Study On Separation And Purification Process Of High Purity Rosmarinic Acid

Rosmarinic acid,which shows high academic research and industrial application value in medicine,food,health care products,skin care products and other fields,is a water-soluble natural antibacterial component,also has pharmacological functions such as anti-oxidation,anti-viral,anti-tumor and the like.In this thesis,the waste liquid of production was taken as raw material,which was came from the extraction process of oil-soluble antioxidants in rosemary.The rosemary acid was separated and purified by the means of ultrafiltration-resin,and the optimal parameters of production process were systematically studied;finally the high-purity product of rosemary acid was made by semi-preparative high performance liquid chromatography,and the optimal solvent system was investigated and the preparation process was optimized.The experimental results are summarized as follows:95%of rosmarinic acid was penetrated by the ultrafiltration membrane with pore diameter of 10k Da,at the same time,80.3%of the macromolecular impurity was retained in the waste solution,the optimal process parameters for the separation of rosinic acid are as follows:the dilution ratio of waste liquid/water is 1:5,the p H value is4,and the insoluble impuritiy is removed by centrifugation at a speed of 14000r/min for15min;the pressure of ultrafiltration is 0.4 MPa,and the stable time of ultrafiltration working is 80min for one time.The content of rosmarinic acid in the dry matter of the final filter solution raises to 39.7%,which is 17.7%higher than of the waste solution,and the yield is 95%.MCI（35-75μm）resin was selected for the purification of rosmarinic acid,the optimal parameters of production process are as follows:The maximum loading capacity is 19.756mg/g resin,the loading concentration is 1mg/m L,and the flow rate of loading is 2BV/h.The eluent was 60%ethanol,the flow rate of eluent is 2BV/h,the p H value of eluent is 4,and the time of eluent is 140min.Under the optimal conditions,sample with rosmarinic acid content of 67.9%is prepared,which is 28.2%higher than those obtained by ultrafiltration separation,and the yield reaches 90.77%.The optimum chromatographic conditions for the purification of rosemary acid by semi-preparative HPLC are as follows:C₁₈ reversed-phase preparative column,the detection wavelength is 280nm,column temperature is fixed at 30℃,the mobile phase is methanol（A）-0.1%acetic acid aqueous solution（B）,flow rate is 15m L/min,concentration of rosmarinic acid in sample is 6mg/m L,injection volume is lower than20 m L,condition of gradient elution:0～10 min:40%→60%A,60%B→40%B;10～15min:60%A→80%A,40%B→20%B;15～20min:80%A→100%A,20%B→0%B.The content of rosmarinic acid in the sample obtained after the preparation under this optimized condition is 92.47%,24.57%higher than the sample purifyed by MCI resin,and the yield is 99.9%.