Formation of Multilayered Vesicles from Water/Organic-Solvent (W/O) Emulsions: Theory and Practice

Multilayered liposome (MLV) formation from water/organic-solvent (W/O) emulsions was studied. A fundamental liposome population parameter, the E^ratio, was defined and used to estimate the bilayer number and water spacing in MLV liposomes. MLVs prepared from W/O emulsions have optimum drug entrapment at an emulsion-lipid/emulsion-water ratio of 0.33. Drug entrapment is typically 50 to 65% under these optimal conditions.     

Predicting the Effect of Nonionic Surfactants on Dispersed Droplet Radii in Submicron Oil-in-Water Emulsions

A novel theoretical model which describes the mass mean radius of oil droplets in an oil-in-water emulsion is described. A modified form of the Langmuir adsorption isotherm is used to account for nonlinear adsorption of surfactant to an oil-water film and its effect on interfacial tension and oil droplet radius. On the basis of this model, the mass mean oil droplet radius may be related to bulk surfactant concentration for a nonionic surfactant. An analysis of the mineral oil-Triton X405-water system shows that aqueous solutions of Triton X405 against mineral oil behaved in accord with the proposed model. _m, the maximum interfacial tension lowering by Triton X405, was estimated to be 20.9 dyne/cm. A surfactant specific apparent constant (B) which relates the rate constant for adsorption to the interface to the rate constant for desorption from the interface into the aqueous bulk was estimated as 5.44E4 cm³/g. The pressure across the curved interface, P, was estimated as 1.05E6 dyne/cm². The theoretical model appears to be consistent with experimentally observed oil droplet radii and is considered to be an accurate representation of the mechanics of dispersed droplet radii under conditions of moderate load of nonionic surfactant.     

The single dose kinetics of chloroquine and its major metabolite desethylchloroquine in healthy subjects

Summary The kinetics and disposition of chloroquine (CQ) and its metabolite monodesethylchloroquine (CQM) were investigated in 5 healthy volunteers after incremental (150–300–600 mg CQ base) single oral doses of CQ. The analytical method used (HPLC and fluorescence detection) is the most sensitive known method for CQ and CQM. Plasma and whole blood concentrations of CQ, CQM and a third metabolite, bidesethylchloroquine (CQMM), were determined. The kinetics of CQ was found to be unique. The best fit was obtained by a multicompartmental model. The biological half-life appeared to be between 30–60 days; the volume of distribution (V_d) was 800l/kg, and the clearance 11/h/kg when calculated from plasma data. The whole blood concentrations were 8–10 times higher than in plasma, and consequently the V_d and whole blood clearance were 10 times lower. The kinetics changed as the dose was increased. An indication of capacity-limited steps in CQ disposition was found, as the rate constants decreased even though the clearance remained the same. The intrinsic half-life of CQM was 1/4 of that of CQ, but was prolonged after the highest dose of CQ. The present knowledge of CQ kinetics could provide a basis for revision of current dosage regimens in malaria suppression and rheumatoid disease to ensure efficacious and safe therapy.     

PLG Microsphere Size Controls Drug Release Rate Through Several Competing Factors

Purpose. Although the rate of drug release from poly(D,L-lactide-co-glycolide) (PLG) microspheres is often modulated by changing fabrication conditions or materials, the specific factors directly controlling the release profiles are often unclear. We have fabricated uniform rhodamine- and piroxicam-containing microspheres, 10 to 100 m in diameter, to better understand how microsphere size controls drug release. Methods. Drug distribution within the microspheres was examined using confocal fluorescence microscopy. The rate of polymer degradation was determined as the change in molecular weight, measured by gel permeation chromatography, during in vitro degradation experiments. Further, changes in the surface and interior morphology of the particles during in vitro degradation were investigated by scanning electron microscopy. Results. Microsphere size greatly affected drug distribution. Small (10-m) microspheres showed an essentially uniform drug distribution. Larger (100-m) microspheres showed redistribution of drug to specific regions of the microspheres. Rhodamine partitioned to the surface and piroxicam partitioned to the interior of large PLG microspheres. Further, the rate of polymer degradation increased with microsphere size, possibly the result of a more acidic interior caused by increased accumulation of hydrolyzed polymer products in larger particles. Finally, larger microspheres developed a more porous interior structure during the drug release. Conclusions. Microsphere size affects drug release not only through changes in diffusion rates but also through secondary effects including drug distribution in the particle, polymer degradation rate, and microsphere erosion rates.     

Einfluß von Pharmaka auf energieliefernde Reaktionen des Stoffwechsels

Ohne ZusammenfassungAbkürzungen NAD = DPN Nicotinamid-Adenin-Dinucleotid - ATP Adenosintriphosphat - 2,4-DNP 2,4-Dinitrophenol - P₁ anorganisches Phosphat; bezeichnet eine energiereiche Bindung - FP Flavoproteid Mit 8 Textabbildungen.     

Electrorepulsion Versus Electroosmosis: Effect of pH on the lontophoretic Flux of 5-Fluorouracil

Purpose. To delineate the contributions of electrorepulsion and electroosmosis to the iontophoretic flux of 5-FU across porcine skin in vitro. Also, the isoelectric point (pI) of the skin model was determined. Methods. The electrotransport of 5-FU, anode-to-cathode ('anodal') and cathode-to-anode ('cathodal') was determined as a function of the pH of the electrolyte bathing the skin. Results. At pH 8.5, the drug (pK_a 8) is negatively charged and 'cathodal', viz. electrorepulsive, transport is much greater than that in the opposite direction. At pH 7.4, where 25% of 5-FU is charged, electrorepulsive and electroosmotic ('anodal') fluxes are balanced. Decreasing the pH to 6, and then 5, reduces the percentage of ionized 5-FU such that 'anodal' electroosmosis dominates across the negatively-charged membrane. But, at pH 4, 'anodal' and 'cathodal' fluxes are again equal suggesting neutralization of the skin (i.e., pI 4). This is confirmed at pH 3, where 'cathodal' electroosmosis dominates across the now net-positively charged barrier. Conclusions. Electrotransport is sensitive, mechanistically, to the properties of the permeant and of the skin; interactions of, for example, the drug or constituents of a formulation, that alter the barrier's net charge, can affect iontophoretic delivery. The pI of porcine ear skin is 4.     

Microdialysis Evaluation of the Ocular Pharmacokinetics of Propranolol in the Conscious Rabbit

Purpose. This study was conducted to assess the effects of anesthesia and aqueous humor protein concentrations on ocular disposition of propranolol. Methods. Rabbits were anesthetized and a microdialysis probe was inserted into the anterior chamber of one eye; the contralateral eye served as a control. At timed intervals after probe placement, a 100-l sample of aqueous humor was aspirated from each eye to determine protein concentration. In vitro protein binding parameters were used to simulate the impact of protein concentration on propranolol disposition. To assess the influence of anesthesia, probes were implanted in the anterior chamber of each eye. After >5-day stabilization, conscious and anesthetized rabbits (n = 3/group) received a 200-g topical dose of [³H] DL-propranolol in each eye; propranolol was assayed in probe effluent. Results. Changes in aqueous humor protein concentrations were observed following probe insertion. Simulations demonstrated that the unbound propranolol AUC (2.4-fold) in aqueous humor should be reduced due to protein influx. Intraocular propranolol exposure in anesthetized rabbits was 8-fold higher than in conscious rabbits, and 1.9-fold higher than in rabbits without a post-surgical recovery period. Conclusions. Anesthesia and time-dependent aqueous humor protein concentrations may alter ocular pharmacokinetics, and must be taken into account in the design of microdialysis experiments.     

Phase Transitions of Glycine in Frozen Aqueous Solutions and During Freeze-Drying

Purpose. To study the solid-state and phase transitions of glycine, (i) in frozen aqueous solutions, and (ii) during freeze-drying. Methods. X-ray powder diffractometry (XRD) and differential scanning calorimetry (DSC) were used to analyze the frozen systems. In situ freeze-drying in the sample chamber of the diffractometer enabled characterization of phase transitions during freeze-drying. Results. Transitions in frozen systems. Rapid (20°C/min) or slow (2°C/min) cooling of aqueous solutions of glycine (15% w/w) to –70°C resulted in crystallization of -glycine. Annealing at –10°C led to an increase in the amount of the crystalline phase. When quench-cooled by immersing in liquid nitrogen, glycine formed an amorphous freeze-concentrate. On heating, crystallization of an unidentified phase of glycine occurred at \-65°C which disappeared at –55°C, and the peaks of -glycine appeared. Annealing caused a transition of - to the - form. The extent of this conversion was a function of the annealing temperature. Slower cooling rates and annealing in frozen solutions increased the crystalline -glycine content in the lyophile. Freeze-drying of quench-cooled solutions led to the formation of -glycine during primary drying resulting in a lyophile consisting of a mixture of - and -glycine. The primary drying temperature as well as the initial solute concentration significantly influenced the solid-state of freeze-dried glycine only in quench-cooled systems. Conclusions. The cooling rate, annealing conditions and the primary drying temperature influenced the solid-state composition of freeze-dried glycine.     

Effect of Surfactant Phase in Perfluorocarbon Emulsification Efficiency

The effect of preparation temperature on the emulsification efficiency of perfluoro-3-butyltetrahydrofuran (FC-75) was investigated. Polyoxyethylene (POE) oleyl ether surfactants were used as the emulsifier(s) in a range of HLB values of 7.5 to 9.5. The emulsions were prepared by paddle mixing as a method of low-shear emulsification. After centrifugation of the resulting O/W emulsions, the volume of FC-75 which separated was utilized as a measure of the emulsification efficiency. In general, emulsions prepared at temperatures where the surfactant was in a lamellar-to-isotropic surfactant solution transition, L L₃, displayed a better emulsification efficiency than those prepared with other surfactant phases.     

Optimized Inhalation Aerosols. I. The Effects of Spherical Baffle Size and Position upon the Output of Several Pressurized Nonaqueous Suspension Formulations

Baffles contained in conventional actuators may be a convenient alternative to some of the extension devices used presently with metered-dose inhalers (MDIs). Actuators were modified to determine whether baffles could be used to decrease the output of large nonrespirable droplets. These actuators were tested using a series of nonaqueous suspension aerosols containing 0.1 to 2.0% micronized disodium fluorescein (DF) as the model drug, stabilized by sorbitan trioleate in a constant blend of fluorocarbons 11, 12, and 114. A 25-µl metering volume was used throughout. Aerosol output was characterized by cascade impaction. Baffle size and position had pronounced effects on actuator retention and aerosol output. Increasing baffle size resulted in increased retention in the actuator. The total output of the MDI in the respirable range (aerodynamic diameter, D _ae, <5.5 µm) was greater in the unbaffled actuator than in all baffled actuators. However, all baffles increased the respirable fraction (DF with D _ae <5.5 µm: total DF leaving the actuator), R, when compared to their unbaffled controls. For example, for a 0.1% DF, 0.14% surfactant formulation, R was increased from 0.40 (unbaffled) to 0.71 by incorporation of a 0.6-cm-diameter sphere 1.3 cm from the jet of the actuator. In these cases, aerosol segregation occurred due to droplet inertia in the high velocity gas flows. Increasing the respirable fraction at the expense of the total respirable output may obviate undesirable clinical effects.     

Hepatic clearance of local anesthetics in man

Two independent methods of calculating hepatic drug clearance were applied to data from studies of the human pharmacokinetics of lidocaine, bupivacaine, and etidocaine. Within experimental limitations, agreement was good between estimates obtained by measurement of areas under blood drug concentration-time curves after rapid intravenous injection and by direct measurement of arterial and hepatic venous drug concentrations. Apparent hepatic extraction ratios of the agents followed the order etidocaine (0.73)/s>lidocaine (0.68>bupivacaine (0.37).Pharmacokinetic implications of increases in hepatic blood flow induced by the agents are discussed.     

Solid-State Emulsions: The Effects of Process and Storage Conditions

The effects of process and storage conditions of solid-state emulsions were studied. Oil-in-water emulsions may be prepared from solid state emulsions by adding an aqueous phase to the solid. Solid-state emulsions are prepared by processing an oil phase and an aqueous solution of matrix material via a solvent removal process. Sucrose, the carrier material utilized in this report, results in a metastable solid or glass, which can transform upon aging to a more stable thermodynamic state. Aging was determined by monitoring the crystallinity as a function of time, temperature, relative humidity, and grinding. The crystallinity of solid-state emulsions was determined with X-ray diffraction and differential scanning calorimetry. Results indicate that solid-state emulsions should be stored between 15 and 25% relative humidity at 25°C. Grinding has no apparent effect on the crystallinity of the sample, as detected by X-ray diffraction, although the microcrystallinity is increased. The utilization of silinized glassware enabled the sample-to-sample microcrystalline variability to be reduced.     

Intracranial Delivery of Recombinant Nerve Growth Factor: Release Kinetics and Protein Distribution for Three Delivery Systems

Purpose. Three different polymeric delivery systems, composed of either poly(ethylene-co-vinyl acetate) (EVAc) or poly(lactide-co-gly-colide) (PLGA), were used to administer recombinant human nerve growth factor (rhNGF) intracranially in rats. Methods. The delivery systems were characterized with respect to release kinetics, both in the brain and in well-stirred buffer solutions. Results. During incubation in buffered saline, the delivery systems released rhNGF in distinct patterns: sustained (EVAc), immediate (PLGA₁), and delayed (PLGA₂). One 10-mg delivery system was implanted in each rat and an ELISA technique was used to determine the amount of rhNGF in 1-mm coronal brain slices produced immediately after removal of the delivery system. High levels of rhNGF (as high as 60,000 ng in a brain slice of 50 L) were recovered from the brain tissue at 1,2, and 4 weeks after implantation. With all three delivery systems, the amount of rhNGF in each brain slice decreased exponentially with distance from the implant site; the distance over which concentration decreased by 10-fold was 2–3 mm for all delivery systems. When rhNGF release was moderate (10 to 200 ng rhNGF/ day), the total amount of rhNGF in the brain increased linearly with release rate, suggesting an overall rate of rhNGF elimination of 0.4 hr^–1 or a half-life of 1.7 hr. With higher release rates (500 to 50,000 ng rhNGF/day), total amounts of rhNGF in the brain were considerably higher than anticipated based on this rate of elimination. Conclusions. Polymeric controlled release can provide high, localized doses of rhNGF in the brain. All of the experimental data were consistent with penetration of rhNGF through the brain tissue with a diffusion coefficient 8 X 10^–7 cm²/s, which is 50% of the diffusion coefficient in water.     

Submicron Cationic Emulsions as a New Delivery System for Oligonucleotides

Purpose. The main purpose of the present study was to investigate submicron emulsions as potential oligonucleotide (ON) delivery system. Methods. Submicron emulsions containing various concentrations of stearylamine (SA) were prepared by microfluidization. After association with model oligothymidylates, these emulsions were characterized in terms of particle size, -potential, association efficiency and release upon dilution. The interactions between ON and SA were investigated by partitioning studies between water and oily phases, with ON of three different lengths (pdT₁₆, pdT₃₀, pdT₅₀). The stability of pdT₁₆ in the presence of nucleases was evaluated by incubation in cell culture medium supplemented with 10% of foetal calf serum. Results. The ON association efficiency was much higher with emulsions containing SA (E_SA) than with control emulsions (E₀), whatever the ON length. In addition, E_SA was shown to protect ON against degradation for up to 3 hours in culture medium. ON and SA were able to form ion-pairs and the resulting complex was found to be insoluble both in water and in oil. Zeta potential was maintained constant when increasing the ON concentration, until flocculation occurred (up to 250M in the case of pdT₁₆ for example). This has been explained by the presence of SA in excess, soluble in the oily core of droplets, able to migrate towards the interface and replacing SA neutralized in ion-pairs. Conclusions. E_SA appears to be a valuable system for delivery of ON and might even be improved by selecting an oily phase in which the SA/ON complex would be soluble.Affiliated with the David R. Bloom Center for Pharmacy at the Hebrew University of Jerusalem.     

Liquid Chromatographic Analysis of Di(2-ethylhexyl) Phthalate: Application to Pharmacokinetic Studies in the Mongrel Dog

A high-performance liquid chromatographic (HPLC) assay was developed for the determination of di(2-ethylhexyl) phthalate (DEHP) in serum or plasma. Plasma DEHP concentrations that were measured by HPLC in specimens obtained from hemodialysis patients were in good agreement with corresponding concentrations that were measured by gas chromatography with selected ion monitoring (GC-SIM) (r ² = 0.996). Plasma DEHP concentrations were measured after intravenous DEHP administration (1.2–4.4 mg DEHP/kg body weight) to determine the effect of bilateral ureteral ligation on DEHP elimination in the mongrel dog. DEHP plasma clearance (6.3 ml/min/kg), steady-state distribution volume (0.2l/kg), and terminal half-life (50 min) were unchanged in two dogs following bilateral ureteral ligation. DEHP terminal half-life and steady-state distribution volume were substantially smaller (25- to 70-fold) than reported previously in the rat or dog.     

The Mechanism of Uptake of Biodegradable Microparticles in Caco-2 Cells Is Size Dependent

Purpose. To study the uptake of biodegradable microparticles in Caco-2 cells. Methods. Biodegradable microparticles of polylactic polyglycolic acid co-polymer (PLGA 50:50) of mean diameters 0.1 m, 1 m, and 10 m containing bovine serum albumin as a model protein and 6-coumarin as a fluorescent marker were formulated by a multiple emulsion technique. The Caco-2 cell monolayers were incubated with each diameter microparticles (100 g/ml) for two hours. The microparticle uptake in Caco-2 cells was studied by confocal microscopy and also by quantitating the 6-coumarin content of the microparticles taken up by the cells. The effects of microparticle concentration, and incubation time and temperature on microparticle cell uptake were also studied. Results. The study demonstrated that the Caco-2 cell microparticle uptake significantly depends upon the microparticle diameter. The 0.1 m diameter microparticles had 2.5 fold greater uptake on the weight basis than the 1 m and 6 fold greater than the 10 m diameter microparticles. Similarly in terms of number the uptake of 0.1 m diameter microparticles was 2.7 × 10³ fold greater than the 1 m and 6.7 × 10⁶ greater than the 10 m diameter microparticles. The efficiency of uptake of 0.1 m diameter microparticles at 100 g/ml concentration was 41% compared to 15% and 6% for the 1 m and the 10 m diameter microparticles, respectively. The Caco-2 cell microparticle (0.1 m) uptake increased with concentration in the range of 100 g/ml to 500 g/ml which then reached a plateau at higher concentration. The uptake of microparticles increased with incubation time, reaching a steady state at two hours. The uptake was greater at an incubation temperature of 37°C compared to at 4°C. Conclusions. The Caco-2 cell microparticle uptake was microparticle diameter, concentration, and incubation time and temperature dependent. The small diameter microparticles (0.1 m) had significantly greater uptake compared to larger diameter microparticles. The results thus suggest that the mechanism of uptake of microparticles in Caco-2 cell is particle diameter dependent. Caco-2 cells are used as an in vitro model for gastrointestinal uptake, and therefore the results obtained in these studies could be of significant importance in optimizing the microparticle-based oral drug delivery systems.     

Preparation of Uniform-Sized Multiple Emulsions and Micro/Nano Particulates for Drug Delivery by Membrane Emulsification

Much attention has in recent years been paid to fine applications of drug delivery systems, such as multiple emulsions, micro/nano solid lipid and polymer particles (spheres or capsules). Precise control of particle size and size distribution is especially important in such fine applications. Membrane emulsification can be used to prepare uniform-sized multiple emulsions and micro/nano particulates for drug delivery. It is a promising technique because of the better control of size and size distribution, the mildness of the process, the low energy consumption, easy operation and simple equipment, and amendable for large scale production. This review describes the state of the art of membrane emulsification in the preparation of monodisperse multiple emulsions and micro/nano particulates for drug delivery in recent years. The principles, influence of process parameters, advantages and disadvantages, and applications in preparing different types of drug delivery systems are reviewed. It can be concluded that the membrane emulsification technique in preparing emulsion/particulate products for drug delivery will further expand in the near future in conjunction with more basic investigations on this technique.     

Solid-State Emulsions: Evaluation by 1H and 13C Solid-State Nuclear Magnetic Resonance

The molecular environment of sucrose and mineral oil within sucrose and mineral oil solid state emulsions was investigated by NMR techniques. The ¹³C and ¹H chemical shifts of sucrose and mineral oil to those observed in solid state emulsions (comprised of sucrose and mineral oil) were equivalent, indicating that the local structure of sucrose is unaffected by the presence of mineral oil in the solid-state emulsion. Cross-polarization, magic angle spinning ¹³C (CP-MAS) in conjuncton with single-pulse studies indicated that the ¹H-¹³C dipole-dipole interactions are very weak, i.e., mineral oil is highly mobile. Spinning side bands were observed, however, in ¹H single-pulse, magic angle spinning (SPMAS) spectra of the solid-state emulsion, indicating that the mineral oil has solid properties. Although the mineral oil was shown to be highly mobile, it also appears to be constrained or included by the sucrose.     

Blockade of spatial learning by the M1 muscarinic antagonist pirenzepine

Two experiments were conducted to determine the effects of the M₁ muscarinic receptor antagonist pirenzepine on place navigation in a water maze. In the first experiment rats were required to learn the location of a hidden platform following intracerebroventricular injections of equimolar doses of pirenzepine or scopolamine methylbromide. Both drugs dose-dependently impaired spatial learning according to both escape latency data and transfer test analysis. Pirenzepine was approximately 3 times less potent than scopolamine, a potency ratio which suggests M₁ receptor mediation of the impairment. In the second experiment pirenzepine (192.3 g/rat ICV) was injected prior to training on a simultaneous place dicrimination task in the water maze. Impairments of choice accuracy were found with a dose of 20 g/rat in the absence of any marked increases in either errors of omission or choice latency. These data suggest that M₁ receptor blockade impairs processes which are involved in spatial learning.     

Distribution of Gacyclidine Enantiomers in Spinal Cord Extracellular Fluid

Purpose. Determination of the pharmacokinetics of gacyclidineenantiomers, a non-competitive NMDA antagonist, in plasma and spinal cordextracellular fluid (ECF) of rats. Methods. Implantation of microdialysis probes in spinal cord (T9).Serial collection of plasma samples and ECF dialysates over 5 hoursafter IV bolus administration of (±)-gacyclidine (2.5 mg/kg). Plasmaprotein binding determined in vivo by equilibrium dialysis. ChiralGC/MS assay. Results. Plasma concentrations of (+)-gacyclidine were 25% higherthan those of (–)-gacyclidine over the duration of the experiment inall animals. Plasma concentrations decayed in parallel in a biphasicmanner (t_1/2 9 min; t_1/2 90 min) with no significant differencebetween enantiomers. Clearance and volume of distribution of(–)-gacyclidine were approximately 20% higher than those of its opticalantipode (CL: 248 vs 197 ml.kg^–1.min^–1;Vd: 31.6 vs 23.5 l/kg).Protein binding (90%) was not stereoselective. Both gacyclidineenantiomers were quantifiable in spinal cord ECF 10 min after drugadministration and remained stable over the duration of the experimentin spite of changing blood concentrations. Penetration of(–)-gacyclidine was significantly higher (40%) than that of (+)-gacyclidine inall animals. Yet, exposure of spinal cord ECF was similar for bothenantiomers, and not correlated with plasma AUCs. Conclusions. The disposition of gacyclidine enantiomers isstereoselective. Both enantiomers exhibit a high affinity for spinal cord tissueand their distribution may involve a stereoselective and active transportsystem. This hypothesis could also explain the discrepancy betweendrug concentrations in plasma and spinal cord ECF.