ARTICLE IN PRESS
Biomaterials 24 (2003) 3915–3919
A novel solvent system for blending of polyurethane and heparin
Qiang Lv, Chuanbao Cao*, Hesun Zhu
Research Center of Material Science, Beijing Institute of Technology, Beijing 100081, China
Received 17 November 2002; accepted 8 April 2003
Abstract
To improve the blood-compatibility of polyurethane, the co-solvent of tetrahydrofuran and water, a new solvent system for
blending polyurethane and heparin, was proposed. After solvent casting, heparin was blended in a polyurethane film. The ATRFTIR was used to analyze the surface chemical element and the contact angle was measured to investigate the hydrophilicity of the
surface of the PUs. As the amount of heparin increased, the surface hydrophilicity was increased and all the clot times exceeded the
measurement limit of the clot detection instrument when the heparin loaded on the polyurethane films was 3%, 5% and 7%. After
the films were immersed in the phosphate buffered saline for 30 days, the activated partial thromboplastin time and thrombin time
still exceeded the measurement limit of the clot detection instrument.
r 2003 Elsevier Science Ltd. All rights reserved.
Keywords: Blood compatibility; Heparin; Blending; Polyurethane
1. Introduction
Segmented polyurethanes have been extensively used
for the construction of cardiovascular devices because of
their desirable physical properties and relative thromboresistance compared to other materials used in
cardiovascular application [1,2], but thrombus formation still exists in small diameter tubing. To enhance the
blood compatibility of polyurethanes, many different
approaches have been studied.
Heparin is an important anticoagulant, used clinically
to minimize thrombus formation on artificial surfaces
[3]. It has the ability to interact strongly with
antithrombin III to prevent the formation of fibrin clot
[4,5]. There are two general methods to develop bloodcompatible polymeric materials using heparin. One
method uses chemical immobilization of heparin and
the other is the heparin-releasing system.
Covalent immobilization of heparin onto a polymer
surface can provide long lasting antithrombogenicity.
Heparin can be directly bound to the surface by
insertion of either functional groups [6–8] or spacer
arms [9], but the activity of heparin was significantly
*Corresponding author. Tel.: +86-68913469; fax: +86-68915023.
E-mail address: cbcao@bit.edu.cn (C. Cao).
decreased compared to raw heparin. In a heparinreleasing system, heparin is slowly released for a short
duration and the bioactivity of heparin is maintained at
a high level. Therefore, a heparin-releasing system is
more suitable for a short-term clinical application than
the covalently coated heparin system. There are several
methods studied for the formulation of a heparinreleasing system [10,11]. In those methods, most
releasing systems are useless because heparin was
released within several hours, so a novel formulation
for controlled release of heparin is necessary. Hyum Tae
Moon et al. [12] reported that the heparin-DOCA,
having an amphiphilic property, was homogeneously
mixed with polyurethane in the co-solvent of dioxane,
propanol and water. After casting the film, heparinDOCA was homogeneously dispersed as nanoparticles
in polyurethane films. However, the losing of the activity
of heparin was inevitable and the system could not be
used in covalent immobilization of heparin because
heparin has grafted with DOCA.
In this study, we utilize polyurethane and the new
solvent system to prepare a heparin-releasing system by
simple solvent casting. It was found that heparin
homogeneously mixed with polyurethane in the cosolvent of tetrahydrofuran and water if the ratio was
suitable. Then heparin was blended in the polyurethane
0142-9612/03/$ - see front matter r 2003 Elsevier Science Ltd. All rights reserved.
doi:10.1016/S0142-9612(03)00266-7
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Q. Lv et al. / Biomaterials 24 (2003) 3915–3919
films by solvent casting. The release rate, surface
property and antithrombogenicity were studied.
2. Experiment
2.1. Materials
The polyurethane was presented from the polymer
laboratory of our university. The synthesis method
employed here was a two-step method, in which the first
step was to use poly(ethylene oxide-tetramethylene
oxide) provided by LiMing chemical research institute
and isophorone diisocyanate (IPDI) to synthesize a
prepolymer, and the second step was to use 1,4butanediol as an extender to react with the prepolymer
to produce the final product. The average molecular
weight of poly(ethylene oxide-tetramethylene oxide) was
4850 and the ratio of ethylene oxide/tetramethylene
oxide was 50/50. As a result, the average molecular
weight of PU determined by gel permeation chromatography (Wasters 150-C, 1 ml/min, 30 C) was about
64,000. The mechanical properties were measured on the
Instron-6022 Test machine at 20 C and 65% RH and
the head speed was 10 mm/min. The tensile strength of
PU was about 8.34 MPa and ultimate elongation was
636%.
2.2. Preparation of heparin blended polyurethane film
Polyurethane (1.3 g) was dissolved in 15 ml tetrahydrofuran, and then a 2 ml different concentration
heparin water solution was mixed with it. The weight
ratio of heparin was 3%, 5% and 7% compared with
polyurethane, respectively. The mixing solution was
spread on a glass plate (6 6 cm2) and the solvent was
evaporated at room temperature. The films were dried in
a drying oven at 60 C for 24 h for testing.
2.4. Release test of heparin from polyurethane film
The amount of heparin released from the film was
determined using the toluidine blue method as reported
in the literature [8,13,14]. For keeping the thickness of
the films identical to each other, all films tested were
made at the same experimental conditions. A known
amount of heparin aqueous solution (2 ml) was added to
the toluidine blue solution (3 ml) and the mixed solution
was adequately vibrated. Hexane (3 ml) was then added,
and the mixture was well shaken so that the toluidine
blue–heparin complex was extracted into the organic
layer. The toluidine blue that remained in the aqueous
phase was determined by measuring the absorbance at
631 nm and the concentration of heparin in the aqueous
solution was obtained and used as a calibration curve to
determine the amount of released heparin.
To study the release rate of heparin, the polyurethane
film (3 cm 4 cm) containing heparin was immersed in
PBS solution (12 ml, pH 7.4) for 24 h. Periodically, the
supernatant (2 ml) of the PBS solution was replaced with
a fresh one in order to maintain a sink condition. And
then the absorbance at 631 nm of the supernatant was
measured and the released amount of heparin was
calculated according to the standard curve. The
standard curve (Fig. 3) was obtained according to the
method of Smith et al. The standard curve was linear in
the range of 0–40 mg heparin/ml and the heparin content
was obtained directly from the standard curve.
2.5. In vitro coagulation time tests for films
The nephelometry measurements, including PT,
activated partial thromboplastin time (APTT) and
thrombin time (TT), were performed with the coagulation instrument Coag-A-Mate XM (Organon Teknika,
USA) which measures the change of luminosity when
a
2.3. Characterizations of heparin blended polyurethane
film
The surfaces of PUs were investigated using attenuated total reflectance infrared spectroscope (ATRFTIR, SYSTEM-2000, PE, USA). After the film was
immersed in phosphate buffered saline (PBS) for 1 h, the
surface morphologies were studied to compare with the
initial morphologies by scanning electron microscope
(SEM, JSM-35C, JEOL, Japan).
The water contact angles of the film surfaces were
measured by putting a droplet of deionized water on the
surface of the polymer films using the JY-82 contact
angle apparatus (ChengDe, China). With each specimen, the measurement was repeated at different sites,
and average values were obtained for the contact angles.
b
c
d
e
3500
3000
2500
2000
1500
1000
Wavenumber cm-1
Fig. 1. ATR-FTIR spectra of: (a) PU, (b) PU-3 wt%He, (c) PU5 wt%He, (d) PU-7 wt%He and (e) FTIR spectra of heparin.
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Q. Lv et al. / Biomaterials 24 (2003) 3915–3919
light traverses the plasma sample. Briefly, the tested
films were incubated with healthy human blood plasma
in a transparent plastic tube, and the reagents for each
coagulation time test were added to the tube immediately.
3. Results and discussion
3.1. Surface infrared analysis
Fig. 1 showed ATR-FTIR spectra of PU-0%He(a),
PU-3%He(b), PU-5%He(c), PU-7%He(d) and the
Table 1
Water contact angles of different polyurethane films
Heparin rate
Contact angle ( )
3% Heparin
5% Heparin
7% Heparin
7272
6871
5473
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FTIR spectra of heparin as typical examples. In the
spectrum of PU(b)–(d), the peak in 2944 cm 1 increased
and the peak in 1460 cm 1 moved to low wave number
which may be due to the effect of heparin. On the other
hand, the new peak in 891 cm 1 which exists in the
spectra of the heparin was also found in the spectra of
(b)–(d) and the peak in 2358 cm 1, though it had no
absorption in heparin, represented the stretching vibration of sulfonamide (–NHSO3) reported by some
researchers [15,16] and the result may mean there are
some interaction between polyurethane and heparin
such as the hydrogen bond. At the same time, the point
that the area of peaks in 2358 cm 1 augment when the
ratio of heparin increases partly suggests the surface
content of heparin increases.
3.2. Water contact angle
To investigate the hydrophilicity of the PUs, water
contact angle measurements were carried out (Table 1).
The PUs blended with heparin were relatively
(a)
10 µm
10 µm
10
10 m
µm
µm
1010 m
10 µm
10 µm
10 µm
10 µm
(b)
(c)
(d)
Fig. 2. SEM of surface morphologies of heparin loaded polyurethane films before and after immersing in PBS for 1 h: (a) 0 wt%heparin,
(b) 3 wt%heparin, (c) 5 wt%heparin, (d) 7 wt%heparin.
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hydrophilic; PUs 1–3 exhibited contact angles of 7272,
6871 and 5473, respectively. These results agree with
the increase of heparin content in the polyurethane.
3.3. Release of heparin from polyurethane films
As shown in Fig. 2, when the heparin-loaded film was
soaked in PBS for 1 h, there were many ridges at the
surface when heparin was released out of the films, but
the size and density of the ridges were not increased with
the increasing amount of loaded heparin. The density of
the ridges was most crowded for the 5 wt% heparin
loading films and most sparse for the 7 wt% heparin
loading one. Furthermore, the SEM images have the
change accordant with the percentage of released
amount of heparin shown in Fig. 4.
For 3 wt%, 5 wt% and 7 wt% heparin loading films
(Fig. 4), the percentages of released amounts of heparin
for 24 h were 6.13%, 7.19% and 3.61%, respectively.
This may be due to the difference in the interlaced
structure of the different content of heparin and
polyurethane. For the heparin-loaded films, heparin
was quickly released within the first one hour, followed
Absorbance at 631nm
2.5
2.0
1.5
1.0
0.5
0.00
0.05
0.10
0.15
by the relative sustained release rate for one day. This
might be because of the fact that heparin located near
the surface would have had a short distance to travel to
the film surface. Although the relation of heparin
content and release rate needed further research, the
blended amount of heparin was an important parameter
in controlling the release rate.
3.4. In vitro blood compatibility
The PT, APTT and TT were widely used for the
clinical detection of the abnormality of blood plasma. In
recent times, they were applied in the evaluation of
in vitro antithrombogenicity of biomaterials. The
normal ranges of PT, APTT and TT for a healthy
blood plasma were regarded as 1173, 2874 and
1675 s, respectively. The results are given in Table 2.
The three times all exceeded the measurement limit of
the clot-detection instrument indicated that heparin
blended with polyurethane kept excellent blood-compatibility and the plasma was not coagulated with
polyurethane since the released heparin was enough to
prevent the formation of fibrin clot.
Table 3 shows the APTT, PT and TT of the films
containing 5% heparin, which was immersed in PBS for
one month. After 30 days, although PT had decreased to
12 s, the APTT and TT still exceeded the measurement
limit of the clot-detection instrument. Since we have
studied that polyurethane containing heparin reduced
the PT time to 15 s when it was located in the desiccator
for five days at 20 C, the reduction of PT may be partly
due to the loss of heparin bioactivity as the films still
preserved enough heparin to keep the films’ bloodcompatibility within about 30 days.
0.20
Table 2
Activated partial thromboplastin time (APTT), thrombin time (TT)
and prothrombin time (PT) of Pu films containing heparin
Heparin amount(mg)
Fig. 3. Grades curve of heparin amount.
Heparin relative release percent (%)
Heparin rate
Clot time (s)
8
7
b
6
a
0%
3%
5%
7%
5
Heparin
Heparin
Heparin
Heparin
TT
APTT
PT
15.1
>150
>150
>150
46.8
>200
>200
>200
25.0
>150
>150
>150
4
c
3
Table 3
Clot times of Pu films containing 5 wt% heparin immersed in PBS for
different number of days
2
1
Immerse time
Clot time (min)
0
0
5
10
15
20
25
Time (h)
Fig. 4. Release profiles of heparin from polyurethane films: (a)
3 wt%heparin, (b) 5 wt%heparin, (c) 7 wt%heparin.
Five days
Thirty days
TT
APTT
PT
>150
>150
>200
>200
11.8
12.4
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Q. Lv et al. / Biomaterials 24 (2003) 3915–3919
4. Conclusion
In this study, the appropriate solvent was used to
dissolve heparin and polyurethane. The simple solvent
casting method allows heparin to be released slowly
from the polymeric film and the rate of drug release can
be controlled by the amount of drug being loaded. In the
films, heparin can keep excellent bioactivity and in a
relatively long time the films shows favorable blood
compatibility. So we think that it could be especially
useful when applied to devices designed for mediumtime use.
Acknowledgements
This study is supported by 973 project (G1999064705)
of China and 863 project (2002AA326030) of China.
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