CN110251499B - Dandelion extract and application thereof in preparing medicine for preventing and treating woman vaginal diseases - Google Patents
Dandelion extract and application thereof in preparing medicine for preventing and treating woman vaginal diseases Download PDFInfo
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- CN110251499B CN110251499B CN201910498473.1A CN201910498473A CN110251499B CN 110251499 B CN110251499 B CN 110251499B CN 201910498473 A CN201910498473 A CN 201910498473A CN 110251499 B CN110251499 B CN 110251499B
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- 229940067866 dandelion extract Drugs 0.000 title claims abstract description 43
- 235000020691 dandelion extract Nutrition 0.000 title claims abstract description 43
- 239000001845 taraxacum officinale leaf extract Substances 0.000 title claims abstract description 43
- 239000003814 drug Substances 0.000 title claims abstract description 15
- 208000013464 vaginal disease Diseases 0.000 title claims abstract description 10
- 241000222122 Candida albicans Species 0.000 claims abstract description 27
- 229940095731 candida albicans Drugs 0.000 claims abstract description 27
- NGSWKAQJJWESNS-UHFFFAOYSA-N 4-coumaric acid Chemical compound OC(=O)C=CC1=CC=C(O)C=C1 NGSWKAQJJWESNS-UHFFFAOYSA-N 0.000 claims abstract description 10
- MWDZOUNAPSSOEL-UHFFFAOYSA-N kaempferol Natural products OC1=C(C(=O)c2cc(O)cc(O)c2O1)c3ccc(O)cc3 MWDZOUNAPSSOEL-UHFFFAOYSA-N 0.000 claims abstract description 10
- REFJWTPEDVJJIY-UHFFFAOYSA-N Quercetin Chemical compound C=1C(O)=CC(O)=C(C(C=2O)=O)C=1OC=2C1=CC=C(O)C(O)=C1 REFJWTPEDVJJIY-UHFFFAOYSA-N 0.000 claims abstract description 9
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- 229940093681 4-coumaric acid Drugs 0.000 claims abstract description 5
- ZVOLCUVKHLEPEV-UHFFFAOYSA-N Quercetagetin Natural products C1=C(O)C(O)=CC=C1C1=C(O)C(=O)C2=C(O)C(O)=C(O)C=C2O1 ZVOLCUVKHLEPEV-UHFFFAOYSA-N 0.000 claims abstract description 5
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- KSEBMYQBYZTDHS-HWKANZROSA-N ferulic acid Chemical compound COC1=CC(\C=C\C(O)=O)=CC=C1O KSEBMYQBYZTDHS-HWKANZROSA-N 0.000 claims abstract description 5
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- 239000003480 eluent Substances 0.000 claims description 12
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- 241000245665 Taraxacum Species 0.000 claims description 5
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- KRZBCHWVBQOTNZ-RDJMKVHDSA-N (3r,5r)-3,5-bis[[(e)-3-(3,4-dihydroxyphenyl)prop-2-enoyl]oxy]-1,4-dihydroxycyclohexane-1-carboxylic acid Chemical compound O([C@@H]1CC(O)(C[C@H](C1O)OC(=O)\C=C\C=1C=C(O)C(O)=CC=1)C(O)=O)C(=O)\C=C\C1=CC=C(O)C(O)=C1 KRZBCHWVBQOTNZ-RDJMKVHDSA-N 0.000 abstract description 4
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- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
- A61K31/19—Carboxylic acids, e.g. valproic acid
- A61K31/192—Carboxylic acids, e.g. valproic acid having aromatic groups, e.g. sulindac, 2-aryl-propionic acids, ethacrynic acid
-
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- A61K31/21—Esters, e.g. nitroglycerine, selenocyanates
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- A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
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- A61K36/28—Asteraceae or Compositae (Aster or Sunflower family), e.g. chamomile, feverfew, yarrow or echinacea
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Abstract
The invention discloses a dandelion extract and application thereof in preparing a medicine for preventing and treating female vaginal diseases, wherein the dandelion extract mainly comprises 4-coumaric acid, ferulic acid, quercetin pentaglucoside, 3, 5-di-O-caffeoylquinic acid, 4, 5-di-O-caffeoylquinic acid and luteolin. The test result shows that the dandelion extract has good inhibition effect on candida albicans and gardnerella, the minimum inhibitory concentration on the candida albicans and the gardnerella is respectively 0.4mg/mL and 0.2mg/mL, and the dandelion extract can be used for preparing the medicine for preventing and treating the vaginal diseases of women caused by the candida albicans and the gardnerella. The dandelion extract has the advantages of convenient material acquisition of raw materials, low production cost, good product quality, low price, greenness and environmental protection, and is suitable for popularization and application. Therefore, the traditional Chinese medicine can be used as a novel traditional Chinese medicine for treating the vaginosis.
Description
Technical Field
The invention belongs to the technical field of medicines, and particularly relates to a dandelion extract and application of the dandelion extract in preparing a medicine for preventing and treating female vaginal diseases caused by candida albicans and gardnerella.
Background
Vaginitis is also known as vaginitis, and is a common and frequent symptom in gynecologic outpatient clinic. The female healthy vagina has natural defense capacity against invasion of pathogenic microorganisms such as bacteria, fungi and the like, and when the natural defense system of the healthy vagina is damaged, pathogens enter while being deficient, the original ecological balance between flora and the vagina is broken, and certain pathogens rapidly grow under the proper growth condition, so that vagina inflammation is caused.
Because the diagnosis in clinical work is incorrect, the treatment standard understanding is not thorough, new vaginal infection and mixed infection are not known enough, women in daily life lack comprehensive and correct knowledge of gynecological diseases, relative self-health care, various bad life work and rest and the like, the physiological health is gradually improved, so that some women are wonderful and fascinated all the year round, and the women are recurrent or not cured for a long time, and the inconvenience is brought to the normal work, study and family life of the individual.
Clinically common vaginosis are: bacterial vaginosis, candida vaginosis, trichomonas vaginosis, senile vaginosis and juvenile vaginosis. Wherein, the bacterial vaginosis and the candida vaginosis are two common vaginosis which are respectively mainly caused by gardner bacteria and candida albicans. In recent years, abuse of antibiotics causes many biological safety problems such as drug resistance of bacteria and fungi, so that the search for a natural medicinal component with good antibacterial property as a vaginal disease treatment medicament is extremely important.
Disclosure of Invention
The invention aims to provide a dandelion extract and provide a new application of the dandelion extract.
Aiming at the purposes, the dandelion extract comprises the following components in percentage by weight: 11 to 12 percent of 4-coumaric acid, 3.5 to 4.5 percent of ferulic acid, 10 to 11 percent of quercetin pentaglycoside, 34 to 35 percent of 3, 5-di-O-caffeoylquinic acid, 3.5 to 4.5 percent of 4, 5-di-O-caffeoylquinic acid and 11 to 12 percent of luteolin.
The dandelion extract is obtained by the following method:
(1) Water extraction
Crushing the dried whole dandelion herb, sieving the crushed whole dandelion herb by a sieve of 60-80 meshes, adding distilled water according to a material-liquid ratio of 1g.
(2) Preliminary purification
And (3) carrying out suction filtration on the supernatant by using a filter membrane with the aperture of 0.45 mu m, then carrying out ultrasonic treatment for 5-10 min, then loading the supernatant on a pretreated D101 macroporous adsorption resin column, washing the column for 2 column volumes, eluting the column volume by using an ethanol aqueous solution with the volume concentration of 50-70%, collecting the eluent, and concentrating the eluent by 4-6 times.
(3) Purifying again
Extracting the concentrated eluent with ethyl acetate of the same volume, collecting the extract, concentrating by 5-6 times, and drying in vacuum to obtain the dandelion extract.
The dandelion extract is used for preparing the medicine for preventing and/or treating the female vaginal diseases caused by candida albicans and gardnerella. Test results show that the dandelion extract has good inhibition effect on candida albicans and gardnerella, the minimum inhibitory concentrations on the candida albicans and the gardnerella are respectively 0.4mg/mL and 0.2mg/mL, and the dandelion extract can be used for preparing medicines for preventing and treating women vaginal diseases caused by the candida albicans and the gardnerella.
The dandelion extract has the advantages of convenient raw material acquisition, low production cost, good product quality, low price, greenness and environmental protection, and is suitable for popularization and application. Therefore, the medicine can be a novel pure traditional Chinese medicine for treating the vaginosis.
Drawings
FIG. 1 is a HPLC analysis chart of the dandelion extract obtained in example 1, wherein 1 is 4-coumaric acid; 2 is ferulic acid; 3 is quercetin pentaglycoside; 4 is 3, 5-di-O-caffeoylquinic acid; 5 is 4, 5-di-O-caffeoylquinic acid; 6 is luteolin.
FIG. 2 shows the Candida albicans inhibition test of the dandelion extract obtained in example 1.
FIG. 3 is a test of the inhibition of Gardner bacteria by the dandelion extract obtained in example 1.
Detailed Description
The present invention will be described in further detail with reference to the drawings and examples, but the scope of the present invention is not limited to these examples.
Example 1
Preparation of Taraxacum Mongolicum extract
(1) Water extraction
Drying fresh dandelion whole herb to constant weight in vacuum at 60 ℃, crushing after drying, sieving by a 80-mesh sieve, adding distilled water according to a material-liquid ratio of 1 g.
(2) Preliminary purification
And (2) performing suction filtration on the supernatant collected in the step (1) by using a filter membrane suction filter with the aperture of 0.45 mu m, performing ultrasonic treatment for 10min, then putting the supernatant on a pretreated D101 macroporous adsorption resin column, washing the column by 2 column volumes, eluting the column by using an ethanol aqueous solution with the volume concentration of 60%, collecting eluent, and concentrating the eluent by 4-6 times.
(3) Purifying again
Extracting the eluent concentrated in the step (2) by using ethyl acetate with the same volume, collecting extract liquor, concentrating by 5-6 times, and drying in a vacuum drying oven at 60 ℃ to obtain the dandelion extract.
The composition of the obtained dandelion extract was determined by High Performance Liquid Chromatography (HPLC), the HPLC system consisting of an Agilent1100HPLC system equipped with a DiKMADIAMONSil C18 column (250 mm. Times.4.6 mm,5 μm) and a Diode Array Detector (DAD). The sample was dissolved in 80 vol% aqueous methanol, the column temperature was maintained at 40 ℃, the mobile phase consisted of acetonitrile (eluent A) and 0.1 vol% aqueous phosphoric acid (eluent B), the isocratic eluent flow rate was 1.0mL/min, the elution was carried out at 20-80% concentration for 47min, and the sample size was 10. Mu.L. The peak area of all polyphenolic compounds was determined at a wavelength of 280 nm. As shown in FIG. 1, 6 compounds, namely 4-coumaric acid, ferulic acid, quercetin valeryl glycoside, 3, 5-di-O-caffeoylquinic acid, 4, 5-di-O-caffeoylquinic acid and luteolin, were detected by HPLC, and the relative percentage content of the 6 compounds in the extract was 11.45%, 3.96%, 10.48%, 34.24%, 3.91% and 11.80%, respectively.
Example 2
The inhibition of Candida albicans and Gardnerella by the Dandelion extract of example 1
(1) Test for bacteriostatic Activity
The bacteriostatic action of the dandelion extract on Candida albicans (Candida albicans, CMCC 85021) and Gardnerella vaginalis (ATCC 14018) was determined by a paper sheet method. The experimental strain was revived and activated 2 times. The Candida albicans is subcultured in a sand type liquid culture medium and cultured for 12h at 37 ℃ and 190 r/min; gardnerella using HTM agar medium plates at 37 ℃ 5% CO2Culturing for 12h under the condition, and washing the bacterial colony by using a liquid culture medium to prepare bacterial suspension. The OD of each cell was measured with a spectrophotometer600nmThe value is obtained. When OD is reached600nmAn optimum coating concentration is 0.6 to 0.7; if OD600nm>At 0.7, the cells were diluted with a liquid medium and applied. Preparing a sand agar culture medium and an HTM agar culture medium, sterilizing at 115 ℃ and 121.3 ℃ for 20min respectively, cooling a sterile coenzyme I (NAD) solution in the culture medium to 50 ℃, adding the HTM agar culture medium, mixing uniformly, and preparing plates respectively. And sucking 100 mu L of activated bacterial suspension with proper concentration on the surface of the plate by using a liquid transfer gun, and uniformly coating the bacterial liquid by using a sterile L-shaped glass rod. Experimental groups: 10 μ L of the sample solution dissolved in 40% dimethyl sulfoxide was pipetted and dropped onto a 6mm diameter sterile filter paper sheet, baked appropriately and attached to the surface of the plate. Blank group: 10 μ L of sterile 40% dimethyl sulfoxide (DMSO) was added dropwise to sterile 6mm diameter filter paper sheets, baked appropriately, and applied to the surface of the plates. Respectively at 37 deg.C and 5% CO2And (4) carrying out inverted culture for 18-36 h under the condition, observing an experimental result, and measuring the diameter of the inhibition zone.
The test results showed that there was essentially no zone of inhibition around the blank group 40% dmso paper, indicating that dimethyl sulfoxide had no effect on fungal and bacterial growth; the periphery of the dandelion extract in the experimental group has an antibacterial ring, which shows that the dandelion extract has obvious antibacterial effect on candida albicans and gardnerella (see figure 2). When the concentration of the dandelion extract is 0.40-2.28 mg/mL, the diameter of the bacteriostatic circle of the candida albicans is 6.00-10.53 mm, and when the concentration of the dandelion extract is 2.28mg/mL, the diameter of the average bacteriostatic circle of the candida albicans is 10.38 +/-0.15 mm; when the concentration of the dandelion extract is 0.20-3.80 mg/mL, the diameter of the inhibition zone of Gardner bacteria is 6.00-11.85 mm, and when the concentration of the dandelion extract is 3.80mg/mL, the diameter of the average inhibition zone of Candida albicans is 11.86 +/-0.17 mm. The dandelion extract has good inhibition effect on candida albicans and gardnerella.
(2) Minimum inhibitory concentration
Culturing Candida albicans and Gardnerella according to the method (1), adjusting the concentration of the bacterial liquid to 10 with liquid culture medium7CFU/mL, and preparing 4mg/mL dandelion extract mother liquor. Adding herba Taraxaci extract mother liquor into test tubes containing 2mL liquid culture medium respectively to make the final concentration of herba Taraxaci extract in the culture medium be 0, 0.05, 0.10, 0.20, 0.40, 0.60, 0.80mg/mL, inoculating at 4% (v/v) concentration of 107CFU/mL bacterial solution, and paraffin Vaseline (V) in Gardnerella test tubeParaffin oil:VVaselineMix = 1) the liquid level was sealed. Culturing Candida albicans and Gardnerella at 37 deg.C, 190r/min and 37 deg.C for 24 hr, respectively, when OD of sample is600nmThe concentration of the dandelion extract which changes for the first time by less than 5 percent is the minimum inhibitory concentration, namely MIC. Analysis shows that when the concentration of the dandelion extract is more than 0.4mg/mL, the Candida albicans OD600nmThe change of the value is less than 5% for the first time, and when the concentration of the dandelion extract is more than 0.2mg/mL, the Gardnerella OD600nmThe value of the change is less than 5% for the first time, and the MIC values of the dandelion extract to the Candida albicans and the Gardnerella respectively are 0.4mg/mL and 0.2mg/mL.
As can be seen from the above, the dandelion extract has good inhibition effect on candida albicans and gardnerella, the MIC values on candida albicans and gardnerella are respectively 0.4mg/mL and 0.2mg/mL, and the dandelion extract can be used for preparing medicines for preventing and/or treating the female vaginal diseases caused by candida albicans and gardnerella.
Claims (1)
1. The application of dandelion extract in preparing medicine for preventing and/or treating women's vagina diseases caused by candida albicans is characterized by that:
the dandelion extract comprises the following components in percentage by weight: 11 to 12 percent of 4-coumaric acid, 3.5 to 4.5 percent of ferulic acid, 10 to 11 percent of quercetin pentaglucoside, 34 to 35 percent of 3, 5-di-0-caffeoylquinic acid, 3.5 to 4.5 percent of 4, 5-di-0-caffeoylquinic acid and 11 to 12 percent of luteolin;
the dandelion extract is obtained by the following method:
(1) Water extraction
Crushing the dried whole dandelion herb, sieving the crushed dandelion herb with a 60-80-mesh sieve, adding distilled water according to the material-liquid ratio of 1g to 25-35 mL, leaching for 30-40 min at 50-70 ℃, leaching for 2 times, filtering, combining the filtrates, concentrating the filtrate by 2-3 times, performing centrifugal separation, and collecting the supernatant;
(2) Preliminary purification
Carrying out suction filtration on the supernatant by using a filter membrane with the aperture of 0.45 mu m, then carrying out ultrasonic treatment for 5-10 min, then putting the supernatant on a pretreated D101 macroporous adsorption resin column, washing the column by 2 column volumes, eluting the column by using an ethanol aqueous solution with the volume concentration of 50-70%, collecting eluent, and concentrating the eluent by 4-6 times;
(3) Purifying again
Extracting the concentrated eluent with ethyl acetate of the same volume, collecting the extract, concentrating by 5-6 times, and drying in vacuum to obtain the dandelion extract.
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