European Journal of Pharmaceutics and Biopharmaceutics (v.64, #1)

APV diary (S1-S4).

Transdermal drug delivery system (TDDS) adhesion as a critical safety, efficacy and quality attribute by Anna M. Wokovich; Suneela Prodduturi; William H. Doub; Ajaz S. Hussain; Lucinda F. Buhse (1-8).
Transdermal drug delivery systems (TDDS), also known as “patches,” are dosage forms designed to deliver a therapeutically effective amount of drug across a patient’s skin. The adhesive of the transdermal drug delivery system is critical to the safety, efficacy and quality of the product. In the Drug Quality Reporting System (DQRS), the United States Food and Drug Administration (FDA) has received numerous reports of “adhesion lacking” for transdermal drug delivery systems. This article provides an overview of types of transdermals, their anatomy, the role of adhesion, the possible adhesion failure modes and how adhesion can be measured. Excerpts from FDA reports on the lack of adhesion of transdermal system products are presented. Pros and cons of in vitro techniques, such as peel adhesion, tack and shear strength, in vivo techniques used to evaluate adhesive properties are discussed. To see a decrease in “adhesion lacking” reports, adhesion needs to become an important design parameter and suitable methods need to be available to assess quality and in vivo performance. This article provides a framework for further discussion and scientific work to improve transdermal adhesive performance.
Keywords: Transdermal drug delivery system; Transdermal system; Drug delivery; Patch; Adhesion; Tack; Peel; Shear;

A novel modified film-hydration–dilution method was employed to prepare highly encapsulated interferon-α-2b containing liposomes for intramuscular sustained release. The liposomes produced by this technique were a mixture of mainly unilamellar vesicles and a small number of multilamellar vesicles. The trapping efficiency was above 80%. With at least 60-fold dilution, Triton X-100 at the concentration of 0.3% (w/v) in phosphate buffered saline (PBS) was able to solubilize phospholipids without denaturing the protein and/or interfering with the enzyme-linked immunoassay (ELISA). After three homogenization cycles under a pressure of 70 MPa the size of liposomes was reduced from 978 to 101 nm while the activity of interferon-α-2b decreased by 9.9% compared to the control. Although liposomes were physically stable for 22 months at 4 °C the mean size of the liposomes increased slightly from 101 to 122 nm. The levels of free interferon-α-2b at the site of intramuscular injection decreased rapidly with only 4.15% of initial dose retained at the injection site after 0.33 h following injection of an interferon-α-2b solution (nonencapsulated). In contrast, interferon-α-2b encapsulated in liposomes was retained at the site of intramuscular injection at higher levels than free interferon-α-2b (p  < 0.05). Larger liposomes containing interferon-α-2b (978 nm) were the most effective for local retention because 27.8% of interferon-α-2b was retained after 24 h. These liposomes have the potential to be topically injected for treating genital herpes with prolonged interferon levels at the local injection site. Since the smaller liposomes (75.8 and 101 nm) retained interferon-α-2b at the injection site for shorter times while enhancing the blood circulation of the drug, they are potentially good carriers for systemic therapy with higher bioavailability and liver targeting.
Keywords: Liposome; Interferon-α-2b; Film-hydration–dilution; Sustained-release; Intramuscular injection;

The effects of N-trimethylchitosan (TMC) on the transcorneal transport of dexamethasone, taken as a marker of the transcellular penetration route, and of tobramycin, a marker of the paracellular route, were studied by assessing the TMC effect on the intraocular pharmacokinetics of each marker. The drugs were topically applied via erodible inserts (weight, 20 mg; diameter, 6 mm; drug dose, 0.3 mg) based on poly(ethylene oxide), containing 10% w/w medicated TMC microspheres (diameter <2.5 μm). Before application, drug release and insert erosion kinetics, and release mechanism were studied in vitro. With either drug, introduction of 10% TMC into insert did not substantially alter the release and erosion rates, hence this formulation was apt to isolate the transcorneal penetration enhancing effect of TMC. Ocular pharmacokinetics were determined in the rabbit model. TMC produced significant increases of dexamethasone C max (5.69 ± 0.49 vs. 3.07 ± 0.31 μg/ml) and AUC (619.3 ± 32.5 vs. 380.5 ± 32.0 μg min/ml) in the aqueous with respect to the reference TMC-free insert. On the other hand, TMC was unable to yield tobramycin concentrations in the aqueous exceeding the determination limit (0.5 μg/ml). In conclusion, TMC enhances transcorneal transport via the transcellular route, whereas it is unable to effectively open the tight junctions between corneal cells.
Keywords: Dexamethasone; Tobramycin; N-Trimethylchitosan; Transcorneal absorption; Absorption enhancer; Transcellular transport; Paracellular transport; Ocular drug delivery; Mucosal delivery; Polymeric biomaterials;

Pharmacoscintigraphic evaluation of lipid dry powder budesonide formulations for inhalation by Thami Sebti; Gabrielle Pilcer; Bernard Van Gansbeke; Serge Goldman; Alain Michils; Francis Vanderbist; Karim Amighi (26-32).
Lung deposition of new formulations of budesonide, using solid lipid microparticles (SLmP) as a pharmaceutically acceptable filler and carrier for inhalation aerosols, and administered from a dry powder inhaler (Cyclohaler®), were compared with that from Pulmicort® Turbuhaler®. Six healthy volunteers took part in a three-way randomized cross-over study, and inhaled a nominal dose of 400 μg budesonide, labelled with 99mTc, on each study day. Lung deposition was determined by gamma scintigraphy and by a pharmacokinetic method. The percentage of dose (SD) in the whole lung was 49.9 (3.7)% for the lipidic matricial form (M) and 62.8 (4.9)% for the lipidic physical blend formulation (PB). These results corresponded well with the in vitro fine particle assessment. In comparison with data recorded in literature for in vivo deposition obtained with Pulmicort® Turbuhaler®, it was estimated that lung deposition was 1.5 and 2.0 times higher for the M and PB formulations, respectively. Furthermore, the relative drug availability obtained from the pharmacokinetic evaluation, expressed as the percentage of pulmonary absorption of the comparator product, was 154% and 220% for M and PB, respectively.The results of the present study indicate that pulmonary administration using SLmP gives a prominent and significant increase in budesonide lung deposition.
Keywords: Solid lipid particles; Dry powder inhaler; Gamma scintigraphy; Total and regional lung deposition; Pharmacokinetics;

Development and validation of a capillary electrophoresis method for the determination of sulfate in effervescent tablets by Helen Ho Sung; Evelyne Laborde-Kummer; Karen Gaudin; Jean-Pierre Dubost (33-37).
A suitable capillary electrophoresis (CE) method was developed and validated for sulfate anion determination in effervescent tablets of Digedryl®. The large excess of other ions in the matrix (i.e. excipients) constituted the main difficulty of this method’s development. So an original analytical procedure for both the conditioning and rinsing of the capillary was purposed including a running electrolyte constituted by boric acid 20 mM and hexamethonium dibromide 0.75 mM at pH 8.00. Separation was carried out on a 60.2 cm (50 cm to the detector) × 0.75 μm i.d. fused-silica capillary at a potential of −29 kV and 35 °C. Indirect UV detection was performed at a wavelength of 254 nm using a background electrolyte containing potassium chromate. Nitrate anion was used as an internal standard for quantification. This CE method was validated in terms of selectivity, linearity, accuracy and precision.
Keywords: Capillary electrophoresis; Sulfate anion; Validation; Pharmaceutical analysis; Quality control;

Quantitative measurements of localized density variations in cylindrical tablets using X-ray microtomography by Virginie Busignies; Bernard Leclerc; Patrice Porion; Pierre Evesque; Guy Couarraze; Pierre Tchoreloff (38-50).
Direct compaction is a complex process that results in a density distribution inside the tablets which is often heterogeneous. Therefore, the density variations may affect the compact properties. A quantitative analysis of this phenomenon is still lacking. Recently, X-ray microtomography has been successfully used in pharmaceutical development to study qualitatively the impact of tablet shape and break-line in the density of pharmaceutical tablets. In this study, we evaluate the density profile in microcrystalline cellulose (Vivapur 12®) compacts obtained at different mean porosity (ranging from 7.7% to 33.5%) using X-ray tomography technique. First, the validity of the Beer–Lambert law is studied. Then, density calibration is performed and density maps of cylindrical tablets are obtained and visualized using a process with colour-scale calibration plot which is explained. As expected, important heterogeneity in density is observed and quantified. The higher densities in peripheral region were particularly investigated and appraised in regard to the lower densities observed in the middle of the tablet. The results also underlined that in the case of pharmaceutical tablets, it is important to differentiate the mechanical properties representative of the total volume tablet and the mechanical properties that only characterize the tablet surface like the Brinell hardness measurements.
Keywords: X-ray tomography; Compaction; Tablet; Density distribution; Anisotropy; Quantitative analysis; 3D imaging; Mechanical properties;

Investigation and modelling approach of the mechanical properties of compacts made with binary mixtures of pharmaceutical excipients by V. Busignies; B. Leclerc; P. Porion; P. Evesque; G. Couarraze; P. Tchoreloff (51-65).
Three pharmaceutical excipients (microcrystalline cellulose, lactose, anhydrous calcium phosphate) and their binary mixtures were compacted to form compacts of various mean porosities. Some mechanical properties (Young’s modulus, tensile strength and Brinell hardness) were studied on these compacts. The mechanical properties of the binary mixtures were not proportional to the mixture composition expressed in mass. More, for all the properties, a negative deviation was always observed from this linear relationship. In reference to a composition percolation phenomenon, critical mass fractions were detected from the graph mechanical property vs. mass composition of a mixture. The results obtained with Brinell hardness differed from the results of the Young’s modulus and the tensile strength, i.e. the most plastic material in the binary mixture controlled the mixture behaviour. Secondly, a predictive model based on a statistical approach was proposed for the Young’s modulus and the tensile strength. The validity of this model was verified on experimental data, and an interaction parameter used to characterize the affinity of the two compounds was calculated. Finally, the X-ray tomography technique was applied to the compacts of cellulose/phosphate mixtures to obtain cross-sections images of the compacts. The analysis of the cross-sections images allowed explaining the no linear relationship of the different mechanical properties results observed on these binary mixtures.
Keywords: Binary mixtures; Compactibility; Tensile strength; Young’s modulus; Brinell hardness; Modelling; X-ray tomography; Percolation;

Compaction behaviour and new predictive approach to the compressibility of binary mixtures of pharmaceutical excipients by V. Busignies; B. Leclerc; P. Porion; P. Evesque; G. Couarraze; P. Tchoreloff (66-74).
The compressibility of three pharmaceutical excipients (microcrystalline cellulose, lactose and anhydrous calcium phosphate) and their binary mixtures was studied. The aim of this work was to observe the impact of the mass composition of the mixture on the compressibility. The single-compound materials and their mixtures were compacted using instrumented presses. It allowed obtaining compression cycles (i.e., force–displacement curves) which were associated with energy measurements (specific compaction energy, E sp cp and specific expansion energy, E sp exp). It was observed that for the mixtures studied, the change of E sp cp with the mass composition could be fitted using a linear relationship (it was not the case with E sp exp). A linear relationship between the porosity of mixture’s compacts and the mass composition was also obtained. Heckel’s plots were then obtained for the three excipients and the mixtures. The mean yield pressure was calculated with the “in-die-method” and the “out-of-die method”. A proportional relationship was not valid for the mean yield pressures. But, a predictive approach was proposed in order to obtain indirectly the mean yield pressure of a binary mixture if the data of the single materials were known. It used the linear mixing rule observed with the porosity. The validity was verified and compared with the experimental values. This comparison showed that it was possible to predict the mean yield pressure of binary mixtures from the accessible data of the single excipients.
Keywords: Binary mixtures; Compressibility; Porosity; Energetic parameters; Yield pressure; Modelling;

The use of Agar as a novel filler for monolithic matrices produced using hot melt extrusion by John G. Lyons; Declan M. Devine; James E. Kennedy; Luke M. Geever; Patrick O’Sullivan; Clement L. Higginbotham (75-81).
The use of filler materials in an extended release monolithic polymer matrix can lead to a vastly altered release profile for the active pharmaceutical ingredient. A range of excipients for use in monolithic matrices have been discussed in the literature. The body of work described in this research paper outlines the use of agar as a novel filler material in a hot melt extruded polymer matrix. Several batches of matrix material were prepared with Diclofenac sodium used as the active pharmaceutical ingredient (API). Agar and microcrystalline cellulose were used as the filler materials in varying ratios, to examine the effect of % filler content as well as filler type on the properties of the hot melt extruded matrix. The resultant extrudates were characterised using steady state parallel plate rheometry, differential scanning calorimetry (DSC) and dissolution testing. The rheometry analysis concluded that the fillers used resulted in an increase in the matrix viscosity. The DSC scans obtained showed negligible effects on the melting behavior of the matrix as a result of the filler inclusion. Dissolution analysis showed that the presence of the fillers resulted in a slower release rate of API than for the matrix alone. The results detailed within this paper indicate that agar is a viable filler for extended release hot melt produced dosage forms.
Keywords: Agar; Polyethylene oxide; Eudragit; Matrix; Hot melt extrusion; Controlled drug delivery devices;

Enhancing mechanism of Labrasol on intestinal membrane permeability of the hydrophilic drug gentamicin sulfate by Kenjiro Koga; Yoichi Kusawake; Yukako Ito; Nobuyuki Sugioka; Nobuhito Shibata; Kanji Takada (82-91).
The aim of this study was to clarify the mechanism by which caprylocaproyl macrogol-8 glyceride (Labrasol) enhances the intestinal absorption of gentamicin sulfate (GM), a drug that has poor permeability but relatively high solubility. We studied the following characteristics: (i) the phase behavior of Labrasol in aqueous solution, (ii) the affinity of GM to Labrasol micelles, and (iii) the interaction between Labrasol and membrane lipids. We measured the critical micelle concentration of Labrasol in aqueous solution to be approximately 0.01%. The average diameters of Labrasol micelles in 2% and 25% solutions were approximately 10 nm and 20 nm, respectively, indicating that micelles increase in size with increasing Labrasol concentration. Although GM dissolved in 5% Labrasol solution was dialyzable, GM dissolved in either 25% or 50% Labrasol solutions was not, suggesting that GM exists in the hydrophilic region of the Labrasol micelle or in the high affinity region of the micelle surface where GM is retained. In membrane permeability experiments and electrophysiological studies conducted with rat ileum, only 25% Labrasol solution enhanced GM permeability, but did not remarkably affect membrane resistance. Furthermore, Labrasol increased membrane lipid fluidity as determined by fluorescence anisotropy in porcine intestinal brush border membrane liposomes. These results suggest that high concentrations of Labrasol solution enhance membrane permeability of GM via a transcellular rather than a paracellular route. We propose the following mechanism: Labrasol micelles grow when the concentration of Labrasol exceeds 20%, at which point GM shows high affinity for the hydrophilic region of the micelles. Since Labrasol micelles increase membrane lipid fluidity, the membrane permeability of GM is concomitantly enhanced.
Keywords: Caprylocaproyl macrogol glyceride; Micelle shape; Fluorescence anisotropy; Intestinal absorption; Membrane lipid fluidity;

Application of an acceptance sampling plan for post-production quality control of chemotherapeutic batches in an hospital pharmacy by Isabelle Borget; Isabelle Laville; Angelo Paci; Stefan Michiels; Lionel Mercier; Romain-Pacôme Desmaris; Philippe Bourget (92-98).
About 26,000 chemotherapeutic batches were produced in 2003 in the Institute Gustave Roussy and 83% were qualitatively and quantitatively assessed in post-production controls via an analytical platform. The rate of non-conformity (outside the specification limits of the target concentration ±10%) decreased from 8.9% to 2.2% between years 2001 and 2003. A cost- and time-saving acceptance sampling plan was applied to assay fewer batches whilst maintaining an accurate estimate of the quality level.The opportunity to apply a single sampling plan by attributes with an acceptance quality level of 2.2% was evaluated. A prognostic study using a logistic regression model was performed for some drugs to identify risk factors associated with the non-conformity rate of preparations.Out of 26 drugs, 17 have not been sampled, since they were prepared less than 400 times per year. For six drugs, a reduction of about 50% in the number of assays was estimated. Three drugs were “at risk” of being non-conform: for these drugs, all batches were analysed.The sampling plan allowed a reduction of almost 8000 analyses with respect to the number of batches analysed for 6 drugs. For the 3 drugs with the higher risk to be non-conform, associated risk factors were identified to set up corrective actions.
Keywords: Chemotherapy; Quality assurance; Sampling plan; Post-production controls; High performance thin-layer chromatography (HPTLC); Cancer;

Evaluation of the antioxidant activity of soybean extract by different in vitro methods and investigation of this activity after its incorporation in topical formulations by Sandra Regina Georgetti; Rúbia Casagrande; Fabiana Testa Moura-de-Carvalho Vicentini; Waldiceu A. Verri; Maria José Vieira Fonseca (99-106).
Chemoprevention by natural products is an emerging therapeutic approach for free radical-mediated diseases including cancer. This is a consequence of its wide applicability and acceptance. In the present study, the antioxidant activity of the soybean extract (Isoflavin Beta®) and of formulations added with this extract were evaluated using stable free radical 2,2-diphenyl-1-pycrylhydrazyl (DPPH) and deoxyribose as well as the lipid peroxidation inhibition assays. For all the assays the extract showed a dose-dependent activity, and IC50 of 21.03 μg/mL in lipid peroxidation inhibition, 161.8 μg/mL in DPPH, and 33.5 ng/mL in hydroxyl radical scavenging assay. The antioxidant activity of the extract added in the formulations could not be assessed using the deoxyribose assay. However, the lipid peroxidation inhibition and DPPH scavenging assays could be successfully applied for the antioxidant activity evaluation of the formulations added with soybean extract to protect the skin against free radicals, which can be generated by the ultraviolet radiation exposure.
Keywords: Soybean extract; Antioxidant; Polyphenols; Topical formulation; DPPH; Deoxyribose; Lipid peroxidation;

Assessing the re-crystallization behaviour of amorphous lactose using the RH-perfusion cell by Inga-Lis Timmermann; Hartwig Steckel; Michael Trunk (107-114).
Many different reports have studied the crystallization behaviour of lactose, e.g., by exposing samples of amorphous lactose to different relative humidity at constant temperatures. However, only few reports are available investigating the formation of α-lactose monohydrate and β-lactose during re-crystallization.Applying the static ampoule method in the microcalorimeter, the enthalpies of amorphous lactose were reported to be constantly 32 and 48 J/g, respectively, considering the mutarotation of lactose at 25 °C and 58% RH, 75% RH and 100% RH. In this study, an alternative microcalorimetric technique, the relative humidity-perfusion cell (RH-perfusion cell) was chosen. The RH-perfusion cell is able to deliver a constant and controlled flow of humidified air to the sample. Investigated compounds were purely amorphous lactose and different powder mixtures of lactose. They consisted of α-lactose monohydrate (Pharmatose 325M®), β-lactose (Pharmatose DCL21®) or a combination (1:1) thereof as carriers, and different concentrations of amorphous lactose. The determination of the enthalpy of desorption of the just re-crystallization lactose by the RH-perfusion cell was used to discriminate whether the monohydrate or the anhydrous form of lactose was produced. Differences in the re-crystallization behaviour of lactose at 25 °C and 58–100% RH were found. At 60–80% RH purely amorphous lactose showed a high heat of desorption which can be attributed to a very high content of formed β-lactose. Powder mixtures containing high contents of amorphous lactose (8% and 15%, respectively) blended with α-lactose monohydrate as a carrier resulted in similar results at the same RH ranges. The high amount of β-lactose can be due to the equilibrium anomeric composition. Whereas powder mixtures containing β-lactose as a carrier and amorphous lactose in a concentration of 1%, 8% and 15%, respectively, formed less β-lactose than the mixtures containing α-lactose monohydrate as a carrier. At a relative humidity of 90% none of the powder mixtures showed desorption as to the fact that in all cases only α-lactose monohydrate was formed at the surface of the re-crystallized lactose. Furthermore, mixtures of α-lactose monohydrate and β-lactose (1:1) and 8% amorphous lactose were investigated. An increase in formed α-lactose monohydrate by increasing RH was found. To consolidate the results, the same mixtures were re-crystallized at different RH in desiccators and subsequently investigated in the solution calorimeter. The results of the pre-mix were confirmed by the solution calorimeter. In summary, purely amorphous lactose and mixtures containing α-lactose monohydrate as a carrier show different re-crystallization behaviour compared to mixtures containing β-lactose as a carrier.
Keywords: α-Lactose monohydrate; β-Lactose; Amorphous lactose; Microcalorimetry; Solution calorimeter; Perfusion cell;

The aim of the present study was to prepare pulsatile release formulations consisting of two-layered tablets appropriate for preventing ischemic heart diseases. For this reason the active core was constituted by a FELO/PVP 10/90 w/w solid dispersion while for the adjustment of the drug release time the coating layer was composed of PVP/HPMC blends at different compositions, acting as a stimulus responsible layer. These blends as was found by DSC studies are miscible in the entire composition range, ensured by the interactions taking place between hydroxyl groups of HPMC and carbonyl groups of PVP. The miscibility of the system enhances the mucoadhesive properties of the blends, compared with those of pure HPMC, which is desired for such applications. The enhancement was attributed to the higher rate of wetting and flexibility of the new matrices due to the faster dissolution of the PVP macromolecules. Upon exposure of the prepared tablets to the release medium it was found that the coating layer disintegrates first, followed by the immediate release of FELO from the active core. The delaying time is based on a complicated mechanism, which is a combination of swelling and erosion of the PVP/HPMC polymer blends. Varying the PVP/HPMC blend ratios, the exact time that FELO is released during a daytime can be effectively adjusted and this ability is expressed mathematically by the equation t  = 0.028 C 1.5, where C is the concentration of HPMC in the blend.
Keywords: Felodipine; Poly(vinyl pyrrolidone); Hydroxypropyl methyl-cellulose; Miscible blends; Pulsatile chronotherapeutics; Mucoadhesive polymers;

by R. Troschütz (127).