Mass spectrometry interfacing with HPLC

Instrumental developments facilitate the miniaturization opportunity. Advances in chromatography led to the use of capillary HPLC techniques (Liu et al., 1993 Kassel et al., 1994 Arnott et al., 1995) for the powerful separations of increasingly smaller samples. For mass spectrometry, developments with the LC/MS interface that resulted in increased ion transmission and ion sampling helped to... 

Frequently industrial hygiene analyses require the identification of unknown sample components. One of the most widely employed methods for this purpose is coupled gas chromatography/ mass spectrometry (GC/MS). With respect to interface with mass spectrometry, HPLC presently suffers a disadvantage in comparison to GC because instrumentation for routine application of HPLC/MS techniques is not available in many analytical chemistry laboratories (3). It is, however, anticipated that HPLC/MS systems will be more readily available in the future ( 5, 6, 1, 8). HPLC will then become an even more powerful analytical tool for use in occupational health chemistry. It is also important to note that conventional HPLC is presently adaptable to effective compound identification procedures other than direct mass spectrometry interface. These include relatively simple procedures for the recovery of sample components from column eluate as well as stop-flow techniques. Following recovery, a separated sample component may be subjected to, for example, direct probe mass spectrometry infra-red (IR), ultraviolet (UV), and visible spectrophotometry and fluorescence spectroscopy. The stopped flow technique may be used to obtain a fluorescence or a UV absorbance spectrum of a particular component as it elutes from the column. Such spectra can frequently be used to determine specific properties of the component for assistance in compound identification (9). 

From the viewpoint of trace analysis by mass spectrometry interfaced online with HPLC, flow-FAB interfaces... 

Reference has been made to the problems associated with the presence of highly involatile analytes. Many buffers used in HPLC are inorganic and thus involatile and these tend to compromise the use of the interface, in particular with respect to snagging of the belt in the tunnel seals. The problem of inorganic buffers is not one confined to the moving-belt interface and, unless post-column extraction is to be used, those developing HPLC methods for use with mass spectrometry are advised to utilize relatively volatile buffers, such as ammonium acetate, if at all possible. 

The method for chloroacetanilide soil metabolites in water determines concentrations of ethanesulfonic acid (ESA) and oxanilic acid (OXA) metabolites of alachlor, acetochlor, and metolachlor in surface water and groundwater samples by direct aqueous injection LC/MS/MS. After injection, compounds are separated by reversed-phase HPLC and introduced into the mass spectrometer with a TurboIonSpray atmospheric pressure ionization (API) interface. Using direct aqueous injection without prior SPE and/or concentration minimizes losses and greatly simplifies the analytical procedure. Standard addition experiments can be used to check for matrix effects. With multiple-reaction monitoring in the negative electrospray ionization mode, LC/MS/MS provides superior specificity and sensitivity compared with conventional liquid chromatography/mass spectrometry (LC/MS) or liquid chromatography/ultraviolet detection (LC/UV), and the need for a confirmatory method is eliminated. In summary,... 

The mass spectrometer is mainly used as a mass detector in chromatography (GC, SFC, HPLC, SEC, TLC). With the great variety of interfaces, ionisation modes and mass spectrometers, chromatography-mass spectrometry is highly diversified. High-resolution separations combined with accurate mass measurements and element-sensitive detection (MIP, ICP) have been reported. 

Perhaps the most mechanically complex solution ever developed for uniting HPLC with mass spectrometry was the moving belt interface [54]. The heart of this system was a mechanically driven continuous belt (analogous to an escalator or moving walkway) to which the HPLC eluent was applied. The majority of the mobile phase was evaporated by a heat source (ideally hot enough to vaporize the solvents but not to... 

Figure 14 Interfacing Liquid Chromatography with Electrospray Ionization Mass Spectrometry HPLC...

Interfacing HPLC or HPCE (capillary electrophoresis) to mass spectrometry is technically more complex than. with a GC because these techniques use a solvent that is often aqueous (water is a poison to mass spectrometers). The use of microcolumns in HPLC is desirable for coupling to MS because micro-columns operate at very low flow rates. They are also compatible with different ionisation techniques for the analysis of high molecular-weight species. 

FAB and LSIMS are matrix-mediated desorption techniques that use energetic particle bombardment to simultaneously ionize samples like carotenoids and transfer them to the gas phase for mass spectrometric analysis. Molecular ions and/or protonated molecules are usually abundant and fragmentation is minimal. Tandem mass spectrometry with collision-induced dissociation (CID) may be used to produce abundant structurally significant fragment ions from molecular ion precursors (formed using FAB or any suitable ionization technique) for additional characterization and identification of chlorophylls and their derivatives. Continuous-flow FAB/LSIMS may be interfaced to an HPLC system for high-throughput flow-injection analysis or on-line LC/MS. 

The use of liquid chromatography-mass spectrometry (LC-MS) is becoming more popular because of the increasing number of LC-MS interfaces commercially available thermospray (TSP), particle beam (PB), and atmospheric pressure ionization (API). Coupled with mass spectroscopy, HPLC provides the analyst with a powerful tool for residue determination. 

In thermospray interfaces, the column effluent is rapidly heated in a narrow bore capillary to allow partial evaporation of the solvent. Ionisation occurs by ion-evaporation or solvent-mediated chemical ionisation initiated by electrons from a heated filament or discharge electrode. In the particle beam interface the column effluent is pneumatically nebulised in an atmospheric pressure desolvation chamber this is connected to a momentum separator where the analyte is transferred to the MS ion source and solvent molecules are pumped away. Magi and Ianni (1998) used LC-MS with a particle beam interface for the determination of tributyl tin in the marine environment. Florencio et al. (1997) compared a wide range of mass spectrometry techniques including ICP-MS for the identification of arsenic species in estuarine waters. Applications of HPLC-MS for speciation studies are given in Table 4.3. 

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