Application of red date extract containing cyclic adenosine monophosphate in preparation of food for improving premature ovarian failure

Abstract

The invention belongs to the technical field of medicinal and edible foods, and discloses an application of a red date extract containing cyclic adenosine monophosphate in preparation of a food for improving premature ovarian failure. The high-content cAMP in the red date extract is used as a core efficacy carrier, and animal experiments prove that the cAMP can accurately act on the pathologic link of premature ovarian failure. The limitation of single conditioning is broken through, and the dual effects of ovarian function improvement and qi and blood conditioning are achieved. The cAMP in the red date extract is combined with the nourishing effects of the Chinese wolfberry fruits, the rhizoma polygonati and the like, so that the premature ovarian failure can be improved in a targeted manner, and anemia can be effectively corrected. Through high-activity component targeting, multi-effect synergistic conditioning and safe process guarantee, side effects and dependency of western medicines are avoided, technical shortages of traditional products are made up, a safe, efficient and accurate conditioning scheme is provided for women with premature ovarian failure, clinical application value and industrial transformation potential are both achieved, and the pharmaceutical composition is worthy of popularization and application. The policy orientation of high-quality development of medicinal and edible functional foods is met.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of food with medicinal value, and particularly relates to application of a red date extract containing cyclic adenosine monophosphate in preparation of food for improving premature ovarian failure. BACKGROUND

[0002] With the rapid development of social economy, the lifestyle of residents has changed significantly, and bad habits such as staying up late, long-term mental stress, lack of exercise, high-fat diet, etc. are becoming increasingly common. Coupled with the widespread exposure to endocrine disruptors in the environment, the incidence of premature ovarian failure (POI) in women is on the rise year by year, and the age of onset is becoming younger and younger. According to the medical diagnostic criteria, premature ovarian failure refers to the occurrence of ovarian failure in women before the age of 40, which is manifested as persistent amenorrhea, persistent elevation of follicle-stimulating hormone (FSH) level (>40 IU / L), and significant reduction of estrogen level. The prevalence of premature ovarian failure in women of childbearing age has reached 1%~3%, which has become an important disease affecting the reproductive health and quality of life of women.

[0003] Premature ovarian failure not only directly leads to the disorder of six indicators of fertility (follicle-stimulating hormone, luteinizing hormone, estradiol, progesterone, testosterone, and prolactin) (the core manifestations are elevated gonadotropins and low estrogen and progesterone), but also induces a series of linked health risks: on the endocrine level, it leads to the premature occurrence of menopausal symptoms such as menstrual irregularity (cycle disorder, abnormal menstrual flow, and even amenorrhea), hot flashes, night sweats, and mood swings. Some patients also have thyroid function abnormalities, insulin resistance, and other complex endocrine disorders. On the skeletal system, estrogen deficiency can accelerate bone loss, significantly increasing the risk of osteoporosis and fractures. Five to ten years after the onset, the incidence of osteoporosis is 3~5 times higher than that of normal women. In addition, it is also closely related to cardiovascular diseases (such as hypertension and coronary heart disease), cognitive decline, skin aging, vaginal dryness, and other problems, which seriously affect the physical and mental health of women and their social adaptability.

[0004] At present, the intervention means for premature ovarian failure in clinical is mainly western medicine, the core is exogenous hormone replacement therapy (HRT), and the commonly used drugs include nearly dozens of estradiol valerate, 17β-estradiol, dydrogesterone, progesterone capsules, etc. The administration mode is mainly oral and external use (gel, patch). Although such chemical synthetic hormones can quickly improve hormone levels and relieve menopausal symptoms, they have obvious limitations: first, the side effects are clear, long-term use may cause gastrointestinal discomfort, breast pain, headache and dizziness, and may significantly increase the risk of serious diseases such as endometrial cancer, breast cancer and thrombosis; second, the essence of treatment is "replacement and supplementation" rather than "functional repair", which cannot reverse the functional decline of the ovary, and once the drug is stopped, the symptoms are easy to rebound quickly, and long-term dependence on exogenous hormones may further inhibit the hormone secretion function of the ovary, forming a vicious cycle; third, the applicable population is limited, and for patients with hormone-dependent tumor history, thrombosis history, and severe liver and kidney dysfunction, there are clear contraindications, and the clinical application scene is strictly limited.

[0006] However, the currently disclosed related technical solutions (such as the patent CN201010280287.X discloses a compound containing wolfberry, female privet fruit, prepared rehmannia, and many other Chinese medicines and peanuts, red dates, and soybeans) still have obvious shortcomings: first, the composition is complex, and many are simple compounding of more than 10 original medicinal and edible materials, without clear core effective components (such as phytoestrogens, polysaccharides, and flavonoids), resulting in unclear mechanism and difficult to achieve precise regulation; second, there is a lack of targeted extraction and enrichment process of effective components, and the raw materials are mostly used in the form of raw powder or crude extract, with low concentration of effective components and poor bioavailability, affecting the regulation effect; third, the formula design is mostly based on the traditional Chinese medicine theory of "tonifying kidney and essence", without optimization of the action target of modern pharmacology on ovarian function repair (such as antioxidant stress and inhibition of ovarian granulosa cell apoptosis), resulting in insufficient targeting and difficulty in product quality control, and the efficacy stability of different batches is difficult to guarantee.

[0007] Therefore, in view of the side effect risk of existing western medicine and the technical defects of medicinal and edible formula, it is an important research direction to develop an ovarian premature failure regulating product with clear components, advanced extraction process, strong targeting and safety without side effects, and to solve the pain points of women's reproductive health. SUMMARY

[0008] In view of the above technical problems, the purpose of the present application is to provide an application of a red date extract containing cyclic adenosine monophosphate in the preparation of food for improving premature ovarian failure. The food is mainly prepared from a red date extract containing cyclic adenosine monophosphate with a content of not less than 30 mg / 100 mL. In combination with the synergistic effect of homologous medicinal and edible components such as medlar and polygonatum, multi-dimensional targeted conditioning of premature ovarian failure is achieved. Not only is premature ovarian failure improved, but anemia is also effectively corrected. Zebrafish anemia experiments have confirmed that the hemoglobin content of the sample treatment group is significantly increased compared with the model control group (P<0.05), and the 1% concentration is the maximum safe concentration without toxic effects, which can improve various problems of low blood and gas. This multi-effect synergistic effect not only relieves symptoms such as irregular menstruation, hot flashes and night sweats caused by premature ovarian failure, but also reduces the risk of osteoporosis, cardiovascular disease and other complications through tonifying qi and blood, achieving comprehensive protection of female reproductive health and overall health.

[0009] In order to achieve the above purpose, the present application adopts the following technical solutions: An application of a red date extract containing cyclic adenosine monophosphate in the preparation of food for improving premature ovarian failure, wherein the cyclic adenosine monophosphate content of the red date extract is not less than 30 mg / 100 mL.

[0010] As a preferred, the food is used for regulating hormone level disorder and improving ovarian tissue pathological damage.

[0011] As a preferred, the regulation and improvement of hormone level disorder includes improving anemia and increasing hormone content. The improvement of ovarian tissue pathological damage includes increasing the number of oocytes and improving oocyte development.

[0012] As a preferred, the hormone includes anti-Mullerian hormone, estradiol hormone, prolactin, follicle-stimulating hormone, testosterone hormone, luteinizing hormone and progesterone.

[0013] A homologous medicinal and edible food for improving premature ovarian failure, comprising the red date extract, wherein the cyclic adenosine monophosphate content of the red date extract is not less than 30 mg / 100 mL.

[0014] As a preferred, the mass fraction of the red date extract containing cyclic adenosine monophosphate in the food is 95-96%.

[0015] As a preferred, the food further comprises 1-1.25% of medlar powder, 1-1.25% of polygonatum powder, 1-1.25% of lily powder and 1-1.25% of spina date seed powder.

[0016] A preparation method of a homologous medicinal and edible food for improving premature ovarian failure, comprising the following steps: S1, preparation of a red date extract: A, select the rich cyclic adenosine monophosphate, moisture content of 12~15%, no mildew, no insect damage, no impurities jujube varieties; B, with 25~30℃ warm water to wash jujube 2~3 times, remove the surface dust, sand and pesticide residues, after draining the surface moisture, jujube broken to 2~4mm particles and remove the jujube peel, jujube meat and other impurities immediately into the extraction process; The extraction process is: jujube and extraction liquid with solid-liquid ratio of 1kg:12~15L, under the constant temperature of 60~65℃, through constant temperature stirring countercurrent extraction and ultrasonic assisted multistage extraction, the obtained extraction liquid is mixed, filtered through 100~200 mesh filter screen, remove jujube meat, jujube peel, 0.3~0.5MPa pressure filtration, obtain clear primary filtrate; The constant temperature stirring countercurrent extraction process is: stirring rate is 50~60r / min, countercurrent level is 3, ultrasonic power is 180~200W, ultrasonic frequency is 30~40kHz, ultrasonic 10min every 20~30min; The multistage extraction includes first stage extraction 50~60min; second stage extraction 30~40min; third stage extraction 20~30min; C, the primary filtrate is filtered through ceramic membrane with pore size of 50~80nm, pressure of 0.2~0.3MPa, temperature of 30~35℃, filtration flow rate of 10~15L / h m 2 To obtain the refined filtrate; The refined filtrate is passed through macroporous adsorption resin at a flow rate of 2BV / h (bed volume / hour), and the pH value of the sample solution is controlled at 5.5~6.5; then the resin column is first washed with 3BV purified water to remove unabsorbed sugars, inorganic salts and other impurities; then eluted with 40~50% ethanol solution at a flow rate of 1BV / h, and the eluate is collected; D, the eluate is concentrated to 1 / 6~1 / 8 of the original eluate volume through a nanofiltration membrane with a molecular weight cut-off of 200~300Da at a pressure of 1.0~1.2MPa and a temperature of 30~35℃, then subjected to double-effect concentration treatment at 50~55℃, and finally filtered through a 0.22μm polyether sulfone filter membrane and sterilized to obtain the jujube extract; S2, according to the corresponding mass fraction, select wolfberry powder, polygonatum sibiricum powder, lily powder and sour jujube kernel powder, add purified water at 78~80℃, the mass volume ratio of mixed powder and purified water is 1:5~1:8, shear for 10~20min with high-speed shear emulsifier, and prepare uniform powder slurry; Mix the jujube extract and the powder slurry at a volume ratio of 9:1, stir at a speed of 60~80r / min for 25~30min, then homogenize 2~3 times at a pressure of 20~25MPa and a temperature of 50~60℃, filter through a 0.22μm polyether sulfone filter membrane and sterilize to obtain the food with medicinal and edible properties.

[0017] As preferably, the red jujube category includes any one of grey jujube, long jujube and Jun jujube.

[0018] As preferably, in S1, the extraction solution is purified water and vitamin C mixed at a mass ratio of 1:0.001.

[0019] Compared with the prior art, the present application has at least the following technical effects: The application provides application of a red jujube extract containing cyclic adenosine monophosphate in preparation of food for improving premature ovarian failure, which takes a red jujube extract containing cyclic adenosine monophosphate (cAMP >= 30 mg / 100 mL) as a core, matches Chinese wolfberry and rhizoma polygonati and other homologous medicinal and edible components, and realizes precise regulation and multiple effects of improving premature ovarian failure through an advanced extraction and compounding process, and the technical effect is remarkable and the advantage is outstanding.

[0020] The food is mainly prepared from a red jujube extract containing cyclic adenosine monophosphate with a content of not less than 30 mg / 100 mL, and has the following advantages: 1) Targeted repair of ovarian function, and significant improvement of core indicators The present application takes a high-content cAMP in a red jujube extract as a core efficacy carrier, and proves through animal experiments that the red jujube extract can precisely act on pathological links of premature ovarian failure. In a cyclophosphamide-induced POF model of mice, the high-dose and low-dose groups of the red jujube extract can significantly improve the level of anti-Mullerian hormone (AMH), effectively protect the preantral follicle and small antral follicle of the ovary, reduce follicle atresia, and improve the ovarian reserve function from the root; meanwhile, the red jujube extract can reduce abnormally elevated follicle-stimulating hormone (FSH) and luteinizing hormone (LH), make the hypothalamic-pituitary-ovarian axis (HPO axis) function tend to be normal, and correct the hormone disorder state. The ovum microscopic examination result shows that the number of ovum of the mice in the administration groups is significantly increased compared with the model group, the 2-cell ratio is improved (the high-dose group reaches 91%), the ovum development tends to be normal, the fertility potential is effectively improved, and the core pain point of infertility caused by premature ovarian failure is solved.

[0021] 2) Multiple effects of synergistic regulation, and consideration of qi and blood and overall health The present application breaks through the limitation of single regulation, realizes the dual effects of improvement of ovarian function and regulation of qi and blood. The cAMP in the red jujube extract, in combination with the nourishing effect of Chinese wolfberry, rhizoma polygonati and other components, not only improves premature ovarian failure, but also effectively corrects anemia. Zebrafish anemia experiments prove that the hemoglobin content of the sample treatment group is significantly increased compared with the model control group (P < 0.01). pIt contains less than 0.05%, and 1% is the maximum safe concentration, with no toxic effects. It can improve various problems caused by low Qi and blood. This multi-effect synergistic effect not only alleviates symptoms such as menstrual irregularities, hot flashes and night sweats caused by premature ovarian failure, but also reduces the risk of complications such as osteoporosis and cardiovascular disease by replenishing Qi and blood, thus achieving comprehensive protection for women's reproductive health and overall health.

[0022] III. Safe with no side effects and widely applicable. Compared to traditional Western medicine-based exogenous hormone replacement therapy (HRT), this invention adheres to the principle of food and medicine sharing the same origin, with a formula composed of natural raw materials and no added chemically synthesized hormones. In animal experiments, mice in all groups treated with jujube extract showed no abnormal toxic side effects, and their mental state and weight gain were superior to the model group. Long-term use did not pose any risks such as gastrointestinal discomfort or increased tumor risk. Furthermore, because it does not contain exogenous hormones, it is also suitable for individuals with a history of hormone-dependent tumors, thrombosis, or liver and kidney dysfunction—conditions contraindicated by Western medicine—significantly broadening its applicability and meeting the requirements for food safety and universality.

[0023] IV. Highly enriched active ingredients, stable and controllable process. This invention achieves efficient retention of core components through optimized extraction processes: After the jujubes are crushed, multi-stage filtration removes impurities such as the skin and pulp, enriching the effective components. A combination of low-temperature filtration extraction and multi-stage enrichment ensures a stable cAMP content of ≥30mg / 100mL, significantly higher than ordinary jujube products. The compounding process employs 80℃ warm water pre-dispersion followed by high-speed mixing to prevent powder agglomeration and improve bioavailability. A standardized quality control system is established throughout the entire process, from raw material selection (high-cAMP jujube varieties) to finished product testing, ensuring stable efficacy between batches. This addresses the technical shortcomings of traditional food-medicine homology formulas, such as complex ingredients, low concentrations of effective components, and large quality fluctuations.

[0024] In summary, this invention, through "targeted action of highly active ingredients + synergistic conditioning with multiple effects + safe process assurance," avoids the side effects and dependence of Western medicine while making up for the technical shortcomings of traditional products. It provides women with premature ovarian failure with a safe, efficient, and precise conditioning solution, which has both clinical application value and industrial transformation potential, and is in line with the policy guidance for the high-quality development of functional foods that are both food and medicine. Attached Figure Description

[0025] Figure 1 This is a schematic diagram showing the results of estradiol (E2) hormone concentration detection. Figure 2 This is a schematic diagram showing the results of follicle-stimulating hormone (FSH) concentration testing. Figure 3 This is a schematic diagram showing the results of prolactin (PRL) hormone concentration testing. Figure 4The figure is a result of the detection of testosterone (T) hormone concentration; Figure 5 The figure is a result of the detection of luteinizing hormone (LH) hormone concentration; Figure 6 The figure is a result of the detection of anti-Mullerian hormone (MIS AMH) hormone concentration; Figure 7 The figure is a result of the detection of progesterone (PROG) hormone concentration; Figure 8 The figure is a result of the detection of ovum under a microscope, and the magnification is 10x; Figure 9 The figure is a result of the detection of ovary HE staining; Figure 10 The figure is a result of the detection of hemoglobin content; Figure 11 The figure is a result of the detection of zebrafish blood cell staining phenotype, and the magnification is 80x. DETAILED DESCRIPTION

[0026] In order to more clearly illustrate the technical solutions of the embodiments of the present application or the prior art, the present application will be briefly introduced below in combination with the drawings and the description of the embodiments or the prior art. Obviously, the following description of the structure of the drawings is only some embodiments of the present application, and other drawings can be obtained by those skilled in the art without creating labor. It should be noted that the description of these embodiments is used to help understand the present application, but does not constitute a limitation on the present application.

[0027] The technical solution of one specific embodiment of the present application is: Preparation of jujube extract: A. Select jujube species rich in cyclic adenosine monophosphate, with water content of 12-15%, no mildew, no insect damage, no impurities; B. Wash the jujube with 30℃ warm water for 3 times to remove surface dust, sand and pesticide residues, and then drain the surface water. Immediately after crushing the jujube to 4mm particles, it is put into the extraction process; The extraction process is: jujube and extraction liquid are mixed at a solid-liquid ratio of 1kg:15L, and multi-stage extraction is carried out by constant temperature stirring countercurrent extraction and ultrasonic assistance at a constant temperature of 65℃. The obtained extraction liquid is mixed, filtered through a 200 mesh sieve, and the jujube meat and jujube skin are removed. The clear primary filtrate is obtained by 0.5MPa pressure filtration; The constant temperature stirring countercurrent extraction process is: the stirring rate is 60r / min, the countercurrent level is 3, the ultrasonic power is 200W, the ultrasonic frequency is 40kHz, and the ultrasonic time is 10min every 30min; The multi-stage extraction includes first-stage extraction for 50min, second-stage extraction for 30min, and third-stage extraction for 20min; C. The initial filtrate is filtered through a membrane with a pore size of 80 nm, a pressure of 0.3 MPa, a temperature of 35 °C, and a flow rate of 15 L / h. m 2 The fine filtrate was obtained after filtration through a ceramic membrane; The fine filtrate was passed through the macroporous adsorption resin at a flow rate of 2 BV / h (bed volume / hour), and the pH of the loading solution was controlled at 5.5~6.5. Then, the resin column was rinsed with 3 BV of purified water to remove unadsorbed sugars, inorganic salts and other impurities. Then, 50% ethanol solution was used to elute at a flow rate of 1 BV / h, and the eluent was collected. D. The eluent is passed through a 200-300 Da nanofiltration membrane and concentrated to 1 / 6 to 1 / 8 of the original eluent volume at a pressure of 1.2 MPa and a temperature of 35 °C. Then, it is subjected to double concentration at 55 °C, filtered through a 0.22 μm polyethersulfone membrane, and sterilized to obtain the jujube extract.

[0028] The jujube extract containing cyclic adenosine monophosphate (cAMP) can be used to prepare food products that improve premature ovarian failure, including: improving various types of anemia, blood deficiency, premature ovarian failure, reversing AMH levels, and improving six fertility indicators.

[0029] Principle: The content of cyclic adenosine monophosphate (cAMP), a special component in jujubes, is thousands or even tens of thousands of times higher than that in ordinary foods, making it the richest source of cAMP in nature. Jujubes are among the most abundant foods in nature in terms of cAMP content. Traditional Chinese medicine has long recorded that jujubes can replenish qi and blood, regulate menstruation, and have excellent effects on maintaining youthful appearance. Modern medical and nutritional research has found that these effects of jujubes mainly rely on cAMP, which is known as the body's second messenger. Therefore, jujube extract is used as the main raw material, supplemented by other medicinal and edible substances such as wolfberry, polygonatum, lily powder, and jujube seed, to improve qi and blood, correct various types of anemia, improve premature ovarian failure, and improve the levels of six fertility hormones.

[0030] Dosage form: can be tablets, granules, syrups, oral liquids, powders, etc.

[0031] Dosage: After preparation, the daily adult dosage is 20mL~80mL, which can be divided into 1~2 doses; based on the single bottle capacity of 10mL~40mL, take 1~2 bottles each time.

[0032] Experimental Verification 1: The following experiments were all completed and reported by Feifan Standard Technical Service (Suzhou) Co., Ltd. 1. Experimental Principle: Cyclophosphamide is a commonly used chemotherapy drug. Its active metabolites can damage rapidly proliferating cells (such as granulosa cells and oocytes in the ovary), inducing apoptosis and oxidative stress, thus mimicking the pathological state of premature ovarian failure in humans, manifested as estrous cycle disorders, decreased serum sex hormone levels, and reduced follicle count. Jujube extract contains active ingredients and potential mechanisms of action. Jujubes are rich in polysaccharides, flavonoids, vitamin C, and cyclic adenosine monophosphate (cAMP), which have strong antioxidant capabilities. These components can scavenge excess reactive oxygen species generated by cyclophosphamide, reducing oxidative stress damage to ovarian granulosa cells. Jujube extract may inhibit excessive apoptosis of granulosa cells by regulating apoptosis-related signaling pathways (such as Bcl-2 / Bax and Caspase-3), thereby maintaining normal follicle development and survival. Some active components in jujubes may improve ovarian endocrine function by affecting the function of the hypothalamus-pituitary-ovarian axis, promoting the normal secretion of follicle-stimulating hormone (FSH) and luteinizing hormone (LH), thereby regulating estradiol and progesterone levels. Traditional Chinese medicine believes that jujubes have the effects of "tonifying the middle energizer, replenishing qi, nourishing blood, and calming the mind." Modern research suggests that they may provide nutrition and support to damaged ovaries by improving local microcirculation and cellular energy metabolism in the ovaries. Pre-administration of jujube extract before modeling can reduce the damage of cyclophosphamide to ovarian tissue in mice through its antioxidant and anti-apoptotic effects, improve their reproductive endocrine function, and delay or improve the progression of premature ovarian failure.

[0033] 2. Experimental instruments and reagents

[0034] 3. Experimental Methods 3.1 Healthy female SPF-grade C57 mice (e.g., 8 weeks old) were selected and acclimatized for one week. They were randomly divided into 4 groups (n=15 per group): Blank control group: The same volume of solvent was administered by gavage and the same volume of physiological saline was injected intraperitoneally.

[0035] Model group: administered an equal volume of solvent by gavage and intraperitoneal injection of cyclophosphamide (50 mg / kg for several consecutive days); Low-dose jujube extract group: low-dose jujube extract was administered by gavage (200 μL / animal / day), and cyclophosphamide was injected intraperitoneally (same as the model group). High-dose jujube extract group: High-dose jujube extract was administered by gavage (400 μL / animal / day). Cyclophosphamide was injected intraperitoneally (same as the model group).

[0036] Except for the blank control group, the other groups were given intraperitoneal injections of cyclophosphamide to establish POF models as planned.

[0037] The jujube extract was administered via gavage to each dosage group starting one week before modeling and continuing until one week after modeling (approximately 2 weeks in total). The control group and the model group were simultaneously administered an equal volume of solvent via gavage.

[0038] 3.2 Sample Collection and Indicator Detection Serum collection and hormone detection: Blood was collected from the eyeballs of 3 birds in each group after administration on days 7 and 14, and the serum was separated by centrifugation. AMH and six estrogen levels were detected using an ELISA kit.

[0039] Ovarian tissue collection: Three mice in each group were sacrificed on day 14, and bilateral ovaries were quickly removed, photographed, and weighed (to calculate the ovarian index).

[0040] The ovaries were fixed with 4% paraformaldehyde, routinely embedded in paraffin, sectioned, and stained with hematoxylin and eosin (HE).

[0041] On day 14, ovaries of 9 birds in each group were collected, oocytes were extracted and cultured in vitro, and oocyte development was observed.

[0042] 3.3 Statistical Analysis All data are expressed as mean ± standard deviation. One-way ANOVA was performed using SPSS or GraphPad Prism software, and LSD-t test was used for comparisons between groups. P < 0.05 was considered statistically significant.

[0043] 4. Experimental Results 4.1 Effects on the general condition and body weight of mice: The model group mice exhibited lethargy, dull fur, reduced activity, and slow or decreased weight gain. These symptoms were significantly improved in all dosage groups of jujube extract, especially the high-dose group.

[0044] This experiment successfully replicated a cyclophosphamide-induced mouse model of premature ovarian failure, characterized by hormonal imbalances and pathological damage to ovarian tissue.

[0045] 4.2 Testing of various hormone indicators 4.2.1 Detection of estradiol (E2) hormone concentration, results are as follows: Figure 1 As shown.

[0046] like Figure 1 The diagram shows the results of estradiol (E2) hormone concentration detection.

[0047] Estradiol (E2) is mainly secreted by developing follicles and is a core hormone for assessing ovarian function and the menstrual cycle.

[0048] The results of estradiol (E2) testing were contrary to expectations: the E2 level in the model group did not decrease significantly, and the E2 level in the jujube extract treatment group was not significantly higher than that in the model group.

[0049] 4.2.2 Detection of follicle-stimulating hormone (FSH) concentration, results are as follows: Figure 2 As shown.

[0050] like Figure 2 The diagram shows the results of follicle-stimulating hormone (FSH) concentration testing.

[0051] Follicle-stimulating hormone (FSH) is secreted by the pituitary gland and is used to "encourage" the ovaries to develop follicles. When ovarian function fails, the ovaries respond less to FSH and secrete more FSH.

[0052] Model group: Elevated FSH levels. This is a diagnostic indicator of premature ovarian failure.

[0053] The FSH level in the jujube extract treatment group was lower than that in the model group. The decrease in FSH level is another extremely strong piece of evidence. It indicates that as ovarian function recovers (follicles can develop normally and secrete E2 and inhibin), the pituitary gland receives a negative feedback signal that "the signal has been received and the task has been completed," thereby reducing excessive FSH secretion.

[0054] This reflects an improvement in the function of the hypothalamus-pituitary-ovarian axis (HPO axis).

[0055] 4.2.3 Prolactin (PRL) hormone concentration detection, results are as follows Figure 3 As shown; 4.2.4 Testosterone (T) concentration detection, results are as follows Figure 4 As shown.

[0056] like Figure 3 The diagram shows the results of prolactin (PRL) hormone concentration detection.

[0057] like Figure 4 The diagram shows the results of testosterone (T) hormone concentration detection.

[0058] The changes in prolactin (PRL) and testosterone (T) in premature ovarian failure are not as typical as those of the aforementioned indicators. Hyperprolactinemia itself can suppress ovarian function, so the results are as expected.

[0059] In women, testosterone is primarily secreted by ovarian stromal cells and the adrenal glands, and its changes are relatively minor in this model. The results were not significantly different in this study.

[0060] 4.2.5 Luteinizing hormone (LH) concentration detection, results are as follows: Figure 5 As shown.

[0061] like Figure 5 The diagram shows the results of luteinizing hormone (LH) concentration detection.

[0062] Luteinizing hormone (LH) works synergistically with FSH to trigger ovulation and corpus luteum formation.

[0063] Model group: LH levels increased.

[0064] The LH levels in the jujube extract treatment group tended to normalize. The normalization of the LH / FSH ratio is also a reference for assessing the recovery of HPO axis function.

[0065] 4.2.6 Detection of anti-Müllerian hormone (MIS AMH) concentration, results are as follows: Figure 6 As shown.

[0066] like Figure 6 The diagram shows the results of anti-Müllerian hormone (MIS AMH) concentration detection.

[0067] Anti-Müllerian hormone (AMH) is secreted by the granulosa cells of preantral and small antral follicles in the ovary and is the most direct and sensitive indicator for assessing ovarian reserve (i.e., the number of remaining follicles in the follicular pool).

[0068] Results analysis: Model group: AMH levels were lower than those in the normal control group. This indicates successful modeling, a sharp decrease in the number of usable follicles in the ovary, and a decline in ovarian reserve.

[0069] The jujube extract treatment group showed higher AMH levels than the model group, and possibly approached the levels of the normal control group. This is one of the key pieces of evidence proving the effectiveness of the jujube extract.

[0070] The rebound in AMH levels strongly suggests that the jujube extract may have protected existing follicles and inhibited excessive follicular atresia. It promoted the development of preantral and small antral follicles, increasing the number of secretory follicles and fundamentally improving ovarian reserve.

[0071] 4.2.7 Progesterone (PROG) hormone concentration detection, results as follows Figure 7 As shown.

[0072] like Figure 7 The image shows a schematic diagram of the progesterone (PROG) hormone concentration test results.

[0073] Progesterone (P) is mainly secreted by the corpus luteum that forms after ovulation.

[0074] Expected Outcome Analysis: If treatment promotes follicle development and induces ovulation, then the corpus luteum can form, and progesterone levels may rise. In the POF model, progesterone levels are typically very low.

[0075] If jujube extract can increase progesterone levels, it indicates that its effect is not only on follicle growth, but may also promote the complete ovulation process and corpus luteum function.

[0076] The progesterone (PROG) test results did not meet expectations: there was no statistically significant difference in progesterone levels between the model group and the jujube extract treatment group, and the expected upward trend in the treatment group was not observed.

[0077] 4.3 Microscopic images of oocytes

[0078] like Figure 8 The image shown is a schematic diagram of oocyte microscopic examination, magnified to 10×.

[0079] Results combined Figure 8 Analysis of oocyte microscopic images revealed that the model group mice had fewer oocytes and a lower 2-cell ratio, indicating abnormal oocyte development. In contrast, the drug-treated group mice had more oocytes and an increased 2-cell ratio, with development gradually returning to normal. The above research results indicate that jujube extract can effectively improve ovarian function in a mouse model of premature ovarian failure.

[0080] 4.4 Ovarian HE like Figure 9 The image shown is a schematic diagram of HE staining of the ovary. The three smaller images in each group of figures represent stained sections (80×) that were validated in three parallel experiments within that group.

[0081] Blank control group: normal ovarian structure, with intact follicles at all stages and no pathological damage.

[0082] Model group: exhibited pathology of premature ovarian failure, with a sharp decrease in follicles and abnormal structure.

[0083] Low-dose jujube extract group: partial repair of ovarian damage, increased number of follicles and improved structure.

[0084] High-dose jujube extract group: ovarian damage was significantly repaired, and the structure and follicles were nearly normal.

[0085] In summary, cyclophosphamide successfully induced a premature ovarian failure model, and jujube extract was able to repair ovarian pathological damage in a dose-dependent manner. The high-dose group showed the most significant repair effect, restoring normal ovarian structure and follicular development, thus confirming its effectiveness in improving premature ovarian failure.

[0086] Experimental Verification 2: Experiment on Improving Anemia 1. Detection principle: Anemia is a decrease in the number of red blood cells in the peripheral blood. Zebrafish develop blood circulation 24 hours after fertilization, and blood cells can be directly observed under a microscope. The hematopoietic process is conserved throughout vertebrate evolution, and due to the transparent nature of zebrafish, it can be clearly observed. The positive control drug is Shengxuening tablets. 2. Materials and Methods 2.1 Experimental Instruments and Reagents Fluorescence microscope (SZX7, Olympus, Japan), o-anisidine (Ron Reagent, Shanghai), Shengxuening tablets (Wuhan United Pharmaceutical Co., Ltd.). 2.2 Detection system and sample size Experimental system: wild AB strain zebrafish Zebrafish age: 3 days after fertilization Sample size per group: 30 samples per group Adult fish rearing and breeding methods: Follow our company's standard rearing and breeding methods. 3. Experimental Procedure The zebrafish were divided into four groups: a blank control group, a model control group, a positive control group, and a sample treatment group. The normal control group received no treatment, the model control group received phenylhydrazine, the positive control group received Shengxuening tablets, and the sample treatment group received both the sample and phenylhydrazine. After a period of time, tissue was extracted from the zebrafish and stained.

[0087] II. Applicability and Limitations For testing the blood-tonifying efficacy of health products, food and their raw materials, the sample must be soluble in water or prepared into a uniformly dispersed suspension in water. III. Data Processing Methods SPSS 26 software was used for analysis, and GraphPad Prism 8 was used for plotting. Data statistics are expressed as mean ± standard deviation. Experimental results were tested for normality using SPSS software. If the data conformed to normality, the significance was determined by the T-parameter test or analysis of variance (ANOVA). If the data did not conform to normality, nonparametric tests were used (P-value < 0.05 indicates statistical significance). IV. Judgment Criteria Based on the premise that the experiment meets the validity requirements, compared with the blank control group, the hemoglobin content in the model control group was significantly reduced ( p <0.05) indicates that the model was successfully created. Compared with the model control group, the hemoglobin content in the sample treatment group was significantly increased ( p <0.05), indicating that the sample has blood-tonifying effects. V. Preliminary Experimental Test Results There was no significant difference between the 1% concentration sample group and the blank control group, therefore 1% concentration is the maximum safe concentration.

[0088] VI. Test Results

[0089] Result data: Zebrafish hemoglobin staining area (pixels)

[0090] like Figure 10 The image shows a bar chart of hemoglobin content.

[0091] like Figure 11 The image shown is a zebrafish blood cell staining phenotype, magnified 80×.

[0092] Results combined Figure 10 It can be seen that the model group had low hemoglobin levels, while the sample group had significantly higher levels than the model group.

[0093] Combination Figure 11 It can be seen that the blood cells in the model group are sparse, while the number of blood cells in the sample group is significantly increased.

[0094] This experiment proved that phenylhydrazine successfully constructed an anemia model, and that jujube extract could significantly increase hemoglobin and blood cell count, demonstrating a clear blood-tonifying effect.

[0095] Finally, it should be noted that the above description is merely a preferred embodiment of the present invention and is not intended to limit the scope of protection of the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. Application of a cyclic adenosine monophosphate-containing red date extract in the preparation of a food for improving premature ovarian failure, characterized in that, The cyclic adenosine monophosphate content in the red jujube extract is not less than 30 mg / 100 mL. 2.The application of the A-AMP-containing red date extract in the preparation of food for improving premature ovarian failure according to claim 1, characterized in that, The food is used for adjusting hormone level disorder and improving ovary tissue pathological damage. 3.The application of the A-5G in the preparation of food for improving premature ovarian failure according to claim 2, characterized in that, The adjusting and improving hormone level disorder includes improving anemia and increasing hormone content. The improving ovary tissue pathological damage includes increasing the number of ova and improving ova development.

4. The use of the adenosine cyclic phosphate-containing red date extract according to claim 3 in the preparation of food for improving premature ovarian failure, characterized in that, The hormone includes anti-Mullerian hormone, estradiol hormone, prolactin, follicle-stimulating hormone, testosterone hormone, luteinizing hormone and progesterone.

5. A functional food for improving premature ovarian failure, characterized by comprising the composition according to claim 1. The food contains the red jujube extract with cyclic adenosine monophosphate content not less than 30 mg / 100 mL.

6. The pharmafood of claim 5, wherein the pharmafood is a composition for improving premature ovarian failure. The mass fraction of the red jujube extract containing cyclic adenosine monophosphate in the food is 95-96%.

7. The phicochemical food of claim 6, wherein the phicochemical food is a food or a food ingredient for improving premature ovarian failure. The food further contains wolfberry powder 1-1.25%, rhizoma polygonati powder 1-1.25%, lily powder 1-1.25% and semen ziziphi spinozae powder 1-1.25%.

8. A process for preparing the functional food for improving premature ovarian failure according to any one of claims 5 to 7, which is characterized by, The method comprises the following steps: S1, preparation of red jujube extract: A, select red jujube with rich cyclic adenosine monophosphate, water content of 12-15%, no mildew, no insect damage, no impurities; B, rinse the red jujube with 25-30℃ warm water for 2-3 times to remove surface dust, sand and pesticide residues, then drain the surface water, crush the red jujube to 2-4mm particles, remove the jujube peel, jujube meat and other impurities, and immediately put into the extraction process; The extraction process is: the red jujube and the extraction liquid are extracted by constant temperature stirring countercurrent extraction and ultrasonic assistance at a solid-liquid ratio of 1kg:12-15L and a constant temperature of 60-65℃, the obtained extraction liquids are mixed, filtered through a 100-200 mesh filter screen to remove jujube meat and jujube peel, and then filtered under a pressure of 0.3-0.5MPa to obtain clear primary filtrate; The constant temperature stirring countercurrent extraction process is: the stirring speed is 50-60r / min, the countercurrent level is 3, the ultrasonic power is 180-200W, the ultrasonic frequency is 30-40kHz, and the ultrasonic time is 10min every 20-30min; The multi-stage extraction includes first-stage extraction for 50-60min, second-stage extraction for 30-40min and third-stage extraction for 20-30min; C. The primary filtrate is filtered through a ceramic membrane with a pore size of 50-80 nm, a pressure of 0.2-0.3 MPa, a temperature of 30-35℃, and a filtration flow rate of 10-15 L / h m 2 to obtain a fine filtrate; The filtrate is passed through a macroporous adsorption resin at a flow rate of 2BV / h (bed volume / hour), and the pH value of the sample solution is controlled at 5.5-6.5; then the resin column is washed with 3BV purified water to remove unabsorbed sugars, inorganic salts and other impurities; then 40-50% ethanol solution is used to elute at a flow rate of 1BV / h, and the eluate is collected; D, the eluate is concentrated to 1 / 6-1 / 8 of the original eluate volume through a nanofiltration membrane with a molecular weight cut-off of 200-300Da under a pressure of 1.0-1.2MPa and a temperature of 30-35℃, then subjected to double-effect concentration treatment at 50-55℃, filtered through a 0.22μm polyether sulfone filter membrane, and sterilized to obtain the red jujube extract; S2, select wolfberry powder, rhizoma polygonati powder, lily powder and semen ziziphi spinozae powder according to the corresponding mass fraction, mix them, add purified water at 78-80℃, and mix the powder and the purified water at a mass to volume ratio of 1:5-1:8, then shear for 10-20min with a high-speed shear emulsifier to prepare a uniform powder slurry. The red jujube extract and the powder slurry are mixed at a volume ratio of 9:1, stirred at a speed of 60-80 r / min for 25-30 min, homogenized 2-3 times at a pressure of 20-25 MPa and a temperature of 50-60 DEG C, filtered through a 0.22 mu m polyether sulfone filter membrane, sterilized, and thus a food with medicinal and edible effects is obtained.

9. A method for preparing a food-medicine homology product for improving premature ovarian failure according to claim 8, characterized in that, The red jujube category includes any one of grey jujube, long jujube and Jun jujube.

10. A method for preparing a food-medicine homology product for improving premature ovarian failure according to claim 8, characterized in that, In the S1, the extraction liquid is a mixture of purified water and vitamin C at a mass ratio of 1:0.001.

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