HPLC plays an important role in analytical research and quality control laboratory. Important decisions regarding the manufacturing process and publication of study results are made based on the high degree of reliability of the data generated by the HPLC system. Because HPLC is a multi-component system, the accuracy and reliability of analysis results depend on the performance level of each component. This article introduces some suggestions to improve the authenticity of the results.
Mobile phase
The main function of mobile phase is to transport the injected sample to the chromatographic column, and transfer each component to the detector for identification and quantification after separation in the chromatographic column. The flow should be freshly prepared before the start of analysis and HPLC, and the solvent should not contain any suspended particles. Use the solvent inlet filter to filter the solvent to avoid impurities entering the system. uHPLCs suggests further using uHPLCs inline filter or hplc guard column installed in front of and behind the chromatographic column, which can effectively prevent the column from being polluted by particles generated in the system.
The mobile phase solvents shall be completely miscible to each other, and the injected sample shall be miscible. When using a buffer as a mobile phase component, do not allow the buffer to dry out inside the system, as the salt formed during drying can deposit and prevent free flow of the mobile phase and damage to the pump component.
Sample
Large volume injection will lead to distortion of peak shape and overload of hplc column. The compatibility of the sample with the mobile phase should be checked prior to analysis. If solubility is low, try using a solvent or solvent combination that matches the polarity of the mobile phase mixture.
Chromatographic column is the key component of HPLC system. Its correct maintenance and use will greatly help to provide high reliability results. The operating flow rate and pressure should change gradually so that these changes do not interfere with the stationary phase packing. The use of uHPLCs pre guard column can also prevent impurities from entering the hplc column. Since the flow rate of the mobile phase depends on the temperature, it is strongly recommended to use the column temperature box to maintain a constant column temperature to maintain the consistency of flow rate.
Pump
In the case of gradient analysis, a pump is required to maintain the composition of the mobile phase according to the software program. Such conditions are achieved by preventing damage to pump parts and replacing such parts in a timely manner when any visible damage is found. Flush the pump gradually with medium polar solvent or pure water as appropriate.
Light source
Most HPLC analyses can be performed using UV light sources. This light source has a useful life. Replacing the lamp as recommended by the supplier will help to improve the reproducibility of the absorbance value.
Detector
The detector is the final component of the HPLC system, confirming the presence and quantifying of different sample components. uHPLCs recommends regularly cleaning the detector offline and rinsing with water and mobile phase before reuse. Do not exceed the operating pressure range to avoid possible damage to the detector unit wall.
The suggestions provided by the uHPLCs will help to improve the quality and accuracy of the results. It is also very important to regularly clean and calibrate the instrument and check whether the instrument is in normal use. The instrument is expensive and needs careful use. Some daily inspection and cleaning processes can help to prolong the service life of the instrument.
Post time: Apr-14-2022
