| Candida albicans is the main pathogenic bacteria that causes oral candidiasis,and it is one of the main pathogenic microorganisms in nosocomial infections.Fluconazole is clinically preferred drug for the treatment of candida albicans,whose action mechanism is to inhibit the synthesis of ergosterol,an important structure on the cell membrane of fungi.In recent years,with the long-term high-dose use of this antifungal drug,the fluconazole-resistant Candida albicans has been clinically increasing year by year.At the same time,due to the narrow antibiotic spectrum of other traditional antifungal drugs,they are not difficult to produce drug resistance,and their toxic and side effects are limited,which limits their long-term clinical use.Therefore,it is urgent to develop a broad-spectrum,low-toxic,and high-efficiency new antifungal drug to fight fungal infections.The output of persimmons in China is extremely large.Due to its seasonal nature and impatience to storage,persimmons can be removed from astringent softening in the short term,so they are mostly stored in low temperatures.Studies have shown that tannins of persimmon have a strong antifungal activity.In this experiment,we studied the effects of persimmon water extracts on the synthesis of ergosterol from Candida albicans,and preliminarily explored its antifungal mechanism and played a positive role in the oral clinical application of tannin compounds.Part oneResearch on the effects of cryopreservation on persimmontannin contentObjective:Persimmon fruit is rich in tannin.The optimal conditions for the determination of persimmon tannin content by Phosphorus molybdenum tungstate-casein method were investigated.Taking the content of tannin as the index,the effect of cryopreservation on the content of tannin in the fruit of the green persimmon was observed,and the optimal extraction time of the tannin of the persimmon was discussed.Method:Usling ethly ethanol as the extracting agent,persimmon tannin were subjected to ultrasonic extraction,and crude persimmon tannin extracts were desugarized and then concentrated under reduced pressure.Using gallic acid as reference substance,the orthogonal test method was used to screen the optimal conditions for the determination of persimmon tannin content by phosphomolybdic tungstic acid-casein method.The casein method was used to determine the content of tannin in the persimmon fruit under different conditions.Result:1.The optimum conditions for the determination of persimmon tannin content by Phosphorus molybdenum tungstate-casein method were as follows:sodium carbonate solution concentration 20%,adding volume 12 mL,and color development time 30 min.2.The standard solution of gallic acid showed good linearity in the range of 0.002 mg/mL~0.012 mg/mL.The standard curve regression equation was y=114.44x-0.0403,and the correlation coefficient was R~2=0.9997.3.This method measured no significant changes in the content of tannin in frozen persimmon and fresh persimmon fruit(P>0.05).The content of total tannin in persimmon fruit was significantly lower than that in fresh persimmon fruit after 6 hours of thawing and 24 hours of thawing.The difference was statistically significant(P<0.01);and there was a significant difference in total tannin content between persimmon fruit after thawing for 6 h and thawing for24 h(P<0.05).Part two Effect of Persimmon Tannin Water Extract on the Synthesis ofErgosterol from Candida AlbicansObjective:The persimmon tannin water extract had a significant inhibitory effect on candida albicans.Ergosterol is an important component of the cell membrane of candida albicans,and it plays an important role in membrane integrity,membrane binding enzyme activity and so on.In this paper,we examined the effect of drug on C.albicans biosynthesis of ergosterol,the antibacterial mechanism of Persimmon tannin water extract on Candida albicans was preliminarily investigated.Method:Persimmon tannin trace dilution method is used to determine the water extract of candida albicans minimum bacteriostatic concentration(MIC),the ethanol-KOH method to extract the cell walls of candida albicans ergosterol content determination by ultraviolet spectropHotometry.Result:1.The minimum inhibitory concentration of persimmon tannin water extract and fluconazole was 84μg/mL and 1μg/mL respectively.2.The percentages of ergosterol in the blank group,the 42μg/mL and 84μg/mL persimmon tannin extracts and the 1μg/mL fluconazole-treated groups were 0.154±0.007,0.140±0.006,0.087±0.010 and 0.043±0.007 respectively.Compared with the blank control group,the final concentration of 42μg/mL,84μg/mL persimmon tannin group and 1μg/mL fluconazole treatment group all reduced ergosterol percentage,and the differences were statistically significant(P<0.01);final concentration of 84μg/mL compared with 42μg/mL of persimmon tannin water extract group,the percentage of ergosterol was significantly decreased,and the difference was statistically significant(P<0.01).Conclusion:1.There was no significant change in the content of tannin in persimmon fruit during cryopreservation,but the thawing process would destroy the tannin,and the tannin content showed a decreasing trend with the prolonged thawing time.2.Persimmon tannin water extract can exert antifungal effects by reducing the biosynthesis of Candida albicansergosteroland show a concentration dependence. |
