Preparation Of In Situ Polylactic Acid Gel Implants Containing Medroxy Progesterone Acetate (mpa-pla) And Studies On The Anti-fertility Effect Of Mpa-pla In Mice

Several studies have shown that Medroxy Progesterone Acetate is progestogen,it is a derivate of the progesterone which can direct action at demic hypothalamic-pituitary-ovarian,accordingly to restrain demic follicular development and ovulation.Medroxy Progesterone Acetate enters blood and depot in adipose tissue,first of all,it influences endometrium,then secretory phase has changed,when the level of progesterone degrade and haemorrhagia uterine,endometrium is influenced again by Medroxy Progesterone Acetate from adipose tissue,change the proliferative phase to last contraception effect.Polylactic acid is biodegradable polymer which is used as the matrix to prepare prolonged action.Controlled release preparation can release the drug with permanent rate and prevent the untoward reaction which is caused by fluctuation of blood drug level with oral and injection.The development of new injectable drug delivery systems has received considerable attention over the past few years.This interest has been sparked by the advantages these delivery systems possess,which include ease of application,localized delivery for a site-specific action,prolonged delivery periods,decreased body drug dosage with concurrent reduction in possible undesirable side effects common to most forms of systemic delivery,and improved patient compliance and comfort. The development of a sustained release(SR) delivery system for MPA-PLA should therefore minimize the frequency of dosing,lengthen half-life,increases tability and improve the compliance.Now there are several kinds of formulations of contraceptives in our country,Such as injection,implant,transdermal patch,oral tablet,but we can not find any studies and reports about sustained release MPA-PLA.It is neeessary to explore a novel formulation for sustained release MPA-PLA,which will have enormous market prospects and good economic benefits.To estimate the sustained release efficiency of in situ gel implants containing medroxy progesterone acetate and investigate effection of reproduction rate in mice and detect the level of E₂ and LH in serum of rats.The results show that MPA-PLA could significantly prolong the anti-birth duration,suggest that MPA-PLA has long-term anti-fertility effect.Objectives:1.To study the preparation and releasing profiles in vitro of sustained-release MPA-PLA.To establish an HPLC method for the determination of in situ Gel Implants containing Medroxy Progesterone Acetate(MPA-PLA).2.To detect the level of E₂ and LH in serum of rats were assigned to in situ Gel Implants containing Medroxy Progesterone Acetate(MPA-PLA).3.To estimate the sustained release efficiency of in situ Gel Implants containing Medroxy Progesterone Acetate(MPA-PLA)and investigate effection of reproduction rate in miceMethods:1.Study on preparation of sustained-release MPA-PLA and its release in vitro:PLA was used as the matrix to prepare sustained- release MPA.Thein situ Gel Implants containing Medroxy Progesterone Acetate(MPA-PLA) was determined by HPLC method.The test conditions as follows:The content of MPA was determined by HPLC.The separation eolunm was VP-ODS.The mobile pase was methanol:water(70:30,V/V).The flow rate was 1.0mL·min^-1,the temperature of the column 30℃.and the dctective wavelength was 254nm.2.Anti-fertility effect of sustained- release MPA-PLA in rats.To evaluate the effect of MPA-PLA on sex hormone levels of female rats.35 SD rats were assigned to 7 groups,different doses of MPA were administered intramuscular(i.m.) injection to the adult female SD rats,respectively,the levels of luteinizing hormone(LH),estradiol(E₂) were determined by radioimmunoassay(RI).3.Anti-fertility effect of sustained- release MPA-PLA in mice.Seventy healthy female NIH mice were used to study the sustained-release of MPA-PLA;70 animals were randomly divided into 7 equal groups with 10 animals in each group.Mice of the trial group 1(n = 10 ) were given each 0mg·kg^-1 of MPA-PLA administered by intramuscular(i.m.). Mice of the trial groups 2～4 were given each 5mg·kg^-1,10mg·kg^-1 and 20 mg·kg^-1 of MPA-ethyl-alcohol administered by intramuscular(i.m.),respectively,Mice of the controll group 5～7 were given each 5mg·kg^-1,10mg·kg^-1 and 20 mg·kg^-1 of MPA-PLA in the same manner.Results:1.A HPLC method for determination of MPA in formulation was established.the linearity was in the range of 1.0ug·mL^-1～500.0ug·mL^-1.The relative standard deviations of intra-day and inter-day assay were 1.60%and 1.33%respectively.The average recovery was 98.30%,its relative standard deviation was 0.51%.The in vitro studies showed a controlled drug release over a period of 40d.2.In radio-immunity(RI) test of SD rats,compared to those in the control MPA-PLA 0 mg·kg^-1 group,the groups 2～7 markedly decreased in the rats administrated to MPA for 15 days.In addition, E₂ and LH levels in homogenates markedly decreased in the rats administrated MPA-Ethyl-alcohol to 10mg·kg^-1 and 20mg·kg^-1 groups for 15 days respectively.Serum levels of LH and E₂ decreased with the doses of MPA-PLA increased,with a significant dose-dependent decrease in E₂ levels,the MPA-PLA of 20 mg·kg^-1 group decreased significantly.LH and E₂ levels in serum also decreased with the doses of MPA increased,but not significantly.3.MPA-PLA,a chemical degraded from biomaterials,has been subjected to the tests of anti-fertility effect of mice.The studies demonstrated that PLA had fine biologieal compatibility without side effect issue.The results suggest that MPA-PLA injection has long-term anti-fertility effect.Reproduction of female mice in the laboratory was significantly prohibited by this contraceptive;the time of littering was delayed about 40 days longer and the proportion of littering females was reduced to 50%.The proportions of littering females and the numbers of littering of mice were also significantly reduced by this contraceptive within 1 months after animals were given MPA-PLA in laboratory.Conclusions:1.The HPLC way for determination of MPA-PLA was simple,reliable and provided a scientific basis for the quality controlling of MPA-PLA.which can be used to determination of MPA-PLA.The sustained- release MPA-PLA were successfully prepared.In vitro release studies showed a controlled drug release over a period of 40d..2.The sustained-release MPA-PLA showed good anti-fertility effect,can reduce the blood of E₂ and LH level.MPA-PLA could significantly inhibit the production of progesterone on female rats, and can alter E₂ and LH levels in serum.3.MPA-PLA not only can prolong the anti-birth duration,but also decrease the dose and side-effects.Furthermore,the MPA-PLA do not interfere with the sexual behavior and the reproductive function in mice.MPA-PLA was shown to be an effective anti-fertility drug and its contraceptive effect is reversible.