Aqueous Two-phase/Three-liquid-phase Extraction of Natural Products
|School||Dalian University of Technology|
|Keywords||Aqueous two-phase three-liquid-phase extraction natural product bioactive component|
Bioactive components of natural products are extremely rich, including flavones, saponins, alkaloids, terpenes, coumarins, steroidal, quinones and other compounds. Solvent extraction is usually used for the extraction and separation of natural products on laboratorial and industrial scales. However, the separation efficiency of this method is low and its cost is high. Some technical problems in separation of bioactive components in natural products, such as Vaccinium uLiginosum Linn, Salia miltiorrhiza Bunge and Capsicum annuum L., were investigated in this work as follows:To begin with, the partition behavior of anthocyanidin, capsaicin and capsanthin was studied in an aqueous two-phase system (ATPS) composed of hydrophilic short-chain organic solvent and inorganic salt. The extraction conditions were determined and optimized. When ATPS was composed of 30% (w/w) ethanol and 19% (w/w) ammonium suLfate, the partition coefficient (K), recovery (R) and yield(Y) for anthocyanidins was 7.06, 94.3% and 91.2%, respectively. For capsaicin and capsanthin, the optimum condition was 20% (w/w) ethanol and 20% (w/w) dipotassium hydrogen phosphate ATPS. Compared with ethanol extraction containing 0.5% (w/w) sodium carbonate and thermal reflux by n-hexane, the yield of capsaicin and capsanthin was 78.1% and 20.5%, respectively. These resuLts demonstrated that aqueous two-phase extraction (ATPE) was effective for the extraction of strongly polar bioactive components in natural products.Next, three-liquid-phase extraction (TLPE) of tanshinone IIA and salvianolic acid B in Salia miltiorrhiza Bunge and capsaicin and capsanthin in Capsicum annuum L. was investigated by using a three-liquid-phase system (TLPS) composed of hydrophobic organic solvent / hydrophilic organic solvent/inorganic salt / water. The effect of hydrophobic organic solvent on partition of bioactive components in TLPS was determined under their optimum conditions of ATPE. Furthermore, the operation parameters of TLPE were optimized. When TLPS was composed of 25% (w/w) n-hexane, 60% (w/w) ethanol extract of Salia miltiorrhiza Bunge and 15% (w/w) ammonium suLfate, the partition coefficient (K) and recovery (R) of tanshinone IIA and salvianolic acid B was 2.74 and 134.82, 89.0% and 97.2%, respectively. For capsaicin and capsanthin, the optimum condition was 10% (w/w) n-hexane, 18% (w/w) ethanol, 18% (w/w) dipotassium hydrogen phosphate and 54% (w/w) water TLPS. In this system, capsanthin was enriched in the top phase (n-hexane phase), while capsaicin was mainly in the middle phase (ethanol phase). A higher yield couLd be got by TLPE than traditional extraction. The results revealed that different polar bioactive components exhibited different partition behavior in TLPS. Weakly polar components are enriched to the top phase, whiles strongly polar bioactive components are mainly in the middle phase and water-soluble impurities are in the bottom phase. According to difference in polarity, TLPE can effectively separate bioactive components in natural product, and simplify the latter purification step.Furthermore, the ATPE and TLPE had been scaled up taking Salia miltiorrhiza Bunge extract for example. When the ATPS and TLPS were scaled up to 400 times of the original system, the extraction efficiency showed little change. This illuminated that ATPE and TLPE were suited for the industrial production of bioactive components in natural product.Finally, ATPE combined with column chromatography was examined to purify anthocyanidins. TLPE combined with column chromatography was for purification of salvianolic acid B. A new process for separation of bioactive components in natural product was put forward: pretreatment→ATPE/TLPE→column chromatography→drying—product. Compared with traditional methods, the new one is a simple and integrated process with low cost. It is especially fit for industrial production of bioactive natural products.