Determination Of Creatinine In Urine Research Articles

Determination of creatinine in urine and blood serum human samples by CZE-UV using on-column internal standard injection.

Creatinine is a well-stablished biomarker for kidney malfunctions and for normalization parameter of urinary quantitative information. Recently, metabolic studies have been discovering other functionalities for creatinine tests in human urine and blood serum. In this work we present an enhanced capillary electrophoresis (CE) based protocol for determination of creatinine. CE is a high-throughput separation technique that have been getting attention through the last decades and might be considered to be adopted as an analytical instrumentation for clinical purposes. In the proposed method, we performed a short injection program with on-column addition of internal standard. Additionally, the method allows a simultaneous screening of non-proteinogenic amino acids that could be considered for metabolomics purposes. We design a pilot study that successfully estimated the creatinine value in 100 urine samples with (2.85±1.78) mg dL-1 LOD; (8.24±5.93) mg dL-1 LOQ and 82.4% accuracy. Considering that serum creatinine is also included in the clinical laboratory routines for estimated Glomerular Filtration Rate dosage, the method was complementary applied to 10 blood serum samples, which resulted in a model with (0.4±0.2)mgdL-1 LOD; (2.0±0.6)mgdL-1 LOQ and 83.8% of accuracy. All results were in agreement with reference values. The proposed method promotes a great analytical frequency and reproducibility with enhanced specificity compared with the ongoing protocol by Jaffe's reaction, thereby proving to be useful as an alternative for creatinine exams that might help complete a diagnosis of a series of health-related issues.

Determination of Creatinine in Urine by Voltammetry and Glassy Carbon Electrode Modified with Functionalized Multi‐walled Carbon Nanotubes and Copper

AbstractA square‐wave voltammetric method was developed for the determination of creatinine using glassy carbon electrode functionalized multi‐walled carbon nanotubes and doped with copper. Since the creatine‐creatinine balance is dependent on pH, the linear responses at either pH 4.0 or 7.0 were evaluated allowing versatility in application, covering analyte concentration from limit of quantification (0.3 μmol L−1) up to 1.200 μmol L−1 (pH 4) or 660 μmol L−1 (pH 7). Electrode is simple to prepare and robust for routine analysis and it was used for the determination of creatinine in urine after simple thin‐layer separation procedure to avoid interference from creatine.

Construction of Au@Metal-organic framework for sensitive determination of creatinine in urine

Creatinine level in urine is an important biomarker for renal function diseases, such as renal failure, glomerulonephritis, and chronic nephritis. The Au@MIL-101(Fe) was prepared by in situ growth of Au nanoparticles in MIL-101(Fe) as a selective SERS substrate. The Au@MIL-101(Fe) offers the great local surface plasmon resonance (SPR) effect due to gold nanoparticles aggregation inside metal-organic frameworks. The framework structure could enrich trace target samples and drag them into SPR hot spots. The optimal Au@MIL-101(Fe) composite substrate is used for analyzing creatinine in urine and the limit of detection is down to 0.1[Formula: see text][Formula: see text]mol/L and a linear relationship is ranging from 1[Formula: see text][Formula: see text]mol/L to 100[Formula: see text][Formula: see text]mol/L.

Low-Cost Synthesis of Gold Nanoparticles from Reused Traditional Gold Leaf and its Application for Sensitive and Selective Colorimetric Sensing of Creatinine in Urine

Background: Gold nanoparticles (Au NPs) are normally prepared using standard gold (III) trichloride which is much expensive and irritant. This work is aimed at demonstrating simple and low-cost synthesis of Au NPs from the reused traditional gold leaf which is cost-free and less toxic. Methods: The reused gold leaf was donated by the local temple. It was digested and used as the precursor for the preparation of the Au NPs by Turkevich method. Poly (vinyl alcohol) (PVA) was employed as a stabilizer. The as-prepared Au NPs were applied for the colorimetric determination of creatinine in urine without any sample pretreatment. Results: Long-term stability of the gold colloids was achieved for at least 3 months. Morphology and purity of the as-prepared Au NPs were the same as the ones prepared from standard gold (III) salt and standard gold foil. Colorimetric response of the Au NPs was linear to the standard creatinine up to 200 mg L-1. The limit of detection (0.16 mg L-1 or 1.41 μM) was enough sensitive for urinary creatinine detection in patients with kidney disease. Good recoveries (97-108%) and fast analysis time (3 min) were achieved. The developed method was successfully validated against the HPLC method. Conclusion: Facile and cost-effective synthesis of the Au NPs from the reused traditional gold leaf, was accomplished. The as-prepared Au NPs were successfully applied for the determination of urinary creatinine with high sensitivity and selectivity.

Comparison of urinary vitamin D binding protein with albumin-creatinine ratio in type 2 Diabetes Mellitus as an early screening tool for diabetic nephropathy

ABSTRACT
 
 Introduction:-
 Diabetes mellitus (DM) is a group of metabolic disorders. Diabetic nephropathy is one of the chronic complications of DM, leading to End stage renal disease (ESRD). A current diagnostic criterion for diabetic nephropathy is measurement of microalbuminuria, which is 30 to 300 mg of albumin/24 hour’s urine or ratio of albumin to creatinine (ACR) in the range of 30 to 300 mg/g in random urine sample but it shows inadequate sensitivity for the early detection of diabetic nephropathy.
 Methodology:- This was a comparative cross sectional study, which comprised of seventy five study subjects and were distributed into three study groups with 25 subjects in each group, having age in the range of 40-50 years.Group-1 comprising controls (without diabetes mellitus),Group-2 contained diabetes mellitus patients with normoalbuminuria. Group-3 composed of diabetes mellitus patients with microalbuminuria. Vitamin D binding protein, urine creatinine and albumin was measured from the urine sample of each study subjects. Determination of creatinine in urine was performed by jaffe method and albumin in urine was determined by immunoturbidimetric method. Vitamin D binding protein was measured by ELISA method. Levels of VDBP and albumin were normalized with urine creatinine and expressed as VDBP creatinine ratio as (ng/mg) and albumin creatinine ratio as (mg/g) in the spot urine sample.
 Results:-Results of this study showed that level of Vitamin D binding protein was significantly increased in diabetes mellitus in comparison to control subjects.
 Conclusion:- Our results proposes that urinary vitamin D binding protein levels is likely to become a useful biomarker for the early detection of diabetic nephropathy in diabetic patients which can be helpful in early treatment and will help to manage diabetic nephropathy.
 ABSTRACT
 Introduction: Diabetes mellitus (DM) is a group of metabolic disorders. Diabetic nephropathy is one of the chronic complications of DM, leading to end stage renal disease (ESRD). A current diagnostic criterion for diabetic nephropathy is measurement of microalbuminuria, which is 30 to 300 mg of albumin/24 hour’s urine or ratio of albumin to creatinine (ACR) in the range of 30 to 300 mg/g in random urine sample but it shows inadequate sensitivity for the early detection of diabetic nephropathy .Current diagnostic criterion for diabetic nephropathy (DN) is detection of microalbuminuria, which is 30 – 300 mg/24 hours of albumin excretion in urine or albumin to creatinine ratio (ACR) in the range of 30 – 300 mg/g in the random urine sample but, it shows inadequate sensitivity for the early detection of DN. It has been observed that increased excretion of UVDBP is related to tubular dysfunction. This protein is excreted in urine earlier than albumin. Hence it can be used as a tool to early detection of DN in type 2 diabetic patients.
 Methodology: This was a comparative cross sectional study, which comprised of seventy five study subjects and were distributed into three study groups with 25 subjects in each group, having age in the range of 40-50 years. Group-1 comprising controls (without diabetes mellitus), Group-2 had diabetes mellitus with normoalbuminuria while Group-3 comprised of diabetes mellitus patients with microalbuminuria. Vitamin D binding protein, urine creatinine and albumin were measured from the random urine sample preferably early in the morning urine sample of each study subjects using ELISA, Jaffe and immunoturbidimetric methods respectively. Levels of VDBP and albumin were normalized with urine creatinine and expressed as VDBP creatinine ratio as (ng/mg) and albumin creatinine ratio as (mg/g) in the spot urine sample.
 Results: Urinary VDBP levels among the three groups were as the highest Median, IQR (Q1- Q3) values were observed in group 3 as 1056 ng/mg,(905 ng/mg – 1215 ng/mg) followed by group 2 as 442 ng/mg,(381.50 ng/mg – 523 ng/mg) and group 1 as 98 ng/mg,(73.50 ng/mg – 149 ng/mg) respectively, and a statistical significant difference was observed among the three groups with a p- value of 0.000. Results of this study showed that level of Vitamin D binding protein was significantly increased in diabetes mellitus in comparison to control subjects.
 Conclusion: Results suggest that urinary vitamin D binding protein level is likely to become a useful biomarker for the early detection and management of diabetic nephropathy in Type 2 diabetic patients.

Simple and fast fabrication of microfluidic paper-based analytical device by contact stamping for multiple-point standard addition assay: Application to direct analysis of urinary creatinine

In this work, a microfluidic paper-based analytical device (μPAD) for simultaneous multiple-point standard addition assay was fabricated by rubber stamping the hydrophobic barrier pattern onto a laboratory filter paper. The μPAD has a central zone from which an applied sample flows into eight surrounding narrow channels to which had been added the standard solutions. Each channel is connected to a circular area loaded with the reagent. The opposite end of this reagent zone is connected by second narrow channel to the final circular detection zone. The μPAD was applied to the measurement of creatinine in human urine. After addition of a urine sample, the orange-colored product arising from the Jaffé reaction is formed at the eight detection zones. A digital image of the μPAD is then recorded and the ratio of the red/green (R/G) intensity obtained using the ImageJ™ program is used in the quantitation of creatinine. The normal standard addition calibration line is constructed using the intensity ratio against the added creatinine concentrations (50–1000 mg L−1). Good linearity was achieved (r2 ˃ 0.99). There were no significant differences between the creatinine content using an HPLC method (paired t-test at 95% confidence, tstat = 1.78, tcritical = 2.26, n = 10). The use of simultaneous multiple-point standard addition calibration allows rapid determination of creatinine in urine with elimination of matrix interference.

Ionophore-Based Optical Sensor for Urine Creatinine Determination.

Creatinine is a metabolite present in urine, and its concentration is used to diagnose and monitor kidney performance. For that reason, the development of new sensors to analyze this metabolite and obtain accurate results in a short period of time is necessary. An optical disposable sensor for monitoring creatinine levels in urine is described. The system, based on a new aryl-substituted calix[4]pyrrole synthetic receptor, has an unusual coextraction scheme. Due to the low p Ka values of creatininium (p Ka 4.8), a careful selection of a lipophilic pH indicator that works in acid medium is required. The sensor components were optimized, and the new sensor displays a good response time to creatinine (approximately 3 min) over a wide dynamic range (from 1 × 10-5 to 1 × 10-2 M). Moreover, the optical selectivity coefficients obtained for creatinine over common cations present in urine meet the requirements for real sample measurements. With a good sensor-to-sensor reproducibility (RSD, 5.1-6.9% in the middle of the range), this method provides a simple, quick, cost-effective, and selective alternative to the conventional methodology based on Jaffé's reaction.

FIA-MS/MS determination of creatinine in urine samples undergoing butylation

Flow injection analysis-tandem mass spectrometry has become widely used for analysis of many biomarkers in various biological matrices. To improve the sensitivity, the compounds are often determined as their butylesters. Since the concentration of urinary excreted compounds are generally reported after normalization to creatinine, the aim of this study was to investigate the possibility of creatinine determination in urine samples which underwent butylation. The impact of derivatization on urinary creatinine determination was investigated by measuring of underivatized and derivatized samples. The 10% creatine to creatinine conversion was observed during butylation, what above 700 μmol creatine/mmol creatinine caused significant creatinine overestimation. In that case, correction for creatine conversion rate was done. QC samples at six concentration levels were examined and precision and accuracy values fulfill the European Medicine Agency validation requirements. The elaborated method was applied for determination of creatinine in 41 real human urine samples. Determined creatinine concentrations were in the range of 0.27–22.3 mmol/L, linearity was confirmed within the concentration range of 0.27–31.7 mmol/L. Obtained results highly correlated with routinely used enzymatic assay for all tested samples and proposed method provide reliable determination of creatinine in butylated urine in a single run with butylesters of other analytes of interest.

Study on nephroprotective effect of Ebselen in cisplatin induced renal damage in rats

Background: Renal dysfunction arises as a result of exposure to medicines, industrial or environmental chemicals. Cisplatin is a major antineoplastic drug used for the treatment of solid tumors. Its chief dose limiting side effect is nephrotoxicity; 20% of patients receiving high-dose cisplatin have severe renal dysfunction. Ebselen a promising antioxidant, was used to explore the nephroprotective effect.Methods: The rats were divided into five groups; each group consisting of 6 animals. The experimental design included one control group and four experimental groups. The study was carried out for a period of 7 wks. The test drug Ebselen in group 4 and 5 and the reference standard drug Amifostine in group 3 was administered once a week intraperitoneally for 5 weeks. Nephrotoxicity was induced by cisplatin (5mg/kg IP) in the 6th week, following this the drug Amifostine in group 3 and Ebselen in group 4 and 5 will be continued twice a day for 5 consecutive days post induction. Urine samples were collected and sent for determination of urine creatinine and albumin.Results: The Urine creatinine level and albumin level estimation in group II show significant renal damage as compared to control group. The statistical reduction in urine creatinine and urine albumin level in Ebselen treated group I (10mg/kg), Ebselen group V (20mg/kg) as compared to Cisplatin group II show a potential reduction in renal damage. Ebselen treated group V showed a reduction in urine creatinine and urine albumin as same as in group III.Conclusions: This study brings to a close that Ebselen lessens Cisplatin induced renal damage.

Characterization of a new ionophore-based ion-selective electrode for the potentiometric determination of creatinine in urine

The optimization, analytical characterization and validation of a novel ion-selective electrode for the highly sensitive and selective determination of creatinine in urine is presented. A newly synthesized calix[4]pyrrole-based molecule is used as an ionophore for the enhanced recognition of creatininium cations. The calculation of the complex formation constants in the polymeric membrane with creatininium, potassium and sodium confirms the strong selective interactions between the ionophore and the target. The optimization of the potentiometric sensor presented here yields an outstanding analytical performance, with a linear range that spans from 1µM to 10mM and limit of detection of 10−6.2M. The calculation of the selectivity coefficients against most commonly found interferences also show significant improvements when compared to other sensors already reported. The performance of this novel sensor is tested by measuring creatinine in real urine samples (N=50) and comparing the values against the standard colorimetric approach (Jaffé's reaction). The results show that this sensor allows the fast and accurate determination of creatinine in real samples with minimal sample manipulation.

Intensified training periods: when much is to too much?

[b]Introduction.[/b] This study has been undertaken in order to examine the usefulness of selected biomarkers as commonly used for monitoring of biological responses to an intensified training period.[b]Material and methods.[/b] The subject was monitored day-by-day throughout an a 2-wk training period, which preceded a pre competition 10-day taper period. Morning blood biochemical indices, perceived sleep quality (SQ), hand-grip strength (H-GS), visuo-attentional motor level (VAM) were assayed and provided to the coach no later than 3 hours after blood sampling and just before the morning training session. Overnight collected urine samples were stored frozen until determination of urinary creatinine, urea and free cortisol.[b]Results. [/b]During the first half of the training period plasma creatine kinase tended to increase, whereas T tended to decrease. In the morning of the 10th day of the period the testosterone rapidly decreased under the lower physiological limit, that was associated with strongly elevated urinary excretion cortisol and urea, deteriorated SQ, H-GS, VAM and reported malaise. Judoka had to stop training. After 2 days of rest period all these psycho-physiological indices returned to the baseline values, judoka felt better and declared his willingness to resume the training.[b]Conclusions.[/b] The results showed usefulness of comprehensive biometric monitoring across an intensified training period. Excessive training with inadequate rest periods resulted in unpredictable, acute psycho-physical indispositions along with changes in the physiological markers similar to those noted during chronic fatigue. Considering very short length of the duration of that syndrome, it does meet criteria of overtraining/overreaching.

A facile low-cost enzymatic paper-based assay for the determination of urine creatinine

Creatinine is one of many markers used to investigate kidney function. This paper describes a low-cost enzymatic paper-based analytical device (enz-PAD) for determining urine creatinine. The disposable dead volumes of creatinine enzyme reagents from an automatic analyser cassette were utilised. Whatman No. 3 paper was cut into long rectangular shapes (4×40mm2) on which the enzyme reagents, R1 and R2, were adsorbed in two consecutive regions. The assay was performed by immersing test strips into urine samples contained in microwells to allow creatinine in the sample to react with immobilised active ingredients and, then, traverse via capillary action to the detection area where chromogen products accumulated. The method is based on hydrogen peroxide (H2O2) formation via creatinine conversion using creatininase, creatinase, and sarcosine oxidase. The liberated H2O2 reacts with 4-aminophenazone and 2,4,6-triiodo-3-hydroxybenzoic acid to form quinoneimine with a pink-red colour at the detection zone. The linear range of the creatinine assay was 2.5–25mgdL−1 (r2=0.983), and the detection limit was 2.0mgdL−1. The colorimetric enz-PAD for the creatinine assay was highly correlated with a conventional alkaline picrate method when real urine samples were evaluated (r2=0.977; n=40). This simple and nearly zero-cost paper-based device provides a novel alternative method for screening urinary creatinine and will be highly beneficial for developing countries.

Validation of a multi-analyte HPLC-DAD method for determination of uric acid, creatinine, homovanillic acid, niacinamide, hippuric acid, indole-3-acetic acid and 2-methylhippuric acid in human urine

During the last decades exposure sciences and epidemiological studies attracts more attention to unravel the mechanisms for the development of chronic diseases. According to this an existing HPLC-DAD method for determination of creatinine in urine samples was expended for seven analytes and validated. Creatinine, uric acid, homovanillic acid, niacinamide, hippuric acid, indole-3-acetic acid, and 2-methylhippuric acid were separated by gradient elution (formate buffer/methanol) using an Eclipse Plus C18 Rapid Resolution column (4.6mm×100mm). No interfering signals were detected in mobile phase. After injection of blank urine samples signals for the endogenous compounds but no interferences were detected. All analytes were linear in the selected calibration range and a non weighted calibration model was chosen. Bias, intra-day and inter-day precision for all analytes were below 20% for quality control (QC) low and below 10% for QC medium and high. The limits of quantification in mobile phase were in line with reported reference values but had to be adjusted in urine for homovanillic acid (45mg/L), niacinamide 58.5(mg/L), and indole-3-acetic acid (63mg/L). Comparison of creatinine data obtained by the existing method with those of the developed method showing differences from -120mg/L to +110mg/L with a mean of differences of 29.0mg/L for 50 authentic urine samples. Analyzing 50 authentic urine samples, uric acid, creatinine, hippuric acid, and 2-methylhippuric acid were detected in (nearly) all samples. However, homovanillic acid was detected in 40%, niacinamide in 4% and indole-3-acetic acid was never detected within the selected samples.

Fast vaporization solid phase microextraction and ion mobility spectrometry: A new approach for determination of creatinine in biological fluids

In this work a rapid and simple method for creatinine determination in urine and plasma samples based on aqueous derivatization of creatinine and complete vaporization of sample (as low as 10µL), followed by ion mobility spectrometry analysis has been proposed. The effect of four important parameters (extraction temperature, total volume of solution, desorption temperature and extraction time) on ion mobility signal has been studied. Under the optimized conditions, the quantitative response of ion mobility spectrometry for creatinine was linear in the range of 0–500mgL−1 with a detection limit of 0.6mgL−1 in urine and 0–250mgL−1 with a detection limit of 2.6mgL−1 in plasma sample. The limit of quantitation of creatinine was 2.1mgL−1 and 8.7mgL−1 in urine and plasma samples, respectively. The relative standard deviation of the method was found to be 13%. The method was successfully applied to the analysis of creatinine in biological samples, showing recoveries from 92% to 104% in urine and 101–110% in plasma samples.

Direct Determination of Creatinine in Urine and Analysis of Pure Aniline by Extractive Electrospray Ionization Mass Spectrometry

The direct mass spectrometric determination of highly concentrated analytes in human urine was demonstrated using extractive electrospray ionization without sample dilution or complex preparation. By increasing the distance between the extractive electrospray source and ion inlet of the mass spectrometer from 5 millimeters to 15 centimeters, the fraction of free analyte ions and charged microdroplets introduced into the mass spectrometer was substantially reduced. Consequently, detector saturation, instrument contamination, and space charge effects were greatly diminished for the analysis of highly concentrated samples. Under the optimized experimental conditions, pure aniline and creatinine (>1 millimolar) in human urine were directly characterized by extractive electrospray ionization without any pretreatment. The urinary creatinine concentrations from two adults were 424 ± 30 and 635 ± 32 micrograms per milliliter and were in good agreement with those obtained by a spectrophotometric method based on the Jaffe reaction. The results show that extractive electrospray ionization is suitable for the direct determination of highly concentrated analytes or even pure compounds, allowing rapid characterization of samples in the chemical industry and clinical studies.

Evaluating an Alternative Method for Rapid Urinary Creatinine Determination

Creatinine (CR) is an endogenously produced chemical that is routinely assayed in urine specimens to assess kidney function and sample dilution. The industry-standard method for CR determination, known as the kinetic Jaffé (KJ) method, relies on an exponential rate of a colorimetric change, and can therefore require automated processing equipment for moderate- to high-throughput analysis (hundreds to thousands of samples per day). This study evaluates an alternative colorimetric method, the “plateau Jaffé” (PJ) method, which utilizes the chemistry of the KJ method, a commercially available kit, and a multipoint calibration curve. This method is amenable to moderate-throughput sample analysis and does not require automated processing equipment. Thirty-two spot urine samples from healthy adult volunteers were analyzed for creatinine concentration (CRc) using the KJ and PJ methods. Samples were also analyzed using a liquid chromatography time-of-flight mass spectrometry (LC-TOF/MS) method, which acted as an analytical control. Replicate measurements of spot samples (natural log-transformed values) were used to estimate method precision, and linear regression models were used to evaluate method accuracy (LC-TOF/MS measurements were considered the analytical benchmark). Measurement precision was comparable across all three methods, with coefficent of variation estimates ranging from 3 to 6%. Regression models generally showed good agreement across methods with R2 estimates ranging from .996 to .998, slope estimates ranging from .944 to .986, and y-intercept estimates ranging from 0.111 to 0.303. Minor bias (between 2 and 16%) was observed across methods at the tails of the measurement distributions. The provided regression equations can be used to adjust for this bias and to improve CR measurement comparisons across studies employing different methods. Considering these results, the PJ method is a suitable alternative to the industry standard KJ method for urinary CRc determination. It can be implemented for moderate-throughput sample analysis using modest and commonly available lab instrumentation and manual sample preparation techniques.

Photochemical decoration of magnetic composites with silver nanostructures for determination of creatinine in urine by surface-enhanced Raman spectroscopy

In this study, silver nanostructures decorated magnetic nanoparticles for surface-enhanced Raman scattering (SERS) measurements were prepared via photoreduction utilizing the catalytic activity of ZnO nanostructure. The ZnO/Fe3O4 composite was first prepared by dispersing pre-formed magnetic nanoparticles into alkaline zinc nitrate solutions. After annealing of the precipitates, the formed ZnO/Fe3O4 composites were successfully decorated with silver nanostructures by soaking the composites into silver nitrate/ethylene glycol solution following UV irradiations. To find the optimal condition when preparing Ag@ZnO/Fe3O4 composites for SERS measurements, factors such as the reaction conditions, photoreduction time, concentration of zinc nitrate and silver nitrate were studied. Results indicated that the photoreduction efficiency was significantly improved with the assistance of ZnO but the amount of ZnO in the composite is not critical. The concentration of silver nitrate and UV irradiation time affected the morphologies of the formed composites and optimal condition in preparation of the composites for SERS measurement was found using 20mM of silver nitrate with an irradiation time of 90min. Under the optimized condition, the obtained SERS intensities were highly reproducible with a SERS enhancement factor in the order of 7. Quantitative analyses showed that a linear range up to 1µM with a detection limit lower than 0.1µM in the detection of creatinine in aqueous solution could be obtained. Successful applying of these prepared composites to determine creatinine in urine sample was obtained.

Determination of Urinary Creatinine in Washington State Residents via Liquid Chromatography/Tandem Mass Spectrometry.

A viable, quick, and reliable method for determining urinary creatinine by liquid chromatography/tandem mass spectrometry (LC/MS/MS) was developed and used to evaluate spot urine samples collected for the Washington Environmental Biomonitoring Survey (WEBS): part of the Washington State Department of Health, Public Health Laboratories (PHL). 50 µL of urine was mixed with a 1 : 1 acetonitrile/water solution containing deuterated creatinine as the internal standard and then analyzed by LC/MS/MS. Utilizing electrospray ionization (ESI) in positive mode, the transition ions for creatinine and creatinine-d3 were determined to be 114.0 to 44.1 (quantifier), 114.0 to 86.1 (qualifier), and 117.0 to 47.1 (creatinine-d3). The retention time for creatinine was 0.85 minutes. The linear calibration range was 20–4000 mg/L, with a limit of detection at 1.77 mg/L and a limit of quantitation at 5.91 mg/L. LC/MS/MS and the colorimetric Jaffé reaction were associated significantly (Pearson r = 0.9898 and R 2 = 0.9797, ρ ≤ 0.0001). The LC/MS/MS method developed at the PHL to determine creatinine in the spot urine samples had shorter retention times, and was more sensitive, reliable, reproducible, and safer than other LC/MS/MS or commercial methods such as the Jaffé reaction or modified versions thereof.

Direct Determination of Urinary Creatinine by Reactive-Thermal Desorption-Extractive Electrospray-Ion Mobility-Tandem Mass Spectrometry.

A direct, ambient ionization method has been developed for the determination of creatinine in urine that combines derivatization and thermal desorption with extractive electrospray ionization and ion mobility-mass spectrometry. The volatility of creatinine was enhanced by a rapid on-probe aqueous acylation reaction, using a custom-made thermal desorption probe, allowing thermal desorption and ionization of the monoacylated derivative. The monoacyl creatinine [M + H]+ ion (m/z 156) was subjected to mass-to-charge selection and collision induced dissociation to remove the acyl group, generating the protonated creatinine [M + H]+ product ion at m/z 114 before an ion mobility separation was applied to reduce chemical noise. Stable isotope dilution using creatinine-d3 as internal standard was used for quantitative measurements. The direct on-probe derivatization allows high sample throughput with a typical cycle time of 1 min per sample. The method shows good linearity (R2 = 0.986) and repeatability (%RSD 8–10%) in the range of 0.25–2.0 mg/mL. The creatinine concentrations in diluted urine samples from a healthy individual were determined to contain a mean concentration of 1.44 mg/mL creatinine with a precision (%RSD) of 9.9%. The reactive ambient ionization approach demonstrated here has potential for the determination of involatile analytes in urine and other biofluids.

Amperometric Determination of Creatinine with a Dialysis Membrane‐Covered Nitrobenzene/Water Interface for Urine Analysis

AbstractThe ion transfer of creatinine (CRE) at a polarized nitrobenzene (NB)/water (W) interface has been studied. When the pH of the W phase is in the range of 1.2 to 4.0, a well‐defined voltammetric wave is observed for a simple transfer of CRE+ (protonated form) at the NB/W interface. This transfer reaction has been applied to develop an amperometric method for the determination of CRE in urine. In this system, the NB/W interface is covered with a dialysis membrane to prevent possible interference from urine proteins. The concentration of CRE in a urine control has successfully been determined.