Characterization of steroid production in cultured human choriocarcinoma cells.

Progesterone is the major steroid synthesized by the JEG-3, BeWo, and JAR cell lines of choriocarcinoma. A lesser amount of pregnenolone is produced. The 17alpha-hydroxy derivatives of these steroids are only minimally present in three lines. The addition of fetal calf serum to the culture medium modestly increases the synthesis of these steroids, but increases the quantity of 17beta-estradiol produced by 30- to 90-fold. The addition of dehydroepiandrosterone, dehydroepiandrosterone sulfate, androstenedione, androstenediol, and testosterone was shown to stimulate 17beta-estradiol synthesis. There is a clear dose-response relationship between the amount of testosterone added and the quantity of 17beta-estradiol produced. These results indicate that 3beta-hydroxysteroid dehydrogenase-isomerase, 17beta-ol dehydrogenase, and aromatase are active in cultured choriocarcinoma cells, whereas 17beta-hydroxylase and 17-20 desmolase do not appear to be functional in these cells. It is concluded that the stereoidogenic capabilities of choriocarcinoma cells in culture are similar to those of the in vivo placenta and support their use as an experimental model of placental steroidogenesis.

Inhibition of pitted morning glory (Ipomoea lacunosa L.) and certain other weed species by phytotoxic components of wheat (Triticum aestivum L.) straw.

This study was conducted to determine if well-known phytotoxic effects of plant residues on crop growth could also be responsible for observed reductions of certain weed species in no-till cropping systems. An aqueous extract of field-grown wheat (Triticum aestivum L.) reduced the germination and root length of pitted morning glory (Ipomoea lacunosa L.) and common ragweed (Ambrosia artemisiifolia L.). Phytotoxicity was increased by about 70% when bioassays with the wheat extract on morning glory and ragweed were conducted in the presence of light. Phytotoxic substances were extracted from wheat with 2 N NaOH. The hydrolyzed extract was fractionated by thin-layer chromatography (TLC). The compound isolated by TLC having the greatest inhibitory effects on morning glory germination was identified using mass spectrometry and determined to be ferulic acid (4-hydroxy-3-methoxycinnamic acid). Ferulic acid at 5 × 10(3) M inhibited the germination and root length of morning glory 23 and 82%, respectively, and prickly sida (Sida spinosa L.) with carpels 85 and 82%, respectively. Crabgrass (Digitaria sanguinalis L.) germination was inhibited 100%. Ferulic acid had no effect on ragweed or prickly sida without carpels. Morning glory root and shoot biomass were reduced 52 and 26%, respectively, when morning glory was grown in sand and watered with a 5 × 10(3) M solution of ferulic acid. Ferulic acid in the presence of prickly sida seed carpels was found to undergo decarboxylation, forming a styrene derivative, 2-methoxy-4-ethenylphenol. The more phytotoxic styrene compound was produced by a bacterium isolated from the carpels of prickly sida seed. The study showed that ferulic acid and other compounds may indeed play a role in reducing the growth of certain weeds in no-tillage cropping systems.

Modification of allelopathic effects ofp-coumaric acid on morning-glory seedling biomass by glucose, methionine, and nitrate.

Studies of allelopathy have emphasized primarily the identification and quantification of phytotoxins in soils, with only limited attention directed toward how organic (carbon) and inorganic constituents (nutrients) in the soil may modify the action of such phytotoxins. In the present study, increasing carbon (C) levels (up to 108µg C/g soil) supplied as glucose, phenylalanine, orp-hydroxybenzoic acid did not alter morning-glory biomass, but similar C levels supplied as leucine, methionine, orp-coumaric acid were inversely related to morning-glory biomass. Similar joint action and multiplicative analyses were used to describe morning-glory biomass response to various C sources and to generate dose isolines for combinations ofp-coumaric acid and methionine at two NO3-N levels and for combinations ofp-coumaric acid and glucose at one NO3-N level. Methionine, glucose, and NO3-N treatments influenced the inhibitory action ofp-coumaric acid on biomass production of morning-glory seedlings. For example, results from the multiplicative analysis indicated that a 10% inhibition of morning-glory biomass required 7.5µgp-coumaric acid/g soil, while the presence of 3.68µg methionine/g soil thep-coumaric acid concentration required for 10% inhibition was only 3.75µg/ g soil. Similar response trends were obtained forp-coumaric acid and glucose. The higher NO3-N (14 vs. 3.5µg/g) treatments lowered the methionine and increased thep-coumaric acid concentrations required for 10% inhibition of morning-glory biomass. These results suggested that allelopathic interactions in soil environments can be a function of interacting neutral substances (e.g., glucose), promoters (e.g., NO3-N), and/or inhibitors (e.g., methionine andp-coumaric acid) of plant growth.

Allelopathic activity in wheat-conventional and wheat-no-till soils: Development of soil extract bioassays.

The primary objective of this research was to determine if soil extracts could be used directly in bioassays for the detection of allelopathic activity. Here we describe: (1) a way to estimate levels of allelopathic compounds in soil; (2) how pH, solute potential, and/or ion content of extracts may modify the action of allelopathic compounds on germination and radicle and hypocotyl length of crimson clover (Trifolium incarnatum L.) and ivyleaved morning glory (Ipomoea hederacea L. Jacquin.); and (3) how biological activity of soil extracts may be determined. A water-autoclave extraction procedure was chosen over the immediate-water and 5-hr EDTA extraction procedures, because the autoclave procedure was effective in extracting solution and reversibly bound ferulic acid as well as phenolic acids from wheat debris. The resulting soil extracts were used directly in germination bioassays. A mixture of phenolic acids similar to that obtained from wheat-no-till soils did not affect germination of clover or morning glory and radicle and hypocotyl length of morning glory. The mixture did, however, reduce radicle and hypocotyl length of clover. Individual phenolic acids also did not inhibit germination, but did reduce radicle and hypocotyl length of both species. 6-MBOA (6-methoxy-2,3-benzoxazolinone), a conversion product of 2-o-glucosyl-7-methoxy-1,4-benzoxazin-3-one, a hydroxamic acid in living wheat plants, inhibited germination and radicle and hypocotyl length of clover and morning glory. 6-MBOA, however, was not detected in wheat debris, stubble, or soil extracts. Total phenolic acids (FC) in extracts were determined with Folin and Ciocalteu's phenol reagent. Levels of FC in wheat-conventionaltill soil extracts were not related to germination or radicle and hypocotyl length of either species. Levels of FC in wheat-no-till soil extracts were also not related to germination of clover or morning glory, but were inversely related to radicle and hypocotyl length of clover and morning glory. FC values, solute potential, and acidity of wheat-no-till soil extracts appeared to be independent (additive) in action on clover radicle and hypocotyl length. Radicle and hypocotyl length of clover was inversely related to increasing FC and solute potential and directly related to decreasing acidity. Biological activity of extracts was determined best from slopes of radicle and hypocotyl length obtained from bioassays of extract dilutions. Thus, data derived from the water-autoclave extraction procedure, FC analysis, and slope analysis for extract activity in conjunction with data on extract pH and solute potential can be used to estimate allelopathic activity of wheat-no-till soils.

Use of water and EDTA extractions to estimate available (free and reversibly bound) phenolic acids in Cecil soils.

Sterile and microbe reinfested Cecil Ap and Bt soil materials amended with 0 to 5 µmol/g of ferulic acid,p-coumaric acid,p-hydroxybenzoic acid, or vanillic acid were extracted after varying time intervals with water, EDTA, or NaOH to characterize sorption of cinnamic and benzoic acid derivatives and to determine the effectiveness of water and EDTA extractions in estimating concentrations of free and reversibly bound phenolic acids in soils. Basic EDTA (0.5 M, pH 8) extractions and water extractions provided good estimates of both free and reversibly bound cinnamic acid derivatives, but not of benzoic acid derivatives. Neutral EDTA (0.25 M, pH 7) and water extractions, however, were effective for both cinnamic and benzoic acid derivatives Rapid initial sorption of both cinnamic and benzoic acid derivatives was followed by slow long-term sorption of the cinnamic acid derivatives. Slow long-term sorption was not observed for the benzoic acid derivatives. The amount of sorption of phenolic acids in soil materials was directly related to the concentration of phenolic acids added to soil materials. The addition of a second phenolic acid to the soil materials did not substantially affect the sorption of each individual phenolic acid. Sodium hydroxide extractions, which were made only after phenolic acids in phenolic acid-amended and non-amended soil material were depleted by microbes, confirmed that neutral EDTA and water extractions of soils can be used to make accurate estimates of baseline (residual) levels of free and reversibly bound phenolic acids available to soil microbes and, thus, potentially to seeds and roots.

Phenolic acid content of soils from wheat-no till, wheat-conventional till, and fallow-conventional till soybean cropping systems.

Soil core (0-2.5 and/or 0-10 cm) samples were taken from wheat no till, wheat-conventional till, and fallow-conventional till soybean cropping systems from July to October of 1989 and extracted with water in an autoclave. The soil extracts were analyzed for seven common phenolic acids (p-coumaric, vanillic,p-hydroxybenzoic, syringic, caffeic, ferulic, and sinapic; in order of importance) by high-performance liquid chromatography. The highest concentration observed was 4 µg/g soil forp-coumaric acid. Folin & Ciocalteu's phenol reagent was used to determine total phenolic acid content. Total phenolic acid content of 0- to 2.5-cm core samples was approximately 34% higher than that of the 0- to 10-cm core samples. Phenolic acid content of 0- to 2.5-cm core samples from wheat-no till systems was significantly higher than those from all other cropping systems. Individual phenolic acids and total phenolic acid content of soils were highly correlated. The last two observations were confirmed by principal component analysis. The concentrations were confirmed by principal component analysis, tions of individual phenolic acids extracted from soil samples were related to soil pH, water content of soil samples, total soil carbon, and total soil nitrogen. Indirect evidence suggested that phenolic acids recovered by the water-autoclave procedure used came primarily from bound forms in the soil samples.