The hydrolysis of indomethacin in aqueous solutions of polysorbates

The effect of surfactants on the rate of hydrolysis of indomethacin is reported for homologous series of polyoxyethylene (20) sorbitan fatty acid esters (polysorbates). The degradation of the solubilized drug follows a first-order process in which the rate decreases with the increasing surfactant concentration. The increasing length of hydrophobic chain of surfactant molecule has little effect on the rate of reaction. The solubilized solutions of indomethacin can be stored for at least one year. The influence of pH on the solubilizing power of polysorbate 80 is also studied.     

Micellar Solubilization of Timobesone Acetate in Aqueous and Aqueous Propylene Glycol Solutions of Nonionic Surfactants

The micellar solubilization of timobesone acetate, a novel topical corticosteroid, was studied in aqueous and aqueous propylene glycol solutions of 1 to 5% nonionic surfactants at 25°C. The surfactants used were polyoxyethylene (POE) sorbitan monofatty acid esters (polysorbates), fatty acid esters (Myrj), and fatty alcohol ethers (Brij), as well as sucrose monolaurate (Crodesta SL40). The increase in the solubility of timobesone acetate in the micellar solutions was dependent on the type and concentration of surfactant. The solubilizing capacity of the surfactant micelles and the distribution coefficient of timobesone acetate in aqueous micellar solutions were found (1) to increase with increasing length of the hydrophobic fatty acid group; (2) to increase according to the structure of the hydrophilic group in the order of POE sorbitan ester, sucrose ester, POE ester, and POE ether; (3) to be unaffected by the increase in POE chain length; and (4) to tend to decrease in surfactant containing unsaturated fatty acid groups. In aqueous propylene glycol solution, the solubilizing capacity increased slightly, i.e., up to 1.5-fold in 50% propylene glycol solution, for the ester-type surfactants (polysorbates and Myrj). But this increase was not observed in the ether-type surfactant (Brij) solution. The distribution coefficient decreased logarithmically with increasing concentrations of propylene glycol in the solution. This was caused by the logarithmic increase in the timobesone acetate solubility in the bulk phase, while the solubility in the micellar phase was practically unchanged. The results support the equilibrium distribution model of micellar solubilization.     

Influence of surface-active food additives on the integrity and permeability of rat intestinal mucosa

The influence of two surface-active food additives on the integrity and permeability of rat ileal mucosa has been studied. We determined the activity of N-acetyl-beta-glucosaminidase, a lysosomal enzyme, in the rat intestinal lumen after deposition of polyoxyethylene (20) sorbitan monostearate (polysorbate 60; Tween 60) or polyoxyethylene (20) sorbitan monooleate (polysorbate 80; Tween 80) in a section of ligated, cannulated gut. We also determined the activities of N-acetyl-beta-glucosaminidase, alkaline phosphatase, 5'-nucleotidase and phospholipase A2 in mixtures of isolated mucosal cells and polysorbate 60 or polysorbate 80. The activity of N-acetyl-beta-glucosaminidase was increased in the luminal contents of the cannulated gut 15 min after deposition of either polysorbate 60 or polysorbate 80 (10 mg/ml fluid instilled into gut). It was also increased in mixtures of mucosal cells and polysorbate 60 or polysorbate 80 (0.1-10 mg/ml). In contrast, the activities of alkaline phosphatase and 5'-nucleotidase were unaffected and that of phospholipase A2 was decreased by the presence of either polysorbate. These findings indicated that polysorbate 60 and polysorbate 80 released lysosomal enzymes from the intestinal mucosal cells and that these agents might damage the intestinal mucosa and increase its permeability. We therefore determined the intestinal permeability to sodium fluorescein in the absence and presence of polysorbate 60 or 80 and found that the permeability was slightly increased in the presence of either of the compounds at concentrations of 10 mg/ml fluid instilled into gut. It is possible therefore that surface-active food additives might impair the function of the mucosal barrier and increase the permeability of the gut to potentially toxic and pathogenic molecules.     

Formulation of a protein with propellant HFA 134a for aerosol delivery

The purpose of this study was to investigate the formulation and delivery of a protein in a pressurized metered-dose inhaler (pMDI) containing HFA 134a as the propellant for aerosol delivery. Ethanol and surfactants, including polyoxyethylene 10 oleyl ether (Brij 97), polyoxyethylene 20 oleyl ether (Brij 98), polyoxyethylene sorbitan monooleate (Tween 80) and Aerosol OT (AOT), were investigated as formulation adjuvants to improve the dose delivery characteristics of the model protein (bovine serum albumin) containing pMDI formulations. The aqueous solution of a surfactant and protein was lyophilized to obtain a solid carrier system of the protein. Readily dispersible suspensions were obtained by suspending this solid carrier system in HFA 134a with ethanol as a dispersing aid. The formulations containing Tween 80 resulted in the highest respirable fraction. This study suggested a potential formulation containing a lyophilized complex of surfactant and protein readily dispersible in HFA 134a for delivering a therapeutic protein to the respiratory tract by inhalation.     

Remarkably high inhibitory effects of docosahexaenoic acid incorporated into hybrid liposomes on the growth of tumor cells along with apoptosis

Inhibitory effects of hybrid liposomes composed of l-alpha-dimyristoylphosphatidylcholine (DMPC) and polyoxyethylene (20) sorbitan monooleate (Tween 80) including polyunsaturated fatty acids or their ethyl esters (HL-PUFA) on the growth of human tumor cells were examined in vitro. Remarkably high inhibitory effects of HL including docosahexaenoic acid (HL-DHA) and alpha-linolenic acid ethyl ester (HL-ALAE) on the growth of lung carcinoma (RERF-LC-OK and A549) cells, colon tumor (WiDr) cells and stomach tumor (MKN45) cells were obtained. The addition of vitamin E (alpha-tocopherol) to HL-DHA and -ALAE prevented almost completely the growth inhibition of A549 cells distinct from the other tumor cells used in this study. On the other hand, fluorescence microscopic and flow cytometric analyses indicated that the inhibitory effects of HL-DHA on the growth of RERF-LC-OK, WiDr and MKN45 cells could be attained through the induction of apoptosis.     

Comparison of cytotoxicity of various surfactants tested on normal human fibroblast cultures using the neutral red test, MTT assay and LDH release.

We used the neutral red test, MTT assay and lactate dehydrogenase (LDH) release to compare the potential cytotoxicity of six surfactants belonging to different classes--three non-ionic surfactants (Triton x100, octylphenoxypolyethoxy alcohol, from Orion; Tween 60, polyoxyethylene (20) sorbitan monostearate, from ICI Speciality Chemicals; Tween 80, polyoxyethylene (20) sorbitan monolaurate, from Labosi), two anionic surfactants (Texapon K1298, sodium lauryl sulphate, from Henkel; Texapon N40, sodium laurylether sulphate, from Henkel) and one cationic surfactant (benzethonium chloride, from Siber Hegner)--on human fibroblast cultures. According to the LC50 (microg ml(-1)), the tested surfactants can be classified in the following order of increasing cytotoxicity: Tween 80 < Texapon N40 < Tween 60 < Texapon K1298 < Triton x100 < benzethonium chloride.     

The Degradation of Polysorbates 20 and 80 and its Potential Impact on the Stability of Biotherapeutics

Purpose

To study the potential impact of the degradation of Polysorbates (PS) 20 and 80 on the stability of therapeutic proteins in parenteral formulations.

Method

First, degradation products of PS20 and 80 were identified. Subsequently, the effect of degraded polysorbate on physical characteristics and long-term stability of protein formulations was assessed. Further, the impact of polysorbate degradation on protein stability was evaluated via shaking stress studies on formulations spiked with artificially degraded polysorbate or degradants like fatty acids. Additionally, aged formulations with reduced polysorbate content were shaken.

Results

The degradation of polysorbate leads to a buildup of various molecules, some of which are poorly soluble, including fatty acids and polyoxyethylene (POE) esters of fatty acids. Spiking studies showed that the insoluble degradants could potentially impact protein stability and that the presence of sufficient intact polysorbate was crucial to prevent this. End-of-shelf-life shaking of protein formulations showed that the stability of various monoclonal antibodies was, however, not affected.

Conclusions

Although some degradants can potentially influence the stability of the protein (as discerned from spiking studies), degradation of polysorbates did not impact the stability of the different proteins tested in pharmaceutically relevant temperature and storage conditions.     

Comparison of impact of the different hydrophilic carriers on the properties of piperazine-containing drug.

The objective of this study was to determine the impact of a series of nonionic surfactants on the solubility of piperazine-containing drug (meclizine, MZ) in comparison to that of natural cyclodextrins (alpha-CD and beta-CD) and dimethyl-beta-cyclodextrin (DM-beta-CD). The solubility of the drug was studied in either CDs solutions or nonionic surfactant solutions. Three classes of nonionic surfactants were used namely; polyoxyethylene (POE) sorbitan fatty acid esters (polysorbates), POE fatty acid esters (Myrjs) and polyethylene oxide (PEO) fatty alcohol ethers (Brijs and Eumulgins). The solubility of MZ was increased linearly with the increasing surfactant concentration, indicating that micellar solubilization follows the partition model. It was found that the longer the hydrocarbon chain in a homologous series, the more efficient is the solubilizing power of surfactant. For example, polysorbate 80 (Tween-80) is a more efficient solubilizer than polysorbate 20 (Tween-20), indicating that the drug was incorporated in the core of micelle more than the capsular region of the micelle. On the other hand, in case of POE fatty acid esters, the solubilizing power increased with decreasing polyoxyethylene chain as Myrj 53 was more efficient than Myrj 59. In class of PEO fatty alcohol ethers, the shorter the hydrophilic chain and longer lipophilic chain, the more efficient was the solubilizing capacity. Thus, Brij 58 was more efficient solubilizer than Brij 35 and Eumulgin C1000 was more active than Eumulgin C1500. Comparatively, Eumulgin C1000 had the highest solubilizing power for MZ among the studied PEO fatty alcohol ethers and other groups of surfactants. The solubility action of surfactants toward MZ was increased by raising the temperature of the surfactant solutions from 30 to 45 degrees C. Hydrophilic macromolecules (PEG 1000 and PEG 6000) or cosolvents (glycerol and propylene glycol) have a very slight effect on the solubility of MZ and confirm the predominance of hydrophobic interaction between the drug and nonionic surfactants. A(L)-type phase solubility diagrams were obtained for the drug with alpha-, beta- and DM-beta-CDs showing that the solubility of MZ was enhanced through inclusion complexation. Comparatively, DM-beta-CD had the highest solubilizing efficiency for the drug among the investigated CDs, which could be attributed to its larger hydrophobic cavity size.     

Formation and characterization of three-component-sorbitan monoester surfactant, oil and water-creams

The effect of molecular structure of four sorbitan monoester surfactants (sorbitan monolaurate, sorbitan monopalmitate, sorbitan monostearate and sorbitan monooleate) on the formation of simple three-component creams is presented. Interfacial properties of the surfactants were determined using a du Nouy tensiometer and rheological properties of selected creams with oscillation stress sweep, creep recovery and viscosity tests. Depending on the composition, sorbitan monolaurate and sorbitan monooleate formed both o/w creams and w/o creams, while sorbitan monopalmitate and sorbitan monostearate formed only o/w creams. Sorbitan monostearate and sorbitan monopalmitate had the smallest cmc and A(cmc) values and they were the most effective surfactants in lowering the interfacial tension. These surfactants formed the most stable and elastic creams with clear linear viscoelastic regions and small compliance values. Sorbitan monolaurate and sorbitan monooleate formed viscous creams without elastic properties.     

Some considerations about the hydrophilic-lipophilic balance system

The methods and results obtained by Griffin et al. in the determination of the hydrophilic-lipophilic balance (HLB) values of non-ionic surfactants and of required HLB values of oil mixtures are reviewed in the present work. HLB values published by Griffin were compared with those obtained by calculations from theoretic chemical formulas. Griffin HLB values of polyoxyethylene alkyl ethers, polyoxyethylene monoesters and propylene glycol monoesters coincide with those obtained from such theoretical chemical formulations. These results demonstrate that, for these surfactants, Griffin did not experimentally obtain their HLB values, but instead calculated them from theoretic formulae. For the calculation of the HLB values of glycerol monostearate, sorbitan fatty acid esters and polyoxyethylene sorbitan fatty acid esters, Griffin's assumptions were possibly based upon the mean saponification values of the ester and the acid of the fatty acid. It is concluded that the HLB values of non-ionic surfactants were not rigorously defined. Moreover, Griffin could not demonstrate the validity of the assumption that individual required HLB values can be added up to obtain the overall required HLB value of an oil mixture. The HLB and required HLB values published by Griffin should only be taken as approximate guidelines.     

Kinetics of hydrolysis of polyoxyethylene (20) sorbitan fatty acid ester surfactants

The kinetics of the hydrolysis of the oleate ester of polyoxyethylene (20) sorbitan (Tween 80) in aqueous buffers were studied at an initial concentration of 0m?020% (w/v) and over the pH range of 1m?10 to 10m?28. The hydrolysis appears to be specific acid-catalysed at pH values below 3 and specific base-catalysed at pH values greater than 7m?6. The pseudo first-order rate constants for hydrogen and hydroxyl ion catalysis were determined, and the temperature and ionic strength dependence of the acid-catalysed reaction was studied. Both the initial, acid- and base-catalysed hydrolysis of Tween 80 exhibited an unusual initial, micellar surfactant concentration-rate dependence, opposite to that previously reported for the hydrolysis of anionic surfactants of the n-alkyl sulphate-type. Specifically, as the initial concentration of Tween 80 was increased above its reported critical micellar concentration, there was a progressive, marked decrease in the rate of the reaction, with the rate eventually reaching a plateau value between 0m?100 and 1m?00% (w/v). It is suggested that this behaviour is due to alterations in the micellar state of the surfactant as its concentration is increased. The influence of the chemical structure of the Tween surfactant on the acid-catalysed hydrolysis reaction at 80° was also examined using the oléate (Tween 80), stéarate (Tween 60), and palmitate (Tween 40) esters of polyoxyethylene (20) sorbitan.     

Polysorbates as novel lipid-modulating candidates for reducing serum total cholesterol and low-density lipoprotein levels in hyperlipidemic C57BL/6J mice and rats

Polysorbates are amphiphilic, non-ionic surfactants composed of fatty acid esters of polyoxyethylene sorbitan which are widely used in the cosmetic, food and pharmaceutical industries owing to these special characteristics and their low toxicity profiles. In the present study, polysorbates were investigated for their hypolipidemic activity. C57BL/6J mice and Sprague-Dawley rats were fed a high-fat diet for four weeks, then were divided into several groups, normal saline, polysorbates and positive control drugs such as lovastatin and colestyramine were administered orally to the animals for another four weeks. Complete lipid profiles of the experimental animals were determined by assessing the serum levels of total cholesterol, triglycerides, high-density lipoprotein cholesterol and low-density lipoprotein cholesterol. The results indicate that polysorbates significantly lowered the lipid components. Polysorbates are potential candidates for preventing intestinal absorption of redundant lipid from daily intake and subsequently for preventing hyperlipidemia as well as atherosclerosis.     

The novel formulation design of self-emulsifying drug delivery systems (SEDDS) type O/W microemulsion II: stable gastrointestinal absorption of a poorly water soluble new compound, ER-1258 in bile-fistula rats

The stabilization effect of the novel self-emulsifying drug delivery systems (SEDDS) type O/W microemulsion on the gastrointestinal absorption of a poorly water soluble new compound, ER-1258 was examined by bile-fistula model rats. In the components of this formulation, medium chain fatty acid triglyceride (MCT), diglyceryl monooleate (DGMO-C), polyoxyethylene hydrogenated castor oil 40 (HCO-40) and ethanol were used as an oil, a lipophilic surfactant, a hydrophilic surfactant and a solubilizer at the mixture ratio of 25/5/45/25 w/w%, respectively. The ratios of AUC in the non-treated rats to that in the bile-fistula rats were 5.1, 12.1 and 3.0 for the suspension, the oily solution and the SEDDS type O/W microemulsion, respectively. The risk from which the difference between individuals of the compound absorption amounts resulting from the flow of the bile secretion serves as the maximum was high in order of oily solution>suspension>SEDDS type O/W microemulsion. Therefore, it was verified that the SEDDS type O/W microemulsion was able to reduce this risk, compared with the other formulations. When short chain fatty acid triglyceride (Triacetin) was used as an oil, the similar effect was demonstrated in the formulation composed of sorbitan sesquioleate (SO-15) as a lipophilic surfactant and polyoxyethylene hydrogenated castor oil 60 (HCO-60) or polyoxyethylene 20 sorbitan monooleate (TO-10M) as a hydrophilic surfactant.     

维生素D2的增溶与稳定性的研究

3种结构类型不同的增溶剂对维生素D₂的增溶效应,聚氧乙烯蓖麻油类大于聚氧乙烯脱水山梨醇脂肪酸酯类及聚氧乙烯脂肪醇醚类。在聚氧乙烯蓖麻油类C-125中维生素D₂的稳定性亦大于其余的两种增溶剂。在吐瘟80胶团中丁基羟基茴香醚主要位于胶团的栅层深部,由于可与维生素D₂处于胶团的相同部位,故较位于栅层浅部及水相之中的没食子酸丙酯,有效地延缓了维生素D₂的自动氧化作用。     

Phase diagram studies for microencapsulation of pharmaceuticals using cellulose acetate trimellitate

Phase diagrams were prepared to indicate the region of microcapsule formation for the following system: cellulose acetate trimellitate, light mineral oil, and the solvent mixture (acetone:ethanol), using chloroform as the hardening agent. The effect of sorbitan monoleate, sorbitan monolaurate, and sorbitan trioleate on the region of the phase diagram for the formation of microcapsules was investigated. The results indicate that microcapsules are readily formed when the polymer concentration is in the 0.5-1.5% range and the solvent concentration is in the 5-10% range. Aggregation of microcapsules was minimized by using lower solvent concentration. Low concentrations of sorbitan monooleate in mineral oil (less than or equal to 1%) gave products that had smoother coats and more uniform particle size. Surfactants with low hydrophile:lipophile balance produced larger regions on the phase diagram for microencapsulation compared with a surfactant with higher hydrophile:lipophile balance. A mechanism for microencapsulation is described. Tartrazine microcapsules produced using different concentrations of surfactant were tested for dissolution characteristics in both acidic and neutral conditions. Tartrazine-containing microcapsules prepared by using 3% sorbitan monooleate had the lowest release in acidic conditions. The effect of surfactant and formulation concentration on microcapsule size was studied by analyzing the particle size distribution for both blank and tartrazine-containing microcapsules. The smallest microcapsule size was obtained when the sorbitan monooleate concentration was 3%. It appears that there is an upper limit for the surfactant concentration that could be used to achieve successful microencapsulation.     

Effect of Tween 20 on freeze-thawing- and agitation-induced aggregation of recombinant human factor XIII

Agitation- and freeze-thawing-induced aggregation of recombinant human factor XIII (rFXIII) is due to interfacial adsorption and denaturation at the air-liquid and ice-liquid interfaces. The aggregation pathway proceeds through soluble aggregates to formation of insoluble aggregates regardless of the denaturing stimuli. A nonionic surfactant, polyoxyethylene sorbitan monolaurate (Tween 20), greatly reduces the rate of formation of insoluble aggregates as a function of surfactant concentration, thereby stabilizing native rFXIII. Maximum protection occurs at concentrations close to the critical micelle concentration (cmc), independent of initial protein concentration. To study the mechanistic aspects of the surfactant-induced stabilization, a series of spectroscopic studies were conducted. Electron paramagnetic resonance spectroscopy indicates that binding is not occurring between Tween 20 and either the native state or a folding intermediate state of rFXIII. Further, circular dichroism spectroscopy suggests that Tween 20 does not prevent the secondary structural changes induced upon guanidinium hydrochloride-induced unfolding. Taken together, these results imply that Tween 20 protects rFXIII against freeze-thawing- and agitation-induced aggregation primarily by competing with stress-induced soluble aggregates for interfaces, inhibiting subsequent transition to insoluble aggregates.     

Non-ionic micellar affinity capillary electrophoresis for analysis of interactions between micelles and drugs

Micellar affinity capillary electrophoresis (MACE) was introduced to evaluate the affinity of various kinds of drugs as benzoic acid, salicylic acid, trinitrophenol, p-hydroxybenzoic acid and o-acetylsalicylic acid. Non-ionic micelles as Brij 35 (polyethylenglycol dodecylether), Tagat (polyoxyethylene (20) glycerol monooleate) and Tween 20 (polyoxyethylen sorbitan monolaurate) were used as a pseudostationary phase in capillary electrophoresis. For polyvinyl alcohol (PVA) coated capillary was used in this examinations. The drugs had negative electrophoretic mobilities at a pH value of pH 7.2. The negatively charged drugs migrated toward the anode and were related by their interaction with the micelles. The difference in the mobility of the drugs owing to the presence of the micelles reflected the interaction between these drugs and the micelles. Equations were derived to calculate the capacity factor k' from the migration times in the presence of micelles t' and in the absence of micelles t, the partition coefficients Pwm and the Gibbs free energy. The drugs show different interaction and affinity with the micelles in the systems. Strong interaction was observed between benzoic acid and the micelles. Furthermore, a linear relationship (R = 0.999) was obtained between deltaG(o) and ln Pwm in the micellar solubilization of drugs. These results show that deltaG(o) can give us information on the affinity and on the partition behaviour of the drugs in these systems.     

Effect of some ethoxylated sorbitan alkanoate on hydrocortisone release from methylcellulose gels

The process of hydrocortisone release from the nonionic polymer--methylcellulose hydrogels with the addition of 1% and 3% ethoxylated sorbitan alkanoate: polysorbate 20 or polysorbate 80, in the presence of 1,2-propylene glycol or PEG 200 has two stages. In the first stage the release rates are higher in comparison with the second stage. Concerning the hydrogel as a two-compartment system with micellas of polysorbates, the release rates of the first stage are in the range 4.27 x 10(-4)-6.23 x 10(-4) h-1 and in the second stage are in the range 8.29 x 10(-6)-1.16 x 10(-5) h-1 what can be affected by HLB of mentioned polysorbates.     

Physical and biophysical effects of polysorbate 20 and 80 on darbepoetin alfa

We studied the physical and biophysical affects of the nonionic surfactants polysorbate 20 and 80 and their mechanism of interaction using darbepoetin alfa, a 4-helix bundle protein, as the exemplary protein. Differences were observed between the abilities of the polysorbates to prevent surface loss/aggregation and correlated with each polysorbates initiation of micelle formation prior to the critical micelle concentration (CMC). The biophysical properties monitored by far-UV circular dichroism (CD) and tryptophan (Trp) fluorescence showed effects due to polysorbates, but were not correlated with their CMC. At a constant protein concentration PS-80 induced α-helix in the protein with a maximal effect at 15:1 molar ratio of PS-80/protein. PS-20 initially induced α-helix with a maximal effect at 1.5:1 ratio followed by a decrease in the α-helix content. PS-80 had no effect on near-UV CD but increased Trp fluorescence only at the 150:1 polysorbate/protein ratio. PS-20 decreased the near-UV CD and Trp fluorescence. Thermodynamic studies by isothermal titration calorimetry (ITC) demonstrated that the protein interacts with monomeric polysorbate, but not with polysorbate micelles. The data suggest that the polysorbates differentially interact with the protein and that the biophysical effects are dependent on the structure of the polysorbate and the polysorbate to protein ratio. © 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:3200–3217, 2009