Electrochemical dynamics of cytochrome P450-3A4 isoenzyme biosensor for protease inhibitor antiretroviral drug

Philosophiae Doctor - PhDThis thesis firstly reports on the development and characterization of reagent-less and cobalt(III) sepulchrate[Co(Sep)³⁺] mediated biosensor platforms (biosensor platform 1 and biosensor platform 2), with human recombinant heme thiolate, cytochrome P450 3A4 isoenzyme (CYP3A4), as biorecognition component. Secondly, each biosensor platform was evaluated by using an entirely different category of compound as model substrate, with the overall objective being the development of alternative analytical method for the detection and quantification of each of these substrates,by amperometric transduction method. In this regard biosensor platform 1 was evaluated for the detection of 2,4-dichlorophenol, whereas biosensor platform 2 was evaluated for the detection of protease inhibitor (PI) HAART drug, indinavir. Fourthly, this dissertation also reports on the use of genetic engineering as complimentary method during biosensor investigations, as source for continuous supply of catalytically active biological recognition component. With respect to the preparation of the biosensors in particular, biosensor platform 1 was constructed by entrapping the commercially sourced full-length, wild type CYP3A4 on a pre-formed electroactive carrier matrix, consisting of Co(Sep)³⁺–modified nafion membrane on a glassy carbon electrode. In this regard, the nafion-Co(Sep)³⁺ composite was prepared by integrating the Co(Sep)³⁺ species into a pre-formed nafion film through manual drop-coating and mixing methods

Familial needs: Comparing traditional and non-traditional families of public officials

BACKGROUND: Recognising traditional and non-traditional families in social policy is not sufficient if the policy implementation choices continue to accord more status to traditional families in comparison to non-traditional families. If unattended, this can create discriminatory practices, and human rights on an equal basis can be threatened between traditional and non-traditional families. This descriptive study compared traditional and non-traditional families of public officials with a focus on (1) form and income, (2) familial needs as per key propositions, (3) families perceptions/experiences at community and broader societal levels and (4) familial needs government must assist them with. METHODS: A quantitative research paradigm, a cross-sectional survey design was electronically administered to 600 public officials and culminated in a final sample of 70 respondents, was implemented. The study was contextualized within a contemporary family discourse, primarily influenced by a feminist perspective as well as a critique of the nuclear or traditional family grounded in functionalist theory. RESULTS: The study showed that traditional and non-traditional families of public officials are more the same than different. Public officials’ families, both traditional and non-traditional families, are affected by their inter-connectivity with communities and broader society in terms of how they experience negative treatment/ discrimination on the basis of a variety of equality issues as well as fulfillment of their socio-economic rights as stipulated in the Bill of Rights as enshrined in the South African Constitution. CONCLUSION: The study indicated that public officials, as members of families, live in both traditional and non-traditional families. Public officials have familial needs similar to any other family and are also influenced by similar factors in broader society. As both rights holders and duty bearers they can improve their own family lives and also better serve families in broader society. However they too need to be supported with their own family needs

Social resilience, structural vulnerability and capabilities in Genadendal, South Africa

The town Genadendal or Genaal as it is called by the residents, is situated in the Western Cape, and was founded by the Moravian Church in 1837. The Faculty of Community and Health Sciences at the University of the Western Cape (UWC) has had a longstanding relationship with the Genadendal community through UWC’s community engagement activities

Conductive composite biosensor system for electrochemical indinavir drug detection

Indinavir is a protease inhibitor antiretroviral (ARV) drug, which forms part of the highly active antiretroviral therapy during the treatment of HIV/AIDS. Indinavir undergoes first-pass metabolism through the cytochrome P450 (CYP) enzymes in the human liver, of which CYP3A4 is the most influential isoenzyme. Multidrug combination therapy and, as such, therapeutic drug monitoring (TDM) during HIV/AIDS treatment are therefore critical, to prevent adverse interactions. The conventional sensitive and specific assays available for quantifying ARV drugs, however, suffer from distinct disadvantages. In this regard, biosensors can be used to provide real time information on the metabolic profile of the drug. In this study, a biosensor with cobalt(III) sepulchrate trichloride {Co(Sep) 3+} as diffusional mediator was constructed. The biosensor platform consisted of CYP3A4 immobilized onto a gold nanoparticle (GNP) overoxidized polypyrrole (OvOxPpy) carrier matrix. The biosensor exhibited reversible electrochemistry, with formal potential determined as −624 ± 5 mV, from voltammetric analysis, with overall electron transfer being diffusion controlled. The biosensor showed typical electrocatalytic response to dioxygen (O2), exemplified by the distinct increase in the cathodic peak current (,). A concentration-dependent increase in , was observed in response to consecutive additions of Indinavir

Electrochemical immunosensor based on polythionine/gold nanoparticles for the determination of Aflatoxin B1

An aflatoxin B1 (AFB1) electrochemical immunosensor was developed by the immobilisation of aflatoxin B1-bovine serum albumin (AFB1-BSA) conjugate on a polythionine (PTH)/gold nanoparticles (AuNP)-modified glassy carbon electrode (GCE). The surface of the AFB1-BSA conjugate was covered with horseradish peroxidase (HRP), in order to prevent non-specific binding of the immunosensors with ions in the test solution. The AFB1 immunosensor exhibited a quasi-reversible electrochemistry as indicated by a cyclic voltammetric (CV) peak separation (ΔEp) value of 62 mV. The experimental procedure for the detection of AFB1 involved the setting up of a competition between free AFB1 and the immobilised AFB1-BSA conjugate for the binding sites of free anti-aflatoxin B1 (anti-AFB1) antibody. The immunosensor’s differential pulse voltammetry (DPV) responses (peak currents) decreased as the concentration of free AFB1 increased within a dynamic linear range (DLR) of 0.6 - 2.4 ng/mL AFB1 and a limit of detection (LOD) of 0.07 ng/mL AFB1. This immunosensing procedure eliminates the need for enzyme-labeled secondary antibodies normally used in conventional ELISA–based immunosensors

The application of high capacity ion exchange absorbent material, synthesized from fly ash and acid mine drainage, for the removal of heavy and trace metals from secondary co-disposed process waters

Magister Scientiae - MScThe objective of this study was to investigate the feasibility of the application of low cost high capacity inorganic ion exchange material, synthesized form collected fly ash and acid mine drainage solid residues, for the decontamination of secondary co-disposal process waters, with emphasis on investigating the processes governing the solid/solution interface.South Afric

Isoform-dependent lysosomal degradation and internalization of apolipoprotein E requires autophagy proteins.

The human apolipoprotein E4 isoform (APOE4) is the strongest genetic risk factor for late-onset Alzheimer's disease (AD), and lysosomal dysfunction has been implicated in AD pathogenesis. We found, by examining cells stably expressing each APOE isoform, that APOE4 increases lysosomal trafficking, accumulates in enlarged lysosomes and late endosomes, alters autophagic flux and the abundance of autophagy proteins and lipid droplets, and alters the proteomic contents of lysosomes following internalization. We investigated APOE-related lysosomal trafficking further in cell culture, and found that APOE from the post-Golgi compartment is degraded through autophagy. We found that this autophagic process requires the lysosomal membrane protein LAMP2 in immortalized neuron-like and hepatic cells, and in mouse brain tissue. Several macroautophagy-associated proteins were also required for autophagic degradation and internalization of APOE in hepatic cells. The dysregulated autophagic flux and lysosomal trafficking of APOE4 that we observed suggest a possible novel mechanism that might contribute to AD pathogenesis. This article has an associated First Person interview with the first author of the paper