Determination of urea and ammonia in leaf extracts: application to ureide metabolism

1985 ◽  
Vol 63 (6) ◽  
pp. 1135-1140 ◽  
Author(s):  
Barry J. Shelp ◽  
Konrad Sieciechowicz ◽  
Robert J. Ireland ◽  
Kenneth W. Joy
Keyword(s):  
Leaf Tissue ◽  
Exchange Chromatography ◽  
Methionine Sulfoximine ◽  
Plant Tissues ◽  
Leaf Extracts ◽  
Reliable Technique ◽  
Urea Metabolism ◽  
Soybean Leaves ◽  
Production Of Ammonia

Methods are described for the routine determination of urea and ammonia in plant tissues. Ureido compounds such as allantoin and allantoate interfere with the urea assay but can be conveniently removed by ion-exchange chromatography prior to the color reaction with α-isonitrosopropiophenone. This assay was used to examine the effect of the urease inhibitor acetohydroxamate on leaf urea levels. A survey of methods for ammonia determination showed that for plant extracts and assay mixtures containing amides, microdiffusion followed by nesslerization provided a convenient and reliable technique. This procedure was used to determine asparaginase and glutaminase activities as well as ammonia levels in leaf tissue. In a study of detached soybean leaves, 5 mM acetohydroxamate, supplied via the transpiration stream, inhibited the production of ammonia normally observed in the presence of methionine sulfoximine (a glutamine synthetase inhibitor) and caused an accumulation of urea. The data were interpreted as evidence that leaf ammonia can arise from urea metabolism and supported a role for urea, probably derived from ureide breakdown, in the provision of nitrogen for amino acid and protein synthesis.

Accumulation of germanium (Ge) in plant tissues of grasses is not solely driven by its incorporation in phytoliths

2020 ◽  
Author(s):  
Sabine Kaiser ◽  
Christin Moschner ◽  
Oliver Wiche
Keyword(s):  
Leaf Tissue ◽  
Grass Species ◽  
Control Element ◽  
Plant Tissues ◽  
Plant Parts ◽  
Icp Ms ◽  
Dry Ashing ◽  
Leaf Tissues ◽  
Canary Grass

<p>Until recently it has been generally assumed that Ge taken up by plants is stored in phytoliths together with Si. This assumption is mostly based on the geochemical similarities between Ge and Si, while a scientific proof was lacking. The aim of the present study is to i) compare the uptake of Si and Ge in three grass species, ii) localize Ge and Si stored in above-ground plant parts and iii) evaluate the amounts of Ge and Si sequestrated in phytoliths and plant tissues. Mays (<em>Zea mays</em>), oat (<em>Avana sativa</em>) and reed canary grass (<em>Phalaris arundinacea</em>) were cultivated in the greenhouse on soil and sand to control element supply. Leaf phytoliths were extracted by dry ashing. Total elemental composition of leaves, phytoliths, stems and roots were measured by ICP-MS. For the localization of phytoliths and the determination of Ge and Si within leaf tissues and phytoliths scanning electron microscopy (SEM), energy dispersive x-ray spectroscopy (EDX) and laser ablation ICP-MS (LA-ICP-MS) was used. The amounts of Si and Ge taken up by the species corresponded with biomass formation and decreased in the order <em>Z. mays </em>><em>P. arundinacea, A. sativa</em>. Results from LA-ICP-MS revealed that Si was mostly localizedin phytoliths, while Ge was disorderly distributed within the leaf tissue. In fact, from the total amounts of Ge accumulated in leaves only 10% was present in phytoliths highlighting the role of organic Ge species in plant tissues and the necessity for using bulk Ge/Si instead of Ge/Si in phytoliths to trace biogeochemical cycling of Si. Moreover, our results represent important background data for the optimization of a phytomining of Ge.</p>

Isoforms of glutamine synthetase in Panicum species having C3, C4, and intermediate photosynthetic pathways

1983 ◽  
Vol 61 (8) ◽  
pp. 2257-2259 ◽  
Author(s):  
B. Hirel ◽  
D. B. Layzell ◽  
B. McCashin ◽  
S. F. McNally ◽  
D. T. Canvin
Keyword(s):  
Glutamine Synthetase ◽  
Leaf Tissue ◽  
Relative Activity ◽  
Exchange Chromatography ◽  
Leaf Extracts ◽  
Photosynthetic System ◽  
Photosynthetic Pathways ◽  
System P ◽  
Single Genus ◽  
Second Peak

The number, relative activity, and immunological nature of glutamine synthetase (GS) isoforms were studied in three species of Panicum, each differing in the nature of their photosynthetic system: P. bisulcatum, a C3 plant; P. milioides, a C3–C4 intermediate; and P. miliaceum, a C4 plant. Ion-exchange chromatography of leaf extracts for all three species revealed a peak of GS activity eluting at 0.23–0.32 M NaCl. These peaks had similar elution characteristics to GS isolated from chloroplasts of barley. Also, as in the barley chloroplastic GS, they were poorly recognized by antibodies from barley cytosolic GS (GS1). These peaks were therefore identified as GS2 and were thought to be chloroplastic in origin. A second peak of GS activity eluting at 0.18 M NaCl was found to account for 0, 21, and 45% of the total GS activity in the C3, C3–C4 intermediate, and C4 leaves, respectively. It had similar elution characteristics to the cytosolic GS from barley and was readily bound by the antibodies made against barley cytosolic GS (GS1). This enzyme, identified as GS1, was thought to be cytosolic in origin. The results demonstrate that even within a single genus, the GS isoform composition is closely related to the type of photosynthetic pathway present within the leaf tissue.

scholarly journals Insight into Glutamatergic Involvement in Rewarding Effects of Mephedrone in Rats: In Vivo and Ex Vivo Study

2021 ◽  
Author(s):  
Olga Wronikowska ◽  
Maria Zykubek ◽  
Agnieszka Michalak ◽  
Anna Pankowska ◽  
Paulina Kozioł ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Interdisciplinary Approach ◽  
Nmda Antagonist ◽  
Exchange Chromatography ◽  
Resonance Spectroscopy ◽  
Monoamine Transporters ◽  
Ex Vivo Study ◽  
Insight Into

AbstractMephedrone is a widely used drug of abuse, exerting its effects by interacting with monoamine transporters. Although this mechanism has been widely studied heretofore, little is known about the involvement of glutamatergic transmission in mephedrone effects. In this study, we comprehensively evaluated glutamatergic involvement in rewarding effects of mephedrone using an interdisciplinary approach including (1) behavioural study on effects of memantine (non-selective NMDA antagonist) on expression of mephedrone-induced conditioned place preference (CPP) in rats; (2) evaluation of glutamate concentrations in the hippocampus of rats following 6 days of mephedrone administration, using in vivo magnetic resonance spectroscopy (MRS); and (3) determination of glutamate levels in the hippocampus of rats treated with mephedrone and subjected to MRS, using ion-exchange chromatography. In the presented research, we confirmed priorly reported mephedrone-induced rewarding effects in the CPP paradigm and showed that memantine (5 mg/kg) was able to reverse the expression of this effect. MRS study showed that subchronic mephedrone administration increased glutamate level in the hippocampus when measured in vivo 24 h (5 mg/kg, 10 mg/kg and 20 mg/kg) and 2 weeks (5 mg/kg and 20 mg/kg) after last injection. Ex vivo chromatographic analysis did not show significant changes in hippocampal glutamate concentrations; however, it showed similar results as obtained in the MRS study proving its validity. Taken together, the presented study provides new insight into glutamatergic involvement in rewarding properties of mephedrone.

scholarly journals An HPLC-automated Derivatization for Glutathione and Related Thiols Analysis in Brassica rapa L.

Agronomy ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 1157
Author(s):  
Francesco Nacca ◽  
Concetta Cozzolino ◽  
Petronia Carillo ◽  
Pasqualina Woodrow ◽  
Amodio Fuggi ◽  
...  
Keyword(s):  
Brassica Rapa ◽  
High Performance ◽  
Internal Standard ◽  
Plant Tissues ◽  
Cellular Functions ◽  
Sulphur Compounds ◽  
Stress Protection ◽  
Automated Method ◽  
Reduced Forms

The high content of glucosinolates and glutathione makes the Brassicaceae an important healthy food. Thiols and especially glutathione and γ-Glu-Cys-Gly tripeptide are involved in many fundamental cellular functions such as oxidative stress protection. Although several methods for sulphur compounds analysis in biological samples are actually used, the determination of glutathione and other sulphur derivatives in plant tissues is rather problematic due to their extreme susceptibility to oxidation, which can lead to their overestimation. The aim of this work was the improvement and validation of an automated method for determination of reduced and oxidised glutathione, cysteine and γ-glutamylcysteine in plant tissues. The method consists of a fully automated pre-column derivatization of thiols based on monobromobimane reagent, a high-performance liquid chromatography derivatives separation, and a fluorimetric detection and quantification. The method was successfully applied for determination of the oxidized and reduced forms of Cys, γ-GC and GSH content in leaves, petioles, inflorescences and roots of Brassica rapa L. subsp. Sylvestris. At harvest, in freshly cut plants, the average contents of GSH/2GSSG were 840/45, 345/70 and 150/70 nmol g−1 FW for the florets, leaf blades and stems, respectively; those of Cys/2Cys were 80/12, 29/12 and 24/6 nmol g-1 FW; while those of γ-GC/γ-GCCG-γ were 8.0/4.0, and 6.0/3.0, 3.0/2.0 nmol g−1 FW, respectively. Such amounts were lower in low-sulphur-grown plants at harvest. The very low coefficient of variation between repeated tests (maximum 1.6%), the high recovery of internal standard (>96%) and the linear correlation coefficient of the calibration (R2 > 0.99) support the efficiency of this method that allowed analysing about 50 samples/die in a totally automated manner with no operator intervention. Our results show that the reported method integrations can significantly improve thiols detection via HPLC.

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