International Journal of Pharmaceutics (v.410, #1-2)

The purpose of this study was to assess the porosity variation of roller compacted ribbons made using different process parameters; in addition, the feasibility of using near-infrared chemical imaging (NIR-CI) to evaluate porosity variations was examined. Ribbons of neat microcrystalline cellulose were compacted using a range of roll pressures (RP), roll speeds (RS) and feed screw speeds (FSS). The ribbon porosity decreased as RP increased with the exception of ribbons produced by the combination of high RS and low FSS where increasing RP increases the porosity of the ribbons. Lower RS was found to produce ribbons with lower porosity and the porosity increases as the RS increased. Increased FSS will decrease ribbon porosity at higher RS while it slightly increase the ribbon porosity at lower RS. A simple linear regression model showed NIR-CI was able to predict the ribbon porosity with a correlation of 0.9258. NIR-CI is able to characterize differences in porosity as a function of position on the ribbon where regions with lower porosity show higher absorbance. Nevertheless, NIR-CI is able to show sinusoidal variation in intensities along the roller compacted ribbon among all settings studied.
Keywords: Roller compaction; NIR chemical imaging; Porosity; Density; Roller compacted ribbon; Feed screw speed, Roller pressure, Roller speed, Quality by design, NIR-CI;

Vinyl polymer-coated lorazepam particles for drug delivery to the airways by Matthew J. Traynor; Yanjun Zhao; Marc B. Brown; Stuart A. Jones (9-16).
A particle engineering method that adsorbs a microfine vinyl polymer coat to crystalline drug microparticles has been shown to be an effective way to control delivery. However, the means by which the functional performance of such microparticles is altered by the behaviour of the polymers in the microparticle coat remains unclear. The aim of this study was to determine the influence of vinyl polymer coating on the in vitro delivery characteristics of intranasal lorazepam microparticles. A series of four, similarly sized (ca. 10 μm), lorazepam-rich microparticles with different polymer coats were generated. The absorption of the polymer coats appeared to disrupt lorazepam solid state dimer formation in the microparticles, which manifested in a reduction in drug melting point. Mildly cohesive particles (aerodynamic diameter of 32 μm) that allowed rapid drug release (ca. 80% in 5 min) were generated when partially hydrolysed PVA dominated the microparticle coat, whilst fully hydrolysed PVA reduced particle cohesion and retarded drug release (ca. 15% release in 5 min). Infrared analysis showed that the properties of the microparticles were dictated by the strength of the hydrogen bonding in the polymer coat and not the strength of coat adsorption that was facilitated by hydrogen bond formation between the hydroxyl groups of the PVA and the hydroxyl group at position C3 of the lorazepam diazepine ring.
Keywords: Lorazepam; Intranasal delivery; Coating; Microparticles; Poly(vinyl alcohol); PVP;

Recently, many people have developed rheumatoid arthritis (RA), and prednisolone (PD) is often used for treatment; however, long use and a large dose of PD can cause toxic side effects. In this study, in order to enhance the therapeutic effects and to suppress the toxic side effects, the conjugate (GC–SP) was prepared by coupling between glycol-chitosan (GC) and succinyl-prednisolone (SP). The drug-release properties of GC–SP were examined and analyzed kinetically. The plasma concentration–time profiles of GC–SP and released PD were investigated after i.v. injection to normal rats, and their pharmacokinetic profiles were analyzed. PD was stable and released gradually (ca. 1%/h) from GC–SP at physiological pH, while PD was unstable at basic pH and the release from GC–SP was accelerated at basic pH. GC–SP showed good systemic retention (more than 16-fold area under the plasma concentration–time curve (AUC) as compared to PD alone), and released PD gradually in vivo. The in vivo release rate was calculated to be much faster than the in vitro rate. From these results, it is expected that GC–SP will be accumulated at inflammatory sites based on enhanced permeability and retention (EPR) effects, and release PD there effectively.
Keywords: Prednisolone; Glycol-chitosan; Conjugate; Drug release; Plasma concentration;

The stabilization mechanism of latanoprost by Akiko Ochiai; Kazumi Danjo (23-30).
The content of latanoprost is likely to decrease in solution because of the adsorption to eye drop containers and hydrolysis. We reduced these problems and established a formulation of latanoprost eye drops which is stable at room temperature. We assume that the additive surfactants form micelles and stabilize latanoprost in this formulation. In this study, we elucidated the latanoprost stabilization mechanism.It was revealed by Arrhenius analysis that the adsorption to eye drop containers and hydrolysis of latanoprost were temperature-dependent. In addition, polyethylene glycol monostearates inhibited the adsorption and hydrolysis of latanoprost at 1 mg/mL, which exceeded the critical micelle concentration. By the fluorescent probe method, it was suggested that the surfactants were associated with benzalkonium chloride and formed complex micelles consisting of about 10 molecules, and latanoprost interacted with the micelles at 1:1. By 1H NMR, it was revealed that adsorption was inhibited by arranging the hydrophobic group toward the center of complex micelles and that hydrolysis was inhibited by interaction between the ester group and the complex micelles.It was shown that the latanoprost is stabilized by the interaction with complex micelles. It was effective for the inhibition of both adsorption and degradation.
Keywords: Latanoprost; Adsorption; Hydrolysis; Stabilization mechanism; Micelle;

Design and evaluation of baicalin-containing in situ pH-triggered gelling system for sustained ophthalmic drug delivery by Haoyun Wu; Zhidong Liu; Junjie Peng; Lin Li; Nan Li; Jiawei Li; Hao Pan (31-40).
CSLM micrograph of FITC-labeled in situ pH-triggered gelling system. (A) In situ gelling system under pH 5.8, (B) After instilled distilled water and (C) After instilled 0.2 M NaOH and adjusted pH to 6.8.Baicalin has been reported to have anti-inflammatory and anti-cataract effects on eye tissues, but it has a low bioavailability partly due to its poor stability of baicalin, the special anatomic structure and efficient protective mechanism of eyes. The aim of this study was to investigate the correlation between the stability of baicalin and in situ pH-triggered gelling system. Carbopol® 974P (0.3%, w/v) was used as the gelling agent combined with hydroxypropylmethylcellulose E4M (0.6%, w/v) which acted as a viscosity enhancing agent. In vitro and in vivo evaluations were performed using several techniques, namely confocal scanning light microscopy analysis, rheometry, Gamma scintigraphic technique and microdialysis method. The rheological behavior showed a significant enhancement in gel strength under physiological conditions, and the formulation provided sustained release of the drug over an 8-h period. In elimination studies, the radioactivity of formulation was always higher than that of the control solution. Additionally, the AUC and C max values were 6.1-fold and 3.6-fold higher than those of the control solution, respectively. The results demonstrated that an in situ pH-triggered gelling system have better ability to keep baicalin stable and retain drug release than marketed baicalin eye drops to enhance the ocular bioavailability.
Keywords: Sustained ophthalmic delivery; In situ gelling system; Baicalin; Carbopol; Hydroxypropylmethylcellulose;

Design of an expert system for the development and formulation of push–pull osmotic pump tablets containing poorly water-soluble drugs by Zhi-hong Zhang; Hong-ye Dong; Bo Peng; Hong-fei Liu; Chun-lei Li; Min Liang; Wei-san Pan (41-47).
The purpose of this article was to build an expert system for the development and formulation of push–pull osmotic pump tablets (PPOP). Hundreds of PPOP formulations were studied according to different poorly water-soluble drugs and pharmaceutical acceptable excipients. The knowledge base including database and rule base was built based on the reported results of hundreds of PPOP formulations containing different poorly water-soluble drugs and pharmaceutical excipients and the experiences available from other researchers. The prediction model of release behavior was built using back propagation (BP) neural network, which is good at nonlinear mapping and learning function. Formulation design model was established based on the prediction model of release behavior, which was the nucleus of the inference engine. Finally, the expert system program was constructed by VB.NET associating with SQL Server. Expert system is one of the most popular aspects in artificial intelligence. To date there is no expert system available for the formulation of controlled release dosage forms yet. Moreover, osmotic pump technology (OPT) is gradually getting consummate all over the world. It is meaningful to apply expert system on OPT. Famotidine, a water insoluble drug was chosen as the model drug to validate the applicability of the developed expert system.
Keywords: Push–pull osmotic pump (PPOP); Expert system; Artificial neural network; Formulation design; Poorly water-soluble drugs;

Development of interferon alpha-2b microspheres with constant release by Zhiping Li; Lin Li; Yan Liu; Hui Zhang; Xueru Li; Fang Luo; Xingguo Mei (48-53).
Interferon alpha-2b was released from microspheres with a zero-order profile except the burst release both in vitro and in vivo.Interferon alpha-2b (IFNα-2b) is an important immune regulator used widely in clinic. However, frequent subcutaneous injection and substantial toxicity decrease patients’ compliance. So, drug delivery with more precisely controlled drug release is urgent for IFNα-2b. Microsphere is a promising sustained drug delivery system, which has been studied widely for delivery of proteins. However, it was found difficult to keep proteins’ activity and guarantee complete release. In this study, we solidified IFNα-2b as microparticles firstly by co-lyophilizing it with gelatin and ZnSO4. Microspheres were then prepared. The preparing procedure and formulation were optimized with encapsulation efficiency and in vitro release as main parameters. Finally, the microspheres were prepared by S/O/W method with microparticle size about 5 μm and PEGT/PBT-PLGA (9:1, w/w) as matrix material. The diameter of microspheres was 28.94 μm, the encapsulation efficiency was 86.01%, the burst release was 16.69%, the cumulative release was 83.06% at 23th day, and IFNα-2b was released from microspheres with a zero-order profile. These microspheres also demonstrated sustained and steady release for about 13 days in rats. In conclusion, the procedure and formulation used in this study were supposed to be successful to keep IFNα-2b active and released constantly and completely.
Keywords: Interferon alpha-2b; Microspheres; Zero-order; Complete release;

Inhaled powder formulation of a new VIP derivative, [R15, 20, 21, L17, A24,25, des-N28]-VIP-GRR (IK312548), attenuated inflammatory symptoms in experimental asthma/COPD model rats as evidenced by a decrease of infiltrated granulocytes.Vasoactive intestinal peptide (VIP) has been considered as a promising drug candidate for asthma and COPD because of its potent immunomodulating and anti-inflammatory activities. Recently, our group developed a new VIP derivative, [R15, 20, 21, L17, A24,25, des-N28]-VIP-GRR (IK312548), with improved chemical and metabolic stability. In the present study, a dry powder inhaler system of IK312548 was designed for inhalation therapy with minimal systemic side effects, the physicochemical properties of which were also evaluated with a focus on morphology, particle size distribution, inhalation performance, and peptide stability. Laser diffraction and cascade impactor analysis suggested high dispersion and deposition in the respiratory organs with a fine particle fraction of 31.2%. According to UPLC/ESI-MS and circular dichroic spectral analyses, no significant changes in the purity and structure of VIP derivative were observed during preparation of respirable formulation. Anti-inflammatory properties of IK312548 respirable powder (RP) were characterized in antigen-sensitized asthma/COPD-model rats. There were marked inflammatory cells infiltrated into the lung tissues of experimental asthma/COPD-model rats; however, intratracheal administration of IK312548-RP led to significant reductions of recruited inflammatory cells in lung tissues and BALF by 72 and 78%, respectively. Thus, respirable powder formulation of IK312548 might be a promising medication for asthma, COPD, and other airway inflammatory diseases.
Keywords: Asthma; Chronic obstructive pulmonary disease; Dry powder inhaler; Vasoactive intestinal peptide;

This study evaluated the physical stability and molecular mobility of a poorly water-soluble amorphous drug, 2-benzyl-5-(4-chlorophenyl)-6-[4-(methylthio)phenyl]-2H-pyridazin-3-one (K-832), adsorbed onto silica mesopores. K-832–Sylysia 740 and K-832–Sylysia 350 formulations, prepared by adsorbing K-832 onto porous silica Sylysia 740 (2.5-nm-diameter pores) and Sylysia 350 (21-nm-diameter pores) and stored at 60 °C/80%RH (open and closed conditions), were investigated. Differential scanning calorimetry revealed that crystallization of K-832 in the K-832–Sylysia 350 formulation stored at 60 °C/80%RH (open and closed conditions) was faster than that of the other formulation stored under identical conditions. Raman spectroscopy revealed shifts to higher wavenumbers in the K-832–Sylysia 350 and K-832–Sylysia 740 formulations (1497 and 1493 cm−1, respectively) in comparison to amorphous K-832 (1481 cm−1); however, no distinct differences were observed in the spectra of the two formulations. Solid-state 13C NMR spectroscopy revealed a difference in spin–lattice relaxation time in the rotating frame (T ) between the two formulations, suggesting the lower molecular mobility of K-832 in the 2.5-nm-diameter pores than in the 21-nm-diameter pores. Thus, the crystallization rate of amorphous K-832 in the K-832–Sylysia 740 formulation was much slower. These results will be useful in estimating the physical stability of amorphous drugs in mesopores.
Keywords: Porous silica; Amorphous; Relaxation time; Solid-state NMR; Raman spectroscopy;

Danshen (Salvia miltiorrhiza) contains tanshinones, which inhibit P-glycoprotein (P-gp) and the cytochrome P450 (CYP) system. In the present study, we evaluated the possible pharmacokinetic interactions of Danshen extract with docetaxel and clopidogrel in rats. Docetaxel (5 mg/kg intravenously and 40 mg/kg orally) or clopidogrel (30 mg/kg orally) was administered to rats with or without oral co-administration of Danshen (400 mg/kg). Co-administration of Danshen did not affect the plasma concentration profiles and pharmacokinetic parameters of docetaxel and clopidogrel, whereas cyclosporine A, a P-gp and CYP3A inhibitor, significantly influenced the pharmacokinetics of co-administered docetaxel and clopidogrel. Orally administered Danshen had no substantial effect on the pharmacokinetics of docetaxel and clopidogrel, suggesting the negligible safety concern of Danshen in P-gp- and CYP3A-mediated interactions in vivo.
Keywords: Danshen; Docetaxel; Clopidogrel; Pharmacokinetic interaction; P-glycoprotein; Cytochrome P450 3A;

Protective effect of drug delivery systems against the enzymatic degradation of dermally applied DNAzyme by Thomas Schmidts; Dorota Dobler; Sylvia von den Hoff; Peggy Schlupp; Holger Garn; Frank Runkel (75-82).
DNAzymes are a group of RNA-cleaving DNA oligonucleotides that contain a catalytic domain and represent a novel class of antisense molecules. Although single-stranded DNAzymes may represent the most effective nucleic acid drug to date, the sensitivity to nuclease degradation is challenging. Therefore, it is important to develop a drug delivery system, which protects the molecule against degradation during dermal application. In the present study, the potential protective effect, regarding the dermal application of DNAzyme, of multiple (W/O/W) emulsions, W/O emulsions, submicron emulsion and microemulsions were investigated using a HPLC method. The HPLC method enables the quantitative analysis of DNAzyme as well as the detection of degradation products. The differences between the activity of DNase I and the activity of nucleases located in the porcine skin were compared. It was found that the degradation of an aqueous solution of DNAzyme is depending on the DNase I activity as well as on the incubation time. Furthermore, the activity of neutral and acid nucleases in skin tissue was determined to be 5.2 and 14.8 U per 1 g of porcine skin tissue, respectively. Investigation of the protective character of different delivery systems revealed that formulations containing DNAzyme in the outer water phase (submicron emulsion and microemulsion) did not exhibit any form of protective effect, whereas formulations containing DNAzyme in the inner water phase (multiple emulsion and W/O emulsion) were able to prevent the DNAzyme degradation to a considerable degree. Consequently, these formulations are promising candidates for the dermal drug delivery of oligonucleotides.
Keywords: DNAzyme; Oligonucleotide; DNase; HPLC; W/O/W multiple emulsion; Submicron emulsion; Microemulsion;

pH triggered injectable amphiphilic hydrogel containing doxorubicin and paclitaxel by Lingling Zhao; Lijun Zhu; Fuyong Liu; Chenyang Liu; Shan-Dan; Qian Wang; Chengliang Zhang; Jiaoli Li; Jiguang Liu; Xiaozhong Qu; Zhenzhong Yang (83-91).
Injectable hydrogel with hydrophobic microdomains for incorporating both hydrophilic and hydrophobic drugs, herein doxorubicin hydrochloride (DOX) and paclitaxel (PTX), was synthesized through dynamic bonding of glycol chitosan and benzaldehyde capped poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol) via Schiff's reaction triggered by environmental pH. Rheology tests show that the inclusion of hydrophilic drug decreases the gelation time and gains more robust gel, while the addition of hydrophobic drug has opposite influences. Dual-drug release from the DOX + PTX loaded gels was observed and the release rate can be accelerated by decreasing the environmental pH from physiological (7.4) to weak acidic pH (6.8). In vivo investigation proved that the gels were able to diminish the amount of DOX in blood circulation and limit the DOX-induced cardiotoxicity. By intratumoral administration, the hydrogel-drug formulations resulted in efficient growth inhibition of subcutaneous tumor (B16F10) on C57LB/6 mouse model. The advantage of the current system for DOX + PTX combination therapy was demonstrated by a prolongation of survival time in comparison with the single drug therapy.
Keywords: Injectable gel; pH triggered; Combination delivery; Cardiotoxicity; Antitumor activity;

Original predictive approach to the compressibility of pharmaceutical powder mixtures based on the Kawakita equation by Vincent Mazel; Virginie Busignies; Stéphane Duca; Bernard Leclerc; Pierre Tchoreloff (92-98).
In the pharmaceutical industry, tablets are obtained by the compaction of two or more components which have different physical properties and compaction behaviours. Therefore, it could be interesting to predict the physical properties of the mixture using the single-component results. In this paper, we have focused on the prediction of the compressibility of binary mixtures using the Kawakita model. Microcrystalline cellulose (MCC) and l-alanine were compacted alone and mixed at different weight fractions. The volume reduction, as a function of the compaction pressure, was acquired during the compaction process (“in-die”) and after elastic recovery (“out-of-die”). For the pure components, the Kawakita model is well suited to the description of the volume reduction. For binary mixtures, an original approach for the prediction of the volume reduction without using the effective Kawakita parameters was proposed and tested. The good agreement between experimental and predicted data proved that this model was efficient to predict the volume reduction of MCC and l-alanine mixtures during compaction experiments.
Keywords: Binary mixtures; Compaction; Compressibility; Excipients; Kawakita equation; Modeling;

Variability in commercial carbamazepine samples – Impact on drug release by Felicia Flicker; Veronika Anna Eberle; Gabriele Betz (99-106).
The aim of this study was to characterize the variability of commercial carbamazepine (CBZ) samples and to investigate the influence of two commonly used tablet fillers, i.e., mannitol and microcrystalline cellulose (MCC) on the CBZ sample variability. Polymorphism and morphology of CBZ samples were analyzed by differential scanning calorimetry, X-ray powder diffraction, sieve analysis, and scanning electron microscopy. Drug release from CBZ samples and binary mixtures (30–90% drug load) was characterized by a unidirectional dissolution method measuring disk intrinsic dissolution rate (DIDR) and drug release, respectively. All CBZ samples were of p-monoclinic form but differed in their polymorphic purity, particle size, morphology and intrinsic dissolution rate. Two characteristic inflection points, determined in the DIDR profiles, characterized the specific transformation behavior of each CBZ sample. The variability in CBZ samples was also exhibited in the drug release profiles from their binary mixtures. Mannitol increased initial drug release of CBZ samples up to 10-fold in mixtures of 30% drug load. The presence of MCC resulted in reduced variability in drug release. The unidirectional dissolution method is presented as a straightforward monitoring tool to characterize variability of CBZ raw materials and effect of tablet fillers.
Keywords: Carbamazepine; Dihydrate; Preformulation; Morphology; Polymorphism; Tablet filler;

Applicability of sucrose laurate as surfactant in solid dispersions prepared by melt technology by Angéla Szűts; Péter Láng; Rita Ambrus; Lóránd Kiss; Mária A. Deli; Piroska Szabó-Révész (107-110).
This study focused on an investigation of the applicability of sucrose laurate as surfactant in solid dispersions. Although this surfactant has a US Drug Master File, it has not been used so far in internal pharmaceutical products. High drug-loaded solid dispersion systems consisting of gemfibrozil as a model drug and PEG 6000 as a carrier, with or without sucrose laurate (D1216), were prepared by the melting method. Cytotoxicity studies on Caco-2 monolayer cells were also performed, in order to gain information on the applicability of D1216 in oral formulations. The results showed that the presence of the surface-active agent did not affect the solid-state characteristics of the model drug significantly. A markedly improved dissolution of gemfibrozil from the ternary solid dispersion systems was observed as compared with the binary solid dispersion systems. The optimum concentration range of the D1216 in the formulations was determined to be 5–10%. The effective final concentrations of D1216 in the dissolution experiments proved to be non-toxic towards CaCo-2 cells. The results suggest the potential use of D1216 in innovative internal pharmaceutical formulations.
Keywords: Sucrose laurate; Surfactant; Solid dispersion; Melt technology; Cytotoxicity; Caco-2 cells; Gemfibrozil; PEG 6000;

New pH-sensitive controlled release system of EGCG using Eudragit® S100 is expected to be effective to avoid gastric degradation and improve metabolic stability in the intestinal tract after oral administration.The main purpose of the present investigation was to develop a novel enteric mucoadhesive formulation of (−)-epigallocatechin-3-gallate (EGCG), a green tea catechin, with the aim of avoiding its severe degradation in the gastrointestinal tract and therefore improving pharmacological effects. The EGCG-loaded microspheres (EGCG/MS), containing Eudragit® S100, were prepared with an emulsion solvent diffusion method in aqueous PVA solution. The EGCG/MS with a diameter of 16 μm exhibited pH-dependent controlled release of EGCG with limited initial burst release, and the Eudragit® S100-based MS also had moderate bioadhesive property in isolated small intestine of rats. There appeared to be marked degradation of EGCG in acidic solution (pH 1.2) and neutral buffer (pH 6.8) containing intestinal microsomal fraction, although significant improvement in chemical and metabolic stability of EGCG was observed in the EGCG/MS, possibly due to the controlled release and/or bioadhesion. From these findings, newly prepared EGCG/MS might be of clinical importance in both stabilizing and delivering EGCG for treatment of intestinal diseases.
Keywords: EGCG; Microspheres; Controlled release; Eudragit S100; Stability;

α-Glucosyl hesperidin induced an improvement in the bioavailability of pranlukast hemihydrate using high-pressure homogenization by Hiromasa Uchiyama; Yuichi Tozuka; Fusatoshi Asamoto; Hirofumi Takeuchi (114-117).
The α-glucosyl hesperidin (Hsp-G)-induced improvement of both the dissolution and absorption properties of pranlukast hemihydrate (PLH) was achieved by means of a high-pressure homogenization (HPH) processing. The average particle size in the HPH-processed suspension was decreased significantly after 50 cycles of processing and reached a constant size of ca. 300 nm. The amount of dissolved PLH gradually increased with the pass number of HPH processing, and was extremely higher than the PLH solubility (0.8 μg/mL at 37 °C) after the HPH processing. On a dissolution study of the freeze-dried sample of HPH-processed PLH/Hsp-G (1/10), the apparent solubility of PLH was at least 2.5-fold more than that of untreated PLH crystals. The transport study showed that the amount of PLH that had permeated through the Caco-2 cell monolayers was improved in the case of HPH-processed PLH/Hsp-G (1/10). The bioavailability of PLH from HPH-processed PLH/Hsp-G (1/10) showed a 3.9- and 2.2-fold improvement over the PLH crystal in terms of C max and AUC values, respectively. Hsp-G formed an associated structure in aqueous media. High-pressure homogenization provides a good opportunity for molecular-level interaction of PLH and the associated structure of Hsp-G to occur. The use of Hsp-G under HPH processing was a promising way to enhance the dissolution and absorption of PLH without using an organic solvent.
Keywords: α-Glucosyl hesperidin; Pranlukast hemihydrate; High-pressure homogenization; Dissolution enhancement; Absorption enhancement;

The purpose of this study was to develop spherical mesocellular foam (MCF) loaded with a poorly water-soluble drug, intended to be orally administered, able to improve the dissolution rate and enhance the drug loading capacity. Spherical MCF with a continuous 3-D pore system was synthesized using Pluronic 123 triblock polymer (P123) as a surfactant coupled with cetyltrimethyl ammonium bromide (CTAB) as a co-surfactant. A model drug, simvastatin (SV), was loaded onto spherical MCF via a procedure involving a combination of adsorption equilibrium and solvent evaporation. The drug release rate and the drug loading efficiency of spherical MCF were compared with those of fibrous SBA-15. Investigations using nitrogen adsorption, scanning electron microscopy (SEM), transmission electron microscopy (TEM), powder X-ray diffraction (PXRD), differential scanning calorimetry (DSC), thermogravimetric analysis (TGA) and HPLC demonstrated the successful incorporation of SV into the MCF host. It was found that spherical MCF has a high drug loading efficiency up to 37.5%, and higher than that of fibrous SBA-15 with a pore diameter of 6.5 nm. It is worth noting that fast release rate of SV was obtained from spherical MCF compared with SBA-15 and pure crystalline SV using enzyme-free simulated intestinal fluid (SIF, pH 6.8).
Keywords: Drug delivery; Mesocellular foam; Poorly water-soluble drugs; Simvastatin; Drug loading; Enhanced dissolution;

Spermine grafted galactosylated chitosan for improved nanoparticle mediated gene delivery by Susan M. Alex; M.R. Rekha; Chandra P. Sharma (125-137).
Despite multitude of beneficial features, chitosan has poor water solubility and transfection ability which affect its gene delivery efficacy. The two features are improved when certain chemical modifications are incorporated into the chitosan parent backbone. This strategy is adopted here, by coupling galactose and spermine into the chitosan backbone. The conjugation was determined with FTIR and 1H NMR and nanoparticle morphology was assessed by TEM and AFM techniques. Particle size, zeta potential, buffering capacity and DNA binding ability gave encouraging result of enhanced solubility and stability. In vitro studies of GCSM in HepG2 cell lines displayed low cytotoxicity and improved transfection. We also identified the preference of receptor mediated internalization for nanoparticles cellular uptake by treating with cellular uptake inhibitors. The results evidently led us to comprehend that galactosylated chitosan-g-spermine could be considered as a promising chitosan derivative for conducting nanoparticle mediated gene delivery.
Keywords: Nanoparticle; Spermine; Endocytosis; Galactosylated chitosan; Gene delivery;

Pharmacokinetics, tissue distribution and relative bioavailability of puerarin solid lipid nanoparticles following oral administration by Cheng-Feng Luo; Mu Yuan; Min-Sheng Chen; Shi-Ming Liu; Liu Zhu; Bi-Yun Huang; Xia-Wen Liu; Wen Xiong (138-144).
Puerarin has various pharmacological effects; however, poor water-solubility and low oral bioavailability limit its clinical utility. A delivery system of solid lipid nanoparticles could enhance its oral absorption. The objective of this study was to investigate the pharmacokinetics, tissue distribution and relative bioavailability of puerarin in rats after a single dose intragastric administration of puerarin solid lipid nanoparticles (Pue-SLNs). The puerarin concentrations in plasma and tissues were determined by rapid resolution liquid chromatography electrospray ionization–tandem mass spectrometry. The C max value of puerarin after the administration of Pue-SLNs was significantly higher than that obtained with puerarin suspension (0.33 ± 0.05 μg/mL vs. 0.16 ± 0.06 μg/mL, P  < 0.01). The T max value after the administration of the Pue-SLNs was significantly shorter than that after puerarin suspension administration (40 ± 0 min vs. 110 ± 15.49 min, P  < 0.01). The AUC0→t values of puerarin were 0.80 ± 0.23 mg h/L, and 2.48 ± 0.30 mg h/L after administration of the puerarin suspension and Pue-SLNs, respectively. Following administration of the Pue-SLNs, tissue concentrations of puerarin also increased, especially in the target organs such as the heart and brain. These data suggest that SLNs are a promising delivery system to enhance the oral bioavailability of puerarin.
Keywords: Puerarin; Solid lipid nanoparticles; Oral bioavailability; Pharmacokinetics; Rat; Liquid chromatography tandem mass spectrometry;

Chitosan nanoparticles as a dual growth factor delivery system for tissue engineering applications by Merlin Rajam; S. Pulavendran; Chellan Rose; A.B. Mandal (145-152).
Growth factors are essential in cellular signaling for migration, proliferation, differentiation and maturation. Sustainable delivery of therapeutic as well as functional proteins is largely required in the pharmacological and regenerative medicine. Here we have prepared chitosan nanoparticles (CNP) and incorporated growth factors such as epidermal growth factor (EGF) and fibroblast growth factor (FGF), either individually or in combination, which could ultimately be impregnated into engineered tissue construct. CNP was characterized by Fourier transform infrared (FTIR) spectroscopy, Zeta sizer and high resolution transmission electron microscope (HRTEM). The particles were in the size range of 50–100 nm with round and flat shape. The release kinetics of both EGF and FGF incorporated CNP showed the release of growth factors in a sustained manner. Growth factors incorporated nanoparticles did not show any toxicity against fibroblasts up to 4 mg/ml culture medium. Increased proliferation of fibroblasts in vitro evidenced the delivery of growth factors from CNP for cellular signaling. Western blotting results also revealed the poor inflammatory response showing less expression of proinflammatory cytokines such as IL-6 and TNFα in the macrophage cell line J774 A-1.
Keywords: Chitosan nanoparticle; Growth factors; Cytocompatibility; Proinflammatory response; Tissue engineering;

Agitation during lipoplex formation improves the gene knockdown effect of siRNA by Jose Mario Barichello; Shinji Kizuki; Tatsuaki Tagami; Tomohiro Asai; Tatsuhiro Ishida; Hiroshi Kikuchi; Naoto Oku; Hiroshi Kiwada (153-160).
The successful delivery of therapeutic siRNA to the designated target cells and their availability at the intracellular site of action are crucial requirements for successful RNAi therapy. In the present study, we focused on the siRNA-lipoplex preparation procedure and its effect on the gene-knockdown efficiency of siRNA in vitro. Agitation (vortex-mixing) during siRNA-lipoplex (vor-LTsiR) preparation and its effect on the gene-knockdown efficiency of stably expressed cell GFP was investigated, and their efficiency was compared with that of spontaneously formed lipoplex (spo-LTsiR). A dramatic difference in size between lipoplexes was observed at the N/P ratio of 7.62 (siRNA dose of 30 nM), even though both lipoplexes were positively charged. With the siRNA dose of 30 nM, vor-LTsiR accomplished a 50% gene-knockdown, while spo-LTsiR managed a similar knockdown effect at the 120 nM level, suggesting that the preparation procedure remarkably affects the gene-knockdown efficacy of siRNA. The uptake of vor-LTsiR was mainly via clathrin-mediated endocytosis, whereas that of spo-LTsiR was via membrane fusion. In addition, by inhibiting clathrin-mediated endocytosis, the gene-knockdown efficiency was significantly lowered. The size of the lipoplex, promoted by the preparation procedure, is likely to define the entry pathway, resulting in an increased amount of siRNA internalized in cells and an enhanced gene-knockdown efficacy. The results of the present study definitively show that a proper siRNA-lipoplex preparation procedure makes a significant contribution to the efficiency of cellular uptake, and thereby, to the gene-knockdown efficiency of siRNA.
Keywords: siRNA; Cationic liposome; Complex formation; Gene silencing; Vortex-mixing;

Chitosan enhances transfection efficiency of cationic polypeptides/DNA complexes by Praneet Opanasopit; Jintana Tragulpakseerojn; Auayporn Apirakaramwong; Tanasait Ngawhirunpat; Theerasak Rojanarata (161-168).
The aim of this research was to investigate the effect of cationic polypeptides mixed with chitosan (CS) on in vitro transfection efficiency and cytotoxicity in human cervical carcinoma cells (HeLa cells). The polypeptides/DNA complexes and ternary complexes (CS, polypeptides and DNA) at varying weight ratios were formulated and characterized by using gel electrophoresis. Their particle sizes and charge were evaluated. The effect of the type and molecular weight (MW) of polypeptides, the weight ratio, order of mixing, the pH and serum on transfection efficiency and cytotoxicity were evaluated in HeLa cells. Three types of polypeptides (poly-l-lysine; PLL, poly-l-arginine; PLA and poly-l-ornithine; PLO) were able to form complete complex with DNA at weight ratio above 0.1. The PLA MW >70 kDa showed the highest transfection efficiency. The order of mixing between CS, PLA and DNA affected the transfection efficiency. The highest transfection efficiency was observed in ternary complexes of PLA/DNA/CS (2:1:4) equal to PEI/DNA complex. For cytotoxicity studies, over 80% the average cell viabilities of the complexes were observed by MTT assay. This study suggests that the addition of CS to PLA/DNA is easy to prepare, safe and exhibits significantly improved DNA delivery potential in vitro.
Keywords: Cationic polypeptides; Poly-l-arginine; Chitosan; Gene delivery;

Improved solubility and pharmacokinetics of PEGylated liposomal honokiol and human plasma protein binding ability of honokiol by Xian-Huo Wang; Lu-Lu Cai; Xiao-Yan Zhang; Lin-Yu Deng; Hao Zheng; Chong-Yang Deng; Jiao-Lin Wen; Xia Zhao; Yu-Quan Wei; Li-Juan Chen (169-174).
PEGylated liposomal honokiol had been developed with the purpose of improving the solubility and pharmacokinetics compared with free honokiol. Human plasma protein binding ability of honokiol was also investigated. PEGylated liposomal honokiol was prepared by thin film evaporation-sonication method. Its mean particle size was 98.68 nm, mean zeta potential was −20.6 mV and encapsulation efficiency were 87.68 ± 1.56%. The pharmacokinetics of PEGylated liposomal honokiol was studied after intravenous administration in Balb/c mice. There were significant differences of parameters T1/2β and AUC0→∞ between them and liposome lengthened T1/2β and AUC0→∞ values. The mean T1/2β value of PEGylated liposomal honokiol and free honokiol were 26.09 min and 13.46 min, respectively. The AUC0→∞ ratio of PEGylated liposomal honokiol to free honokiol was about 1.85-fold (219.24 μg/mL min/118.68 μg/mL min) (P  = 0.000). Examination of protein binding ability showed that honokiol with 0.5, 8.0 and 20 μg/mL concentrations in human plasma achieved the percent of bound between 60% and 65%. The results suggested that PEGylated liposomal honokiol improved the solubility, increased the drug concentration in plasma, and withstanded the clearance. Besides, the percent of protein bound of honokiol in human plasma was between 60% and 65%.
Keywords: Honokiol; PEGylated liposome; Solubility; Pharmacokinetics; Human plasma protein binding;

Controlled antisolvent precipitation of spironolactone nanoparticles by impingement mixing by Yuancai Dong; Wai Kiong Ng; Shoucang Shen; Sanggu Kim; Reginald B.H. Tan (175-179).
Continuous antisolvent precipitation of spironolactone nanoparticles were performed by impingement mixing in this work. In the range of Reynolds numbers (Re) 2108–6325 for the antisolvent water stream and 1771–5313 for the solvent stream, i.e. acetonic drug solution, 302–360 nm drug nanoparticles were achieved. Increasing drug concentration from 25 to 50 and 100 mg/ml led to a significant size increase from 279.0 ± 2.6 to 302.7 ± 4.9 and 446.0 ± 17.3 nm, respectively. “Two-step crystallization” was first observed for spironolactone in the water/acetone system: the drug was precipitated initially as spherical cluster, which rearranged into ordered cuboidal nanocrystals finally. The nanoformulation showed faster dissolution rate in comparison with the raw drug. By combining the impingement mixing and an on-line spray drying, a fully continuous process may be developed for mass-production of dried drug nanoparticles.
Keywords: Spironolactone; Nanoparticles; Antisolvent precipitation; Impingement mixing; Oral bioavailability;

Injectable nimodipine-loaded nanoliposomes: Preparation, lyophilization and characteristics by Tingting Guan; Yuqiang Miao; Lishuang Xu; Shenshen Yang; Jing Wang; Haibing He; Xing Tang; Cuifang Cai; Hui Xu (180-187).
The main purpose of this study was to prepare nimodipine-loaded nanoliposomes for injection and evaluate their characteristics after lyophilization. Nimodipine-loaded nanoliposomes were prepared by the emulsion-ultrasonic method with sodium cholesterol sulfate (SCS) as the regulator and then lyophilized by adding different cryoprotectants. SCS was used as a blender of regulator and surfactant and helped to prepare smaller liposomes due to the steric hindrance of the sulfate group. The results showed that nimodipine-loaded nanoliposomes with a 20:1 of egg yolk lecithin PL-100M vs. SCS ratio had a particle size of 86.8 ± 42.007 nm, a zeta potential of −13.94 mV and an entrapment efficiency (EE) of 94.34% and could be stored for 12 days at 25 °C. Because of the good bulking effect of mannitol and the preservative effect of trehalose, they were used to obtain suitable lyophilized nanoliposomes. The lyophiles containing 10% mannitol and 20% trehalose had a good appearance and a slightly altered particle size after rehydration. In addition, the lyophilized products were characterized by differential scanning calorimetry, X-ray diffraction and scanning electron microscopy, which confirmed the morphous state of trehalose, mannitol and the mixture. Trehalose could inhibit mannitol crystallization to some extent. The drug release from nanoliposomes before and after lyophilization in pH 7.4 phosphate buffer containing 30% ethanol was also examined and both profiles were found to fit the Viswanathan equation. This means that the drug release was controlled by the pore diffusion resistance.
Keywords: Nimodipine; Nanoliposomes; Sodium cholesterol sulfate; Lyophilization; Cryoprotectants; In vitro release;

A model poorly aqueous-soluble drug, ibuprofen (IBU), was co-spray dried with mesoporous silica materials having different pore sizes and particle sizes for dissolution enhancement. Drug molecules were entrapped inside the mesoporous channels at a high drug loading of 50:50 (w/w). The pore sizes were found to affect the physical state and particle size of IBU in mesoporous structures, which influenced the dissolution profiles. When IBU was co-spray dried with MCM-41 and SBA-15 with pore size smaller than 10 nm, amorphous state of IBU was obtained due to nano space confinement. In contrast, nanocrystals were obtained when ibuprofen was co-spray dried with large pore SBA-15-LP with pore size above 20 nm. The physical state of ibuprofen played a key role in affecting the dissolution of IBU from the solid dispersion. IBU in the amorphous state exhibited a higher dissolution rate than nanocrystalline IBU, even though the larger pore size could facilitate diffusion from the host matrix. The particle size of mesoporous silica showed a less pronounced effect on the dissolution of IBU. Thus, the amorphous/nanocrystalline state of ibuprofen was the most important influence on drug dissolution followed by the diffusion kinetics, particle size of IBU and path length from host matrix to dissolution medium.
Keywords: Nanocrystals; Amorphous; Co-spray drying; Mesoporous silica; Dissolution;

Molecular modeling of gel nanoparticles with cyclosporine A for oral drug delivery by Jonáš Tokarský; Tomáš Andrýsek; Pavla Čapková (196-205).
Structure and behavior of amphiphilogel nanoparticles as drug carriers for cyclosporine A have been studied by the molecular modeling using empirical force field.Structure and behavior of amphiphilogel nanoparticles as a drug carriers for cyclosporine A (CsA) have been studied by the molecular modeling using empirical force field. Five atomistic models of a gel-based emulsions (GEM) with various gel compositions have been investigated in order to find a system most similar to a sixth atomistic model of self-microemulsifying drug delivery system (SMEDDS) taken as an exemplar of CsA delivery system. Structural parameters and energy characteristics (i.e. non-bond interaction energy between CsA and whole remaining components of a gel nanoparticle, CsA/gel nanoparticle intermolecular non-bond interaction energy, CsA–gel molecule pair interaction energy, volume fraction, concentration profiles and number of pervaded water molecules) of these six models in a waterless form and in a water containing form have been studied in dependence on the composition. The Flory–Huggins theory as implemented in the Accelrys Materials Studio 4.2 modeling environment was used to study the pair interactions of cyclosporine A with various types of surfactants. Structural parameters and energy characteristics of all systems have been compared and one composition was selected as a very promising for further experimental study.
Keywords: Cyclosporine A; Molecular modeling; Drug delivery; Structure characterization;

In this article, inulin and poly(acrylic acid) grafted inulin copolymer were used to enhance the dissolution of poorly water-soluble Irbesartan drug and to control its drug release rate, respectively. Topological structure of inulin showed sleazy separable flower-like platelets and granules accumulated above each other, which adapt it to physically bind Irbesartan drug and enhance its dissolution. Consequently, the increase of inulin content in the polymeric matrix was found to increase the drug dissolution gradually until it reaches its maximum (∼90%) within the first 60 min. The release rate had followed zero-order transport mechanism. On the other hand, the poly(acrylic acid) grafted inulin copolymer, characterized using 1H NMR, FTIR, TGA, and SEM techniques, was found to form highly consistent amorphous systems of two-dimensional surfaces with some voids topology. Such features adapted it to control Irbesartan drug dissolution (∼33%) and show Fickian diffusion mechanism.
Keywords: Inulin; Poly(acrylic acid) grafted inulin; Poorly water-soluble drug; Control of drug release; Irbesartan;