Doctoral Degrees (Chemistry)

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Structure-activity relationships of novel anti-diabetic ruthenium compounds : synthesis, characterization, mechanistic and in vitro studies.
(2024) Makanyane, Madikoloha Daniel.; Booysen, Irvin Noel.; Mambanda, Allen.
Type 2 Diabetes Mellitus (T2DM) is a chronic metabolic disorder which is globally responsible for millions of fatalities per year. Management of T2DM typically involves orally administered anti-hyperglycaemic drugs in conjunction with dietary interventions. However, the current conventional therapy seems to be largely ineffective as patients continue to develop complications such as cardiovascular diseases, blindness and kidney failure. Existing alternative treatment entails the administration of organic therapeutic pharmaceuticals, but these drugs have various side effects such as nausea, headaches, weight gain, and respiratory and liver damage. Transition metal complexes have shown promise as anti-diabetic agents owing to their diverse mechanisms of activity. In particular, selected ruthenium compounds have exhibited intriguing biological behaviours as Protein Tyrosine Phosphatase (PTP) 1B and Glycogen Synthase Kinase 3 (GSK-3) inhibitors, as well as aggregation suppressants for the human islet amyloid polypeptide (hIAPP). The introduction chapter served as a survey on studies pertaining to ruthenium compounds as metallo-drugs for T2DM. Herein, we also provide perspectives on directions to fully elucidate in vivo functions of this class of potential metallopharmaceuticals. More specifically, there is still a need to investigate the pharmacokinetics of ruthenium drugs in order to establish their biodistribution patterns which will affirm whether these metal complexes are substitutionally inert or serve as pro-drugs. In addition, embedding oral-administered ruthenium complexes into bio-compatible polymers can be a prospective means of enhancing stability during drug delivery. This chapter was concluded with a descriptive rationale of the research study as well as specifying the specific research aims and objectives. Our prior studies have illustrated that the uracil ruthenium(II) diimino complex, [Ru(H3ucp)Cl(PPh3)] (1) (H4ucp = 2,6-bis-((6-amino-1,3-dimethyluracilimino)methylene)pyridine) displayed high hypoglycaemic effects in diet-induced diabetic rats. To rationalize the anti-diabetic effects of 1 in the first experimental chapter, three new derivatives have been prepared, cis-[Ru(bpy)2(urdp)]Cl2 (2) (urdp = 2,6-bis-((uracilimino)methylene)pyridine), trans-[RuCl2(PPh3)(urdp)] (3), and cis-[Ru(bpy)2(H4ucp)](PF6)2 (4). Various physicochemical techniques were utilized to characterize the structures of the novel ruthenium compounds. Prior to biomolecular interactions or in vitro studies, the stabilities of 1 – 4 were monitored in anhydrous DMSO, aqueous phosphate buffer containing 2% DMSO, and dichloromethane (DCM) via UV-Vis spectrophotometry. Time-dependent stability studies showed ligand exchange between DMSO nucleophiles and chloride co-ligands of 1 and 3, which was suppressed in the presence of an excess amount of chloride ions. In addition, the metal complexes 1 and 3 are stable in both DCM and an aqueous phosphate buffer containing 2% DMSO. In the case of compounds 2 and 4 with no chloride co-ligands within their coordination spheres, high stability in aqueous phosphate buffer containing 2% DMSO was observed. Fluorescence emission titrations of the individual ruthenium compounds with bovine serum albumin (BSA) showed that the metal compounds interact non-discriminately within the protein's hydrophobic cavities as moderate to strong binders. The metal complexes were capable of disintegrating mature amylin amyloid fibrils. In vivo, glucose metabolism studies in the liver (Chang) cell lines confirmed enhanced glucose metabolism as evidenced by the increased glucose utilization and glycogen synthesis in liver cell lines in the presence of complexes 2 – 4. The second experimental chapter, reports on the formation and characterization of new diamagnetic ruthenium uracil mono-imine compounds: [(η6-p-cymene)RuII(L)Cl] (L = urpda = 5-((pyridin-2-yl)methyleneamino)-6-aminouracil) for 1, urdpy = 6-amino-1,3-dimethyl-5-((pyridin-2-ylmethylene)amino)uracil) for 2 or urqd = 5-((quinolin-2-yl)methyleneamino)-6-aminouracil) for 3); cis-[RuII(L)(bipy)2] (L = urpy = 5-((pyridin-2-yl)methyleneamino)uracil) for 4 and H2dadp = 5,6-diaminouracil for 5); trans-[RuII(L)(PPh3)Cl2] (L = urpda for 6) are described. Various physicochemical techniques were utilized to characterize the structures of the novel ruthenium compounds. Furthermore, the DPPH and NO radical scavenging capabilities of metal complexes (2 – 10) were investigated. UV-Vis spectrophotometry data of the time-dependent (for 24 h) studies show that 4 and 5 are stable in aqueous phosphate buffer containing 2% DMSO. Similarly, the stabilities of 1 - 3 and 6 monitored in chloro-containing and non-coordinating solvent dichloromethane show that they are kinetically inert, whereas, in a high nucleophilic environment, the chloride co-ligands of 1 - 3 and 6 were rapidly substituted by DMSO. In contrast, the substitution of the labile ligand of the complexes by DMSO molecules from its solution with a high chloride content was suppressed. Solution chemical reactivities of the different metal complexes were rationalized by density functional theory computations. Furthermore, the binding affinities and strengths between BSA and the respective metal complexes were monitored using fluorescence spectroscopy. Mutually, these metal complexes showed comparable capabilities of denaturing mature BSA aggregates which was established by fluorescence spectroscopy and Transmission Electron Microscopy (TEM). The final experimental chapter entails the encapsulation of the ruthenium complexes 1 - 10 into separate organic chitosan (CS)-polyvinyl alcohol (PVA) blends and the subsequent nanofabrication of their electrospun nanofiber (ENF) conjugates, Ru-CS-PVA ENFs. Intravenous injections of insulin can be regarded as a primitive method for Diabetes Mellitus management which characteristically leads to patients developing insulin resistance while oral-administered anti-diabetic organodrugs such as Metformin have exhibited low bio-availability and typically induce gastrointestinal (GI) side-effects. Although the intravenous injections of selected metal compounds in Streptozocin (STZ)-diabetic results have delivered promising results, limited work has been done to evaluate their efficiencies during oral administration. Herein, the fabricated chitosan (CS)-polyvinyl alcohol (PVA) electrospun nanofibers (CS-PVA ENFs) of the leading insulin-enhancing ruthenium complex 1, cis-[Ru(bipy)2(H4ucp)](PF6)2 and its analogs: 2 – 10. The Ru-CS-PVA ENFs nanocomposites were characterized by using (SEM-EDX), powder X-ray diffraction, and FTIR spectroscopy. The Ru-CS-PVA ENF nanohybrids exhibited randomly oriented fiber mat morphology with mean diameters in the range of 118 - 280 nm. Metal-based drug release kinetics of 1 - 10 from the ENF polymer matrix were measured spectrophotometrically at pH 1.5 and 7.4, respectively. Electronic spectral trends and data analysis over a 24-hour data collection period reveals variable dissolution rates with first-order rate (kobs) constants ranging from 0.0146 to 2.74 μM h-1 with accompanying hyperchromism effects between 5.69 to 37.6% at a pH of 1.5 while at a pH of 7.4, kobs value limits were 0.0104 and 3.89 μM h-1 rendering corresponding 19.14 and 87.32% hypochromic shifts. The release kinetics data of 1 - 10 were spontaneously released into the aqueous media from the Ru-CS-PVA ENFs, with the highest and releasing rates recorded for complexes 8 and 4, respectively.
Harnessing the power of Microalgae and Daphnia for bioremediation of nutrients, pharmaceuticals, and heavy metals in wastewater.
(2024) Tenza, Ntombiphumile Perceverence.; Mahlambi, Precious Nokwethemba.
Excess nutrients in aquatic ecosystems promote eutrophication, which significantly affects oxygen-dependent organisms. Furthermore, toxic microalgae, such as microcystin, cylindrospermopsis, etc., thrive during eutrophication, releasing poisonous compounds harmful to human health and other aquatic organisms. Pharmaceutical compounds and heavy metals in aquatic environments further exacerbate these global concerns. Thus, addressing such problems is paramount and aligns with Sustainable Development Goals (SDGs) 6 - Clean Water and Sanitation, 12 - Responsible Consumption and Production, and 14 - Life Below Water. Bioremediation of wastewater with biological microorganisms such as microalgae and daphnia provides an excellent solution due to their remarkable properties that enable them to efficiently eliminate many contaminants, including heavy metals, nutrients, and pharmaceuticals. So, this study explored the novel approach to nutrient removal by combining Chlorella spp. and Daphnia magna (D. magna). Chlorella spp. completely removed nitrate and nitrite from wastewater by converting them into compounds like amino acids, proteins, and lipids. When D. magna was employed alone, it faced significant challenges due to the absence of primary producers like bacteria and microalgae, which they mainly feed on. However, combining it with Chlorella spp. proved exceptionally effective, as 100% removal was obtained for nitrate and nitrite, possibly driven by D. magna grazing on Chlorella spp. that had assimilated nitrate and nitrite into their biomass. Challenges arose for ammonia and phosphate removal as they achieved up to 27% removals. This can be ascribed to nutrient release, Chlorella’s saturation capacity, and environmental changes such as pH. For pharmaceuticals, the study successfully developed and validated the LC-PDA method for separating sulfamethoxazole, nevirapine, and efavirenz. The optimum conditions were a mobile phase (90:10% acetonitrile: water with 0.1% formic acid), run time (8 minutes), flow rate (0.4 mL/min), and wavelength (220 nm). The study also developed and validated the solid-phase extraction (SPE) method for extracting analytes of interest in water matrices. The optimum conditions were conditioning solvent (mixture of acetonitrile and methanol in a 70:30 (v/v) ratio), sample volume (50 mL), and pH 7. The study then assessed Chlorella's capacity to remove sulfamethoxazole, nevirapine, and efavirenz. The obtained removal efficiencies for efavirenz, nevirapine, and sulfamethoxazole ranged from 0–60%, 5–51%, and 10–50%, respectively. Furthermore, the results exhibited lower removal efficiency (up to 15%) for higher concentrations (1 and 5 mg/L), whereas lower initial concentrations (0.5 and 0.25 mg/L) showed higher removal rates (up to 60%). Low removals could be due to factors like toxic metabolite accumulation and pharmaceutical toxicity. This study also explored copper, lead, and zinc adsorption capacity on Chlorella spp. biomass. Batch cultures were assessed in triplicate at 150 rpm in an orbital shaker under different biomass dosages, pH levels, contact times, and metal concentrations. The optimum conditions were pH 7, 60 minutes contact time, biomass dosage of 12.5 mg, and 0.5 m/L concentration. The optimal conditions yielded complete removal of lead and zinc, with copper reaching up to 80% removal. The study also assessed the adsorption of target heavy metals employing Freundlich, Langmuir, and Temkin isotherms, with the Langmuir isotherm better fitting copper (R2 = 0.9888) while the Freundlich isotherm best-fitted lead (R2 = 0.976) and zinc (R2 = 0.968). Lead and zinc favoured the pseudo-first-order kinetic model, whereas copper favoured the pseudo-second-order kinetic model. Thermodynamic studies exhibited an endothermic and spontaneous process for copper and zinc. The results of this PhD underscored Chlorella's potential as an environmentally safe and effective option for removing nutrients, pharmaceutical compounds, and heavy metals. Mechanisms for removal included surface adsorption, photodegradation, bioaccumulation, and enzymatic degradation. The Fourier transform infrared spectroscopy (FTIR) confirmed the existence of functional groups like alkene, amide, carbonyl, carboxyl, ethers, hydroxyl, and methyl, which participate in the adsorption of these contaminants through various interactions. Surface morphology analysis through scanning electron microscopy (SEM) shows changes in Chlorella spp. cells after exposure to target compounds (nutrients, pharmaceutical compounds, and heavy metals), suggesting the possibility of interaction that aids their removals. Thus, this study contributed valuable insights for improving wastewater treatment strategies and addressing water scarcity concerns. Additionally, it promotes a circular economy as Chlorella spp. and daphnia biomass can be harvested at the end of the treatment process for diverse uses, including biogas production, organic fertiliser, animal feed, etc. Going forward, future research should focus on optimising bioremediation by exploring different combinations of microalgae and other biological agents to enhance the removal efficiencies of heavy metals and pharmaceuticals. Moreover, genetic modification of Chlorella spp. to improve resilience and uptake capacity is crucial, and integrating advanced monitoring technologies like biosensors are promising directions. In-depth studies on removal mechanisms, such as adsorption, photodegradation, bioaccumulation, and enzymatic degradation, are essential. Also, scaling up to pilot and full-scale applications is crucial for evaluating feasibility and economic viability. Lastly, collaboration with industrial partners and policymakers can help develop regulatory frameworks and incentives. These efforts can advance bioremediation and support global SDGs related to clean water, responsible production, and life below water.
Adsorption of pharmaceuticals by nano-molecularly imprinted polymers (nano-MIPs) from wastewater: kinetics, isotherms, and thermodynamics studies.
(2024) Nxumalo, Nonhlazeko Loveday.; Mahlambi, Precious Nokwethemba.; Mahlambi, Mphilisi.; Mngadi, Sihle.; Chokwe, Tlou.
It has been reported that pharmaceuticals are not entirely removed or broken down during the wastewater treatment process, allowing them to escape into effluent water. This stems from the pharmaceuticals widespread use and the inefficient wastewater treatment methods. Therefore, the objective of this study was to develop more effective methods for removing pharmaceuticals from wastewater systems. Adsorption-based pharmaceutical removal is one of the most promising approaches because it is easily incorporated into current water treatment systems. The first part of this work reports on literature studies for recent advancements in the adsorption process involving the incorporation of an artificial molecularly imprinted polymer (MIP), that is an effective molecular receptor that can selectively recognize and remove pollutants. In magnetic solid-phase extraction, dispersive solid-phase microextraction, and solid-phase extraction, MIPs can be used as a selective adsorbent for analyte cleanup and preconcentration. Moreover, MIPs can be produced by combining nanoparticles to develop composite nanomaterials (nanoMIPs). In comparison to conventional bulk adsorbents, the enhanced selective adsorption capacity and kinetics are attributed to the large surface area per unit volume and specific functionality of nanomaterials. Nonetheless, some significant barriers to the application of nanomaterials are their dispersive qualities, difficulty in cycling, and secondary pollution from the loss of adsorbent during treatment. Another way to use nanoparticles for detectability enhancement is to modify the molecularly imprinted polymers chemical or physical characteristics. The nanoparticles' embedding in the MIP enhances the material's surface area or gives the adsorbent new features. This study describes a method for creating reusable, economical, and effective polymer-based silver nanoparticles-adsorbents. Notably, silver nanoparticles have a wide range of applications due to their unique properties which include their large surface area, shape and size. Plant-mediated synthesis plays a significant role in their synthesis. Remarkably, the synthesis of silver nanoparticles from plant extracts is inexpensive, easily scalable, and harmless for the environment. Plant extracts can be used to produce nanoparticles with controlled sizes and shapes. The molecular imprinting technique was used to create species-specific functionalities like carboxylic acid (-COOH) on a polymer surface. MIPs offer several advantages, including large surface area, targeted functionalities for high reactivity, and the ability to minimise nanoparticles from leaking into the surrounding environment when MIP-based adsorbents are being handled. To further comprehend the behaviour of adsorbents and the adsorption process, kinetics, thermodynamics, and isotherm models were explored. The second part of the work involved synthesizing the MIPs for efficient and selective removal of pharmaceuticals from specific groups. All target compounds were employed as multiple templates in a bulk polymerization process carried out at 70 °C to synthesize MIPs. Additional reagents utilized in the synthesis included toluene as a porogenic solvent, ethylene glycol dimethacrylate as cross-linker, 1,1'-azobis-(cyclohexane carbonitrile) as an initiator and 2- vinyl pyridine as functional monomer, respectively. The synthesis of a non-imprinted polymer (NIP) was conducted without templates, using reaction conditions similar to those of MIP. Furthermore, following the synthesis, the polymers were characterized using X-ray diffraction, thermogravimetric analysis, differential scanning calorimetry, Fourier transform infrared spectroscopy, and scanning electron microscopy. Liquid chromatography-mass spectrometry (LC-MS) was successfully used to develop an analytical method for detection and quantification of the target pharmaceuticals. The method yielded quantification limits of 0.42 to 0.75 mg L-1 and detection limits of 0.14 to 0.25 mg L-1 for the target pharmaceuticals. The synthesized polymer attained maximum matrix-matched adsorption capacities of 3.89, 4.97 and 3.40 mg g-1 for sulfamethoxazole, nevirapine and ibuprofen, respectively within 10 minutes. Competitive adsorption of the target pharmaceuticals demonstrated a link between adsorption and the pharmaceuticals pKa, log Kow, and molecular size. Studies on batch adsorption and kinetics revealed that the binding of pharmaceuticals to the MIP particles suited the pseudo-second order kinetics, leading to various interactions through chemisorption. The data also fitted well in Langmuir isotherm which meant that the target pharmaceuticals adsorption occurred on the homogeneous binding sites of the MIP. Furthermore, the thermodynamic data demonstrated the adsorption process's endothermic and spontaneous nature. Notably, the synthesized MIP was highly selective and its application in environmental studies led to the development of a less expensive analytical method. Moreover, the MIP particles that had been generated were recovered to be reusable up to five cycles with removal efficiency >90%. The third part involved incorporating silver nanoparticles (AgNPs) into MIPs using ibuprofen, nevirapine, and sulfamethoxazole as templates. In this part, starch (St) and macadamia nutshells (MCD) were employed in the synthesis of AgNPs as reducing and stabilizing agents. Following that, each of these AgNPs was incorporated with MIP, and the most effective combination was identified through comparison. The synthesized adsorbents were further optimized for the adsorptive removal of selected target pharmaceuticals. The % removal efficiencies were greater than 70%, indicating that the adsorbents are suitable for use in water treatment processes. The material's adsorption mechanisms and performance were examined through the application of various kinetics and isotherm models. Both the St and MCD-AgNPs experimental data fit to Freundlich and Langmuir adsorption isotherms. However, based on the somewhat higher correlation coefficients, the Langmuir isotherm model provided a better fit. The St/MCD nanoMIPs best suited the Freundlich model, indicating that the adsorption occurred on the multilayer heterogeneous surface. Further, both the St/MCD nanoMIP adsorbents underwent spontaneous, endothermic adsorption, as demonstrated by the thermodynamic data, whereas the behaviour of the kinetics was effectively anticipated by pseudo-second order model, which suggested adsorption through chemisorption. Accordingly, large internal surface area, greater loading capacity, thermal stability, and reusability were among the advantageous properties of the nanoMIPs adsorbent materials. Moreover, both adsorbents showed improved qualities and were highly selective and effective in removing the selected pharmaceuticals in wastewater. As a result, during the course of five adsorption/desorption cycles, the St/MCD nanoMIPs show a removal efficiency of more than 90%. As a result, they demonstrated proficiency in efficient application. The fourth part involved the incorporation of MIP with Platanus acerifolia and Moringa oleifera silver nanoparticles. Using plants to synthesize AgNPs is a more cost-effective and lowmaintenance method; in contrast, using other organisms requires a particular medium and a specific amount of time. Therefore, the leaves of both the platanus acerifolia (PL) and moringa oleifera (MO) served as stabilizing and reducing agents during the synthesis of AgNPs. Each optimized parameter that could influence the adsorption potential, such as temperature, adsorbate concentration, pH, adsorbent dose, and contact time, was examined in relation to the removal effectiveness of the MO/PL nanoMIP adsorbents. These evaluated parameters were optimum at pH 7, concentration of 0.2 mg/L and contact time of 10 minutes for both MO and PL-nanoMIPs, mass dosage of 30 mg and 20 mg, and temperature of 40 and 30 °C for MO and PL-nanoMIP, respectively. Further, the maximum removal efficiencies obtained at these optimum conditions were >97% for both MO-nanoMIP and PL-nanoMIP. The adsorption experimental data for both MO/PL-AgNPs and MO/PL-nanoMIPs nano-adsorbents fitted with the linear Langmuir model which suggests that the binding took place on the homogenous monolayer surface. Additionally, compared to MO/PL-AgNPs, the MO/PL-nanoMIPs adsorption capacities for the target pharmaceuticals were higher, suggesting that the nanoMIPs larger surface areas contribute to their enhanced adsorption capacity. The linear pseudo-second order kinetic model best fitted on MO/PL-nanoMIPs which implied adsorption through chemisorption, whereas the thermodynamic data demonstrated that the adsorption process was endothermic and spontaneous. Moreover, the values of ΔH° for the MO/AgNPs were less than 40 kJ/mol and more than 40 kJ/mol for the MO/PL-nanoMIPs. This therefore confirmed that the MO/AgNPs was dominated by physical adsorption whereas the MP/PL-nanoMIPs was dominated by chemical adsorption. The MO/PL-nanoMIPs confirmed the high efficiency for the removal of target pharmaceuticals in wastewater. Upon recycling the adsorbents for five cycles, it was noted that the MO-nanoMIP adsorbent was effective continued to remove 86.7- 88.8% and 97-98% for PL-nanoMIP even in the fifth cycle. Indeed, the removal of sulfamethoxazole, nevirapine, and ibuprofen by nanoMIP adsorbents has demonstrated the importance of the surface area, structural stability, pore size and the electrostatic interactions brought about by the charges on the nanoMIPs surface. Consequently, among the investigated nanoMIP adsorbents, PL-nanoMIP demonstrated strong adsorption capacities for the targeted pharmaceuticals due to it large surface area and narrow size distribution as compared to the other nanoMIP adsorbents. The usability of plant leaves as a reducing and capping agent for nanoparticles as well as the recycling of nanoMIPs has the potential to transform waste that is no longer useful into valuable pollutants adsorbents. This would solve the problem of waste disposal and have beneficial impacts on the environment pollution and the economy. Notably, the nanoMIPs synthesized in this study are highly selective, reusable adsorbents that are cost effective and environmentally friendly. In contrast, as a substitute for more costly synthetic materials, these nanoMIPs are a promising material for the removal of different classes of pharmaceuticals in wastewater treatment plants and they can possibly be applied on a large scale.
Extraction of pesticides using selected analytical methods from soil and maize segments : cumulative and health risks assessment.
(2024) Zondo, Sandisiwe Gladness.; Mahlambi, Precious Nokwethemba.
Increased agricultural operations result in increased usage of various pesticides to safeguard agricultural crops, however this is done without paying attention to the effects of the amounting potential harm both humans and the environment are exposed to. In this present study, a structured study was conducted to investigate the uptake of atrazine, mesotrione, 2,4- dichlorophenoxyacetic acid and glyphosate herbicides from contaminated soil and their translocation into different maize segments. Soil profile and quality of irrigation water were also assessed as they are crucial resources required in agricultural crop production due to their ability to influence the yield and quality of the agricultural products. Various physicochemical parameters were measured in an attempt to monitor the soil profile, irrigation water and maize quality harvested from Buhle farm located in Howick, KwaZulu-Natal Province. The irrigation water physicochemical parameters considered were the pH, electrical conductivity, alkalinity and chloride concentration. The soil physicochemical parameters considered were moisture content, pH, electrical conductivity, texture, total nitrogen as well Mg, Na, K, Zn, Mn, P and N elements. Maize was analysed for nutrition content and medicinal health promoting compounds. Based on the attained results, the soil texture contained high clay content (56.4%), followed by sand (40.6%) and silt (2.98%). The concentrations for total nitrogen, phosphorus and potassium which were translated to high soil fertility were 2700, 19 and 222 mg L-1, respectively. These particular elements are essential for agricultural plantation processes and consequently maize quality and maize yield. The levels of sodium, sodium adsorption ratio and electrical conductivity found in irrigation water were 0.05 mg L-1, 2 and 1.81 μS m-1, respectively. The findings showed that maize harvested from Buhle farm had high starch content of 58.6%. Fibre, protein and fat contents in maize were 23.4, 9.01 and 4.55%, respectively. Furthermore, the total anthocyanin, total flavonoids and total phenolic acid content were 8.5, 49.5 and 100 mg L-1, respectively. High amounts of phenolic acid detected indicated therapeutic ability of the maize since phenolic acids are essential for cancer prevention to the consumer. The presence of anthocyanin, flavonols and phenolic acids in maize crop is associated with its quality that could benefit livestock and human after consumption. The analysis of herbicides in soil and maize samples require sample pre-treatment due to their low concentration and complex matrix hence an ultrasonic extraction, microwave-assisted extraction (MAE), Soxhlet extraction (SE) and QuEChERS methods were investigated. The optimization and application of ultrasonic extraction, MAE, SE and QuEChERS methods were conducted for the effective extraction of pesticides from maize and their corresponding soil samples. The analysis of pesticides (atrazine, glyphosate, 2,4-dichlorophenoxyacetic acid and mesotrione) was done with gas chromatography-flame ionization detector. Factors influencing the efficiency of the extraction methods such as the extraction solvent, extraction time, solvent volume, sample wetting and spiking concentration were assessed. Under the optimum experimental conditions, the relative standard deviation (RSD), coefficient of determination (R2), limit of detection (LOD), limit of quantification (LOQ), and percentage recoveries were the quantitative characteristics of the current methods assessed. All calibration curves showed a high correlation coefficient (R2) ≥0.996, indicating good linearity. The LODs and LOQs ranged between 0.22-0.32 μg L-1 and 2.0-2.9 μg L-1 for SE , 0.1-0.25 μg L-1 and 1.1-2.2 μg L- 1 for MAE, 0.02 – 0.15 μg L-1 and 0.2 - 0.5 μg L-1 for UE and 0.01 – 0.23 μg L-1 and 0.13 – 0.8 μg L-1 for QuEChERS. The maize and soil analytes recoveries for SE, MAE, EU and QuEChERS ranged between 62-80% and 70-81%, 80-98% and 85-101%, 100-104% and 91- 97 % and 94-115% and 92-101%, respectively with the repeatability, articulated as RSD values of which are within the acceptable range as they are lower than 20%. MAE method showed higher sensitivity compared to SE while, UE and QuEChERS both showed high sensitivity for the extraction and quantification of the target analytes at low concentrations found in soil and maize cob. It was observed that 2,4-dichlorophenoxyacetic acid (2.4-D) was least absorbed by the soil, however, all the studied herbicides showed high absorption in the leafy segment of the maize plant due to the high polarity of the leaf cuticle. Glyphosate showed high absorption rate in soil, roots, stalk and leaves while mesotrione was highly absorbed in corn and tassels in all treatments. The absorption rate of analyte increased with increasing growth days. The higher treatment concentration (0.75 g L-1) showed elevated accumulation with the highest concentration (1.02 μg L-1) observed for glyphosate in leaves after 140 days and high mesotrione in corn (0.51 μg L-1) and tassel (0.42 μg L-1) observed after 120 days. Even though all maize treatment showed a pesticide toxicity index (PTI) values of <1, the health risk index (HI) data were below 100% threshold as well indicating no possible health risk linked with the intake of these crops by both adults and children.
Phytochemistry of some South African medicinal Rubiaceae and Curtisiaceae species.
(2024) Moyo, Prince Nqaba.; van Heerden, Fanie Retief.
The investigation began by identifying which taxonomic groups from southern Africa are most prominently featured in ethnopharmacological practices. This involved collecting data from ethnopharmacological surveys and performing statistical analyses. Over-represented southern African taxa and the concept of over-represented genera are presented for the first time. The families and genera with the highest margins of over-representation were found to be Loganiaceae and Albizia in southern Africa as a whole and Sapotaceae and Solanum in South Africa. The families and genera with the highest margins of over-representation across disease categories are Ebenaceae and Albizia, Canellaceae and Dicoma, Combretaceae and Pterocelastrus, Ebenaceae and Bersama, Francoaceae and Erythrina, and Aristolochiaceae and Strychnos for plants used in the treatment of STIs, febrile and mosquito-vector diseases, microbial infections, pain, skin conditions, and female sexual/reproductive problems, respectively. The Rubiaceae family was found to be one of the 25 most over-represented taxonomic groups in Southern Africa. Plants in the family were targeted for investigation due to the availability of plant material. The study focused on the phytochemical investigations of three southern African medicinal plants, Coddia rudis (E.Mey. ex Harv.) Verdc., Keetia gueinzii (Sond.) Bridson and Curtisia dentata (Burm.f.) C.A.Sm. The ethnopharmacology and phytochemistry of the Rubiaceae species were first reviewed. The review was intended to be comprehensive, recording and discussing the notable findings of investigations on Rubiaceae species. Hundred and thirteen Rubiaceae species with ethnobotanical applications. Thirty-five species have nutritional applications. Twenty-three out of 34 anti-inflammatory and analgesic species, 17 out of 31 anti-viral and anti-bacterial species, and 13 of 27 Rubiaceae species used for sexual and reproductive problems have not undergone preparative phytochemical analysis. Only 52 southern African Rubiaceae were found to have undergone bioactivity investigations. Two hundred and thirty-three isolated compounds have been reported from 39 Rubiaceae species of southern Africa. Coddia rudis and Keetia gueinzii were selected based on proximal availability in the Pietermaritzburg region. Curtisia dentata was chosen due to findings during the review that plants in the Cornales order (of which C. dentata is a member) produce iridoid glycosides similarly to the Rubiaceae family members. The study yielded eleven bioactive known compounds. Gardenoside, geniposidic acid, and astragalin were isolated from C. rudis leaves and kaempferitrin was isolated from K. gueinzii leaves. Phlorizin and (-)-epicatechin were isolated from C. dentata bark, and loganic acid, secologanoside, geniposide, juglalin, and sweroside were isolated from C. dentata leaves. The study represents the first report of the isolation of these compounds from the species in question. The isolation of phlorizin, a known diuretic with anti-diabetic activities, supports the use of the C. dentata bark to treat diabetes in traditional medicine. Furthermore, obtaining juglalin, which has been reported to inhibit the senescence of dermal fibroblast cells, corroborates the application of the plant in the treatment of various skin conditions. The results of the study are consistent with findings reported in literature that show the presence of biologically active iridoid glycosides in Rubiaceae and Cornales species.
Synthesis elaboration of fragments that potentially inhibit the HOP-HSP90 protein-protein interaction.
(2024) Molefe, Patience Snenhlanhla Sthembile.; Sithebe, Siphamandla.; Veale, Clinton Gareth Lancaster.
Heat Shock Protein 90 (HSP90) is a molecular chaperone that mediates the stability and maturation of many important proteins for oncogenesis. There is an overexpression of the Heat Shock Protein 70/Heat Shock Protein 90 Organising Protein- HSP90 (HOP-HSP90) protein-protein interaction (PPI) complex in tumour tissues unlike in healthy cells. This PPI complex of HSP90 displayed a potential druggable target because of the crucial role it plays in cancer development. However, the challenge is the development of HOP-HSP90 PPI inhibitors. The literature showed the activity of valsartan (27) for the inhibition of HOP-HSP90 PPI as it entails the features of ortho-biphenyl tetrazole fragments that were obtained from the Structural-Binding Relationship (SBR) of the active fragments using fragment-based drug discovery (FBDD). These fragments bound to the tetratricopeptide repeat 2A (TPR2A) domain of HOP and inhibited the PPI of HOP-HSP90. As a result, this study aimed to synthesise and assay ortho-biphenyl tetrazole fragments as inhibitors of HOP-HSP90 for novel anticancer inhibitors, triple-negative breast cancer (TNBC). Valsartan (27) and its analogues were synthesised following reported procedures and modified methods. A series of 13 ortho-biphenyl tetrazole desired fragments were successfully synthesised using a Suzuki-Miyaura cross-coupling reaction and [3 + 2] cycloaddition of nitrile with sodium azide. The cross coupling of 2-iodobenzonitrile or 2-(2-bromophenyl) acetonitrile with para-substituted phenylboronic acid was conducted using different substrates including Cl, Br, F, CH3, CF3, H, and OCH3. Cycloaddition was done after the cross-coupling to skip the protection step of the tetrazole. With the desired ortho-biphenyl tetrazole fragments in hand, the PPI inhibition activity was evaluated at different concentrations from 0 mM to 2.0 mM. It is interesting to observe that some of these fragments showed PPI activity at different concentrations including compounds 76, 80, 82, 83 and 84. No activity was observed following the incorporation of the benzylic carbon. The data presents the successful lead optimisation for the development of HOP-HSP90 novel PPI inhibitors for the treatment of TNBC.
Quantum effects in the nerves and brain.
(2024) Adams, Betony Lascelles.; Sinayskiy, Ilya.; Petruccione, Francesco Portia.
Quantum biology is often referred to as an emerging field of research. In theory it shares its roots with the more general field of quantum physics. Many of the founding figures of quantum theory were intrigued as to whether its insights into the structure of matter might equally offer insights into living matter. Experimental evidence for quantum effects in biological systems took longer to emerge, with tunnelling in enzymes observed in the 1960s. More recently, advances in ultrafast experimental techniques have led to extensive investigation into the role of quantum coherence in energy and charge transfer in photosynthesis. Despite this long history, the role that quantum effects play in biological systems is still very much up for debate. Even more debatable is the role that quantum effects may play in that most complex biological system: the brain. Penrose and Hameroff, for example, have suggested that consciousness cannot be explained by classical processes, and their Orchestrated Objective Reduction theory has generated both interest and critique. Consciousness is the brain’s most profound secret and it remains to be seen whether quantum mechanics will prove a likely explanation. But, less ambitiously, the brain can also be described as a collection of nerve cells, whose function involves physiological processes similar to those in other cells. The aim of this thesis is to investigate how progress made in quantum biology might be applied to the specific context of neurology. To this end, the thesis revisits two of the models currently employed in quantum biological research. The first of these is the Posner molecule model of cognition, developed by Matthew Fisher. This hypothesis involves the entangled spins of phosphorus nuclei in calcium phosphate molecules, which have an influence on the balance of free calcium ions and thus neural activation. This original model is further developed here to investigate how entanglement and coherence are altered by the inclusion of lithium isotopes, and whether this might offer an explanation for the mode of action of lithium in treating bipolar disease. The second model investigated in this thesis is the vibration-assisted tunnelling model first developed in the context of olfaction. The hypothesis here is that olfactory receptors are potentially activated by an electron transfer that is facilitated by the vibrational modes of the olfactant. Ligand-receptor interactions are ubiquitous in biological systems and not least in the effective functioning of the nervous system. This thesis thus re-examines the vibration-assisted tunnelling model to determine how generalisable it might be, by taking the specific case of infection with the SARS-CoV-2 virus. While this virus-host interaction is not neurological, intriguing evidence that antidepressants can have antiviral effects as well as the profound effects that COVID-19 can have on the nervous system, suggests that this timely example might offer valuable neurological insights.
Derivatised phenanthroline transition metal chelates : targeted chemotherapeutic agents.
(2024) Hunter, Leigh André.; Akerman, Matthew Piers.
The derivatisation of 1,10-phenanthroline at the 2-position afforded two classes of compounds with two different bridging groups in this study. The first group comprised two amide-bridged tetradentate N4-donor ligands and were chelated to copper(II), nickel(II) and palladium(II). The ligand chelation occurred with concomitant deprotonation of the amide N-H, resulting in a monoanionic ligand and monocationic complexes when coordinated to the divalent metal ions. The ligands N-(quinolin-8-yl)-1,10-phenanthroline-2-carboxamide, HL1, and N-(pyridin-2-ylmethyl)-1,10-phenanthroline-2-carboxamide, HL2, were characterised by NMR, IR and UV/vis spectroscopy as well as mass spectrometry. The second class of compounds were imine-bridged copper(II) chelates. These chelates were synthesised via a templating condensation reaction between various salicylaldehyde derivates and 1,10-phenanthrolin-2-ylmethanaminium chloride, yielding eight additional copper(II) chelates. The metal chelates were characterised by IR, UV/vis and EPR spectroscopy, and mass spectrometry. HL1, [Cu(L4)(NO3)] and [Cu(L7)](NO3) were further studied by X-ray diffraction. The copper(II) chelates exhibit two different solid-state structures with the nitrate counter ion coordinated to the metal centre in [Cu(L4)(NO3)], but in the outer coordination sphere for [Cu(L7)](NO3). The paramagnetic copper(II) chelates were studied with EPR spectroscopy, which confirmed the square planar coordination geometries of these chelates in solution. The metal chelates were designed to be chemotherapeutic agents, exerting their cytotoxicity through DNA intercalation and, for the copper(II) chelates, DNA cleavage through the catalytic production of ROS. The ability of the copper(II) chelates to catalyse the production of hydroxyl radical in situ in the presence of ascorbic acid and hydrogen peroxide was studied via a hydroxyl radical assay using Rhodamine B as an analogue for the aromatic DNA bases. Competitive binding studies determined the affinity of the metal chelates towards ct-DNA, [Cu(L1)](PF6) has the highest binding constant: 5.91 × 106 M-1. DFT calculations were performed on the ligands and metal chelates to determine the geometry-optimised structures, vibrational frequencies, 1H and 13C NMR chemical shifts and electronic transitions. The B3LYP/6-311G (d,p) level of theory was used for the ligands, copper(II) and nickel(II) chelates and the B3LYP/LanL2DZ level of theory for the palladium(II) chelates. The TD-DFT method was used for the energy calculations. The experimental and calculated results were compared where possible, and a reasonable correlation was found. The cytotoxicity of five amide-based chelates was evaluated against four human cancer cell lines, namely A549, TK-10, HT29 and U251, using an MTT assay. The screened chelates exhibited favourable anticancer activity with the mean IC50 values against the four cancer cell lines ranging from ca. 12 to 35 μM. Importantly, it was found that the combination of the copper(II) ion and the ligand was essential for enhanced cytotoxicity. The complex [Cu(L1)](PF6) was identified as the lead drug candidate based on the high DNA affinity and cytotoxicity. This compound was most cytotoxic towards the glioblastoma cell line U251 with an IC50 value of 7.59 μM. The imine-based chelates were screened against three human cancer cell lines: MDA-MB, HELA, and SHSY5Y, and a healthy human cell line, HEK293. The selectivity index of these chelates for neoplastic versus the healthy cell line was calculated. The imine-based chelates showed a high selectivity towards the triple-negative breast cancer MDA-MB, an order of magnitude more toxic to the tumour cell than the healthy one. This selectivity index is significantly improved over that of cisplatin. A gel mobility shift assay investigated the interactions between the copper(II) chelates and plasmid DNA. The in vivo biodistribution of [Cu(L1)](PF6) was determined using the copper-64 radiolabelled analogue of [Cu(L1)]Cl and microPET-CT scanning. The initial biodistribution studies suggested that the complex has good serum stability and showed that there was no significant accumulation in any organs. The subsequent study involved a xenograft model using the A549 cell line and showed significant uptake and retention of the complex in the tumour. The cytotoxicity of the chelate when synthesised with the non-radioactive isotopes of copper and the uptake of the radiolabelled equivalent in a tumour model suggest that this complex could have application as a “theranostic agent”.
Metallophthalocyanine-based electrochemical sensors for accurate qualitative and quantitative analysis of emerging pollutants in water resources.
(2024) Shoba, Siyabonga Blessing.; Booysen, Irvin Noel.; Mambanda, Allen.
Water is a precious resource and safeguarding it from pollution is paramount to ensure the well-being of both the environment and human health. Emerging contaminants such as pharmaceuticals and heavy metals pose significant threats, necessitating vigilant monitoring and appropriate action. Traditional laboratory-based analytical techniques like Gas Chromatography, ICP-OES and HPLC have been instrumental in quantifying pollutants. However, their high operational costs, maintenance requirements and the need for specialized personnel limit their widespread use, especially in resource-constrained countries. Electrochemical sensors have emerged as a promising solution. They provide real-time, portable and cost-effective options for on-site detection of pollutants in water. Current advancements in electrochemical sensors are centred around achieving selective detection using chemical modifiers, all while maintaining electrocatalytic sensitivity and reproducibility. These sensors can be tailored to target specific contaminants, making them highly efficient tools for monitoring water quality and ensuring the sustainability of this invaluable resource. In the first experimental chapter, a glassy carbon electrode (GCE) was modified by an asymmetric metallophthalocyanine (MPc) complex, A3B-CoPc-flav (where A = flavonyloxy substituent and B = an alkynyloxy substituent/molecular mast). The modification of an electrode was achieved via electrochemical grafting followed by clicked chemistry between the diazonium-functionalized GCE and the a-CoPc-flav3 to afford the GCE|clicked-a-CoPc-flav3. The chemically modified electrodes (CME) were utilized as electrocatalytic detectors for dopamine (DA) under optimized conditions. The response of the GCE|clicked-a-CoPc-flav3 was linear in the concentration range of 2 μM to 14 μM, attaining limits of detection and quantification of 0.311 and 0.942 μM, respectively, and high reproducibility (%RSD of 2.25%, N = 3). Interference studies were conducted, revealing a marginal shift in the DA peak potential in the presence of interfering substances. Despite this shift, the peak current intensity of DA remained largely unaffected, affirming the selectivity and accuracy of the CME. The analytical capabilities of the CME were further assessed using real water samples. The obtained percentage recoveries of (97.1%) of DA by the GCE|clicked-a-CoPc-flav3 and the well-established HPLC-MS method (113%) are both within the acceptable range of 80-120%. In the second experimental chapter, a platinum electrode (Pt) was modified via the electropolymerization of polypyrrole (PPy) after its co-electrodeposition of tetra-[4-((1H benzotriazole)methoxy)phthalocyaninato]cobalt(II) (CoPc-Bzt). The electrodeposition of CoPc-Bzt was performed in 1:1 DMF/acetonitrile containing 1 M tetrabutylammonium hexafluorophosphate (NBu4PF6) electrolyte over 20 cycles using cyclic voltammetry to afford a Pt|PPy/CoPc-Bzt (Bzt = benzotriazole). The resultant CME was prone to fouling by the analyte of interest, mercury(II). Due to fouling the differential pulse anodic stripping voltammetry (DPASV) was used to detect Hg(II) using the Pt|PPy/CoPc-Bzt within 10 μM to 100 μM linear range. The LOD and LOQ were found to be 3.11 and 10.00 μM, respectively. Interference studies illustrated that the detection capabilities of the CME were not affected by the presence of other heavy metal cations. The analytical performances of Pt|PPy/CoPc-Bzt (97.4%) and Inductively coupled plasma – optical emission spectroscopy (ICP-OES) (112.3%) are both within the acceptable range of 80-120%. In the third experimental chapter, the Pb electrocatalytic sensing capability of a gold electrode modified via the adsorption of electrospun nanofibers (ENFs) and Nafion (Nf) as an annealed conductive top-layer was evaluated. The fabricated ENFs comprised of a core polymeric nanocomposite of tetra-4-(3-oxyflavonephthalocyaninato)cobalt(II) (CoPc-flav), the carboxylic acid functionalized multiwalled carbon nanotubes (f-MWCNTs) and polyaniline (PANI) encapsulated in a polyvinyl acetate (PVA) ENFs. The resultant CME, Au|ENFs-1-Nf was not prone to fouling as was found when using the bare and the other constructed CMEs whose signal stabilities were compromised by background electrolyte currents. The Au|ENFs-1-Nf electrode could detect the Pb(II) cations in a reproducible manner (%RSD of 3.92%, N = 3) ranging from 8 to 125 μM, and limits of detection and quantification of 0.51 and 1.55 μM were obtained, respectively. However, the interference studies illustrated that the detection capabilities of the CME are severely compromised by the presence of other heavy metal cations. The analytical performance of the CME rendered a comparable percentage recovery (103%) with that of the ICP-OES (115%). In the fourth experimental chapter, the nanofabrication and characterization of new conductive materials, PANI-CoPc-fur (1) ((PANI = polyaniline and CoPc-fur = tetra-4-(2-furanmethylthiophthalocyaninato)Co(II)) and PANI-CoPc-fur-f-MWCNTs (2) are reported. Subsequently, an electrospun nanofiber (ENF) composite was fabricated where the core comprised of 2 that was encapsulated with a PVA shell. The resultant nanoconjugate, ENFs-2 was adsorbed on a glassy carbon electrode (GCE) followed by the immobilization of a permeable adhesion top layer of Nafion (Nf) to render the chemically modified electrode, GCE|ENFs-2-Nf. The classical physical properties of the electron-mediating layer for the CME synergistically aided in promoting its electrocatalytic activities. Consequently, the CME showed greater anodic and cathodic cyclic voltammetry (CV) peak currents compared to the bare GCE and other modified electrodes, indicating its higher sensitivity to acetaminophen (APAP), an emerging water pollutant of concern. Limits of detection and quantification (LOD and LOQ) values for APAP attained by squarewave voltammetry (SWV) were lower compared to those acquired using other electrochemical techniques. The detection of APAP at the GCE|ENFs-2-Nf attained by squarewave voltammetry (SWV) was linear from 10 to 200 μM of APAP and was reproducible (%RSD of 3.2%, N = 3). The respective calculated LOD and LOQ values of 0.094 and 0.28 μM were lower compared to those acquired using other electrochemical techniques. Analysis of APAP in the presence of commonly associated interferences metronidazole (MTZ) and dopamine (DA) illustrated a significant separation between the SWV peak potentials of APAP and MTZ, whereas there was some degree of overlap between the SWV current responses of APAP and DA. The analytical performance of the GCE|ENFs-2-Nf rendered a comparable percentage recovery (103.8%) with that of liquid chromatography–mass spectrometry (LC–MS) (106%).
Cytotoxic and mechanistic studies of novel phenanthroline-derived oxovanadium(IV) complexes.
(2022) Barry, Kristy-Lyn.; Munro, Orde Quentin.
In this work, a new series of ternary oxovanadium(IV) complexes of the type [VO(ONO)(DPQ/DPPZ)], [VO(ONN)(PHEN/DPQ)](PF6) and [VO(ONN)(DPPZ)](Cl) have been synthesised and characterised for the purpose of developing novel anticancer agents. ONO represents a tridentate Schiff base ligand derived from salicylaldehyde and various 4-substituted-2-aminophenols. ONN represents a tridentate Schiff base ligand derived from 1-methyl-2-imidazolecarboxaldehyde and various 4-substituted-2-aminophenols. PHEN represents 1,10-phenanthroline, DPQ represents dipyrido[3,2-d:2′,3′-f]quinoxaline and DPPZ represents dipyrido[3,2-a:2′,3′-c]phenazine. The bidentate N,N-donor polypyridyl co-ligands were incorporated to enhance DNA binding and to stabilise the vanadium(IV) metal centre. The known N,N-bidentate ligands were synthesised and characterised by ESI-mass, 1H and 13C NMR spectroscopy. The tridentate O,N,O′ and O,N,N′ ligands were synthesised and characterised by ESI-mass, NMR, IR, and UV/visible spectroscopic techniques, elemental analysis and single crystal X-ray crystallography. Single crystal X-ray diffraction, 1H NMR and DFT simulations confirmed the O,N,N′ ligands in this work form dimeric hydrogen-bonded supramolecular structures that are stable in solution. The oxovanadium(IV) complexes were characterised by elemental analysis, UV/visible, FT-IR and EPR spectroscopy as well as mass spectrometry. Elemental analysis and mass spectrometry confirmed the identity and purity of the complexes. EPR spectroscopy confirmed the monomeric VIVO-bound species. IR and UV/visible spectroscopy confirmed coordination of the ligands to the metal centre. Highresolution solid-state structures were elucidated for the cationic complexes with PHEN coligands and the 4-chloro and 4-tert-butyl derivatives of the [VO(ONO)(DPQ)] complexes. The crystal structures of the complexes showed a monomeric vanadium(IV) species with the bidentate N,N-donor co-ligand and respective dianionic O,N,O′- or monoanionic O,N,N′-tridentate Schiff base ligand coordinated to the oxovanadium centre in a distorted octahedral geometry. The respective neutral VIVO3N3 and monocationic VIVO2N4+ species were formed. A least-squares fit of the solid-state and DFT-simulated (B3LYP-/6-311G(dp)) structures of the oxovanadium chelates indicate the experimental and simulated structures are in good agreement. DFT-simulated structures were determined for those complexes where X-ray data are not available. The geometry-optimised structures for the neutral and cationic complexes all indicate that the respective bidentate polypyridyl ligands are free from steric hindrance by the tridentate ligand and should be available to bind DNA, which is their proposed cellular target. DFT simulations indicated the neutral complexes have larger HOMO-LUMO energy gaps than the corresponding cationic complexes, suggesting that the neutral complexes are more stable with respect to ligand substitution than the cationic complexes. Experimental mass spectrometry and UV/visible spectroscopy confirmed slower solvolysis processes for the neutral complexes versus the cationic analogues. 51V NMR studies indicate partial oxidation of the vanadium(IV) species in DMSO to VVO2(ONO/ONN)(DMSO) analogues. The cationic complexes with PHEN and DPQ co-ligands were deemed suitable to proceed with absorption DNA binding studies. The cytotoxicity screening of the oxovanadium complexes in this work revealed that, in general, the neutral complexes with DPQ co-ligands are cytotoxic against the triple-negative breast cancer MDA-MB and neuroblastoma SH-SY5Y tumour cell lines and non-toxic towards the cervical cancer HeLa cell line. The charge of the complexes was found to influence the cytotoxic properties. The cationic complexes with PHEN and DPQ co-ligands are cytotoxic towards the HeLa cell line as well as the MDA-MB and SH-SY5Y cell lines. The neutral DPQ and cationic complexes with PHEN and DPQ co-ligands were found to be more cytotoxic towards MDA-MB cell lines than cisplatin and the cationic complexes were found to be more cytotoxic towards the HeLa cell line than cisplatin. Steric bulk of the Schiff base functional group influences cytotoxicity with larger functional groups, such as tert-butyl and sulfonyl, leading to lower cytotoxicity. The N,N-donor co-ligand and steric bulk of the Schiff base functional group also influenced the selectivity index of the cationic complexes. The cationic oxovanadium-DPQ complex with a methyl substituent on the tridentate ligand is significantly more toxic to the carcinoma cell lines than the healthy renal cell line HEK293. In comparison, the cationic oxovanadium-PHEN analogue with a methyl substituent and the cationic oxovanadium-DPQ analogue with a bulky tert-butyl substituent are less selective in their cytotoxicity. The DNA binding studies show that the neutral and cationic DPQ compounds do have an affinity for DNA. A positive correlation between antitumour activity and DNA binding affinity was found. The [VO(ONN)(DPQ)](PF6) analogue with a bulky tert-butyl substituent has a lower intrinsic ct-DNA binding constant than the [VO(ONN)(DPQ)](PF6) analogue with a methyl substituent (1.3 × 104 M–1 and 2.8 × 104 M–1 respectively). The cationic DPQ derivatives also bind more strongly to DNA than the cationic complexes with PHEN co-ligands. The steric effect is also evident in the neutral complexes. The [VO(ONO)(DPQ)] complex with a tert-butyl substituent has a lower apparent binding constant than the [VO(ONO)(DPQ)] complex with no substituents on the Schiff base ligand. The cationic charge also led to a higher apparent binding constant for the [VO(ONO)(DPQ)](PF6) complex with a tert-butyl functional group than for the corresponding neutral [VO(ONO)(DPQ)] analogue with a tert-butyl functional group. Absorption and fluorescence spectroscopic and DNA viscosity studies indicate at least a partial DNA intercalative ability for the cationic oxovanadium-DPQ derivatives and the neutral oxovanadium-DPQ complexes with less bulky substituents. Molecular docking studies further highlighted the affinity of the metal chelates towards DNA, including interactions between DNA and the tridentate ligand. The lowest energy molecular docking poses range from ca. -48 to -67 kJ mol–1. Gel electrophoresis studies showed the cationic vanadium complexes with DPQ co-ligands (unlike the neutral DPQ and cationic PHEN analogues) were able to cleave plasmid DNA without adding external oxidising or reducing reagents. Experimental data suggest a singlet oxygen pathway is the most likely. It was also shown that the combination of metal ion and ligand is needed to induce DNA cleavage. The neutral [VO(ONO)(DPQ)] derivative with a methyl functional group on the Schiff base was shown to oxidatively cleave supercoiled plasmid DNA in the presence of H2O2 through the generation of hydroxyl radicals. EPR spintrapping studies with DMPO further support the idea that hydroxyl radicals are formed from reaction of the oxovanadium complex and H2O2. In summary the charge of the complex, type of substituent on the tridentate ligand and the identity of the N,N-donor heterocyclic ligand affected the stability, cytotoxic properties, selectivity, DNA binding, DNA cleavage abilities and DNA cleavage mechanism of the oxovanadium compounds in this study.
Comparison of extraction methods efficiency for the extraction of polycyclic aromatic hydrocarbons and phenolics in water matrices, sludge and sediment: sources of origin and ecological risk assessment.
(2023) Ndwabu, Sinayo.; Mahlambi, Precious Nokwethemba.; Malungana, Mncedisi.
Polycyclic aromatic hydrocarbons (PAHs) and phenolic compounds (PCs) are persistent and environmentally toxic compounds. This study therefore aimed to determine the levels of both PAHs and PCs in river water, wastewater, sludge and sediment samples. The evaluation of their origin source and ecological risk was also determined. The status of both these contaminants in South African environment is still not fully investigated, which is a gap this study intended to fill together with previous studies that have been carried-out. The PAHs and PCs were extracted using different extraction methods which include a solid phase extraction (SPE) and dispersive liquid-liquid micro-extraction (DLLME) in water matrices. The microwave assisted extraction (MAE) and Ultrasonication (UE) coupled with either filtering (F) or F + SPE as a clean-up technique was used for extraction of solid samples. The analytes extracted form water or sediment were determined using GC-MS. The PAH %recoveries obtained under optimum conditions in liquid samples were determined to be 72.1 - 118% for SPE and 70.7 – 88.4% for DLLME while the LOD and LOQ were 5.00 – 18.0 ng/L and 10.0 – 44.0 ng/L for SPE while they were 6.00 – 20.0 ng/L and 11.0 – 63.0 ng/L for DLLME. The recovery test for PAHs in solid samples gave a range of 93.7% - 121% for UE and 79.6% - 122% for MAE while the LOD and LOQ ranged from 0.0250 μg/kg to 1.21 μg/kg & 0.0800 μg/kg to 3.54 μg/kg for MAE and from 0.0840 μg/kg to 0.215 μg/kg & 0.0190 μg/kg to 0.642 μg/kg for UE respectively. The LOD and LOQs for PCs in both water and solid matrices were 0.01 – 2.00 μg/L and 0.02 – 6.07 μg/L for SPE, 0.05 – 1.20 μg/kg and 0.17 – 3.17 μg/kg for MAE and 0.09 – 1.33 μg/kg and 0.26 - 3.54 μg/kg for UE correspondingly, their %recovery test gave ranges of 75.2 – 112% (SPE), 80.9 – 110% (MAE) and 79.3 – 119% (UE).The optimization and validation of these methods indicated that they can be used for the extraction of PAHs or PCs in liquid samples, however, SPE when compared to DLLME showed to be more accurate and sensitive. Moreover, in solid samples the clean-up method was a deciding factor, with F + SPE cleaned samples giving higher concentrations of both PCs and PAHs than the filtered ones in both MAE and UE. The concentrations of PAHs ranged from nd (not detected) to 1046 ng/L in river water and nd to 778 ng/L in wastewater samples with naphthalene showing dominance over all other PAHs in both water matrices. The PC concentrations at 4.12 to 1134 μg/L for wastewater and nd to 98.0 μg/L for river water were high but still within the maximum allowable limit except for 2.4-DCP (2.4 dichlorophenol) at Wdv4. The concentrations obtained from F + SPE cleaned samples were higher for both PAHs and PCs with a range from 95.96 to 926.0 μg/kg and 1.30 to 310 μg/kg compared to concentrations from filtered only samples at 21.61 to 380.6 μg/kg and 0.90 to 266 μg/kg respectively. Pyrene showed dominance over all other PAHs in both sludge and sediments while 2.4-DCP and PCP dominated the sludge and sediment samples respectively. PAHs were determined to be of petrogenic (water matrices) and pyrolytic (solid samples) origin and on average posed low (water matrices) and a medium to high (solid matrices) ecological risk. The ILCRderm values at 4.98 x 10-1 and 2.62 x 10-1 (DahA) and 5.92 x 10-2 and 5.34 x 10-2 (PCP) were highfor adults compared to that of children at 1.92 x 10-1 and 1.01 x 10-1 (DahA) and 1.39 x 10-2 and 1.26 x 10-2 (PCP) for both sediment and sludge samples respectively. The low values of ILCRderm for children indicates that the have a high risk exposure even at low concentrations of the contaminants. The findings of this study showed that both areas (uMsunduzi river and Darvill wastewater works (WWW) of interest are polluted with PAHs and PCs therefore, more regulations such as the National Environmental Management: Waste Act (NEMWA) are needed to ensure environmental, human and animal safety.
Targeting the tumour extracellular environment through rational modification of the SNX class of HSP90 inhibitors.
(2023) Mathenjwa, Gciniwe Sindiswa.; Akerman, Matthew Piers.; Veale, Clinton Garenth Lancaster.; Bode, Moira Leanne.
HSP90 remains a valuable target for cancer therapy. Unfortunately, targeting intracellular HSP90 has proven not to be a viable chemotherapeutic approach. Compensatory HSR induction and HSP70 overexpression are the main limitations of this approach. A growing body of evidence suggests that targeting the extracellular environment would be of advantage and devoid of the drawbacks observed with intracellular HSP90 inhibition. As a result, the development of extracellular HSP90 inhibitors represents a novel opportunity for cancer therapeutics. In view of this hypothesis, we aimed to design and synthesise extracellular inhibitors and to assay these compounds against HSP90. To develop extracellular HSP90 inhibitors, cell-impermeable analogues of the well-developed benzamide HSP90 inhibitor (SNX 2112) were designed, synthesised and biologically evaluated. The desired target compounds were synthesized using developed methodology, as well as modified methodology. In Chapter 3 we compared and evaluated a variety of reported synthetic methods to deliver the analogues of SNX 2112. Interested in a general procedure for the synthesis of our analogues, we initially attempted to afford both the methyl and the trifluoromethyl containing analogues via a β- triketone mediated procedure. Despite the success observed with the methyl analogues, the instability of a trifluoromethyl containing β-triketone, deemed this procedure not feasible for this class of compounds. Our continued effort towards a general procedure led to the investigation of a tosylhydrazone mediated tetrahydroindazolone condensation; unexpectedly attempts to synthesise the methyl containing analogues via this procedure led to a 1—5 nitrogen to carbon tosyl migration, which was further investigated for varying substrates, and these results are explained in detail in this thesis. It then became apparent that each of the reported methods had its merits and shortcomings, there was no one best method, rather the synthetic approach was mainly determined by the C-3 substituent. The key intermediates were then converted into the desired targeted compounds by tethering the HSP90 pharmacophore to flexible alkyl groups, attached to polar sulfonate and phosphonate functionalities. Hypothetically, introduction of polar alkyl groups, would inhibit cell penetration thus limiting them to the extracellular environment. Based on the goals of our study we were interested in three biological evaluations; to confirm that our modified compounds were still capable of inhibiting HSP90s ATPase activity, to evaluate if our modifications reduced intracellular HSP90 activity, whether they stimulated the pro-oncogenic HSR, and to evaluate their cytotoxicity. Preliminary biological assessment of our compounds was consistent with our hypothesis. Here we showed that our compounds did not inhibit intracellular HSP90, and did not stimulate HSP70 expression, a marker of induction of the compensatory HSR. Furthermore, our analogues displayed cytotoxicity in the nanomolar range against the HeLa cell line. These preliminary data support the feasibility of targeting extracellular HSP90 as a novel anticancer strategy.
Syntheses of mixed donor homogeneous and immobilized palladium(II) complexes catalysts for methoxycarbonylation and hydrogenation reactions.
(2021) Akiri, Saphan Owino.; Ojwach, Stephen Otieno.
Reactions of ligands (E)-N'-(2,6-diisopropylphenyl)-N-(4-methylpyridin-2-yl)benzimidamide (L1), (E)-N'-(2,6-diisopropylphenyl)-N-(6-methylpyridin-2-yl)benzimidamide (L2), (E)-N'-(2,6-dimethylphenyl)-N-(6-methylpyridin-2-yl)benzimidamide (L3), (E)-N'-(2,6-dimethylphenyl)-N-(4-methylpyridin-2-yl)benzimidamide (L4) and (E)-N-(6-methylpyridin-2-yl)-N'-phenylbenzimidamide (L5) with [Pd(NCMe)2Cl2] furnished the corresponding palladium(II) pre-catalysts (Pd1-Pd5), in good yields. Molecular structures of Pd2 and Pd3 revealed an N^N bidentate coordination mode to afford square planar compounds. Activation of the palladium(II) complexes with para tolyl sulfonic acid (PTSA) afforded active catalysts in the alkenes methoxycarbonylation. The resultant catalytic activities were controlled by both the complex structure and alkene substrate. While aliphatic substrates favoured the formation of linear esters (>70%), styrene substrate resulted in predominantly branched esters of up to 91%. The water-soluble ligands; sodium 4-hydroxy-3-((phenylimino)methyl)benzenesulfonate (L6), sodium 3-(((2,6-dimethylphenyl)imino)methyl)-4-hydroxybenzenesulfonate (L7) and sodium 3-(2,6-diisopropylphenyl)imino)methyl)-4-hydroxybenzenesulfonate (L8) reacted with with Pd(OAc)2 afford their respective palladium(II) complexes [Pd(6)2] (Pd6), [Pd(L7)2] (Pd7) and [Pd(L8)2] (PdL8). In addition, treatment of the non-water-soluble ligands 2-((phenylimino)methyl)phenol (L9), 2-(((2,6-dimethylphenyl)imino)methyl)phenol (L10) and 2-((2,6 diisopropylphenyl)imino)methyl)phenol (L11) with Pd(OAc)2 yielded complexes [Pd(L9)2] (Pd9), [Pd(10)2] (Pd10) and [Pd(L11)2] (Pd11), respectively in good yields. Solid-state structures of compounds Pd6 and Pd9 revealed bis(chelated) square planar neutral compounds. All the complexes formed active catalysts in the methoxycarbonylation of 1- hexene, affording yields of up to 92% within 20 h and regioselectivity of 73% in favour of linear esters. The activities and selectivities of the compounds depended on the steric encumbrance around the coordination centre. The water-soluble complexes displayed comparable catalytic behaviour to the non-water-soluble systems. The complexes could be recycled five times with minimal changes in both the catalytic activities and regio-selectivity. Reactions of (amino)phenyl ligands, (E)-N-((Z)-4-(phenylamino)pent-3-en-2-ylidene)aniline (L12) and N,N'E,N,N'E)-N,N'-(3-(3 (triethoxysilyl)propyl)pentane-2,4-diylidene)dianiline (L13) with [Pd(NCMe)2Cl2] led to the formation of homogeneous complexes Pd13 and Pd14. Besides, supporting of complex Pd14 with either MCM-41, SBA-15, or Fe3O4 magnetic nanoparticles gave immobilized complexes P15-Pd17, respectively. Using varying metal loading in the MCM-41 immobilization of complex Pd14 produced complexes Pd18 and Pd 19. In addition, calcination of complex Pd16 at 150oC and 200oC led to the formation of complexes Pd20 and Pd21, respectively. All the complexes were received in good yields. The catalytic activities and selectivities of the homogeneous complexes were influenced by the coordination sphere, with the complexes predominantly forming linear esters. On the other hand, the catalytic behaviours of the immobilized catalysts depended on the nature of support and calcination temperatures. In addition, the catalytic activities were observed to depend on the reaction temperature, catalyst loading, amounts of PPh3 and acid promoters. The immobilized complexes Pd15, Pd16 and Pd17, were recycled up to five times. The homogeneous and silica immobilized palladium(II) complexes of ligands (2-phenyl-2-((3(triethoxysilyl)propyl)imino)ethanol) (L14), (4-methyl-2-((3(triethoxysilyl)propyl)imino)methyl)phenol ) (L15 ), [L14-MCM-41 (L16), and [L15- MCM-41 (L17)]. The homogeneous complexes [Pd(L14)2] (Pd22), [Pd(L14)2] (Pd23), [Pd(L14)(Cl2)] (Pd24), [Pd(L15)(Cl2)] (Pd25) were obtained from homogenous ligands L14, L15, L16 and L17 respectively. In addition, the silica immobilized compounds [Pd(L14)2]-MCM-41] (Pd26) and [Pd(L15)2)-MCM-4] (Pd27) were obtained through convergence immobilization of complexes Pd22 and Pd23, respectively. Comparatively, immobilized complexes [Pd (L14)(Cl2)-MCM-41] (Pd28) and [Pd(L15)(Cl2)]-MCM-41] (Pd29) were obtained from the complexation of immobilized ligands L16 and L17. Both sets of complexes gave active catalysts in molecular hydrogenation of alkenes, alkynes and functionalized benzenes. The catalytic activities and product distribution in these reactions were largely dictated by the nature of the substrate. The kinetic studies revealed reaction orders dependence on styrene for both the homogeneous and supported catalysts. Significantly, the selectivity of both sets of catalysts was comparable in the hydrogenation of alkynes and multi-functionalized benzenes. The supported catalysts could be recycled up to four times with minimum reduction in catalytic activities and showed the absence of any leaching from hot filtration experiments. Kinetics and poisoning studies established the presence of active homogeneous species for complexes Pd22-Pd5 and Pd(0) nanoparticles for the immobilized complexes Pd26-Pd29, respectively.
Carboxamide ruthenium(II) and manganese(II) complexes: structural, kinetic, and mechanistic studies in the transfer hydrogenation of ketones.
(2022) Kumah, Robert Tettey.; Ojwach, Stephen Otieno.
The carboxamide ligands N-(benzo[d]thiazol-2-yl)pyrazine-2-carboxamide (HL1), N-(1H-benzo[d]imidazol-2-yl)pyrazine-2-carboxamide (HL2), were prepared by condensation of pyrazine-carboxylic acid and appropriate heteroaromatic amines. Reactions of HL1 and HL2 with ruthenium(II) precursors, [RuH(CO)Cl(PPh3)3] and [RuH2(CO)(PPh3)3] afforded the mononuclear complexes [RuL1(PPh3)2(CO)Cl] (Ru1), [RuL1(PPh3)2(CO)H] (Ru2), [RuL2(PPh3)2(CO)Cl] (Ru3), [RuL2(PPh3)2(CO)H] (Ru4). The solid-state structures of complexes Ru1, Ru2, and Ru4 reveal bidentate modes of coordination of the ligands and distorted octahedral geometries around the Ru(II) centre. The complexes formed active catalysts in the transfer hydrogenation of ketones and achieved turnover number (TON) up to 530 in 6 h. The ruthenium(II)–hydride complexes, Ru2 and Ru4, were capable of catalysing transfer hydrogenation of ketones reactions under base free reaction conditions and demonstrated higher catalytic activities compared to the corresponding non-hydride analogues (Ru1 and Ru3). An inner sphere monohydride mechanism involving dissociation of one PPh3 group was proposed from in situ 31P{1H} NMR spectroscopy studies. Dipicolinamide ligand system, N,N'-(1,4 phenylene)dipicolinamide (H2L3), N,N'-(1,2-phenylene)dipicolinamide (H2L4), N,N'-(4,5-dimethyl-1,2-phenylene)dipicolinamide (H2L5), N,N'-(4-methoxy-1,2-phenylene)dipicolinamide (H2L6) were synthesised following a similar protocol described for HL1 and HL2. Treatment of the ligands H2L3 and H2L4 with RuH(CO)Cl(PPh3)3 afforded bimetallic complexes [Ru2(H2L3)(PPh3)4(CO)2][2Cl] (Ru5), [Ru2(H2L3)(H)2(PPh3)4(CO)2] (Ru5b), [Ru2(HL4)(PPh3)3(CO)2Cl3] (Ru6) and a mononuclear complex [RuCl2L4(PPh3)2(CO)] (Ru7). The solid-sate structure of the dinuclear ruthenium(II) complexes confirmed a bidentate coordinate mode, with PPh3, CO, and chlorido auxiliary ligands occupying the remaining coordinating sites to afford distorted trigonal bipyramidal geometries (Ru5 and Ru6) while the mononuclear complex Ru7 adopted a distorted octahedral geometry around its ruthenium(II) atom. The reaction of the ligands H2L4-H2L6 with the [RuCl2-η6-p-cymene]2 precursor produces half-sandwich diruthenium complexes [{Ru(η6-p-cymene)}2-μ-Cl(L4)][Ru(η6-p-cymene)Cl3] (Ru8), [{Ru(η6-p-cymene)}2-μ-Cl(L4)][PF6] (Ru9), [{Ru(η6-p-cymene)}2-μ-Cl(L5)][PF6] (Ru10), and [{Ru(η6-p-cymene)}2-μ-Cl (L6)][PF6] (Ru11). The molecular structure of cationic complexes, Ru8-Ru11, was confirmed by single-crystal X-ray crystallography analysis. The complexes Ru8-Ru11 display a bidentate Npyridine ^ Namidate mode of coordination to give pseudo-octahedral geometry (piano-stool-like geometry). The ruthenium(II) complexes demonstrated remarkable enhanced catalytic activity (TON values up to 1.71 x 104) in the transfer hydrogenation of ketones at a very low catalyst loading of 2.75 x10-2 mol% (275 ppm). The dinuclear ruthenium(II) complexes showed higher catalytic activity compared to the corresponding mononuclear complex Ru5. The half-sandwich diruthenium complexes Ru8-Ru11 displayed relatively higher catalytic activity than the ruthenium complexes Ru5 and Ru6 bearing the PPh3 co-ligands. Monohydride inner-sphere catalytic cycle was proposed for the transfer hydrogenation of ketones catalysed by both Ru1 and Ru9, and the formation of the reactive intermediates was supported with low-resolution mass spectrometry data. The dinuclear ruthenium complexes of pyridine and pyrazine-carboxamide bearing quinolinyl motif were synthesised by reacting, N-(quinolin-8-yl)pyrazine-2-carboxamide, (HL7), 5-methyl-N-(quinolin-8-yl)pyridine-2-carboxamide, (HL8), 5-chloro-N-(quinolin-8-yl)pyridine-2-carboxamide, (HL9), and 2-pyrazine-carboxylic acid (HL10) with equimolar [RuCl2(η6-p-cymene)]2 to afford the dinuclear complexes [{Ru(η6-p-cymene)}2Cl3(L10)] (Ru12), [{Ru(η6-p-cymene)Cl}2(L7)] [PF6] (Ru13), [{Ru(η6-p-cymene)Cl}2(L8)][Ru(L8)Cl3] (Ru14), and [{Ru(η6-p-cymene)Cl}2(L9)][PF6] (Ru15), respectively. The solid-state structures of the dinuclear complexes Ru12 and Ru13 reveal a typical piano-stool geometry around the Ru(II) ions. The dinuclear ruthenium complexes Ru12-Ru15 were used as catalysts in the transfer hydrogenation of a broad spectrum of aldehydes and ketones and demonstrated excellent catalytic activity, TON values up to 4.8 x 104, using catalyst loading of 2.0 x10-3 mol% (20 ppm). The catalytic performance of the complexes was affected by the ligand architecture and the substituents on the pyridyl ring. Complexes Ru13-15 exhibited higher catalytic activities compared to the complex Ru12 which could be ascribed to the role of quinoline in stabilising the complexes. The pyridine and pyrazine motifs have a significant impact on the reactivity and the catalytic activity of the complexes. In-situ low-resolution ESI-MS analyses of the reactive intermediates aided in proposing a monohydride inner-sphere mechanism for the transfer hydrogenation of ketones catalysed by Ru15. To develop a more sustainable, environmentally compatible and cost-efficient protocol for transfer hydrogenation of ketones, a new catalytic system based on manganese(II) metal was synthesised. New manganese(II) complexes Mn1-Mn4, ligated on dipicolinamide ligands were synthesized by treating the N,N'-(1,4-phenylene)dipicolinamide (H2L3), N,N'-(1,2-phenylene)dipicolinamide (H2L4), N,N'-(4-methoxy-1,2-phenylene)dipicolinamide (H2L5) and N,N'-(4,5-dimethyl-1,2-phenylene)dipicolinamide (H2L6) with MnCl2.4H2O salt to afford dinuclear manganese(II) complexes [Mn2(H2L3)2Cl4] (Mn1), [Mn2(H2L4)2Cl4] (Mn2), [Mn2(H2L5)2(Cl)4] (Mn3) and [Mn2(H2L6)2Cl4] (Mn4). The solid-state structure of complex Mn2 showed a six-coordinate dinuclear complex with the two Mn(II) ions adopting a distorted octahedral environment surrounded by two tetradentate ligands and chlorido co-ligands, respectively. The Mn(II) complexes formed active catalysts in transfer hydrogenation of ketones to achieve TON values up to 5.12 x 104. The presence of electron-donating substituents -OCH3 and -CH3 in complexes Mn3 and Mn4 displayed minor effects in the transfer hydrogenation of ketones. The new carboxamide-manganese(II) complexes are among the most active manganese-based catalysts capable of hydrogenating a large scope of ketones ranging from aliphatic to aromatic ketones. A dihydride catalytic cycle has been proposed and supported with in-situ low-resolution mass spectrometry data.
Assessment of antiretroviral drugs uptake by vegetables from contaminated soil and their adsorption by exfoliated graphite in river and wastewater.
(2022) Kunene, Philisiwe Nganaki.; Mahlambi, Precious Nokwethemba.
This study was directed toward vegetable uptake of the commonly used antiretroviral drugs (ARVDs), abacavir, nevirapine, and efavirenz. Antiretroviral drugs are used to treat the human immune-deficiency virus (HIV). South Africa (SA) is one of the countries with a high number of infected people on ARV therapy, therefore, the ARVDs are anticipated to be existing at high concentrations in the South African environment than in other countries worldwide. In recent years, the presence of ARVDs in the environment has drawn attention; hence studies have reported their presence in aquatic environments while very few studies have been conducted on their uptake using vegetables. This work was therefore based on the optimization and application of sensitive, simple, cost-effective, and robust techniques for quantifying ARVDs in vegetables. Based on this information, ultrasonic extraction (UE) and microwave-assisted extraction (MAE) were used to isolate target compounds from vegetable samples to the aqueous phase. Dispersive liquid-liquid microextraction (DLLME) and solid-phase extraction (SPE) were utilized to preconcentration and clean up the extracts from UE and MAE, respectively. A liquid chromatography photodiode array detector (LC-PDA) was utilized to detect and quantify the extracted compounds. The UE with and without DLLME cleanup were compared with each other, also, MAE with and without SPE cleanup were compared with each other. The methods comparison was done in terms of their detection (LOD) and quantification limits (LOQ), extraction efficiencies (%Recovery), relative standard deviations (%RSD), and concentrations of ARVDs found in vegetable samples. In comparison of UE and ultrasonic-assisted dispersive liquid-liquid microextraction (UADLLME), the LOD and LOQ obtained ranged between 0.0081 - 0.015 μg/kg and 0.027 - 0.049 μg/kg for UE and 0.0028 -0.0051 μg/kg and 0.0094 - 0.017 μg/kg for UADLLME respectively. High recoveries ranging from 93 to 113% in UE and 85 to 103% in UADLLME with less than 10% RSD in both procedures were obtained. These results indicated that UADLLME is more sensitive than the UE method, although they are both accurate and precise. The UE can be recommended for routine analysis as UADLLME showed the inability to extract analytes from root vegetables. The optimized UE and UADLLME methods were applied to extract ARVDs from vegetables bought from local fruit and veggie supermarket. Vegetables were categorized as root (carrot, potato, and sweet potatoes), leaf (cabbage and lettuce), and fruit (green paper, butternut, and tomato). The target ARVDs were quantified in most samples with concentrations up to 8.18 μg/kg. The concentrations obtained were slightly high in UADLLME than in UE as a result of its high sensitivity. Efavirenz was the most dominant drug, while the potato was the most contaminated vegetable. In the comparison of MAE and MAESPE, the obtained LOD and LOQ ranged from 0.020 to 0.032 μg/kg and 0.068 to 0.109 μg/kg for MAE and 0.019 to 0.066 μg/L and 0.065 to 0.22 μg/L for MAE-SPE. The obtained recoveries ranged from 85 to 103% for MAE and from 82 to 98 % for MAE-SPE, respectively, and the RSDs were all less than 6%. These results showed that both methods have comparable sensitivity; however, the recoveries values for MAE were slightly higher than those obtained in MAE-SPE, which signals MAE’s high accuracy. The optimized MAE and MAE-SPE methods were applied to remove ARVDs in the root (potatoes, onions, and beetroot), leaf (lettuce, and spinach), and fruit (green paper, cucumber, and eggplant) vegetables bought from local fruit and veggie supermarket. The obtained ARVDs concentration range was 1.48 ± 0.5 - 27.9 ± 1.2 μg/kg. The MAE-SPE resulted in low concentration compared to MAE without cleanup. Beetroot exhibited high concentrations of the target ARVDs, while nevirapine was found to have high concentration and as a dominant compound. The results obtained revealed that the vegetables from the studied area are contaminted with ARVDs, which could indicate their possible irrigation with wastewater effluent or the use of sludge as biosolids in the agricultural areas. This is a concern as it leads to unintentional consumption by consumers which could lead to drug resistance by the human body or have human health effects. The study was then expanded by conducting the phytoremediation approach to investigate the uptake of abacavir, nevirapine, and efavirenz by beetroot, spinach, and tomato from the contaminated soil. The three selected vegetable plants were planted and irrigated with ARVDs spiked (at 2000 and 5000 μg/L) water over a period of three months. The optimized UE and LC-PDA methods were used to extract and quantify the selected ARVDs from the target vegetables and soil. The obtained results showed that the studied vegetables have the potential to take up abacavir, nevirapine, and efavirenz from contaminated soil, be absorbed by the root, and translocate to the aerial part of the plants. Abacavir was found at high concentrations to a maximum of 40.21 μg/kg in the root, 18.43 μg/kg in the stem, and 6.77 μg/kg in the soil, while efavirenz was the highest concentrations, up to 35.44 μg/kg in leaves and 8.86 μg/kg in fruits. Spinach root accumulated more ARVDs than beetroot and tomato. The bio-accumulation factor ranged from 2.0-14 μg/kg in beetroot, 3.6 - 15 μg/kg in spinach, and 6 – 10 μg/kg in tomato. The root concentration factor range was 0.047 – 17.6 μg/kg; 0.34-5.9 μg/kg, and 0.14-2.82 μg/kg in beetroot, spinach, and tomato, respectively. The translocation factor range obtained was 0.40 – 38 μg/kg, 0.08 – 19 μg/kg, and 0.14 – 49 μg/kg in beetroot, spinach, and tomato, respectively. However, the accumulation of ARVDs in all studied plants showed that they could be used in phytoremediation. The results obtained in the phytoremediation approach revealed that the utilization of the contaminated water has an influence on the presence ARVDs in vegetables; hence this work also focused on evaluating the exfoliated graphite adsorption of ARVDs in water. Natural graphite was intercalated with acids and exfoliated with thermal shock to obtain the exfoliated graphite. The scanning electron microscopy images showed that the exfoliated graphite had increased c-axis distance between the layers with accordion-like structure which were confirmed by the lower density of exfoliated graphite material (0.0068 gmL-1) compared to the natural graphite (0.54 g mL-1). Fourier Transformed Infrared Spectroscopy results showed the C=C in natural and exfoliated graphite at 1635 cm-1 stretching. The phenolic, alcoholic, and carboxylic groups were observed from 1000 to 1700 cm-1 for the intercalated and exfoliated graphite. The Energy-dispersive X-ray results further confirmed these results, which showed carbon and oxygen peaks in the intercalated and exfoliated graphite spectrum, whereas natural graphite showed only a carbon peak. Raman spectroscopy results showed that the material’s crystallinity was not affected by the intercalation and exfoliation processes as observed from the ratios of the G and D peaks and the G' and D'. Natural, intercalated and exfoliated graphite contained the D, G, D', and G' peaks at about 1350 cm-1, 1570 cm-1, 2440 cm-1, and 2720 cm-1, respectively. The exfoliated graphite material showed the characteristic of a hexagonal phase graphitic structure by (002) and (110) reflections in the X-ray diffraction results. The exfoliated graphite adsorption method was optimized based on the pH of a solution, adsorbent dosage, and adsorption time prior to application to water samples. The optimum pH solution, adsorbent dosage, and adsorption time were 7, 30 mg, 0.01 μg/L, and 30 minutes respectively. The kinetics and isotherm studies were conducted to assess the model that best fit and explain the experimental data obtained. The kinetic model and adsorption isotherm studies showed that the experimental data fit well pseudo-second-order kinetics and is well explained by Freundlich’s adsorption isotherm. The maximum adsorption capacity of the exfoliated graphite (EG) for ARVDs ranges between 1.660-197.0, 1.660-232.5, and 1.650-237.7 mg/g for abacavir, nevirapine, and efavirenz, respectively. These results showed that under proper operating conditions, the EG adsorbent could potentially be applied as a water purifying tool for the removal of ARVDs pollutants.
Phytochemistry and quorum sensing inhibitory studies of four vernonia species growing in Nigeria.
(2017) Aliyu, Abubakar Babando; Moodley, Brenda.
Abstract This thesis contains the phytochemical analysis of four medicinal Vernonia species growing in Nigeria and used in ethnomedicine to treat a variety of medical conditions. The four Vernonia species studied were Vernonia blumeoides, Vernonia perrottetii, Vernonia ambigua and Vernonia glaberrima. The thesis also contains a comprehensive review of the sesquiterpene lactones from the genus Vernonia, their structural diversity and biosynthetic considerations. For each of the plants, the crude extracts and selected compounds were tested for their antibacterial activity using the traditional disc diffusion and broth microdilution as well as anti-quorum sensing assays. Three of the four plants studied yielded sesquiterpene lactones, Vernonia blumeoides yielded four novel eudesmanolide sesquiterpene lactones (blumeoidolides A-D), Vernonia perrottettii yielded a novel keto-hirsutinolide 13-acetoxy-1(4β),5(6)β-diepoxy-8α-(senecioyloxy)-3-oxo-1,7(11)-germacradiene-12,6-olide B1 and the known keto-hirsutinolide 13-acetoxy-1,4β-epoxy-8α-(senecioyloxy)-3-oxo-1,5,7(11)-germacratriene-12,6-olide B2 and Vernonia ambigua yielded a novel glaucolide sesquiterpene, 5,6-dehydrobrachycalyxolide. The structures of the sesquiterpenes were determined from their 1H, 13C and 2D NMR spectra along with mass spectra. The crystal structure of one of the eudesmanolide sesquiterpenes allowed the configuration of the stereocentres in the molecule to be determined. In addition to the sesquiterpene lactones, some common sterols and flavonoids were isolated from the plants: stigmasterol was isolated from V. blumeoides, lupeol was isolated from V. blumeoides, V. ambigua and V. perrottettii and lupeol acetate from V. ambigua and V. perrottetti. The flavonoid apigenin was found in V. blumeoides, V. perrottetti and V. glaberrima, luteolin in V. blumeoides and V. perrottetti, velutin in V. perrottetti and V. glaberrima and chrysoeriol in V. ambigua and V. glaberrima. Chrysin was found only in V. blumeoides and luteolin 3',4'-dimethyl ether in V. glaberrima. Several of the isolated sesquiterpene lactones showed good anti-quorum sensing inhibition (QSI). QSI ≥80% was obtained for blumeoidolide A at a concentration ≥ 0.071 mg mL-1, blumeoidolide B (≥ 3.6 mg mL-1) and B1 (1.31 mg mL-1), QSI ≥75% for B2 (0.33 mg mL-1) and QSI ≥84% for 5,6-dehydrobrachycalyxolide (2.6 mg mL-1). The sterols, lupeol and lupeol acetate, were also found to have QSI ≥84% at 2.6 mg mL-1. Molecular docking studies carried out on blumeodolides A-D in the binding sites of CviR and CviR' (transcription activator proteins) suggested that these molecules are able to bind to certain domains in the target protein, thus eliciting an effect. The current work adds to the library of sesquiterpene lactones from the genus Vernonia and provides some lead compounds to antibacterial activity via quorum sensing inhibition.
NCC-based SERS substrate: fundamentals, preparation and applications.
(2018) Ogundare, Segun Ajibola.; Van Zyl, Werner Ewald.
This study reports the isolation and characterization of nanocrystalline cellulose (NCC) produced from discarded cigarette filters (DCF). The DCF were processed into cellulose via ethanolic extraction, hypochlorite bleaching, alkaline deacetylation, and then converted into NCC by sulphuric acid hydrolysis. The morphological structures of the isolated NCC, established with TEM, showed that the nanocrystals were needle-like with length in the range of between 62–258 nm. FEGSEM showed the morphological transition of the micro-sized DCF to a self-assembled NCC, while EDX revealed the presence of Ti (as TiO2) in DCF, which was retained in the NCC. A NCC sample that was freeze-dried showed a specific surface area of ~8 m2/g. The crystallinity of the NCC film and the freeze-dried samples were ~97% and ~94%, respectively. Crystallite sizes of the freeze-dried (8.4 nm) and film (7.6 nm) samples correlated with the mean width (8.3 nm) of the NCC, observed with the TEM. The isolated NCC was used in the dual role as a reducing- and stabilizing agent in the formation of silver nanoparticles (AgNPs). By this method, a notable size variation of the synthesized AgNPs was found over the pH range of between 5–10, ranging from 4.61 nm at pH = 9 and increasing to 19.93 nm diameter at pH = 5. The size and yield of the AgNPs were also affected by the reaction time and concentration. The spherically shaped AgNPs induced a localized surface plasmon resonance (LSPR) at around 416 nm. The Ag content in the dry AgNPs was 81.9 wt%, which correlated with 82.1 wt% mass left at 600 ºC. Further analysis showed that the dry AgNPs were macroporous with reduced surface area and porosity upon calcination. The sensitivity of the AgNPs showed excellent surface-enhanced Raman scattering (SERS) of riboflavin. The limit-of-detection (LOD) for riboflavin, based on a signal-to-background ratio of 3:1, was found to be 3 x 10-7 M. The intensities of SERS signals increased with increase in concentration. In addition, clusters of AgNPs were synthesized with NCC isolated from Whatman cellulose filter paper, which behaved as a dual reducing- and stabilizing agent, and Stӧber silica (SiO2) provided a suitable anchoring surface. The synthesized nanocomposite (AgNPs/SiO2/NCC) was evaluated as a substrate for surface-enhanced Raman scattering (SERS) of malachite green (MG) and compared with AgNPs/NCC nanocomposite. The FTIR spectra of both nanocomposites showed a weak carbonyl band (1754 cm-1), indicating partial oxidation of the NCC. The UV-vis spectrum of AgNPs/NCC showed a narrow peak at 412 nm, characteristicof LSPR of monodispersed AgNPs. However, this peak was broad with a shoulder at 490 nm in the spectrum of the AgNPs/SiO2/NCC, which indicated clustering of the plasmonic nanoparticles. TEM micrographs showed that the plasmonic nanoparticles were monodispersed with a mean diameter of 19.5 nm in AgNPs/NCC, while they aggregated into clusters on SiO2 in AgNPs/SiO2/NCC resulting in an approximately 20 nm increase in the mean diameter of the SiO2. The SEM/EDX spectra and XRD diffractograms of the nanocomposites showed Ag as the predominant element. The SERS performance of the nanocomposites was evaluated by using MG as a probe, showed AgNPs/SiO2/NCC as a superior substrate with significant improvement in intensities of Raman peaks of MG and high sensitivity as the LOD was 0.9 nm, while AgNPs/NCC showed a LOD of 5.2 nm, based on a signal-to-background ratio of 3:1. This result underscores the huge contribution of SERS “hot spots” as AgNPs assembled into clusters in contrast to monodispersed AgNPs in the absence of SiO2.
Production of nanocellulose composites and catalytic and microbial applications.
(2018) Moodley, Vashen.; Van Zyl, Werner Ewald.
This study describes the preparation, isolation, characterization and application of polysaccharide based nanocrystalline cellulose (NCC) from two source materials (filter paper and bleached pulp). The isolated NCCs were utilized as a composite/support material. Hydroxyapatite (HAp), titanium(IV) oxide (anatase phase, TiO2) and biologically synthesized silver nanoparticles were chosen as the preferred candidates for the incorporation of NCC into their respective matrices, which allowed for the preparation of three new materials. These newly prepared composites were applied in catalytic and anti-microbial studies. NCC formed the basis of this investigation and was prepared via a common acid hydrolysis treatment, using sulfuric acid as the preferred acid hydrolytic medium. The isolated NCCs were obtained in reasonable yields and were characterized using the following techniques: ATR-FTIR, XRD, TEM, HRTEM, FEGSEM equipped with EDX detector, TGA/DTA, NTA (zeta potential) and BET. The rod-shaped particles revealed a high crystallinity, small crystallite sizes and good thermal stabilities. These results led to use the prepared NCC’s as a composite material in the pursuit to prepare a new class of materials with a potential array of applications. The preparation of HAp and subsequently the NCC/HAp (in varied wt%) composite allowed for the use of this newly synthesized material as a versatile catalyst. The catalyst was well characterized and used in the preparation of the two-component one-pot synthesis of triazolidine-3-one derivatives. Preliminary reaction optimization established that the 40 w/w% NCC/HAp composite catalyst returned the best results. The eleven new triazolidine-3-one derivatives (4a - 4k) were synthesized in good yields and maintained good atom economy. The catalyst proved to be an effective tool in this protocol with the supplementary advantage of being recyclable. This approach to organic multicomponent reactions (MCRs) proved to be a cost-effective strategy and allowed for an easy work up with environment-friendly reaction conditions. Compared to non-catalytic protocols this approach required shorter reaction times. With the prospect of employing the optical properties of the prepared NCC, TiO2 was then considered for the use in the second newly prepared composite material as a potential photo-catalyst. The NCC/TiO2 material was synthesized in varying wt% and was characterized via a number of optical, spectroscopic and microscopic techniques to establish if it possessed the potential to be used as a photo-catalyst. Proceeding this evaluation, the NCC/TiO2 material was used in the solar-driven photo-degradation mineralization of o-chloranil (2,3,5,6-tetrachloro-2,5-cyclohexadiene-1,4-dione), a commonly used pesticide. The successful decomposition of o-chloranil led to the identifiable products to 2,3-dichloro-4,5-dioxohex-2-enedoic acid (DCA), 2,3-dioxosuccinic acid (DSA) and oxalic acid (OA). This proved that the 20 w/w% NCC/TiO2 composite could be employed as a successful photo-catalyst, and in particular that NCC could be used as a successful composite material together with TiO2. The rate of degradation was influenced by various parameters such as substrate concentration and photo-catalyst loading. The intermediate product (DCA) formed during the decomposition process was assumed to slow down the progression of the reaction and provided a useful insight into the degradation pathway of the contaminant. The final study demonstrated the synthesis of silver nanoparticles (AgNPs) via a biological (phyto-mediated) route using Lippia javanica plant extract (LPE). The preparation of the colloidal AgNPs involved a variation in the LPE (100 – 400 μL) and AgNO3 (1 – 10 mM) concentration to determine the ideal morphology of AgNPs formed. This biosynthetic approach proved successful in the formation of AgNPs in colloidal form, with superior advantages over the chemical formation. These AgNPs were used in several applications as demonstrated with the inclusion of NCC as a support. Samples were characterized via optical, spectroscopic and microscopic analyses, with the ideal colloidal solution C9 (400 μL LPE, 10 mM AgNO3) established as providing the greatest number of AgNPs with the lowest size. This colloid was chosen for the further incorporation of NCC. NCC (filter paper source) was then incorporated into the quasi-spherical shaped nanoparticle matrix and further characterized, analysed and applied as a catalyst to the synthesis of benzylidene-bis-(4-hydroxycoumarin) derivatives and as a potential bactericidal agent. All colloidal samples were biologically tested against 5 bacterial strains and demonstrated good activity, however, samples with the highest concentration of AgNPs were chosen for further Minimal Bactericidal Concentration (MBC) testing. The results showed that all samples were superior in relation to their anti-biotic counterparts used as standards. Sample C9e (1000 mg NCC, 4 mL of LPE in 10 mL of 10 mM AgNO3 and made up to 50 mL), proved to be an efficient catalyst for a three-component reaction. This led to the catalytic preparation of nine benzylidene-bis-(4-hydroxycoumarin) derivatives (6a – 6i).
Catalytic ring-opening polymerization of cyclic esters to biodegradable polyesters using N,N'- and N,O-ligand supported Cu(II), Mg(II) and Zn(II) complexes.
(2018) Munzeiwa, Wisdom.; Owaga, Bernard Omondi.; Nyamori, Vincent Onserio.
Over the past decades, there has been a tremendous increase in market demand for polyesters and their co-polymers. Of interest, polycaprolactone (PCL) and polylactides (PLA) which are biodegradable have found widespread applications in the packaging and biomedical fields. Polyesters are produced via ring-opening polymerization (ROP) process using metal-based metal-catalyst/initiators, with industrial production relying on tin(II) compounds. Despite the intense research efforts devoted to this area, there are still considerable limitations. For example, in case of chiral lactides monomers very few catalytic systems are capable of stereoselective synthesis. In addition, there is also lack of control of the polymerization process to curb side reactions which results in low molecular weight polymers with broad molecular distributions. Furthermore, the toxicological effects associated with tin compounds pose a danger if polymers are applied in the biomedical field since it is difficult to completely remove remnant catalyst from the polymer matrices. Thus, this thesis investigated the synthesis of less toxic metal complexes such as zinc, copper and magnesium supported by strategically designed ligands and their application in ROP. Four different class of ligands were explored as supports namely formamidine, N-hydroxy formamidine, Schiff base phenoxide and chiral amino pyridyl ligands and thirty complexes were synthesized and reported in this thesis. The steric and electronic properties of the ligands were fine-tuned to influence the catalytic activity and the polymer properties. The effect of the nature of the metal—oxygen bond which is prerequisite for ring-opening polymerization was investigated. Complexes with acetate and alkoxide reactive ligands were synthesised where the oxygen was not part of the ligand system. N-hydroxy formamidine and Schiff base phenoxide ligands contain the oxygen heteroatom as part of the ligand backbone. All the complexes polymerized caprolactone and lactides with appreciable activity, however for hydroxy formamidine ligands the polymerization complexes were more active only in the presence of co-initiator. The effect of auxiliary ligands such as acetates, alkoxides was also investigated. The polymerization data showed that catalytic activity depended on the metal identity, steric crowding and auxiliary ligands. Generally, zinc acetate complexes were more active achieving complete monomer conversion within 68 h compared to 120 h for the copper analogues.Magnesium-amino phenolate complexes showed greater activity, attaining 99% monomer conversion in less than 32 h as compared to 55 h for the zinc analogues. The zinc pyridyl alky and alkoxide complexes showed excellent activity, achieving 100% monomer conversion within 1 min at room temperature. Bulk substituents and electron withdrawing substituent resulted in reduced catalytic activity. All catalytic systems produced low molecular weight polymers ranging from 1200 to 10 500 g mol-1 with relatively broad molecular weight distributions and PDIs that lie between 1.2 and 2 pointing to semi-living polymerization. Chiral ligand supported catalysts showed good stereoselectivity in polymerization of rac-lactide (rac-LA) with Pr values ca 0.70. The role of the solvent was studied, and it was observed that coordinating solvent such as THF retards the polymerization as they compete with the monomer for catalytic active sites. Detailed abstracts are given in each of chapters 3 to 6.
Phytochemical, elemental and biological studies of three ficus species (moraceae) found in KwaZulu-Natal, South Africa.
(2017) Ogunlaja, Olumuyiwa Olufisayo.; Jonnalagadda, Sreekantha Babu
ABSTRACT Ficus (Moraceae), with over 800 species, is one of the understudied genera in modern pharmacognosy. Rural households depend on their fruits for food while other plant parts such as leaves and bark are utilised for medicinal purposes. Phytochemical analyses and biological activities of different plant parts, as well as the nutritional value of the edible fruits of many of the species are yet to be investigated. This study aimed at investigating three Ficus species (Ficus burtt-davyi, Ficus sur Forssk and Ficus sycomorous Linn) that produce edible fruits and are indigenous to KwaZulu-Natal, South Africa, as a source of secondary metabolites and essential dietary elements, due to their claimed medicinal and nutritional value. Plant material was subjected to chromatographic analyses and isolated compounds were identified using spectroscopic techniques and by comparison with previously reported data. Fruit and soil samples that were collected from sites within KwaZulu-Natal, were digested and analysed for macro, micro and toxic elements by Inductively Coupled Plasma-Optical Emission Spectrometry (ICP-OES). The phytochemical investigation of F. burtt-davyi revealed the bark to be rich in pentacyclic triterpenoids (lupeol and lupeol acetate) and the known antioxidant flavonoid ((+)-catechin) whilst the leaves were rich in sterols (β-sitosterol, campesterol, and stigmasterol). Both leaves and fruits also contained phaeophytin a, lutein and α-amyrin. This is the first phytochemical report on this species. The cytotoxic results indicated that lupeol and (+)-catechin, the most abundant bioactive compounds in the stem bark, were responsible for its synergistic cytotoxic effects against breast and colorectal adenocarcinoma cell lines. This study supports the use of this plant species as a substitute for antioxidant supplements and as an alternative medicine for oxidative stress related iv non-communicable chronic diseases in vulnerable communities. The phytochemical analysis of F. sur revealed two pharmacologically active triterpenoids (lupeol and sitosterol), one pheaophytin (pheaphytin a) and one flavonoid (epicatechin). The analytical results indicated that the fruits of F. burtt-davyi, F. sur and F. sycomorous are good sources of essential dietary elements and can contribute significantly (p < 0.05) to the recommended dietary allowances (RDAs) for most nutrients. The fruits of F. sur and F. burttdavyi are good dietary sources of Se and Mn, respectively. The concentrations of As, Cd, and Pb were below the instrument detection limits in all three figs indicating that the species do not tend to accumulate these toxic elements. Data from this study showed that metal interactions in soil influenced their availability, but uptake was to a greater extent controlled by the plant. Statistical analyses revealed synergistic relationships in the plants, thereby confirming that uptake of elements is controlled to meet metabolic needs. Overall, this study validates the ethnomedicinal use of these figs and reveals the nutritional and medicinal benefits of consuming the indigenous edible fruits. It also addressed the need for analytical information on the elemental concentrations in indigenous edible fruits consumed in South Africa.

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