Reliable low-cost capillary electrophoresis device for drug quality control and counterfeit medicines

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Abstract

The proportion of counterfeit medicines is dramatically increasing these last few years. According to numerous official sources, in some pharmaceutical wholesalers in African countries, the proportion has reached 80%. Unfortunately, this situation is far to be improved due to lack of suitable analytical equipment allowing rapid actions of the Regulatory Agencies based on scientific consideration, at affordable cost and all over the drug supply chain.

For that purpose, a network group considered that mater by building a low-cost original capillary electrophoresis (CE) equipment equipped with a new deep UV detector based on LED technology.

The generic conditions for analysis were investigated: capillary zone electrophoresis (CZE) performed at acidic pH for basic drug molecules (i.e., quinine, highly used as the last antimalarial rampart), basic pH for compounds such as furosemide (a common diuretic drug) and at neutral pH for a well known antibiotic combination, trimethoprim/sulfamethoxazol.

To evaluate the ability of the CE equipment for quantification, a full validation and a method comparison study were carried out for the CZE method dedicated to quinine determination. The validation involved the use of accuracy profile and total error concept to monitor the adequacy of the results obtained by the new prototype. The method comparison was based on the Bland and Altman approach by comparing results obtained by the low-cost CE and a conventional set-up. Subsequent validation studies were realized with neutral and acidic drug molecules, each focusing on a single concentration level calibration curve in order to maintain as low as possible the expenses due to reagents and thus the cost of analysis, as important advantages of CE for drug quality control.

Introduction

A counterfeit medicine is a medicine that is deliberately and fraudulently mislabelled with respect to identity and/or source [1], [2], [3]. Both branded and generic products can be concerned by the counterfeiting. Counterfeits may include products (i) with correct ingredients/components, (ii) with wrong ingredients/components, (iii) without active ingredients, (iv) with incorrect amounts of active ingredients or (v) with fake packaging [1], [3]. The consequences of counterfeiting are mainly observed among patients, the medicines end-user's supposed to be the beneficiaries. They are exposed to risk of consuming such counterfeit products, i.e., therapeutic failure or drug resistance [4]. Some extreme cases can lead to death. Dramatic examples are reported in Panama and Nigeria. In 2006, more than 100 patients have been killed in Panama by medicines manufactured with counterfeit glycerine; in 2008, about 100 babies died because of absorbing a false paracetamol syrup [3], [5]. The proportion of counterfeit medicines is dramatically increasing these days [6], [7], [8], [9], in particular in some African countries pharmaceutical wholesalers where it was reported up to 80% of counterfeiting [10]. Unfortunately consumers and prescribers are unable to assess the quality, safety and efficacy of medical products.

Considerable efforts are deployed in order to truly fight and prevent trade in counterfeit medical products [1], [3], [7], [10], [11], [12], [13]. One of them is based on the dissemination of information useful for assessing technologies aimed at preventing, deterring, or detecting counterfeit medicinal products. This is somewhat paradoxical in particular for poor/emerging countries since such practices require adequate/large infrastructure or facilities, i.e. well equipped laboratory that is very often lacking or simply not functioning. Unfortunately, lack/insufficient of such facilities reduces the capacity of the Regulatory Agencies to react rapidly and adequately based on scientific considerations, thus leading to ineffective control. Several analytical methods, including near infrared spectroscopy, Raman spectroscopy, refractometry, colorimetry, X-ray powder diffraction analysis, nuclear magnetic resonance [14], [15], [16], [17], [18], [19], [20], [21], [22] and separations techniques such as liquid chromatography (LC), thin layer chromatography, gas chromatography and capillary electrophoresis (CE) [23], [24], [25], [26], [27], [28], [29], [30] are used to analyse pharmaceutical substances, taking into account their variety of structure and chemical properties such as polarity and acidity. These methods need straightforward sample preparation and rapidity to support decisions in pharmaceutical fields, i.e., batch release or rejection, etc. However, for most of these techniques, financial expenses are not affordable to allow an easy implementation for a regular, systematic and wide-spread (or extended) control. Furthermore, the apparatus maintenance is one of the crucial issues allowing a long-term use of an analytical device in emerging countries.

Last 20 years, CE has gained importance for its ability to analyze several compounds with good selectivity [27], [28]. Several aspects can largely contribute to the CE implementation in poor/emerging countries: simple, reliable and (cost-)efficient drug control methods, financial expenses, i.e., solvent and reference material consumption [31], water is often the solvent of choice, ease of operation (no need of complex solvent gradients [32]). Finally, several generic conditions have been reported for separating molecules. In this context, the University of Applied Sciences Western Switzerland, College of Engineering and Architecture of Fribourg (UAS-WS-FR) has developed in collaboration with the School of Pharmaceutical Sciences, University of Geneva-Lausanne and the Geneva University Hospital (HUG) a low cost analytical device, based on capillary electrophoresis (CE), with the aim to use it for educational purpose in developing and transitional countries. Three prototypes were built, the first one, within the period 2006–2007, where the mechanical and electronical issues were assessed, including an original detection device built on the basis of diode technology. The second, within the period 2007–2008, for the software and ergonomic optimisations and the third one (2009), was used for the development of methods of drugs analysis, selected toward a list submitted by African partners located in Mali, where it is now located. In the method development stage, in collaboration with the Institute of Pharmacie, University of Liège, a particular attention was paid to the robustness of the system in order to anticipate the problems that can be encountered when dealing with analytical methods in Africa as well as the quantitative aspects of quality controls of drugs.

In this paper, the analytical performance and the ability of the developed low-cost CE equipment for quantification is presented. A complete validation study with minimal requirement in regards to calibration purposes, were performed on representative drugs with different physico-chemical properties: quinine (QUN), furosemide (FUR) and the combination trimethoprime (TMP)/sulfamethoxazol (SMX) (see chemical structures in Fig. 1). These medicines represent one of the most targeted pharmacological groups by counterfeiting namely the antimalarial (21% of counterfeiting), diuretic (9%) and anti-effective (12%) drugs, respectively [35]. A strategy based on the total error of measurement was applied [33], [34] and completed with a method comparison in order to evaluate the performance of the low-cost CE equipment to another one commonly available on the market.

Section snippets

Instrumentation

Experiments for method development and validation of QUN were performed on an Agilent HP3DCE system (Hewlett-Packard, Waldbronn, Germany) equipped with an on-column diode-array detector, an autosampler, a high-velocity air-cooled capillary cartridge, a power supply able to deliver up to 30 kV and an external pressure system. A CE ChemStation software version Rev. A.10.02 was used to control the CE instrument, to acquire and to handle the data. Separations were performed in bare fused-silica

CE method development

In this study, CE methods were developed for identification of counterfeit drugs, with a focus on the determination of the presence of the active ingredient and the possibility to rapidly decipher regarding the correct amount of the active ingredient. Thus, several aspects needed to be taken into account: (i) simple and generic methods, in order to analyze a high number of compounds and make easier the method selection for each analyte with basic chemistry knowledge; (ii) low cost methods,

Conclusion

Drug counterfeits is a major public health issue in some poor and emerging countries. In order to fight this problem a robust and low cost analytical device was developed. Three methods were developed for the quantitative analysis of active substances present in pharmaceutical formulations subjected to be counterfeited and selected according to their therapeutic uses. For these methods, among the counterion tested, Tris+ gave the best results. Suitable parameter values were obtained allowing

Acknowledgements

A research grant from the Belgium National Fund for Scientific Research (FRS-FNRS) to E. Rozet is gratefully acknowledged as well as the research grant from the Belgian Coopération Universitaire au Développement (CUD, R.D. Marini) and Walloon Project PPP (Convention OPTIMAL DS No. 917007, R.D. Marini). Research grant from University of applied sciences of western Switzerland (Fribourg) Projet d’institut iTIN, Sagex No. 11480/19870 were received for the low-cost CE device development (C.

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