Thin Layer Chromatography-Mass Spectrometry Techniques

3 THIN LAYER CHROMATOGRAPHY-MASS SPECTROMETRY TECHNIQUES 

Acceptable agreement between the migration distance of standards and sample components, and application of specific derivatizing reagents are commonly used 

Of more immediate interest are approaches that permit offline TLC-MS in which the spots are scraped out from the layer and the analytes are either extracted from the sorbent to be transferred to the mass spectrometer as discrete samples or are introduced without sorbent removal into the spectrometer on a direct insertion probe (51). TLC-MS quantification and confirmation efficiency can be further enhanced by submitting the TLC extract to an additional chromatographic separation using a different technique prior to the final MS analysis. Advantages of this approach over direct TLC-MS include extra cleanup through the additional chromatographic separation (52). This has been realized in the TLC-GC-MS analysis of eggs and meat for chloramphenicol residues (49). 

A simpler means for unequivocal identification of substances by TLC or HPTLC combined with MS, can be provided by online procedures (54, 55). The ability to perform MS on analytes directly on the chromatographic plate removes the need to recover them prior to identification. This greatly reduces the amount of work needed to confirm identity. The determination of midazolam in serum (56) and the identification of tetracycline residues in honey (57) are examples of TLC coupled in situ with FAB-MS. 

However, tandem mass spectrometry, as a separation technique, does have limitations. It cannot easily differentiate between isomeric and isobaric species, and, in complex matrices, the presence of components with a high surface activity can suppress the ionization of components with a lower surface activity, leading to the nondetection of analytes (66). Therefore, the combination of MS-MS with a readily available chromatographic separation method such as TLC affords analysts real benefits. 

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PCB materials in field-collected bald eagle specimens have been identified utilizing combined gas chromatography-mass spectrometry techniques in conjunction with thin-layer chromatographic separations of standard materials as well as tissue extracts (26). A total of 19 PCB compounds were detected in the field samples. Analyses were performed employing a spiral glass gas chromatographic column, 9 ft X 0.25 inch o.d., packed with 1% SE-30 on 100/120 mesh Gas Chrom Q. Mass spectra of the major components of Aroclor 1254 PCB standard were presented in this report as well as relative retention data for the individual components. 

See also-. Extraction Supercritical Fluid Extraction. Forensic Sciences Gunshot Residues Thin-Layer Chromatography. Fourier Transform Techniques. Gas Chromatography Mass Spectrometry Forensic Applications. Microscopy Appiications Forensic. X-Ray Absorption and Diffraction X-Ray Absorption X-Ray Diffraction - Powder. 

The on-line principle has also been extended into the field of detection (Fig. 8). Thus, it is now possible to record FTIR [27-31] and Raman spectra in situ [32, 33], and there have been considerable advances in the on-line coupling of thin-layer chromatography with mass spectrometry. Here it has been, above all, the research groups of Wilson [34-36] and Busch [37-40] that have made the necessary instrumental and methodological advances, so that TLC must no longer be viewed as merely a clean-up method. Rather it forms the essential central point for all these on-line coupling techniques. 

All previous discussion has focused on sample preparation, i.e., removal of the targeted analyte(s) from the sample matrix, isolation of the analyte(s) from other co-extracted, undesirable sample components, and transfer of the analytes into a solvent suitable for final analysis. Over the years, numerous types of analytical instruments have been employed for this final analysis step as noted in the preceding text and Tables 3 and 4. Overall, GC and LC are the most often used analytical techniques, and modern GC and LC instrumentation coupled with mass spectrometry (MS) and tandem mass spectrometry (MS/MS) detection systems are currently the analytical techniques of choice. Methods relying on spectrophotometric detection and thin-layer chromatography (TLC) are now rarely employed, except perhaps for qualitative purposes. 

The conventional approach to solvent extraction is the batch method. Early work with this method was hampered by the low concentration of the compounds present and the relative insensitivity of the methods of characterization. Thus lipids and hydrocarbons have been separated from seawater by extraction with petroleum ether and ethyl acetate. The fractionation techniques include column and thin-layer chromatography with final characterisation by thin-layer chromatography, infrared, and ultra-violet spectroscopy and gas chromatography. Of these techniques, only gas chromatography is really useful at levels of organic matter present in seawater. With techniques available today such as glass capillary gas chromatography and mass spectrometry, much more information could be extracted from such samples [20]. 

The laboratory devised a special vacuum pump and filters to take samples from various sections of the garage. The laboratory was able to confirm the presence of heroin, using rigorous isolation techniques combined with fluor-imetry, thin-layer chromatography, and mass spectrometry. The laboratory s work played a major role in the successful prosecution of this case. 

FSIS laboratories also use chemical techniques and instrumentation to identify select antibiotic residues. The tetracyclines of interest are identified by thin layer chromatography. Sulfonamides are detected and quantified by fluorescence thin lay chromatography and confirmed by gas chromatography/mass spectrometry. Amoxicillin and gentamycin are identified and/or quantified by high pressure liquid chromatography. Similar techniques are used to identify ionophores and other antimicrobials of interest. 

Sulfatide in urine can be quantified by high-performance liquid chromatography (HPLC) [39] and tandem mass spectrometry techniques [63]. Here, a simple semi-quantitative thin-layer chromatography (TLC) method is described, as modified from Rail et al. [46]. 

H Oka, Y Ikai, F Kondo, N Kawamura, J Hayakawa, K Masuda, KI Harada, M Suzuki. Development of a condensation technique for thin-layer chromatography fast-atom bombardment mass spectrometry of non-visible compounds. Rapid Commun Mass Spectrom 6 89-94, 1992. 

Other techniques that have been used to determine polycyclic aromatic hydrocarbons in soil extracts include ELISA field screening [86], micellar elec-tr okinetic capillary chromatography [ 87], supersonic jet laser-induced fluorescence [88,89], fluorescence quenching [90], thermal desorption gas chromatography-mass spectrometry [81,90,100], microwave-assisted extraction [91], thermal desorption [92], immunochemical methods [93,94], electrophoresis [96], thin layer chromatography [95], and pyrolysis gas chromatography [35]. 

Thompson et al. [113] emphasise that even the use of two dissimilar gas chromatographic columns does not ensure irrefutable compound identification. For example, if the retention characteristics of a given peak obtained from two dissimilar columns suggest the possibility of the presence of a compound which appears wholly out of place in a specific sample, further confirmation is clearly indicated by such techniques as specific detectors, coulometry, p values, or gas chromatography-mass spectrometry or thin layer chromatography. 

Chapter 14 discusses the very useful technique of thin layer chromatography. It has extensive applications in the analysis of complex mixtures of organic compounds and also has found limited applications in the analysis of organometallic compounds. No applications to anions and cations have been reported to date. The technique can also be used to prepare extracts suitable for subsequent examination by infrared spectroscopy or mass spectrometry. 

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