CN110721239A

CN110721239A - Application of Fuzhuan tea extract in preparation of antifungal drugs

Info

Publication number: CN110721239A
Application number: CN201911190691.5A
Authority: CN
Inventors: 傅文燕; 丁敏
Original assignee: Fengchao Pharmaceutical Technology (shanghai) Co Ltd
Current assignee: Fengchao Pharmaceutical Technology (shanghai) Co Ltd
Priority date: 2019-11-28
Filing date: 2019-11-28
Publication date: 2020-01-24
Also published as: WO2021104213A1

Abstract

The invention discloses an application of a Fuzhuan tea extract in preparation of antifungal medicines, wherein fungi refer to fungi of Ascomycota, Basidiomycota, Deuteromycota and zygomycota. The Fuzhuan tea extract provided by the invention has a strong killing effect on cryptococcus and the like, does not influence mammalian cells, does not generate hemolytic reaction, has the potential of being developed into a medicine for preventing or treating diseases caused by fungal infection, and can be used as an antifungal medicine.

Description

Application of Fuzhuan tea extract in preparation of antifungal drugs

Technical Field

The invention belongs to the technical field of medicines and health-care foods, and particularly relates to application of a Fuzhuan tea extract in preparation of antifungal medicines.

Background

The Fu tea is a post-fermented tea which is prepared by using raw dark green tea or crude old green tea as a raw material and reprocessing, belongs to one of six basic tea types in China, and is also a fully fermented tea. Fu tea has a long history, and is called Fu tea because it is processed and prepared in summer overnight and has fragrance and action similar to that of Poria. The main form of the Fuzhuan tea is compressed tea, namely, in the manufacturing process, in order to facilitate transportation and storage, a producer compresses the Fuzhuan tea into a brick shape, so that the Fuzhuan tea looks like a brick, and is also named as the Fuzhuan tea. At present, the Fu tea is also sold in the forms of loose tea, cake tea and the like. Fu tea is mainly sold in areas of frontier regions, such as Xinjiang, Gansu, Tibet and the like, is commonly called as 'side tea' or 'side selling tea', and is known as 'mystery tea on ancient silk roads' and 'black and gold on silk roads' in more than forty countries, such as Zhongya, Western Asia and the like, along the 'silk roads'.

Modern pharmacological research shows that the Fu tea has obvious health effects of promoting digestion, reducing blood fat, nourishing stomach, losing weight, regulating glycometabolism, inhibiting tumor cells, enhancing immunity and the like. The tuckahoe tea can promote metabolism of human bodies, improve immunity, strengthen physique and delay senility after being drunk for a long time, and has obvious pharmacological health care and pathological prevention effects.

Research shows that the pharmacological activity of the Fuzhuan tea is the result of the combined action of the active ingredients of the tea and a dominant bacterium, namely Eurotium cristatum, which is naturally formed in the fermentation process. The eurotium cristatum plays an important role in the performance of the quality and the pharmacological effect of the Fu tea. Eurotium cristatum called as golden flower fungus in Fuzhuan tea, belongs to Eurotium cristatum of Tricholomataceae of Ascomycetales, and is suitable for growing in soil, Fuzhuan tea, Cordyceps, Chinese medicinal tablet and sawdust. Eurotium cristatum consists of ascocarp and hyphae. The eurotium cristatum can produce rich natural active ingredients, thereby endowing the Fuzhuan tea with unique color, fragrance and taste, and being the material basis of the Fuzhuan tea with the health-care effects of promoting digestion, reducing fat and losing weight, inhibiting tumor cells, treating cardiovascular diseases and the like. The eurotium cristatum also generates a large amount of special bioactive substances in the Fu tea in the process of co-fermentation of the Fu tea and the tea.

At present, there are many reports on the research of Fu tea. For example, the process of preparing instant black tea by liquid fermentation is reported by the lu courser (the master thesis of agriculture university in Anhui, Eurotium cristatum liquid fermentation and functional research thereof, and the director Yuepeng Xiang). Yuan Yong (thesis of Master academic thesis of Hunan agriculture university, the preliminary research on pharmacological efficacy and chemical components of Fuzhuan tea golden flower fungus extract, and the guiding teacher Liu Zhong Hua) reported the research on the extraction and separation of Eurotium cristatum spore powder. Eichhornia et al (Eichhornia, Liusu pure, Yuanqian, Zhao le, research on wall breaking method of Ascomyces guanfaciei ascospores, food and machinery 2011,27 (3): 137-. Lu et al (Advance Journal of Food Science and technology 2016, 10 (8): 591-596.) reported the study of the active ingredients in Eurotium cristatum fermented black tea. Zou et al (Molecules 2014, 19, 17839-17847) reported alkaloid compounds produced by fermentation of Eurotium cristatum. Fu et al (Food Research International 2011, 44: 2999-3005.) report the mechanism of hypolipidemic activity of Fu tea. Yan et al (Food and Agricultural Immunology 2017, 28 (3): 388-402) report the effect and mechanism of Eurotium cristatum in Fuzhuan tea for activating immunity.

According to 26.09.2013, the original national quality inspection and inspection bureau approves the application of geographical sign product protection to Jingyang Fuzhuan tea (national quality supervision and inspection and quarantine bureau announcements about approval of geographical sign product protection to salt pond large salt (river east large salt), salt pond black mud, Jingyang Fuzhuan tea, Yijun walnut and Hongcuan wine (No. 137 in 2013)) makes clear requirements on the quality and the technology of the Fuzhuan tea, and facilitates subsequent research on the Fuzhuan tea.

Disclosure of Invention

The invention aims to provide application of a Fuzhuan tea extract in preparation of antifungal medicines.

In order to achieve the purpose, the technical scheme adopted by the invention is as follows:

the invention provides an application of a Fuzhuan tea extract in preparing an antifungal medicine.

The Fuzhuan tea extract has broad-spectrum sterilization and bacteriostasis effects on fungi of Ascomycetes, Basidiomycetes, Deuteromycetes and Zygomycota. For example, the fungi that the Fuzhuan tea extract of the present invention can inhibit is selected from the group consisting of Cryptococcus (Cryptococcus), Candida (Candida sp.), Saccharomyces (Saccharomyces sp.), Malassezia (Malassezia), Aspergillus (Aspergillus sp.), Coccidioides (Coccidioides), Paracoccus (Paracoccidiaides), Histoplasma (Histoplasma), Blastomyces (Blastomyces), Neurospora crassa (Neurospora crassa), and the like. Preferably, the fu tea extract has the capability of inhibiting the growth of or killing the following fungi: cryptococcus neoformans (Cryptococcus neoformans), Cryptococcus gattii (Cryptococcus gattii), Saccharomyces cerevisiae (Saccharomyces cerevisiae), Candida glabrata (Candida glabrata), Candida albicans (Candida albicans), Malassezia terrestris, Candida krusei (Candida krusei), Candida tropicalis (Candida atropicalis), Candida parapsilosis (Candida parapsilosis), Trichosporon bailii (Trichosporon beigelii), and Malassezia furfur (Malassezia furfur).

In some preferred embodiments of the present invention, the extract of Fuzhuan tea has an inhibitory effect on superficial fungi which are common to human skin, and the superficial fungi comprise Trichophyton rubrum, Trichophyton gypseum, Trichophyton floccosum, Microsporum gypseum, Microsporum canis, Pityrosporum ovale and Candida. In some preferred embodiments of the present invention, the fu tea extract has especially the use of preparing products for fungal skin diseases, such as tinea capitis, tinea manuum and tinea pedis, tinea cruris, tinea corporis, fungal skin of nail (onychomycosis), eczema-like ringworm, tinea versicolor and the like.

The Fuzhuan tea extract refers to a water-soluble and/or fat-soluble extract. In other embodiments of the present invention, the fu tea extract refers to supercritical CO₂Extract, organic solvent extract, aqueous extract or mixture thereof.

The organic solvent is ethanol.

The preparation method of the organic solvent extract comprises the following steps: grinding Fuzhuan tea into powder, wherein the diameter of the powder is not more than 0.5cm, adding 85% -95% ethanol into the Fuzhuan tea powder according to the solid-to-liquid ratio of 1:20(m/v), ultrasonically breaking the wall until no tea powder and thallus can be seen by naked eyes, then carrying out cold soaking for 18-28 hours, collecting filtrate, repeating for 2-5 times, carrying out reduced pressure distillation on the filtrate, concentrating the filtrate, carrying out low-temperature drying to obtain a crude extract, adsorbing and decoloring the crude extract by macroporous resin (AB-8 macroporous adsorption resin), eluting by 85% -95% ethanol, concentrating and evaporating to dryness, and carrying out low-temperature freeze drying to obtain the organic solvent extract of the Fuzhuan tea.

The preparation method of the water extract comprises the following steps: grinding Fuzhuan tea into powder, wherein the diameter of the powder is not more than 0.5cm, adding water into the Fuzhuan tea powder according to the solid-to-liquid ratio of 1:10(m/v), uniformly stirring, ultrasonically breaking the wall until no tea powder and thallus can be seen, standing for 2 hours at room temperature, and boiling for 2 hours in a boiling water bath to obtain a mixed solution; taking out the mixed solution, cooling, ultracentrifuging, and taking the supernatant; filtering and sterilizing the supernatant with 0.22 μm filter membrane to obtain water extract of Fuzhuan tea. Concentrating and evaporating to dryness if necessary, and freeze-drying at low temperature.

The supercritical CO₂The preparation method of the extract comprises the following steps: grinding Fuzhuan tea into powder with diameter not more than 0.5cm, completely crushing tea powder and thallus, adding water vapor for pre-soaking until water content is increased to 40%, placing the pre-soaked Fuzhuan tea powder into an extraction tank, and continuously feeding CO into the tank₂Gradually extracting the crude extract with CO of the crude extract₂Sending to a cleaning tank, allowing the Fuzhuan tea extract to enter water phase, and passing through supercritical CO₂Extracting to obtain crude extract, hydrolyzing starch and fiber (cellulose, hemicellulose, etc.) with mixture of cellulase and alpha-amylase (concentration of 0.5% each), heating at 65 deg.C for 30min to denature the enzyme, and centrifuging to remove impurities; removing residual protein (exudate) with 15kDa ultrafiltration column, bleaching extract with activated carbon and filtering and recovering, sterilizing the product with 0.2 μm filter membrane, and lyophilizing or packaging to obtain Fuzhuan tea supercritical CO₂And (3) extracting.

The antifungal medicine takes the Fuzhuan tea extract as the only active component, or a medicine composition containing the Fuzhuan tea extract.

The pharmaceutical composition containing the Fuzhuan tea extract is a pharmaceutical composition consisting of the Fuzhuan tea extract and one or more pharmaceutically-allowable auxiliary materials.

The content of the Fuzhuan tea extract in the antifungal medicine is 0.1-99 wt%.

The content of the Fuzhuan tea extract in the antifungal medicine is 0.5-90 wt%.

The antifungal medicine is a medicine for preventing or treating fungal infection.

The auxiliary material is at least one of diluent, excipient, adhesive, filler, disintegrating agent, flavoring agent and sweetener.

The Fuzhuan tea extract can be prepared into a medicinal preparation with a conventional pharmaceutic adjuvant in pharmaceutics.

The pharmaceutical formulation is at least one of a capsule, a suspension, a tablet, a powder, an emulsion, a solution, a syrup, or an injection.

The administration mode of the pharmaceutical preparation is oral administration and injection.

In the description of the present invention, the fu tea is preferably jing yang fu tea which meets the technical requirements for the quality of fu tea as described in "quality inspection bureau of the national quality supervision, inspection and quarantine bureau" bulletin of the ministry of quality inspection for approving the geographical indication product protection of the salt pond salt (large salt in river east), the black mud in the salt pond in canal city, jing yang fu brick tea, lijun walnut and hong chuan wine (No. 137 in 2013).

The fu tea extract can be administered to a subject in any suitable dosage without limitation.

For example, the above dosage may be determined and/or adjusted by routine experimentation by one of ordinary skill in the art based on the desired effect of administration and the condition of the subject, among others.

In some embodiments, the present invention provides a fu tea extract comprising the fu tea extract described above in an effective concentration. Preferably, the effective concentration is not less than 1 mg/ml.

In some embodiments, the antifungal agent comprises the fu tea extract in a therapeutically effective concentration. The term "therapeutically effective concentration" as used herein means a concentration that is capable of slowing or inhibiting the growth of a fungus in clinical therapeutics and thus treating or slowing the rate of progression of a fungus-induced disease. The clinician can determine and adjust the therapeutically effective concentration of the fu tea extract for the age, sex, health condition, disease severity, diet, etc. of the subject to be treated, and can use the antifungal drug in combination with any other known drug or adjuvant based on the expected clinical benefit.

The invention also comprises a composition containing the Fuzhuan tea extract as an active ingredient according to the type of application. The composition may further comprise at least one pharmaceutically acceptable carrier. Examples of carriers include, but are not limited to: excipients, diluents, granulating agents, lubricants, binders, disintegrating agents, wetting agents, emulsions, colorants, mold release agents, coating agents, sweeteners, flavoring agents, fragrances, preservatives, antioxidants, plasticizers, gelling agents, thickening agents, hardening agents, solidifying agents, suspending agents, surfactants, humectants, carriers, stabilizers, and combinations thereof; for example, water, polyethylene glycol, starch, sugar, microcrystalline cellulose, talc, povidone (PVP), and the like.

In addition, the antifungal agent of the present invention may contain one or more of the following substances: hinokitiol, herb, phellodendron, coptis, scutellaria, liquorice, pseudo-ginseng, Korean ginseng, crude drug extract, cypress oil, jojoba oil, plant essential oil, horse oil, lard, eicosapentaenoic acid, docosahexaenoic acid, perilla oil, garlic oil, synthetic vitamin E, natural vitamin E, gamma-oryzanol, beta-carotene, vitamin B1, vitamin B2, vitamin B6, vitamin 12, vitamin C, polyphenol, lycopene, mushroom mycelium culture extract, mushroom fruiting body culture extract, ganoderma lucidum, agaricus mushroom extract, phyllostachys major extract, grifola frondosa extract, chlorella, spirulina, tannin, alginic acid, catechin, glycyrrhizin, propolis, ginkgo leaf extract, Korean ginseng extract, American ginseng extract, acanthopanax extract, salvia leaf extract, ginseng extract, aloe vera extract, chitosan, inositol, peptide, ceramide, nicotinic acid, total isoflavone, pantothenic acid, biotin, total carotenoid, selenium, zinc, chromium, collagen, chondroitin sulfate, mucopolysaccharin, calcium, royal jelly, millet, field horsetail, saw palmetto, turmeric extract, silybum marianum extract, garcinia extract, isopropyl alcohol, sodium hyaluronate, chlorhexidine gluconate, povidone iodine, ethanol, benzalkonium cetyl phosphate, triclosan, chloroxylenol, isopropylmethylphenol, benzalkonium chloride, etc., thereby further preparing antifungal agents having high antifungal effects.

In addition, the antifungal medicine of the present invention may also contain one or several kinds of coenzyme Q10, alpha-lipoic acid, vitamin P, vitamin E, gamma-oryzanol, beta-carotene, tannin, vitamin C and heme iron as antioxidant and antisenility preparation.

In some embodiments, the extract of Fuzhuan tea of the present invention can also be combined with other antifungal agents known in the art to make pharmaceutical compositions, or by co-administration to achieve a faster or stronger fungal inhibitory effect. Antifungal agents that can be used in combination with the fu tea extract of the present invention include, but are not limited to: 5-flucytosine, amphotericin B, anidulafungin, bifonazole, butoconazole, caspofungin, chlordantoin, chlorphenesin, ciclopirox olamine, clotrimazole, eberconazole, econazole, fluconazole, flutrimazole, isavuconazole, isoconazole, itraconazole, ketoconazole, micafungin, miconazole, nitroaldoxime, posaconazole, tioconazole, terconazole, undecylenic acid, and pharmaceutically acceptable salts or esters of the above antifungal agents.

The antifungal agent of the present invention may contain an emulsifier and/or a plasticizer, and may be in the form of a solid, gel, emulsion, paste or liquid. Here, coconut oil is an example of the emulsifier, and beeswax is an example of the plasticizer.

In addition, the amount of beeswax added to the antifungal agent of the present invention may be 50mg to 600mg, and the amount of purified coconut oil may be 50mg to 600 mg.

The fu tea extract described herein can be administered directly or can be formulated into any dosage form known in the art, such as, but not limited to: tablet, capsule, injection, powder, granule, aerosol, dripping pill, suppository, tincture, lozenge, syrup, suspension, nanometer preparation, etc.

The fu tea extract described herein can be administered continuously or at constant or varying intervals over a period of time, for example, once daily, twice daily, once weekly, twice weekly, monthly, every 1 month, or in alternating order every 1 month and every 2 months.

In some embodiments, the extract of fu tea of the present invention may be administered by any route, e.g., via intravenous administration, intradermal administration, intramuscular administration, intraarterial administration, intralesional administration, transdermal administration, subcutaneous administration, intraperitoneal administration, intracerebroventricular administration, or by aerosol administration. In some embodiments, the administration is therapeutic or prophylactic.

The fu tea extract of the present invention is capable of preventing and/or treating a fungal infection in a subject. The subject is preferably a vertebrate, including a human, a non-human primate (e.g., chimpanzee, macaque, spider monkey, etc.), a mouse, rat, guinea pig, rat, dog, cat, pig, horse, sheep or cow, fish (e.g., zebrafish, etc.), or a bird (e.g., chicken, emu, etc.), but is not limited to these examples. In addition, the Fuzhuan tea extract of the present invention can also be used for preventing and/or treating fungal infections in the above animals, such as dogs, cats, pigs, etc.

The Fuzhuan tea extract can also be used for sterilizing a sample. Preferably, the sample comprises a sample obtained from or derived from an environmental source comprising: food processing (e.g., beverages, dairy products), agricultural products, crops, poultry, livestock, meat, fish, beverages, dairy products, water (including wastewater), ponds, rivers, reservoirs, swimming pools, soil, food processing and/or packaging sites, slaughter sites, agricultural sites, farms and plantations (including hydroponic food plantations), pharmaceutical manufacturing plants, or any combination of the foregoing environmental sources.

In other aspects of the invention, the invention also includes the following uses of the fu tea extract: inducing apoptosis of fungal cells; inhibiting fungal cell proliferation; inhibit the synthesis of the ergosterol of the fungus and reduce the germination rate of the fungal spores. Namely, the antifungal medicine is a product for inducing apoptosis of fungal cells, a product for inhibiting proliferation of fungal cells, a product for inhibiting synthesis of fungal ergosterol and a product for reducing germination rate of fungal spores. The fungi is selected from Cryptococcus neoformans, Cryptococcus gatus, Saccharomyces cerevisiae, Candida glabrata, Candida albicans, Candida krusei, Candida tropicalis, Malassezia equi, Candida parapsilosis, Trichophyton baijili, Malassezia furfur, Trichophyton rubrum, Trichophyton gypseum, Trichophyton floccosum, Microsporum gypseum, Microsporum canis, Pityrosporum, Candida

Due to the adoption of the technical scheme, the invention has the following advantages and beneficial effects:

the Fuzhuan tea extract provided by the invention has a strong killing effect on cryptococcus and the like, does not influence mammalian cells, does not generate hemolytic reaction, has the potential of being developed into a medicine for preventing or treating diseases caused by fungal infection, namely can be used as an antifungal medicine, and can be safely used for a human body because the Fuzhuan tea extract is a natural source. Moreover, the Fuzhuan tea extract is used as an antifungal drug, so that the infection of dogs in the treatment of malassezia infection which causes troubles in medical treatment fields can be prevented. In addition, in the care site or the medical site, the finger or the affected part is cleaned with ethanol for sterilization, a disinfectant, and a bactericide, but when such cleaning is performed at a high frequency, there is a very high possibility that the skin is dried, penetrates into the wound, and the skin is damaged such as deterioration of dermatitis. The Fuzhuan tea extract can be used as an antifungal medicine for treating dermatitis, and can preserve moisture and improve the state of skin.

Drawings

FIG. 1 is a schematic diagram of the statistical results of flow cytometry of PI staining after the treatment of cryptococcus neoformans with Pu' er tea extract and Fuzhuan tea extract (1 mg/ml).

FIG. 2 is a graph showing the hemolytic toxicity of Fuzhuan tea extract.

Detailed Description

In order to more clearly illustrate the invention, the invention is further described below in connection with preferred embodiments. It is to be understood by persons skilled in the art that the following detailed description is illustrative and not restrictive, and is not to be taken as limiting the scope of the invention.

In the example of statistical calculation, p-values are calculated by nonparametric T-test between two groups, p-values are calculated by square difference analysis and comparison between groups, and p-values are calculated by chi-square test for rate comparison.

Example 1

The preparation method of the fu tea organic solvent extract comprises the following steps: grinding Fuzhuan tea into powder, wherein the diameter of tea powder is not more than 0.5cm, adding 85% -95% ethanol into the Fuzhuan tea powder according to the solid-to-liquid ratio of 1:20(m/v), and performing ultrasonic wall breaking until the tea powder and thalli are not visible to naked eyes. Then cold soaking for 18-28 hours, collecting filtrate, repeating for 2-5 times, distilling the filtrate under reduced pressure, concentrating the filtrate, drying at low temperature to obtain crude extract, adsorbing and decolorizing the crude extract by macroporous resin (AB-8 macroporous adsorption resin, Donghong chemical industry), eluting by 85% -95% ethanol, concentrating and evaporating to dryness, and freeze-drying at low temperature to obtain the fu tea organic solvent extract.

Example 2

The preparation method of the Fuzhuan tea water extract comprises the following steps: grinding Fuzhuan tea into powder, wherein the diameter of tea powder is not more than 0.5cm, adding water into the Fuzhuan tea powder according to the solid-to-liquid ratio of 1:10(m/v), stirring uniformly, and breaking the wall by ultrasonic waves until the tea powder and thalli are not visible to naked eyes. Then standing for 2 hours at room temperature, and boiling for 2 hours in a boiling water bath to obtain a mixed solution; taking out the mixed solution, cooling, ultracentrifuging, and taking the supernatant; filtering and sterilizing the supernatant with 0.22 μm filter membrane to obtain water extract of Fuzhuan tea. Concentrating and evaporating to dryness if necessary, and freeze-drying at low temperature.

Example 3

The preparation method of the Fuzhuan tea extract comprises the following steps:

grinding Fuzhuan tea into powder with diameter not more than 0.5cm, completely crushing the Fuzhuan tea powder with a dry wall breaking machine, adding water vapor for pre-soaking until the water content is increased to 40%, filling the pre-soaked Fuzhuan tea powder into an extraction tank, and continuously feeding CO into the extraction tank₂And the crude extract is gradually extracted. CO with crude extract₂Sending to a cleaning tank, allowing the Fuzhuan tea extract to enter water phase, and passing through supercritical CO₂Extracting to obtain crude extract, hydrolyzing starch and fiber (cellulose, hemicellulose, etc.) with mixture of cellulase and alpha-amylase (concentration of 0.5% each), and reacting at the temperature ofHeat-treating at 65 deg.C for 30min to denature the enzyme, centrifuging to remove impurities; removing residual protein (exudate) with 15kDa ultrafiltration column, bleaching extract with activated carbon and filtering and recovering, sterilizing the product with 0.2 μm filter membrane, and lyophilizing or packaging to obtain Fuzhuan tea supercritical CO₂And (3) extracting.

Pu her tea extract is obtained by similar method.

Example 4

Inhibition effect of Fuzhuan tea extract on saccharomyces cerevisiae

In this example, the test subject Saccharomyces cerevisiae (Saccharomyces cerevisiae) was BY4741(invitrogen), and the genotype was MAT α ura3 Δ met15 Δ leu2 Δ his3 Δ.

Saccharomyces cerevisiae was cultured to log phase in YPD medium (10g yeast extract, 20g peptone in water, autoclaved at 121 ℃ for 20min, cooled and added to 100mL 10 Xglucose stock). Subsequently, the Fuzhuan tea extract prepared in example 3 and the control Pu' er tea extract were added to a final concentration of 1mg/ml, and treated for 2 hours. Propidium Iodide (PI) staining of yeast cells was performed to clarify the apoptotic effect of each extract on yeast cells and then analyzed by flow cytometry (BECKMAN MoFlo XDP). The results show that the Fuzhuan tea extract has stronger effect of mediating the apoptosis of the yeast cells (shown in the table 1). The prepared Saccharomyces cerevisiae was then spread on a plate at a density of 10⁶CFU/ml. Then, 1mg/ml Fuzhuan tea extract and 5ml of control Pu' er tea extract as described in example 3 were uniformly spread on a plate and cultured at 37 ℃ for 2 days. Plate colony counts showed that 69.8% of the Saccharomyces cerevisiae cells were killed by 1mg/ml of Fuzhuan tea extract (as shown in Table 2).

TABLE 1 staining of yeast cells with PI

	PI％	SD	p value
				Blank control	6.57	0.25
Pu' er tea extract	7.11	0.16	p＞0.05
				Fuzhuan tea extract	71.52	5.26	p＜0.05

TABLE 2 Yeast cell plate count

	Relative count%	SD	p value
				Blank control	100	3.35
Pu' er tea extract	98.55	6.56	p＞0.05
				Fuzhuan tea extract	30.21	4.85	p＜0.05

Example 5

Inhibition effect of Fuzhuan tea extract on malassezia

Will modulate to about 10⁶CFU/ml of Malassezia (Thermo # R4604023) was plated on Mueller-Hinton medium (Bio-Rad # 3564884). Then, 1mg/ml of the Fuzhuan tea extract prepared in example 3 and 5ml of the control Pu' er tea extract were uniformly spread on Mueller-Hinton medium and cultured at 37 ℃ for 2 days. Plate colony counts showed that 80.50% of malassezia cells were killed by the fu tea extract.

TABLE 3 Malassezia cell plate count

	Relative count%	SD	p value
				Blank control	100	5.51
Pu' er tea extract	106.67	10.22	p＞0.05
				Fuzhuan tea extract	19.50	4.35	p＜0.05

Example 6

Inhibition of cryptococcus by Fuzhuan tea extract

Cryptococcosis is an infectious mycosis of zoonosis mainly caused by cryptococcus neoformans, cryptococcus gatae and the like. With the advent of antibiotic drugs such as amphotericin B, cryptococcosis is considered a disease within the control of humans. However, with the widespread use of invasive procedures and the increasing use of immunosuppressive agents, particularly in AIDS patients, not only cryptococcosis infection rates continue to rise, but also new features such as high mortality rates, and an increase in infection rates in immunocompromised persons have emerged (Bielska, Ewa, "Whole masks Cryptococcus gattii a pathogen." FEMSyeast research 16.1(2015): fov 106.). The 2016 world health organization data showed that about 625,000 people died of cryptococcal meningitis, the east and south of Africa, and the south of Asia as the major disease areas (https:// www.who.int/gho/publications/world _ health _ statistics/2016/en /). Currently, the anti-cryptococcus drugs commonly used in clinical therapy for cryptococcus are polyenes, fluorocytosines and azoles (Coelho, Carolina, and Altartur Casadev. "Cryptococcal peptides and drug targets: the old, the new and the breeding." Cellular microbiology 18.6(2016): 792) 792 799.). However, the above drugs have significant side effects and may cause damage to normal cells and organs of the human body. In addition, cryptococcus is highly susceptible to the development of resistance to the above-mentioned drugs. In clinical practice, some patients cannot be effectively treated by using the existing medicines. In view of the high mortality rate of cryptococcosis, the altered epidemiological characteristics and the emergence of resistant strains, there is a need to find new drugs that can replace traditional antibiotics.

Cryptococcus neoformans (Cryptococcus neoformans) JEC21, which was the test subject in this example, was obtained from the institute of microorganisms of the Chinese academy of sciences. RPMI1640 medium: RPMI-1640 (Sigma) was added 0.165M/L mops (Sigma) and sterile filtered. The minimum inhibitory concentration of the Fuzhuan tea extract was determined by the method described in Makovitzki, Arik, Dorit Avramami, and Yeachiel Shai, "Ultrashort antibacterial and antibacterial peptides," Proceedings of the national Academy of Sciences 103.43(2006):15997 16002. by microbubmery dilution (M27-A2).

The cryptococcus neoformans was cultured in the RPMI1640 medium to the logarithmic phase, and after diluting to OD600 of 0.1 with the fresh RPMI1640 medium, the diluted cryptococcus neoformans culture solution was added to 96-well plates, and the Fuzhuan tea extract solution prepared in example 3 was added to each well to give final concentrations of 0mg/ml, 0.125mg/ml, 0.25mg/ml, 0.5mg/ml, 1mg/ml, 2mg/ml, and 5mg/ml, respectively, and the total volume of each sample was 100. mu.l. Three replicate wells were set for each concentration to exclude the effects of fungal contamination in the experiment. The 96-well plates were incubated at 37 ℃ for 24h with RPMI1640 medium as a blank, OD600 values were determined (full-automatic growth curve analyser, Finland Bioscreen, with blank as OD600 reference), and duplicate samples were determined and averaged three times. The minimum concentration required for growth of the cryptococcus neoformans to be 50% below the initial OD600 after 24h of incubation was defined as the Minimum Inhibitory Concentration (MIC). According to this algorithm, in this example, the minimal inhibitory concentration of the Fuzhuan tea extract is such that the OD600 of the Cryptococcus neoformans after 24h is less than 0.15. The result shows that the minimal inhibitory concentration of the Fuzhuan tea extract is 0.5 mg/ml.

Experimental example 7

Propidium Iodide (PI) staining determination of Fuzhuan tea extract bactericidal effect

The fungicidal effect of Fuzhuan tea extract was quantitatively determined by PI staining according to the method described by Hernlem, Bradley, and Sui-Sheng hua, "Dual fluorochrome flow microbiological assessment of last vision." Current microbiology 61.1(2010):57-63. Cryptococcus in log phase was diluted to 1X10 using fresh RPMI1640 medium⁶Adding the Fuzhuan tea extract and the control Pu' er tea extract prepared in example 3 at a final concentration of 1mg/ml, culturing at 37 ℃ for 30 minutes, adding PI (amresco company) at a final concentration of 5 μ g/ml, standing in the dark for 15 minutes, and counting the number of fungi which are positive for PI staining by using a flow cytometer (BECKMAN MoFlo XDP), wherein the counted total number of cells is 50,000. FIG. 1 is a schematic diagram of the statistical results of flow cytometry of PI staining after the treatment of cryptococcus neoformans with Pu' er tea extract and Fuzhuan tea extract (1 mg/ml). Since PI can not pass through living cell membranes but can pass through damaged cell membranes to stain nuclei, the high staining ratio of PI indicates that the killing effect is remarkable. As can be seen from FIG. 1, the Fuzhuan tea extract has a significant killing effect on cryptococcus neoformans.

Experimental example 8

Effect of Fuzhuan tea extract on mammalian cells

At 5% CO₂Hela cells (from ATCC chamber) were cultured in a 37 ℃ incubator in a DMEM high-glucose medium (sigma). The Fuzhuan tea extract and the Pu' er tea extract prepared in example 3 and amphotericin B at a final concentration of 1mg/ml were added to the cell culture and treated for 6 hours, respectively. After the supernatant was aspirated, PI staining was performed (amresco, final concentration 20. mu.g/ml). The number of Hela cells positively stained for PI was counted by flow cytometry (BECKMANMoFlo XDP) and the total number of cells was 10,000. Flow cytometry results show that the Fuzhuan tea extract disclosed by the invention does not have a killing effect on mammalian cells, and the toxicity is far lower than that of amphotericin B (shown in Table 4).

TABLE 4 staining of mammalian cells Hela cells with PI

	PI％	SD	p-value (vs. amphotericin B)
				Blank control	4.48	0.56	p＜0.05
Pu' er tea extract	3.64	0.15	p＜0.05
				Fuzhuan tea extract	2.55	0.11	p＜0.05
Amphotericin B	55.58	6.77

Experimental example 9

Hemolytic toxicity of Fuzhuan tea extract

Blood samples to be tested in this example were obtained from C57BL mice.

The degree of hemolysis is an important index of the damage degree of the sample to be tested to the red blood cells, and the higher the hemoglobin content is, the higher the degree of hemolysis is, which indicates that the damage degree of the red blood cells is higher. The degree of damage of the red blood cells is indicated by detecting the content of hemoglobin in the supernatant.

To further investigate whether the Fuzhuan extract prepared In example 3 caused hemolysis In mammals, the degree of hemolysis of blood samples after 4 hours of treatment with 0mg/ml, 0.125mg/ml, 0.25mg/ml, 0.5mg/ml, 1mg/ml, 2mg/ml, 5mg/ml Fuzhuan extract was determined according to the method of Sung, Woo Sang, In-Seon Lee, and Dong Gun Lee. The absorbance at a wavelength of 540nm was measured with a microplate reader (SynergyH 4, Bolten USA) as an indication of the relative content of hemoglobin. The absorbance of the blood sample after 4h of addition of pure water (i.e., complete lysis of erythrocytes) was defined as the degree of hemolysis of 100%. As shown in FIG. 2, FIG. 2 is a schematic diagram showing the hemolytic toxicity result of Fuzhuan tea extract. When the concentration of the fu tea extract is 1mg/ml, the relative hemolysis degree of the fu tea extract sample is only about 2%, and the fu tea extract has no hemolytic toxicity to mammals at the concentration.

In conclusion, the Fuzhuan tea extract has a strong killing effect on cryptococcus and the like, does not influence mammalian cells, and does not generate hemolytic reaction. The Fuzhuan tea extract has the potential of being developed into a medicament for preventing or treating diseases caused by fungal infection. The Fuzhuan tea extract is a natural source, so that the Fuzhuan tea extract can be safely used for a human body. Moreover, the poria cocos tea extract can prevent the infection of dogs in the treatment of malassezia infection which causes troubles in medical treatment fields. In addition, in the care site or the medical site, the finger or the affected part is cleaned with ethanol for sterilization, a disinfectant, and a bactericide, but when such cleaning is performed at a high frequency, there is a very high possibility that the skin is dried, penetrates into the wound, and the skin is damaged such as deterioration of dermatitis. Although a disinfectant containing a moisturizing agent such as glycerin or a lubricant is available, it has a poor moisturizing effect and a reduced bactericidal effect, and is therefore not used in the care and/or medical field. In addition, such disinfectants cannot exert a recovery effect on already damaged skin, and there is no countermeasure to be taken. Therefore, the Fuzhuan tea extract can be used as an antifungal medicine for treating dermatitis, and can preserve moisture and improve the state of skin.

Example 10

Biological synthesis effect of Fuzhuan tea extract on cryptococcus ergosterol

Ergosterol is an important constituent of fungal cell membranes, and it plays an important role in maintaining the fluidity of the plasma membrane and stabilizing the structure of membrane molecules.

0.5ml of cryptococcus suspension with OD600 of 0.15 was added to the medium containing 1mg/ml of the Fuzhuan tea extract prepared in example 3, and the control group was a medium containing the same concentration of Pu her tea extract. After 3 days of shaking culture (28 ℃, 150rpm), centrifuging the culture solution to obtain thalli, weighing 1g of mycelia, putting into a mortar, adding 5ml of PBS, grinding into homogenate, adding 12ml of freshly prepared saponifier, uniformly mixing, and performing condensation reflux water bath saponification at 80 ℃ for 60 min. Extracting with 50ml of petroleum ether (30-60 ℃), washing the petroleum ether phase with 50ml of distilled water for 2 times, naturally volatilizing the petroleum ether phase, dissolving with absolute ethyl alcohol to a constant volume of 50ml, and repeating 3 times for each group to be tested. Preparing ergosterol standard solution with concentration of 30, 24, 18, 12, 6ug/ml, measuring absorption peak at 282nm, measuring the concentration of ergosterol of the sample to be measured by taking the absorbance as ordinate and the concentration as abscissa, and then calculating the content of ergosterol in each treatment group to obtain the content of ergosterol in each gram of mycelium (as shown in table 5).

TABLE 5 ergosterol content

	Ergosterol (mg/g)	SD	p value (vs. blank)
				Blank control	1.315	0.015
Pu' er tea extract	1.291	0.010	P＞0.05
				Fuzhuan tea extract	0.811	0.007	p＜0.05

As can be seen from the data in Table 5, the content of ergosterol in cryptococcus was significantly reduced after the treatment with the Fuzhuan tea extract, which indicates that the Fuzhuan tea extract affects the synthesis of ergosterol in cryptococcus and the permeability of the biofilm.

Example 11

Effect of Fuzhuan tea extract on Cryptococcus spores

Spores are an important reproductive organ of fungi, and when the fungi grow in an adverse environment, the spores are produced to resist the adverse environment. Activated cryptococcus was inoculated into a well-plated Sambucus agar medium plate (AMRESCO) containing 1mg/ml of the Fuzhuan tea extract prepared in example 3, and a control group containing 1mg/ml of a control Pu' er tea extract was inoculated in 3 replicates each. After culturing at 28 deg.C for 14 days, under aseptic conditionsThe surface of the plate was washed with sterilized brushes and 20m L sterile water to obtain the total number of spores washed, and the average density (pieces/cm) of spores was calculated²) (ii) a Then 50 mu L of each group of spore suspension is taken out and added into a clean concave slide, the concave slide is placed into a moisturizing culture dish for 6 hours at 28 ℃, the spore germination condition is observed under a microscope, and each concave slide has 3 random visual fields. Each group was repeated 3 times. According to the literature [ Xujie, Dengloxing, Hu national yuan, etc.. ] the artemisinin derivative antibacterial mechanism research [ J]Natural product research and development, 2018(5).]The average spore density and spore germination rate were calculated (as shown in tables 6 and 7).

TABLE 6 average spore densities

TABLE 7 spore germination Rate

	The germination rate%	SD	p value (vs. blank)
				Blank control	95.64	3.25
Pu' er tea extract	98.85	1.26	P＞0.05
				Fuzhuan tea extract	65.33	0.78	P＜0.05

As can be seen from the data in tables 6 and 7, the cryptococcus was cultured with the Fuzhuan tea extract, which did not promote sporulation, but significantly affected the germination of the spores, possibly destroying the spore structure.

Example 12

Fuzhuan tea extract has effects on common fungus and superficial dermatophyte.

The Fuzhuan tea extract prepared in example 3 was tested for its antibacterial activity against common fungi (Cryptococcus gatus, Candida glabrata, Candida krusei, Candida tropicalis, Candida parapsilosis, Trichophyton virginica, Malassezia furfur) and dermatophytes (Trichophyton rubrum, Trichophyton mentagrophytes, Microsporum canis, Microsporum gypseum, Epidermophyton floccosum, Candida albicans), and the control Pu' er tea extract was used as a control and the antibacterial activity of clotrimazole was used as a control of the currently used drugs.

Candida glabrata, Candida krusei, Candida tropicalis, Candida parapsilosis, Trichophyton baijili, Malassezia furfur, Trichophyton rubrum, Trichophyton mentagrophytes, Microsporum canis, Microsporum gypseum, Epidermophyton floccosum are derived from ATCC; the Candida albicans is from the institute of microorganisms of Chinese academy of sciences; the Cryptococcus gatherensis is from China medical fungi collection management center.

Preparing a medicine base: peptone agar base (4% glucose, 1% peptone, 2% agar) containing Fuzhuan tea extract was prepared, wherein Fuzhuan tea extract and control were diluted to 4mg/ml, 2mg/ml, 1mg/ml, 0.5mg/ml, 0.25mg/ml, 0.125mg/ml, 0.0625mg/ml, 0.0312mg/ml, 0.0156mg/ml, 0.0078mg/ml, 0.0039mg/ml, 0.0019mg/ml, subpackaged with test tubes (5 ml per tube), placed on a slant and numbered for use.

Inoculating the strain: selecting the activated strain, adding into 0.9% sodium chloride solution, grinding mycelium into unit cell with sterile grinder, and adjusting the bacterial suspension concentration to 1 × 10 with hemocytometer⁶CFU/ml, 100ul of bacterial suspension was inoculated into each tube, 2 tubes were inoculated at each concentration, and in addition to the above drugs, glucose peptone agar base and ethanol-containing glucose peptone agar base were used as placebo 1 and placebo 2, respectively. The growth condition is observed and recorded every day in a constant temperature box at 25 ℃, and the result is read after 7 days.

Interpretation of MIC values: the growth of each tube was compared to positive and negative control tubes, and the Minimal Inhibitory Concentration (MIC) end point was determined on the substrate tube with the lowest drug concentration and no culture growth.

Determination of MFC value: taking 2ml of glucose peptone liquid base, respectively adding into test tubes for 2h of bacteria-free growth in an antibacterial experiment, shaking, taking out 100ul, inoculating into glucose peptone agar base, incubating at 25 ℃ for 7 days, and taking the minimum concentration non-growth as the minimum bactericidal concentration (MFC) end point.

Quality control: positive and negative controls were set up separately and the procedure was repeated once as described above.

Statistical treatment: data were analyzed using SPSS13.0 software.

TABLE 8 inhibition of common strains by Fuzhuan tea extract (mg/ml)

The data in table 8 show that the fu tea extract has broad-spectrum antibacterial effect and stronger skin superficial antibacterial effect.

Example 13

Preparation of external film agent of Fuzhuan tea extract

Firstly, respectively adding 30g of polyvinyl alcohol 05-88 and 10g of sodium carboxymethylcellulose into 100ml of purified water for wetting, stirring, soaking for 48h to fully swell, heating in a water bath at 90 ℃ until the materials are dissolved, and respectively obtaining polyvinyl alcohol 05-88 glue serous fluid and sodium carboxymethylcellulose glue serous fluid for later use.

And secondly, weighing 1g of the Fuzhuan tea extract prepared in the embodiment 3, putting the Fuzhuan tea extract into a mortar, sequentially adding 5g of azone, 5g of Tween-80 and 3g of glycerol, the polyvinyl alcohol 05-88 glue serous fluid and the sodium carboxymethyl cellulose glue serous fluid prepared in the first step, grinding while adding until the grinding is uniform, putting the mixture into a water bath at 60 ℃ for heat preservation for 30min, and removing bubbles to obtain medicine membrane slurry.

And thirdly, pouring the medicinal membrane slurry prepared in the second step on a sterile glass plate, uniformly spreading, drying in an oven at 65 ℃, demoulding after film forming, cutting into small blocks of 6cm multiplied by 8cm, 20 blocks in total, irradiating the medicinal membrane for 15min by an ultraviolet lamp in a sterile operating room for sterilization, and packaging in a sterile bag to obtain the exterior-applied film agent of the fu tea extract.

Fourthly, preparing an external blank film agent: except that the second step is not added with the Fuzhuan tea extract, the types and the dosages of other steps and auxiliary materials are the same.

Fifthly, preparing a contrast external film agent: except that 1g of the Pu' er tea extract is added in the second step, the types and the dosages of other steps and auxiliary materials are the same.

Therapeutic test research of poria cocos tea extract on trichophyton mentagrophytes infected guinea pig model

Selecting trichophyton mentagrophytes (ATCC) as pathogenic bacteria, recovering the pathogenicity of the trichophyton mentagrophytes before the experiment, inoculating the trichophyton mentagrophytes into a sandcastle agar (SDA) inclined test tube, culturing at 26 ℃, carefully scraping colonies after 7-10 days, and preparing an infectious fungus suspension by using physiological saline for later use. SDA consisted of 4% glucose, 1% peptone and 2% agar.

48 healthy white guinea pigs are used for both male and female, and the weight of the guinea pigs ranges from 250 g to 350 g. Shearing long hairs on the back and abdomen of all white guinea pigs, removing hairs by a shaver to form a hair removing area of 8cm multiplied by 10cm, coating a proper amount of glycerin on the hair removing area to prevent skin from drying and cracking, repeatedly rubbing the skin on the hair removing area by abrasive paper, wherein the damaged surface is about 3cm multiplied by 5cm, applying the prepared trichophyton mentagrophytes suspension on the damaged skin, observing the pathological change degree of the skin in the infected area of the white guinea pigs on the 10 th day after the infection of fungi, scraping skin rash, scales or crusts, and detecting hypha or spore of the fungi by a microscope. Proves the success of the scrub wound infection method for infecting the guinea pig animal model.

After the trichophyton mentagrophytes infected guinea pig animal model was prepared, the infected model animals were randomly divided into four groups of 12 animals each having half male and female, each consisting of clotrimazole cream group (applied with clotrimazole cream, national standard character H50020636, specification 10 g: 3%), test group (applied with the exterior film preparation of fu tea extract prepared in this example), control group (applied with the exterior film preparation of control prepared according to the method of this example), and blank control group (applied with the exterior blank film preparation of this example). The clotrimazole cream group is administered 200ul per day for smearing skin lesions, the test group, the control group and the blank control group are directly applied with 1 piece of medical adhesive tape for fixing the medicinal film, the medicine is applied to the lesion site 1 time per day, and the medicinal film is replaced every day. After 4 days of administration, the curative effect is observed and checked every day, and recovery conditions of rash, scales or crusts at the lesion are recorded until 14 days. The healing is the regression of skin lesions, the negative result of the fungus microscopic examination is 2 times continuously and the negative result of the fungus culture is obtained; the ineffectiveness is that the skin lesion is not faded away and the fungus is positive by microscopic examination. And recording the cure number of each group of animals, calculating the cure rate, and observing the relapse condition of the cured animals after stopping the medicine.

The clotrimazole cream, the external film preparation of the Fuzhuan tea extract, the control external film preparation and the external blank film preparation are respectively administered to each component, and the treatment observation is carried out for 14d continuously, wherein the cure rates of 7, l0 and 14d are shown in table 9. The test statistics results in table 9 show that the external film preparation containing the fu tea extract has a significant curative effect, is superior to the positive control drug clotrimazole cream, and indicates that the external preparation containing the fu tea extract as an active ingredient has a good curative effect on a trichophyton mentagrophytes infection model of guinea pig.

Table 9: comparison of therapeutic effects of Trichophyton mentagrophytes in various groups on Guinea pig infection

	n	7d cure rate (%)	10d cure rate (%)	14d cure rate (%)
					External blank film agent	12	0	0	0
Clotrimazole cream group	12	16.67	25	33.33
					External film agent of Fuzhuan tea extract	12	50	83.33	91.67
Control topical film	12	0	0	0

As can be seen from the data in Table 9, the Fuzhuan tea extract external film agent has extremely outstanding and obvious curative effect.

Example 14

Animal systemic fungal infection experiment of Fuzhuan tea extract

Female C57BL/6 mice were selected as experimental animals (about 20 g) and administered 5X 10 by tail vein injection⁶Cryptococcus neoformans 0.1ml (5X 10) in CFU/ml concentration⁵CFU/ml) to create a systemic fungal infection model. Groups were then divided into 10 mice each, and treated groups were administered with the Fuzhuan tea extract (50mg/kg in PBS) prepared in example 3 once a day, the blank group was administered with PBS alone, and the control group was administered with Pu' er tea extract. The administration was performed for 5 days, and on day 5, mice were sacrificed, brains were collected, brain tissues were homogenized, the homogenate was diluted by a certain factor and added to a peptone agar-based smear, colonies on the medium were counted, and the amount of mouse brain fungi, which were charged, was calculated, and the results are shown in table 10.

Table 10: brain tissue derived colony count

	Relative colony (%)	SD	p value
				Blank group	100	5.51
Control group	106.91	7.52	p＞0.05
				Fuzhuan tea extract group	41.36	3.22	p＜0.05

As can be seen from Table 10, systemic administration of the Fuzhuan tea extract can effectively inhibit the growth of fungi in brain tissues, and the curative effect is obvious.

Example 15

Clinical curative effect of poria cocos tea extract for treating tinea pedis after soaking

Laboratory examination of 100 patients in the test group showed that they had erosive tinea pedis in summer (the patients' dandruff was digested with 10% KOH solution and examined microscopically, and there was hypha in the dandruff). The clinical results of 120 patients in the control group, summer erosive tinea pedis, were also diagnosed by laboratory examination, and there was no significant difference in the basic condition of the two groups of patients (P > 0.05).

The fu tea extract prepared in example 3 was dissolved in clear water at a mass volume ratio of 5% to obtain a liquid medicine. The patients in the test group are soaked in the liquid medicine for 0.5 hour, 1 dose for each time and 2 times for 1 day, and then are wiped dry after soaking. The control group was soaked in the same clean water without Fuzhuan tea extract, the protocol was the same as that of the test group, and 3% clotrimazole ointment was applied to the affected part. The treatment course is 7 days for both groups.

And (3) curing: the red and swollen part is completely removed after treatment, exudation stops, the erosion face is healed, the symptom disappears, and no skin damage is caused; the effect is shown: the area of the skin lesion is reduced by more than 2/3 after treatment, and the symptoms are obviously improved; the method has the following advantages: the symptoms are relieved, and the area of the skin lesion is reduced by more than 1/3; and (4) invalidation: after 2 treatment courses, symptoms and skin lesions are not obviously changed, and the curative effect is shown in table 11.

TABLE 11

As can be seen from the data in Table 11, the effect of treating tinea pedis by soaking with the Fuzhuan tea extract is quick, the method is simple and convenient, and the curative effect is satisfactory.

Although the present invention has been described with reference to a preferred embodiment, it should be understood that various changes, substitutions and alterations can be made herein without departing from the spirit and scope of the invention as defined by the appended claims.

Claims

1. An application of Fuzhuan tea extract in preparing antifungal medicine is provided.

2. The use of the Fuzhuan extract in preparing an antifungal agent according to claim 1, wherein the fungus is selected from the group consisting of Ascomycota, Basidiomycota, Deuteromycota, and zygomycota.

3. The use of an extract of Fuzhuan tea of claim 2, wherein the fungus is selected from the group consisting of Cryptococcus, Candida, Saccharomyces, Malassezia, Aspergillus, Coccidioides, Paracoccidiodes, Histoplasma, Blastomyces, and Neurospora crassa.

4. The use of the Fuzhuan extract according to claim 3, wherein the fungus is selected from the group consisting of Cryptococcus neoformans, Cryptococcus gatus, Saccharomyces cerevisiae, Candida glabrata, Candida albicans, Candida krusei, Candida tropicalis, Malassezia serrata, Candida parapsilosis, Trichophyton bailii, Malassezia furfur, Trichophyton rubrum, Trichophyton gypseum, Trichophyton floccosum, Microsporum gypseum, Microsporum canis, Pityrosporum, Candida.

5. The use of the Fuzhuan tea extract of claim 1, wherein the Fuzhuan tea extract is water soluble and/or fat soluble.

6. The use of the Fuzhuan tea extract of claim 5, wherein the Fuzhuan tea extract is supercritical CO₂Extract, organic solvent extract, aqueous extract or mixture thereof.

7. The use of Fuzhuan extract in preparing antifungal medicine according to claim 6, wherein the preparation method of the organic solvent extract comprises the following steps: grinding Fuzhuan tea into powder, wherein the diameter of the powder is not more than 0.5cm, adding 85% -95% ethanol into the Fuzhuan tea powder according to the solid-to-liquid ratio of 1:20, carrying out ultrasonic wall breaking until no tea powder and thalli can be seen by naked eyes, then carrying out cold soaking for 18-28 hours, collecting filtrate, repeating for 2-5 times, carrying out reduced pressure distillation on the filtrate, concentrating the filtrate, drying at low temperature to obtain a crude extract, adsorbing and decoloring the crude extract by macroporous resin, eluting by 85% -95% ethanol, concentrating and drying by distillation, and carrying out low-temperature freeze drying to obtain the organic solvent extract of the Fuzhuan tea.

8. The use of Fuzhuan tea extract in the preparation of antifungal drugs according to claim 6, wherein the preparation method of the water extract comprises the following steps: grinding Fuzhuan tea into powder, wherein the diameter of the powder is not more than 0.5cm, adding water into the Fuzhuan tea powder according to the solid-to-liquid ratio of 1:10, uniformly stirring, ultrasonically breaking the wall until no tea powder and thallus can be seen, standing for 2 hours at room temperature, and boiling for 2 hours in a boiling water bath to obtain a mixed solution; taking out the mixed solution, cooling, ultracentrifuging, and taking the supernatant; filtering and sterilizing the supernatant with 0.22 μm filter membrane to obtain water extract of Fuzhuan tea. Concentrating and evaporating to dryness if necessary, and freeze-drying at low temperature.

9. The use of the Fuzhuan tea extract according to claim 6 for preparing antifungal drugs,characterized in that the supercritical CO₂The preparation method of the extract comprises the following steps: grinding Fuzhuan tea into powder with diameter not more than 0.5cm, completely crushing tea powder and thallus, adding water vapor for pre-soaking until water content is increased to 40%, placing the pre-soaked Fuzhuan tea powder into an extraction tank, and continuously feeding CO into the tank₂Gradually extracting the crude extract with CO of the crude extract₂Sending to a cleaning tank, allowing the Fuzhuan tea extract to enter water phase, and passing through supercritical CO₂Extracting to produce a crude extract, hydrolyzing the starch and fiber with a mixture of cellulase and alpha-amylase, treating the mixture at 65 deg.C for 30 minutes to denature the enzyme, and centrifuging to remove impurities; removing residual protein with 15kDa ultrafiltration column, bleaching extract with activated carbon and filtering and recovering, sterilizing the product with 0.2 μm filter membrane, and lyophilizing or packaging to obtain Fuzhuan tea supercritical CO₂And (3) extracting.

10. The use of the fu tea extract of claim 1 for the preparation of an antifungal medicament, wherein the antifungal medicament is a fu tea extract as the only active ingredient, or a pharmaceutical composition comprising a fu tea extract;

the antifungal medicine is a product for inducing apoptosis of fungal cells, a product for inhibiting proliferation of fungal cells, a product for inhibiting synthesis of fungal ergosterol and a product for reducing germination rate of fungal spores.