Determining Iron(III) Concentration in a Green Chemistry Experiment

Laboratory Experiment Cite This: J. Chem. Educ. XXXX, XXX, XXX−XXX

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Determining Iron(III) Concentration in a Green Chemistry Experiment Using Phyllanthus emblica (Indian Gooseberry) Extract and Spectrophotometry Parawee Rattanakit* and Rasimate Maungchang School of Science, Walailak University, Nakhon Si Thammarat 80160, Thailand

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S Supporting Information *

ABSTRACT: A laboratory experiment utilizing locally available Indian Gooseberry (Phyllanthus emblica Linn.) extract as a bioreagent to determine iron(III) is described with the goal of providing students with hands-on experience in spectrophotometry and green chemistry. Spectrophotometry was used to measure the concentration of the complex formed between iron(III) and gallic acid, an active compound found in Indian Gooseberry extract. Excellent results from the quantitative analysis of iron(III) in water were achieved. In this case, the bioreagent was used to replace the conventional hazardous chemicals that are usually employed as chromogenic reagents. The use of Indian Gooseberry extract is simple, inexpensive, and environmentally friendly. This experiment is appropriate for use as a simple spectrophotometric protocol for undergraduate teaching in an analytical- or generalchemistry laboratory. KEYWORDS: Second-Year Undergraduate, Analytical Chemistry, Environmental Chemistry, Hands-On Learning/Manipulatives, Green Chemistry, UV−Vis Spectroscopy

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INTRODUCTION Green chemistry, also known as environmentally benign chemistry or clean chemistry, is a new chemistry that has been extensively developed over the past two decades. This new approach aims to reduce or, ideally, eliminate the use and generation of substances hazardous to human health and the environment; to make laboratory practices more environmentally friendly and cost-effective; and also to improve laboratory safety. To achieve this goal, development and implementation of chemical products and processes need to be made.1−3 Spectrophotometric laboratories are mandatory for students in science and health science. Students learn how to use spectrophotometers to determine the amount of a substance in various sample solutions. For example, the detection of iron(III) is determined by the absorbance of a solution after mixing with a chromogenic reagent that is typically toxic or expensive, such as 2,2′-bipyridine,4 thiocyanate,5 glycine,6 8-hydroxyquinoline,7 2(5-bromo-2-pyridylazo)-5-diethylaminophenol (5-BrPADAP),8 1,2-dihydroxy-3,4-diketocyclo-butene,9 or 1,10phenanthroline.10 In a college or university where many students conduct a basic spectrophotometry laboratory experiment every year, a large amount of toxic waste is generated, leading to high wastetreatment costs. To reduce these problems, we introduced a green chemistry experiment that is safer, simpler, and cheaper and uses fewer chemicals. Several publications on green © XXXX American Chemical Society and Division of Chemical Education, Inc.

laboratory experiments have been proposed in many branches of chemistry.11−15 For example, liquid CO2 has been used instead of a solvent in the extraction of limonene from orange rind,11 and gold nanoparticles have been synthesized using tea extract.14 In this experiment, we have developed a spectrophotometric method for detecting iron(III) using Indian Gooseberry extract as a green reagent to replace 1,10-phenanthroline,16 a conventional hazardous chemical that we previously used. The main active compound in the extract is gallic acid, which reacts with iron(III) to form an Fe−polyphenol complex.17 The proposed mechanism of complex formation suggests a 2:1 complex of gallic acid with iron(III), with two water molecules acting as coligands to complete the octahedral coordination sphere of the iron−(gallic acid)2 complex (see Figure 1). The Indian Gooseberry extract can be purchased as a tea powder at any herbal medicine store in Thailand as well as widely throughout Asia and online. It is normally used to relieve coughing. A package costs about $1 and contains 20 small tea packs. No extraction and purification are needed in the preparation step, which eliminates the use of a toxic solvent. This is the first time green chemistry has been used in a student laboratory in our university. Creating a simple laboratory Received: October 6, 2018 Revised: March 3, 2019

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DOI: 10.1021/acs.jchemed.8b00817 J. Chem. Educ. XXXX, XXX, XXX−XXX

Journal of Chemical Education

Laboratory Experiment

Figure 1. Formation of the gallic acid−iron(III) complex.

unknowns were tap water or spiked water samples that could be conveniently made from a standard iron(III) solution after appropriate dilution. The acetate buffer at pH 5.6 and the unknown solutions were prepared by the instructor. The acetate buffer was used to control the pH of the solution, which is important for stable complex formation between the iron(III) ions and the ligands. In their groups, students helped each other prepare solutions of the Indian Gooseberry extract, serial-dilution solutions from an iron(III) standard solution (50 mg L−1), and sample solutions. The bioreagent was prepared by adding a small bag of tea powder (1.0 g) into 150 mL of deionized water. The solution was left for 10 min at room temperature and mixed well before removal of the tea bag. The extract solution was used without any further filtration or purification. For spectrophotometric detection, students mixed 15.00 mL of the acetate buffer, 15.00 mL of the Indian Gooseberry extract, and an appropriate volume of iron(III) solution into a 50.00 mL volumetric flask, with the final volume adjusted with DI water. For the unknown solution, the mixture was prepared as above, but the iron(III) solution was replaced by 5.00 mL of the unknown sample. In all cases, the mixture was left for 10 min to ensure complete reaction. For absorbance measurements, students calibrated the UV−vis spectrophotometers by setting the absorbance to 0.000 for the blank extract solution. Then, the absorbance was recorded over the range of 400−750 nm in order to obtain the maximum wavelength of the complex using 2.50 mg L−1 iron(III), the maximum concentration used. After fixing the maximum wavelength, each group prepared their own standard calibration curve over an iron(III)-concentration range of 0.50−2.50 mg L−1. Data analysis was performed using MS Excel. The curve should follow the Beer−Lambert law and fit the data well. A group was not allowed to move on until their R2 value was over 99%, and this should be emphasized by the instructor to ensure that students are sufficiently careful in solution preparation. Each group used the calibration curve to obtain the iron(III) concentration of the given unknown solution. The experimental details are fully documented in the Supporting Information. After completing the experiment, all groups submitted their laboratory reports to the instructor. To check the accuracy of the students’ experiment, the instructor compared the concentration of iron(III) in each group’s unknown to the known concentration and then handed the reports back to the students. Other learning outcomes were assessed from students’ reports using the lab-report rubric provided in the Supporting Information.

procedure that integrates spectroscopy and green chemistry into our analytical-chemistry-laboratory curriculum was initially a challenge. The experiment allows students to prepare the complex solution formed between Indian Gooseberry extract and iron(III), which produces a color change that can be measured by UV−vis spectroscopy to determine the amount of iron(III) in the water samples. Instructors may find the framework useful for teaching the concepts of green chemistry, molecular absorption in UV−vis spectroscopy, and quantitative analysis of iron(III) in water samples. The proposed method is a simple, inexpensive, and environmentally friendly procedure for iron(III) detection and satisfies all the requirements of green chemistry.

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LEARNING OUTCOMES After completing this experiment, students should be able to • prepare a solution of desired concentration using a pipet and dilution techniques together with basic concentration calculations, • create a calibration graph presenting the relationship between the concentration of a substance of interest and the absorbance of the solution, • interpolate the concentration of a substance of interest using the created calibration curve, • explain how and why the Beer−Lambert law is used to determine concentration and the limitations of the Beer− Lambert law, • explain the reason the solution changes color after mixing with the Indian Gooseberry extract solution, and • appreciate the use of Indian Gooseberry extract as a green reagent in the determination of iron(III) concentration.

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MATERIALS All chemicals, except the Indian Gooseberry extract, were of analytical-reagent grade and used without any further purification. Deionized water was used in the preparation of all aqueous solutions. Sodium acetate (CH3COONa) was obtained from Loba Chemie Pvt. Ltd. Acetic acid (CH3COOH) and a standard solution of 1000 mg L−1 iron(III) nitrate solution in 0.5 M HNO3 were purchased from Merck. Commercial Indian Gooseberry tea powder was purchased from a local herbal medical store in Nakhon Si Thammarat, Thailand. The analysis was performed using a Halo UV−vis 20 spectrometer (Dynamica Scientific Ltd.) with a 1.0 cm width glass cuvette. The pH was measured using a pH meter (Starter 3100, Ohanus) equipped with a combined glass electrode.

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HAZARDS The iron(III) standard solution may cause skin irritation and strong eye irritation. The use of personal protective equipment including laboratory coats, gloves, and goggles is required throughout the experiment.

EXPERIMENTAL PROCEDURE The experiment was 3 h long and consisted of 12 groups of 2−3 students. Each group was given a small pack of Indian Gooseberry tea powder and one unknown water sample. The B



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RESULTS AND DISCUSSION In this traditional experiment, an Indian Gooseberry aqueous extract was utilized as a green complexing agent for iron(III) determination. Gallic acid, present in Indian Gooseberry, is responsible for chelating with the iron(III) ion.17 Mixing of iron(III) with the extract in an acetate buffer at pH 5.6, leads to an easily observable color change in the solution from light yellow to dark purple, indicating complex formation (Figure 2).

various iron(III) concentrations are added to the extract, as shown in Figure 3 (left). A representative calibration curve obtained by a group of students is presented in Figure 3 (right). As expected, the absorbance was directly proportional to iron(III) concentration, with an excellent linear relationship of over 0.99. Next, the data was used to determine the amount of iron(III) in the unknown sample. Typical results compared well with the actual values, and the percentage errors were less than 10%. Table 1 summarizes the results of the students who performed the experiment in 2017. A discussion about students’ performance follows. About 11% of students needed to repeat the making of the solutions because they had R2 values less than 99%. These students used about 45 min more than the others, but they were able to complete the experiment within 3 h. About 90% of these students understood why they had an error and discussed it in their lab report, whereas about 10% did not describe any error sources in the report. Less than 5% of all students did not get the correct unknown concentration. This error could be from methodological error or personal error. For example, students may have done a wrong calculation, pipetted chemicals with a wrong volume, not left the mixed solutions long enough (10 min before use), or not mixed the solutions in the order indicated in the instructions. Common mistakes and advice are also provided in the Supporting Information.

Figure 2. UV−vis spectrum of the complex. Inset is the solutions of Indian Gooseberry extract and its complex with iron(III) in an acetate buffer at pH 5.6.

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IMPLEMENTATION AND CONCLUSION The 3 h experiment described herein was designed for secondyear undergraduate students who take analytical chemistry classes at Walailak University, Thailand. There were approximately 400 students from the Chemistry, Health Science, and Agricultural departments. Students worked in groups of two or three. The objectives in designing this experiment were (1) to teach students the basic principles of spectrophotometry and its practical applications as they use the method to determine the amount of iron(III) in water samples and (2) to introduce students to the concept of green chemistry by substituting natural reagents for some hazardous chemicals. Because many undergraduate students have limited experience in a chemistry laboratory, employing natural extracts as reagents in the laboratory greatly reduces the risks of the experiment. The economic benefit of using a natural reagent is also significant.

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A spectral scan of the complex formed by a 2.50 mg L iron(III) solution from 400 to 750 nm allowed the students to identify the maximum absorption wavelength. A representative absorption spectrum obtained by a group of students is shown in Figure 2. In this example, the spectrum exhibits a maximum absorption at 570 nm. The results of all groups revealed that the maximum wavelength varied in the range from 560 to 570 nm, depending on the UV−vis spectrophotometer. However, the absorbance of the complex in all groups was about the same at approximately 0.20. After each group found the maximum absorption wavelength, they used it to construct a standard calibration curve by recording the absorbance of the complex at different iron(III) concentrations. At this stage, students should notice a change in the solution color to different shades of dark purple when

Figure 3. (Left) Digital image demonstrating the color change in the solution with iron(III) concentration. (Right) Sample student calibration curve for iron(III) determination using Indian Gooseberry extract and spectrophotometric detection. C




Table 1. Comparative Student Experimental Results Iron(III) Concentration, mg L−1

Student Group, N

Iron(III) Determined (Average), mg L−1

Standard Deviation

Recovery,%

Relative Error, %

5.00 15.00 25.00

39 38 44

4.81 13.79 23.85

0.62 1.57 1.46

96.20 91.93 95.40

3.80 8.07 4.60

acknowledge the time and valuable suggestions of the reviewers who helped to improve the article.

The cost of running the green chemistry laboratory at Walailak University was 100 times less than that of the traditional laboratory in which 1,10-phenanthroline was used as a complexing reagent for iron(II) determination. We found that the inclusion of a green experiment into the curriculum provides a platform for the discussion of environmental problems in the classroom. Because it is simple, environmentally friendly, and cost-effective, this method can be used as a green-laboratory model for undergraduate teaching in an analytical-chemistry course. The method can also be extended to other courses, such as general and physical chemistry, where an inquiry-based approach may be used. Moreover, students can bring their own water samples for testing. Although this experiment was concerned with water analysis, other applications (e.g., food, pharmaceutical, agricultural, clinical, and environmental analysis) are possible. Lastly, we would like to discuss the possibility of modifying this investigation for a home science experiment or a simple screen for iron(III). Using a very simple setup, one can use only Indian Gooseberry extract and a water sample with a suspected excessive amount of iron(III) to form a complex. However, the observation of the color change should be done within 10−15 min because the stability of the complex is low in the absence of the acetate buffer, and the color will fade more rapidly. This method is similar to the use of guava leaves in iron detection by some northern Thai villagers when they put the leaves in groundwater and observe the change in water color to a darker tone if a high iron concentration is present.18

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ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available on the ACS Publications website at DOI: 10.1021/acs.jchemed.8b00817.

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REFERENCES

(1) Anastas, P. T.; Kirchhoff, M. M. Origins, Current Status, and Future Challenges of Green Chemistry. Acc. Chem. Res. 2002, 35 (9), 686−694. (2) Hjeresen, D. L.; Boese, J. M.; Schutt, D. L. Green Chemistry and Education. J. Chem. Educ. 2000, 77 (12), 1543−1547. (3) Haack, J. A.; Hutchison, J. E. Green Chemistry Education: 25 Years of Progress and 25 Years Ahead. ACS Sustainable Chem. Eng. 2016, 4 (11), 5889−5896. (4) Bopegedera, A. M. R. P.; Coughenour, C. L.; Oswalt, A. J. Quantitative Determination of Iron in Limonite Using Spectroscopic Methods with Senior and General Chemistry Students: GeologyInspired Chemistry Lab Explorations. J. Chem. Educ. 2016, 93 (11), 1916−1922. (5) Nyasulu, F.; Barlag, R. Colorimetric Determination of the Iron (III)−Thiocyanate Reaction Equilibrium Constant with Calibration and Equilibrium Solutions Prepared in a Cuvette by Sequential Additions of One Reagent to the Other. J. Chem. Educ. 2011, 88 (3), 313−314. (6) Prasad, R.; Prasad, S. Spectrophotometric Determination of Iron (III)−Glycine Formation Constant in Aqueous Medium Using Competitive Ligand Binding. J. Chem. Educ. 2009, 86 (4), 494−497. (7) Adebayo, B. K.; Ayejuyo, S.; Okoro, H. K.; Ximba, B. J. Spectrophotometric Determination of Iron (III) in Tap Water Using 8Hydoxyquinoline as a Chromogenic Reagent. Afr. J. Biotechnol. 2011, 10 (71), 16051−16057. (8) Filik, H.; Giray, D. Cloud Point Extraction for Speciation of Iron in Beer Samples by Spectrophotometry. Food Chem. 2012, 130 (1), 209− 213. (9) Stalikas, C. D.; Pappas, A. Ch.; Karayannis, M. I.; Veltsistas, P. G. Simple and Selective Spectrophotometric Method for the Determination of Iron (III) and Total Iron Content, Based on the Reaction of Fe (III) with 1, 2-Dihydroxy-3, 4-Diketocyclo-Butene (Squaric Acid). Microchim. Acta 2003, 142 (1−2), 43−48. (10) Jones, D. R.; Jansheski, W. C.; Goldman, D. S. Spectrophotometric Determination of Reduced and Total Iron in Glass with 1, 10Phenanthroline. Anal. Chem. 1981, 53 (6), 923−924. (11) Buckley, H. L.; Beck, A. R.; Mulvihill, M. J.; Douskey, M. C. Fitting It All In: Adapting a Green Chemistry Extraction Experiment for Inclusion in an Undergraduate Analytical Laboratory. J. Chem. Educ. 2013, 90 (6), 771−774. (12) Graham, K. J.; Jones, T. N.; Schaller, C. P.; McIntee, E. J. Implementing a Student-Designed Green Chemistry Laboratory Project in Organic Chemistry. J. Chem. Educ. 2014, 91 (11), 1895− 1900. (13) Jones, D. R.; DiScenza, D. J.; Mako, T. L.; Levine, M. Environmental Application of Cyclodextrin Metal−Organic Frameworks in an Undergraduate Teaching Laboratory. J. Chem. Educ. 2018, 95 (9), 1636−1641. (14) Sharma, R. K.; Gulati, S.; Mehta, S. Preparation of Gold Nanoparticles Using Tea: A Green Chemistry Experiment. J. Chem. Educ. 2012, 89 (10), 1316−1318. (15) van Opstal, M. T.; Nahlik, P.; Daubenmire, P. L.; Fitch, A. Physicians as the First Analytical Chemists: Gall Nut Extract Determination of Iron Ion (Fe2+) Concentration. J. Chem. Educ. 2018, 95 (3), 456−462. (16) Harris, D. C. Quantitative Chemical Analysis, 6th ed.; WH Freeman and Company: New York, 2003.

Student handouts and instructor materials (PDF, DOCX)

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. ORCID

Parawee Rattanakit: 0000-0003-1065-3403 Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS The authors would like to thank Natta Jaikrajang for summarizing the data collected from the student reports and Adcharawadee Chooyimpanit for calculating and comparing the running costs of the laboratory experiment. Special thanks are expressed to Chitnarong Sirisathitkul for his suggestions on the manuscript and his support and encouragement. The authors would also like to thank David J. Harding for his assistance in the correction of grammar and writing and his additional advice concerning the manuscript. Lastly, we would like to gratefully D




(17) Jaikrajang, N.; Kruanetr, S.; Harding, D. J.; Rattanakit, P. A Simple Flow Injection Spectrophotometric Procedure for Iron (III) Determination Using Phyllanthus Emblica Linn. as a Natural Reagent. Spectrochim. Acta, Part A 2018, 204 (5), 726−734. (18) Settheeworrarit, T.; Hartwell, S. K.; Lapanatnoppakhun, S.; Jakmunee, J.; Christian, G. D.; Grudpan, K. Exploiting Guava Leaf Extract as an Alternative Natural Reagent for Flow Injection Determination of Iron. Talanta 2005, 68 (2), 262−267.

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Determining Iron(III) Concentration in a Green Chemistry Experiment

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