Reduced peak height

[AnaChem1983]

Hi folks,

The heights of the peaks in my application have somewhat abruptly decreased to approximately 60% of their usual height. This is based on recoveries in a spiked sample being around 60% of the true value (normal values usually hover around 100%). The application is the separation of formic, acetic and propionic acid on an Agilent Hi-Plex H column (300mm x 7.7mm, 8um with the use of matching guard column) with 5mM H2SO4 as the mobile phase. 40 deg C column temperature, 0.7 ml/min flow rate. RID detection at 55 deg C.

I have eliminated a simple sample prep issue by preparing fresh SST and spike samples and observing the same results. The retention of the compounds has not changed and matches up well with previous runs which suggests that the mobile phase is OK.

The performance of the column has slowly been going down over the past couple of months when looking at parameters such as resolution of a critical pair, plate number and tailing of a specific peak. I initially thought, therefore, that the reduction of peak height was linked. However, upon changing the guard column, the parameters have been restored to values we recorded when first using the column. The peak heights however have remained at their depreciated levels.

System pressure has not increased or decreased to a significant degree.

Going from memory (I'm sat at home at the moment) I can't remember if the solvent peak is the same size as usual as my next step for the morning is to see if there's a problem with the actual injection volume. I have a few ideas for things to try out next but thought I would post here to see if anyone has any advice or if they've seen anything like this before.

[tkubowicz]

Hello

If you observe peak area lower than "normal" value (or let say normal range) I would say it is problem could be linked with "injection" system. So I'd recommend to check:
- RSD for 5-6 injection. If it is far away from 1% I would check sampler components: sample loop, needle seat, needle, syringe, rotor seal (perhaps blocked or worn out),
-if RSD is less than 1-2% and peaks area ar still smaller check injector parameters like washing solvents or washing steps, check vials, perhaps metering device or check method for any changes in injection volume or sample loop volume

Please note that problem could be with RI detector as well - make sure that analytical and reference cells are properly flushed and diodes are balanced. Also make sure that baseline is stable and there is no drift due to temperature fluctuations.

Regards

Tomasz Kubowicz

[AnaChem1983]

Hi,

Thank you for your input. It transpired that it was indeed the RI detector. I gave the reference cell a good 20 minute purge this morning and it has solved the problem. It's funny because the software was showing the RID as being OK for analysis prior to this purge.

Anyway, problem solved