Effect of water treatment on analyte and matrix ion yields in matrix-assisted time-of-flight secondary ion mass spectrometry: the case of insulin in and on hydroxycinnamic acid

A systematic study was performed to identify the origin of surprisingly high analyte-to-matrix yield ratios recently observed in time-of-flight secondary ion mass spectrometry (TOF-SIMS) analysis of oligo- and polypeptides mixed in matrices of alpha-cyano-4-hydroxycinnamic acid (4HCCA). Several sets of samples of porcine insulin in 4HCCA (1:3100 molar) were prepared from liquid solutions by a nebuliser technique, with more than one order of magnitude variation in sprayed material (substrate silicon). Following different periods of storage in air and/or vacuum as well as exposure to high-purity water, TOF-SIMS analysis was performed under oblique impact of 22 keV SF5+. Treatment with water involved either deposition of a droplet covering the whole sample for times between 1 and 20 min or spraying with water in droplet equivalent quantities. The analyte and matrix molecules were detected as protonated molecules (insulin also in doubly protonated form). Even the as-prepared samples usually showed insulin-to-4HCCA yield ratios exceeding the molar ratio of the mixed material. Upon ageing in vacuum the matrix ion yields remained constant but the analyte yields decreased, partly due to break-up of intrachain disulfide bonds. Water treatment resulted in a pronounced decrease in the 4HCCA yield, typically by a factor of five, in parallel with an increase of the insulin yield, by up to a factor of four. Evidence is provided that these changes occur concurrently with a partial dissolution of 4HCCA at the sample surface. The enhanced insulin yield was not correlated with the Na+ yield. The typically 20-fold increase in the insulin-to-4HCCA yield ratio, generated by water exposure of the samples, provides the explanation for the high yield ratios observed previously with water-treated samples. Spraying with water or repeated exposure to water droplets caused a pronounced degradation of the insulin parent yields in combination with an increasing appearance of signals due to the B- and A-chains of insulin. To clarify the issue of surface segregation, a few samples were prepared by spraying acetone-diluted solutions of insulin on previously deposited layers of 4HCCA. Whereas the insulin yields from as-prepared samples were rather low, the yields observed after water treatment were comparable with those observed with samples of insulin in 4HCCA. The results suggest that a large amount of insulin is present at the surface of samples prepared from liquid mixtures of insulin in 4HCCA. With both methods of sample preparation, however, high secondary ion yields of insulin were only obtained after exposure of the samples to water. The chemical changes responsible for this beneficial effect still need to be identified.     

Information on measurement uncertainty in test and product standards

According to ISO/IEC 17025, accreditation bodies generally accept that test procedures in normative documents have been validated. This contribution investigates, with respect to measurement uncertainty, the methodical basis of this practice in general and examines specifically two very different groups of products: asphalt, with its major components mineral aggregates and bitumen, and metal wire cloth. The author concludes that both the general and the specific situations are incoherent. The practice of accepting standardised test procedures a priori as validated does not have a sound basis in all cases. Both the concept, and the practical application, of validation or fitness for purpose vary in different product groups. The impact of this situation on the practice of accreditation should be taken into account in order to reduce market distortions in single-product groups. Laboratories face fundamental consequences from this practice.Presented at the Metro Trade Workshop on Traceability and Measurement Uncertainty in Testing, 30–31 January 2003, Berlin, Germany     

Studie zum Thermischen Abbau von PtBr4 an der Thermowaage

Study of the Thermal Decomposition of PtBr₄ on the Thermobalance If the product of a thermal decomposition has a complicated structure, the decomposition (e. g. of PtBr₄) leads only under special conditions — ≧1 atm pressure, raising of temperature small (e. g. 0.2°/min), capillary opening of the container for the effusion of gases — to equilibrium states, which can be utilized thermodynamically.     

Molecular complexes—IV: Effect of additional unspecific shielding (aus). Aus correction of NMR data applied to the 1:1 complex formation between 1,3,5-triacetyl benzene and benzene

Shifts of the two proton NMR signals of 1,3,5-triacetylbenzene (TAB) both relative to external and internal reference have been measured in CCl₄ solutions as function of the donor (C₆D₆) concentrations. A very large range of donor concentrations [D_o]_i yielded plots of Δ_i/[D_o]_i vs Δ_i (simple Scatchard plots) which were distinctly curved. External referencing yielded upward curvatures, internal referencing even stronger downward curvatures.The external referenced shifts were subjected to a linearizing procedure according to the modified Scatchard equation developed from the AUS concept. These independently from one another linearized Scatchard plots (for methyl and for aromatic signal) were exactly parallel, i.e. they gave exactly the same equilibrium quotient K. Performing the linearizing steps in the Cresswell-Allred procedure instead of the Scatchard procedure gave negligible deviations from the modified Scatchard K.     

Gas chromatographic enantiomer separation with modified cyclodextrins: Carboxylic acid esters and epoxides

The wide range of application of 2,6-di-O-methyl-3-O-pentyl-β-and y-cyclodextrins is demonstrated by the resolution of the enantiomers of the methyl esters of chiral carboxylic acids: α and β hydroxy acids with up to 18 carbon atoms, hydroxy di- and tricarboxylic acids, and alkyl/aryl-substituted carboxylic acid methyl esters, including the plant hormone abscisic acid, insect juvenile hormones, and some non-steroidal anti-inflammatory drugs can be resolved with generally large separation factors. The new cyclodextrin derivatives also exhibit high selectivity for epoxides.     

Liquid chromatography/electrospray tandem mass spectrometry characterization of styrene metabolism in man and in rat

Liquid chromatography with electrospray tandem mass spectrometry was used to characterize the metabolism of styrene in man and in rat. To improve identification and characterization of minor styrene metabolites, rats were co-exposed to styrene and styrene-d(8). In addition to the main styrene metabolites, mandelic acid and phenylglyoxylic acid, and specific mercapturic acids, phenylhydroxyethylmercapturic acids (PHEMAs), other minor metabolites, including phenylglycine, N-acetyl-S-(phenacyl)cysteine, 4-vinylphenol and styreneglycol conjugates (glucuronides and sulfates) were identified and determined both in human and rat urine. Phenylglycine and N-acetyl-S-(phenacyl)cysteine have been hypothesized to occur, but never detected in human or rat urine after styrene exposure. 4-Vinylphenol and styrene glycol had already been recognized as styrene metabolites, but never determined as intact glucuronide and sulfate conjugates. Failure to identify 1- and 2-phenylethanol conjugates suggests that phenylethanol might be an intermediate metabolite, but it is not a conjugated catabolite. A method for the simultaneous determination of mandelic acid, phenylglyoxylic acid, phenyglycine and the four PHEMA diastereoisomers has been developed and validated. For those glucuronide and sulfate conjugates whose standards are not commercially available, a method for semiquantitative analysis, based on the use of structurally similar compounds as standards, has been developed. This approach was found to be valid for the determination of 4-vinylphenol glucuronide and 4-vinylphenol sulfate.     

Ligand and solvent effects on cobalt(I)-catalysed reactions: Alkyne dimerisation versus [2+2+2]-cyclotrimerisation versus Diels-Alder reaction versus [4+2+2]-cycloaddition

The cobalt catalysed conversion of phenyl acetylene led to linear enyne dimerisation products when CoBr₂(dppe) was activated with magnesium in the absence of a Lewis acid. In contrast, in the presence of a Lewis acid the cyclotrimerisation process is favoured. Among several ligand systems and solvents tested the best results were obtained using a catalyst system consisting of a diimine cobalt bromide complex, zinc and zinc iodide in acetonitrile. With 2-5 mol% of the cobalt catalyst at ambient temperatures 1,2,4-triphenylbenzene could be obtained in 99% yield and in excellent regioselectivity (95:5) in 10 min reaction time. Competition experiments of phenylacetylene and isoprene were performed. A preference for the cyclotrimerisation reaction was found for the diimine cobalt complex in acetonitrile, while the Diels-Alder reaction is favoured with the cobalt(dppe) complex in dichloromethane. Also a regioselectively substituted cyclooctatriene product was formed in a [4+2+2]-cycloaddition process and isolated which allows assumptions on the reaction mechanism.