Incorporation and antimicrobial activity of nisin Z within carrageenan/chitosan multilayers.

An antimicrobial peptide, nisin Z, was embedded within polyelectrolyte multilayers (PEMs) composed of natural polysaccharides in order to explore the potential of forming a multilayer with antimicrobial properties. Using attenuated total reflection Fourier transform infrared spectroscopy (ATR FTIR), the formation of carrageenan/chitosan multilayers and the inclusion of nisin Z in two different configurations was investigated. Approximately 0.89 µg cm-2 nisin Z was contained within a 4.5 bilayer film. The antimicrobial properties of these films were also investigated. The peptide containing films were able to kill over 90% and 99% of planktonic and biofilm cells, respectively, against Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA) strains compared to control films. Additionally, surface topography and wettability studies using atomic force microscopy (AFM) and the captive bubble technique revealed that surface roughness and hydrophobicity was similar for both nisin containing multilayers. This suggests that the antimicrobial efficacy of the peptide is unaffected by its location within the multilayer. Overall, these results demonstrate the potential to embed and protect natural antimicrobials within a multilayer to create functionalised coatings that may be desired by industry, such as in the food, biomaterials, and pharmaceutical industry sectors.

Odd-even effects on hydration of natural polyelectrolyte multilayers: An in situ synchrotron FTIR microspectroscopy study.

HYPOTHESIS: Odd-even effects in polysaccharide polyelectrolyte multilayers influence their hydration content and the chemical environment of the water within them. EXPERIMENTS: Polysaccharide polyelectrolyte multilayers (PEMs) composed of pharmaceutical grade fucoidan and chitosan were studied under confinement using synchrotron FTIR microspectroscopy at increasing pressure, in order to isolate and measure infrared spectra of water within the PEM, without interference from bulk water. Complementary studies of the PEMs were carried out using lab-based in situ attenuated total reflectance Fourier transform spectroscopy (ATR FTIR) and quartz crystal microbalance with dissipation monitoring (QCM-D), as well as zeta potential measurements, to determine the quantity of adsorbed polymer, hydration content, film thickness, viscoelastic properties and surface charge during layer-by-layer deposition. FINDINGS: The hydration of the PEM followed a saw-tooth profile, known as the odd-even effect, where the film increased hydration with fucoidan adsorption and dehydrated/densified with chitosan adsorption. The water structure within the film showed a lower degree of hydrogen bonding than water in the bulk electrolyte. However, the water structure/environment was independent of the terminating layer of the PEM, in spite of the alteration in percentage hydration water, indicating only a partial proof of the initial hypothesis for this multilayer system (hydration amount changes, hydration water environment does not).

The influence of pH on the interfacial behaviour of Quillaja bark saponin at the air-solution interface.

The interfacial behavior of surfactants present in a natural extract from Quillaja saponaria Molina bark at the air-solution interface is studied by measurements of interfacial tension, interfacial elasticity, and interfacial reflectance FTIR spectroscopy. The active molecule, saponin, is observed directly at the air-solution interface (via reflectance FTIR spectroscopy) above and below the pKa of the molecule, and spectra confirm the altered charge of the interfacial layer at the two solution conditions. For all concentrations of saponin studied, and at pH values below and above pKa (i.e. pH 3 and 7), a reduction in interfacial tension as a function of time is observed, with some differences in early time-scale adsorption and with lower values of quasi-equilibrium interfacial tension for pH 3. The interfacial layer is seen to be elastic, as determined from measurements of hydrostatic expansion, with some variation at the two pH values, and as a function of concentration. In addition to interfacial layer characterisation, the interaction between two air-solution interfaces is probed using bubble collisions with an air-solution interface. This experiment allows for observation of thin film drainage kinetics and determination of the final foam film thickness for the case when one of the interfaces is at equilibrium while the dynamic adsorption layer is being established at the other. This is the first time when the interactions between such interfaces (i.e. only one being at equilibrium) have been studied. This is of particular importance for the formation stage of foams, during which time many of the interfaces are not at equilibrium. When two interfaces interact across a thin liquid film, pH is seen to significantly influence foam film thickness.

A Novel Soft Contact Piezo-Controlled Liquid Cell for Probing Polymer Films under Confinement using Synchrotron FTIR Microspectroscopy.

Soft polymer films, such as polyelectrolyte multilayers (PEMs), are useful coatings in materials science. The properties of PEMs often rely on the degree of hydration, and therefore the study of these films in a hydrated state is critical to allow links to be drawn between their characteristics and performance in a particular application. In this work, we detail the development of a novel soft contact cell for studying hydrated PEMs (poly(sodium 4-styrenesulfonate)/poly(allylamine hydrochloride)) using FTIR microspectroscopy. FTIR spectroscopy can interrogate the nature of the polymer film and the hydration water contained therein. In addition to reporting spectra obtained for hydrated films confined at the solid-solid interface, we also report traditional ATR FTIR spectra of the multilayer. The spectra (microspectroscopy and ATR FTIR) reveal that the PEM film build-up proceeds as expected based on the layer-by-layer assembly methodology, with increasing signals from the polymer FTIR peaks with increasing bilayer number. In addition, the spectra obtained using the soft contact cell indicate that the PEM film hydration water has an environment/degree of hydrogen bonding that is affected by the chemistry of the multilayer polymers, based on differences in the spectra obtained for the hydration water within the film compared to that of bulk electrolyte.

Inorganic surface chemistry and nanostructure controls lipolytic product speciation and partitioning during the digestion of inorganic-lipid hybrid particles.

HYPOTHESIS: Solid-state lipid formulations, whereby liquid lipids are encapsulated in inorganic particle matrices, have attracted significant interest for drug/nutrient delivery in recent years. We hypothesized that the surface chemistry of the inorganic material used to encapsulate lipids impacts the lipase-mediated digestion and partitioning of lipolytic species between the solubilized aqueous and insoluble pellet phases. EXPERIMENTS: Medium chain triglycerides were spray dried with silica nanoparticles, montmorillonite or laponite platelets to form inorganic-lipid hybrid particles. In vitro lipolysis studies were conducted under gastric (pH 1.6) and intestinal (pH 7.5) conditions, and the speciation and partitioning of lipolytic products between the aqueous and pellet phases was characterized using solution-state proton nuclear magnetic resonance and fourier transform infrared spectroscopy. FINDINGS: Under gastric conditions, greater than 80% of all lipid species remained adsorbed within each lipolysis pellet after 60 min. Approximately 40%, 50-60% and 80-90% of all lipid species were adsorbed from solution by silica-, montmorillonite- and laponite-based particle matrices during intestinal lipolysis. Monoglycerides were preferentially adsorbed by silica, whereas triglycerides and fatty acids were adsorbed by montmorillonite and laponite. Adsorption of lipolytic products from solution is expected to impact significantly on drug/nutrient solubilization and absorption in vivo. To the best of our knowledge, this is the first report characterizing the speciation and phase behavior of lipolytic products released from solid-state lipid formulations during in vitro lipolysis studies.

Formation and enzymatic degradation of poly-l-arginine/fucoidan multilayer films.

A polyelectrolyte multilayer (PEM) system based on biopolymers has been constructed and studied in its formation and enzymatic breakdown. The multilayer is composed of fucoidan (a proven antimicrobial/anti-inflammatory seaweed-based polysaccharide) and poly-l-arginine (a polypeptide that can be readily degraded with trypsin to yield arginine, a known NO donor), thus making the multilayer a potential dual action surface treatment for wound dressings. Studies on the formation of the multilayer revealed that the film built-up in the expected stepwise manner with consistent reversal of the zeta potential upon the adsorption of each subsequent polyion. The completed film (8 bilayers) was seen to have low hydration (30% water), as determined by H2O/D2O solvent replacement studies using the quartz crystal microbalance, with an adsorbed mass (without hydration water) of approx. 4.8µgcm-2, as determined by quantitative attenuated total reflectance Fourier transform infrared (ATR FTIR) spectroscopy. The enzymatic breakdown of the film in response to exposure to trypsin was also investigated, and the film was seen to release both polymers over time, with a projected complete film removal period of approximately 24h. Critically, this information was determined using ATR FTIR spectroscopy experiments, which allowed unambiguous deconvolution of the removal rates of the two polyions, which is information that cannot be obtained from other methodologies used to study enzymatic breakdown of surface films.

The influence of polyanion molecular weight on polyelectrolyte multilayers at surfaces: elasticity and susceptibility to saloplasticity of strongly dissociated synthetic polymers at fluid-fluid interfaces.

We studied the interfacial mechanical properties of polyelectrolyte multilayer assemblies of poly(diallylamine hydrochloride) (PAH) and poly(4-styrenesulfonate)sodium salt (PSS) at the air-water interface using axisymmetric drop shape analysis (ADSA) during hydrostatic inflation as a function of aqueous salt concentration and two different polyanion molecular weights (Mw ∼ 13 and 70 kDa). Surface elastic moduli (Gs) ranged from 50 to 300 mN m-1. Using the measured film thickness, the bulk moduli (G) ranged from 10 to 90 MPa consistent with elastomeric solids. This solid-like interface was evidenced by a systematic departure of the inflated shape from the Young-Laplace equation, which assumes a liquid-like interface. Surface elastic moduli increased and bulk elastic moduli decreased with increasing nanomembrane transverse dimension, and multilayers with the lower molecular weight anion were more transversely compact than those of higher molecular weight and displayed a larger elastic modulus. The bulk moduli of both types of multilayer assemblies asymptotically approach a constant value for films with more than two bilayers of polyelectrolyte, consistent with the observed transition from a 'glassy' to 'rubbery' state. Both types of multilayer assemblies displayed plasticization with increasing sodium chloride concentration in the adjoining aqueous phase, i.e. saloplasticity, and exhibited a transition from elastic to plastic response to deformation. The restored mobility of the polyelectrolyte resulting from the shift from intrinsic to extrinsic charge complexation, restores fluidity to the interface and is evidenced by experimental observation of a liquid-like interface when loaded. The higher molecular weight polyanion multilayers plasticized at lower salt concentrations suggesting that the lower melting point of the higher molecular weight polyanion assembly is attributable to a lesser extent of electrostatic cross-linking underscoring the unconventional dependence of molecular weight on saloplasticity in strongly dissociated polyelectrolytes.

The influence of polyanion molecular weight on polyelectrolyte multilayers at surfaces: protein adsorption and protein-polysaccharide complexation/stripping on natural polysaccharide films on solid supports.

Two different fucoidan polymers (unfractionated Fucus vesiculosus fucoidan, and fractionated low molecular weight Fucus vesiculosus fucoidan) have been used to create substrates for protein adsorption studies. Polyelectrolyte multilayers were formed using the fucoidans (polyanions) with chitosan as the corresponding polycation. Multilayer formation was studied using zeta potential measurements, quartz crystal microbalance with dissipation monitoring (QCM-D) and attenuated total reflectance (ATR) FTIR spectroscopy. The formation studies reveal that the low molecular weight (LMW) fucoidan produces a less hydrated multilayer, with a significantly increased adsorbed mass, and with fucoidan as the diffusing species during formation. Protein adsorption studies using bovine serum albumin (BSA) were undertaken for solution conditions designed to mimic biological conditions, and to minimise the role of electrical double layer forces in influencing adsorption. Under these conditions, and as revealed by ATR FTIR spectroscopy, BSA is seen to adsorb less substantially to multilayers formed with the LMW fucoidan, and to cause extraction/stripping of the LMW fucoidan from the multilayer. FTIR spectra reveal that the protein adopts a different conformation when adsorbed to the LMW fucoidan multilayer, both relative to the protein in solution and when adsorbed at the surface of the multilayer formed from unfractionated fucoidan.