Long-term delivery enhances in vivo osteogenic efficacy of bone morphogenetic protein-2 compared to short-term delivery

In this study, heparin-conjugated poly(l-lactide-co-glycolide) (PLGA) nanospheres (HCPNs) suspended in fibrin gel (group 1) were developed for a long-term delivery of BMP-2, and then used to address the hypothesis that a long-term delivery of BMP-2 would enhance ectopic bone formation compared to a short-term delivery at an equivalent dose. Fibrin gel containing normal PLGA nanospheres (group 2) was used for short-term delivery of BMP-2. The in vitro release of BMP-2 from group 1 was sustained for 4 weeks with no initial burst release. In contrast, 83% of BMP-2 loaded in group 2 was released only for the first 3 days. BMP-2 released from group 1 stimulated an increase in alkaline phosphatase (ALP) activity of osteoblasts for 9 days in vitro. In contrast, BMP-2 released from group 2 induced a transient increase in ALP activity for the first 5 days and a decrease thereafter. Importantly, group 1 induced bone formation to a much greater extent than did group 2, with 2.0-fold greater bone formation area and 3.5-fold greater calcium content, upon implantation into rat hind limb muscle. These results show that long-term delivery of BMP-2 enhances in vivo osteogenic efficacy of the protein compared to short-term delivery at an equivalent dose.     

Biological activity of rhBMP-2 released from PLGA microspheres

Human recombinant bone morphogenetic protein-2 (rhBMP-2) has been proven effective in stimulating the regeneration of bone in both skeletal and extraskeletal locations. Through encapsulation within, and release from, biodegradable poly(DL-lactic-co-glycolic acid) (PLGA) microspheres, a proven vehicle for sustained delivery of various proteins, the local concentrations of rhBMP-2 could be maintained at optimal levels to stimulate bone regeneration and remodeling at the site of healing in diverse clinical settings. Thus the purpose of this work was to investigate the encapsulation of rhBMP-2 in PLGA microspheres and its biologic activity upon release. Using in vitro tests in simulated body fluids, the effect of rhBMP-2 released from PLGA microspheres upon osteoblast cell cultures was found to be statistically similar to the effect produced by positive controls consisting of nonencapsulated aqueous rhBMP-2 in simulated body fluids. This clarifies an important step in skeletal tissue engineering strategies aimed at the use of encapsulated rhBMP-2 to stimulate bone regeneration and remodeling.     

Preparation,in vitro release,in vivo absorption and biocompatibility studies of insulin-loaded microspheres in rabbits

The purpose of this study was to develop a single-dose insulin delivery system based on poly (lactide-co-glycolide) (PLGA) microspheres to provide basal insulin level for a prolonged period. Insulin-loaded PLGA microspheres were prepared by water-in-oil-in-water double emulsion (batch A) and solid-in-oil-in-water emulsion (batch B) methods. Microspheres were characterized for physical characteristics and in vitro release. In vivo absorption of insulin and biocompatibility of insulin-loaded PLGA microspheres were performed in diabetic New Zealand white rabbits. Light and transmission electron microscopy were performed on the skin tissues excised from microspheres injected sites in order to study the biocompatibility. The burst release of insulin was high (47%) from batch B and low (5%) from batch A. Therefore, we mixed microspheres of batch A and B in ratio of 3:1 w/w, which produced desirable in vitro release profile. In vivo absorption study showed that insulin-loaded microspheres provided a serum insulin level of 20-40 microU/ml up to 40 days. Biocompatibility study provided evidence of normal inflammatory and foreign body reactions, which were characterized by the presence of macrophages, fibroblasts and foreign body giant cells. Neither necrosis nor tissue damage was identified. At the end of 12 weeks, no distinct histological differences were observed in comparison to the control tissue samples. In conclusion, insulin-loaded PLGA microspheres controlled the in vivo absorption of insulin to maintain the basal insulin level for longer period and the delivery system was biocompatible.     

Effect of a freeze-dried CMC/PLGA microsphere matrix of rhBMP-2 on bone healing

The hypothesis of this research was that implants of poly(lactide-co-glycolide) (PLGA) microspheres loaded with bone morphogenetic protein-2 (rhBMP-2) and distributed in a freeze-dried carboxymethylcellulose (CMC) matrix would produce more new bone than would matrix implants of non-protein-loaded microspheres or matrix implants of only CMC. To test this hypothesis it was necessary to fashion microsphere-loaded CMC implants that were simple to insert, fit precisely into a defect, and would not elicit swelling. Microspheres were produced via a water-in-oil-in-water double-emulsion system and were loaded with rhBMP-2 by soaking them in a buffered solution of the protein at a concentration of 5.4 mg protein per gram of PLGA. Following recovery of the loaded microspheres by lyophilization matrices for implantation were prepared by lyophilizing a suspension of the microspheres in 2% CMC in flat-bottom tissue culture plates. Similar matrices were made with 2% CMC and with 2% CMC containing blank microspheres. A full-thickness calvarial defect model in New Zealand white rabbits was used to assess bone growth. Implants fit the defect well allowing for direct application. Six weeks postsurgery, defects were collected and processed for undecalcified histology. In vitro, 60% of the loaded rhBMP-2 released from devices or microspheres in 5 to 7 days. With the unembedded microspheres releasing faster than those embedded in CMC In vivo. the rhBMP-2 microspheres greatly enhanced bone healing, whereas nonloaded PLGA microspheres in the CMC implants had little effect. The results showed that a lyophilized device of rhBMP-2 PLGA microspheres in CMC was an effective implantable protein-delivery system for the use in bone repair. Published: October 7. 2001.     

Repair effect of diabetic ulcers with recombinant human epidermal growth factor loaded by sustained-release microspheres

In this study the w/o/w extraction-evaporation technique was adopted to prepare poly(lactic-co-glycolic acid) (PLGA) microspheres loading recombinant human epidermal growth factor (rhEGF). The microspheres were characterized for morphology by transmission electron microscopy (TEM) and particle size distribution. The release performances, the proliferation effects and therapeutic effects of rhEGF-loaded PLGA microspheres were all studied. The results showed that these spherical microspheres had a narrow size distribution and a high drug encapsulation efficiency (85.6%). RhEGF-loaded microspheres enhanced the growth rate of fibroblasts and wound healing more efficiently than pure rhEGF. The number of the proliferating cell nuclear antigen (PCNA) in the epidermis layer with the microsphere treatment was significantly larger than those of the control groups. Overall locally sustained delivery of rhEGF from biodegradable PLGA microspheres may serve as a novel therapeutic strategy for diabetic ulcer repair.     

Preparation and quality by design assisted (Qb-d) optimization of bioceramic loaded microspheres for periodontal delivery of doxycycline hyclate

PLGA (Lactic- co-glycolic acid) coated chitosan microspheres loaded with hydroxyapatite and doxycycline hyclate complex were developed in the present study for periodontal delivery. A modified single emulsion method was adopted for the development of microspheres. Formulation was optimized on the basis of particle size, drug loading and encapsulation efficiency with the central composite design using 2³ factorial design. Microspheres were optimized and electron microscopy revealed their spherical shape and porous nature. In-vitro study showed initial burst and then sustained release behavior of the formulation for 14 days. Further, in-vitro antibacterial study performed on E. coli (ATCC-25922) and S. aureus (ATCC-29213) revealed concentration dependent activity. Also, in-vitro cyto-toxicity assessment ensures biocompatibility of the formulation with the fibroblast’s cells. Overall, the quality by design assisted PLGA microspheres, demonstrated the desired attributes and were found suitable for periodontal drug delivery.     

Repair effect of diabetic ulcers with recombinant human epidermal growth factor loaded by sustained-release microspheres

In this study the w/o/w extraction–evaporation technique was adopted to prepare poly(lactic-co-glycolic acid) (PLGA) microspheres loading recombinant human epidermal growth factor (rhEGF). The micro-spheres were characterized for morphology by transmission electron microscopy (TEM) and particle size distribution. The release performances, the proliferation effects and therapeutic effects of rhEGF-loaded PLGA microspheres were all studied. The results showed that these spherical micro-spheres had a narrow size distribution and a high drug encapsulation efficiency (85.6%). RhEGF-loaded microspheres enhanced the growth rate of fibroblasts and wound healing more efficiently than pure rhEGF. The number of the proliferating cell nuclear antigen (PCNA) in the epidermis layer with the mi-crosphere treatment was significantly larger than those of the control groups. Overall locally sustained delivery of rhEGF from biodegradable PLGA microspheres may serve as a novel therapeutic strategy for diabetic ulcer repair.     

De novo adipose tissue generation through long-term, local delivery of insulin and insulin-like growth factor-1 by PLGA/PEG microspheres in an in vivo rat model: a novel concept and capability

This study was undertaken to characterize the duration of long-term growth factor delivery by poly(lactic-co-glycolic-acid)-polyethylene glycol (PLGA/PEG) microspheres and to evaluate the potential of long-term delivery of insulin and insulin-like growth factor-1 (IGF-1) for the de novo generation of adipose tissue in vivo. PLGA/PEG microspheres containing insulin and IGF-1, separately, were produced by a double-emulsion solvent-extraction technique. In the first phase of the experiment, the in vitro release kinetics of the microspheres were evaluated for the optical density and polyacrylamide gel electrophoresis of solutions incubated with insulin-containing microspheres for four different periods of time (n = 1). The finding of increased concentrations of soluble insulin with increased incubation time confirmed continual protein release. In the second stage of the experiment, 16 rats were divided equally into four study groups (insulin, IGF-1, insulin + IGF-1, and blank microspheres) (n = 4). Insulin and IGF-1 containing microspheres were administered directly to the deep muscular fascia of the rat abdominal wall to evaluate the potential for de novo adipose tissue generation via adipogenic differentiation from native nonadipocyte cell pools in vivo. Animals treated with blank microspheres served as an external control group. At the 4-week harvest period, multiple ectopic islands of adipose tissue were observed on the abdominal wall of the animals treated with insulin, IGF-1, and insulin + IGF-1 microspheres. Such islands were not seen in the blank microsphere group. Hematoxylin and eosin-stained sections of the growth factor groups demonstrated mature adipocytes interspersed with fibrous tissue superficial to the abdominal wall musculature and continuous with the fascia. Oil-Red-O stained sections demonstrated that these cells contained lipid. Computer-aided image analysis of histologic sections confirmed that there were statistically significant increases in the amount of "ectopic" adipose neotissue developed on the abdominal wall of animals treated with growth factor microspheres. In conclusion, this study confirms the long-term release of proteins from PLGA/PEG microspheres up to 4 weeks and demonstrates the potential of long-term local insulin and IGF-1 to induce adipogenic differentiation to mature lipid-containing adipocytes from nonadipocyte cell pools in vivo at 4 weeks.     

Alginate-PEG sponge architecture and role in the design of insulin release dressings

Wound healing is a natural process involving several signaling molecules and cell types over a significant period of time. Although current dressings help to protect the wound from debris or infection, they do little in accelerating the healing process. Insulin has been shown to stimulate the healing of damaged skin. We have developed an alginate sponge dressing (ASD) that forms a hydrogel capable of providing a moist and protective healing environment. By incorporating insulin-loaded poly(d,l-lactide-co-glycolide) (PLGA) microparticles into ASD, we successfully stabilized and released insulin for up to 21 days. Insulin release and water absorption and transfer through the ASD were influenced by altering the levels of poly(ethylene glycol) (PEG) in the dressing matrix. Bioactivity of released insulin can be maintained for at least 10 days, demonstrated using a human keratinocyte migration assay. Results showed that insulin-loaded PLGA microparticles, embedded within PEG-ASD, functioned as an effective long-term delivery platform for bioactive insulin.     

Porous bone morphogenetic protein-2 microspheres: Polymer binding and in vitro release

This research compared the binding and release of recombinant human bone morphogenetic protein 2 (rhBMP-2) with a series of hydrophobic and hydrophilic poly-lactide-co-glycolide (PLGA) copolymers. Porous microspheres were produced via a double emulsion process. Binding and incorporation of protein were achieved by soaking microspheres in buffered protein solutions, filtering, and comparing protein concentration remaining to nonmicrosphere-containing samples. Protein release was determined by soaking bound microspheres in a physiological buffer and measuring protein concentration (by reversed-phase high-performance liquid chromatography) in solution over time. Normalized for specific surface area and paired by polymer molecular weight. microspheres made from hydrophilic 50∶50 or 75∶25 PLGA bound significantly more protein than microspheres made from the corresponding hydrophobic PLGA. Increased binding capacity correlated with higher polymer acid values. With certain polymers, rhBMP-2 adsorption was decreased or inhibited at high protein concentration, but protein loading could be enhanced by increasing the protein solution:PLGA (volume:mass) ratio or by repetitive soaking. Microspheres of various PLGAs released unbound protein in 3 days, whereas the subsequent bound protein release corresponded to mass loss. RhBMP-2 binding to PLGA was controlled by the acid value, protein concentration, and adsorption technique. The protein released in 2 phases: the first occurred over 3 days regardless of PLGA used and emanated from unbound, incorporated protein, while the second was controlled by mass loss and therefore was dependent on the polymer molecular weight. Overall, control of rhBMP-2 delivery is achievable by selection of PLGA microsphere carriers. Published: October, 7, 2001.     

Novel biodegradable and pH-sensitive poly(ester amide) microspheres for oral insulin delivery

Biodegradable and pH-sensitive PEAs based on dual amino acids are designed, synthesized, and characterized. Insulin can be loaded into the PEA microspheres by a solid-in-oil-in-oil technique with high encapsulation efficiency. The feasibility of PEA microspheres as oral insulin delivery carriers is evaluated in vitro and in vivo. The hydrophobic leucine groups on PEA seem to play an important role in the pH-dependent release mechanism and cytotoxicity of PEA microspheres. Oral administration of insulin-loaded PEA microspheres to streptozotocin-induced diabetic rats at 60 IU kg(-1) is able to reduce fasting plasma glucose levels to 49.4%. These results indicate that PEA microspheres are potential new vehicles for insulin oral delivery.     

重组人粒细胞集落刺激因子缓释微球的研究

目的：研究固体／油／水法制备重组人粒细胞集落刺激因子缓释微球,为开发其缓释剂型进行初步研究。方法：以聚乳酸．聚羟乙酸共聚物（PLGA）为载体材料：用固体／油／水法和水／油／水法制备载rhG-CSF缓释微球;考察粒径大小、外观、包封率等理化性质;用MieroBCA法考察微球的体外释药特性及影响因素;用SEC-HPLC和MTT比色法初步评价了微球制备工艺过程对rhG-CSF稳定性的影响。结果：两种方法制得的微球形态圆整、分散性良好,包封率均超过80％。固／油／水法制得的微球体外释放在2周内可超过90％,而水／油／水法制得的微球在相同的时间内仅释放30％。对于固／油／水法制备过程,SEC-HPLC法测定蛋白无明显聚集体出现,MTT法测定蛋白活性无明显损失。结论：实验证明了固／油／水法制备的PLGA微球可以实现2周以上的体外缓释。     

Optimized bone regeneration based on sustained release from three-dimensional fibrous PLGA/HAp composite scaffolds loaded with BMP-2

Contemporary treatment of critical bone defect remains a significant challenge in the field of orthopedic surgery. Engineered biomaterials combined with growth factors have emerged as a new treatment alternative in bone repair and regeneration. Our approach is to encapsulate bone morphogenetic protein-2 (BMP-2) into a polymeric matrix in different ways and characterize their individual performance in a nude mouse model. The main objective of this study is to examine whether the PLGA/HAp composite fibrous scaffolds loaded with BMP-2 through electrospinning can improve bone regeneration. The hypothesis is that different loading methods of BMP-2 and different HAp contents in scaffolds can alternate the release profiles of BMP-2 in vivo, therefore modify the performance of scaffolds in bone regeneration. Firstly, mechanical strength of scaffolds and HAp nanoparticles distribution in scaffolds were investigated. Secondly, nude mice experiments extended to 6 weeks were carried out to test the in vivo performance of these scaffolds, in which measurements, like serum BMP-2 concentration, ALP activity, X-ray qualification, and H&E/IHC tissue staining were utilized to monitor the growth of new bone and the changes of the corresponding biochemical parameters. The results showed that the PLGA/HAp composite scaffolds developed in this study exhibited good morphology/mechanical strength and HAp nanoparticles were homogeneously dispersed inside PLGA matrix. Results from the animal experiments indicate that the bioactivity of BMP-2 released from the fibrous PLGA/HAp composite scaffolds is well maintained, which further improves the formation of new bone and the healing of segmental defects in vivo. It is concluded that BMP-2 loaded PLGA/HAp composite scaffolds are promising for bone healing.     

Delivery of Bioactive Lipids from Composite Microgel-Microsphere Injectable Scaffolds Enhances Stem Cell Recruitment and Skeletal Repair

In this study, a microgel composed of chitosan and inorganic phosphates was used to deliver poly(lactic-co-glycolic acid) (PLAGA) microspheres loaded with sphingolipid growth factor FTY720 to critical size cranial defects in Sprague Dawley rats. We show that sustained release of FTY720 from injected microspheres used alone or in combination with recombinant human bone morphogenic protein-2 (rhBMP2) improves defect vascularization and bone formation in the presence and absence of rhBMP2 as evaluated by quantitative microCT and histological measurements. Moreover, sustained delivery of FTY720 from PLAGA and local targeting of sphingosine 1-phosphate (S1P) receptors reduces CD45+ inflammatory cell infiltration, promotes endogenous recruitment of CD29+CD90+ bone progenitor cells and enhances the efficacy of rhBMP2 from chitosan microgels. Companion in vitro studies suggest that selective activation of sphingosine receptor subtype-3 (S1P₃) via FTY720 treatment induces smad-1 phosphorylation in bone-marrow stromal cells. Additionally, FTY720 enhances stromal cell-derived factor-1 (SDF-1) mediated chemotaxis of CD90+CD11B-CD45- bone progenitor cells in vitro after stimulation with rhBMP2. We believe that use of such small molecule delivery formulations to recruit endogenous bone progenitors may be an attractive alternative to exogenous cell-based therapy.     

Controlled release of modified insulin glargine from novel biodegradable injectable gels

The objective of this study was to investigate the duration of biological effects of modified insulin glargine released from a novel biodegradable injectable gel in type II diabetic Zucker diabetic fatty (ZDF) rats. Modified insulin glargine was purified from the marketed formulation by process of dialysis followed by freeze-drying, and the purity was confirmed by the single peak, corresponding to insulin glargine in the HPLC chromatogram. To determine and to compare the biological activity of purified insulin glargine with marketed formulation, it was suspended in isotonic saline solutions and administered subcutaneously to ZDF rats at a dose of 10 IU/kg of insulin and the blood glucose levels were measured. The blood glucose levels of ZDF rats after a subcutaneous injection of a suspension of purified insulin glargine decreased below 200 mg/dL within 2 h and remained at this level up to 6 h, then steadily raised above 400 mg/dL in 12 h. Insulin glargine particles were loaded into a novel biodegradable injectable gel formulation prepared from a blend of polylactic-co-glycolic acid (PLGA) and biocompatible plasticizers. Approximately 0.1 mL of insulin glargine-loaded gel prepared with PLGA was administered subcutaneously to the ZDF rats, and blood glucose levels were measured. The PLGA gel formulations prepared with insulin glargine particles had duration of action of 10 days following a single subcutaneous injection. The addition of zinc sulfate to the formulations prepared with purified insulin glargine particles further slowed down the drop in blood glucose concentrations.     

Synthesis and characterization of hydroxyapatite-ciprofloxacin delivery systems by precipitation and spray drying technique

This investigation synthesized and characterized hydroxyapatite (HAP) microspheres, agglomerated microspheres, and implants containing ciprofloxacin. This delivery system is to be used as an implantable drug delivery system for the treatment of bone infections. The HAP microspheres were made by chemical precipitation followed by a spray-drying technique. Agglomerated microspheres were prepared by a wet granulation process using a granulator. Implants were prepared by direct compression of the granules on a Carver press. Ciprofloxacin was analyzed by high-performance liquid chromatography. Characterization of the HAP microspheres include particle size, size distribution, physical state of the drug in the microsphere, and microstructure of the drug delivery system before and after in vitro release. The particle size, porosity, and morphology of the microspheres were dependent on viscosity and concentration of the slurry as well as the atomization pressure used during spray drying. Even at the highest drug load (2% wt/wt), the drug was present in a noncrystalline state. The drug release from the agglomerated microspheres was quick and almost complete within 1 hour. However, compressing the same amount of agglomerated microspheres into an implant greatly reduced the rate of ciprofloxacin release. Only 12% (wt/wt) of the drug was released from the implant within 1 hour. The in vitro release of ciprofloxacin from these implants follows a diffusion-controlled mechanism. This method provides a unique way of producing various shapes and drug loads of HAP microspheres that can be easily manufactured on a commercial scale. Published: January 28, 2002.     

New route for synthesizing poly(ethylene glycol)-acrylic acid hydrogels using γ-irradiation for drug delivery carriers

Biocompatible and pH-responsive poly(ethylene glycol) (PEG)-acrylic acid (AAc) hydrogels were prepared by new technique using γ-irradiation for controlled oral drug delivery. The gel fraction was over 80% and the equal amounts of PEG and AAc blended hydrogel had efficient insulin loading using equilibrium swelling. These hydrogels exhibited unique pH-responsive characteristics in which interpolymer complexes were formed in acidic media and dissociated in neutral or basic environments. The insulin release from the gel was significantly retarded in acidic media while rapid release occurred under neutral/basic conditions. At the high pH solution, the gels swelled rapidly and over 70% of the insulin loaded was released over a period of 10 h. Within 2 h of administration of the insulin-containing gels, significant blood glucose reduction effects were observed in diabetic rats. The blood glucose reduction lasted for up to 10 h following administration.     

Preparation and characterization of poly(D,L-lactide-co-glycolide) microspheres for controlled release of human growth hormone

The purpose of this research was to assess the physicochemical properties of a controlled release formulation of recombinant human growth hormone (rHGH) encapsulated in poly(D,L-lactide-co-glycolide) (PLGA) composite microspheres. rHGH was loaded in poly(acryloyl hydroxyethyl) starch (acHES) microparticles, and then the protein-containing microparticles were encapsulated in the PLGA matrix by a solvent extraction/evaporation method. rHGH-loaded PLGA microspheres were also prepared using mannitol without the starch hydrogel microparticle microspheres for comparison. The detection of secondary structure changes in protein was investigated by using a Fourier Transfer Infrared (FTIR) technique. The composite microspheres were spherical in shape (44.6±2.47 μm), and the PLGA-mannitol microspheres were 39.7±2.50 μm. Drug-loading efficiency varied from 93.2% to 104%. The composite microspheres showed higher overall drug release than the PLGA/mannitol microspheres. FTIR analyses indicated good stability and structural integrity of HGH localized in the microspheres. The PLGA-acHES composite microsphere system could be useful for the controlled delivery of protein drugs.     

Formulation and in vitro transfection efficiency of poly (D, L-lactideco-glycolide) microspheres containing plasmid DNA for gene delivery

The stability, in vitro release, and in vitro cell transfection efficiency of plasmid DNA (pDNA) poly (D,L.-lactide-co-glycolide) (PLGA) microsphere formulations were investigated. PLGA microspheres containing free and polylysine (PLL)-complexed pDNA were prepared by a water-oil-water solvent extraction/evaporation technique. Encapsulation enhanced the retention of the supereoiled structure of pDNA as determined by gel electrophoresis. PLL complexation of pDNA prior to encapsulation increased both the stability of the supercoiled form and the encapsulation efficiency. Free pDNA was completely degraded after exposure to DNase while encapsulation protected the pDNA from enzymatic degradation. Rapid initial in vitro release of pDNA was obtained from microspheres containing free pDNA. while the release from microspheres containing PLL-complexed pDNA was sustained for more than 42 days. Bioactivity of encapsulated pDNA determined by in vitro cell transfection using Chinese hamster ovary cells (CHO) showed that the bioactivity of encapsulated pDNA was retained in both formulations but to a greater extent with PLL-complexed pDNA microspheres. These results demonstrated that PLGA microspheres could be used to formulate a controlledrelease delivery system for pDNA that can protect the pDNA from DNase degradation without loss of functional activity.     

Synthesis and characterization of alginate coated zinc calcium phosphate nanoparticles for intestinal delivery of insulin

Nanosized calcium phosphates studied as drug delivery systems are highly compatible with the various drugs like insulin, antibiotics etc. Zinc is an essential trace element that plays a crucial role in the synthesis, storage and release of insulin in a human body. Therefore, an attempt has been made to develop zinc modified calcium phosphate nanoparticles (less than 100 nm) as carriers for intestinal delivery of insulin. The insulin loaded nanoparticles were coated with pH sensitive alginate. These pH sensitive nanoparticles released insulin in the intestinal medium, and the conformation of released insulin was stable. The blood glucose level of diabetic rats came to normal on administration of the formulation. With the beneficial effect of zinc reported on diabetic patients, the present system seems to be an excellent carrier for intestinal delivery of insulin.