Laboratory Experiment pubs.acs.org/jchemeduc
Quantitative Analysis of Bisphenol A Leached from Household Plastics by Solid−Phase Microextraction and Gas Chromatography− Mass Spectrometry (SPME−GC−MS) Bettie Obi Johnson,* Fernanda M. Burke, Rebecca Harrison, and Samantha Burdette Department of Chemistry, University of South Carolina Lancaster, Lancaster, South Carolina 29720, United States S Supporting Information *
ABSTRACT: The measurement of trace levels of bisphenol A (BPA) leached out of household plastics using solid-phase microextraction (SPME) with gas chromatography−mass spectrometry (GC−MS) is reported here. BPA is an endocrine-disrupting compound used in the industrial manufacture of polycarbonate plastic bottles and epoxy resin can liners. This experiment allows students to use modern instrumentation and analytical techniques to investigate a timely and relevant issue involving the contamination of food products by a packaging component. The impact of handling conditions and container type on the quantity of BPA released from the plastics is explored. This experiment is suitable for an undergraduate analytical chemistry or instrumental analysis course and requires two, 3-h laboratory periods to complete. KEYWORDS: Upper-Division Undergraduate, Analytical Chemistry, Laboratory Instruction, Hands-On Learning/Manipulatives, Problem Solving/Decision Making, Consumer Chemistry, Gas Chromatography, Mass Spectrometry, Student-Centered Learning, Toxicology
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resins.6 The structures for these polymers are illustrated in Figure 1. Polycarbonate plastics have been used to make products such as plastic bottles, storage containers, CDs, and dental fillings. Epoxy resins are used for the linings in soda cans and vegetable tins. BPA has been found to migrate out of these plastics, especially under severe conditions.7,8 BPA levels detected in water from these containers ranged from 2.4 to 14.3 μg/L under normal use conditions and up to 228−521 μg/L after heating at 70 °C for six days.8 The levels varied based on extraction time, extraction temperature, container dimensions, water volume, and the quantity of residual BPA present in the plastic bottle. BPA has also been detected at low μg/L levels in a variety of prepackaged products including canned liquid infant formulas, soft drinks, and foods.9−11 The issue of chemical monomers, plasticizers, and other chemicals migrating out of plastic packaging into foodstuffs is a common concern for manufacturers as they attempt to choose an appropriate packaging material for a given food.12 Although BPA has been used for over 50 years, its use has become controversial in recent years due to its established role as an endocrine disruptor, a chemical that mimics hormones in the body and disrupts normal functions.7,13 Population studies have shown that most people in the developed world have partper-billion levels of BPA in their body fluids, arising primarily from the consumption of foods and beverages that leach BPA from their packaging.14,15 The FDA has estimated average exposures to BPA at 2.42 μg/kg/day for infants and 0.185 μg/ kg/day for adults in the United States.16 These exposures are well below the FDA “no observed adverse effect level”
he incorporation of laboratory experiments that have realworld relevance has been shown to increase student engagement in lab.1−4 This can be accomplished by introducing experiments in areas such as pharmacology,2 environmental science,3 and forensics.4 Using these context-based experiments improves student attitudes toward lab and increases their interest in analytical chemistry.1 Once students are engaged in an experiment, they are better able to learn new laboratory techniques, develop critical thinking skills, and begin designing aspects of their experiments; all student learning outcomes desired by college chemistry faculty.5 In this experiment, students are asked to assess the degree to which the chemical monomer, bisphenol A (BPA), is leached from containers that are stored and treated under a variety of harsh conditions. Owing to BPA’s recent and extensive coverage in the news media, many students are familiar with and interested in this topic. BPA (Figure 1) is an aromatic compound used in the commercial preparation of polycrbonate plastics and epoxy
Figure 1. Chemical structures of bisphenol A (top), polycarbonate polymer (middle), and epoxy resin polymer (bottom). © 2012 American Chemical Society and Division of Chemical Education, Inc.
Published: August 14, 2012 1555
dx.doi.org/10.1021/ed2003884 | J. Chem. Educ. 2012, 89, 1555−1560
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(NOAEL) of 5,000 μg/kg/day and the acceptable daily reference dose level (RfD) of 50 μg/kg/day. However, the FDA stated in a January 2010 report that although the current levels of BPA exposure are safe, there is some concern about the potential low-level effects of BPA on the brain, behavior, and development of fetuses, infants, and young children.17 Further studies are underway to clarify these potential effects. In a 2008 study of 1455 adults aged 18−74 years, higher concentrations of BPA in urine were associated with cardiovascular disease, diabetes, and high levels of certain liver enzymes.18 In response to these concerns, regulating agencies and select corporations have taken actions to limit the use of BPA in consumer food packaging products.17 Various techniques have been used to measure BPA levels directly in plastics as well as in liquids contained inside the plastics. These methods often include an extraction or concentration step such as liquid−liquid extraction, solid− phase extraction (SPE) or solid−phase microextraction (SPME) followed by an analysis step using gas chromatography−mass spectrometry (GC−MS), liquid chromatography− mass spectrometry, fluorescence, or electrochemical detection.19,20 One such experiment published in this Journal involves the identification of part-per-million levels of BPA in water samples using SPE followed by derivatization and GC− MS detection.21 The experiment presented here differs from the previously published experiment in that the analytical samples are plastic leachate solutions containing lower part-perbillion levels of BPA. In addition, SPME is used instead of SPE and an internal standard calibration is employed. SPME sampling was chosen because it provides a relatively simple and inexpensive method for extracting and concentrating lowlevel organic compounds out of aqueous solutions. GC−MS detection was used because of its wide availability in college laboratories and the sufficient level of volatility of BPA for this technique. The use of the mass spectrometer allows low limits of detection and the ability to separate BPA from other trace compounds by using selected ion monitoring (SIM). Previous studies have demonstrated the SPME−GC−MS method is capable of detection limits of BPA between 0.04 and 1.0 μg/L with good reproducibility in the range of 8% relative standard deviation.20 Various quantitative analysis experiments utilizing SPME combined with GC−MS have been published in this Journal including cinnamaldehyde in cinnamon,22 acrolein and acrylonitrile in environmental water samples,23 nicotine and cotitine in urine and sputum,24 bromoform in swimming pool water,25 and caffeine in beverages.26 In these experiments, the analytes are typically determined at part-per-million or high part-per-billion levels. In contrast, BPA in this experiment is present at less than 100 parts-per-billion in the leachate solutions. Detecting an analyte at these low levels presents the analytical chemist with additional challenges that can be overcome by the application of specific techniques to address each source of variation. For example, SIM is used to enhance the analyte’s signal and to avoid contaminant signals. The SPME fiber is thermally conditioned for the entire run time to avoid analyte carryover between runs. An internal standard is used to compensate for the variation in analyte recovery in the SPME sampling and desorption steps. These procedures help students learn how measurement variability can be addressed in the development of an analytical method.
Laboratory Experiment
LEARNING OUTCOMES After completing this experiment, students should be able to (1) select appropriate experimental conditions for an analytical test method; (2) prepare calibration standards using an internal standard; (3) extract trace level organic contaminants in an aqueous sample with SPME; (4) use GC−MS to measure BPA concentration; and (5) calculate BPA exposure levels from experimental results to evaluate potential impact on human health.
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EXPERIMENTAL PROCEDURE
Overview
The experiment was conducted in quantitative analysis and organic chemistry laboratories on a small regional campus with approximately 1,700 students. Second-year students majoring in chemistry, biology, and preprofessional programs were enrolled in these classes, which ranged in size from 4 to 12 students per class. This experiment was conducted in two adjacent laboratories: a chemistry lab where standards and samples were prepared and an instrument room where the Fourier transform infrared (FTIR) and SPME−GC−MS analyses were performed. This experiment requires two, 3-h laboratory sessions with students working in groups of 2−4 students. In the first week, students assemble the SPME fibers, set up the GC−MS method, prepare stock solutions and calibration standards, verify plastic composition by FTIR, design sample treatment plans, and treat the plastic and control sample containers. In the second week, students run the calibration standards and sample solutions by SPME−GC−MS, set up calibration curves, and quantify their results. If students need additional time to complete the calibration and data analysis, this can be done outside of the normal laboratory sessions. As part of their reports, students evaluate the accuracy and precision of the method and the impact of treatment conditions and container type on the quantity of BPA leached. They also calculate human exposure levels based on their measured BPA concentrations. Standard Preparation
All chemical reagents were obtained from Fisher Scientific Company. Reagent grade bisphenol A (Fisher AC15824) and 2-phenylphenol (Fisher AC13076) were used as received to prepare the stock solutions in GC-Resolv grade methanol (Fisher A457−4). ACS Reagent grade water (Fisher 91511) was used to prepare the diluted calibration standards and sample solutions. Separate stock solutions of 100,000 μg/L 2phenylphenol (2PP) and 100,000 μg/L bisphenol A (BPA) were prepared by diluting 0.0100 g of each to 100.0 mL with methanol. Calibration standards were prepared in 15 mL SPME tubes by dispensing 10.0 mL of reagent grade water into each tube and then adding the appropriate microliter volume of each stock solution to achieve the desired concentration. The final concentrations for the aqueous calibration standards ranged between 1 and 100 μg/L. Sample Preparation
Plastic containers with the #7 recycling code were chosen for the sample containers. The #7 recycling code represents the “other” category of plastics that includes acrylic, acrylonitrile butadiene styrene, fiberglass, nylon, polycarbonate, and polylactic acid. Students could use containers from home (older water bottles and baby bottles), purchase containers 1556
dx.doi.org/10.1021/ed2003884 | J. Chem. Educ. 2012, 89, 1555−1560
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Laboratory Experiment
from the fiber to prevent carryover between injections. The effectiveness of this cleansing method for the SPME fiber was demonstrated by injecting blank fibers after calibration standard runs. The blank fibers produced little or no detectable 2PP or BPA, with area counts of less than 5,000 (
