title: Synthesis and Identification of FITC-Insulin Conjugates Produced Using Human Insulin and Insulin Analogues for Biomedical Applications.
abstract: Human insulin was fluorescently labelled with fluorescein isothiocyanate (FITC) and the conjugate species produced were identified using high performance liquid chromatography and electrospray mass spectroscopy. Mono-labelled FITC-insulin conjugate (A1 or B1) was successfully produced using human insulin at short reaction times (up to 5 h) however the product always contained some unlabelled native human insulin. As the reaction time was increased over 45 h, no unlabelled native human insulin was present and more di-labelled FITC-insulin conjugate (A1B1) was produced than mono-labelled conjugate with the appearance of tri-labelled conjugate (A1B1B29) after 20 h reaction time. The quantities switch from mono-labelled to di-labelled FITC-insulin conjugate between reaction times 9 and 20 h. In the presence of phenol or m-cresol, there appears to be a 10 % decrease in the amount of mono-labelled conjugate and an increase in di-labelled conjugate produced at lower reaction times. Clinically used insulin analogues present in commercially available preparations were successfully fluorescently labelled for future biomedical applications.
authors.affiliation: Leicester School of Pharmacy, Faculty of Health and Life Sciences, De Montfort University, The Gateway, Leicester, LE1 9BH, UK.
authors.forename: Dolly
authors.initials: D
authors.lastname: Jacob
authors.affiliation: Leicester School of Pharmacy, Faculty of Health and Life Sciences, De Montfort University, The Gateway, Leicester, LE1 9BH, UK.
authors.forename: M
authors.initials: M
authors.lastname: Joan Taylor
authors.affiliation: Leicester School of Pharmacy, Faculty of Health and Life Sciences, De Montfort University, The Gateway, Leicester, LE1 9BH, UK.
authors.forename: Paul
authors.initials: P
authors.lastname: Tomlins
authors.affiliation: Leicester School of Pharmacy, Faculty of Health and Life Sciences, De Montfort University, The Gateway, Leicester, LE1 9BH, UK. ssahota@dmu.ac.uk.
authors.forename: Tarsem S
authors.initials: TS
authors.lastname: Sahota
mesh_terms: D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000302:isolation & purification; D005456:Fluorescent Dyes / Q000737:chemistry*; D006801:Humans; D007328:Insulin / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry* / Q000302:isolation & purification; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D005456:Fluorescent Dyes; D007328:Insulin; C031573:insulin, fluorescein-isothiocyanated-; D016650:Fluorescein-5-isothiocyanate
keywords: FITC-insulin; Fluorescent labelling; HPLC; Insulin; Mono-labelled