Feed containing quercetagetin and preparation method thereof

Abstract

The invention discloses feed containing quercetagetin and a preparation method of the feed, and belongs to the field of feed. By adding the quercetagetin into the basic feed, the adverse effect of heat stress on the female rabbits can be effectively relieved, the conception rate, the comprehensive litter size and the comprehensive live litter size of the female rabbits can be increased, the reproductive performance is optimized, the growth of the young rabbits is promoted, and meanwhile the breeding economic benefits are improved; the feed prepared by the invention can effectively reduce the content of MDA in animal serum, reduce the damage of lipid peroxidation reaction to organisms, effectively improve the diversity of animal intestinal microorganisms under heat stress, and provide an effective nutrition regulation strategy for animal breeding in summer.

Description

Technical Field

[0001] The present invention belongs to the field of feeds, and particularly relates to a feed containing quercetagetin and a preparation method thereof. Background Art

[0002] As one of the environmental stresses that cannot be ignored in modern animal husbandry, heat stress has an obvious impact on the growth performance of rabbits. High temperature will cause a decrease in the feed intake of rabbits, affect nutrient metabolism, resulting in slow growth and reduced feed utilization rate. It will also damage the intestinal barrier function, increase the risk of disease, and have a negative impact on slaughter and reproductive performance. Under high temperature conditions, the physiological functions of female rabbits are challenged, leading to a decline in a series of reproductive performance indicators. Specifically, heat stress will reduce the conception rate of female rabbits, that is, the proportion of successful pregnancies is significantly reduced. This is mainly due to the disorder of hormone levels in female rabbits under high temperature environment, which affects the normal implantation process of fertilized eggs after mating. In addition, heat stress will also reduce the litter size of female rabbits, making the number of rabbits born per litter significantly decrease. This not only affects the reproductive efficiency of female rabbits, but also increases the breeding cost. More seriously, heat stress will also increase the mortality rate of rabbit pups. The high temperature environment poses a great threat to the growth and development of rabbit pups, resulting in a significant reduction in the survival rate of rabbit pups. In view of the negative impact of heat stress on the reproductive performance of female rabbits, it is particularly important to find effective mitigation measures.

[0003] In recent years, plant active substances have attracted much attention due to their excellent antioxidant capacity. Some studies have shown that adding cardamonin to the feed can regulate some serum biochemical indexes of heat-stressed Danzhou chickens, promote the development of immune organs, enhance the body's immune capacity, relieve liver damage caused by heat stress, improve the body's antioxidant capacity, and improve immune stress and inflammation induced by heat stress; heat stress will also affect the production performance and antioxidant performance of Wenchang chickens. Adding ethanol extract of cassava leaves to the diet can relieve the negative impact of heat stress on experimental chickens. And plant polyphenols have also been widely used to relieve animal oxidative damage and enhance the body's immunity due to their antiviral, antioxidant and anti-inflammatory properties. Existing studies have found that curcumin, as a common polyphenol substance, can effectively improve H2O2-induced oxidative stress, intestinal epithelial damage and mitochondrial damage of porcine intestinal epithelial cells by activating adenosine monophosphate-activated protein kinase and inducing Parkin (a protein related to mitochondrial autophagy) -dependent mitochondrial autophagy. The above studies show that natural plant extracts play an important role in relieving animal oxidative stress. Therefore, exploring the effects of quercetagetin (QG) on the reproductive performance of female rabbits and the growth performance of rabbit pups under heat stress can provide new strategies and theoretical basis for the rabbit industry to cope with heat stress problems. Summary of the Invention

[0004] The technical problem to be solved by the present invention is: to provide a feed containing quercetagetin and a preparation method thereof, so as to solve the technical problem of the reduced reproductive ability of animals under heat stress.

[0005] To achieve the above object, the technical solution adopted by the present invention is: to provide a feed containing quercetagetin, which is composed of a basal diet and quercetagetin (QG) mixed in a mass ratio of 1 kg: 100 - 600 mg. The raw materials used in the basal diet include, by weight: 10 - 15 parts of corn, 3 - 7 parts of wheat middlings, 3 - 7 parts of corn bran, 5 - 10 parts of wheat bran, 0.5 - 2.5 parts of soybean oil, 10 - 20 parts of soybean meal, 3 - 9 parts of rapeseed meal mixture, 5 - 13 parts of distillers grains, 5 - 8 parts of corn germ meal, 0.5 - 2.5 parts of artemisia powder, 3 - 9 parts of rice hull powder, 3 - 7 parts of peanut shell powder, 5 - 10 parts of white rod powder, 5 - 10 parts of alkali grass, and 3 - 7 parts of premix.

[0006] On the basis of the above technical solution, the present invention can also be improved as follows:

[0007] Further, the feed is composed of a basal diet and quercetagetin mixed in a mass ratio of 1 kg: 600 mg.

[0008] Further, the raw materials used in the basal diet include, by weight: 12 parts of corn, 5 parts of wheat middlings, 5 parts of corn bran, 8.5 parts of wheat bran, 1.5 parts of soybean oil, 14 parts of soybean meal, 6 parts of rapeseed meal mixture, 9 parts of distillers grains, 6.5 parts of corn germ meal, 1.5 parts of artemisia powder, 6 parts of rice hull powder, 5 parts of peanut shell powder, 7 parts of white rod powder, 8 parts of alkali grass, and 5 parts of premix.

[0009] Further, the raw materials used in the basal diet include, by weight: 10 parts of corn, 3 parts of wheat middlings, 3 parts of corn bran, 10 parts of wheat bran, 2.5 parts of soybean oil, 20 parts of soybean meal, 3 parts of rapeseed meal mixture, 5 parts of distillers grains, 5 parts of corn germ meal, 0.5 parts of artemisia powder, 3 parts of rice hull powder, 7 parts of peanut shell powder, 10 parts of white rod powder, 10 parts of alkali grass, and 7 parts of premix.

[0010] Further, the raw materials used in the basal diet include, by weight: 15 parts of corn, 7 parts of wheat middlings, 7 parts of corn bran, 5 parts of wheat bran, 0.5 parts of soybean oil, 10 parts of soybean meal, 9 parts of rapeseed meal mixture, 13 parts of distillers grains, 8 parts of corn germ meal, 2.5 parts of artemisia powder, 9 parts of rice hull powder, 3 parts of peanut shell powder, 5 parts of white rod powder, 5 parts of alkali grass, and 3 parts of premix.

[0011] Furthermore, per kilogram of the premix includes: 50 - 100 mg of Fe, 10 - 30 mg of Cu, 50 - 100 mg of Zn, 5 - 15 mg of Mn, 0.1 - 0.2 mg of Co, 0.1 - 0.3 mg of I, 0.2 - 0.3 mg of Se, 9000 - 11000 IU of VA, 800 - 1000 IU of VD, 30 - 70 mg of VE, 1 - 3 mg of VK, 1 - 3 mg of VB1, 4 - 8 mg of VB2, 30 - 70 mg of VB5, 1 - 3 mg of VB6, VB 12 0.01 - 0.03 mg, 30 - 70 mg of VB3, 30 - 60 mg of VB9, 900 - 1100 mg of VB4, and 0.1 - 0.3 mg of VB7.

[0012] Furthermore, per kilogram of the premix includes: 70 mg of Fe, 20 mg of Cu, 70 mg of Zn, 10 mg of Mn, 0.15 mg of Co, 0.2 mg of I, 0.25 mg of Se, 10000 IU of VA, 900 IU of VD, 50 mg of VE, 2 mg of VK, 2 mg of VB1, 6 mg of VB2, 50 mg of VB5, 2 mg of VB6, VB 12 0.02 mg, 50 mg of VB3, 44 mg of VB9, 1000 mg of VB4, and 0.2 mg of VB7.

[0013] The present invention also discloses a method for preparing the above - mentioned feed containing quercetagetin, comprising the following steps: mixing the raw materials evenly in proportion to obtain the feed containing quercetagetin.

[0014] The beneficial effects of the present invention are as follows:

[0015] 1. By adding QG to the basal diet, the present invention can effectively alleviate the adverse effects of heat stress on female rabbits, improve the conception rate, total litter size, and total live litter size of female rabbits. While optimizing the reproductive performance and promoting the growth of young rabbits, it can improve the breeding economic benefits, providing an effective nutritional regulation strategy for meat rabbit breeding in summer.

[0016] 2. By adding QG to the basal diet, it can significantly increase the levels of P, SOD, T - AOC, GSH - Px, and IL - 10 in the serum of breeding female rabbits, and at the same time significantly reduce the content of MDA in the serum (P < 0.05). This shows that quercetagetin can effectively reduce the content of MDA in the serum of female rabbits, reduce the damage caused by lipid peroxidation reaction to the body; at the same time, enhance the activity of antioxidant enzymes, improve the body's own ability to scavenge free radicals, thereby effectively maintaining the balance state of oxidation and antioxidation in the body and reducing the adverse effects of heat stress on female rabbits.

[0017] 3. Adding QG to the basal diet can effectively improve the intestinal microbial diversity of female rabbits under heat stress, have a certain impact on the composition of the intestinal flora at the phylum and genus levels, change the relative abundances of some dominant flora. In addition, LEfSe analysis further identifies the microbial taxa with significant differences between the two groups, revealing the potential of QG in regulating the intestinal microbial community structure and providing a theoretical basis for the application of QG in animal husbandry. Description of the Drawings

[0018] Figure 1 are the data of the temperature-humidity index in the rabbit house;

[0019] Figure 2 is the statistical chart of the heat stress state;

[0020] Figure 3 are the concentrations of P in the sera of female rabbits in the control group and the experimental group on the 15th day of pregnancy under heat stress;

[0021] Figure 4 are the concentrations of E2 in the sera of female rabbits in the control group and the experimental group on the 15th day of pregnancy under heat stress;

[0022] Figure 5 are the concentrations of SOD in the sera of female rabbits in the control group and the experimental group on the 15th day of pregnancy under heat stress;

[0023] Figure 6 are the concentrations of T-AOC in the sera of female rabbits in the control group and the experimental group on the 15th day of pregnancy under heat stress;

[0024] Figure 7 are the concentrations of GSH-Px in the sera of female rabbits in the control group and the experimental group on the 15th day of pregnancy under heat stress;

[0025] Figure 8 are the concentrations of MDA in the sera of female rabbits in the control group and the experimental group on the 15th day of pregnancy under heat stress;

[0026] Figure 9 are the concentrations of IL-6 in the sera of female rabbits in the control group and the experimental group on the 15th day of pregnancy under heat stress;

[0027] Figure 10 are the concentrations of IL-10 in the sera of female rabbits in the control group and the experimental group on the 15th day of pregnancy under heat stress;

[0028] Figure 11 is the Venn diagram of the intestinal microbiota OUT of heat-stressed rabbits;

[0029] Figure 12 is the effect of QG on the Chao 1 index of the intestinal microbiota of heat-stressed rabbits;

[0030] Figure 13Effect of QG on Shannon index of intestinal microbiota in heat-stressed female rabbits;

[0031] Figure 14 Effect of QG on Simpson index of intestinal microbiota in heat-stressed female rabbits;

[0032] Figure 15 Effect of QG on β-diversity of intestinal microbiota in heat-stressed female rabbits;

[0033] Figure 16 Effect of QG on the composition of intestinal flora at the phylum level in heat-stressed female rabbits;

[0034] Figure 17 Relative abundance values of Firmicutes in the control group and the experimental group;

[0035] Figure 18 Relative abundance values of Bacteroidetes in the control group and the experimental group;

[0036] Figure 19 Relative abundance values of Cyanobacteria in the control group and the experimental group;

[0037] Figure 20 Relative abundance values of Verrucomicrobia in the control group and the experimental group;

[0038] Figure 21 Relative abundance values of Patellibacter in the control group and the experimental group;

[0039] Figure 22 Relative abundance values of Proteobacteria in the control group and the experimental group;

[0040] Figure 23 Effect of QG on the composition of intestinal flora at the genus level in heat-stressed female rabbits;

[0041] Figure 24 Relative abundance values of unclassified_f_Muribaculaceae in the control group and the experimental group;

[0042] Figure 25 Relative abundance values of unclassified_f Lachnospiraceae in the control group and the experimental group;

[0043] Figure 26 Relative abundance values of unclassified_o_Clostridia_vadinBB60_group in the control group and the experimental group;

[0044] Figure 27 Relative abundance values of Bacteroides in the control group and the experimental group;

[0045] Figure 28is the relative abundance value of unclassified_o_Clostridia_UCG-014 in the control group and the experimental group;

[0046] Figure 29 is the relative abundance value of Ruminococcus in the control group and the experimental group;

[0047] Figure 30 is the LEfSe analysis chart of the control group and the experimental group;

[0048] Figure 31 is the phylogenetic branching diagram of the control group and the experimental group. Specific implementation manners

[0049] The specific implementation manners of the present invention will be described below to facilitate those skilled in the art to understand the present invention. For those conditions not specified in the examples, they are carried out according to conventional conditions or the conditions recommended by the manufacturer. For the reagents or instruments not indicating the manufacturer, they are all conventional products that can be obtained through commercial purchase. However, it should be clear that the present invention is not limited to the scope of the specific implementation manners. For those ordinary skilled in the art in the technical field, as long as various changes are within the spirit and scope of the present invention defined and determined by the appended claims, these changes are obvious, and all inventions and creations using the concept of the present invention are within the scope of protection.

[0050] Example 1

[0051] A feed containing quercetagetin, which is a mixture of a basal diet and quercetagetin in a mass ratio of 1 kg: 600 mg. The raw materials used in the basal diet include, by weight: 12 parts of corn, 5 parts of wheat middlings, 5 parts of corn bran, 8.5 parts of wheat bran, 1.5 parts of soybean oil, 14 parts of soybean meal, 6 parts of rapeseed meal mixture, 9 parts of black brewer's grains, 6.5 parts of corn germ meal, 1.5 parts of artemisia powder, 6 parts of rice hull powder, 5 parts of peanut hull powder, 7 parts of white rod powder, 8 parts of alkali grass, and 5 parts of premix.

[0052] Each kilogram of the premix includes: 70 mg of Fe, 20 mg of Cu, 70 mg of Zn, 10 mg of Mn, 0.15 mg of Co, 0.2 mg of I, 0.25 mg of Se, 10000 IU of VA, 900 IU of VD, 50 mg of VE, 2 mg of VK, 2 mg of VB1, 6 mg of VB2, 50 mg of VB5, 2 mg of VB6, VB 12 0.02 mg, 50 mg of VB3, 44 mg of VB9, 1000 mg of VB4, and 0.2 mg of VB7.

[0053] Example 2

[0054] A feed containing quercetagetin, which is prepared by mixing a basal diet and quercetagetin at a mass ratio of 1 kg: 400 mg. The raw materials used in the basal diet, by weight, include: 10 parts of corn, 3 parts of wheat middlings, 3 parts of corn bran, 10 parts of wheat bran, 2.5 parts of soybean oil, 20 parts of soybean meal, 3 parts of rapeseed meal mixture, 5 parts of spent dark beer grains, 5 parts of corn germ meal, 0.5 part of artemisia powder, 3 parts of rice hull powder, 7 parts of peanut hull powder, 10 parts of white rod powder, 10 parts of alkali grass, and 7 parts of premix.

[0055] Each kilogram of the premix contains: 50 mg of Fe, 10 mg of Cu, 50 mg of Zn, 15 mg of Mn, 0.2 mg of Co, 0.3 mg of I, 0.3 mg of Se, 9000 IU of VA, 800 IU of VD, 70 mg of VE, 1 mg of VK, 1 mg of VB1, 4 mg of VB2, 30 mg of VB5, 3 mg of VB6, VB 12 0.03 mg, 70 mg of VB3, 60 mg of VB9, 1100 mg of VB4, and 0.3 mg of VB7.

[0056] Example 3

[0057] A feed containing quercetagetin, which is prepared by mixing a basal diet and quercetagetin at a mass ratio of 1 kg: 100 mg. The raw materials used in the basal diet, by weight, include: 15 parts of corn, 7 parts of wheat middlings, 7 parts of corn bran, 5 parts of wheat bran, 0.5 part of soybean oil, 10 parts of soybean meal, 9 parts of rapeseed meal mixture, 13 parts of spent dark beer grains, 8 parts of corn germ meal, 2.5 parts of artemisia powder, 9 parts of rice hull powder, 3 parts of peanut hull powder, 5 parts of white rod powder, 5 parts of alkali grass, and 3 parts of premix.

[0058] Each kilogram of the premix contains: 100 mg of Fe, 30 mg of Cu, 100 mg of Zn, 5 mg of Mn, 0.1 mg of Co, 0.1 mg of I, 0.2 mg of Se, 11000 IU of VA, 1000 IU of VD, 30 mg of VE, 3 mg of VK, 3 mg of VB1, 8 mg of VB2, 70 mg of VB5, 1 mg of VB6, VB 12 0.01 mg, 30 mg of VB3, 30 mg of VB9, 900 mg of VB4, and 0.1 mg of VB7.

[0059] Comparative Example

[0060] The difference between this comparative example and Example 1 is that quercetagetin (QG) is omitted, and the other components are the same as those in Example 1 to prepare a basal diet.

[0061] Experimental Example 1

[0062] A total of 460 Yila multiparous female rabbits with similar body weights and parities were selected as the experimental subjects and randomly divided into 5 groups (92 rabbits in each group), namely:

[0063] (1) Control group (NC group): fed the basal diet prepared in the comparative example;

[0064] (2) QG100 group: fed the basal diet prepared in the comparative example + 100 mg / kg QG;

[0065] (3) QG200 group: fed the basal diet prepared in the comparative example + 200 mg / kg QG;

[0066] (4) QG400 group: fed the basal diet prepared in the comparative example + 400 mg / kg QG;

[0067] (5) QG600 group: fed the basal diet prepared in the comparative example + 600 mg / kg QG.

[0068] Starting from the day when the light in the rabbit house was increased (i.e., 6 days before breeding), the female rabbits in each experimental group were fed the corresponding feed from this time until the 35th day after parturition, with a total cycle of 71 days. The nutritional levels of the basal diet used are shown in Table 1.

[0069] Table 1 Nutritional levels of the basal diet for female rabbits (air-dried basis)

[0070]

[0071]

[0072] In this experiment, a 42-day breeding model was adopted, that is, the female rabbits were bred 12 days after parturition. On the day of breeding, artificial insemination technology was used to perform the breeding operation on the female rabbits; on the 15th day after breeding, a pregnancy check was carried out by palpation to determine whether the female rabbits were successfully pregnant and to count the conception rate; on the day of parturition of the female rabbits, various reproductive indexes were counted in detail, and then the relevant indexes such as the growth of the offspring rabbits were continuously recorded to provide detailed data support for subsequent research.

[0073] 1. Evaluation of heat stress status

[0074] (1) Measurement of temperature-humidity index

[0075] A temperature-humidity recorder was placed in the rabbit house to automatically collect the temperature and humidity data of the rabbit house. From the day of breeding to the end of parturition of the female rabbits, the data was recorded every 30 minutes during this period, and the temperature and humidity values at 14:00, 16:00, and 18:00 every day were selected. The temperature-humidity index (THI) was calculated according to the values of the dry-bulb temperature and relative humidity. The calculation formula is:

[0076] THI = db - [(0.31 - 0.31RH)(db - 14.4)];

[0077] Wherein, THI is the temperature - humidity index, %; db is the dry - bulb temperature, °C; RH is the relative humidity, %.

[0078] (2) Test results

[0079] The data of the temperature - humidity index (THI) are as Figure 1 shown. The threshold THI of the heat stress of rabbits is 27.8. According to the THI evaluation value, the degree of heat stress faced by rabbits is divided into the following categories:

[0080] When THI < 27.8, the rabbits are not in a heat - stress state;

[0081] When 27.8 ≤ THI < 28.9, it indicates that the rabbits are in a mild heat - stress state;

[0082] When 28.9 ≤ THI < 30.0, the rabbits are in a severe heat - stress state;

[0083] When THI ≥ 30.0, the rabbits are in an extreme heat - stress state.

[0084] It is assumed that if more than 50% of the time reaches the heat - stress state during the entire test period, it is determined that the test environment meets the test conditions of heat stress.

[0085] From Figure 2 it can be seen that during the entire test process, the time proportion of the female rabbits in the mild heat - stress state is 20%, the time proportion in the severe heat - stress state is 13.33%, and the time proportion in the extreme heat - stress state reaches 53.33%. It shows that the time proportion of the female rabbits suffering from different degrees of heat stress is as high as 86.66%, far exceeding 50%. Therefore, it can be clearly determined that the test conditions of this test have reached the heat - stress standard, laying a foundation for the subsequent research on the impact of heat stress on female rabbits.

[0086] 2. The effect of quercetagetin on the feed intake of female rabbits

[0087] (1) Recording of the feed intake of female rabbits

[0088] The feed intake of female rabbits was recorded separately in the early pregnancy stage (from the 0th day to the 10th day of pregnancy), the middle pregnancy stage (from the 11th day to the 20th day of pregnancy), the late pregnancy stage (from the 21st day to the 30th day of pregnancy), and the lactation period. The method of recording the feed intake of female rabbits is: add the weight of the test feed fed in the afternoon every day to the weight of the supplementary feed in the morning of the next day, and then calculate the difference from the remaining feed, which is the feed - intake data of the female rabbits on the same day.

[0089] (2) Test results

[0090] The feed intake data of female rabbits are shown in Table 2.

[0091] Table 2 Effects of adding quercetagitrin on the feed intake of female rabbits under heat stress

[0092]

[0093] Note: Different lowercase letters after the values in the same row in the table indicate significant differences, P < 0.05.

[0094] As can be seen from Table 2, among all the experimental groups, only the QG400 group and the QG600 group could significantly increase the feed intake of female rabbits during the entire pregnancy period (P < 0.05), showing obvious advantages compared with the control group and other experimental groups. The other two experimental groups (QG100 group and QG200 group) had no significant differences compared with the control group. In terms of the daily average feed intake during pregnancy, except for the QG100 group, the values of the other three experimental groups were significantly higher than those of the control group (P < 0.05); during the lactation period, adding 600 mg / kg of QG significantly increased the feed intake of female rabbits (P < 0.05).

[0095] In summary, under certain dosage conditions, QG has a positive effect on improving the feed intake of female rabbits. Especially the addition amounts of 400 mg / kg and 600 mg / kg can effectively promote the feeding of female rabbits at specific stages during the entire pregnancy period and lactation period, which is of great significance for ensuring the nutrient intake of female rabbits and improving reproductive performance.

[0096] 3. Effects of quercetagitrin on the reproductive performance of female rabbits

[0097] (1) Determination of the reproductive performance of female rabbits

[0098] On the first day of the pre-feeding period, light synchronization estrus treatment was carried out on the female rabbits participating in this experiment. After the female rabbits were in estrus, artificial insemination was used for breeding on the first day of the formal test period, and the number of female rabbits participating in breeding was accurately marked. On the 15th day after breeding, palpation was carried out to determine whether the female rabbits were pregnant, and the conception rates of each group were recorded. After the female rabbits gave birth, the litter-related situations of each female rabbit were recorded, including a series of key indicators such as the littering rate, comprehensive litter size, comprehensive number of live-born offspring, litter size, number of live-born offspring per litter, litter weight, weight of live-born offspring per litter, and number of dead fetuses. Among them, the conception rate refers to the proportion of female rabbits that are actually successfully pregnant after breeding, and the littering rate is the proportion of female rabbits that actually give birth after pregnancy. The comprehensive litter size refers to the product of the litter size and the littering rate, and the comprehensive number of live-born offspring refers to the product of the number of live-born offspring per litter and the littering rate. The specific calculation formulas are as follows:

[0099] A = (a / m) × 100%;

[0100] B = (b / m) × 100%;

[0101] C = c × B;

[0102] D = d × B.

[0103] Among them, A is the conception rate, %; a is the number of pregnant female rabbits, unit: only; m is the number of female rabbits mated, unit: only; B is the littering rate, %; b is the number of female rabbits giving birth, unit: only; C is the comprehensive litter size, unit: only; c is the litter size, unit: only; D is the comprehensive number of live-born offspring, unit: only; d is the number of live-born offspring per litter, unit: only.

[0104] (2) Test results

[0105] The data of the reproductive performance of female rabbits are shown in Table 3.

[0106] Table 3 Effects of adding quercetin marigold on the reproductive performance of female rabbits under heat stress during the whole pregnancy period

[0107]

[0108] Note: Different lowercase letters after the same-row values in the table indicate significant differences, P < 0.05; control group (basic diet), QG 100 group (basic diet + 100 mg / kg QG), QG200 group (basic diet + 200 mg / kg QG), QG400 group (basic diet + 400 mg / kg QG), QG600 group (basic diet + 600 mg / kg QG).

[0109] It can be seen from the data in Table 3 that the conception rate of female rabbits in the QG600 group was significantly improved compared with that of the control group (P<0.05), increasing by 15.03%, while the difference in the conception rate between the other dose experimental groups and the control group was not obvious. In terms of the litter size, there was no obvious difference between each experimental group and the control group. In terms of the number of live births per litter, the QG100 group and the QG600 group were significantly increased compared with the control group (P<0.05), increasing by 1.77 and 0.97 respectively. In terms of the number of live born offspring per litter, the QG100 group, the QG200 group and the QG600 group were significantly higher than the control group (P<0.05), increasing by 2.1, 0.4 and 1.23 respectively. In terms of the comprehensive number of live born offspring per litter, compared with the control group, the four experimental groups with different doses all showed significant differences (P<0.05), increasing by 0.53, 0.47, 0.6 and 1.63 respectively. In terms of the comprehensive number of live born offspring, all the experimental groups supplemented with quercetin marigoldin were higher than the control group (P<0.05), increasing by 0.73, 0.59, 0.63 and 1.74 respectively. In terms of the number of stillbirths, compared with the control group, the QG100 group showed a significant decrease (P<0.05), decreasing by 0.33, while the differences between the other three experimental groups and the control group were not significant. Compared with the control group, the QG100 group, the QG400 group and the QG600 group significantly increased the total weight of live born offspring per litter (P<0.05); all experimental groups significantly increased the weight of live born offspring per litter (P<0.05); the QG400 group and the QG600 group significantly increased the average weight of live born offspring (P<0.05).

[0110] In summary, under heat stress conditions, adding 600 mg / kg QG to the basic diet of female rabbits can effectively improve the conception rate of female rabbits (increased by 15.03%, P<0.05), significantly increase the number of live born offspring per litter (increased by 0.97), the number of live born offspring per litter (increased by 1.23), the comprehensive number of live born offspring per litter (increased by 1.63), the comprehensive number of live born offspring (increased by 1.74), as well as the total weight of live born offspring per litter (increased by 76.66 g), the weight of live born offspring per litter (increased by 81 g) and the average weight of live born offspring (increased by 2.5 g, P<0.05). Therefore, 600 mg / kg QG is the optimal addition amount to relieve heat stress and improve the reproductive performance of female rabbits.

[0111] 4. Effects of quercetin marigoldin on the growth of young rabbits

[0112] (1) Determination of the growth performance of young rabbits

[0113] After the birth of the young rabbits, in order to ensure that the growth conditions of each litter of young rabbits are consistent, the litter equalization operation should be carried out as early as possible to make the young rabbits in each litter balanced in size and number. Generally, the young rabbits start to grow hair about 5 days after birth and open their eyes around 12 days. After 18 days of birth, the birth box can be opened to allow the young rabbits to start eating feed. After more than ten days of feed adaptation period, the young rabbits have a certain ability to survive independently, so they are weaned at 35 days. During the growth process of the young rabbits, relevant data are recorded, including the number of young rabbits in each litter at 7 days, 14 days, 21 days and 35 days, and the litter weight is weighed when the young rabbits are fasting. Based on the recorded number of young rabbits in the litter and the weighed litter weight, the average weight of the young rabbits is further calculated.

[0114] (2) Test results

[0115] The data of the growth performance of the young rabbits are shown in Table 4.

[0116] Table 4 Effects of adding quercetin marigoldin during the whole pregnancy on the growth of young rabbits under heat stress

[0117]

[0118] Note: Different lowercase letters after the same row of values in the table indicate significant differences, P < 0.05.

[0119] As can be seen from Table 4, on the 7th day after the birth of the young rabbits, compared with the control group, the litter size of the QG200 group, QG400 group, and QG600 group showed a significant increase (P<0.05). In terms of the litter weight of the young rabbits, only the experimental groups with 400 mg / kg and 600 mg / kg QG added showed a level significantly higher than that of the control group (P<0.05), while there were no significant differences in the average weight of the young rabbits among the experimental groups. When the young rabbits grew to the 14th day, the average litter size of the four experimental groups was significantly higher than that of the control group (P<0.05). In terms of the litter weight index of the young rabbits, except for the QG100 group, the values of the other experimental groups were significantly higher than those of the control group (P<0.05). It is worth noting that in terms of the average weight of the young rabbits, only the experimental groups with 400 mg / kg and 600 mg / kg QG added showed an advantage significantly higher than that of the control group (P<0.05). By the 21st day after the birth of the young rabbits, compared with the control group, the average litter size of the four-dose experimental groups showed a significant increase (P<0.05). In terms of the litter weight of the young rabbits, except for the QG100 group, the values of the other experimental groups were significantly higher than those of the control group (P<0.05). In terms of the average weight index of the young rabbits, the four-dose experimental groups were all significantly higher than the control group (P<0.05), showing an obvious promoting effect. When the young rabbits were weaned at the 35th day of growth, compared with the control group, the litter size of the experimental groups with 200 mg / kg, 400 mg / kg, and 600 mg / kg QG added was significantly increased (P<0.05). In terms of the litter weight of the young rabbits, the four-dose experimental groups all showed a significant increase (P<0.05). In terms of the average weight of the young rabbits, the QG200 group, QG400 group, and QG600 group showed a significant increase compared with the control group (P<0.05).

[0120] In summary, adding 400 mg / kg and 600 mg / kg QG to the diet of female rabbits has a more significant effect on promoting the growth of young rabbits under heat stress conditions, can effectively improve multiple growth indexes of young rabbits, can effectively reduce the breeding cost, improve the market competitiveness, and provide strong support for the sustainable development of the rabbit farming industry.

[0121] Experimental Example 2

[0122] At 8:00 am on the 15th day of pregnancy in female rabbits, 6 female rabbits were randomly selected from each group, and blood samples were collected from the auricular veins of them. Then the blood was centrifuged at a speed of 2500 revolutions per minute for 10 minutes, and the serum obtained after centrifugation was collected and stored at -80 °C for detecting multiple key indicators, including progesterone (P), estradiol (E2), superoxide dismutase (SOD), total antioxidant capacity (T-AOC), glutathione peroxidase (GSH-Px), malondialdehyde (MDA), interleukin-6 (IL-6) and interleukin-10 (IL-10). The information of the manufacturers of the reagents used in the detection is shown in Table 5.

[0123] Table 5 Information of the manufacturers of main reagents

[0124] Serial Number Name Manufacturer 1 Progesterone (P) Detection Kit Northern Biotechnology Research Institute 2 Estradiol (E2) Detection Kit Northern Biotechnology Research Institute 3 Superoxide Dismutase (SOD) Assay Kit Nanjing Jiancheng Bioengineering Institute 4 Total Antioxidant Capacity (T-AOC) Detection Kit Nanjing Jiancheng Bioengineering Institute 5 Glutathione Peroxidase (GSH-Px) Assay Kit Nanjing Jiancheng Bioengineering Institute 6 Malondialdehyde (MDA) Detection Kit Nanjing Jiancheng Bioengineering Institute 7 Interleukin-6 (IL-6) Detection Kit Thermo Fisher Scientific 8 Interleukin-10 (IL-10) Detection Kit Thermo Fisher Scientific

[0125] 1. Effects of QG on serum sex hormones of female rabbits on the 15th day of pregnancy

[0126] It can be seen from Figure 3 that on the 15th day of pregnancy, female rabbits in the heat stress environment, compared with the control group, adding 100 mg / kg, 200 mg / kg, 400 mg / kg and 600 mg / kg QG to the basal diet could significantly increase the concentration of progesterone (P) in the serum of breeding female rabbits (P < 0.05), and the improvement effects of the doses of 100 mg / kg and 600 mg / kg were particularly significant.

[0127] As Figure 4 shown, in terms of the concentration of estradiol (E2), according to the serum detection data, there were no significant differences between the four experimental groups adding different doses of QG and the control group.

[0128] The above results indicate that supplementing QG in the diet of female rabbits has a positive effect on increasing the concentration of P in the serum of female rabbits, can effectively alleviate the negative effects of the reproductive endocrine system of female rabbits under heat stress conditions, and improve the reproductive performance of female rabbits.

[0129] 2. Effects of QG on serum oxidative stress indexes of female rabbits on the 15th day of pregnancy

[0130] As Figures 5 - 7As shown, through the analysis of serum sample data, it was found that compared with the control group, adding 200 mg / kg, 400 mg / kg, and 600 mg / kg QG to the basal diet could significantly increase the contents of superoxide dismutase (SOD), total antioxidant capacity (T-AOC), and glutathione peroxidase (GSH-Px) in the serum (P<0.05). Among them, compared with the control group, the experimental group adding 100 mg / kg QG could also significantly increase the contents of SOD and GSH-Px in the serum of female rabbits (P<0.05).

[0131] As Figure 8 shown, compared with the control group, the experimental groups adding 200 mg / kg, 400 mg / kg, and 600 mg / kg QG could all significantly reduce the content of MDA in the serum (P<0.05). Among the experimental groups, the QG400 group and the QG600 group were particularly prominent in improving the oxidative stress-related indicators.

[0132] The above results indicate that adding 600 mg / kg QG to the diet significantly increased the contents of superoxide dismutase (SOD), total antioxidant capacity (T-AOC) in the serum, and glutathione peroxidase (GSH-Px), while significantly reducing the content of malondialdehyde (MDA) in the serum. It shows that under heat stress conditions, QG plays an important antioxidant role in female rabbits. The mechanism of alleviating oxidative stress caused by heat stress is mainly through reducing the content of MDA in the serum of female rabbits and reducing the damage caused by lipid peroxidation reaction to the body; at the same time, enhancing the activity of antioxidant enzymes and improving the body's own ability to scavenge free radicals, so as to effectively maintain the balance between oxidation and antioxidant in the body and reduce the adverse effects of heat stress on female rabbits.

[0133] 3. Effects of QG on serum inflammatory indexes of female rabbits on the 15th day of pregnancy

[0134] As Figure 9 shown, compared with the control group, when adding 100 mg / kg, 200 mg / kg, and 400 mg / kg QG to the basal diet, the content of interleukin-6 (IL-6) in the serum of female rabbits decreased significantly (P<0.05). As Figure 10 shown, four different doses of QG could all significantly increase the content of interleukin-10 (IL-10) in the serum to a certain extent (P<0.05), and the experimental group adding 600 mg / kg QG was extremely prominent in increasing the content of serum IL-10, showing a highly significant difference (P<0.001). It shows that for female rabbits in a heat stress state, adding QG to the feed can effectively alleviate the adverse effects of heat stress on the inflammatory-related indexes in the body of female rabbits.

[0135] Experimental Example 3

[0136] In this experimental example, the control group (NC group) fed only the basal diet and the group with 600 mg / kg QG added to the basal diet (QG 600 group) were used as examples to study the effects of quercetagetin on the intestinal microbial community of female rabbits under heat stress from multiple aspects such as the number of OTUs, diversity index, and microbial community composition.

[0137] 1. Effects of QG on the number of OTUs in the intestinal microbiota of female rabbits

[0138] Figure 11 The OTUs shared by the control group (NC) and the QG600 group and the unique OTUs of each group are shown. The number of OTUs shared by the two groups is 1,571, the number of unique OTUs in the control group is 4,141, and the number of unique OTUs in the QG600 group is 3,809. The numbers of unique OTUs in the two groups are similar.

[0139] 2. Effects of QG on the α-diversity of the intestinal microbiota of female rabbits

[0140] α-diversity is used to reflect the richness and evenness of a community. The commonly used measurement indices mainly include the Chao 1 index that focuses on reflecting the richness of the community, and the Shannon index and Simpson index that take into account both the diversity and evenness of the community.

[0141] The effects of QG on the α-diversity of the intestinal microbiota of female rabbits are shown in Table 6. Figures 12 - 14 As shown, there were no significant differences in the Chao 1 index, Shannon index, and Simpson index between the two groups (P > 0.05). However, the Simpson index in the QG600 group was higher than that in the control group, indicating that adding 600 mg / kg QG to the diet can improve the diversity of the intestinal microbiota of female rabbits under heat stress.

[0142] Table 6 Effects of QG on the α-diversity of the intestinal microbiota of female rabbits under heat stress

[0143] Item Control Group QG600 Group P Value Chao 1 Index 1437.00±163.19 1273.83±284.61 0.25 Simpson Index 0.008±0.002 0.018±0.012 0.12 Shannon Index 5.88±0.13 5.57±0.34 0.09

[0144] 3. Effects of QG on the β-diversity of the intestinal microbiota of female rabbits

[0145] β-diversity is used to compare the similarity in species diversity among different samples. PCA is a technique for analyzing and simplifying a data set. By decomposing the variance and taking the two eigenvalues that can best reflect the variance as the coordinate axes, the differences among multiple groups of data are reflected on a two-dimensional coordinate graph. The closer the distance between two samples, the more similar the composition of the two samples.

[0146] PCA plot ( Figure 15) It was shown that the samples in the control group were significantly aggregated internally, presenting similar bacterial communities; while the samples in the QG600 group were dispersed internally, indicating certain differences among different individuals within the group. The addition of QG led to an increase in the differences in the intestinal microbiota composition among different individuals within the group. However, the samples of the control group and the QG600 group had a certain overlap and dispersion on the confidence ellipse, indicating that the addition of QG had an impact on the intestinal microbiota composition of heat-stressed female rabbits, but did not form a completely independent community structure.

[0147] 4. Effects of QG on the composition of the intestinal flora of female rabbits at the phylum level

[0148] Figure 16 The microbial community compositions of the control group and the QG600 group at the phylum level are shown. It can be seen that the microbial communities of both groups are composed of multiple phyla, and the dominant phyla are similar. The dominant phyla mainly include Firmicutes, Bacteroidota, Cyanobacteria, Verrucomicrobiota, Patescibacteria, and Proteobacteria. There are also certain proportions of other phyla, but the relative abundances are relatively low. Figures 17 - 22 The relative abundances of the top six dominant phyla (Firmicutes, Bacteroidota, Cyanobacteria, Verrucomicrobiota, Patescibacteria, and Proteobacteria) in the control group and the QG600 group were compared more intuitively.

[0149] 5. Effects of QG on the composition of the intestinal flora of female rabbits at the genus level

[0150] Figure 23 The compositions of the intestinal flora at the genus level of the control group and the QG600 group are presented. It can be seen that the samples of both groups are composed of multiple genera, and there are some dominant genera with relatively high abundances, such as unclassified_f_Muribaculaceae, unclassified_f_Lachnospiraceae, etc. However, the proportional distributions of different genera are different between the two groups, indicating that the microbial community structures of the two groups of samples are different ( Figures 24 - 29 ).

[0151] 6. Effects of QG on the intestinal microbiota composition of female rabbits

[0152] LEfSe (Linear discriminant analysis Effect Size) is the LDA Effect Size analysis, which is an analytical tool for discovering and interpreting biological markers (taxonomic units, pathways, genes) in high-dimensional data. It can compare two or more groups and also perform comparative analysis between subgroups within a group to find species with significant differences in abundance between groups (i.e., biomarkers).

[0153] From the LDA scores ( Figure 30 ), microorganisms with an LDA value greater than 2 were significantly enriched in the QG600 group. For example, microorganisms of the order Bacteroidales that were not classified to a lower level, such as g_unclassified_o_Bacteroidales, s_unclassified_o_Bacteroidales, and f_unclassified_o_Bacteroidales, had higher abundances in the QG600 group, indicating that the environment of this group might be more suitable for the survival of such microorganisms. g_Lysinibacillus (genus Lysinibacillus), its species s_unclassified_g_Lys inibacillus that was not classified to a species level, as well as s_unclassified_f_Acholeplasmataceae and g_unclassified_f_Acholeplasmataceae (unclassified microorganisms of the family Acholeplasmataceae) all showed a dominant position in the QG600 group, meaning that the treatment with 600 mg / kg QG promoted the growth or proliferation of these microorganisms.

[0154] Microorganisms with an LDA value less than 2 in the control group were significantly enriched in the control group. It can be seen that methanogenic microorganisms such as g__Eubacterium__n odatum_group (genus Eubacterium nodatum group), g_Methanosphaera (genus Methanosphaera), and its species s_Methanosp haera_cuniculi were dominant in the control group, indicating that heat stress conditions were more conducive to the growth of such microorganisms. In addition, s_unclassified_g_Christensenellaceae_R (unclassified genus R of the family Christensenellaceae), g_Sanguibacteroides (genus Sanguibacteroides), etc. were also significantly enriched in the control group, indicating that heat stress conditions promoted the proliferation of these microorganisms.

[0155] The phylogenetic tree shows different species and their evolutionary relationships. The concentric circles radiating from the inside out represent taxonomic levels from phylum to genus (or species). Each small circle at different taxonomic levels represents a taxon at that level, and the diameter of the small circle is proportional to the relative abundance. As Figure 31 shown, the distribution of microbial taxa in the two groups of samples can be more intuitively seen through the phylogenetic tree. The QG600 group and the control group showed different dominant microbial taxa on different phylogenetic branches, which not only reflected the differences in microbial composition between the two groups of samples but also implied their differences in ecological functions. For example, there were more Bacteroidales-related microorganisms in the QG600 group, which might play an important role in ecological processes such as material metabolism in this group; while the dominance of methanogenic microorganisms such as Methanosphaera in the control group might be closely related to the energy metabolism and gas production processes in this group. Combining with the LEfSe results, from an evolutionary perspective, the Bacteroidales-related microorganisms enriched in the QG600 group (such as g_unclassified_o_Bacteroidales, s_unclassified_o_Bacteroidales, etc.) had a certain evolutionary correlation, and they might share some characteristics adapted to the QG600 group environment. The methanogenesis-related microorganisms enriched in the control group (such as g_Methanosphaera, s_Methanosphaera_cuniculi) were also closely connected on the phylogenetic branch, indicating that they might have similar evolutionary strategies in adapting to the control group environment. This association between the evolutionary relationship and the differences in microbial abundances between groups helps to understand the formation mechanism of differences in microbial community structure under different conditions.

[0156] In summary, the addition of QG had various effects on the intestinal microbiota of heat-stressed female rabbits, including changes in the number of OTUs, improvement of α-diversity, differences in β-diversity, subtle adjustments in the composition at the phylum and genus levels, and significantly different microorganisms revealed by LEfSe analysis. These results provide important clues for further exploring the mechanism of action of QG on intestinal microbiota and its potential impact on the health of female rabbits.

Claims

1. A feed containing quercetagetin, characterized in that, The feed is prepared by mixing a basic diet and quercetin marigoldin in a mass ratio of 1 kg: 100 - 600 mg. The raw materials used in the basic diet include, by weight: 10 - 15 parts of corn, 3 - 7 parts of wheat middlings, 3 - 7 parts of corn bran, 5 - 10 parts of wheat bran, 0.5 - 2.5 parts of soybean oil, 10 - 20 parts of soybean meal, 3 - 9 parts of rapeseed meal mixture, 5 - 13 parts of distillers grains, 5 - 8 parts of corn germ meal, 0.5 - 2.5 parts of artemisia annua powder, 3 - 9 parts of rice hull powder, 3 - 7 parts of peanut shell powder, 5 - 10 parts of white rod powder, 5 - 10 parts of alkali grass, and 3 - 7 parts of premix.

2. The feed containing quercetagetin according to claim 1, wherein The feed is prepared by mixing a basic diet and quercetin marigoldin in a mass ratio of 1 kg: 600 mg.

3. The feed containing quercetagetin according to claim 1, wherein The raw materials used in the basic diet include, by weight: 12 parts of corn, 5 parts of wheat middlings, 5 parts of corn bran, 8.5 parts of wheat bran, 1.5 parts of soybean oil, 14 parts of soybean meal, 6 parts of rapeseed meal mixture, 9 parts of distillers grains, 6.5 parts of corn germ meal, 1.5 parts of artemisia annua powder, 6 parts of rice hull powder, 5 parts of peanut shell powder, 7 parts of white rod powder, 8 parts of alkali grass, and 5 parts of premix.

4. The feed containing quercetagetin according to claim 1, characterized in that, The raw materials used in the basic diet include, by weight: 10 parts of corn, 3 parts of wheat middlings, 3 parts of corn bran, 10 parts of wheat bran, 2.5 parts of soybean oil, 20 parts of soybean meal, 3 parts of rapeseed meal mixture, 5 parts of distillers grains, 5 parts of corn germ meal, 0.5 parts of artemisia annua powder, 3 parts of rice hull powder, 7 parts of peanut shell powder, 10 parts of white rod powder, 10 parts of alkali grass, and 7 parts of premix.

5. The feed containing quercetagetin according to claim 1, characterized in that, The raw materials used in the basic diet include, by weight: 15 parts of corn, 7 parts of wheat middlings, 7 parts of corn bran, 5 parts of wheat bran, 0.5 parts of soybean oil, 10 parts of soybean meal, 9 parts of rapeseed meal mixture, 13 parts of distillers grains, 8 parts of corn germ meal, 2.5 parts of artemisia annua powder, 9 parts of rice hull powder, 3 parts of peanut shell powder, 5 parts of white rod powder, 5 parts of alkali grass, and 3 parts of premix.

6. The feed containing quercetagetin according to claim 1, characterized in that, Per kilogram of the premix includes: Fe 50 - 100 mg, Cu 10 - 30 mg, Zn 50 - 100 mg, Mn 5 - 15 mg, Co 0.1 - 0.2 mg, I 0.1 - 0.3 mg, Se 0.2 - 0.3 mg, VA 9000 - 11000 IU, VD 800 - 1000 IU, VE 30 - 70 mg, VK 1 - 3 mg, VB1 1 - 3 mg, VB2 4 - 8 mg, VB5 30 - 70 mg, VB6 1 - 3 mg, VB 12 0.01 - 0.03 mg, VB3 30 - 70 mg, VB9 30 - 60 mg, VB4 900 - 1100 mg, and VB7 0.1 - 0.3 mg.

7. The feed containing quercetagetin according to claim 6, characterized in that, Per kilogram of the premix includes: 70 mg of Fe, 20 mg of Cu, 70 mg of Zn, 10 mg of Mn, 0.15 mg of Co, 0.2 mg of I, 0.25 mg of Se, 10000 IU of VA, 900 IU of VD, 50 mg of VE, 2 mg of VK, 2 mg of VB1, 6 mg of VB2, 50 mg of VB5, 2 mg of VB6, VB 12 0.02 mg, 50 mg of VB3, 44 mg of VB9, 1000 mg of VB4 and 0.2 mg of VB7.

8. The preparation method of the feed containing quercetagetin according to claim 1, characterized in that, It includes the following steps: Mix the raw materials evenly in proportion to obtain the feed containing quercetin marigoldin.

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