Conjugated polymers as robust carriers for controlled delivery of anti-inflammatory drugs

Electrochemical measurements and conditioning were performed in a three-electrode cell by use of CH Instruments 620 electrochemical workstation. The electrochemical synthesis of polymers was performed by means of cyclic voltammetry; the process of conditioning was carried out potentiostatically. The concentration of released IBU was monitored by means of Hewlett Packard 8453 UV–Vis Diode Array Spectrophotometer.

A three-electrode electrochemical cell was set up in a 3-ml electrochemical glass cell with 1 cm² platinum foil working electrode, Ag/AgCl as a reference electrode and glassy carbon rod as an auxiliary electrode. The polymer films containing IBU (PEDOT/LiClO₄/IBU) were obtained by means of cyclic voltammetry (CV) in aqueous solution containing 10 mM EDOT, 10 mM IBU, 0.1 M LiClO₄, which was sonicated prior to the experiment for 30 min. Typical polymer film was formed on Pt electrode within the potential range of 0–1.1 V (vs. Ag/AgCl) at the scan rate v = 100 mV s⁻¹ for 25 potential cycles.

In the three-step synthesis, the same three-electrode electrochemical cell was used as that described in the previous section. In the first step, the polymer film was obtained from aqueous solution containing 10 mM EDOT and 0.1 M LiClO₄ in the similar procedure employing cyclic voltammetry with a narrower potential range of 0–1.0 V (vs. Ag/AgCl), scan rate v = 100 mV s⁻¹, and number of potential cycles equal to 50.

In the second step, the original ClO₄ ⁻ dopant was removed from the polymer matrix by conditioning it at a reduction potential E _red = −0.7 V (vs. Ag/AgCl) for 10 min in 0.1 M LiClO₄ aqueous solution. The IBU immobilisation occurred in the third step, when the polymer matrix after rinsing with deionised water was oxidised at a potential E _ox = 0.8 V (vs. Ag/AgCl) in 0.4 M IBU aqueous solution for 10 min.

The concentration of IBU released from PEDOT/LiClO₄/IBU matrix was determined using UV–Vis spectroscopic measurements under potentiostatic conditions. The three-electrode electrochemical cell was set up in a 2-mm quartz cuvette, in which the PEDOT/LiClO₄/IBU-modified platinum-working electrode was mounted together with Ag wire used as a pseudoreference electrode, and graphite rod as an auxiliary electrode. Before measurements, the electrode covered with PEDOT/LiClO₄/IBU was soaked in PBS solution for 10 min to wash off any loosely attached IBU and unreacted monomer. In order to release IBU from PEDOT/LiClO₄/IBU matrix, a constant potential in the range −0.8 to 0.8 V (vs. Ag/AgCl) was stepped for a specified period of time. After each period of time at the given potential, the UV–Vis spectra were recorded for IBU released to the solution. All these measurements of the controlled release of the immobilised IBU were performed in the pH 6.5 phosphate buffer solution, PBS, containing 0.15 M KCl, 0.006 M K₂HPO₄, 0.001 M KH₂PO₄.

The stability of PEDOT/LiClO₄/IBU matrix exposed to a given electrode potential was determined in terms of the charge storage capacity, CSC, of the matrix. That factor was calculated as the electric charge integrated under corresponding CV curve during one CV cyclewhere t ₁ is the beginning of CV cycle, t ₂ is the end of CV cycle.

The CVs were recorded for the modified platinum electrode in PBS after 20 min exposition to a given potential.

As many other anions, IBU in its ionic form (Scheme 1) is able to participate in the polymer doping process, which is a part of the electropolymerisation reaction. Molecules like EDOT when oxidised (chemically or electrochemically) extend their conjugated bonding system, forming positively charged polymer chain that is stabilised by doping of anions from the reaction environment. Under the electrochemical conditions, the polymerisation/doping process occurs at positive potentials applied to the electrode in the electrolyte solution containing monomer. Thus, a presence of IBU⁻ in the electrolyte solution may lead to its immobilisation in the polymer matrix, which is created on the electrode surface. This process is shown in Scheme 2.

The current versus potential curves recorded in subsequent scans make it possible to observe the progress of electropolymerisation process, as shown in Fig. 1. Two sets of CV curves were collected, the first for the solution containing EDOT, LiClO₄ and IBU (Fig. 1a) and the second for the case, when IBU was not present in the solution (Fig. 1b). In both cases, the increase of current indicates that the monomer is irreversibly oxidised near 1.1 V (vs. Ag/AgCl). Also for both data sets, the gradually increasing currents confirm formation of conductive deposits, typical for the electropolymerisation process. At the same number of potential scans, the final level of the resulting currents and corresponding electric charge are evidently different, indicating a slower process of electrochemical polymerisation of EDOT in case of IBU presence in solution (Fig. 1a). This may be due to the fact that the molecule of IBU is significantly larger than the molecule of electrolyte, thus the incorporation of drug into the structure of the polymer is less spatially favourable. The preliminary results indicated that the molar ratio of EDOT to IBU equal to 1:1 produces a polymer with the highest drug content, approximately 60 μg cm⁻² (mass of IBU immobilised in the unit surface area of Pt electrode). The charge storage capacity, CSC, calculated for PEDOT/LiClO₄/IBU (22.3 mC) was however not as large as for PEDOT/LiClO₄ film (43.6 mC) polymerised under analogous conditions as indicate the CV curves shown in Fig. 2. The CSC factor used in here is simply current integrated along the potential axis recorded for the polymer-coated electrode placed in the pure electrolyte solution. It was found that the use of higher IBU concentrations suppresses the formation of polymer film, therefore it has adverse effects on its charge storage capacity. When IBU was present at lower concentration in the monomer solution (1, 2, or 5 mM), the amount of IBU immobilised in the polymer film was as low as 5, 6 and 11 μg cm⁻², respectively.

The three-step synthesis of PEDOT/LiClO₄/IBU matrix allows to separate the processes of electropolymerisation and immobilisation. In this procedure, the polymer film is formed in pure electrolyte solution, thus it is doped only with perchlorate ions. Electrochemical reduction of the film eliminates to some extent that dopant from the polymer. Oxidation of the polymer is then carried out in the aqueous solution of IBU only. The CVs recorded on Pt electrodes modified with PEDOT/LiClO₄ and PEDOT/LiClO₄/IBU obtained via one-step (1) and three-step (3) procedures are compared in Fig. 2. The average current magnitude of PEDOT/LiClO₄/IBU(3) is larger than for PEDOT/LiClO₄/IBU(1) and only slightly smaller than that for PEDOT/LiClO₄. Hence, it may be concluded that a three-step synthesis route results in a matrix with a higher CSC (40.9 mC) than matrix synthesised according to the one-step procedure (22.3 mC), only slightly smaller than CSC of PEDOT/LiClO₄ film (43.6 mC).

During the synthesis, anionic compounds are immobilised in a positively charged conducting polymer matrix to maintain a neutral charge of the polymer backbone. When the sufficiently negative voltage is applied to the matrix, the anionic dopant becomes unnecessary, therefore it is released to the solution. The schematic representation of the electrically triggered drug release from conducting polymer matrix is shown in Scheme 3.

The process of IBU release from PEDOT/LiClO₄/IBU matrix was monitored under potentiostatic conditions using UV–Vis spectroscopy. Calibration curve was plotted for absorbance versus IBU concentration (where y is the absorbance and x is the IBU concentration in mM). A linear relationship was observed between 0.01 and 1 mM IBU concentration satisfying the equation y = 0.0176x + 1.9365 (R ² = 0.9992).

In order to release IBU from PEDOT/LiClO₄/IBU matrix, a constant potential in the range between −0.8 and 0.8 V (vs. Ag/AgCl) was applied to the matrix placed in the cuvette. The absorbance at λ = 222 nm, which is characteristic for IBU, was recorded versus time at a given potential. The changes of absorbance spectra in time for the matrix stimulated with E = −0.5 V (vs. Ag/AgCl) are presented in Fig. 3.

The time profiles of IBU release from the matrix stimulated with E = −0.5 V (vs. Ag/AgCl) and matrix maintained under open circuit conditions are presented in Fig. 4. As it can be seen, for the non-stimulated matrix (open circuit potential), IBU was released in small concentration, six times lower than for the matrix stimulated with the reducing potential E _red = −0.5 V (vs. Ag/AgCl). This result evidently confirms the effect of the applied potential on the process of the drug release. The greatest quantity of IBU appears to be released during the first 2 min of the electric stimulation, as compared to longer times. It suggests that only a small portion of IBU molecules is imbedded deeply within the polymeric structure of the matrix; also the process of full reduction of PEDOT is practically completed within the applied stimulation time. Under the described conditions, a total time below 10 min is required to reach equilibrium (t _equil) in the system, then the solution concentration of IBU is stable.

Since the ratio of the amounts of oxidised and reduced forms of the conjugated polymer is strictly determined by the potential applied to the polymer film, therefore one can expect variation in the concentration of IBU released to the solution as a function of the value of stimulating potential. These data were collected for t _equil = 10 min for polymer films of approximately the same amount of immobilised IBU. The resulting concentration of IBU released as a function of applied potential is shown in Fig. 5. The maximum concentration of IBU released, 0.66 (±0.10) mM, was observed when the potential E = −0.5 V (vs. Ag/AgCl) was applied. When PEDOT/LiClO₄/IBU was subjected to more negative potentials, the concentration of released IBU decreased. This is caused by the effect of n-doping of negatively charged polymer matrix with cations present in PBS solution: the incorporation of potassium cations occurs instead of the release of the anionic form of IBU. The concentration of IBU released decreases as the value of applied potential increases since the positively charged film holds the anionic drug; the concentration of IBU released at E = 0.8 V (vs. Ag/AgCl) was 0.21 (±0.05) mM, which is one-third of the concentration released at E = −0.5 V (vs. Ag/AgCl). These results are consistent with the mentioned above effects of the applied potential on the redox state of conjugated polymer. The application of negative potentials results in facilitated release of anionic co-dopant, IBU, while the application of positive potential results in the drug retention.

The capacity of the polymer matrix towards the amount of immobilised IBU generally depends on the amount of polymer; the last in turn is easy to control in the process of electropolymerisation by variation of the number of CV cycles. The relation between the drug capacity of the matrix and the conditions of electrochemical polymerisation, in terms of number of CV cycles, is presented in Fig. 6. The data indicate nearly linear relation between the electropolymerisation time and the drug capacity; the first is represented by the number of CV scans, whereas the other is expressed in terms of the concentration of released IBU. These results demonstrate that it is possible to synthesise PEDOT/LiClO₄/IBU matrix of a specified IBU content that can be further effectively released.

Since the three-step synthesis of PEDOT/LiClO₄/IBU allows to separate the processes of electropolymerisation and immobilisation, it should be possible to apply the immobilisation-release procedure several times for the same matrix. Again, the curves representing the concentration of IBU released versus time were plotted for the PEDOT/LiClO₄/IBU matrix that was subjected to four consecutive immobilisation-release cycles. The plot of the concentration of released IBU calculated from the corresponding UV–Vis data is shown in Fig. 7. After the first cycle, the concentration of released IBU was equal to 0.70 (±0.05) mM, whereas each subsequent immobilisation/release cycle appeared to be less efficient; in the forth cycle the concentration of IBU released was reduced to 0.34 (±0.02) mM. The observed effect is caused by the gradual degradation of PEDOT/LiClO₄/IBU matrix that usually occurs when the polymer is exposed to oxidative potential exceeding its optimal value (overoxidation). If, however, the potential is carefully chosen, the polymer degradation during its destructive oxidation can be optimised. On the other side, the matrix exposed to negative potentials for longer times (or at no potential applied) keeps its CSC factor practically unchanged.

The gradual loss of the matrix loading effectiveness, as observed in the spectral measurements, is likely caused by a degradation of the polymer film. The CV curves recorded after each IBU load/release cycle reveal decreasing electric charge. These data shown in terms of CSC values (Table 1) are consistent with the spectral results, which all together give a proof of a limited durability of the PEDOT matrix when multiple IBU- loadings are attempted.