Creatinine In Human Urine Samples Research Articles

Nitrogen doped carbon dots and gold nanoparticles mediated FRET for the detection of creatinine in human urine samples

Serum creatinine (CR) is regarded as one of the most sought out prognostic biomarkers in medical evaluation of chronic kidney disease (CKD). In light of the diagnostic significance of CR, the utility of a fluorescence biosensor for its detection in human urine specimens has been explored based on Förster resonance energy transfer (FRET) across nitrogen-doped carbon dots (N-CDs) and gold nanoparticles (GNPs). A straightforward microwave-assisted synthesis procedure has been adopted to prepare N-CDs (λexcitation = 400 nm, λemission = 540 ± 5 nm) with bright green emissions. On addition of pre-synthesized GNPs, the radiative emanation of the N-CDs is completely suppressed on account of FRET across the N-CDs and the GNPs. About 77 % of their fluorescence intensity is recovered after adding CR to GNPs@N-CDs nanocomposite. The limit of detection for CR sensing is estimated as 0.02 µg•mL−1. This biosensor is selective enough to recognize CR in the existence of potential interfering substances (e.g., ascorbic acid, glucose, glutathione, urea, and electrolytes). Its practical utility for CR detection has been validated further on the basis of satisfactory correlation with the benchmark Jaffe method, as observed in artificial/human urine specimens. Consequently, this manuscript marks a pioneering report on employing CDs and GNPs-based FRET for identifying CR in urine specimens of CKD patients.

Portable smartphone integrated 3D-Printed electrochemical sensor for nonenzymatic determination of creatinine in human urine

3D printing technologies are an attractive for fabricating electrochemical sensors due to their ease of operation, freedom of design, fast prototyping, low waste, and low cost. We report the fabrication of a simple 3D-printed electrochemical sensing device for non-enzymatic detection of creatinine, an important indicator of renal function. To create the 3D-printed electrodes (3DE), carbon black/polylactic acid (CB/PLA) composite filament was used. The 3DE was activated using 0.5 M NaOH via amperometry prior to use to improve electrochemical performance. To give selectivity for creatinine, the activated 3DE was modified with a copper oxide nanoparticle-ionic liquid/reduced graphene oxide (CuO-IL/rGO) composite. The modified 3DE was characterized using microscopy and electrochemistry. Cyclic voltammetry and amperometry were used to evaluate sensor performance. The modified 3DE provided electrocatalytic activity towards creatinine without enzymes. Under optimal conditions, the modified 3DE directly coupled with a portable smartphone potentiostat exhibited the linear detection range of 0.5–35.0 mM, and the limit of detection was 37.3 μM, which is sufficient for detecting creatinine in human urine samples. Furthermore, the other physiological compounds present in human urine were not detected on the modified 3DE. Therefore, the modified 3DE could be a tool for effective creatinine screening in the urine.

Comparison of three parallelismassessment methods ofbiomarker quantification by LC-MS/MS: a case study of the bioanalysis of creatinine in human urine samples.

Background: Currently, no regulatory guidelines are available for parallelism assessment for LC-MS biomarker quantification. Spike recovery, standard addition and dilutional linearity are recommended with no mention of the implications of applying these approaches. Results: Here, using human urine creatinine, the authors compared spike recovery and standard addition in LC-MS biomarker quantification, and evaluated a new hybrid approach: parallelism QCs. The authors drew different conclusions based on which approach was used (<15% cutoff). Conclusion: Current recommended approaches may lead to different conclusions and are not equivalent and interchangeable. The authors recommend that standard addition should be the universal 'go-to'method for LC-MS biomarker parallelism assessment; parallelism QCs, which considerthe total concentration as the theoretical value, can be used if theauthentic matrix is limited.

Comprehensive profile of DNA adducts as both tissue and urinary biomarkers of exposure to acrylamide and chemo-preventive effect of catechins in rats

Two representative DNA adducts from acrylamide exposure, N7-(2-carbamoyl-2-hydroxyethyl) guanine (N7-GA-Gua) and N3-(2-carbamoyl-2-hydroxyethyl) adenine (N3-GA-Ade), are important long-term exposure biomarkers for evaluating genotoxicity of acrylamide. Catechins as natural antioxidants present in tea possess multiple health benefits, and may also have the potential to protect against acrylamide-induced DNA damage. The current study developed an ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC−MS/MS) method for simultaneous analysis of N7-GA-Gua and N3-GA-Ade in tissues and urine. The validated UHPLC−MS/MS method showed high sensitivity, with limit of detection and limit of quantification ranging 0.2–0.8 and 0.5–1.5 ng/mL, respectively, and achieved qualified precision (RSD<14.0%) and spiking recovery (87.2%–110.0%) with elution within 6 min, which was suitable for the analysis of the two DNA adducts in different matrices. The levels of N7-GA-Gua and N3-GA-Ade ranged 0.9–11.9 and 0.6–3.5 μg/g creatinine in human urine samples, respectively. To investigate the interventional effects of catechins on the two DNA adducts from acrylamide exposure, rats were supplemented with three types of catechins (tea polyphenols, epigallocatechin gallate, and epicatechin) 30 min before administration with acrylamide. Our results showed that catechins effectively inhibited the formation of DNA adducts from acrylamide exposure in both urine and tissues of rats. Among three catechins, epicatechin performed the best inhibitory effect. The current study provided evidence for the chemo-preventive effect of catechins, indicating that dietary supplement of catechins may contribute to health protection against exposure to acrylamide.

Mass spectrometry-based biosensing using pencil graphite rods

Pencil graphite rods are inexpensive, eco-friendly, and readily available materials that can be useful as probes for biosensing using mass spectrometry (MS). In this sense, we developed a method named graphite-MS in which a pencil graphite rod is used to collect a small amount of sample and is subsequently submitted to MS analysis. The method was optimized and applied for the analysis of several analytes in solution, including creatinine in human urine samples. Figures of merits were obtained by analyzing the insecticide chlorpyrifos in tomato peel extracts, and a calibration curve with R2 > 0.99 was achieved, along with a limit of detection of 80 µg L−1 and limit of quantification of 200 µg L−1. Values of accuracy and precision were below 5%. In tests with solid samples, apples were analyzed by simply puncturing the fruits for 10 s and submitting the rods for MS analysis. Mouse brains were also analyzed (ex vivo and in vivo) without any sample preparation. Discrimination between regions of the mouse brain such as cortex, cerebellum, and ventricle, as well as differentiation between female and male mice, were successfully achieved by using lipid profiles in combination with principal component analysis. In summary, this study increases the range of knowledge about simple and readily available materials that can be used by the scientific community specializing in analytical chemistry and biosensors.

Novel amperometric flow-injection analysis of creatinine using a molecularly-imprinted polymer coated copper oxide nanoparticle-modified carbon-paste-electrode

We report a novel amperometric flow-injection (FI) analysis of creatinine based on a sensor comprising copper oxide nanoparticles (CuO) coated with a molecularly-imprinted polymer (CuO@MIP) and decorating a carbon-paste electrode (CPE) to form the CuO@MIP/CPE electrode. The CuO@MIP was synthesized by using CuO as the supporting core, creatinine as the template, methacrylic acid (MAA) as monomer, N, N′-(1,2-dihydroxyethylene)bis(acrylamide) (DHEBA) as cross-linker, and 2,2′-azobis (2-methylpropionitrile) (AIBN) as initiator. Morphology and structural characterization reveal that CuO nanoparticle imprinted sites (CuO) synthesized using a precipitation method, exhibits features that are well suited to creatinine detection: high surface area, good analyte diffusion and adsorption characteristics that provide shorter response times, and large numbers of specific cavities for enhanced analyte capacity and sensitivity. Cyclic voltammetric measurements indicate that our sensor provides excellent performance toward electro-oxidation of creatinine. The amperometric FI system was used to quantitatively determine creatinine at the CuO@MIP/CPE sensor, in a phosphate buffer carrier. The imprinted sensor exhibits excellent performance for creatinine oxidation at an applied potential of +0.35 V and flow rate of 0.6 mL.min−1. The as-prepared sensor exhibits a linear dynamic range for creatinine detection from 0.5 to 200 μM (r2 = 0.995) with a limit of detection of 0.083 μM (S/N = 3). The system exhibits satisfactorily good precision (%RSD = 1.94%, n = 30) and selectivity toward creatinine. There is only approximately 20% loss from initial response after 2 weeks when stored at 4 oC. We successfully applied the FI detection system to detect creatinine in human urine samples.

A fully automated high-throughput liquid chromatography tandem mass spectrometry method for measuring creatinine in urine

Reliable creatinine measurements are important to evaluate kidney function and for creatinine correction to reduce biological variability of other urinary analytes. A high-throughput, accurate liquid chromatography tandem mass spectrometry method for quantitation of human urinary creatinine has been developed and validated.Sample preparation was fully automated including cryovial decapping, sample ID scanning and two serial dilution steps. Quantitation was performed using a stable isotope-labeled internal standard. Multiplexed chromatographic separation of creatinine was achieved within a one-minute analysis and followed by tandem mass spectrometry in positive electrospray ionization mode. The precursor and product ions of creatinine and D3-creatinine were monitored in selected reaction monitoring mode.Method validation results showed reproducibility with within-run precision of 3.59, 3.49 and 2.84% and between-run precision of 4.01, 3.28 and 3.57% for low, medium and high quality control materials prepared from pooled donor urine, respectively. The method showed excellent accuracy with a bias of −1.94%, −0.78% and −1.07% for three levels of certified reference material. The calibration curve was linear throughout a 7.50–300 mg/dL (0.663–26.5 mmol/L) measurement range (R2 = 0.999), with the mean slope of 0.0115 (95%CI, 0.0108–0.0122) and intercept of 0.0027 (95%CI, 0.0003–0.0051). The limit of detection (LOD) of the method was 3.17 mg/dL (0.280 mmol/L). Analytical specificity was achieved by chromatographically separating creatinine from potentially interfering creatine within a one-minute run and monitoring the Quantitation Ion/Confirmation Ion (QI/CI) ratios in samples.A simple, accurate, high-throughput method was successfully developed for measuring creatinine in human urine samples.

Capillary electrophoresis with dual diode array detection and tandem mass spectrometry to access cardiovascular biomarkers candidates in human urine: Trimethylamine-N-Oxide and l-carnitine

A capillary electrophoresis with diode array and tandem mass spectrometry detection (CE-UV-MS/MS) method has been developed for the targeted assessment of cardiovascular biomarkers candidates, trimethylamine-N-Oxide (TMAO) and l-carnitine, and creatinine in human urine samples. The dual detection was applied due to the high concentration of creatinine (monitored by UV detection at 200 nm) in relation to TMAO and l-carnitine (quantified by selected reaction monitoring (SRM) mass spectrometry), in human urine. All instrumental parameters, sheath liquid (SHL) and background electrolyte (BGE) compositions were optimized with a pool of urine provided by adult healthy volunteers and evaluated by signal-to-noise ratio (SNR) and peak shape of TMAO. The compositions for the optimized BGE was formic acid at concentration of 0.10 mol L−1, and for SHL was 70:30 MeOH:H2O containing 0.05% (v/v) formic acid, delivered at a flow rate of 5 μL min−1. Limits of detection for TMAO, l-carnitine and creatinine were 0.76, 0.54 and 303 μmol L−1, respectively. Limits of quantification were 2.5, 1.8 and 1000 μmol L−1, respectively. Linearity was evaluated by ANOVA and presented R2 from 0.993 to 0.997. Precision and accuracy were evaluated at three concentration levels. Coefficients of variation (CV) from 1 to 21% were obtained for the intra-day precision evaluation and from 2 to 16% for the inter-day precision evaluation. The recovery ranged from 75 to 116%. Quantitation of TMAO and l-carnitine in infarcted patients urine in comparison to healthy individuals indicated a 2.2 fold increase of TMAO and a 7.0 fold increase of l-carnitine. These results showed the potential applicability of the proposed method for the evaluation of TMAO and l-carnitine in urine within a panel of candidate metabolites in targeted metabolomics studies of cardiovascular diseases among other conditions.

Protein-protected red emittive copper nanoclusters as a fluorometric probe for highly sensitive biosensing of creatinine

We describe red emittive bovine serum albumin-modified copper nanoclusters (BSA-CuNCs) applied as a precise fluorescent nanoprobe for the optical recognition of creatinine in human urine samples.

Calorimetric Biosensing System for Quantification of Urinary Creatinine.

In this work, we demonstrate the quantification of creatinine in human urine samples using a microcalorimetric sensing system. The calorimetric sensor is based on an array of microfabricated Y-cut quartz resonators. The piezoelectric quartz is etched down to a thickness of 10 μm and exhibits a bulk acoustic resonance of 166 MHz. The temperature sensitivity of this high-frequency quartz resonator is 14 600 Hz/K due to the high phenomenological sensitivity of quartz. Most importantly, the quartz sensors and the analyte fluidics are decoupled providing a significantly more robust calorimetric sensing system than directly contacted chip calorimeters. A reference resonator, consisting of a suspended structure held by four arms, was realized to thermal isolation from the bulk quartz by using focused ion beam etching. We employ alginate entrapped creatinine deiminase to transduce urinary creatinine into temperature signatures, permitting the quantification of creatinine. Fairly good agreement with the measured creatinine values in the 5 urine samples using calorimetric and HPLC methods is obtained.

Paper-based Platform for Urinary Creatinine Detection

A new paper platform was developed for the colorimetric detection of creatinine. The filter paper was coated with 3-propylsulfonic acid trimethoxysilane and used as the platform. Creatinine in a cationic form was extracted onto the paper via an ion-exchange mechanism and detected through the Jaffé reaction, resulting in a yellow-orange color complex. The color change on the paper could be observed visually, and the quantitative detection of creatinine was achieved through monitoring the color intensity change. The color intensity of creatinine complexes on the paper platform as a function of the creatinine concentration provided a linear range for creatinine detection in the range of 10 - 60 mg L(-1) and a detection limit of 4.2 mg L(-1). The accuracy of the proposed paper-based method was comparable to the conventional standard Jaffé method. This paper platform could be applied for simple and rapid detection of creatinine in human urine samples with a low consumption of reagent.

A simple and rapid creatinine sensing via DLS selectivity, using calix[4]arene thiol functionalized gold nanoparticles

A new, simple, ultra-sensitive and selective approach has been reported for the “on spot” colorimetric detection of creatinine based on calix[4]arene functionalized gold nanoparticles (AuNPs) with excellent discrimination in the presence of other biomolecules. The lower detection limit of the method is 2.16nM. The gold nanoparticles and p-tert-butylcalix[4]arene were synthesized by microwave assisted method. Specifically, in our study, we used dynamic light scattering (DLS) which is a powerful method for the determination of small changes in particle size, improved selectivity and sensitivity of the creatinine detection system over colorimetric method. The nanoassembly is characterized by Transmission electron microscopy (TEM), DLS, UV–vis and ESI–MS spectroscopy, which demonstrates the binding affinity due its ability of hydrogen bonding and electrostatic interaction between –NH group of creatinine and pSDSC4. It exhibits fast response time (<60s) to creatinine and has long shelf-life (>5 weeks). The developed pSDSC4–AuNPs based creatinine biosensor will be established as simple, reliable and accurate tool for the determination of creatinine in human urine samples.

Chemiluminescence of creatinine/H2O2/Co(2+) and its application for selective creatinine detection.

Creatinine is an important biomarker in clinical diagnosis and biomonitoring programs as well as urinary metabolomic/metabonomics research. Current methods are either nonselective, time consuming or require heavy and expensive instruments. In this study, chemiluminescence of creatinine with hydrogen peroxide has been reported for the first time, and its chemiluminescence is remarkably enhanced in the presence of cobalt ions. By utilizing these phenomena, we have developed a sensitive and selective chemiluminescence method for creatinine determination by coupling with flow injection analysis. The calibration curve is linear in the range of 1×10(-7)-3×10(-5)mol/L with a limit of detection (S/N=3) of 7.2×10(-8)mol/L, which is adequate for detecting creatinine in the clinically accepted range. The relative standard deviation for seven measurements of 3×10(-5)mol/L creatinine is 1.2%. The chemiluminescence method was then utilized to detect creatinine in human urine samples after simple dilution with water. It takes less than 1min each measurement and the recoveries for spiked urine samples were 100-103%. The interference study demonstrates that some common species in urine, such as amino acids, ascorbic acid and creatine, have negligible effects on creatinine detection. The present method does not use expensive instruments, enzymes and separation technique. This method has the advantages of sensitivity, selectivity, simplicity, rapidity, and low cost. It holds great promise for basic or comprehensive metabolic panel, drug screening, anti-dopping, and urinary metabolomic/metabonomics research.

Detection of urinary creatinine using gold nanoparticles after solid phase extraction

Label-free gold nanoparticles (AuNPs) were utilized in the detection of creatinine in human urine after a sample preparation by extraction of creatinine on sulfonic acid functionalized silica gel. With the proposed sample preparation method, the interfering effects of the urine matrix on creatinine detection by AuNPs were eliminated. Parameters affecting creatinine extraction were investigated. The aggregation of AuNPs induced by creatinine resulted in a change in the surface plasmon resonance signal with a concomitant color change that could be observed by the naked eye and quantified spectrometrically. The effect of AuNP concentration and reaction time on AuNP aggregation was investigated. The method described herein provides a determination of creatinine in a range of 15–40mgL−1 with a detection limit of 13.7mgL−1 and it was successfully used in the detection of creatinine in human urine samples.

Development of a fast capillary electrophoresis method for determination of creatinine in urine samples

The aim of this work was to develop a fast method using capillary electrophoresis for the determination of creatinine in human urine samples. The pH and constituents of the background electrolyte were selected by inspection of effective mobility of creatinine and candidate urine interferents versus pH curves. The tendency of the analyte to undergo electromigration dispersion and the buffer capacity were evaluated by the Peakmaster software and considered in the optimization of the background electrolyte, composed by 10 mmol L −1 tris(hydroxymethyl)aminomethane and 20 mmol L −1 2-hydroxyisobutyric acid (HIBA) at pH 3.93. Separation was conducted in a fused-silica capillary (32 cm total length and 8.5 cm effective length, 50 μm I.D.), with short-end injection configuration and direct UV detection at 215 nm. The migration time of creatinine was only 22 s. A few figures of merit of the method are as follows: good linearity in the concentration interval of 5–70 mg L −1 ( R 2 > 0.99), limit of detection of 0.5 mg L −1, inter-day precision better than 2.7% ( n = 9) and recovery in the range 99.0–103.7% at three concentration levels (50, 100 and 150 mg L −1). Urine samples were prepared by deproteination with acetonitrile (1:3 sample:acetonitrile, v/v), centrifugation and dilution of a deproteinated aliquot with 12.5 mmol L −1 HIBA (1:4, v/v). Creatinine concentrations between 489 and 1063 mg L −1 were obtained in the urine of four healthy volunteers.

Simultaneous determination of urinary creatinine, calcium and other inorganic cations by capillary zone electrophoresis with indirect ultraviolet detection

A method for the rapid analysis of calcium and creatinine in urine has been developed using capillary zone electrophoresis without sample pretreatment other than dilution. Cations of NH 3, K, Na, Mg, Ca, Li, Ba and creatinine in human urine samples could be separated within 7 min. Background electrolyte (BGE) was composed of 5 mmol/l pyridine (chromophore), 3.6 mmol/l tartaric acid and 2 mmol/l 18-crown-6, having a pH of 4.05. An indirect UV detection mode was employed at 255 nm. Efforts were made to eliminate the interaction between cations and proteins and to minimize the electromigrative dispersion (EMD) for the analytes of interest by selecting a suitable BGE and sample dilution solution. Quantitative analyses were performed for calcium and creatinine. The calibration plots showed good linearity over the concentration range of interest to clinical analysis. Data on recoveries and reproducibilities are also reported. Results for urine samples (previously collected and frozen) from a variety of healthy and pathological individuals were in good agreement with those obtained by the Technicon SMA II calcium and creatinine methods.