Department of Experimental and Clinical Pharmacology, University of Rzeszów, Kopisto 2a, 35-959 Rzeszów, Poland.
Department of Inorganic Chemistry, Medical University of Lublin, Chodźki 4a, 20-093 Lublin, Poland.
Molecules. 2019 Jul 16;24(14):2580. doi: 10.3390/molecules24142580.
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. 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. 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). 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.
识别和定量测定野靛碱，特别是在生物样品和药物制剂中，仍然是一项具有挑战性的分析任务。野靛碱是一种具有小而非常极性分子的生物碱。由于这个原因，它在反相（RP）固定相上的保留非常弱，例如常用的烷基键合相。野靛碱的非常弱保留导致它与生物基质的成分一起洗脱。 通过高效液相色谱（HPLC）与二极管阵列（DAD）、荧光（FLD）和质谱（MS）检测，比较和评估各种色谱系统在不同基质（血清、唾液和药物制剂）中分析野靛碱的性能。 分析采用反相（RP）、亲水相互作用液相色谱（HILIC）和离子交换色谱（IEC）模式进行。测试了不同的样品预处理方法：蛋白质沉淀（用丙酮、甲醇（MeOH）或乙腈（ACN），以及使用十八烷基（C18）、亲水-疏水平衡共聚物（HLB）或强阳离子交换吸附剂（Strata X-C）的固相萃取（SPE）试剂盒。 使用不同的色谱系统时，观察到保留参数有显著差异。固定相的各种性质导致分析物保留、峰形和系统效率的差异。在 RP 系统中观察到的保留最弱；然而，极性 RP 相的使用可以作为绿色色谱应用的替代方法。在使用强阳离子交换（SCX）相时观察到最强的保留。在经过新的 SPE 处理程序对药物制剂、血清和唾液进行样品预处理后，选择了最优化的系统用于分析野靛碱。由于灵敏度高，HPLC-DAD 或 HPLC-FLD 用于药物制剂中的药物分析是最优化的，而 HPLC-MS 适合于生物样品中野靛碱的分析。
