Enhanced stability and skin permeation of ibuprofen-loaded solid lipid nanoparticles based binary solid lipid matrix: Effect of surfactant and lipid compositions.

Hypothesis: The type of emulsifier selected has an impact on the physicochemical properties of solid lipid nanoparticles (SLNs). This study was designed to compare the effects of emulsifiers on the physicochemical properties and in vitro skin performance of SLNs prepared from a binary mixture of Softisan® 378 (S378) and cetyl palmitate (CP) to those of SLNs prepared from only CP and S378. Experiments: SLNs were prepared from CP, S378, or a binary mixture of CP and S378 (1:1 w/w) as the lipid phase and stabilized with Tego®Care 450 (TG450) or poloxamer 188 (P188) containing 1.0% w/w ibuprofen loading. The physicochemical properties including the particle size, polydispersity index (PDI), zeta potential (ZP), encapsulation efficiency (E.E.), crystallinity (%CI), and polymorphism were determined after production and after storage for 180 days under different conditions. In addition, in vitro drug release and permeation through human skin was studied after production and storage at room temperature for 180 days. Finding: The particle sizes of ibuprofen-loaded SLNs (IBSLNs) stabilized with P188 (IBSLN-P188) were smaller than those of SLNs stabilized with TG450 (IBSLN-TG450) (p < 0.05). After 180 days, the particle sizes of the IBSLNs were slightly increased compared to those at the initial time but were <250 nm. The IBSLN-TG450 sample showed a higher %CI than IBSLN-P188 prepared with similar propotions of CP and S378, and ibuprofen crystals were observed in the IBSLN1-TG450 sample after storage at 4 °C for 180 days. Based on the result of the in vitro release study and the in vitro skin permeation test, the addition of S378 into the CP-matrix modified ibuprofen release and skin permeation both permeated ibuprofen through the epidermis and retained ibuprofen in the epidermis. In addition, the storage time affected the release and skin permeation of ibuprofen from the SLNs, which depended on the composition of the IBSLNs.

Formulation and Pharmaceutical Evaluation of Extemporaneous α-arbutin Creams for the Treatment of Melasma.

Concentrated 7% w/w a-arbutin cream was formulated and evaluated using O/W and W/O emulsion bases as an extemporaneous preparation for melasma treatment. Cream bases were formulated with two pH values, 4.0 and 5.5, using a hot process. The stability of the creams was studied for 60 days under three storage conditions (i.e., 2°C to 8°C, 30°C, 40°C). Cream characteristics and all aspects of product stability including physical, chemical, and microbial were investigated. Stability was defined as no dramatic change in color, viscosity, pH, and no visible microbial growth. For stability, at least 90% of the initial a-arbutin concentration quantified by stability-indicating high-performance liquid chromatography must be obtained. It was found that pH had no influence on the a-arbutin or formulations' stability. All formulations had a-arbutin remaining higher than 90% (approximately 92%) after being stored for 60 days in all storage conditions with no significant changes in pH or viscosity. All samples complied with the microbial limits test for nonsterile pharmaceutical preparation for cutaneous products. However, a color change was detected in O/W and W/O emulsions, especially at 40°C storage condition within 28 and 14 days, respectively. Drug crystals were observed in W/O emulsion stored at 2°C to 8°C. Concerning the in vitro drug release, a-arbutin was released from O/W emulsion but not from W/O emulsion. From the above results, the O/W emulsion that was developed in this study can be used as a cream base for concentrated a-arbutin as an extemporaneous preparation. The developed a-arbutin cream prepared using O/W emulsions can be used as an extemporaneous preparation with a beyond-use date of 60 days when stored at room temperature (30°C) and in the refrigerator (2°C to 8°C).

Putative hydrophobins of black poplar mushroom (Agrocybe cylindracea).

Hydrophobin proteins were extracted from Agrocybe cylindracea mycelia, the culture media (potato dextrose broth, PDB), and fruiting bodies. The putative hydrophobins obtained showed approximate sizes ranging from 8.0 to 25.0 kDa, dependent on their source. Multiple hydrophobin protein bands were detected in fruiting bodies. The hydrophobin yielded from aerial mycelia, or fruiting bodies, was approximately 6 mg/g dried weight. The crude extracts were examined for their properties in regards to surface modification, emulsification, and surface activity. Coating of hydrophobic Teflon sheet with crude extract made the surface significantly hydrophilic, whereas exposure of glass surfaces to extracts resulted in enhanced hydrophobicity. Crude extracts from culture media of A. cylindracea displayed emulsifying activity when mixed with hexane and could significantly reduce the surface tension of 60% ethanol and deionised water. The putative hydrophobin protein band from culture media (9.6 kDa), as analysed using LC-MS/MS, contained an amino acid fragment structurally similar to class I hydrophobin proteins from Basidiomycetes.

Effect of binary solid lipid matrix of wax and triglyceride on lipid crystallinity, drug-lipid interaction and drug release of ibuprofen-loaded solid lipid nanoparticles (SLN) for dermal delivery.

HYPOTHESIS: The physicochemical properties of solid lipid nanoparticles (SLN) depend on lipid compositions. An addition of secondary solid complex triglycerides (Softisan 378; S378) into solid wax (cetyl palmitate; CP) is expected to influence the properties of obtained SLN compared to SLN prepared from sole CP. EXPERIMENTS: Ibuprofen-loaded SLN (IBSLN-TG) composed of different ratios of CP and S378 were prepared and evaluated in term of size, zeta potential (ZP), entrapment efficiency (E.E.), crystallinity, lipid-drug interaction and in vitro drug release. FINDINGS: After production, all developed IBSLN-TG prepared from different ratios of CP and S378 had the particle size in the nanometer range (180-200nm) with the ZP values of higher than |-40mV| and possessed approximately 100% E.E. The release of IBSLN-TG demonstrated the biphasic pattern with a fast release followed by sustained release, which was fitted to Higuchi's kinetics. The addition of S378 into CP-matrix led to a slight decrease in particle size and surface charge, and distortion of CP crystallization. The results from 1H-NMR indicated the formation of tiny liquid S378 nanocompartments within CP-matrix. The localization of ibuprofen in the S378 nanocompartments and the interaction between ibuprofen and S378 had an impact on the release profiles of IBSLN-TG depending on the ratios of CP and S378.

Nasal absorption and local tissue reaction of insulin nanocomplexes of trimethyl chitosan derivatives in rats.

OBJECTIVES: The objective of this work was to explore the potential and safety of trimethyl chitosan (TMC) and PEGylated TMC for improved absorption of insulin after nasal administration. METHODS: The nasal absorption of insulin nanocomplexes of TMC or PEGylated TMC was evaluated in anaesthetized rats. Concomitantly, the histopathological effects of these nanocomplexes on rat nasal mucosa were studied using a perfusion fixation technique. KEY FINDINGS: All insulin nanocomplexes containing TMC or PEGylated TMC showed a 34-47% reduction in the blood glucose concentration, when the insulin absorption through the rat nasal mucosa was measured indirectly. In addition, the relative pharmacodynamic bioavailability (F(dyn)) of the formulations was found to be dependent upon the charge ratio of insulin and polymer, regardless of polymer structure. The F(dyn) apparently decreased with increasing charge ratio of insulin : polymer. Although acute alterations in nasal morphology by the formulations were affected by the charge ratio of insulin and polymer, the formulation of insulin/PEGylated TMC nanocomplexes was shown to be less toxic to the nasal epithelial membrane than insulin/TMC nanocomplexes. CONCLUSIONS: PEGylated TMC nanocomplexes were a suitable absorption enhancer for nasal delivery of insulin.

The role of mucoadhesion of trimethyl chitosan and PEGylated trimethyl chitosan nanocomplexes in insulin uptake.

The aim of this work was to investigate the role of mucoadhesion in the insulin uptake of nanocomplexes (NC) based of trimethyl chitosan (TMC) and poly(ethylene glycol) (PEG)-graft-TMC copolymers. Self-assembled insulin NC were prepared by polyelectrolyte complexation. The effects of PEGylation and positive charge density on mucoadhesion were assessed using a mucin assay and mucus-secreting HT29-MTX-E12 (E12) monolayers. The behaviors of corresponding insulin NC after adhesion to E12 were also established. All PEGylated TMC copolymers showed significantly higher levels of adhesion to mucus than unmodified TMC. The copolymer composed of 298 PEG chains per TMC macromolecules exhibited the highest level of mucoadhesion, being 3.4 times higher than TMC. The higher mucoadhesive properties of PEGylated TMC copolymers resulted from the synergistic effects of interpenetration of PEG chains into the mucus and electrostatic interaction between positive charged TMC and anionic glycoproteins present in the mucus layer. Compared to TMC, insulin NC based on PEGylated TMC copolymers demonstrated no evidence of insulin uptake improvement due to complete release of insulin from NC after adhering to mucus. CLSM revealed the localization of TMC and its corresponding insulin NC at cell surface membranes of E12.

Physicochemical properties and biocompatibility of N-trimethyl chitosan: effect of quaternization and dimethylation.

The aim of this research was to investigate the effect of degrees of quaternization (DQ) and dimethylation (DD) on physicochemical properties and cytotoxicity of N-trimethyl chitosan (TMC). TMC was synthesized by reductive methylation of chitosan in the presence of a strong base at elevated temperature and polymer characteristics were investigated. The number of methylation process and duration of reaction were demonstrated to affect the DQ and DD. An increased number of reaction steps increased DQ and decreased DD, while an extended duration of reaction increased both DQ and DD. The molecular weight of TMC was in the range of 60-550kDa. From the Mark-Houwink equation, it was found that TMC in 2% acetic acid/0.2M sodium acetate behaved as a spherical structure, approximating a random coil. The highest solubility was found with TMC of an intermediate DQ (40%) regardless of DD and molecular weight. The effect of DD on the physicochemical properties and cytotoxicity was obviously observed when proportion of DD to DQ was higher than 1. TMC with relatively high DD showed reduction in both solubility and mucoadhesion and hence decreased cytotoxicity. However, the influence of DD was insignificant when DQ of TMC was higher than 40% at which physicochemical properties and cytotoxicity were mainly dependent upon DQ.

Peroral delivery of insulin using chitosan derivatives: a comparative study of polyelectrolyte nanocomplexes and nanoparticles.

Polymeric delivery systems based on nanoparticles (NP) have emerged as a promising approach for peroral insulin delivery. Using a trimethyl chitosan (TMC) and a PEG-graft-TMC copolymer, polyelectrolyte complexes (PEC) and nanoparticles were prepared and their properties were compared. The amount of insulin was quantified by HPLC and the stability of PEC and NP upon exposure to simulated gastrointestinal (GI) fluid was monitored by dynamic laser light scattering. It was shown that polymer/insulin (+/-) charge ratio played an important role in PEC and NP formation. Stable, uniform, and spherical PEC/NP with high insulin association efficiency (AE) were formed at or close to optimized polymer/insulin (+/-) charge ratio, depending on polymer structure. All PEC were more stable in pH 6.8 simulated intestinal fluid (SIF) than NP. The PEC also appeared to play some role in protecting insulin from degradation at higher temperature and with proteolytic enzyme more efficiently than NP. On the basis of these results, polyelectrolyte complexation can be suggested as a potentially useful technique for generating insulin delivery systems for peroral administration.

Self-assembled polyelectrolyte nanocomplexes between chitosan derivatives and insulin.

Polyelectrolyte complexes (PEC) formed from chitosan derivatives and insulin was prepared and parameters influencing complex formation were characterized. Turbidimetric titration, in combination with dynamic light scattering (DLS) and laser doppler anemometry (LDA), were used to study the complexation process. The morphology of the PECs was determined using atomic force microscopy (AFM). PEC formation was predominantly pH-dependent. Complexation with insulin occurred only above critical pH value (pHc) of 6.0 for all the chitosan derivatives investigated. Soluble PECs in the size range of 200-500 nm with spherical or subspherical morphology and smooth surface structure were obtained at optimized polymer/insulin charge ratios. Optimal conditions were obtained when the pH of PECs was in the range of 6.5-8.0, depending on polymer structure. The stability of PECs was influenced by polymer chain length. Only when the MW of the polymers was > or =25-kDa PEC precipitation could be avoided. An increase in the ionic strength of the medium accelerated complex dissociation. Chitosan methylation and PEGylation significantly improved the stability of insulin in the PECs. Moreover, the PEC could protect insulin from degradation even at 50 degrees C for at least 6 h. All complexes could be lyophilized without influencing the particle size, complex concentration, and stability of insulin. On the basis of our results, we suggest that interactions involved in PEC formation were predominantly electrostatic in nature, involving the positively charged amino groups of chitosan and the negatively charged insulin above its isoelectric point. Intranasal absorption of the polyelectrolyte nanocomplexes will be studied in vivo.