Udo Blum

Microbial populations and phenolic acids in soil

Abstract Populations of bacteria, fungi and actinomycetes in Portsmouth A 1 - and B,-soil material were affected in different ways by repeated enrichment with ferulic, p -coumaric, p -hydroxybenzoic or vanillic acids. Responses varied with type of soil material and phenolic acid, phenolic acid concentration, and inorganic nutrient status of the soil. Populations changed more frequently in B 1 - than in A 1 -soil material. Phenolic acids were readily metabolized by microorganisms, sometimes without detectable population changes, when adequate mineral nutrients were present. Induction of enzymes or selection of organisms capable of degrading individual phenolic acids were clearly evident. Results imply that microbial activity in bioassay systems should be defined for allelopathic studies, particularly when results from various bioassay systems are to be compared

Effects of microbial utilization of phenolic acids and their phenolic acid breakdown products on allelopathic interactions

Reversible sorption of phenolic acids by soils may provide some protection to phenolic acids from microbial degradation. In the absence of microbes, reversible sorption 35 days after addition of 0.5–3 μmol/g of ferulic acid or p-coumaric acid was 8–14% in Cecil Ap horizon and 31–38% in Cecil Bt, horizon soil materials. The reversibly sorbed/solution ratios (r/s) for ferulic acid or p-coumaric acid ranged from 0.12 to 0.25 in Ap and 0.65 to 0.85 in Bt horizon soil materials. When microbes were introduced, the r/s ratio for both the Ap and Bt horizon soil materials increased over time up to 5 and 2, respectively, thereby indicating a more rapid utilization of solution phenolic acids over reversibly sorbed phenolic acids. The increase in r/s ratio and the overall microbial utilization of ferulic acid and/or p-coumaric acid were much more rapid in Ap than in Bt horizon soil materials. Reversible sorption, however, provided protection of phenolic acids from microbial utilization for only very short periods of time. Differential soil fixation, microbial production of benzoic acids (e.g., vanillic acid and p-hydroxybenzoic acid) from cinnamic acids (e.g., ferulic acid and p-coumaric acid, respectively), and the subsequent differential utilization of cinnamic and benzoic acids by soil microbes indicated that these processes can substantially influence the magnitude and duration of the phytoxicity of individual phenolic acids.

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 & Ciocalteus 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.

Induction and/or Selection of Phenolic Acid-Utilizing Bulk-Soil and Rhizosphere Bacteria and Their Influence on Phenolic Acid Phytotoxicity

Bulk-soil and rhizosphere bacteria are thought to exert considerable influence over the types and concentrations of phytotoxins, including phenolic acids, that reach a root surface. Induction and/or selection of phenolic acid-utilizing (PAU) bacteria within the bulk-soil and rhizosphere have been observed when soils are enriched with individual phenolic acids at concentrations ≥0.25 μmol/g soil. However, since field soils frequently contain individual phenolic acids at concentrations well below 0.1 μmol/g soil, the actual importance of such induction and/or selection remains uncertain. Common bacteriological techniques (e.g., isolation on selective media, and plate dilution frequency technique) were used to demonstrate in Cecil Ap soil systems: (1) that PAU bacterial communities in the bulk soil and the rhizosphere of cucumber seedlings were induced and/or selected by mixtures composed of individual phenolic acids at concentrations well below 0.25 μmol/g soil; (2) that readily available carbon sources other than phenolic acids, such as glucose, did not modify induction and/or selection of PAU bacteria; (3) that the resulting bacterial communities readily utilize mixtures of phenolic acids as a carbon source; and (4) that depending on conditions (e.g., initial PAU bacterial populations, and phenolic acid concentration) there were significant inverse relationships between PAU bacteria in the rhizosphere of cucumber seedlings and absolute rates of leaf expansion and/or shoot biomass. The decline in seedling growth could not be attributed to resource competition (e.g., nitrogen) between the seedlings and the PAU bacteria in these studies. The induced and/or selected rhizosphere PAU bacteria, however, reduced the magnitude of growth inhibition by phenolic acid mixtures. For a 0.6 μmol/g soil equimolar phenolic acid mixture composed of p-coumaric acid, ferulic acid, p-hydroxybenzoic acid, and vanillic acid, modeling indicated that an increase of 500% in rhizosphere PAU bacteria would lead to an approximate 5% decrease (e.g., 20–25%) in inhibition of absolute rates of leaf expansion. As far as we know, this is the first time that such a relationship has been quantified.

Allelopathic substances in ecosystems : Effectiveness of sterile soil components in altering recovery of ferulic acid.

Recovery studies were conducted with ferulic acid, a common allelopathic agent, using various soils and soil components. Ferulic acid was placed into sterile soil components (gibbsite, geothite, Georgia kaolin, and Utah bentonite), and different sterile soil materials (from different horizons in the same profile) varying in mineralogy and in organic matter content. The initial concentration of ferulic acid added to the soil materials was 1000 μg/g (5.149 mmol/g). The pH of the soil materials was adjusted and maintained at approximately 4.5 or 7.5. Samples were extracted with 0.03 M EDTA at days, 1, 4, 7, 10, and 13 after addition of ferulic acid. Concentrations of ferulic acid in the extracts were determined with a high performance liquid chromatograph. No breakdown products were detected. Models were developed to describe the recovery of ferulic acid from each soil material and soil component over time. Organic matter was the most active soil component involved in the irreversible retention of ferulic acid. The inorganic soil components were much less active than organic matter but appeared to be similar to each other in activity. Irreversible retention of ferulic acid by soil and soil components was greatest as pH 7.5.

A study of the potential ways in which ozone could reduce root growth and nodulation of soybean

Abstract The possible mechanisms by which ozone reduces root growth and nodulation of soybean were investigated. Ozone did not appreciably penetrate the plant growth substrates nor did it oxidize soil organic matter to form compounds inhibitory to Rhizobium . When ozone was excluded from the plant foliage, but not from the soil, root growth and nodulation were not reduced. However, when plant tops were directly exposed to ozone, root growth and nodulation were reduced. These results indicated that observed reductions in root growth and nodulation did not occur by way of the soil, but resulted from an effect of ozone on the plant foliage.

Effects of ferulic acid, an allelopathic compound, on net P, K, and water uptake by cucumber seedlings in a split-root system.

Since distribution of allelopathic compounds in soils is highly variable, injurious effects by such compounds should be related to the frequency of contact with roots. Experiments were conducted to determine how P, K, and water uptake of cucumber seedlings were affected as the fraction of roots in contact with ferulic acid (FA) was increased. Seedlings were grown in Hoaglands nutrient solution for 14 days and then transferred to 0.5 mM CaSO4 solution for 24 hr before being placed into a split-root culture system. The containers in the system were filled with 0.5 mM concentrations of KH2PO4 and CaSO4 or 0.5 mM concentrations of KH2PO4, CaSO4, and ferulic acid (FA). Net uptake of P by seedlings (milligrams per seedling) decreased in a curvilinear (concave) manner as the fraction of the roots in contact with FA increased. Net uptake of K (milligrams per seedling) and water (milliliters per seedling) by seedlings decreased linearly as the fraction of the roots in contact with FA increased. Net uptake of P, K, and water by seedlings was reduced 57, 75, and 29%, respectively, when the whole root system was exposed to FA. Net P and K uptake of roots (milligrams per gram root fresh weight) not in contact with FA decreased in a linear and curvilinear (convex) manner, respectively, as the fraction of roots in contact with FA increased. Net P and K uptake of roots in contact with ferulic acid increased in a linear and curvilinear (convex) manner, respectively. Net water uptake of roots (milliliters per gram root fresh weight) not in contact with FA increased in a curvilinear (concave) manner as the frequency of the roots in contact with FA increased. Net water uptake of roots in contact with FA did not show a trend. Transpiration (milliliters per square centimeter) was reduced in a linear manner as the fraction of roots in contact with FA increased. A very slight compensation by roots not in contact with FA for roots in contact with FA was observed for net water uptake rates. No compensation for P and K uptake rates was observed.

Inhibition and recovery of cucumber roots given multiple treatments of ferulic acid in nutrient culture

Ferulic acid, a frequently cited allelopathic agent, inhibited photosynthesis, leaf expansion, and root elongation of cucumber seedlings grown in aerated nutrient cultures in a growth chamber. Other effects were a reduction in the proportion of radioactivity fixed by photosynthesis translocated to roots, a stimulation in secondary root initiation, and an increase in root-shoot ratios. Inhibition of leaf expansion and root elongation induced by multiple ferulic acid treatments was rapidly lost once ferulic acid was removed from the root environment. The changes in general root morphology, i.e., average root length and root number, associated with ferulic acid treatments, were partially reversed or not affected when ferulic acid was removed from the root environment.

Effects of various mixtures of ferulic acid and some of its microbial metabolic products on cucumber leaf expansion and dry matter in nutrient culture

Cucumber seedlings (Cucumis sativus cv. ‘Early Green Cluster’) ranging from 6 to 16 days of age were treated with various concentrations (0– 1 mM) of caffeic, ferulic,p-coumaric,p-hydroxybenzoic, protocatechuic, sinapic, syringic, and vanillic acids and mixtures of ferulic acid and one or two of the other phenolic acids. Seedlings were grown in full-strength Hoaglands solution which was changed every other day. Phenolic acid treatments were given with each nutrient solution change starting at day 6 or given once when seedlings were 13 or 14 days old. Leaf area, mean relative rates of leaf expansion, transpiration rates, water utilization, and the concentrations of the phenolic acids in nutrient solution were determined at one- or two-day intervals. Seedling dry weight was determined at final harvest. Seedling leaf area and dry weight were linearly related. Since leaf areas can be easily obtained without destructive sampling and leaf area expansion responds rapidly to phenolic acid treatments, it was utilized as the primary indicator of plant response. The resulting data suggested that a number of ferulic acid microbial metabolic products, as well as two other phenolic acids observed in soils (p-coumaric and syringic acid), can reduce seedling dry weight, leaf expansion, and water utilization of cucumber seedlings in a similar manner. The magnitude of impact of each of the phenolic acids, however, varied with phenolic acid and concentration. It appears that the inhibitory activity of these phenolic acids involved water relations of cucumber seedlings, since the phenolic acid treatments resulted in closure of stomata which then remained closed for several days after treatment. The data also demonstrated that the effects of mixtures of phenolic acids on cucumber seedlings may be synergistic, additive, or antagonistic. The type of response observed appeared to be related to the factor measured, the compounds in the nmixture, and the magnitued of inhibition associated with each compounds. The data also indicated that the effects of the various phenolic acids were reversible, since seedling leaf area increased rapidly once phenolic acids were removed from the root environment. Mean relative rates of leaf expansion recovered even in the presence of the various phenolic acids.

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.