A method of improving reproductive performance

Abstract

This invention belongs to the field of agriculture and animal husbandry, specifically relating to a method for improving the reproductive performance of mammals under stress conditions. This invention discovers that tryptophan improves fetal weight and placental weight in mammals under heat stress conditions. By adding tryptophan to the basal diet, this invention effectively alleviates the adverse effects of heat stress on mice, improving their reproductive performance, promoting fetal and placental weight, and thus enhancing the economic benefits of animal husbandry. Tryptophan is readily available, easy to process, and suitable for large-scale production. Furthermore, it is highly safe as an additive, can be directly added to the feed of pregnant mammals, is convenient to operate, does not require injection or implantation, and avoids other stress responses in mammals.

Description

Technical Field

[0001] This invention belongs to the field of agriculture and animal husbandry, and specifically relates to a method for improving the reproductive performance of mammals under stress conditions. Background Technology

[0002] Stress is a biological response of an animal to stimuli that disrupt its physiological balance or homeostasis. With global warming, heat stress has become a key environmental factor restricting the sustainable development of animal husbandry. Heat stress refers to the body's inability to maintain a balance between heat production and heat dissipation; it is the result of the combined effects of various factors such as high temperature, high humidity, heat radiation, and air velocity, with high temperature having a particularly significant effect. Heat stress can lead to disorders in the secretion of key reproductive hormones (such as GnRH, LH, and FSH), impairing ovarian function, follicle development, endometrial receptivity, and placental blood supply, ultimately causing a series of serious reproductive damages such as embryo implantation failure, developmental delays, increased abortion rates, and decreased reproductive performance.

[0003] Currently, common heat stress mitigation measures in farms and laboratories involve physical cooling to alleviate the adverse effects of heat stress, such as preventing solar radiation, improving ventilation, using sprinkler fans, evaporative cooling pads, air coolers, and roof sprinkler systems. While lowering the ambient temperature is the most effective way to alleviate heat stress, its effectiveness in actual production is limited or prohibitively expensive, it is ineffective in humid areas, and it cannot resolve existing physiological cascade damage.

[0004] Currently, common additives for combating heat stress include electrolytes, vitamins, sedatives, antibiotics, organic acids, and traditional Chinese medicine powders. For example, adding vitamin C and vitamin E to feed can reduce heat stress-induced oxidative damage by scavenging reactive oxygen species (ROS). The technical problem it solves is alleviating systemic oxidative stress, but it lacks specific protection for the reproductive system (such as the embryo and placenta), resulting in limited effectiveness. Furthermore, sedatives and antibiotics can produce toxic side effects and drug residues. Adding plant extracts, such as those containing flavonoids and polyphenols, utilizes their antioxidant and anti-inflammatory properties. The technical problem it solves is providing exogenous antioxidants, but it faces challenges such as complex composition, difficulty in standardization, high cost, and potential impact on palatability.

[0005] Tryptophan (Trp) is the most widely distributed indole derivative in nature (Comai S, Bertazzo A, Brughera M, et al. Tryptophan in health and disease[J]. Advances in clinical chemistry, 2020, 95: 165-218.). Its chemical name is α-amino-β-indolepropionic acid, and its structural formula is shown in Figure 1. Tryptophan includes three isomers: L-form, D-form, and racemic DL-form. L-tryptophan is an essential amino acid for monogastric animals (such as humans, pigs, dogs, mice, and chickens) and pre-weaned ruminants (such as calves and lambs), and its endogenous synthesis in vivo cannot occur. In addition to participating in protein synthesis as a substrate, it can be metabolized through various pathways to produce biologically important compounds. D-Tryptophan is present in very small amounts in animals and undergoes almost no metabolic activity (Li Jianxin, Zhang Xumei, Xu Qishou. Physiological and biochemical effects and applications of tryptophan [J]. Amino Acids and Biological Resources, 2005, (03): 58-62.). Tryptophan and its metabolites play important roles in health and various diseases.

[0006] In reality, building a factory involves large investments and high risks, and the prices of some antioxidants remain high, making them unsuitable for widespread industrial application. Therefore, finding an additive that effectively improves reproductive performance and is suitable for large-scale production is a key, green, environmentally friendly, and safe method to address the damage caused by heat stress. Summary of the Invention

[0007] This invention discovers that tryptophan improves the reproductive performance of mammals under heat stress, specifically fetal weight and placental weight. Based on this discovery, this invention was completed.

[0008] In a first aspect, the present invention provides the use of tryptophan in the preparation of additives that improve the reproductive performance of mammals.

[0009] Furthermore, the mammals include mice, rats, pigs, cattle, sheep, horses, deer, foxes, mink, and rabbits.

[0010] Furthermore, the reproductive performance includes fetal weight and placental weight.

[0011] Furthermore, the mammal in question is a mammal subjected to stress.

[0012] Furthermore, the stress condition is heat stress.

[0013] Furthermore, the amount of tryptophan added is selected from 0.295-0.43%.

[0014] Furthermore, the amount of tryptophan added is 0.295%.

[0015] In one embodiment of the present invention, tryptophan is added directly to the diet of pregnant mammals, which is convenient to operate and does not require injection or implantation, thus avoiding stress response in animals.

[0016] In a second aspect, the present invention provides the use of tryptophan in the preparation of medicaments that improve the reproductive performance of mammals.

[0017] Furthermore, the mammals include mice, rats, pigs, cattle, sheep, horses, deer, foxes, mink, and rabbits.

[0018] Furthermore, the reproductive performance includes fetal weight and placental weight.

[0019] Furthermore, the mammal in question is a mammal subjected to stress.

[0020] Furthermore, the stress condition is heat stress.

[0021] Furthermore, the amount of tryptophan added is selected from 0.295-0.43%.

[0022] Furthermore, the amount of tryptophan added is 0.295%.

[0023] Thirdly, the present invention provides a method for improving the reproductive performance of mammals, the method comprising adding tryptophan to the diet of pregnant mammals.

[0024] Furthermore, the mammals include mice, rats, pigs, cattle, sheep, horses, deer, foxes, mink, and rabbits.

[0025] Furthermore, the reproductive performance includes fetal weight and placental weight.

[0026] Furthermore, the mammal in question is a mammal subjected to stress.

[0027] Furthermore, the stress condition is heat stress.

[0028] Furthermore, the amount of tryptophan added is selected from 0.295-0.43%.

[0029] Furthermore, the amount of tryptophan added is 0.295%.

[0030] In one embodiment of the present invention, tryptophan is added directly to the diet of pregnant mammals, which is convenient to operate and does not require injection or implantation, thus avoiding stress response in animals.

[0031] Beneficial effects This invention effectively alleviates the adverse effects of heat stress on mice by adding tryptophan to the basic feed, thereby improving the reproductive performance of mice, promoting fetal weight and placental weight, and enhancing the economic benefits of animal husbandry.

[0032] Tryptophan is easy to obtain and simple to process, making it suitable for large-scale production. Furthermore, it is highly safe as an additive, easily added directly to the feed of pregnant mammals without the need for injection or implantation, thus avoiding other stress responses in the mammals. Attached Figure Description

[0033] Figure 1 This is a statistical result of the food intake of breeding female mice under heat stress conditions.

[0034] Figure 2 This is a schematic diagram of obtaining a fetus through dissection.

[0035] Figure 3 A schematic diagram of placental dissection. Detailed Implementation

[0036] The specific embodiments of the present invention will be further described below. It should be noted that these descriptions are for the purpose of aiding understanding the present invention, but do not constitute a limitation thereof. Furthermore, the technical features involved in the embodiments described below can be combined with each other as long as they do not conflict with each other.

[0037] Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods, and the experimental materials used in the following embodiments are all available through conventional commercial channels.

[0038] ICR Mice: In 1954, the ICR closed colony mouse was introduced to the Roswell Park Cancer Institute by Dr. Hauschka. Originating in 1947 from the Swiss mouse population, the ICR mouse was bred by Dr. Hauschka at the Philadelphia Cancer Institute. The ICR mouse population was selected for its high reproductive rate, and this strain is one of the most prolific closed colony mouse breeds. Dr. Hauschka used the Swiss mouse population for selective breeding with a focus on high productivity. The ICR mice are preserved at the Institute for Cancer Research and distributed to various countries, hence the abbreviation ICR. ICR mouse strain characteristics: Albino fur, docile, highly adaptable, robust, highly reproductive, fast-growing, with good experimental reproducibility, and a low incidence of spontaneous tumors.

[0039] Example 1: Test Samples and Materials mice Forty male mice and eighty female mice were housed together; after 0.5 days, 70 female mice were confirmed to have successfully mated.

[0040] Healthy 8-week-old female ICR (Institute of Cancer Research) mice, weighing approximately 32g on average, were selected and randomly divided into 5 groups; all female mice complied with all relevant laws and regulations; the groups were as follows: Control group (fed at room temperature, with 0.25% tryptophan in their basal diet and free access to water); Heat stress group (HS group, heat stress, basal diet containing 0.25% tryptophan, free access to water); Heat stress + 0.295% tryptophan intervention group (HS + 0.295% Trp group, the basal feed contains 0.25% tryptophan, and 0.045% tryptophan solution is supplemented through drinking water); Heat stress + 0.34% tryptophan intervention group (HS + 0.34% Trp group, the basal feed contains 0.25% tryptophan, and 0.09% tryptophan solution is supplemented through drinking water). Heat stress + 0.43% tryptophan intervention group (HS + 0.43% Trp group, the basal feed contains 0.25% tryptophan, and 0.18% tryptophan solution is supplemented through drinking water).

[0041] Feeding time: Feed with a solution containing tryptophan throughout the day. To ensure the activity of tryptophan, prepare a fresh solution and replace it daily.

[0042] cycle Record the heat stress observed when rats are caged together and tethered; continue the experiment for 13 days.

[0043] Test instruments and equipment Table 1 shows the main experimental instruments and equipment; Table 2 shows the main reagents and consumables.

[0044] Table 1 Main Test Instruments and Equipment Table 2 Main Reagents and Experimental Consumables Basic feed formulation The basic feed diet in this application is a commonly used formula in the industry. The nutritional levels of the basic feed formula are shown in Table 3. The basic feed ingredients include corn, soybean meal, fish meal, flour, wheat bran, sodium sulfate, calcium ammonium phosphate, limestone powder, various vitamins, various trace elements and amino acids, etc.

[0045] Table 3 Nutritional levels of basal feed formulations Reproductive performance The day after the animals were placed in the same cage, tethered mice were identified. A heat stress test was conducted on the tethered mice, and samples were taken at day 12.5 (embryonic day 12.5). Placental weight, fetal weight, number of live fetuses, and number of stillborn fetuses were recorded.

[0046] Test conditions All embodiments of this invention were conducted under heat stress conditions. A constant-temperature biological incubator was used, with the temperature set at 38.5 degrees Celsius for 2 hours, to induce heat stress in female mice (Wu JJ, Zheng X, Wu C, Ma W, Wang Y, Wang J, Wei Y, Zeng X, Zhang S, Guan W, Chen F. Melatonin alleviates high temperatureexposure induced fetal growth restriction via the gut-placenta-fetus axis inpregnant mice. J Adv Res. 2025 Feb;68:131-146. doi: 10.1016 / j.jare.2024.02.014. Epub 2024 Feb 20. PMID: 38382594; PMCID: PMC11785557.).

[0047] Example 2: Effect of tryptophan on feed intake in breeding female rats under heat stress conditions like Figure 1 The table shows the statistical results of food intake in breeding female mice under heat stress. Compared with the control group, there were no significant differences in food intake during gestation (E0.5-E12.5) among the experimental groups (HS, 0.295%Trp, 0.34%Trp, 0.43%Trp) (P>0.05); there were also no significant differences in the average daily food intake during gestation between the control group and the experimental groups (P>0.05).

[0048] Example 3: Effects of tryptophan on reproductive performance of female rats under heat stress. A. Test methods 1. Cage closing operation At 18:00 on day E0, male mice were placed in the female mice's cages for pairing. Pairing was conducted at a male-to-female ratio of 1:2. A total of 40 male mice and 80 female mice were used for pairing.

[0049] 2. Mating inspection At 9:00 AM on day E0.5, mating was checked on the female mice, a process known as "plug inspection." If a milky-white, firm solid was found at the vaginal opening of the female mouse, it was considered a vaginal plug, confirming successful mating. A total of 70 female mice were confirmed to have successfully mated.

[0050] 3. Dissection and Sample Collection Preparation: Prepare dissection instruments (scissors, forceps), precision electronic balance, petri dishes, physiological saline, filter paper, etc.

[0051] Sacrifice and fixation: Pregnant mice were euthanized using methods that comply with animal ethics, such as cervical dislocation, and then fixed in a supine position on a dissection board.

[0052] Open the abdominal cavity: At 12.5 days, disinfect the abdominal skin with 75% alcohol, cut along the midline of the abdomen to expose the uterus.

[0053] Uterine separation: Separate the surrounding tissues, completely remove both uterine horns, and place them in a culture dish lined with moistened filter paper.

[0054] 4. Separation of fetus and placenta The uterine wall is cut along the opposite side of the mesentery, and the intact fetus and placenta are removed one by one. The amniotic sac is carefully dissected with forceps, the umbilical cord is cut, and the fetus is separated from the placenta. The fetus is gently rinsed with saline solution to remove blood, and the surface liquid is blotted dry with filter paper.

[0055] 5. Indicator Measurement Weigh the placenta: Weigh the dried placenta and record the data.

[0056] Weigh the fetus: Weigh the individual fetal mice after they have been dried and record the data.

[0057] Record the number of live births and stillbirths.

[0058] B. Reproductive performance testing of female mice like Figure 2 The image shows a schematic diagram of the fetus obtained through dissection, with fetal weight measured and results recorded. Figure 3The diagram shows a schematic of the placenta obtained through dissection. Placental weight was measured and the results were recorded (Table 4). As shown in Table 4, compared with the control group (0.25% Trp), the fetal weight and placental weight of the heat stress group (0.25% Trp) were significantly lower (P<0.05). The fetal weight and placental weight of the 0.295% Trp group were the highest, significantly higher than all other groups (P<0.05). The fetal weight of the 0.34% Trp group and the 0.43% Trp group were significantly higher than the heat stress group, but significantly lower than the control group (P<0.05). The placental weight of the 0.34% Trp group and the 0.43% Trp group was significantly higher than the heat stress group (P<0.05), but there was no significant difference compared with the control group (P>0.05). There were no significant differences among the groups in terms of the number of live fetuses and stillbirths (P>0.05). In conclusion, under the conditions of this experiment, the addition of 0.295% tryptophan to the diet had a significant effect on the weight of the fetus and placenta in response to heat stress.

[0059] Table 4 Reproductive performance results of female mice

Claims

1. Use of tryptophan in the preparation of additives to improve the reproductive performance of mammals.

2. The use as described in claim 1, wherein the mammal is a mammal under stress; the mammal includes mice, rats, pigs, cattle, sheep, horses, deer, foxes, mink, and rabbits; the reproductive performance includes fetal weight and placental weight; and the amount of tryptophan added is selected from 0.295-0.43%.

3. The use as described in claim 2, wherein the stress condition is heat stress; and the amount of tryptophan added is 0.295%.

4. Use of tryptophan in the preparation of drugs that improve the reproductive performance of mammals.

5. The use as described in claim 4, wherein the mammal is a mammal under stress; the mammal includes mice, rats, pigs, cattle, sheep, horses, deer, foxes, mink, and rabbits; the reproductive performance includes fetal weight and placental weight; and the amount of tryptophan added is selected from 0.295-0.43%.

6. The use as described in claim 5, wherein the stress condition is heat stress; and the amount of tryptophan added is 0.295%.

7. A method for improving the reproductive performance of mammals, the method comprising adding tryptophan to the diet of pregnant mammals.

8. The method of claim 7, wherein the mammal is a mammal under stress; the mammal includes mice, rats, pigs, cattle, sheep, horses, deer, foxes, mink, and rabbits; the reproductive performance includes fetal weight and placental weight; and the amount of tryptophan added is selected from 0.295-0.43%.

9. The method of claim 8, wherein the stress condition is heat stress; and the amount of tryptophan added is 0.295%.

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