International Journal of Pharmaceutics (v.421, #2)

Automatic encapsulation machines have two dosing principles: dosing disc and dosator. Dosator-based machines compress the powder to plugs that are transferred into capsules. The encapsulation process in dosator-based capsule machines was modeled in this work. A model was proposed to predict the weight and length of produced plugs. According to the model, the plug weight is a function of piston dimensions, powder-bed height, bulk powder density and precompression densification inside dosator while plug length is a function of piston height, set piston displacement, spring stiffness and powder compressibility. Powder densification within the dosator can be achieved by precompression, compression or both. Precompression densification depends on the powder to piston height ratio while compression densification depends on piston displacement against powder. This article provides the theoretical basis of the encapsulation model, including applications and limitations. The model will be applied to experimental data separately.
Keywords: Encapsulation; Filling; Powder technology; In silico modeling; Compression; Excipients; Formulation; Mathematical model; Solid dosage form; Capsules; Quality by design (QbD);

A theoretical model was previously derived to predict powder encapsulation in dosator-based machines. The theoretical basis of the model was discussed earlier. In this part; the model was evaluated experimentally using two powder formulations with substantially different flow behavior. Encapsulation experiments were performed using a Zanasi encapsulation machine under two sets of experimental conditions. Model predicted outcomes such as encapsulation fill weight and plug height were compared to those experimentally obtained. Results showed a high correlation between predicted and actual outcomes demonstrating the model's success in predicting the encapsulation of both formulations. The model is a potentially useful in silico analysis tool that can be used for capsule dosage form development in accordance to quality by design (QbD) principles.
Keywords: Encapsulation; Filling; Powder technology; In silico modeling; Compression; Excipients; Formulation; Mathematical model; Solid dosage form; Capsules;

Combining an amino acid and a sugar is a known strategy in the formulation of spray or freeze dried biomolecule powder formulations. The effect of the amino acid leucine in enhancing performance of spray-dried powders has been previously demonstrated, but interaction effects of several constituents which may provide multiple benefits, are less well-understood. A 3 factor 2 level (23) factorial design was used to study the effects of leucine, glycine and alanine in a mannitol-based dry powder formulation on particle size, aerosolisation, emitted dose and cohesion. Other qualitative tests including scanning electronic microscopy and X-ray powder diffraction were also conducted on the design of experiment (DoE) trials. The results show that the use of glycine and/or alanine, though structurally related to leucine, did not achieve similar aerosol performance enhancing effects, rather the particle formation was hindered. However, when used in appropriate concentrations with leucine, the combination of amino acids produced an enhanced performance regardless of the presence of glycine and/or alanine, yielding significantly modified particle properties. The results from the DoE analyses also revealed the lack of linearity of effects for certain responses with a significant curvature in the model which would otherwise not be discovered using a trial-and-error approach.
Keywords: Dry powder inhalers; Aerosolisation; Pulmonary delivery; Design of experiment; Factorial design; Spray-drying;

Characterization and evaluation of miconazole salts and cocrystals for improved physicochemical properties by Shunichirou Tsutsumi; Motoo Iida; Norio Tada; Takashi Kojima; Yukihiro Ikeda; Toshiya Moriwaki; Kenjirou Higashi; Kunikazu Moribe; Keiji Yamamoto (230-236).
Miconazole salts and cocrystals were studied to improve the physicochemical properties of miconazole. Maleate, hemifumarate, and hemisuccinate were prepared and characterized by powder X-ray diffractometry, differential scanning calorimetry, and single crystal X-ray diffractometry. The intrinsic dissolution rate and stability of each miconazole crystal form were compared to those of freebase and nitrate to evaluate the optimal crystal form. Crystal structure analysis indicated that maleate was a salt formed by proton transfer from the acid to the imidazole group of miconazole. Hemifumarate and hemisuccinate were determined to be cocrystals formed by hydrogen bonding between the acids and the base in their crystal lattices. Intrinsic dissolution tests showed that the formation of salts and cocrystals improved the dissolution rate of miconazole. Stability tests of preliminary formulations prepared with each crystal form indicated that maleate and hemifumarate were unstable at 80 °C and generated a specific degraded product, i.e., a Michael adduct, between miconazole and the acids. Hemisuccinate had a superior intrinsic dissolution rate and stability, and is thus considered a promising crystal form of miconazole
Keywords: Miconazole; Salt; Cocrystal; Crystal structure; Intrinsic dissolution rate; Stability;

Real-time predictions of drug release and end point detection of a coating operation by in-line near infrared measurements by Claire Gendre; Mathieu Boiret; Muriel Genty; Pierre Chaminade; Jean Manuel Pean (237-243).
The aim of this work was to carry out real-time near infrared (NIR) predictions of drug release from sustained release coated tablets and to determine end point of coating operation.In-line measurements were ensured by implementation of a NIR probe inside a pan coater. Tablets were coated using a functional aqueous dispersion of ethylcellulose blended with PVA–PEG graft copolymer to obtain a controlled drug release dosage form over 16 h. Samples were collected at regular intervals and subjected to a standardized curing step. Percentages of released drug at 4 h, 8 h and 12 h were selected to describe the controlled drug release of cured tablets. These dissolution criteria were used as reference values to calibrate NIR spectral information and to develop three partial least squares regressions.Low predictive errors of 1.7%, 1.9% and 1.5%, respectively, were obtained. The coating operation was stopped while desired dissolution criteria were achieved, corresponding to a coating level around 10%.The present study demonstrated that real-time NIR measurements could be performed on non-finished drug products to predict dissolution properties of cured coated tablets. This novel and innovative approach fulfils the expectations of ICH Q8 guideline on pharmaceutical development, in terms of process understanding and process analytical technology (PAT) control strategy. This approach should be however adapted to curing operation to allow a real-time release testing.
Keywords: Coating operation; Dissolution testing; Sustained release tablets; Process analytical technology (PAT); End point determination; Near infrared spectroscopy (NIRS);

Improvement of low bioavailability of a novel factor Xa inhibitor through formulation of cationic additives in its oral dosage form by Yoshimine Fujii; Taro Kanamaru; Hiroshi Kikuchi; Hiroaki Nakagami; Shinji Yamashita; Mitsuru Akashi; Shinji Sakuma (244-251).
A clinical trial of (2S)-2-[4-[[(3S)-1-acetimidoyl-3-pyrrolidinyl]oxy]phenyl]-3-(7-amidino-2-naphtyl) propanoic acid (DX-9065) revealed that its oral bioavailability was only 3% when it was administered as a conventional capsule formulation. The low bioavailability of DX-9065 was likely caused by both its poor membrane permeability and its electrostatic interaction with anionic bile acids. We hypothesized that DX-9065 absorption would be enhanced when the cationic drug was free from the complex through its replacement with other cationic substances. Polystyrene nanospheres coated with cationic poly(vinylamine) and cholestyramine, which is clinically used as a cholesterol-lowering agent, dramatically prevented DX-9065 from interacting with chenodeoxycholic acid in vitro. Successive animal experiments showed that bioavailability of DX-9065 administered with these cationic substances was 2–3 times that of DX-9065 administered solely. A dry syrup formulation with one-half of a minimal cholesterol-lowering equivalent dose of cholestyramine was designed, and the clinical trial was resumed. A 1.3-fold increase in bioavailability of DX-9065 was observed when the dry syrup was administered. We successfully demonstrated that DX-9065 absorption was enhanced when the drug was administered with cationic additives; however, it appeared that the absorption-enhancing function of cholestyramine largely depended on its dose. The dose escalation is probably prerequisite for the significant improvement of DX-9065 absorption in humans.
Keywords: Factor Xa inhibitor; Bioavailability; Bile acid; Nanosphere; Cholestyramine; Electrostatic interaction;

The effect of water-soluble polymers, PEG and PVP, on the solubilisation of griseofulvin in aqueous micellar solutions of Pluronic F127 by Cristiane P. Oliveira; Maria Elenir N.P. Ribeiro; Nágila M.P.S. Ricardo; Ticiane V. de P. Souza; Carolina Lima Moura; Chiraphon Chaibundit; Stephen G. Yeates; Keith Nixon; David Attwood (252-257).
The purpose of this study was to investigate the possibility of enhancing the solubilisation capacity of micellar solutions of Pluronic F127 for the poorly water-soluble drug griseofulvin by co-formulating with a water-soluble polymer. The effect of the addition of the polyethylene glycols PEG6000 and 35000, and the poly(vinylpyrrolidone)s PVP K30 and K90, on the solubilisation capacity of 1 wt% solutions of Pluronic F127 was related to the effect of these additives on particle size as determined by dynamic light scattering measurements. The addition of PEG35000 to 1 wt% F127 solutions significantly increased the solubility capacity expressed in terms of unit weight of F127; PVP K90 had a smaller effect but no enhancement was noted following the addition of PEG6000 or PVP K30. Solubilisation enhancement was thought to be a consequence of the association of the polymers with the E-blocks of the micelle corona so providing an expanded region of reduced polarity for drug solubilisation.
Keywords: Solubilisation; Micelles; Pluronic F127; PEG; PVP;

Supercritical fluid assisted atomization introduced by hydrodynamic cavitation mixer (SAA-HCM) was used to produce lysozyme microparticles with controlled particle size distribution in the range for aerosol drug delivery. The process is based on the atomization effect of carbon dioxide. The solubilization of certain amount of carbon dioxide in the solution plays the key role and the HCM can intensify mass transfer between carbon dioxide and liquid feedstock greatly. Water was used as the solvent to solubilize lysozyme and thus no organic residual was detected. The influences of process parameters on particle formation were investigated including temperature in the precipitator, pressure and temperature in the mixer, concentration of the solution and feed ratio CO2/solution. The particles were characterized with respect to their morphologies and particle size: well defined, spherical and separated particles with diameters ranging between 0.2 and 5 μm could be always produced at optimum operating conditions. Bio-activity assay showed that good activity maintenance of higher than 85% for lysozyme was usually achieved. Solid state characterizations were further performed to investigate the changes of lysozyme in the process. Fourier transform infrared spectroscopy indicated that no change in secondary structure had occurred for processed lysozyme. X-ray diffraction analysis showed that the lysozyme particles produced remained similarly amorphous as the raw material. Differential scanning calorimetry and thermogravimetry analysis revealed that there was no significant difference in water association but with the increase of water content after processing.
Keywords: Supercritical fluid assisted atomization; Hydrodynamic cavitation mixer; Lysozyme; Micronization; Bio-activity;

Display OmittedDevelopment of quality estimation models using near infrared spectroscopy (NIRS) and multivariate analysis has been accelerated as a process analytical technology (PAT) tool in the pharmaceutical industry. Although linear regression methods such as partial least squares (PLS) are widely used, they cannot always achieve high estimation accuracy because physical and chemical properties of a measuring object have a complex effect on NIR spectra. In this research, locally weighted PLS (LW-PLS) which utilizes a newly defined similarity between samples is proposed to estimate active pharmaceutical ingredient (API) content in granules for tableting. In addition, a statistical wavelength selection method which quantifies the effect of API content and other factors on NIR spectra is proposed. LW-PLS and the proposed wavelength selection method were applied to real process data provided by Daiichi Sankyo Co., Ltd., and the estimation accuracy was improved by 38.6% in root mean square error of prediction (RMSEP) compared to the conventional PLS using wavelengths selected on the basis of variable importance on the projection (VIP). The results clearly show that the proposed calibration modeling technique is useful for API content estimation and is superior to the conventional one.
Keywords: PAT; NIR; API content estimation; Locally weighted partial least squares; Wavelength selection;

Series of monooleate-modified PEG with active carboxylic terminus on the other end (MO–PEG–COOH) were used to modify the lipid emulsions surface to prepare a sterically stabilized lipid emulsions for carrying Traditional Chinese Medicine – breviscapine. Based on the research of relationship between polymer structure and prolonged circulation activity, we developed an optimized formulation and a technological method to prepare the sterile and stable MO–PEG10,000–COOH (Bre–LE–PEG10,000) coated breviscapine lipid emulsions (Bre–LE) for intravenous administration. Follow the optimum preparation, the average particle size, polydispersity index, zeta potential, Ke value and content of final product were determined to be (207.1 ± 8.5) nm, 0.197 ± 0.005, (−33.6 ± 2.0) mV, (21.1 ± 2.3)% and (95.0 ± 1.8)% respectively (n  = 3). The characteristics, stability and safety of Bre–LE–PEG10,000 were also studied with Bre–LE as a control. Increased plasma concentration by surface modification of the lipid emulsions may enhance the pharmacological activity of breviscapine to promote blood circulation.
Keywords: Breviscapine; Lipid emulsions; PEG; Prolonged circulation; Stability; Toxicity;

Ionically cross-linked chitosan microspheres for controlled release of bioactive nerve growth factor by Wen Zeng; Jinghui Huang; Xueyu Hu; Wei Xiao; Mengyao Rong; Zhi Yuan; Zhuojing Luo (283-290).
Controlled release of neurotrophic factors to target tissue via microsphere-based delivery systems is critical for the treatment strategies of diverse neurodegenerative disorders. The present study aims to investigate the feasibility of the controlled release of bioactive nerve growth factor (NGF) with ionically cross-linked chitosan microspheres (NGF–CMSs). The microspheres were prepared by the emulsion-ionic cross-linking method with sodium tripolyphosphate (STPP) as an ionic cross-linking agent. The size and distribution of the microspheres, SEM images, Fourier transform infra red spectroscopy (FT-IR), encapsulation efficiency, in vitro release tests and bioactivity assay were subsequently evaluated. We found that the microspheres had relatively rough surfaces with mean sizes between 20 and 31 μm. FT-IR results provided evidence of ionic interaction between amino groups and phosphoric groups of chitosan and STPP. The NGF encapsulation efficiency ranged from 63% to 88% depending on the concentration of STPP. The in vitro release profiles of NGF from NGF–CMSs were influenced by the concentration of STPP. NGF–CMSs which were cross-linked with higher concentration of STPP showed slower but sustained release of NGF. In addition, the released NGF from NGF–CMSs was capable of maintaining the viability of PC12 cells, as well as promoting their differentiation. Taken together, our findings suggest that NGF–CMSs are capable of releasing bioactive NGF over 7 days, thus having potential application in nerve injury repair.
Keywords: Chitosan microspheres; Nerve growth factor; Controlled release; Peripheral nerve regeneration; STPP;

Development of lysolipid-based thermosensitive liposomes for delivery of high molecular weight proteins by Xin Zhang; Paul F. Luckham; Alun D. Hughes; Simon Thom; Xiao Yun Xu (291-292).
The purpose of the present investigation was to determine the feasibility of using lysolipid-based thermal sensitive liposomes (145 nm) for high molecular weight molecule delivery. Fluorescein isothiocyanate conjugate-albumin was used as a model drug (MW 66 kDa). Thermal sensitive liposomes, which encapsulated the protein were prepared using a passive encapsulation methodology involving freeze-thawing cycles followed by extrusion. In vitro release studies at various temperatures indicated rapid release behaviour of the encapsulated protein at 42 and 44.5 °C but a good stability at 37.5 °C. The current findings suggest that lysolipid-based thermal sensitive liposomes can be used to deliver high molecular weight molecules.
Keywords: Lysolipid incorporated thermal sensitive liposome; Macromolecular therapeutics; Mild-hyperthermia;

Layered lipid microcapsules for mesalazine delayed-release in children by Anna Giulia Balducci; Gaia Colombo; Giuseppe Corace; Cristina Cavallari; Lorenzo Rodriguez; Francesca Buttini; Paolo Colombo; Alessandra Rossi (293-300).
The goal was to make available a delayed-release dosage form of mesalazine to be dispersed in water to facilitate swallowing in adults and children. Mesalazine microparticles containing carnauba wax were prepared by spray-congealing technique. A second step of spray-congealing of carnauba microparticles dispersed in liquefied stearic acid gave rise to mesalazine lipid microcapsules in which several carnauba microparticles remained embedded as cores in a reservoir structure. In order to favor their water dispersion, the lipid microcapsules were dry coated by tumbling them with different ratios of mannitol/lecithin microparticles prepared by spray-drying.Release rate measurements showed a delayed-release behavior, in particular a pH-dependence with less than 10% of drug released in acidic medium and complete release in phosphate buffer pH 7.4 in 4–5 h. The layering with hydrophilic excipient microparticles allowed manufacturing of a pH-dependent dosage form suitable for extemporaneous oral use in adults and children.
Keywords: Mesalazine; Lipid; Microcapsules; Spray-congealing; Excipient microparticles; Delayed-release;

Inclusion complexes of isoflavones with two commercially available dendrimers: Solubility, stability, structures, release behaviors, cytotoxicity, and anti-oxidant activities by Chen Zhao; Yitong Wang; Yunzhang Su; Hongfeng Zhang; Lingxiao Ding; Xiaofeng Yan; Di Zhao; Naimin Shao; Xiyun Ye; Yiyun Cheng (301-309).
We prepared and characterized the inclusion complexes of daidzein with poly(amidoamine) (PAMAM) and poly(propylene imine) (PPI) dendrimers. Aqueous solubility of daidzein was significantly enhanced by both PAMAM and PPI (186- and 650-fold at 0.36 mM, respectively). Daidzein in G3 PAMAM solution is more stable than that in G4 PPI. NMR studies reveal the encapsulation of daidzein within the interior cavities of PPI through hydrophobic interactions. Daidzein exhibits a slower release behavior from PPI than that from PAMAM. PPI/daidzein complex is much more toxic than PAMAM/daidzein complex on several cell lines. PAMAM/daidzein complexes showed similar protective effect on oxidative stress-induced cytotoxicity as compared to free daidzein. These results suggest that the inclusion of daidzein with dendrimer can effectively improve the solubility, prolong the delivery, and maintain the anti-oxidant activity of daidzein. This research provides new insights into dendrimer-based drug delivery systems and will be helpful for the design of novel dendrimer/drug formulations.
Keywords: Isoflavones; Daidzein; Dendrimer; PAMAM; PPI;

Controlled green tea polyphenols release from electrospun PCL/MWCNTs composite nanofibers by Shijun Shao; Long Li; Guang Yang; Jingrong Li; Chao Luo; Tao Gong; Shaobing Zhou (310-320).
Poly(ɛ-caprolactone)/multi-walled carbon nanotubes (PCL/MWCNTs) composite nanofibers with various content of green tea polyphenols (GTP) were successfully fabricated via an electrospinning technology to maintain the chemical structural stability of GTP. The non-covalent interaction between MWCNTs and GTP was measured by UV–vis spectrophotometer and FT-IR. The topographical features of the nanofibers were characterized by scanning electron microscopy (SEM). The dispersibility of MWCNTs and the distribution of GTP in nanofibers were observed by transmission electron microscopy (TEM) and laser scanning confocal microscope (LSCM), respectively. In vitro degradation was also characterized in terms of the morphological change and the mass loss of the nanofiber meshes. In vitro GTP release behavior was investigated in phosphate-buffered solution (PBS) at 37 °C. Alamar blue assays were performed to estimate the cytotoxicity of the nanofibers with normal osteoblast cells and the antiproliferative effects to A549 and Hep G2 tumor cells. The results exhibited that the GTP-loaded composite nanofibers possessed a significant inhibition effect to tumor cells. Therefore, GTP, as a multifunctional drug, encapsulated into polymer composite nanofibers, must have broad application prospects in cancer therapy.
Keywords: Electrospinning; Biodegradable; Nanofibers; Controlled release;

Delivery systems for natural antioxidant compounds: Archaeosomes and archaeosomal hydrogels characterization and release study by Ana González-Paredes; Beatriz Clarés-Naveros; Mª Adolfina Ruiz-Martínez; Juan José Durbán-Fornieles; Alberto Ramos-Cormenzana; Mercedes Monteoliva-Sánchez (321-331).
The aim of this study was to use archaeosomes, a novel kind of liposomes made up by archaeal polar lipids, both multilamellars (MLVs) and unilamellars (SUVs), as a topical delivery system for natural antioxidant compounds recovered from olive mill waste. For comparative purpose an analogue formulation of phosphatidylcholine liposomes was prepared. SUVs were smaller than MLVs ones, showing size values smaller than 200 nm, which was maintained during the stability study. Transmission electron microscopy showed spherical morphology for conventional liposomes while archaeosomes had more irregular membranes. Vesicle encapsulation efficiency was quite similar in both formulations and was enough to ensure a good antioxidant activity. Stability studies were performed one month after the preparation of formulations, which showed a high stability with no change in the initial characteristics of the suspensions. Furthermore, the possibility of incorporating the liposomal suspensions in different excipients (Carbopol-940® and Pluronic-127®) for topical administration was studied. In order to evaluate the release behaviour of the different systems prepared, in vitro diffusion studies were carried out using vertical diffusion Franz cells. In both cases the incorporation of the vesicles into the gels lead in a sustained release for 24 h. Archaeosome gels released a similar amount of phenolic compounds regardless the excipient used, while in liposomal gels great release differences were found between carbopol and pluronic gel.
Keywords: Antioxidant phenolic compound; Archaeal polar lipid; Archaeosome; Drug release;

Vitamin E TPGS coated liposomes enhanced cellular uptake and cytotoxicity of docetaxel in brain cancer cells by Madaswamy S. Muthu; Sneha A. Kulkarni; Jiaqing Xiong; Si-Shen Feng (332-340).
The aim of this work was to develop a drug delivery system of liposomes, which are coated with d-alpha-tocopheryl polyethylene glycol 1000 succinate (TPGS), a PEGylated vitamin E, with docetaxel as a model drug for enhanced treatment of brain tumour in comparison with the nude liposomes as well as with the so-called stealth liposomes, i.e. those coated with polyethylene glycol (PEG), which have been intensive investigated in the literature. Docetaxel or coumarin-6 loaded liposomes were prepared by the solvent injection method and characterized for their particle size, polydispersity, zeta potential and drug encapsulation efficiency. C6 glioma cells were employed as an in vitro model to access cellular uptake and cytotoxicity of the drug or coumarin-6 loaded liposomes. The particle size of the PEG or TPGS coated liposomes was ranged between 126 and 191 nm. High-resolution field-emission transmission electron microscopy (FETEM) confirmed the coating of TPGS on the liposomes. The IC50 value, which is the drug concentration needed to kill 50% cells in a designated time period, was found to be 37.04 ± 1.05, 31.04 ± 0.75, 7.70 ± 0.22, and 5.93 ± 0.57 μg/ml for the commercial Taxotere®, the nude, PEG coated and TPGS coated liposomes, respectively after 24 h culture with C6 glioma cells. The TPGS coated liposomes showed great advantages in vitro than the PEG coated liposomes.
Keywords: Cancer nanotechnology; PEGylation; Molecular biomaterials; Nanomedicine; Stealth liposomes; Taxotere®;

Most of the time HIV virus escape immunological burden exerted by antiretroviral drugs and develops resistance against therapy. For complete eradication of virus from body one has to use long term chemotherapies, which results in drug toxicity, drug resistance and eventually poor patient compliance. Nevirapine (NNRTI, non nucleoside reverse transcriptase inhibitor) nanosuspensions were developed and surface modified with serum albumin, polysaccharide and polyethylene glycol to enhance its targeting potential. The biodistribution studies revealed improved antiretroviral drug accumulation in various organs of rat for nanosuspensions as compared to the plain drug solution when administered intravenously. Nanosuspension after surface modification showed further enhancement in accumulation. Higher MRT values of surface coated nanosuspension in brain, liver and spleen as compared to pure drug solution ensured enhanced bioavailability and prolonged residence of the drug at the target site.
Keywords: Nevirapine; HIV/AIDS; Nanosuspensions; High pressure homogenization; Pharmacokinetics; Targeting;

Three novel amino acid based amphiphilic copolymers, poly(sodium N acryloyl-l-aminoacidate-co-dodecylacrylamide) (where aminoacidate = glycinate, leucinate, and phenylalaninate) were synthesized and characterized. These hydrophobically modified polyelectrolytes (HMPs) formed spheroidal nanoparticular aggregates above a critical aggregation concentration with average diameter 20–200 nm and overall negative charges as indicated by dynamic light scattering (DLS) and zeta-potential (ζ  ≈ −10.2 to −25.2) measurements, respectively. The size and shape of the nanostructures was confirmed by transmission electron microscopic images. The micropolarity and microviscosity of the nanosize aggregates were investigated by fluorescence probe method using extrinsic probes like N-phenyl-1-naphthylamine, pyrene, and 1,6-diphenyl-1,3,5-hexatriene. The stability of the copolymer micelles was investigated as a function of pH and temperature using fluorescence and DLS techniques. Both fluorescence probe and DLS data suggest that the copolymer micelles are highly stable under physiological condition (pH 7.4, 37.4 °C). These HMP micelles were evaluated primarily as a drug delivery system. The ability of the copolymers to encapsulate hydrophobic drug was investigated using a poorly water-soluble antifungal drug, griseofulvin. Biocompability of the HMPs were examined by hemolytic and cytotoxicity assay. All three HMPs were non-hemolytic up to the tested concentration of about 1.0 g/L. In vitro biological assay indicated that these new copolymers were also less toxic against 3T3 mammalian cell line. The studies suggest that these newly conceived amino acid based biocompatible polymeric nanoparticles might have potential application as injectible drug delivery systems which can enhance the therapeutic index of poorly water-soluble clinically challenging drugs.
Keywords: Hydrophobically modified polyelectrolytes; Nanoparticles; Griseofulvin; Solubilization; Fluorescence; Toxicity;

Nanostructured lipid carriers as nitroxide depot system measured by electron paramagnetic resonance spectroscopy by S.F. Haag; M. Chen; D. Peters; C.M. Keck; B. Taskoparan; A. Fahr; C. Teutloff; R. Bittl; J. Lademann; M. Schäfer-Korting; M.C. Meinke (364-369).
Various nanometer scaled transport systems are used in pharmaceutics and cosmetics to increase penetration or storage of actives. Nanostructured lipid carriers (NLCs) are efficient drug delivery systems for dermatological applications. Electron paramagnetic resonance (EPR) spectroscopy was used for the determination of TEMPO (2,2,6,6-tetramethyl-1-piperidinyloxy) distribution within the carrier and to investigate the dynamics of skin penetration. Results of ex vivo penetration of porcine skin and in vivo data – forearm of human volunteers – are compared and discussed to previously obtained results with invasomes under comparable conditions. W-band measurements show 35% of TEMPO associated with the lipid compartments of the NLC. Application of TEMPO loaded NLC to skin ex vivo increases the observation time by 12 min showing a stabilisation of the nitroxide radical. Moreover, stabilisation is also seen with data generated in vivo. Thus, same as invasomes NLCs are a suitable slow release depot system.
Keywords: Drug partitioning; Spin probe; Microenvironment; Drug stabilisation; TEMPO; EPR; ESR; Invasomes; NLC; Aminoxyl radical;

This article reports the development of a multifunctional silica nanoparticle system for targeted delivery of hydrophobic imaging and therapeutic agents. Normally, silica nanoparticles have been widely used to deliver hydrophilic drugs such as doxorubicin while difficult to carry hydrophobic drugs. A strategy for loading hydrophobic drugs onto silica nanoparticles via covalent attachment was developed in this study as a universal strategy to solve this problem. Docetaxel, one of the most potent therapeutics for cancer treatment is selected as a model hydrophobic drug and quantum dots (QDs) are used as a model imaging agent. Such a multifunctional delivery system possesses high drug loading capacity, controlled drug release behavior and stable drug reservation. A mixed layer of polyethylene glycol conjugated phospholipids is formed on the nanoparticle surface to further enhance the biocompatibility and cell fusion capability of the delivery system. Folic acid as ligand is then conjugated onto the surface layer for targeting. Such a multifunctional system for targeting, imaging and therapy is characterized and evaluated in vitro. Fluorescent confocal microscopy is used to monitor the cellular uptake by specific cancer cells. Cytotoxicity studies are conducted by using MTT assay.
Keywords: Nanomedicine; Cancer nanotechnology; Docetaxel; Phospholipid; Drug targeting; Surface modification;

A facile preparation method of a PFC-containing nano-sized emulsion for theranostics of solid tumors by Kouichi Shiraishi; Reiko Endoh; Hiroshi Furuhata; Masamichi Nishihara; Ryo Suzuki; Kazuo Maruyama; Yusuke Oda; Jun-ichiro Jo; Yasuhiko Tabata; Jun Yamamoto; Masayuki Yokoyama (379-387).
Theranostics means a therapy conducted in a diagnosis-guided manner. For theranostics of solid tumors by means of ultrasound, we designed a nano-sized emulsion containing perfluoropentane (PFC5). This emulsion can be delivered into tumor tissues through the tumor vasculatures owing to its nano-size, and the emulsion is transformed into a micron-sized bubble upon sonication through phase transition of PFC5. The micron-sized bubbles can more efficiently absorb ultrasonic energy for better diagnostic images and can exhibit more efficient ultrasound-driven therapeutic effects than nano-sized bubbles. For more efficient tumor delivery, smaller size is preferable, yet the preparation of a smaller emulsion is technically more difficult. In this paper, we used a bath-type sonicator to successfully obtain small PFC5-containing emulsions in a diameter of ca. 200 nm. Additionally, we prepared these small emulsions at 40 °C, which is above the boiling temperature of PFC5. Accordingly, we succeeded in obtaining very small nano-emulsions for theranostics through a very facile method.
Keywords: Theranostics; Tumor targeting; Ultrasound; Perfluorocarbon; Emulsion;

Facile synthesis of pH sensitive polymer-coated mesoporous silica nanoparticles and their application in drug delivery by Hongyan Tang; Jia Guo; Yang Sun; Baisong Chang; Qingguang Ren; Wuli Yang (388-396).
pH-responsive polymer shell chitosan/poly (methacrylic acid) (CS-PMAA) was coated on mesoporous silica nanoparticles (MSN) through the facile in situ polymerization method. The resultant composite microspheres showed a flexible control over shell thickness, surface charges and hydrodynamic size by adjusting the feeding amount of MSN and the molar ratio of [–NH2]/MAA. The MSN/CS-PMAA composite microspheres were stable in the pH range of 5–8 as well as in the physiological saline (0.15 M NaCl). Doxorubicin hydrochloride (DOX) was applied as a model drug to investigate the drug storage and release behavior. The results demonstrated that DOX could be effectively loaded into the composite microspheres. The cumulative release of DOX-loaded composite microspheres was pH dependent and the release rate was much faster at low pH (5.5) than that of pH 7.4. The cytotoxicity test by MTT assay showed that the blank carrier MSN/CS-PMAA microspheres were suitable as drug carriers. The cellular uptake of composite microspheres was investigated by confocal laser scanning microscopy (CLSM), which indicated that MSN/CS-PMAA could deliver the drugs into HeLa cell. The above results imply that the composite microspheres are a promising drug delivery system for cancer therapy.
Keywords: Mesoporous silica nanoparticle; pH sensitive; Composite microsphere; Drug delivery;

Heparin-coated superparamagnetic nanoparticle-mediated adeno-associated virus delivery for enhancing cellular transduction by Jun-Ho Hwang; Slgirim Lee; Eunmi Kim; Jung-Suk Kim; Chang-Ha Lee; Ik-Sung Ahn; Jae-Hyung Jang (397-404).
Superparamagnetic iron oxide nanoparticles (SPIONs) have been exploited as an elegant vehicle to enhance gene delivery efficiencies in gene therapy applications. We developed a magnetically guided adeno-associated virus (AAV) delivery system for enhancing gene delivery to HEK293T and PC12 cell lines. Wild-type AAV2 and a novel AAV vector, AAVr3.45, which was directly evolved in a previous study to possess diverse cell tropisms, were used as gene carriers. Additionally, the affinity of each viral vector to heparin was employed as a moiety to immobilize virus onto heparin-coated SPIONs (HpNPs). Magnetically guided AAV delivery resulted fast and efficient cellular transduction. Importantly, a short exposure of virus to target cells under a magnetic field (<180 min) yielded comparable transduction produced by the conventional gene-delivery protocol (i.e., 24 h-incubation of virus with target cells prior to replacing with fresh medium). Additionally, magnetic guidance of AAV encoding nerve growth factor (NGF) produced sufficient functional NGF, leading to robust neurite elongation by PC12 as compared to direct NGF protein delivery or non-magnetic delivery. The successful establishment of a magnetically guided AAV delivery system, with the ability to efficiently and rapidly infect target cells, will provide a powerful platform for a variety of gene therapy applications.
Keywords: Heparin; Adeno-associated virus; Iron oxide nanoparticle; Magnetism;