In Situ Formation of a Relatively Transparent Ion-Associate Liquid Phase from an Aqueous Phase and Its Application to Microextraction/High-Performance Liquid Chromatography-Fluorescence Detection of Bisphenol A in Water.

The design of a simple approach enabling the detection of bisphenol A (BPA) in water samples without the need for large amounts of solvents is of utmost importance. This paper reports a simple method for the separation, concentration, and quantification of BPA in water samples using high-performance liquid chromatography with fluorescence detection (HPLC-FLD) after its microextraction into an in situ formed organic ion-associate (IA) liquid phase (LP). Novel IA phase components without conjugated double bonds, such as benzene rings, were investigated. Ethylhexyloxypropylamine hydrochloride and sodium dodecyl sulfate solutions were added to the water samples to form IAs. The aqueous phase and ion-associate liquid phase (IALP) were separated by centrifugation. The aqueous phase was removed, and the liquid phase was recovered and measured using HPLC-FLD or HPLC-electrochemical detection (ECD). The concentrated phase (IALP) had a relatively low viscosity and could be injected directly into the chromatograph without dissolving it in organic solvents. The detection limits for BPA by HPLC-FLD and HPLC-ECD were 0.009 and 0.3 µg L-1, respectively.

Effects of paddy irrigation-drainage system on water quality and productivity of small rivers in the Himi region of Toyama, Central Japan.

The aim of this study was to clarify the impact of differences between historical and recently introduced irrigation and drainage management systems on water quality in the rivers around paddy fields. We investigated the seasonal variation in nutrients concentration and dissolved organic carbon (DOC) components in single- (used for intake only) and dual-purpose (used for both intake and drainage) channels during a 4-year period in the Himi region of Toyama, Central Japan. The system of dual-purpose channel has traditionally been used in the region of this study. A total of 197 three-dimensional excitation-emission matrix (3DEEM) fluorescence spectra of DOM in waters were applied for the parallel factor analysis (PARAFAC) modeling. Based on the 3DEEM and PARAFAC, the abundance of terrestrial humic-like components in the dual-purpose channel was significantly higher than that in the single-purpose channel. The even long-chain n-fatty acids derived associated with rice cropping in sediments of the dual-purpose channels were 22-30-fold higher than that of the single-purpose channel. In addition, the turbidity values of the river waters had significantly positive linear correlations with concentrations of K+, DOC, and humic-like components. These observations indicate that the dissolved nutrient concentrations in the river water were higher in the dual-purpose channel compared to those of the single-purpose channel, which may be supplied by leaching from the inflow of soil particles from the paddy fields. During the mid-irrigation period, the quantity of epiphytic chlorophyll a on artificial substrate tiles in the dual-purpose channel were 3.1-4.1-fold higher than that in the single-purpose channel. This study clear that the input of paddy drainage during the irrigation season significantly changes the DOC components in river waters and irrigation management is strongly linked to the primary production in agricultural channels. Therefore, it is important to consider the impact of the introduction of different irrigation and drainage management systems on water quality and productivity in order to maintain the riverine ecosystems around rice paddies, which are based on historical water use systems.

Simple solid-phase colorimetry for trace Cr(VI) by combination of complexation with diphenylcarbazide and ion-pair solid-phase extraction with sedimentable dispersed particulates.

A highly sensitive and simple solid-phase colorimetry for Cr(VI) was proposed. It was based on the ion-pair solid-phase extraction of Cr-diphenylcarbazide (DPC) complex with sedimentable dispersed particulates. The concentration of Cr(VI) was measured from the color tones obtained by image analysis of the photo of sediment. Various conditions, e.g., material and amounts of adsorbent particulates, chemical properties and concentration of counter ions, and pH, were optimized for the formation and quantitative extraction of the complex. In the recommended procedure, 1 mL of sample was put into a 1.5 mL microtube where powder form adsorbent and reagents, i.e., XAD-7HP particles, DPC, sodium dodecyl sulfate, amido sulfuric acid, and sodium chloride had been packed. The analytical operation was completed within 5 min by gently shaking the microtube and allowing it to stand until enough amounts of particulates were deposited to take a picture. Chromium (VI) up to 2.0 ppm was determined, and the detection limit was 0.0034 ppm. The sensitivity was enough to determine Cr(VI) at lower concentrations than the water quality of standard (0.02 ppm). This method was successfully applied to the analysis of simulated industrial wastewater samples. The stoichiometry of the extracted chemical species was also investigated by applying the same equilibrium model as the ion-pair solvent extraction.

Removal of humic acid interference in soil enzymatic analysis using poly-γ-glutamic acid.

Soil enzymes are biological indicators in environmental and agricultural monitoring. However, brownish humic acid (HA) in samples interferes significantly with various analytical methods, especially in optical-based techniques. Here, we implemented a coagulation-flocculation process to carry out continuously an enzymatic reaction without separation and transfer of a sample solution. The elimination of HA in a soil suspension using poly-γ-glutamic acid (PGA) by coagulation to minimize the HA interference in soil enzymatic analysis was investigated. As a result of the optimization of preliminary parameters, the removal efficiency of HA was > 92% in 100 mg L-1 HA in neutral pH, using 100 mg L-1 PGA and aluminum trivalent as a coagulant aid. However, the fluorescent intensity of the enzyme product (i.e., 4-methylumbelliferone) decreases by about 50% as HA was removed under the conditions used. A decrease in the enzymatic detection of 3,3',5,5'-tetramethylbenzidine (TMB) was not observed from treated samples even though the initial level of HA was different. The results suggested that the coagulation-flocculation approach is suitable for the reduction of HA interference, while maintaining target analyte detection. Therefore, the proposed sample treatment can be used to examine enzyme activity based on TMB product detection without regular standard addition calibration.

Colorimetric analysis based on solid-phase extraction with sedimentable dispersed particulates: demonstration of concept and application for on-site environmental water analysis.

A novel simple and functional colorimetric methodology for on-site environmental water analysis was proposed. This method combines coloration of the analyte and extraction of the colored species on dispersed particulates during their sedimentation in the same container. The whole analysis can be performed within 15 min by comprising the addition of 1 mL of sample solution into a 1.5-mL microtube containing the powders of coloring reagents and the sedimentable fine particulates as an adsorbent. The analyte is determined by comparing the sediment color with the standard color by visual inspection or the color information of the photo image. The potential of this methodology was demonstrated through developing colorimetry for Fe2+ with o-phenanthroline, NO2- by azo-dye formation, HCHO by the MBTH method, and PO43- by the 4-aminoantipyrine method based on the enzyme reactions. The material, size, amount of the adsorbent particles, and other conditions were optimized for each analytes. The advantages of the methodology were as follows: high sensitivity, easy controllability of the sensitivity over the wide range by the amount, size, and material of the particulates, lower interference from the colored matrix components due to obtaining the color data from not the aqueous phase but the sedimented particulates, and acceleration of the color development rate by the particulates as seen in NO2- determination as consequence shorten the operation time. A simple device equipped with twin cells was proposed for on-site analysis which contains two successive different coloring operations. The developed methods were successfully applied to the environmental water samples with the good agreement of the results with those by the usual instrumental methods.

Investigation and modeling of diurnal variation in suburban ambient formaldehyde concentration.

Formaldehyde (HCHO) is a naturally occurring compound found in ambient air which can induce cancer and sick-building syndrome. It plays an important role in the formation of OH radicals, which are connected to the formation of various airborne chemicals. Herein, we present a simple modeling for the simulation of diurnal variations in the HCHO concentration of ambient air. This was achieved using data collected during different seasons from November 2015 to March 2017 at a suburban location in Toyama City (Japan), where non-methane hydrocarbon (NMHC) levels were low at sub carbon ppm (ppmC) order. The modeling was based on the assumption that photochemical reactions of methane were the major factor of secondary HCHO formation. The model took into account the production and decomposition of HCHO by photochemical reactions as well as its loss due to other reactions such as dry deposition. Accordingly, the model's equation contained terms for solar radiation, temperature, and methane concentration. The results predicted using the model showed good agreement with the experimental data observed on fine days, i.e., except rainy, foggy, and heavily cloudy days. The relationships between HCHO concentration and solar radiation/temperature on different days as well as the seasonal variation of HCHO concentration were also interpreted by the proposed model. This study contributes to the evaluation of the pollution levels of formaldehyde. Moreover, the model may be used to demonstrate the impact of increasing methane levels, with regard to global warming and the background levels of HCHO in the atmosphere.

Effect of humic acids on the toxicity of pollutants to Chlamydomonas reinhardtii: Investigation by a microscale algal growth inhibition test.

Dissolved humic substances (DHSs) are the major components of organic matter in the aquatic environment. DHSs are well known to considerably affect the speciation, solubility, and toxicity of a wide variety of pollutants in the aquatic environment. In this study, the effects of the toxicity of heavy metals and hydrophobic organic pollutants (HOPs) on Chlamydomonas reinhardtii in the presence of humic acid (HA) were examined by a microscale algal growth inhibition (µ-AGI) test based on spectrophotometric detection. To clarify the relationship between the chemical properties of HAs and the toxicity change of pollutants, eight HAs from different sources were prepared and used. HAs were responsible for mitigating the toxicity of Hg, Cu, pesticides (γ-HCH, 2,4-D, and DDT), and polycyclic aromatic hydrocarbons (PAHs) such as naphthalene (Nap), anthracene (Ant), and benzo[a]pyrene (BaP). In particular, an approximately 100-fold decrease in the toxicity of BaP was observed in the presence of 10 ppm HAs extracted from tropical peat. The results indicated that the carboxylic group content and the HA molecular weight are correlated to the changes in the heavy metal toxicity. For HOPs, the aromaticity and polarity of HAs are crucial for mitigating their toxicity. Furthermore, it was clearly shown that the lake water including a high concentration of DHSs collected from Central Kalimantan, Indonesia, reduced the toxicity of Hg and γ-HCH on Chlamydomonas reinhardtii. Graphical abstract.

High-heat Effects on the Physical and Chemical Properties of Soil Organic Matter and Its Water-soluble Components in Japan's Forests: A Comprehensive Approach Using Multiple Analytical Methods.

Wildfires that expose the soil organic layer to high heat levels can alter soil organic matter (SOM), which includes water-soluble organic matter (WSOM) components. Various evaluation methods were used to characterize and quantify the effects of high heat levels on SOM and WSOM, including ion chromatography, thermogravimetry-differential thermal analysis (TG-DTA), colorimetry, elemental analysis, pyrolysis-gas chromatography-mass spectrometry using tetramethylammonium hydroxide (TMAH-py-GC/MS), total organic carbon (TOC) analysis, three-dimensional excitation-emission matrix (3DEEM) spectroscopy, and high-performance size-exclusion chromatography. In this study, we applied each of these evaluation methods using soil samples that were collected from broadleaf, coniferous, and bamboo forests and peatland in Japan and exposed to different initial high heat levels. Based on the TG-DTA results, the remaining mass in select soil samples markedly decreased when reheated to approximately 200°C. Comparatively, the TMAH-py-GC/MS results indicated a drastic change in SOM composition and the production of low molecular organic components (

Organic Ion-associate Phase Extraction/Back-microextraction for the Preconcentration and Determination of Lithium Using 2,2,6,6-Tetramethyl-3,5-heptanedione by Liquid Electrode Plasma Atomic Emission Spectrometry and GF-AAS in Environmental Water.

We developed an ion-associate phase (IAP)-extraction/acid back-extraction system for the preconcentration and atomic spectrometric determination of lithium trace amounts in water. The chelating reagent for lithium also works as a constituent of the extraction phase. The lithium in a 10 mL sample solution was converted through a chelate complex reaction with 2,2,6,6-tetramethyl-3,5-heptanedione (HDPM). The addition of a benzyldimethyltetradecylammonium ion caused the formation of IAP suspension in the solution. Centrifugation of the solution led to the isolation of a liquid organic phase and the lithium complex was extracted as the upper phase from the centrifuge tube. After the aqueous phase was removed, lithium was back-extracted with a 400 µL nitric acid solution from the IAP. The acid phase was measured using liquid-electrode-plasma atomic-emission-spectrometry (LEP-AES) or graphite-furnace atomic-absorption spectroscopy (GF-AAS). The detection limits were 0.02 mg/L for LEP-AES and 0.02 µg/L for GF-AAS. This system was applied to the determination of environmental water. The HDPM in the organic phase was reusable.

Effects of forest fire on the properties of soil and humic substances extracted from forest soil in Gunma, Japan.

Increases in global wildfires and fire severity are expected to result from global warming. Severe wildfires not only burn surface vegetation but also affect forest soil. Humic substances play key roles in the transport of nutrients and the carbon cycle in terrestrial ecosystems. In this study, we evaluated the effects of forest fires on the chemical properties of fulvic acid (FA) and humic acid (HA) extracted from non-burned and burned forest soils in Gunma, Japan. The differential thermal analysis of FA indicated that the intensity of exothermic reaction peak at 400 °C was 2-fold higher than that from non-burned soil. Based on pyrolysis-gas chromatography-mass spectrometry analysis with tetramethyl ammonium hydroxide, the amount of pyrolysate compounds in FA from burnt soil was significantly lower than that in FA from non-burnt soil. Therefore, we can conclude that the forest fire caused the significant change in the properties of FA such as increasing the aromaticity and refractory. In addition, the concentration of dissolved organic carbon with low molecular weight in surface soil increased after forest fire. This study suggests that the denaturation of soil organic matter by wildfire can affect the carbon cycle in terrestrial ecosystems.

Potential risk of coupling products between tetrahalobisphenol A and humic acid prepared via oxidation with a biomimetic catalyst.

Tetrahalobisphenol A (TXPBAs, X = Br or Cl), TBBPA and TCBPA, which are widely used as flame retardants, ultimately disposed of in landfills. In landfills, enzymatically oxidized TXBPAs can be covalently incorporated into humic acids (HAs) to form coupling products (HA-TXBPAs). In the present study, HA-TXBPAs were prepared by catalytic oxidation with iron(III)-phthalocyanine-tetrasulfate as a model of oxidative enzymes. The stability of HA-TXBPAs was evaluated by incubating them under physicochemical conditions of landfills (pH 9 and 50 °C). For HA-TBBPA, 18-26% of TBBPA was released from HA-TBBPA, due to the acid dissociation of the loosely bound TBBPA. However, no additional release was observed, even after 30 days, indicating that 74-82% of the TBBPA was incorporated into the HA. For HA-TCBPA, 3-4% of TCBPA and a major byproduct, 4-(2-hydroxyisopropyl)-2,6-dichlorophenol, was found to be loosely incorporated into HA. For both TBBPA and TCBPA, covalently bound organo-halogens were not released during the 30 days of incubation. Inhibition of the growth of Chlamydomonas reinhardtii was indicated when trace levels of TXBPAs (approximately 0.1 µM) were present. These results suggest that HA-TXBPAs contain not only covalently incorporated TXBPAs but also loosely bound TXBPAs and halophenols. The latter in HA-TXBPAs have the potential to leach from landfills and affect aquatic ecosystems.

Determination of heavy metal toxicity by using a micro-droplet hydrodynamic voltammetry for microalgal bioassay based on alkaline phosphatase.

We developed an electrochemical microalgal bioassay for the determination of heavy metal toxicity in water on the basis of the alkaline phosphatase (ALP) enzyme inhibition of Chlamydomonas reinhardtii. Five heavy metals were chosen as toxicants: Hg, Cd, Pb, Zn, and Cu. The induced ALP activity of C. reinhardtii was inhibited using the phosphate starvation method, and the results were evaluated by measuring the electrochemical oxidation of p-aminophenol (PAP) following the enzymatic conversion of p-aminophenyl phosphate (PAPP) as a substrate. The rapid determination of enzymatic activity was achieved using hydrodynamic voltammetry in a 50 µL micro-droplet with a rotating disk electrode (RDE). Enzymatic activity over a PAPP substrate is affected by heavy metal ions, and this phenomenon decreases the chronoamperometric current signal. The concentrations of Hg, Cd, Pb, Zn, and Cu in which the ALP activity was half that of the control (EC50) were found to be 0.017, 0.021, 0.27, 1.30, and 1.36 µM, respectively. The RDE system was demonstrated to be capable of detecting enzymatic activity by using a small amount of regent, a reaction time of only 60 s, and a detection limit of 5.4 × 10-7 U.

Hydrodynamic voltammetry as a rapid and simple method for evaluating soil enzyme activities.

Soil enzymes play essential roles in catalyzing reactions necessary for nutrient cycling in the biosphere. They are also sensitive indicators of ecosystem stress, therefore their evaluation is very important in assessing soil health and quality. The standard soil enzyme assay method based on spectroscopic detection is a complicated operation that requires the removal of soil particles. The purpose of this study was to develop a new soil enzyme assay based on hydrodynamic electrochemical detection using a rotating disk electrode in a microliter droplet. The activities of enzymes were determined by measuring the electrochemical oxidation of p-aminophenol (PAP), following the enzymatic conversion of substrate-conjugated PAP. The calibration curves of ß-galactosidase (ß-gal), ß-glucosidase (ß-glu) and acid phosphatase (AcP) showed good linear correlation after being spiked in soils using chronoamperometry. We also performed electrochemical detection using real soils. Hydrodynamic chronoamperometry can be used to assess the AcP in soils, with a detection time of only 90 s. Linear sweep voltammetry was used to measure the amount of PAP released from ß-gal and ß-glu by enzymatic reaction after 60 min. For the assessment of soil enzymes, the results of hydrodynamic voltammetry assay compared favorably to those using a standard assay procedure, but this new procedure is more user-friendly, rapid and simple.

Effects of peat fires on the characteristics of humic acid extracted from peat soil in Central Kalimantan, Indonesia.

When peat forest fires happen, it leads to burn soil and also humic acids as a dominant organic matter contained in peat soil as well as the forest. The structure and properties of humic acids vary depending on their origin and environment, therefore the transformation of humic acid is also diverse. The impacts of the peat fires on peat soil from Central Kalimantan, Indonesia were investigated through the characterization of humic acids, extracted from soil in burnt and unburnt sites. The characterization of humic acids was performed by elemental composition, functional groups, molecular weight by HPSEC, pyrolysate compounds by pyrolysis-GC/MS, fluorescence spectrum by 3DEEM spectrofluorometer, and thermogravimetry. The elemental composition of each humic substance indicated that the value of H/C and O/C of humic acids from burnt sites were lower than that from unburnt sites. The molecular weight of humic acids from burnt sites was also lower than that from unburnt sites. Pyrolysate compounds of humic acids from unburnt sites differed from those of humic acids from burnt soil. The heating experiment showed that burning process caused the significant change in the properties of humic acids such as increasing the aromaticity and decreasing the molecular weight.

Electrochemical genotoxicity assay based on a SOS/umu test using hydrodynamic voltammetry in a droplet.

The SOS/umu genotoxicity assay evaluates the primary DNA damage caused by chemicals from the ß-galactosidase activity of S. typhimurium. One of the weaknesses of the common umu test system based on spectrophotometric detection is that it is unable to measure samples containing a high concentration of colored dissolved organic matters, sediment, and suspended solids. However, umu tests with electrochemical detection techniques prove to be a better strategy because it causes less interference, enables the analysis of turbid samples and allows detection even in small volumes without loss of sensitivity. Based on this understanding, we aim to develop a new umu test system with hydrodynamic chronoamperometry using a rotating disk electrode (RDE) in a microliter droplet. PAPG when used as a substrate is not electroactive at the potential at which PAP is oxidized to p-quinone imine (PQI), so the current response of chronoamperometry resulting from the oxidation of PAP to PQI is directly proportional to the enzymatic activity of S. typhimurium. This was achieved by performing genotoxicity tests for 2-(2-furyl)-3-(5-nitro-2-furyl)-acrylamide (AF-2) and 2-aminoanthracene (2-AA) as model genotoxic compounds. The results obtained in this study indicated that the signal detection in the genotoxicity assay based on hydrodynamic voltammetry was less influenced by the presence of colored components and sediment particles in the samples when compared to the usual colorimetric signal detection. The influence caused by the presence of humic acids (HAs) and artificial sediment on the genotoxic property of selected model compounds such as 4-nitroquinoline-N-oxide (4-NQO), 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX), 1,8-dinitropyrene (1,8-DNP) and 1-nitropyrene (1-NP) were also investigated. The results showed that the genotoxicity of 1-NP and MX changed in the presence of 10 mg∙L-1 HAs. The genotoxicity of tested chemicals with a high hydrophobicity such as 1,8-DNP and 1-NP were decreased substantially with the presence of 1 g∙L-1 sediment. This was not observed in the case of genotoxins with a low log K(ow) value.

The evaluation for alterations of DOM components from upstream to downstream flow of rivers in Toyama (Japan) using three-dimensional excitation-emission matrix fluorescence spectroscopy.

The dissolved organic matter (DOM) is one of the important factors for controlling water quality. The behavior and constitutions of DOM is related to the risk of human health because it is able to directly or indirectly affect the behavior, speciation and toxicity of various environmental pollutants. However, it is not easy to know the contents of DOM components without using various complicated and time consuming analytical methods because DOM is a complex mixture and usually exists at low concentration. Here, we describe the fluorescence properties of DOM components in water samples collected from four rivers in Toyama, Japan by means of the three-dimensional excitation-emission matrix (3DEEM) fluorescence spectroscopy. In order to evaluate the alterations of DOM components in each of the river during the flow from upstream to downstream, the patterns of relative fluorescence intensity (RFI) at six peaks which are originated from fluorophores including humic-like and protein-like components were investigated. The changes in the patterns of RFI values at each of the peak and the concentration of dissolved organic carbon (DOC) for each river water sample were discussed in connection with the differences of land use managements and basic water quality parameters, such as pH, EC, turbidity, Fe(3+), T-N, NO(3)-N, T-P, PO(4)-P, chlorophyll a, DOC and N/P ratio. The DOC concentrations in the water samples collected from these rivers were relatively low (0.63-1.16 mg/L). Two main peaks which have a strong RFI value expressed a positive correlation with the DOC concentration (r = 0.557, 0.535). However, the correlations between the RFI values for other four peaks and the DOC concentration were below 0.287. The alterations of DOM components during the flow of a river from upstream to downstream were investigated from the changes in the patterns of RFI values for six fluorescent peaks. It was clarified that the great increase of RFI values in peak A and peak T from river water located in urban area showed high concentration of PO(4)-P and Fe(3+), and low N/P ratio due to the high biological activities. The values of fluorescence index (FIX) and biological index (BIX) were as high as 1.60 and 0.72, respectively.