Volume 44, Issue 3-4

The essential root oils of the Pim pinella saxifraga subspecies eusaxifraga, alpestris and nigra were investigated by means of capillary GC and spectroscopic m ethods. A total of 48 com ponents was identified. Characteristic constituents of all oils are pseudoisoeugenol derivatives, sesquiterpenoids and trinor-sesquiterpene hydrocarbons, but the qualitative and quantitative patterns of the individual oils exhibit significant differences. The results obtained were discussed in regard to a chemotaxonomic reliable delimitation of the 3 described subspecies of Pimpinella saxifraga L.

The composition of the essential root oil of Peucedanum lancifolium Lange (A piaceae), growing in the Atlantic coast regions of France, North-West Spain und Portugal was investigated by chromatographic and spectroscopic methods. Forty five terpenoids, many of them of already known structure, and several aliphatic esters have been identified. Besides the main constituent, the irregular sesquiterpene alcohol trans-sesquilavandulol, representing 53.37% of the total oil, some hitherto unknown sesquiterpenoids with an irregular carbon skeleton were found. The possible chemotaxonomic value of the essential oil com position with regard to the close botanical relationship of Peucedanum lancifolium Lange and Peucedanum palustre (L .) M oench is briefly discussed. With the exception of these two species, trans-sesquilavandulol was not found in any other member of the genus Peucedanum.

In Plagiomnium cuspidatum the three new dihydrobiflavones 2,3-dihydro-5′-hydroxyam entoflavone, 2,3-dihydro-5′,3‴-dihydroxyamentoflavone and 2,3-dihydro-5′-hydroxyrobustaflavone were detected.

The chemical composition of the leaf surface wax of Euphorbia lathyris L. was analysed using TLC, GC, GC-MS and NMR. A predominance of pentacyclic triterpenoids and primary alcohols was observed. They together constituted 60% of the total wax. Seven triterpenols: taraxerol, β-amyrin, lupeol, isomotiol, a-fernenol, simiarenol. Ψ-taraxasterol and eight triterpenones: taraxerone, β-amyrinone, lupenone, isomotione, a- and β-fernenone, simiarenone and filicanone were isolated. Among them , β-amyrin and lupeol were found esterified with homologous series of fatty acids. The minor part of wax was formed by long chained and predominantly saturated alkanes, wax esters, aldehydes and free fatty acids.

The cyanobacterium Synechococcus leopoliensis (Anacystis nidulans, strain L 1402-1) grown at 39 °C and 2 vol. % CO : could be synchronized by a light/dark regime of 3:5 h (white light intensity 1.5 × 10 4 erg cm -2 sec -1 ). Content of pigments (chlorophyll a. phycocyanin and carotenoids), R N A and proteins increased linearly up to 100% at the end of the light period while DNA synthesis was lower. Chlorophyll a synthesis was correlated to the photosystem I activity of the isolated thylakoids and to the formation of MGD G . Galacto lipids were synthesized in the light period, only. A lag phase of 2h was observed in the biosynthesis of SQDG and PG. No significant differences were found between the cell and thylakoid fractions. Palmitic (C 16:0 ), hexadecenoic (C 16:1 ) and octadecenoic (C 18:1 ) acid as major com ponents accounted for more than 90% of total fatty acids in MGD G , DGDG and SQDG . PG contains a small amount of stearic (C 18:0 ) and heptadecenoic (C 17:1 ) acid. No significant variations in the fatty acid distribution of all lipids could be detected in the cell fraction during the division cycle. Changes in the ratio of saturated to unsaturated fatty acids were found in isolated thylakoids. only. In experiments with [ 14 C]bicarbonate main radioactivity was measured in galacto lipids while using [ 14 C]acetate SQDG and PG were markedly [ 14 C]labelled. Results were discussed with reference to the findings of eucaryotic algae and the formation of photosynthetic membranes.

Superoxide reacts with catechol in dimethylsulfoxide to form a yellow product. The structure of this product was established by different methods. UV-spectra of the yellow compound corresponded under all circumstances to those of enzymatically produced 2-hydroxymuconic acid semialdehyde. In the presence of ammonia the product was converted to picolinic acid. The UV - spectra, the retention times in GC and HPLC and the mass spectra after methylation corresponded to those of authentic picolinic acid. Thus, the yellow reaction product was unequivocally identified as 2-hydroxymuconic acid semialdehyde. The formation of the me/a-cleavage product was only observed at low catechol concentrations (<2 mM) , at higher concentrations the product was not formed. The reaction was complete within 10 min at room temperature. Spectroscopic data suggest a first intermediate in the reaction with λ max = 804 nm, 772 nm and 660 nm.

Semiempirical CNDO/2 calculations of the electronic and molecular structures have been car­ried out for six antitumor anthracycline antibiotics. The strong correlation between their biological activity and such molecular properties as chromophore dipole moment, energy of frontier orbitals, and steric volum es was found and the proper QSAR equations were constructed. The molecular mechanics model showed the weak dependence the DNA-intercalation energy on differences in chromophore constituents (C4 and C9 positions). The expression for the intercala­tion energy has also been obtained on the ground of a quantum-statistical approach: it was applied to the DNA -intercalation energy calculations for the same six anthracyclines. The results agree with the molecular mechanics ones quite satisfactory.

A quantitative structure-activity relationship (QSAR ) approach by use of the minimal topological difference (MTD) method including 46 derivatives of 4-androstene-3,17-dione and 5a-androstane- 3,17-dione is applied to give indications about the role of the C 4 = C5 double bond in competitive inhibition of human placental aromatase and about sterical requirements in steroidaromatase interactions. The inhibitory activity is found to correlate with the sterical MTD variable, hydrophobicity, and π-system conjugation in the A,B-ring region. A comparison of the MTD results reveals a good agreement with interpretations based on freeenergy data derived from inhibition constants. By means of MM2 molecular mechanics and PCILO quantum-chemical calculations, the 4-ene structure is shown to significantly influence conformational features of C19 substituents which are important in enzym atic transformations. While 19-hydroxy-5a-androstane-3,17-dione favour a conformation having the hydroxyl group in the enzyme-directed out-of-ring position, the C 4 = C5 double bond energetically enables the steroid to adopt a conformation which can be hydroxylated without internal rotations. According to present theoretical findings, the 4-ene unsaturation thus exerts an indirect conformational influence by hydroxyl positioning appropriate to aromatase interactions and a direct electronic influence by π conjugation.

A series of N-phenylcarbamates induced flowers in one-month-old seedlings of Asparagus officinalis L. Ninety to 100% of the plants flowered when the seeds were germinated in the presence of the most potent members of this class. The flowering occurred only once at the top of the seedlings, which then continued to grow normally. This made it possible to select the commer­cially preferred m ales of this dioecious plant at the seedling stage. Both male and female flowers were fertile, so cross-breeding was possible between flowering seedlings as well as between flowering seedlings and adults that had grown normally. Activity of flowering induction was not related with inhibition of photosystem II activity.

Akademie der Wissenschaften der DDR , Institut für Biochemie der Pflanzen, Weinberg 3, Halle/Saale. The metabolism of [1- 14 C]2-(2,4-dichlorophenoxy)-propionic acid was studied in excised plants and cell suspension cultures of tomato. It was rapidly taken up and metabolized by both the plants and the cultured cells. The metabolites, isolated by extraction with aqueous acetone, separated and purified by TLC and HPLC, were identified by chemical and spectrometric methods. Conjugates with carbohydrates were detected. Glucose, diglucose as the main conjugating moiety, and triglucose were found as carbohydrate components within the conjugates. Almost the same conjugates occurred in plants and cultured cells.

In tissue cultures from shoot apex meristems with leaf primordias of Vicia faba cv. TP667, addition of low concentration of auxins (0.01 mg ・1 -1 ) induced regeneration of whole plants at high frequency (100%). The combination of NAA and kinetin or GA , also induced a high yield of plant regeneration. Regenerated plants from various cultivars on a medium with 2 ,4 D (0.01 mg ・1 -1 ) were infected with Botrytis cinerea, Phytophthora megasperma and R hizoctonia solani. Accumulation of phytoalexins, ethylene production and the resistance to fungal diseases were studied. In general, production of phytoalexins occurred at a high level in all cultivars infected with B. cinerea. Ethylene production varied more in the seven cultivars studied than phytoalexin accumulation. No cultivar was resistant to B. cinerea. The highest resistance and the low est concentration of phytoalexin was found after infection by R. solani, and phytoalexin accumulation and resistance were interm ediate in plants infected by P. megasperma. The data suggest that only low to medium concentrations of phytoalexin in faba beans are correlated with resistance of regenerated plants.

Fluorescence induction m easurem ents indicate a PS II α,β heterogeneity in the unicellular green alga Chlorella fusca. Under normal light the relative amount of PS α II is decreasing from 63% to 43% during the first 8 h of the cell cycle. The rate constant k α shows a concomitant decrease by 20% in the first 4 h. The relative amount and antenna size of PS II α is restored in the dark. In contrast, PS II β - centres show the same rate constant during the whole cell cycle. Under low light k α is increased by approximately 30% over the cell cycle, whereas no change of k β is observed. It is concluded that the PS II organization of Chlorella is changed both during the cell cycle and the adaptation to light intensity by a complex regulation (i) involving the antenna size of α-centres and (ii) changing the quantitative relationship of fast and slow reaction centres.

Im mobilized tobacco cells excreted only a small proportion of their main secondary metabolite caffeoylputrescine into the culture medium . Experiments designed to release more of the com ­ pound by permeabilization caused the loss of caffeoylputrescine, probably by oxidative reactions. Moreover, rather mild treatments with permeabilizing agents (e.g. n-propanol) resulted in severe growth inhibition. The ability of permeabilized cells to form caffeoylputrescine and other hy-droxycinnam oyl conjugates from phenylalanine decreased considerably even when such cells were still able to metabolize phenylalanine into various ethyl acetate extractable com pounds (e.g. hydroxycinnamic acids and acetophenones). The formation of new biotransformation products suggests that permeabilized cells could be used as a tool for testing the enzymatic capabilities of a cell culture.

2-A zido-s-T riazines, Connectivity Indices. Inhibition of Hill Reaction. Photosystem II, OSAR Modelling This study was undertaken to find a simple and accurate structural parameters for the quantita­tive description of inhibitory potency of 2-azido-s-triazines in Hill reaction and to gain more information about the mechanism of inhibition on molecular level. A very good correlation (r = 0.946) was obtained between the pl 50 values (the negative logarithm of the molar concentra­tion that causes 50% inhibition) and the valence zero-order and the difference between the second-order and the valence second-order molecular connectivity indices. This model, when com pared with the empirical models based on the 1-octano/water partition coefficients and the chromatographic retention data, shows superior performance in accuracy and range of applicabili­ty. In addition, the direct correspondence between molecular structure and above connectivity indices makes it possible to locate structural features responsible for the inhibitory potency of 2-azido-s-triazines in Hill reaction. From our OSAR analysis, the interaction between the chloro­plast receptor site and 2-azido-s-triazines, which causes inhibition of Hill reaction, is primarily influenced by the size of alkylamino substituents and it accounts for the most variation in the pl 50 data. The structural features of secondary importance that control the magnitude of pl 50 ’s are the polarity of alkylamino chains and the degree of branching on alpha carbon atom of R:alkylamino substituent. Com pared with the main factor, the size of alkylamino substituents, they can be viewed as a fine tuning elements for the inhibitory potency of 2-azido-s-triazines.

Chlorophyll Fluorescence. Fluorescence Quenching, Hydrogen Peroxide. Active Oxygen. Ascorbate Peroxidase Chlorophyll fluorescence quenching induced by H 20 2 in intact spinach chloroplasts was investi­gated with a modulation fluorometer which allows to distinguish between photochemical and non­ photochemical quenching components by the so-called saturation pulse method. Residual catalase activity was removed by washing and percoll gradient centrifugation. H 2 O 2 was found to induce pronounced photochem ical and non-photochemical quenching, characteristic for the action of a Hill reagent, with a half-maximal rate already observed at 5 × 10 -6 m . The saturation characteris­tics and maximal rate of H 2 O 2 -reduction were very similar to those of methylviologen reduction. H 2 O 2 -dependent quenching was stimulated by ascorbate and inhibited by cyanide and azide in agreement with previous findings by other researchers that H 2 O 2 -reduction involves the ascorbate peroxidase scavenging system and that the actual “Hill acceptor” is an oxidation product of ascorbate, i.e. monodehydroascorbate or dehydroascorbate. With well-coupled intact chloro­plasts reducing CO 2 at 150 (μmol (mg Chl) -1 h -1 , iodoacetamide stopped CO 2 -dependent O 2 -evolution and consequent addition of 10″3 m H 2 O 2 produced an O 2 -Solution rate of 240 (μmol (mg Chl) -1 h -1 .It is concluded that light-dependent H 20 2 reduction is a very efficient reaction in intact chloroplasts. As H 2 O 2 formation and consequent reduction also occur in vivo, the corre­sponding quenching should be considered when assimilatory electron flow is estimated from quenching coefficients. It is suggested that proton flux associated with H 2 O 2 -formation and reduc­tion may be important for the adjustment of appropriate ATP /NADPH ratios required for CO 2 -fixation in vivo. Furthermore, H 2 O 2 -reduction may serve as a valve reaction whenever Calvin cycle activity is limited by factors different from NADPH supply, thus protecting against photo-inhibitory damage.

The mode of action of three types of conjugated enamine compounds was investigated by means of thermoluminescence measurement. Cyanoacrylate and 2-(l-ethoxyethylam inom ethylidene)- 4-dodecyl-5,5-dim ethyl-cyclohexane-1,3-dione (ACm12) converted the B-band (30 °C) arising from S 2 Q B - charge recombination to a downshifted 6 °C-band. This band was proved to be identical with the DCM U-induced Q-band (6 °C) arising from S 2 Q A - recombination, indicating that these two compounds block Q A to Q B electron transport. 3-(1-dodecylam inopropyridene)-6- methyl-2H-pyran-2,4-dione (APp12 ), on the other hand, induced an abnormal band peaking at 15 °C between the Q-band and B-band. From the gradual downshift of its peak temperature in titration experiments, this band was assigned to arise from a modified S 2 Q B - charge pair, in which the properties of either Q B - or S 2 is altered. The 15 °C-band showed normal oscillation during the first 2 flashes, but the oscillation was interrupted thereafter. Another therm oluminescence analysis by use of post flash low temperature illumination protocol revealed that APp 12 affects neither Q A to Q B nor Q B 2- to PQ electron transport, but specifically blocks S 3 to S 0 transition. These results indicate that APp12 is a new-type PS II inhibitor.

The residual motion of spin labels bound to cysteine β93 of methemoglobin and oxyhemoglobin has been analyzed as a function of temperature and hydration. The rotational diffusion of the whole protein molecule has been prevented or restricted by crystallization, lyophilization or by high viscosity of the solution. The residual motion of the labels is characterized by an angle of the limited motion cone and their rotational correlation time using computer simulations of the E PR spectra. Two types of motion can be separated due to different correlation times and different dependenceson temperature and hydration. One of these motional mechanisms can be shown to be determined by protein fluctuations. Correlation times of these fluctuations decrease from 2 × 10 -8 at T = 220 K to 10 -9 s at T = 300 K in the samples of high water concentration. Strong correlation between the properties of the hydration shell and these fluctuations are observed.

Rat, Intestinal Absorptive Cell, Basolateral Plasma Membrane, Protein Isolation. Chemical Characterization The protein pattern of the basolateral membrane (BLM) of the rat small intestinal absorptive cell shows about 20 major and a multitude of minor bands. A simple and efficient method is described for isolation and purification of a major protein in the 17 kDa molecular weight (M W)-range called Prot 17 . The isolated BLM of intestinal epithelial cells was dissolved in buffer 1 (Tris/HCl, 2% SD S, 10% glycerol, 5% β-m ercaptoethanol. pH 6.8) and subsequently dialyzed for 4 h against buffer 2 (Tris/glycine, pH 8.3) and then for 12 h against buffer 2 containing 25% methanol. The resulting precipitate contained Prot 17 and phospholipids in the form of liposom es. A ll other BLM proteins remained dissolved in the supernatant. Chemical characterization of Prot 17 suggested that it is an integral membrane protein amounting to about 5% of the total BLM protein. Amino acid analysis revealed a MW of 17.6 kDa. The Prot 17 molecule did not contain any PAS-positive carbohydrates. In its isolated form, and apparently also in the BLM . Prot 17 occurred as a polymerized structure with a MW of about 90 kD a. By dissolution in buffer 1 and heating to 100 °C for 1 min the complex was split into its 17 kD a subunits. By oxidation with performic acid it was also broken down into its subunits. A specific antiserum against Prot 17 was obtained from immunized Balb/c mice. Immunofluorescence labelling of rat small intestinal sec­tions with this serum showed that Prot 17 was not a BLM -specific protein. It occurred in both plasma membrane domains of the intestinal absorptive cell.

As part of a continuing study on the origin of genetic coding and the process of protein synthesis, we have compared sequences of a large number of transfer RNAs and several 5S ribosomal RNAs using automated routines. Transfer RNAs were found to exhibit a high degree of matching with 5S rRNA s. These matches are considered to be indicative of sequence homology, reflecting common ancestry of the two molecules. Other possible explanations for the matches (convergence, transfection) are discussed and found to be highly implausible. Matches are also found between 5S rRNA and the introns of yeast precursor tRNAs. Many matches extend from before the 3′ end to after the 5′ end of circularized 5S rRNA sequences. Data is presented which indicates a tandemly duplicated or circular molecule could have served as the precursor to both 5S rRNA and tRNA . A derivative of this molecule may have functioned as a universal translator before the evolution of the highly specialized tRNAs.

In the presence of sulfhydryl compounds nitracrine, an anticancer drug, binds covalently to DNA . The accessibility of DNA in chromatin both to nitracrine and to 8-methoxypsoralen. which was used as a reference compound in this study, when assayed in NaCl concentrations from 0 to 2 m show similar characteristics. The initial decrease reaches a minimum at 0.15 m NaCl above which dissociation of non-histone proteins and histones at higher ionic strengths is demonstrated by an increase in accessible sites. The relative accessibility of DNA in chromatin to nitracrine is, however, lower than that found for 8-methoxypsoralen. Partial dissociation of chromatin with 0.7 m NaCl increases the accessibility of DNA in chromatin when assayed in the absence of NaCl but has no apparent influence when estimated at ionic strength close to physiological conditions.

An algorithm is described for the construction of phylogenetic trees. The algorithm is based on the progressive correction of data along the tree construction. For the correction, the average value of the difference between each pair of neighbour elements to the rest of the table is considered.

The ED 50 for moulting inhibition by injected azadirachtin A is for fourth instar larvae of all the triatomines, Triatoma vitticepes, T. pseudomaculata, T. maculata, T. brasiliensis, T. lecticularis, T. matogrossensis, T. infestans, Rhodnius prolixus, R. neglectus, R. robustus, Panstrongvlus megi-stus, and P. herrera in the range of 10-25 ng/larva. In Rhodnius prolixus, the survival of T. cruzi was studied after treatment with the drug. If the trypomastigotes were fed in presence of 1.0 (.μg aza­ dirachtin A/ml blood, the number of parasites decreased near to the limit of detection within 30 days. If the drug was applied 20 days after T. cruzi infection, it still completely abolished the parasite in the host’s gut within the subsequent 20 days. The same holds true if the insect larvae were pretreated with azadirachtin A 20 days before the subsequent infection with T. cruzi. Azadirachtin A, if applied at the dose of 1 (μg/ml, did not affect the hemolytic activity of the crop contents or the proteinase content of the intestine. A parallel between azadirachtin effects on the hormone balance of the host and growth inhibition of the parasite is discussed on the basis of the present results.

The monotypic genus Leucocyclus, consisting of only two subspecies, was investigated for its exudate flavonoids. Major flavonoid aglycones were polymethoxyderivatives of 6-hydroxyflavones and, to a lesser extent, of 6-hydroxyflavonols. The two subspecies differed in their accumula­tion trends, but populational variation reduced the significance of taxonomic implications. Cultivated material corre­sponded largely to the original material collected from nature. The accumulation trends of Leucocyclus coincide with those of the closely related genus Achillea.

From workers of Mourella caerulea, an extremely rare and primitive species of neotropical stingless bees, 101 cephalic volatiles were identified by GC/MS analyses.

Treatment of cell smears, paraffin, and Epon tissue sec­tions with aqueous solutions of curcumin results in a green fluorescence reaction in chromatin under violet-blue exci­tation which is abolished after extraction procedures with DNase and TCA. The selective fluorescence characteris­ tics of curcumin support the possibility of employing this dye as a new fluorochrome.

Reticulosphaera socialis n. gen.. n. sp., a protist deriving from the Caribbean Sea. could be isolated and cultivated. The life cycle consists of a benthic and a pelagic phase. Spherical cells, characteristic of the benthic phase, are united by a common reticulopodium by means of which they capture and digest diatoms (“reticulopodial digestion”). When food gets exhausted, the cellular association (plasmodium) disintegrates into single cells resembling amoebae (“individualization”). After some time, they transform into heliozoan-like cells which float in the seawater. Under unfavorable conditions the spherical cells may encyst. When the diatoms run short and illumination is sufficient, they can survive by means of their plastids. though for a limited time only. Electron microscopical investigations showed that the heliozoan-like cells have not only pseudopodia used as floating projections, but also two flagella. One of the flagella induces an eyespot in a nearby plastid. This situation is characteristic of the so-called heterokont algae (Chrysophyceae, Xanthophyceae). Classification of Reticulosphaera socialis - and a light-microscopically similar species: Chlorarachnion reptans - is discussed.