A fast and simple headspace gas chromatographic technique for quantitatively analyzing urea in human urine.

A fast and convenient headspace gas chromatographic (HS-GC) approach was described for the estimation of urea in human urine. The HS-GC could detect the generated carbon dioxide derived from the urease-catalyzed hydrolysis of urea. It was found that the hydrolysis of urea catalyzed by urease was completed within 40 min at 35 °C. The results proved the great accuracy (relative errors ≤ 8.48%) and precision (RSD ≤ 2.66%) of the HS-GC approach. Moreover, the recoveries ranged from 97.9% to 101.5%. The new approach is rapid and automated, which provides a new way to routinely analyze urea in urine for the control of metabolic disease.

A New Calibration Circuit Design to Reduce Drift Effect of RuO2 Urea Biosensors.

The goal of this study was to reduce the drift effect of RuO2 urea biosensors. A new calibration circuit (NCC) based on the voltage regulation technique with the advantage of having a simple structure was presented. To keep its simplicity, the proposed NCC was composed of a non-inverting amplifier and a voltage calibrating circuit. A ruthenium oxide (RuO2) urea biosensor was fabricated to test the calibrating characteristics of the drift rate of the proposed NCC. The experiment performed in this study was divided into two main stages. For the first stage, a sound RuO2 urea biosensor testing environment was set-up. The RuO2 urea sensing film was immersed in the urea solution for 12 h and the response voltage was measured using the voltage-time (V-T) measurement system and the proposed NCC. The results of the first stage showed that the RuO2 urea biosensor has an average sensitivity of 1.860 mV/(mg/dL) and has a linearity of 0.999 which means that the RuO2 urea biosensor had been well fabricated. The second stage of the experiment verified the proposed NCC's functions, and the results indicated that the proposed NCC reduced the drift rate of RuO2 urea biosensor to 0.02 mV/hr (98.77% reduction).

The EuBIVAS: Within- and Between-Subject Biological Variation Data for Electrolytes, Lipids, Urea, Uric Acid, Total Protein, Total Bilirubin, Direct Bilirubin, and Glucose.

BACKGROUND: The European Federation of Clinical Chemistry and Laboratory Medicine European Biological Variation Study (EuBIVAS) has been established to deliver rigorously determined data describing biological variation (BV) of clinically important measurands. Here, EuBIVAS-based BV estimates of serum electrolytes, lipids, urea, uric acid, total protein, total bilirubin, direct bilirubin, and glucose, as well as their associated analytical performance specifications (APSs), are presented. METHOD: Samples were drawn from 91 healthy individuals (38 male, 53 female; age range, 21-69 years) for 10 consecutive weeks at 6 European laboratories. Samples were stored at -80 °C before duplicate analysis of all samples on an ADVIA 2400 (Siemens Healthineers). Outlier and homogeneity analyses were performed, followed by CV-ANOVA on trend-corrected data, when relevant, to determine BV estimates with CIs. RESULTS: The within-subject BV (CVI) estimates of all measurands, except for urea and LDL cholesterol, were lower than estimates available in an online BV database, with differences being most pronounced for HDL cholesterol, glucose, and direct bilirubin. Significant differences in CVI for men and women/women <50 years of age were evident for uric acid, triglycerides, and urea. The CVA obtained for sodium and magnesium exceeded the EuBIVAS-based APS for imprecision. CONCLUSIONS: The EuBIVAS, which is fully compliant with the recently published Biological Variation Data Critical Appraisal Checklist, has produced well-characterized, high-quality BV estimates utilizing a stringent experimental protocol. These new reference data deliver revised and more exacting APS and reference change values for commonly used clinically important measurands, thus having direct relevance to diagnostics manufacturers, service providers, clinical users, and ultimately patients.

An update report on the harmonization of adult reference intervals in Australasia.

The Australasian Association of Clinical Biochemists (AACB) has over the past 5 years been actively working to achieve harmonized reference intervals (RIs) for common clinical chemistry analytes using an evidence-based checklist approach where there is sound calibration and metrological traceability. It has now recommended harmonized RIs for 18 common clinical chemistry analytes which are performed in most routine laboratories and these have been endorsed by the Royal College of Pathologists of Australasia (RCPA). In 2017 another group of analytes including urea, albumin and arterial blood gas parameters were considered and suggested harmonized RIs proposed. This report provides an update of those harmonization efforts.

Determination of exposure multiples of human metabolites for MIST assessment in preclinical safety species without using reference standards or radiolabeled compounds.

A simple, reliable, and accurate method was developed for quantitative assessment of metabolite coverage in preclinical safety species by mixing equal volumes of human plasma with blank plasma of animal species and vice versa followed by an analysis using high-resolution full-scan accurate mass spectrometry. This approach provided comparable results (within (±15%) to those obtained from regulated bioanalysis and did not require synthetic standards or radiolabeled compounds. In addition, both qualitative and quantitative data were obtained from a single LC-MS analysis on all metabolites and, therefore, the coverage of any metabolite of interest can be obtained.

CALIPER paediatric reference intervals for the urea creatinine ratio in healthy children & adolescents.

BACKGROUND: The urea creatinine ratio (UCR) is important in the clinical assessment of several medical conditions, including acute kidney injury and gastrointestinal bleeding. However, accurate and robust paediatric reference intervals (RIs) for this ratio have not been well established. Here, we determined age- and sex-specific discrete and continuous RIs for UCR in the Canadian Laboratory Initiative on Paediatric Reference Intervals (CALIPER) cohort of healthy children and adolescents for the first time. METHODS: UCR was calculated for approximately 1030 CALIPER participants using retrospective urea and creatinine (both Jaffe and enzymatic methods) normative data. Partitions were determined using the Harris & Boyd statistical method. Discrete RIs were established in accordance with the Clinical and Laboratory Standards Institute (CLSI) guidelines. Continuous RIs were established using nonparametric quantile regression. RESULTS: Several age- and sex-specific partitions were necessary to capture dynamic physiological trends associated with this ratio throughout childhood and adolescence, highlighting the benefit of continuous RI establishment. Established UCR RIs also demonstrated marked differences between Jaffe and enzymatic assay methods. CONCLUSION: Our results clearly demonstrate the critical need for RI stratification by important covariates such as age, sex, and creatinine assay methodology for paediatric UCR test result interpretation. These data contribute to our understanding of normative UCR values in childhood and adolescence and can be expected to improve paediatric test result interpretation in clinical laboratories that report this ratio.

Development and Validation of a HPLC-UV Method for Urea and Related Impurities.

Urea is used in biopharmaceutical manufacturing processes for the purification of therapeutic proteins, for cleaning columns, and for refolding proteins after purification. The urea used for such purposes is typically USP grade material obtained from commercial sources and further characterization is required prior to use, such as determination of purity and identity. For this purpose, a robust analytical method is needed that can characterize the known organic impurities of urea. However, the existing methods show high assay variability and are not able to resolve all known organic impurities as desired for accurate quantification. In the present manuscript we developed a new high-performance liquid chromatography method with UV detection for the separation of urea and its impurities (biuret, cyanuric acid, and triuret). The method performance characteristics evaluated for urea and biuret were specificity, linearity, accuracy, identity, precision, and robustness and the newly developed method met all predefined performance acceptance criteria.

Metal-organic frameworks derived magnetic porous carbon for magnetic solid phase extraction of benzoylurea insecticides from tea sample by Box-Behnken statistical design.

Ferric oxide/carbon (Fe2O3@C) was fabricated via direct carbonization of metal-organic framework of iron (MOF-235) under argon atmosphere. The magnetic Fe2O3 nanoparticles are evenly embedded in porous carbon matrix, while original morphology of MOF-235 was well-maintained. The synthesized Fe2O3@C was used as magnetic sorbent for extracting five benzoylurea insecticides (BUs). The materials exhibited excellent extraction performance, which benefited not only from the strong π-π interaction and hydrophobic interaction (π-conjugated system), but also to the abundant adsorption sites and flexible transport channel (the interconnected 3D porous structure). A three-factor-three-level Box-Behnken design (BBD) was selected to optimize three greatly influential parameters: amount of adsorbent (A), desorption time (B) and volume of desorption solvent (C) by response surface methodology. The established method coupled to HPLC-UV detection showed wide linearity with the range of 0.2-450 µg•L-1, relatively low limits of detection (0.05-0.10 µg•L-1) with the relative standard deviation (RSD) (n = 7) lower t than 5.47%. Moreover, the proposed method was successfully applied to analyze BUs in tea samples and investigate the removal effect of different washing on BUs residues from tea leaf. These results indicated that the synthesized Fe2O3@C is a promising adsorbent material for magnetic solid phase extraction of BUs at trace concentrations from tea samples.

Method for improved accuracy in endogenous urea recovery marker calibrations for microdialysis in tumors.

INTRODUCTION: Urea has been proposed as an endogenous recovery marker for microdialysis for absolute concentration calculations of analytes in microdialysis samples. Previously we demonstrated a linear relationship between urea concentrations in a rat mammary carcinoma and that in plasma, validating its use as a recovery marker for that particular tumor. In this paper, we have extended the validation to two other tumor lines, thereby providing confidence that the calibration is constant across tumor types. To improve the accuracy in the determination of the plasma/tumor urea relationship from no net flux calibrations, we extended the range of the calibration by adding exogenous urea to tumor bearing animals. This method enabled more accurate calculations of absolute recovery from plasma and dialysate urea concentrations. We confirm that by using this method the calibration is valid across three different tumor lines. The existence of a common calibration between tumors provides rationale for using plasma urea as a recovery marker for clinical trials. The existence of a common calibration between tumor types bypasses the need to perform time consuming calibrations for each patient. This makes the procedure much more practical for clinical studies. METHODS: The no net flux technique was used to determine the plasma vs. tumor urea relationship for the R3230Ac mammary carcinoma, 9 L glioma, and a fibrosarcoma (FSa), grown in Fischer 344 rats. Plasma urea was stably increased beyond the normally occurring concentration for some of the data points by subcutaneous bolus administration to extend the range of data for the no net flux calibration. RESULTS: Urea recovery was unaffected by plasma urea concentration and was consistent with other reported values. The relationship between plasma and tumor urea was fit by a line, and linear regressions of the data with the extended plasma urea range had better R2 values than we reported previously. Statistical comparison of the regressions suggests that within reasonable uncertainty limits, they are the same for the different tumor types. DISCUSSION: Increasing the plasma urea concentration range for no net flux calibrations of urea as an endogenous recovery marker in tumors resulted in more accurate determination of the plasma/tumor urea relationship. A single linear regression may describe the relationship between plasma and tumor urea concentration across tumor lines for a given set of microdialysis parameters.

Small amounts of urea and guanidine hydrochloride can be detected by a far-UV spectrophotometric method in dialysed protein solutions.

The quantization of small amounts of chemical denaturants as urea or guanidine hydrochloride in protein solutions after dialysis is a difficult task in the molecular biology laboratory practice. Refractometric methods are useful to quantify a denaturant in the molar range but this methodology is not helpful when the denaturant is present in small amounts. The method herein described is a new comparative method that requires, a priori, the quantification of the stock solutions of urea (8 M) and guanidine hydrochloride (6 M) by refractometry to prepare by sequential dilution the standards used for comparison in the spectropolarimeter. The method is based on the observation that the wavelengths, at which the absorbance of polarized light increases in the far-UV region, as observed by spectropolarimetry, is related to the concentration of the chemical denaturant present in the protein solution. In the quantitation method herein reported, the urea and guanidine hydrochloride detection limits range from 1.2 x 10(-4) to 6 x 10(-6) M depending on the protein dialysis buffer used for a standard cell path length of 1 cm. The sensibility of this method results to be comprised in a range 4-5 orders of magnitude higher than that measured by refractometry. The determinations in both the sample and the control preparations are virtually completed within approximately 10 min.

Validation of a technique for the preparation of individual patient doses of 14C-urea.

A breath test employing 14C-urea is commonly used to detect the presence of Helicobacter pylori in the stomach of patients with peptic ulcer. 14C-urea is not available commercially as a radiopharmaceutical. It is available only as non-pharmaceutical grade material. A technique for preparing individual patient doses was therefore validated and methods for demonstrating the quality of the 14C-urea raw material and final product were developed. Individual patient doses, each containing 100 kBq 14C and 350 mg urea in 1 ml, were prepared and stored at -20 degrees C. Each batch consisted of approximately 400 doses. The activities of samples from each batch were measured by liquid scintillation counting immediately after preparation (99.6 +/- 4.9 kBq) and 6 months later (98.2 +/- 4.7 kBq). The chemical identity of the 14C-urea was demonstrated by a thin-layer chromatographic technique in which the 14C was shown to have the same Rf value as stable urea. The high radiochemical purity of the final product was demonstrated by the presence of only one peak on the thin-layer chromatogram. The radionuclide identity of the 14C-urea was demonstrated by beta-ray spectroscopy. This technique of preparing individual patient doses of 14C-urea results in a product that is stable for at least 6 months.

High-lysine corn as a source of protein and energy for finishing calves.

Three trials evaluated the protein and energy value of high-lysine corn for finishing calves. In Trial 1, 60 finishing steer calves were used to evaluate corn source (high-lysine vs control) and protein source (urea, blood meal [BM], corn gluten meal [CGM]) and level (BM and CGM addition; low, medium, high). Calves were individually fed using Calan gates for 102 d, and then were pen-fed (two pens per corn treatment) the remaining 83 d. During the initial 102 d, calves fed high-lysine corn had similar gains but were 6% more efficient (P < .10) compared with calves fed control corn. Performance did not differ (P > .10) among sources or levels of protein supplementation. Over the entire feeding period (185 d), calves fed high-lysine corn were 10% more efficient (P < .10) than calves fed control corn. In the second study, in situ starch disappearance was faster (P < .10) and the proportion of CP degraded by 12 h was 27% greater (P < .10) for high-lysine corn than for control corn. In a metabolism trial, five steers fitted with ruminal, duodenal, and ileal cannulas were used in a randomized block design to evaluate the effect of corn source on site and extent of digestion. Intake and ruminal and total tract digestibility of OM and N did not differ (P > .10) between corn sources. Steers fed high-lysine corn tended to have greater ruminal (P = .23) and postruminal (P = .18) starch digestion, resulting in greater (P < .10) total tract starch digestibility.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of degradable and escape protein and roughage type on performance and carcass characteristics of finishing yearling steers.

We evaluated protein sources for finishing steers in two randomized complete block design experiments. Experiment 1 used 144 steers (334 kg) with 2 x 3 factorially arranged treatments. Basal diets contained .9% urea or 5.6% soybean meal (SBM) and were either not supplemented or supplemented with additional protein (2%) from blood meal-corn gluten meal (BMCG) or SBM. Steers fed urea-containing diets consumed 4.6% (P < .10) more feed than those fed SBM-supplemented basal diets. On the basis of carcass weights, steers fed diets containing SBM as the basal protein source were 3.8% (P < .10) more efficient than those fed urea-containing diets; supplying additional SBM improved gain efficiency (G/F) 4.3% (P < .10) compared with BMCG. In Exp. 2, 384 steers (367 kg) were fed diets containing 1.0% urea (DM basis) and 10% roughage as either sorghum silage (four diets) or alfalfa hay (two diets). Additional protein was either not provided or provided (2%) as SBM, sunflower meal (SFM), or a 50:50 (N basis) SBM:SFM blend in silage-containing diets; for diets containing alfalfa, additional protein was either not provided or provided (2%) as SBM. Averaged across roughage source, added SBM tended (P = .16) to increase ADG. Dressing percent decreased (P = .09) with added SBM but was higher (P = .04) with alfalfa as roughage source. Feeding alfalfa vs sorghum silage as the roughage source increased carcass adjusted ADG 4.3% (P = .06) and G/F 4.8% (P = .02). Supplementing high-grain diets with SBM enhanced diet utilization, but BMCG was of little value.

Composition of ruminal bacteria harvested from steers as influenced by dietary forage level and fat supplementation.

The objective of this study was to examine the effects of dietary forage level and fat supplementation on the chemical composition of mixed ruminal bacteria (MRB). Six ruminally cannulated beef steers (354 kg +/- 18) were given ad libitum access to six diets (13.2% CP; DM basis) that were offered twice daily in a 6 x 6 Latin square design. Treatments were arranged as a 2 x 3 factorial with two forage levels (70 vs 30% of dietary DM as corn silage) and three forms of fat supplementation including no canola seed or canola seed added at 10% of dietary DM as whole treated with alkaline hydrogen peroxide or untreated crushed. Canola seed contributed 5% added fat to the total diet. The remaining dietary ingredients were corn, canola meal, molasses, and urea. No interactions (P > .05) between dietary forage level and canola seed supplementation were observed. Concentrations of OM, N, and all amino acids were higher (P < .05) in MRB from steers fed low forage than in MRB from steers fed high forage. Concentrations of purines and GE and the N:purines ratio in MRB were not affected (P > .05) by dietary forage level or canola seed supplementation. Canola seed supplementation did not affect (P > .05) concentrations of OM, N, or most of the amino acids in MRB. Concentrations of four essential amino acids (i.e., isoleucine, leucine, lysine, and phenylalanine) in MRB were decreased (P < .05) due to canola seed supplementation. Dietary forage level did not affect (P > .05) concentrations of long-chain fatty acids in MRB.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of protein source on nitrogen metabolism in continuous culture and intestinal digestion in vitro.

Eight dual flow continuous culture fermenters were used in four replicated periods to study the effects of protein supplements on ruminal fermentation and CP digestion. A basal diet was supplemented with isonitrogenous amounts of urea and tryptone (control; CTRL), soybean meal (SBM), lignosulfonate-treated SBM (LSBM), corn gluten meal (CGM), blood meal (BM), hydrolyzed feather meal (HFM), fish meal (FM), or meat and bone meal (MBM). Digestion of DM, OM, and carbohydrates was not affected by treatment. Ammonia N concentration was highest (P < .05) for CTRL and lowest (P < .05) for LSBM, CGM, BM, HFM, and FM. Nonammonia N flow was lowest (P < .05) for CTRL. Dietary N flow was lowest (P < .05) for CTRL, intermediate for SBM, and highest (P < .05) for LSBM, CGM, BM, HFM, FM, and MBM. Total N flow, bacterial N flow, and efficiency of microbial protein synthesis were not affected by treatment. Protein degradation was highest (P < .05) for CTRL. Flow of total amino acids (AA) was lowest (P < .05) for CTRL, SBM, and MBM. Diets containing BM provided the largest (P < .05) amounts of essential AA and lysine, and FM provided the largest (P < .05) amounts of methionine in fermenter effluent. Supplementation of diets with proteins low in ruminal degradability increased flows of nonammonia N, dietary N, and total and essential AA and modified the AA profile flowing out of fermenters.

Protein requirements of growing steers limit-fed corn-based diets.

In Exp. 1, six steers (254 kg) were used in a 6 x 4 incomplete Latin square to determine the effects of solvent-extracted soybean meal alone or in combination with rumen-protected methionine and lysine on N balance in steers limit-fed a high-corn diet to gain 1.1 kg/d. The basal diet contained (DM basis) 80% rolled corn, 15% alfalfa, and .9% urea (13.9% CP), and 2 or 4% soybean meal replaced corn to give CP concentrations of 14.8 and 15.6%, respectively. Each diet was fed with and without 5 g/d of Smartamine-ML (.75 and 2.0 g of rumen-protected methionine and lysine, respectively). Nitrogen retention increased linearly (P = .09) with level of soybean meal. Rumen-protected methionine and lysine had no effect on N balance. In Exp. 2, seven steers (233 kg) were used in a 7 x 4 incomplete Latin square experiment to investigate optimal levels and sources of CP for steers limit-fed to gain 1 kg/d. Treatments included a negative-control diet (urea; 11.7% CP) and six diets containing either 13.5, 15.4, or 17.2% CP with either solvent-extracted or expeller-processed soybean meal. Diets provided 75, 87.5, 100, or 112.5% of estimated CP requirement for a gain of 1 kg/d. The basal diet contained 83% rolled corn, 15% alfalfa, and .2% urea. Nitrogen retention increased linearly (P = .006) with soybean meal addition, and no differences were observed between CP sources. The CP system underpredicted the protein requirements of limit-fed steers under our conditions.

[Determining the levels of selected pesticides, nitrates, nitrites, ammonium ions, sulfates, chlorides and urea in surface and underground waters. VII]. / Oznaczanie wybranych pestycydów, azotanów, azotynów, jonów amonowych, siarczanów, chlorków i mocznika w wodach powierzchniowych i podziemnych. Cz. VII.

The subjects of the study was determination of the residues of pesticides: lindane, metoxychlor and chlorphenvinphos, nitrates, nitrites, ammonium ions, sulphates, chlorides and urea in surface waters and underground waters in the southeastern part of the Province of Szczecin in 1983-1988. The certain pesticides and urea in none of determined samples of waters were present. The concentrations of nitrates, ammonium and chlorides below the permitted value have been. Nitrites in surface waters and in underground waters in 48 per cent and 82 percent of the samples have been respectively.

Raman-on-chip device and detection fibres with fibre Bragg grating for analysis of solutions and particles.

An all-fibre based Raman-on-chip setup is introduced which enables analysis of solutions and trapped particles without microscopes or objectives. Beside the novel quartz microfluidic chip, innovative multi-core single-mode fibres with integrated fibre Bragg gratings are used for detection. The limit of quantitation is 7.5 mM for urea and 2.5 mM for nicotine with linear Raman spectroscopy. This is an improvement of more than two orders of magnitude compared with previous fibre-based microfluidic Raman detection schemes. Furthermore, our device was combined with optical traps to collect Raman-on-chip spectra of spherical polymer beads.

Labeled vs actual concentration of bleaching agents.

The purpose of this study was to determine if the actual concentration of bleaching agents available in four different countries were the same as the label indicated and within the recommendations of the International Standard on Tooth Whitening. The method recommended for assaying peroxide by the United States Pharmacopeia was used to determine concentrations. All products in the United States and China were within the standard when products were tested immediately upon delivery at testing sites. One product in Saudi Arabia and three products in Brazil had greater than 30% concentration loss. Three of 24 products in the United States did not meet the International Standard when they were tested at month of expiration.

Wavelet analysis used for spectral background removal in the determination of glucose from near-infrared single-beam spectra.

Wavelet analysis is developed as a preprocessing tool for use in removing background information from near-infrared (near-IR) single-beam spectra before the construction of multivariate calibration models. Three data sets collected with three different near-IR spectrometers are investigated that involve the determination of physiological levels of glucose (1-30 mM) in a simulated biological matrix containing alanine, ascorbate, lactate, triacetin, and urea in phosphate buffer. A factorial design is employed to optimize the specific wavelet function used and the level of decomposition applied, in addition to the spectral range and number of latent variables associated with a partial least-squares calibration model. The prediction performance of the computed models is studied with separate data acquired after the collection of the calibration spectra. This evaluation includes one data set collected over a period of more than 6 months. Preprocessing with wavelet analysis is also compared to the calculation of second-derivative spectra. Over the three data sets evaluated, wavelet analysis is observed to produce better-performing calibration models, with improvements in concentration predictions on the order of 30% being realized relative to models based on either second-derivative spectra or spectra preprocessed with simple additive and multiplicative scaling correction. This methodology allows the construction of stable calibrations directly with single-beam spectra, thereby eliminating the need for the collection of a separate background or reference spectrum.