A validated liquid chromatography/tandem mass spectrometry method for 4,4'-methylenedianiline quantitation in human urine as a measure of 4,4'-methylene diphenyl diisocyanate exposure.

RATIONALE: 4,4'-Methylene diphenyl diisocyanate (MDI) is a highly reactive isocyanate used in the production of polyurethanes. Workers exposed to these products may develop sensitization to the diisocyanate compounds, leading to occupational asthma. Quantifying MDI levels is necessary to ensure workplace safety. MDI is metabolized by acetylation and/or conjugation to macromolecules for excretion into urine. All metabolites can be chemically hydrolyzed to form the free diamine 4,4'-methylenedianiline (MDA) as a urinary biomarker of MDI exposure. Current methods involve long sample preparation, or have been designed using costly automation. There is therefore a need to develop a new practical method for assessing exposure to MDI. METHODS: Urine samples were acidified and heated to form MDA, followed by neutralization and liquid-liquid extraction. Extracts were separated by reversed-phase chromatography on a HSS T3 column followed by analysis on a triple quadrupole mass spectrometer in multiple reaction monitoring (MRM) mode. RESULTS: 13 C15 N-MDA was selected as the internal standard (IS) of choice following an investigation of internal standard stability. The hydrolysis efficiency, forming free MDA from conjugated metabolites in vivo, was evaluated using 4,4'-methylenebis(acetanilide) spiked into urine and complete hydrolysis occurred after 1 h. A dynamic range of 5 to 500 nM was achieved, and was useful for monitoring MDI exposure considering the biological guidance value (BGV) of 10 µg/L (~50 nM) proposed by the German Research Foundation (DFG). The limit of detection (LOD) and limit of quantification (LOQ) of the method were 0.8 and 2.7 nM, respectively. The intra-day and inter-day precisions were 4.33% and 4.27%, respectively. Finally, the method was tested with inter-laboratory samples from the German External Quality Assessment Scheme (G-EQUAS) program and the results submitted were all within the allowable tolerance range. CONCLUSIONS: A practical and validated method for the analysis of small- to medium-sized batches of samples has been developed for the biological monitoring of MDI exposure in human urine.

[Determination of methylenedianiline in urine by high performance liquid chromatography-tandemmass spectrometry].

Objective: To develop a method for determination of metabolites of diphenylmethane diisocyanate (MDI) in urine, i.e. methylenedianiline (MDA) by high performance liquid chromatography-tandem mass (LC-MS-MS) . Methods: Urine samples were prepared by hydrolyzation with sulfuric acid and extraction by acetonitrile, and then separated on a Shim-pack XR-ODS column, analyzed with high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) . The external solvent standard calibration were tested. Results: The linearity ranges were 0.05~20.00 µg/L, The related coefficients were 0.999 5. The limit of detection was 0.02 µg/L. The rats of recovery were 91.0%~103.4%. The relative standard deviations were between 2.7%~7.3%. Conclusion: The method was sensitive, accurate and suitable for the MDA determination in urine of MDI exposed population.

Effect of CYP3A4 genetic polymorphisms on the genotoxicity of 4,4'-methylene-bis(2-chloroaniline)-exposed workers.

OBJECTIVES: We investigated the relationship between 4,4'-methylene-bis(2-chloroaniline) (MBOCA) exposure and micronucleus (MN) frequency, and how this association was affected by genetic polymorphism of the cytochrome P450 enzyme (CYP3A4). METHODS: We divided the study population into an exposed group (n=44 with total urine MBOCA ≥20 µg/g creatinine) and a control group (n=47 with total urine MBOCA <20 µg/g creatinine). Lymphocyte MN frequency (MNF) and micronucleated cell (MNC) frequency were measured by the cytokinesis-block MN assay method. MNF reported as the number of micronuclei in binucleated cells per 1000 cells, and MNC reported as the number of binucleated cells with the presence of MN per 1000 cells. CYP3A4 alleles were measured by PCR-based restriction fragment length polymorphism (PCR-RFLP). RESULTS: The mean MNF (6.11 vs 4.46 MN/1000 cells, p<0.001) and MNC (5.75 vs 4.15 MN/1000 cells, p<0.001) in the exposed workers was significantly higher than that in the controls. The CYP3A4 polymorphism A/A+A/G influenced the difference in the mean MNF (5.97 vs 4.38 MN/1000 cells, p<0.001) and MNC (5.60 vs 4.15 MN/1000 cells, p<0.001) between the MBOCA-exposed and control groups. After adjusting risk factors, the MNF level in the MBOCA-exposed workers was 0.520 MN cells/1000 cells (p<0.001) higher than the control group among the CYP3A4 A/A+A/G genotype. Similarly, the MNC level in the MBOCA-exposed workers was 0.593 MN/1000 cells (p<0.001) higher than the control group among the CYP3A4 A/A+A/G genotype. However, the difference in adjusted MNF and MNC between the exposed and control groups was not significant for the CYP3A4 polymorphism with the G/G genotype. CONCLUSIONS: We recommend that lymphocytes MNF and MNC are good indicators to evaluate MBOCA genotoxicity. Individuals with the CYP3A4 polymorphism A/A and A/G genotypes appear to be more susceptible to MBOCA genotoxicity.

Two liquid chromatographic approaches for the simultaneous determination of xipamide and its degradation product (2,6-xylidine) using time-programmed fluorescence detection.

A study of the performance of reversed-phase chromatography with a programmable multiwavelength fluorimetric technique using either conventional hydro-organic or micellar eluent is established for the determination of xipamide (XIP) in the presence of its degradation product, 2,6-xylidine (XY). In conventional liquid chromatography (CLC), the analyses were carried out on a Promosil ODS 100 Å column (250 mm × 4.6 mm i.d., 5 µm) using a mobile phase consisting of methanol/0.1 M phosphate buffer (65: 35, v/v) at pH 4.0. For micellar liquid chromatography (MLC), a short Spherisorb column (150 mm × 4.6 mm i.d., 5 µm) was employed in conjunction with a greener mobile phase (pH 5.0) containing 0.1 M sodium dodecyl sulfate and 15% n-propanol. CLC proved to be superior to MLC in terms of sensitivity for the determination of the degradation product because it could detect trace amounts down to 10.0 ng/ml of XY as a degradation product in XIP. However, MLC represents an eco-friendly approach for the simultaneous determination of XIP and XY. In addition, the opportunity for the direct introduction of biological matrices into the chromatographic system is one of the distinctive benefits of MLC. The proposed methods were applied for the determination of XIP in its tablets, human urine and content uniformity testing. The results of the proposed methods were statistically compared with those obtained using the comparison fluorimetric method, revealing no significant differences in the performance of the methods regarding accuracy and precision.

Isotope Dilution UPLC-APCI-MS/MS Method for the Quantitative Measurement of Aromatic Diamines in Human Urine: Biomarkers of Diisocyanate Exposure.

Urinary diamines are biomarkers of diisocyanate exposure. Diisocyanates are considered as skin and respiratory sensitizers and are the most frequently reported cause of occupational asthma. Herein we report on the development and validation of an ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the measurement of five aromatic diamines, 4,4'-methylenedianiline (MDA), 2,4-toluenediamine (4TDA), 2,6-toluenediamine (6TDA), 1,5-naphthalenediamine (NDA), and p-phenylenediamine (PPDA) in human urine. The method incorporates sample preparation steps, which include a 4 h acid hydrolysis followed by high-throughput solid-phase extraction prior to chromatographic separation. Chromatographic separation was achieved using a C18 reversed phase column with gradient elution of basic mobile phases (pH 9.2). The duty cycle of the method was less than 5 min, including both the column equilibration and autosampler movement. Analytical detection was performed using positive ion atmospheric pressure chemical ionization tandem mass spectrometry (APCI-MS/MS) in scheduled multiple reaction monitoring (sMRM) mode. Excellent linearity was observed over standard calibration curve concentration ranges of 3 orders of magnitude with method detection limit ranging from 10 to 100 pg/mL. The interday and intraday reproducibility and accuracy were within ±15%. This method is fast, accurate, and reproducible and is suitable for assessment of exposure to the most common aromatic diisocyanates within targeted groups as well as larger population studies such as the National Health and Nutrition Examination Survey (NHANES).

Human metabolism and excretion kinetics of aniline after a single oral dose.

Aniline is an important source material in the chemical industry (e.g., rubber, pesticides, and pharmaceuticals). The general population is known to be ubiquitously exposed to aniline. Thus, assessment of aniline exposure is of both occupational and environmental relevance. Knowledge on human metabolism of aniline is scarce. We orally dosed four healthy male volunteers (two fast and two slow acetylators) with 5 mg isotope-labeled aniline, consecutively collected all urine samples over a period of 2 days, and investigated the renal excretion of aniline and its metabolites by LS-MS/MS and GC-MS. After enzymatic hydrolysis of glucuronide and sulfate conjugates, N-acetyl-4-aminophenol was the predominant urinary aniline metabolite representing 55.7-68.9 % of the oral dose, followed by the mercapturic acid conjugate of N-acetyl-4-aminophenol accounting for 2.5-6.1 %. Acetanilide and free aniline were found only in minor amounts accounting for 0.14-0.36 % of the dose. Overall, these four biomarkers excreted in urine over 48 h post-dose represented 62.4-72.1 % of the oral aniline dose. Elimination half-times were 3.4-4.3 h for N-acetyl-4-aminophenol, 4.1-5.5 h for the mercapturic acid conjugate, and 1.3-1.6 and 0.6-1.2 h for acetanilide and free aniline, respectively. Urinary maximum concentrations of N-acetyl-4-aminophenol were reached after about 4 h and maximum concentrations of the mercapturic acid conjugate after about 6 h, whereas concentrations of acetanilide and free aniline peaked after about 1 h. The present study is one of the first to provide reliable urinary excretion factors for aniline and its metabolites in humans after oral dosage, including data on the predominant urinary metabolite N-acetyl-4-aminophenol, also known as an analgesic under the name paracetamol/acetaminophen.

Ethnic comparison of pharmacokinetics of (18)F-florbetaben, a PET tracer for beta-amyloid imaging, in healthy Caucasian and Japanese subjects.

PURPOSE: (18)F-Florbetaben is a positron emission tomography (PET) tracer indicated for imaging cerebral beta-amyloid deposition in adult patients with cognitive impairment who are being evaluated for Alzheimer's disease and other causes of cognitive decline. The present study examined ethnic comparability of the plasma pharmacokinetics, which is the input to the brain, between Caucasian and Japanese subjects. METHODS: Two identical phase I trials were performed in 18 German and 18 Japanese healthy volunteers to evaluate the plasma pharmacokinetics of a single dose of 300 MBq (18)F-florbetaben, either of low (≤5 µg, LD) or high (50-55 µg, HD) mass dose. Pharmacokinetic parameters were evaluated based on the total (18)F radioactivity measurements in plasma followed by metabolite analysis using radio-HPLC. RESULTS: The pharmacokinetics of (18)F-florbetaben was characterized by a rapid elimination from plasma. The dose-normalized areas under the curve of (18)F-florbetaben in plasma as an indicator of the input to the brain were comparable between Germans (LD: 0.38 min/l, HD: 0.55 min/l) and Japanese (LD: 0.35 min/l, HD: 0.45 min/l) suggesting ethnic similarity, and the mass dose effect was minimal. A polar metabolite fraction was the main radiolabelled degradation product in plasma and was also similar between the doses and the ethnic groups. CONCLUSION: Absence of a difference in the pharmacokinetics of (18)F-florbetaben in Germans and Japanese has warranted further global development of the PET imaging agent.

The metabolism of 4-bromoaniline in the bile-cannulated rat: application of ICPMS ((79/81)Br), HPLC-ICPMS & HPLC-oaTOFMS.

1. An excretion balance study was performed following i.p. administration of 4-bromoaniline (50 mg kg(-1)) to bile-cannulated rats, using bromine-detected ((79/81)Br) ICPMS for quantification. Approximately 90% of the dose was recovered in urine (68.9 ± 3.6%) and bile (21.4 ± 1.4%) by 48 h post-administration. 2. HPLC-ICPMS ((79/81)Br) was used to selectively detect and profile the major urinary and biliary-excreted metabolites and determined that the 0-12 h urine contained at least 21 brominated metabolites with 19 bromine-containing peaks observed in the 6-12 h bile samples. 3. The urinary and biliary metabolites were subsequently profiled using HPLC-oaTOFMS. By exploiting the distinctive bromine isotope pattern ca. 60 brominated metabolites were detected in the urine in negative electrospray ionisation (ESI) mode while bile contained ca. 21. 4. While a large number of bromine-containing metabolites were detected, the profiles were dominated by a few major components with the bulk of the 4-bromoaniline-related material in urine accounted for by 4-bromoanaline O-sulfate (∼75% of the total by ICPMS, 84% by TOFMS). In bile a hydroxylated N-acetyl compound was the major metabolite detected, forming some ∼65% of the 4-bromoaniline-related material by ICPMS (37% by TOFMS).

Human exposure to airborne aniline and formation of methemoglobin: a contribution to occupational exposure limits.

Aniline is an important starting material in the manufacture of polyurethane-based plastic materials. Aniline-derived methemoglobinemia (Met-Hb) is well described in exposed workers although information on the dose-response association is limited. We used an experimental design to study the association between aniline in air with the formation of Met-Hb in blood and the elimination of aniline in urine. A 6-h exposure of 2 ppm aniline in 19 non-smoking volunteers resulted in a time-dependent increase in Met-Hb in blood and aniline in urine. The maximum Met-Hb level in blood (mean 1.21 ± 0.29 %, range 0.80-2.07 %) and aniline excretion in urine (mean 168.0 ± 51.8 µg/L, range 79.5-418.3 µg/L) were observed at the end of exposure, with both parameters rapidly decreasing after the end of exposure. After 24 h, the mean level of Met-Hb (0.65 ± 0.18 %) returned to the basal level observed prior to the exposure (0.72 ± 0.19 %); whereas, slightly elevated levels of aniline were still present in urine (means 17.0 ± 17.1 vs. 5.7 ± 3.8 µg/L). No differences between males and females as well as between slow and fast acetylators were found. The results obtained after 6-h exposure were also comparable to those observed in four non-smoking volunteers after 8-h exposure. Maximum levels of Met-Hb and aniline in urine were 1.57 % and 305.6 µg/L, respectively. Overall, our results contribute to the risk assessment of aniline and as a result, the protection of workers from aniline-derived adverse health effects at the workplace.

Dispersive liquid-liquid microextraction based on solidification of floating organic drop combined with field-amplified sample injection in capillary electrophoresis for the determination of beta(2)-agonists in bovine urine.

Dispersive liquid-liquid microextraction based on solidification of floating organic drop (DLLME-SFO) was for the first time combined with field-amplified sample injection (FASI) in CE to determine four ß(2)-agonists (cimbuterol, clenbuterol, mabuterol, and mapenterol) in bovine urine. Optimum BGE consisted of 20 mM borate buffer and 0.1 mM SDS. Using salting-out extraction, ß(2)-agonists were extracted into ACN that was then used as the disperser solvent in DLLME-SFO. Optimum DLLME-SFO conditions were: 1.0 mL ACN, 50 µL 1-undecanol (extraction solvent), total extraction time 1.5 min, no salt addition. Back extraction into an aqueous solution (pH 2.0) facilitated direct injection of ß(2)-agonists into CE. Compared to conventional CZE, DLLME-SFO-FASI-CE achieved sensitivity enhancement factors of 41-1046 resulting in LODs in the range of 1.80-37.0 µg L(-1). Linear dynamic ranges of 0.15-10.0 mg L(-1) for cimbuterol and 15-1000 µg L(-1) for the other analytes were obtained with coefficients of determination (R(2)) ≥ 0.9901 and RSD% ≤5.5 (n = 5). Finally, the applicability of the proposed method was successfully confirmed by determination of the four ß(2)-agonists in spiked bovine urine samples and accuracy higher than 96.0% was obtained.

Urinary bio-monitoring of aromatic amine derivatives by new needle trap device packed with the multi-component adsorbent.

Aromatic amines are a large group of chemical compounds that have attracted the attention of researchers due to their toxicity and carcinogenicity. This study aimed to develop an efficient method for sampling and analysis of aromatic amines (Aniline, N, N-dimethylaniline, 2-chloroaniline, and 3-chloroaniline) from the vapour phase (headspace) of urine samples. For the implementation of this plan, a needle trap device packed with the three-component adsorbent consisting of nano-Hydroxy Apatite (nHA), Zeolite (Ze), and Metal-Organic Framework (MOF) equipped with GC-FID was employed for the first phase. Examination of the prepared adsorbents was performed by FT-IR, PXRD, and FE-SEM techniques. The optimal value of considerable parameters such as time and temperature of extraction, salt content, and pH were established using the Response Surface Methodology-Central Composite Design (RMS-CCD) method. In this way, the optimal extraction of targeted analytes was accomplished in 41 min at 41 °C with NaCl content of 33.0% (w/v) and pH: 13.0, respectively. Also, the repeatability and reproducibility of the method were calculated to be in the range of 2.2-7.1% and 3.9-8.1%, respectively, which indicates the acceptable precision of the method. Also, the limit of detection (LOD) and limit of quantification (LOQ) were determined in the range of 0.3-32.0 ng.L-1 and 0.8-350.0 ng.L-1, respectively, which proves the high sensitivity of the proposed method. Furthermore, the recovery percent of the extracted analytes was concluded in the range of 97.0-99.0% after 6 and 30 days of the sampling and storage at 25 °C and 4 °C, respectively. Finally, the designed procedure was employed in the analysis of the above-mentioned aromatic amines in the real urine samples. The achieved results illustrate that the three-component absorbent system (nHA;Ze;MOF@NTD) can be introduced as an efficient, fast-response, sensitive, and versatile procedure for trace analysis of the different aromatic amine compounds in public and occupational health.

Differences in the pharmacokinetics of 4-amino-3-chlorophenyl hydrogen sulfate, a metabolite of resatorvid, in rats and dogs.

The pharmacokinetics of 4-amino-3-chlorophenyl hydrogen sulfate, M-III of resatorvid, in rats and dogs were investigated using radiolabeled M-III ([(14)C]M-III). The elimination half-life of (14)C in the plasma of rats was approximately 1/30 of that of dogs after intravenous dosing of [(14)C]M-III at 0.5 mg/kg to rats and dogs. The in vitro and in vivo plasma protein binding ratios of M-III were relatively high and were the same in both species. The intrinsic clearance (CL(int)) of M-III in rats was much higher than the glomerular filtration rate in rats. Furthermore, the concentration of [(14)C]M-III in the kidney of rats was much higher than that in the plasma. On the contrary, in dogs, the concentration of [(14)C]M-III in the kidney was very much lower than that in the plasma. These results indicated that M-III was effectively taken up into the kidney and was excreted into the urine in rats; however, in dogs, ineffective renal uptake of M-III was presumed. When [(14)C]M-III and probenecid were simultaneously and continually infused intravenously to rats, the CL(int) of M-III decreased with increasing plasma concentrations of probenecid, indicating that kidney uptake of M-III in rats was inhibited by probenecid. It was also thought that uptake by the organic anion transport system(s) in the basolateral membrane is involved in the renal uptake of M-III in rats. The pharmacokinetic differences of M-III between rats and dogs are considered to be mainly caused by the difference in the urinary excretion via the renal distribution processes.

Metabolism and disposition of 2-methoxy-4-nitroaniline in male and female Harlan Sprague Dawley rats and B6C3F1/N mice.

The disposition of 2-Methoxy-4-nitroaniline (MNA) was investigated in male and female Harlan Sprague Dawley rats and B6C3F(1)/N mice following oral, intravenous, and dermal exposure to [(14)C]MNA at 2, 15, or 150 mg/kg. Clearance of MNA was investigated in male and female rat, mouse, and human hepatocytes. MNA was cleared slowly in hepatocytes from rat (t(1/2) = 152-424 min) and human (t(1/2) = 118-403 min) but faster in mouse (t(1/2)= 70-106 min). MNA was well-absorbed in rats and mice following oral administration and eliminated chiefly in urine (rats, 75-79%; mice, 55-68%) 72 h post dosing. Less than 1% of the radioactivity remained in tissues at 72 h. MNA was poorly absorbed following dermal application in rats (5.5%) and mice (10%) over 24 h. The major pathway of metabolism of MNA was via hydroxylation of the phenyl ring to form 6-hydroxy MNA; major metabolites detected were sulfate and glucuronide conjugates of 6-hydroxy MNA. Following oral administration, the percent of total radioactivity bound in tissues bound was highest in liver (43%) and red blood cells (30%), whereas the radioactivity bound to DNA was highest in cecum (160 pmol/mg DNA).

A liquid chromatography-tandem mass spectrometry method for the analysis of primary aromatic amines in human urine.

Aromatic amines are widely used in personal care products and human exposure to this class of chemicals is widespread. Bioanalytical methods to determine trace levels of aromatic amines in human urine are scarce. In this study, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed to determine 39 primary aromatic amines (AAs) along with nicotine and cotinine in human urine. Chromatographic separation of the 41 analytes was achieved on an Ultra Biphenyl (100 mm × 2.1 mm, 5 µm) column. Mass spectrometry was operated in electrospray ionization positive ion multiple reaction monitoring (MRM) mode. The method exhibited excellent linear dynamic range (0.1-50 ng/mL) with correlation coefficients (r) > 0.999 for all analytes. Urine samples (2 mL) were hydrolyzed using 10 M NaOH at 95 °C for 15 h and target analytes were extracted using methyl-tert-butyl ether (MTBE). Addition of 15 µL of 0.25 M HCl to the sample extracts improved the recoveries of several target analytes. The method was validated through the analysis of fortified quality control (QC) samples and a certified standard reference material (SRM). Relative recoveries (%) of target analytes fortified in QC samples were in the range of 75-114% for 37 of the 41 analytes while the other analytes exhibited lower recoveries (16-74%). The limits of detection (LOD) and limits of quantification (LOQ) of target analytes were in the range of 0.025-0.20 ng/mL and 0.1-1.0 ng/mL, respectively. Intra-day and inter-day precision of the method assessed through the analysis of fortified urine QC samples at three different concentrations were < 11.7% and < 15.9% (measured as RSD), respectively. The method was applied in the analysis of urine samples from the general population and known smokers; aniline, para-anisidine, para-toluidine, ortho/meta-toluidine, 3-chloroaniline, 4-chloroaniline, 3,4-dichloroaniline, and 4,4'-methylenedianiline were found in all smoker's urine at sum concentrations ranging from 0.04 to 9.16 ng/mL.

Determination of osimertinib in human plasma, urine and cerebrospinal fluid.

Aim: Osimertinib (Tagrisso, AZD9291) has been approved for the treatment of patients with metastatic EGFRm T790M non-small-cell lung cancer. Results: Rapid and sensitive LC-MC/MS methods were developed for osimertinib and its metabolites, AZ13597550 and AZ13575104, in human plasma (low- and high-range), urine and cerebrospinal fluid. We discuss the challenges of these multi-analyte and multiple matrix assays. The methods have been successfully validated and used for the analysis of over 20,000 clinical samples, with successful incurred sample reproducibility. Conclusion: The assays have been shown to be selective, accurate and robust, providing high-throughput analysis during the clinical development of osimertinib.

Aniline in hydrolyzed urine and plasma--possible biomarkers for phenylisocyanate exposure.

There are few studies on phenylisocyanate (PhI) exposure, although there are studies indicating that PhI is a very potent chemical sensitizer. The aim of this study was to evaluate aniline in urine and plasma as possible biomarkers of exposure to PhI. Occupational airborne exposure to PhI was measured during one day for 11 workers exposed to thermal degradation products from polyurethane with filters impregnated with 2-methoxyphenyl piperazine. A urine sample was collected from each worker on measurement day, and plasma samples were collected within the following 2 weeks. Urine and plasma samples also were collected from four unexposed subjects. The biological samples were hydrolyzed and analyzed with gas chromatography mass spectrometry. The time-weighted averages (TWA) for the workers were between 0.1 and 1.6 microg/m3. Aniline levels in urine were in the same range for the exposed and unexposed workers, but there was a significant correlation between air and urinary levels (Pearson's correlation coefficient r = 0.518; p = 0.05). All exposed workers had higher levels in the plasma samples than the highest control, and there was a significant correlation between the plasma levels and measured air levels (r = 0.675; p = 0.008). The conclusion is that aniline in hydrolyzed urine and plasma are possible biomarkers of exposure to PhI, and that the plasma biomarker is more sensitive, at least at this rather low exposure.

The point-of-care colorimetric detection of the biomarker of phenylamine in the human urine based on Tb3+ functionalized metal-organic framework.

Phenylamine has been recognized as one of the most important industrially relevant ingredient and a crucial intermediate in chemical products. Yet, its internal exposure detection in human remains largely elusive due to the lack of potent monitoring method. Hereby this issue is addressed with a probe based on lanthanide functionalized organic-inorganic hybrid material Al(OH)(bpydc) (1) through post-synthetically modified metal-organic framework. The as-synthesized Tb3+@1 exhibits the strong luminescence of Tb3+ originated from efficient energy transfer from the ligand, which can sense the biological metabolite p-aminophenol (PAP) of the phenylamine in the human urine. Linear correlation between the integrated fluorescence intensity and the concentration of PAP was investigated, enabling quantitative analysis of PAP in physiologically ranges (0.005-5 mg mL-1) with low detection limit (5 µg mL-1). This probe demonstrates excellent sensitivity, high selectivity, good reusability and quick response to PAP. Furthermore, a simple and rapid smartphone-based medical portable test paper was developed, whose quantitative color change can be easily distinguished visually. Hence, the PAP sensing platform can serve as a potential diagnostic tool for home monitoring of PAP.

Detection of aminophenylnorharman, a possible endogenous mutagenic and carcinogenic compound, in human urine samples.

Mutagenic/carcinogenic 9-(4'-aminophenyl)-9H-pyrido[3,4-b]indole [aminophenylnorharman (APNH)] is formed from norharman and aniline in the presence of cytochrome P450 3A4/1A2. Because both precursors are widely distributed in the environment, human exposure is unavoidable. To clarify APNH formation in the human body, amounts of the compound in 24-h human urine collected from smokers and nonsmokers, eating a normal diet, were analyzed by liquid chromatography/electrospray ionization tandem mass spectrometry. In addition, norharman and aniline were also analyzed by high-performance liquid chromatography and gas chromatography, respectively. APNH could be detected in all urine samples at levels 49 to 449 pg for smokers and 21 to 594 pg for nonsmokers per 24-h urine, respectively. The amounts of norharman and aniline were 46 to 185 ng and 0.70 to 8.10 microg for smokers and 52 to 447 ng and 0.49 to 5.72 microg for nonsmokers, respectively, per 24-h urine (none of the levels differing significantly between smokers and nonsmokers). To exclude exogenous exposure to norharman and aniline, we analyzed the levels of APNH, norharman, and aniline in urine samples collected from inpatients receiving parenteral alimentation. Similar to the healthy volunteers, all urine samples contained 12 to 338 pg of APNH, 6 to 75 ng of norharman, and 0.33 to 1.86 microg of aniline per 24-h urine. These results suggest that APNH should be considered as a novel endogenous mutagen/carcinogen; thus, it is very important to determine the biological significance of this carcinogen for human cancer development.

[Occupational and environmental exposure to anilide and dicarboximide pesticides]. / Esposizione ambientale e professionale a pesticidi propionanilidici e dicarbossimidici.

Dicarboximide fungicides (DF) such as vinclozolin, iprodione, procymidone are widely used on vines, fruit and vegetables, and anilide herbicides (AH) such as diuron, linuron, propanil are used to control weeds on hard surfaces, such as, roads, railway tracks, paths, and in crops, forestry. Italian reports on food safety found many samples contaminated by pesticides belonging to these categories, even though only few exceeding L.M.R. Since adverse effects on human health, such as endocrine disruption, have been reported, biological monitoring is essential for exposure assessment both of occupationally exposed subjects and of the general population. Common metabolites of DF and AH are dichloroanilinines such as 3,4-DCA and 3,5-DCA, urine samples from 153 subjects living in Novafeltria, central Italy, were collected for analysis of 3,4- and 3,5-DCAs, each participant was invited to complete a very detailed questionnaire. A total of 151 out of 153 samples were found to be positive for 3,5-DCA, and 81.7% were positive for 3,4-DCA. Also 33 workers, engaged in application of propanil on rice in northern Italy, were involved in the study and 3,4-DCA was determined as marker of exposure. 3,4 and 3,5 dichloroaniline are useful and promising biological indicators for monitoring occupational and environmental exposure to these classes of pesticides.

Mass spectrometric characterization of efaproxiral (RSR13) and its implementation into doping controls using liquid chromatography-atmospheric pressure ionization-tandem mass spectrometry.

Efaproxiral (2-[4-[[(3,5-dimethylanilino)carbonyl]methyl]phenoxyl]-2-methylpropionic acid, formerly referred to as RSR13) is prohibited in sports according to the World Anti-Doping Agency (WADA). The drug as well as structurally related compounds and a stable isotope-labeled derivative have been synthesized to elucidate the fragmentation pathway of efaproxiral, using electrospray ionization (ESI) and tandem mass spectrometry by employing a novel linear ion trap--orbitrap hybrid mass spectrometer--in positive and negative ionization modes. The elimination of 2-methyl acrylic acid (-86 u) has been identified as a major fragmentation process in both charge states. Negative ionization and collision-induced dissociation (CID) caused an additional release of carbon dioxide (-44 u), and positive ionization the loss of formic acid (-46 u). Efaproxiral was incorporated into an existing screening procedure for doping controls using solid-phase extraction (SPE) followed by liquid chromatography-tandem mass spectrometry, enabling a limit of detection of 2.5 ng/ml and interday precisions ranging from 7.9 to 13.0%.