| Purpose: At present,Ginkgo biloba extract is the most widely used medical extract of plant Ginkgo biloba L.in Ginkgo,Ginkgoaceae.Ginkgo lactone components are the most active components except ginkgo flavonoids.The structure of ginkgo lactone components is very rare in natural products and is a unique structure in nature,unique to ginkgo.Ginkgolides can be divided into diterpenoids and sesquiterpenes.Ginkgolide A,B and C belong to diterpenoid compound and are platelet activating factor antagonists.Bilobalide is the only sesquiterpene compound,which has certain protective effect on the central nervous system and high medicinal value.It is clinically used for treating the central nervous system,improving cerebral ischemia,resisting platelet aggregation,resisting oxidation,resisting inflammation,resisting shock,etc.In 1991,Harvard University scholars won the Nobel Prize for discovering the molecular structure of ginkgolide B.However,the artificial synthesis of this kind of compounds costs a lot of cost and expense.At present,extraction and separation are still the main preparation methods.In the extraction and production process of ginkgo biloba leaves(taking the production process of ginkgo biloba ketone ester as an example in this study),the transfer rate of ginkgo flavonoids is about 80%,while the transfer rate of ginkgolides is only about 50%,and nearly half of lactone components are discarded.Every year,the industrial waste of ginkgo leaf extract reaches several thousand tons,causing waste of resources.Therefore,a preparation process of lactone components in ginkgo biloba extract industrial waste is established to obtain high-purity ginkgolide monomer compounds so as to improve the comprehensive utilization efficiency of ginkgo biloba resources.The prepared ginkgolides components can be further developed into related products to improve the utilization value and efficiency of ginkgo leaf resources,which have certain guiding significance for industrialization and green manufacturing,and conform to the development concept of circular economy.Methods: In this study,Ginkgolide A,ginkgolide B,ginkgolide C and bilobalide are used as target compounds.The content of ginkgolide in waste is determined by high performance liquid chromatography(HPLC-ELSD),and the parts with relatively high content are selected for enrichment,separation and purification.The main research methods are as follows: 1.Determination of lactone components in industrial waste of Ginkgo biloba extract: HPLC-ELSD method for determination of total lactone in industrial waste of Ginkgo biloba extract was established,and waste collection points with relatively high content were selected for enrichment and preparation process research.2.Establishment and optimization of process for enriching ginkgolides by macroporous adsorption resin column chromatography: the static and dynamic adsorption of ginkgolides by different types of macroporous resins were investigated,and the key process parameters such as diameter-height ratio,adsorption ratio,elution solvent and the like were investigated by single factor and orthogonal experiments to finally determine the optimal enrichment process.3.Establishment,optimization and structural confirmation of the preparation process of ginkgolides: investigate different chromatographic methods,establish the preparation process,investigate the main parameters of the process,and finally determine the optimal process.The structure of the target compound was confirmed by spectral method and literature comparison.Results: Firstly,the content of ginkgolides in different industrial waste parts was determined by HPLC-ELSD,and the parts with relatively high content of ginkgolides were selected: methanol was used as mobile phase A,water was used as mobile phase B,gradient elution was performed,and the flow rate was 1.0 m L/min;.The column temperature is 40?,the drift tube temperature of the evaporative light detector is 70?,and the carrier gas flow rate is 1.6 L/min.The detection results show that the contents of ginkgolides in S2(settling sediment)and S7(water eluent)waste parts are significantly higher than those in other parts.Therefore,S2 and S7 are selected for further enrichment and preparation process research.Secondly,macroporous adsorption resin column chromatography was used to enrich ginkgolides: the combination of static and dynamic adsorption was used to investigate different types of macroporous resins with the content(%)and recovery(%)of ginkgolides as indicators.The results showed that D-101 macroporous adsorption resin was suitable for enrichment of ginkgolides.Through orthogonal test,the optimum enrichment process parameters were obtained as follows: diameter to height ratio 1:5,adsorption ratio 1:50,eluent water and 60% ethanol respectively 5 column volumes eluted sequentially,60% ethanol eluate was vacuum concentrated and dried,the content at this time could reach about 20%,and the recovery rate could reach more than 80%.Finally,a series process of normal phase silica gel chromatography,recrystallization and reverse phase silica gel chromatography is adopted to investi GAte the main parameters of the process and determine the optimal process: taking normal phase silica gel column chromatography(200-300 mesh)with a diameter-to-height ratio of 1:10 and an adsorption ratio of 1:50,sequentially eluting 5 column volumes with dichloromethane: methanol(40:1 V/V),eluting 5 column volumes with dichloromethane-methanol(35:1 V/V),collecting and combining fractions containing ga,GB,GC and BB,dissolving the obtained fractions in 50% acetone for recrystallization,and the total lactone content reaches 80% Conclusion: In this project,the preparation technology of lactone components in ginkgo biloba extract industrial waste was established for the first time.The process is stable,feasible,simple in process equipment requirements,large in sample processing capacity,low in production cost,and has potential for industrial production and application.The research results of this subject fully utilize the effective components in the waste and realize industrialization,improve the utilization added value of the waste,make it turn waste into treasure,and at the same time provide valuable reference for optimizing the production process of ginkgo ketone ester bulk drug. |
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