Abstract of thesis presented to the Senate of Universiti Putra Malaysia in
fulfillment of the requirement for the degree of Master of Science.
DEVELOPMENT OF NANOANTIBIOTIC DELIVERY SYSTEM USING COCKLE SHELL DERIVED ARAGONITE NANOPARTICLES FOR TREATMENT OF OSTEOMYELITIS
By
LAMIN SAIDYKHAN
June 2015
Chairman: Professor Md Zuki Bin Abu Bakar @ Zakaria, PhD
Faculty: Institute of Bioscience
Osteomyelitis is one of the most devastating orthopedic diseases especially during its chronic stages where avascularity and loss of bone tissue are quite prevalent. Avascularity or vascular insufficiency hinders drugs from reaching infected tissues, thereby rendering parenteral antibiotics administration inefficacious. This bone infection problem has been aggravated by Stapylococcus aureus (S. aureus) which has been further compounded by the emergence of methicillin resistant S. aureus (MRSA). Local drug delivery system (LDDS) with biodegradable drug vehicles has been recognized to be the most effective therapeutic approach for the treatment of osteomyelitis. However, the design of biodegradable LADS with high therapeutic efficacy is too costly and demanding. An ideal LDDS should most importantly be able to sustainably supply antibiotics at concentrations many times higher than the minimum inhibitory concentration (MIC) of the pathogen and simultaneously promote bone regeneration. In this research, a low cost and facile method was used to design vancomycin-loaded CaCO3 (aragonite) nanoparticles (VCNP) from cockle shells with the aim of understanding its potential in developing a therapeutic bone implant for the treatment of osteomyelitis.
The nanoparticles were synthesized via mechanical stirring of micron-sized cockle shell powder in the presence of a surfactant-N-dodecyl-N, N-dimethyl-3-ammonio-1-propanesulfonate (SB-12) for 90 minutes at 80ºC. Vancomycin-loaded nanoparticles were prepared with several nanoparticles to antibiotic ratios and the formulation with the highest drug content and encapsulation efficiency (EE) was used for in vitro release study and antimicrobial evaluation. Physiochemical characterization of naked CNP and VCNP was performed by transmission electron microscopy (TEM), scanning electron microscopy (SEM), Fourier transform infrared (FT-IR) spectroscopy, X-ray powder diffraction (XRD), and Zetasizer. Finally, MTT-cytotoxicity assay was performed with human osteoblast (hFOB 1.19) cell line to investigate the cytotoxicity of VCNP.
Cubic-shaped nanoparticles with average particle sizes of 34 ± 5 nm and pure aragonite polymorphs were obtained as a result of the influence of inter-particle collision and temperature. The formulation with nanoparticles to drug ratio of 1:4 has the highest EE of 54.05%. No significant differences were observed between the naked CNP and VCNP in terms of size and morphology as both samples were in cubic-shaped with sizes of approximately 35 nm. Successful loading was attributed to the porosity of the nanoparticles, negative charge density (-19.4 ± 3 mV) and interactions with vancomycin molecules as indicated by SEM, zetasizer, and FTIR analysis respectively. VCNP displayed a 120 hours (5 days) release profile of vancomycin, which expressed high bactericidal effect at approximately 145 times MIC of MRSA. The cell proliferation assay showed 80% cell viability of VCNP at the highest concentration (250 µg/mL) indicating good biocompatibility of VCNP. The findings of this research confer VCNP to be of high potential in the development of nanoantibiotic bone implants that can be used in local antibiotic delivery therapy against osteomyelitis with optimal antibacterial efficacy, good bone resorbability and biocompatibility. This study deduced that cockle shell-based nanoparticles with different morphologies could be synthesized by simple manipulation of experimental variables like temperature and surfactants. In addition, CNP can be tapped for its potential as a drug vehicle for many MRSA-induced and bone diseases, especially if the work is extrapolated in an in vivo study.
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