Characterisation

Morphology and structure characterization of nanocomposite fibres is essential to understand the effect of nanoparticles and drug within the nanocomposites, as well as to understand the drug release mechanism from nanofibre mats, which may help todevelop drug delivery systems. Various characterization methods, including scanning electron microscopic (SEM),transmission electron microscopy (TEM), X-ray diffraction (XRD), differential scanning calorimetry (DSC), Fourier transform infrared spectroscopy (FTIR), viscometry, electrical conductivity measurement, gel permeation chromatography (GPC) and UV-vis spectrophotometry, were performed on some typical mat samples. The specific arrangements and investigation conditions are explained in the following subsections.

3.3.1 Scanning electron microscopy (SEM)

SEM was carried out to investigate the morphological structures of nanofibre mats when blending PLA with PCL and incorporating HNTs, MPs and drug (TCH) with the blending polymers. SEM employs a focused beam of high energy electrons to produce a diversity of signals at the solid specimens’ surface (Bozzola, 1992). The signals that are received from the interaction between the nanofibres and electrons are reformatted to form microscopic images.

The morphology of electrospun nanofibres was studied via an EVO 40XVP scanning electron microscope from Zeiss (Germany) at an accelerating voltage of 5 kV. Before SEM observation, the samples were sputter-coated with platinum. Fibre diameter was calculated from the SEM images by using an image analysis tool in the Zeiss Smart SEM software. For each sample, measurements were made with a minimum of 150 fibres from multiple scanned SEM images and at a rate 15 fibres per image.

3.3.2 Transmission electron microscopy (TEM)

Transmission electron microscopy was utilized to examine the morphology of HNT and MP dispersion within the nanofibres in order to confirm the XRD data. TEM is a broadly utilized characterization method for solid materials. In contrast to SEM, the beam of TEM electrons is transmitted through a thin sample of the material and assists in revealing the detailed structures that are embedded within the material (Williams and Carter, 2009).

The nanofibre mat was embedded in Araldite® epoxy resin. The following method was used to control the homogeneity and distribution of the fibres in the mat sample: a 1×1×1 cm3 volume of epoxy was polymerized for 24 h, then the fibre mat was macerated in epoxy solution and placed on the solid pre-polymerized epoxy, and subsequently the composite was re-polymerized. The composite was sectioned into 100 nm layers using a diamond knife with a Leica EM UC6 microtome, and the sections were later mounted onto carbon grids. The TEM observations were performed using a JEOL 2011 at an accelerating voltage of 200 kV to study the dispersion level of HNT and MP within the composite fibre mats.

3.3.3 X-ray diffraction (XRD)

X-ray diffraction (XRD) allows phase identification of the crystalline material structure, including degree of crystallinity, crystallite size, and structural changes.XRD measurements of prepared samples were performed in a Bruker Discover 8 X-ray diffractometer (Germany) operated at 40 kV and 40 mA using Cu-Kα radiation that was monochromatised with graphite sample monochromators in a 2θ range from 5° to 40° with a scanning rate of 0.05°/s. The d-spacing (d) corresponding to the silicate XRD peak was determined from Bragg’s equation:

nλ=2dsinθ

where θ is the diffraction position, λ is the wavelength, which is 1.54 Å for a Cu target, and n is an integer. The crystallite size (L) was calculated from the Scherrer relation:

L = Kλ / (Bcosθ)

where B is the complete width at half maxima of the crystalline peak in radians and K is the shape factor of the average crystallite. The crystallinity was determined by applying the area integration method from diffracted intensity over the range of 2θ data from 8° to 27° (Ning et al., 2007).

3.3.4 Differential scanning calorimetry (DSC)

DSC is a common thermo-analytical technique that is used to understand effects of heating and cooling on polymer behaviour and also study a polymer’s thermal transitions (Menczel et al., 2009). This technique was employed to determine the impact of changing the PCL concentration, blending ratio of PLA to PCL, embedding of HNTs, embedding of MPs and the addition of drugs on the thermal properties of the composites. Thermal analysis was performed using a DSC6000 from PerkinElmer (USA) with Cryofill liquid nitrogen cooling system. Approximately 10mg of fibre mat was sealed in aluminium pans and its thermal behaviour was analysed during heating between 30°C and 200°C with a ramp rate of10°C/min for PLA nanofibres, while samples with PCL were heated from –100°C to 200°C. The glass transition temperature (Tg), crystallisation temperature (Tc) and melting temperature (Tm) were obtained for PLA: PCL nanofibres from analysis of the DSC data.

3.3.5 Fourier transform infrared spectroscopy (FTIR)

FTIR is a method of materials analysis that produces the ‘fingerprint’ of a material with absorption peaks that match the frequencies of vibrations of the material’s atomic bonds. It can identification of different compounds because each material is a unique combination of atoms (Smith, 2009). FTIR was used to investigate the interaction level between biopolymers (PLA: PCL), HNTs, MPs and drugs. It was performed in a Spectrum 100 FTIR Spectrometer from PerkinElmer (Japan). Spectra were recorded in the range between 4000–550 cm−1 with 4 cm−1 resolution by using an attenuated total reflectance (ATR) technique (Chittur, 1998).

3.3.6 Viscometry and electrical conductivity measurement

Solution viscosity, which is a very important variable that influences nanofibre diameters, was measured by using a Visco 88 portable viscometer from Malvern Instruments (UK). It has a built-in temperature sensor and is supplied with a double gap measuring geometry to provide extra sensitivity when measuring low-viscosity fluids. All the samples were measured under constant temperature of 23°C. The electrical conductivity and pH value of the solution were measured by using a WP-81 Waterproof pH and conductivity meter (TPS, Australia).

3.3.7 Gel permeation chromatography (GPC)

Gel permeation chromatography (GPC) was utilized to find the effect of impurities that accompany the MPs on the molecular weight of PLA: PCL blending. GPC analysis was performed to measure the molecular weight of polymers at 40°C using a Shodex DS-4 pump, one Shodex KF-805L and three KF803L columns connected in series, and a UV detector. Tetrahydrofuran (THF) was used as the eluent at a flow rate of 1.0 ml/min.

3.3.8 UV-vis spectrophotometry

UV-vis spectrophotometry is a method that has been used to identify and measure the organic and inorganic materials in many processes. The amount of TCH present in the release buffer (PBS) was obtained by means of a UV-vis spectrophotometer (JascoV-67) at wavelengths of 360 nm and 319 nm of TCH and IMC respectively.