Multiplex binary gradient elution profile for quantitation of trimethylamine-N-oxide (TMAO) by LC/MS/MS. Two different solvents, A, 0.1% propanoic acid in water; B, 0.1% acetic acid in methanol, were used to generate the gradient. a1 and a2, time point for sample injection on column 1 and 2 respectively; b1 and b2, time point for start of data collection on column 1 and 2 respectively; c1, c2, time point for data collection to end and column 1 and 2 to be switched to waste, respectively.