Transformation of atmospheric NO2 absorbed in spinach leaves

NO₂ fumigation at 8 ppm of spinach plants resulted in nitriteaccumulation in the leaves in the dark but not in the light.When spinach plants were fumigated with ₁₅N-labeled NO₂ in thelight, amide nitrogen of glutamine, glutamic acid, -amino butyricacid and aspartic acid, in this order, were highly labeled with¹⁵N and nitrate was also labeled. These results suggest thatNO₂-nitrogen (at least some of it) is converted into nitriteand nitrate, and then actively assimilated into amino acidsthrough the glutamine synthetase/glutamate synthase pathwayin spinach leaves. ¹This work was conducted as a part of the special research project"Studies on evaluation and amelioration of air pollution byplants" (1976–1978) at the National Institute for EnvironmentalStudies. (Received July 24, 1978; )     

Ethylisocyanide equilibria of hemoglobins M Iwate, M Boston, M Hyde Park, M Saskatoon, and M Milwaukee-I in half-ferric and fully reduced states.

The ethylisocyanide equilibria of all the five known hemoglobins M, namely Hb M Iwate (alpha287 Tyrbeta2), Hb M Boston (alpha258 Tyrbeta2), Hb M Hyde Park (alpha2beta292 Tyr), Hb M Saskatoon (alpha2beta263 tyr), and Hb M Milwaukee-I (alpha2beta267 Glu), were studied both in the half-ferric and fully reduced heme states. In the half-ferric state, no heme-heme interaction was observed for Hb M Iwate, Hb M Boston, and Hb M Hyde Park, but Hb M Saskatoon and Hb M Milwaukee-I show small but definite heme-heme interaction with Hill's n of 1.3. The beta chain mutants, Hb M Hyde Park and Hb M Saskatoon, have almost normal affinity for ethylisocyanide and a normal Bohr effect, whereas the alpha chain mutants, Hb M Iwate and Hb M Boston, have abnormally low affinity and almost no Bohr effect. Hb M Milwaukee-I showed a large Bohr effect and low affinity. These results are consistent qualitatively with those on oxygen equilibria reported previously. In the fully reduced state, in which all four hemes were in the ferrous state and capable of binding ethylisocyanide distinct differences were found in the extent of heme-heme interaction. Namely, the n values for proximal histidine mutants, Hb M Iwate and Hb M Hyde Park, were 1.1 and 1.0, respectively, whereas the distal histidine mutants, Hb M Boston and Hb M Saskatoon, showed high n values of 2.4 and 1.6, respectively. Hb M Milwaukee-I also exhibited a high n value of 2.0 The ethylisocyanide affinity of the four histidine mutants was high compared with that of Hb A, while that for Hb M Milwaukee-I was almost normal. All five Hbs M had approximately normal magnitudes of Bohr effect. In the half-ferric state, the proximal and distal histidine mutants of the same chain showed similar affinity for ethylisocyanide and Bohr effect, rather different from those of the mutants of the opposite chain. These differences seem to be derived from the difference of abnormal bonding of ferric iron to tyrosine or glutamic acid. On the other hand, the reduction of iron, which abolished the abnormal bonding and made all of the chains capable of binding ligand, extinguished the differences of alpha and beta chains, and the effect of amino acid side chains close to iron on ligand binding properties became clear. Proximal histidine, which is considered to trigger the transition between the T and R states, seems to be essential to the heme-heme interaction.     

Interaction of poly- ,L-glutamic acid with acridine orange

Interaction of poly-α,L -glutamic acid (PGLA) with acridine orange (AO) was studied with circular dichroism and absorption spectra measurements. The following results were observed: (1) the addition of a comparable amount of AO with the glutamyl residue to the PLGA solution at pH of 4.5 reduced the fraction of helix of the polymer; (2) when AO was added to the PGLA solution, the pH range of the helix-coil transition of the polymer shifted toward higher pH regions; and (3) when the mixture of the same amount of AO and the glutamyl residue was brought to the neutral and alkaline pH region, some induced circular dichroism bands were observed. In this case, it was assumed that PLGA in the system takes a helical form due to the neutralization of the anionized side chains by the cationic species of AO. We concluded that AO molecules bound to the carboxylate groups of the side chains of PLGA arrange to from a righthanded super-helix which surrounds the core of the righthanded α-helix of PLGA.     

Changes in intermediate haemoglobins during autoxidation of haemoglobin.

Initial-rate measurements were made of the oxidations of pyridine-3-methanol and glycerol by NADP+ and of the reduction of the corresponding aldehydes by NADPH catalysed by pig kidney aldehyde reductase. In addition, a brief survey of the specificity of the enzyme towards aldehyde substrates and its sensitivity to the inhibitors ethacrynic acid, sodium barbitone and warfarin was made. The detailed kinetic work indicates a compulsory mechanism for aldehyde reduction, with NADPH binding before aldehyde. For alcohol oxidation, however, it is necessary to postulate the formation of kinetically significant amounts of binary complexes of the type enzyme-alcohol to explain the results. Thus, for alcohol oxidation random-order addition of substrates may occur. Inhibition studies of the kinetics of aldehyde reduction in the presence of the corresponding alcohol product provide further evidence for the existence of enzyme-alcohol complexes. Finally, detailed kinetic studies were made of the inhibition of pyridine-3-aldehyde reduction by sodium barbitone. The mechanism of the inhibition is discussed.     

Effects of pH and Osmotic Stress on Cellular Polyamine Contents in the Soybean Rhizobia Rhizobium fredii P220 and Bradyrhizobium japonicum A1017

Homospermidine is a polyamine present in its highest concentrations in root nodule bacteria. By using the soybean rhizobia Rhizobium fredii P220 and Bradyrhizobium japonicum A1017, the effects of the pH and osmolarity of the medium on rhizobial growth and cellular polyamine contents were investigated. Elevation of medium pH repressed the growth of slowly growing B. japonicum A1017 and resulted in a slight increase in cellular putrescine, while homospermidine content was not significantly affected. In contrast, in fast-growing R. fredii P220, which showed good growth over a wide range of the medium pHs from 4.0 to 9.5, homospermidine content increased with the lowering of the medium pH. Under the acid-stressed conditions, cellular Mg²⁺ content in strain P220 also increased. Strain P220 was able to grow in NaCl concentrations up to 0.4 M, while strain A1017 did not grow in media containing 0.15 M NaCl. Glutamic acid and K⁺ contents of salt-tolerant P220 cells increased in response to NaCl concentrations, but homospermidine and Mg²⁺ contents were inversely related to the NaCl concentrations. External salinity had no effect on the contents of other polyamines in P220 cells. On the basis of osmotic strength, NaCl, KCl, sucrose, or glycerol induced similar decreases in cellular homospermidine content. These results suggested that the cellular levels of homospermidine in strain P220 may be regulated by mechanisms related to their pH and osmotic tolerance.     

Respiration and nitrogen fixation of hydroponically cultured Phaseolus vulgaris L. cv. OAC Rico and a supernodulating mutant

This study was undertaken to determine how carbon utilization of fruit production might affect symbiotic activity in hydroponically cultured white bean (Phaseolus vulgaris L.) cv. Rico and its supernodulating genotype R32-BS15 (abbreviated as RBS15). Total plant biomass production of both genotypes was similar. Nodule dry weight of RBS15 consistently scored approximately twice the amount recorded for Rico, while nodule numbers in the mutant were almost six times as high. Nodule respiration on a per-plant basis and specific respiration was initially (day 37) disproportionally higher in the mutant, reaching up to three times the values recorded for Rico, while plant N₂ fixation estimated by ¹⁵N dilution was almost identical. This indicates a lower fixation efficiency of RBS15 nodules, which we suggest to be a result of the larger number of smaller nodules with a higher proportion of growth and maintenance respiration per unit nodule mass. During reproductive development, specific respiration of the mutant dropped below that of Rico without a reduction in fixation, indicating a change in relative efficiency of fixation. Continuous removal of fruits from day 37 onwards stimulated respiration of nodules in both genotypes with highest values per plant being documented for RBS15, while specific activity was higher for Rico. The results indicate that symbiotic activity was not detrimentally affected by competition for carbohydrates between fruits and nodules. It appeared that nodules did not possess excess N₂-fixation capacity which could be stimulated by additional provision of photosynthate. Hence, the early onset of reproduction during the life cycle of P. vulgaris is unlikely to be responsible for inadequate fixation performance in the field.The authors are grateful to Ms. M. Sudoh (National Institute of Animal Industry, Kukizaki, Tsukuba, Ibaraki 305, Japan) for the inductively coupled plasma analyses and to Ms. M. Takebe, Ms. T. Iso (National Agriculture Research Center, Kannondai, Tsukuba, Ibaraki 305, Japan), Ms. K. Kouno (Department of Agricultural Chemistry, Nihon University, Setagaya, Tokyo, Japan) and Mr. K. Hiraoka (Fruit Tree Research Station, Fujimoto, Tsukuba, Ibaraki 305, Japan) for their skilled technical assistance. Drs. B. Buttery and S.J. Park (Agriculture Canada, Research Station Harrow, Ontario, Canada) are thanked for the provision of mutant seeds. One of us (A.P.H.) is indebted to the Alexander von Humboldt-Stiftung and the Japanese International Science and Technology Exchange Center for the provision of an STA Research Fellowship which allowed this collaboration to become possible.     

Effects of aqueous extract of rice plants (Oryza sativa L.) on seed germination and radicle elongation of Monochoria vaginalis var. plantaginea

Laboratory experiments were conducted to examine the allelopathic potentials of rice plants (Oryza sativa L.) on seed germination and radicle elongation of Monochoria vaginalis (Burm. f.) Presl var. plantaginea (Roxb.) Solms-Laubat. To compare the relative allelopathic potentials of rice plants at different growth stages, aqueous extracts of dried tissues were obtained from the seeds, husks, and seedlings harvested 10, 20, 40 and 120 days after seeding. When M. vaginalis seeds were incubated in solutions containing aqueous extracts, significant increases in the germination rate were observed for all the aqueous extracts except for the extract of 10 d-old shoots at higher dosages. When partitioned with ethyl acetate, the germination-promoter(s) in the aqueous extracts remained in the aqueous phase while some inhibitors were extracted with ethyl acetate. In contrast, aqueous shoot extracts added to the incubation media at the same dosage for promotion of seed germination inhibited radicle elongation of M. vaginalis, indicating that the sensitivity of the weed varies between these two growth stages.     

Effect of grandinol and phloroglucinol derivatives on gibberellin-inducible agr;-amylase synthesis in barley aleurone cells

Grandinol derivatives, which show ABA-like activity in biological tests of seed germination, transpiration and stomatal closure, inhibit gibberellin inducible amylase synthesis. Although these chemicals also inhibit photosynthetic electron transport, the structure activity relationships in inhibiting amylase synthesis was different from that in the inhibition of photosynthetic electron transport. This implies that the action mechanism of these chemicals is different in the two processes. Northern blot analysis showed that the most potent compound, 1,3,5-trihydroxy-4-nitro-2-phenylacetylresorcinol (9), inhibits the expression of agr;-amylase gene, pHv19, as does ABA. These results suggest that compound 9 may act in the same way as ABA or an inhibitor of the signal transduction pathway in barley aleurone cells.     

Spectral reflectance ratio of rice canopy for estimating crop nitrogen status

A portable meter for measuring the intensity of the green color of a rice canopy in the field by using spectral reflectance at 550 and 800 nm was newly devised. The measurements were found to be affected by reflection of sunshine on the leaf surface. With such limitations being taken into account, rice canopy green color intensity could be evaluated with this meter.     

Natural 15N abundance in shrub and tree legumes,Casuarina, and non N2 fixing plants in Thailand

The leaves and nodules from the shrub and tree legumes, particularly, Aeschynomene spp., Sesbania spp., Mimosa spp. and Leucaena spp., and Casuarina spp. and the leaves from neighbouring non-fixing plants were analyzed for their natural abundances of ¹⁵N ( ¹⁵N).The ¹⁵N in the leaves of non-fixing plants was +5.9% on average, whereas those from shrub legumes and Casuarina spp. were lower and close to the values of atmospheric N₂, suggesting the large contribution of N₂ fixation as the N source in these plants. The ¹⁵N values of the leaves from tree legumes except for Leucaena spp. were between the shrub legumes and non-fixing plants, which suggests that the fractional contribution of fixed N₂ in tree legumes may be smaller than that in the shrub legumes. Casuarina spp. was highly dependent on N₂ fixation. The ¹⁵N values of the nodules from most of the shrub legumes investigated were higher than those of the leaves.