Innovative Self-Assembling Nanodelivery Systems to Combat Bacterial Infections by Mohammed Abdeen Mohammed Salih (M

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Innovative Self-Assembling Nanodelivery Systems to Combat Bacterial Infections by Mohammed Abdeen Mohammed Salih (M Innovative Self-assembling Nanodelivery Systems to Combat Bacterial Infections by Mohammed Abdeen Mohammed Salih (M. Pharm, National Ribat University, Sudan) Submitted as fulfilment of the requirements for the degree of Doctor of Philosophy in Pharmaceutics at the Discipline of Pharmaceutical Sciences of the School of Health Sciences at the University of KwaZulu-Natal Supervisor: Professor Thirumala Govender (PhD, University of Nottingham, United Kingdom) Date submitted: 23 July, 2020 “The ultimate pursuit of learning is to be able to ask better questions. The one who is able to do so, will equip himself to explore the answers” -Ahmed Salem- "This thesis is dedicated to my beloved parents and family for their endless love, support and encouragement.” Declaration 1-Plagiarism I, Mr. Mohammed Abdeen Mohammed Salih, declare that 1. The research data reported in this thesis, except where otherwise indicated is my own original work. 2. This thesis has not been submitted for any degree or examination at any other university. 3. This thesis does not contain data, pictures, graphs, or other information belonging to other people, unless specifically acknowledged as being sourced from other people. 4. This thesis does not contain any other person‘s writing, unless specifically acknowledged as being sources from other researchers. Where other written sources have been quoted, then: a. Their words have been rephrased but the general information attributed to them has been referenced; b. Where their exact words have been used, their writing has been placed inside quotation marks, and referenced. 5. Where I have reproduced a publication of which I am an author, co-author, or editor, I have indicated in detail which part of the publication was written by myself alone and have fully referenced such publications. 6. This dissertation does not contain any graphics, text or tables copied from the internet, unless specifically acknowledged, and the source being detailed in the reference sections of the dissertation. Signed: Date: 23 July 2020 I, Professor Thirumala Govender as supervisor of the Ph.D. studies hereby consent to the submission of this Ph.D. thesis. Signed: Date: 23 July 2020 i Declaration 2-Publications Mr. Mohammed Salih contributed to the literature search and evaluations, conceptualization, design of the projects, modification, optimization of methods, preparation, characterization in terms of synthesis and structural elucidation techniques such as Fourier- transform infrared (FT-IR) spectroscopy, Proton and 13Carbon nuclear magnetic resonance spectroscopy (1H NMR and C13 NMR), 2D Nuclear Overhauser Effect measurements (NOESY spectrum), and the bio-safety of all the materials synthesized for the studies in this thesis. He also contributed to the formulation and characterization of the nano-based drug delivery system that was formulated from the synthesized materials in terms of particle size, polydispersity index, zeta potential, surface morphology, entrapment efficiency, in vitro drug release, differential scanning calorimetry (DSC), protein binding with Microscale Thermophoresis (MST) technique, in vitro and in vivo antimicrobial activity, the mathematical modeling in terms of the in vitro release kinetics data and stability studies and molecular dynamics (MD) simulations studies. Mr. Mohammed Salih also wrote the first drafts of all the first authored papers and undertook all revisions. Dr. Mr. C.A Omolo assisted with the inception, overall design of the project, solving any technical problems and supervision of the studies. Dr. Nikita Devnarain assisted in performing the cytotoxicity studies of the synthesized materials, assisted by performing MD simulation studies and proof reading of manuscripts and thesis chapters. Dr. Ayman Y. Waddad assisted in the characterization of the synthesized materials and proofreading of manuscripts. Dr. Pavan Walvekar assisted in the synthesis of the PEGylated fusidic acid and formulation. Dr. C. Mocktar supervised the in vitro and in vivo antibacterial activity studies. Prof. T. Govender served as supervisor and was responsible for project conceptualization, problem-solving, editing of papers and abstracts as well as general supervision of all studies. Research output from the dissertation 1. First authored Publications The following research papers were published as results generated from specific objectives from this study and they include: Salih M, Omolo CA, Agrawal N, Walvekar P, Waddad A.Y, Mocktar C, Ramdhin C, Govender T. 2020. Supramolecular amphiphiles of Beta-cyclodextrin and Oleylamine for enhancement of vancomycin delivery. International Journal of Pharmaceutics. 574:118881. (Impact factor = 4.320). DOI: 10.1016/j.ijpharm.2019.118881. ii Salih, M., Omolo, C. A., Devnarain, N., Elrashedy, A. A., Mocktar, C., Soliman, M. E. S., & Govender, T. (2020). Supramolecular Self-Assembled Drug Delivery System (SADDs) of Vancomycin and Tocopherol Succinate as an Antibacterial Agent: In vitro, In silico and In vivo Evaluations. Pharmaceutical Development and Technology. (Impact factor 2.374) doi:10.1080/10837450.2020.1797786 Salih M, Walvekar P, Omolo CA, Elrashedy A. A, Devnarain N, Waddad A.Y, Fasiku V, Mocktar C, Govender T. A Self-Assembled Polymer Therapeutic for Simultaneously Enhancing Solubility and Antimicrobial Activity and Lowering Serum Albumin Binding of Fusidic acid. Journal of Biomolecular Structure & Dynamics (Impact Factor 3.22) (manuscript ID: TBSD-2020-1529). (In Press). 2. Conference Presentations The following local and international conference presentations were produced from the data generated during the doctoral study: Salih M, Omolo CA, Agrawal N, Walvekar P, Waddad A.Y, Mocktar C, Ramdhin C, Govender T. 2020. Supramolecular amphiphiles of Beta-cyclodextrin and Oleylamine for enhancement of vancomycin delivery. UKZN Nanotechnology Platform Symposium, 9th October 2018, Durban, South Africa (Poster Presentation). Salih M, Omolo CA, Agrawal N, Walvekar P, Waddad A.Y, Mocktar C, Ramdhin C, Govender T. 2020. Supramolecular amphiphiles of Beta-cyclodextrin and Oleylamine for enhancement of vancomycin delivery. ICONAN 2019, 16-18 October 2019, Munich, Germany (Poster Presentation). Salih M, Omolo CA, Devnarain N, Elrashedy A. A, Mocktar C, Mahmoud E. S. Soliman M. E, Govender T. Supramolecular Self-Assembled Drug Delivery System (SADDs) of Vancomycin and Tocopherol Succinate as an Antibacterial Agent: in vitro, in silico and in vivo Evaluations. College of Health Sciences Annual Research Symposium, University of KwaZulu Natal, 1st November 2019 Durban, South Africa. (Oral presentation). 3. Co-authored publications The following are co-authored publications related to the studies on the synthesis of advanced materials for combating antimicrobial resistance. iii Walvekar P, Gannimani R, Salih M, Makhathini S, Mocktar C, Govender T. Self- assembled oleyl amine grafted hyaluronic acid polymersomes for delivery of vancomycin against methicillin-resistant Staphylococcus aureus (MRSA). Colloids and Surfaces B: Biointerfaces. 2019 Oct 1;182:110388. (Impact Factor = 3.973) DOI: 10.1016/j.colsurfb.2019.110388. Nikita Devnarain, Nawras Osman, Victoria Fasiku, Sifiso Makathini, Mohammed Salih, Usri Ibrahim and Thirumala Govender. (2020). Intrinsic Stimuli-Responsive Nanocarriers for Smart Drug Delivery of Antibacterial Agents – An In-Depth Review of the Last Two Decades. WIREs Nanomedicine & Nanobiotechnology. Manuscript ID: NANOMED-651. (Impact Factor = 7.689) (In press). iv Abstract The rising surge of bacterial resistance puts an immense economic and social strain on the healthcare system worldwide. Unfortunately, the production of new antibiotics is significantly outstripped by existing therapies that are losing their effectiveness. Limitations associated with conventional dosage forms are one of the main contributing factors for increasing antimicrobial resistance. Novel nano-drug delivery systems have immense potential for overcoming antimicrobial resistance. This study broadly aimed to design advanced materials and explore nano-based strategies for preparation of self-assembling delivery systems to combat Staphylococcus aureus (S. aureus) and methicillin-resistant Staphylococcus aureus (MRSA) infections. In this study, three novel self-assembling systems; supramolecular amphiphilic Beta-cyclodextrin and Oleyl amine (BCD-OLA), supramolecular self-assembled drug delivery system (SADDs) for enhancement of vancomycin (VCM) delivery, and self-assembling PEGylated Fusidic acid (PEG-FA) as a polymer therapeutic were designed, synthesized and employed for the formulation of nano- drug delivery systems for efficient delivery of antibiotics. All the newly synthesized systems were confirmed and characterized by FTIR, DSC, NMR and molecular dynamic (MD) simulations. The synthesized materials and the formulated delivery system were found to be biosafe after exhibiting cell viability above 75% in all human cell lines tested using the MTT assay. The formulated nano-based systems were evaluated for sizes, polydispersity indices (PDI), zeta potential (ZP), surface morphology, drug release, in vitro and in vivo antibacterial activity. The formulated BCD-OLA nanovesicles size was shown to be 119.8 ± 1.12 nm with a PDI of 0.220 ± 3.98, and ZP of 25.8 ± 6.96 mV. The formulated SADDs for VCM delivery displayed a size of 85.15 ± 0.4 nm with PDI of 0.131 ± 0.017 and ZP of -27 ± 1.3 mV. The encapsulation efficiency of VCM in both formulated BCD-OLA and VCM/TS nano-system was 40.2 ± 4.5% and
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