Analytical methods electrospray ionization

A sensitive and rapid chromatographic procedure using a selective analytical detection method (electrospray ionization-mass spectrometry in SIM mode) in combination with a simple and efficient sample preparation step was presented for the determination of zaleplon in human plasma. The separation of the analyte, IS, and possible endogenous compounds are accomplished on a Phenomenex Lima 5-/rm C8(2) column (250 mm x 4.6 mm i.d.) with methanol-water (75 25, v/v) as the mobile phase. To optimize the mass detection of zaleplon, several parameters such as ionization mode, fragmentor voltage, m/z ratios of ions monitored, type of organic modifier, and eluent additive in the mobile phase are discussed. Each analysis takes less than 6 min. The calibration curve of zaleplon in the range of 0.1-60.0 ng/ml in plasma is linear with a correlation coefficient of >0.9992, and the detection limit (S/N = 3) is 0.1 ng/ml. The within- and between-day variations (RSD) in the zaleplon plasma analysis are less than 2.4% (n = 15) and 4.7% (n = 15), respectively. The application of this method is demonstrated for the analysis of zeleplon plasma samples [14]. 

Electrospray mass spectrometry has developed into a well-established method of wide scope and potential over the past 15 years. The softness of electrospray ionization has made this technique an indispensable tool for biochemical and biomedical research. Electrospray ionization has revolutionized the analysis of labile biopolymers, with applications ranging from the analysis of DNA, RNA, oligonucleotides, proteins as well as glycoproteins to carbohydrates, lipids, gly-colipids, and lipopolysaccharides, often in combination with state-of-the-art separation techniques like liquid chromatography or capillary electrophoresis [1,2]. Beyond mere analytical applications, electrospray ionization mass spectrometry (ESMS) has proven to be a powerful tool for collision-induced dissociation (CID) and multiple-stage mass spectrometric (MSn) analysis, and - beyond the elucidation of primary structures - even for the study of noncovalent macromolecular complexes [3]. 

A connnon feature of all mass spectrometers is the need to generate ions. Over the years a variety of ion sources have been developed. The physical chemistry and chemical physics communities have generally worked on gaseous and/or relatively volatile samples and thus have relied extensively on the two traditional ionization methods, electron ionization (El) and photoionization (PI). Other ionization sources, developed principally for analytical work, have recently started to be used in physical chemistry research. These include fast-atom bombardment (FAB), matrix-assisted laser desorption ionization (MALDI) and electrospray ionization (ES). 

Aerosols can be produced as a spray of droplets by various means. A good example of a nebulizer is the common household hair spray, which produces fine droplets of a solution of hair lacquer by using a gas to blow the lacquer solution through a fine nozzle so that it emerges as a spray of small droplets. In use, the droplets strike the hair and settle, and the solvent evaporates to leave behind the nonvolatile lacquer. For mass spectrometry, a spray of a solution of analyte can be produced similarly or by a wide variety of other methods, many of which are discussed here. Chapters 8 ( Electrospray Ionization ) and 11 ( Thermospray and Plasmaspray Interfaces ) also contain details of droplet evaporation and formation of ions that are relevant to the discussion in this chapter. Aerosols are also produced by laser ablation for more information on this topic, see Chapters 17 and 18. 

Electrospray ionization mass spectrometry (ESI-MS) is an analytical method for mass determination of ionized molecules. It is a commonly used method for soft ionization of peptides and proteins in quadmpole, ion-trap, or time-of-flight mass spectrometers. The ionization is performed by application of a high voltage to a stream of liquid emitted from a capillaty. The highly charged droplets are shrunk and the resulting peptide or protein ions are sampled and separated by the mass spectrometer. 

Electrospray ionization occurs by the same four steps as listed above for thermospray (see Section 4.6). In contrast to thermospray, and most other ionization methods nsed in mass spectrometry, it shonld be noted that electrospray ionization nnnsnally takes place at atmospheric pressure. A similar process carried out under vacuum is known as electrohydrodynamic ionization and gives rise to qnite different analytical results. This technique has not been developed into a commercial LC-MS interface and will not be considered further. 

In this study, the effect of mobile-phase flow rate, or more accurately, the rate of flow of liquid into the LC-MS interface, was not considered but as has been pointed out earlier in Sections 4.7 and 4.8, this is of great importance. In particular, it determines whether electrospray ionization functions as a concentration-or mass-flow-sensitive detector and may have a significant effect on the overall sensitivity obtained. Both of these are of great importance when considering the development of a quantitative analytical method. 

The fact that APCl and electrospray are soft ionization techniques is often advantageous because the molecular ion alone, in conjunction with HPLC separation, often provides adequate selectivity and sensitivity to allow an analytical method to be developed. Again, method development is important, particularly when more than one analyte is to be determined, when the effect of experimental parameters, such as pH, flow rate, etc., is not likely to be the same for each. Electrospray, in particular, is susceptible to matrix effects and the method of standard additions is often required to provide adequate accuracy and precision. 

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,... 

There is a recent hybrid between AP-MALDI and ESI, matrix-assisted laser desorption electrospray ionization (MALDESI) [202], where species desorbed from a MALDI target are subjected to an electrospray before entering the mass spectrometer. The method is similar to ELDI except that the analyte is embedded in a matrix as in MALDI. 

The year 2002 was an extraordinary year for liquid chromatography-mass spectrometry (LC/MS) practitioners. On October 9, 2002, the Royal Swedish Academy of Sciences annonnced their decision to award the Nobel Prize in Chemistry to John B. Fenn, Koichi Tanaka, and Kurt Wiithrich for their development of analytical methods for the identification and structnral analysis of biological macromolecnles. Fenn and Tanaka shared the prize for developing electrospray and soft-laser desorption, respectively. These soft-ionization techniqnes allow macromolecules to be ionized withont fragmentation. 

Direct analysis of solid samples or analytes present on solid surfaces without any sample preparation has always been a topic of interest. Desorption electrospray ionization (DESI) is an atmospheric pressure desorption ionization method introduced by Cooks et al., producing ions directly from the surface to be analyzed, which are then sampled with the mass spectrometer [22, 37]. DESI is based on charged liquid droplets that are directed by a high velocity gas jet (in the order of 300 m s ) to the surface to be analyzed. Analytes are desorbed from the surface and analyzed by mass spectrometer (Eig. 1.15). 

Atmospheric pressure chemical ionization APCI is a method closely related to electrospray ionization. It uses ion-molecule reactions to produce ions from analyte molecules. The sample is electrohydrodynamically sprayed into the source (Figure 14.3). The evaporation of the solvent is often supported by a heated gas at temperatures between 80 and 400°C. Within the source, a plasma is created using a corona discharge needle that is placed close to the end of the metal capillary. In this plasma, proton transfer reactions occur, leading to the ionization of the analyte, mainly by the formation of [M+H]+ ions. Compared to ESI MS, APCI MS is very well suited for the analysis of less-polar components and can therefore... 

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