Surveying Iodine Nutrition Using Kinetic Spectrophotometry: An

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Surveying Iodine Nutrition Using Kinetic Spectrophotometry: An Integrative Laboratory Experiment in Analytical Chemistry for Population Health Adriana Nori de Macedo, Stellena Mathiaparanam, Ritchie Ly, and Philip Britz-McKibbin* Department of Chemistry and Chemical Biology, McMaster University, 1280 Main Street West, Hamilton, Ontario L8S 4M1, Canada S Supporting Information *

ABSTRACT: New experiments for undergraduate students are needed to stimulate experiential learning in the laboratory while providing valuable training for future career development. Iodine deficiency remains a major public health concern that is monitored by measuring the median urinary iodide concentration of a population. In this context, we have introduced a kinetic spectrophotometric experiment based on the classic Sandell−Kolthoff reaction for second-year undergraduate students in analytical chemistry. This two-day laboratory experiment incorporates principles of quantitative chemical analysis, redox chemistry, reaction kinetics, optical spectroscopy, sample pretreatment, external calibration, method validation, statistical analysis, and quality assurance. Additionally, students gain real-world experience of implementation of a reliable analytical method used in global health initiatives to combat iodine deficiency, including participation in a round-robin study with the CDC. This colorimetric assay is widely used for continuous monitoring of mandatory iodized table salt programs to prevent the risk of iodine deficiency disorders, including developmental delays and intellectual impairment in children. KEYWORDS: Analytical Chemistry, Laboratory Instruction, Second-Year Undergraduate, Hands-On Learning/Manipulatives, Inquiry-Based/Discovery Learning, Interdisciplinary/Multidisciplinary, Kinetics, UV−Vis Spectroscopy, Oxidation/Reduction, Quantitative Analysis

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INTRODUCTION Early experiences in the laboratory that address timely socioeconomic, health, and environmental problems are important components to a successful chemistry undergraduate research program.1 Recent curriculum renewal efforts in our department have included the development of stand-alone laboratory courses decoupled from traditional lectures, with emphasis on inquiry-based laboratory experiments that bridge the gap between explicitly prescribed experiments and independent senior thesis research projects.2 There is growing interest in renewing the curriculum to enhance the jobreadiness of undergraduate students as required for future careers in analytical chemistry within a regulatory environment in government or industry.3 Indeed, the top skill sets in chemistry-related sectors for employment of chemistry graduates not only require in-depth knowledge of modern instrumental techniques (LC−MS, GC−MS, UV−vis),4 but also training in method development and quality assurance/ quality control (QA/QC) along with effective communication skills and collaborative abilities as required for reliable decision making.5 Analytical chemistry plays an important role in preventative medicine based on the development of simple, sensitive, yet specific, methods for clinical diagnostic testing and populationbased screening. For instance, iodine is an essential inorganic micronutrient in human health that is required for biosynthesis © XXXX American Chemical Society and Division of Chemical Education, Inc.

of thyroid hormones. Inadequate iodine intake is associated with goiter and other adverse chronic health conditions, and it remains the most common preventable cause of neurocognitive impairment in children worldwide.6 As a result, public health policies have established table salt iodization programs as a cost-effective strategy to reduce the risk of iodine deficiency disorders that still impact about 30% of school-aged children globally.6 However, continuous monitoring is needed to ensure the efficacy of iodized table salt policies by measuring the median urinary iodide concentration (UIC) for detection of excessive or deficient intake of iodine in a population. It is important to note that iodine is an essential element used in the context of human nutrition, whereas iodide is the principal bioavailable species of iodine that is uptaken by the thyroid gland with excess excreted in urine. According to the World Health Organization (WHO), adequate iodine intake in a population is indicated by median UIC of 0.79−1.57 μM (100−199 μg/L).7 Although several methods have been developed for reliable UIC measurements, including inductively coupled plasma-mass spectrometry (ICP-MS) and capillary electrophoresis,8 kinetic spectrophotometry based on the Sandell−Kolthoff (S−K) reaction is widely used in epidemioReceived: September 14, 2017 Revised: April 3, 2018

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directly with CDC results for external validation in the context of an interlaboratory method comparison. The Supporting Information provides details for obtaining proficiency samples from the CDC. Alternatively, an instructor may opt to use artificial/synthetic urine samples spiked with iodide (refer to Supporting Information). For students who did not collect their own urine sample (about 15−20% of class), an extra CDC urine specimen was provided to them in the laboratory for analysis. Detailed instructions on a standardized urine collection procedure and disposal are described in the Supporting Information.

logical studies due to its low cost, robustness, and high sample throughput when using a microtiter robotic system.9 This colorimetric assay relies on the catalytic role of iodide for the reduction of ceric ions, Ce(IV), by arsenous acid/arsenite, As(III), under acidic conditions.9 In this case, heat digestion of urine samples is needed to avoid various chemical interferences, including endogenous reducing agents (e.g., thiocyanate, nitrite, ascorbic acid or ferrous ion), organic chelators of cerium, and spectral interferences (e.g., excess riboflavin due to vitamin supplementation).9 Herein we introduce a two-day integrative analytical chemistry experiment for second-year undergraduate students using the S−K reaction for assessment of iodine nutrition from random single-spot urine samples (i.e., a single/untimed urine specimen) collected from student volunteers, and/or proficiency urine test specimens provided by the Centers for Disease Control and Prevention (CDC).10 Students work in pairs to acquire an external calibration curve for iodide using the S−K kinetic spectrophotometric assay, and then determine UIC after sample pretreatment of urine specimens. Additionally, the median UIC of the class is calculated and classified on the basis of recommended iodine nutrition ranges from WHO guidelines. Results are also compared to urine specimens independently measured by ICP-MS at the CDC for quality assurance.

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Data Analysis and Calculations

Students are required to complete a series of prelab questions and tables in their notebook after reading the laboratory experiment (refer to Supporting Information) to assist in preparation of their calibrant solutions, and coordinate the acquisition of time-dependent absorbance measurements using conventional Genesys 20 spectrophotometers. After the first day, students process their data to generate external calibration curves for iodide determination using linear regression after performing blank replicate measurements in order to assess key analytical parameters (e.g., slope, linearity, precision). Students then determine the UIC for various urine samples using their external calibration curve, including calculation of the recovery of iodide after heat digestion, as well as interday precision for the blank performed over 2 days (n = 6). Each pair of students then submits a summary of their results in a report sheet the following week (refer to Supporting Information), which can be subsequently discussed in class to present the calculated median UIC and iodine nutritional status of students, as well as a comparison of UIC results from the round-robin trial with the CDC. It is important to note that this assay is not a diagnostic test for thyroid dysfunction on an individual level due to biological variations associated with recent iodine intake from the diet, hydration status, and diurnal cycle. As a result, this method is only used in the context of risk assessment of iodine deficiency in the population.11 It is recommended that instructors familiarize students with nonparametric statistics, including median, interquartile range (IQR), and Mann− Whitney U tests (equivalent to student’s t test) used in the case of skewed and non-normally distributed data frequently found in population health studies involving trace micronutrients. Also, the role of urinary iodide as a biochemical indicator of recent dietary iodine intake is emphasized, and students are also requested to record iodine-rich foods from their diet over 24 h in their report summary (e.g., seaweed, fish, dairy products, and food prepared with iodized table salt). Overall, a median UIC below 0.79 μM (or 100 μg/L) is associated with nutritional deficiency and classified as mild, moderate, or severe12 as summarized in Table 1.

EXPERIMENTAL OVERVIEW

Laboratory Practical

This integrative analytical laboratory experiment was designed to be optimal for 2 × 3 h laboratory periods (refer to Supporting Information). Although this was initially introduced as a single laboratory experiment in the Fall 2014, it was found that students working in pairs required additional time to troubleshoot and familiarize themselves with the S−K method, including preparation of standard solutions, coordination of the mixing and timing of reactions using trial runs, as well as preparation of heat digested urine specimens. As a result, in the Fall 2015 and 2017, we expanded this experiment over two periods with the first day devoted to the preparation of iodide calibrant solutions (as KI), acquisition of data for external calibration curve (10 min intervals at 420 nm), and assessment of technical precision using a reagent blank (n = 3). The second day is primarily dedicated to the processing of four urine samples after repeating blank technical replicates, including analysis of each student’s own urine specimen, a CDC proficiency test sample, and a spiked (1 μM iodide) urine sample to evaluate iodine recovery after heat digestion with ammonium persulfate for 30 min at 90 °C. Students work collaboratively and are directly involved in all preanalytical steps of this experiment, including sample collection and workup, and serial dilution of calibration standards. Also, students have the option to collect their own random/single-spot urine specimen in the morning prior to the start of the experiment using sterilized plastic tubes provided and/or analyze urine specimens provided by the CDC as part of their Ensuring the Quality of Urinary Iodine Procedures (EQUIP) program.10 These blinded urine specimens were independently measured by ICP-MS as the reference method used at the CDC, as well as various other methods (including variants of the S−K reaction) in over 120 different laboratories worldwide as part of a round-robin study.10 As a result, UIC data from students using the S−K reaction are compared

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HAZARDS The major hazards of the experiment include potential exposure to inorganic arsenic (i.e., arsenous acid) that is used as a reagent in the S−K reaction. All students receive stringent safety training and complete prelab questions prior to starting the experiment to ensure safe handling and disposal of solutions (e.g., inorganic waste container in fume hood) using disposable micropipet tips, as well as mandatory use of gloves, lab coats, and safety goggles while applying proper hygiene practices (i.e., washing of hands). Additionally, S−K reactions in this experiment were optimized using microliter B

DOI: 10.1021/acs.jchemed.7b00710 J. Chem. Educ. XXXX, XXX, XXX−XXX

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Table 1. WHO Recommendations for Assessing Iodine Nutrition in a Population among Nonpregnant Adults/ School Children12 Iodine Intake Deficient Insufficient Insufficient Adequate More than adequate Excessive

UIC in μM (μg/L)

Iodine Status

300)

Figure 1. Kinetic spectrophotometric method based on the S−K reaction for assessment of iodine nutrition in a population as an integrative laboratory experiment for second-year undergraduate students in analytical chemistry. This coupled redox reaction takes advantage of iodide-catalyzed reduction of ceric ion that results in a time-dependent decrease in absorbance response at 420 nm in the presence of arsenite under acidic conditions. This UV−vis spectral overlay can be acquired by students as an option with access to scanning UV−vis spectrophotometers in order to measure the firstorder kinetics for the S−K reaction, as well as defining an optimal time range where there is a linear absorbance response decrease ( 0.99) as shown in the Supporting Information. For the iodide-catalyzed reaction, the solution absorbance (at 420 nm measured after 10 min) is inversely related to the iodide concentration with a linear dynamic range from 0.3 to 3.3 μM. This external calibration curve is thus suitable for the determination of iodine nutritional status based on analysis of iodide in heat digested urine specimens as shown in Figure 2. The external calibration curve for iodide was pooled from a cohort of 130 undergraduate students in our Chemistry and Chemical Biology programs working in pairs using fresh batches of chemical reagents (arsenite, ceric ion) and primary stock solutions for iodide over two years, which demonstrates good method precision and robustness with an average RSD = 12% and equal variance (i.e., no weighting required in the linear

Figure 2. Pooled results of an external calibration curve for quantitative measurement of urinary iodide concentration from second-year undergraduate students working in pairs over two consecutive years (n = 65, error bars represent ±1σ) using the S−K reaction with kinetic spectrophotometry under standardized conditions. Urine samples require heat digestion with ammonium persulfate for 30 min at 90 °C in order to avoid chemical interferences prior to analysis by the S−K reaction for 10 min at room temperature.

regression model). Since this colorimetric assay is dependent on reaction conditions, careful preparation of iodide calibrant solutions and reagents together with consistent reaction timing is required. As students work together in pairs, good communication and organizational skills are also essential. In this case, all S−K reactions were performed at room temperature (≈21 °C) in order to simplify the overall procedure. On the first day of experiments, students first familiarize themselves with the S−K reaction by assessing their precision based on three independent trials for the blank. Then, student pairs proceed with subsequent acquisition of an external calibration curve once all iodide calibration solutions have been prepared. During the second laboratory period the following week, students prepare urine specimens that require heat digestion prior to the S−K reaction using ammonium persulfate as a safe oxidizing agent for matrix interferences.9 C

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Figure 3. External validation for urinary iodine measurements by a large cohort of undergraduate student pairs (n = 65) using kinetic spectrophotometry via the S−K reaction (McMaster University) as compared to ICP-MS (CDC) as reference method based on analysis of 11 different blinded urine samples. Each average urinary iodide measurement (■) was derived from 4−8 student pairs over a two-year period, whereas ICP-MS data were provided by the CDC in support of their EQUIP program. Overall, a modest mean bias (−16%) was detected from this roundrobin study when comparing UIC measured independently by two different analytical methods, where all data were located within upper/lower agreement limits (±2σ, p = 0.05).

Table 2. Summary of External Validation of S−K Kinetic Spectrophotometric Method for Urinary Iodine Determination by Undergraduate Students As Compared to ICP-MS Results from CDC’s EQUIP Program CDC Sample 608 613 623 643 650 657 667 668 669 671 696

S−K Assay Mean ±1σ (μM) (1.29 (2.18 (0.58 (0.86 (0.24 (1.81 (1.03 (0.87 (0.54 (0.99 (3.22

± ± ± ± ± ± ± ± ± ± ±

0.50) 0.98) 0.34) 0.48) 0.17) 0.56) 0.95) 0.82) 0.32) 0.63) 0.81)

RSD, % (Student Pairs, n) 38 45 59 56 73 31 92 93 58 64 25

(5) (4) (5) (4) (5) (5) (7) (7) (5) (6) (8)

CI at 95% (μM)

ICP-MS (μM) [Bias, %]a

0.68−1.91 0.63−3.74 0.22−0.94 0.09−1.64 0.02−0.46 1.10−2.51 0.15−1.92 0.12−1.63 0.15−0.94 0.33−1.67 2.54−3.90

1.93 2.71 0.70 1.16 0.51 2.83 0.77 1.08 0.30 1.53 2.42

[−33] [−20] [−17] [−26] [−53] [−36] [+34] [−19] [+81] [−35] [+33]

CDC Acceptable Range (95% CI)

Conclusions, Significant at p = 0.05

1.64−2.22 2.30−3.12 0.52−0.88 0.93−1.40 0.39−0.64 2.40−3.25 0.54−1.01 0.86−1.29 0.21−0.39 1.23−1.84 2.06−2.78

Outside CDC range Outside CDC rangeb Within CDC rangeb Outside CDC rangeb Outside CDC range Outside CDC range Outside CDC rangeb Within CDC rangeb Outside CDC rangeb Outside CDC rangeb Outside CDC rangeb

a

Bias calculated by relative difference of mean of S−K urinary iodide concentrations measured by students relative to the CDC reference method of ICP-MS for the same urine sample with an average overall bias of −11%. bSignificant at p = 0.05 based on technical variance of S−K assay even if data were outside the reported CDC acceptable range based on ICP-MS as reference method (95% CI).

undergraduate program. This experiment was introduced near the end of the term so that students had become sufficiently skilled at calibrant solution preparations, micropipeting, volumetric-based titrations, as well as statistical analysis/data interpretation. Overall, students generated largely consistent calibration curves with an average slope (sensitivity) of −(0.0734 ± 0.024) μM−1 with good linearity (R2 = 0.994) along with acceptable interday precision (n = 6) for blank samples measured over 2 days (median RSD = 5.6%, ranging from 0.2% to 33%). Also, the average recovery for 1 μM iodide spiked into pooled urine was about 82% (ranging from 49% to 162%). Additionally, good mutual agreement in UIC was demonstrated when comparing 11 proficiency test urine samples from EQUIP measured by students using the S−K reaction as compared to ICP-MS from the CDC with a mean bias of −16% (ranging from −70% to +60%).10 Figure 3 provides an overview of an interlaboratory method comparison when using a Bland−Altman (percent difference) plot, which highlights that all data were located within the upper and lower mutual agreement limits at a 95% confidence level (±2σ, p = 0.05).

Student pairs processed four different urine specimens, including a morning random/single-spot urine sample in order to determine the iodine nutritional status of the class, as well as a blinded urine specimen from the CDC for external validation. Additionally, students also perform a spike/recovery study in one of their own urine specimens to assess method accuracy. We have found that heat digestion of urine samples is preferably conducted in tightly parafilmed 1.5 mL microcentrifuge tubes on a compact heat block heater for better temperature control, which also allows for processing of large numbers of samples simultaneously while reducing reagent consumption. While urine samples undergo heat digestion at 90 °C for 30 min, students repeat blank measurements to determine method reproducibility (i.e., interday precision performed by both students). After cooling, aliquots of heattreated urine samples are then analyzed in the same way as blanks and iodide calibrants while following their standard operating procedure. Overall, students successfully completed this experiment over two laboratory periods (2 × 3 h); this was performed by two different cohorts of second-year Honor’s Chemistry (Fall 2015) and Honor’s Chemical Biology (Fall 2017) students in our D

DOI: 10.1021/acs.jchemed.7b00710 J. Chem. Educ. XXXX, XXX, XXX−XXX

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In fact, 8 of the 11 samples were found to have statistically equivalent mean UIC relative to ICP-MS when considering the technical variance of data generated by students (i.e., betweensubject variance for UIC determination as reflected by an RSD ranging from 25% to 93%) when performing the S−K method. This extent of technical variance is greater than interday precision for a blank given the additional steps involved with sample workup of heat digested/oxidized urine samples by a diverse cohort of sophomore students. This laboratory experience provides students unique insights into the rigors of external validation when evaluating the “trueness” of different analytical methods by participating in a robin-robin study. For instance, only 2 of the 11 samples satisfied the CDC’s reported acceptable range based on the lower technical variance of ICP-MS measurements likely performed by a single trained technician as the reference method for UIC as summarized in Table 2. A series of mandatory prelab questions were implemented to better prepare students before starting the experiment, whereas a final report was included in the experimental handout to ensure students were aware of the major objectives of their investigation, including interday assay precision, calibration curve (slope, linearity), spike/recovery and round-robin study (accuracy), as well as UIC results for the class as derived from individual urine samples (refer to Supporting Information). The median UIC determined for a cohort of young Canadian adults (average age of 20 years, n = 65) at McMaster University was found to be 0.85 μM (IQR ± 1.2 μM) with a range from 0.050 to 3.88 μM, which was consistent with adequate iodine nutrition for the population based on WHO criteria described in Table 1. A Shapiro−Wilks test confirmed that UIC measured for undergraduate students was not normally distributed (p = 2.05 × 10−4). As a result, a Mann−Whitney U test (two-tailed) was performed to determine whether there was a sex difference in recent dietary iodine intake (37 males, median UIC = 1.03 μM; 28 females, median UIC = 0.58 μM), which was concluded not to be statistically significant (p = 0.118) likely due to inadequate study power as a result of large biological variance. A larger sample size is thus required to detect a significant effect of sex-dependence on iodine intake. For instance, the iodine nutritional status of women is critical during the early stages of pregnancy in order to ensure proper fetal development. Recent evidence suggests that pregnant and lactating women have adequate iodine nutrition in countries with successful and sustained iodized salt programs.11 Figure 4 depicts the iodine nutritional status for sophomore students based on their measured UIC distribution, which highlights that collectively the group of students have adequate iodine intake (0.79−1.57 μM).

Figure 4. Summary of results from urinary iodide measurements using the S−K assay from random single-spot urine specimens collected from a cohort of second-year undergraduate students (n = 65; 37 males; 28 females; average age = 20 years) at McMaster University. The median UIC was 0.85 μM (mean = 1.10 μM), which is consistent with adequate iodine intake on a group level based on WHO criteria.

disorders, due its low operating costs with adequate selectivity, precision, and accuracy. However, several systematic errors were also apparent in this experiment as related to improper solution preparation (i.e., no sulfuric acid added), dilution errors for iodide calibrants, as well as overheating (>100 °C) or inadequate sealing of urine sample vials whose lids could snap open, which can be minimized by following a standard operating procedure with use of automation (e.g., robotic liquid handling system) on a microplate reader.9 Overall, this experiment provides a technically challenging yet meaningful real-world experience in analytical chemistry as applied to population health using inexpensive reagents and simple instrumentation. In fact, many undergraduate students are likely unfamiliar with the successful history of iodized table salt to prevent goiter and cretinism in iodine-deficient countries, as well as the need for continuous monitoring to prevent excessive iodine intake that may also contribute to hyperthyroidism.12 Students are reminded that UIC is not a diagnostic test per se and reflects only recent dietary intake of iodine that vary significantly from day to day based on time of collection, habitual diet, and hydration status.13 Repeat 24 h urine specimens from the same subject or random urine samples collected from a larger cohort provide more insight of iodine nutritional status for risk assessment of iodine deficiency on a population level.13 Overall, there was acceptable reproducibility based on repeated measurements of blanks and calibration curves over a wide range of iodide concentrations (RSD < 12%). Similarly, accuracy was deemed acceptable for a diverse cohort of sophomore undergraduate students over a two-year period when relying on manual solution preparations and timed kinetic reactions without temperature control as reflected by an average mean bias of −16% as compared to ICP-MS. Future efforts will aim at developing integrative laboratory experiments that provide students deeper appreciation of the important roles that quantitative chemical analysis plays in populationbased screening, preventative medicine, and clinical diagnostic testing. Alternative reducing agents that can replace the role of arsenous acid in the S−K reaction will be considered in future inquiry-based student projects with an emphasis on principles of green chemistry. An emphasis on providing students rigorous training in method validation, statistical analysis, QC/QA, and quality by design is also a critical aspect for future careers in analytical chemistry as required for reliable measurements that guide decision making and public health policy.

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REFLECTIONS The major learning objective of this experiment was to provide students a practical experience in the implementation of a kinetic spectrophotometric assay for measurement of low micromolar levels of iodide in complex human biofluids. Additionally, students are directly involved in method calibration, method validation, and nonparametric statistics while participating in an international round-robin study used for ensuring quality assurance for UIC. The kinetic spectrophotometric assay based on the S−K reaction is still widely used by accredited laboratories around the globe to monitor the efficacy of universal iodized table salt programs to ensure adequate nutrition for prevention of iodine deficiency E

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(8) (a) Shelor, C. P.; Dasgupta, P. K. Review of analytical methods for the quantification of iodine in complex matrices. Anal. Chim. Acta 2011, 702, 16−36. (b) Nori de Macedo, A.; Teo, K.; Mente, A.; McQueen, M. J.; Zeidler, J.; Poirier, P.; Lear, S. A.; Wielgosz, A.; BritzMcKibbin, P. A robust method for iodine status determination in epidemiological studies by capillary electrophoresis. Anal. Chem. 2014, 86, 10010−10015. (c) Nori de Macedo, A.; Macri, J.; Hudecki, P.; Saoi, M.; McQueen, M. J.; Britz-McKibbin, P. Validation of a capillary electrophoresis assay for monitoring iodine nutrition in populations for prevention of iodine deficiency: An inter-laboratory method comparison. J. Appl. Lab. Med. 2017, 1, 649−660. (9) (a) Sandell, E. B.; Kolthoff, I. M. Chronometric catalytic method for the determination of microquantities of iodine. J. Am. Chem. Soc. 1934, 56, 1426−1426. (b) Mina, A.; Favaloro, E. J.; Koutts, J. A robust method for testing urinary iodide using a microtitre robotic system. J. Trace Elem. Med. Biol. 2011, 25, 213−217. (c) Pino, S.; Fang, S.-L.; Braverman, L. E. Ammonium persulfate: a safe alternative oxidizing agent for measuring urinary iodine. Clin. Chem. 1996, 42, 239−243. (d) Haap, M.; Roth, H. J.; Huber, T.; Dittmann, H.; Wahl, R. Urinary iodine: comparison of a simple method for its determination in microplates with measurement by inductively-coupled plasma mass spectrometry. Sci. Rep. 2017, 7, 39835. (10) Caldwell, K. L.; Makhmudov, A.; Jones, R. L.; Hollowell, J. G. EQUIP: a worldwide program to ensure the quality of urinary iodine procedures. Accredit. Qual. Assur. 2005, 10, 356−361. (11) (a) Eastman, C. J. Screening for thyroid disease and iodine deficiency. Pathology 2012, 44, 153−159. (b) Bath, S. C.; Steer, C. D.; Golding, J.; Emmett, P.; Rayman, M. P. Effect of inadequate iodine status in UK pregnant women on cognitive outcomes in their children: results from the Avon Longitudinal Study of Parents and Children (ALSPAC). Lancet 2013, 382, 331−337. (c) Sullivan, K. M.; Perrine, C. G.; Pearce, E. N.; Caldwell, K. L. Monitoring the iodine status of pregnant women in the United States. Thyroid 2013, 23, 520−521. (12) (a) WHO/UNICEF/ICCIDD. Assessment of Iodine Deficiency Disorders and Monitoring Their Elimination: A Guide for Programme Managers, 3rd ed.; World Health Organization: Geneva, 2007. (b) McClure, R. D. Goitre prophylaxis with iodized salt. Science 1935, 82, 370−371. (13) (a) Rasmussen, L. B.; Ovesen, L.; Christiansen, E. Day-to-day and within-day variation in urinary iodine excretion. Eur. J. Clin. Nutr. 1999, 53, 401−407. (b) European Food Safety Authority. Scientific opinion on dietary reference values for iodine. EFSA J. 2014, 12, 3660.

ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available on the ACS Publications website at DOI: 10.1021/acs.jchemed.7b00710. Student experiment handout, report summary, TA notes, and reagents/equipment list (PDF, DOCX)

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. ORCID

Adriana Nori de Macedo: 0000-0002-4125-3755 Philip Britz-McKibbin: 0000-0001-9296-3223 Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS Financial support was provided by the Natural Sciences and Engineering Council of Canada (NSERC). The authors would like to thank Karen Neuman, Leah Allan, Trish Martin, and Kevin Wyszatko for support in running this experiment at McMaster University. The second-year undergraduate students in the Chemistry (2015) and Chemical Biology (2017) Honor’s program are also thanked for performing this experiment enthusiastically while providing helpful feedback.

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REFERENCES

(1) (a) Read, D. A U.K. approach to counter declining enrollment in chemistry and related disciplines at the university level. J. Chem. Educ. 2010, 87, 898−900. (b) Scott, A. Chemistry makes a comeback in UK universities, schools. Chem. Eng. News 2013, 91, 79−82. (c) Wenzel, T. J.; Larive, C. K.; Frederick, K. A. Role of undergraduate research in an excellent and rigorous undergraduate chemistry curriculum. J. Chem. Educ. 2012, 89, 7−9. (2) Bernard, E.; Gernigon, N.; Britz-McKibbin, P. Reservatrol photoisomerization: an integrative guided-inquiry experiment. J. Chem. Educ. 2007, 84, 1159−1161. (3) (a) Windsor, S. A. M.; Rutter, K.; McKay, D. B.; Meyers, N. Embedding graduate attributes at the inception of a chemistry major in a bachelor of science. J. Chem. Educ. 2014, 91, 2078−2083. (b) Claycomb, G. D.; Venable, F. A. Selection, evaluation, and modification of a standard operating procedure as a mechanism for introducing an undergraduate student to chemical research: A case study. J. Chem. Educ. 2015, 92, 256−261. (4) Headrick, K. L. Want ads, job skills, and curriculum: A survey of 1998 chemistry help-wanted ads. J. Chem. Educ. 2001, 78, 1281−1282. (5) (a) Bell, S. C.; Moore, J. Integration of quality assurance/quality control into quantitative analysis. J. Chem. Educ. 1998, 75, 874. (b) Schazmann, B.; Regan, F.; Ross, M.; Diamond, D.; Paull, B. Introducing quality control in the chemistry teaching laboratory using control charts. J. Chem. Educ. 2009, 86, 1085. (6) (a) Pearce, E. N.; Andersson, M.; Zimmermann, M. B. Global iodine nutrition: where do we stand in 2013? Thyroid 2013, 23, 523− 528. (b) Zimmermann, M. B.; Andersson, M. Update on iodine status worldwide. Curr. Opin. Endocrinol., Diabetes Obes. 2012, 19, 382−387. (c) Zimmermann, M. B.; Moretti, D.; Chaouki, N.; Torresani, T. Development of a dried whole-blood spot thyroglobulin assay and its evaluation as an indicator of thyroid status in goitrous children receiving iodized salt. Am. J. Clin. Nutr. 2003, 77, 1453−1458. (7) World Health Organization. Recommended iodine levels in salt and guidelines for monitoring their adequacy and effectiveness. Nutrition Unit, Division of Food and Nutrition; WHO: Geneva, 1996. F

DOI: 10.1021/acs.jchemed.7b00710 J. Chem. Educ. XXXX, XXX, XXX−XXX