Effects of 6PPD-Quinone on Human Liver Cell Lines as Revealed with Cell Viability Assay and Metabolomics Analysis.

N-(1,3-Dimethyl butyl)-N'-phenyl-phenylenediamine-quinone (6PPD-Q) is a derivative of the widely used rubber tire antioxidant 6PPD, which was first found to be acutely toxic to coho salmon. Subsequent studies showed that 6PPD-Q had species-specific acute toxicity in fishes and potential hepatotoxicity in mice. In addition, 6PPD-Q has been reported in human urine, demonstrating the potential widespread exposure of humans to this chemical. However, whether 6PPD-Q poses a higher risk to humans than its parent compound, 6PPD, and could cause adverse effects in humans is still unclear. In this study, we utilized two human liver cell models (the human proto-hepatocyte model L02 and the human hepatocellular carcinoma cell line HepG2) to investigate the potentially differential effects of these two chemicals. Cell viability curve analysis showed that 6PPD-Q had lower IC50 values than 6PPD for both liver cell lines, suggesting higher toxicity of 6PPD-Q to human liver cells than 6PPD. In addition, L02 cells are more sensitive to 6PPD-Q exposure, which might be derived from its weaker metabolic transformation of 6PPD-Q, since significantly lower levels of phase I and phase II metabolites were detected in 6PPD-Q-exposed L02 cell culture medium. Furthermore, pathway analysis showed that 6PPD-Q exposure induced changes in phenylalanine, tyrosine, and tryptophan biosynthesis and tyrosine metabolism pathways in L02 cells, which might be the mechanism underlying its liver cell toxicity. Gene expression analysis revealed that exposure to 6PPD-Q induced excessive ROS production in L02 cells. Our results further supported the higher risk of 6PPD-Q than 6PPD and provided insights for understanding the effects of 6PPD-Q on human health.

Distribution patterns of rubber tire-related chemicals with particle size in road and indoor parking lot dust.

In recent years, concerns have arisen from the chemicals incorporated into tire material which are of potential to leach with the tire and road wear particles (TRWP) into the environment. In this study, the distributions of substituted benzothiazoles (BTHs) and p-phenylenediamines (PPDs), two groups of representative TRWP-related chemicals, were investigated in various size fractions (<20, 20-53, 53-125, 125-250, 250-500, 500-1000 µm) of dust samples from open roads and indoor parking lots in the urban region of Guangzhou (Guangdong, China). Mass weight distribution of the dust samples showed that fractions of <250 µm accounted for >72% of the total dust in both microenvironments. Widespread occurrence was observed with >80% detection frequency for almost all target compounds in all the particle fractions. Concentrations of BTHs and PPDs were similar between the two dust matrices. In addition, the newly defined transformation product of 6PPD, 6PPD-Q was at the median concentration of 122 ng/g in road dust and 154 ng/g in indoor parking lot dust. Overall, concentrations of the target BTHs and PPDs varied in different size fractions, which were mostly dominated in fine particle sizes (<53 µm). Specially, >70% of the target compounds were in the size fractions of <250 µm, suggesting the necessity of using <250 µm fractions of particles for monitoring and evaluating contamination levels and exposure risks of BTHs and PPDs from dust in future studies.

Simultaneous quantification of plasma immunoglobulin subclasses for assessment of maternal and fetal immune response during pregnancy.

Measurement of immunoglobulin subclasses is a useful tool for exploring humoral immune deficiency in the presence of total immunoglobulins within reference intervals. Conventional methods for immunoglobulin measurement are mostly immunoassays, which are of low throughput and laborious to run multiple immunoglobulin subclass tests. Liquid chromatography-mass spectrometry (LC-MS) has emerged as a promising technology for the measurement of protein biomarkers in biological matrices, owing to its high specificity, selectivity, multiplexing, and wide dynamic range. In fully taking these advantages, we developed here a LC-MS based methodology for simultaneous quantitation of the primary immunoglobulin isotypes (IgG, IgA, IgM) and their subclasses in human plasma. Method validation demonstrated that the proposed method showed good linearity with R > 0.99, high precision with coefficients of variation for inter- and intra-assay less than 15%, as well as high accuracy with relative errors less than 8.7%. The developed method was further applied to maternal and cord blood collected at delivery for assessment of maternal and fetal immune status. The immunoglobulin profiles and the features of transplacental transport of maternal immunoglobulin subclasses were comparable to the findings from previous reports, further demonstrating the reliability of this method. Therefore, our method provides a competitive approach to high-throughput detection of multiple immunoglobulin subclasses for biomonitoring or epidemiological studies.