Extending the use of certified reference sediments for assessment of accuracy in determinations of trace metals

Straightforward analysis for components in a single certified reference sediment is of limited use for assessing the accuracy of environmental determinations. A systematic approach requires mixing of certified sediments, one with another and with environmental samples, and the preparation of secondary reference material by the laboratory. Use of Youden pairs, reference material embedded in samples and linear models should enable valid accuracy statements to be made based on well known statistical concepts. For assessing accuracy, reference sediments which are matched in particle size, and are end-members for components or sediment types are most useful.     

A new photochemically removable protecting group for amines

A new arylmethylsulfonyl chloride reacts with secondary or primary amines to give sulfonamides which can be photochemically cleaved, making it suitable for use as a photoremovable protecting group.     

Investigation of parameters affecting the on-line combination of supercritical fluid extraction with capillary gas chromatography

Two different injectors, a split/splitless injector and a programmed temperature vaporizer (PTV) injector were investigated as the interface in on-line supercritical fluid extraction (SFE)-capillary gas chromatography (cGC). The parameters affecting the chromatographic peak shapes as well as the quantitative performance of the interfaces in on-line SFE-cGC were identified and studied. Particular attention was paid to the case where modified extraction fluids were used. Experiments were performed on two different samples. The first sample consisted of PAHs spiked on sand at different concentration levels. The other sample was a polymeric material.     

General deoxyribozyme-catalyzed synthesis of native 3'-5' RNA linkages

An elusive goal for nucleic acid enzymology has been deoxyribozymes that ligate RNA rapidly, sequence-generally, with formation of native 3'-5' linkages, and in preparatively useful yield. Using in vitro selection, we have identified Mg2+- and Zn2+-dependent deoxyribozymes that simultaneously fulfill all four of these criteria. The new deoxyribozymes operate under practical incubation conditions and have modest RNA substrate sequence requirements, specifically D downward arrowRA for 9DB1 and A downward arrowR for 7DE5 (D = A, G, or U; R = A or G). These requirements are comparable to those of deoxyribozymes such as 10-23 and 8-17, which are already widely used as biochemical tools for RNA cleavage. We anticipate that the 9DB1 and 7DE5 deoxyribozymes will find immediate practical application for RNA ligation.     

Decarboxylative aldol reactions of allyl beta-keto esters via heterobimetallic catalysis

Mild and selective heterobimetallic-catalyzed decarboxylative aldol reactions involving allyl beta-keto esters have been developed. The reaction is promoted by Pd(0)- and Yb(III)-DIOP complexes at room temperature and involves the in situ formation of a ketone enolate from allyl beta-keto esters followed by addition of the enolate to aldehydes. The reaction is a new example of heterobimetallic catalysis in which the optimized reaction conditions require the addition of both metals.     

Open-tubular capillary electrochromatography/electrospray ionization-mass spectrometry using polymeric surfactant as a stationary phase coating

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.     

Average peptide score: a useful parameter for identification of proteins derived from database searches of liquid chromatography/tandem mass spectrometry data

The quantity and variable quality of data that can be generated from liquid chromatography (LC)/mass spectrometry (MS)-based proteomics analyses creates many challenges in interpreting the spectra in terms of the actual proteins in a complex sample. In spite of improvements in algorithms that match putative peptide sequences to MS/MS spectra, the assembly of these lists of possible or probable peptides into a 'correct' set of proteins is still problematic. We have observed a trend in a simple relationship, derived from standard database search outputs, which can be useful in assessing the quality of a MS/MS-based protein identification. Specifically, the ratio of the protein score and number of non-redundant peptides, or average peptide score (APS), can facilitate initial filtering of database search results in addition to providing a useful measure of confidence for the proteins identified. This parameter has been applied to results from the analysis of multi-protein complexes derived from pull-down experiments analyzed using a two-dimensional LC/MS/MS workflow. In particular, the complex list of protein identifications derived from a drug affinity pull-down with immobilized ampicillin and an E. coli lysate was greatly simplified by applying the APS as a filter, allowing for facile identification of the penicillin-binding proteins known to interact with ampicillin. Furthermore, an APS threshold can be used for any data sets derived from electrospray ionization (ESI)- or matrix-assisted laser desorption/ionization (MALDI)-MS/MS experiments and is also not specific to any database search program.     

(18)O]-oxygen incorporation reveals novel pathways in spiroacetal biosynthesis by Bactrocera cacuminata and B. cucumis

The origins of the oxygen atoms in 1,7-dioxaspiro[5.5]undecane (1) and hydroxyspiroacetal (2) from Bactrocera cacuminata, and in 2,8-dimethyl-1,7-dioxaspiro[5.5]undecane (3) and hydroxyspiroacetal (4) from B. cucumis, have been investigated by incorporation studies from both [(18)O(2)]-dioxygen and [(18)O]-water. Combined GC-MS examination and high-field NMR analysis have demonstrated that all oxygen atoms in 1 and 2 from B. cacuminata are dioxygen derived, but in contrast, the spiroacetals 3 and 4 from B. cucumis incorporate one ring oxygen from water and one ring oxygen (and the hydroxyl oxygen in 4) from [(18)O(2)]-dioxygen. These results reveal not only the generality of monoxygenase mediation of spiroacetal formation in Bactrocera sp., but also an unexpected complexity in their biosynthesis. A general paradigm accommodating these and other observations is presented.     

Anatoxins and degradation products, determined using hybrid quadrupole time-of-flight and quadrupole ion-trap mass spectrometry: forensic investigations of cyanobacterial neurotoxin poisoning

The potent neurotoxins from cyanobacteria, anatoxin-a (AN), its methyl analogue, homoanatoxin-a (HMAN), and their degradation products, have been studied using nano-electrospray hybrid quadrupole time-of-flight mass spectrometry (QqTOF-MS). The anatoxin degradation products, which are readily produced in vivo by either reduction or epoxidation, were also examined in this study. The high mass accuracy QqTOF-MS data was used to confirm formula assignments for major product ions and quadrupole ion-trap (QIT)-MS was used to construct fragmentation pathways for anatoxins. Significant differences between these fragmentation pathways were observed. Comparisons between the spectra of compounds that differ in side-chain length (the AN and HMAN series) were used to identify ions that are characteristic of the homologues. The application to forensic samples in which the principal neurotoxin had undergone rapid biodegradation has been demonstrated and used to confirm anatoxin poisoning of dogs.     

Viability of the antigen determines whether DNA or urocanic acid act as initiator molecules for UV-induced suppression of delayed-type hypersensitivity

UV radiation suppresses the immune response, and UV-induced immune suppression contributes to UV-induced photocarcinogenesis. For UV-induced immune suppression to occur, electromagnetic energy (i.e. UV radiation) must be converted to a biological signal. Two photoreceptors have been identified in the skin that serves this purpose, epidermal DNA and trans-urocanic acid (UCA). Although compelling evidence exists to support a role for each pathway (UV-induced DNA damage or photoisomerization of UCA) in UV-induced immune suppression, it is not clear what determines which photoreceptor pathway is activated. To address this question, we injected UV-irradiated mice with a monoclonal antibody with specificity for cis-UCA or applied liposomes containing DNA repair enzymes to the skin of UV-irradiated mice. The effect that each had on UV-induced suppression of delayed-type hypersensitivity was measured. We asked whether the light source used (FS-40 sunlamps vs solar-simulated UV radiation) altered whichever pathway of immune suppression was activated. Different doses of UV radiation and the viability of the antigen were also considered. Neither the dose of UV nor the light source had any influence on determining which pathway was activated. Rather, we found that the viability of the antigen was the critical determinant. When live antigens were used, UV-induced immune suppression was blocked with monoclonal anti-cis-UCA but not with T4 endonuclease V-containing liposomes. The reverse was observed when formalin-fixed or killed antigens were used. Our findings indicate that antigen viability dictates which photoreceptor pathway predominates after UV exposure.