flavor characteristics of the two groups of muscle samples. From Figure 3 A in it, it can be seen that after adding sodium propionate, the volatile organic compounds changed. From Figure 3 the PCA analysis result shown in C in it, it can be seen that P = 0.024, and the cumulative contribution of PCA1 (92.23%) and PCA2 (6.35%) accounted for 98.58%, indicating that they can cover most of the odor changes. It should be noted that under the influence of sodium propionate, as shown in Figure 3 B in it, sensors WIC, W3C, W5C, and W2W showed higher sensitivities to volatile flavor compounds, directly affecting the mutton flavor and resulting in significant differences in taste composition (P < 0.05). It can be concluded from this that supplementing sodium propionate in the basal diet significantly increased the levels of umami substances, ammonia levels, and olefin content, and at the same time decreased certain organosulfides.

[0049] IV. Effects of Adding Sodium Propionate on Amino Acids in Mutton 0.5 g of frozen meat sample was added with 2.5 mL of 8% sulfosalicylic acid and ground in a freezer grinder at 4 °C for 180 s. Subsequently, the mixture was centrifuged at 4 °C, 10,000 g / min for 15 min. The supernatant (1 mL) was extracted, hexane (0.25 mL) was added and shaken, and the non-organic phase was filtered with a 0.22 μm aqueous filter membrane after standing and stratification, and analyzed using an amino acid autoanalyzer. LA8080 amino acid analyzer: analytical column size (4.6 mm×60 mm, 3 μm). The injection volume was 20 μL, operated at a pump flow rate of 0.1 mL / min, a cycle time of 20 min, a run time of 1.4 min, a column temperature of 57 °C and a reaction column temperature of 135 °C. Electronic nose and amino acids were evaluated by one-way ANOVA, in which the feeding group (P vs C) was considered as a fixed factor. Homogeneity of variance was assessed by Levene's test, and normality was tested by the Shapiro-Wilks test. Data are presented as mean ± standard error, and significance was defined as P < 0.05.

[0050] After determination, 15 common amino acids were detected in the two groups of sheep samples, as shown in Table 4. They include essential amino acids (Thr, Val, Met, Ile, Leu, Phe, Lys) and non-essential amino acids (Asp, Ser, Glu, Gly, Ala, Tyr, His, Arg). The Thr content in group C was significantly higher than that in group L, while the Glu content was significantly lower than that in group L (P<0.05). Free amino acids are considered to have better meat flavor (You, Z. et al. A review of taste-active compounds in meat: Identification, influencing factors, and taste transduction mechanism. Journal of food science). Among the free amino acids in mutton muscle, phenylalanine and arginine are the main bitter ones, serine, glycine, threonine, etc. are sweet, and aspartic acid, glutamic acid, asparagine and glutamine are the ones that bring umami taste (Gilbreath, K. et al. Amino Acid Nutrition and Reproductive Performance in Ruminants. Advances in experimental medicine and biology). Therefore, it is important to increase the level of free amino acids in mutton. Results The results of metabolomics and proteomics focused on the key metabolic pathways in muscle, which are protein synthesis and degradation. Previous studies have found that the addition of probiotics to the diet can change the amino acid profile of the longissimus dorsi muscle of fattening pigs, thereby improving meat quality (Weng, G. et al.. Effects of dietary Brevibacillus laterosporus BL1 supplementation on meat quality, antioxidant capacity, and the profiles of muscle amino acids and fatty acids in finishing pigs. Meatscience).In addition, the addition of Clostridium butyricum to the diet regulates the microbial composition of the intestinal group and significantly increases the content of flavor amino acids in fattening goats (Zhang, M. et al. Effects of dietary Clostridium butyricum and rumenprotected fat on meat quality, oxidative stability, and chemical composition of finishing goats. Journal of animal science and biotechnology). It is currently known that glutamate is one of the free amino acids that induce umami taste. The taste synergy between it and inosinic acid is a sign of umami taste, and the intensity of umami taste will be significantly enhanced (Rifkin, B., & Bartoshuk, LM (1980). Taste synergism between monosodium glutamate and disodium 5'-guanylate. Physiology & behavior). From this study, it can be seen that the supplementation of sodium propionate in the basic diet increases the level of glutamate in the longissimus dorsi muscle, increases the production of umami substances in mutton, and improves the umami taste of mutton.

[0051] Table 4 Effects of sodium propionate on amino acids in sheep muscle

[0052] 5. Effect of sodium propionate addition on protein metabolism-related genes in mutton muscle Real-time PCR was used to quantitatively analyze gene expression in sheep muscle. First, total RNA was extracted from sheep muscle tissue using TaKaRa RNAisoPlus reagent, and the concentration and quality of the sample were determined by a micro-nucleic acid protein analyzer. The sample was then diluted to 500 ng / μL, and the quality of RNA was evaluated by agarose gel electrophoresis. After confirming the purity of RNA, all samples were reverse transcribed using TaKaRa PrimeScriptTMRT kit and gDNA Eraser, and the final concentration of cDNA obtained was 50 ng / μL. Specific fluorescent quantitative primers for sheep PI3K, AKT, mTOR, TSC2, Rheb, S6K, eIF4B, TelO2, LC3, P62 and β-actin genes were designed using conventional methods, and the diluted cDNA samples were used together with specific fluorescent quantitative primers to prepare real-time quantitative PCR reaction systems. Amplification efficiency curves and standard curves were generated according to the Ct value to evaluate the primer effect. The DNA fragment size of the amplified product was determined by electrophoresis analysis. Real-time quantitative PCR amplification was performed under the following conditions: pre-denaturation: 95 °C, 30 s; denaturation: 95 °C, 5 s; annealing: 60 °C, 30 s (temperature determined by primer design), and final extension at 72 °C, 30 s. Relative gene expression was calculated by normalizing sheep muscle genes to β-actin, which was used as an internal reference gene. Then 2 -ΔΔCt Methods The data were analyzed.

[0053] The analysis of metabolites and differentially expressed proteins in sheep muscle revealed the main metabolic pathways for protein metabolism, including protein synthesis and degradation. The effects of sodium propionate on the expression of genes related to the longest muscle of sheep back are shown in Table 5. It can be seen that the levels of AKT, 4EBP and TELO2 genes in group P were significantly reduced compared with group C (P<0.05). It is worth noting that the expression level of LC3 gene in group C was significantly reduced compared with group P (P<0.05). Previous studies have demonstrated that autophagy acts as a cellular stress response in which microtubule-associated protein light chain 3 (LC3-I) is converted to microtubule-associated protein light chain 3-II (LC3-II). LC3 is also considered to be a reliable marker for autophagosome formation and autophagy initiation (Klionsky DJ, Abdalla FC, Abeliovich H, et al. Guidelines for the use and interpretation of assays for monitoring autophagy[J]. Autophagy). In mammals, amino acid metabolism and protein autophagy are complexly regulated, and lysine (Lys), leucine (Leu), arginine (Arg), and glutamine (Gln) have been extensively studied in relation to autophagy.

[0054] In this study, the lysine level in the longissimus dorsi muscle of the sodium propionate-fed group was lower compared to the control group. This suggests that insufficient lysine supply may activate the autophagy-lysosomal pathway, causing skeletal muscle cells to undergo autophagy and degrade their own proteins as a response to amino acid deficiency (Sato, T., Ito, Y., & Nagasawa, T. (2013). Regulation of skeletal muscle protein degradation and synthesis by oral administration of lysine in rats. Journal of nutritional science and vitaminology). mTOR kinase is a key regulator that plays an important role in various signaling pathways that induce autophagy and regulate cell apoptosis (Zi, D., Zhou, ZW, Yang, YJ, Huang, L., Zhou, ZL, He, SM, He, ZX,&Zhou, SF (2015). Danusertib Induces Apoptosis, Cell Cycle Arrest, and Autophagy but Inhibits Epithelial to Mesenchymal Transition Involving PI3K / Akt / mTOR Signaling Pathway in Human Ovarian Cancer Cells. International journal of molecular sciences). In previous studies, the effect of Lys treatment on the phosphorylation levels of downstream targets S6K1 and 4EBP1 in muscle was examined, which was mediated by mTORC1 (Sato, T., Ito, Y., & Nagasawa, T. (2014). Lysine suppresses myofibrillar protein degradation by regulating the autophagic-lysosomal system through phosphorylation of Aktin C2C12 cells. SpringerPlus). After treatment in a medium containing 10 mmol / L lysine for 30 min, the ratio of LC3-II to LC3-I was significantly reduced by 30%, while the phosphorylation levels of S6K1 and 4EBP1 were significantly increased.These findings suggest that lysine regulates autophagy-lysosome activity in C2C12 myotubes through an mTORC1-dependent pathway (Lang, SM, Kazi, AA, Hong-Brown, L., & Lang, CH (2012). Delayed recovery of skeletal muscle mass following hindlimb immobilization in mTOR heterozygous mice. PloSone). Lysine supplementation can enhance the phosphorylation level of 4EBP1, indicating that lysine addition can inhibit the activity of the autophagy-lysosome system and promote protein synthesis in muscle cells by activating the mTORC1 signaling pathway. In addition, LC3-II also showed a significant decrease under this treatment condition; however, the mechanism of lysine-mediated inhibition of muscle protein degradation is still unclear. As a special autophagy receptor, SQSTM1 / p62 interacts with the Atg8 family (LC3 / GABARAP) and ubiquitin chains in the process of aggregation-induced autophagy, mitochondrial autophagy and xenophagy (Dikic, I., & Elazar, Z. (2018). Mechanism and medical implications of mammalian autophagy. Nature reviews. Molecular cellbiology), playing a key role. Therefore, supplementing sodium propionate in sheep muscle can increase the level of protein autophagy, affect the free amino acid composition in sheep muscle, and increase the content of umami substances in lamb meat.

[0055] Table 5 Effects of sodium propionate on gene expression in sheep muscle

[0056] 6. Effects of sodium propionate supplementation on sheep muscle metabolites Each sample weighed 50 mg, added 300 μL of 80% methanol, and mixed and crushed evenly in a tissue disruption device. 700 μL of the 80% methanol mixture was treated with ice bath ultrasound for 20 min. The resulting solution was then placed at -20 °C for 2 h, followed by centrifugation at 16,000 g and 4 °C for 20 min. Finally, the supernatant was collected. The supernatant was shaken dry by a high-speed vacuum concentrator centrifuge. For mass spectrometry analysis, 100 μL of 50% methanol was added to re-dissolve the sample, and the supernatant was injected for UHPLC-QTOF-MS analysis by centrifugation at 20,000 g for 15 min at 4 °C. Sample analysis was performed using a SHIMADZU-LC30 ultra-high performance liquid chromatography system (UHPLC) with an ACQUITY UPLC® HSS T3 column (2.1×100 mm, 1.8 µm). 4 μL of sample was injected into the chromatographic system, and the column temperature was set to 40 °C and the flow rate was maintained at 0.3 mL / min. The mobile phase consisted of two components: A was 0.1% formic acid solution (supplied by Merck, Darmstadt, Germany) and B was acetonitrile (also from Merck). To ensure the reliability and stability of the experimental data, quality control (QC) samples were inserted into the sample queue for monitoring and evaluation. Each individual sample was analyzed by electrospray ionization (ESI) in both positive and negative ion modes. The samples were effectively separated by UPLC and analyzed by mass spectrometry using a QE Plus mass spectrometer (Thermo Scientific).

[0057] The mass spectrometry acquisition time was set to 15 min, and the parent ion scan range was limited to 70-1050 m / z. Secondary mass spectrometry analysis was performed as follows: full scan was triggered to collect secondary mass spectra (MS2 scans) of the 10 most intense parent ions. Raw data were acquired using MSDIAL software, and peak alignment, retention time correction, and peak area extraction were performed. For metabolite structure identification, accurate mass matching (mass tolerance <10 ppm) and secondary spectrum matching (mass tolerance <0.01 Da) were used. To ensure the accuracy of the data, ion peaks with more than 50% missing values ​​within the group were excluded during the statistical analysis stage. In addition, the total peak area of ​​positive and negative ion data was normalized, and the positive and negative ion peaks were integrated and applied to Python software for pattern recognition. Finally, data analysis was performed after unit variance scaling (UV) preprocessing. The stability of the non-targeted metabolomics system was evaluated and analyzed by comparing the base peak spectra of QC samples, PCA statistical analysis of samples, correlation of QC samples, and Hotelling's T2 test of all samples.

[0058] Orthogonal partial least squares discriminant analysis (OPLS-DA) was used to analyze the differential metabolites between the two groups (OPLS-DA VIP>1, and P<0.05). The OPLS-DA model was verified, and the Q2 score of the original model was compared with the Q2 score obtained after 200 permutation tests. The quality of the sheep samples fed to the two groups was tested, and the OPLS-DA score results of the metabolites of the two groups of samples were obtained as follows Figure 4 As shown, A is the OPLS-DA score diagram of metabolites in the two sample groups. The OPLS-DA score and permutation test were performed on the overall sample using the negative ion detection mode. The horizontal axis represents the correlation between the Y of the random group and the Y of the original group, and the vertical axis represents the R2 and Q2 scores. The QC sample is represented by a triangle; (B) The principal component analysis diagram shows the obvious intra-group aggregation and inter-group separation in the muscle sample, and the ellipse represents the 95% confidence interval. The results show that the Q2 score after the permutation test is lower than the original model, indicating that the OPLS-DA model established based on the experimental data is not overfitted, thus ensuring the reliability of the model, see Figure 4 After removing the quality control sample, it can be observed that dietary sodium propionate supplementation leads to obvious clustering and separation between muscle groups, see Figure 4 B. Based on the variable importance projection (VIP) derived from the OPLS-DA model, the influence and interpretability of each metabolite expression pattern on sample classification and differentiation of different groups were further evaluated. The differential metabolites in the muscles of the two groups of sheep are shown in Figure 5 As shown, Figure 5 A is a bubble diagram of differential metabolite enrichment; B is the main metabolic pathways enriched by differential metabolites; C is the classification of differentially expressed proteins in muscle; and D is a volcano diagram of differential metabolites in muscle. Figure 5 Differential metabolites with significant biological significance can be determined. By selecting metabolites with multivariate statistical analysis VIP>1 and univariate statistical analysis P value <0.05 under simultaneous detection of positive and negative ions, 2614 muscle metabolites were identified in the database. These muscles identified 170 differential metabolites associated with various KEGG metabolic pathways. These included metabolic pathways, glycerophospholipid metabolism, and cysteine ​​and methionine metabolism that were enriched in specific pathways (see Figure 5 In addition, most of these diverse metabolites are involved in ovine protein metabolism (see Figure 5 Figure 3 (B), indicating that sodium propionate supplementation may induce metabolic changes in lamb muscle.

[0059] To ensure comprehensive and actionable data, all metabolites were further detected in positive and negative ion modes. Muscle metabolites mainly include lipid and lipid-like molecules (52, 31.52%), organic heterocyclic compounds (32, 19.39%), organic acid derivatives (26, 15.76%), benzo compounds (17.10, 3%), and polyketones (17, 10.3%) (see Figure 5 These results may have important implications for fatty acid metabolism and flavor in mutton. Compared with the C group, the P group showed 7 significantly down-regulated differential metabolites and 3 significantly up-regulated differential metabolites (see Figure 5 D in the article). We chose to focus on the possible benefits of acetyl-L-carnitine for meat quality. Acetyl-L-carnitine is an antioxidant present in sheep, and it was significantly upregulated in the P group. In addition, clinical studies have shown that acetyl-L-carnitine promotes mitochondrial metabolism, which can prevent oxidative stress and rapidly improve insulin resistance (Bigio B, Azam S, Mathé AA, Nasca C. (2024) The neuropsychopharmacology of acetyl-L-carnitine (LAC): basic, translational and therapeutic implications. Discov Ment Health). Among muscle metabolites, the differential metabolites of sheep muscle samples after feeding sodium propionate were mainly enriched in glycerophospholipid metabolism, cysteine ​​and methionine metabolism, which play a key role in protein synthesis and degradation.

[0060] VII. Effect of sodium propionate addition on differential protein in sheep muscle This experiment was tested through the following process, specifically protein extraction and purification → protein digestion → peptide desalting and quantitative analysis of peptides → DIA mass spectrometry detection.

[0061] This proteomics used the Majorbio platform (https: / / cloud.majorbio.com) to perform bioinformatics analysis on Astral proteomics (Ren et al., 2022). Spectronaut software was used to search DIA raw data, and the R package "t-test" was used to calculate the P value and fold change of the two groups of proteins. The fold change threshold (FC>1.2 or FC<0.83) and P value <0.05 were used to identify differentially expressed proteins (DEPs). Finally, GO (http: / / geneontology.org / ) and Kyoto Encyclopedia of Genes and Genomes (KEGG, http: / / www.genome.jp / kegg / / ) were used to functionally annotate all identified proteins, DEPs were used for GO and KEGG enrichment analysis, and String v11.5 software tool set was used for protein interaction analysis. After measurement, the effect of adding sodium propionate on the differential proteins in sheep muscle samples was obtained. Figure 6 As shown, A is a volcano plot of two groups of differential proteins (P value + VIP screening of differential metabolites), the upregulated "↑" is red, the downregulated "↓" is blue, and the numerical annotation shows the number of each category; B is a VENN diagram showing the characteristic proteins and shared proteins in the muscle tissues of the two groups of sheep; C is the metabolic pathway analysis of differential protein enrichment; D is the protein quantification of different functional pathways.

[0062] Based on the mass spectrometry results, the top 20 key differentially expressed proteins were identified by visualizing the volcano plot and performing differentially expressed protein (DEP) analysis using two criteria (see Figure 6A in the data set). Compared with the database, the difference fold FC>1.2, FC<0.86, and P value<0.05. Therefore, the main focus is on differentially expressed proteins related to protein degradation, such as A0A836A510 (methylsulfoxide reductase), which can reduce methylsulfoxide to methionine and whose expression is significantly downregulated in the P group (Chatterjee, A.,&Sepuri, NBV (2024). Methionine sulfoxide reductase2 regulates Cvt autophagic pathway by altering the stability of Atg19 andApe1 in Saccharomyces cerevisiae. The Journal of biological chemistry). According to the amino acid results, the methionine content in the BL group was lower than that in the C group; however, this difference did not reach statistical significance. Autophagy is involved in protein degradation (Akabane, T., et al. Heme deficiency in skeletal muscle exacerbates sarcopenia and impairs autophagy by reducing AMPK signaling. Scientific reports). A0A6P3EBK7 is a ubiquitin-binding protein that positively regulates ligand-activated EGFR internalization by specifically recognizing and binding to "Lys-63"-linked ubiquitin, and binds to the Ub group on the ubiquitinated EGFR on the cell membrane. EGFR dimerization leads to the formation of an intracellular tyrosine kinase domain, which promotes the conversion of ATP to ADP and phosphorylates TK (tyrosine kinase) to activate the catalytic activity of EGFR (Needham, SR, et al. EGFR ligomerization organizes kinase-active dimers into competent signalling platforms. Nature communications).Activated EGFR controls protein synthesis and degradation by regulating proliferation and anti-apoptotic signals through the PI3K-AKT-mTOR signaling pathway (Battaglioni, S., et al. mTORC1phosphorylates and stabilizes LST2 to negatively regulate EGFR. Proceedingsof the National Academy of Sciences of the United States of America). It was observed in the samples that after supplementation with sodium propionate, the expression of A0A6P3EBK7 protein in the P group was significantly decreased, suggesting that it may have a regulatory effect on the protein degradation rate. A0A836AKT6 and W5NRR3, which are related to energy metabolism and apoptosis, were significantly upregulated in the P group. The former corresponds to ATP synthase subunit α in mitochondria, while the latter promotes DNA damage-induced apoptosis (Roberts, JZ, Crawford, N., & Longley, DB (2022). The role of Ubiquitination in Apoptosis and Necroptosis. Cell death and differentiation). The serine / threonine protein phosphatase CPPED1 (A0A835ZPE9) was significantly downregulated in the P group and specifically targeted the AKT family kinase Ser-473 site for dephosphorylation, which blocked cell cycle progression and promoted apoptosis.

[0063] In the volcano plot, A0A836AKT6 and W5NRR3 showed more significant significance in the P group compared with A0A835ZPE9. Sodium propionate may act on ADP release and protein synthesis mediated by the mTOR signaling pathway by inhibiting EGFR activation, and promote protein degradation with the help of A0A836AKT6 and W5NRR3 proteins (Haapalainen, AM, Daddali, R., Hallman, M., & Rämet, M. (2021). Human CPPED1 belongs to calcineurin-likemetallophosphoesterase superfamily and dephosphorylates PI3K-AKT pathwaycomponent PAK4. Journal of cellular and molecular medicine). Ultimately, this leads to different amino acid profiles. The effect of sodium propionate on the expression level of sheep muscle protein was evaluated using DIA quantitative proteomics analysis. The results are shown in Figure 6 As shown in A, the results showed that a total of 214 differentially expressed proteins (DEPs) were identified in muscle, of which 83 were up-regulated and 131 were down-regulated. The main differentially expressed proteins were involved in the inner mitochondrial membrane protein complex, active transmembrane transporter activity, transmembrane transporter complex, and other signaling pathways closely related to material transport (see Figure 6 In the Venn diagram, 3206 common proteins were identified in both groups (accounting for 99.72% of the total), while 5 proteins were found exclusively in the P group (accounting for 0.16% of the total), and 4 proteins were found exclusively in the C group (accounting for 0.12% of the total), which indicates that the addition of sodium propionate can cause changes in differential expression in sheep muscle (see Figure 6 The differentially expressed proteins were analyzed for KEGG pathway enrichment, and the P value was less than 0.05 after correction. The results of KEGG enrichment analysis showed that more than 22 proteins were enriched in the functions of interest, especially in oxidative phosphorylation (see Figure 6D in). During active protein metabolism, ATP is hydrolyzed into ADP and Pi, and quickly enters the mitochondria through oxidative phosphorylation for efficient utilization (Kummer, E., & Ban, N. (2021). Mechanisms and regulation of protein synthesis inmitochondria. Nature reviews. Molecular cell biology). In addition, oxidative phosphorylation also plays an important regulatory role in protein degradation (Mandic, M., Paunovic, V., Vucicevic, L., Kosic, M., Mijatovic, S., Trajkovic, V., & Harhaji-Trajkovic, L. (2024). No energy, noautophagy-Mechanisms and therapeutic implications of autophagic responseenergy requirements. Journal of cellular physiology), and is related to the energy pathway required for protein autophagy. Differentially expressed proteins may affect the variability of mutton taste. Therefore, sodium propionate can regulate these differential proteins and these differential proteins can serve as potential markers for regulating mutton flavor, further regulating the flavor of mutton, which has application value in the field of sheep farming.

[0064] Although the content of the present invention has been described in detail through the above preferred embodiments, it should be appreciated that the above description should not be considered as a limitation of the present invention. After reading the above content, it will be apparent to those skilled in the art that various modifications and substitutions of the present invention will occur. Therefore, the protection scope of the present invention should be limited by the appended claims.

Claims

1. The application of sodium propionate in improving sheep slaughtering performance and mutton flavor, characterized in that: The slaughter rate and the content of umami substances in sheep can be improved by supplementing sodium propionate in the basal diet.

2. The use according to claim 1, characterized in that: The umami substances are free amino acids.

3. The use according to claim 2, characterized in that: The free amino acids include glutamic acid.

4. The use according to claim 1, characterized in that: The supplement amount of sodium propionate is 20g / each / day.

5. The use according to claim 1, characterized in that: The basic diet comprises, by weight, 44 parts of corn, 5.24 parts of soybean meal, 10.93 parts of wheat bran, 34.96 parts of compound feed, 2.97 parts of premix, 1.05 parts of sodium carbonate, and 1.14 parts of sodium chloride; The compound feed comprises, by weight, 54-55 parts of flaked corn, 3 parts of grass meal, 7-9 parts of soybean meal, 8 parts of cottonseed meal, 20 parts of stone powder, 5.4-6 parts of calcium hydrogen phosphate, 0.25 parts of vitamin A, 0.15 parts of vitamin D3, and 0.3 parts of vitamin E; The premix comprises, by weight, 58 parts of corn, 7 parts of soybean meal, 10 parts of cottonseed meal, 8 parts of rapeseed meal, 12 parts of DOGS, 0.15 parts of vitamin A acetate, 0.03 parts of vitamin D3, 0.8 parts of vitamin E, 0.16 parts of EDL-α-tocopherol acetate, 0.15 parts of copper sulfate, 0.6 parts of magnesium sulfate, 1.2 parts of zinc sulfate, 0.5 parts of manganese sulfate, 0.4 parts of calcium phosphate, 0.01 parts of sodium selenite, 0.1-0.2 parts of saccharin sodium, 0.5 parts of ethoxyquinoline, 1-1.5 parts of sodium bicarbonate, 0.2 parts of pentose, 0.18 parts of sodium chloride, and 1-1.5 parts of stone powder.

6. A sodium propionate-added feed for improving sheep slaughter performance and mutton flavor, characterized in that: The sodium propionate-added feed is a feed in which 20 g of sodium propionate is added to the sheep's basic diet per sheep per day. The basic diet comprises, by weight, 44 parts of corn, 5.24 parts of soybean meal, 10.93 parts of wheat bran, 34.96 parts of compound feed, 2.97 parts of premix, 1.05 parts of sodium carbonate, and 1.14 parts of sodium chloride; The compound feed comprises, by weight, 54-55 parts of flaked corn, 3 parts of grass meal, 7-9 parts of soybean meal, 8 parts of cottonseed meal, 20 parts of stone powder, 5.4-6 parts of calcium hydrogen phosphate, 0.25 parts of vitamin A, 0.15 parts of vitamin D3, and 0.3 parts of vitamin E; The premix comprises, by weight, 58 parts of corn, 7 parts of soybean meal, 10 parts of cottonseed meal, 8 parts of rapeseed meal, 12 parts of DOGS, 0.15 parts of vitamin A acetate, 0.03 parts of vitamin D3, 0.8 parts of vitamin E, 0.16 parts of EDL-α-tocopherol acetate, 0.15 parts of copper sulfate, 0.6 parts of magnesium sulfate, 1.2 parts of zinc sulfate, 0.5 parts of manganese sulfate, 0.4 parts of calcium phosphate, 0.01 parts of sodium selenite, 0.1-0.2 parts of saccharin sodium, 0.5 parts of ethoxyquinoline, 1-1.5 parts of sodium bicarbonate, 0.2 parts of pentose, 0.18 parts of sodium chloride, and 1-1.5 parts of stone powder.

7. Use of the sodium propionate added feed as claimed in claim 6 in the field of sheep breeding.

8. The use according to claim 7, characterized in that: The application includes improving sheep slaughtering performance and mutton flavor.

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