ESI mass spectrometer

Currently PCR and mass spectrometry are performed by two separate instruments. However, there is no reason why PCR followed by simple automated cleanup and mass spectrometry cannot be incorporated into a single integrated instrument. Essentially every configuration of the modern ESI mass spectrometer has been used successfully for the analysis of PCR products, from the highest to the lowest resolution involving. Fourier transform ion cyclotron resonance (FTICR), triple quadrupole, quadrupole-time of flight (Q-TOF), and ion trap.22-24 MS discriminates between two structurally related PCR products by MW difference. Mass accuracy is needed to differentiate the... 

ESI Mass Spectrometer ESI, APCI, Photodissociation, Positive/Negative Ionization... 

Set up HPLC system and begin running mobile-phase solution at a flow rate of 200 (il/rnin into the APCI or ESI mass spectrometer. 

An ESI mass spectrometer coupled online to a microreactor was used to intercept the catalytically active cationic intermediates of the Ziegler-Natta polymerization of ethylene with the homogeneous catalyst system [Cp2Zr(Me)Cl]-MAO (MAO = methylaluminoxane). For the first time these intermediates were studied directly in the solution and their catalytic activity proved.60... 

Although MALDI-MS plays an outstanding role in dendrimer analysis, additional use is also made of modern ESI mass spectrometers for monitoring syntheses, for determination of relative molecular masses, and for studying the purity and polydispersity of dendrimers, including those of higher generations [34]. 

An ESI mass spectrometer equipped with a standard or a nanospray source and with the option for MSn may be used (Q-TOF, Ion Trap, etc.) as well as appropriate software. Ion Trap mass spectrometer (Esquire-LC, Bruker) equipped with a nanospray source was used. 

An example for the separation for flavonoids with HP-RPC is the screening method employed for the systematic identification of glycosylated flavonoids and other phenolic compounds in plant food materials by Lin et al20 These authors used an analytical 4.6 mm x 250 mm 5 pm C18 silica column at 25 °C with linear gradient elution (eluent A (0.1% FA in water and eluent B 0.1% FA in ACN) at 1.0 ml min-1. DAD was performed at 270, 310, 350, and 520 nm to monitor the UV/VIS absorption. The LC system was directly coupled to an ESI mass spectrometer without flow splitting and the mass spectra acquired in the positive and negative ionization mode. The same analytical scheme (aqueous MeOH extraction, reversed-phase liquid chromatographic separation, and diode array and mass spectrometric detection) can be applied to a wide variety of samples and standards and therefore allows the cross-comparison of newly detected compounds in samples with standards and plant materials previously identified in the published literature. 

Data dependent scanning is a MS technique that utilizes algorithms to automatically make real-time decisions to control the acquisition mode of the mass spectrometer. The data system executes the next scan type based on previously collected data. -Eor example, an ESI mass spectrometer can be programmed to implement the following experiment. 

On the other hand, there are limitations to the LC-MS analysis. The two most common ionization techniques available in LC-MS, TSP and electrospray ionization (ESI), yield primarily molecular weight information i.e., little fragmentation is observed to conhrm the structure of the analyte. Thermally induced decomposition " and in-source collision-induced dissociation have been utilized to produce structurally signihcant ions. However, these techniques are often unreliable and can suffer signihcant losses in sensitivity. Alternatively, the on-line photolysis can be used to induce photolytic dissociation of different types of compounds. Volmer et al. have reported the simultaneous detection and conhrmation of several A-nitrosodialkylamines by on-line coupling of a photolysis reactor with an ESI mass spectrometer. 

Molecular mass determination by MS is also useful for analysis of substrate specificities and cleavage patterns of enzymes, including chitinases, chitosanases, lysozymes, and chitin deacety-lases, as summarized with a few representative examples in Table 11.4. Continuous infusion of reaction mixtures into an ESI mass spectrometer (so called real time monitoring) was nsed to analyze the hydrolysis of D (z = 4 - 6) by several chitosanases and some mutants (Dennhart et al. 2008). ESl-MS is also snitable for simnltaneous measurement of individual kinetic constants of enzymes in mixtures of substrates, as reported for a bacterial sulfotransferase with A (z = 2 - 5) (Pi and Leary 2004). 

Mass spectrometry with electrospray ionization (ESl-MS) is especially powerful when combined with HPLC because the two techniques can be used in tandem. With such an instrument the effluent from the HPLC is introduced directly into an ESI mass spectrometer. Thus, chromatographic separation of peptides in a mixture and direct structural information about each of them are possible using this technique. 

Figure 5.38 Electrospray ionization peak areas measured as a function of flow rate for a solution of diltiazem injected (no HPLC column) into a flow of 50 50 acetonitrile water with 1 % acetic acid. A true mass flow dependent detector is predicted (Equation [4.5], Appendix 4.1) to yield peak areas independent of flow rate but this is clearly not observed in (a). A concentration dependent detector should give peak areas proportional to flow rate , and this is observed in (b) at lower flow rates but the peak area response falls below the extrapolated prediction as flow rate increases. Thus in this flow rate range the ESI mass spectrometer did not behave in accord with either of these idealized models.

Metzger and coworkers have studied reactive intermediates of chemical reactions in solution by using a microreactor coupled to an ESI mass spectrometer. The highly stereo- and regioselective dimerization of trows-anethole to give the head-to-head trans, anti, trans-cydobutane initiated by aminium salt proceeds by a radical cation chain mechanism (Scheme 4.4) and this method was further used to study the transient radical cations intermediates in electron transfer-initiated D-A reactions [12-14]. 

Figure 43 The effects of lithium hydroxide concentration on ionization of different lipid classes. A GPL mixture consisting of dil5 0 and di22 6 PG, dil4 l and dil8 l PC, and dil5 0 and di20 4 PE molecular species in a molar ratio of 1 1 10 10 15 15 in 1 1 (v/v) chloro-form/methanol was prepared and analyzed by varying the addition of a different amount of LiOH (as a modifier) as indicated in both negative- (a) and positive-ion (b) modes. ESI-MS analysis of the GPL mixture was conducted utilizing a TSQ ESI mass spectrometer (Thermo Fisher Scientific, San Jose, CA).

Figure 44 The effects of flow rate and hpid solution concentration on the normalized ion count density (i.e., ionization efficiency) of each examined GPL class in the negative-ion mode. The GPL mixture was comprised of dil5 0 and di22 6 PG, dil4 l and dil8 l PC, and dil5 0 and di20 4 PE molecular species in a molar ratio of 1 1 10 10 15 15 in 1 1 (v/v) chloroform/methanol. Ion intensities in ion current of each GPL species were determined from ESI-MS analyses of three independent injections utilizing a TSQ ESI mass spectrometer (Thermo Fisher Scientific, San Jose, CA). The ionization efficiencies of PG (a) and the broken line in (b), PC (b), and PE (b) were calculated from the normahzation of the total ion current of a class to a unit concentration (i.e., pmol/pL) of each class at the indicated lipid concentration and flow rate. The ionization efficiency of PE or PC represents the mean SD of the ionization efficiencies of the class at the flow rates of 1, 2, 4, 6, 8, 10, and 15 pL/rnin. The ionization efficiency of PG in (b) represents the mean SD of the ionization efficiencies in (a) at the flow rates of 4, 6, 8, 10, and 15 pL/min. Some of the error bars are within the symbols. Han et al. [31]. Adapted with permission of Springer.

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