Andreas Rabenstein

Detection of traces of oxidized and reduced sulfur compounds in small samples by combination of different high-performance liquid chromatography methods☆

Depending on the sulfur species, picomoles of different inorganic sulfur compounds can be detected and separated by HPLC in one arrangement in a sample volume less than 50 μl. The combination of fluorescence labelling of reduced inorganic sulfur compounds such as sulfide (S2−), sulfite(SO32− and thiosulfate (S2O32−) with monobromobimane followed by an extraction of elemental sulfur (S°) by chloroform treatment enables the detection of all mentioned sulfur compounds as well as sulfate (remaining aqueous phase) in the same sample. While the derivatized sulfur compounds could be detected by their fluorescence emission at 480 nm, elemental sulfur is identified by its UV absorption at 263 nm. Sulfate in the remaining aqueous phase is detected by HPLC with indirect UV detection at 254 nm. Detection ranges for the different sulfur compounds examined are as follows: sulfide (5 μM to 1.5 mM), sulfite (5 μM to 1.0 mM), thiosulfate (1 μM to 1.5 mM), elemental sulfur (2 μM to 32 mM) and sulfate (5 μM to >1 mM).

Determination of low thiourea concentrations in industrial process water and natural samples using reversed-phase high-performance liquid chromatography.

A rapid LC method was developed to determine thiourea in natural samples and freshly prepared as well as used tin-baths from the galvanic industry, with LC on a C18-bonded silica column and with doubly distilled water as eluent. The retention time of thiourea is 1.35 min +/- 5%. UV detection is at 236 nm. Small sample volumes of 10-50 microl allow detection down to 2 microg/l thiourea without any interference by heavy metals or organic compounds and in the case of marine samples by mineral salts. The presented technique should be highly useful for industrial purposes.

SULPHITE AS INTERMEDIATE SULPHUR COMPOUND IN ANAEROBIC SULPHIDE OXIDATION TO THIOSULPHATE BY MARINE CYANOBACTERIA

Abstract Anaerobic sulphide oxidation in the light was studied with seven cyanobacterial strains isolated from the southern Baltic Sea. All strains oxidized sulphide (1.2-3.5 mM) stoichio-metrically to thiosulphate. No elemental sulphur could be detected as end product of anoxy-genic photosynthesis. Sulphite as intermediate sulphur compound was released into the growth medium during anaerobic sulphide oxidation by these cyanobacteria. The formation of sulphite from sulphide was inhibited by the photosynthesis inhibitor DCMU. The cyanobacterium Oscillatoria sp. strain BO 32 produced another so far not identified intermediate sulphur compound which was probably stored within the cells.

Dimethyl sulphoxide reduction with reduced sulphur compounds as electron donors by anoxygenic phototrophic bacteria

The marine purple non-sulphur bacterium Rhodovulum euryhalinum strain DSM 4868 reduced dimethyl sulphoxide (DMSO) to dimethyl sulphide (DMS) chemotrophically with sulphide as electron donor. The oxidation of sulphide was correlated with the formation of polysulphides. R. euryhalinum reduced DMSO phototrophically with sulphide as well, but the amount of DMSO reduced in relation to sulphide oxidized was lower. The marine green sulphur bacterium Chlorobium vibrioforme strain DSM 8327 reduced DMSO to DMS phototrophically with sulphide and thiosulphate as electron donors. The extent of DMSO reduction was much less in the dark. Eight strains of purple sulphur bacteria - marine, brackish water and freshwater isolates - and another marine green sulphur bacterium showed a very weak capacity for DMSO reduction with sulphide or thiosulphate as electron donors in the light and dark, respectively.

Biodegradation of a high molecular weight aliphatic ether – indications of an unusual biodegradation pathway

An aliphatic ether (1-phytanyl-1-octadecanyl-ether) of high molecular weight was used as a sole carbon source in degradation experiments with different aerobic bacteria. The enriched culture B5, obtained from fuel contaminated soils, was able to degrade the substance for more than 90%. A culture of Rhodococcus ruber was similarly effective. Detailed investigation of the metabolites allowed us to characterize an unusual degradation pathway via a mid-chain oxidation mechanism (`internal oxidative pathway). Obviously, formation of intermediate alkenes mainly at the unbranched side chain was a prerequisite for bacterial degradation of the added substrate. Degradation proceeded – in spite of the usually preferred terminal oxidation – via oxidation of the internal double bond and was followed by an ester cleavage. In turn, a series of alcohols was formed which were subsequently oxidized to the respective carboxylic acids and were further metabolized via the normal β-oxidation pathway.

Isolation of methanotrophic bacteria from termite gut.

The guts of termites feature suitable conditions for methane oxidizing bacteria (MOB) with their permanent production of CH4 and constant supply of O2 via tracheae. In this study, we have isolated MOB from the gut contents of the termites Incisitermes marginipennis, Mastotermes darwiniensis, and Neotermes castaneus for the first time. The existence of MOB was indicated by detecting pmoA, the gene for the particulate methane monooxygenase, in the DNA of gut contents. Fluorescence in situ hybridization and quantitative real-time polymerase chain reaction supported those findings. The MOB cell titer was determined to be 10(2)-10(3) per gut. Analyses of the 16S rDNA from isolates indicated close similarity to the genus Methylocystis. After various physiological tests and fingerprinting methods, no exact match to a known species was obtained, indicating the isolation of new MOB species. However, MALDI-TOF MS analyses revealed a close relationship to Methylocystis bryophila and Methylocystis parvus.

Application of MALDI-TOF-MS and nested SAPD-PCR for discrimination of Oenococcus oeni isolates at the strain level

Oenococcus oeni is the most important lactic acid bacterium during vinification and is frequently used as a commercial starter culture for malolactic fermentation. Since different strains exhibit a high heterogeneity concerning wine-related physiological characteristics, reliable methods for their differentiation at the strain level are needed, especially for the development of starter cultures. Microbial identification by using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) has increased over the last few years. In the present study, we used MALDI-TOF-MS for the differentiation of 11 O. oeni isolates originating from 5 different wine-growing regions. Furthermore, these strains were analyzed by a DNA fingerprinting method (nested specifically amplified polymorphic DNA (nSAPD)-PCR). Both methods supplied fast and reproducible results allowing a reliable differentiation of the O. oeni strains studied. MALDI-TOF-MS showed a slightly higher discriminatory ability, since two genetically very similar strains could be separated only by the application of this technique. However, nSAPD-PCR and the subsequent cluster analysis permitted a correlation between genetic similarities and the geographical origin of the strains.

The influence of cell counts, cell size, EPS and microbial inclusions on the lubrication properties of microorganisms

Metalworking fluids (MWF) play an essential role in many machining applications and can be classified e.g. according to DIN 51385 in non-water miscible- and water miscible metalworking fluids. Most of the several billion liters of the worldwide and annually consumed MWF are water-based and thus exposed to a microbial contamination, which leads to a deterioration of MWF compounds and therefore to a loss of quality and technical performance. To maintain the MWF quality, biocides are used to reduce the microbial load, regardless of their potential risk for health and environment. A further limitation poses the high consumption of raw materials (mainly mineral oils) for the production of MWF, with approximately 50,000 t of raw materials consumed annually in Germany. To overcome these limitations, the paradigm shift of using microorganisms as a lubricant in a manufacturing process is investigated in this paper. Preliminary investigations on a Brugger-tribotester indicated the great potential of microorganisms as replacement for conventional metalworking fluid components. Against this background, the approach presented here intends to investigate the influence of cell counts, cell size, extracellular polymeric substances and microbial storage products on the lubrication properties of selected microorganisms. The results of the tribological tests show, that some microorganism exhibit superior lubrication properties (up to 68xa0% higher) compared to a highly concentrated conventional metalworking fluid. Therefore, the potential of microorganisms to substitute conventional MWF components can be derived.

Pseudochelatococcus lubricantis gen. nov., sp. nov. and Pseudochelatococcus contaminans sp. nov. from coolant lubricants.

Two Gram-negative, rod-shaped, non-spore-forming bacteria, isolated from metal working fluids were investigated to determine their taxonomic positions. On the basis of 16S rRNA gene sequence phylogeny, both strains (MPA 1113(T) and MPA 1105(T)) formed a distinct cluster with 97.7u200a% sequence similarity between them, which was in the vicinity of members of the genera Methylobacterium, Camelimonas, Chelatococcus, Bosea, Salinarimonas and Microvirga to which they showed low sequence similarities (below 94u200a%). The predominant compounds in the polyamine pattern and in the quinone system of the two strains were spermidine and ubiquinone Q-10, respectively. The polar lipid profiles were composed of the major compounds: phosphatidylmonomethylethanolamine, phosphatidylglycerol, phosphatidylcholine, major or moderate amounts of diphosphatidylglycerol, two unidentified glycolipids and three unidentified aminolipids. Several minor lipids were also detected. The major fatty acids were either C19u200a:u200a0 cyclo ω8c or C18u200a:u200a1ω7c. The results of fatty acid analysis and physiological and biochemical tests allowed both, the genotypic and phenotypic differentiation of the isolates from each other, while the chemotaxonomic traits allowed them to be differentiated from the most closely related genera. In summary, low 16S rRNA gene sequence similarities and marked differences in polar lipid profiles, as well as in polyamine patterns, is suggestive of a novel genus for which the name Pseudochelatococcus gen. nov. is proposed. MPA 1113(T) (u200a=u200aCCM 8528(T)u200a=u200aLMG 28286(T)u200a=u200aCIP 110802(T)) and MPA 1105(T) (u200a=u200aCCM 8527(T)u200a=u200aLMG 28285(T)) are proposed to be the type strains representing two novel species within the novel genus, Pseudochelatococcus gen. nov., for which the names Pseudochelatococcus lubricantis sp. nov. and Pseudochelatococcus contaminans sp. nov. are suggested, respectively.

Alkali-labile precursors of dimethyl sulfide in marine benthic cyanobacteria

Abstract By the method of cold alkali hydrolysis, 29 marine benthic cyanobacteria were screened for production of alkali-labile precursors of dimethyl sulfide (DMS) including dimethylsulfoniopropionate (DMSP), a compound of significant importance in marine environments. Concentrations of DMS precursors ranged from undetectable to 0.8 mmol (g Chl a)–1. The data correspond to some previous investigations concerning DMSP content of marine cyanobacteria and suggest that marine benthic cyanobacteria are only minor producers of DMSP.