Usage and Precautions of HPLC Column

Before using an HPLC column, the HPLC column is best to perform column performance tests and save the results as a reference for future evaluation of column performance changes. However, HPLC column should be noted that the column performance may be different due to the differences in the samples, liquid chromatography column, and column temperature; in addition, the column performance test is performed according to the conditions in the liquid chromatography column factory report ( the conditions used in the factory test are the best conditions), only in this way, the measured results are comparable.

High Purity HPLC Columns


  1. Column Installation:
    • Follow the manufacturer’s guidelines for column installation, ensuring proper connections and sealing.
    • Use appropriate fittings and tools to avoid damage during installation.
  2. Mobile Phase Preparation:
    • Prepare the mobile phase according to the method requirements.
    • Ensure that the mobile phase is filtered to remove particulate matter that could affect column performance.
  3. Column Equilibration:
    • Equilibrate the column with the mobile phase before sample injection.
    • Follow recommended equilibration times to ensure stable and reproducible results.
  4. Sample Preparation:
    • Ensure that samples are properly prepared and filtered to prevent contamination.
    • Choose an appropriate injection volume to avoid overloading the column.
  5. Flow Rate:
    • Operate the HPLC system at the recommended flow rate for the specific column.
    • Avoid exceeding the maximum pressure limit specified by the manufacturer.
  6. Temperature Control:
    • Maintain the column at a consistent temperature using a column heater or temperature-controlled environment.
    • Follow temperature recommendations to ensure reproducibility.
  7. Column Storage:
    • Store columns in a cool and dry environment when not in use.
    • Follow the manufacturer’s guidelines for long-term storage.
  8. Column Selection:
    • Select the appropriate column type and dimensions based on the separation requirements.
    • Choose the stationary phase and particle size suitable for the analytes of interest.


  1. Pressure Limits:
    • Avoid exceeding the maximum pressure limits specified by the manufacturer to prevent column damage.
    • Monitor system pressure regularly.
  2. pH Compatibility:
    • Consider the pH stability of the column material and choose a compatible mobile phase.
    • Avoid extreme pH conditions if not recommended by the manufacturer.
  3. Injection Volume:
    • Do not exceed the recommended sample injection volume to prevent overloading the column.
    • Use appropriate dilution if needed.
  4. Avoid Contamination:
    • Minimize sample contamination by using clean glassware and proper sample handling techniques.
    • Flush the system thoroughly between runs to avoid carryover.
  5. Solvent Compatibility:
    • Ensure that the solvents used are compatible with the column material.
    • Avoid using aggressive solvents that may damage the column.
  6. Temperature Control:
    • Do not exceed the recommended temperature limits to prevent damage to the column.
    • Avoid sudden temperature changes.
  7. Backpressure Monitoring:
    • Regularly monitor backpressure during HPLC runs.
    • A sudden increase in backpressure may indicate column blockage or deterioration.
  8. Column Cleaning:
    • Follow recommended cleaning procedures provided by the manufacturer.
    • Use compatible solvents for cleaning.
  9. Avoid Impact:
    • Handle columns with care to avoid physical damage.
    • Protect columns during transportation and storage.
  10. Regular Maintenance:
    • Perform routine maintenance checks on the HPLC system, including column inspection.
    • Replace columns as recommended by the manufacturer or based on performance criteria.

Following these guidelines and precautions helps ensure optimal performance, longevity, and reproducibility in HPLC column usage. Always refer to the specific instructions provided by the column manufacturer for the best results.

Notes on the use of liquid chromatography columns

a. Because methanol is cheap, the methanol system is recommended for reversed-phase columns (except where acetonitrile must be used).
b. For normal phase columns, it is recommended to use petroleum ether with a boiling range of 30-60 °C or purified hexane as the liquid chromatography column. Unpurified hexane should not be used. It is better to use ultrapure water (resistivity greater than 18 megohms). Deionized water and double distilled water contain phenolic impurities, which may affect the analysis results.
c. The aqueous HPLC column should be prepared well before the experiment, especially if the buffer solution is used as the liquid chromatography column in summer. Add good sodium azide to prevent bacterial growth.
d. The liquid chromatography column requires a 0.45μm filter membrane to remove particulate impurities.
e. Use HPLC grade solvent to prepare the HPLC column, use a suitable liquid chromatography column can prolong the service life of the liquid chromatography column, and improve column performance.
You also can choose C18 Universal HPLC Columns, Phenyl-Ether HPLC Columns, C8 Standard HPLC Columns, and other HPLC Columns using.