Mass spectra, direct comparison

FTICR-MS is capable of powerful mixture analysis, due to its high mass range and ultrahigh mass resolving power. However, in many cases it is still desirable to couple a chromatographic interface to the mass spectrometer for sample purification, preconcentration, and mixture separation. In the example given above, DTMS under HRMS conditions provides the elementary composition. Apart from DTMS, PyGC-MS can be performed to preseparate the mixture of molecules and to obtain the MS spectrum of a purified unknown. Direct comparison with the pure reference compound remains the best approach to obtain final proof. 

The mass spectrum of 3-citroneIlyl-2,5-dimethylpyrazine (201) exhibits a parent peak at m/z 246 and a normal base peak (484a) at m/z 122, indicative of a dimethylpyrazine having an unsaturated Cjq alkyl chain without branching at the a-methylene group. The pyrazine is identified by direct comparison with the compounds synthesized on the assumption that the alkyl chain is isoprenoid 142). 

Aristolochic acid I (143), isolated from whole bodies of swallowtail butterflies of the genus Pachlioptera, is the already known substance and was identified by the mass spectrum (75). One of the two acidic components obtained from ap-osematic butterflies of the genus Zerynthia has been identified as aristolochic acid C (145) by direct comparison with an authentic sample. The other, first isolated from this butterfly, has been identified as aristolochic acid la (144) by mass spectral analysis and its derivation to aristolochic acid I methyl ester (146) (74). 

The utility of UPLC has been demonstrated for both qualitative and quantitative analyses. In 2005, Castro-Perez et al. (2005) compared the performance of a HPLC with that of a UPLC and showed that improved chromatographic resolution and peak capacity attained with UPLC lead to reduction in ion suppression and increased MS sensitivity. Comparison of the mass spectrum obtained using HPLC with that from UPLC (Fig. 1.15) revealed that the higher resolving power of the UPLC-MS system resulted in a much cleaner mass spectrum than that obtained using the HPLC-MS system. The sensitivity improvement directly resulted in a higher ion count in the UPLC mass spectrum (855 vs. 176). 

Hydroxycanthin-6-one (16) has been isolated from the root bark of Simarouba amara (34) and from cell cultures of A. altissima 12,15). Since its UV spectrum was similar to that of 5-methoxycanthin-6-one (25), Lassak et al. (34) assumed it to be 4- or 5-hydroxycanthin-6-one. The prominent peak at M -56 (mlz 180, M -2CO) in the mass spectrum supported structure 16 having an OH adjacent to the carbonyl group. The final confirmation of structure 16 was made through direct comparison of methylated 16 with authentic 7. 

The structural assignment of the above cytochalasins was based primarily on their characteristic spectral data and direct comparison with previously known compounds. The mass spectrum of all metabolites showed a tropylium ion as the base peak (91 m/z) which is characteristic of 10-phenylcytochalasins and the molecular formulae of the six new compounds were established via high resolution MS and elemental analyses. Extensive H and 3C NMR analysis using COSY, HETCOR and DEPT NMR experiments, was also carried out in order to establish the reported structures. 

This approach has been used to quantify individual species of over 40 lipid classes directly from lipid extracts of biological samples with or without derivatization [16, 18, 29, 37]. For example, in the analysis of SM species present in lipid extracts of mouse cortices, which were treated with lithium methoxide as previously described [38], the quantities of lithiated N18 l SM at miz 735.5, N18 0 SM at m/z 737.5, and N24 l SM at m/z 819.6 were accurately measured with ratiometric comparisons to the selected internal standard (i.e., N14 0 SM at m/z 653.5) after de-isotoping of the mass spectrum acquired in the full MS mode (Trace a of Figure 14.2). The contents of all other low-abundant SM species were determined with the second step of quantitation utilizing the mass spectrum of NLS 183 (Trace b of Figure 14.2) or NLS213 (as previously demonstrated [20]) with N14 0, N18 l, and N24 l SM as internal standards. 

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