| 2-methoxyestradiol(2-ME) is a natural metabolite of main estrogen which wasdiscovered in the metabolism of estradiol in human urine in the1970s and has shownpotent inhibitory effects on proliferating cells and angiogenesis.In addition, it hassome advantages, such as high efficiency and low toxicity, less adverse reactions andnot easy to generate cross tolerance, which is studied at clinical phaseⅡabroadcurrently.2-ME is an anticancer drug with dependence of concentration and time,however, it has low solubility in water, be lack of correlation between the drugconcentration in plasma and the oral dose, strong first-pass metabolism and excretesat a high speed, which result in shorter biological half-life and lower oralbioavailability. Besides, it is difficult for the conventional dosage forms to meet thepharmacological characteristics dependence of concentration and time. Therefore,how to improve its oral absorption efficiency and bioavailability to main its effectivedrug concentration at the target site is worthy of studying. In this paper, we prepared2-ME phospholipid complex which was made of2-ME as the main drug and soybeanlecithin as the isotopic carrier, thus2-ME could integrate with phospholipid intophospholipid complex under certain condition. In view of amphipathy ofphospholipid, we hope that2-ME phospholipid complex prepared could overcome theshortcomings of2-ME in the aspect of biological pharmacy and finally meet therequirements of clinical oral dose.In the first place, a HPLC method in the study was established to determine thecontent of2-ME in its phospholipid complex.The optimum condition was as follows:Akasil C18column(5μm,4.6×250mm);mobile phase consisted of methanol andwater(70:30);flow rate1ml/min;column temperature30℃; UV detector at288nm.Theregression equation was A=0.2876C+0.1537(R2=0.9999), precision in days andbetween days were1.35%and2.02%, mean recovery was0.88%. The resultsindicated this method was fitted for the demands of sample analysis and detection. Atthe same time, we determined the solubility of2-ME, the results showed that2-MEwas insoluble in water (approximately0.225±0.016μg·ml-1), slightly soluble in dichloromethane and ether, easily soluble in tetrahydrofuran and methanol.Itsapparent oil-water partition coefficient was about3.61.Secondly,2-ME phospholipid complex was prepared by the solvent evaporationmethod using proportion as evaluation parameter, we investigated the formulation andpreparation techniques such as the reactive solvent, the concentration of reactants, theratio of lecithin to drugs, the reactive temperature and the reactive time. Finally theoptimized formulation and preparation techniques of preparing2-ME phospholipidcomplex were confirmed.The best process was as follows: the reactive solvent wastetrahydrofuran, the concentration of2-ME was5mg·ml-1, the molar ratio of lecithinto drugs was1:1.2,the temperature was controlled at40℃, the reactive time was threehours.Finally,2-ME phospholipid complex was prepared based on the abovecondition and its proportion was up to92.4%.In addition,the physicochemical properties of2-ME phospholipid complex wereevaluated. SEM and XRD showed that the complex exhibited an amorphouscharacteristic of soybean phospholipid and the crystal feature of2-ME disappearedcompletely. The characteristic endothermic peaks of2-ME also disappeared in theDSC spectrum of the complex.UV demonstrated that the chromophore of2-ME in thecomplex was unchanged. The results of IR and1H-NMR suggested that theinteraction between2-ME molecular and the polar end of soybean lecithin molecularcontributed to the formation of the phospholipid complex. Next,we investigated thesolubility of2-ME phospholipid complex in water and apparent partition coefficientsin the system of n-octyl alcohol and water.The outcome showed that both of themenhanced2.12and1.11times respectively.The release of2-ME,the physical mixtureand2-ME phospholipid complex in vitro reflected that2-ME phospholipid complexreleased faster than its physical mixture,2-ME released the slowest.The study on thestability of phospholipid complex showed that the complex should be becomeunstable under the condition of high temperature, strong illumination and highhumidity, hence, it should be stored under the condition of room temperature and dryair and dark condition.Furthermore, the effects of2-ME phospholipid complex on the MCF-7cellspharmacodynamics in vitro were studied by SRB. The data showed that2-ME phospholipid complex had obvious inhibition effect on MCF-7cells and thisinhibition effect could be heightened with time and concentration increment.Finally, the pharmacokinetic of2-ME phospholipid complex in big rats wasinvestigated in this paper.We established fluorescence HPLC with high sensitivity andgood selectivity to examine the2-ME blood concentration in the biologicalsamples.The rats were administrated by oral and got orbital blood and then detectedthe blood concentration.Compared to the AUC,Tmax,Cmaxand t1/2β (25.099±3.004μg·ml-1·min,8.872±0.886min,0.405±0.031μg·ml-1,73.693±4.579min),2-MEphospholipid complex were37.367±4.212μg·ml-1·min,8.865±0.187min,0.535±0.077μg·ml-1and121.912±10.812min. We could draw the conclusion that the oralbioavailability of2-ME phospholipid complex enhanced1.49times and its biologicalhalf-life extended50min, showing that2-ME phospholipid complex, a new kind ofdosage forms, could improve its oral absorption efficiency and bioavailability. |
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