工业制备色谱在中药分离制备中的应用

概述了工业制备色谱柱型结构、填料、色谱技术特性的研究进展,对目前普遍使用的工业制备色谱的工作原理作了较为全面的评述,总结了动态轴向压缩色谱和模拟移动床色谱技术在中药分离制备中的应用和发展。     

一种多通道匀浆装填毛细管色谱柱的新装置及应用

针对目前毛细管色谱柱装柱效率低、不同批次装填的毛细管色谱柱之间性能差异大的问题,我们发展了一种多通道匀浆装填毛细管色谱柱的新装置。该装置以液相色谱泵提供压力、采用磁力搅拌保持匀浆液均匀分散,一次可装填多达6根毛细管色谱柱。以牛血清白蛋白(BSA)的胰蛋白酶酶切肽段混合物为样本,选择峰容量、蛋白覆盖率、3个特定离子的保留时间以及毛细管色谱柱柱压为指标,在毛细管液相色谱-质谱联用系统上对装填的反相毛细管色谱柱的性能进行了评价。分别考察了一次装填的6根毛细管色谱柱、两次装填的12根毛细管色谱柱以及一次装填1根与一次装填6根毛细管色谱柱的性能及稳定性。实验结果表明:同一批次装填的6根毛细管色谱柱的性能相近;不同批次装填的12根毛细管色谱柱的峰容量和覆盖率没有明显的区别,但保留时间和毛细管色谱柱柱压的稳定性较差;一次装填1根和一次装填6根毛细管色谱柱柱性能的稳定性与两次分别装填6根毛细管色谱柱的稳定性相近,即采用本装置可显著提高毛细管色谱柱的装填效率且每次装填毛细管色谱柱的数量不会对柱性能产生影响。     

柱吸附容量对蔗糖还原糖色谱分离过程产率和回收率的影响

本文采用了能反映流体轴向扩散的大型液相色谱分离过程数学模型,应用计算机模拟分析蔗糖和还原糖的色谱分离过程,从吸附剂吸附容量和柱装填密度两个方面;考察柱吸附容量对大型色谱分离的产率和回收率的影响。研究结果表明：在色谱柱中流体线速度恒定的条件下对多组分分离,回收率是随吸附剂吸附容量以及床层装填密度的增大而增大的;产率先随吸附剂吸附容量的增大而增大,而在出现峰值后下降;随着床层装填密度的增大,产率增加,但与此同时轴向扩散系数也增大,从而降低分离效率,导致在较高装填密度的范围内产率增力。的幅度减少。在相同的吸附剂用量下,采用短柱高装填密度的色谱柱将比长柱低装填密度色谱柱能获得更高的回收率和产率。     

半制备色谱柱装填条件的选择

填充技术是影响柱性能的重要因素,但关于半制备柱装填的报道很少.本论文主要对干法和匀浆法装填的半制备柱(φ10×200mm)的性能进行了对比,并对两种方法的装填条件进行了选择.实验结果表明匀浆法装填柱的重现性和柱性能明显优于干法装填的柱.在匀浆法填充中,乙醇为匀浆溶剂,匀浆体积为120mL,冲洗流速为330mL/min,装填时间为20min时所填柱的性能最好.     

一种简单的毛细管色谱柱电加热装置的制作方法及其在液相色谱-质谱联用系统中的应用

为了实现蛋白质组的深度覆盖,特别是低丰度蛋白质的定性鉴定和定量分析,目前常用的方法是采用更长或装填更小粒径填料的毛细管色谱柱,但因此带来的问题是色谱柱反向柱压显著升高。针对以上问题发展了一种简单的毛细管色谱柱电加热装置制作方法,并将该装置安装于液相色谱-质谱联用系统,分别以牛血清白蛋白(BSA)酶切肽段混合物和酵母蛋白(yeast)酶切肽段混合物为样品,从柱压和柱效两方面对该装置的性能进行了评价。实验结果表明,所制作的毛细管柱电加热装置安装在装填粒径为3 μm反相色谱填料的毛细管柱上,在最佳电流(100 mA)下对BSA及yeast酶切肽段混合物进行分离时的柱压比不加电流时的柱压降低至少50%,柱效略有升高。这说明所制作的毛细管色谱柱电加热装置能显著降低柱压,为在较低的柱压条件下选择更小粒径色谱颗粒填料的毛细管色谱柱提供了一种有效的方法。     

自动真空液相色谱装置的研制及其在五味子成分分离中的应用

为满足天然产物高效分离的需求,本文研制了自动真空液相色谱(AUTO-VLC)装置并用于五味子石油醚萃取物的分离。该装置由自主设计的流动相储备系统、10通分流切换阀、3通切换阀、3个不同规格的动态轴向压缩色谱柱、10通馏分收集阀和馏分收集器组成,采用可编程逻辑控制器(PLC)S7-200实现了分离过程的不同比例流动相切换、不同规格色谱柱选择、分离时间设定及馏分收集的自动控制及监测。应用于五味子成分分离的结果表明:采用150 mm直径动态轴向压缩色谱柱的VLC,从100 g五味子石油醚萃取物中一次分离得到6个组成明显不同的样品(S1~S6)。建立了通过多次薄层色谱(TLC)展开筛选VLC分离条件的方法,并进行了分离验证。VLC分离条件选择方法:自动真空液相色谱的初始洗脱剂的比例以首次TLC展开时全部目标化合物的比移值(R_f)介于0~0.45时展开剂的组成为宜;梯度洗脱比例变化以k值(展开次数(n)与R_f的线性函数斜率)和TLC分离度为依据选择;不同R_f范围内洗脱次数由n≈ ΔR_f/k计算。采用选择的条件从样品S5中分离得到了13个组分和4个纯度在85%以上的化合物,仅耗时17 h。AUTO-VLC的研制对于中药成分的自动和系统性分离具有重要价值。     

自然沉降法制备常压色谱柱

提出了自然沉降法制备常压色谱柱,它与传统方法制备的色谱柱相比,柱效和分离效果均有明显提高。此法装填的色谱柱还具有良好的稳定性和重复性。     

离子交换水处理技术文摘

标题的装置包括装有预涂中空纤维的过滤器的过滤一脱盐箱，一个装填离子交换树脂的柱塔，用于监测徨水电导率或CL~-浓度的装置和依靠来自监测器的反馈信号来控制离子交换树脂数值的装置。阴离子交换树脂对阳离子交换树脂的数值的装置。阴离子交换树脂对阳离子交换树脂的混合比例控制在>0.5:1,离子交换树脂的平     

用于变性蛋白质复性及同时纯化的简易型色谱柱的研制及其应用

研制出一种简易型色谱柱并通过装填大颗粒的疏水色谱填料对色谱柱的性能进行了考察,该柱的外形和操作如同传统的液相色谱柱,但它却在生物大分子分离方面有着和高效液相色谱相似的分辨率。另外,只要给该柱装填合适的固定相,比如疏水相互作用色谱固定相,便可用于蛋白质的复性及同时纯化。实验考察了该色谱柱的结构、操作和性能,包括柱压、柱寿命及对蛋白质的分辨率等。以溶菌酶为模型蛋白质,实验测得其在初始浓度为50.0 g/L时的质量回收率和活性回收率分别为(96.6±1.3)%和(101.1±6.0)%。这种简易型色谱柱价格低廉     

时间域内拟合法测定固定床流体轴向扩散系数

使用惰性电解质溶液作为示踪剂,在001×7Ca~(2+)型树脂固定床上进行扰动应答实验,提出用时间域内拟合法测定固定床液相流体轴向扩散系数D_(ax)。结果表明:轴向扩散系数D_(ax)随流体线速增大而增大,干法装填的固定床层比湿法装填的固定床层有较大的轴向返混系数。采用Lax-Wendroff差分格式对固定床对流扩散方程进行数值解,理论流出曲线与实验流出曲线相吻合。     

Behavior of packing materials in axially compressed chromatographic columns

The behavior of a packing material (Luna C18 from Phenomenex, Torrance, CA, USA) was studied during the consolidation of a column bed under axial compression stress. The kinetics of this consolidation, the permeability and efficiency of the columns obtained, and the reproducibility of these column properties were measured under different conditions. The consolidation process and the column properties are considerably affected by the friction between the packing material in the bed and the column wall. Clear evidence of this wall effect was demonstrated. The apparent permeability of columns consolidated under the same axial stress increases with increasing column length. The apparent modulus of elasticity of the beds increases with increasing column length. The shear resistance between the packed bed and the column wall was measured for columns of different lengths. It increases rapidly with increasing bed length. The column efficiency for thiourea (unretained) and phenyloctane (retention factor, k' approximately 1) was much poorer after recompression than after the first compression. It depended little on the compression stress. The effect of the column length was small.     

Use of ultrasound to monitor the packing of large-scale columns,the monitoring of media compression and the passage of molecules,such as monoclonal antibodies,through the column bed during chromatography

The novel use of ultrasound as a detector in pilot- and production-scale chromatography is described. The difficulties in packing production scale chromatography columns using an integral packing valve are reviewed. Results are presented from the packing of 400- and 600-mm diameter columns with various medias. From these results it is proposed that when packing large columns using a packing valve, for a given medium and column size, there is an ideal rate and pattern (or control "corridor") by which the bed builds in order to give optimum performance. Ultrasound was shown to be able to monitor the building of such a column bed as the medium was pumped into the column. It was found that the ultrasound detector was sensitive to bed compression, mobile phase composition and components such as acetone, albumin, casein and monoclonal antibodies while on the chromatography bed. This enabled the visualisation, by ultrasound, of these components as they were chromatographed through the column.     

高效制备液相色谱柱技术的研究进展

本文概述了高效制备液相色谱柱的柱型结构、填料以及柱填充方法等研究的最新进展,讨论了制备柱与分析柱的不同特征,对目前普遍使用的压缩型制备柱的类型、结构及填充方法作了较为全面的评述,总结比较了工业化制备色谱填料不同于分析色谱填料的特点,探讨了高效制备液相色谱柱技术的应用和发展前景.     

The reverse auto-compression system: a new type of column for preparative liquid chromatography

Axial compression of the stationary phase is an easy technique for packing preparative liquid chromatography columns. However, the compression piston often needs an important external device to move the external jack. A new type of column is described which uses an auto-compression system to move the piston and compress the packing material. The system is fitted with a new annular injector which is known to reduce the peak tailing. The results of the tests demonstrate the ease of use of this column, its efficiency and stability.     

Visualization of bed compression in an axial compression liquid chromatography column

The consolidation of a packed bed undergoing axial compression was studied in glass columns using an on-column visualization process. In this visualization process the refractive indices of the mobile phase (carbon tetrachloride) and the stationary phase (YMC C18 silica) matched perfectly, hence the otherwise opaque stationary phase became transparent to the eye. Alumina layers, which have a different refractive index, were placed at regular intervals along the column bed. These layers were therefore visible and their movement could be tracked during the axial compression of the bed. Consequently, the Young's modulus could be measured at three radial locations and at four bed depths below the head fitting. Theresults showed that the bed was heterogeneous in packing density, both radially (with the bed density increasing from the column center toward the wall) and axially (with the density increasing from the column top toward its center). Furhermore, the bed was shown to be non-symmetrical about the column axis. This was thought to be due to the column inlet head fitting making contact with the packing material on one side of the column first, rather than making contact with the entire cross section of the packing simultaneously.     

Conformational changes of poly(N-isopropylacrylamide) chains at air/water interface: effects of temperature, compression rate, and packing density

This study concerns the effects of temperature, compression rate, and packing density on conformational properties of polystyrene-block-poly(N-isopropylacryamide) (PS-b-PNIPAM) diblock copolymer monolayers at the air/water interface using the Langmuir balance technique. The dependency of surface pressure changes on temperature and compression rate is strongly influenced by the conformations of PNIPAM chains at the interface, which can be adjusted by varying the packing density. Specifically, when loops or tails are formed at the interface, PNIPAM chains display thermosensitive properties due to hydration/dehydration and obvious dependence on compression rate. When PNIPAM chains take train conformation at the air/water interface, however, the surface pressure changes are nearly independent of temperature and compression rate because almost all segments of the PNIPAM chains are adsorbed at the interface and the nonpolar isopropyl groups are preferentially oriented toward the air. Our work reveals that one could manipulate stimuli-responsive properties of PNIPAM chains at the interface simply by adjusting the conformations of PNIPAM chains.     

Demonstration of the Feasibility of Radially Compressed Microbore HPLC Columns

Abstract

This work involved the development of radially compressed, microbore high performance liquid chromatography (HPLC) columns. The design of the overall system and the column are described, and the problems associated with the design features are reported. Variables examined during the course of this work included the column material, column length, packing method, flow rate, radial compression pressure, and internal column pressure. Efficiencies (expressed as plates/meter) are shown for various combinations of those variables and are compared to those obtained using a commercial, steel microbore column.     

Hydrodynamic impact of particle shape in slurry packed liquid chromatography columns

We report on a series of flow velocity and efficiency profiles, which were measured across the cross section of preparative chromatographic columns packed with different stationary phase materials using computed tomography. It is shown that this non-invasive technique is very useful for visualization of the inner part of a packed column and measurement of the spatial resolved column packing properties. For evaluation of the influence of the particle shape on the velocity distribution and column performance, irregular and spherical reversed phases were studied in detail. The results showed a decreasing velocity towards the column wall most certainly due to a lower permeability. This effect was much less pronounced in the case of spherical particles, indicating a more homogenous packing structure. The influence of the column packing pressure, as a possible measure for improvement of the packing homogeneity was also studied. It was shown that under the same packing conditions spherical particles always lead to a more homogeneous packing. The overall results of this work contribute to the origin of the fact that spherical material is superior to irregular one from the hydrodynamic point of view.