小鼠口服多糖包覆胰岛素脂质体的降血糖作用小鼠口服多糖包覆胰岛素脂质体的降血糖作用

目的 研究壳聚糖和海藻酸钠两种多糖包覆胰岛素脂质体的小鼠po降血糖作用。方法 用逆相蒸发法制备胰岛素脂质体;用透射电镜和激光粒度仪测定它们的形态和粒径;用HPLC法和超速离心法测定包封率;用胃蛋白酶和胰蛋白酶溶液试验多糖包覆脂质体对胰岛素的保护作用;用酶-苯酚法测定小鼠po多糖包覆胰岛素脂质体后降血糖作用。结果小鼠po 0.1%壳聚糖和0.1%海藻酸钠包覆的胰岛素脂质体具有较好的降血糖作用。结论壳聚糖或海藻酸钠包覆的脂质体能减少胃蛋白酶或胰蛋白酶对胰岛素的降解并促进胰岛素po吸收。     

包裹在脂质体中胰岛素的二级结构

目的　以单纯胰岛素为对照 ,采用傅立叶红外转换光谱 (FourierTransformInfraredFTIR)研究包裹在脂质体内部胰岛素二级结构的变化。方法　分别对单纯胰岛素、胰岛素与空白脂质体混合物 (样品I)及包裹胰岛素的脂质体 (样品II)样品进行FTIR测定。结果　与单纯胰岛素相比 ,样品Ⅰ和Ⅱ与胰岛素的FTIR谱图形状基本一致 ,仅其中的α -螺旋略有下降 (由36 %分别到 32 %～ 31% ) ,β 折叠略有增加 (由 4 8%分别到 5 3%～ 5 1% )。样品I同样品II相比 ,胰岛素的二级结构无明显差别(α 螺旋分别为 32 %～ 31% ,β 折叠分别为 5 3%～ 5 1% )。说明包裹在脂质体内的胰岛素未与脂质体的膜发生插膜作用 ,所产生与单纯胰岛素二级结构之间微小差别的原因是由于部分胰岛素铺展在脂质体膜表面所致的。结论　FTIR测定被载体包裹的蛋白多肽药物的二级结构 ,具有快速直接和非破坏性的特点。经FTIR测定 ,包裹在脂质体内的胰岛素与单纯胰岛素的二级结构相比无明显的变化 ,仍保持原胰岛素的二级结构     

胰岛素脂质体的结构特点

目的研究胰岛素脂质体中胰岛素所处的位置及结构特点.方法采用激光散射测定其粒径大小与分布,用透射电镜测定形态;用HPLC法、荧光扫描、胰蛋白酶降解和聚丙烯胺凝胶电泳等实验手段研究胰岛素在脂质体中所处的位置.结果胰岛素脂质体平均粒径为218.3nm,多分散度为0.073,拟合度为7.2,形态多为圆球或近圆球形.HPLC测定结果显示,胰岛素脂质体中含有胰岛素;荧光扫描结果显示胰岛素脂质体中无胰岛素荧光发射峰;胰岛素脂质体可抵抗胰蛋白酶对胰岛素的降解;胰岛素脂质体的聚丙烯胺凝胶电泳谱图中未见胰岛素的条带.结论胰岛素脂质体中胰岛素被包裹在脂质体的内部,因此可抵抗胰蛋白酶的降解.     

胰岛素经口腔给药对正常大鼠的降血糖作用

目的　研究胰岛素溶液 (insulinsolution ,INS SOL)经正常大鼠口腔给药后的降血糖作用。方法　以血糖水平为指标 ,考察各种吸收促进剂经正常大鼠口腔给药后对INS SOL降血糖作用的影响 ,以皮下注射为对照 ,计算不同条件下INS SOL的药理生物利用度 (pharmacologicalbioavailability ,PA)。 结果　不加吸收促进剂的条件下 ,10U·kg-1的INS SOL经口腔给药后的生物利用较低 (PA =6 9% )。十二烷基硫酸钠 (5 % ,PA =14 5 % ) ,苄泽 78(5 % ,PA =2 0 6 % ) ,脱氧胆酸钠 (5 % ,PA =16 5 % )和卵磷脂(10 % ,PA =13 8% )均增加INS SOL的降血糖作用。苄泽78(5 % )可使INS SOL(5U·kg-1)的PA最高达到 33%。结论　在适当的吸收促进剂的作用下INS SOL经口腔给药后具有明显的降血糖效果。     

HPLC法测定胰岛素脂质体中胰岛素的含量

目的胰岛素脂质体中胰岛素含量的测定.方法采用高效液相色谱法,色谱柱为Hypersil ODS2,流动相为0.1mol L-1磷酸二氢钠-0.05mol L-1硫酸钠-乙睛=36∶36∶28,流速为1ml min-1,检测波长为214nm.结果高、中、低的回收率分别为99.55%,98.23%,100.9%,RSD分别为1.1%,1.7%和0.61%.在5～100μg ml-1测定范围内,线性良好,r=0.9999.结论方法可靠,准确.     

HPCL法测定胰岛素脂质体中胰岛素的含量

目的：胰岛素脂质体中胰岛素含量的测定。方法：采用高效液相色谱法,色谱柱为HypersilODS2,流动相为0．1molL^-1磷酸二氢钠－0．05molL^-1硫酸钠－乙睛＝36：36：28,流速为1mlmin^-1,检测波长为214nm。结果：高、中、低的回收率分别为99．55％,98．23％,100．9％,RSD分别为1．1％,1．7％和0．61％。在5－100μgml^-1测定范围内,线性良好,r＝0．9999。结论：方法可靠、准确。     

口服胰岛素脂质体吸收的细胞学机制

目的:在细胞水平研究胰岛素脂质体的口服吸收机制.方法:以Caco-2模型为小肠上皮细胞模型.在不同浓度下进行通透实验,研究动力学机制;用ELISA方法评价小肠黏膜胞内途径吸收药物的能力;用EDTA、维拉帕米处理细胞,研究胞旁途径,以及P-糖蛋白的外排作用.结果:脂质体可以促进胰岛素的吸收;药物的吸收机制不是简单的被动扩散;药物可以从胞内途径、胞外途径通透;维拉帕米对药物的外排作用不显著;药物作用、通透作用对Caco-2细胞单层完整性的改变是暂时的.结论:脂质体包裹的胰岛素可以通过胞内途径、胞外途径吸收.     

PEG修饰水飞蓟素脂质体的制备及体外释放研究

目的:合成一种PEG硬脂酸单酯,应用Box-Behnken设计优化PEG修饰水飞蓟素脂质体的处方和工艺.方法:采用IR、1H-NMR法表征PEG硬脂酸单酯的结构,采用改进的薄膜分散法制备修饰脂质体,血浆透析法比较脂质体与修饰脂质体的体外释放曲线.结果:水飞蓟素修饰脂质体最佳处方含大豆磷脂:胆固醇:PEG硬脂酸单酯:水飞蓟素为20:4.8:0.9:1(w/w),修饰脂质体的平均粒径为100.9nm,包封率为91.2%,体外释放动力学符合Weibull方程.结论:PEG修饰水飞蓟素脂质体体外释放缓慢,注射给药体内能达到长循环作用.     

胰岛素脂质体药物含量及包封率的测定

目的：建立胰岛素脂质体含量及包封率的测定方法。方法：采用葡聚糖凝胶柱色谱法分离胰岛素脂质体及游离胰岛素,RP-HPLC法测定胰岛素含量,色谱柱：Hypersil ODS2柱（250mm×4．6mm,5μm）,流动相：0．05mol·L^-1硫酸盐缓冲液-乙腈（72：28）;流速：1．0ml·min^-1。结果：辅料及溶剂不干扰胰岛素的含量测定,可准确求得胰岛素脂质体的药物含量及包封率。结论：所用方法简便准确,可用于胰岛素脂质体的含量及包封率测定。     

胰岛素气雾剂经正常及糖尿病大鼠肺部给药后的降血糖作用

目的：研究胰岛素气雾剂经肺部给药后的降血糖作用。方法：建立了大鼠的肺部给药模型,通过经口有入及气管内给药两种途径将溶液型胰岛素气雾剂导入大鼠肺内,采用葡萄糖氧化酶法测定给药后正常大鼠及糖尿病大鼠的血糖水平,并计算其药理生物利用度。结果：在同一种剂量下,经口吸入和气管内给药两种途径在正常及糖尿病大鼠体内均呈现显著的降血糖作用,其药理生物利用度在正常大鼠体内分别为6．0％和11．4％,在糖尿病大鼠体内分别为5．6％和8．7％,结论：胰岛素气雾剂能有效地降低正常及糖尿病大鼠血糖,为进一步开发临床治疗糖尿病的新剂型打下基础。     

Pharmacodynamics of Insulin Following Intravenous and Enteral Administrations of Porcine-Zinc Insulin to Rats

Previous work from this laboratory showed site-dependent variations in the apparent permeability of insulin as measured using the everted rat gut sac technique, with the greatest permeability in the distal jejunum and the lowest in the duodenum (5). To quantify better the rate and extent of insulin absorption from the small intestine, closed-loop in situ experiments were performed in nondiabetic rats. Results correlated with the everted gut sac technique in that the absolute bioavailability determined in situ was higher for insulin solution administered to the more distal region of the intestine (0.133%) than that absorbed from an earlier portion of the intestine (0.059%). While the difference in regional bioavailabilities was not significant (P = 0.08), the blood glucose response showed highly significant differences (P = 0.0015), with severe and prolonged hy-poglycemia resulting from insulin delivered to the distal jejunum/ proximal ileum. Insulin administered iv followed a two-compartment pharmacokinetic model. Whole-body elimination rate constants were similar for both iv and enteral insulin. Although therapeutic quantities of insulin were absorbed from the distal small intestine, absorption enhancers would be necessary to decrease the dose of insulin required.     

Pulmonary Delivery of Free and Liposomal Insulin

The effects of oligomerization and liposomal entrapment on pulmonary insulin absorption were investigated in rats using an intratracheal instillation method. The results indicated that both dimeric and hexameric insulins can be rapidly absorbed into the systemic circulation, producing a significant hypoglycemic response. Intratracheal instillation of insulin in two different oligomerized states has not resulted in any significant difference in the duration of hypoglycemic effect. However, the initial hypoglycemic response (first 10 min) obtained from intratracheal administration of 25 IU/kg hexameric insulin appears to be slower than that from the 25 IU/kg dimeric insulin, thereby suggesting that hexameric insulin may have a lower permeability coefficient across alveolar epithelium than the dimeric insulin. Intratracheal administration of insulin liposomes (dipalmitoylphosphatidyl choline:cholesterol, 7:2) led to facilitated pulmonary uptake of insulin and enhanced the hypoglycemic effect. Nevertheless, similar insulin uptake and pharmacodynamic response were obtained from both the physical mixture of insulin and blank liposomes and liposomally entrapped insulin.     

Facilitation of Pulmonary Insulin Absorption by H-MAP: Pharmacokinetics and Pharmacodynamics in Rats

Purpose. Several low molecular weight amino acids have previously been reported to enable the oral delivery of proteins. In the present studies, the effect of H-MAP (hydroxy methyl amino propionic acid) on the pharmacokinetics (PK) and pharmacodynamics (PD) of porcine insulin delivered to the lungs of rats by spray-instillation (SI) has been determined. Methods. Aliquots (100 l) of increasing doses of porcine insulin alone (0.26, 1.3, 2.6, 13, and 26 U/kg) or combined with increasing doses of H-MAP (5, 10, 16, and 25 mg/kg), at pH 7.2-7.6 were administered intratracheally to fasted anesthetized rats using a micro spray-instillator. Blood samples were collected from the jugular vein at specified intervals and the plasma concentrations of insulin and glucose were determined. The PK and PD of porcine insulin alone following subcutaneous (SC) administration of increasing doses were also determined. Results. The PK of insulin administered either by SI to the lungs or SC injection were absorption rate dependent, resulting in post-peak half-lives 10 to 25-fold greater than the reported intravenous elimination half-life (3 min). The relative bioavailability (F') of insulin administered alone by SI varied from 23.8 to 80% for the lowest and highest insulin dose, respectively. Co-administration of H-MAP and insulin to the lungs significantly changed the PK and PD of insulin in a dose dependent fashion. Maximum PK and PD responses were obtained at an H-MAP dose of 16 mg/kg and an insulin dose of 1.3 U/kg. At this combination, the relative bioavailability of insulin was increased more than 2.5 fold, maximum concentration (C_max) increased 2-fold and the minimum plasma glucose concentration (%MPGC) was reduced more than 2-fold with respect to same dose of insulin alone. A greater total reduction in plasma glucose (%TRPG_0t) was achieved for H-MAP/insulin combination (66 ± 5 %) compared to insulin alone (47 ± 10 %). Conclusion. H-MAP has potential for increasing the pulmonary bioavailability of insulin administered through the lungs.     

高分子化学材料负载短效胰岛素对糖尿病大鼠的降血糖作用

目的研究高分子化学材料制备水凝胶负载短效胰岛素对糖尿病大鼠的降糖作用。方法大鼠以65mg·kg^-1剂量腹腔注射链脲佐菌素制备糖尿病大鼠模型,随机分为模型组、常规组(常规胰岛素注射)、水凝胶组(水凝胶负载短效胰岛素注射),另设正常对照组,每组10只。监测各组大鼠药物干预后24 h内血糖的变化;高糖灌胃后血糖的变化;观察药物干预后肝肾功能水平变化。结果 24h内模型组大鼠血糖无明显变化,常规组与水凝胶组的血糖均于3h时显著下降至最低点后上升,但水凝胶组的血糖上升至12 h后再次下降,并于24 h内维持在原水平的49.3%~55.6%,显著低于同时间常规组的血糖(P<0.05)。高糖灌胃后常规组大鼠血糖于0.5 h下降,于1.5~2.5 h维持于原水平的59.8%~66.4%,显著低于同时间段模型组血糖(P<0.05),水凝胶组血糖值自0.5h持续显著下降(P<0.05),2.5~3.5h时血糖值均较同时间常规组血糖值明显降低(P<0.05),实验过程中无明显低血糖。同时,水凝胶组大鼠的谷丙转氨酶及谷草转氨酶显著低于模型组,其中谷草转氨酶明显低于常规组,但高于正常对照组(P<0.05)。结论高分子化学材料水凝胶负载短效胰岛素后使其不仅降糖效果优于常规胰岛素,并且作用时间明显延长,可进一步应用于胰岛素新剂型的开发。     

维生素A联合地特胰岛素-门冬胰岛素降糖方案对糖尿病患儿糖脂代谢和胰岛素抵抗的影响

目的:分析维生素A联合地特胰岛素-门冬胰岛素降糖方案对糖尿病患儿糖脂代谢和胰岛素抵抗的影响,为临床应用提供参考。方法:选择我院2019-2021年收治的114例1型糖尿病患儿随机分为观察组和对照组各57例,对照组给予地特胰岛素+门冬胰岛素治疗,观察组给予地特胰岛素+门冬胰岛素联合维生素A治疗,两组患儿均持续治疗12周作为治疗后观察时间点,比较两组患儿残存胰岛β细胞功能、糖脂代谢、氧化应激水平、血糖达标时间、胰岛素日用量、低血糖发生情况、并发症发生情况。结果:治疗后观察组胰岛β细胞功能指数(HOMA-β)、低密度脂蛋白胆固醇(HDL-C)、一氧化氮(NO)、超氧化物歧化酶(SOD)水平高于对照组,胰岛素抵抗指数(HOMA-IR)、甘油三酯(TG)、总胆固醇(TC)、低密度脂蛋白胆固醇(LDL-C)、空腹血糖(FPG)、丙二醛(MDA)水平及尿液中8-羟基脱氧鸟苷(8-OhdG)和8-异前列腺素(8-iso-PGF2α)水平低于对照组(P<0.05);治疗后观察组低血糖发生率低于对照组,且低血糖患儿中的低血糖发生次数低于对照组,而低血糖水平高于对照组(P<0.05);治疗后观察...     

壳聚糖-半胱氨酸轭合物对胰岛素酶降解和降血糖作用的影响

目的研究壳聚糖-半胱氨酸轭合物对胰岛素消化酶降解的抑制作用,及其对胰岛素口服降血糖的促进作用.方法合成壳聚糖-半胱氨酸轭合物,体外实验考察其对α-糜蛋白酶和胰蛋白酶降解胰岛素的抑制作用.O1/O2乳化溶剂挥发法制备胰岛素肠溶微球,考察壳聚糖-半胱氨酸轭合物对胰岛素溶液和肠溶微球大鼠灌胃后降血糖作用的影响.结果所合成的壳聚糖-半胱氨酸轭合物巯基含量为200μmol·g-1polymer,体外实验对酶降解胰岛素有明显的保护作用:不加壳聚糖-半胱氨酸轭合物的对照组胰岛素溶液可分别在1 h和5 h内被α-糜蛋白酶和胰蛋白酶完全降解;含4mg·mL-1轭合物的同浓度的胰岛素溶液在相同浓度的酶溶液中经过相同时间,胰岛素剩余量均大于75%.所制备的胰岛素肠溶微球载药量7%,大鼠灌胃给予壳聚糖-半胱氨酸轭合物85 mg·kg-1,可增强胰岛素溶液或肠溶微球的口服降血糖作用.结论壳聚糖-半胱氨酸轭合物对胰岛素口服制剂降血糖作用的增强可能与其酶抑制作用有关.     

米力农诱导的胰岛素抵抗对大鼠糖脂代谢的影响

目的　探讨选择性磷酸二酯酶Ⅲ (PhosphdiesteraseⅢ ,PDE3)抑制剂米力农 (milrinone)对大鼠胰岛素分泌、血糖、血浆游离脂肪酸 (freefattyacid ,FFA)的影响和剂量依赖关系 ,及其在胰岛素钳夹状态下对大鼠糖代谢和胰岛素敏感性的影响。方法　由植入导管给予大鼠不同剂量的米力农 ( 1,5 ,2 5 μmoL·kg- 1 )在不同时相测定血糖、血浆FFA和胰岛素水平并与对照组比较。在意识清醒状态下建立大鼠高胰岛素 正常血糖钳夹技术 ,并在钳夹 12 0min时分别经导管给予米力农 ( 2 5 μmoL·kg- 1 )和 2 5 %二甲基亚砜 (DMSO ,对照组 )。采用气相色谱 -质谱仪 (GC MS)测定糖代谢率。结果　 3个不同剂量米力农组血浆FFA浓度明显高于对照组和给药前 ,在注射后 2min ,各组FFA升高的百分数为 :5 0 %、5 2 %、5 5 % ( 1,5 ,2 5 μmoL·kg- 1 )。在 ( 5 ,2 5 )μmoL·kg- 1 组血浆胰岛素水平也明显高于对照组和给药前 ,仅 2 5 μmoL·kg- 1 组血糖浓度高于对照组和给药前。在胰岛素钳夹研究中 ,米力农处理组大鼠血浆FFA明显高于给药前 ( 173± 15vs 6 34± 87μmoL·kg- 1 ) ,肝糖输出 (HGP)也明显高于给药前( - 5 1± 3 1vs 7 5± 2 0mg·kg- 1 ·min)。葡萄糖输注率 (Glucoseinfusionrate,GIR)明显低于对照组和给药前 (     

Study on the Influencing Factors of Hypoglycemic Effect of Folate Targeted Polymersomes Encapsulating Insulin

PurposeTo study the effects of the density of folic acid (FA) on the hypoglycemic ability of FA-targeted polymersomes as oral insulin carriers. Also to study the change of the hypoglycemic effect of FA-targeted mixed polymersomes added with various mass ratio of d-α-tocopherol polyethylene glycol 1000 succinate (TPGS).MethodsThe FA-targeted polymersomes with different FA molar contents were prepared. The in vitro insulin release experiments in different media for FA-targeted polymersomes with various FA contents were studied. Their quantitative cellular uptake in Caco-2 cells was examined. The in vivo hypoglycemic activity of FA-targeted polymersomes was also studied with diabetic rats. The polymersomes with the optimal FA molar content was chosen to prepare mixed polymersomes with various TPGS contents.ResultsAmong insulin-loaded FA-targeted polymersomes with four different FA molar contents, insulin-loaded polymersomes with 10% FA molar content (insulin-loaded 10%FA-Ps) showed the hightest cellular uptake and the best hypoglycemic response. In addition, the insulin-loaded FA-Ps/TPGS5:1 mixed polymersomes exhibited higher cellular uptake and better hypoglycemic response than the other two insulin-loaded mixed polymersomes adding TPGS did.ConclusionsFA-Ps/TPGS5:1 could be a promising formulation for the oral administration of insulin.     

Effect of Receptor Up-Regulation on Insulin Pharmacokinetics in Streptozotocin-Treated Diabetic Rats

The present study investigated the mechanism by which the disposition of insulin is altered in streptozotocin (STZ)-treated diabetic rats as compared with 48-hr-fasted normal (control) rats. It was shown by an indocyanine green infusion method that the hepatic plasma flow rate (Q _H) in diabetic rats (1.64 ml/min/g liver) is significantly higher than that in control rats (0.982 ml/min/g liver). The portal injection technique revealed that the unidirectional clearance (CL_on), which represents the binding of A₁₄-¹²⁵ I-insulin to surface receptors in the liver, is significantly elevated in diabetic rats, suggesting an increase in the surface receptor number R _T), i.e., up-regulation in the liver. In both control and diabetic rats, the total-body clearance (CL_tot) and steady-state volume of distribution (Vd _ss) of labeled insulin decreased significantly with a simultaneous injection of unlabeled insulin (8 U/kg), confirming that the disposition of insulin is affected largely by specific, saturable receptor-mediated processes. The CL_tot and Vd _ss increased significantly in diabetic rats, while nonspecific portions of these parameters were not changed. From the increases in CL_tot (80%) and Q _H (67%) in diabetic rats, a pharmacokinetic analysis has revealed a 40% increase in the hepatic intrinsic clearance (CL_{int sp}) of A₁₄-¹²⁵ I-insulin via a specific mechanism in diabetic rats. In conclusion, we have provided in vivo evidence for a small increase in CL_{int sp} of insulin in STZ-diabetic rats compared with control rats, which may be caused by an increase in the surface receptor number in the livers of diabetic rats.     

Pharmacokinetic and Pharmacological Profiles of Free and Liposomal Recombinant Human Erythropoietin After Intravenous and Subcutaneous Administrations in Rats

Purpose. Recombinant human erythropoietin (Epo) is used frequently through intravenous (i.v.) and subcutaneous (s.c.) administration for the clinical treatment of the last stage of renal anemia. We encapsulated Epo in liposomes to develop an alternative administration route. The purpose of our study was to evaluate the pharmacokinetics and the pharmacological effects of liposomal Epo in comparison with the Epo after i.v. and s.c. administration to rats. Methods. Epo was encapsulated in liposomes composed of dipalmitoylphosphatidylcholine (DPPC) and soybean-derived sterol mixture (SS) prepared by the reversed-phase evaporation vesicle method. After filtration through a 0.1 m polycarbonate membrane, liposomes were gel filtered (Epo/liposomes). Results. Epo/liposomes showed higher pharmacological activity than Epo/liposomes before gel filtration after i.v. administration to rats. Non-encapsulated Epo lost its activity, whereas encapsulated Epo in liposomes retained it. The pharmacological effects of Epo/liposomes were greater than those of Epo after i.v. administration. Epo/liposomes afforded 3–9 times higher AUC, lower clearance and lower steady-state volume of distribution than Epo after both i.v. and s.c. administrations. Epo/liposomes had an improved pharmacokinetic profile compared with Epo. S.c. administration of Epo/liposomes at 7 h may penetrate primarily (40% of dose) through the blood as a liposome and partly (7% of dose) in lymph. Conclusions. Epo/liposomes may reduce the frequency of injections required for a certain reticulocyte effect in comparison to Epo. The lower clearance of Epo/liposomes may increase the plasma concentrations of Epo, which increases the efficacy.