Showing papers in "Analytical Chemistry in 1991"

Interactive self-modeling mixture analysis

Abstract: In the analytical environment, spectral data resulting from the analysis of samples often represent mixtures. To extract information about pure components often is a major problem, especially when reference spectra are not available. For this type of problem, self-modeling mixture analysis techniques have been developed. Examples will be shown of Raman spectroscopic data of a rection, where spectra of intermediates are extracted, and of FT-IR microscopic data of a polymer laminate, where it will be shown that spectra of layers below the resolution of the FT-IR microscope can be calculated

Photophysics and photochemistry of oxygen sensors based on luminescent transition-metal complexes

Abstract: A detailed study of the photophysics and photochemistry of polymer-immobilized luminescent transition-metal complex oxygen sensors is presented. Emphasis is on understanding the underlying origin of the nonlinear Stern-Volmer quenching response. Microheterogeneity is important in both photophysical and photochemical behavior, and the nonlinear quenching responses in RTV 118 silicone rubber can be adequately described by a two-site model, although detailed lifetime measurements suggest a more complex Underlying system. Counterion studies with quenching counterions are shown to be useful probes of the structure of the complex in the polymer. While oxygen enhances photochemical instability, singlet oxygen is not directly implicated in sensor decomposition. In the photochemistry there is at least one reactive and one much less reactive site, although the photochemistry and quenching measurements probably sample different populations of sites. The existence of reactive sites suggests that stability can be enhanced by a preliminary photolysis to eliminate the more reactive sites.

Design and in vitro studies of a needle-type glucose sensor for subcutaneous monitoring.

Abstract: A new miniaturized glucose oxidase based needle-type glucose microsensor has been developed for subcutaneous glucose monitoring. The sensor is equivalent in shape and size to a 26-guage needle (0.45-mm o.d.) and can be implanted with ease without any incision. The novel configuration greatly facilitates the deposition of enzyme and polymer films so that sensors with characteristics suitable for in vivo use (upper limit of linear range greater than 15 mM, response time less than 5 min, and sensitivity yielding a 5:1 signal-to-background ratio at normal basal glucose levels) can be prepared in high yield (greater than 60%). The sensor response is largely independent of oxygen tension in the normal physiological range. It also exhibits good selectivity against common interferences except for the exogenous drug acetaminophen.

Electrophoresis in fused-silica capillaries: the influence of organic solvents on the electroosmotic velocity and the .zeta. potential

Abstract: 3390. (44) De Le RkM J. H. Ar-h s ~ b s ~ ~ Nftrm Academic Press Inc.: New York, 1959. (45) mine, H. J. RMCW m n / s m in orgenic ”/sby, A r m & ReanangementS: Monograph 6, Eaborn, C., Chapman, N. E., Eds.: Elsevier Publishing Company: New York. 1967. (46) Mer&, J. ~b~~~ orgenic cmiSw, RMcbbn Mechanism Structure, 3rd. ed.: John Wlley 8 Sons: New York, 1985; p 501. p. D. RECEIVED for review February 11, 1991. Accepted June 7, 1991. The support of the U.S. Environmental Protection Agency (EPA CK-815749-01-0) is acknowledged. The Support provided by the Emil Aaltonen Foundation and Suomen Kulttuurirahasto is greatly appreciated (T.K.).

Structural characterization of protein tryptic peptides via liquid chromatography/mass spectrometry and collision-induced dissociation of their doubly charged molecular ions

Abstract: The formation of multiply charged molecular ions via the field-assisted ion evaporation mechanism during electrospray ionization enables the use of an atmospheric pressure ionization quadrupole mass spectrometer system for characterizing biologically important peptides. The straightforward implementation of high-performance liquid chromatography (HPLC) into this new strategy to determine the molecular weight of tryptic peptides via the pneumatically assisted electrospray (ion spray) interface is presented. Examples utilizing both microbore (1.0 mm) and standard bore (4.6 mm) inside diameter columns are shown for the LC/MS molecular weight determination of tryptic peptides in methionyl-human growth hormone (met-hGH). Injected levels from 50 to 75 pmol of tryptic digest onto 1 mm i.d. HPLC columns provided full-scan LC/MS or LC/MS/MS results without postcolumn splitting of the effluent. When standard 4.6 mm i.d. HPLC columns were used, a 20:1 postcolumn split was utilized, which required from 1 to 5 nmol of injected tryptic digest for full-scan LC/MS or LC/MS/MS results. Collision-induced dissociation (CID) mass spectra resulting from either "infusion" or on-line LC/MS/MS analysis of the abundant doubly charged ions that predominate for tryptic peptides under electrospray conditions provided structurally useful sequence information for met-hGH and human hemoglobin tryptic digests. The slower mass spectrometer scan rate used during infusion of sample provides more accurate mass assignments than on-line LC/MS or LC/MS/MS, but the latter on-line experiments preclude ambiguities caused by matrix or component interferences. However, in some instances very weak CID product ions preclude complete tryptic peptide structural characterization based upon the CID data alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Statistical treatment for rejection of deviant values : critical values of Dixon's Q parameter and related subrange ratios at the 95% confidence level

Abstract: Critical values at the 95% confidence level for the two-tailed Q test, and related tests based upon subrange ratios, for the statistical rejection of outlying data have been interpolated by applying cubic regression analysis to the values originally published by Dixon. Corrections to errors in Dixon's original tables are also included. It is recommended that the newly generated 95% critical values be adopted by analytical chemists as the general standard for the rejection of outlier values

Integrated fluid handling system for biomolecular interaction analysis.

Abstract: An integrated fluid handling system used for multichannel biomolecular interaction analysis is described. Reactions between biological molecules are monitored in real time by measuring changes in the angular position where surface plasmon resonance occurs at a biospecific active surface. The adsorption efficiency of the analyte onto the biospecific active surface is up to approximately 3%, due to the low channel height, 50 microns, in the flow cell. When a large part of the total biospecific active surface for surface plasmon resonance probing (approximately 0.15 mm2) is used, the sensitivity is high. Sample sizes in the order of 1-50 microL can be injected. The sample zone dispersion is minimized by the low dead volume in the system (approximately 0.4 microL) accomplished by using integrated sample loops and thin conduits. An asset of this integration is the low reagent consumption. The sensor chip with the biospecific active surface is reusable and easily exchanged. Experimental results obtained with a theophylline monoclonal antibody as the analyte are compared with a theoretical model. The standard deviation for the repeatability is approximately 5% typically with 50 microL of 250 pM analyte, and the assay time is 10 min. The detection limit is approximately 10 pg of the analyte on the probed spot of the surface. Possible improvements of the sensitivity and detection limit are discussed.

Amperometric glucose microelectrodes prepared through immobilization of glucose oxidase in redox hydrogels.

Abstract: Glucose microelectrodes have been formed with glucose oxidase immobilized in poly[(vinylpyridine)Os(bipyridine)2Cl] derivative-based redox hydrogels on beveled carbon-fiber microdisk (7 microns diameter) electrodes. In the resulting microelectrode, the steady-state glucose electrooxidation current density is 0.3 mA cm-2 and the sensitivity is 20 mA cm-2 M-1. The current density and sensitivity are 10 times higher than in macroelectrodes made with the same hydrogel. Furthermore, the current is less affected by a change in the partial pressure of oxygen. The higher current density and lower oxygen sensitivity point to the efficient collection of electrons through their diffusion in the redox hydrogel to the electrode surface. These results contrast with those observed for enzyme electrodes based on diffusing mediators, where loss of the enzyme-reduced mediator by radial diffusion to the solution decreases the current densities of microelectrodes relative to similar macroelectrodes.

Capillary electrophoretic separations of proteins using nonionic surfactant coatings.

Abstract: Capillary zone electrophoretic separations of proteins have been achieved by using nonionic surfactant coated capillaries. Capillaries were prepared by derivatization of the silica surface with octadecylsilane followed by the deposition of a layer of nonionic surfactant from an aqueous solution above the critical micelle concentration. This coating is of sufficient thickness and hydrophilicity to reduce both protein adsorption and electroosmotic pumping. This hydrophilic coating reduces electroosmotic pumping 5-8-fold while resolving proteins quickly and efficiently with good recovery. The coating provides a stable and reproducible means of deactivation, while the rate of electroosmotic pumping stays relatively constant throughout the pH range 4-11. This allows the pH to be varied to enhance selectivity without adversely affecting the flow rate.

Improvements in the gaseous hydrogen-water equilibration technique for hydrogen isotope ratio analysis

Abstract: Improved precision in the H 2 -H 2 O equilibration method for δD analysis has been achieved in an automated system. Reduction in 1-δ standard deviation of a single mass-spectrometer analysis to 1.3‰ is achieved by (1) bonding catalyst to glass rods and assigning use to specific equilibration chambers to monitor performance of catalyst, (2) improving the apparatus design, and (3) reducing the H 3 + contribution of the mass-spectrometer ion source

Application of (3-mercaptopropyl)trimethoxysilane as a molecular adhesive in the fabrication of vapor-deposited gold electrodes on glass substrates

Abstract: This paper presents fabrication and characterization of durable thin-film gold electrodes prepared by Au vapor deposition on glass substrates pretreated with (3-mercaptopropyl) trimethoxysilane. We characterize the electrodes with regard to (1) the Au adhesion, (2) the voltammetry of Au oxidation, and (3) the self-assembly of octadecyl mercaptan monolayers, and with regard to the extent to which they passive electrode surfaces

Stable carbon isotope analysis of amino acid enantiomers by conventional isotope ratio mass spectrometry and combined gas chromatography/isotope ratio mass spectrometry

Abstract: The application of a combined gas chromatography/isotope ratio mass spectrometry (GC/IRMS) method for stable carbon isotope analysis of amino acid enantiomers is presented. Amino acids require derivatization prior to GC/IRMS analysis. Derivatization of amino acid standards of known stable carbon isotope compositions in conjunction with natural samples, however, permits computation of the original, underivatized amino acid δ 13 C values through use of an empirical correction for the carbon introduced during the derivatization process

Luminescence quenching mechanism for microheterogeneous systems

Abstract: Determining quenching mechanisms for luminescent species adsorbed or bound to a variety of heterogeneous systems (e.g., silicas, organic, inorganic, and biopolymers) is quite difficult In the absence of detailed information on system heterogeneity. A method for assessing the relative contributions of static and dynamic quenching in heterogeneous systems is presented. While the method does not provide direct information on the details of system heterogeneity, it requires no a priori information on the nature of the heterogeneity. This approach is based on a comparison of intensity quenching data with lifetime quenching data using a preexponential weighted lifetime, τ_m. τ_m is calculated by fitting the observed decay curves to a sum of a relatively small number (2-4) of exponentials. For time-correlated single-photon counting the parameters obtained from a statistically acceptable fit can be used to accurately estimate τ_m, even though the computed model may bear no resemblance to the true decay kinetics. Simulations confirm that the method works for a wide range of heterogeneous systems. The technique is applied to oxygen quenching of a luminescent metal complex on a silica surface.

Three DNA sequencing methods using capillary gel electrophoresis and laser-induced fluorescence.

Abstract: Capillary gel electrophoresis is demonstrated for the four-spectral-channel sequencing technique of Smith, the two-spectral-channel sequencing technique of Prober, and the one-spectral-channel sequencing technique of Richardson and Tabor. Sequencing rates up to 1000 bases/h are obtained at electric field strengths of 465 V/cm. At lower electric field strengths, capillary electrophoresis produces useful data for fragments greater than 550 nucleotides in length with 2 times better resolution than slab gel electrophoresis. An on-column detector produces detection limits of 200 zmol (1 zmol = 10(-21) mol = 600 molecules) for the four-spectral-channel technique. A postcolumn detector, based on the sheath flow cuvette, produces detection limits of 20 and 2 zmol for the two- and one-spectral-channel techniques, respectively.

Electrospray and tandem mass spectrometric characterization of acylglycerol mixtures that are dissolved in nonpolar solvents

Abstract: This paper presents electrospray mass spectrometric analysis of mixtures containing monoglycerides, diglycerides, and triglycerides. Sample compounds were dissolved in concentrations of 1-50 pmol/microL in chloroform:methanol (70:30, v:v), which was modified by the addition of alkall-metal or ammonium salts or by addition of formic acid to favor the addition of a cationic species to the sample molecules. Electrospray mass spectrometric analysis of acylglycerol standards yielded positive-ion current signals for (M + Na)+ or (M + NH4)+ of all the species that were present at low picomole per microliter concentrations with no fragmentation. For equimolar concentrations of these sample compounds, there was a general decrease in ion current response as the analyte polarity decreased. Therefore, acylglycerols that contained unsaturated fatty acid chains were observed to exhibit a response in the mass spectrum greater than those with saturated chains, and ion signals resulting from the molecular adduct ions of monoglycerides were more abundant than those of diglycerides, which were more abundant than those of triglycerides in the mass spectrum. Electrospray mass spectrometric analysis of an unknown lipid material recovered from a mammalian cell culture reactor revealed a mixture of triglycerides containing mostly C14, C16, and C18 fatty acids with varying degrees of unsaturation. The results obtained by electrospray mass spectrometry compared favorably to those obtained by gas chromatography after saponification and methylation of fatty acid components of the triglycerides. MS/MS fragmentation of sodiated acylglycerols required a dissociation energy significantly greater than that required for fragmentation of ammoniated acylglycerols, so MS/MS characterization of acylglycerols was generally performed on the ammoniated compounds.(ABSTRACT TRUNCATED AT 250 WORDS)

Integration of mass spectrometry in analytical biotechnology.

Abstract: Mass spectrometry (MS) has become an indispensable tool for peptide and protein structure analysis because of three unique capabilities that enable it to be used to solve structural problems not easily handled by conventional techniques. First, MS is able to provide accurate molecular weight information on low-picomole amounts of peptides and proteins independent of covalent modifications that may be present. Second, this information is obtainable for peptides present in complex mixtures such as those that result from a proteolytic digest of a protein. Third, by using tandem MS, partial to complete sequence information may be obtained for peptides containing up to 25 amino acid residues, even if the peptides are present in mixtures. Sensitivity and speed of the MS-based approaches now equal (and in some cases exceed) that of Edman-based sequence analysis. In this perspective we discuss how MS, tandem high-performance MS, and on-line liquid chromatography/MS using fast atom bombardment or electrospray ionization have been integrated with more conventional techniques in order to increase the accuracy and speed of peptide and protein structure characterization. The expanding role of matrix-assisted laser desorption MS in protein analysis is also described. The unique niche that MS occupies for locating and structurally characterizing posttranslational modifications of proteins is emphasized. Examples chosen from the authors' laboratory illustrate how MS is used to sequence blocked proteins, define N- and C-terminal sequence heterogeneity, locate and correct errors in DNA- and cDNA-deduced protein sequences, identify sites of deamidation, isoaspartyl formation, phosphorylation, oxidation, disulfide bond formation, and glycosylation, and define the structural class of carbohydrate at specific attachment sites in glycoproteins.

Lifetime of neutral-carrier-based liquid membranes in aqueous samples and blood and the lipophilicity of membrane components.

Abstract: On the basis of previously reported correlations between the lipophilicity of membrane components, their partition coefficient between the membrane and the sample, and the lifetime of corresponding neutral-carrier-based sensors, the lipophilicities of ionophores and plasticizers in analytically relevant ion-selective electrodes, ISFETs, and optodes are analyzed and reported. Equations for the estimation of the lifetimes of liquid membranes in continuous-flow systems are presented, and the experimental determination of the lipophilicity values by thin-layer chromatography (TLC) is described. The required lipophilicities for the lifetimes of liquid membranes over 30 24-h days for different applications in aqueous solutions as well as in blood are presented. A comparison of the experimental results of lifetime measurements with calculated theoretical values is given. The experimental results of the determination of the lipophilicity by TLC are compared with the lipophilicities estimated on the basis of Hansch parameters.