Electrospinning of Hyaluronic Acid (HA) and HA/Gelatin Blends

114 DOI: 10.1002/marc.200500726 Communication 

Summary: In the present study, electrospinning of hyaluronic 

acid (HA) and hyaluronic acid/gelatin (HA-GE) blends 

in N,N-dimethylformamide (DMF)/water-mixed solvents 

have been investigated. When the volume ratio of DMF to 

water was in the range of 1.5–0.5, HA solutions could be 

electrospun into fibrous membranes successfully. The average 

diameter of HA fibers was about 200 nm. The HA-GE 

composite nanofibrous membranes with varied HA/GE 

weight ratio in the range of 100/20–100/100 have also been 

successfully fabricated. The average diameter of HA-GE 

fibers was in the range of 190–500 nm. The decrease in surface 

tension could promote fiber formation. Thus, an introduction 

of DMF that could decrease the surface tension 

distinctively, without significant change or increase in viscosity 

of the solution, could bypass the use of blowing-assisted 

electrospinning. Our postulated picture is that the lower 

surface tension could help the ejection of stream with 

relatively high viscosity and reduce or prevent the droplet 

formation during the spinning process. 

Electrospinning of Hyaluronic Acid (HA) and HA/ 

Gelatin Blends 

Junxing Li, 1 Aihua He,* 1 Charles C. Han,* 1 Dufei Fang, 2 Benjamin S. Hsiao, 2,3 Benjamin Chu 2,3 

1 State Key Laboratory of Polymer Physics and Chemistry, Joint Laboratory of Polymer Science and Materials, 

Institute of Chemistry, Chinese Academy of Sciences, Beijing 100080, China 

Fax: (þ86) 10 62521519; E-mail: aihuahe@iccas.ac.cn; c.c.han@iccas.ac.cn 

2 Department of Chemistry, Stony Brook University, Stony Brook, NY 11794-3400 

3 Stonybrook Technology and Applied Research (STAR), Stony Brook, New York, 11794-3400, USA 

Received: October 25, 2005; Accepted: November 10, 2005; DOI: 10.1002/marc.200500726 

Keywords: electrospinning; gelatin (GE); hyaluronic acid (HA); nanofibers 

Introduction 

The electrospinning technique is a simple and effective 

method for production of ultra-thin fibers with diameters 

ranging from submicrons to a few nanometers; and the electrospun 

fibrous products possess useful characteristics such 

as, high-specific surface area, high aspect ratio, and high 

porosity with very small pore size, which can mimic the 

extracellular matrix (ECM) and enhance the cell migration 

and proliferation, and are especially suitable for biomedical 

applications, including tissue engineering scaffolds, wound 

dressings, drug delivery, medical implants, and others. [1–12] 

In recent years, studies on electrospinning of new biopolymers 

and with the use of novel apparatus have drawn a 

great deal of attention. Although most studies of electro- 

HA/GE (100/80) nanofibrous membrane produced by 

electrospinning. 

spinning focus on synthetic polymers, such as PLA, [13] 

PCL, [11,14–16] PLGA, [17,18] PLLA, [19] PVA, [20] PU, [21] and 

PEO, [1,22] only a limited number of natural biopolymers 

have been successfully electrospun. Compared with 

those synthetic polymers, natural biopolymers generally 

offer better biocompatibility and biodegradability, and 

are more suitable for biomedical applications. However, 

their processability is, in general, fairly poor. Electrospinning 

of these natural biopolymers is often more 

difficult when compared with synthetic polymers. [23] Some 

natural biopolymers have been successfully electrospun, 

including silk fibroin, [24,25] collagen, [26] gelatin (GE), [23,27] 

chitin and chiston, [28–30] DNA, [31] and hyaluronic acid 

(HA). [9] GE/PCL [23] and PLGA/chitin [32] have been coelectrospun 

for the purpose of improving electrospinning 

Macromol. Rapid Commun. 2006, 27, 114–120 ß 2006 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Electrospinning of Hyaluronic Acid (HA) and HA/Gelatin Blends 115 

processability to fabricating into novel nanofibrous 

composites. 

Hyaluronic acid, as a naturally occurring linear polysaccharide, 

consists of repeating disaccharide units (b-1,4- 

D-glucuronic acid and b-1,3-N-acetyl-D-glucosamine), is 

the main component of the ECM of connective tissues, and 

has many important biological functions. [33–35] Due to its 

excellent properties of biocompatibility and biodegradability, 

HA and its derivatives have been used extensively in 

biomedical field, for example in wound dressings, tissue 

scaffolds, arthritis treatment, drug delivery, and components 

of implant materials. [35–38] However, medical studies 

on HA nanofibrous membranes have hardly been reported 

in spite of its importance in medical applications. 

Generally, it was very difficult to carry out electrospinning 

of HA aqueous solution. The molecular weight of 

native HA is typically several million. The unusually high 

viscosity and surface tension of HA were thought to be the 

key factors that hinder the electrospinning of HA solution. 

Additionally, the strong water retention ability of HA might 

lead to the fuse of nanofibers electrospun on the collector 

due to the insufficient evaporation of solvents in electrospinning. 

The fabrication of HA into nanofibrous nonwoven 

membranes from aqueous solution was successfully 

carried out only after the development of blowing-assisted 

electrospinning. [9] 

In this paper, the electrospinning process of HA and 

hyaluronic acid/gelatin (HA-GE) blends in mixed solvents 

has been investigated. The main objectives are as follows: 

(a) Explore the electrospinning process of pure HA 

solution in water and mixed solvents, find the key factor 

which hinders the electrospinning of HA aqueous solution, 

and provide the solution to this spinning difficulty. 

(b) Find a novel and simple way to electrospin HA 

solutions with the common electrospinning setup. 

(c) Investigate the effects of GE on the electrospinning 

process of HA solutions. The characteristics of GE, such as 

relatively lower molecular weight and amphiphilic property, 

might favor the electrospinning of HA solution. 

(d) Obtain HA-GE nanofibrous membranes with various 

blending ratios. The HA-GE nanofibrous membranes with 

different compositions and sized scale could have different 

clinical applications to drug delivery, wound healings, and 

tissue engineering scaffolds. 

The present report shows that with the aid of N,Ndimethylformamide 

(DMF), HA can be electrospun successfully 

with common electrospinning setup at neutral pH, 

and HA-GE aqueous solutions were first co-electrospun to 

produce HA-GE composite fibrous non-woven membranes. 

Experimental Part 

Materials 

Hyaluronic acid (sodium salt, Mw ¼ 2 000 000) was purchased 

from Dali Co. (Nanning, China). Polymers of GE Type A 

(Approx. 220 Bloom, Mn ¼ 80 000), extracted from porcine 

skin by acidic process, were purchased from Sanhesheng 

Gelatin Co. (Wenzhou, China). DMF and ethanol were 

obtained from Beijing Chem. Co. (Beijing, China). All the 

solvents were used without further purification. 

Preparation of Spinning Solution 

HA Solutions in Water System 

Transparent aqueous solutions of HA were prepared by dissolving 

a specific amount of HA powder in the mixed solvents 

consisting of distilled water (w) and ethanol (e) (w/e ¼ 9/1, 

volume ratio). Pure HA solutions with concentrations of 1.3 w/ 

v% (w in grams and v in milliliters) and 1.5 w/v% were 

obtained. 

1.5 w/v% HA Solutions in DMF/Water System 

Dissolve certain amounts of HA powder in DMF under gentle 

stirring for 10 min, and then add a specific amount of distilled 

water into the HA solution according to the volume ratio of 

DMF to water (2, 1.5, 1 and 0.5), respectively, and continue to 

stir the solution for 20 min until the solution became 

transparent. The spinning solution of 1.5 w/v% HA in DMF/ 

water system (2, 1.5, 1 and 0.5) was prepared. 

HA-GE Blends Solutions 

1.875 w/v% HA in DMF/water (volume ratio ¼ 1.5) solution 

was prepared using the same procedure as above. 1.5, 3, 4.5, 6, 

and 7.5 w/v% GE solutions with the solvent being pure water 

was prepared at 40 8C under gentle stirring for 20 min. Add the 

GE solution into the HA solution with specific volumes to 

obtain the HA-GE solutions (HA/GE ¼ 100/20, 100/40, 100/ 

60, 100/80, 100/100, weight ratio), and in each solution the 

concentration of HA was fixed at 1.5 w/v%. 

Electrospinning 

All electrospinning solutions, spinneret, and the environmental 

temperatures were controlled at 40 3 8C. The electrospinning 

solutions were placed into a 5 ml syringe with a capillary tip 

having an inner diameter of 0.3 mm. A syringe pump was used 

to feed the polymer solution and the feeding rate was fixed 

at 60 ml min 1 . A high-voltage power supply was employed 

to generate the electric field (0–50 kV). The applied voltage 

was fixed at 22 kV. The tip-to-collector distance was fixed at 

15 cm. 

Hyaluronic acid has strong water retention ability. The residual 

water could dissolve the electrospun fibers on the collector 

if the aluminum foil was used. In this study, the ethanol bath 

was used as a collector because ethanol is a poor solvent for 

both HA and GE. A piece of aluminum foil, connecting to the 

ground, was immersed in a vessel filled with ethanol. After 

electrospinning, the vessel was put into the vacuum oven at 

50 8C for 30 min to dry off ethanol and other solvents. Then the 

aluminum foil can be taken out. Through this method, HA and 

HA-GE films can be peeled off easily. 

Macromol. Rapid Commun. 2006, 27, 114–120 www.mrc-journal.de ß 2006 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

116 J. Li, A. He, C. C. Han, D. Fang, B. S. Hsiao, B. Chu 

Characterization 

The morphologies of the electrospun fibers were observed 

by means of scanning electron microscopy (SEM, HITACHI 

S-4300) at an accelerating voltage of 5 or 15 kV. The 

conductivities and the surface tensions of spinning solutions 

were measured at 40 8C by conductivity meter (DDS-307A, 

Shanghai Rex Instruments) and surface tension meter (DCAT 

21, Dataphysics) with the Wilhelmy plate method, respectively. 

The shear viscosities of the spinning solutions were 

measured by using Advanced Rheometric Expansion System 

(Ares, TA Instruments) at 40 8C. The 17 mm 16 mm couette 

geometry was used for the measurements. The shear rate was 

controlled from 1 to 1 000 s 1 . 

Results and Discussion 

Electrospinning of Pure HA 

Hyaluronic acid could be dissolved in water easily, but 

could not be dissolved in most organic solvents. In the present 

study, water-ethanol mixed solvent was used to dissolve 

HA in order to increase the evaporation rate and to 

decrease the surface tension of the HA solution. The applied 

voltage was fixed at 22 kV, and the distance between the 

spinneret tip and the collector was set at 25 cm. The electrospinning 

of HA solution at different HA concentrations 

was investigated. The electrospinning process was rather 

discontinuous as the SEM results being shown in Figure 1(a) 

and (c). It can be seen from SEM images that many large 

irregular beads with few fibers were produced on the collector. 

However, when the distance between the electrode and 

the collector was decreased to 15 cm, the membrane morphology 

became more uniform when compared with that 

electrospun at the 25 cm distance, as shown in Figure 1(b) 

and (d). The bead-string structure was formed at both 1.3 

and 1.5 w/v% HA aqueous solution and the beads were 

more like spindles in Figure 1(b) and (d). Therefore, an 

increase in the electric field strength could benefit the fiber 

formation. However, the electrospinning process under 

these conditions was not satisfactory, as large bead formation 

occurred even when the voltage was increased to 40 kV. 

It meant that it was impossible to improve the electrospinning 

process of HA aqueous solution by adjusting the 

distance and applied voltage. 

The above results are well consistent with that in ref. [9] 

Although with the assistance of air-blowing HA aqueous 

solution could be electrospun fluently, what we want to 

know is that which is the key factor hindering the electrospinning 

of HA solution with common electrospinning 

setup. 

In our study, the physical properties of HA solution have 

been studied in detail. A solvent mixture of DMF-water was 

used. DMF was chosen because it is a polar solvent with 

poor solubility for HA. HA/DMF-water solutions at a 

volume ratio of DMF to water in the range of 0.5–2:1 were 

used for electrospinning. The morphologies of obtained 

membranes are shown in Figure 2. At a volume ratio of 

Figure 1. Effect of the distance of tip to collector (d) on the morphology of electrospun 

fibers of pure HA solutions in water system ( 5k). Distilled water and ethanol mixture was 

used as solvent (volume ratio of water to ethanol was 9/1), the voltage was fixed at 22 kV. 

(a) HA 1.3 w/v%, d ¼ 25 cm; (b) HA 1.3 w/v%, d ¼ 15 cm; (c) HA 1.5 w/v%, d ¼ 25 cm; (d) 

HA 1.5 w/v%, d ¼ 15 cm. 



Figure 2. SEM images of HA (1.5 w/v%) fibers at different volume ratio of DMF to water 

( 5k). The voltage was fixed at 22 kVand the distance of tip to collector was fixed at 15 cm. 

(a) DMF/water ¼ 0.5, (b) DMF/water ¼ 1, (c) DMF/water ¼ 1.5, and (d) DMF/water ¼ 2. 

DMF to water was 0.5, the fibers were rather thin, but large 

polymer beads occurred on the collector, as shown in 

Figure 2(a). The average fiber diameter was about 120 nm. 

When the volume ratio of DMF to water was 1, the electrospinning 

process could proceed very smoothly and the 

average fiber diameter was in the order of 200 nm. When the 

volume ratio of DMF to water was 1.5, the solution still 

remained transparent (implying no HA precipitation). SEM 

images showed that uniform fibers without beads could be 

produced with an average fiber diameter of about 250 nm. It 

was found that HA could not be completely dissolved in the 

mixed solvent when the volume ratio of DMF to water 

was 2. Therefore, the proper volume ratio of DMF to water 

was 0.5–1.5:1 for HA with a molecular weight in the two 

million range. 

Table 1. Physical data of HA and HA/GE spinning solutions. 

It was found that both the surface tensions and conductivities 

of HA solutions decreased with increasing in DMF 

content, as shown in Table 1 (samples 2, 3, 4, 5, and 6). It can 

be concluded that the decrease in surface tension could 

benefit the electrospinning process and make the beads 

disappear. The average diameter of HA fibers increased 

when DMF was introduced. 

Electrospinning of HA/Gelatin Blends 

Gelatin is a natural biopolymer derived from collagen by 

controlled hydrolysis and has been widely used in biomedical 

fields. [39–43] Studies on blends of HA and GE have 

indicated that GE can endow HA with protein properties to 

improve the cell attachment and the blends of HA-GE have 

Sample HA/GE weight HA w/v% DMF/water in HA Surface tension Conductivity Average diameter 

number ratios 

solution v/v 

mN m 1 

mS cm 1 

nm 

1 100/0 1.3 0 52.99 2 870 – 

2 100/0 1.5 0 52.33 3 170 – 

3 100/0 1.5 0.5 50.95 1 612 120 

4 100/0 1.5 1 47.85 1 178 200 

5 100/0 1.5 1.5 45.91 970 250 

6 100/0 1.5 2 44.66 773 – 

7 100/20 1.5 1.5 47.46 1 250 190 

8 100/40 1.5 1.5 46.12 1 214 260 

9 100/60 1.5 1.5 44.97 1 210 320 

10 100/80 1.5 1.5 46.55 1 116 350 

11 100/100 1.5 1.5 39.83 1 067 500 



the potential use in tissue repair. [44] GE is also an ampholyte 

polymer with interesting protein characteristics and generally 

has lower molecular weight when compared with that 

of HA. The purpose of the GE used in this study was to 

improve the electrospinning processability and to endow 

the fibrous membranes with protein characteristic. Therefore, 

HA-GE/DMF-water solutions were prepared in this 

study to investigate the electrospinning process of this two 

natural biopolymer blend. The concentration of HA was 

fixed at 1.5 w/v% and the volume ratio of DMF to water was 

1.5. The electrospinning of HA-GE solutions could be 

carried out successfully with the electrospinning processes 

becoming smoother on increasing the GE content. SEM 

images at both low and high magnifications indicated that 

uniform HA-GE nanofibrous membranes could be fabricated 

(Figure 3). HA-GE fibrous membranes at different 

Figure 3. SEM images of HA-GE composite fibrous membranes. HA ¼ 1.5 w/v%, DMF/ 

water ¼ 1.5, volume ratio. Weight ratio of (a) HA/GE ¼ 100/20, (b) HA/GE ¼ 100/40, 

(c) HA/GE ¼ 100/60, (d) and (f) HA/GE ¼ 100/80, (e), (g), and (h) HA/GE ¼ 100/100. In 

each solution, the concentration of HA was fixed at 1.5 w/v%. Magnification: panel (a–e, 

10k); panel (f, 200); panel (g, 100), and panel (h, 50). 



HA/GE compositions and different average diameters 

ranging from 190 to 500 nm could be produced by electrospinning. 

In the present procedure, the overall polymer 

concentration increased with increasing GE content, and 

the higher polymer concentration led to an increase in the 

average diameter of HA-GE nanofibers as well as the decrease 

in conductivity of HA solution, as shown in Table 1. The 

results of surface tension data (Table 1) also showed that 

the addition of GE, acting as a kind of surfactant, decreased 

the surface tension of the solution (samples 5, 7, 8, 9, 10, and 

11 in Table 1) and rendered better electrospinning 

processability. 

Rheological measurements showed that the viscosity 

changed very little after DMF was added into the HA 

solution, as shown in Figure 4(a). Both HA and HA-GE 

solutions in DMF-water had similarly high viscosities, 

which meant that electrospinning of HA solution with 

unusually high viscosity could be proceeded successfully. 

The normal force of spinning solutions, as shown in 

Figure 4(b), could reflect the viscoelastic part of the solutions, 

which was a necessary factor in the spinning process 

a) 

Normal force (N) Viscosity (Pa*s) 

100 

10 

1 

0.1 

1 10 100 1000 

Shear rate (s -1 ) 

b) 

0.1 

0.01 

1E-3 

1E-4 

HA1.3w/v% 


HA in DMF/water=1.5 

HA/GE=100/100 



HA in DMF/water=1.5 

HA/GE=100/100 

10 100 1000 

Shear rate (s -1 ) 

Figure 4. Viscosity (a) and normal force (b) of pure HA and HA- 

GE solutions. 

including dry spinning and electrospinning. The viscoelasticity 

of HA (and HA-GE) in DMF/water was higher than 

that of HA in the water-ethanol solvent mixture. 

Therefore, the decrease in surface tension could play an 

important role in the electrospinning of HA and the formation 

of smooth fibers. In contrast, for the electrospinning of 

HA in water-ethanol solution at a 15 cm distance, the 

relatively high surface tension led to the bead-on-string 

structure, as shown in Figure 1. 

The HA-GE fibrous membranes obtained from electrospinning 

looked like gauzes with ultra-thin fibers at low 

magnifications. This kind of nanostructures could have 

many potential clinical applications, especially in the field 

of artificial skin. Also, HA-GE fibrous membranes at 

different HA/GE compositions obtained by electrospinning 

are expected to have different cell proliferation properties 

and biodegradability, and have potential applications in 

wound dressings, such as hemostatic gauzes, tissue engineering 

scaffolds, and drug delivery. 

Conclusion 

In this study, pure HA solution could be successfully 

electrospun in DMF/water system without the assistance of 

air blowing. The electrospinning processability of HA-GE 

solutions was also investigated. It was found that the 

processability of HA had been improved greatly by using a 

DMF-water solvent mixture or/and by adding GE into the 

HA solution. Nanofibrous membranes with different 

average fiber diameters and different HA/GE compositions 

could be obtained. The average diameters of HA or HAbased 

nanofibrous membranes became larger and the 

conductivity of HA or HA-based solution decreased with 

increasing DMF content in the solvent mixture or increasing 

GE content. Measurements on viscosity indicated that 

the viscosity of HA solution in DMF-water mixed solvent 

did not change much when compared with that in waterethanol. 

HA solution with high viscosity could be electrospun. 

The decrease in surface tension contributed to the 

fiber formation of HA and HA/GE by electrospinning. 

Therefore, the study provided not only a novel and simpler 

way to electrospin the natural polyanion HA solution but 

also the fundamental physical insight and solution to this 

spinning difficulty. The HA-GE nanofibrous membranes 

at different HA/GE compositions are expected to be 

useful in the biomedical field as novel scaffolds for many 

applications. 

Acknowledgements: This work was financially supported by 

the National Nature Science Foundation of China (Funds No. 

50503023 and 50373048), the National ‘‘973’’ Project (G2003 

CB615605), the MSC Creative Project of CAS (CMS-CX200503), 

the National Institutes of Health SBIR grant (GM63283-02) in the 

U.S, administered by the Stonybrook Technology and Applied 

Research, Inc. 



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Electrospinning of Hyaluronic Acid (HA) and HA/Gelatin Blends

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