Statistical precision of the intensities retrieved from constrained fitting of overlapping peaks in high-resolution mass spectra

[摘要] Least-squares fitting of overlapping peaks is often needed to separatelyquantify ions in high-resolution mass spectrometer data. A statisticalsimulation approach is used to assess the statistical precision of theretrieved peak intensities. The sensitivity of the fitted peak intensitiesto statistical noise due to ion counting is probed for synthetic datasystems consisting of two overlapping ion peaks whose positions arepre-defined and fixed in the fitting procedure. The fitted intensities aresensitive to imperfections in the m/Q calibration. These propagate as alimiting precision in the fitted intensities that may greatly exceed theprecision arising from counting statistics. The precision on the fitted peakintensity falls into one of three regimes. In the "counting-limitedregime" (regime I), above a peak separation χ ~ 2 to 3half-widths at half-maximum (HWHM), the intensity precision is similar tothat due to counting error for an isolated ion. For smaller χ andhigher ion counts (~ 1000 and higher), the intensity precisionrapidly degrades as the peak separation is reduced ("calibration-limitedregime", regime II). Alternatively for χ < 1.6 but lower ioncounts (e.g. 10–100) the intensity precision is dominated by the additionalion count noise from the overlapping ion and is not affected by theimprecision in the m/Q calibration ("overlapping-limited regime", regimeIII). The transition between the counting and m/Q calibration-limited regimesis shown to be weakly dependent on resolving power and data spacing and canthus be approximated by a simple parameterisation based only on peakintensity ratios and separation. A simple equation can be used to findpotentially problematic ion pairs when evaluating results from fittedspectra containing many ions. Longer integration times can improve theprecision in regimes I and III, but a given ion pair can only be moved outof regime II through increased spectrometer resolving power. Studiespresenting data obtained from least-squares fitting procedures applied tomass spectral peaks should explicitly consider these limits on statisticalprecision.

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