Helge Egsgaard

Enzymatic Evidence for the Key Role of Arginine in Nitrogen Translocation by Arbuscular Mycorrhizal Fungi

Key enzymes of the urea cycle and 15N-labeling patterns of arginine (Arg) were measured to elucidate the involvement of Arg in nitrogen translocation by arbuscular mycorrhizal (AM) fungi. Mycorrhiza was established between transformed carrot (Daucus carota) roots and Glomus intraradices in two-compartment petri dishes and three ammonium levels were supplied to the compartment containing the extraradical mycelium (ERM), but no roots. Time courses of specific enzyme activity were obtained for glutamine synthetase, argininosuccinate synthetase, arginase, and urease in the ERM and AM roots. 15NH4+ was used to follow the dynamics of nitrogen incorporation into and turnover of Arg. Both the absence of external nitrogen and the presence of l-norvaline, an inhibitor of Arg synthesis, prevented the synthesis of Arg in the ERM and resulted in decreased activity of arginase and urease in the AM root. The catabolic activity of the urea cycle in the roots therefore depends on Arg translocation from the ERM. 15N labeling of Arg in the ERM was very fast and analysis of its time course and isotopomer pattern allowed estimation of the translocation rate of Arg along the mycelium as 0.13 μg Arg mg−1 fresh weight h−1. The results highlight the synchronization of the spatially separated reactions involved in the anabolic and catabolic arms of the urea cycle. This synchronization is a prerequisite for Arg to be a key component in nitrogen translocation in the AM mycelium.

Carvone : an overlooked contact allergen cross-reacting with sesquiterpene lactones ?

The terpene l‐carvone is one of the main constituents of spearmint oil. The sensitizing potential of l‐carvone has been considered low, but it has occasionally caused contact allergy in users of spearmint toothpaste and chewing gum. l‐Carvone is also an oxidation product of d‐limonene that occurs in solvents used increasingly in industry. We included l‐carvone 5% pet, in the standard patch test series. In the 1st year, 541 patients were tested and 15 (2.77%) had positive, and 12 doubtful positive (?+) reactions to l‐carvone. The strongest reactions were observed in 9 patients with concomitant Compositae sensitivity. The key clinical Features and other contact allergies of the patients are presented. When re‐testing with l‐carvone in the same or lower concentrations, only 2 out 8 patients had positive reactions. Possible reasons for this discrepancy are discussed in terms of cross‐reactions, concomitant sensitization, excited skin syndrome, irritancy and facilitated immunological response.

Determination of tetramethyllead and tetraethyllead in the atmosphere by a two-step enrichment method and gas chromatographic—mass spectrometric isotope dilution analysis

Abstract The development of a specific and sensitive technique for determining tetramethyllead (TML) in air is described. The method has been tested in different areas under differing meteorological conditions to determine the atmospheric content of TML and tetraethyllead (TEL). The advantages and limitations of the method are critically discussed. The tetraalkyllead compounds are collected on Porapak QS or N at ambient temperature, desorbed, re-collected at -80°C on a small column containing 4% Apiezon M on Chromosorb PAW-DCMS, and analysed by gas chromatography—mass spectrometry with single ion monitoring. The isotope dilution technique is used by adding known amounts of d 12 -TML and d 20 -TEL to the sampling columns in advance. This makes it possible to correct for decomposition during the sampling and/or during the analytical procedure. The detection limit for TML is 20 pg m -3 .

Exploring Symbiotic Nitrogen Fixation and Assimilation in Pea Root Nodules by in Vivo 15N Nuclear Magnetic Resonance Spectroscopy and Liquid Chromatography-Mass Spectrometry

Nitrogen (N) fixation and assimilation in pea (Pisum sativum) root nodules were studied by in vivo 15N nuclear magnetic resonance (NMR) by exposing detached nodules to15N2 via a perfusion medium, while recording a time course of spectra. In vivo 31P NMR spectroscopy was used to monitor the physiological state of the metabolically active nodules. The nodules were extracted after the NMR studies and analyzed for total soluble amino acid pools and 15N labeling of individual amino acids by liquid chromatography-mass spectrometry. A substantial pool of free ammonium was observed by 15N NMR to be present in metabolically active, intact nodules. The ammonium ions were located in an intracellular environment that caused a remarkable change in the in vivo 15N chemical shift. Alkalinity of the ammonium-containing compartment may explain the unusual chemical shift; thus, the observations could indicate that ammonium is located in the bacteroids. The observed15N-labeled amino acids, glutamine/glutamate and asparagine (Asn), apparently reside in a different compartment, presumably the plant cytoplasm, because no changes in the expected in vivo15N chemical shifts were observed. Extensive15N labeling of Asn was observed by liquid chromatography-mass spectrometry, which is consistent with the generally accepted role of Asn as the end product of primary N assimilation in pea nodules. However, the Asn 15N amino signal was absent in in vivo 15N NMR spectra, which could be because of an unfavorable nuclear Overhauser effect. γ-Aminobutyric acid accumulated in the nodules during incubation, but newly synthesized 15N γ-aminobutyric acid seemed to be immobilized in metabolically active pea nodules, which made it NMR invisible.

Slow alkyl, alkene, and alkenyl loss from primary alkylamines: Isomerization of the low-energy molecular ions prior to fragmentation in the μsec timeframe

Abstract Isomerization of the molecular ion precedes alkyl radical elimination from many primary aliphatic amines. Rearrangement of the carbon skeleton is initiated by 1,5-hydrogen abstraction by the -NH 2 +· and converts primary-alkyl amines to sec - and tert -alkyl amines; fragmentation is then by α-cleavage of the rearranged molecular ions. Isomerization is also encountered for amines branched at the α-carbon atom. The elimination of propene from n -pentylamine and of a butenyl radical from neo -pentylamine is discussed. Remote (δ or e) cleavage does not contribute to the low-energy reactions.

Leaf surface wax is a source of plant methane formation under UV radiation and in the presence of oxygen

The terrestrial vegetation is a source of UV radiation-induced aerobic methane (CH4 ) release to the atmosphere. Hitherto pectin, a plant structural component, has been considered as the most likely precursor for this CH4 release. However, most of the leaf pectin is situated below the surface wax layer, and UV transmittance of the cuticle differs among plant species. In some species, the cuticle effectively absorbs and/or reflects UV radiation. Thus, pectin may not necessarily contribute substantially to the UV radiation-induced CH4 emission measured at surface level in all species. Here, we investigated the potential of the leaf surface wax itself as a source of UV radiation-induced leaf aerobic CH4 formation. Isolated leaf surface wax emitted CH4 at substantial rates in response to UV radiation. This discovery has implications for how the phenomenon should be scaled to global levels. In relation to this, we demonstrated that the UV radiation-induced CH4 emission is independent of leaf area index above unity. Further, we observed that the presence of O2 in the atmosphere was necessary for achieving the highest rates of CH4 emission. Methane formation from leaf surface wax is supposedly a two-step process initiated by a photolytic rearrangement reaction of the major component followed by an α-cleavage of the generated ketone.

Kinetic study of the formation of isotopically substituted ozone in argon

The kinetics of the formation of ozone was studied by using pulse radiolysis coupled with time-resolved UV absorption at 275 nm and at T = 294.9±0.6 K. The rate constant for the formation of ozone 16O16O16O in argon was determined to be k3a = (3.38±0.04) × 10−34 cm6 molecule−2 s−1. The rate constants for the reactions 18O + 16O16O (k3b), 16O + 16O18O (k3c), 16O + 18O18O (k3d), 18O + 16O18O (k3e), and 18O + 18O18O (k3f) were studied, and the following parameters were determined: (k3b + k3d)/(2k3a) = (1.184±0.037), (k3c + k3e)/(2k3a) = (1.155±0.062), and k3f/k3a = (0.977±0.021). The values for (k3b + k3d)/(2k3a) and (k3c + k3e)/(2k3a) obtained here are equal to the values derived from the product studies and the recently reported relative rate study but higher than the reported values for (k3b + k3d)/(2k3a) and (k3c + k3e)/(2k3a) obtained by using CO2 as a third body. The parameter k3f/k3a = (0.977±0.021) is lower than the value of k3f/k3a obtained by using CO2 as a third body and the value derived from the product studies. These different values of k3f may be partly due to changes in third body efficiency or due to resonance interactions between the excited ozone molecules and the third body. The absolute measurements reported here together with literature data suggest that the nature of the third body is an important factor in controlling the enhancements of the rate constants for ozone formation and that asymmetry of neither ozone nor dioxygen ensure a fast ozone formation rate.

First direct kinetic study of isotopic enrichment of ozone

The formation kinetics of ozone has been studied using isotopes and pulse radiolysis combined with time-resolved UV absorption spectroscopy. An enhancement of (9.8±2.6)% was found for the rate constant for the reaction of 18O with 18O18O relative to that for the 16O+ 16O16O reaction. The average formation rate for unsymmetric 18O16O16O and 16O18O18O from O2 and 18O18O respectively was enhanced by (14.7±2.8)%. For the formation of a mixture of symmetric and unsymmetric ozone species from O18O the enhancement was (3.6±2.4)%. This leads to the conclusion that both mass and symmetry affect the rate constant for formation of isotopic ozone. The results are compared with recent enhancement studies from the literature, and an apparent conflict is discussed.

An effective approach to flash vacuum thermolytic studies

Abstract Flash vacuum thermolysis in combination with field ionization mass spectrometry, supplementary with collision activated spectra of the single field ionized molecules, is shown to be a facile and highly informative method for studying even very complex mixtures of primarily formed products in the gas phase thermolysis of organic molecules. The method allows quantitative detection of substances with half lifes t 1 2 > 10 −3 sec. A detailed description of the apparatus, which offers the possibility of studying gas phase thermolyses over a wide range of temperatures (300—1400 K), and the method is given, and possible applications discussed.

Contact allergy to 2‐hydroxy‐5‐tert‐butyl benzylalcohol and 2,6‐bis(hydroxymethyl)‐4‐tert‐butylphenol, components of a phenolic resin used in marking pens

2‐hydroxy‐5‐tert‐butyl benzylalcohol and 2,6‐bis(hydroxymethyl)‐4‐tert‐butylphenol were identified as contact allergens in a phenolic resin used as a tackifier in the ink of a marking pen, which, after being used directly on the skin, caused an acute contact dermatitis on the hand of a 13‐year‐old boy. The patient also reacted to 4‐tert‐butylphenol‐formaldehyde resin (BPF resin) 1% pet, included in the European standard series.