Visible light-induced ion-selective optodes based on a metastable photoacid for cation detection

Abstract

A new platform of ion-selective optodes is presented here to detect cations under thermodynamic equilibrium via ratiometric analysis. This novel platform utilizes a ‘one of a kind’ visible light-induced metastable photoacid as a reference ion indicator to achieve activatable and controllable sensors. These ion-selective optodes were studied in terms of their stability, sensitivity, selectivity, and theoretical aspects.

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Ion selective optodes (ISOs) have been extensively studied to monitor different cations for biomedical and environmental applications.^1–11 In recent years, there has been considerable interest to study ISOs that are activatable, reversible and controllable for their use in ion sensing applications that require localized detection of free ions without perturbing their surroundings.^12–15 Attempts have been made to convert the traditional passive mode ISOs into active mode by the use of photoactive compounds such as spiropyrans and photoacid generators.^16–19 However, the use of spiropyrans in an ISO requires UV light to activate the sensor, which can result in cellular damage when applied in biomedical applications.²⁰ In addition, UV light photodegrades the active compound in the ISO, shortening the sensor's lifetime.^13–17 Moreover, photoacid generators in an ISO undergo photolysis, making the sensor irreversible.^18,19 To overcome photodegradation and irreversibility for such ISOs, we propose to use a ‘one of a kind’ visible light activatable and controllable ISO based on a metastable photoacid (mPAH) to detect cations under equilibrium conditions by ratiometric analysis.

A typical ISO consists of a neutral basic indicator (In) that is selective to a reference ion (H⁺), an ionophore (L) that is selective to the cation of interest, and anionic additives (R⁻) that maintain electroneutrality in the plasticized polymer matrix (Scheme 1A).^21–23 The response mechanism of these ISOs is dictated by mass transfer equilibria (cation-exchange process) between an organic (ISO) and an aqueous phase.^24,25 Consequently, the detection of the cation of interest, after achieving thermodynamic equilibrium, is made by measuring the changes in the indicators’ optical properties, either by absorption or fluorescence spectroscopy.²⁶ Nevertheless, prior to cation detection, these ISOs need an external source of protons for the cation-exchange process to proceed, resulting in a response mechanism with multiple steps. In contrast, the proposed tricyanofuran based mPAH (CF₃PhTCF-PAH) acts as a neutral acidic indicator (InH), providing protons for the cation-exchange process. Considering that mPAH is a photoactive compound, it photodissociates its proton upon irradiation and thermally undergoes proton reassociation with a dissociated state that is sufficiently long-lived,^27,28 thus eliminating the need for an external source of protons, while the dissociated state of the mPAH acts as an anionic additive to maintain electroneutrality within the ISO as shown in Scheme 1B.²² Hence, an ISO based on the proposed mPAH exhibits a one-step response mechanism.

Scheme 1 Cation-exchange process of ISOs that utilize a (A) neutral basic indicator, and a (B) neutral acidic indicator. (In: neutral basic indicator; L: cation selective ionophore; R⁻: anionic additive; InH: neutral acidic indicator).

We have recently shown that merocyanine based mPAH linked to an acrylate polymer backbone can be utilized in an ISO to detect calcium ions.²⁹ This merocyanine based mPAH is a neutrally charged acidic indicator which exhibits a longer equilibrium response time (in the order of hours). Consequently, the detection of calcium ions was performed under non-equilibrium conditions.²⁹ Moreover, we have shown that by modifying the merocyanine based mPAH with an appropriate functional group (electron donating group), the equilibrium response time can be shortened to the order of minutes.³⁰ However, the charged nature of the merocyanine based mPAH was optimal only with a polar plasticizer, such as 2-nitrophenyl octyl ether (o-NPOE), to reduce ion-pair formation that affects selectivity.²⁹o-NPOE absorbs under 400 nm, interfering with the absorbance peak of the deprotonated state merocyanine based mPAH, and inhibited ratiometric analysis. In contrast, the non-charged CF₃PhTCF-PAH has shown ideal compatibility with a non-polar plasticizer, such as bis(2-ethylhexyl)sebacate (DOS) which does not interfere optically, allowing ratiometric analysis to further increase the sensor sensitivity and signal reproducibility.³¹

As shown in Scheme 2, it is expected that CF₃PhTCF-PAH within the ISO would undergo oxidative photoreaction under visible light irradiation, resulting in a stable carbanion state (CF₃PhTCF-PA⁻) of the mPAH along with its photodissociated proton.³² Likewise, the photodissociated protons would be exchanged when exposed to cations of interest, as the CF₃PhTCF-PA⁻ state is sufficiently long-lived to allow for the diffusion mediated cation-exchange process.

Scheme 2 Photoresponsive behaviour of CF₃PhTCF-PAH when incorporated in an ISO.

CF₃PhTCF-PAH was synthesized according to a literature procedure.^32–34 The calcium and sodium ISOs proposed here contain CF₃PhTCF-PAH (7.5 mmol kg⁻¹), and calcium ionophore IV (22.5 mmol kg⁻¹) or sodium ionophore X (7.5 mmol kg⁻¹) within poly(vinyl chloride) (33 wt%) and DOS (66 wt%).

At first, these ISOs were exposed to their respective buffer solutions without any cation of interest. As shown in Fig. 1, the ISOs based on CF₃PhTCF-PAH were stable over repeated activation cycles between the ON (deprotonated form) and OFF states (protonated form) without any loss of the absorbance signal, indicating no observable photodegradation.

Fig. 1 Stability of CF₃PhTCF-PAH in ISOs for (A) calcium sensor in 0.5 M formate buffer at pH 4.5, and (B) sodium sensor in 0.3 M magnesium acetate buffer at pH 5.5. Absorbance was recorded at 470 nm. (ON state: after 1 minute irradiation with 470 nm; OFF state: after 25 minutes in the dark).

Fig. 2 shows the absorption spectra of the ISOs based on CF₃PhTCF-PAH towards different concentrations of calcium (Fig. 2A) and sodium (Fig. 2B) ions at thermodynamic equilibrium (25 minutes in the dark) once the ISOs were activated by visible light (470 nm) for 1 minute. From both absorption spectra, it is observed that as the concentration of the cation of interest increases, there is a gradual decrease in the CF₃PhTCF-PAH peak (470 nm) and a gradual increase in the CF₃PhTCF-PA⁻ peak (318 nm). These absorbance changes allow for ratiometric analysis (protonated and deprotonated absorbance peaks of the CF₃PhTCF-PAH) to indirectly correlate the activity of the cation of interest.

Fig. 2 Absorption spectra of ISOs for 25 minutes in the dark after activation for different concentrations of (A) calcium ions in 0.5 M formate buffer at pH 4.5; concentrations (a) 0, (b) 1.0 × 10⁻⁸, (c) 1.0 × 10⁻⁷, (d) 1.0 × 10⁻⁶, (e) 1.0 × 10⁻⁵, (f) 1.0 × 10⁻⁴, and (g) 1.0 × 10⁻³ M, and (B) sodium ions in 0.3 M magnesium acetate buffer at pH 5.5; concentrations (a) 0, (b) 1.0 × 10⁻⁶, (c) 1.0 × 10⁻⁵, (d) 1.0 × 10⁻⁴, (e) 1.0 × 10⁻³, (f) 1.0 × 10⁻², (g) 1.0 × 10⁻¹, and (h) 1.0 M.

The experimental data obtained were then compared to the theoretical response function. The theoretical response function (eqn (1)), in terms of the activity of protons and the cation of interest, was generated from the ion-exchange equilibria (Scheme 1B). This was derived by utilizing mass balance (eqn (2)), charge balance (eqn (3)), degree of deprotonation using the ratio of absorbance at 470 nm and 318 nm (eqn (4)), and the ion-exchange equilibrium constant (eqn (5)) equations. This is analogous to that of traditional ISO theory which was established in the early 1990s.^21–23

Theoretical response function for cation:

Mass balance equations:

Charge balance equation:

[In⁻] = z[L_xM^z+] (3)

Degree of deprotonation (α):

Cation-exchange equilibrium constant (K_exch) for the neutral acidic indicator:

The subscript “T” indicates the total concentration of CF₃PhTCF-PAH ([InH]_T) and the ionophore ([L]_T). The activity of the cation of interest and the proton is denoted as a_M^z+ and a_H⁺. Also, “x” and “z” denote the value of the ionophore chelating with the cation of interest and charge of the cation of interest, respectively. The ratio of absorbance for CF₃PhTCF-PAH is denoted as A, the protonated state of CF₃PhTCF-PAH in 1 M hydrochloric acid as A_P, and the deprotonated state of CF₃PhTCF-PAH in 1 M sodium hydroxide as A_D; for 25 minutes in the dark after activation.

As shown in Fig. 3, the experimental data present a strong correlation with the theoretical response curve. From the resulting cation response curves, the experimental limit of detection (LOD) for calcium and sodium ions was 2.6 × 10⁻⁶ M and 2.3 × 10⁻³ M, respectively. These values were obtained by intersecting two extrapolated segments of the response curve as indicated in the literature.³⁵ Furthermore, the cation-exchange constant (log K_exch) for calcium and sodium ISOs was −9.3 ± 0.3 and −5.3 ± 0.1, respectively. The ionophore–cation complex ratio for calcium ISO was 3 to 1 and for sodium ISOs was 1 to 1.^36,37

Fig. 3 Experimental ( and □) and theoretical (lines) responses of ISOs based on CF₃PhTCF-PAH for the detection of (A) calcium, and (B) sodium ions for 25 minutes in the dark after activation (n = 3).

It was hypothesized that ISOs which contain no anionic additive, cannot maintain constant ionic strength within the ISO.²³ As a result, the cation-exchange constant cannot be retained, due to each change in the activity of protons and the activity of the cation of interest.²³ However, the changes in the ionic strength within the ISO based on CF₃PhTCF-PAH, containing no additional anionic additive, were negligible as observed in the kinetic data for different concentrations of sodium ions (Fig. 4), where stable responses over time were obtained at thermodynamic equilibrium. It is noteworthy that the interactions between the negatively and positively charged components for such an ISO do occur, after a certain threshold (1 − α > 0.5), as the ionic strength within the ISO changes drastically. Accordingly, higher concentrations of cations are not shown in Fig. 3 because the thermodynamic equilibrium was not maintained, and thus the upper detection limit was 6.3 × 10⁻⁴ M and 0.6 M for calcium and sodium ions, respectively (at α = 0.5).

Fig. 4 Kinetic study for ISOs based on CF₃PhTCF-PAH at different concentrations of sodium ions in 0.3 M magnesium acetate buffer at pH 5.5. Scans every 30 second after activation; concentrations (a) 1.0 × 10⁻⁶, (b) 1.0 × 10⁻⁵, (c) 1.0 × 10⁻⁴, (d) 1.0 × 10⁻³, (e) 1.0 × 10⁻², (f) 1.0 × 10⁻¹, and (g) 1.0 M.

Furthermore, these ISOs based on CF₃PhTCF-PAH were studied in terms of selectivity towards interfering cations (J) by a separate solution method.^25,38 As shown in Fig. 5, all interfering ions were highly discriminated for both ISOs, calcium (Fig. 5A) and sodium (Fig. 5B). Thus, the selectivity coefficient values for the calcium ISO towards magnesium, sodium and potassium ions were −3.8, −3.1 and −3.3, respectively. When compared to other ISOs, containing the same ionophore and spiropyran as the reference indicators, the selectivity coefficients were −2.9 (magnesium ion), −6.6 (sodium ion) and −8.9 (potassium ion).¹⁶ The log K_M,J values for the sodium ISO towards potassium, calcium and magnesium ions were −4.5, −6.6 and −2.7, respectively. These values are comparable to the selectivity coefficient of sodium against the potassium ion (−2.4), calcium ion (−4.0) and magnesium ion (−4.1) containing spiropyran as a reference indicator.¹⁶ The selectivity coefficient (log K_M,J) values were calculated using eqn (6), and is analogous to that of ISO theory at α = 0.5.²³

Fig. 5 Selectivity response for (A) calcium (buffer: 0.5 M formate at pH 4.5), and (B) sodium (buffer: 0.3 M magnesium acetate at pH 5.5) ISOs towards interfering cations for 25 minutes in the dark after activation (n = 3).

Selectivity coefficient for interfering cations:

The high discrimination observed may be due to the proton affinity towards CF₃PhTCF-PA⁻ (deprotonated form) to be greater than the binding energy of interfering cations by the respective ionophores. Consequently, these ISOs inhibit the competitive behaviour between the interfering cations and protons during the cation-exchange process, even at high concentrations. As a result, an ISO based on CF₃PhTCF-PAH may also be utilized for practical purposes which demands negligible interactions towards interfering cations.

In conclusion, this novel visible light activatable and controllable ISO based on CF₃PhTCF-PAH that does not exhibit photodegradation, showed good stability, selectivity and reproducibility. Furthermore, the responses of these ISOs towards calcium and sodium ions were standardized following the cation-exchange equilibria. Similarly, other cations can be detected using this platform by interchanging the ionophore within the polymer matrix. Also, we aim to optimize this type of ISO in terms of size reduction and sensitivity. Likewise, we expect that this novel mPAH (CF₃PhTCF-PAH) may act as a substitute to neutral basic indicators for their use in cation sensing applications that provide control using visible light as needed.

The authors acknowledge the Colleges of Sciences and the Department of Chemistry at the University of Central Florida (USA) for financial support for this research. We also acknowledge Dr Yi Liao (Florida Institute of Technology, USA) to provide tricyanofuran based metastable photoacids for preliminary studies, Dr Eric Bakker (University of Geneva, Switzerland) and Dr Philippe Buhlmann (University of Minnesota, USA) for advice on ion-selective optode theory.

Notes and references

1. W. E. Morf, K. Seiler, B. Rusterholz and W. Simon, Anal. Chem., 1990, 62, 738.
2. K. Wang, K. Seiler, W. E. Morf, U. E. Spichiger, W. Simon, E. Lindner and E. Pungor, Anal. Sci., 1990, 6, 715.
3. K. Seiler, K. Wang, E. Bakker, W. E. Morf, B. Rusterholz, U. E. Spichiger and W. Simon, Clin. Chem., 1991, 37, 1350.
4. M. Lerchi, E. Bakker, B. Rusterholz and W. Simon, Anal. Chem., 1992, 64, 1534.
5. H. Hisamoto, N. Miyashita, K. Watanabe, E. Nakagawa, N. Yamamoto and K. Suzuki, Sens. Actuators, B, 1995, 29, 378.
6. M. Lerchi, F. Orsini, Z. Cimerman, E. Pretsch, D. A. Chowdhury and S. Kamata, Anal. Chem., 1996, 68, 3210.
7. M. R. Shortreed, S. Dourado and R. Kopelman, Sens. Actuators, B, 1997, 38, 8.
8. J. M. Dubach, D. I. Harjes and H. A. Clark, J. Am. Chem. Soc., 2007, 129, 8418.
9. B. Kuswandi, Nuriman, H. H. Dam, D. N. Reinhoudt and W. Verboom, Anal. Chim. Acta, 2007, 591, 208.
10. A. A. Ensafi and M. Fouladgar, Sens. Actuators, B, 2009, 136, 326.
11. M. T. Bamsey, A. Berinstain and M. A. Dixon, Sens. Actuators, B, 2014, 190, 61.
12. G. Mistlberger, G. A. Crespo, X. J. Xie and E. Bakker, Chem. Commun., 2012, 48, 5662.
13. X. J. Xie, G. Mistlberger and E. Bakker, J. Am. Chem. Soc., 2012, 134, 16929.
14. X. J. Xie, G. Mistlberger and E. Bakker, Anal. Chem., 2013, 85, 9932.
15. X. J. Xie and E. Bakker, ACS Appl. Mater. Interfaces, 2014, 6, 2666.
16. G. Mistlberger, X. J. Xie, M. Pawlak, G. A. Crespo and E. Bakker, Anal. Chem., 2013, 85, 2983.
17. G. Mistlberger, M. Pawlak, E. Bakker and I. Klimant, Chem. Commun., 2015, 51, 4172.
18. A. Shvarev, J. Am. Chem. Soc., 2006, 128, 7138.
19. X. J. Xie, G. Mistlberger and E. Bakker, Sens. Actuators, B, 2014, 204, 807.
20. V. Adler, A. Schaffer, J. Kim, L. Dolan and Z. Ronai, J. Biol. Chem., 1995, 270, 26071–26077.
21. K. Seiler and W. Simon, Sens. Actuators, B, 1992, 6, 295.
22. K. Seiler and W. Simon, Anal. Chim. Acta, 1992, 266, 73.
23. E. Bakker and W. Simon, Anal. Chem., 1992, 64, 1805.
24. X. J. Xie and E. Bakker, Anal. Bioanal. Chem., 2015, 407, 3899.
25. E. Bakker, P. Buhlmann and E. Pretsch, Chem. Rev., 1997, 97, 3083.
26. P. Buhlmann, E. Pretsch and E. Bakker, Chem. Rev., 1998, 98, 1593.
27. Z. Shi, P. Peng, D. Strohecker and Y. Liao, J. Am. Chem. Soc., 2011, 133, 14699.
28. V. K. Johns, Z. Z. Wang, X. X. Li and Y. Liao, J. Phys. Chem. A, 2013, 117, 13101.
29. V. K. Johns, P. K. Patel, S. Hassett, P. Calvo-Marzal, Y. Qin and K. Y. Chumbimuni-Torres, Anal. Chem., 2014, 86, 6184.
30. P. K. Patel, V. K. Johns, D. M. Mills, J. E. Boone, P. Calvo-Marzal and K. Y. Chumbimuni-Torres, Electroanalysis, 2015, 27, 677.
31. S. Peper, I. Tsagkatakis and E. Bakker, Anal. Chim. Acta, 2001, 442, 25.
32. V. K. Johns, P. Peng, J. DeJesus, Z. Z. Wang and Y. Liao, Chem. – Eur. J., 2014, 20, 689.
33. M. Q. He, T. M. Leslie and J. A. Sinicropi, Chem. Mater., 2002, 14, 2393.
34. S. Liu, M. A. Haller, H. Ma, L. R. Dalton, S. H. Jang and A. K. Y. Jen, Adv. Mater., 2003, 15, 603.
35. E. Bakker, M. Willer and E. Pretsch, Anal. Chim. Acta, 1993, 282, 265–271.
36. P. Gehrig, B. Rusterholz and W. Simon, Chimia, 1989, 43, 377.
37. D. Diamond, G. Svehla, E. M. Seward and M. A. Mckervey, Anal. Chim. Acta, 1988, 204, 223.
38. Y. Umezawa, P. Buhlmann, K. Umezawa, K. Tohda and S. Amemiya, Pure Appl. Chem., 2000, 72, 1851.

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Footnote

† Electronic supplementary information (ESI) available: Experimental methods. See DOI: 10.1039/c5an02206a

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