Untersuchungen zur biotischen Bildung von Methylquecksilber aus anorganischem Quecksilber durch intestinale Sulfat-reduzierende Bakterien und die Bestimmung alkylierter Quecksilberverbindungen mittels GC-ICP-MS und Isotopenverdünnungsanalyse

Alkylierte Quecksilberspezies sind hundertfach toxischer als anorganisches Quecksilber (Hg) und werden in der Nahrungskette mit zunehmender Trophieebene im Gewebe von Tieren und dem Menschen akkumuliert. Aufgrund der Relevanz für die Umwelt und den Effekt auf die menschliche Gesundheit kommt der biotischen Transformation von anorganischem Hg zu Monomethylquecksilber (MeHg) eine große Bedeutung zu. Es ist bekannt, dass Sulfat-reduzierende Bakterien zu den Hauptproduzenten von MeHg gehören. Darüber hinaus gibt es jedoch nur wenige Untersuchungen über die biologischen Mechanismen und die Zusammenhänge in terrestrischen und insbesondere in intestinalen Systemen. Die vorliegende Arbeit leistet daher einen wichtigen Beitrag zur Abschätzung des Potentials zur Hg-Methylierung durch intestinale Bakterien und vertieft die Kenntnisse zu der damit verbundenen Akkumulation der organischen Schwermetallverbindung im Gewebe des Kompostwurms Eisenia foetida (E. foetida). rnIm Rahmen dieser Arbeit wurde erstmals unter Anwendung der Gas Chromatographie mit induktiv gekoppelter Massenspektrometrie (GC-ICP-MS) und Isotopenverdünnungsanalyse verschiedene Kulturen intestinaler Sulfat-reduzierender Bakterien auf die Bildung von organischem Monomethylquecksilber aus Hg(II) untersucht. Da in komplexen bakteriellen Nährlösungen mit hohem Sulfidgehalt Matrixeffekte auftreten und die Analyse von MeHg im Ultraspurenbereich erschweren können, erfolgte die Probenvorbereitung mittels der Methanol-Kaliumhydroxid-Extraktion unter Verwendung eines Maskierungsreagenzes und der Derivatisierung mit Natriumtetrapropylborat. Das Detektionslimit für MeHg in bakteriellen Nährlösungen betrug 0,03 ng/mL. Die Wiederfindung von zertifiziertem Referenzmaterial ERM® CE-464 Tuna Fish war sehr gut und lag in einem Bereich zwischen 98 – 105%. rnDie Resultate der Untersuchung von 14 verschiedenen Rein- und Anreicherungskulturen Sulfat-reduzierender Bakterien zeigten, dass neun Kulturen innerhalb von 12 h nach einer Inkubation mit 0,1 mg/L Hg2+ im Durchschnitt 100 bis 1200 pg/mL MeHg produzierten. Darunter waren zwei Desulfovibrio sp. Stämme, die Spezies Desulfovibrio piger, Desulfovibrio giganteus, Desulfovibrio termitidis, Desulfotomaculum ruminis, Desulfobulbus propionicus sowie Anreicherungskulturen aus dem Intestinaltrakt einer Zygoptera-Larve Zy1 und E. foetida EF4. Die Fähigkeit zur Hg-Methylierung durch eine Spezies der Ordnung Desulfotomaculum aus der Gruppe der Gram-positiven Firmicutes wurde hiermit erstmals beobachtet.rnWeiterhin wurde gezeigt, dass im Intestinaltrakt von E. foetida im Gegensatz zu mikrobiellen Bodenproben eine signifikante biotische Methylierung von Hg(II) durchgeführt wird. Dass diese Transformationen in hohem Maße von der intestinalen Region ausgeht und somit zur Akkumulation von MeHg im Gewebe beiträgt, konnte durch weiterführende Experimente mittels Laserablations-ICP-MS an histologischen Gefrierschnitten des Invertebraten darge-stellt werden. rn

Entwicklung von chiralen- sowie RP-HPLC-Methoden in Verbindung mit hochauflösender MS und deren Anwendung zur Analyse sekundärer organischer Aerosole in der Atmosphäre

Die Erdatmosphäre besteht hauptsächlich aus Stickstoff (78%), Sauerstoff (21%) und Edelga¬sen. Obwohl Partikel weniger als 0,1% ausmachen, spielen sie eine entscheidende Rolle in der Chemie und Physik der Atmosphäre, da sie das Klima der Erde sowohl direkt als auch indirekt beeinflussen. Je nach Art der Bildung unterscheidet man zwischen primären und sekundären Partikeln, wobei primäre Partikel direkt in die Atmosphäre eingetragen werden. Sekundäre Partikel hingegen entstehen durch Kondensation von schwerflüchtigen Verbindungen aus der Gasphase, welche durch Reaktionen von gasförmigen Vorläufersubstanzen (volatile organic compounds, VOCs) mit atmosphärischen Oxidantien wie Ozon oder OH-Radikalen gebildet werden. Da die meisten Vorläufersubstanzen organischer Natur sind, wird das daraus gebil¬dete Aerosol als sekundäres organisches Aerosol (SOA) bezeichnet. Anders als die meisten primären Partikel stammen die VOCs überwiegend aus biogenen Quellen. Es handelt sich da¬bei um ungesättigte Kohlenwasserstoffe, die bei intensiver Sonneneinstrahlung und hohen Temperaturen von Pflanzen emittiert werden. Viele der leichtflüchtigen Vorläufersubstanzen sind chiral, sowohl die Vorläufer als auch die daraus gebildeten Partikel werden aber in den meisten Studien als eine Verbindung betrachtet und gemeinsam analysiert. Die mit Modellen berechneten SOA-Konzentrationen, welche auf dieser traditionellen Vorstellung der SOA-Bil¬dung beruhen, liegen deutlich unterhalb der in der Atmosphäre gefundenen, so dass neben diesem Bildungsweg auch noch andere SOA-Bildungsarten existieren müssen. Aus diesem Grund wird der Fokus der heutigen Forschung vermehrt auf die heterogene Chemie in der Partikelphase gerichtet. Glyoxal als Modellsubstanz kommt hierbei eine wichtige Rolle zu. Es handelt sich bei dieser Verbindung um ein Molekül mit einem hohen Dampfdruck, das auf Grund dieser Eigenschaft nur in der Gasphase zu finden sein sollte. Da es aber über zwei Alde¬hydgruppen verfügt, ist es sehr gut wasserlöslich und kann dadurch in die Partikelphase über¬gehen, wo es heterogenen chemischen Prozessen unterliegt. Unter anderem werden in An¬wesenheit von Ammoniumionen Imidazole gebildet, welche wegen der beiden Stickstoff-He¬teroatome lichtabsorbierende Eigenschaften besitzen. Die Verteilung von Glyoxal zwischen der Gas- und der Partikelphase wird durch das Henrysche Gesetz beschrieben, wobei die Gleichgewichtskonstante die sogenannte Henry-Konstante ist. Diese ist abhängig von der un¬tersuchten organischen Verbindung und den im Partikel vorhandenen anorganischen Salzen. Für die Untersuchung chiraler Verbindungen im SOA wurde zunächst eine Filterextraktions¬methode entwickelt und die erhaltenen Proben anschließend mittels chiraler Hochleistungs-Flüssigchromatographie, welche an ein Elektrospray-Massenspektrometer gekoppelt war, analysiert. Der Fokus lag hierbei auf dem am häufigsten emittierten Monoterpen α-Pinen und seinem Hauptprodukt, der Pinsäure. Da bei der Ozonolyse des α-Pinens das cyclische Grund¬gerüst erhalten bleibt, können trotz der beiden im Molekül vorhanden chiralen Zentren nur zwei Pinsäure Enantiomere gebildet werden. Als Extraktionsmittel wurde eine Mischung aus Methanol/Wasser 9/1 gewählt, mit welcher Extraktionseffizienzen von 65% für Pinsäure Enan¬tiomer 1 und 68% für Pinsäure Enantiomer 2 erreicht werden konnten. Des Weiteren wurden Experimente in einer Atmosphärensimulationskammer durchgeführt, um die Produkte der α-Pinen Ozonolyse eindeutig zu charakterisieren. Enantiomer 1 wurde demnach aus (+)-α-Pinen gebildet und Enantiomer 2 entstand aus (-)-α-Pinen. Auf Filterproben aus dem brasilianischen Regenwald konnte ausschließlich Pinsäure Enantiomer 2 gefunden werden. Enantiomer 1 lag dauerhaft unterhalb der Nachweisgrenze von 18,27 ng/mL. Im borealen Nadelwald war das Verhältnis umgekehrt und Pinsäure Enantiomer 1 überwog vor Pinsäure Enantiomer 2. Das Verhältnis betrug 56% Enantiomer 1 zu 44% Enantiomer 2. Saisonale Verläufe im tropischen Regenwald zeigten, dass die Konzentrationen zur Trockenzeit im August höher waren als wäh¬rend der Regenzeit im Februar. Auch im borealen Nadelwald wurden im Sommer höhere Kon¬zentrationen gemessen als im Winter. Die Verhältnisse der Enantiomere änderten sich nicht im jahreszeitlichen Verlauf. Die Bestimmung der Henry-Konstanten von Glyoxal bei verschiedenen Saataerosolen, nämlich Ammoniumsulfat, Natriumnitrat, Kaliumsulfat, Natriumchlorid und Ammoniumnitrat sowie die irreversible Produktbildung aus Glyoxal in Anwesenheit von Ammoniak waren Forschungs¬gegenstand einer Atmosphärensimulationskammer-Kampagne am Paul-Scherrer-Institut in Villigen, Schweiz. Hierzu wurde zunächst das zu untersuchende Saataerosol in der Kammer vorgelegt und dann aus photochemisch erzeugten OH-Radikalen und Acetylen Glyoxal er¬zeugt. Für die Bestimmung der Glyoxalkonzentration im Kammeraerosol wurde zunächst eine beste¬hende Filterextraktionsmethode modifiziert und die Analyse mittels hochauflösender Mas¬senspektrometrie realisiert. Als Extraktionsmittel kam 100% Acetonitril, ACN zum Einsatz wo¬bei die Extraktionseffizienz bei 85% lag. Für die anschließende Derivatisierung wurde 2,4-Di¬nitrophenylhydrazin, DNPH verwendet. Dieses musste zuvor drei Mal mittels Festphasenex¬traktion gereinigt werden um störende Blindwerte ausreichend zu minimieren. Die gefunde¬nen Henry-Konstanten für Ammoniumsulfat als Saataerosol stimmten gut mit in der Literatur gefundenen Werten überein. Die Werte für Natriumnitrat und Natriumchlorid als Saataerosol waren kleiner als die von Ammoniumsulfat aber größer als der Wert von reinem Wasser. Für Ammoniumnitrat und Kaliumsulfat konnten keine Konstanten berechnet werden. Alle drei Saataerosole führten zu einem „Salting-in“. Das bedeutet, dass bei Erhöhung der Salzmolalität auch die Glyoxalkonzentration im Partikel stieg. Diese Beobachtungen sind auch in der Litera¬tur beschrieben, wobei die Ergebnisse dort nicht auf der Durchführung von Kammerexperi¬menten beruhen, sondern mittels bulk-Experimenten generiert wurden. Für die Trennung der Imidazole wurde eine neue Filterextraktionsmethode entwickelt, wobei sich ein Gemisch aus mit HCl angesäuertem ACN/H2O im Verhältnis 9/1 als optimales Extrak¬tionsmittel herausstellte. Drei verschiedenen Imidazole konnten mit dieser Methode quanti¬fiziert werden, nämlich 1-H-Imidazol-4-carbaldehyd (IC), Imidazol (IM) und 2,2‘-Biimidazol (BI). Die Effizienzen lagen für BI bei 95%, für IC bei 58% und für IM bei 75%. Kammerexperimente unter Zugabe von Ammoniak zeigten höhere Imidazolkonzentrationen als solche ohne. Wurden die Experimente ohne Ammoniak in Anwesenheit von Ammoni¬umsulfat durchgeführt, wurden höhere Imidazol-Konzentrationen gefunden als ohne Ammo¬niumionen. Auch die relative Luftfeuchtigkeit spielte eine wichtige Rolle, da sowohl eine zu hohe als auch eine zu niedrige relative Luftfeuchtigkeit zu einer verminderten Imidazolbildung führte. Durch mit 13C-markiertem Kohlenstoff durchgeführte Experimente konnte eindeutig gezeigt werden, dass es sich bei den gebildeten Imidazolen und Glyoxalprodukte handelte. Außerdem konnte der in der Literatur beschriebene Bildungsmechanismus erfolgreich weiter¬entwickelt werden. Während der CYPHEX Kampagne in Zypern konnten erstmalig Imidazole in Feldproben nach¬gewiesen werden. Das Hauptprodukt IC zeigte einen tageszeitlichen Verlauf mit höheren Kon¬zentrationen während der Nacht und korrelierte signifikant aber schwach mit der Acidität und Ammoniumionenkonzentration des gefundenen Aerosols.

Synthesis and metal complexes of C2 symmetric ligands obtained from (R)-(+)-Betti and dialdehydes for asymmetric induction reactions

The aim of this master’s research thesis was the employment of an enantiopure 1,3-aminoalcohol, the 1-(α-aminobenzyl)-2-naphthol, known as Betti base, for the synthesis of some novel compounds which show a C2 symmetry. Some of these compounds, after derivatization, were used as ligands in association with transition metals to prepare some catalysts for enantioselective catalytic reactions. Some aminoalcohol (Salan-type) derivatives of these compounds were obtained upon reduction and in some cases it was possible to obtain complexes with transition metals such as Mn, Ni, Co and Cu. Furthermore a novel 6-membered analogue bisoxazoline ligand, 2,6-bis((R)-1-Phenyl-1H-naphtho[1,2-e][1,3]oxazin-3-yl)pyridine, was obtained and from it two Cu-complexes were prepared. The metal complexes were employed in some reactions to test the asymmetric induction, which was in some cases up to discrete values.

Probing The Active And Allosteric Binding Sites Of Soybean Lipoxygenase-1

Soybean lipoxygenase-1 is a model for lipoxygenase activity. While the mechanism of oxygenation is understood, the substrate binding mechanism has not yet been elucidated. Two putative binding mechanisms are the ¿head-first¿ and ¿tail-first¿ models, in which the carboxy-terminus or the methyl terminus of the fatty acid substrate is inserted into the active site while the remainder of the molecule protrudes from the surface, respectively. Previous work has demonstrated that derivatization of fatty acid substrates with D-tryptophan increases active site affinity. It has also been shown that while polyunsaturated fatty acids are the natural substrates of lipoxygenases, monounsaturated fatty acids can be oxygenated at a much slower rate. Starting with a monounsaturated fatty acid, oleic acid, as a platform, the molecule N-oleoyl-D-tryptophan (ODT) was synthesized with the anticipation of it being a potent competitive substrate-analogue inhibitor that could be used to discern the substrate binding mechanism. Inhibition kinetics demonstrated that this molecule functions as a partially competitive inhibitor, through an unknown mechanism. The implication behind partially competitive inhibition is that substrate and inhibitor molecules can bind simultaneously to the enzyme, which alludes to the presence of an allosteric binding domain. To investigate the possibility of an inhibitor binding site on the non-catalytic subunit, limited proteolysis was used to cleave the subunits apart which should have eliminated inhibition. Interestingly, it was observed that at high substrate concentrations the inhibitor was completely ineffective, but at low substrate concentrations the inhibitor maintained its standard efficacy. A satisfactory explanation for these results has not yet been determined.

TERPENE AND TERPENOID EMISSIONS AND SECONDARY ORGANIC AEROSOL PRODUCTION

Approximately 90% of fine aerosol in the Midwestern United States has a regional component with a sizable fraction attributed to secondary production of organic aerosol (SOA). The Ozark Forest is an important source of biogenic SOA precursors like isoprene (> 150 mg m-2 d-1), monoterpenes (10-40 mg m-2 d-1), and sesquiterpenes (10-40 mg m-2d-1). Anthropogenic sources include secondary sulfate and nitrate and biomass burning (51-60%), vehicle emissions (17-26%), and industrial emissions (16-18%). Vehicle emissions are an important source of volatile and vapor-phase, semivolatile aliphatic and aromatic hydrocarbons that are important anthropogenic sources of SOA precursors. The short lifetime of SOA precursors and the complex mixture of functionalized oxidation products make rapid sampling, quantitative processing methods, and comprehensive organic molecular analysis essential elements of a comprehensive strategy to advance understanding of SOA formation pathways. Uncertainties in forecasting SOA production on regional scales are large and related to uncertainties in biogenic emission inventories and measurement of SOA yields under ambient conditions. This work presents a bottom-up approach to develop a conifer emission inventory based on foliar and cortical oleoresin composition, development of a model to estimate terpene and terpenoid signatures of foliar and bole emissions from conifers, development of processing and analytic techniques for comprehensive organic molecular characterization of SOA precursors and oxidation products, implementation of the high-volume sampling technique to measure OA and vapor-phase organic matter, and results from a 5 day field experiment conducted to evaluate temporal and diurnal trends in SOA precursors and oxidation products. A total of 98, 115, and 87 terpene and terpenoid species were identified and quantified in commercially available essential oils of Pinus sylvestris, Picea mariana, and Thuja occidentalis, respectively, by comprehensive, two-dimensional gas chromatography with time-of-flight mass spectrometric detection (GC × GC-ToF-MS). Analysis of the literature showed that cortical oleoresin composition was similar to foliar composition of the oldest branches. Our proposed conceptual model for estimation of signatures of terpene and terpenoid emissions from foliar and cortical oleoresin showed that emission potentials of the foliar and bole release pathways are dissimilar and should be considered for conifer species that develop resin blisters or are infested with herbivores or pathogens. Average derivatization efficiencies for Methods 1 and 2 were 87.9 and 114%, respectively. Despite the lower average derivatization efficiency of Method 1, distinct advantages included a greater certainty of derivatization yield for the entire suite of multi- and poly-functional species and fewer processing steps for sequential derivatization. Detection limits for Method 1 using GC × GC- ToF-MS were 0.09-1.89 ng μL-1. A theoretical retention index diagram was developed for a hypothetical GC × 2GC analysis of the complex mixture of SOA precursors and derivatized oxidation products. In general, species eluted (relative to the alkyl diester reference compounds) from the primary column (DB-210) in bands according to n and from the secondary columns (BPX90, SolGel-WAX) according to functionality, essentially making the GC × 2GC retention diagram a Carbon number-functionality grid. The species clustered into 35 groups by functionality and species within each group exhibited good separation by n. Average recoveries of n-alkanes and polyaromatic hydrocarbons (PAHs) by Soxhlet extraction of XAD-2 resin with dichloromethane were 80.1 ± 16.1 and 76.1 ± 17.5%, respectively. Vehicle emissions were the common source for HSVOCs [i.e., resolved alkanes, the unresolved complex mixture (UCM), alkylbenzenes, and 2- and 3-ring PAHs]. An absence of monoterpenes at 0600-1000 and high concentrations of monoterpenoids during the same period was indicative of substantial losses of monoterpenes overnight and the early morning hours. Post-collection, comprehensive organic molecular characterization of SOA precursors and products by GC × GC-ToFMS in ambient air collected with ~2 hr resolution is a promising method for determining biogenic and anthropogenic SOA yields that can be used to evaluate SOA formation models.

Directed Metalation Cascade To Access Highly Functionalized Thieno2,3-fbenzofuran and Exploration as Building Blocks for Organic Electronics

A tandem directed metalation has been successfully applied to the preparation of thieno2,3-fbenzofuran-4,8-dione, providing an efficient and facile approach to symmetrically and unsymmetrically functionalize the thieno2,3-fbenzofuran core at the 2,6 positions as well as to introduce the electron-withdrawing or -donating groups (EWG or EDG) at its 4,8 positions. The presence of various functional groups makes late-stage derivatization attainable.

Structures and Concentrations of Surfactants in Gut Fluid of the Marine Polychaete Arenicola Marina

Marine invertebrate deposit feeders secrete surfactants into their gut fluid in concentrations sufficient to induce micelle formation, enhancing solubilization of sedimentary lipids. We isolated and identified 3 related surfactant molecules from the deposit-feeding polychaete lugworm Arenicola marina. Surfactants were isolated and separated by a combination of solvent extraction and thin-layer and gas chromatography. Identification was performed using mass and infrared spectrometry, coupled to various derivatization and hydrolysis reactions. A. marina produces a mixture of related yet distinct anionic surfactants composed of branched, C9, saturated and unsaturated fatty acids that are amide linked to leucine or glycine residues, showing some similarity to crustacean surfactants. The critical micelle concentration of the mixture of these surfactants in gut fluid was about 2 mM, and total concentrations ranged from 5.5 to 19.5 mM. The hydrophilic amide linkage helps to explain previous observations that gut surfactants do not adsorb onto sediment transiting the gut.

A new sensitive method for the quantification of glyoxal and methylglyoxal in snow and ice by stir bar sorptive extraction and liquid desorption-HPLC-ESI-MS

In this study, the development of a new sensitive method for the analysis of alpha-dicarbonyls glyoxal (G) and methylglyoxal (MG) in environmental ice and snow is presented. Stir bar sorptive extraction with in situ derivatization and liquid desorption (SBSE-LD) was used for sample extraction, enrichment, and derivatization. Measurements were carried out using high-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS). As part of the method development, SBSE-LD parameters such as extraction time, derivatization reagent, desorption time and solvent, and the effect of NaCl addition on the SBSE efficiency as well as measurement parameters of HPLC-ESI-MS/MS were evaluated. Calibration was performed in the range of 1–60 ng/mL using spiked ultrapure water samples, thus incorporating the complete SBSE and derivatization process. 4-Fluorobenzaldehyde was applied as internal standard. Inter-batch precision was <12 % RSD. Recoveries were determined by means of spiked snow samples and were 78.9 ± 5.6 % for G and 82.7 ± 7.5 % for MG, respectively. Instrumental detection limits of 0.242 and 0.213 ng/mL for G and MG were achieved using the multiple reaction monitoring mode. Relative detection limits referred to a sample volume of 15 mL were 0.016 ng/mL for G and 0.014 ng/mL for MG. The optimized method was applied for the analysis of snow samples from Mount Hohenpeissenberg (close to the Meteorological Observatory Hohenpeissenberg, Germany) and samples from an ice core from Upper Grenzgletscher (Monte Rosa massif, Switzerland). Resulting concentrations were 0.085–16.3 ng/mL for G and 0.126–3.6 ng/mL for MG. Concentrations of G and MG in snow were 1–2 orders of magnitude higher than in ice core samples. The described method represents a simple, green, and sensitive analytical approach to measure G and MG in aqueous environmental samples.

Synthesis and cellular characterization of novel isoxazolo- and thiazolohydrazinylidene-chroman-2,4-diones on cancer and non-cancer cell growth and death

Coumarins are extensively studied anticoagulants that exert additional effects such as anticancerogenic and even anti-inflammatory. In order to find new drugs with anticancer activities, we report here the synthesis and the structural analysis of new coumarin derivatives which combine the coumarin core and five member heterocycles in hydrazinylidene-chroman-2,4-diones. The derivatives were prepared by derivatization of the appropriate heterocyclic amines which were used as electrophiles to attack the coumarin ring. The structures were characterized by spectroscopic techniques including IR, NMR, 2D-NMR and MS. These derivatives were further characterized especially in terms of a potential cytotoxic and apoptogenic effect in several cancer cell lines including the breast and prostate cancer cell lines MCF-7, MDA-MB-231, PC-3, LNCaP, and the monocytic leukemia cell line U937. Cell viability was determined after 48 h and 72 h of treatment with the novel compounds by MTT assay and the 50% inhibitory concentrations (EC50 values) were determined. Out of the 8 novel compounds screened for reduced cell viability, 4c, 4d and 4e were found to be the most promising and effective ones having EC50 values that were several fold reduced when compared to the reference substance 4-hydroxycoumarin. However, the effects were cancer cell line dependent. The breast cancer MDA-MB-231 cells, the prostate cancer LNCaP cells, and U937 cells were most sensitive, MCF-7 cells were less sensitive, and PC-3 cells were more resistant. Reduced cell viability was accompanied by increased apoptosis as shown by PARP-1 cleavage and reduced activity of the survival protein kinase Akt. In summary, this study has identified three novel coumarin derivatives that in comparison to 4-hydroxycoumarin have a higher efficiency to reduce cancer cell viability and trigger apoptosis and therefore may represent interesting novel drug candidates

Optimization of a DNSH passive sampling method to measure airborne carbonyls

Various airborne aldehydes and ketones (i.e., airborne carbonyls) present in outdoor, indoor, and personal air pose a risk to human health at present environmental concentrations. To date, there is no adequate, simple-to-use sampler for monitoring carbonyls at parts per billion concentrations in personal air. The Passive Aldehydes and Ketones Sampler (PAKS) originally developed for this purpose has been found to be unreliable in a number of relatively recent field studies. The PAKS method uses dansylhydrazine, DNSH, as the derivatization agent to produce aldehyde derivatives that are analyzed by HPLC with fluorescence detection. The reasons for the poor performance of the PAKS are not known but it is hypothesized that the chemical derivatization conditions and reaction kinetics combined with a relatively low sampling rate may play a role. This study evaluated the effect of absorption and emission wavelengths, pH of the DNSH coating solution, extraction solvent, and time post-extraction for the yield and stability of formaldehyde, acetaldehyde, and acrolein DNSH derivatives. The results suggest that the optimum conditions for the analysis of DNSHydrazones are the following. The excitation and emission wavelengths for HPLC analysis should be at 250nm and 500nm, respectively. The optimal pH of the coating solution appears to be pH 2 because it improves the formation of di-derivatized acrolein DNSHydrazones without affecting the response of the derivatives of the formaldehyde and acetaldehyde derivatives. Acetonitrile is the preferable extraction solvent while the optimal time to analyze the aldehyde derivatives is 72 hours post-extraction. ^

A standard operation procedure and a pilot water testing for herbicides in northwest Harris County, Texas

Herbicides are used to control the growth of weeds along highways, power lines, and many other urban locations. Exposure to herbicides has been linked to adverse health outcomes. This study was initiated to pretest for the presence of herbicides in multiple water sources near intersections in a corridor in the Northwest Harris County (specifically in the Highway 6/FM 1960, North Freeway 45, US 290 and S 99 corridor). Roadside water and tap water samples were collected and analyzed for herbicides using the established Environmental Protection Agency (EPA) Method 515.4: "Determination of Chlorinated Acids in Drinking Water by Liquid-Liquid Micro-extraction, Derivatization, and Fast Gas Chromatography with Electron Capture Detection." A standard operating procedure (adapted from the US EPA Method 515.4) was developed for subsequent, larger studies of environmental fate of herbicides and non-occupational exposure risks. Preliminary testing of 16 water samples was performed to pretest the existence of trace herbicides; all concentrations that were greater than the minimum reporting limits of each analyte are reported with a 99 percent confidence. This study failed to find concentrations above the limits of detection of the method in any of the samples collected on June 15, 2008. However, this does not indicate that the waters around the NW Harris County are free of herbicides and metabolites. A larger and repeated sampling in the region would be necessary to make that claim. ^

Proceedings of the 24th Annual Biochemical Engineering Symposium

The 24th Biochemical Engineering Symposium was held 9-10 September 1994 at the YMCA of the Rockies conference center in Estes Park, Colorado, under the sponsorship of the Department of Chemical Engineering at the University of Colorado. Previous symposia in this series have been hosted by Kansas State University (1st, 3rd, 5th, 9th, 12th, 16th, 20th), University of Nebraska-Lincoln (2nd, 4th), Iowa State University (6th, 7th, 10th, 13th, 17th, 22nd), University of Missouri-Columbia (8th, 14th, 19th), Colorado State University (11th, 15th, 21st), University of Colorado (18th), and the University of Oklahoma (23rd). The next symposium is scheduled to be held at the University of Missouri-Columbia. The symposia are devoted to talks by students about their ongoing research. Because final publication usually takes place elsewhere, the papers included in the proceedings are brief, and often cover work in progress. ContentsIn-Well Aeration: An Innovative Subsurface Remediation TechnologyPrashant Gandhi, X. Yang, L.E. Erickson, and L. T. Fan; Kansas State University Expression of an Antimicrobial Peptide Analog in Eacherlchill coliChris Haught and Roger G. Harrison; University of Oklahoma Using High-frequency Backpulaing to Maximize Croasflow Filtration PerformanceSanjeev G. Redkar and Robert H. Davis; University of Colorado Low Molecular Weight Organic Compositions of Acid Waters from Vegetable Oil SoapstocksSteven L. Johansen, Arunthathi Sivasothy, Peter J. Reilly, and Earl G. Hammond; Iowa State University; Michael K. Dowd; U.S. Department of Agriculture Gas Phase Composition Effects on Suspension Cultures of Taxus cuspidata Noushin Mirjalili and James C. Linden; Colorado State University Cybernetic Modeling of Spontaneous Oscillations in Continuous Cultures of Ssccharomyces cerevisiaeKenneth D. Jones and Dhinakar S. Kompala; University of Colorado The Effect of Turbulent Shear on Calcium Mobilization in Mammalian CellsChristopher M. Cannizzaro, Pradyumna K. Namdev, and Eric H. Dunlop; Colorado State University Experimental Studies of Droplet Ejection at the Free Surface In Sparged ReactorsT. Y. Yiin, L A. Glasgow, and L. E. Erickson; Kansas State University The Role of Domain E (Starch-Binding Region) on the Activity of a Bacillus macersns Cyclodextrln GlucanotransferaseHai-yin Chang, Trang Le, and Zivko L. Nikolov; Iowa State University Use of the Rotating Wall Vessel for Study of Plant Cell Suspension CulturesXinzhi Sun and James C. Linden; Colorado State University A Novel Counter-Current Distribution Apparatus for the Study of Multi-Stage Aqueous Two-Phase Extraction of Biomolecules and Cell ParticlesMartin R. Guinn and Paul Todd; University of Colorado The Dynamics of Unhooking and Contraction of a Polyelectrolyte Chain Around an Isolated PostLin Zhang and Edith M. Sevick; University of Colorado A Laboratory Study of the Fate of Trichloroathylene and 1,1,1-Trlchloroathane In the Presence of Alfalfa PlantsMuralidharan Narayanan, Ryan M. Green, Lawrence C. Davis, and Larry E. Erickson; Kansas State University Modeling the Fate of Pyrene In the RhIzosphereS.K. Santharam, LE. Erickson, and L. T. Fan; Kansas State University Derivatization of MaltooligosaccharidesDaniela Prinz, Peter J. Reilly, and Zivko L. Nikolov; Iowa State University Probing Surfactant-Protein Binding by EPA SpectroscopyNarendra B. Bam, Yale University; Theodore W. Randolph; University of Colorado Optimization of a Stir-Cell Bioreactor for In Vitro Production of RNANeal T. Williams, Kim A. Wicklund, and Robert H. Davis; University of Colorado

Measurements of seawater and aerosol particles during POLARSTERN cruise ANT-XXVII/4

An analytical method for the determination of the alpha dicarbonyls glyoxal (GLY) and methylglyoxal (MGLY) from seawater and marine aerosol particles is presented. The method is based on derivatization with o-(2,3,4,5,6-Pentafluorobenzyl)-hydroxylamine (PFBHA) reagent, solvent extraction and GC-MS (SIM) analysis. The method showed good precision (RSD < 10%), sensitivity (detection limits in the low ng/l range), and accuracy (good agreement between external calibration and standard addition). The method was applied to determine GLY and MGLY in oceanic water sampled during the Polarstern cruise ANT XXVII/4 from Capetown to Bremerhaven in spring 2011. GLY and MGLY were determined in the sea surface microlayer (SML) of the ocean and corresponding bulk water (BW) with average concentrations of 228 ng/l (GLY) and 196 ng/l (MGLY). The results show a significant enrichment (factor of 4) of GLY and MGLY in the SML. Furthermore, marine aerosol particles (PM1) were sampled during the cruise and analyzed for GLY (average concentration 0.19 ng/m**3) and MGLY (average concentration 0.15 ng/m**3). On aerosol particles, both carbonyls show a very good correlation with oxalate, supporting the idea of a secondary formation of oxalic acid via GLY and MGLY. Concentrations of GLY and MGLY in seawater and on aerosol particles were correlated to environmental parameters such as global radiation, temperature, distance to the coastline and biological activity. There are slight hints for a photochemical production of GLY and MGLY in the SML (significant enrichment in the SML, higher enrichment at higher temperature). However, a clear connection of GLY and MGLY to global radiation as well as to biological activity cannot be concluded from the data. A slight correlation between GLY and MGLY in the SML and in aerosol particles could be a hint for interactions, in particular of GLY, between seawater and the atmosphere.

Occurrence and analysis of selected pharmaceutical compounds in soil from Spanish agricultural fields

This work describes the analysis of 15 pharmaceutical compounds, belonging to different therapeutic classes (anti-inflammatory/analgesics, lipid regulators, antiepileptics, ?-blockers and antidepressants) and with diverse physical?chemical properties, in Spanish soils with different farmland uses. The studied compounds were extracted from soil by ultrasound-assisted extraction (UAE) and determined, after derivatization, by gas chromatography with mass spectrometric detection (GC?MS). The limits of detection (LODs) ranged from 0.14 ng g?1 (naproxen) to 0.65 ng g?1 (amitriptyline). At least two compounds where detected in all samples, being ibuprofen, salicylic acid, and paracetamol, the most frequently detected compounds. The highest levels found in soil were 47 ng g?1 for allopurinol and 37 ng g?1 for salicylic acid. The influence of the type of crop and the sampling area on the levels of pharmaceuticals in soil, as well as their relationship with soil physical?chemical properties, was studied. The frequent and widespread detection of some of these compounds in agricultural soils show a diffuse contamination, although the low levels found do not pose a risk to the environment or the human health.

(4-Aminomethyl)phenylguanidine derivatives as nonpeptidic highly selective inhibitors of human urokinase

Increased expression of the serine protease urokinase-type plasminogen activator (uPA) in tumor tissues is highly correlated with tumor cell migration, invasion, proliferation, progression, and metastasis. Thus inhibition of uPA activity represents a promising target for antimetastatic therapy. So far, only the x-ray crystal structure of uPA inactivated by H-Glu-Gly-Arg-chloromethylketone has been reported, thus limited data are available for a rational structure-based design of uPA inhibitors. Taking into account the trypsin-like arginine specificity of uPA, (4-aminomethyl)phenylguanidine was selected as a potential P1 residue and iterative derivatization of its amino group with various hydrophobic residues, and structure–activity relationship-based optimization of the spacer in terms of hydrogen bond acceptor/donor properties led to N-(1-adamantyl)-N′-(4-guanidinobenzyl)urea as a highly selective nonpeptidic uPA inhibitor. The x-ray crystal structure of the uPA B-chain complexed with this inhibitor revealed a surprising binding mode consisting of the expected insertion of the phenylguanidine moiety into the S1 pocket, but with the adamantyl residue protruding toward the hydrophobic S1′ enzyme subsite, thus exposing the ureido group to hydrogen-bonding interactions. Although in this enzyme-bound state the inhibitor is crossing the active site, interactions with the catalytic residues Ser-195 and His-57 are not observed, but their side chains are spatially displaced for steric reasons. Compared with other trypsin-like serine proteases, the S2 and S3/S4 pockets of uPA are reduced in size because of the 99-insertion loop. Therefore, the peculiar binding mode of the new type of uPA inhibitors offers the possibility of exploiting optimized interactions at the S1′/S2′ subsites to further enhance selectivity and potency. Because crystals of the uPA/benzamidine complex allow inhibitor exchange by soaking procedures, the structure-based design of new generations of uPA inhibitors can rely on the assistance of x-ray analysis.