We are frequently asked about issues with reduced peak size in gas chromatography (GC), and I’m guessing this is related to just how difficult this problem is to troubleshoot. There are so many potential causes that an inexperienced GC user may not know where to begin the troubleshooting process. Fear not. What follows is our logical guide to locating and fixing the issues with loss of sensitivity, and we’ve tried to cover as many of the instrument and application issues that we can think of.
We are frequently asked about issues with reduced peak size in gas chromatography (GC), and I’m guessing this is related to just how difficult this problem is to troubleshoot. This isn’t necessarily related to the complexity of the problem. It has more to do with the number of possible causes. There are so many potential causes that an inexperienced GC user may not know where to begin the troubleshooting process.
Fear not. What follows is our logical guide to locating and fixing the issues with loss of sensitivity, and we’ve tried to cover as many of the instrument and application issues that we can think of.
Step 1 – Define the Problem
Sensitivity problems can be grouped by way of their presentation within the chromatogram:
1. All peaks sizes (heights and areas) decrease– retention times do not change
2. All peaks sizes (heights and areas) decrease– retention times shift, no evidence of loss of efficiency (peak broadening)
3. All peaks sizes (heights and areas) decrease –peaks are broadened, retention time shift may occur
4. Early-eluting peak sizes are reduced, later-eluting peaks remain constant
5. Later-eluting peaks appear smaller, earlier peak sizes remain constant
6. Specific peaks within the chromatogram are smaller, whilewhile others remain constant (or become relatively larger)
We should attempt to identify which of the above categories our problem falls into, as this directly influences the sources of the potential problem, and consequently, the actions we take to investigate and solve the problem.
All peaks sizes (heights and or areas) decrease–retention times do not change
Figure 1: Reduction in peak area for all analytes in a chromatogram, retention time and peak width remain constant
As the description of this problem suggests, whatever the issue, it seems to be affecting all analytes equally, which leads us to think about physical problems with the instrument settings (including the autosampler) rather than thinking about chemical issues.
As always, it’s best to start with the most obvious possible causes, as they are typically easier to check, verify, and correct.
Figure 2: Reduction in peak area of all chromatographic peaks with accompanying shift in retention time
Where peak height or area reductions are accompanied by shifts in retention time but without noticeable reduction in efficiency, for example, peaks do not broaden significantly, we begin to think about carrier gas flows and the column dimensions.
All peaks sizes (heights and or areas) decrease–peaks are broadened
Figure 3: Reduction in peak height or area with accompanying band broadening
When all peak heights or areas reduce and the peaks broaden, the most obvious cause is a loss of efficiency within the chromatographic system. This phenomenon should also lead a reduction in signal-to-noise ratio for analyte peaks.
When all peak heights or areas reduce and the peaks broaden, the most obvious cause is a loss of efficiency within the chromatographic system. This phenomenon should also lead a reduction in signal-to-noise ratio for analyte peaks.
Figure 4: Lower data sampling rate causing issues with reduced peak height and increased peak width
As shown in Figure 4, incorrect detector settings may also cause a reduction in peak height, accompanied by peak broadening.
Early-eluting poeak sizes are reduced, later-eluting peaks remain constant
Figure 5: Injection of n-alkanes in which early-eluting peaks are significantly reduced in height and area, which is accompanied by poor peak shape for the early-eluting compounds. Top chromatogram starts with oven temperature too high to achieve solvent focussing, the problem is alleviated in the bottom chromatogram as the initial over temperature is lowered
The reduction in peak height or area of early-eluting peaks may point to some specific problems within the GC inlet or method acquisition conditions. For some of these problems, the reduction in peak size of the early-eluting compounds is also accompanied by a deterioration in peak shape
Later-eluting peaks appear smaller, earlier peak sizes remain constant
Figure 6: Loss of later-eluting compounds due to analyte discrimination with in the split or splitless inlet
Again, the loss of sensitivity for later-eluting peaks can lead to specific troubleshooting investigations which focus on the GC inlet and sample introduction processes. This phenomenon is known as sample discrimination, and occurs due to the higher boiling nature of later-eluting analytes.
Specific peaks within the chromatogram are smaller, while others remain constant (or become relatively larger)
Figure 7: Loss of sensitivity with certain analytes due to GC system activity and adsorption of more polar analyte species
When certain analytes (often initially appearing to be randomly spaced throughout the chromatogram or just one or two analyte peaks which are not closely eluting) then this symptom can appear very mysterious. Further, this problem can often be accompanied by a deterioration in peak shape of the affected peaks. When these symptoms occur, then our thoughts typically turn to matters of chemistry, and to finding a link between the analytes that have been affected. Typically, one may be able to establish a link such as a similar analyte chemistry, and in particular, more polar analytes are seen to be the linking factor. When polar analytes encounter absorptive groups (silanol groups from silica-based materials or active metal surfaces) within the inlet liner or GC column, then analyte adsorption may occur, significantly reducing the number of analytes that reach the GC detector.
As I said in the introduction, a bewildering number of issues can lead to a reduction in instrument sensitivity when using GC or GC–MS instruments, and it is important to identify the symptoms as described above in order to perform the correct troubleshooting checks and remedial actions. This guide should also help you to perform the troubleshooting investigations in the most efficient manner, starting with the most common or simplest checks initially, to save both time and effort in identifying and correcting the problems that have led to the reduction in peak height or area.
Tony Taylor is the Chief Scientific Officer of Arch Sciences Group and the Technical Director of CHROMacademy. His background is in pharmaceutical R&D and polymer chemistry, but he has spent the past 20 years in training and consulting, working with Crawford Scientific Group clients to ensure they attain the very best analytical science possible. He has trained and consulted with thousands of analytical chemists globally and is passionate about professional development in separation science, developing CHROMacademy as a means to provide high-quality online education to analytical chemists. His current research interests include HPLC column selectivity codification, advanced automated sample preparation, and LC–MS and GC–MS for materials characterization, especially in the field of extractables and leachables analysis.
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