Liquid Urine Research Articles

Analysis of Globotriaosylceramide (Gb3) in Liquid Urine: A Straightforward Assay Using Tandem Mass Spectrometry.

Fabry disease (FD) is a lysosomal storage disorder caused by variants in the GLA gene encoding α-galactosidase A, an enzyme required for catabolism of globotriaosylceramide (Gb3). Accumulation of Gb3 in patients' cells, tissues, and biological fluids causes clinical manifestations including ventricular hypertrophy, renal insufficiency, and strokes. This protocol describes a methodology to analyze urinary Gb3 and creatinine. Samples are diluted with an internal standard solution containing Gb3(C17:0) and creatinine-D3, centrifuged, and directly analyzed by ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) using an 8.7-min method. Eight Gb3 isoforms [C16:0, C18:0, C20:0, C22:1, C22:0, C24:1, C24:0, and (C24:0)OH] are analyzed and the total is normalized to creatinine. Confirmation ions are monitored to detect potential interferences. The Gb3 limit of quantification is 0.023 µg/ml. Its interday coefficients of variation (3 concentrations measured) are ≤15.4%. This method minimizes matrix effects (≤6.5%) and prevents adsorption or precipitation of Gb3. Urine samples are stable (bias <15%) for 2 days at 21°C, 7 days at 4°C, and 4 freeze/thaw cycles, whereas prepared samples are stable for 5 days at 21°C, and 14 days at 4°C. The Gb3/creatinine age-related upper reference limits (mean + 2 standard deviations) are 29 mg/mol creatinine (<7 years) and 14 mg/mol creatinine (≥7 years). This simple, robust protocol has been fully validated (ISO 15189) and provides a valuable tool for diagnosis and monitoring of FD patients. © 2024 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Analysis of urinary globotriaosylceramide (Gb3) and creatinine by UHPLC-MS/MS Support Protocol 1: Preparation of the urinary quality controls Support Protocol 2: Preparation of the urine matrix used for the Gb3 calibration curve Support Protocol 3: Preparation of the Gb3 calibrators Support Protocol 4: Preparation of the working solution containing the internal standards Support Protocol 5: Preparation of the creatinine calibrators Support Protocol 6: Preparation of the UHPLC solutions and mobile phases.

Accuracy and precision of dried urine spot method for the detection of Schistosoma mansoni circulating cathodic antigens in resource-limited settings

BackgroundThe World Health Organization recommends the use of Schisto point-of-care circulating cathodic antigens (Schisto POC-CCA) for screening of Schistosoma mansoni as it offers better sensitivity than microscopy. However, there are limitation facing the use of this method including timely availability of the test cassettes. The aim of this study was to determine the reliability of dried urine spot (DUS) method for collection of urine and detection of S. mansoni using Schisto POC-CCA cassettes in a resource-limited settings.MethodsA cross-sectional study was conducted between October and November 2022 among 250 primary school children in Sengerema District, northwestern Tanzania. S. mansoni CCA was detected in filter paper-based DUS, liquid urine using DUS Schisto POC-CCA (index), and direct urine Schisto POC-CCA (comparator) methods respectively. S. mansoni eggs in stool were detected using duplicate Kato-Katz (KK) method. The measures of accuracy were computed and compared between the index and comparator methods. The strength of agreement between inter-raters precisions was tested using Cohen’s kappa (k).ResultsThis study revealed S. mansoni prevalence rates of 28.8%, 54.0% and 50.8% by duplicate KK, direct urine Schisto POC-CCA and DUS Schisto POC-CCA methods respectively. The mean intensity of infection among infected participants was 86.3 eggs per gram of stool (EPG) ranging from 12.0 EPG to 824.0 EPG. The sensitivity of DUS Schisto POC-CCA and direct urine Schisto POC-CCA was 94.44% (95% CI: 89.15–99.74%) and 97.22% (95% CI: 93.43–100.00%) respectively. The DUS Schisto POC-CCA method had slightly higher specificity (66.85%) than direct urine Schisto POC-CCA method (63.48%). The accuracy of the DUS Schisto POC-CCA was found to be slightly high (74.80%, 95% CI: 68.94–79.06%) compared to that of direct urine Schisto POC-CCA (73.20%, 95% CI: 67.25–78.59%). There was good agreement between two laboratory technologists who performed the DUS Schisto POC-CCA method on similar samples (k = 0.80, 95% CI: 0.59–0.95).ConclusionsThe DUS Schisto POC-CCA method had comparable S. mansoni detection accuracy to direct urine Schisto POC-CCA. This suggests that the method could be a potential alternative to direct urine Schisto POC-CCA for screening S. mansoni in resource-limited situations.Graphical

Cancer detection in dogs using rapid Raman molecular urinalysis.

The presence of cancer in dogs was detected by Raman spectroscopy of urine samples and chemometric analysis of spectroscopic data. The procedure created a multimolecular spectral fingerprint with hundreds of features related directly to the chemical composition of the urine specimen. These were then used to detect the broad presence of cancer in dog urine as well as the specific presence of lymphoma, urothelial carcinoma, osteosarcoma, and mast cell tumor. Urine samples were collected via voiding, cystocentesis, or catheterization from 89 dogs with no history or evidence of neoplastic disease, 100 dogs diagnosed with cancer, and 16 dogs diagnosed with non-neoplastic urinary tract or renal disease. Raman spectra were obtained of the unprocessed bulk liquid urine samples and were analyzed by ISREA, principal component analysis (PCA), and discriminant analysis of principal components (DAPC) were applied using the Rametrix®Toolbox software. The procedure identified a spectral fingerprint for cancer in canine urine, resulting in a urine screening test with 92.7% overall accuracy for a cancer vs. cancer-free designation. The urine screen performed with 94.0% sensitivity, 90.5% specificity, 94.5% positive predictive value (PPV), 89.6% negative predictive value (NPV), 9.9 positive likelihood ratio (LR+), and 0.067 negative likelihood ratio (LR-). Raman bands responsible for discerning cancer were extracted from the analysis and biomolecular associations were obtained. The urine screen was more effective in distinguishing urothelial carcinoma from the other cancers mentioned above. Detection and classification of cancer in dogs using a simple, non-invasive, rapid urine screen (as compared to liquid biopsies using peripheral blood samples) is a critical advancement in case management and treatment, especially in breeds predisposed to specific types of cancer.

A new adsorptive 3D–printed sampling device for simultaneous determination of 63 urinary organic acids by LC–MS/MS

BackgroundThe detection and quantification of urinary metabolites play an important role in disease diagnosis. In most cases, urinary analyses are done with liquid urine samples, which must be quickly transported to the laboratory to avoid metabolites degradation that is associated with temperature fluctuations. Consequently, dried sampling devices have emerged to minimize analyte degradation. However, most commercial dried sampling devices are expensive, aggregate low volumes, and need better analytical sensitivity. Therefore, a new dry urine sampling device that is inexpensive, suitable for domestic sampling operation, and efficient for quantifying metabolites without requiring high–resolution instruments is proposed in the present study. ResultsThe newly designed dry urine sampling device was produced by 3D printing that efficiently determines 63 urinary organic acids using liquid chromatography coupled with mass spectrometry (LC–MS/MS). The system's efficiency was demonstrated with analytical figures of merit, such as precision, accuracy, and stability of analytes after the sampling and storing of ordinary urine samples. The limits of quantification ranged from 0.01 to 0.42 ng mL–1. Precision and accuracy tests showed relative standard deviations of less than 15 %. The urine stability in the sampling device was high within seven days without any significant degradation of the metabolites. The method was applied to the analysis of 10 human urine samples and compared to a conventional method without the use of the sampling device. The results showed no statistically significant differences, demonstrating the method's efficiency. SignificanceThe proposed 3–D printing device was developed with fast, low–cost manufacturing features and can be manufactured with different volumetric capacities, adaptable to the needs of each user. Furthermore, it is innovative because this is the first sampling device that is effective for the simultaneous storage and preservation of several important urinary metabolites. Thus, it is anticipated that its application would contribute significantly to the identification of metabolic disorders.

Dried urine spot (DUS) applied for sampling prior to the accurate HILIC-MS/MS determination of 14 amino acids

Urine amino acid analysis has proven valuable for an array of clinical or nutritional studies. However, transportation of liquid urine sample shows certain disadvantages, such as possible leakage, need for cold chain and thus higher costs for their transport. Utilization of dried urine spots (DUS) can offer an interesting alternative. In the present study, a method was developed for the determination of 14 amino acids in DUS including the testing of in-house collection device and drying of the sample before analysis. Normal filter paper was tested as the means for sample collection. Absorption and extraction experiments were performed on 3 different types of filter paper, with 3 different extraction solvents and two different solvent volumes. The solvents used were mixtures of common analytical solvents (methanol, water, acetonitrile) using total volumes of 1 mL and 1.5 mL. Finally, 1 mL of acetonitrile: methanol: water 40:40:20 (v/v/v) was chosen as the optimal system. Analysis was performed on a UHPLC-MS system, using stable isotope labeled internal standards. Method validation included the study of limits of detection (LOD) and quantification (LOQ), linearity ranges, precision, matrix effect, extraction recovery, precision, and stability for each analyte. The obtained results were satisfactory, thus enabling application of the proposed method as an alternative to the analysis of liquid urine. Further utilization of DUS can offer advantages by enabling patient centric sampling even in long distances far from the analytical laboratories.

Fate of enteric bacteria and viruses in silt loam soil amended with biofertilizers made from human feces and urine for crop production

Human excreta can be used as biofertilizers due to their nutrient and organic matter content. Nevertheless, the behavior of microorganisms should be investigated, as enteric pathogens can accumulate in the soil. Therefore, we evaluated the survival and transport of two enteric bacteria (E. coli and Salmonella enterica) and two enteric virus surrogates (MS2 and ΦX-174 bacteriophages) when applied as contaminants of four biofertilizers on Lactuca sativa production. The study was carried out in lysimeters outdoors with repacked silt loam soil, using a randomized block design (RBD) with six treatments and three replicates. Biofertilizers were urea-treated feces, feces composted with organic waste, stored liquid urine, and struvite and were inoculated at 106–107 cfu/pfu g−1 of E. coli and bacteriophages strains and 107–109 of S. enterica. Soil amended with composted feces exhibited the lowest decimal reductions (T90) for the microorganisms evaluated, with 10.2 days for E. coli, 11.9 days for S. enterica, 22.2 days for MS2, and 3.7 days for ΦX-174. Urea-treated feces temporarily hindered the growth and survival of E. coli and S. enterica in the soil. However, both bacteria were present after one month, while MS2 showed a stable concentration with this biofertilizer. E. coli presented a stable behavior when applied with urine, whereas MS2 was stable with urea-treated feces, urine, and struvite. ΦX-174 and S. enterica were not considered good representatives of most resistant enteric pathogens’ behavior in the soil during food production since they were not detected in leachate and showed a rapid die-off in soil. E. coli leached faster and in higher concentrations than MS2, which presented higher concentrations in deeper soil layers, from 10 to 20 cm. Therefore, using urine, struvite, and urea-treated faces implied a higher chance of infection, as a decimal reduction time was not observed during the 36 days of the experiment. As a result, in short-term cultures, it is recommended that additional hygiene barriers are taken to avoid infection since E. coli and MS2 were still detected at the end of the experiment. An interval of 90 days between fertilizing and harvesting is recommended to prevent pathogen infection. These results are the basis for a quantitative risk assessment of human excreta-based biofertilizers for food production.

In-vial dried urine spot collection and processing for quantitative analyses.

Analysis of dried urine spots (DUSs) is becoming an emerging technique in clinical, toxicological, and forensic chemistry due to the fully non-invasive collection, facile transportation, and simple storage of DUS samples. Correct DUS collection and elution is of the utmost importance because inadequate DUS sampling/processing may have direct consequences on quantitative DUS analyses and these aspects were, for the first time, comprehensively investigated in this contribution. Various groups of endogenous and exogenous species were selected as model analytes and their concentrations were monitored in DUSs collected on standard cellulose-based sampling cards. Strong chromatographic effects were observed for most analytes having a crucial impact on their distribution within the DUSs during sampling. Concentrations of target analytes were up to 3.75-fold higher in the central DUS sub-punch in comparison to the liquid urine. Consequently, substantially reduced concentrations of these analytes were determined in peripheral DUS sub-punches demonstrating that sub-punching, often applied to dried material spots, is not acceptable for quantitative DUS analyses. Hence, a simple, rapid, and user-friendly procedure was suggested, which employed an in-vial collection of a known urine volume on a pre-punched sampling disc (using a low-cost micropipette designed for patient-centric clinical sampling) and in-vial processing of the whole DUS. Excellent accuracy (0.20%) and precision (0.89%) of liquid transfers were achieved by the micropipette, which was also applied to remote DUS collection by laic and expert users. The resulting DUS eluates were analysed by capillary electrophoresis (CE) for the determination of endogenous urine species. The CE results demonstrated no significant differences between the two user groups, elution efficiencies of 88-100% (in comparison to the liquid urine), and precision better than 5.5%.

Application of Dried Urine Spots for Non-Targeted Quadrupole Time-of-Flight Drug Screening.

The use of dried urine spots (DUS) can simplify sample handling, shipment and storage when compared to liquid urine samples. To prepare DUS, a small amount of urine is pipetted on a filter paper card. The subsequent drying of the specimen can prevent the post-sampling formation or degradation of substances (e.g., caused by bacteria). To evaluate the potential of DUS screening, 17 authentic urine samples, containing a broad range of substances, were extracted and analyzed on a Sciex TripleTOF® 5600+ System using a non-targeted screening and library searching approach. The screening results were compared to the analysis of the same urine sample in liquid form, using the same high-resolution liquid chromatography--quadrupole time-of-flight mass spectrometry method. More than 65 different legal and illegal drugs were successfully identified within the investigated 17 urine samples using the DUS screening approach. When compared to the analysis of liquid urine, the following compounds could not be identified: 1x ecgonine methyl ester, 1x nicotine, 1x promazine and 1x 11-nor-9-carboxy-∆9-tetrahydrocannabinol. Overall, 95.2% of the target substances that have been detected in liquid urine were identified correctly using the DUS approach. In conclusion, DUS screening offers a simple, cost-effective and easier sample handling alternative to the traditional use of liquid urine and provides the detection of the most important substances for forensic requirements. Furthermore, the DUS sample preparation can be fully automated (sample documentation, internal standard application and extraction).

Development of a kit for urine collection on filter paper as an alternative for Pompe disease screening and monitoring by LC-HRMS.

Pompe disease (PD) is an inborn error of metabolism caused by α-glucosidase acid enzyme deficiency. It significantly impacts patients' health and life quality and may lead to death in the first few years of life. Among the well-established diagnostic methods, urinary glucose tetrasaccharide (Glc4) screening by high performance-liquid chromatography has been helpful in monitoring Glc4 levels in patients on enzyme replacement therapy, demonstrating therapy efficacy. However, the specimen shipping process from a sample collecting location to a specialized laboratory for monitoring the Glc4 is costly and presents preanalytical challenges. In this work, we developed a filter paper based-urine collection kit to facilitate specimen shipment, and liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) analysis to determine Glc4 and creatinine in dried urine on filter paper. The LC-HRMS was based on a combination of targeted and untargeted screening on the same specimen injection and was successfully developed and validated. Bland-Altman statistics revealed a good relationship between dried and liquid urine samples and Glc4 and creatinine. Glc4 and other metabolites in dried urine showed stability for at least 7 days at 4 and 22 °C, and 3 days at 50 °C. The stability of the analytes and the efficiency of the kit were tested simulating real conditions by sending it by post. After two days in transit without refrigeration, the stability of compounds was maintained, showing the reliability of the urine collection kit and analysis method to determine the PD biomarker Glc4.

Quantitation of ethanol in UTI assay for volatile organic compound detection by electronic nose using the validated headspace GC-MS method.

Disease detection through gas analysis has long been the topic of many studies because of its potential as a rapid diagnostic technique. In particular, the pathogens that cause urinary tract infection (UTI) have been shown to generate different profiles of volatile organic compounds, thus enabling the discrimination of causative agents using an electronic nose. While past studies have performed data collection on either agar culture or jellified urine culture, this study measures the headspace volume of liquid urine culture samples. Evaporation of the liquid and the presence of background compounds during electronic nose (e-nose) device operation could introduce variability to the collected data. Therefore, a headspace gas chromatography-mass spectrometry method was developed and validated for quantitating ethanol in the headspace of the urine samples. By leveraging the new method to characterize the sample stability during e-nose measurement, it was revealed that ethanol concentration dropped more than 15% after only three measurement cycles, which equal 30 minutes for this study. It was further shown that by using only data within the first three cycles, better accuracies for between-day classification were achieved, which was 73.7% and 97.0%, compared to using data from within the first nine cycles, which resulted in 65.0% and 81.1% accuracies. Therefore, the newly developed method provides better quality control for data collection, paving ways for the future establishment of a training data library for UTI.

High resolution, high accuracy non-targeted LC-HR-MS/MS dried urine spot screening for drug of abuse testing

Screening for drugs of abuse in urine samples collected during roadside traffic testing is common practice in Switzerland. The use of an onsite rapid test for the detection of drugs and their metabolites is generally limited by a restricted panel of detectable drugs or substance classes (e.g. buprenorphine, benzodiazepines, cocaine, EDDP, morphine or opiates) with limited sensitivity and specificity. To improve the screening for legal and illegal drugs, the already collected urine sample typically undergoes an additional screening routine in a forensic laboratory. By using dried urine spots (DUS), sample handling, shipment, and storage could be substantially simplified, when compared to the sample handling of liquid urine. To prepare a DUS, the driver's urine is pipetted on a prepared paper card. The subsequent drying of the specimen, starting immediately after its collection, can prevent the post-sampling formation or degradation of substances (e.g. caused by bacteria). Furthermore, the DUS can be stored and shipped at ambient temperature and need no cooling or freezing. To evaluate the potential of DUS screening, 17 authentic urine samples containing multiple substances, were pipetted (10 μL) onto Autocollect TFN filter paper cards (Ahlstrom, France). After drying, the DUS cards were extracted (6 mm Ø extraction head) into glass vials with an insert (100 μL, methanol/water, 70/30, v/v), using a fully automated CAMAG DBS-MS 500 HCT autosampler in combination with a PAL RSI 3 robot. Subsequently, the prepared extracts were analyzed on a Sciex 5600 TOF instrument using a non-targeted screening and library searching approach. Thereby, 1 μL of the extract was injected onto a Kinetex C8 column and a ballistic gradient was used (mobile phase A: H2O+ 0.1% HCOOH, mobile phase b: MeCN + 0.1% HCOOH). A run time of 15 minutes was applied. To evaluate the performance of DUS, the screening results were compared to the analysis of the same urine sample in liquid form, using the same method on the same LC-MS instrument (injection of 1 μL of a 1/10 diluted urine). More than 50 different legal and illegal drugs were successfully identified within the investigated 17 urine samples using the DUS screening approach: 6-MAM, alprazolam, amoxicillin, amphetamine + metabolite, bisoprolol, bupropion + metabolite, caffeine + metabolites, carbamazepine + metabolites, citalopram + metabolite, cocaethylene, cocaine + BE + EME, codeine + metabolite, fentanyl, gabapentin, ketamine + metabolites, lacosamide, lamotrigine, levetiracetam, lidocaine, lorazepam + metabolite, losartan, metamizole-metabolites, metformin, methadone + EDDP, methylphendiate, metoclopramide, metoprolol, midazolam + metabolite, morphine + metabolite, nicotine + cotinine, ondansetron, oxazepam, paracetamol + metabolites, phenytoin, pregabalin + metabolite, promazine, propranolol, quetiapine + metabolites, sildenafil + metabolite, THC-COOH, torasemide, tramadol + metabolite, tranexamic acid, trazodone-metabolite, trimipramine + metabolites, venlafaxine-metabolite, zolpidem + metabolite. When compared to the analysis of liquid urine, the following compounds could not be identified: 2× nicotine, 1× promazine, 1× THC-COOH. Overall, 96% of the target substances that have been detected in liquid urine were identified correctly using the DUS approach. The DUS contain 10 μL of urine, therefore the signal intensity compared to liquid urine analysis, where we used 30 μL, is on average 68% lower. In most cases all substances were identified correctly using the DUS screening approach. Because of lower signal intensities for the DUS in one case of nicotine, promazine and THC-COOH were not properly identified. DUS Screening offers a simple, cost-effective and easier sample handling alternative to the traditional use of liquid urine. Based on the obtained results, a pilot study has been initiated (during roadside traffic control by the local police) in order to verify the usability in the forensic workflow.

Study on the Bioelectricity of Human’s Urine

Human urine can be used to test the urine bioelectricity by taking a graphite electrode pair out of dry cells, putting fresh urine into a small cup, and assembling a urine bioelectricity. A number of conditions of the human body were analysed, including family relationships, age, gender, health status, diet habits, and how much water one drinks during a day. Additionally, we examined the electrical properties of not fresh urine depending on the number of days it was stored with or without refrigeration. A cotton cloth soaked with urine is used to change the form of urine for testing to prevent liquid urine inconvenience. By examining the discharge characteristics of human urine, we can better understand our health status through this study.

Detection of Cytomegalovirus in Urine Dried on Filter Paper.

Urine is the best specimen for the diagnosis of congenital cytomegalovirus, but collection and processing of liquid urine are impractical for screening. Urine dried on filter paper was processed by the same convenient, low-cost method used by newborn screening to test blood spots and showed high sensitivity and specificity.

Reliability of a dried urine test for comprehensive assessment of urine hormones and metabolites

BackgroundMass spectrometry allows for analysis of multiple hormone and organic acid metabolites from small urine volumes; however, to assess the full extent of daily hormone production, 24-h urine collections are usually required. The aims of this study were, first, to confirm that mass spectrometric analysis of an array of hormones and organic acids would yield similar results in both liquid and dried urine, and, second, to determine if collection of four dried spot urine samples could be substituted for a 24-h collection when measuring reproductive hormones.MethodsTwo study populations were included in this prospective observational study. Twenty individuals collected both a spot liquid urine and dried urine on filter paper to analyze eight organic acids. A second group of 26 individuals collected both a 24-h urine and four dried spot urines during waking hours throughout the same day for evaluation of 17 reproductive hormones and metabolites; data from 18 of these individuals were available to compare liquid versus dried urine results. Dried urine was extracted, hydrolyzed, and derivatized before analysis by mass spectrometry; all analytes from dried urine were normalized to urine creatinine.ResultsReproductive hormone results from dried and liquid urine were in excellent agreement with intraclass correlation coefficients (ICCs) greater than 0.90; comparison of dried to liquid urine for organic acids showed good to excellent agreement (ICC range: 0.75 to 0.99). Comparison between the 4-spot urine collection and 24-h urine collection methods showed excellent agreement (ICC > 0.9) for 14 of the 17 urine metabolites and good agreement for the others (ICC 0.78 to 0.85) with no systematic differences between the two methods of collection.ConclusionsThe burden of urine collection can be reduced using collection of four spot dried urines on filter paper without compromising comparability with hormone results from a 24-h urine collection. A large number of urine analytes can be assessed from the dried urine with similar results to those from liquid urine. Given the ease of sample handling, this 4-spot dried urine assay would be useful for both clinical assessment of patients and for large epidemiologic studies.

Dried urine and salivary profiling for complete assessment of cortisol and cortisol metabolites

Aims/objectivesThe primary aim of this study was to determine the utility of dried urine sampling in obtaining measures of cortisol and cortisol metabolites. Additional aims were to evaluate if a 4-spot dried urine collection is representative of a 24-hour urine collection and if expected diurnal cortisol patterns can be observed in samples from both urine and saliva. MethodsData from individuals with cortisol measures available from both a 4-spot dried urine collection and a 24-hour urine collection (n = 28) were evaluated. Of these 28, 20 also had concurrent liquid and dried 24-hour urine measures. Consistency between these methods was evaluated using paired t-tests and intraclass correlation coefficients (ICCs). In addition, data from individuals with concurrent measures of both urinary and salivary cortisol (n = 68) were assessed for consistency in the diurnal pattern of change in cortisol. ResultsNear ideal consistency was observed between liquid and dried urine for measures of total urine free cortisol, total urine cortisone, and total cortisol metabolites (n = 20; ICCs = 0.99, 0.97 and 0.96, respectively). Good to excellent consistency was observed between the 4-spot method and the 24-hour collection (n = 28; ICCs = 0.89, 0.95 and 0.92, respectively). In mixed model analysis, no difference was seen in the diurnal pattern of cortisol between salivary and urinary free cortisol (n = 68; P = 0.83). ConclusionDried urine is a viable alternative to liquid urine for the measurement of cortisol and cortisol metabolites. Additionally, if the 4 measures are added together, 4-spot urine collections can be representative of diurnal cortisol patterns commonly assessed using saliva and 24-hour urine collections.

Long-term follow-up with filter paper samples in patients with propionic acidemia.

BackgroundPropionic acidemia (PA) is an inherited disorder caused by deficiency of propionyl CoA carboxylase. Most patients with this disorder are diagnosed during the neonatal period because of severe metabolic acidosis and hyperammonemia. Patients are required to undergo blood and urine analysis at least 3 to 4 times per year, depending on age and metabolic control.MethodsWe designed a prospective study in which we investigated the results from blood and urinary samples collected monthly in filter paper from 10 PA patients followed in a single metabolic reference center from January 2015 to September 2017. The aim of this study was to evaluate the usefulness of filter paper samples in the follow‐up of the PA patients.ResultsDuring the follow‐up period, 163 dried blood spot (DBS) and 119 urine dried spot samples were analyzed and compared with 160 plasma and 103 liquid urine specimens; 64 specimens of plasma were analyzed for odd‐numbered long‐chain fatty acids (OLCFAs). A total of 40 metabolic crises, 18 of them with hyperammonemia were documented. We observed a strong correlation between the filter paper and the urine/plasma samples for the main PA parameters both in stable metabolic conditions as well as in acute decompensations. Also, there was a strong correlation between OLCFAs measured in plasma and quantification of odd number acylcarnitines in DBS.ConclusionsWe conclude that filter paper blood and urinary samples can be used for the follow‐up of the patients with PA, correctly reflecting their metabolic situation.

Raman chemometric urinalysis (Rametrix) as a screen for bladder cancer.

Bladder cancer (BCA) is relatively common and potentially recurrent/progressive disease. It is also costly to detect, treat, and control. Definitive diagnosis is made by examination of urine sediment, imaging, direct visualization (cystoscopy), and invasive biopsy of suspect bladder lesions. There are currently no widely-used BCA-specific biomarker urine screening tests for early BCA or for following patients during/after therapy. Urine metabolomic screening for biomarkers is costly and generally unavailable for clinical use. In response, we developed Raman spectroscopy-based chemometric urinalysis (Rametrix™) as a direct liquid urine screening method for detecting complex molecular signatures in urine associated with BCA and other genitourinary tract pathologies. In particular, the RametrixTM screen used principal components (PCs) of urine Raman spectra to build discriminant analysis models that indicate the presence/absence of disease. The number of PCs included was varied, and all models were cross-validated by leave-one-out analysis. In Study 1 reported here, we tested the Rametrix™ screen using urine specimens from 56 consented patients from a urology clinic. This proof-of-concept study contained 17 urine specimens with active BCA (BCA-positive), 32 urine specimens from patients with other genitourinary tract pathologies, seven specimens from healthy patients, and the urinalysis control SurineTM. Using a model built with 22 PCs, BCA was detected with 80.4% accuracy, 82.4% sensitivity, 79.5% specificity, 63.6% positive predictive value (PPV), and 91.2% negative predictive value (NPV). Based on the number of PCs included, we found the RametrixTM screen could be fine-tuned for either high sensitivity or specificity. In other studies reported here, RametrixTM was also able to differentiate between urine specimens from patients with BCA and other genitourinary pathologies and those obtained from patients with end-stage kidney disease (ESKD). While larger studies are needed to improve RametrixTM models and demonstrate clinical relevance, this study demonstrates the ability of the RametrixTM screen to differentiate urine of BCA-positive patients. Molecular signature variances in the urine metabolome of BCA patients included changes in: phosphatidylinositol, nucleic acids, protein (particularly collagen), aromatic amino acids, and carotenoids.

Automated analysis of dried urine spot (DUS) samples for toxicology screening

BackgroundDried specimens have been proposed in multiple environments to minimize costs associated with specimen storage and shipping in clinical studies. This report describes the development and validation of an automated method for qualitative toxicology screening of dried urine samples using LC–MS/MS. MethodsUrine standards containing 41 compounds were prepared and applied to filter paper cards. Dried urine was eluted from the cards using a Dried Blood Spot (DBS) autosampler from Spark Holland, which was plumbed inline with a Thermo Scientific Turboflow chromatography system for subsequent MS/MS detection with selected reaction monitoring. Limits of detection, precision of peak areas, repeatability, and carryover studies were conducted. Concordance with a reference LC–MS/MS method using liquid samples was evaluated using remnant discarded specimens. ResultsThe limit of detection ranged from 5 to 75 ng/mL for most compounds. At the LOD for each analyte, the peak area precision ranged from 8 to 29%. For 20 repeat injections of samples spiked at ±25% of the LOD, there was a 4% false positive rate for the 75% × LOD samples, and a 0.4% false negative rate for the +125% × LOD samples. In comparing 40 known positive specimens analyzed with the DUS method and a liquid urine reference method, there was 88% agreement. Analysis of 10 known negative specimens yielded negative results. There was no significant carryover detected up to 2000 ng/mL for any of the analytes in the assay. ConclusionUsing a robotic DUS sampling an inline HTLC–MS/MS system, we have developed and validated a fully-automated and robust method for multi-analyte detection of drugs of abuse in dried urine specimens.

Pengaruh Pemberian Pupuk Cair Urin Kambing Etawa terhadap Pertumbuhan Bawang Merah

This study aims to determine the effect of giving urine liquid fertilizer Etawa on the growth of shallots (Allium ascalonicum L.). The research method used was an experiment using a completely randomized design (CRD), consisting of 5 treatments and 4 replications, namely treatment A (0 ml of goat urine / polybag), treatment B (50 ml of goat urine / polybag), treatment C (100 ml of goat urine / polybag), D treatment (150 goat urine / polybag), E treatment (200 ml / l goat urine / polybag). Observations were made on red onion plant height, wet western tubers, number of tubers, and tuber diameter. The results of the study, F count 1.38 &lt; F table 3.06 (5%), wet weight F count 0.8 &lt; F table 3.06 (5%), number of tubers F count 0.5 &lt; F table 3. 06 (5%), and diameter of shallot bulbs F count 0.51 &lt; F table 3.06 (5%). Conclusion, the administration of liquid urine fertilizer Etawa goat did not affect the height of the shallot plant&#x0D; Keywords: etawa goat urine, shallots

Feasibility of measuring sodium, potassium and creatinine from urine sample on dried filter paper.

Aim: To develop a bioassay for estimation of sodium, potassium and creatinine in dried urine strips and comparing with their respective concentration in liquid urine samples. Materials & methods: Urine was collected on filter paper strips, dried at room temperature and, eluted for estimation of sodium, potassium by indirect ion selective electrode method and creatinine by Jaffé method. Result: This bioassay was validated based on the US FDA guidelines for bioanalytical method validation and was linear, sensitive, accurate and precise with acceptable recovery and matrix effects. Analytes were stable in dried urine strips during 1 year of storage at 4°C. Conclusion: We conclude that the dried urine is suitable for analysis of sodium, potassium and creatinine and offers a convenient alternative for monitoring dietary salt intake.