Study On Extraction Of Quercetin And Its Glycosides In Guava Leaf By Microemulsion

ObjectiveGuava leaf is the leaf of Psidium guajava Linn..Guava leaf has astringent flavor and is neutral in nature,which was commonly used in astringing to stop diarrhea and diminishing inflammation to stop bleeding,on which there're many studies about flavonoids,such as quercetin,ursolic acid and volatile oil.Chinese traditional medicine always contains many effective components.Components with similar polarity were extracted by using single solvent according to the principle of similitude of polarity and solubility parameter.Microemulsion,with average particle at the nanometer scale,which is a highly dispersed system that can be propitious to contact the medicinal materials in its huge phase contact area,increases the extraction ratio of effective components.Microemulsion makes it possible to extract the water-soluble constituent and liposoluble constituent simultaneously,for it has several areas of different polarity,such as oil-phase,water-phase and oil-water interfacial film.The main content of the papers is the extraction of quercetin and its glycosides in Guava Leaf by microemulsion,which has the ability to dissolve the water-soluble constituent and liposoluble constituent simultaneously.Methods and Results1 the study of the analytical method to the extract of Guava Leaf by microemulsionTo the extract by microemulsion which was constitute by ethyl acetate-H2O-OP(oil phase/water phase=3:7,w/w),methodological study results showed that the standard Quercetin had a good linear correlation within the concentration of 0.606-36.36μg/mL, with correlation coefficient 0.9995(n=8).The analysis precision standard deviation was 0.0015,relative standard deviation was 1.17%(n=6),the repeatability precision standard deviation was0.20,relative standard deviation was 1.54%(n=6),the stability precision standard deviation was 0.18,relative standard deviation was 1.30%(n=9),the stability experiment revealed that the sample solution is stable in 48 hours.The average recovery was 99.53%,relative standard deviation was 1.46%(n=9),the recovery experiment showed that the recovery of Quercetin was fine.2 the study of screening the formulasFind out 100 microemulsion formulas which were stable,clarifying,with a large microemulsion area,which can be used in extraction.Established an analysis method detecting the Quercetin extracted from Guava Leaf by microemulsion with high-performance liquid chromatography(HPLC).Using the method of HPLC to detect the Quercetin in formulas,the results showed that, traditional solvent extraction methods were used to extracted quercetin in Guava Leaf,from which the content of quercetin extracted with methanol was the highest(4.72μg/mL).But the contents of Quercetin extracted by 72%of the formulas' were higher than methanol's, 98%of formulas' were higher than ethanol's,all formulas' were higher than ethyl acetate's. The formula with highest content(10.43μg/mL) of Quercetin which was constitute by ethyl acetate-30%isopropyl alcohol-RH60(oil phase/water phase=3:7,w/w),its content was more than double as the methanol's.The formula with lowest content(2.12μg/mL) of Quercetin which was constitute by n-hexane- 70%isopropyl alcohol-OP(oil phase/water phase=3:7,w/w),its content was more than the ethyl acetate's.Compared with traditional solvent extraction methods,microemulsion can effectively improve the extraction of quercetin from guava leaf.The results showed that,the fifteen formulas that can extract the higher content of quercetin in all 100 formulas,also extracted the higher content of its glycosides.Water and ethanol,which can extract more quercetin and its glycosides simultaneously than other traditional solvents,are compared with the formulas.Water,as a polar solvent,can extract amount of glycosides,but less quercetin.As to ethanol,the content of quercetin extracted with ethanol was higher than that with water,though it's still lower than that with 72%of all 100 formulas.But at the same time,the content of glycosides extracted with ethanol was lower than that with water.3 the study of optimizing the extraction technics,hydrolysis technics and prescription of microemulsoin ingredientsEthyl acetate- 30%isopropyl alcohol-RH60(oil phase/water phase=3:7,w/w) and ethyl acetate- 30%PEG400-OP(oil phase/water phase=7:3,w/w) were selected in this study.(1) In the study of extraction technics,the single factor test and orthogonal experiment design methods L₉(3⁴) were applied to analyze the influence of each factor in ultrasonic time and solid-liquid ratio on the extraction of Quercetin from guava leaf.The results showed that the optimum extraction technics of ultrasonic time and solid-liquid ratio is respectively 10min and 1:10.(2) In the study of hydrolysis technics,the single factor test and orthogonal experiment design methods L₉(3⁴) were applied to analyze the influence of each factor in ultrasonic time and the consistency of hydrochloric acid on the hydrolysis of quercetin's glycosides. The results showed that the optimum hydrolysis technics of ultrasonic time and the concentration of hydrochloric acid were respectively 15min and 3 mol/L. (3) Sample solutions were confected according to the results of optimizing the extraction technics and hydrolysis technics,the formula which was constituted by ethyl acetate- H₂O-OP(oil phase/water phase=3:7,w/w) with highest content of total weighted quercetin concentration was assigned as the working formula in the following study which was the preliminary separation of the microemulsion extraction.4 the study of the preliminary separation of the microemulsion extractionA method was established to separate the quercetin and its glycosides initially in the microemulsion extraction from the guava leaf by the silica gel column chromatography gradient eluted with chloroform- methanol-H2O as eluent.In this way,the quercetin and its glycosides were separated preferably.ConclusionThe study results showed that,compared with traditional solvent extraction methods, microemulsion can effectively improve the extraction of quercetin and its glycosides from guava leaf simultaneously.The contents of Quercetin extracted by microemulsion were higher than the traditional solvents'.And the quercetin and its glycosides were separated preferably by the silica gel column chromatography.