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How to Develop GC Methods That Meet Both Technical and Business Demands

Use Pro EZGC Software to Quickly Compare Columns, Conditions, and MS versus Non-MS Detection

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Many factors go into GC method development: fast run times, adequate separations, symmetric peak shapes, good sensitivity, and many other chromatographic issues that need to be optimized. But business factors also need to be considered. Lab managers need to budget time for method development and weigh which instruments they want to use for specific methods in order to maximize productivity and profitability. Fortunately, using Restek’s free Pro EZGC chromatogram modeling software makes it easy to quickly evaluate methods virtually without tying up an instrument in the lab. And now it can also be used to help lab managers decide whether an MS or non-MS platform will be used.

Getting Started

Let’s say, for example, that we need to develop a new method for the 56 volatile compounds listed below, and we also want to determine whether GC-MS or GC with a non-MS detector is the best approach for our lab. Simulating separations with different columns and conditions using Pro EZGC software has been a useful tool for evaluating chromatographic parameters for years, but a new GC-MS mode that automatically targets isobaric compounds for separation allows us to more efficiently compare MS and non-MS methods.

Target Analytes

  • 2-Propanol
  • 1-Methoxypropan-2-ol
  • Octanal
  • Diisopropyl ether
  • Heptane
  • Cyclohexane
  • Ethyl acetate
  • Allyl alcohol
  • 2-(2-Butoxyethoxy)ethanol
  • 1,2-Dichlorobenzene
  • Methyl ethyl ketone
  • Toluene
  • Acetone
  • Limonene
  • Benzene
  • Pentyl acetate
  • Chloroform
  • Naphthalene
  • 1-Propanol
  • Decane
  • 2-(2-Methoxyethoxy)ethanol
  • Octane
  • Isopropyl acetate
  • Benzyl alcohol
  • Ethyl methacrylate
  • Pyridine
  • 2-Chlorotoluene
  • Isobutanol
  • Vinyl acetate
  • Acetonitrile
  • Methyl acrylate
  • Bromobenzene
  • n-Butyl acetate
  • 2-Butoxyethyl acetate
  • Pentane
  • Tetrahydrofuran
  • Nonane
  • Methylcyclohexane
  • o-Xylene
  • 4-Ethyltoluene
  • Ethylbenzene
  • 2-Hexanone
  • Acrylonitrile
  • Carbon tetrachloride
  • Propyl acetate
  • 1,1-Dichloroethane
  • 2-Butoxyethanol
  • Hexane
  • 2,2,4-Trimethylpentane
  • n-Butyl acrylate
  • 2-Phenylpropene
  • Ethanol
  • p-Xylene
  • iso-Pentane
  • Cumene
  • Dichloromethane

To get started, we go to ez.restek.com/proezgc; enter our analyte list in the Compounds tab (no experimental data needed); select a detection mode; and click the Solve button. GC mode will attempt to separate all compounds and will produce modeled results and conditions that could be used with a non-MS detector (e.g., FID or ECD). In contrast, GC-MS mode will automatically target only isobars because, when using MS, they are the only compounds that require chromatographic resolution. In both cases, the software will generate a list summarizing which column stationary phases could be used and how many compounds (GC) or isobars (GC-MS) they will separate (Figure 1).

Figure 1: In the Compounds tab, enter your analytes, select a detection mode, and click Solve to generate a list of stationary phases and a summary of the compounds they will separate.

GC Mode

modeler interface showing the compound tab and GC mode

 

GC-MS Mode

modeler interface showing the compound tab and GC-MS mode

Evaluating Initial Results

Initial results for the non-MS model show 41 out of 56 compounds can be fully resolved (Rs ≥1.5) on an Rtx-VMS column in an eight-minute run (Figure 2). This may be acceptable depending on which specific compounds individual labs are analyzing, what degree of resolution is needed, and whether a screening or quantitative method is being developed. For the MS model, nine isobaric ions are present, and the compounds sharing them are all resolved to baseline in a slightly longer nine-minute analysis (Figure 3). As shown in Figure 4, individual isobaric ions can be selected, and the modeler will highlight the separation of all compounds sharing that ion.

Figure 2: GC Model–Initial

Initial model output in GC mode

 

Figure 3: GC-MS Model–Initial

Initial model output in GC-MS mode

 

Figure 4: Selecting an ion from the Available Isobars field highlights the separation of compounds sharing that ion (e.g., m/z 43) and gives the resolution value of the closest eluting pair.

modeled chromatogram with an isobar (m/z 43) highlighted

 

From the initial simulated chromatogram results, we could select GC-MS for developing our final method in the lab since all the isobars are resolved in a reasonable analysis time, but what if we prefer to use a non-MS method for business reasons (e.g., the GC-MS is often unavailable, or we want to keep it open for more profitable samples)? Using the Pro EZGC chromatogram modeler, we can further optimize both methods to better inform our decision. 

Fine-Tuning Methods

For our purposes, we will select the Rtx-VMS column recommended by the software on the Compounds tab for both the GC and GC-MS models before we optimize them, but it is important to note that users can choose other columns from that list if desired. This can be useful if you want to evaluate columns that you already have, or if you want to see results on a different stationary phase before buying a new column.

To further refine our methods, we will now move to the Conditions tab. Here, models can be fine-tuned manually by adding gradients and/or modifying any of the input fields, but the simplest way to optimize a method is to use the Refine Oven Program button (Figure 5). As method parameters are changed, the results summary at the bottom of the Conditions tab and the simulated chromatogram and peak information are automatically updated. Users can also save models for easy reference and comparison.

Figure 5: Change parameters on the Conditions tab to optimize your separations (example shown is in GC-MS mode).

modeler interface showing the conditions tab

 

For our purposes, we used the Refine Oven Program button to generate the final optimized models shown in Figures 6 (GC mode) and 7 (GC-MS mode). The Refine Oven Program button can be clicked multiple times if run time or resolution can be improved with different temperature ramps, and it will disable and turn gray when no further automatic refinements are available.

Comparing the initial and final GC mode models, we can see that a slightly faster overall run time was achieved in the final optimized model and resolution was improved for some compounds but diminished for others. For example, 2-propanol and hexane (peaks 6 and 7) were not adequately resolved (Rs = 1.2) under the initial modeled conditions but were separated to baseline (Rs = 1.5) using the optimized conditions. In contrast, resolution between cyclohexane and chloroform (peaks 15 and 16) passed in the initial model (Rs = 1.7) but decreased in the optimized model (Rs = 1.2), highlighting the importance of labs evaluating results for their specific analyte lists against their own chromatographic requirements.

Figure 6: GC Model–Optimized

optimized model in GC mode

 

Similarly, the GC-MS model was further optimized using the Refine Oven Program button. In this case, as shown in Figure 7, baseline resolution was maintained for all the isobaric compounds, and the overall analysis time (termed “oven time” in the Results section of the Conditions tab) was reduced from 9.33 minutes to 8.10 minutes. While there are some coeluting compounds (e.g., allyl alcohol and vinyl acetate) in this model, it is not a concern because none of the coeluting compounds are isobars so the MS detector will be able to distinguish them. 

Figure 7: GC-MS Model–Optimized

optimized model in GC-MS mode

Choosing a Method

Deciding on a final method to establish in the lab should factor in both chromatographic performance and business interests. In our examples, the non-MS method was faster but contained coeluting compounds. If these compounds are critical analytes for a quantitative method, then the GC-MS method may be the preferred technical choice because it separates all isobars in a similar analysis time. However, if more profitable samples can be run on GC-MS instruments and labs can accept the degree of coelution among target compounds, developing the method on a non-MS instrument instead may be the better business decision. Using Pro EZGC chromatogram modeling software to examine different options before making a final decision allows labs to quickly and efficiently balance all interests and choose a sound method that meets technical requirements and business goals.

GNAR4407-UNV