Biodegradable biomaterials have been wildly considered and used in current
for medical and related application (Piskin, 1995).
Generally, it divided into synthetic and natural polymers. The development of
biodegradable biomaterials specifically developed for novel biomedical technologies
including tissue engineering, regenerative medicine, gene therapy, controlled
drug delivery and bionanotechnology (Nair and Laurencin,
Advantages of polymeric materials such as biocompatibility, biomimetic structures,
mechanical and degradation properties are requires. Synthetic biodegradable
polymers have been commercially developed including poly (α-ester), polyurethanes,
poly (ester amide), poly (ortho esters), polyanhydrides, poly (anhydride-co-imide),
cross-linked polyanhydrides, poly (propylene fumarate), pseudopoly (amino acid),
poly (alkyl cyanoacrylates), polyphosphazenes and polyphosphoester. On the other
hand, natural polymers have been reported including protein and poly (amino
acids) and polysaccharides (Nair and Laurencin, 2007).
Silk, a natural protein polymer has numerous studied as technical and biomedical
applications (Min et al., 2004). It is produced
by Lepidopteran insects of the family Bombycidae and Saturniidae (Mandal
et al., 2009). SF can be used in various forms including gel, powder,
fiber or membrane, depending on application (Park et
al., 2004). Each silk fiber is composed of fibroin core protein coated
with sericin (Servoli et al., 2005). Silk Fibroin
(SF) has been used as biotechnological and biomedical resources since their
unique properties including nontoxicity, biocompatibility and biodegradability
were reported (Foo and Kaplan, 2002; Altman
et al., 2003).
Gelatin (G) is a derivative collagen. It is suitable as a biomaterial for tissue
engineering. Gelatin shows excellent properties including low cost, good biocompatibility,
biodegradability, low immunogenicity, increased cell adhesion, migration, differentiation
and proliferation (Thein-Han et al., 2009). In
the two past decades, gelatin has been wildly used as sealants for vascular
prostheses, carriers for drug delivery, wound dressings, health caring devices
and tissue engineering scaffolds (Huang et al., 2004;
Gil et al., 2007; Lee et
al., 2008). Moreover, it can be applied alone or as a blend polymer
(Huang et al., 2004).
In this study, SF/G blend film was prepared and investigated for their morphology, secondary structures as well as thermal properties. The goal of this work is extended to know about two biomaterials characteristics for material design and applications.
MATERIALS AND METHODS
This study was done from July 1, 2009 to November 10, 2009. Most of the experiment was carried out at Department of Chemistry and the Central Instrument Center, Faculty of Science, Mahasarakham University.
Materials: Bombyx mori (B. mori; locally called Nang-Lai) silk cocoons were kindly supplied from Silk Innovation Center (SIC), Mahasarakham University, Thailand. The cocoons were kept in air-dried room until use. Gelatin powder was purchased from Fluka. All used chemicals were analytical grade obtained commercially.
Preparation of SF solution: The B. mori cocoons were firstly
striped into small pieces and then washed with distilled water to exclude those
of impurity, dried in oven at 40°C. The sericin protein was extracted from
the cocoons by boiling twice with 0.5% (w/v) Na2CO3 solution.
Each time, the cocoons were thoroughly rinsed in distilled water. They were
then air-dried at room temperature. The dried cocoons were dissolved with the
tertiary solvent system of CaCl2-Ethanol-H2O (1:2:8 by
mole ratio). The mixture of solvent and striped cocoons was boiled at 90-95°C
for 1 h and stirring. The SF solution was obtained and then dialyzed in dialysis
bag (Mw. cut off 7500 kDa) against distilled water for 3 days. SF concentration
was calculated by evaporation method and adjusted to 1% (w/v).
Preparation of gelatin solution: 1% (w/v) of gelatin solution was prepared by dissolving 1 g of gelatin powder using distilled water. The mixture was stirred and stand at room temperature until absolutely dissolved.
SF/G blend films: Various ratios of SF/G blend films were prepared by casting on the 5 cm polystyrene plates at room temperature. The plates were dried at 40°C in oven for 3 days.
Morphology observation: The blend films were cut and sputter coated with gold to enhance their surfaces conductivity before observation under Scanning Electron Microscope (SEM) (JEOL, JSM-6460LV, Tokyo, Japan).
Secondary structure analysis: Secondary structure of the blend films were analyzed using Fourier transform infrared (FTIR) spectrometer (Perkin Elmer-Spectrum Gx, USA) with resolution of 2 cm-1 and 32 scans. The intensity at wave numbers from 500 to 4000 cm-1 was recorded.
Thermal behavior measurement: The SF/G blend films were loaded in a platinum crucible. The Thermo Gravimetric Analysis (TGA) was performed using TA instruments, SDT Q600 (Lukens drive, New Castle, DE). The samples were non-isothermal heated from 50°C to 700°C at a heating rate of 20°C min-1. The TGA was carried out in nitrogen with the flow rate of 100 mL min-1. The differential TG (DTG) was recorded with TA instruments Q series explorer software. The analyses of the data were done using TA Instruments Universal Analysis 2000 software (version 3.3B).
Fourier transform-infrared (FT-IR) spectroscopy: SF film showed many absorption bands at 1690, 1654 and 1636 cm-1 (amide I), 1558 and 1522 cm-1 (amide II) and 1242 cm-1 (amide III) (Fig. 1a). The FT-IR spectrum of gelatin (G) film showed absorption bands at 1621, 1567, 1546 and 1242 cm-1 (Fig. 1c). In SF/G blend film, the spectrum indicated similar characteristic bands of the native polymers (Fig. 1b). Blend with G led to shifting of amide I bands (1678, 1653 and 1634 cm-1), amide II (1559 and 1526 cm-1), except amide III (1242 cm-1) in the spectrum of SF.
Scanning Electron Microscopy (SEM): The morphology of SF, G and SF/G films were observed by SEM. It was found that the surface of the blend film (Fig. 2b) has uniform without phase separation, compared to both SF (Fig. 2a) and G (Fig. 2c). The surface area of SF film is highly rough in comparison with other. In addition, SF/G blend film showed smoother surface than that of SF film.
Thermal analysis by TG and DTG: The function of %wt loss vs. temperature
of SF, G and SF/G were studied by TG. In case of SF, two-stage decomposition
was occurred in the temperature range 225-310°C and 310-340°C (Fig.
3). In addition, minor peak was also found in the temperature range 340-400°C.
In case of G film, also two-stage decomposition was found in the temperature
range 225-330°C and 330-360°C, with the shoulder peak in temperature
range 360-440°C. However, In case of SF/G blend film, at least four-stage
decomposition was found in temperature range 225-300°C, 300-338°C, 338-355°C
and 355-440°C, respectively (Fig. 3). DTG analysis of
all films was also studied as a function of rate of weight loss vs temperature.
||SEM micrographs of SF/G blend films; (a) 3/0, (b) 1/1 and
(c) 0/3 (w/w). Bars = 10 μm
||FT-IR spectra of SF/G blend films; (a) 3/0, (b) 1/1 and (c)
||TG curves of SF/G blend films; (a) 3/0, (b) 1/1 and (c) 0/3
||TG and DTG curves of SF film
and DTG curves of film
The maximum decomposition temperature (Td, max) of SF
film was found at 298, 315 and 392°C, respectively (Fig. 4),
whereas G film was found at 323, 335 and 362°C, respectively (Fig.
5). Mixture characteristic between SF and G was found in SF/G blend film
since the Td, max was indicated at 294, 309, 334 and 346°C, respectively
||TG and DTG curves of SF/G blend film
In this study, SF of B. mori from Thai strain (locally called Nang Lai)
solution was blended with G in order to study their morphology, secondary structure
and thermal properties. The films were successfully prepared by using evaporation
technique. Both SF and G have been reported as an excellent biomaterials used
in various fields (Thein-Han et al., 2009; Patel
et al., 2008; Fan et al., 2008;
Gil et al., 2007). SF indicated various excellent
properties, especially mechanically robust. It was found that the amino compositions
and their structures are main factor on this property. If the structures were
changed the SF properties were changed too. Generally, the secondary structures
of protein are indicated by the amide groups of the silk composition (Kweon
et al., 2000; Hino et al., 2003). In
this work, FT-IR data indicated that the secondary structures of SF film showed
1690, 1655 and 1636 cm-1 (amide I, C=O stretching), 1558 and 1522
cm-1 (amide II, N-H deformation), 1242 cm-1 (amide III,
C-N stretching), assigned to mixture of α-helix and β-sheet forms
(Li et al., 2003; Mandal
et al., 2009). The gelatin film was characterized by those carbonyl
and amino peaks at 1631, 1545 and 1244 cm-1 which are due to C =
O stretching, amide II and amide V, respectively (Thein-Han
et al., 2009). In SF/G blend, FT-IR spectrum depicts similar characteristic
bands of the native materials. The intensity of the absorption bands concerned
the C = O and amide II at about 1678, 1653, 1634 cm-1 and 1559, 1526
cm-1. This result suggested a chemical interaction between SF and
G molecular chains and intermolecular bonding of hydrogen of SF and G formation.
It is might be suggested that SF provides as area for G in the binding and structure
in the blend film (Thein-Han et al., 2009).
SEM micrographs revealed uniform throughout the surface area. The micrographs reflected that SF and G form a continuous phase in the blend film. Thermal decomposition of SF film took place in a similar step with G film whereas SF/G blend film has different points. The blend film took maximum decomposition temperature characteristics of both SF and G indicated that the interactions between two materials are formed. However, DTG studies are evident that SF/G blend film is not thermally more stable than the native films.
The morphology, secondary structure and thermal properties of SF, G and SF/G blend films were reported in this study. The results showed that all of films showed uniform throughout surface area under SEM. The secondary structure of SF slightly changed after blending with G which was observed by the FT-IR. It was suggested that SF and G could be formed interaction together. TG and DTG showed that SF/G blend is not enhancing thermal stability of the film since it has decomposition temperature pattern as similar as native materials.
We appreciate to thank Division of Support and Development, Mahasarakham University for financial support of this study.