Understanding of Relationship Among Pumps, Columns, and Detectors
At present, many scientific and technological workers who develop and use HPLC have not figured out or have not fully figured out the relationship between the pump, column, and detector in HPLC. Many domestic and foreign manufacturers only produce pumps and detectors, and chromatographic columns are purchased parts. When we say HPLC is a system, we mean a complete system composed of pumps, columns, and detectors. Therefore, the HPLC system produced in this way is always not easy to use.
However, the majority of users, because the HPLC pump, column, and detector are not fully understood, cannot use HPLC to the highest level (to obtain the highest analysis and detection data with the smallest error). Based on HAWACH long-term practical experience in the development and use of HPLC, it’s believed that the following issues are particularly worthy of attention when developing and using HPLC:
1. Pumps:
Role: Pumps are responsible for delivering the mobile phase through the chromatographic system.
Connection to Columns: The pump is connected to the chromatography column, providing a continuous flow of the mobile phase.
Importance: Precise and consistent flow rates are essential for reproducible chromatographic separations. The pump ensures a constant mobile phase flow, allowing for accurate elution of analytes from the column.
2. Columns:
Role: Columns are the separation beds where compounds in the sample are separated based on their interaction with the stationary phase.
Connection to Pumps and Detectors: The column is connected to both the pump (for the delivery of the mobile phase) and the detector (for signal detection).
Importance: The column’s stationary phase is selective, causing different compounds to interact differently and elute at distinct times. The choice of column type (e.g., reversed-phase, normal phase) and dimensions (length, diameter) influences the separation efficiency and selectivity.
3. Detectors:
Role: Detectors monitor the eluent leaving the column and generate signals in response to the presence of analytes.
Connection to Columns: Detectors are connected downstream from the column to capture and analyze the separated compounds.
Importance: Detectors provide information on the identity and concentration of analytes. Common detectors include UV-Visible detectors, fluorescence detectors, mass spectrometers, and refractive index detectors. The choice of detector depends on the properties of the compounds being analyzed.
4. Data System (optional):
Role: A data system or chromatography software is often used to control instrument parameters, acquire and process data from the detector, and generate chromatograms.
Connection to Pumps, Columns, and Detectors: The data system interacts with the pump, column, and detector to ensure coordinated operation and accurate data acquisition.
Importance: Chromatography data systems facilitate automation, data analysis, and reporting, enhancing the efficiency and reliability of the chromatographic process.
Correctly understand and handle the relationship among HPLC pumps, columns, and detectors:
(1) HPLC is a system, and it can be called HPLC only by combining the three parts of pump, column, and detector. Only when the quality of the pump, column, and detector are all qualified, and the test results are also qualified after online, can this HPLC be qualified.
(2) All three components are important, and one cannot be emphasized. It should be said that the three are a whole, one is indispensable, and they are all important.
(3) The three components must be connected for inspection and testing, and the quality index requirements must be met before the HPLC can be said to meet the quality requirements. Otherwise, it can only be said that the components are qualified, and the whole system may not be qualified. Why? Because:
①The pump will have unstable pressure and unstable flow due to column problems, which will affect the quality of the HPLC system;
②The column has a great influence on the pump: column blockage, column contamination, column joint leakage, etc., will affect the pressure fluctuation of the pump, make the flow rate unstable, affect the system instability, make RSD worse, peak tailing, and reduce sensitivity ;
③The column has a great influence on the detector: reduced column efficiency (decrease in the number of column trays), column blockage, the degree of newness of the column, joints, pipeline leakage, etc., will affect the detection limit, resolution, noise, and sensitivity of the detector.
Please note: the chromatographic column is a quick-wear part. Users will often replace the chromatographic column with a new one after long-term use. At this time, the pump and detector of the HPLC system basically enter the bottom of the “electronic component failure theory” and are relatively stable.
Therefore, the problem that the instrument cannot be used due to the replacement of a new column will not occur. In particular, it should be pointed out that after the user changes a new bar, the comprehensive indicators of the system are generally not checked. If the indicators are tested, the original factory indicators may not be reached (because the quality of the pump and detector will decline after a long period of use, and the old pump, new column, and old detector cannot make the best-equipped equipment), but not affect the usage.
