Boosting narrow bore column performance

The benefits of high-efficiency, narrow-bore columns can only be fully realised when sample introduction is also optimised. Dr Mark Badger explores a novel way to improve chromatography and reproducibility.

Faster analysis times can improve lab productivity, but only if chromatographic performance still allows accurate peak identification and quantification. Narrow-bore GC columns (<0.25 mm ID) speed up analysis because they have greater chromatographic efficiency, which produces tall, narrow, symmetrical peaks that are easy to identify. While using narrow-bore columns can be a good approach to creating fast, highly efficient chromatography with a typical GC-MS setup, their effectiveness is limited by sample introduction.

A variety of sample introduction options are available but split/splitless inlets are the most common. For these inlets, liners with internal diameters of ~2 mm or ~4 mm are typically used. The smaller volume liners (~2 mm ID) transfer sample onto the column faster and in a narrow band, which can improve resolution, decrease the splitless hold time, and minimise adverse interactions, such as adsorption and reactivity. However, lower liner volume means less sample can be injected, which can reduce sensitivity and reproducibility. In contrast, larger volume liners (~4 mm ID) provide greater sample capacity, which can improve sensitivity and reproducibility, but their lower flow rates can cause band broadening, wider peaks, poor resolution, and analyte degradation due to the longer residence time in the liner. Ultimately, when choosing between larger and smaller volume liners, tradeoffs must be made in terms of capacity and chromatographic performance.

A more balanced solution

To give narrow-bore column users more flexibility and allow additional sample to be injected, without risking backflash or compromising on peak characteristics, Restek has developed an intermediate-volume (IV) liner, that has a 3 mm ID. Compared to smaller volume liners, IV liners allow injection volumes to be doubled because they have significantly more room to contain the solvent vapor cloud. And, since IV liners have faster sample loading capabilities than larger volume liners, good chromatographic performance is maintained because the analytes spend less time in the inlet.

Improved chromatography

To assess the performance of IV liners versus larger and smaller liners when used with narrow-bore columns, we compared peak characteristics and resolution of 51 semivolatile compounds analyzed on an Rxi-SVOCms column (20 m x 0.15 mm ID x 0.15 µm). To avoid exceeding maximum liner capacities, six 0.5 µL injections were made on the IV and smaller liners, and six 1 µL injections were made on the IV and larger liners.

When comparing the 1 µL injections, peak area and height were significantly improved (p < 0.05) for most compounds when using the IV liner, making peak identification and quantitation easier and more accurate. As shown in Figure 1, 92% of compounds showed greater average peak areas and 80% showed a greater average peak height when using the IV liner. Peak width, resolution, and symmetry were not statistically different from results for the larger volume liner for most compounds, which was attributed to solvent effects causing poor peak shape for some early eluting compounds using both liners. Performance did improve for early eluting compounds when less sample was injected as well as overall for later eluting compounds, which is likely the result of the IV liner’s faster sample transfer onto the column (Figure 2).

For the 0.5 µL injections, the difference was more dramatic: every peak parameter that was investigated showed statistically significant improvements when using the IV liner versus the smaller volume liner (Figure 1). These benefits can be attributed to the IV liner providing a narrow sample transfer band and maintaining resolution from the solvent peak. Improved performance was particularly beneficial for separating closely eluting isobaric semivolatiles, such as benzo[b]fluoranthene and benzo[k]fluoranthene (resolution of 1.83 vs. 1.93 on the smaller and IV liner, respectively).

Increased reproducibility

To evaluate how consistent chromatographic performance was, the relative standard deviations (%RSD) of averaged parameters across all compounds in all experiments were compared across the different liner sizes and injection volumes. As shown in Table I, the intermediate-volume IV liner demonstrated statistically better reproducibility for peak area and height on narrow-bore columns for both injection volumes and at both partial and full capacity. Results for peak symmetry were also more reproducible using an IV liner, except for the 0.5 µL injections where there was no statistical difference, and the mid-capacity comparison where symmetry was only 0.1% better using the larger volume liner.

Overall assessment

Table I summarises the statistical results of all the experiments and provides a high-level evaluation of the effects of liner choice on chromatographic performance when using narrow-bore GC columns with splitless injection. Of the 50 comparisons, 43 (86%) showed the intermediate-volume IV liner provided improved (22/50) or equivalent (21/50) performance compared to smaller and larger volume liners. Results were most pronounced for area and height, which are critical for accurate peak identification and quantification. Effects were more varied for width, resolution, and symmetry, but these parameters can also be strongly affected by the other method conditions (isothermal temperatures, column phase, solubility, etc.) and elution time.

Based on this data, IV liners provide more balanced and improved chromatographic performance compared to smaller and larger volume liners. When using IV liners with narrow-bore columns, more sample can be injected compared to smaller liners, reducing the negative effects caused by solvent and liner capacity. In addition, compared to larger volume liners, the sample is introduced onto the column more quickly and in a narrower band, taking better advantage of the column’s intrinsic high efficiency. In conclusion, the intermediate volume of IV liners contributes to fast, sensitive, and highly reproducible analyses, which allows labs to maximize the benefits of narrow-bore columns.

Dr Mark Badger is Product Manager at Restek.

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