Chromatography Forum

Hello all,

I am using a Waters 2695 along with a Waters 2996 PDA. I am quantifying an extraction, and suspect peak coeultion.

When peaks coelute, do their areas add up?

As an example, say two peaks have areas of 5 units and 40 units. If these two peaks were to elute, would the total area of the single peak be 45 units? Or, would the smaller peak get 'absorbed' into the larger peak, thus only reporting a single peak with an area of 40 units?

Thanks in advance for any insight provided!

Take a look at the Beer-Lambert Law and assume you are not deviating from it.

They add up. Whether its the same substance or two different substances - the more of it/them the more absorption i.e. higher peak i.e. larger peak area. The prerequisite is though you are working in the linear range.

Best Regards

Great! Thanks you guys for the helpful information.

What if you have negative abs for one peak?
I have seen cases where baseline dropped due to unknown component in the product.
Just another possibility...
Alfred

A negative peak usually means eluting substance that is more transparent than the mobile phase - under the given conditions. If a negative peak coincide a positive peak it will result in an overall reduction of the absorbance hence smaller positive peak. No miracles in such a case.

Best Regards