3'-Azido-3'-deoxy-5'-O-isonicotinoylthymidine: a novel antiretroviral analog of zidovudine. II. Stability in aqueous media and experimental and theoretical ionization constants

Degradation of 3'-azido-3'-deoxy-5'-O-isonicotinoylthymidine (AZT-Iso), an antiretroviral derivative of zidovudine, was investigated in buffer pH 7.4, mu = 300 mOsm at 37, 50 and 60 degrees C, and in water (pH 6.6, 37 degrees C), giving zidovudine (AZT) and isonicotinic acid (INA) as products. The rate constants were determined by reversed-phase HPLC showing pseudo-first-order kinetics related to the residual amount of AZT-Iso. In this way, the studied compound was demonstrated to be 153 times more stable in water than in buffer solution at 37 degrees C. The analytical method was conveniently validated demonstrating to be a rapid and accurate stability-indicating technique. In addition, experimental and theoretical values of pKa were determined.     

3′-Azido-3′-deoxy-5′-O-isonicotinoylthymidine: A Novel Antiretroviral Analog of Zidovudine. II. Stability in Aqueous Media and Experimental and Theoretical Ionization Constants

Abstract

Degradation of 3′-azido-3′-deoxy-5′-O-isonicotinoylthymidine (AZT-Iso), an antiretroviral derivative of zidovudine, was investigated in buffer pH 7.4, µ = 300 mOsm at 37, 50 and 60°C, and in water (pH 6.6, 37°C), giving zidovudine (AZT) and isonicotinic acid (INA) as products. The rate constants were determined by reversed-phase HPLC showing pseudo-first-order kinetics related to the residual amount of AZT-Iso. In this way, the studied compound was demonstrated to be 153 times more stable in water than in buffer solution at 37°C. The analytical method was conveniently validated demonstrating to be a rapid and accurate stability-indicating technique. In addition, experimental and theoretical values of pKa were determined.     

Reaction Kinetics of Immobilized α-Chymotrypsin in Organic Media 1. Influence at Solvent Polarity

Esterification of N-acetyl phenylalanine with ethanol catalyzed by immobilized α-chymotrypsin (EC 3.4.21.1) was studied in various reaction media. The effect of reaction medium polarity on enzymatic activity as well as equilibrium yield was measured. The reaction rate increased with increasing amounts of added water, reaching an optimum corresponding to water saturation of the reaction medium. Further additions of water resulted in decreased activity. Bell-shaped activity profiles were obtained for all reaction media tested. Reaction media consisting of pure solvents and of mixtures of solvents were used. The enzymatic activity and the equilibrium yield increased with decreased polarity of the medium. Maximum activity was found in a reaction medium consisting of 80% diisopropyl ether and 20% heptane. The enzymatic activity obtained at optimal water additions in the different solvents and solvents mixtures could be correlated to the solubility of water and the log P of the medium. The equilibrium yield of the reaction was much more closely correlated to the solubility of water than the log P. Much lower enzymatic activity was obtained when solvent mixtures producing water-miscible media were created, than in mixtures producing water-immiscible media, such as mixtures of acetonitrile and diisopropyl ether. The equilibrium yield could be increased by decreasing the water content in the reaction medium, which reduced the water activity.     

Effect of ethanol on the thermal stability of tRNA molecules

The thermal denaturation of E. coli unfractionated tRNA in ethanol/water mixtures has been studied as a function of alcohol concentration in the water-rich region (mole fraction of co-solvent chi 2 less than 0.2). The results show that with increasing alcohol concentration the melting temperature of tRNA first reaches a minimum at an intermediate composition chi *2 approximately equal to 0.055 and then increases with increasing chi 2. The value of chi *2 is close to that at which structural changes in the mixture occur as inferred from compressibility and optical absorption measurements. The present experimental data support the assumptions that the dominant mechanism by which ethanol affects the thermal stability of tRNA molecules is through its effect on the structure of water.     

Study of the hydrolysis and ionization constants of Schiff base from pyridoxal 5''-phosphate and n-hexylamine in partially aqueous solvents. An application to phosphorylase b.

Formation and hydrolysis rate constants as well as equilibrium constants of the Schiff base derived from pyridoxal 5'-phosphate and n-hexylamine were determined between pH 3.5 and 7.5 in ethanol/water mixtures (3:17, v/v, and 49:1, v/v). The results indicate that solvent polarity scarcely alters the values of these constants but that they are dependent on the pH. Spectrophotometric titration of this Schiff base was also carried out. We found that a pKa value of 6.1, attributed in high-polarity media to protonation of the pyridine nitrogen atom, is independent of solvent polarity, whereas the pKa of the monoprotonated form of the imine falls from 12.5 in ethanol/water (3:17) to 11.3 in ethanol/water (49:1). Fitting of the experimental results for the hydrolysis to a theoretical model indicates the existence of a group with a pKa value of 6.1 that is crucial in the variation of kinetic constant of hydrolysis with pH. Studies of the reactivity of the coenzyme (pyridoxal 5'-phosphate) of glycogen phosphorylase b with hydroxylamine show that this reaction only occurs when the pH value of solution is below 6.5 and the hydrolysis of imine bond has started. We propose that the decrease in activity of phosphorylase b when the pH value is less than 6.2 must be caused by the cleavage of enzyme-coenzyme binding and that this may be related with protonation of the pyridine nitrogen atom of pyridoxal 5'-phosphate.     

Biowaiver or Bioequivalence: Ambiguity in Sildenafil Citrate BCS Classification

The aim of the present study is to contribute to the scientific characterization of sildenafil citrate according to the Biopharmaceutics Classification System, following the World Health Organization (WHO) guidelines for biowaivers. The solubility and intestinal permeability data of sildenafil citrate were collected from literature; however, the experimental solubility studies are inconclusive and its “high permeability” suggests an API in the borderline of BCS Class I and Class II. The pH-solubility profile was determined using the saturation shake-flask method over the pH range of 1.2–6.8 at a temperature of 37 °C in aqueous media. The intestinal permeability was determined in rat by a closed-loop in situ perfusion method (the Doluisio technique). The solubility of sildenafil citrate is pH-dependent and at pH 6.8 the dose/solubility ratio obtained does not meet the WHO criteria for “high solubility.” The high permeability values obtained by in situ intestinal perfusion in rat reinforce the published permeability data for sildenafil citrate. The experimental results obtained and the data available in the literature suggest that sildenafil citrate is clearly a Class II of BCS, according to the current biopharmaceutics classification system and WHO guidance.     

Solubility enhancement of cox-2 inhibitors using various solvent systems

This study examined the solubility enhancement of 4 cox-2 inhibitors, celecoxib, rofecoxib, meloxicam, and nimesulide, using a series of pure solvents and solvent mixtures. Water, alcohols, glycols, glycerol, and polyethylene glycol 400 (PEG 400) were used as solvents and water-ethanol, glycerol-ethanol, and polyethylene glycol-ethanol were used as mixed-solvent systems. A pH-solubility profile of drugs was obtained in the pH range 7.0 to 10.9 using 0.05M glycine-sodium hydroxide buffer solutions. Lower alcohols, higher glycols, and PEG 400 were found to be good solvents for these drugs. The aqueous solubility of celecoxib, rofecoxib, and nimesulide could be enhanced significantly by using ethanol as the second solvent. Among the mixed-solvent systems, PEG 400-ethanol system had highest solubilization potential. In the case of meloxicam and nimesulide, solubility increased significantly with increase in pH value. Physico-chemical properties of the solvent such as polarity, intermolecular interactions, and the ability of the solvent to form a hydrogen bond with the drug molecules were found to be the major factors involved in the dissolution of drugs by pure solvents. The greater the difference in the polarity of the 2 solvents in a given mixed solvent, the greater was the solubilization power. However, in a given mixed-solvent system, the solubilization power could not be related to the polarity of the drugs. Significance of the solubility data in relation to the development of formulations has also been discussed in this study.     

Transesterification of canola oil in mixed methanol/ethanol system and use of esters as lubricity additive

Transesterification of canola oil was carried out with methanol, ethanol, and various mixtures of methanol/ethanol, keeping the molar ratio of oil to alcohol 1:6 and using KOH as a catalyst. Mixtures of alcohol increased the rate of transesterification reaction and produced methyl as well as ethyl esters. The increased rate was result of better solubility of oil in reaction mixture due to better solvent properties of ethanol than methanol and equilibrium due to methanol. With 3:3 molar ratio of methanol to ethanol {MEE (3:3)} the amount of ethyl ester formed was 50% that of methyl ester. Properties (acid value, viscosity, density) of all esters including mixed esters were within the limits of ASTM standards. Lubricities of these esters are in the order: ethyl ester>methyl ethyl ester>methyl ester.     

THE SOLUBILITIES OF THE l-DIHALOGENATED TYROSINES IN ETHANOL-WATER MIXTURES AND CERTAIN RELATED DATA

1. The solubilities of l-tyrosine, dichloro-l-tyrosine (hydrated), dibromo-l-tyrosine (anhydrous), and diiodo-l-tyrosine in ethanol-water mixtures at 25° have been determined. 2. It was found that the solubilities of the dihalogenated substitution products of l-tyrosine are increased by addition of ethanol up to a certain concentration. Further addition of ethanol leads to a decrease in solubility. The solubility of l-tyrosine is decreased by addition of ethanol. 3. Dichloro-l-tyrosine (hydrated) was found to change to the anhydrous form when allowed to stand in the presence of ethanol. 4. The apparent heat of solution of diiodo-l-tyrosine in an ethanol-water mixture has been determined. 5. The solubility of dl-thyroxine at 30° has been determined in urea solution, ethanol, dioxan, ethylene glycol, and propylene glycol.     

Copper(II) interaction with 3,5-diisopropylsalicylic acid (Dips): new insights on its role as a potential *OH inactivating ligand

The purpose of this study was to identify the low molecular mass complexes formed between copper(II) and 3,5-diisopropylsalicylic acid (Dips) in physiological conditions. Copper(II)-Dips complex equilibria were determined using glass electrode potentiometry and their solution structures checked by UV-visible (UV-vis) spectrophotometry. Because of the low solubility of Dips in water, the equilibria were investigated in different water/ethanol mixtures. Formation constants were extrapolated to 100% water and then compared with the values obtained for the other anti-inflammatory drugs previously studied. Given the prime role of histidine as the copper(II) ligand in blood plasma, copper(II)-histidine-Dips ternary equilibria were studied under similar experimental conditions. Computer simulations of copper(II) distribution relative to different biofluids, gastrointestinal (g.i.) fluid and blood plasma, show that like salicylic and anthranilic acids, Dips favors g.i. copper absorption, but cannot exert any significant influence on plasma copper distribution. Moreover, Dips can mobilize increasing fractions of copper(II) as the pH decreases. In conclusion, Dips seems to correspond to the notion of *OH-inactivating ligand (OIL) as determined for anthranilic acid.     

Chemical properties of bile acids. IV. Acidity constants of glycine-conjugated bile acids

The dissociation constants for the carboxyl group of a series of glycine (N-acyl)-conjugated and unconjugated bile acids were determined by potentiometric titration using dimethylsulfoxide-water and methanol-water mixtures of varying proportions. The pKa values in water were calculated by extrapolating the experimental values determined in different mole fractions of the organic solvent mixtures. The following values were obtained: 3.9 +/- 0.1 for glycine-conjugated bile acids and 5.0 +/- 0.1 for unconjugated bile acids, as general pKa values for the two classes of bile acids, respectively. The amidation of bile acids with glycine lowers the pKa value because of the proximity of the amide bond to the terminal carboxyl group. Bile acid dissociation constants are independent of the substituents in the steroid nucleus, since inductive effects of the hydroxyl groups on the steroid nucleus are too distant from the acidic group at the end of the side chain to influence its ionization.     

Prediction of Drug Solubility in Lipid Mixtures from the Individual Ingredients

The purpose of this research was to investigate the relationship of drug solubility in a complex lipid mixture to that of the individual ingredients with the goal of substantiating a quantitative equation that can be applied in formulation development of lipid dosage forms. To this end, the solubility of four drugs, which span a large range of physicochemical properties, was evaluated in 18 lipid ingredients that cover the major lipid classes. To assess the solubility relation in complex lipid mixtures in an unbiased manner, the experiments were created as an experimental design with the ability to detect cubic curvature in the solubility-lipid composition space. The results demonstrated that for all drugs, irrespective of their significantly distinct physicochemical properties, solubility in lipid mixtures can be readily estimated as a simple weighted average of the drug solubility in the individual ingredients. This result is of great value to formulators who can minimize a large number of solubility experiments once a basis set of solubility is determined in individual lipids.     

The solubilization of progesterone by mixed bile,salt-phospholipid sols

The solubility of progesterone was determined in several different bile salt-phospholipid mixtures, and it is concluded that: (1) The solubility in unconjugated bile salts is greater than in the conjugated analogues, and the solubility in deoxycholate solutions is twice that in cholate solutions. (2) Substitution of hydroxyl groups in the 11 and 21 positions of progesterone increases solubility, whilst substitution in the 17-position decreases solubility in bile salt solutions. (3) Progesterone solubility in mixed bile salt solutions is proportional to the mole ratio of the surfactant mixture. (4) Sodium deoxycholate (SDC)-phospholipid sols show no such linear solubilizing properties; a minimum occurring at a mole ratio of SDC to phospholipid of 1 : 4. (5) There is a break in the solubility curve of progesterone in lysophosphatidycholine (LPC)/phosphatidylcholine (PC) mixtures at a mole ratio of 65 : 35 coincident with maximum viscosity. (6) Introduction of SDC into LPC/PC mixtures results in decreased progesterone solubility.     

Solid–Liquid Equilibrium of Maltitol Aqueous Solutions—Implications on the Crystallization Behavior and Process

The solubility of maltitol in pure water and industrial syrup was measured in a temperature range from 10 to 90 °C. Maltitol is highly soluble in water, and this yields high viscosity values for the saturated aqueous solutions at different temperatures. In addition, solubility of maltitol in ethanol/water mixtures was followed at 30, 35, 45, and 55 °C. Results show that maltitol solubility is highly dependent on water content in the solvent mixture. Moreover, it increases monotonically with temperature. The logarithm of viscosity changes linearly against the mole fraction of maltitol in the aqueous solutions up to saturation. The saturated solutions showed a Newtonian behavior in a temperature range from 20 to 90 °C. Maltitol is also characterized in supersaturated solutions by a narrow metastable zone, which slightly increases as temperature is raised. The density of aqueous solutions of maltitol was measured as a function of molality up to saturation at 20 °C, and results show that density can be correlated with concentration according to a linear relation. The obtained results were used to explain maltitol crystallization, which exhibits a high nucleation rate and a slow growth leading to small size crystals.     

Anion Effect on the Binary and Ternary Phase Diagrams of Chiral Medetomidine Salts and Conglomerate Crystal Formation

The binary phase diagrams of hydrogen halides salts of medetomidine (Med.HX, X:Br,I) and hydrogen oxalate salt of medetomidine (Med.Ox) were determined based on thermogravimetric/differential thermal analysis (TGA/DTA) and their crystal structure behavior was confirmed by comparison of the X‐ray diffractometry and FT‐IR spectroscopy of the racemate and pure enantiomer. All hydrogen halide salts presented racemic compound behavior. Heat of fusion of halides salt of (rac)‐medetomidine decreased with ionic radius increase. Eutectic points for Med.HCl (previously reported), Med.HBr, and Med.HI rest were unchanged approximately. The solubility of different enantiomeric mixtures of Med.HBr and Med.HI were measured at 10, 20, and 30°C in 2‐propanol showing a solubility increase with ionic radius. A binary phase diagram of Med.Ox shows a racemic conglomerate behavior. The solubility of enantiomeric mixtures of Med.Ox were measured at 10, 20, 30, and 40°C. The ternary phase diagram of Med.Ox in ethanol conforms to a conglomerate crystal forming system, favoring its enantiomeric purification by preferential crystallization. Chirality 26:183–188, 2014. © 2014 Wiley Periodicals, Inc.     

Determination of glucose and ethanol effective diffusion coefficients in Ca-alginate gel.

Glucose and ethanol diffusion coefficients in 2% Ca-alginate gel were measured using the experimental technique based on solute diffusion into or out of gel beads in a well-stirred solution. The aim of the study was to make the measurements under typical conditions found in alcoholic fermentations, such as the concentrations of glucose (100 g l-1) and ethanol (50 g l-1), the simultaneous counter-diffusion of glucose and ethanol, and the presence of cells in the gel beads at a level of 10(9) cells g-1 of beads. Previously, an evaluation of the error associated with the methodology used indicated how the experimental procedure would minimize the error. The individual measurement of glucose and ethanol coefficients in 2% Ca-alginate with no cells gave values of 5.1 and 9.6 x 10(-6) cm2 s-1, respectively, which are lower than those in water. When the effect of counter-diffusion was investigated, both coefficients decreased: glucose by 14% and ethanol by 28%. When cells were incorporated into the beads, only the ethanol coefficient decreased significantly, while the glucose coefficient apparently increased its value to 6.9 10(-6) cm2 s-1.     

The solubility of sickle and non-sickle hemoglobins in concentrated phosphate buffer.

A new turbidimetric method for the direct measurement of the solubility of oxy- and deoxyhemoglobins (Hb) in concentrated phosphate buffer has been established. The principle of the method is the formation of a homogeneous emulsion when hemoglobin is introduced in concentrated phosphate buffer. The solubility of the oxy and deoxy forms of Hb A, Hb S, Hb C, Hb F, and Hb CHarlem (beta 6Glu leads to Val, beta 73Asp leads to Asn) has been studied. The solubility of deoxy-Hb S was the lowest and the solubility curve was broader than those of the other hemoglobins indicating that the aggregates of deoxy Hb S require more water to be dissolved. The solubility of oxy- and deoxyhemoglobins depends on temperature and pH. The solubility of hemoglobins is increased as the temperature is lowered and the pH is raised. The pH dependency of the solubility of deoxy-Hb S in high phosphate buffer was opposite to that of the minimum gelling concentration of deoxy-Hb S. The order of the solubility of Hb CHarlem, Hb FS, Hb AS, Hb CS, and Hb S in concentrated phosphate buffer corresponds to the order of minimum gelling concentration of these hemoglobins or hemoglobin mixtures. Solubility studies of a 1:1 mixture of deoxy-Hb A and deoxy-Hb S show that deoxy-Hb A aggregates in 2.42 M phosphate buffer in which pure deoxy-Hb A is totally soluble. This result indicates that deoxy-Hb S interacts with deoxy-Hb A and decreases its solubility.     

A comparison of force fields for ethanol–water mixtures

Aqueous ethanol mixtures are studied through molecular dynamics simulations with the focus on exploring how various force field models reproduce the association and its influence on selected thermo-physical properties of these mixtures. The most important conclusion seems to be the inadequacy of all classical force fields to reproduce the very peculiar shape of the excess enthalpy of these mixtures, as a function of the ethanol concentration, neither quantitatively nor qualitatively. The Kirkwood–Buff (KB) integrals calculated using the simulation data follow the same trends as the experimental ones. This suggests complicated correlation of the excess enthalpy with the concentration fluctuation and clustering in these mixtures. The KB force field shows better overall agreement with experimental results than the other studied models.     

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_as, 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.     

Incorporation of phosphonic acid diesters into lipid model membranes. Part II. X-ray and neutron diffraction studies.

Mixtures of egg phosphatidylcholine and phosphonic acid diethyl or dibutyl esters of the general type RP(O)(OR')2 with R = hexane or dodecane were studied at room temperature in the fluid lamellar state by X-ray and by neutron diffraction. Generally a molar ratio of lipid and ester of 1:0.5 was used. Additionally an equimolar lipid/ester mixture of hexane phosphonic acid diethyl ester was studied. Depending on the ester used and its concentration a single L alpha-phase was observed above a certain water content which changes to an L alpha + water two phase system at high water concentration. Despite the large amounts of the amphiphilic ester molecules incorporated in the membrane and their high molecular asymmetry, the mixtures qualitatively show the typical hydration and swelling behaviour of non-charged lipid membranes. However, the incorporation of the esters induces a higher hydration capacity, a lateral extension and a decrease in membrane thickness. The position of the ester molecules and their orientation in the membrane were determined by neutron diffraction using partially deuterated esters. The esters were found to be located with their phosphonic moiety near or in the lipid/water interface. The lamellar structure contradicts this location of the cone-shaped ester molecules which should increase the tendency to form hexagonal structures. However, the experimental findings can be understood if one considers a partial interdigitation of the last hydrocarbon groups of the lipid chains accompanied by a larger disorder in the hydrophobic centre of the membrane. In the case of hexane phosphonic acid dibutyl ester, a vertical translocation of the ester takes place below a certain water content where it is distributed between two locations at the lipid water interface and the centre of the membrane.