Now showing 1 - 10 of 13
  • Publication
    PK/PD modelling of combed-shaped PEGylated salmon calcitonin conjugates of differing molecular weights
    Salmon calcitonin (sCT) was conjugated via cysteine-1 to novel comb-shaped end-functionalised (poly(PEG) methyl ether methacrylate) (sCT-P) polymers, to yield conjugates of total molecular weights (MW) inclusive of sCT: 6.5, 9.5, 23 and 40 kDa. The conjugates were characterised by HPLC and their in vitro and in vivo bioactivity was measured by cAMP assay on human T47D cells and following intravenous (i.v.) injection to rats, respectively. Stability against endopeptidases, rat serum and liver homogenates was assessed. There were linear and exponential relationships between conjugate MW with potency and efficacy respectively, however the largest MW conjugate still retained 70% of Emax and an EC50 of 3.7 nM. In vivo, while free sCT and the conjugates reduced serum [calcium] to a maximum of 15–30% over 240 min, the half-life (T1/2) was increased and the area under the curve (AUC) was extended in proportion to conjugate MW. Likewise, the polymer conferred protection on sCT against attack by trypsin, chymotrypsin, elastase, rat serum and liver homogenates, with the best protection afforded by sCT-P (40 kDa). Mathematical modelling accurately predicted the MW relationships to in vitro efficacy, potency, in vivo PK and enzymatic stability. With a significant increase in T1/2 for sCT, the 40 kDa MW comb-shaped PEG conjugate of sCT may have potential as a long-acting injectable formulation.
      527Scopus© Citations 25
  • Publication
    Nutraceutical formulation, characterisation, and in-vitro evaluation of methylselenocysteine and selenocystine using food derived chitosan: zein nanoparticles
    Selenoamino acids (SeAAs) have been shown to possess antioxidant and anticancer properties. However, their bioaccessibility is low and they may be toxic above the recommended nutritional intake level, thus improved targeted oral delivery methods are desirable. In this work, the SeAAs, Methylselenocysteine (MSC) and selenocystine (SeCys2) were encapsulated into nanoparticles (NPs) using the mucoadhesive polymer chitosan (Cs), via ionotropic gelation with tripolyphosphate (TPP) and the NPs produced were then coated with zein (a maize derived prolamine rich protein). NPs with optimized physicochemical properties for oral delivery were obtained at a 6: 1 ratio of Cs:TPP, with a 1:0.75 mass ratio of Cs:zein coating (diameter ~260 nm, polydispersivity index ~0.2, zeta potential >30 mV). Scanning Electron Microscopy (SEM) analysis showed that spheroidal, well distributed particles were obtained. Encapsulation Efficiencies of 80.7% and 78.9% were achieved, respectively, for MSC and SeCys2 loaded NPs. Cytotoxicity studies of MSC loaded NPs showed no decrease in cellular viability in either Caco-2 (intestine) or HepG2 (liver) cells after 4 and 72 h exposures. For SeCys2 loaded NPs, although no cytotoxicity was observed in Caco-2 cells after 4 h, a significant reduction in cytotoxicity was observed, compared to pure SeCys2, across all test concentrations in HepG2 after 72 h exposure. Accelerated thermal stability testing of both loaded NPs indicated good stability under normal storage conditions. Lastly, after 6 h exposure to simulated gastrointestinal tract environments, the sustained release profile of the formulation showed that 62 ± 8% and 69 ± 4% of MSC and SeCys2, had been released from the NPs respectively.
      744Scopus© Citations 20
  • Publication
    Formulation, Characterisation and Evaluation of the Antihypertensive Peptides, Isoleucine-Proline-Proline and Leucine-Lysine-Proline in Chitosan Nanoparticles Coated with Zein for Oral Drug Delivery
    Isoleucine-Proline-Proline (IPP) and Leucine-Lysine-Proline (LKP) are food-derived tripeptides whose antihypertensive functions have been demonstrated in hypertensive rat models. However, peptides display low oral bioavailability due to poor intestinal epithelial permeability and instability. IPP and LKP were formulated into nanoparticles (NP) using chitosan (CL113) via ionotropic gelation and then coated with zein. Following addition of zein, a high encapsulation efficiency (EE) (>80%) was obtained for the NP. In simulated gastric fluid (SGF), 20% cumulative release of the peptides was achieved after 2 h, whereas in simulated intestinal fluid (SIF), ~90% cumulative release was observed after 6 h. Higher colloidal stability (39–41 mV) was observed for the coated NP compared to uncoated ones (30–35 mV). In vitro cytotoxicity studies showed no reduction in cellular viability of human intestinal epithelial Caco-2 and HepG2 liver cells upon exposure to NP and NP components. Administration of NP encapsulating IPP and LKP by oral gavage to spontaneously hypertensive rats (SHR) attenuated systolic blood pressure (SBP) for 8 h. This suggests that the NP provide appropriate release to achieve prolonged hypotensive effects in vivo. In conclusion, chitosan-zein nanoparticles (CZ NP) have potential as oral delivery system for the encapsulation of IPP and LKP.
      41Scopus© Citations 10
  • Publication
    Antibacterial effects of poly(2-(dimethylamino ethyl)methacrylate) against selected Gram-positive and Gram-negative bacteria
    Antimicrobial coatings can reduce the occurrence of medical device-related bacterial infections. Poly(2-(dimethylamino ethyl)methacrylate)) (pDMAEMA) is one such polymer that is being researched in this regard. The aims of this study were to (1) elucidate pDMAEMA’s antimicrobial activity against a range of Gram-positive and Gram-negative bacteria and (2) to investigate its antimicrobial mode of action. The methods used include determination of minimum inhibitory concentration (MIC) values against various bacteria and the effect of pH and temperature on antimicrobial activity. The ability of pDMAEMA to permeabilise bacterial membranes was determined using the dyes 1-N-phenyl-naphthylamine (NPN) and Calcein-AM. Flow cytometry was used to investigate pDMAEMA’s capacity to be internalised by bacteria and to determine effects on bacterial cell cycling. pDMAEMA was bacteriostatic against Gram-negative bacteria with MIC values between 0.1–10 mg/ml. MIC values against Gram-positive bacteria were variable. pDMAEMA was active against Gram-positive bacteria around its pKa and at lower pH values, while it was active against Gram-negative bacteria around its pKa and at higher pH values. pDMAEMA inhibited bacterial growth by binding to the outside of the bacteria, permeabilising the outer membrane and disrupting the cytoplasmic membrane. By incorporating pDMAEMA with erythromycin, it was found that the efficacy of the latter was increased against Gram-negative bacteria. Together, the results illustrate that pDMAEMA acts in a similar fashion to other cationic biocides.
      1362Scopus© Citations 210
  • Publication
    Formulation, Characterization and Stability Assessment of a Food‐Derived Tripeptide, Leucine‐Lysine‐Proline Loaded Chitosan Nanoparticles
    The chicken‐ or fish‐derived tripeptide, leucine‐lysine‐proline (LKP), inhibits the angiotensin converting enzyme and may be used as an alternative treatment for prehypertension. However, it has low permeation across the small intestine. The formulation of LKP into a nanoparticle (NP) has the potential to address this issue. LKP‐loaded NPs were produced using an ionotropic gelation technique, using chitosan (CL113). Following optimization of unloaded NPs, a mixture amount design was constructed using variable concentration of CL113 and tripolyphosphate at a fixed LKP concentration. Resultant particle sizes ranged from 120 to 271 nm, zeta potential values from 29 to 37 mV, and polydispersity values from 0.3 to 0.6. A ratio of 6:1 (CL113:TPP) produced the best encapsulation of approximately 65%. Accelerated studies of the loaded NPs indicated stability under normal storage conditions (room temperature). Cytotoxicity assessment showed no significant loss of cell viability and in vitro release studies indicated an initial burst followed by a slower and sustained release.
    Scopus© Citations 8  331
  • Publication
    Comparative study of the structural and physicochemical properties of two food derived antihypertensive tri-peptides, Isoleucine-Proline-Proline and Leucine-Lysine-Proline encapsulated into a chitosan based nanoparticle system
    Food derived tri-peptides; Leucine-Lysine-Proline (LKP) and Isoleucine-Proline-Proline (IPP) are angiotensin converting enzyme inhibitors and may have potential to alleviate hypertension. The aim of this work was to understand the interactions of IPP and LKP when formulated into a chitosan nanoparticle (NP) to help improve permeation. Our findings indicate different mean inhibitory concentrations (LKP: 0.36 ± 0.01 μM and IPP: 3.1 ± 0.6 μM) and encapsulation efficiencies at different ratios of chitosan: tripolyphosphate (LKP NPs: 65% at 6:1 and IPP NPs: 43% at 4:1). Molecular modelling and circular dichroism showed different stable amino side-chain-specific conformations for each peptide. IPP showed more steric hindrances to intra-chain rotations, resulting in an unordered peptide structure, whereas LKP showed more flexibility associated with a (disordered) β-strand-like conformer. In-vitro release kinetics showed a slower release for LKP NPs in acidic pH compared to IPP NPs. In conclusion, LKP NPs were found to have better binding compatibility with chitosan.
      534Scopus© Citations 14
  • Publication
    Oral delivery strategies for nutraceuticals: Delivery vehicles and absorption enhancers
    Lifestyle issues contribute to the development of obesity, type 2 diabetes, and cardiovascular disease. Together with appropriate diet and exercise, nutraceuticals may contribute to managing prevention at an early stage prior to therapeutic intervention. However, many useful food-derived bioactive compounds will not sufficiently permeate the small intestine to yield efficacy without appropriate oral delivery technology. The pharmaceutical industry uses commercialised approaches for oral delivery including solubilizing technologies for small molecules, which could be applied to selected nutraceuticals with solubility issues. Systems currently being studied for labile and poorly permeable hydrophilic peptides and macromolecules include nanoparticles, intestinal permeation enhancers (PE) and mucolytics. These may also have potential for application to nutraceuticals with similar sub-optimal physicochemical characteristics. Scope and approach We introduce factors which effect oral delivery of four types of nutraceuticals, namely fatty acids, bioactive peptides, micronutrients, and phytochemicals. Factors preventing oral absorption can arise from molecule physicochemical characteristics, which influence solubility, stability, and epithelial permeability in the gastrointestinal tract (GIT). We highlight the potential of selected delivery strategies to improve oral bioavailability of different types of nutraceuticals. Key findings and conclusions There is an opportunity for the nutraceutical industry to leverage the pharmaceutical industry's progress in oral drug delivery. The use of delivery approaches using formulation with excipients or substances with a history of use in man has potential to improve solubility, stability, or permeability of nutraceuticals, leading to improved oral bioavailability. Leveraging oral delivery formulation approaches across nutraceutical and pharmaceutical molecules will lead to synergies for both fields.
      1509Scopus© Citations 96
  • Publication
    An intra-articular salmon calcitonin-based nanocomplex reduces experimental inflammatory arthritis
    Prolonged inappropriate inflammatory responses contribute to the pathogenesis of rheumatoid arthritis (RA) and to aspects of osteoarthritis (OA). The orphan nuclear receptor, NR4A2, is a key regulator and potential biomarker for inflammation and represents a potentially valuable therapeutic target. Both salmon calcitonin (sCT) and hyaluronic acid (HA) attenuated activated mRNA expression of NR4A1, NR4A2, NR4A3, and matrix metalloproteinases (MMPs) 1, 3 and 13 in three human cell lines: SW1353 chondrocytes, U937 and THP-1 monocytes. Ad-mixtures of sCT and HA further down-regulated expression of NR4A2 compared to either agent alone at specific concentrations, hence the rationale for their formulation in nanocomplexes (NP) using chitosan. The sCT released from NP stimulated cAMP production in human T47D breast cancer cells expressing sCT receptors. When NP were injected by the intra-articular (I.A.) route to the mouse knee during on-going inflammatory arthritis of the K/BxN serum transfer model, joint inflammation was reduced together with NR4A2 expression, and local bone architecture was preserved. These data highlight remarkable anti-inflammatory effects of sCT and HA at the level of reducing NR4A2 mRNA expression in vitro. Combining them in NP elicits anti-arthritic effects in vivo following I. A. delivery.
    Scopus© Citations 61  982
  • Publication
    Evaluation of Selenomethionine Entrapped in Nanoparticles for Oral Supplementation Using In Vitro, Ex Vivo and In Vivo Models
    Selenium methionine (SeMet) is an essential micronutrient required for normal body function and is associated with additional health benefits. However, oral administration of SeMet can be challenging due to its purported narrow therapeutic index, low oral bioavailability, and high susceptibility to oxidation. To address these issues, SeMet was entrapped in zein-coated nanoparticles made from chitosan using an ionic gelation formulation. The high stability of both the SeMet and selenomethionine nanoparticles (SeMet-NPs) was established using cultured human intestinal and liver epithelial cells, rat liver homogenates, and rat intestinal homogenates and lumen washes. Minimal cytotoxicity to Caco-2 and HepG2 cells was observed for SeMet and SeMet-NPs. Antioxidant properties of SeMet were revealed using a Reactive Oxygen Species (ROS) assay, based on the observation of a concentration-dependent reduction in the build-up of peroxides, hydroxides and hydroxyl radicals in Caco-2 cells exposed to SeMet (6.25–100 μM). The basal apparent permeability coefficient (Papp) of SeMet across isolated rat jejunal mucosae mounted in Ussing chambers was low, but the Papp was increased when presented in NP. SeMet had minimal effects on the electrogenic ion secretion of rat jejunal and colonic mucosae in Ussing chambers. Intra-jejunal injections of SeMet-NPs to rats yielded increased plasma levels of SeMet after 3 h for the SeMet-NPs compared to free SeMet. Overall, there is potential to further develop SeMet-NPs for oral supplementation due to the increased intestinal permeability, versus free SeMet, and the low potential for toxicity.
  • Publication
    Stability, toxicity and intestinal permeation enhancement of two food-derived antihypertensive tripeptides, Ile-Pro-Pro and Leu-Lys-Pro
    Two food-derived ACE inhibitory peptides, Ile-Pro-Pro (IPP) and Leu-Lys-Pro (LKP), may have potential as alternative treatments for treatment of mild- or pre-hypertension. Lack of stability to secretory and intracellular peptidases and poor permeability across intestinal epithelia are typical limiting factors of oral delivery of peptides. The stability of IPP and LKP was confirmed in vitro in rat intestinal washes, and intestinal and liver homogenates over 60min. A positive protein control for peptidases, insulin, was significantly digested in each format over the same period. Neither tripeptide showed cytotoxic activity on Caco-2 and Hep G2 cells using the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay, even after chronic exposure. The basal Papp of fluorescein isothiocyanate (FITC)-labeled IPP and FITC-LKP across isolated rat jejunal and colonic mucosae were low, but were significantly increased in each tissue type by the medium chain fatty acids (MCFA) permeation enhancers, sodium caprate (C10) and the sodium salt of 10-undecylenic acid (uC11). IPP and LKP were therefore stable against intestinal and liver peptidases and were non-cytotoxic; their Papp values across rat intestinal mucosae were low, but could be increased by MCFA. There is potential to make on oral dosage form once in vivo pharmacology is confirmed. 
      672Scopus© Citations 37