Band 39, Heft 7-8

Three violet terphenylquinone pigments, 4′-hydroxyphlebiarubrone (2), 4′,4″-dihydroxy-phlebiarubrone (3), 3′,4′,4″-trihydroxyphlebiarubrone (4), and the already known pigment phlebiarubrone (1) were isolated from cultures of Punctularia atropurpurascens. The structures of the phlebiarubrone derivatives were determined by spectral evidence. The structure of 3 was confirmed by synthesis from atromentin (6). Two strains of the genus Punctularia obtained from CBS, were also found to produce these compounds, whereas with a third strain only the production of 1 was observed. 3 did not exhibit any antimicrobial or cytotoxic activity. The other pigments showed weak antibacterial and cytotoxic activity.

The hydrocarbon fractions of the essential oils from leaves of Cistus ladanifer, C. palhinhae, C. laurifolius and C. monspeliensis were analysed by GLC and GCMS techniques.

A survey of chemical composition of forty-four species of Heterotropa growing in Japan and Formosa showed that four chemical groups could be distinguished and that two routes of distribution to Japan from China may be suspected on the basis of phenylpropane components.

Leaves of a number of Ericaceous plants have a thin epicuticular layer that consists mostly of triterpenes. In Kalmia angustifolia, K. latifolia and K. polifolia as well as in Gaultheria procumbens and G. shallon this material also contains trace amounts of flavonoid aglycones. In Kalmia they are the 3-OMe-derivatives of four C-methyl flavones reported previously as the typical leaf-wax flavonoids of Eucalyptus. The new compounds, 5-OH-3,7,4′-triOMe-6,8-diCH 3 -flavone (kalmiatin), 5-OH-3,7,4′-triOMe-6-CH 3 -flavone (8-desmethyl-kalmiatin), 5,4′-diOH-3,7-diOMe-6,8-diCH 3 -flavone (latifolin) and 5,4′-diOH-3,7-diOMe-6-CH 3 -flavone (8-desmethyl-latifolin) are novel natural products. They could not be detected in eight species of Eucalyptus. A screening of 15 species of Ericaceae revealed that 8-desmethyl-sideroxylin and 8-desmethyl-latifolin are present also in the epicuticular layer on leaves of Gaultheria procumbens, while on G. shallon and on Andromeda polifolia traces of galangin-3-methyl ether were found. In the other species checked no external flavonoid aglycones could be detected.

Soluble enzyme preparations from flower buds of Matthiola incana catalysed the conversion of dihydrokaempferol to kaempferol and of dihydroquercetin to quercetin. The reaction required 2-oxoglutarate, ascorbate and Fe 2+ as cofactors and had a pH-optimum at about 7.0. Highest enzyme activity was already present in the youngest buds followed by a rapid decline during bud and flower development. Furthermore, a substantial correlation was observed between the enzyme activity for flavonol formation and the flavonol content of the buds and flowers.

From an equimolar dipeptide/xanthosine mixture in water/methanol, only the nucleoside crystallized in anhydrous form, space group P2 1 2 1 2 1 with a = 17.406 (5) Å, b = 12.378 (4), c = 5.350 (2) Å, Z = 4, C 10 H 12 O 6 N 4 , FW = 284.26, D x = 1.639 g · cm -3 . The structure was determined by direct methods on the basis of 984 X-ray data and refined to an agreement index of R = 3.9%. Xanthosine occurs in the syn conformation with χ = -132.7° which is stabilized by an intramolecular hydrogen bond N (3) - H ... O(5′). The pseudorotation parameters for sugar puckering are P = 156° and τ m = 38.7° corresponding to C(2′)-endo form. The crystal packing of xanthosine in anhydrous and dihydrate modifications are very different, yet the molecular conformations are similar. In the anhydrous form, an unusual intramolecular hydrogen bond O (2′) - H ... O(3′) is observed.

Nitrogen dioxide, formed by γ-radiolysis in deaerated aqueous nitrate/nitrite solutions, is capable of oxidizing Gly-Tyr in favourable competition with the natural decay of NO 2 • by dimerization and disproportionation. 2,2′-Biphenolic tyrosine dimers and nitro-tyrosine were identified spectroscopically as stable products. The results suggest that NO 2 • reacts with the peptide by electron abstraction, generating Gly-Tr phenoxyl radicals (PheO • ) which terminate by dimerization (2 PheO • → 2,2′-biphenol) and NO 2 • -scavenging (PheO • + NO 2 • → Nitro-Tyr).

Three different methods for the determination of phenylalanine hydroxylase activity have been compared: a) Differential photometric assay of the increase in tyrosine concentration in the presence of phenylalanine; b) Product separation by thin layer chromatography and scintillation counting of the [ 14 C]tyrosine formed; c) HPLC separation and spectrofluorometric quantification of derivatized amino acids. A comparison of the activities of phenylalanine hydroxylase in rat liver and Euglena gracilis clearly showed that only rat liver contains this enzymic activity as shown by methods b) and c) although pseudo-activity of Euglena gracilis preparations was found during the spectrophotometric test a). The HPLC method proved to be the fastest, most reliable and convenient method for direct tyrosine determination and thus for measuring phenylalanine hydroxylase activity.

The enzymic hydroxylation of phenylalanine by phenylalanine hydroxylase (E.C. 1.14.16.1.) in vitro is dependent on the presence of hydrogen peroxide removing processes. The loss of phenylalanine hydroxylase activity can be prevented to the same extent by catalase as well as the presence of optimized amounts of both peroxidase and superoxide dismutase. Peroxidase alone exhibited only two third of the maximal protective effect of catalase whereas superoxide dismutase alone was not able to exert any protective influence on phenylalanine hydroxylase. These findings suggest that the termination of phenylalanine hydroxylation in the absence of hydrogen peroxide removing reactions is probably due to destructive oxygen species generated at the active site iron of phenylalanine hydroxylase in the presence of H 2 O 2 and the tetrahydropterin cofactor.

1 H double-quantum and COSY NMR-spectra of the porcine protein adenylate kinase have been obtained in order to assign aromatic spin systems within this protein, which contains seven tyrosyl, five phenylalanyl, and two histidyl residues. Double-quantum spectroscopy in its simple form allowed the assignment of five connectivities in the aromatic region and therefore proved inferior to COSY, which allowed the assignment of connectivities within nine aromatic residues.

Previous phytochrome models were based on experimental results obtained only with proteolytically degraded phytochrome. The reactivity of native phytochrome, in both the Pr and the Pfr form, towards 8-anilinonaphthalene-1-sulfonate (ANS), urea, and ethylene glycol is described here. Whereas native phytochrome in the Pfr form is only very slowly bleached by ANS, immediate bleaching occurs with ANS treated Pr during red irradiation. The bleached form, Pr 660 , can be converted into Pr 660 by prolonged far-red irradiation. A tentative model for native phytochrome is proposed which explains the effects of phototransformation, ANS binding and partial proteolysis.

The indol alkaloid gramine inhibited photophosphorylation, Pi-ATP exchange reaction and proton gradient, and enhanced electron transport in spinach thylakoids with I 50 around 0.2 mᴍ. It thus behaves as a typical uncoupler of photophosphorylation.

The incubation of erythrocyte ghosts with mixtures of vitamins A + D 3 , A + E, A + K 1 , produces decrease or increase of fluorescence anisotropy r of the DPH probe, depending on the vitamin/vitamin ratio. We found a correlation between the order parameter S and cell fusion phenomena observed by phase contrast microscopy.

The effects of auxin and cytokinin on light dependent carotenoid synthesis were studied in callus cultures derived from Ricinus endosperm tissue. Chloroplasts differentiate when calli are grown in the light in the presence of cytokinin and auxin. When cultures are transferred to an auxin-free medium, chromoplast differentiation is initiated, i.e. rhodoxanthin accumulates whereas lutein and chlorophyll content decrease and the plastid morphology changes from that of a typical chloroplast to that of a globular-type chromoplast. These changes are fully reverted by readdition of auxin. Plastid differentiation into either chloroplasts or chromoplasts in Ricinus endosperm cultures therefore appears to be controlled by auxin and cytokinin.

The protein complement of chloroplasts, prochromoplasts, chromoplasts, and mitochondria of the daffodil (Narcissus pseudonarcissus) has been investigated comparatively by gel electrophoresis. Mitochondria do not share common proteins with plastids. On the other hand, there is a broad overlap of the protein complement of the three plastid types investigated, in spite of their different fine structures. There are, however, remarkable differences regarding the relative amounts of many protein species, and certain plastid proteins are entirely absent either from the chloroplasts (e.g., 2 chromoplast proteins in the 60 kDa range) or the chromoplasts (e.g., LHCP). The protein complement of prochromoplasts, which contain chlorophylls and the LHCP, is nevertheless quite more similar to the protein pattern of the chromoplasts than to the one of the chloroplasts.

At lower concentrations, polyarginine, polylysine, protamine, histones H 1, H2A, H2B and H3 cause lysis of human erythrocytes, whereas at higher concentrations inner histones are not hemolytic but induce only surface condensation and alterations in the cell-shape. Antibody coated erythrocytes treated with polyarginine result in ghost-like spheres having globular bodies 1 μm in diameter on the surface. Human fibroblasts and lymphocytes, and Ehrlich ascites cells treated with polyarginine also form surface globular bodies similar in size. Nucleate cell-poly-arginine mixtures with lower polycation doses result in cytolysis, while higher polycation doses produce pyknosis of the cell surface accompanied by reorganization of a membrane-like structure. Changes in spectrofluorometric values result from 1,6 -diphenyl-1,3,5-hexatrien binding to cell lipids, match the plasma membrane alterations. Reciprocal shake incubation am plifies and/or conditions these polycation-induced alterations. The homogeneity of pyknotic surface bodies and the apparent polycationic membrane reorganization requiring oscillatory friction forces suggest the preexistence of a multizonal glycocalyx distribution corresponding to plasma membrane compartments. The possible role of this compartmentalization in receptor and membrane recycling, as well as the involvement of reversible catalytic-like polycation condensation in macromolecular changes are discussed.

Polymyxin B produced dose dependent changes in the surface topography of the goat erythrocyte cells. Transformation from the normal biconcave discs through crenated structures to the final rounded or spherical shape was recorded by scanning electron microscopy. A maxim um of three to four crenations per cell was recorded corresponding to a polymyxin dose of 15.62 ng/ml. Transmission electron microscopy of the ultrathin sections of treated or untreated erythrocytes indicated that the crenations were formed by protrusions of the plasma membrane, occurring presumably because of the local increase of membrane fluidity after polymyxin treatment. Changes in the shape of the erythrocytes to the ultimate rounded forms were also indicated by the transmission electron microscopy.

A statistical analysis of current quantum responses (“bumps”) to the absorption of single photons in the ventral photoreceptor of Limulus is presented. The analysis is performed in terms of bump parameters among which two groups are distinguished: “time-like parameters” with distributions independent of individual cells and “current parameters” the distribution of which differ for different cells. The analysis yields a set of “primary” parameters with mutually uncorrelated fluctuations. The results are interpreted by a new model formulation on the basis of a transmitter hypothesis for the transduction. The model separates latency from amplification and postulates the existence of different sources of fluctuations among which one includes individual cell properties.

Three periodicities in the circadian range are observed when measuring circadian parameters of unicellular organisms in long running experiments (more than 15 days). This is demonstrated for different organisms (Chlamydomonas, Euglena, Chlorella) and different parameters (auto­ kinesis, extracellular pH, CO 2 - and O 2 -partial pressure). Having excluded analytical and experimental artefacts (i.e. filter leakage and subpopulation effects), the multiple periodicities have to be interpreted in a physiological model. The three periodicities always exhibit two common features: The locations of the side peaks are symmetrical to the middle peak and their energy contribution is always the same. We therefore favour the model of multiplicative coupling between the circadian oscillator and a low frequency oscillator modulating the amplitude of the circadian rhythm. Since the low frequency rhythm is not correlated to any exogenously running periodicity of the experimental surroundings, it is considered as generated by an endogenous oscillator. This shows the existence of different biological long time oscillators in one single cell and contradicts the so-called master-clock hypothesis stating that one cell has only one clock related oscillator.

Motility of Euglena gracilis shows free running circadian rhythms. The circadian system is sensitive to light and temperature signals, but it is always masked by direct responses of motility to light (photokinesis) and temperature (thermokinesis). By means of a compartimental model which defines the interrelations between the pathways of thermokinesis, photokinesis and the circadian system a unifying view of effects of temperature and light input signals is outlined. According to the model, and using double sine input signals the dynamics of thermokinesis is described by a differential amplifier with constant gain. Although thermokinesis heavily masks circadian responses to temperature signals, the limited range of circadian entrainment is indirectly demonstrated by a limited reappearance of free running circadian oscillations after stopping the temperature program. Free running circadian oscillations do reappear only after pretreatment with temperature periods near the circadian eigenperiod. A white mutant lacking photosynthesis is used to investigate the role of photosynthesis in the signal processing. Although light synchronizes the circadian rhythms of the white mutant if applied as single input, it does not affect the motility if applied together with temperature inputs near the circadian eigenperiod. These results indicate frequency dependent mutual interactions between the model compartments.

The fine structure of haptocysts, characteristic organelles of most suctorians, is described during the resting state and the discharge after contact with the prey. In Ephelota, there is no fusion of the plasma membranes of predator and prey, as has been suggested for the haptocysts of other suctorians. But the membrane covering the organelle becomes continuous with the plasma membrane of the suctorian tentacle, and inner structures of the haptocyst attach to the perforated pellicle of the prey.

In contrast with the outmost uniform aspect of Trichoplax there is a clear cut differentiation between marginal and central cell territories. Dissection and transplantation as well as vital staining show that the marginal material is restricted to a seam approximately 20 μm in width, surrounding the much larger mass of central cells. Isolated marginal or central cell material can not regenerate a complete individual and is therefore doomed to die. Regeneration requires the presence of both components because the cells propagate true to type only, meeting the Ruthmann model (A Ruthmann, Cytobiol. 15, 58 (1977)). There is no redifferentiation between marginal and central cells. Marginal pieces transplanted into the central area produce an inverted inner edge, surrounding an open hole. Dissection or transplantation borne frag­ments containing sufficient cells of both marginal and central provenience reassort their cells to the normal relative position. Unbalanced compositions of fragments may be regulated by detachment of surplus material or (in the case of marginal surplus) also by fission, even in very small individuals. This way fission is involved in the regulation of the quantitative relation between these two kinds of cells. A monstrous modification of the shape and physiologic properties demonstrates a disturbance of these regulations. The marginal versus central differentiation manifests a radial polarity, till now unknown as to the relation to the histologically discerned cell types.

The hawkweed Hieracium intybaceum is covered with glandular trichomes that produce a sticky resinous exudate. This material, assumedly of terpenoid nature, contains trace amounts of flavonoid aglycones. They could be identified to be known flavones and flavanones. This is the first time that an alpine Asteraceae was found to exude flavonoid aglycones.

Semithin and thin sections of glutaraldehyde-fixed, epoxy resin-embedded animal tissues were treated with solutions of oxidized p-phenylenediamine (PPD). This method is suitable to reveal the general morphology of tissues in light microscopy, showing a high staining degree in some polyanion containing components. Posttreatments of thin sections with gold chloride solutions give consider­ able electron opacity in PPD positive structures.

The toxic amino acid β-cyano-L-alanine is shown to be present in 21 species of the Zygaenidae, which are known to produce and store the cyanoglucosides linamarin and lotaustralin. β-Cyano-alanine is also reported for the first time from other cyanogenic Lepidoptera, i.e. the Acraeinae and the Heliconiinae of the nymphalid family and from other species of the Nymphalidae, which have not been known to be cyanogenic before. Cyanogenesis, including the presence of β-cyano-alanine and the cyanoglucosides linamarin and lotaustralin is demonstrated for the first time for the Heterogynidae.

Species-specific sex-attractants are reported for western Palearctic Zygaena moths of the carniolica, viciae, filipendulae, and purpuralis species groups. The attractants were established by combined electrophysiological and field behavioural studies and consist of ternary combinations of (Z)-7-dodecenyl/(Z)-9-tetradecenyl acetates with a particular further (Z)-alkenyl acetate. Data on other Zygaenini taxa are included.

Binary combinations of (Z)-7-dodecenyl acetate and (Z)-9-tetradecenyl acetate in a 1/10 ratio were highly attractive to Rh. baja males in field trapping tests conducted in southern Germany. Trap captures increased further on adding 0.1 or 0.3% of (Z)-5-dodecenyl acetate to this lure; higher amounts of this chemical were strongly inhibitory, as were additions of > 3% of (Z)-11-hexa-decenyl acetate. The trapping data are considered in terms of electrophysiological responses of five types of antennal receptor cells.

The antennal Sensilla trichodea of male Eulia ministrana (Tortricidae: Cnephasiini) contain specialist receptor cells for the sex-attractant components (Z)-9-and (Z)-11-tetra-decenyl acetate and the attraction-inhibitors (E)-11 -tetra-decenyl acetate, (Z)-7-dodecenyl acetate, and (Z)-11 -hexa-decenyl acetate. The significance of the latter two compounds as potential sex pheromone components in Tortri­cidae is considered.

Out of several media tested, KCl medium containing BSA was found to be most suitable for isolation of functionally efficient mitochondria from earthworms. Respiration and oxidative phosphorylation in isolated mitochondria are demonstrated in the presence of cytochrome c and BSA in reaction medium. Introduction - The details of the respiratory metabolism in earthworm are yet to be established. There are only a few contradictory reports on the nature of its respiratory enzyme components [1]. While, cytochrome oxidase, succinoxidase, succinate dehydrogenase were identified in earthworm tissue homogenates, the presence of Kreb′s cycle enzymes could not be demonstrated. Very little information is available on earthworm mitochondria except one report where it is isolated in 0.44 ᴍ mannitol [2]. Mitochondria from invertebrates like insects and crustaceans are isolated and characterized [3, 4]. But such methods cannot be extended to earthworms as it is known that conditions for isolation and assay of mitochondria are highly system specific [5].

The patterns of the anterior lateral-line afferents of Lampetra fluviatilis as revealed by transganglionic trans­port of horseradish peroxidase are described. The afferents form two roots in entering the rhombencephalon. Fibers of the dorsalmost root can be traced to a short dorsal fascicle which runs along the dorsal nucleus. The ventral roots form two fascicles adjacent to the nucleus intermedius. Comparison with urodeles indicates that lampreys, like urodeles and other anamniotic vertebrates are electroreceptive.

Metabolism, [4- 13 C]Phenacetin, Continuous Flow 13 C NMR Continuous flow 13 C NMR spectroscopy has been used for the first time to monitor the metabolism of a 13 C labeled drug in an isolated liver. Continuous and almost immediate information on the metabolite formation could be obtained using 13 C labeled phenacetin without alteration of the biological system. The data are consistent with those observed by conventional techniques (HPLC, aliquot 13 C NMR measurements). From the biological point of view the sensitivity of continuous flow 13 C NMR spectroscopy is still low (10 -3 ᴍ). The results presented demonstrate however that non-invasive and non-radio-active real time monitoring of drug metabolism in intact organs is possible.