The hydrolysis of triglycerides by immobilized lipase in a hydrophilic membrane reactor

In the present article a method is described to immobilize lipase from Candida rugosa on a hollow fiber membrane, and the use of such a system for the hydrolysis of lipids is reported. The membranes were ENKA hydro philic Cuprophan((R))-type hollow fibers, having a large specific surface area. The immobilized lipase exhibited a high stability: the half-life time was 43 days at a temperature of 30 degrees C. Furthermore, it is proved that kinetic studies can be carried out with this system, operated in a batch or continuous mode. The relation between conversion rate and degree of hydrolysis was determined. On this basis, a dynamic model of the process was developed that describes the relation between reaction conditions and the conversion rate.     

Compartmentation of glutamate metabolism in brain. Evidence for the existence of two different tricarboxylic acid cycles in brain

1. (14)C from [1-(14)C]glucose injected intraperitoneally into mice is incorporated into glutamate, aspartate and glutamine in the brain to a much greater extent than (14)C from [2-(14)C]glucose. This difference for [1-(14)C]glucose and [2-(14)C]glucose increases with time. The amount of (14)C in C-1 of glutamate increases steadily with time with both precursors. It is suggested that a large part of the glutamate and aspartate pools in brain are in close contact with intermediates of a fast-turning tricarboxylic acid cycle. 2. (14)C from [1-(14)C]acetate and [2-(14)C]acetate is incorporated to a much larger extent into glutamine than into glutamate. An examination of the time-course of (14)C incorporated into glutamine and glutamate reveals that glutamine is not formed from the glutamate pool, labelled extensively by glucose, but from a small glutamate pool. This small glutamate pool is not derived from an intermediate of a fast-turning tricarboxylic acid cycle. 3. It is proposed that two different tricarboxylic acid cycles exist in brain.     

The ubiquitin conjugation system is required for ligand-induced endocytosis and degradation of the growth hormone receptor.

The ubiquitin-dependent protein degradation system has recently been implicated in downregulation of signal transducing receptors. Growth hormone receptor (GHR) cDNA was transfected into Chinese hamster ovary cells, which exhibit a temperature-sensitive defect in ubiquitin conjugation (CHO-ts20), as well as into wild-type cells (CHO-E36). Upon binding of growth hormone (GH), two GHR polypeptides dimerize and initiate signal transduction. In CHO-E36 and in CHO-ts20 at the permissive temperature the GHR was ubiquitinated and degraded in a GH-dependent fashion. However, at the non-permissive temperature in CHO-ts20 cells, neither GH-dependent uptake nor degradation of the GHR was observed, while in CHO-E36 cells both GHR uptake and degradation were accelerated. Incubation of CHO-E36 cells with inhibitors of endosomal/lysosomal function (NH4Cl, bafilomycin A1) markedly reduced ligand-induced GHR degradation. Our results indicate that a functional ubiquitin conjugating system is required for GH-induced endocytosis and that degradation of both the exoplasmic and cytoplasmic portions of the GHR occurs within the endosomal/lysosomal compartment.     

Reductive Dehalogenation of Brominated Phenolic Compounds by Microorganisms Associated with the Marine Sponge Aplysina aerophoba

Marine sponges are natural sources of brominated organic compounds, including bromoindoles, bromophenols, and bromopyrroles, that may comprise up to 12% of the sponge dry weight. Aplysina aerophoba sponges harbor large numbers of bacteria that can amount to 40% of the biomass of the animal. We postulated that there might be mechanisms for microbially mediated degradation of these halogenated chemicals within the sponges. The capability of anaerobic microorganisms associated with the marine sponge to transform haloaromatic compounds was tested under different electron-accepting conditions (i.e., denitrifying, sulfidogenic, and methanogenic). We observed dehalogenation activity of sponge-associated microorganisms with various haloaromatics. 2-Bromo-, 3-bromo-, 4-bromo-, 2,6-dibromo-, and 2,4,6-tribromophenol, and 3,5-dibromo-4-hydroxybenzoate were reductively debrominated under methanogenic and sulfidogenic conditions with no activity observed in the presence of nitrate. Monochlorinated phenols were not transformed over a period of 1 year. Debromination of 2,4,6-tribromophenol, and 2,6-dibromophenol to 2-bromophenol was more rapid than the debromination of the monobrominated phenols. Ampicillin and chloramphenicol inhibited activity, suggesting that dehalogenation was mediated by bacteria. Characterization of the debrominating methanogenic consortia by using terminal restriction fragment length polymorphism (TRFLP) and denaturing gradient gel electrophoresis analysis indicated that different 16S ribosomal DNA (rDNA) phylotypes were enriched on the different halogenated substrates. Sponge-associated microorganisms enriched on organobromine compounds had distinct 16S rDNA TRFLP patterns and were most closely related to the δ subgroup of the proteobacteria. The presence of homologous reductive dehalogenase gene motifs in the sponge-associated microorganisms suggested that reductive dehalogenation might be coupled to dehalorespiration.     

Detection and characterization of a dehalogenating microorganism by terminal restriction fragment length polymorphism fingerprinting of 16S rRNA in a sulfidogenic, 2-bromophenol-utilizing enrichment

Terminal restriction fragment length polymorphism analysis of reverse-transcribed 16S rRNA during periods of community flux was used as a tool to delineate the roles of the members of a 2-bromophenol-degrading, sulfate-reducing consortium. Starved, washed cultures were amended with 2-bromophenol plus sulfate, 2-bromophenol plus hydrogen, phenol plus sulfate, or phenol with no electron acceptor and were monitored for substrate use. In the presence of sulfate, 2-bromophenol and phenol were completely degraded. In the absence of sulfate, 2-bromophenol was dehalogenated and phenol accumulated. Direct terminal restriction fragment length polymorphism fingerprinting of the 16S rRNA in the various subcultures indicated that phylotype 2BP-48 (a Desulfovibrio-like sequence) was responsible for the dehalogenation of 2-bromophenol. A stable coculture was established which contained predominantly 2BP-48 and a second Desulfovibrio-like bacterium (designated BP212 based on terminal restriction fragment length polymorphism fingerprinting) that was capable of dehalogenating 2-bromophenol to phenol. Strain 2BP-48 in the coculture could couple reductive dehalogenation to growth with 2-bromophenol, 2,6-dibromophenol, or 2-iodophenol and lactate or formate as the electron donor. In addition to halophenols, strain 2BP-48 appears to use sulfate, sulfite, and thiosulfate as electron acceptors and is capable of simultaneous sulfidogenesis and reductive dehalogenation in the presence of sulfate.     

Characterizing man-made and natural modifications of microbial diversity and activity in coastal ecosystems

The impacts of growing coastal pollution and habitat alteration accompanying human encroachment are of great concern at the microbial level, where much of the ocean's primary production and biogeochemical cycling takes place. Coastal ecosystems are also under the influence of natural perturbations such as major storms and flooding. Distinguishing the impacts of natural and human stressors is essential for understanding environmentally-induced change in microbial diversity and function. The objective of this paper is to discuss the applications and merits of recently developed molecular, ecophysiological and analytical indicators and their utility in examining anthropogenic and climatic impacts on the structure and function of coastal microbial communities. The nitrogen-limited Neuse River Estuary and Pamlico Sound, North Carolina are used as examples of ecosystems experiencing both anthropogenic (i.e., accelerating eutrophication) and climatic stress (increasing frequencies of tropical storms and hurricanes). Additional examples are derived from a coastal monitoring site (LEO) on the Atlantic coast of New Jersey and Galveston Bay, on the Gulf of Mexico. In order to assess structure, function, and trophic state of these and other coastal ecosystems, molecular (DNA and RNA-based) characterizations of the microbial taxa involved in carbon, nitrogen and other nutrient transformations can be combined with diagnostic pigment-based indicators of primary producer groups. Application of these methods can reveal process-level microbial community responses to environmental variability over a range of scales. Experimental approaches combined with strategic monitoring utilizing these methods will facilitate: (a) understanding organismal and community responses to environmental change, and (b) synthesizing these responses in the context of ecosystem models that integrate physical, chemical and biotic variability with environmental controls. This revised version was published online in August 2006 with corrections to the Cover Date.     

Spatial distribution of total, ammonia-oxidizing, and denitrifying bacteria in biological wastewater treatment reactors for bioregenerative life support

Bioregenerative life support systems may be necessary for long-term space missions due to the high cost of lifting supplies and equipment into orbit. In this study, we investigated two biological wastewater treatment reactors designed to recover potable water for a spacefaring crew being tested at Johnson Space Center. The experiment (Lunar-Mars Life Support Test Project-Phase III) consisted of four crew members confined in a test chamber for 91 days. In order to recycle all water during the experiment, an immobilized cell bioreactor (ICB) was employed for organic carbon removal and a trickling filter bioreactor (TFB) was utilized for ammonia removal, followed by physical-chemical treatment. In this study, the spatial distribution of various microorganisms within each bioreactor was analyzed by using biofilm samples taken from four locations in the ICB and three locations in the TFB. Three target genes were used for characterization of bacteria: the 16S rRNA gene for the total bacterial community, the ammonia monooxygenase (amoA) gene for ammonia-oxidizing bacteria, and the nitrous oxide reductase (nosZ) gene for denitrifying bacteria. A combination of terminal restriction fragment length polymorphism (T-RFLP), sequence, and phylogenetic analyses indicated that the microbial community composition in the ICB and the TFB consisted mainly of Proteobacteria, low-G+C gram-positive bacteria, and a Cytophaga-Flexibacter-Bacteroides group. Fifty-seven novel 16S rRNA genes, 8 novel amoA genes, and 12 new nosZ genes were identified in this study. Temporal shifts in the species composition of total bacteria in both the ICB and the TFB and ammonia-oxidizing and denitrifying bacteria in the TFB were also detected when the biofilms were compared with the inocula after 91 days. This result suggests that specific microbial populations were either brought in by the crew or enriched in the reactors during the course of operation.     

Neuroticism in the elderly. The utility of the shortened DPQ-scales

This article reports on the relation between aging and personal adjustment. Current personality scales are not developed for older persons. Scales contain items which are not valid for an aging population and contain too many items for administration in older populations. As part of the Longitudinal Aging Study Amsterdam (LASA) Neuroticism in older persons was measured with a shortened version of the Inadequacy (IN) and Social Inadequacy (SI) scales of the Dutch Personality Questionnaire (DPQ). The utility of these shortened scales was assessed based on internal consistency, inter-item correlations, test-retest reliability and factor analysis. The consistency of the personality dimension Neuroticism was assessed based on cohort-differences and a 6-year longitudinal comparison. The research-population contained 2118 respondents at baseline, aged between 55 and 85 years, 49% were male and they were not living in an institution. The shortened scales appeared to be reliable and valid instruments to measure Neuroticism in the elderly. The gaining of time due to the administration of the shortened scales enlarges the feasibility of the scales for measuring Neuroticism in older persons. Results showed no significant age-difference on the IN-scale, but revealed a significant difference on the SI-scale (p < .01). The 65+ elderly (65-74 and 75-85) have higher scores on Social Inadequacy than the youngest elderly (55-64). Longitudinal analyses showed an interaction between age at baseline and the stability and change of the level of Neuroticism. On both scales the youngest age-group showed a significant decline in mean level of Neuroticism (p < .01). The mean level of Social Inadequacy in the oldest age-group showed an increase during the 6-year follow-up period (p < .05). However, the differences were very small. Future research is needed to assess the effect of related variables on Neuroticism in older persons.     

Identification of a new mutation,S305C,in exon 7 of the low-density lipoprotein receptor gene in a Brazilian family with homozygous familial hypercholesterolemia

DNA analysis of the low-density lipoprotein receptor gene of a young Brazilian man, suffering from an extreme form of hypercholesterolemia, revealed the presence of two mutations, thereby confirming the diagnosis of homozygous familial hypercholesterolemia (FH). The first mutation was a mutation frequently found in Brazilian patients with FH, termed C660X or FHLebanese. The second mutation, in which a serine residue was replaced by a cysteine at amino acid position 305 (S305C) was a new mutation never described before. S305C was inherited from the proband's mother, who was of Italian descent. The occurrence of LDL receptor gene mutations of Lebanese and Italian origin in Brazil underlines the genetic heterogeneity of the Brazilian population.