Research On The Extraction Conversion Of Jujuboside From Ziziphi Spinosae Semen And Their Antioxidant Activities

Ziziphi Semen Spinosae?ZSS?has a high nutritional and medicinal value.It has the effect of anti-lipid peroxidation and bacteriostasis.It is closely related to jujuboside,flavonoids,alkaloids,polysaccharides,unsaturated fatty acids and polyphenols in ZSS.The high biological activity of jujuboside has been widely and deeply studied.In order to improve the extraction rate of jujuboside,the extraction and purification method of jujuboside was changed from traditional single type extraction and purification to integrated extraction and purification.The extraction efficiency of ZSS was greatly improved under the integrated extraction and purification model.Pharmacological and biological activity of jujuboside were lower than jujube;jujube obtained only by acid or bioconversion.The former leads to environmental pollution and increases the cost of extraction,and the latter reduces the utilization of jujuboside.To solve these problems,pressure and ultrasound-assisted solvent extraction?PUSE?was used to extract jujuboside from ZSS in this paper.The breaking of glycosidic bond and acetal bond in jujuboside was achieved by hydrothermal method.The conversion of jujubes was achieved by selective oxidation of Fe Mn catalyst.Silica gel chromatography was used to separate and purify the the total jujuboside obtained by PUSE.Combination of related structural characterization,their antioxidant mechanisms were studied in detail.The main contents and results include:?1?The thermal decomposition behaviors of powder and ethanol extract of Ziziphi Spinosae Semen?ZSS,a traditional Chinese medicine?in air atmosphere were investigated by TGA and DSC.It was found that the thermal stability of active ingredients was stronger than powder,and the decomposition temperature of jujuboside exceeded 240°C.The thermal decompositions of both powder and ethanol extract of ZSS belong to zero-order reaction.Furthermore,kinetic parameters?the activation energies?E?,pre-exponential factors?A??of the thermal decompositions of them based on the DSC curves obtained at different heating rates were calculated following three different methods,the Kissinger method,the Friedman-Reich-Levi?FRL?method and the Flynn-Wall-Ozawa?FWO?method.The results show that the correlation coefficient of thermal decomposition of ZSS ethanol extract is better than its powder.R²_Kissinger?0.9968?and R²_FWO?0.9833?were larger than R²_FRL?0.9684?,it indicated that Kissinger and the FWO methods are more suitable than FRL to describe the thermal degradation behavior of powder and ethanol extract.The Kissinger method obtains the average values of reaction kinetic parameters;the FWO method obtains the information of entire decomposition process??:0.05–0.95?under linear heating rate conditions.Whereas the calculating results of the FWO method are more accurate than FRL when the ethanol extract decomposition percent conversion is between 0.05 and 0.8.FWO is more suitable for describing the thermal degradation kinetics of ethanol extract.The less the components,the closer to the real value the kinetic parameters obtained by Kissinger and FWO methods.The isothermal weightlessness times of powder and ethanol extract were long at room temperature.The weightlessness times of315.5 K is 6.5 times less than that of room temperature.The active components were thoroughly decomposed when the temperature reaches 513.15 K.The test results of TGA and DSC can well master the thermal stability of drgus and calculate the kinetic parameters of their thermal degradation.It can be used as the main technology for drug production and evaluation.?2?The total jujubosides were separated from crude extract of ZSS by using nonpolar macroporous resin?D-101?.The macroporous resin?D-101?was pretreated by ultrasound,vacuum drying and immersing in methanol.The separation and purification behavior of resin was analyzed.The resin is characterized by SEM,BET and DSC.Composition and its concentration are measured by spectrophotometry and mass spectrometry,and the absolute configuration of jujuboside A is calculated by quantum chemistrry methods.The results show that the pore size and water content in the resin are the main factors affecting the purify of jujuboside,which are controlled by ultrasonic and moisture control.Adsorption and desorption between jujuboside and resins are also affected by the spatial conformation of jujuboside.The content of total jujubosides is more than 43%after purification by pretreatment resin,and the optimal pretreatment process of the resin was that the resin of 21%water content was treated under 175W for 30 min in the methanol.?3?Pressure and ultrasound-assisted solvent extraction?PUSE?,a new extraction technology,was used to extract jujuboside from Ziziphi Spinosae Semen?ZSS?and the transformation of jujuboside structure was performed synchronously with the extraction.Compared with CSE,USE and PSE,PUSE showed the distinctive extraction power,the highest extraction yield and content of jujuboside.The synergic effect of ultrasound,temperature and pressure in PUSE could have enhanced broken intensity of cell walls by SEM.Basing on the results of extraction and combining with transformation of jujuboside structure,the optimal conditions of the PUSE were following:powder particle size 0.18-0.25 mm,ethanol concentration 80%,extraction temperature 130 ^oC,extraction pressure 4 atm,ultrasound power 480 W and extraction time 60 min,iron oxide?0.01%?and manganese ions(10^-3 mol/L).Under these conditions,the extraction yield and content of jujuboside achieved 30.46±0.46%and 30.82±0.40 mg/g,which are obviously better than 22.65±1.05%and 9.55±0.61 mg/g of the heat reflux method,14.92±0.72%and 11.62±1.21 mg/g of the ultrasonic extraction method and 26.97±0.90%and 13.24±1.41 mg/g of the high pressure extraction method.?4?New mixtures were structurally characterized by high performance liquid chromatography?HPLC?,thin layer chromatography?TLC?,Fourier transform-infrared spectroscopy analysis?FT-IR?.HPLC and TLC showed that new mixtures obtained by PUSE different from mixtures obtained by CSE,USE and PSE.The FT-IR spectrum proved the new mixtures characteristic band:Acetal bond and glycoside bond disappeared?Hydrothermal method?and conjugated double bonds and unsaturated ketones at the ends of conjugated double bond chains appeared?Ferromanganese base catalyst?.The results indicated that the total antioxidant activity and total reductive activity of the jujuboside were positively related to the content of the reductive functional group and oxidizing functional group.Jujuboside obtained by PUSE existed in the form of allyl free radical in the free radical scavenging process,which could effectively remove the OH and DPPH free radicals.But their ability to scaveng OH radicals of jujuboside obtained by PUSE is much stronger than DPPH radicals.The reasons are as follows:first,the space hindrance of the DPPH radical is larger than that of the OH radical.Secondly,the space hindrance effect leads to a significant reduction in the effective active sites of the free radical of saponins.Finally,the free radicals can not be realized at the termination stage of the free radical chain reaction.The formation of free radicals is terminated by effective collision.At the same time,under the condition of PUSE,ester bond of linoleic ester breaks up and a lot of linoleic acid were produced.