Disposable Glassy Carbon Stencil Printed Electrodes for Trace Detection of Cadmium and Lead

Abstract

Cadmium (Cd) and lead (Pb) pollution is a significant environmental and human health concern, and methods to detect Cd and Pb on site are valuable. Stencil-printed carbon electrodes (SPCEs) are an attractive electrode material for point-of-care (POC) applications due to their low cost, ease of fabrication, disposability and portability. At present, SPCEs are exclusively formulated from graphitic carbon powder and conductive carbon ink. However, graphitic carbon SPCEs are not ideal for heavy metal sensing due to the heterogeneity of graphitic SPCE surfaces. Moreover, SPCEs typically require extensive modification to provide desirable detection limits and sensitivity at the POC, significantly increasing cost and complexity of analysis. While there are many examples of chemically modified SPCEs, the bulk SPCE composition has not been studied for heavy metal detection. Here, a glassy carbon microparticle stencil printed electrode (GC-SPE) was developed. The GC-SPEs were first characterized with SEM and cyclic voltammetry and then optimized for Cd and Pb detection with an in situ Bi-film plated. The GC-SPEs require no chemical modification or pretreatment significantly decreasing the cost and complexity of fabrication. The detection limits for Cd and Pb were estimated to be 0.46 μg L⁻¹ and 0.55 μg L⁻¹, respectively, which are below EPA limits for drinking water (5 μg L⁻¹ Cd and 10 μg L⁻¹ Pb).[1] The reported GC-SPEs are advantageous with their low cost, ease of fabrication and use, and attractive performance. The GC-SPEs can be used for low-level metal detection at the POC as shown in the report herein.

Graphical Abstract

1. Introduction

Heavy metal pollution, originating from both natural and industrial sources, is a significant human and environmental health concern. Heavy metals are persistent in the environment, non-biodegradable, and bioaccumulate.[2] While several heavy metals are essential for biochemical and physiological pathways, other heavy metals such as cadmium (Cd) and lead (Pb) are highly toxic and serve no biochemical function.[3] Cd and Pb accumulate in human tissues, adversely affecting numerous organs and organ systems causing respiratory, cardiovascular, renal, neurological, developmental, reproductive, and hematological symptoms.[2, 4] As a result, federal agencies have set strict limits and regulations on the acceptable levels of Cd and Pb in media including water, soil, and air in both environmental and occupational settings.[5, 6] Since distribution of Cd and Pb in the environment is widespread, these metals should be continuously monitored.[2]

Atomic absorption spectroscopy (AAS), inductively coupled plasma mass spectrometry (ICP-MS) and variations thereof are the certified, standard methods for highly sensitive Cd and Pb determination in a variety of complex sample matrices.[7–9] However, these techniques are not suitable for large scale environmental studies or on-site monitoring. Samples must be distributed to a centralized laboratory and analyzed by trained personnel. Therefore, the time to answer is slow and costs are high using conventional methods.[10] Electrochemical techniques including potentiometry and anodic/cathodic stripping voltammetry are beneficial when rapid, on-site determination of Cd and Pb is necessary. With portable, miniaturized instrumentation becoming more accessible, electrochemical methods are well suited for POC monitoring of Cd and Pb, providing fast analysis times (e.g. several minutes) with minimal training.[11–13] Commercially available ion selective electrodes (ISEs) provide detection of limits of 10 μg L⁻¹ and 210 μg L⁻¹ for Cd and Pb, respectively.[14, 15] However, these detection limits are not relevant in many applications. For example, 5 μg L⁻¹ Cd and 10 μg L⁻¹ Pb are the maximum permissible levels in drinking water.[16] In addition, multiplexing is difficult because different ISEs are required for the detection of various ions.

Anodic stripping voltammetry (ASV) is widely used in trace and ultra-trace metal determination.[17, 18] Historically, mercury (Hg) (mercury film or hanging mercury drop electrode) was the standard electrode material for stripping voltammetry.[19, 20] Hg provides a wide negative potential window, which is useful for the large negative potentials required to deposit metals. Hg forms a stable amalgam with other metals and multiplexed detection of up to six metals is possible. In optimized systems, ASV can provide as low as 10⁻¹¹ M detection limits.[10, 19] However, Hg is a toxic metal, and its use has been banned and/or restricted in certain countries.[21] Significant efforts have been made to construct alternative, non-toxic, electrode materials. In 2000, Joseph Wang and coworkers reported the first bismuth-film electrode. In this work, bismuth (Bi) was deposited in situ onto a glassy carbon electrode (GCE), providing similar stripping performance to Hg-film electrodes.[19] Bi-film GCEs are attractive for trace metal analysis since Bi forms stable alloys with other metals, is non-toxic, and Bi-film GCEs are not sensitive to dissolved oxygen, eliminating the need to purge solutions prior to analysis.[19, 22] However, macro GCEs are not suitable for POC detection because they are bulky, non-disposable, expensive, and require large sample volume.

Stencil printed carbon electrodes (SPCEs), a common composite carbon electrode material, have been widely adopted for POC testing of metals. SPCEs are inexpensive, simple to fabricate, mass producible, require small sample volumes (e.g. < 100 μL), and disposable.[12, 23] Moreover, carbon composite electrodes are frequently integrated with both traditional and paper-based microfluidics, with the goal of increasing portability and field use.[24, 25] Several reports performed ASV carried with Bi-film SPCEs. However, Bi-film SPCEs have displayed a number of undesirable characteristics including double stripping peaks for a single metal, poor resolution of neighboring metal peaks, high background currents and lower sensitivities than macro GCEs.[26–29] Graphite powders are exclusively employed in SPCE fabrication, yet graphite is inherently heterogeneous, with both edge and basal planes exposed, resulting in heterogeneous electrochemical activity at a single electrode surface.[30, 31] The heterogeneity is suggested to contribute to the aforementioned problems with stripping voltammetry.[20] While glassy carbon is also a form of graphitic carbon, the microstructure differs from graphite and Bi-film GCEs provide single, well resolved stripping peaks for Cd and Pb detection.[19] Significant work has been done to overcome the problems with Bi-film SPCEs by chemically modifying the electrode surface with nanomaterials and/or polymers such as silver nanoparticle/Bi/Nafion,[26] Nafion/ionic liquid/Graphene,[32] multiwalled carbon nanotubes,[33] and many others.[34] Cd and Pb quantification has been successful with these methods, nevertheless, chemical modification is complex, time consuming, expensive, and often reduces electrode stability.

To date, there are no published works studying the use of other carbon materials and/or graphite types for the SPCE bulk electrode composition for Cd and Pb detection. There are a large number of reports on modifying the surface of SPCEs with alternative forms of carbon.[35–38] However, modifying the bulk material is of interest since there are many forms of carbon available for fabricating carbon composite electrodes, including SPCEs. Here, we developed a glassy carbon microparticle stencil printed electrode (GC-SPE). The proposed GC-SPE is similar to macro GCEs for the detection of Cd and Pb in terms of stripping peak characteristics, allowing an inexpensive, simple, disposable and portable detection platform. The GC-SPEs were fabricated in the same fashion as traditional SPCEs; glassy carbon microparticles were mixed with a commercial ink, then printed through a stencil onto a polyethylene terephthalate (PET) substrate. Electrochemical characteristics including solvent window, double layer capacitance, and cyclic voltammetry of several redox mediators, both surface sensitive and insensitive, are discussed. Next, the optimal detection conditions for square wave anodic stripping voltammetry (SWASV) for trace detection of Cd and Pb with the GC-SPEs are presented. Here, our GC-SPE is capable of rapid, sensitive, and simultaneous detection of Cd and Pb. Detection limits were determined based on 3S/N and found as 0.46 μg L⁻¹ (Cd) and 0.54 μg L⁻¹ (Pb) after 6 min deposition.[39] The system was then used to determine Cd and Pb in soil extractants. This work provides steps toward simplified fabrication of stencil printed electrodes for ASV of heavy metals, whereby the carbon type can be tailored to improve the performance of this class of electrodes for POC applications.

2. Experimental

2.1. Reagents and Materials

Potassium chloride (KCl) and glacial acetic acid (TraceMetal™ Grade) were purchased from Fisher Scientific (New Jersey, USA). Potassium hexacyanoferrate(III) (K₃Fe(CN)₆), potassium hexacyanoferrate(II) trihydrate (K₄Fe(CN)₆), sodium hydroxide (NaOH, 99.99% trace metal basis), atomic absorption spectroscopy standards of Bi, Cd, and Pb (1000 mg L⁻¹ in 5 wt% nitric acid), and certified reference standard soil (Trace metals sand 1) were purchased from Sigma-Aldrich. P-aminophenol (pAP), glassy carbon microparticles (spherical, 0.4 – 12 μm) and glassy carbon microparticles (spherical, 10 – 20 μm) were purchased from Alfa-Aesar (Massachusetts, USA). Commercial carbon ink (E3178) was purchased from Ercon Incorporated (Massachusetts, USA). Silver (Ag) paint was purchased from SPI supplies (Pennsylvania, USA). Silver/Silver Chloride ink was purchased from Gwent Group (Torfean, UK). Ferrocenylmethyl trimethylammonium hexafluorophosphate (FcTMA⁺) was synthesized in house using a previously reported method.[40] All solutions were prepared fresh daily using 18.2 MΩ·cm water purified using a Milli-Q system. Transparency film (polyethylene terephthalate/PET) and double-sided adhesive (467 MP) were purchased from 3M (Minnesota, USA). The electrode stencil and solution reservoir were designed using CorelDRAW (Corel; Ontario, Canada) and cut using a 30 W Epilog Engraver Zing CO₂ laser cutter and engraver (Colorado, USA). Carbon electrodes (ItalSens IS-C) were purchased from Basi Inc. (Indiana, USA). The stir plate (Equatherm™, 267-914) used in stirred ASV experiments was purchased from Curtin Matheson Scientific, Inc (Texas, USA). Graphite powder (MG-1599) was purchased from Great Lakes Graphite (Ontario, Canada). Soil samples (#14, #21, and Gold King Mind (GKM) were obtained from commercial and private sources.

2.2. GC-SPE and SPCE Fabrication

The GC-SPEs were fabricated according to a previously reported three electrode cell design which was laser cut into a PET sheet to form the stencil.[26, 35] The fabrication scheme is shown in Fig. S1. To investigate the effect of particle size on electrochemical performance of GC-SPEs, two different GC microparticle sizes, 0.4 – 12 μm and 10 – 20 μm, were used for fabrication of GC-SPEs. A hand-mixed composite of GC microparticles and commercial carbon ink (Ercon) (0.8 g ink : 1.0 g GC) was then stencil printed onto a second PET sheet. This ratio was chosen due to the best consistency of the ink for stencil printing. The electrodes were then cured in a 65° C oven for 30 minutes. The working electrode (WE) was 3 mm in diameter. Both WE and the counter electrode (CE) were GC-SPEs. Ag/AgCl ink (Gwent) was painted onto the exposed GC-SPE surface to serve as the pseudo-reference electrode (RE). Ag/AgCl ink produced the most stable potential over time compared to a C RE and Ag paint RE (SPI) as discussed in the supporting information (Fig. S2). The same procedure was carried out to fabricate Ercon ink only SPCEs and graphite SPCEs (3.5 g ink : 2.5 g graphite). To fabricate the solution reservoir, a PET sheet was sandwiched between two layers of double-sided adhesive (3M) and laser cut. The solution reservoir was attached to the SPEs via the double-sided adhesive and aligned with the electrode connection pads to define a reproducible interelectrode surface area of 0.112 cm². The solution volume required to fill the solution reservoir was 50 μL.

2.3. Electrode Characterization

All electrochemical experiments were carried out using a CH Instruments 660B potentiostat (Texas, USA) at room temperature (22 ± 1 °C). For electrode characterization, Ag/AgCl ink served as the RE, a GC-SPE served as the CE, and all solutions contained 0.1 M KCl as the supporting electrolyte. A solution volume of 50 μL was used to fill the solution reservoir. Cyclic voltammetry experiments were carried out with 1.0 mM FcTMA⁺, 1.0 mM Fe(CN)₆³⁻/Fe(CN)₆⁴⁻ and 0.5 mM pAP at a scan rate of 100 mV s⁻¹. Solvent windows were recorded in 0.1 M KCl (pH 6.5) at a scan rate of 200 mV s⁻¹ between +2.0 and −2.0 V. Capacitance measurements were carried out in 0.1 M KCl at scan rate of 100 mV s⁻¹ between +0.1 and −0.1 V for a total of 10 cycles. The electrodes were imaged using a JSM-6500F field emission scanning electron microscope (JEOL, Tokyo, Japan) with a 2 kV acceleration voltage. Cd and Pb Detection

The RE used was either a saturated calomel electrode (SCE) or Ag/AgCl ink. The Ag/AgCl was compared to the SCE to demonstrate the feasibility of these sensors for in field use. pH optimization studies for ASV were carried out in 0.1 M acetate buffers with pH values varying from pH 3.6 to 5.5. All metal (Cd, Pb, and Bi) solutions were prepared by diluting the AAS standards in acetate buffer. A deposition potential of −1.2 V and −1.4 V vs. SCE and Ag/AgCl were used respectively for ASV. In SWV, the frequency was 14 Hz, the amplitude was 80 mV, and the increment was 20 mV (Fig. S4). The Bi-film was plated in situ by spiking the Cd and Pb solutions with the appropriate concentrations of Bi. Cd and Pb were extracted from the real soil samples using a previously described extraction method.[41] Standard addition was used to quantify Cd and Pb in the extraction solutions. 3 μM Fe(CN)₆³⁻ was added to the standard addition solutions to remove copper (Cu) interference as previously reported.[26] To generate a calibration curve in stirred solutions, the stir plate was set to 5.

3. Results and Discussion

3.1. Electrochemical Characterization and Imaging of GC-SPEs

To investigate which GC microparticle size provides better electrochemical performance, cyclic voltammetry (CV) was used to determine peak-to-peak separation (ΔE_p) and peak currents of several well understood redox mediators. First, CVs of Fe(CN)₆³⁻/Fe(CN)₆⁴⁻ were obtained (Fig. 1A). Fe(CN)₆³⁻/Fe(CN)₆⁴⁻ is an electrochemically reversible redox species, however, it has been shown to undergo a surface sensitive electrochemical reaction and irreversible voltammetry is often observed at carbon electrode materials.[24, 30, 31] Here, the measured ΔE_p values were 441 ± 12 mV and 512 ± 7 mV for the 0.4 – 12 μm and 10 – 20 μm GC-SPEs respectively. The large peak separations for both are likely a result of deactivation of the electrode surface due to the ink coating as well as ohmic resistance effects.[12, 42, 43] A scan rate study was carried out with each electrode type. The peak separation of Fe(CN)₆⁴⁻/Fe(CN)₆³⁻ increases with increasing scan rate at both electrode types, however, peak separations at the 10 – 20 μm GC-SPE exhibited greater dependence on the scan rate than the smaller particle size (Fig. S5). CV of pAP, an electrochemically reversible but surface sensitive redox species is shown in Fig. 1B. The measured ΔE_p values were 246 ± 14 and 421 ± 18 mV for the 0.4 – 12 μm and 10 – 20 μm GC-SPEs respectively, again indicating lower electrochemical activity of the 10 – 20 μm GC-SPE. Next, CV of FcTMA⁺ was investigated as shown in Fig. 1C. FcTMA⁺ is surface insensitive and undergoes an “outer-sphere” electron transfer reaction.[44] Therefore, the voltammetry is insensitive to the electronic and chemical structure of the electrode surface and Nernstian behavior is observed at sufficiently conductive electrode materials.[44, 45] Here, ΔE_p values were 107 ± 8 mV and 105 ± 15 mV for the 0.4 – 12 μm and 10 – 20 μm GC-SPEs respectively. There is no difference in FcTMA⁺ voltammetry between the two electrodes, indicative of the surface insensitive electron transfer reaction of FcTMA⁺.[46] The peak separations indicate ohmic resistance effects due to the lower conductivity associated with composite electrode materials.[47] Since FcTMA⁺ is an electrochemically reversible redox probe, an ideal peak-to-peak separation of 59 (±10) mV is expected under diffusion limited conditions.[44, 48] The 0.4 – 12 μm GC-SPE outperformed the 10 – 20 μm GC-SPE indicated by the smaller ΔE_p values and higher peak currents obtained for both surface sensitive species. Therefore, the 0.4 – 12 μm GC-SPE was selected for further studies. We hypothesize that the smaller particle size outperforms the larger particle size due to easier mixing of the particles and the ink, producing better particle to particle contact and reduced ohmic resistance effects.[49]

Figure 1.

Cyclic voltammograms recorded at a 0.4 – 12 μm (gold traces) and 10 – 20 μm (blue traces) GC-SPE in (a) 1 mM Fe(CN)₆³⁻/Fe(CN)₆⁴⁻ (b) 0.5 mM pAP and (c) 1 mM FcTMA⁺ in 0.1 M KCl. (d) Solvent windows recorded with the two GC-SPE types and a homemade SPCE (MG1599) in 0.1 M KCl, pH 6.5.

The HER and oxygen reduction reaction (ORR) are typical reactions occurring in the cathodic potential region employed in ASV, and can have deleterious effects on both the deposition and stripping of Cd and Pb.[8, 19] Therefore, both electrochemical reactions must be considered when selecting an electrode material for ASV. The solvent window for each type of GC-SPE was measured and compared to a homemade SPCE (MG1599) (Fig. 1D). Here, both GC-SPE solvent windows are almost featureless with only a small background peak at ~1 V, which is attributed to sp² carbon oxidation.[44] The SPCE current density for hydrogen evolution (HER) at potentials negative of −1.5 V is significantly greater than both GC-SPEs. HER is a catalytic reaction and requires surface binding sites to occur to an appreciable extent. Greater HER at the graphite SPCE indicates the presence of a higher density of catalytic sites, likely on graphite edges.[30, 44] Additionally, a reduction wave for ORR is not observed at the GC-SPE. Fig. S8b shows the potential windows of a macro GCE, a GC-SPE, and an SPCE. At the macro-GCE, ORR was observed at an onset potential of ~ −1.5 V. These results indicate the GC-SPE has a wider operational cathodic potential window than graphite SPCEs and macro GCEs, rendering the GC-SPE suitable for ASV.

The background current is an important parameter to consider in electrochemical sensing applications as it limits the achievable detection limits.[44] The capacitance of an electrode contributes significantly to the background current; therefore, the capacitance of the GC-SPEs was measured experimentally (Fig. S6). Equation 1 was used to calculate the apparent capacitance of the GC-SPEs (0.4 – 12 μm and 10 – 20 μm), traditional SPCEs, and a macro GCE.

$${\text{C}}_{\text{dl}}\text{=}\frac{{\text{i}}_{\text{average}}}{\text{vA}}$$ (1)

i_average is the average absolute value of the measured anodic and cathodic currents in A, v is the scan rate in V/s and A_geometric is the geometric surface area in cm².[45] C_dl values for the GC-SPEs were 6.05 ± 0.59 μF cm⁻² and 6.60 ± 1.21 μF cm⁻² for the 0.4 – 12 μm and 10 – 20 μm GC-SPEs respectively. The low capacitance obtained is due to the low fraction of electrochemically reactive (GC) surface area in contact with solution, while the remainder of the surface is occupied by the ink.[30] This low capacitance is a highly desirable property for sensing applications as it directly effects the signal to noise characteristics. The experimentally measured capacitance of a macro GCE was 66.1 μF cm⁻² (Fig. S6b). This large capacitance is likely a result of polishing the electrode, which can increase the number of redox active moieties on the electrode surface. Commercially available SPCEs have capacitance around 37 μF cm⁻² which is consistent with the data presented in Fig. 3.[42] The MG1599 had a measured capacitance of 7.95 ± 0.59 μF cm⁻².

Fieure 3.

SWASV for a 100 μg L⁻¹ solution of Cd and Pb in 0.1 M acetate buffer (pH 3.6) with an in situ plated Bi-film GCSPE (blue line), SPCE (green line), Ercon ink only SPCE (gold line) and PalmSens SPCE (black line). Deposition time for all ASVs was 3 min.

Scanning electron microscopy (SEM) was used to image the surface morphology of the 0.4 – 12 μm and the SPCE as shown in Fig. 2. The GC particles appear as smooth spheres (Fig 2A), while the graphitic flakes appear rougher and both basal and edge planes are visible. The surface of the outermost GC particles do not all appear to be uniformly coated in the ink. However, the coating on the graphite particles on the SPCE surface appear to be thinner and more homogeneous, and a much higher magnification is required to image uncoated particles (Fig. 2B inset). We hypothesize that the difference in particle morphology clearly evident in the SEM images contributes to the stripping behavior shown in Fig. 3.

Figure 2.

SEM images of (a) 0.4 – 12 μm GC-SPE and (b) the MG1599 SPCE.

In order to compare the behavior of the GC-SPEs for ASV to conventional stencil printed electrode materials, SWASV was carried out with four different electrode types: 1) GC-SPE, 2) SPCE (MG-1599), 3) Ercon ink only SPCE, and 4) PalmSens SPCE as shown in Fig. 3. Here, the GC-SPE produces symmetric and well-resolved peaks for stripping voltammetry of Cd and Pb at −0.87 V and −0.65 V (vs Ag/AgCl) respectively. In contrast, the stripping peaks for Cd and Pb obtained using the homemade graphite SPCE are split into unresolved double peaks indicated by the small shoulder on the Cd peak at about −0.88 V and the broad double peaks obtained for Pb at about −0.65 V and −0.59 V. These results are indicative of Cd and Pb being deposited and subsequently stripped from different chemical environments, possibly the basal and edge planes of the graphite.[26, 50, 51] These double peaks would likely become more evident at lower concentrations, as larger peaks obscure this effect.[51] Stripping peaks are observed for the Ercon ink SPCE, however, the peaks are not well resolved and quantification via SWASV of Cd and Pb at this SPCE material without further modification would be difficult. Cd and Pb stripping peaks are also evident with the PalmSens electrode, however, in contrast to all other SPCE types, the background current is an order of magnitude higher, significantly affecting the achievable signal-to-noise ratio. The increase is likely a result of the higher capacitance of commercial SPCEs compared to the GC-SPEs as discussed previously.[42] A high background and low signal to noise ratio is undesirable in electrochemical sensing as the detection limit is calculated using this parameter.[39] Here, the GC-SPE provides the most sensitive SWASV characteristics for the simultaneous detection of Cd and Pb compared to the other SPCE types as evident in the voltammograms shown in Fig. 3.

Prior to generating calibration curves for Cd and Pb with the GC-SPE, we first optimized the SWV parameters and solution conditions. The square wave frequency, amplitude, increment and deposition time were selected for optimal reproducibility and maximum peak currents while still retaining well-resolved peaks. The optimal square wave parameters of 80 mV for the amplitude, 20 mV for the increment, and 14 mV for the frequency were selected. While the peak current increased at higher frequencies (i_p ∝ 1/f^1/2), higher frequencies also increased the peak width, decreasing Cd and Pb peak resolution. We selected a deposition time of 6 min. While sensitivity increases at increasing deposition times and we did not observe saturation up to 15 min deposition times, we chose 6 min to minimize analysis time and maintain sensitivity. Deposition time optimization data is shown in the SI (Fig. S7).

The concentration of Bi precursor present in the metal plating solution has been shown to be a critical parameter for optimal SWASV sensitivity. We hypothesize this to be a result of the thickness of the Bi film formed during the deposition step.[52] As demonstrated previously, the Bi to Cd and Pb ratio has significant effects on the sensitivity of the stripping peak signals. For ASV with carbon paste electrodes (CPEs), the optimal Bi to Cd and Pb ratio has been shown to range between 1 and 10, with a decrease in sensitivity at ratios greater than 10.[52] To study these effects at GC-SPEs, the Bi ion concentration in the Cd and Pb plating solution was varied from 1 to 5 mg L⁻¹. The resulting peak currents for 100 μg L⁻¹ Cd and Pb are shown in Fig. 4A. Here, the peak current reaches a maximum at 2.5 mg L⁻¹ Bi, or a Bi to Cd and Pb ratio of 12.5. At higher Bi concentrations, both peak current and reproducibility of Cd and Pb decrease. This is likely a result of increasing Bi-film thickness at high concentrations, leading to Cd and Pb occlusion during co-deposition.[52] While the Bi to Cd and Pb ratio of 12.5 was optimal for 100 μg L⁻¹ Cd and Pb, this concentration is at the higher end of the concentrations we are interested in detecting, therefore, a ratio of 10 Bi to Cd and Pb (2 mg L⁻¹ Bi to 100 μg L⁻¹ Cd and Pb) was selected for calibration of Cd and Pb in the 7.5 to 200 μg L⁻¹ range. Figure 4B shows the peak currents for 10 μg L⁻¹ Cd and Pb co-deposited with varying concentrations of Bi. Again, the greatest peak currents were obtained at a Bi to metal ratio of 12.5 (250 μg L⁻¹ Bi). Therefore, we selected 250 μg L⁻¹ Bi for the calibration of 2.5 to 50 μg L⁻¹ Cd and Pb.

Figure 4.

SWASV peak currents obtained for (a) 100 μg L⁻¹ Cd and Pb and (b) 10 μg L⁻¹ Cd and Pb as a function of Bi precursor concentration carried out in 0.1 M acetate buffer, pH 3.6. Deposition times were 3 min and 12 min for 100 μg L⁻¹ and 10 μg L⁻¹ Cd and Pb respectively.

Solution chemistry, especially pH, is critical to the sensitive detection of Cd and Pb with Bi-film electrodes.[53–56] Therefore, SWASV was carried out in 0.1 M acetate buffers ranging in pH from 3.6 to 5.5. The peak currents for 100 μg L⁻¹ Cd and Pb as a function of pH are shown in Fig. 5A. Interestingly, the highest peak currents occurred at a pH of 3.6. This diverges from the literature where acetate buffer, pH 4.5, is widely accepted as the optimal supporting electrolyte for both in situ and ex situ Bi-film electrodes.[19] For further investigation, the Bi-film stripping characteristics were studied as a function of solution pH shown in Fig. 5B and 5C. Here, the peak current for Bi is significantly higher at a pH of 3.6 than all other pH conditions, decreasing exponentially as a function of increasing pH. The pH dependence of Cd and Pb ASV is likely controlled by the properties of the Bi-film formed on the GC-SPE surface. Based on the voltammetry shown in Fig. 5B, Bi-film deposition at a pH of 3.6 yields the thickest Bi-film, which is likely more uniform, and provides a reproducible surface for Cd and Pb co-deposition. Higher pH conditions have been reported in the literature for ASV of Cd and Pb but not when employing Bi-film electrodes. For example, a copper electrode provided the greatest sensitivity in pH 5.5 acetate buffer.[8] As mentioned previously, the HER limits the cathodic potential window for an electrode material. HER is an inner-sphere electron transfer reaction and sensitive to the electrocatalytic properties of the electrode surface, producing H₂ gas via the reduction of H⁺. At lower pH (increasing H⁺ concentration), HER is increasingly thermodynamically favorable and occurs at less negative over potentials. In ASV, significant HER is detrimental due to the formation of gas bubbles on the electrode surface which interferes with metal deposition. Moreover, Bi hydrolysis occurs in solution pHs as low as 1 to 2.[57] Metal speciation, including Bi speciation, controls lability and consequently, deposition efficiency.[34, 54, 56] To mitigate HER while maintaining the highest percentage of labile Bi in solution, pH 4.5 is typically employed in SWASV with in situ Bi-film carbon electrodes. However, HER does not occur to an appreciable extent at the GC-SPEs compared to graphite (Fig. 1D) and macro GC electrodes (Fig. S8a and b). Potential windows were recorded in acetate buffer pH 3.6 to 5.5 as shown in Fig S8b. Here, the HER did not increase with acidity at the GC-SPEs. The GC-SPEs are functional in more acidic solutions, and the deposition step can be carried out in increasingly acidic conditions, increasing the stability of Bi in the plating solutions.[8]

Figure 5.

SWASV peak currents obtained in 0.1 M acetate buffers ranging from pH 3.6 to 5.5 for (a) 100 μg L⁻¹ Cd and Pb. SWASV results obtained for 2 mg L⁻¹ Bi plated in 0.1 M acetate buffers ranging from pH 3.6 to 5.5, (b) voltammograms and (c) peak currents.

Next, we generated a calibration curve for Cd and Pb with the optimized SWASV parameters and solution conditions described above. While the GC-SPEs are disposable, it was also found that they are reusable up to at least 10 times when the electrode was cleaned between ASV experiments as shown in Fig. S9. Therefore, we generated two sets of calibration curves for 7.5 to 200 μg L⁻¹ Cd and Pb. The first calibration curve was generated with a brand new electrode for every run (n = 3 per calibration point). In these experiments, an SCE RE was employed and the calibration curve is shown in Fig. S3 a and c. Next, we generated a calibration curve with a total of 3 electrodes. The electrodes were reused after applying a cleaning step between each ASV experiment in acetate buffer, pH 3.6, by holding the potential at +0.4 V for 100 seconds. One electrode was employed to test each standard concentration once. The Ag/AgCl ink RE was employed to demonstrate the applicability and reusability of the sensor for use in the field. The sensitivities of the calibration curves generated with an SCE RE and an Ag/AgCl RE were similar but statistically different for both Cd and Pb. The observed difference in sensitivities is attributed to reusing the electrodes for the Ag/AgCl calibration curve. Over the course of many ASV experiments, the GC-SPE undergoes significant cathodic polarization to deposit the metals, then is anodically polarized at +0.4 V in acetate buffer to clean the surface for the next experiment. Electrochemical polarization at high anodic potentials generates reactive oxygen species such as O₂ and CO₂, that can further oxidize the carbon electrode surface. Also, oxidation of organic surface impurities (e.g. the ink) and their subsequent removal from the electrode surface can occur, resulting in further activation of the composite electrode surface.[58] While some change in the surface chemistry is evident, the correlation for all calibration curves is greater than 0.99, and both methods are suitable for Cd and Pb calibration as shown in Fig. S3. As the results in Fig. 6 show, the GC-SPEs provide sharp, well-resolved, and reproducible peak currents (coefficient of variation ranges from 2 to 12%) for the simultaneous detection Cd and Pb over a wide linear range of 7.5 to 200 μg L⁻¹ Cd and Pb. The sensitivities were found as 0.122 μA L μg⁻¹ (R² = 0.992) and 0.101 μA L μg⁻¹ (R² = 0.992) for Cd and Pb, respectively. The obtained sensitivities of the developed electrodes are higher than other SPCEs reported in the literature. For example, the sensitivities achieved for Cd and Pb with a graphitic Bi-film SPCE modified with multi-walled carbon nanotubes were 0.0066 μA L μg⁻¹ and 0.0068 μA L μg⁻¹, respectively.[35] The deposition time in this study was slightly shorter at 4 min, however, this small difference would not account for the almost 20 times increase in sensitivity at the GC-SPEs. Detection limits were calculated using 3σ/m, where σ is the standard deviation of the background current for the lowest measured concentration and m is the slope of the calibration curve. Detection limits were estimated as 0.46 μg L⁻¹ and 0.55 μg L⁻¹ for Cd and Pb respectively.

Figure 6.

Calibration curves of 7.5 to 200 μg L⁻¹ Cd and Pb using SWASV. (a) Shows the voltammograms and (b) shows the linear regressions. Analysis was performed in 0.1 M acetate buffer pH 3.6, 360 s deposition time, −1.4 V deposition potential.

To the best of our knowledge, these detection limits have only been reported for chemically modified Bi-film SPCEs and/or bare Bi-film SPCEs operated under hydrodynamic conditions. In hydrodynamic conditions, the deposition step is carried out under forced convection.[24, 25, 59] Therefore, mass transport is not limited by diffusion, deposition is more efficient, and higher sensitivities are achieved. To further increase sensitivity and lower detection limits, while also decreasing deposition time, the GC-SPEs could be adapted into portable systems that operate under flow such as traditional and paper-based microfluidics.[60–63] To illustrate the effect of forced convection on stripping analysis at the GC-SPEs, calibration curves were generated for Cd and Pb in stirred solution. The sensitivity of the GC-SPEs increased by a factor of ~ 6 for both Cd and Pb utilizing a 3 min deposition. The increase in sensitivity resulted in lower detection limits of 0.098 μg L⁻¹ Cd and 0.084 μg L⁻¹ Pb (Fig. S10). A third calibration curve (Fig. S11) for 2.5 to 50 μg L⁻¹ Cd and Pb employing 250 μg L⁻¹ Bi and a 20 min deposition was generated to demonstrate the flexibility of the sensor in terms of sensitivity and detection limits by tuning the deposition parameters in quiescent solution. Under these conditions, detection limits were lowered by a factor of two to 0.18 μg L⁻¹ and 0.29 μg L⁻¹ for Cd and Pb, respectively. Again, a high degree of reproducibility was achieved and the obtained coefficients of variation ranged from 1 to 10%.

Finally, the GC-SPEs were employed for quantitative determination of Cd and Pb in several soil samples via the standard addition method. The amounts of Cd and Pb were validated using ICP-MS and the results obtained using ASV and ICP-MS are shown in Table 1. Prior to ASV detection the samples were exposed to a mild extraction solution that is designed for POC or in-field use with quick extraction times and low hazard. Previous work in our group determined the interference other metal ions commonly present in environmental samples have on the ASV determination of Cd and Pb at in situ Bi-film electrodes. Copper ions (Cu) were found to interfere at relatively low Cd/Pb to Cu ratios.[23] In the previous work, adding 3 μM Fe(CN)₆³⁻ to the sample was shown to eliminate the interference effect of Cu on the stripping voltammetry of Cd and Pb.[23] Therefore, all standard addition measurements were carried out in the presence of 3 μM Fe(CN)₆³⁻. The amounts found using the mild extraction solution with ASV detection were lower than those found using ICP-MS, as expected. Traditionally, samples are fully digested using strong acidic conditions at high temperatures in order to free all Cd and Pb and measure total amounts. The extraction method used here likely extracts other organic matter that in turn complex Cd and Pb in the sample.[56] However, ASV only measures labile Cd and Pb.[56] Despite the differences observed, the concentrations of Cd and Pb determined by ASV are comparable to those obtained with ICP and are appropriately sensitive for POC or in-field detection.

Table 1.

Concentrations of Cd and Pb determined via ASV and ICP-MS sample extractants.

Sample	Cd (μg L−1) ASV	Cd (μg L−1) ICP-MS	Pb (μg L−1) ASV	Pb (μg L−1) ICP-MS
Standard Soil	2630 ± 110	4080	1410 ± 180	2310
Soil Sample #14	46.3 ± 9.3	63.2	2440 ± 200	7150
Soil Sample #21	Below LOD	0	213 ± 58	450
Gold King Mine	Below LOD	0	79.7 ± 31.3	121

4. Conclusions

In this work, we report the fabrication and characterization of a glassy carbon stencil printed electrode (GC-SPE) for the first time. The GC-SPEs exhibit highly desirable properties for sensing the heavy metals Cd and Pb. These properties include low background and capacitive currents, inhibited hydrogen evolution reaction (HER) in acidic pH conditions, and a more homogeneous electrochemically active surface for metal deposition leading to single, well resolved peaks for the anodic stripping voltammetry of Cd and Pb. Fabrication of the GC-SPEs is field detection. simple and no pre-treatment or chemical modification is required to detect trace levels of Cd and Pb which bodes well for a point-of-care sensor as it significantly reduces the cost of the sensor. The use of inexpensive glassy carbon microparticles, commercial carbon ink and transparency film to fabricate the GC-SPE lead to a cost of about $0.25 per sensor. Further work can include the integration of the GC-SPEs into both traditional and paper-based microfluidics, permitting the deposition step to be carried out under hydrodynamic conditions, further increasing the sensitivity of Cd and Pb detection at the POC.

Highlights.

• A low-cost and disposable glassy carbon stencil printed electrode (GC-SPE) was developed and characterized.

• GC-SPEs exhibit desirable characteristics for anodic stripping voltammetry compared to conventional graphite stencil-printed carbon electrodes (SPCEs).

• The GC-SPEs were applied to the sensitive detection of Cadmium (Cd) and Lead (Pb).