Application Of Steroid Hormone Drug Analysis Method

Steroid hormone drugs refer to the hormone drugs containing steroidal structure in molecular structure. Nowadays steroid hormone drugs have been widely used in clinic medicine because of their many physiological activities, and have become an important category of medicine. Moreover the quantity of the drugs directly affects people’s life health and safety. Therefore it is necessary to comprehensively monitor drug quality in the clinic and medical treatment. The goals of steroid hormone drug analysis include (1) To monitor the quality of the drug itself in raw material or in pharmaceutical formulation with accurate and reliable method; (2) To determine drugs and their metabolites in biological samples with sensitive and accurate technique; (3) To study the interaction of drugs and protein or amino acid. In order to meeting the sensitivity and accuracy of analysis requests, some sensibilization or preconcentration/seperation techniques are usually adopted in pharmaceutical analysis.The aim of this work is to explore simpler and more sensitive analysis methods of steroid hormone drugs, with using spectrophotometry and high performance liquid chromatography combining with some ordered molecular assembly sensitizing agents and liquid phase extraction techniques as research means. The relevant sensitization or extraction mechanism was preliminarily discussed. The major contents are summarized as follows:1. β-cylodextrin sensitized spectrofluorimetry for the determination of abiraterone acetate and abirateroneA novel spectrofluorimetric method to determine androgen abiraterone acetate and its active metabolite, abiraterone was developed, based on the fact that fluorescence intensity of abiraterone acetate and abiraterone could be enhanced in ordered molecular assembly (3-cyclodextrin (β-CD) due to the formation of the inclusion complex. The inclusion interaction of β-CD and abiraterone acetate and the β-cyclodextrin sensitized spectrofluorimetry was examined. The various factors influencing fluorescence were discussed in details. The results showed that under the optimized conditions, the linear range of calibration curve for the determination of biraterone acetate and abiraterone was 0.20-6.0 μg/mL, and the detection limit (LOD) was 6.8 (r=0.997) or 6.6 ng/mL (r=0.996), respectively. No interference was observed from common co-existing substances or pharmaceutical excipient. The method was successfully applied to the analysis of abiraterone acetate in pharmaceutical formulation and abiraterone in human serum/urine.2. Surfactant/ionic liquid aqueous two-phase system extraction coupled with spectrofluorimetry for the separation/analysis of dutasteride in pharmaceutical formulation and biological samplesA novel separation/analysis method was developed for the determination of androgen dutasteride in a pharmaceutical formulation and biological samples, which based on ordered molecular assembly surfactant dodecyl sodium sulfate (SDS)/ionic liquid 1-hexyl-3-methylimidazolium bromide (IL, [C6min]Br) aqueous two-phase system (ATPS) extraction coupled with spectrofluorimetry. The ATPS was formed owing to new ordered molecular assembly SDS/IL, which not only extracted dutasteride but also sensitized its fluorescence intensity due to synergistic sensitizing effect. Under optimum conditions, a linear calibration curve was obtained in the range of 0.016-0.64 μg/mL with r= 0.999 (n=8). The detection limit was 2.2 ng/mL. The preconcentration factor was 7.5. The proposed green analytical procedure was satisfactorily applied to the analysis of dutasteride in dutasteride soft gelatin capsule and human serum with the recovery of 96.7%-108.0%. The obtained results of this work were in good agreement with the results of HPLC. The extraction mechanism ATPS-based was also discussed.3. Dispersive solvent-free ultrasound-assisted ionic liquid dispersive liquid-liquid microextraction coupled with HPLC for the separation/analysis of ulipristal acetateIn this paper, a simple and efficient ultrasound-assisted ionic liquid dispersive liquid-liquid microextraction (UA IL-DLLME) coupled with high-performance liquid chromatography for the analysis of progesterone ulipristal acetate (UPA) was developed. UPA could be easily migrated into ordered molecular assembly l-octyl-3-methylimidazolium hexafluorophosphate [C8mimPF6] IL phase without dispersive solvent. Several important parameters affecting the extraction recovery were optimized. Under the optimized conditions,25-fold enrichment factor was obtained and the limit of detection (LOD) was 6.8 ng/mL (tablet) or 9.3 ng/mL (serum) at a signal-to-noise ratio of 3. The calibration curve was linear over the range of 0.03-6.0 μg/mL. The proposed method was successfully applied to the real mice serum samples and UPA tablets with good spiked recoveries in the range of 103.5%-109.7%. The research of extraction mechanism showed that hydrophobic interaction force played a key role in the extraction of UPA.4. Miniaturized ionic liquid dispersive liquid-liquid microextraction in a coupled-syringe system combined with UV for the separation/analysis of danazol in danazol capsule and mice serumIn this study, for the first time, a coupled 1-mL syringe system was utilized to perform a miniaturized ionic liquid dispersive liquid-liquid microextraction method. Androgen danazol was extracted and determined via the developed method followed by micro volume ultraviolet spectroscopy (UV). The extraction process was carried out by the injection of extraction solvent ionic liquid 1-octyl-3-methylimidazolium hexafluorophosphate [C8mimPF6] into sample solution (syringe A), and then rapid shoot the solution into syringe B. After that the shooting was repeated several times at a rate of 1 cycle/s. The extraction procedure was induced by the formation of cloudy solution, which was composed of fine drops of ordered molecular assembly [C8mimPF6] dispersed entirely into sample solution with the help of shooting without any dispersive solvent, ultrasonication or high temperature, and which promoted the analytes more easily migrate into the ionic liquid phase. Several important parameters affecting the extraction efficiency were studied and optimized, including the type and volume of extraction solvent, sample pH, shooting times and salt concentration. Under the optimized conditions, the limit of detection (LOD) was 0.055 μg/mL (capsule) or 0.054 μg/mL (serum) at a signal-to-noise ratio of 3. The calibration curve was linear over the range of 0.62-25.0 μg/mL. The proposed method was successfully applied to the real mice serum samples and danazol capsule and good spiked recoveries in the range of 90.5-103.4% were obtained. The obtained results of this work were in good agreement with the results of HPLC.5. Ultrasound-assisted emulsification of solidified floating organic drop micro extraction (USE-SFODME) coupled to UV spectrometry for the separation/analysis of epristeride in tablet and biological fluidsA simple, sensitive, and effective method of ultrasound-assisted emulsification of solidified floating organic drop microextraction (USE-SFODME) coupled to micro volume UV spectroscopy for the determination of androgen epristeride (EP) in tablet and human serum was established. The extraction procedure was induced by the emulsification of ordered molecular assembly 1-undecanol, which was composed of fine drops of 1-undecanol dispersed entirely into sample solution with the help of ultrasonication and EP can be extracted into 1-undecanol. The solidification of 1-undecanol after extraction the solution was cooled at 5℃ made it was easy to separate from aqueous solution. The extraction efficiency was over 90.0% and 20-fold enrichment factor was obtained. In examination it was also found that the fluorescence intensity of L-tryptophan could be quenched by EP, and fluorescence quenching value (A F=FL. tryptophan-FEP-L-tryptophan) showed a good linear relationship with adequate EP concentration. Hence in this study not only 1-undecanol extraction spectrophotometric method but also a novel spectrofluorimetric method was developed to determine EP. The two developed methods were compared and simultaneously successfully applied to the analysis of EP in EP tablets and biological fluids. The obtained results were in good agreement with the results of the official method (acid-base titration).