International Journal of Pharmaceutics (v.468, #1-2)

Tamoxifen nanostructured lipid carriers: Enhanced in vivo antitumor efficacy with reduced adverse drug effects by Harshad K. Shete; Nilakash Selkar; Geeta R. Vanage; Vandana B. Patravale (1-14).
Display OmittedA novel approach of enhancing the Tamoxifen uptake via Intestinal Lymphatic System is executed by developing long chain lipid and oil based nanostructured lipid carrier system (Tmx-NLC). The aim was to achieve improved systemic bioavailability of Tamoxifen, prevent systemic and hepatotoxicity and enhance antitumor efficacy. Following the proof of concept achieved in cell culture experiments and in vivo pharmacokinetic and biodistribution study, the current work focuses on investigation of antitumor efficacy and treatment associated toxicity in murine mammary tumor mice model. The efficacy study demonstrated greater tumor suppression and 100% survival with 1.5 and 3 mg/kg Tmx-NLC compared to 3 mg/kg Tamoxifen suspension and Mamofen® (Khandelwal Pharmaceuticals, Mumbai, India). Tmx-NLC treatment for a month demonstrated improved systemic toxicity profile and no evidences of hepatotoxicity. Thus, developed Tmx-NLC could prove to be a promising delivery strategy to confer superior therapeutic efficacy and ability to address the biopharmaceutical and toxicity associated issues of drug.
Keywords: Tamoxifen; Nanostructured lipid carriers; Intestinal lymphatic system; Murine model; Hepatotoxicity;

Direct comparison of two albumin-based paclitaxel-loaded nanoparticle formulations: Is the crosslinked version more advantageous? by Chunlei Li; Yanhui Li; Yuqing Gao; Na Wei; Xi Zhao; Caixia Wang; Yongfeng Li; Xian Xiu; Jingxia Cui (15-25).
Display OmittedNon-crosslinked albumin-bond paclitaxel nanoparticles were unstable, but could deliver more paclitaxel into tumor than the crosslinked (A), resulting in enchanced efficacy (B).Nanoparticles using albumin as particle matrix have entered the mainstream of drug delivery. It was reported that non-crosslinked albumin nanoparticles were unstable in circulation and could deliver drugs into tumor through gp60/SPARC pathway; in contrast, the delivery of drugs with stable nanoparticles was dependent on enhanced permeability and retention effect. Thus, it is questionable which kind of nanoparticles was more advantageous. Two versions of albumin-bound paclitaxel nanoparticles were prepared. In vitro, the non-crosslinked particles could rapidly disintegrate and the crosslinked was stable. The pharmacokinetics of both formulations was different especially at early time and the non-crosslinked particles were cleared rapidly. After non-crosslinked particle treatment paclitaxel had a tendency to accumulate into heart and kidney and following therapy with the crosslinked particles, paclitaxel was liable to be delivered into lung, spleen and liver. The delivery efficiency of paclitaxel into tumor following the non-crosslinked particle treatment was greater than that of the crosslinked (p  < 0.05), thus resulting in a considerably improved antineoplastic activity. Moreover, the non-crosslinked formulation was only slightly more toxic. It was concluded that the non-crosslinked formulation was more advantageous for the delivery of paclitaxel and our conclusion might be generalized to other lipophilic drugs delivered with albumin nanoparticles.
Keywords: Nanoparticles; Paclitaxel; Albumin; Stability; Pharmacokinetics; Efficacy;

Increased tumor targeted delivery using a multistage liposome system functionalized with RGD, TAT and cleavable PEG by Ling Mei; Ling Fu; Kairong Shi; Qianyu Zhang; Yayuan Liu; Jie Tang; Huile Gao; Zhirong Zhang; Qin He (26-38).
Display OmittedThough PEGylation has been widely used to enhance the accumulation of liposomes in tumor tissues through enhanced permeability and retention (EPR) effects, it still inhibits cellular uptake and affects intracellular trafficking of carriers. Active targeting molecules displayed better cell selectivity but were shadowed by the poor tumor penetration effect. Cell penetrating peptides could increase the uptake of the carriers but were limited by their non-specificity. Dual-ligand system may possess a synergistic effect and create a more ideal drug delivery effect. Based on the above factors, we designed a multistage liposome system co-modified with RGD, TAT and cleavable PEG, which combined the advantages of PEG, specific ligand and penetrating peptide. The cleavable PEG could increase the stability and circulation time of liposomes during circulation. After the passive extravasation to tumor tissues, the previously hidden dual ligands on the liposomes were exposed in a controlled manner at the tumor site through exogenous administration of a safe reducing agent l-cysteine. The RGD specifically recognized the integrins overexpressed on various malignant tumors and mediated efficient internalization in the synergistic effect of the RGD and TAT. In vitro cellular uptake and 3D tumor spheroids penetration studies demonstrated that the system could not only be selectively and efficiently taken up by cells overexpress ingintegrins but also penetrate the tumor cells to reach the depths of the avascular tumor spheroids. In vivo imaging and fluorescent images of tumor section further demonstrated that this system achieved profoundly improved distribution within tumor tissues, and the RGD and TAT ligands on C-R/T liposomes produced a strong synergistic effect that promoted the uptake of liposomes into cells after the systemic administration of l-cysteine. The results of this study demonstrated a tremendous potential of this multistage liposomes for efficient delivery to tumor tissue and selective internalization into tumor cells.
Keywords: Tumor targeting; Multistage liposome; Cleavable PEG; Synergistic effect;

Display OmittedUsing a procedure of emulsification–lyophilization (PEL), adjuvant lipid A-cochleates (LACs) were prepared as a carrier for model antigen bovine serum albumin (BSA). With phosphatidylserine and lipid A as emulsifiers dissolved in oil phase (O), sucrose and CaCl2 in the inner water phase (W1), and BSA, sucrose and PEG2000 in the outer water phase (W2), the W1/O/W2 emulsions were prepared and subsequently lyophilized to form a dry product which was stable enough to be stored at room temperature. Upon rehydration of the dry products, cochleates formed with a size of 800 nm and antigen association rates of 38%. After vaccination of mice through oral mucosal (o.m.) administration, LACs showed no side effects but induced potent immune responses as evidenced by high levels of IgG in the sera and IgA in the salivary, intestinal and vaginal secretions of mice. In addition, high levels of IgG2a and IFN-γ in the sera or culture supernatants of splenocytes of the immunized mice were also detected. These results revealed that LACs induced a mixed Th1/Th2 response against the loaded antigens. Thus, the LACs prepared by PEL were able to induce both systemic and mucosal immune responses and may act as a potent cold-chain-free oral mucosal vaccine adjuvant delivery system (VADS).
Keywords: Cochleate; Lyophilization; Immune response; Mucosa associated lymphoid tissue; Pathogen-associated molecular pattern; Vaccine adjuvant delivery system;

A comparative ex vivo drug permeation study of beta-blockers through porcine buccal mucosa by Sonia Amores; Jacinto Lauroba; Ana Calpena; Helena Colom; Alvaro Gimeno; José Domenech (50-54).
Display OmittedApparent permeability coefficients (k p) of a series of beta-blockers: acebutolol, atenolol, labetalol, metoprolol, oxprenolol and propranolol, through porcine buccal mucosa were determined. The aim of the study was to determine the permeation parameters (apparent permeability coefficient, k p; flux, J; and lag time, TL ) as a measure of the intrinsic permeability of porcine buccal mucosa to these drugs, in order to predict the efficacy of their possible administration through human buccal mucosa. A positive linear correlation was observed between the apparent permeability coefficient, k pand the partition coefficient, P. Oxprenolol and propranolol are the drugs that presented the highest values of k p: 0.3231 × 102  cm/h and 0.5666 × 102  cm/h, respectively. Multiple linear regression (MLR) using least square estimation was performed on the data set with log k p as dependent variable and the descriptors as predictor variables.The potential systemic capacity after a buccal administration was predicted by estimating the plasma concentrations at steady-stated (C ss). Considering the entire process of permeation ex vivo, propranolol and oxprenolol would seem to be the best candidates for administration through the buccal mucosa.
Keywords: Buccal permeation; Beta-blockers; Porcine buccal mucosa; Ex vivo drug permeation;

Preliminary pharmaceutical development of antimalarial–antibiotic cotherapy as a pre-referral paediatric treatment of fever in malaria endemic areas by Alexandra Gaubert; Tina Kauss; Mathieu Marchivie; Boubakar B. Ba; Martine Lembege; Fawaz Fawaz; Jean-Michel Boiron; Xavier Lafarge; Niklas Lindegardh; Jean-Louis Fabre; Nicholas J. White; Piero L. Olliaro; Pascal Millet; Karen Gaudin (55-63).
Display OmittedArtemether (AM) plus azithromycin (AZ) rectal co-formulations were studied to provide pre-referral treatment for children with severe febrile illnesses in malaria-endemic areas. The target profile required that such product should be cheap, easy to administer by non-medically qualified persons, rapidly effective against both malaria and bacterial infections. Analytical and pharmacotechnical development, followed by in vitro and in vivo evaluation, were conducted for various AMAZ coformulations. Of the formulations tested, stability was highest for dry solid forms and bioavailability for hard gelatin capsules; AM release from AMAZ rectodispersible tablet was suboptimal due to a modification of its micro-crystalline structure.
Keywords: Artemether; Azithromycin; Malaria; Acute respiratory infections; Pediatric; Rectal route;

Display OmittedIn current pharmaceutical drug discovery, most candidates are poorly soluble in water, which can result in poor bioavailability. To overcome this problem, formulations that create supersaturation of the drug are a well-studied alternative. Characterizing the dissolution from these systems is challenging because conventional methods, such as sampling with a syringe then filtering with a 0.2–0.45 μm filter before an HPLC assay, can overestimate the concentration of dissolved drug by allowing nuclei or small precipitated particles to pass, which then dissolve in the HPLC mobile phase. Nuclei and small particles can also cause overestimation of the dissolved concentration when using optical methods. Such overestimations can lead to failure of in vivo prediction of drug bioavailability from supersaturated systems. This paper reports a novel method to determine the free dissolved drug concentration in a dissolution medium using pulsatile microdialysis (PMD). Ibuprofen was used as a model drug for determining precipitation and supersaturation. Supersaturation was induced chemically by changing pH, and also by dissolution/release from an in-house formulation. The adaptation of a previously developed PMD model is summarized, and experimental results comparing dissolved concentrations determined using PMD and direct sampling by syringe and filtering are presented.
Keywords: Supersaturation; Precipitation; Poorly soluble; Pulsatile microdialysis; PMD; Dissolution;

Insight into the fabrication of polymeric particle based oxygen carriers by Bin Li; Shasha He; Yanxin Qi; Zhigang Xie; Xuesi Chen; Xiabin Jing; Yubin Huang (75-82).
Display OmittedFor the sake of protein stability and targeted application as blood substitutes, formulation customization of hemoglobin-loaded polymeric particles (HbP) was conducted via a double emulsion method. Screening of the emulsification parameters was firstly performed for the stability of Hb, and the structure and functions of recovered Hb could be well preserved via CD and UV–vis spectroscopy investigation. In the optimized conditions, Hb was loaded into the polymeric matrix formed of three material compositions. They were poly(ϵ-caprolactone)(PCL), poly(ethylene glycol)-block-poly(allyl glycidyl ether) (functionalized with mercaptopropionic acid)-block-poly(ϵ-caprolactone) (PEG-PAGE(MPA)-PCL), and the blend of the two polymers. The morphology, internal structure, in vitro leakage and hemocompatibility of the HbP products were characterized in detail, and the encapsulation mechanism was explored by the combined analysis of the encapsulation efficiency, non-specific protein adsorption and in vitro leakage studies. Results showed that the burst release effect found in homopolymers could be alleviated by use of block copolymers due to the reduced protein adsorption, and completely avoided by further cross-linking of particles through carbonyl-amino condensation reactions. The amphiphilic copolymers showed relatively high stability in blood and no interference with blood components compared with hydrophobic PCL. These results suggest that both the optimization of emulsion formation and material composition are prerequisite for stable formulations of Hb encapsulated in polymeric particles.
Keywords: Hemoglobin; Emulsification; Polymeric particle; Burst release; Cross-linking; Blood compatibility;

New amphiphilic N-phosphoryl oligopeptides designed for gene delivery by Yunfei Sun; Long Chen; Fude Sun; Xibo Tian; Shi-Zhong Luo (83-90).
Display OmittedGene therapy is a potent tool for the treatment of cancer and other gene defect diseases, which involves using DNA that encodes a functional, therapeutic gene to replace a mutated gene. However, the DNA transfection efficiency is restricted by its negative charges and low susceptibility to endonucleases which prevent them penetrating tissue and cellular membranes. Both viral and non-viral vectors have been used for gene delivery, but the former are limited by their immunogenicity, while the latter are less efficient than their viral counterpart. Cationic amphiphilic lipopeptides whose structures can be easily modified and transformed have been used as non-viral vectors in gene delivery system due to their low cytotoxicity and high transfection efficiency. In this study, a series of cationic amphiphilic N-phosphoryl oligopeptides with varied lengths of hydrophobic tails and oligopeptide headgroups (C12-K6, C14-K6, C16-K6, Chol-K6 and C12-H6) were synthesized and used as gene delivery vectors. The affinities, abilities to condense pDNA and transfection efficiencies of the K6-lipopeptides were better than those of the H6-lipopeptides. In addition, the hydrophobic chains of the lipopeptides also affected their transfection efficiencies. The K6-lipopeptide with a hydrophobic chain of twelve carbons (C12-K6) showed the highest transfection efficiency in all these synthetic lipopeptides. At an optimal P/N ratio of 20, C12-K6 showed comparable pDNA transfection efficiency to PEI-25k, a well-defined gene delivery vector, but the cytotoxicity of C12-K6 was much lower. With acceptable gene transfection efficiency and low cytotoxicity, this cationic amphiphilic lipopeptide will have promising applications in gene therapy.
Keywords: Gene delivery; Non-viral vector; Cationic lipopeptide; Transfection;

Display OmittedThe pulverization of poorly water-soluble drugs and drug candidates into nanoscale particles is a simple and effective means of increasing their pharmacological effect. Consequently, efficient methods for pulverizing compounds are being developed. Femtosecond lasers, which emit ultrashort laser pulses, can be used to generate nanoscale particles without heating and are finding in various fields, including pharmaceutical science. Laser ablation holds promise as a novel top-down pulverization method for obtaining drug nanoparticles. We used a poorly water-soluble compound, curcumin (diferuloyl methane), to understand the characteristics of femtosecond laser pulverization. Various factors such as laser strength, laser scan speed, and the buffer solution affected the size of the curcumin particles. The minimum curcumin particle size was approximately 500 nm; the particle size was stable after 30 days. In vitro studies suggested that curcumin nanoparticles exhibited a cytotoxic effect on C6 rat glioma cells, and remarkable intracellular uptake of the curcumin nanoparticles was observed. The results suggest that femtosecond laser ablation is a useful approach for preparing curcumin nanoparticles that exhibit remarkable therapeutic effects.
Keywords: Femtosecond laser; Laser ablation; Pulverization; Curcumin; Glioma;

Display OmittedRecently, great attention has been paid to nanocapsules. The interest of these structures is due to their promising applications as drug delivery systems. The objective of this study was to develop novel enteric coating technique based on instantaneous encapsulation of the acid-labile drug, omeprazole in innovative enteric nanocapsules. Omeprazole enteric nanocapsules were formulated by varying the type and amount of the enteric polymer. The particle size (PS), polydispersity index (PDI), zeta potential (ZP) and encapsulation efficiency (EE) values of the prepared enteric nanocapsules were determined. A full 21  × 31 factorial design was used for planning and analysis of the experimental trials to select the optimized formulation. The highest desirability value was 0.7463 for formula E3 (containing 200 mg hydroxypropyl methylcellulose phthalate (HPMCP)). The stability of omeprazole was reflected by the absence of the exothermal peak when the drug was encapsulated as detected by differential scanning calorimetry (DSC) thermograms. In vitro drug release study confirmed the USP specifications required to meet the key formulation characteristics of gastro-resistance. In vivo pharmacological assessment showed that the optimized nanocapsules were able to protect rat stomach against ulcer formation compared to the aqueous suspension of the drug which showed less significant protection.
Keywords: Enteric nanocapsules; Omeprazole; Hydroxypropyl methylcellulose phthalate; In vivo antiulcer activity;

Dextran–protamine coated nanostructured lipid carriers as mucus-penetrating nanoparticles for lipophilic drugs by Ana Beloqui; María Ángeles Solinís; Anne des Rieux; Véronique Préat; Alicia Rodríguez-Gascón (105-111).
Display OmittedThe main objectives of the present study were (i) to evaluate the effect of the mucus layer on saquinavir-loaded nanostructured lipid carriers (SQV-NLCs) uptake and (ii) to evaluate the mucopenetrating properties of dextran–protamine (Dex–Prot) coating on NLCs as per SQV permeability enhancement. Three different NLC formulations differing on particle size and surfactant content were obtained and coated with Dex–Prot complexes. SQV permeability was then evaluated across Caco-2 cell monolayers (enterocyte-like model) and Caco-2/HT29-MTX cell monolayers (mucus model). In the Caco-2 monolayers, Dex–Prot–NLCs increased up to 9-fold SQV permeability in comparison to uncoated nanoparticles. In the Caco-2/HT29-MTX monolayers, Dex–Prot–NLCs presenting a surface charge close to neutrality significantly increased SQV permeability. Hence, Dex–Prot complex coating is a promising strategy to ensure successful nanoparticle mucus-penetration, and thus, an efficient nanoparticle oral delivery. To our knowledge, this is the first time that Dex–Prot coating has been described as a nanoparticle muco-penetration enhancer across the intestinal mucus barrier.
Keywords: NLCs; Mucus; HT29-MTX; Caco-2; Saquinavir; Permeability;

Evaluating optimal combination of clodronate and bioactive glass for dental application by Kirsi Rosenqvist; Sari Airaksinen; Marko Vehkamäki; Anne Mari Juppo (112-120).
Display OmittedBoth clodronate and bioactive glass are mostly used alone as treatment in various bone diseases but, they are also known to have beneficial effects in dental application. The same processes that lead to loss of bone can also result in alveolar bone loss. The object of this study was to define the optimal combination of clodronate and bioactive glass (BAG) to be used locally in dentistry. The evaluation was based on measurements and solid state properties obtained with pH, scanning electron microscopy (SEM), differential scanning calorimetric (DSC), X-ray powder diffraction (XRPD), Fourier transform infrared spectroscopy (FTIR) and Focused-ion beam (FIB) and energy dispersive X-ray spectroscopic (EDS) mapping. The results indicate that if too much calcium clodronate precipitation is formed, the activity of BAG is affected negatively. As there is more reaction surface to form calcium clodronate, similar to the amount of clodronate present, this reduces the bioactivity of BAG. Therefore, in dental treatment the most suitable BAG and clodronate combination product would have apatite (HA, hydroxyapatite) formation ability and amount of clodronate enough to enhance the bioactivity of BAG allowing HA formation. Based on combinations investigated, the one with 200 mg clodronate and 1 g BAG with particle size 0.5–0.8 mm was chosen to be the most promising for local dental application.
Keywords: Bioactive glass; Clodronate; Apatite formation; Granule size; Calcium clodronate;

Quantifying the release of lactose from polymer matrix tablets with an amperometric biosensor utilizing cellobiose dehydrogenase by Patrik Knöös; Christopher Schulz; Lennart Piculell; Roland Ludwig; Lo Gorton; Marie Wahlgren (121-132).
Display OmittedThe release of lactose (hydrophilic) from polymer tablets made with hydrophobically modified poly(acrylic acid) (HMPAA) have been studied and compared to the release of ibuprofen, a hydrophobic active substance. Lactose is one of the most used excipients for tablets, but lactose release has not been widely studied. One reason could be a lack of good analytical tools. A novel biosensor with cellobiose dehydrogenase (CDH) was used to detect the lactose release, which has a polydiallyldimethylammonium chloride (PDADMAC) layer that increases the response. A sample treatment using polyethylenimine (PEI) was developed to eliminate possible denaturants. The developed methodology provided a good approach to detect and quantify the released lactose. The release was studied with or without the presence of a model amphiphilic substance, sodium dodecyl sulphate (SDS), in the release medium. Ibuprofen showed very different release rates in the different media, which was attributed to hydrophobic interactions between the drug, the HMPAA and the SDS in the release medium. The release of hydrophilic lactose, which did not associate to any of the other components, was rapid and showed only minor differences. The new methodology provides a useful tool to further evaluate tablet formulations by a relatively simple set of experiments.
Keywords: Controlled release; Polymer matrix; Biosensors; Cellobiose dehydrogenease; Lactose release; Release mechanisms;

RGD-fatty alcohol-modified docetaxel liposomes improve tumor selectivity in vivo by Yinghuan Li; Xuelian Zheng; Yi Sun; Zhao Ren; Xuemei Li; Guohui Cui (133-141).
Display OmittedThe tripeptide arginine-glycine-aspartate (RGD) was conjugated with various fatty alcohols to obtain RGDOC n H2n  + 1 (n  = 8, 10, 12, 14, 16, 18), which were incorporated into the bilayer of docetaxel liposomes to improve their tumor specificity. The fatty alcohols were accepted as linking groups to insert the tetrapeptide RGDX (X = amino acid) into the bilayer of liposomes. RGDX was previously shown to be a potent ligand to target tumor cell-surface integrin receptors, whereas RGD was not shown to have this ability. We hypothesized that RGD-fatty alcohol conjugates lacking the fourth amine X can guide liposomes to tumors without reducing their binding affinity to integrins. Antitumor activity, pharmacokinetics and biodistribution studies were evaluated in mice inoculated with S180 sarcoma. Compared with unmodified liposomes, RGD-fatty alcohol-modified liposomes successfully delivered significantly more docetaxel to tumors, which led to significant tumor weight loss and increased tumor docetaxel concentrations accompanied by reduced liver accumulation. Improved affinity of RGD-fatty alcohols to integrins was also confirmed on A375 cell model. Further comparisons among the tumor-targeting capacities of liposomes containing RGD-fatty alcohols, RGDF-fatty alcohols and RGDV-fatty acids demonstrated that RGD-fatty alcohols were as effective as the other two tetrapeptide derivatives. Therefore, a simplified tumor-targeting delivery system using RGD-fatty alcohols was developed.
Keywords: RGD; Fatty alcohol; Docetaxel liposomes; Tumor targeting; Pharmacokinetics; In vivo;

Display OmittedAlthough gemcitabine (Gem) constitutes first-line therapy for pancreatic cancer, its clinical outcome suffers from rapid metabolism and acquired drug resistance. To overcome its limitations, several lipophilic prodrugs including 4-(N)-stearoyl Gem (GemC18) have been studied for their efficacy over Gem. Herein, we aimed to prepare and characterize the GemC18-loaded poly(ethylene glycol)–poly(d,l-lactide) (PEG–PLA) polymeric micelles (PMs) as well as its self-assembled nanoparticles (NPs). A D-optimal design was also utilized to investigate the effects of formulation variables, namely initial drug/polymer ratio, total solid content, and the type of organic solvent on properties of GemC18-loaded PMs. The optimized formulation showed a particle size of about 120 nm, encapsulation efficiency >90%, and a sustained release behavior of the drug. Alternatively, the prodrug NPs were harvested in larger size (∼300 nm) and more negative zeta potential, but less chemical stability compared to the optimized PMs. In Panc-1 and AsPC-1 cell lines, both GemC18-loaded PMs and NPs were significantly more cytotoxic than GemC18 solution. Chiefly, they could effectively reduce the viability of Gem high-resistant AsPC-1 cells in culture, whereas the molar equivalent doses of Gem did not show any acceptable cytotoxicity. Thus, these results suggest a promising direction for alternative Gem delivery systems for future therapeutic applications.
Keywords: Polymeric micelles; Stearoyl gemcitabine; Poly(ethylene glycol)–poly(d,l-lactide); Self-assembly; Pancreatic cancer;

Extended duration local anesthetic agent in a rat paw model by D.E. Ickowicz; L. Golovanevski; A.J. Domb; C.F. Weiniger (152-157).
Display OmittedEncapsulated local anesthetics extend postoperative analgesic effect following site-directed nerve injection; potentially reducing postoperative complications. Our study aim was to investigate efficacy of our improved extended duration formulation – 15% bupivacaine in poly(DL-lactic acid co castor oil) 3:7 synthesized by ring opening polymerization. In vitro, around 70% of bupivacaine was released from the p(DLLA-CO) 3:7 after 10 days. A single injection of the optimal formulation of 15% bupivacaine-polymer or plain (0.5%) bupivacaine (control), was injected via a 22G needle beside the sciatic nerve of Sprague-Dawley rats under anesthesia; followed (in some animals) by a 1 cm longitudinal incision through the skin and fascia of the paw area. Behavioral tests for sensory and motor block assessment were done using Hargreave’s hot plate score, von Frey filaments and rearing count. The 15% bupivacaine formulation significantly prolonged sensory block duration up to at least 48 h. Following surgery, motor block was observed for 48 h following administration of bupivacaine-polymer formulation and rearing was reduced (returning to baseline after 48 h). No significant differences in mechanical nociceptive response were observed. The optimized bupivacaine-polymer formulation prolonged duration of local anesthesia effect in our animal model up to at least 48 h.
Keywords: Bupivacaine; Injectable local anesthetics; Polymer; Prolonged release;

Synthesis and in vitro localization study of curcumin-loaded SPIONs in a micro capillary for simulating a targeted drug delivery system by Mohammed Anwar; Mohammed Asfer; Ayodhya P. Prajapati; Sharmistha Mohapatra; Sohail Akhter; Asgar Ali; Farhan J. Ahmad (158-164).
Display OmittedNano-sized curcumin-loaded super-paramagnetic iron oxide nanoparticles (CUR-OA-SPIONs) were synthesized chemically by co-precipitation method using oleic acid as a stabilizer and Myrj 52 as a surfactant. The synthesized nanoparticles were characterized for their shape, size, surface morphology, electrokinetic potential, magnetic properties, crystalinity, chemical interactions and thermal transitions. The synthesized CUR-OA-SPIONs were spherical, mono-dispersed, physically stable and super-paramagnetic in nature. In vitro localization study and aggregation dynamics of CUR-OA-SPIONs were studied with a flow of blood inside a square glass capillary (500 × 500 μm2 cross section) in the presence of an externally applied magnetic field (Ms = 1200 mT). This research which is first of its kind showed the fluorescent imaging of CUR-OA-SPIONs with respect to time to understand the aggregation dynamics of magnetic nanoparticles in a micro capillary simulating the case of targeted drug delivery system. The size of the aggregation increases with respect to time (t  = 0+  s to t  = 500 s), while no significant change in the size of the aggregate was observed after time t  = 500 s.
Keywords: Curcumin; Superparamagnetic iron oxide nanoparticles; SPIONs; Drug localization; Drug targeting; Imaging;

Preparation and evaluation of colon adhesive pellets of 5-aminosalicylic acid by Meixia Xu; Minjie Sun; Hongzhi Qiao; Qineng Ping; Eltayeb Suliman Elamin (165-171).
Display OmittedOral modified-release delivery systems, such as bio-adhesive one, enable drug delivery to affected regions and minimize the side effects by reducing the systemic absorption. Our aim was to develop colon adhesive pellets of 5-aminosalicylic acid (5-ASA) for the treatment of ulcerative colitis. The core of the pellet was formulated from bioadhesive agents, Carbomer 940 and hydroxypropyl cellulose (HPC), by extrusion/spheronization method and coated with Surelease® as inner layer for waterproof and with Eudragit® S100 as outer layer for pH control. The rat model of ulcerative colitis was used to evaluate the efficiency of our loaded pellets as a drug carrier. Microcrystalline cellulose 101 (PH 301) was found to be the best agent for pellet core. The ratio of CP940 to HPC should be kept as (1:1) to achieve high bioadhesion. When the amount of Surelease® was from 16% to 20% and of Eudragit® S100 was 28%, the dissolution profiles of coated pellets revealed no drug release in the artificial gastric fluid (pH 1.0) within 2 h and less than 10% was released in phosphate buffer (pH 6.0) within 2 h whereas complete dissolution was observed in colonic fluid of pH 7.4 for 20 h. The animal experiment showed that 5-ASA loaded colon adhesive pellets had optimal therapeutic effect. We showed a novel approach to prepare effective bioadhesive pellets as colon targeted drug delivery system.
Keywords: 5-Aminosalicylic acid; Bioadhesion; Extrusion spheronization; Colon-specific delivery; Pharmacodynamics;

Accelerating the dissolution of enteric coatings in the upper small intestine: Evolution of a novel pH 5.6 bicarbonate buffer system to assess drug release by Felipe J.O. Varum; Hamid A. Merchant; Alvaro Goyanes; Pardis Assi; Veronika Zboranová; Abdul W. Basit (172-177).
Display OmittedDespite rapid dissolution in compendial phosphate buffers, gastro resistant (enteric coated) products can take up to 2 h to disintegrate in the human small intestine, which clearly highlights the inadequacy of the in vitro test method to predict in vivo behaviour of these formulations. The aim of this study was to establish the utility of a novel pH 5.6 bicarbonate buffer, stabilized by an Auto pH™ System, as a better surrogate of the conditions of the proximal small intestine to investigate the dissolution behaviour of standard and accelerated release enteric double coating formulations. Prednisolone tablets were coated with 3 or 5 mg/cm2 of partially neutralized EUDRAGIT® L 30 D-55, HP-55 or HPMC adjusted to pH 6 or 8. An outer layer of EUDRAGIT® L 30 D-55 was applied at 5 mg/cm2. For comparison purposes, a standard single layer of EUDRAGIT® L 30 D-55 was applied to the tablets. Dissolution was carried out using USP II apparatus in 0.1 M HCl for 2 h, followed by pH 5.6 bicarbonate buffer. EUDRAGIT® L 30 D-55 single-coated tablets showed a slow drug release with a lag time of 75 min in buffer, whereas release from the EUDRAGIT® L 30 D-55 double-coated tablets was accelerated. These in vitro lag times closely match the in vivo disintegration times for these coated tablets reported previously. Drug release was further accelerated from modified double coatings, particularly in the case of coatings with a thinner inner layer of HP-55 or HPMC (pH 8 and KH2PO4). This study confirms that the pH 5.6 bicarbonate buffer system offers significant advantages during the development of dosage forms designed to release the drug in the upper small intestine.
Keywords: Gastro resistant coatings; Bicarbonate buffers; Biorelevant dissolution; pH responsive polymers; Hypromellose phthalate; Enteric polymers;

Display Omitted In situ coating of 5-fluorouracil pellets by ethylcellulose and pectin powder mixture (8:3 weight ratio) in capsule at simulated gastrointestinal media provides colon-specific drug release in vitro. This study probes into pharmacodynamic and pharmacokinetic profiles of intra-capsular pellets coated in vivo in rats with reference to their site-specific drug release outcomes. The pellets were prepared by extrusion–spheronization technique. In vitro drug content, drug release, in vivo pharmacokinetics, local colonic drug content, tumor, aberrant crypt foci, systemic hematology and clinical chemistry profiles of coated and uncoated pellets were examined against unprocessed drug. In vivo pellet coating led to reduced drug bioavailability and enhanced drug accumulation at colon (179.13 μg 5-FU/g rat colon content vs 4.66 μg/g of conventional in vitro film-coated pellets at 15 mg/kg dose). The in vivo coated pellets reduced tumor number and size, through reforming tubular epithelium with basement membrane and restricting expression of cancer from adenoma to adenocarcinoma. Unlike uncoated pellets and unprocessed drug, the coated pellets eliminated aberrant crypt foci which represented a putative preneoplastic lesion in colon cancer. They did not inflict additional systemic toxicity. In vivo pellet coating to orally target 5-fluorouracil delivery at cancerous colon is a feasible therapeutic treatment approach.
Keywords: Colon cancer; Colon-specific drug delivery; 5-Fluorouracil; In vivo pellet coating;

Display OmittedOil-in-water emulsions consisting of squalene, tween and/or span have shown significant benefits for the prevention and control of influenza, with their adjuvant efficacy enhancing the immunogenicities of influenza vaccines in high-risk groups. However, concerns have been raised following reports that post-immunization reactions associated with these adjuvanted vaccines are more frequent. In this work, a stable and biocompatible oil-in-water emulsion adjuvant containing squalene, egg lecithin and sodium oleate has been developed. Animal studies demonstrated that this adjuvant could induce strong immune responses in BALB/c mice, as measured by hemagglutinin inhibition titers, influenza-specific serum antibody titers and cytokine levels (IFN-γ and IL-4). Different oil compositions, including squalene, medium chain triglyceride and long chain triglyceride, were also evaluated. Furthermore, in contrast to MF59® which can only be sterilized by aseptic filtration, this adjuvant remained stable during autoclaving, showing minimal changes in pH, particle size and lysolecithin concentration.
Keywords: Adjuvants; Oil-in-water emulsion; Influenza vaccine; Egg lecithin; Squalene;

Taste acceptability of pulverized brand-name and generic drugs containing amlodipine or candesartan by Peter Uestuener; Alessandra Ferrarini; Maristella Santi; Chiara Mardegan; Mario G. Bianchetti; Giacomo D. Simonetti; Gregorio P. Milani; Sebastiano A.G. Lava (196-198).
Display OmittedTrials with pulverized brand-name antihypertensive drugs suggest that, from the perspective of taste acceptability, crushed candesartan, chlortalidon, hydrochlorothiazide, lercanidipine and lisinopril should be preferred to pulverized amlodipine, atenolol, bisoprolol, enalapril, irbesartan, losartan, ramipril, telmisartan and valsartan. Brand-name antihypertensive drugs and the corresponding generic medicines have never been compared with respect to their taste acceptability. We therefore investigated among healthy health care workers the taste acceptability of a pulverized 1 mg-test dose of the brand-name and two generics containing either the dihydropyridine calcium-channel blocker amlodipine (Norvasc®, Amlodipin-Mepha® and Amlodipin Pfizer®) or the angiotensin receptor antagonist candesartan (Atacand®, Cansartan-Mepha® and Pemzek®). For this purpose, a smiley-face scale depicting four degrees of pleasure was used. Between November and December 2013, the taste test was performed among 19 nurses (15 female and 4 male subjects) and 12 physicians (5 female and 7 male subjects) aged between 25 and 49 years. Pulverized brand-names and generics containing either amlodipine or candesartan did not differ with respect to their taste acceptability.
Keywords: Angiotensin receptor antagonists; Calcium-channel blockers; Hypertension; Medication adherence; Taste perception;

The effect of shear and extensional viscosity on atomization in medical inhaler by L. Broniarz-Press; M. Ochowiak; M. Matuszak; S. Włodarczak (199-206).
Display OmittedThe paper contains the results of experimental studies of water, aqueous solutions of glycerol and aqueous solutions of glycerol-polyethylene oxide (PEO) atomization process in a medical inhaler obtained by the use of the digital microphotography method. The effect of the shear and extensional viscosity on the drop size, drop size histogram and mean drop diameter has been analyzed. The obtained results have shown that the drop size increases with the increase in shear and extensional viscosity of liquid atomized. Extensional viscosity has a greater impact on the spraying process. It has been shown that the change in liquid viscosity leads to significant changes in drop size distribution. The correlation for Sauter mean diameter as function of the shear and extensional viscosity was proposed.
Keywords: Shear viscosity; Extensional viscosity; Atomization; Inhaler; Drop size histogram; Sauter mean diameter;

Preparation and evaluation of ritodrine buccal tablets for rational therapeutic use by Hiraku Onishi; Kei Yumoto; Osamu Sakata (207-213).
Display OmittedRitodrine hydrochloride (RD-HCl) tablets containing alginate (AL) and lactose (LC) with or without microcrystalline cellulose (MC) as excipients were produced as a buccal dosage form. The RD-HCl (2 mg) tablets with AL/LC but no MC swelled and dissolved gradually in the in vitro dissolution test. The tablet showing the fastest dissolution and highest drug release rate, called Tablet A1, was selected as a tablet to show rapid and prolonged absorption. However, in the in vivo buccal absorption test using rats, it could not give a plasma concentration over the human minimal effective level (15 ng/mL). The modified tablet containing AL, LC, MC and RD-HCl (4 mg), named Tablet B/MC, showed better hardness and faster drug release. Tablet B/MC gave a plasma concentration over the human effective level within 15 min, and the plasma concentration was maintained at >15 ng/mL over 4 h. Moreover, the deconvolution analyses demonstrated that a prolonged high absorption rate could be achieved in vivo best with Tablet B/MC. Tablet B/MC improved the pharmacokinetic profile in comparison with Tablet A1 and the solution dosage form. The RD-HCl buccal tablets with AL, LC and MC as excipients are suggested to be possibly useful for the treatment of premature labor.
Keywords: Ritodrine hydrochloride; Buccal tablet; Plasma level; Absorption rate; Dissolution; Hardness;

Studies on drug-polymer interaction, in vitro release and cytotoxicity from chitosan particles excipient by Thandapani Gomathi; C. Govindarajan; Maximas H. Rose H.R.; P.N. Sudha; P.K. Mohamed Imran; Jayachandran Venkatesan; Se-Kwon Kim (214-222).
Display OmittedNonobvious controlled polymeric pharmaceutical excipient, chitosan nanoparticles (CS-NPs) for lenalidomide encapsulation were geared up by the simple ionic cross linking method to get better bioavailability and to reduce under as well as overloading of hydrophobic and sparingly soluble drug lenalidomide towards cancer cells. Lenalidomide loaded chitosan nanoparticles (LND-CS-NPs) were in the size range of 220–295 nm and characterized by DLS, TEM, FT-IR, TGA and XRD. Encapsulation of lenalidomide over chitosan nanoparticles was observed about 99.35% using UV spectrophotometry method. In vitro release and the cytotoxic studies were performed using LND-CS-NPs. This study implies the new drug delivery route for lenalidomide and illustrates that the CS-NPs serves as the effective pharmaceutical carrier for sustained delivery of lenalidomide.
Keywords: Pharmaceutical carrier; Biodegradable polymer; New drug delivery route; Nanoparticles; Nanoencapsulation; Lenalidomide;

Display OmittedIn this work, one-dimensional population balance models (PBMs) have been developed to model a pulsed top-spray fluidized bed granulation. The developed PBMs have linked the key binder solution spray operating factors of the binder spray rate, atomizing air pressure and pulsed frequency of spray with the granule properties to predict granule growth behaviour in the pulsed spray fluidized bed granulation process at different operating conditions with accuracy. A multi-stage open optimal control strategy based on the developed PBMs was proposed to reduce the model mismatch, in which through adjusting the trajectory of the evolution of the granule size distribution at predefined sample intervals, to determine the optimal operating variables related to the binder spray including the spray rate of binding liquid, atomizing air pressure and pulsed frequency of spray. The effectiveness of the proposed modelling and multi-stage open optimal control strategies has been validated by experimental and simulation tests.
Keywords: Top-spray fluidized bed granulation; Population balance model; Optimal control; Particle size distribution;

Impact of active ingredients on the swelling properties of orally disintegrating tablets prepared by microwave treatment by Syusuke Sano; Yasunori Iwao; Susumu Kimura; Shuji Noguchi; Shigeru Itai (234-242).
Display OmittedThe impact of different active pharmaceutical ingredients (APIs) loading on the properties of orally disintegrating tablets (ODTs) prepared according to our previously reported microwave (MW) treatment process was evaluated using famotidine (FAM), acetaminophen (AAP), and ibuprofen (IBU). None of the APIs interrupted the tablet swelling during the MW treatment and the tablet hardness were improved by more than 20 N. MW treatment, however, led to a significant increase in the disintegration time of the ODTs containing IBU, but it had no impact on that of the ODTs containing FAM or AAP. This increased disintegration time of the ODTs containing IBU was attributed to the relatively low melting point of IBU (T m  = 76 °C), with the IBU particles melting during the MW treatment to form agglomerates, which interrupted the penetration of water into the tablets and delayed their disintegration. The effects of the MW treatment on the chemical stability and dissolution properties of ODTs were also evaluated. The results revealed that MW treatment did not promote the degradations of FAM and AAP or delay their release from the ODTs, while dissolution of the ODTs containing IBU delayed by MW treatment. Based on these results, the MW method would be applicable to the preparation of ODTs containing APIs with melting points higher than 110 °C.
Keywords: Orally disintegrating tablet; Microwave; Famotidine; Acetaminophen; Ibuprofen; Fluorescence;

Display OmittedLight- and temperature-responsive liposomes were prepared by immobilizing cinnamoyl Pluronic F127 (CP F127) on the surface of egg phosphatidylcholine liposomes. CP F127 was prepared by a condensation reaction, and the molar ratio of cinnamoyl group to Pluronic F127 was calculated to be 1:1.4 on 1H NMR spectrum. The cinnamoyl group of CP F127 was readily dimerized under the irradiation of a UV light (254 nm, 6 W). CP F127 decreased the absolute value of the zeta potential of liposome possibly because it can shift the hydrodynamic plane away from the liposome surface. The size of liposome decorated with CP F127, measured on a dynamic light scattering machine and observed on a TEM, was larger than that of bare liposome. The liposome bearing CP F127 seemed to fuse and aggregate each other. The liposome released calcein, a fluorescence dye, in response to a UV irradiation, possibly because the photo-dimerization of cinnamoyl group perturbs the liposomal membrane. Moreover, the liposome released the dye in response to a temperature change, possible due to the phase transition of Pluronic F127 layer on the liposomal surface or the hydrophobic interaction of the polymer with liposomal membrane.
Keywords: Liposomes; Cinnamoyl Pluronic F127; Light-responsive; Temperature-responsive; Release;

Evaluation of P(L)LA-PEG-P(L)LA as processing aid for biodegradable particles from gas saturated solutions (PGSS) process by D.R. Perinelli; G. Bonacucina; M. Cespi; A. Naylor; M. Whitaker; G.F. Palmieri; G. Giorgioni; L. Casettari (250-257).
Display OmittedA series of biodegradable P(L)LA-PEG1.5 kDa-P(L)LA copolymers have been synthesized and compared as processing aid versus Poloxamer 407 (PEO–PPO–PEO), in the formulation of protein encapsulated microparticles, using supercritical carbon dioxide (scCO2). Bovine serum albumin (BSA) loaded microcarriers were prepared applying the particles from the gas saturated solutions (PGSS) technique using scCO2 and thus, avoiding the standard practice of organic solvent encapsulation. Four triblock copolymers were synthesized and characterized, particularly in terms of thermal properties and behaviour when exposed to scCO2. The effects of the inclusion of these copolymers in the formulation of poly(α-hydroxy acids) based microparticles – e.g. poly(d,l-lactic-co-glycolic acid) (PLGA) and poly(d,l-lactide) (PLA) – were analysed in terms of yield, particle size, morphology and drug release. The use of P(L)LA-PEG1.5 kDa-P(L)LA triblock copolymers were found to increase the yield of the PGSS-based process and to decrease the size of the microparticles produced, in comparison with the formulation containing the Poloxamer 407. Moreover the microparticles formulated with the triblock copolymers possessing the higher hydrophobic character were able to maintain a controlled drug release profile.
Keywords: Block copolymers; PGSS; Biodegradable microparticles; scCO2; BSA;

β-Cyclodextrin-dextran polymers for the solubilization of poorly soluble drugs by Massimiliano di Cagno; Thorbjørn Terndrup Nielsen; Kim Lambertsen Larsen; Judith Kuntsche; Annette Bauer-Brandl (258-263).
Display OmittedThe aim of this study was to assess the potential of novel β-cyclodextrin (βCD)-dextran polymers for drug delivery. The size distribution of βCD-dextrans (for eventual parenteral administration), the influence of the dextran backbones on the stability of the βCD/drug complex, the solubilization efficiency of poorly soluble drugs and drug release properties were investigated. Size analysis of different βCD-dextrans was measured by size exclusion chromatography (SEC) and asymmetrical flow field-flow fractionation (AF4). Stability of drug/βCD-dextrans was assessed by isothermal titration calorimetry (ITC) and molar enthalpies of complexation and equilibrium constants compared to some commercially available βCD derivatives. For evaluation of the solubilization efficiency, phase-solubility diagrams were made employing hydrocortisone (HC) as a model of poorly soluble drugs, whereas reverse dialysis was used to detect potential drug supersaturation (increased molecularly dissolved drug concentration) as well as controlled release effects. Results indicate that all investigated βCD-polymers are of appropriate sizes for parenteral administration. Thermodynamic results demonstrate that the presence of the dextran backbone structure does not affect the stability of the βCD/drug complex, compared to native βCD and commercially available derivatives. Solubility studies evidence higher solubilizing abilities of these new polymers in comparison to commercially available βCDs, but no supersaturation states were induced. Moreover, drug release studies evidenced that diffusion of HC was influenced by the solubilization induced by the βCD-derivatives.
Keywords: Cyclodextrins; Calorimetry (ITC); βCD-dextran polymers; Solubilization; Poorly soluble drugs;

Construction and physiochemical characterisation of a multi-composite, potential oral vaccine delivery system (VDS) by Marie W. Pettit; Paul D.R. Dyer; John C. Mitchell; Peter C. Griffiths; Bruce Alexander; Beatrice Cattoz; Richard K. Heenan; Stephen M. King; Ralf Schweins; Frank Pullen; Stephen R. Wicks; Simon C.W. Richardson (264-271).
Display OmittedAn increasing human population requires a secure food supply and a cost effective, oral vaccine delivery system for livestock would help facilitate this end. Recombinant antigen adsorbed onto silica beads and coated with myristic acid, was released (∼15% (w/v)) over 24 h at pH 8.8. At pH 2, the myristic acid acted as an enteric coating, protecting the antigen from a variety of proteases. The antigen adsorbed onto silica particles, coated in myristic acid had a conserved secondary structure (measured by circular dichroism (CD) spectroscopy) following its pH-triggered release. Small angle neutron scattering (SANS) was used to measure the thickness of the adsorbed antigen, finding that its adsorbed conformation was slightly greater than its solution radius of gyration, i.e. 120–160 Å. The addition of myristic acid led to a further increase in particle size, with scattering data consistent with an acid thickness slightly greater than a monolayer of fully extended alkyl chains and a degree of hydration of around 50%. Whilst adsorbed onto the silica and coated in myristic acid, the protein was stable over 14 days at 42 °C, indicating a reduced need for cold chain storage. These data indicate that further investigation is warranted into the development of this technology.
Keywords: Vaccine; Enteric; Immobilisation; Silica; Synthetic protein;

Display OmittedThe present study aimed to develop an in situ gel formulation for intranasal delivery of tacrine (THA), an anti-Alzheimer’s drug. Thermosensitive polymer Pluronic F-127 was used to prepare THA in situ gels. Sol–gel transition temperature (T sol–gel), rheological properties, in vitro release, and in vivo nasal mucociliary transport time were optimized. The pharmacokinetics and brain dispositions of in situ gel were compared with that from THA oral solution in rats. The in situ gel demonstrated a liquid state with Newtonian fluid behavior under 20 °C, while it exhibited as non-flowing gel with pseudoplastic fluid behavior beyond its T sol–gel of 28.5 °C. Based on nasal mucociliary transport time, the in situ gel significantly prolonged its retention in nasal cavity compared to solution form. Moreover, the in situ gel achieved 2–3 fold higher peak plasma concentration (C max) and area under the curve (AUC) of THA in plasma and brain tissue, but lowered C max and AUC of the THA metabolites compared to that of oral solution. The enhanced nasal residence time, improved bioavailability, increased brain uptake of parent drug and decreased exposure of metabolites suggested that the in situ gel could be an effective intranasal formulation for THA.
Keywords: Tacrine; In situ gel; Intranasal; Pluronic F-127; Pharmacokinetics;