What are the common plant extract manufacturing detection methods in the plant extraction industry? Today we will bring you HPLC detection methods. Let’s see what is HPLC? What is the HPLC process?
What is HPLC?
HPLC stands for High Performance Liquid Chromatography. It also known as “high pressure liquid chromatography,” “high speed liquid chromatography,” “high resolution liquid chromatography,” “modern column chromatography” และอื่นๆ.
High performance liquid chromatography (HPLC) is an analytical method in the late 1960s. In recent years, it widely uses in the separation and determination of functional components of health food, nutritional fortifiers, vitamins and proteins. About 80% of organic compounds in the world can be analyzed by HPLC.
Characteristics of HPLC
High performance liquid chromatography (HPLC) has the characteristics of high pressure, high efficiency, high speed, high sensitivity and high selectivity. It widely uses in pharmaceutical, environmental protection, petrochemical, food, chemical and other industries. And is now an important method for separation and analysis.
Flow Chart for High Performance Liquid Chromatography
The solvent in the liquid storage bottle is sucked into the chromatography system by the pump. And then output, and after the flow and pressure measurement, it imports into the sampler. The detected object injects into the sample injector and passes through the column with the mobile phase. After separation on the column, the detected signal collects and processes by the data processing equipment, and the chromatogram records. Waste liquid flows into the waste liquid bottle. The gradient controller can also use for gradient elution in the case of complex mixture separation. This is similar to the programmed temperature of gas chromatography, except that gas chromatography changes the temperature. While HPLC changes the polarity of the mobile phase, so that the sample components can separate under the best conditions.
The separation process of high performance liquid chromatography
Like other chromatographic processes, HPLC is a continuous multi-exchange process between the stationary phase and the mobile phase. It separates different solutes by the difference of partition coefficient, affinity, adsorption force or molecular size between two phases.
Start the sample by adding it to the stigma. Assume that the sample contains 3 components, ก, B and C, which enter the column along with the mobile phase and begin to be divided between the stationary phase and the mobile phase. The component A with small partition coefficient is not easily blocked by the stationary phase and flows out of the column earlier. The component C with a large partition coefficient stayed on the stationary phase for a long time and left the column later. The partition coefficient of component B is between A and C, and the second flows out of the column. If a mixture containing multiple components enters the system, each component in the mixture flows out of the column. According to its partition coefficient between the two phases in different order to achieve the purpose of separation.
The separation of different components in the chromatographic process depends first on whether there are differences in the partition coefficient, adsorption capacity and affinity of each component between the two phases, which is a thermodynamic equilibrium problem and the primary condition for separation. Secondly, when the different components move in the column, the spectral band is broadened with the column length, and the separation is related to the diffusion coefficient between the two phases, the size of the stationary phase, the packing of the column and the flow rate of the mobile phase. Therefore, the final effect of separation is the comprehensive benefit of both thermodynamics and kinetics.