International Journal of Pharmaceutics (v.407, #1-2)

Why and how to prepare biodegradable, monodispersed, polymeric microparticles in the field of pharmacy? by Van-Thanh Tran; Jean-Pierre Benoît; Marie-Claire Venier-Julienne (1-11).
Drug delivery via biodegradable microparticles benefits from both the protection of the encapsulated drug from hazardous conditions and the controlled release of the encapsulated drug, thereby reducing the administration frequency and improving patient compliance. Microsphere-size particle distribution is considered as being an important factor that affects the choice of the administration route and the drug-release rate. Significant research efforts have been directed towards the production of monodispersed “designer” particles. Amongst various techniques, some have been examined from lab-scale to industrial-scale. This review provides a global overview of monodispersed microparticle production methods and then focuses on recent processes being used to produce biodegradable microparticles applied in the pharmaceutical field. Further discussion about the choice of process according to the microparticle objectives of use is suggested.
Keywords: Monodispersed microparticles; Biodegradable polymer; Controlled delivery; Pharmaceutical field;

Computational and experimental investigation of needle-shaped crystal breakage by Zdenĕk Grof; Carl M. Schoellhammer; Pavol Rajniak; František Štĕpánek (12-20).
Needle-shaped crystals are a common occurrence in many pharmaceutical and fine chemicals processes. Even if the particle size distribution (PSD) obtained in a crystallization step can be controlled by the crystal growth kinetics and hydrodynamic conditions, further fluid–solid separation steps such as filtration, filter washing, drying, and subsequent solids handling can often lead to uncontrolled changes in the PSD due to breakage. In this contribution we present a combined computational and experimental methodology for determining the breakage kernel and the daughter distribution functions of needle-shaped crystals, and for population balance modeling of their breakage. A discrete element model (DEM) of needle-shaped particle breakage was first used in order to find out the appropriate types of the breakage kernel and the daughter distribution functions. A population balance model of breakage was then formulated and used in conjunction with experimental data in order to determine the material-specific parameters appearing in the breakage functions. Quantitative agreement between simulation and experiment has been obtained.
Keywords: DEM; Population balance modeling; Fragmentation; Daughter length distribution; Breakage kernel;

Microparticle uptake in the small intestine is relevant to both the delivery of pharmaceutics and exposure to environmental pollutants. The Caco-2 enterocyte model is a useful tool to study the parameters that affect epithelial microparticle permeability and the mechanisms controlling them. The current study used this model to explore further the different effects of 10% ethanol v/v or ice on transepithelial resistance (TER), microparticle uptake and immunofluorescent labelling of intercellular junctions. The same exposure times for both treatments were used, rather than those shown in the literature to produce demonstrable changes induced by each. The effects of both pre-treatments were greater after 60 min than after 15 min. Ethanol pre-treatment for 60 min decreased TER, increased particle uptake and was associated with a disorganisation of tight and adhering junctional proteins. Pre-treatment with ice for 60 min however, increased TER, decreased particle uptake and was associated with concentration of intercellular junctional proteins in a more constrained manner. These findings on the effects of pre-treatment with ethanol or ice for 60 min suggest that the extent of uptake is influenced by changes in the distribution of intercellular junctional proteins.
Keywords: Caco-2; Latex microparticle uptake; Ethanol; Low temperature; Transepithelial resistance (TER); Occludin; ZO-1; E-cadherin;

Protamine modified metal ion–protein chelate microparticles for sustained release of interferon by Yanbo Jiang; Kai Shi; Dengning Xia; Hongze Piao; Peng Quan; Tao Song; Fude Cui (31-37).
This study focuses on extending the release period of zinc–protein chelate through protamine modification. Recombinant human interferon-α-2b (rhIFN), a highly pleiotropic cytokine with a short intrinsic pharmacokinetic half-life when injected subcutaneously (∼2–6 h), was used as a model drug. Protamine modified zinc–rhIFN chelate microparticles were prepared by co-precipitating rhIFN with zinc and protamine. Introduction of protamine (2.5–20 mg/mL) into the chelation system had several prominent effects. First, percentage of chelated rhIFN was lowered (from >99% to ∼90%); second, particle size was gradually increased (from ∼0.45 μm to ∼2 μm); last but important, it extended the release period of the chelate both in vitro (complete release was retarded from 8 h to 48 h) and in vivo (t 1/2 was prolonged from 4.5 h to 15.5 h and mean residence time from 9.4 h to 29.6 h). Size-exclusion liquid chromatography and cytopathic effect inhibition assay indicated rhIFN preserved its structural and functional integrity in these chelates.
Keywords: Interferon; Zinc; Protamine; Sustained release; In vitro release; Pharmacokinetics;

Development and in vitroin vivo relationship of controlled-release microparticles loaded with tramadol hydrochloride by Muhammad Naeem Aamir; Mahmood Ahmad; Naveed Akhtar; Ghulam Murtaza; Shujaat Ali Khan; Shahiq-uz-Zaman; Ali Nokhodchi (38-43).
In conclusion, the controlled-release microparticles of TmH can be developed via phase separation method. The development and optimization of controlled-release microparticles of tramadol hydrochloride (TmH) for the oral delivery and their in vitro and in vivo correlation was prime objective of the present study. Four formulations of controlled-released microparticles were developed and optimized in terms of encapsulation efficiency, dissolution study and release kinetics. Among all formulated microparticles F-3 (ratio of TmH:EC 1:2) and F-4 (ratio of TmH:EC 1:3) presented the better characteristics in reference to entrapment efficiency, release kinetics and dissolution profile compared to other formulations (F-1, F-2). For in vivo analysis a new HPLC analytical method was developed and validated. The optimized formulations were subjected to in vivo studies to calculate various pharmacokinetic parameters, i.e., C max, t max, AUC0–∞ and MRT. The in vitro dissolution and in vivo absorption data was correlated with the help of Wagner–Nelson method. F-3 showed a good in vitro–in vivo correlation with a correlation determination of 0.9957. Moreover, lower T max, t 1/2 and MRT, and higher values of C max and K e were observed for F-3. The control formulation (immediate-release) presented lowest values of t 1/2, MRT and T max but the highest values of C max and K e. The controlled-release microparticles (F-3 and F-4) could sustain the drug release within therapeutic level up to 24 h and good IVIVC is expected from them.
Keywords: Controlled-release microparticles; HPLC method; Ethyl cellulose; Tramadol; In vitroin vivo relationship;

Sustained-release of protein from biodegradable sericin film, gel and sponge by Ayumu Nishida; Masaki Yamada; Takanori Kanazawa; Yuuki Takashima; Kiyohisa Ouchi; Hiroaki Okada (44-52).
The film, gel and sponge forms of silk protein, sericin, were examined to determine the characteristics of sustained release of a drug protein model and the biodegradability.A silk protein, sericin, contains 18 kinds of amino acids, mostly polar side chains forming a complex of three principal polypeptides. The major polypeptides exhibit hydrophobic characteristics by forming a β-sheet structure in a hydrate state. As a drug-releasing biomaterial made by an aqueous process without using any cross linker, sericin is expected to form various hydrophobic dosage forms. However, its dosage form, with respect to the molecular weight and concentration of sericin, and its biodegradation behavior has not been studied in detail. In this study, the film, gel and sponge of sericin were prepared and examined to determine the release properties of the charged protein, fluorescein isothiocyanate–albumin (FA). The film and gel, as solid and semisolid forms, respectively, were also evaluated for their biodegradation behavior. For in vitro release, FA was sustained-released from these preparations. The concentration and dosage form markedly affected FA release. For in vivo biodegradation, the sericin preparations implanted subcutaneously in rats gradually decreased in size and weight. Histological examination indicated no marked inflammation at the site. As for in vivo release, FA remained for 3–6 weeks or more in rats. These findings suggest that sericin is suitable for use as a drug-releasing biomaterial.
Keywords: Sericin; Sustained release; Film; Gel; Sponge; Biodegradable;

Time-oriented experimental design method to optimize hydrophilic matrix formulations with gelation kinetics and drug release profiles by Sangmun Shin; Du Hyung Choi; Nguyen Khoa Viet Truong; Nam Ah Kim; Kyung Rok Chu; Seong Hoon Jeong (53-62).
A new experimental design methodology was developed by integrating the response surface methodology and the time series modeling. The major purposes were to identify significant factors in determining swelling and release rate from matrix tablets and their relative factor levels for optimizing the experimental responses. Properties of tablet swelling and drug release were assessed with ten factors and two default factors, a hydrophilic model drug (terazosin) and magnesium stearate, and compared with target values. The selected input control factors were arranged in a mixture simplex lattice design with 21 experimental runs. The obtained optimal settings for gelation were PEO, LH-11, Syloid, and Pharmacoat with weight ratios of 215.33 (88.50%), 5.68 (2.33%), 19.27 (7.92%), and 3.04 (1.25%), respectively. The optimal settings for drug release were PEO and citric acid with weight ratios of 191.99 (78.91%) and 51.32 (21.09%), respectively. Based on the results of matrix swelling and drug release, the optimal solutions, target values, and validation experiment results over time were similar and showed consistent patterns with very small biases. The experimental design methodology could be a very promising experimental design method to obtain maximum information with limited time and resources. It could also be very useful in formulation studies by providing a systematic and reliable screening method to characterize significant factors in the sustained release matrix tablet.
Keywords: Matrix tablet; Robust design; Response surface methodology; Drug releases; swelling;

Hansen solubility parameter as a tool to predict cocrystal formation by Mohammad Amin Mohammad; Amjad Alhalaweh; Sitaram P. Velaga (63-71).
The objective of this study was to investigate whether the miscibility of a drug and coformer, as predicted by Hansen solubility parameters (HSPs), can indicate cocrystal formation and guide cocrystal screening. It was also our aim to evaluate various HSPs-based approaches in miscibility prediction. HSPs for indomethacin (the model drug) and over thirty coformers were calculated according to the group contribution method. Differences in the HSPs between indomethacin and each coformer were then calculated using three established approaches, and the miscibility was predicted. Subsequently, differential scanning calorimetry was used to investigate the experimental miscibility and cocrystal formation. The formation of cocrystals was also verified using liquid-assisted grinding. All except one of the drug-coformers that were predicted to be miscible were confirmed experimentally as miscible. All tested theoretical approaches were in agreement in predicting miscibility. All systems that formed cocrystals were miscible. Remarkably, two new cocrystals of indomethacin were discovered in this study. Though it may be necessary to test this approach in a wide range of different coformer and drug compound types for accurate generalizations, the trends with tested systems were clear and suggest that the drug and coformer should be miscible for cocrystal formation. Thus, predicting the miscibility of cocrystal components using solubility parameters can guide the selection of potential coformers prior to exhaustive cocrystal screening work.
Keywords: Hansen solubility parameters (HSPs); Predicting cocrystal formation; Cohesive energy density; Group contribution method; Miscibility;

Enhanced permeation of fentanyl from supersaturated solutions in a model membrane by P. Santos; A.C. Watkinson; J. Hadgraft; M.E. Lane (72-77).
Permeation fluxes of fentanyl through silicone membranes after application of saturated and supersaturated solutions, for finite doses of PG/Et (60:40) (▪), and finite dose (▪) and infinite dose (▪) conditions for PG/H2O (60:40).The aim of the present study was to investigate the permeation of fentanyl from supersaturated formulations when applied to silicone membrane. Silicone was chosen in order to separate the effects of supersaturation from other possible influences of volatile formulation components on biological membranes. Supersaturated formulations containing either propylene glycol/water (PG/H2O) or propylene glycol/ethanol (PG/Et) were prepared containing varying degrees of saturation (DS) of fentanyl. Permeation of finite and infinite doses of the PG/H2O formulations, and finite doses of the PG/Et formulations was investigated using Franz-type diffusion cells. For the PG/H2O formulations a good correlation between the flux and the DS of the formulation up to 5 DS for infinite dose studies (r 2  = 0.99), and up to 7 DS for finite dose studies (r 2  = 0.98), was evident. Similarly, for the PG/Et formulations there is a good correlation between the mean flux and the theoretical DS of the formulation (r 2  = 0.95). Except for the 2 DS formulations, no significant differences were seen in the mean flux between PG/H2O and PG/Et finite dose studies. The larger fluxes observed for infinite doses of the PG/H2O formulations versus finite doses reflect changes in the effective area of diffusion over the time of the experiment for the latter set of experiments. The permeation enhancement observed for PG/Et formulations confirms that enhanced drug thermodynamic activity was induced by ethanol evaporation.
Keywords: Supersaturation; Fentanyl; Permeation; Silicone; Propylene glycol; Ethanol;

Design and realization of drug delivery systems based on polymer matrices could be greatly improved by modeling the phenomena which take place after the systems administration. Availability of a reliable mathematical model, able to predict the release kinetic from drug delivery systems, could actually replace the resource-consuming trial-and-error procedures usually followed in the manufacture of these latter.In this work, the complex problem of drug release from polymer (HPMC) based matrices systems was faced. The phenomena, previously observed and experimentally quantified, of water up-take, system swelling and erosion, and drug release were here described by transient mass balances with diffusion. The resulting set of differential equations was solved by using finite element methods.Two different systems were investigated: cylindrical matrices in which the transport phenomena were allowed only by lateral surfaces (“radial” case), and cylindrical matrices with the overall surface exposed to the solvent (“overall” case).A code able to describe quantitatively all the observed phenomena has been obtained.
Keywords: Hydrogels; Swelling; Erosion; Drug release; Mathematical modeling;

Deposition of aerosols delivered by nasal route with jet and mesh nebulizers by Laurent Vecellio; Ruth De Gersem; Sandrine Le Guellec; Gregory Reychler; Laurent Pitance; Deborah Le Pennec; Patrice Diot; Gilles Chantrel; Pierre Bonfils; François Jamar (87-94).
To quantify the amount of aerosol deposited in different parts of the airways with a commercially available nasal sonic jet nebulizer (NJN) using a sound effect, and to compare its performance with a new nasal mesh nebulizer (NMN).Seven healthy non-smoking male volunteers aged 21–36 years with a mean weight of 77 ± 10 kg were included in this single-center study. Both nebulizer systems were loaded with 99mTc-DTPA and scintigraphies were performed with a gamma camera. Particle size distribution of the aerosols produced by the two nebulizer systems was measured.There was no statistical difference between the two nebulizers in terms of fraction of particles smaller than 5 μm (44 ± 4% vs 45 ± 2%) (p  > 0.9). Aerosol deposition in the nasal region was 73 ± 10% (% of aerosol deposited in airways) with the NJN, and 99 ± 3% with the NMN (p  = 0.01). Total nasal deposition was 9.6 ± 1.9% of the nebulizer charge with the NJN and 28.4 ± 8.9% with the NMN (p  = 0.01). 0.5 ± 0.3% of the nebulizer charge was deposited in the maxillary sinuses with the NJN, compared to 2.2 ± 1.6% with the NMN (p  = 0.01).Although the two nebulizers had the same particle size, NMN significantly improved aerosol deposition in nasal cavity and prevents deposition into the lungs.
Keywords: Nebulizer; Aerosol; Nasal; Deposition; Scintigraphy;

Optimization of microdermabrasion for controlled removal of stratum corneum by Samantha N. Andrews; Vladimir Zarnitsyn; Brian Bondy; Mark R. Prausnitz (95-104).
Microdermabrasion has been shown to increase skin permeability for transdermal drug delivery by damaging or removing skin's outer layer, stratum corneum. However, relationships between microdermabrasion parameters and effects on the stratum corneum barrier have not been developed. In this study, we determined the effect of microdermabrasion crystal flow rate, time, and suction pressure applied in both static and dynamic modes on the extent of stratum corneum removal from excised porcine skin. In addition to controlling the depth of tissue removal by microdermabrasion parameters, we also controlled the area of tissue removal by applying a metal mask patterned with 125- or 250-μm holes to selectively expose small spots of tissue to microdermabrasion. We found that the extent of stratum corneum removal depended strongly on the crystal flow rate and exposure time and only weakly on pressure or static/dynamic mode operation. Masking the skin was effective to localize stratum corneum removal to exposed sites. Overall, this study demonstrates that optimized microdermabrasion in combination with a mask can be used to selectively remove stratum corneum with three-dimensional control, which is important to translating this technique into a novel method of transdermal drug delivery.
Keywords: Microdermabrasion; Skin; Stratum corneum; Transdermal drug delivery;

Pretreatment effects of moxibustion on the skin permeation and skin and muscle concentrations of salicylate in rats by Dianxiu Cao; Yuko Tazawa; Hiroshi Ishii; Hiroaki Todo; Kenji Sugibayashi (105-110).
The effect of moxibustion on the in vitro and in vivo skin permeation of salicylate was evaluated in rats. First, the effect of moxibustion pretreatment on the elimination pharmacokinetics of salicylate after i.v. injection in rats was determined: no clear difference was observed in the plasma profiles of salicylate (SA) with or without moxibustion pretreatment. However, much higher skin and muscle concentrations of salicylate were observed after its i.v. injection. Next, an in vitro skin permeation study of SA was performed after moxibustion pretreatment. Moxibustion pretreatment increased the skin permeation of SA, and the extent of the increase in SA skin permeation was related to the strength of moxibustion ignition. More intense treatments produced higher skin permeation. A similar enhancement effect on the skin permeation of SA was observed in in vivo studies. Interestingly, the skin/plasma and muscle/plasma ratios of SA were markedly increased by moxibustion pretreatment. These results were due to the induction of enhanced skin permeation and lower clearance into the cutaneous vessels by moxibustion ignition. Combination treatment involving moxibustion and the topical application of drugs such as NSAID may be useful for increasing local pharmaceutical effects by enhancing the drug concentration in the skin and muscle underneath the topical application site.
Keywords: Moxibustion; Topical formulation; Skin permeation enhancement; Skin concentration enhancement; Salicylate;

To clarify the contribution of drug–polymer interaction to the physical stability of amorphous solid dispersions, we studied the crystallization rates of nitrendipine (NTR) enantiomers with identical physicochemical properties in the presence of hydroxypropylmethylcellulose (HPMC), hydroxypropylmethylcellulose phthalate (HPMCP) and polyvinylpyrrolidone (PVP). The overall crystallization rate at 60 °C and the nucleation rate at 50–70 °C of (+)-NTR were lower than those of (−)-NTR in the presence of 10–20% HPMC or HPMCP. In contrast, similar crystallization profiles were observed for the NTR enantiomers in solid dispersions containing PVP. The similar glass transition temperatures for solid dispersions of (−)-NTR and (+)-NTR suggested that the molecular mobility of the amorphous matrix did not differ between the enantiomers. These results indicate that the interaction between the NTR enantiomers and HPMC or HPMCP is stereoselective, and that differences in the stereoselective interaction create differences in physical stability between (−)-NTR and (+)-NTR at 50–70 °C. However, no difference in physical stability between the enantiomers was obvious at 40 °C. Loss of the difference in physical stability between the NTR enantiomers suggests that the stereoselective interaction between NTR and the polymers may not contribute significantly to the physical stabilization of amorphous NTR at 40 °C.
Keywords: Nitrendipine; Enantiomer; Chiral polymer; Solid dispersions; Crystallization;

Porphyrin-containing polyaspartamide gadolinium complexes as potential magnetic resonance imaging contrast agents by Guo-Ping Yan; Zhen Li; Wei Xu; Cheng-Kai Zhou; Lian Yang; Qiao Zhang; Liang Li; Fan Liu; Lin Han; Yuan-Xing Ge; Jun-Fang Guo (119-125).
Porphyrin-containing polyaspartamide ligands (APTSPP–PHEA–DTPA) were synthesized by the incorporation of diethylenetriaminepentaacetic acid (DTPA) and 5-(4′-aminophenyl)-10,15,20-tris(4′-sulfonatophenyl) porphyrin, trisodium salt (APTSPP) into poly-α,β-[N-(2-hydroxyethyl)-l-aspartamide] (PHEA). These ligands were further reacted with gadolinium chloride to produce macromolecule-gadolinium complexes (APTSPP–PHEA–DTPA–Gd). Experimental data of 1H NMR, IR, UV and elemental analysis evidenced the formation of the polyaspartamide ligands and gadolinium complexes. In vitro and in vivo property tests indicated that APTSPP–PHEA–DTPA–Gd possessed noticeably higher relaxation effectiveness, less toxicity to HeLa cells, and significantly higher enhanced signal intensities (SI) of the VX2 carcinoma in rabbits with lower injection dose requirement than that of Gd–DTPA. Moreover, APTSPP–PHEA–DTPA–Gd was found to greatly enhance the contrast of MR images of the VX2 carcinoma, providing prolonged intravascular duration, and distinguished the VX2 carcinoma and normal tissues in rabbits according to MR image signal enhancements. These porphyrin-containing polyaspartamide gadolinium complexes can be used as the candidates of contrast agents for targeted MRI to tumors.
Keywords: Magnetic resonance imaging (MRI); Gadolinium complex; Polyaspartamide; Porphyrin; Relaxivity; VX2 carcinoma;

Incidence of low bioavailability of leuprolide acetate after percutaneous administration to rats by dissolving microneedles by Yukako Ito; Hiroshi Murano; Noriyuki Hamasaki; Keizo Fukushima; Kanji Takada (126-131).
Two-layered dissolving microneedles of which acral portion contained leuprolide acetate (LA) as solid dispersion were prepared with sodium chondroitin sulfate as the base and the systemic absorption efficiency of LA was studied in rats after administration to their abdominal skin. A patch contained 100 dissolving microneedles of which length and basement diameter were 469.8 ± 4.7 μm and 284.5 ± 9.8 μm, where LA content was 14.3 ± 1.6 μg. In vitro dissolution experiment showed that LA was released from dissolving microneedle patch within 3 min. LA was stable in the patch, % recoveries for 3 months were 102.2 ± 1.9–95.3 ± 1.9%. One and half-patch of LA dissolving microneedles were administered to the rat skin and plasma LA concentrations were measured by LC–MS/MS. Plasma LA concentrations increased immediately after administration, and reached to the maximum level at 15 min, where C max were 6.0 ± 0.7 and 16.4 ± 0.9 μg/ml, respectively. The AUC were 5.8 ± 0.8 and 14.5 ± 0.4 μg h/ml and BA were 33.8 ± 4.3% and 31.7 ± 0.8%. When LA solution was subcutaneously (s.c.) injected to rats, 50 μg/kg, the BA was 32.0 ± 2.1%. Relative BA of LA from dissolving microneedles against s.c. solution was 105.6 ± 13.5%. The degradation rate of LA in the rat skin tissue homogenate was very fast where the half-life was 16.3 ± 5.7 min. The degradation of LA in the skin tissue was the cause of the low BA of LA after percutaneous administration to rats.
Keywords: Leuprolide acetate; Dissolving microneedles; Low bioavailability; Degradation in the skin tissue; Rat;

Novel chitosan−magnesium aluminum silicate nanocomposite film coatings for modified-release tablets by Wanwisa Khunawattanakul; Satit Puttipipatkhachorn; Thomas Rades; Thaned Pongjanyakul (132-141).
Chitosan (CS), a positively charged polysaccharide, and magnesium aluminum silicate (MAS), a negatively charged clay with silicate layers, can electrostatically interact to form nanocomposite films. In this study, CS–MAS nanocomposite films were evaluated for use in tablet film coating. Effects of CS–MAS ratio and coating level on water uptake and drug release from the coated tablets were investigated. Surface and film matrix morphology of the coated film and the effect of enzymes in the simulated gastro-intestinal fluid on drug release were also examined. The results demonstrated that the CS–MAS coated tablets had a rough surface and a layered matrix film, whereas a smooth surface and dense matrix film on the CS coated tablets was found. However, the CS–MAS coated tablets provided fewer film defects than the CS coated tablets. Nanocomposite formation between CS and MAS could retard swelling and erosion of CS in the composite films in acidic medium. The higher MAS ratio of the CS–MAS coated tablets gave lower water uptake and slower drug release when compared with the CS coated tablets. Moreover, the CS–MAS films on the tablets presented good stability towards enzymatic degradation in simulated intestinal fluid. The release of drug from the CS–MAS coated tablets could be modulated by varying CS–MAS ratios and coating levels. Additionally, drug solubility also influenced drug release characteristics of the CS–MAS coated tablets. These findings suggest that the CS–MAS nanocomposites displays a strong potential for use in tablet film coating intended for modifying drug release from tablets.
Keywords: Chitosan; Magnesium aluminum silicate; Nanocomposite film; Film coating; Drug release;

Design of a composite drug delivery system to prolong functionality of cell-based scaffolds by Ainhoa Murua; Enara Herran; Gorka Orive; Manoli Igartua; Francisco Javier Blanco; José Luis Pedraz; Rosa Mª Hernández (142-150).
Co-implantation of DXM-loaded PLGA microspheres resulted in improved functionality of the cell-based grafts during 45 days in allogeneic mice.Cell encapsulation technology raises hopes in medicine and biotechnology. However, despite important advances in the field in the past three decades, several challenges associated with the biocompatibility are still remaining. In the present study, the effect of a temporary release of an anti-inflammatory agent on co-administered encapsulated allogeneic cells was investigated. The aim was to determine the biocompatibility and efficacy of the approach to prevent the inflammatory response. A composite delivery system comprised of alginate-poly-l-lysine-alginate (APA)-microencapsulated Epo-secreting myoblasts and dexamethasone (DXM)-releasing poly(lactic-co-glycolic acid) (PLGA) microspheres was implanted in the subcutaneous space of Balb/c mice for 45 days.The use of independently co-implanted DXM-loaded PLGA microspheres resulted in an improved functionality of the cell-based graft, evidenced by significantly higher hematocrit levels found in the cell-implanted groups by day 45, which was found to be more pronounced when higher cell-doses (100 μL) were employed. Moreover, no major host reaction was observed upon implantation of the systems, showing good biocompatibility and capability to partially avoid the inflammatory response, probably due to the immunosuppressive effects related to DXM. The findings of this study imply that DXM-loaded PLGA microspheres show promise as release systems to enhance biocompatibility and offer advantage in the development of long-lasting and effective implantable microencapsulated cells by generating a potential immunopriviledged local environment and an effective method to limit the structural ensheathing layer caused by inflammation.
Keywords: Alginate; PLGA; Biocompatibility; Microencapsulation; Dexamethasone; Erythropoietin;

Decoration of fibrinogen γ-chain peptide on adenosine diphosphate-encapsulated liposomes enhances binding of the liposomes to activated platelets by Koji Tokutomi; Toshiaki Tagawa; Maki Korenaga; Masatoshi Chiba; Tomohiro Asai; Naohide Watanabe; Shinji Takeoka; Makoto Handa; Yasuo Ikeda; Naoto Oku (151-157).
For the purpose of efficient hemostasis, we previously developed ADP-encapsulated liposomes modified with a dodecapeptide (HHLGGAKQAGDV, H12), H12-(ADP)Lipo. This liposome actually enhanced platelet aggregation in vitro, and showed significant hemostatic effect in vivo. Since fibrinogen (Fbg) is abundant in the bloodstream, it is unclear why this liposome binds platelets so efficiently, overcoming the competition with Fbg. Therefore, we investigated the relationship between H12 density on the liposome and the binding ability to platelets, and evaluated the inhibitory effect of Fbg on the binding of H12-(ADP)Lipo to platelets. As a result, the binding ability to platelets steeply increased depending on H12 density until it reached about 3 × 1015 H12 molecules/m2. The 50% inhibition concentration of Fbg on the binding of H12-(ADP)Lipo to platelets was about 25-fold over the concentration of H12 molecules on the liposome. Moreover, almost no inhibition by Fbg was observed at the physiological concentration of it. This result suggests that the ability of H12 to bind to GPIIb/IIIa increased overwhelmingly by the anchoring to the liposome that enabled the cooperative binding of H12 peptides to the platelets.
Keywords: Hemostasis; Liposome; Fibrinogen; Platelet aggregation; ADP;

An oil-free microemulsion for intravenous delivery of diallyl trisulfide: Formulation and evaluation by Xinru Li; Yuanting Yue; Yanxia Zhou; Yating Fan; Chao Fan; Yanqing Huang; Fei Wu; Yan Liu (158-166).
The aim of the present study was to develop an oil-free o/w microemulsion, Cremophor EL:ethanol–propylene glycol:saline, for diallyl trisulfide (DATS) for intravenous (i.v.) administration to modify the safety and pharmacokinetics of DATS. The ternary diagram was constructed to identify the regions of dilutable microemulsions, and the optimal composition of microemulsion was determined by evaluation of injection safety such as hemolysis, intravenous stimulation and injection anaphylaxis compared to commercial formulation Chentian®. Promising microemulsion with modified injection safety was developed that could incorporate 100 mg/g of DATS. The droplet size of the microemulsion was about 26 nm in diameter with narrow distribution (polydispersity index: 0.14). Acute toxicity test showed that median lethal dose (LD50) of DATS microemulsion was 1.69-fold higher than that of Chentian®. Pharmacokinetics was assessed by comparing with the commercial injection after intravenous administration to rats at a dose of 30 mg/kg. The developed microemulsion showed significant higher area under the drug concentration–time curve and lower clearance and distribution volume than those of Chentian® (p  < 0.05). This helped DATS to reach higher level in vessel, and circulate in the blood stream for a longer time resulting in better therapeutic effect. In conclusion, microemulsion would be a promising intravenous delivery system for DATS.
Keywords: Diallyl trisulfide; Microemulsion; Safety; Pharmacokinetics;

Keywords: Silicone based drug delivery systems; PH dependent drug delivery; Hybrid macromolecules; Nanocarriers; Dendritic micells;

Self-assembly of cyclodextrin complexes: Aggregation of hydrocortisone/cyclodextrin complexes by Martin Messner; Sergey V. Kurkov; Marcus E. Brewster; Phatsawee Jansook; Thorsteinn Loftsson (174-183).
A permeability profile of hydrocortisone from HPβCD solution saturated with the drug through semi permeable cellophane membrane with MWCO 15 kDa. Negative deviation (J exp) from the theoretical flux (J theo) is due to hydrocortisone/HPβCD complex aggregation. J D is the flux of the free drug.Cyclodextrins (CDs) are well known functional excipients for solubilization and stabilization of drugs in aqueous formulations as well as enabling adjuncts for increasing the oral bioavailability of solid dosage forms. More recently a number of the valuable properties of these CDs have been ascribed to nanoparticulate aggregation in addition to its ability to form molecular inclusion complexes. The purpose of this study is to identify and characterize the aggregation of CD inclusion complexes with a model drug, hydrocortisone, in saturated solutions which are more relevant to drug formulation than highly dilute systems. Penetration studies of complexes through membranes and phase solubility relationships were assessed for saturated hydrocortisone solutions with the parent CDs, namely αCD, βCD, γCD or with various water-soluble derivatives, i.e., 2-hydroxypropyl-βCD (HPβCD), 2-hydroxypropyl-γCD (HPγCD) or sulfobutyl ether-β-CD (SBEβCD). The data indicate that βCD and γCD form micro-aggregates with hydrocortisone resulting in non-linear phase-solubility relationships. By contract, the other studies of CDs or CD derivatives were found to form nanoaggregates with hydrocortisone resulting in linear solubilization relationships. Permeability profiles were evaluated for the systems formed and are described in three sections specifically a section (section I) where flux is linear (Fickian) as a function of CD concentration, a section (section II) where flux deviates in a negative fashion from linearity but still increases as the CD concentration increases and a section (section III) where flux is independent of the cyclodextrin concentration. Diminished values of flux can be interpreted based on the formation of nanoaggregates of hydrocortisone/CD complexes. Extrapolation of section I data made it possible to obtain theoretical flux values which could be used to estimate the fraction of complexes and drug which participate in aggregation. The CDs which appeared to demonstrate the lowest tendency to form complex aggregates were αCD and SBEβCD, due to their low complexation efficacy and repulsive forces, respectively. Complex aggregates with these CDs are also smaller with maximum size between 50 and 100 kDa. HPβCD and HPγCD complex aggregates manifested a maximum size above 100 kDa and the fraction of drug which participates in complex aggregation with these species is higher than for the other materials assessed. In the case of 90 mM HPγCD solution, data suggest that 87% of all hydrocortisone is tied up in the form of aggregates. These high concentrations were confirmed by TEM which found most particles in the 3–5 nm range but rarely particles as large as 10 and 20 nm. Speculation on the mechanism of the aggregation processes and equilibrium constants are provided but these tend to punctuate our limited understanding of these potentially important processes.
Keywords: Cyclodextrins; Permeation; Complex aggregates; Nanoparticles; Hydrocortisone;

Preparation of Tacrolimus loaded micelles based on poly(ɛ-caprolactone)–poly(ethylene glycol)–poly(ɛ-caprolactone) by YingJing Wang; Cheng Wang; ShaoZhi Fu; Qing Liu; DanYang Dou; Hang Lv; Min Fan; Gang Guo; Feng Luo; ZhiYong Qian (184-189).
Self-assembled polymeric micelles are widely applied in drug delivery system. In this study, Tacrolimus (FK506) loaded micelles were prepared based on biodegradable poly(ɛ-caprolactone)–poly(ethylene glycol)–poly(ɛ-caprolactone) (PCEC) copolymers. Micelles were prepared by self-assembly of triblock copolymer PCEC in distilled water triggered by its amphiphilic characteristics. Drug loading and encapsulation efficiency were determined by adjusting the weight ratio of FK506 and PCEC. The particle size distribution and variation of obtained micelles were determined using Malvern laser particle size analyzer, while the spherical geometry was observed on transmission electron microscope (TEM), and the crystallographic assays were fulfilled by X-ray diffractometer (XRD). Besides, in vitro release profile demonstrated a significant difference between rapid release of free Tacrolimus and much slower and sustained release of FK506 loaded micelles. These results suggested that we have successfully prepared Tacrolimus loaded micelles in an improved method which is safer and more efficient. The prepared micelles might be potential carriers for Tacrolimus delivery in immunosuppressive therapy.
Keywords: Tacrolimus; PCL-PEG-PCL; Polymeric micelles; Drug slow-release;

The aim of this study was to formulate nanoparticles from three different hyperbranched polymers, namely an unmodified dendritic polyester (Boltorn H40™), a lipophilic, fatty acid modified dendritic polymer (Boltorn U3000™) and an amphiphilic dendritic polymer (Boltorn W3000™) for drug delivery of paclitaxel and to investigate their properties. A solvent displacement method allowed preparation of nanoparticles from all three hyperbranched polymers. Nanoparticle sizes ranged from 70 to 170 nm. The lipophilic Boltorn U3000™ formed the biggest nanoparticles and the amphiphilic Boltorn W3000™ formed the smallest ones. Nanoparticles of amphiphilic Boltorn W3000™ displayed only a slightly negative zeta potential, while more negative zeta potentials were measured for nanoparticles based on the other two polymers. Degradation profiles were investigated by short time pH-stat titration. Boltorn H40™ showed a faster degradation rate then the two other fatty acid containing polymers. For Boltorn H40™, degradation rate was also investigated in longer term mass loss studies resulting in 30% degradation during 3 weeks. Cytotoxicity of the nanoparticles was studied by MTT assay displaying low cytotoxicity for all three polymers. All three types of nanoparticles were loaded with paclitaxel and their release profiles were studied. Sizes and zeta potentials remained stable after loading and did not change significantly. These three types of hyperbranched polymers show potential as nanoparticulate delivery systems and should be further studied. Due to their high loading efficiency, Boltorn U3000 and W3000 represent the most interesting candidates.
Keywords: Hyperbranched polyesters; Nanoparticles; Degradation; Toxicity; Paclitaxel;

Water state effect on drug release from an antibiotic loaded polyurethane matrix containing albumin nanoparticles by Andrea Martinelli; Lucio D’Ilario; Iolanda Francolini; Antonella Piozzi (197-206).
The water state in an antibiotic-loaded composite drug delivery system consisting of albumin nanoparticles dispersed into a carboxylated polyurethane has been investigated. The non freezing water fraction was accountable for a slower drug release along with the nanoparticle/polyurethane interactions. This latter also improved the system mechanical properties.Water mobility plays a crucial role in determining transport properties of small molecules in polymer matrices. In particular, in drug delivery systems, water state affects the pharmacokinetics, especially drug absorption, diffusion and release. In the present study, the state of water in an antibiotic-loaded composite consisting of albumin nanoparticles (BSAnp) dispersed into a carboxylated polyurethane (PEUA) has been investigated and compared with that of the single drug-loaded components. The antibiotic cefamandole nafate was used as a model drug. DSC analysis, used to evaluate the freezing and non-freezing water fractions in the hydrated samples, showed that in BSAnp water can adsorb both in the inter-particles regions and inside the particles. With increasing of total adsorbed water amount, the contribution of the freezing water fraction was higher than the non-freezing one. As for PEUA, the majority of water molecules absorbed is in a mobile freezing state (about 60% of the W tot).As for the PEUA/BSAnp composite, the higher polyurethane phase segregation induced by the nanoparticles as well as the higher non-freezing water fraction significantly enhanced drug uptake with respect to PEUA. Moreover, the greater non-freezing water fraction allowed the drug to penetrate within BSA nanoparticles and to give rise then to a controlled drug release. Indeed, the diffusion barrier exerted by nanoparticles and the matrix prolonged the antimicrobial activity from 4 to 9 days.Finally, the higher polyurethane phase segregation also improved composite mechanical properties, as evidenced in stress–strain experiments and dynamic mechanical analysis.
Keywords: Freezing and non freezing water; Nano-composites; Nanoparticles; Drug delivery systems; Albumin; Polyurethanes;

Three novel amino acid based anionic amphiphilic copolymers poly(sodium N-acryloyl-l-valinate-co-alkylacrylamide) (where, alkyl = octyl and dodecyl) with either 9 or 16 mol% hydrophobic substitution were synthesized. These hydrophobically modified polyelectrolytes (HMPs), above a critical concentration, self-assemble in aqueous solution through inter-chain hydrophobic aggregation, forming micelle-like aggregates having hydrodynamic diameter in the range of 50–200 nm. The HMPs were found to undergo conformational changes with the change in solution pH, electrolyte and additive concentration, and temperature. The polymeric micelles were observed to be stable under biological conditions (pH 7.4, [NaCl] = 150 mM and temperature (37 °C)). The solubilization capacity of the polymeric micelles for six important non-steroidal anti-inflammatory drugs of different hydrophobicity was evaluated. Depending upon the hydrophobicity the solubilities of the drugs were observed to increase ca. 2–10 times in the presence of 1.0 g/L copolymers. The in vitro release kinetics of the loaded drug was studied under physiological pH. To explore their potential application in pharmaceutical industries hemocompatibility and cytotoxicity studies were carried out using hemolytic and MTT assay, respectively. The anionic HMPs were found to be not directly toxic to mammalian cells.
Keywords: Polymeric micelles; Solubilization; In vitro release; Hemocompatibility; Cytotoxicity;