Nano-LC-ESI

Disadvantages of microbore columns are that the increase in sensitivity is partially negated by the smaller injection volume that can be accommodated (although volumes up to 10 xl can sometimes be injected onto 0.3 mm columns), internal dead volumes have to be minimized, and there is increased experimental difficulty. Nano/LC/ESI/MS is more applicable to the analysis of very small... 

K. Vanhoutte, W. van Dongen, E.L. Esmans, On-line nano-LC-ESI-MS Effect of the mobile-phase composition and the ESI tip design on the performance of a NanoFlow ESI, Rapid Conunun. Mass Spectrom., 12 (1998) 15. 

M.W.H. Pinkse, P.M. Uitto, M.J. Hilhorst, B. Ooms, A.J.R. Heck, Selective isolation at the femtomole level of phosphopeptides from proteolytic digests using 2D-nano-LC-ESI-MS-MS and titanium oxide precolumns. Anal. Chem., 76 (2004) 3935. 

J. Fmbrechts, F. Femiere, W. Van Dongen, F.F. Fsmans, Equilenin-2 -deoxynucleoside adducts analysis with nano-LC-ESI-MS-MS, J. Mass Spectrom., 36 (2001)317. 

Recently, by performing direct nano-LC/ESI-MS sample injection onto a reverse phase capillary without immunoaffinity clean-up, LOD and LOQ in grape extracts of 1 and 2 pg/g, respectively, were reported (Timperio et al., 2006). HPLC analysis was carried out at flow 200nL/min and introducing l xL sample onto the column. Figure 4.9 shows a nano-LC/ESI-MS chromatogram recorded in positive mode in the analysis of acetonitrile solutions containing various concentrations of OTA. 

Nano-LC/ESI-MS has been successfully applied in analysis of peptides from wine proteins (Kwon, 2004). A volume of 2p,L of acetonitrile/H20/acetic acid (2 97.9 0.1 v/v/v) peptide solution (buffer A) is loaded into the capillary C18 column (50 mm x 75 pan i.d., 5 pan particle size, 300 A pore diameter) and peptides are eluted with a gradient from 5% to 80% of buffer B [acetonitrile/H20/acetic acid (90 9.9 0.1 v/v/v)] in buffer A in 10 min at flow rate 0.3p,L/min. Fragmentation of the three strongest parent ions of full MS spectrum is performed, and m/z 700-1300 spectra recorded. The MS/MS spectra... 

Methoxypyrazines 97,106,109,110 Micro- and Nano-LC/ESI/MS 25 MIKE spectrum 78 Mousy N-Heterocycles 269 Mousy off-flavor of wines 268 MS/MS in space 80 MS/MS in time 83... 

Fernandez-Arroyo, S., Gdmez-Martfnez, A., Rocamora-Reverte, L., Quirantes-Pine, R., Segura-Carretero, A., Femandez-Gutidrrez, A., and Ferragut, J. A. 2012. Application of nano LC-ESI-TOF-MS for the metabolomic analysis of phenolic compounds from extra-virgin olive oil in treated colon-cancer cells. J. Pharm. Biomed. Anal. 63 128-134. 

IDENTIFICATION OF HNE-MODIFIED PEPTIDES IN BIOLOGICAL SAMPLES BY SOLID-PHASE ENRICHMENT AND NANO-LC-ESI-MS/MS... 

This protocol describes a procedure for the identification of PTM by HNE to reveal protein targets and specific sites of covalent attachment. Identification is performed by combining proteolytic digestion followed by SPH enrichment and nanoscale liquid chromatography-electrospray ionization tandem mass spectrometry (nano-LC-ESI-MS/MS).The modified proteins are subjected to reduction, alkylation, and subsequent digestion by a proteolytic enzyme. The peptides thus obtained are desalted and the substoichiometric quantities of HNE-modified peptides are fractionated from unmodified species using hydrazide-coated glass beads-based enrichment technique that selectively captures peptide carbonyls (as hydrazones) that are subsequently... 

Fig. 7. Application in practice. The spots of both gels are cut out after 2D-PAGE and Coomassie G-250 staining. The protein spots of one gel are analyzed by on-line nano-LC-ESI-MS/MS after tryptic digestion (a), whereas protein spots of the other gel are analyzed using off-line nano-LC and MALDI-TOF-MS/MS analysis (b).

Table 2. Comparison of the identified a-crystallin A peptides using nano-LC-ESI-MS/MS or nano-LC-MALDI-MS/MS, respectively.

The peptides exclusively identified by nano-LC-ESI-MS/MS are marked red, those detected by nano-LC-MALDI-MS/MS are labeled blue. 

To sum up, nano-LC-ESI-MS/MS and -MALDTMS prove to be complementary, depending on the actual problem. Comprehensive mass spectrometric analysis is ensured by the application of a combination of both methods, resulting in satisfactory information about the proteins in a given sample. 

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