An electrospun nanofiber colorimetric probe for detection of Alkaline Phosphatase for diagnosis of liver toxicity
- Authors: Mohale, Mamello
- Date: 2014
- Language: English
- Type: text , Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/54532 , 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.
- Full Text:
- Date Issued: 2014
- Authors: Mohale, Mamello
- Date: 2014
- Language: English
- Type: text , Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/54532 , 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.
- Full Text:
- Date Issued: 2014
Development of styrene based imprinted sorbents for selective clean-up of metalloporphyrins in organic media
- Authors: Awokoya, Kehinde Nurudeen
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/54521 , vital:26580
- Description: Most crude oils contain traces of vanadium and nickel complex with porphyrins (VTPP and NTPP) within their asphaltene fraction. Although these metals are only present in trace quantities, they have a significant and detrimental impact on the refining process by degrading the quality of intermediate and end products. Therefore, their selective removal is highly desirable. This thesis presents the development of nickel porphyrin, nickel vanadyl porphyrin imprinted nanofibers and vanadyl porphyrin imprinted polymer (MIP) particles for application as selective sorbents. Computational model based on the combination of molecular dynamics simulations and quantum mechanics was successfully applied to the styrene functional monomer selection. The particle was prepared by bulk polymerization and the nanofibers by a novel approach combining molecular imprinting and electrospinning technology. The morphologies, thermal stabilities and porosities of the imprinted sorbents were studied using SEM, TGA, and BET nitrogen gas adsorption. Chloroform was found to swell the polymer particles to a higher degree than methanol and acetonitrile. The adsorption characteristics of the imprinted sorbents best fitted with Freundlich isotherm model. The imprinted sorbents recorded high extraction efficiencies (EEs) of > 99 % in selectively extracting the metalloporphyrins. The impact of the template on the affinity of recognition for NTPP was evaluated. The results showed that the NTPP adsorption capacity increased as the molar ratio of NTPP to styrene was increased from 1:1 to 3:1. The optimal ratio of template to functional monomer which yielded the best specific affinity and highest recovery (99.9 %) was 3:1. The effects of trifluoroacetic acid (TFA), dichloromethane (DCM), dimethyl sulphoxide (DMSO), pentane (PEN) on electro-spinnability of the polymer solutions and the morphological appearance of the nanofibers were investigated. The imprinted nanofibers exhibited the same selectivity specialism for both NTPP and VTPP. A remarkable stability in relation to reusability was observed when imprinted nanofibers were used, as they could be reused nine times without incurring any significant loss in removal efficiency. The results were validated by analysing a certified reference material. The imprinted sorbents were therefore found to be selective sorbents that are well suited for handling trace metals in organic media.
- Full Text:
- Date Issued: 2014
- Authors: Awokoya, Kehinde Nurudeen
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/54521 , vital:26580
- Description: Most crude oils contain traces of vanadium and nickel complex with porphyrins (VTPP and NTPP) within their asphaltene fraction. Although these metals are only present in trace quantities, they have a significant and detrimental impact on the refining process by degrading the quality of intermediate and end products. Therefore, their selective removal is highly desirable. This thesis presents the development of nickel porphyrin, nickel vanadyl porphyrin imprinted nanofibers and vanadyl porphyrin imprinted polymer (MIP) particles for application as selective sorbents. Computational model based on the combination of molecular dynamics simulations and quantum mechanics was successfully applied to the styrene functional monomer selection. The particle was prepared by bulk polymerization and the nanofibers by a novel approach combining molecular imprinting and electrospinning technology. The morphologies, thermal stabilities and porosities of the imprinted sorbents were studied using SEM, TGA, and BET nitrogen gas adsorption. Chloroform was found to swell the polymer particles to a higher degree than methanol and acetonitrile. The adsorption characteristics of the imprinted sorbents best fitted with Freundlich isotherm model. The imprinted sorbents recorded high extraction efficiencies (EEs) of > 99 % in selectively extracting the metalloporphyrins. The impact of the template on the affinity of recognition for NTPP was evaluated. The results showed that the NTPP adsorption capacity increased as the molar ratio of NTPP to styrene was increased from 1:1 to 3:1. The optimal ratio of template to functional monomer which yielded the best specific affinity and highest recovery (99.9 %) was 3:1. The effects of trifluoroacetic acid (TFA), dichloromethane (DCM), dimethyl sulphoxide (DMSO), pentane (PEN) on electro-spinnability of the polymer solutions and the morphological appearance of the nanofibers were investigated. The imprinted nanofibers exhibited the same selectivity specialism for both NTPP and VTPP. A remarkable stability in relation to reusability was observed when imprinted nanofibers were used, as they could be reused nine times without incurring any significant loss in removal efficiency. The results were validated by analysing a certified reference material. The imprinted sorbents were therefore found to be selective sorbents that are well suited for handling trace metals in organic media.
- Full Text:
- Date Issued: 2014
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