Simple and Inexpensive Microfluidic Thread Based Device for

The miniaturization of flow injection analysis devices presents several improvements related to analytical procedures. However, a cost reduction and ...
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Simple and Inexpensive Microfluidic Thread Based Device for Teaching Microflow Injection Analysis and Electrochemistry Deonir Agustini, Maŕ cio F. Bergamini, and Luiz Humberto Marcolino-Junior* Laboratório de Sensores Eletroquímicos (LabSensE), Departamento de Química, Universidade Federal do Paraná (UFPR), CEP 81.531-980 Curitiba, PR, Brazil

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S Supporting Information *

ABSTRACT: The miniaturization of flow injection analysis devices presents several improvements related to analytical procedures. However, a cost reduction and simplification are required to allow extensive use in undergraduate teaching laboratories. Here we present a low cost microfluidic electroanalytical device composed only of interlocking toy blocks, double sided tape, pencil leads, and hydrophilic cotton threads for performing microflow injection analysis (μFIA). With adequate analytical performance and a manufacturing cost of only $1.0/unit, the proposed device can be easily employed in undergraduate laboratories for teaching content related to μFIA and electrochemistry in a simple and inexpensive way.

KEYWORDS: Second-Year Undergraduate, Analytical Chemistry, Hands-On Learning/Manipulatives, Calibration, Electrochemistry, Microscale Lab, Quantitative Analysis

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out the experiments and discuss the results. The group activities were completed within a period of 90 min. In order to connect the fundamental information received by the students throughout the course with the laboratory techniques, the experiments carried out with the proposed device allowed exploration of the phenomena and concepts related to μFIA (such as transport of small volumes of solutions and samples in micrometric channels, low waste generation, detection in the form of transient signals, repeatability in sample injections) and electrochemical techniques of cyclic voltammetry and amperometry (system of measurements based on three electrodes, function of each electrode and supporting electrolyte, principle of operation of the electrochemical techniques, and interpretation of the obtained signals). In addition, concepts and routines related to analytical chemistry were taught, which include the use of software to organize the data in the form of graphs, construction of a relation between analytical signal and analyte concentration, and figures of merit (such as limit of detection, LOD; limit of quantification, LOQ; linear dynamic range, LDR; among others).

icroflow injection analysis (μFIA) with miniaturized devices is an important strategy to facilitate the use of this technique in undergraduate laboratories, due to low reagent and sample consumption, small waste generation, rapid analysis, and high portability.1−3 However, the use of μFIA is very limited due the high cost and complex manufacturing process of devices traditionally based on inorganic or polymeric materials,4−6 as well as the low mechanical strength and the inconvenient step of constructing hydrophobic barriers on low cost devices made with paper.7−9 Recently, microfluidic devices based on cotton threads have emerged as an interesting alternative for μFIA in a simple and accessible way.10−12 This material presents low cost, high mechanical strength, direct construction of the microchannels without the use of hydrophobic barriers, and transport of solutions without the need of mechanical pumping.13−15 In this way, we describe here a low cost electroanalytical microfluidic device based on cotton threads and composed only of simple materials present in the daily routine of people to be employed in undergraduate teaching laboratories to carry out μFIA with amperometric detection.

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CONTEXTUALIZATION AND EDUCATIONAL RELEVANCE The proposed device was applied in an undergraduate chemistry course in the matter related to instrumental analytical techniques (including electrochemical techniques) taught after the students have completed both general chemistry and analytical chemistry as prerequisites. The students (15−18) were arranged in groups (3−4) to carry © XXXX American Chemical Society and Division of Chemical Education, Inc.

ASSEMBLY AND OPERATION OF THE DEVICE The device manufacture was done by manual assembly of the components. Interlocking toy blocks were used as base and as Received: March 23, 2018 Revised: June 20, 2018

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DOI: 10.1021/acs.jchemed.8b00211 J. Chem. Educ. XXXX, XXX, XXX−XXX

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reservoirs of the solutions, dispensing the acquisition and use of specific components for this purpose. In order to perform μFIA, initially, supporting electrolyte was added in the inlet reservoir (Figure 1B), which fills the entire microfluidic channel (including the electrochemical cell) by action of the capillary force generated in the cotton threads due to their high polarity and the interfiber and interthread gaps.16 In the vertical region of the microfluidic channel, a siphon effect was generated by the gravitational force due to the height difference between the inlet and outlet reservoirs, creating a continuous and stable movement of the solutions along the device.17,18 The injection of the sample, with a volume of 2 μL, was performed (with a micropipette) directly in the microfluidic channel at a distance of 10 mm from the electrochemical cell (more detailed in the Supporting Information, Figure S2 and video), and transported (due to the continuous flow of supporting electrolyte in the microchannel) to the region of the electrodes previously connected to a potentiostat through jumper wire ribbon cables, with the generation of the electrochemical response of the analyte (through a redox reaction) in the form of a transient signal (example shown in the Supporting Information, Figure S3). Finally, the sample followed the flow direction of the supporting electrolyte to the outlet reservoir, allowing the injection of a new sample aliquot.

inlet and outlet reservoirs (both with a volume of 6 mL) (Figure 1A). The electrochemical cell was mounted on the

Figure 1. (A) Assembly process with indication of the materials that make up the proposed microfluidic device. (B) Main regions responsible for the microflow analyses with the device.

upper region of the base, with the addition of a piece of double sided tape (sticky acrylic adhesive foam tape), in which were fixed three pencil leads with diameters of 0.5 mm for the working and pseudoreference electrodes and 0.7 mm for the counter electrode, and two pieces of double sided tape (with a distance of 6 mm between them) over the central region of the pencil leads. The effective surface area of the electrodes (corresponding to the separation between the two pieces of double sided tape) was controlled with a ruler and can be optimized taking into account the obtained signal and the time required for complete cleaning of the electrochemical cell after detection of the analyte. For the completion of the assembly process, a piece of double sided tape was added next to the inlet reservoir to prevent possible leaks, followed by attaching the microfluidic channel (composed by 9 cotton threads removed from hydrophilic gauze) from the inlet reservoir to the outlet reservoir and placement of pieces of cotton threads over the electrode region. More detailed explanation about the device assembly are presented in the Supporting Information, Figure S1. Compared to the miniaturized system reported in a previous work,11 the proposed device presented a greater ease of construction and a smaller amount of components, since the base of the device was formed by the simple fitting of the interlocking toy blocks, which also function as inlet and outlet

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DEVICE APPLICATION Experiments were conducted with the proposed device to demonstrate its analytical performance and to evaluate some characteristics related to μFIA and electrochemical techniques that can be explored in practical classes with undergraduate students. The electrochemical analyses were performed with the device connected to a potentiostat μAutolab Type III; however, it is possible to replace the commercial equipment by low cost measurement platforms, such as the one presented by Meloni,19 which has a cost less than $30. In all analyses, 0.1 M phosphate-buffered saline (PBS) pH 7.0 was used as the supporting electrolyte. Initially, the students were trained in the software used in the potentiostat in relation to the choice of the electrochemical technique and the insertion of the measurement parameters. After that, the cyclic voltammetry technique, which is based on the application of a potential scan in one direction and the sequence in the reverse direction returning to the initial potential value, was used to obtain signals in the absence

Figure 2. (A) Typical chronoamperometric responses obtained with the proposed device for injections (n = 3) of standard AA with concentrations between 1.0 μM and 1.0 mM. The red transient signals (S) correspond to the injections of the orange juice sample (4-fold dilution in supporting electrolyte). In detail, the responses to the six lowest injected concentrations of standard AA (1.0−25.0 μM) in the device. (B) Linear relationship between the current values obtained in A and the AA concentrations. Interval time: 100 ms. Applied potential: 0.5 V. Injected volume: 2.0 μL. B

DOI: 10.1021/acs.jchemed.8b00211 J. Chem. Educ. XXXX, XXX, XXX−XXX

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Additionally, analytical curves for AA concentrations from 5.0 × 10−5 to 1.0 × 10−3 mol L−1 were constructed by students, followed by injection of a solution from a pharmaceutical sample containing AA. Typical results are presented in the Supporting Information, Figure S7A. Despite good results, a typical problem observed by instructors is related to the interval between injections as shown in the Supporting Information, Figure S7B. It is important to emphasize the return of the baseline to a low value before the next injection since it can provide difficulties in measuring the current value of the transient signal. With adoption of the approach, previously mentioned pharmaceutical samples were analyzed for ascorbic acid determination. Sample preparation was realized by direct dilution using PBS solution in order to provide a concentration value between 2.5 × 10−4 and 7.5 × 10−4 mol L−1 (C3−C5). The diluted sample was injected on the microchannel, and the determination of ascorbic acid content was performed using a calibration curve. Typical values found are present in the Supporting Information, Table S1.

(blank) and the presence of 1 mM ascorbic acid (AA) between −0.1 and 0.8 V, a potential range that was selected to visualize all redox processes that occurred on the electrodes of the device. The CV of the blank did not show any faradaic processes, indicating the efficiency in the assembly and operation of the device and the absence of contaminants. The CV of the AA presented an irreversible oxidation process in 0.34 V (Supporting Information, Figure S4), with the formation of dehydroascorbic acid by the loss of 2e− and 2H+ (Supporting Information, Figure S5). Subsequently, a μFIA assay was performed with amperometric detection, which is based on the application of a constant and sufficient potential for the generation of redox reactions of the analyte, with the monitoring of the current obtained from these reactions for the sample injections over the time of the analysis. An analytical curve employing a potential of 0.5 V (sufficient for complete oxidation of AA according to CV analysis) was constructed for the detection of 13 standard solutions of AA with concentrations between 1.0 μM and 1.0 mM, and obtained signals in the form of current transients corresponding to the oxidation of the analyte that occurred during the passage through the electrochemical cell (Figure 2A), with its signal intensities related to the injected AA concentrations. According to the current generated, a linear response was observed for the AA concentration range studied, with a linear regression equation of I (current, nA) = −6.67 (nA) + 3.24 × CAA (ascorbic acid concentration, μM), correlation coefficient (R2) of 0.9994, limit of detection (LOD) of 0.22 μM (three times the baseline standard deviation divided by the slope of the analytical curve), and a deviation of only 2.83% in the sensitivity values of the curve obtained for injections of increasing and decreasing concentrations of AA, indicating an excellent stability in the responses of the device and absence of electrode poisoning. The injections of a commercial orange juice sample (S in Figure 2) without any previous treatment were performed in the interval between injections of increasing and decreasing concentrations of standard AA, with calculation (through interpolation in the standard AA analytical curve) of an AA concentration of 0.35 mg mL−1 present in orange juice, similar to the value presented in the product package (0.36 mg mL−1). The transient signals obtained for the AA in Figure 2 showed similar widths (30 s), indicating a reproducible transport of the injected analyte, with absence of AA and juice sample retention in the microfluidic channel and in the electrochemical cell. In addition, an analytical frequency of 120 injections per hour was calculated, higher than reported by other studies with microfluidic devices,20−22 indicating the need for a short period of time to perform flow analyses (such as a complete analytical curve) with the proposed device, allowing its use in practical classes conducted in undergraduate laboratories. Finally, experiments using the proposed device were conducted (in three sections) with undergraduate students in chemistry divided into groups, who performed sequential injections of a solution containing 0.1 mM AA. Relative standard deviation (RSD) values of 1.43−6.48% were observed in the injections of each group (Supporting Information, Figure S6A), and an RSD of 10.23% was calculated for AA signals obtained between the groups (Supporting Information, Figure S6B). These results demonstrate the ease of construction and employment of the proposed microfluidic device and the potential for use in undergraduate laboratories.

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CONCLUSION We have reported the construction and use of a low cost electroanalytical microfluidic device based on cotton threads for μFIA, which presented a simple assembly and was composed entirely of easily obtainable materials, with a cost of only $1.0/device. The transport of solutions in the microchannel of the device removed the use of any pumping system; the transport was done only by the action of the capillary and gravitational forces. On the basis of its rapid response (30 s per injection), good sensitivity, robustness, and low deviation between the analyses, the proposed device presented the necessary requirements for use as a simple and accessible alternative for the exploration of concepts related to the μFIA and electrochemical techniques in undergraduate laboratories.

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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.8b00211.

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Response obtained in assays showing the formation of transient signal, cyclic voltammogram obtained for AA, reaction of electrochemical oxidation of ascorbic acid, transient signals obtained with the proposed device by 5 groups of undergraduate students, transient signals performed by two groups with different waiting time between each injection, and determination of ascorbic acid in pharmaceutical sample using the microfluidic device (PDF, DOCX) Video of sample injection (AVI)

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. ORCID

Luiz Humberto Marcolino-Junior: 0000-0002-6279-469X Notes

The authors declare no competing financial interest. C

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determination of copper and zinc. Anal. Methods 2015, 7 (20), 8757− 8762. (19) Meloni, G. N. Building a Microcontroller Based Potentiostat: A Inexpensive and Versatile Platform for Teaching Electrochemistry and Instrumentation. J. Chem. Educ. 2016, 93 (7), 1320−1322. (20) Wisitsoraat, A.; Sritongkham, P.; Karuwan, C.; Phokharatkul, D.; Maturos, T.; Tuantranont, A. Fast cholesterol detection using flow injection microfluidic device with functionalized carbon nanotubes based electrochemical sensor. Biosens. Bioelectron. 2010, 26 (4), 1514−20. (21) Fonseca, A.; Raimundo, I. M., Jr.; Rohwedder, J. J.; Lima, R. S.; Araujo, M. C. A microfluidic device with integrated fluorimetric detection for flow injection analysis. Anal. Bioanal. Chem. 2010, 396 (2), 715−23. (22) Xu, Z. R.; Zhong, C. H.; Guan, Y. X.; Chen, X. W.; Wang, J. H.; Fang, Z. L. A microfluidic flow injection system for DNA assay with fluids driven by an on-chip integrated pump based on capillary and evaporation effects. Lab Chip 2008, 8 (10), 1658−63.

ACKNOWLEDGMENTS We gratefully acknowledge the undergraduates enrolled in chemistry courses and financial support from Brazilian foundations: CAPES and CNPq (process number 402943/ 2016-3).

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DOI: 10.1021/acs.jchemed.8b00211 J. Chem. Educ. XXXX, XXX, XXX−XXX