摘要:

AbstractThe problem of stationary electrolysis of a single‐component ampholyte solution is analyzed. The effect of nonconstant relative mobility is taken into account. It is demonstrated that an incorrect dissociation model (i.e., expressing the resulting ampholyte flux by the arithmetic sum of the fluxes for “cationic” and “anionic” species) may lead to the violation of the so‐called “law of

ISSN:0173-0835    

DOI:10.1002/elps.1150191303

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractWe applied best fitting procedures to capillary electrophoresis (CE) mobility values, measured at varying acidic pH, of a set of 21 peptides with a molecular mass ranging from about 350 to 1850 Da. This method allowed the contemporary measurements ofC‐terminus and carboxylic group of the side‐chain of aspartic and glutamic acid dissociation constants and of peptide Stokes radius at different protonation stages. Stokes radius was related to peptide molecular massMat the power of a fractional coefficient, and best correlation was found at pH 2.25, the fractional coefficient being equal to 0.68. This value is close to that proposed by R. E. Offord (Nature1966,211, 591–593), who suggested a proportionality between the polymer Stokes radius andM2/3. The coefficient value decreases at higher pH, reaching a value of 0.58 at pH 4.25, corresponding to a mean peptide conformational transition towards more compact structures as a consequence ofC‐terminus dissociation. The measurement of the dissociation constants of each peptide allowed us to determine the percentage error on peptide charge predictions performed utilizing mean dissociation constants. Even for the charge, the best predictive performance is obtained at the most acidic edge of the range of the pH studied, mainly at pH 2.25. Conclusively, this study shows that the best performance of predictive models for peptide CE mobility is obtainable in the very acidic pH range (2.25–2.50) and in the absence of electroosmotic flow, and that a satisfactory predictive equation of peptide electrophoretic mobility (m2V−1S−1) is given by μ = 85.

ISSN:0173-0835    

DOI:10.1002/elps.1150191304

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractFused‐silica capillary columns having cationic surfactant moieties (CSM) covalently attached to the capillary inner walls were introduced for the separation of proteins by capillary zone electrophoresis (CZE). The CSM capillary coating proved to be useful in the separation of basic and acidic proteins when modified hydroxypropylcellulose (HPC), namely epoxybutane‐HPC (EBHPC), was adsorbed to the primary CSM coating, yielding a hybrid coating (i.e., coating consisting of a covalenty bound ligand and an adsorbed ligand). The EBHPC layer rendered the capillary surface highly hydrophilic, thus permitting the separation of basic proteins with relatively high plate counts. Also, the CSM coating was useful for the separation of acidic proteins when the capillary wall had, in addition, a negatively charged polymer adsorbed to the surface,e.g., hyaluronic acid. In general, neutral and charged polymeric compounds could be readily adsorbed by the CSM coating, thus altering the zeta potential of the capillary and diminishing solute adsorption to the capillary surface. In other words, the sign of the zeta potential of the capillary surface could be tailored to be of the same sign as the charge of the analytes. Under these conditions, little or no solute‐wall interaction could be observed due to electrostatic repulsion. Although the capillary surface was charged, the presence of adsorbed polymer suppressed or, in most cases, even eliminated the electroosmotic flow

ISSN:0173-0835    

DOI:10.1002/elps.1150191305

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractThe selectivity in the capillary zone electrophoresis (CZE) of a variety of acidic and basic proteins including α‐chymotrypsinogen A, cytochromec, lysozyme, ribonuclease A, ovalbumin, and β‐lactoglobulins A and B, was altered by adding 6‐monodeoxy‐6‐monoamino‐β‐cyclodextrin or carboxymethylated β‐cyclodextrin to the electrophoretic medium of aqueous 50 mMsodium phosphate, pH 2.5. On the other hand, no significant improvement was obtained in the separation upon addition of heptakis (2,6‐di‐O‐methyl)‐β‐cyclodextrin. Whereas protein adsorption on the wall of raw silica capillaries was significant in the absence of cyclodextrin, by addition of β‐cyclodextrin or its derivatives to the background electrolyte, wall adsorption was reduced with concomitant enhancement of the recovery. The results confirm that in various separation techniques, particularly those which employ microcolumns, certain cyclodextrin additives can be useful selectivity enhancers not only in the separation of small sample molec

ISSN:0173-0835    

DOI:10.1002/elps.1150191306

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractCapillary isoelectric focusing (CIEF) is a high resolution technique for protein separation. The on‐column single point detector requires a mobilization step which lengthens the analysis time and causes an uneven resolution along the separation column. The real time and whole column imaging detection has been developed for performing CIEF without mobilization. Three types of imaging detection systems have been developed: optical absorption, refractive index gradient, and laser induced fluorescence. This technique provides a fast analysis speed (about 6 min) and a good resolution of 0.03 pH unit level. Using the absorption imaging detector, ampholyte‐free IEF in tapered capillary is being demonstrated, which eliminates the interference of the expensive carrier ampholytes for protein detection in UV region. Recent advancements in this imaged CIEF technique as well as its applications are revie

ISSN:0173-0835    

DOI:10.1002/elps.1150191307

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractCapillary isoelectric focusing is a highly effective method for the separation of proteins due to focusing as a function of their pIvalues in the separation process. This technique is also effective for certain types of peptides that focus well. Fluorescence labeling and subsequent detection by laser‐induced fluorescence farther enhance the sensitivity of this technique. This paper demonstrates the utility of this technique in an enzyme assay. A synthetic nona peptide, H‐Gly‐Cys‐His‐Glu‐Ala‐Arg‐Ala‐Glu‐Glu‐OH, was labeled with an iodoacetyl derivative of Lissamine rhodamine B at the thiol group of the cysteine residue as a substrate for trypsin. Trypsin catalyzed the cleavage of the Arg‐Ala bond of the labeled substrate, which focused at pH 4.8, and liberated a shortened, labeled product, H‐Gly‐*Cys‐His‐Glu‐Ala‐Arg‐OH that focused at pH 6.9 (*indicates the label). The product peptide at 3–300 pMwas determined with a relative standard deviation of 5.5% (n= 5) by fluorescence detection at 590 nm with excitation by a green line of He‐Ne laser. Incubation of trypsin with the substrate for 10 min at 37°C allowed the determination of 50–250 pg of trypsin, with

ISSN:0173-0835    

DOI:10.1002/elps.1150191308

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractChemiluminescence (CL) detection based on the horseradish peroxidase (HRP) catalyzed reaction of luminol with peroxide was investigated as a post‐separation detection scheme for microchip‐based capillary electrophoresis. An integrated injector, separator and post‐separation reactor was fabricated on planar glass wafers. The fluorescein conjugate of HRP (HRP‐F1) was used as a sample for optimization of the CL detector response. In devices etched 10 μm deep, with an aluminum mirror integrated onto the backside of the detection zone to enhance collection efficiency, the detection limit, estimated at 3 standard deviations (SD) above background noise, for 1 nL injected sample plugs was 35 nMin HRP‐F1. In devices etched 40 μm deep, 8 nL plugs gave a detection limit of 7 nM. Separation and CL detection of the products of an immunological reaction of a F(ab')2fragment of the HRP conjugate of goat anti‐mouse immunoglobulin G (IgG) with mouse IgG was performed on‐chip. A linear calibration curve was obtained for the decrease in peak height of the HRP conjugate (53 μg/mL) with increasing mouse IgG (0–60 μg/mL). When microperoxidase was used as an internal standard, the R2value of a linear least‐squares fit was 0.9867, and the relative errors in the slope and intercept were ± 5.8 an

ISSN:0173-0835    

DOI:10.1002/elps.1150191309

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractElectrophoretic conditions to separate sodium dodecyl sulfate (SDS) complexes of human plasma proteins according to their size differences, by capillary electrophoresis employing linear polyacrylamide as a sieving matrix (LPA‐CE), have been examined. Using the optimized separation conditions, SDS complexes of human plasma proteins not treated with reducing agents were separated into about 40 peaks and shoulders within 60 min. The molecular mass values of major peaks in a separation pattern were estimated from a plot of molecular mass and migration time for standard proteins and some of the major plasma proteins have been identified on the pattern. The electrophoretic conditions were successfully applied for the analysis of proteins in immunoglo‐bulin G (IgG) myeloma s

ISSN:0173-0835    

DOI:10.1002/elps.1150191310

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractA fast and reproducible method was developed to characterize cell lysates by their electrophoretic profiles using capillary electrophoresis (CE). Characteristic and reproducible patterns were recorded for each bacterial strains when “dynamic sieving” CE, using a polymer solution in the capillary, was applied to distinguish four strains of theEnterobacteriaceaefamily. The electropherograms showed distinct differences when comparing them to the sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) protein profiles. This is certainly a result of the differences in the separation principles and in the detection methods of the two tec

ISSN:0173-0835    

DOI:10.1002/elps.1150191311

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY

摘要:

AbstractPersistent inhibition of bacterial growth, called postantibiotic effect (PAE), after a short exposure to a new carbapenem, meropenem, was determined in different strains of theEnterobacteriaceaefamily. Capillary electrophoresis (CE), as well as sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) were used to study the outer membrane protein (OMP) profiles before and after meropenem treatment. CE proved to be suitable for the characterization of the OMP profiles of bacteria. Significant changes in the electrophoretic patterns were observed, showing the consequential effect of meropenem on bacte

ISSN:0173-0835    

DOI:10.1002/elps.1150191312

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1998

数据来源: WILEY