Nutrition additive for preventing and relieving livestock transportation stress and application

Abstract

The invention relates to the technical field of livestock transportation stress, and particularly discloses a nutrition additive for preventing and relieving livestock transportation stress and application of the nutrition additive. The feed additive comprises the following components in parts by weight: 1-10 parts of vitamin B1, 1-10 parts of vitamin B6, 1-18 parts of vitamin C, 1-3 parts of ATP, 1-6 parts of sodium bicarbonate, 1-3 parts of sodium chloride, 1-5 parts of L-lysine hydrochloride, 1-5 parts of inosine, 10-30 parts of glucose, 4-12 parts of astragalus polysaccharide, 4-12 parts of N-carbamylglutamic acid and 1-6 parts of a traditional Chinese medicine composition. The astragalus polysaccharide in the components can relieve the influence of transportation stress on the meat quality of livestock; moreover, the N-carbamylglutamic acid in the component can reduce the adverse effect of the transportation stress on the meat quality of the livestock, and by supplementing the N-carbamylglutamic acid, the muscle energy metabolism can be adjusted to weaken the influence of the transportation stress on the muscle metabolism of the livestock, so that the effect of preventing and relieving the transportation stress is achieved, and the meat quality of the livestock is ensured.

Description

Technical Field

[0001] The invention relates to the technical field of livestock transportation stress, and in particular to a nutritional additive for preventing and alleviating livestock transportation stress and an application thereof. Background Art

[0002] "Stress" refers to the biological response that occurs when livestock are affected by stressors and their internal balance is threatened. The physiological response of livestock to stressors includes three stages: immediate response stage, acute stress response stage and chronic stress response stage. The immediate response stage is an adrenaline-dependent mechanism, in which liver glycogen is decomposed to produce glucose (GLU) to provide immediate energy needs. This reaction affects metabolic rate, heart function, blood pressure, peripheral blood circulation, respiration, visual acuity and energy utilization, so that livestock can cope with physical and emotional stimulation. If the stressor does not disappear, the second stage reaction - acute stress response - occurs. During this period, the increase in the body's metabolic level meets the increase in energy demand, and the body reaches a steady state. This stress state disappears with the elimination of the stimulus. In addition to consuming stored energy, it does not cause body pathology. Under the action of this mechanism, livestock can adapt to changes in the environment. The third stage is the chronic stress response stage, which may have an adverse effect on the growth, reproduction and health of livestock.

[0003] The impact of transportation on livestock is usually called transportation stress, which refers to the instinctive adaptive and defensive reactions of livestock under the combined effects of stressors such as fasting, environmental changes, bumps, and psychological pressure during transportation. It is one of the important factors affecting meat quality. Unreasonable transportation methods and transportation time can cause livestock to lose water and lose weight; strong stress reactions can lead to glycogen loss in livestock, which in turn affects the quality of meat after slaughter; livestock will lose weight and get scratched when transported by car or train; poor air circulation and narrow space during transportation can also cause livestock to suffocate or even die. Therefore, how to arrange and choose reasonable transportation time and transportation methods is very important for improving meat quality.

[0004] Livestock respond to stressors during transportation through behavioral, immune, hematological and metabolic changes. Studies have shown that after transportation, the plasma GLU concentration of donkeys increases, the CK and LDH activities increase, and the Cor and ACTH concentrations increase. In addition, transportation stress affects the blood indicators of broilers, pigs, sheep, cattle and other livestock and poultry, changes the levels of hormones such as plasma CC and muscle Cor, affects the body's glycogen metabolism through energy metabolism-related enzymes such as plasma CK and LDH, and causes changes in meat quality indicators such as pH, meat color, and drip loss after slaughter.

[0005] Therefore, the research on the regulation of anti-transportation stress before slaughtering livestock is of great significance to ensure the supply of high-quality livestock products. Although there are studies on relieving stress before slaughtering poultry in the existing technology, for example, the Chinese patent with publication number CN109315629A provides an anti-stress agent containing glucose, guanidinoacetic acid glycine, glutamine, theanine, betaine, L-ascorbic acid, tannin, potassium citrate, sodium propionate, sodium diacetate, lactic acid, potassium chloride and sodium bicarbonate. However, it is not effective in preventing and relieving livestock transportation stress, and has no obvious effect on relieving transportation stress and ensuring the high-quality supply of livestock products. Summary of the invention

[0006] The purpose of the present invention is to solve the prior art problems raised in the above background technology and to provide a nutritional additive and application for preventing and alleviating livestock transportation stress.

[0007] The above-mentioned purpose of the present invention is achieved like this:

[0008] In one aspect, the present invention provides a nutritional additive for preventing and alleviating livestock transportation stress, wherein the nutritional additive is composed of the following components, measured in weight percentage:

[0009] 1-10 parts of vitamin B1, 1-10 parts of vitamin B6, 1-18 parts of vitamin C, 1-3 parts of ATP, 1-6 parts of sodium bicarbonate, 1-3 parts of sodium chloride, 1-5 parts of L-lysine hydrochloride, 1-5 parts of inosine, 10-30 parts of glucose, 4-12 parts of astragalus polysaccharide, 4-12 parts of N-carbamylglutamate, and 1-6 parts of traditional Chinese medicine composition.

[0010] Furthermore, the nutritional additive is composed of the following components, in weight percentage:

[0011] 2-9 parts of vitamin B1, 2-9 parts of vitamin B6, 2-17 parts of vitamin C, 2-2.5 parts of ATP, 2-5 parts of sodium bicarbonate, 2-2.5 parts of sodium chloride, 2-4 parts of L-lysine hydrochloride, 2-4 parts of inosine, 11-29 parts of glucose, 5-11 parts of astragalus polysaccharide, 5-11 parts of N-carbamylglutamate, and 2-5 parts of traditional Chinese medicine composition.

[0012] Furthermore, the nutritional additive is composed of the following components, in weight percentage:

[0013] 1 part of vitamin B1, 1 part of vitamin B6, 1 part of vitamin C, 1 part of ATP, 1 part of sodium bicarbonate, 1 part of sodium chloride, 1 part of L-lysine hydrochloride, 1 part of inosine, 10 parts of glucose, 4 parts of astragalus polysaccharide, 4 parts of N-carbamylglutamic acid, and 1 part of traditional Chinese medicine composition.

[0014] Furthermore, the nutritional additive is composed of the following components, in weight percentage:

[0015] 10 parts of vitamin B1, 10 parts of vitamin B6, 18 parts of vitamin C, 3 parts of ATP, 6 parts of sodium bicarbonate, 3 parts of sodium chloride, 5 parts of L-lysine hydrochloride, 5 parts of inosine, 30 parts of glucose, 12 parts of astragalus polysaccharide, 12 parts of N-carbamylglutamic acid, and 6 parts of traditional Chinese medicine composition.

[0016] Furthermore, the nutritional additive is composed of the following components, in weight percentage:

[0017] 5 parts of vitamin B1, 5 parts of vitamin B6, 9 parts of vitamin C, 2 parts of ATP, 3 parts of sodium bicarbonate, 2 parts of sodium chloride, 2.5 parts of L-lysine hydrochloride, 2.5 parts of inosine, 20 parts of glucose, 10 parts of astragalus polysaccharide, 10 parts of N-carbamylglutamic acid, and 3 parts of traditional Chinese medicine composition.

[0018] Furthermore, the Chinese medicine composition is composed of the following components, measured in weight percentage:

[0019] 6-16 parts of Anemarrhena Rhizoma, 8-12 parts of Pollen Typhae, 4-16 parts of Talcum Powder, 10-20 parts of Rhubarb, 5-12 parts of Gentiana Scabra, 8-20 parts of Isatis Radix, 3-12 parts of Olive Root, and 5-12 parts of Clove.

[0020] Furthermore, the Chinese medicine composition is composed of the following components, measured in weight percentage:

[0021] 12 parts of Rhizoma Anemarrhenae, 10 parts of Pollen Typhae, 8 parts of Pulvis Talci, 12 parts of Radix Rhei, 8 parts of Radix Gentianae, 12 parts of Radix Isatidis, 5 parts of Radix Oleae, and 8 parts of Cloves.

[0022] Another aspect of the present invention provides a use of a nutritional additive for preventing and alleviating livestock transportation stress in preparing a product for preventing and alleviating livestock transportation stress.

[0023] Furthermore, the product is a feed additive or a medicine.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] 1. The astragalus polysaccharide in the nutritional additive component of the scheme of the present invention can alleviate the impact of transportation stress on the quality of livestock meat; and the N-carbamylglutamate in the component can reduce the adverse impact of transportation stress on the quality of livestock meat. By supplementing N-carbamylglutamate, muscle energy metabolism can be regulated to reduce the impact of transportation stress on livestock muscle metabolism, thereby achieving the effect of preventing and alleviating transportation stress and ensuring the quality of livestock meat;

[0026] 2. The Chinese medicine composition of the present invention is based on the theory of dialectical treatment and is used to resist livestock transportation stress reaction, enhance the ability of livestock to resist transportation stress, and reduce the adverse reactions of livestock due to stress during transportation;

[0027] 3. The components of the present invention can supplement the nutrition of livestock during transportation by adding vitamin B1, vitamin B6, vitamin C, ATP, sodium bicarbonate, sodium chloride, L-lysine hydrochloride, inosine and glucose, and can prevent and alleviate the stress symptoms of livestock such as mental depression, fear of strangers, anorexia, vomiting, diarrhea, weight loss and the like due to transportation stress. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 is the change of rectal temperature in the embodiment of the present invention ((a) the change trend of rectal temperature before and after transportation and after resting in the case of APS supplementation; (b) the comparison of rectal temperature in the case of APS supplementation and no APS supplementation at three time points before and after transportation and after resting);

[0029] Figure 2 The effects of transportation and rest after APS supplementation on the quality of the longest back muscle of sheep in the embodiment of the present invention (a: the effects of transportation and rest after APS supplementation on the slaughter rate and the quality of the longest back muscle of Mongolian sheep, b: pH (0h), c: pH (24h), d: L value, e: a value, f: b value, g: cooking loss, h: shear force);

[0030] Figure 3 is the effect of APS supplementation on the slaughter rate and meat quality of sheep in the embodiment of the present invention (a: the effect of APS supplementation on the slaughter rate and longissimus dorsi muscle quality of Mongolian sheep, b: pH (0h), c: pH (24h), d: L value, e: a value, f: b value, g: cooking loss, h: shear force);

[0031] Figure 4 is the effect of supplementing APS in the embodiment of the present invention on the quality of mutton slaughtered directly after transportation (a: slaughter rate, b: pH (0h), c: pH (24h), d: L value, e: a value, f: b value, g: cooking loss, h: shear force);

[0032] Figure 5 The effect of APS supplementation on the quality of mutton after 24 hours of rest after transportation in the embodiment of the present invention;

[0033] Figure 6 is the PCA analysis of plasma metabolites in the embodiment of the present invention;

[0034] Figure 7 is the OPLS-DA and RPT analysis of plasma metabolites in the embodiments of the present invention;

[0035] Figure 8is the PCA analysis of plasma metabolites in the embodiment of the present invention;

[0036] Fig. 9 is the OPLS-DA and RPT analysis of the Cvs.A comparison group in the embodiment of the present invention;

[0037] Fig.10 is the PCA analysis of muscle metabolites in the embodiment of the present invention;

[0038] Fig.11 is the OPLS-DA and RPT analysis of metabolites in each comparison group in the examples of the present invention;

[0039] Fig.12 is the PCA analysis of plasma metabolites in the embodiment of the present invention;

[0040] Fig.13 is the OPLS-DA and RPT analysis of the Cvs.A comparison group in the embodiment of the present invention;

[0041] Fig.14 is the change of rectal temperature in the embodiment of the present invention ((a): the change trend of rectal temperature before and after transportation and after resting in the case of NCG supplementation, (b): the comparison of rectal temperature in the case of NCG supplementation and no NCG supplementation at three time points before and after transportation and after resting);

[0042] Fig.15 The effect of transportation and resting after supplementation with NCG on the quality of the longissimus dorsi muscle of sheep in the embodiment of the present invention;

[0043] Fig.16 is the effect of NCG supplementation on sheep quality in the embodiment of the present invention;

[0044] Fig.17 is the effect of supplementing NCG in the embodiment of the present invention on the quality of Mongolian mutton slaughtered directly after transportation;

[0045] Fig.18 is the effect of supplementing NCG on the quality of resting mutton after transportation in the embodiment of the present invention;

[0046] Fig.19 is the PCA analysis of plasma metabolites in the embodiment of the present invention;

[0047] Fig. 20 is the OPLS-DA and RPT analysis of plasma metabolites in each comparison group in the examples of the present invention;

[0048] Fig.21 is the PCA analysis of plasma metabolites in the embodiment of the present invention;

[0049] Fig. 22 It is the OPLS-DA and RPT analysis of plasma metabolites in the Cvs.N comparison group in the embodiment of the present invention;

[0050] Fig.23 is the PCA analysis of muscle metabolites in the embodiment of the present invention;

[0051] Fig.24 is the OPLS-DA and RPT analysis of the metabolites of the longissimus dorsi muscle in each comparison group of the embodiments of the present invention;

[0052] Fig.25 is the PCA analysis of muscle metabolites in the embodiment of the present invention;

[0053] Fig.26 This is the OPLS-DA and RPT analysis of the longissimus dorsi muscle metabolites in the Cvs.N comparison group in the example of the present invention. DETAILED DESCRIPTION

[0054] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0055] The implementation of the present invention is described in detail below in conjunction with specific embodiments.

[0056] The following are preferred embodiments of the present invention.

[0057] Example 1: This example 1 provides a nutritional additive for preventing and alleviating livestock transportation stress. The nutritional additive is composed of the following components, calculated by weight percentage:

[0058] 1 part of vitamin B1, 1 part of vitamin B6, 1 part of vitamin C, 1 part of ATP, 1 part of sodium bicarbonate, 1 part of sodium chloride, 1 part of L-lysine hydrochloride, 1 part of inosine, 10 parts of glucose, 4 parts of astragalus polysaccharide, 4 parts of N-carbamylglutamic acid, and 1 part of traditional Chinese medicine composition.

[0059] The Chinese medicine composition is composed of the following components, calculated by weight percentage:

[0060] 6 parts of Rhizoma Anemarrhenae, 8 parts of Pollen Typhae, 4 parts of Pulvis Talcum, 10 parts of Radix Rhei, 5 parts of Gentiana, 8 parts of Radix Isatidis, 3 parts of Root of Olive, and 5 parts of Clove. The preparation method of the Chinese medicine composition in this embodiment is as follows:

[0061] (1) Weighing the above raw materials according to the above weight parts;

[0062] (2) The raw materials weighed above are mixed, crushed, sieved, and then decocted in water three times, the first decoction time is 1.5h-2h, and the second and third decoction times are 1h-1.5h respectively. The decoctions are then combined, filtered, and the filtrate is concentrated into an extract. Appropriate amounts of sucrose, benzoic acid, and ethylparaben are added, water is added, stirred, allowed to stand, and filtered to obtain a liquid preparation (1g / ml) of a traditional Chinese medicine composition.

[0063] Example 2: This example 2 provides a nutritional additive for preventing and alleviating livestock transportation stress. The nutritional additive is composed of the following components, calculated by weight percentage:

[0064] 10 parts of vitamin B1, 10 parts of vitamin B6, 18 parts of vitamin C, 3 parts of ATP, 6 parts of sodium bicarbonate, 3 parts of sodium chloride, 5 parts of L-lysine hydrochloride, 5 parts of inosine, 30 parts of glucose, 12 parts of astragalus polysaccharide, 12 parts of N-carbamylglutamic acid, and 6 parts of traditional Chinese medicine composition.

[0065] The Chinese medicine composition is composed of the following components, calculated by weight percentage:

[0066] 16 parts of Rhizoma Anemarrhenae, 12 parts of Pollen Typhae, 16 parts of Pulvis Talci, 20 parts of Radix Rhei, 12 parts of Radix Gentianae, 20 parts of Radix Isatidis, 12 parts of Root of Olive, and 12 parts of Cloves.

[0067] The preparation method of the Chinese medicine composition in this embodiment 2 is:

[0068] (1) Weighing the above raw materials according to the above weight parts;

[0069] (2) The raw materials weighed above are mixed, crushed, sieved, and then decocted in water three times, the first decoction time is 1.5h-2h, and the second and third decoction times are 1h-1.5h respectively. The decoctions are then combined, filtered, and the filtrate is concentrated into an extract. Appropriate amounts of sucrose, benzoic acid, and ethylparaben are added, water is added, stirred, allowed to stand, and filtered to obtain a liquid preparation (1g / ml) of a traditional Chinese medicine composition.

[0070] Example 3: This example 3 provides a nutritional additive for preventing and alleviating livestock transportation stress. The nutritional additive is composed of the following components, calculated by weight percentage:

[0071] 5 parts of vitamin B1, 5 parts of vitamin B6, 9 parts of vitamin C, 2 parts of ATP, 3 parts of sodium bicarbonate, 2 parts of sodium chloride, 3 parts of L-lysine hydrochloride, 3 parts of inosine, 20 parts of glucose, 8 parts of astragalus polysaccharide, 8 parts of N-carbamylglutamic acid, and 3 parts of traditional Chinese medicine composition.

[0072] The Chinese medicine composition is composed of the following components, calculated by weight percentage:

[0073] 11 parts of Rhizoma Anemarrhenae, 10 parts of Pollen Typhae, 10 parts of Pulvis Talci, 15 parts of Radix Rhei, 7 parts of Radix Gentianae, 14 parts of Radix Isatidis, 9 parts of Radix Oleae, and 10 parts of Cloves.

[0074] The preparation method of the Chinese medicine composition in this embodiment 3 is:

[0075] (1) Weighing the above raw materials according to the above weight parts;

[0076] (2) The raw materials weighed above are mixed, crushed, sieved, and then decocted in water three times, the first decoction time is 1.5h-2h, and the second and third decoction times are 1h-1.5h respectively. The decoctions are then combined, filtered, and the filtrate is concentrated into an extract. Appropriate amounts of sucrose, benzoic acid, and ethylparaben are added, water is added, stirred, allowed to stand, and filtered to obtain a liquid preparation (1g / ml) of a traditional Chinese medicine composition.

[0077] The nutritional additives of the above embodiments of the present invention are used to prevent and alleviate livestock transportation stress, wherein the core components are astragalus polysaccharides and N-carbamylglutamate.

[0078] The following is an experiment of the embodiment of the present invention:

[0079] 1. Mechanism of Astragalus Polysaccharide in Anti-Stress and Regulating Meat Quality of Livestock

[0080] Astragalus polysaccharide (APS), as a Chinese herbal medicine additive, is a bioactive substance composed of a variety of monosaccharides. It is extracted from the dried roots of Astragalus mongolica or Astragalus membranaceus, a leguminous plant. After APS is fed to livestock, it mainly affects livestock growth performance, antioxidant capacity, immune response and serum metabolic indicators. In this experiment, 18 Mongolian sheep waiting to be marketed were used as the research objects. All sheep were continuously fed APS for 7 days before slaughter. Three treatment groups were set up: direct slaughter without transportation, direct slaughter after 800km transportation, and slaughter after 24h of rest after 800km transportation. The mechanism of APS in anti-stress and regulation of meat quality was analyzed from the aspects of live body temperature physiology, mutton quality, plasma and muscle metabolomics. The effects of the same transportation strategy on the above indicators of mutton sheep with or without APS supplementation were compared. The purpose was to explore the role of APS in the formation of mutton quality after long-distance transportation, so as to reduce the adverse effects of long-distance transportation on mutton sheep.

[0081] 1. Experimental Materials

[0082] 1.1. Livestock and regulatory agents

[0083] 1.1.1. Experimental sheep and APS

[0084] 18 healthy Mongolian sheep aged 2 years old with good body condition and average weight (47.26±4.09) kg were selected as experimental sheep after passing deworming and disease detection. The experimental sheep had no transportation experience. APS was purchased from Shanghai MacLean Biochemical Technology Co., Ltd., item number C832678, in powder form with a purity of 70%.

[0085] 1.1.2 Experimental feed

[0086] A self-prepared complete mixed granular diet was used. The diet composition and nutrient content are shown in Table 1.

[0087] Table 1 Diet composition and nutrient content

[0088]

[0089] 1.2 Experimental instruments

[0090] The main experimental instruments used in this experiment are shown in Table 2, and also include: scissors, tweezers, scalpels, beakers, flat dishes, electronic thermometers, bench scales, transport carts, etc.

[0091] Table 2 Main experimental instruments

[0092]

[0093] 1.3 Experimental reagents and consumables

[0094] The main experimental reagents used in the experiments of the present invention are shown in Table 3.

[0095] Table 3 Main experimental reagents

[0096]

[0097] The main experimental consumables used in the present invention include: syringes, blood collection needles, blood collection tubes, centrifuge tubes, cryopreservation tubes, sealing bags, markers, surgical blades, etc.

[0098] 2. Experimental methods

[0099] 2.1. Grouping and treatment of livestock This experiment was conducted at the Animal Welfare and Healthy Breeding Research and Pilot Base of the Animal Husbandry Research Institute of the Inner Mongolia Academy of Agricultural and Animal Husbandry Sciences (Geerletuya Breeding Cooperative, Siziwang Banner) from May 15 to June 5, 2022. The sheep were divided into 3 groups according to their weight, with 6 sheep in each group and the average weight of the sheep in each group was not significantly different. The groups are shown in Table 4.

[0100] Table 4 Livestock Grouping

[0101]

[0102] Each group of sheep was isolated for 15 days before transportation, and each sheep had about 2m of activity space.2 All sheep were fed with the same formula and equal amount of complete mixed granular diet at 8:00 and 16:00 every day. The management conditions and daily diet supply were consistent before the experiment.

[0103] The three groups of sheep were continuously supplemented with APS for 7 days before the start of the transportation experiment. Each sheep was gavaged with 7.14 g APS at 7:30 am every day (the purity of APS used in the experiment was 70%, that is, each sheep was supplemented with 5.00 g APS per day).

[0104] Before the start of transportation, the sheep in each group were fasted for 12 hours and deprived of water for 6 hours. The vehicle was a long cargo transport vehicle. The transport vehicle was thoroughly disinfected and dried before transportation. The deck of the vehicle was laid with woven straw mats, and partitions were set up inside the vehicle. Each group of sheep had a partition, with a density of about 0.65m2 / sheep. The transportation was 800km. The average temperature on the day of transportation was 22℃, the average relative humidity was 21%, and there was no extremely bad weather. After the transportation, the AT group was slaughtered directly, and the ATR group was slaughtered at the same time as the A group after resting for 24 hours without food or water after the transportation. The weights of the A group, AT group, ATR group, C group, CT group and CTR group were similar.

[0105] 2.2. Measurement of body temperature and weight

[0106] The method is: measure the rectal temperature of sheep before loading and unloading, and before slaughter. When measuring, the thermometer probe is inserted about 5 cm into the anus, and the measurement and reading are carried out according to the instructions for use of the thermometer. Weigh the live weight and carcass weight before slaughter. The determination method is carried out in accordance with the provisions of NY / T1236-2006.

[0107] 2.3 Sample collection

[0108] In addition to the samples required for muscle ultrastructure analysis, other sample collection methods are:

[0109] (1) Plasma: Before slaughtering the sheep, 10 mL of blood was collected from the jugular vein and placed in a sodium citrate anticoagulant tube. After standing to precipitate plasma, the blood was centrifuged at 3000 rpm for 10 min. The separated plasma was divided into 2 mL cryotubes, stored in liquid nitrogen, and then transferred to a -80°C refrigerator for standby use. (2) Muscle: After slaughtering the sheep, according to the requirements of different indicators, the longissimus dorsi muscle on the 12th-13th intercostal spine of the left carcass was selected for the detection of relevant indicators, and the samples were placed under different conditions for standby use according to different detection requirements. The specific sampling site and storage conditions are as follows: 100 g of the longissimus dorsi muscle was selected, the connective tissue was removed, and it was frozen in a ziplock bag for cooking loss and shear force determination. For muscle ultrastructure analysis, the longissimus dorsi muscle between the 6th and 7th vertebrae of the carcass was selected. A newly opened surgical blade was used for sampling to reduce mechanical damage such as pulling, contusion and squeezing. The meat sample was cut into 1 mm × 1 mm × 1 mm pieces, fixed in 2.5% glutaraldehyde, and stored at 4°C for standby use. Metabolomics analysis The longissimus dorsi muscle between the 6th and 7th vertebrae of the carcass was selected, cut into 5 mm × 5 mm × 10 mm pieces, placed in 2 mL sterile enzyme-free cryotubes, stored in liquid nitrogen, brought back to the laboratory and transferred to a −80 °C refrigerator for use.

[0110] 2.4 Meat quality determination

[0111] The method is as follows: (1) Dressing rate: weigh the sheep before slaughter and the carcass after slaughter, and calculate the dressing rate: Dressing rate = carcass weight after slaughter / sheep weight before slaughter × 100%; (2) pH value: 45 minutes after slaughter, use a pH meter to measure the pH value of the longissimus dorsi muscle on the 12th-13th intercostal spine of the carcass, recorded as pH (0h); the sample at this position is stored at 4°C for 24 hours and then measured again, recorded as pH (24h). The measurement is repeated 3 times, and the average value is taken as the result. (3) Color: remove the connective tissue and visible fat of the cross section of the longissimus dorsi muscle on the 12th-13th intercostal spine, use a colorimeter to measure the meat color of the sample at 3 different positions, and take the average value as the result. The indicators include L* value, a* value and b* value.

[0112] 2.5. Blood metabolomics analysis

[0113] LC-MS analysis was performed using an ACQUITY UPLC I-Class plus ultra-high performance liquid chromatography tandem QE high-resolution mass spectrometer (liquid chromatography tandem time-of-flight mass spectrometry) produced by Waters.

[0114] 2.6 Muscle metabolomics analysis

[0115] Sample pretreatment: (1) Take out the sample stored at -80℃, thaw at room temperature, weigh 30mg and put it into a 1.5mL sterile enzyme-free centrifuge tube, add 400μL methanol-water (V:V=4:1, containing L-2-chlorophenylalanine, 4μg / mL) and 2-3 small steel balls; (2) Place the sample in a -40℃ refrigerator, precool for 2min, then put it in a grinder and grind it at 60Hz for 2min; (3) Ultrasonic extraction for 10min in an ice water bath and let it stand at -40℃ for 30min. The subsequent operations are the same as the blood sample treatment in 2.3.7.1. Then use liquid chromatography-mass spectrometry for analysis.

[0116] 3. Results and Discussion

[0117] 3.1. Regulation of body temperature of sheep subjected to transport stress by APS

[0118] In this experimental study, the rectal temperature of the sheep in the APS group was measured before transportation, after transportation, and after transportation and resting for 24 hours.

[0119] The results showed that the core body temperature of the sheep increased significantly after transportation (P<0.05), and decreased significantly after resting for 24 hours (P<0.01) ( Figure 1 ). Compared with the sheep without APS (C group, CT group, CTR group), there was no significant difference in body temperature between group A and group C, the AT group was significantly higher than the CT group (P < 0.05), and there was no significant difference between group A and group CTR. Existing studies have shown that feeding Chinese herbal medicine to dairy cows during heat stress can significantly reduce the rectal temperature of dairy cows. The results of the present invention are close to this, that is, 24 hours after the end of transportation, the rectal temperature of sheep was significantly lower than the temperature measured immediately after the end of transportation. Existing studies on horses have shown that there is no difference in the change of rectal temperature before and after transportation with or without antioxidant supplementation, but the rectal temperature 15 minutes after transportation is significantly higher than the temperature before transportation and 1-21 days after transportation; the rectal temperature of mice also increased significantly after transportation; during the 8-hour transportation of piglets treated in a thermoneutral environment before weaning, the core (abdominal) temperature began to rise 2 hours after transportation, reaching the highest at 7 hours, and then began to decline, and was higher than the baseline at 8 hours. This result is consistent with the trend of body temperature changes in this study.

[0120] 3.2. Regulation of APS on mutton quality

[0121] Meat quality indicators such as meat color (L* value, a* value and b* value), drip loss and tenderness are all related to the pH value. The pH value depends on the rate and degree of LD production by glycolysis after slaughter. Dietary additives can affect meat quality. Existing studies have shown that the addition of garcinol to pig diets can improve muscle antioxidant capacity, reduce the activity of PFKFB3, PFK1, GAPDH and PGK1 by inhibiting PCAF (P300-CBP binding factor)-mediated protein acetylation, slow down post-muscle glycolysis and LD production, and improve meat quality. APS can improve the body's antioxidant function and prevent liver glucose and lipid metabolism disorders induced by transportation stress through the PGC1α / SIRT1 / AMPK / PPARα / PPARγ signaling system. There are also reports that APS can change meat production performance and meat quality such as slaughter rate, cooked meat rate and shear force. In the experimental study of the present invention, after supplementing APS, transportation and rest after transportation had a series of effects on slaughter rate and meat quality (Table 5, Figure 2 ), including slaughter rate, cooking loss, shear force, pH (0h) value, pH (24h) value, L* value, a* value and b* value, all decreased after transportation but the value was not significant; while L* value, a* value and b* value increased significantly after resting after transportation (P<0.05), while slaughter rate, cooking loss, shear force, pH (0h) value and pH (24h) value increased but the value was not significant.

[0122] Table 5 Effects of APS supplementation on slaughter rate and meat quality

[0123]

[0124] Note: a, b: Data without the same letters in the same column indicate significant differences (P<0.05); A, B: Data without the same letters in the same column indicate extremely significant differences (P<0.01).

[0125] Compared with the group without APS supplementation, the pH (24h) value of the directly slaughtered mutton increased significantly after the addition of APS, and the slaughter rate, cooking loss, shear force, L* value, and a* value increased but not significantly, while the pH (0h) value and yellowness value decreased, which was also not significant ( Figure 3 ).

[0126] For sheep slaughtered directly after transportation, compared with the group without APS supplementation, APS supplementation before slaughter can lead to a significant increase in cooking loss, and at the same time, the L* value and a* value are also significantly increased, improving the sensory quality of meat. In addition, APS supplementation before slaughter can also increase the slaughter rate, pH (0h) value and b* value, reduce the shear force and pH (24h) value of meat, and improve meat quality ( Figure 4 ).

[0127] For sheep that were rested for 24 hours before slaughter after transportation, supplementation with APS led to a significant increase in pH (24h) (P < 0.01), which increased the slaughter rate, cooking loss, and shear force but not significantly. Similarly, pH (0h) decreased. When pH (24h) increased significantly, the quality of mutton was improved ( Figure 5 ).

[0128] The pH value of pork is a key factor in determining meat quality, directly or indirectly determining meat quality traits such as meat color, water holding capacity and shear force. The reason for the decrease in muscle pH after slaughter is that LD produced by glycolysis accumulates in the muscle. The rate and degree of pH decrease both affect meat quality. If the ultimate pH value of muscle after slaughter is too low, its water holding capacity will be worse, resulting in increased hardness; if the pH drops too quickly, the meat will turn pale and soft, producing PSE meat; if the pH drops too slowly, the meat will turn black and hard, with high water holding capacity, producing DFD meat. The pH (0h) and pH (24h) of sheep slaughtered directly after APS supplementation and transportation were lower than those without transportation or resting after transportation. The pH value of the longissimus dorsi muscle of sheep without APS supplementation had the same trend, but except for the pH (0h) of sheep without APS supplementation (Group C), which was higher than that of sheep supplemented with APS (Group A), the rest were lower than those of sheep supplemented with APS. Therefore, APS supplementation can effectively reduce the adverse effects of long-distance transportation before slaughter on meat quality.

[0129] 3.3 Effects of transportation and resting on plasma metabolism in sheep after APS supplementation

[0130] Supplementation of APS can improve the stress of livestock to adverse stimuli and have a certain effect on plasma metabolism. The existing technology has studied the effect of APS on the blood metabolomics of dairy cows, showing that supplementation of hawthorn and APS 21 days before delivery of dairy cows resulted in 31, 27, 20 and 42 differential metabolites 7 days before delivery, on the day of calving, 7 days after delivery and 21 days after delivery, respectively. The results of enrichment analysis showed that the addition of hawthorn and APS to the diet can alleviate the negative energy balance by regulating the amino acid metabolism and lipid metabolism of peripartum dairy cows.

[0131] In this study, PCA analysis was performed on plasma metabolites to reflect the variability of plasma metabolism between and within groups after APS supplementation, and the results are as follows: Figure 3-6 As shown in the figure, the total contribution rates of the two principal components PCA1 and PCA2 after dimensionality reduction of the three groups of data to the spectra are 21.1% and 11.31% respectively ( Figure 6 In a), in the PCA1 axis, the AT group and the A group were distributed in the negative and positive directions respectively, with the farthest distance, while the ATR group was in the middle but biased towards the A group. This trend indicates that after the addition of APS, the sheep plasma metabolic profile also changed significantly due to transportation, and quickly returned to the pre-transportation state after rest. Pairwise comparisons of the metabolites in the three groups showed that the plasma metabolic profiles were significantly different between the groups ( Figure 6bd in.

[0132] In order to filter out the noise irrelevant to the classification information, improve the analytical ability and effectiveness of the model, and maximize the differences between different groups within the model to distinguish the overall differences in the metabolic profiles between the groups, OPLS-DA analysis was performed. At the same time, the model quality was examined using the 200 response ranking test method. The results showed that the Avs.AT comparison group was completely separated in two-dimensional space ( Figure 7 a), the RPT scoring model is R2=0.922, Q2=-0.269( Figure 7 b); Avs.ATR comparison groups are completely separated in two-dimensional space ( Figure 7 c), the RPT scoring model is R2=0.922, Q2=-0.147( Figure 7 b); ATvs.ATR comparison groups are also completely separated in two-dimensional space ( Figure 7 e), the RPT scoring model is R2=0.978, Q2=-0.153( Figure 7 b), indicating that there were significant differences in the metabolic profiles among the groups, and the scoring model had good predictive ability without overfitting tendency.

[0133] 3.4 Effects of APS supplementation on sheep plasma metabolism

[0134] PCA analysis of plasma metabolites with and without APS supplementation showed that the total contribution rates of the two principal components PCA1 and PCA2 of the Cvs.A comparison group data after dimensionality reduction were 18.17% and 15.38%, respectively ( Figure 8 In a), the two axial metabolites were not well separated, indicating that in the absence of transport, the addition of astragalus polysaccharides had little effect on plasma metabolism and the difference in metabolic spectrum was not significant. The total contribution rates of the two principal components PCA1 and PCA2 of the CTvs.AT comparison group data after dimensionality reduction to the spectrum were 19.67% and 15.21%, respectively ( Figure 8 b), the CT group and AT group in the PCA1 axis are distributed in the positive and negative directions, respectively, and the distance is the farthest, indicating that the effects of transportation stress on the plasma metabolic profile of mutton sheep supplemented with or without astragalus polysaccharide are different. The total contribution rates of PCA1 and PCA2 to the spectrum in the CTRvs.ATR comparison group were 17.58% and 15.52%, respectively ( Figure 8 In c), the CTR group and the ATR group were distributed in the negative direction and the positive direction, respectively, in the PCA1 axis, and were basically able to be separated, indicating that a short period of rest after transportation had a significant effect on the plasma metabolic profiles of mutton sheep with or without the addition of astragalus polysaccharide.

[0135] The Cvs.A comparison group data were analyzed by OPLS-DA, and the results showed that ( Fig. 9The two groups of data in a) are completely separated in two-dimensional space, and there are significant differences in the metabolic profiles between the groups. RPT analysis shows that the scoring model of this comparison group is R2=0.968, Q2=-0.042 ( Fig. 9 b), the model has good predictive ability.

[0136] 3.5 Effects of transportation and resting after APS supplementation on sheep muscle metabolism

[0137] The results of the second chapter showed that the metabolic profiles of the three groups, namely, no transportation, transportation, and resting after transportation, changed significantly due to transportation and resting. In particular, the metabolic profiles of the transportation treatment group and the resting after transportation group were significantly different from those of the non-transportation group. However, resting after transportation for 24 hours did not fully restore the body, and the metabolic profiles of the transportation treatment group and the resting after transportation group were not significantly different. In this study, in order to generally reflect the variability of muscle metabolism between and within groups after the addition of astragalus polysaccharides, transportation and resting after transportation were performed, PCA analysis of muscle metabolites was performed. The results are as follows: Figure 3-23 As shown in the figure, the total contribution rates of the two principal components PCA1 and PCA2 after dimensionality reduction of the three groups of data to the spectra are 12.78% and 11.17% respectively ( Fig.10 a), the metabolite distribution of the AT group and the ATR group in the PCA1 axis overlapped, while they were distributed negatively and positively with the A group, respectively. This trend indicates that after the addition of APS, the metabolic profile of mutton also changed significantly due to transportation, and resting for 24 hours still failed to restore the body. From the comparison of the three groups ( Fig.10 bd), the metabolites among the comparison groups were completely separated, indicating that although the body was still not able to recover after 24 hours of rest, the metabolic profiles were still significantly different compared with the transport group.

[0138] OPLS-DA analysis was performed on the data between the comparison groups, and the results showed that ( Fig.11 a, c, e in the figure), the comparison groups were completely separated in two-dimensional space, and there were significant differences in the metabolic profiles between the groups. The RPT analysis results showed that the scoring model of the Avs.AT comparison group was R2=0.955, Q2=-0.005 ( Fig.11 b), Avs.ATR comparison group R2 = 0.968, Q2 = -0.102 ( Fig.11 d), ATvs.ATR comparison group R2=0.993, Q2=-0.052 ( Fig.11 f), indicating that the scoring model has good predictive ability and no overfitting tendency.

[0139] 3.6 Effects of transportation and resting after APS supplementation on sheep muscle metabolism

[0140] The research results of the inventors of this application show that the metabolic profiles of the three groups of no transportation, transportation and rest after transportation changed significantly due to transportation and rest. In particular, the metabolic profiles of the transportation treatment group and the rest after transportation group were significantly different from those of the non-transportation group. Rest after transportation for 24 hours did not fully restore the body, and the metabolic profiles of the transportation treatment group and the rest after transportation group were not significantly different. In this study, in order to generally reflect the variability of muscle metabolism between and within groups after the addition of astragalus polysaccharides, transportation and rest after transportation were used to analyze muscle metabolites. The results are as follows: Fig.12 As shown in the figure, the total contribution rates of the two principal components PCA1 and PCA2 after dimensionality reduction of the three groups of data to the spectra are 12.78% and 11.17% respectively ( Fig.12 a), the metabolite distribution of the AT group and the ATR group in the PCA1 axis overlapped, while they were distributed negatively and positively with the A group, respectively. This trend indicates that after the addition of APS, the metabolic profile of mutton also changed significantly due to transportation, and resting for 24 hours still failed to restore the body. From the comparison of the three groups ( Fig.12 bd), the metabolites among the comparison groups were completely separated, indicating that although the body was still not able to recover after 24 hours of rest, the metabolic profiles were still significantly different compared with the transport group.

[0141] OPLS-DA analysis was performed on the data between the comparison groups, and the results showed that ( Fig.13 a, c, e in the figure), the comparison groups were completely separated in two-dimensional space, and there were significant differences in the metabolic profiles between the groups. The RPT analysis results showed that the scoring model of the Avs.AT comparison group was R2=0.955, Q2=-0.005 ( Fig.13 b), Avs.ATR comparison group R2 = 0.968, Q2 = -0.102 ( Fig.13 d), ATvs.ATR comparison group R2=0.993, Q2=-0.052 ( Fig.13 f), indicating that the scoring model has good predictive ability and no overfitting tendency.

[0142] 3.7 Effects of APS supplementation on mutton metabolism

[0143] PCA analysis was performed on muscle metabolites with or without astragalus polysaccharide. The results showed that the total contribution rates of the two principal components PCA1 and PCA2 of the Cvs.A comparison group data after dimensionality reduction were 16.39% and 14.77% respectively ( Fig.14 In a), the two axial metabolites can be basically separated, indicating that the addition of astragalus polysaccharide has an effect on plasma metabolism without transport, but the effect is small. The total contribution rates of the two principal components PCA1 and PCA2 of the CTvs.AT comparison group data after dimensionality reduction to the spectrum are 21.93% and 16.11%, respectively ( Fig.14In b), the spectra of the CT group and the AT group can be well distinguished, indicating that the addition of astragalus polysaccharides can lead to significant changes in the metabolic profile of sheep muscle under transportation stress conditions. The total contribution rates of PCA1 and PCA2 to the spectrum in the CTRvs.ATR comparison group were 18.38% and 14.35%, respectively ( Fig.14 In c), the CTR group and the ATR group were distributed in the negative and positive directions in the PCA1 axis, respectively, and could be completely separated, indicating that under the condition of adding astragalus polysaccharides, the muscle metabolic profile of the sheep also changed significantly after a short period of rest after transportation.

[0144] The OPLS-DA method was used to analyze the Cvs.A comparison group data. The results showed that ( Fig.15 a), there were significant differences in the metabolic profiles between the two groups. RPT analysis showed that the scoring model of this comparison group was R2 = 0.966, Q2 = -0.166 (Figure Fig.15 b), the model has good predictive ability.

[0145] From the above, it can be seen that adding APS before slaughtering and transporting mutton sheep is beneficial to improving the color and tenderness of meat after slaughter, and can effectively alleviate the impact of transportation stress on meat quality. Under the same transportation treatment method before slaughtering mutton sheep, adding APS intervention is beneficial to improving meat quality. The lipid metabolism of mutton sheep may be greatly affected during transportation stress, and the metabolic pathway involving L-acetylcarnitine may be an important way for APS to intervene in metabolism and improve meat quality. Supplementation of APS can change the plasma metabolites of sheep, which may mainly improve the impact of transportation stress on meat quality by regulating sphingomyelin metabolism.

[0146] 2. Mechanism of N-carbamylglutamate in resisting stress and regulating mutton quality

[0147] 1.2 Experimental Materials

[0148] 1.2.1. Livestock and regulatory agents

[0149] 1.2.1.1. Experimental sheep

[0150] 18 healthy Mongolian sheep aged 2 years old with good body condition and average weight (46.23±4.09) kg were selected as experimental sheep after passing deworming and disease detection. The experimental sheep had no transportation experience. NCG was purchased from Huake Jiarui (Beijing) Technology Co., Ltd., item number HWG57681-500g, powder, purity 97%.

[0151] 1.2.1.2 Experimental feed

[0152] A self-prepared complete mixed granular diet was used. The diet composition and nutrient content are shown in Table 2-1.

[0153] 1.2.2 Experimental instruments

[0154] The main experimental instruments used in this experiment are the same as those mentioned above.

[0155] 1.2.3 Experimental reagents and consumables

[0156] The main experimental reagents used in this experiment are the same as the above experimental reagents except NCG.

[0157] 2. Experimental methods

[0158] 2.1. Livestock grouping and treatment

[0159] This experiment was conducted at the Animal Welfare and Healthy Breeding Research and Pilot Base of the Animal Husbandry Research Institute of the Inner Mongolia Academy of Agricultural and Animal Husbandry Sciences (Geerletuya Breeding Cooperative, Siziwang Banner) from May 15 to June 5, 2022. The sheep were divided into 3 groups according to their weight, with 6 sheep in each group and the average weight of the sheep in each group was not significantly different. The groups are shown in the table.

[0160] Table 6 Livestock Grouping

[0161]

[0162] Each group of sheep is isolated separately for 15 days before transportation, and each sheep has an activity space of about 2m 2 , fed at 8:00 and 16:00 every day, all sheep were fed with the same formula and equal amount of complete mixed granular diet, and the management conditions and daily diet supply were consistent before the experiment. Seven days before the start of the transportation experiment, the three groups of sheep were continuously supplemented with NCG, and 2.58g of NCG was administered orally at 7:30 every day (the purity of NCG used in the experiment was 97%, that is, each sheep was supplemented with 2.50g of NCG every day. Before the start of transportation, each group of sheep fasted for 12 hours and deprived of water for 6 hours. The vehicle was a long cargo transport vehicle. The transport vehicle was thoroughly disinfected and dried before transportation, the deck of the vehicle was covered with woven straw mats, and partitions were set up in the vehicle. Each group of sheep had a partition fence with a density of about 0.65m 2 / head. The transportation was 800km. The average temperature on the day of transportation was 22℃, the average relative humidity was 21%, and there was no extreme bad weather. After transportation, the NT group was slaughtered directly, and the NTR group was slaughtered at the same time as the N group after resting for 24 hours without food or water. The NT group was transported and slaughtered at the same time as the CT group in Chapter 2 and the AT group in Chapter 3. The NTR group was transported and slaughtered at the same time as the CTR group in Chapter 2 and the ATR group in Chapter 3.

[0163] 2.2. Measurement of body temperature and weight

[0164] The method is the same as the above experiment.

[0165] 2.3 Sample collection

[0166] The method is the same as the above experiment.

[0167] 2.4 Meat quality determination

[0168] The method is the same as the above experiment.

[0169] 2.5. Blood metabolomics analysis

[0170] 2.6 Muscle metabolomics analysis

[0171] The method is the same as the above experiment.

[0172] 3. Results and Discussion

[0173] 3.1 Effects of NCG on the body temperature of sheep subjected to transportation stress

[0174] This experimental study measured the rectal temperature of sheep before transportation, after transportation, and after transportation and resting for 24 hours under the premise of NCG supplementation. The results showed that the core body temperature of sheep increased significantly after transportation, and was significantly lower than after transportation after resting for 24 hours, and there was no significant difference from before transportation. The trend of body temperature change was consistent with that when APS was supplemented in Chapter 3, and different from the trend of body temperature change of sheep without regulator supplementation. The rectal temperature of all sheep was within the normal range. The body temperature of sheep dropped after transportation without regulator supplementation, which may be because the heat dissipation was greater than the heat production. When APS or NCG was added, the transportation conditions were the same as those without regulator supplementation, that is, the heat dissipation was consistent, but the rectal temperature rose after transportation. It may be that the regulator promoted metabolic heat production during transportation, which is beneficial for sheep to resist the increase in heat dissipation during transportation ( Fig.14 ).

[0175] 3.2. Regulation of mutton quality by NCG

[0176] Adding NCG to feed can promote muscle growth. In this study, transportation after NCG supplementation and rest after transportation had a series of effects on slaughter rate and meat quality (Table 7, Fig.15 ). Among them, compared with slaughtering directly after transportation, the cooking loss of slaughtering after transportation was significantly reduced (P < 0.05), and compared with slaughtering directly without transportation, the cooking loss of slaughtering after transportation was extremely significantly reduced (P < 0.01). At the same time, the a value of meat after transportation can be significantly reduced. In addition, supplementation of NCG can increase the slaughter rate after transportation, reduce shear force, and improve the quality of meat. The pH (0h) of sheep slaughtered immediately after transportation was significantly higher than that of sheep not transported and those resting after transportation. Studies have shown that an important reason for the change in pH value in the early post-slaughter period is that cell respiration changes from aerobic to anaerobic after slaughter, resulting in a rapid decrease in cell energy and activation of glycolytic enzyme activity to enhance glycolysis and generate LD. The pH (0h) of the NT group was the highest, indicating that its glycolysis reaction was slower than that of the N group and the NTR group, which may be due to low muscle glycogen content or low activity of related enzymes; while the pH (24h) of the NT group was lower than that of the N group and the NTR group, that is, its pH value decreased faster and to a greater extent, which was easy to have an adverse effect on meat quality.

[0177] Table 7 Effects of NCG supplementation on dressing rate and meat quality

[0178]

[0179] Note: Data in the same row without letter headers or with the same letters indicate no significant difference, without the same lowercase letters indicate significant difference (P<0.05), and without the same uppercase letters indicate extremely significant difference (P<0.01).

[0180] Compared with the group without NCG supplementation, the shear force of mutton decreased after NCG supplementation, especially in the groups slaughtered directly after transportation (CT and NT), where the shear force decreased significantly, while the other indicators did not change significantly ( Fig.16 , Fig.17 , Fig.18 ). Therefore, supplementation of NCG can improve meat quality and reduce the adverse effects of long-distance transportation on meat quality.

[0181] 3.3 Effects of transportation and rest on sheep plasma metabolism after NCG supplementation

[0182] Supplementation of NCG in feed can affect plasma metabolism, thereby improving the quality of mutton after transportation. Liu Luo used the LC-MS method to perform non-targeted metabolomics analysis on horse serum after transportation and recovery after transportation, indicating that 68 differential metabolites were screened after transportation, 47 differential metabolites were found between resting for 12 hours and not resting, and 74 differential metabolites were screened between resting and before transportation. The pathway enrichment results showed that neurochemical metabolism was mainly involved in the process of transportation and recovery. The results of the metabolomics study of serum and urine of sheep with different transportation durations and recovery durations using hydrogen magnetic resonance spectroscopy showed that GLU, lipids (lipoproteins, betaine) and amino acids (leucine, isoleucine) in the blood, and betaine and sarcosine in the urine changed significantly, indicating that the intestinal and energy metabolism of sheep changed and muscles decomposed during transportation. In this study, in order to generally reflect the variability of plasma metabolism between and within groups after NCG supplementation, transportation and resting after transportation, PCA analysis of plasma metabolites under different treatment conditions was performed. The results are as follows: Fig.19 As shown in the figure, the total contribution rates of the two principal components PCA1 and PCA2 after dimensionality reduction of the three groups of data to the spectra are 16.52% and 12.71% respectively ( Fig.19 In a), the NT group and the N group were distributed in the positive and negative directions, respectively, with the farthest distance in the PCA1 axis, while the NTR group was in the middle but biased towards the N group. This trend indicates that after supplementation with NCG, the plasma metabolic profile of sheep changed significantly due to transportation, and quickly returned to the pre-transportation state after rest. Pairwise comparisons of the metabolites in the three groups showed that the plasma metabolic profiles were significantly different between the groups ( Fig.19 bd in.

[0183] The OPLS-DA method was used to analyze the overall differences in metabolic profiles between the groups, and the RPT method was used to examine the quality of the model. The results showed that the Nvs.NT comparison group was completely separated in two-dimensional space ( Fig. 20 a), the RPT scoring model is R2=0.903, Q2=-0.37( Fig. 20 b); Nvs.NTR comparison groups are completely separated in two-dimensional space ( Fig. 20 c), the RPT scoring model is R2=0.996, Q2=0.036( Fig. 20 d); NTvs.NTR comparison groups are also completely separated in two-dimensional space ( Fig. 20 e), the RPT scoring model is R2=0.902, Q2=-0.563( Fig. 20 f), indicating that there were significant differences in the metabolic profiles among the groups, and the scoring model had good predictive ability without overfitting tendency.

[0184] 3.4 Effects of NCG supplementation on sheep plasma metabolism

[0185] PCA analysis of plasma metabolites with and without NCG addition showed that the total contribution rates of the two principal components PCA1 and PCA2 to the spectrum of the Cvs.N comparison group data after dimensionality reduction were 21.82% and 17.28%, respectively ( Fig.21 a), the PCA2 axis of the C group and the N group were distributed in the negative and positive directions, respectively, indicating that the addition of N-carbamylglutamate also had a certain effect on plasma metabolism without transport. The total contribution rates of the two principal components PCA1 and PCA2 of the CTvs.NT comparison group data after dimensionality reduction to the spectrum were 18.4% and 13.56%, respectively ( Fig.21 b), the PCA1 axis of the CT group and the NT group distributed negatively and positively, respectively, indicating that NCG supplementation can lead to significant changes in the plasma metabolic profile of sheep under transportation stress conditions. The total contribution rates of PCA1 and PCA2 to the spectrum in the CTRvs.NTR comparison group were 21.05% and 13.7%, respectively ( Fig.21 In c), the PCA1 axis of the CTR group and the NTR group were distributed in the negative and positive directions, respectively, indicating that under the condition of adding NCG, the plasma metabolic profile of the sheep also changed significantly after a short period of rest after transportation.

[0186] The Cvs.N comparison group data were analyzed by OPLS-DA, and the results showed that ( Fig. 22 The two groups of data in a) are completely separated in two-dimensional space, and there are significant differences in the metabolic profiles between the groups. RPT analysis shows that the scoring model of this comparison group is R2=0.911, Q2=-0.503 ( Fig. 22 b), the model has good predictive ability.

[0187] 3.5 Effects of transportation and resting on sheep muscle metabolism after NCG supplementation

[0188] The research results of the inventors of the present application show that the muscle metabolic profiles of the three groups, namely, no transportation, transportation, and resting after transportation, changed significantly due to transportation and resting. In particular, the metabolic profiles of the transportation-treated group and the resting group after transportation were significantly different from those of the non-transported group. Resting for 24 hours after transportation did not fully restore the body, and there was no significant difference in the metabolic profiles between the transportation-treated group and the resting group after transportation. The results of the third chapter show that the metabolic profile of mutton also changed significantly due to transportation after APS supplementation, and resting for 24 hours still failed to restore the body. In this study, in order to generally reflect the variability of muscle metabolism between and within groups after the addition of NCG, transportation and resting after transportation, PCA analysis of muscle metabolites was performed, and the results are as follows: Fig.23 As shown in the figure, the total contribution rates of the two principal components PCA1 and PCA2 after dimensionality reduction of the three groups of data to the spectra are 13.38% and 11.65% respectively ( Fig.23 In a), the metabolites of the N and NT groups in the PCA1 axis were distributed in the negative and positive directions, respectively, while those of the NTR group were in between. This trend indicates that after supplementation with NCG, the metabolic profile of mutton also changed significantly due to transportation. Resting for 24 hours can quickly restore the body, but it did not return to the level before transportation. From the comparison of the three groups ( Fig.23 bd), the metabolites among the comparison groups were completely separated, and the metabolic profiles were significantly different.

[0189] OPLS-DA analysis was performed on the data between the comparison groups, and the results showed that ( Fig.24 a, c, e in the figure), the comparison groups were completely separated in two-dimensional space, and there were significant differences in the metabolic profiles between the groups. The RPT analysis results showed that the scoring model of the Nvs.NT comparison group was R2=0.958, Q2=-0.268 ( Fig.24 b), Nvs.NTR comparison group R2 = 0.978, Q2 = -0.312 ( Fig.24 d), NT vs. NTR comparison group R2 = 0.953, Q2 = -0.322 ( Fig.24 f), indicating that the scoring model has good predictive ability and no overfitting tendency.

[0190] 3.6 Effects of NCG supplementation on sheep muscle metabolism

[0191] PCA analysis of muscle metabolites in patients with or without NCG supplementation showed that the total contribution rates of the two principal components PCA1 and PCA2 to the spectrum of the Nvs.A comparison group data after dimensionality reduction were 21.48% and 15.7%, respectively ( Fig.25In a), the two axial metabolites can be effectively separated, indicating that the addition of N-carbamylglutamate affects muscle metabolism without transport and makes muscle metabolism tend to be uniform. The total contribution rates of the two principal components PCA1 and PCA2 of the CTvs.NT comparison group data after dimensionality reduction to the spectrum were 20.94% and 15.36%, respectively ( Fig.25 In b), the spectra of the CT group and the NT group can be well distinguished, indicating that NCG supplementation can lead to significant changes in the metabolic profile of sheep muscle under transportation stress conditions. The total contribution rates of PCA1 and PCA2 to the spectrum in the CTRvs.NTR comparison group were 18.81% and 15.6%, respectively ( Fig.25 c), the PCA1 axis of the CTR group and the NTR group were distributed in the negative and positive directions, respectively, and could be completely separated, indicating that under the condition of NCG supplementation, the sheep had a short rest after transportation, and the muscle metabolic profile also changed significantly. The OPLS-DA method was used to analyze the Cvs.N comparison group data, and the results showed that ( Fig.26 a), there were significant differences in the metabolic profiles between the two groups. RPT analysis showed that the scoring model for this comparison group was R2 = 0.974, Q2 = -0.149 ( Fig.26 b), the model has good predictive ability.

[0192] In summary, in the above experiments, adding NCG before transportation can effectively improve the tenderness of mutton and reduce cooking loss; and, regardless of whether it is transported or not, NCG supplementation has a significant effect on muscle metabolism, and after NCG supplementation, the muscle metabolic profile tends to be homogenized; in addition, the study of key plasma metabolites showed that 16-methylheptadecanoylcarnitine (16-Methylheptadecanoylcarnitine), octadec-6-enoylcarnitine (Octadec-6-enoylcarnitine) and palmitoylcarnitine (Palmitoylcarnitine) play an important role in regulating transportation stress; (4) NCG supplementation may attenuate the impact of transportation stress on muscle metabolomics by regulating muscle energy metabolism.

[0193] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A nutritional additive for preventing and alleviating livestock transportation stress, characterized in that: The nutritional additive is composed of the following components, in weight percentage: 1-10 parts of vitamin B1, 1-10 parts of vitamin B6, 1-18 parts of vitamin C, 1-3 parts of ATP, 1-6 parts of sodium bicarbonate, 1-3 parts of sodium chloride, 1-5 parts of L-lysine hydrochloride, 1-5 parts of inosine, 10-30 parts of glucose, 4-12 parts of astragalus polysaccharide, 4-12 parts of N-carbamylglutamate, and 1-6 parts of traditional Chinese medicine composition.

2. A nutritional additive for preventing and alleviating livestock transportation stress according to claim 1, characterized in that: The nutritional additive is composed of the following components, in percentage by weight: 2-9 parts of vitamin B1, 2-9 parts of vitamin B6, 2-17 parts of vitamin C, 2-2.5 parts of ATP, 2-5 parts of sodium bicarbonate, 2-2.5 parts of sodium chloride, 2-4 parts of L-lysine hydrochloride, 2-4 parts of inosine, 11-29 parts of glucose, 5-11 parts of astragalus polysaccharide, 5-11 parts of N-carbamylglutamate, and 2-5 parts of traditional Chinese medicine composition.

3. A nutritional additive for preventing and alleviating livestock transportation stress according to claim 1, characterized in that: The nutritional additive is composed of the following components, in weight percentage: 1 part of vitamin B1, 1 part of vitamin B6, 1 part of vitamin C, 1 part of ATP, 1 part of sodium bicarbonate, 1 part of sodium chloride, 1 part of L-lysine hydrochloride, 1 part of inosine, 10 parts of glucose, 4 parts of astragalus polysaccharide, 4 parts of N-carbamylglutamic acid, and 1 part of traditional Chinese medicine composition.

4. A nutritional additive for preventing and alleviating livestock transportation stress according to claim 1, characterized in that: The nutritional additive is composed of the following components, in percentage by weight: 10 parts of vitamin B1, 10 parts of vitamin B6, 18 parts of vitamin C, 3 parts of ATP, 6 parts of sodium bicarbonate, 3 parts of sodium chloride, 5 parts of L-lysine hydrochloride, 5 parts of inosine, 30 parts of glucose, 12 parts of astragalus polysaccharide, 12 parts of N-carbamylglutamic acid, and 6 parts of traditional Chinese medicine composition.

5. A nutritional additive for preventing and alleviating livestock transportation stress according to claim 1, characterized in that: The nutritional additive is composed of the following components, in weight percentage: 5 parts of vitamin B1, 5 parts of vitamin B6, 9 parts of vitamin C, 2 parts of ATP, 3 parts of sodium bicarbonate, 2 parts of sodium chloride, 2.5 parts of L-lysine hydrochloride, 2.5 parts of inosine, 20 parts of glucose, 10 parts of astragalus polysaccharide, 10 parts of N-carbamylglutamic acid, and 3 parts of traditional Chinese medicine composition.

6. A nutritional additive for preventing and alleviating livestock transportation stress according to claim 1, characterized in that: The Chinese medicine composition is composed of the following components, calculated by weight percentage: 6-16 parts of Anemarrhena Rhizoma, 8-12 parts of Pollen Typhae, 4-16 parts of Talcum Powder, 10-20 parts of Rhubarb, 5-12 parts of Gentiana Scabra, 8-20 parts of Isatis Radix, 3-12 parts of Olive Root, and 5-12 parts of Clove.

7. A nutritional additive for preventing and alleviating livestock transportation stress according to claim 6, characterized in that: The Chinese medicine composition is composed of the following components, calculated by weight percentage: 12 parts of Rhizoma Anemarrhenae, 10 parts of Pollen Typhae, 8 parts of Pulvis Talci, 12 parts of Radix Rhei, 8 parts of Radix Gentianae, 12 parts of Radix Isatidis, 5 parts of Radix Oleae, and 8 parts of Cloves.

8. Use of a nutritional additive for preventing and alleviating livestock transportation stress according to any one of claims 1 to 7 in the preparation of a product for preventing and alleviating livestock transportation stress.

9. The use according to claim 8, characterized in that: The product is a feed additive or a medicine.

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