鹿藿正丁醇提取物体外杀精作用及机制探讨

A study on spermicidal effect in vitro and the mechanism of n-Butanol extract of Rhynchosia volubilis Lour

目的研究鹿藿正丁醇提取物（11-ButanolextractofRhynchosiavolubilisLour，BERVL）的体外杀精子作用及机制。同时观察该提取物体外对嗜酸乳酸杆菌的影响，初步评估BERVL作为杀精子剂的可行性和安全性。方法体外杀精子实验以生理盐水为阴性对照，壬苯醇醚（nonoxynolum-9，N-9）为阳性对照，显微镜下观察浓度为250mg／ml，125mg／ml，100mg／ml，90ms／ml，62．5mg／ml的BERVL对人精子活力的影响；体外抑茵实验用试管稀释法和琼脂稀释法；体外杀精子机制探讨：用伊红Y水试验和Hoechst33342／PI荧光双染，显微镜下检测精子膜功能和精子凋亡。结果BERVL在90mg／ml浓度组：体外杀精实验，原地活动精子百分率为（4．730±3．231）％；体外抑菌试验，对嗜酸乳酸杆菌抑制率为32．96％-35．70％；伊红Y水溶液染色，精子头部未着色百分率为（7．846±4．706）％。Hoechst33342／PI荧光双染．凋亡精子百分率为（4．977±2．267）％，死精子百分率为（90．111±4．509）％。结论90mg／ml为BERVL较安全理想的体外杀精浓度。BERVL有显著的体外杀精子作用，可能与破坏精子膜和加速精子凋亡及死亡有关。

Objective To explore spermicidal effect in vitro of n - Butanol Rhynchosia volubilis Lour （BERVL） and its mechanism as well as its influence to lactobacillus acidophilus. Methods Spermicidal experiments were classified nonoxynolum- 9 （N- 9）as positive control group, O. 9% sodium chloride （Nacl） as negative control group and BERVL group. Based on different spermicida[ concentrations of BERVL group ,they were divided into 250 mg/ml, 125 mg/ml, 100 mg/ml,90 mg/ml and 62. 5 mg/ml. After BERVL was mixed with sperm and continued for two minutes,the moving sperm was calculated under microscope. Bacteriostasis tests in vitro were studied by Tubes and AGAR dilution methods. To probe into Spermicidal mechanism, sperm membrane and sperm apoptosis under the microscope by Eosin Y and Hoechst 33342/PI stain were observed. Results After 90 mg/ml of the BERVL was mixed with sperm,there was no moving forward sperm, only （4. 730 ＋ 3.231 ） % of sperm was moving around. In the baeteriostasis experiment, the rate of inhibitory effect to laetobacillus acidophilus was 35.7 % or 32. 96%. After Eosin Y and Hoechst33342/PI stain, （7. 846 ＋_ 4. 706）% of the sperm head was not stained and the sperm apoptosis rate was （4. 977 -＋ 2. 267 ） % as well as the necrosis sperm was （90. 111 ＋ 4. 509 ） %. Conclusion 90 mg/ml of BERVL is the minimal spermicidal.and safe dose in vitro. BERVL had obvious spermicidal function in vitro, and the possible mechanism maybe related to destroy sperm membrane and accelerate sperm apoptosis and necrosis.