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Showing papers on "Urea published in 1998"


Journal ArticleDOI
TL;DR: Intracellular urease activity is regulated by external pH, defends against gastric acidity by increasing periplasmic pH and membrane potential, and stimulates protein synthesis at acidic pH.

270 citations


Journal ArticleDOI
01 May 1998-Proteins
TL;DR: The data support a model in which urea denatures proteins by decreasing the hydrophobic effect and by directly binding to the amide units via hydrogen bonds, and indicate that the enthalpy of amide hydrogen bond formation in water is considerably higher than previously estimated.
Abstract: The effects of urea on protein stability have been studied using a model system in which we have determined the energetics of dissolution of a homologous series of cyclic dipeptides into aqueous urea solutions of varying concentration at 25 degrees C using calorimetry. The data support a model in which urea denatures proteins by decreasing the hydrophobic effect and by directly binding to the amide units via hydrogen bonds. The data indicate also that the enthalpy of amide hydrogen bond formation in water is considerably higher than previously estimated. Previous estimates included the contribution of hydrophobic transfer of the alpha-carbon resulting in an overestimate of the binding between urea and the amide unit of the backbone and an underestimate of the binding enthalpy.

192 citations


Journal ArticleDOI
TL;DR: It is concluded that selective alterations in IMCD transport occur in hypercalcemia, permitting the body to dispose of excess calcium without forming calcium-containing renal stones.
Abstract: To investigate how hypercalcemia blunts renal concentrating ability, alterations in basal and arginine vasopressin (AVP)-elicited osmotic water (P f) and urea (P urea) permeabilities were measured ...

143 citations


Journal ArticleDOI
TL;DR: The metabolic range of acid-adapted H. pylori is between an environmental pH of 3.5 and 8.6 and is an acid-tolerant neutralophile due to internal urease activity, which allows metabolism in the pH range between 4.0 and 2.0.

135 citations


Journal ArticleDOI
TL;DR: The hydrophobic sites in α-crystallin were evaluated using a fluorescent probe 1,1′-bi(4-anilino)naphthalenesulfonic acid (bis-ANS) and photolysis of bis-ANS resulted in incorporation of the probe to both A- and B-subunits, indicating that both subunits in native α- Crystallin contribute to the surface hydrophobicity of the protein.

135 citations


Journal ArticleDOI
TL;DR: Results are consistent with TMAO counteraction of the effects of urea on LDH kinetic parameters, supporting the premise that counteraction is a property of the solvent system and is independent of the evolutionary history of the protein.

134 citations


Journal ArticleDOI
TL;DR: The effects of inorganic N and organic manure, applied to a loamy arable soil, on CH4 oxidation were investigated in laboratory incubation experiments as discussed by the authors, showing that NH4Cl, (NH4)2SO4, and urea treatments were more effective in reducing CH4 consumption.
Abstract: The effects of inorganic N and organic manure, applied to a loamy arable soil, on CH4 oxidation were investigated in laboratory incubation experiments. Applications (40 mg N kg–1) of NH4Cl, (NH4)2SO4, and urea caused strong instantaneous inhibition of CH4 oxidation by 96%, 80%, and 84%, respectively. After nitrification of the added N the inhibitory effect was not fully reversible, resulting in an residual inhibition of 21%, 16%, and 25% in the NH4Cl, (NH4)2SO4, and urea treatments, respectively. With large NH4 + applications [240 mg N kg–1 as (NH4)2SO4] the residual inhibition was as high as 64%. Exogenous NO2 – (40 mg NO2 –-N kg–1) initially inhibited CH4 oxidation by 84%, decreasing to 41% after its oxidation. Therefore, applied NO2 – was a more effective inhibitor of CH4 consumption than NH4 +. Temporary accumulation of NO2 – during nitrification of added N was small (maximum: 1.9 mg NO2 –-N kg–1) and thus of minor importance with respect to the persistent inhibition after NH4 + or urea application. CH4 oxidation after NaNO3 (40 mg N kg–1) and NaCl addition did not differ to that of the untreated soil. The effect of organic manures on CH4 oxidation depended on their C/N ratio: fresh sugar beet leaves enhanced mineralization, which caused an instantaneous 20% inhibition, whereas after wheat straw application available soil N was rapidly immobilized and no effect on CH4 oxidation was found. The 28% increase in CH4 oxidation after biowaste compost application was not related to its C/N ratio and was probably the result of an inoculation with methanotrophic bacteria. Only with high NH4 + application rates (240 mg N kg–1) could the persistent inhibitory effect partly be attributed to a pH decrease during nitrification. The exact reason for the observed persistent inhibition after a single, moderate NH4 + or urea application is still unknown and merits further study.

119 citations


Journal ArticleDOI
Antonello Cannas1, A. Pes, R. Mancuso, B. Vodret, Anna Nudda1 
TL;DR: It is suggested that milk or blood urea N concentrations can be used as indicators of protein metabolism and intake of lactating ewes and less closely correlated with dietary CP intake.

114 citations


Journal ArticleDOI
TL;DR: Hyperosmolar NaCl and urea activate independent and complementary cellular programs that confer enhanced osmotolerance to renal medullary epithelial cells, which is not associated with a restoration of normal biosynthetic rates or cell cycle progression.
Abstract: Physiological adaptation to the hyperosmolar milieu of the renal medulla involves a complex series of signaling and gene expression events in which NaCl and urea activate different cellular processes. In the present study, we evaluated the effects of NaCl and urea, individually and in combination, on the viability of murine inner medullary collecting duct (mIMCD3) cells. Exposure to hyperosmolar NaCl or urea caused comparable dose- and time-dependent decreases in cell viability, such that 700 mosmol/kgH2O killed >90% of the cells within 24 h. In both cases, cell death was an apoptotic event. For NaCl, loss of viability at 24 h paralleled decreases in RNA and protein synthesis at 4h, whereas lethal doses of urea had little or no effect on these biosynthetic processes. Cell cycle analysis showed both solutes caused a slowing of the G2/M phase. Surprisingly, cells exposed to a combination of NaCl + urea were significantly more osmotolerant such that 40% survived 900 mosmol/kgH2O. Madin-Darby canine kidney cells but not human umbilical vein endothelial cells also exhibited a similar osmotolerance response. Enhanced survival was not associated with a restoration of normal biosynthetic rates or cell cycle progression. However, the combination of NaCl + urea resulted in a shift in Hsp70 expression that appeared to correlate with survival. In conclusion, hyperosmolar NaCl and urea activate independent and complementary cellular programs that confer enhanced osmotolerance to renal medullary epithelial cells.

113 citations


Journal ArticleDOI
TL;DR: Because of the large variation among and within cows, the monitoring of protein utilization of an individual cow was inaccurate, however, milk urea concentration in bulk milk is a valuable tool to monitor the rumendegraded protein balance in the ration.

107 citations


Journal ArticleDOI
01 May 1998-Fuel
TL;DR: In this paper, activated carbons were prepared by carbonization and steam activation of nitrogen-enriched low-rank coals by reaction of ammonia or its derivatives with the carboxyl groups either naturally occurring in coal or artificially introduced by performic oxidation.

Journal ArticleDOI
TL;DR: In this paper, a polyaniline−Nafion composite electrode was used to detect ammonium ion in urea biosensors, achieving a detection limit as low as 0.5 μM and a response time as short as 40 s.
Abstract: A new procedure for urea determination was developed. By cross-linking urease onto a polyaniline−Nafion composite electrode which sensed the ammonium ion effectively, a very sensitive urea biosensor was formed. The effects of applied potential, pH of buffer solutions, flow rate of carrier, and possible interferences on the response of urea biosensor were studied. With the developed urea biosensor, a detection limit as low as 0.5 μM and a response time as short as 40 s were obtained in an flow injection analysis system. A relative standard deviation of 2.2% (n = 15) was obtained for the successive analysis of a 0.03 mM standard urea solution. Applicability of the urea biosensor for urea analysis was demonstrated by the analysis of NIST standard reference material and urine samples.

Journal ArticleDOI
TL;DR: The results suggest that the quality of soy protein is inferior to that of casein protein.
Abstract: For routine evaluation of the quality of dietary protein, amino acid scoring patterns were used. Evaluation of this pattern for soy and casein revealed that these proteins are of almost equal quality. However, in vivo studies showed a large difference. To study the biological effects of meals with casein and soy protein, the contributions of individual amino acids to net protein retention and amino acid kinetics in gut, liver and muscle in healthy pigs were investigated. Isonitrogenous enteral nutrition, infused at a rate of 10 mL. kg body wt-1. h-1 and consisting of maltodextrin (137 g/L) with added casein (53 g/L) or soy protein (68 g/L), was given to conscious, healthy female multicathetized pigs (20-22 kg, n = 12). A primed-constant infusion protocol with L-[ring-2,6-3H]phenylalanine, L-[3,4-3H]valine and [15N-15N]urea was used to measure amino acid and urea kinetics in gut, liver and muscle. Measurements were done postabsorptively and 2-6 h after initiation of the enteral nutrition. During the meal, appearance of amino acids into the portal vein and the uptake by the liver was lower with casein infusion. Muscle uptake did not differ. Gut protein synthesis tended to be lower with soy infusion (P = 0.1). Liver protein synthesis and degradation were higher with casein infusion (P < 0.05), while in muscle, soy infusion stimulated protein turnover (P < 0.05). In comparison to the postabsorptive condition, liver urea production was unchanged after casein infusion, while it was significantly increased after soy infusion. These results suggest that the quality of soy protein is inferior to that of casein protein.

Journal ArticleDOI
TL;DR: Simultaneous TG-MS measurement, a mass spectrometer connected with a thermogravimetry by a transfer line and an interface or by the skimmer coupling, was used to investigate decomposition of urea as mentioned in this paper.
Abstract: Simultaneous TG-MS (TG: thermogravimetry, MS: mass spectrometry) measurement, a mass spectrometer connected with a thermogravimetry by a transfer line and an interface or by the skimmer coupling, was used to investigate decomposition of urea.A simple decomposition reaction of urea to form ammonia and cyanic acid (or isocyanic acid) was determined. The low intensity ratio of m/z 43 to m/z 17 was attributed to that the cyanic acid react with water to form carbon dioxide.Polymerization and decomposition of the polymers were more predominant later than the simple decomposition after the first stage.

Journal ArticleDOI
TL;DR: In this paper, the measurement of urea (CO(NH2)2) in rainwater samples from predominantly marine-influenced locations in Bermuda, and Ireland, and in rains and aqueous aerosol extracts from a rural site at UEA, Norwich indicates that urea is not generally a major contributor to atmospheric water-soluble organic nitrogen.

Journal ArticleDOI
TL;DR: The present model predicted that 95% of the label movements across the GIT could be accommodated by three or fewer entries and returns of urea-N and 99% by recycling for a maximum of six occasions, and offered a relatively simple means of estimating anabolic reuse of Urea by digestive tract micro-organisms.
Abstract: The effect of intake on urea production, entry into the digestive tract and return of N to the ornithine cycle was studied in four sheep. Each sheep received 0–6, 1–2 and 1.8 × estimated maintenance energy intake quantities of grass pellets for 9 d. After 4 d of adjustment, N balance measurements were conducted between days 5 and 8. From day 7 to day 9 animals were continuously infused, via the jugular vein, with [15N15N]urea and three urine samples were collected at approximately 2h intervals 48–54h after the start of infusion. Total urea and enrichments of [15N15N]- and [14N15N]urea in the urine samples were determined. Urea production was calculated from the isotopic dilution of [15N15N]urea and entry into the gastrointestinal tract (GIT) obtained from the difference between this and urinary urea elimination. Urea which enters the GIT undergoes hydrolysis to liberate NH3 which may be reabsorbed and enter the ornithine cycle, in which case the product is [14N15N]urea, based on the probabilities of labelled and unlabelled N providing ureagenic precursors. The quantity of urea-N which returns to the ornithine cycle from the GIT can thus be calculated. Existing models based on this approach yield overestimates of the fate of individual urea molecules due to a failure to allow for multiple recycling of [14N15N]urea species through the GIT. Refinements introduced to cover this resulted in a 33–48 % reduction in calculated return of label for the current study. The present model also predicted that 95 % of the label movements across the GIT could be accommodated by three or fewer entries and returns of urea-N and 99 % by recycling for a maximum of six occasions. Urea-N production increased with intake (P < 0.001) and exceeded digestible N values at all intakes. Urea which entered the digestive tract, both in absolute terms (P < 0.001) and as a proportion of production (0.62, 0.69, 0.73; P = 0.027), increased with intake. The proportion of entry into the digestive tract which was returned to the ornithine cycle remained reasonably constant (0.37–0.41) across all intakes but the absolute amount increased (5.6, 9.2 and 15.0gN/d; P < 0.001) with intake. If allowance is included for losses of 15N in faeces then the approach offers a relatively simple means of estimating anabolic reuse of urea by digestive tract micro-organisms and can complement data obtained from the technically more demanding arterio-venous and multiple-isotope techniques used hitherto.Urea: Gastrointestinal tract: [15N]kinetics: Sheep

Journal ArticleDOI
TL;DR: Results indicate that UT3 functions as a urea/water channel utilizing a common aqueous pathway and the water transporting function and low urea reflection coefficient of UT3 in vasa recta may be important for the formation of a concentrated urine by countercurrent exchange in the kidney.

Journal ArticleDOI
01 Sep 1998-Talanta
TL;DR: The biosensor with best response characteristics, obtained by procedure (B), showed the following characteristics of response to urea: linearity in the 10(-4) to 10(-2) M range; slope of up to 56 mV per decade; and lifetime of 2 months.

Journal ArticleDOI
10 Apr 1998-Virology
TL;DR: This system offers not only a model for investigation of the intrinsic interactions that occur during L1 assembly, but also a potential route for convenient manufacture of highly purified VLP vaccines.

Journal ArticleDOI
TL;DR: It is concluded that the dogfish apical membrane is extremely tight to urea, but the low apparent branchial permeability may also relate to the presence of an active urea transporter on the basolateral membrane that returns urea to the blood and hence reduces the apical urea gradient.
Abstract: We used a perfused gill preparation from dogfish to investigate the origin of low branchial permeability to urea. Urea permeability (C-14 urea) was measured simultaneously with diffusional water pe ...

Patent
18 Dec 1998
TL;DR: In this paper, a cyclic urea prepolymer comprising urea, formaldehyde, and ammonia or a primary amine is used to obtain a useful binder for a phenol-formaldehyde or melamine-formaline-based resin.
Abstract: The preparation of phenol-formaldehyde and melamine-formaldehyde resin-based binders extended with a cyclic urea-formaldehyde prepolymer and to products prepared using the binders. More particularly, the invention relates to a cyclic urea prepolymer comprising urea, formaldehyde, and ammonia or a primary amine which, when added to a phenol-formaldehyde or melamine-formaldehyde based resin, results in a useful binder for the manufacturer numerous articles.

Journal ArticleDOI
TL;DR: The results suggest that biogeochemical cycling in natural waters is influenced by sunlight, due to changes of microbially available components that were not reported previously, including amino acids, carbohydrates, nitrate and urea.

Journal ArticleDOI
TL;DR: In this article, the authors investigated the performance of metallophthalocyanine (M-Pc) catalysts for the simultaneous reduction of carbon dioxide and nitrite ions.

Journal ArticleDOI
TL;DR: It is concluded that cytoplasmic urease activity alone is not sufficient to allow survival of H. pylori in an acid environment and that the activity of surface-localized ure enzyme is essential for resistance ofH.
Abstract: Helicobacter pylori, an important etiologic agent in a variety of gastroduodenal diseases, produces large amounts of urease as an essential colonization factor. We have demonstrated previously that urease is located within the cytoplasm and on the surface of H. pylori both in vivo and in stationary-phase culture. The purpose of the present study was to assess the relative contributions of cytoplasmic and surface-localized urease to the ability of H. pylori to survive exposure to acid in the presence of urea. Toward this end, we compared the acid resistance in vitro of H. pylori cells which possessed only cytoplasmic urease to that of bacteria which possessed both cytoplasmic and surface-localized or extracellular urease. Bacteria with only cytoplasmic urease activity were generated by using freshly subcultured bacteria or by treating repeatedly subcultured H. pylori with flurofamide (1 μM), a potent, but poorly diffusible urease inhibitor. H. pylori with cytoplasmic and surface-located urease activity survived in an acid environment when 5 mM urea was present. In contrast, H. pylori with only cytoplasmic urease shows significantly reduced survival when exposed to acid in the presence of 5 mM urea. Similarly, Escherichia coli SE5000 expressing H. pylori urease and the Ni2+ transport protein NixA, which expresses cytoplasmic urease activity at levels similar to those in wild-type H. pylori, survived minimally when exposed to acid in the presence of 5 to 50 mM urea. We conclude that cytoplasmic urease activity alone is not sufficient (although cytoplasmic urease activity is likely to be necessary) to allow survival of H. pylori in acid; the activity of surface-localized urease is essential for resistance of H. pylori to acid under the assay conditions used. Therefore, the mechanism whereby urease becomes associated with the surface of H. pylori, which involves release of the enzyme from bacteria due to autolysis followed by adsorption of the enzyme to the surface of intact bacteria (“altruistic autolysis”), is essential for survival of H. pylori in an acid environment. The ability of H. pylori to survive exposure to low pH is likely to depend on a combination of both cytoplasmic and surface-associated urease activities.

Journal ArticleDOI
TL;DR: Low pH and urea are able to stimulate KCl loss from sickle cells, leading to cellular dehydration, even in regions of low PO2, significant for the pathophysiology of sickle cell disease.
Abstract: 1 K+ influx and efflux were measured in normal (HbA) and sickle (HbS) red blood cells to investigate the interaction of swelling, H+ ions and urea with O2 (0 to 150 mmHg O2) in the presence of ouabain and bumetanide (both 100 μM). 2 In HbA cells, K+-Cl− cotransport was O2 dependent. At low oxygen tensions (PO2s) the transporter was inactive and refractory to low pH, swelling or urea. 3 Cl−-independent K+ influxes in sickle cells were elevated at low PO2s, as previously reported. Cl−-dependent K+ influxes were large at both high and low PO2s, whether stimulated by swelling, H+ ions or urea. In the absence of O2, Cl−-dependent K+ influxes were similar in magnitude to those measured at high PO2s. The minimum for Cl−-dependent K+ influx was observed at PO2s of about 40-70 mmHg. 4 K+ efflux from HbS cells was stimulated by the addition of urea (500 mM). The rate constants were of similar magnitude whether measured at high PO2 or in the absence of O2, and were predominantly Cl− dependent under both conditions. 5 In HbS red blood cells, reduction of extracellular Ca2+, addition of 1 mM Mg2+ or nitrendipine (10 μM) to the saline had no effect. Inhibitors of K+-Cl− cotransport, [(dihydroindenyl)oxy] alkanoic acid (DIOA; 100 μM) or calyculin A (0·1 μM), inhibited influxes by a similar magnitude to Cl− substitution. 6 Results are significant for the pathophysiology of sickle cell disease. Low pH and urea are able to stimulate KCl loss from sickle cells, leading to cellular dehydration, even in regions of low PO2.

Journal ArticleDOI
TL;DR: Molecular studies have revealed that a specialised urea transport protein is present in kidney tissue of elasmobranchs, similar to the facilitated urea transporter found in the mammalian inner medulla of the kidney.
Abstract: Teleosts appear to have retained the genes for the urea cycle enzymes. A few species express the full complement of enzymes and are ureotelic (e.g., Lake Magadi tilapia) or ammoniotelic (e.g., largemouth bass), whereas most species have low or non-detectable enzyme activities in liver tissue and excrete little urea (e.g., adult rainbow trout). It was surprising, therefore, to find the expression of four urea cycle enzymes during early life stages of rainbow trout. The urea cycle may play a role in ammonia detoxification during a critical time of development. Exposure to alkaline water (pH 9.0-9.5) or NH4Cl (0.2 mmol/l) increased urea excretion by several-fold in trout embryos, free embryos and alevin. Urea transport is either by passive simple diffusion or via carried-mediated transport proteins. Molecular studies have revealed that a specialised urea transport protein is present in kidney tissue of elasmobranchs, similar to the facilitated urea transporter found in the mammalian inner medulla of the kidney.

Journal ArticleDOI
TL;DR: The urea cycle is essentially the simultaneous operation of two linear pathways, both primitive and widespread among animals; one is for arginine synthesis and the other is forArginine degradation to ornithine and urea.
Abstract: 1. The urea cycle is essentially the simultaneous operation of two linear pathways, both primitive and widespread among animals; one is for arginine synthesis and the other is for arginine degradation to ornithine and urea. 2. All animals may have the genetic capacity to express a urea cycle and many diverse groups of animals, from flatworms to mammals, have a functional urea cycle. 3. Evolutionary changes in vertebrates of carbamylphosphate synthetase (CPS) are directed from glutamine-dependent (CPSIII) towards NH3-dependent (CPSI) ureagenesis. Invertebrates, cartilagenous fish and the coelacanth have CPSIII (i.e. glutamine-dependent), whereas lungfish, amphibians and amniote vertebrates have CPSI; the teleost Heteropneustes has CPSI-like activity. That the coelacanth has CPSIII and Heteropneustes has 'CPSI' suggests that the form of CPS may by physiologically related (CPSIII in a balancing solute role and CPSI in a terrestrial, air-breathing excretion role) rather than being phylogenetically constrained. 4. Urea is a major balancing osmolyte in marine cartilagenous fish, the coelacanth and a few amphibians and some aestivating terrestrial amphibians. It is a storage osmolyte in cocoon-forming aestivating lungfish and amphibians. 5. Urea contributes towards positive buoyancy in marine cartilagenous fish. 6. Urea functions for non-toxic N transport in ruminant and pseudoruminant mammals. 7. Urea is a major solute in the mammalian (but not avian) kidney, contributing to a renal medullary osmotic gradient; it is substantially reabsorbed by mammalian nephrons. 8. Urea is used as a preferred nitrogenous waste compared with ammonia at high ambient pNH3 or pH, with water restriction, or air breathing. 9. Urea synthesis maintains acid-base balance by the 1:1 stoichiometry of removal of HCO3- and NH4+.

Journal ArticleDOI
TL;DR: In this article, a light-addressable potentiometric sensor for detection of glucose, penicillin and urea as model compounds is described, and the sensitivity of the glucose sensor was improved by the presence of hexacyanoferrate(III) as an alternative to O 2, which reacts with the coenzyme of glucose oxidase.

Patent
02 Jun 1998
TL;DR: In this paper, a method and apparatus for calculating the mass of a composition in a fluid volume or efficiency of exchange of said composition with an exchange fluid, especially urea in the body of a dialysis patient is presented.
Abstract: A method and apparatus for calculating the mass of a composition in a fluid volume or efficiency of exchange of said composition with an exchange fluid, especially urea in the body of a dialysis patient. Calculations are based on total mass of urea in the body. The concentration cd of urea in the effluent dialysate is measured, and the total removed mass U of urea is calculated by integrating the product of the urea concentration cd and the dialysate flow Qd. The momentary relative efficiency of the removal (K/V) is determined essentially by calculating the slope of the logarithm of the concentration curve and the momentary mass is determined therefrom. Then, the pre-treatment mass of urea in the body can be determined very accurately. Moreover, the momentary relative efficiency in any point is determined by using the removed urea U. The dialysis dose is calculated by integrating the momentary efficiencies.

Journal ArticleDOI
TL;DR: Urea-grown Ni-deprived plants showed reduced growth and accumulated large amounts of urea owing to the lack of urease activity, which suggests that the Ni requirement strongly depends on the N source employed.
Abstract: Nickel is considered to be an essential micronutrient in plants because of its role in the metalloenzyme urease. In order to characterize the metabolic consequences of Ni deprivation, the significance of Ni supply for growth and N metabolism of rice plants grown with either NH 4 NO 3 or urea as sole N source was evaluated. Growth of plants receiving NH 4 NO 3 was not affected by the Ni status, and neither were the activities of arginase and glutamine synthetase. However, urease activity was not detectable in leaves of low-Ni plants, which in conjunction with arginase action, led to the accumulation of urea in plants grown with NH 4 NO 3 . Amino acid contents and mineral nutrient status (except Ni) were not affected by the Ni treatment. Urea-grown Ni-deprived plants showed reduced growth and accumulated large amounts of urea owing to the lack of urease activity. These plants were further characterized by low amino acid contents indicating impaired usage of the N supplied. They also exhibited reduced levels of the urea precursor arginine, which is merely attributed to the overall N economy in these plant. When urea-grown plants were supplied with 0.5 mmol m -3 Ni in the nutrient solution, the dry weight and the amino acid N contents were increased at the expense of the urea contents, indicating efficient use of urea N in Ni-supplemented plants. A critical Ni concentration in the shoot regarding dry matter production of NH 4 NO 3 -grown plants could not be deduced, while 25 μg Ni kg -1 DW is certainly inadequate for urea-grown plants. This suggests that the Ni requirement strongly depends on the N source employed.