Product-Ion Analysis

TABLE 2.2 Comparisons among the Scan Modes in Tandem Mass Spectrometry 

Mode Mass Analyzer 1 Mass Analyzer 2 Application 

Product ion Selecting Scanning To obtain structural information about the precursor ions 

PIS Scanning Selecting To detect the analytes yielding an identical fragment ion after CID 

NLS Scanning Scanning To detect the analytes losing a common neutral fragment after CID 

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The four-sector mass spectrometer is the ultimate in MS-MS instrumentation and consists of two high-resolution mass spectrometers in series. The strength of these instruments is in terms of their high-mass and high-resolution capabilities for both precursor-ion selection and product-ion analysis. Their cost, however, precludes their primary use for LC-MS and therefore they will not be considered any further here [12]. 

Tian, Q. et al., Screening for anthocyanins using high-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry with precursor-ion analysis, product-ion analysis, common-neutral-loss analysis, and selected reaction monitoring, J. Chromatogr. A, 1091, 72, 2005. 

Ion trap MS is particularly suited for chemical structure elucidation, as it allows for simultaneous ion storage, ion activation and fragmentation, and product ion analysis. The fragmentation pathway of selected ions and the fragmentation products provide information on the molecular structure. Compared with triple-quadrupole and especially with sector instruments, the ion trap instrument provides more efficient conversion of precursor ion into product ions. However, the CID process via resonance excitation, although quite efficient in terms of conversion yield, generally results in only one (major) product ion in the product-ion mass spectrum. MS/MS with a quadrupole ion trap offers a number of advantages ... 

An important issue is the need to acquire both MS and MS-MS data for the unknowns. In general, this requires two injections with data-processing in between. The first run is done in full-spectrum LC-MS mode. Precmsor /w/z-values for relevant peaks in this chromatogram have to be determined in (manual) data processing. The /w/z-values found are then used in a time-scheduled product-ion MS-MS procedure using multiple precmsor-ions. Alternatively, data-dependent acquisition (DDA, Ch. 2.4.2) can be used, as demonstrated by Drexler et al. [106]. An alternative to DDA in a triple-quadrupole instrument is the RF product-ion analysis mode (RFD), proposed by Kienhuis and Geerdink [107]. 

Kaufmann, R., Spengler, B., and Lutzenkirchen, F. (1993). Mass spectrometric sequencing of linear peptides by product-ion analysis in a reflectron time-of-flight mass spectrometer using matrix-assisted laser desorption ionisation, Rapid Commun. Mass Spectrom., 7,902. 

REACTANT ION PREPARATION REACTION REGION PRODUCT ION ANALYSIS... 

Vance and Bailey [94] also carried out measurements of velocity distributions of product ions using a double mass-spectrometer system similar to the one used by Giese and Maier [95] combined with the retardation product-ion analysis by Menendez et al. [96]. They studied the charge transfer and dissociative charge transfer reactions of Hj and N2 with H2, as well as the reaction + H2 H3 + H, and the results yielded considerable new information on the reaction mechanism. 

The precursor ion selection, fragmentation, and product ion analysis can be separated in space or in time, as shown in Figure 1.29. Separation in time requires trapped ions, as available in the quadrupole ion trap or the ion cyclotron resonance trap. Separation in space necessitates at least two physically distinct mass analyzing devices, one for precursor ion selection (MS-1) and one for product ion analysis (MS-2). The simplest in-space tandem instruments are the triple quadrupole mass spectrometer (QqQ), the double-focusing sector tandem mass spectrometer (EB or BE), and the reflectron time-of-flight mass spectrometer. In a triple quadrupole, the first and third quadrupoles (Q) are mass analyzers, while the center quadrupole iq) serves as the collision cell. In sector instruments, a collision cell is situated... 

Linked-field scan at constant B/E. This scan mode is used to obtain a product-ion spectrum on forward- and reverse-geometry inslruments [37]. In this scan, the value of V is fixed and B and E are both scanned while the ratio BIE is held constant. Fragmentation occurs in the first FFR. The mass resolution of the product-ion analysis is much higher selection of the precursor ion is at a lower resolution. 

Quadrupole-Orthogonal Acceleration TOF Instrument As of today, quadrupole (Q)-orthogonal acceleration (oa) TOF (oa-TOF) instrument is the most popular hybrid instrument [58,59] a simphstic pictorial representation is shown in Figure 4.11. The quadrupole section consists of a normal massresolving quadrupole and an rf-only quadrupole. The latter serves as a collision cell and as an ion-accumulation device. For precursor-ion scan, the ions desired are mass-selected by the main quadrupole, accumulated in the collision cell, and a packet of the CID product ions is pushed into the TOF analyzer. The precursor-ion selection by the quadrupole is at a medium resolution, but the product-ion analysis by the TOF section is at a reasonably high resolution. 

Name all types of tandem instruments that can be used for product ion analysis. 

Instrument operation in SRM measurements is exactly identical to that used in the product-ion analysis (Section 4.2) except that instead of acquiring a full-scan spectrum, only the products selected are monitored. A triple-quadrupole instrument is ideally suited for SRM experiments, although magnetic-sector and Qrr instruments have also been used. 

Mass analyzers capable of MS/MS experiments have been classified (Johnson 1990) as either tandem-in-space or tandem-in-time . Tandem-in-space experiments are performed using ion beam instruments (typically a triple quadrupole or a QqTOF) in which the ions are transported from one analyzer or other device to another, each of which is responsible for one of the precursor selection, ion fragmentation and product ion analysis functions. In contrast, ion traps conduct precursor ion selection plus fragmentation and product ion analysis by a tandem-intime sequence of events all within one spatial trapping region. 

A pentaquadrupole (PQ-MS) instrument, thus featuring Q-qjoii-C -q.oii" has been developed and extensively used for the study of gas-phase ion-molecule reactions [58, 59]. The PQ-MS instruments obviously allow sequential product-ion analysis, i.e., MS experiments, but other types of experiments are possible as well. The PQ-MS instrument can be used to perform reactive colhsion of selected ions either in the first or the second collision cell, as illustrated with a wide variety of examples [59]. 

For optimal selectivity, particularly for quantitation with ion trap or triple quadrupole analyzers, MS/MS scanning techniques can be utilized in GCMS. With these techniques, instruments are operated to perform one of three basic experiments including product ion analysis, precursor ion analysis or neutral loss analysis. Ion traps are normally limited to product ion scans in which a particular ion of interest is isolated in the ion trap, subjected to collisional activation and its fragments are detected. Triple quadrupoles can additionally be operated to detect all precursors that generate a common fragment or the analyzer can be set to detect a specific neutral loss characteristic of an analyte functional group. All these methods add specificity to mass spectral detection. 

The development of hybrid instruments has improved product ion analysis to a great extent in comparison to both QqQ and ion-trap mass analyzers. For example, QqTOF instruments have good mass accuracy and resolving power for determining product ions, whereas QqLIT instruments allow for MS" analysis in addition to NLS and PIS analyses [59]. It should be noted that QqTOF mass spectrometers are incapable of virtual NLS and PIS analyses, but can extract NLS- and PlS-like dataset from the array of product ion analysis data. 

There are four main MS/MS modes (including product ion analysis, NLS, PIS, and SRM) that are particularly useful in lipidomics (Figure 2.7). The general principles of these MS/MS techniques can be easily explained by using a QqQ-type mass spectrometer, as briefly described below. Although different parameters are used for mass spectrometers with different hybrid mass analyzers, the underlying chemical principles are quite similar. Comparisons among these techniques are summarized in Table 2.2. 

It should be emphasized that the transitions must be pre-determined, and much work goes into ensuring that the transitions selected have maximum specificity. Moreover, SRM could be considered a speeial ease of PIS in which the first analyzer is fixed at a certain m/z, or a special case of product ion analysis in which the second analyzer is focused at a specific product ion, or a special case of NLS in which both analyzers are separately fixed at the ions of a transition. Accordingly, from an integrated chemical perspective, it should be recognized that the SRM mode only represents a special case of the other three MS/MS techniques with particular advantages associated with and essential to LC-MS analysis where only a limited amount of time is available for data acquisition, and therefore effective duty cycles are critical. The SRM/MRM techniques have been widely used for quantitative analysis of individual lipid species in lipidomics when a mass spectrometer is coupled with LC [57, 85-87]. 

The connection of SRM/MRM mode to other MS/MS modes is briefly mentioned earlier. In fact, the other MS/MS techniques (i.e., product-ion analysis, NLS, and PIS) are also interrelated. This interrelationship provides a foundation to multidimensional mass spectrometry-based shotgun lipidomics (MDMS-SL) and can be schematically illustrated with a simplified model system that comprises three molecular ions (/ i, m2, and m ) of a lipid class (Figure 2.8). 

In this model, each of the three molecular ions has a different m/z, and therefore each yields a different mass spectrum in the product ion analysis mode after CID. Because these molecular ions belong to the same lipid class, these ions possess virtually identical fragmentation patterns. We assume that the fragmentation pattern of these molecule ions shows three features of product ions as highlighted in the broken-line box (Figure 2.8). First, these molecule ions yield product ions that correspond to the loss of a common neutral fragment with a mass of a. This loss gives rise to product ions / 2a from the molecular ions m, m2,... 

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