Letter pubs.acs.org/jchemeduc
Comment on “Analysis of Citric Acid in Beverages: Use of an Indicator Displacement Assay” Krzysiek Konski,† Jessica Saw,† and Angel A. J. Torriero*,†,‡ †
School of Life and Environmental Sciences, Deakin University, Burwood, Victoria 3125, Australia Centre for Chemistry and Biotechnology, Deakin University, Burwood, Victoria 3125, Australia
‡
S Supporting Information *
ABSTRACT: This letter comments on the paper “Analysis of Citric Acid in Beverages: Use of an Indicator Displacement Assay” (J. Chem. Educ. 2010, 87 (8), 832−835). Discrepancies in figures and host:indicator complex behavior are discussed and an alternative experimental protocol presented. KEYWORDS: First-Year Undergraduate/General, Upper-Division Undergraduate, Laboratory Instruction, Analytical Chemistry, Physical Chemistry, Hands-On Learning/Manipulatives, Molecular Recognition, Noncovalent Interactions, UV−Vis Spectroscopy
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experiment introduces an innovative way to visualize the binding events between host and guest molecules, which is essential for students learning supramolecular chemistry
e read with interest the undergraduate laboratory experiment reported by Umali et al. in the August 2010 issue of this Journal.1 This informative laboratory
Figure 1. (A) Student data (2016) from the spectrophotometric titration with standard citric acid following Umali et al. protocol.1 Inset in part A: the respective calibration plot (n = 1). (B) UV−vis spectra of the cuvette solution (red) before and (blue) after the addition of MES buffer pH 5.5 (50:50 v/v methanol in water) following Umali et al. protocol.1 (C) Student data (2016) from the spectrophotometric titration with standard citric acid following the improved protocol presented in this letter. Inset in C: average calibration plot (n = 3) obtained following the improved protocol (error bars = ±1 standard deviation). Received: November 9, 2016 Revised: January 30, 2017 © XXXX American Chemical Society and Division of Chemical Education, Inc.
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DOI: 10.1021/acs.jchemed.6b00864 J. Chem. Educ. XXXX, XXX, XXX−XXX
Journal of Chemical Education
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concepts. Following the procedure provided by Umali et al., we implemented the experiment in an undergraduate practical class during 2015 and 2016. Only less than 10% of the total cohort (2015 = 112 students; 2016 = 100 students) obtained a linear (R2 > 0.970; n = 1−3) calibration plot. The best result obtained by students performing the procedure provided by Umali et al. in our laboratory is shown in Figure 1A. It should be noted that these results differ both in absorbance magnitude and peak wavelength (λmax) compared to Figures 3 and 4 presented by Umali et al.1 For example, an absorbance change at 503 nm of ca. 0.05 was observed after 1.2 mM citric acid was added. This is significantly different than the ∼0.18 change shown in Figure 3 of the published paper. Moreover, a signal with a λmax = 467 ± 5 nm, obtained by our students for the host:indicator binding, indicates a weaker interaction than that expected for a system free of guest molecules, which was given as λmax = 450 nm by Umali et al.1 Figure 1B shows that the dilution of the host:indicator complex in the MES (pH = 5.5; 50:50 v/v methanol in water) buffer may be responsible for the weak interaction observed. To address these issues, we modified the published protocol by preparing a cuvette solution composed only of solution B (indicator) and C (host) without the dilution used by Umali et al. (see the Supporting Information). Likewise, the titrant solution was also improved. It was prepared by diluting citric acid in MES (pH = 5.5; 50:50 v/v methanol in water) buffer without the addition of indicator and host. Due to these changes, it is very important to perform the titration using small volumes of titrant solution (in this case, 5 μL). Thereby, the change in the host and indicator concentrations in the cuvette solution with the addition of the titrant is small enough to have little impact on the measured absorbance. It was calculated that the concentration of the indicator decreases by 1.67% after 10 consecutive additions (total volume added = 50 μL) using the modified protocol. This implies that the error in the absorbance reading related to dilution effect is within the experimental error and smaller than ±1 standard deviation. Students from the 2016 cohort were randomly selected to participate in a trial of this new protocol. Figure 1C shows the results obtained for one student using the modified protocol, which resemble Umali et al.’s Figures 3 and 4 in regards to λmax, peak intensity, and the presence of a well-defined isosbestic point. The resulting linear plot is highly reproducible (Figure 1C, inset; n = 3), and it shows an increased sensitivity (0.092 absorbance units/mM) with respect to the results obtained in Figure 1A using the previously published protocol (0.045 absorbance units/mM). It is worth mentioning that all the selected students obtained similar outcomes. An absorbance change of 0.11 was observed after the addition of 1.2 mM citric acid using the new protocol, allowing observation of more significant absorbance changes after small concentrations of citric acid are added. The Umali et al. laboratory document previously published1 was modified (see Supporting Information) to incorporate the optimal conditions found in our laboratory with the aim of helping those who are experiencing our same problem.
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Letter
AUTHOR INFORMATION
Corresponding Author
*E-mail address:
[email protected]. ORCID
Angel A. J. Torriero: 0000-0001-8616-365X Notes
The authors declare no competing financial interest.
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REFERENCES
(1) Umali, A. P.; Anslyn, E. V.; Wright, A. T.; Blieden, C. R.; Smith, C. K.; Tian, T.; Truong, J. A.; Crumm, C. E.; Garcia, J. E.; Lee, S.; Mosier, M.; Nguyen, C. P. Analysis of Citric Acid in Beverages: Use of an Indicator Displacement Assay. J. Chem. Educ. 2010, 87 (8), 832− 835.
ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available on the ACS Publications website at DOI: 10.1021/acs.jchemed.6b00864. Student handouts and notes for the instructor (PDF, DOCX) B
DOI: 10.1021/acs.jchemed.6b00864 J. Chem. Educ. XXXX, XXX, XXX−XXX
