Quantification of Trimethylamine-N-Oxide and Trimethylamine in Fish Oils for Human Consumption

Supplementing fish oil is one of the strategies to reduce the risk of cardiovascular disease, the leading cause of death around the world. Contradictorily, fish oil may also contain trimethylamine-N-oxide, a recently emerged risk factor for cardiovascular disease, as well as one of its precursors, trimethylamine. A method suitable for routine quantification of trimethylamine-N-oxide and trimethylamine in fish oil with a quick and easy liquid extraction without derivatization has been developed. Liquid chromatography with tandem mass spectrometry detection was employed along with a zwitterionic hydrophilic interaction liquid chromatography column and a gradient elution with eluents containing 50 mmol/L of ammonium formate. An internal standard (triethylamine) was used for quantification by mass spectrometry with an external calibration. The assay proved high linearity in the ranges of 10 to 100 ng/mL and 100 to 1000 ng/mL for trimethylamine-N-oxide and trimethylamine, respectively. The lowest limit of quantification was determined to be 100 µg/kg for trimethylamine and 10 µg/kg for trimethylamine-N-oxide, with the limit of detection at 5 µg/kg and 0.25 µg/kg, respectively. Accuracy ranged from 106-119%. Precision was below 7% the relative standard deviation for both analytes. The method was successfully applied for the determination of trimethylamine-N-oxide and trimethylamine contents in nine commercially available liquid fish oils and three commercially available fish oil capsules, showing that trimethylamine and trimethylamine-N-oxide are not present in highly refined fish oils.