Factors affecting solubilization of a poorly soluble novel tubulin-binding agent.

JCA112 is a novel tubulin-binding agent with limited aqueous solubility and high hydrophobicity. Three strategies; cyclodextrin inclusion complexation, solid dispersion (SD) formation, and liposome incorporation were evaluated to enhance the solubility of JCA112. Phase-solubility studies were carried out with hydroxypropyl ß-cyclodextrin (HPßCD), SDs were prepared by solvent evaporation method and liposomes were prepared by thin-film hydration method. Saturation solubility of the prepared formulations resulted in a significant increase in the solubility of JCA112 by all three methods. Cyclodextrin complexation resulted in a higher order complex formation increasing the aqueous solubility by 30-fold up to 105 µg/mL. Amongst the selected polymers, Poloxamer 188 (Pluronic(®) F68) was the most effective polymer in enhancing the aqueous solubility via SD, resulting in an equilibrium solubility of 50 µg/mL, independent of the drug loading. Liposomes were the most effective amongst all three techniques, with a saturation solubility of 1.8 mg/mL contributing to greater than 500-fold increase in the solubility of JCA112. The solubility enhancement by liposome was directly proportional to the drug loading. All the three strategies were successful in enhancing the solubility of the drug. Solubility enhancement by the three techniques can be attributed to the geometry/structure and the lipophilicity of the drug.

Physico chemical characterization of a novel anti-cancer agent and its comparison to Taxol(®).

Every year several thousand compounds are screened for their anti-cancer activity by a general test procedure amongst which only few selected move past the in vitro screening process. This may be due to the intrinsic property of the drug substance. Therefore, a complete physicochemical characterization of a New Chemical Entity (NCE) is essential to understand the effect of these properties on the in vitro and possibly in vivo behavior of these compounds. Various physicochemical properties such as dissociation constant, octanol-water partition co-efficient, pH solubility, stability, thermal characterization and membrane permeability were evaluated for a novel tubulin-binding agent JCA112 and were compared to that of Taxol(®). The drug exhibited a pKa value of 10.9, log P value of 2.3, pH dependent solubility, and low artificial membrane permeability. Stability of the drug substance in the in vitro screening media suggested a significant degradation during the 48-hour study duration. The results demonstrate that due to low aqueous solubility, limited membrane permeability and due to insufficient stability of JCA112 in the in vitro screening media, the drug exhibited limited anti-cancer activity. Along with challenging physicochemical characteristics, a generalization of the in vitro testing procedure may also result in loss of important anti-cancer agents. As a result, a complete understanding of the physico-chemical properties of the drug leading to prototype formulation with acceptable physico-chemical properties may be required for successful in vitro screening.

Physicochemical characterization of NPC 1161C, a novel antimalarial 8-aminoquinoline, in solution and solid state.

NPC 1161C is a novel antimalarial drug of interest because of its superior curative and prophylactic activity, and favorable toxicity profile against in vivo and in vitro models of malaria, pneumocystis carinii pneumonia, and leishmaniasis. The preformulation studies performed included determination of pK(a)s, aqueous and pH solubility, cosolvent solubility, log P, pH stability, thermal analysis, and preliminary hygroscopicity studies. The mean pK(a1), pK(a2), and pK(a3) were determined to be 10.12, 4.07, and 1.88, respectively. The aqueous solubility was found to be 2.4×10(-4) M having a saturated solution pH of 4.3-5.0 and a low intrinsic solubility of 1.6×10(-6) M. A mathematical model of the pH-solubility profile was derived from pH 2.2 to 8.0. An exponential decrease in solubility was observed with increasing pH. The excess solid phase in equilibrium with the solution in aqueous buffers was determined to be the free-base form of the drug. A significant increase in solubility was observed with all the cosolvents studied, in both unbuffered and buffered systems. Mean log P of the salt and the free base were estimated to be 2.18 and 3.70, respectively. The compound had poor stability at pH 7.0 at 37 °C, with a t (90) of 3.58 days. Thermal analysis of the drug using DSC and TGA revealed that the drug is present as a semi-crystalline powder, which transformed into the amorphous state after melting. The drug was also found to sublime at higher temperatures. Determination of physicochemical properties of NPC 1161C provided useful information for the development of a dosage form and preclinical evaluation.

Wax-based sustained release matrix pellets prepared by a novel freeze pelletization technique II. In vitro drug release studies and release mechanisms.

A novel freeze pelletization technique was evaluated for the preparation of wax-based sustained release matrix pellets. Pellets containing water-soluble drugs were successfully prepared using a variety of waxes. The drug release significantly depended on the wax type used and the aqueous drug solubility. The drug release decreased as the hydrophobicity of wax increased and the drug release increased as the aqueous drug solubility increased. In glyceryl monostearate (GMS) pellets, drug release rate decreased as the loading of theophylline increased. On the contrary, the release rate increased as the drug loading of diltiazem HCl increased in Precirol pellets. Theophylline at low drug loads existed in a dissolved state in GMS pellets and the release followed desorption kinetics. At higher loads, theophylline existed in a crystalline state and the release followed dissolution-controlled constant release for all the waxes studied. However, with the addition of increasing amounts of Brij 76, theophylline release rate increased and the release mechanism shifted to diffusion-controlled square root time kinetics. But the release of diltiazem HCl from Precirol pellets at all drug loads, followed diffusion-controlled square root time kinetics. Therefore, pellets capable of providing a variety of release profiles for different drugs can be prepared using this freeze pelletization technique by suitably modifying the pellet forming matrix compositions.

Wax-based sustained release matrix pellets prepared by a novel freeze pelletization technique I. Formulation and process variables affecting pellet characteristics.

A novel freeze pelletization technique was evaluated for the preparation of wax-based matrix pellets. Pellets containing either theophylline or diltiazem HCl were prepared using various waxes. In this technique, molten waxes along with a dispersed active ingredient were introduced as droplets into an inert and immiscible column of liquid to form pellets. An 80% (w/w) aqueous glycerol solution was found to be the most suitable column liquid for preparing spherical wax pellets. The physical stability of the molten wax suspensions was substantially improved by the addition of a 5% (w/w) colloidal silica gel. Pellet size obtained was directly proportional to the cubic root of the outer radius of the needle tip used to form pellets. Pellet size increased as the ratio of interfacial tension (gamma(LL)) to the density difference (Deltarho) between the molten matrix and the column liquid increased. Moreover, an increase in the drug load of theophylline increased the pellet size. However, an addition of a surfactant to the matrix slightly decreased the pellet size. Microscopic studies indicated that theophylline was homogenously dispersed throughout the matrix and existed in a crystalline state at higher drug loads. The percent drug recoveries ranged from 90.7 to 102.3% with acceptable drug loads up to 20% (w/w). Therefore, wax pellets containing drugs of varying aqueous solubility were successfully prepared using this technique.

Development and validation of a stability-indicating reversed-phase high performance liquid chromatography method for NPC 1161C, a novel 8-aminoquinoline anti-malarial drug.

NPC 1161C (+/-8-[(4-amino-1-methylbutyl)amino-5-(3,4-dichlorophenoxy)-6-methoxy-4-methylquinoline succinate]) is a novel investigational antimalarial drug of interest for its in vivo oral potency, activity against blood and tissue stage parasites, favorable toxicity profile, long duration of action, and utility in both prophylaxis and treatment models. The pharmaceutical development of NPC 1161C warranted the availability of an assay for the detection and quantification of the drug and its separation from the impurities and degradation products. A simple and rapid stability-indicating reversed-phase HPLC method was developed and validated according to ICH guidelines. The method was found to be linear, precise and accurate. It also proved to be selective in the presence of impurities and degradation products during forced degradation studies. The method was found to be robust by factorial experimental design and was well within the recommended parameters of system suitability testing. Degradants of the drug during stress studies were also identified using high resolution mass spectrometry.

A mathematical model to predict the size of the pellets formed in freeze pelletization techniques: parameters affecting pellet size.

A mathematical model was developed based on the theory of drop formation to predict the size of the pellets formed in the freeze pelletization process. Further the model was validated by studying the effect of various parameters on the pellet size such as viscosity of the pellet forming and column liquids, surface/interfacial tension, density difference between pellet forming and column liquids; size, shape, and material of construction of the needle tips and temperatures maintained in the columns. In this study, pellets were prepared from different matrices including polyethylene glycols and waxes. The column liquids studied were silicone oils and aqueous glycerol solutions. The surface/interfacial tension, density difference between pellet forming and column liquids and needle tip size were found to be the most important factors affecting pellet size. The viscosity of the column liquid was not found to significantly affect the size of the pellets. The size of the pellets was also not affected by the pellet forming liquids of low viscosities. An increase in the initial column temperature slightly decreased the pellet size. The mathematical model developed was found to successfully predict the size of the pellets with an average error of 3.32% for different matrices that were studied.

Content analysis and stability evaluation of selected commercial preparations of St. John's wort.

Content analysis and stability studies were performed for the commercial products of St. John's wort. Six marketed formulations were analyzed for their hypericin and pseudohypericin content. These products were standardized to contain 0.3% hypericin. Results revealed total hypericin as 7.72-38.57% of the labeled claim with varying concentrations of pseudohypericin. Stability studies were carried out under three different storage conditions: 1) 25+/-2 degrees C, 60+/-5%RH for six months, 2) 40+/-2 degrees C, 75+/-5%RH for six months, and 3) 50 degrees C for one month. Tablet formulations were also analyzed for their hardness and friability. Stability studies revealed significant decrease in the content of the marker compounds with time.

Rheological characterization of Microcrystalline Cellulose/Sodiumcarboxymethyl cellulose hydrogels using a controlled stress rheometer: part I.

Rheological properties of two different commercial grades of Microcrystalline Cellulose/Sodiumcarboxymethyl Cellulose (MCC/NaCMC) hydrogels were investigated. A controlled stress rheometer fitted with parallel plate geometry was used. Application of the Cross Model relating the viscosity and shear rate data indicated the gels are extremely shear thinning. The two grades of Avicel (RC-591 and CL-611) made of varying MCC and NaCMC concentrations, exhibited distinguishable changes in yield stress and shear thinning behavior attributable to the individual composition. The hydrogels reached structural equilibrium in 1 week after manufacture. Lot to lot variability of Formula A hydrogels had minimal influence on the rheological properties of the resulting hydrogels. The yield stress and/or initial viscosity values observed were proportional to the concentration or phase volume of the MCC/NaCMC in water.

Rheology of Microcrystalline Cellulose and Sodiumcarboxymethyl Cellulose hydrogels using a controlled stress rheometer: part II.

Rheological properties of two different commercial grades of Microcrystalline Cellulose/Sodiumcarboxymethyl Cellulose (MCC/NaCMC) hydrogels were investigated. Viscoelastic characterization of the hydrogels using a controlled stress rheometer revealed that structure formation in the gels could be detected at a concentration as low as 1.0% w/w MCC/NaCMC in purified water. The elastic modulus (G') and the linear viscoelastic region (LVR) increased with increase in hydrogel concentration. The frequency sweep study of the hydrogels exhibited a flat G', indicating a stable structure at 1.5% w/w and 2.0% w/w concentrations. The oscillation time sweep study indicated that the rate of structure build up was dependent on the concentration of hydrogel. Structure buildup at various temperatures indicated that structure formation was rapid at higher temperature (40 degrees C), and the gel point was reached fairly quickly. Phase volume of the hydrogel significantly influenced structural recovery at different temperatures.

Isothermal titration calorimetry method for determination of cyclodextrin complexation thermodynamics between artemisinin and naproxen under varying environmental conditions.

A novel isothermal titration calorimetry method was used to determine the complexation thermodynamics for hydroxypropyl-beta-cyclodextrin with artemisinin and naproxen at varying temperature and pH. The new method is very useful for studying complexation reactions between cyclodextrin and drugs with poor solubility and all the thermodynamic parameters of the cyclodextrin complexation were determined. The analysis of the thermodynamic data reveals involvement of hydrophobic bonding in the cyclodextrin complexes studied. The data also reveals the presence of enthalpy-entropy compensation in the system and provide information as to the orientation of the drug molecule inside the cyclodextrin cavity. From the thermodynamic parameters for dissociation of HPBCD complexes of artemisinin and naproxen at pH 2 it is concluded that the complexation is primarily driven by enthalpy with entropic assistance at all temperatures studied. From the dissociation studies of HPBCD complexes of naproxen at pH 10 it is concluded that the complexation is predominantly driven by entropy and moderately by enthalpy at lower temperatures and by enthalpy with entropic assistance at higher temperatures.

Interaction of artemisinin and its related compounds with hydroxypropyl-beta-cyclodextrin in solution state: experimental and molecular-modeling studies.

Hydroxypropyl-beta-cyclodextrin (HPBCD) was investigated as a possible solubilizer for a series of poorly water-soluble antimalarial drugs. The solubilities of artemisinin, artether, dihydroartemisinin, and 10-deoxoartemisinin in HPBCD solutions were studied. The phase-solubility profile of these drugs in HPBCD solutions, in the concentration range studied, can be classified as type A(L) or soluble 1:1 complexes. The solubilities of artemisinin, artether, dihydroartemisinin, and 10-deoxoartemisinin in 20% w/v solutions of HPBCD are 4.5, 1.3, 6.0, and 5.2 mg/mL, respectively. The stability constants of artemisinin, dihydroartemisinin, artether, and 10-deoxoartemisinin complexes with HPBCD are 475, 405, 327, and 146 M(-1), respectively. Three different docking methods, SYBYL DOCK, FlexiDock, and DOCK 4.0.1 were evaluated to further understand the complexation modes and applicability of the docking programs for the modeling of inclusion complexes. The results showed that DOCK 4.0.1 offers a better correlation in terms of orientation of molecules inside the cyclodextrin cavity and also in terms of docking scores.

Carrier-mediated partitioning of artemisinin into Plasmodium falciparum-infected erythrocytes.

The purpose of the present study was to characterize the partitioning of artemisinin into both uninfected and Plasmodium falciparum-infected red blood cells (RBCs). The partitioning of [(14)C](+)-artemisinin into RBCs was studied at four different hematocrit levels and eight time periods. At the optimum time of 2 h, the partitioning process was investigated with eight different drug concentrations ranging from 0.88 to 3.52 microM at 37 and 4 degrees C. The effect of the presence of unlabeled artemisinin on the partitioning of the same concentration of [(14)C]artemisinin was studied. About 35 to 40% of the drug was seen to partition into uninfected RBCs at a hematocrit of 33%, irrespective of the incubation period or the drug concentration used. In contrast, infected RBCs showed an increase in partitioning of the drug with time until saturation was achieved at 1 h. While the partitioning of artemisinin into parasitized RBCs at 37 degrees C was found to be significantly higher than that in nonparasitized RBCs, at 4 degrees C both parasitized and nonparasitized RBCs showed identical partitioning of the drug. The partitioning of [(14)C]artemisinin into parasitized RBCs was completely inhibited in the presence of the same concentration of unlabeled artemisinin. However, no such effect was observed in nonparasitized cells, and no evidence suggesting that binding of the drug in parasitized RBCs is reversible was found. The partitioning of artemisinin into parasitized RBCs was found to be rapid, saturable, temperature dependent, irreversible, and subject to competitive inhibition with unlabeled artemisinin. The results obtained suggest the involvement of carrier mediation in the partitioning of artemisinin across the parasitized RBC membrane. In contrast, simple passive diffusion of artemisinin was seen in nonparasitized RBCs.