Gas‐phase acid–base chemistry and its effects on mass isotopomer abundance measurements of biomolecular ions

Various parameters which affect mass isotopomer abundance measurements of derivatives of palmitic acid ionized by electron ionization (EI) and electron‐capture negative chemical ionization (ECNCI) were tested on a sector‐field double‐focusing mass spectrometer. Results on methyl palmitate ionized by EI are as follows: (i) sample size had a significant effect on mass isotopomer abundance ratios (MIARs); (ii) the electron multiplier gain of the detector also had an effect on MIARs; and (iii) ion scattering by ion–neutral collisions in the mass analyzer did not appear to have any significant effect on MIARs (under standard analysis conditions). However, ‘reagent’ gas pressure (methane) had a significant effect on MIARs of pentafluorylbenzyl palmitate ionized by ECNCI. It was concluded that there are two compensatory effects which alter MIARs of methyl palmitate ionized by EI: (i) gas‐phase acid–base chemistry in the source (specifically, proton transfer between fragment cations and neutral molecules); and (ii) detector non‐linearities, specifically, underestimation of less abundant isotopomers due to their signal disproportionately falling within the signal‐to‐noise ratio level of the electron multiplier. Gas‐phase chemistry is the dominant cause of inaccuracy in MIAR measurements for large sample sizes, while detector non‐linearity is the dominant cause of inaccuracy in MIAR measurements at small sample sizes. However, in a narrow intermediate range of sample size, these two effects balance each other and result in MIARs which are ‘acceptable’ when compared with the known MIAR values. It is emphasized that these two effects are present regardless of the type of mass analyzer used, e.g. quadrupole, sector‐field. Improvements in the accuracy of MIAR measurements will require developments in mass spectrometry aimed at eliminating each of the contributory effects. © 1998 John Wiley & Sons, Ltd.