Band 48, Heft 1-2

By a comparative thin layer chromatographic screening of the methanol-soluble leaf exudates from more than 400 Aloe plants (183 species), 5-hydroxyaloin A was identified in 20 species. Whilst 13 of the 20 species revealed interindividual variations concerning to the occurrence of 5-hydroxyaloin A, this anthrone-C-glucosyl was unambiguously detected in each individual of 6 Aloe species. In the leaf exudates from A. marlothii Berger 5-hydroxyaloin A was only traceable in the aloin-containing chemivars. The complete anthrone-C-glucosyl pattern of these 7 clearly characterized species has been determined additionally by qualitative and quantitative high performance liquid chromatography: The results obtained demonstrate that 5-hydroxyaloin only occurs in the more stable A-configuration (10 R, 1′S), thus being till now the only anthrone-C-glycosyl which has not been found as diastereomeric pair genuinely in plants. As well, 5-hydroxyaloin A characterizes a quantitatively significant hydroxylating pathway in biosynthesis of anthranoids. It is discussed as a chemotaxonomic marker of the genus Aloe, especially of the sections Pachydendron and Eualoe.

The epicuticular wax of Euphorbia helioscopia was fractionated into fatty acids, hydrocarbons, wax esters, aldehydes, methyl esters, triterpenol acetates, alcohols, sterols, and polar components. The composition of the fractions was determined by GC, GC-MS, HPLC. Main components within these lipid classes are hentriacontane, wax esters C 46 and C 48 , octacosanal, hexacosanol and octacosanol, hexadecanoic acid, and β-sitosterol. Lupeol and its acetate were also confirmed.

The results of tracer experiments performed with anthers of Tulipa cv. Apeldoorn have already shown that a high level of incorporation into the sporopollenin fraction was achieved, when [ 14 C]phenylalanine was applicated as a precursor. In order to investigate whether the substantial incorporation of [ 14 C]phenylalanine is a unique phenomenon restricted to Tulipa, tracer experiments were carried out on anthers of Cucurbita maxima. The sporopollenin fraction was isolated and purified by a gentle method including extractions with various solvents, incubations with hydrolysing enzymes and fractionated saponifications. The remaining, as well as the released radioactivity, was determined after each purification step. After the application of [U- 14 C]phenylalanine, a substantial incorporation into the sporopollenin fraction was determined. The values were clearly higher than those obtained with [1- 14 C]glucose or those from corresponding experiments on Tulipa anthers. After potash fusion of sporopollenin fractions labeled via [U- 14 C]phenylalanine, p-hydroxybenzoic acid was shown to be the main com ponent among the ether soluble acids; moreover it showed the highest level of radioactivity. No radioactivity was detected in the degradation products oxalic acid, benzoic acid, phthalic acid and terephthalic acid or octanedioic acid- and decanedioic acid-dimethylester.

7-O-Methyl-luteone was metabolized in a buffer solution by resting cells of Botrytis cinerea which is known as a fungus detoxifying prenylated isoflavones into further oxygenated metabolites, and we could isolate an unstable epoxy-intermediate. The epoxy-intermediate was easily cyclized under acidic conditions to yield the corresponding isoflavone with a hydroxychroman part structure (= dihydropyrano-isoflavone). The optically active (dextrorotatory) dihydropyrano-isoflavone metabolite was derivatized into the [R]-a-methoxy-a-trifluoromethyl- phenylacetic acid ester and its absolute configuration [S] was unambiguously elucidated by the modified Mosher’s method.

In crude extracts of Chlorella kessleri two forms of fructosebisphosphatase can be separated by ion exchange chromatography or by acid precipitation. FBPase I is eluted from DEAE cellulose at 200mᴍ KC1 and precipitated at pH 4.5, FBPase II is correspondingly eluted at 310 mᴍ KC1 and soluble at pH 4.5. Both enzymes differ in substrate affinity and degree of cooperativity. Based on literature data, FBPase I is assumed to be of cytosolic and FBPase II of chloroplastic origin. The mole mass of FBPase I is identical (51-65 kDa) at pH 6.5 and pH 8.5. T hat of FBPase II is however, about four times larger at pH 6.5 (257 kDa) than at pH 8.5 (67 kDa). Other pH values have only been tried with crude cell extracts in which still larger mole masses of FBPase resulted at more acidic pH (1349 kD a at pH 6.0). The lower mole mass form of FBPase II shows three times higher catalytic activity. Reducing agents, such as DTT, also increase the activity of FBPase II in vitro. In vivo, alkalization and production of reducing power occurs in the chloroplast stroma during illumination. If the above alterations exist in vivo, they would be a means to activate FBPase in the light. Oligomerization of FBPase II to aggregates with altered catalytic activities and kinetic properties is discussed as result of the action of specific wavelengths and to be responsible for differences in carbohydrate metabolism of Chlorella exposed to red or blue light.

Hemidiscoidal and hemiellipsoidal phycobilisomes have been determined in cells of the complementary chromatically adapting cyanobacterium Phormidium sp. C86 . They could be isolated from red and green light-adapted cells, respectively. Hemidiscoidal red light phycobilisomes show molar pigment ratios of allophycocyanin: phycocyanin of 1:4.5 with phycoerythrin lacking. Hemiellipsoidal phycobilisomes induced by green light present allophycocyanin: phycocyanin: phycoerythrin ratios of 1:1:6.8. The differences between the two phycobilisome types could additionally be demonstrated by their ultrastructure and sedimentation values. Isolated red light phycobilisomes have six rods, show dimensions of 70×30×15nm and a sedimentation value of 66 S whereas green light phycobilisomes are nearly twice larger. They contain ten rods and present dimensions of 70×40×25nm and a sedimentation value of 98 S. The number of phycobilisomes in red light cells is almost twice as large as in green light cells. There is evidence that cells grown under white light contain both types as well as “intermediate” forms.

Binding of tritiated indole-3-acetic acid ([ 3 H]IAA) to symbiosome membranes of soybean nodules occurred in a protein-dependent manner and was competitively inhibited by unlabeled indole-3-acetic acid (IAA), 1-naphthaleneacetic acid (1-NAA) and dithiothreitol (DTT), but not by tryptophan and benzoic acid. The symbiosome membranes bound IAA with a K D of 1 × 10 -6 m. Photoaffinity labeling identified an auxin-binding protein (ABP) in the symbiosome membrane with an apparent molecular mass of 23 kDa. This 23 kDa protein was labeled either with 5-azido-[7- 3 H]indole-3-acetic acid ([ 3 H]N 3 IAA) or with 5′-azido-[3,6- 3 H 2 ]-1-naphthylphthalamic acid ([ 3 H 2 ]N 3 NPA). Labeling of the 23 kDa protein with [ 3 H]N 3 IAA was competitively inhibited by unlabeled IAA and 1-NAA. NPA and quercetin, inhibitors of polar auxin transport, as well as rutin, a glycosylated derivative of quercetin, competed with IAA for binding. Conversely, [ 3 H 2 ]N 3 NPA labeling was inhibited by unlabeled IAA and NPA. The 23 kDa symbiosome membrane protein was partially solubilized with Triton X-100 and nearly completely using Triton X-114. The observation that auxin transport inhibitors compete with IAA for binding suggests that the symbiosome membrane ABP could be part of an auxin efflux carrier system required to control the auxin concentration in infected soybean nodule cells.

The hydrogenases of the green algae Scenedesmus obliquus and Chlamydomonas reinhardtii were activated under anaerobic conditions. Exposure of whole cells and cell-free homogenates to air lead to a complete inactivation of the hydrogenases. The inactivation in whole cells of Scenedesmus is faster than the inactivation of the cell-free homogenate. Inactivation of the hydrogenases could be reversed by anaerobic readaptation in whole cells. The inactivation of the hydrogenase in homogenates seems to be irreversible. Neither the removal of oxygen nor the addition of ATP, NAD(P)H, sodium dithionite, dithiothreitol, ferredoxin and thioredoxin to homogenates facilitated the reactivation of the hydrogenase. The occurrence of a hydrogenase regulating factor is discussed.

Excitation spectra of chlorophyll fluorescence from intact rye leaves were registered at different steps of the induction of photosynthesis after dark adaptation. Analysis of these spectra indicates that at least two processes related to spectroscopic features are responsible for a fluorescence quenching. The first one, active during the first 100 s of illumination, was interpreted to consists in an overall decrease of the fluorescence quantum yield of antenna pigments and chlorophylls, in particular close to the reaction centers. The second type of a fluorescence decrease (between 100 s and 300 s of illumination) was found to be in large extent related to decrease of the rate of an excitation energy transfer between accessory xanthophyll pigments and chlorophylls emitting fluorescence. This latter molecular mechanism is discussed as being related to violaxanthin availability to de-epoxidation in the xanthophyll cycle.

Several benzoic, cinnamic and phenylacetic acid derivatives were screened with 20 microorganisms, mainly fungi, for the reduction of their carboxylic function. For all organisms several compounds were reduced in fairly good yields up to 80% to the corresponding alcohol. No general rule could be established, concerning the substitution pattern, as to which compounds were transformed to the alcohol. Generally the reactions were accomplished within 48-70h. Only minor, if any, side products were detected. Dicarboxylic acids, such as phthalic or phenylglutaric acids and similar compounds could not be reduced by the microorganisms tested.

The sulfur amino acid homocysteine has recently been addressed as marker for vessel damaging and atherosclerotic dispositions. The atherogenic index has been correlated with the one of cholesterol and is significantly higher in cholesterinemic as compared to normal lipidemic persons. In the present communication biochemical model reactions are presented indicating the prooxidative activity of homocysteine where a cooperative effect with the transition-metals copper and iron is indicated.

The miscoding potential of N 2 ,3-ethenoguanine (∈G), one of the carcinogen vinyl chloride adducts to DNA bases, has been examined by copying of poly (A, ∈G ) templates with DNA-dependent RNA polymerase and reverse transcriptase. In contrast to the results previously obtained with poly (C, ∈G) templates where ∈G acts as G and A, in poly (A, ∈G) templates ∈G acts alm ost exclusively as A. These results suggest that mutagenic potential of ∈G in vivo can depend on the nature of neighboring bases.

The flavonoids present in beeswax produced in “La Alcarria” region were analyzed by HPLC. Pinocembrin, pinobanksin, pinobanksin 3-acetate, chrysin, galangin and techtochrysin were detected as the main flavonoid constituents. This is the first detailed report on the flavonoids of beeswax. These substances are already present when wax scales are secreted by bees. The same flavonoid compounds were generally present in honey, propolis and Populus nigra bud exudates collected in the same geographical region. These results indicate that beeswax flavonoids originate from those of honey and/or propolis, and suggest that analysis of beeswax flavonoids could be used as an adjunct in the detection of beeswax adulterations.

Isomeric compounds reportedly released by mature males of Nezara viridula as attractants for conspecific females are evidently artifacts formed by dimerization of (E)-4-oxo-2-hexenal. Additional analyses of the male-specific volatiles from Italian, Australian, Brazilian, and Japanese populations of N. viridula verify that at least two distinctive pheromone strains exist, but an active synthetic pheromone has not yet been developed. Analyses of volatiles from N. antennata and Acrosternum aseadum males are also reported showing that the native Japanese Nezara sp., and species in the sister genus Acrosternum, produce species-specific blends based on the same compounds as N. viridula: (Z)-α-bisabolene (1-methyl-4-(1,5-dim ethyl-(Z)-1,4- hexadienyl)-cyclohexene), and trans- and cis-1,2-epoxides of (Z)-α-bisabolene. The trans-/cis- 1,2-epoxide ratio of N. antennata is within the range found for most N. viridula populations, but the blend from Japanese N. viridula males deviates radically from those of other conspecific populations.

Cast skins of various snakes and in addition one lizard were found to contain 3.5-8.6% extractable lipid material. Lipids obtained from a cast skin of the Indian python were analyzed in detail indicating the presence of hydrocarbons (squalene, cholestadiene and alkanes), monoester waxes, sterol esters, diester waxes, triglycerides, sterols, free fatty acids and even more polar lipids. Among the monoester wax constituents odd-numbered secondary alkanols were found. It is assumed that the above lipids originate from cells of the past integumental generation and that they play an important facilitating role during the sloughing process.

Glycogen synthesis in isolated perfused livers or livers of anesthetized rats (in situ), was studied using radioactively labelled fructose, lactate, and inositol as substrates. The specific radioactivity of glucose and glycogen was measured at various times and compared with that of some intermediates. The results suggest that liver glycogen is formed from the pool of free glucose which in turn is fed by the so-called “direct and indirect pathway” of glycogen synthesis. This points to an important role of glucose-6-phosphatase, an enzyme complex subject to regulation by glucocorticoids, well known promoters of hepatic glycogen synthesis.

The elementary responses, the “bumps”, of the ventral photoreceptor of Limulus polyphemus were measured under voltage clamp conditions. We observed a type of bump which differs in size and time parameters from the “standard” bumps described previously. The average value of the amplitude of these “small, slow” bumps is about 20-fold smaller than that of the standard bumps (about 1 nA) and the average latency is about 25% longer. The duration of these bumps is 2 -5 -fold longer than that of a standard bump with the same amplitude. The small, slow bumps are probably light-induced and arise with about the same frequency after a light flash as standard bumps. The different kinetics of the two bump types can be explained by the assumption that the transduction chain branches into at least two pathways.

Wavelength discrimination in the flower visiting blowfly Lucilia spec, was investigated in an attempt to elucidate the mechanisms underlying colour vision in this insect. The flies were subjected to a classical conditioning procedure in which they had to discriminate between a rewarded and an unrewarded monochromatic light stimulus. The results reveal large wavelength ranges within which no discrimination occurs, between which, however, a very distinct discrimination is found. The first range consists of the UV region up to 400 nm (UV). The second range comprises wavelengths between 400 nm and 515 nm (BLUE) and the third range all wavelengths longer than 515 nm (YELLOW). A simple model consisting of two colour opponent subsystems (R7p /R8p and R7y/R8y) can explain these results. Each of the two subsystems is assumed to evaluate only whether the sign of the difference between the excitations of R7 and R8 is positive or negative. For the whole system there are thus four possible conditions: p + y + , p + y - , p - y + , p - y - . Three of them correspond to the experimentally obtained wavelength ranges. The fourth condition (p + y - ) might represent a still hypothetical PURPLE category in which the stimulus is made up of both short and long wavelengths.

Hydrolyzed alcoholic extracts of Viscum album L. were analyzed by HPLC for their flavonoid content. Besides the known flavonoid aglyca 4′,5,7-trihydroxy-3′- methoxyflavanone (homoeriodictyol), 5,7-dimethoxy-4′- hydroxyflavanone, 3′-methylquercetin (isorhamnetin) and 3′,7-dimethylquercetin (rhamnazin) another flavanone derivate, 4′,5-dihydroxy-7-methoxyflavanone (sakuranetin) was detected in mistletoe extracts for the first time. With the described method it is possible to distinguish the three mistletoe subspecies abietis, album and austriacum.

The venom gland of the braconid parasitoid, Apanteles kariyai, was morphologically observed with photomicroscope and aromatic amino acids in its venom fluid were chemically analyzed with a three-dimensional HPLC svstem-coulometric ECD. Tyrosine (TYR-4) → tyramine (TYRA) → hydroxyphenylacetic acid (HPAC-4) and tryptophan (TRP) → kynurenine (KYN) were the detected metabolic pathways. This report of venom components of a braconid wasp outlines qualitative differences between this and venom from social wasps.

By means of GC, GC-MS and GC-combined EAG recordings (9Z,12E)-9,12-tetradecadien-1-ol (1) and (9Z,12E)-9,12-tetradecadienyl acetate (2) in a ratio of 4:1 were identified as the pheromone components of the female pyralid moth Euzophera punicaella M. originating from Armenia. Determination of EAG activity with male moth antennae and synthetic test chemicals revealed best responses with a mixture of both compounds.

Infection of Chlorella-like green algae with freshwater phycoviruses is associated with a large and rapid demand for DNA precursors which cannot be met by the algal deoxyribonucleotide-synthesizing enzymes. We have demonstrated in these cells an up to ten-fold increase of the key enzyme, ribonucleotide reductase, 1-2 h post infection. The enzyme activity has been partially enriched from cell extracts. In vitro, it differs from that of uninfected algae in three characteristic parameters, viz. eight-fold higher resistance to millimolar hydroxyurea concentrations, much higher optimum concentration of an allosteric effector nucleotide, thymidine triphosphate, and an unusually low temperature optimum at 20 °C. We conclude that the large DNA phycoviruses, like Herpes and pox viruses, code for their own specific ribonucleotide reductase.