Interface continuous flow FAB

El may be used with the moving-belt and particle-beam interfaces. Cl with the moving-belt, particle-beam and direct-liquid-introduction interfaces, and FAB with the continuous-flow FAB interface. A brief description of these ionization methods will be provided here but for further details the book by Ashcroft [8] is recommended. 

Two forms of interface have been commercially developed [7] which allow analytes in a flowing liquid stream - it has to be pointed out, not necessarily from an HPLC system (see below) - to be ionized by using FAB. These are essentially identical except for that part where the HPLC eluate is bombarded with the heavy-atom/ion beam. Both of these interfaces consist of a probe in the centre of which is a capillary which takes the flowing HPLC eluate. In the continuous-flow FAB interface (Figure 4.3), the column eluate emerges from the end of the capillary and spreads over the probe tip, while in the frit-FAB interface the capillary terminates in a porous frit onto which the atom/ion beam is directed. 

The development of the continuous-flow FAB interface by Caprioli et al. [8] was primarily directed at the analysis of peptides (Ch. 4.6). Continuous-flow FAB provided reduced suppression of hydrophilic peptides in peptide mixtures. It was widely applied in the field of peptide characterization and the analysis of proteolytic digests, e.g., in the analysis of a tryptic digest of bovine ribonuclease B before and after treatment with N-glycanase [9]. A single injection of 100 pmol provided ca. 70% sequence coverage. 

Therefore three principal strategies for handling the effluents of the LC-columns were under research 1. Removal of solvent by vaporization and subsequent ionisation of the analytes first led to the moving-belt interface which, later, was followed by the development of particle beam ionisation. 2. Direct ionisation was the basic principle of the continuous flow-FAB interface, whereas 3. nebuliza-tion of the column effluent was the basic principle of DLI, TSP, APCI or ESI ionisation [1]. 

In 1981, Barber and co-workers at the University of Manchester Institute of Science and Technology introduced fast atom bombardment (FAB), which produced a continuous source of ions from nonvolatile molecules dissolved in a liquid matrix. Such sources were easily retrofitted to existing sector and quadrupole mass spectrometers, and they profoundly improved capabilities for the structural analysis of peptides, carbohydrates, and other biological molecules. In addition, a continuous flow FAB interface was later introduced that enabled online interfacing with hi ih-performance liquid chromatography (HPLC). Thus, this powerful, new ionization technique continued to reinforce the preeminence of sector and quadrupole mass spectrometers, and did little to advance either the technology or the popularity of time-of-flight instruments. 

Dynamic/continuous-flow FAB allows a continuous stream of liquid into the FAB source hence it constitutes an LC/MS interface for analyses of peptide mixtures. 

Figure 4.3 Schematic of a continuous-flow FAB LC-MS interface. From applications literature published by Kratos Analytical Ltd, Manchester, UK, and reproduced by permission of Mass Spectrometry International Ltd.

Many interfaces have been developed to meet these demanding challenges. Some of these coupling methods, such as the moving belt or the particle beam interface, are based on the concomitant elimination of the solvent before it enters the mass spectrometer. Other methods such as direct liquid introduction (DLI) or continuous flow FAB rely on splitting the flow of the liquid that is introduced into the interface in order to obtain a flow that can be directly infused into the ionization source. However, these types of interfaces can only handle a fraction of the liquid flow from the LC. 

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. 

Since APCI and ESI interfaces operate at atmospheric pressure and do not depend upon vacuum pumps to remove solvent vapor, they are compatible with a wide range of HPLC flow rates. HPLC methods that have been developed using conventional detectors such as UV/VIS, IR, or fluorescence are usually transferable to LC/MS systems without adjustment. However, the solvent system should contain only volatile solvents, buffers, or ion-pair agents to reduce fouling of the mass spectrometer ion source. In the case of chlorophyll solvent systems, isocratic and gradient combinations of methanol, acetonitrile, water, acetone, and/or ethyl acetate have been used for APCI or ESI LC/MS. Unlike continuous-flow FAB/LSIMS, no sample matrix is necessary. 

During the 1990s, electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) became the standard interfaces for LC/MS. Unlike thermospray, particle beam, or continuous-flow FAB, ESI and APCI inter-... 

Different methods are used to tackle these problems [10-13], Some of these coupling methods, such as moving-belt coupling or the particle beam (PB) interface, are based on the selective vaporization of the elution solvent before it enters the spectrometer source. Other methods such as direct liquid introduction (DLI) [14] or continuous flow FAB (CF-FAB) rely on reducing the flow of the liquid that is introduced into the interface in order to obtain a flow that can be directly pumped into the source. In order to achieve this it must be reduced to one-twentieth of the value calculated above, that is 5 pi min. These flows are obtained from HPLC capillary columns or from a flow split at the outlet of classical HPLC columns. Finally, a series of HPLC/MS coupling methods such as thermospray (TSP), electrospray (ESI), atmospheric pressure chemical ionization (APCI) and atmospheric pressure photoionization (APPI) can tolerate flow rates of about 1 ml min 1 without requiring a flow split. Introducing the eluent entirely into the interface increases the detection sensitivity of these methods. ESI can accept flow rates from 10 nl min-1 levels to... 

Like continuous-flow FAB, the popularity of particle beam interfaces is diminishing, but systems are still available from commercial sources. During particle beam LC-MS, the HPLC eluate is sprayed into a heated chamber... 

Note MB moving belt DLL direct liquid introduction ISP ion spray HNI heated nebuliser interface cfFAB continuous-flow FAB RP reversed phase NP normal phase IE ion evaporation API atmospheric pressure ionization RV relatively volatile RIV relatively involatile IV involatile. The rest of the abbreviations as in Fig. 1. 

However, the great improvement to solve the possibility of coupling HPLC to MS was the development of interfaces that allow the elimination of the solvent and the vaporization of the solute before this can be analyzed in the mass spectrometer. Several interfaces are now available, such as continuous flow FAB (CF-FAB), thermospray (TSP), moving belt (MB), direct liquid introduction (LDI), and atmospheric pressure ionization (API) in the field of API, three different, but fundamentally similar, techniques are available ion evaporation, electrospray, and ion spray. In each case, the removal of... 

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