In-source fragmentation

The presence of in-source fragmentation could have the following consequences  

Accordingly, elimination of the in-source fragmentation process should be a task of a researcher who develops a method in lipidomics. 

Here are some tips that should aid in elimination of this process. A standard solution of lipids should be further used to fine-tune the instrument after an instrument is tuned with a standard solution as recommended by the instrumental vendor(s) since those standard solutions are generally in favor of proteomics. This measure alone could enhance the ionization efficiency by an order of magnitude in many cases in the experiences of author s laboratory. Alternatively, a less harsh ionization condition (temperature, voltage, etc.) can be purposely set when one begins a tune/calibration experiment. 

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The ionization techniques most widely used for LC-MS, however, are termed soft ionization in that they produce primarily molecular species with little fragmentation. It is unlikely that the molecular weight alone will allow a structural assignment to be made and it is therefore desirable to be able to generate structural information from such techniques. There are two ways in which this may be done, one of which, the so-called cone-voltage or in-source fragmentation, is associated specifically with the ionization techniques of electiospray and APCl and is discussed later in Section 4.7.4. The other, termed mass spectrometry-mass spectrometry (MS-MS) or tandem mass spectrometry, is applicable to all forms of ionization, provided that appropriate hardware is available, and is described here. 

Van Breemen, R.B. et al.. Liquid chromatography/mass spectrometry of carotenoids using atmospheric pressure chemical ionization, J. Mass Spectrom., 31, 975, 1996. Tian, Q., Duncan C.J.G., and Schwartz S. J., Atmospheric pressure chemical ionization mass spectrometry and in-source fragmentation of lutein esters, J. Mass Spectrom., 38, 990, 2003. 

Unfortunately, this technique is not selective and all components are affected in the same way so that if exact co-elution of sample components occurs, this may not be detected. Using a single quadrupole instrument limits the user to using in-source fragmentation, but in many cases this can provide enough information to identify unknowns. 

Fragmentation patterns of standards can help to determine whether observed peaks are the result of in-source fragmentation or natural occurrence in the sample. Flavonol glycoside standards are available from Extrasynthese and Indo Fine Chemical. 

TABLE 9.1. A Summary of in-Source Fragmentation and Diagnostic MS/MS Fragments Generated Using a TSQ Quantum Mass Spectrometer... 

For a compound which contains dimethylpyperidine substructure (shown on the left), isomeric metabolites oxygenated at positions 1, 2, and 3 can be easily distinguished without NMR by conducting a single LC-APCI-MS experiment on the TSQ Quantum. Loss of 16 is observed for oxidation at position 1 (/V-oxide), loss of 18 (H20) for aliphatic hydroxylation at position 3, while for aromatic hydroxylation at position 2 no significant in-source fragmentation is observed. 

Dihydroxacephenone (DHAP) Mixed with diammonium hydrogen citrate to limit cation adduction, offers improved ionization of sialylated glycopeptides with limited in-source fragmentation compared to DHB and HCCA36... 

Gangoliside in-source fragmentation can be reduced but not eradicated, by doping the matrix with large cations such as cesium, by permethylation,191 conversion of the carboxyl group into a methyl ester,195 and the use of a novel ionic-liquid matrix which has soft desorption capabilities.144... 

ESI, like MALDI is compatible with negative- and positive-mode analyses and for analysis of native or permethylated neutral glycosphingolipids. The most abundant signal is also often the singly sodiated ion [M + Na]+. In-source fragmentation can be tuned by varying the ion source temperature and skimmer voltage. 

Mazsaroff, I. Yu, W. Kelley, B. D. Vath, J. E. 1997. Quantitative comparison of global carbohydrate structures of glycoproteins using LC-MS and in-source fragmentation. Anal Chem., 69,2517-2524. 

Xian, Q. Duncan, C.J.G. Schwartz, S.J. 2003. Atmospheric pressure chemical ionization mass spectrometry and in-source fragmentation of lutein esters. J. Mass Spectrom. 38 990-995. 

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