TECHNICAL FIELD
The present invention generally relates to the use of a solvent free, single step
process for generation of three-dimensional porous scaffolds. The invention
more particularly relates to the generation of three-dimensional porous
scaffolds through an in-place porosity generation process during ring opening
polymerization involving homo-polymerization or cross-linking polymerization
techniques. The porous scaffolds thus produced can be used as supports or
culture matrices for in-vitro tissue cultures e.g. as biomedical materials.
BACKGROUND OF THE INVENTION
The technique of particulate leaching to generate three dimensional porous
scaffolds is well known; however it is a two step process in which a preformed
polymer is first dissolved in a solvent having dispersed particulates followed by
leaching of particulates in a media in which particulate is soluble while the
polymer is not. In addition the technique cannot be applied to a cross-linked
polymer as a cross-linked polymer doesn't dissolve in any solvent (it only
swells).
Discussed in this section are some of the attempts in the instant state of art
where various articles and research papers attempts to build a three
dimensional porous scaffolds. Japanese patent 2007-187558, describes a
method which involves the use of a preformed polymer and the porosity is
generated via phase separation mechanism and has prepared scaffold material
for living bodies, biocompatible water-soluble polymer fibers are mixed and
dispersed into a water-insoluble biocompatible material.
Similarly another Japanese Patent 2011-002204, discloses the preparation of
porous scaffold made of a biodegradable polymer such as a polylactide, and a
method for homogeneously seeding cells inside the porous scaffold. The
process described in this patent uses an organic solvent (dioxane) and is a
multistep process involving inclusion of a preformed polymer.
Other Japanese Patent 2006-010335, provides a process of preparation of a
multilayer porous structure comprising a biodegradable polymer capable of
forming a multilayer porous structure. The said method comprises mixing a
biodegradable polymer with a material incompatible with the biodegradable
polymer to prepare a coating fluid, forming the prepared coating fluid into a
coating film with a phase-separated structure in which the biodegradable
polymer and the incompatible material are phase-separated, and extracting the
incompatible material from the coating film with the phase-separated structure.
The process involves preformed polymer and mixing of this polymer with
incompatible substance to form a coating over it.
Another Japanese Patent 2004-110328, describes a process for generating an
implantable, biocompatible scaffold that has a biocompatible, porous
polymeric matrix, a biocompatible, porous fibrous mat encapsulated by and
disposed within the polymeric matrix. A preformed polymer is used for this
purpose.
The subject matter of Japanese Patent 2003-032045, pertains to a porous
scaffold of high porosity, pore size and high strength, to provide an implantable
medical device, especially an orthopedic implant. The process involves the
formation of unsintered metal foam from the polymer and metal skin which is
then coated with bond solution followed by sintering. This is altogether a
different approach than in-situ porosity generation during polymerization or
cross-linking.
Another International Patent application WO2006091921, describe about the
micro-carrier bead having a porous three-dimensional core having at least 99%
interconnected pores and an outer protective layer for growing cells in a
bioreactor using higher sparging rates than cells would ordinary withstand. The
method involves making of an artificial scaffold wherein a scaffolding material
is extruded into a coolant and thereby creating a porous material.
Another International Patent Application WO 2006082270 teaches the
preparation of the three dimensional porous polymer scaffolds for tissue
regeneration, having a porous structure which is the inverse or negative of a
three-dimensional porogen or template resulting from the sintering of stacked
meshes of nylon fibers. The polymer is obtained by infiltration and
polymerization of the cross-linking material in the template, elimination of the
porogen by means of leaching, and drying of the resulting porous matrix. Free
radical polymerization was used in this method and vinyl monomers are used
in this case where the porosity is coming from a 3-D template based on Nylon.
European Patent EP 1664168 claims the use of hydrogel micro particles with
entrapped liquid as the porogen. In the process, a biodegradable unsaturated
polymer, a cross-linking agent, and a porogen comprising biodegradable
hydrogel microparticles are mixed together and allowed to form a porous
scaffold in a mold or in a body cavity. The cross-linking agent used here is a
free radical initiator, or may include a free radical initiator and a monomer
capable of addition polymerization.
International Patent Application WO 2002060508, discloses the method for the
preparation of a porous body of a biodegradable material comprising the steps
of mixing a copolymer of polyalkylene glycol terepthalate and an aromatic
polyester with particles that are soluble in a solvent in which the copolymer
essentially does not dissolve, and subjecting the obtained mixture to heat
and/or pressure sufficiently long for forming the body. The process involved
use of a preformed polymer and also an organic solvent.
The Subject matter of US patent 7943678 pertains to the production of porous
foam from polyurethane. The method is based on salt leaching out and phase
separation techniques. First macrodiols react with macro diisocyanate for
producing polyurethane foam. At the next step salt leaching technique was
followed for porosity generation.
Another US patent 7846466, teaches a process in which scaffolds containing
porous polymer material are prepared by a process of gas foaming/particulate
leaching and a wet granulation step prior to gas foaming and particulate
leaching. A preformed polymer is used in such process and porosity is mainly
generated via gas foaming.
Other US patent 7842305 claims a process of preparing biodegradable dual
pore polymer scaffolds which was prepared from previously synthesized
biodegradable polymer. They had dissolved the polymer in an effervescent
mixture of carbonate and organic acid and solvent. Then scaffold was produced
by salt leaching and phase separation technique. This invention also was
subjected to preformed polymer.
Yet another US patent US 6586246 describes a multi-step process of threedimensional
scaffold synthesis of biodegradable polymer. At first an
effervescent salt was mixed with polymer-solvent system for obtaining certain
gel point. Subsequently solvent was removed from mixture. Then gel was
treated with acid to cause the salt to effervesce at room temperature, thus threedimensional
porous structure was produced.
Another US patent US 6562374 claims a three step process for porous
biodegradable scaffold by subsequent solubilizing, effervescing and drying
techniques. It produced an interconnected network structure.
The subject matter of Korean Patent application KR20040101787 pertains to a
method for preparing biodegradable porous polymer support for tissue
engineering without organic solvent. A preformed polymer was used in this
case which was melt pressed along with a water soluble salt to form a
composite construct. The salt was then leached out from the construct after
washing with water. The process presented in this patent described use of a
preformed polymer and the same process was not shown to be useful for crosslinked
systems. Thus the porosity generated here is not an in-situ method of
porosity generation during polymerization.
Yet Another US application US2007178159, teaches about the formation of
porous scaffold by making use of the viscous gel formed from a combination of
a biodegradable polymer and a biocompatible solvent that also includes a
hydrophilic porogen. The said process is useful for only preformed polymer
and involves use of a solvent.
These listed above prior arts describes porous structures prepared by different
process involving solvent/non-solvent induced phase separation method, gas
foaming, leaching, coagulation method etc. but the existing processes and
structures obtained from those methods are still associated with the following
limitations:-
• Use of organic solvents
• Lengthy, time consuming and round about process
• Multi step process
• Not suitable for cross-Linked polymers
• No in-situ porosity generation
Hence there is requirement of new methodology to overcome above limitations
and applicant has developed a new method of preparing porous scaffolds via
in-situ porosity generation during polymerization suitable for both homopolymers
and cross-linked polymers without use of organic solvents.
OBJECTS OF THE INVENTION
The primary object of the present invention is to provide a single step process
for generation of three-dimensional porous scaffolds via in-situ porosity
generation during polymerization involving homo-polymerization or
cross-linking.
Another object of the present invention is to provide a simple method for the
production of three-dimensional porous scaffolds without the use of the organic
solvents.
Another object of the present invention is to provide a three-dimensional
porous scaffold where mode of polymerization or cross-linking developed is
based on ring-opening polymerization.
It is yet another object of the present invention to use lactones, lactides or
cyclic carbonates as monomers and organometallic compounds as catalysts for
polymerization.
It is yet another object of the present invention to provide an efficient process
for the preparation of three-dimensional porous scaffolds suitable for both
homo-polymers and cross-linked polymers.
These and other objects and advantages of the present invention will become
readily apparent from the following detailed description taken in conjunction
with the accompanying drawings.
SUMMARY OF THE INVENTION
The various embodiments of the present invention provide a single step, solvent
free process of preparing a three-dimensional porous scaffold through a ringopening
polymerization process, the said process comprising the steps of
preparing a homogeneous mixture of monomer (e.g. CL) and catalyst (e.g.
Sn(0ct)2) of known quantity in a mould of desired dimensions and adding a
known quantity of pre-dried and pre-heated porogen (e.g. NaCl crystals) in the
mould and heating the mould to a temperature of 160 ± 20°C for 2.0 ± 0.5 hours
in an oven without any inert gas atmosphere. The mould is then taken out from the
oven and cooled to room temperature and then finally porogen is leached out in
water.
According to an embodiment of the present invention, a cross-linking agent (e.g.
bis(s-caprolactone-4yl) - BCY) is added along with monomer (e.g. CL) and
catalyst (e.g. Sn(0ct)2) in the mould with the similar process as described above to
obtain the cross-linked porous three-dimensional porous scaffolds .
According to an embodiment of the present invention the molar ratio of monomer
and cross-linking agent is taken in a range using equation (1) to vary the crosslink
density (p) of the final three-dimensional porous scaffold synthesized.
P = ^ ^ 1 0 0 Equation (1)
Where n is the mole fraction of cross-linking agent and m is the mole fraction of
monomer.
According to an embodiment of the present invention, porosity is mainly
generated in-situ, by the addition of porogen within the process during
polymerization.
According to an embodiment of the present invention, the polymerization
process does not require the use of organic solvents for producing threedimensional
porous scaffolds.
According to an embodiment of the present invention monomers or catalyst
having 93-100% purity are selected to suit the applicability of the process at
commercial level.
These and other aspects of the embodiments herein will be better appreciated
and understood when considered in conjunction with the following description
and the accompanying drawings. It should be understood, however that the
following description while indicating preferred embodiments and numerous
specific details thereof are given by way of illustration and not of limitation.
Many changes and modifications may be made within the scope of the
embodiments herein without departing from the spirit thereof, and the
embodiments herein include all such modifications.
BRIEF DESCRIPTION OF THE DRAWINGS
The other objects, features and advantages will occur to those skilled in the art
from the following description of the preferred embodiment and the
accompanying drawings
FIG.l is a representative 'H-NMR spectrum depicting complete monomer (ecaprolactone)
conversion during synthesis of uncross-linked three-dimensional
porous scaffolds.
FIG.2 is a representative DSC thermogram confirming formation of PCL
during synthesis of uncross-linked three-dimension porous scaffolds.
FIG.3 is a representative SEM image depicting porous surface of uncrosslinked
three-dimensional porous scaffold.
FIG.4 is a Representative SEM image depicting porous cross-section of
uncross-linked three-dimensional porous scaffold.
FIG.5 is representative swelling behavior characteristics of cross-linked porous
scaffolds based on PCL in chloroform (1-3) and water (4-6).
FIG.6 is a representative FTIR spectrum of cross-linked three-dimensional
porous scaffold based on PCL; characteristic peaks due to functional groups
present in the cross-linked polymer are: 2941 cm"'(asymmetric CHi stretching),
1729 cm"'(carbonyl stretching), 1357 cm"'(C-0 and C-C stretching in the
crystalline phase), 1235 cm"'(asymmetric C-O-C stretching ) and 1164 cm"
'(asymmetric C-O-C stretching).
FIG.7 is a representative SEM image depicting porous surface of cross-linked
three-dimensional porous scaffold.
FIG.8 is a representative SEM image depicting porous cross-section of crosslinked
three-dimensional porous scaffold.
Although specific features of the present invention are shown in some drawings
and not in others. This is done for convenience only as each feature may be
combined with any or all of the other features in accordance with the present
invention.
DETAILED DESCRIPTION OF THE INVENTION
The invention will now be described in detail with reference to a few preferred
embodiments, as illustrated in accompanying drawings. In the following
description, numerous specific details are set forth in order to provide a
thorough understanding of the invention. However, it will be apparent to one
skilled in the art that the invention may be practiced without some or all of
these specific details. In other instances, well-known features and/or process
steps have not been described in detail in order to not unnecessarily obscure the
invention. The features and advantages of the invention may be better
understood with reference to the drawings and discussions that follow.
10
The various embodiments of the present invention provide a process of
preparing a three-dimensional porous scaffold through a ring-opening
polymerization, the method comprising the steps of preparing a homogenous
mixture comprising of monomer, e-caprolactone (CL) of at least 0.5 to 10
grams and catalyst, Sn(0ct)2 of at least 0.01 to 0.1 mol% of CL in a pre dried
mould, and adding a known quantity of a pre-meshed and a pre-heated porogen
(NaCl crystals) which is at least 5 to 10 wt% of CL to the mould containing the
homogeneous mixture wherein the mould than immediately placed inside an
oven with a preset temperature of at least 160 ± 20°C for at least 2.0 ± 0.5 hour
and a porous film so formed, is cooled to the room temperature in a desiccator
and removed and then the porogen is leached out from the film in a water bath
for at least 48 hours with several rounds of changing the water and the film
obtained finally is dried under vacuum and stored for characterization.
In the present invention, the preparation of three-dimensional porous scaffolds
for tissue culture is based on ring opening polymerization.
In the preferred embodiment of the present invention, Uncross-linked threedimensional
porous scaffolds are synthesized without the use of any crosslinking
agent using the same process as described above.
In other preferred embodiment, cross-linked three-dimensional porous
scaffolds are synthesized with the use of any cross-linking agent where a crosslinking
agent (bis(e-caprolactone-4yl) - BCY) is mixed along with monomer
and catalyst in the mould and a porogen is added in a mould and the similar
process as described above is followed.
Previously existing methodologies provide technique of particulate leaching to
generate three dimensional porous scaffolds which is a well known art.
However it is a two step process in which a preformed polymer is first
11
dissolved in a solvent having dispersed particulates followed by leaching of
particulates in a media in which particulate is soluble while the polymer is not.
In addition the technique cannot be applied to a cross-linked polymer as a
cross-linked polymer doesn't dissolve in any solvent (it only swells).
The present invention provides a single step process for generation of threedimensional
porous scaffolds via in-situ porosity generation during
polymerization involving homo-polymerization or cross-linking.
According to the embodiment, the process of obtaining single step process for
generation of three-dimensional porous scaffolds comprises a homogeneous
mixture of monomer (e.g. CL), catalyst (e.g. Sn(0ct)2) and cross-linking agent
(e.g. bis(s-caprolactone-4yl) - BCY) prepared in a mould of desired
dimensions. The molar ratio of monomer and cross-linking agent is taken in a
range using equation (1) to vary the cross-link density ip) of the final threedimensional
porous scaffold synthesized.
^" X 100 Equation (1)
'^ (2n+7n)
Where n is the mole fraction of cross-linking agent and m is the mole fraction
of monomer. To the mould containing mixture of monomer, catalyst and crosslinking
agent, a known quantity of pre-dried porogen (e.g. NaCl crystals) is
added and the mould is heated to a temperature of 160 ± 20°C for 2.0 ± 0.5
hours in an oven without any inert gas atmosphere. The mould is then taken out
from the oven, cooled to room temperature and then porogen is leached out in
water.
The porous film obtained is finally dried under vacuum and stored for
characterization. The film is analyzed by various characterization techniques
such as the extent of polymerization, degree of cross-linking and morphology
of three-dimensional porous scaffolds is determined using spectroscopy,
calorimetric analysis, gravimetric analysis and microscopy.
12
The three-dimensional scaffolds developed are comprised of aliphatic
polyesters or aliphatic carbonates using lactones, lactides or cyclic carbonates
as monomers and organometallic compounds as catalysts for polymerization.
The purity of the monomers or catalyst is selected over a wide range to suit the
applicability of the process at commercial level. Preferably, monomers or
catalyst having purity of 93- 100% is used in the present invention.
In one aspect of the present invention, the process as being described above is a
solvent free process and no inert atmosphere is required for synthesizing the
materials and the said process has the potential to convert a batch process of
generating a three dimensional porous scaffold via particulate leaching
technique into a continuous process.
In an embodiment of the present invention, monomer used in the process is
preferably in liquid form (at room temperature or at temperature of
polymerization) and the process does not involve the use of an organic solvent.
Monomer is preferably selected from the group consisting of Lactones (4 to 16
C cyclic esters), Lactides (L-, D- and D,L- Lactide) and Cyclic Carbonates
(Trimethylene carbonate {TMC} and substituted TMC).
Catalyst is preferably selected from the group consisting of Sn(0ct)2 (Stannous
Octoate), Al(iPro)3 (Aluminum Isopropoxide) or their substituted derivatives.
In one aspect of the Invention, the porogen is selected such that it imparts
porosity to the porous scaffold in situ. Porogen is preferably selected from the
group consisting of any thermally stable (up to 180°C) common lab salt which
is not soluble in CL but soluble in water e.g. NaCl, Na2S04, MgS04, NaiCOs
etc. The porogen used in the present invention is hygroscopic, preferably in a
crystal form. The presence of hygroscopic compound such as NaCl (used as
13
porogen) doesn't affect the polymerization and the porogen (salt) is leached out
from the construct after washing with water. Thus the porosity generated here
is an in-situ method of porosity generation during polymerization.
CHARACTERIZATION STUDIES
The various characteristics of three dimensional porous scaffolds such as the
extent of polymerization, degree of cross-linking and morphology are analyzed
and determined using characterization techniques as described below:
Nuclear Magnetic Resonance (NMR) Spectroscopy
The monomer conversion and degree of polymerization was determined by 'H
NMR. A Bruker-300 NMR instrument operating at 300 MHz was used for this
purpose. CDCI3 was used as solvent as well as internal standard (5 = 7.26 ppm).
As shown in FlG.l, a representative 'H-NMR spectrum depicts complete
monomer (e-caprolactone) conversion during synthesis of uncross-linked threedimensional
porous scaffolds according to an embodiment of the present
invention. According to the embodiment, ' H-NMR was used to determine the
monomer conversion and degree of polymerization (DP) of the PCL
homopolymer (uncross-linked) based porous scaffold formed. The spectrum is
presented in figure 1. No peak due to unreacted CL monomer was observed
implying that all the monomer is converted into polymer and monomer
conversion is ~ 100%. The DP was calculated considering the intensities of
oxy-methylene protons (-CH2-O-) at -4.0 ppm to the intensity of terminal oxymethylene
protons (-CH2-OH) at -3.6 ppm.
Differential Scanning Calorimetry (DSC)
The thermal properties of the polymers were determined by DSC using a
Perkin Elmer Instrument under a nitrogen atmosphere with a sample mass of 5
14
± 1 mg and a heating rate of 10°C/min. The relative crystallinity of different
samples of PCL was calculated according to the equation 2
Wc _— -^—^f^ xlOO Equation (2)
where Wc is the crystallinity, AHf is the heat of fusion of the sample, and AHf
is the heat of fusion of 100% crystalline PCL. The value of AH f used for the
calculations was 139.5 J/g.
FIG.2 shows a representative DSC thermogram confirming formation of PCL
during synthesis of uncross-linked three-dimension porous scaffolds according
to an embodiment of the present invention. According to the embodiment, DSC
thermogram of a representative porous scaffold based on PCL homopolymer
(uncross-linked) showed a melting peak at ~60°C suggesting that PCL
homopolymer has formed under chosen set of polymerization conditions. The
relative crystallinity (Wc) of the homopolymer was calculated according to
equation (2).
Thermogravimetric Analysis (TGA)
Thermal stability was evaluated by TGA using a Perkin Elmer
thermogravimetric analyzer under a nitrogen atmosphere with a sample mass
of 5 ± 1 mg and a heating rate of 10°C /min.
Fourier Transform Infra-red Spectroscopy (FTIR)
FTIR measurements were performed on a Perkin-Elmer spectrometer,
equipped with a golden gate single reflection ATR unit with a diamond crystal.
-1
The spectra were taken as an average of 20 scans at a resolution of 4 cm .
F1G.6 shows the FTIR spectrum of cross-linked three-dimensional porous
scaffold based on PCL; characteristic peaks due to functional groups present in
the cross-linked polymer are: 2941 cm"' (asymmetric CH2 stretching), 1729 cm"
' (carbonyl stretching), 1357 cm"' (C-0 and C-C stretching in the crystalline
15
phase), 1235 cm" (asymmetric C-O-C stretching) and 1164 cm" (asymmetric
C-O-C stretching)according to an embodiment of the present invention.
According to the embodiment, the figure shows that the porous cross-linked
scaffold formed is based on PCL. As cross-linked PCL could not be dissolved
in a solvent to record its IH-NMR, this technique (FTIR) was used to show
that the polymer formed here is PCL.
Scanning Electron Microscopy (SEM)
SEM images were acquired using a Zeiss Evo 50 SEM (Scanning Electron
Microscope) & ImageJ software was used for measuring the pore size
variations in porous films.
As shown in FIG.3, a SEM image depicts the porous surface of uncross-linked
three-dimensional porous scaffold according to an embodiment of the present
invention. According to the embodiment, the scanning electron microscopy
image shows the surface of a representative porous scaffold based on PCL
homopolymer (uncross-linked). The pores formed on the surface clearly
showed the porous nature of the scaffold.
As seen in FIG.4, a SEM image depicts porous cross-section of uncross-linked
three-dimensional porous scaffold according to an embodiment of the present
invention. According to the embodiment, the scanning electron microscopy
image shows the cross-section of a representative porous scaffold based on
PCL homopolymer (uncross-linked). The uniformity and distribution of the
pores clearly shows that pores were also formed inside the scaffold and they
were uniformly present in the scaffold.
FIG.7 shows the SEM image depicting porous surface of cross-linked threedimensional
porous scaffold according to an embodiment of the present
invention. According to the embodiment, the scanning electron microscopy
image shows the surface of a representative porous scaffold based on cross-
16
linked PCL. The pores formed on the surface clearly showed the porous nature
of the scaffold.
FIG. 8 is a SEM image which shows the porous cross-section of cross-linked
three-dimensional porous scaffold according to an embodiment of the present
invention. According to the embodiment, the scanning electron microscopy
image shows the cross-section of a representative porous scaffold based on
cross-linked PCL. The uniformity and distribution of the pores clearly shows
that pores were also formed inside the scaffold and they were uniformly present
in the scaffold.
Degree of swelling (DS)
The degree of swelling of the Cross-linked films was determined
gravimetrically. A piece of the film was weighed and kept in a sealed flask
containing chloroform (or water). At regular intervals, the film was taken out
and the excess solvent was removed from the surface with the help of a tissue
paper. The film was then weighed and returned to the medium. This procedure
was continued until a constant weight was attained. The equilibrium degree of
swelling (DS) was calculated according to equation (3).
DS= ^^^^^xlOO Equation (3)
Wo
where ^ is the initial weight of the dry sample and ^ i s the final weight of the
swollen sample.
Swelling behavior of cross-linked porous scaffolds is shown in FIG.5, based on
PCL in chloroform (1-3) and water (4-6) according to an embodiment of the
present invention. According to the embodiment, the degree of swelling was
calculated for porous cross-linked scaffolds of PCL according to the procedure
described under experimental section using equation 3. The degree of swelling
was measured in two different media i.e. chloroform and water. This figure
17
shows actual pictures of swollen porous cross-linked scaffolds of PCL in
chloroform (figure 5: 1-3) and water (figure 5: 4-6). The value of degree of
swelling for scaffolds, as expected, is found to be higher when chloroform was
used as the medium.
Gel Content (GC)
The gel content of the cross-linked films was determined gravimetrically. A
piece of the film was weighed and kept in a sealed flask containing chloroform
for 48 hours with intermittent shaking of the flask. The films was taken out
from the flask and left for drying under air for 48 hours before vacuum drying
for 24 hours. The weight of the dried film (W) was recorded and it was
subtracted from the weight of the original film (Wo) to determine the weight of
the soluble portion (Ws) of the sample according to equation 4.
Ws = WQ-W Equation(4)
The gel content is calculated according to equation 5
GC = ^\^'^ X 100 Equation (5)
where Wo is the initial weight of the sample and Ws is the weight of soluble
fraction of the sample.
The Table-1 as shown below provides the result of reproducibility of a crosslinked
three-dimensional porous scaffold synthesized using CL (monomer),
Sn(0ct)2 (catalyst) and BCY (cross-linker) at constant cross-link density and
extent of porosity.
18
Table-1:
s
•
N
o
•
1
2
3
4
Samp
lelD
PI
P2
P3
P4
CL
(mole
s)
0.056
0.056
0.056
0.056
Sn(Oc
t)2
(moles
)
0.0002
0.0002
0.0002
0.0002
BCY
(mole
s)
0.001
5
0.001
5
0.001
5
0.001
5
Theoreti
calcrosslink
density
(%)
5
5
5
5
NaCl
(wt %
wrt
CL)
5
5
5
5
Degre
eof
swelli
ng
(%) in
CHCI3
1642
1589
1404
1468
Gel
Conte
nt (%)
91.0
93.5
89.9
90.8
Avera
ge
pore
size
(Hm)
37.24
35.96
34.54
35.45
The above table summarize the results obtained for degree of swelling (in
chloroform), gel content and average pore size (obtained from SEM) for FOUR
porous cross-linked scaffolds synthesized using identical feed formulation. The
theoretical cross-linked density is calculated from equation (1). The value of
degree of swelling calculated from equation (3) for all the four samples is
found to be close to each other (with an average value of 1525 % and standard
deviation of 109%).This shows that almost equal cross-linking has happened in
all the four samples under the polymerization conditions chosen in our
experiments. The gel content calculated from equation (5) was also found to be
very close (average 91.3 % and standard deviation 1.5%) for all the four
samples suggesting that the process is effectively reproducible. Similarly the
average pore size for all the four samples is also very close, which shows that
porous scaffolds bearing uniform pore sizes could be reproduced.
19
The foregoing description of the specific embodiments will so fully reveal the
general nature of the embodiments herein that others can, by applying current
knowledge, readily modify and/or adapt for various applications such specific
embodiments without departing from the generic concept, and, therefore, such
adaptations and modifications should and are intended to be comprehended
within the meaning and range of equivalents of the disclosed embodiments. It
is to be understood that the phraseology or terminology employed herein is for
the purpose of description and not of limitation. Therefore, while the
embodiments herein have been described in terms of preferred embodiments,
those skilled in the art will recognize that the embodiments herein can be
practiced with modification within the spirit and scope of the appended claims.
ADVANTAGES OF THE INVENTION
As described above, the method of preparing three dimensional porous
scaffolds of the present invention is advantageous due to the following reasons:
The process is simple and the pore size can be easily controlled, also the
problem caused by the secretion and existence of the toxic substance can be
avoided and high efficiency can be achieved.
The other advantage of the process is that it is a single step process and use of
organic solvents can be avoided in making three-dimensional porous scaffolds
wherein the process is suitable not only for homo-polymers but also for crosslinked
polymers.
The Polymerization is based on ring-opening polymerization method and the
presence of a hygroscopic compound such as NaCl (used a as porogen) doesn't
affect the polymerization,
In the present invention no inert atmosphere is required for synthesizing the
materials and Lactones, lactides and cyclic carbonates are used as monomers.
20
Monomers or catalyst having purity of up to 93-100 % can be used and the
process have the potential to convert a batch process of generating a three
dimensional porous scaffold via particulate leaching technique into a
continuous process.
The present invention has been described in an illustrative manner, and it is to
be understood that the terminology used is intended to be in the nature of
description rather than of limitation. Many modifications and variations of the
present invention are possible in light of the above teachings. It is also to be
understood that the following claims are intended to cover all of the generic
and specific features of the embodiments described herein and all the
statements of the scope of the embodiments which as a matter of language
might be said to fall there between. Therefore, it is to be understood that within
the scope of the appended claims, the invention may be practiced otherwise
than as specifically described.
21

CLAIMS
What is claimed is:
1. A single step, solvent free method of preparing a three-dimensional porous
scaffold through a ring-opening polymerization process, comprising the step
of:
a. preparing a homogenous mixture comprising of monomer, scaprolactone
(CL) of at least 0.5 to 10 grams and catalyst, Sn(0ct)2 of
at least 0.01 to 0.1 mol% of CL in a pre dried mould
b. adding a known quantity of a pre-meshed and a pre-heated porogen
(NaCl crystals) which is at least 5 to 10 wt% of CL to the mould
containing the homogeneous mixture
wherein the above mould is than immediately placed inside an oven with a
preset temperature of at least 160 ± 20°C for at least 2.0 ± 0.5 hour and a
porous film so formed, is cooled to the room temperature in a desiccator
and removed and then the porogen is leached out from the film in a water
bath for at least 48 hours with several rounds of changing the water and the
film obtained finally is dried under vacuum and stored for characterization.
2. The method as claimed in claim 1, wherein a cross-linking agent (bis(ecaprolactone-
4yl) - BCY is added along with monomer and catalyst in the
mould with the similar process to obtain a cross-linked porous film.
3. The method as claimed in claim 1, wherein monomer in the homogenous
mixture is at least one selected from the group consisting of Lactones (4 to 16
C cyclic esters), Lactides (L-,D- and D,L- Lactide) and Cyclic Carbonates
(Trimethylene carbonate {TMC} and substituted TMC).
4. The method as claimed in claim 1, wherein catalyst in the homogenous
mixture is at least one selected from the group consisting of Sn(0ct)2
(Stannous Octoate), Al(iPro)3 (Aluminum Isopropoxide) or their substituted
derivatives.
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5. The method as claimed in claim 1, wherein porogen added in the homogenous
mixture is hygroscopic and at least one selected from the group consisting of
any thermally stable (up to 180°C) common lab salt which is not soluble in CL
but soluble in water such as NaCl, Na2S04, MgS04, and NaaCOs.
6. The method as claimed in claim 1, wherein the porosity is generated in-situ,
by the addition of porogen during the polymerization process.
7. The method as claimed in claim 1, wherein the process is ring-opening
polymerization involving both homo-polymerization and cross-link
polymerization.
8. The method as claimed in claim 1, wherein the process is single step, solvent
free process and is carried out without the use of organic solvents.
9. The method as claimed in claim 1, wherein the monomers or catalyst having
93-100% purity are used for generation of three-dimensional porous scaffolds.
10. The method as claimed in claim 1, wherein three-dimensional scaffolds
developed comprises of aliphatic polyesters or aliphatic carbonates using
lactones, lactides or cyclic carbonates as monomers and organometallic
compounds as catalysts for polymerization.
11. The method as claimed in claim 1, wherein the pore size of the porous scaffold
is controlled and no inert atmosphere is required for synthesizing the three
dimensional porous scaffold.
12. The method as claimed in claim 1, wherein the process can be run in a batch
or continuous manner to generate three-dimensional porous scaffolds.