A semi-automated instrument for cellular oxidative potential evaluation (SCOPE) of water-soluble extracts of ambient particulate matter

Abstract

Abstract. Several automated instruments exist to measure the acellular oxidative potential (OP) of ambient particulate matter (PM). However, cellular OP of the ambient PM is still measured manually, which severely limits the comparison between two types of assays. Cellular assays could provide a more comprehensive assessment of the PM-induced oxidative stress, as they incorporate more biological processes involved in the PM-catalyzed reactive oxygen species (ROS) generation. Considering this need, we developed a semi-automated instrument, the first of its kind, for measuring the cellular OP based on a macrophage ROS assay using rat alveolar macrophages. The instrument named SCOPE – semi-automated instrument for cellular oxidative potential evaluation – uses dichlorofluorescein diacetate (DCFH-DA) as a probe to detect the OP of PM samples extracted in water. SCOPE is capable of analyzing a batch of six samples (including one negative and one positive control) in 5 h and is equipped to operate continuously for 24 h with minimal manual intervention after every batch of analysis, i.e., after every 5 h. SCOPE has a high analytical precision as assessed from both positive controls and ambient PM samples (coefficient of variation (CoV)<17 %). The results obtained from the instrument were in good agreement with manual measurements using tert-butyl hydroperoxide (t-BOOH) as the positive control (slope =0.83 for automated vs. manual, R2=0.99) and ambient samples (slope =0.83, R2=0.71). We further demonstrated the ability of SCOPE to analyze a large number of both ambient and laboratory samples and developed a dataset on the intrinsic cellular OP of several compounds, such as metals, quinones, polycyclic aromatic hydrocarbons (PAHs) and inorganic salts, commonly known to be present in ambient PM. This dataset is potentially useful in future studies to apportion the contribution of key chemical species in the overall cellular OP of ambient PM.

On the Relative Contribution of Iron and Organic Compounds, and Their Interaction in Cellular Oxidative Potential of Ambient PM2.5

“…Because during the elution from the Chelex column, Fe(II) is oxidized to Fe(III) (see Figure S4), which has very low activity in the macrophage ROS assay, 29 ascorbate was used to recover Fe(II) in the metallic hydrophilic fraction. Then, 188 μL of ascorbate [concentration varying on the basis of the sample (see Figure S5 for more details about choosing the concentration of ascorbate)] mixed with 642 μL of 3 mg/L dithiothreitol (DTT, to prevent the consumption of ascorbate by Cu) was added to 1100 μL of the metallic hydrophilic fraction, and the mixture was diluted by DI to 5000 μL.…”

An Automated Online Field Instrument to Quantify the Oxidative Potential of Aerosol Particles via Ascorbic Acid Oxidation

“…Traditionally, acelluar ROS and OP measurement methods are based on the collection of PM filters, with quantification of OP taking place days, weeks or even months later (Venkatachari et al, 2005;Godri et al, 2011;Salana et al, 2021;Zhang et al, 2021). This results most likely in an underestimation of PM OP because many highly reactive aerosol components, including particle-bound ROS (e.g.…”