Volume 46, Issue 3-4

The basidiomycete Lepista irina (F r.) Bigelow (strains CBS 366.47 and CBS 458.79) was cultivated on a biomalt (2% )-liquid medium for 28 days. The steam distillate was analyzed by GLC and GLC-S. The structure elucidation of the predominant constituent resulted from spectroscopic methods, particularly from its 13 CNMR spectrum, nuclear Overhauser enhancement 1 HNMR data, and from 2d 1 H homonuclear shift correlation. The bisabolane derivative was nam ed lepistirone and is described for the first time as a natural product. Two minor related com pounds could be characterized by their MS data.

Callus cultures of Sophora favescens var. angustifolia established on Murashige-Skoog medium containing 1 μм 2,4-dichlorophenoxyacetic acid and 1 μм kinetin produced the prenylated flavanones (2S)-5,7,2′,4′-tetrahydroxy-8-lavandulylflavanone (sophoraflavanone G) and (25)-7,2′,4′-trihydroxy-8-lavandulylflavanone (lehmannin). In addition, maackiain and its 3-O-β-glucoside (trifolirhizin) were also produced in the callus. Up on transfer to White’s medium or M9 medium, the content of prenylated flavanones, in particular lehmannin, was increased, whereas that of pterocarpans was decreased. Time-course experiments indicated that the production of pterocarpans was closely related with cell growth. On the other hand, an inverse relationship existed between cell growth and the production of prenylated flavanones.

From the leaf exudate of Aloe lateritia ENGLER the C-glucosyl com pounds homonataloin, aloeresin A and aloesin (synon. aloeresin B) were isolated together with the anthraquinone nataloeem odin-8-methylether and spectroscopically identified. Hom onataloin, widely distributed in Aloe species, was separated into homonataloin A and B by combined TLC and DCCC. In their 1 D and 2D 1 H NMR spectra only the shifts of the 2′-hydroxyl protons of both glucosyl residues differ significantly, indicative of 10 S (A) resp. 10 S (B) configurations. In both com pounds the anthrone is in β-position of the D-glucopyranosyl, as determined by the large coupling constants of the anomeric protons. The 13 C NMR signals are unambiguously assigned by the use of DEPT, APT and gated-decoupling methods. Only the chemical shifts of C -11 and C -14 show significant differences between both diastereomers due to the adjacent 2′-sugar hydroxyls. The two homonataloins differ mostly in optical rotation and circulardichroism due to different configurations at C - 10 of the anthrone part. The absolute configurations of the diastereomers are determined by correlation of their CD spectra with the CD spectra of the structural analogues 7-hydroxyaloins A and B, which shows that hom onataloin A is the 10 S, 1′S-compound and that homonataloin B has 10 R, 1′S-configuration.

An axenic culture of Symphyogyna brongniartii has been initiated from spores on modified Benecke medium. Growth was best on Gam borg B5 medium with 0.3% sucrose, with a doubling time of 36 days. The culture produced the same terpenoids as plants growing in their natural habitat, perrottetianal A being the major constituent. Also found were bicyclogermacrene, β-barbatene, δ-selinene, β-cubebene, spathulenol, and (-)-geosm in . The latter compound was hitherto only known from microorganisms. Synthesis rate of terpenoids during the passage period and quantitative differences in terpenoid production on different mineral media were investigated.

Sliced turnip root (Brassica campestris L. ssp rapa) was irradiated for a total of 20 min with a 15 W germicidal lamp and the tissue incubated at 25 °C. The effects of such treatment on indole phytoalexins (methoxybrassinin (I); brassinin (II); cyclobrassinin (III); spirobrassinin (IV) and glucosinolates were determined using high performance liquid chromatography procedures. Accumulation of phytoalexins I - III was evident within 8 h of irradiation, whilst formation of spirobrassinin was evident only after 24 h. Maximal levels of III and IV (> 100 μg g -1 freeze dried tissue) were greater than those of I and II (27 and 17 μg g -1 , respectively). The individual glucosinolate levels were affected in a complex manner; whilst most glucoinolates decreased on storage, the levels of indole glucosinolates, glucobrassicin (XI) and 1-methoxyglucobrassicin (XIII), increased until 5 to 6 days after irradiation and thereafter declined. Whilst structural features of I - IV , XI and XIII suggest close biosynthetic relationships between these classes of biologically-active indoles, further studies are needed to establish this point unambiguously.

Pseudomonas fluorescens Y 4 grown in an iron deficient medium produces increased amounts of 2,9-di- and 2,3,9-trihydroxyphenazine-1-carboxylic acid when Be 2+ is added to the culture. The significance of the formation of these compounds is discussed.

Three bacterial strains of marine origin were isolated during a screening for biosurfactants among n-alkane degrading microorganisms. One strain - identified as Alcaligenes sp. MM 1 - produced a novel glucose lipid. In the case of Arthrobacter sp. EK 1 the well-known trehalose tetraester was found as major component. From another pure culture classified as Arthrobacter sp. SI 1, extracellular emulsifying agents with properties indicating high molecular weight substances were detected. Furthermore trehalose corynomycolates were found at up to 2 g/1. The isolated biosurfactants showed good interfacial and emulsifying properties.

Within a screening for biosurfactants we could isolate various n-alkanes utilizing marine bacteria which were capable of synthesizing glycolipids. One strain was identified as Arthrobacter sp. EK 1 which produced trehalose lipids. After purification by column and thick layer chromatography the main fraction, an anionic 2,3,4,2′-trehalose tetraester, was obtained. The chain lengths of fatty acids ranged from 8 up to 14, furthermore succinate could be detected. Since the place of substitution of succinate has so far not been cited in literature, a definitive structural elucidation was carried out chemically by hydroboration and by 1 H, 2 D 1 H, 13 C and 13 C - 1 H correlation NMR measurements. All investigations confirmed the exact position of succinate at C2 atom of trehalose. After improvement of growth conditions the production of the trehalose tetraester increased up to 4.8 g/1 during a fermentation in a 20 1 bioreactor under nitrogen limitation.

Eight synthetic and nine biogenetic surfactants were tested on their toxicity. Because of their possible application as oil dispersants against oil slicks on sea. the test organisms used were marine microorganisms (mixed and pure cultures of bacteria, microalgae, and protozoa). Bacterial growth was hardly effected or stimulated, whilst that of algae and flagellates was reduced. All substances tested were biodegradaded in sea water. The bioluminescence of Photobacter phosphoreum (Microtox test) was the most sensitive test system used. A ranking shows that most biogenetic surfactants were less toxic than synthetic surfactants. No toxicity could be detected with the glucose-lipid GL. produced by the marine bacterium Alcaligenes sp. MM 1.

Physiological experiments indicate that Anabaena variabilis can express either a V- or a Fe-nitrogenase in addition to the conventional, M o-containing enzyme complex. The occurrence of the Fe-nitrogenase in A. variabilis can also be concluded from DNA - DNA hybridization experiments using cloned anfW or nifW probes coding for the smaller subunit (= nitrogenase reductase) of the Fe-nitrogenase from Azotobacter vinelandii or for the same subunit of the Mo-nitrogenase from Klebsiella pneumoniae. The cyanobacterium A. variabilis is the first phototroph found to contain all three nitrogenases wich has been described only for A. vinelandii as yet. Vanadium cannot substitute for Mo in nitrate reduction of A. variabilis.

Specific enzyme activities of the non-oxidative pentose phosphate pathway in Aspergillus oryzae mycelia grown on different carbon sources were determined. Mycelia grown on glucose, mannitol and ribose show the highest specific activities, ribose 5-phosphate isomerase being specially very enhanced. Moreover, transketolase, transaldolase, ribose 5-phosphate isomerase and ribulose 5-phosphate 3-epimerase were determined in different developmental stages of mycelia grown on glucose, mannitol and ribose. The non-oxidative pentose phosphate pathway is more active during conidiogenesis, except for ribulose 5-phosphate 3-epimerase, suggesting a fundamental role of this pathway during that stage to supply pentoses for nucleic acids biosynthesis. A general decrease of the enzyme activities was found in sporulated mycelia. Arabinose 5-phosphate was tested as metabolite of the pentose pathway. This pentose phosphate was not converted into hexose phosphates or triose phosphates and inhibits significantly the ribose 5-phosphate utilization, being therefore unappropriate to support the Aspergillus oryzae growth.

Protein-free and protein-containing chlorosomes from Chloroflexus aurantiacus, strain Ok-70 fl, were studied by resonance Raman (RR) spectroscopy. Both preparations gave the same spectra of the bacteriochlorophyll c (BChl c) chromophores in the range of 1200-1750 cm -1 . This strongly corroborates previous evidence [Griebenow et al., Z. Naturforsch. 45c, 823-828 (1990), and references therein] that the three-dimensional structure of the antenna complexes is not determined by direct interaction with protein but rather is due to BChl c selforganization. The analysis of the coordination-sensitive marker bands of the chlorin macrocycle reveals a mixed six- and fivefold ligation of the Mg ion. Based on two C = O stretching vibrations originating from a free and a Mg-bound C-9 keto group, it is concluded that only in the six-coordinated state the keto group serves as an axial ligand to the Mg ion of a neighbouring chlorin. The second permanently bound axial ligand is attributed to the C-2a hydroxyl group.

Light-induced absorbance changes in the near-infrared (NIR ) spectral region (700-950 nm) are analyzed in isolated spinach chloroplasts using pulsed light-emitting diodes as modulated measuring light sources. With chloroplasts under coupled conditions the light-induced signal is dominated by a relatively slow scattering change displaying a flat difference spectrum. More specific changes can be distinguished by faster induction and relaxation kinetics and in the presence of valinomycin/nigericin which prevents scattering changes. Besides the dominant P700 change, with a narrow bleaching band around 703 nm and a broad absorbance increase between 750 and 850 nm (peak at 815 nm) the following absorbance changes were identified in the NIR: 1) An absorbance increase caused by plastocyanin (PC) oxidation, with a relatively flat spectrum. 2) An absorbance decrease gaining amplitude towards longer wavelengths, which reflects reduction of a low potential acceptor of PSI different from ferredoxin. 3) A field indicating absorbance decrease peaking around 730 nm, the properties of which correspond to those of P515. 4) An uncoupler insensitive absorbance decrease stimulated by dark adaptation and anaerobic conditions, the difference spectrum of which resembles that of ferredoxin reduction. The relative contributions of P700 and PC to the overall oxidized-reduced difference spectrum are determined by redox titration. At 706 nm, 815 nm and 950 nm the P 700/PC is - 119/ 19, 67/37 and 31/69, respectively. From these ratios and the molar extinction coefficients a molar P 700/PC stoichiometry of 1 /3 is determined.

A new method is presented to follow the turn over of the D-1 reaction center polypeptide of photosystem II in intact spruce needles or spinach leaves by incorporation of [ 14 C]leucine. Like in other organisms the synthesis and degradation of this protein is strongly light-dependent and very fast. The synthesis rate under moderate light intensities in healthy spruce needles is about 9 nmol [ 14 C]leucine mg -1 Chl h -1 . The half-life time of the D-1 protein under these conditions is less than 30 min. In order to demonstrate the usefulness of this method for the stress physiology of photosynthesis, first experiments are shown comparing the turnover of the D-1 protein in spruce needles with different age and different degree of dam age.

A boronated analogue of the antiestrogen U 23,469-M (D. Lednicer, D. W. Emmert, S. C. Lyster, and G. W. Duncan, J. Med. Chem . 12, 881 (1969)) was prepared, for possible use in neutron capture therapy of estrogen receptor-positive tumors. In this analogue, the terminal OH group was replaced by a B-decachloro-o-carboranyl residue. This com pound showed a large, non-specific uptake in ZR75-1 breast cancer-derived cells. It could partially inhibit the uptake of estradiol in these cells. Accumulation in the cells at physiologically obtainable concentrations was, however, too low to envisage a therapeutic effect following thermal neutron irradiation.

(Z ,Z )-3,9-cis-6,7-Epoxyheptadecadiene, and (Z,Z)-6,9-cis -3,4-epoxyheptadecadiene were identified by gas chromatography/flame ionization detection-electroantennographic detection (GC/FID-EAD) and coupled gas chromatography-mass spectrometry (GC-MS) from abdominal tip extracts of Tephrina arenacearia (Lepidoptera: Geometridae), an alfalfa pest. In gas chromatography/flame ionization-single sensillum detection (GC/FID-SC) analyses, specific olfactory receptor cells were found for (Z,Z)-6,9-cis-3,4-epoxyheptadecadiene, and tricosane. Synthetic samples of racemic (Z,Z )-6,9-cis-3,4-epoxyheptadecadiene attracted large numbers of male T. arenacearia into traps. Best catches were observed at 100-1000 μg dosages. The addition of (Z,Z,Z)-3,6,9-heptadecatriene did not influence catches. (Z,Z,Z)-3,6,9-Heptadecatriene and (Z,Z)-6,9-cis-3,4-epoxyheptadecadiene were also identified in the pheromone gland extract of Chiasma clathrata, an other geometrid pest of alfalfa.

Male predaceous stink bugs (Pentatom idae: Asopinae) in the genus Podisus release longrange attractant pheromones from a pair of hypertrophied glands opening underneath the wings. Pherom one com positions are reported for four additional Podisus spp.: two Neotropical species (P. connexivus and an undetermined Podisussp.), and two Nearctic species (P. placidus and P. mucronatus). Males of each species release (E)-2-hexenal, plus species-specific major components that include α-terpineol, linalool, 9-hydroxy-2-nonanone, and (E)-2-hexenyl tiglate. The pheromonal chemistry of the Neotropical species closely resembles that for the previously studied Nearctic species, P. macuiiventris and P. fretus.

Heart tissue contains two different thioredoxins. One is a specific mitochondrial protein and is best prepared from pre-isolated, intact heart mitochondria (mt-thioredoxin) whereas mitochondria-depleted tissue homogenates contain the major cellular thioredoxin of cytoplasmic origin (c-thioredoxin). Both heat-stable proteins are clearly differentiated chrom atographically. They exhibit slightly different molecular weights (12300 vs. 12000) and isoelectric points (4.7 vs. 4.8) but differ remarkably in their cysteine content: mt-Thioredoxin has two cysteine residues like the bacterial proteins, and c-thioredoxin possesses six cysteines. Heart extracts were also show n to contain a NADPH-specific thioredoxin reductase of the known mammalian type. A specific function or target enzyme of mt-thioredoxin has not as yet been established.

Conditions of oxidative stress may lead to cataract formation. Reaction of certain flavoproteins, the NADH :oxidoreductases, with different quinones is well known to form hydrogenperoxide. This reaction was investigated to get more information on cataract induction by naphthalene and its quinone metabolites. Protein extracts from bovine lens cortex exhibit “diaphorase” activity, indicated as dye reduction in the presence of NADH and dichlorophenol-indophenol (DCPIP) or ferricyanide. Different redox cycling com pounds are shown to be active in this “diaphorase” reaction by lens protein extract (LCE): Oxygen consumption can be detected in the presence of pyrroloquinoline quinone and juglone whereas 1,4-naphthoquinone, menadione and paraquat are no redox cyclists in this flavoprotein catalyzed reaction.

Naphthalene cataract is probably due to peroxide production through naphthoquinone (NQ) redox cycling and/or glutathione conjugation. Both mechanisms yield losses of essential SH-groups in cristallins and are thus probably involved in protein modification finally visible as lens opacity. 1,2-N aphthoquinone produces H 2 O 2 in the presence of either ascorbate, glutathione, NADH or - to a lesser extend - by homogenates of lens protein preparations. In the presence of 1,2-naphthoquinone and the above reductive additions, both, oxygen uptake and H 2 O 2 formation can be observed. Reductive oxygen activation in these systems are diminuated by iodide in a concentration-dependent manner. Since maleimide-treated proteins are less capable to activate oxygen by 1,2-naphthoquinone, a direct oxygen activation by the interactions of 1,2-naphthoquinone with protein-SH is indicated. Catalysis of “diaphorase”-type (dia) enzymes via NADH - dia - 1,2-NQ - O 2 seems not to operate in hydrogenperoxide production during 1,2-naphthoquinone lens toxicity.

We present quantum bumps obtained from flash experiments at the Limulus ventral nerve photoreceptor under voltage clamp conditions. The results are shown and discussed in form of histograms for the latency, amplitude and net charge transfer (current time integral) of the bump current responses. We argue that the experimental latency histogram s cannot be described satisfactorily by chemical models if one assumes that not more than one photon is captured per flash. Instead of, one has to take into account the Poisson statistics of the captures of 0,1,2 ,... photons released by a single flash. We show that the inclusion of Poisson statistics makes the effective latency histograms of flash responses typically asymmetric and skewed to wards short latencies as compared to that of model histograms for one-photon responses. Our conjecture also implies that under our experimental conditions a fraction of up to 20% of the bump responses evoked by a flash should be suspected to be superpositions of two ore more one-photon responses which cannot be separated by any kind of evaluation analysis. Consequently, the average values of amplitudes and net charge transfers of the light-evoked bump responses are expected to be overestimated as compared to that of true one-photon responses. This hypothesis is confirm ed by a numerical simulation of light-evoked bump responses using experimentally recorded spontaneous bumps (at times larger than 1 s after the flash) as the simulation material. We show that the superposition of one-photon events in the light-evoked bump responses due to Poisson statistics settles the question why their amplitudes and net charge transfers are found to be larger than that of the spontaneous bumps. We suggest that true one-photon responses evoked by a light flash and spontaneous bumps start from the same activated rhodopsin state and take the same biochemical pathway.

This paper analyzes the relationships between the genetic code coevolution hypothesis and the physicochemical hypothesis by means of a comparative study of the precursor-product amino acid pairs on which the former hypothesis is based. Even if the coevolution between the biosynthetic relationships of amino acids and the organization of the genetic code is not questioned in this paper, the results and the arguments used lead us to believe that the selective pressures considered essential by the physicochemical postulates, played a more active role than that of the precursor-product relationships in defining the allocation of these amino acids in the genetic code. It is furthermore pointed out that the two evolutionary hypothesis might be aspects of the same selective pressure, and thus difficult to differentiate.

Carbon-13 and deuteron spin lattice relaxation times T 1 of concentrated solutions of trehalose and sucrose in heavy water are given as function of temperature. From the temperature dependence of the T 1 ideal glass transitions temperatures T o are derived. For both compounds the T o derived for the disaccharides are at high concentrations larger than the T o of the water molecules.

The mutation of the nifA gene of Bradyrhizobium japonicum leads to a pathogenic-like response of the host plant. Soybean nodules induced by the nifA mutant A9 exhibited symptoms of a hypersensitive reaction (HR) normally observed in plant pathogen interactions as localized death of infected cells and the accumulation of the phytoalexin glyceollin I. Little or no glyceollin I was present in nodules elicited by wild type B. japonicum 110 spc 4.

Synthetic peptides selected from HLA-DQ and HLADP glycoproteins were coupled to Sepharose, and used for the isolation of anti-HLA Class II antibodies from the immune sera of rabbits immunized with human lymphoblastoid cells expressing Class II antigens. Antibodies from early and late bleedings displayed remarkable differences in affinity for peptides and for soluble membrane proteins: these differences might be due to an early immune response directed preferentially against surface linear determinants, and to a late response to assembled (discontinuous) sites. The possibility that such antibodies might be used for the identification of amino acid stretches involved in the formation of the same assembled determinant is considered.