Resolution in life and mass spectrometers? It really depends…

By January 25, 2016

“How few there are who have courage enough to own their faults, or resolution enough to mend them.” Benjamin Franklin


As I sat one morning before work staring into my green tea leaves at the bottom of my cup and reflecting on my resolutions achieved and resolutions that evaded me in 2015, I resolved to make 2016 a better year. The ‘missed’ resolutions outweighed the ‘achieved’ and my mind, to protect itself, meandered and latched on to the word ‘resolution’. The meaning was not clear to me. The context seemed to be important. I peered deeper into the leaves. “Yes. It depends,” the leaves suggested. “Are you asking about life or mass spectrometers?”

In mass spectrometry, measured resolution is a function of the molecular weight of the compound but this number only tells a small part of the story. Published research has shown that mass resolution is a complex parameter and changes with mass and scan rate depending on the type of MS technology being utilized.

As shown in the graph below from a recent LCGC webinar by Bertrand Rochat, Research Project Leader, Faculte de Biologe et de Medecene of the Centre Hospitalier Universitraire Vaudois at Lausanne, for a trap-based system, the resolution drops off as the mass increases. The graph shows that the resolution is optimal at lower molecular weights (70, 000 at 200 m/z) and drops down to less than 40,000 at 800 m/z. This is due to the first principles of how a trap-based mass spectrometer functions (resolution is inversely proportional to the square root of the mass).

Resolution vs Mass v2

Time of flight (Tof) mass spectrometers, on the other hand, although possessing a lower resolution at molecular weights less than 200 m/z, maintain a consistent resolution and are not as impacted by further increases in molecular weight (40,000 at 800 m/z). The charge on the molecular ion affects the resolution observed (a higher charge state is consistent with a higher calculated resolution).

Mesmerized by the tea leaves, I wondered how fast the goodness from the tea would be absorbed. I wanted the results fast! I know I needed to do several experiments using a combination of techniques. Now I needed to consider separation techniques, sample complexity, scan rates, and software to interpret the data. Pondering about mass resolution had led me to think about a complete analysis—I digress.

It so happens that some instruments will be able to run faster analyses and maintain their high resolution better than others. Due to how trap-based instruments operate, any change in scan rate affects the resolution. As shown in the graph below (from a recent presentation from Mark Wrona, Senior Manager, Pharma Sciences at Waters Corporation, given at the European Bioanalysis forum), resolution falls off at higher scan rates.


Further, in any LC-MS experiment, the width of the chromatographic peak needs to be considered. The width of the peak will dictate the number and the types of mass spectrometric experiments one can perform across the eluting peak. If the duty cycle of the scan is longer, then fewer experiments can be executed and less data can be collected within that chromatographic window.

As with any analytical technique, there are trade-offs. Some analysts increase the time of the chromatographic separation to take maximum use of the slower scan speed on the instrument. This is one way to extract the best from the technology.

However, the growing demand to be more efficient strains this approach. Thankfully, the outcome of a more efficient, higher resolution, ultra-performance chromatographic run is a sharper chromatographic peak requiring a faster scanning instrument like a Tof-HRMS or tandem quad MS whose performances are not encumbered by the scan speed needed to complete the experiments or collect enough data points required for confident, high quality analytical data. There are alternatives—I can collect fast, confident data and provide insight in a more efficacious manner. I can determine if there was a pleasant surprise in my tea this morning in a timely manner.

It is interesting to see what the tea leaves have to say about resolution and its impact or lack thereof on mass accuracy and selectivity. Some business development managers have to work and can’t stare at leaves all morning long. Tune-in next time.

In the meantime, look at these links for more details on the relationships of resolution with molecular weight and scan speed.


Categories: Pharmaceutical