Analysis of urinary potassium isotopes and association with pancreatic health: healthy, diabetic and cancerous states.

Background: More than 700 million people worldwide suffer from diseases of the pancreas, such as diabetes, pancreatitis and pancreatic cancer. Often dysregulation of potassium (K+) channels, co-transporters and pumps can promote development and progression of many types of these diseases. The role of K+ transport system in pancreatic cell homeostasis and disease development remains largely unexplored. Potassium isotope analysis (δ41K), however, might have the potential to detect minute changes in metabolic processes relevant for pancreatic diseases. Methods: We assessed urinary K isotope composition in a case-control study by measuring K concentrations and δ41K in spot urines collected from patients diagnosed with pancreatic cancer (n=18), other pancreas-related diseases (n=14) and compared those data to healthy controls (n=16). Results: Our results show that urinary K+ levels for patients with diseased pancreas (benign and pancreatic cancer) are significantly lower than the healthy controls. For δ41K, the values tend to be higher for individuals with pancreatic cancer (mean δ41K = -0.58 ± 0.33‰) than for healthy individuals (mean δ41K = -0.78 ± 0.19‰) but the difference is not significant (p=0.08). For diabetics, urinary K+ levels are significantly lower (p=0.03) and δ41K is significantly higher (p=0.009) than for the healthy controls. These results suggest that urinary K+ levels and K isotopes can help identify K disturbances related to diabetes, an associated factors of all-cause mortality for diabetics. Conclusion: Although the K isotope results should be considered exploratory and hypothesis-generating and future studies should focus on larger sample size and δ41K analysis of other K-disrupting diseases (e.g., chronic kidney disease), our data hold great promise for K isotopes as disease marker.

Energy Expenditure of a Male and Female Tennis Player during Association of Tennis Professionals/Women's Tennis Association and Grand Slam Events Measured by Doubly Labeled Water.

METHODS: Doubly labeled water assessed TEE during a 17-d period analyzed by days 1 to 7 (P1) and 7-17 (P2) which included a Women's Tennis Association/Association of Tennis Professionals tournament and culminated at the Wimbledon Championships. Daily training and match loads were assessed using a 10-point Borg scale multiplied by time. Match data were provided by video analysis and player tracking technology. RESULTS: The TEE during P1 for the female player was 3383 kcal·d-1 (63.5 kcal·kg-1) fat-free mass (FFM) with 362 points played over 241 min in three matches covering a distance of 2569 m, with an additional 875 min training. During P2, TEE was 3824 kcal·d-1 (71.7 kcal·kg-1) FFM with 706 points played over 519 min during five matches, covering a distance of 7357 m with an additional 795 min training. The TEE during P1 for the male player was 3712 kcal·d-1 (56.3 kcal·kg-1) FFM with 133 points played over 88 min during one match covering 1125 m, with an additional 795 min training. During P2, TEE was 5520 kcal·d-1 (83.7 kcal·kg-1) FFM with 891 points played over 734 min during five matches, covering 10,043 m, with an additional 350 min training. CONCLUSIONS: This novel data positions elite tennis, played at the highest level, as a highly energetic demanding sport, highlighting that nutritional strategies should ensure sufficient energy availability during competition schedules.

[Urinary protein detection by iTRAQ® associated with renal transplant complications and its modification with therapy]. / Detección de proteínas urinarias por iTRAQ® asociadas a complicaciones del trasplante renal y su modificación con la terapia.

BACKGROUND: After renal transplant, surgical, infection complications, as well as graft rejection may occur; early detection through non-invasive markers is the key to change therapy and avoid biopsy. OBJECTIVE: The aime of the study is to determine urine protein profiles in patients undergoing renal transplant with complications and detect its variation when therapy is modified. MATERIAL AND METHODS: Urine samples were collected from patients prior the transplant and various postoperative stages. Urinary protein profiles were obtained by peptide labeling using isobaric isotopes for relative quantification (iTRAQ(®)). RESULTS: A total of 22 patients were included, of whom 12 developed post-transplant complication: 2 with graft rejection (one male and one female) and 10 (6 males and 4 females) in the group of post-transplant infections. Using iTRAQ(®) 15/345 and 28/113 proteins were identified and fulfilled the acceptance criteria, in graft rejection and post-transplant infections group, respectively. CONCLUSIONS: Albumin was the only protein found in both groups, the remaining proteins were different. The 5 proteins with higher scores in graft rejection were: alpha-1-microglobulin, 5'-nucleotidase cytosolic III, retinol-binding protein 4, membrane protein palmitoylated 4, and serine carboxypeptidase, while post-transplant infections were: mitochondrial acetyl-coenzyme A synthetase, putative adenosyl homocysteinase 2, zinc finger protein GLIS1, putative protein FAM157B, and zinc finger protein 615. It remains to elucidate the involvement of each of these in patients with renal transplantation.

Biologic monitoring and surveillance results for the department of veterans affairs' depleted uranium cohort: Lessons learned from sustained exposure over two decades.

BACKGROUND: A small group of Gulf War I veterans wounded in depleted uranium (DU) friendly fire incidents have been monitored in a clinical surveillance program at the Veterans Affairs Medical Center, Baltimore since 1994. METHODS: An in-patient clinical surveillance protocol was performed on 35 members of the cohort, including exposure monitoring for total and isotopic uranium concentrations in urine and a comprehensive assessment of health outcomes. RESULTS: Although urine U concentrations continue to be elevated in this group, illustrating on-going in situ mobilization of U from embedded fragments, no consistent U-related health effects have been observed. CONCLUSIONS: Now more than 20 years since first exposure to DU, an aging cohort of military veterans continues to show no U-related health effects in known target organs of U toxicity. As tissue concentrations continue to accrue with exposure duration, critical tissue-specific U concentration thresholds may be reached, thus recommending on-going surveillance of this veteran cohort.

Measurement of isotope abundance variations in nature by gravimetric spiking isotope dilution analysis (GS-IDA).

Subtle variations in the isotopic composition of elements carry unique information about physical and chemical processes in nature and are now exploited widely in diverse areas of research. Reliable measurement of natural isotope abundance variations is among the biggest challenges in inorganic mass spectrometry as they are highly sensitive to methodological bias. For decades, double spiking of the sample with a mix of two stable isotopes has been considered the reference technique for measuring such variations both by multicollector-inductively coupled plasma mass spectrometry (MC-ICPMS) and multicollector-thermal ionization mass spectrometry (MC-TIMS). However, this technique can only be applied to elements having at least four stable isotopes. Here we present a novel approach that requires measurement of three isotope signals only and which is more robust than the conventional double spiking technique. This became possible by gravimetric mixing of the sample with an isotopic spike in different proportions and by applying principles of isotope dilution for data analysis (GS-IDA). The potential and principle use of the technique is demonstrated for Mg in human urine using MC-TIMS for isotopic analysis. Mg is an element inaccessible to double spiking methods as it consists of three stable isotopes only and shows great potential for metabolically induced isotope effects waiting to be explored.

Profiling of mercapturic acids of acrolein and acrylamide in human urine after consumption of potato crisps.

SCOPE: Acrolein (AC) and acrylamide (AA) are food contaminants generated by heat treatment. We studied human exposure after consumption of potato crisps by monitoring excretion of mercapturic acids (MAs) in urine. METHODS AND RESULTS: MA excretion was monitored in human urine collected up to 72 h after ingestion of a test meal of experimental (study 1: 1 mg AA/150 g) or commercially available (study 2: 44 µg AA plus 4.6 µg AC/175 g) potato crisps. MA contents were analysed after purification via SPE using HPLC-ESI-MS/MS. On the basis of the area under the curve values of MAs excreted in urine, the total excretion of AC-related MAs exceeded that of AA-related MAs up to 12 times in study 1 and up to four times in study 2. Remarkably, AC content of potato crisps of study 2 was found to be only about 1/10 the AA content, as determined by isotope dilution headspace GC/MS. CONCLUSION: Our results indicate substantially higher exposure to AC from potato crisps than to AA. Total AC in such foods may encompass bioavailable AC forms not detected by headspace GC/MS. Both findings may also apply to other heat processed foods.

Human urinary excretion and metabolism of (82)Se-enriched selenite and selenate determined by LC-ICP-MS.

Urinary excretion of selenium after ingestion of isotope labeled selenite and selenate was studied in seven healthy volunteers, 4 men and 3 women (age 28-50 years). An aqueous solution containing 330 µL (82)Se-selenate (corresponding to 74.3 µg (82)Se) was given orally and urine samples were subsequently collected during the following 24 hours. The scheme was repeated four weeks later with a 280 µL (82)Se-selenite solution (corresponding to 74.4 µg (82)Se). The amount of total Se in the urine samples was determined by inductively coupled mass spectrometry. The mean total urinary excretion of (82)Se following (82)Se-selenate administration was 33.7% (range 15.6-42.5%) while the mean total excretion of (82)Se after (82)Se-selenite administration was 3.2% (range 2.8-3.9%) of the ingested amount. LC-ICPMS analysis of the urine samples showed that the majority of the selenium excreted after selenate ingestion was unchanged selenate for 6 of the individuals while one individual had metabolized a fraction (approx. 20%) of the selenate to selenosugar. Ingestion of 10 times larger doses of selenite in two individuals resulted in 13-23% excretion primarily excreted as selenosugar. These results show that the human metabolic pathways of selenite and selenate are different and indicate that not all selenate, although well absorbed, may be available for the beneficial health effects.

Biological fractionation of lead isotopes in Sprague-Dawley rats lead poisoned via the respiratory tract.

OBJECTIVES: It was considered that lead isotope ratios did not change during physical, chemical, or biological processes. Thus, lead isotope ratios have been used as fingerprints to identify possible lead sources. However, recent evidence has shown that the lead isotope ratios among different biological samples in human are not always identical from its lead origins in vitro. An animal experiment was conducted to explore the biological fractionation of lead isotopes in biological systems. METHODS: 24 male Sprague-Dawley (SD) rats were divided into groups that received acute lead exposure (0, 0.02, 0.2, or 2 mg/kg body weight of lead acetate) via the respiratory route every day for 5 days. Biological samples (i.e., blood, urine, and feces) were collected for comparison with the lead acetate (test substance) and the low-lead animal feed (diet) administered to the rats. The lead isotope ratios were determined by inductively coupled plasma mass spectrometry (ICP-MS). RESULTS: There are significant differences (p<0.05) in lead isotope ratios between blood, urine, and feces. Moreover, a nonlinear relationship between the blood lead concentration and the blood lead isotope ratios was observed. There is also a threshold effect to the fractionation function. Only the blood isotope ratio of (204)Pb/(206)Pb matches the test substance well. As for feces, when (204)Pb/(206)Pb ratio is considered, there is no significant difference between feces-test substance pairs in medium and high dose group. CONCLUSIONS: The biological fractionation of lead isotopes in SD rats was observed. Moreover, there might be a threshold for the biological fractionation of lead isotopes which is depending on whole blood lead level. It is considered to be more reliable that we compared the isotope ratios of potential lead hazards with both blood and feces lead fingerprints especially for (204)Pb/(206)Pb ratio under high-dose exposure.

Rapid speciation and quantification of selenium compounds by HPLC-ICP MS using multiple standards labelled with different isotopes.

Speciation analysis using high-performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC-ICP MS) is now commonly used to investigate metabolic and toxicological aspects of some metals and metalloids. We have developed a rapid method for simultaneous identification and quantification of metabolites of selenium (Se) compounds using multiple standards labelled with different isotopes. A mixture of the labelled standards was spiked in a selenised garlic extract and the sample was subjected to speciation analysis by HPLC-ICP MS. The selenised garlic contains γ-glutamyl-methylselenocysteine, methylselenocysteine, and selenomethionine and the concentrations of those Se compounds were 723.8, 414.8, and 310.7 ng Se ml(-1), respectively. The isotopically labelled standards were also applied to the speciation of Se in rat urine. Selenate, methylselenonic acid, selenosugar, and trimethyselenium ions were found to be excreted by the present speciation procedure. Multiple standards labelled with different stable isotopes enable high-throughput identification and quantitative measurements of Se metabolites.

Influence of the consumption pattern of magnesium from magnesium-rich mineral water on magnesium bioavailability.

It is generally considered that the absorption of Mg is inversely related to the ingested dose. The objective of the present study was to determine if the mode of administration (bolus v. consumption throughout the day) could influence Mg bioavailability from Mg-rich natural mineral water comparing the same nutritional Mg amount (126 mg). Using a 2 d cross-over design, twelve healthy men were asked to drink 1·5 litres Mg-rich mineral water either as 2 × 750 ml or 7 × 212 ml throughout the day. Two stable isotopes ((25)Mg and (26)Mg) were used to label the water in order to distinguish both regimens. Fractional apparent Mg absorption was determined by faecal monitoring and Mg retention was determined by measuring urinary excretion of Mg isotopes. Higher Mg absorption (50·7 (SD 12·7) v. 32·4 (SD 8·1) %; P = 0·0007) and retention (47·5 (SD 12·9) v. 29·0 (SD 7·5) %; P = 0·0008) from Mg-rich mineral water were observed when it was consumed in seven servings compared with larger servings. Thus, regular water consumption throughout the day is an effective way to increase Mg bioavailability from Mg-rich mineral water.

Intake risk and dose evaluation methods for workers in radiochemistry labs of a medical cyclotron facility.

The aim of this paper is to evaluate the risks and doses for the internal contamination of the radiochemistry staff in a high workload medical cyclotron facility. The doses from internal contamination derive from the inhalation of radioactive gas leakage from the cells by personnel involved in the synthesis processes and are calculated from urine sample measurements. Various models are considered for the calculation of the effective committed dose from the analysis of these urine samples, and the results are compared with data obtained from local environmental measurement of the radioactivity released inside the lab.

Selenium metabolism in rats with long-term ingestion of Se-methylselenocysteine using enriched stable isotopes.

Se-methylselenocysteine (MeSeCys) is not only a selenium (Se) supplement but also a more promising precursor of an anti-tumor drug containing Se than selenomethionine, which is currently used as Se supplement. In this study, the metabolism of MeSeCys labeled with an Se isotope, 82Se, in rats depleted of endogenous natural abundance isotopes with another Se isotope, 78Se, was traced for 21 days when MeSeCys was continuously and perorally ingested at a supplemental dose. The tracer experiment was performed with our improved method that utilized an inductively coupled plasma-deuterium reaction-mass spectrometer. The substitution of endogenous Se with a single isotope, 78Se, facilitated the detection of exogenous labeled Se. Exogenous Se in the form of MeSeCys preferably accumulated and/or assimilated in the liver, kidneys and testes with long-term ingestion of MeSeCys and was utilized for the synthesis of selenoproteins, i.e., extracellular and cellular glutathione peroxidases and selenoprotein P. Meanwhile, intact MeSeCys was not excreted into urine although trimethylselenonium was detected in addition to selenosugar. The results suggest that MeSeCys was transformed into selenide via methylselenol by beta-lyase. Consequently, it is surmised that MeSeCys is a precursor of methylselenol under long-term ingestion.

Urinary isotopic analysis in the UK Armed Forces: no evidence of depleted uranium absorption in combat and other personnel in Iraq.

OBJECTIVES: To assess the distribution and risk factors of depleted uranium uptake in military personnel who had taken part in the invasion of Iraq in 2003. METHODS: Sector field inductively coupled plasma-mass spectrometry (SF-ICP-MS) was used to determine the uranium concentration and (238)U/(235)U isotopic ratio in spot urine samples. The authors collected urine samples from four groups identified a priori as having different potential for exposure to depleted uranium. These groups were: combat personnel (n = 199); non-combat personnel (n = 96); medical personnel (n = 22); and "clean-up" personnel (n = 24) who had been involved in the maintenance, repair or clearance of potentially contaminated vehicles in Iraq. A short questionnaire was used to ascertain individual experience of circumstances in which depleted uranium exposure might have occurred. RESULTS: There was no statistically significant difference in the (238)U/(235)U ratio between groups. Mean ratios by group varied from 138.0 (95% CI 137.3 to 138.7) for clean-up personnel to 138.2 (95% CI 138.0 to 138.5) for combat personnel, and were close to the ratio of 137.9 for natural uranium. The two highest individual ratios (146.9 and 147.7) were retested using more accurate, multiple collector inductively coupled plasma-mass spectrometry (MC-ICP-MS) and found to be within measurement of error of that for natural uranium. There were no significant differences in isotope ratio between participants according to self-reported circumstances of potential depleted uranium exposure. CONCLUSIONS: Based on measurements using a SF-ICP-MS apparatus, this study provides reassurance following concern for potential widespread depleted uranium uptake in the UK military. The rare occurrence of elevated ratios may reflect the limits of accuracy of the SF-ICP-MS apparatus and not a real increase from the natural proportions of the isotopes. Any uptake of depleted uranium among participants in this study sample would be very unlikely to have any implications for health.

Diagnosis and management of Wilson's disease: results of a single center experience.

AIMS: To report on the diagnostic features, management, and clinical outcome after different treatments of Wilson's disease patients followed over a mean period of 15 years. PATIENTS: Thirty-five patients with Wilson's disease referred to the University of Padova's Department of Gastroenterology for diagnosis or treatment were observed for a mean 15 years. The diagnosis was based on clinical symptoms, laboratory tests (ceruloplasmin, urinary, and hepatic copper concentrations), and uptake of the radiostable isotope Cu into the plasma protein pool. Hepatic Cu content was measured by regular follow-up biopsies. Neurologic outcome after therapy was assessed using a newly developed scoring system. RESULTS: Twenty-three (65.7%) patients presented with liver disease; 12 (34.3%) had mixed neurologic and hepatic involvement. All patients had been initially treated with either penicillamine (23) or zinc sulfate (12). The neurologic symptoms became worse or remained stationary in 75% of those treated with penicillamine, whereas zinc treatment improved these symptoms in 90% of treated cases. Both treatments were effective in improving the hepatic symptoms. No differences in hepatic Cu content emerged between follow-up biopsies in either treatment group. Six patients (26%) had to abandon the penicillamine treatment due to side effects. In all, 4 patients underwent liver transplantation, which was successful in 3, with a mean survival after transplantation of 4.6 years; the fourth, who had a severe neurologic impairment, died of central pontine myelinolysis. CONCLUSIONS: Penicillamine and zinc can effectively treat Wilson's disease, though the side effects of penicillamine may be severe enough to prompt its suspension. Liver transplantation remains the treatment of choice for end-stage liver disease.

Development of a multianalyte method for the determination of anabolic hormones in bovine urine by isotope-dilution GC-MS/MS.

A sensitive, specific and selective multianalyte GC-MS/MS method has been developed for the determination of 11 anabolic hormones in bovine urine. After adjusting the urine pH to 4.8, the samples were spiked with deuterated internal standards and submitted to enzymatic hydrolysis with beta-glucuronidase/arylsulfatase. Hormones were eluted with methanol through a C18 solid phase cartridge and submitted to a liquid-liquid extraction. Analytes were derivatized by adding N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) with 1% trimethylchlorosilane and GC-MS data were obtained in the positive electron impact tandem mass mode. Under these conditions, no matrix effects were observed and limit of detection values were in the range of 0.005 ng/mL (diethylstilbestrol) to 0.38 ng/mL (17alpha-methyltestosterone and 17alpha-ethynylestradiol). Recoveries from 81% (alpha-zeranol) to 149% (17alpha-methyltestosterone) were found under the selected conditions. These results were better than those found using heptafluorobutyric anhydride (HFBA) as derivative reagent and those measured in full scan and selective ion monitoring modes.

Flow-injection technique for determination of uranium and thorium isotopes in urine by inductively coupled plasma mass spectrometry.

A sensitive and efficient flow-injection (FI) preconcentration and matrix-separation technique coupled to sector field ICP-mass spectrometry (SF-ICP-MS) has been developed and validated for simultaneous determination of ultra-low levels of uranium (U) and thorium (Th) in human urine. The method is based on selective retention of U and Th from a urine matrix, after microwave digestion, on an extraction chromatographic TRU resin, as an alternative to U/TEVA resin, and their subsequent elution with ammonium oxalate. Using a 10 mL sample, the limits of detection achieved for 238U and 232Th were 0.02 and 0.03 ng L(-1), respectively. The accuracy of the method was checked by spike-recovery measurements. Levels of U and Th in human urine were found to be in the ranges 1.86-5.50 and 0.176-2.35 ng L(-1), respectively, well in agreement with levels considered normal for non-occupationally exposed persons. The precision obtained for five replicate measurements of a urine sample was 2 and 3% for U and Th, respectively. The method also enables on-line measurements of the 235U/238U isotope ratios in urine. Precision of 0.82-1.04% (RSD) was obtained for 235U/238U at low ng L(-1) levels, using the FI transient signal approach.

Effects of sulphate- and bicarbonate-rich mineral waters on net and fractional intestinal absorption and urinary excretion of magnesium in rats.

Magnesium (Mg) intake is below the recommended daily allowances in many developed countries. Mg-rich mineral waters can provide significant amounts of energy-free Mg and thus help to meet Mg requirements. We assessed the effects of different Mg-rich mineral waters on overall intestinal Mg absorption and urinary Mg excretion in 40 rats split into four groups: one received distilled water, another a solution of MgCl(2) and the others two different mineral waters, sulphated water (Hépar) and carbonated water (Badoit) mixed with the diet and as drinking water, for four weeks. The rats were given 3 mg of (26)Mg orally and 0.5 mg of (25)Mg intravenously. They were placed in metabolic cages, and diet consumption, and faeces and urine excretion were monitored during the last four days of the experiment. The rats were then sacrificed and blood was sampled. Mg levels in the diet, faeces, urine and biological samples were measured by atomic absorption spectrometry. Mg stable isotope measurements were performed by ICP/MS. Mg-rich mineral waters significantly increased net intestinal absorption of Mg by more than 30%, but the proportions of both apparent and true intestinal absorption of Mg were similar in all four groups. Thus, net and fractional retention of Mg were similar in the three Mg-supplemented groups. In conclusion, both types of Mg-rich mineral waters studied similarly increased both absorption and urinary excretion of Mg with no positive effect on the overall retention of Mg, probably because the Mg status of the rats was already satisfactory.

Measurement of magnesium absorption and retention in type 2 diabetic patients with the use of stable isotopes.

BACKGROUND: Magnesium deficiency has been associated with type 2 diabetes and may reduce insulin sensitivity and impair glucose tolerance. The etiology of magnesium depletion in diabetes is unclear. Animal studies suggest that diabetes may impair magnesium absorption; however, there are no published data on magnesium absorption in humans with diabetes. OBJECTIVE: Magnesium absorption from a test meal and the excretion and retention of magnesium were compared between patients with type 2 diabetes and healthy control subjects. DESIGN: A meal labeled with 10 mg (26)Mg isotopic label was administered, and stool and urine samples were collected for 10 and 6 d, respectively. Apparent absorption was calculated as the difference between the oral dose of (26)Mg isotopic label and the total amount of the isotopic label excreted in the feces. Magnesium retention was calculated from the apparent absorption and urinary excretion of (26)Mg isotopic label in the 6 d after administration. RESULTS: Mean (+/- SD) values for fractional magnesium absorption in the diabetic patients and the control subjects were 59.3 +/- 7.0% and 57.6 +/- 8.5%, respectively (NS). Mean (+/- SD) urinary magnesium excretion values in the diabetic patients and the control subjects were 11.2 +/- 2.6% and 11.7 +/- 3.8%, respectively (NS); retention values were 54.2 +/- 7.1% and 51.4 +/- 6.1%, respectively (NS). CONCLUSION: Dietary magnesium absorption and retention are not impaired in patients with reasonably well-controlled type 2 diabetes.

The quantitative analysis of depleted uranium isotopes in British, Canadian, and U.S. Gulf War veterans.

The purpose of this work was to determine the concentration and ratio of uranium isotopes in allied forces Gulf War veterans. The 27 patients had their 24-hour urine samples analyzed for 234U, 235U, 236U, and 238U by mass spectrometry. The urine samples were evaporated and separated into isotopic dilution and concentration fraction by the chromatographic technique. The isotopic composition was measured by a thermal ionization mass spectrometer using a secondary electron multiplier detector and ion-counting system. The uranium blank control and SRM960 U isotopic standard were analyzed by the same procedure. Statistical analysis was done by an unpaired t test. The results confirm the presence of depleted uranium (DU) in 14 of 27 samples, with the 238U:235U ratio > 207.15. This is significantly different from natural uranium (p < 0.008) as well as from the DU shrapnel analysis, with 22.22% average value of DU fraction, and warrants further investigation.