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Capillary gas chromatographic (GC) methods using sulphur and mass selective detection for the qualitative and quantitative determination of tetrahydrophtalimide (THPI) and 2-thiothiazolidine-4-carboxylic acid (TTCA), urinary metabolites of the fungicide captan in rat and humans, were developed. Urinary detection limits were 2.7 μg/l for THPI and 110 μg/l for TTCA. Intraperitoneal and oral administration of captan to rats resulted in a 48-h cumulative urinary excretion of THPI of 1%-2% and 3%-9% of the dose, respectively. Cumulative urinary excretion of TTCA over 48 h ranged from 2% to 5% of the captan dose for the respective routes of administration. In urine of non-exposed human subjects, neither THPI nor TTCA could be detected. In urine of fruit-growers who were occupationally exposed to captan, both THPI and TTCA could be detected. Based on these results, THPI and TTCA are proposed as promising parameters for the biological monitoring of occupational exposure to captan. [ABSTRACT FROM AUTHOR]

Background: Patients with chronic obstructive pulmonary disease (COPD) have an independent increased risk of cardiovascular (CV) disease. Cardiovascular risk (CVR) assessment should be offered to all patients with COPD, according to the new Dutch CVR management (CVRM) guideline (May 2019)., Aim: To evaluate the impact of the new CVRM guideline on the care of patients with COPD in primary care., Design & Setting: A retrospective study took place within five primary healthcare centres located in The Netherlands., Method: In accordance with the guideline, the CVR of all patients with COPD was estimated and categorised. Data from 2014-2019 were used for the qualitative risk assessment based on comorbidities, and the quantitative Systematic Coronary Risk Evaluation (SCORE). In addition, the guideline-based follow-up was investigated., Results: Of the 391 patients with COPD, 84.1% ( n = 329) had complete data on CVR assessment: 90.3% ( n = 297) had a (very) high risk, and 9.7% ( n = 32) a low-to-moderate risk. Of the patients with (very) high risk, 73.4% ( n = 218) received guideline-based follow-up (primary care: 95.4%, secondary care: 4.6%). In 15.9% ( n = 62) of all patients with COPD, the CVR profile was not measured and of the (very) high-risk patients, 26.6% ( n = 79) were not enroled in a CV care programme., Conclusion: Whereas in the majority of patients with COPD the CVR is already known, for one out of six patients this CVR still has to be assessed according to the recently updated guideline. Moreover, once a (very) high risk has been assessed, as a consequence CV treatment of risk factors should be intensified in one out of four patients with COPD. Adherence to the new CVRM guideline could provide improvement in CVRM in more than a third of all patients with COPD., (Copyright © 2020, The Authors.)

The European Bioanalysis Forum dried blood spots/microsampling consortium is reporting back from the experiments they performed on further documenting the potential hurdles of the DBS technology. Their experiments focused on the impact of hematocrit changes, IS addition, spot homogeneity, aging of spots and stability of fresh blood and cards. Results from these experiments demonstrate that the issues of DBS in regulated bioanalysis are real and that the technology will need additional improvements to be ready for use as a general tool for regulated bioanalysis. In addition, results on fresh blood and card stability were shared at international meetings and will be reported at a later date.

Background: The European Bioanalysis Forum dried blood spots (DBS)/microsampling consortium is reporting back from the experiments they performed on further documenting the potential hurdles of the DBS technology. This paper is focused on the impact of hematocrit changes on DBS analyses., Results: The hematocrit can have an effect on the size of the blood spot, on spot homogeneity and on extraction recovery in a compound-dependent manner. The extraction recovery can change upon aging in an hematocrit-dependent way. Different card materials can give different outcomes., Conclusions: The results from the conducted experiments show that the issues of DBS in regulated bioanalysis are real and that the technology will need improvements to be ready for use as a general tool for regulated bioanalysis.

Background: At the start of their work, the European Bioanalysis Forum dried blood spots microsampling consortium did not form a dedicated team to investigate the spot homogeneity. However, two teams performed experiments that produced results relating to sample homogeneity., Results: The data, which were produced via two different approaches (a radiolabeled and a nonradiolabeled approach), are highly complementary and demonstrate clear effects on sample inhomogeneity due to the substrate type, compound and hematocrit levels., Conclusion: The results demonstrate that sample inhomogeneity is a significant hurdle to the use of dried blood spots for regulated bioanalysis that should be investigated further in the method establishment phase if the whole spot is not sampled.

Background: This study investigated the effect of injection of rocuronium or sugammadex alone and rocuronium + sugammadex on urea, creatinine, electrolyte levels, and histopathological findings in rats., Methods: Thirty-six Sprague-Dawley male rats were divided to receive intravenously 16 or 96 mg/kg sugammadex, 1 mg/kg rocuronium, 1 mg/kg rocuronium + 16 mg/kg sugammadex, or 1 mg/kg rocuronium + 96 mg/kg sugammadex. The control group received an equal volume of physiological serum. Rats receiving rocuronium were ventilated until resumption of spontaneous ventilation and followed for 72 h. Blood samples were withdrawn from the tail vein to measure urea, creatinine, and electrolyte values; then both kidneys were excised, and the tissues were used for histopathological examination., Results: Rats receiving rocuronium and high doses of sugammadex (96 mg/kg) showed increased glomerular vacuolation, tubular dilatation, vascular vacuolation and hypertrophy, lymphocyte infiltration, and tubular cell sloughing compared to the control group (p = 0.002). Biochemical markers of renal function were not significantly altered after treatment with high doses of sugammadex., Conclusion: The elimination half-life of the rocuronium-sugammadex complex was found to be greater than that of free rocuronium or sugammadex, which led to marginal histopathological changes in the kidney without affecting any renal functions.

Two simple and straightforward analytical procedures for the screening of sediment samples are reported. They involve extraction with ethyl acetate or methanol and subsequent analysis by means of gas chromatography-mass spectrometry (GC-MS) using large-volume injection (LVI) or solid-phase extraction (SPE). The latter, which was originally developed for the analysis of aqueous samples, can be used without any modification. In general, 10 ml of organic solvent were added to 2 g of sediment, and the mixture was shaken and allowed to stand overnight. The methanolic extracts were then diluted in water and subjected to preconcentration and analysis using on-line SPE-GC-MS. The ethyl acetate extracts were injected directly into the GC using LVI. Both methods were used for the detection and identification of microcontaminants during a monitoring study of the river Nitra (Slovak Republic). They included polyaromatic hydrocarbons (PAHs), chlorofluorohydrocarbons, alkoxylated and alkylated phenols and benzothiazole derivatives. Semi-quantitative profiles of the contaminants were constructed and provisionally interpreted. The results indicate that SPE-GC-MS, and also LVI-GC-MS, have good potential for a rapid screening of sediment samples and the identification of microcontaminants. The analytical procedures pose no problems, and the on-line set-up is user-friendly.

Chemical analysis for the characterisation of micro-organisms is rapidly evolving, after the recent advent of new ionisation methods in mass spectrometry (MS): electrospray (ES) and matrix-assisted laser desorption/ionisation (MALDI). These methods allow quick characterisation of micro-organisms, either directly or after minimum sample preparation. This review provides a brief introduction to ES and MALDI MS and a discussion of micro-organism characterisation capabilities. Some attention is devoted to the analysis of mixtures of proteins, lipids and other compounds, to the combination of polymerase chain reaction technology and MS, and to the analysis of whole bacteria and their lysates. The review of results produced hitherto is concluded with an outlook on future developments.

Cholera toxin, one of the toxins that may be generated by various strains of the bacterium Vibrio cholerae, can be considered as a substance possibly used in biological warfare. The possibilities of characterising the toxin by liquid chromatography electrospray mass spectrometry (LC-ES-MS) were investigated. The toxin can be detected by flow-injection (FIA) ES-MS of a dialysed solution and observation of the charge envelope signals of its A-unit and B-chain protein; sufficient information for identification by the molecular mass of either protein could be obtained for quantities in the order of 10 fmol. Confirmatory analysis was carried out by 2-mercaptoethanol reduction and FIA-ES-MS detection of the product proteins or by tryptic digest LC-ES-MS with ion chromatogram detection of most of the tryptic fragments of the A-unit and B-chain from the singly, doubly or triply charged ion signals. The confirmatory tryptic digest LC-ES-MS analysis could be achieved with quantities as low as 1 pmol. Possible biovariations in the toxin can mostly be determined by sequencing, where the amino acid composition of tryptic fragments of the A1-chain, T5 and T15, and of the B-chain, T1, T4 and T5, cover all known biovariations. Partial sequencing of cholera toxin, originating from a classical strain, O1/569B, was achieved by LC-ES-MS/MS of most tryptic fragments larger than three amino acid residues.

One of the major processes that occur as a result of radical-induced oxidative stress is lipid peroxidation (LPO). Degradation of lipid peroxides results in various products, including a variety of carbonyl compounds. In the present study eight different lipid degradation products, i.e., formaldehyde, acetaldehyde, acetone, propanal, butanal, pentanal, hexanal and malondialdehyde were identified and measured simultaneously and quantitatively in rat urine after derivatization with O-(2,3,4,5,6-pentafluorbenzyl)hydroxylamine hydrochloride, extraction with heptane and using gas chromatography-electron-capture detection (GC-ECD). The identity of the respective oximes in urine was confirmed by gas chromatography-negative ion chemical ionization mass spectrometry (GC-NCI-MS). Simultaneously measured standard curves were linear for all oxime-products and the detection limits were between 39.0 +/- 5.3 (n=9) and 500 +/- 23 (n=9) fmol per microl injected sample. Recoveries of all products from urine or water were 73.0 +/- 5.2% and higher. In urine of CCl4-treated rats an increase in all eight lipid degradation products in urine was found 24 h following exposure. ACON showed the most distinct increase, followed by PROPA, BUTA and MDA. It is concluded that the rapid, selective and sensitive analytical method based on GC-ECD presented here is well suited for routine measurement of eight different lipid degradation products. These products appear to be useful as non-invasive biomarkers for in vivo oxidative stress induced in rats by CCl4.

Epichlorohydrin (ECH) is used in many industrial processes. Different toxic effects of ECH were found in rodents. The metabolism of ECH was investigated before in rats using [14C]ECH. The aim of this investigation was the development of non-radioactive quantitative analytical methods for measuring two urinary metabolites of ECH, namely 3-chloro-2-hydroxypropylmercapturic acid (CHPMA) and alpha-chlorohydrin (alpha-CH). The identity of CHPMA and alpha-CH excreted in urine of rats treated with 5 to 35 mg/kg ECH was confirmed by GC-MS. The quantitative analysis of CHPMA, involving ethyl acetate extraction from acidified urine and subsequent methylation and analysis by gas chromatography-flame photometric detection (GC-FPD), showed a method limit of detection of 2 micrograms/ml. The analysis of alpha-CH based on ethyl acetate extraction and subsequent analysis by GC-ECD, showed a method limit of detection of 2 micrograms/ml. CHPMA and alpha-CH derivatives could be determined quantitatively down to concentrations of 0.5 and 0.4 micrograms/ml urine, respectively, by selected-ion monitoring GC-MS under EI conditions. Cumulative urinary excretion of CHPMA and alpha-CH by rats treated with ECH were found to be 31 +/- 10 and 1.4 +/- 0.6% (n = 13) of the ECH dose, respectively. For CHPMA, the dose-excretion relationship suggested partially saturated ECH metabolism. For alpha-CH, the doe-excretion relationship was linear. With fractionated urine collection it was found that approximately 74 and 84% of the total cumulative excretion of CHPMA and alpha-CH, respectively, took place within the first 6 h after administration of ECH. From these investigations it is concluded that the GC-FPD and GC-ECD based methods developed are sufficiently sensitive to measure urinary excretion of CHPMA and alpha-CH in urine from rats administered 5 to 35 mg/kg ECH. It is anticipated that the analysis of CHPMA and alpha-CH based on GC-MS may be sufficiently sensitive to investigate urinary excretion from humans occupationally exposed to ECH.

1. Potential sulphur-containing metabolites of the anticancer agent, fotemustine, were synthesized, namely thiodiacetic acid (TDA), S-2-hydroxyethyl N-acetyl-L-cysteine (2-HE-NAC), N-acetyl-L-cysteine (NAC), S-methyl N-acetyl-L-cysteine (M-NAC), S-carboxymethyl-L-cysteine (CM-Cys), S-carboxymethyl N-acetyl-L-cysteine (CM-NAC), their corresponding sulphoxides and sulphones. Their chemical structures and stabilities were confirmed and derivatization methods were developed for their analysis by sulphur-selective g.l.c. (g.l.c.-FPD) and g.l.c.-mass spectrometry. 2. Four methods for isolation of potential metabolites of fotemustine were developed. Quantification of metabolites, derived in various ways was carried out by g.l.c.-atomic emission detection (AED) or g.l.c.-mass spectrometry. 3. Male Wistar rats (n = 4) were given a single i.p. dose of 40 mg/kg fotemustine. Urine excretion of TDA (18.4 +/- 1.9% in 24 h) and TDA sulphoxide (12.0 +/- 1.6% in 24 h) was significant; 32.7 +/- 4.6% of the fotemustine dose was excreted as TDA, and TDA sulphoxide in 48 h. NAC was excreted in rat urine at 1% of the dose. No other potential glutathione-derived metabolites of fotemustine were excreted. 4. Male Wistar rats (n = 4) were also treated i.p. with fotemustine at 5, 20 and 40 mg/kg, to investigate dose dependency and the time course of excretion of TDA. Excretion of TDA in 48 h urine decreased from 32 +/- 2 to 17 +/- 2% dose (mean +/- SD) with increasing dose of fotemustine.

Cisplatin is a widely used antitumor drug, highly effective in the treatment of several tumors. Cisplatin exerts its antitumor activity through an interaction with DNA, which results in the formation of bidentate adducts. An important side-effects of cisplatin is nephrotoxicity. Selenite can reduce the nephrotoxicity of cisplatin without reducing the antitumor activity of the drug. We have studied the mechanism of selenite protection against cisplatin-induced nephrotoxicity. The protection correlates with higher levels of selenium in the kidney (about eight times) and with higher levels of glutathione in the kidney, both compared to tumors. Selenite is metabolized into selenols, specifically into methylselenol and glutathionylselenol; this bioactivation of selenite into selenols is a glutathione-dependent process. HPLC with on-line radioactivity detection of 195mPt showed that methylselenol is capable of forming a complex with cisplatin in vitro. 1H-NMR gave evidence that the complex contains one or more Pt-Se-CH3 bonds. Attempts to obtain further structural information by Desorption Chemical Ionization and Fast Atom Bombardment mass-spectrometry failed. It is proposed that the formation of a cisplatin-selenol complex also takes place in vivo, especially in the kidney, thereby preventing cisplatin exerting its nephrotoxic activity.