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  本文選題：2-甲氧基雌二醇 + 磷脂復(fù)合物��； 參考：《鄭州大學(xué)》2014年碩士論文

【摘要】：2-甲氧基雌二醇（2-methoxyestradiol，簡稱2-ME）是20世紀(jì)70年代，在研究雌二醇體內(nèi)代謝過程時，尿液中發(fā)現(xiàn)的主要雌激素的天然代謝產(chǎn)物，已研究證實，該化合物對細(xì)胞增殖和血管新生具有潛在抑制作用，且具有高效低毒、不良反應(yīng)少、不易產(chǎn)生交叉耐藥性等優(yōu)點，目前國外正處于Ⅱ期臨床研究階段。2-ME作為一種濃度時間依賴性的新型廣譜抗腫瘤藥，但由于它難溶于水，口服血藥濃度和劑量之間缺乏相關(guān)性，肝首過效應(yīng)強，，代謝速率快，導(dǎo)致體內(nèi)生物半衰期短和口服生物利用度低，而且常規(guī)給藥劑型很難滿足其濃度時間依賴性藥理學(xué)特征。因此，如何提高2-ME的口服吸收效率，提高生物利用度，以維持其有效的血藥濃度是臨床應(yīng)用的關(guān)鍵。本研究是欲以2-ME為主藥，大豆卵磷脂為載體，利用二者相互作用形成磷脂復(fù)合物，希望該磷脂復(fù)合物能夠解決2-ME生物藥劑學(xué)方面存在的不足，以達到臨床口服給藥目的。 本研究首先建立了高效液相色譜測定2-ME磷脂復(fù)合物中2-ME含量的分析方法，選用Akasil C18色譜柱，甲醇：水(70:30)為流動相，流速為1ml/min，柱溫為30℃，紫外檢測波長為288nm。該方法的線性回歸方程A=0.2876C+0.1537(R2=0.9999)，日內(nèi)和日間精密度分別為1.35%和2.02%，平均回收率達到0.88%，符合樣品分析要求。2-ME溶解度測定結(jié)果顯示：2-ME難溶于水(約0.225±0.016μg·ml-1)，略溶于二氯甲烷和乙醚，易溶于四氫呋喃和甲醇等有機溶劑，表觀油水分配系數(shù)：log P=3.61。 其次采用溶劑揮發(fā)法制備了2-ME磷脂復(fù)合物，以2-ME與大豆卵磷脂的復(fù)合率為評價指標(biāo)，綜合考察了處方因素(反應(yīng)溶劑、反應(yīng)藥物濃度、藥物與磷脂投料比)及工藝因素(反應(yīng)溫度、反應(yīng)時間)對復(fù)合率的影響，最終確定了最優(yōu)處方及工藝為：以四氫呋喃為反應(yīng)溶劑，2-ME質(zhì)量濃度為5mg·ml-1，2-ME與磷脂投料摩爾比為1:1.2，反應(yīng)溫度40℃，反應(yīng)時間3h，制得復(fù)合率可達92.4%的淡黃色黏性固體物。 再次對2-ME磷脂復(fù)合物進行表征，SEM和XRD法分析結(jié)果顯示2-ME晶體特征消失，磷脂復(fù)合物表現(xiàn)出無定型態(tài)；DSC法測定顯示復(fù)合物中2-ME熔點峰消失；UV法測定顯示復(fù)合物中2-ME的發(fā)色基團未發(fā)生改變；IR和1H-NMR表征結(jié)果顯示2-ME磷脂復(fù)合物是磷脂分子的極性端與2-ME分子之間發(fā)生相互作用的結(jié)果。復(fù)合物水中溶解度(0.477±0.021μg·ml-1)和表觀油水分配系數(shù)(log P4.02)分別提高了2.12倍和1.11倍。槳法考察了復(fù)合物體外溶出度，結(jié)果顯示2-ME磷脂復(fù)合物累積溶出度高于其物理混合物，2-ME原料藥溶出最慢。初步穩(wěn)定性試驗考察結(jié)果顯示2-ME磷脂復(fù)合物在高溫、光照和濕度條件下穩(wěn)定性會降低，應(yīng)當(dāng)貯藏在低溫干燥避光處。 此外，以MCF-7細(xì)胞株為研究對象，SRB法評價了2-ME磷脂復(fù)合物的體外細(xì)胞生長抑制作用，結(jié)果顯示2-ME磷脂復(fù)合物對MCF-7細(xì)胞的增殖抑制作用呈時間濃度依賴性。2-ME磷脂復(fù)合物在48h和72h的IC50值分別為3.26μmol·l-1和0.98μmol·l-1，而2-ME原料藥的為5.28μmol·l-1和2.72μmol·l-1。 最后以大鼠為給藥模型，建立了熒光高效液相色譜測定復(fù)合物生物樣品中2-ME濃度的分析方法。大鼠灌胃給藥，體內(nèi)藥代動力學(xué)測定結(jié)果顯示：2-ME磷脂復(fù)合物的AUC、Tmax、Cmax和t1/2β分別為(37.367±4.212) μg·ml-1·min、(8.865±0.187)min、(0.535±0.077)μg·ml-1、(121.912±10.812)min，相對應(yīng)的2-ME原料藥AUC、Tmax、Cmax和t1/2β依次為(25.099±3.004) μg·ml-1·min、(8.872±0.886) min、(0.405±0.031)μg·ml-1、(73.693±4.579)min，2-ME磷脂復(fù)合物的口服生物利用度約是2-ME的1.49倍，生物半衰期延長近50min，說明將2-ME制成磷脂復(fù)合物可提高其口服吸收效率。
[Abstract]:2- methoxy estradiol (2-methoxyestradiol, 2-ME) is a natural metabolite of the main estrogen found in the urine during the metabolic process of estradiol in 1970s. It has been confirmed that the compound has a potential inhibition effect on cell proliferation and angiogenesis, with high toxicity and low toxicity, and is not easy to produce. At present,.2-ME is a new type of broad-spectrum antitumor drug with time dependence in phase II clinical study. However, because it is difficult to dissolve in water, there is a lack of correlation between the concentration and dose of oral blood, strong head over effect and fast metabolic rate, resulting in short biological half-life in the body and oral bioavailability. It is low, and it is difficult to meet the time dependent pharmacological characteristics of the conventional drug type. Therefore, how to improve the oral absorption efficiency of 2-ME, improve the bioavailability and maintain its effective blood concentration is the key to the clinical application. This study is intended to use 2-ME as the main drug, soybean lecithin as the carrier, and use the interaction of the two to form phospholipids. It is hoped that the phospholipid complex can solve the deficiency of 2-ME Biopharmaceutics, so as to achieve the goal of oral administration.
In this study, a high performance liquid chromatography (HPLC) method for the determination of 2-ME in 2-ME phospholipid complex was established by using Akasil C18 chromatographic column, methanol: water (70:30) as the mobile phase, the flow velocity of 1ml/min, the column temperature of 30 C, and the linear regression square A=0.2876C+0.1537 (R2=0.9999) of the UV detection wavelength for 288nm., and the intra day and day precision points. The average recovery rate of 1.35% and 2.02% is 0.88%. The results show that the solubility of.2-ME shows that 2-ME is difficult to dissolve in water (about 0.225 + 0.016 mu g. ML-1), slightly dissolved in dichloromethane and ether, easily dissolved in organic solvents such as tetrahydrofuran and methanol, and the apparent oil water distribution coefficient: log P=3.61.
Secondly, 2-ME phosphatide complex was prepared by solvent evaporation, and the compound ratio of 2-ME and soybean lecithin was used as evaluation index. The effect of prescription factors (reaction solvent, reaction drug concentration, drug and phospholipid ratio) and process factors (reaction temperature and reaction time) on the compound rate were investigated, and the optimal formulation and process were finally determined. With tetrahydrofuran as the reaction solvent, the mass concentration of 2-ME is 5mg / ml-1,2-ME and the molar ratio of phospholipid is 1:1.2, the reaction temperature is 40 and the reaction time is 3h, and the light yellow sticky solid with the compound rate of up to 92.4% is obtained.
The 2-ME phospholipid complex was characterized again. The results of SEM and XRD analysis showed that the characteristics of 2-ME crystal disappeared, the phospholipid complex showed the amorphous state, and the DSC method showed that the peak of 2-ME melting point in the compound disappeared, and the UV method showed that the chromophore group of 2-ME in the complex was not changed; IR and 1H-NMR characterization showed 2-ME phospholipid complex. It is the result of interaction between the polar end of phospholipid molecule and 2-ME molecule. The solubility of the composite water (0.477 + 0.021 G. ML-1) and the apparent oil and water distribution coefficient (log P4.02) increased by 2.12 times and 1.11 times respectively. The paddle method examined the exsolution degree of the compound object, and the results showed that the cumulative dissolution of the 2-ME phospholipid complex was higher than that of its physical mixture. The initial stability test results show that the stability of the 2-ME phospholipid complex will decrease under the conditions of high temperature, light and humidity, and should be stored at the low temperature drying and light avoiding place. The preliminary stability test results show that the stability of 2-ME phospholipid complex will decrease.
In addition, the inhibitory effect of 2-ME phospholipid complex on the proliferation of 2-ME phospholipid complex was evaluated by SRB method. The results showed that the inhibitory effect of 2-ME phospholipid complex on the proliferation of MCF-7 cells was time dependent.2-ME phospholipid complex at 48h and 72h in IC50 value of 3.26 Mu mol. L-1 and 0.98 micron mol. 5.28 mol. L-1 and 2.72 Mu mol. L-1.
At last, a method for the determination of the concentration of 2-ME in the biological samples of the compound was established with the rat as the drug delivery model. The rats were administered by GC and the pharmacokinetics of 2-ME phospholipid complex showed that the AUC, Tmax, Cmax and t1/2 beta of the phospholipid complex were (37.367 + 4.212) mu g. Ml-1. Min, (8.865 + 0.187) min, (0.535 + 0.077) micron. G. Ml-1, (121.912 + 10.812) min, the corresponding 2-ME AUC, Tmax, Cmax and t1/2 beta are (25.099 + 3.004) mu g ml-1 min, (8.872 + 0.886) min, (0.405 + 0.031) micron, (73.693 + 4.579), and the bioavailability of the phospholipid complex is about 1.49 times that of that, and the biological half-life is prolonged, indicating that the phospholipid will be made into phospholipid. The compound can improve its oral absorption efficiency.

【學(xué)位授予單位】：鄭州大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R943

【共引文獻】

相關(guān)期刊論文 前1條

1 馬偉峰;胡海英;連曉培;張正全;張振中;;2-甲氧基雌二醇的大鼠在體胃腸吸收特性[J];鄭州大學(xué)學(xué)報(醫(yī)學(xué)版);2011年02期

本文編號：1804186