Portable Colorimetric Paper-Based Biosensing ... - ACS Publications

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Environmental Science & Technology

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A Portable Colorimetric Paper-Based Biosensing Device for

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the Assessment of Bisphenol A in Indoor Dust

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Ramiz S. J. Alkasir†, Alan Rossner,ǁ Silvana Andreescu†

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13699 - 5810, United States

Department of Chemistry and Bimolecular Science, Clarkson University, Potsdam, New York

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United States

Institute for a Sustainable Environment, Clarkson University, Potsdam, New York 13699 - 5715,

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Table of Content

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22 Test Zone of Colorimetric Biosensing Device

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PVC Filter Membrane

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Abstract

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Bisphenol A (BPA) is found in polycarbonate plastic and epoxy resins and is used in a

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variety o f commercial and consumer products. The leaching of BPA c a n r e s u l t i n human

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exposure via inhalation, ingestion, and dermal routes. As a result, humans have been exposed in

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their home and work environment to BPA. Conventional methods for BPA exposure assessment

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rely on cumbersome laboratory instrumentation with high capital and operational expenditures

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which limit the number of samples that can be analyzed. We report here the design of a compact

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portable colorimetric paper based biosensing device with integrated sampling/analysis units for

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field-based measurements of BPA in indoor dust. The system employs interchangeable low cost

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paper based enzyme sensors as a test zone for BPA detection interfaced with an air sampling

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cassette as a sample collection area. The sensor response was concentration dependent with a

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detection limit of 0.28 µg/g.

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Chromatography (GC) method and used to detect BPA exposure in household dust. BPA

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concentrations ranged from 0.05 to 3.87 µg/g in 57 household dust samples when both methods

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were used. The potential of this method for field measurements of dust samples is discussed.

The sensor was validated with the conventional Gas

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Keywords: bisphenol A, paper biosensor, portable device, indoor dust, exposure assessment

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INTRODUCTION

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Bisphenol A (BPA) is a phenolic compound commonly used in the plastics industry since the

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1960s.1 Worldwide BPA production is extensive; in 2013, 4.7 million tons of BPA were

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manufactured worldwide.2 BPA is generally used as plasticizer for plastic consumer products

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that include drinking bottles, water supply pipes, eyeglass lenses, dental sealants, plastic wraps,

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compact discs, and toys.3 Moreover, BPA is used in the production of paints, carpets, recycled

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toilet papers, and thermal papers such as store receipts.4 Recently, public attention has been

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drawn to BPA in the United States, Europe, and Japan because of its estrogenic activity and

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widespread presence in the environment.5 BPA is considered an endocrine disruptor; in very low

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doses it can impact biological systems and lead to behavioral, reproductive, developmental, and

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neurological disorders.6 BPA enters the human body through different pathways, including daily

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oral exposure, inhalation and dermal absorption. Scientists from the U.S. Center for Disease

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Control and Prevention (CDC) found BPA in more than 90% of the U.S. population in 2009.1, 7

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This high percentage is related to the ingestion of foods stored in BPA-bearing packaging8 and in

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the air we breathe.9, 10 Most studies in the literature have focused on BPA exposure from dietary

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sources. Other potential sources, such as air and dust, have received far less attention. Animal

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studies suggest that BPA exposures, even at low doses well below the established U.S.

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Environmental Protection Agency (EPA) reference dose of 50 µg/kg BW/day, are associated

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with breast and prostate cancer, cardiovascular disease, and reproductive and behavioral

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abnormalities.6, 11, 12, 13

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While relatively high concentrations of BPA are reported to be released in the living

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environment, there is a lack of temporal and spatial resolution data to characterize BPA



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exposure. Traditional analytical techniques for indoor measurements of BPA rely on expensive

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laboratory based instrumentation that requires specialized expertise, sample collection and

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subsequent laboratory analysis. Conventionally, BPA is collected on filters or sorbent tubes and

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analyzed using GC or high pressure liquid chromatography (HPLC) coupled with MS.14 These

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procedures require multiple steps, use hazardous chemicals and are time consuming. An

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improved understanding of the exposure-response effects of BPA requires field deployable

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instrumentation that can be used for site monitoring of BPA levels in the living environment. To

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further this understanding and facilitate spatiotemporal measurements of BPA exposure, we

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report design, development and field demonstration of a biosensing based sampling/analysis

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instrument for BPA exposure assessment in indoor dust. This instrument makes use of low cost

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paper based portable sensors biofunctionalized with a BPA responsive enzyme deposited on a

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paper platform .

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Paper based bioanalytical devices are emerging as a promising low cost technology for

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field monitoring in the clinical and environmental fields.13, 16 Paper-based devices have a flexible

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design, are easy to carry, and straightforward to operate. The cellulosic fibers of paper can be

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modified with materials for sensing, biorecognition and signal amplification functions. Examples

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include optically responsive nanoparticles,15 polymers and biological reagents such as

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enzymes,16 aptamers and DNA. These devices provide the required selectivity and sensitivity and

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have a performance comparable to that of laboratory based methods.17 Quantitative analysis

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involving paper technology is usually based on color measurements. Color intensity can be

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expressed digitally when red, green and blue colors are numerically quantified as expressions of

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analyte concentration.10, 18 Digital image scanners, high-resolution smart phone cameras,19 field


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portable color readers like Pantone/X-Rite20 and graphic design apps are frequently used for

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color analysis,21 adding convenience and flexibility in analysis.

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Despite extensive research in the sensing field use of these platforms for the field analysis

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of aerosol and dust contaminants has been limited. One example of a paper sensor for oxidative

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activity measurements in urban air environments has been designed using a dithiothreitol assay.22

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Other examples include a paper-based device with colorimetric and electrochemical detection of

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metals including Fe, Ni, Cr, Cu, Pb and Cd in indoor and outdoor air samples23 and for

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measuring the metals content in welding fumes24 and the total Cr in airborne particulate matter.25

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Here we describe the development and field performance of a compact device for field-based

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BPA measurements that incorporates interchangeable low cost portable sensors with sample

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collection, extraction and analysis, all in one unit. In previous work we introduced a reagentless

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enzyme based colorimetric method on paper to measure BPA, phenol, catechol, and cresols in

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aqueous mediums. The limit of detection (LOD) for BPA was 0.83 µg/L.16 In this work, we

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expand this method to dust specimens for the following reasons: (1) the low LOD suggests the

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method could be applied to dust measurements, (2) the paper sensor can be interfaced with air

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sampling units and (3) sample analysis through measurement of color intensity can be made

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directly on the sensing pad, immediately following sample aspiration without any sample

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preparation.

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The field sampling/analysis biosensing system was designed to be a robust detection

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method and a sample collection procedure for BPA in surface and airborne measurements in

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household or commercial buildings. The device comprises two main parts: (1) an air-sampling

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cassette, and (2) the colorimetric paper-based enzyme sensor, shaped as a circular disc with a

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diameter of 0.6 cm. The plastic cassette is used as a platform for the paper sensor and allows for



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the efficient collection of sample whereas the paper sensor is used for BPA detection based on

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color change (Figure 1). The sensor contains in built detection in which BPA is first oxidized

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enzymatically to its corresponding quinone, which then reacts with a chitosan layer to form a

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charge transfer quinone-imine complex with strong absorption in the visible range. All the

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reagents necessary for analysis (except for the sample) are immobilized on paper. The system

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requires only addition of a small amount of liquid for analysis, buffer or water, to activate the

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enzyme, initiate the biorecognition reaction and generate the colorimetric response on the paper

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surface. Exposure of the sensing surface to the sample is done by aspiration of an air flow to the

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responsive surface. Sensor data is gathered as a visual color change with the naked eye and by

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using common image analysis software such as Adobe or Image J, or with an iPhone. A color

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database was generated based on an (RGB) color space. The database numerically quantifies the

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color intensity and permits an estimate of BPA concentration. The system was used to assess

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BPA exposure in 57 household locations, as well as a local day care, and the results were

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validated against the conventionally GC/MS method. To our knowledge, this is the first report of

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a field-deployable enzyme based analytical device for BPA detection in surface dust.

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EXPERIMENTAL SECTION

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Chemicals and Materials. Bisphenol A (BPA; 4,4'-(1-methylethylidene)bis-phenol) (99%+

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purity), tyrosinase (E.C.1.14.18.1, 3130 units/mg), chitosan from crab shells, alginic acid

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(sodium salt), phenol (99% purity), and sodium triphosphate pentabasic, NaTPP (practical grade)

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were purchased from Sigma-Aldrich (St. Louis, MO, USA). Methanol (HPLC Grade, 99.9%),

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sodium phosphate monobasic, sodium phosphate dibasic anhydrous, and Fisherbrand filter paper


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(P5; 09-801C) with a diameter of 11 cm and a medium porosity were supplied by Fisher

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Scientific. A 37 mm air sampling cassette (3-Piece w/ 5.0 µm, PVC Filter) was purchased from

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Zefon International, Inc. (Ocala, FL). A portable battery powered air sampling pump (model BGI

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400S, BGI, INC., Waltham, MA) was used as the vacuum source. A nonpyrogenic/ nontoxic 1

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ml syringe, which was obtained from NORM-JECT ® Tuberkulin (Henke Sass Wolf GmbH,

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Germany), was used as the hose extension with a cut at a 45o angle. A fine air cleaner test dust

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(Batch No. 1761) classified from the natural Arizona dust was prepared by AC Spark Plug Div.

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(GM corporation, Flint, Michigan, USA) with a particle size of 44 µM. Household dust with a

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particle size of 45 µm was obtained after sieving.

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Stock solutions of chitosan at 1.25% (w/v) and sodium alginate at 2.0% (w/v) were

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prepared by dissolving chitosan and sodium alginate in 0.1 M acetic acid and distilled water

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(DI), respectively. The stock solution of 1 x 10-2 M BPA was prepared in methanol. Solution of

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NaTPP (1.5% w/v) and 0.1 M phosphate buffer (PBS) at pH 6.5 were prepared in DI. All

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reagents were used without further purification, unless noted, and all solutions were prepared

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with DI water (Millipore, Direct-Q-system) with a resistivity of 18.2 MΩ.

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Fabrication of the Colorimetric Paper-Based Biosensor. The device was designed as a

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portable measurement tool comprised of a sampling unit with a flow-through chamber connected

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to an aerosol sampling cassette. The cassette contained the paper sensor with direct reading

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ability of BPA concentrations. The sensor was fabricated using a layer-by-layer assembly

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approach with the enzyme tyrosinase immobilized in between the alternating layers of chitosan

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and alginate as described.16 Spectroscopic analysis confirmed the formation of a colored

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quinone-imine compound between the quinone carbonyl groups and the nucleophilic amino



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groups of chitosan.16,

A strong absorption band was present at around 610 nm for BPA27,

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which is the basis of the colorimetric detection using paper-based tyrosinase biosensing system.

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The sensors can be mass produced by inkjet printing and stored in dry state for up to one year at -

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20 oC, 4 oC or at room temperature (~ 25 oC), respectively.16 To ensemble the sampling/analysis

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device in a compact unit, the PVC filter membrane, already installed on the lower section of the

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sampling cassette, was used as a pad for mounting the paper sensor disk. The upper part was

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attached directly to a hose 6.0 cm long, whereas the lower part was connected to a portable

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aerosol pump with a flow rate 2.5 L/min through a rubber tube of a length 25 cm (Figure 1). In

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our first set of experiments, we used a Mixed Cellulose Ester (MCE) membrane filter, but the

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buffer solution was rapidly adsorbed during the test time. This was likely due to the

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hydrophilicity of this membrane. Therefore all experiments were performed with a PVC

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membrane that is hydrophobic and can retain the aqueous solution on the paper sensor.

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Analysis and Field Sampling. To perform analysis, the sample zone of the sensor (0.6 cm

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diameter) was placed into the air flow of the sampling cassette connected to a sampling pump for

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collecting dust at a controlled rate as shown in Figure 1A. Sampling was initiated by air flow

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aspiration of dust samples across the filter. The common sampling area selected for vacuuming

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dust was about 100 cm2 (L 10 cm × W 10 cm), based on the ASTM E1973 procedures.28

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After exposure, 25 µl of 0.1 M PBS at pH 6.5 were added to the test zone to activate the sensing

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materials and initiate the enzymatic reaction. Because of the hydrophobic property of PVC

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membrane, the solution was completely retained on the surface of the paper sensor during the

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test time. Once the enzyme paper sensor is in contact with the BPA-dust sample, a greenish

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color started to appear within the first 60 s after addition of buffer. The color stabilized after 30

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min (Figure 2). The color formation was monitored each 30 s for 60 min using an iPhone 4

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camera. The digital images of the test zones (0.28 cm2 with r = 0.3 cm) were analyzed by Adobe

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Photoshop. For each sample measurement, we used a new NORM-JECT syringe and air-

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sampling cassette. The sensor response was measured on the active side of the paper containing

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the enzyme on which dust particles are collected. Reading of the backside of the sensor was also

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considered but the scanned image of the back paper did not show a green color. Dust sampling

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was performed in 57 locations in 4 houses and a day-care center in Potsdam, New York. In each

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location 6-10 samples were collected on carpets, fabric furniture and hard surface floors. These

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sample locations were selected because they were horizontal surfaces where particle bound BPA

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may accumulate. Dust samples were aspirated in field directly into the sampling cassette

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containing the paper sensor. Sample analysis was performed without any extraction or sample

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treatment. Samples collected on filters for validation purposes were extracted and analyzed using

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a GC/MS method.

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Color Analysis. The conventional RGB color space of red, green and blue intensities was used

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to quantify the color on the paper platform.16, 30 BPA showed a specific greenish color on the

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paper sensor which is easy to measure using the sRGB color space and can be conveniently

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quantified by Adobe Photoshop. Synthetic RGB color spaces provided by Adobe Photoshop

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include sRGB, Apple RGB, ColorMatch RGB, Adobe RGB (1998), Wide Gamut RGB, and

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ProPhoto RGB.31 Each is mathematically designed to provide flexible editing options for image

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quality. In this work, we have selected the color profile: sRGB IEC61966-2.1 which was

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installed in Adobe®Photoshop®CS for assignment of numerical values to color intensities. sRGB

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was chosen for several reasons: (1) a neutral color was easy to define, (2) sRGB was derived

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from HDTV standard; thus, all LCD and CRT displays can produce sRGB if properly calibrated,

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(3) sRGB has a flexible color gamut size compatible with the gamut sizes of consumer-level

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digital cameras, and scanners produced by Hewlett-Packard. A desktop HP ScanJet 3970

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Scanner with a resolution of 2400 dot per inch (dpi) was used to obtain a high-resolution scanned

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image of the paper sensor disk and record the RGB light intensities. A desktop scanner is

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preferable to other imaging tools because it prevents variable lighting conditions (daylight and

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reflection) from distorting the intensity of the scanned image.21, 32 We used the eyedropper tool

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in Adobe Photoshop with an average of 5 ×5 pixels to analyze the RGB color channels of the

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image and to obtain the digital values. Each 8-bit digital value of the RGB color channels ranged

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from 0 to 255 for each channel. We selected a digital value of red color channel for BPA

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detection which is the complementary color of green on the color wheel chart.

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Method Validation using GC and GC/MS. Validation of the colorimetric biosensing method

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was carried out using filter extracts of BPA collected at the same time and locations as with the

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sensor. Each filter collected in the field was divided into two equal parts: one for analysis using

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the colorimetric biosensing device and the other analyzed using GC/MS. Samples for GC

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analysis were dust accumulated on the PVC filter membrane followed by extraction with

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methanol using a Soxhlet-extracted apparatus. The methanol extract was then concentrated by

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Kuderna-Danish evaporation. Finally, the concentrated aliquot was analyzed for BPA by

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GC/MS. All GC analysis of sample extracts and standard solutions were done on a Thermo trace

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GC ultra, equipped with a ChromQuest 5.0 data system. The GC column was a TR-5 fused silica

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capillary column (30 m × 0.32 mm ID; 0.25 µm film thickness). Following injection, the GC

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column was held at 80oC for 1 min, then programmed to 230 oC at 15 oC/min, and then to 290 oC


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at 10oC/min. For BPA quantification in household dust samples, GC peak areas were monitored

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at a retention time of 15.1 min. These quantifications correspond to internal standards of the

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target analyte that were generated from standard calibrations. The fragment peaks of the dust

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components were identified with a Hewlett-Packard GC/MS, operated in SIM mode (70 eV EI)

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with a ChemStation data software. The peaks represent the abundances of molecular ions in the

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dust. The GC column DB-5 has the same parameters as a TR-5 capillary column and was used

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for GC/MS measurements.

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RESULTS AND DISCUSSION

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Analytical Performance of the Colorimetric Biosensor for BPA Detection in Dust. We first

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established the ability of the paper sensor to determine BPA in dust. Calibration curves were

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initially carried out using Arizona road dust which is considered to be a “clean” natural dust and

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therefore is commonly used as a standard. The dynamic ranges of the calibration curve show

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linear responses with the BPA concentration (Figure S1). However, Arizona dust has a light

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brown-reddish color, whereas household dust is uniformly gray. Therefore, the household dust

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was a more representative dust to use in our experiments. The Arizona dust was tested and can

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be successfully used with this sensor. The measurements can be expanded to outdoors dust

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measurements but we caution that background color of the dust should be considered. Household

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dust used for calibration was washed nine times with methanol to dissolve and remove other

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chemicals that might interfere with analysis and affect the accuracy of the sensing device

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(Figure S2). The methanol solution acquired a yellow color after the first wash, but became

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colorless after the ninth. GC/MS analysis of washed solutions shown in Figure S3 does not show

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any detectable level of BPA and other constituents. Cleaned household dust with a particle size 12 | P a g e ACS Paragon Plus Environment


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of 45 µm was dried and stored at room temperature. Colorimetric analysis on standard specimens

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of washed household dust spiked with BPA resulted in an increase in color intensity with

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increased BPA concentration in the range of 1-500 µg/g. The calibration curve shows two

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dynamic ranges: the first one in the range of 1-25 µg/g, and the second concentration range of

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50-500 µg/g (Figure 2A). The colored digital images in Figure 2B represent the test zones used

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to create the calibration curve. The limit of quantification (LOQ) was 0.95 µg/g, and the

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detection limit (LOD) was 0.28 µg/g (determined using the 3σs/S criteria, where S is the slope of

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the linear calibration curve, and σs is the standard deviation of the color intensity of the blank).

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The reproducibility was evaluated for three identical test zones. The average color intensity was

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112.6 (±2.082) with a calculated RSD value of 0.018 (n=3) for 100 µg/g BPA. The system was

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tested with two wetting agents: phosphate buffer solution and tap water. The sensor showed only

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slightly higher colorimetric response to 100 µg/g BPA in phosphate buffer (112.6±2.082; n=3) as

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compared to tap water (110.3±3.055; n=3) (Figure S4). This indicates that water can equally be

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used to initiate the enzymatic/colorimetric reaction. The results demonstrate the ability of the

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device to measure concentrations of BPA in dust at exposure concentration levels as reported

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using traditional GC methods for BPA in dust.12-14

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A color database was specifically designed to

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Color Database for BPA Quantification.

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recognize BPA from a mixture of colors that appear in the test zone after exposure to dust, and

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to quantify the intensity of the red color, a complementary color of green. The digital values of

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the intensities of red, green, and blue were manually recorded from the website of Valspar paint

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® (Chicago, IL, USA),33 based on the ID of a particular color (Table S1). The color intensity of

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a real dust sample was matched with a compatible color ID in the database to determine BPA


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concentration. The arithmetic mean of pixel intensity obtained from the Histogram option in

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Adobe Photoshop was used to determine the matching color. Figure 3 shows the red color

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channel histograms with the mean of pixel intensities. One can match the color of a dust sample

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with the color obtained from the database. For example, the mean of pixel red intensity was

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140.40 (with Std Dev 74.23) for sample 57, whereas it was 140 for a color of ID (EB 10-1). As a

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result, the mean difference of pixel red intensity was (±0.40). The dust sample, however, had a

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relatively high standard deviation. This might be due to a mixture of colors that can be found in

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the dust specimen; a broad range in red color intensity was clearly observed in the histogram of a

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dust sample (Figure 3B). In addition, narrow red color intensity was shown in the histogram of

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blank B1 of clean household dust (Figure 3D). Table S2 summarizes the digital images of each

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test zone exposed to dust as well as the RGB values with their color ID. To determine the BPA

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concentration in dust, blank subtraction was applied. The mean pixel intensity of a real sample

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was subtracted from the mean of pixel red intensity of the blank, which was 175 (n=3).

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Study of Interferences. To assess the effect of interferences originating from other compounds,

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we studied the response of the sensor to phenol, catechol and ascorbic acid in single and binary

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mixtures. The colorimetric sensors display a brown color for phenol, dark brown for catechol and

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red for ascorbic acid, while the color for BPA is blue/green.34 The different colors generated by

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the individual compounds (with measurement of their complementary color) provide selectivity

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to the sensor. The colorimetric profile for BPA detection follows a similar trend with and without

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the interfering compound. The presence of phenol, catechol or ascorbic acid in the mixture had

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little effect on the intensity of the greenish color produced by BPA for BPA content up to 50%

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when this was compared with that obtained in the absence of interferent (Figure S5). For higher



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amount of interferent in the mixture, a reduction in color intensity was observed. However,

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according to the GC/MS profile of household dust (described in the following section), BPA is

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present at much higher levels as compared to other phenolics which can be attributed to the low

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vapor pressure of BPA promoting BPA adsorption on the particles35 and thus, such interferences

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would be minimal. Other compounds such as uric acid, ascorbic acid and phenylalanine have no

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effect on the biosensor response, as previously demonstrated.16 We have also studied the effect

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of potential inhibitors of enzyme activity, such as organophosphate pesticides36 that may be

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present in the environment and effect on enzyme activity (Figure S6). Concentrations of

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paraoxon below 50 µg/mL had no effect on the sensor response. A small decrease (below 15%)

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was observed for very high paraoxon concentrations (between 100- 250 µg/mL). However, such

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high concentrations are not expected in indoor dust.

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Field Sampling and Quantitative BPA Measurements in Dust. Dust collection was done in

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four homes and a Day care center in Potsdam, New York. Four family members lived in each

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house: two adults between 40 and 55 and two children ranging in age from 2 to 10 years. Two

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sources of household dust were selected: carpet dust and couch dust. Because BPA is typically

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used as a component of epoxy resins in the manufacture of floorings and furniture, we

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hypothesized that BPA may accumulate on these locations. Table 1 summarizes the sampling

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locations in each house (H1 through 4) and the day care center (DC). Figure S7 shows digital

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images and color change of the colorimetric test zones in household dust as compared to clean

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dust, used as blank. BPA concentrations in carpet dust of the living rooms in H1 and H2 were

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2.28, and 3.13 µg/g, respectively. BPA in carpet dust of family rooms in H3 and H4 were 1.25

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and 0.38 µg/g, respectively. The family room of H1 yielded a BPA concentration of 2.10 µg/g in 15 | P a g e ACS Paragon Plus Environment

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dust from couch fabric. The presence of old unclean carpet and plastic beverage containers

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resulted in higher BPA levels. The cleaned carpets of family rooms in H3 and H4 showed lower

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BPA levels. No BPA was detected in the living room carpet of H3; the carpet was an antique

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wool-made manually most likely without epoxy resins. The method was further used to quantify

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the concentration of BPA in carpet dust at a day care center that housed about 60 children,

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ranging from infants to 10 years-olds. BPA concentration ranged from 1.28 to 3.78 µg/g in

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carpet dust from seven different rooms. We expected to find high levels of BPA in the day care

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center because of the abundance of the plastic toys, bed mattresses, shelves and clothes

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containers. In addition, children (who stay up to 8 hours per day) often wear plastic and rubber

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shoes and boots. Workers vacuumed the carpet in each room daily; nonetheless, BPA was found

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in the vacuumed dust. We further investigated whether the BPA in the vacuumed dust originates

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from the carpet itself. BPA analysis in a new carpet, similar to the one installed in the day care

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center revealed presence of a BPA concentration of 0.8(±0.18) using the colorimetric method

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and of 0.83(±0.15) using the GC method. This analysis shows that part of BPA exposure is due

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to the presence and release of BPA from the manufactured carpet.

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Method Validation. A GC method was developed to validate the colorimetric biosensing

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method for household dust analysis. Figure S8 shows the calibration curve in the range of 1-500

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µg/ml. GC chromatograms of standard solutions of BPA showed well-defined peaks at a

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retention time of 15.1 min. GC analysis of 57 dust samples indicates that BPA is present in the

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dust. For example, sample 57 showed a well-defined peak at retention time of 15.102 min

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whereas the blank showed no peak at that retention time (Figure S9). The results of the

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comparative analysis of BPA in 57 household dust samples by the colorimetric and GC methods



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are summarized in Table S3. Comparative BPA analysis in three samples vacuumed from a new

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carpet revealed presence of BPA at concentrations of 0.8±(0.18) and 0.83±(0.15) by the

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colorimetric and GC method respectively (Table S4). Comparative performance between the two

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methods showed a linear regression: (y = 0.9484x + 0.2508; R2= 0.9743) for both methods

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(Figure 4).

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concentration of BPA, however, this bias is well within tolerance given the acceptahble field

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sampling and analytical errors for methods used for environmental and occupational monitoring.

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Side-by-side comparison of BPA concentrations found by the colorimetric and GC in the

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household dust samples at the same locations shows good correlation between the two methods

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(Figure 5). These results indicate the ability of the colorimetric method to measure BPA in dust

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at concentrations comparable to the traditional GC method, which validates our colorimetric

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method.

The slope is approximately 0.95, hence slightly underestimating the “actual”

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GC/MS was further used to characterize the components of dust and confirm the presence

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of BPA. Household dust specimens analyzed by GC/MS were collected from the same locations

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as with the sensor. MS results indicated abundance intensity at m/z 228for a standard solution of

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BPA (Figure S10A), a value related to the molecular weight of BPA at 228 g/mol.37 The

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fragment ion at 91 m/z corresponds to the benzylic cleavage from the alkyl substituted benzenes,

362

while the fragment ions at 77 and 65 m/z are related to the formation of phenyl cation and the

363

loss of CHO from the phenolic rings, respectively. No abundance intensity was observed at the

364

m/z position of a blank sample containing methanol (Figure S10B). The MS of Arizona road

365

dust showed abundance intensities at different m/z positions that are related to silane, esters,

366

amides, and hydrocarbons (Figure S11). The MS of household dust collected with a vacuum

367

cleaner indicated the presence of phenolics and ketones (Figure S12). Components of Arizona


Page 19 of 31


368

road dust and the household dust found by GC-MS are summarized in Table S5. The

369

components of the cleaned household dust, which was used as a blank, are shown in Table S6.

370

The results of our analysis of 57 samples of indoor dust by the colorimetric and GC methods

371

showed BPA concentration that ranged from 0.05 to 3.87 µg/g. The range was similar (0.567- to

372

3.26 µg/g) to that obtained by Wilson et al12 who analyzed floor dust from two day care centers

373

in Raleigh-Durham, North Carolina. Another study found BPA concentrations ranging from

374

0.535 to 9.729 µg/g in 18 samples of indoor dust collected from 18 homes in Belgium.13 This

375

group reported two offices with BPA concentrations of 4.69 and 8.38 µg/g.35 Liao et al have

376

found BPA in the range of 0.496-12.3 µg/g in 19 indoor dust samples collected from five cities

377

in Japan.38 Loganathan et al reported BPA in the range of 0.135-0.232 µg/g in 9 samples from 9

378

houses in Albany, New York.39

379 380

Implications in the Environmental Monitoring and Exposure Assessment. Our design of the

381

colorimetric paper-based biosensing device represents a new approach for the detection of BPA

382

in dust specimens. The sensor performance compared well to the more established methods.

383

Advantages of the system include on-site measurements and ease-of-use . The device can be

384

used in homes, day care centers, schools and the work environment. Two unique aspects of the

385

work include (1) the reagentless operation with all in one sampling and analysis and (2) a

386

database color ID that quantifies color intensity based on sRGB color profile. A smartphone app

387

can be further interfaced with the sensor to enable direct in site image quantification based on a

388

pre-calibrated color scheme. This sensor design could be used to establish a procedure for indoor

389

surface sampling for BPA. The system is not limited to detecting BPA. The same design could



Page 20 of 31

390

be adapted for detecting other chemicals that have potential human health concerns such as

391

VOCs, phthalates, and pesticides.

392

There is a great deal of interest in conducting large scale epidemiological studies to

393

evaluate exposure risks to a variety of chemicals, including endocrine disrupting chemicals like

394

BPA, pesticides and volatile organic compounds. Although many low cost devices have been

395

created, these sensors have not been used for aerosol analysis. This study was designed to

396

demonstrate the feasibility of low cost paper based devices with biomolecular recognition as

397

reliable detection tools for field based assessment of BPA exposure in dust samples. These data

398

can provide a basis for implementation of other types of sensors for aerosol measurements.

399

Further development and adoption of these devices can accelerate risk assessment and

400

epidemiological studies of BPA exposure (and potentially of other chemicals by changing the

401

sensing strip) in a variety of environments and conditions.

402 403 404

ASSOCIATED CONTENT

405 406

Supporting Information. Additional calibration curves of Arizona dust, the color analysis

407

database and the results of the GC and GC/MS analysis for the household samples tested and

408

controls. This material is available free of charge via the Internet at http://pubs.acs.org.

409 410

AUTHOR INFORMATION

411

Corresponding

412

[email protected] (AR) and [email protected] (SA).

Authors.

*Phone

315

268

2394.

Fax:

315

268

6610.

E-mails:


Page 21 of 31


413 414

Author Contributions. The manuscript was written through contributions of all authors. All

415

authors have given approval to the final version of the manuscript.

416 417

Funding Sources. This material is based upon work supported by the National Science

418

Foundation under Grant No. 0954919. Any opinions, findings, and conclusions or

419

recommendations expressed in this material are those of the author(s) and do not necessarily

420

reflect the views of the National Science Foundation.

421 422 423

ACKNOWLEDGMENT

424

The authors would like to thank Dr. Andrea Ferro for providing the Arizona dust and for help

425

with setting up household collection procedures.

426 427

REFERENCES

428 429 430 431 432 433 434 435 436 437 438 439 440 441 442

1. Lakind, J. S.; Naiman, D. Q., Daily intake of bisphenol A and potential sources of exposure: 20052006 National Health and Nutrition Examination Survey. Journal of exposure science & environmental epidemiology 2011, 21 (3), 272-9. 2. www.mindbodygreen.com/, BPA what is it and do we really need to worry about it/ Healthy Child Health World 2013. 3. Liao, C. Y.; Kannan, K., Widespread Occurrence of Bisphenol A in Paper and Paper Products: Implications for Human Exposure. Environ Sci Technol 2011, 45 (21), 9372-9379. 4. Zimmers, S. M.; Browne, E. P.; O'Keefe, P. W.; Anderton, D. L.; Kramer, L.; Reckhow, D. A.; Arcaro, K. F., Determination of free Bisphenol A (BPA) concentrations in breast milk of US women using a sensitive LC/MS/MS method. Chemosphere 2014, 104, 237-243. 5. Services, U. S. D. o. H. a. H., Annual Report; President’s Cancer panel. Reducing Enviromental Cancer risk. National Institutes of Health, National Cancer Institute 2008-2009 2, 1-240. 6. Maffini, M. V.; Rubin, B. S.; Sonnenschein, C.; Soto, A. M., Endocrine disruptors and reproductive health: The case of bisphenol-A. Mol Cell Endocrinol 2006, 254, 179-186.



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7. Centers for Disease Control and Prevention, Bisphenol A: Biomonitoring Summary. National Biomonitoring Program. 2013. 8. Kubwabo, C.; Kosarac, I.; Stewart, B.; Gauthier, B. R.; Lalonde, K.; Lalonde, P. J., Migration of bisphenol A from plastic baby bottles, baby bottle liners and reusable polycarbonate drinking bottles. Food Addit Contam A 2009, 26 (6), 928-937. 9. Rudel, R. A.; Camann, D. E.; Spengler, J. D.; Korn, L. R.; Brody, J. G., Phthalates, alkylphenols, pesticides, polybrominated diphenyl ethers, and other endocrine-disrupting compounds in indoor air and dust. Environ Sci Technol 2003, 37 (20), 4543-4553. 10. Dungchai, W.; Chailapakul, O.; Henry, C. S., Use of multiple colorimetric indicators for paperbased microfluidic devices. Analytica Chimica Acta 2010, 674 (2), 227-233. 11. Vandenberg, L. N.; Colborn, T.; Hayes, T. B.; Heindel, J. J.; Jacobs, D. R., Jr.; Lee, D. H.; Shioda, T.; Soto, A. M.; vom Saal, F. S.; Welshons, W. V.; Zoeller, R. T.; Myers, J. P., Hormones and endocrinedisrupting chemicals: low-dose effects and nonmonotonic dose responses. Endocrine reviews 2012, 33 (3), 378-455. 12. Wilson, N. K.; Chuang, J. C.; Lyu, C.; Menton, R.; Morgan, M. K., Aggregate exposures of nine preschool children to persistent organic pollutants at day care and at home. J Expo Anal Env Epid 2003, 13 (3), 187-202. 13. Geens, T.; Roosens, L.; Neels, H.; Covaci, A., Assessment of human exposure to Bisphenol-A, Triclosan and Tetrabromobisphenol-A through indoor dust intake in Belgium. Chemosphere 2009, 76 (6), 755-760. 14. Baxter, L. K.; Dionisio, K. L.; Burke, J.; Sarnat, S. E.; Sarnat, J. A.; Hodas, N.; Rich, D. Q.; Turpin, B. J.; Jones, R. R.; Mannshardt, E.; Kumar, N.; Beevers, S. D.; Ozkaynak, H., Exposure prediction approaches used in air pollution epidemiology studies: Key findings and future recommendations. J Expo Sci Env Epid 2013, 23 (6), 654-659. 15. Ornatska, M.; Sharpe, E.; Andreescu, D.; Andreescu, S., Paper Bioassay Based on Ceria Nanoparticles as Colorimetric Probes. Anal Chem 2011, 83 (11), 4273-4280. 16. Alkasir, R. S.; Ornatska, M.; Andreescu, S., Colorimetric paper bioassay for the detection of phenolic compounds. Anal Chem 2012, 84 (22), 9729-37. 17. von Lode, P., Point-of-care immunotesting: Approaching the analytical performance of central laboratory methods. Clin Biochem 2005, 38 (7), 591-606. 18. Martinez, A.; Phillips, S.; Butte, M.; Whitesides, G., Patterned Paper as a Platform for Inexpensive, Low-Volume, Portable Bioassays. Angewandte Chemie International Edition 2007, 46 (8), 1318-1320. 19. Sicarda, C.; Glenb, C.; Aubie, B.; Wallace, D.; Jahanshahi-Anbuhi, S.; Pennings, K.; Daigger, G. T.; Pelton, R.; Brennan, J. D.; Filipe, C. D. M., Tools for water quality monitoring and mapping using paperbased sensors and cell phones. Water Research 2015, 70. 20. Sharpe, E.; Bradley, R.; Frasco, T.; Jayathilaka, D.; Marsh, A.; Andreescu, S., Metal oxide based multisensor array and portable database for field analysis of antioxidants. Sensors and actuators. B, Chemical 2014, 193, 552-562. 21. Byrne, L.; Barker, J.; Pennarun-Thomas, G.; Diamond, D.; Edwards, S., Digital imaging as a detector for generic analytical measurements. Trac-Trend Anal Chem 2000, 19 (8), 517-522. 22. Sameenoi, Y.; Panymeesamer, P.; Supalakorn, N.; Koehler, K.; Chailapakul, O.; Henry, C. S.; Volckens, J., Microfluidic Paper-Based Analytical Device for Aerosol Oxidative Activity. Environ Sci Technol 2013, 47 (2), 932-940. 23. Dungchai, W.; Sameenoi, Y.; Chailapakul, O.; Volckens, J.; Henry, C. S., Determination of aerosol oxidative activity using silver nanoparticle aggregation on paper-based analytical devices. The Analyst 2013, 138 (22), 6766-6773.


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24. Cate, D. M.; Nanthasurasak, P.; Riwkulkajorn, P.; L'Orange, C.; Henry, C. S.; Volckens, J., Rapid Detection of Transition Metals in Welding Fumes Using Paper-Based Analytical Devices. Ann Occup Hyg 2014, 58 (4), 413-423. 25. Rattanarat, P.; Dungchai, W.; Cate, D. M.; Siangproh, W.; Volckens, J.; Chailapakul, O.; Henry, C. S., A microfluidic paper-based analytical device for rapid quantification of particulate chromium. Analytica Chimica Acta 2013, 800, 50-55. 26. Kumar, G.; Smith, P. J.; Payne, G. F., Enzymatic grafting of a natural product onto chitosan to confer water solubility under basic conditions. Biotechnol Bioeng 1999, 63 (2), 154-165. 27. Ispas, C. R.; Ravalli, M. T.; Steere, A.; Andreescu, S., Multifunctional biomagnetic capsules for easy removal of phenol and bisphenol A. Water Research 2010, 44 (6), 1961-1969. 28. Creek, K. L.; Whitney, G.; Ashley, K., Vacuum sampling techniques for industrial hygienists, with emphasis on beryllium dust sampling. J Environ Monitor 2006, 8 (6), 612-618. 29. (NIOSH), U. N. I. f. O. S. a. H., NIOSH Manual of Analytical Methods. Methods 7102 and 7300 1994, NIOSH, Cincinnati, OH, USA. 30. Lin, H. W.; Suslick, K. S., A Colorimetric Sensor Array for Detection of Triacetone Triperoxide Vapor. J Am Chem Soc 2010, 132 (44), 15519-15521. 31. Incorporated, A. S., The role of working spaces in Adobe applications. . 2006. 32. Shishkin, Y. L.; Dmitrienko, S. G.; Medvedeva, O. M.; Badakova, S. A.; Pyatkova, L. N., Use of a scanner and digital image-processing software for the quantification of adsorbed substances. J Anal Chem+ 2004, 59 (2), 102-106. 33. Corporation., w. v. c. e. e.-c. p. c.-s. h. T. V., 2014. 34. Calafat, A. M.; Weuve, J.; Ye, X. Y.; Jia, L. T.; Hu, H.; Ringer, S.; Huttner, K.; Hauser, R., Exposure to Bisphenol A and Other Phenols in Neonatal Intensive Care Unit Premature Infants. Environ Health Persp 2009, 117 (4), 639-644. 35. Vandenberg, L. N.; Hauser, R.; Marcus, M.; Olea, N.; Welshons, W. V., Human exposure to bisphenol A (BPA). Reprod Toxicol 2007, 24 (2), 139-177. 36. de Albuquerque, Y. D. T.; Ferreira, L. F., Amperometric biosensing of carbamate and organophosphate pesticides utilizing screen-printed tyrosinase-modified electrodes. Analytica Chimica Acta 2007, 596 (2), 210-221. 37. Szyrwinska, K.; Kolodziejczak, A.; Rykowska, I.; Wasiak, W.; Lulek, J., Derivatization and gas chromatography-low-resolution mass spectrometry of bisphenol A. Acta Chromatogr 2007, 18, 49-58. 38. Liao, C. Y.; Liu, F.; Guo, Y.; Moon, H. B.; Nakata, H.; Wu, Q.; Kannan, K., Occurrence of Eight Bisphenol Analogues in Indoor Dust from the United States and Several Asian Countries: Implications for Human Exposure. Environ Sci Technol 2012, 46 (16), 9138-9145. 39. Loganathan, S. N.; Kannan, K., Occurrence of Bisphenol A in Indoor Dust from Two Locations in the Eastern United States and Implications for Human Exposures. Arch Environ Con Tox 2011, 61 (1), 6873.

527 528 529



530

Page 24 of 31

Tables

531 Table 1. BPA concentrations found in household dust specimens at various sampling locations using the colorimetric biosensor and GC methods. Colorimetric Method [BPA]

[BPA]

[BPA]

GC Method

[BPA]

Location

SD

RSD

[BPA]

[BPA]

[BPA]

[BPA]

(µg/g)

(µg/g)

(µg/g)

(n=3)

Location

SD

RSD

0.41

0.1002

0.2423

1.92

2.00

0.1562

0.0781

1.92

2.06

2.01

0.0781

0.0389

2.70

2.48

2.95

2.71

0.2352

0.0868

CP-LR-H 2

3.23

2.60

2.75

2.86

0.3291

0.1151

0.0881

CO-LR-H2

0.75

0.54

0.95

0.75

0.2050

0.2746

0.0849

0.7714

CP-LR-H3

0.01

0.02

0.004

0.01

0.0113

0.9428

1.25

0.1877

0.1506

CP-FR-H3

0.84

0.61

0.73

0.73

0.1150

0.1583

0.11

0.23

0.1644

0.7046

CO-LR-H3

0.06

0.18

0.01

0.08

0.0874

1.0484

0.48

0.30

0.38

0.0917

0.2412

CP-FR-H1

0.39

0.46

0.24

0.36

0.1124

0.3094

2.90

3.15

3.51

3.19

0.3066

0.0962

CP-DC-R4

3.04

3.14

3.37

3.18

0.1692

0.0532

CP-DC-R2

2.05

1.93

2.24

2.07

0.1563

0.0754

CP-DC-R2

2.05

1.96

2.16

2.06

0.1002

0.0487

CP-DC-R1

3.87

3.51

3.69

3.69

0.1800

0.0488

CP-DC-R1

3.78

3.40

3.57

3.58

0.1904

0.0531

CP-DC-R5

2.24

2.72

2.42

2.46

0.2425

0.0986

CP-DC-R5

2.19

2.65

2.08

2.31

0.3024

0.1311

CP-DC-R6

3.33

3.15

2.96

3.15

0.1850

0.0588

CP-DC-R6

3.26

3.10

3.00

3.12

0.1311

0.0420

CP-DC-R7

1.51

1.75

1.81

1.69

0.1587

0.0939

CP-DC-R7

1.36

1.45

1.59

1.47

0.1159

0.0790

CP-DC-R5

1.63

1.57

1.45

1.55

0.0917

0.0591

CP-DC-R5

1.76

1.32

1.28

1.45

0.2663

0.1833

(µg/g)

(µg/g)

(µg/g)

(n=3)

CP-FR-H1

0.60

0.72

0.90

0.74

0.1510

0.2040

CP-FR-H1

0.31

0.42

0.51

CO-FR-H1

2.24

1.93

2.12

2.10

0.1563

0.0746

CO-FR-H1

1.90

2.18

CP-LR-H1

2.42

2.30

2.12

2.28

0.1510

0.0662

CP-LR-H1

2.05

CP-DR-H2

2.84

2.78

2.72

2.78

0.0600

0.0216

CP-DR-H2

CP-LR-H2

3.33

3.09

2.96

3.13

0.1877

0.0600

CO-LR-H2

1.02

0.96

1.14

1.04

0.0917

CP-LR-H3

0.09

0.17

0.05

0.11

CP-FR-H3

1.45

1.08

1.21

CO-LR-H3

0.17

0.42

CP-FR-H4

0.36

CP-DC-R4

CP: Carpet ; CO: Coach ; FR: Family Room ; LR: Living Room ; DR: Dinning Room ; H: Home ; DC: Day care center ; R: Room. Three different spots in flooring carpet or couch are selected for each sampling location.

532 533 534 535 536 23 | P a g e ACS Paragon Plus Environment

Page 25 of 31

537


Captions for Figures

538 539

Figure 1. Design of the colorimetric paper-based biosensing device for detection of BPA in

540

household dust specimens (A) Air sampling cassette with the paper-based sensor, (B) The

541

compact device included the test zone and the hose extension, (C) The sensing device connected

542

to the air sampling pump by a rubber tube, (D) The accumulation of BPA-dust particles on the

543

test zone, (E) The color change after the exposure to BPA-dust sample, (F) The scanned image

544

of the test zone clearly showed the formation of greenish color.

545 546

Figure 2. (A) The response calibration curve and dynamic range (inset) of the colorimetric

547

paper-based analytical device for detection of BPA in household dust (triplet analysis), (B)

548

Digital images of the tests zones of the colorimetric devices shows the colorimetric responses in

549

the range of 1-500 µg/g BPA (3 trials). The ﬁrst column shows a blank, which is the cleaned

550

household dust.

551 552

Figure 3. Histograms of red color channels of (A) Color ID EB 10-1, (B) sample 57, (C) color

553

ID 6004-1C, (D) blank_B1. The cleaned dust was used as a blank. Color ID is obtained from

554

color database; each histogram was analyzed using Adobe Photoshop. The x-axis of image

555

histogram represents the mean value of red color channel, while y-axis represents the number of

556

pixels.

557 558

Figure 4. Linear correlation between the colorimetric BioPAS and GC methods for the detection

559

of BPA in household dust specimens.



Page 26 of 31

560 561

Figure 5. Comparison between the colorimetric BioPAS and GC methods for detection of BPA

562

in household dust specimens at the same sampling locations.

563 564


Page 27 of 31

565


Figures

566 567 A

568

B

C

Test Zone of Colorimetric Biosensing Device

569 570 571 572 573 574

PVC Filter Membrane

575 576 577 578

F

E

D

579 580 581 582 583 584 585 586 587 588

Figure 1.

589 590 591 592 593 26 | P a g e ACS Paragon Plus Environment


Page 28 of 31

594 595 596 160 140

598

120 Color Intensity

597

599 600

y = 0.0919x +102.4 R2 = 0.9935

100

150

80 100

60 50

40

601

y = 16.486x + 10.129 R2 = 0.9968

20

602

0 0

20

40

60

0 0

603 604

100

200

300 [BPA] (µg/g)

400

500

600

A

605 606

1st trial

607 2nd trial

608 609 610

3rd trial

B

Blank

1

5

10

25

50

100

250

500 µg/g BPA

611 612 613

Figure 2.

614


Page 29 of 31


615 616 617 618 619 620 621

A

B

C

D

622 623 624 625 626 627 628 629 630 631 632 633

Figure 3.

634 635



Page 30 of 31

636 637 4.5 [BPA] µg/g ; Colorimetric Method

638 639 640 641 642 643

4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5

y = 0.9484x + 0.2508 R² = 0.9743

0.0

644

0.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0

[BPA] µg/ml ; GC Method

645 646 647

Figure 4.

648


Page 31 of 31


649 4.50

GC Method

Colorimetric Method

4.00 3.50

[BPA] (µg/g)

3.00 2.50 2.00 1.50 1.00 0.50 0.00

650 651 652 653

Figure 5.


Portable Colorimetric Paper-Based Biosensing ... - ACS Publications

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