Improved bioavailability of orally delivered insulin using Eudragit-L30D coated PLGA microparticles

Large porous microparticles of PLGA entrapping insulin were prepared by solvent evaporation method and evaluated in diabetes induced rat for its efficacy in maintaining blood sugar level from a single oral dose. Incorporation of Eudragit L30D (0.03% w/v) in the external aqueous phase resulted in formation of pH responsive enteric coated polymer particles which release most of the entrapped insulin in alkaline pH. At acidic pH, release of insulin from uncoated PLGA microparticles and Eudragit L30D coated PLGA microparticles was 31.62 +/- 1.8% and 17.5 +/- 1.29%, respectively, for initial 30 min. However, in 24 h, in vitro released insulin from uncoated PLGA and Eudragit coated particles was 96.29 +/- 1.01% and 88.30 +/- 1%, respectively. Released insulin from composite polymer particles were mostly in monomer form without aggregation and was stable for a month at 37 degrees C. Oral administration of insulin loaded PLGA (50 : 50) and Eudragit L30D coated PLGA (50 : 50) microparticles (equivalent to 25 IU insulin/kg of animal weight) in alloxan induced diabetic rats resulted in 37.3 +/- 11% and 62.7 +/- 3.8% reduction in blood glucose level, respectively, in 2 h. This effect continued up to 24 h in the case of Eudragit L30D coated PLGA microparticles. Results demonstrate that use of stabilizers during PLGA particle formulation, large porous particle for quick release of insulin and coating with Eudragit L30D resulted in a novel oral formulation for once a day delivery of insulin.     

Toxicological assessment of orally delivered nanoparticulate insulin

Subacute toxicological assessment on diabetic rats was conducted after 15 days of daily oral administration of nanoparticulate insulin. Haematological and biochemical analyses were conducted on blood and urine, biopsies performed on organs and tissues, and histology analysed by optical microscopy. Insulin-loaded nanoparticles alone did not change liver or kidney functions. The increase of some hepatic parameters was attributed to diabetes physiopathology and to chemical inducement of diabetes and not to the nanoparticle composition since diabetic controls showed the same variations. In terms of kidney function, parameters such as urea nitrogen and creatinine, were also similar to normal rats with the exception of glycosuria. This single effect was due to diabetes physiopathology and the method of induction, and not to the nanoparticle composition, since non-dosed diabetic rats showed the same alteration. Even so, glycosuria levels of animals dosed with insulin-loaded nanoparticles were lower than control diabetic rats which may indicate an effective hypoglycaemic response. Nanoparticles did not exhibit toxicity in haematological parameters. Finally, organ histology was similar between dosed animals and normal rats with the exception of pancreas histology.     

Release optimization of epidermal growth factor from PLGA microparticles

Abstract

The objective of this study was to prepare poly lactic-co-glycolic acid (PLGA)-based microparticles as potential carriers for recombinant human epidermal growth factor (rhEGF). In order to optimize characteristic parameters of protein-loaded microspheres, bovine serum albumin (BSA) was selected as the model protein. To reduce burst release as a common problem of microspheres, a proper alteration in the particle composition was used, such as addition of poly vinyl alcohol and changes in initial drug loading. The effects of these parameters on particle size, encapsulation efficiency and in vitro release kinetics of BSA in PLGA microspheres were investigated using a Box–Behnken response surface methodology. The biological activity of the released rhEGF was assessed using human skin fibroblasts cell proliferation assay. The prepared rhEGF-loaded microspheres had an average size of 6.44?±?2.45?µm, encapsulation efficiency of 97.04?±?1.13%, burst release of 13.06?±?1.35% and cumulative release of 22.56?±?2.41%. The proliferation of human skin fibroblast cells cultivated with rhEGF releasate of microspheres was similar to that of pure rhEGF, indicating the biological activity of released protein confirming the stability of rhEGF during microsphere preparation. These results are in agreement with the purpose of our study to prepare rhEGF-entrapped PLGA microparticles with optimized characteristics.     

Enhancement of oral insulin bioavailability: in vitro and in vivo assessment of nanoporous stimuli-responsive hydrogel microparticles

ABSTRACT

Objective: Oral insulin administration suffers gastrointestinal tract (GIT) degradation and inadequate absorption from the intestinal epithelium resulting in poor bioavailability. This study entails in vitro and in vivo assessment of stimuli-responsive hydrogel microparticles (MPs) in an attempt to circumvent GI barrier and enhance oral insulin bioavailability.

Methods: Bacterial cellulose-g-poly(acrylic acid) (BC-g-P(AA)) hydrogel MPs were evaluated for morphology, swelling, entrapment efficiency (EE), in vitro insulin release and enzyme inhibition. The ex vivo mucoadhesion, insulin degradation and transport were investigated in excised intestinal tissues. The effect of MPs on paracellular transport was studied in Caco-2/HT29-MTX monolayers. The in vivo hypoglycemic effect and pharmacokinetics of insulin-loaded MPs were investigated in diabetic rats.

Results: Hydrogel MPs efficiently entrapped insulin (EE up to 84%) and exhibited pH-responsive in vitro release. The MPs decreased the proteolytic activity of trypsin (up to 60%). Insulin transport across monolayers was increased up to 5.9-times by MPs. Histological assessment of GI tissues confirmed the non-toxicity of MPs. Orally administered insulin-loaded MPs showed higher hypoglycemic effect as compared to insulin solution and enhanced relative oral bioavailability of insulin up to 7.45-times.

Conclusion: These findings suggest that BC-g-P(AA) MPs are promising biomaterials to overcome the barriers of oral insulin delivery and enhancing its bioavailability.     

Influence of the primary emulsification procedure on the characteristics of small protein-loaded PLGA microparticles for antigen delivery

Microparticles prepared from poly(lactic-co-glycolic acid) (PLGA) using a W₁/O/W₂ double emulsion solvent evaporation method are suitable vehicles for the delivery of proteins to antigen presenting cells, e.g. dendritic cells. In this study, the influence of different techniques for the preparation of the primary W₁/O emulsion was investigated with respect to the protein localization within the microparticles, morphological characteristics of these particles, protein burst release and the native state of the released protein. Bovine serum albumin bearing fluorescein isothiocyanate (FITC-BSA) was used as model protein. A static micromixer was applied for the preparation of the W₁/O/W₂ double emulsion. Employing a rotor-stator homogenizer (Ultra-Turrax®) for primary emulsification, microcapsules with a high burst release were produced, because nearly all FITC-BSA was attached to the outside of the particle wall. Using a high pressure homogenizer or an ultrasonic procedure resulted in the formation of microspheres with homogeneous protein distribution and a reduced burst release.     

混合膜材及添加剂对包衣膜性质的影响

目的制备肠溶性包衣膜,考察混合膜材以及添加剂对膜材性质的影响。方法采用平面铸膜法制备Eudragit NE30D/L 30D-55及Eudragit FS 30D/L 30D-55游离膜,并以膜的透湿性、机械性能为指标,考察膜材比例及添加剂[增塑剂柠檬酸三乙酯(TEC)、抗黏剂单硬脂酸甘油酯(GMS)、乳化剂吐温80(Tween-80)]对游离膜的影响。结果 Eudragit NE 30D及Eudragit FS 30D的加入能够增加Eudragit L 30D-55膜材的延展性、弹性和耐撞击负载的能力,同时也会减小膜材的强度、增加其透湿性。Eudragit L 30D-55与Eudragit NE 30D的混合膜材透湿性较大,对于湿度敏感性药物,应尽量避免选择透湿性大的包衣膜材。TEC和Tween-80的加入可改善膜材的柔韧性和弹性,降低膜材的强度,同时,TEC的加入可以增加膜材的透湿性,而GMS对膜材各项机械性质和透湿性影响不大。结论通过简单的调节混合膜材的种类、比例、添加剂的用量可以制得符合要求的肠溶游离膜。     

Eudragit NE30D膜的制备及其性能参数的标准化研究

目的考察Eudragit NE30D膜的制备条件,探索膜性能参数的标准化方法。方法分别使用质量分数5%、7.5%、10%、12.5%、15%的Eudragit NE30D水分散液,考察了1、2 m L两种体积和25、40、55℃温度对Eudragit NE30D膜制备的影响,比较了其厚度、均匀度、表面平滑度、含水量、透湿量、抗张强度、断裂伸长率。结果 Eudragit NE30D适宜的成膜温度为40℃,成膜时间4~6 h,操作体积为1 m L,质量分数为5%~10%,控制含水量5%。制得的膜标准化参数、透湿性和机械性能的重现性良好。结论本法简单可行,为研究Eudragit NE30D性能提供参考。     

Preparation of preformed porous PLGA microparticles and antisense oligonucleotides loading

The objective of this study was to load preformed highly porous microparticles with drug. The microparticles were prepared by a modified multiple emulsion (w/o/w) solvent evaporation method with the addition of pore formers (NaCl into the internal aqueous phase or of glycerol monooleate to the poly(lactide-co-glycolide) (PLGA) polymer phase). The drug-free solidified microparticles were then washed with either water (for NaCl) or hexane (for glycerol monooleate) to extract the pore formers. The drug was then loaded into the preformed porous microparticles by incubation in aqueous drug solutions followed by air- or freeze-drying. The drug was strongly bound to the polymeric surface with air-dried microparticles. A biphasic drug release with an initial rapid release phase (burst effect) was followed by a slower release up to several weeks. The initial burst was dependent on the drug loading and could be significantly reduced by wet (non-aqueous) temperature curing.     

Effects of alginate coated on PLGA microspheres for delivery tetracycline hydrochloride to periodontal pockets

The effects of alginate coated on tetracycline (Tc) loaded poly (D, L-lactic-co-glycolic acid) (PLGA) microspheres fabricated by double emulsion solvent evaporation technique for local delivery to periodontal pocket were investigated. Alginate coated PLGA microspheres showed smoother surface but enlarged their particle sizes compared with those of uncoated ones. In addition, alginate coated microspheres enhanced Tc encapsulation efficiency (E.E.) from 11.5?±?0.5% of uncoated ones to 17.9?±?0.5%. Moreover, all of the coated PLGA microspheres even fabricated at different conditions could prolong Tc release from 9–12 days with 50% or higher in cumulative release of Tc compared with those of uncoated ones. The swelling ratios of PLGA microspheres for alginate coated or uncoated ones, one of the possible mechanisms for enhancing Tc release for the coated ones, were measured. The results showed that 20% or higher in swelling ratio for the coated microspheres at the earlier stage of hydration (e.g.?≤?24?h) could be an important factor to result in high Tc release compared to the uncoated ones. In conclusion, alginate coated Tc loaded PLGA microspheres could enhance Tc delivery to periodontal pocket by enhancing drug encapsulated efficiency, released quantities and sustained release period compared with uncoated ones.     

门冬胰岛素30与精蛋白生物合成人胰岛素30R对2型糖尿病治疗效果比较

目的研究门冬胰岛素30(诺和锐30)与精蛋白生物合成人胰岛素30R(预混30R)治疗2型糖尿病(T2DM)的效果。方法选取某院T2DM患者92例(2017年1月~2019年1月),按照治疗方法分为A组、B组,各46例。A组采用诺和锐30治疗,B组采用预混30R治疗。比较两组治疗前、治疗3个月后血糖[空腹血糖(FBG)、餐后2 h血糖(2hPBG)、睡前血糖、糖化血红蛋白(HbA1c)]水平、不良反应及低血糖发生率。结果治疗3个月后A组FBG、2hPBG、睡前血糖、HbA1c水平均低于B组(P<0.05);A组不良反应总发生率为8.70%,B组不良反应总发生率为17.39%,组间比较差异无统计学意义(P>0.05);A组低血糖发生率4.35%低于B组17.39%(P<0.05)。结论与精蛋白生物合成人胰岛素30R对比,门冬胰岛素30治疗T2DM能显著降低血糖水平及低血糖发生率,二者安全性高。     

Microencapsulation of insulin using a W/O/W double emulsion followed by complex coacervation to provide protection in the gastrointestinal tract

Abstract

Microcapsules containing insulin were prepared using a combination of a W/O/W double emulsion and complex coacervation between WPI (used as a hydrophilic emulsifier) and CMC or SA with further spray drying of the microcapsules in order to provide protection in the gastrointestinal tract. The microcapsules prepared exhibited high encapsulation efficiency and showed the typical structure of a double emulsion. After spray drying of these microcapsules, the integrity of the W/O/W double emulsion was maintained and the biological residual activity remained high when using the combination of 180?°C inlet air temperature and 70?°C outlet air temperature. The microcapsules exhibited low solubility at pH 2 and high solubility at pH 7 so they might protect insulin at acid pH values in the stomach and release it at intestinal pH values. The microcapsules developed in this study seem to be a promising oral delivery vehicle for insulin or other therapeutic proteins.     

Design,development and in vitro-in vivo study of a colon-specific fast disintegrating tablet

Objective: Targeted delivery systems for the treatment of Inflammatory bowel disease (IBD) are designed to increase local tissue concentrations of anti-inflammatory drugs from lower doses compared with systemic administration. The objective of this study was to formulate and evaluate an oral system designed to achieve site-specific and instant drug release in the colon for effective treatment of IBD.

Materials and methods: The system consists of a core tablet containing the model drug diclofenac sodium, superdisintegrant sodium starch glycolate, and coated with enteric polymer Eudragit FS 30 D to achieve different total percentage weight gain. Drug release studies were carried out using a changing pH method. A placebo formulation containing barium sulphate in the tablet was administered to human volunteers for in vivo X-ray studies. SEM studies were performed to determine coating thickness and film topography.

Results: In vitro studies revealed that the tablet with 10% coating level released the drug after 5?h lag time corresponding to the colonic region. Tablets with 10% coating level could maintain their integrity in human volunteers for 5?h, approximating colon arrival time and release the drug instantaneously.

Discussion: Colon-targeting and instant drug release for 10% coating level was due to the dissolution of the Eudragit FS 30 D and the immediate release effect of superdisintegrant. It was observed that as the coating level increased, the lag time also increased. This was because of increased diffusion path length and tortuosity at higher coating levels.

Conclusion: An in vivo-in vitro study revealed that not only the sensitivity of the polymer to the pH environment but also the thickness of coating plays an important role in colon delivery and the tablet with 5% superdisintegrant and 10% coating level achieved the desired performance of the colon targeting.     

Enhanced oral bioavailability of insulin using PLGA nanoparticles co-modified with cell-penetrating peptides and Engrailed secretion peptide (Sec)

Abstract

Biodegradable polymer nanoparticle drug carriers are an attractive strategy for oral delivery of peptide and protein drugs. However, their ability to cross the intestinal epithelium membrane is largely limited. Therefore, in the present study, cell-penetrating peptides (R8, Tat, penetratin) and a secretion peptide (Sec) with N-terminal stearylation were introduced to modify nanoparticles (NPs) on the surface to improve oral bioavailability of peptide and protein drugs. In vitro studies conducted in Caco-2 cells showed the value of the apparent permeability coefficient (P_app) of the nanoparticles co-modified with Sec and penetratin (Sec-Pen-NPs) was about two-times greater than that of the nanoparticles modified with only penetratin (Pen-NPs), while the increase of transcellular transport of nanoparticles modified together with Sec and R8 (Sec-R8-NPs), or Sec and Tat (Sec-Tat-NPs), was not significant compared with nanoparticles modified with only R8 (R8-NPs) or Tat (Tat-NPs). Using insulin as the model drug, in vivo studies performed on rats indicated that compared to Pen-NPs, the relative bioavailability of insulin for Sec-Pen-NPs was 1.71-times increased after ileal segments administration, and stronger hypoglycemic effects was also observed. Therefore, the nanoparticles co-modified with penetratin and Sec could act as attractive carriers for oral delivery of insulin.     

Bioavailability enhancement,Caco-2 cells uptake and intestinal transport of orally administered lopinavir-loaded PLGA nanoparticles

Nanoparticles (NPs) can be absorbed via M cells of Peyer’s patches after oral delivery leading to passive lymphatic targeting followed by systemic drug delivery. Hence, the study was aimed to formulate PLGA NPs of lopinavir. The NPs were prepared by nanoprecipitation, optimized by 3³ factorial design and characterized by TEM, DSC, FTIR studies and safety was assessed by MTT assay. In vivo pharmacokinetic studies were performed in rats. The NPs were discrete spherical structures having particle size of 142.1?±?2.13?nm and entrapment of 93.03?±?1.27%. There was absence of drug-polymer interaction. Confocal images revealed the penetration and absorption of coumarin-loaded NPs in Caco-2 cells and intestine after oral delivery. There was 3.04 folds permeability and 13.9 folds bioavailability enhancement from NPs. The NPs can be promising delivery system for antiretroviral drug by delivering the drug to lymph (major HIV reservoir site) via direct absorption through intestine before reaching systemic circulation.     

Cyclodextrins in the production of large porous particles: Development of dry powders for the sustained release of insulin to the lungs

The aim of this work was to develop dry powders intended for insulin pulmonary delivery. To this purpose, large porous particles (LPP) made of poly(lactide-co-glycolide) (PLGA) were produced by the double emulsion-solvent evaporation technique. Hydroxypropyl-beta-cyclodextrin (HPbetaCD), also known as absorption enhancer for pulmonary protein delivery, was tested as aid excipient to optimize the aerodynamic behaviour of the microparticles. Several microsphere formulations, differing in HPbetaCD and insulin loadings, were produced and their properties compared. A contemporary release of insulin and HPbetaCD from the system can be achieved by selecting appropriate formulation conditions. HPbetaCD-containing LPP with flow properties and dimensions suitable for aerosolization and deposition in deep regions of the lung following inhalation were produced. In conclusion, the developed system turns to be of great potential for the combined delivery of the protein and the adsorption promoter in the respiratory tract.     

Evaluation of hydrophobic nanoparticulate delivery system for insulin

Insulin loaded hydrophobic nanoparticles were prepared by solvent diffusion followed by lyophilization. Nanoparticles were characterized for mean size by dynamic laser scattering and for shape by scanning electron microscopy. Insulin encapsulation efficiency, in vitro stability of nanoparticles in presence of proteolytic enzymes and in vitro release were determined by high pressure liquid chromatography analysis. The biological activity insulin from the nanopraticles was estimated by enzyme-linked immunosorbant assay and in vivo using Wister diabetic rats. Nanoparticles ranged 0.526±0.071 μm in diameter. Insulin encapsulation efficiency was 95.7±1.2%. Insulin hydrophobic nanoparticles suppressed insulin release promoted sustained release in pH 7.4 phosphate buffer and shown to protect insulin from enzymatic degradation in vitro in presence of chymotripsin. Nanoencapsulated insulin was bioactive, demonstrated through both in vivo and in vitro.     

诺和锐30对初诊2型糖尿病的疗效观察

目的观察每日3次诺和锐30对初诊2型糖尿病的疗效。方法选择许昌市中心医院2009年10月至2010年10月新诊断的T2DM住院患者(诊断按WTO1999年标准)60例,随机分为MDI组及诺和锐30组治疗两周,比较PG、c肽、血糖达标时间、胰岛素日用量和低血糖发生人数。结果两组组内比较均有明显改善,差异具有统计学意义(P<0.01)。两组间无统计学意义(P>0.05)。结论每日3次诺和锐30能有效降低血糖,改善β细胞功能,有效降低低血糖事件的发生,可作为初诊2型糖尿病的强化治疗方案。     

An investigation into the characteristics of chitosan/Kollicoat SR30D free films for colonic drug delivery

The purpose of the study was to establish the physico-mechanical, digestibility, permeability and swelling properties of chitosan/Kollicoat SR30D films as potential coatings for colonic drug delivery. Free films containing different ratios of chitosan to Kollicoat SR30D were prepared by casting/solvent evaporation method. The resultant mixed films were characterized in terms of puncture strength and elongation (%), glass transition temperature, swellability, polymer miscibility, permeability, and digestibility under different media. The mixed films possessed good mechanical properties, which could be used as film-coating materials for drug delivery. The extent of digestion was directly proportional to the amount of chitosan present within the film. No apparent miscibility was detected between the chitosan and Kollicoat SR30D, regardless of the film composition. The films were found to be susceptible to digestion by bacterial or β-glucosidase enzymes in simulated colonic fluid (SCF). The SCF with rat cecal bacterial enzymes had a more profound hydrolytic activity than that with β-glucosidase enzyme for the digestion of chitosan within the mixed films. Overall, the results indicated that such chitosan/Kollicoat SR30D films had potential as a coating system for drug delivery to the colon.     

Development of smart delivery system for ascorbic acid using pH-responsive P(MAA-co-EGMA) hydrogel microparticles

pH-Responsive P(MAA-co-EGMA) hydrogel microparticles were prepared and their feasibility as intelligent delivery carriers was evaluated. P(MAA-co-EGMA) hydrogel microparticles were synthesized via dispersion photopolymerization. There was a drastic change in the swelling ratio of P(MAA-co-EGMA) microparticles at a pH of ~ 5 and, as the amount of MAA in the hydrogel increased, the swelling ratio increased at a pH above 5. The loading efficiency of the ascorbic acid into the hydrogel was affected more by the degree of swelling of the hydrogel than the electrostatic interaction between the hydrogel and the loaded ascorbic acid. The P(MAA-co-EGMA) hydrogel microparticles showed a pH-sensitive release behavior. Thus, at pH 4 almost none of the ascorbic acid permeated through the skin while at pH 6 relatively high skin permeability was obtained. The ascorbic acid loaded in the hydrogel particles was hardly degraded and its stability was maintained at high temperature.     

黄杨宁口腔速崩片人体药代动力学及相对生物利用度的研究

目的：研究黄杨宁口腔速崩片的人体相对生物利用度和生物等效性。方法：22名健康男性受试者，随机分组双交叉给予黄杨宁口腔速崩片（受试制剂）和黄杨宁片（参比制剂），剂量均为2mg，间隔为2周。分别于给药后120h内多点抽取静脉血，血浆样品经固相萃取（SPE）后用液质联用技术测定其中环维黄杨星D浓度。DAS2．0药动学程序计算相对生物利用度并评价两者的生物等效性。AUC0-120、AUC0-inf和Cmax经方差分析和双单侧t检验，tmax进行秩和检验。结果：单剂量给予受试制剂、参比制剂后血浆中的环维黄杨星D的Cmax分别为（121±55）、（122±52）ng／L；tmax分别为（10±7）、（12±8）h；AUC0-120分别为（4398±1656）、（4524±1760）ng·L^-1·h；AUC0→inf分别为（5292±2034）、（5440±2446）ng·L^-1·h。Cmax的90％可信区间为82．7％～117．3％；AUC－120的90％可信区间为83．9％。112．9％；AUC0→inf的90％可信区间为83．4％～117．6％。结论：受试制剂的人体相对生物利用度为（97．7±14．7）％，与参比制剂相比，两者具有生物等效性。