Effects of Alkaline Extraction pH on Amino Acid Compositions, Protein Secondary Structures, Thermal Stability, and Functionalities of Brewer's Spent Grain Proteins.

A high alkaline pH was previously demonstrated to enhance the extraction yield of brewer's spent grains (BSG) proteins. The effects of extraction pH beyond the extraction yield, however, has not been investigated before. The present work examined the effects of extraction pH (pH 8-12) on BSG proteins' (1) amino acid compositions, (2) secondary structures, (3) thermal stability, and (4) functionalities (i.e., water/oil holding capacity, emulsifying, and foaming properties). The ideal extraction temperature (60 °C) and BSG-to-solvent ratio (1:20 w/v) for maximizing the extraction yield were first determined to set the conditions for the pH effect study. The results showed that a higher extraction pH led to more balanced compositions between hydrophilic and hydrophobic amino acids and higher proportions of random coils structures indicating increased protein unfolding. This led to superior emulsifying properties of the extracted proteins with more than twofold improvement between pH 8 and a pH larger than 10. The extraction pH, nevertheless, had minimal impact on the water/oil holding capacity, foaming properties, and thermal denaturation propensity of the proteins. The present work demonstrated that a high alkaline pH at pH 11-12 was indeed ideal for both maximizing the extraction yield (37-46 wt.%) and proteins' functionalities.

Mechanical Properties of 3-Hydroxybutyric Acid-Induced Vesicles.

The vesicle mechanical behaviors were studied upon its exposure to 3-hydroxybutyric acid using an atomic force microscope (AFM). Dipalmitoylphosphatidylcholine (DPPC) and 3-hydroxybutyric acid were used to manufacture the vesicles at their desired ratio. The deflection of an AFM probe with respect to its displacement was measured after characterizing the vesicle adsorption. The movement was analyzed with the Hertzian model to understand the physical behavior of the vesicles. However, in the deflection just prior to the first penetration, the model was a good fit, and the vesicle mechanical moduli were calculated. The moduli became lower with the higher ratio of 3-hydroxybutyric acid to DPPC, but the moduli were saturated at 0.5 of the ratio. These results appear to be the basis for the function of the metabolism associated with 3-hydroxybutyric acid, i.e., anesthetization and glycemic control, on the physical properties of cell membranes.

Deep Eutectic Solvent as Green Solvent in Extraction of Biological Macromolecules: A Review.

Greater awareness of environmental sustainability has driven many industries to transition from using synthetic organic solvents to greener solvents in their manufacturing. Deep eutectic solvents (DESs) have emerged as a highly promising category of green solvents with well-demonstrated and wide-ranging applications, including their use as a solvent in extraction of small-molecule bioactive compounds for food and pharmaceutical applications. The use of DES as an extraction solvent of biological macromolecules, on the other hand, has not been as extensively studied. Thereby, the feasibility of employing DES for biomacromolecule extraction has not been well elucidated. To bridge this gap, this review provides an overview of DES with an emphasis on its unique physicochemical properties that make it an attractive green solvent (e.g., non-toxicity, biodegradability, ease of preparation, renewable, tailorable properties). Recent advances in DES extraction of three classes of biomacromolecules-i.e., proteins, carbohydrates, and lipids-were discussed and future research needs were identified. The importance of DES's properties-particularly its viscosity, polarity, molar ratio of DES components, and water addition-on the DES extraction's performance were discussed. Not unlike the findings from DES extraction of bioactive small molecules, DES extraction of biomacromolecules was concluded to be generally superior to extraction using synthetic organic solvents.

Lyophilization of Curcumin-Albumin Nanoplex with Sucrose as Cryoprotectant: Aqueous Reconstitution, Dissolution, Kinetic Solubility, and Physicochemical Stability.

An amorphous curcumin (CUR) and bovine serum albumin (BSA) nanoparticle complex (nanoplex) was previously developed as a promising anticancer nanotherapy. The CUR-BSA nanoplex had been characterized in its aqueous suspension form. The present work developed a dry-powder form of the CUR-BSA nanoplex by lyophilization using sucrose as a cryoprotectant. The cryoprotective activity of sucrose was examined at sucrose mass fractions of 33.33, 50.00, and 66.66% by evaluating the lyophilized nanoplex's (1) aqueous reconstitution and (2) CUR dissolution and kinetic solubility. The physicochemical stabilizing effects of sucrose upon the nanoplex's 30-day exposures to 40 °C and 75% relative humidity were examined from (i) aqueous reconstitution, (ii) CUR dissolution, (iii) CUR and BSA payloads, (iv) amorphous form stability, and (v) BSA's structural integrity. The good cryoprotective activity of sucrose was evidenced by the preserved BSA's integrity and good aqueous reconstitution, resulting in a fast CUR dissolution rate and a high kinetic solubility (≈5-9× thermodynamic solubility), similar to the nanoplex suspension. While the aqueous reconstitution, CUR dissolution, and amorphous form were minimally affected by the elevated heat and humidity exposures, the treated nanoplex exhibited a lower BSA payload (≈7-26% loss) and increased protein aggregation postexposure. The adverse effects on the BSA payload and aggregation were minimized at higher sucrose mass fractions.

A Potential Quorum-Sensing Inhibitor for Bronchiectasis Therapy: Quercetin-Chitosan Nanoparticle Complex Exhibiting Superior Inhibition of Biofilm Formation and Swimming Motility of Pseudomonas aeruginosa to the Native Quercetin.

Quercetin (QUE)-a plant-derived flavonoid, is recently established as an effective quorum sensing (QS) inhibiting agent in Pseudomonas aeruginosa-the main bacterial pathogen in bronchiectasis lungs. Successful clinical application of QUE, however, is hindered by its low solubility in physiological fluids. Herein we developed a solubility enhancement strategy of QUE in the form of a stable amorphous nanoparticle complex (nanoplex) of QUE and chitosan (CHI), which was prepared by electrostatically driven complexation between ionized QUE molecules and oppositely charged CHI. At its optimal preparation condition, the QUE-CHI nanoplex exhibited a size of roughly 150 nm with a 25% QUE payload and 60% complexation efficiency. The complexation with CHI had no adverse effect on the antibacterial and anticancer activities of QUE, signifying the preservation of QUE's bioactivities in the nanoplex. Compared to the native QUE, the QUE-CHI nanoplex exhibited superior QS inhibition in suppressing the QS-regulated swimming motility and biofilm formation of P. aeruginosa, but not in suppressing the virulence factor production. The superior inhibitions of the biofilm formation and swimming motility afforded by the nanoplex were attributed to (1) its higher kinetic solubility (5-times higher) that led to higher QUE exposures, and (2) the synergistic QS inhibition attributed to its CHI fraction.

High-Payload Buccal Delivery System of Amorphous Curcumin-Chitosan Nanoparticle Complex in Hydroxypropyl Methylcellulose and Starch Films.

Oral delivery of curcumin (CUR) has limited effectiveness due to CUR's poor systemic bioavailability caused by its first-pass metabolism and low solubility. Buccal delivery of CUR nanoparticles can address the poor bioavailability issue by virtue of avoidance of first-pass metabolism and solubility enhancement afforded by CUR nanoparticles. Buccal film delivery of drug nanoparticles, nevertheless, has been limited to low drug payload. Herein, we evaluated the feasibilities of three mucoadhesive polysaccharides, i.e., hydroxypropyl methylcellulose (HPMC), starch, and hydroxypropyl starch as buccal films of amorphous CUR-chitosan nanoplex at high CUR payload. Both HPMC and starch films could accommodate high CUR payload without adverse effects on the films' characteristics. Starch films exhibited far superior CUR release profiles at high CUR payload as the faster disintegration time of starch films lowered the precipitation propensity of the highly supersaturated CUR concentration generated by the nanoplex. Compared to unmodified starch, hydroxypropyl starch films exhibited superior CUR release, with sustained release of nearly 100% of the CUR payload in 4 h. Hydroxypropyl starch films also exhibited good payload uniformity, minimal weight/thickness variations, high folding endurance, and good long-term storage stability. The present results established hydroxypropyl starch as the suitable mucoadhesive polysaccharide for high-payload buccal film applications.

Proof-of-concept preparation and characterization of dual-drug amorphous nanoparticle complex as fixed-dose combination of poorly soluble drugs.

OBJECTIVES: To carry out a proof-of-concept study on the development of dual-drug amorphous nanoparticle complex (nanoplex in short) as a potential formulation platform for fixed-dose combination (FDC) of poorly-soluble drugs. SIGNIFICANCE: FDC has been proven effective in improving patient compliance for treatment that requires complex multidrug regimen. Currently, there is growing interest to develop FDC of poorly-soluble drugs due to the increased number of drugs exhibiting poor solubility thus low bioavailability. METHODS: The dual-drug nanoplex was prepared by electrostatically-driven co-complexation of drug molecules with oppositely charged dextran sulfate, using ciprofloxacin (CIP) and itraconazole (ITZ) as the model poorly-soluble drugs. RESULTS: We first verified that the co-complexation products were dual-drug CIP-ITZ nanoplex, and not binary mixtures of the single-drug CIP and ITZ nanoplexes, by demonstrating their distinct thermal behaviors and dissolution characteristics. Depending on the preparation condition, the dual-drug nanoplex exhibited size and zeta potential of 160-410 nm and -35-50 mV, respectively. The individual drug payloads were readily manipulated by varying the CIP/ITZ mass ratio in the feed, resulting in CIP and ITZ payloads in the range of 60-30% and 15-45%, respectively. The CIP-ITZ nanoplex, however, exhibited diminished CIP supersaturation generation, thus lower CIP solubility enhancement, compared to the single-drug CIP nanoplex. The CIP-ITZ nanoplex, nonetheless, remained capable of generating high ITZ supersaturation level. CONCLUSION: Dual-drug nanoplex was successfully prepared with a high degree of control over its physical characteristics. Nevertheless, whether dual-drug nanoplex always exhibits diminished solubility enhancement compared to its single-drug counterparts needs to be investigated using different poorly-soluble drugs.

Effects of chitosan molecular weight on the physical and dissolution characteristics of amorphous curcumin-chitosan nanoparticle complex.

OBJECTIVE: To investigate the effects of varying molecular weight (MW) of chitosan (CHI) used in the complexation with curcumin (CUR) on the physical and dissolution characteristics of the amorphous CUR-CHI nanoparticle complex produced. SIGNIFICANCE: Amorphous CUR-CHI nanoparticle complex (or CUR nanoplex in short) recently emerged as a promising bioavailability enhancement strategy of CUR attributed to its fast dissolution, supersaturation generation capability, and simple preparation. Existing CUR nanoplex prepared using low MW CHI, however, exhibited poor colloidal stability during storage. Herein we hypothesized that the colloidal stability could be improved by using CHI of higher MW. The effects of this approach on the nanoplex's other characteristics were simultaneously investigated. METHODS: The CUR nanoplex was prepared by electrostatically driven self-assembled complexation between CUR and oppositely charged CHI of three different MWs (i.e. low, medium, and high). Besides colloidal stability, the effects of MW variation were investigated for the nanoplex's (1) other physical characteristics (i.e. size, zeta potential, CUR payload, amorphous state stability), (2) preparation efficiency (i.e. CUR utilization rate, yield), and (3) dissolutions under sink condition and supersaturation generation. RESULTS: CUR nanoplex prepared using CHI of high MW exhibited improved colloidal stability, larger size, superior morphology, and prolonged supersaturation generation. On the other hand, the effects of MW variation on the payload, amorphous state stability, preparation efficiency, and dissolution under sink condition were found to be insignificant. CONCLUSIONS: Varying MW of CHI used was an effective means to improve certain aspects of the CUR nanoplex characteristics with minimal adverse effects on the others.

Complexation of Polyelectrolytes with Hydrophobic Drug Molecules in Salt-Free Solution: Theory and Simulations.

The delivery and dissolution of poorly soluble drugs is challenging in the pharmaceutical industry. One way to significantly improve the delivery efficiency is to incorporate these hydrophobic small molecules into a colloidal polyelectrolyes(PE)-drug complex in their ionized states. Despite its huge application value, the general mechanism of PE collapse and complex formation in this system has not been well understood. In this work, by combining a mean-field theory with extensive molecular simulations, we unveil the phase behaviors of the system under dilute and salt-free conditions. We find that the complexation is a first-order-like phase transition triggered by the hydrophobic attraction between the drug molecules. Importantly, the valence ratio between the drug molecule and PE monomer plays a crucial role in determining the stability and morphology of the complex. Moreover, the sign of the zeta potential and the net charge of the complex are found to be inverted as the hydrophobicity of the drug molecules increases. Both theory and simulation indicate that the complexation point and complex morphology and the electrostatic properties of the complex have a weak dependence on chain length. Finally, the dynamics aspect of PE-drug complexation is also explored, and it is found that the complex can be trapped into a nonequilibrium glasslike state when the hydropobicity of the drug molecule is too strong. Our work gives a clear physical picture behind the PE-drug complexation phenomenon and provides guidelines to fabricate the colloidal PE-drug complex with the desired physical characteristics.

Amorphous nanoparticle complex of perphenazine and dextran sulfate as a new solubility enhancement strategy of antipsychotic perphenazine.

OBJECTIVE: The objective of this study is to develop a new solubility enhancement strategy of antipsychotic drug - perphenazine (PPZ) - in the form of its amorphous nanoparticle complex (or nanoplex) with polyelectrolyte dextran sulfate (DXT). SIGNIFICANCE: Poor bioavailability of PPZ necessitated the development of fast-dissolving PPZ formulations regardless of delivery routes. Existing fast-dissolving formulations, however, exhibited low PPZ payload. The high-payload PPZ-DXT nanoplex represents an attractive fast-dissolving formulation, as dissolution rate is known to be proportional to payload. METHODS: The nanoplex was prepared by electrostatically driven complexation between PPZ and DXT in a simple process that involved only ambient mixing of PPZ and DXT solutions. We investigated the effects of key variables in drug-polyelectrolyte complexation (i.e. pH and charge ratio RDXT/PPZ) on the physical characteristics and preparation efficiency of the nanoplex produced. Subsequently, we characterized the colloidal and amorphous state stabilities, dissolution enhancement, and supersaturation generation of the nanoplex prepared at the optimal condition. RESULTS: The physical characteristics of nanoplex were governed by RDXT/PPZ, while the preparation efficiency was governed by the preparation pH. Nanoplex having size of ≈80 nm, zeta potential of ≈(-) 60 mV, and payload of ≈70% (w/w) were prepared at nearly 90% PPZ utilization rate and ≈60% yield. The nanoplex exhibited superior dissolution than native PPZ in simulated intestinal juice, resulting in high and prolonged apparent solubility with good storage stabilities. CONCLUSIONS: The simple yet efficient preparation, excellent physical characteristics, fast dissolution, and high apparent solubility exhibited by the PPZ-DXT nanoplex established its potential as a new bioavailability enhancement strategy of PPZ.

Amorphization strategy affects the stability and supersaturation profile of amorphous drug nanoparticles.

Amorphous drug nanoparticles have recently emerged as a promising bioavailability enhancement strategy of poorly soluble drugs attributed to the high supersaturation solubility generated by the amorphous state and fast dissolution afforded by the nanoparticles. Herein we examine the effects of two amorphization strategies in the nanoscale, i.e., (1) molecular mobility restrictions and (2) high energy surface occupation, both by polymer excipient stabilizers, on the (i) morphology, (ii) colloidal stability, (iii) drug loading, (iv) amorphous state stability after three-month storage, and (v) in vitro supersaturation profiles, using itraconazole (ITZ) as the model drug. Drug-polyelectrolyte complexation is employed in the first strategy to prepare amorphous ITZ nanoparticles using dextran sulfate as the polyelectrolyte (ITZ nanoplex), while the second strategy employs pH-shift precipitation using hydroxypropylmethylcellulose as the surface stabilizer (nano-ITZ), with both strategies resulting in >90% ITZ utilization. Both amorphous ITZ nanoparticles share similar morphology (∼300 nm spheres) with the ITZ nanoplex exhibiting better colloidal stability, albeit at lower ITZ loading (65% versus 94%), due to the larger stabilizer amount used. The ITZ nanoplex also exhibits superior amorphous state stability, attributed to the ITZ molecular mobility restriction by electrostatic complexation with dextran sulfate. The higher stability, however, is obtained at the expense of slower supersaturation generation, which is maintained over a prolonged period, compared to the nano-ITZ. The present results signify the importance of selecting the optimal amorphization strategy, in addition to formulating the excipient stabilizers, to produce amorphous drug nanoparticles having the desired characteristics.

Biofilm-like Lactobacillus rhamnosus probiotics encapsulated in alginate and carrageenan microcapsules exhibiting enhanced thermotolerance and freeze-drying resistance.

Microcapsules containing high-density biofilm-like Lactobacillus rhamnosus probiotics, in place of planktonic cells, are developed in order to enhance the cell viability upon exposures to stresses commonly encountered during food lifecycle (i.e., heating, freeze-drying, refrigerated storage, and acid). The high-density (HD) capsules are prepared by in situ cultivation of the planktonic cells in the confined space of polysaccharide-based capsules (i.e., chitosan-coated alginate and carrageenan capsules). Compared to their planktonic counterparts, the HD capsules exhibit higher freeze-drying resistance (~40×) and higher thermotolerance upon prolonged wet heat exposures at 60 and 70 °C (~12-8000×), but not at higher temperatures even for short exposures (i.e., 80 and 100 °C). The enhanced viability of the HD capsules, however, is not observed during the refrigerated storage and exposure to the simulated gastric juice. The alginate capsules are superior to carrageenan owed to their better cell release profile in the simulated intestinal juice and storage viability.

Binding Behavior between Transforming-Growth-Factor-Beta1 and Its Receptor Reconstituted in Biomimetic Membranes.

Transforming growth factor ß1 (TGF-ß1) is critical to cell differentiation, proliferation, and apoptosis. It is important to understand the binding affinity between TGF-ß1 and its receptors. In this study, their binding force was measured using an atomic force microscope. Significant adhesion was induced by the interaction between the TGF-ß1 immobilized on the tip and its receptor reconstituted in the bilayer. Rupture and adhesive failure occurred at a specific force around 0.4~0.5 nN. The relationship of the force to loading rate was used to estimate the displacement where the rupture occurred. The binding was also monitored in real time with surface plasmon resonance (SPR) and interpreted with kinetics to acquire the rate constant. Using the Langmuir adsorption, the SPR data were analyzed to estimate equilibrium and association constants to be approximately 107 M-1 and 106 M-1 s-1. These results indicated that the natural release of the binding seldom occurred. Furthermore, the degree of binding dissociation, confirmed by the rupture interpretation, supported that the reverse of the binding hardly happened.

Green amorphous nanoplex as a new supersaturating drug delivery system.

The nanoscale formulation of amorphous drugs represents a highly viable supersaturating drug-delivery system for enhancing the bioavailability of poorly soluble drugs. Herein we present a new formulation of a nanoscale amorphous drug in the form of a drug-polyelectrolyte nanoparticle complex (or nanoplex), where the nanoplex is held together by the combination of a drug-polyelectrolyte electrostatic interaction and an interdrug hydrophobic interaction. The nanoplex is prepared by a truly simple, green process that involves the ambient mixing of drug and polyelectrolyte (PE) solutions in the presence of salt. Nanoplexes of poorly soluble acidic (i.e., ibuprofen and curcumin) and basic (i.e., ciprofloxacin) drugs are successfully prepared using biocompatible poly(allylamine hydrochloride) and dextran sulfate as the PE, respectively. The roles of salt, drug, and PE in nanoplex formation are examined from ternary phase diagrams of the drug-PE complex, from which the importance of the drug's charge density and hydrophobicity, as well as the PE ionization at different pH values, is recognized. Under the optimal conditions, the three nanoplexes exhibit high drug loadings of ~80-85% owing to the high drug complexation efficiency (~90-96%), which is achieved by keeping the feed charge ratio of the drug to PE below unity (i.e., excess PE). The nanoplex sizes are ~300-500 nm depending on the drug hydrophobicity. The nanoplex powders remain amorphous after 1 month of storage, indicating the high stability owed to the PE's high glass-transition temperature. FT-IR analysis shows that functional groups of the drug are conserved upon complexation. The nanoplexes are capable of generating prolonged supersaturation upon dissolution with precipitation inhibitors. The supersaturation level depends on the saturation solubility of the native drugs, where the lower the saturation solubility, the higher the supersaturation level. The solubility of curcumin as the least-soluble drug is magnified 9-fold upon its transformation to the nanoplex, and the supersaturated condition is maintained for 5 h.

Benchmarking the Solubility Enhancement and Storage Stability of Amorphous Drug-Polyelectrolyte Nanoplex against Co-Amorphous Formulation of the Same Drug.

Amorphization, typically in the form of amorphous solid dispersion (ASD), represents a well-established solubility enhancement strategy for poorly soluble drugs. Recently, two amorphous drug formulations, i.e., the amorphous drug-polyelectrolyte nanoparticle complex (nanoplex) and co-amorphous system, have emerged as promising alternatives to circumvent the issues faced by ASD (i.e., large dosage requirement, high hygroscopicity). In the present work, the nanoplex was benchmarked against the co-amorphous system in terms of the preparation efficiency, drug payload, thermal stability, dissolution rate, supersaturation generation, and accelerated storage stability. Weakly acidic curcumin (CUR) and weakly basic ciprofloxacin (CIP) were used as the model poorly soluble drugs. The CUR and CIP nanoplexes were prepared using chitosan and sodium dextran sulfate as the polyelectrolytes, respectively. The co-amorphous CUR and CIP were prepared using tannic acid and tryptophan as the co-formers, respectively. The benchmarking results showed that the amorphous drug nanoplex performed as well as, if not better than, the co-amorphous system depending on the drug in question and the aspects being compared. The present work successfully established the nanoplex as an equally viable amorphous drug formulation as the more widely studied co-amorphous system to potentially serve as an alternative to ASD.

Changes in Mechanical Properties of Vesicles by Mucin in Aqueous Solution.

The mechanical properties of vesicles were investigated as they were prepared, according to the ratio of mucin to dipalmitoylphosphatidylcholine (DPPC), using an atomic force microscope (AFM). After the confirmation of the vesicle adsorption on a mica surface, an AFM-tip deflection, caused by the interaction between the tip and the vesicle, was measured. The deflection showed that the tip broke through into the vesicle twice. Each break meant a tip-penetration into the upper and lower portion of the vesicle. Only the first penetration allowed the Hertzian model available to estimate the vesicle mechanical moduli. Two moduli reduced as the ratio of mucin to DPPC increased to 0.5, but the moduli were little changed above the 0.5 ratio. These results seem to be a platform for the effect of the mucin on the plasma-membrane anchoring and cellular signaling.

Formulation Strategies to Improve the Stability and Handling of Oral Solid Dosage Forms of Highly Hygroscopic Pharmaceuticals and Nutraceuticals.

Highly hygroscopic pharmaceutical and nutraceutical solids are prone to significant changes in their physicochemical properties due to chemical degradation and/or solid-state transition, resulting in adverse effects on their therapeutic performances and shelf life. Moisture absorption also leads to excessive wetting of the solids, causing their difficult handling during manufacturing. In this review, four formulation strategies that have been employed to tackle hygroscopicity issues in oral solid dosage forms of pharmaceuticals/nutraceuticals were discussed. The four strategies are (1) film coating, (2) encapsulation by spray drying or coacervation, (3) co-processing with excipients, and (4) crystal engineering by co-crystallization. Film coating and encapsulation work by acting as barriers between the hygroscopic active ingredients in the core and the environment, whereas co-processing with excipients works mainly by adding excipients that deflect moisture away from the active ingredients. Co-crystallization works by altering the crystal packing arrangements by introducing stabilizing co-formers. For hygroscopic pharmaceuticals, coating and co-crystallization are the most commonly employed strategies, whereas coating and encapsulation are popular for hygroscopic nutraceuticals (e.g., medicinal herbs, protein hydrolysates). Encapsulation is rarely applied on hygroscopic pharmaceuticals, just as co-crystallization is rarely used for hygroscopic nutraceuticals. Therefore, there is potential for improved hygroscopicity reduction by exploring beyond the traditionally used strategy.

Assessing the impact of counterion types on the sustained release characteristics of high-payload drug-ion complex: A case study on tetracycline hydrochloride.

Complexation of ionized hydrophilic drugs with counterions (e.g. polyelectrolytes, ionic amphiphiles, multivalent salt ions) represents a well-established formulation approach to produce sustained release of highly soluble drugs while maintaining a high drug payload. This renders the drug-ion complex an attractive alternative to the conventional polymer matrix systems. The effects of the counterion's type on the sustained release characteristics of drug-ion complexes, however, have not been investigated before under the same dissolution environment. Using antibiotic tetracycline hydrochloride (TC•HCl) as the model hydrophilic drug, we investigated the effects of three types of counterions, sodium dextran sulfate (DXT), sodium dodecyl sulfate (SDS), and K2HPO4, on (1) the sustained release characteristics, (2) long-term storage stability, (3) preparation efficiency (i.e. yield, payload), and (4) antibiotic activity of the resultant (TC•HCl)-ion complexes. The results showed that the three complexes exhibited comparable TC•HCl payloads at approximately 80% (w/w) and yield between 40 and 60% (w/w). They also exhibited good storage stability after 18 months and uncompromised antibiotic activity compared to the native drug. In the intestinal fluid, all three complexes could produce sustained drug release profiles, albeit at different rates ((TC•HCl)-DXT > (TC•HCl)-SDS > (TC•HCl)-HPO4), whereas in the gastric fluid, only the (TC•HCl)-DXT complex could produce a sustained release profile suitable for oral delivery. The different sustained release profiles among the complexes were attributed to their different solid forms (amorphous versus crystalline), hydrophobicity, solubility, and drug release mechanisms. The present work highlighted the importance of selecting the most suitable counterion to achieve the desired sustained drug release profile.

Antibacterial efficacy of inhalable levofloxacin-loaded polymeric nanoparticles against E. coli biofilm cells: the effect of antibiotic release profile.

PURPOSE: To investigate the effect of the antibiotic release profiles of levofloxacin-loaded polymeric nanoparticles on their antibacterial efficacy against E. coli biofilm cells. METHODS: Three distinct antibiotic release profiles are produced by encapsulating levofloxacin in PCL and PLGA nanoparticles by nanoprecipitation and emulsification-solvent-evaporation methods. The antibacterial efficacy is examined over six days by time-dependent biofilm susceptibility testing that takes into account the effects of the biofilm age, antibiotic exposure history, and simulated drug removal. RESULTS: Biofilm cells that survive the initial antibiotic exposure exhibit a higher antibiotic tolerance than fresh biofilm cells, where the lower the initial exposure, the higher the tolerance of the surviving biofilm cells. The lower antibiotic susceptibility of the surviving biofilm cells is transferred to their planktonic cell progeny, which can subsequently form new biofilm colonies having a higher antibiotic tolerance, hence exacerbating the infections. A biphasic extended release profile at an appropriate dose can inhibit the biofilm growth for four days, therefore reducing the dosing frequency. The importance of a high initial antibiotic exposure renders a slow release profile ineffective despite the same dosing amount. CONCLUSIONS: The antibiotic release profile has an equally significant influence on the biofilm eradication rate as the antibiotic dose.

Ternary nanoparticle complex of antibiotic, polyelectrolyte, and mucolytic enzyme as a potential antibiotic delivery system in bronchiectasis therapy.

Antibiotic-polyelectrolyte nanoparticle complex (or nanoplex in short) has been recently demonstrated as a superior antibiotic delivery system to the native antibiotic in bronchiectasis therapy owed to its ability to overcome the lung's mucus barrier and generate high localized antibiotic exposure in the infected sites. The present work aimed to further improve the mucus permeability, hence the antibacterial efficacy of the nanoplex, by incorporating mucolytic enzyme papain (PAP) at the nanoplex formation step to produce PAP-decorated antibiotic-polyelectrolyte nanoplex exhibiting built-in mucolytic capability. Ciprofloxacin (CIP) and dextran sulfate (DXT) were used as the models for antibiotics and polyelectrolyte, respectively. The results showed that the PAP inclusion had minimal effects on the physical characteristics, preparation efficiency, and dissolution of the CIP-DXT nanoplex. The optimal CIP-(DXT-PAP) nanoplex exhibited size and zeta potential of approximately 200 nm and -50 mV with CIP and PAP payloads of 60% and 32% (w/w), respectively. The nanoplex was prepared at high efficiency with larger than 80% CIP and PAP utilization rates. The CIP-(DXT-PAP) nanoplex exhibited tenfold improvement in the mucus permeability compared to its CIP-DXT nanoplex counterpart, resulting in the former's superior bactericidal activity against clinical Pseudomonas aeruginosa biofilm in the presence of mucus barrier. A trade-off, nevertheless, existed between antibacterial efficacy and cytotoxicity towards human lung epithelium cells upon the incorporation of PAP above a certain concentration threshold. Therefore, the optimal dosing of the CIP-(DXT-PAP) nanoplex must be carefully determined.