Human Insulin Absorption from the Intestine in Diabetic Rats

Abstract

The ability of human insulin to produce hypoglycemia in streptozocin-diabetic rats (average weight 500 g) was investigated. A simple solution of human insulin (insulin in hypotonic buffer solution) was administered intraduodenally. Rats received a dose of insulin of either 200 U/kg, 400 U/kg, or 600 U/kg. The hypoglycemic response was most significant when 600 U/kg solution of insulin was administered. The 200 U/kg dose was of two forms; one form was a solution of insulin with a concentration of 25 U/ml, 4 ml, and the other was a solution of insulin with a concentration of 50 U/ml, 2 ml. The dose of 25 U/ml, 4 ml produced a more significant hypoglycemic response than that of 50 U/ml, 2 ml. Carrier-insulin systems were also introduced intraduodenally in streptozocin-diabetic rats. The results indicate that the carrier systems without insulin had glucogenic effect. Despite this glucogenic effect, carrier-insulin systems at 200 U/kg dose were as effective as that of 600 U/kg of insulin solution. We conclude that insulin absorption from the intestine can cause a significant hypoglycemic state if given in a large enough dose, even without the presence of other absorption enhancers in the dosage form.     

Human Insulin Absorption from the Intestine in Diabetic Rats

The ability of human insulin to produce hypoglycemia in streptozocin-diabetic rats (average weight 500 g) was investigated. A simple solution of human insulin (insulin in hypotonic buffer solution) was administered intraduodenally. Rats received a dose of insulin of either 200 U/kg, 400 U/kg, or 600 U/kg. The hypoglycemic response was most significant when 600 U/kg solution of insulin was administered. The 200 U/kg dose was of two forms; one form was a solution of insulin with a concentration of 25 U/ml, 4 ml, and the other was a solution of insulin with a concentration of 50 U/ml, 2 ml. The dose of 25 U/ml, 4 ml produced a more significant hypoglycemic response than that of 50 U/ml, 2 ml. Carrier-insulin systems were also introduced intraduodenally in streptozocin-diabetic rats. The results indicate that the carrier systems without insulin had glucogenic effect. Despite this glucogenic effect, carrier-insulin systems at 200 U/kg dose were as effective as that of 600 U/kg of insulin solution. We conclude that insulin absorption from the intestine can cause a significant hypoglycemic state if given in a large enough dose, even without the presence of other absorption enhancers in the dosage form.     

Effectiveness of Rectal Insulin Suppositories Containing Sodium Cholate in Normal and Insulin Dependent Diabetic Subjects

The effectiveness of 100-U porcine insulin suppository containing 5% sodium cholate was examined in 3 normal volunteers (controls) and 15 diabetics. The hypoglycaemic response, in controls, started at 15 min, peaked at 30 min and returned to the initial level after 45 min, while in diabetics, started at 15 min, peaked at 120 min and persisted for the duration of the study (150 min). The maximum percent reduction in plasma glucose was found to be significantly greater in controls than in diabetics, while the AUC for percent glucose reduction was greater in diabetics than in controls. This was accompanied by a significant rise in serum insulin in controls than in diabetics. The serum insulin peaked at 15 min and remained above the initial value for 45 and 120 min, post administration, in controls and diabetics, respectively. Thus, rectal insulin could be considered as alternative to existing therapies.     

Enhancement of Absorption of Insulin-Loaded Polyisobutylcyanoacrylate Nanospheres by Sodium Cholate After Oral and Subcutaneous Administration in Diabetic Rats

Polyisobutylcyanoacrylate (PIBCA) nanospheres were employed as biodegradable polymeric carriers for oral (p.o.) and subcutaneous (s.c.) delivery of insulin. The polymerization technique used was able to hold 65%–95% of insulin added 30 min after initiation of polymerization. The percentage drug loading was monomer concentration dependent. Insulin adsorption to the nanospheres was measured by radioimmumoassay. Although Pluronic F68 (0.5%) did not significantly alter the in vitro insulin degradation half-life T_50%, sodium cholate (0.5%) increased the degradation T_50% of insulin by 56% (from 13.6 ± 1.6 to 22.1 ± 2 min). This study also investigated the in vivo performance of insulin-loaded PIBCA in aqueous suspension with or without sodium cholate (0.5%) and Pluronic F68 (0.5%) surfactants after oral and subcutaneous administration to alloxan-induced diabetic rats. Insulin absorption was evaluated by its hypoglycemic effect. Insulin associated with PIBCA nanospheres retains its biological activity up to 15 h and 24 h after oral and subcutaneous administrations, respectively. Administered orally, insulin-loaded (75 U/kg) nanospheres, in the presence of surfactants, significantly reduced the mean blood glucose level from 392 ± 32 to 80 ± 13 mg/dl within 2 h and maintained it at 100 mg/dl or less for more than 8 h. On the other hand, the subcutaneous administration of insulin-loaded (25 U/kg) nanospheres significantly decreased the blood glucose level from 406 ± 33 to 88.5 ± 12.8 mg/dl within 1 h, and the lowered glucose level was maintained at 100 mg/dl or less for more than 12 h; it returned to its initial value 24 h after administration. Insulin-loaded nanospheres with surfactants showed significant P < 05) pharmacological availability (PA%) of 37.6% ± 3.7% and 65.2% ± 2.7% after oral and subcutaneous dosages, respectively. The existence of surfactants with PIBCA nanospheres improved the oral PA% by 49.2%. These findings suggest that the developed PIBCA, in the presence of surfactants, would be useful not only in improving insulin gastrointestinal absorption, but also in sustaining its systemic action by lowering the blood glucose to an acceptable level.     

Relative Hypoglycemia of Rectal Insulin Suppositories Containing Deoxycholic Acid, Sodium Taurocholate, Polycarbophil, and Their Combinations in Diabetic Rabbits

In this study, insulin suppositories containing 50 U insulin incorporated with 50 mg of deoxycholic acid, sodium taurocholate, or both were placed in the rectum of alloxan-induced hyperglycemic rabbits. A large decrease in plasma glucose concentrations was observed, and the relative hypoglycemias were calculated to be 38.0%, 34.9%, and 44.4%, respectively, compared with insulin subcutaneous (s.c.) injection (40 U). Insulin suppositories containing 50 mg polycarbophil alone or mixed with 50 mg deoxycholic acid produced relative hypoglycemia of 43.1% and 42.2%, respectively. The most pronounced effect was observed with the addition of polycarbophil to the suppository formulation containing a combination of deoxycholic acid and sodium taurocholate, which produced a 56% relative hypoglycemia compared with subcutaneous injection. These suppository formulations could be very promising alternatives to the current insulin injections, being roughly half as efficacious as subcutaneous injection.     

Relative Hypoglycemia of Rectal Insulin Suppositories Containing Deoxycholic Acid,Sodium Taurocholate,Polycarbophil, and Their Combinations in Diabetic Rabbits

In this study, insulin suppositories containing 50 U insulin incorporated with 50 mg of deoxycholic acid, sodium taurocholate, or both were placed in the rectum of alloxan-induced hyperglycemic rabbits. A large decrease in plasma glucose concentrations was observed, and the relative hypoglycemias were calculated to be 38.0%, 34.9%, and 44.4%, respectively, compared with insulin subcutaneous (s.c.) injection (40 U). Insulin suppositories containing 50 mg polycarbophil alone or mixed with 50 mg deoxycholic acid produced relative hypoglycemia of 43.1% and 42.2%, respectively. The most pronounced effect was observed with the addition of polycarbophil to the suppository formulation containing a combination of deoxycholic acid and sodium taurocholate, which produced a 56% relative hypoglycemia compared with subcutaneous injection. These suppository formulations could be very promising alternatives to the current insulin injections, being roughly half as efficacious as subcutaneous injection.     

Effects of Low-Viscosity Sodium Hyaluronate Preparation on the Pulmonary Absorption of rh-Insulin in Rats

Purpose: A low-viscosity formulation for pulmonary delivery of rh-insulin as model peptide drugs was developed using a solution of sodium hyaluronate. Method: The effects of different concentrations and pH values of low-viscosity solutions of hyaluronate on the pulmonary absorption of rh-insulin were examined after intratracheal administration in rats. The permeation of fluorescein isothiocyanate (FITC)-dextran(molecular weight 4300; FD-4) and insulin through excised rat trachea in vitro were also examined. Results: The hyaluronate (2140 kDa) solutions (0.1% and 0.2%w/v) at pH 7.0 significantly enhanced the pharmacological availability (PAB) of insulin compared to the aqueous solution of insulin at pH 7.0. The absorptionenhancing effect at a concentration of 0.1% w/v hyaluronate was greater than that at a concentration of 0.2% w/v hyaluronate. Furthermore, the greatest absorptionenhancing effect was obtained, regardless of the molecular weight of hyaluronate, when the concentration of hyaluronate was adjusted to 0.47 µM. Absorption-enhancing effects were consistent with the effect of a 0.1% w/v hyaluronate preparation at pH 4.0 and 7.0 on the permeation of FITC-dextran and insulin through excised rat trachea in vitro. Conclusion: Low-viscosity hyaluronate preparation was shown to be a useful vehicle for pulmonary delivery of peptide drugs.     

Absorption of Thyrotropin-Releasing Hormone in Rats Using a Mucoadhesive Buccal Patch

Mucoadhesive buccal patches were evaluated in vitro and in vivo using rats for release of thyrotropin-releasing homne (TRH). TRH (10% w/w) was incorporated into mucoadhesive buccal patches that were custom coformulated with silicone and organic polymers (Dow Coming, Midland, MI) and its release profile was characterized in vitro using a modified Franz diffusion cell. TRH released into pH = 7.0 phosphate buffered saline at 37°C under sink conditions was detected using high-performance liquid chromatography (HPLC). Release of TRH in vitro from the buccal patches was rapid during the first 2 hr, with 51% of the total amount of TRH incolporated into the patches released after 24 hr. HPLC analysis indicated that TRH extracted from buccal patches thermally stressed at 40°, 55°, and 70°C showed negligible degradation after 6 months. In contrast, an aqueous TRH solution stored at 70°C showed degradation of TRH as soon as 10 days following incubation at this temperature. TRH patches placed on the buccal mucosa of anesthetized rats demonstrated rapid stimulation and release of thyroid-stimulating hormone (TSH) from the anterior pituitary. Thirty minutes after patch application, plasma concentrations of TSH fluctuated but remained approximately 4-to 7-fold greater than baseline (prior to patch application) TSH concentrations. Therefore, this preliminary study has demonstrated that physiologically active TRH was released from the TRH mucoadhesive buccal patches and systemically absorbed. Thus, the TRH mucoadhesive buccal patches may represent a convenient delivery system for therapeutic peptides.     

Absorption of Thyrotropin-Releasing Hormone in Rats Using a Mucoadhesive Buccal Patch

Abstract

Mucoadhesive buccal patches were evaluated in vitro and in vivo using rats for release of thyrotropin-releasing homne (TRH). TRH (10% w/w) was incorporated into mucoadhesive buccal patches that were custom coformulated with silicone and organic polymers (Dow Coming, Midland, MI) and its release profile was characterized in vitro using a modified Franz diffusion cell. TRH released into pH = 7.0 phosphate buffered saline at 37°C under sink conditions was detected using high-performance liquid chromatography (HPLC). Release of TRH in vitro from the buccal patches was rapid during the first 2 hr, with 51% of the total amount of TRH incolporated into the patches released after 24 hr. HPLC analysis indicated that TRH extracted from buccal patches thermally stressed at 40°, 55°, and 70°C showed negligible degradation after 6 months. In contrast, an aqueous TRH solution stored at 70°C showed degradation of TRH as soon as 10 days following incubation at this temperature. TRH patches placed on the buccal mucosa of anesthetized rats demonstrated rapid stimulation and release of thyroid-stimulating hormone (TSH) from the anterior pituitary. Thirty minutes after patch application, plasma concentrations of TSH fluctuated but remained approximately 4–to 7-fold greater than baseline (prior to patch application) TSH concentrations. Therefore, this preliminary study has demonstrated that physiologically active TRH was released from the TRH mucoadhesive buccal patches and systemically absorbed. Thus, the TRH mucoadhesive buccal patches may represent a convenient delivery system for therapeutic peptides.     

Effect of Different Bile Salts on the Relative Hypoglycemia of Witepsol W35 Suppositories Containing Insulin in Diabetic Beagle Dogs

Insulin suppositories were formulated using Witepsol W35 as a base to investigate the effect of various bile salts/acids on the plasma glucose concentration of diabetic beagle dogs. Comparison of the effect of these formulations was made with that produced by insulin subcutaneous injections. Of the bile salts/acids studied, incorporation of 100 mg of deoxycholic acid (DCA), sodium cholate (NaC), or sodium deoxycholate (NaDC) with insulin (10 U/Kg) showed that suppositories containing NaDC produced the highest area under the curve (AUC) and relative hypoglycemia (RH) of 290 ± 83 mg%h and 28% ± 8.1%, respectively. To study the optimum amount of NaDC in insulin suppositories to produce the highest RH, 50-200 mg/suppository were used, and we found that 150 mg NaDC produced 35% ± 13% RH. We also studied the influence of different doses of insulin (5-20 U/kg) in the presence of NaDC (100 mg). It was found that increase of the insulin dose was accompanied by an increase in AUC and maximum reduction in plasma glucose level C_max. A combination of NaDC (100 mg) and NaC (50 mg) produced an AUC of 252 ± 13 mg% h and an RH of 49% ± 2.6%, which were higher than produced by either of its individual components (NaC 50 mg or NaDC 100 mg) when used alone or when compared with an equivalent amount of NaDC (150 mg). When the effect of sodium taurocholate (NaTC) and sodium taurodeoxycholate (NaTDC) was studied, it was found that an insulin suppository containing 100 mg of either NaTC or NaTDC produced an RH equivalent to that produced previously with a mixture of NaDC (100 mg) and NaC (50 mg). On the other hand, NaC (50 mg) did not improve the hypoglycemic effect of NaTC any further. In conclusion, a relative hypoglycemia of about 50% can be reached using insulin suppositories containing Witepsol W35 as a base and NaDC plus NaC (100 mg plus 50 mg, respectively), NaTDC (100 mg), or NaTC (100 mg) as rectal absorption enhancers of insulin. A desirable hypoglycemia, expressed as C_max, and/or AUC can be reached by adjusting the insulin dose in the formulation according to the degree of hyperglycemia.     

The Relationship Between the Rigidity of the Liposomal Membrane and the Absorption of Insulin After Nasal Administration of Liposomes Modified with an Enhancer Containing Insulin in Rabbits

The relationship between the rigidity of the liposomal membrane and the absorption of insulin after nasal administration of liposomes modified with an enhancer containing insulin was investigated for the nasal delivery of peptide drugs in rabbits. The rigid liposomal membrane makes liposomes stable, protecting insulin from enzymatic degradation. Soybean-derived sterol (SS) or its sterylglucoside (SG) was used as an enhancer. Dipalmitoylphosphatidylcholine (DPPC) liposomes modified with SG had increased fluidity of the hydrophobic group of the liposome bilayer compared with the liposomes modified with cholesterol (Ch) or SS, as shown by measurements of the steady-state fluorescence anisotropy of 1,6-diphenyl-1,3,5,-hexatriene (DPH); however, the fluidity of the polar group of the liposome bilayer was decreased according to measurements of steady-state fluorescence anisotropy of dansylhexadecylamine (DSHA) at 37°C. These findings suggest that the fluidity of the hydrophobic group of the liposome bilayer is responsible for the increase of liposomal leakage and instability of the liposomes. When insulin was administered nasally to rabbits as a solution, no hypoglycemic effect was observed. The administration of insulin contained in DPPC/SG (7/4, mole) liposomes with high fluidity caused a high glucose reduction of long duration (8 hr). DPPC/SS and DPPC/Ch (7/4) liposomes with low fluidity caused low glucose reductions. These results demonstrated that liposomes modified with SG can be useful as carriers of insulin administered nasally.     

Prelimnaky Evaluation of Cissus Root Gum as a Binder in Sodium Salicylate Tablet Formulations

Abstract

Cissus root gum was processed and evaluated as a binder in lactose-based tablets each containing 100 mg of sodium salicylate as the active ingredient. Acacia binder was used as basis for comparison. Tablet hardness, friability, disintegration time and dissolution rate were the parameters investigated. The cissus gum gave hard and non-friable tablets at 1 - 3% w/w concentration of the tablet formula. Tablets containing above 2% w/w of the cissus gum gave high disintegration time values and the pattern of dissolution of the incorporated drug suggests that the gum may be useful in prolonged release tablet formulations. No significant changes in the tablet properties was observed after storage at 30°C for 16 weeks.     

钯对钛膜吸氢能力的抗污染作用的研究

为寻求一种防止因表面污染而导致钛膜吸氢能力下降的途径，用表面分析方法检测样品表面状态，用质谱仪测量样品吸氧能力，研究了表面状态和吸氢能力的相互关系。采用在钛膜上淀积钯膜（蒸发或溅射）的方法，可使受碳、氧污染的钛膜吸氢能力得以恢复。这种钯／钛复合结构在吸氢能力上对碳、氧污染并不灵敏。对样品的近费米能级处的占有电子态密度（densityofstate，DOS）的测量证明，凡吸氢能力良好的样品，DOS呈峰形结构。具有抗污染能力的钯／钛结构，其DOS因污染而导致的变化很小，而无抗污染能力的钛膜，其峰形结构受污染作用而消失。这种峰形结构能提供氢分子解离吸附过程中所需的电子。     

Quantitation of Nizatidine in Capsules Using High-Performance Liquid Chromatography

A stability-indicating high-performance liquid chromatography method for the quantitation of nizatidine in capsules has been developed. The method is accurate and precise with a percent relative standard deviation of 0.34 based on 6 readings. A number of inactive ingredients present in the capsules did not interfere in the assay procedure. The recovery from the synthetic mixtures was quantitative. The extraction procedure from the capsules is very simple. The drug appears to be very sensitive to bases (such as sodium hydroxide) since 100% of the drug decomposed on boiling for 35 minutes. The drug was very stable when boiled with sulfuric acid.     

Development of Diclofenac Sodium Controlled Release Solid Dispersion Powders and Capsules by Freeze Drying Technique Using Ethylcellulose and Chitosan as Carriers

Abstract

Controlled-release, solid dispersions of diclofenac sodium (DS) were prepared by freeze-drying technique, using ethylcellulose (EC) and chitosan (CS) as single and combined carriers. Factorial design was applied as an experimental design to study the main and interactive effects of EC and CS on drug dissolution from the controlled release solid dispersion. All DS solid dispersions showed slower drug dissolution than did DS powder. The equations of dissolution parameters as functions of EC and CS contents were established through multiple regression. The contour plots of the established equations were constructed. The 10: (2.4 + 0.05) DS:(EC + CS) solid dispersion was prepared and developed into a capsule dosage from, using lactose as diluent. The effect on capsule dissolution of a disintegrant, sodium starch glycolate (Explotab®), in concentrations of 2%, 5%, and 8% was studied. The solid-dispersion capsule containing 5% Explotab was found to provide the most similar dissolution profile to the one obtained with the 10:(2.4 + 0.05) DS:(EC + CS) solid-dispersion powder. The dissolutions of the 10:(2.4 + 0.05) solid-dispersion powder and capsules were closer to a first-order model than to a zero-order or diffusion control model.     

奥氏体不锈钢在含铀碳酸钠溶液中的腐蚀行为

用动电位极化法和均匀腐蚀法研究了奥氏体不锈钢(1Cr18Ni9Ti)在含铀碳酸钠溶液中的腐蚀行为.结果表明:铀酰离子对碳酸钠溶液中奥氏体不锈钢电极的阳极反应有抑制作用,95 ℃温度下,经过2 000 h,奥氏体不锈钢在0.06 mol/L Na4UO2(CO3)3 1.1 mol/L Na2CO3混合溶液中未发生可观测的腐蚀.给出了奥氏体不锈钢电极在含铀碳酸钠溶液中的腐蚀电位、腐蚀电流密度.腐蚀样品经XRD分析表明沉积物由UO3和Na4UO2(CO3)3组成,XPS分析表明其中铀的氧化态为铀(Ⅵ).初步探讨了碳酸铀酰离子的作用机制,讨论了奥氏体不锈钢耐蚀的原因.     

用石墨炉原子吸收光谱法测定次磷酸钠中微量铅

试样直接用水溶解,以磷酸氢二铵作测铅基体改进剂,考察了基体改进剂用量,原子化温度以及基体元素对测定的影响,在最佳测定条件下,用石墨炉原子吸收光谱法测定了次磷酸钠中微量铅,测试结果为:线性范围为0～60×10-3μg/mL,检出限为1.6×10-3μg/mL,回收率为94.0%～102.6%,RSD<5%。方法简便,灵敏度高,重现性较好。     

Use of CACO-2 Cells as an in vitro Intestinal Absorption and Metabolism Model

Abstract

The Caco-2 cell line, a human colorectal carcinoma cell line, is an established in vitro model for the study of drug transport in the human intestine. We have routinely utilized this in vitro model to 1) elucidate intestinal absorption mechanisms of small drug molecules and peptide-like therapeutic agents (e.g. paracellular/transcellular passive diffusion and carrier-mediated active transport), 2) screen and select orally active therapeutic agents, 3) identify optimum luminal pH's for drug absorptions, 4) address dissolution rate-related absorption problems, 5) assess mucosal toxicity of therapeutic agents, and 6) evaluate prodrug approaches for enhanced drug absorptions. We have also utilized this in vitro model to assess the metabolic stability of therapeutic agents in the intestinal epithelium. Demonstrated in this report are primarily the techniques for the elucidation of absorption mechanisms. Examples of the characterization of paracellular/ transcellular passive diffusion pathways and carrier-mediated active transport will be given. Application of the Caco-2 model to the process of drug development will also be discussed.     

Use of CACO-2 Cells as an in vitro Intestinal Absorption and Metabolism Model

The Caco-2 cell line, a human colorectal carcinoma cell line, is an established in vitro model for the study of drug transport in the human intestine. We have routinely utilized this in vitro model to 1) elucidate intestinal absorption mechanisms of small drug molecules and peptide-like therapeutic agents (e.g. paracellular/transcellular passive diffusion and carrier-mediated active transport), 2) screen and select orally active therapeutic agents, 3) identify optimum luminal pH's for drug absorptions, 4) address dissolution rate-related absorption problems, 5) assess mucosal toxicity of therapeutic agents, and 6) evaluate prodrug approaches for enhanced drug absorptions. We have also utilized this in vitro model to assess the metabolic stability of therapeutic agents in the intestinal epithelium. Demonstrated in this report are primarily the techniques for the elucidation of absorption mechanisms. Examples of the characterization of paracellular/ transcellular passive diffusion pathways and carrier-mediated active transport will be given. Application of the Caco-2 model to the process of drug development will also be discussed.