Jasmine flower extract and application thereof

Abstract

The invention relates to a jasmine flower extract and application thereof. The jasmine flower extract can be used for improving mood and preventing or treating diseases related to neurotransmission impairment. The jasmine flower polypeptide extract does not depend on fragrance, and the effect is remarkable and lasting.

Description

Technical Field

[0001] This application relates to jasmine flower extract and its applications, belonging to the field of biochemical and pharmaceutical technology. Background Technology

[0002] Jasmine (Jasminum sambac (L.) Ait.), also known as jasmine flower, is a plant belonging to the genus Jasminum in the Oleaceae family. It is a known traditional Chinese medicine, with its roots, leaves, and flowers all possessing medicinal value. Jasmine flowers are also edible; in Fujian and Guangxi provinces, there is a tradition of using jasmine flowers in cooking (such as jasmine porridge and jasmine flower scrambled eggs). In Southeast Asian countries, jasmine flowers are widely used in desserts and drinks. Therefore, jasmine is considered a plant with both medicinal and edible properties.

[0003] In recent years, emotional health and diseases or symptoms related to impaired neurotransmission have received increasing attention from society. Because of the pleasant fragrance and safety of jasmine flowers, they are frequently used to make jasmine essential oil or as a fragrance agent to regulate aromas (such as in jasmine tea), and to soothe the mind and relieve depression. In the preparation of jasmine essential oil, jasmine flowers are the sole source of aroma, releasing volatile aromatic compounds such as benzyl acetate (sweet fruity scent), linalool (fresh floral and woody scent), benzyl benzoate, methyl anthranilate, indole (which imparts a mysterious quality in trace amounts), and jasmone (characteristically sweet and warm floral scent). A common method is to soak fresh jasmine flowers in a volatile organic solvent (such as petroleum ether or other fat-soluble solvents), allowing the aromatic oils (fat-soluble) in the petal cells, as well as some waxes and pigments, to dissolve in the solvent, thus obtaining a richly fragrant jasmine essential oil. In the preparation of jasmine tea, the tea leaves absorb the volatile aromatic compounds (such as esters and alcohols) released by fresh jasmine flowers through a scenting process. Then, the withered flowers (i.e., the jasmine flowers that have turned from fresh flowers to wilted) are sieved out of the tea leaves, so that the tea leaves themselves still retain the aroma of jasmine.

[0004] However, because the fragrance comes from volatile aromatic compounds released by jasmine flowers, it quickly dissipates and changes over time, making the pleasant effect of the fragrance short-lived. Summary of the Invention

[0005] To address the aforementioned technical problems, this application provides a "jasmine flower polypeptide extract that is fragrance-independent or fragrance-free," which can be used to regulate or improve mood, and to prevent or treat diseases or conditions related to impaired neurotransmission. Since preparing polypeptide extracts from plant tissues is a conventional technique in this field, such as acid-base hydrolysis, enzymatic hydrolysis, and microbial fermentation; those skilled in the art also know how to remove volatile aromatic substances from polypeptide extracts (e.g., distillation, open-top operation, defatting, etc.), therefore, the "jasmine flower polypeptide extract that is fragrance-independent or fragrance-free" in this application can refer to any jasmine flower polypeptide extract prepared from jasmine flowers using any method and containing little or no volatile aromatic substances. Although crude jasmine flower polypeptide extracts already possess preliminary activity (see...), Figure 4 However, the jasmine flower polypeptide extract, after further extraction or purification (with high polypeptide purity), exhibits more significant activity (see...). Figure 5-7 ).

[0006] Generally, polypeptides are water-soluble (except for a very few hydrophobic peptides), while aromatic compounds are lipid-soluble. In open-air operation, based on the principle of "like dissolves like," polypeptides can be extracted using a highly polar extractant (such as water), removing almost all volatile aromatic substances (which are volatile, insoluble in polar solutions, and have a fragrance). The jasmine flower polypeptide extract of this application was obtained through open-air extraction. Unless otherwise specified, any methods or steps described in this application are carried out in a non-enclosed, non-light-protected indoor or outdoor open environment. Extraction of jasmine flowers using a highly polar extractant yields a crude extract, which is a product form of jasmine flower polypeptide extract or a polypeptide mixture. In some preferred embodiments, the jasmine flowers are pulverized before extraction with the extractant. In some preferred embodiments, heating and / or ultrasonic treatment may be used during extraction with the extractant for more complete extraction. Optionally, a step to remove organic solvents (such as an alcohol removal step, which is not required for water extracts) may be added after the extraction process. Therefore, the jasmine flower extract of this application contains little or no jasmine flower essential oil or other ingredients and has no fragrance.

[0007] Those skilled in the art know how to balance the practical needs of purity and total quantity, such as increasing / decreasing extraction or purification steps, replacing different reagents with similar functions in a certain step, or simply repeating a certain step. The core of this application lies in the discovery that a mixture of peptides extracted from jasmine flowers can be used to improve mood-related disorders without relying on the aroma of jasmine flowers, and not in how the peptides are extracted from jasmine flowers. In some embodiments, although most lipid-soluble substances (such as essential oils) or jasmine flower aroma substances have been removed from the crude extract while water-soluble substances (such as peptides) are retained, the inventors may optionally further purify the crude extract through an extraction step (i.e., enriching the peptides). In some embodiments, to further purify the peptides, the peptides are (optionally) precipitated after extraction. In some embodiments, to further purify the peptides, the peptides are (optionally) concentrated and dried after precipitation. In some embodiments, the peptides are optionally identified, for example, by adding deionized water to dissolve the dried product and identifying the peptides using instruments.

[0008] A method for preparing jasmine flower polypeptide extract includes: extracting jasmine flowers once or multiple times with a highly polar extractant to form a crude extract. In some preferred embodiments, further extraction with an organic solvent (e.g., sequentially with solvents of increasing polarity) is performed once or multiple times. In some preferred embodiments, further precipitation with a protein precipitant is performed once or multiple times. In some specific embodiments, the method for preparing jasmine flower polypeptide includes the following steps:

[0009] (1) Pulverize dried jasmine flowers, add water for extraction (preferably using ultrasound) to obtain a crude extract. For example: add 8 times the volume of water solvent to each kilogram of herbal powder, and extract by ultrasound at 40°C for 30 minutes each time in 2-3 batches to obtain jasmine flower extract and filter residue;

[0010] (2) Extraction is carried out by adding an extractant, preferably using extractants with decreasing polarity to increasing polarity sequentially. For example: jasmine flower extract is added to a suspension at a volume ratio of 1:1 with petroleum ether to obtain an extract containing petroleum ether and an extract without petroleum ether; the extract without petroleum ether is then added to a suspension at a volume ratio of 1:1 with ethyl acetate to obtain an extract containing ethyl acetate and an extract without ethyl acetate; the extract without ethyl acetate is then added to a suspension at a volume ratio of 1:1 with n-butanol to obtain an extract containing n-butanol and an extract without n-butanol.

[0011] (3) Add protein or polypeptide precipitant. For example: concentrate the extract containing n-butanol to dryness, suspend the resulting dried product I in deionized water, precipitate it with an organic mixed reagent (n-butanol: dichloromethane = 3:1) at a volume ratio of 4:1, let it stand, take the protein precipitate layer, concentrate it to dryness to obtain dried product II; dissolve dried product II in 10% ACN solution, centrifuge, divide it into 2 equal parts (dried product II-1, dried product II-2), concentrate and dry under nitrogen protection.

[0012] In some preferred embodiments, the obtained polypeptide extract or polypeptide mixture may be further identified. Methods for identifying polypeptides are known in the art, such as mass spectrometry. In some embodiments, the mass spectrometry identification step specifically includes:

[0013] (A) Add 100 μl of ammonium bicarbonate solution (50 mM) to the obtained dried product II-2, sonicate to dissolve, add 5 μl of dithiothreitol (DTT) (1 M, 50 mM ammonium bicarbonate buffer), and incubate at 95 °C for 8 minutes or at 56 °C for 60 minutes. Add 20 μl or 40 μl of iodoacetamide (IAM) (500 mM, 8 M urea solution), and incubate at room temperature in the dark for 30 minutes. Desalt the product using a C18 column, recover the 70% acetonitrile-water solution (containing 0.1% trifluoroacetic acid) fraction, and freeze-dry to obtain dried product III. Dissolve dried product II-1 and dried product III separately in 100 μl of 2% ACN (containing 0.1% trifluoroacetic acid) using sonication, centrifuge at 13300 rpm for 60 minutes, and take 40 μl of each into a liquid chromatography vial.

[0014] (B) Perform Nano-Orbitrap Exploris 480LC-MS / MS analysis:

[0015] The elution was set to gradient elution at a flow rate of 0.4 μl / min. The elution mobile phase composition was: A: 20% H2O (0.1% FA) B: 80% ACN (0.1% FA); 2% B eluted for 0-4 min, 2-26% B eluted for 4-15 min, 26-35% B eluted for 15-30 min, 35-70% B eluted for 30-55 min, 70-95% B eluted for 55-60 min, and 95% B eluted for 60-70 min. The injection volume was 1 μl.

[0016] (C) Mass spectrometry settings:

[0017] Set the positive ion acquisition mode as follows: First-level scan range: m / z 350-2000, fragmentation mode HCD, resolution 120000; Second-level scan range: m / z 50-1500, fragmentation mode HCD, fragmentation energy 30V, acquisition time point: all ions within a 1.5s cycle, resolution 60000; Add FAIMS, set voltage to -45V and -65V.

[0018] (D) Proteome Discoverer parameter settings

[0019] No enzymatic hydrolysis was performed. The mass deviation of the parent ion was 10 ppm, and the mass deviation of the daughter ion was 0.02 Da. PTMs: Carboxymethyl (+58.005 Da), Acetyl (+42.011 Da), Oxidation (+15.995). The ion results were screened according to the molecular weight of 2.5-5 KD and the charge number of 2-4.

[0020] This application demonstrates that jasmine flower polypeptide extract has the function of regulating or improving mood, and preventing or treating diseases or conditions related to impaired neurotransmission (see [link to relevant documentation]). Figure 5-7 It does not rely on the fragrance emitted by jasmine flowers. Attached Figure Description

[0021] Figure 1 Mass spectrometry results of polypeptides in jasmine flower polypeptide extract.

[0022] Figure 2 Liquid phase separation of fractions from jasmine flower polypeptide extract.

[0023] Figure 3 Testing of the binding activity of each fraction separated by liquid phase with cells.

[0024] Figure 4 Both crude extracts of jasmine flower polypeptides (water extract and alcohol extract) have cell-binding activity.

[0025] Figure 5 Results of antidepressant activity of jasmine flower polypeptide extract (shuttle box experiment).

[0026] Figure 6 Results of antidepressant activity of jasmine flower polypeptide extract (forced swimming test).

[0027] Figure 7 Detection and analysis of the interaction between jasmine flower polypeptide extract and dopamine receptor. Detailed Implementation

[0028] This application provides a jasmine flower polypeptide extract and its uses. Specifically, this application provides a jasmine flower polypeptide extract and products containing the extract, such as foods (e.g., foods for special medical purposes (see the "Regulations for the Registration and Management of Foods for Special Medical Purposes"), functional foods, health foods, etc.), beverages, health products, nutritional products, dietary supplements, cosmetics, compositions, drugs, preparations, etc. This application also provides the use of the jasmine flower polypeptide extract and products containing the extract for regulating or improving mood, and for preventing or treating diseases or conditions related to impaired neurotransmission. In some embodiments, the jasmine flower polypeptide extract is an active ingredient in any of the above-mentioned products, such as the sole active ingredient, the main active ingredient, or the directly active ingredient.

[0029] In some embodiments, the jasmine flower polypeptide extract accounts for more than 0.1%, 1%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98%, or 99% of any of the above products, where the percentage is a weight percentage, a molar percentage, or a volume percentage.

[0030] In some embodiments, the proportion of polypeptides in the jasmine polypeptide extract or any of the above products is greater than or at least not less than 0.001%; ​​preferably 0.01% to 100% (0.1% to 1%, 1% to 10%, 10% to 20%, 20% to 30%, 30% to 40%, 40% to 50%, 50% to 60%, 60% to 70%, 70% to 80%, 80% to 90%, 90% to 100%), or 0.01% to 99%, or greater than 0.01%; or greater than 1%, or greater than 10%, or greater than 20%, or greater than 30%, or greater than 40%, or greater than 50%, or greater than 60%, or greater than 70%, or greater than 80%, or greater than 90%; the percentage is expressed as mass / volume concentration (ratio) or mass ratio or molar (number) ratio.

[0031] Jasmine flower polypeptide extract can be a crude extract obtained by extracting jasmine flowers with a highly polar extractant (to remove volatile aromatic substances). In some embodiments, the highly polar (or very polar) extractant is preferably an extractant with a polarity close to that of water or a hydrophilic solvent, such as water, alcohols (preferably lower alcohols with 1 to 4 carbon atoms), or alcohol-water mixtures (preferably with more than 50% alcohol); in some preferred embodiments, the highly polar extractant is water, methanol, ethanol, methanol-water, or ethanol-water. In some embodiments, the amount of highly polar extractant added is equal to or similar to the amount of jasmine flowers used, or the amount of highly polar extractant added is more than 2, 3, 4, 5, 6, 7, 8, 9, or 10 times the amount of jasmine flowers used; for example, 2-4 times, 2-6 times, 2-8 times, 2-10 times, 4-6 times, 4-8 times, 4-10 times, 6-8 times, 6-10 times, or 8-10 times. In some embodiments, the jasmine flower polypeptide extract is an extract soluble in polar solvents, such as an alcohol extract or a water extract. When the extractant contains alcohol, the extract may optionally be concentrated to remove the alcohol. If the extractant is water, the alcohol removal step is unnecessary. In some preferred embodiments, the highly polar extractant is water, an alcohol with 1-2 or 1-3 carbon atoms, or a mixture of water and an alcohol with 1-2 or 1-3 carbon atoms. In some embodiments, the volume ratio of alcohol in the mixture of water and alcohol with 1-2 or 1-3 carbon atoms is ≥50%.

[0032] The jasmine flower polypeptide extract can also be an enriched product formed by further extraction of the crude extract (this is an optional step). In some embodiments, the crude extract can be further extracted with an organic solvent. In some embodiments, a low- to medium-polarity organic solvent can be used for extraction. The low- to medium-polarity solvent is preferably an organic solvent with a polarity lower than acetone, such as petroleum ether, n-hexane, ethyl acetate, or n-butanol. If it is desired to particularly enrich polypeptide components with a polarity between n-butanol and water in the crude extract, a high- to medium-polarity organic solvent, i.e., a reagent with a polarity approximately between acetone and n-butanol, can also be used for extraction, such as acetone (preferably cold acetone) or n-butanol. Those skilled in the art can select the extractant as needed. In some embodiments, the extraction step can be repeated once, twice, or more. In some preferred embodiments, extraction is performed sequentially using organic solvents with gradually increasing polarity, such as sequentially using the lowest polarity extractant, a lower polarity extractant, and a higher polarity extractant for one, two, or more extractions. In some embodiments, extraction is performed once using petroleum ether, ethyl acetate, and n-butanol in sequence; or once using ethyl acetate and n-butanol in sequence. In some embodiments, the same organic solvent may be used for repeated extractions. The extraction step further removes low-polarity compounds (such as pigments) from the crude extract and yields an enriched product, which is also a product form of jasmine flower polypeptide extract.

[0033] The jasmine flower polypeptide extract can also be further purified by precipitating the enriched product after extraction (this is an optional step). In some embodiments, the product is further precipitated with a protein precipitant. In some embodiments, the protein precipitant may be an organic solvent, acid, alkaloid, or salt. In some embodiments, the protein precipitant includes methanol, ethanol, acetone (preferably cold acetone), dimethyl sulfoxide, acetonitrile, an aqueous solution of trichloroacetic acid, picric acid, ammonium sulfate, sodium sulfate, zinc sulfate, a mixture of n-butanol and dichloromethane, or a mixture of n-butanol and trichloromethane. The purified product obtained after precipitation is also a product form of the jasmine flower polypeptide extract.

[0034] Jasmine flower polypeptide extract can also be a dried product obtained by further concentrating and drying the purified product after precipitation (this is an optional step). Therefore, the dried purified product (or dried product) is also a product form of jasmine flower polypeptide extract.

[0035] The concentration or proportion of jasmine flower polypeptide extract in the pharmaceutical composition or nutritional composition is at least 0.001%; ​​preferably 0.01% to 100% (0.1% to 1%, 1% to 10%, 10% to 20%, 20% to 30%, 30% to 40%, 40% to 50%, 50% to 60%, 60% to 70%, 70% to 80%, 80% to 90%, 90% to 100%), more preferably 25%; the percentage is expressed as mass / volume concentration (ratio) or mass ratio or molar (number) ratio.

[0036] In some implementations, jasmine flowers can be fresh flowers, dried flowers, or processed flowers.

[0037] The compositions of this application (such as pharmaceutical or nutritional compositions) can be in various oral or non-oral dosage forms, such as injections, lyophilized powders for injection, aerosols, creams, drops, ointments, or pastes, capsules, effervescent tablets, chewable tablets, lozenges, granules, ointments, syrups, nasal drops, oil-water mixtures, suspensions, (rubs, lotions, granules, sprays, or patches), pills, suppositories, emulsions, or tablets. In the case of formulation, they are prepared using commonly used fillers, expanders, binders, wetting agents, disintegrants, surfactants, and other diluents or excipients.

[0038] In some embodiments, compositions (such as pharmaceutical compositions or nutritional compositions) are used to produce 50 to 500 mg of jasmine flower polypeptide extract per dose unit. In some embodiments, the nutritional composition comprises a protein source, a carbohydrate source, a lipid source, and / or minerals (such as micronutrients and trace elements).

[0039] In some embodiments, the pharmaceutical composition or nutritional composition is formulated or designed with one or more other pharmaceuticals (or adjunct therapeutic agents) for continuous, simultaneous, sequential, alternating, spaced-out, or single administration.

[0040] In some embodiments, a purification and / or identification method for jasmine flower polypeptides according to this application includes the following steps: a) preparing a jasmine flower extract; b) optionally, adding an extractant A to the jasmine flower extract to obtain an extract containing extractant A and an extract without extractant A; c) optionally, adding an alcohol extractant B to the extract without extractant A to obtain an extract containing alcohol extractant B and an extract without alcohol extractant B; d) optionally, concentrating the extract containing alcohol extractant B to obtain a dried product I, and then contacting the dried product I with a mixed extract C to obtain the jasmine flower extract. The precipitate is concentrated to obtain dried product II; e) Optionally, the dried product II is divided into two equal parts, dried product II-1 and dried product II-2; f) Optionally, a urea solution containing dried product II-2 is mixed with dithiothreitol and reacted to obtain precursor 1; g) Optionally, precursor 1 is contacted with iodoacetamide to perform an alkylation reaction to obtain precursor 2; h) Optionally, dithiothreitol is added to precursor 2, and after the reaction is completed, the salt is removed and dried to obtain dried product III; i) Optionally, the dried product II-1 and the dried product III are analyzed for peptide detection using mass spectrometry.

[0041] In a preferred embodiment, the mass spectrometry data of the dried product II-1 is used as an internal control for the mass spectrometry data of the disulfide-bonded alkylated polypeptide of the dried product III. In a preferred embodiment, the extractant A is selected from at least one of petroleum ether, ethyl acetate, cyclohexane, and dichloromethane. Preferably, the volume ratio of extractant A to the jasmine flower extract is 1:1. In a preferred embodiment, the alcohol extractant B is n-butanol. Preferably, the volume ratio of extractant B to the extract of extractant A is 1:1. In a preferred embodiment, the mixed extract C is characterized by being composed of the alcohol extractant n-butanol and dichloromethane or chloroform. Preferably, the volume ratio of the alcohol extractant to dichloromethane in the mixed extract C is 3-5:1. Preferably, the extractant is selected from n-butanol. Preferably, the volume ratio of the mixed extract C to the solution of the dried product I is 1:4. In a preferred embodiment, in step f), the urea solution containing the dried product II-2 is mixed with dithiothreitol and reacted at 50–70°C for 0.5–1.5 hours. Preferably, in step f), the urea solution containing the dried product II-2 is mixed with dithiothreitol and reacted under water bath conditions. In a preferred embodiment, in step g), the precursor 1 is reacted with iodoacetamide at room temperature for 3–5 hours. In a preferred embodiment, in step h), dithiothreitol is added to the precursor 2, and the reaction is carried out at room temperature for 0.5–1.5 hours. In a preferred embodiment, the jasmine flower extract is an ethanol-water solution of a solid jasmine flower sample; preferably, the volume ratio of ethanol in the ethanol-water solution is 70%. In a preferred embodiment, the mass spectrometry technique includes Nano-Orbitrap Exploris 480LC-MS / MS loaded with ion mobility FAIMS Pro. Preferably, the mass spectrometry technique is Nano-Orbitrap Exploris 480LC-MS / MS.

[0042] In a preferred embodiment, the Nano-Orbitrap Exploris 480LC-MS / MS includes at least the following steps: (a) isocratic elution using a Trap column, wherein the elution mobile phase consists of phase A of 75%–98% H2O and phase B of 2%–25% acetonitrile, at a flow rate of 0.1–1 μl / min; (b) gradient elution using a Nano column, wherein the flow rate is 0.5 μl / min, and the elution mobile phase consists of phase A of 75%–98% H2O and phase B of 2%–25% acetonitrile; followed by the following sequential elutions: 2% B elution for 0–4 min, 2–26% B elution for 4–15 min, 26–35% B elution for 15–30 min, 35–70% B elution for 30–55 min, 70–95% B elution for 55–60 min, and 95% B elution for 60–70 min. Preferably, the test injection volume is 1 μl.

[0043] Terminology Definition

[0044] The abbreviations used herein have their conventional meanings in the fields of chemistry and biology. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains; the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of this application; the terms “comprising” and “having”, and any variations thereof, in the specification, claims, and foregoing description of the drawings are intended to cover non-exclusive inclusion.

[0045] "Polypeptide extract" has a clear meaning even though it is a mixture, such as sea cucumber polypeptide extract in patent CN115054581B and milk polypeptide extract in CN109380728B; the separation or extraction of polypeptides is a conventional technique.

[0046] "Jasmine flower polypeptide extract" refers to a mixture of active peptides with a molecular weight ≤10,000 Da that are selectively extracted from jasmine flowers. In the jasmine flower polypeptide extract of this application, the polypeptide is the main or core component of the extract, and is also the direct active ingredient, sole active ingredient, or main active ingredient in the composition for regulating or improving mood and preventing or treating diseases or conditions related to impaired neurotransmission.

[0047] "Not dependent on aroma or contains no aroma" means that it contains little or no jasmine aroma components or volatile aromatic substances. Those skilled in the art know how to remove volatile substances, such as by distillation to remove volatile aromatic compounds, or by dissolving aromatic compounds in volatile organic solvents (such as hexane or petroleum ether), and other conventional methods.

[0048] "Contains no or very little" means that a certain component has been effectively and as much as possible removed during the preparation process.

[0049] "Regulating or improving emotions" refers to regulating or improving one or more of the following emotional problems: (1) rapid mood changes; sudden sadness, anger or rage; (2) persistent and severe anger and anxiety; (3) anxiety and tension; (4) depression and despair; (5) lack of interest in daily life; (6) fatigue or low energy; (7) lack of concentration; (8) changes in appetite; (9) sleep disorders; (10) difficulty in controlling emotions; (11) nervousness; (12) memory difficulties; (13) melancholy, so as to restore the subject to a positive, optimistic and healthy state of mind and body.

[0050] "Mood disorder" is a disturbance of mood, including persistent excessive sadness, excessive joy, or both. Mood disorders can occur in children and adolescents (such as childhood and adolescent depression, childhood and adolescent bipolar disorder, etc.).

[0051] "Diseases or conditions associated with impaired neurotransmission" include neurocognitive impairment, depression (such as unipolar depression, bipolar depression, acute depression, chronic depression, subchronic depression, psychotic depression, postpartum depression, menopausal depression, mild depression, or severe depression), anxiety disorders (such as premenstrual anxiety syndrome, social anxiety disorder), aggression, attention deficit disorder (ADS), seasonal affective disorder, fibromyalgia syndrome, chronic fatigue, sleep disorders (i.e., insomnia), post-traumatic stress disorder, panic disorder, obsessive-compulsive disorder, ADHD, nervousness, migraine / primary headache and general pain, vomiting, bulimia, anorexia nervosa, eating disorders, gastrointestinal disorders, psychosomatic exhaustion syndrome, irritability, and fatigue.

[0052] "Depressive disorder" is characterized by persistent and severe sadness, decreased interest in activities or feelings of pleasure, and impaired social functioning. The exact cause is unclear, but may involve genetics, changes in neurotransmitter levels, alterations in neuroendocrine function, and sociopsychological factors. Treatment typically includes medication, psychotherapy, or a combination of both, and sometimes includes electroconvulsive therapy or rapid transcranial magnetic stimulation (rTMS).

[0053] "Depression" can refer to any of several depressive disorders. In the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition (DSM-IV), it is classified according to specific symptoms: depression (such as major depressive disorder), persistent depressive disorder (dysthymia), and other specific or non-specific depressive disorders. Based on etiology, it can be classified as: premenstrual dysfunctional disorders, physical condition-related depressive disorders, and substance / medical-related depressive disorders. Depression often manifests as mood disturbance symptoms such as fatigue, apathy, loss of interest, low mood, and slowed reaction time, and is often accompanied by insomnia.

[0054] "Polar solvent" is a common term in this field. The polarity order of commonly used solvents is: water (highest) > formamide > trifluoroacetic acid > DMSO > acetonitrile > DMF > hexamethylphosphoramide > methanol > acetic acid > ethanol > isopropanol > pyridine > tetramethylethylenediamine > acetone > triethylamine > n-butanol > dioxane > tetrahydrofuran > methyl formate > tributylamine > methyl ethyl ketone > ethyl acetate > chloroform > trioctylamine > dimethyl carbonate > diethyl ether > isopropyl ether > n-butyl ether > trichloroethylene > diphenyl ether > dichloromethane > dichloroethane > benzene > toluene > carbon tetrachloride > carbon disulfide > cyclohexane > hexane > kerosene (petroleum ether) (lowest). Water is an example of a highly polar solvent.

[0055] "Extractant" refers to any reagent used in the preparation of jasmine flower polypeptide extracts (such as extraction-precipitation). In some embodiments, the extractant is a physiologically acceptable or edible solvent. Edible means a physiologically acceptable solvent that can be absorbed by the human body.

[0056] "Extraction," also known as solvent extraction or liquid-liquid extraction, is a unit operation that uses the different solubilities of components in a solvent to separate a mixture.

[0057] "Precipitation" refers to the steps that promote the aggregation and deposition of proteins (or polypeptides). In the extraction of polypeptides from plant tissues, organic solvent precipitation is generally used.

[0058] As used herein, the term "pharmaceutically acceptable" means approved for use in animals, preferably in humans, by regulatory authorities such as the CFDA (China), EMEA (Europe), and / or FDA (US) and / or any other national regulatory authority. When referring to compositions, excipients, carriers, diluents, and reagents, the terms "pharmaceutically acceptable," "physiologically tolerable," and their synonyms are used interchangeably and indicate substances that can be administered to subjects or patients, particularly human patients. For example, "pharmaceutically acceptable salt" includes both acid and base addition salts.

[0059] Those skilled in the art can determine the edible or drug dosage of the active ingredient in the food or pharmaceutical composition of this application based on known techniques. For example, the daily dosage can be 0.1-5000 mg / kg / day, more specifically 50-500 mg / kg / day, but is not limited thereto. The dosage may vary depending on various factors such as the age and physical condition of the person in need, the presence of complications, etc.

[0060] There are no particular restrictions on hydrophilic organic solvents; they can be selected appropriately according to the purpose. Examples include: lower alcohols with 1 to 5 carbon atoms such as methanol, ethanol, propanol, and isopropanol; lower aliphatic ketones such as acetone and methyl ethyl ketone; and polyols with 2 to 5 carbon atoms such as 1,3-butanediol, propylene glycol, and glycerol. They can be used alone or in combination with two or more.

[0061] There are no particular limitations on the mixed solvent of water and the hydrophilic solvent, and appropriate selection can be made according to the purpose. When using the lower alcohol as the hydrophilic solvent, it is preferable to use 1 to 90 parts by mass of the hydrophilic solvent relative to 10 parts by mass of water. When using the lower aliphatic ketone as the hydrophilic solvent, it is preferable to use 1 to 40 parts by mass of the hydrophilic solvent relative to 10 parts by mass of water. When using a polyol as the hydrophilic solvent, it is preferable to use 1 to 90 parts by mass of the hydrophilic solvent relative to 10 parts by mass of water. They can be used alone or in combination with two or more.

[0062] In some embodiments, this application uses a mixed organic solvent to precipitate and extract peptides. The mixed organic solvent is a mixture of dichloromethane / trichloromethane and n-butanol, and the volume ratio of n-butanol to dichloromethane / trichloromethane can be 3:1, 4:1 or 5:1.

[0063] "Extract" can refer to any substance extracted from natural products, but after using a highly polar extractant, the jasmine flower polypeptide extract of this application no longer contains or contains almost no essential oil substances, and is also different from the components in jasmine leaves.

[0064] "Direct active ingredient," also known as effective ingredient, refers to an ingredient that has a direct causal relationship with the outcome (such as the effect of improving mood or treating disease) relative to auxiliary or secondary ingredients.

[0065] "Sole active ingredient" or "major active ingredient" means that its content accounts for more than 50%, 60%, 70%, 80%, 90%, or 100% of all active ingredients, and the percentage is a mass ratio or molar ratio; or "as the sole active ingredient or major active ingredient" means that its contribution to the result (such as the effect of improving mood or treating disease) is more than 50%, 60%, 70%, 80%, 90%, or 100%.

[0066] As used in this article, “individual,” “subject,” or “patient” refers to humans and non-human animals, animals, or mammals, including but not limited to humans, monkeys, farm animals (such as sheep, pigs, cattle, and horses); pet animals (such as dogs, cats, monkeys, etc.); and laboratory animals (such as mice, rats, rabbits, chickens, and non-human primates).

[0067] The term "excipient" refers to any pharmaceutically acceptable additive, carrier, diluent, adjuvant, or other component other than the active pharmaceutical ingredient (API), which is typically included in the formulation and / or administered to the patient.

[0068] In the description of this application, unless otherwise stated, "a plurality of" means two or more. References to "one or more" herein indicate the presence of at least one of the elements; a plurality of such elements may be present unless otherwise expressly specified.

[0069] In the description of this application, the term "and / or" is merely a description of the relationship between related objects, indicating that three relationships can exist. For example, A and / or B can represent: A existing alone, A and B existing simultaneously, or B existing alone. Additionally, the character " / " in this document generally indicates that the preceding and following related objects have an "or" relationship.

[0070] In this specification and the appended claims, unless the context clearly specifies otherwise, the singular forms, including the singular forms “a,” “an,” and “the,” specifically cover the plural indicators of the terms they refer to. Additionally, as used herein, unless otherwise specifically indicated, the word “or” means “inclusive” rather than “exclusive” in the sense of “and / or.”

[0071] As used herein, references to the numerical range of variables are intended to convey that this application can be implemented with variables equal to any value within that range. Thus, for intrinsically discontinuous variables, the variable may be equal to any integer value within the numerical range, including the endpoints of the range. Similarly, for intrinsically continuous variables, the variable may be equal to any real value within the numerical range, including the endpoints of the range. For example, a variable described as having a value between 0 and 2 may be 0, 1, or 2 for intrinsically discontinuous variables, and may be 0.0, 0.1, 0.01, 0.001, or any other real value for intrinsically continuous variables.

[0072] As used herein, the term “about” will be understood by those skilled in the art and will vary to some extent depending on the context in which it is used. If the use of this term is unclear to those skilled in the art, then in the context of its use, “about” will mean a value within the range of the listed value plus or minus 10%.

[0073] "Container" refers to a vessel for holding things, such as a box, basket, enamel basin, glass, etc.; a storage container (such as a box, can, jar) or a shaped or soft and non-shaped covering material used to package or load things (see Chinese Dictionary), emphasizing the functions of packaging and storage.

[0074] "Device" refers to a complex object in a machine, instrument, or equipment that has a certain independent function (see the Chinese Dictionary). In this application, "device" can release drugs or has drug delivery function or potential drug delivery capability, focusing on the function of administration.

[0075] "Administering" the jasmine flower polypeptide extract of this application to a subject includes any route by which they are introduced into or delivered to the subject to perform their intended function. This can be performed by any suitable route, including but not limited to oral, intraocular, intranasal, parenteral (via intravenous, intramuscular, intraperitoneal, or subcutaneous), systemic, local, non-invasive, or ablation-free administration.

[0076] Regarding "single administration", taking drug administration as an example, it means that only one method of administration is used at each stage of the entire medication process, while the method of administration can be changed at different stages of the medication process (but there is no alternation).

[0077] Regarding "simultaneous administration," taking drug administration as an example, it refers to administering at least two active ingredients via the same or different routes (such as oral and injection) at the same or substantially the same time; or administering drugs and performing surgery at the same or substantially the same time; or administering drugs and applying a treatment device at the same or substantially the same time.

[0078] Regarding "continuous administration," taking drug administration as an example, it refers to maintaining a stable drug concentration in the body through continuous infusion of drugs (such as intravenous drip).

[0079] Regarding "interval administration," taking drug administration as an example, it refers to the periodic fluctuation of drug concentration by administering the drug in multiple doses.

[0080] Regarding "sequential administration," taking drug administration as an example, it refers to administering at least two active ingredients at different times during treatment, with the same or different routes of administration, using different drugs or treatment regimens in sequence, one after another, usually starting the next stage only after the previous stage is completed.

[0081] Regarding "alternating use", taking drug administration as an example, it refers to the periodic rotation of two or more different drugs or treatment regimens.

[0082] As used herein, the term "effective dose" refers to a dose sufficient to suppress, stop, improve, alleviate, delay, or produce improvement in the symptoms of one or more treatable conditions. For example, a dose that induces prevention or reduction of symptoms associated with depression. The amount of medication administered to the subject will depend on the type and severity of the illness and the individual's characteristics, such as general health, age, sex, weight, degree of depression, rate of depression progression, and tolerance to the medication. A qualified physician will be able to determine the appropriate dosage based on these and other factors.

[0083] "Mixture," "(peptide) composition," "formulation," "pharmaceutical composition," or "nutritional composition" refers to a mixture comprising two or more compounds, elements, or molecules. In some respects, these terms can be used to refer to mixtures comprising one or more active agents (active ingredients) with a carrier or other excipients, which can take almost any physical form, including solid, liquid (e.g., solution), or gas.

[0084] The term "dosage form" can include one or more formulations or compositions provided in a form intended for administration to a subject. For example, a composition can be in the form of a non-oral administration such as an injection, or an oral administration such as a tablet, capsule, granule, powder, syrup, etc., and may contain excipients, binders, extenders, dispersants, or preservatives permitted in the formulation.

[0085] The products of this application can be formulated as health foods such as tablets, capsules, powders, granules, and health drinks (including solutions and suspensions), or as dairy products, seasonings, processed foods, desserts, and confectionery (e.g., chewing gum, candy, jelly), such as soft drinks, tea drinks, yogurt, or lactic acid bacteria drinks. The food products of this application may also contain excipients, binders, coating agents, disintegrants, additives, etc., permitted as food products.

[0086] The products of this application may be mixed with other physiologically active ingredients, such as minerals; vitamins such as vitamin E, vitamin C, and vitamin A; nutrients; flavorings; colorings, and other additives, as needed. Any of the above-mentioned additives may be those commonly used in food and beverage products.

[0087] As used in this document, the term "prevention" refers to proactive preventative measures and behaviors taken before the onset of the disease or the appearance of its related symptoms, with the aim of effectively and / or actively avoiding the occurrence of the disease or its related symptoms. The terms "treatment," "control," "suppression," "relief," "reduction," "prevention," or "mitigation," and similar descriptions, all refer to measures to cure the disease and / or to hinder its progression. Therefore, the means described by these terms are all aimed at preventing or alleviating the target symptoms or disorders, or even eliminating or reversing them. Treatment also includes preventative treatment and palliative care. "Relief" can refer to complete or partial relief. For example, in some implementations, "treatment" does not necessarily require 100% elimination or reversal of depressive mood or symptoms.

[0088] The term "predisposition to depression (individual)" used in this article can refer to a situation where depressive mood has already occurred but has not yet developed into clinical depression; it can also refer to a person who is predicted or identified by an authoritative institution or professional physician as having a predisposition to depression or being at high risk; it can also refer to an individual with a family history of mental illness; it can also refer to an individual who has been exposed to a lot of negative information or significant stress for a long time but lacks opportunities for understanding or communication; it can also refer to an individual who may experience relapses even after taking medication or other depression interventions; and it can also refer to a situation where the individual will experience depressive mood or clinical depression if no medication or other depression treatment (prevention) interventions are used.

[0089] The jasmine polypeptide extract of this application can be administered to a patient alone or as part of a pharmaceutical composition in a therapeutically effective amount. Various non-limiting methods of administering the polypeptide and related compositions to a patient include oral, parenteral (intravenous, intramuscular, or subcutaneous), topical (powder, ointment, or drops), or as a sublingual or spray formulation. Additionally, the polypeptide or composition containing the active substance can be administered once, for example by bolus injection, or multiple times, for example by a series of tablets, or delivered substantially uniformly over a period of time, for example using transdermal delivery. It should be noted that the dosage of the substance may vary over time.

[0090] The jasmine polypeptide extract of this application can be administered alone or in combination with other pharmaceutically active substances. These other pharmaceutically active substances may be intended to treat the same or different diseases or conditions as those of the substances in this application. If a patient is receiving or is receiving multiple pharmaceutically active substances, these substances may be administered simultaneously or sequentially. For example, in the case of tablets, the active substances may be present in a single tablet or in separate tablets, which may be administered once or sequentially in any order. Furthermore, it should be recognized that the compositions may be in different forms. For example, one or more substances may be delivered via tablets, while another may be administered orally via injection or syrup. This application includes all combinations, methods of delivery, and sequences of administration.

[0091] "Pharmaceutical composition" refers to a compound suitable for administration in pharmaceutical or veterinary use. It should be understood that determining the appropriate dosage form, dosage, and route of administration for a specific patient is within the level of a person of ordinary skill in the pharmaceutical and medical field. A pharmaceutical composition may include one or more pharmaceutically acceptable excipients and a therapeutically effective amount of a jasmine flower polypeptide extract.

[0092] This application also relates to a kit or device that may include one or more separate pharmaceutical compositions, such as the jasmine flower polypeptide extract of this application and a second pharmaceutical substance. The kit or device may also include containers, such as separate bottles or separate blister packs. Blister packs are known and widely used in the packaging industry for packaging unit dosage forms of pharmaceuticals (tablets, capsules, etc.). Other examples of containers include syringes, boxes, and bags. The container contains the jasmine flower polypeptide extract of this application and / or the second pharmaceutical substance.

[0093] The peptides and peptide extracts of this application can also benefit from a variety of drug delivery systems, including timed release, delayed release or sustained release drug delivery systems, or novel drug delivery systems (such as those using nanocarriers).

[0094] When implementing the method of this application, the jasmine flower polypeptide extract of this application is administered at a therapeutically effective amount. Procedural experiments in clinical trials can determine the specific range that produces the best therapeutic effect.

[0095] The polypeptide compositions, polypeptide extracts, or pharmaceutical compositions of this application may be used for first-line, second-line, or third-line treatment; or may be used to treat patients who have received unsatisfactory previous treatments or who have developed drug resistance.

[0096] The polypeptide compositions, polypeptide extracts, or pharmaceutical compositions of this application may be used for the prevention and / or treatment of psychiatric disorders or conditions, particularly depression, including severe depression and depressive disorder in bipolar disorder, and psychiatric conditions requiring medication, including schizophrenia, psychosis in bipolar disorder, and irritant drug poisoning.

[0097] The polypeptide compositions, polypeptide extracts, or pharmaceutical compositions of this application may be used in combination with other treatment methods (including drug therapy, psychotherapy, and electroconvulsive therapy).

[0098] A method for preparing jasmine flower extract involves extracting jasmine flowers with a highly polar extractant to form a crude extract. In some embodiments, the highly polar extractant can be water, alcohols (such as methanol or ethanol), or an alcohol-water mixture (more than 50% alcohol).

[0099] This application further removes low-polarity compounds (such as pigments) from the crude extract described above, including sequentially using the extractant with the lowest polarity, the extractant with relatively low polarity, and the extractant with relatively high polarity (extractants with gradually increasing polarity). In some embodiments, petroleum ether and ethyl acetate are used sequentially for extraction.

[0100] In order to enrich the polypeptide components (with polarity between n-butanol and water) in the crude extract, this application uses medium to high polarity organic solvents, i.e., reagents with polarity between acetone (containing) and n-butanol (containing), such as n-butanol and acetone (preferably cold acetone).

[0101] To further precipitate polypeptide components (such as proteins or amino acids), this application uses organic solvents (such as a mixture of n-butanol and dichloromethane) or salts to precipitate the polypeptides. These extraction methods are all conventional techniques in the art.

[0102] The polypeptides in the jasmine flower polypeptide extract prepared in this application have a molecular weight of 50-5000 Daltons or contain 2-100 amino acids.

[0103] Example 1: Preparation and Detection of Jasmine Flower Peptide Extract (Example 1)

[0104] ① Preparation of jasmine flower polypeptide extract: 1.0 kg of dried jasmine flowers were crushed and added to 8 L of water. The mixture was ultrasonically extracted twice at 35 °C for 30 min each time. The extract was filtered to obtain jasmine flower polypeptide extract and jasmine flower residue. Petroleum ether was added to the jasmine flower polypeptide extract at a volume ratio of 1:1 for extraction. The extract without petroleum ether was then extracted with ethyl acetate. The extract without ethyl acetate was extracted with n-butanol at a volume ratio of 1:1. The extract containing n-butanol was concentrated to dryness to obtain dried product Ia. Dried product Ia was suspended in deionized water and precipitated at a volume ratio of deionized water to organic reagent (n-butanol: dichloromethane = 3:1) of 4:1. The precipitate was collected and concentrated to dryness to obtain dried product IIa. Dried product IIa was dissolved in 10% ACN, centrifuged, and the supernatant was collected. Optionally, the supernatant was divided into two equal parts (dried product II-1a and dried product II-2a) and freeze-dried.

[0105] Optionally, ②, alkylation treatment of jasmine flower polypeptide extract samples facilitates mass spectrometry detection: Add 100 μl of 50 mM ammonium bicarbonate solution to dried product II-2a, sonicate until dissolved, add 5 μl of dithiothreitol (DTT) (1 M, 50 mM ammonium bicarbonate solution), incubate at 95℃ for 8 min, add 20 μl of iodoacetamide (IAM) (500 mM, 6 M urea solution), and place at room temperature in the dark for 30 min. After C18 desalting, recover the 80% acetonitrile-water solution (containing 0.1% trifluoroacetic acid) fraction, freeze-dry, and obtain dried product IIIa. Optionally, dissolve dried products II-1a and IIIa separately in 100 μl of 10% ACN using sonication, centrifuge at 133000 rpm for 60 min, and take 40 μl of each into a liquid chromatography vial.

[0106] Example 2: Preparation and Detection of Jasmine Flower Peptide Extract

[0107] ① Preparation of jasmine flower polypeptide extract: 1.0 kg of dried jasmine flowers were crushed and added to 8 L of 70% ethanol aqueous solution. The mixture was ultrasonically extracted twice at 40℃ for 1 h each time. The extract was filtered to obtain jasmine flower polypeptide ethanol extract and jasmine flower residue. The jasmine flower polypeptide ethanol extract was concentrated to the alcohol-free volume. Deionized water was added to suspend the extract, and ethyl acetate was added to the suspension at a volume ratio of 1:1. The extract without ethyl acetate was taken and n-butanol was added at a volume ratio of 1:1. The extract containing n-butanol was concentrated to dryness to obtain dried product Ib. Dried product Ib was suspended in deionized water and extracted at a volume ratio of deionized water:organic reagent (n-butanol:dichloromethane = 4:1) of 3:1. The protein precipitate was collected and concentrated to dryness to obtain dried product IIb. Dried product IIb was centrifuged with 10% ACN. Optionally, the supernatant was divided into two equal parts (dried product II-1b and dried product II-2b) and lyophilized.

[0108] Optionally, ② alkylation treatment of jasmine flower polypeptide extract samples facilitates mass spectrometry detection: Add 100 μl of 50 mM ammonium bicarbonate solution to dried product II-2b, sonicate until dissolved, add 5 μl of dithiothreitol (DTT) (1 M, 50 mM ammonium bicarbonate solution), incubate in a water bath at 56 °C for 60 min, add 40 μl of iodoacetamide (IAM) (500 mM, 6 M urea solution), and place at room temperature in the dark for 30 min. After C18 desalting treatment, recover the 80% acetonitrile-water solution (containing 0.1% trifluoroacetic acid) fraction, freeze-dry, and obtain dried product IIIb; Optionally, dissolve dried product II-1b and dried product IIIb separately with 100 μl of 10% ACN using sonication, centrifuge at 133000 rpm for 60 min, and take 40 μl of each into a liquid chromatography vial.

[0109] Example 3: Preparation and Detection of Jasmine Flower Peptide Extract

[0110] ① Preparation of jasmine flower polypeptide extract (i.e., enrichment of polypeptide compounds in jasmine flowers): 0.5 kg of dried jasmine flowers were crushed and added to 4 L of 70% ethanol aqueous solution. The mixture was filtered to obtain jasmine flower polypeptide ethanol extract and jasmine flower residue. The jasmine flower polypeptide extract was concentrated to an alcohol-free volume and suspended in deionized water. Ethyl acetate was added to the suspension at a volume ratio of 1:1 to obtain an extract containing ethyl acetate and an extract without ethyl acetate. The extract without ethyl acetate was then mixed with n-butanol at a volume ratio of 1:1. An extract containing n-butanol and an extract without n-butanol were obtained; the extract containing n-butanol was concentrated to dryness to obtain dried product Ib; dried product Ib was suspended in deionized water to obtain dried product Ic; dried product Ic was suspended in deionized water and extracted with an organic reagent (n-butanol: dichloromethane = 5:1, volume ratio) to obtain the protein precipitate, and concentrated to dryness to obtain dried product IIc; dried product IIc was centrifuged with 10% ACN, and the supernatant was divided into two equal parts (dried product II-1c and dried product II-2c), and lyophilized.

[0111] Optionally, ② alkylation treatment of jasmine flower polypeptide extract samples facilitates mass spectrometry detection: Add 100 μl of 50 mM ammonium bicarbonate solution to dried product II-2c, sonicate until dissolved, add 5 μl of dithiothreitol (DTT) (1 M, 50 mM ammonium bicarbonate solution), incubate at 95℃ for 8 min, add 40 μl of iodoacetamide (IAM) (500 mM, 6 M urea solution), and place at room temperature in the dark for 30 min. After C18 desalting treatment, recover the 80% acetonitrile-water solution (containing 0.1% trifluoroacetic acid) fraction, freeze-dry to obtain dried product IIIc; Dissolve dried product II-1c and dried product IIIc separately with 100 μl of 10% ACN using sonication, centrifuge at 133000 rpm for 60 min, and take 40 μl of each into liquid chromatography vials.

[0112] Example 4: Mass Spectrometry Detection Example

[0113] The alkylated jasmine flower polypeptide extract sample from Example 3 was analyzed by LC-MS / MS: ① LC-MS settings: flow rate 0.4 μl / min, elution mobile phase composition A: 20% H2O (0.1% FA) B: 80% ACN (0.1% FA); 2% B elution 0-4 min, 2-26% B elution 4-15 min, 26-35% B elution 15-30 min, 35-70% B elution 30-55 min, 70-95% B elution 55-60 min, 95% B elution B. Elution for 60-70 minutes, injection volume 2 μl; ② Mass spectrometry settings: Set positive ion acquisition mode, primary scan range m / z: 350-2000, fragmentation mode HCD, resolution 120000; secondary scan range 50-1500, fragmentation mode HCD, fragmentation energy 30V, acquisition time point 1.5s for all ions within the cycle, resolution 60000; Add FAIMS, set voltage -45V and -65V; ③ ProteomeDiscoverer library search parameters: no enzymatic digestion, parent ion mass deviation 10ppm, daughter ion mass deviation 0.02Da; PTMs: Carboxymethyl (+58.005Da), Acetyl (+42.011Da), Oxidation (+15.995), ion results were screened according to molecular weight 2.5-5KD and charge number 2-4, results are shown in [link to results]. Figure 1 .

[0114] Example 5: Fractionation and Activity Determination of Jasmine Flower Peptide Extract

[0115] 1) Sample separation: The jasmine flower polypeptide extract sample (the dried purified product from Example 3) was dissolved in 3 ml of 5% methanol (MeOH). After sonication at room temperature for 30 min, the solution was filtered through a 0.22 μM membrane. The resulting filtrate was then transferred to a liquid chromatography vial and subjected to liquid chromatography (using a C18 OBD Prep Column). Separation was performed using a 5 μM, 10 mm × 250 nm concentration. The sample loading volume was 500 μl. Mobile phase A was ultrapure water containing 0.1% formic acid, and mobile phase B was methanol. A gradient elution program was used according to Table 1. The detection wavelength was 254 nm, and the flow rate was 7 mL / min. The separation results are shown below. Figure 2 As shown. The fractionally collected fractions were then recovered, freeze-dried, and subjected to cell viability testing.

[0116] Table 1 Gradient elution program

[0117]

[0118] 2) Cell Culture: 1> Resuspend the pre-cultured cell line into a cell suspension, determine the concentration, dilute to a suitable concentration, mix well, and place in the sample well; 2> Select an appropriate number of 96-well plates as needed to construct a glutamate-glycine injury model (Model), evenly seed the cells, with 3000-5000 cells per well, and leave 6 replicates for each test concentration; 3> After labeling the wells, use a 100µl pipette to aspirate the cell suspension from the sample well, trying to aspirate from the center area, and add it to the 96-well plate. Incubate at 37℃ until the cells adhere; 4> After cell adhesion, add fractions 1-8 (J1-J8) at a concentration of 250µg / ml, and continue incubation; 5> After completing the above model, gently dry the culture medium containing the drug / GLU / GLY in the wells, and add a 10%... Complete culture medium containing CCK8 reagent was incubated at 37°C for 1 hour in the dark. After the reaction, the OD values ​​of each well in the 96-well plate were measured using a microplate reader at 450 nm. The results were exported and statistically processed. The results showed that fractions 2, 4, and 6 exhibited better activity (see...). Figure 3 ).

[0119] Example 6: Crude extracts (water or alcohol extracts) of jasmine flower polypeptides are active.

[0120] To detect the cell-binding activity of crude extracts (water or alcohol extracts) of jasmine flower peptides: 1. Resuspend the pre-cultured cell line as a cell suspension, determine the concentration, dilute to an appropriate concentration, mix well, and place in a sample well; 2. Select an appropriate number of 96-well plates as needed, construct a glutamate-glycine injury model, and evenly seed the cells, with 3000-5000 cells per well. Six replicates were left for each test concentration; 3. After labeling the wells, use a 100µl pipette to aspirate the cell suspension from the sample wells, trying to aspirate from the center area, and add it to the 96-well plate. Incubate at 37°C until the cells adhere; 4. After cell adhesion, perform a two-fold serial dilution at the highest concentration of 250µg / ml (i.e., 250µg / ml, 125µg / ml, 62.5µg / ml, 31.25µg / ml, 15.625µg / ml, 7.8125µg / ml) and add the polypeptide water extract (H) and polypeptide alcohol extract (E) respectively. After drug addition, continue incubation; 5. After completing the above model, gently dry the culture medium mixed with the drug / GLU / GLY in the wells, and add 10%... Complete culture medium containing CCK8 reagent was incubated at 37°C for 1 hour in the dark. After the reaction, the OD values ​​of each well in the 96-well plate were measured using a microplate reader at 450 nm. The results were exported and statistically processed. The results showed that both the water extract (H) and the alcohol extract (E) of jasmine flower peptides were active (see...). Figure 4 ).

[0121] Example 7: Antidepressant Activity Analysis

[0122] This study used learned helplessness (LH) to establish a mouse model of depression. Modeling method: Five C57BL / 6 mice were housed per cage and exposed to unavoidable foot shocks for a period of time. Eventually, the animals became unable to avoid the shocks. The shocks were administered once daily for three consecutive days.

[0123] ① Shuttle Box Experiment. Stimulation method: The experimental group mice were exposed to one side of an electrified shuttle box and subjected to 60 unavoidable electrical stimulations with a current of 0.3 mA. Each shock lasted randomly for 3-10 seconds, with a random interval of 1-15 seconds between each shock. While the experimental group was being shocked, the control group (con) was placed in the shuttle box for 30 minutes without being shocked. The day after the last stimulation, the shuttle box experiment (SBT) was performed to measure learned helplessness behavior.

[0124] Measurement method: Mice were placed in a shuttle box for 5 minutes to acclimatize, followed by a 5-second, 200 Hz, 80 dB noise stimulus. If a mouse escaped during the noise stimulus, it was considered a successful active avoidance. If the mouse did not escape, it was given a 5-second, 0.3 mA electric shock stimulus. If the mouse escaped during the shock, it was considered a successful passive avoidance. If the mouse remained in the original shuttle box after the shock time ended, it was considered an avoidance failure (no response). A 3-second interval was observed before each noise stimulus, for a total of 30 cycles. When replacing mice, the shuttle box was cleaned of feces and alcohol was sprayed to remove the odor of the previous mouse. The number of active avoidances and the number of failed avoidances were recorded and counted. Results showed that administration of jasmine flower polypeptide extract (1 mg / kg by gavage) had a good antidepressant effect (see...). Figure 5 ).

[0125] ② Forced Swimming Test (FST). Two hours after drug administration, mice were individually placed in an plexiglass tank (25 cm high, 13 cm in diameter, 15 cm deep, 23 ± 2 °C) and allowed to swim for 6 minutes, followed by 2 minutes of acclimatization. The cumulative immobility time (the duration of cessation of struggling, floating, and only occasional limb movements to keep the head afloat) within the last 4 minutes was recorded. One day prior to the experiment, each mouse underwent a 15-minute pre-swimming test (pre-swimming acts as a stressor to induce despair in the mice during the next swim). Twenty-four hours later, the mice were placed back into the water for the formal swimming test. The results showed that administration of jasmine flower extract (1 mg / kg by gavage) had a good antidepressant effect (see...). Figure 6 In this embodiment, jasmine flower polypeptide extract (dried purified product) samples were dissolved in physiological saline to different concentrations and used as experimental groups (J).

[0126] Example 7: Study on the antidepressant mechanism

[0127] Experimental Procedure: ① Based on the properties of the target molecule, a super streptavidin (SSA) sensor chip was selected. The chip surface underwent special treatment to stably couple the target molecule. After biotinylation of the dopamine protein, it was immobilized on the BLI probe surface by the sensor. ② A 0.002% Tween PBS solution (PBST) was prepared and the sensor was pre-wetted. The sensor head was rinsed with buffer solution to obtain blank baseline data for establishing the sensor chip baseline. ③ Jasmine polypeptide extract (dried purified product) was prepared into a liquid using PBST solution. The sensor was immersed in the jasmine polypeptide extract liquid. After observing that a stable signal level was reached, a new baseline was recorded to record the initial state of the sensor under ligand-targeted protein binding saturation. ④ After completing the instrument preparation, the binding and dissociation times were set. The protein-linked sensor was placed in jasmine polypeptide extract solutions of different concentrations. The instrument recorded the changes in the BLI signal over time and recorded data at different stable concentrations for testing. ⑤ After completing the stable test, the equilibrium dissociation constant (KD) was calculated by automatically fitting the data of the binding and dissociation stages based on the concentration gradient analysis software.

[0128] Results analysis: Jasmine flower polypeptide extract has the ability to target dopamine receptor binding. For example... Figure 7 As shown, the X-axis represents the test time, and the Y-axis represents the response intensity of the BLI instrument. Its KD value is 9.326E-06M, suggesting that its antidepressant effect can be achieved by targeting dopamine receptors.

[0129] The above description is merely a few embodiments of this application and is not intended to limit this application in any way. Although this application discloses preferred embodiments as described above, it is not intended to limit this application. Any changes or modifications made by those skilled in the art without departing from the scope of the technical solution of this application using the disclosed technical content are equivalent to equivalent implementation cases and fall within the scope of the technical solution.

Claims

1. Use of jasmine flower polypeptide extract in the preparation of a composition, wherein the jasmine flower polypeptide extract contains little or no volatile aromatic substances; the use of the composition is for at least one of the following purposes for an individual in need: ① prevention or treatment of diseases or conditions related to impaired neurotransmission; ② regulation or improvement of mood, reduction of mental stress, reduction of fatigue, promotion of restful sleep, regulation of hunger and overeating, regulation of motor nerve activity, increase of energy or improvement of overall health; and / or ③ prevention and treatment of mental stress, alleviation of physical symptoms associated with mental stress, and improvement of resistance or tolerance to mental stress.

2. The use of claim 1, wherein the disease or condition associated with impaired neurotransmission is neurocognitive impairment, depression (such as unipolar depression, bipolar depression, acute depression, chronic depression, subchronic depression, psychogenic depression, postpartum depression, menopausal depression syndrome, mild depression, or severe depression), anxiety disorder (such as premenstrual anxiety syndrome, social anxiety disorder), aggression, attention deficit disorder (ADS), seasonal affective disorder, fibromyalgia syndrome, chronic fatigue, sleep disorder (i.e., insomnia), post-traumatic stress disorder, panic disorder, obsessive-compulsive disorder, ADHD, nervousness, migraine / primary headache and general pain, vomiting, bulimia, anorexia nervosa, eating disorders, gastrointestinal disorders, burnout syndrome, irritability and fatigue; Alternatively, the regulation or improvement of mood refers to regulating or improving an individual's (1) rapid mood changes (such as sudden sadness, anger, or rage); (2) persistent and severe anger and anxiety; (3) anxiety and tension; (4) depression and despair; (5) lack of interest in daily life; (6) fatigue or low energy; (7) poor concentration; (8) changes in appetite; (9) sleep disorders; (10) difficulty in controlling emotions; (11) nervousness; (12) memory difficulties; and / or (13) melancholy, so as to restore the individual to a positive, optimistic, and healthy state of mind and body.

3. The use as described in any of the preceding claims, wherein the jasmine flower polypeptide extract is the direct active ingredient, the main active ingredient, or the sole active ingredient in the composition.

4. The use as described in any of the preceding claims, wherein the composition is a pharmaceutical composition, a food for special medical purposes, a functional food or beverage, a dietary supplement, a complete nutritional formula food, a health food, a health care product, a nutritional composition, or a cosmetic; or the individual is a person (e.g., an infant, child, adolescent, middle-aged or elderly person), a companion animal, a laboratory animal, or a farm animal.

5. The use as described in any of the preceding claims, wherein the composition is administered systemically, topically, parenterally (e.g., via mucosal administration, transdermal administration, microneedle administration) or non-invasively; Alternatively, the composition may be a topical preparation, an oral preparation, or an injection.

6. The use as claimed in any of the preceding claims, wherein the composition is used in combination with one or more other drugs; said other drugs include, for example, selective serotonin reuptake inhibitors (SSRIs), serotonin modulators (5-HT2 blockers), 5-HT-norepinephrine reuptake inhibitors, dopamine-norepinephrine reuptake inhibitors, heterocyclic antidepressants, monoamine oxidase inhibitors (MAOIs), melatonin-producing antidepressants, or ketamine analogs.

7. The use as claimed in claim 6, wherein the composition is formulated with one or more other drugs for administration alone, concurrently, continuously, sequentially, alternately, or at intervals.

8. A jasmine flower polypeptide extract, a product containing the jasmine flower polypeptide extract, or a kit or device containing the jasmine flower polypeptide extract, characterized in that, The preparation method of the jasmine flower polypeptide extract is as follows: ① Extract jasmine flowers with a highly polar extractant to remove volatile aromatic substances, thereby forming a crude extract (such as a water extract or an alcohol extract); optionally, ② further extract the crude extract; optionally, ③ precipitate the collected extract after extraction; optionally, ④ concentrate and dry the precipitate.

9. The jasmine flower polypeptide extract as described in claim 8, characterized in that, The extraction reagent is a C3-C6 alcohol (preferably n-butanol), and the precipitation reagent is a mixed solvent of dichloromethane / trichloromethane and n-butanol.

10. A jasmine flower polypeptide extract, a product containing the jasmine flower polypeptide extract, or a kit or device containing the jasmine flower polypeptide extract, characterized in that, The jasmine flower polypeptide extract includes any one or more fractions from fractions 2-6 eluted by gradient elution of the chromatographic column, all fractions eluted within 50 minutes, or all fractions eluted within 55 minutes; preferably, it includes any or more fractions from fractions 2, 4, 5, and 6; preferably, it includes any or more fractions from fractions 2, 4, and 6. The chromatographic column is a C18 OBD Prep Column. A 5 μM, 10 mm × 250 nm chromatographic column was used. The sample loading volume for gradient elution of the column was 500 μl of jasmine flower polypeptide extract. The gradient elution conditions were as follows: mobile phase A was ultrapure water containing 0.1% formic acid, mobile phase B was methanol, and the gradient elution program was: 0 min: 80% A and 20% B; 50 min: 30% A and 70% B; 55 min: 2% A and 98% B; 60 min: 80% A and 20% B; 65 min: 80% A and 20% B; the detection wavelength was 254 nm, the flow rate was 7 mL / min, and fractions 2-6 were obtained by stepwise separation. The jasmine flower polypeptide extract was prepared by dissolving the extract sample in 3 ml of 5% methanol, then sonicating it at room temperature for 30 min, and finally passing it through a 0.22 μM membrane to obtain the filtrate. The extract sample is the solid obtained after concentration and alcohol removal of the ethanol-water extract of jasmine flower polypeptide; Fractions 2-6 correspond to the fractions collected by column chromatography from minutes 21 to 54; wherein, fraction 2 corresponds to the fraction collected by column chromatography from minutes 21 to 28, fraction 3 corresponds to the fraction collected by column chromatography from minutes 28 to 39, fraction 4 corresponds to the fraction collected by column chromatography from minutes 39 to 44, fraction 5 corresponds to the fraction collected by column chromatography from minutes 44 to 49, and fraction 6 corresponds to the fraction collected by column chromatography from minutes 49 to 54.

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