Ozone‐induced dissociation on a traveling wave high‐resolution mass spectrometer for determination of double‐bond position in lipids

Abstract

Rationale The position of C=C within fatty acyl chains affects the biological function of lipids. Ozone induced dissociation-mass spectrometry (OzID-MS) has great potential in determination of lipid double bond position, but has generally been implemented on low resolution ion trap mass spectrometers. In addition, most of the OzID-MS experiments carried out so far were focused on the sodiated adducts of lipids, fragmentation of the most commonly observed protonated ions generated in LC-MS based lipidomics workflow has been less explored. Methods Ozone generated in line from an ozone generator was connected to the trap and transfer gas supply line of a Synapt G2 high resolution mass spectrometer. Protonated ions of different phosphatidylcholines (PC) were generated by electrospray ionization through direct infusion. Different parameters, including traveling wave height and velocity, trap entrance and DC potential were adjusted to maximize the OzID efficiency. sn- positional isomers and cis/trans isomers of lipids were compared for their reactivity with ozone. Results Traveling Wave height and velocity were tuned to prolong the encounter time between lipid ions and ozone, and resulted in improved OzID efficiency, as did increasing trapping region DC and entrance potential. Under optimized settings, at least 1000 times enhancement in OzID efficiency was achieved compared to that under default settings for monounsaturated PC standards. Monounsaturated C=C in the sn-2 PC isomer reacted faster with ozone than the sn-1 isomer. Similarly, the C=C in trans PC reacted faster than in cis PC. Conclusions This is the first implementation of OzID in the trap and transfer region of a traveling wave enabled high resolution mass spectrometer. The OzID reaction efficiency is significantly improved by slowing down ions in the trap region for their prolonged interaction with ozone. This will facilitate application of high resolution OzID-MS in lipidomics.

Structural Analysis of Unsaturated Glycosphingolipids Using Shotgun Ozone-Induced Dissociation Mass Spectrometry

“…The OzID-MS experiments were performed on a modified Synapt™ G2 HDMS instrument (Waters, Manchester, UK) as reported previously [52]. Briefly, O 3 MEGA integrated ozone delivery system (MKS Inc., Andover, MA) was connected to the trap and transfer regions of the instrument through a three-way valve.…”

“…Traveling wave in the trap and transfer regions were operated at the following settings as optimized previously [52]: Entrance, 5.0 V; Bias, 2.0; Trap DC, 0.2; Exit, 0; trap wave velocity, 8 m/s; wave height, 0.2 V; transfer wave velocity, 247 m/s; wave height, 0.4 V. Default instrument settings for all the remaining parameters were applied. Under these settings, the reaction time between ozone and ions is estimated to be ~16.65 ms [52]. Instrument was calibrated daily in Resolution mode using sodium formate following manufacturer’s instructions obtaining less than 0.6 ppm (0.5 mDa) RMS residual mass.…”

“…Recently, we reported the modification and optimization of parameters of a traveling wave-based quadrupole time-of-flight (Q-ToF) mass spectrometer (Synapt™ G2 HDMS, Waters ® , UK) to allow the use of ozone as collision gas in the trap and transfer regions [52]. Herein, the said instrument, along with the previously optimized OzID settings [52], was applied to study the fragmentation of glycosphingolipids and demonstrate its capability to differentiate isobaric and isomeric glycosphingolipid species using shotgun approach.…”

“…Herein, the said instrument, along with the previously optimized OzID settings [52], was applied to study the fragmentation of glycosphingolipids and demonstrate its capability to differentiate isobaric and isomeric glycosphingolipid species using shotgun approach.…”

Collision-induced dissociation of doubly-charged barium-cationized lipids generated from liquid samples by atmospheric pressure matrix-assisted laser desorption/ionization provides structurally diagnostic product ions

“…It has been demonstrated that OzID is functional on chromatographic timescales, which is of importance for the separation of structural isomers by liquid chromatography (LC) and quantitative lipidomics [20,21]. Ozone was introduced to a high-pressure region of the mass spectrometer in order to perform online ozonolysis at the fatty acid double bonds [22,23]. Characteristic product ions, including radical cations, were produced from rapidly formed reaction products.…”