Nanospray

The HPLC system comprised a 75 ftm x 15 cm PepMap column with a linear gradient of acetonitrile/0.1% aqueous formic acid (5 to 50% acetonitrile over 45 min) at a flow rate of 250 nlmin . Positive-ion electrospray ionization was employed using a nanospray interface. MS-MS specna were acquired over the range m/z 40 to 2000 at a rate of 1 s per scan. 

Different options are available for LC-MS instruments. The vacuum system of a mass spectrometer typically will accept liquid flows in the range of 10-20 p,L min-1. For higher flow-rates it is necessary to modify the vacuum system (TSP interface), to remove the solvent before entry into the ion source (MB interface) or to split the effluent of the column (DLI interface). In the latter case only a small fraction (10-20 iLrnin ) of the total effluent is introduced into the ion source, where the mobile phase provides for chemical ionisation of the sample. The currently available commercial LC-MS systems (Table 7.48) differ widely in characteristics mass spectrometer (QMS, QQQ, QITMS, ToF-MS, B, B-QITMS, QToF-MS), mass range m/z 25000), resolution (up to 5000), mass accuracy (at best <5ppm), scan speed (up to 13000Das-1), interface (usually ESP/ISP and APCI, nanospray, PB, CF-FAB). There is no single LC-MS interface and ionisation mode that is readily suitable for all compounds... 

Variations may also be introduced at the CE end of the coupling. CZE is the most widely practised mode of CE. Nanospray CZE-ESI-MS [889],... 

Estimating the amount of a metabolite when an authentic reference standard is not available is still a challenge. Yu et al.191 described a procedure that uses the results of an in vitro metabolite identification based on a test compound that produces 14C-labelled metabolites essentially the 14C-labelled metabolites are used to provide a correction factor for the MS response when assaying samples that contain the same metabolite in a study that did not use the 14C-labelled test compound. Flop192 described another novel approach for metabolite quantitation based on the observation that the MS responses for most compounds are very similar to responses from nanospray ESI. Valaskovic et al.193 also reported equimolar MS responses for multiple compounds when the flow rate to the nanospray ESI source was set to about 10 nl/min. It is too soon to know whether these intriguing findings can be readily applied to discovery metabolite identification studies. 

Hsieh, F. Baronas, E. Mnir, C. Martin, S.A. A Novel Nanospray CE/MS Interface. Rapid Commun. Mass Spectrom. 1999,13, 67-72. 

Several reports concerning the development of stable and rugged sheathless interfaces were proposed. The first sheathless interface was developed by Olivares et al., and two types of sheathless interfaces are currently distinguished. The first one consists of a nanospray needle, which is inserted with a connection unit to the CE capillary. This setup allows changing the spray needle alone independently on the capillary exchange.The second approach involves the use of the end of capillary tip as an emitter with the help of a capillary-outlet conductive coating " or by inserting a conductive wire into the capillary outlet. 

Lord et al. analyzed a mixture of steroids by CEC-ESI/MS and interfaced externally tapered CEC columns in both sheathless and sheath-flow arrangement. Sensitivity was found 20-fold higher in the sheathless configuration. The same conclusion was drawn by Warriner et ah, who evaluated CEC-nanospray/MS vs. CEC-microspray/MS with an ion trap using five corticosteroids. Cahours et al. used CEC-ESI/MS for a drug metabolism study and obtained a simultaneous baseline separation of flunitrazepam and its major metabolites. For CEC-ESI/MS coupling, the commercially available packed-CEC column was connected... 

Microchips fabrication with integrated tips can result in improved spray repeatability and efficiency since alignment and dead volume are not a critical issue anymore. However, production of fine and robust nanospray emitters as an integral part of a microdevice is not trivial, and highly specialized microfabrication procedures are required. Microfluidic devices with integrated ESI tips have been produced for infusion experiments, but to date, no microchips with such a design was fabricated for CE separation prior to MS detection. 

Viberg, P., Nilsson, S., and Skog, K. (2004). Nanospray mass spectrometry with indirect conductive graphite coating. Anal. Chem. 76, 4241 —4244. 

Wetterhall, M., Nilsson, S., Markides, K. E., and Bergquist, J. (2002). A conductive polymeric material used for nanospray needle and low-flow sheathless electrospray ionization applications. Anal. Chem. 74, 239-245. 

Vuorensola, K., Siren, H., Kostiainen, R., and Kotiaho, T. (2002). Analysis of catecholamines by capillary electrophoresis and capillary electrophoresis-nanospray mass spectrometry. Use of aqueous and non-aqueous solutions compared with physical parameters. /. Chromatogr. A 979, 179—189. 

Warriner, R. N., Craze, A. S., Games, D. E., and Lane, S. J. (1998). Capillary electrochromato-graphy/mass spectrometry a comparison of the sensitivity of nanospray and microspray ionization techniques. Rapid Commun. Mass Spectrom. 12, 1143-1149. 

Use of Automated Nanospray for Greater Sensitivity and Smaller Sample Size (Less Protein/Drug)... 

Henion Characterizatioin of a fiilly automated nanospray system with mass spectrometric detection for proteomic analyses, J. Biomol. Tech. 2002, 13, 72-84. 

A number of different types of ESI sources, known as nanospray sources, have been designed that can operate at lower sample flow rates (10-200 nL min ). These generate smaller droplets and improve the signal intensity of the protein-ligand noncovalent complexes further, with the added benefit of reducing protein consumption up to 100-fold compared to standard ESI flow rates. Nanospray has also been reported to be more tolerant to nonvolatile cations in solution [37]. Recently, an automated fabricated chip nanospray source has been developed. This chip-based device has improved the ease-of-use for nanospray, while the design eliminates carryover effects as the spray is produced directly from an orifice on each sample well of the chip [38]. 

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