Atmospheric pressure ionization API

The advent of atmospheric-pressure ionization (API) provided a method of ionizing labile and nonvolatile substances so that they could be examined by mass spectrometry. API has become strongly linked to HPLC as a basis for ionizing the eluant on its way into the mass spectrometer, although it is also used as a stand-alone inlet for introduction of samples. API is important in thermospray, plasmaspray, and electrospray ionization (see Chapters 8 and 11). 

After being formed as a spray, many of the droplets contain some excess positive (or negative) electric charge. Solvent (S) evaporates from the droplets to form smaller ones until, eventually, ions (MH+, SH+) from the sample M and solvent begin to evaporate to leave even smaller drops and clusters (S H n = 1, 2, 3, etc.). Later, collisions between ions and molecules (Cl) leave MH+ ions that proceed into the mass analyzer. Negative ions are formed similarly. 

Since ions and neutral molecules are formed close together in an API source, many ion/molecule collisions occur as in Cl, and so the ion evaporation process also has impressed upon it the characteristics of Cl. Therefore, API is usually thought to involve a mix of ion evaporation and chemical ionization. 

The mix of ions, formed essentially at or near ambient temperatures, is passed through a nozzle (or skimmer) into the mass spectrometer for mass analysis. Since the ions are formed in the vapor phase without having undergone significant heating, many thermally labile and normally nonvolatile substances can be examined in this way. 

An example of proton (H ) transfer from a protonated solvent molecule (SH ) or cluster to form a quasi-molecular ion (MH ) of the substrate (M). 

Ionization at atmospheric pressure was for many years considered to be an experimental curiosity despite excellent early work (Homing 1974, 1974a Carroll 1975). However, API techniques nowadays dominate on-line LC-MS work. There are currently three kinds of API sources in common use for trace quantitative analyses, namely, atmospheric pressure chemical ionization (APCI), atmospheric pressure photoionization (APPI) and electrospray ionization (ESI). In addition, atmospheric pressure MALDI has been investigated but not as an on-line approach to quantitative LC-MS. Since the highly practical discussions of Chapters 9 and 10 will be concerned mainly with LC-MS analyses (since GC-MS is now almost routine), an extended discussion of API methods is appropriate here. 

The major problem with all API techniques for mass spectrometry concerns the transfer of the ions from the atmospheric pressure ion source into the vacuum required for operation of the miz analyzer itself, a pressure drop by a factor 10 Such a transfer involves a sudden expansion of the gas at some stage and this tends to enhance the condensation of solvent molecules (particularly water) on the ions to produce clusters of various sizes that redistribute the total ion current among several species thus compUcatmg the spectra and reducing S/B values. An interface between any atmospheric pressure ionization (API) source and a mass spectrometer must be able to deal with the pressure ratio (pumping speed is a crucial factor here. Section 6.6.1) and the de-clustering of the analyte ions before m z analysis and detection. 

Despite these difficulties (discussed in Section 5.3.3a) API sources are currently used widely in combination with a wide range of liquid chromatography methods, i.e. normal and reverse phase, isocratic and gradient, normal bore (4.6 mm i.d.) and capillary columns, as well as ESI with capillary electrophoresis and APCI with GC. In the context of trace level quantitation, API techniques are most often used in combination with reverse phase HPLC, and it is this combination that will be the main focus of discussions of matrix effects (Sections 5.1.1 and 5.3.6a) and of the more practical aspects in Sections 9.6 and 10.10.4.1d. In this regard it is worth noting here that in reverse phase chromatography using e.g., a Cjg-derivatized silica 

A phenomenon common to all API techniques is the production of ionized intact analyte molecules in a variety of forms of the type [A-l-X], where X can be one of several ionic species the most common is X = H , but others are commonly observed (see Table 5.2 for those observed in APCI and ESI). It is possible to use any of the adduct ions hsted in Table 5.2 as the species monitored in quantitative LC-MS, but it is important to ensure that only one form (one adducting ion X) contributes significantly to the ion current derived from the analyte (to maximize sensitivity) and also that the concentration of X is constant under the conditions used (to maintain a constant sensitivity for [A -l-X] ). 

Many new applications in mass spectrometry concern either the measurement of elements or the study of unstable and polar biomolecules (M 2000Da). These applications have become possible with the advent of atmospheric pressure ionization (API) techniques applicable to compounds in solution following nebulization. 

Argon plasmas are used in optical emission spectrometry (cf. section 14.3.1) to atomize and ionize elements in order to provoke the emission of characteristic spectral lines. Hence, it is not surprising that the same plasma torches are employed to ionize inorganic samples in mass spectrometry. Thermal ionization is induced at high temperatures in a gaseous sample with microwave or an inductively coupled plasma. 

Hyphenated techniques as micro-HPLC/SM or HPCE/SM have been the driving force in the development of different highly sensitive devices, which have in common the nebulization of the mobile liquid phase, issued from the separative 

This mode of ionization is associated with photon induced ionization at around 10 eV. In place of the corona assembly, a UV lamp is used as for the photoionization detection (cf section 2.7.5). This process, which yields few fragments, is only interesting for molecules of low polarity. 

The advantage of these soft methods of ionization is to obtain pseudomolecular and multicharged ions (z can be greater than 30), formed prior to entry into the spectrometer. With the exception of APCI they are able to extend the range 

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For a more detailed description of the ionization process inherent in electrospray, please see Chapter 9, which discusses atmospheric pressure ionization (API), The reader also should compare electrospray with thermospray (see Chapter 11). 

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... 

Another big advance in the appHcation of ms in biotechnology was the development of atmospheric pressure ionization (API) techniques. There are three variants of API sources, a heated nebulizer plus a corona discharge for ionization (APCl) (51), electrospray (ESI) (52), and ion spray (53). In the APCl interface, the Ic eluent is converted into droplets by pneumatic nebulization, and then a sheath gas sweeps the droplets through a heated tube that vaporizes the solvent and analyte. The corona discharge ionizes solvent molecules, which protonate the analyte. Ions transfer into the mass spectrometer through a transfer line which is cryopumped, to keep a reasonable source pressure. 

Atmospheric-pressure ionization (API) A general term used for all forms of ionization mat take place at atmospheric pressure. 

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,... 

As with GC, the combination of MS and MS/MS detection with LC adds an important confirmatory dimension to the analysis. Thermospray (TSP) and particle beam (PB) were two of the earlier interfaces for coupling LC and MS, but insufficient fragmentation resulted in a lack of structural information when using TSP, and insufficient sensitivity and an inability to ionize nonvolatile sample components hampered applications using PB. Today, atmospheric pressure ionization (API) dominates the LC/MS field for many environmental applications. The three major variants of API... 

HPLC, ConstaMetric 3500 MS and ConstaMetric 3200 MS Mass spectrometer, TSQ 7000 with atmospheric pressure ionization (API) electrospray interface Robotcoupe, Model RSI 25 Syringes, Luer lock, 10-mL... 

Hewlett-Packard 1100 Series LC-MSD equipped with an atmospheric pressure ionization (API) source (APcI or ESI)... 

TSQ 7000 mass spectrometer (Finnigan MAT) with atmospheric pressure ionization (API) interface (ESI mode) or equivalent... 

As a more sensitive detection method, MS can be very useful in amino acid determinations. For example, S-carboxymethyl-(R) cysteine or SCMC, is a mucolytic agent used in the treatment of respiratory diseases. The development of a method utilizing high performance IEC and atmospheric pressure ionization (API) mass spectrometry to quantify SCMC in plasma has been described.66 This method is simple (no derivatization needed), rapid (inn time 16 min.), sensitive (limit of quantification 200 ng/mL in human plasma), and has an overall throughput of more than 60 analyses per day. API-MS was used successfully with IEC to determine other sulfur-containing amino acids and their cyclic compounds in human urine.67 IEC has also been used as a cleanup step for amino acids prior to their derivatization and analysis by gas chromatography (GC), either alone or in conjunction with MS.68 69... 

The advent of the atmospheric pressure ionization (API) source in the early 1990s allowed direct coupling of LC to MS. By the mid-1990s, this technology was a common in drug metabolism laboratories. The enhanced selectivity of tandem mass spectrometry (MS/MS) experiments reduced the need for exhaustive chromatographic separations prior to detection and this feature was exploited to significantly reduce analysis times. 

Atmospheric pressure chemical ionization (APCI) was introduced in 1973 by Horning et al. [38, 42, 43] and coupled to GC. This is also the introduction of atmospheric pressure ionization (API) in general. The next year corona discharge was introduced for ion generation as well as successful coupling to LC [44, 45]. In APCI of a liquid, a pneumatic nebulizer induces the flow of liquid to form a spray at atmospheric pressure. The spray droplets pass a corona discharge electrode situated close to the orifice, which... 

D. I. Carroll, I. Dzidic, R. N. Stillwell, M. G. Homing, and E. C. Homing. Subprogram Detection System for Gas Phase Analysis Based upon Atmospheric Pressure Ionization (API) Mass Spectrometry. Anal. Chem., 46(1974) 706-710. 

Solutes were tentatively identified by atmospheric pressure ionization (API)-electrospray-mass selective detector (gas temperature 350°C, flow rate 101/min, nebulizer pressure 30 psi, quadrupole temperature 30°C, capillary voltage 3 500 V). 

In principle, mass spectrometry is not suitable to differentiate enantiomers. However, mass spectrometry is able to distinguish between diastereomers and has been applied to stereochemical problems in different areas of chemistry. In the field of chiral cluster chemistry, mass spectrometry, sometimes in combination with chiral chromatography, has been extensively applied to studies of proton- and metal-bound clusters, self-recognition processes, cyclodextrin and crown ethers inclusion complexes, carbohydrate complexes, and others. Several excellent reviews on this topic are nowadays available. A survey of the most relevant examples will be given in this section. Most of the studies was based on ion abundance analysis, often coupled with MIKE and CID ion fragmentation on MS " and FT-ICR mass spectrometric instruments, using Cl, MALDI, FAB, and ESI, and atmospheric pressure ionization (API) methods. 

With external ion sources it became feasible to interface any ionization method to the QIT mass analyzer. [171] However, commercial QITs are chiefly offered for two fields of applications i) GC-MS systems with El and Cl, because they are either inexpensive or capable of MS/MS to improve selectivity of the analysis (Chap. 12) and ii) instruments equipped with atmospheric pressure ionization (API) methods (Chap. 11) offering higher mass range, and some 5-fold unit resolution to resolve isotopic patterns of multiply charged ions (Fig. 4.47). [149,162,172,173]... 

Nowadays, ESI is the leading member of the group of atmospheric pressure ionization (API) methods and the method of choice for liquid chromatography-mass spectrometry coupling (LC-MS, Chap. 12). [10-13] Currently, ESI and MALDI (Chap. 10) are the most commonly employed ionization methods and they opened doors to the widespread biological and biomedical application of mass spectrometry. [5,10,11,13-17] Moreover, ESI serves well for the analysis of ionic metal complexes [18,19] and other inorganic analytes. [20-22]... 

Atmospheric pressure ionization (API) was the first technique to directly interface solution phase with a mass analyzer. [26] In API, a solution of the analyte is injected into a stream of hot nitrogen to rapidly evaporate the solvent. The vapor passes through a Ni source where electrons emitted from the radioactive Ni isotope initiate a complex series of ionizing processes. Beginning with the ioniza-... 

Homing, E.C. Carroll, D.I. Dzidic, I. Haegele, K.D. Homing, M.G. Stillwell, R.N. Atmospheric Pressure Ionization (API) MS. Solvent-Mediated Ionization of Samples Introduced in Solution and in a Liquid Chromatograph Effluent Stream. J. Chromatogr. Sci. 1974, 72,725-729. 

Analytes must be liberated from their associated solvent molecules as well as be ionized to allow mass separation. Several ionization methods enable ion production from the condensed phase and have been used for the coupling of CE to MS. Among them, atmospheric pressure ionization (API) methods, matrix-assisted laser desorption/ionization (MALDI), and inductively coupled plasma (ICP) ionization are mainly used. API techniques are undoubtedly the most widespread ionization sources and cover different analyte polarity ranges. 

Eiceman et d. [23] determined that mixtures of product ions, M 02 and (M-H), can be observed when ion formation and determination are fast, as with an atmospheric pressure ionization (API) mass spectrometer. In contrast, usually (M—H) or M-02 (but not both) is observed with explosives in IMS drift mbes where residence times for ions are Sms or greater, enough time for proton abstraction to be complete [24]. Alternatively, the M-02 ion may undergo dissociation with charge retention by the analyte molecule as shown in Eq. (3) and Figure 6 ... 

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