This Information Applies To: Agilent OpenLab CDS (ver 2.x)
Issue:
How to integrate non-ideally separated peaks starting above the zero baseline in OpenLab CDS
Resolution
Integrating merged peaks is a classical challenge in chromatography in cases where two compounds cannot be fully separated. This post outlines some key steps and references to helpful resources on how to address this problem.
What are merged peaks?
Merged peaks occur when a new peak begins before the end of peak is found. Figure 1 illustrates how the integrator deals with merged peaks.
Figure 1. Merged Peaks Example
How to integrate merged peaks in OpenLab CDS?
General recommendation is to optimize the chromatography to fully separate peaks.
In case you cannot avoid coelution, optimize the automated integration to avoid unnecessary use of manual integration, which is prone to human error and variation in interpretation
Begin by defining starting integration events, then fine-tune the integration results. (See Figure 2)
Figure 2. Baseline construction
Peak Valley Ratio
The Peak to valley ratio is a measure of quality, indicating how well the peak is separated from other substance peaks. This user-specified parameter is a constituent of advanced baseline tracking mode.
By adjusting the peak to valley ratio, you can decide if two peaks that do not show baseline separation are separated using a drop line or a valley baseline. (See video in Figure 4)
The peak-to-valley ratio is calculated using the following equations:
H1 ≥ H2, Peak valley ratio = H2/Hv
and
H1 < H2, Peak valley ratio = H1/Hv;
where H1=peak height of the first peak, H2=peak height of the second peak, Hv= height of the valley between the two split peaks. (see Figure 3)
Figure 4. Integrating merged peaks
If cannot be avoided, you can use manual integration, see OpenLab CDS tutorial - Manually integrate a peak | Agilent
Please note that in regulated environments you always need to refer to the guidelines outlined in your SOPs for integration parameters.
Learn how to effectively operate your OpenLab CDS: HPLC-OLII-1140e - Agilent Infinity LC systems with OpenLab CDS - Integration and viewing results GC-OLII-2134eV2 - Basic Integration of GC Data Using OpenLab CDS Data Analysis (Ver. 2.4) e-learning courses available from Agilent education |