Shotgun lipidomics approaches

Based on these unique features, at least three different approaches of shotgun lipidomics are developed and well documented in the literature, including tandem MS-based shotgun lipidomics, high mass accuracy-based shotgun lipidomics, and multidimensional MS-based shotgun lipidomics. 

2 High Mass Accuracy-Based Shotgun Lipidomics Currently, commercially available hybrid instruments (e.g., quadrupole-time-of-flight (Q-TOF) or Q-Exactive (i.e., quadrupole-Orbitrap) mass spectrometers) offer an improved duty cycle that increases the detection sensitivity and/or high mass resolution/accuracy [18, 19]. These instruments can thus be employed to quickly perform product-ion 

The differential charge properties of different lipid classes (which are predominant with the head groups of polar lipid classes) are exploited to selectively ionize a certain category of lipid classes under multiplexed experimental conditions to separate many lipid classes in the ion source (i.e intrasource separation) [33]. This separation method is analogous to the electrophoretic separation of different compounds that possess different pi values [33] (see Chapter 2 for details). 

The concept of building blocks in lipid structure (see above) is fully employed for the identification of individual lipid species in MDMS [11, 34] because these building blocks can be determined with two powerful tandem MS techniques 

In theory, to fully investigate the effects of ionization conditions on ionization efficiency and/or the effects of collision conditions on fragmentation processes or other effects, a variety of ionization voltages, ionization temperatures, collision energies, collision gas pressures, etc., should be employed in an experiment (see Chapter 4). These variables can all be logically varied unit by unit within a certain range. Therefore, from a new set of spectra when an individual variable of MS is ramped, a new dimension is added to the basic 2D MS. All of these dimensions form the family of MDMS [11]. Specifically, MDMS is defined as the comprehensive MS analyses conducted under a variety of instrumental variables that collectively comprise an MDMS spectrum. 

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Through global lipidomics, each distinct lipid species present in a cell s lip-idome can be identified. A shotgun lipidomics approach, which uses an ESI-intrasource separation of Upids from a complex extract, multidimensional mass spectrometry, and computer-assisted array analysis, is described. 

MDMS-based shotgun lipidomics overcomes the majority of the limitations of other shotgun lipidomics approaches and possesses many obvious advantages as follows ... 

Limitations in MDMS-based shotgun lipidomics along with other shotgun lipidomics approaches include the following ... 

Although MDMS-based shotgun lipidomics identifies and quantifies all individual species of a characterized lipid class in an unbiased manner within the limits of instrumentation sensitivities, the approach is not ideal for identification and quantitation of species of an unknown or uncharacterized lipid class since identification of the building blocks of a lipid class has to be pre-determined. Its throughput is relatively lower compared to the other two shotgun lipidomics approaches. 

Bowden, J.A., Bangma, J.T. and Kucklick, J.R. (2014) Development of an automated multi-injection shotgun lipidomics approach using a triple quadrupole mass spectrometer. Lipids 49, 609-619. 

Jiang, X. and Han, X. (2006) Characterization and direct quantitation of sphingoid base-1-phosphates from lipid extracts A shotgun lipidomics approach. J. Lipid Res. 47, 1865-1873. 

Wang, M., Hayakawa, J., Yang, K. and Han, X. (2014) Characterization and quantification of diacylgjycerol species in biological extracts after one-step derivatization A shotgun lipidomics approach. Anal. Chem. 86, 2146-2155. 

In the high mass accuracy/resolution mass spectrometry-based shotgun lipidomics approach, the overlaps between SM and M-f1 C isotopologue of PC species, and between PC subclasses can be resolved [35]. Moreover, the linkages of alkyl vs. alkenyl at the sn-1 position of glycerol and the fatty acyl chains of diacyl species are also identified by product-ion analysis [35,36]. Clearly, this approach provides much more structural information than that of the tandem MS approach as aforementioned. 

S LipidSearch LipidSearch is the commercial software (Thermo Fisher Scientific) developed jointly by Prof. Ryo Taguchi and MKI (Tokyo, Japan). It is a powerful new tool for automatic identification and relative quantification of cellular lipid species from a large amount of mass spectrometric data obtained from both LC-MS and shotgun lipidomics approaches. A lipid database containing... 

Yang, K., Zhao, Z., Gross, R.W., Han, X. (2011) Identification and quantitation of unsaturated fatty acid isomers by electrospray ionization tandem mass spectrometry A shotgun lipidomics approach. Anal. Chem. 83, 4243-4250. 

Yang, K., Dilthey, B.G. and Gross, R.W. (2013) A shotgun lipidomics approach using charge switch derivatization Analysis of fatty acid double bond isomers. J. Am. Soc. Mass Spectrom. 24(S1), 228. 

Many lipid classes can be quantified by this improved shotgun lipidomics approach. Quantification of individual species of a lipid class is conducted based on the summed abundance of its major fragment(s) in comparison to the counterpart of the spiked internal standard of the class. The effects of different isotopologue... 

Note that this shotgun lipidomics approach is based on tandem MS analysis. As discussed earlier, due to the differential kinetics and/or thermodynamics of different species of a class during CID, two or more internal standards of a class spiked during lipid extraction are necessary to minimize any effects of differential fragmentation patterns on quantification. This point is particularly important to those species containing polyunsaturated fatty acyl substituents [20]. Moreover, identification and quantification of low-abundant isomeric/isobaric species overlapped with an abundant species may be missed due to the limitation of a relatively narrow dynamic range. 

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