Separation of C18:1 fatty acid isomers using cyclic ion mobility mass spectrometer

Authors: HAMEED, AHSAN - Arkansas Children'S Nutrition Research Center (ACNC); HAILEMARIAM, ABRHA ASSRESS - Arkansas Children'S Nutrition Research Center (ACNC); LAN, RENNY - Arkansas Children'S Nutrition Research Center (ACNC); FERRUZZI, MARIO - Arkansas Children'S Nutrition Research Center (ACNC); MORRIS, ANDRER - Arkansas Children'S Nutrition Research Center (ACNC)

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 3/6/2023
Publication Date: N/A
Citation: N/A

Interpretive Summary: Positional and geometrical isomers (isobaric compounds) have same molecular formula due to which it is challenging to identify and quantify them using conventional GC/LC-MS. However, as these isobaric compounds have different molecular structure so it is potentially possible completely identify and quantify them using recently introduced multi-pass WATERS SELECT SERIES cyclic ion mobility (Q-cIM-TOF) mass spectrometer. To demonstrate this, six fatty acid (FA) C18:1 positional and geometric isomers were separated. Cis and trans '9-C18:1 FA were well separated after 20 passes with a resolution of 290 O/'O. The arrival time distribution (ATD) for the cis and trans '9-C18:1 isomers were centered at 211.19 ms and 215.68 ms, respectively. Cis and trans '11-C18:1 was also separated after 20 passes with a resolution of 290 O/'O. However, ATD was slightly different: 209.68 and 214.16 ms for cis and trans '11-C18:1, respectively. Moreover, for cis and trans '6-C18:1, the ATD was slightly higher (i.e.,428.79 and 434.6) due to higher number of passes required for resolution (410 O/'O) with 60% valley. Separation of positional isomers of C18:1 FA was also tested using the cIM instrument. Interestingly, sufficient resolution of such isomers was achieved, except for cis '9-C18:1 and cis '11-C18:1, with fewer number of passes than the cis and trans isomers of the FA with same double bond position. Cis '6-C18:1 and cis '9-C18:1 was baseline resolved following 16 passes at ATD of 178.52 and173.77 ms, respectively. Similarly, the mobility spectrum of cis '6-C18:1 and '11-C18:1 showed two different components at ATD at 179.65 and 174.30 ms, respectively, following 16 passes. Likewise, sufficient resolution among trans-trans binary isomer mixture was also achieved further evidencing the capability of cIM to separate positional and geometric isomers of FA.

Technical Abstract: Introduction Fatty acid (FA) C18:1 has six positional and geometrical isomers namely oleic & elaidic acids ('9-cis C18:1 & '9-trans C18:1), vaccenic & transvaccenic acid ('11-cis C18:1 & '11-trans C18:1), and petroselinic & transpetroselinic acid ('6-cis C18:1 & '6-trans C18:1). These isomers not only have different physicochemical properties and biological functions but also involved in the etiology of several diseases. Therefore, it is very important to accurately identify and quantify them in biological samples. However, resolution of these isomers using conventional GC-MS/LC-MS is a significant research challenge. Recently, we demonstrated the complete separation and identification of these lipids isomers is possible using multi-pass WATERS SELECT SERIES cyclic ion mobility (Q-cIM-TOF) mass spectrometer. Methodology Six isomers of C18:1 FA were purchased and derivatized using AMP+ MaxSpec® Kit following the manufacturer’s protocol. IM separation of the FA isomers was performed on Q-cIM-TOF-MS. FA derivatives were infused directly and separation was achieved using a multi-pass traveling wave cIM separator. The instrument was operated in the positive mode using a capillary voltage of 3kV and desolavtion gas flow of 800 L/h. The desolvation and source temperatures were 550 and 120 °C, respectively. Following separation in the cIM, MS/MS spectra for the target mass was acquired for 2 min with trap collision energy of 55 V in a V-mode TOF geometry. The separation times for the cIM device were varied to acquire data from 1 to 50 passes of the cIM. The traveling wave static height was set to 22 V at a velocity of 375 m/s. MassLynx XX was used to control the instrument and UNIFI xx and DriftScope XX to process the obtained data. Preliminary data Binary FA C18:1 isomers were directly infused and looked upon for [M+H-AMP]+ (m/z = 449.3520) to confirm the sufficient resolution of isomers. For identification, MS/MS spectra was acquired post cIM separation in the transfer region between the cIM and the TOF entrance. Moreover, arrival order of each isomer was confirmed by infusing individual FA isomer and subjected to equal number of passes as mixture in the cIM cell. Overall, a different number of cycles through the IM separator was required to achieve resolution among different positional and/or geometric isomers. Cis and trans '9-C18:1 were well separated after 20 passes with a resolution of 290 O/'O. The arrival time distribution (ATD) for the cis and trans '9-C18:1 isomers were centered at 211.19 ms and 215.68 ms, respectively. Cis and trans '11-C18:1 were also separated after 20 passes with a resolution of 290 O/'O. However, ATD was slightly different: 209.68 and 214.16 ms for cis and trans '11-C18:1, respectively. Moreover, for cis and trans '6-C18:1, the ATD was slightly higher (i.e.428.79 and 434.6) due to higher number of passes required for resolution (410 O/'O) with 60% valley. Separation of positional isomers of C18:1 FA was also tested using the cIM instrument. Interestingly, sufficient resolution of such isomers was achieved, except for cis '9-C18:1 and cis '11-C18:1, with fewer number of passes than the cis and trans isomers of the FA with same double bond position. Cis '6-C18:1 and cis '9-C18:1 were baseline resolved following 16 passes at ATD of 178.52 and173.77 ms, respectively. Similarly, the mobility spectrum of cis '6-C18:1 and '11-C18:1 showed two different components at ATD at 179.65 and 174.30 ms, respectively, following 16 passes. Likewise, sufficient resolution among trans-trans binary isomer mixture was also achieved further evidencing the capability of cIM to separate positional and geometric isomers of FA. Novelty This work for the first time demonstrated the capability and application of cIM-TOF-MS for the separation of positional and geometrical isomers of FA.