How do I troubleshoot HPLC?
- Check system for loose fittings. Check pump for leaks, salt buildup, unusual noises.
- Check make-up of mobile phase.
- Purge air from pump head or check valves.
- Use reliable column oven.
- Inject smaller volume (e.g., 10 μL vs.
- Adjust solvent.
- Substitute new column of same type to confirm column as cause.
How do I clear a blocked HPLC column?
First, wash the column with varying solvents at a low flow rate. However, if your column is so clogged that it can’t flow at all, flip the column and run it in the opposite direction. If all else fails, then you might need a new column.
How do I start an HPLC instrument?
For setting up the HPLC machine:
- Make sure you have all your buffers set up.
- Open the purge valve and purge the system for 5 minutes.
- Add your samples into the autosampler tray.
- Stop the purge.
- Close the purge valve.
- Run the system at a normal flow rate (1 ml/min) with your buffer to equilibrate the column for 10 minutes.
How do you clean HPLC lines?
Flush the column with appropriate solvent found in the column manual. Remove and seal column, and store according to good laboratory practice if needed. Flush system with water to remove buffer. Remove all samples from the sampler and store according to good laboratory practice.
How do you handle an HPLC instrument?
What is purging in HPLC?
Answer: “When setting up an HPLC system, the aim of the purge is simply to flush through all the lines so that any remaining solvent in them from a previous analysis or wash is replaced with the new mobile phase. As you would expect, the amount of new solvent required depends on the volume of the tubing in the system.
How do you bleed air from HPLC pump?
Treatment 1: Open a purge valve. Prime the pump with degassed solvent. Treatment 2: Open the compression fitting at the top of the outlet check valve with a slow pump flow until solvent leaks around the fitting. Tap the side of the check valve with a small wrench until small air bubbles come out with the liquid.
How do you stop peak tailing?
There are a few methods that can be used to avoid peak tailing:
- Operate at a lower pH.
- Use a highly deactivated column.
- Consider the possibility of mass overload.
- Consider the possibility of column bed deformation.
- Work at high pH when analyzing basic compounds.
- Use a sample clean-up procedure.
How do you solve problems in an HPLC system?
In an HPLC system, problems can arise from many sources. First define the problem, then isolate the source. Use HPLC Problem Index to determine which component (s) may be causing the trouble. A process of elimination will usually enable you to pinpoint the specific cause and correct the problem.
What are the most common HPLC pump problems?
HPLC pumps incorporate single or dual piston, syringe, or diaphragm pump designs. Pumping system problems are usually easy to spot and correct. Some of the more common symptoms are erratic retention times, noisy baselines, or spikes in the chromatogram. Leaks at pump fittings or seals will result in poor chromatography.
What are the important segments of an HPLC system?
The important segments of an HPLC system are the same, whether you use a modular system or a more sophisticated unit. Problems affecting overall system performance can arise in each component.
Where can I find online HPLC training courses?
Analytical Training Solutions, by Separation Science, offers online liquid chromatography (HPLC) training courses for analytical scientists. Skip to content Home Blog 日本語 Chromatography Forum