Note: Descriptions are shown in the official language in which they were submitted.
~3~3~
POROUS MICROSPHERES FOR DRUG DELIVERY
AND METHOD FOR MAKING SAME
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BACKGROUND OF THE INVENTION
(1) Field of the Invention
The present invention relates generally
to spherical polymer matrices for the controlled
releAse of various drug(s) or other selected agents.
More particularly, this invention describes
methodology for preparing highly porous spherical
polymer matrices with preselected incorporated agents,
e.g., thexapeutics, dispersed within the confines of
the pores therein for controlled delivery to target
physiological systems and resulting biodegradable
microspheric drug carrier or controlled delivery
; 15 systems.
(2) State of the Art
A wide variety of microencapsulation drug
delivery systems have been developed heretofore for
the rate controlled release of therapeutics or other
agents. For instance, considerable research has been
devoted to incorporating therapeutic agents into
polyesters such as poly( E -caprolactone), poly( E -
caprolactone-CO-DL-lactic acid), poly(DL-lactic acid),
poly(DL-lactic acid-CO-glycolic acid) and poly( E -
caprolactone-CO-glycolic acid) in which relea~e was
diffusion controlled. See, lfor example, Pitt, C.G.
(Pitt, C.G., Gratzl, M.M., Jeffcoat, A.R., Zweidinger,
R., Schindler, A., Sustained Drug Delivery Systems.
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II. Factors Affecting Release Rates from Poly(~-
caprolactone) and Related Biodegradable Polyesters.
J. Pharm. Sci., 68, 1534 (1979). These systems were
fabricated as films and capsules and the results
suggest that the devices can be prepared to erode
after release of the drug is essentially completed.
Degradation of at least the polyesters has been
reported to proceed by random hydrolytic cleavage of
ester linkages by an autocatalytic process the rate of
chain cleavage being influenced by chemical and
morphological factors.
Sustained release systems of antimalarial
agents and sulfadiazine in glycolic-lactic acid
copolymers have also ~een reported. Wise, D.L.,
Gesser, J.D., McCormick, G.J., Sustained Release of a
Dual Antimalarial System, J Pharm. Pharmacol., 31,
201 (1979). Wise, D.L., McCormick, G.J., Willet,
G.P., Anderson, L.C., Sustained Release of an
Antimalarial Drug Using a Copolymer of Glucolic/Lactic
Acid, Life Sci., 19, 867 (1976). Wise, D.L.,
McCormick, G.J., Willet, G.P., Anderson, L.C., Howes,
J.F., J Pharm. Pharmacol., 30, 686 (1978). Methods
reported by the foregoing investigators involved
dissolving the agents in a suitable solvent and either
spray drying or casting films according to usual
methods and evaporating the solvent. Various
narcotic antagonists and steroids have been
incorporated i.n films and implanted in rats [e.g., see
Woodland, J.H.R., Yolles, S., Blake, D.A., Helrich,
M., Meyer, F.J., Long-Acting Delivery Systems for
Narcotic Antagonists: I. J Med. Chem., 16, 897
~1973). Jackanicz, T.M., Nash, H.A., Wise, D.L.,
Gregory, J.B., Polylactic Acid as a Biodegradable
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--3--
Carrier for Contraceptive Steroids, Contraceptionl 8,
227 (1973~. Anderson, L.C., Wise, D.L., Howes, JoF~
An Injectable Sustained Release Fertility Control
System, Contraception, 13, 375 (1976)] and
incorporated into particles injected subcutaneously
[Yolles, S., Tim~-Release Depot for Anticancer Drugs:
Release of Drugs Covalently Bonded to Polymers, J
Parent, Drug Assoc., 32, 188 (1978)]. The release of
a number of anti-tumor agents has been evaluated in
implantable systems as reported in Yolles, S., Time
Release Depot for Anticancer Drugs: Release of Drugs
Covalently Bonded to Polymers, J. Parent, Drug Assoc.,
32~ 188 (1978), and the antibiotic Mitomycin C has
been encapsulated in microspherical carriers of
gelatin and administered intraveneously [~oshioka,
T., Hashida, M., Muranishi, S., and Sezaki, H.,
Specific Delivery of Mitomycin C to Liver, Spleen and
Lung: Nano-and Micxospherical Carriers of Gelatin.
Intern J. Pharm., 81, 131 (1981)] and the effect of
size on in vivo distribution and the potential for
antibiotic targeting discussed. The size distribution
of the microspheres (i.e. 5-30 ~m) reported in the
last mentioned publication was very broad, especially
for intravenous administration. Recently ~he in-vitro
release of local anesthetics from polylactic acid
spheres prepared by a solvent evaporation process has,
likewise, been reported [Wakiyama, N., Kaxuhiko, J.,
Nakano, M., Influence of Physicochemical Properties of
Polylactic Acid on the Characteristics and In Vitro
Release Patterns of Polylactic Acid Microspheres
Containing Local Anesthetics, Chem. Phaxm. Bull., 30,
2621 l1982)]. The pattexns of release from these
polylactic acid spheres were characterized by the
various degrees of degradation o the polymer as well
5~
as solubilities of loaded drugs although no attempt
was apparently made to evaluate this parameter.
Additionally, it is apparent that the solubility of
the drug played an important role in the rate and
extent of release. Scanning electron photomicrographs
also revealed varying degrees of erosion and
deformation of the spheres after release.
It will be seen from the foregoing that while
the controlled release delivery of pharmaceuticals or
other agents from heretofore described polymeric
systems has been principally limited to oral, topical
or implantable systems in which the considerations
relative to pore size and/or cell size within the
carrier matrix as well as the overall dimensions of
the microspheres to be administered along with the
rate of release and the relative absorption rate from
a bioavailability standpoint are distinctly different
from the evaluation parameters involved in the
utilization of these microsphere delivery systems for
parenteral, i.e., intravenous, intraarterial,
intraocular or inhalation administration routes to
which the present invention is particularly
applicable.
SUMMARY OF T~E INVENTION
It is, therefore, a primary object of the
present invention to afford novel porous microspheres
for the controlled delivery of drugs or other matrix
confined materials to target organs or systems in
warm-blooded animals in need thereof and to methods
for making such microspheres.
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A further object of the present invention is
to provide methods for preparing porous microspheres
of heretofore unattainable narrow-range size
distribution particularly suitable for use as
parenterally administerable drug delivery systems for
injectable and inhalation dosage forms as well as
facilitating sustained drug release via more
conventional oral administration routes.
It is a still further object of the present
invention to provide porous microsphere matrices
wherein the accessability of the drug or other
incorporated agent is not dependent upon the physical
or chemical erosion of the polymer for release.
Another object of the present invention is to
provide chemically modified polymer compositions
: suitable for use in the spherical polymer matrices of
: the invention whereby porosity a~ well as degradation
of the polymer substrate after release of the matrix
confined agent for release can be predetermined and
controlled.
A still further object of the present
invention is to provide porous polymeric microspheric
~rug delivery systems which allow targeting of drugs
or other agents to specific host tissues or cells via
injection or inhalation providing high localized
concentrations, sustained activity, systemic
administration and treatment not possible by other
methods thereby minimizing undesirable systemic
effects of toxic drugs administered directly into the
circulation.
~3~
These and other similar objects, advantages
and features are accomplished according to the
methods, products and compositions of the present
invention.
BRIEF DESCRIPTION OF DRAWINGS
FIGURE 1 is a drawing of a polymer with a low
degree of crystallinity in accordance with the
practice of the present invention and a drawing of a
polymer with a high degree of crystallinity.
FIGURE 2 is a graph of half-life in months
versus various ratios of polyglycolic (PGA) and
polylactic ~PLA) as copolymer implanted in rat
tissues.
FIGURE 3 is a graph of percent water uptake
versus percent glycolic acid for glycolide/lactide
copolymers~
FIGURES 4 and 4A generally depict the
preparative methods of the present invention.
FIGURE 5 depicts the shape and surface
appearance of polyglycolic (PGA) microspheres prepared
by Dilution-Precipitation Method.
FIGURE 6 depicts the shape and surface
appearance of PGA microspheres prepared by Freeze Dry
Method.
FIGURE 7 is a graph of the release profile
from matrices which were prepared by Precipitation
Method and contain different amounts of marker.
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FIGURE 8 is a graph of the release profile
from matrices which were prepared by Freeze Dry Method
and contain different amounts of marker.
FIGU~E 9 is a graph of the release profile
from matrices which were prepared by Freeze Dry Method
and contain prednisolone acetate.
FIGURE 10 depicts scanning electron
micrography (SEM) micrographs of the PGA matrix
manufactured by Freeze Dry Method 72 hours following
drug release.
FIGURE 11 depicts SEM micrographs of the PGA
matrix manufactured by Freeze Dry Method after 120
hours following drug release.
FIGURE 12 depicts SEM micrographs of the PGA
matrix manufactured by Freeze Dry Method 168 hours
following drug release.
FIGURE 13 is a graph of the release of dye
from polymer in plasma.
FIGURE 14 depicts PGL microspheres containing
blue dye manufactured by Dilution-Precipitation
Method.
FIGURE 15 depicts gelatin microspheres
manufactured by a modified Dilution-Precipitation
Method.
~5 DESCRIPTION OF THE PREFERRED EMBODIMENTS
The porous polymeric microspheres of the
present invention are derived from copolymeric and
homopolymeric polyesters containing hydrolyzable ester
linkages which are, therefore, biodegxadable.
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Typically preferred of such polyesters are
polyglycolic (PGA) and polylactic (PLA) acids, and co-
polymers of glycolide and L(-lactide) (PGL). The
aforementioned polyesters are particularly suited for
the methods and compositions of the present invention
by reason of their characteristically low human
toxicity and virtually complete biodegradability. Of
course, it will be understood that the particular
polyester or other polymer, oligomer, copolymer~ etc.,
utilized as the microspheric polymer matrix is not
critical and a variety of polymers may be utilized as
a conse~uence of the novel processing methods o~ the
invention which yield the desired microspheres of the
porosity, consistency, shape and size distribution
essentially irrespective of the source of polymer
utilized. Accordinglyl other biodegradable or
bioerodable polymers or copolymers evidencing the
necessary low degree of toxicity suitable for use in
the present invention include, for example, gelatin,
agar, starch, arabinogalactan, albumin, collagen,
natural and synthetic materials or polymers, such as,
poly(~-caprolactone), poly(~ -caprolactone-CO-lactic
acid), poly(s -caprolactone-CO-glycolic acid),
poly( ~-hydroxy butyric acid) t polyethylene oxide,
polyethylene, poly(alkyl-2-cyanoacrylate), (e.g.,
methyl, ethyl, butyl, etc.), hydrogels such as
poly~hydroxyethyl methacrylate), polyamides (e~g.,
polyacrylamide), poly(amino acids) (i.e., L-leucine,
L-aspartic acid,~ -methyl-L-aspartate r ~ -benzyl-L-
aspartate, glutamic acid and the likej,poly(2-hydroxyethyl DL-aspartamide), poly(ester urea),
poly(L-phenylalaninetethylene glycol/1,6-
diisocyanatohexane) and poly(methyl methacrylate).
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The foregoing exemplary natural and synthetic
polymers suitable for use in the present invention
are, of course, either readily available commercially
or are obtainable by condensation polymerization
reactions from the suitable monomers or, comonomers or
oligomers. For instance, homopolymers and copolymers
of glycolic and lactic acids can be prepared by direct
polycondensation or by reacting glycolide and lactide
monomers as disclosed by Gilding, D.K~, Reed, A.M.,
Biodegradable Polymers for Use in Surgery -
Polyglycolic/Poly(lactic acid) Homo~ and Copolymers:
1, Polymer, 20, 1459 (1979). Structurally,
polyglycolic acid (PGA) has the following structure:
H O H O
I 1l I n
_ -o -- C -- C -- o -- C -- C- _
H H n
whereas the related copolymer polyglycolic
acid/polylactic acid (PGL) has the structure depicted
below:
r H 1l H ~oll ~ CH3 1I CH3 01
_ ~O - C - C - O - C - C _ ~ -O - C - C - O - Cl - C- _
H H n _ H H _ n
--10--
~ oth of the foregoing are polyester type
polymers which readily degrade via hydrolysis at the
ester linkages and by appropriately selecting suitable
molecular weight polyesters, modifying ~he degree of
crosslinking and the degree of crystallinity, the
biodegradation properties of such polymers may be
advantageously controlled. As pointed out previously,
however, in accordance with the present invention the
necessity for biodegradation or bioerosion of the
polymer matrix for release of the agent incorporated
therein to occur is obviated by reason of the
intrinsic porosity characteristics of the polymer
matrices of the invention and the fact that the
incorporated agent or agents are matrix confined
within the interconnecting channels or pores of the
spherical polymer. However, in accordance with
alternative and preferred embodiments of the present
invention the possibility that the matrix could be
coated with a film or crosslinking agent to inhibit or
control releaser thereby allowing bioerosion to
influence release is not in any way precluded and
may, in fact, depending upon the nature of the
incorporated agent as well as the rate of release
required in the target organ system may be desirable
or advantageous. For example, in those instances
where it may be desirable to inhibit or retard drug
release rates, more extensive cross-linking of the
copolymer or polymer may be achieved by the addition
of higher concentratiQns of suitable cross-linking
agents such as glyoxal, succinaldehyde,
glutaraldehyde, 3-methylglutaraldehyde,
methyleneacrylamide, bisacrylamide and similar cross-
linking agents. Likewise, the reduction or
elimination of crosslinks in the copolymers or
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polymers of the invention will result in enhanced
biodegradability. On the basis o~ such polymer
modifications, it is evident that the release of the
incorporated agent or agents will be essentially
complete, i.e., 90~ before any erosion or degradation
of the polymer matrix occurs, and, thus, the polymer
composition can be preselected to permit controlled
clearance from the target system after release of the
incorpoxated drug.
The polymers utilized in accordance with the
invention exist in the crystalline form with amorphous
regions interdispersed between the crvstalline areas
as shown, for example, in Figure 1. Hydrolysis rates
have been shown to be higher in the amorphous regions.
For the copolymers of PLA/PGA, the degree of
crystallinity is reduced at a composition of equal
amounts of PLA and PGA. As shown in Figure 2, the
half-life for the degradation of polymer in rat tissue
was lowest at a 50-50 composition. Figure 3 shows
that the water uptake is highest in this range which
constitutes the amorphous region. Therefore
bioerosion occurs in the amorphous regions initially
and eventually the backbone is destroyed and the
matrix will collapse, thereby accelerating the
bioerosion and elimination of the polymer.
Consistent with the controlled conditions of
the methods of the present invention, spherical
polymer matrices or microspheres having a diameter
range between about 0.5 to 150 microns (~m) can be
prepared in narrow size ranges for targeting to
various organ or organ systems via parenteral
injection or inhalation. A more preferred range for
the spherical polymer matrices or microspheres is
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between 0.5 to 50 microns. The integratable methods
for preparing porous spherical matrices consis~ent
with the present invention result in microspheres in
which essentially all of the agent(s) incorporated
within the pores of the drug delivery system is
readily available for release. Essentially the
foregoing principle objective of the invention is
accomplished by forming emulsified droplets or spheres
consisting of a homogeneous mixture of polymer (or
copolymer), solvent and matrix incorporated agent from
a solution of a preselected polymer and agent
dispersed in a continuous (non-solvent phase).
Removal of the solvent from the sphere by either
freeze drying or dilution-extraction-precipitation or
a combination thereof creates the interconnecting
network of pores wherein the incorporated agent is
confined within the walls and channels of the pores as
opposed to random distribution within the more poorly
defined interstices of the polymer. As used in the
specification and claims, the expression "pore
incorporated agent" is used to define the relative
specific location of the agent confined essentially
completely inside the pores of the porous microspheres
of the invention. ~imilarly, the term "agent"
specifically encompasses any diagnostic or pharma-
cologically active material which would be generally
classifiable as a drug suitable for introduction into
a human or other warm-blooded animal host, as well as
other materials or compositions including, for
instance, dyes, antigens, antibodies, enzymes,
flavors, comestibles and the like and mixtures
thereof.
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The drug delivery systems in accordance wi~h
the present invention are ideally suited for
administration by parenteral or inhalation routes. It
will be appreciated by those skilled in the art that
the porous microspheres of the present invention
containing pore incorporated drugs for release to
target cells or tissues, therefore, may be
administered alone or in admixture with appropriate
pharmaceutical diluents, carriers, excipients or
adjuvants suitably selected with respect to the
intended route of administration and conventional
pharmaceutical practices. For example, ~or parenteral
injection, dosage unit forms may be utilized to
accomplish intravenous, intramuscular or subcutaneous
administration, and for such parenteral administra-
tion, suitable sterile aqueous or non-aqueous
solutions or suspensions, optionally containing
appropriate solutes to e~fectuate isotonicity, will be
employed. Likewise for inhalation dosage unit forms,
for administration through the mucus membranes of the
nose and throat or bronchio-pulmonary tissues,
suitable aerosol or spray inhalation compositions and
devices will be utilized.
Consistent with other preferred embodiments of
~5 the present invention, the porous microspheric drug
delivery systems of the invention may be additionally
coated or modified to advantageously influence the
targeting of the release of the incorporated drug
therein to preselected target cells, tissues or
organs. For example, the drug delivery microspheres
may ~e coated with various agents, e.g., proteins,
surfactants, antibodies or receptor site specific
drugs which may be the same or different from those
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-14-
incorporated in the porous microsphere whereby the
release of the incorporated drug is concentrated at
the targeted system.
The preparative methods of the present
S invention are generally depicted in Figures 4 and 4AD
In accordance with the methods for making ~he
porous microspheres of the invention, the desired
polymer or copolymer and the drug(s~ or other agentts)
are dissolved separately in a suitable solvent. The
polymer and drug solutions are mixed together in the
appropriate manner to provide a polymer concentration
ranging between about 205 to 18~ w/w and a
drug:polymer ratio ranging between about 1:1 ~o 1:10.
The temperature of the resultant solution is
controlled between about 30-45C. The drug-polymer
solution comprising the dispersed phase is dispersed
into the continuous phase containing an appropriate
surface active agent at a thermostatically controlled
temperature generally in the range of 10-20C. The
foregoing is accomplished by forcing the dispersed
phase under pressure through a fine orifice nozzle.
The continuous phase which is 10-20 times by weight of
the dispersed phase is then agitated by a dispersator.
Following the introduction of the dispersed phase,
one of two recovery methods ~see Figure 4) is utilized
to stabilize and recover the drug-loaded microspheres
for final processing.
More specifically, consistent with the freeze-
dry method of the invention, following dispersion, the
temperature is maintained at 10-20C, preferably 15C,
for two minutes then increased to 45-55C, preferably
50C, over a three minute period. Vigorous agitation
of the mixture is continued during this period. When
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the temperature reaches 50C, either a refrigerant
solution is circulated through the jacket from the
bath or the container is immersed in dry ice-methanol
and cooled to a temperature which will freeze the
drug-polymer-solvent phase and not the continuous
phase. The suspension or emulsion (solid dispersion
phase in liquid continuous phase) is quickly
transferred to precooled vials (-40~ to -60C) and
cooled to -40 to -60C in a freeze dryer, freezer or
dry ice-acetone bath. The solvent in the suspended
droplets (microspheres) and the ~ontinuous phase
solvent are removed by freeze drying. Upon completion
of the freeze dry cycle the microspheres are washed
with a suitable solvent, filtered and air dried.
In the dilution-extraction-precipitation
method of the invention, following dispersion, the
temperature is maintained at 10-20C, preferably 15C,
for two minutes, then increased to 45-55C, preferably
50C, over a three minute period. The dispersion is
then transferred to a vessel containing a diluent
solvent at room temperature as depicted in Figure 4.
Agitation is continued for 30 minutes using a
vibromixer. During the pxocess the dispersed phase
solvent is removed from the drug-polymer-solvent
~5 emulsion droplets by extraction causing solidification
of the droplets. The solid spheres are then removed
by filtration, washed with a suitable solvent and air
dried.
Solvents for the dispPrsed phase and the
continuous phase will of course differ in order to
attain phase ~eparation and, are therefore, selected
based upon the solvent requirements for each phase.
More particularly, the solvent for the dispersed
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phase should dissolve the polymer and the incorporated
agent and remain in the emulsified droplets with the
drug and polymer in the continuous phase until leached
out by a diluent solvent or removed by vaporization or
evaporation. In this way pores are formed in the
drug-polymer matrix. In the case of PGA polymer
into which water soluble markers or agents are
incorporated, hexafluoroacetone sesquihydrate (HFA) is
an appropriate solvent. Other solvents which can he
used, depending upon characteristics of polymer and
incorporated agents, include water,
hexafluoroisopropanol (HFIP), methylene chloride,
tetrahydrofuran, hexane, benzene and the like.
Solvents for the continuous phase should not dissolve
the polymer and should emulsify the dispersed phase.
Solvents include benzene, dioxane, acetone, methylene
chloride, chloroform, carbon tetrachloride, toluene,
ethyl alcohol, acetonitrile, p-xylene, tetrahydrofuran
and mixtures of these solvents.
A diluent (non-solvent) phase can also be
employed to dilute the continuous phase following
dispersion of the polymer-agent solution. The diluent
should be miscible with the continuous phase and
dispersed phase solvents but not dissolve the polymer
or incorporated agent. Examples of solvents include
1,4-dioxane, cyclohexanone, acetone, ethanol,
acetonitrile, dimethylformamide, tetrahydrofuran and
cyclohexanol.
The concentration of polymer in the dispersed
phase directly influences the porosity or l'void" space
in the final microsphere product as well as the shape
of the microsphere. A concentration of 2.5~ to 10%
w/w polymer yields dimensionally suitable spherical
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particles. With respect to the concentration of the
pore incorporated agent, up to 50~ by weight of the
polymer has been achiPved with consistent results.
In accordance with another preferred
embodiment of the present invention, hydrophilic
colloids are employed to improve the yield and prevent
phase inversion in the continuous and diluent phases.
Substances which can be utilized in concentrations
ranging between about 0.5 to 5% include anionic
surfactants such as sorbitan, gelatin and gelatin
derivatives, polyvinyl alcohol, polystyrene sulfonate,
hydroxyethylcellulose, hydroxypropylcellulose and
related colloids with suitable hydrophilicity.
It has been determined that certain processing
parameters influence the recovery methods as well as
the resultant microspheres of the present invention.
Identifiable parameters include the concentration of
polymer in the dispersed phase, the ~emperature of the
dispersed phase at the time of dispersion, the
concentration of surfactants in the dispersed phase as
well as the ratio of incorporated agent to polymer in
the dispersed phase. It will be appreciated that the
concentrations, temperatures and ratios referred to
hereinabove and in the Examples set forth operable
ranges and that other numerical expressions may apply
as different solvents, polymers, incorporated agents,
etc. are selected.
The shape and surface appearance of
microspheres prepared in accordance with the recovery
methods of the invention were assessed by Scanning
Electron Microscopy (SEM), Figures 5-6 as well as by
optical micrography.
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Typical release profiles are also shown in
Figures 7-~. The very water soluble FD&C dye Blue #1
releases within 1-3 days depending upon the
concentration of dye (i.e., marker). In
all instances release is completed before discernible
degradation or erosion of the matrix occurs. Figures
10 and 11 show SEM micrographs of the spheres
following 72 hours and 120 hours, respectively, in the
dissolution media. The spheres were essentially
intact indicating minimal erosion. The release of a
less-soluble compound, prednisolone acetate is shown
in Figure 9. Essentially 90% of the drug was released
after 7 days and degradation of the matrix was very
evident after 7 days as shown by the fragmentation in
Figure 12.
The following non-limiting Examples are
afforded in order that those skilled in the art may
more readily understand the present invention and
specific preferred embodiments thereof with respect to
the methods and compositions in accordance with the
foregoing description.
Example 1 FD&C Blue #l - PGA microspheres (freeze
dry recovery)
l. 0.1 g of FD~C Blue #1 was dissolved in 9.9 g of
HFA to make a 1% (w/w) solution.
2. 1.0 g of PGA was dissolved in 9.0 g of HFA to make
a 10% (w/w) solution.
3. Equal weights of the above are mixed together to
form the dispersed phase. The resultant spheres
from this combination are very porous having 94.5
"voidl' space and a dye~polymer ratio of 1:10. In
this example 2.0 g of each solution were mixed
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together and maintained at 37C. The spherical
microporous polymeric network has a degree of
porosity of between about 80 to 98 percent as
determined by relative void space in relation to
the starting concentration of the polymer~
Dispersed Phase Concentrations:
DYE POLYMER SOLVENT
20 mg 200 mg 3780 mg
0.5% 5.096 94.5
4. The continuous phase constituted 160g of CC14
containing 0.1% sorbitan sesquioleate (SO-15)
which was maintained at 15C in
a 500 ml jacketed vessel. A dispersator was
located at the center of the vessel for
mixing.
5. The dve-polymer-solvent solution was then
dispersed via pressure through a fine orifice
into the continuous phase which was agitated
vigorously with the dispersator. The
temperature was maintained at 15C and the
mixing continued for 2 minutes. The
temperature was then increased to 5 0 ~C over a
3 minute period by either circulating 70C
water through the jacket (or immersing the
vessel in a 70C water bath).
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. When the temperature reached 50, a refrigerant
solution at -22C was circulated through the
jacket to freeze the dispersed phase and not the
continuous phase (f.p. of CCl~=-22.6).
7. The above suspension was quickly transferred to
pre-cooled (ca -45C) 50 ml vials and cooled to
-40 to -50 on the shelves in a freeze dryer which
had been precooled to -50C.
8. The suspension was maintained at -50 for 1
hour. Vacuum was applied and the shelves
heated to -10C and maintained at this
temperature for 24 hours to remove the CC14.
~he temperature of shelves was increased to
20C for 24 hours to remove the HFA. The
temperature of the shelves was increased to
35 and maintained for 2 hours to ensure
removal of all solvent.
9. The vials containing the spheres were removed from
the chamber and stoppered pending washing and
evaluation~
Example 2 Prednisolone acetate - PGA microspheres
tfreeze dry recoveryl
1. 0.1 g of prednisolone acetate was dissolved in
9.9 g of HFA to make a 1% w/w solution~
2. 1.0 g of PGA was dissolved in 9.0 g of HFA to
make a 10% w/w solution.
3. 2.0 g of each solution were mixed together and
maintained at 37C.
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Dispersed Phase Concentrations:
DRUG POLYMER SOLVENT
20 mg 200 mg 3780 mg
0.5% 5.0~ 94.5%
4-9. Steps four through nine were the same as in
Example 1.
Spheres obtained by the freeze dry method were
washed twice with 125 ml volum~s of acetone and
collected on 0.8, 10, & 50 ~m filters. The spheres
obtained by the precipitation method were washed with
acetone in the size ranges previously obtained by
filtration and collected on 0~8, 10, & 50 ~m filters.
Washing removes approximately 8.5~ of the dye or drug
from the sphere.
Example 3 Characterization of Microporous
Microspheres and Release of Model
Compounds
SEM Photomicrographs of Examples 1 and 2 are
shown in Figur~s 5-6 at lO-fold differences in
magnification. The porous nature is evident from the
topography of the magnified surfaces of both methods
of preparation~
In-vitro release from the microspheres was
determined in 0.1 M phosphate buffer (pH 7.4). The
spheres were quantitatively transferred to a 15 ml
cuvette tube with a screw cap and the buffer added.
The tubes were placed on a rocker-type shaker in an
oven at 37C. The tubes were centrifuged at various
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~36~
-22-
times and solution samples were removed for
spectrophotometric analysis at 630 nm for FD&C Blue #1
and at 245 nm ~or prednisolone acetate. Release
profiles are shown in Figures 7-9 for Examples
l, 2 & 3 along with the profiles of other
compositions. The release of water soluble dye was
essentially complete in 2 to 3 days with spheres
prepared by the dilution-extraction-precipitation and
freeze dry methods while the less-soluble prednisolone
acetate releases much more slowly, 90% in 7 days.
In experiments at a fixed level of dye, i.e.,
4~ (by weight of polymer) and variable polymer
concentration in the dispersed phase (2.5%, 5~ & 10%),
the release rate was reduced in relation to the
polymer concentration (Figure 13). The "void"
space or the porosity is controlled by the polymer (or
solvent) concentration of the dispersed phase.
Example 4 FD&C Blue #1 - PGL microspheres
(Dilution-Extraction-Precipitation
Recovery)
l. 0.1 g of FD&C Blue #l was dissolved in 9.9 g of
HFA to make a 1% (w/w) solution.
2. 0~5 g of polyglactin 910 (VicrylR) was dissolved
in 4.5 g of HFA to make a 10% (w/w) solution.
3. Equal weights of the above were mixed together to
form the dispersed phase and maintained at 37C.
.' . ~ ~ -' '
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Dispersed Phase Concentrati~ns:
DYE POLYMER SOLVENT
20 mg 200 mg 3780 mg
0.5% 5.0% 94O5~
The resultant spheres from this combination ratio
will be very porous having 94.5~ "void" space and
a dye-polymer ratio of 1:10.
4-5. Steps four and five are the same as in
~xample 1.
6-8. Steps six through eight are the same as in
Example 2. See Figure 14 for topography of
the microspheres.
Example 5 FD&C Blue #1 -~ Gelatin microspheres
~Dilu~ion-Extraction-Precipitation
Recovery)
1. 0.1 g of FD&C Blue #1 was dissolved in 10.0 g of
10% w/w aqueous gelatin solution. 3.0 g of this
mixture was maintained at 37C as a dispersed
phase.
Dispersed Phase Concentrations:
DYE POLYMER SOLVENT
30 mg 300 mg 2670 mg
: 1.0~ 10.0% 8g.0~
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2. The continuous phase constituted 120 g of CC14
containing 2% S0-15 which was maintained at 40 in
a 500 ml jacketed vessel. A dispersator was
located at the center of the vessel for mixing.
3. 3.0 g of the dye-gelatin-water solution was
dispersed via pressure through a fine orifice into
the continuous phase which was agitated
vigorously. The temp. was maintained at 40C and
the mixing continued for 3 min.
4. lO0 g of 1,4-dioxane containing 2% S0-15 was added
slowly to the emulsified dye-gelatin-water system
in CC14 to harden the gelatin matrix and agitation
continuedfor30minutes.
5. A refrigerant solution was circulated through the
jacket and the system cooled to 14.
6. 50 g of a curing solution, consisting of 10 g of
50~ glutaraldehyde and 40 g of 1,4-dioxane
containing 2% S0-15, was added dropwise (4 ml/min)
to the dye-gelatin-water system in CC14.
Agitation continued at 14C for 30-60 minutes.
7. ~he suspension was then filtered through a series
of filters to collect the spheres in various size
ranges, followed by washing. See Figure 15 for
topography of the microspheres.
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~D8;~
-25-
While the invention has been described and
illustrated with reference to certain preferred
embodiments thereof, those skilled in the art will
appreciate that various changes, modifications and
substitutions can be made therein without departing
from the spirit of the invention. It is intended,
therefore, that the invention be limited only by the
scope of the claims which follow.
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