Mercury-free and modification-free electroanalytical approach towards bromazepam and alprazolam sensing: A facile and efficient assay for their quantification in pharmaceuticals using boron-doped diamond electrodes
Introduction
Benzodiazepines (BDZs) have become one of the most widely used drugs, since the chlordiazepoxide and diazepam were launched on global market in 1960s. These drugs were found to exhibit anticonvulsant, hypnotic, sedative and muscle-relaxant effects [1]. For this reason, BDZs have been indicated in medical use for the treatment of anxiety, insomnia, depression, psychiatric disorders and alcohol withdrawal syndromes [2], [3]. Nowadays, these drugs are mostly abused due to their toxic effects and widespread availability [4]. Based on the pharmaceutical relevance and medicinal objectives of BDZs, advanced and perspective analytical approaches of high efficiency for the rigid control of these important substances in pharmaceutical formulations and different biological fluids are still required.
Many analytical methods have been recently developed for determination of BDZs, in particular, high performance liquid chromatography (HPLC) coupled with diode array detector (DAD) [5], [6] and tandem mass spectrometry (MS/MS) [7], [8], gas chromatography (GC) in combination with MS [9], electrophoresis [10] and spectrofluorimetry [11]. These remarkable analytical methods are high sensitive and selective requiring low sample consumption and enabling simultaneous determination of plenty of miscellaneous analytes. On the other hand, they suffer from expensive instrumentation, time consuming and very costly analysis as well as need for the sample derivatization is required prior to analysis.
Electroanalytical methods represent financially unassuming, temporally efficient and sensitive tool for detection and determination of various structurally and biologically interesting substances [12], [13]. The literature survey states that hanging mercury drop electrodes (HMDE) [14], [15], [16] and silver solid amalgam electrode (AgSAE) [17] have been applied to sensitive determination of several BDZs derivatives based on their electrochemical reduction. Besides, various chemically modified carbon-based electrodes such as glassy carbon electrode (GCE) modified with multi-walled carbon nanotubes (MWCNT) [18] and 3-methylthiophene [19] as well as carbon paste electrode (CPE) modified with MWCNT [20] and bentonite [21] have been exploited as mercury-free electrochemical platforms for quantifying BDZs. In order to achieve low detection limit (LOD), a modification of carbon-based electrode substrates appeared to be favorable for supporting electron transfer in redox reaction, for decreasing peak potential and principally for increasing sensitivity. Nevertheless, bare (unmodified) conventional carbon-based electrode materials are known for their higher background with limited usable potential window in negative area due to hydrogen evolution reaction. For electroanalytical determination of BDZs, differential pulse voltammetry (DPV) and square-wave voltammetry (SWV) coupled with stripping steps such as square-wave adsorptive cathodic stripping voltammetry (SWAdCSV), square-wave cathodic stripping voltammetry (SWCSV), differential pulse stripping voltammetry (DPSV), differential pulse adsorptive cathodic stripping voltammetry (DPAdCSV) and linear sweep adsorptive cathodic stripping voltammetry (LSAdCSV) have been commonly used. In this respect, stripping voltammetry is an efficient technique routinely capable of achieving LODs lower than 10−8 mol/L due to preconcentration of analyte on particular working electrode [22]. In spite of convenient application of HMDEs and modified carbonaceous electrodes in BDZs sensing, researchers are still forced to search the advanced and perspective electrode materials to be applied as sensitive electrochemical sensors, even without performing any surface modification and using toxic mercury electrodes. It is worth noting that the authors of the present work have recently demonstrated the use of meniscus modified silver solid amalgam electrode (m-AgSAE) as prospective electrode material for the quantification of selected BDZs with LODs of 10−7 mol/L [17].
Boron-doped diamond (BDD) is a perspective and “green” carbon-based electrode material exhibiting many advantages when compared to conventional materials such as CPE, GCE or HMDE. BDD electrode (BDDE) renders high chemical and electrochemical stability owing to sp3 hybridization of carbon in diamond structure (chemically inert character), wide potential window in the aqueous and non-aqueous media as well as low and stable background current [23], [24]. Apart from these properties, it yields excellent biocompatibility, mechanical robustness and stability in alkaline and acidic media [25], [26]. BDD electrode surface can be treated electrochemically by applying very negative and positive potentials (cathodic and anodic pretreatment) to obtain predominantly hydrogen and oxygen terminated surface (hydrophobic and hydrophilic nature), respectively, which may change overall chemical properties of this working electrode [13]. Recently, interesting reviews on practical guide to usage of BDDE in electrochemical research [27] as well as possibilities of chemical modification of this material for biosensors and biosensing [28] have also been published. In addition, BDDE has seemed to be an efficient electrochemical sensor for detection and determination of significant biologically active compounds used in protection of human health [29], environment [30] and food analysis [31].
This paper reports the novel application of BDDEs as advanced, mercury-free and modification-free electrochemical platforms suitable for the individual determination of the selected electrochemically reducible BDZs (Fig. S1), namely bromazepam (7-Bromo-5-(2-pyridyl)-3H-1,4-benzodiaxepin-2(1H)-one, BZ) and alprazolam (8-Chloro-1-methyl-6-phenyl-4H-[1,2,4]triazolo[4,3-a][1,4]benzodiazepine, ALZ). The feasibility of the developed assays was demonstrated by the analysis of commercial pharmaceuticals. Moreover, as far as we know, these procedures could be considered as the first usage of BDD as advanced carbon-based electrode material for the reliable quantification of BDZs in pharmaceutical samples.
Section snippets
Chemicals
BZ (CAS No. 1812-30-2, purity ≥ 98%, in all figures presented as A) and ALZ (CAS No. 28981-97-7, purity ≥ 98%, in all figures presented as B) were purchased from Sigma Aldrich (Czech Republic). 1 × 10−3 mol/L stock solutions were prepared by dissolution of appropriate amount of BZ and ALZ in 25 mL methanol (p.a., Lachema, Czech Republic) without any further purification. These solutions were stored in the fridge for following use. Britton-Robinson buffer (BR) was prepared by mixing of 0.04 mol/L H3BO3
Effect of pH and reversibility study
The choice of the suitable pH of supporting electrolyte is an important stage in the electroanalytical studies because it affects the properties of the solution and the electrode-solution interface, thus modifying the thermodynamics and kinetics of the particular charge transfer process. The effect of pH on the peak current density (JBZ and JALZ) for the individual BDZ solutions of 1 × 10−4 mol/L was investigated using differential pulse voltammetry (DPV) in BR buffers (pH 2–12). The results
Conclusion
Recent analytical methods dedicated to BDZs quantification are relatively of high cost, generate a high amount of toxic organic solvent with time consuming analysis and need for sample derivatization. In this work, the commercial and lab-made BDDEs with various boron-doping levels were exploited for the first time in connection with DPV technique to elaborate the novel and advanced electrochemical protocols for simple analytical determination of the selected BDZs. CV experiments revealed that
Acknowledgements
This work has been supported by the Student grant competition (University of Ostrava)SGS07/PřF/2015 and SGS03/PřF/2016, the Grant Agency of the Slovak Republic (grant No. 1/0489/16), the Slovak Research and Development Agency (APVV-0365-12), the Ministry of Education and Science of the Republic of Serbia (projects No. OI 172030) and MagBioVin project. ĽŠ also thanks the STU Grant scheme for Support of Excellent Teams of Young Researchers.
MSc. Petr Samiec is a young researcher at the Ostrava University, Czech Republic who is interested in electroanalytical chemistry of pharmaceuticals and application of various types of electrode materials for their analysis.
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MSc. Petr Samiec is a young researcher at the Ostrava University, Czech Republic who is interested in electroanalytical chemistry of pharmaceuticals and application of various types of electrode materials for their analysis.
Assoc. Prof. Ľubomír Švorc, Ph.D is a researcher under the age of 35 who received Ph.D. in 2009 and habilitated (Associate Professor) in 2015 in Analytical Chemistry at the Slovak University of Technology in Bratislava. His research interest mostly covers the development and application of perspective electrochemical sensors for clinical, pharmaceutical, food and environmental analysis. He has authored and co-authored more than 80 journal papers with h-index 16. He is the head of the Division of Analytical Chemistry of the Slovak Chemical Society and the Laureate of Young Scientist Award 2014 in Slovakia.
Dr. Dalibor M. Stanković is a young researcher whose research interest is electroanalytical chemistry and electrochemistry of biological active compounds, application of novel materials for modification of electrodes and their application in environmental analysis and biomedical analysis. He is authored and co-authored around 60 publications in this topic.
Dr. Marian Vojs received his Ph.D. in Electronics in 2009 at the Slovak University of Technology in Bratislava. Since 2009 he is working as researcher at the Institute of Electronics and Photonics, Faculty of Electrical Engineering and Information Technology, Slovak University of Technology in Bratislava. His main interests include deposition of boron doped diamond thin films, carbon structures and characterization of morphology, electrical and electrochemical properties and Raman spectroscopy.
Dr. Marián Marton received his Ph.D. in electronics and vacuum technology at the Slovak University of Technology in Bratislava. Currently his research deals with synthesis, analysis and applications of carbon nanomaterials, e.g. diamond, CNTs, CNWs and DLC. He is an expert in CVD technology and Raman spectroscopy.
Assoc. Prof. Zuzana Navrátilová, Ph.D received Ph.D. in 1990 and habilitated (Associate Professor) in 2006 in Analytical Chemistry at the University of Pardubice, Czech Republic. She is experienced in the field of study of sorption and ion-exchange properties of clay minerals by means of modified electrodes and analysis of environmentally significant substances (metals, gas emissions, humic substances). She guarantees the master and doctoral study programmes of Analytical Chemistry at the Ostrava University, Czech Republic.