LCMS vs GCMS, as a method of analysis for Extractables and Leachables : Part 1 GC-MS analysis
Updated: Jan 7, 2021
Hi, time for another Blog Post. This time I like to focus on a couple of the analytical “workhorses” which are used for extractable and leachable studies.
When I started writing this post, I thought I could cover both GCMS and LCMS in the same post. However, as I began to write I realised there was a quite a lot to cover so I have decided I will split this up. This is Part 1, it covers GC-MS.
So why these two methods of analysis, well I haven’t done a full poll, but I wouldn’t mind betting that the vast majority of E&L testing done involves one or both these analytical methods. Why is that?
Leachable Analysis and most of the time Extractable Analysis is an exercise in trace analysis.
That is, the substances we are looking for are mostly found in the ppm or ppb range.
This means if we represent 1 ppm as a micro-gram per gram value we would be looking to detect 1 µg of a substance in 1 g of sample (and for 1 ppb it’s is 1 ng in 1 gram). That is a very small amount and it becomes even smaller when we consider how most chromatography is conducted.
For a typical Gas Chromatography method rather than inject all of your sample, you are looking to introduce around 1 µL onto the column to ensure you do not overload the injection port (LCMS is a similar volume 1-50 µL). This means if your solution has a concentration of 10 µg/mL (10 ng /µL) you will be introducing just 10 ng onto the column to be detected by the detector following chromatography.
Consequently as you can see, the amounts are very small and therefore detectors such as a Mass Spectrometer are needed to enable detection at the trace level.
There is a second important reason for use of GC-MS and LC-MS and that is selectivity.
As you can seen from the name LC-MS and GC-MS are hyphenated techniques, there is a coupling of Chromatography (LC or GC) with the Mass Spectrometer. The chromatography can separate the substances from the sample matrix and other substances present as they travel through the column and also act as a convenient method of introduction into the Mass Spectrometer (MS). When the substance enters the MS, it is also then possible to further separate the substances on the basis of their mass to charge ratio (m/z). Depending on the type of MS this can achieve a very high specificity for a given substance in the original sample.
This combination of sensitivity and selectivity, together with the possibility of quantification, marks out these techniques compared to other analytical techniques.
But I said I would discuss LC-MS vs GC-MS. So, what is better?
Well the best and simply answer is neither. Both have their place and I would not suggest you can completely ignore one and just use the other. However, I do want to point out some properties of each one and you might then decide for your application that one is more relevant.
So, starting with GC-MS what is important to realise is what the limits of each technique are and consider carefully what that means for your E&L project.
First point to consider:
The substance has to reach the detector to be detected
Sounds obviously doesn’t it, but it’s surprising how many factors there are to consider here.
First (and this is also true for LC-MS) the substances must be introduced into the GC-MS. If the substances don’t make it over the 1st step there is zero chance they will be detected.
In GC, that means the sample must be in the gas phase as it passes down the GC column. If it is not, the mobile phase (carrier gas) cannot transport the substances from the injection port on to column and no chromatography partitioning between (non-volatile liquid mostly but sometimes solid) phase and mobile phase can occur. To make it into the gaseous phase the substance either needs to be a gas to begin with or be converted into a gas (heated in the injection port or in the case of head-space or thermal desorption prior to introduction).
Only a relatively small proportion of all organic substances are volatile enough to achieve this, there is also the issue of stability to consider the substances must be stable enough not to react or degrade as they pass from the injection port and down the column. This is the Achilles’ Heel of GC.
There are ways to mitigate this, such as derivatisation to a more volatile form or high temperature columns but not all substances will elute from a GC column. So we are looking for extractables or leachables which are classified as volatile or semi-volatile. If you believe non-volatiles need to be detected you need to consider other methods of analysis (LCMS).
The second part of the process starts when the substance reaches the end of the chromatography column. In GCMS they are then subject to another process – ionisation. Without successful ionisation the substances will never reach the detector since it is the ionised substance which is then focused and directed towards the detector. Un-ionised substances simply go to waste. The exact mechanism of ionisation can vary. Both chemical ionisation (CI) and electron impact (EI) ionisation sources are used but the most common is EI. After ionisation the ions are directed to the mass analyser which again can vary depending on the type and there are many choices potentially here; Single Quadrupole, Triple Quadrupole, Ion Trap, Time of Flight and combination such as QToF.
So, with all these choices and factors in play the substance finally makes it to the detector and is recorded!
What happens after the substance is detected?
As stated, the most common ionisation is done with EI. With GCMS it is possible to generate consistent electron impact (EI) mass spectra operating at 70eV.
This universal standard leads to the ability to generate mass spectra which can be readily compared between runs and between different instruments, in turn this means that you can compare to commercial libraries of MS spectra. This is the “killer application” for GCMS allowing quick comparison and therefore the potential for identification of unknowns. This leads to other possibilities too, such as non-targeted screening due to the robustness of the equipment and repeatability between different manufacturer sources.
This is source is a blessing and a curse, the advantages are as stated but EI ionisation is hard ionisation. This causes the substance to be broken apart in the source to generate those characteristic ions represented in a mass spectrum. The downside of this process is it is sometimes very difficult to determine what the molecular weight of the intact substance is, since no molecular ion remains and for certain substance classes this leads to the inability to differentiate and identify between substances.
Here a quick example with a couple a spectra courtesy of NIST Webbook
Here is spectrum of C14 Hydrocarbon Tetradecane
and here is the spectrum of 1-Tetradecanol
See the problems here?
In both spectra the molecule ion is very small indeed, in fact in the case of the alcohol it is missing altogether.
The molecule weight of tetradecane is 198.3880, the alcohol is 214.3874 whilst you can just about see the 198 ion the 214 is not present, so it the absence of further information it is very difficult from the MS spectra to tell the difference, that is true of corresponding thiol too.
If I show you one more spectrum, the slightly more branched 3-methyl-tridecane (but still 198 molecule weight)
Then you can see the molecule ion is absence altogether.
Chemical ionisation sources can be used to induce a softer ionisation, but this does not guarantee a molecular ion and is challenging to setup without experience and skill.
However, the examples above do have different retention times on a capillary GC column thanks to the excellent resolving power of capillary GC, so all is not lost - identification is possible.
Well it is if you happen to have reference standards available to confirm your identity. In the world of E&L that may or mayn’t be the case. The life of the analytical chemist is a hard one!
Finally, I like to discuss briefly the quantitation possible with GC-MS. I would like to start by saying it is very possible to conduct quantitative analysis using GC-MS for Extractables and Leachables. There are as ever some caveats, firstly some elements of successful quantitation will depend on what type on mass analyser you are using. As a lot of people will be using single quadrupoles (SQ) I will discuss those. Other mass analysers present there are issues too, for example the limited linear range of some ToFs.
The SQ can be operated in two modes, Scan Mode or Single Ion Monitoring (SIM) Mode. In Scan Mode, the MS rapidly changes the voltages applied to quadrupoles over a short period of time so than a wide range of mass to charge ratios pass through the analyser in turn constructing the Mass Spectrum. This means a loss of sensitivity as a compromise as not much data is available per ion.
In SIM mode, you preselect a small number of ions you wish the analyser to let through to the detector. This enhances the sensitivity and collects much more data points for these ions but does not allow the generation of a full spectrum as you are not collecting data on other ions.
Generally speaking, quantitation is better when using SIM mode, signal to noise levels are higher and more data is collected across a typical GC peak (general rule of at least ten data points across the peak for good results).
Using SIM, it is possible to get excellent quantitative results; good precision, accuracy, linearity and limits of quantitation. However, you must be mindful of selection of correct ions and as with any trace analysis method where you are working close to the limits of detection there are many factors which can derail achieving satisfactory results.
If you need a fully quantitative method SIM is the method of choice, if you can live with semi-quantitative then SCAN mode remains on the table, since you are collecting all the ions in the molecule the response factor variation between substances is reduced. This is important for screening methods where you are looking to see if substances are breaching a generic Analytical Evaluation Threshold (AET) which perhaps in linked to a safety concern threshold.
So that is it for GCMS. There are a few areas here which I might revisit in future posts (Choice of Columns, Methods of Sample Introduction, Identification Classification etc). Use of GCMS in E&L analysis is a big topic area, so there is plenty more which might be discussed. I hope you find this content useful.
Comments and questions contact Maven E&L at www.MavenEandL.com
Next up Part 2: LC-MS for Extractable and Leachable Analysis