Model Hydrophobic Compound Research Articles

Evaluation of the Stability of Resveratrol Pluronic®Micelles Prepared by Solvent Casting and Simple Equilibrium Methods

Background: Many different methods have been developed to incorporate hydrophobic compounds into polymeric micelles, including simple equilibrium (SE) and solvent casting (SC). Thus far, no studies have compared the SE and SC methods for drug loading and stability. Methods: Resveratrol (RES), a model hydrophobic compound, loaded in various Pluronics® (F88, F98, F108, and F127) is used to determine the differences between SE and SC methods. Results: Micelles prepared by SE method demonstrated RES loading in proportion to their respective CMC values with F127 having the lowest CMC and highest loading at Day 14 of 5.39±0.19 mg/mL, followed by F98 at 3.65±0.13 mg/mL, F108 at 3.29±0.06, and F88 at 2.35±0.05 mg/mL. Initial RES loading in SC method approaches 10 mg/mL in all the polymers, however, the micelles are unable to retain RES at this concentration for more than a few hours to a day. At the lower loading of 5 mg/mL a higher stability is seen in the Pluronic® micelles. In all micelles prepared by SC method, whether high or low loading, over a 14 day period the RES concentration in the micelles approached the equilibrium RES solubility. Conclusion: Our results demonstrate that micelles prepared by SC result in meta-stable micelles when the amount of the compound loaded exceeds the equilibrium loading and the rate at which the compound in the micelles approaches equilibrium solubility is determined by the hydrophobicity of the compound and the hydrophile- lipophile balance (HLB) of the polymer. Keywords: Drug loading and retention, micelle stability, pluronics®, polymeric micelles, resveratrol, simple equilibrium method, solvent casting method.

Nanosized Camptothecin Conjugates for Single and Combined Drug Delivery

Camptothecin (CPT), a natural DNA topoisomerase 1 inhibitor, is commonly used as a model hydrophobic compound for anticancer drug delivery research. Nanoparticle loaded with active drugs has the potential to selectively deliver the therapeutic agents into tumor cells without affecting the normal tissues, and hence is attracting tremendous attention from biomedical scientists. Nanoparticles can benefit from the enhanced permeability and retention effect (EPR) of the tumor tissues that allows nanoparticles to be transported through the leaky blood vessels and retained for a longer time compared with traditional small molecule drugs. In particular, covalent camptothecin conjugates are of extensive interest due to their robust stability by covalent chemical bonding. In this mini-review, recent progress of nanosized camptothecin conjugates will be discussed with the focus on the difference of delivery materials as well as their potential of combined drug delivery for the therapy of cancer.

A green method for the preparation of fluorescent hybrid structures of gold and corrole

Gold/soap nanostructures were prepared by a green methodology using saponified household sunflower oil, as reducing and organic dispersing agent of auric acid. The incorporation of hydrophobic molecules on the novel water-soluble gold nanoparticles was followed by fluorescence lifetime imaging microscopy, using as model hydrophobic compound 5,10,15-tris-(pentafluorophenyl)corrolatogallium(III)(pyridine) (GaPFC), a highly fluorescent corrole. The results showed the hydrophobic GaPFC located between the organic bilayer surrounding several Au nanoparticles, which in turn were coated with fatty acids salts anchored by the double bond at the gold’s surface.

Absorption and biodegradation of toluene: Optimization of its initial concentration and the biodegradable non-aqueous phase liquid volume fraction

Di (2-EthylHexyl) Phthalate (DEHP) was selected as a biodegradable organic solvent to be implemented in a two-phase partitioning bioreactor (TPPB) dedicated to remove a model hydrophobic volatile organic compound (VOC), toluene. In a first step, the absorption capacity of toluene in the selected organic solvent was examined according to the partition coefficients H. In a second step, toluene biodegradation in DEHP by non-acclimated activated sludge was carried out for different volume fractions of DEHP in water and five different toluene concentrations (4.3, 43, 106, 212 and 430 mg l−1). Toluene showed high affinity for DEHP with H = 0.99 Pa m3 mole−1. Both toluene and DEHP were completely consumed for 4.3 mg l−1 (initial toluene concentration) and a volume ratio of 0.1% DEHP in water. For an initial toluene concentration of 106 mg l−1 and a volume ratio of 0.1%, total toluene consumption and 87% DEHP degradation yield were obtained after seven days of incubation.

Passive dosing versus solvent spiking for controlling and maintaining hydrophobic organic compound exposure in the Microtox® assay

Microbial toxicity bioassays such as the Microtox® test are ubiquitously applied to measure the toxicity of chemicals and environmental samples. In many ways their operation is conducive to the testing of organic chemicals. They are of short duration, use glass cuvettes and take place at reduced temperatures in medium lacking sorbing components. All of these are expected to reduce sorptive and volatile losses, but particularly for hydrophobic organics the role of such losses in determining the bioassay response remains unclear. This study determined the response of the Microtox® test when using solvent spiking compared to passive dosing for introducing the model hydrophobic compounds acenaphthene, phenanthrene, fluoranthene and benzo(a)pyrene. Compared to solvent spiking, the apparent sensitivity of the Microtox® test with passive dosing was 3.4 and 12.4 times higher for acenaphthene and phenanthrene, respectively. Furthermore, fluoranthene only gave a consistent response with passive dosing. Benzo(a)pyrene did not result in a response with either spiking or passive dosing even at aqueous solubility. Such differences in the apparent sensitivity of the Microtox® test can be traced back to the precise definition of the dissolved exposure concentrations and the buffering of losses with passive dosing. This highlights the importance of exposure control even in simple and short-term microbial bioassays such as the Microtox® test.

PEGylated Liposomes as Carriers of Hydrophobic Porphyrins.

Sterically stabilized liposomes (SSLs) (PEGylated liposomes) are applied as effective drug delivery vehicles. Understanding the interactions between hydrophobic compounds and PEGylated membranes is therefore important to determine the effectiveness of PEGylated liposomes for delivery of drugs or other bioactive substances. In this study, we have combined fluorescence quenching analysis (FQA) experiments and all-atom molecular dynamics (MD) simulations to study the effect of membrane PEGylation on the location and orientation of 5,10,15,20-tetrakis(4-hydroxyphenyl)porphyrin (p-THPP) that has been used in our study as a model hydrophobic compound. First, we consider the properties of p-THPP in the presence of different fluid phosphatidylcholine bilayers that we use as model systems for protein-free cell membranes. Next, we studied the interaction between PEGylated membranes and p-THPP. Our MD simulation results indicated that the arrangement of p-THPP within zwitterionic membranes is dependent on their free volume, and p-THPP solubilized in PEGylated liposomes is localized in two preferred positions: deep within the membrane (close to the center of the bilayer) and in the outer PEG corona (p-THPP molecules being wrapped with the polymer chains). Fluorescence quenching methods confirmed the results of atomistic MD simulations and showed two populations of p-THPP molecules as in MD simulations. Our results provide both an explanation for the experimental observation that PEGylation improves the drug-loading efficiency of membranes and also a more detailed molecular-level description of the interactions between porphyrins and lipid membranes.

Light-responsive azobenzene-based glycopolymer micelles for targeted drug delivery to melanoma cells

Light-responsive glycopolymer micelles were produced by the self-assembly of amphiphilic block copolymers containing azobenzene and β-galactose units. These well-defined block copolymers were synthesised firstly by the RAFT polymerisation of an azobenzene methacrylate monomer (AzoMA) to produce two short azobenzene macroRAFT agents containing 7 and 15 monomer units. Chain extension with a second block of ∼150 or ∼250 sugar units comprising of a protected β-galactose monomer (β-AcGalEtMA) generated four block copolymers, which were converted to amphiphilic structures by deprotection of the acetyl groups on the sugar units. Micelles with well-defined sizes of 26–50nm were produced by self-assembly in water. The azobenzene units isomerised very rapidly to their more polar cis isomers under UV irradiation, reaching the photostationary state within 2min, with reversion to the trans state taking several hours in the dark. This transition to the more polar cis state is an important criteria for aiding expulsion of a hydrophobic payload. In cell studies, unloaded micelles showed low cytotoxicity, and micelles loaded with the model hydrophobic compound Nile red demonstrated high cellular uptake in human melanoma A375 cells, demonstrating their suitability as a potential drug delivery system for melanoma.

Immobilization of unspecific peroxygenases (EC 1.11.2.1) in PVA/PEG gel and hollow fiber modules

The immobilization of enzymes has many advantages, such as higher stability, easier handling, and reuse of the catalyst. Here we report, for the first time, two effective methods for the immobilization of unspecific peroxygenase (UPO; EC 1.11.2.1). This biocatalyst type comprises heavily glycosylated heme-thiolate proteins that catalyze various biotechnologically relevant oxyfunctionalizations. Both the encapsulation in cryogel and the retention of the enzyme in hollow fiber modules were found to be efficient methods for their immobilization. After encapsulation, the enzyme still exhibited 60% of its initial activity. Interestingly, we did not find differences in the kinetic parameters of free and immobilized UPOs. In long-term experiments, the conversion of the pharmaceutical diclofenac with immobilized UPOs in different reactor types yielded between 62mg and 154mg of the major human drug metabolite 4′-hydroxydiclofenac. The maximal total turnover number was about 60-fold higher compared to the free enzyme. A test over 5 months showed that storage of encapsulated UPOs in non-polar solvents (e.g., cyclohexane) helps to preserve the enzyme stability and increases their relative activity (by about ∼150%, in the case of diclofenac hydroxylation). In addition to the hydrophilic substrate diclofenac, encapsulated UPOs also oxidized the hydrophobic model compound cyclohexane.

Transpapillary drug delivery to the breast.

The study was aimed at investigating localized topical drug delivery to the breast via mammary papilla (nipple). 5-fluorouracil (5-FU) and estradiol (EST) were used as model hydrophilic and hydrophobic compounds respectively. Porcine and human nipple were used for in-vitro penetration studies. The removal of keratin plug enhanced the drug transport through the nipple. The drug penetration was significantly higher through the nipple compared to breast skin. The drug’s lipophilicity had a significant influence on drug penetration through nipple. The ducts in the nipple served as a major transport pathway to the underlying breast tissue. Results showed that porcine nipple could be a potential model for human nipple. The topical application of 5-FU on the rat nipple resulted in high drug concentration in the breast and minimal drug levels in plasma and other organs. Overall, the findings from this study demonstrate the feasibility of localized drug delivery to the breast through nipple.

Self‐Assembly of Amphiphilic Block Copolypeptoids with C2‐C5 Side Chains in Aqueous Solution

Nowadays, amphiphilic molecules play an important role in our life. In medical applications, amphiphilic block copolymers have attracted much attention as excipients in drug delivery systems. Here, the polymers are used as emulsifiers, micelles, or polymersomes with a hydrophilic corona block and a hydrophobic core or membrane. The aggregation behavior in aqueous solutions of a series of different amphiphilic block copolypeptoids comprising polysarcosine as a hydrophilic part is here reported. The formation of aggregates is investigated with 1H NMR spectroscopy and dynamic light scattering, and the determination of the critical micelle concentration (cmc) is performed using pyrene fluorescence spectroscopy. For the different block copolypeptoids cmc values ranging from 0.6 × 10−6m to 0.1 × 10−3m are found. The tendency to form micelles increases with increasing hydrophobicity at the nitrogen side chain in the hydrophobic moiety. Furthermore, in the case of the same hydrophobic side chain, a decreasing hydrophilic/lipophilic balance leads to the formation of larger aggregates. The aggregates formed in the buffer are able to solubilize the hydrophobic model compounds Reichardt's dye and pyrene, and exhibit versatile microenvironments. Final investigations about the cytotoxicity reveal that the block copolypeptoids are well tolerated by mammalian cells up to high concentrations. image

Novel biocompatible nanocapsules for slow release of fragrances on the human skin

There is a growing demand for fragranced products, but due to the poor aqueous solubility and instability of fragrance molecules, their use is limited. Nowadays, fragrance encapsulation in biocompatible nanocontainer material is emerging as a novel strategy to overcome the evaporation of volatile molecules and to prolong the sensory characteristics of fragrance molecules and the longevity of perfumes. The objective of this study was to develop an innovative sustained release system of perfume, by entrapping fragrance molecules in a polymeric nanocarrier; the impact of this strategy on the human axillary microbiome was further assessed. Stabilised poly-l-lactic acid nanocapsules (PLA-NCs) with a diameter of approximately 115 nm were prepared through nanoprecipitation. Size and morphology of the capsules were evaluated using Transmission Electron Microscopy (TEM) and Dynamic Light Scattering (DLS). Two model hydrophobic compounds, chlorobenzene and fluorescein, representing two different types of functionalised molecules, were encapsulated in PLA-NCs with an efficiency rate of 50%. Different release behaviours were seen, dependent on hydrophobicity. For hydrophobic compounds, a steady release was observed over 48hours. The polymeric nanocarriers did not impact the human axillary microbiome. Because of the slow and sustained release of fragrances, encapsulation of molecules in biocompatible NCs can represent a revolutionary contribution to the future of toiletries, body deodorant products, and in washing and cleaning sectors.

Toluene biodegradation in a solid/liquid system involving immobilized activated sludge and silicone oil as pollutant reservoir

A solid/liquid system involving activated sludge immobilized in an agar medium and a non-aqueous phase liquid containing the target pollutant has been considered to treat a model hydrophobic volatile organic compound, toluene. The positive impact of the use of a multiphase bioreactor is that the organic phase constitutes a pollutant reservoir and also helps to overcome possible pollutant toxicity. In addition and to overcome the drawbacks of the use of a solid organic phase (high pressure drop and low mass transfer) instead of a liquid organic phase, the considered solid phase was the aqueous. Consequently, silicone oil (polydimethylsiloxane) which showed its relevance for implementation in multiphase bioreactors was used. Promising results were observed from the analysis of toluene in the gaseous phase; for an initial amount of 2 g L−1 related to the organic phase, a v/v ratio of 0.5 of the organic phase to the aqueous agar phase, total toluene consumption was observed in about 9 days, leading to a global biodegradation rate of approximately 3.1 mg L−1 h−1, namely in the range of values previously observed in liquid/liquid systems.

PH-Sensitive Nanocapsules with Barrier Properties: Fragrance Encapsulation and Controlled Release

A facile synthesis method for polymer nanocapsules with high diffusion barrier and stimuli-responsive release properties is presented. The highly volatile fragrance α-pinene was used as hydrophobic model compound for the encapsulation process, which is based on a miniemulsion-analogous free radical polymerization process. The copolymer composition was systematically varied, and increasing contents of methacrylic acid as functional monomer in combination with high glass transition temperatures enabled unusually high encapsulation efficiencies of ≥90% for capsules with z-average diameters of <200 nm. Temperature and pH change can be used as trigger to open the capsules, and the release kinetics can be tailored depending on the polymer shell composition. In contrast to more frequently applied barrier microcapsules the nanocapsules provide drastically improved colloidal stabilities. Furthermore, the barrier nanocapsule approach is principally not restricted to fragrances and is expected to be compatible with other hydrophobic actives.

Synthesis via RAFT polymerization of thermo- and pH-responsive random copolymers containing cholic acid moieties and their self-assembly in water

Here we report the design and synthesis of polymers containing cholic acid pendant groups, a major bile acid synthesized in the liver from cholesterol. Random copolymers derived from 2-(methacryloyloxy)ethyl cholate (MAECA) with polyethylene glycol methyl ether methacrylate (PEGMA) and N,N-dimethylaminoethyl methacrylate (DMAEMA) have been synthesized by reversible addition–fragmentation chain transfer (RAFT) polymerization. Compositions of co-monomers in the copolymers were determined by 1H NMR spectroscopy, which were further used to determine reactivity ratios of the monomers using the extended Kelen–Tudos method at high conversions. Copolymers from MAECA with PEGMA (P(MAECA-co-PEGMA)) series displayed lower critical solution temperatures (LCSTs) up to 23% MAECA content in the copolymer, which was studied by UV-visible spectroscopy. Both thermo- and pH-responsiveness were observed for the P(MAECA-co-DMAEMA) copolymers with an MAECA unit in the copolymer up to 11.8%. The critical aggregation concentration (CAC) of water soluble copolymers was determined by fluorescence spectroscopy. Self-assembly of random copolymers was investigated by 1H NMR spectroscopy, dynamic light scattering (DLS), atomic force microscopy (AFM) and scanning electron microscopy (SEM), which indicated micellar structures in water from cholic acid based random copolymers. The polymeric aggregates were further explored for their drug encapsulation capabilities in aqueous medium using nile red as a hydrophobic model compound.

Formation and characterization of quercetin-loaded chitosan oligosaccharide/β-lactoglobulin nanoparticle

Food grade materials including chitosan and β-lactoglobulin need to be developed for the application of nanoparticles to food science. It is hypothesized that hydrophobicity of chitosan oligosaccharide and β-lactoglobulin may be a key factor to affect the physicochemical properties of nanoparticle. The aim of this study was to investigate how substitution of chitosan oligosaccharide with linoleic acid (LA) and sub-ambient temperature treatment, which can influence hydrophobicity, affect the formation and physicochemical properties of LA-modified chitosan oligosaccharide/β-lactoglobulin (CSO-LA/β-lg) nanoparticles. CSO-LAs with different charged amounts of LA (0, 10, 20, and 30%) and β-lg mixtures at pH4.4 were sub-ambient temperature treated at 5, 10, 15, and 20°C for 30min followed by treatment with 0.5mM tripolyphosphate. Quercetin was used as a model hydrophobic bioactive compound. Spherical shape nanoparticles in the range of 170 and 350nm were successfully formed. An increase in charged amount of LA and temperature resulted in an increase in the size of CSO-LA/β-lg nanoparticles. Increasing charged amount of LA and decreasing sub-ambient temperature treatment led to an increase in the association efficiency of CSO and β-lg, respectively. Encapsulation efficiency of quercetin was enhanced with increased charged amount of LA and decreased temperature. It was concluded that charged amounts of LA and sub-ambient temperature were major key-factors affecting the size of nanoparticles and encapsulation efficiency of quercetin.

Facile synthesis of metallized nanocontainers for the entrapment and controlled release of hydrophobic compounds

Template assisted synthesis of metallized nanocontainers (MNCs) is reported by in situ synthesis of gold nanoparticles (GNPs) in the bilayer of soft lecithin micelles using aminoantipyrene as a weak reducing agent. The exact bilayer mimicry of MNCs was confirmed by electron microscopic (SEM and TEM) characterization. These MNCs were further coated with thiolated polyethylene glycol (PEG-SH) to improve their stability in aqueous media. The solubility of MNCs in polar and non-polar solvents can be tuned simply by varying their pH values. These MNCs were then used for the entrapment and controlled release of a model hydrophobic compound, pyrene, to demonstrate their potential applications for the controlled release of hydrophobic drugs.

Polypeptide-based star-block quadripolymers as unimolecular nanocarriers for the simultaneous encapsulation of hydrophobic and hydrophilic guests

Star-block quadripolymers PEI-g-(PLF-b-PLL-b-PEG) and PEI-g-(PLF-b-PLG-b-PEG) [i.e., a polyethylenimine (PEI) core, an amphiphilic copolypeptide poly(l-phenylalanine)-b-poly(l-lysine) (PLF-b-PLL) or poly(l-phenylalanine)-b-poly(l-glutamic acid) (PLF-b-PLG) inner shell, and a poly(ethylene glycol) (PEG) outer shell] were synthesized, characterized, and evaluated as drug nanocarriers. The star-block quadripolymers were obtained by sequential ring-opening polymerizations of l-phenylalanine N-carboxyanhydride and ε-benzyloxycarbonyl-l-lysine N-carboxyanhydride or γ-benzyl-l-glutamate N-carboxyanhydride initiated by the terminal primary amines of PEI. Subsequently, the periphery was PEGylated, and the poly(l-lysine) or poly(l-glutamic acid) side chains were deprotected. The synthesized star-block quadripolymers were characterized with 1H NMR, gel permeation chromatography (GPC), and laser light dynamic scattering (DLS). These polymers were well dispersed in aqueous solutions and resembled amphiphilic unimolecular micelles. The encapsulation study demonstrated that these polymers can solubilize nonpolar model compounds through hydrophobic interactions. Dialysis and spectrophotometric titration experiments indicated that these polymers could efficiently encapsulate hydrophilic model compounds via electrostatic interactions. Furthermore, the synthesized quadripolymers could entrap hydrophobic and hydrophilic model compounds in the site-isolated state simultaneously. The entrapped hydrophilic model compounds demonstrated sustained release at physiological pH and accelerated release when the pH was either increased or decreased. The simultaneous encapsulation of versatile guest molecules as well as the pH-responsive releasing properties of these star-block quadripolymers could be potentially useful in the controlled drug co-delivery applications.

Influence of Formulation Factors on the Preparation of Zein Nanoparticles

The main objective of the present study was to investigate the influence of various formulation parameters on the preparation of zein nanoparticles. 6,7-dihydroxycoumarin (DHC) was used as a model hydrophobic compound. The influence of pH of the aqueous phase, buffer type, ionic strength, surfactant, and zein concentration on particle size, polydispersity index, and zeta potential of DHC-loaded zein nanoparticles were studied. Smaller nanoparticles were formed when the pH was close to the isoelectric point of zein. DHC-loaded zein nanoparticles prepared using citrate buffer (pH 7.4) was better than phosphate buffer in preventing particle aggregation during lyophilization. The ionic strength did not have a significant influence on the particle size of DHC-loaded zein nanoparticles. A combination of Pluronic F68 and lecithin in 2:1 ratio stabilized the zein nanoparticles. An increase in zein concentration led to increase in particle size of DHC-loaded zein nanoparticles. The use of optimal conditions produced DHC-loaded nanoparticles of 256 ± 30 nm and an encapsulation efficiency of 78 ± 7%. Overall, the study demonstrated the optimal conditions to prepare zein nanoparticles for drug encapsulation.

Tuning the sugar‐response of boronic acid block copolymers

AbstractA detailed study of the pH‐ and sugar‐responsive behavior of poly(3‐acrylamidophenylboronic acid pinacol ester)‐b‐poly(N,N‐dimethylacrylamide) (PAPBAE‐b‐PDMA) block copolymers is presented. Reversible addition‐fragmentation chain transfer (RAFT) polymerization of the pinacol ester of 3‐acrylamidophenylboronic acid resulted in homopolymers with molecular weights between 12,000 and 37,000 g/mol. The resulting homopolymers were employed as macro‐chain transfer agents during the polymerization of N,N‐dimethylacrylamide (DMA). Successful chain extension and removal of the pinacol protecting groups to yield poly(3‐acrylamidophenylboronic acid)‐b‐PDMA (PAPBA‐b‐PDMA) with free boronic acid moieties resulted in pH‐ and sugar‐responsive block copolymers that were subsequently investigated for their behavior in aqueous solution. The PAPBA‐b‐PDMA block copolymers were capable of solution self‐assembly due to the PAPBA block being water‐insoluble below its pKa. The resulting aggregates were demonstrated to solubilize and release model hydrophobic compounds, as demonstrated by fluorescence studies. Dissociation of the aggregates was induced by raising the pH above the pKa of the boronic acid residues or by adding sugars capable of forming boronate esters. Aggregate size, dissociation kinetics, and the effect of various sugars were considered. The critical sugar concentration needed to induce aggregate dissociation was tuned by incorporation of hydrophilic DMA units within the PAPBA responsive segment to yield PDMA‐b‐poly(3‐acrylamidophenylboronic acid‐co‐DMA) block copolymers. © 2012 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2012

Yeast cells as microcapsules. Analytical tools and process variables in the encapsulation of hydrophobes in S. cerevisiae

Yeast cells can be used as biocompatible and biodegradable containers for the microencapsulation of a variety of actives. Despite the wide application of this process, e.g. in the food industry, mechanism and controlling factors are yet poorly known. In this study we have studied kinetics and mechanistic aspects of the spontaneous internalization of terpenes (as model hydrophobic compounds) in Saccharomyces cerevisiae, quantifying their encapsulation through HPLC analysis and fluorescent staining of lipidic bodies with Nile Red, while in parallel monitoring cell viability. Our results showed that this encapsulation process is essentially a phenomenon of passive diffusion with negligible relevance of active transport. Further, our evidence shows that the major determinant of the encapsulation kinetics is the solubility of the hydrophobe in the cell wall, which is inversely related to partition coefficient (log P).