Biomedical subjects
Publications and source records attributed to Isiah M Warner.
The determination of cysteine and homocysteine levels is of great current interest for the monitoring of desease states. A new colorimetric method for the simultaneous detection of l-cysteine and l-homocysteine has been developed. A fluorescein derivative reacts with the above amino acids, producing their respective thiazolidines resulting in color changes. Interference from other amino acids and proteins is minimal.
The effect of temperature on the chiral recognition of binaphthyl derivatives in the presence of poly sodium N-undecanoyl-LL-leucyl-leucinate (poly LL-SULL) is examined using electrokinetic chromatography (EKC) and steady-state fluorescence anisotropy. An examination of the effect of temperature suggests that the chiral recognition of 1,1'-binaphthyl-2,2'-diol enantiomers improves with increasing temperature, whereas lower temperatures resulted in better enantiosolectivity in the case of 1,1'-binaphthyl-2,2'-diyl hydrogen phosphate enantiomers. In addition, steady-state fluorescence anisotropy results show that the anisotropy of the two enantiomers are different when complexed to poly-(LL) SULL. As would be expected, the enantiomer that binds stronger to the chiral pseudostationary phase, as evidenced by EKC experiments, had higher anisotropy values. The results of this study suggest that steady-state fluorescence anisotropy can be used to gain further insight into chiral recognition.
One disadvantage of amino acid-based chiral selectors for micellar electrokinetic chromatography (MEKC) is that either they have very low solubility or are insoluble at acidic pHs. In order to increase solubilities at lower pHs, we have synthesized a highly water-soluble achiral surfactant and copolymerized it with an amino acid-based chiral surfactant. These two surfactants were polymerized either separately or at various molar rations of binary solutions, yielding pure molecular or copolymerized surfactant (CoPS), respectively. All surfactants were characterized by use of several analytical techniques prior to using them as novel pseudostationary phases in MEKC. The chromatographic performance of the CoPS in MEKC was tested with chiral and achiral analytes. The highly soluble sulfate head group significantly increased the solubility of amino acid-based CoPS over a wide range of pH. Three chiral binaphthyl derivatives were tested and each surfactant system was found to have different selectivity.
We describe the use of the polymeric surfactant poly(sodium undecylenic sulfate) (poly-SUS) as a stationary phase coating in open-tubular capillary electrochromatography (OT-CEC) coupled with electrospray ionization-mass spectrometry (ESI-MS) for the analysis of beta-blocker and benzodiazepine analytes. The production of a polymeric surfactant coating on the capillary inner wall involves (i) adsorption of the cationic polymer poly(diallyldimethylammonium chloride) (PDADMAC) to the inner surface of capillary, and (ii) adsorption of the negatively charged poly-SUS onto the cationic polymer layer via strong physical interaction of the two polymer layers. As compared with micellar electrokinetic chromatography (MEKC) coupled with ESI-MS, the main advantage of this proposed method is minimization of introduction of the monomeric or polymeric surfactant into the mass spectrometer, thus avoiding the interference of the nonvolatile micelle in ESI-MS. The effects of buffer pH and applied voltage on the separation of the analytes are also discussed. Under optimum conditions, four of the five beta-blockers and four benzodiazepines are separated.
A cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) method was applied to the enantioseparation of three binaphthyl derivatives using neutral CDs (i.e., beta- and gamma-CD) in combination with various chiral amino acid-based polymeric surfactants (PSs). Both the D- and L-configurations of poly(sodium N-undecanoyl alaninate), poly(sodium N-undecanoyl leucinate), and poly(sodium N-undecanoyl valinate) (poly(L-SUV)) were synthesized. The retention behavior of the three binaphthyl derivatives under optimum electrophoretic conditions using a single chiral additive (PS or CD) is discussed. In addition, the effect of CD cavity size and stereochemical configuration of polymeric surfactants on selectivity (alpha) and resolution (Rs) was investigated. The enantioseparation of (+/-)1,1'-binaphthyl-2,2'-diamine gave a reversal of enantiomeric order when using beta-CD in combination with any of the three D-configuration PS. However, better enantioseparation is obtained when using the corresponding L-configuration PS with beta-CD. A reversal of migration order (RMO) for the enantiomers of (+/-)1,1'-bi-2-naphthol was observed upon the addition of 10 mM gamma-CD to poly(L-SUV). However, no RMO of (+/-)1,1'-bi-2-naphthol was seen when either beta-CD or gamma-CD was combined with D-PS. The enantiomers of (+/-)1,1'-binaphthyl-2,2'-diyl hydrogen phosphate showed little enantioselective behavior toward the PS alone. However, combined D- or L-PS and beta-CD or gamma-CD systems gave increased Rs and alpha values. The chiral recognition of binaphthyl derivatives observed resulting from the various combinations of two chiral selectors is discussed.
The effectiveness of Triton X-102 (TX-102), as a structural modifier of the polymeric surfactant sodium undecanoyl-L-leucinate (L-SUL) was investigated for enhanced enantiomeric recognition of various chiral compounds in micellar electrokinetic chromatography (MEKC). Increasing concentrations of TX-102 were separately added into the micellar solutions of L-SUL and then polymerized to form poly-L-SUL. The resulting polymers were purified by use of 3500 molecular-weight-cutoff (MWCO) dialysis membranes. Fluorescence and pulsed field gradient-nuclear magnetic resonance (PFG-NMR) techniques were used to elucidate the structural effects of TX-102 on poly-L-SUL. Evaluation of data from fluorescence measurements suggested an increase in polarity with increasing concentration of TX-102. However, the polarity decreased at higher concentrations of TX-102. Evaluation of data from PFG-NMR suggested an increase in hydrodynamic radius upon increasing the concentration of TX-102. The racemates of coumarinic and phenythiohydantoin amino acid derivatives, and pindolol were used as test analytes in MEKC. A notable increase in resolution and capacity factors of the test analytes was observed when the modified poly-L-SUL was used in MEKC measurements. Examination of the data obtained from fluorescence, PFG-NMR, and MEKC suggests a strong correlation between the polarity and the hydrodynamic radii of TX-102 modified micelles and the enantiomeric resolution of the test analytes.
The detection of saccharides in biological media is of great current importance for the monitoring of disease states. We have previously reported that solutions of boronic acid-functionalized macrocycles form acyclic oligomeric materials in situ. The oligomers contain fluorescent xanthene moieties. Current efforts are aimed at modulating the spectroscopic responses of these materials for the analysis of specific sugars. We describe conditions whereby the xanthene boronic acids exhibit high colorimetric fructose selectivity. In contrast, at physiological levels selective glucose monitoring can be achieved via fluorescence. Additionally, we describe a method which exhibits promise for detecting both glucose and fructose at dual wavelengths in the UV-Vis region. Mechanistic rationale for each of these findings is presented.
As part of an ongoing study on atherosclerotic arteries, the concentrations of 12 elements in a native and by-pass human coronary artery plaque deposits from five human hearts were determined using inductively coupled plasma-mass spectrometry (ICP-MS). These elements were Ca, P, Na, K, Mg, Zn, Cu, Pb, Fe, Al, Si, and S. For Zn, Ca, Pb, Fe, Al, and Si, the levels were at the fractional micromol/g levels and they probably played an unimportant role in plaque development. Sulfur levels varied from 23 to 140 micromols indicative of the possible presence of homocysteine, but there appeared to be no consistent relationship between by-pass and native concentrations. The calcium and phosphorus concentrations were relatively high in all cases, but the ratio of their molecular concentrations did not correspond to hydroxyapatite, which is conventionally considered to be the chemical form of calcium in heart plaque. In the mature native plaque, high calcium/phosphorus ratios indicated calcium in chemical forms other than hydroxyapatite. In undeveloped by-pass plaque, the phosphorus concentration was too high to be as hydroxyapatite but may be phospholipids. Because it is difficult to get suitable samples, only five heart samples were available. Therefore, these results should be treated as preliminary.
In this study, we report the use of ionic liquids as modifiers in the separation of achiral and chiral analytes in micellar electrokinetic chromatography. In this investigation, polymeric surfactants and ionic liquids were added to a low-conducting buffer solution. The polymeric surfactants used in this study were poly(sodium N-undecylelinic sulfate) and poly(sodium oleyl-l-leucylvalinate). The ionic liquids used in this study were chosen because of their high conductivity, hydrophobicity, and good solvating properties. Thus, it was expected that these ionic liquids would have the ability to assist in the separation of hydrophobic mixtures while maintaining adequate background current. Three analyte mixtures were separated using various buffer combinations of polymeric surfactant and ionic liquids. The ionic liquids were shown to improve the resolution and peak efficiency of the analytes while maintaining adequate background current.
In this study, fused-silica capillaries are modified using a polyelectrolyte multilayer (PEM) coating procedure in open-tubular capillary electrochromatography. The PEM coating was constructed in situ with alternating rinses of positively and negatively charged polymers. The quaternary ammonium salt poly (diallyldimethylammonium chloride) was used as the cationic polymer, and the polymeric surfactant poly (sodium N-undecanoyl-l-leucylvalinate) was used as the anionic polymer. Previous studies have shown that the PEM-coated capillaries used for achiral separations have excellent reproducibilities and high stabilities against extreme pH values. In the current study, this PEM coating approach was applied to chiral separations of 1,1'-binaphthyl-2,2'-dihydrogenphosphate (BNP), 1,1'-bi-2-naphthol, secobarbital, pentobarbital, and temazepam. However, the PEM coating procedure used in the achiral studies needed to be significantly modified in order to achieve chiral separations. Optimal conditions were established by varying the additives (sodium chloride, 1-ethyl-3-methyl-1H-imidazolium hexafluorophosphate, 1-butyl-3-methylimidazolium tetrafluoroborate) in the polymer deposition solutions, the salt concentration, the column temperature, and the bilayer number. Reproducibilities were evaluated by use of the relative standard deviation (RSD) values of the electroosmotic flow (EOF) and the first peak ((R)-(+)-BNP). In all cases, the run-to-run and capillary-to-capillary RSD values of EOF were less than 0.5%, and the run-to-run RSD values of the (R)-(+)-BNP peak were less than 1%. In addition, more than 230 runs were performed on a single PEM-coated capillary.
Dipeptide micelle polymers are a new class of polymeric surfactants of which the polysodium undecanoyl-L-leucylvalinate (poly-L-SULV) was found to be a broadly applicable chiral selector for micellar electrokinetic chromatography. This negatively charged dipeptide micelle polymer is a high molecular weight compound with large countercurrent mobility, zero critical micelle concentration, low aggregation number, and high solubility in water or water-organic solvents. In an extensive chiral screening program, enantioseparation of 75 racemic compounds was tested with poly-L-SULV as chiral pseudostationary phase in neutral pH and basic pH background electrolytes. A total of 58 out of 75 racemic compounds could be resolved after choosing an appropriate concentration of poly-L-SULV. Although anionic chiral analytes are difficult to resolve using poly-L-SULV, the percent success rate for chiral resolution of cationic (77%) and neutral (85%) racemates was very high. Aspects regarding electrostatic, steric, hydrophobic, and hydrogen-bonding interactions of this dipeptide micelle polymer with various classes of chiral analytes are discussed.
A novel method of modifying sodium undecanoyl-L-leucinate (SUL) micelles employed in chiral separation of analytes in micellar electrokinetic chromatography (MEKC) to enhance selectivity toward specific analytes is discussed. The current study aimed at modifying the SUL micelles by introducing different alcohols into the mono-SUL micelles. The micellar solutions were then polymerized in the presence of alcohols followed by postpolymerization extraction of the alcohols to yield alcohol-free polymeric surfactants (poly-L-SUL). The effects of hexanol (C(6)OH) and undecylenyl alcohol (C(11)OH) on micellar properties of this surfactant were investigated by use of surface tensiometry, fluorescence spectroscopy, pulsed field gradient-nuclear magnetic resonance (PFG-NMR), and MEKC. The surface tension and PFG-NMR studies indicated an increase in the critical micelle concentration (cmc) and micellar size upon increasing the alcohol concentration. Fluorescence measurements suggested that alcohols induce closely packed micellar structures. Coumarinic and benzoin derivatives, as well as (+/-)-1, 1'-binaphthyl-2,2'-dihydrogen phosphate (BNP) were used as test analytes for MEKC experiments. Examination of MEKC data showed remarkable resolutions and capacity factors of coumarinic derivatives obtained with modified poly-L-SUL as compared to the unmodified poly-L-SUL. Evaluation of fluorescence, PFG-NMR, and MEKC data suggest a strong correlation between the polarity and hydrodynamic radii of alcohol-modified micelles and the resolution of the test analytes.
A chiral amino acid-based monomeric and polymeric surfactant, sodium oleyl-L-leucylvalinate) (L-SOLV) and poly(sodium oleyl-L-leucylvalinate) (poly-L-SOLV) were synthesized and used for chiral separations in micellar electrokinetic chromatography (MEKC). Poly-L-SOLV was used successfully in the separation of various enantiomers of neutral, acidic, and basic analytes such as 1,1'-bi-2-napthol, 1,1'-binaphthyl-2,2'-diamine, benzoin, hydrobenzoin, benzoin methylether, warfarin, and coumachlor obtaining well-resolved peaks but with only partial separation of temazepam. In addition, the atropisomer 1,1'-binaphthyl-2, 2'-dihydrogen phosphate was chosen to study the applicability of the polymeric surfactant over a wide range of parameters such as concentration, temperature, voltage, and pH. The most striking characteristic of this new surfactant is its high hydrophobicity. It is favorable to interactions with hydrophobic chiral analytes, and thus may provide better chiral recognition for the compounds.
An achiral monomeric surfactant (sodium 10-undecenyl sulfate, SUS) and a chiral surfactant (sodium 10-undecenoyl L-leucinate, SUL) were synthesized and polymerized individually to form poly-SUS and poly-SUL. These surfactants were then copolymerized at various molar ratios to produce a variety of copolymerized surfactants (CoPSs), possessing both achiral (sulfate) and chiral (leucinate) head groups. The CoPSs, poly-SUS, poly-SUL, and sodium dodecyl sulfate were characterized using several analytical techniques. The aggregation numbers of the polymeric surfactants and the partial specific volumes were determined by the use of fluorescence quenching and density measurements, respectively. These polymeric surfactants were investigated as novel pseudostationary phases in micellar electrokinetic chromatography (MEKC) for the separation of chiral and achiral solutes. Solute hydrophobicity was found to have major influence on the MEKC retention of alkyl phenyl ketones. In contrast, hydrogen-bonding ability of benzodiazepines is the major factor that governs their retention, but hydrophobicity has an insignificant effect on MEKC retention of benzodiazepines.
This review represents a summary of recent progress in open-tubular capillary electrochromatography (OT-CEC) for chiral and achiral separations. The OT-CEC approach is an alternative to packed-CEC that could eliminate the problems associated with retaining frits and silica particles. In OT-CEC, the stationary phase is immobilized on the inner walls of the capillary. Preparation of the stationary phase is critical for OT-CEC. The preparation methods for capillary columns include (i) adsorption, (ii) covalent bonding and/or cross-linking, (iii) porous layers, (iv) chemical bonding after etching, (v) sol-gel, and (vi) molecular imprinting. Major developments, potential applications, technical difficulties and advantages associated with these wall coatings in OT-CEC are presented. In addition, the coupling of OT-CEC with mass spectrometry (MS) is briefly reviewed. Several applications of this hyphenated technique for analytical separations are also summarized.
This study examines the role of steric hindrance near the stereogenic centers of four glutamic acid based polymeric surfactants. The single amino acid surfactants of glutamic acid, glutamic acid methyl ester, glutamic ethyl ester, and glutamic acid tert-butyl ester were investigated. The micellar electrokinetic chromatography (MEKC) separation of three binaphthyl derivatives and three benzodiazepines were used to study these steric factors. In addition, the hydrophobicity of these polymers as a function of pH was investigated by use of fluorescence measurements.
A stable polyelectrolyte multilayer (PEM) coating was investigated for use in open-tubular capillary electrochromatography (o-CEC). In this approach, the PEM consisted of the cationic polymer of a quaternary ammonium salt, poly(diallyldimethylammonium chloride) and the anionic polymeric surfactant, poly(sodium undecylenic sulfate). Both the cationic and anionic polymers were physically adsorbed to the surface of a fused-silica capillary by use of a simple coating procedure. This procedure involved an alternate rinse of the positively and negatively charged polymers. The performance of the PEM coating as a dynamic stationary phase was evaluated by use of electrochromatographic experiments and showed good selectivity for both phenols and benzodiazepines. Reproducibility of the PEM coating was also evaluated by calculating the relative standard deviations (RSDs) of the electroosomotic flow (EOF). The run-to-run and capillary-to-capillary RSD values of the EOF were less than 1.5%. The endurance of the coating was more than 100 runs. The importance of the PEM coating was illustrated by comparing separations on a bare uncoated capillary with the coated capillary. In addition, the chromatographic performance using o-CEC and micellar electrokinetic chromatography (MEKC) was compared for the separation of benzodiazepines.
Human atherosclerotic plaques in both native and bypass arteries have been visualized using microtomography to provide additional information on the nature of coronary artery disease. Plaques contained within arteries removed from three white males aged 51, 55 and 70 are imaged in three-dimensions with monochromatic synchrotron x-ray radiation. Fields of view are 658 x 658 x 517 voxels. with cubic voxels ranging from 12 to 13 microm on a side. X-ray energies range from 11 to 15 keV (bandpass approximately 10 eV). At lower energies, high local absorption tends to generate reconstruction artefacts, while at higher energies the arterial wall is scarcely visible. At all energies, calcifications are clearly visible and differences are observed between plaques in native arteries (lifetime accumulations) versus bypass arteries (plaques developing in the interval between the heart bypass operation and the autopsy). In order to characterize coronary calcification, a microfocused, 50 microm2, 25 keV x-ray beam was used to acquire powder diffraction data from selected calcifications. Also, large calcifications were removed from the native arteries and imaged with 25 keV x-ray energy. Calcifications are composed of hydroxyapatite crystallites and an amorphous phase. In summary, native calcifications are larger and have a higher fraction of hydroxyapatite than calcifications from the bypass arteries.