Study Of Emulsification's Effect On The Quality Of Angelica Volatile Oil And Its Emulsifying Mechanism

BackgroundVolatile oil of traditional Chinese medicine(TCM),which derived from aromatic TCM,is a kind of volatile oily liquid.It can be distilled with steam and is immiscible with water.As an important intermediate of TCM preparation,pharmaceutical enterprises generally extract volatile oil using steam distillation.However,the unexpected emulsification phenomenon in the extracting process makes the extraction rate low,the product quality would be affected,and finally the emulsion needed to be further treated after extraction.This is a common problem in the modern preparation process of TCM,and also one of the bottleneck problems in the industrialization and modernization of TCM.It is of great theoretical significance and practical value to explore the formation mechanism of volatile oil emulsification phenomenon and find effective oil-water separation technology,to improve the oil extraction rate of volatile oil,ensure the quality of volatile oil and realize the oil-water separation successfully.Angelica sinensis,an aromatic TCM,which is used frequently and in large dosage.Volatile oil is its main pharmacodynamic basis.Due to the serious emulsification phenomenon in the process of steam distillation,the extraction rate of angelica volatile oil was low or even could not be collected,the quality of volatile oil was affected,and the emulsification mechanism was not clear.To solve these problems,to forecast the quality marker as the basis of the follow-up research,combine with the physicochemical properties and chemical composition to clarify the impact on the quality of volatile oil,take the emulsion in the extraction process of angelica volatile oil as the analysis object to find the emulsification rule,and use molecular simulation technology to explore the internal mechanism.Finally,the appropriate method is selected to reduce the emulsification phenomenon and realize the oil-water separation.The purpose of this study provides research reference for the emulsification of volatile oil in traditional Chinese medicine and promote the emulsification solution.Object1.To predict the quality marker(Q-marker),and provide effective evaluating indicator for the quality evaluation of angelica volatile oil.2.To evaluate the effect of emulsification on the quality of angelica volatile oil and provide a data support for emulsification research.3.To investigate the relationship between external factors and internal components of emulsification and explore the emulsification rule and material basis of angelica volatile oil.4.To investigate the role of emulsification material base in volatile oil emulsion via the molecular simulation technology and explain the mechanism of emulsification.5.To verify the emulsification rule and find effective and feasible emulsification method.Methods1.To predict the Q-marker of angelica volatile oil,angelica volatile oil was extracted by steam distillation,and the chemical components were analyzed via GC-MS.The co-existing components from multiple batches of angelica volatile oil were used as candidate components of Q-marker.By using network pharmacology,the targets of common component were searched in public database.By constructing the network of component-target,protein-protein interaction,component-target-pathway,and analyzing the enrichment of G0 and KEGG,to get the chemical components which were most closely related to multi-target,multi-channel,multi-organization as the Qmarker of angelica volatile oil.Finally,the biological activity of Q-marker was explained and confirmed by using molecular docking and consulting the literature.2.The volatile oil extracted from angelica sinensis by steam distillation of small-scale experiment was used as control,the effect of emulsification on angelica sinensis volatile oil was evaluated from three aspects: extraction rate,physicochemical properties including relative density,aroma,properties,refractive index,optical rotation,ethanol miscibility and acid value,and chemical components were analyzed by GC-MS qualitative analysis and quantitative analysis of Q-marker.3.Taking the temperature,turbidity,and chemical composition of the emulsion in the extraction process of angelica volatile oil as the research focus,using the partial least squares(PLS)method to fit and explain the influence of temperature on chemical composition of the emulsion,chemical composition on turbidity,and the relationship between temperature and turbidity.At the same time,the change of Q-marker content in the emulsion was analyzed to find the emulsification rule.The chemical components positively correlated with turbidity were screened out and combined with the hydrophile-lipophile balance(HLB)value,material basis of emulsification was determined according to the HLB value range of surfactant.4.Molecular simulation technology and molecular dynamics were used to simulate the surface tension and Gibbs free energy of various concentrations of emulsified key component to investigate its surface properties.By simulating the effect of emulsified key component on the surface tension of volatile oil/water system,observing the dispersion and aggregation rules of the system by the polar surface area and radius of gyration of solvent,and analyzing the change of solubility of volatile oil in water and the cause of solubility,to explain the mechanism of emulsification.5.The influence of external factors and internal components on emulsification was tested to confirm the emulsification rule.The effective and feasible demulsification methods were explored from the following three aspects: the influence of temperature on extraction rate was investigated by controlling condenser temperature in extraction process,turbidity change of emulsion was investigated by introducing new material porous-low-density-polystyrene and studying on oil-water separation of emulsion by coalescence method.Results1.There were 18 batches of volatile oil derived from radix angelica sinensis in Gansu,Yunnan,and Qinghai province,and five common components were identified by GCMS.Through network pharmacology,112 targets were obtained by searching database and the network was constructed and analyzed.Trans-ligustilide and 3-butylidene-1(3H)-isobenzofuranone were considered as Q-marker of angelica volatile oil.The pharmacodynamic mechanism of Q-marker was as follows: they acted on the core targets PIK3R1,CHRM1,CHRM2,SRC,CREBBP,via the regulation of actin cytoskeleton,and mainly distributed in cerebral cortex,gallbladder,smooth muscle,hippocampus and other tissues,the highest expression level was found in the cerebral cortex.Molecular docking technology verified the biological activity of the Q-marker,whether compared with the control group,or the affinity value of Q-marker with targets less than-4 kcal/mol,which showed good evaluation results of Q-marker.At the same time,some literatures confirmed biological activities of trans-ligustilide and 3-butylidene-1(3H)-isobenzofuranone.2.Compared with the quality of angelica sinensis volatile oil extracted in the smallscale experiment,the loss of the volatile oil in the pilot-scale experiment with obvious emulsification was as high as 79%,and the reduction of the extraction rate would waste the resources of medicinal materials seriously.The analysis of pilotscale experiment volatile oil' physical and chemical properties showed that the color was deepened,the transparency of liquid was poor,the relative density and refractive index decreased,the optical rotation turned from right to left,and the acid value increased.The chemical components of type and percentage was changed,the content of trans-ligustilide decreased and 3-butylidene-1(3H)-isobenzofuranone increased.In summary,emulsification affected the appearance and internal quality of angelica volatile oil.3.During the extraction process of angelica volatile oil,81 groups of angelica volatile oil emulsions were collected.Temperature,turbidity,and chemical components were determined,respectively.PLS analysis showed that temperature was positively correlated with alkanes and lactones in the emulsion,and these components were positively correlated with turbidity.When the collector temperature was higher than50 ?,angelica volatile oil was easy to emulsify.Due to the high content of Q-marker in emulsion and the difficulty of oil-water separation,the loss of trans-ligustilide and3-butylidene-1(3H)-isobenzofuranone in angelica volatile oil was as high as 89% and91%,respectively.7 kinds of chemical components of positive correlation with turbidity,the HLB value were calculated.The HLB value of 2-methoxy-4-vinylphenol was 4.4,which was in the emulsification range of surfactant 3-6.At the same time,2-methoxy-4-vinylphenol was the chemical component with positive correlation with turbidity in emulsions from three producing areas.So,2-methoxy-4-vinylphenol was determined as the material basis of angelica volatile oil'emulsification.4.The condition of molecular simulation was set at 50 ? and 1 atmospheric pressure in the canonical ensemble.The effect of 2-methoxy-4-vinylphenol on angelica volatile oil emulsion was simplified as the effect on ligustilide/water system.The results of surface tension and Gibbs free energy showed that 2-methoxy-4-vinylphenol could reduce the surface tension with water at low concentration.Through the polar surface area and radius of gyration of solvent,2-methoxy-4-vinylphenol could increase the dispersing ability of ligustilide in water and reduce the surface tension between ligustilide and water.Hydrophobic aromatic ring of 2-methoxy-4-vinylphenol could stack with the ligustilide aromatic ring via Pi-Pi bond,and hydrophilic phenolic hydroxyl and methoxy group could form intermolecular hydrogen bond with water molecules to increase the solubility of ligustilide in water phase.Finally,a relatively stable emulsion was formed.5.Validation experiments showed that turbidity of angelica volatile oil/water solution increased with the increasement of temperature in a certain range,2-methoxy-4-vinylphenol could increase turbidity of angelica volatile oil/water solution and ligustilide/water solution,which confirmed that temperature and 2-methoxy-4-vinylphenol affected the degree of emulsification.In the exploration of demulsification scheme,the extraction rate of volatile oil could be improved by controlling the collector temperature below 50 ?.The new material porous lowdensity polystyrene and coalescence method could reduce the turbidity of volatile oil emulsion and realize oil-water separation preliminary.Conclusion1.Combined with the analysis of chemical components and biological activity,the predicted Q-marker trans-ligustilide and 3-butylidene-1(3H)-isobenzofuranone provided evaluation index for the quality research of Angelica volatile oil,and also provided reference for the Q-marker prediction of volatile oil of traditional Chinese medicine.2.According to the comparative analysis of extraction rate,physicochemical properties and chemical components,emulsification reduced the extraction rate and quality of Angelica volatile oil.This provides data support for the study of emulsification phenomenon of volatile oil of TCM,and lays an experimental foundation for the establishment of quality standard of volatile oil of TCM.3.The relationship among temperature,turbidity,chemical components and the percentage change of Q-marker in the emulsion were obtained by the analysis of emulsification rule.The reason of emulsification affecting the quality of Angelica volatile oil was explained.The selected 2-methoxy-4-vinylphenol as emulsifying material basis determined the research scope for exploring the emulsifying mechanism.4.Molecular dynamics simulation technology was used to analyze the surface tension of 2-methoxy-4-vinylphenol and the influence of 2-methoxy-4-vinylphenol on oil /water system,and the mechanism of emulsification was explained microscopically.5.The effects of temperature and 2-methoxy-4-vinylphenol on the emulsification degree of Angelica volatile oil / water system were confirmed.By controlling the collector temperature,applying new material porous low-density polystyrene and adopting coalescence method,the simple and feasible de-emulsification method was obtained,which provides a reference for oil-water separation of emulsion.