International Journal of Pharmaceutics (v.313, #1-2)

Stability of Beriplast® P fibrin sealant: Storage and reconstitution by Ulrich Eberhard; Martin Broder; Günther Witzke (1-4).
This study was performed to investigate the stability of Beriplast® P fibrin sealant (FS) across a range of storage conditions, both pre- and post-reconstitution. Storage stability of the FS was evaluated during long-term refrigeration (24 months) with or without interim storage at elevated temperatures (40 °C for 1 week and 25 °C for 1 and 3 months). Stability of individual FS components was assessed by measuring: fibrinogen content, Factor XIII activity (FXIII), thrombin activity and aprotinin potency. The package integrity of each component was also checked (sterility testing, moisture content and pH). Storage stability was also evaluated by testing the reconstituted product for adhesion (tearing force testing after mixing the solutions) and sterility. Reconstitution stability was evaluated following 3-months’ storage, for up to 50 h post-reconstitution using the same tests as for the storage stability investigations. Pre-defined specifications were met for fibrinogen content, Factor XIII activity, and thrombin activity, demonstrating storage stability. Package integrity and the functionality and sterility of the reconstituted product were confirmed throughout. Reconstitution stability was demonstrated for up to 50 h following reconstitution, in terms of both tearing force and sterility tests. In conclusion, the storage stability of Beriplast® P was demonstrated over a range of 24-month storage schedules including interim exposure to elevated temperature, and the reconstituted product was stable for up to 50 h.
Keywords: Fibrin tissue adhesive; Fibrin sealant; Drug storage; Drug stability;

Statistical evaluation of physiological variability of rifampicin in fixed dose combinations by Ramesh Panchagnula; Jitendra Parmar; Kanwaljit Kaur; Inderjit Singh; Shantaram Ramdos Bade; Yasvanth Ashokraj (5-13).
Tuberculosis is one of the microbial diseases having a long history of its occurrence and yet to be eradicated from the world. Due to the development of bacterial resistance, treatment has changed from monotherapy to combotherapy to fixed dose combinations (FDCs). Rifampicin has been found one of the most important anti-tubercular drugs, however variable bioavailability of rifampicin in some FDCs as well as separate formulations has been reported in the literature, and led to the development of WHO model protocol for evaluation of FDCs for bioequivalence trials. In present investigation, role of physiological variability in rifampicin bioequivalence was studied. Influence of subject's body weight, inter/intra-individual variability of elimination rate and impact of outliers on the decision of bioequivalence were investigated. Normalization of pharmacokinetic measures for bioequivalence (AUC and C max) were carried out as per body weights and elimination rate constants of subjects, then different statistical tests like two-way ANOVA, hauschke analysis, normal and log-transformed confidence interval were applied to check for the change in bioequivalence decision. It was found that normalization as per body weights did not play a significant role in the outcome of bioequivalence endpoint. Similarly, elimination rate variability and outliers have been found insignificant regarding final outcome of bioequivalence study. Hence, it has been concluded that physiological variability did not play a significant role in bioequivalence of rifampicin in FDCs.
Keywords: Rifampicin; Variable bioavailability; Physiological factors;

Co-deposition of salmeterol and fluticasone propionate by a combination inhaler by Andrew Theophilus; Alison Moore; Dave Prime; Simona Rossomanno; Brandon Whitcher; Henry Chrystyn (14-22).
The combination of the long-acting β2-agonist, salmeterol xinafoate (salmeterol) and inhaled corticosteroid, fluticasone propionate (FP) (Seretide®/Advair®) has shown enhanced efficacy compared with concurrent administration of the two drugs from individual inhalers at the same dose. A possible explanation for this increased effect is a higher degree of co-deposition of the two drugs from the combination (Seretide) inhaler compared with the component drugs administered separately.Raman laser spectroscopy, a technique capable of identifying individual drug particles, has been used with novel statistical methodology that we have developed, to determine whether there is any co-association between drug particles and whether this occurs in the Seretide formulation rather than by chance. Samples from a combined Seretide metered dose inhaler (MDI, 25/50 mcg) and salmeterol (25 mcg) with FP (50 mcg) from separate MDI's taken from Plate 4 of an Anderson Cascade Impactor were analysed.Using a statistical test based on the bootstrap technique, it was found that the co-deposition of FP and salmeterol particles from the combination MDI was significantly greater than from the separate inhalers group (p  < 0.001). A higher degree of co-deposition on the same cells of the airways may possibly account for the increased efficacy observed in patients prescribed Seretide MDI.
Keywords: Synergy; Salmeterol; Fluticasone propionate; Combination therapy;

Investigating the moisture-induced crystallization kinetics of spray-dried lactose by D.J. Burnett; F. Thielmann; T. Sokoloski; J. Brum (23-28).
Gravimetric water sorption experiments were performed to study the crystallization behavior of amorphous spray-dried lactose over a wide range of temperature and humidity conditions. Experiments performed at 25 °C between 48 and 60% relative humidity (RH) showed that the onset time to crystallization increased dramatically with decreasing humidity. At 55% RH and above, crystallization occurred in a single detectible step, while below a two-step process was observed. Experiments performed at 51% RH between 22 and 32 °C indicated the induction time to crystallization onset increased with decreasing temperature. Above 25 °C at 51% RH, crystallization occurred in one measurable step, while below crystallization occurred in two steps. The constant RH with varying temperature results were modeled to determine the crystallization mechanism. Above 25 °C a mechanism consisting of two competing reaction sequences fit the data with a 0.9997 correlation coefficient. Both reaction sequences have two steps: an auto-catalytic first step is followed by a three-dimensional diffusion controlled water loss step.
Keywords: Vapor sorption; Phase transition; Humidity; Amorphous; Crystallization kinetics; Spray-drying;

Lactococcus lactis subsp. lactis transformed with Plasmid ss80 (encoding the production and secretion of TEM β-lactamase) was used for the delivery of β-lactamase through the C-33A (cervix cell) monolayer. The viability of the cell monolayers co-cultured with L. lactis was examined by the trypan blue exclusion method. The integrity of the monolayers was monitored by measuring the transport of mannitol and propranolol as well as the transepithelial electrical resistance. The transport rate of β-lactamase through C-33A monolayer was increased by four- and nine-folds (p  < 0.05) at the first hour by the transformed L. lactis compared to the free solution with or without presence of the untransformed L. lactis, respectively. This increase was gradually diminished after the 1st hour: it became 30 and 50% (p  < 0.05) at 10 h. The presence of the untransformed L. lactis with free solution delivery also increased the transport rate by 100% at 1 h (p  < 0.05) and 15% at 10 h (p  > 0.05). The increase in transport rate by the transformed L. lactis is most probably due to the concentrate of β-lactamase on C-33A monolayer. When co-cultured with the L. lactis, the C-33A cell viability and the monolayer TEER remained steady for 10 h. The presence of L. lactis did not change the transport of propranolol and mannitol through the monolayers. In conclusion, the transformed L. lactis significantly (p  < 0.05) increased the transport of β-lactamase through the cervical monolayers, indicating probiotic bacteria delivery may be a promising approach for protein delivery through the vagina.
Keywords: C-33A; Lactococcus lactis; TEM β-lactamase; Protein delivery; Normal flora; Bioavailability; Vaginal delivery;

Synthesis and evaluation of the mucoadhesivity of a CD-chitosan derivative by J.P. Venter; A.F. Kotzé; R. Auzély-Velty; M. Rinaudo (36-42).
Combining mucoadhesive characteristics of a biodegradable polymer such as chitosan with the potential to enhance drug release by increasing the solubility of poorly water-soluble drugs has great potential for pharmaceutical technology and drug delivery design. Polymeric delivery systems have been extensively researched in an attempt to achieve modified drug release. Cyclodextrins (CD) offer an alternative approach. These cyclic oligosaccharides have the ability to form non-covalent complexes with a number of drugs altering their physicochemical properties. In the continuing challenge to improve the properties of delivery systems, this paper focuses on the modification of chitosan by introducing β-cyclodextrin and to test the mucoadhesive strength and inclusion properties of this synthesised cyclodextrin-polymer. β-Cyclodextrin was successfully grafted onto a chitosan chain polymer with a cyclodextrin grafting yield of 7% and a CD-chitosan yield of 85%. Although the complexation of (+)-catechin by the grafted β-CD was found to be about five times weaker than that by the β-CD monoaldehyde and natural β-CD, the inclusion properties of the chitosan-CD remain promising. The mucoadhesive properties of chitosan-CD were compared to that of pectin (reference) and the parent chitosan with the use of a tensile separation test. The chitosan-CD showed mucoadhesive strengths of 12% stronger than pectin, but 13.5% weaker than the parent chitosan. The synthesised chitosan-CD-polymer exhibits characteristics of a possible mucoadhesive drug delivery system with some inclusion properties from β-cyclodextrin.
Keywords: Chitosan; Drug delivery; Bioabsorption; Mucosal adhesion;

Cast films composed of mixtures of Eudragit® S100:L100 (1:1) and plasticized with triethyl citrate (TEC) were prepared from aqueous dispersions and organic solutions, and the physicochemical properties and the weight loss of cast films during dissolution testing were examined. The tensile strength of the organic cast films was significantly higher and the percent elongation was lower than that of the aqueous cast films. The weight loss of the organic films was also lower than that of the aqueous films. Furthermore, leaching of the TEC from the aqueous films was rapid and the TEC was found to diffuse from the films within one hour at pH 6.0, the pH at which the Eudragit® S100:L100 (1:1) films were insoluble. In contrast to the aqueous films, minimal levels of the TEC diffused from the organic cast films, and the disintegration of acrylic polymers occurred simultaneously with the release of TEC from the film during dissolution testing at pH 7.0. For Eudragit® L100-55, which could form films at lower TEC levels than Eudragit® S100:L100, both the organic and aqueous films showed the same weight loss after four hours in pH 5.0 media. These results demonstrated that for Eudragit® S100:L100 cast films, the high levels of TEC needed for film formation from aqueous dispersions resulted in rapid dissolution and disintegration at pH 6.0 and above. While aqueous dispersions are preferred for the coating of solid substrates, for Eudragit® S100:L100 film coatings, a change from organic solutions to aqueous dispersions in the coating process will impact film properties and product performance.
Keywords: Organic cast films; Aqueous cast films; Eudragit® S100; Eudragit® L100; Eudragit® L100-55; Triethyl citrate;

Modulation of intestinal P-glycoprotein function by polyethylene glycols and their derivatives by in vitro transport and in situ absorption studies by Qi Shen; Yulian Lin; Takahiro Handa; Masamichi Doi; Masami Sugie; Kana Wakayama; Naoki Okada; Takuya Fujita; Akira Yamamoto (49-56).
We examined the effect of polyethylene glycols (PEGs) with different molecular weights and their derivatives on the intestinal absorption of rhodamine123, a P-glycoprotein (P-gp) substrate, across the isolated rat intestinal membranes by an in vitro diffusion chamber system. The serosal to mucosal (secretory) transport of rhodamine123 was greater than its mucosal to serosal (absorptive) transport, indicating that the net movement of rhodamine123 across the intestinal membranes was preferentially secretory direction. The secretory transport of rhodamine123 was inhibited by the addition of PEGs with average molecular weights of 400, 2000 and 20,000, irrespective of its molecular weight. The inhibitory effects of these PEGs for the intestinal P-gp function were concentration dependent over the range 0.1–20% (v/v or w/v). Similar inhibitory effect for the intestinal P-gp function was observed when PEG derivatives including PEG monolaurate, PEG monooleate and PEG monostearate were added to the mucosal site of the chambers. Furthermore, we also examined effect of PEG20,000 on the intestinal absorption of rhodamine123 by an in situ closed loop method. The intestinal absorption of rhodamine123 was enhanced in the presence of PEG20,000. These findings suggest that PEGs and their derivatives are useful excipients to inhibit the function of intestinal P-gp, thereby improving the intestinal absorption of P-gp substrates, which are secreted by a P-gp-mediated efflux system.
Keywords: Intestinal absorption; P-glycoprotein; Transporter; Pharmaceutical excipient; Oral absorption; Polyethylene glycol;

Initial studies of water granulation of eight grades of hypromellose (HPMC) by Jenny Herder; Åsa Adolfsson; Anette Larsson (57-65).
The purpose of this study is to investigate the water granulation mechanism of the hydrophilic matrix polymer HPMC in a high shear mixer and to relate the properties of the granules and tablets to the molecular weight and the degree of substitution for eight HPMC grades. Although the hydrophilic matrix system is a well known drug delivery one, there is a difficulty in that the desirable water granulation technique often causes problems in the presence of relatively large amounts of HPMC due to its hydrophilicity. The results of this study show that the properties of the granules and the tablets fall into two groups according to whether the molecular weight of the polymer is high or low. The granules of low molecular weight were smaller and more compact, with better flow properties but with less tensile strength of the compacts, whereas the opposite was valid for granules of high molecular weight. The explanation for these differences is linked to the proposed granulation mechanism of HPMC, in which the properties of the gel layer are important. The dominant factors governing the properties are the molecular weight and, to lesser extent, the degree of substitution.
Keywords: Hydroxypropyl methylcellulose; Water granulation; Molecular weight; Degree of substitution; High shear granulation;

Metolose–PEG interaction as seen by positron annihilation spectroscopy by K. Pintye-Hódi; G. Regdon; I. Erős; K. Süvegh; T. Marek; I. Kéry; R. Zelkó (66-71).
The plasticizing effects of poly(ethylene glycol) (PEG 400) on methylcellulose (Metolose) cast films were studied by conventional physicochemical methods and positron annihilation spectroscopy. The PEG concentrations relative to the total polymer content were varied within the range 0–75% (w/w). At low concentrations (below 33.3%, w/w), the plasticizer was found to build in into the methylcellulose structure. On the other hand, at higher concentrations (above 50%, w/w), it formed small separate phases in the films.Positron annihilation spectroscopy (PALS) was applied to track the Metolose–PEG interaction. Controlled ageing of Metolose–PEG films at room temperature and at 75% RH revealed a significant difference between the ageing processes of the monophase and those of the separate phase films. The ageing involves two steps in both cases: a fast and a slow one. The PALS measurements demonstrated that the slow process is hindered in the phase-separated samples.
Keywords: Deformation process; Thermal behaviour; Positron annihilation spectroscopy;

Effect of plasticizers on properties of pregelatinised starch acetate (Amprac 01) free films by Giulia Bonacucina; Piera Di Martino; Martina Piombetti; Angela Colombo; Francesco Roversi; Giovanni F. Palmieri (72-77).
Film coating is a technique widely used in the pharmaceutical field to improve and modify technological and release characteristics of capsules, tablets and granules.In this paper physical and mechanical properties of free films of Amprac 01, obtained by the solvent cast method, were studied in order to investigate the film forming ability of this modified starch and the effects of the addition of different plasticizers.A morphological microscopical analysis (SEM) was performed to study surface properties of the films, while thermal analysis (DSC) was carried out to investigate the influence of different types of plasticizers on the glass transition temperature of the polymer.Then a mechanical characterization permitted to evaluate important parameters such as film crack resistance and deformation at break. Extensional creep/relaxation tests were also performed to investigate the viscoelastic characteristics.As clearly demonstrated by the T g values, the residual water present in the films acted as plasticizers, making possible the formation of free films characterised by good macroscopical and mechanical properties.Except glycerol, the kind and amount of the other tested plasticizers did not markedly improve the mechanical and crack resistance of the films.
Keywords: Amprac 01; Free films; Plasticizer;

Gamma-irradiation of lyophilised wound healing wafers by K.H. Matthews; H.N.E. Stevens; A.D. Auffret; M.J. Humphrey; G.M. Eccleston (78-86).
Lyophilised wafers are being developed as drug delivery systems that can be applied directly to the surface of suppurating wounds. They are produced by the freeze-drying of polymer solutions and gels. This study investigates the possibility of sterilising these glassy, solid dosage forms with gamma-irradiation and determining the rheological properties of rehydrated wafers post-irradiation. One series of wafers was formulated using sodium alginate (SA) modified with increasing amounts of methylcellulose (MC), the other being composed of xanthan gum (XG) and MC. Batches were divided into three lots, two of which were exposed to 25 and 40 kGrays (kGy) of Cobalt-60 gamma-irradiation, respectively, the third being retained as a non-irradiated control. Apparent viscosities of solutions/gels resulting from the volumetric addition of distilled water to individual wafers were determined using continuous shear, flow-rheometry. Flow behaviour on proprietary suppurating surfaces was also determined. Large reductions in viscosity were apparent for irradiated SA samples while those of XG appeared to be largely unaffected. In addition, an increase in the yield stress of xanthan formulations was observed. Xanthan wafers appeared to withstand large doses of irradiation with no detrimental effect on the rheology of reconstituted gels. This offers the possibility of manufacturing sterilisable delivery systems for wounds.
Keywords: Lyophilised wafer; Wound healing; Sodium alginate; Methylcellulose; Xanthan; Gamma-irradiation;

Dehydration of trehalose dihydrate at low relative humidity and ambient temperature by Matthew D. Jones; Jennifer C. Hooton; Michelle L. Dawson; Alan R. Ferrie; Robert Price (87-98).
The physico-chemical behaviour of trehalose dihydrate during storage at low relative humidity and ambient temperature was investigated, using a combination of techniques commonly employed in pharmaceutical research. Weight loss, water content determinations, differential scanning calorimetry and X-ray powder diffraction showed that at low relative humidity (0.1% RH) and ambient temperature (25 °C) trehalose dihydrate dehydrates forming the α-polymorph. Physical examination of trehalose particles by scanning electron microscopy and of the dominant growth faces of trehalose crystals by environmentally controlled atomic force microscopy revealed significant changes in surface morphology upon partial dehydration, in particular the formation of cracks. These changes were not fully reversible upon complete rehydration at 50% RH. These findings should be considered when trehalose dihydrate is used as a pharmaceutical excipient in situations where surface properties are key to behaviour, for example as a carrier in a dry powder inhalation formulations, as morphological changes under common processing or storage conditions may lead to variations in formulation performance.
Keywords: Atomic force microscopy; Crystals; Dry powder inhalers (DPIs); Polymorphism; Surface morphology; Transition;

Mass transfer resistance of the dry layer during the primary drying phase of a lyophilizaton cycle is probably the most important factor affecting maximum product temperature and drying time. Product resistance parameters should be determined for each formulation because of their dependence of formulation composition and concentration. The purpose of this study was to determine the dry layer mass transfer resistance, using a simple and rapid method, for various pharmaceutical formulations during primary drying in a laboratory dryer, using monitored product temperature profiles. The mathematical tools used for the determination were a primary drying simulation program in conjunction with Powell's optimization algorithm. For each formulation studied, primary drying was performed using a shelf temperature of −15 or −20 °C and the chamber pressure controlled at 100 mTorr (0.1 Torr). The product temperature profiles (T b) during primary drying were recorded and became the input data for the parameter estimation. The normalized product resistance, R pN, as a function of the dry layer thickness, ℓ , can be described by: R pN = R 0 + A 1 ℓ / ( 1 + A 2 ℓ ) , where the constants R 0, A 1 and A 2 are product resistance parameters of water vapor through the dry layer. Even when the parameter A 1 was negative, indicating that product temperature atypically decreased over time, the dry layer product resistance parameters of the various pharmaceutical formulations could be rapidly and successfully determined using the proposed approach. The product resistance equation obtained in this work for 5% marmitol, expressed as R pN = 0.0002025 + 20.23 ℓ , is similar to that obtained by Pikal [Pikal, M.J., 1985. Use of laboratory data in freeze drying process design: heat and product resistance parameters and the compute simulation of freeze drying. J. Parent. Sci. Technol. 39, 115–138.] using the microbalance method, expressed as R pN = 1.40 + 16.0 ℓ . The product resistance values obtained for the 3% lactose–LDH formulation are also very close to those obtained by (Milton, N., Pikal, M.J., Roy, M.L., Nail, S.L., 1997. Evaluation of manometric temperature measurement as a method of monitoring product temperature during lyophilization. PDA J. Pharm. Sci. Technol. 51, 7–16.) for 5% lactose using the MTM (manometric temperature measurement) method. With the obtained values of the parameters R 0, A 1, and A 2, simulations can be performed to determine the maximum product temperature and the drying time during primary drying. As such, optimum cycle parameters can be determined to avoid collapse of the product. The proposed approach requires only accurately measured product temperature profiles, easily obtained in a laboratory dryer.
Keywords: Collapse temperature; Dry layer product resistance parameters; FORTRAN; Lactose dehydrogenase (LDH); 5% Mannitol; Mass transfer resistance; Micro-collapse; Newton–Ralphson iteration; Powell's optimization algorithm; Primary drying; Primary drying subroutine;

Powder dissolution kinetics have shown that for particles in the so called “large” size regime (more than about 50 μm), the dissolution rate scales as the specific surface area, i.e. rate proportional to d −1 where d is the particle diameter. This is consistent with an effective diffusion boundary layer width h EFF that is constant with respect to particle size. However, for particles in the so called “small” size regime (d less than about 50 μm), the dissolution rate has a stronger dependence than proportional to d −1 [Bisrat, M., Anderberg, E.K., Barnett, M.I., Nystroem, C., 1992. Physicochemical aspects of drug release. XV. Investigation of diffusional transport in dissolution of suspended, sparingly soluble drugs. Int. J. Pharm., 80, 191–201; Mosharraf, M., Nystroem, C., 1995. The effect of particle size and shape on the surface specific dissolution rate of microsized practically insoluble drugs. Int. J. Pharm., 122, 35–47]. In this regime, Prandtl boundary layer theory predicts an h EFF approximately equal to the particle radius or diameter. This paper presents the first experimental determination of h EFF for particles less than about 2 μm. The powder dissolution kinetics of six suspensions over the particle diameter range of 5.9 ± 0.1 to 0.53 ± 0.05 μm are analyzed to yield h EFF values of 8.5 ± 1.9 to 0.34 ± 0.14 μm. The theoretical expectation for mass transport, dissolution time proportional to d 2.0, is in good agreement with the experimental results of dissolution time proportional to d 2.3. An understanding of these mass transfer mechanisms allows pharmaceutical scientists to achieve targeted release rates with minimum ensemble instability.
Keywords: Hydrodynamic boundary layer; Nanoparticles; Powder dissolution; Solubility limited; Prandtl boundary layer theory; Diffusion limited;

This study focuses on the design and evaluation of chitosan-based films intended for wound dressing application. Films of chitosan and their blends with cornstarch and dextran were developed to improve the films’ physical strength. Polypropylene glycol at concentrations of 0.5, 1.0 and 1.5% (w/v) was added to improve the films’ flexibility. Some properties required for successful wound dressing, such as liquid adsorption, vapor and oxygen penetration, bioadhesiveness, and film elasticity, were examined. Chitosan films showed the highest liquid adsorption and the adsorption tended to decrease with addition of cornstarch and dextran. Moisture vapor and oxygen were found to be able to penetrate through all film formulations, and those films with cornstarch and dextran showed increased penetration rates through the films. The bioadhesiveness test using a pig gut model did not show significantly different bioadhesive properties with the addition of cornstarch and dextran. The film elasticity of the formulation containing only chitosan exhibited the lowest elongation of the film at a force of 2 N, but increased with the addition of cornstarch and dextran, respectively. In conclusion, the design and development of chitosan as a wound dressing can be improved by the addition of cornstarch or dextran, and propylene glycol to obtain the films with optimal properties for wound management.
Keywords: Wound dressing; Chitosan hydrogels; Cornstarch; Dextran; In vitro evaluation;

Radiosterilisation of indomethacin PLGA/PEG-derivative microspheres: Protective effects of low temperature during gamma-irradiation by Ana Fernández-Carballido; Patricia Puebla; Rocío Herrero-Vanrell; Pilar Pastoriza (129-135).
Currently, γ-irradiation seems to be a good method for sterilising drug delivery systems made from biodegradable polymers. The γ-irradiation of microspheres can cause several physicochemical changes in the polymeric matrix. These modifications are affected by the temperature, irradiation dose and nature of the encapsulated drug and additives. This study has aimed to evaluate the influence of temperature during the sterilisation process by gamma irradiation in indomethacin PLGA microspheres including a PEG-derivative. Microspheres were prepared by the solvent evaporation method from o/w emulsion and were then exposed to γ-irradiation. A dose of 25 kGy was used to ensure effective sterilisation. Some microspheres were sterilised with dry ice protection that guaranteed a low temperature during the process whilst others were sterilised without such dry ice protection. The effects of γ-irradiation on the characteristics of non-loaded PLGA/PEG-derivative and indomethacin loaded PLGA/PEG-derivative microspheres with and without protection were studied. Non-protected microspheres showed changes in their morphological surface, polymer glass transition temperature, molecular weight and release rate of indomethacin after sterilisation. However, microspheres sterilised with protection did not show significant differences after γ-irradiation exposure. The sterilisation method was satisfactory when the indomethacin loaded microspheres including a PEG-derivative were exposed to γ-irradiation at low temperature.
Keywords: Microspheres; PLGA; Intraarticular; PEG-derivative; Gamma irradiation;

Modified polysaccharides have been used widely to increase physico-chemical stability of liposomes. However, the stability and pharmacokinetic studies on the polysaccharides modified anchored liposomes containing hydrophobic drugs which exist in lipid bilayer membranes were insufficient as compared with the liposomes carrying hydrophilic or ionic drugs in inner aqueous phase. In the present study, a hydrophobic drug, dipyridamole (DIP), was entrapped into liposomes through film hydration. Amylopectin was palmitoylated and anchored on the surface of plain DIP liposomes. Subsequently, the stabilities of DIP ethanol solution, plain DIP liposomes (PDL) and anchored DIP liposomes (ODL) against irradiation, disperse medium, biofluid, long-term storage were determined and compared. The concentrations of DIP in plasma of rats and its pharmacokinetic behaviors after intravenous administration of DIP injection, PDL and ODL were studied by RP-HPLC. The pharmacokinetic parameters were computed by software 3p97 programme. The results showed that ODL could increase stabilities more of DIP in vitro as compared with PDL. The plasma concentration–time curves of DIP after intravenous administration of DIP injection, PDL and ODL were all in accordance with open two-compartment model. Pharmacokinetic parameters of DIP injection, PDL and ODL in rats were significantly different. The present findings suggest that anchored liposomes could increase stabilities of DIP in vitro as compared with plain liposomes. Furthermore, the difference of pharmacokinetic profiles was due to the targetability of anchored liposomes.
Keywords: O-Palmitoyl amylopectin; Liposomes; Dipyridamole; Stabilization; Pharmacokinetics;

Orally administered tamoxifen undergoes a first-pass metabolism and substrates for multidrug resistance (MDR) transporters efflux in the liver and intestines, which obstructs its systemic exposure. This study investigated the effect of quercetin, a dual inhibitor of CYP3A4 and P-gp, on the bioavailability and pharmacokinetics of tamoxifen and one of its metabolites, 4-hydroxytamoxifen, in rats. The pharmacokinetic parameters of tamoxifen and 4-hydroxytamoxifen in plasma were determined after orally administering tamoxifen (10 mg/kg) with or without quercetin (2.5, 7.5 and 15 mg/kg). The coadministration of quercetin (2.5 and 7.5 mg/kg) significantly (p  < 0.05) increased the absorption rate constant (K a), peak concentration (C max) and the areas under the plasma concentration-time curve (AUC) of tamoxifen. The absolute bioavailability (AB%) of tamoxifen with 2.5 and 7.5 mg/kg quercetin ranged from 18.0% to 24.1%, which was significantly higher than the control group, 15.0% (p  < 0.05). The relative bioavailability (RB%) of tamoxifen coadministered with quercetin was 1.20–1.61 times higher than the control group. The coadministration of quercetin caused no significant changes in the terminal half-life (t 1/2) and the time to reach the peak concentration (T max) of tamoxifen.Compared with the control group, the coadministration of 7.5 mg/kg quercetin significantly (p  < 0.05) increased the AUC of 4-hydroxytamoxifen. However, the metabolite ratios (MR; AUC of 4-hydroxytamoxifen to tamoxifen) were significantly lower (p  < 0.05). This suggests that quercetin inhibits the both MDR transporters efflux and first-pass metabolism of tamoxifen. The enhanced bioavailability of tamoxifen as a result of its coadministration with quercetin might be due to the effect of quercetin promoting the intestinal absorption and reducing the first-pass metabolism of tamoxifen. If the results are further confirmed in the clinical trials, the tamoxifen dosage should be adjusted when tamoxifen is administered with quercetin or quercetin-containing dietary supplements in order to avoid potential drug interactions.
Keywords: Tamoxifen; 4-Hydroxytamoxifen; Quercetin; Bioavailability; Pharmacokinetics;

A blend of floating and pulsatile principles of drug delivery system would have the advantage that a drug can be released in the upper GI tract after a definite time period of no drug release. A multiparticulate floating-pulsatile drug delivery system was developed using porous calcium silicate (Florite RE®) and sodium alginate, for time and site specific drug release of meloxicam. Meloxicam was adsorbed on the Florite RE® (FLR) by fast evaporation of solvent from drug solution containing dispersed FLR. Drug adsorbed FLR powder was used to prepare calcium alginate beads by ionotropic gelation method, using 32 factorial design. Developed formulations were evaluated for yield, entrapment efficiency, image analysis, surface topography, mechanical strength, apparent density, buoyancy studies and dissolution studies. Entrapment efficiency of different formulations varied from 70% to 94%. Formulations show a lag period ranging from 1.9 to 7.8 h in acidic medium followed by rapid release of meloxicam in simulated intestinal fluid USP, without enzymes (SIF). Complete drug release in SIF occurred in less than 1 h from the formulations. The size of beads varied from 2.0 to 2.7 mm for different batches. Prepared beads were spherical with crushing strength ranging from 182 to 1073 g. Floating time was controlled by density of beads and hydrophobic character of drug. A pulsatile release of meloxicam was demonstrated by a simple drug delivery system which could be useful in chronopharmacotherapy of rheumatoid arthritis.
Keywords: Floating-pulsatile drug delivery system; Porous calcium silicate; Meloxicam; Rheumatoid arthritis;

Oral delivery of diclofenac sodium using a novel solid-in-oil suspension by Hongyu Piao; Noriho Kamiya; Junji Watanabe; Hideakira Yokoyama; Akihiko Hirata; Takeru Fujii; Ichiro Shimizu; Susumu Ito; Masahiro Goto (159-162).
The present work reports on a new pharmaceutical formulation for oral delivery of diclofenac sodium (DFNa), a non-steroidal anti-inflammatory drug (NSAID). Although DFNa itself is water-soluble at neutral pH, it was readily suspended in soybean oil via complex formation with an edible lipophilic surfactant and a matrix protein. The resulting solid-in-oil (S/O) suspension containing stably encapsulated DFNa in an oil phase markedly reduced the risks for gastrointestinal ulcers upon oral administration even at the LD50 level in rats (ca. 50 mg/kg DFNa). In addition, plasma concentration of DFNa upon administration of an S/O suspension was comparable with that of the aqueous counterpart at the same DFNa dose. These results indicate the potential use of S/O suspensions as novel oil-based pharmaceutical formulations for oral delivery of water-soluble drugs without causing severe mucitis.
Keywords: Diclofenac sodium; Gastric ulcer; NSAID; Oral administration; Solid-in-oil suspension;

Potential interactions between model drugs (benzoates, diltiazem, cyanocobalamin, dextrans) and a thermoresponsive poly(N-isopropylacrylamide) (PNIPA) hydrogel and corresponding linear polymer were investigated. The influence of the drugs on the equilibrium swelling level of the hydrogel was examined and drug–hydrogel binding isotherms were established where appropriate. Differential scanning calorimetry (DSC) was used to investigate the influence of the drugs on the lower critical solution temperature (LCST) of the linear polymer solution. Phase solubility studies were preformed to investigate binding. Drug–polymer co-precipitated blends were also prepared and analysed by X-ray diffraction (XRD), thermal analysis and Fourier transform infrared (FT-IR) spectroscopy. Hydrophobic binding was apparent between PNIPA and the aromatic ring/ester side chain of the unionised benzoate. The effect of this binding on hydrogel swelling was clarified in terms of the influence of the binding on the LCST of the system. The drug release rates of the benzoates from the hydrogel were shown to be dependent on drug binding properties. Ionisation of the benzoate prevented such hydrophobic binding, with a weaker salting out effect apparent with sodium benzoate. Significant interactions between diltiazem, cyanocobalamin (Vitamin B12) or the dextrans and PNIPA were not apparent. High concentrations of the hydrophilic drugs did, however, interfere with the magnitude of hydrogel equilibrium swelling. Hydrophobic binding, the salting out effect and the influence of the drugs on hydrogel swelling under non-sink conditions were therefore shown to be important effects which depended on the chemical nature of the drug present.
Keywords: Drug–hydrogel interactions; Thermoresponsive poly(N-isopropylacrylamide); Binding Isotherm; Hydrophobic binding; Hydrogel swelling;

The biopharmaceutical aspect of the fluoroquinolones–metal cations interaction, which reduces antibacterial agents bioavailability and the mechanism of the fluoroquinolone intestinal efflux are still poorly understood. The purpose of this work was to gain further insights into these two issues by measuring the permeability of ciprofloxacin through the rat small intestine in side-by-side diffusion chambers using different incubation media and transport inhibitors. The permeability of ciprofloxacin from the mucosal to the serosal side was low. It was not influenced by the different concentrations of Ca2+ and Mg2+ in the donor solution. The active efflux of ciprofloxacin was the highest in the region of the rat small intestine excised proximal to the ileo-caecal junction or when the pH value of the incubation saline was slightly acidic. Thus ciprofloxacin appears to be transported in its cationic or in its zwitterionic form. The efflux was not inhibited by verapamil, benzbromarone or quinidine, which were added to the mucosal side of the intestinal tissue. It was however inhibited by quinidine added to the serosal side. The active secretion is therefore most probably a consequence of the organic cation transporter 1 activity at the basolateral membrane of enterocytes.
Keywords: Fluoroquinolones; Ciprofloxacin; Side-by-side diffusion chambers; Metal cations; Efflux proteins; Intestinal elimination;

In vivo distribution and antitumor activity of heparin-stabilized doxorubicin-loaded liposomes by Hee Dong Han; Aeri Lee; Chung Kil Song; Taewon Hwang; Hasoo Seong; Chong Ock Lee; Byung Cheol Shin (181-188).
The purpose of this study was to investigate the effect of heparin conjugation to the surface of doxorubicin (DOX)-loaded liposomes on the circulation time, biodistribution and antitumor activity after intravenous injection in murine B16F10 melanoma tumor-bearing mice. The heparin-conjugated liposomes (heparin-liposomes) were prepared by fixation of the negatively charged heparin to the positively charged liposomes. The existence of heparin on the liposomal surface was confirmed by measuring the changes in the particle size, zeta potential and heparin amount of the liposomes. The stability of the heparin-liposomes in serum was higher than that of the control liposomes, due to the heparin-liposomes being better protected from the adsorption of serum proteins. The DOX-loaded heparin-liposomes showed high drug levels for up to 64 h after the intravenous injection and the half-life of DOX was approximately 8.4- or 1.5-fold higher than that of the control liposomes or polyethyleneglycol-fixed liposomes (PEG-liposomes), respectively. The heparin-liposomes accumulated to a greater extent in the tumor than the control or PEG-liposomes as a result of their lower uptake by the reticuloendothelial system cells in the liver and spleen. In addition, the DOX-loaded heparin-liposomes retarded the growth of the tumor effectively compared with the control or PEG-liposomes. These results indicate the promising potential of heparin-liposomes as a new sterically stabilized liposomal delivery system for the enhancement of the therapeutic efficacy of chemotherapeutic agents.
Keywords: Heparin conjugation; Liposome; Circulation time; Biodistribution;

In the present study predictable pulsatile chronotherapeutics of felodipine (FELO), which is a poorly-water soluble drug, were prepared in the form of two layered tablets. As active core PVP/FELO nanodispersion was used while as effective coating layer different PVP/HPMC blends were added. From dissolution studies of FELO nanodispersions it was revealed that PVP/FELO 90/10 w/w dispersion is an ideal system for pulsatile formulations since the whole amount of FELO is released within the first 30 min. This dissolution enhancement and fast release was attributed to FELO amorphisation, as was found from XRD and DMTA techniques and the effective particle size reduction. Transmission electron microscopy (TEM) studies revealed that FELO creates amorphous nanodispersions into the PVP matrix while particle sizes are directly dependable upon FELO concentration. Drug particles with sizes lower than 150 nm may be the optimal level for a substantial enhancement of FELO dissolution rate. The time of FELO release initiated by the two-layered tablets was adequately adjusted by using different PVP/HPMC blends as coating layer, which is a swellable and erodible barrier. The delaying time of FELO release is directly depended by HPMC concentration and this correlation was mathematically expressed. The significance of these blends is that they are completely miscible over the entire compositional range, thus forming a new matrix with different physicochemical properties, contrary to the initial polymers.
Keywords: Felodipine; Amorphous nanodispersions; Miscible blends; Pulsatile chronotherapeutics;

Diffusion and mathematical modeling of release profiles from nanocarriers by Letícia Cruz; Leonardo U. Soares; Teresa Dalla Costa; Graziela Mezzalira; Nadya P. da Silveira; Sílvia S. Guterres; Adriana R. Pohlmann (198-205).
The aim of this work was to establish models and to differentiate the kinetic release behavior of drug models from nanocapsules, nanoemulsion and nanospheres by physico-chemical characterization and release experiments. SAXS analysis showed that the polymer is organized in the nanocapsules, while in the nanospheres the sorbitan monostearate is organized and acts as an impurity of the poly(ɛ-caprolactone) suggesting that constituents in these nanocarriers are differently organized. Formulations presented particle sizes ranging from 178 to 297 nm, probe content from 0.981 to 0.997 mg/mL, pH values from 4.90 to 5.10 and zeta potential from −37.9 to −51.9 mV. The kinetic experiments showed that the nanostructures present similar behaviors when the probe is adsorbed on the nanocarriers (indomethacin-loaded formulations). However, when the probe is entrapped within the nanocarriers (indomethacin ethyl ester-loaded formulations), nanocapsules, nanospheres and nanoemulsion presented different kinetic behaviors. Mathematical modeling of the release profiles was conducted, showing that the presence of the polymer increases the half-lives of the burst phases (5.9, 4.4 and 2.7 min) while the presence of the oil increases the half-lives of the sustained phases (288.8, 87.7 and 147.5 min) for nanocapsules, nanospheres and nanoemulsion, respectively.
Keywords: Nanoparticles; Nanocapsules; Indomethacin; Alkaline hydrolysis; Poly(ɛ-caprolactone); Indomethacin ethyl ester;

Lecithin-based cationic nanoparticles as a potential DNA delivery system by Zhengrong Cui; Fu Qiu; Brian R. Sloat (206-213).
Previously, we have reported a novel nanoparticle-based DNA vaccine delivery system, which elicited strong immune responses against antigens of interest encoded by the DNA. The nanoparticles were engineered by cooling pre-formed warm microemulsions comprised of emulsifying wax as the oil phase and hexadecyltrimethyl ammonium bromide (CTAB) as the surfactant. However, the poor aqueous stability of the nanoparticles and the emulsifying wax in the nanoparticles may severely limit the applications of the nanoparticles. In the present study, we used lecithin, a more biocompatible material, instead of emulsifying wax, to prepared lecithin-based cationic nanoparticles. The 50% growth inhibition concentration (IC50) of the lecithin-based nanoparticles was found to be more than 1000-fold higher than that of the emulsifying wax-based nanoparticles. Moreover, the stability of the lecithin nanoparticles was also significantly increased. The size of the nanoparticles did not significantly change during a 6-month storage period at room temperature. Finally, when plasmid DNA was adsorbed on their surface, the lecithin nanoparticles successfully transfected cells in culture. These lecithin-based nanoparticles may hold great potentials as a DNA (vaccine) delivery system.
Keywords: Cytotoxicity; Stability; Transfection;

Identification of degradation products from aqueous carboplatin injection samples by electrospray mass spectrometry by Kannan Vivekanandan; Madugundu Guru Swamy; Sudhanand Prasad; Golak Chandra Maikap; Rama Mukherjee; Anand C. Burman (214-221).
Four different carboplatin injection samples in water (∼10 mg/ml) stored at room temperature were investigated for degradation products by electrospray liquid chromatography-mass spectrometry (ESI/LC–MS). A mass spectrometer compatible mobile phase system with 0.02% formic acid and methanol was used to resolve the impurities. Possible chemical structures of the unknown impurities and its degradation mechanism were proposed based on its experimental results and literature findings.
Keywords: Carboplatin; Electrospray; LC–MS;

Noticeboard (222-224).