桑叶黄酮提取纯化及其降血脂作用研究
摘要
桑叶(mulberry leaf)为桑树(Morus alba L.)的叶,黄酮是其中主要的生物活性成分,为深度利用资源,本文对黄酮的提取分离技术进行了工艺研究,并后研究了桑叶黄酮对高尿酸血症模型的降尿酸的作用。
研究通过高压液相色谱法对桑叶黄酮类化合物进行成分分析,结果表明,其主要成分为芦丁,杨梅素,槲皮素,山奈素等。采用正交试验法对提取桑叶黄酮的工艺进行优化,结果表明,最佳的浸提条件为：在80℃下70%乙醇回流提取2次,每次1.5h,料液比为1：20(w/v)。
为得到纯度较高的桑叶总黄酮,采用大孔树脂吸附法进行黄酮纯化,以桑叶总黄酮的得率和纯度为指标,比较了五种大孔树脂的吸附性能,结果显示HPD826大孔树脂对桑叶总黄酮吸附性能最好；进一步采用正交试验法对HPD826大孔树脂纯化桑叶黄酮的工艺进行优化,所获最佳条件组合为：取浓度为4mg/ml左右的桑叶提取液上样,上样体积为1.5树脂体积数(BV),上样速率控制在2BV/h左右,之后用3 BV,60%的乙醇,以1 BV/h的速率洗脱,纯化后的桑叶黄酮得率为1.95%,纯度55.41%。
药理学研究表明：桑叶黄酮具有调节α-葡萄糖苷酶、降血糖、抗肿瘤等药理活性。而且黄酮类化合物对高尿酸血症有效。所以选择体外实验探讨桑叶黄酮(MLF)对大鼠肝脏匀浆液中黄嘌呤氧化酶(XO)活性的影响,结果显示：MLF剂量依赖性地抑制大鼠肝脏匀浆液XO活性,在100.00 mg/L抑制率与别嘌醇(AP)无显著差异,提示MLF可能具有降尿酸活性。进一步研究其对正常小鼠血清UA含量和肝脏匀浆液XO活性的影响,以50、100、200mg/kg*d-1剂量MLF灌胃正常小鼠14d,测定相关指标。结果显示：MLF三种剂量与对照相比对正常小鼠体内血清UA含量和肝脏匀浆XO活性均无显著性影响,对小鼠体重和脏器系数无显著影响。
采用氧嗪酸钾诱导的高尿酸血症小鼠模型评价了MLF的降尿酸活性,以50、100、200mg/kg*d-1剂量MLF灌胃模型小鼠14d,检测相关指标,结果显示：200mg/kg bw MLF可极显著(P<0.01)降低模型小鼠血清尿酸含量,下降16.67%；并能显著(P<0.05)抑制肝脏XO活性,抑制率达6.05%。而50mg/kg bw MLF对肝脏XO活性的无明显影响(P>0.05),但能极显著降低模型小鼠血清尿酸含量(P<0.01)。
长期高尿酸血症导致肾损害和痛风,肾损害亦可致尿酸排泄障碍。为进一步探讨桑叶黄酮对病理状态下的尿酸代谢及相关肾损伤的影响,本实验继续研究了MLF对腺嘌呤联合高糖饲料诱导的大鼠高尿酸血症、肾损伤的防治作用。在腺嘌呤联合高糖饲料诱导的高尿酸病理状态下,分别给予200、100、50mg/kg* d-1的MLF,以别嘌醇为阳性对照药物。结果显示：MLF中、高剂量组在给药1w后,血清UA水平分别降低17.83%和21.17%；2w后降低21.09%和27.15%。3w后MLF中剂量组血清尿酸接近于正常大鼠水平,XO活力比HC组显著降低25.01%。200mg/kg*d-1 MLF可使血清TG,FFA水平降低41.95%,42.05%,同时MLF可减小腺嘌呤造模引发的肝脏、心脏系数的异常。降低高尿酸病理状态下肌酐和尿素氮含量,并通过电镜观察,MLF对腺嘌呤所引起肾脏损伤有一定的保护作用。
综合以上结果：推断MLF具有明显的降尿酸活性,并对高尿酸病理状态下的肾脏损伤有一定的保护作用；MLF可能通过抑制XO和调节脂质紊乱来调节血清UA水平。
Leaves from Morus alba L.is a kind of traditional Chinese medicine. Mulberry leaf flavonoids are the mainly active ingredient in mulberry leaves. The technique of extraction and separation of active ingredient has been investigated in this paper. Thenceforth, the hypouricemic activity of the mulberry leaves flavonoids was explored.
Mulberry leaf flavonoids were analysed with high performance liquid chromatography. Rrutin, myricetin, quercetin, kaempferol are the main ingredients in it. Factors of temperature, extraction time, ethanol concentration,volume of solvent and extraction frequency were evaluated for their effects on extraction process by orthogonal designed experiments. The results showed that the optimum conditions were 80℃,2 times,1.5 hours,70% ethanol-herb ratio of volume to mass was 20:1 respectively.
In order to get purer mulberry leaf flavonoids, macroporous resin was adopted. The adsorption ability of macroporous resin was evaluated by yield and purity of the flavonoids. The kind of HPD826 was the best of the five macroporous resin. The optimum process conditions were 3 BV 60% ethanol as eluting solvent, adding sample rate in about 2 BV/h, eluting rate in about 1 BV/h. The yield of Mulberry leaf flavonoids by this process was 1.95%,and the purity was 55.41%.
Pharmacology study showed that mulberry leaf flavonoids could adjust the activity of a-glucosidase, lower blood glucose, antitumor, etc. Results indicated that MLF inhibit rat liver XO activity on dose dependent. Normal mice were administered orally with MLF at doses of 50,100,150 mg./kg*d-1.The results showed that mulberry leaf flavonoids had no significant effect on uric acid levels and liver XO activity in normal mice.
Experimental animal model of hyperuricemia was induced by uricase inhibitor potassium oxonate. Potassium oxonate-induced hyperuricemic mice were administered orally with MLF at doses of 50,100,150 mg/kg*d-1, while distilled water and allopurinol at 50 mg/kg*d-1 were given to the hyperuricemic control group and positive control group. The results showed that oral administration of 200 mg/kg*d-1 MLF to hyperuricemic mice exhibited significant reduction in the blood uric acid levels 16.67% and in XO activity 6.05%. The oral administration of 50 mg./kg*d-1 MLF also caused remarkable reduction in the blood uric acid levels, but without remarkable reduction in XO activity.
Long-term hyperuicemia caused kidney damage and gout, renal damage cause uric acid excretion obstacles. We studied the hypouricemic effect of MLF by using model of adenine-reduced hyperuricemia in rats. In this test, Adenine-induced hyperuricemic mice were administered orally with MLF at doses of 50,100,150 mg/kg*d-1, while distilled water and allopurinol at 50 mg/kg*d-1 were given to the hyperuricemic control group and positive control group. The results showed that oral administration of 100 mg/kg*d-1 MLF to hyperuricemic mice exhibited significant reduction in the blood uric acid levels at different time periods:17.83% at 7th d, 21.17% at 14th d, and oral administration of 200 mg/kg*d-1 MLF caused remarkable reduction in the blood uric acid levels at different time periods:21.09% at 7th d, 27.15% at 14th d. Oral administration of 200 mg/kg*d-1 MLF also caused remarkable reduction in the blood triglyceride levels 41.95% and in the blood free fatty acid levels 42.05% at 21th d. Transmission electron microscopy (TEM) observation results showed that MLF protected kidney cells of rats which treated by adenine.
All the results mentioned above showed that MLF could decrease the levels of blood uric acid. It also protect kidney cell and prevent it from further damage, improve the metabolism of triglyceride and adjust body weight in hyperuricemic mice. It was concluded that MLF had good activity on adenine-reduced hyperuricemic rats, but its molecular mechanism need further study.
研究通过高压液相色谱法对桑叶黄酮类化合物进行成分分析,结果表明,其主要成分为芦丁,杨梅素,槲皮素,山奈素等。采用正交试验法对提取桑叶黄酮的工艺进行优化,结果表明,最佳的浸提条件为：在80℃下70%乙醇回流提取2次,每次1.5h,料液比为1：20(w/v)。
为得到纯度较高的桑叶总黄酮,采用大孔树脂吸附法进行黄酮纯化,以桑叶总黄酮的得率和纯度为指标,比较了五种大孔树脂的吸附性能,结果显示HPD826大孔树脂对桑叶总黄酮吸附性能最好；进一步采用正交试验法对HPD826大孔树脂纯化桑叶黄酮的工艺进行优化,所获最佳条件组合为：取浓度为4mg/ml左右的桑叶提取液上样,上样体积为1.5树脂体积数(BV),上样速率控制在2BV/h左右,之后用3 BV,60%的乙醇,以1 BV/h的速率洗脱,纯化后的桑叶黄酮得率为1.95%,纯度55.41%。
药理学研究表明：桑叶黄酮具有调节α-葡萄糖苷酶、降血糖、抗肿瘤等药理活性。而且黄酮类化合物对高尿酸血症有效。所以选择体外实验探讨桑叶黄酮(MLF)对大鼠肝脏匀浆液中黄嘌呤氧化酶(XO)活性的影响,结果显示：MLF剂量依赖性地抑制大鼠肝脏匀浆液XO活性,在100.00 mg/L抑制率与别嘌醇(AP)无显著差异,提示MLF可能具有降尿酸活性。进一步研究其对正常小鼠血清UA含量和肝脏匀浆液XO活性的影响,以50、100、200mg/kg*d-1剂量MLF灌胃正常小鼠14d,测定相关指标。结果显示：MLF三种剂量与对照相比对正常小鼠体内血清UA含量和肝脏匀浆XO活性均无显著性影响,对小鼠体重和脏器系数无显著影响。
采用氧嗪酸钾诱导的高尿酸血症小鼠模型评价了MLF的降尿酸活性,以50、100、200mg/kg*d-1剂量MLF灌胃模型小鼠14d,检测相关指标,结果显示：200mg/kg bw MLF可极显著(P<0.01)降低模型小鼠血清尿酸含量,下降16.67%；并能显著(P<0.05)抑制肝脏XO活性,抑制率达6.05%。而50mg/kg bw MLF对肝脏XO活性的无明显影响(P>0.05),但能极显著降低模型小鼠血清尿酸含量(P<0.01)。
长期高尿酸血症导致肾损害和痛风,肾损害亦可致尿酸排泄障碍。为进一步探讨桑叶黄酮对病理状态下的尿酸代谢及相关肾损伤的影响,本实验继续研究了MLF对腺嘌呤联合高糖饲料诱导的大鼠高尿酸血症、肾损伤的防治作用。在腺嘌呤联合高糖饲料诱导的高尿酸病理状态下,分别给予200、100、50mg/kg* d-1的MLF,以别嘌醇为阳性对照药物。结果显示：MLF中、高剂量组在给药1w后,血清UA水平分别降低17.83%和21.17%；2w后降低21.09%和27.15%。3w后MLF中剂量组血清尿酸接近于正常大鼠水平,XO活力比HC组显著降低25.01%。200mg/kg*d-1 MLF可使血清TG,FFA水平降低41.95%,42.05%,同时MLF可减小腺嘌呤造模引发的肝脏、心脏系数的异常。降低高尿酸病理状态下肌酐和尿素氮含量,并通过电镜观察,MLF对腺嘌呤所引起肾脏损伤有一定的保护作用。
综合以上结果：推断MLF具有明显的降尿酸活性,并对高尿酸病理状态下的肾脏损伤有一定的保护作用；MLF可能通过抑制XO和调节脂质紊乱来调节血清UA水平。
Leaves from Morus alba L.is a kind of traditional Chinese medicine. Mulberry leaf flavonoids are the mainly active ingredient in mulberry leaves. The technique of extraction and separation of active ingredient has been investigated in this paper. Thenceforth, the hypouricemic activity of the mulberry leaves flavonoids was explored.
Mulberry leaf flavonoids were analysed with high performance liquid chromatography. Rrutin, myricetin, quercetin, kaempferol are the main ingredients in it. Factors of temperature, extraction time, ethanol concentration,volume of solvent and extraction frequency were evaluated for their effects on extraction process by orthogonal designed experiments. The results showed that the optimum conditions were 80℃,2 times,1.5 hours,70% ethanol-herb ratio of volume to mass was 20:1 respectively.
In order to get purer mulberry leaf flavonoids, macroporous resin was adopted. The adsorption ability of macroporous resin was evaluated by yield and purity of the flavonoids. The kind of HPD826 was the best of the five macroporous resin. The optimum process conditions were 3 BV 60% ethanol as eluting solvent, adding sample rate in about 2 BV/h, eluting rate in about 1 BV/h. The yield of Mulberry leaf flavonoids by this process was 1.95%,and the purity was 55.41%.
Pharmacology study showed that mulberry leaf flavonoids could adjust the activity of a-glucosidase, lower blood glucose, antitumor, etc. Results indicated that MLF inhibit rat liver XO activity on dose dependent. Normal mice were administered orally with MLF at doses of 50,100,150 mg./kg*d-1.The results showed that mulberry leaf flavonoids had no significant effect on uric acid levels and liver XO activity in normal mice.
Experimental animal model of hyperuricemia was induced by uricase inhibitor potassium oxonate. Potassium oxonate-induced hyperuricemic mice were administered orally with MLF at doses of 50,100,150 mg/kg*d-1, while distilled water and allopurinol at 50 mg/kg*d-1 were given to the hyperuricemic control group and positive control group. The results showed that oral administration of 200 mg/kg*d-1 MLF to hyperuricemic mice exhibited significant reduction in the blood uric acid levels 16.67% and in XO activity 6.05%. The oral administration of 50 mg./kg*d-1 MLF also caused remarkable reduction in the blood uric acid levels, but without remarkable reduction in XO activity.
Long-term hyperuicemia caused kidney damage and gout, renal damage cause uric acid excretion obstacles. We studied the hypouricemic effect of MLF by using model of adenine-reduced hyperuricemia in rats. In this test, Adenine-induced hyperuricemic mice were administered orally with MLF at doses of 50,100,150 mg/kg*d-1, while distilled water and allopurinol at 50 mg/kg*d-1 were given to the hyperuricemic control group and positive control group. The results showed that oral administration of 100 mg/kg*d-1 MLF to hyperuricemic mice exhibited significant reduction in the blood uric acid levels at different time periods:17.83% at 7th d, 21.17% at 14th d, and oral administration of 200 mg/kg*d-1 MLF caused remarkable reduction in the blood uric acid levels at different time periods:21.09% at 7th d, 27.15% at 14th d. Oral administration of 200 mg/kg*d-1 MLF also caused remarkable reduction in the blood triglyceride levels 41.95% and in the blood free fatty acid levels 42.05% at 21th d. Transmission electron microscopy (TEM) observation results showed that MLF protected kidney cells of rats which treated by adenine.
All the results mentioned above showed that MLF could decrease the levels of blood uric acid. It also protect kidney cell and prevent it from further damage, improve the metabolism of triglyceride and adjust body weight in hyperuricemic mice. It was concluded that MLF had good activity on adenine-reduced hyperuricemic rats, but its molecular mechanism need further study.
引文
1、中国桑树品种志[M].北京：农业出版社，1993：5.
2、国家药典委员会.中国药典,Ⅰ部[S].北京：化学工业出版社，2002：244.
3、宋立人,洪恂,丁绪亮,等.现代中药学大辞典[M].北京：人民卫生出版社,2001：1821.
4、高学敏.中药学[M].北京：人民卫生出版社,2000：309.
5、Andallu B, Suryakantham V,Lakshmi Srikanthi B, et al. Effect of mulberry (Morus indica L.) therapy on plasma and erythrocyte membrane lipids in patients with type 2 diabetes[J].Clin Chim Acta,2001,314(1-2):47.
6、Andallu B, Varadacharyulu NCh. Control of hyperglycemia and retardation of cataract by mulberry (Morus indica L.) leaves in streptozotocin diabetic rats [J]. Indian JExp Bio,2002,40(7):791.
7、Doi K, Kojima T, Fujimoto Y. Mulberry leaf extract inhibits the oxidative modification of rabbit and human low density lipoprotein [J]. BiolPharm Bul,2000, 23(9):1066.
8、Kim SY, Gao JJ, Kang HK. Two flavonoids from the leaves of Morus alba induce differentiation of the human promyelocytic leukemia (HL-60) cell line [J]. BiolPharm Bull,2000,23(4):451.
9、刘利,殷志琦,张现涛等.桑叶几种化学物质的分离鉴定[J].蚕业科学,2005,31(3)：331-333.
10、刘利,殷志琦,张雷红等.桑叶次生代谢产物分析[J].蚕业科学,2005,31(4)：413～417.
11、邵继红,沈洪兵,莫宝庆等.社区人群高尿酸血症危险因素的病例对照研究[J].中华流行病学杂志,2004，25：688～690.
12、Johnson R J, Feig D I, Herrera-Acosta J,t al. Resurrection of uric acid as a causal risk factor in essential hypertension[J]. Hypertension,2005,45(1):28-33.
13、Thomas B.Current management of gout in patients unresponsive or allergic to allopurinol[J]. Joint Bone Spine,2004,71:481-485.
14、陈晓红,赵志刚,魏丽荣.嘌醇的不良反应文献分析及探讨[J].实用药物与临 床,2010,13(2)：149～151.
15、L.D.Kong, Y. Cai, W.W.Huang, et al. Inhibition of xanthine oxidase by some Chinese medicinal plants used to treat gout[J].Journal of Ethnopharmacology,73 (2000):199-207.
16、Dewanto V,Wu X,Adom KK,et al.Thermal processing enhances the nutritional value of tomatoes by increasing total antioxidant activity[J]. J Agric Food Chem, 2002,50:3010-3014.
17、李静,聂继云,李海飞,等Folin-酚法测定水果及其制品总多酚含量的条件[J].果树学报,2008,25(1)：126-131.
18、赵道辉,金玉玲,倪蕾.枸杞及其制品中功效成分多糖的测定研究[J].中华医学实践杂志,2004,3(6)：542-543.
19、邹士玉,马方励,丁博.苯酚-硫酸法测定“润和津露”粗多糖含量[J].中药材,2007,30(11)：1740—1742.
20、余启荣,晏利芝,谭湘湘HPLC法测定银杏叶提取物中总黄酮醇苷的含量[J].首都师范大学学报,2009,30(2)：32-34.
21、孙斌,瞿伟菁,张小玲等.高效液相色谱法测定沙棘籽渣中黄酮苷元含量[J].中国药学杂志,2005,40(2)：139-141.
22、刘咏,行春丽,成战胜.从桑叶中提取多酚、黄酮和多糖的优化试验[J].林产化工通讯.2005,39(1)：10-13.
23、原爱红,黄哲,马骏.桑叶黄酮的提取及其降糖作用的研究.从桑叶中提取多酚、黄酮和多糖的优化试验[J].中草药.2004,35(11)：1242-1243.
24、赵凤琴,李先佳,任丽平.正交试验优选桑叶黄酮及其降糖作用研究安徽农业科学[J].2009,37(20)：9485-9486.
25、倪学文,吴谋成.大孔吸附树脂提取纯化银杏酚酸的研究[J].中草药.2003,34(3)：231.
26、邹节明,陆浩,何斌.苦玄参、黄芩与黄柏的大孔树脂提取研究[J].中草药,2003,34(3)：222.
27、Kong L D, Yang C, Ge F,et al. A Chinese herbal medicine Ermiao wan reduces serum uric acid level and inhibits liver xanthine dehydrogenase and xanthine oxidase in mice[J].J Ethnopharmacol,2004,93:325-330.
28、杨澄,朱继孝,王颖等.盐制对黄柏抗痛风作用的影响[J].中国中药杂志,2005, 30：145-148.
29、赵赞,赵洪军,瞿伟菁等.越桔果渣黄酮对腺嘌呤诱导大鼠高尿酸血症肾损伤的防治作用[J].营养学报,2009,31(2)：149-151.
30. Zhu JX, Wang Y, Kong LD,et al. Effects of Biota orientalis extract and its flavonoid constituents, quercetin and rutin on serum uric acid levels in oxonate-induced mice and xanthine dehydrogenase and xanthine oxidase activities in mouse liver[J]. J Ethnopharmacol,2004,93:133-140.
31、屠晓钢,王舒然,乔凌.银杏黄酮对卡铂所致大鼠肾损伤的防护作用[J].中国实用医学,2006,4：34-36.
32、Francesca V, Giovanna L, Elena R, et al. Mild hyperuricemia and subclinical renal damage in untreated primary hypertension[J]. Am J Hypertens,2007,20:1276-1282.
33、Kinugasa Y, Ogino K, Tomikura Y, et al. Hyperuricemia is a new therapeutic target of heart failure[J].J Cardiac Failure,2004,10:192.
34、曹汉民.生物电子显微镜技术[M].华东师范大学出版社,1990：3-8.
35、Wu-X, Wakamiya M, Vaishnav S, et al. Hyperuricemia and urate nephropathy in urate oxidase-deficient mice [J]. Proc Nat,1994, (2):7421.
36、王宗涛,管春梅,车会莲,等.高尿酸血症大鼠模型的实验研究[J].中华内分泌代谢杂志,2004,20(5)：4741.
37、陈光亮,孙秀霞,王钦茂等.小鼠高尿酸血症模型的研究[J].中国药理学通报,2001,17(3)：350-352.
38、刘小青,张冰,刘春梅等.鹌鹑高尿酸血症模型建立与初探[J].中国病理生理杂志,2001,17(10)：1038-1040.
39、徐曼,孙铭柱.生长因子在实验性尿酸盐肉芽肿形成和转归中的作用[J].重庆医科大学学报,2003，28(6)：769-774.
40、张超,曹克光.高尿酸血症及尿酸性肾病动物模型的建立及应用[J].实验动物科学与管理,1999,16(4)：18.
41、林伟青,谢建祥.高尿酸血症动物模型研究进展[J].中国药物与临床,2005,5(2)：115.
42、苏友新,陈伟宏,王和鸣等.痛风宁颗粒对实验性鸡痛风性关节炎的影响[J].中国骨伤,2003，16(5)：275-277.
43、徐立,时乐.小鼠代谢性高尿酸血症模型的复制方法初探[J].中国比较医学杂 志,2006,16(1)：1-4.
44、唐灿,杨奎.高尿酸血症动物模型初探[J].中药新药与临床药理,2000,9,11(5)：292-294
45、申海莉.酸脂清胶囊治疗尿酸性肾病及高脂血症的实验研究[J].辽宁中医学院学报,2005,7(1)：71.
46、申海莉,文绍敦.酸脂清胶囊治疗尿酸性肾病的实验研究[J].青海医学院学报,2005,26(1)：57.
47、何立群,聂永红,邹士林.新型尿酸性肾病动物模型的建立[J].中华国际医学杂志,2003，3(3)：213.
48、何立群,邹士林,聂永红等.降尿酸方对尿酸性肾病动物肾小管间质病变干预作用[J].中华国际医学杂志,2003,3(3)：213.
49、陈光亮,张清林,马晓芹等.酵母致小鼠高尿酸血症模型[J].中国药理学通报,2003,19(4)：467-469.
50、杨红莲,张冰,刘小青等.雌性鹌鹑高尿酸血症模型研究[J].辽宁中医杂志,2009,36(1)：6-8.
51、张冰,刘小青,丁正磊等.鹌鹑高尿酸血症模型发病机理研究[J].北京中医药大学学报,2006,29(9)：595-598.
52、刘小青,张冰,胡京红,等.鹌鹑高尿酸血症模型建立及莴苣提取物对其影响的实验研究[J].中国中西医结合杂志,2003,23(1)：44.
53、Johnson W. J, Stavric B. Chartrand A, Uricase inhibition intheratbys-triazines:an anima 1 mode 1 for hyperuicemia and hyperuri-cosuria [J]. Proc. Soc. Expt. Bio. Med,1969,131 (1):8.
54、Iris H. H, John P. S, Dvid J. R, et al. Substitutedcy-clic imides as potential anti-gout agents [J]. Life sciense,1990,46(26):1923.
55、Kazumi IVAKI,Yukio YONETANI. Hyperuricemic effects of cholinergica-gentsin rats [J]. Japan. J. Pharmco,1982,32:343.
56、徐立,时乐,小鼠代谢性高尿酸血症模型的复制方法初探[J].中国比较医学杂志,2006,16(1)：1-4.
57、曹瑞竹,张三印，代勇等.秦皮总香豆素降低小鼠急性高尿酸血症血尿酸水平及机理研究[J].辽宁中医杂志,2010,37(2)：362-364.
58、王海东,葛飞,郭玉松等.金钱草提取物对高尿酸血症小鼠的影响[J].中国中 药杂志,2002,27(12)：939.
59、喻志峰,杨澄,仇熙等.瑞香苷对高尿酸血症小鼠的影响[J].中国药科大学学报,2002,33(2)：142.
60、徐立,时乐,仲文欣.尿酸酶抑制剂在大鼠急性高尿酸血症模型复制中的应用[J].中国比较医学杂志，2007,17(3)：142-146.
61、陈文照,徐红,谷焕鹏.尿酸利仙冲剂治疗痛风性肾病的实验研究.中国中西医结合肾病杂志,2002,3(7)：422.
62、陈心智,黄晓巍,欧喜燕.痛风安胶囊对高尿酸血症大鼠血尿酸水平影响的实验研究[J].长春中医药大学学报,2007,23(2)：8-9.
63、熊湘明,曲竹秋,贾锡莲.大鼠高尿酸血症肾损害模型的建立[J].中国比较医学杂志,2005,15(4)：206-209.
64、金沈锐,郑军，刘绍唐.小鼠高尿酸血症模型初探[J].成都中医药大学学报,1999,3,22(10).
65、Kelly SJ, Delnomdedieu M, Oliverio M I, et al. Diabetes insip idus in uricase-deficientmice:A model for evaluating therapywith poly(ethylene glycol)-modified uricase [J]. J Am Soc Nephrol,2001,12(5):1001.
66、李会芳,孙晓波,方芳.高尿酸血症动物模型研究[J].辽宁中医学院学报.2004,6(5)：415-416.
67、孟永会,曾俞霖.高尿酸血症动物模型研究[J].中国民族民间医药.2010,11：124-125.
68、李东晓,迟家敏.高尿酸血症与代谢综合征[J].国外医学内分泌学分册,2004,24(6)：386-388.
69、符彬,陈新.高尿酸血症与心血管疾病关系的Meta分析[J].中国全科医学,2007,10(6)：451-452.
70、刘梅颜,胡大一.高尿酸血症与高纤维蛋白原血症对冠状动脉风险的联合评估价值探讨[J].中华医学杂志,2006,86(10)：678-680.
71、LIU J, LI L. Pathogenesy and pharmacotherapy of hyperuricem ia:research advances [J]. J Int Pharm Res, Vol.37, No.1, February,2010:24-28.
72、Rafey MA, Lipkowitz MS, Leal-Pinto E, et al. Uric acid transport. Current Opinion in [J].Nephrology Hypertension,2003,12 (5):511-516.
73、Lipkowitz MS, Leal-Pinto E, Rappoport JZ, et al. Functional reconstitution, membrane targeting,genomic structure and chromosomal localization of a human urate transporter[J]. J Clin Invest,2001,107:1103-1115.
74、Enomoto A, Kimura H, Chairoungdua A, et al. Molecular identification of a renal urate anion exchanger that regulates blood urate levels[J]. Nature,2002,417:447-452.
75、Ichida K, Hosoyamada M, Kimura H, et al. Urate transport via human PAH transporter hOAT1 and its gene structure [J]. Kidney Int,63:143-155.
76、Cha SH, Sekine T, Fukushima J I, et al. Identification and characterization of human organic anion transporter 3 expressing predominantly in the kidney [J]. Mol Pharmacol,2001,59:1277-1286.
77、Okada H, Danoff TM, Kalluri R, et al. Early role of Fspl in epithelial-mesenchymal transformation [J]. Am J Physiol,1997,273 (4p t2): F563-F574.
78、Ikenouchi J, Matsuda M, Furuse M, et al. Regulation of tight junctions during the ep ithelium-mesenchyme transition:direct repression of the gene exp ression of claudins/occluding by Snail [J].J Cell Sci,2003,116 (p t10):1959-1967.
79、Baker JF, Krishnan E, Chen L, et al. Serum uric acid and cardiovascular disease: recent developments, and where do they leave us [J]. Am J Med,2005,118 (8):816-826.
2、国家药典委员会.中国药典,Ⅰ部[S].北京：化学工业出版社，2002：244.
3、宋立人,洪恂,丁绪亮,等.现代中药学大辞典[M].北京：人民卫生出版社,2001：1821.
4、高学敏.中药学[M].北京：人民卫生出版社,2000：309.
5、Andallu B, Suryakantham V,Lakshmi Srikanthi B, et al. Effect of mulberry (Morus indica L.) therapy on plasma and erythrocyte membrane lipids in patients with type 2 diabetes[J].Clin Chim Acta,2001,314(1-2):47.
6、Andallu B, Varadacharyulu NCh. Control of hyperglycemia and retardation of cataract by mulberry (Morus indica L.) leaves in streptozotocin diabetic rats [J]. Indian JExp Bio,2002,40(7):791.
7、Doi K, Kojima T, Fujimoto Y. Mulberry leaf extract inhibits the oxidative modification of rabbit and human low density lipoprotein [J]. BiolPharm Bul,2000, 23(9):1066.
8、Kim SY, Gao JJ, Kang HK. Two flavonoids from the leaves of Morus alba induce differentiation of the human promyelocytic leukemia (HL-60) cell line [J]. BiolPharm Bull,2000,23(4):451.
9、刘利,殷志琦,张现涛等.桑叶几种化学物质的分离鉴定[J].蚕业科学,2005,31(3)：331-333.
10、刘利,殷志琦,张雷红等.桑叶次生代谢产物分析[J].蚕业科学,2005,31(4)：413～417.
11、邵继红,沈洪兵,莫宝庆等.社区人群高尿酸血症危险因素的病例对照研究[J].中华流行病学杂志,2004，25：688～690.
12、Johnson R J, Feig D I, Herrera-Acosta J,t al. Resurrection of uric acid as a causal risk factor in essential hypertension[J]. Hypertension,2005,45(1):28-33.
13、Thomas B.Current management of gout in patients unresponsive or allergic to allopurinol[J]. Joint Bone Spine,2004,71:481-485.
14、陈晓红,赵志刚,魏丽荣.嘌醇的不良反应文献分析及探讨[J].实用药物与临 床,2010,13(2)：149～151.
15、L.D.Kong, Y. Cai, W.W.Huang, et al. Inhibition of xanthine oxidase by some Chinese medicinal plants used to treat gout[J].Journal of Ethnopharmacology,73 (2000):199-207.
16、Dewanto V,Wu X,Adom KK,et al.Thermal processing enhances the nutritional value of tomatoes by increasing total antioxidant activity[J]. J Agric Food Chem, 2002,50:3010-3014.
17、李静,聂继云,李海飞,等Folin-酚法测定水果及其制品总多酚含量的条件[J].果树学报,2008,25(1)：126-131.
18、赵道辉,金玉玲,倪蕾.枸杞及其制品中功效成分多糖的测定研究[J].中华医学实践杂志,2004,3(6)：542-543.
19、邹士玉,马方励,丁博.苯酚-硫酸法测定“润和津露”粗多糖含量[J].中药材,2007,30(11)：1740—1742.
20、余启荣,晏利芝,谭湘湘HPLC法测定银杏叶提取物中总黄酮醇苷的含量[J].首都师范大学学报,2009,30(2)：32-34.
21、孙斌,瞿伟菁,张小玲等.高效液相色谱法测定沙棘籽渣中黄酮苷元含量[J].中国药学杂志,2005,40(2)：139-141.
22、刘咏,行春丽,成战胜.从桑叶中提取多酚、黄酮和多糖的优化试验[J].林产化工通讯.2005,39(1)：10-13.
23、原爱红,黄哲,马骏.桑叶黄酮的提取及其降糖作用的研究.从桑叶中提取多酚、黄酮和多糖的优化试验[J].中草药.2004,35(11)：1242-1243.
24、赵凤琴,李先佳,任丽平.正交试验优选桑叶黄酮及其降糖作用研究安徽农业科学[J].2009,37(20)：9485-9486.
25、倪学文,吴谋成.大孔吸附树脂提取纯化银杏酚酸的研究[J].中草药.2003,34(3)：231.
26、邹节明,陆浩,何斌.苦玄参、黄芩与黄柏的大孔树脂提取研究[J].中草药,2003,34(3)：222.
27、Kong L D, Yang C, Ge F,et al. A Chinese herbal medicine Ermiao wan reduces serum uric acid level and inhibits liver xanthine dehydrogenase and xanthine oxidase in mice[J].J Ethnopharmacol,2004,93:325-330.
28、杨澄,朱继孝,王颖等.盐制对黄柏抗痛风作用的影响[J].中国中药杂志,2005, 30：145-148.
29、赵赞,赵洪军,瞿伟菁等.越桔果渣黄酮对腺嘌呤诱导大鼠高尿酸血症肾损伤的防治作用[J].营养学报,2009,31(2)：149-151.
30. Zhu JX, Wang Y, Kong LD,et al. Effects of Biota orientalis extract and its flavonoid constituents, quercetin and rutin on serum uric acid levels in oxonate-induced mice and xanthine dehydrogenase and xanthine oxidase activities in mouse liver[J]. J Ethnopharmacol,2004,93:133-140.
31、屠晓钢,王舒然,乔凌.银杏黄酮对卡铂所致大鼠肾损伤的防护作用[J].中国实用医学,2006,4：34-36.
32、Francesca V, Giovanna L, Elena R, et al. Mild hyperuricemia and subclinical renal damage in untreated primary hypertension[J]. Am J Hypertens,2007,20:1276-1282.
33、Kinugasa Y, Ogino K, Tomikura Y, et al. Hyperuricemia is a new therapeutic target of heart failure[J].J Cardiac Failure,2004,10:192.
34、曹汉民.生物电子显微镜技术[M].华东师范大学出版社,1990：3-8.
35、Wu-X, Wakamiya M, Vaishnav S, et al. Hyperuricemia and urate nephropathy in urate oxidase-deficient mice [J]. Proc Nat,1994, (2):7421.
36、王宗涛,管春梅,车会莲,等.高尿酸血症大鼠模型的实验研究[J].中华内分泌代谢杂志,2004,20(5)：4741.
37、陈光亮,孙秀霞,王钦茂等.小鼠高尿酸血症模型的研究[J].中国药理学通报,2001,17(3)：350-352.
38、刘小青,张冰,刘春梅等.鹌鹑高尿酸血症模型建立与初探[J].中国病理生理杂志,2001,17(10)：1038-1040.
39、徐曼,孙铭柱.生长因子在实验性尿酸盐肉芽肿形成和转归中的作用[J].重庆医科大学学报,2003，28(6)：769-774.
40、张超,曹克光.高尿酸血症及尿酸性肾病动物模型的建立及应用[J].实验动物科学与管理,1999,16(4)：18.
41、林伟青,谢建祥.高尿酸血症动物模型研究进展[J].中国药物与临床,2005,5(2)：115.
42、苏友新,陈伟宏,王和鸣等.痛风宁颗粒对实验性鸡痛风性关节炎的影响[J].中国骨伤,2003，16(5)：275-277.
43、徐立,时乐.小鼠代谢性高尿酸血症模型的复制方法初探[J].中国比较医学杂 志,2006,16(1)：1-4.
44、唐灿,杨奎.高尿酸血症动物模型初探[J].中药新药与临床药理,2000,9,11(5)：292-294
45、申海莉.酸脂清胶囊治疗尿酸性肾病及高脂血症的实验研究[J].辽宁中医学院学报,2005,7(1)：71.
46、申海莉,文绍敦.酸脂清胶囊治疗尿酸性肾病的实验研究[J].青海医学院学报,2005,26(1)：57.
47、何立群,聂永红,邹士林.新型尿酸性肾病动物模型的建立[J].中华国际医学杂志,2003，3(3)：213.
48、何立群,邹士林,聂永红等.降尿酸方对尿酸性肾病动物肾小管间质病变干预作用[J].中华国际医学杂志,2003,3(3)：213.
49、陈光亮,张清林,马晓芹等.酵母致小鼠高尿酸血症模型[J].中国药理学通报,2003,19(4)：467-469.
50、杨红莲,张冰,刘小青等.雌性鹌鹑高尿酸血症模型研究[J].辽宁中医杂志,2009,36(1)：6-8.
51、张冰,刘小青,丁正磊等.鹌鹑高尿酸血症模型发病机理研究[J].北京中医药大学学报,2006,29(9)：595-598.
52、刘小青,张冰,胡京红,等.鹌鹑高尿酸血症模型建立及莴苣提取物对其影响的实验研究[J].中国中西医结合杂志,2003,23(1)：44.
53、Johnson W. J, Stavric B. Chartrand A, Uricase inhibition intheratbys-triazines:an anima 1 mode 1 for hyperuicemia and hyperuri-cosuria [J]. Proc. Soc. Expt. Bio. Med,1969,131 (1):8.
54、Iris H. H, John P. S, Dvid J. R, et al. Substitutedcy-clic imides as potential anti-gout agents [J]. Life sciense,1990,46(26):1923.
55、Kazumi IVAKI,Yukio YONETANI. Hyperuricemic effects of cholinergica-gentsin rats [J]. Japan. J. Pharmco,1982,32:343.
56、徐立,时乐,小鼠代谢性高尿酸血症模型的复制方法初探[J].中国比较医学杂志,2006,16(1)：1-4.
57、曹瑞竹,张三印，代勇等.秦皮总香豆素降低小鼠急性高尿酸血症血尿酸水平及机理研究[J].辽宁中医杂志,2010,37(2)：362-364.
58、王海东,葛飞,郭玉松等.金钱草提取物对高尿酸血症小鼠的影响[J].中国中 药杂志,2002,27(12)：939.
59、喻志峰,杨澄,仇熙等.瑞香苷对高尿酸血症小鼠的影响[J].中国药科大学学报,2002,33(2)：142.
60、徐立,时乐,仲文欣.尿酸酶抑制剂在大鼠急性高尿酸血症模型复制中的应用[J].中国比较医学杂志，2007,17(3)：142-146.
61、陈文照,徐红,谷焕鹏.尿酸利仙冲剂治疗痛风性肾病的实验研究.中国中西医结合肾病杂志,2002,3(7)：422.
62、陈心智,黄晓巍,欧喜燕.痛风安胶囊对高尿酸血症大鼠血尿酸水平影响的实验研究[J].长春中医药大学学报,2007,23(2)：8-9.
63、熊湘明,曲竹秋,贾锡莲.大鼠高尿酸血症肾损害模型的建立[J].中国比较医学杂志,2005,15(4)：206-209.
64、金沈锐,郑军，刘绍唐.小鼠高尿酸血症模型初探[J].成都中医药大学学报,1999,3,22(10).
65、Kelly SJ, Delnomdedieu M, Oliverio M I, et al. Diabetes insip idus in uricase-deficientmice:A model for evaluating therapywith poly(ethylene glycol)-modified uricase [J]. J Am Soc Nephrol,2001,12(5):1001.
66、李会芳,孙晓波,方芳.高尿酸血症动物模型研究[J].辽宁中医学院学报.2004,6(5)：415-416.
67、孟永会,曾俞霖.高尿酸血症动物模型研究[J].中国民族民间医药.2010,11：124-125.
68、李东晓,迟家敏.高尿酸血症与代谢综合征[J].国外医学内分泌学分册,2004,24(6)：386-388.
69、符彬,陈新.高尿酸血症与心血管疾病关系的Meta分析[J].中国全科医学,2007,10(6)：451-452.
70、刘梅颜,胡大一.高尿酸血症与高纤维蛋白原血症对冠状动脉风险的联合评估价值探讨[J].中华医学杂志,2006,86(10)：678-680.
71、LIU J, LI L. Pathogenesy and pharmacotherapy of hyperuricem ia:research advances [J]. J Int Pharm Res, Vol.37, No.1, February,2010:24-28.
72、Rafey MA, Lipkowitz MS, Leal-Pinto E, et al. Uric acid transport. Current Opinion in [J].Nephrology Hypertension,2003,12 (5):511-516.
73、Lipkowitz MS, Leal-Pinto E, Rappoport JZ, et al. Functional reconstitution, membrane targeting,genomic structure and chromosomal localization of a human urate transporter[J]. J Clin Invest,2001,107:1103-1115.
74、Enomoto A, Kimura H, Chairoungdua A, et al. Molecular identification of a renal urate anion exchanger that regulates blood urate levels[J]. Nature,2002,417:447-452.
75、Ichida K, Hosoyamada M, Kimura H, et al. Urate transport via human PAH transporter hOAT1 and its gene structure [J]. Kidney Int,63:143-155.
76、Cha SH, Sekine T, Fukushima J I, et al. Identification and characterization of human organic anion transporter 3 expressing predominantly in the kidney [J]. Mol Pharmacol,2001,59:1277-1286.
77、Okada H, Danoff TM, Kalluri R, et al. Early role of Fspl in epithelial-mesenchymal transformation [J]. Am J Physiol,1997,273 (4p t2): F563-F574.
78、Ikenouchi J, Matsuda M, Furuse M, et al. Regulation of tight junctions during the ep ithelium-mesenchyme transition:direct repression of the gene exp ression of claudins/occluding by Snail [J].J Cell Sci,2003,116 (p t10):1959-1967.
79、Baker JF, Krishnan E, Chen L, et al. Serum uric acid and cardiovascular disease: recent developments, and where do they leave us [J]. Am J Med,2005,118 (8):816-826.
© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号 地址：北京市海淀区学院路29号 邮编：100083 电话：办公室：(+86 10)66554848；文献借阅、咨询服务、科技查新：66554700 |