几种中药醇沉过程颗粒生长特征与沉降模型研究
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摘要
醇沉工艺是目前我国中药制药企业最常用的分离精制方法,近年来关于中药醇沉工艺的研究报道主要集中在工艺参数优化和过程质量检测等方面,缺乏颗粒生长特征、微观形态和静置沉降模型等方面的研究。本论文以丹参、赤芍等中药浸膏为研究对象,对其醇沉过程颗粒生长特征、微观形态变化规律及静置沉降动力学等进行了系统的研究。本论文主要研究内容和结果如下：
1.采用FBRM和PVM技术研究了丹参醇沉过程颗粒生长团聚行为。结果表明浸膏初始密度和初醇浓度对颗粒生长特征和团聚行为均存在明显的影响。当初醇浓度为95%时,丹参醇沉过程总颗粒数量呈明显的双峰分布,颗粒生长过程可分为诱导期、生长期、聚结期、破碎分散期以及相对稳定期。浸膏初始密度和初醇浓度增大,诱导期缩短,颗粒平均生长速率和最大生长速率增大。当初醇浓度小于90%时,颗粒数量呈逐渐增大然后相对稳定的平台分布,表明不同条件下,丹参醇沉过程颗粒析出动力学存在差异。PVM和有效成分测定结果表明,有效成分的损失与颗粒形态存在相关性,颗粒团聚结块程度越高,混合液中液滴越多,有效成分包裹损失越大。颗粒电位分析与蛋白质和多糖测定结果表明,丹参醇沉过程形成的颗粒带负电荷且随着混合液乙醇浓度增大,电荷量增大,表明颗粒团聚不是由颗粒静电吸引力导致,可能与颗粒的微观形态和沉淀之间分子间吸附结合作用力有关,其具体机理还有待进一步研究。
2.以丹参、青蒿、益母草、金银花和枳壳为对象,采用FBRM-PVM联用技术,考察其醇沉过程颗粒数量、弦长分布和微观形态的变化规律,并对醇沉过程固液两相总蛋白和总糖的含量分布进行了初步研究。FBRM和PVM结果表明不同浸膏醇沉过程颗粒生长经历不同的阶段。总蛋白和总糖分析结果表明,不同浸膏醇沉过程固液两相总蛋白和总糖的含量分布存在明显差异。此外,不同浸膏醇沉过程颗粒数量和生长速率的动力学之间也存在差异,且与总糖和总蛋白在固液两相中的含量分布存在关联。其中丹参醇沉过程颗粒数量和生长速率均与液相中总糖浓度存在良好二次方抛物线关系。青蒿、益母草和金银花三种浸膏醇沉过程颗粒数量和生长速率均与固相中总糖浓度存在良好线性关系。枳壳浸膏醇沉过程颗粒数量和生长速率均与液相中总糖／总蛋白比例存在良好线性关系。以丹参为对象的颗粒数量与生长速率动力学验证结果表明所建立的模型预测性能良好,具有潜在应用价值。
3.以丹参二次醇沉混合液静置沉降过程为研究对象,考察关键醇沉工艺参数对其颗粒沉降过程的影响。结果表明丹参醇沉混合液沉降曲线与污泥沉降过程相似,主要包括区域沉降和压缩沉降。在Vesilind方程的基础上,建立了区域沉降阶段的速率模型,其表达式为：
Vzs=8.75×10-35e195.4p
同时考察了醇沉混合液密度对区域沉降过程的影响,结果表明随着混合液密度增大,区域沉降速率增大。当醇沉混合液密度ρ从0.916g·mL-1增大到0.934g·mL-1时,对应的区域沉降速率Vzs从0.262cm·min-1增大到3.23cm·min-1,增大幅度达11.3倍。
此外,建立了丹参二次醇沉混合液静置沉降过程压缩沉降阶段界面高度模型,并提出了模型参数的预测公式,同时导出了压缩沉降速率公式,提出了判断区域沉降与压缩沉降临界点的方法。
最终建立了丹参二次醇沉混合液静置沉降全过程的分段描述模型,其表达式为：
验证结果表明所建立的分段描述模型能准确地对丹参二次醇沉混合液静置沉降过程界面高度进行预测。该模型的建立对醇沉设备的设计和醇沉工艺运行控制具有一定的指导意义。
4.以赤芍一次醇沉和二次醇沉为对象,考察了不同因素对醇沉混合液沉降过程的影响。结果表明药液含醇量、浸膏初始密度和初醇浓度三个因素对赤芍醇沉混合液静置沉降过程具有明显影响。随着药液含醇量增加,沉降速度逐渐增大,沉淀沉降比逐渐减小,沉降性能得到改善。当浸膏密度为1.140g·mL-1时,沉降速度最快,最终形成的固体沉淀体积最小。初醇浓度为95%条件下颗粒沉降速率最大。研究结果表明,在更高的药液含醇量条件下,同样体积的醇沉混合液在沉降终点时沉淀的体积更小,减少了固液分离时沉淀的处理量,有利于提高固液分离效率。
在批沉降实验基础上,建立了赤芍醇沉混合液压缩沉降过程速率预测模型：Vs=(93.52-0.946PSV)×[1-(0.00153+0.0081PVI)×x]4.65
预测结果表明所建立的压缩沉降速率模型可同时准确描述赤芍水提浸膏一次醇沉和一次醇沉浸膏二次醇沉压缩沉降过程。此外,模型对生产规模醇沉样本的预测结果良好,具有较好的生产应用意义。
Alcohol precipitation is the most widely used technology in the production of TCM to achieve initial purification and separation of a crude extract. Unfortunately, most of the reported researches about alcohol precipitation of TCM mainly paid attention to parameters optimization and content analysis in the precipitation stage, while few researches focused on particle growth characteristics, morphology and settling model. Danshen (Salvia Miltiorrhiza Bunge) and Chishao (Radix Paeoniae Rubra) and other extracts were used to systematical study the particle growth characteristics, morphology and settling model of the alcohol precipitation process. The contents and results of the dissertation were described as follows:
1. The focused beam reflectance measurement (FBRM) and particle video microscope (PVM) technologies were introduced to study the particle growth and agglomeration processes during alcohol precipitation process of Danshen extract. The results showed that both initial extract density and initial alcohol concentration have significant effects on the particle growth and agglomeration. The variation of particle counts during the alcohol precipitation process of Danshen extract showed a significant bimodal distribution at initial alcohol concentration of95%. The particle growth process can be divided into five stages according to the variation of particle counts: induction, growth, agglomeration, breakage and relative stable stage. As the initial extract density and initial alcohol concentration increase, the induction time reduces, while the average and maximum particle growth rates increase. The particle counts increases gradually and finally reaches to almost a flat distribution at the initial alcohol concentration lower than90%. The result indicates different particle precipitation kinetics under different conditions. It can be inferred from the particle PVM images that there are some correlations between active component loss and particle morphology. Higher agglomeration leads to more droplets and greater active components loss. The results of zeta potential show that increase charge on the particles in agree with the increase ratio of polysaccharide/protein in the solid phase, indicating that electrostatic attraction is not the main reason for particle agglomeration during the alcohol precipitation process of Danshen. The mechanism of agglomeration is probably related to the particle morphology and the binding forces between particles. Further research is needed to gain a better understanding of the mechanism of agglomeration.
2. FBRM combined with PVM were applied for studying the variation of particle number, CLD and micron morphology characteristics in the alcohol precipitation process of Danshen (Salvia Miltiorrhiza Bunge), Qinghao (Artemisiae Annua), Yimucao (Leonurus japonicus), Jinyinhua (Flos Lonicerae), Zhiqiao (Fructus Aurantii). The distribution of total protein and polysaccharide in the solid and liquid phase during the alcohol precipitation process was also investigated. The results showed that the particle growth behavior is different for different extracts. The kinetics of particle counts and growth rate are also different for different alcohol precipitation processes. During the alcohol precipitation process of Danshen extract, the chord counts and growth rate of particles are found to be a quadratic parabola function of polysaccharide concentration in the liquid phase. The kinetics of chord counts and growth rates for Qinghao, Yimucao and Jinyinhua are very similar, which shows a negative and positive linear function of polysaccharide concentration in the precipitate phase, respectively. The kinetics of chord counts and growth rates for Zhiqiao are linear functions of polysaccharide/protein in the liquid phase. The validation results of the kinetics of particle counts and growth rate for the alcohol precipitation process of Danshen extract show that the developed models performance good prediction and have potential applications.
3. The second alcohol precipitation mixture of Danshen was used to investigate the effects of alcohol precipitation parameters on the settling process of mixture. The settling results indicated that the settling process of second alcohol precipitation mixture of Danshen can be divided into zone settling and compression settling, which is similar to the settling process of activated sludge. Based on the Vesilind function, a modified zone settling velocity model of second alcohol precipitation mixture of Danshen was developed and the expression is:Vzs=8.75×10-35e195.4p. The effects of alcohol precipitation mixture density on zone settling were investigated. The results showed that the zone settling velocity increases as the increasing of mixture density. The zone settling velocity increased from0.262to3.23cm·min-1as the mixture density increased from0.916to0.934g·mL-1, increased by as much as1130%. Meanwhile, a model describing the interface height was developed and the model parameters prediction formulas were provided. A velocity function for the compression settling was deduced from the model and a method for determining the critical time when the zone settling stage ends was established. Finally, a segmented model for describing the complete settling process was developed. The interface height equations for zone settling and compression settling stages are respectively given in the following:
The method was applied to predict the interface height during the complete settling process of second alcohol precipitation mixture, the results showed that the settling processes could be simulated well.
4. In order to study the settling characteristics for different alcohol precipitation process of TCM. The first and second alcohol precipitation of Chishao (Radix Paeoniae Rubra) extract were used to investigate the effects of alcohol concentration of mixture, the extract density, the initial alcohol concentration, the ratio of height to diameter (H/D) and the settling temperature on the settling process. The results showed that three factors, including mixture alcohol concentration, initial extract density and initial alcohol concentration, have significant influence on the settling process. The settling velocity increases as mixture alcohol concentration increases. Meanwhile, the Precipitate settling volume ratio (PSV) decreases, indicating settling performance improvements. Smallest precipitate volume and greatest settling velocity were obtained at initial extract density of1.140g-mL"1and initial alcohol concentration of95%, respectively. The results also showed that the higher the mixture alcohol concentration, the smaller the precipitate settling volume for same settling mixture volume. This will reduce the process load of precipitate and help to improve the efficiency of the solid-liquid separation. Therefore, settling performance of the alcohol precipitation mixture of TCM can be taken as an evaluation index in the optimization and industrial scale-up research. Under the condition of required retention rate of the active components, parameters which obtain better settling performance will accelerate the particle settling velocity and reduce the settling time, as well as the process load of precipitate.
Furthermore, a model describing the compression settling process of first and second alcohol precipitation processes of Chishao was developed based on batch tests. The equation is:
Vs=(93.52-0.946PSV0.5h) x [1-(0.00153+0.008IP Ⅵ)×x]4.65
The model was validated by experiments and the results showed that the compression settling processes could be simulated well by the developed model. In addition, the model shows good prediction for the production scale alcohol precipitation sample and potential industrial application.
1.采用FBRM和PVM技术研究了丹参醇沉过程颗粒生长团聚行为。结果表明浸膏初始密度和初醇浓度对颗粒生长特征和团聚行为均存在明显的影响。当初醇浓度为95%时,丹参醇沉过程总颗粒数量呈明显的双峰分布,颗粒生长过程可分为诱导期、生长期、聚结期、破碎分散期以及相对稳定期。浸膏初始密度和初醇浓度增大,诱导期缩短,颗粒平均生长速率和最大生长速率增大。当初醇浓度小于90%时,颗粒数量呈逐渐增大然后相对稳定的平台分布,表明不同条件下,丹参醇沉过程颗粒析出动力学存在差异。PVM和有效成分测定结果表明,有效成分的损失与颗粒形态存在相关性,颗粒团聚结块程度越高,混合液中液滴越多,有效成分包裹损失越大。颗粒电位分析与蛋白质和多糖测定结果表明,丹参醇沉过程形成的颗粒带负电荷且随着混合液乙醇浓度增大,电荷量增大,表明颗粒团聚不是由颗粒静电吸引力导致,可能与颗粒的微观形态和沉淀之间分子间吸附结合作用力有关,其具体机理还有待进一步研究。
2.以丹参、青蒿、益母草、金银花和枳壳为对象,采用FBRM-PVM联用技术,考察其醇沉过程颗粒数量、弦长分布和微观形态的变化规律,并对醇沉过程固液两相总蛋白和总糖的含量分布进行了初步研究。FBRM和PVM结果表明不同浸膏醇沉过程颗粒生长经历不同的阶段。总蛋白和总糖分析结果表明,不同浸膏醇沉过程固液两相总蛋白和总糖的含量分布存在明显差异。此外,不同浸膏醇沉过程颗粒数量和生长速率的动力学之间也存在差异,且与总糖和总蛋白在固液两相中的含量分布存在关联。其中丹参醇沉过程颗粒数量和生长速率均与液相中总糖浓度存在良好二次方抛物线关系。青蒿、益母草和金银花三种浸膏醇沉过程颗粒数量和生长速率均与固相中总糖浓度存在良好线性关系。枳壳浸膏醇沉过程颗粒数量和生长速率均与液相中总糖／总蛋白比例存在良好线性关系。以丹参为对象的颗粒数量与生长速率动力学验证结果表明所建立的模型预测性能良好,具有潜在应用价值。
3.以丹参二次醇沉混合液静置沉降过程为研究对象,考察关键醇沉工艺参数对其颗粒沉降过程的影响。结果表明丹参醇沉混合液沉降曲线与污泥沉降过程相似,主要包括区域沉降和压缩沉降。在Vesilind方程的基础上,建立了区域沉降阶段的速率模型,其表达式为：
Vzs=8.75×10-35e195.4p
同时考察了醇沉混合液密度对区域沉降过程的影响,结果表明随着混合液密度增大,区域沉降速率增大。当醇沉混合液密度ρ从0.916g·mL-1增大到0.934g·mL-1时,对应的区域沉降速率Vzs从0.262cm·min-1增大到3.23cm·min-1,增大幅度达11.3倍。
此外,建立了丹参二次醇沉混合液静置沉降过程压缩沉降阶段界面高度模型,并提出了模型参数的预测公式,同时导出了压缩沉降速率公式,提出了判断区域沉降与压缩沉降临界点的方法。
最终建立了丹参二次醇沉混合液静置沉降全过程的分段描述模型,其表达式为：
验证结果表明所建立的分段描述模型能准确地对丹参二次醇沉混合液静置沉降过程界面高度进行预测。该模型的建立对醇沉设备的设计和醇沉工艺运行控制具有一定的指导意义。
4.以赤芍一次醇沉和二次醇沉为对象,考察了不同因素对醇沉混合液沉降过程的影响。结果表明药液含醇量、浸膏初始密度和初醇浓度三个因素对赤芍醇沉混合液静置沉降过程具有明显影响。随着药液含醇量增加,沉降速度逐渐增大,沉淀沉降比逐渐减小,沉降性能得到改善。当浸膏密度为1.140g·mL-1时,沉降速度最快,最终形成的固体沉淀体积最小。初醇浓度为95%条件下颗粒沉降速率最大。研究结果表明,在更高的药液含醇量条件下,同样体积的醇沉混合液在沉降终点时沉淀的体积更小,减少了固液分离时沉淀的处理量,有利于提高固液分离效率。
在批沉降实验基础上,建立了赤芍醇沉混合液压缩沉降过程速率预测模型：Vs=(93.52-0.946PSV)×[1-(0.00153+0.0081PVI)×x]4.65
预测结果表明所建立的压缩沉降速率模型可同时准确描述赤芍水提浸膏一次醇沉和一次醇沉浸膏二次醇沉压缩沉降过程。此外,模型对生产规模醇沉样本的预测结果良好,具有较好的生产应用意义。
Alcohol precipitation is the most widely used technology in the production of TCM to achieve initial purification and separation of a crude extract. Unfortunately, most of the reported researches about alcohol precipitation of TCM mainly paid attention to parameters optimization and content analysis in the precipitation stage, while few researches focused on particle growth characteristics, morphology and settling model. Danshen (Salvia Miltiorrhiza Bunge) and Chishao (Radix Paeoniae Rubra) and other extracts were used to systematical study the particle growth characteristics, morphology and settling model of the alcohol precipitation process. The contents and results of the dissertation were described as follows:
1. The focused beam reflectance measurement (FBRM) and particle video microscope (PVM) technologies were introduced to study the particle growth and agglomeration processes during alcohol precipitation process of Danshen extract. The results showed that both initial extract density and initial alcohol concentration have significant effects on the particle growth and agglomeration. The variation of particle counts during the alcohol precipitation process of Danshen extract showed a significant bimodal distribution at initial alcohol concentration of95%. The particle growth process can be divided into five stages according to the variation of particle counts: induction, growth, agglomeration, breakage and relative stable stage. As the initial extract density and initial alcohol concentration increase, the induction time reduces, while the average and maximum particle growth rates increase. The particle counts increases gradually and finally reaches to almost a flat distribution at the initial alcohol concentration lower than90%. The result indicates different particle precipitation kinetics under different conditions. It can be inferred from the particle PVM images that there are some correlations between active component loss and particle morphology. Higher agglomeration leads to more droplets and greater active components loss. The results of zeta potential show that increase charge on the particles in agree with the increase ratio of polysaccharide/protein in the solid phase, indicating that electrostatic attraction is not the main reason for particle agglomeration during the alcohol precipitation process of Danshen. The mechanism of agglomeration is probably related to the particle morphology and the binding forces between particles. Further research is needed to gain a better understanding of the mechanism of agglomeration.
2. FBRM combined with PVM were applied for studying the variation of particle number, CLD and micron morphology characteristics in the alcohol precipitation process of Danshen (Salvia Miltiorrhiza Bunge), Qinghao (Artemisiae Annua), Yimucao (Leonurus japonicus), Jinyinhua (Flos Lonicerae), Zhiqiao (Fructus Aurantii). The distribution of total protein and polysaccharide in the solid and liquid phase during the alcohol precipitation process was also investigated. The results showed that the particle growth behavior is different for different extracts. The kinetics of particle counts and growth rate are also different for different alcohol precipitation processes. During the alcohol precipitation process of Danshen extract, the chord counts and growth rate of particles are found to be a quadratic parabola function of polysaccharide concentration in the liquid phase. The kinetics of chord counts and growth rates for Qinghao, Yimucao and Jinyinhua are very similar, which shows a negative and positive linear function of polysaccharide concentration in the precipitate phase, respectively. The kinetics of chord counts and growth rates for Zhiqiao are linear functions of polysaccharide/protein in the liquid phase. The validation results of the kinetics of particle counts and growth rate for the alcohol precipitation process of Danshen extract show that the developed models performance good prediction and have potential applications.
3. The second alcohol precipitation mixture of Danshen was used to investigate the effects of alcohol precipitation parameters on the settling process of mixture. The settling results indicated that the settling process of second alcohol precipitation mixture of Danshen can be divided into zone settling and compression settling, which is similar to the settling process of activated sludge. Based on the Vesilind function, a modified zone settling velocity model of second alcohol precipitation mixture of Danshen was developed and the expression is:Vzs=8.75×10-35e195.4p. The effects of alcohol precipitation mixture density on zone settling were investigated. The results showed that the zone settling velocity increases as the increasing of mixture density. The zone settling velocity increased from0.262to3.23cm·min-1as the mixture density increased from0.916to0.934g·mL-1, increased by as much as1130%. Meanwhile, a model describing the interface height was developed and the model parameters prediction formulas were provided. A velocity function for the compression settling was deduced from the model and a method for determining the critical time when the zone settling stage ends was established. Finally, a segmented model for describing the complete settling process was developed. The interface height equations for zone settling and compression settling stages are respectively given in the following:
The method was applied to predict the interface height during the complete settling process of second alcohol precipitation mixture, the results showed that the settling processes could be simulated well.
4. In order to study the settling characteristics for different alcohol precipitation process of TCM. The first and second alcohol precipitation of Chishao (Radix Paeoniae Rubra) extract were used to investigate the effects of alcohol concentration of mixture, the extract density, the initial alcohol concentration, the ratio of height to diameter (H/D) and the settling temperature on the settling process. The results showed that three factors, including mixture alcohol concentration, initial extract density and initial alcohol concentration, have significant influence on the settling process. The settling velocity increases as mixture alcohol concentration increases. Meanwhile, the Precipitate settling volume ratio (PSV) decreases, indicating settling performance improvements. Smallest precipitate volume and greatest settling velocity were obtained at initial extract density of1.140g-mL"1and initial alcohol concentration of95%, respectively. The results also showed that the higher the mixture alcohol concentration, the smaller the precipitate settling volume for same settling mixture volume. This will reduce the process load of precipitate and help to improve the efficiency of the solid-liquid separation. Therefore, settling performance of the alcohol precipitation mixture of TCM can be taken as an evaluation index in the optimization and industrial scale-up research. Under the condition of required retention rate of the active components, parameters which obtain better settling performance will accelerate the particle settling velocity and reduce the settling time, as well as the process load of precipitate.
Furthermore, a model describing the compression settling process of first and second alcohol precipitation processes of Chishao was developed based on batch tests. The equation is:
Vs=(93.52-0.946PSV0.5h) x [1-(0.00153+0.008IP Ⅵ)×x]4.65
The model was validated by experiments and the results showed that the compression settling processes could be simulated well by the developed model. In addition, the model shows good prediction for the production scale alcohol precipitation sample and potential industrial application.
引文
[1]曹光明.中药制药工程学[M].北京：化学工业出版社,2004：213.
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