The interprocess NIR sampling as an alternative approach to multivariate statistical process control for identifying sources of product-quality variability.

We are presenting a new approach of identifying sources of variability within a manufacturing process by NIR measurements of samples of intermediate material after each consecutive unit operation (interprocess NIR sampling technique). In addition, we summarize the development of a multivariate statistical process control (MSPC) model for the production of enteric-coated pellet product of the proton-pump inhibitor class. By developing provisional NIR calibration models, the identification of critical process points yields comparable results to the established MSPC modeling procedure. Both approaches are shown to lead to the same conclusion, identifying parameters of extrusion/spheronization and characteristics of lactose that have the greatest influence on the end-product's enteric coating performance. The proposed approach enables quicker and easier identification of variability sources during manufacturing process, especially in cases when historical process data is not straightforwardly available. In the presented case the changes of lactose characteristics are influencing the performance of the extrusion/spheronization process step. The pellet cores produced by using one (considered as less suitable) lactose source were on average larger and more fragile, leading to consequent breakage of the cores during subsequent fluid bed operations. These results were confirmed by additional experimental analyses illuminating the underlying mechanism of fracture of oblong pellets during the pellet coating process leading to compromised film coating.

Use of factorial design for evaluation of factors affecting the chemical stability of sirolimus (rapamycin) in solid dosage form.

Effects of four process and formulation parameters (spraying rate of ethanol solution, drying and tablet hardness and hydroxypropyl methyl cellulose (HPMC) content) were evaluated in terms of initial quality of tablets using factorial design approach. For determination of stability of final drug product, the tablets were exposed to stress testing conditions and the three most significant factors were investigated (spraying rate of ethanol solution, drying and HPMC content). Considering the chemical stability of Sirolimus, the following responses were found to be most important: total sum of degradation products, levels of impurity I and assay of isomer C. Investigated factors and their interactions most significantly affected the assay of isomer C in initial and in stressed stability testing samples. The factorial design approach is a very economic way of obtaining the maximum amount of information in a short period of time, which is especially important in studies that include a variety of different factors and their interactions.

Freeze-drying and release characteristics of polyelectrolyte nanocarriers for the mucosal delivery of ovalbumin.

Polyelectrolyte complexes (PEC) consisting of an alginate core entrapping the protein ovalbumin and the chitosan coating were prepared by the self-assembly of oppositely charged polyelectrolytes. The PEC were prepared at pH 4.0 and consisted of alginate, ovalbumin and chitosan in a concentration of 0.5, 0.5 and 0.05 mg/ml, respectively, having a particle size of around 300nm, a zeta potential of -44 mV and a protein association efficiency of 80%. The release of ovalbumin from PEC was mostly dependant on the pH of release medium and the presence of strong electrolytes contributed to higher release. Approximately 90% of the ovalbumin was released in a phosphate buffer media, pH 7.4. The release was lower in media with pH 4.0, reaching the value of app. 40% and 60% of ovalbumin released in water (pH 4.0) and NaCl solution (0.9% w/v, pH 4.0), respectively. In an acidic saline solution, pH 3.0, there was only 5% of ovalbumin release, however, increasing the pH to 6.8, approximately 70% of ovalbumin immediately released from the PEC. The PEC were freeze-dried aided by various excipients. Their efficiency on the redispersibility of the freeze-dried product was evaluated according to the mean particle diameter, polydispersity, average scattering intensity (particle concentration) and visual appearance of the PEC (Tyndal effect). In the presence of trehalose and mannitol, the aggregation and integrity of the PEC were prevented, yielding properties similar to the PEC dispersion before lyophilisation. The surface hydrophobicity of the ovalbumin either free or formulated in the nanocomplexes was determined by the bis-ANS fluorescence intensity, indicating a higher surface hydrophobicity for the PEC. The mild formulation conditions, nanometre-sized particles, high protein association efficiency, pH-dependant release, and modified surface properties are promising factors towards the development of an oral delivery system for protein made by the self-assembly of oppositely charged polyelectrolytes.

Study of Immediate Release Spherical Microparticles Containing Clarithromycin using a Hot-melt Fluid Bed Technique.

The aim of the study was to evaluate a hot-melt technique for preparation of immediate release spherical microparticles containing clarithromycin with acceptable characteristics and process yield. A modified fluid bed apparatus with rotor insert was used to prepare spherical microparticles in the size range of 125-355 µm. Poloxamer 188, PEG-32 glyceryl laurate (Gelucire 44/14) and a mixture of polyethylene glycol (PEG) 4000 with PEG 400 were used as meltable binders. Key process parameters were identified and optimized and their influence on process yield and microparticles characteristics was determined. Microparticles with poloxamer 188 and PEG exhibited relatively good mechanical properties. Process yield was around 70% and 60% in the case of PEG and poloxamer 188 respectively. Microparticles prepared with PEG-32 glyceryl laurate exhibited poor mechanical properties and process yield compared to other microparticles. The process was shown to be limited by the bed temperature, exhibiting the best process stability with poloxamer 188 followed by PEG and PEG-32 glyceryl laurate. Dissolution rate and equilibrium concentration of clarithromycin released from prepared microparticles was improved compared to similar particles prepared by wet granulation.

Self-assembled Polyelectrolyte Nanocomplexes of Alginate, Chitosan and Ovalbumin.

Polyelectrolyte complex (PEC) nanoparticles for delivering model protein drug ovalbumin were prepared from two polysaccharide polymers, alginate and chitosan. The parameters influencing the complex formation were characterised using colloid titration in combination with dynamic light scattering. The polyelectrolyte interactions and morphology of the formed complexes were verified by differential scanning calorimetry and scanning electron microscopy, respectively. The PEC formation was predominantly pH- and concentration-dependent. The complexation of ovalbumin with a negatively charged alginate occurred only at a pH below the isoelectric point of the ovalbumin. After the complexation, negatively charged complexes of alginate and ovalbumin were further coated with chitosan. The optimal composition of the PEC, yielding 280 nm sized particles having a zeta potential -40 mV, was determined for alginate:ovalbumin:chitosan in a mass ratio 1: 1: 0.1, respectively, giving their final concentration 0.5: 0.5: 0.05mg/ml. The loading of ovalbumin in the PEC depended on the initial amount of ovalbumin used to produce the PEC, and ranged from 7-38% for different formulations, however, the association efficiency remained pretty similar for all formulations, i.e. 80-85%. Mild formulation conditions, nanometre-sized particles, and a high protein association efficiency are promising factors towards the development of a delivery system for proteins.

High celecoxib-loaded nanoparticles prepared by a vibrating nozzle device.

Drug delivery research has resulted in the availability of several enabling technologies for formulating poorly water-soluble compounds. In this study the vibrating nozzle device, originally used for encapsulation of drugs, cells and microorganisms, has been used to formulate nanoparticles (NP) with high loading capacity. Celecoxib was incorporated in NP of polylactic acid (PLA) and poly(lactic-co-glycolic acid) (PLGA) and the influence of polymers, initial drug : polymer ratio and stabilizer concentration on NP size and surface properties, entrapment efficiency, drug loading and in vitro release profile were investigated. NP were in the size range of 230-270 nm, with a polydispersity index less than 0.25 and a spherical shape. The highest celecoxib loading (13% w/w) was obtained at initial ratio celecoxib : Resomer RG 502 (PLA/PGA = 50/50) of 1 : 5 and 0.1% w/w polyvinyl alcohol concentration. Thermal analysis and X-ray diffraction suggested that celecoxib was amorphous or molecularly dispersed in the polymeric matrix. The release profile exhibited an initial burst followed by sustained release. The freeze-dried NP could be completely dispersed on addition of lyoprotectants. The production of NP by the vibrating nozzle device is highly reproducible, time saving, can be performed under aseptic conditions and offers the possibility of scale-up.

Preparation and evaluation of celecoxib-loaded microcapsules with self-microemulsifying core.

The purpose of this study was to prepare alginate microcapsules with a self-microemulsifying system (SMES) containing celecoxib in the core. An Inotech IE-50 R encapsulator equipped with a concentric nozzle was used to prepare the microcapsules. The encapsulated SMES was shown to increase celecoxib solubility over that of the pure drug more than 400-fold. Microcapsules prepared with a high SMES:celecoxib ratio exhibited distinct core vesicles containing liquid SMES. By modifying the SMES and including an additional chitosan coating, drug loading in the range from 12-40% could be achieved with the degree of encapsulation ranging from 60-82%. Alginate microcapsules loaded with SMES and celecoxib showed increased dissolution rate of celecoxib over that of alginate microcapsules loaded with celecoxib or of the celecoxib alone. Compared to the previous report, drug loading capacity was significantly improved, enabling the formulation of dosage forms which are of suitable size for peroral application.

Effect of surface hydrophobicity of therapeutic protein loaded in polyelectrolyte nanoparticles on transepithelial permeability.

Oral delivery of protein drugs is greatly limited by low hydrophobicity, an important determinant for intestinal epithelial permeation and bioavailability. Herein, surface properties of recombinant erythropoietin were investigated using the fluorescent dye bis-ANS to monitor relative hydrophobicity for correlation with permeabilities with Caco-2 cells. At various pHs, bis-ANS fluorescence intensity indicated different surface hydrophobicities of erythropoietin molecules. Erythropoietin incorporated in chitosan or chitosan-trimethylchitosan (CS-TMC) nanoparticles prepared by polyelectrolyte complexation and ionotropic gelation with tripolyphosphate also showed different surface hydrophobicities. Chitosan nanoparticles with erythropoietin provided the most hydrophobic surface, followed by free erythropoietin (in water) and that loaded into CS-TMC nanoparticles. Chitosan nanoparticles were more effective than CS-TMC nanoparticles for permeation of erythropoietin across Caco-2 cell monolayers; the lowest permeability was shown by erythropoietin itself. Thus, hydrophilic protein molecules complexed with polyelectrolytes can provide more hydrophobic surfaces that enhance transepithelial permeability. This bis-ANS method also provides valuable information for the design of polyelectrolyte nanoparticules for oral delivery of protein drugs.

A new amoxicillin/clavulanate therapeutic system: preparation, in vitro and pharmacokinetic evaluation.

A new peroral amoxicillin/clavulanate therapeutic system composed of immediate release tablet and controlled release floating capsule was developed and evaluated by in vivo bioavailability study. Pharmacokinetic (PK) parameters for amoxicillin and clavulanic acid of the new therapeutic systems: AUCt, AUCi, (AUCt/AUCi), Cmax, Tmax, kel, T(1/2) and additionally for amoxicillin T(4) and T(2) were calculated from the plasma levels. The study confirmed enhanced pharmacokinetic parameters of a newly developed therapeutic system containing 1500 mg of amoxicillin and 125 mg of clavulanic acid. Prolonged time over MIC of amoxicillin in relation to a regular immediate release amoxicillin/clavulanate formulation was confirmed.

The evaluation of some pharmaceutically acceptable excipients as permeation enhancers for amoxicillin.

The aim of the present study was to evaluate different pharmaceutically acceptable excipients as permeation enhancers for a low permeability drug, amoxicillin. As a model for the intestinal epithelium excised rat jejunum, mounted in side-by-side diffusion cells, was used. Amoxicillin was actively transported across the intestine in the serosal-to-mucosal direction, but only if glucose was present at the mucosal side. This effect of glucose was abolished by a multridrug resistance associated protein (MRP) inhibitor benzbromarone (0.04 mM), but not by verapamil (0.2 mM). Among the tested pharmaceutically acceptable excipients only sodium lauryl sulfate (0.2 mg/ml) increased the permeability of amoxicillin in the mucosal-to-serosal direction, which was accompanying with the abolishment of the secretory oriented transport of amoxicillin. Other excipients (0.07 2mg/ml Pluronic F68, 0.2 mg/ml Lutrol F127, 0.2 mg/ml Cremophor EL or 0.2 mg/ml Carbopol 934) have no influence on the permeability of amoxicillin. The effect of sodium lauryl sulfate on the active secretion of amoxicillin was mainly attributed to the reversible cellular ATP depletion. We concluded that sodium lauryl sulfate can be considered as a relatively safe permeation enhancer for amoxicillin in drug delivery systems intended to improve oral bioavailability of this drug.

In-line NIR monitoring of key characteristics of enteric coated pellets.

We describe the development of an in-line monitoring approach for the fluid-bed drying and coating steps for the production of enteric coated pellets by NIR. Our results show that key pellet characteristics can be monitored in-line. Likewise, the finished product acidic resistance is in excellent agreement to the in-line NIR predictions. Samples were collected at regular intervals and analyzed by several reference methods to characterize both process steps. In-line NIR models for pellets size sieve fractions, residual solvent content, and amount of coating layer have been constructed. Both the pellet coating layer amount and the in-vitro enteric performance demonstrate low variability which represents a challenge to the usual chemometric model development approach. To overcome this challenge a hierarchical PLS model for predicting acidic resistance was successfully constructed using time-evolving spectral data from 22 batches. Moreover, a novel multivariate meta-analysis of the PLS loadings of individual in-line models and the hierarchical PLS model has identified which pellet characteristics correlate most significantly with the observed enteric performance of the finished product. Additionally, the meta-analysis pointed toward the presence of further mechanisms unrelated to studied characteristics that also significantly influence the acidic resistance.

Influence of polymers on the bioavailability of microencapsulated celecoxib.

Celecoxib, a selective COX-2 inhibitor, primarily used in treatment of osteoarthritis, rheumatoid arthritis and acute pain was encapsulated in microparticles composed of various polyesters, polymethacrylates or cellulose derivatives used alone or blended. The influence of polymers on microparticle mean diameter, encapsulation efficiency and in vitro and in vivo celecoxib release was investigated. Microparticles were in the size range 11-37 microm. Encapsulation efficiency was optimal due to poor aqueous solubility of celecoxib. Considering in vitro release, microparticles could be divided into drug delivery systems with fast and slow release profiles. Microparticles prepared with poly-epsilon-caprolactone, Eudragit RS and low viscosity ethylcellulose, together with physical mixture of celecoxib with lactose and Celebrex, were tested in vivo. Relative bioavailability of celecoxib was below 20% in all cases and was probably the consequence of a slow in vivo release of celecoxib from microparticles or low wettability in the case of Celebrex and physical mixture.