High-resolution mass spectrometry and chromatographyEdexcel International A Level Chemistry: Revision notes

A low-resolution mass spectrum gives molecular ion peaks to the nearest whole number. A high-resolution mass spectrometer measures m/z to four decimal places. Isotopes have accurate masses that are not whole numbers (¹²C is defined as exactly 12.0000), so different molecular formulae with the same nominal Mr give different accurate Mr.

In the data booklet: H 1.0078, C 12.0000, N 14.0031, O 15.9949. For example CO = 27.9949, N₂ = 28.0062 and C₂H₄ = 28.0312, all nominally 28.

To calculate accurate Mr, add the accurate relative atomic masses of each atom and give the answer to four decimal places: propanone, C₃H₆O = 36.0000 + 6.0468 + 15.9949 = 58.0417.

The molecular ion peak of a high-resolution spectrum is matched against the calculated accurate Mr of each candidate formula. Example: a peak at m/z 149.1201 fits C₁₀H₁₅N (120.0000 + 15.1170 + 14.0031 = 149.1201) but not C₉H₁₁NO (108.0000 + 11.0858 + 14.0031 + 15.9949 = 149.0838).

The molecular formula can then be combined with fragment peaks, ¹³C and ¹H NMR and other data to suggest a structure. The molecular ion peak is the one at the highest m/z (ignoring small M+1 peaks from ¹³C).

Chromatography separates the components of a mixture using a mobile phase that moves over or through a stationary phase. Components separate because they differ in how strongly they are attracted to the stationary phase and how soluble they are in the mobile phase. A component that is more soluble in the mobile phase and less attracted to the stationary phase travels further or faster.

In paper chromatography the stationary phase is water bound to the cellulose fibres; in thin-layer chromatography (TLC) it is a thin layer of silica or alumina on a plate. The mobile phase is a solvent that rises up by capillary action.

For one-way paper or thin-layer chromatography, spots are measured from the start line (drawn in pencil so it does not dissolve) to the centre of each spot, and the solvent front is marked immediately.

Rf = distance moved by the spot ÷ distance moved by the solvent front

Rf is between 0 and 1 and has no units. Example: spot 5.6 cm, front 8.0 cm gives Rf = 0.70. Differences in Rf reflect differences in polarity or solubility: on polar silica with a non-polar solvent, a polar compound that hydrogen bonds strongly to the silica has a low Rf; a non-polar compound has a high Rf. Compare Rf values with known standards under identical conditions to identify components.

Gas chromatography (GC) has an inert gas (such as helium) as the mobile phase, a long column containing a liquid stationary phase on a solid support, and an oven. The sample is vaporised, so GC suits volatile compounds.

High-performance liquid chromatography (HPLC) pumps a liquid mobile phase at high pressure through a column packed with a solid stationary phase. It suits non-volatile or heat-sensitive compounds.

In both, each component leaves the column at a characteristic retention time, the time from injection to the peak at the detector, which is compared with standards. Peak areas show the relative amounts of each component.

In GC-MS or HPLC-MS the components leaving the column pass one at a time into a mass spectrometer. Chromatography separates the mixture and the mass spectrometer identifies each component from its accurate Mr and fragmentation pattern, which is more reliable than the retention time alone because different compounds can share the same retention time.

Applications include forensics, where traces at a crime scene are identified, and drug testing in sport, where banned substances in blood or urine are detected even at very low concentrations.