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Papers by Norberto Guzman
Journal of Chromatography a, Oct 1, 1991
A procedure to improve on-column fluorescence detection for capillary zone electrophoresis is rep... more A procedure to improve on-column fluorescence detection for capillary zone electrophoresis is reported. A fluorescence detector was built using an epillumination fluorescence microscope and an argon-ion air-cooled laser. The 488-nm line was isolated with a band pass filter to ...
Analytical and Bioanalytical Chemistry, Jan 30, 2007
Several CE methodologies have been described for the analysis of rHuEPO in concentrated solutions... more Several CE methodologies have been described for the analysis of rHuEPO in concentrated solutions, but the inherently limited concentration sensitivity of CE precludes the detection of EPO at the levels found in biological fluids. In this work, we have investigated an on-line immunoaffinity solid-phase extraction capillary electrophoresis (IA-CE) methodology for the selective preconcentration of EPO in diluted solutions. The preliminary results obtained using a custom-made immunoaffinity sorbent prepared from an anti-human EPO polyclonal antibody and glutaraldehyde-glass beads show the potential of this novel approach. The summarized findings are discussed in detail as a starting point for our ongoing investigations.
The techniques of CE and on-line capillary electrophoresis-mass spectrometry (CE-MS) have been wi... more The techniques of CE and on-line capillary electrophoresis-mass spectrometry (CE-MS) have been widely used for the analysis of many chemically diverse molecules. These methods of analysis allow analyte separations in aqueous solutions that are complementary to classical techniques such as HPLC and HPLC-MS. However, the one major limitation of CE is the fact that the best performance is normally obtained in analyzing small sample volumes (typically < 50 nL for a 50-micron-i.d. capillary). Ultimately, this leads to a relatively poor concentration limit of detection (CLOD) for CE and CE-MS when compared to that of HPLC or HPLC-MS. Recently, the analyte concentrator and membrane preconcentration cartridge have been described for use on-line with the CE capillary. Common to many of the approaches that led to the development of these devices is the incorporation of a suitable stationary phase at the inlet of the CE capillary. This relatively simple modification permits the introduction of much larger sample volumes (> 100 microL) into the CE capillary, and lowers both the CE and CE-MS CLOD. Detection of analytes present in complex mixtures at concentrations of < 200 fg/mL is reported to be possible when using such techniques in conjunction with CE and CE-MS. Additionally, the analyte concentrator has been developed to analyze microreactions on-line with the CE capillary. Recently, analyte derivatization and enzymatic protein digestion on-line with CE separations were reported. The purpose of this manuscript is to discuss and critically review these techniques and all described attempts at improving the CLOD of CE and CE-MS techniques using an adsorptive mechanism at the inlet of the CE capillary.
Lc Gc North America, 2001
... Prefix Symbol Factor Numerical Value tera T 1012 1 000 000 000 000 giga G 109 1 000 000 000me... more ... Prefix Symbol Factor Numerical Value tera T 1012 1 000 000 000 000 giga G 109 1 000 000 000mega M 106 1 000 000 kilo k 103 1000 hecto† h 102 100 deka† da 101 10 — — 100 1 deci† d 10 1 0.1 centi† c 10 2 0.01 milli m 10 3 0.001 micro µ 10 6 0.000001 ...
Analytical Chemistry, Nov 1, 2003
The separation of the glycoforms of erythropoietin (EPO) by capillary electrophoresis (CE) was re... more The separation of the glycoforms of erythropoietin (EPO) by capillary electrophoresis (CE) was recently published as a monograph by the European Pharmacopoeia (European Pharmacopoeia 4 2002, 1316, 1123-1128). Although the experimental CE conditions employed a background electrolyte containing additives suitable for on-line UV-absorption detection, they were not appropriate for on-line mass spectrometry (MS) detection. In this work, an attempt was made to investigate experimental conditions employing volatile electrolyte systems to achieve the separation and characterization of EPO glycoforms using CE and ESI-MS methodologies. The influence of several operating conditions, such as the coating of the internal walls of the capillary as well as the composition, concentration, and the pH of the separation buffer were investigated. The results demonstrated that when the internal walls of the capillaries were permanently coated with Polybrene and a buffer electrolyte containing 400 mM of HAc-NH4Ac (acetic acid-ammonium acetate), pH 4.75, was used, a significantly reproducible separation was achieved for EPO glycoforms. Intact EPO was characterized by two mass spectrometry techniques: electrospray ionization (ESI-MS) and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF-MS). The data demonstrated that MALDI-TOF-MS provided a good approximation to an average molecular mass of the EPO molecule. However, it was still necessary to carry out further separation of the intact EPO glycoforms in order to obtain molecular mass information when ESI-MS was used.
Journal of Liquid Chromatography Related Technologies, Sep 23, 2006
A quantitative spectrophotometrc assay for the determination of proline and hydroxyproline is des... more A quantitative spectrophotometrc assay for the determination of proline and hydroxyproline is described. The assay is based on the derivatization of the imino acids with the fluorogenic chromophore fluorescamine and the separation of the derivatized analytes by capillary electrophoresis. Fluorescamine reacts efficiently with secondary amino acids to form nonfluorescent aminoenone type chromophores which are easily detected at the low UV
J Liq Chromatogr Relat Techno, 1990
Abstract A quantitative ultraviolet detection method for the determination of urinary metabolites... more Abstract A quantitative ultraviolet detection method for the determination of urinary metabolites is described using capillary zone electrophoresis. The determination of these metabolites is simple, fast. reproducible and utilizes very small amounts of sample. This ...
Http Dx Doi Org 10 1080 01483919108049300, Oct 23, 2006
J Liq Chromatogr Relat Techno, 1993
... JUAN P. ADVIS' AND NORBERT0 A. GUZMAN2f 'Deparhnent of A nid Sciences Rutge... more ... JUAN P. ADVIS' AND NORBERT0 A. GUZMAN2f 'Deparhnent of A nid Sciences Rutgers University New Brunnuick, New Jersey 08903 2Protein Research Unit Princeton Biochemicals, Inc. Princeton, New Jersey 08543 ABSTRACT ...
J Liq Chromatogr Relat Techno, 1993
... LUIS HERNANDEZ', JOSE ESCALONA', PHILLIPE VERDEGUER2, AND NORBERT0 A. GUZMAN3* ... more ... LUIS HERNANDEZ', JOSE ESCALONA', PHILLIPE VERDEGUER2, AND NORBERT0 A. GUZMAN3* 'Department of Physwlogv, School of Medicine Los Andes University ... In this experiment, the optimal molar excess to la-bel low concentrations of glutamate was measured. ...
Enzyme, 1984
The physicokinetic and immunologic properties of the purified low and high uptake forms of the hu... more The physicokinetic and immunologic properties of the purified low and high uptake forms of the human lysosomal hydrolase, alpha-L-iduronidase, have been determined and compared. The apparent Km and Vmax values for the low and high uptake forms were similar toward two artificial substrates, 4-methylumbelliferyl-alpha-L-iduronide (0.07 and 0.06 mmol/l; 16.15 and 14.85 mumol/min/mg, respectively), and phenyl-alpha-L-iduronide (1.42 and 1.66 mmol/l; 0.83 and 1.05 mumol/min/mg, respectively), and one natural substrate, anhydro-[3H]-mannitol-iduronide (0.86 and 1.04 mmol/l; 2.50 and 2.79 mumol/min/mg, respectively). The pH optima for both purified forms also were similar for each of the three substrates ( approximately 3.50, approximately 3.50, and approximately 4.50, respectively). Heparin markedly inhibited the 4-methylumbelliferyl-alpha-L-iduronide activities of both the low and high uptake forms, while dermatan sulfate and heparan sulfate were more inhibitory toward the low uptake act...
Organization and Assembly of Plant and Animal Extracellular Matrix, 1990
Journal of Liquid Chromatography, 1995
A method to perform on-line sample preconcentration of serum immunoglobulin E by affinity capture... more A method to perform on-line sample preconcentration of serum immunoglobulin E by affinity capture is described. Purified anti-IgE antibodies were covalently bound to an analyte concentrator-reaction chamber or cartridge. The immunoglobulins (IgE) were bound to and eluted from the cartridge by the optimum dissociating buffer system, and the eluent(s) were then subjected to capillary electrophoresis.The first design used was a
The New England journal of medicine, Jan 12, 1979
Journal of capillary electrophoresis and microchip technology, 2008
Journal of capillary electrophoresis and microchip technology, 2007
Journal of Chromatography a, Oct 1, 1991
A procedure to improve on-column fluorescence detection for capillary zone electrophoresis is rep... more A procedure to improve on-column fluorescence detection for capillary zone electrophoresis is reported. A fluorescence detector was built using an epillumination fluorescence microscope and an argon-ion air-cooled laser. The 488-nm line was isolated with a band pass filter to ...
Analytical and Bioanalytical Chemistry, Jan 30, 2007
Several CE methodologies have been described for the analysis of rHuEPO in concentrated solutions... more Several CE methodologies have been described for the analysis of rHuEPO in concentrated solutions, but the inherently limited concentration sensitivity of CE precludes the detection of EPO at the levels found in biological fluids. In this work, we have investigated an on-line immunoaffinity solid-phase extraction capillary electrophoresis (IA-CE) methodology for the selective preconcentration of EPO in diluted solutions. The preliminary results obtained using a custom-made immunoaffinity sorbent prepared from an anti-human EPO polyclonal antibody and glutaraldehyde-glass beads show the potential of this novel approach. The summarized findings are discussed in detail as a starting point for our ongoing investigations.
The techniques of CE and on-line capillary electrophoresis-mass spectrometry (CE-MS) have been wi... more The techniques of CE and on-line capillary electrophoresis-mass spectrometry (CE-MS) have been widely used for the analysis of many chemically diverse molecules. These methods of analysis allow analyte separations in aqueous solutions that are complementary to classical techniques such as HPLC and HPLC-MS. However, the one major limitation of CE is the fact that the best performance is normally obtained in analyzing small sample volumes (typically < 50 nL for a 50-micron-i.d. capillary). Ultimately, this leads to a relatively poor concentration limit of detection (CLOD) for CE and CE-MS when compared to that of HPLC or HPLC-MS. Recently, the analyte concentrator and membrane preconcentration cartridge have been described for use on-line with the CE capillary. Common to many of the approaches that led to the development of these devices is the incorporation of a suitable stationary phase at the inlet of the CE capillary. This relatively simple modification permits the introduction of much larger sample volumes (> 100 microL) into the CE capillary, and lowers both the CE and CE-MS CLOD. Detection of analytes present in complex mixtures at concentrations of < 200 fg/mL is reported to be possible when using such techniques in conjunction with CE and CE-MS. Additionally, the analyte concentrator has been developed to analyze microreactions on-line with the CE capillary. Recently, analyte derivatization and enzymatic protein digestion on-line with CE separations were reported. The purpose of this manuscript is to discuss and critically review these techniques and all described attempts at improving the CLOD of CE and CE-MS techniques using an adsorptive mechanism at the inlet of the CE capillary.
Lc Gc North America, 2001
... Prefix Symbol Factor Numerical Value tera T 1012 1 000 000 000 000 giga G 109 1 000 000 000me... more ... Prefix Symbol Factor Numerical Value tera T 1012 1 000 000 000 000 giga G 109 1 000 000 000mega M 106 1 000 000 kilo k 103 1000 hecto† h 102 100 deka† da 101 10 — — 100 1 deci† d 10 1 0.1 centi† c 10 2 0.01 milli m 10 3 0.001 micro µ 10 6 0.000001 ...
Analytical Chemistry, Nov 1, 2003
The separation of the glycoforms of erythropoietin (EPO) by capillary electrophoresis (CE) was re... more The separation of the glycoforms of erythropoietin (EPO) by capillary electrophoresis (CE) was recently published as a monograph by the European Pharmacopoeia (European Pharmacopoeia 4 2002, 1316, 1123-1128). Although the experimental CE conditions employed a background electrolyte containing additives suitable for on-line UV-absorption detection, they were not appropriate for on-line mass spectrometry (MS) detection. In this work, an attempt was made to investigate experimental conditions employing volatile electrolyte systems to achieve the separation and characterization of EPO glycoforms using CE and ESI-MS methodologies. The influence of several operating conditions, such as the coating of the internal walls of the capillary as well as the composition, concentration, and the pH of the separation buffer were investigated. The results demonstrated that when the internal walls of the capillaries were permanently coated with Polybrene and a buffer electrolyte containing 400 mM of HAc-NH4Ac (acetic acid-ammonium acetate), pH 4.75, was used, a significantly reproducible separation was achieved for EPO glycoforms. Intact EPO was characterized by two mass spectrometry techniques: electrospray ionization (ESI-MS) and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF-MS). The data demonstrated that MALDI-TOF-MS provided a good approximation to an average molecular mass of the EPO molecule. However, it was still necessary to carry out further separation of the intact EPO glycoforms in order to obtain molecular mass information when ESI-MS was used.
Journal of Liquid Chromatography Related Technologies, Sep 23, 2006
A quantitative spectrophotometrc assay for the determination of proline and hydroxyproline is des... more A quantitative spectrophotometrc assay for the determination of proline and hydroxyproline is described. The assay is based on the derivatization of the imino acids with the fluorogenic chromophore fluorescamine and the separation of the derivatized analytes by capillary electrophoresis. Fluorescamine reacts efficiently with secondary amino acids to form nonfluorescent aminoenone type chromophores which are easily detected at the low UV
J Liq Chromatogr Relat Techno, 1990
Abstract A quantitative ultraviolet detection method for the determination of urinary metabolites... more Abstract A quantitative ultraviolet detection method for the determination of urinary metabolites is described using capillary zone electrophoresis. The determination of these metabolites is simple, fast. reproducible and utilizes very small amounts of sample. This ...
Http Dx Doi Org 10 1080 01483919108049300, Oct 23, 2006
J Liq Chromatogr Relat Techno, 1993
... JUAN P. ADVIS' AND NORBERT0 A. GUZMAN2f 'Deparhnent of A nid Sciences Rutge... more ... JUAN P. ADVIS' AND NORBERT0 A. GUZMAN2f 'Deparhnent of A nid Sciences Rutgers University New Brunnuick, New Jersey 08903 2Protein Research Unit Princeton Biochemicals, Inc. Princeton, New Jersey 08543 ABSTRACT ...
J Liq Chromatogr Relat Techno, 1993
... LUIS HERNANDEZ', JOSE ESCALONA', PHILLIPE VERDEGUER2, AND NORBERT0 A. GUZMAN3* ... more ... LUIS HERNANDEZ', JOSE ESCALONA', PHILLIPE VERDEGUER2, AND NORBERT0 A. GUZMAN3* 'Department of Physwlogv, School of Medicine Los Andes University ... In this experiment, the optimal molar excess to la-bel low concentrations of glutamate was measured. ...
Enzyme, 1984
The physicokinetic and immunologic properties of the purified low and high uptake forms of the hu... more The physicokinetic and immunologic properties of the purified low and high uptake forms of the human lysosomal hydrolase, alpha-L-iduronidase, have been determined and compared. The apparent Km and Vmax values for the low and high uptake forms were similar toward two artificial substrates, 4-methylumbelliferyl-alpha-L-iduronide (0.07 and 0.06 mmol/l; 16.15 and 14.85 mumol/min/mg, respectively), and phenyl-alpha-L-iduronide (1.42 and 1.66 mmol/l; 0.83 and 1.05 mumol/min/mg, respectively), and one natural substrate, anhydro-[3H]-mannitol-iduronide (0.86 and 1.04 mmol/l; 2.50 and 2.79 mumol/min/mg, respectively). The pH optima for both purified forms also were similar for each of the three substrates ( approximately 3.50, approximately 3.50, and approximately 4.50, respectively). Heparin markedly inhibited the 4-methylumbelliferyl-alpha-L-iduronide activities of both the low and high uptake forms, while dermatan sulfate and heparan sulfate were more inhibitory toward the low uptake act...
Organization and Assembly of Plant and Animal Extracellular Matrix, 1990
Journal of Liquid Chromatography, 1995
A method to perform on-line sample preconcentration of serum immunoglobulin E by affinity capture... more A method to perform on-line sample preconcentration of serum immunoglobulin E by affinity capture is described. Purified anti-IgE antibodies were covalently bound to an analyte concentrator-reaction chamber or cartridge. The immunoglobulins (IgE) were bound to and eluted from the cartridge by the optimum dissociating buffer system, and the eluent(s) were then subjected to capillary electrophoresis.The first design used was a
The New England journal of medicine, Jan 12, 1979
Journal of capillary electrophoresis and microchip technology, 2008
Journal of capillary electrophoresis and microchip technology, 2007