Development of chromatographic methods for thebaine detection and quantification along with some of related alkaloid derivatives

Marija Spasevska, Jane B Bogdanov, Hristina Babunovska


Thebaine is naturally occurring alkaloid  which is of great importance in the pharmaceutical industry and  up to now there is not a Pharmacopoeial monograph for this compound.  There is a definite need for development of  simple procedures for routine quality assessment  based on physical and spectroscopic properties (melting point, UV-Vis, IR, specific optical rotation) and even more importantly a need for development of chromatographic methods for determination of  purity.The main objective of this work was to develop RP-HPLC-UVD method for quantification of thebaine and also to develop complementary GC-MS method for qualitative analysis. The developed HPLC method utilized RP-C18 monolithic column, gradient elution with trifluoroacetic acid and formic acid in water and in acetonitrile and detection at 285nm. The effects of the relevant chromatographic conditions was investigated and the optimized and validated method was usеd for analysis of  three batches of thebaine.  

A procedure that contained different methods for determination and identification of thebaine, was performed with main focus on the HPLC method suitable for quantitative and qualitative determination.  In parallel, to complete the purity profile,  the three batches of thebaine were analyzed by IR, UV-Vis spectroscopy and specific optical rotation and melting point determination.  The analysis of thebaine can be performed in few steps by simple and reproducible methods like IR spectroscopy, specific optical rotation, melting point and the developed HPLC method, later proved by its validation. The developed HPLC-UV method is suitable for transfer to HPLC-MS method. The GC-MS method is quite suited for identification of thebaine  and of the impurities, but is limited to free bases. Additionally, pseduomorphine  and N-oxide derivatives of the related alkaloids are not suitable for GC-MS analysis.


thebaine, HPLC, purity, GC-MS, physical properties, spectroscopic properties

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