Evidence for Naturally Occurring Inhibitors of Archegonial Differentiation in Lygodium japonicum

Archegonial differentiation in prothallia of Lygodium japonicum was inhibited when the filtrate of conditioned medium or the extracts of prothallia with organic solvents were added to the medium. By varying the timing of treatment with the methanol extract, archegonial differentiation was shown to start at least 4 days before microscopically detectable change. The inhibitory effect of methanol extract was nullified by transferring the treated plants to a fresh medium omitting the methanol extract, so that the archegonial formation became discernible 6 days after the transfer. The inhibitory activity was stable in both acidic and basic solutions at room temperature, and was partially lost by boiling at pH 3 or 11 for 30 min. The inhibitor, which could be retrieved from the filtrate and the methanol extract, was fractionated into the neutral ethyl acetate fraction, but was not found in the acidic ethyl acetate fraction and in the aqueous residue. At least two active zones were separated on thin layer chromatograms of the ethyl acetate extracts from the filtrate and the methanol extract, and the relative flow-rates of each active zone from these two sources were very similar. The evidence described above indicates that specific inhibitors of archegonial differentiation may be produced in the tissue of prothallia of Lygodium and eventually be secreted to the medium.     

The Effect of Stratification on the Endogenous Cytokinin Levels of Seed of Protect compacta and Leucadendron daphnoides

The soybean callus assay was used to study the effect of stratification on the cytokinin levels of the embryo dormant seed of Protea compacta R.Br. and the seed of Leucadendron dapbnoides Meisn., where dormancy is coat imposed. Chilling the seed for 30 days increased germination significantly, and resulted in a simultaneous increase in the butanol soluble cytokinins of both species. It would appear as if these compounds are either synthesized or released from a bound form in embryo dormant seed. In contrast, an interconversion from water soluble to butanol soluble cytokïnins appears to account for the increase where dormancy is coat imposed. The results also indicate that for germination to take place a threshold concentration of cytokinin may be required. It is suggested that the increase in butanol soluble cytokinins may lead to the breaking of dormancy, probably by increasing radicle elongation and/or cotyledon expansion.     

Gibberellin-induced dormancy in Begonia aerial tubers. Increase of the growth inhibitor content by gibberellin treatment

Changes in the amount of growth inhibitors by gibberellin (GA)treatment in tubers of Begonia evaniana was studied. Acidic,neutral and basic ethyl acetate-soluble and n-butanol-solublegrowth inhibitors increased with GA treatment and also withthe progress of natural dormancy. These inhibitors suppressedthe sprouting of nondormant aerial tubers. The increase of neutralethyl acetate-soluble inhibitors in the GA-treated tubers wassuppressed by incubation in low-O₂ atmosphere. In early dormantstage tubers, the inhibitor increased remarkably during incubationat 28?C in light. The aerial tubers probably enter a dormantstate when sufficient amounts of inhibitors accumulate in them. (Received February 9, 1973; )     

小珊瑚藻对赤潮异弯藻的化感效应

研究了不同浓度的小珊瑚藻组织4种不同极性有机溶剂(甲醇、丙酮、乙醚、氯仿)提取物对赤潮异弯藻的生长抑制作用.结果表明：小珊瑚藻组织甲醇提取物对赤潮异弯藻的生长抑制活性最强,并且在较高浓度下能使赤潮异弯藻完全死亡,其他3种有机溶剂提取物对赤潮异弯藻的生长无明显影响.表明小珊瑚藻组织中含有的对赤潮异弯藻有抑制作用的活性物质具有较高的极性.对小珊瑚藻的甲醇提取物进行液液萃取,将其分离为石油醚相、乙酸乙酯相、正丁醇相和蒸馏水相, 并对赤潮异弯藻进行生物活性检测.结果发现石油醚相和乙酸乙酯相具有较强的杀藻活性,表明脂肪酸可能是小珊瑚藻组织内抑制赤潮异弯藻生长的化感物质的重要组成成分之一.     

The role of the cotyledons in the photocontrol of hypocotyl extension in Cucumis sativus L.

Summary When pea seedlings lose about 5% of their water content the abscisic acid ((+)-ABA) level of the shoots increases ca. 20 times and the level of bound ABA, in all probability ABA-glucose, ca.7-10 times. After watering both ABA and bound ABA contents decrease within 24–48 h to the level in the control plants.After application of (±)-[2-¹⁴C] ABA to wilted pea shoots at the time of watering radioactive substances appear in the water-soluble, ether-insoluble fraction of ethanolic extracts and increase with time whereas radioactivity in the acidic ether fraction decreases. The neutral ether fraction remains free of radioactivity. Three radioactive zones, A, B, and C, are seen on chromatograms of the water-soluble fraction. A increases considerably within the entire experimental time, whereas B increases in the first 4–8 h after application and subsequently decreases. The third substance, C, which releases free ABA after hydrolytic treatment, does not change during the experiment. Chromatograms of the acidic ether fraction yield ABA and a substance staying at the origin, possibly phaseic acid and/or dihydrophaseic acid. Only the activity of ABA decreases during the experiment.     

Inhibiteurs non acides et dormance embryonnaire chez Taxus baccata

Non-acidic inhibitors and embryo dormancy in Taxus baccata L. Embryo dormancy of Taxus baccata is eliminated when the embryos are continuously kept in sterile nutritive liquid medium. After 3 weeks of culture, an important non-acidic inhibitory complex can be extracted from this liquid medium. At least three substances are involved: two pigments and a compound with some properties that suggest xanthoxin. These substances are neither found in embryos taken directly from the seeds, nor in liquid medium after 8 days of culture, which is the time necessary and sufficient to allow germination after transfer on agar medium. Such behaviour is quite different from that of ABA previously studied and indicates that these non-acidic inhibitors appear late during the culture and are not directly involved in embryo dormancy.     

Isolation of a factor stimulatory to Bradyrhizobium japonicum in broth culture

Previous experiments indicated that water extracts of lambsquarters (Chenopodium album) and green foxtail (Setaria viridis), among others, stimulated growth of Bradyrhizobium japonicum in broth culture. Water extracts of lambsquarters shoots collected before or after anthesis were equally stimulatory. The stimulatory effect of extracts of lambsquarters when heated to 100°C for 30 min or autoclaved for 15 min was reduced by about 20% compared to untreated extracts. Extracts of green foxtail were less affected by higher temperature under similar conditions. Extracts of both green foxtail and lambsquarters completely lost their stimulatory effect following exposure to aerial microflora for 120 h. Water extract of lambsquarters shoots was more stimulatory than methanol extract, and neither ether nor butanol extracts resulted in stimulation. Both shoots and roots of lambsquarters and green foxtail were sequentially extracted first by water followed by methanol and vice-versa. The bioassay of these extracts indicated that there could be two components of the growth factor-one, larger component is soluble in water, the other, smaller component is soluble in methanol. After fractionation of the crude aqueous extract of lambsquarters shoots by four organic solvents, the residual aqueous extract retained the growth factor. Dialysis of the residual aqueous extract of lambsquarters shoots through a membrane (MWCO 1000) indicated that the molecular weight of the growth factor was less than 1000. The fraction having molecular weight <1000 was separated by paper chromatography using 6% acetic acid as developer. The fraction with Rf 0.91 showed the highest stimulation of the bacterium.     

The Mechanism of Low Temperature Dormancy in Mature Seeds of Syringa Species

The mechanism of seed dormancy at low temperatures (15-9°C) was investigated in the seeds of Syringa josikaea, S. reflexa and S. vulgaris. Low temperature dormancy in Syringa species was mainly imposed by endosperm embedding the radicle. Different degrees of embryo dormancy may occur in S. reflexa seeds. In most cases the low temperature dormancy was broken completely by removing the endosperm around the radicle. The endosperm did not seem to contain significant quantities of germination inhibitors, and the results indicate that it prevents germination mainly due to its mechanical resistance. The mechanical resistance of endosperm did not change during chilling or during induction of dormancy by high temperature incubation. The strength of the endosperm decreased rapidly in non-dormant seeds before visible germination. Similar changes were not observed in dormant seeds. Generally, the strength of the endosperm was lower in the non- (or less) dormant species S. josikaea and S. vulgaris than in the more dormant S. reflexa seeds. The growth potential of the embryos, measured as their ability to germinate in osmotic solutions (mannitol or polyethylen glycol 4000), was increased by chilling and by GA₃-treatment. The growth potential of untreated S. josikaea and S. vulgaris embryos was generally higher than that of S. reflexa embryos. Acid ethyl acetate fractions of methanol extracts from embryos of all three species contained substances with GA³-like activity in the lettuce hypocotyl test. The activity was found at Rf 0.9–1.0 on paper chromatograms run in distilled water. No significant changes in the activity were detected during chilling or prior to visible germination.     

GROWTH-INHIBITORS AND GROWTH-PROMOTERS IN ENTEROMORPHA COMPRESS A (CHLOROPHYTA)1

Methanol extracts from the alga Enteromorpha compressa (L.) Grev. contain substances which inhibit the elongation of Lepidium roots. Chromatographic separation of the inhibiting substances revealed that one of the inhibitory zones of the chromatograms had properties of the so-called inhibitor β. Neither abscisic acid (ABA) nor lunularic acid proved to be responsible for the growth-inhibiting property of this zone. Moreover, the extracts contain substances which promote the elongation of Avena coleoptile segments. One of these substances could be tentatively identified as indole-3-acetic acid by thin-layer and gas-liquid chromatography. (In addition to indole-3-acetic acid a second growth-promoting factor with the properties of the so-called accelerator α could be detected.)     

Antioxidant Activities of Some Extracts of Thymus zygis

The antioxidant activities of methanol and ethyl ether extracts obtained from Thymus zygis, collected during the flowering or non-flowering period, were evaluated and compared. To investigate this potential, extracts were tested on their capacity to react with diphenyl-picrylhydrazyl (DPPH) in a homogeneous medium, and to inhibit Fe²⁺/ascorbate-induced membrane lipid peroxidation, as estimated by the formation of thiobar-bituric acid-reactive substances (TBARS). Although methanol extracts reduce DPPH radicals more efficiently than ethyl ether extracts, suggesting a potent radical scavenger activity, the ethyl ether extracts were found to be most active in inhibiting lipid peroxidation in sarcoplasmic reticulum (SR) membranes. In addition, both extracts present peroxyl and superoxide radical scavenging activities. Peroxyl radicals were generated by the water soluble 2, 2A-azobis(2-amidinopropane) dihydrochloride (AAPH) azoinitiator, and the scavenging activities of the extracts were measured by the inhibition of cis-parinaric acid (PnA) fluorescence decay in SR. Superoxide radicals were generated either by an enzymatic or a non-enzymatic system, and the scavenger ability was evaluated by the inhibition of nitrob-lue tetrazolium reduction. Methanolic extracts are more potent as scavengers of peroxyl and super oxide radicals than the ethyl ether extracts. Apparently, there is a relationship between antioxidant potency and the total phenolic groups content in each extract.     

Alterations in Endogenous Levels of Gibberellin-like Substances during Germination of Phaseolus vulgaris Seeds

The endogenous gibberellin-like substances were determined in mature dry and germinating bean seeds, Phaseolus vulgaris L. cv. Alabaster. Methanol extracts were partitioned against ethyl acetate and butanol at neutral and acid pH. Each phase was individually chromatographed on a silica gel column. The gibberellin activity was measured with the Tan-ginbozu dwarf rice micro-drop bioassay. Each extract was tested in two dilutions. Extracts from dry seeds showed the highest gibberellin activity, largely attributable to ethyl acetate-soluble substances. The activity was considerably reduced in extracts from seeds imbibed for 1 day. Gibberellin-like substances soluble in butanol appeared in extracts from seeds soaked for 1 or more days.     

Growth Regulators in Picea abies

The occurrence of growth regulators active in the Avena coleoptile straight-growth test in sprouting buds and seedlings of Norway spruce (Picea abies Karst.) was investigated. The acid ether fraction contained a growth stimulator, the Rf of which in isopropanol: ammonia: water was 0.2–0.4. This substance behaved as indole-3-acetic acid (IAA) in elec-trophoresis, in chromatography in various solvent systems on paper and on a Sephadex column. It gave the colour typical of IAA when sprayed with Ehrlich reagent and its fluorescence characteristics corresponded to IAA. Acid ether-soluble inhibitors showed most activity at Rf 0.4–0.7, but due to tailing they interfered with the determination of the stimulator at the Rf of IAA in the bioassay. They also masked the activity of other stimulators. Colour reactions were obtained with Ehrlich reagent in the inhibiting chromatogram zone. When eluates from this zone were tested in high dilutions or after gel filtration growth stimulation was obtained. The acid fraction of seedling shoots also contained a stimulator with Rf 0.7–0.8. In the neutral-basic ether-soluble fraction growth stimulation was obtained at Rf 0.5–0.7. The extracts also contained stimulatory substances insoluble in ether but soluble in n-butanol and partly in ethyl acetate. When the butanol fraction was hydrolyzed in 1 M NaOH a substance behaving as IAA when chromatographed was released.     

Effects of Oxygen and Respiratory Inhibitors on Induction and Release of Dormancy in Aerial Tubers of Begonia evansiana

Effects of O₂ and some respiratory inhibitors on the induction and release of bud dormancy were examined with the aerial tubers of different ages of Begonia evansiana Andr. Oxygen was needed not only for tuber sprouting but also during the chilling process at 2 to 5 C to break tuber dormancy. If the mature tubers were exposed to blue light during the chilling period, their dormancy was strikingly released even by the chilling given under an O₂ concentration as low as 3%. Blue light pretreatment promoted photo-sprouting of immature tubers only when given under lower O₂ concentrations. On the other hand, red light became effective in inducing dormancy in the immature tubers and in prolonging dormancy in the mature tubers as O₂ tension was increased. This was also the case with the induction of dormancy in the immature tubers by exposing them to a lower temperature (17 C) in the dark. The development of dormancy was suppressed by 2,4-dinitrophenol, p-nitrophenol, and sodium azide.     

Screening of terrestrial and freshwater halotolerant cyanobacteria for antifungal activities

Cyanobacterial cultures tolerating 200 mmol l⁻¹ sodium chloride isolated from terrestrial and freshwater habitats of North Maharashtra region of India were evaluated for antifungal activity. Aqueous, methanol, n-propanol, and petroleum ether extracts of 40 cyanobacterial isolates belonging to nine genera were examined for inhibitory activity against five fungal pathogens. Eighteen isolates belonging to genus Oscillatoria dominated the population of halotolerant cyanobacterial cultures. Four antifungal bioassays viz. double layer agar method, disc diffusion assay, silica gel method, and minimum inhibitory concentration (MIC) were used to screen the cultures for antifungal activity. Among the solvents used, methanol extracts showed 34.9% inhibition followed by n-propanol, petroleum ether, and water exhibiting 30.2%, 18.6% and 16.2% inhibition, respectively. The double agar layer method was found to be a suitable method in preliminary screening for handling large number of cultures without extraction of compounds. However, in later screening experiments, silica gel method was seen to be advantageous over MIC and agar disc diffusion methods.     

Changes in the levels of free IAA and cytokinins in potato tubers during dormancy and sprouting

Changes in the levels of free indol-3-ylacetic acid (IAA) and free cytokinins were determined in the course of dormancy and sprouting period in potato tubers(Solanum tuberosum L., cv. Nevskii) stored at 4 °C. The same analyses were performed in potato tubers after Ethrel application, which prolongs dormancy. No significant changes were found in free IAA level during dormancy followed by a rapid decrease during sprouting. After Ethrel application a significant lower IAA level was found 3 weeks after application, but further on no changes in free IAA level between treated and non-treated tubers were detected. Cytokinin level was relatively low and constant till sprouting and increases then by about 55 %, mainly due to an increase in the level of zeatin riboside and isopentenyladenosine. Ethrel application decreased cytokinin level during dormancy slightly, but postpones the increase coupled with sprouting by about 1 month. Thus, IAA does not seem to have a significant effect on tuber dormancy, while cytokinins are probably necessary for sprouting initiation.     

Multiple allelopathic activity of the crustose coralline algaLithophyllum yessoenseagainst settlement and germination of seaweed spores

A study was made to investigate possible formation by the crustose coralline algaLithophyllum yessoenseof multiple allelopathic-related substances against the settlement and germination of spores of various seaweeds. Seven different solvents (n-hexane, diethyl ether, acetone, ethyl acetate, acetonitrile, methanol, distilled water) and seawater were used to obtain crude extracts and secretory exudates from the coralline alga. The extracts and the algal conditioned seawater were tested for inhibitory activity against the settlement and germination of spores from 17 species representing 15 genera. Spore settlement of 14 species was inhibited over 90% by one or more extracts of the six organic solvents and conditioned seawater. The germination of spores from 13 species was inhibited by one or more extracts of all seven solvents and conditioned seawater. The species where spore settlement was not significantly affected showed strong inhibition of germination, andvice versa.     

Ein Beitrag zur Analyse der Gibberelline in Samen von Pisum sativum L.

Summary Unripe seeds of Pisum sativum were analyses for gibberellins and gibberellin-like substances. Forty kg of seeds were extracted with methanol, the methanol was evaporated and the residue distributed between water and ethylacetate, and between water and butanol at different pH-values. The acidic ethylacetate and the butanol fractions were separated by thin layer chromatography. In the acidic ethylacetate fraction a new substance tentatively named gibberellin A_x was isolated by preparative thin layer chromatography. In different solvent mixtures this substance migrates closely behind gibberellin A₃. It is biological active in a dwarf pea and in dwarf maize mutant bioassays. IR and fluorescense spectra point to a gibban structure. Furthermore gibberellin A₅ was found to be the main gibberellin in pea seeds. It was identified by its R_f-values and the R_f-values of its methylester in different solvent systems as well as by its characteristic relative biological activity in three biological test systems. Most probably gibberellin A₆ is also present in the pea seeds, but because the amounts detected were small, no clear evidence can be presented.In the butanol phase two gibberellin-like substances could be separated by thin layer chromatography.

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Mit Hilfe der Deutschen Forschungsgemeinschaft.     

Chloroindolyl-3-acetic Acid and Its Methyl Ester Incorporation of 36Cl in Immature Seeds of Pea and Barley

Immature seeds of pea and barley were harvested on plants grown in solutions containing ³⁶Cl^?, but no other chlorides. Autoradiography of two-dimensional thin layer chromatograms (silicagel) of butanol extracts of freeze-dried seeds showed the presence in both species of several radioactive compounds besides Cl^?. One compound, present in pea and probably in barley, cochromatographed with a mixture of 4- and 6-chloroindolyl-3-acetic acid methyl esters. Another, detected in pea, but probably not in barley, cochromatographed with a mixture of 4-and 6-chloroindolyl-3-acetic acids.     

Antioxidant Activities of Some Extracts of Thymus zygis

The antioxidant activities of methanol and ethyl ether extracts obtained from Thymus zygis, collected during the flowering or non-flowering period, were evaluated and compared. To investigate this potential, extracts were tested on their capacity to react with diphenyl-picrylhydrazyl (DPPH) in a homogeneous medium, and to inhibit Fe²⁺/ascorbate-induced membrane lipid peroxidation, as estimated by the formation of thiobar-bituric acid-reactive substances (TBARS). Although methanol extracts reduce DPPH radicals more efficiently than ethyl ether extracts, suggesting a potent radical scavenger activity, the ethyl ether extracts were found to be most active in inhibiting lipid peroxidation in sarcoplasmic reticulum (SR) membranes. In addition, both extracts present peroxyl and superoxide radical scavenging activities. Peroxyl radicals were generated by the water soluble 2, 2A-azobis(2-amidinopropane) dihydrochloride (AAPH) azoinitiator, and the scavenging activities of the extracts were measured by the inhibition of cis-parinaric acid (PnA) fluorescence decay in SR. Superoxide radicals were generated either by an enzymatic or a non-enzymatic system, and the scavenger ability was evaluated by the inhibition of nitrob-lue tetrazolium reduction. Methanolic extracts are more potent as scavengers of peroxyl and super oxide radicals than the ethyl ether extracts. Apparently, there is a relationship between antioxidant potency and the total phenolic groups content in each extract.     

栎属7种植物种子的发芽抑制物质研究

运用系统溶剂法和生物测定法,以栎属7种植物种子为材料,研究了其种皮、胚(胚及周围部分子叶)和子叶(远离胚端2/3子叶)1.0 g/mL和0.5 g/mL浓度甲醇等浸提液以及各有机相对白菜种子萌发的影响。结果表明:栓皮栎、锐齿栎、蒙古栎、沼生栎和麻栎甲醇浸提液均能显著降低白菜种子发芽率、根长和苗高,且对发芽率的抑制作用逐渐降低;夏栎和房山栎甲醇浸提液对白菜种子发芽率没有显著影响,但对根长和苗高有一定的抑制作用;种子不同部位甲醇浸提液的抑制作用表现为胚>子叶>种皮,且甲醇高浓度浸提液的抑制作用大于低浓度的抑制作用;甲醇相对白菜种子发芽率、苗高或根长的抑制作用最强,其次是乙酸乙酯相,其它溶剂相萃取液的抑制作用不明显。栎属种子甲醇浸提液及各有机相对白菜种子苗高和根长的抑制作用强于对发芽率的影响,说明栎属种子中所含抑制物质主要是限制自身根和芽的生长,可能是造成延迟萌发和出苗不整齐的原因。