- Title
- An electrospun nanofiber colorimetric probe for detection of Alkaline Phosphatase for diagnosis of liver toxicity
- Creator
- Mohale, Mamello
- ThesisAdvisor
- Torte, Nelson
- Date
- 2014
- Type
- text
- Type
- Thesis
- Type
- Masters
- Type
- MSc
- Identifier
- http://hdl.handle.net/10962/54532
- Identifier
- vital:26581
- Description
- A novel electrospun nanofiber colorimetric probe for the detection of Alkaline Phosphatase (ALP) for diagnosis of liver toxicity was developed through electrospinning of a para nitrophenyl phosphate (pNPP) functionalised nylon polymer. The chemical stability of the enzyme substrate (pNPP) and its two products (para nitrophenol (pNP) and para nitrophenolate (pNPL)) was evaluated in biological (pH 7.4), acidic (pH 2) and alkaline (pH 9) pH, respectively. Enzyme kinetics models of Michaelis Menten (MM) and Lineweaver Burk (LB) were used to characterise free ALP. Solution and nanofiber assaying of free ALP and 10x diluted serum (spiked with ALP) was also carried out. The results demonstrated that pNPP and pNP were colourless while pNPL was yellow which indicated that all reagents were chemically stable. In addition, the chromophore of pNPL exhibited a strong molar extinction coefficient (ε) of 18,458 M-1 cm-1. LB plot being the most accurate compared to MM showed Vmax, Km and excess substrate concentration of 5.5 × 10-3 μmol/min-1, 0.025 mM and 0.25 mM respectively. Solution and nanofiber assaying of free ALP and serum confirmed a direct proportional correlation between the pNPL yellow colour intensity and enzyme activity up to 858 IU/L and 820 IU/L respectively. The dipping of the nanofiber layer into solution showed that the leaching rate of pNPP was extremely high at 1.37 × 10-3 A/min-1 as was observed after only the first 0.25 min interval. However, this was not of great concern since it was also observed that administration of the sample (20 μL) by a dropwise method minimised leaching compared to dipping. The preliminary findings on the effect of temperature on the chemical stability of pNPP indicated that it was stable below temperatures of 40°C while it hydrolysed at 80°C. Therefore a sensitive, rapid and simple colorimetric probe for the detection of ALP was developed. The probe exhibited characteristics that make it suitable to be incorporated into point of care colorimetric liver toxicity diagnostic devices for applications in resource poor settings and telemedicine.
- Format
- 63 leaves, pdf
- Publisher
- Rhodes University, Faculty of Science, Chemistry
- Language
- English
- Rights
- Mohale, Mamello
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- Downloads: 366
Collections
RU Department of Chemistry
| Thumbnail | File | Description | Size | Format | |||
|---|---|---|---|---|---|---|---|
| View Details | SOURCE1 | Microsoft Word - MASTERS THESIS. MAMELLO MOHALE -corrections Copy 2007.doc | 1 MB | Adobe Acrobat PDF | View Details |