Application of qi-tonifying and blood-nourishing formula in preparation of medicine for preventing and treating premenstrual syndrome

Abstract

The invention provides application of a qi-tonifying and blood-nourishing formula in preparation of a medicine for preventing and treating premenstrual syndrome, and relates to the technical field of medicines. In the application, the active ingredients of the formula for tonifying qi and maintaining blood comprise pig blood extract, astragalus membranaceus and Chinese dates. The medicine for preventing and treating the premenstrual syndrome is prepared from the pig blood extract, the astragalus membranaceus and the Chinese dates, and the compatibility advantages of reinforcing qi and nourishing blood are achieved. According to the composition, the defects that traditional hormones and psychotropic drugs are single in target spot and large in side effect are overcome, by cooperatively regulating and controlling a blood coagulation-inflammation-neuroendocrine network, neurotransmitters can be up-regulated, emotion abnormalities such as depression and anxiety can be improved, the blood hypercoagulable state can be reduced, microcirculation can be improved, and multi-dimensional and multi-system safe comprehensive intervention is achieved.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, and more specifically, to the application of a Qi-nourishing and blood-nourishing formula in the preparation of a drug for the prevention and treatment of premenstrual syndrome. Background Technology

[0002] Premenstrual syndrome (PMS) refers to a complex of symptoms affecting a woman's mental, behavioral, and physical state that recur during the luteal phase of her menstrual cycle. These symptoms usually subside or disappear spontaneously with the start or end of the next menstrual cycle. PMS is a relatively common condition among women of reproductive age, with epidemiological data from both domestic and international sources indicating a high incidence. In some patients, physical discomfort and emotional symptoms such as depression and anxiety can severely impact their normal life and work. Therefore, the prevention and effective intervention of PMS have received widespread clinical attention.

[0003] The exact etiology and pathogenesis of premenstrual syndrome (PMS) are not fully understood. It is generally believed that an imbalance of endocrine hormones, such as estrogen and progesterone, during the luteal phase may be a significant contributing factor. Current clinical interventions for this condition are relatively limited, primarily relying on hormone-regulating drugs and psychotropic medications. For example, conventional methods often involve regulating the menstrual cycle using progesterone or synthetic progestins, or combining this with oral contraceptives. Furthermore, for patients experiencing significant mood swings, selective serotonin reuptake inhibitors are commonly used for symptomatic treatment; in rare severe cases, potent regulators such as gonadotropin-releasing hormone agonists may be considered.

[0004] However, the aforementioned existing intervention methods have many limitations and shortcomings in practical clinical applications. On the one hand, although hormonal drugs such as progestins or contraceptives can regulate hormone levels to some extent, their exact efficacy is highly controversial in many randomized controlled studies, and long-term exogenous hormone intervention can easily disrupt the body's original endocrine and physiological homeostasis, triggering a series of adverse reactions. On the other hand, although psychotropic drugs are somewhat effective in relieving some symptoms of depression and anxiety, their targets are relatively singular, often accompanied by corresponding tolerance problems or side effects, and they cannot improve the complex physical and physiological microcirculatory abnormalities of patients with premenstrual syndrome as a whole.

[0005] In summary, due to the complex pathological changes involving multiple systems such as the nervous and endocrine systems in premenstrual syndrome (PMS), existing single-target drugs or hormone replacement therapies are insufficient for comprehensive and multi-dimensional regulation, and their long-term safety and efficacy stability are inadequate. Clinically, the limited selection of drugs and unsatisfactory comprehensive intervention effects remain challenges, necessitating the search for a drug formulation that can safely, systematically, and through multiple pathways improve the various complex symptoms of PMS.

[0006] In view of this, the present invention is hereby proposed. Summary of the Invention

[0007] The purpose of this invention is to provide an application of a Qi-nourishing and blood-enriching formula in the preparation of a drug for the prevention and treatment of premenstrual syndrome. The application achieves multi-dimensional and safe regulation of the coagulation-inflammation-neuroendocrine network of premenstrual syndrome through the synergistic effect of pig blood extract, astragalus, and jujube, effectively improving emotional abnormalities and microcirculatory disorders, and overcoming the shortcomings of existing single-target drugs with large side effects.

[0008] In order to achieve the above-mentioned objectives of the present invention, the following technical solution is adopted: This invention provides the application of a Qi-nourishing and Blood-nourishing formula in the preparation of a drug for the prevention and treatment of premenstrual syndrome. The active ingredients of the Qi-nourishing and Blood-nourishing formula include pig blood extract, astragalus, and jujube.

[0009] In an optional embodiment, the pig blood extract includes heme iron; and / or, The dosage form of the drug is an oral preparation; preferably, the oral preparation includes at least one of tablets, capsules, granules, pills, powders, decoctions, oral liquids, and syrups.

[0010] In an optional embodiment, the equivalent dose of the drug is 150 mg / kg to 750 mg / kg.

[0011] In an optional implementation, the drug is used to improve depressive-like behaviors, anxiety-like behaviors, or anhedonia symptoms caused by the premenstrual syndrome.

[0012] In an optional implementation, the drug treats or prevents premenstrual syndrome by modulating the coagulation-inflammation-neuroendocrine axis and / or by improving systemic oxidative stress and neuroinflammation and regulating brain-gut axis homeostasis.

[0013] In optional embodiments, the regulation of the coagulation-inflammation-neuroendocrine axis includes at least one of the following pharmacological effects: A. Upregulating the levels of central neurotransmitters and neurotrophic factors, wherein the central neurotransmitters include serotonin and norepinephrine, and the neurotrophic factors include brain-derived neurotrophic factor; B. Upregulating the levels of sex hormones, wherein the sex hormones include estradiol and progesterone; C. Improving blood circulation and reducing hypercoagulable states and blood stasis tendencies; D. Improving systemic oxidative stress and neuroinflammation.

[0014] In an optional embodiment, the improvement in blood circulation is manifested as an improvement in at least one of the following pharmacological indicators: A. prolongation of at least one of prothrombin time, thrombin time, and activated partial thromboplastin time; B. reduction of fibrinogen levels; C. reduction of at least one of whole blood viscosity, plasma viscosity, hematocrit, and erythrocyte sedimentation rate.

[0015] In an optional implementation, the regulation of the coagulation-inflammation-neuroendocrine axis further includes improving immune-inflammatory markers, specifically manifested as an improvement in at least one of the following pharmacological markers: A. Decreases serum interleukin-6; B. Upregulates the percentage of monocytes in the blood; C. Decreases serum malondialdehyde levels; D. Upregulates serum superoxide dismutase levels and / or the ratio of reduced glutathione to oxidized glutathione.

[0016] In an optional implementation, the drug is used to upregulate the subject's mean red blood cell volume and / or mean red blood cell hemoglobin concentration.

[0017] In an optional embodiment, the drug is used to upregulate at least one of the following indicators: A, serum F9 levels in the target subject; B, mRNA expression levels of PLCD3 in hippocampal and uterine tissues; C, protein expression levels of CAMK2G and SNCAIP in hippocampal tissues; D, mRNA expression levels of NFE2L2, HMOX1, and SOD1 in the hippocampal tissues of the target subject.

[0018] Compared with existing technologies, this invention uses active ingredients containing pig blood extract, astragalus, and jujube to prepare a medicine for the prevention and treatment of premenstrual syndrome, fully leveraging the advantages of the combination of medicinal and edible ingredients in "tonifying qi and nourishing blood, harmonizing the liver and spleen." Pig blood extract is rich in highly bioavailable heme iron, which can quickly nourish yin and blood; astragalus and jujube complement each other, both tonifying qi and raising yang to promote the absorption of nutrients, and nourishing blood and calming the mind. The synergistic effect of the three ingredients corrects the pathological state of qi imbalance and bodily environment disorder in patients with premenstrual syndrome from a macroscopic and holistic perspective of qi and blood interaction, effectively overcoming the shortcomings of traditional exogenous hormone replacement therapy, which easily disrupts the body's endocrine homeostasis and causes adverse reactions.

[0019] This drug formulation effectively overcomes the limitations of existing clinical interventions with their single mechanism of action, enabling comprehensive, multi-dimensional, and multi-systemic intervention for premenstrual syndrome (PMS). At the neuroendocrine level, this combination of active ingredients significantly upregulates the levels of specific neurotransmitters and neurotrophic factors in the central nervous system and restores normal sex hormone secretion, thereby effectively improving severe emotional and behavioral abnormalities such as depression, anxiety, and anhedonia caused by PMS. At the physiological microcirculation level, this combination effectively reduces the expression of specific coagulation factors, prolongs clotting time, and reduces blood viscosity, improving the hypercoagulable state and inflammatory tendency caused by abnormal endocrine fluctuations, achieving multi-target synergistic regulation of the coagulation-inflammation-neuroendocrine network.

[0020] Furthermore, the drug prepared in this invention can also inhibit hippocampal NOX4-NFE2L2 pathway-mediated neuroinflammation and oxidative stress by regulating the "brain-gut axis" homeostasis. Through the synergistic effect of multiple components in the compound, this drug can effectively regulate the intestinal microecology, reduce the generation of harmful metabolites such as p-cresol and their accumulation in the brain as p-cresol sulfate (PCS), thereby alleviating neuronal oxidative stress damage from the source (specifically manifested as a significant downregulation of serum malondialdehyde (MDA) levels and an upregulation of superoxide dismutase (SOD) and reduced glutathione (GSH) levels). Simultaneously, this drug can significantly upregulate the mRNA expression of core antioxidant regulatory factors and target genes (such as NFE2L2, HMOX1, and SOD1) in hippocampal tissue. This mechanism forms a positive feedback loop of neuroprotection in the subject's body, effectively reducing the release of core pro-inflammatory factors such as interleukin-1β, further consolidating the improvement effect on emotional and behavioral abnormalities such as depression and anxiety.

[0021] This approach, utilizing a combination of specific Chinese herbal extracts to prepare drugs for the prevention and treatment of premenstrual syndrome (PMS), not only demonstrates comprehensive efficacy comparable to conventional targeted chemotherapy drugs, but also fundamentally avoids the tolerance issues and side effects associated with long-term use of psychotropic drugs due to the natural and mild nature of its active ingredients. Its multi-pathway, systemic intervention characteristics provide a novel and definitively effective solution to the clinical lack of safe, effective, and comprehensive PMS treatments that can improve complex symptoms. Attached Figure Description

[0022] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0023] Figure 1This is a representative movement trajectory diagram of the rat open field experiment in Example 2 of this application; Figure 2 This is a bar chart showing the statistical results of various indicators (total movement distance, activity time, number of standing times, number of times the grid is threaded, time spent in the central area, and time spent in the four sides) of the rat open field experiment in Example 2 of this application; Figure 3 This is a bar chart showing the statistical results of the behavioral effects of open field experiments on rats in Example 5 of this application, used for mechanism verification. Figure 4 This is a bar chart showing the statistical results of the percentage of rats' preference for sucrose water in Example 2 of this application; Figure 5 This is a bar chart showing the statistical results of the number of marbles buried in the rat marble burial experiment in Example 2 of this application; Figure 6 This is a bar chart showing the statistical results of the static time of the rat forced swimming experiment in Example 2 of this application; Figure 7 This is a bar chart showing the statistical results of the forced swimming and sucrose preference experiments of rats used for mechanism verification in Example 2 of this application (A is the resting time, B is the percentage of sucrose preference). Figure 8 This is a bar chart showing the statistical results of the interaction time in the three-box social experiment of rats in Example 2 of this application; Figure 9 This is a bar chart showing the statistical results of four coagulation indicators (FIB, APTT, PT, TT) in rats in Example 3 of this application; Figure 10 This is a bar chart showing the statistical results of low / medium / high shear viscosity, plasma viscosity, and low / medium shear reduced viscosity of whole blood in rats in Example 3 of this application; Figure 11 This is a bar chart showing the statistical results of high-shear reduced viscosity, hematocrit, erythrocyte aggregation / electrophoresis / deformation index, and erythrocyte sedimentation rate of whole blood in rats in Example 3 of this application; Figure 12 This is a bar chart showing the statistical results of relevant indicators of rat blood routine in Example 3 of this application; Figure 13 This is a bar chart showing the statistical results of interleukin-6 (IL-6) levels in the serum of rats in each experimental group in Example 3 of this application; Figure 14 This is a bar chart showing the statistical results of serum sex hormone (estradiol E2, progesterone PROG) levels in rats in Example 4 of this application; Figure 15 This is a bar chart showing the statistical results of the levels of monoamine neurotransmitters (5-HT, NE) in the rat hippocampus in Example 4 of this application; Figure 16This is a bar chart showing the statistical results of brain-derived neurotrophic factor (BDNF) levels in rat hippocampal tissue in Example 4 of this application. Figure 17 This is a bar chart showing the statistical results of the levels of neurobiochemical indicators (5-HT, NE, BDNF) in the rat hippocampal tissue of the batch used for mechanism verification in Example 4 of this application (C is 5-HT, D is NE, E is BDNF). Figure 18 This is a principal component analysis (PCA) diagram of the rat hippocampal transcriptome in Example 5 of this application; Figure 19 This is a heatmap of differential gene expression clustering in the transcriptome of rat hippocampus tissue in Example 5 of this application; Figure 20 This is a volcano plot showing the differential transcriptome analysis of rat hippocampal tissue in Example 5 of this application; Figure 21 This is a bubble chart showing the enrichment analysis of differentially expressed genes in the transcriptome of rat hippocampus tissue in Example 5 of this application. Figure 22 This is a heatmap showing the correlation between gene expression of key target sites in the core regulatory network and multidimensional pharmacodynamic indicators in Example 5 of this application. Figure 23 These are immunohistochemical images of the core targets (CAMK2G, SNCAIP) in Example 5 of this application; where A is the immunohistochemical image of CAMK2G and B is the immunohistochemical image of SNCAIP. Figure 24 This is a bar chart showing the quantitative analysis and expression level detection results of the core targets (CAMK2G, SNCAIP, PLCD3, F9) in Example 5 of this application; where C and D are the semi-quantitative analysis results of CAMK2G and SNCAIP immunohistochemistry, respectively; E and F are the relative mRNA expression levels of PLCD3 in uterine tissue and hippocampal tissue, respectively; G is the relative mRNA expression level of F9 in liver tissue; and H and I are the protein expression levels of coagulation factor IX (F9) in liver tissue and serum, respectively. Figure 25 This is a bar chart showing the statistical results of interleukin-1β (IL-1β) and interleukin-6 (IL-6) levels in rat serum in Example 6 of this application; where A represents the statistical results of interleukin-1β (IL-1β) levels and B represents the statistical results of interleukin-6 (IL-6) levels. Figure 26This is a bar chart showing the statistical results of malondialdehyde (MDA) levels, superoxide dismutase (SOD) levels, and reduced / oxidized glutathione (GSH / GSSG) ratios in rat serum in Example 6 of this application; where A represents the statistical results of MDA levels, B represents the statistical results of SOD levels, and C represents the statistical results of the reduced / oxidized glutathione (GSH / GSSG) ratio. Figure 27 This is a bar chart showing the statistical results of the relative mRNA expression levels of SOD1, NFE2L2, and HMOX1 in rat hippocampus tissue in Example 6 of this application; where A represents the statistical results of the relative mRNA expression level of SOD1, B represents the statistical results of the relative mRNA expression level of NFE2L2, and C represents the statistical results of the relative mRNA expression level of HMOX1. Detailed Implementation

[0024] The embodiments of the present invention will be described in detail below with reference to examples. However, those skilled in the art will understand that the following examples are for illustrative purposes only and should not be considered as limiting the scope of the invention. Unless otherwise specified in the examples, conventional conditions or conditions recommended by the manufacturer are followed. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0025] This application provides an application of a Qi-nourishing and Blood-nourishing formula in the preparation of a drug for the prevention and treatment of premenstrual syndrome. The active ingredients of the Qi-nourishing and Blood-nourishing formula include pig blood extract, astragalus, and jujube.

[0026] In some specific implementation schemes, the pig blood extract, astragalus, and jujube can be prepared using modern traditional Chinese medicine extraction processes to obtain effective active ingredients. For example, through appropriate solvent extraction, separation, purification, and drying steps, components capable of exerting synergistic preventive and therapeutic effects can be obtained. The pig blood extract provides a direct active material basis for the drug, and when combined with astragalus and jujube extracts, it effectively achieves synergistic effects among the components.

[0027] When preparing the above-mentioned active ingredients into a drug for the prevention and treatment of premenstrual syndrome, the preparation process includes, but is not limited to, mixing the above-mentioned active ingredients with pharmaceutically acceptable excipients. The pharmaceutically acceptable excipients can be selected according to conventional formulation processes to suit different routes of administration and dosage form requirements. The preparation process may include steps such as pulverization, sieving, mixing, granulation, drying, and molding to obtain a structurally stable drug product with high bioavailability. The dosage form of the drug product is not strictly limited; for example, it can be prepared as an oral formulation for the convenience of patients.

[0028] The drug prepared in this embodiment is specifically targeted for the prevention and treatment of premenstrual syndrome (PMS). In practical applications, PMS typically manifests as cyclical physiological and behavioral abnormalities in women. To verify the drug's efficacy in preventing and treating this disease, those skilled in the art can use recognized animal models of PMS for evaluation. For example, by constructing a disease model through specific hormonal intervention combined with stress treatment, and administering the drug prepared above, standardized experiments (such as open field tests, forced swimming tests, sucrose preference tests, and other behavioral tests, as well as physiological indicator detection) can significantly observe a marked improvement and reversal of the related abnormal symptoms in the model animals, thereby confirming the effectiveness and practical value of the active ingredient combination in the preparation of a drug for the prevention and treatment of PMS.

[0029] In some embodiments, the pig blood extract includes heme iron.

[0030] In a preferred embodiment, the active ingredient of the Qi-nourishing and Blood-nourishing Formula of the present invention includes heme iron in the pig blood extract. Using a pig blood extract containing heme iron as the core material basis, its bioavailability is significantly higher than that of conventional inorganic iron components. In practical pharmaceutical applications, this heme iron component can be rapidly absorbed and utilized by the body, quickly exerting its effect of nourishing Yin and blood, thereby specifically improving the pathological basis of Qi and blood deficiency in patients with premenstrual syndrome, and exhibiting good gastrointestinal tolerance and drug safety. To ensure the stability of the drug's efficacy, in the industrial preparation process, the pig blood extract can be obtained through conventional biological extraction and purification processes (such as enzymatic hydrolysis, separation, concentration, and drying steps), and its heme iron content can be standardized and controlled.

[0031] In some embodiments, the dosage form of the drug is an oral preparation; preferably, the oral preparation includes at least one of tablets, capsules, granules, pills, powders, decoctions, oral liquids, and syrups.

[0032] Regarding the specific dosage form design of the drug, the drug described in this embodiment is configured as an oral preparation suitable for clinical application and patient self-management. Premenstrual syndrome is characterized by periodic attacks, and oral administration can significantly improve patient medication adherence. Preferably, depending on different formulation processes and clinical medication needs, the oral preparation can be formulated as at least one of tablets, capsules, granules, pills, powders, decoctions, oral liquids, and syrups.

[0033] For example, in preparing solid dosage forms (such as tablets, capsules, or granules), the pig blood extract, astragalus extract, and jujube extract can be mixed in a certain proportion, and pharmaceutically acceptable solid dosage form excipients (such as fillers, binders, disintegrants, and lubricants) can be added. After pulverization, mixing, granulation, and drying, the mixture is finally compressed into tablets or filled into capsule shells. In preparing liquid dosage forms (such as oral liquids or syrups), the above-mentioned active ingredients can be dissolved or suspended in a suitable solvent, and preservatives, flavoring agents, and stabilizers can be added as needed. After filling and sterilization, the product is obtained. The above-mentioned wide range of dosage forms allows the drugs of the present invention to adapt to the taking habits and storage needs of different patient groups.

[0034] In some embodiments, the equivalent dose of the drug is 150 mg / kg to 750 mg / kg. For example, it can be 150 mg / kg, 180 mg / kg, 250 mg / kg, 359 mg / kg, 450 mg / kg, 550 mg / kg, 650 mg / kg, 718 mg / kg, 750 mg / kg, etc. In this embodiment of the invention, after the drug is prepared, its dosage in the test body needs to be controlled to ensure its effective prevention and treatment of premenstrual syndrome. Preferably, the equivalent dose of the drug is controlled within the range of 150 mg / kg to 750 mg / kg. In actual pharmaceutical industrial applications and product development, defining the above-mentioned equivalent dose range provides a scientific basis for the specification design and clinical usage of the drug. This dose range establishes an effective therapeutic window for drug intervention. Within this therapeutic window, the active ingredients of the Yiqi Weixue formula can fully exert their systemic regulatory effect, effectively intervening in the coagulation-inflammation-neuroendocrine axis, thereby significantly improving the physiological and emotional abnormalities related to premenstrual syndrome.

[0035] Those skilled in the art will understand that the aforementioned equivalent dosage is typically calibrated based on the weight of standardized laboratory animals (such as rat models). When converting the drug for use in different species (such as human clinical applications), it can be converted using conventional dosage conversion formulas based on body surface area (e.g., target subject dose = original subject dose × original subject body surface area coefficient / target subject body surface area coefficient) to calculate the corresponding single or daily total dose for clinical administration. Pharmaceutical companies can then formulate the drug into an orally produced dosage form with appropriate unit volume based on the calculated total dose and formulation process to ensure that patients can accurately ingest a drug dose within the aforementioned effective range. The dosage can also be adaptively adjusted within the aforementioned limits according to the severity and urgency of symptoms.

[0036] In some embodiments, the drug is used to improve depressive-like behaviors, anxiety-like behaviors, or anhedonia symptoms caused by the premenstrual syndrome.

[0037] In the context of this application, the term "improvement" refers to the reduction or alleviation of the severity of abnormal symptoms such as depressive-like behavior, anxiety-like behavior, or anhedonia exhibited by a subject suffering from premenstrual syndrome or under the induction of specific pathogenic factors, or a benign recovery of related physiological and behavioral indicators towards a normal physiological state. Specifically, the "improvement" described in the embodiments of this application can be quantitatively defined through an objective standardized assessment model, that is, compared with the disease model group that did not receive the drug intervention of this invention, the quantitative parameters (such as total movement distance, time spent in a specific area, resting time, percentage of sucrose preference, etc.) measured in specific behavioral evaluation tests (such as open field test, forced swimming test, sucrose preference test, and marble burial test, etc.) of the test group receiving the drug intervention of this invention show a statistically significant (e.g., P<0.05) benign reversal or trend towards the level of the normal control group.

[0038] It should be noted that premenstrual syndrome is often accompanied by significant mood disturbances, manifesting as periodic low mood (depressive-like behavior), excessive tension and irritability (anxiety-like behavior), and loss of interest in daily activities (anhedonia). These psychobehavioral symptoms severely impact the patient's quality of life. The drug provided by this invention can effectively exert a "liver-regulating and mind-calming" effect, crossing systemic barriers to produce a definite regulatory and reversal effect on the aforementioned abnormal emotional states.

[0039] To objectively verify the effect of the drug of the present invention on the aforementioned psychosocial symptoms, those skilled in the art can use recognized standardized behavioral evaluation models for confirmation. For example, the assessment of depressive-like behavior can be measured through a forced swimming test. In this paradigm, when the subject model is in a state of behavioral despair, it exhibits a significantly prolonged period of immobility; after administration of the drug of the present invention, this immobility period can be significantly shortened, and the active escape behavior can be prolonged, thereby confirming its inhibitory effect on behavioral despair.

[0040] For the assessment of anxiety-like behaviors, open field tests and marble burial tests can be introduced. The drug of this invention can effectively increase the total movement distance, central area dwell time, and number of upright movements of the test model in the open field, while increasing its willingness to explore, indicating that it can effectively alleviate anxiety and obsessive-compulsive tendencies in novel environments or under stress.

[0041] The assessment of anhedonia symptoms typically relies on a sucrose preference test. The evaluation index is based on the formula: sucrose preference rate = sucrose intake / (sucrose intake + regular water intake) × 100%. Premenstrual syndrome models usually show a significant decrease in the sucrose preference rate, while the drug intervention of this invention can significantly increase this preference percentage, objectively demonstrating the effective regulatory ability of the drug of this invention on the central reward system and anhedonia symptoms.

[0042] In summary, the drug prepared in this embodiment provides a safe and well-documented new treatment approach for clinical intervention of depression, anxiety, and anhedonia caused by premenstrual syndrome.

[0043] In some embodiments, the drug treats or prevents premenstrual syndrome by modulating the coagulation-inflammation-neuroendocrine axis and / or by improving systemic oxidative stress and neuroinflammation, and regulating brain-gut axis homeostasis.

[0044] In a further implementation scheme, the core pharmacological mechanism by which the drug described in this application exercisees its therapeutic or preventive effect on premenstrual syndrome lies in its ability to systematically regulate the body's coagulation-inflammation-neuroendocrine axis.

[0045] The pathophysiological process of premenstrual syndrome is highly complex, often manifesting simultaneously as abnormal fluctuations in endocrine hormones, metabolic disorders of central neurotransmitters, and changes in coagulation and inflammation in the peripheral blood microcirculation. The drug described in this application overcomes the limitations of traditional single-target interventions, implementing overall regulation of the aforementioned intertwined pathological network through multi-component synergistic effects to restore the body's systemic homeostasis.

[0046] In the context of this application, the term "regulation of the coagulation-inflammation-neuroendocrine axis" refers to the ability of a drug to simultaneously exert statistically significant benign interventions on the blood coagulation cascade system, immune and inflammatory signaling pathways, and the hypothalamic-pituitary neuroendocrine regulatory network in the test subject. This regulatory effect can be objectively confirmed using modern transcriptomics techniques and systems biology methods.

[0047] For example, in specific evaluation embodiments, those skilled in the art can extract specific target organs (such as hippocampus) from premenstrual syndrome model animals for transcriptome sequencing (RNA-seq). Combined with bioinformatics analysis methods, a large number of differentially expressed genes involved in coagulation factor abnormalities, inflammation-related pathway alterations, and neuroendocrine dysfunction can be observed in the model animals. After administration of the drug intervention of the present invention, the transcriptional profiles of these differentially expressed genes can be significantly reversed. Furthermore, by constructing a protein-protein interaction (PPI) network model and performing module analysis, the core target group of the drug intervention can be clearly identified as being widely and closely distributed in the coagulation, inflammation, and neuroendocrine pathways, thereby confirming at the molecular network level that the drug achieves a multi-system integrated effect in treating premenstrual syndrome by synergistically intervening in the coagulation-inflammation-neuroendocrine axis.

[0048] In some embodiments, the regulation of the coagulation-inflammation-neuroendocrine axis includes at least one of the following pharmacological effects. That is, the specific pharmacological effects of regulating the coagulation-inflammation-neuroendocrine axis can manifest at the microscopic and systemic levels as a benign improvement in at least one of the following physiological and biochemical indicators: A. Upregulating the levels of central neurotransmitters and neurotrophic factors, wherein the central neurotransmitters include serotonin and norepinephrine, and the neurotrophic factors include brain-derived neurotrophic factor (BDNF). In upregulating the levels of central neurotransmitters and neurotrophic factors, the drug can effectively reverse the insufficient expression of central neurotransmitters and neurotrophic factors caused by premenstrual syndrome. Specifically, the central neurotransmitters include serotonin and norepinephrine, and the neurotrophic factors include BDNF. Those skilled in the art can extract specific brain tissue (such as hippocampus) from subjects and use quantitative techniques such as enzyme-linked immunosorbent assay (ELISA) to find that the drug intervention can significantly increase the concentration of the above substances in brain tissue. This pharmacological effect provides a direct neurochemical basis for the drug's ability to improve mood and behavioral disorders such as depression and anxiety.

[0049] B. Upregulating sex hormone levels, including estradiol and progesterone. In upregulating sex hormone levels, the drug can intervene in and correct the hypothalamic-pituitary-gonadal axis imbalance associated with premenstrual syndrome. Specifically, it upregulates the secretion levels of sex hormones such as estradiol and progesterone in the subjects. Confirmation by collecting serum samples and detecting the concentration of relevant hormones demonstrates that the drug of this invention can effectively alleviate the abnormally low hormone levels in the disease model, thereby helping to regulate physiological fluctuations before and after the menstrual cycle from the endocrine source.

[0050] C. Improves blood circulation and reduces hypercoagulability and blood stasis tendency. Regarding improving peripheral microcirculation, the drug effectively reduces hypercoagulability and blood stasis tendency. Premenstrual syndrome is often accompanied by an increased risk of microthrombus formation or blood viscosity induced by endocrine and stress signals. The drug of this invention improves tissue microcirculation by intervening in coagulation pathways and blood rheological properties, thereby alleviating physical symptoms such as tissue edema and local congestion.

[0051] D. Improvement of Systemic Oxidative Stress and Neuroinflammation. In improving systemic oxidative stress and neuroinflammation, the drug exhibits remarkable synergistic protective capabilities against the brain-gut axis and the central nervous system microenvironment. Patients with premenstrual syndrome often experience drastic endocrine fluctuations and physical stress, leading to intestinal microecological imbalance and increased levels of harmful metabolites (such as para-cresol). These substances accumulate in the brain regions via systemic circulation, crossing the blood-brain barrier. They abnormally activate the NOX4 pathway in nerve cells, triggering a strong oxidative stress and neuroinflammation cascade, which in turn damages neurons in core brain regions such as the hippocampus and exacerbates negative emotions such as depression and anxiety. The multi-component drug of this invention can effectively intervene in this pathological process: on the one hand, by significantly downregulating the excessive release of core pro-inflammatory factors such as interleukin-1β (IL-1β) and interleukin-6 (IL-6) in the central and peripheral systems, it effectively suppresses systemic and local neuroinflammation; on the other hand, it can block the excessive generation of free radicals at the source, reduce lipid peroxidation damage, and restore normal redox homeostasis of cells. This dual approach of "anti-inflammatory" and "antioxidant" not only eliminates the toxic pathological factors that lead to decreased synaptic plasticity, but also provides core neuroprotective mechanisms to support the improvement of complex mental and behavioral abnormalities in patients with premenstrual syndrome.

[0052] The aforementioned pharmacological effects A, B, C, and D can occur independently, but under the multi-component drug intervention in the embodiments of this application, they often produce synergistic effects through a complex system network, jointly achieving the overall prevention and treatment efficacy for premenstrual syndrome.

[0053] In some embodiments, to objectively quantify and confirm the pharmacological effects of the drug of the present invention in "improving blood circulation and reducing hypercoagulable state and tendency to blood stasis," those skilled in the art can use clinically standard coagulation function tests and blood rheology testing systems for evaluation. The improvement in blood circulation is manifested as an improvement in at least one of the following pharmacological indicators: A. Prolongation of at least one of prothrombin time, thrombin time, and activated partial thromboplastin time. This refers to the prolongation of extrinsic or intrinsic coagulation time. Specifically, it manifests as prolongation of at least one of prothrombin time (PT), thrombin time (TT), and activated partial thromboplastin time (APTT). The stress associated with premenstrual syndrome often leads to an abnormally shortened coagulation time, indicating a hypercoagulable state where microthrombi are easily formed. Measurements using a fully automated coagulation analyzer on plasma samples from subjects revealed that administration of the drug of this invention significantly prolongs the time of these indicators, demonstrating its definite anticoagulant activity.

[0054] B. Reduce fibrinogen levels. The above reduces the absolute level of fibrinogen (FIB) in plasma. Fibrinogen is a core macromolecule mediating blood coagulation and erythrocyte aggregation. In vitro experiments, including the thrombin method, have confirmed that the drug of this invention can effectively inhibit abnormally elevated fibrinogen expression, fundamentally blocking the excessive amplification of the coagulation cascade and improving blood stasis tendencies.

[0055] C. Reduce at least one of whole blood viscosity, plasma viscosity, hematocrit, and erythrocyte sedimentation rate. The above-mentioned improvements in blood rheological properties reduce blood viscosity. Specifically, this manifests as a reduction in at least one of the following: whole blood viscosity (covering high, medium, and low shear rates), plasma viscosity, hematocrit, and erythrocyte sedimentation rate (ESR). Abnormally elevated levels of these indicators typically indicate increased microcirculatory perfusion resistance. Detection using a blood rheometer confirms that the drug intervention of this invention can significantly reduce blood viscosity under different shear conditions, reduce abnormal accumulation of red blood cells, thereby effectively restoring hemodynamic abnormalities caused by disease and promoting microcirculatory blood supply to peripheral tissues.

[0056] In some embodiments, premenstrual syndrome is not only characterized by simple endocrine or neurotransmitter disorders, but its pathophysiological process is also deeply accompanied by systemic immune stress and inflammatory network imbalance. The drug of this invention, in regulating the coagulation-inflammation-neuroendocrine axis, also exhibits a definite improving effect on immune-inflammatory markers, thereby exerting a comprehensive preventive and therapeutic effect at the neuroimmunological and peripheral immune defense levels. The regulation of the coagulation-inflammation-neuroendocrine axis also includes improving immune-inflammatory markers, specifically manifested as an improvement in at least one of the following pharmacological indicators: A. Reduces serum interleukin-6. Improves serum inflammation levels and inhibits neuroinflammation. Specifically, it reduces serum interleukin-6 (IL-6) levels in subjects. Modern medicine believes that abnormal accumulation of pro-inflammatory cytokines in the central nervous system directly interferes with the plasticity of neural synapses, thereby inducing depressive and anxiety-like behaviors. Those skilled in the art have confirmed through serum extraction and analysis using enzyme-linked immunosorbent assay (ELISA) and other techniques that the drug of this invention can significantly inhibit the overexpression of pro-inflammatory factors, thus confirming its material basis for improving premenstrual syndrome-related mood disorders from an anti-inflammatory mechanism perspective.

[0057] B. Upregulating the percentage of monocytes in the blood. This regulates peripheral immune homeostasis and upregulates the percentage of monocytes in the blood. The occurrence of stress-related disorders such as depression is associated with multiple peripheral immune phenotypes. Premenstrual syndrome modeling stress typically leads to a significant decrease in the percentage of monocytes in peripheral blood leukocyte differential counts, weakening the body's basic immune capacity. Conventional complete blood cell analyzer measurements revealed that the drug intervention of this invention effectively restored and significantly upregulated the percentage of monocytes in the whole blood of test animals. This result objectively reflects that the drug of this invention achieves the overall therapeutic goal of "tonifying qi to strengthen the foundation" by upregulating the function of the peripheral immune system, and this peripheral indicator provides a basis for convenient monitoring of clinical efficacy.

[0058] C. Reduce serum malondialdehyde (MDA) levels; D. Upregulate serum superoxide dismutase (SOD) levels and / or the ratio of reduced glutathione to oxidized glutathione. Regarding the confirmation of the above-mentioned improvements in oxidative stress indicators, the pathological process of premenstrual syndrome is often accompanied by systemic redox homeostasis imbalance and excessive oxidative stress. Among these, the level of malondialdehyde (MDA) directly reflects the degree of lipid peroxidation damage in the body; superoxide dismutase (SOD) represents the activity of core antioxidant enzymes in the body; and the ratio of reduced glutathione to oxidized glutathione (GSH / GSSG) indicates the cell's reducing capacity. Those skilled in the art have confirmed through serum collection and standard colorimetric methods that subjects in a disease model state exhibit a severe oxidative stress state characterized by significantly abnormally elevated MDA levels and significantly decreased SOD activity and GSH / GSSG ratio. After administration of the drug intervention of this invention, serum MDA levels can be significantly downregulated, while SOD levels and the GSH / GSSG ratio are simultaneously upregulated. The improvement in these pharmacological indicators objectively verifies, from the perspective of systemic redox homeostasis, that the drug of this invention can effectively alleviate oxidative stress damage and enhance the body's antioxidant defense network at the systemic level. This provides solid peripheral mechanism evidence and data support for the drug to intervene in and improve central nervous system oxidative stress and nerve cell damage through the "brain-gut axis".

[0059] In some embodiments, the drug is used to upregulate the subject's mean red blood cell volume and / or mean red blood cell hemoglobin concentration.

[0060] The Qi-nourishing and Blood-tonifying formula described in this application embodiment not only regulates the neuroendocrine network but also demonstrates a definite effect on improving the morphology and quality of red blood cells, thus confirming its "blood-nourishing" and "blood-tonifying" efficacy at the microscopic hematological level. In a specific implementation, the drug is specifically used to upregulate the mean erythrocyte volume (MCV) and / or mean erythrocyte hemoglobin concentration (MCHC) of the subject.

[0061] Patients with premenstrual syndrome often experience hormonal fluctuations and psychological stress, resulting in a sub-optimal state of their blood system, frequently manifested as a decrease in parameters related to the oxygen-carrying capacity of red blood cells. The reduction in red blood cell volume and hemoglobin concentration directly leads to insufficient oxygen supply to the body, especially to the brain and peripheral tissues, thereby inducing or aggravating symptoms such as fatigue, dizziness, and low mood.

[0062] To verify the effect of the drug of this invention on improving red blood cell quality, those skilled in the art can evaluate it using a conventional complete blood cell analyzer. In specific experimental tests, anticoagulated whole blood was collected from test animals and analyzed using routine blood tests. The results showed that in subjects under a premenstrual syndrome model, the mean corpuscular volume (MCV) and mean corpuscular hemoglobin (MCH) both exhibited an abnormally decreasing pathological trend; however, after receiving intervention with the drug of this invention, the active ingredients such as heme iron in the drug could be efficiently utilized by the body, significantly upregulating and restoring the above two key red blood cell parameters. This pharmacological effect effectively improves the oxygen-carrying capacity of red blood cells and improves the body's microcirculatory oxygen supply environment, providing a solid objective basis for the prevention and treatment of premenstrual syndrome from a hematological perspective. It also provides a reliable biological indicator for convenient monitoring of clinical efficacy (such as through routine blood tests).

[0063] To further elucidate the underlying molecular mechanisms by which the drug of this invention regulates the coagulation-inflammation-neuroendocrine axis, the embodiments of this application further reveal specific regulatory targets of the drug at the molecular genetics and proteomics levels. In some embodiments, the drug is used to upregulate at least one of the following indicators: A. Serum and liver coagulation factor IX levels in target subjects. Regarding the confirmation of the upregulation of serum coagulation factor IX levels, the F9 gene encodes coagulation factor IX, which is involved in the intrinsic coagulation pathway. The premenstrual syndrome model exhibits dysregulation of coagulation-related factors, leading to a significant decrease in serum coagulation factor IX levels. Extraction of serum and liver tissue homogenate supernatants from test subjects and detection using enzyme-linked immunosorbent assay (ELISA) confirmed that the drug of this invention can effectively reverse and significantly increase the protein concentration of coagulation factor IX in serum, thereby verifying its regulatory effect on the coagulation-anticoagulation balance at the systemic circulation level.

[0064] B. mRNA expression levels of PLCD3 in hippocampal and uterine tissues and F9 in liver tissue. Regarding the confirmation of the upregulation of PLCD3 and F9 mRNA expression levels in hippocampal and uterine tissues and liver tissue, PLCD3, as a core gene regulating calcium signaling and inflammatory metabolism, is closely related to local tissue microcirculatory disturbances and inflammation. The F9 gene encodes coagulation factor IX, which participates in the intrinsic coagulation pathway. Total RNA was extracted from hippocampal, uterine, and liver tissues, reverse transcribed, and amplified by real-time quantitative PCR (RT-qPCR), with relative quantification calculated using the 2^-ΔΔCt method. The results confirmed that the drug intervention of this invention can specifically upregulate the abnormally suppressed PLCD3 transcription level in hippocampal and uterine tissues and the abnormally suppressed F9 transcription level in liver tissue, thereby blocking the excessive cascade of inflammatory signaling pathways and helping to maintain normal coagulation function, preventing abnormal bleeding or microthrombus formation, and thus improving tissue microcirculation.

[0065] C. Protein expression levels of CAMK2G and SNCAIP in hippocampal tissue. Regarding the confirmation of the upregulation of CAMK2G and SNCAIP protein expression levels in the hippocampus, CAMK2G is associated with synaptic plasticity and neurotransmitter release, while SNCAIP is directly related to neuronal survival and neuroprotective function. Those skilled in the art can use immunohistochemical (IHC) staining to perform morphological and semi-quantitative analysis of the hippocampus. Microscopic observation and image analysis (such as analysis of positive expression area and average optical density) show that the expression of these two proteins in the cytoplasm and processes of hippocampal neurons is significantly reduced in the model state; however, the drug intervention of this invention can significantly enhance the positive expression of the above-mentioned target proteins. This targeting effect provides direct morphological and molecular evidence at the protein level for drug repair of central nervous system damage and improvement of negative emotional behaviors.

[0066] D. mRNA expression levels of NFE2L2, HMOX1, and SOD1 in the hippocampus of the target subjects. Regarding the confirmation of the upregulation of NFE2L2, HMOX1, and SOD1 mRNA expression levels in the hippocampus, NFE2L2, as a core transcriptional regulator of the cellular antioxidant defense system, works in conjunction with its downstream target genes HMOX1 (heme oxygenase-1) and SOD1 (superoxide dismutase 1) to construct a key defense against oxidative stress. The pathological state of premenstrual syndrome and the accumulation of harmful intestinal metabolites in the brain easily induce excessive oxidative stress and neuroinflammation in hippocampal neurons, leading to abnormal consumption and inhibition of the expression of the aforementioned antioxidant genes. Those skilled in the art have confirmed through extraction of total RNA from the hippocampus of the subjects and real-time quantitative PCR (RT-qPCR) that the drug intervention of this invention can effectively reverse this pathological depletion process, specifically and significantly upregulating the transcriptional levels of NFE2L2 and its classic target genes HMOX1 and SOD1. This targeting effect provides direct molecular evidence at the gene transcription level for the drug of this invention to systematically initiate the endogenous antioxidant defense response of central nervous cells, scavenge excess reactive oxygen species, and reduce nerve cell damage and apoptosis.

[0067] The present invention will be further illustrated below with specific embodiments. However, it should be understood that these embodiments are merely for the purpose of more detailed illustration and should not be construed as limiting the present invention in any way.

[0068] For ease of explanation and correspondence with the markings in the accompanying drawings, the English abbreviations of experimental groups, testing methods, physiological and biochemical indicators, and target genes involved in the following embodiments and related drawings of the present invention are as follows: 1. Experimental group abbreviations: Control: Normal control group (or blank control group), no modeling treatment, only given an equal volume of physiological saline; Vehicle: Solvent control group, only injected with sesame oil and given an equal volume of physiological saline; PMS: Premenstrual Syndrome model group, modeled by progesterone injection combined with restraint stress; KTT (or PMS+KTT): Kuntai Capsule group (positive control drug); HXT (or PMS+HXT): Honghua Xiaoyao Tablet group (positive control drug); YQWX (or PMS+YQWX): Yiqi Weixue Formula administration group; where YQWX-L represents the low-dose group, YQWX-M represents the medium-dose group, and YQWX-H represents the high-dose group.

[0069] 2. Abbreviations for behavioral tests: FST: Forced Swimming Test; SPT: Sucrose Preference Test; MBT: Marble Burying Test.

[0070] 3. Neuroendocrine system indicators: 5-HT: serotonin; NE: norepinephrine; BDNF: brain-derived neurotrophic factor; E2: estradiol; PROG: progesterone.

[0071] 4. Blood circulation, immune and biochemical indicators: PT: Prothrombin Time; TT: Thrombin Time; APTT: Activated Partial Thromboplastin Time; FIB: Fibrinogen; WBV: Whole Blood Viscosity, including Low-shear, Medium-shear and High-shear; PV: Plasma Viscosity; ESR: Erythrocyte Sedimentation Rate; HCT: Hematocrit; MCV: Mean Corpuscular Volume; MCHC: Mean Corpuscular Hemoglobin Concentration; WBC: White Blood Cell; RBC: Red Blood Cell. Cell); HGB: Hemoglobin; MONO%: Monocyte Percentage; IL-6: Interleukin-6.

[0072] 5. Core Molecular Targets and Analytical Methods: F9: Coagulation factor IX; PLCD3: Phospholipase C delta 3; CAMK2G: Calcium / calmodulin-dependent protein kinase II gamma; SNCAIP: Synuclein alpha interacting protein; ELISA: Enzyme-linked immunosorbent assay; RT-qPCR: Real-time quantitative PCR; IHC: Immunohistochemistry; PCA: Principal Component Analysis; PPI: Protein-Protein Interaction; KEGG / GO: Kyoto Encyclopedia of Genes and Genomes / Gene Ontology Functional Annotation. Ontology).

[0073] Experimental Example 1: Construction and Dosing Regimen of a Complex Animal Model of Premenstrual Syndrome (PMS) This experimental example provides a method for constructing an animal model to verify the efficacy of drugs in preventing and treating premenstrual syndrome. It also specifies the grouping of animals and the administration intervention plan of Yiqi Weixue formula, laying the experimental foundation for subsequent evaluation of various indicators.

[0074] 1. Experimental materials and animal grouping: (1) Animals: SPF-grade female Wistar rats, 6-8 weeks old, weighing 170-220g. They were acclimatized under specific pathogen-free conditions before the formal experiment.

[0075] (2) Drugs, reagents and instruments: Yiqi Weixue Formula (provided by Guangdong Hongshanhu Pharmaceutical Co., Ltd.); positive control drug Kuntai Capsules (Guizhou Xintian Pharmaceutical Co., Ltd.), raw material of Honghua Xiaoyao Tablets (Jiangxi Puzheng Pharmaceutical Co., Ltd.). Modeling reagents: progesterone (Shanghai Yuanye Biotechnology Co., Ltd., catalog number: s30586), sesame oil (Beijing Solarbio Technology Co., Ltd.). Detection reagents: 5-hydroxytryptamine (5-HT), norepinephrine (NE), brain-derived neurotrophic factor (BDNF), coagulation factor IX (F9) rat ELISA kit (Shanghai Enzyme-Linked Biotechnology Co., Ltd.); CAMK2G rabbit antibody (Wuhan Aibote Biotechnology Co., Ltd., catalog number: A21668); SNCAIP rabbit antibody (Shanghai Aibite Pharmaceutical Technology Co., Ltd., PA2411S); total RNA extraction kit, reverse transcription and SYBR quantitative kit and other conventional molecular biology reagents. Main instruments: fully automated microplate reader (Thermo Fisher Scientific, USA, model MK3), microscope (OLYMPUS, Japan, model BX51), QuantStudio3 real-time fluorescence quantitative gene amplification system (Thermo Fisher Scientific, USA), fully automated coagulation analyzer, and blood rheometer, etc.

[0076] (3) Grouping and Model Establishment (Progesterone Injection + Restraint Stress): After eliminating rats with behavioral scores significantly below the average level through the open field test, the rats were randomly divided into 8 groups: normal (blank) group, solvent group, model group, positive drug Kuntai capsule group (KTT group), positive drug Honghua Xiaoyao tablet group (HXT group), and low, medium, and high dose groups of Yiqi Weixue Fang. Except for the normal group and solvent group, the other groups were injected intraperitoneally with progesterone suspension (6 mg / rat) and restrained with a restraint device for 5 hours every day to establish a composite PMS animal model. Specifically, the progesterone suspension was prepared by dissolving progesterone in sesame oil and stirring overnight at 37°C. The total volume of each intraperitoneal injection was 0.2 mL / rat.

[0077] 2. Dosage regimen: The dosage was calculated based on the animal's body weight and the intervention lasted for two weeks. (1) Low-dose group of Yiqi Weixue Formula: 180 mg / kg was administered by gavage; (2) Medium-dose group of Yiqi Weixue Formula: 359 mg / kg was administered by gavage; (3) High-dose group of Yiqi Weixue Formula: 718 mg / kg was administered by gavage; (4) Positive drug group (e.g., KTT group): Kuntai capsules (540 mg / kg) of equivalent dose were administered by gavage; (5) Normal group and solvent group: an equal volume of physiological saline was given. After the intervention, behavioral tests were performed, and then whole blood, serum, whole brain (hippocampus), uterus and other tissues of rats were collected for subsequent tests.

[0078] Experimental Example 2: Investigation on the effect of Yiqi Weixue Formula on the improvement of behavioral abnormalities in PMS rats This experimental example aims to investigate the therapeutic effect of Yiqi Weixue Formula on improving the core mental symptoms such as depressive-like behavior, anxiety-like behavior, anhedonia, and social impairment induced by PMS through a series of standardized animal ethological experiments.

[0079] 1. Experimental methods: (1) Open field test: The rats were placed in the center of an open field box with dimensions of 100 cm × 100 cm × 40 cm, and the total movement distance, activity time, standing times, and residence time in the central area within 5 minutes were recorded. After each test, the box was sprayed with 5% acetic acid aqueous solution to remove the smell. (2) Sucrose preference test: The animals were housed individually. First, a bottle of 2% sucrose solution and a bottle of ordinary water were provided for 24-hour sucrose exposure training and adaptation. Water was withheld for 12 hours before the baseline test, and then a bottle of sucrose solution and a bottle of ordinary water were provided. The water intake within 1 - 2 hours was recorded, and the sucrose preference rate (sucrose intake / total water intake × 100%) was calculated. (3) Marble burying test: To evaluate the anxiety and compulsive tendencies of the animals (recording the number of buried marbles). (4) Forced swimming test: A single rat was placed in a cylindrical transparent glass tank (diameter 25 cm, water depth 30 cm), and the water temperature was controlled at about 25°C. The rat swam for 6 minutes, and the total passive floating time (immobile time / stationary time) during the last 4 minutes, when the rat stopped active struggling and only maintained its head above the water surface, was recorded. (5) Three-chamber social test: The interaction time of the rats was recorded to evaluate social ability and social novelty preference.

[0080] 2. Experimental results and analysis: (Subheading 1) Improvement of anxiety and reduction of exploratory willingness: Referring to Figure 1 , Figure 2 and Figure 3 , in the model group, indicators such as the total movement distance in the open field significantly decreased, and the residence time in the four-week area significantly increased (P < 0.01). Compared with the model group, the medium and high-dose groups of Yiqi Weixue could significantly increase the total movement distance, activity time, and threading times, and the high-dose group significantly reduced the residence time in the four-week area. This indicates that the drug can increase the activity of rats and enhance their exploratory willingness. (Subheading 2) Improvement of anhedonia: Referring to Figure 4 , the percentage of sucrose preference in the model group significantly decreased. The percentage in the medium and high-dose administration groups of Yiqi Weixue significantly rebounded (P < 0.05, P < 0.01), effectively regulating the anhedonia in the core symptoms of depression. (Subheading 3) Alleviation of anxiety and compulsive behavior: Referring to Figure 5 , the number of buried marbles in the model group significantly increased. All low, medium, and high-dose groups of Yiqi Weixue could significantly reduce (improve) the number of glass marbles buried by rats (P < 0.05 - P < 0.01). (Subheading 4) Improvement of behavioral despair (depressive-like state): Referring to Figure 6 and Figure 7The immobility time in the model group was significantly longer than that in the solvent group (P<0.001). The immobility time in each dose group of the Qi-invigorating and blood-nourishing treatment was significantly shorter than that in the model group (P<0.01), and the efficacy was comparable to that of the positive control drug, effectively alleviating "behavioral despair". (5) Improvement of social barriers: Reference Figure 8 The interaction time of rats in the model group decreased significantly. The interaction time of the low- and medium-dose groups of Yiqi Weixue Fang increased significantly, effectively improving social impairment.

[0081] Experimental Example 3: Investigation on the effect of Yiqi Weixue Formula on blood circulation and immune indicators in PMS rats This experimental case focuses on examining the specific manifestations of the "Qi- and Blood-regulating" effects of the Yiqi Weixue formula in improving coagulation function, blood rheological properties, and routine blood immune-related indicators, thereby reducing hypercoagulable state and blood stasis tendency.

[0082] 1. Experimental Methods: Whole blood and serum were collected from rats. A complete blood count (CBC) instrument was used to measure blood count (red blood cell count, white blood cell differential, hemoglobin concentration, etc.). A coagulation analyzer was used to measure coagulation parameters (FIB, APTT, PT, TT). A blood rheometer was used to measure whole blood viscosity and plasma viscosity at different shear rates. In addition, the concentration of interleukin-6 (IL-6) in rat serum was detected using an enzyme-linked immunosorbent assay (ELISA) kit.

[0083] 2. Experimental Results and Analysis: (1) Improvement of coagulation function: Reference Figure 9 In the model group, the level of fibrinogen (FIB) was significantly increased, and the values ​​of APTT, PT, and TT were significantly shortened, indicating a severe hypercoagulable state. After intervention, the high-dose group of Yiqi Weixue significantly prolonged APTT, PT, and TT (all P<0.01) and significantly reduced FIB level (P<0.05). Compared with the positive drug KTT, its prolongation effect on APTT and PT was more significant. This result indicates that compared with simple blood-activating drugs, Yiqi Weixue formula, through the combination characteristics of "invigorating qi and promoting blood circulation, and replenishing qi to promote blood circulation", can not only activate blood circulation and unblock the meridians, but also fundamentally correct the coagulation dysfunction and fibrinolytic system imbalance caused by "qi not governing blood" in premenstrual syndrome. (2) Improve blood rheology abnormalities: refer to Figure 10 and Figure 11 In the model group, the viscosity of whole blood at various shear rates, plasma viscosity, hematocrit, and erythrocyte sedimentation rate increased significantly. The medium and high dose groups of Yiqi Weixue could significantly reverse the above-mentioned blood viscosity-related indicators (P<0.05~P<0.001), and significantly improve the concentration, viscosity, and aggregation of peripheral blood. (3) Improvement of immune and erythrocyte indicators (the effect of Weixue): Reference Figure 12The percentage of monocytes in the model group decreased significantly. Each dose of Yiqi Weixue significantly increased the percentage of monocytes (P<0.05~P<0.01). At the same time, the treatment group significantly increased the mean corpuscular volume and mean corpuscular hemoglobin concentration, reflecting the effect of Yiqi to enhance the function of the immune system and improve the ischemic and hypoxic state. Studies have shown that stress signals are transmitted from the central nervous system to immune cells, and monocytes are positively correlated with the occurrence of depression. Yiqi Weixue formula significantly increased the percentage of monocytes and the core parameters of erythrocytes, which proved that it can improve the immune system defense function and the oxygen-carrying capacity of peripheral blood by increasing the function of the immune system defense and improving the oxygen-carrying capacity of peripheral blood, and effectively verified the overall therapeutic effect of the compound "Yiqi" and "Weixue". (4) Improve the body's inflammatory state: refer to Figure 13 In the model group, the serum interleukin-6 (IL-6) level in rats was significantly higher than that in the normal group (P<0.001), indicating that the body was in a state of significant inflammatory stress. All doses of the Yiqi Weixue formula significantly reduced IL-6 levels (P<0.001). This result, combined with the aforementioned upregulation of monocyte percentage by the drug, fully confirms that the drug can effectively improve the immune-inflammatory imbalance associated with premenstrual syndrome, and exert a comprehensive protective effect in anti-neuroinflammatory and peripheral immune homeostasis regulation.

[0084] Experiment Example 4: Investigation on the effect of Yiqi Weixue formula on endocrine and central neurotransmitter improvement in PMS rats This experimental case investigated the material basis of the Qi-invigorating and blood-nourishing formula's effects on "harmonizing the liver and spirit" and regulating endocrine imbalances, specifically assessing changes in sex hormone levels, monoamine neurotransmitters, and neurotrophic factors.

[0085] 1. Experimental methods: The levels of estradiol (E2) and progesterone (PROG) in rat serum were detected by ELISA; and the concentrations of 5-hydroxytryptamine (5-HT), norepinephrine (NE) and brain-derived neurotrophic factor (BDNF) were detected by ELISA kits after extracting hippocampal tissue homogenate.

[0086] 2. Experimental Results and Analysis: (1) Restoration of Sex Hormone Levels: Reference Figure 14 The concentrations of E2 and PROG in the model group decreased significantly. The medium and high doses of Yiqi Weixue significantly upregulated the levels of E2 and PROG in rats (P<0.05~P<0.001), and the regulatory effect of the high dose was significantly better than that of the positive control drug KTT group. (2) Upregulation of neurotransmitters and neurotrophic factors: Reference Figure 15 , Figure 16 and Figure 17The levels of 5-HT, NE, and BDNF in the hippocampus of the model group were significantly reduced, which confirms the traditional Chinese medicine pathogenesis of "insufficient liver blood leading to malnourishment of the spirit" in PMS. The medium-dose group of the Qi-tonifying and Blood-nourishing formula significantly upregulated 5-HT and BDNF levels (P<0.01), while the high-dose group significantly upregulated NE levels (P<0.001). This confirms the advantages of the drug's combination of "Qi generates blood, and blood carries Qi"—by nourishing liver blood and regulating Qi, it specifically regulates the neurotransmitter network of 5-HT and NE and neural plasticity. Its efficacy is highly consistent with the traditional Chinese medicine principle of "harmonizing the liver and spirit," providing precise intervention for the emotional abnormalities associated with "deficiency of both Qi and blood" in premenstrual syndrome.

[0087] Experiment Example 5: Confirmation of the multi-target mechanism of the Qi-tonifying and Blood-nourishing formula regulating the "coagulation-inflammation-neuroendocrine axis" based on multi-omics. This experimental case used transcriptome sequencing and systems biology methods to reveal the core molecular network of Yiqi Weixue formula in the treatment of PMS, and rigorously validated the core key gene / protein targets screened by RT-qPCR, ELISA and immunohistochemistry (IHC) techniques.

[0088] 1. Experimental methods: (1) Transcriptome sequencing: RNA was extracted from hippocampal tissue and DNBSEQ-T7 paired-end sequencing was performed. Differentially expressed genes (DEGs) were screened using edgeR, and GO and KEGG enrichment analysis and PPI network construction were performed. (2) Molecular validation: mRNA expression of PLCD3 in hippocampal and uterine tissues and F9 in liver tissue was detected by RT-qPCR: Total RNA was extracted and reverse transcribed into cDNA. GAPDH was used as an internal reference gene. The cDNA was amplified on a real-time PCR instrument using the primers shown in Table 1 and the SYBR quantitative kit. -ΔΔCt The relative expression level was calculated using the method described above. The primer sequences used for RT-qPCR detection are shown in Table 1 (NO. represents SEQ ID: NO.): Table 1. Primer sequences used for RT-qPCR detection

[0089] ELISA detection: Serum from rats in each group and liver tissue homogenate supernatant were extracted, and the concentration of coagulation factor IX protein was detected strictly according to the kit instructions.

[0090] Immunohistochemical (IHC) staining: Hippocampal tissue was fixed and embedded, sections were prepared, and after antigen retrieval, primary antibodies (CAMK2G or SNCAIP diluted 1:50) were added and incubated overnight at 4°C. The next day, secondary antibodies were added and DAB staining was performed. The positive expression area and average optical density of the target protein were analyzed using image processing software.

[0091] 2. Experimental Results and Analysis: (1) Transcriptional Profile Retrospective and Pathway Enrichment: Reference Figure 18 , Figure 19 , Figure 20 and Figure 21 PCA analysis showed that the gene expression profile of the hippocampus in the model group was significantly disordered, while the drug-treated group showed a trend of returning to normal. 123 core intersection target genes were screened out. KEGG analysis showed that the regulated genes were widely enriched in multiple pathways such as coagulation, inflammation and neuroendocrine. (2) Construction of core regulatory network: refer to Figure 22 Through PPI network analysis, two core signal axes were identified: A) the "coagulation-inflammation balance regulation axis" with F9 and PLCD3 as the core; B) the "neuroendocrine-emotional behavior regulation axis" with CAMK2G and SNCAIP as the core. Changes in target expression were significantly correlated with multidimensional pharmacodynamic indicators. (3) Target expression reversal verification: Refer to Figure 23 and Figure 24 In the middle (C, D) groups, immunohistochemistry confirmed that the expression of CAMK2G and SNCAIP proteins in the hippocampus was significantly reduced in the model group; after administration of medium and high doses of Yiqi Weixue Fang, the positive expression was significantly enhanced (all P<0.001), confirming neuroprotection and the recovery of synaptic plasticity. (Reference) Figure 24 (E~I), RT-qPCR and ELISA results showed that the serum and liver tissue coagulation factor IX protein concentrations of PLCD3 and F9 transcription levels, which were significantly inhibited in the model group, were significantly or extremely significantly upregulated after intervention with Yiqi Weixue Fang (P<0.05 or P<0.001), confirming the precise regulation of coagulation and inflammation-related targets at both systemic and local levels.

[0092] Experimental Example 6: Confirmation of the mechanism by which the Qi-nourishing and Blood-nourishing formula based on the brain-gut axis inhibits oxidative stress and neuroinflammation in the NOX4-NFE2L2 pathway. This experimental case, building upon the aforementioned foundation, further explores the "brain-gut axis" mechanism, focusing on verifying the effects of Yiqi Weixue formula on systemic oxidative stress and neuroinflammation, as well as its gene transcriptional activation effect on the NFE2L2 antioxidant defense pathway. Studies have shown that p-cresol from the gut is converted to p-cresol sulfate (PCS) and accumulates in the brain, inducing severe central oxidative stress and neuronal damage. Yiqi Weixue formula can intervene through a "central-peripheral-central" feedback loop.

[0093] 1. Experimental methods: (1) Detection of core inflammatory factors (ELISA): Serum from rats in each group was extracted, and the concentrations of interleukin-1β (IL-1β) and interleukin-6 (IL-6) were detected strictly according to the instructions of the enzyme-linked immunosorbent assay kit. (2) Detection of oxidative stress and reduction homeostasis indicators (colorimetric method): Serum from rats in each group was collected, and the level of malondialdehyde (MDA), a lipid peroxidation marker, the activity of superoxide dismutase (SOD), and the ratio of reduced glutathione to oxidized glutathione (GSH / GSSG) were measured using a standard colorimetric assay kit. (3) Detection of antioxidant gene mRNA expression in hippocampus (RT-qPCR): Total RNA was extracted from rat hippocampus and reverse transcribed into cDNA. GAPDH was used as an internal reference gene and amplified on a real-time PCR instrument using a SYBR quantitative PCR kit. -ΔΔCt The relative expression levels of NFE2L2, HMOX1, and SOD1 were calculated using a method. Specific primer sequences used for RT-qPCR detection were designed and synthesized using standard primer design software.

[0094] 2. Experimental results and mechanism analysis: (1) Significantly downregulated core pro-inflammatory factors and inhibited neuroinflammatory levels: such as Figure 25 As shown, compared with the solvent group, the serum levels of IL-6 and IL-1β in the model group rats showed extremely significant abnormal increases (both P<0.001). Meanwhile, the medium and high doses of Yiqi Weixue Fang significantly downregulated the levels of IL-6 and IL-1β in rats (P<0.001). This result confirms that Yiqi Weixue Fang can effectively inhibit systemic and neuroinflammatory responses caused by stress and the accumulation of intestinal metabolites. (2) Comprehensive improvement of systemic redox homeostasis: MDA level reflects the degree of lipid peroxidation damage, SOD represents antioxidant enzyme activity, and the GSH / GSSG ratio indicates cellular reducing capacity. Figure 26 As shown, compared with the solvent group, the serum SOD level and GSH / GSSG ratio of the model group rats decreased significantly (P<0.05, P<0.01), and the MDA level increased significantly (P<0.001), indicating severe oxidative stress damage. After intervention, the medium dose of Yiqi Weixue Fang significantly upregulated the serum SOD level and GSH / GSSG ratio of rats (P<0.05), and the low, medium and high dose groups significantly downregulated the serum MDA level of rats (P<0.001). The experimental results show that Yiqi Weixue Fang can effectively alleviate oxidative stress and enhance the body's antioxidant defense network at the overall level. (3) Systematically activate the NFE2L2 antioxidant defense pathway at the gene transcription level: such as Figure 27As shown, compared with the solvent group, the mRNA expression levels of SOD1, NFE2L2 and HMOX1 in the hippocampus of the model group showed a significant pathological decrease (P<0.05). After intervention with Yiqi Weixue Fang, the high-dose administration group was able to significantly reverse this process and specifically increase the mRNA expression levels of SOD1, NFE2L2 and HMOX1 (P<0.01, P<0.001, P<0.001). This molecular-level evidence suggests that Yiqi Weixue Fang can systematically initiate the cellular antioxidant program at the gene transcription level. (4) Summary and discussion of the mechanism: Combining the above objective biochemical and molecular experimental data, and combining the previous metabolomics and target analysis (verified by target molecular docking), it can be reasonably concluded that Yiqi Weixue Fang effectively downregulated NOX4 protein expression and blocked excessive oxidative stress by reshaping intestinal homeostasis, reducing the generation and brain conversion of harmful metabolites such as p-cresol (PCS). The reduction of ROS simultaneously activates the NFE2L2 pathway and its downstream genes (HMOX1, SOD1), effectively exerting anti-inflammatory and neuroprotective effects. This "central-peripheral-central" pharmacological loop provides extremely solid multidimensional mechanistic evidence for the compound's ability to alleviate the depressive and anxiety symptoms associated with premenstrual syndrome.

[0095] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features therein. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. The application of a Qi-tonifying and blood-nourishing formula in the preparation of a drug for the prevention and treatment of premenstrual syndrome, characterized in that, The active ingredients of the Qi-boosting and blood-nourishing formula include pig blood extract, astragalus, and jujube.

2. The application as described in claim 1, characterized in that, The pig blood extract includes heme iron; and / or, The dosage form of the drug is an oral preparation; preferably, the oral preparation includes at least one of tablets, capsules, granules, pills, powders, decoctions, oral liquids, and syrups.

3. The application as described in claim 1, characterized in that, The equivalent dose of the drug is 150 mg / kg to 750 mg / kg.

4. The application as described in claim 1, characterized in that, The drug is used to improve symptoms of depression-like behavior, anxiety-like behavior, or anhedonia caused by the premenstrual syndrome.

5. The application as described in claim 1, characterized in that, The drug treats or prevents premenstrual syndrome by regulating the coagulation-inflammation-neuroendocrine axis and / or by improving systemic oxidative stress and neuroinflammation and regulating brain-gut axis homeostasis.

6. The application as described in claim 5, characterized in that, The regulation of the coagulation-inflammation-neuroendocrine axis includes at least one of the following pharmacological effects: A. Upregulate the levels of central neurotransmitters and neurotrophic factors, wherein the central neurotransmitters include serotonin and norepinephrine, and the neurotrophic factors include brain-derived neurotrophic factors. B. Upregulate sex hormone levels, including estradiol and progesterone; C. Improves blood circulation and reduces hypercoagulability and blood stasis tendency; D. Improves systemic oxidative stress and neuroinflammation.

7. The application as described in claim 6, characterized in that, The improvement in blood circulation is manifested by an improvement in at least one of the following pharmacological indicators: A. Prolonging at least one of prothrombin time, thrombin time, and activated partial thromboplastin time; B. Reduce fibrinogen levels; C. Reduce at least one of whole blood viscosity, plasma viscosity, hematocrit, and erythrocyte sedimentation rate.

8. The application as described in claim 6, characterized in that, The regulation of the coagulation-inflammation-neuroendocrine axis also includes improving immune-inflammatory markers, specifically manifested as an improvement in at least one of the following pharmacological indicators: A. Reduce serum interleukin-6; B. Upregulates the percentage of monocytes in the blood; C. Reduce serum malondialdehyde levels; D. Upregulate serum superoxide dismutase levels and / or the ratio of reduced glutathione to oxidized glutathione.

9. The application as described in claim 1, characterized in that, The drug is used to upregulate the mean red blood cell volume and / or mean red blood cell hemoglobin concentration in subjects.

10. The application as described in claim 1, characterized in that, The drug is used to upregulate at least one of the following indicators: A. Serum and liver tissue coagulation factor IX levels in the target subjects; B. mRNA expression levels of PLCD3 in hippocampal and uterine tissues; C. Protein expression levels of CAMK2G and SNCAIP in hippocampal tissue; D. MRNA expression levels of NFE2L2, HMOX1, and SOD1 in the hippocampus of the target subjects.

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