Development of a Retention Time Interpolation scale (RTi) for liquid chromatography coupled to mass spectrometry in...

The accuracy and sensitivity of high resolution mass spectrometry (HRMS) enables the identification of candidate compounds with the use of mass spectrometric databases among other tools. However, retention time (RT) data in identification workflows has been sparingly used since it could be strongly affected by matrix or chromatographic performance. Retention Time Interpolation scaling (RTi) strategies can provide a more robust and valuable information than RT, gaining more confidence in the identification of candidate compounds in comparison to an analytical standard. Up to our knowledge, no RTi has been developed for LC-HRMS systems providing information when acquiring in either positive or negative ionization modes. In this work, an RTi strategy was developed by means of the use of 16 isotopically labelled reference standards, which can be spiked into a real sample without resulting in possible false positives or negatives. For testing the RTi performance, a mixture of several reference standards, emulating suspect analytes, were used. RTi values for these compounds were calculated both in solvent and spiked in a real matrix to assess the effect of either chromatographic parameters or matrix in different scenarios. It has been demonstrated that the variation of injection volume, chromatographic gradient and initial percentage of organic solvent injected does not considerably affect RTi calculation. Column aging and solid support of the stationary phase of the column, however, showed strong effects on the elution of several test compounds. Yet, RTi permitted the correction of elution shifts of most compounds. Furthermore, RTi was tested in 47 different matrices from food, biological, animal feeding and environmental origin. The application of RTi in both positive and negative ionization modes showed in general satisfactory results for most matrices studied. The RTi developed can be used in future LC-HRMS screening analysis giving an additional parameter, which facilitates tedious processing tasks and gain more confidence in the identification of (non)-suspect analytes.