Voltammetric Determination of Tannic Acid in Medicinal Plants Using Multi-Walled Carbon Nanotube-Modified Electrode ^†

Abstract

Tannins are a class of natural phenolic antioxidants represented by oligomers and polymers. Tannic acid (gallotannin) (TA) is one of the most typical compounds widely distributed in plants. It has a wide application area in food technology and as a part of traditional herbal medicine in the treatment of various diseases. TA is one of the standards used in medicinal plant quality control for plants rich in tannins. Therefore, the development of sensitive and simple methods for TA quantification is of practical interest. Glassy carbon electrode modified with multi-walled carbon nanotubes (GCE/MWCNTs) has been developed for the determination of TA in medicinal plants. An improvement in TA voltammetric response has been achieved using modified electrode due to the high electroactive surface area and electron transfer rate vs. bare electrode. TA electrooxidation at the GCE/MWCNTs is an irreversible surface-controlled process involving the transfer of two electrons and two protons in the first step. In differential pulse mode using Britton–Robinson buffer pH 2.0 as the supporting electrolyte, a linear dynamic range of 0.10–7.5 μM with a detection limit of 0.038 μM has been obtained. The method has been applied for the analysis of infusions and decoctions of tannin-containing medicinal plants and compared to the spectrophotometric method. A positive correlation has been observed with ferric reducing power reflecting the total content of phenolic compounds in the sample.

1. Introduction

Tannins are a class of natural phenolic antioxidants represented by oligomers and polymers. Tannic acid or gallotannin (TA) (Figure 1) is one of the most typical compounds widely distributed in plants [1]. It has a wide application area in food technology [2,3] and as a part of traditional herbal medicine in the treatment of various diseases [4,5,6]. TA is one of the standards used in medicinal plant quality control for plants rich in tannins. Therefore, the development of sensitive and simple methods for TA quantification is of practical interest.

The presence of phenolic moieties in the TA structure makes it electroactive and able to be measured using voltammetry. Nevertheless, a lack of electrochemical methods has been reported for TA determination compared to other natural phenolics. Traditional carbon-based [7,8] and platinum [9] electrodes show low sensitivity and selectivity of TA response, which are the main limitations of their practical application. Various chemically modified electrodes have been developed to date to solve this problem. Carbon nanomaterials [10,11,12,13], porous nanomaterials [14,15,16], metal nanoparticles [17,18], electrodeposited nickel(II) hexacyanoferrate [19], polymeric coverages [20,21], and 1-benzoyl-3-(pyrrolidine) thiourea film [22] have been shown to be effective electrode surface modifiers for TA quantification. In most cases, adsorptive preconcentration for 2.5–5 min is used to improve the analytical characteristics of TA. This step increases the measurement duration and can lead to the co-adsorption of other components from the real samples. Furthermore, the analytical characteristics of the existing methods are not impressive (the linear ranges cover mainly µM concentrations with the detection limits of n × 10^–7 M).

The current work is focused on the development of a fast and sensitive voltammetric approach for TA determination using glassy carbon electrode modified with multi-walled carbon nanotubes (GCE/MWCNTs) and its application in medicinal plant analysis.

2. Materials and Methods

TA (ACS reagent grade) from Sigma-Aldrich (Saint Louis, MO, USA), 99% gallic and 99% ascorbic acids, 98% quercetin dihydrate from Sigma (Steinheim, Germany), and 97% rutin trihydrate from Alfa Aesar (Heysham, UK) were used. Their standard 10 mM (1.0 mM for rutin) solutions were prepared in ethanol (rectificate). Exact dilution was used for the preparation of less concentrated solutions.

MWCNTs (o.d. 40–60 nm, i.d. 5–10 nm, and l = 0.5–500 μm) from Sigma-Aldrich (Steinheim, Germany) were applied as the electrode surface modifier. Sodium lauryl sulfate (97.2% purity) from Panreac (Barcelona, Spain) was used as the dispersive agent for MWCNTs. A homogeneous suspension of MWCNTs in 1% sodium lauryl sulfate (0.5 mg mL⁻¹) was prepared through sonication for 30 min in an ultrasonic bath (WiseClean WUC-A03H) (DAIHAN Scientific Co., Ltd., Wonju-si, Republic of Korea).

Commercial medicinal plant materials (Quercus sp. cortex, Bergenia crassifolia (L.) Fritsch rhizomata, Potentilla erecta (L.) Raeusch. rhizomata, Alnus incana (L.) Moench and Alnus glutinosa (L.) Gaertn. fructus, Sanguisorba officinalis L. rhizomata et radices) were studied. Infusions and decoctions were prepared using a standard pharmacopoeia procedure [23].

Other reagents were c.p. grade and used as received. Distilled water was used for the supporting electrolyte preparation. The laboratory temperature was 25 ± 2 °C.

Electrochemical measurements were conducted on the potentiostats/galvanostats µAutolab Type III (Eco Chemie B.V., Utrecht, The Netherlands) with Nova 1.7.8 software and Autolab PGSTAT 302N with the FRA 32M module (Metrohm Autolab B.V., Utrecht, The Netherlands) and NOVA 1.10.1.9 software. A glassy electrochemical cell of 10 mL with a three-electrode system (working GCE of ø = 3 mm (CH Instruments, Inc., Bee Cave, TX, USA) or an MWCNT-modified GCE, an Ag/AgCl reference electrode, and a platinum wire as the auxiliary electrode) was used.

An “Expert-001” pH meter (Econix-Expert Ltd., Moscow, Russia) with a glassy electrode was used for the pH measurements.

The ferric reducing power of the decoctions and infusions was measured through coulometric titration with electrogenerated ferricyanide ions [24] using a coulometric analyzer “Exper-006” (Econix-Expert, Moscow, Russia) with four platinum electrodes (two of them as working and auxiliary electrodes in the generating circuit and another two polarized needle electrodes in the indicator circuit).

3. Results and Discussion

3.1. Electrooxidation of TA

The voltammetric characteristics of TA at the bare GCEs and GCE/MWCNTs have been studied using cyclic voltammetry in Britton–Robinson buffer pH 2.0 (Figure 2). Electrode surface modification provides a significant improvement in the voltammogram shape due to the 20.3-fold increase in the oxidation currents. Such an effect can be explained by the increase in the electroactive surface area of GCE/MWCNTs (75 ± 2 vs. 8.9 ± 0.3 mm² for bare GCE on the basis of electrochemical data for 1.0 mM ferrocyanide ions oxidation) as well as the preconcentration of TA on the electrode surface. The last assumption has been confirmed using cyclic voltammetry data for a study of the potential scan rate effect.

A 30 mV cathodic shift in the TA oxidation potential has been observed at the GCE/MWCNTs, which is caused by the electrocatalytic effect of the MWCNTs leading to the higher electron transfer rate at the modified electrode (5.23 × 10^–5 and 3.11 × 10^–4 cm s^–1/c for bare and modified GCEs, respectively, as the electrochemical impedance spectroscopy data show).

TA electrooxidation is accompanied by proton transfer since the oxidation potentials in both steps are shifted to a lower value with the pH increase from 2.0 to 7.0. An equal number of electrons and protons participate in the electrode reaction. The voltammetric response of TA is decreased with the increase in the pH value and fully disappears in basic medium due to the oxidation with air oxygen. Therefore, Britton–Robinson buffer pH 2.0 has been used in further study.

The linear plot I_p vs. potential scan rate and slope of 1.08 for ln I_p vs. lnυ indicate a surface-controlled electrode reaction. The number of electrons has been calculated using the Laviron equation to be two. Thus, TA electrooxidation is an irreversible two-electron and two-proton process, which is in good agreement with results reported previously [16,17].

3.2. TA Quantification

TA determination has been performed in differential pulse mode using a modulation amplitude of 75 mV and modulation time of 50 ms, providing the highest oxidation currents of the analyte. The oxidation peak at 0.49 V and a shoulder at 0.58–0.60 V are clearly pronounced on the differential pulse voltammograms of TA (Figure 3a).

TA oxidation currents are linearly increased with the concentration in the range of 0.10–7.5 µM (Figure 3b). The detection limit of 0.038 μM has been obtained. These analytical characteristics are improved compared to those reported for the electrodes also modified with MWCNTs in [10,12,13]. Moreover, the absence of a preconcentration step in the current work makes the procedure faster and more accurate as far as the co-adsorption of coexisting components is excluded.

The method developed shows a high accuracy of TA determination (recovery for model systems is 98.5–100%) and reproducibility (RSD for five measurements each on the new electrode is less than 2.0%).

3.3. Real Samples Analysis

The approach has been successfully applied in the analysis of infusions and decoctions of tannin-containing medicinal plant materials (Quercus sp. cortex, Bergenia crassifolia (L.) Fritsch rhizomata, Potentilla erecta (L.) Raeusch. rhizomata, Alnus incana (L.) Moench and Alnus glutinosa (L.) Gaertn. fructus, Sanguisorba officinalis L. rhizomata et radices). A well-defined oxidation step at 0.48–0.49 V and a shoulder at 0.58–0.60 V have been observed for all samples. The signal corresponds to TA, as a standard addition method indicates. The absence of matrix effects is confirmed by the recovery of 96.1–101%.

The results for the TA contents in medicinal plant infusions and decoctions are presented in Figure 4. The voltammetric method has been validated with UV-spectroscopy [25] and a good agreement of the results has been obtained.

A positive correlation (r = 0.8098) of TA contents in medicinal plant infusions and decoctions with ferric reducing power has been observed. Ferric reducing power reflects the total content of phenolic compounds in the sample.

4. Conclusions

Thus, a novel voltammetric approach has been developed for direct TA quantification using GCE/MWCNTs. The procedure is simple and fast, needs a low sample volume (5 µL), and excludes additional reagent consumption. The method developed can be applied for the analysis and standardization of tannin-containing medicinal plants.