Improved micro-method for the high-performance liquid chromatographic determination of caffeine and paraxanthine in biological fluids

A high-performance liquid chromatographic procedure is reported for reproducibly and sensitively quantitating caffeine and its N-demethylated metabolite paraxanthine in micro-samples. A 5-μm reversed-phase radial compression column and 214-nm fixed wavelength ultraviolet detector were used to attain a sensitivity sufficient to quantitate these compounds at concentratios as low as 80 ng/ml using only 25 μl of sample. The assay is applicable to microliter samples of whole blood, serum, plasma, saliva, amniotic, cerebro-spinal and gastric fluids such as might be obtained in studies involving small animals or neonates. The utility of the assay is illustrated with caffeine and paraxanthine levels measured in several maternal and fetal fluids following constant-rate intravenous infusion of caffeine into a rabbit throughout pregnancy.     

Zeitaufwand für das Ernten und Verstecken von Kiefernsamen bei Dünnschn?bligen Tannenh?hern (Nucifraga caryocatactes macrorhynchos) im Fernen Osten Russlands

Zusammenfassung In einem Vorkommen des Dünnschnäbligen Tannenhähers an der Küste des Ochotskischen Meeres im Fernen Osten Sibiriens wurde das Ernten und Verstecken von Samen aus den Zapfen der Zwergzirbelkiefer (Pinus pumila) untersucht. Der Inhalt von durchschnittlich 2,8 Zapfen, das sind etwa 80 Samen, wurde in der gefüllten Kehltasche transportiert und auf eine Anzahl unter niedriger Zwergstrauchvegetation gelegener Bodenverstecke verteilt. Die Verstecke wurden in annähernd linearer Anordnung ohne Bevorzugung einer bestimmten Himmelsrichtung angelegt. Die Versteckserien enthielten im Median 79, maximal mehr als 120 Samen, das Einzelversteck durchschnittlich 19,6 Samen. Das Ernten und Leeren eines Zapfens geschah im Schnitt innerhalb von 47 s. Für das Verstecken einer Füllung des Kehlsacks benötigten die Vögel ca. 170 s. Für das gesamte Beschaffen und Verstecken eines einzelnen Kiefernsamens errechnet sich ein durchschnittlicher Zeitbedarf von 3,26 s. Nach 20 Tagen war der Zapfenvorrat in der lokalen Kiefernpopulation erschöpft. Jeder Häher hat nach den Hochrechnungen bis zu 100.000 Samen vergraben.

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Harvesting and caching capacities of Thin-billed Nutcrackers in the Russian Far East

Summary At the Ochotskian sea coast Thin-billed Eurasian Nutcrackers (Nucifraga caryocatactes macrorhynchos) harvested seeds of ripe cones of the brush pinePinus pumila in late summer. The mean number of seeds carried in their sublingual pouch was 80, which respresents the harvestable contents of 2.8 cones. These were distributed in an average of 5 caches, exclusively in the soil under low tundra vegetation. Caches were organized in nearly straight lines. Series contained a mean of 82.7 seeds, single caches a mean of 19.6 seeds. Plucking one cone and harvesting its seeds took 47 seconds on average. The caching of a complete pouchful took on average 123.4 seconds. The time invested for harvesting and caching one single seed was calculated at 3.26 seconds. Within three weeks in July, an average individual bird was calculated to have cached a total of up to 100,000 seeds.

     

Dielectric spectroscopy for noninvasive examination of corneal tissue]

Dielectric spectroscopy is a non-invasive contact technique that permits the in vivo measurement of the specific electrical properties of biological tissue induced by an external electrical field. Permittivity, relaxation time and specific conductivity as a function of corneal hydration (wet weight/dry weight) and temperature were measured in 10 porcine corneas. Variation of tissue hydration has a minor influence on the signal, with a significant variation of the signal being detectable only for relatively dry tissue. A much greater influence was found for temperature, in particular on relaxation times. Dielectric spectroscopy provides us with an opportunity to detect structural, in particular temperature-induced, changes in living tissue. In the frequency range investigated, hydration has only a small influence on the dielectric properties of the tissue.     

Die Entwicklung der Laut?u?erungen bei der Graugans (Anser anser)

Zusammenfassung Das Lautrepertoire von sechs handaufgezogenen Graugänsen wurde in individuellen Längsschnitten vom Schlüpfen bis zum Alter von 100 Tagen mit sonagraphischer Methode analysiert. Die Einteilung des Repertoires erfolgte nach drei verschiedenen Kriterien: Klang, Sonagramm und Kontext. Im jugendlichen Repertoire ließen sich 4 Rufklassen voneinander trennen. Im Laufe der Jugendentwicklung veränderten sich vor allem die phonetischen Parameter der Rufe, während die syntaktischen weitgehend konstant blieben. Nach einem Anstieg in der ersten Woche fiel die Tonhöhe der Rufe bei allen untersuchten Individuen und Rufklassen gleichmäßig ab. Der Stimmbruch, meßbar an der Zunahme geräuschhafter Komponenten, war ein kontinuierlicher, langfristiger Prozeß. Die Individualität war nicht in einem einzelnen, wohl aber in der Kombination mehrerer Merkmale faßbar. Die Geschlechter waren schon früh an Unterschieden in der Tonhöhe erkennbar. An sonstigen ontogenetischen Veränderungen innerhalb des Untersuchungszeitraumes konnten die Erweiterung des Repertories durch Aufspaltung und Reifung sowie ein möglicher Verlust von Rufklassen beobachtet werden.

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Juvenile development of vocalizations in Greylag Geese (Anser anser)

Summary The juvenile vocal development of six Greylag Geese from hatching up to the age of 100 days was analysed by using sonagraphic methods. The calls were classified by means of the acoustic phenomenon, sonagrams, and context criteria. The juvenile repertoire included four main call types. During ontogeny, changes of call parameters mainly referred to phonetic characteristics, whereas syntactic parameters remained more or less constant. The frequency pitch of calls changed to higher levels during the first week of development while in older goslings this parameter decreased constantly. This held true for each individual and call type. Breaking of the voice, defined as an increment of noisy parts in the vocalizations, is described as a continuous long term process. Structural individuality of calls apparently was not based on one single parameter, but on the combination of several ones. Sex differences were found in the pitch of vocalizations, correlating with different body weights of male and female geese. Additional developmental processes of the vocal repertoire included differentiation, maturation of new call types, and apparent suppression of juvenile ones. The functional organization of the juvenile call system in Greylags is discussed.

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Herrn Dr. Hans Löhrl zum 75. Geburtstag gewidmet     

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) enhances antibody production and protein kinase activity in murine B cells

Treatment of murine spleen cells with 30 nM TCDD resulted in an approximately 3 fold increase in unstimulated antibody production after 3 days in culture. This response was not accompanied by increased cellular proliferation and may represent an effect of TCDD on B cell activation or differentiation. Since PMA is capable of activating B cells, presumably via PKC, we have compared the effects of PMA and TCDD on protein kinase activation and phosphorylation of endogenous proteins in a highly purified preparation of B cells. In contrast to a reduction of cytosolic PKC activity, the expected effect of PMA, TCDD caused an increase in basal kinase activity with no effect on PKC activity. Addition of either PMA or TCDD resulted in enhanced phosphorylation of a similar profile of proteins, including proteins of Mr 12.2, 14.6, 29.2, 52.3 and 62.7 KDa. Addition of TCDD also resulted in the increased phosphorylation of a protein of Mr 45.2, which was unaffected by PMA. Combined treatment with PMA and TCDD resulted in additive responses. The additive effects of PMA and TCDD suggest an interaction at the level of protein phosphorylation which is mediated by different kinases. Therefore, TCDD may be stimulating B cells via an early effect on an unidentified protein kinase.     

Efficient oligonucleotide-directed construction of mutations in expression vectors by the gapped duplex DNA method using alternating selectable markers.

An efficient method for the construction of multiple mutations in a sequential manner is described. It is based on the gapped duplex DNA approach to oligonucleotide-directed mutagenesis (Kramer et al. 1984, Nucl. Acids Res. 12, 9441-9456) and a set of newly constructed phasmid vectors. These are characterized by the following features. Presence of the phage fl replication origin permits ready conversion to the single stranded DNA form. An amber mutation within, alternatively, the bla or cat gene provides a means for ready selection of the strand into which the mutagenic oligonucleotide has been incorporated. By means of the alternating antibiotic resistance markers any number of mutations can be constructed in consecutive rounds of mutagenesis. The optional presence of gene expression signals allows the direct overproduction of structurally altered proteins without re-cloning. Both the mutagenesis and expression aspects were tested using the lacZ gene as a model.     

Structure determination of the UDP-disaccharide fragment of cytoplasmic cofactor isolated from Methanobacterium thermoautotrophicum

The methylcoenzyme M methylreductase reaction has an absolute requirement for 7-mercaptoheptanoylthreonine phosphate or component B, which is the active component of the intact molecule previously referred to as cytoplasmic cofactor. A hydrolytic fragment of cytoplasmic cofactor has been purified and identified as uridine 5'-(O-2-acetamido-2-deoxy-beta-manno-pyranuronosyl acid (1----4)-2-acetamido-2-deoxy-alpha-glucopyranosyl diphosphate) by high resolution NMR and fast atom bombardment mass spectro-metry. It is postulated that UDP-disaccharide may function to anchor 7-mercaptoheptanoyl threonine phosphate at the active site of the methyl-reductase enzyme complex.     

The phosphorylation state of the reticulocyte 90-kDa heat shock protein affects its ability to increase phosphorylation of peptide initiation factor 2 alpha subunit by the heme-sensitive kinase

The rabbit reticulocyte Mr 90,000 protein associated with the heme-sensitive eIF-2 alpha kinase has been identified previously as the mammalian heat shock protein of this size class (hsp 90). Purified reticulocyte hsp 90 when added exogenously to the kinase increases its activity. This stimulatory effect is abolished after incubation of hsp 90 with a highly purified type 1 phosphoprotein phosphatase isolated from reticulocytes. Phosphorylation of dephosphorylated hsp 90 by casein kinase II but not by cAMP-dependent protein kinase restores the biological activity of hsp 90 to stimulate eIF-2 alpha phosphorylation.