Band 47, Heft 7-8

The structures of two pyoverdins (Pp 1 and Pp 2) and one dihydropyoverdin (dihydro-Pp 2) from a strain of Pseudomonas putida have been elucidated by spectroscopic methods and degradation studies. The pyoverdins Pp 1 and Pp2 consist of a chromophore which was identified as (1 S)-5-amino-2,3-dihydro-8,9-dihydroxy-1 H-pyrimido[1,2-a]quinoline-1-carboxylic acid substituted at the amino group with a 3-carboxypropanoyl or a succinamoyl residue and at the carboxy group with the N-terminus of L-Ser-L-Thr-D-Ser-L-Orn-L-threo-(OH)Asp-[D-Gln + L-Dab]*-L-Ser-D-allo-Thr-L-c(OH)Orn. Dihydro-Pp 2 differs from Pp 2 only in the chromophore, which is saturated at carbons 5 and 6. All compounds contain a tetrahydropyrimidine moiety ([D-Gln + L-Dab]*) resulting from the condensation of 2,4-diaminobutyric acid and glutamine. The structures of two pyoverdins (Pp 1 and Pp 2) and one dihydropyoverdin (dihydro-Pp 2) from a strain of Pseudomonas putida have been elucidated by spectroscopic methods and degradation studies. The pyoverdins Pp 1 and Pp2 consist of a chromophore which was identified as (1 S)-5-amino-2,3-dihydro-8,9-dihydroxy-1 H-pyrimido[l,2-a]quinoline-1-carboxylic acid substituted at the amino group with a 3-carboxypropanoyl or a succinamoyl residue and at the carboxy group with the N-terminus of L-Ser-L-Thr-D-Ser-L-Orn-L-threo-(OH)Asp- [D-Gln + L-Dab]*-L-Ser-D-allo-Thr-L-c(OH)Orn. Dihydro-Pp 2 differs from Pp 2 only in the chromophore, which is saturated at carbons 5 and 6. All compounds contain a tetrahydropyrimidine moiety ([D-Gln + L-Dab]*) resulting from the condensation of 2,4-diaminobutyric acid and glutamine.

Column chromatography of the acetone extract of the lichen Stereocaulon azoreum afforded several substances identified by chemical and spectral means; α-amyrin, lupeol, taraxerol, ursolic acid, ergosterol peroxide, brassicasterol, cerevisterol, stictic acid and an acetone-condensation derivative, lobaric acid. Furthermore atranorin, methyl β-orsellinate, atranol and, for the first time from the genus Stereocaulon, cryptostictic acid and the dibenzofuran, strepsilin were isolated.

Several triterpenes were isolated from the lipophilic frond exudates of three Notholaena species. In the terpenoid portion of N. Candida we identified two novel cycloartane type triterpenes. N. rigida yielded protolyofoligenic acid, a cycloartane type product that has been reported once before from a higher plant. From N. schaffneri, a novel dammarane type compound was isolated. These products were identified by spectroscopic methods.

Analysis of bud exudate of Populus tristis by GC/MS revealed 61 phenolic components representing 55 phenolic compounds. Caffeic acid esters and chalcones made up the bulk of the exudate.

Epicuticular leaf waxes of Ginkgo biloba, Magnolia grandiflora and Liriodendron tulipifera contain homologous series of hydrocarbons, wax esters, benzyl acyl esters, aldehydes, primary alcohols, and fatty acids. None of these lipid classes is found to contain any main component dominating. In addition to these usual wax lipids, in G. biloba leaf wax a secondary alcohol namely nonacosan-10-ol (15.0%), γ-tocopherol (1.7%) and acetates (0.3%) is also present. The wax of L. tulipifera, however, contains hentriacontan-16-one (23%) and several triterpenols (10%), additionally. On G. biloba leaves a dense arrangement of tubular wax crystalloids are found on the lower as well as on the upper leaf surface. The openings of the tubules can be seen very well in the SEM figures at a magnification of 20000. The small tubules are a clear indication for the presence of nonacosan-10-ol as also reported previously for conyferyl waxes. Leaves of M . grandiflora have an abaxial epidermis with a continuous wax layer without any crystalloids or sculptures. The adaxial epidermis also shows a continuous wax layer but with little irregular granular sculptures. L. tulipifera leaves show an abaxial epidermis with a continuous wax layer superimposed by a dense arrangement of crystalloids in shape of angular rodlets which are composed of several piled up layers. The adaxial leaf surface is also superimposed with wax crystalloids, the rodlets of which, however, are not sculptured in such definite way. They usually appear melted up and also form granular sculptures. The wax crystalloids in shape of angular rodlets on the abaxial surface are formed by hentriacontan-16-one. The abaxial and adaxial leaf surfaces show most different micromorphological wax ultrastructures, as shown for all trees studied.

From Bartramia pomiformis bartramic-acid (luteolin-8,2′′-protocatechuic acid), bartramiatriluteolin, the first triflavone and six known biflavonoids (bartramiaflavone, anhydrobartramiaflavone, 2,3-dihydrophilonotisflavone, philonotisflavone, dicranolomin and 5′,3′′′-dihydroxyamentoflavone) could be isolated. The structures of the new compounds were elucidated spectroscopically.

Seven phenolic compounds were isolated from the aerial parts of U. picroides. They were identified by UV spectra, TLC and HPLC-DAD as luteolin, luteolin-7-glucoside, quercetin, quercetin-3-galactoside, kaempferol-3-galactoside, chlorogenic and isochlorogenic acids. Other phenolics were characterized by HPLC-DAD analysis: gallic, protocatechuic, caffeic, ferulic and isoferulic acids, quercetin-3-glucoside and luteolin-4′-glucoside.

A histochemical analysis was carried out on the distribution of flavonoid derivatives in Sophora flavescens var. angustifolia plant. Prenylated flavonoids such as kushenol I, kurarinone, sophoraflavanone G and des-O-methylanhydroicaritin were mainly localized in the periderm (cork layers, and cork layer-like tissues scattered in parenchymatous tissues of the root system). Pterocarpan derivatives, maackiain, maackiain-3-O-glucoside (trifolirhizin) and its 6′-O-malonyl ester, were distributed in the cortex, cambium and pith as an ester form. In the aerial parts of the plant, neither pterocarpan nor prenylated flavonoid derivatives, but only flavone monoglucosides such as luteolin-7-O-glucoside and apigenin-7-O-glucoside were detected.

In an earlier publication we reported on the role of arginine for the development of nitrate reductase activity in cells of the filamentous cyanobacterium Oscillatoria chalybea (Bednarz and Schmid, Z. Naturforsch. 46c, 591 (1991)). In the present paper we present further evidence that arginine is the natural inducer for nitrate reductase activity. Thus, we show that the induction is regulated by transcription, probably related to the apoprotein or the molybdenumcofactor. We also examined the influence of arginine on nitrate reductase activity in the filamentous cyanobacterium Anabaena PCC7120. In contrast to Oscillatoria chalybea this cyanobacterium forms heterocysts and shows nitrogen fixation activity. Like in Oscillatoria chalybea nitrate reductase activity in Anabaena PCC7120 is stimulated in the presence of arginine as the sole nitrogen source. However, this stimulation is limited to an early growth stage. Subsequently, nitrogen fixation activity appears and nitrate reductase activity decreases.

Cells from photoautotrophic cultures of Chenopodium rubrum were fractionated for the isolation of purified chloroplasts and mitochondria. The subcellular localization of the enzymatic activities involved in the metabolism of malic acid was investigated. Highly purified chloroplasts were obtained from the protoplasts, whereas peroxisomes were still present in the mitochondrial fraction. NAD - and NADP-dependent malate dehydrogenase and malic enzyme activities were found in the mitochondrial and chloroplast fractions, respectively. Exogenously supplied [ 14 C]labelled malate was metabolized by the photoautotrophic cell suspension, obviously to the greater part in mitochondria.

Flavonol synthase (FLS) was demonstrated in crude extracts from flower buds of Dianthus caryophyllus (carnation). The enzyme catalyzed the conversion of dihydrokaempferol and ihydroquercetin to kaempferol and quercetin, respectively. The reaction required 2-oxoglutarate, ferrous ion and ascorbate as co-factors and had a pH optimum at about 7.4. The demonstration of FLS activity allowed comparative studies on flavonol and anthocyanin biosynthesis during bud and flower development. Besides FLS the flavonoid enzymes chalcone synthase (CHS), flavanone 3-hydroxylase (FH T) and dihydroflavonol 4-reductase (DFR) were measured. DFR is specifically involved in anthocyanin synthesis, while CHS and FHT provide dihydroflavonol, the common substrate for both FLS and DFR . Maximum expression of CHS, FHT and FLS activity was already observed in small buds, whereas DFR activity started to increase much later and reached its highest level in opened flowers. A substantial correlation was observed between the time courses of FLS and DFR activity and the accumulation of flavonols and anthocyanins, respectively. The competition of FLS and DFR for dihydroflavonols was found to be largely circumvented by different substrate specificities and by the sequential expression of the two enzymes. Both flavonols and anthocyanins are obviously not, or only to some extent, subject to degradation.

Cultures of the basidiomycete Mycena avenacea TA 8480 were shown to metabolize entachlorophenol (PCP), 2,3,5,6-tetrachloro-p-hydroquinone (TeCHQ), and 2,3,5,6-tetrachloro-p- benzoquinone (TeCBQ). The first metabolite of the PCP degradation pathway was identified as TeCBQ which in a second reaction is reduced to the hydroquinone TeCHQ. Subsequently dechlorination of TeCHQ yielded 3,5,6-trichloro-2-hydroxy-p-benzoquinone (TCOHBQ). The specific degradation rate for PCP was 1.4 mg x g dry m ycelia -1 x day -1 . The initial dechlorination rate of TeCHQ was 5.9 mg x g dry mycelia -1 x hour -1 . None of the compounds supported growth of the fungus.

Keywords Ascorbic acid radical, EPR, Herbicides, Damage to Forest, Antioxidative System Treatment of young spruces with the herbicides aminotriazole and paraquat leads to oxidative stress resulting in a drastically increased production of ascorbic acid radical (m onodehydroascorbic acid, MDAA) which could be demonstrated by in vivo EPR-analysis. While paraquat action is accompanied by a rapid decrease in reduced ascorbic acid content, no change in ascorbic acid content was observed by aminotriazole treatment. Thus MDAA-measurements can undoubtedly verify an existing oxidative stress of different origin in spruce needles. A decision between both radical generating mechanisms based on the radical kinetics is possible.

S-triazines are photosynthetic inhibitors. They have been substituted with ω-aminoundecanoic acid. The coupling products have been transformed into triazine aldehydes. These compounds displace radioactive terbutryn and have inhibitory effects on photosynthesis in plants and bacteria. Triazine aldehydes were shown to be effective substrates for bacterial luciferase. A competitive assay between photosystem-II-herbicides and aldehyde-labeled triazines is discussed.

Several herbicides block the photosystem II electron transfer because they compete with Q B , the second stable electron acceptor of photosystem II for binding to the D 1 subunit. We have previously isolated a mutant of Synechocystis 6714 in which Asn is replaced by Thr at position 266 of D 1 (G. Ajlani, I. Meyer, C. Vernotte, and Astier, FEBS Lett. 246, 207-210 (1989)) and presenting resistance to ioxynil but not to DCMU . In this report we present selection, from this mutant, of a double mutant with an additional substitution at position 264 (Ser by Ala). The sensitivity of this mutant toward several herbicides is given and compared to those of the mutants having only one substitution at 266 and one substitution at 264. It was also compared to a mutant of Chlamydomonas having the same substitutions. This allows us to discuss the interaction of various herbicides with the D 1 protein and to compare the herbicide binding niches of Chlamydomonas and Synechocystis.

During photoinhibitory light treatment of the green alga, Chlamydobotrys stellata and Pisum sativum leaves the amplitude of the B thermoluminescence band (S 2 O B - charge recombination) decreased faster than that of the Q band (S 2 Q A - charge recombination). Consistently, in the alga cells the electron transport rate from water to oxidized diaminodurene (electron acceptor after Q B ) was also diminished faster than that measured from water to phenyl- p-benzoquinone (electron acceptor after Q A ). These observations suggest that in intact photosynthetizing organisms at high light intensities on the acceptor side of photosystem II photoinhibition is initiated at the binding site of the secondary quinone acceptor, Q B .

Using in situ RNA hybridization and immunohistochemical techniques, we investigated the tissue-specific and light-dependent expression of four genes involved in phenylpropanoid metabolism in various parts of parsley (Petroselinum crispum) seedlings. The genes encode phenylalanine ammonia-lyase (PAL) and 4-coum arate: CoA ligase (4CL), two enzymes of general phenylpropanoid metabolism; chalcone synthase (CHS), the key enzyme of flavonoid biosynthesis; and bergaptol O-methyltransferase (BMT), a late enzyme of the furanocoumarin pathway. In young leaves, PAL and 4 CL genes were expressed predominantly and in a coordinated fashion in epidermal cells, young xylem cells of vascular bundles, and epithelial cells of oil ducts. The gene(s) encoding BMT was active exclusively in the epithelial cells of oil ducts, whereas CHS gene expression was largely confined to the epidermis. In shoot and root apices, PAL, 4CL and CHS m RNAs were detectable at low levels without distinct patterns. The corresponding proteins, however, accumulated preferentially in the protoderm and pith meristem of the shoot apex and in root-cap cells. A gene encoding pathogenesis-related protein 1 (PR 1) was analyzed in roots for comparison and was found to be expressed predominantly in the cortical cells of root tips. The expression of all genes investigated was, to a greater or lesser extent, dependent on light conditions and tissue age, with highest levels occurring in newly differentiated, light-exposed tissue. In the leaf epidermis of seedlings illuminated for 20 h, PAL and CHS mRNAs and proteins, as well as 4CL protein, were particularly abundant.

Pentachlorophenol (PCP) is a widely used pesticide, particularly for the preservation of wood. Given its high toxicity and resistance to degradation it has become a dangerous environmental pollulant. Due to its high lipophilicity, PCP is able to partition into the lipid bilayer of cell membranes disrupting several vital functions. The present research was concerned with the effects that the chronic administration of PCP could produce in vivo to the sciatic nerve of rats. X-ray diffraction patterns obtained from freshly dissected and dried sciatic nerves of PCP treated rats did not show significant differences in their reflections with respect to those present in the patterns from untreated animals. However, morphological studies performed by optical and electron microscopy showed degenerative changes in about 10% of the A and B type of nerve fibers.

The effects of resorcinolic lipids (5-n-alk(en)ylresorcinols) isolated from cereal grains on the phospholipase A 2 catalyzed hydrolysis of phospholipid vesicles were examined. Studied compounds inhibited the apparent enzyme activity at the molar fraction in the membrane as low as 0.025, which is equivalent to the concentration of 3 μᴍ . This effect was visualized by dramatic increase (over ten-fold) of the latency period of the reaction progress. This makes resorcinolic lipids one of the most potent inhibitors of phospholipase A 2 among already studied compounds. Highest inhibitory activities were shown for dienoic and monoenoic homologs of 17 carbon atoms in the aliphatic chain. Both saturation of the chain and the increase of its length reduced inhibitory properties of resorcinolic lipids. The data suggest that the compounds studied in this paper like other known amphiphilic inhibitors of phospholipase A 2 owe most of their effects to the ability to modify the quality of the substrate interface. These are the alteration of the enzyme binding, velocity of the formation and redistribution of the products. However part of the effect seems to be attributed to direct interaction and modification of enzyme properties by alk(en)ylresorcinols.

In field trapping tests male Orthosia gothica (Lepidoptera: Noctuidae) were captured in traps baited with blends of (Z)-9-tetradecenyl acetate, (Z)-9-tetradecenol, (Z)-5-tetradecenyl acetate and (Z)-l 1-tetradecenyl acetate. The highest catches were recorded in traps baited with a 20:20:1:1 blend. The above quaternary blend is a potent sex attractant of this species, and it can be recommended in dosages of 10-100 μg per dispenser for monitoring purposes.

Analysis of the marking fluid of two tigers (one Bengal and one Sumatran) by GC using an amine-specific column and a nitrogen-specific detector has shown the presence of the following amines: ammonia, methylamine, dimethylamine, trimethylamine, triethylamine, propylamine, and butane-1,4-diamine (putrescine). In contrast to previously published reports, we were unable to detect 2-phenylethylamine. The anal sac secretion was found to have a similar amine content.

The effect of low intensity microwaves on the conformational state of the genome of X-irradiated E. coli cells was studied by the method of viscosity anomalous time dependencies. It has been established that within the ranges of 51.62-51.84 GHz and 41.25-41.50 GHz the frequency dependence of the observed effect has a resonance nature with a resonance halfwidth of the order of 100 MHz. The power dependence of the microwave effect within the range of 0.1-200 (μW/cm 2 has shown that a power density of 1 μW/cm 2 is sufficient to suppress radiation-induced repair of the genome conformational state. The effect of microwave suppression of repair is well reproduced and does not depend on the sequence of cell exposure to X-rays and microwave radiation in the millimeter band. The results obtained indicate the role of the cell genome in the resonant interaction of cells with low intensity millimeter waves.

To explain the high specificity, high reaction rate, and high thermodynamic efficiency in enzymatic processes, cooperation of the enzyme with a molecular transfer unit is assumed. A “kinetic equilibrium of forces” is suggested, which enables high reaction rates to occur under equilibrium conditions and a thorough examination of the substrate to be made without consumption of free energy. In case of ATPases, ion-binding proteins are the most probable transfer units. By analyzing the elementary effect in muscle contraction it is shown that the new theorem may be of substantial value in elucidating biochemical processes.

Propolis and propolis balsam from two different Spanish Regions were analyzed by GC-MS. The majority of the compounds were typical of poplar bud exudate of P. nigra. No significant differences were found between the balsam and the entire propolis analyzed, apart from the components of beeswax added to the plant exudate by the bees.

Myricetin-3,7,3′,4′-tetramethyl ether has been isolated as an aglycone from the acetone wash of Tillandsia usneoides. It is thus shown to be deposited externally, on leaf and stem surfaces. We suggest that the previously reported lipophilic flavonoids from this and other species are also exudate constituents.