Metabolism of nitrate and ammonium in seedlings of Norway spruce (Picea abies) measured by in vivo ¹⁴N and ¹⁵N NMR spectroscopy

Abstract

S. Metabolism of nitrate and anunonium in seedlings of Norway spruce (Picea ahies) measured by in vivo "N and ' In vivo "N and '*N nuclear magnetic resonance spectroscopy was used to investigate the assimilation of nitrate and ammonium in seedlings of Norway spruce (Picea ahies [L.] Karst,), The main objective was to study accumulation of free NHJ and examine to what extent the nitrogen source affects the composition of the free amino acid pools in roots, stems and needles, NHJ concentrations in plants growing in the presence of 0,5-50 mM ammonium were quantified using ' "^N NMR. The NH:I values in tissues ranged from 6 to 46;(mol (g fresh weight)'', with highest concentrations in roots and needles. The tissue NHl peaked at 5,0 mAf NHJ in the medium, and failed to increase when NH4 in the medium was increased to 50 mM, indicating metabolic control of the concentration of this cation in tissues. The ^""N NMR spectra were used to estimate pH of the NH4 storage pools. Based on the pH sensitivity of the quintet of "NHJ resonance, we suggest that the pH of the ammonium storage compartments in the roots and stems should be 3,7-3.8, and in needles 3.4-3.5, representing extremely low pH values of the tissue, "N from nitrate or ammonium was first incorporated into the amide group of glutamine and then into a-amino groups, confirming that the glutamine synthetase/ glutamate synthase cycle is the major route of nitrogen assimilation into amino acids and thus plays a role in lowering the levels of NH* in the eytoplasm. NHJ can also be assimilated in roots in piants growing in darkness. The main "N-labelled amino acids were glutamine, arginine and alanine. Almost no ''N signals from needles were observed. Double labelling (dN + (o,a>N) of arginine is consistent with the operation of the omithine cycle, and enrichment indicates that this cycle is a major sink of newly assimilated nitrogen. Nitrogen assimilation in roots in the presence of added methionine sulphoximine and glutamate indicated the catabolic action of glutamate dehydrogenase. The ' •'N NMR spectra of plants grown on '^N-urea showed a marked increase in the labelling of ammonium and glutamine, indicating high urease activity. Amino acids were also quantified using high pressure liquid chromatography. Arginine was found to be an important transport form cf nitrogen in the stem.

Rapid 15N uptake and metabolism in fine roots of Norway spruce

“…In the present study, rather low concentrations of Gln and Asp were measured in the roots, in contrast to Glu, while other studies have described Gln as one of the main amino acid in the roots of Norway spruce seedlings (Chalot et al 1991;Aarnes et al 1995;Gessler et al 1998). The low Gln concentrations could be due to N depletion in our laboratory study, as no N was applied before the tracer application.…”

“…The differences between the at% 15 N enrichment values in roots and shoots demonstrated a dilution effect already described by Aarnes et al (1995) in white spruce. The amino acid concentrations increased after the nitrate fertilisation mainly in the shoot of the Norway spruce seedlings, while concentrations in the roots remained un-changed.…”

“…Previous studies of short-term ammonium 15 N labelling of mycorrhizal eucalypt (Turnbull et al 1995) or spruce (Chalot et al 1991;Aarnes et al 1995) seedlings reported the greatest flow of 15 N via the amide group of Gln, with a label also in the amino group of Gln, Glu, Ala and GABA. While the latter was not measured in the present study, the high enrichment patterns of Gln, Glu and Ala were confirmed in non-mycorrhizal spruce seedlings after 4 h to 1 day.…”

Regulation of nitrate uptake at the whole-tree level: interaction between nitrogen compounds, cytokinins and carbon metabolism

“…In annual and perennial herbaceous plants, signaling of N status plays an important role during particular developmental stages, e.g., during seed production (Imsande and Touraine 1994). Regulation of N nutrition is more complex in trees than in herbaceous plants and, as a result, an integrated signaling system would fulfil several functions (Gezelius and Näsholm 1993, Aarnes et al 1995, Fotelli et al 2002. Because of the long life span of trees in a changing environment, there is an advantage in reducing the dependency of tree mineral nutrition on current mineral supply.…”

Retrospective chemical analysis of tree rings by means of the scanning electrochemical microscopy with shear force feedback

“…The local determination of the content of chemically bound nitrogen in tree rings of wood samples is from large interest in the analysis of growth influences. 3,4 By means of a chemical analysis of tree rings, it is possible to make quantitative statements about the amount of nitrogen in the wood and its distribution as function of the respective growth year of the examined tree. These data contribute to a better knowledge about the nutrient supply in trees and thus indirectly about the chemical environmental conditions at the time of the growth of the respective tree ring.…”