Liquid Chromatography

A reader recently submitted this question: “Most HPLC methods call for preparing the sample in mobile phase as the solvent. I like to avoid using some of the organic solvents for sample preparation because of toxicity issues, so most of the time I...

A reader asks, I would like to split HPLC column effluent between an evaporative light scattering detector (ELSD) and a fraction collector. What is the best way of doing this, particularly as we may want a biased split, i.e., 10% to detector and 90%...

A reader asks, “I am using a shared equipment where somebody used tetrabutyl ammonium (TBA) as an ion pair in their mobile phase for chromatography. It looks to me like the ion pairing reagent was not washed completely from the column, even though I...

A reader recently asked how to determine the column volume of a column packed with superficially porous particles. You’ll recall that these are a newer type of HPLC packing material, variously referred to as superficially porous, core-shell,...

A reader observed baseline drift in his gradient method and wondered where it came from and what could be done to eliminate it. There are two main sources of baseline drift – temperature and mobile phase. If the column is operated in a column oven,...

One of the topics in my HPLC Troubleshooting: A Performance Qualification Approach Master Class is the importance of knowing how your HPLC system works when it is working correctly. Knowing the normal detector noise and drift can help you to realize...

In HPLC Solutions #109 we discussed a technique to measure noise and drift for an HPLC detector. There are several possible sources of noise in the detector, and the approach to reducing or eliminating such noise depends on the noise source. For the...

In HPLC Solutions #110 we looked at a problem of baseline noise created by a pump malfunction. In this instalment, we’ll look at what can be done about electronic noise in the chromatogram.

In the last HPLC Solutions #111, we looked at the influence of signal averaging on baseline noise for a simulated chromatogram. Let’s look now at how the detector time constant can be used to reduce baseline noise. The detector time constant (also...

In the last instalment of HPLC Solutions #112, we looked at one of the two most common ways to filter the detector signal to reduce baseline noise – the detector time constant. The second technique is to adjust the data system data rate. The data...

For the last few instalments of HPLC Solutions , we’ve been looking at ways to reduce baseline noise. We’ve been particularly focused on noise from electrical sources which tend to reflect the frequency of the power delivered to the laboratory. This...

A question that commonly arises in my Master Classes is the proper technique to integrate chromatograms where the peaks are not fully baseline resolved. For this, I most commonly draw on what I call the 10% Rule. This applies when a minor peak is...

Q: I have just developed a gradient HPLC method and need to set the acceptance criteria for system suitability. The column efficiency is one of the measurements we usually include in system suitability, but I don’t know which peak I should use to...

Most of us take advantage of the autozero function on our HPLC data systems to make the chromatogram look more pleasing to the eye. The autozero function is quite simple – it just takes whatever signal the data system is receiving and sets it at the...

Q: I’m wondering why most of scientists prefer to use a manual injector?! I know that autosamplers are used more to improve the productivity and it is standard with many HPLC systems. I was working with both and the autosampler produced better...