| Traditional Chinese medicine Salvia miltiorrhiza Bunge is the roots and rhizomes of Salvia miltiorrhiza bunge,which has an important effect in the treatment of cardiovascular diseases.The active components of Salvia miltiorrhiza are the key to its efficacy.The traditional extraction process has disadvantages such as low efficiency,high energy consumption and poor quality,so determining a set of green and efficient extraction process of active components of Salvia miltiorrhiza has important application value for realizing multi-level comprehensive utilization of Salvia miltiorrhiza resources.Enzyme assisted extraction hydrolyses plant cell walls using enzymes as catalysts to promote the diffusion of active ingredients from the intracellular extraction medium with accurate specificity,regioselectivity and the ability to react under mild conditions.Because of the poor solubility and easy inactivation of biological enzymes in traditional organic solvents,it is crucial to find a suitable solvent with good solubility and biocompatibility for enzyme assisted extraction and transformation of poorly water-soluble compounds.Deep eutectic solvents have many advantages such as low toxicity,green color,dynamic tunability,non flammability and strong solubility to compounds,and are considered as good alternatives to traditional organic solvents.Based on this,18 natural deep eutectic solvents(NADES)were prepared in this study,which were combined with biological enzymes,under ultrasound assisted extraction technology,to explore the extraction efficiency of this system for the active components of Salvia miltiorrhiza,then to use macroporous adsorption resin method for the enrichment and separation of tanshinone ⅡA in deep eutectic solvent extraction solution,and finally to evaluate the antioxidant activity of deep eutectic solvent extract through DPPH radical scavenging experiment.The main findings are as follows:1.The three species choline chloride(Ch Cl),betaine(Bet)and L-proline(L-Pro)are used as the hydrogen bond acceptors HBA;Six such as ethylene glycol(EG),triol(Gly),1,4-butanediol(1,4-bu),urea(U),lactic acid(LA),and glucose(D-Glu)were used as hydrogen bond donors to HBD.By ultrasonic heating method,the HBA and HBD components were synthesized to obtain 18 natural deep eutectic solvents,and their physicochemical properties were determined.The effects of different deep eutectic solvent components,component molar ratios,as well as solvent water contents on the extraction yields of active ingredients of Salvia miltiorrhiza were examined.The results showed that deep eutectic solvent synthesized from choline chloride: 1,4-butanediol at a molar ratio of 1:4 at a water content of 30% had the highest extraction yields for salvianolic acid B,tanshinone ⅡA and cryptotanshinone.2.Combined with ultrasound-assisted extraction technology,the best enzymatic hydrolysis system was selected as cellulase: pectinase(2:1).The effects of deep eutectic solvent and complex enzyme on the extraction rate of active components of Salvia miltiorrhiza were studied.The results showed that the optimal extraction system was 70% choline chloride :1,4-butanediol,30% citrate buffer(p H=5)and 1.5 mg/m L complex enzyme.Single factor test and response surface method BBD test were used to optimize the extraction process.The results showed that when the solid-liquid ratio was 1:22 g/m L,the enzyme concentration was 1.7mg/m L,the extraction temperature was 54℃,and the extraction time was 43 min,the yield of salvianolic acid B and tanshinone ⅡA reached 3.85% and 0.86%,respectively.NADES system has higher extraction efficiency,less solvent consumption and better economic benefits.3.The adsorption and desorption properties of tanshinone ⅡA from deep eutectic solvent extract were examined using five kinds of macroporous adsorption resins including AB-8,D-101,DM-130,HPD-100 and HPD-400 A.The results showed that HPD-400 A was enriched and separated the best.Under the optimal separation conditions,HPD-400 A showed high separation efficacy with 78.23% adsorption and 79.8% desorption of tanshinone ⅡA from deep eutectic solvent.The deep eutectic solvent after extraction was reused after cycled 3 times.The results showed that tanshinone ⅡA increments were almost consistent in the deep eutectic solvent extraction solution which was recycled 3 times,indicating that this deep eutectic solvent was stable and reproducible.4.Antioxidant activity of natural deep eutectic solvents extract of Salvia miltiorrhiza was evaluated by DPPH free radical scavenging experiment.The results showed that the antioxidant activity of natural deep eutectic solvents extract was higher than that of ethanol extract,and the free radical scavenging rate reached 90%,indicating that the prepared natural deep eutectic solvents extract had good antioxidant activity. |
