Table 2. Summary of Methods Used within Lipidomics.
| sample introduction | benefits/drawbacks |
|---|---|
| UHPLC | |
| reverse phase | + provides separation based on class and provides acyl selectivity |
| + usually results in class coelution | |
| + robust and reliable | |
| – may suffer from variable ionization throughout chromatogram | |
| HILIC | + provides separation based on class. Usually provides discrete class separation |
| + can provide more accurate quantitation through bracketing and single standard approaches | |
| – limited acyl selectivity | |
| SFC | |
| BEH | + provides separation based on class; usually provides discrete class separation. |
| + can provide rapid class-based lipid separation. | |
| – limited acyl selectivity for most current SFC column chemistries. | |
| HSS C18 | + provides separation based on class and provides a degree of acyl selectivity. |
| + usually provides discrete class separation. | |
| + can provide more accurate quantitation through bracketing and single standard approaches. | |
| – chromatographic behavior is sensitive to mobile phase and SFC parameters. | |
| direct infusion | + can provide rapid “shotgun” lipidomics analysis. |
| traditionally used with targeted triple quadrupole methods. | |
| + can be quantitative with bracketing. | |
| – limited structural detail. | |
| MALDI/DESI | + can provide in situ lipidomics imaging of samples. |
| traditionally coupled to a TOF detector. | |
| – usually not quantitative. | |
| – usually limited structural information. |
| instrument | benefits/drawbacks |
|---|---|
| triple quadrupole | + provides the highest sensitivity and linear range. |
| +/– usually operated in a targeted mode such as MRM or precursor scanning; not usually suited for exploratory lipidomics. | |
| time of flight | + mass accuracy is scan speed independent, providing high-resolution mass accuracy. |
| + faster scan speed is compatible with fast chromatography. | |
| – usually has a reduced linear range. | |
| – usually has a lower resolution than ion traps. | |
| ion trap | + can provide increased resolution. |
| + some setups provide MS3 and above. | |
| – mass accuracy is scan speed dependent. The number of data points versus mass accuracy needs to be considered. |
| technology | benefits/drawbacks |
|---|---|
| DDA acquisition | + can provide the cleanest mass spectral fragmentation for identification. |
| – commonly used with ion traps. Top “n” scans can result in complex multicompound spectra; can also increase duty cycle. | |
| – software can fail to fragment peaks; can require inclusion/exclusion lists. | |
| MSe, Sonar, SWATH | + data independence allows for all fragmentation data to be acquired with minimal setup. |
| usually found on TOF systems. | |
| – data requires deconvolution via software. Fragmentation data may still contain multiple compounds, dependent on the sample matrix. | |
| Ion mobility | + rapidly emerging, powerful technology, which provides complementary separation based on cross-sectional area; allows for a degree of separation even without chromatography. |
| + likely to be incorporated into modern instruments. |