Faculty of Applied Sciences
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Item Interaction studies of nanomaterials with plasma protein using experimental and computational methods(2018) Sabela, Myalowenkosi Innocent; Bisetty, Krishna; Sagrado, Salvador; Mwakikunga, B.Nanomaterials have received considerable attention due to their unique physicochemical properties and various applications. The present study attempts to fill in the knowledge gaps by investigating the synthesis, stability, antimicrobial, and antioxidant properties, along with their sensing capabilities. For this purpose, chemical and green methods were used to synthesise silver nanoparticles, titanium dioxide nanomaterials and zinc oxide nanoparticles. For the chemical synthesis, bio-discrimination of macromolecules were investigated by calorimetric methods. Whereas for green synthesis, silver nanoparticles and zinc oxide nanoparticles were synthesized using plant extracts and tested for cytotoxicity and antioxidant properties followed by the electrochemical sensing of silymarin. The synthesized nanomaterials were analyzed by high-resolution transmission electron microscopy, X-ray diffraction, photoluminescence, fourier transform infrared spectroscopy, thermogravimetric analyzer, dynamic light scattering and zeta potential analyzer to better understand the morphology and capping. In addition, computational modelling techniques were implemented to assess the adsorption and interaction of nanomaterial with biomolecules through docking. The results arising from this work are presented in a case-study fashion ranging from colorimetry, protein- nanomaterial interactions through to biosynthesis. For the colorimetric discrimination an extensive literature survey revealed that silver nanoparticles and gold nanoparticles based colorimetric assays, are widely used for screening in biosciences and metal analysis. A detailed review of colorimetric-based assays and identification of the fundamental parameters that influences such strategies are presented in this work. Accordingly, colorimetric discrimination of bovine serum albumin, lysozyme, single-stranded deoxyribonucleic acid, double-stranded deoxyribonucleic acid, and polynucleotides using silver nanoparticles was performed. This work demonstrates the importance of the starting material for colorimetric measurements preferably, with particles that are active in a wider absorbance region. Furthermore, an in-depth statistical analysis of the colorimetric data using principal component analysis provided a framework for sensing of the selected proteins, deoxyribonucleic acid, and polynucleotides by simultaneously varying the concentration of a salt and biomacromolecules from 10 to 100 mM and from 0.1 to 10 µg mL-1 respectively. In the case of the protein-nanomaterial interaction, the bioconjugate formation was studied between human serum albumin and TiO2 nanostructures. The TiO2 nanostructures were prepared by microwave assisted calcination at 200, 400, 700, and 900 °C. The results revealed that calcination above 400 °C had a significant impact on the surface of the nanomaterial, resulting in a lower surface adsorption of the protein confirmed by characterization techniques mentioned above. To further elucidate interactions between human serum albumin and titanium dioxide nanostructures, computational molecular docking studies were performed, resulting in the characteristic shapes of TiO2 nanostructures correlating well with the experimental observations. The nanowires and nanotubes showed the much greater binding energy of –2.33 Kcal mol-1 and -2.85 Kcal mol-1, respectively. In contrast to the nanotubes, the docked conformation of the nanowires showed a greater number of interactions. In regard to the biosynthesis of silver nanoparticles prepared from aqueous plant extracts of Iboza riparia leaf and Ilex Mitis root bark, the formation of nanoparticles were due to the reduction of silver ions caused by electron transfer from the electron rich atoms of the extracts such as saponins and diterpenes. Therefore, relevant cytotoxicity studies were conducted with the MTT assay on HEK293T followed by an antioxidant activity with ABTS. Antimicrobial activity analysis depicted susceptibility of microorganisms in the order of nanoparticles by diterpenes (156 nm) followed by nanoparticles by saponins (50 nm). Another biosynthesis of zinc oxide nanoparticles was performed using the extracts from jacaranda mimosifolia flowers and the carica papaya seeds. The composition of both extracts was analyzed by gas chromatography–mass spectrometry. Oleic acid was found to be the major component in the extract of jacaranda mimosifolia flowers. The microwave assisted biosynthesis of nanoparticles was successfully completed within 5 min resulting in 2–4 nm particle size. Further characterization was performed using the aforementioned instruments. The zinc oxide nanoparticles from jacaranda mimosifolia flowers were used to test the viability of gram-negative Escherichia coli and gram-positive Enterococcus faecium bacteria, which was found to be 48% and 43% respectively. In order to better understand the influence of oleic acid on the size and shape of nanoparticles, its adsorption on the different facets was explored computationally. The preferential passivation of the (101) facet was evident from the concentration profiles with an average distance of 1.4 Å for oleic acid. On the other hand, facets (100) and (002) were located at varying distances of 4 and 9 Å respectively. Overall the results indicated that the presence of oleic acid could drive the preferential growth of zinc oxide nanomaterials to nanorod array and other structures. Furthermore, the composition of the zinc oxide nanoparticles from the carica papaya seed extracts was also identified using the aforementioned techniques. The results highlighted the role of oleic acid as a capping agent for these nanoparticles. In relation to their electrochemical applications, they were also tested for sensing activities of silymarin by incorporating them with multiwalled carbon nanotubes immobilized on the surface of a glassy carbon electrode. The electrochemical signals obtained from the modified electrode was 2-fold higher than that of a bare glassy carbon electrode. The electrode based on zinc oxide nanoparticle was able to detect silymarin in the commercial Milk Thistle tablet.Item Biosynthesis of ZnO nanoparticles using Jacaranda mimosifolia flowers extract: Synergistic antibacterial activity and molecular simulated facet specific adsorption studies(Elsevier, 2016) Sharma, Deepali; Sabela, Myalowenkosi Innocent; Kanchi, Suvardhan; Mdluli, Phumlani Selby; Singh, Gulshan; Stenström, Thor-Axel; Bisetty, KrishnaThe naturally occurring biomolecules present in the plant extracts have been identified to play an active role in the single step formation of nanoparticles with varied morphologies and sizes which is greener and environmen-tally benign. In the present work, spherical zinc oxide nanoparticles (ZnO NPs) of 2–4 nm size were synthesized using aqueous extract of fallen Jacaranda mimosifolia flowers (JMFs), treated as waste. The microwave assisted synthesis was completed successfully within 5 min. Thereafter, phase identification, morphology and optical band gap of the synthesized ZnO NPs were done using X-ray diffraction (XRD), high resolution transmission elec-tron microscopy (HRTEM) and UV–Visible spectroscopy techniques. The composition of JMFs extract was ana-lyzed by gas chromatography–mass spectrometry (GC–MS) and the ZnO NPs confirmation was further explored with fourier transform infrared spectroscopy (FTIR). The GC–MS results confirmed the presence of oleic acid which has high propensity of acting as a reducing and capping agent. The UV–Visible data suggested an optical band gap of 4.03 eV for ZnO NPs indicating their small size due to quantum confinement. Further, facet specific adsorption of oleic acid on the surface of ZnO NPs was studied computationally to find out the im-pact of biomolecules in defining the shape and size of NPs. The viability of gram negative Escherichia coli and gram positive Enterococcus faecium bacteria was found to be 48% and 43%, respectively at high concentration of NPs.Item Fabrication of copper nanoparticles decorated multiwalled carbon nanotubes as a high performance electrochemical sensor for the detection of neotame(Elsevier, 2015) Bathinapatla, Ayyappa; Kanchi, Suvardhan; Singh, Parvesh; Sabela, Myalowenkosi Innocent; Bisetty, KrishnaA highly sensitive and novel electrochemical sensor for the detection of neotame using differential pulse voltammetry with a modified glassy carbon electrode is presented. The method was further customized by the fabrication of the electrode surface with copper nanoparticles–ammonium piperidine dithiocar-bamate–mutiwalled carbon nanotubes assimilated with β-cyclodextrin. The multiwalled carbon nano-tubes assimilated with β-cyclodextrin/glassy carbon electrode exhibited catalytic activity towards the oxidation of neotame at a potential of 1.3 V at pH 3.0. The transmission electron microscopy, thermogravimetric analysis, frontier transform infrared spectroscopy and cyclic voltammetry were employed to characterize the electrochemical sensor. The sensitivity and detection limits of the electrode increased two-fold in contrast to the β-CD-MWCNTs/GCE sensor. The developed method was successfully applied for the determination of neotame in food samples, with results similar to those achieved by our modified capillary electrophoresis method with a 96% confidence level.Item Analytical evaluation of steviol glycosides by capillary electrophoresis supported with molecular docking studies(Iranian Chemical Society, 2014-05-05) Ayyappa, Bathinapatla; Kanchi, Suvardhan; Singh, Parvesh; Dovey, Martin; Sabela, Myalowenkosi Innocent; Bisetty, KrishnaThis paper reports on a newly developed elec-trokinetic chromatographic method for the simultaneous separation and determination of steviol glycosides in real stevia samples by capillary electrophoresis and supported by molecular docking studies. Our results obtained using 30-mM heptakis-(2,3,6-tri-o-methyl betacyclodextrin) as a separating agent, suggest that at optimum experimental conditions the detection limits of 2.017 9 10-5 and 7.386 9 10-5 M and relative standard deviations (n = 5) of 1.10 and 1.17 were obtained for rebaudioside-A and stevioside, respectively. In addition, the molecular docking studies explained to a certain extent why the separation was successful. The calculated binding free energy results for the rebaudioside-A and stevioside complexes formed with the separating agent showed that although both ligands penetrated deeply into the hydrophobic cavity of the sep-arating agent, the presence of additional hydrogen bonding in the case of stevioside is probably responsible for its stronger binding affinity than that of rebaudioside-A.Item A Box-Behnken Design and Response Surface Approach for the Simultaneous Determination of Chromium (III) and (VI) Using Catalytic Differential Pulse Polarography(ESG, 2004-09-29) Sabela, Myalowenkosi Innocent; Kanchi, Suvardhan; Ayyappa, Bathinapatla; Bisetty, KrishnaThe present paper describes an optimized Box-Behnken design using a catalytic-differential pulse polarograhic technique for the simultaneous determination of chromium (III) and (VI) in wastewater samples using ammonium piperidine dithiocarbamate as a complexing agent. The optimization strategy was carried out using a two level full factorial design. The results of the experimental design were based on an analysis of variance and demonstrated that only the pH, concentrations of the buffer and the complexing agent were statistically significant throughout the experiment. The optimal values for the three variables were 8.0, 0.2 mM and 5.0 mM for pH, concentrations of the buffer and the complexing agent respectively. Under optimum experimental conditions the detection limit of the proposed method was found to be 0.0841 µg L-1 while the linear range was 1.0-10.0 and 0.5-25.0 µg L- for chromium (III) and (VI) respectively. The present method was also applied for the simultaneous determination of chromium in the presence of some foreign ions with satisfactory analytical responses.Item Polarographic catalytic hydrogen wave technique for the determination of copper(II) in leafy vegetables and biological samples†(ESG, 2013-03-01) Kanchi, Suvardhan; Singh, Parvesh; Sabela, Myalowenkosi Innocent; Naidu, N. Venkatasubba; Bisetty, KrishnaAn economical, novel, eco-friendly and robust method for the quantification of copper(II) in various leafy vegetables, milk and blood samples has been developed using direct current catalytic hydrogen wave (DC-CHW) technique involving the formation of ammonium salts of piperidine/morpholine dithiocarbamates metal complex. Ammonium piperidine/morpholine dithiocarbamates complexed with copper(II) in the presence of NH4Cl-NH4OH medium at pH 6.5±1 produces CHW’s at -0.35±1 V vs SCE. Consequently, various optimal parameters such as preparation medium, effect of pH, ligand concentration, metal ion concentration and indifferent ions effect on peak height were optimized to enhance the sensitivity and selectivity. The novel aspect in the work address the interaction of ammonium piperidine/morpholine dithiocarbamates with copper(II) were confirmed with cyclic voltammetry (CV) and supported by computational calculations using Density Functional Theory (DFT) methods. Furthermore, the student “t”-test and variance ratio “f”-tests indicated no significant difference between the present method and the differential pulse polarographic (DPP) method.Item Evaluation of enantioresolution of (±)-catechin using electrokinetic chromatography and molecular docking(SRP, 2012) Sabela, Myalowenkosi Innocent; Singh, Parvesh; Gumede, Njabulo Joyfull; Bisetty, Krishna; Sagrado, SagradoThis study involves the enantioresolution of (±) catechin with the highly sulphated beta cyclodextrin (HS-β-CD) as a chiral selector using capillary electrophoresis (CE). The purpose of this study was to be tter understand enantioresolution amongst host-guest interactions. Furthermore, molecular docking was carried out to elucidate the mechanism of the enantioselective separations of (±) catechin enantiomers obtained in Electrokinetic chroma tography (EKC). A large difference in the interaction energies observed between the two enantiomers represents significant enantiodifferentiation. Our results also suggest that the host-guest interactions between the phenyl ring of the ligand and the open cavity of the HS-β-CD are due mainly to hydrophobic interactions. Interestingly, the stronger interactions observed with (+)-catechin is consistent with the elution order observed in the CE experiments.Item Study of interaction between polyphenolic compounds and protein using computational and capillary electrophoresis techniques(2013-07-30) Sabela, Myalowenkosi Innocent; Bisetty, KrishnaThe present work involves the interaction studies of chiral compounds with the Human Serum Albumin (HSA) protein using computational and experimental methods. The HSA protein has multiple binding sites that forms the basis for its exceptional ability to interact with many organic and inorganic molecules, which makes this protein an important regulator of intercellular fluxes and the pharmacokinetic behaviour of many drugs. This study was undertaken to evaluate the related pharmacokinetic and enantioselective binding parameters of the racemic catechin enantiomers with the HSA. Accordingly, this work involved a method development for the chiral separation of a racemic compound, by capillary electrophoresis-electrokinetic chromatography (CE-EKC) with a highly sulphated beta-cyclodextrin (HS--CD) as a chiral selector. The experimental work was supported by two molecular docking studies. The first included the mimicking of the host-guest interactions between a chiral selector and an enantiomeric compound. The second study included the estimation of the pseudo enantioselective (ES) binding of catechin to HSA. Overall, it was found that CE-EKC is the preferred method for the(±)-catechin binding to HSA protein evaluation. Moreover, the technique used in this work is not restricted to HSA or polyphenols, but can also be applied to other proteins and ligands that possess chirality. Furthermore, the molecular docking approaches also proved to be very useful for the evaluation of chiral recognition systems and for elucidation of the ligand-protein interactions.Item Preparation, spectrochemical, and computational analysis of L-Carnosine (2-[(3-Aminopropanoyl)amino]-3-(1H-imidazol-5- yl)propanoic Acid) and Its Ruthenium (II) coordination complexes in aqueous solution(MDPI, 2011-12-09) Branham, Michael Lee; Bisetty, Krishna; Sabela, Myalowenkosi Innocent; Govender, Thirumala; Singh, ParveshThis study reports the synthesis and characterization of novel ruthenium (II) complexes with the polydentate dipeptide, L-carnosine (2-[(3-aminopropanoyl)amino]-3-(1H-imidazol-5-yl)propanoic acid). Mixed-ligand complexes with the general composition [MLp(Cl)q(H2O)r]·xH2O (M = Ru(II); L = L-carnosine; p = 3 − q; r = 0–1; and x = 1–3) were prepared by refluxing aqueous solutions of the ligand with equimolar amounts of ruthenium chloride (black-alpha form) at 60 °C for 36 h. Physical properties of the complexes were characterized by elemental analysis, DSC/TGA, and cyclic voltammetry. The molecular structures of the complexes were elucidated using UV-Vis, ATR-IR, and heteronuclear NMR spectroscopy, then confirmed by density function theory (DFT) calculations at the B3LYP/LANL2DZ level. Two-dimensional NMR experiments (1H COSY, 13C gHMBC, and 15N gHMBC) were also conducted for the assignment of chemical shifts and calculation of relative coordination-induced shifts (RCIS) by the complex formed. According to our results, the most probable coordination geometries of ruthenium in these compounds involve nitrogen (N1) from the imidazole ring and an oxygen atom from the carboxylic acid group of the ligand as donor atoms. Additional thermogravimetric and electrochemical data suggest that while the tetrahedral-monomer or octahedral-dimer are both possible structures of the formed complexes, the metal in either structure occurs in the (2+) oxidation state. Resulting RCIS values indicate that the amide-carbonyl, and the amino-terminus of the dipeptide are not involved in chelation and these observations correlate well with theoretical shift predictions by DFT.