Automated multiple development thin-layer chromatography for separation of opiate alkaloids and derivatives
Introduction
Opiates are the most powerful known pain relievers, they are derivatives of opium and can produce euphoria but also they are used as analgesics. Opiates can be classified according to three series. The first one is constituted by the poppy alkaloids: morphine, codeine, thebaine, noscapine and papaverine (Fig. 1); the second category included mainly semi-synthetic derivatives of morphine or synthetic compounds are used in therapy as antitussives and analgesics: pholcodine, ethylmorphine (codethyline), dextromethorphan (Fig. 2); at last, the third class is composed of narcotic compounds: diacetylmorphine (heroin), and other opiates; they are employed as substitutes in treatment of addiction: buprenorphine, and methadone (Fig. 3).
Automated multiple development (AMD) is an instrumental technique of planar chromatography [1], [2], [3], [4] which uses an eluent gradient starting from the most polar to the least polar [5]; the migration is performed by successive steps (15–25) and at each new development the proportions of the eluent constituents change; so the polarity is decreasing when the distance increases (Fig. 5).
Gradient development with linear eluotropic profile [6] leads to a band re-concentration improving the separation [1]. A successful separation depends mainly on the choice of the solvent components, optimisation of the shape of the gradient, the stepwise movement of the elution front [3], [7] and the repeated developments increase the resolution [8].
This chromatographic technique is available for separation of plant extracts [9], [10], [11], [12], [13], [14], [15], [16], mainly herbal drugs which present an interest with therapy activity and chemical substances belonging to various classes as: essential oils, alkaloids, resins, phenolic compounds. Besides, this method has permitted to analyse various chemical classes of alkaloids and phenolic compounds [10], [11], [12] with similar chemical properties and also isomeric position compounds [11]. In the field of detection of abuses, AMD was used with success for the chromatographic analysis of cannabinoids [13].
The aim of this work was to apply the performances of AMD to the separation of opium alkaloids, antitussives and narcotic compounds and to obtain a clean separation for each group studied.
Section snippets
Apparatus and materials
Chromatographic separations were performed using the AMD system (Camag, Muttenz, Switzerland).
Plates used were HPTLC silica gel F254 10 cm × 20 cm on glass, layer thickness 0.1 mm, Article 11764, and HPTLC silica gel F254 10 cm × 20 cm on glass, layer thickness 0.2 mm, Article 5642 (Merck, Darmstadt, Germany).
Samples were applied with a Linomat IV system (Camag, Muttenz, Switzerland).
All the solvents were analytical grade and purchased from Carlo Erba Reactifs (Val de Reuil, France). Before use, the
Separation of opiate alkaloids by classical TLC
The classical thin-layer chromatography analysis of opium alkaloids is not easy because these alkaloids belong to two different groups: morphinan (morphine, codeine and thebaine) and benzylisoquinoline (noscapine, papaverine) and their chromatographic behaviour is very different; the morphinan group presents low hRF and isoquinoline group very high hRF. For these reasons the authors use generally complex eluents, for example: diisopropyl ether–ethanol–diethylamine (97:2:1) [18],
Conclusion
Planar chromatography is always a choice method for the analysis of natural products and medicinal plants. It has become a modern technique with the arrival of automated apparatus for the application of samples and automated development chambers (OPLC and AMD) which are available techniques particularly to separate constituents in crude extracts.
OPLC and AMD are more appropriate in many instances because these techniques give better reproducible hRF values in well-defined experimental
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2017, Journal of Chromatography ACitation Excerpt :They have been used directly and indirectly (via semi-synthesis) as valuable drugs due to potent pharmacological activities, for example analgesic properties of codeine and morphine, the coronary vasodilator function of papaverine, the potentially anti-cancer drug and cough suppressant features of noscapine [1–3]. An overview on the reported separation approaches indicated that there are numerous appropriate techniques available for qualification and quantitative analysis of opium alkaloids such as thin-layer chromatography [4], capillary electrophoresis [5,6], gas chromatography [7] and high-performance liquid chromatography (HPLC), with a vast diversity of detection methods [8–10], particularly mass spectrometry (MS) [11–14], UV absorbance and/or fluorescence detections [8], and chemiluminescence [9]. Analysis of opium alkaloids is typically performed by applying reversed-phase high-performance liquid chromatography (RP-HPLC), accompanied by ion pairing reagents.