Optimisation of glutathione conjugation to liposomes quantified with a validated HPLC assay.

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dc.contributor.author Reginald-Opara, Joy N en
dc.contributor.author Svirskis, Darren en
dc.contributor.author OCarroll, Simon en
dc.contributor.author Sreebhavan, Sreevalsan en
dc.contributor.author Dean, Justin en
dc.contributor.author Wu, Zimei en
dc.date.accessioned 2019-10-01T02:45:06Z en
dc.date.issued 2019-08 en
dc.identifier.issn 0378-5173 en
dc.identifier.uri http://hdl.handle.net/2292/48192 en
dc.description.abstract Glutathione (GSH) grafted onto nanoliposomes (GSH-liposomes) have the potential to enhance drug delivery into the brain. GSH is known to be an unstable tripeptide, however, despite widespread use to promote active transport its stability has been largely ignored to date. Therefore this study focuses on the optimisation of GSH conjugation with liposomes, supported with a validated HPLC assay for GSH. An isocratic stability-indicating HPLC assay of GSH was developed after derivatisation of GSH with 5,5'-dithio-bis-2-nitrobenzoic acid and applied for efficient conjugation of GSH to DSPE-PEG2000-maleimide lipid, either in solution or in preformed liposomes (4% molar ratio) at pH 7.4. The conjugation rate was monitored by the HPLC assay to optimise the conjugation conditions, including GSH concentration, GSH:lipid ratio, reaction time, temperature and medium. The physiochemical properties of the resulting GSH-liposomes and their GSH densities were characterised. The HPLC method was linear in the range of 0.05-50 µg/ml, highly sensitive (limit of quantification 50 ng/ml), and accurate (98-102% recoveries) with less than 4% intra-day and inter-day variability. Interestingly, enhanced GSH stability was observed at higher GSH concentrations ≥2 mg/ml and mass spectroscopy confirmed that GSH degradation occurred predominantly by oxidation. Both the proton nuclear magnetic resonance (1H NMR) spectra and HPLC analysis of GSH concentrations confirmed the formation of GSH-PEG-DSPE conjugate. Under the optimal conditions, complete conjugation was attained either by post-insertion or direct conjugation methods with the resulting GSH-liposomes attaining a GSH density of 4% with similar size (120 nm) and zeta potential (-26.7 ± 0.9 mV or -29.8 ± 1.5 mV). The study provides useful information on GSH stability for the optimisation of its conjugation in liposomal formulation. en
dc.format.medium Print-Electronic en
dc.language eng en
dc.relation.ispartofseries International journal of pharmaceutics en
dc.rights Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. Previously published items are made available in accordance with the copyright policy of the publisher. en
dc.rights.uri https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm en
dc.subject Glutathione en
dc.subject Liposomes en
dc.subject Chromatography, High Pressure Liquid en
dc.subject Reproducibility of Results en
dc.title Optimisation of glutathione conjugation to liposomes quantified with a validated HPLC assay. en
dc.type Journal Article en
dc.identifier.doi 10.1016/j.ijpharm.2019.118451 en
pubs.begin-page 118451 en
pubs.volume 567 en
dc.rights.holder Copyright: The author en
pubs.publication-status Published en
dc.rights.accessrights http://purl.org/eprint/accessRights/RestrictedAccess en
pubs.subtype Journal Article en
pubs.elements-id 775926 en
pubs.org-id Medical and Health Sciences en
pubs.org-id Medical Sciences en
pubs.org-id Anatomy and Medical Imaging en
pubs.org-id Physiology Division en
pubs.org-id Pharmacy en
dc.identifier.eissn 1873-3476 en
pubs.record-created-at-source-date 2019-06-24 en
pubs.dimensions-id 31229530 en

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