Quantitative analysis of chemoresistance-inducing fatty acid in food supplements using UHPLC-ESI-MS/MS.

Polyunsaturated fatty acids are important signaling molecules. A recent study reported hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid, 12-oxo-5Z,8E,10E-heptadecatrienoic acid, and (12S)-hydroxy-5Z,8E,10E-heptadecatrienoic acid as chemotherapy resistance-inducing factors when tumor cells were treated with cisplatin. Marine-based food supplements like fish oil or algae extracts are rich in polyunsaturated fatty acids and can contain large amounts of hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid. Thus, it was concluded that oral uptake of hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid might induce chemoresistance as shown in a mouse model. Cancer patients tend to consume food supplements containing polyunsaturated fatty acids on a regular basis. The uptake of hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid and (12S)-hydroxy-5Z,8E,10E-heptadecatrienoic acid should be controlled, because even low concentrations of 0.5 ng mL-1 showed chemoresistance-inducing effects in animal experiments. For accurate analysis of hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid and (12S)-hydroxy-5Z,8E,10E-heptadecatrienoic acid a validated method was developed by using ultrahigh-performance liquid chromatography hyphenated to quadrupole time of flight mass spectrometry via electrospray ionization and sample preparation by solid-phase extraction (SPE) with 3-aminopropyl silica. A combined targeted/untargeted approach was utilized using MS/MS by data-independent acquisition with SWATH and applied to commercial food supplements (refined fish oil, fish oil capsules, algae oil capsules, and flaxseed capsules). Accurate quantification of hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid and (12S)-hydroxy-5Z,8E,10E-heptadecatrienoic acid on the MS/MS level with simultaneous untargeted fatty acid screening revealed additional information. The LODs for hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid and (12S)-hydroxy-5Z,8E,10E-heptadecatrienoic acid were 0.036 ng mL-1 and 0.054 ng mL-1, respectively. Since hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid was present in the samples in large amounts and (12S)-hydroxy-5Z,8E,10E-heptadecatrienoic was not expected to be present in high concentrations, two calibration ranges, namely, 0.5-20 ng mL-1 and 5-200 ng mL-1, were validated. An untargeted screening identified 18-39 free fatty acids being present in the lipid extracts of the food supplement samples. Graphical abstract ᅟ.

Free fatty acid profiling in marine algae extract by LC-MS/MS and isolation as well as quantification of the ω-3 fatty acid hexadeca-4,7,10,13-tetraenoic acid.

Undaria pinnatifida (Wakame) alga contains high amounts of hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid which was reported to decrease the efficiency of cisplatin chemotherapeutics. To obtain a fatty acid enriched extract of this ω-3 poly-unsaturated fatty acid as an analytical standard, Wakame was used as source material for its extraction. A two-step extraction protocol consisting of a liquid-liquid extraction followed by solid-phase extraction with 3-aminopropyl silica in accordance to a normal-phase elution mode was developed. An ultra high performance liquid chromatography with electrospray ionization tandem mass spectrometry method based on sequential windowed acquisition of all theoretical fragment ion mass spectra allowed a simultaneous comprehensive group selective fatty acids profiling in untargeted manner and quantitative analysis of the targeted fatty acid. Hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid was identified using high-resolution product ion spectra. The quantitative method was based on d5-deuterated hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid which was employed as surrogate calibrant. Preliminary method validation was performed by evaluating detection and quantification limits, linear range, intra-assay and inter-day precision. Finally, a concentration of 421.2 ± 14.9 ng/mL (4% CV) of hexadeca-4Z,7Z,10Z,13Z-tetraenoic acid was determined in the extract which was further used as analytical standard.