Particle beam interface HPLC

Reverse-phase (RP)-HPLC is probably the best system for purifying triterpenoids, principally when mixtures of isomers are present [35]. Gunther and Wagner in 1996 [36] carried out the separation and quantification of active triterpenes from Centella asiatica employing an RP system with acetonitrile-water as mobile phase. Recently, Gaspar et al. [37] described the complete separation of a mixture of triterpenoid isomers from the fruit of Arbutus unedo by HPLC coupled to a mass spectrophotometer by means of a particle beam interface (HPLC-PBMS). The separation of different quassinoids from crude bark of Quassia amara was developed by Vitanyi et al. [38] using a reverse-phase HPLC-MS... 

The nebulization and evaporation processes used for the particle-beam interface have closely similar parallels with atmospheric-pressure ionization (API), thermospray (TS), plasmaspray (PS), and electrospray (ES) combined inlet/ionization systems (see Chapters 8, 9, and 11). In all of these systems, a stream of liquid, usually but not necessarily from an HPLC column, is first nebulized... 

The particle-beam interface provides El spectra from HPLC eluates and this is of great advantage over other interfaces which provide only molecular weight information. Why then, is it of advantage to be able to generate Cl spectra from the particle-beam interface ... 

The particle-beam interface has been developed primarily to provide El spectra from HPLC eluates but may be combined with other ionization techniques such as CL If quantitative studies are being undertaken, a detailed study of experimental conditions should be undertaken. Isotope-dilution methodology is advocated for the most accurate results. 

The performance of the particle-beam interface deteriorates as the percentage of water in the HPLC mobile phase increases. 

Only the particle-beam interface produces El spectra for direct comparisons with computerized library spectra of fragmentation patterns. The other systems enable the relative molecular mass (RMM) of analytes up to 105 and above to be established. An example of an HPLC-APCI separation and identification of some benzodiazepine tranquillizers is shown in Figure 4.39. The most appropriate choice of LC-MS interface for a particular... 

The particle beam interface [55] borrowed and built upon some of the key elements and concepts of its predecessors. Eluent from the HPLC was nebulized into a spray of small droplets by a flow of helium. The spray of droplets entered a heated chamber where evaporative processes further reduced the droplet size creating an aerosol. The next step of the process involved the spraying of the aerosol (i.e., the... 

Figure 16.14—HPLC/MS interface using a particle beam interface. The angular diffusion of heavy molecules under vacuum is narrower that that of smaller molecules of solvent, thus they have a higher probability of being transported to the mass spectrometer along the central axis of the system.

The particle beam interface is an example of an HPLC/MS interface (see Fig. 16.14). In this interface, the mobile phase is vaporised in the form of a spray into a desolvation chamber before entering the area where the sample is concentrated by evaporation. Because solvent molecules are lighter, their angular dispersion is wider than that of the analyte, which can be transferred into the transfer capillary. This type of interface is slowly being abandoned in favour of others that have a greater sensitivity. 

Urea pesticides are structurally similar to carbamates. Some common pesticides of this class are listed in Table 2.19.2. These substances can be determined by reverse phase HPLC method. Aqueous samples can be analyzed by U.S. EPA Method 553 using a reverse phase HPLC column interfaced to a mass spectrometer with a particle beam interface. The outline of the method is described below. 

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. 

Earlier methods of ionization applied to carotenoids, including electron impact (El), chemical ionization (Cl), a particle beam interface with El or Cl, and continuous-flow fast atom bombardment (CF-FAB), have been comprehensively reviewed elsewhere (van Breemen, 1996, 1997 Pajkovic and van Breemen, 2005). These techniques have generally been replaced by softer ionization techniques like electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI), and more recently atmospheric pressure photoionization (APPI). It should be noted that ESI, APCI, and APPI can be used as ionization methods with a direct infusion of an analyte in solution (i.e. not interfaced with an HPLC system), or as the interface between the HPEC and the MS. In contrast, matrix-assisted laser desorption ionization (MALDI) cannot be used directly with HPEC. 

The main components of an LC-MS are the HPLC apparatus, an optional UV or photodiode array detector, the interface, the mass spectrometer and a computer system for data management and evaluation. The interface is the key component of the LC-MS system. All other components must be adapted to the particular interface that is used. Most commercially available systems work with thermospray, electrospray, or particle beam interfaces. Each interface has a distinct mode of action and its own operational parameters. 

The particle beam interface is a system in which the solvent of the mobile phase is evaporated from the sample using reduced pressure and an elevated temperature. This requirement places some restrictions on the HPLC-part of the system. The working range, for the flow rate of the HPLC, is specified (19) at O.l-l.OmL/min. 

Chromatographic columns with an inner diameter of 2 mm are frequently used for LC-MS applications. We have found that columns with a 3 mm inner diameter are more robust and ideally suited for use with a particle beam interface. Flow rates around 0.5mL/min can be used with conventional HPLC systems as well as LC-MS systems having a particle beam interface. Using this size column and flow rate, no adaptation of the separation method is necessary. 

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... 

These efforts are centered around the use of techniques including enzymatic hydrolysis [5,6,7], physical or chemical degradation [8,9], and monodisperse aerosol generation interface (particle beam) for HPLC/MS [10,11] to solve specific problems. This paper discusses the implementation of several HPLC/MS methods which offer a combination of sensitivity and specificity for compounds such as peptides, pharmaceuticals, and pesticides in complex matrices. 

Nowadays, interfacing of a liquid chromatograph or a capillary electrophoresis instrument with a mass spectrometer is used too, although technically more complex. The presence of water in elution solvents is an undesirable compound for the mass spectrometer. With HPLC, the use of micro-columns is desirable, to have very low flow rates. They are also well-suited with different ionization techniques for the analysis of high molecular-weight compounds. A rather old device, whose sensitivity is now judged to be very poor, is the particle beam interface... 

Figure 16.16 HPLC/MS interface using a particle beam interface. Solvent molecules are lighter than analytes, so their angular dispersion is wider. Heavier molecules, less deflected by the eflect of the vacuum, have a greater probahility of remaining on the axis of the system and being transported to the MS. This baUistic approach was much used formerly in other devices, though now is all hut abandoned.

A schematic of a particle beam interface is shown in Figure 21.13. The eluent from the HPLC column is nebulized using helium gas to form an aerosol in a reduced pressure chamber heated at 70°C. A cone with a small orifice is at the end of the chamber, which leads into a lower pressure area. The difference in pressure causes a supersonic expansion of the aerosol. The hehum and the solvent molecules are lighter than the analyte molecules and tend to diffuse out of the stream and are pumped away. The remaining stream passes through a second cone into a yet lower pressure area, and then the analyte vapor passes into the ion source. The particle beam interface produces electron ionization (El) spectra similar to those of GC-MS, so the vast knowledge of El spectra can be used for analyte identification. 

Figure 7.12 Particle beam HPLC-MS interface and data system reports (a) VG particle beam line interface (b) total ion chromatogram for a series of pesticides obtained using the LINC Particle Beam interface, in the El mode. Peaks A and B are identified below as Rotenone and Atrazine by the library search.

Ionisation methods investigated earlier, including thermospray, fast atom bombardment, and particle beam interfaces have been replaced by electrospray imiiza-tion (ESI) interfaces. Atmospheric pressure chemical ionisation (APCI) has also been applied but is less sensitive towards the PANOs. A combination of ESI with reversed-phase HPLC using acid mobile phases to protonate the PA molecules has most often been used [41]. 

The obvious alternative for the in-line flow-through cell in HPLC-FTIR is mobile-phase elimination ( transport interfacing), first reported in 1977 [495], and now the usual way of carrying out LC-FTIR, in particular for the identification of (minor) constituents of complex mixtures. Various spray-type LC-FTIR interfaces have been developed, namely, thermospray (TSP) [496], particle-beam (PB) [497,498], electrospray (ESP) [499] and pneumatic nebulisers [486], as compared by Som-sen et al. [500]. The main advantage of the TSP-based... 

LC-MS inlet probes support all conventional HPLC column diameters from mobile phase must be eliminated, either before entering or from inside the mass spectrometer, so that the production of ions is not adversely affected. The problem of removing the solvent is usually overcome by direct-liquid-introduction (DLI), mechanical transport devices, or particle beam (PB) interfaces. The main disadvantages of transport devices are that column... 

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