Ralf Kautenburger

Staphylococcus aureus ClpC ATPase is a late growth phase effector of metabolism and persistence

Staphylococcus aureus Clp ATPases (molecular chaperones) alter normal physiological functions including an aconitase‐mediated effect on post‐stationary growth, acetate catabolism, and entry into death phase (Chatterjee et al., J. Bacteriol. 2005, 187, 4488–4496). In the present study, the global function of ClpC in physiology, metabolism, and late‐stationary phase survival was examined using DNA microarrays and 2‐D PAGE followed by MALDI‐TOF MS. The results suggest that ClpC is involved in regulating the expression of genes and/or proteins of gluconeogenesis, the pentose‐phosphate pathway, pyruvate metabolism, the electron transport chain, nucleotide metabolism, oxidative stress, metal ion homeostasis, stringent response, and programmed cell death. Thus, one major function of ClpC is balancing late growth phase carbon metabolism. Furthermore, these changes in carbon metabolism result in alterations of the intracellular concentration of free NADH, the amount of cell‐associated iron, and fatty acid metabolism. This study provides strong evidence for ClpC as a critical factor in staphylococcal energy metabolism, stress regulation, and late‐stationary phase survival; therefore, these data provide important insight into the adaptation of S. aureus toward a persister state in chronic infections.

Influence of metal concentration and the presence of competing cations on europium and gadolinium speciation with humic acid analysed by CE-ICP-MS

Nowadays, there is a broad consensus on the technical merits of the disposal of high-level nuclear waste (HLW) in deep and stable geological formations. For the safety assessment of a waste disposal it is important to understand the radionuclide migration in the near- and far-field of a repository caused by an incident. In this environment, natural organic matter such as humic substances can play an important role by their complexation behaviour for metal ions. Capillary electrophoresis hyphenated with inductively inductively coupled plasma mass spectrometry (CE-ICP-MS) have been used to study the complexation behaviour of Eu and Gd (as homologues of the actinides americium and curium) with humic acid. The influence of lanthanide concentration as well as the presence of competing cations like Ca, Mg and Al on the HA-complexation have been analysed. The lanthanide speciation by CE-ICP-MS reveals weak and strong HA binding sites for the used trivalent lanthanides subject to the given lanthanide concentration. The influence of the competing alkaline earth ions can be assumed as relevant at very high concentrations only while aluminium at already low concentration represents a strong competitor to Eu and Gd in HA-complexation, and may affect toxic metal speciation and thus metal mobility in the geological barrier of a future disposal.

Complexation of europium and uranium by humic acids analyzed by capillary electrophoresis-inductively coupled plasma mass spectrometry.

Investigations of the mobility of radioactive and nonradioactive substances in the environment are important tasks for the development of a future disposal in deep geological formations. Dissolved organic matter (DOM) can play an important role in the mobilization of metal ions due to complexation. In this study, we investigate the complexation behavior of humic acid (HA) as a model substance for DOM and its influence on the migration of europium as homologue for the actinide americium and uranium as the principal component of nuclear fuel. As speciation technique, capillary electrophoresis (CE) was hyphenated with inductively coupled plasma mass spectrometry (ICP‐MS). For the study, 0.5 mg·L−1 of the metals and 25 mg·L−1 of (purified Aldrich) HA and an aqueous solution sodium‐perchlorate with an ionic strength of 10 mM at pH 5 were used. CE–ICP‐MS clearly shows the different speciation of the triple positively charged europium and the double positively charged uranyl cation with HA.

Influence of metal loading and humic acid functional groups on the complexation behavior of trivalent lanthanides analyzed by CE-ICP-MS

The complexation behavior of Aldrich humic acid (AHA) and a modified humic acid (AHA-PB) with blocked phenolic hydroxyl groups for trivalent lanthanides (Ln) is compared, and their influence on the mobility of Ln(III) in an aquifer is analyzed. As speciation technique, capillary electrophoresis (CE) was hyphenated with inductively coupled plasma mass spectrometry (ICP-MS). For metal loading experiments 25 mg L(-1) of AHA and different concentrations (cLn(Eu+Gd)=100-6000 μg L(-1)) of Eu(III) and Gd(III) in 10mM NaClO4 at pH 5 were applied. By CE-ICP-MS, three Ln-fractions, assumed to be uncomplexed, weakly and strongly AHA-complexed metal can be detected. For the used Ln/AHA-ratios conservative complex stability constants log βLnAHA decrease from 6.33 (100 μg L(-1) Ln(3+)) to 4.31 (6000 μg L(-1) Ln(3+)) with growing Ln-content. In order to verify the postulated weaker and stronger humic acid binding sites for trivalent Eu and Gd, a modified AHA with blocked functional groups was used. For these experiments 500 μg L(-1) Eu and 25 mg L(-1) AHA and AHA-PB in 10mM NaClO4 at pH-values ranging from 3 to 10 have been applied. With AHA-PB, where 84% of the phenolic OH-groups and 40% of the COOH-groups were blocked, Eu complexation was significantly lower, especially at the strong binding sites. The log β-values decrease from 6.11 (pH 10) to 5.61 at pH 3 (AHA) and for AHA-PB from 6.01 (pH 7) to 3.94 at pH 3. As a potential consequence, particularly humic acids with a high amount of strong binding sites (e.g. phenolic OH- and COOH-groups) can be responsible for a higher metal mobility in the aquifer due to the formation of dissolved negatively charged metal-humate species.

Influence of geochemical parameters on the sorption and desorption behaviour of europium and gadolinium onto kaolinite

In this study we investigated the sorption and desorption behaviour of europium and gadolinium (homologues of the actinides americium and curium) onto the clay mineral kaolinite KGa-1b. In the model system metal/kaolinite, sorption isotherms and pH-edges were determined in different batch experiments. Calcium and magnesium as competing cations were used to simulate the influence of water hardness on the sorption and desorption processes. After centrifugation, the free metal ions in the supernatant solution were analysed by ICP-MS. With increasing lanthanide concentration, especially the relative desorption of these metals from kaolinite rises significantly before the cation exchange capacity (CEC) of 0.94 meq per 100 g is reached. This indicates that low lanthanide concentrations will result in a relatively lower metal migration due to sorption reactions at higher active sites in comparison with higher lanthanide concentrations. The K(d)-values and sorption isotherms of Eu(iii) and Gd(iii) were determined at pH 5.0 (+/-0.02) and analysed with Freundlich and Langmuir sorption models. The experimental data can best be fitted by Langmuir sorption isotherm. The pH-value has only a minor influence on the sorption onto kaolinite. Only at low pH-values (<pH 4), relevant changes in migration behaviour have to be expected. One important influence factor on lanthanide sorption found in our study is naturally occurring competing cations such as alkaline earth metals. With Ca and Mg concentrations close to those found in nature, lanthanide sorption decreases dramatically. Desorption experiments show that after four consecutive equilibration steps, nearly all the lanthanide has been mobilised from kaolinite to the aquifer.

Waste Disposal in Clay Formations: Influence of Humic Acid on the Migration of Heavy‐Metal Pollutants

The deep geological disposal of high-level radioactive waste is assumed to be a way of providing adequate protection for humans and the environment. For the safe isolation of radioactive waste, a low permeability and high sorption capacity of the geological barrier is important. Clay minerals play a major role in different concepts for the disposal of high-level nuclear waste in deep geological formations. Geological clay formations are considered to be a host rock for waste repository. Natural clays can contain natural organic matter, for example, humic substances. In most aquatic systems, components of natural organic matter such as humic acids can act as ligands for complexing metal ions and may affect the speciation and thus the mobility of the metal. By forming soluble complexes and colloids with numerous toxic heavy metals, including radio ACHTUNGTRENNUNGnuclides, humic acids can influence the migration behavior of these pollutants in hematite or geological clay formations, as reported, for example, for bentonite, kaolinite, silica, and montmorillonite. Kaolinite is a significant component of many soils and fracture-filling materials of crystalline hard rock. These clay formations are part of the natural barrier protecting the environment from the possible impact of radionuclide contamination. Herein we report the sorption behavior of the trivalent lanthanides europium and gadolinium (as homologues of the actinides americium and curium), under conditions close to those of nature, in a binary system consisting of these heavy metals and well-crystallized kaolinite (KGa-1b) as a model mineral, and in the ternary system metal/kaolinite/humic acid. The influence of the concentration of lanthanide ion, pH value, or concentrations of different competing metal cations (water hardness, represented by calcium and magnesium) on the sorption of the lanthanide ion onto kaolinite in the presence or absence of humic acid was investigated. After the sorption experiments, free metal ions in the supernatant solution were analyzed by inductively coupled plasma mass spectrometry (ICP-MS). Speciation of the lanthanide–humic acid complexes are reported elsewhere. The ternary system described in Figure 1 is most suitable to study metal migration under natural conditions. Research on the three binary systems can be seen as preliminary tests to understand partial aspects of the complete system. With humic acid, there is the possibility that metal humates could adsorb on the kaolinite surface through hydrophobic interactions, thus increasing retention of the lanthanide ion. Negatively charged complexes could inhibit sorption of the lanthanide and accelerate migration through clay minerals. Sorption behavior is strongly dependent on the experimental conditions as tests performed in the ternary system clearly show. The experiments that were performed here are based on the system metal/kaolinite with humic acid as an additional influencing factor. With humic acid present, the sorption of the lanthanide ions onto kaolinite is suppressed, especially at low concentrations of Eu and Gd (Figure 2). As lanthanides are complexed by humic acid and as there are not enough positively charged ions present to neutralize the complexes at pH 5, a significant amount of the complexes stays in solution. With rising lanthanide concentration, which also means a higher amount of metal complexed by humic acid, the humic acid partially loses its negative charge and is able to adsorb on the kaolinite surFigure 1. The ternary system, consisting of lanthanide ions, humic acid, and kaolinite, and important influential factors.

New approach of a transient ICP-MS measurement method for samples with high salinity

In the near future it is necessary to establish a disposal for high level nuclear waste (HLW) in deep and stable geological formations. In Germany typical host rocks are salt or claystone. Suitable clay formations exist in the south and in the north of Germany. The geochemical conditions of these clay formations show a strong difference. In the northern ionic strengths of the pore water up to 5M are observed. The determination of parameters like Kd values during sorption experiments of metal ions like uranium or europium as homologues for trivalent actinides onto clay stones are very important for long term safety analysis. The measurement of the low concentrated, not sorbed analytes commonly takes place by inductively coupled plasma mass spectrometry (ICP-MS). A direct measurement of high saline samples like seawater with more than 1% total dissolved salt content is not possible. Alternatives like sample clean up, preconcentration or strong dilution have more disadvantages than advantages for example more preparation steps or additional and expensive components. With a small modification of the ICP-MS sample introduction system and a home-made reprogramming of the autosampler a transient analysing method was developed which is suitable for measuring metal ions like europium and uranium in high saline sample matrices up to 5M (NaCl). Comparisons at low ionic strength between the default and the transient measurement show the latter performs similarly well to the default measurement. Additionally no time consuming sample clean-up or expensive online dilution or matrix removal systems are necessary and the analysation shows a high sensitivity due to the data processing based on the peak area.

Local Effects on Airway Inflammation and Systemic Uptake of 5 nm PEGylated and Citrated Gold Nanoparticles in Asthmatic Mice

Nanotechnology is showing promise in many medical applications such as drug delivery and hyperthermia. Nanoparticles administered to the respiratory tract cause local reactions and cross the blood-air barrier, thereby providing a means for easy systemic administration but also a potential source of toxicity. Little is known about how these effects are influenced by preexisting airway diseases such as asthma. Here, BALB/c mice are treated according to the ovalbumin (OVA) asthma protocol to promote allergic airway inflammation. Dispersions of polyethylene-glycol-coated (PEGylated) and citrate/tannic-acid-coated (citrated) 5 nm gold nanoparticles are applied intranasally to asthma and control groups, and (i) airway resistance and (ii) local tissue effects are measured as primary endpoints. Further, nanoparticle uptake into extrapulmonary organs is quantified by inductively coupled plasma mass spectrometry. The asthmatic precondition increases nanoparticle uptake. Moreover, systemic uptake is higher for PEGylated gold nanoparticles compared to citrated nanoparticles. Nanoparticles inhibit both inflammatory infiltrates and airway hyperreactivity, especially citrated gold nanoparticles. Although the antiinflammatory effects of gold nanoparticles might be of therapeutic benefit, systemic uptake and consequent adverse effects must be considered when designing and testing nanoparticle-based asthma therapies.

Recovery rate and homogeneity of doping europium into luminescent metal hydrides by chemical analysis

During the investigation of concentration dependent properties of phosphors, such as emission intensities, the knowledge of the activator ion concentration is of great importance. Herein we present a study on recovery rate and homogeneity of the activator ion concentration by chemical analysis in luminescent europium doped metal hydrides. The analysis method was established on the model system EuxSr1−xH2 and applied to brightly emitting hydridic perovskites LiMH3:Eu2+ and LiMD3:Eu2+ (M = Sr, Ba). The nominal activator ion concentrations calculated from initial weights are in good agreement with those determined via ICP-MS for 10−3% ≤ x both for the hydrides (deuterides) and the parent alloys. The synthesis protocol thus allows a reliable method for reproducibly doping europium into metal hydrides. Luminescence spectra of LiEuxSr1−xH3 with x = 0.0037% show a broad band emission at room temperature typical for Eu2+ 4f65d1–4f7 transitions. Below 80 K, a vibronic fine structure is observed with vibrational coupling frequencies of approximately 100, 370 and 970 cm−1.

Speciation via Hyphenation-Metal Speciation in Geological and Environmental Samples by CE-ICP-MS

This review deals with different aspects of analytical speciation methods developed for metals and metal-containing biomolecules or metal complexes with natural organic and inorganic matter in geological and environmental samples. An overview of recent studies dealing with metal speciation in various geological and environmental sample matrices is presented. The focus of analytical speciation methods is limited to capillary electrophoresis (CE) hyphenated with Inductively Coupled Plasma (ICP) detection techniques. The main aspects discussed in this review are the separation and detection step for the metal species. Possible influences of the applied separation technique on the metal speciation have been considered. Prominent exampes are, the metal-NOM (natural organic matter) complex stability during the separation step by the use of CE or the problem of ion-exchange and adsorption processes in the case of silica tubes as CE-capillaries. Additionally, different matrix effects in ICP-MS analysis will be discussed. Furthermore, the major drawback of speciation analysis is the lack of suitable reference or standard materials for the purpose of quality assurance. Therefore, quantification and verification of separation efficiency and accuracy for both the separation and detection step of the species have been evaluated, too.