Toxicity of Aged Paint Particles to Soil Ecosystems: Insights from Caenorhabditis elegans.

Despite the extensive global consumption of architectural paint, the toxicological effects of aged exterior paint particles on terrestrial biota remain largely uncharacterized. Herein, we assessed the toxic effect of aged paint particles on soil environments using the nematode Caenorhabditis elegans (C. elegans) as a test organism. Various types of paint particles were generated by fragmentation and sequential sieving (500-1000, 250-500, 100-250, 50-100, 20-50 µm) of paint coatings collected from two old residential areas. The paint particles exerted different levels of toxicity, as indicated by a reduction in the number of C. elegans offspring, depending on their size, color, and layer structure. These physical characteristics were found to be closely associated with the chemical heterogeneity of additives present in the paint particles. Since the paint particle sizes were larger than what C. elegans typically consume, we attributed the toxicity to leachable additives present in the paint particles. To assess the toxicity of these leachable additives, we performed sequential washings of the paint particles with distilled water and ethanol. Ethanol washing of the paint particles significantly reduced the soil toxicity of the hydrophobic additives, indicating their potential environmental risk. Liquid chromatography-mass spectrometry analysis of the ethanol leachate revealed the presence of alkyl amines, which exhibited a high correlation with the toxicity of the paint particles. Further toxicity testing using an alkyl amine standard demonstrated that a paint particle concentration of 1.2% in soil could significantly reduce the number of C. elegans offspring. Our findings provide insights into the potential hazards posed by aged paint particles and their leachable additives in the terrestrial environment.

Improving liquid chromatography-mass spectrometry sensitivity for characterization of lignin oligomers and phenolic compounds using acetic acid as a mobile phase additive.

Characterization of complex lignin degradation products by liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) requires a high sensitivity, which can be improved by use of optimized mobile phase additive. In this study, chromatographic separation and ionization efficiency for LC-MS analysis of lignin oligomers and lignin-derived phenolic compounds were compared among six different mobile phase additives. We found that acetic acid exhibits analytical performance superior to the other mobile phase additives for characterization of lignin degradation products by LC-(-)-ESI-MS/MS. The retention of phenolic compounds (PCs) containing carboxylic group was improved by using acidic additives instead of their ammonium salts. The use of acetic acid as an LC mobile phase additive enhanced the ionization efficiency of PCs in (-)-ESI-MS about 2 to 4 times higher than the other additives. The improved MS sensitivity with acetic acid yielded a higher coverage of fragment ions in LC-MS/MS analysis, leading to identification of four additional synthetic G-type lignin oligomers (LOs) than ammonium acetate. A similar degree of sensitivity enhancement was also demonstrated for degradation products from rice straw lignin using acetic acid additive over the others. The increased chromatographic retention and enhanced sensitivity of PCs and LOs provided by the use of acetic acid as an LC-(-)-ESI-MS mobile phase additive will facilitate structural analysis of lignin degradation products without relying on additional instrumentation.

Synthesis of Alternating Polyisocyanate Copolymers by Anionic Polymerization for Mimicking Amphiphilic Helical Peptides.

The amphiphilic conformation of α-helical peptides has important biological functions, such as ion transport, antifreeze, and innate immunity, which can be mimicked by alternating polyisocyanate copolymers. We synthesized poly(allyl isocyanate-alt-(S)-(-)-α-methylbenzyl isocyanate (P(AIC-alt-SMBIC)) and ammonium-containing P(AIC-alt-SMBIC) (N-P(AIC-alt-SMBIC)), ensuring the amphiphilic helical conformation. The benzyl group of SMBIC plays an important role in alternating copolymerization with its steric and electron-withdrawing effects, while AIC provides an alkene group capable of introducing a customized functional group. The P(AIC-alt-SMBIC) with predominantly alternating sequence was acquired at fSMBIC /fAIC =8 with a controlled molecular weight and narrow dispersity. N-P(AIC-alt-SMBIC)s were synthesized from thiol-ene radical addition with P(AIC-alt-SMBIC).

Characterization of Spray Modes and Factors Affecting the Ionization Efficiency of Paper Spray Ionization.

In this study, we systematically evaluated the factors affecting the ionization efficiency of paper spray ionization (PSI), such as electric field, solvent supply rate, and paper thickness and hydrophobicity. The observed paper spray plume was classified into three modes: single cone-jet, multi-jet, and rim-jet modes. With the increase in the spraying voltage, the spray plume appeared in order of single cone-jet, multi-jet, and rim-jet modes. The rim-jet mode exhibited the lowest standard deviation and high ionization efficiency among the three spray modes. The main parameter determining the spray mode was the charge density of the droplets generated by paper spray, which depends on the electric field and solvent supply rate. A thicker paper reduced the electric repulsion between the jets and lowered the threshold voltage to reach the rim-jet mode. Lowering the solvent supply rate caused mode transitions from the single cone-jet to the rim-jet, possibly due to the increased droplet charge density. The hydrophobic modification on a paper substrate led to a different ionization mechanism or electrostatic spray ionization at low applied voltages.

Fluorescent paper strip immunoassay with carbon nanodots@silica for determination of human serum amyloid A1.

A fluorescent paper strip immunoassay in conjunction with carbon nanodots@silica (CND@SiO2) as a label was developed for the quantitative measurements of human serum amyloid A1 (hSAA1) in serum at clinically significant concentrations for lung cancer diagnosis. Monodispersed CND@SiO2 was prepared by cohydrolysis between silane-crosslinked carbon nanodots and silica precursors via the Ströber method and further attached covalently to anti-hSAA1 (14F8) monoclonal antibody [anti-hSAA1(14F8)] specific to the hSAA1 target. The hSAA1 concentrations were then determined by quantifying the blue fluorescence intensity upon 365 nm excitation of the captured hSAA1 with anti-hSAA1(14F8)-CND@SiO2 conjugates in the test line on a paper strip where anti-hSAA1 (10G1) monoclonal antibody was physisorbed. The developed fluorescent paper strip with CND@SiO2 can detect hSAA1 at concentrations ranging from 0.1 to 5 nM (R2 = 0.995), with a limit of detection of  0.258 nM in 10 mM phosphate buffer pH 7.4 containing human serum albumin. The performance of  recovery (90.98-109.17%) and repeatability (coefficients of variation < 8.46%) obtained was also acceptable for quantitative determinations. The platform was employed for direct determination of hSAA1 concentrations in undiluted serum samples from lung cancer patients (relative standard deviation (RSD) < 7.46%) and healthy humans (RSD < 3.96%). The results were compared with those obtained using a commercially available enzyme-linked immunosorbent assay alongside liquid chromatography with tandem mass spectrometry measurements.

Development of an optimized sample preparation method for quantification of free fatty acids in food using liquid chromatography-mass spectrometry.

Accurate and precise determination of free fatty acid (FFA) contents is essential for quality control and assurance in food production. Herein, a mass spectrometric study was performed to develop a sample preparation protocol that can minimize exogenous FFA contamination during the quantification of FFA in food. The quantities of exogenous FFAs were measured using various combinations of seven pretreatment methods for a sample tube, three extraction methods, and four types of sample tubes. Methanol washing could effectively reduce exogenous palmitic acid (PA) and stearic acid (SA) by 73 and 64%, respectively, in contrast to furnace baking resulting in a decrease in the amount of PA and SA contaminants by 50 and 37%, respectively. A lower amount of FFA contaminants was extracted from glass tubes during comparative analysis of the four different sample tubes studied. A combination of a methanol-washed glass tube and chloroform extraction solvent was determined to be the optimal method for minimizing the error in FFA quantification. The optimized sample preparation protocol used for FFA quantification can lower the amount of foreign palmitic acid and stearic acid to the sub-nanomolar level in the analysis of FFAs in skimmed milk.

Microfluidic ultrafine particle dosimeter using an electrical detection method with a machine-learning-aided algorithm for real-time monitoring of particle density and size distribution.

Growing concerns related to the adverse health effects of airborne ultrafine particles (UFPs; particles smaller than 300 nm) have highlighted the need for field-portable, cost-efficient, real-time UFP dosimeters to monitor individual exposure. These dosimeters must measure both the particle density and size distribution as these parameters are essential to the determination of where and how many UFPs will be deposited in human lungs. However, though various kinds of laboratory-grade instruments and hand-held monitors have been developed, they are expensive and only capable of measuring particle size distribution. A microfluidic UFP dosimeter is proposed in this study to address these limitations. The proposed sensor, based on an electrical detection method with a machine-learning-aided algorithm, can simultaneously measure the size distribution (number concentration, mean mobility diameter, geometric standard deviation) and particle density, and is compact owing to the microelectromechanical systems (MEMS) technology. In a comparison test using physically synthesised Ag and di-ethyl-hexyl sebacate (DEHS) aerosols, the mean measurement errors of the proposed sensor compared to the reference system were 6.1%, 4.5%, and 7.3% for number concentration, mean mobility diameter, and particle density, respectively. Moreover, when the machine-learning aided algorithm was operated, the geometric standard deviation could be deduced with a 7.6% difference. These results indicate that the proposed device can be successfully used as a field-portable UFP sensor to assess individual exposure, an on-site monitor for ambient air pollution, an analysis tool in toxicological studies of inhaled particles, for quality assurance of nanomaterials engineered via aerosol synthesis, etc.

Monitoring the Effective Density of Airborne Nanoparticles in Real Time Using a Microfluidic Nanoparticle Analysis Chip.

Determining the effective density of airborne nanoparticles (NPs; particles smaller than 100 nm in diameter) at a point of interest is essential for toxicology and environmental studies, but it currently requires complex analysis systems comprising several high-precision instruments as well as a specially trained operator. To address these limitations, a field-portable and cost-efficient microfluidic NP analysis device is presented, which provides quantitative information on the effective density and size distribution of NPs in real time. Unlike conventional analysis systems, the device can operate in a standalone mode because of the chip operating principle based on the electrostatic/inertial classification and electrical detection methods. Moreover, the device is both compact (16.0 × 10.9 × 8.6 cm3) and light (950 g) owing to the hardware strip down enabled by integrating the essential functions for effective density analysis on a single chip. Quantitative experiments performed to simulate real-life applications utilizing effective density (i.e., effective density-based morphology analysis on engineered NPs and multi-parametric NP monitoring in ambient air) demonstrate that the developed device can be used as an analysis tool in toxicological studies as an on-site sensor for the monitoring of individual NP exposure and environments, for quality monitoring of engineered NPs via aerosol synthesis, and other applications.

A novel pathway for initial biotransformation of dinitroaniline herbicide butralin from a newly isolated bacterium Sphingopyxis sp. strain HMH.

Butralin (N-sec- Butyl-4-tert-butyl-2,6-dinitroaniline) is a highly persistent dinitroaniline herbicide frequently detected in the environment. In this study, butralin-degrading soil bacterium, Sphingopyxis sp. strain HMH was isolated from agricultural soil samples. Based on whole genome sequence analysis of the strain HMH, the gene encoding a nitroreductase NfnB was identified and expressed in Escherichia coli (E. coli), and protein was purified to homogeneity. NfnB is a flavin-nitroreductase, found to be a functional tetramer, composed of subunit molecular mass of 25 kDa. The metabolites from butralin degradation by strain HMH and purified NfnB were identified using ultra performance liquid chromatography high resolution mass spectrometry (UPLC-HRMS), and a novel mechanism of butralin degradation was proposed. NfnB selectively nitro-reduced butralin into N- (sec-Butyl)-4-(tert-butyl)-6-nitrobenzene- 1,2-diamine, followed by formation of 5-(tert-Butyl)-3 -nitrobenzene-1,2-diamine and butanone by N- dealkylation through possible hydroxylation reaction onto the carbon linked amine of the N-(sec-Butyl) moiety. In our study, we could not detect the hydroxylated product 2-(2-Amino-4-tert-butyl-6-nitro- phenylamino)-butan-2-ol) (carbinolamine), instead its Schiff base product (E)-2-(Butan-2-yildeneamino)-5- (tert-butyl)-3-nitroaniline was detected. The release of butanone was further confirmed by derivatization with 2,4- dinitrophenylhydrazine (DNPH) followed by MS analysis. In conclusion, this study explores a novel multi-functional flavin- nitroreductase family enzyme NfnB, catalyzing unique and sequential nitroreduction and N-dealkylation through oxidative hydroxylation of dinitroaniline herbicide butralin.

Physiological responses of the abalone Haliotis discus hannai to daily and seasonal temperature variations.

Organisms inhabiting tidal mixing-front zones in shallow temperate seas are subjected to large semidiurnal temperature fluctuations in summer. The ability to optimize energy acquisition to this episodic thermal oscillation may determine the survival, growth and development of these ectotherms. We compared the physiological and molecular responses of Haliotis discus hannai cultivated in suspended cages to fluctuating or stable temperature conditions. Several physiological indicators (respiration, excretion rates and O:N) were measured in both conditions, and alterations in the proteome during thermal fluctuations were assessed. No summer mortality was observed in abalone cultivated in fluctuating temperatures compared with that at stable high temperatures. Metabolic rates increased sharply during stable warm summer conditions and fluctuated in accordance with short-term temperature fluctuations (20-26 °C). Ammonia excretion rates during acute responses were comparable in both conditions. When abalone were exposed to fluctuating temperatures, enzyme activities were downregulated and structure-related protein expression was upregulated compared with that at an acclimation temperature (26 °C), highlighting that exposure to low temperatures during fluctuations alters molecular processes. Our results reveal that modulation of physiological traits and protein expression during semidiurnal thermal fluctuations may buffer abalone from the lethal consequences of extreme temperatures in summer.

Thermoalkaliphilic laccase treatment for enhanced production of high-value benzaldehyde chemicals from lignin.

Enzymatic conversion of lignin into high-value chemicals is a key step in sustainable and eco-friendly development of lignin valorization strategies. In the present study, a novel thermoalkaliphilic laccase, CtLac, from Caldalkalibacillus thermarum strain TA2.A1 was tested for the depolymerization of lignin and the production of value-added chemicals, using three different lignocellulosic biomass, organosolv lignin (OSL), and Kraft lignin. Seven valuable lignin monomers were identified from the CtLac-treated samples using high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). Remarkably, increases of 22.0%, 65.6%, and 27.3% of p-hydroxybenzaldehyde and increases of 111.1%, 93.5%, and 238.1% of vanillin were observed from rice straw, corn stover, and reed, respectively. Comparative analysis of lignin monomers released from rice straw, using Trametes versicolor laccase (TvL) and CtLac indicated efficient depolymerization of lignin by CtLac. CtLac treatment resulted in 2.3 fold and 5.6 fold, and 1.9 fold and 2.8 fold higher amounts of p-hydroxybenzaldehyde and vanillin from OSL and Kraft lignin, respectively, compared to CtLac-treated rice straw samples after 12 h reaction. OSL was the best substrate for the production of benzaldehyde chemicals using CtLac treatment. The results demonstrated potential application of bacterial laccase CtLac for valorization of biomass lignin into high-value benzaldehyde chemicals under thermoalkaliphilic conditions.

A novel laccase from thermoalkaliphilic bacterium Caldalkalibacillus thermarum strain TA2.A1 able to catalyze dimerization of a lignin model compound.

In the present study, the gene encoding a multicopper oxidase, more precisely a laccase from the thermoalkaliphilic aerobic bacterium Caldalkalibacillus thermarum strain TA2.A1 (CtLac), was cloned and expressed in Escherichia coli. CtLac is a monomeric protein with a molecular weight of 57 kDa as determined by native polyacrylamide gel electrophoresis. The optimum pH and temperature for 2,6-dimethoxyphenol (2,6-DMP) oxidation were 8.0 and 70 °C, respectively. The kinetic constants Km and kcat for 2,6-DMP were of 200 µM and 23 s-1, respectively. The enzyme was highly thermostable at 80 °C and retained more than 80% of its activity after 24 h preincubation under thermoalkaliphilic conditions. Remarkably, it showed a half-life of about 12 h at 90 °C. The enzyme activity was significantly enhanced by Cu2+ and Mn2+ and was not affected in the presence of most of the other metal ions. CtLac activity was stimulated in the presence of halides, organic solvents, and surfactants. Furthermore, the activity of CtLac on a dimeric lignin model compound, guaiacylglycerol-ß-guaiacyl ether (GGGE) was investigated. Liquid chromatography-mass spectrometry analysis indicated that CtLac catalyzes dimerization of GGGE to form a C5-C5 biphenyl tetramer. The stability and activity of CtLac characterized herein under thermoalkaliphilic conditions make it a highly suitable biocatalyst for various biotechnological and industrial applications.