Studies On Estradiol Nasal Spray

Estradiol (E₂) is a nature sex steroids hormone, used to improve the symptom of menopause, reduce the risk of bone fractures occurring after menopause, prevent and cure femme ovarian dysfunction primarily or after ovariectomy. The most common forms of E₂ supplementation currently used are the oral, transdermal and provagina. However, the three methods have several limitations, respectively. In order to avoid the first pass effect and elevate the bioavailability of E₂, and improve the compliance of the patients, the E₂ nasal spray was prepared.The HPLC method was established to determine the E₂ concentration in E₂ nasal spray and the solubility in different medium. The character of E₂ was investigated, too.E₂ is a lipid soluble drug which is micromolecular, if wanted to nasal administration, its solubility in water must be increased, so the randomly methylatedβ-cyclodextrin (RAMEB) complex was prepared to develop the solubility in water. The preparation method was studied with factor investigation, based on which, the solubility of E₂ in water and entraption ratio were taken as indexes to optimize the preparation technology with orthogonal design experiment. Finally determined the optimal technology. The complex prepared by this technology had the better solubility in water and better physical stability. The complex was identified by DSC and X- ray diffraction method. Rat in situ nasal perfusion methods was used as an experimental model to investigate the regulation of nasal absorption. The effects of perfusion volume, flow rate, and perfusion concentration on nasal absorption of E₂ were studied. The results were that the larger the perfusion volume was, the less of the percent absorbed was each minute. The faster the flow rate was, the more of the percent absorbed each minute. The mechanism of E₂ nasal absorption is active transport, following zero-order kinetics.By rat in situ nasal perfusion methods, the optimal amount of chitosan bolted was 0.5 %; and the influence of the formulation pH on the E₂ nasal absorption was investigated, the results indicated that in the pH range of 4.0-6.0, there was no influence on the E₂ nasal absorption.After Making sure the stable pH range of the E₂ nasal spray, the influential factor study and initial stability investigation were carried out, the results indicated the stable pH range was 4.0～6.0; the formulation was stable to heat and light; and had good stability in accelerated test within 3 months and long-term place test within 4 months.To assess the nasal ciliary toxicity of E₂ nasal spray, the hoptoad in situ maxilla mucosa method was used by determining the ciliary movement time after in. administration. The toxicity is very little and the preparation can be used in nasal delivery system. Based on the researches above, the dosage form of E₂ nasal spray were: the complex was prepared with E₂ and RAMEB in 1:2(mol ratio), the dosage of chitosan and benzalkonium chloride was 0.5% and 0.01%, respectively, sodium chloride was used to adjust iso-osmia, pH range was 4.0～6.0.HPLC fluorometric method was performed to detect the concentration of E₂ in rats plasma, which was simple and precise. Physical solution of the E₂-RAMEB combination, and 70% ethanol solution of E₂ and E₂ nasal spray in 3 different doses (0.16,0.48,1.60 mg·Kg^-1) had been nasally administrated to rats, respectively. Non-compartment statistical moment method had been used in pharmacokinetics study. Absolute bioavailability degree had been compared with E₂ i.v. T_max, C_max values and absolute bioavailability degree of groupâ… (E₂ nasal spray 0.16,0.48,1.60 mg·Kg^-1),â…¡(Physical solution of the E₂-RAMEB combination),â…¢(70% ethanol solution of E₂) are 30, 25, 13, 20 and 5 min; 72.41, 74.72, 259.21, 20.51 and 60.65 ng·ml^-1; 86.87%, 65.83%, 48.99%, 56.44% and 24.38%, respectively. Absolute bioavailability degree of both 0.16å’Œ0.48 mg.kg^-1 administration in groupâ… are higher than that in groupâ…¡, which showed nasal spray enhanced medicine absorption compared with combination solution.HPLC fluorometric method was performed to detect the concentration of E₂ in rats CSF, which was simple and precise. E₂ concentration in CSF and plasma were examined, probe recovery was 61.64% by concentration difference method. Non-compartment statistical moment method had been used in pharmacokinetics study. Absolute bioavailability degree of test preparation had been compared with E₂ i.v., T_max, C_max and AUC values in CSF of i.n. and i.v. administration were 20 and 60min; 37.05 and 17.91 ng·ml^-1; 4445.74 and 3228.48 ng·min·ml^-1, respectively. AUC_CSF/AUC_plasma value were 1.08 and 0.41, the results of t-test indicated the data was significant difference between the i.n. and i.v. administration(P＜0.05). It was clear that E₂ could directly enter brain through olfactory channel because of the shorter of T_max, it implement with two pathway, this conclusion was conformity with the research before. The site of action and recipient of female hormone were in the central nervous system, this research supplied the evidence of E₂ nasally administration to cure Alzheimer.