title: Formulation of abalone hemocyanin with high antiviral activity and stability.
abstract: Hemocyanin has been shown to have potential antiviral activity against herpes simplex virus type-1. However, current liquid formulations have short shelf life and high risk of bacterial contamination. The aim of our study was to develop a stable functional formulation. Analytical techniques (nano-differential scanning calorimetry and spectroscopy) and biological assays (cytotoxicity and plaque reduction) were employed to measure the effect of sugar addition on the physical properties and shelf life of the solid formulated hemocyanin. Sucrose improved thermal stability significantly by both increasing the aggregation onset temperature (70°C to>78 °C) and enhancing the activation energy (18%). Lyophilisation without trehalose caused degradation and unfolding of the α-helices of hemocyanin. However, the addition of an optimal proportion of trehalose:protein (5:1 by weight) prevented the degradation and unfolding during lyophilisation, hence maintained the protein solubility. The estimated ED50 values of the formulated solid (0.43±0.1) and liquid samples (0.37±0.06) were similar in magnitude, and were significantly lower than the respective controls; thus, confirming enhanced antiviral activity of the formulation. Formulated compounds were stable for six months at 5 °C storage. The enhanced shelf life and stable antiviral activity of the formulation offers its significant potential as effective therapeutic agent in future clinical applications.
authors.affiliation: School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Negar Talaei
authors.initials: NT
authors.lastname: Zanjani
authors.affiliation: School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Fareed
authors.initials: F
authors.lastname: Sairi
authors.affiliation: School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Gavin
authors.initials: G
authors.lastname: Marshall
authors.affiliation: Centre for Virus Research, Westmead Millennium Institute, Westmead, Sydney, NSW, Australia.
authors.forename: Monica Miranda
authors.initials: MM
authors.lastname: Saksena
authors.affiliation: School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Peter
authors.initials: P
authors.lastname: Valtchev
authors.affiliation: School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, NSW, Australia.
authors.forename: Vincent G
authors.initials: VG
authors.lastname: Gomes
authors.affiliation: Centre for Virus Research, Westmead Millennium Institute, Westmead, Sydney, NSW, Australia.
authors.forename: Anthony L
authors.initials: AL
authors.lastname: Cunningham
authors.affiliation: School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, NSW, Australia. Electronic address: fariba.dehghani@sydney.edu.au.
authors.forename: Fariba
authors.initials: F
authors.lastname: Dehghani
mesh_terms: D000818:Animals; D000998:Antiviral Agents / Q000737:chemistry* / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D002626:Chemistry, Pharmaceutical; D002522:Chlorocebus aethiops; D004355:Drug Stability; D005612:Freeze Drying; D049851:Gastropoda; D006433:Hemocyanins / Q000737:chemistry* / Q000494:pharmacology; D006561:Herpes Simplex; D018259:Herpesvirus 1, Human / Q000187:drug effects; D013395:Sucrose / Q000737:chemistry*; D014199:Trehalose / Q000737:chemistry*; D014709:Vero Cells
chemical_list: D000998:Antiviral Agents; D013395:Sucrose; D006433:Hemocyanins; D014199:Trehalose
keywords: Glycoprotein; HSV-1; Secondary structure; Thermal stability