Peptides electropherograms

FIGURE 16.2 Representative base peak electropherograms from CZE runs of RPLC fractions, (a) Fraction 15 (5 peptide identifications) and (b) fraction 20 (19 peptide identifications). Column, bare fused silica capillary, 60 cm x 180 pm ODx30pm i.d. separation voltage, 15 kV observed CZE current, 1.91 p.A running electrolyte, 200 mm acetic acid + 10% isopropanol temperature, 22°C injection time, 10 s at 2 psi ( 4 nL total injection volume) supplementary pressure, 2 psi flow rate, 25nL/min spray voltage, 1.5 kV (reprinted with permission from Electrophoresis). 

Fig. 4 Epitope mapping by ACE/MS in the positive ESI mode. (A) Tryptic digest, 4.6 pmol//j,E, 10-s pressure injection (B) tryptic digest, 10-s injection followed by 25-s injection of antibody (4.7 pmol//xL). Selected ion electropherograms for the m/z. indicated and the total ion electropherograms, respectively. It is obvious that peptide 1 (Tyr-Gly-Gly-Phe-Met-Thr-Ser-Glu-Lys) is captured by the antibody. See text for further explanation. (Reprinted with permission from Ref. 40. Copyright 1997 American Chemical Society.)... 

Figure 8.13 The expanded electropherograms of 12-angiotensin peptides on NCE with a 4.5 cm separation channel [64],...

Peptide mapping studies, generated by the cleavage of a protein into peptide fragments, must be highly reproducible and quantitative. Several electropherograms of protein digests have been obtained when chicken ovalbumin was cleaved by trypsin... 

In Fig. 2a, we see that a 7.5 kDa peptide purified by reversed-phase HPLC shows several peaks by CE, indicating that at this stage in the purification process, the sample is not pure. Further fractionation by cation-exchange HPLC yielded a much purer sample, as shown in Fig. 2c. However, this sample shows a minor component just before the main component the same component is seen after the main component by cation-exchange HPLC. This is explained by the fact that positively charged analyte appears first in the electropherogram, whereas, the same peak would elute last from a... 

While all of these devices used normal zone electrophoresis separation techniques, isoelectric focusing (IEF) methods on microchips have also been interfaced with ESI-MS for high-resolution separations of proteins [37], Figure 5 shows the electropherogram and corresponding mass spectra generated by this device. For on-chip sample preconcentration before separation, a polarityswitching technique was employed to achieve subnanomolar detection limits for many peptide standards [38],... 

Figure 13.9. Affinity capillary electrophoresis-UV-raass spectrometry of a 100-tetrapep-tide library screened for binding to vancomycin (104 fxM in the electrophoresis buffer), (a) The elution of peptides was monitored with UV absorbance during capillary electrophoresis, and the elution time irrieased with increasing affinity for vancomycin. (b) Positive ion electrospray mass spectrum with CID of the Tris adduct of the proton-ated peptide detected at —5 min in the electropherogram shown in a (Reproduced from Ref 52 by permission of the American Chemical ardety.)...

An off-line approach that is simple and useful for peptide/protein sequencing using 5-10 picomoles of material has been demonstrated. Peptide and protein samples were first separated by capillary electrophoresis. Selected peaks were fraction collected and analyzed by both nano-electrospray mass spectrometry and Edman sequencing. A standard peptide mixture, a tryptic-digested protein and intact proteins were used to illustrate this method. Successful fraction collection of each component required reproducible electropherograms, the ability to automatically switch the outlet buffer vessel and the ability to maintain electrophoretic integrity while eluting a peak of interest into a small outlet buffer... 

Figure 8 Stepwise elimination of noninteracting peptides from a mixture of 32 peptides and identification of one tight-binding ligand for vancomycin. Interpretation of each electropherogram is described in the text. Specific experimental conditions are described elsewhere (86a). (Reproduced with permission of the copyright holder, publisher and Journal of Organic Chemistry.)...

Totals of 21 and 16 peaks were visually recognized and matched in the electropherograms obtained from the ovine and bovine/ovine cheese, respectively (Table 16.7). The identification of the peaks corresponding to intact casein and the peptide release during the cheese ripening has been previously described by our research group (32,33). 

Fig. 15 Capillary electrophoresis-mass spectrometry profile of two peptides preconcentrated on-line prior to separation. (A) Electropherogram of neurotensin (1) and angiotensin (2), concentrated and purified by C-18 immobilized to porous beads and monitored at 195 nm after separation by CE. (B) Total ion-current electropherogram of the separated peptides. The experimental conditions were similar to those described in Ref. 120 for gonadotropin-releasing hormone. The limits of detection for the peptides were approximately 1 to 5 ng/mL, depending primarily on the quality of the analyte concentrator-microreactor.

Figure 10.6 Upper panel shows a two-dimensional separation contour plot showing the separation of peptides bradykinin, substance p and bombesin on a PMMA chip. The x axis represents m/z values, the y axis represents the separation and readout times and the gray shade shows the ion intensities. The panel below is a representative electropherogram generated by integrating ion intensities from a two-dimensional plot.

The electropherogram shown in Figure 10.6 represent a separation of three peptides with corrected migration times on the x axis. The separation efficiencies averaged 10s plates per meter. Compared to capillary electrophoresis, lower field strength is employed in PMMA microchip to minimize the Joule heating effect. Nonetheless, separations still occurred in far less time on the chip because of the short channel and generated comparable separation efficiencies. 

Figure 5.1-2. Electropherogram of a mixture of bioactive peptides separated at pH 2.5. Peptides (1) reference (2) bradykinin (3) bradykinin fragment 1-5 (4)substance P (5) [arg]-vasopressin (6) luteinizing hormone releasing hormone (7) bombesin (8) leucine enkephalin (9) methionine enkephalin (10) oxytocin. (Reprinted from Ref. 13, with permission.)...

FIGURE 9.4 (a) Schematic diagram of a CCD-based wavelength-resolved CE-LIF instrument, (b) Wavelength-resolved electropherogram of a mixture of seven peptides containing tryptophan and tyrosine residues. (Adapted from Timperman, A. T., et al., Anal. Chem., 67, 3421, 1995. With permission.)... 

Figure 4.86 Capillary electropherogram for the separation and detection of three peptides from a Model 270A-HT (Applied Biosystems).

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