Optimizing Sensitivity in GC–FID Analysis

It is often possible to achieve better sensitivity and lower limits of detection and quantitation using standard gas chromatography (GC) equipment, such as a standard split–splitless injection port and a flame ionization detector (FID). Paying attention to fundamental variables can lead to much improved method performance, Here, we consider some important aspects necessary to optimize GC–FID performance.

Sample Solvent

By choosing a sample solvent that matches the stationary phase polarity as closely as possible, peak shape will be optimized with no tailing or broadening, which might otherwise reduce the signal-to-noise ratio. Here, we think of choosing nonpolar solvents such as n-hexane with a non-polar stationary phase such as 1% polydimethyl siloxane (DB1, RTX-1) or methanol with polar stationary phases such as waxes.

High Head-Pressure (Pressure Pulsed) Injection

The inlet liner is designed to hold a certain volume of gas (typically 500 to 950 mL), and we need to avoid overfilling the liner with sample gas, or problems will occur such as poor quantitative reproducibility and carry over. By choosing a sample solvent with a lower expansion coefficient, and increasing the pressure within the liner during the injection, to constrain the volume of sample gas created, larger injection volumes can be made. Calculators, typically known as backflash or vapor volume calculators, can be used to determine the maximum sample volume that can be injected before liner overfilling occurs.

Optimize Splitless Time

In splitless injection mode, the split vent valve is closed, and all of the contents of the inlet are directed into the GC column. After a predetermined length of time (the splitless time), the split vent value is opened, and the remaining inlet contents are vented to the atmosphere. This helps to prevent wide and badly tailing solvent peaks, increased baseline noise, and a rising baseline during the analysis. If the split vent is opened too early, there is a risk of analyte loss, and, if opened too late, the broad and tailing solvent peak may reduce the sensitivity of early eluting analytes; and noisier baselines will reduce the signal-to-noise ratio for all analytes. One must experimentally determine the optimum splitless time to achieve the best reproducibility and sensitivity from the splitless injection. For proper solvent focussing to occur, the initial oven temperature must be maintained around 20 °C below the boiling point of the sample solvent during the injection phase of the analysis.

Initial Oven Temperature Hold

On-column sample focusing during splitless injection takes a finite amount of time, and, during this time, the oven temperature must be held constant. However, if this initial hold time is too long, some degree of analyte dispersion may occur, reducing the peak efficiency, and, hence, the signal-to-noise ratio. The initial hold time should be carefully evaluated if optimum sensitivity is to be achieved.

GC Column Choice

Shorter columns (10–15 m) with narrow i.d. (0.18–0.25 mm) and thin films (<0.3 mm) will give the best peak efficiencies, and therefore the optimum signal-to-noise ratio. Further, less-polar stationary phase chemistries will show less inherent bleed, again improving the signal-to-noise ratio, although the choice of stationary phase chemistry will be sample dependent. One should choose the least polar column with the thinnest film that results in a satisfactory separation, in order to reduce column bleed to the minimum amount possible.

Carrier Gas Operating Mode

Ensure that the carrier gas operating mode is set to constant flow, rather than constant pressure, to ensure that the carrier flows at the same linear velocity during the whole temperature program. This approach prevents the carrier from slowing as the temperature increases, which can lead to broadening of later-eluted peaks and suboptimal detector response.

FID Optimization

Optimize the fuel (hydrogen) to oxidizer (air) ratio of the FID to ensure the best response for your analytes; typically, start with a 10:1 ratio and adjust the fuel gas in steps of +/- 5 mL/min. Also, ensure that the make-up gas flow rate (nitrogen is recommended by many manufacturers as the most effective make-up gas) is optimized, and note that the make-up gas flow rate can have a pronounced effect on analyte sensitivity. Start with a 1:1 ratio of make-up gas to fuel gas (typically hydrogen), and adjust in +/- 5 mL/min. steps to investigate the optimum range.

Thermal Gradients

Ballistic (short and fast) thermal gradients will produce the sharpest peaks and the best signal-to-noise ratios, but ensure the oven heater motor can follow the required temperature profile in a reproducible way. Of course, the use of ballistic thermal gradients will depend upon the separation requirements and the complexity of the sample.