Abstract
Background: Identification and quantitative determination of cytisine, especially in biological samples and pharmaceutical formulations, is still a difficult analytical task. Cytisine is an alkaloid with a small and very polar molecule. For this reason, it is very weakly retained on reversed phase (RP) stationary phases, such as commonly used alkyl-bonded phases. The very weak retention of cytisine causes it to be eluted together with the components of biological matrices. Objective: Comparison and evaluation of various chromatographic systems for analysis of cytisine in different matrices—serum, saliva and pharmaceutical formulation—by high performance liquid chromatography (HPLC) with diode array (DAD), fluorescence (FLD) and mass spectrometry (MS) detection. Methods: The analyses were performed using HPLC in reversed phase (RP), hydrophilic interaction liquid chromatography (HILIC) and ion exchange chromatography (IEC) modes. Different sample pre-treatment methods were tested: Protein precipitation (with acetone, methanol (MeOH) or acetonitrile (ACN), and solid phase extraction (SPE) using cartridges with octadecyl (C18), hydrophilic-lipophilic balanced copolymer (HLB) or strong cation exchange sorbents (Strata X-C). Conclusion: Significant differences were observed in retention parameters with a change of the used chromatographic system. The various properties of stationary phases resulted in differences in analyte retention, peaks’ shape and systems’ efficiency. The weakest retention was observed using RP systems; however, the use of the Polar RP phase can be an alternative for application in green chromatography. In the strongest retention was observed using a strong cation exchange (SCX) phase. The most optimal systems were chosen for the analysis of cytisine in the pharmaceutical preparation, serum and saliva after sample pre-treatment with the new SPE procedure. Due to the sensitivity, the use of HPLC-DAD or HPLC-FLD is the most optimal for drug analysis in pharmaceutical preparations, whereas HPLC-MS is suitable for analysis of cytisine in biological samples.
Highlights
Cytisine is a quinolizidine alkaloid originating from plants of the Leguminosae (Fabaceae) family.The Cytisine greatest is amount of the alkaloid was found in from the seeds of of Laburnum anagyroides (GoldenRain a quinolizidine alkaloid originating plants the Leguminosae (Fabaceae) family. acacia; about 1%–5%) [1].Plants containing cytisine have been used as a natural remedy andThe greatest amount of the alkaloid was found in the seeds of Laburnum anagyroides (Golden Raina medicinal agent for[1].various forcytisine centuries [2,3]. 1–5%)Plantspurposes containing have been used as a natural remedy and a medicinal
The intensity of the signal was comparable to that obtained with the DAD detection at the maximum wavelength at about 303 nm (Figure 2)
Chromatographic systems for analysis of cytisine were optimised with the application of a large number of various stationary phases and mobile phases of different compositions
Summary
Cytisine is a quinolizidine alkaloid originating from plants of the Leguminosae (Fabaceae) family. The Cytisine greatest is amount of the alkaloid was found in from the seeds of of Laburnum anagyroides Rain a quinolizidine alkaloid originating plants the Leguminosae (Fabaceae) family. The greatest amount of the alkaloid was found in the seeds of Laburnum anagyroides Cytisine is an alkaloid with a small and very polar molecule For this reason, it is very weakly retained on reversed phase (RP). Methods: The analyses were performed using HPLC in reversed phase (RP), hydrophilic interaction liquid chromatography (HILIC) and ion exchange chromatography (IEC) modes. The various properties of stationary phases resulted in differences in analyte retention, peaks’ shape and systems’
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