Determination Of Urea Research Articles

Inflammation-associated responses in piglets induced with post-weaning colibacillosis are influenced by dietary protein level

Forty piglets (average body weight = 5.32 kg) were used to investigate the effect of dietary crude protein (CP) content on immunological responses following a challenge with enterotoxigenic Escherichia coli (ETEC) K88. Pigs, housed 4 per pen, were randomly allotted to 2 diets: 1) a high, 225 g/kg CP diet (HCP) or 2) a low, 176 g/kg CP diet (LCP) supplemented with crystalline amino acids. Pigs were orally challenged with 6 mL of an ETEC K88 suspension containing 10 10 cfu/mL on d 8 after weaning. Blood samples were collected from 10 pigs (1 pig/pen) on d 7 (at weaning), −24 h, 8 h, 72 h and 7 d after the challenge for determination of plasma urea N (PUN) and serum concentrations of tumor necrosis factor alpha (TNF-α) and interleukin 1 beta (IL-1β) and haptoglobin (Hp). Tumor necrosis factor alpha, IL-1β and Hp were measured as indicators of inflammatory responses. The concentrations of serum TNF-α at 8 h, 72 h and 7 d after challenge were similar to the level observed at 24 h before challenge but higher ( P < 0.05) than the weaning level. Pigs fed the LCP diet had lower ( P = 0.032) concentrations of IL-1β (72 vs. 116 pg/mL) at 8 h post-challenge compared with those fed the HCP diet. Likewise, pigs fed the LCP diet tended to have lower ( P = 0.088) concentration of Hp (9 vs. 25 mg/dL) compared with those fed the HCP diet at 8 h post-challenge. Compared with the weaning concentration, PUN concentration at 72 h after ETEC challenge was higher ( P < 0.05) in pigs fed the HCP diet. The results indicate that the LCP diet supplemented with crystalline amino acids reduced inflammatory responses, as indicated by serum IL-1β, in piglets infected with ETEC K88.

Second-Derivative Spectrometric Determination of Urea in Milk Using the Diacetyl Monoxime Reagent

The application of derivative spectrometry to resolve overlapping spectra and improve the sensitivity and selectivity of the colorimetric determination of urea in milk using diacetyl monoxime is presented. With first-derivative (1D) spectrometry, the lambda(max) of the colored complex was established to be 525 nm. The absorption band at lambda(max) = 525 nm in normal absorption (0D) spectrometry was resolved into three clearly distinct spectral bands with minima at 497, 530, and 566 nm with second-derivative (2D) spectrometry. With the second-derivative (2D530) technique, the depth of the trough of the strongest signal at 530 nm was used to determine urea in milk samples. Linear calibration curves for urea were obtained over a concentration range of 0.2-1.4 mg/L, with correlation coefficients greater than 0.999. The LOD and LOQ of the method were 0.01 and 0.03 mg/L, respectively. Recoveries of 99-100% of spiked urea in milk with RSD values below 2.0% were obtained. The second-derivative method is simple, affordable, sensitive, and selective for the determination of urea in milk.

Determination of Derivatized Urea in Exhaled Breath Condensate by LC-MS

Elevation in one or more compounds in exhaled breath condensate (EBC) has been reported to be related to one or another lung disease. The increased concentration might be caused by increased chemicals in the airway surface liquid. However, it might also be due to an increased delivery of liquid samples into the airstream. Being evenly distributed throughout the body, urea is a likely candidate for a marker of such dilution. A liquid chromatography-tandem mass spectrometry method was developed for determination of EBC urea. Urea in EBC samples was converted to 2-hydroxypyrimidine (2-HPM) through a one step reaction, along with (15)N(2)-urea added as an internal standard. The product ion m/z 97/42 was selected for quantification with m/z 99/43 from (15)N(2)-2-HPM as a standard. Concentrations of urea in EBC from five lung cancer patients were found to be 35.1, 2.2, 103.5, 19.3, and 3.6 microM, respectively. The highest values were in patients dying of respiratory distress, whose lungs were filled with fluid. Lower values were seen in patients whose conditions were improving. Lately, one of the low EBC urea values was observed in a patient whose airway status did not contribute to his poor clinical condition.

Array-based titanium dioxide biosensors for ratiometric determination of glucose, glutamate and urea

A novel optical array-based titanium dioxide (TiO 2) biosensor combining with simple vapor deposition method was developed to simultaneously determine multianlaytes including glucose, urea and glutamate in serum samples. Carboxy seminaphthorhodamine-1-dextran (SNARF-1-dextran), serving as a single-molecule fluorescent probe, was co-immobilized with different enzymes including urease, glucose dehydrogenase (GDH), and glutamate dehydrogenase (GLDH) for determination of urea, glucose and glutamate simultaneously. The SNARF-1-dextran was a useful pH sensitive dye and the fluorescence ratio at 585 and 630 nm was used to characterize the change in solution pH. Atomic force microscopic (AFM) and scanning electron microscopy (SEM) images showed that TiO 2 is a crack-free stable sol–gel material, which can effectively trap enzymes in the sol–gel-derived matrices prepared by vapor deposition method. The surface morphology of the sol–gel-derived TiO 2 films changed significantly after the immobilization of macromolecules and enzymes. The array-based TiO 2 biosensors show good analytical performance on the simultaneous determination of multiple samples for multianalytes without obvious cross-interference. The analytical ranges of the three analytes in water samples were between 0.01 and 10 mM and limit of detections (LODs) were in the range 2.4–5.5 μM. In addition, satisfactory dynamic ranges of 2–3 orders of magnitude with LODs of 3.1–7.8 μM in serum samples were obtained. These results demonstrate the first report of combining a sol–gel-derived TiO 2 array biosensor with SNARF-1-dextran for ratiometric determination of multianalytes simultaneously and provide a viable alternative to current silica technology for the fabrication of array-based optical biosensors.

Regulation of arginase pathway in response to wall shear stress

ObjectiveAlterations of wall shear stress can predispose the endothelium to the development of atherosclerotic plaques. Ample evidence indicates that arginase expression and/or activity correlates with several risk factors for cardiovascular disease including atherosclerosis. We investigated the regulation of arginase pathway in response to distinct patterns of wall shear stress. MethodsIsolated porcine endothelial cells and carotid arterial segments were perfused under unidirectional high shear stress (HSS) or oscillatory shear stress (OSS) for 1 and 3 days. Arginase I and II expression, cellular localization and enzyme activity were, respectively, assessed by Western blot, immunohistochemistry and colorimetric determination of urea. The contribution of arginase to the processes of endothelial dysfunction, cell proliferation and arterial remodeling induced by OSS was evaluated by administration of the arginase inhibitor N-ω-hydroxy-nor-l-arginine (nor-Noha). ResultsOnly arginase II isoform was detected on porcine carotid endothelial cells and on carotid artery. Exposure of arteries to OSS increased arginase II expression and activity as compared to HSS. Inhibition of arginase by nor-Noha improved NO-dependent endothelial function and decreased total vascular ROS formation in arteries submitted to OSS. In addition, inhibition of arginase activity decreased smooth muscle cell proliferation rate with no effect on collagen content after OSS. ConclusionsExposure of carotid artery to oscillatory flow induced a more pronounced activation of arginase as compared to HSS. Inhibition of arginase in arteries exposed to OSS improved NO-dependent endothelial function and decrease smooth muscle cell proliferation rate, both processes are important for the focal development of atherosclerotic plaque.

LTCC microreactor for urea determination in biological fluids

This article presents design, fabrication and testing of an enzymatic microreactor with integrated heater and a temperature sensor. The microsystem was fabricated using low temperature co-fired ceramics (LTCCs) technology, which has a few advantages over silicon-based fabrication processes. Computational fluid dynamics (CFD) and finite element method (FEM) analysis was used to arrive at an optimum microreactor and heater design. The heater performance was evaluated using thermographic measurements, and the temperature resistance characteristics of the temperature sensor revealed good electrical properties and stability at higher temperatures (up to 140°C). As an enzyme carrier, the glass beads coated with polyacrylonitrile layer and porous glass beads were used. The microreactor was used to measure urea concentration with high output signal (ca. 2.5 pH units) and large scale. The maximal output signal enabled to apply presented microreactor for determination of urea in biological or environmental fluids. The properties of the LTCC-based enzymatic microreactor were comparable with the features of a similar one made in silicon.

Protective effect of oleanolic acid on gentamicin induced nephrotoxicity in rats

Oleanolic acid is a molecule of current therapeutic interest. In the present study, oleanolic acid isolated from the cuticular epithelium of Viscum articulatum Burm. f. (Viscaceae) was investigated for its protective effects on gentamicin-induced renal damage in rats. Nephrotoxicity was induced in rats by intraperitoneal injection of gentamicin at a dose of 100 mg/kg/day for 8 days. The effect of Oleanolic acid administered orally at doses 40, 60 and 80 mg/kg/day was assessed biochemically by determination of albumin, urea and creatinine in serum and urine samples and also through histopathological examination of the kidneys. Oleanolic acid protected the rat kidneys from gentamicin-induced nephrotoxicity as evident from a decrease in the serum and urine levels of creatinine, albumin and urea. Oleanolic acid also protected the rat kidneys from histological alterations induced by gentamicin and also improved the glomerular filtration rate. Compared with an earlier report on intraperitoneal administration of oleanolic acid in paracetamol-induced nephrotoxicity in rats, the data show that orally administered oleanolic acid also exerted a nephroprotective effect even in the case of a nephrotoxicant such as gentamicin, which directly deteriorates the kidney function without prior metabolism.

Catalytic systems based on the organic nickel(ii) complexes in chronoamperometric determination of urea and creatinine

The organic nickel(ii) complexes with catalytic activity in the electrochemical oxidation of creatinine and urea were synthesized and studied. The signals of electrocatalytic oxidation of the studied carbonyl-containing amines in model solutions were obtained. The detection limit (by the 3σ-criterion) is 8.7·10−6 and 2.7·10−5 mol L−1 for urea and creatinine, respectively.

Quantitative determination of urea concentrations in cell culture medium

Urea is the major nitrogenous end product of protein metabolism in mammals. Here, we describe a quantitative, sensitive method for urea determination using a modified Jung reagent. This assay is specific for urea and is unaffected by ammonia, a common interferent in tissue and cell cultures. We demonstrate that this convenient colorimetric microplate-based, room temperature assay can be applied to determine urea synthesis in cell culture.

The Impact of Samh Seeds on Blood Parameters of Experimental Animals

Samh seeds are cultivated in some regions of KSA and some people think that it can improve health. We aimed to find out the health benefits of Samh seeds. The seeds were obtained from Al Jouf, KSA and roasted for 5 min and grounded into flour. Its content from protein, fat and carbohydrate were determined. Thirty six normal male albino rats (100±5.0 g) were classified into 6 groups; control group that fed standard diet only, 5% Samh group received 5% of the seeds; 15% Samh group fed 15% of the seeds; high lipid diet group received high lipid diet (15% fat and 1% cholesterol); 5% Samh and high lipid group received high lipid diet plus 5% of the seeds and 15% Samh and high lipid diet group received high lipid diet plus 15% of the seeds. After 60 days, rats were fasted then anesthetized and blood samples were collected for determination of glucose, blood lipids, urea, creatinine, albumin, total protein, billirubin, calcium and iron. The results showed that the seeds are rich in protein (22.3±2.1%) and the administration of 5% of the seeds decreased glucose and BUN, while it elevated creatinine, albumin, total protein, cholesterol, triglycerides, HDL, calcium and iron. On the other hand the administration of 15% of the seeds decreased creatinine, albumin, cholesterol, triglycerides and LDL, while increased glucose, BUN, total bilirubin, HDL and iron. In conclusion the administration of 15% of Samh seeds flour reduce creatinine concentration and improve blood lipids especially cholesterol, HDL and LDL.

Potentiometric biosensor for urea determination in milk

A potentiometric urea sensitive biosensor, using a NH 4 + ion sensitive electrode in double matrix membrane (DMM) technology as the transducer, has been described. Thick film screen printing technique was utilized for the fabrication of the basal conducting track of the potentiometric electrode and also for the development of Ag/AgCl reference electrode. The ion sensitive polymer matrix membrane could be formed in the presence of an electrochemically inert filter paper. The electrochemical response behavior of the NH 4 + ion sensitive electrode has been studied initially. Later, a urea biosensor has been developed by immobilizing the urease enzyme, through entrapping, onto the ion sensitive membrane using a polymer matrix of poly(carbamoylsulphonate) (PCS) and polyethyleneimine (PEI). The urea biosensor responded rapidly and in a stable manner to the changes in the test urea concentrations. The sensor exhibited a detection limit of 2.5 × 10 −5 mol/L. The average slope in the linear range has been found to be 51.7 ± 0.5 mV/decade. The biosensor system has, then, been tested for the detection of urea levels in the real samples of milk. A good agreement could be observed between the urea concentrations detected by the developed biosensor and the spectrophotometric technique.

Stabilization of Sensing Assay within Polyelectrolyte-Coated Alginate Microspheres for Optical Urea Sensing

A new absorbance-based enzymatic biosensor for determination of urea (in the range 0.01 to 6.7 mM) is described. Quantification using cresol red dye, immobilized in the nanofilm coatings assembled on alginate microspheres to immobilize the urease enzyme, has been accomplished using ratiometric absorbance measurements. The effect of salt concentration in polyelectrolyte nanofilms (on the stability of dye molecules) and buffer pH (on the enzyme stability) are reported. The results demonstrate excellent stability of sensing assay within alginate microspheres. Urea-sensing experiments demonstrate the potential to develop an optical urea sensor that is stable over a month.

An amperometric urea biosensor based on covalently immobilized urease on an electrode made of hyperbranched polyester functionalized gold nanoparticles

An amperometric biosensor was fabricated for the quantitative determination of urea in aqueous medium using hematein, a pH-sensitive natural dye. The urease (Urs) was covalently immobilized onto an electrode made of gold nanoparticles functionalized with hyperbranched polyester-Boltron ® H40 (H40–Au) coated onto an indium–tin oxide (ITO) covered glass substrate. The covalent linkage between the Urs enzyme and H40–Au nanoparticles provided the resulting enzyme electrode (Urs/H40–Au/ITO) with a high level of enzyme immobilization and excellent lifetime stability. The response studies were carried out as a function of urea concentration with amperometric and photometric measurements. The biosensor based on Urs/H40–Au/ITO as the working electrode showed a linear current response to the urea concentration ranging from 0.01 to 35 mM. The urea biosensor exhibited a sensitivity of 7.48 nA/mM with a response time of 3 s. The Michaelis–Menten constant for the Urs/H40–Au/ITO biosensor was calculated to be 0.96 mM, indicating the Urs enzyme immobilized on the electrode surface had a high affinity to urea.

Gold Nanoparticles as Assisted Matrices for the Detection of Biomolecules in a High-Salt Solution through Laser Desorption/Ionization Mass Spectrometry

Citrate-capped gold nanoparticles (AuNPs) serve as matrices for the determination of biomolecules in a high-salt solution through matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). In the case of using 2,5-dihydroxybenzoic acid (2,5-DHB) as a matrix, the signal intensities of neutral steroids were severely suppressed in a high-salt solution. A high concentration of NaCl caused the formation of the sodium adduct ions during the desorption/ionization process, resulting in a decrease of the signal intensities of the protonated ions. In comparison, by applying AuNP-assisted LDI-TOF-MS, the signal intensities of neutral steroids remained almost constant when the concentration of NaCl was increased to 500 mM. Because the use of citrate-capped AuNPs as matrices primarily offers alkali metal ion adducts, AuNP matrices have a higher tolerance to high NaCl concentrations relative to that of 2,5-DHB matrices. The relevant phenomena are also discovered in the case of analysis of neutral carbohydrate, monosialoganglioside, indolamine, and angiotensin I. The quantification of small molecules in a high-salt solution has been accomplished by AuNP-assisted LDI-TOF-MS coupled to a unique sample preparation, in which samples are deposited onto the sample plate before AuNPs. The present method has been further applied to the determination of urea, creatinine, uric acid, and glucose in a urine sample.

Long term cultures of primary human hepatocytes as an alternative to drug testing in animals

Due to species differences, primary human hepatocytes are still the in vitro system of choice to analyse liver specific processes and functions. Human hepatocytes were cultured for several weeks in a serum-free two-dimensional culture system, which was used to study the effects of acetaminophen (APAP) on hepatocellular functions and vitality. Non-invasive determinations of albumin, urea and lactate dehydrogenase concentrations in cell culture supernatants allowed continuous monitoring for at least two weeks. APAP was applied every 4 days for 24 h. Each application reduced urea production by 25% and albumin synthesis by approximately 70% without any effects on cellular viability. After removal of the substance, hepatocellular functions returned to control levels within one (urea) to three (albumin) days. The repetitive analyses of APAP-mediated effects on cellular metabolism led to identical results for up to five cycles. The drug also caused reversible and repetitive ultrastructural modifications, in particular an almost complete replacement of rough endoplasmic reticulum by smooth endoplasmic reticulum and a massive degradation of glycogen stores. The data demonstrate the suitability of the culture system to serve as a model for repetitive testing of drug-mediated changes on hepatocellular functions, thereby reducing animal studies during drug development.

Instructions to Authors

Live lobsters are considered a fish product by European laws but no invertebrates welfare reference exists in European legislation. In this work the effect of commercial storage stress considering two different commercial maintenance methods (air exposure vs seawater resubmission) were evaluated. Thirteen different physiological variables of the hemolymph were analyzed: glucose, lactate, total protein, cholesterol, triglycerides, ammonia, chloride, calcium, magnesium, sodium, potassium and urea concentration, total haemocyte counts. Bacteriological analysis of hemolymph was performed in order to determine the possible penetration of microorganisms in the tissues. In this work, for the first time, the urea concentration was determined in these species: at arrival this parameter in hemolymph was 2.74 ± 1.04 mg dl−1, after 12 h in seawater the urea level decreased to 1.17 ± 0.72 mg dl−1. Thanks to this study, a single global parameter useful to measure the distress status was not evidenced, even if the determination of protein concentration was the easiest and cheapest one, although influenced by several factors. Ammonia and urea determination were the most specific parameters for the determination of the air transport distress.

Miniaturized module with biosensors for potentiometric determination of urea

We describe the construction of a miniaturized module which allows carrying out potentiometric urea determination with the usage of biosensors. The module was fabricated using new hybrid technologies developed in our group which combine ceramic and polymeric materials. Its simplicity and easy way of preparation makes the system very useful for analytical measurements in a flow mode. Application of urease-modified polymer membranes deposited on silver screen-printed electrodes in the module allows to determine urea concentration in clinical range. Moreover, it is a very promising construction for other applications in that other enzymes can be immobilized and various bioanalytes can be determined using this module.

Voltammetric Detection of Urea on an Ag-Modified Zeolite-Expanded Graphite-Epoxy Composite Electrode.

In this paper, a modified expanded graphite composite electrode based on natural zeolitic volcanic tuff modified with silver (EG-Ag-Z-Epoxy) was developed. Cyclic voltammetry measurements revealed a reasonably fast electron transfer and a good stability of the electrode in 0.1 M NaOH supporting electrolyte. This modified electrode exhibited moderate electrocatalytic effect towards urea oxidation, allowing its determination in aqueous solution. The linear dependence of the current versus urea concentration was reached using square-wave voltammetry in the concentrations range of urea between 0.2 to 1.4 mM, with a relatively low limit of detection of 0.05 mM. A moderate enhancement of electroanalytical sensitivity for the determination of urea at EG-Ag-Z-Epoxy electrode was reached by applying a chemical preconcentration step prior to voltammetric/amperometric quantification.

Spectrophotometric Determination of Urea in Sugar Cane Distilled Spirits

Urea is an important precursor in the formation of ethyl carbamate, a known carcinogen in alcoholic beverages. Ethyl carbamate has recently been detected at high concentrations in sugar cane distilled spirits, but little is known about the concentration of urea in these beverages. The objectives of this study were to validate methodology for the determination of urea in sugar cane distilled spirits, to determine the levels in 68 samples from different regions within the state of Minas Gerais, Brazil, and to examine the relationship between the concentrations of urea and ethyl carbamate. The method, based on the reaction of urea with 1-phenyl-1,2-propanodione-2-oxime and spectrophotometric quantification at 540 nm, provided linear response from 0.5 to 15.0 mg/L. No purification of the sample was required. The limits of detection and quantification were 0.1 and 0.5 mg/L, respectively. Urea was detected in 69% of the samples at levels varying from 0.50 to 5.10 mg/L. There was no significant difference on the levels of urea in samples from different regions of the state. No significant correlation between the levels of urea and ethyl carbamate was observed for the samples analyzed.

Spectrophotometric Determination of Urea in Dermatologic Formulations and Cosmetics

A rapid, relatively sensitive, and low-cost method for the determination of water-soluble urea content in dermatological therapy products and cosmetics is proposed using a new spectrophotometric assay with water as the only extraction solvent. Spectrophotometric methods involve addition of a known excess of bromate to urea in an acid medium, followed by the determination of residual bromine and chlorine reacting with methyl orange and measurement of absorbance at 505 nm. The absorbance increases linearly with urea concentration (r = 0.9998). The systems obey Beer's law for 6 - 90 microg ml(-1). The calculated apparent molar absorbance values are found to be 4.537 x 10(3) dm(3) mol(-1) cm(-1) and the Sandell's sensitivity is 0.013 microg cm(-2). The variables affecting the rate of the reaction were investigated. The relative standard deviation for five-replication determination of 60 microg ml(-1) urea was 2.1% and the detection limit of the method is 0.34 ng ml(-1).