四种中药有效成分对D-半乳糖诱导大鼠醛糖还原酶活性和蛋白非酶糖化的作用

Effect of active component of four types of Chinese herbs on aldose reductase activity and non-enzymatic protein glycation in D-galactose induced rats

目的:观察葛根素、小檗碱、黄芩苷和甘草酸对D-半乳糖诱导大鼠体内醛糖还原酶活性、糖耐量和蛋白非酶糖基化反应的作用,并与可拮抗糖基化反应的阳性对照药氨基胍做比较。方法:实验于2005-03/07在暨南大学药学院药理教研室动物室完成。①选用6周龄清洁级SD大鼠105只。取15只作为正常对照组:腹腔注射生理盐水,1次/d,持续8周。其余90只动物均腹腔注射150mg/kgD-半乳糖,1次/d,持续8周。于第3周,按大鼠性别和体质量均衡随机分为6组(每组15只):模型组、氨基胍组、葛根素组、小檗碱组、黄芩苷组和甘草酸组,各组分别灌胃氨基胍(Sigma公司产品)、葛根素(购自北京安琪药业,批号:030112)、小檗碱(广东万基药业有限公司,批号:20040704)、黄芩苷(广东惠州中药厂,批号:20040521)和甘草酸(江苏省天晟药业有限公司,批号:20040803)300mg/(kg·d),10mL;同时模型组和正常对照组动物均灌胃等量蒸馏水,1次/d,持续6周。②上述动物于第8周末,用血糖仪测定空腹血糖和口服葡萄糖耐量试验后2h血糖。用荧光分光光度计和马斯亮蓝法测定血浆晚期糖基化终末产物水平。采用荧光法测定红细胞醛糖还原酶活性。采用硫代巴比妥酸比色法,按试剂盒说明书测定血浆糖化血红蛋白含量。采用硝基四氮唑蓝比色法,按试剂盒说明书测定血浆果糖胺含量。③计量资料分别采用配对t检验和组间t检验。结果:大鼠105只均进入结果分析。①D-半乳糖处理8周后,各组大鼠空腹血糖未见明显变化(P>0.05);模型组大鼠口服葡萄糖耐量试验后2h血糖及空腹血糖与口服葡萄糖耐量试验后2h血糖差值明显高于正常对照组(P<0.01),而氨基胍组、葛根素组、小檗碱组、黄芩苷组明显低于模型组(P<0.05～0.01);甘草酸组也低于模型组,但差异不明显(P>0.05)。②D-半乳糖处理8周后,模型组红细胞醛糖还原酶活性明显高于其他6组(P<0.05～0.01),果糖胺、晚期糖基化终末产物含量明显高于正常对照组、氨基胍组、葛根素组、小檗碱组(P<0.01),糖化血红蛋白含量明显高于正常对照组、氨基胍组、葛根素组、小檗碱组、黄芩苷组(P<0.05～0.01)。结论:氨基胍、葛根素、小檗碱、黄芩苷和甘草酸均能抑制D-半乳糖诱导大鼠体内醛糖还原酶活性,氨基胍、葛根素与小檗碱对D-半乳糖诱致的大鼠糖基化反应具有明显的抑制作用,而黄芩苷对大鼠糖基化反应具有部分抑制作用,甘草酸对大鼠糖基化反应未见抑制作用。

AIM： To observe the effect of puerarin （Pue）, berberine （Ber）, baicalin （Bai） and glycyrrhizic acid （Gly） on the activity of aldose reductase, glucose tolerance and products of protein non-enzymatic glyeation induced by Dgalactose （D-gal） in rats. METHODS： The experiment was conducted in the animal laboratory of Teaching and Research Section of Pharmacology, College of Pharmacy, Jinan University from March to July 2005. ①105 clean grade SD rats aged 6 months were selected. Fifteen rats were selected as the control group, which were intraperitoneally injected with normal saline, once daily for 8 weeks. The rest 90 rats were given intraperitoneal injection of 150 mg/kg D-gal, once daily for 8 weeks. At the 3rd week, the D-gal-treated rats were randomly divided into 6 groups with 15 rats in each group： model control, aminoguanidine （AG）, Pue, Ber, Bai and Gly groups, which were intragstrointestinally infused AG （products of Sigma Company）, Pue （purchased from the Beijng Anqi Pharmaceutical Co., Ltd., No. 030112）, Ber （Wanji Pharmaceutical Co., Ltd., No. 20040704）, Bai （Guangdong Huizhou Chinese Medicine Business, No. 20040521） and Gly （Jiangsu Tianeheng Pharmaceutical Co., Ltd., No. 20040803） with 300 mg/kg daily, 10 mL, respectively; meanwhile, the rats in the model and control groups were intragstrointestinally infused with the same dose of distilled water, once daily for 6 weeks. ②At the end of the 8th week, the fasting blood glucose and the glucose 2 hours after the orally taken glucose tolerance test were measured by the Blood Glucose Meter;, end products of protein glyeation by spectrophotofluorometer and coomassie brilliant blue method; activity of aldose reductase in red blood cells by fluorometrie method; plasm glueosylated hemoglobin by the thio-malonylurea eolormetrie method according to the kit; and the plasm fructosamine by the nitroblue tetrazolium colormetric method according to the kit. ③The pairing t-test and t-test between groups were used for handling the measurement data. RESULTS： All the 105 rats entered the result analysis. ①After 8 weeks of D-gal treatment, no obvious change of the fasting blood glucose was found in each group （P 〉 0.05）; the blood glucose 2 hours after the orally taken glucose tolerance test and the difference between the fasting blood glucose and the glucose 2 hours after the test in the model group were all higher than the control group （P 〈 0.01） and the AG, Pue, Bet, Bai groups （P 〈 0.05-0.01）; the Gly was lower than the model group, but there was nosignificant difference （P 〉 0.05）. ②After 8 weeks of D-gal treatment, the activity of aldose reduetase in the model group was obviously higher than the other 6 groups （P 〈 0.05-0.01）, the fruetosamine and the end glyeation products of the model group were markedly higher than the control, AG,Pue and Bet groups （P 〈 0.01）; the content of glucosylated hemoglobin was higher than the control, AG, Pue, Ber and Gly groups （P 〈 0.05-0.01）. CONCLUSION： The results confirm that all the AG, Pue, Ber, Bai and Gly can inhibit the activity of aldase reductase; moreover, the AG, Pue and Ber have obviously inhibitory effects on the protein glyeation, and the Bai can partly inhibit the glycation, but the Gly has no effect on the glycated end products.