Volume 42, Issue 1-2

Mass spectra, X-ray data and high resolution 13 C and 1 H NMR spectra of 8-(dichloroacetyl)-5-hydroxy-2,7-dim ethyl-1,4-naphthoquinone are reported. The antibiotic active com pound was isolated from the fungus Mollisia sp.

The structures of azadirachtin B and of detigloylazadirachtin have been determined on the basis of detailed 1 H and 13 C NMR spectroscopic analysis. Comparison of the data from both com pounds unequivocally proves esterification by tiglic acid at position 3. Our data thus confirm the structure as proposed by Klenk et al., J. Chem. Soc., Chem. Commun. 1986, 523-524.

The isolation of n-alkylphenols from the drupes of Schinus terebinthifolius is described for the first time. The 1 H -NMR parameters, especially those for the triolefinic and monoolefinic components, are presented. Structures were proved by calculated spectra. Skin irritating activities were determined

The methylating capability of methamidophos, assayed by the formation of [7- 14 C]methylguanine in mouse liver, was investigated using a 14 C-insecticide labelled at the O -CH 3 group. Following i.p. administration of the toxicant, [7- 14 C]methylguanine could be isolated from liver nucleic acids of treated mice. The amount of 14 C-label reached its maximum 6 h follow ing administration of the insecticide. At maximum 14 C-labelling, the amount of 7-methylguanine calculated as fraction of applied dose, was 20-22 × 10 -4 and 98 -104 x -4 , for DNA and RNA , respectively. The results obtained indicate also, that an appreciable amount of I4 C-activity is incorporated via the C-1 pool.

The cytogenetic effect of the insecticide methamidophos (0,S-dimethylphosphoroamidothiolate) was studied in mouse bone marrow and mouse spleen cells in culture. In vivo the ability of methamidophos to induce micronuclei and sisterchromatid exchange in mouse bone marrow was investigated. In vitro mouse spleen cells in culture were used to assess the ability of the insecticide to induce chromosomal aberrations and sister chromatid exchange. Three different routes of application for the pure insecticide were tested so as to cover the different possibilities for human exposure to the insecticide. Intraperitoneal, oral and dermal treatment with metham idophos caused toxicity to marrow as indicated bv a significant increase in the percentage of polychromatic erythrocytes (PEs) over that of the control. Methamidophos showed mutagenic potential as evidenced by a positive response in the micronucleus and chromosomal aberrations assays. Thus, single and multiple i.p. injections at 6 and 4.5 mg methamidophos/kg body w t., oral administration of the insecticide for 14 consecutive days at a dietary level of 100 ppm and multiple dermal treatments (total 4) with 24 mg/kg body wt. induced a statistically significant increase in the frequency of PEs with micronuclei in mouse bone marrow. Moreover, the tested concentrations of m etham idophos as low as 0.25 ng/ml induced a high percentage of metaphases with chromosomal aberrations in cultured mouse spleen cells. Methamidophos is a weak inducer of SCEs in mouse bone marrow and cultured mouse spleen cells.

The quinone 2-methoxy-6-n-pentyl-2,5-cyclohexadien-1,4-dion (Primin) was found to possess molluscicidal activity against the snail Planorbis corneus. The synthesis of homologous and isomers and the results in the biotests are reported.

Four plant bioregulators were tested for their effects on production of valepotriates in Valeriana wallichii and Fedia cornucopiae cell suspension cultures. Concentrations of more than 10 ppm reduced valepotriate yield. At lower concentrations production was increased. For optimal activity, bioregulators had to be applied during early exponential growth, up to day 8 of the growth cycle. At equimolar concentrations dim ethylm orpholinium bromide (4 ppm) and dimethylpiperidinium chloride (3 ppm) significantly im proved total valepotriates in V. wallichii (up to 23%) and in F cornucupiae (up to 50% ) 2-(3,4-dichlorophenoxy ) - triethylamine (6 ppm ) and 2-(3,5-diisopropylphenoxy)-triethylam ine (6.4 ppm) increased valepotriate production in both cell cultures up to 40%. With dimethylpiperidinium chloride and dimethylmorpholinium bromide the ratio of m onoene to diene valepotriates in both cell systems was significantly shifted to the m onoene com pounds. A general use of these bioregulators to increase production of terpenoid secondary m etabolites in plant tissue cultures is indicated.

Modulation of the N-azo coupling betw een ring-substituted arenediazonium chlorides and premixed m ethylam ine-form aldehyde leads not only to 1-aryl-3-hydroxymethyl-3-methyltriazenes and their dimers, but also to unexpected cyclic and com plex products. The syntheses comprise reactions with arenediazonium chlorides bearing both - M and + M substituents at para and ortho/para positions of the phenyl ring. One of the major constituents isolated from a mixture of products is the O-acetate ot 3-hydroxymethyl-3-methyl-1-(2,4,6>-trichlorophenyl)triazene. This product was obtained from the reaction of 2,4,6-trichlorobenzenediazonium chloride and methylamine- formaldehyde mixture which was then stabilized by acetylation. The structures of the isolated products could be derived from reactive methylolamines and electrophilic intermediates that possibly occur in vivo and thereby offer a plausible mechanistic explanation for the carcinogenic and tumour-inhibitory activity associated with the open-chain triazene com pounds in the living cell.

Non-epithelial intermediate filament (IF) subunit proteins show a high and specific affinity for core histones at physiological ionic strength. When IF proteins are titrated with a mixture of core histones and linker histone H1, in general the latter is totally excluded from com plexation and in the adducts formed the moderately-arginine-rich histones H2A and H2B are progressively replaced by the very-arginine-rich histones H3 and H4. At histone saturation, 2 molecules of nonneuronal IF protein bind 1 histone HI molecule or 8 core histone m olecules, whereas due to its glutamic acid rich, C terminal extensions one dimer of thp 68 kD npnrofilament nrotein associates with 3 molecules of histone H1 or 24 molecules of core histones. The salt stability of the insoluble association products is dependent on the amount and arginine content of the constituent histone species. Rem oval of the non-α-helical N- and C-terminal polypeptides from IF proteins by partial chymotryptic digestion does not affect their histone-binding characteristics. Since core histones are only partially inactivated by limited tryptic digestion, they also appear to react through their a-helix-rich central domains; the limit peptide derived from histone H1 is com pletely inactive at physiological ionic strength. Affinity chromatography of rod domains of IF proteins on core histone-Sepharose 4B and of histones and their limit peptides on vim entin-Sepharose 4B has shown that the interactions involving fractions of histones H3 and H4 are extrem ely resistant to salt and can be dissociated only with arginine or salt under denaturing conditions. In general, the experim ental results revealed close parallels between the association of histones with IF proteins and their interaction with DNA.

The metabolization of trans-2-hexenal - one of the main components of plant wound gases with antibiotic activity - was investigated for 5 different isolates of Botrytis cinerea PERS. The transformation products as well as the kinetics of their formation were analyzed. Isolates exclusively mycelium forming (Bc 1 and Bc 13) transformed tr-2-hexenal into tr-2- hexenol, while sporulating isolates (Bc 3, Bc 9 and Bc 10) converted tr-2-hexenal to hexanol-1. Basically the metabolization of tr-2-hexenal proceeded in the same way via the aqueous phase as in the gas phase. The transformation products tr-2-hexenol and hexanol-1 showed significantly lower toxicity against the tested B. cinerea isolates than tr-2-hexenal. In each isolate the end product of tr-2- hexenal conversion had the weakest inhibitory activity. The transformation reactions thus represent detoxification mechanisms for these fungi.

The growth of root hairs from stems of Nicotiana tabacum and Spartium junceum was induced by Agrobacterium rhizogenes. Stem roots of Nicotiana contained nicotine and a second alkaloid, probably anabasein. Root hairs from stems of Spartium contained cytisine, N methylcytisine and anagyrine as major alkaloids. The alkaloid content of the tumors was 3-12 times higher than that of the respective healthy host tissue.

The resinous material excreted by aerial parts of snakeweed, Gutierrezia sarothrae, was analyzed for flavonoid aglycones. Six major flavonoids were characterized by their spectral data; nine minor flavonoids were identified by direct comparisons with markers. The major flavones form three pairs of isomeric products, five of which are 6,8-methoxylated.

Labelled 3β-hydroxy-5β-pregnan-20-one was incorporated by Digitalis purpurea plants into digitoxin while 3β-hydroxy-5β-pregn-8(14)-en-20-one was not. This result excluded the intermediary role of the latter compound as precursor of cardenolides in the mentioned plant.

Humulene was transform ed by strains of Chaetomium cochlioides and Diplodia gossypina ATCC 10936 affording 33 new humulanes. The first step is a non-enantioselective epoxidation of the 1,2-double bond, whereas the subsequent hydroxylations were performed preferably at the 1R ,2R -epoxide. To proof these results the anti-1,2;8,9-diepoxide was synthesized and used as substrate. While the 1R ,2R ,8S,9S-antipode of this diepoxide was hydroxylated the other one was recovered unchanged. Diplodia gossypina ATCC 10936 gave the same products but differing in relative and absolute configurations.

Cell-free extracts from oak (Quercus robur) leaves catalyze the transfer of the galloyl-moiety of β-glucogallin (1-O-galloyl-β-D-glucopyranose) specifically to the 6-position of the same compound, yielding 1,6-di-O-galloyl-β-D-glucopyranose, an intermediate of gallotannin biosynthesis, β- Glucogallin thus functions as both donor and acceptor molecule in this reaction. The partial purification and some general properties of this new acyltransferase are reported.

PAPS-reductase from Escherichia coli was em ployed to detect thioredoxins from pro- and eukaryotic organisms. A simple method for the isolation of this enzyme and properties of the enzymatic assay were described. A comparison betw een thioredoxins detected by the PAPS-reductase and the Fructose-bisphosphatase or NADP malate dehydrogenase was used to assess the validity of the test. The high cross-reactivity of the bacterial enzyme was useful in the purification of heterologous thioredoxins from spinach, Synechococcus, and Saccharomyces cerevisiae.

The energy requirement for protein transport into chloroplast was assayed under conditions that perm it to distinguish whether the posttranslational translocation is dependent on ATP or whether a membrane potential across the chloroplast envelope can drive this transport event. A membrane potential is not required for translocation. ATP can support protein transport in the presence of protonophores and ionophores. Non-hydrolyzable ATP analogues and GTP, CTP, UTP cannot serve as ATP substitutes. Translocation could be observed when an ATP generating system was used to supply ATP. In contrast ATP degrading systems completely abolished translocation. The inner envelope mem brane localized ATP-ase is probably not involved in the transport event. The results suggest that ATP is needed at the outer chloroplast envelope. Inhibition of protein transport by ADP, pyrophosphate and NaF is studied and its consequences discussed.

Interaction of the antenna protein B 800-850 with the lipid environment and with the reaction center of the photosynthetic bacterium Rhodopseudomonas sphaeroides is studied by fluorescence spectroscopy, transient absorption techniques, light scattering and electron microscopy. Using vesicles of synthetic phospholipids it is shown that solidification of the m em brane causes a structural protein change evident from a reduction in fluorescence quantum yield. The change occurs at a tem perature up to 5 °C below that corresponding to the gel/fluid transition tem perature and indicates local melting. The structural change is not specific for the lipid head group nor chain length (investigated for lengths of 12 to 16 CH 2 groups) and can be understood applying a simple elastic model. It can also be included isothermally by changing the ionic milied and thus varying the lipid phase state. Energy transfer LHCP→RC is proven to be highly efficient in model m em branes and is not affected by the existence of a phase transition. This indicates two LHCP fractions one tightly - and one non-bound to the RC.

In daffodil, development of chloroplasts or chromoplasts is not accompanied by plastid DNA modification. This has been shown by comparing the fragment pattern of different restriction endonucleases, and by the lack of methylation of CCGG sequences. A physical map has been constructed for the plastome using the restriction endonucleases Sal I, Pst I, and Bgl I. The fragments containing the genes for the large subunit of ribulose bisphosphate carboxylase/oxygenase (rbcL), the alpha, beta, and epsilon subunits of the ATP synthase complex (atpA , atpB, atpE ), cytochrome f(petA ), and for the 51 kDa chlorophyll apoprotein of photosystem II (psbB) have been identified. The respective gene locations correspond to those on spinach chloroplast DNA.

The uptake of protoporphyrin IX by Propionibacterium acnes in suspension has been studied by fluorescence spectroscopy. Protoporphyrin, after it was injected into a cell suspension, was firstly bound to receptors on the cell surface and in this state protoporphyrin was non-fluorescent. Subsequently, probably as a result of lateral diffusion in the cell wall, these protoporphyrinreceptor complexes formed dimers. The final step in the overall uptake process of protoporphyrin by the cells from the surroundings consisted in a jum p of such dimers from waterlike to lipidlike com partm ents in the cell mem brane where protoporphyrin became fluorescent. The lipidlike com partm ents in the cells had a limited binding capacity of protoporphyrin. The fraction of surviving cells versus light dose has also been studied for varying amounts of protoporphyrin added to the cell suspensions. The survival curves were exponentially decaying with the irradiation time and there was a direct proportionality between the inverse slope of the survival curves and the intensity of protoporphyrin fluorescence from the lipidlike compartm ents. The relevance of these results to the therapy of Acne vulgaris is also discussed.

Two E. coli tRNA Ile sequences were compared against those of 36 other E. coli tRNAs. tRNA Ile 1 was found to bear high similarity with tRNA Val 1 (E = 1.11 x 10 -18 ) while tRNA Ile 2 had the greatest match ( E = 3 .4 0 x 10 -19 ) with tRNA Lysl (E is the expected number of such matches, per search, based on coincidence). These matches, which we consider to represent homologies, extend from base 7 to base 67 in the former and base 7 to the end (76) in the latter pair. These results coupled wim otners on tne lower activity ot isoleucine in reactions postulated to be important in primitive protein synthesis (i.e. esterification reactions and non-enzymatic activation by ATP [1-3]) lead us to propose that isoleucine was included among the proteinaceous amino acids, and received its anticodonic assignment, relatively late in evolution through mutation of tRNAs previously employed for other amino acids.

Hydroxyl radicals were generated by pulse radiolysis of N 2 O-saturated aqueous solutions. The OH radicals react with thiols such as 1,4-dithiothreitol by abstracting S-bound H-atoms (k = 1.5 × 10 10 dm 3 mol -1 s -1 ). The 1,4-dithiothreitol radical HSCH 2 (CHOH) 2 CH 2 S (2) closes the ring (k - 1.5 × 10 6 s -1 ) thereby forming the three-electron-bonded species (3) in equilibrium (2, 3: ε(380 nm) = 450 dm 3 mol -1 cm -1 ) which has a reported pK a value of 5.2. At pH 7.2 3 deprotonates (k = 10 5 s -1 ) yielding the ring-closed radical anion 4 which is characterized by a strong absorption at 390 nm (ε = 5900 dm 3 mol -1 cm -1 ). The deprotonation of 3 is catalyzed by HPO 2 4 (k - 6 .8 × 10 7 dm 3 mol -1 s -1 ). The ring-closed radical anion 4 has also been generated by reacting trans-4,5-dihydroxy- 1,2-dithian with the solvated electron and the H atom in Ar-saturated solution. Its protonation by protons yielding 3 occurs with k - 1.8 × 10 10 dm 3 mol -1 s -1 , with H 2 PO - 4 with k = 3.8 × 10 7 dm 3 mol -1 s -1 . In its reaction towards tetranitrom ethane 3 differs from its non-complexed analogue HOCH 2 -CH 2 S‘ (1) in that 3 is rapidly oxidized (k = 2 × 10 8 dm 3 mol s -1 ) while 1 is not. Isoenergetic H transfer from 1,4-dithiothreitol to 1 occurs with a rate constant of k = 1.7 × 10 7 dm 3 mol -1 s -1 . 1,4-Dithiothreitol has pK a values at 9.12 and 10.15. At pH 9.1 thiolate ions com pete with ringclosure by complexing with 2 thereby forming an open-chain disulphide radical anion 6 which has very similar spectral properties as its ring-closed analogue 4.

The volatile components of the Dufour gland secretion of workers cf the ant Harpagoxenus sublaevis have been analysed by gas chrom atography and mass spectrometry. Over 20 com ponents have been identified, consisting of linear and terpenoid hydrocarbons. Each gland contains approximately 6 ng of hydrocarbons with n-heptadecene (40% ) and /n-heptadecadiene (30% ) as the major components. Two terpenoid hydrocarbons, (E)-β-farnesene and a homofarnesene isomer were also identified.

Silylated phenolic fraction from two propolis samples was investigated by capillary GC/MS. Fifteen phenolic acids (seven of them new for propolis), two ketones, new for propolis, eight esters of phenolic acids, new for propolis, six flavanones and two flavonols, were tentatively detected.

The functional properties of the light adaptation system in the superposition eye of the moth Agrotis segetum have been investigated by reflection spectroscopy. The spectrum of the reflected light from the tapetum of dark adapted eyes had a peak at about 580 nm corresponding to a spacing between the reflecting layers in the tapetum of 145 nm. During light adaptation of the eye the observed reflectance changes could be explained by light extinction in one screening pigment. The shape of the extinction spectrum of the screening was constant throughout the adaptation process and after a lag phase the optical density of the pigment in the light path increased linearly with time. The screening pigment caused light extinction both by absorption and to some degree also by scattering. The absorption spectrum of the screening pigment had a broad maximum about 590 nm and the scattering efficiency of the pigment particles seemed to be nearly independent of the wavelength.

The epicuticular wax of Cistus albanicus was investigated. Homologous series of alkanes, wax esters, alcohols and long chain fatty acids were identified. The alcohol fraction consisted of primary and most probably of secondary alcohols.

Ceratocystis coerulescens RWD 451 (Ascomycetes) was cultivated on defined synthetic culture medium containing glucose (3.0% ), asparagine (0.1% ), mineral salts and thiamine. 6 weeks oid cultures produced methyl heptenyl compounds, citronellol. citronellyl acetate, dihydro-farnesol and a hitherto unknown monoterpene alcohol. By GC/MS, 1 H and 13 C NMR spectroscopy, this alcohol could be identified as ( -)isopulegol. So far, this monocyclic monoterpene alcohol has not been described as a fungal metabolite.

Inhibition of 3′:5′-cyclic-AMP-5′-nucleotidohydrolase (EC 3.1.4.17) type II (cAMP-phosphodiesterase) by sodium molybdate was studied: While determination of inorganic phosphate and 5′-ribonucleotide phosphohydrolase (5′-nucleotidase) (EC 3.1.3.5) activity was not disturbed by sodium molybdate in concentrations up to 10 mM , cAMP-phosphodiesterase was inhibited by millimolar concentrations of molybdate. The half maximal effect was observed at about 2 mM sodium molybdate (0.75 mM cAMP in the assay).

During field screening of polyene compounds, conducted at different localities of Hungary, in 1984, sex attractants were found for Costaconvexa polygrammata Bkh., as (3Z , 6Z , 9Z)-3,6,9-eicosatriene; Eupithecia subnotata Hbn., as (3Z , 6Z , 9Z)-3,6,9-nonadecatriene; Synthomis phegea L., as (3Z , 6Z , 9Z)-3,6,9-heneicosatriene and for Dysauxes ancilla L., as (3Z , 6 Z, 9Z )-l,3,6,9-nonadecatetraene. (7E , 9Z)-7,9-dodecadienyl acetate was found to be a sex attractant for Sterrha rubraria Stgr.

From the volatile constituents of the solitary bee species Panurgus banksianus Kirby and P. calcaratus Scopuli 31 compounds could be identified. The cephalic secretion of P. banksianus is a sex specific m ulticomponent mixture of terpenes and aliphatic carboxylic acid esters while P. calcaratus contains monoterpenes only. The major component in the Dufour’s gland secretion of P. banksianus is a macrocyclic lactone, eicosanolide, which is absent in P. calcaratus. In contrast, the new terpenoid ester, geranyl citronellate, is present in large amounts in the Dufour’s gland secretion of P. calcaratus.