Ion chromatograms

Computers, often combined with transputers, are used for three main functions when connected to a mass spectrometer. The foremost requirements involve the acquisition and preprocessing of basic data and the control of the instrument s scanning operations. Additional software programs are available to manipulate the preprocessed data in a wide variety of ways depending on what is required, e.g., a mass spectrum or a total ion chromatogram. 

Ion-beam lithography Ion beam mixing Ion beam processing Ion beams Ion-beam sputtering Ion channels Ion chromatogram Ion chromatography... 

Fig. 4. Chromatograms of Russian coriander oil injected into a serially configured gc /ir /ms ( a) total ion chromatogram (TIC) (b) total response...

Figure 15.10 Primary (a) and secondary (b) separation of unleaded gasoline, where (a) shows the IRD chromatogram, and (b) shows the MSD total ion chromatogram of heart cut c. Adapted from Analytical Chemistry, 65, N. Ragunathan et al., Multidimensional gas chromatography with parallel cryogenic tr aps , pp. 1012-1016, copyright 1993, with permission from the American Chemical Society.

Ions at m/z 55, 60, 214 and 236 are observed but do some or all of these arise from the background and are present throughout the analysis, or are they present in only a few scans, i.e. are they from a component with insufficient overall intensity to appear as a discrete peak in the TIC trace An examination of reconstructed ion chromatograms (RICs) from these ions generated by the data system may enable the analyst to resolve this dilemma. The TIC shows the variation, with time, of the total number of ions being detected by the mass spectrometer, while an RIC shows the variation, with time, of a single ion with a chosen m/z value. The RICs for the four ions noted above are shown in Figure 3.15. These ions have similar profiles and show a reduction in intensity as analytes elute from the column. The reduction in intensity is a suppression effect. 

Figure 3.15 Reconstructed ion chromatograms obtained for ions in the background mass spectrum from the LC-MS analysis of a pesticide mixture From applications literature published by Micromass UK Ltd, Manchester, UK, and reproduced with permission.

Why is the sensitivity obtained when using reconstructed ion chromatograms (RICs) for quantitation less than that achieved when employing selected-ion monitoring (SIM) to monitor the same ions ... 

Figure 5.10 Reconstructed ion chromatograms used to quantify the levels of the principal oligosaccharides found in rituximab process samples. Reprinted from J. Chromatogr., A, 913, Wan, H. Z., Kaneshiro, S., Frenz, J. and Cacia, J., Rapid method for monitoring galactosylation levels during recombinant antibody production by electrospray mass spectrometry with selective-ion monitoring , 437-446, Copyright (2001), with permission from Elsevier Science.

The results of this analysis are shown in Figure 5.16, which contains the UV trace, at 220 nm, and the equivalent mass spectral trace. In this case, the mass spectral data are represented by a modification of a reconstructed ion chromatogram (RIC) known as the base-peak trace, i.e. a plot of the intensity of the base peak in each scan (the m jz value of this may well vary from scan to scan) as a function of time. 

What is a reconstructed ion chromatogram (RiC) and how does it aiiow a specific compound to be iocated in the totai-ion current (TiC) trace from an LC-MS anaiysis ... 

Figure 5.28 LC-electrospray-MS total ion chromatogram of sulfated oligosaccharides from mucins purified from the porcine large intestine, where the annotations indicate the molecular ions observed from each component. Reprinted with permission from Thoms-son, K. A., Karlsson, H. and Hansson, G. C., Anal. Chem., 72, 4543-4549 (2000). Copyright (2000) American Chemical Society.

While there is a vast range of different drug structures, there are only a relatively small number of chemical reactions, some of which are shown below in Table 5.13 (p. 199), involved in the production of metabolites. Based on the structure of the drug, it is therefore possible to predict the most likely metabolites. Use may then be made of reconstructed ion chromatograms (RlCs) of mlz values corresponding to the predicted molecular weights of these metabolites to locate them within the LC-MS data obtained. 

Figure 5.41 The total-ion-current (TIC) trace and reconstructed ion chromatograms from the predicted pseudomolecular ions of Indinavir m/z 614) and its mono- (m/z 630) and dihydroxy metabolites (m/z 646), generated from full-scan LC-MS analysis of an incubation of Indinavir with rat liver S9. Reprinted by permission of Elsevier Science from Identification of in vitro metabolites of Indinavir by Intelligent Automated LC-MS/MS (INTAMS) utilizing triple-quadrupole tandem mass spectrometry , by Yu, X., Cui, D. and Davis, M. R., Journal of the American Society for Mass Spectrometry, Vol. 10, pp. 175-183, Copyright 1999 by the American Society for Mass Spectrometry.

One approach to drug metabolism studies is therefore to predict the molecular weights of possible metabolites of the drug under consideration, to use reconstructed ion chromatograms to locate any components that have the appropriate molecular weights and then use MS-MS to effect fragmentation of the (M - - H)+ ions from these metabolites, and then to finally link the m jz values of the ions observed with ions of known structure from the parent drug or from other metabolites whose structures have been elucidated. 

The general procednre is to nse reconstrncted ion chromatograms at appropriate m/z values in an attempt to locate componnds of interest and then look at the mass spectrum of the unknown to determine its molecnlar weight. MS-MS can then be employed to obtain spectra from this and related compounds to find ions that are common to both and which may therefore contain common stmctmal features. Having the same m/z value does not necessarily mean the ions are identical and further MS-MS data or the elemental composition may be required. If these data do not allow unequivocal structure identification, then further MS" information may be required. 

Figure 5.57 Reconstructed ion chromatograms from the negative-ion LC-MS-MS analysis of (A) G-fenozide, (B) hydroxyfenozide, and (C) methoxyfenozide (for structures,...

Figure 5.58 Reconstructed LC-MS-MS ion chromatograms for selected-reaction monitoring of methoxyfenozide using the m/z 367 to m/z 149 transition from the continual post-column infusion of a standard solution of analyte during the HPLC analysis of a...

Figure 5.67 Reconstructed ion chromatograms for Idoxifene and internal standard (ds-Idoxifene using LC-ToF-MS for (a) double-blank human plasma extract, (b) extract of blank human plasma containing internal standard (IS), and (c) control-blank human plasma spiked with Idoxifene at 5 gml , the LOQ of the method. Reprinted from 7. Chromatogr., B, 757, Comparison between liquid chromatography-time-of-flight mass spectrometry and selected-reaction monitoring liquid chromatography-mass spectrometry for quantitative determination of Idoxifene in human plasma , Zhang, H. and Henion, J., 151-159, Copyright (2001), with permission from Elsevier Science.

A reconstructed ion chromatogram is a plot showing the variation in intensity of an ion of a particular m/z ratio as a function of analysis time, while the total-ion-current trace shows the variation in the intensity of all ions being produced as a function of analysis time. Simplistically, the TIC will show an increase as a compound elutes from an HPLC column and is ionized. If an ion with a particular m/z value is found to be diagnostic of a compound or series of compounds of interest, then an RIC of this m/z will show where its intensity increases and, therefore, where a compound of interest may have eluted. The mass spectrum at this point can then be examined for further confirmation that it is of significance. 

Reconstructed ion chromatogram A plot of the intensity of an ion of chosen m/z ratio as a fnnction of analysis time. This is produced by computer analysis of mass spectral data acqnired over an extended mass range. 

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