Quantitative screening method for hydrolases in microplates using pH indicators: determination of kinetic parameters by dynamic pH monitoring.

The presented pH-dyn assay serves as a versatile tool for screening enzymatically catalyzed reactions consuming or producing acids. The method is based on material balances of substrates and products. Ion balances relate concentrations of acids and bases to pH. pH-changes caused by the enzymatically catalyzed reaction in a well-defined buffer system are recorded by light-absorption measurements of a pH-indicator. Kinetic parameters are estimated by fitting the modeled pH changes to the experimentally observed ones. The enzymatically catalyzed hydrolysis of 4-nitrophenol is used as a model system. A pH indicator, bromothymol blue, is used to monitor the reaction progress. The reaction is monitored until the limiting substrate is completely consumed. This allows the estimation of the parameters of the Michaelis-Menten kinetics, K(M) and k(cat), in a single run. The results agree well with conventional spectrophotometric experiments and values reported in literature. Around pH 7, environmental CO(2) influences pH. Carbon dioxide influence was included in the model. Thus it was possible to estimate initial CO(2) concentrations as a model parameter, and therefore automatic correction for the CO(2) disturbances was achieved. This was important to detect low conversions at low buffer concentrations.     

Mitochondrial phylogeography and population history of pine martens Martes martes compared with polecats Mustela putorius

The flora and fauna of Europe are linked by a common biogeographic history, most recently the Pleistocene glaciations that restricted the range of most species to southern refugial populations. Changes in population size and migration, as well as selection, have all left a signature on the genetic differentiation. Thus, three paradigms of postglacial recolonization have been described, inferred from the patterns of DNA differentiation. Yet some species, especially wide-ranging carnivores, exhibit little population structuring between the proposed refugia, although relatively few have been studied due to the difficulty of obtaining samples. Therefore, we investigated mitochondrial variation in pine martens, Martes martes, in order to understand the extent to which they were affected by glacial cycles, and compared the results with an analysis of sequences from polecats, Mustela putorius. A general lack of ancient lineages, and a mismatch distribution that is consistent with an expanding population, is evidence that the present-day M. martes and Mu. putorius in central and northern Europe colonized from a single European refugium following a recent glaciation. There has also been interspecific mitochondrial introgression between M. martes and the sable M. zibellina in Fennoscandia.     

The Kiss of Death: Chagas Disease in the Americas

The Kiss of Death: Chagas Disease in the Americas. Joseph William Bastien. Salt Lake City: University of Utah Press. 1998. 301 pp.     

Parasympathetic innervation of canine tracheal smooth muscle

To investigate whether efferent parasympathetic fibers to the trachealsmooth muscle course through the pararecurrent nerve rather than therecurrent or the superior laryngeal nerve, we stimulated all threenerves in anesthetized dogs. We also recorded the pararecurrentnerve activity response to bronchoconstrictor stimuli and compared itwith pressure changes inside a saline-filled cuff of an endotrachealtube. Electrical stimulation (30 s, 100 Hz, 0.1 ms, 10 mA) increasedtracheal cuff pressure by 21.0 ± 3.2 and 1.3 ± 0.7 cmH₂O for the pararecurrent and the recurrent laryngealnerve, respectively. Stimulation of the superior laryngeal nerveincreased tracheal cuff pressure before, but not after, sectioning ofthe ramus anastomoticus, which connects it to the pararecurrent nerve.Intravenous administration of sodium cyanide increased pararecurrentnerve activity by 208 ± 51% and tracheal cuff pressure by14.4 ± 3.5 cmH₂O. Elevation of end-tidalPCO₂ to 50 Torr increased pararecurrent nerveactivity by 49 ± 19% and tracheal cuff pressure by 8.4 ± 3.6 cmH₂O. Further elevation to 60 Torr increasedpararecurrent nerve activity by 101 ± 33% and tracheal cuffpressure by 11.3 ± 2.9 cmH₂O. These results lead usto the conclusion that parasympathetic efferent fibers reach the smoothmuscle of the canine trachea via the pararecurrent nerve.