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(high performance liquid chromatography, HPLC) is also called high pressure liquid chromatography, high speed liquid chromatography, high resolution liquid chromatography or the like or Efficient liquid chromatography spectrometry. Based on the classical liquid chromatography method, the gas chromatography theory was introduced in the late 1960s and rapidly developed. The difference between it and the classical liquid chromatography method is that the filler particles are small and uniform, and the small particles have high column efficiency, but cause high resistance. It is required to transport the mobile phase with high pressure, so it is also called high pressure liquid chromatography. It is also called high-speed liquid chromatography because of its fast analysis speed.
High performance liquid chromatography is currently the most widely used chromatographic method. The high performance liquid chromatography system consists of a mobile phase storage liquid bottle, an infusion pump, an injector, a column, a detector and a recorder. The overall composition is similar to gas chromatography. However, many adjustments have been made to the characteristics of the mobile phase as a liquid. HPLC infusion pump requires constant and stable infusion volume; the injection system requires convenient injection and switching; because the liquid mobile phase viscosity is much higher than gas, in order to reduce column pressure, the HPLC column is generally thicker and the length is much smaller. Gas chromatography column. HPLC applications are very broad and cover almost every area of ??quantitative and qualitative analysis.
When using high performance liquid chromatography, the liquid analyte is injected into the column and moved through the pressure in the stationary phase. Due to the different interactions between different substances and the stationary phase of the tested species, different substances are sequentially separated from the column and are different through the detector. The peak signal is finally analyzed by comparing these signals to determine the substance contained in the analyte. As an important analytical method, high performance liquid chromatography is widely used in chemical and biochemical analysis. High-performance liquid chromatography is not fundamentally different from classical liquid chromatography. It is characterized by high-pressure infusion pump, high-sensitivity detector and high-efficiency particulate stationary phase. It is suitable for analysis of high boiling point, non-volatile, large molecular weight and different. Polar organic compounds.
The system of high performance liquid chromatography consists of a reservoir, a pump, an injector, a column, a detector, a recorder, and the like. The mobile phase in the reservoir is driven into the system by a high pressure pump. The sample solution enters the mobile phase through the injector and is loaded into the column (stationary phase) by the mobile phase, since each component in the sample solution has two phases. Different partition coefficients, when doing relative motion in two phases, after repeated multiple adsorption-desorption distribution processes, the components have a large difference in the moving speed, and are separated into individual components and sequentially flow out from the column. When passing through the detector, the sample concentration is converted into an electrical signal and transmitted to the recorder, and the data is printed in the form of a map.
Chinese name: High performance liquid chromatography
Type: liquid-solid adsorption chromatography, bonded phase chromatography, etc.
Foreign name: high performance liquid chromatography
Development time: 1960s
Composition: reservoir, pump, injector, column, etc.
Features: high pressure, high speed, high efficiency
What is the principle of high performance liquid chromatography:
The partition coefficient is related to the thermodynamic properties of the components, mobile phase and stationary phase, and also to temperature and pressure. Among the different chromatographic separation mechanisms, K has different concepts: adsorption chromatography is the adsorption coefficient, ion exchange chromatography is the selectivity coefficient (or exchange coefficient), and gel chromatography is the permeation parameter. However, the general case can be expressed by a distribution coefficient.
When the conditions (mobile phase, stationary phase, temperature, pressure, etc.) are certain, when the sample concentration is very low (Cs, Cm is small), K depends only on the nature of the component, regardless of the concentration. This is only the chromatographic condition under ideal conditions. Under these conditions, the obtained chromatographic peak is a normal peak; in many cases, as the concentration increases, K decreases, and the chromatographic peak is a tailing peak; As the solute concentration increases, K also increases, at which point the chromatographic peak is the pre-extension peak. Therefore, normal peaks can only be obtained if the injection volume is reduced as much as possible so that the concentration of the components in the column is lowered and K is constant.
Under the same chromatographic conditions, the component with a large K value in the sample has a long residence time in the stationary phase and then flows out of the column; the component with a small K value has a short residence time and flows out of the column first. The greater the difference in the partition coefficients of the components in the mixture, the easier it is to separate, so the partition coefficient of each component in the mixture is a prerequisite for chromatographic separation.
In HPLC, after the stationary phase is determined, K is primarily affected by the nature of the mobile phase. In practice, the composition ratio and pH value of the mobile phase are mainly adjusted to obtain the difference of partition coefficient between components and the appropriate retention time to achieve the purpose of separation.
This is the end of the introduction of Introduction of HPLC. I hope it can help you.