Farmworker and nonfarmworker Latino immigrant men in North Carolina have high levels of specific pesticide urinary metabolites.

This article compares detections and concentrations of specific organophosphate (OP), bis-dithiocarbamate, and pyrethroid pesticide urinary metabolites among Latino male farmworkers and nonfarmworkers in North Carolina. Data are from interviews and urine samples collected in 2012 and 2013. Farmworkers and nonfarmworkers frequently had detections for OP and pyrethroid pesticide urinary metabolites. Detection of bis-dithiocarbamate urinary metabolites was less frequent, but substantial among the nonfarmworkers. The concentrations of organophosphate, bis-dithiocarbamate, and pyrethroid pesticide urinary metabolites were high for farmworkers and nonfarmworkers compared to National Health and Nutrition Examination Survey results. Pesticide urinary metabolite detection was not associated with occupation in nonfarmworkers. Research for reducing pesticide exposure among farmworkers remains important; research is also needed to determine pesticide exposure pathways among Latino nonfarmworkers.

Protection of humans by plant glucosinolates: efficiency of conversion of glucosinolates to isothiocyanates by the gastrointestinal microflora.

Plant-based diets rich in crucifers are effective in preventing cancer and other chronic diseases. Crucifers contain very high concentrations of glucosinolates (GS; ß-thioglucoside-N-hydroxysulfates). Although not themselves protective, GS are converted by coexisting myrosinases to bitter isothiocyanates (ITC) which defend plants against predators. Coincidentally, ITC also induce mammalian genes that regulate defenses against oxidative stress, inflammation, and DNA-damaging electrophiles. Consequently, the efficiency of conversion of GS to ITC may be critical in controlling the health-promoting benefits of crucifers. If myrosinase is heat-inactivated by cooking, the gastrointestinal microflora converts GS to ITC, a process abolished by enteric antibiotics and bowel cleansing. When single oral doses of GS were administered as broccoli sprout extracts (BSE) to two dissimilar populations (rural Han Chinese and racially mixed Baltimoreans) patterns of excretions of urinary dithiocarbamates (DTC) were very similar. Individual conversions in both populations varied enormously, from about 1% to more than 40% of dose. In contrast, administration of ITC (largely sulforaphane)-containing BSE resulted in uniformly high (70%-90%) conversions to urinary DTC. Despite the remarkably large range of conversion efficiencies between individuals, repeated determinations within individuals were much more consistent. The rates of urinary excretion (slow or fast) were unrelated to the ultimate magnitudes (low or high) of these conversions. Although no demographic factors affecting conversion efficiency have been identified, there are clearly diurnal variations: conversion of GS to DTC was greater during the day, but conversion of ITC to DTC was more efficient at night.

The principal urinary metabolite of allyl isothiocyanate, N-acetyl-S-(N-allylthiocarbamoyl)cysteine, inhibits the growth and muscle invasion of bladder cancer.

Naturally occurring allyl isothiocyanate (AITC) was recently shown to be selectively delivered to bladder cancer tissue via urinary excretion and to inhibit bladder cancer growth and muscle invasion in an animal model. AITC is excreted in urine mainly as N-acetyl-S-(N-allylthiocarbamoyl)cysteine, more commonly known as the N-acetylcysteine conjugate (NAC-AITC). We show here that treatment of human bladder cancer UM-UC-3 cells or rat bladder cancer AY-27 cells with NAC-AITC at 15 µM results in significant inhibition of cell growth and proliferation, together with cell cycle arrest and apoptosis. We also show that NAC-AITC administered orally at 10 µmol/kg body wt inhibits cancer growth by 40% and muscle invasion by 49% in an orthotopic rat bladder cancer model. Furthermore, the anticancer activity of NAC-AITC is associated with the modulation of several important molecular targets, including downregulation of both α-tubulin and ß-tubulin, activation of caspase-3 and downregulation of vascular endothelial growth factor. These results are similar to those shown previously for AITC and are consistent with the understanding that NAC-AITC is a carrier of AITC. Furthermore, comparison of the pharmacokinetic and physical properties of NAC-AITC with those of AITC suggests that NAC-AITC is superior to AITC for potential use for prevention and therapy of bladder cancer.

High-throughput sample preparation for the quantitation of acephate, methamidophos, omethoate, dimethoate, ethylenethiourea, and propylenethiourea in human urine using 96-well-plate automated extraction and high-performance liquid chromatography-tandem mass spectrometry.

Acephate, methamidophos, o-methoate, and dimethoate are organophosphorus pesticides, and ethylenethiouria and propylenethiourea are two metabolites from the bisdithiocarbamate fungicide family. They are some of the most widely used pesticides and fungicides in agriculture both domestically and abroad. The existing high-performance liquid chromatography (HPLC)-tandem mass spectrometry (MS/MS) method for the measurement of these compounds in human urine was improved by using a 96-well plate format sample preparation; the use of HPLC-MS/MS was comparable with a concentration range of 0.125 to 50 ng/ml. Deuterium-labeled acephate, ethylenethiouria, and methamidophos were used as internal standards. The sample preparation procedure, in the 96-well format with a 0.8-ml urine sample size, uses lyophilization of samples, followed by extraction with dichloromethane. The analytes were chromatographed on a Zorbax SB-C3 (4.6 × 150 mm, 5.0-µm) column with gradient elution by using 0.1% formic acid in aqueous solution (solvent A) and 0.1% formic acid in methanol (solvent B) mobile phase at a flow rate of 1 ml/min. Quantitative analysis was performed by atmospheric pressure chemical ionization source in positive ion mode using multiple-reaction monitoring of the precursor-to-product ion pairs for the analytes on a TSQ Quantum Ultra HPLC-MS/MS. Repeated analyses of urine samples spiked with high (15 ng/ml), medium (5 ng/ml), and low (1 ng/ml) concentrations of the analytes gave relative SDs of <13%. The limits of detection were in the range of 0.004-0.01 ng/ml. The method also has high accuracy, high precision, and excellent extraction recovery. Furthermore, the improved sample preparation method decreased the cost and labor required while effectively doubling the analytic throughput with minimal matrix effect.

Cruciferous vegetable intake questionnaire improves cruciferous vegetable intake estimates.

OBJECTIVE: To develop a validated, focused Cruciferous Vegetable Food Frequency Questionnaire (FFQ) as an assessment tool for specific quantification of dietary cruciferous vegetable exposure. DESIGN/METHODS: Participants (n=107; 18 to 76 years old) completed a standard FFQ and the Cruciferous Vegetable FFQ twice over a 2-week period. Repeat dietary recalls were collected on 3 days over the same 2-week period. Urinary dithiocarbamate was determined as a biomarker of cruciferous vegetable intake. STATISTICAL ANALYSES: Descriptive statistics of intake; paired t tests and sign tests for comparison of intake estimates between instruments; Spearman correlations to assess reliability and associations between diet instruments and urinary dithiocarbamate. RESULTS: Cruciferous vegetable intake was significantly correlated between the two FFQs (r(s)=0.58), although the Cruciferous Vegetable FFQ estimated intake 35 g higher than the standard FFQ. The Cruciferous Vegetable FFQ was reliable, with a repeated measures correlation of 0.69 (P

Characterizing the influence of structure and route of exposure on the disposition of dithiocarbamates using toluene-3,4-dithiol analysis of blood and urinary carbon disulfide metabolites.

Differences in the toxicities observed for dithiocarbamates have been proposed to result from the influence of nitrogen substitution, oxidation state, and route of exposure. To better characterize the fate of dithiocarbmates in vivoas a function of structure and route of exposure, rats were administered equimolar doses of carbon disulfide (CS2), N-methyldithiocarbamate, pyrrolidine dithiocarbamate, N,N-diethyldithiocarbamate, or disulfiram daily for five days, either po or ip, and sequential blood samples obtained. Protein dithiocarbamates formed by the in vivo release of CS2, parent dithiocarbamate, and protein-bound mixed disulfides were assessed in plasma and hemolysate by measuring toluene trithiocarbonate generated upon treatment with toluene-3, 4-dithiol (TdT). To aid in determining the bioavailability of CS2 from the administered dithiocarbamates, the urinary CS2 metabolites, 2-thiothiazolidine-4-carboxylic acid (TTCA) and 2-thiothiazolidin-4-ylcarbonylglycine (TTCG), were also determined. The levels of TdT-reactive moieties detected depended upon both the compound administered and the route of exposure. Parent dithiocarbamates, with the exception of disulfiram, were eliminated from blood within 24 h; but protein associated TdT-reactive moieties persisted and accumulated with repeated exposure, regardless of the route of exposure. N-Methyldithiocarbamate demonstrated the greatest potential to produce intracellular globin modifications, presumably through its unique ability to generate a methylisothiocyanate metabolite. Urinary excretion of TTCA and TTCG was more sensitive than TdT analysis for detecting dithiocarbamate exposure, but TdT analysis appeared to be a better indicator of in vivo release of CS2 by dithiocarbamates than were urinary CS2 metabolites. These data suggest that CS2 is a more important metabolite, following oral exposure, than are other routes of exposure, e.g., inhalation or dermal. In addition, data also suggest that acid stability, nitrogen substitution, and route of exposure are important factors governing the toxicity observed for a particular dithiocarbamate.

Involvement of active transport systems in the mobilization of cadmium by dithiocarbamates in vivo.

Previously, we reported that the action of cadmium (Cd) complexing dithiocarbamates, such as N-benzyl-D-glucamine dithiocarbamate (BGD) and N-p-hydroxymethylbenzyl-D-glucamine dithiocarbamate (HBGD), in removing Cd from the kidney involves a probenecid-sensitive organic anion transport system. However, other mechanisms responsible for Cd mobilizing effects of BGD and HBGD are still unclear. Therefore, in the present study we examined the effects of phloretin (an inhibitor of plasma membrane glucose carrier), phloridzin (an inhibitor of Na(+)-dependent active hexose transport) and alpha-aminoisobutyric acid (AIB, an inhibitor of amino acid transport) on the excretion and distribution of the chelating agents and Cd in mice. Phloretin pretreatment markedly decreased the biliary and urinary excretions of BGD and HBGD. Phloridzin pretreatment also decreased the biliary and urinary excretions of HBGD, but had no effect on the BGD. AIB pretreatment had no effect on the excretions of either BGD or HBGD. Phloretin pretreatment increased the hepatic and renal contents of BGD and HBGD. Contrary to this, phloridzin pretreatment decreased the hepatic content of BGD and hepatic and renal contents of HBGD, while AIB pretreatment decreased the renal contents of BGD and HBGD. The mobilizing effects of BGD and HBGD on the hepatic and renal Cd was also investigated using Cd-exposed mice. Phloretin or phloridzin pretreatment decreased the mobilizing effect of BGD and HBGD on the hepatic Cd, but had no effect on the renal Cd. These results suggest that BGD and HBGD are taken up into the liver and kidney by phloridzin- and phloretin-sensitive transport system, respectively; that Cd-BGD and Cd-HBGD complexes formed in the hepatic cells are secreted to the bile by phloridzin- and phloretin-sensitive transport systems; and that free BGD and HBGD secreted from these organ to the bile and urine might have occurred, at least in part, by different mechanisms.

Quantitative determination of dithiocarbamates in human plasma, serum, erythrocytes and urine: pharmacokinetics of broccoli sprout isothiocyanates in humans.

BACKGROUND: Humans are exposed to substantial quantities of isothiocyanates and glucosinolates from vegetables. Since dietary isothiocyanates are widely regarded as potentially important chemoprotectors against cancer, reliable methods for measuring the plasma and tissue pharmacokinetics of isothiocyanates and their dithiocarbamate metabolites are essential for defining dosing regimens. METHODS: Isothiocyanates (ITC) and dithiocarbamates (DTC) react quantitatively with 1,2-benzenedithiol to produce 1,3-benzodithiole-2-thione that can be quantified spectroscopically. Although this cyclocondensation reaction has been highly useful for analyzing plant material and urine samples, the determination of DTC/ITC (the total quantity of DTC and ITC components in a sample that react in the cyclocondensation reaction) in blood and tissues has been hampered by their low levels and the high concentrations of proteins that interfere with the cyclocondensation reaction. The protein content of blood and tissues was reduced by the precipitation with polyethylene glycol (PEG) or ultrafiltration, and the sensitivity of the method was increased substantially by the solid phase extraction of the cyclocondensation product. RESULTS: Pharmacokinetic measurements were made in four human volunteers who received single doses of about 200 micromol of broccoli sprout isothiocyanates (largely sulforaphane, with lesser amounts of iberin and erucin). Isothiocyanates were absorbed rapidly, reached peak concentrations of 0.943-2.27 micromol/l in plasma, serum and erythrocytes at 1 h after feeding and declined with first-order kinetics (half-life of 1.77+/-0.13 h). The cumulative excretion at 8 h was 58.3+/-2.8% of the dose. Clearance was 369+/-53 ml/min, indicating active renal tubular secretion. CONCLUSION: A sensitive and specific method for quantifying DTC levels in human plasma, serum, and erythrocytes has been devised. Determinations of ITC/DTC levels are important because: (i) dietary isothiocyanates are of potential value in reducing the risk of cancer, and (ii) humans are extensively exposed to DTC as fungicides, insecticides, pesticides and rubber vulcanization accelerators.

Urinary excretion of dithiocarbamates and self-reported Cruciferous vegetable intake: application of the 'method of triads' to a food-specific biomarker.

OBJECTIVE: Greater intake of Cruciferous vegetables (e.g. broccoli) may prevent cancer at several sites. Urinary excretion of isothiocyanate conjugates (dithiocarbamates, DTC) provides a specific biomarker of Cruciferous vegetable consumption suitable for epidemiological investigations. However, no gold-standard referent is available for evaluating urinary DTC levels as an estimator of Cruciferous vegetable consumption. We compared urinary DTC levels to intake as measured by two self-reported dietary assessment techniques. DESIGN: Cruciferous vegetable consumption was measured before and after a behavioural dietary intervention using multiple 24-hour recalls (24HR), a food-counting questionnaire (VFQ) and urinary DTC excretion levels. Analysis included a structural equation approach (Method of Triads) combining these three assessment techniques to estimate the relationship between DTC level and the study population's 'true' Cruciferous vegetable intake. SETTING: The intervention curriculum assisted participants in consuming about 2 servings per day for a 6-week period. Participants attended four classes emphasising problem-solving skills, dietary counselling and vegetable preparation skills. There were no dietary restrictions. SUBJECTS: Thirty-three healthy, free-living, post-menopausal women. RESULTS: Although few participants reported Cruciferae consumption prior to the intervention, 30 participants reported Cruciferae consumption after the intervention (Post-intervention). Urinary DTC levels were correlated with estimated intake derived from either the 24HR (r = 0.57; 95% confidence interval (95% CI) 0.28, 0.76) or VFQ (r =0.49; 95% CI 0.17, 0.71). The validity coefficient (Method of Triads) between urinary DTC excretion and an index of true Cruciferous intake was stronger than the Pearson correlation (r(v) = 0.65; 95% CI 0.35, 0.90), and comparable to estimates derived from the 24HR (r(v) = 0.82; 95% CI 0.65, 1.00) or VFQ (r(v) = 0.76; 95% CI 0.47, 0.92) method. These associations were not affected by adjustment for body mass index, energy intake, or social approval or desirability response sets. CONCLUSIONS: Food-frequency questionnaires (FFQ) suitable for large epidemiological studies may not be designed to measure all Cruciferae, and cannot capture exposure to phytochemicals derived from those vegetables. Urinary DTC measurement was significantly correlated with Cruciferae intake derived from two dietary assessment approaches, and urinary DTC levels could supplement traditional FFQ data by providing an index of recent Cruciferous vegetable intake not susceptible to reporting biases.

The measurement of 2-thiothiazolidine-4-carboxylic acid as an index of the in vivo release of CS2 by dithiocarbamates.

Dithiocarbamates and their disulfides are used extensively as agricultural fungicides, as accelerators of the vulcanization process of rubber in industry, and as therapeutic agents in medicine. The widespread uses of these compounds in agriculture, industry, and medicine provide many avenues of exposure to the human population. Subchronic to chronic exposures to some dithiocarbamates have resulted in the development of neuropathy in humans and experimental animals. Decomposition to CS2 presents a potential mechanism through which the toxicity of dithiocarbamates may be mediated. The purpose of this study was to determine the potential of dithiocarbamates to release CS2 in vivo. The ability to release CS2 was assessed by measuring urinary 2-thiothiazolidine-4-carboxylic acid (TTCA), which is used in industry to measure the exposure of workers to CS2. In this study, rats were housed individually in metabolic cages and given daily equimolar ip or po doses (1.5 mmol/kg) of N,N-diethyldithio-carbamate (DEDC), disulfiram (DS), N-methyldithiocarbamate (NMDC), or CS2 for 5 days, and TTCA was measured in urine collected at 24 h intervals. For each compound administered, TTCA was produced in all of the treated animals and the amount of TTCA eliminated in urine from po administration was significantly greater than that from ip administration. The relative rates of TTCA elimination in urine were DS > DEDC approximately equal to CS2 > NMDC for both routes of administration. Following administration of N,N-diethyl[13C = S] dithiocarbamate, carbon-13 enrichment at the thiocarbonyl carbon of TTCA was demonstrated using 13C NMR. Analysis of urinary TTCA proved to be useful both for establishing the in vivo release of CS2 by dithiocarbamate containing compounds and for evaluating the bioavailability of CS2. The results appear especially relevant to disulfiram, which is given orally for sustained periods in the treatment of alcoholism and has resulted in the development of neuropathy in susceptible individuals.

Identification and quantification of the N-acetylcysteine conjugate of allyl isothiocyanate in human urine after ingestion of mustard.

Allyl isothiocyanate (AITC) is a constituent of cruciferous vegetables. It occurs widely in the human diet as a natural ingredient or food additive. AITC possesses numerous biochemical and physiological activities. It is cytotoxic and tumorigenic at high doses and also is a modulator of enzymes involved in metabolism of xenobiotics, including carcinogens. It is plausible that the wide consumption of dietary AITC may have profound effects on human health. To facilitate investigations of the effects of dietary AITC in humans, a method of measuring its uptake is needed. In this study, a urinary marker was developed for quantifying AITC uptake in humans. Four adult volunteers were asked to eat a meal containing brown mustard as the source of AITC. The 48-h urine samples were collected from these individuals and analyzed by reverse phase high performance liquid chromatography. A major urinary metabolite was found, which was identified as N-acetyl-S-(N-allylthiocarbamoyl)-L-cysteine, the N-acetylcysteine conjugate of AITC, by comparing its retention time and UV, nuclear magnetic resonance, and mass spectra with those of the synthetic standard. After ingestion of mustard, the AITC conjugate was detected in urine collected from 0 to 12 h. No conjugate was found in urine samples collected after 12 h. The major portion of this metabolite was excreted within 8 h. The average total excretion of AITC conjugate was 5.4 +/- 1.7 (SD) mg after consumption of 10 g of mustard and 12.8 +/- 2.0 mg when 20 g of mustard was consumed. Thus, a dose-dependent excretion of this metabolite was demonstrated.(ABSTRACT TRUNCATED AT 250 WORDS)

Metabolism of the thiocarbamate herbicide SUTAN in rats.

1. The metabolism of the thiocarbamate herbicide SUTAN (butylate) was studied after administration of single oral doses of [isobutyl-1-14C]SUTAN to male and female rats. 2. The radiolabelled dose was rapidly absorbed and excreted, with 79% of the dose excreted in the urine in 72 h. The small percentages of radioactivity excreted in the faeces and as 14CO2 were significantly higher (P less than or equal to 0.05) in males than in females. 3. SUTAN was extensively metabolized, and no unmetabolized SUTAN was found in the urine. A total of 18 of the 29 urinary metabolites were identified, and identified metabolites represented 83-88% of the urinary radioactivity. 4. Diisobutylamine was the major urinary metabolite in both males and females, averaging 51% of the urinary radioactivity. 5. Other significant urinary metabolites included primary hydroxylated and tertiary hydroxylated diisobutylamines and a series of mercapturic acid pathway metabolites, including an S-glucuronide and several hydroxylated and unhydroxylated mercapturates. 6. Oxidations at the three alkyl groups produced a variety of minor urinary metabolites, and hydroxylation of the primary or tertiary carbon on the isobutyl groups, followed by an intramolecular reaction, generated a series of minor cyclized metabolites.

Determination of N-acetyl-S-(N-alkylthiocarbamoyl)-L-cysteine, a principal metabolite of alkyl isothiocyanates, in rat urine.

A simple and rapid analytical procedure is described for N-acetyl-S-(N-alkylthiocarbamoyl)-L-cysteine (alkyl = benzyl, allyl, methyl, ethyl or n-butyl), a mercapturic acid with an unstable dithiocarbamic acid ester structure, which is found in rat urine as the principal metabolite of the corresponding alkyl isothiocyanate. Because such mercapturic acids decompose at pH values greater than 5 to N-acetylcysteine and alkyl isothiocyanate, the free isothiocyanate is converted with n-butylamine into the corresponding disubstituted thiourea, and, after extraction, measured by high-performance liquid chromatography using an ultraviolet detector. The recovery is ca. 100% and the precision is very good. The lower limit of detection is ca. 0.5 microgram of thiourea. The 24-h renal excretion of these mercapturic acids was determined in rats after administration of benzyl, allyl, methyl, ethyl or n-butyl isothiocyanate.

Metabolism of some naturally occurring isothiocyanates in the rat.

The metabolism of methyl, ethyl, butyl and allyl isothiocyanate, which occur as glucosinolates in a number of plants, was studied. Oral administration of the substances to the rat was followed by their renal excretion as mercapturic acids, which were isolated as dicyclohexylamine salts. Chemical structure was determined by synthesis and 1H-n.m.r. spectra. The mercapturic acids were very labile dithiocarbamidic acid esters, formed by the addition of the isothiocyanate group to the SH group of the cysteine component.

[Urinary excretion of diethyldithiocarbamate (DDC) after antabuse administration to workers chronically exposed to carbon disulfide]. / Wydalanie dwuetylodwutiokarbaminianów (DDC) z moczem po podaniu antabusu u pracowników przewlekle narazonych na dwusiarczek wegla.

Djuricz's antabus test has been applied in a group chronically exposed to carbon disulphide. Sixty eight viscose industry workers hospitalized in the Clinic of Occupational Diseases, Institute of Occupational Medicine, Lódz, have been examined along with controls. In all subjects, after oral administration of antabus, the content of diethyldithiocarbamates (DDC) was determined in a 5 hr urine sample. The mean values of excreted DDC have indicated a statistically significant decrease in those chronically CS2 exposed, as compared to the controls. The dependence of DDC excretion on the duration of work in exposure has indicated a clear decrease in the values in those chronically exposed as compared to those exposed for a short time. The obtained results confirm the observations of other authors on DDC excretion decrease in CS2 exposed workers, demonstrating simultaneously some effects of chronic CS2 exposure on antabus metabolism.

Dose dependence of the excretion of maneb metabolites in urine of rats.

14C-activity in the urine of female rats appeared to be independent of dose when single oral doses of [14C]maneb, a fungicide, manganese ethylene-bis(dithiocarbamate), were given. This was found with [14C]maneb synthesized from [U-14C]ethylenediamine. In the dose range, 23 microgram/kg to 1.4 g/kg of maneb, the excreted activity in urine of 39 female rats was 48.8 +/- 12.6% (n=39).