1H-NMR-based urinary metabolomic analysis for the preventive effects of gushudan on glucocorticoid-induced osteoporosis rats.

Gushudan (GSD), a traditional Chinese medicine with a history of more than 15 years, has been shown to have anti-osteoporosis effects, but the specific therapeutic mechanism behind it is still unclear. To further elucidate the pathogenesis of osteoporosis and the preventive mechanism of GSD on glucocorticoid-induced osteoporosis (GIOP) rats, a rapid and comprehensive 1H NMR metabolomics method was established to detect urinary metabolic profiles in the control group, model group and GSD treatment group in this study. The orthogonal partial least squares discriminant analysis (OPLS-DA) was performed to investigate changes in the metabolites, and related metabolic pathways were discovered using MetaboAnalyst platform. As a result, a total of 27 differential metabolites were identified. Of these, 17 metabolites such as formate, allantoin and l-threonate were newly discovered as GIOP potential biomarkers. Energy metabolism, intestinal flora metabolism, amino acid metabolism and oxidative stress response were significantly changed in the urinary profiles of GIOP rats, and GSD could play an anti-osteoporosis role by regulating these metabolic pathways. This study compliments the earlier LC-MS based urine metabolomics research, and helps further understand the pathogenesis of osteoporosis and the potential preventive effects of GSD on GIOP rats.

Multi-Compartment Profiling of Bacterial and Host Metabolites Identifies Intestinal Dysbiosis and Its Functional Consequences in the Critically Ill Child.

OBJECTIVES: Adverse physiology and antibiotic exposure devastate the intestinal microbiome in critical illness. Time and cost implications limit the immediate clinical potential of microbial sequencing to identify or treat intestinal dysbiosis. Here, we examined whether metabolic profiling is a feasible method of monitoring intestinal dysbiosis in critically ill children. DESIGN: Prospective multicenter cohort study. SETTING: Three U.K.-based PICUs. PATIENTS: Mechanically ventilated critically ill (n = 60) and age-matched healthy children (n = 55). INTERVENTIONS: Collection of urine and fecal samples in children admitted to the PICU. A single fecal and urine sample was collected in healthy controls. MEASUREMENTS AND MAIN RESULTS: Untargeted and targeted metabolic profiling using 1H-nuclear magnetic resonance spectroscopy and liquid chromatography-mass spectrometry or urine and fecal samples. This was integrated with analysis of fecal bacterial 16S ribosomal RNA profiles and clinical disease severity indicators. We observed separation of global urinary and fecal metabolic profiles in critically ill compared with healthy children. Urinary excretion of mammalian-microbial co-metabolites hippurate, 4-cresol sulphate, and formate were reduced in critical illness compared with healthy children. Reduced fecal excretion of short-chain fatty acids (including butyrate, propionate, and acetate) were observed in the patient cohort, demonstrating that these metabolites also distinguished between critical illness and health. Dysregulation of intestinal bile metabolism was evidenced by increased primary and reduced secondary fecal bile acid excretion. Fecal butyrate correlated with days free of intensive care at 30 days (r = 0.38; p = 0.03), while urinary formate correlated inversely with vasopressor requirement (r = -0.2; p = 0.037). CONCLUSIONS: Disruption to the functional activity of the intestinal microbiome may result in worsening organ failure in the critically ill child. Profiling of bacterial metabolites in fecal and urine samples may support identification and treatment of intestinal dysbiosis in critical illness.

First and second trimester urinary metabolic profiles and fetal growth restriction: an exploratory nested case-control study within the infant development and environment study.

BACKGROUND: Routine prenatal care fails to identify a large proportion of women at risk of fetal growth restriction (FGR). Metabolomics, the comprehensive analysis of low molecular weight molecules (metabolites) in biological samples, can provide new and earlier biomarkers of prenatal health. Recent research has suggested possible predictive first trimester urine metabolites correlating to fetal growth restriction in the third trimester. Our objective in this current study was to examine urinary metabolic profiles in the first and second trimester of pregnancy in relation to third trimester FGR in a US population from a large, multi-center cohort study of healthy pregnant women. METHODS: We conducted a nested case-control study within The Infant Development and the Environment Study (TIDES), a population-based multi-center pregnancy cohort study. We identified 53 cases of FGR based on the AUDIPOG [Neonatal growth - AUDIPOG [Internet]. [cited 29 Nov 2016]. Available from: http://www.audipog.net/courbes_morpho.php?langue=en ] formula for birthweight percentile considering maternal height, age, and prenatal weight, as well as infant sex, gestational age, and birth rank. Cases were matched to 106 controls based on study site, maternal age (± 2 years), parity, and infant sex. NMR spectroscopy was used to assess concentrations of four urinary metabolites that have been previously associated with FGR (tyrosine, acetate, formate, and trimethylamine) in first and second trimester urine samples. We fit multivariate conditional logistic regression models to estimate the odds of FGR in relation to urinary concentrations of these individual metabolites in the first and second trimesters. Exploratory analyses of custom binned spectroscopy results were run to consider other potentially related metabolites. RESULTS: We found no significant association between the relative concentrations of each of the four metabolites and odds of FGR. Exploratory analyses did not reveal any significant differences in urinary metabolic profiles. Compared with controls, cases delivered earlier (38.6 vs 39.8, p < 0.001), and had lower birthweights (2527 g vs 3471 g, p < 0.001). Maternal BMI was similar between cases and controls. CONCLUSIONS: First and second trimester concentrations of urinary metabolites (acetate, formate, trimethylamine and tyrosine) did not predict FGR. This inconsistency with previous studies highlights the need for more rigorous investigation and data collection in this area before metabolomics can be clinically applied to obstetrics.

Evaluation of genotoxicity in workers exposed to low levels of formaldehyde in a furniture manufacturing facility.

Formaldehyde (FA) is a chemical widely used in the furniture industry and has been classified as a potential human carcinogen. The purpose of this study was to evaluate the occupational exposure of workers to FA at a furniture manufacturing facility and the relationship between environmental concentrations of FA, formic acid concentration in urine, and DNA damage. The sample consisted of 46 workers exposed to FA and a control group of 45 individuals with no history of occupational exposure. Environmental concentrations of FA were determined by high-performance liquid chromatography. Urinary formic acid concentrations were determined by gas chromatography with flame ionization detector. DNA damage was evaluated by the micronucleus (MN) test performed in exfoliated buccal cells and comet assay with venous blood. The 8-h time-weighted average of FA environmental concentration ranged from 0.03 ppm to 0.09 ppm at the plant, and the control group was exposed to a mean concentration of 0.012 ppm. Workers exposed to higher environmental FA concentrations had urinary formic acid concentrations significantly different from those of controls (31.85 mg L(-1) vs. 19.35 mg L(-), p ≤ 0.01 Mann-Whitney test). Significant differences were found between control and exposed groups for the following parameters: damage frequency and damage index in the comet assay, frequency of binucleated cells in the MN test, and formic acid concentration in urine. The frequency of micronuclei, nuclear buds, and karyorrhexis did not differ between groups. There was a positive correlation between environmental concentrations of FA and damage frequency (Spearman's rank correlation coefficient [r s] = 0.24), damage index (r s = 0.21), binucleated cells (r s = 0.34), and urinary formic acid concentration (r s = 0.63). The results indicate that, although workers in the furniture manufacturing facility were exposed to low environmental levels of FA, this agent contributes to the observed increase in cytogenetic damage. In addition, urinary formic acid concentrations correlated strongly with occupational exposure to FA.

Assessment of lipid peroxidation and p53 as a biomarker of carcinogenesis among workers exposed to formaldehyde in the cosmetic industry.

Despite the wide use of cosmetic products, they exert a number of health effects on tissues ranging from irritation to cancer. Our study aimed at assessing the effect of formaldehyde on lipid peroxidation and verifying the susceptibility to carcinogenesis using p53 as a biomarker among workers exposed to formaldehyde in cosmetic industry. Our entire exposed group (n = 40) and the controls (n = 20) were subjected to estimation of formate in urine, serum malondialdehyde (MDA), and p53. Also, complete blood picture, liver, and kidney function tests were carried out. The study revealed significant increase in the levels of formate, MDA, and p53 in the exposed group compared with their control group. Our results showed that workers in cosmetic industry had significant exposure to formaldehyde. Furthermore, the study pointed to the negative impact of formaldehyde as a cause of oxidative stress and suspicious carcinogen.

Divergent Urinary Metabolic Phenotypes between Major Depressive Disorder and Bipolar Disorder Identified by a Combined GC-MS and NMR Spectroscopic Metabonomic Approach.

Bipolar disorder (BD) is a complex debilitating mental disorder that is often misdiagnosed as major depressive disorder (MDD). Therefore, a large percentage of BD subjects are incorrectly treated with antidepressants in clinical practice. To address this challenge, objective laboratory-based tests are needed to discriminate BD from MDD patients. Here, a combined gas chromatography-mass spectrometry (GC-MS)-based and nuclear magnetic resonance (NMR) spectroscopic-based metabonomic approach was performed to profile urine samples from 76 MDD and 43 BD subjects (training set) to identify the differential metabolites. Samples from 126 healthy controls were included as metabolic controls. A candidate biomarker panel was identified by further analyzing these differential metabolites. A testing set of, 50 MDD and 28 BD subjects was then used to independently validate the diagnostic efficacy of the identified panel using an area under the receiver operating characteristic curve (AUC). A total of 20 differential metabolites responsible for the discrimination between MDD and BD subjects were identified. A panel consisting of six candidate urinary metabolite biomarkers (propionate, formate, (R*,S*)2,3-dihydroxybutanoic acid, 2,4-dihydroxypyrimidine, phenylalanine, and ß-alanine) was identified. This panel could distinguish BD from MDD subjects with an AUC of 0.913 and 0.896 in the training and testing sets, respectively. These results reveal divergent urinary metabolic phenotypes between MDD and BD. The identified urinary biomarkers can aid in the future development of an objective laboratory-based diagnostic test for distinguishing BD from MDD patients.

Identification and validation of urinary metabolite biomarkers for major depressive disorder.

Major depressive disorder (MDD) is a widespread and debilitating mental disorder. However, there are no biomarkers available to aid in the diagnosis of this disorder. In this study, a nuclear magnetic resonance spectroscopy-based metabonomic approach was employed to profile urine samples from 82 first-episode drug-naïve depressed subjects and 82 healthy controls (the training set) in order to identify urinary metabolite biomarkers for MDD. Then, 44 unselected depressed subjects and 52 healthy controls (the test set) were used to independently validate the diagnostic generalizability of these biomarkers. A panel of five urinary metabolite biomarkers-malonate, formate, N-methylnicotinamide, m-hydroxyphenylacetate, and alanine-was identified. This panel was capable of distinguishing depressed subjects from healthy controls with an area under the receiver operating characteristic curve (AUC) of 0.81 in the training set. Moreover, this panel could classify blinded samples from the test set with an AUC of 0.89. These findings demonstrate that this urinary metabolite biomarker panel can aid in the future development of a urine-based diagnostic test for MDD.

An isotope-dilution, GC-MS assay for formate and its application to human and animal metabolism.

Formate, a crucial component of one-carbon metabolism, is increasingly recognized as an important intermediate in production and transport of one-carbon units. Unlike tetrahydrofolate-linked intermediates, it is not restricted to the intracellular milieu so that circulating levels of formate can provide insight into cellular events. We report a novel isotope-dilution, GC-MS assay employing derivatization by 2,3,4,5,6-pentafluorobenzyl bromide for the determination of formate in biological samples. This assay is robust and sensitive; it may be applied to the measurement of formate in serum, plasma and urine. We demonstrate how this method may be applied by providing the first characterization of formate levels in a human population; formate levels were higher in males than in females. We also show how this procedure may be applied for the measurement of in vivo kinetics of endogenous formate production in experimental animals.

Formate: an essential metabolite, a biomarker, or more?

Plasma and urinary formate concentrations were recently found to be elevated during vitamin B12 and folate deficiencies. It was proposed that formate may be a valuable biomarker of impaired one-carbon metabolism. Formate is an essential intermediary metabolite in folate-mediated one-carbon metabolism and, despite its importance, our knowledge of its metabolism is limited. Formate can be produced from several substrates (e.g., methanol, branched chain fatty acids, amino acids), some reactions being folate-dependent while others are not. Formate removal proceeds via two pathways; the major one being folate-dependent. Formate is a potentially toxic molecule and we suggest that formate may play a role in some of the pathologies associated with defective one-carbon metabolism.

Formate can differentiate between hyperhomocysteinemia due to impaired remethylation and impaired transsulfuration.

Formate can differentiate between hyperhomocysteinemia due to impaired remethylation and impaired transsulfuration. Am J Physiol Endocrinol Metab 301: E000-E000, 2011. First published September 20, 2011; 10.1152/ajpendo.00345.2011.-We carried out a (1)H-NMR metabolomic analysis of sera from vitamin B(12)-deficient rats. In addition to the expected increases in methylmalonate and homocysteine (Hcy), we observed an approximately sevenfold increase in formate levels, from 64 µM in control rats to 402 µM in vitamin B(12)-deficient rats. Urinary formate was also elevated. This elevation of formate could be attributed to impaired one-carbon metabolism since formate is assimilated into the one-carbon pool by incorporation into 10-formyl-THF via the enzyme 10-formyl-THF synthase. Both plasma and urinary formate were also increased in folate-deficient rats. Hcy was elevated in both the vitamin B(12)- and folate-deficient rats. Although plasma Hcy was also elevated, plasma formate was unaffected in vitamin B(6)-deficient rats (impaired transsulfuration pathway). These results were in accord with a mathematical model of folate metabolism, which predicted that reduction in methionine synthase activity would cause increased formate levels, whereas reduced cystathionine ß-synthase activity would not. Our data indicate that formate provides a novel window into cellular folate metabolism, that elevated formate can be a useful indicator of deranged one-carbon metabolism and can be used to discriminate between the hyperhomocysteinemia caused by defects in the remethylation and transsulfuration pathways.

(1)H nuclear magnetic resonance metabolomics analysis identifies novel urinary biomarkers for lung function.

Chronic obstructive pulmonary disease (COPD), characterized by chronic airflow limitation, is a serious public health concern. In this study, we used proton nuclear magnetic resonance ((1)H NMR) spectroscopy to identify and quantify metabolites associated with lung function in COPD. Plasma and urine were collected from 197 adults with COPD and from 195 without COPD. Samples were assayed using a 600 MHz NMR spectrometer, and the resulting spectra were analyzed against quantitative spirometric measures of lung function. After correcting for false discoveries and adjusting for covariates (sex, age, smoking) several spectral regions in urine were found to be significantly associated with baseline lung function. These regions correspond to the metabolites trigonelline, hippurate and formate. Concentrations of each metabolite, standardized to urinary creatinine, were associated with baseline lung function (minimum p-value = 0.0002 for trigonelline). No significant associations were found with plasma metabolites. Urinary hippurate and formate are often related to gut microflora. This could suggest that the microbiome varies between individuals with different lung function. Alternatively, the associated metabolites may reflect lifestyle differences affecting overall health. Our results will require replication and validation, but demonstrate the utility of NMR metabolomics as a screening tool for identifying novel biomarkers of pulmonary outcomes.

(1)H NMR-based metabonomic investigation of the effect of two different exercise sessions on the metabolic fingerprint of human urine.

Physical exercise modifies animal metabolism profoundly. Until recently, biochemical investigations related to exercise focused on a small number of biomolecules. In the present study, we used a holistic analytical approach to investigate changes in the human urine metabolome elicited by two exercise sessions differing in the duration of the rest interval between repeated efforts. Twelve men performed three sets of two 80 m maximal runs separated by either 10 s or 1 min of rest. Analysis of pre- and postexercise urine samples by (1)H NMR spectroscopy and subsequent multivariate statistical analysis revealed alterations in the levels of 22 metabolites. Urine samples were safely classified according to exercise protocol even when applying unsupervised methods of statistical analysis. Separation of pre- from postexercise samples was mainly due to lactate, pyruvate, hypoxanthine, compounds of the Krebs cycle, amino acids, and products of branched-chain amino acid (BCAA) catabolism. Separation of the two rest intervals was mainly due to lactate, pyruvate, alanine, compounds of the Krebs cycle, and 2-oxoacids of BCAA, all of which increased more with the shorter interval. Metabonomics provides a powerful methodology to gain insight in metabolic changes induced by specific training protocols and may thus advance our knowledge of exercise biochemistry.

Characterization of inflammatory bowel disease with urinary metabolic profiling.

OBJECTIVES: Distinguishing between the inflammatory bowel disease (IBD), Crohn's disease (CD), and ulcerative colitis (UC) is important for both management and prognostic reasons. Discrimination using noninvasive techniques could be an adjunct to conventional diagnostics. Differences have been shown between the intestinal microbiota of CD and UC patients and controls; the gut bacteria influence specific urinary metabolites that are quantifiable using proton high-resolution nuclear magnetic resonance (NMR) spectroscopy. This study tested the hypothesis that such metabolites differ between IBD and control cohorts, and that using multivariate pattern-recognition analysis, the cohorts could be distinguished by urine NMR spectroscopy. METHODS: NMR spectra were acquired from urine samples of 206 Caucasian subjects (86 CD patients, 60 UC patients, and 60 healthy controls). Longitudinal samples were collected from 75 individuals. NMR resonances specific for metabolites influenced by the gut microbes were studied, including hippurate, formate, and 4-cresol sulfate. Multivariate analysis of all urinary metabolites involved principal components analysis (PCA) and partial least squares discriminant analysis (PLS-DA). RESULTS: Hippurate levels were lowest in CD patients and differed significantly between the three cohorts (P<0.0001). Formate levels were higher and 4-cresol sulfate levels lower in CD patients than in UC patients or controls (P=0.0005 and P=0.0002, respectively). PCA revealed clustering of the groups; PLS-DA modeling was able to distinguish the cohorts. These results were independent of medication and diet and were reproducible in the longitudinal cohort. CONCLUSIONS: Specific urinary metabolites related to gut microbial metabolism differ between CD patients, UC patients, and controls. The emerging technique of urinary metabolic profiling with multivariate analysis was able to distinguish these cohorts.

Formate excretion in urine of rats fed dimethylaminoazobenzene-rich diets: the possibility of formate formation from D-lactate.

This experiment was carried out to evaluate the possibility of degradation of d-lactate into formate and acetaldehyde. In order to induce hyperproduction of d-lactate in rats. Donryu male albino rats were fed diets containing 0.064% 3'-methyl-4-dimethylaminoazobenzene (3'-MDAB), 4'-methyl-4-dimethylaminoazobenzene (4'-MDAB) or 2-methyl-4-dimethylaminoazobenzene (2-MDAB) for 10 weeks. During the experiment, body mass, food and water intake and volume of urine were documented. Methylglyoxal, D-lactate and formate in the urine samples were determined. On the first day of the eleventh week, methylglyoxal, D-lactate, glutathione and enzymatic activities of demethylation and glyoxalase I and II in liver were measured. Methylglyoxal, D-lactate and clinical chemistry parameters of blood plasma were also measured. The levels of methylglyoxal and D-lactate in livers of rats fed 3'-MDAB were very high, while those of 2-MDAB fed-rats and the control group were the same. The fact that glyoxalase I activity and the level of glutathione, a cofactor of glyoxalase I, were high in the livers of the 3'-MDAB-fed rats can explain the elevated levels of methylglyoxal and D-lactate in the liver. The most striking results were the elevated formate levels in the urine of rats fed 3'- and 4'-MDAB in a precancerous state. The degradation of D-lactate, an end product of the methylglyoxal bypass, into acetaldehyde and formate was suggested as a possible way to explain the results.

'Real-world' compensatory behaviour with low nicotine concentration e-liquid: subjective effects and nicotine, acrolein and formaldehyde exposure.

AIMS: To compare the effects of (i) high versus low nicotine concentration e-liquid, (ii) fixed versus adjustable power and (iii) the interaction between the two on: (a) vaping behaviour, (b) subjective effects, (c) nicotine intake and (d) exposure to acrolein and formaldehyde in e-cigarette users vaping in their everyday setting. DESIGN: Counterbalanced, repeated measures with four conditions: (i) low nicotine (6 mg/ml)/fixed power; (ii) low nicotine/adjustable power; (iii) high nicotine (18 mg/ml)/fixed power; and (iv) high nicotine/adjustable power. SETTING: London and the South East, England. PARTICIPANTS: Twenty experienced e-cigarette users (recruited between September 2016 and February 2017) vaped ad libitum using an eVic Supreme™ with a 'Nautilus Aspire' tank over 4 weeks (1 week per condition). MEASUREMENTS: Puffing patterns [daily puff number (PN), puff duration (PD), interpuff interval (IPI)], ml of e-liquid consumed, changes to power (where permitted) and subjective effects (urge to vape, nicotine withdrawal symptoms) were measured in each condition. Nicotine intake was measured via salivary cotinine. 3-Hydroxypropylmercapturic acid (3-HPMA), a metabolite of the toxicant acrolein, and formate, a metabolite of the carcinogen formaldehyde, were measured in urine. FINDINGS: There was a significant nicotine concentration × power interaction for PD (P < 0.01). PD was longer with low nicotine/fixed power compared with (i) high nicotine/fixed power (P < 0.001) and (ii) low nicotine/adjustable power (P < 0.01). PN and liquid consumed were higher in the low versus high nicotine condition (main effect of nicotine, P < 0.05). Urge to vape and withdrawal symptoms were lower, and nicotine intake was higher, in the high nicotine condition (main effects of nicotine: P < 0.01). While acrolein levels did not differ, there was a significant nicotine × power interaction for formaldehyde (P < 0.05). CONCLUSIONS: Use of a lower nicotine concentration e-liquid may be associated with compensatory behaviour (e.g. higher number and duration of puffs) and increases in negative affect, urge to vape and formaldehyde exposure.

Determination of thiazolidine-4-carboxylates in urine by chloroformate derivatization and gas chromatography-electron impact mass spectrometry.

The derivatization method of thiazolidine-4-carboxylic acid (TZCA) and methyl-thiazolidine-4-carboxylic acid (Me-TZCA) in urine with alcohol/chloroformate was achieved. TZCA and Me-TZCA were derivatized in one step in urine with ethyl chloroformate in 1 min at room temperature. The derivatives of TZCA and Me-TZCA had very good chromatographic properties and offered very sensitive response for gas chromatography-electron impact ionization-mass spectrometry (GC-EI-MS). On the basis of derivatization, the method for simultaneous determination of TZCA and Me-TZCA in human urine was developed. Deuterated Me-TZCA (Me-TZCA-d(4)) was synthesized as the internal standard (IS) for the analysis of urine samples. TZCA and Me-TZCA were derivatized and extracted from urine at pH 9.5 with toluene, and then the dried extract was dissolved with 100 microl ethyl acetate and injected in GC/MS system. The recoveries of TZCA and Me-TZCA were about 102 and 103%, respectively, at the concentration of 0.05 mg/l. The method detection limits (MDL) were 1.0 and 0.5 microg/l, respectively, for TZCA and Me-TZCA in 1 ml human urine. The coefficients of variation of TZCA and Me-TZCA were less than 6% at the concentrations of 0.05 and 0.2 mg/l, respectively. To assess the formation of TZCA during inhalation with formaldehyde (FA) (about 3.1 and 38.1 ppm FA in air), urine samples from rats were taken during 3 days after initiation of treatment. The mean amount of TZCA determined was 0.07 mg/l in control group and 0.18 mg/l during treatment with 3.1 ppm. The TZCA levels increased up to about 1.01 mg/l during treatment with 38.1 ppm. It is planned to study whether urinary TZCA can be used as an indicator in the biological monitoring of exposure to FA.

Extremely slow formate elimination in severe methanol poisoning: a fatal case report.

UNLABELLED: Methanol poisoning is a potentially fatal medical emergency because of its metabolism to formic acid. The half-life of formate has been reported in the range of 2.5-12.5 hours, but the degree of inter-individual variation is not known. We studied methanol and formate kinetics in a case of late diagnosed methanol poisoning with persisting metabolic acidosis and circulatory failure. CASE REPORT: A 63-year-old man was referred to our hospital with a tentative diagnosis of stroke. He was awake on admission, but he soon deteriorated in the emergency department and a metabolic acidosis was revealed. Methanol poisoning was then suspected approximately five hours after admission but in spite of intensive treatment he died after six days. RESULTS: The S-methanol half-lives during treatment with fomepizole before and during hemodialysis were 49.5 and 4.1 hours, respectively, while the similar half-lives of S-formate were 77.0 and 2.9 hours. S-fomepizole was measured and found to be within the therapeutic range during treatment. DISCUSSION: The patient was treated with the established dosing regimen for fomepizole and the measured S-fomepizole levels throughout the treatment were adequate; the S-methanol elimination also suggests that methanol metabolism was blocked. Hence, other explanations for this exceptionally long formate half-life include slow formate metabolism, due to small hepatic folate stores or to genetic deficiencies in formate-metabolizing enzymes, or slow formate excretion, due to renal tubular acidosis, to a non-oliguric renal failure, or to genetic deficiencies in the renal formate transporters. CONCLUSION: This case report indicates that the half-life of S-formate may have greater inter-individual variation than earlier expected, being by far the longest half-life reported in the medical literature. These results support the use of hemodialysis in the treatment of such patients.

Trichloroethylene-induced formic aciduria in the male C57 Bl/6 mouse.

1, 1, 2-Trichloroethylene (TCE) is of environmental concern, due to evaporation while handling, chemical processing and leakage from chemical waste sites, leading to its contamination of ground water and air. For several decades there has been issues about possible long term health effects of TCE but recently the International Agency for Research on Cancer (IARC) and the US Environmental Protection Agency classified TCE as a human carcinogen. Links having been established between occupational exposures and kidney cancer and possible links to non-Hodgkin lymphoma and liver cancer, but there is more still more to learn. In male rats, TCE produces a small increase in the incidence of renal tubule tumours but not in female rats or mice of either sex. However, chronic renal injury was seen in these bioassays in both sexes of rats and mice. The mechanism of kidney injury from TCE is thought to be due to reductive metabolism forming a cysteine conjugate that is converted to a reactive metabolite via the enzyme cysteine conjugate ß-lyase. However, TCE also produces a marked and sustained formic aciduria in male rats and it has been suggested that long term exposure to formic acid could lead to renal tubule injury and regeneration. In this study we have determined if TCE produces formic aciduria in male mice following a single and repeat dosing. Male C57 Bl/6OlaHsd mice were dosed with 1000mg/kg by ip injection and urine collected overnight 24, 48, 72 and 96h after dosing. Formic acid was present in urine 24h after dosing, peaked around 48h at 8mg formic acid excreted/mouse, and remained constant over the next 24h and was not back to normal 96h after dosing. This was associated with a marked acidification of the urine. Plasma creatinine and renal pathology was normal. Plasma kinetics of formic acid showed it was readily cleared with an initial half-life of 2.42h followed by a slower rate with a half-life of 239h. Male mice were then dosed twice/week at 1000mg/kg TCE for 56days, as anticipated there was a marked and sustained formic aciduria over the duration of the study. This was associated with acidification of the urine, mild diuresis and a marked fall in urinary ammonia. Six biomarkers of renal injury KIM-1, NGAL, NAG, Cystatin-c, Albumin and IL-18 were measured in urine over time and they all showed a small increase at the later time points indicative of early markers of renal injury. However, there was no histological evidence of renal damage or renal tubule cell proliferation after 8 weeks' exposure to TCE. The concentration of formic acid in plasma at the end of the study was 1.05±0.61mM compared to control, 0.39±0.17mM. In the liver, formic acid was present at a concentration of 1mM in both control and treated mice while in the kidney it was higher at 2mM in both treated and controls. We also report that trichloroacetic acid (TCA) a metabolite of TCE also causes formic aciduria, at doses likely to arise in vivo after 1000mg/kg TCE namely 16 and 32mg/kg. Urinary formic acid peaked 24h after dosing at 4mg formic acid excreted/mouse. Thus, as in male and female rats (Yaqoob et al., 2013) male mice show a marked formic aciduria following TCE which after 8 weeks' exposure did not produce renal injury, but the small rise in renal biomarkers suggest renal damage may occur following longer exposure. Thus, TCE-induced formic aciduria may be a contributor factor in the chronic renal injury seen in male and female rats and mice.

Metabolomic profiles investigation on athletes' urine 35 minutes after an 800-meter race.

BACKGROUND: The aim of this study was to identify possible biological pathways of the metabolite profile changes in athletes' urine samples before and after 800-m runs. METHODS: We used an NMR-based metabolomics analysis to evaluate the metabolite profile changes in 19 young male athletes' urine samples after 800-m runs and provide an overall picture of its impact. Various multivariate data analysis methods, including principal component analysis (PCA), partial least squares-discrimination analysis (PLS-DA), and orthogonal projection of latent-structure-discrimination analysis (OPLS-DA) were applied to analyze the NMR data and thus identify possible correlations between the metabolite profile changes and the alterations in biological pathways. RESULTS: The potential biological mechanism of an 800-m race was finally elucidated based on the multivariate statistical analysis results. The levels of blood lactate (Lac), 2-hydroxyisovalerate (2HIV), leucine, 2-hydroxyisobutyrate (2HIB), alanine, N-acetyl-glucoprotein, pyruvate, creatinine, fumarate, inosine (Ino) and hypoxanthine (Hyx) were up-regulated in the post samples, whereas the levels of certain metabolites, including 3-hydroxyisovalerate, citrate, taurine, glycine and formate were down-regulated in the postsamples. CONCLUSIONS: Our study provides novel insights into the 800-m race metabolic characteristic. Separation of pre- from postexercise samples was related to the Krebs cycle, Cori cycle, Cahill cycle, HIFs and ROS. Besides the Lac change, the increased concentrations of Ino, 2HIV concentrations in the postexercise urine samples represent potential indices which indicate the high percent of glycolysis during the 800-m run. The increase of concentrations of Hyx, 2HIB may indicated oxidative stress with concomitant ROS generation in the athletes' bodies during the 800-m race.

Environmental and biological monitoring of occupational formaldehyde exposure resulting from the use of products for hair straightening.

The evaluation of formaldehyde (FD) exposure in beauty salons, due to the use of hair straightening products, and its relation with genotoxicity biomarkers was performed in this study. Regardless of official recommendations, the inappropriate use of homemade hair creams has became a popular practice in Brazil, and high formaldehyde content in the "progressive straightening" creams can contain mutagens that could increase the incidence of neoplasia in those people who use them. Damage to DNA was assessed by conducting a micronuclei test (MNT) on buccal cells and the comet assay on heparinized venous blood samples. A total of 50 volunteers were recruited at six different beauty salons (labeled A to F). At two salons that used products that did not contain FD (salons D and E), environmental FD concentrations were 0.04 and 0.02 ppm. In contrast, the products used at salons A, B, C, and F contained 5.7, 2.61, 5.9, and 5.79% of FD, and these salons had environmental FD concentrations of 0.07, 0.14, 0.16, and 0.14 ppm, respectively. Comparison of the beauty salon workers from each of the six beauty salons revealed significant differences in urinary formic acid (FA) concentration before exposure (p = 0.016), urinary FA after exposure (p = 0.004), variation in FA concentration before and after exposure (p = 0.018), environmental FD concentration (p < 0.001), cytogenetic damage detected by the comet assay according to both damage index (p < 0.001) and frequency of damage (p < 0.001), and for karyorrhexis only according to the MNT (p = 0.001).