Study On The Separation Of RA By Solvcnting-out Crystallization And Extraction

Steviol glycosides are a group of diterpene glycoside isolated from the leaves of Steviarebaudiana Bertoni. Amongst the steviol glycosides, stevioside, rebaudioside A andrebaudioside C are the main components. Because of their high sweetness and lower calorificvalue, steviol glycosides has great advantage of natural sweeteners for diabetic patients,obesity group and the old group. The taste quality of rebaudioside A close to sucrose and beused as ideal sweeter. This thesis unfolds around isolate high purity rebaudioside A, studiedon the separation of rebaudioside A by solventing-out crystallization and liquid-liquidextraction. Besides, the analytical method for determination of rebaudioside A by thin layerchromatography and dissolving selectivity of solvents on rebaudioside A were preliminarilyinvestigated. The main content as follows.1. Established the analytical method for determination of rebaudioside A by thin layerchromatography. The results showed that developer was isopropanol/ethyl acetate/acetone/water （30/52/3/15）, steviol glycosides were divided into two parts, one was RA, the otherwas X, resolution was2.4. Visualization reagent was aniline/diphenylamine/phosphoric acid/methanol （2ml/2g/10ml/100ml）, scanning wavelength of rebaudioside A spots was330nm.The sampling amount of rebaudioside A from2×10^-5g to4.5×10^-5g showed linear correlation,the correlative equation was Y=8776+4.1×10⁸X （R²=0.96）. The average recovery was108.8%,relative error was8.8%. The coefficient of variation in reproducibility test was3.9%.2. The dissolving selectivity of solvents on rebaudioside A were investigated. The resultsshowed that the higher concentration of rebaudioside A in solution correspond to the highervalue of separation coefficient of rebaudioside A and other components in steviol glycosidesin the dissolution process. Dissolving selectivity of methanol and ethanol on rebaudioside Aweaker than other alcoholic solvents. Solubility of steviol glycosides in alcoholic solventdecreased with decreasing polarity of the solvent. Separation coefficient of rebaudioside Aand other components in steviol glycosides in the crystallization process of steviol glycosidesmethanol solution from40g/L to120g/L was less than1. Separation coefficient ofrebaudioside A and other components in steviol glycosides in the dissolution process steviolglycosides by alcoholic solvents polarity less than methanol was more than1.3. Studied on the separation of rebaudioside A by solventing-out crystallization. Theresults showed that with the level changes of each factor, the trend of yield of steviolglycosides was contrary to separation coefficient of RA and other components in steviolglycosides. Separation coefficient of rebaudioside A and other components in steviolglycosides decreased and then increased with ethyl acetate content increasing at40%⁶0%.Separation coefficient of rebaudioside A and other components in steviol glycosides decreasedwith the steviol glycosides initial concentration increasing at20g/L⁴0g/L, crystallizationtemperature increasing at8℃²8℃,crystallization time increasing at6h¹4h. Under theoptimal conditions, that is, steviol glycosides initial concentration was20g/L, crystallizationtemperature was18℃,crystallization time was8h, ethyl acetate content was50%, the yield ofsteviol glycosides was34%and separation coefficient of rebaudioside A and other components in steviol glycosides was3.8.4. Studied on the separation of rebaudioside A by liquid-liquid extraction. The resultsshowed that with the level changes of chloroform content and oil water ratio, the trend ofyield of rebaudioside A was contrary to separation coefficient of rebaudioside A and othercomponents in steviol glycosides in the extraction process. Separation coefficient ofrebaudioside A and other components in steviol glycosides decreased with the chloroformcontent increasing at20%⁶0%. Separation coefficient of rebaudioside A and othercomponents in steviol glycosides increased with the oil water ratio increasing at5∶7⁷∶5.Yield of rebaudioside A and separation coefficient of rebaudioside A and other components insteviol glycosides decreased and then increased with the concentration increasing at0.6g/L¹.4g/L. Yield of rebaudioside A and separation coefficient of rebaudioside A and othercomponents in steviol glycosides increased and then decreased with the extractiontemperature increasing at3℃²3℃. Under the optimal conditions, that is, chloroform contentwas40%, concentration was1.5g/L, oil-water ratio was5∶5, extraction temperature was18℃,the yield of rebaudioside A was78%and separation coefficient of rebaudioside A andother components in steviol glycosides was1.8.