We report on a paper-based 2,2-diphenyl-1-(2,4,6-trinitrophenyl)hydrazyl (DPPH) assay for a simple, inexpensive, low reagent and sample consumption and high throughput analysis of antioxidant activity. The paper-based device was fabricated using a lamination method to create a 5-mm in diameter circular test zone that was embedded with a DPPH reagent. The analysis was carried out in one-step by dropping an antioxidant/sample onto the test zone. After reduction by the antioxidant, the DPPH radicals become stable DPPH molecules, resulting in a change in color from deep violet to pale yellow. The violet color intensity of DPPH was inversely proportional to the antioxidant activity of the samples, and was measured using imaging software. A high precision and a low limit of detection were found in the analysis of six standard antioxidants including gallic acid, trolox, ascorbic acid, caffeic acid, vanilliic acid and quercetin. The device was then validated against the traditional spectrophotometric DPPH assay by analyzing the antioxidant activity of 7 tea samples. The results showed no significant difference for gallic acid equivalent for all 7 samples obtained from the two methods at the 95% confidence level, indicating that the developed method was reliable for antioxidant activity analysis of real samples. Finally, the paper-based DPPH device was found to be stable over 10 days when stored in a refrigerator (2 - 4°C), making it an easy-to-use device for end-users.

We report a simple, low-cost, one-step fabrication method for microfluidic paper-based analytical devices (μPAD) using only polystyrene and a patterned screen. The polystyrene solution applied through the screen penetrates through the paper, forming a three-dimensional hydrophobic barrier, defining a hydrophilic analysis zone. The optimal polystyrene concentration and paper types were first investigated. Adjusting polystyrene concentration allows for various types of paper to be used for successful device fabrication. Using an optimized polystyrene concentration with Whatman#4 filter paper, a linear relationship was found to exist between the design width and the printed width. The smallest hydrophilic channel and hydrophobic barrier that can be obtained are 670 ± 50 μm and 380 ± 40 μm, respectively. High device-to-device fabrication reproducibility was achieved yielding a relative standard deviation (%RSD) in the range of 1.12-2.54% (n = 64) of the measured diameter of the well-shaped fabricated test zones with a designed diameter of 5 and 7 mm. To demonstrate the significance of the fabricated μPAD, distance-based and well-based paper devices were constructed for the analysis of H2O2 and antioxidant activity, respectively. The analysis of H2O2 in real samples using distance-based measurement with CeO2 nanoparticles as the colorimetric agent produced the same results at 95% confidence level, as those obtained using KMnO4 titration. A proof-of-concept antioxidant activity determination based on the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay was also demonstrated. The results verify that the polymer screen-printing method can be used as an alternative method for μPAD fabrication.

Particulate matter (PM) air pollution has a significant impact on human morbidity and mortality; however, the mechanisms of PM-induced toxicity are poorly defined. A leading hypothesis states that airborne PM induces harm by generating reactive oxygen species (ROS) in and around human tissues, leading to oxidative stress. We report here, a system employing a microfluidic electrochemical sensor coupled directly to a Particle-into-Liquid-Sampler (PILS) system to measure aerosol oxidative activity in an on-line format. The oxidative activity measurement is based on the dithiothreitol assay (DTT assay) where after oxidized by PM, the remaining reduced DTT was analyzed by the microfluidic sensor. The sensor consists of an array of working, reference, and auxiliary electrodes fabricated in a poly(dimethylsiloxane) (PDMS)-based microfluidic device. Cobalt (II) phthalocyanine (CoPC)-modified carbon paste was used as the working electrode material allowing selective detection of reduced DTT. The electrochemical sensor was validated off-line against the traditional DTT assay using filter samples taken from urban environments and biomass burning events. After off-line characterization, the sensor was coupled to a PILS to enable on-line sampling/analysis of aerosol oxidative activity. Urban dust and industrial incinerator ash samples were aerosolized in an aerosol chamber and analyzed for their oxidative activity. The on-line sensor reported DTT consumption rates (oxidative activity) in good correlation with aerosol concentration (R2 from 0.86–.97) with a time-resolution of approximately 3 minutes.

Human exposure to particulate matter (PM) air pollution has been linked with respiratory, cardiovascular, and neurodegenerative diseases, in addition to various cancers. Consistent among all of these associations is the hypothesis that PM induces inflammation and oxidative stress in the affected tissue. Consequently, a variety of assays have been developed to quantify the oxidative activity of PM as a means to characterize its ability to induced oxidative stress. The vast majority of these assays rely on high-volume, fixed-location sampling methods due to limitations in assay sensitivity and detection limit. As a result, our understanding of how personal exposure contributes to the intake of oxidative air pollution is limited. To further this understanding, we present a microfluidic paper-based analytical device (μPAD) for measuring PM oxidative activity on filters collected by personal sampling. The μPAD is inexpensive to fabricate and provides fast and sensitive analysis of aerosol oxidative activity. The oxidative activity measurement is based on the dithiothreitol assay (DTT assay), uses colorimetric detection, and can be completed in the field within 30 min following sample collection. The μPAD assay was validated against the traditional DTT assay using 13 extracted aerosol samples including urban aerosols, biomass burning PM, cigarette smoke and incense smoke. The results showed no significant differences in DTT consumption rate measured by the two methods. To demonstrate the utility of the approach, personal samples were collected to estimate human exposures to PM from indoor air, outdoor air on a clean day, and outdoor air on a wildfire-impacted day in Fort Collins, CO. Filter samples collected on the wildfire day gave the highest oxidative activity on a mass normalized basis, whereas typical ambient background air showed the lowest oxidative activity.

We report the first use of a paper-based device coated with nanoceria as a simple, low-cost and rapid detection platform for the analysis of organophosphate (OP) pesticides using an enzyme inhibition assay with acetylcholinesterase (AChE) and choline oxidase (ChOX). In the presence of acetylcholine, AChE and ChOX catalyze the formation of H2O2, which is detected colorimetrically by a nanoceria-coated device resulting in the formation of a yellow color. After incubation with OP pesticides, the AChE activity was inhibited, producing less H2O2, and a reduction in the yellow intensity. The assay is able to analyze OP pesticides without the use of sophisticated instruments and gives detection limits of 18 ng mL(-1) and 5.3 ng mL(-1) for methyl-paraoxon and chlorpyrifos-oxon, respectively. The developed method was successfully applied to detect methyl-paraoxon in spiked vegetables (cabbage) and a dried seafood product (dried green mussel), obtaining ∼95% recovery values for both sample types. The spiked samples were also analyzed using LC-MS/MS as a comparison to the developed method and similar values were obtained, indicating that the developed method gives accurate results and is suitable for OP analysis in real samples.