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1
PADOAN, ANDREA. "Statistical methods for mass spectrometry data analysis and identification of prostaste cancer biomarkers." Doctoral thesis, Università degli Studi di Milano-Bicocca, 2014. http://hdl.handle.net/10281/50248.
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BACKGROUND
Prostate Cancer (PCa) is the most common cancer among males in Europe. Patients developing early PCa sometimes refer non-specific symptoms, namely lower urinary tract symptoms (LUTS), and they usually undergo medical investigations based on Prostate Specific Antigen (PSA) and Digital Rectal Examination (DRE). Suspicious results of one or both testings are prerequisite to Prostate Biopsy. However, due to PSA low sensitivity/specificity in predicting positive prostate biopsy, the identification of new PCa biomarkers is actually a real need. MALDI-TOF/MS protein profiling could be a valuable technology for biomarkers identification. However, up to now its use is laden with lack of reproducibility that confounds scientific inferences and limits its broader use.
AIMS
Goal of this study is to analyze urine collected after prostatic massage in patients referring LUTS, to identify candidate biomarker for PCa, by using MALDI-TOF/MS. We considered important aspects of MALDI-TOF/MS label-free proteomic profiling, in order to assess features reproducibility and to propose appropriate strategy to handle both measurement error and limit of detection (LOD) problems. The study results should aid in reducing the number of worthless first-biopsied and assist Urologists on differential diagnosis of PCa.
METHODS
In a cross-sectional study, we collected urine obtained after DRE from 205 patients that referred LUTS to consultants at the Urological Unit at University of Padova. All patients undergone to prostate biopsy for suspicious PCa. Urines were dialyzed and analyzed by MALDI-TOF/MS in reflectron mode. For the MALDI-TOF/MS reproducibility evaluation, we analyzed a urine pooled from 10 reference samples, spiked with 12.58 pmol of a 1589.9 m/z internal standard (IS) peptide. For the inter-run variability assessment, 14 aliquots were dialyzed by MALDI-TOF/MS. For the intra-run study, an aliquot was divided into 26 separate sub-aliquots and analyzed by MALDI- TOF/MS. To estimate the signal detection limit (sLOD), serial dilution up to 1/256 of a urine pool were analyzed in triplicate. We evaluated the sLOD and adjusted the data appropriately to reduce its variability. We investigated six data normalization approaches - the mean, median, internal standard, relative intensity, total ion current and linear rescaling normalization. Between-spectrum and the overall spectra variability were evaluated by the coefficient of variation (CV). An optimized signal detection strategy was also evaluated to overcome peak detection algorithms errors. Measurement errors and with-in subject variances were evaluated by an external dataset, made of urine repeatedly collected from 20 reference subjects. Intra class correlation coefficient (ICC), Regression Calibration (RCAL) and SIMEX analyses were used to estimate unbiased logistic regression coefficients relating MALDI-TOF/MS features with Patients biopsy outcome. Monte Carlo simulations were used to estimate influence of different LOD adjustment methods on ICC and RCAL.
RESULTS
Initially, we evaluated the intra- and inter-run on data obtained from automatic peak detection. Normalization methods performed almost similarly in both studies, except IS, which resulted in an increased CV. Calculated sLOD varied with spectra m/z. After sLOD adjustment, raw and normalized data showed a reduction in CVs, while median and mean normalizations performed better, especially in the intra-assay study. However, by optimizing the peak signal detection, the overall features variability drastically decreased. Median normalization with sLOD correction remained the preferable choice for further analyses. Evaluating the external dataset, we found that most of the MALDI-TOF/MS variability is intrinsic to the biological matrix. By using substitution of below LOD values by LOD/2, simulation studies showed that ICC estimations were poorly affected by LOD, when measurement error σ is less that 0.36 and values below LOD are less that 50%. Comparing results from naïve logistic regression, RCAL and SIMEX, measurement error appeared to cause a "bias toward the null". However, SIMEX estimations seemed to correct for a smaller amount of bias than RCAL. Overall, we found eight MALDI-TOF/MS features associated with positive biopsy results.
CONCLUSION
Findings from the reproducibility study showed that the major contributing factor for MALDI-TOF/MS profiling variability is the peak detection process. So, a new algorithm suited for MALDI-TOF reflectron mode is desirable for its applications in profiling studies. However, normalization strategies aid in increasing MALDI-TOF/MS label-free data reproducibility, especially with sLOD correction. Despite urine does not seem to be a promising biological fluid for proteomic biomarker discovers, RCAL and SIMEX appeared valuable approaches to obtain regression coefficients adjusted for biological and instrumental errors on MALDI-TOF/MS features.
2
Rocha, Diogo Librandi da. "Desenvolvimento de procedimento analítico em fluxo com multicomutação para a determinação espectofotométrica de ácido úrico em urina." Universidade de São Paulo, 2009. http://www.teses.usp.br/teses/disponiveis/46/46133/tde-05102009-105307/.
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A mecanização de procedimentos analíticos em análises clínicas traz vantagens tais como minimização de erros sistemáticos e do tempo das análises. Sistemas de análises em fluxo com multicomutação apresentam grandes potencialidades nesse sentido, atendendo às necessidades da mecanização de procedimentos analíticos de maneira versátil e robusta. Estes sistemas permitem minimizar o consumo de reagentes e a geração de resíduos, devido ao gerenciamento preciso de pequenos volumes de soluções por dispositivos controlados eletronicamente, tais como microbombas solenoide. O fluxo pulsado proporcionado pelas microbombas e a estratégia da amostragem binária melhoram a mistura entre amostra e reagentes. O ácido úrico é o principal produto final do metabolismo de purinas. A determinação deste analito em amostras de urina apresenta importância clínica, uma vez que sua concentração pode auxiliar no diagnóstico de disfunções no organismo humano, como a gota e o mau funcionamento dos rins. Um procedimento analítico empregando sistema de análises em fluxo com microbombas solenoide foi desenvolvido para a determinação de ácido úrico em amostras de urina. Os íons Cu(II) são reduzidos pelo ácido úrico a íons Cu(I), que podem ser quantificados por espectrofotometria na presença do ácido 4,4\'-dicarboxi-2,2\'- biquinolínico (BQA). Resposta linear foi observada entre 10 e 100 µmol L-1 de ácido úrico, sendo a curva analítica representada pela equação A=(0,0063±0,0002)CAU + (0,0285±0,0040), r = 0,999, em que CAU é a concentração de ácido úrico em µmol L-1. O limite de detecção foi estimado em 3,0 µmol L-1 (99,7% de nível de confiança; n = 20). O coeficiente de variação foi estimado em 1,2% com 20 medidas de uma solução de ácido úrico 75 µmol L-1 e a frequência de amostragem foi de 150 h-1. As principais espécies concomitantes presentes na urina não interferem na determinação de ácido úrico em concentrações até 5 vezes maiores que as usualmente encontradas. Recuperações entre 91 e 112% foram estimadas e os resultados das análises de 4 amostras de urina concordaram com os obtidos pelo procedimento enzimático para a determinação de ácido úrico (95% de nível de confiança). O alto grau de diluição da amostra necessário (100 vezes) minimiza o volume de amostra utilizado e os efeitos de matriz. Uma simples reconfiguração do sistema e a reotimização das frações volumétricas permitiram que a amostra fosse diluída em linha por reamostragem na zona dispersa. Resposta linear foi observada até 5,0 mmol L-1 de ácido úrico, sendo a curva analítica obtida representada pela equação A=(0,105±0,001) CAU + (0,023±0,003), r=0,999, em que CAU é a concentração de ácido úrico em mmol L-1. O coeficiente de variação, o limite de detecção e a frequência de amostragem foram estimados em 1,0%, 0,2 mmol L-1 e 95 h-1, respectivamente. Os resultados da análise de 3 amostras de urina concordaram com os obtidos pelo procedimento enzimático, com nível de confiança de 95%Mechanization of analytical procedures in clinical analysis brings advantages such as minimization of systematic errors and analysis time. Multicommuted flow systems attain the requirements to mechanization of analytical procedures in a versatile and robust way, minimizing reagent consumption and waste generation, due to the low solution volumes handled by electronically controlled devices, such as solenoid micro-pumps. The pulsed flow characteristic of the micro-pumps and the binary sampling approach improve sample and reagent mixing. Uric acid is the main end product of purine metabolism and its determination in urine shows clinical importance, because its concentration can be related to human organism dysfunctions, such as gout and renal disorders. An analytical procedure employing a flow system with solenoid micro-pumps was developed, aiming the determination of uric acid in urine samples. Cu(II) ions are reduced by uric acid to Cu(I) ions that can be quantified by spectrophotometry in the presence of 2,2´-biquinoline 4,4´-dicarboxylic acid (BCA). Linear analytical response was observed between 10 and 100 µmol L-1 uric acid and the analytical curve corresponds to the equation A=(0.0063±0.0002) CUA + (0.0285±0.0040), r = 0.999, in which CUA is the uric acid concentration in µmol L-1. The detection limit was estimated as 3.0 µmol L-1 (99.7% confidence level; n = 20). The coefficient of variation was estimated in 1.2% with 20 replicates of a 75 µmol L-1 uric acid solution and sampling rate of 150 h-1 was achieved. The main concomitant species does not interfere in uric acid determination in concentrations up to 5-fold higher than that usually found in urine samples. Recoveries from 91 to 112% were estimated and the results for 4 urine samples agreed with those obtained by the commercially available enzymatic kit for determination of uric acid (95% confidence level). The 100-fold sample dilution minimizes sample consumption and matrix effects. A simple system reconfiguration and a re-optimization of volumetric fractions attained on-line sample dilution by zone sampling. Linear response was observed up to 5.0 mmol L-1 uric acid and the analytical curve corresponds to the equation A=(0.105±0.001) CUA\' + (0.023±0.003), r = 0.999, in which CUA\' is the uric acid concentration in mmol L-1. The coefficient of variation, detection limit and sampling frequency were estimated as 1.0%, 0.2 mmol L-1 and 95 h-1, respectively. The results of the analysis of 3 urine samples also agreed with those obtained with the enzymatic procedure at the 95% confidence level
3
Lough, Patricia Schechter. "Use of urine samples for ethanol analysis." CSUSB ScholarWorks, 1989. https://scholarworks.lib.csusb.edu/etd-project/446.
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4
Paquin, Olivier. "La microalbuminurie chez le sujet âgé." Bordeaux 2, 1994. http://www.theses.fr/1994BOR2M113.
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Jansson, Emelie. "Gaskromatografisk metod för analys av GHB i urin." Thesis, Department of Physics, Chemistry and Biology, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-19651.
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En metod för detektering och kvantifiering av gamma-hydroxysmörsyra (GHB) i urin med gaskromatografi (GC) är framtagen på Sahlgrenska universitetssjukhuset. Metoden är relativt unik då den inte kräver upparbetning i form av derivatisering, indunstning eller extraktion. Urinen surgörs med koncentrerad saltsyra och internstandard, gamma-valerolakton, tillsätts. GHB övergår då till laktonformen, gamma-butyrolakton (GBL). Därefter injiceras provet direkt på en GC-FID med en kapillärkolonn för glykoler och alkoholer. Detektion ner till 100 μmol/L är möjligt med en variationskoefficient mellan 6 och 12 %. Provsvar erhålls efter 6,5 minuter. Metoden är dock inte fullständig då en del frågetecken kvarstår. Bland annat bör det undersökas om andra föreningar, som kan förekomma i urin, kan eluera samtidigt som GHB. Om ja så bör vidare analyser genomföras för att separera GHB och den andra föreningen. Metoden kan däremot användas i nuläget som en screeninganalys för att snabbt få ett svar på om GHB finns närvarande eller inte. Verifiering kan sedan ske med GC-MS.
A method for determination and quantification of gamma-hydroxyburyric acid (GHB) in urine samples is developed at Sahlgrenska universitetssjukhus. No time consuming procedures as derivatization and exctration is required, which makes the method fairly unique. Hydrochloric acid and internal standard, gamma-valerolakton, is added to the urine sample before the sample is injected to a gas chromatograph with a flame ionization detector and a column for glycols and alcohols. The hydrochloric acid makes the GHB convert into gamma-butyrolactone (GBL) which is easier to separate in the gas chromatograph. Limit of detection was found to be 100 μmol/L and test result is received after 6,5 minutes. There are still some question marks around the method, for example, there is a possibility that another substance elute at the same time as GHB. More tests are required to determine whether or not it is so. For now the method can be used as a screening analysis to hastily detect GHB presence. Verification can be done with GC-MS.
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Cooper, Mark Thomas. "A chromatographic method for detecting phenolic metabolites of carbosulfan in urine." Thesis, Queensland University of Technology, 1989. https://eprints.qut.edu.au/35977/1/35977_Cooper_1989.pdf.
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The inability of the conventional blood cholinesterase
test to reliably detect carbamate pesticide poisoning in
humans prompted the investigation of an alternative
surveillance method. Rapid, micro-scale sample treatment
procedures were developed to extract the phenolic
metabolites of carbosulfan from urine, convert these
compounds to their dinitrophenyl ether derivatives and
determine their concentrations quantitatively by nitrogen
selective gas-liquid chromatography. This method was
capable of detecting micro-gram levels of metabolites and
performed to an accuracy of<+/ 10% and precision of<
6% RSD.
In vivo experiments we re undertaken in which carbosul fan
was administered to laboratory rats and the effects of
dosage and sampling time on the level of phenolic
metabolites in urine were examined. These results
provide guidelines for human exposure however absolute
confidence in these thresholds will only occur when the
data base of human experience is collected and correlated
to metabolite levels in urine.
Urine samples were drawn and analyzed from potentially
exposed personnel handling carbosulfan and in all cases
no phenolic metabolites were detected.
7
Chen, Hui-Chuen. "The urinary excretion of mercapturic acids in free-living adult males." Thesis, This resource online, 1991. http://scholar.lib.vt.edu/theses/available/etd-12052009-020010/.
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Allen, Robert Douglas III. "Development of an assay for the detection of cytomegalovirus in urine." Thesis, Georgia Institute of Technology, 1993. http://hdl.handle.net/1853/25410.
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Hoang, Tiffany Truc. "Speciation and identification of low molecular weight organoselenium metabolites in human urine." Diss., Georgia Institute of Technology, 2003. http://hdl.handle.net/1853/30671.
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Silva, Júnior Jarbas Miguel da. "Excreção urinária de derivados de purinas e de compostos nitrogenados de zebuínos em pastejo." Universidade Federal de Viçosa, 2014. http://locus.ufv.br/handle/123456789/5825.
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This study aimed evaluating the excretion of purine derivatives and nitrogen compounds in zebu cattle grazing, on different days and times within days. The experiment was conducted in the cattle department of the Federal University of Viçosa / MG, using five Nelore heifers with an average body weight of 300 ± 15 kg and 20 months of age, in 5x5 Latin square design. The experimental treatments were defined to represent those commonly used in the dry season, as follows: control (mineral salt), concentrated with 20.31% crude protein (CP) on dry matter (DM) being offered (OF) level of 0.5 to 1% of body weight fasted (BWF) OF5 and OF10, respectively; and two concentrated self- regulating (SR) consumption, containing 69.38% CP on a DM basis (20% urea and 20% salt) offered ad libitun (SR70) and other concentrate containing 39.73% CP based on MS being offered ad libitum (SR40). The experimental periods was 18 days, with one day to perform 14 hours of fasting for weighing and adjustment of the quantities supplied, 12 days for adaptation to the experimental diets and five for total collection of urine and stool sample at the times of 0h00a.m. to 4h00a.m, 4h00a.m. to 8:00a.m., 8:00a.m. to 12:00p.m., 12:00p.m. to 4:00p.m., 4:00p.m. to 8:00p.m. and 20:00p.m. to 0:00a.m. For total collection of urine was used probe Folley number 26, coupled to polyethylene hose leading to a urine collection bag for urine closed system, which was emptied every two hours in the range of 8:00a.m. to 8:00p.m., and every four hours in the range from 8:00p.m. to 8:00a.m. and subsequently homogenized and cooled. The collected urine sampling was performed every four hours, measuring the volume, and withdrawing one sample were diluted in H 2 SO 4 at 0,036N and another don't diluted. To estimate fecal output, used the titanium dioxide, provided the total daily amount of 15g, between 9 th and 18 th day of each period. To estimate the intake of pasture, used the indigestible neutral detergent fiber (iNDF) as internal indicator. Was performed by collecting pasture technique for determining the square potentially digestible dry matter (PDDM) on the third day of each experimental period, and on days 14 th and 18 th was held grazing simulation to estimate the consumption of constituents of diets. In urine samples the concentrations of creatinine, total nitrogen, urea, uric acid and allantoin. For statistical analysis we used the statistical program SAS Proc Mixed. Dry matter intake 10was higher (P<0.05) for the treatment OF10 compared to SR70, SR40 and control treatments but was not different (P>0.05) treatment OF5. The CP intake increased by supplementation (P<0.05), which caused no effect on DM intake from pasture. Excretions of creatinine did not change treatment, day and sampling period (P>0.05) and had a mean of 23.03 ± 0.30 mg / kgPC. Urinary relations of allantoin (Al) and uric acid (UA) with creatinine were not affected (P>0.05) by treatments, collection days and times of collection. The total nitrogen relations:creatinine and urea nitrogen:creatinine in urine showed interaction (P<0.05) between treatment and sampling period. The relationship between urea nitrogen:total nitrogen was influenced (P<0.05) only at time of collection. The nitrogen balance (NB) in g/day did not differ between treatments OF10, SR70 and SR40, however these had higher retention of N (P<0.05) than treatments OF5 and control, which were not different. The NB, in g/ging, showed differences (P<0.05) between treatments with concentrated, which did not differ (P> 0.05), and control treatment, with the lowest NB. The production of microbial N was not affected (P>0.05) by treatments. The microbial efficiency gPBmic/kgMOD and gPBmic/kgNDT was affected (P<0.05) by supplementation, being higher (P<0.05) for OF5, OF10 and SR70 treatments, which did not differ. The control and OF5, treatments had the lowest values were similar. The lack of effect of day and the collection period on allantoin and uric acid compared with creatinine has wide practical application, enabling use spot urine sample to calculate the excretion of purine derivatives at any time of day or night, and consequently the microbial production. Depending on the variations observed for total nitrogen and urea nitrogen relations with creatinine over 24 hours is not recommended the use of a single spot urine sample for determination of these nitrogen compounds.
O presente trabalho foi desenvolvido com os objetivos de avaliar a excreção dos derivados de purinas e de compostos nitrogenados em zebuínos em pastejo, em diferentes dias e períodos dentro de dias. O experimento foi conduzido no setor de gado de corte da Universidade Federal de Viçosa/MG, utilizando-se cinco novilhas Nelore com peso corporal médio de 300 ± 15kg e 20 meses de idade, distribuídas em quadrado latino 5x5. Os tratamentos experimentais foram definidos para representar aqueles normalmente utilizados na época seca do ano, sendo eles: controle (sal mineral), concentrado com 20,31% de proteína bruta (PB) com base na matéria seca (MS) sendo oferecido (OF) em nível de 0,5 e 1% do peso corporal em jejum (PCJ), OF5 e OF10, respectivamente; e dois concentrados autorreguladores (AR) de consumo, um contendo 69,38% PB com base na MS (20% de ureia e 20% de sal) ofertado ad libitun (AR70) e outro concentrado contendo 39,73% PB com base na MS sendo ofertado ad libitum (AR40). Os períodos experimentais possuíram 18 dias, sendo o dia um para realização de jejum de 14 horas para pesagem e ajuste das quantidades fornecidas, 12 para adaptação dos animais às dietas experimentais e cinco para a coleta total de urina e amostral de fezes, nos horários das 0h00 às 4h00, 4h00 às 8h00, 8h00 às 12h00, 12h00 às 16h00, 16h00 às 20h00 e 20h00 às 24h00. Para a coleta total de urina utilizou-se sonda de Folley no26, acoplada a mangueira de polietileno que conduziu a urina até uma bolsa coletora de urina por sistema fechado, que foi esvaziada a cada duas horas no intervalo das 8h00 às 20h00, e a cada quatro horas no intervalo das 20h00 às 8h00, sendo posteriormente homogeneizadas e resfriadas. A amostragem da urina coletada foi realizada a cada 4 horas, medindo-se o volume e retirando-se duas amostras, uma diluída com solução H2SO4 0,036N e não diluida. Para determinação da excreção fecal, utilizou-se o dioxido de titânio, fornecido na quantidade total diária de 15g, entre os dias 9 e 18 de cada período. Para estimativa do consumo de pasto, utilizou-se a fibra indigestível em detergente neutro (FDNi), como indicador interno. Realizou-se coleta de pasto pela técnica do quadrado para determinação da matéria seca potencialmente digestível (MSpd) no terceiro dia de cada período experimental, e nos dias 14o e 18o realizou-se simulação de pastejo para estimar os consumos dos constituintes das dietas. Nas amostras de urina foram determinadas as concentrações de creatinina, nitrogênio total, ureia, acido úrico e alantoína. Para análise estatística utilizou-se o programa estatístico Proc Mixed do SAS. O consumo de MS foi superior (P<0,05) para o tratamento OF10 em relação aos tratamentos AR70, AR40 e controle, mas não diferiu (P>0,05) do tratamento OF5. O consumo de PB aumentou com a suplementação (P<0,05), que não causou efeito sobre o consumo de MS do pasto. As excreções de creatinina não sofreram efeito de tratamento, dia e período de coleta (P>0,05) e apresentaram média de 23,03 ± 0,30 mg/kgPC. As relações urinárias da alantoína (Al) e do ácido úrico (AU) com a creatinina não foram influenciadas (P>0,05) pelos tratamentos, dias de coleta e horários de coleta. As relações nitrogênio total:creatinina e nitrogênio ureico:creatinina na urina apresentaram interação (P<0,05) entre tratamento e período de coleta. A relação entre nitrogênio ureico:nitrogênio total foi influenciada (P<0,05) apenas pelo horário de coleta. O balanço de compostos nitrogenados (BN), em g/dia, não diferiu entre os tratamentos OF10, AR70 e AR40, contudo esses apresentaram maiores retenções de N (P<0,05) que os tratamentos OF5 e controle, que não foram diferentes. O BN, em g/ging, apresentou diferença (P<0,05) entre os tratamentos com concentrado, que não diferiram entre si (P>0,05), e tratamento controle, que apresentou o menor BN. A produção de compostos nitrogenados microbianos não foi alterada (P>0,05) pelos tratamentos. A eficiência microbiana, em gPBmic/kgMOD e gPBmic/kgNDT foi afetada (P<0,05) pela suplementação, sendo maior (P<0,05) para os tratamentos OF5, OF10 e AR70, que não diferiram entre si. Os tratamentos controle e OF5, apresentaram os menores valores e foram semelhantes entre si. A ausência de efeito de dia e do período de coleta sobre a relação alantoína e ácido úrico com a creatinina tem grande aplicação prática, possibilitando utilizar a amostra spot de urina para calcular a excreção de derivados de purinas em qualquer horário do dia ou da noite, e consequentemente a produção microbiana. Em função das variações observadas para as relações nitrogênio ureico e nitrogênio total com a creatinina ao longo do período de 24 horas não se recomenda o uso de uma única amostra spot de urina para determinação destes compostos nitrogenados.
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Abdelrazig, Salah M. A. "Mass spectrometry for high-throughput metabolomics analysis of urine." Thesis, University of Nottingham, 2015. http://eprints.nottingham.ac.uk/30600/.
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Direct electrospray ionisation-mass spectrometry (direct ESI-MS), by omitting the chromatographic step, has great potential for application as a high-throughput approach for untargeted urine metabolomics analysis compared to liquid chromatography-mass spectrometry (LC-MS). The rapid development and technical innovations revealed in the field of ambient ionisation MS such as nanoelectrospray ionisation (nanoESI) chip-based infusion and liquid extraction surface analysis mass spectrometry (LESA-MS) suggest that they might be suitable for high-throughput metabolomics analysis. In this thesis, LC-MS and high-throughput direct ESI-MS methods using high resolution orbital trap mass spectrometer were developed and validated for untargeted metabolomics of human urine. Three different direct ESI-MS techniques were explored and compared with LC-MS: flow injection electrospray ionisation-MS (FIE-MS), chip-based infusion and LESA-MS of dried urine spots on a cell culture slide. A high-throughput sample preparation protocol was optimised using in-house artificial urine. Urine samples after consumption of green tea and healthy controls were used as a model to explore the performance and classification ability of the direct ESI-MS. High-throughput data pre-processing and multivariate analysis protocols were established for each method. The developed methods were finally applied for the analysis of clinical urine samples for biomarker discovery and to investigate the metabolic changes in osteoarthritis and malaria. Also, the methods were applied to study the effect of oligofructose diet on the gut microbial community of healthy subjects. The analytical performance of the methods for urine metabolomics was validated using quality control (QC) and principal component analysis (PCA) approaches. Rigorous validation including cross-validation, permutation test, prediction models and area under receiver operating characteristic (ROC) curve (AUC) was performed across the generated datasets using the developed methods. Analysis of green tea urine samples generated 4128, 748, 1064 and 1035 ions from LC-MS, FIE-MS, chip-based infusion and LESA-MS analysis, respectively. A selected set of known green tea metabolites in urine were used to evaluate each method for detection sensitivity. 15 metabolites were found with LC-MS compared to 8, 5 and 6 with FIE-MS, chip-based infusion and LESA, respectively. The developed methods successfully differentiated between the metabolic profiles of osteoarthritis active patients and healthy controls (Q2 0.465 (LC-MS), 0.562 (FIE-MS), 0.472 (chip-based infusion) and 0.493 (LESA-MS)). The altered level of metabolites detected in osteoarthritis patients showed a perturbed activity in TCA cycle, pyruvate metabolism, -oxidation pathway, amino acids and glycerophospholipids metabolism, which may provide evidence of mitochondrial dysfunction, inflammation, oxidative stress, collagen destruction and use of lipolysis as an alternative energy source in the cartilage cells of osteoarthritis patients. FIE-MS, chip-based infusion and LESA-MS increased the analysis throughput and yet they were able to provide 33%, 44% and 44%, respectively, of the LC-MS information, indicating their great potential for diagnostic application in osteoarthritis. Malaria samples datasets generated 9,744 and 576 ions from LC-MS and FIE-MS, respectively. Supervised multivariate analysis using OPLS-DA showed clear separation and clustering of malaria patients from controls in both LC-MS and FIE-MS methods. Cross-validation R2Y and Q2 values obtained by FIE-MS were 0.810 and 0.538, respectively, which are comparable to the values of 0.993 and 0.583 achieved by LC-MS. The sensitivity and specificity were 80% and 77% for LC-MS and FIE-MS, respectively, indicating valid, reliable and comparable results of both methods. With regards to biomarker discovery, altered level of 30 and 17 metabolites were found by LC-MS and FIE-MS, respectively, in the urine of malaria patients compared to healthy controls. Among these metabolites, pipecolic acid, taurine, 1,3-diacetylpropane, N-acetylspermidine and N-acetylputrescine may have the potential of being used as biomarkers of malaria. LC-MS and FIE-MS were able to separate urine samples of healthy subjects on oligofructose diet from controls (specificity/sensitivity 80%/88% (LC-MS) and 71%/64% (FIE-MS)). An altered level of short chain fatty acids (SCFAs), fatty acids and amino acids were observed in urine as a result of oligofructose intake, suggesting an increased population of the health-promoting Bifidobacterium and a decreased Lactobacillus and Enterococcus genera in the colon. In conclusion, the developed direct ESI-MS methods demonstrated the ability to differentiate between inherent types of urine samples in disease and health state. Therefore they are recommended to be used as fast diagnostic tools for clinical urine samples. The developed LC-MS method is necessary when comprehensive biomarker screening is required.
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Bordin, Keliani. "Avaliação de biomarcadores da exposição humana à fumonisina B1 nos alimentos em municípios dos estados de São Paulo e Santa Catarina, Brasil." Universidade de São Paulo, 2015. http://www.teses.usp.br/teses/disponiveis/74/74132/tde-23042015-140349/.
Full textAbstract:
A fumonisina B1 (FB1) e uma micotoxina produzida pelo metabolismo secundário de espécies de Fusarium, principalmente F. verticillioides e F. proliferatum, os quais contaminam diversos alimentos antes e apos o processamento, sobretudo o milho e derivados, gerando graves problemas para a Saúde Pública e a qualidade dos alimentos. O objetivo deste trabalho foi avaliar a exposição humana a FB1 presente nos alimentos através da estimativa de ingestão da toxina na dieta e da análise de diferentes biomarcadores presentes em amostras de sangue, urina e cabelo. Além disso, foram investigados os efeitos da toxina através da avaliação de ácido fólico presentes em alimentos e em soro, e os níveis de uréia e creatinina presentes em soro. O estudo foi realizado em dois municípios dos Estados de São Paulo e Santa Catarina, cujos respectivos voluntários foram categorizados como de baixo consumo de derivados de milho (Grupo A, voluntários de Pirassununga/SP) e de alto consumo de derivados de milho (Grupo B, voluntários de Erval Velho/SC). As amostras de alimentos do Grupo A (Pirassununga/SP) foram fornecidas pelos voluntários (n=100) nos meses de Junho/2011, Setembro/2011, Dezembro/2011 e Marco/2012. Os voluntários do Grupo B (Erval Velho/SC) (n=20) forneceram amostras de alimentos no mês de Abril/2012. Em cada grupo, uma lista com 20 alimentos a base de milho foi entregue aos voluntários, para fornecimento de amostras daqueles disponíveis em suas respectivas residências em cada mês de amostragem, totalizando 122 amostras de derivados de milho no Grupo A e 17 amostras no Grupo B coletadas durante o estudo. Adicionalmente, aplicou-se um Questionário de Frequência Alimentar (QFA) e um Inquérito Recordatório de 24 horas (QIR - 24 h) no momento das coletas de amostras. Em cada mês de amostragem de alimentos, foram coletadas amostras de sangue, urina (somente Grupo A) e cabelo dos voluntários, sendo as amostras armazenadas a -20ºC (urina e cabelo) ou -80ºC (sangue) até o momento das análises. As amostras de alimentos foram submetidas a análise de FB1, sendo que as de farinha de milho foram também analisadas quanto ao teor de ácido fólico. Ambas as análises foram feitas através de cromatografia líquida de alta eficiência (CLAE). Em soro, foram avaliadas a relação esfinganina/esfingosina (Sa/So), resíduos de FB1, ácido fólico, uréia e creatinina. Em urina, foram analisados os níveis de FB1, creatinina para correção do volume urinário e a relação Sa/So. Em cabelo, foram analisados os resíduos de FB1 através de CLAE acoplada a espectrometria de massas. Todos os métodos de análise foram submetidos a procedimento de otimização e validação intra--laboratorial. A incidência de FB1 nos alimentos foi, em média, 72% (n=122) nas amostras do Grupo A (Pirassununga/SP) e 35% (n=17) no Grupo B (Erval Velho/SC). Os maiores níveis foram encontrados em amostras de pipoca provenientes do Grupo B, com uma amostra excedendo o limite de tolerância estabelecido no Brasil (2,500 µg kg-1). A ingestão diária provável média (IDPM) de FB1 no Grupo A foi de 63,3 ng kg-1 peso corpóreo (p.c.) dia-1, que corresponde a 3,1% da ingestão provisória máxima tolerável (IPMT) recomendada para fumonisinas (2.000 ng kg-1 p.c. dia-1). A IDPM do Grupo B apresentou uma média de 190,1 ng kg-1 p.c. dia-1 o que corresponde a 9,5% da IDMT. As concentrações de ácido fólico nas amostras de farinha de milho variaram de < 0,3 µg kg-1 (limite de quantificação do método) a 1.705 µg kg-1, com média de 713 ± 435 µg kg-1. Somente uma amostra apresentou nível de ácido fólico acima do valor mínimo estabelecido pela ANVISA. Em urina, a incidência de FB1 foi de 33,4% (n=251), com níveis médios de 3,19 ± 3,15 ng mg-1 de creatinina. Não houve correlação (P>0,05) entre as concentrações de FB1 na urina e nos alimentos. Os níveis de esfinganina foram mais elevados em mulheres, com 25,0% (n=116) de amostras positivas, em comparação à urina de homens, 10,4% (n=96). A relação Sa/So apresentou em média 0,91, 0,77 e 0,89 para urina de mulheres, homens e em combinação, respectivamente. Em soro, os níveis de esfingosina foram em média 2,48 ng mL-1 para o Grupo A e 5,01 ng mL-1 para o Grupo B. A relação Sa/So variou de 0,06 a 3,19 com média de 0,79 para o Grupo A e 0,78 para o Grupo B. Embora tenha havido correlação positiva (r=0,574, P<0,05) entre a relação Sa/So no soro e os dados de consumo de milho e derivados obtidos no QIR-24 h, não foram observadas correlações (P>0,05) entre a ingestão de FB1 e a relação Sa/So na urina ou soro. A concentração de ácido fólico no soro variou de 6,7 a 24,0 ng mL-1 (média de 13,4 ± 5,4 ng mL-1), com ambos os grupos (A e B) apresentando resultados dentro dos valores de referências. Não foram observados níveis detectáveis de FB1 nas amostras de soro. No entanto, FB1 foi detectada em 4 amostras de cabelo humano (7,2%) dos Grupos A e B, cuja concentração média foi de 21,3 ± 12,1 ng g-1. Em síntese, os resultados obtidos nas análises de biomarcadores de FB1 no presente trabalho estão de acordo com os valores de IDPM encontrados, indicando que a exposição a FB1 nas populações estudadas não representa um risco a saúde.Fumonisin B1 (FB1) is a mycotoxin produced by the secondary metabolism of Fusarium species, mainly F. verticillioides and F. proliferatum, which contaminates foods before and after processing and causes serious problems to public health and food quality. The aim of this study was to evaluate the human exposure to FB1 in food by means of estimated intake of toxin in the diet, and analysis of different biomarkers in serum, urine and hair. In addition, folic acid in food and blood as well urea and creatinin in serum were investigated to evaluate the toxin effects. The study was conducted in two cities of Sao Paulo and Santa Catarina States, where the respective volunteers were categorized as low-consumers of corn products (Group A, volunteers from Pirassununga/SP) and high-consumers of corn products (Group B, volunteers from Erval Velho/SC). Food samples from Group A (Pirassununga/SP) were provided by volunteers (n=100) in June/2011, September/2011, December/2011 and March/2012. The volunteers from Group B (Erval Velho/SC) (n=20) provided food samples in April/2012. In each group, a list of 20 corn products was given to volunteers, to allow them to check and collect the food items available in their homes at each sampling time. The total number of samples of corn products provided by the volunteers were 122 and 17 in Group A and Group B, respectively. Addicionally, a Food Frequency Questionnaire (FFQ) and a 24-Hours Dietary Recall Questionnaire (24h-DRQ) were applied by the time of sample collections. In each month of food samples collection, samples of blood, urine (only Group A) and hair from the volunteers were collected and storage at -20ºC (urine and hair) or -80ºC (blood) until analysis. Food samples were submitted to determination of FB1, and corn meal samples were also evaluated for folic acid levels. Both analysis were performed by high performance liquid chromatography (HPLC). In serum, analyses included sphinganine/sphingosine ratio (Sa/So), FB1 residue, folic acid, urea and creatinine. In urine, the levels of FB1, creatinine to correct urinary volume and Sa/So ratio were evaluated. In hair, FB1 residues were analysed by HPLC coupled to mass spectrometry. All the analytical methods were submitted to optimization and intra-laboratorial validation procedures. The mean incidences of FB1 in corn products were 72% (n=122) in samples of Group A (Pirassununga/SP), and 35% (n=17) of Group B (Erval Velho/SC). The higher levels were found in popcorn from Group B, with one sample exceeding the tolerance limit established in Brazil (2,500 µg kg-1). The mean probable daily intake (PDIM) of FB1 in Group A was 63.3 ng kg-1 body weigh (b.w.) day-1, which corresponds to 3.1% of provisional maximum tolerable intake (PMTDI) recommended for fumonisins (2,000 ng kg-1 b.w. day-1). PDIM of Group B was 190.1 ng kg-1 b.w. day-1, which represents 9.5% of PMTDI. Folic acid levels in corn meal ranged from < 0,3 µg kg-1 (quantification limit) to 1.705 µg kg-1, with a mean of 713 ± 435 µg kg-1. Only one sample had levels of folic acid above the minimum established by ANVISA. In urine, the incidence of FB1 was 33,4% (n=251), at mean levels of 3,19 ± 3,15 ng mg-1 of creatinine. There wasn\'t correlation (P>0.05) between concentrations of FB1 in urine and foods. Sphinganine levels were higher in woman, with 25.0% (n=116) of positive samples in comparison to urine of men, 10.4% (n=96). The mean Sa/So ratios were 0.91, 0.77 and 0.89 for urine of women, men and in combination, respectively. In serum, sphingosine presented a mean of 2.48 ng mL-1 to Group A and 5.01 ng mL-1 to Group B. Sa/So ratio ranged from 0.06 to 3.19 with a mean of 0.79 to Group A and 0.78 to Group B. Although a positive correlation (r=0.574, P<0.05) was found between Sa/So ratio in serum and corn consumption data obtained by 24h-DRQ, no correlation was observed (P>0,05) with FB1 intake and Sa/So ratio in urine or serum. Folic acid concentration in serum ranged from 6.7 to 24.0 ng mL-1 (mean of 13.4 ± 5.4 ng mL-1), with both groups (A and B) presenting levels within the reference valuies. There were no detectable levels of FB1 in serum samples. However, FB1 was detected in 4 human hair samples (7.2%) of Groups A and B, at a mean concentration was 21.3 ± 12.1 ng g-1. In summary, the results obtained in the analyses of FB1 biomarkers in the present study are in agreement with the PDIM values found, hence indicating that FB1 exposure in the populations studied do not represent a health concern.
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West, Robert E. 1952. "Confirmation of urinary benzodiazepines by gas chromatography/mass spectrometry." Thesis, The University of Arizona, 1989. http://hdl.handle.net/10150/277228.
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A new method is described for the quantitative analysis of urinary benzodiazepines by gas chromatography/mass spectrometry. Development work was aimed at satisfying federal requirements for methods used in forensic urine drug testing which have become the standard in the laboratory industry. Trimethylsilyl (TMS), tert-butyl-dimethylsilyl (TBDMS) and benzophenone derivatives were tested in the development of the new assay. TBDMS derivatives were found to be the most suitable for the analysis of six common benzodiazepine metabolites. Precision for all metabolites tested, as measured by the within run coefficient of variation, was less than 7% at 100 ng/ml (n = 15). Assay sensitivity varied with the specific analyte in the range of 5 to 10 ng/ml. Validation of the procedure included the reanalysis of benzodiazepine positive urine specimens obtained from a forensic drug testing laboratory and comparison of the results from the independent assays. These specimens were tested first by radioimmunoassay using a 100 ng/ml cutoff and then confirmed by GC/MS. Sensitivity was sufficient to confirm the presence of benzodiazepine metabolites in all specimens tested.
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Couchman, Lewis. "LC-MS/MS analysis of buprenorphine and norbuprenorphine in urine." Thesis, Queen Mary, University of London, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.511397.
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Hassan, Syed Saeed-Ul. "Rapid immunological methods for analysis of dexamethasone in equine urine." Thesis, University of Sunderland, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.245822.
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Joensuu, Frida. "Behov av surgörning av urin vid analys för kalcium, fosfat och magnesium." Thesis, Örebro universitet, Institutionen för hälsovetenskaper, 2021. http://urn.kb.se/resolve?urn=urn:nbn:se:oru:diva-93001.
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Studier har tidigare visat surgörning urinprover inför analys av kalcium, fosfat och magnesium inte är nödvändigt. Dessa analyter är viktiga att analysera vid njursjukdomar och det görs med instrumentet Advia Chemistry XPT (Siemens AG, München, Tyskland) på Universitetssjukhuset Örebro. Syftet med arbetet var att se om samma resultat erhölls för kalcium, fosfat och magnesium beroende på om urinen var surgjord eller inte inför analys. Tre metoder utvärderades beträffande behov av surgörning av urin: Metod 1; den nuvarande metoden, urinen surgjordes till pH 3-4. Metod 2; utan surgörning, urinen rumstempererades i 30 min före mätning. Metod 3; utan surgörning, urinen värmdes till 36ºC före varje mätning för att lösa upp eventuella saltkristaller. Alla metoderna utfördes på 30 prover och testades efter 0h, 24h, 3 dagar och 7 dagar. Resultaten sammanställdes i ett linjärt regressionsdiagram och ett Bland Altmann-diagram. Ett förutbestämt acceptansmål var att de genomsnittliga analysresultaten inte skulle ha en större skillnad än 10% vid jämförelse mellan metod 1 och metod 2 respektive metod 1 och metod 3, samt att eventuella avvikande enskilda prover skulle undersökas närmare. Fem av proverna höll inte den förutbestämda 10%-gränsen och mikroskoperades för att se om saltkristaller förekom och surgjordes för att se om provsvaren skulle förändras. I tre av fem prover kunde saltkristaller observeras. Detta gav slutsatsen att alla prover fortsatt måste surgöras eftersom det inte går att avgöra i förväg om provet måste surgöras eller inte eftersom ett tydligt samband inte kunde ses mellan proverna som inte höll 10%-gränsen och de som gjorde det.Studies have previously shown that acidification of urine samples before analysis for calcium, phosphate and magnesium may not be necessary. These analytes are important to monitor during kidney disease and are, at USÖ, detected using the instrument Advia Chemistry XPT (Siemens AG, München, Germany). The aim of this study was to examine whether the results from analyzing previously mentioned mineral levels would differ depending on whether the urine had been acidified prior to analysis or not. To examine this, three methods were used. Method 1; the current method, where urine is acidified to pH 3-4. Method 2; without acidification, the urine was warmed at room temperature before analysis. Method 3; without acidification, the urine was heated to 36°C before analysis. All methods were assayed on 30 samples which were all examined after 0h, 24h, 3 days and 7 days. Results were compiled using a linear regression diagram and a Bland Altmann diagram. The predetermined acceptance criterion was a maximum 10% difference between mean analyte levels found using methods 2 or 3, in comparison to using method 1. Five samples deviated from the remainder by breaking the 10% limit and were therefore scrutinized under microscope to search for salt crystals, before acidification and reanalysis. Crystals were detected in three of the five samples. As there was no clear connection between the deviating samples, there is no way of knowing prior to analysis whether acidification will be necessary or not, and it is therefore deemed a necessity to acidify all urine samples.
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Kirk, Jayne Marie. "Mass Spectrometric Analysis of Steroid Hormones for Application in Analysis of Bovine Urine." Thesis, University of York, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.485830.
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Analytical strategies for the identification and quantification of up to 14 androgenic steroids and up to 17 corticosteroids have been evaluated and applied to bovine urine. The two classes ofsteroid have been analysed both as the native species and as Girard P hydrazone derivatives. Triple quadrupole mass spectrometry, operated in multiple reaction monitoring mode, has permitted the development of methods that enable the simultaneous detection of a range ofandrogenic steroids and corticosteroids at the ng mL-1 level. For a non-targeted approach, screening for the presence of corticosteroids was performed on a time of flight mass spectrometer, where confirmation of the identities of corticosteroids was obtained from accurate mass information. Girard P hydrazone derivatives of androgenic steroids and corticosteroids are amenable to analysis by electrospray mass spectrometry. The presence of an ionic group at position C-3 ofthe steroids increases their response relative to the native species by up to 33 times for the androgenic steroids and up to 21 times for the corticosteroids. The derivatisation reaction has been shown to work effectively in bovine urine, with limits ofdetection determined as ::::; 1 ng mL-1 for both classes ofsteroid hydrazone. Ion trap mass spectrometry has proved to be an extremely powerful tool for the elucidation of dissociation pathways of steroids and their hydrazone derivatives. Analysis of androgenic steroids, androgenic steroid hydrazones and corticosteroid hydrazones using multistage tandem mass spectrometry has shown how the varying functionality of the steroids affects their dissociation pathways, and how comparisons between similar structures can aid the assignment of product ions. Multistage tandem mass spectrometry of the hydrazone derivatives provides a wealth of structure-specific product ions arising due to losses from either the steroid or hydrazine moiety, and detailed dissociation sequences have been established, enabling structure assignment. A complete method employing sample extraction and derivatisation followed by analysis using liquid chromatography-multistage tandem mass spectrometry has been developed to allow full characterisation and structure confirmation of the steroids present in urine.
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Stubbs, Christopher. "High performance liquid chromatographic analysis of erythromycin in serum and urine." Thesis, Rhodes University, 1985. http://hdl.handle.net/10962/d1004581.
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Erythromycin, a macrolide antibiotic used mainly against gram-positive bacteria has been in clinical use since 1952 (1). Previous pharmacokinetic data published on this antibiotic have been derived predominantly from microbiological assay techniques. However, these techniques are relatively imprecise as well as being non-specific and extremely tedious to perform. A novel high performance liquid chromatographic analysis of erythromycin in human serum and urine using U.V. detection at 200 nm and/or electrochemical detection using both an amperometric and a coulometric electrochemical detector is presented. The method involves a solid phase extraction procedure followed by a simple phase separation step and chromatography on a reverse phase column. In order to select the optimum U.V. detector for this analysis, five "state of the art" detectors were compared in terms of their signal-to-noise ratios at U.V. wavelengths between 200 and 210 nm. A known metabolite des-N-methylerythromycin is readily detectable using U.V. detection, whilst another metabolite/degradation product anhydroerythromycin is not seen using U.V. detection but is readily observable using an electrochemical detector. The method has a limit of sensitivity of 0.25 μg/mL and 1.00 μg/mL in serum and urine respectively (U.V. detection) and is sufficiently sensitive to monitor serum and urine concentrations of erythromycin in man after administration of a single 500 mg erythromycin stearate tablet.KMBT_363
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Kelly, Barbara M. "The analysis of biological fluids for acylcarnitines." Thesis, Open University, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.326566.
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Dumitrescu, Vlad Andrei. "Comparative analysis of biogas slurry and urine as sustainable nutrient sources for hydroponic vertical farming." Thesis, Linköpings universitet, Tema vatten i natur och samhälle, 2013. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-96368.
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Sustainable alternatives to using mined nutrients in agriculture must be found in order to limit environmental impacts such as eutrophication, habitat destruction and greenhouse gas emis-sions. Biogas slurry and urine recycled to hydroponic food production (a type of soilless agri-culture) have the potential of providing inorganic nitrogen and phosphorus, the main essential nutrients required for plant growth. A Life Cycle Inventory Assessment (LCI) methodology has been used to compare the systems of producing artificial fertilizer, biogas slurry and urine based nutrient solutions for the growth of Brassica rapa L. (Chinese cabbage) in the context of a large scale hydroponic vertical farm. Costs and energy requirements have been the basis of the comparison and results show that both biogas slurry and urine are considerably cheaper than the commercial alternative and based on the nutrient content they have the potential of being successful nutrient solutions after dilution and nutrient supplementation. Filtration might also be required in order to remove suspended particles and pathogens.
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Hannon, Sarrah. "Analysis of cocaine adulterants and their metabolites in real patient urine samples." Thesis, Boston University, 2013. https://hdl.handle.net/2144/12114.
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Thesis (M.S.)--Boston UniversityCocaine is one of the most common drugs of abuse in the United States today, but street-quality cocaine is decreasing in purity each year. This change in purity requires a shift in the focus of cocaine analysis in forensic laboratories. In recent years, many federal agencies have begun testing and profiling for the adulterants and diluents present in cocaine samples submitted as evidence. By analyzing the compounds present in the street-quality samples, forensic chemists may be able to track the cocaine back to its source, based on the unique identities of certain adulterants. Many of the adulterants currently being added to cocaine are dangerous on their own, even though they may enhance the effects of the cocaine. For this reason many doctors and forensic pathologists are interested in the identities of the adulterants present. Often times, a sample of the cocaine ingested may not be available for testing. Thus, there is a need for the development of methods to test for these adulterants and their metabolites in biological samples. The objective of this research is to develop extraction and instrumental analysis methods for several common cocaine adulterant metabolites, in an effort to create a geographical profile of human urine samples that tested positive for benzoylecgonine, a cocaine metabolite, and exploring the possible trends.
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Wu, Ruige, and 吴瑞阁. "Microchip-capillary electrophoresis with two-dimensional separation and isotachophoresis preconcentration for determining low abundanceproteins in human urine and dairy products." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2011. http://hub.hku.hk/bib/B46506044.
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FERREIRA, Pablo Gomes. "Avaliação do efeito da membrana de látex de Hevea brasiliensis no reparo de defeito da parede abdominal de rato." Universidade Federal de Alfenas, 2007. https://bdtd.unifal-mg.edu.br:8443/handle/tede/752.
Full textAbstract:
A hérnia incisional (HI) é uma das complicações tardias das laparotomias, podendo
acometer até 20% dos pacientes operados. A utilização de próteses representou um avanço
sem precedentes no reparo da HI. Contudo, não há ainda tratamento ideal para o reparo e
prevenção desse mal. A membrana de látex da seringueira demonstrou atividade cicatrizante e
angiogênica em estudos in vitro e in vivo. Assim, o objetivo deste trabalho foi avaliar o efeito
da membrana de látex sobre o reparo da parede abdominal lesada de ratos. Foram utilizados
ratos machos Wistar com peso de 340 ± 20 g, nos quais foi induzido um defeito na parede
abdominal de 1 x 2 cm, tendo a linha alba ao centro. Os animais foram divididos em dois
grupos experimentais: Controle (sem reparo) e Látex (reparo com a membrana) e submetidos
à eutanásia aos 3, 5, 7 e 28 dias após a indução do defeito. Os fragmentos da parede
abdominal foram retirados, divididos em duas partes e utilizados para análise histológica e
bioquímica. A análise histológica revelou que o látex induziu a proliferação de vasos
sangüíneos, o aumento de fibras musculares novas e a regeneração tecidual. Observou-se
ainda o aumento da densidade de fibroblastos e deposição de colágeno, o que é importante
para determinar o aumento da resistência a forças tensoras no tecido cicatricial, evitando a
deiscência da ferida e a instalação da HI. Houve aumento gradual de deposição e organização
das fibras colágenas durante o período de estudo. O látex induziu ainda a expressão dos
fatores de crescimento celular, FGF-2 e TGFb; a diminuição da expressão de IGF e PDGF e
não alterou a expressão de TGFa. A regeneração tecidual requer a influência de fatores de
crescimento celular e uma seqüência de eventos celulares, que resultam na regulação celular,
com proliferação, sobrevivência e diferenciação. Assim, a modulação de fatores de
crescimento pela membrana de látex, parece ter contribuído positivamente para a regeneração
do defeito da parede abdominal. Conclui-se que a membrana de látex da seringueira, quando
usada para reconstrução da parede abdominal de ratos, permite a regeneração tecidual, a
formação de tecido conjuntivo fibroso de reparação, podendo ser utilizada efetivamente na
hernioplastia.The incisional hernia (IH) formation is one complication after abdominal surgery, affecting up to 20% of the reoperated patients. The use of prosthesis increased the repair of IH, however there is not yet an ideal treatment to repair or prevent the IH. The latex membrane from rubber tree has demonstrated to be cicatrizing and angiogenic, both in in vitro and in vivo studies. The aim of this work was to evaluate the effect of the latex membrane on the repair of the injuried abdominal wall of rats. Male Wistar rats with 340 ± 20 g were used and it was induced an abdominal wall defect with 1 x 2 cm and the alba line at the centre. The animals were divided into two experimental groups: Control (without repair) and Latex (with membrane repair) and were euthanized at 3, 5, 7 e 28 days after the defect induction. The abdominal wall fragments were removed, divided into two parts and used for histological and biochemical analysis. The histological analysis revealed that the latex membrane induced blood vessels proliferation, new muscle fiber formation and tissue regeneration. It was also observed a fibroblast density and collagen deposition increasing that is important to increase the resistance to tension forces and prevent the incision dehisce and IH formation. There was also a gradual increasing on the deposition and organization of collagen fibers during the experiment. The latex membrane induced the expression of cellular growth factors, FGF-2 and TGFb; decreased the expression of IGF and PDGF and did not alter the TGFa expression. Tissue regeneration requires the influence of growth factors in a sequence of cellular events that results on the cell regulation, proliferation, surviving and differentiation. The modulation of growth factor by the latex membrane may have contributed positively to the abdominal wall defect regeneration. It can be concluded that the latex membrane from rubber tree, when used to reconstruct the rat abdominal wall, allows tissue regeneration, fiber connective tissue formation and may be used effectively in hernioplasty.
Fundação de Amparo à Pesquisa do Estado de Minas Gerais - FAPEMIG
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De, Kock Neil. "The development of direct infusion mass spectrometry method for analysis of small metabolites in urine." Thesis, Stellenbosch : Stellenbosch University, 2013. http://hdl.handle.net/10019.1/80213.
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Thesis (MSc)--Stellenbosch University, 2013.ENGLISH ABSTRACT: This study focused on the development of an analytical method whereby creatinine, creatine and caffeine could be determined quantitatively. Urine is the preferred body fluid for the analysis of metabolites that the body excretes after administration of medicinal and illicit drugs. The detection of these metabolites depends on the volume of water the patient has drunk or, in criminal cases, the amount of water the suspect may deliberately add to their urine to dilute it. Creatinine, whose concentration in urine has been found to correlate with muscle mass, is chosen as an endogenous control substance against which the metabolite concentration is compared. While high performance liquid chromatography with ultraviolet detection (HPLC–UV) is commonly selected for the analysis, the quality of chromatography is affected by the fact that creatinine, being highly polar, is not retained in the reversed-phase columns. Furthermore, urine contains many polar substances that elute with the solvent front along with creatinine, thereby grossly affecting HPLC measurements. Hydrophilic interaction chromatography (HILIC) is a good alternative, although these methods generally require extensive sample preparation. Direct infusion electrospray ionization mass spectrometry (DI–ESI–MS) is ideally suited to highly polar compounds and was selected for this work. Pneumatically assisted ESI is preferred above the standard ionization method of atmospheric pressure chemical ionization (APCI) since pneumatically assisted ESI disperses the solution into ion-containing aerosol droplets which do not promote online conversion of creatinine to creatine. The objective of this study was to develop a simple and sensitive DI–ESI–MS method for the determination of various compounds in urine with creatinine as analytical reference compound and internal standard (IS). The analytical method development includes addition of 1-methyl-3-phenylpropylamine as a primary IS to standard solutions as well as to urine samples, followed by direct infusion of the sample into a mass spectrometer to determine the absolute concentrations of creatinine, creatine and caffeine. After appropriate instrument conditions were established, linear graphs of analyte-IS signal intensity ratios were obtained. The ratio of the concentration of the analyte (drug or metabolite) to that of creatinine (as IS) may be used to determine analyte concentration in artificial samples and/or urine. This method is not affected by change in fluid volume or adulteration of urine samples because the analyte-to-creatinine ratio remains unchanged. As part of this study, the developed DI–ESI–MS method was compared with an LC–UV–MS method developed for this purpose.
AFRIKAANSE OPSOMMING: Hierdie studie fokus op die ontwikkeling van ‘n analitiese metode waardeur kreatinien, kreatien en kaffeïen kwantitatief bepaal kan word. Uriene is die voorkeur liggaamsvloeistof vir die analise van metaboliete wat deur die liggaam, na administrasie van mediese en onwettige middels, uitgeskei word. Die deteksie van hierdie metaboliete hang van die volume water af wat die pasiënt gedrink het, of in strafbare gevalle, die hoeveelheid water wat verdagtes met opset by hul uriene gevoeg het ten einde dit te verdun. Daar is bevind dat die konsentrasie van kreatinien in uriene met spiermassa korreleer, derhalwe is kreatinien as ‘n interne kontrolemiddel gekies waarmee die metaboliet-konsentrasie vergelyk kan word. Hoë-druk vloeistofchromatografie met ultravioletdeteksie (HPLC–UV) word algemeen vir die analise van kreatinien ingespan, maar die gehalte van die chromatografie word deur die hoogs polêre aard van kreatinien beïnvloed en het swak retensie in omgekeerde-fasekolomme tot gevolg. Bowendien, uriene bevat groot hoeveelhede polêre middels wat saam met kreatinien in die oplosmiddelfront elueer en sodoende HPLC-bepalings uitermatig beïnvloed. Hidrofiliese interaksiechromatografie (HILIC) is ‘n goeie alternatief, ofskoon omvangryke monster-voorbereidings algemeen vereis word. Direkte inspuitelektrosproei-ionisasiemassaspektrometrie (DI–ESI–MS) is ideaal geskik vir hoogs polêre stowwe en is vir hierdie studie gekies. Pneumatiese hulp-ESI word bo die standaard ionisasie-metode van lugdruk chemiese ionisasie (APCI) verkies weens pneumatiese hulp-ESI se vermoë om die oplosmiddel in aërosoldruppels wat ione bevat, te versprei – sonder die aanlynomskakeling van kreatinien na kreatien. Die doel van hierdie studie was om ‘n eenvoudige en sensitiewe DI–ESI–MS-metode te ontwikkel wat verskeie stowwe in uriene kan bepaal deur kreatinien as analitiese verwysingsmiddel en interne standaard (IS) vir die opstelling van ‘n IS-kalibrasiekurwe te gebruik. Die analitiese metode-ontwikkeling sluit die gebruik van 1-metiel-3-fenielpropielamien as primêre IS in. Die IS word tot standaard oplossings en urienemonsters gevoeg, gevolg deur direkte inspuiting van die monster in ‘n massaspektrometer om die absolute konsentrasies van kreatinien, kreatien en kaffeïen te bepaal. Lineêre kurwes van die seinintensiteitsverhouding van analiet tot IS is verkry na gepaste instrumentkondisies vasgestel is. Die verhouding van konsentrasie van die analiet (middel of metaboliet) tot dié van kreatinien (as IS) mag gebruik word om die analietkonsentrasie in die standaard oplossings en/of urienemonster te bepaal. Die metode word nie deur veranderinge in die vloeistofvolume of verwatering van urienemonsters beïnvloed nie, weens die analiet-tot-kreatinienverhouding wat onveranderd bly. ‘n LC–UV–MS-metode is voorts ontwikkel om die ontwikkelde DI–ESI–MS-metode se data te vergelyk.
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Low, Ann Stewart. "An evaluation of analytical procedures for detection of drug abuse with particular reference to opiates." Thesis, Robert Gordon University, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.242985.
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Kim, Yuni T. "The effects of broccoli on the excretion of urinary conjugates." Diss., Virginia Tech, 1992. http://hdl.handle.net/10919/38535.
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The effects of dietary broccoli on the body's ability to detoxify were studied in 18 male subjects between the ages of 22-40 years. The biological parameters used for measuring detoxification were the four major urinary conjugates, namely, mercapturates, sulfoconjugates, glucuronides, and amino acid conjugates. Dietary broccoli increased the urinary excretion of mercapturates and sulfoconjugates, but did not influence the excretion of glucuronides and amino acid conjugates. A significant linear trend was observed over the six-day broccoli diet treatment for both urinary mercapturate (P<0.005) and sulfoconjugate (P<0.0001) excretion. The linear trend for the mercapturate excretion was in a dose-dependent manner, resulting in a 1.3 and 2.1 fold increase by the third and sixth days, respectively, of the broccoli diet, compared to the control. For sulfoconjugates, an unexpected decrease was observed on the first day of the broccoli diet. However, within the six-day broccoli dietary treatment, a continuous increase in conjugate excretion was observed, resulting in a 2.5 fold increase by the sixth day compared to the first day. The excretion of sulfoconjugates was not necessarily dose-dependent, and increased excretion at the highest level of broccoli (500 g) could be due to a time effect. Overall, sulfoconjugate excretion was the highest (3.98-8.91 mmole/24 h) followed by the amino acid conjugates (3.06-5.99 mmole/24 h) and glucuronides (2.85-3.54 mmole/24 h). Mercapturate excretion was the lowest (0.16-0.34 mmole/24 h). In spite of its low excretion level, the level of urinary mercapturates appeared to be the most responsive urinary conjugate to the different levels of broccoli diet.Ph. D.
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Sena, Laís Cristina Santana. "Desenvolvimento de método analítico para determinação dos principais adulterantes da cocaína em urina humana." Universidade Federal de Sergipe, 2016. https://ri.ufs.br/handle/riufs/3944.
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Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPqCocaine is a stimulant that features a strong ability to cause dependence. Often adulterants are added to this drug in order to mimic its action or minimize its adverse effects. When there are other pharmacologically active components in the drug composition, severe problems can occur to users’ health, such as intoxication symptoms. Thus, the aim of this study was to develop a method for the determination of the main adulterants of cocaine (caffeine, levamisole, lidocaine, phenacetin, diltiazem, and hydroxyzine) in human urine. The high-performance liquid chromatography with a photodiode array detector and the dispersive liquid-liquid microextraction based on solidification of floating organic drop were used as analysis technique and as sample preparation technique, respectively. The reversed-phase chromatographic separation was obtained with a C18 column (250 x 4.6 mm; 5 μm; 80 Å) in gradient elution mode using acetonitrile-trifluoroacetic acid 0.026% (v/v) at 1 mL min-1 as mobile phase, at 25°C, and detection at 235 nm. The analysis time was 25 min. Under optimum conditions, human urine samples were alkalized with 0.5 mol.L-1 sodium phosphate buffer (pH 10) and added sodium chloride (20% m/v). Acetonitrile (150 μL) and 1-dodecanol (30 μL) were used as dispersive and extraction solvent, respectively. The method presented linear range of 312.5 – 3125 ng.mL−1 for caffeine and levamisole and 187.5 – 1875 ng.mL−1 for lidocaine, phenacetin, diltiazem, and hydroxyzine, with limit of quantification of 187.5 ng.mL-1 to lidocaine, phenacetin, diltiazem, and hydroxyzine and 312.5 ng.mL-1 for caffeine and levamisole. Recovery mean values were between 6.0 and 42.6%. The method showed good precision and accuracy, with within- and between-run relative standard deviation and relative error less than 15%. The samples were stable after freeze-thaw cycle and short-term room temperature stability tests. Additionally, this method was applied in samples of urine of five cocaine users and at least one adulterant was identified in all samples. It is expected that this method will contribute to the precision in the diagnosis of cocaine adulterants’ intoxication and to the proper planning of therapeutic measures.
A cocaína é uma droga estimulante que apresenta capacidade de causar dependência. Frequentemente são adicionados a esta droga adulterantes com o intuito de mimetizar sua ação ou minimizar seus efeitos adversos. Quando há nessa droga outros componentes farmacologicamente ativos, agravos à saúde dos usuários podem ocorrer, como quadros de intoxicação. Assim, o objetivo deste trabalho foi desenvolver um método de determinação dos principais adulterantes da cocaína (cafeína, levamisol, lidocaína, fenacetina, diltiazem e hidroxizina) em urina humana. A cromatografia líquida de alta eficiência com detector de arranjo de fotodiodos foi utilizada como técnica de análise e a microextração líquido-líquido dispersiva com solidificação da gota orgânica flutuante, como técnica de preparo das amostras. A separação cromatográfica dos analitos em fase reversa foi obtida em uma coluna C18 (250 x 4,6 mm; 5 μm; 80 Å) em modo gradiente e usando acetonitrila-ácido trifluoroacético 0,026% (v/v) a 1 mL.min-1 como fase móvel (25°C e detecção a 235 nm). O tempo de análise foi de 25 min. Para o preparo da amostra, a urina foi alcalinizada com tampão fosfato de sódio 0,5 mol.L-1 (pH 10) e adicionada de cloreto de sódio (20% m/v). Acetonitrila (150 μL) e 1-dodecanol (30 μL) foram utilizados como solvente dispersor e extrator, respectivamente. O método apresentou intervalos lineares de concentração de 312,5 – 3125 ng.mL−1 para cafeína e levamisol e de 187,5 – 1875 ng.mL−1 para lidocaína, fenacetina, diltiazem e hidroxizina, com limite de quantificação de 187,5 ng.mL-1 para lidocaína, fenacetina, diltiazem e hidroxizina e 312,5 ng.mL-1 para cafeína e levamisol. Os valores médios de recuperação variaram de 6,0 a 42,6%. O método mostrou boa precisão e exatidão intra e intercorrida com coeficientes de variação e erros relativos menores que 15%. As amostras apresentaram-se estáveis após ciclos de congelamento-descongelamento e após serem mantidas por 24h em temperatura ambiente. Ainda, o método foi aplicado em cinco amostras de urina de usuários de cocaína e pelo menos um adulterante foi identificado em todas as amostras. Espera-se que este método possa contribuir para a precisão no diagnóstico das intoxicações por adulterantes da cocaína e para o adequado planejamento das medidas terapêuticas.
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Corrêa, Cristiana Leslie. "Validação da urina para análise toxicológica de etanol em \"Programas de Controle e Prevenção do uso de álcool e drogas no local de trabalho\"." Universidade de São Paulo, 1997. http://www.teses.usp.br/teses/disponiveis/9/9137/tde-24112015-182937/.
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Nos dias atuais tem havido uma crescente preocupação com o consumo de álcool, bem como de outras drogas de abuso, no local de trabalho. Com isso, começaram a ser implantados programas que visam o controle e a prevenção do uso de álcool e drogas neste ambiente. De um modo geral, esses programas utilizam a urina dos trabalhadores para a realização de análises toxicológicas. O presente trabalho procurou validar a urina como amostra para determinação do etanol, através da (1) padronização de um método de análise e (2) estudo das correlações entre concentrações sanguínea, urinária e no ar exalado, obtidas de 10 voluntários saudáveis, após administração oral única de bebida alcoólica na dose de 0,68 g de etanol por kg de peso corpóreo, em coletas realizadas durante período de 7 horas. Foi utilizado 1 mL de amostra adicionada de n-propanol (padrão interno), separação por head space e cromatografia em fase gasosa com coluna Poraplot Q 25m x 0,32mm, tendo sido obtidos os seguintes resultados: tempo de retenção do etanol 2.718 ± 0,0024 min., limite de detecção em sangue e urina, 0,07 e 0,01g/l, respectivamente, precisão intra e interensaio igual a 11,4 e 10,0% para o sangue e 5,9 e 6,5% para a urina e recuperação relativa, 55,88 ± 8,03 a 146,35 ± 13,07% para o sangue e 91,60 ± 0,81 a 103,28 ± 1,79% para a urina. A comparação da técnica padronizada com a de imunofluorescência polarizada forneceu um r de 0,9976 para o sangue e 0,9949 para a urina. O estudo de conservação demonstrou que não houve produção in vitro de etanol nas amostras de urina, após permanência à temperatura ambiente por 1 semana. Os resultados de estabilidade mostraram que não houve variação estatisticamente significante entre os resultados de análises feitas em intervalo de 30 dias, quando armazenadas em freezer com adição de fluoreto de sódio a 1 % (p=0,1088 e 0,1548, para sangue e urina, respectivamente). A técnica padronizada foi utilizada nas amostras de sangue e urina colhidas dos voluntários, mostrando-se adequada para a detecção de etanol, em concentrações que variaram de 0,03 a 1,44 g/L. Os valores de correlação urina : sangue nos diferentes tempos de coleta mostraram menor variação que os de sangue: ar exalado, sendo que no período entre 2 e 5 horas após a ingestão do álcool houve os menores desvios de valores de correlação, em todos os indivíduos estudados. Assim sendo, a urina colhida no tempo de 3 horas após o início ou reinício da jornada de trabalho pode ser recomendada para análise de etanol, em programas de controle e prevenção do uso de álcool e drogas no local de trabalho.Abstract not available.
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Barbosa, Analívia Martins. "Período de coleta de urina e de fezes para avaliação da excreção de creatinina, produção microbiana e digestibilidade aparente dos nutrientes em Nelore." Universidade Federal de Viçosa, 2005. http://locus.ufv.br/handle/123456789/5002.
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Previous issue date: 2005-03-02This trial was carried out to evaluate the effect of periods of urine and feces collection on urinary excretion of creatinine, urea and purine derivatives, absorbed purine, microbial nitrogen compounds production (Nmic), apparent digestibility of dry matter (DM), organic matter (OM), crude protein (CP), ether extract (EE), neutral detergent fiber (NDF) and nonfiber carbohydrates (CNF) and total digestible nutrients contents (TDN) of Nelore bovines four categories (heifers, steers, bulls and lactating cows) fed 25 or 50% concentrate, total dry matter basis. The plasma N-urea concentrations (NUP) were evaluated and the Nmic obtained in urine spot samples was compared with that of total collection. The experiment was divided in two experimental periods of 28 days each, when the feces and urine total collection were performed at 22nd and 28th day of each experimental period. Feces were colleted directly from the floor after excretion and the urine was obtained using catheters in females and funnels in males. A 25% concentrate-based diet was fed to the animals in the first period and a 50% concentrate-based diet in the second one, all of them with 12% CP. Sixteen Nelore bovines, under feedlot, housed in individual pens, were assigned to a completely randomized design, in a split plot scheme, where the treatments were represented by the plots (2 x 4 factorial scheme), with two levels of concentrate (25 or 50%) and four Nelore categories, and the split plots were represented by the urine collections. The comparisons for digestibility evaluation were made by regression analyses, that were performed considering 4 days of feces collection of each period. The equations were obtained by comparing the nutrients digestibility referent to one (24h), two (48h) or three days (72h) compared to four days (96h of collection). Comparison with six days of total feces collection, using equations (second experimental period [50% concentrate]) was considered reference (144h total feces collection). No interaction (P>0.05) among concentrate levels, Nelore categories and collection days for the urinary volume, the creatinine excretion and Nmic production was observed. Urinary volume was not affected (P>0.05) by the concentrate levels and collection days, but significant effect (P<0.05) was observed for cows. Creatinine excretion was not affected (P>0.05) by treatments and collection days, considering average of 27.1 mg/kg0.75. Absorbed purines and microbial nitrogen compounds production were also not influenced (P>005) by the treatments and collection days. Nmic production estimated by the urinary spot collection differed (P>0.05) neither from that obtained by total collection total, nor among the concentrate levels and Nelore categories. No significant difference (P>0.05) was observed for any evaluated digestibility and TDN contents during the total feces collection period. The results suggest that the coefficients of variation decresed as the period of collection days increased. Considering the results of creatinine excretion and of microbial protein production, it was concluded that a 24-h period is enough for researchs with Nelore, independently of the category, and that urinary spot sample collection can be used to estimate microbial protein production in all Nelore bovines (heifers, steers, bulls and lactating cows). Total feces collection from 1 to 6 days to evaluate nutrients apparent digestibility are precise, but better results could be obtained by increasing the collection period.
O presente trabalho foi conduzido com o objetivo de avaliar o efeito da duração do período de coletas de urina e de fezes sobre a excreção urinária de creatinina, de uréia e de derivados de purinas, as purinas absorvidas, a produção de compostos nitrogenados microbianos (Nmic), as digestibilidades aparentes da matéria seca (MS), matéria orgânica (MO), proteína bruta (PB), extrato etéreo (EE), fibra em detergente neutro (FDN) e de carboidratos não-fibrosos (CNF) e os teores de nutrientes digestíveis totais (NDT) em Nelores de quatro categorias (novilhas, machos castrados, machos inteiros e vacas em lactação) alimentados com 25 ou 50% de concentrado na base da matéria seca total das dietas. Avaliou-se ainda as concentrações de N-uréia plasmática (NUP) e comparou-se também a produção de Nmic obtida em amostras spot de urina com aquela da coleta total. O experimento foi conduzido em dois períodos experimentais com duração de 28 dias cada, sendo as coletas totais de urina e de fezes efetuadas do 22o ao 28o dias de cada período experimental. As fezes foram retiradas do piso imediatamente após excreção e a urina obtida com sondas em fêmeas e funis nos machos. Utilizou-se dieta com 25% de concentrado no primeiro período e com 50% no segundo experimental. Todas as dietas continham aproximadamente 12% de PB. Utilizaram-se 16 animais da raça Nelore, mantidos em confinamento, alojados em baias individuais, distribuídos em delineamento inteiramente casualizado, em esquema de parcelas subdivididas, tendo nas parcelas os tratamentos em esquema fatorial 2 x 4, sendo dois níveis de concentrado (25 ou 50%) e quatro categorias de Nelores e nas subparcelas os seis dias de coletas de urina. Já para a avaliação das digestibilidades, as comparações foram feitas através de análises de regressão, que foram efetuadas considerando quatro dias de coletas de fezes em cada período, sendo as equações obtidas sempre comparando as digestibilidades dos nutrientes referentes a um dia (24 h), 2 dias (48 h) ou 3 dias (72 h) em relação aos 4 dias (96 h de coleta). Foram feitas também comparações através de equações, utilizando os seis dias de coleta total de fezes, referentes ao segundo período experimental (50% de concentrado), sendo nesse caso utilizado como referência os seis dias de coleta total (144 h de coleta total de fezes). Não houve interação (P>0,05) entre níveis de concentrado, categorias de Nelore e dias de coleta para o volume urinário, a excreção de creatinina e a produção de Nmic. O volume urinário não foi influenciado (P>0,05) pelos níveis de concentrado e dias de coleta, contudo foi significativamente maior (P<0,05) para as vacas. A excreção de creatinina não foi afetada (P>0,05) pelos tratamentos e dias de coletas, observando-se média de 27,1 mg/kg0,75. As purinas absorvidas e a produção de compostos nitrogenados microbianos também não foram influenciadas (P>0,05) pelos tratamentos e dias de coleta. A produção de Nmic estimada através de amostra spot de urina não diferiu (P>0,05) daquela obtida pela coleta total, nem entre os níveis de concentrados e categorias de Nelore. Não houve diferença significativa (P>0,05) para nenhuma das digestibilidades avaliadas e também para os teores de NDT entre os dias de coleta total de fezes, contudo, observou-se que os coeficientes de variação diminuíram à medida que se aumentou o número de dias de coleta. Concluiu-se considerando as respostas obtidas para excreção de creatinina e a produção de proteína microbiana, que um período de coletas de urina de 24 horas é suficiente para trabalhos com Nelores, independente de serem novilhas, machos castrados ou inteiros e vacas em lactação e que a coleta de amostra spot de urina também pode ser utilizada para estimar a produção de proteína microbiana em novilhas, machos inteiros ou castrados e vacas lactantes da raça Nelore. Concluiu-se também que para avaliar a digestibilidade aparente dos nutrientes, coletas totais de fezes feitas durante um a seis dias são exatas. Contudo, a precisão é melhorada com o aumento dos dias de coleta.
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Hassen, Imed Eddine. "Analyse de traces et ultratraces (organiques et inorganiques) dans les matrices biologiques." Lyon 1, 2004. http://www.theses.fr/2004LYO10126.
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La majorité des analyses biologiques s'appuient sur les deux matrices urine et sang, qui présentent un inconvénient majeur au niveau de la fenêtre de détection très courte. Cette limitation a contribué ces dernières années à l'essor d'une matrice alternative : les cheveux, support beaucoup plus stable. L'objectif de ce travail est de comparer les résultats d'analyse de xénobiotiques dans l'urine et les cheveux. L'application principale porte sur l'étude le phénomène du tabagisme, examiné sous l'angle du traitement de la dépendance. Un suivi d'un programme de sevrage a été effectué par CG-SM et CLHP-SM en utilisant les deux marqueurs nicotine et cotinine. L'étude d'une corrélation éventuelle entre le tabagisme et la concentration de plomb et de cadmium dans le corps a été réalisée par ICP-MS. Une deuxième partie est enfin consacrée au suivi par CLHP-SM-SM de l'évolution de la concentration d'un principe actif, la tamsulosine, dans les urines de personnes sous traitement
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Diaz, Sílvia de Oliveira. "Pregnancy and newborns disorders followed by urine metabolomics." Doctoral thesis, Universidade de Aveiro, 2014. http://hdl.handle.net/10773/13110.
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Doutoramento em QuímicaChapter 1 introduces the scope of the work by identifying the clinically relevant prenatal disorders and presently available diagnostic methods. The methodology followed in this work is presented, along with a brief account of the principles of the analytical and statistical tools employed. A thorough description of the state of the art of metabolomics in prenatal research concludes the chapter, highlighting the merit of this novel strategy to identify robust disease biomarkers. The scarce use of maternal and newborn urine in previous reports enlightens the relevance of this work. Chapter 2 presents a description of all the experimental details involved in the work performed, comprising sampling, sample collection and preparation issues, data acquisition protocols and data analysis procedures. The proton Nuclear Magnetic Resonance (NMR) characterization of maternal urine composition in healthy pregnancies is presented in Chapter 3. The urinary metabolic profile characteristic of each pregnancy trimester was defined and a 21-metabolite signature found descriptive of the metabolic adaptations occurring throughout pregnancy. 8 metabolites were found, for the first time to our knowledge, to vary in connection to pregnancy, while known metabolic effects were confirmed. This chapter includes a study of the effects of non-fasting (used in this work) as a possible confounder. Chapter 4 describes the metabolomic study of 2nd trimester maternal urine for the diagnosis of fetal disorders and prediction of later-developing complications. This was achieved by applying a novel variable selection method developed in the context of this work. It was found that fetal malformations (FM) (and, specifically those of the central nervous system, CNS) and chromosomal disorders (CD) (and, specifically, trisomy 21, T21) are accompanied by changes in energy, amino acids, lipids and nucleotides metabolic pathways, with CD causing a further deregulation in sugars metabolism, urea cycle and/or creatinine biosynthesis. Multivariate analysis models´ validation revealed classification rates (CR) of 84% for FM (87%, CNS) and 85% for CD (94%, T21). For later-diagnosed preterm delivery (PTD), preeclampsia (PE) and intrauterine growth restriction (IUGR), it is found that urinary NMR profiles have early predictive value, with CRs ranging from 84% for PTD (11-20 gestational weeks, g.w., prior to diagnosis), 94% for PE (18-24 g.w. pre-diagnosis) and 94% for IUGR (2-22 g.w. pre-diagnosis). This chapter includes results obtained for an ultraperformance liquid chromatography-mass spectrometry (UPLC-MS) study of pre-PTD samples and correlation with NMR data. One possible marker was detected, although its identification was not possible. Chapter 5 relates to the NMR metabolomic study of gestational diabetes mellitus (GDM), establishing a potentially predictive urinary metabolic profile for GDM, 2-21 g.w. prior to diagnosis (CR 83%). Furthermore, the NMR spectrum was shown to carry information on individual phenotypes, able to predict future insulin treatment requirement (CR 94%). Chapter 6 describes results that demonstrate the impact of delivery mode (CR 88%) and gender (CR 76%) on newborn urinary profile. It was also found that newborn prematurity, respiratory depression, large for gestational age growth and malformations induce relevant metabolic perturbations (CR 82-92%), as well as maternal conditions, namely GDM (CR 82%) and maternal psychiatric disorders (CR 91%). Finally, the main conclusions of this thesis are presented in Chapter 7, highlighting the value of maternal or newborn urine metabolomics for pregnancy monitoring and disease prediction, towards the development of new early and non-invasive diagnostic methods.
O Capítulo 1 descreve o enquadramento deste trabalho identificando as doenças pré-natais relevantes e os métodos de diagnóstico actualmente disponíveis. É depois apresentada a metodologia seguida, assim como uma breve introdução dos princípios dos métodos analíticos e estatísticos aplicados. O capítulo é concluído com uma descrição do estado da arte na área de metabolómica em investigação pré-natal, identificando o mérito desta inovadora estratégia para a identificação de marcadores robustos de doenças pré-natais. A relevância deste trabalho torna-se clara através do escasso uso de urina materna e do recém-nascido em trabalhos anteriores. O Capítulo 2 descreve os procedimentos experimentais utilizados neste trabalho, incluindo condições de amostragem, recolha e preparação das amostras, protocolos de aquisição e de tratamento dos dados. A caracterização da composição da urina materna, através de espectroscopia de Ressonância Magnética Nuclear (RMN) de protão é apresentada no Capítulo 3. Define-se o perfil metabólico urinário característico para cada trimestre de gravidez, tendo sido encontrado um conjunto de 21 metabolitos descritivo das alterações metabólicas ocorridas ao longo da gravidez. 8 metabolitos foram encontrados a variar com a gravidez, pela primeira vez, tendo sido confirmadas variações metabólicas conhecidas. É ainda estudado o efeito do não-jejum (usado neste trabalho) como possível factor de confusão. O Capítulo 4 apresenta o estudo metabolómico de urina materna do 2º trimestre para o diagnóstico de doenças fetais e previsão de complicações mais tarde desenvolvidas. Este estudo compreende a aplicação de um método de selecção de variáveis desenvolvido no âmbito desta tese. Observou-se que as malformações fetais (e, especificamente, do sistema nervoso central, SNC) e as cromossomopatias (e, especificamente, a trissomia 21, T21) são acompanhadas por alterações nos metabolismos energético, dos aminoácidos, lípidos e nucleótidos, enquanto que as cromossomopatias mostraram ser acompanhadas por uma desregulação adicional dos metabolismos dos açúcares, ciclo da ureia e/ou biossíntese da creatinina. A validação dos modelos multivariados revelou taxas de classificação (CR) de 84% para malformações (87%, SNC) e 85% para CD (94%, T21). Para o parto pré-termo, pré-eclampsia (PE) e restrição de crescimento intrauterino (RCIU) observaram-se perfis que podem ajudar à previsão precoce, com CR 84% para pretermo (11-20 semanas de gestação, g.w. pré-diagnóstico), 94% para PE (18-24 g.w. pré-diagnóstico) e 94% para RCIU (2-22 g.w. pré-diagnóstico). Este capítulo inclui resultados obtidos por cromatografia líquida de ultra eficiência acoplada a espectrometria de massa (UPLC-MS) para pré-pretermo e correlação com os dados de RMN. Um possível composto marcador foi detectado mas a sua identificação não foi possível. O Capítulo 5 descreve o estudo metabolómico por RMN da diabetes mellitus gestacional (DMG), estabelecendo-se um perfil metabólico potencialmente preditivo da doença (CR 83%, 2-21 g.w. pré-diagnóstico). Verificou-se ainda que o espectro de RMN contém informação sobre o fenótipo individual, capaz de prever a necessidade futura de tratamento com insulina (CR 94%). No Capítulo 6 demonstra-se o impacto do tipo de parto (CR 88%) e género do bebé (CR 76%) no perfil da urina do recém-nascido. Verificou-se ainda que a prematuridade, depressão respiratória, crescimento grande para a idade gestacional e malformações induzem perturbações metabólicas relevantes (CR 82-92%), assim como algumas doenças maternas como a DMG (CR 82%) e doenças psiquiátricas (91% CR). Finalmente, no Capítulo 7 apresentam-se as principais conclusões deste trabalho, enfatizando o potencial da metabolómica de urina materna e do bebé para o acompanhamento da gravidez e previsão de doenças, visando o desenvolvimento de novos métodos de diagnóstico precoce e não-invasivo.
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Colombeli, Adriana Scotti da Silva. "Avaliação do potencial de interferência analítica de fármacos na análise química do exame de urina." Florianópolis, SC, 2006. http://repositorio.ufsc.br/xmlui/handle/123456789/88952.
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Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências da Saúde. Programa de Pós-Graduação em Farmácia.Made available in DSpace on 2012-10-22T15:16:05Z (GMT). No. of bitstreams: 1 233101.pdf: 855282 bytes, checksum: 0cc76a508ef9aa577987b2e41271fece (MD5)
O exame de urina proporciona informações sobre patologias renais e do trato urinário, bem como sobre algumas moléstias extra-renais. Pela sua simplicidade, baixo custo e facilidade na obtenção da amostra para análise, é considerado exame de rotina. Fármacos podem alterar exames laboratoriais por meio de mecanismos farmacológicos, físicos, químicos e metabólicos. O objetivo deste trabalho foi avaliar o potencial de interferência analítica de 24 fármacos no exame químico da urina. A análise de todos os parâmetros da fita reativa (Multistix® 10 SG Bayer) foi realizada comparando-se os resultados de urinas controles (sem o fármaco) e urinas às quais foram adicionados os fármacos. A metodologia baseou-se em um protocolo adotado internacionalmente para estudos da interferência analítica. Foi realizada a avaliação do potencial de interferência em concentrações consideradas supraterapêuticas, terapêuticas e subterapêuticas, estimadas com base na dose e em dados farmacocinéticos. Não foram detectadas interferências nas concentrações testadas para aciclovir, amoxicilina, atenolol, diclofenaco potássico, lisinopril, tartarato de metoprolol, metronidazol, ofloxacino, piracetam e zidovudina. No parâmetro cetona, foi verificada interferência falso-positiva para o captopril, em concentrações supraterapêutica e terapêutica. O carbonato de lítio apresentou interferência falso-negativa para o parâmetro densidade, no qual captopril, cefalexina, cimetidina, difosfato de cloroquina, bem como os cloridratos de amantadina, ciprofloxacino, efedrina, metformina, metoclopramida, ranitidina e tetraciclina, além dos sulfatos de atropina e quinina, provocaram interferência falso-positiva nas concentrações testadas. No parâmetro glicose houve interferências falso-negativas dos fármacos captopril, cloridrato de ciprofloxacino e difosfato de cloroquina na concentração supraterapêutica. Os fármacos captopril, cimetidina e cloridrato de tetraciclina promoveram interferência falso-negativa no parâmetro hemoglobina, em concentração supraterapêutica; o difosfato de cloroquina interferiu também em concentração terapêutica e o sulfato de quinina nas três concentrações testadas. O pH sofreu interferências positivas na concentração terapêutica do carbonato de lítio e na supraterapêutica da cimetidina, enquanto os cloridratos de ciprofloxacino e tetraciclina, bem como o sulfato de quinina, acidificaram a urina. No parâmetro proteína, o cloridrato de ciprofloxacino e o sulfato de quinina apresentaram interferência falso-positiva só na concentração supraterapêutica, enquanto o difosfato de cloroquina interferiu também na concentração terapêutica. Não foram observadas interferências positivas nos parâmetros bilirrubina, urobilinogênio e esterase de leucócitos. No parâmetro nitrito, nenhuma interferência foi observada. Algumas das interferências analíticas encontradas estão sendo relatadas pela primeira vez, demonstrando a necessidade de mais estudos nesta área. A qualidade das informações do fabricante do produto utilizado foi considerada inadequada, recomendando-se maior cuidado na sua elaboração. The urinalysis provides information about renal and urinary diseases, as well as about some extra-renal diseases. Due to its simplicity and low cost, it is considered a routine examination. Drugs can modify laboratorial results through pharmacological, physical and chemical mechanisms. The objective of this study was to evaluate the potential of analytical interference of 24 drugs in the chemical examination of urine. The analysis of reagent strips# parameters with Multistix® 10 SG Bayer was done comparing the results of control urines (drug free) and drugs supplemented urines. The methodology was based on an internationally adopted protocol for studies of analytical interferences. The evaluation of interference potencial was done in supratherapeutical, therapeutical and subtherapeutical concentrations, which were estimated based on dose and pharmacokinetical data. Interferences were not detected in the tested concentrations to acyclovir, amoxicillin, atenolol, potassic diclofenac, lisinopril, metoprolol tartarate, metronidazole, ofloxacin, piracetam and zidovudine. The parameter ketone showed false-positive interference for captopril in supratherapeutical and therapeutical concentrations. Urine supplemented with lithium carbonate presented false-negative interference in the parameter density. In this parameter captopril, cefalexin, cimetidine, chloroquine difosfate, as well as the hydrochloride salts of amantadine, ciprofloxacin, ephedrin, metformin, metoclopramide, ranitidin and tetracycline, beyond atropine sulfate and quinine sulfate provoked false-positive interference in the tested concentrations. There were false-negative interferences for glucose in urines containing captopril, ciprofloxacin hydrochloride and chloroquine diphosphate in supratherapeutical concentrations. Captopril, cimetidine and tetracycline hydrochlorides promoted false-negative interference in hemoglobin, in supratherapeutical concentrations; chloroquine diphosphate also interfered in therapeutical concentration and quinine sulphate in all the three tested concentrations. Urines spiked with lithium carbonate (therapeutical) and cimetidine (supratherapeutical) presented positive interferences in pH; ciprofloxacin and tetracycline hydrochlorides, as well quinine sulfate diminished urinary pH. In the parameter protein, ciprofloxacin hydrochloride and quinine sulfate presented false-positive interference only in the supratherapeutical concentration, while chloroquine difosfate also interfered in therapeutical concentration. Positive interferences were not observed in bilirubin, urobilinogen and leukocytes esterase. No interference was observed to nitrite. Some of these analytical interferences are being described for the first time, what showed up the necessity of more studies in this field.
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Pujos, Estelle. "Analyse de traces dans les matrices urinaires : application au contrôle antidopage." Lyon 1, 2004. http://www.theses.fr/2004LYO10120.
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Les progrès pharmaceutiques et les enjeux croissants des résultats sportifs ont permis le développement du dopage. Devenant un problème de santé publique, les organisations établissent des listes d'interdictions et développent des contrôles nécessitant des méhtodes d'analyses fiables. La première partie de ce travail concerne la mise au point de la détection de neuf métabolites de stéroi͏̈des dans l'urine, en vue d'analyses par spectrométrie de masse isotopique. Cette étude nous a permis de développer une analyse globale concernant le sulfate de DHEA, les corticostéroi͏̈des et les androgènes dans les urines. Nous avons montré que la prise de corticostéroi͏̈des entraînait une modification de l'excrétion des stéroi͏̈des. Dans une deuxième partie, nous avons évalué les possibilités de trois techniques alternatives (ELISA, GC/MS, LC/MS) en vue du contrôle des corticostéroi͏̈des et bêtabloquants. Différents protocoles ont été proposés et les limites de quantification ont été déterminées
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Fong, Ka-wah Martin, and 方家華. "Adaptation of a simplified method for urinary iodine for studying the iodine status of local Chinese." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2004. http://hub.hku.hk/bib/B31971726.
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Thomas, Marie Martineau Guy-Pierre. "Prévalence de infections urinaires chez la truie gestante (ITU) selon le stade de gestation et la parité dans deux contextes d'abreuvement différents." [S.l.] : [s.n.], 2007. http://oatao.univ-toulouse.fr/1754/1/debouch_1753.pdf.
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Silva, Adriana Scotti da. "Avaliação da interferência analítica de fármacos na determinação de proteínas e cetonas no exame químico de urina." Florianópolis, 2012. http://repositorio.ufsc.br/xmlui/handle/123456789/100495.
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Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências da Saúde. Programa de Pós-Graduação em Farmácia.Made available in DSpace on 2013-06-25T19:43:19Z (GMT). No. of bitstreams: 1 310058.pdf: 1105263 bytes, checksum: af4972ecbcb412b86bd83dc70f8dc500 (MD5)
Interferências de fármacos em análises laboratoriais podem levar a resultados incorretos, diagnósticos equivocados e procedimentos desnecessários. O exame de urina permite a detecção de processos patológicos intrínsecos e extrínsecos ao sistema urinário, e é importante, sobretudo para diabéticos e pacientes com problemas renais. A partir de interferências em proteínas e cetonas urinárias detectadas em estudo anterior, o objetivo deste trabalho foi aprofundar os estudos de interferência analítica nestes parâmetros utilizando as tiras reagentes Multistix® 10 SG e Combur® 10 Test M, por meio de ensaios in vitro e in vivo. No estudo in vitro tornou-se evidente o potencial interferente de fármacos quinolínicos (derivados de quinina, cloroquina e hidroxicloroquina) e quinolônicos (cloridrato de ciprofloxacino, ofloxacino e levofloxacino) para proteína na tira Multistix ® 10SG e no teste confirmatório com vermelho de pirogalol-molibdato. Fármacos quinolínicos apresentaram interferência em concentrações subterapêuticas. No estudo in vivo, pacientes usuários de captopril apresentaram interferências estatisticamente significativas na detecção de cetonúria, em doses ? 50 mg/dia, mas as interferências não foram necessariamente proporcionais à dose. Para um mesmo paciente, amostras com interferência falso-positiva na tira Multistix® 10SG apresentaram maiores concentrações de captopril na urina (determinadas por método de cromatografia líquida de alta eficiência, adaptado e validado) que amostras sem interferência. Pacientes em uso de ciprofloxacino apresentaram interferências clinicamente significantes na quantificação de proteína urinária com vermelho de pirogagol-molibdato. No estudo transversal, a comparação da frequência de uso de fármacos entre amostras verdadeiro-positivas e falso-positivas para cetona e proteína urinárias não permitiu identificar inequivocamente outros fármacos com potencial de interferência, porém observaram-se associações estatisticamente significativas entre resultados falso-positivos para proteína e uso de cloridrato de metoclopramida, diazepam, dipirona + butilbrometo de escopolamina, heparina ou sulbactam sódico pelo teste de qui-quadrado.
Urinalysis allows detection of pathological processes of urinary system (intrinsic and extrinsic), being especially important for diabetics and patients with renal diseases. Drugs interferences in clinical analysis can lead to incorrect results, false diagnostics and unnecessary procedures. The objective of this study was to extend a previous in vitro interference study on urinary protein and ketone interferences, to include analytical reagent strips Test 10 M,Combur R besides Multistix R 10 SG and to perform also in vivo studies. Potential interferences for urinary protein were observed in vitro with quinolinic drugs (quinine sulfate, chloroquine diphosphate, hydroxychloroquine sulfate) and quinolone antibiotics (ciprofloxacin hydrochloride, ofloxacin and levofloxacin) in tests with Multistix R 10SG reagent strip and the pyrogallol red-molybdate assay; quinolinic drugs interfered in subtherapeutical concentrations. By the in vivo study, statistically significant interferences were observed in patients using captopril (. 50 mg/day). Interference magnitudes were not always proportional to the dosage, but considering intrapersonal variations, urinary concentrations of captopril were higher in samples that showed false-positive interference by MultistixR 10SG reagent strip in comparison to samples that did not show these interferences. Samples from patients taking ciprofloxacin presented clinically significant interference for proteinuria in the quantitative analysis performed with pyrogallol red-molybdate. In the transversal study, comparison of reports for used drugs among patients with true-positive and false-positive tests for urinary protein and ketones, did not allow the identification of other potentially interfering drugs, but statistically significant associations were observed between therapy with metoclopramide hydrochloride, diazepam, dipyrone + scopolamine butylbromide, heparin or sodium sulbactam and false-positive results for proteinuria with chi-square test.
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Shreeram, Devesh Dadhich. "Electrochemical Analysis of Genetically Engineered Bacterial Strains in a Urine-Based Microbial Fuel Cell." University of Cincinnati / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1458814734.
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SANTORELLI, LUCIA. "Proteomic analysis of urine-based liquid biopsy to provide new insights into renal diseases." Doctoral thesis, Università degli Studi di Milano-Bicocca, 2020. http://hdl.handle.net/10281/263397.
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L'area di indagine delle paotlogie renali è un campo ampio e complesso e molte delle cause che sono alla loro base non sono completamente comprensibili o curabili. A tal motivo, oltre ad una migliore comprensione della loro eziopatogenesi, è necessario individuare in maniera precisa i fattori di rischio, al fine di migliorare la prognosi e personalizzare il trattamento terapico.
Lo sviluppo e l’implementazione di nuovi approcci, come l'analisi proteomica basata sulle urine, come biopsia liquida è attualmente una delle scelte più giuste da compiere. Infatti, l'uso dell'analisi proteomica per la scoperta di proteine clinicamente rilevanti, note come proteomica clinica, individuando ad esempio marcatori diagnostici e prognostici, può effettivamente favorire la traslazione delle scoperte di base in applicazioni cliniche a beneficio del paziente.
Un campione biologico facilmente accessibile, come le urine, in tal senso, si rivela essere una preziosa fonte di biomarcatori per patologie renali. Le urine, infatti, possono essere raccolte in grandi quantità e in modo non invasivo; inoltre, sono meno complesse di altri fluidi corporei e per questo più semplici da studiare.
Purtroppo, in diversi casi, le preziose informazioni presenti nelle urine sono spesso tecnicamente difficili da estrarre perché le proteine legate alla malattia sono spesso presenti in concentrazioni molto basse e per di più nascoste da proteine urinarie presenti in grande abbondanza, come l'albumina o l'uromodulina.
In questo contesto, lo studio proteomico degli esosomi urinari (EU) rappresenta una valida alternativa, per rivelare e scoprire questo interessante paesaggio molecolare nascosto. EU sono vescicole di dimensioni nanometriche (>150 nm), che possono provenire da cellule endoteliali, podociti o cellule epiteliali tubolari. La loro composizione molecolare dipende dal tipo e dalla condizione fisiologica della cellula da cui si originano. Inoltre, l'impiego degli EU permette di ridurre la complessità del proteoma urinario: essi, infatti, contengono solo il 3% delle proteine urinarie totali (>3000 specie). Per tutte queste ragioni, alla stregua delle urine, possiamo considerare anche gli EU come una biopsia liquida, modalità non invasiva, in grado di fornire informazioni diagnostiche e prognostiche relative alle più disparate patologie renali.
In questo studio, abbiamo applicato gli approcci di spettrometria di massa (MS) e proteomica applicata allo studio del proteoma e del glicoproteoma di campioni urinari provenienti da pazienti affetti da carcinoma renale a cellule chiare (ccRCC), il più frequente e aggressivo istotipo di carcinoma renale. In aggiunta, abbiamo anche studiato il contenuto proteico di EU di pazienti affetti da sindrome nefrosica idiopatica (INS), la più diffusa malattia glomerulare infantile. Grazie a questo approccio complementare, abbiamo raggiunto il duplice obiettivo di individuare una specifica signature proteica e glicoproteica di progressione tumorale (nel caso di cc-RCC) e di chiarire l'eziopatogenesi della malattia e il meccanismo molecolare che sta alla base della diversa risposta al trattamento farmacologico e dell'insorgenza della farmacoresistenza ai corticosteroidi (nel caso di INS).
Combinando i risultati ottenuti, speriamo di sostenere e rafforzare ulteriormente l'impiego della biopsia liquida nella ricerca clinica, sia da un punto di vista tecnico che da un punto di vista applicativo. Inoltre, sfruttando le possibilità legate all'impiego della MS, speriamo di arrivare alla conversione dei nostri risultati in strumenti diagnostici o prognostici che possano migliorare la gestione dei pazienti affetti da ccRCC e INS.The area of kidney diseases is a wide and complex field, and many conditions leading to them are not fully understood or curable. Along with a better understanding of these pathologies, there is a need for improved risk detection, for determination of prognosis and for improved and personalized treatment. Therefore, it is important to develop and implement new approaches, such as proteomic analysis of urine- based liquid biopsy. Indeed, the use of proteome analysis for the discovery of clinically relevant proteins, known as clinical proteomics, for example for the identification of earlier and prognostic markers may actually foster the translation of basic discoveries into clinical applications for the benefit of the patient. Easily accessible biological sample, such as urine is valuable sources of biomarkers for pathologies related to kidney. In fact, urine can be collected in large quantities and in non-invasive way; additionally, it is less complex than other bodily fluids. Unfortunately, in diverse cases, this information is often technically difficult to be mined because the disease-related proteins are often present in very low concentrations, are frequently labile, and are hidden by high-abundance proteins such as albumin or uromodulin. In this context, the proteomic study of the urinary extracellular vesicles (UEv) represents a valid alternative to reveal and discovery these hidden molecular landscape. UEv are nanometer-sized vesicles (>150 nm), that can originate from endothelial cells, podocytes or tubular epithelial cells. Their molecular composition depends upon the type, and even tatus, of the producer cell. Therefore, the use of UEv allows to reduce the complexity of the urine proteome, because they contain only 3% of total urine proteins (>3000 species). For all these reasons, we can consider also them (as the urine), as a liquid biopsy, non-invasive modality that can provide diagnostic and prognostic information about kidney disease. In this study, we applied the proteomics MS-based approaches to investigate the proteome and glycoproteome in urine samples of patients affected by clear cell Renal Cell Carcinoma (ccRCC), the most frequent and aggressive type of renal carcinoma. Additionally, we also studied the protein content of UEv of Idiopathic Nephrotic Syndrome (INS), the major childhood glomerular disease. By our complementary approaches, we gained the double aim: firstly, to pinpoint a characteristic specific disease protein and glycoprotein signature of tumour progression (in case of cc-RCC); secondly, to clarify the disease etiopathogenesis and the molecular mechanism underling the different response to drug treatment and the onset of pharmacoresistance to corticosteroids (in case of INS). By combining the obtained results, we hope to enforce further the use of liquid biopsy in clinical research, from both a technical and application standpoint. Furthermore, by taking advantage of the possibilities related to the employment of the MS technologies, we also expect that this can lead to the direct translation of our findings into diagnostic or prognostic tools that can improve the clinical management of ccRCC and INS patients.
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BELLINTANI, SANDRA A. "Estudo de uma metodologia dosimetrica para plutonio por meio de analise radiotoxicologica em urina." reponame:Repositório Institucional do IPEN, 1988. http://repositorio.ipen.br:8080/xmlui/handle/123456789/9893.
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Dissertacao (Mestrado)
IPEN/D
Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP
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Rocha, Diogo Librandi da. "Desenvolvimento de procedimentos analíticos em fluxo com multicomutação e foto-oxidação em linha para determinação espectrofotométrica de espécies de interesse ambiental, alimentício e clínico." Universidade de São Paulo, 2013. http://www.teses.usp.br/teses/disponiveis/46/46136/tde-06082013-075926/.
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A mecanização do preparo de amostras minimiza erros sistemáticos, a geração de efluentes e o tempo de análise. Sistemas em fluxo com microbombas solenoide (MPFS) atendem aos requisitos da mecanização de maneira robusta e versátil. Sendo assim, foram desenvolvidos procedimentos analíticos baseados em MPFS e foto-oxidação em linha visando ao fracionamento de fósforo em extratos de alimentos e águas naturais e à determinação de cloreto em águas naturais e urina. Fósforo é um nutriente importante cuja biodisponibilidade depende de sua forma química, tornando importante o fracionamento. O monitoramento de cloreto também é importante porque altas concentrações causam efeitos adversos ao meio ambiente e à saúde. Um MPFS foi proposto para o fracionamento de fósforo solúvel em extratos de alimentos, empregando foto-oxidação em linha do fósforo orgânico a ortofosfato, que foi quantificado pelo método do azul de molibdênio. Foi obtida resposta linear entre 5,0 e 40 mg L-1 para fósforo inorgânico (PI) e orgânico (PO), com limites de detecção de 0,5 e 1,2 mg L-1, respectivamente. Coeficientes de variação foram estimados em 1,2 e 3,6% para PI e PO, respectivamente, com frequência de amostragem de 80 determinações por hora. Por determinação, foram consumidos 380 µg de (NH4)6Mo7O24, 620 µg de ácido ascórbico e 790 µg de K2S2O8, gerando 2,5 mL de efluentes. Os resultados do fracionamento em extratos de alimentos concordaram com os obtidos pelo procedimento de referência (95% de confiança), baseado na digestão nitro-perclórica para a determinação de PO. Um procedimento com MPFS e espectrofotometria de longo caminho óptico foi desenvolvido para o fracionamento de fósforo (inorgânico e orgânico dissolvidos) em águas naturais. A quantificação foi baseada no método do azul de molibdênio, após fotoconversão das espécies a ortofosfato. Resposta linear foi observada entre 10 e 75 µg L-1 P, com limite de detecção de 2,0 µg L-1. Foram obtidos coeficiente de variação de 1,8% e frequência de amostragem de 40 determinações por hora. Por determinação, foram consumidos 160 µg de (NH4)6Mo7O24, 10 µg de SnCl2, 640 de µg K2S2O8 e 10 mg de NaOH, gerando 4,0 mL de efluentes. Os coeficientes angulares das curvas obtidas com 4 tipos diferentes de espécies de PO indicaram conversão quantitativa e os resultados obtidos para amostras de águas de rios foram concordantes com os obtidos pelo procedimento descrito pela AOAC (95% de confiança). Um procedimento limpo foi também desenvolvido para a determinação de cloreto em águas naturais e urina, evitando o uso de reagentes tóxicos. O analito foi foto-oxidado a cloro, que foi determinado por espectrofotometria através da descoloração do alaranjado de metila. Resposta linear foi observada entre 2,0 e 20 mg L-1, com limite de detecção de 0,7 mg L-1. O coeficiente de variação foi estimado em 1,6%, com frequência de amostragem de 75 determinações por hora e consumo de 7,5 µg de corante por determinação. Espécies concomitantes usualmente presentes nas amostras não interferiram mesmo em excesso em relação às concentrações normalmente encontradas. Os resultados das análises das amostras concordaram com os obtidos pelo procedimento de referência (95% de confiança). Os procedimentos propostos são alternativas limpas e rápidas para o fracionamento de fósforo e determinação de cloreto.Mechanization of sample preparation minimizes systematic errors, waste generation and analysis time. Flow-based systems with solenoid micropumps (MPFS) attain the requirements for mechanization in a versatile and robust way. Therefore, analytical procedures based on MPFS and on-line photo-oxidation were developed aiming phosphorus fractionation in foodstuff and river waters and chloride determination in natural waters and urine. Phosphorus is an important nutrient for plants and animals and its bioavailability depends on its chemical form, making fractionation studies important. Chloride monitoring is relevant because the concentration unbalance leads to environmental and health issues. A MPFS was proposed for the fractionation of water soluble phosphorus in foodstuff, incorporating on-line photo-oxidation of organic phosphorus to phosphate, which was quantified by the spectrophotometric molybdenum blue method. Linear response was observed from 5 to 40 mg L-1 for both inorganic (PI) and organic (PO) phosphorus, with detection limits of 0.5 and 1.2 mg L-1, respectively. Coefficients of variation (n = 20) were estimated as 1.2 and 3.6% for PI and PO, respectively, with a sampling rate of 80 determinations per hour. Per determination, 380 µg of (NH4)6Mo7O24, 620 µg of ascorbic acid and 790 µg of K2S2O8 were consumed, generating 2.5 mL of waste. The results for food extracts agreed with those obtained by the reference procedure (95% confidence level) based on nitro-percloric digestion for PO determination. A MPFS procedure with long pathlength spectrophotometry was developed for phosphorus fractionation (dissolved organic and inorganic) in natural waters. Quantification was also based on the formation of molybdenum blue, after on-line photo-convertion of the organic species to orthophosphate. The analytical response was linear within 10 and 75 µg L-1 with a detection limit of 2.0 µg L-1. Coefficient of variation of 1.8% and sampling rate of 40 determinations per hour were achieved. Per determination, 160 µg of (NH4)6Mo7O24, 10 µg of SnCl2, 640 µg of K2S2O8 and 10 mg of NaOH were consumed, generating 4.0 mL of waste. Slopes of analytical curves obtained for four different PO species agreed with those obtained for orthophosphate, indicating quantitative conversion and the results for five freshwater samples agreed with those obtained by the AOAC reference procedure at 95% confidence level. A green procedure for chloride determination in urine and natural waters was also developed, avoiding hazardous chemicals. The analyte was on-line photo-converted to chlorine which was spectrophotometrically detected by methyl orange discoloration. The analytical response was linear from 2.0 to 20 mg L-1 Cl with a detection limit of 0.7 mg L-1. The coefficient of variation was 1.6% with a sampling rate of 75 determinations per hour, consuming 7.5 µg of the dye per determination. Usual concomitant species did not cause significant interference even in excess in relation to their highest concentration expected in the samples. The results for urine and water samples agreed with those obtained by the reference procedures at the 95% confidence level. The proposed procedures are environmentally friendly and fast alternatives for phosphorus fractionation and chloride determination
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Scalabre, Aurélien. "Étude clinique des modifications du profil métabolique urinaire secondaires à une anomalie congénitale de l’écoulement des urines par Résonance Magnétique Nucléaire et analyse métabolomique." Thesis, Lyon, 2017. http://www.theses.fr/2017LYSE1272/document.
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Les dilatations des voies urinaires de diagnostic anténatal (DVU) peuvent être transitoires, ou correspondre à une anomalie significative de l'écoulement des urines pouvant entrainer une dégradation de la fonction rénale. L'identification de biomarqueurs urinaires pourrait contribuer à différencier précocement les uropathies des dilatations transitoires. La métabolomique permet l'identification de toutes les molécules de bas poids moléculaire présentes dans un échantillon biologique et la mise en évidence d'une signature métabolique associée à un événement physiopathologique. L'objectif principal de cette thèse est l'identification de marqueurs urinaires pronostiques chez les nouveaux nés présentant une DVU de diagnostic anténatal, par spectroscopie RMN et analyse métabolomique.Soixante-dix nouveau-nés ayant une DVU unilatérale et 90 témoins ont été inclus dans cette étude prospective. Tout d'abord, nous comparons la précision de différentes classifications échographiques pour l'évaluation du risque d'intervention chirurgicale.Ensuite, nous montrons l'absence de différence significative entre les profils métaboliques urinaires des nouveau-nés ayant une DVU et ceux des témoins. Dans une troisième partie, nous démontrons l'influence significative de l'âge, du poids et de la taille sur le profil métabolique urinaire des nouveau-nés. Enfin, nous montrons l'évolution du profil métabolique urinaire avec la croissance chez les nourrissons ayant une DVU.Ce travail permet une meilleure compréhension de la physiopathologie des DVU et de la maturation métabolique des nouveau-nés. Il contribue à identifier des biais potentiels dans les études métaboliques en néonatologieThe prenatal finding of unilateral Urinary Tract Dilatation (UTD) can be transient or represent a significant urinary flow impairment that would lead to progressive deterioration of renal function. Identifying urinary biomarkers could help to differentiate uropathy requiring surgical management from transient dilatation at an early stage.Metabolic phenotyping studies provide untargeted quantification of all detectable low molecular-weight molecules by profiling without any a priori the metabolic signatures of biological samples in connection to physiopathological events.The main objective of this study is to identify diagnosis and prognosis urinary markers for uropathy in newborns with prenatally diagnosed unilateral UTD using 1H-NMR spectroscopy combined with multivariate statistical analyses.A total of 70 newborns with unilateral UTD and 90 controls were included in this prospective study. First, the usefulness of different ultrasound grading systems in predicting the need for surgical intervention is evaluated. Then, we report the absence of significant difference between the urinary metabolic profiles of newborns with UTD and controls. In thethird part, the influence of age, weight and height on the urinary metabolic profiles of healthy newborns is highlighted for the first time, and key-metabolites responsible for this evolution are identified. Finally, we demonstrate the influence of age on the urinary metabolic profiles of children with UTD. This work allows a deeper understanding of the metabolic maturation of healthy newborns. It contributes to identify potential confounding factors for metabolomics investigations in neonatology. It represents a step toward a better comprehension of thephysiopathology of UTD
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Li, Xin. "Comparative evaluation of the extraction and analysis of urinary phospholipids and lysophospholipids using MALDI-TOF/MS." Doctoral thesis, Kyoto University, 2021. http://hdl.handle.net/2433/265182.
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京都大学新制・課程博士
博士(医学)
甲第23410号
医博第4755号
新制||医||1052(附属図書館)
京都大学大学院医学研究科医学専攻
(主査)教授 村川 泰裕, 教授 長尾 美紀, 教授 柳田 素子
学位規則第4条第1項該当
Doctor of Medical Science
Kyoto University
DFAM
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Van, Buynder Paul G. "The epidemiology of renal disease in Aboriginal Australians." Thesis, The University of Sydney, 1991. https://hdl.handle.net/2123/26311.
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Data from dialysis units in the Northern Territory and, from a rural community screening program in South Australia, suggested there was a high prevalence of renal disease in Aboriginal Australians. This thesis documents the high prevalence of urinary abnormalities in three Aboriginal communities in the Northern Territory and examines factors of aetiological relevance.
After ethical approval from the local institutional ethics committee which has Aboriginal
representation, a randomly chosen group of 327 adults and 180 children consented to participate in pilot studies in three communities. Subsequently a family-based study of 16 families comprising all 219 persons over the age of 10 years was undertaken in the community with the highest prevalence of renal disease.
Subjects with renal disease were identified on the basis of a urinary protein to creatinine ratio of > 50 mg/mmol; this measure was shown to be reproducible; urine was also examined by microscopy and culture. Anthropometric and blood pressure measurements were made under standard conditions. Biochemical parameters were assayed on a sample of blood collected 2 hours after a 75g glucose load (in adults) or at the time of collection of throat and skin swabs for streptococcal surveillance (in children).
In the community with the highest frequency of renal disease, some of the renal disease was not associated with diabetes, but was familial and associated with glomerular haematuria; in this same community glomerular haematuria and significant proteinuria were seen also in children. In all three communities, renal disease was associated with obesity, diabetes mellitus and hypertension. In none of the communities was renal disease associated with HBsAG, or evidence of hepatic disease. Although urinary tract infection was detected in some subjects with diabetes, it was not frequent enough to account for the high prevalence of renal disease in diabetics, nor for the high prevalence of renal disease in the Aboriginal communities studied.
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Yap, Bin Kiat. "Exercise-stress responses of urinary hormones." Thesis, The University of Sydney, 1994. https://hdl.handle.net/2123/26858.
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Knowledge on the effects of episodic or short-term exercise-stress on changes of corticosteroids, androgenic steroids and gonadotropins still remains fragmentary and inconclusive. In this study an alternative approach to investigate these changes, based on the concentration ratios of urinary total testosterone (T), luteinizing hormone (LH), free cortisol(F),cortisone (E) and their primary metabolites tetrahydrocortisol (THF) and tetrahydrocortisone (THE), was developed to profile shortterm exercise-stress responses in healthy, drug-free male athletes and sedentary subjects.
The hormonal concentrations were measured using the sensitive and accurate gas chromatography-mass selective detection (GC MSD) and microparticle enzyme immunoassay (MEIA) analytical techniques which were developed. Stress profiles derived from exercise-stress at V02max, 60-80% VOgmax and 50-55% VOzmax were plotted using the ratios of E/F, THE/F, THE/E, LH/T, LH/F, LH/E, T/F and T/E.
Significant changes between the basal and post-exercise ratios were found to be consistent and more representative of acute changes in stress levels compared to single corticosteroid and testosterone concentrations. Marked changes in LH values were also observed to reflect a dramatic increase in stress— response.
The ratio profiling method was also employed to investigate the urinary hormonal changes of a separate group of athletes who were subjected to mild exercise (50% VOZmax) combined with warm (27°C) and moderate humidity (64%). Variation in temperature and humidity resulted in changes to some of the corticosteroid ratio profiles which were consistent with those found in the earlier trials. In a subsequent field study the swim-stress responses of a group of teenage, male swimmers participating in a typical training session during a competition season were also profiled. Whilst no marked changes were found in the LH and T ratios, some of the corticosteroid ratios showed a trend towards positive significance. The profiles appeared to reflect the level of swim-stress prescribed in the training that was less than an equivalent of treadmill running at 60-80% VOZmax The consistent results obtained from the evaluation of the stress-response profiles showed that the profiling approach based on the proportionate changes of urinary F,E,THF,THE,T and LH levels could be utilised as a reliable method for evaluating the inter-relationship between the various hormones in human stress studies. The method could be of advantage in determining the training status of athletes in competitions and for drug-testing purposes.
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Troster, Charles Micah Smolkin. "Trace analysis of cyclophosphamide and its metabolites in urine by liquid chromatography-tandem mass spectrometry." Thesis, University of British Columbia, 2010. http://hdl.handle.net/2429/29263.
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Cyclophosphamide (CP) is an antineoplastic drug used to treat a wide variety of cancers and immune disorders. CP is also a highly toxic alkylating agent, classified as an IARC Group 1 carcinogen. Workers in health-care environments are vulnerable to occupational exposure to CP, primarily via inhalation and dermal absorption. CP is a prodrug; both its therapeutic effectiveness and toxicity are activated through metabolism. To date, however, no study measuring occupational exposure to CP has successfully analyzed its metabolites. The main objective of this work was to develop an analytical method for CP, as well as its metabolites 4-ketocyclophosphamide (KCP) and carboxyphosphamide (CBP), in urine samples collected from health-care workers at risk of CP exposure.
A liquid chromatography-tandem mass spectrometry (LC/MS-MS) method was optimized for CP, KCP and CBP on two different instruments. Post-column infusion showed that the matrix effects resulting from synthetic urine could be separated from the analyte peaks by LC. Estimated instrument limits of quantitation for CP, KCP and CBP in neat solvent were respectively 4.2, 8.2 and 57 ng/L. These parameters were sufficient to meet a quantitation target of 50 ng/L CP in urine, but suggested a need to reduce sample volume to reach a 2.5 ng/L target for KCP and CBP.
Solid-phase extraction (SPE) was explored as a means to exchange the sample matrix for clean solvent and reduce sample volume. Previously developed SPE methods for CP were not designed to include the more polar metabolites, and thus required modification. The best retention of metabolites was seen on a C¹⁸ sorbent with reduced carbon loading. Retention was improved further under acidic loading conditions, but this had to be controlled carefully since CBP can decompose more rapidly at acidic pH. All three analytes were observed to elute with methanol.
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Cromwell, Wyn Zhang. "An Analysis of the Loops of Henle and Urine Concentrating Mechanisms in the Kangaroo Rat." Thesis, The University of Arizona, 2011. http://hdl.handle.net/10150/144330.
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Batista, Carla Alice Theodoro. "Desenvolvimento de um sensor eletroquímico para análise de lactato em amostras de urina e saliva." Universidade Federal de Juiz de Fora, 2012. https://repositorio.ufjf.br/jspui/handle/ufjf/1532.
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CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior
Neste trabalho são apresentados resultados relacionados a estudos sobre o comportamento eletroquímico do lactato e do peróxido de hidrogênio em diversos eletrodos utilizando voltametria cíclica. O comportamento eletroquímico do peróxido de hidrogênio em eletrodos modificados com filme de Azul da Prússia utilizando eletrodos de ouro, compósito e carbono vítreo também foi investigado. Em uma segunda etapa desenvolveu-se um biossensor eletroquímico para determinar lactato por meio do monitoramento do peróxido de hidrogênio produzido na reação catalisada do lactato com oxigênio na presença da enzima lactato oxidase. Nessa etapa, o trabalho consistiu em imobilizar a enzima lactato oxidase na superfície do eletrodo, previamente, modificado com Azul da Prússia, com o auxílio de Nafion. Após a construção do biossensor e a otimização das condições de análise (vazão, potencial, volume de amostra, quantidade de enzima e volume de Nafion) para obtenção de maior sensibilidade no desenvolvimento do método para a determinação de lactato por análise em fluxo, averiguou-se a repetibilidade das medidas (Clactato = 0,20 mmol.L-1) obtendo-se um desvio padrão de 7,1% para 27 repetições. A frequência analítica foi estimada em 41 injeções por hora. Foi obtido um limite de detecção de 2,95 μmol.L-1 e um limite de quantificação igual a 9,84 μmol.L-1. O biossensor foi aplicado na quantificação de lactato em amostras biológicas (urina e saliva). Os resultados obtidos pelo método proposto foram comparados com aqueles oriundos do uso de método de referência.
This paper presents findings related to studies on the electrochemical behavior of lactate and hydrogen peroxide in various electrodes using cyclic voltammetry. The electrochemical behavior of hydrogen peroxide on modified electrodes with Prussian blue film using gold electrodes, composite and glassy carbon was also investigated. In a second step it was developed an electrochemical biosensor for determining lactate by monitoring the hydrogen peroxide produced in the reaction catalyzed lactate with oxygen in the presence of the enzyme lactate oxidase. At this stage, the work consisted of the enzyme lactate oxidase immobilized on the electrode surface, previously modified with Prussian Blue, with the aid of Nafion®. After the construction and optimization of the biosensor analysis conditions (flow rate, potential, sample volume, amount of enzyme and volume of Nafion) for obtaining greater sensitivity in development of the method for the determination of lactate by flow analysis, it was investigated whether the repeatability of measurements (Clactate = 0.20 mmol.L-1) obtaining a standard deviation of 7.1% to 27 repetitions. The analytical frequency was estimated to 41 injections per hour. It was obtained a detection limit of 2.95 μmol.L-1 and a limit of quantification equal to 9.84 μmol.L-1. The biosensor was applied to the quantification of lactate in biological samples (urine and saliva). The results obtained by the proposed method were compared with those from the use of the reference method.
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Lugogo, Rita de Nicolo. "Quantitative assessment of daily urinary conjugates in an adult male population." Diss., This resource online, 1992. http://scholar.lib.vt.edu/theses/available/etd-06062008-171211/.
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Avery, Thomas W. "Further characterization of the direct injection nebulizer for flow injection analysis and liquid chromatography with inductively coupled plasma spectrometric detection." Virtual Press, 1988. http://liblink.bsu.edu/uhtbin/catkey/539620.
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A direct injection nebulizer (DIN) was constructed in our laboratory and was evaluated as an interface between a liquid chromatography column and an inductively coupled plasma-atomic emission spectrometer (ICP-AES). Optimum operating conditions, detection limits and reproducibility of the DIN closely matched literature data for a somewhat different commercial device. In addition, when using the DIN for sample introduction, the ICP detection exhibited uniform response towards phosphorous compounds of different volatilities.Department of Chemistry
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Prado, Isabelle. "L'examen bactériologique des urines a-t-il encore quelque intérêt ?" Caen, 1991. http://www.theses.fr/1991CAEN3039.
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