An Improved Process for Isolation of Corn Fiber Gum

Sequential alkaline extraction and alkaline hydrogen peroxide (AHP) bleaching have been used to prepare corn fiber gum in yields ranging from 21 to 40%, depending on the pH of the extraction medium. The pH was adjusted by using different ratios of NaOH and Ca(OH)₂ The whitest product was obtained after AHP bleaching of the extract obtained using the lowest pH value. In order for the product gum to give its characteristic clear and low viscosity solutions, it was necessary to remove starch from the corn fiber substrate using α-amylase. The water-insoluble hemicellulose A fraction, a minor component, was removed by neutralizing AHP-treated extracts before ethanol precipitation of the useful hemicellulose B (corn fiber gum) fraction. At ambient temperature, AHP bleaching was near optimal after ≈2 hr under the processing conditions used. High ratios of arabinose (39%) to xylose (50%) were present in the corn fiber gum extracted under various alkaline conditions, and the H₂O₂ processing step did not significantly alter these ratios. The same low levels of galactose (7%) and glucuronic acid (4%) were present regardless of the extraction conditions. Molecular mass of the corn fiber gum preparations ranged from 2.78 × 10⁵ for the material extracted with Ca(OH)₂ to 3.94 × 10⁵ for the material extracted with NaOH. Molecular mass was unaffected by the H₂O₂ present in the second processing step. As expected for a carbohydrate polymer with a rather low uronic acid content, solution viscosities were unaffected by the presence of salt.     

Isolation and Characterization of Cellulose/Arabinoxylan Residual Mixtures from Corn Fiber Gum Processes

White, fluffy cellulose/arabinoxylan mixtures (CAX) were generated from the solid residues remaining after corn fiber gum (CFG) production. Most CAX were produced using variations of a process in which a single alkaline hydrogen peroxide (AHP) step was used for delignification and for CFG (arabinoxylan) extraction. The optimal ratio of H₂O₂ to corn fiber to water was 0.1:1:20. Holding this ratio constant, time and temperature conditions were systematically varied, and yields of CAX and CFG determined. Parallel processes were conducted without H₂O₂ to determine its effect on CAX and CFG yield. CAX prepared under identical conditions but without H₂O₂ retained nearly twice the levels of CFG sugars, as revealed from L‐arabinose, D‐xylose, and D,L‐galactose levels. Even the CAX prepared under extreme AHP conditions (1 hr, 100°C), however, contained 32.9% of these CFG sugars. This CAX was obtained in a 25.1% yield, whereas those produced under less vigorous conditions were obtained in higher yields, because they retained more CFG. CAX prepared in the presence of H₂O₂ hydrated very effectively, as indicated by their high swollen volumes and water absorbance values. This suggests potential food applications for CAX as a bulking agent. In addition, the open structure of the CAX matrix would render these residues suitable for chemical derivatization and enzymatic saccharification.     

Molecular Characteristics of Corn Fiber Gum and Their Influence on CFG Emulsifying Properties

The molecular characteristics of two purified arabinoxylan fractions derived from corn kernels, corn fiber gum-1 and -2 (CFG-1 and -2), have been studied and correlated with emulsifying properties. CFG-1 and -2 fractions were isolated from different corn fiber sources by 1) a sequential alkaline extraction and H₂O₂ bleaching to produce CFG-1; and 2) additional H₂O₂ treatment of the alkali-extracted residue at pH 11.5, yielding CFG-2. Multiangle laser light-scattering and online viscosity were used to measure the molar mass, polydispersity, structure compactness, and intrinsic viscosity of the generated CFG fractions. Emulsification properties in an oil-in-water emulsion system with 10:1 oil-to-gum ratio was investigated by measuring turbidity of an aliquot from the bottom of the diluted emulsion over 10 days. The isolated CFG-2 from each fiber source was higher in weight-average molar mass (M_w) polydispersity) (M_w/M_n) and structure compactness, and also lower in solution weight-average intrinsic viscosity (η_w) than the corresponding CFG-1. Average M_w and η_w values were 244–491 kDa and 1.35–1.84 dL/g, respectively. The emulsion stabilizing capacity of CFG-2 from each fiber source was superior to the corresponding CFG-1.     

THE HYDROLYTIC EXTRACTION OF CARBOHYDRATES FROM SOIL BY SULPHURIC ACID

The parameters of the extraction and hydrolysis of soil carbohydrates by methods involving 24N H₂SO₄ and N H₂SO₄ were studied for a sandy granitic loam. Sugars were measured by alkaline ferricyanide, orcinol, anthrone (hexoses), orcinol-ferric chloride (pentoses), cysteine-sulphuric acid (methyl pentoses), glucose oxidase (glucose), and also by analysis of the individual sugars by paper chromatography. After shaking the soil with 24N H₂SO₄ at 20° C a further period of treatment with N H₂SO₄ at 100° C was required to obtain maximum hydrolysis of the soil carbohydrates. This period decreased from 17 h to 5 h as the time with 24N H₂SO₄ increased from 2 to 16 h. NH₂SO₄ at 100° C alone was less efficient. The extraction of pentoses by 24N H₂SO₄ reached a maximum within 8 h, and methyl pentoses within 4 h, both declining thereafter. The release of hexoses was continuing after 40 h. The best compromise involved extraction with 24N H₂SO₄ for 16 h followed by hydrolysis with N H₂SO₄ for 5 h. By this treatment yields of pentoses and methyl pentoses were respectively 99 and 92 per cent of the maximum obtainable.     

β-Glucanase Activity and Molecular Weight of β-Glucans in Barley After Various Treatments

β-Glucanase activity interferes with molecular characterization of mixed-linkage (1→3)(1→4)-β-d -glucans (β-glucans). Reductions in β-glucanase activity were determined after barley cvs. Azhul, Waxbar, and Baronesse were treated with autoclaving (120°C, 45 min), calcium chloride (0.05M, 1 hr), 70% ethanol (80°C, 4 hr), hydrochloric acid (0.1N, 1 hr), oven heating (120 and 140°C, 40 min), sodium hydroxide (0.0025M, 1 hr), and 5% trichloroacetic acid (TCA) (40°C, 1 hr). High-performance size-exclusion chromatography (HPSEC) of α-amylase-treated aqueous extracts was used to demonstrate the effects of treatments on the molecular weights of β-glucans. The HPSEC system included multiple-angle, laser light scattering, refractive index, and fluorescence detectors. β-Glucanase activities, ranging from 52 to 65 U/kg of barley, were reduced by autoclaving (50–75%), hot alcohol (67–76%), oven heating (40–96%), CaCl₂ (75–95%), NaOH (76–89%), and TCA (92–96%). Some malt β-glucanase activity remained after most treatments. HCl and TCA treatments reduced extraction and molecular weights of β-glucans. Weight-average molecular weights (M_w) for β-glucans extracted with water at 23°C were low (most <8 × 10⁵). Base treatment (pH 9) and extraction at 100°C for 2.5 hr resulted in the greatest extraction of β-glucans and highest M_w. As a result, the conditions seem appropriate for measurement of physical characteristics of β-glucans in cereal products.     

Pretreatment and Enzymatic Hydrolysis of Sorghum Bran

Sorghum bran has potential to serve as a low‐cost feedstock for production of fuel ethanol. Sorghum bran from a decortication process (10%) was used for this study. The approximate chemical composition of sorghum bran was 30% starch, 18% hemicellulose, 11% cellulose, 11% protein, 10% crude fat, and 3% ash. The objective of this research was to evaluate the effectiveness of selected pretreatment methods such as hot water, starch degradation, dilute acid hydrolysis, and combination of those methods on enzymatic hydrolysis of sorghum bran. Methods for pretreatment and enzymatic hydrolysis of sorghum bran involved hot water treatment (10% solid, w/v) at 130°C for 20 min, acid hydrolysis (H₂SO₄), starch degradation, and enzymatic hydrolysis (60 hr, 50°C, 0.9%, v/v) with commercial cellulase and hemicellulose enzymes. Total sugar yield by using enzymatic hydrolysis alone was 9%, obtained from 60 hr of enzyme hydrolysis. Hot water treatment facilitated and increased access of the enzymes to hemicellulose and cellulose, improving total sugar yield up to 34%. Using a combination of starch degradation, optimum hot water treatment, and optimum enzymatic hydrolysis resulted in maximum total sugar yield of up to 75%.     

Mineralisation of Surfactants Using Ultrasound and the Advanced Fenton Process

The destruction of the surfactants, sodium dodecylbenzene sulfonate (DBS) and dodecyl pyridinium chloride (DPC), using an advanced oxidation process is described. The use of zero valent iron (ZVI) and hydrogen peroxide at pH = 2.5 (the advanced Fenton process), with and without, the application of 20 kHz ultrasound leads to extensive mineralisation of both materials as determined by total organic carbon (TOC) measurements. For DBS, merely stirring with ZVI and H₂O₂ at 20°C leads to a 51% decrease in TOC, but using 20 kHz ultrasound at 40°C, maintaining the pH at 2.5 throughout and adding extra amounts of ZVI and H₂O₂ during the degradation, then the extent of mineralisation of DBS is substantially increased to 93%. A similar result is seen for DPC where virtually no degradation occurs at 20°C, but if extra amounts of both ZVI and hydrogen peroxide are introduced during the reaction at 40°C and the pH is maintained at 2.5, then an 87% mineralisation of DPC is obtained. The slow latent remediation of both surfactants and the mechanism of degradation are also discussed.     

Mineral-catalyzed peroxidation of tetrachloroethylene

The treatment of perchloroethylene (PCE) was investigated by the promotion of Fenton-like reactions using the iron oxyhydroxide goethite (α-FeOOH) as the sole source of the iron catalyst. A silica sand-goethite matrix was contaminated with 5 mg L^?1 PCE and the oxidative treatments were conducted with 0.15 mM, 2 mM, 5mM, 10mM, 20mM, and 30mM H₂O₂. Perchloroethylene was effectively degraded within 96 hr and the most efficient treatment stoichiometry was observed using 0.15 mM H₂O₂ at pH 3. The degree of heterogeneous catalysis was evaluated by conducting oxidation reactions in parellel systems with an equivalent concentration of soluble iron. The results showed that, within the first 24 hr, up to 94% of the PCE degradation was attributed to heterogeneous catalysis. This modified Fenton's process, when used to treat 5 mg L^?1 PCE in natural subsurface materials with 2 mM H₂O₂ at pH 3, resulted in a residual of 0.20 mg L^?1 PCE after 96 hr.     

Investigation of the changes in low molecular weight organic acids and other physiological parameters by nitric oxide application in two wheat cultivars exposed to boron stress

Boron (B) is one of the essential nutrients for the growth of plants, but its high concentrations are toxic for plants. Thus, B toxicity is a big challenge in crop cultivation. Nitric oxide (NO) is a small signaling molecule that has cytoprotective roles in plants. We investigated whether exogenous sodium nitroprusside (SNP), which is a NO donor, may succeed to alleviate B-induced toxicity in wheat cultivars. Seedlings were grown for 10 days in a growth chamber at 25°C with 16 hr light–8 hr dark photo cycle. After high B application, the effects of SNP on growth parameters; electrolyte leakage (EL); changes in reactive oxygen species [contents of hydrogen peroxide (H₂O₂), malondialdehyde (MDA), and proline]; the activities of antioxidant enzymes [glutathione peroxidase (GSHPx), glutathione reductase (GR), and glutathione S-transferase (GST)] and nitrate reductase (NR); and low molecular weight organic acid (LMWOAs) contents and also chlorophyll and total carotenoid contents were investigated in both shoots and roots of two different wheat cultivars. All experiments were carried out in triplicate. 0.2 mM SNP application ameliorated the chlorophyll and total carotenoid contents, and growth parameters such as shoot length, root length, and fresh weight in both wheat cultivars exposed to B stress. SNP reduced the B-induced lipid peroxidation, EL, and proline and H₂O₂ content in wheat cultivars. SNP application also increased the activities of NR and antioxidant enzymes, including GSHPx, GR, and GST in wheat cultivars exposed to B toxicity. All of the tested LMWOAs including succinic, propionic, butyric, oxalic, formic, malic, malonic, and benzoic acids were increased by SNP treatment in the shoots and roots of both wheat cultivars exposed to B toxicity. In conclusion, results obtained from this study have demonstrated that interactive effects of SNP with B considerably reduced the toxic effects of B in wheat cultivars.     

Changes in Water Absorption and Swollen Volume in Extruded Alkaline Peroxide Pretreated Rice Hulls

Rice hulls were pretreated with an alkaline (pH 11.5) solution of hydrogen peroxide (1%) and then extruded. Pretreatment of rice hulls (4% db) at 50°C for 12 hr promoted 94.4% silica reduction, caused lignin solubilization and increased water absorption index (54%) and swollen volume (44%). The effects of temperature (125, 175, and 225°C), moisture content (25, 30, and 35%) and screw speed (120, 140, and 160 rpm) on water absorption and swollen volume of rice hulls fiber were evaluated after extrusion in a single-screw extruder. Operational conditions that produced the most modified product with regard to the functional properties were: 125°C, 35% moisture, and 120 rpm. Extruded fiber had a water absorption index 95% higher and swollen volume 138% higher than the unprocessed material. Microscopic examination showed a slight effect on the hulls epidermis after pretreatment, while extrusion promoted cellular structure disruption.     

Oxidative coupling activity in soil extracts

An assay was developed in which 50 mm citrate buffer was used to extract catalysts responsible for oxidative coupling reactions in soil. The assay was based on the formation of the dimerized quinone from the oxidative coupling of 2,6-dimethoxyphenol. Oxidative coupling activity was destroyed by heating the extracts for 15 min at 100°C and was inhibited by H₂O₂ (98% at 10 mm), KCN (96% at 10 mm), dithiothreitol (100% at 0.1 mm) and 2,3-dimercapto-1-propanol (100% at 0.1 mm). In the assay, a pH of 7.0 and a temperature of 55°C were determined as optimal. Freezing the soil extracts provided the best protection against activity loss, whereas storage caused a decline in oxidative coupling activity as a function of increasing temperatures (5–50°C). If soil had been supplemented with sucrose before extraction, activity increased. Soil extracts reacted with syringaldazine, benzidine, o-dianisidine, guaiacol, and p-cresol, substrates previously used to detect phenoloxidase enzymes.     

Sauerstoffabsorption beeinflußende Reaktionen bei Ligninen

Reactions of lignins which influence oxygen absorption During the isolation of lignin by n-dioxane, acetone, dimethyl sulfoxid and acetic acid in presence of anorganic acids (HCl, H₂SO₄), forced by a longer extraction at 80 ± 5°C and especially in air atmosphere, condensed structures, aliphatic double bonds in phenyl cumarone systems and stilbenoide units, as well as α, β-keto groups, phenolic hydroxyl and free radicals increased, while methoxyl and aldehyde groups diminished. Less altered lignin took up relative small amounts of O₂ in the alkaline medium n-dioxane/H₂O/NaOH and in the neutral medium γ-butyrolactone/H₂O/SiO₂. In the acid medium γ-butyrolactone/H₂O/SiO₂/hydroquinone chemical bonds between lignin and the (aut-)oxidation products of hydroquinone decreased the oxygen uptake. These bonds were forced by phenolic hydroxyl and free radicals. The oxidation medium with hydroquinone, being very similar to natural conditions of decomposition is also more effective than the other investigated media in oxidizing less altered lignin like such as in straw. Reaction mechanisms for the alterations of lignin during its isolation as well as for oxidative processes in the choosen media are discussed.     

Formulation,synthesis and characterization of boron phosphate (BPO4) compounds as raw materials to develop slow‐release boron fertilizers

We investigated the use of boron phosphate (BPO₄) as a slow‐release boron (B) source. Boron phosphate compounds were synthesized by mixing boric acid (H₃BO₃) and phosphoric acid (H₃PO₄) and heating at temperatures of 25 to 1000°C for 1 or 24 h. X‐ray diffraction (XRD) patterns and chemical analysis confirmed the formation of BPO₄. The crystallinity of these compounds increased with increasing temperature and heating time. The compounds synthesized at 300°C or less were hygroscopic and clumped together, while those synthesized at 500 to 1000°C were non‐hygroscopic and free‐flowing. The solubility of these compounds was assessed at different pH and P concentrations, and compared to the solubility of ulexite and colemanite. The solubility of the BPO₄ compounds decreased with increasing synthesis temperature and with decreasing pH. The solubility and the kinetics of B release from BPO₄ compounds synthesized at 500 and 800°C were slower than for most commonly used B sources. Given their slow dissolution, the BPO₄ compounds may have potential to continuously supply B to crops in environments where B leaching is a problem. The compounds synthesized at 500 and 800°C show potential for co‐granulation with macronutrient fertilizers such as mono‐ammonium phosphate to produce slow‐release B‐enriched granules.     

Preparation and Solubility of Phosphorylated β-Cyclodextrins

β-Cyclodextrin (CD) was phosphorylated with phosphoryl chloride in aqueous alkaline media at different temperatures and pH values. The phosphorylated cyclodextrin (PCD) were characterized by phosphorus contents and positions of substitution as determined by ³¹P-NMR spectroscopy. Reaction of CD with equivmolar POCl₃ for 3 hr at pH 12 and 45°C yielded in a PCD with a phosphorus content of 5.67%. The ratio of monoand diphosphate esters increased when the reaction temperature was raised from 25 to 60°C. The monoesterified phosphate groups were mainly located at C-6 of the anhydroglucose units when the reaction pH was 11 or 12. Reactions at pH 10, however, led to a higher degree of substitution at C-2 than at C-6. Phosphorylation enhanced the water solubility of CD. Solubility of a PCD (5.65% phosphorus) was 35% at pH 8 and 25°C. Simultaneously, solubility of the PCD in 25% ethanol in water was much greater than unsubstituted CD (22.3 vs. 2.8%). The PCD enhanced the water solubility of nonpolar compounds, such as β-carotene.     

Acid pyrophosphate extraction of soil fulvic acids

A simple three step method is described for isolation of soil fulvic acids in high yield. The complexing agent H₂P₂O₇^2? (at pH 2) is used to release soil-bound fulvic acids. Extraction of humic acids is minimal. Selective separation of the protonated fulvic acids from the ionic extractant is achieved on a non-ionic polyacrylate resin (Amberlite XAD-7); after washing the resin, fulvic acids were retrieved in >98% yield by adjusting the pH to 6.5. Two problems associated with the classical alkali extraction method are avoided: possible alkaline oxidation of phenolic components, and their oxidation by Fe³⁺ under the acidic conditions employed to precipitate humic acids. The product typically has an ash weight of <0.6% after one XAD treatment. The method has been applied to three soils and one IHSS peat sample.     

Comparison of Asian Noodles from Some Hard White and Hard Red Wheat Flours

Asian noodles were prepared by an objective laboratory method that included adding optimum water to the dry ingredients, mixing the ingredients to homogeneous salt distribution, and sheeting of the dough under low shear stress. The lightness (L*) values of alkaline‐ and salt‐noodle doughs made from 65% extraction hard white wheat flours (except KS96HW115 flour at ≈70% extraction) were higher than those from 60% extraction hard red wheat flours (except Karl 92 flour at ≈70% extraction). A hard white spring wheat, ID377s, and a Kansas line of hard white winter wheat, KS96HW115, to be released in 2000, gave the highest L* values for dough sheets stored for 2 and 24 hr at 25°C. Cooking losses were 5–9 percentage points higher for alkaline noodles than salt noodles, but the cooking yields of the two types of Asian noodles were almost the same. Cooked alkaline noodles made from a high‐swelling flour (SP₉₃≈21 g/g) gave higher tensile strength than those made from several low‐swelling flours (SP₉₃ ≈15 g/g) with the same protein contents (≈12.5%). However, the cooked salt noodles gave the same tensile strength.     

Extraction of Wheat Starch with Aqueous Sodium Hydroxide

Wheat starch was extracted with aqueous sodium hydroxide at 30–38% starch solids, pH 11.5–12.3, and 25–42°C for 0.17–24 hr. Stirring wheat starch at pH 12.3 and 25°C for 3 and 24 hr, then washing with water, neutralizing, and washing again, removed 70 and 90% phosphorus (P), respectively. Adding 16% sodium sulfate (dry starch basis) into the alkaline medium removed ≈80% of P at pH 12.0 and 25°C in 3 hr and >95% of P at pH 11.7 and 42°C in 3 hr. Sulfate ion was absorbed strongly by wheat starch in aqueous sodium hydroxide at pH 12.0, and sodium sulfate also increased the starch's uptake of hydroxide ion. Low-P wheat starch (>90% of P removed) retained the fatty acids in the untreated starch, but a fatty acid-amylose complex was not detectable by differential scanning colorimetry. The enthalpy of gelatinization of the low-P wheat starch almost matched that of prime starch, as did its X-ray diffraction pattern. Those data are consistent with saponification of the lysophospholipid in the amorphous phase of the starch to form fatty acid salts and glycerol-choline or glycerol-ethanolamine phosphodiesters that slowly diffused out of the granules. The low-P wheat starch was judged to have less “cereal” odor than the prime starch, and its pasting temperature at 9.3% starch solids was lowered by ≈10°C.     

Kinetic and Thermodynamic Studies of Chlorinated Organic Compound Degradation by Siderite-Activated Peroxide and Persulfate

The efficacy of two oxidant systems, iron-activated hydrogen peroxide (H₂O₂) and iron-activated hydrogen peroxide coupled with persulfate (S₂O₈ ^2?), was investigated for treatment of two chlorinated organic compounds, trichloroethene (TCE) and 1,2-dichloroethane (DCA). Batch tests were conducted at multiple temperatures (10–50 °C) to investigate degradation kinetics and reaction thermodynamics. The influence of an inorganic salt, dihydrogen phosphate ion (H₂PO₄ ^?), on oxidative degradation was also examined. The degradation of TCE was promoted in both systems, with greater degradation observed for higher temperatures. The inhibition effect of H₂PO₄ ^? on the degradation of TCE increased with increasing temperature for the iron-activated H₂O₂ system but decreased for the iron-activated hydrogen peroxide-persulfate system. DCA degradation was limited in the iron-activated hydrogen peroxide system. Conversely, significant DCA degradation (87% in 48 h at 20 °C) occurred in the iron-activated hydrogen peroxide-persulfate system, indicating the crucial role of sulfate radical (SO₄ ^??) from persulfate on the oxidative degradation of DCA. The activation energy values varied from 37.7 to 72.9 kJ/mol, depending on the different reactants. Overall, the binary hydrogen peroxide-persulfate oxidant system exhibited better performance than hydrogen peroxide alone for TCE and DCA degradation.     

Biotreatment of Melanoidin-Containing Distillery Spent Wash Effluent by Free and Immobilized Aspergillus oryzae MTCC 7691

Humic acids (HA) are known as the precursors of carcinogenic compounds formed by the disinfection of drinking water. While conventional treatments were found to be inefficient HA removal processes in drinking water, advanced oxidation processes have been proven to have a significant effect in the treatment of HA. The degradation of HA was investigated using nano-sized zinc oxide (ZnO)/laponite composite (NZLC). The reactions occurred in a UVC reactor by considering following variables: pH, initial HA concentration, catalyst loading, addition of hydrogen peroxide (H₂O₂), and catalyst reuse. Water samples containing HA were analysed by ultraviolet/visible spectrophotometer and high-performance size-exclusion chromatography. Initial HA concentrations were tested by the Langmuir–Hinshelwood model with k and K _ads values, determined to be 0.126 mg/L.min and 0.0257 L/mg, respectively. The change in pH affected the HA degradation efficiency by the photocatalytic activity where it was higher under acidic conditions rather than alkaline ones. Optimal catalyst loading was proved to be a constrained factor in influencing the photocatalytic efficiency: the increase of catalyst concentration enhanced the HA decomposition efficiency up to an optimum value of 20 g/L, where there was no further degradation with excess loading. The addition of H₂O₂ was investigated through homogenous and heterogeneous photocatalysis, and, heterogeneous photocatalysis showed higher removal efficiency due to the combined effect of both catalysts and H₂O₂. Finally, NZLC was effective for reuse and exhibited an excellent stability after six times of usage.     

Effects of chemical and heat treatments on ethylene production in soil

Factors influencing C₂H₄ production in a silt loam were investigated in an effort to determine the source of this gas in soil. Air-dried samples of soil in glass vials were moistened to about ?10 kPa, sealed with rubber septa, and incubated at 30 or 35°C with an original atmosphere of air or O₂-free N₂. C₂H₄ concentrations in the vials were determined by gas chromatography.Addition of the antibacterial agents chloramphenicol or novobiocin to the soil inhibited C₂H₄ production, whereas the antifungal agent cycloheximide had no effect. Sodium azide and sodium cyanide also reduced C₂H₄ production. Treatment of the soil with moist heat (i.e. passing a steam-air mixture through it) at 80°C for 30 min failed to reduce the ability of the soil to produce C₂H₄ during subsequent incubation at 30°C, but autoclaving it twice at 121°C prevented C₂H₄ production. As with nonheated soil, C₂H₄ production from soil treated at 80°C was prevented by novobiocin but not by cycloheximide. Only about 10% of the bacteria isolated from nontreated soil were spore-formers. In contrast, 95–98% and possibly more of the bacteria isolated from heat-treated soil were spore-formers, including those in soil which was heat-treated and then incubated moist at 30°C for an additional 3 days before dilution plating. Addition of methionine had no effect on the production of C₂H₄ in anaerobic soil, whereas ethionine, chlorogenic acid, and ethylenediaminetetraacetic acid (EDTA) all enhanced C₂H₄ production. Ethionine, but not chlorogenic acid or EDTA, also resulted in considerable C₂H₄ accumulation in autoclaved soil; the C₂H₄ detected in ethionine-amended soil was apparently nonmicrobial in origin. Soil samples incubated at constant temperatures of 30, 50, or 70°C all produced C₂H₄.The results collectively indicate that C₂H₄ in soil is most likely produced by facultative or strictly anaerobic bacteria which are probably spore-formers and may also be thermophilic. Several isolates of spore-forming bacteria were inoculated into autoclaved soil, but none produced appreciable amounts of C₂H₄ under the test conditions.