Synthesis of Keratin-based Nanofiber for Biomedical Engineering

Summary

Electrospun nanofibers have a high surface area to weight ratio, excellent mechanical integrity, and support cell growth and proliferation. These nanofibers have a wide range of biomedical applications. Here we fabricate keratin/ PCL nanofibers, using the electrospinning technique, and characterize the fibers for possible applications in tissue engineering.

Abstract

Electrospinning, due to its versatility and potential for applications in various fields, is being frequently used to fabricate nanofibers. Production of these porous nanofibers is of great interest due to their unique physiochemical properties. Here we elaborate on the fabrication of keratin containing poly (ε-caprolactone) (PCL) nanofibers (i.e., PCL/keratin composite fiber). Water soluble keratin was first extracted from human hair and mixed with PCL in different ratios. The blended solution of PCL/keratin was transformed into nanofibrous membranes using a laboratory designed electrospinning set up. Fiber morphology and mechanical properties of the obtained nanofiber were observed and measured using scanning electron microscopy and tensile tester. Furthermore, degradability and chemical properties of the nanofiber were studied by FTIR. SEM images showed uniform surface morphology for PCL/keratin fibers of different compositions. These PCL/keratin fibers also showed excellent mechanical properties such as Young's modulus and failure point. Fibroblast cells were able to attach and proliferate thus proving good cell viability. Based on the characteristics discussed above, we can strongly argue that the blended nanofibers of natural and synthetic polymers can represent an excellent development of composite materials that can be used for different biomedical applications.

Introduction

Electrospinning is recognized as a prevalent method of achieving polymer nanofibers. The fibers can be produced on a nanoscale and the fiber properties are customizable ¹. These developments and the characteristics of electrospun nanofibers have been especially interesting for their applications in biomedical engineering especially in tissue engineering. The electrospun nanofibers possess similarities to the extracellular matrix and thus promote cell adhesion, migration and proliferation². Due to this similarity to the extracellular matrix (ECM), electrospun fibers can be used as materials to assist in wound dressing, drug delivery, and for engin....

Protocol

All protocol follows the guidelines of the North Carolina A&T State University Office of Research Compliance and Ethics.

1. Chemical Preparation for Keratin Extraction ⁴

1. To prepare 1,000 ml of 2% wt/vol peracetic acid solution (PAS), under a fume hood add 20 ml of peracetic acid to 980 ml of Deionized (DI) water.
2. To prepare 1,000 ml of 100 mM Tris base solution (TBS), add 12.2 g of Tris Base to 1,000 ml of DI water and stir until completely dissolved.
3. Prepare diluted hydrochloric acid solution (DHAS) in a fume hood by pouring 4 ml of concentrated hydrochloric acid into 30 ml of DI water.
4. Procure approxima....

Results

Fiber Morphology
SEM images of the fibers were obtained for all the fiber compositions. See Figure 3. Fiber image confirms that the fibers are randomly oriented.

Mechanical Testing
Mechanically strong fibers are generally required for various tissue engineering applications. These fibers should retain sufficient strength and flexibility under certain stress and .......

Discussion

Extraction of keratin from human hair was successfully achieved. The peracetic acid acted as an oxidizing agent on the human hair, allowing the keratin to be extracted by the Tris Base. The production of keratin powder was small scale due to the fact that it was only done for research purposes. This procedure has already been established in industry for large-scale production. The purpose of extracting the small-scale keratin was to control contamination, batch variability, and cost-effectiveness.

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Human Hair			Obtained from Local Barber Shop in Greensboro
Peracetic acid	Sigma Aldrich
PCL (e-caprolactone polymer)	Sigma Aldrich	502-44-3	Mn 70-90 kDa
Trifluoroethanol (TFE)	Sigma Aldrich	75-89-8
Tris Base (TrizmaTM Base Powder)	Sigma Aldrich		>99.9% crystalline
Hydrochloric Acid	Fischer Scientific	A144C-212 Lot 093601	Waltham, MA
Kwik-Sil	World Precision Instruments		Sarasota, FL
Cellulose membrane	Sigma Aldrich		12 - 14 kDa molecular cut off
optical microscope	Olympus BX51M	BX51M	Japan
scanning electron microscope	Hitachi SU8000	SU8000	Japan
Table-Top Shimadzu machine	North America Analytical and Measuring Instruments AGS-X series	AGS-X Series	Columbia, MD
Fourier transform infrared spectroscopy	Bruker Tensor 2 Instrument		Billerica, MA
Microcal Origin software			Northampton, MA
X-ray diffraction (XRD)	Bruker AXS D8 Advance X-ray Diffractometer		Madison, WI
Fibroblast 3T3  cell	American Tissue Type Culture Collection		Manassas, VA
Dulbecco's modified Eagle's medium (DMEM	Invitrogen		Grand Island, NY
Spectra max Gemini XPS microplate reader	Molecular Devices		Sunnyvale, CA
Student- Newman-Keuls post hoc test	SigmaPlot 12 software