ABSTRACT
An automated sample preparation for high throughput accurate mass determinations by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) has been developed. Sample preparation was performed with an automated workstation and automated mass analyses were performed with a commercial MALDI-TOF mass spectrometer. The method was tested with a 41-sample library. MALDI-TOFMS was found to give the needed sensitivity, accurate mass measurement, and soft ionization necessary for structure confirmation, even of mixtures. A mass accuracy of 5 ppm or less was obtained in over 80% of known compound measurements. A mass accuracy better than 10 ppm was obtained for all measurements of known compounds. Analyses of parallel synthesis products resulted in 77% of the measurements with a mass accuracy of 5 ppm or better.
Subject(s)
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods , Automation , Reproducibility of Results , Sensitivity and Specificity , Specimen Handling , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/instrumentation , Time FactorsABSTRACT
Size exclusion chromatography (SEC) isolation of affinity-selected ligands combined with reverse phase liquid chromatography-mass spectrometry (LC-MS) is an effective means for identifying members of mixtures which form tightly bound noncovalent complexes with target proteins. A potential liability of the approach is that the SEC isolation is carried out under nonequilibrium conditions favoring protein/ligand complex dissociation. At long SEC isolation times and/or for complexes with fast off-rates the extent of dissociation can jeopardize the ability to detect the affinity-selected components. Additionally, equilibrium binding affinities cannot be exactly determined from the measured distribution of isolated ligands. We present here an online SEC/LC-MS system for determining affinity-selected members of active mixtures which reduces this liability. A kinetic model of the SEC isolation process is developed to determine the practical limits for the application of the method and to extrapolate equilibrium binding affinities from the nonequilibrium data. The utility of online SEC/LC-MS for identifying affinity-selected ligands and for estimating binding affinities is demonstrated for a small molecule mixture of compounds with known binding affinities and for a simple combinatorial mixture.
Subject(s)
Chromatography, Liquid/methods , Combinatorial Chemistry Techniques , Ligands , Mass Spectrometry/methods , Chemical Fractionation , Matrix Metalloproteinase 3/metabolismABSTRACT
Molecular orbital spectral predictions suggest that 2,5,7,10,12,15,17,20-octaaza-21H, 23H-porphine has a visible spectral range closely matching that of chlorophyll-a. Since the octaazaporphine is, in its core, a simple derivative of an (HCN)12 oligomer, this fact, together with its spectral properties, would suggest that it occupies a high rank as a primordial porphinic solar energy transducer for photochemistry essential to life's formation. The demonstration that the mass 324 hexahydrooctaazaporphine is formed in protic media by the cyclotetramerization of imidazol-4-aminohydroxonium ion or the derived nitrenium ion, and that a mass 318 species consonant with that of the Huckel aromatic octaazaporphine is observed in the course of these studies, strongly supports the proposed octaazaporphine synthesis in a prebiotic hydrocyanic acid milieu.
Subject(s)
Hydrogen Cyanide/chemistry , Hydrogen Cyanide/radiation effects , Nitroimidazoles/chemical synthesis , Origin of Life , Porphyrins/chemical synthesis , Sunlight , Chlorophyll/chemistry , Chlorophyll A , Ions , Nitrogen Compounds/chemistry , Photochemistry , Spectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationABSTRACT
Matrix-assisted laser desorption-ionization (MALDI) time of flight is shown to give a molar peak area response for isolated methylmethacrylate oligomers that have 25 and 50 repeat units when run on three different instruments in reflectron or linear mode and using three different matrix materials. In addition, fragmentation was not observed in any of the three different matrices or at higher laser power. No spectral differences were observed for syndiotactic and isotactic methylmethacrylate oligomers. These results suggest that the low most probable peak values observed for narrow distribution poly(methylmethacrylate) standards by MALDI mass spectrometry are not the result of mass discrimination or fragmentation.
ABSTRACT
Electrospray ionization (ESI) is capable of ionizing many soluble polymers. The ESI spectra are complex because of overlap of the multiply charged ions of the oligomer distribution, causing current computer transform programs to fail. However, it is possible to determine the origin of the multiply charged ions, making it feasible to write a program designed to transform ESI polymer spectra. To assess the value of such a program for polymer analysis, isolated monodisperse methyl methacrylate (MMA) oligomers (25 and 50 repeat units) were used to determine molar signal response and propensity for fragmentation.The sum of the peak areas for the multiply charged MMA 50-mer was found to be only about 66% of the summed peak areas for the 25-mer for the same molar concentration. However, conversion of the multiply charged peak areas to the singly charged representations, with peak area compression taken into account, gave equal signal responses for the 25-and 50-mers. Signal response variations due to the tacticity of the MMA oligomers were not observed. Fragmentation of the MMA oligomers also was shown not to occur under normal ESI conditions. Therefore, transformation of the polymer spectra to the singly charged molecular ion distribution should allow accurate calculation of average molecular weights, polydispersity, end group mass, and repeat unit mass.
ABSTRACT
Perfluoroalkylated nanospheres have been prepared by reaction of fullerenes with a variety of fluoroalkyl radicals. The latter are generated by thermal or photochemical decomposition of fluoroalkyl iodides or fluorodiacyl peroxides. Up to 16 radicals add to C(60) to afford easily isolable fluoroalkylated derivatives. The monosubstituted radical adducts were detected by electron spin resonance in the early stages of the fluoroalkylation reactions. These spheroidal molecules are thermally quite stable, soluble in fluoroorganic solvents, chemically resistant to corrosive aqueous solutions, and more volatile than the parent fullerenes. Films of the sublimed material display properties typical for a perfluoroalkylated material.
ABSTRACT
Electro spray mass spectra of carbonic anhydrase (MW â¼ 29000) and ovalbumin (MW â¼ 45000) were obtained on a double focusing magnetic secter mass spectrometer by using a single stage of mechanical pumping in the interface between atmospheric pressure and high vacuum. Full scan spectra of lysozyme were recorded on 15 fmoles consumed. In addition, accurate mass measurement was demonstrated for peptides and proteins, and resolution in excess of 10,000 (m /â³m, 10% valley) was observed. These results clearly show that high performance magnetic sector mass spectrometers can be advantageously interfaced to an atmospheric pressure electrospray ion source.