Industrial preparative chromatography / dynamic axial compression system in the pharmaceutical industry
1 Introduction With the rapid development of pharmaceutical, biochemical and other industries, preparative liquid chromatography separation technology has been more and more widely developed and applied, and has become an important method for separation and purification of complex mixtures, especially for natural products and large organisms. Separation of molecules (polypeptides, proteins, etc.). In order for the sample to be effectively separated, the column must be packed to meet the requirements for continuity, uniformity, stability and tightness of the bed. Column packing, especially for large-diameter columns of industrial production scale, presents considerable difficulties.
The traditional filling method is divided into a dry method and a homogenizing method. The dry filling method is awkward, the column performance is not good, and the filler particles are easily dispersed in the column wall region during the filling process. The packing density in the packed column of the homogenization method is not uniform, the packing density near the column wall is high, and the reproducibility of the column performance is poor. Filling large diameter columns in the traditional way, the bed is unstable and will reduce separation performance. At present, the flanged end-ends sold in the domestic market are basically filled by homogenization.
Dynamic Axial Compression (DAC) technology is recognized as the best technology for loading and preparing columns (column diameter greater than 50 mm). The principle is to use piston packing (homogenization) and during operation. Keep the bed under compression to ensure its stability. The column packed by the DAC method has uniform column, stable performance, high density, high column efficiency and good column performance reproducibility. Columns loaded with the DAC process are gradually dominated the entire preparative column market.
2 Advantages of Dynamic Axial Compression Columns Up to today, there are three main compression methods:
(1). Axial compression: is a wet filling process that uses a piston to compress the bed;
(2). Radial compression: It is a dry filling process. It uses a double-tube column. The inner tube is made of plastic or rubber. It is filled with dry filler particles and kept in a certain space formed by the two tubes. Pressure, the column is radially compressed;
(3). Circumferential compression: achieved by moving a conical rod along the axial direction of the column.
At present, among the three methods, axial compression and radial compression methods have been used, and the latter has not been studied in depth, and the filling process is complicated, and the filling principle has not been well understood and understood. , less application. The dynamic axial compression (DAC) method has many advantages and has been further researched and developed. Compared to traditional homogenization methods and other filling methods, DAC packed columns have many advantages:
First of all, the DAC column has high efficiency, good reproducibility, and short loading time. It can use smaller particle size fillers, reduce column length, increase column diameter, and thus reduce tube wall effect, which can be obtained almost close to the analytical column. The efficiency of the column can be made more efficient. Compared with the traditional flanged end-capping column, the yield and purity of the product can be greatly improved. The filling process is easy to learn and can be learned quickly even by people who have not been in contact with it. Once the column is contaminated, it can be easily and conveniently unloaded, cleaned, and refilled.
Secondly, in other filling methods, the inlet end of the column is loose, and the packed bed is unstable. After a period of use, it is easy to collapse at the inlet end to form large and small holes, and the liquid will be strongly in the cavity. The local convection leads to a large loss of efficiency. In the DAC method, the piston is applied to the bed at a constant pressure during the filling process, and the stud and the piston end are in close contact with the bed, and no looseness is formed at both ends of the column to form a cavity. During the use of the column, there is still a constant pressure acting on the piston, which acts on the entire bed. Even after a long period of use, the top of the column does not generate holes due to collapse, so the DAC column does not form holes. When the fluid is disturbed, the column performance can be kept stable for a long time. Moreover, even if voids or cracks are formed in the column during use, the initial efficiency can be obtained after the voids or cracks formed in the column are recovered during the unused period.
Finally, although the DAC column is larger than the one-time input of a conventional flanged end-capping column, the DAC column greatly increases the yield and purity of the product, prolongs the life of the column, and can be refilled by itself. From the perspective of overall cost effects, costs are lower. Therefore, the DAC column can increase production efficiency and save production costs. : Simultaneously purchase a conventional preparation column with a diameter of 5cm and a set of column machine. If it is used frequently, due to the limitation of the service life of the conventional preparation column, it needs to be purchased frequently and the equipment is deactivated due to the problem of delivery period. After the month, the cost of the two is roughly equivalent. When it is 2 years, the use of the column packing machine can save about 30,000 euros compared to the use of the conventional preparation column.
3 Influencing factors of the dynamic axial compression column To ensure the performance of the dynamic axial compression column, many factors must be considered.
3.1 Effect of column length:
Under the same conditions, as the column grows, the axial pressure distribution within the column is larger. The longer the column, the greater the pressure difference between the piston and the other end, the more unstable and uneven the resulting bed, the smaller the packing density, resulting in an axial pressure gradient and density gradient.
3.2 Effects of filler properties:
For spherical particles, the filler can reach a steady state quickly and smoothly; for irregularly shaped particles, the collapse of the bed during the filling process is unpredictable (especially when the filler particles are large), and the filling situation is complicated. The influence of irregular packing pressure is large. When the pressure is large, some particles will be broken, which will cause the column length to decrease and the column pressure drop to become larger.
3.3 Effect of pulping solvent:
The nature of the slurry is also a very important factor, which is evaluated by the physicochemical properties, surface properties and geometry of the filler particles. If the solvent is not properly selected, the formed homogenate is unstable, the deposition of the particles is fast, and the friction of the particles is large, so that the packing density formed at the column wall is more uneven.
3.4 Effect of compression pressure:
The performance of the column depends on the strength of the packing pressure and the length of the compression time. The behavior of a dynamic axial compression bed is very complicated. During the compression of the column, although the pressure applied to the piston is equal, within the bed, the pressure transfer is not as uniform as the pressure in the liquid, and the particles contacting the piston are subjected to direct pressure, and the bed The other end is only squeezed by the filter plate and the flange. The pressure of the piston end bed is greater than the pressure of the fixed end. Because there is friction between the column wall and the particles, even along the axis of the column. In the same section, the distribution of pressures is also different, that is, there are pressure gradients in both the axial and radial directions. For the axial compression of irregular packing, the higher the pressure, the higher the degree of particle breakage, so the performance of the column obtained under low pressure is better than that under high pressure. Regardless of the shape of the particle, the pressure increases rapidly to the maximum at the beginning of compression. The performance of the column obtained is gradually higher than that of the gradual increase.
4 Axial compression system Dynamic axial compression column must use the most precise polishing technology and concentric technology to make the column wall finish (≥▽13, ie surface roughness Ra≤0.012μm) exceeds the foreign similar products and guarantees The concentricity of the column tube (0.2μm) minimizes the wall effect during packing and use, and ensures good sealing performance and efficiency when the initial filling is repeated after repeated filling. There are indeed a number of key techniques in the fabrication of the preparation column. It is worth mentioning that the preparation of the column at the inlet end of the sample requires the addition of a distribution plate. Porous linear groove distribution plate, when the sample flows from the narrow pipe into the front end of the column, the plate can accelerate the sample to be evenly distributed to the cylinder plane. This design is effective for reducing the band expansion effect. The key to success.
5 Conclusion One of the most painful and time consuming steps in the separation and purification process is that the separation of analytical scales cannot be directly linearly amplified. In this case, the preparation method needs to be re-developed to achieve or approach the resolution required by the analytical method, thus delaying the important steps from drug discovery to optimization in the development of new drugs. Separation conditions cannot be directly amplified. Usually, the type of filler filled in the preparation column and the analytical column is different (or the preparation column is used at a lower cost than the analytical column, and the direct amplification is only possible in the preparation column and the analysis column. Implemented when they are identical.
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