Role of Azospirillum brasilense nitrate reductase in nitrate assimilation by wheat plants

Summary The nitrogen metabolism of wheat plants inoculated with various Azospirillum brasilense strains and nitrate reductase negative (NR^–) mutants was studied in two monoxenic test tube experiments. The spontaneous mutants selected with chlorate under anaerobic conditions with nitrite as terminal electron acceptor fixed N₂ in the presence of 10 mM NO₃ ^– and were stable after the plant passage. One strain (Sp 245) isolated from surface-sterilized wheat roots produced significant increases in plant weight at both NO₃ levels (1 and 10 mM) which were not observed with the NR^– mutants or with the two other strains. Similar effects were observed in a pot experiment with soil on dry weight and total N incorporation but only at the higher N fertilizer level. In the monoxenic test tube experiments plants inoculated with the mutants showed lower nitrogenase activities than NR⁺ strains at the low NO₃ ^– level (1 = mM) but maintained the same level of activity with 10 mM NO₃ ^– where the activity of all NR⁺ strains was completely repressed. The nitrate reductase activity of roots increased with the inoculation of the homologous strains and with the mutants at both NO₃ ^– levels. At the low NO₃ ^– level this also resulted in increased activity in the shoots, but at the high NO₃ ^– level the two homologous strains produced significantly lower nitrate reductase activity in shoots while the mutants more than doubled it. The possible role of the bacterial nitrate reductase in NO₃ ^– assimilation by the wheat plant is discussed.     

Stabilization of ascorbic acid and its measurement by liquid chromatography in nonfat dry milk

The determination of ascorbic acid by liquid chromatography (LC) was improved by performing the analysis in the presence of solvents that had been purged with argon to reduce the concentration of oxygen. This methodological modification eliminated the oxidation of ascorbic acid during the chromatographic procedure and reduced the minimum detection level to 1 microgram. Solutions of ascorbic acid have been successfully stabilized for 67 days by addition of dithiothreitol to a deaerated solution of water-acetonitrile (25 + 75 v/v), sealed under argon in amber vials and stored at -20 degrees C. In a second independent study, a procedure for the extraction of ascorbic acid from nonfat dry milk in a single step was developed. The ascorbic acid content of Nonfat Dry Milk (SRM 1549) was determined by LC, using the method of standard additions. The mean ascorbic acid content was 54 +/- 5 micrograms/g of sample. Analysis of variance of the analytical results indicates that there is a significant continual increase in the content of the ascorbic acid in each bottle from first to last sample.     

Review of chromatographic methods for chloramphenicol residues in milk, eggs, and tissues from food-producing animals

Although chloramphenicol is not approved for use in food-producing animals in the United States, this broad spectrum antibiotic has been widely used to treat diseases in such animals including the lactating dairy cow. Extremely low ophthalmologic doses of chloramphenicol are known to cause aplastic anemia in humans. The residues in meat, milk, and eggs intended for human consumption cause particular public health concern because the bone marrow aplasia is not dose dependent. Furthermore, chloramphenicol, a known inhibitor of protein synthesis, also retards erythropoiesis, a condition that is dose dependent and may cause allergic hypersensitivity reactions. This paper is a review of sensitive methods that use gas, liquid, thin layer, and simple column chromatography as both determinative and cleanup steps for detecting and quantitating chloramphenicol in edible animal tissues, milk, and eggs.     

Determination of fat in vegetable foods

The fat in vegetable foods--tree nuts, peanuts, sunflower seeds, avocado, and olives--can be determined volumetrically by acid digestion of the material and separation of the fat. The assay can be performed conveniently by using the equipment developed for fat determination of milk (Gerber method). The results agree well with those obtained by Soxhlet extraction. The advantages of using the Gerber method for vegetable foods are simplicity, speed, low operation cost, and elimination of the use of inflammable solvents.     

Effect of Nitrogen Fertilization on Quantity of Flour Protein Components,Dough Properties,and Breadmaking Quality of Wheat

Three winter wheat varieties with differing breadmaking quality were grown at two locations in two years at 0 or 3 × 60 kg of nitrogen application. The effect of nitrogen on amount of different components of gluten proteins was determined by reverse-phase HPLC. A high amount of nitrogen led generally to a significant increase of total protein content. However, this increase was obvious only for the gluten proteins; albumins and globulins remained nearly unaffected. The effect of increased protein content on gliadin to glutenin (gli-glu) ratio was inconsistent. While increased protein content increased the gli-glu ratio in the variety Capo, the opposite was true for the variety Renan. Gli-glu ratio of the variety Lindos showed no discernible tendency. As total protein content increased, the ratio of low molecular weight (LMW) to high molecular weight (HMW) glutenins decreased consistently, i.e., in all varieties, in both years and locations. Change of LMW to HMW ratio showed a significant negative correlation to sedimentation value and bread volume. There was no consistent change in the ratio between x- and y-type HMW subunits due to fertilization, as could be shown by densitometric measurements on SDS-PAGE gels. This ratio appeared to be dependent on the genotype and has decreased with decreasing quality. The amount of x-type subunits correlated closely with sedimentation value and bread volume. These results suggest that ratio of HMW glutenins, especially x-type subunits, to total protein content could be the best early detectable parameter with high predictive value for breadmaking quality.     

Short-term effects of nitrogen on methane oxidation in soils

 The short-term effects of N addition on CH₄ oxidation were studied in two soils. Both sites are unfertilized, one has been under long-term arable rotation, the other is a grassland that has been cut for hay for the past 125 years. The sites showed clear differences in their capacity to oxidise CH₄, the arable soil oxidised CH₄ at a rate of 0.013 μg CH₄ kg^–1 h^–1 and the grassland soil approximately an order of magnitude quicker. In both sites the addition of (NH₄)₂SO₄ caused an immediate reduction in the rate of atmospheric CH₄ oxidation approximately in inverse proportion to the amount of NH₄ ⁺ added. The addition of KNO₃ caused no change in the rate of CH₄ oxidation in the arable soil, but in the grassland soil after 9 days the rate of CH₄ oxidation had decreased from 0.22 μg CH₄ kg^–1 h^–1 to 0.13 μg CH₄ kg^–1 h^–1 in soil treated with the equivalent of 192 kg N ha^–1. A ¹⁵N isotopic dilution technique was used to investigate the role of nitrifiers in regulating CH₄ oxidation. The arable soil showed a low rate of gross N mineralisation (0.67 mg N kg^–1 day^–1), but a relatively high proportion of the mineralised N was nitrified. The grassland soil had a high rate of gross N mineralisation (18.28 mg N kg^–1 day^–1), but negligible nitrification activity. It is hypothesised that since there was virtually no nitrification in the grassland soil then CH₄ oxidation at this site must be methanotroph mediated. Received: 31 October 1997