Light-scattering studies on bile acid salts I: Pattern of self-association of sodium cholate, sodium glycocholate, and sodium taurocholate in aqueous electrolyte solutions

The pattern of association of the trihydroxy bile salts in aqueous electrolyte solutions was investigated utilizing the light-scattering technique. The turbidity of the bile salts sodium cholate, sodium taurocholate, and sodium glycocholate was determined over the concentration range of 0-25 mg/ml at 25 degrees. For sodium cholate, the concentration of the supporting electrolyte was varied from 0.15 to 0.5 M. For all bile salts in 0.15 M electrolyte, the turbidity was determined in sodium fluoride, sodium chloride, sodium bromide, and sodium iodide. Comparison of the light-scattering data with amonomer-micellar model showed that qualitative agreement was obtained; however, quantitative agreement could not be achieved. Further examination of the data showed that the light-scattering results were in good agreement with a model that includes dimers, trimers, and a higher aggregate containing approximately eight monomeric units.     

Light-scattering studies on bile acid salts II: pattern of self-association of sodium deoxycholate, sodium taurodeoxycholate, and sodium glycodeoxycholate in aqueous electrolyte solutions

The pattern of self-association of the bile salts sodium deoxycholate, sodium glycodeoxycholate, and sodium taurodeoxycholate was investigated in aqueous electrolyte solutions by the light-scattering technique. The turbidity of the bile salt solutions was obtained over the concentration range of 0-20 mg/ml at 25 degrees. These data were analyzed according to a monomer-micellar equilibrium model and a stepwise association model. Comparison of the light-scattering data with these models suggests that the monomer-micellar model may be inappropriate. Analysis of the data according to the stepwise association model suggests that the dihydroxy bile salts associate to form dimers, trimers, and tetramers in addition to a larger aggregate which varies in size depending on the degree of conjugation of the bile salt.     

胆酸钠/磷脂混合胶团对环孢素A的增溶作用研究

目的研究胆酸钠/磷脂混合胶团对难溶性多肽环孢素A(CyA)的增溶作用.方法采用共沉淀法制备胆酸钠/磷脂混合胶团,并对影响增溶作用的处方及工艺进行考查.结果相同胆酸钠浓度条件下,混合胶团对CyA的增溶能力远大于胆酸钠胶团,增大混合胶团中的磷脂用量或者降低胆酸钠/磷脂(摩尔比)均有利于提高混合胶团对药物的增溶能力.升高水合温度,增加水合介质的离子强度,加入抗氧化剂维生素E(VE)及胆固醇,均不同程度的降低了混合胶团的增溶能力.通过优化各个影响因素可获得最大的增溶量(＞5mg/mL),增加CyA溶解度100倍以上.结论胆酸钠/磷脂混合胶团可以成为CyA等难溶性多肽药物的一种新型增溶载体.     

Solubilization and pharmacokinetic behaviors of sodium cholate/lecithin-mixed micelles containing cyclosporine A

The purpose of this study was to investigate the solubilization capacity of sodium cholate/lecithin-mixed micelles and to evaluate the potential of mixed micelles as a carrier of cyclosporine A for intravenous infusion. The mixed micelles were prepared by coprecipitation technique. The formulation components and preparation procedures, which may affect the solubilization of cyclosporine A, were studied. The dilution stability of cyclosporine A-containing mixed micelles was investigated. Pharmacokinetic behaviors of mixed micelles in rabbits after intravenous infusion were compared with Sandimmun. Results showed the strategies to increase the solubility of cyclosporine A include lowering the molar ratio of sodium cholate to lecithin, increasing the concentration of lecithin, and reducing the ionic strength of the dispersion medium and temperature. The largest solubility was found to be 5.42 +/- 0.16 mg/ml. The leakage of mixed micelles in 5% glucose (5.84%) was much less than that in saline solution (36.7%). The relative bioavailability of mixed micelles versus Sandimmun was 112 +/- 20%, and statistical analysis demonstrated both preparations were bioequivalent. Sodium cholate/lecithin-mixed micelles are promising carriers in the intravenous delivery of cyclosporine A, considering their capability of large-scale production and low-toxic property.     

A permeation method for detection of self-aggregation of doxorubicin in aqueous environment

For pharmaceutical scientists, it is important to know if dissolved drug molecules are present only as monomers or in the form of aggregates in a test solution or formulation. Amphiphilic or hydrophobic drugs frequently self-associate to form dimers, trimers or higher order aggregates. Doxorubicin aggregation was examined by a previously developed permeation technique to detect oligosaccharide aggregation in aqueous solutions. At very low doxorubicin concentrations dimers and trimers have been observed, but in aqueous 0.5 mg/ml doxorubicin solutions aggregates containing about 40 molecules were observed. The permeation studies were supported by TEM studies. The results indicate that neutral doxorubicin molecules aggregate more readily than the protonated ones. Doxorubicin aggregation is a stepwise process resulting in formation of aggregates of variable sizes are enhanced aggregation with increasing doxorubicin concentration.     

Self-association and solubility behaviors of a novel anticancer agent, brequinar sodium

To aid in the selection of appropriate excipients to formulate brequinar sodium [6-fluoro-2-(2'-fluoro-1,1'-biphenyl-4-yl)-3-methyl-4-quinolinecarboxyli c acid sodium salt; DuP 785], studies were initiated to characterize thoroughly its solubility behavior. The measured solubilities at RT (approximately 23 degrees C) agreed with the theoretical values in the pH range from 0.5 to 7.2, but became significantly greater than theoretical values at pH values above 7.2. This deviation was likely due to the vertical stacking-type self-association between brequinar molecules in water. The NMR and pH methods determined a critical association concentration of 15 mg/mL. Sodium salicylate, which has been proven to interfere with molecular self-association, reduced drug solubility from 116 to 10 mg/mL. But urea, another deaggregative agent, gave about a twofold increase rather than a decrease in solubility. Addition of sodium chloride caused a 226-fold decrease in solubility. The apparent solubility product did not remain constant but decreased as sodium chloride concentration increased, suggesting that the added salt decreased the degree of self-association between brequinar molecules. Among four surfactants examined (a bile salt with a rigid fused ring versus three ordinary surfactants with a flexible chain structure), only sodium cholate significantly increased the aqueous solubility of brequinar sodium.     

Prevention of molecular self-association by sodium salicylate: effect on methylene blue

Sodium salicylate improves the rectal absorption of drugs which exhibit molecular self-association; it is suggested that salicylate may improve drug bioavailability by altering the drug self-association pattern. Methylene blue was chosen as a model molecule for investigating the interference of salicylate with drugs undergoing self-association. The effect of sodium salicylate on the concentration-dependent association of methylene blue as expressed by metachromasy was observed and compared with the effects of other additives: urea, sodium chloride, sodium acetate, sodium sulfate, and sodium benzoate. By increasing the methylene blue concentration from 10(-5) M to 2 X 10(-3) M, the lambda max peak shifts from the longer wavelength region (approximately 660 nm) of the monomer toward the shorter (approximately 600 nm) indicating the presence of dimers and other oligomers. Addition of increased concentrations of sodium salicylate had a deaggregative effect on a 10(-3) M methylene blue solution, shifting the peaks toward the monomer region. On the other hand, the addition of 0.5 M of any of the following salts: sulfate, acetate, or chloride, to a 10(-3) M, aqueous solution of methylene blue had the opposite effect, eliminating the lambda max peak at 660 nm and generating a spectrum with one peak at approximately 600 nm, which indicated a high degree of self-association. The sodium salicylate effect is concentration dependent, with a high excess (approximately 450 times on a molar scale) being necessary to reduce the self-association. At lower concentrations of salicylate, precipitation occurs in the system.(ABSTRACT TRUNCATED AT 250 WORDS)     

Study of partition of nitrazepam in bile salt micelles and the role of lecithin

The effect of trihydroxy (sodium cholate and sodium glycocholate) and dihydroxy (sodium deoxycholate and sodium glycodeoxycholate) bile salt micelles on the spectrophotometric properties and on the solubility of nitrazepam in aqueous solution, at 25.0 degrees C and at ionic strength 0.1 M in sodium chloride, has been assessed. From the results obtained it was possible to calculate the partition coefficients (Kp) of nitrazepam between aqueous and micellar phases. The partition coefficients of nitrazepam have also been determined in mixed micelles of cholate or deoxycholate with lecithin (egg yolk phosphatidylcholine), which were used as a model of the gastrointestinal tract. Drug partition was found to depend on the bile acid (number of hydroxyl groups and conjugation with glycine), and our data indicate further that addition of lecithin to bile salt micelles decreases the values of the partition coefficients in the mixed micelles at physiological pH.     

The effect of cholate on solubilisation and permeability of simple and protein-loaded phosphatidylcholine/sodium cholate mixed aggregates designed to mediate transdermal delivery of macromolecules.

Carriers for non-invasive administration of biologically important antioxidant enzymes Cu,Zn-superoxide dismutase (SOD) and catalase (CAT) were developed. Solubilisation and permeabilities of various soybean phosphatidylcholine/sodium cholate (SPC/NaChol) mixtures, mainly in the form of lipid bilayers, focussing on system properties relevant for non-invasive enzyme delivery were investigated in this work. Static and dynamic light scattering measurements gave information on the behaviour of the systems containing up to 40 mM NaChol and 30.6-1.2 mM SPC in the final suspension. The average size of such mixed aggregates was in the 100-200 nm range. Suspension turbidity decreased by 50% upon increasing nominal molar detergent/lipid ratio to NaChol/SPC = 7 and 1.25, in case of SPC = 1.2 and 19.6 mM, respectively. The effective NaChol/SPC molar ratio in bilayers saturated with the detergent was found to be: R(e)(sat) = 0.70 +/- 0.01; bilayer solubilisation point corresponded to R(e)(sol) = 0.97 +/- 0.02, independently of enzyme loading. Vesicles became very permeable to SOD when membrane bound NaChol concentration exceeded 13.7 mM, in case of total starting lipid concentration of 138 mM diluted to SPC = 19.6 mM. Specifically, we measured a 50% loss of SOD from the vesicles with an aggregate-associated molar detergent ratio NaChol/SPC approximately 0.7, which is near the saturation but well below the solubilisation limit. Calcein efflux from such vesicles was compared with SPC/NaChol/SOD mixed aggregates. Our results should contribute to the future design of vesicle mediated transdermal delivery of antioxidant enzymes.     

Structure and solubility of interleukin-2 in sodium dodecyl sulfate

The solubility of an interleukin-2 (C125A) in an aqueous sodium dodecyl sulfate (SDS) solution was examined as a function of SDS concentration. A stock interleukin-2 solution at 1.1 or 0.31 mg/mL was mixed with one-tenth volume of a stock SDS solution at various concentrations. At intermediate SDS concentrations, the interleukin-2 showed nearly complete precipitation, while lower or higher SDS concentrations resulted in an increase in protein solubility. Circular dichroic analysis indicated that the protein remaining in the supernatant at low or high SDS concentrations has an altered conformation. Sedimentation velocity analysis of interleukin-2 in 0.1%/SDS showed that about half the protein was monomeric, with the remainder as large aggregates with a broad size distribution from 20 to 100 S (Mr? 10⁵-10⁷). In contrast, interleukin-2 in 0.01% SDS has a much smaller distribution of aggregates, sedimenting at ca. 2–10 S.     

Common ion effect on solubility and dissolution rate of the sodium salt of an organic acid

The solubility and the dissolution rate of the sodium salt of an acidic drug (REV 3164; 7-chloro-5-propyl-1H,4H-[1,2,4]triazolo[4,3-alpha]quinoxaline-1,4-dione) decreased by the effect of common ion present in aqueous media. The solubility of the sodium salt of REV 3164 in a buffered medium was much lower than that in an unbuffered medium. Also, the presence of NaCl decreased its solubility in water. The apparent solubility product (K'sp) of the salt, however, did not remain constant when the concentration of NaCl was changed. A decrease in K'sp value with the increase in NaCl concentration was observed; for example, the K'sp values at 0 and 1 M NaCl were 7.84 X 10(-4) and 3.94 X 10(-4) M2, respectively. Even when corrected for the effect of ionic strength, the solubility product decreased. This decrease in the solubility product in the presence of NaCl indicated a decrease in the degree of self-association (increase in activity coefficient) of the drug in aqueous media.     

Enhancement of oral bioavailability of the poorly water-soluble drug silybin by sodium cholate/ phospholipid-mixed micelles

Aim:

To evaluate a mixed micellar drug delivery system composed of sodium cholate and phospholipid for oral administration of silybin, a promising hepatoprotectants.

Methods:

The optimum formulation of sodium cholate/phospholipid-mixed micelles containing silybin was obtained based on the study of pseudo-ternary phase diagram. The dissolution of silybin-mixed micelles was investigated. The pharmacokinetic characteristics and bioavailability after oral administration of silybin-mixed micelles and silybin-N-methylglucamine were compared in dogs.

Results:

The mean particle size of prepared mixed micelles was 75.9±4.2 nm. The largest solubility of silybin was found to be 10.0±1.1 mg/mL in the optimum formulation of mixed micelles. The silybin-sodium cholate/phospholipid-mixed micelles showed a very slow release of silybin 17.5% (w/w) within 72 h in phosphate buffer (pH 7.4) and 15.6% (w/w) in HCl solution (pH 1.2). After oral administration to dogs, the relative bioavailability of mixed micelles versus silybin-N-methylglucamine in dogs was 252.0%.

Conclusion:

Sodium cholate/phospholipid-mixed micelles are promising carriers in orally delivery of silybin, considering their capability of enhancing bioavailability and large-scale production.     

Solubility-Excipient Classification Gradient Maps

This study assessed the effect of excipients (sodium taurocholate, 2-hydroxypropyl-f-cyclodextrin, potassium chloride, propylene glycol, 1-methyl-2-pyrrolidone, and polyethylene glycol 400) on the apparent intrinsic solubility properties of eight sparingly soluble drugs (four bases, two neutrals, and two acids): astemizole, butacaine, clotrimazole, dipyridamole, griseofulvin, progesterone, glibenclamide, and mefenemic acid. Over 1,200 UV-based solubility measurements (pH 3-10) were made with a high-throughput instrument. New equations, based on the "shift-in-pKa" method, were derived to interpret the complicated solubility-pH dependence observed, and poorly predicted by the Henderson-Hasselbalch equation. An intrinsic solubility-excipient classification gradient map visualization tool was developed to rank order the compounds and the excipients. In excipient-free solutions, all of the ionizable compounds formed either uncharged or mixed-charge aggregates. Mefenamic acid formed anionic dimers and trimers. Glibenclamide displayed a tendency to form monoanionic dimers. Dipyridamole and butacaine tended to form uncharged aggregates. With strong excipients, the tendency to form aggregates diminished, except in the case of glibenclamide. We conclude that a low-cost, compound-sparing, and reasonably accurate high-throughput assay which can be used in early screening to prioritize candidate molecules by their eventual developability via the excipient route is possible with the aid of the "self-organized" intrinsic solubility-excipient classification gradient maps.     

Prevention of molecular self-association by sodium salicylate: effect on insulin and 6-carboxyfluorescein

The effect of sodium salicylate on the concentration-dependent self-association of insulin and 6-carboxyfluorescein (CF), as expressed by metachromasy, fluorescence, and changes in aqueous solubility, was learned. By decreasing the CF concentration from 12 to 0.48 microgram.ml-1, lambda max peaks shift from the shorter wavelengths (451, 474 nm), indicating the presence of oligomers, toward the monomer wavelength region (484 nm). Sodium salicylate shifts the peaks of a 12 micrograms.mL-1 CF solution towards the monomer region, eliminating the peak at the lower wavelengths and generating a spectrum with one peak at 490 nm, the effect being concentration dependent. The fluorescence of insulin and CF solutions increases with their concentration. Quenching of these solutions was observed, up to complete elimination of fluorescence, when various concentrations of salicylate were added. The water solubility of both molecules, CF and insulin, was considerably increased with the addition of increasing concentrations of salicylate to the solutions: at 37 degrees C, 2.5 M sodium salicylate solution increases the CF solubility 532 times from 12.2 to 6.5 mg.mL-1, and 1.5 M salicylate increases the solubility of insulin 7875 times, thus an aqueous solution containing 630 mg.mL-1 of insulin may be prepared. The results obtained here, together with our previously reported data, indicate that the interference between sodium salicylate and drug self-association behavior, by increasing drug solubility, may substantially contribute to the improved drug bioavailability mediated by salicylate.     

A comparative study of the effect of bile salts on the absorption of quinalbarbitone sodium in goldfish.

The goldfish has been used as a model membrane to estimate the absorption of quinalbarbitone sodium by measurement of overturn time. The effect of sodium cholate, sodium deoxycholate, sodium chenodeoxycholate and sodium taurodeoxycholate on the quinalbarbitone-induced overturn time has been investigated and differences in capacity to promote absorption have been demonstrated. These differences could not be attributed to variations in solubility or partitioning of the drug within the membrane by the bile salts or to the relative hydrophobicity of each molecule. It is suggested that the bile salts must in someway affect the integrity of the goldfish membrane rendering it more permeable to quinalbarbitone sodium.     

Absorption-excipient-pH classification gradient maps: sparingly soluble drugs and the pH partition hypothesis.

This study of sparingly soluble model drugs assesses (a) how pH and the aqueous boundary layer factors may affect in vitro and in vivo absorption, (b) to what extent single excipients (sodium taurocholate, hydroxypropyl-beta-cyclodextrin, KCl, propylene glycol, methylpyrrolidone, and polyethylene glycol 400) can mitigate adverse absorption effects, and (c) how a novel rank-order visualization tool can be applied in high-throughput screening to identify promising single-excipient effects on the absorption potential of test compounds. The products of accurately measured solubility and artificial-membrane permeability (PAMPA) values at pH 5.0, 6.2, and 7.4, fully taking into account factors such as aqueous boundary layer resistance, membrane retention, and the formation of drug dimers and trimers, were used to define a flux function. A "self-organized" data visualization tool based on the flux function was mined for the promising excipient-drug combinations. In excipient-free solutions, most of the compounds studied formed aggregates. The presence of an excipient predominantly lowered permeability, but most often not by the same amount as solubility was elevated. The compounds with absorption potential most helped by excipients were: clotrimazole>griseofulvin>progesterone>dipyridamole>glibenclamide>mefenamic acid>butacaine>astemizole. The HP-beta-CD effect observed for albendazole and glibenclamide appeared to follow Cmax trends in published pharmacokinetics studies. A surprising outcome of the in vitro measurements was that the classical pH Partition Hypothesis can be "inverted" in its monotonicity by sparingly soluble compounds.     

Self-association of disodium cromoglycate in dilute aqueous solution

The association characteristics of isotropic solutions of disodium cromoglycate in water and aqueous electrolyte solutions have been examined using light-scattering techniques. The association in these solutions may be satisfactorily described using anti-cooperative stepwise association models in which successive association constants decrease with increasing aggregate size. Size distribution curves show that the extent of association is limited mainly to dimers and trimers.     

Evaluation of the surfactant properties of ascorbyl palmitate sodium salt.

In this paper we report on the physicochemical surface properties of ascorbyl palmitate (Asc16) and of its sodium salt (Asc16Na) with a view to their use as surfactants. Asc16Na was synthesized from ascorbyl palmitate by neutralizing the -OH groups in position 3 of the ascorbyl ring. The acid-base properties, thermal analysis and stability of Asc16Na monomers were determined. Self-assembling parameters of micellar aggregates in aqueous dispersions through critical micellar concentration (CMC) and critical micellar temperature (CMT) were measured. Asc16Na micellar dispersions efficiently solubilize poorly soluble drugs such as phenacetin and griseofulvin, and enhance their apparent solubility in aqueous environments. Stability tests showed that Asc16Na is more unstable than ascorbyl palmitate. Ascorbyl palmitate and its sodium salt are insoluble at room temperature in water, but their solubilities strongly depend on temperature, and largely increase above the CMT. Although Asc16Na is insoluble at room temperature, it is more soluble than Asc16, and its CMT significantly lowers in the undissociated acidic form. The apparent solubilities of phenacetin and griseofulvin are increased in Asc16Na aqueous solutions. The Asc16Na potential use as surfactant is restricted by its low stability in water, therefore the addition of some antioxidant species is necessary.     

Mixed micelle proliposomes for preparation of liposomes containing amphotericin B, in-vitro and ex-vivo studies

The aim of this work was to produce a form of injectable liposomes containing amphotericin B derived from mixed micelle proliposomes. Mixed micelles were derived from a mixture of lecithin/sodium cholate in aqueous media. The solubility of amphotericin B in proliposomes was studied as a function of lipid composition (total lipid concentration, molar ratio of lecithin/sodium cholate), and the dispersion media (pH, ionic strength, presence or absence of human serum albumin), and the temperature. The data show that micelle-->liposome transformation occurs during the dilution of proliposomes containing amphotericin B. These transformations could be followed via transmission electron microscopy (TEM). Data related to dilution of proliposomes as well, show that under no circumstance there occurs any precipitation that might be assigned to the decreased solubility of amphotericin B. These indicate that the incorporated drug also participates during the transformation of the proliposomes into liposomes. It is thus concluded that mixed micelle proliposomes are prime candidates for the production of a form of injectable amphotericin B in liposomes.     

Preparation and <Emphasis Type="Italic">in vitro</Emphasis> evaluation of povidone-sodium cholate-phospholipid mixed micelles for the solubilization of poorly soluble drugs

Mixed micelles made of polyvinylpyrrolidone (PVP), sodium cholate, and phospholipids were prepared to improve the solubility of poorly water-soluble drugs. Sylibin, a drug used in treating liver diseases, was incorporated into the mixed micelles. The formulation of sylibin containing PVP-sodium cholate-phospholipid mixed micelles with an optimized composition (PVP/sodium cholate/phospholipid/silybin = 3:3:4:1∼2 by weight) was obtained based on the study of pseudoternary phase diagrams. The critical micelle concentration was used to evaluate the micellar stability towards dilution. The results showed that addition of PVP to sodium-cholate-phospholipid mixed micelles increased stability. The solubility of sylibin in PVP-sodium cholate-phospholipid mixed micelles was higher than that in pure water or in sodium cholate-phospholipid mixed micelles. In a stability study, we found that PVP-sodium cholate-phospholipid mixed micelles showed good stability. After 3 months storage at 40°C, just 2.6% sylibin was lost with only minor changes of the particle size when compared to a reference formulation containing sodium cholate and phospholipid mixed micelles. In addition, the developed formulation significantly improved in vitro drug release. The time required to release 50% sylibin (t50%) from sodium cholate and phospholipid mixed micelles was 326 h, while the t50% from PVP-sodium cholate-phospholipid mixed micelles was only 51.1 h. Our results suggest that these mixed micelles might have significant potential application to the biomedical field.