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Background Glyphosate and its salt formulations are nonselective herbicides that have been extensively used worldwide, both for residential and agricultural purposes. The possible carcinogenicity and teratogenicity of glyphosate remain to be elucidated. We developed a sensitive and high-throughput analytical method for urinary glyphosate using liquid chromatography-tandem mass spectrometry with the aim of contributing to glyphosate exposure assessment in epidemiological studies. Methods After urine dilution (creatinine matching dilution to 0.05 g creatinine/L), glyphosate was extracted using two types of solid phase extraction columns (SCX and NH2) with automated sample preparation instruments. The eluate was dried and dissolved in the mobile phase, followed by liquid chromatography-tandem mass spectrometry analysis. The optimized method was applied to urine samples obtained from 54 Japanese adults and children. Results The results from the validation study demonstrated good recoveries (91.0-99.6%), within- and between-run precisions (< 15%), low detection limits (0.1 μg/L), and lower limit of quantification (0.3 μg/L). The detection frequency and median concentration of the urinary glyphosate in Japanese subjects were 59% and 0.25 μg/L (0.34 μg/g creatinine). Conclusions Our reliable determination method was successful in measuring urinary glyphosate concentration. Moreover, this is the first biomonitoring report of urinary glyphosate levels in the Japanese general population.

Over the past two decades, domestic shipments of glyphosate (Gly), in the form of an ionic salt, have been increasing steadily in Japan. This increase has raising concerns about the effects of chemical exposure on children. The International Agency for Research on Cancer classified Gly as a "probably carcinogenic to humans (Group 2A)" in 2015. The purpose of the current study was to analyze Gly in urine samples of Japanese children to determine temporal changes, seasonal changes, and gender differences.First-morning urine samples were obtained from 50 Japanese children (4-6-year-old) in October of 2006, 2011, and 2015 (total = 150) to investigate the temporal trends in urinary Gly concentrations. Additionally, first-morning urine samples were collected from 3-year-old children in August-September of 2012 (summer; n = 42) and in February of 2013 (winter; n = 42) to investigate the seasonal and gender differences, and the correlations between urinary Gly concentrations and insecticide exposure biomarkers. Urine samples were analyzed to measure for Gly using a liquid chromatography with tandem mass spectrometry (LC-MS/MS).Detectable Gly concentrations were found in 41% of the 234 children. The 75th percentile and maximum concentrations of urinary Gly were 0.20 and 1.33 μg/L, respectively. The urinary Gly concentration in 2015 was significantly higher than in 2006, suggesting that the Gly exposure levels have been increasing. No seasonal or gender-specific differences in urinary Gly concentrations were observed, and no correlation with insecticide exposure biomarkers was found.This study revealed that Gly exposure trends show an increase between 2006 and 2015, and that season and gender were not the exposure-determining factors. Overall, urinary concentrations of Gly were comparable with studies from other countries.

Four pyrethroids (PYRs), metofluthrin, profluthrin, tefluthrin, and transfluthrin, which were newly developed and have relatively high vapor activity at ambient temperature, are now playing a key role in safely controlling insects in our daily lives. We developed a sensitive and high-throughput determination method for urinary metabolites derived from the newly developed PYR, e.g., 2,3,5,6-tetrafluoro-1,4-benzenedimethanol (HOCH2-FB-Al), 2,3,5,6-tetrafluorobenzyl alcohol (FB-Al), and other PYR metabolites such as trans-chrysanthemumdicarboxylic acid (trans-CDCA) and 3-phenoxybenzoic acid (3PBA). After high temperature acid hydrolysis of 2 mL urine sample in 24-deep well plate, the PYR metabolites were extracted by semi-automated liquid-liquid extraction with tert-butyl methyl ether. N,O-Bis (trimethylsilyl) trifluoroacetamide containing 1% trimethylchlorosilane or 1,1,1,3,3,3-hexafluoroisopropanol were used for the derivatization of PYR metabolites, and the derivatized metabolites were analyzed separately by GC-MS/MS equipped with dual injector system (DB-5MS and mid- to high-polarity phase Rtx-65 columns). The derivatization and evaporation conditions were mainly optimized for improving sensitivity and reproducibility. The mean within-run day precisions were less than 18.4% (relative standard deviation, %RSD) with low detection limits ranging from 0.01 μg/L for HOCH2-FB-Al to 0.06 μg/L for trans-CDCA. This method was successfully applied to urine samples obtained from 50 3-year-old children with high detection frequencies (e.g., 82% for HOCH2-FB-Al and 84% for FB-Al). This method may be a pivotal tool for developing risk assessment from PYR exposure in the general population.

It has been reported that the oxygen pressure of a photosensitizer medium decreases during an irradiation leading to decrease in the efficacy of the photosensitization reaction against the target cell in vitro.The aim of this study was to obtain solutions with high dissolved oxygen levels in cultivated wells with perceiving oxygen environment and photosensitizer bleaching for photosensitization reaction studies.We used a 10-mm-wide optical cell cuvette with a 1-mm optical path length as the well. A red blood cell (RBC) suspension with a hematocrit level of 0.625% was employed as the optical sample. The photosensitizer talaporfin sodium was added to a concentration of 30 μg/mL. The optical sample was irradiated by a 663-nm diode laser at 120 mW/cmThe relationship between oxygen saturation and the absorption peak in the Q band from the talaporfin sodium dynamics with increasing radiant exposure was revealed by our method with approximately twofold oxygen-dissolved solution.We could perceive the oxygen environment and the photosensitization reaction progression simultaneously with increasing dissolved oxygen by adding RBCs to the cell medium and measuring the absorption spectrum of it.

To reveal the mechanism of vascular patency in the myocardium after photosensitization immediately after talaporfin sodium (TS) injection in a canine model, we investigated acute injury to vascular endothelial cells (VECs) in vitro and in vivo.There are many reports of vascular shutdown within the target region in photodynamic therapy with TS. Vascular patency within healthy canine myocardium in which a photosensitization reaction starts immediately after injection of TS has been reported.TS fluorescence in human umbilical vein endothelial cells and cell lethality were measured with drug contact time (DCT) up to 120 min at 20 μg/mL. Dependence of radiant exposure on cell lethality with 60 min DCT was investigated using two albumin concentrations that corresponded to those in plasma and interstices. Irradiation (21 mW/cm) outside the adventitia of canine cervical veins for 167 or 667 sec was emitted through a diffuser probe 30 min after intravenous injection of TS (2.5 mg/kg). Veins were extracted ∼30 min after the reaction and stained with von Willebrand factor.Intracellular fluorescence increased, but not cell lethality, with increasing DCT. Cell lethality increased gradually and reached 100% over 20 J/cmVEC injury after a photosensitization reaction to healthy tissue shortly after TS injection might be low enough for the blood vessels to be patent.

Background and aim Metofluthrin, profluthrin, tefluthrin, and transfluthrin are pyrethroid (PYR) insecticides increasingly used to control mosquitoes, flies, and moths in households and public places (hygiene-PYRs). Currently, there is limited data available concerning exposure to these novel hygiene-PYRs. The goal of this study was to monitor exposure to these hygiene-PYRs by analysing their urinary metabolites and to investigate the temporal and seasonal trends in the concentrations of these metabolites. Methods First morning urine samples were obtained from 50 Japanese children (four-six years old) in October of 2006, 2011, and 2015 (total = 150 children) in order to investigate temporal trends. Additionally, first-morning urine samples were collected from 44 three-year-old children in August–September of 2012 (summer) and in February of 2013 (winter) to investigate seasonal differences. The urinary concentrations of 2,3,5,6-tetrafluorobenzyl alcohol (FB-Al; a specific metabolite of transfluthrin), 4-methyl-2,3,5,6-tetrafluorobenzyl alcohol (CH3-FB-Al; a common metabolite of tefluthrin and profluthrin), 4-methoxymethyl-2,3,5,6-tetrafluorobenzyl alcohol (CH3OCH2-FB-Al; a specific metabolite of metofluthrin), and 2,3,5,6-tetrafluoro-1,4-benzenedimethanol (HOCH2-FB-Al; a common metabolite of metofluthrin, tefluthrin, and profluthrin) were measured using GC-MS/MS. Results For the investigated years, rapid increases in the detection rates of the hygiene-PYR metabolites were observed. In 2015, FB-Al was identified in 64% of the samples, CH3-FB-Al in 46%, CH3OCH2-FB-Al in 50%, and HOCH2-FB-Al in 83%. Significant increasing trends were found for the concentrations of all hygiene-PYR metabolites from 2006 to 2015 (Jonckheere-Terpstra test, p Conclusions These findings suggest that, in Japanese children, exposure to hygiene-PYRs has increased over the past decade, and that children are exposed to higher levels of hygiene-PYRs in summer than in winter.

Background We investigated light attenuation at 664 nm, which is the excitation wavelength of photodynamic therapy (PDT) using talaporfin sodium, in a salted cadaver brain. Estimation of therapeutic lesions is important to ensure the effectiveness and safety of brain tumor PDT. Previously reported optical properties of the human brain vary widely. In this study, we measured the light attenuation in brain tissue using a practical method. We employed a salted cadaver brain, in which the mechanical and optical properties can be maintained as close as possible to those under operative conditions. Methods A neuroendoscope was inserted into the brain until the cerebral ventricle was reached. A thin cylindrical diffuser probe was advanced 10 mm from the endoscope tip. By another path from the brain surface, an optical fiber for measurement was inserted into a puncture needle, and a pair of needles was used to puncture the tissue and reach the same cerebral ventricle in which the endoscope tip was positioned. The attenuation of light intensities in the frontal lobe and cerebellum was measured by varying the bundle tip position. The starting positions of the bundle were confirmed by the endoscopic view. The measured light intensity attenuations were fitted with an exponential curve. Results The following attenuation coefficients were obtained: 0.20 ± 0.05 mm−1 in the cerebrum and 0.27 ± 0.05 mm−1 in the cerebellum. Conclusion As conventional spectroscopic measurement may overestimate attenuation in the whole tissue, in situ measurement using the withdrawal technique might be appropriate for measurement of inhomogeneous biological tissues.

Background: An oxygen-enriched photosensitizer solution was created by the addition of red blood cells (RBCs) as an investigative tool for photosensitization reactions (PRs). Although the ...

To investigate hemolysis phenomena during a photosensitization reaction with the reaction condition continuously and simultaneously for a safety assessment of hemolysis side effect, we constructed an optical system to measure blood sample absorption spectrum during the reaction. Hemolysis degree might be under estimated in general evaluation methods because there is a constant oxygen pressure assumption in spite of oxygen depression take place. By investigating hemoglobin oxidation and oxygen desorption dynamics obtained from the contribution of the visible absorption spectrum and multiple regression analysis, both the hemolysis phenomena and its oxygen environment might be obtained with time. A 664 nm wavelength laser beam for the reaction excitation and 475-650 nm light beam for measuring the absorbance spectrum were arranged perpendicularly crossing. A quartz glass cuvette with 1×10 mm in dimensions for the spectrum measurement was located at this crossing point. A red blood cells suspension medium was arranged with low hematocrit containing 30 μg/ml talaporfin sodium. This medium was irradiated up to 40 J/cm2 . The met-hemoglobin, oxygenatedhemoglobin, and deoxygenated-hemoglobin concentrations were calculated by a multiple regression analysis from the measured spectra. We confirmed the met-hemoglobin concentration increased and oxygen saturation decreased with the irradiation time, which seems to indicate the hemolysis progression and oxygen consumption, respectively. By using our measuring system, the hemolysis progression seems to be obtained with oxygen environment information.

Background The analysis methods for fecal short-chain fatty acids (SCFAs) have evolved considerably. Recently, the role of SCFAs in gastrointestinal physiology and their association with intestinal microbiota and disease were reported. However, the intra-fecal variability and storage stability of SCFAs have not been extensively investigated. The aim of this study was to understand the limitations of the measurement of SCFAs in crude feces and develop a useful pre-examination procedure using the freeze-drying technique. Methods SCFAs in crude feces, obtained from healthy volunteers, and freeze-dried feces were determined by derivatization with isobutyl chloroformate, followed by liquid–liquid extraction with hexane, and separation and analysis using gas chromatography–mass spectrometry. Results Among the SCFAS, the maximum intra-fecal variability was observed for iso-butyrate (coefficient of variation of 37.7%), but the freeze-drying procedure reduced this variability (coefficient of variation of 7.9%). Similar improvements were also observed for other SCFAs. Furthermore, significant decreases in the SCFA amounts were observed with storage at 4 °C for 24 h. Conclusions The freeze-drying procedure affords fecal SCFA stability, even with storage at room temperature for 3 d. The freeze-drying procedure allows reliable SCFA measurements without labour-intensive processes. Therefore, the freeze-drying procedure can be applied in basic, clinical, and epidemiological studies.

To reveal hemolysis phenomena induced by a photosensitization reaction with its environment, we measured absorption spectrum of a blood sample to analyze hemoglobin oxidation and resolved oxygen desorption dynamics. The quartz glass cell with 1 mm optical path length was used as a cuvette. Red blood cell suspension medium of 0.625 hematocrit with 30 μg/ml talaporfin sodium was used as a sample. A red diode laser of 664 nm wavelength was emitted to the cuvette with 120 mW/cm2 in irradiance for 40 J/cm2. Absorption spectra of the sample were obtained before and after the photosensitization reaction by a spectrophotometer. Multiple regression analysis was employed to obtain concentrations of various hemoglobin species from measured absorption spectrum. Comparing to 0 and 40 J/cm2, methemoglobin and deoxygenated hemoglobin concentrations increased 0.19 g/dL and 0.02 g/dL, respectively. Oxygenated hemoglobin concentration decreased 0.17 g/dL. Oxygen environment could also be presented by oxygen pressure calculated from the concentrations of oxygenated hemoglobin and deoxygenated hemoglobin. These obtained hemoglobin concentration changes might indicate hemolysis progress and oxygen environment. We think this simple optical measurement could reveal both the hemolysis and oxygen environment.

In order to investigate patency of heart blood vessels by photosensitization reaction shortly after intravenous injection of talaporfin sodium, we performed in vitro endothelial cell lethality study and in vivo study of heart blood vessel patency in canine one week after photosensitization reaction. Cell lethality of human umbilical vein endothelial cells under different albumin concentrations corresponding with blood and interstice concentrations were employed and their lethality 2 hours after the reaction was measured by WST assay in vitro. Almost all cells survived by 40 J/cm2 photosensitization reaction with blood albumin concentration. Laser diffuser made of plastic optical fiber with 70 mm in length was used in vivo. Red diode laser of 664nm wavelength was emitted from this diffuser with 17.1-42.9 mW/cm in 10 minutes. We estimated the fluence rate distribution by a ray-trace simulator using pre-measured optical coefficients of myocardium tissue, μa; 0.12 mm-1 and μs’; 0.36 mm-1. Almost all blood vessels were patent in every irradiation conditions in canine heart. Coronary artery and vein up to 1 mm diameter were patent in typical myocardium sample with 25.7 mW/cm. We estimated fluence rate distribution of this sample and found that blood vessels were patent even fluence rate over 40 J/cm2. This in vivo study could be explained by the result of in vitro study. We suggest that this blood vessel patency after our particular photosensitization reaction might be because of few photosensitizer uptake in the blood endothelial cells and/or reduced oxidation damage by thick albumin concentration in blood.

s / Photodiagnosis and Photodynamic Therapy 12 (2015) 325–375 351 for low and high light dose, respectively for Vero and HEp-2. The results suggest that HY-G may be as potent photosensitizer as HY with the advantage of being more soluble. Acknowledgements: CEPOF, CAPES, FAPESP. http://dx.doi.org/10.1016/j.pdpdt.2015.07.102 Cellular distribution and PDT response of chlorin based photosensitizer in murine melanoma Bruno A. Ono, Sebastiao Pratavieira, Cristina Kurachi Sao Carlos Institute of Physics, University of Sao