Quantitative LC-MS Metabolite Profiling

LC-MS differentiation and detection of Erdman, CDC1551, and mutant strains were performed with an Agilent Accurate mass 6230 time of flight (TOF) device coupled with an Agilent 1290 liquid chromatography system using solvents and configuration as previously described (11 (link), 13 (link)). An isocratic pump was used for continuous infusion of a reference mass solution to allow mass axis calibration. Detected ions were classified as metabolites based on unique accurate mass-retention time identifiers for masses showing the expected distribution of accompanying isotopologues. Metabolites were analyzed using Agilent Qualitative Analysis B.07.00 and Profinder B.06.00 software (Agilent Technologies, Santa Clara, CA, USA) with a mass tolerance of <0.005 Da. Standards of authentic chemicals of known amounts were mixed with bacterial lysates and analyzed to generate the standard curves used to quantify metabolite levels.

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Quinonez C.G., Lee J.J., Lim J., Odell M., Lawson C.P., Anyogu A., Raheem S, & Eoh H. (2022). The Role of Fatty Acid Metabolism in Drug Tolerance of Mycobacterium tuberculosis. mBio, 13(1), e03559-21.

Publication 2022

Accurate mass 6230 time of flight (TOF Agilent Qualitative Analysis B.07.00 Profinder B.06.00

Axis Bacterial lysates Device Ions Liquid chromatography Retention Solvents Strains Tolerance

Corresponding Organization : University of Westminster

Other organizations : University of Lincoln, University of Strathclyde, University of West London

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Variable analysis

independent variables

Erdman strain
CDC1551 strain
Mutant strains

dependent variables

Metabolite levels
Metabolite identification based on accurate mass-retention time

control variables

Solvents and configuration as previously described (11, 13)
Isocratic pump for continuous infusion of reference mass solution to allow mass axis calibration
Mass tolerance of <0.005 Da for metabolite analysis
Standards of authentic chemicals of known amounts mixed with bacterial lysates to generate standard curves for metabolite quantification

controls

Positive control: Standards of authentic chemicals of known amounts
Negative control: Not explicitly mentioned

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