Chromatography Forum

Greetings:
I am helping to develop/validate a method with using a Chirex d-penicillamine column. The conditions are....

mobile phase - 1mM Cu(II) Acetate
Col temp. - 25C thermostatted
flow rate - 1 mL/min
detection - 230 nm
run time - 40 min (to elute acetate peak)

As mentioned in the subject line, the baseline wanders. It has been observed by multiple analysts using different instruments in different labs (countries). Per the vendor info, it is necessary to use the Cu-based mobile phase to impart the selectivity. Does any one have suggestion as to how to stabilize the baseline? And yes this is a chiral column being used for assay/impurity analysis but appears to be our best option.

thanks,
Robotjock

Robotjock,

Most likely the problem with baseline wander is related to temperature sensitivity. Since you are working with an eluent system with a substantial background absorbance at that wavelength, you can make expect it to be quite temperature sensitive. Assuming you do not have the option of switching to a more reasonable eluent system (it's never a great idea to add a detectable component to the eluent system as this sort of problem is always an unavoidable side effect), your best option is to work on finding strategies for better control of temperature. For example, make sure the instrument is well away ventilation vents in the laboratory environment, protect any transfer lines between the column compartment and the detector from exposure to laboratory temperature, consider the option of temperature control of the eluent containers and as a last resort you might benefit from placing the entire instrument in a controlled temperature environment.

Any idea on how much temperature variation is needed to be really noticeable on the baseline, 2°C, 5°C?

Thanks

BenNC,

It's difficult to say just how sensitive a given system will be to temperature as it depends upon the detector, the sensitivity requirements and the magnitude of eluent background at the operating wavelength. If you are trying to operate under conditions where your detector signal to noise ratio is minimal, you will generally need temperature control that is significantly better than +/- 1°C. Also, another thing to keep in mind is that if you have a component in your mobile phase that absorbs at your operating wavelength and is retained by your analytical column, temperature changes can indirectly affect the background by varying the partition coefficient of this component on your column. So, when working near the detection "floor" it is often necessary to control the temperature of your column as well as the temperature of the liquid entering the column.