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First an average mass spectrum is generated from different regions of the chromatogram that contain internal standards (Fig. 3(a)). Since the accurate masses of the standards are known, the average spectrum can be used to re-calibrate the entire run (Fig. 3(b)). 0 mDa after re-calibration (Fig. 3(c)). Samples should be analyzed in random order. Blanks, quality control samples such as pools of individual samples, and calibration controls should be interspersed. The addition of blanks helps to identify contamination peaks that originate from solvents, extraction solutions, impurities from internal standards or the column.

1 ppm mass accuracy when isotope information is not considered. g. the SmartFormula tool from Bruker Daltonics. The quality of the isotopic fit of the experimental pattern versus pattern of a proposed sum formula is assessed by the mSigma value, which is 0 for a perfect match and 1000 in the absence of a match. Algorithms for calculating elemental formulas additionally follow chemical and heuristic rules such as default element ratios or restrictions on the number of elements, to further exclude unlikely elemental combinations (Kind and Fiehn, 2007).

That’s why absolute amounts obtained from newly developed methods should always be cross-validated with established techniques. Stable-isotope-labeled internal standards are, meanwhile, commonplace to meet the requirements for generating accurate and reproducible quantitative data. g. 13C, 15N, or 2H-labeled compounds, is expected to be equal to the ion suppression observed for the analyte, as the chromatographic behavior and ionization are pretty much identical. One should note that deuterated internal standards, depending on the number of deuterium atoms incorporated, might show a partial separation from the unlabeled analyte.