High-throughput method for the analysis of ethylenethiourea with direct injection of hydrolysed urine using online on-column extraction liquid chromatography and triple quadrupole mass spectrometry

Abstract

Ethylenethiourea (ETU) is of major toxicological concern, since in experimental animal studies, ETU has shown a large spectrum of adverse effects. High occupational exposure can be found among agricultural workers or during manufacturing of ethylenbisdithiocarbamates (EBDC). For the general public, sources of environmental exposure may be residues of ETU in commercial products, food and beverages. For the determination of ETU in human urine we present a high-throughput online on-column extraction liquid chromatography triple quadrupole mass spectrometry method using direct injection of hydrolysed urine samples. This method is simple, user- and environmentally friendly and all sample preparation is performed in 96-well plates. A labelled ETU internal standard was used for quantification. The method showed a good sensitivity with a limit of quantification (LOQ) of 0.5ng ETU/mL urine and the calibration curve was linear in the range 0.25-200ng ETU/mL urine. The within-run, between-run and between-batch precision was between 6% and 13%. Alkaline hydrolysis considerably increased the levels of ETU indicating a potential conjugate. The method was applied in an experimental dermal exposure study in humans, with sample concentrations ranging from 0.4 to 5.0ng ETU/mL urine. The excretion in urine was 10% of the applied dose. The elimination profile seemed to differ between the two individuals. The results show an estimated half-life of ETU between 34 and 72h. Although the experiment is limited to two individuals, the data provide valuable and new information regarding the toxicokinetics of ETU after dermal exposure.

Aerial Application of Mancozeb and Urinary Ethylene Thiourea (ETU) Concentrations among Pregnant Women in Costa Rica: The Infants’ Environmental Health Study (ISA)

“…The agricultural work clothes the women most often washed came from family members who worked on banana plantations. In this activity, there was a stronger association of ETU with washing the day before urine sampling compared with washing on the day of urine sampling, which may reflect relatively slow uptake of ETU after dermal exposure ( Ekman et al 2013 ). In contrast, ETU was more strongly associated with reported aerial spraying on the day of sampling than aerial spraying on the previous day, possibly due to faster absorption of mancozeb and ETU through the lungs following respiratory exposure compared with absorption via dermal exposure ( Ekman et al 2013 ).…”

“…A small occupational exposure study ( n = 13) reported associations between dermal mancozeb exposure and urinary ETU concentrations ( Colosio et al 2002 ). A recent experimental dermal exposure study on ETU in humans reported that approximately 10% of the dose of ETU applied on skin was excreted in urine ( Ekman et al 2013 ).…”

“…Urine samples were collected in 100-mL beakers (Vacuette®, sterile), transferred to 15 mL tubes (PerformR™ Centrifuge tubes, Labcon®, sterile), and stored at –20°C until shipment (4°C) to Lund University, Sweden, for analysis. Samples were hydrolyzed in 0.09 M NaOH for 1 hr at 100°C and aliquots of 20 μL were analyzed using a triple quadrupole linear ion trap mass spectrometer (QTRAP 5500; AB Sciex, Foster City, CA, USA) coupled to a two dimensional liquid chromatography system (UFLCRX; Shimadzu Corporation, Kyoto, Japan) ( Ekman et al 2013 ). The analyses were performed in positive atmospheric pressure chemical ionization mode ( Ekman et al 2013 ).…”

“…Samples were hydrolyzed in 0.09 M NaOH for 1 hr at 100°C and aliquots of 20 μL were analyzed using a triple quadrupole linear ion trap mass spectrometer (QTRAP 5500; AB Sciex, Foster City, CA, USA) coupled to a two dimensional liquid chromatography system (UFLCRX; Shimadzu Corporation, Kyoto, Japan) ( Ekman et al 2013 ). The analyses were performed in positive atmospheric pressure chemical ionization mode ( Ekman et al 2013 ). Three urinary quality control samples with known concentrations were added to each analytical batch ( n = 28).…”

Prenatal Mancozeb Exposure, Excess Manganese, and Neurodevelopment at 1 Year of Age in the Infants’ Environmental Health (ISA) Study

“…Specimens were aliquoted and were stored at until their shipment to Lund University, Sweden. Samples were then analyzed for ETU using a two-dimensional liquid chromatography mass spectrometer (LC-MS/MS; UFLCRX; Shimadzu Corporation) with a triple quadrupole linear ion trap (QTRAP 5500; AB Sciex) ( Ekman et al. 2013 ).…”