Proofs of the Structure of Lipid Coated Nanoparticles (SMBV™) Used as Drug Carriers

Purpose. Supramolecular Biovectors (SMBV) consist of cross-linkedcationic nanoparticles surrounded by a lipid membrane. Thepurpose was to study the structure of the lipid membrane and tocharacterise its interaction with the nanoparticles in order to differentiateSMBV from other polymer/lipid associations. Methods. The interaction of lipids with the nanoparticle surface wasstudied using zeta potential, Fluorescence Energy Transfer (FET) andFluorescence Microscopy. SMBV were compared to liposomes andmixtures nanoparticles/liposomes. Finally the structure of SMBVwas visualised by Electron Microscopy. Results. Zeta potential measurements showed that lipids on SMBVhad a pronounced shielding effect on the surface charge. This was notthe case for mixtures of nanoparticles and liposomes. FET experimentsconfirmed these results indicating that, for SMBV, the lipids aremuch closer to the nanoparticle surface. SMBV Fluorescence microscopyon model microparticles showed a lipid crown on SMBV thatwas confirmed by electron microscopy on SMBV nanoparticles. Conclusions. Results show that in case of SMBV lipids are stronglyadsorbed on the polysaccharide core surface probably due to ionic/hydrophobicinteractions. The resulting supramolecular structure is aspherical cationic polysaccharide particle surrounded by a phospholipid/cholesterol layer.     

Characterization of Particle-Interactions by Atomic Force Microscopy: Effect of Contact Area

Purpose. The purpose of this work was to compare adhesion forces, contact area, and work of adhesion of salbutamol sulphate particles produced using micronization and a supercritical fluid technique (solution-enhanced dispersion by supercritical fluids - SEDS) using atomic force microscopy (AFM). Methods. Adhesion forces of individual particles of micronized and SEDS salbutamol against a highly orientated pyrolytic graphite surface were acquired in a liquid environment consistent with that of a pressurized metered dose inhaler. The forces were then related to contact area and work of adhesion. Results. The raw adhesion force data for the micronized and SEDS material were 14.1 nN (SD 2.5 nN) and 4.2 nN (SD 0.8 nN), respectively. After correction for contact area, the forces per unit area were 13 mN/m² (SD 2.3 mN/m²) and 3 mN/m² (SD 0.6 mN/m²). The average work of adhesion was calculated using the Johnson-Kendall-Roberts theory and was found to be 19 mJm^–2 (SD 3.4 mJm^–2) for the micronized particle and 4 mJm^–2 (SD 0.8 mJm^–2) for the SEDS particle. Conclusions. It is possible to produce a three-dimensional representation of the contact area involved in the interaction and make quantitative comparisons between different particles. There was a lower force per unit area and work of adhesion observed for the SEDS material, possibly because of its lower surface free energy.     

Effect of Colonic Lactulose Availability on the Timing of Drug Release Onset in Vivo from a Unique Colon-Specific Drug Delivery System (CODES™)

Purpose. To test the hypothesis that the onset of drug release in vivo from a unique colon-specific drug delivery system (CODES) would depend on the colonic availability rate of lactulose. The site specificity of drug release in canine GI tract was also estimated. Methods. CODES tablets were prepared by tableting the granulation of acetaminophen and lactulose, followed with film coating. The pharmacokinetic performance of different CODES formulations was evaluated in six beagle dogs under fasted conditions. The release of acetaminophen and lactulose was also characterized in vitro. Results. The onset of acetaminophen release in beagle dogs was found to be dependent on the coating level of Eudragit E and lactulose loading in the core tablet. At Eudragit E coating levels of 4%, 8%, and 12% (coating weight gain), the onset of in vivo drug release occurred 5.5 (±1.9) h, 4.8 (±1.0) h. and 7.5 (±1.0) h, respectively, after dosing. A similar trend was observed when the loading of lactulose in the core tablet decreased from 78% to 58% and 38%. However, the rate and extent of acetaminophen absorption did not vary significantly in each situation based on the values of AUC and C_max. Conclusions. The onset of drug release in vivo from CODES tablets is predominantly dependent on colonic availability rate of lactulose because drug release from this system is triggered by localized drop of colonic pH from the fermentation of lactulose.     

Protein Inhalation Powders: Spray Drying vs Spray Freeze Drying

Purpose. To develop a new technique, spray freeze drying, for preparing protein aerosol powders. Also, to compare the spray freeze-dried powders with spray-dried powders in terms of physical properties and aerosol performance. Methods. Protein powders were characterized using particle size analysis, thermogravimetric analysis, scanning electron microscopy, X-ray powder diffractometry, and specific surface area measurement. Aerosol performance of the powders was evaluated after blending with lactose carriers using a multi-stage liquid impinger or an Anderson cascade impactor. Two recombinant therapeutic proteins currently used for treating respiratory tract-related diseases, deoxyribonuclase (rhDNase) and anti-IgE monoclonal antibody (anti-IgE MAb), were employed and formulated with different carbohydrate excipients. Results. Through the same atomization but the different drying process, spray drying (SD) produced small (3 m), dense particles, but SFD resulted in large (8–10 m), porous particles. The fine particle fraction (FPF) of the spray freeze-dried powder was significantly better than that of the spray-dried powder, attributed to better aerodynamic properties. Powders collected from different stages of the cascade impactor were characterized, which confirmed the concept of aerodynamic particle size. Protein formulation played a major role in affecting the powder's aerosol performance, especially for the carbohydrate excipient of a high crystallization tendency. Conclusions. Spray freeze drying, as opposed to spray drying, produced protein particles with light and porous characteristics, which offered powders with superior aerosol performance due to favorable aerodynamic properties.     

Formation of Multilayers in the Caco-2 Cell Culture Model: A Confocal Laser Scanning Microscopy Study

Purpose. To introduce confocal laser scanning microscopy (CLSM)combined with digital image restoration to characterise Caco-2 cellsunder different culture conditions, and thus to define additional validcriteria for the optimisation of culture models. Methods. Growth curves were established and transepithelial electricalresistance (TEER) measured for cells grown in EMEM or DMEMmedium on Cyclopore membranes. Cytoskeleton, cell nuclei and tightjunctions (TJ) were investigated by CLSM. Results. Cultures reached a plateau of 4.5 × 10⁵ cells/cm² after 10 days. At the same time TEER reached 750 cm². An irregular,fairly complete network of TJ was present at confluence (2 d).Between 15 and 30 days a regular TJ network was established. Cellsformed mixed mono- and multilayers under most conditions with twoexceptions: flat monolayers were observed on polycarbonate filterswith EMEM and with the Biocoat intestinal epithelium differentiationenvironment system. In multilayers TJ were found in the upper aswell as in the lower cell layers although the regular vertical polaritywas disturbed. Conclusions. CLSM represents an important tool to investigate thecytoarchitecture of Caco-2 cells. 3D-analysis of confocal data givesimportant clues on the characteristics of cell layers and thus helps tovalidate optimisation strategies.     

Synergistic Effect of Formulated Plasmid and Needle-Free Injection for Genetic Vaccines

Purpose. A plasmid-based gene expression system was complexed with protective, interactive, and non-condensing (PINC) polymer system and administered with Medi-Jector, a needle-free injection device (NFID), to achieve high and sustained levels of antigen-specific antibodies in blood circulation. Methods. Human growth hormone (hGH) or bacterial -galactosidase gene expression plasmids driven by a cytomegalovirus (CMV) promoter were formulated in saline or complexed with a PINC polymer, polyvinylpyrrolidone (PVP), and intramuscularly or subcutaneously administered into dogs and pigs using a 22-gauge needle or a NFID. The hGH-specific IgG titers in serum were measured by an ELISA. -galactosidase expression was measured in injected muscles by an enzymatic assay or immunohistochemistry. The effect of NFID on DNA stability and topology was assessed by gel electrophoresis. Results. Intramuscular (i.m.) or subcutaneous (s.c.) injection of a hGH expression plasmid pCMV-hGH (0.05-0.5 mg/kg) in dogs and pigs elicited antigen-specific IgG antibody titers to expressed hGH. With both routes of injection, pDNA delivery by a NFID was superior to pDNA injection by needle. The magnitude of hGH-specific IgG titers with NFID was 15–20-fold higher than needle injection when pDNA was complexed with PVP, and only 3–4-fold higher with pDNA in saline. The transfection efficiency in the injected muscle, as measured by -galaclosidase expression, following i.m. injection of pCMV--galaclosidase/PVP, was not significantly different between needle and NFID-injected groups. Conclusions. These data demonstrate that the combination of pDNA/ PVP complexes and a NFID act synergistically to achieve high and sustained levels of antigen-specific IgG response to expressed antigen. This gene delivery approach may offer advantage over needle injection of naked DNA for the development of genetic vaccines.     

Spray Dried Powders and Powder Blends of Recombinant Human Deoxyribonuclease (rhDNase) for Aerosol Delivery

Purpose. We have used rhDNase to investigate the feasibility of developing a dry protein powder aerosol for inhalation delivery. Methods. Powders of rhDNase alone and with sodium chloride were prepared by spray drying. Powder blends were obtained by mixing (tumbling and sieving) pure rhDNase powder with 'carrier' materials (lactose, mannitol or sodium chloride). The weight percent of drug in the blends was between 5 and 70%. The particle size distributions and crystallinity of the spray dried powders were obtained by laser diffraction and X-ray powder diffraction, respectively. Particle morphology was examined by scanning electron microscopy. The ability of the powders and powder blends to be dispersed into respirable aerosols was measured using a Rotahaler connected to a multistage liquid impinger operating at 60 L/min. Results. Pure rhDNase powder was quite cohesive with a fine particle fraction (FPF or 'respirable fraction': % wt. of particles < 7 m in the aerosol cloud) of about 20%. When particles also contained NaCl, the powders were dispersed better to form aerosols. A linear relationship was observed between the NaCl content and FPF for a similar primary size (~3 m volume median diameter) of particles. The particle morphology of these powders varied systematically with the salt content. For the blends, SEM revealed a monolayer-like adhesion of the fine drug particles to the carriers at drug contents 50 % wt. An overall 2-fold increase in FPF of rhDNase in the aerosol cloud was obtained for all the blends compared to the pure drug aerosols. Conclusions. The aerosol properties of spray dried rhDNase powders can be controlled by incorporation of a suitable excipient, such as NaCl, and its relative proportion. Coarse carriers can also enhance the performance of rhDNase dry powder aerosols.     

In Vivo Lung Deposition of Hollow Porous Particles from a Pressurized Metered Dose Inhaler

Purpose: PulmoSphere particles are specifically engineered for delivery by the pulmonary route with a hollow and porous morphology, physical diameters < 5 m, and low tap densities (circa 0.1 g.cm^-3). Deposition of PulmoSphere particles in the human respiratory tract delivered by pressurized metered dose inhaler (pMDI) was compared with deposition of a conventional micronized drug pMDI formulation. Methods: Nine healthy nonsmoking subjects (5 male, 4 female) completed a two-way crossover gamma scintigraphic study, assessing the lung and oropharyngeal depositions of albuterol sulfate, formulated as ^99mTc-radiolabeled PulmoSphere particles or micronized particles (Ventolin Evohaler^TM, GlaxoSmithKline, Ltd.) suspended in HFA-134a propellant. Results: Mean (standard deviation) lung deposition, (% ex-valve dose) was doubled for the PulmoSphere formulation compared with Evohaler pMDI (28.5 (11.3) % vs. 14.5 (8.1) %, P < 0.01), whereas oropharyngeal deposition was reduced (42.6 (9.0) % vs. 72.0 (8.0) %, P < 0.01). Both PulmoSphere and Evohaler pMDIs gave uniform deposition patterns within the lungs. Conclusions: These data provided proof of concept in vivo for the PulmoSphere technology as a method of improving targeting of drugs to the lower respiratory tract from pMDIs, and suggested that the PulmoSphere technology may also be suitable for the delivery of systemically acting molecules absorbed via the lung.     

In Vitro Gene Transfection in Human Glioma Cells Using a Novel and Less Cytotoxic Artificial Lipoprotein Delivery System

Purpose. To develop and evaluate a novel artificial lipoprotein delivery system for in vitro gene transfection in human glioma cells. Method. Nanoemulsion was formulated with similar lipid compositions present in natural lipoproteins. The oil phase of nanoemulsion was composed of triolein (70%), egg phosphatidylcholine (22.7%), lysophosphatidylcholine (2.3%), cholesterol oleate (3.0%), and cholesterol (2.0%). To replace the surface protein as in natural lipoprotein, poly-L-lysine was modified to add palmitoyl chains at a basic condition and was incorporated onto the nanoemulsion particles through hydrophobic interaction. A model plasmid DNA, pSV--Gal containing a reporter gene for -galactosidase was carried by the nanoemulsion/poly-L-lysine particles. The charge variation of so-formed complex was examined by agarose gel electrophoresis and zeta potential measurement. In vitro transfection was conducted on human SF-767 glioma cell line using this new system. After standard X-Gal staining, transfected cells were observed under light microscope. The effect of chloroquine on the transfection was examined and, finally, the cytotoxicity of this new system was evaluated in comparison with commercial Lipofectamine gene transfection system. Results. The plasmid DNA was effectively carried by this artificial lipoprotein delivery system and the reporter gene was expressed in the glioma cells. Transfection efficiency was significantly increased by the treatment of chloroquine, indicating that endocytosis possibly was the major cellular uptake pathway. Compared to Lipofectamine system, this new delivery system demonstrated similar transfection efficiency but a much lower cytotoxicity. In the experiment, the cell viability showed up to 75% using this system compared to only 24% using Lipofectamine system. Conclusion. A new artificial lipoprotein delivery system was developed for in vitro gene transfection in tumor cells. The new system showed similar transfection efficiency but a much lower cytotoxicity compared with commercial Lipofectamine system.     

DQAsomes: A Novel Potential Drug and Gene Delivery System Made from Dequalinium™

Purpose. Dequalinium, a drug known for over 30 years, is a dicationic amphiphile compound resembling bolaform electrolytes. The purpose of our work was to determine the state of aggregation of dequalinium in aqueous medium and to investigate both, its ability to bind DNA and its potential to serve as a novel non-viral transfection vector. Methods. The form of aggregation was determined employing electron microscopic techniques. The DNA binding capacity of dequalinium was assayed using SYBR Green I stain. For in vitro cell transfection experiments plasmid DNA encoding for firefly luciferase was used. Results. Dequalinium forms in aqueous medium liposome-like aggregates, which we term DQAsomes. These dequalinium vesicles bind DNA and they are able to transfect cells in vitro with an efficiency comparable to Lipofectin. Conclusions. Based on the intrinsic properties of dequalinium such as the in vivo selectivity for carcinoma cells and selective accumulation in mitochondria we propose DQAsomes as a novel and unique drug and gene delivery system.     

Aerosol Electrostatics I: Properties of Fine Powders Before and After Aerosolization by Dry Powder Inhalers

Purpose. To evaluate the dependence of fine particle dose charge (FPD charge) generated from powder inhalers on physico-chemical properties of the inhalation powder, inhaler type, deaggregation mechanism, dose number and/or retained powder. Methods. Electrostatic charges were determined on micronized powders and aerosolized fine particle doses withdrawn from two, high efficiency, multidose powder inhalers, Turbohalerand prototype Dryhaler. The behavior of terbutaline sulfate, budesonide, albuterol (sulfate and base), beclomethasone dipropionate and lactose was assessed before and after aerosolization. Results. Both inhalers conferred triboelectric FPD charges during aerosolization in the range –400 pC through +200 pC. Specific charges (charge/unit mass) on the fine particle doses of budesonide from Dryhaler were significantly less than those from Turbohaler (p < 0.01). Electrostatic charges on the potentially respirable cloud of terbutaline sulfate generated by Bricanyl Turbohaler were positive and/or negative and unpredictable. With Pulmicort Turbohaler, FPD charges on budesonide were always positive. Dryhaler was used to determine the chemical dependence of fine particle triboelectrification during the aerosolization of pure materials. A triboelectric series was constructed from the Dryhaler results ranking the powders from positive to negative as budesonide > lactose > albuterol sulfate > terbutaline sulfate albuterol beclomethasone dipropionate. Conclusions. While there was no evidence of FPD charge dependence upon dose number with either inhaler, FPD charges were dependent upon the powder under investigation, as well as the construction and deaggregation mechanism of the inhaler. The specific charge on the fine particle dose of budesonide from Turbohaler corresponded to approximately 200 electronic charges per particle, a value which is known to affect both total and regional aerosol deposition in the human lung. Electrostatic charge effects may be important determinants of aerosol behavior and should not be neglected.     

The AERX™ Aerosol Delivery System

Purpose. We describe the AER_X aerosol delivery system, a new, bolus inhalation device that is actuated at preprogrammed values of inspiratory flow rate and inhaled volume. We report on its in vitro characterization using a particular set of conditions used in pharmacokinetic and scintigraphic studies. Methods. Multiple doses of aerosol were delivered from single use collapsible plastic containers containing liquid formulation. The aerosol was generated by forcing the formulation under pressure through an array of 2.5 micron holes. Air was drawn through the device at 70 LPM, and the aerosol was collected onto a filter or Andersen cascade impactor. The emitted dose was quantified from the filter collection data, and the particle size distribution was obtained from the best fit log-normal distribution to the impactor data. Results. 57.0 ± 5.9% of the dose of drug placed as an aqueous solution in the 45 L collapsible container was delivered as an aerosol (n = 40). The best fit size distribution had an MMAD = (2.95 ± 0.06) m and a geometric standard deviation _g = 1.24 ± 0.01 (n = 6). Conclusions. The AER_X aerosol delivery system generates a nearly monodisperse aerosol with the properties required for efficient and repeatable drug delivery to the lung.     

Crystal structure of neotame anhydrate polymorph G

Purpose. To determine the crystal structure of the neotame anhydrate polymorph G and to evaluate X-ray powder diffractometry (XRPD) with molecular modeling as an alternative method for determining the crystal structure of this conformationally flexible dipeptide. Methods. The crystal structure of polymorph G was determined by single crystal X-ray crystallography (SCXRD) and also from the X-ray powder diffraction (XRPD) pattern using molecular modeling (Cerius² , Powder Solve module). Results. From SCXRD, polymorph G crystals are orthorhombic with space group of P2₁2₁2₁ with Z = 4, unit cell constants: a = 5.5999(4), b = 11.8921(8), c = 30.917(2) Å, and one neotame molecule per asymmetric unit. The XRPD pattern of polymorph G, analyzed by Cerius² software, led to the same P2₁2₁2₁ space group and almost identical unit cell dimensions. However, with 13 rigid bodies defined, Cerius² gives a conformation of the neotame molecule, which is different from that determined by SCXRD. Conclusions. For neotame anhydrate polymorph G, the unit cell dimensions calculated from XRPD were almost identical to those determined by SCXRD. However, the crystal structure determined by XRPD closely resembled that determined by SCXRD, only when the correct conformation of the neotame molecule had been chosen before detailed analysis of the XRPD pattern.     

Evaluation of Genetic Stability of Recombinant Human Factor VIII by Peptide Mapping and On-line Mass Spectrometric Analysis

Purpose. The genetic stability of a recombinant human factor VIII (rhFVIII) product expressed in Chinese hamster ovary cells (Recombinate) has been evaluated through comparisons of the protein produced at the beginning, middle and end of a typical production campaign. Methods. Recombinant human factor VIII was incubated with thrombin, the resulting four polypeptides were isolated by RP-HPLC, subjected to proteolysis with trypsin, and the peptide mixtures were resolved by RP-HPLC. Tryptic peptide mixtures were subjected to online mass spectrometric analysis using an electrospray ionization source interfaced to a quadrupole mass analyzer scanning from 1950–200 amu, and the peptide ion data were compared for three lots produced from the beginning, middle and end of a production campaign. Results. The UV elution profiles for each of the rhFVIIIa polypeptides were highly similar for factor VIII isolated from the beginning, middle and end of production. Total ion data from the peptide maps derived from three lots of rhFVIII were compared by MH¹⁺ values as a function of scan range. A total of 918 ions were analyzed for the four polypeptides of rhFVIII produced at the beginning, middle and end of a production campaign. The ions were detected at the same relative retention times, as indicated by the similar scan numbers for the three lots. Conclusions. These observations support that rhFVIII preparations produced from the beginning, middle and end of a production campaign were highly similar, and demonstrate genetic stability in the manufacturing process of Recombinate.     

Size Characterization of Mycobacterium bovis BCG (Bacillus Calmette Guérin) Vaccine,Tice™ Substrain

Reconstituted, lyophilized, attenuated Mycobacterium bovis, Bacillus Calmette Guérin (BCG) vaccine, Tice substrain, was characterized using a Coulter Multisizer and a HIAC/Royco counter. The primary organism has an equivalent spherical diameter approximating 1 µm but the BCG cell suspension is heavily aggregated. The cumulative size distribution of the suspension fits a log-probit plot and this information can be used to determine the total number of particles per ampoule. The instrumental count may be related to the viable count. The state of dispersion was unaffected by mild shear (syringe aspiration or ultrasound) and only slightly affected by the addition of cetylpyridinium chloride or sodium tauroglycolate.     

Nuclear Transport of Oligonucleotides in HepG2-cells Mediated by Protamine Sulfate and Negatively Charged Liposomes

Purpose. The aim of this study was to characterize the intracellular fate and nuclear uptake kinetics of oligonucleotides (ON) that were complexed with protamine sulfate (PS) and negatively charged liposomes at different ratios of ON to PS. Methods. Double-fluorescence labelling of ON and liposomal lipid was applied to simultaneously monitor the interaction as well as the individual fate of active agent and carrier upon intracellular delivery using confocal laser scanning microscopy (CLSM). A DNA-analogue of a 68-mer intramolecular double-stranded RNA:DNA-hybridoligo- nucleotide (chimeraplasts) with unmodified phosphate backbone was employed. This construct was condensed with PS and coated with a liposomal formulation (AVE-3 = artificial viral envelope). Results. PS-ON complexes and AVE-3-coated complexes with a defined composition were very effective in nuclear transport of ON for a ON:PS charge ratio of 1:3. Nucleus:cytosol fluorescence ratios peaked at about 10 hrs and started to decrease again at 21 hrs. Conclusions. AVE associates with PS-condensed ON, and this complex is able to be taken up by cells and to deliver ON to the nucleus. PS-ON complexes are released from the liposomal formulation, mainly as an extranuclear enzymatic degradation of the liposomal phospholipids. The results of the kinetic analysis can be used to optimize transfection protocols with ON in HepG2 cells.     

Influence of Suspension Stabilisers on the Delivery of Protein-Loaded Porous Poly (DL-Lactide-co-Glycolide) (PLGA) Microparticles via Pressurised Metered Dose Inhaler (pMDI)

Purpose

This work investigates the feasibility of delivering large (≈ 25 μm) porous poly (lactide-co-glycolide) (PLGA) microparticles containing a model protein via pressurised metered dose inhaler (pMDI).

Methods

Porous PLGA microparticles were prepared by modified double emulsion method as pMDI suspension based systems containing suspension stabilisers in 1,1,1,2,3,3,3-heptafluoropropane (HFA 227). Physical suspension stability was assessed by visual and optical suspension techniques. Aerosolisation characteristics were investigated using aerosol particle sizing, dose delivery through the valve (DTV) and shot weight.

Results

An optimum concentration of suspensions stabiliser was required to achieve physical pMDI suspension stability; values of; 0.0075%w/w PVP K30 or 0.075%w/w PEG 300 were required. Formulations that exhibited good physical stability also showed optimum aerosolisation characteristics. When employing 0.0075% PVP K30 DTV at the start and end of can life was 98.11(±10.01) % and 75.06 (±7.01) % respectively verses values of 37.39 (±11.12) % and 5.57 (±1.72) % without the inclusion of PVP K30.

Conclusion

Porous PLGA microparticles show potential as macromolecule/protein carrier and also to target lower regions of the lungs when prepared as pMDI suspension formulations in HFA 227 using suspension stabilisers to achieve consistent dose delivery through the life of the pMDI, however, inter-relationship between the device and the formulation need to be considered to achieve suitable respiratory delivery.     

Stability of nystatin in mouthrinses; effect of pH, temperature, concentration and colloidal silver addition, studied using an in vitro antifungal activity.

Alkaline low concentration nystatin mouthrinses extemporanely prepared can be used to treat oropharyngeal candidiasis in immunodeficient patients. However, their expiration dates are not distinctly determined.The stability of nystatin, added (as Mycostatine) at a concentration of 14 400 U/ml in 10^–4 N hydrochloric acid, purified water and 1.4% injectable sodium hydrogen carbonate with or without 0.002% colloidal silver (an antiseptic agent added because of its known antifungal potency) was studied after storage in tinted glass bottles at 5 °C and 22 °C over 11 days, and compared with reconstituted 100 000 U/ml aqueous Mycostatine oral suspension. At 2, 4, 7, 9, and 11 days after preparation. Samples were tested for pH, microbial contamination, and assayed by an in vitro microbiological test.Neither significant variation of pH nor microbial contamination were in evidence. Nystatin 14 400 U/ml maintained at least 90% of its initial concentration for 4 days in acid at both temperatures, for 7 days (5 °C) and 4 days (22 °C) in aqueous and alkaline environments, for 9 days (5 °C) and 7 days (22 °C) in 1.4% injectable sodium hydrogen carbonate containing colloidal silver which showed an antifungal potency. The 100 000 U/ml aqueous Mycostatine oral suspension was stable for 9 days and 4 days at 5 °C and 22 °C respectively.An ambulant patient can keep a low concentration alkaline antifungal mouthrinse at home for a week at 5 °C.     

Reduced UV-Induced Degradation of Doxorubicin Encapsulated in Polyethyleneglycol-Coated Liposomes

Purpose. The aim of this study was to investigate the stability of doxorubicin encapsulated in polyethyleneglycol-coated liposomes (Doxil) under UV-A light. Methods. High performance liquid chromatography and a fluorimetric method were used to quantify doxorubicin in bulk solution and doxorubicin in Doxil formulation. Results. The photodegradation of Doxil was significantly lower in comparison to the photodegradation of the free drug and showed no concentration dependency at the measured concentration range of 5–50 g/ml. During and after UV-A irradiation, there was no leakage of the drug from liposomes to the medium. After induced leakage of doxorubicin from the liposomes by the ionophore nigericin, the degradation kinetics of Doxil were identical to that of free doxorubicin. Conclusions. High intraliposomal doxorubicin concentration and intraliposomal acidic pH are the two critical factors that protect DXR in Doxil from UV-A degradation.     

Evaluation of a filling system for binary pediatric solutions.

The aim of this study was to assess the use of an automatic filling system (Siframix M31 and M32 system) to prepare pediatric parenteral nutrition. Volumetric accuracy was measured for each siframix system loads cells (< 5% for 5 ml with the Siframix M32 and < 5% for 9 ml with the Siframix M31) with sterile water for injection. The minimal 20 ml of flushing sterile water of the common tubulure of the Siframix M32 (p=0.211 for 20 ml and p=0.75 for 500 ml), the use of viscous solutions (70 % dextrose) on the Siframix M31 (p=0.28 for 20 ml and p=0.12 for 500 ml) and the use of a special tubulure for using E.V.A. Luerlock bags (p=0.89 for 20 ml and p=0.103 for 500 ml) do not modify the accuracy. Changing bags or bottles during the filling operation modify the accuracy (p=0.004 for 20 ml and p=0.009 for 500 ml). A flushing operation is necessary to lower the risk of electrolytic pollution for the filling of little bags. The filling speed for each module was also measured (the maximal filling speed was five liters per minute). The Siframix system allows one to prepare pediatric parenteral nutrition bags when volumes are above 4 ml and with adapted source solutions in terms of concentration and conditioning volumes.