Maybe you found a paper in which they used gas chromatography (GC) to analyze a sample that is similar to yours and now you are wondering if you should try it too. If you are hesitant because you know little about GC, then this article will help you with that.
You may already know that chromatography covers a wide range of analytical techniques that are used to separate, identify, and quantify compounds. If you are familiar with other chromatography methods, such as HPLC or GPC, then you already know more than you think!
When it comes to GC, the compounds that are analyzed are in, not surprisingly, the gas phase. This doesn’t mean that your sample has to be a gas (although it could). Instead, the gas chromatograph will vaporize it for you. After this, your material of interest will take a steamy journey through a column, where it is partitioned between a mobile phase and a stationary phase. The nature of this partitioning will dictate how much time a specific compound spends in the column until it eventually reaches the detector. This measure of time is called retention time (tR) and is determined by the detector when a compound exits the column. Figure 1 shows the basic components that make up a GC system.
How GC Works
Because the injector port is contained in an oven (see Figure 1), once you inject your sample it will be immediately vaporized (poof!). The gaseous molecules are then mixed with the mobile phase, which is also known as the carrier gas, because it does exactly that: it carries the sample through the column.
If you inject a mixture, the components will separate as they travel at different speeds. This happens because a compound can be either moving along with the carrier gas or spend some time attached to the stationary phase. The time that a compound spends interacting with the stationary phase will depend on the compound’s unique composition. Therefore, it is expected that a sample containing multiple substances will result in different retention times (each substance with its own tR).
To put into context, you can imagine that a compound that is “held up” by the stationary phase will have a longer tR than another that is traveling freely with the mobile phase. It is because of this principle that GC has its usefulness.
GC Components: A Closer Look
The type of sample can vary greatly depending on the study. Typically, GC samples consist of non-polar, low molecular weight, thermally stable, and vaporizable chemical material. It may be a single substance or a mixture, liquid or solid that is dissolved in solvent and only a tiny amount (few microliters) is injected via syringe. It is also possible to analyze gas samples (air, breath) and injection is usually through a gas valve.
This is where all the action (a.k.a separation of compounds) takes place. The column is a coiled tube made of metal or glass material that can withstand high temperatures and vary in length or diameter. Inside of the tube is the stationary phase. This can be a number of different materials with varying polarities, either solid or liquid that interact with the chemicals passing through. The stationary phase is assembled as a compacted material (packed column) or as a wall-coating film (capillary column).
The carrier gas is introduced from a gas cylinder into the gas chromatograph. It moves through the column at a constant flow rate and exits at the detector outlet. Unlike other methods, the mobile phase in GC does not interact with chemicals and only serves to carry them. Because of this, the carrier gas must be inert. Some examples are helium, nitrogen, and argon. The type of detector on the GC usually determines which gas is used. It is not something that you would have to decide since a working GC should already have a gas tank connected to it.
Like any other oven, a GC oven provides heat. But instead of baking goods, what this oven gives you is vaporized material right after injection. In addition, it keeps the column heated so that you continue to have gaseous molecules traveling through. Its temperature could vary from room temperature to 300 °C, though the limits vary by instrument. A temperature program can be set electronically to maintain a constant temperature or to gradually increase (ramping). The program that you select will depend on the nature of the sample.
This is a device at the end of the column that senses each compound as it comes out. The data recorded by the detector is transmitted into a computer and produces a two-dimensional plot, called chromatogram. There are several types of detectors with varying detection methods and limits. A particular powerful detector is the mass spectrometer (MS). A GC coupled with an MS, known as a GC/MS system, produces a mass spectrum in addition to the chromatogram.
Reading the Chromatogram
A GC chromatogram (Figure 2) is a visual output of the data recorded by the detector. It is presented as a plot of detector response (y-axis) versus retention time (x-axis).
The chromatogram can tell you a number of different things. I will break it down into three main categories:
- The nature of the sample. To evaluate the complexity of your sample you can count the number of peaks. Each compound detected by GC will appear as a single peak positioned at a specific tR. If you injected a mixture and the chromatogram shows three peaks, then this tells you that the sample had three different compounds. Now let’s say that you wanted to confirm the purity of a sample. In this case you expect a single peak, and hopefully that’s what you get!
- The identity of the sample. A substance can be characterized by matching its tR with a literature value or by injecting reference material. The catch with this is that you have to use the same conditions for each case. This is important because the tR depends on many factors other than compound identity, such as gas flow, temperature, and column length. The GC/MS systems are the most powerful for compound characterization as they enable identification by mass.
- The amount of sample. The peak area is proportional to the concentration of a substance in the original sample. Modern GC software will integrate the peak area and provide this information to you. The relative amounts of compounds in a sample can be determined by comparing the peak areas, or you can calculate the actual concentrations by using a standard calibration curve.
GC can be an intimidating technique at first but after giving it a couple of tries you will get the hang of it in no time! I would recommend watching this video for a visual tutorial on GC and reviewing this article for detailed information. Good luck!