- Email: [email protected]
In vitro degradation and solute release from erodible polyanhydride supports containing skeletal β-alanine residues
145
COREl.00687
More efficient means of administerinb drugs at controlled rates are continuously being sought. An important alttrnative is biodegradable polymers which do not have to be removed once implanted. A class of biodegradable polymers studied by several groups is the polyanhydrides. The synthesis of a novel class of polyanhydrides containing skeletal amino acid residues was recently described (Staubli et al., 1990). In the present study, the degradation kinetics and the potential applications in controlled drug release of one of these poiymers, poly(N-trimellitylimido-P&mine-co-sebacic anhydride) (PTIASA; 20:80), were investigated simultaneously for the first time. Drug-polymer discs were prepared by compression molding using either acid orange or sodium salicylate as the model drug ( _ 30% weight loading). In vitro incubation experiments were carried out in buffer solutions of pH 1.4 and 10.0 at 37X The appearance ofp-alanine (a polymer by-product) and drug in solution was followed spectrophotometrically for up to 240 h. Mass balances were then performed to determine the rates of oolymer degradation and drug release. Drug-free PTIASA discs decreased in thickness throughout the incubation period at both @I’s but maintained their shape. The degradation rates (in terms of the appearance of palanine in solution) were relatively constant (zero-order kinetics) and higher at pH 10.0. The drug-free discs degraded comoletelv in 7-8 davs at oH IO.0and in about IO davs at OH 7.4. On the other hand. the drubloaded PTIASA discs swel;e~ initially and disintegrated part&lly iater on. In the acid orange:PTIASA experiments about 90% of the drug was released in the first 15 h of incubation, the remaining 10% being released at a relatively constant and slower rate thereafter. When sodium salicylate was used as the test drug, the magnitude of the initial burst in drug release was reduced and the useful life of the formulations extended. This study clearly shows the potential use of these novel amino acid-containing polyanhydrides in controlled release formulations. Key words: Erodible polymers; Polyanhydrides with skeletal amino acid residues: Quantification of polymer degradation products: Polymer degradation mechanisms; Solute release -
5000, U.S.A.
U S.A.
146
Erodible polymers represent a viable alternative for controlled drug release. Since they can degrade through a variety of mechanisms [I], their erosion kinetics can be controlled by proper choice of chain substituents and crosslinking agents. Also, they do not haveto be removed after implantation and many have shown excellent biocompatibility. Several classes of erodible polymers have been studied in connection with controlled drug release applications. These include poly(amides). poly(esters), poly(orthaesters), poly(acetals), poly(organophosphazenes), poly(urethanes), and polyanhydrides. This last group has the most hydrolytically unstable linkage of any erodible polymer, making it readily degradable, and is the basis ofthe present study. The ltydrolytic reactivity ofpolyanhydrides is due LOthe anhydride linkage. Degradation rates may be controlled by changes in the polymer backbone, and their degradation products (carboxyiic and amino acids) are easily metabolized by the body. Implantation studies with polyanhydrides have shown their good biocompatibility (21. These polymers have also been studied in various controlled release applications including insulin delivery [ 3 ], inhibition of neovascularization 141, cancer chemotherapy [5], and for treating brain cancer [ 61. Recently, a synthetic route which allows incorporation of amino acids into a polymeric backbone via anbydride bonds was described [ 71.The amino acids were first converted into d&ids by condensation with trimeliitic anhydride. These
were then converted to their mixed anhydrides by heating at reflex in acetlc anhydride. Finally, the prepolymers were subjected to melt-polycondensation at elevated temperature (IOO-250°C) under vacuum. Several homopolymers were prepared by this route using different amino acid monomers. Also, copolymers with a spacer molecule, sebacic acid, were prepared. Enhanced degradation rates relative to poly(amino acids) and improved mechanical strength were obtained. In the present study, the degradation and solute release characteristics of poly(N-trimellitylimido-fl-alanine-co-sebacic anhydride) with a copolymerization ratio of 20: 80 (Fig. I ) were studied in vitro. Based on its constituents, the degradation products are expected to be N-trimellitic acid, ,&alanine. and sebacic acid. These could form by first breaking the anhydride linkage and then hydrolyzing the imido group [ 71. Acid orange and sodium salicylate were used as the model drugs because they can be followed spectrophotometrically. This study is the first to use a novel class of polyanhydrides containing skeletal fialanine residues in potential controlled drug release applications. Experimental
Poly(N-trimellitylimido-/I-alanine-co-sehacic anhydride) (PTIASA; 20~80) was kindly provided by Andrea Staubli and Dr. Robert S. Langer, Department of Chemical Engineering, Massachusetts Institute of Technology. Its molecular
hb
C
ba/ij
cw2ciQ(CH2)4CH#i~
0
weight (MW) was obtained by gel permeatiun chromatography using polystyrene standards and gave a number average MW of 18.000, a weight average MW of 41,000, and a polydispersity of 2.3. Its identity was verilied by ‘H-NMR (Varian Gemini 300 BB: 300 MHz) “sing deuterated chloroform as solvent and tetramethylsilane as internal reference. The ‘H-NMR spectrum gave the expected results’ [9]: (CDCI,, 6): 8.48 (s. I-%:),8.44 (d, Hi.), 7.98 (d, Hb.), 4.07 (t, H,), 2.93 (1, H?), 2.67 (t, H, SA-TMA), 2.44 (t, H, SA-SA and SA-/?-Ala), 1.65 (m, H,), 1.32 (m, H,). PTIASA was stored in a desiccator under nitrogen. The test solutes (drugs) were acid orange 8 (MW 364.36; -65% dye content; Sigma Chemical Co., St. Louis, MO) and sodium sali, cylate (MW 160.1 I; Fisher Scientific, Springfield, NJ). These were used as supplied. The ni”hydri” reagent used to determine the concentration of/?&mine (a polymer by-product) in solution was purchased from Sigma. Methods Preparation of drug-F’TIASA matrices Dntg-polymer matrices were prepared by compression molding as described by Rosen et al. [8]. Drug-free PTIASA discs (9.5 mm diameterx0.9 mm thickness) were prepared in a Parr Pellet Press (Parr Instrument Company, Moli”e, IL) by applying 2000 psi for 2 minutes at room temperature. Drug-PTIASA discs were prepared by manually mixing the powdered drug and polymer in a predetermined proportion ( -30% drug by weight) followed by pressing as in the drug-free matrices. Table gives the weight and percent
I
drug loading ofall
the discs used in this study.
Crams
DruE
loading (wcieht %) p”
7.4 WI
SCUCE
0. ,028
PV2 P”3 PAAI”
0. ,024 0.1026 0.0998
PAAZ’ P.&A)”
0.1036
PSSI” PSSZD PSS3’ pH 10.0rcrtcs WI
PV? PV3 PAA!” P&AZ” P*Aj’
PSSI” PSSZ PSSZ
0.0
0.1027 0.1067 0.1041 O.lliO
0.0 0.U 29.7 31.5 30. I 31.6 30.7 33.0
0.1010 0.1028 0.1049 0.1034 0.1027 0.1044 0.1033 0.1037 0.1059
0.0 0.0 0.0 30.6 30.1 31.5 32.0 31.0 32.0
‘Aad orange-PTihSAdac; %dium salicylate-PTIASAdisc. to the cap. Twenty milliliters of one of the followina media were added: 0.1 M sodium uhosphateiuffer (pH 9.4; Sigma) or 0.05 M sddium bicarbonate buffer (pH 10.0). The vials were placed in a” incubator-shaker (Infers AG, Bottmiogen, Switzerland) set at 37°C and 100 rpm. The buffers were changed periodically to approximate perfect sink conditions by removing the discs from the vials and placing them in fresh
The spent media were saved in order lo determine the concentration ofall the species of interest. In the experiments reported here, two types of drug-FTIASA matrices were tested at two
buffer.
PA’S for periods of up to 24Q h. Polymer degradation and drug release experiments The edge and one flat surface of all discs were coated with paraffin leaving only one flat surface exposed. Each disc was then suspended in a glass scintillation vial by attaching it to a needle glued ‘SA=scbacir arid: TMA=trimellitylimido-palanroe: Ala=palanine.
P
Assays Polymer degradation and drug release were followed simultaneouslv bv measurine the absorbance of aiiquots &?“e*spent medal using a Perkin-Elmer UV-Vis Spectraphotometer Model Lambda 48 or a Bausch and Lomb Spectronic 501. The only polymer by-product followed was
&danine for which media samples were reacted with the ninhydrin reagent and read at 570 nm. Likewise, acid orange was read at 490 am and sodium salicylate at 295 nm. Appropriate blanks were used to correct all absorbances. Concenlrations were determined from standard curves prepared for each compound. Each assay was run in triplicate. The@lanine assays were incorporated in mass balances of the polymer and used to express its cumulative fractional loss from the discs as a function of time. These calculations were based on tbc original 20:80 PTIASA composition. Similarly, the cumulative fractional release of each drug was calculated by dividing the total mass of drug released up to a given time by the initial loading of each disc.
Fig. 2. Cumulative fractional 10s of,%alamne from drug-free PTIASA dzrcsas a funciion ofincubation
time at CH 7.4 (
Results Qu~Ilta~va analysis
ofthe incubation
experiments
The drug-free PTIASA discs consistently decreased in thickness throughout the incubation period at both pn’s, but maintained their shape. This is suggestive of a surface erosion mechanism as in Rosen et al. 181. Disc disinteeration or swelling was not observed. However, thi drugloaded discs swelled initially and disintegrated partially later ou with concomitant loss of their shape. The swelling could be due to poor binding of drug and polymer during the compression molding procedure, or to the osmotic gradient created by the trapped drug. Other preparative techniques for drug-loaded discs are presently under evaluation in our !aboratory. .-a
tion occuning in l-8 days. However, complete degradation at pH 7.4 took about 10 days. As can be seen, the degradation rate was nearly constant throughout most of the incubation period at both pH’s suggestive of a surface erosion mechanism. Fig. 3 presents the cumulative fractional loss of/?-alanine and the fractional release of acid orange from acid orange-PTIASA discs at pH 7.4 and 10.0. By comparing the top and lower panels, it can be seen that palanine loss was faster at pH 10.0 in agreement with the data from the drugfree PTIASA discs. However, the polymer degradation process was different from the drug-free
discs in that /?-alanine appeared in solution rapFigure 2 shows the cumulative fractional loss of palanine from drug-free PTIASA discs as a function of incubation time at pH 7.4 and 10.0. The data arc presented as the average of three discs at earh time pomt. Good reproducibility was observed in these and in all other sets ofdiscs. It is evident that the rate of palanine loss was faster at pH 10.0 with nearly complete degrada-
idly at the beginning (60-90% j-alanine loss in the first I5 h of incubation) followed by a much slower appearance rate later on. Complete degradation at pH 10.0 took about 3 days, while at pH 7.4 degradation was still not complete after 7 days. Fig. 3 also shows that the cumulative fractional release of acid orange was not affected by pH. Rather, about 90% of the loaded drug was released in the first 15 h followed by a much slower and relatively constant release rate there-
I I
I ‘s
discs. This initial burst in drug release is likely due to the increase in surface area upon disintegration of the drug-loaded discs. It is evident from Fig. 3 that practically ail the drug loaded was released within the time scale of the experiments. Fig. 4 presents the cumulative fractional loss ofpalanine and the fractional releaseof sodium salicytate from sodium salicylate-PTIASA discs at pH 7.4 and 10.0. Once again, the rate of /a&er.
alanine loss was faster at pH 10.0, but complete degradation was not attained within the time scale of the experiments. Contrary to the acid orange_PTIASA discs, the cumulative frassional reiease of sodium salicyiate was strongiy rifected by pH with complete release occurring in about 3 days at pH 10.0. Also, the magnitude of the initial burst in drug release and the fractional re-
150
lease were lower for sodium salicylate than for acid orange. iscussian The first attempt to use polyanhydrides in an erodihle, controlled drug release system was that by Rosen et al. IS]. Their in vitm and in viva studies employed poiy[bis (pcarbonyphenosy)methane] (PCPM) as a prototype polyanhydride. Cholic acid was selected as the model drug which was melt-pressed with the polymer into discs. Upon incubation, the discs showed an initial induction period followed by one of nearly rcromorder erosion during which they decreased in size while maintaining their shape. In addition, both polymer erosion and drug release proliles d+&&d zero-order kinetics. Finally, drugfree PCPM discs showed a half-life of about 47 d.ays in viva. In a later study, Leong et al. [ 51 found that by incorporating a hydrophilic spacer molecule, sebacic acid, in hydrophobic polyanhydride backbonestheir degradation rate could be increased by a factor of 800. Following a recent report by Staubli et al. [ 71 in which a procedure for incorporating amino acids into polyanhydride backbones was desrribed. we decided to undertake a study of the degradation and drug release characteristics in vitro of poly( Iv-trimellitylimido-P-alanine-cosebacic anhydride) with a copolymerization ratio of 20: 80. This is thus the first study to test this novel class of polymers in controlled drug release applications. Two model drugs, acid orange and sodium salicylate, were compressionmolded with PTIASA at _ 30% weight loading. Incubation media consisted of pH 7.4 and 10.0 buffers at 37°C. Perfect sink conditions were maintained throughout the experiment by hequent media changes. This procedure for testing drug-polymer matrices is standard in the controlled drug release literature. Our. results showed that the rate of @-alanine loss irom the drug-free PTIASA discs was pH-
sensitive with complete loss afpalanine from the backbone occurring in 7-8 days at pki 10.0 and in about 10days at pH 7.4. The rates of/I-alanine
loss were nearly constant over the time scale of the experiments suggestive of a surface erosion mechanism. This confirms earlier results with polyanhydrides by Rosen et al. [ 81. Theacid orange-PTIASA discs behaved similarly to fhe drugrfree PTIASA discs with respect to the pH-dependence of the polymer degradation process, but released about 90% ofthe loaded drug in the first 15h of incubation making this particular formulation unsuitable for controlled drug release applications. It was observed that while the drug-free PTIASA discs maintained their shape during incubation, the drug--loaded P’IIASA discs swelled initially and disintegrated partially later on. No attempts were made to quantify the degree of swelling and disintegra-
tion, but extreme care was exercised of not losing sedimented fragments during sampling or media changes. The swelling/disintegration phenomenon mentioned above could be due to a variety of physical and chemical factors including rhe method of preparation ofthe drug-PTtASA discs. For example, Staubli et al. [7] observed that melt-cast samples of poly (N-trimellitylimidoglycine-co-scbacic anhydride) disintegrated twelve times slower in 0. I M sodium phosphate buffer (pH 7.4) than compression-molded samples similar to the ones used in the present study. Another important factor in determining the extent of the swelling/disintegration phenomenon is the drug itself. In this study, the use of sodium salicylate in the drug-PTIASA discstieduced the mamitude of the initial bunt in drua release
(relative to the acid orange-PTIASA d&s) and extended the useful life of the formulations. The method of preparation, drug loading, and partide size are presently under investigation in our ongoing. studies with these amino acid-containing polyanhydrides.
We gratefully acknowledge Andrea Staubli and Dr. Robert S. Laager, Depafiment of Chemical Engineering, Massachusetts Institute of Tech-
nology, Cambridge, MA, for providing us with
several PTIASA also thank
samples and encouragement.
Elizabeth
Feliir
for
her excellent
WC sec-
retarial assistance. Finally, we would like to thank the NSF-EPSCoR Program for funding this praject through grant RII-8610677.
4
R. Lund, D. Lcong, K, Fokman, ,. Co”lrclled ~C,EBECof macmm~!rx~,~s: Biological studies, 1. CanLanger.
trolled Release, 2 (1985) 331-341. Leon& KW, Bran. BC. Langw. R. Bmerodible polyanhydridcs as drwpcarner matrices. 1. Characlcrization, degradation. and release characteristics, J. Biom~ul. Mater. as.. 19 (1985,94,-955. 6 Langrr. R, Emmaterials in conlrollcd drug delivery: NFW per~paclwer from biolecbnological advances. PharmaCeUticQ,Technology, 13 ( Ron, E. Langer. R. Hydrolytically degradable ammo acld containing polymers, J. Am. Chcm. Sot.. (1950) 4419-4424. 8 Rosen, HB, Chang. J, Wnek, GE, Linhardl. RJ, Langcr, R, B,oe,od,ble polyanhydrides far controlled dNg deli”cry, Biomaterialr,4 (1983) 131-133.
5
7 S,au~4,
i1980) 51-57. 2
Leon& KW, D’Amore. P, Marlcua. M. Langer, R, Bioeredible poiyanbydridcs 8s drug-tamer matrices. II. Biocompatibibty and chemical rcac~irity, I. Bnmcd. Maler. Res..20(1986~51-64. 3 Mathtowtz. E, Luger, R. Polyanhydridr microrpheres ar drUg tamers. I. Hobmcll micrxncapsulatmn, 3. Conlrolled Rclcasc, 5 (1987) J-22.
I
I989,IS-JO.
II2
COREl.00687
More efficient means of administerinb drugs at controlled rates are continuously being sought. An important alttrnative is biodegradable polymers which do not have to be removed once implanted. A class of biodegradable polymers studied by several groups is the polyanhydrides. The synthesis of a novel class of polyanhydrides containing skeletal amino acid residues was recently described (Staubli et al., 1990). In the present study, the degradation kinetics and the potential applications in controlled drug release of one of these poiymers, poly(N-trimellitylimido-P&mine-co-sebacic anhydride) (PTIASA; 20:80), were investigated simultaneously for the first time. Drug-polymer discs were prepared by compression molding using either acid orange or sodium salicylate as the model drug ( _ 30% weight loading). In vitro incubation experiments were carried out in buffer solutions of pH 1.4 and 10.0 at 37X The appearance ofp-alanine (a polymer by-product) and drug in solution was followed spectrophotometrically for up to 240 h. Mass balances were then performed to determine the rates of oolymer degradation and drug release. Drug-free PTIASA discs decreased in thickness throughout the incubation period at both @I’s but maintained their shape. The degradation rates (in terms of the appearance of palanine in solution) were relatively constant (zero-order kinetics) and higher at pH 10.0. The drug-free discs degraded comoletelv in 7-8 davs at oH IO.0and in about IO davs at OH 7.4. On the other hand. the drubloaded PTIASA discs swel;e~ initially and disintegrated part&lly iater on. In the acid orange:PTIASA experiments about 90% of the drug was released in the first 15 h of incubation, the remaining 10% being released at a relatively constant and slower rate thereafter. When sodium salicylate was used as the test drug, the magnitude of the initial burst in drug release was reduced and the useful life of the formulations extended. This study clearly shows the potential use of these novel amino acid-containing polyanhydrides in controlled release formulations. Key words: Erodible polymers; Polyanhydrides with skeletal amino acid residues: Quantification of polymer degradation products: Polymer degradation mechanisms; Solute release -
5000, U.S.A.
U S.A.
146
Erodible polymers represent a viable alternative for controlled drug release. Since they can degrade through a variety of mechanisms [I], their erosion kinetics can be controlled by proper choice of chain substituents and crosslinking agents. Also, they do not haveto be removed after implantation and many have shown excellent biocompatibility. Several classes of erodible polymers have been studied in connection with controlled drug release applications. These include poly(amides). poly(esters), poly(orthaesters), poly(acetals), poly(organophosphazenes), poly(urethanes), and polyanhydrides. This last group has the most hydrolytically unstable linkage of any erodible polymer, making it readily degradable, and is the basis ofthe present study. The ltydrolytic reactivity ofpolyanhydrides is due LOthe anhydride linkage. Degradation rates may be controlled by changes in the polymer backbone, and their degradation products (carboxyiic and amino acids) are easily metabolized by the body. Implantation studies with polyanhydrides have shown their good biocompatibility (21. These polymers have also been studied in various controlled release applications including insulin delivery [ 3 ], inhibition of neovascularization 141, cancer chemotherapy [5], and for treating brain cancer [ 61. Recently, a synthetic route which allows incorporation of amino acids into a polymeric backbone via anbydride bonds was described [ 71.The amino acids were first converted into d&ids by condensation with trimeliitic anhydride. These
were then converted to their mixed anhydrides by heating at reflex in acetlc anhydride. Finally, the prepolymers were subjected to melt-polycondensation at elevated temperature (IOO-250°C) under vacuum. Several homopolymers were prepared by this route using different amino acid monomers. Also, copolymers with a spacer molecule, sebacic acid, were prepared. Enhanced degradation rates relative to poly(amino acids) and improved mechanical strength were obtained. In the present study, the degradation and solute release characteristics of poly(N-trimellitylimido-fl-alanine-co-sebacic anhydride) with a copolymerization ratio of 20: 80 (Fig. I ) were studied in vitro. Based on its constituents, the degradation products are expected to be N-trimellitic acid, ,&alanine. and sebacic acid. These could form by first breaking the anhydride linkage and then hydrolyzing the imido group [ 71. Acid orange and sodium salicylate were used as the model drugs because they can be followed spectrophotometrically. This study is the first to use a novel class of polyanhydrides containing skeletal fialanine residues in potential controlled drug release applications. Experimental
Poly(N-trimellitylimido-/I-alanine-co-sehacic anhydride) (PTIASA; 20~80) was kindly provided by Andrea Staubli and Dr. Robert S. Langer, Department of Chemical Engineering, Massachusetts Institute of Technology. Its molecular
hb
C
ba/ij
cw2ciQ(CH2)4CH#i~
0
weight (MW) was obtained by gel permeatiun chromatography using polystyrene standards and gave a number average MW of 18.000, a weight average MW of 41,000, and a polydispersity of 2.3. Its identity was verilied by ‘H-NMR (Varian Gemini 300 BB: 300 MHz) “sing deuterated chloroform as solvent and tetramethylsilane as internal reference. The ‘H-NMR spectrum gave the expected results’ [9]: (CDCI,, 6): 8.48 (s. I-%:),8.44 (d, Hi.), 7.98 (d, Hb.), 4.07 (t, H,), 2.93 (1, H?), 2.67 (t, H, SA-TMA), 2.44 (t, H, SA-SA and SA-/?-Ala), 1.65 (m, H,), 1.32 (m, H,). PTIASA was stored in a desiccator under nitrogen. The test solutes (drugs) were acid orange 8 (MW 364.36; -65% dye content; Sigma Chemical Co., St. Louis, MO) and sodium sali, cylate (MW 160.1 I; Fisher Scientific, Springfield, NJ). These were used as supplied. The ni”hydri” reagent used to determine the concentration of/?&mine (a polymer by-product) in solution was purchased from Sigma. Methods Preparation of drug-F’TIASA matrices Dntg-polymer matrices were prepared by compression molding as described by Rosen et al. [8]. Drug-free PTIASA discs (9.5 mm diameterx0.9 mm thickness) were prepared in a Parr Pellet Press (Parr Instrument Company, Moli”e, IL) by applying 2000 psi for 2 minutes at room temperature. Drug-PTIASA discs were prepared by manually mixing the powdered drug and polymer in a predetermined proportion ( -30% drug by weight) followed by pressing as in the drug-free matrices. Table gives the weight and percent
I
drug loading ofall
the discs used in this study.
Crams
DruE
loading (wcieht %) p”
7.4 WI
SCUCE
0. ,028
PV2 P”3 PAAI”
0. ,024 0.1026 0.0998
PAAZ’ P.&A)”
0.1036
PSSI” PSSZD PSS3’ pH 10.0rcrtcs WI
PV? PV3 PAA!” P&AZ” P*Aj’
PSSI” PSSZ PSSZ
0.0
0.1027 0.1067 0.1041 O.lliO
0.0 0.U 29.7 31.5 30. I 31.6 30.7 33.0
0.1010 0.1028 0.1049 0.1034 0.1027 0.1044 0.1033 0.1037 0.1059
0.0 0.0 0.0 30.6 30.1 31.5 32.0 31.0 32.0
‘Aad orange-PTihSAdac; %dium salicylate-PTIASAdisc. to the cap. Twenty milliliters of one of the followina media were added: 0.1 M sodium uhosphateiuffer (pH 9.4; Sigma) or 0.05 M sddium bicarbonate buffer (pH 10.0). The vials were placed in a” incubator-shaker (Infers AG, Bottmiogen, Switzerland) set at 37°C and 100 rpm. The buffers were changed periodically to approximate perfect sink conditions by removing the discs from the vials and placing them in fresh
The spent media were saved in order lo determine the concentration ofall the species of interest. In the experiments reported here, two types of drug-FTIASA matrices were tested at two
buffer.
PA’S for periods of up to 24Q h. Polymer degradation and drug release experiments The edge and one flat surface of all discs were coated with paraffin leaving only one flat surface exposed. Each disc was then suspended in a glass scintillation vial by attaching it to a needle glued ‘SA=scbacir arid: TMA=trimellitylimido-palanroe: Ala=palanine.
P
Assays Polymer degradation and drug release were followed simultaneouslv bv measurine the absorbance of aiiquots &?“e*spent medal using a Perkin-Elmer UV-Vis Spectraphotometer Model Lambda 48 or a Bausch and Lomb Spectronic 501. The only polymer by-product followed was
&danine for which media samples were reacted with the ninhydrin reagent and read at 570 nm. Likewise, acid orange was read at 490 am and sodium salicylate at 295 nm. Appropriate blanks were used to correct all absorbances. Concenlrations were determined from standard curves prepared for each compound. Each assay was run in triplicate. The@lanine assays were incorporated in mass balances of the polymer and used to express its cumulative fractional loss from the discs as a function of time. These calculations were based on tbc original 20:80 PTIASA composition. Similarly, the cumulative fractional release of each drug was calculated by dividing the total mass of drug released up to a given time by the initial loading of each disc.
Fig. 2. Cumulative fractional 10s of,%alamne from drug-free PTIASA dzrcsas a funciion ofincubation
time at CH 7.4 (
Results Qu~Ilta~va analysis
ofthe incubation
experiments
The drug-free PTIASA discs consistently decreased in thickness throughout the incubation period at both pn’s, but maintained their shape. This is suggestive of a surface erosion mechanism as in Rosen et al. 181. Disc disinteeration or swelling was not observed. However, thi drugloaded discs swelled initially and disintegrated partially later ou with concomitant loss of their shape. The swelling could be due to poor binding of drug and polymer during the compression molding procedure, or to the osmotic gradient created by the trapped drug. Other preparative techniques for drug-loaded discs are presently under evaluation in our !aboratory. .-a
tion occuning in l-8 days. However, complete degradation at pH 7.4 took about 10 days. As can be seen, the degradation rate was nearly constant throughout most of the incubation period at both pH’s suggestive of a surface erosion mechanism. Fig. 3 presents the cumulative fractional loss of/?-alanine and the fractional release of acid orange from acid orange-PTIASA discs at pH 7.4 and 10.0. By comparing the top and lower panels, it can be seen that palanine loss was faster at pH 10.0 in agreement with the data from the drugfree PTIASA discs. However, the polymer degradation process was different from the drug-free
discs in that /?-alanine appeared in solution rapFigure 2 shows the cumulative fractional loss of palanine from drug-free PTIASA discs as a function of incubation time at pH 7.4 and 10.0. The data arc presented as the average of three discs at earh time pomt. Good reproducibility was observed in these and in all other sets ofdiscs. It is evident that the rate of palanine loss was faster at pH 10.0 with nearly complete degrada-
idly at the beginning (60-90% j-alanine loss in the first I5 h of incubation) followed by a much slower appearance rate later on. Complete degradation at pH 10.0 took about 3 days, while at pH 7.4 degradation was still not complete after 7 days. Fig. 3 also shows that the cumulative fractional release of acid orange was not affected by pH. Rather, about 90% of the loaded drug was released in the first 15 h followed by a much slower and relatively constant release rate there-
I I
I ‘s
discs. This initial burst in drug release is likely due to the increase in surface area upon disintegration of the drug-loaded discs. It is evident from Fig. 3 that practically ail the drug loaded was released within the time scale of the experiments. Fig. 4 presents the cumulative fractional loss ofpalanine and the fractional releaseof sodium salicytate from sodium salicylate-PTIASA discs at pH 7.4 and 10.0. Once again, the rate of /a&er.
alanine loss was faster at pH 10.0, but complete degradation was not attained within the time scale of the experiments. Contrary to the acid orange_PTIASA discs, the cumulative frassional reiease of sodium salicyiate was strongiy rifected by pH with complete release occurring in about 3 days at pH 10.0. Also, the magnitude of the initial burst in drug release and the fractional re-
150
lease were lower for sodium salicylate than for acid orange. iscussian The first attempt to use polyanhydrides in an erodihle, controlled drug release system was that by Rosen et al. IS]. Their in vitm and in viva studies employed poiy[bis (pcarbonyphenosy)methane] (PCPM) as a prototype polyanhydride. Cholic acid was selected as the model drug which was melt-pressed with the polymer into discs. Upon incubation, the discs showed an initial induction period followed by one of nearly rcromorder erosion during which they decreased in size while maintaining their shape. In addition, both polymer erosion and drug release proliles d+&&d zero-order kinetics. Finally, drugfree PCPM discs showed a half-life of about 47 d.ays in viva. In a later study, Leong et al. [ 51 found that by incorporating a hydrophilic spacer molecule, sebacic acid, in hydrophobic polyanhydride backbonestheir degradation rate could be increased by a factor of 800. Following a recent report by Staubli et al. [ 71 in which a procedure for incorporating amino acids into polyanhydride backbones was desrribed. we decided to undertake a study of the degradation and drug release characteristics in vitro of poly( Iv-trimellitylimido-P-alanine-cosebacic anhydride) with a copolymerization ratio of 20: 80. This is thus the first study to test this novel class of polymers in controlled drug release applications. Two model drugs, acid orange and sodium salicylate, were compressionmolded with PTIASA at _ 30% weight loading. Incubation media consisted of pH 7.4 and 10.0 buffers at 37°C. Perfect sink conditions were maintained throughout the experiment by hequent media changes. This procedure for testing drug-polymer matrices is standard in the controlled drug release literature. Our. results showed that the rate of @-alanine loss irom the drug-free PTIASA discs was pH-
sensitive with complete loss afpalanine from the backbone occurring in 7-8 days at pki 10.0 and in about 10days at pH 7.4. The rates of/I-alanine
loss were nearly constant over the time scale of the experiments suggestive of a surface erosion mechanism. This confirms earlier results with polyanhydrides by Rosen et al. [ 81. Theacid orange-PTIASA discs behaved similarly to fhe drugrfree PTIASA discs with respect to the pH-dependence of the polymer degradation process, but released about 90% ofthe loaded drug in the first 15h of incubation making this particular formulation unsuitable for controlled drug release applications. It was observed that while the drug-free PTIASA discs maintained their shape during incubation, the drug--loaded P’IIASA discs swelled initially and disintegrated partially later on. No attempts were made to quantify the degree of swelling and disintegra-
tion, but extreme care was exercised of not losing sedimented fragments during sampling or media changes. The swelling/disintegration phenomenon mentioned above could be due to a variety of physical and chemical factors including rhe method of preparation ofthe drug-PTtASA discs. For example, Staubli et al. [7] observed that melt-cast samples of poly (N-trimellitylimidoglycine-co-scbacic anhydride) disintegrated twelve times slower in 0. I M sodium phosphate buffer (pH 7.4) than compression-molded samples similar to the ones used in the present study. Another important factor in determining the extent of the swelling/disintegration phenomenon is the drug itself. In this study, the use of sodium salicylate in the drug-PTIASA discstieduced the mamitude of the initial bunt in drua release
(relative to the acid orange-PTIASA d&s) and extended the useful life of the formulations. The method of preparation, drug loading, and partide size are presently under investigation in our ongoing. studies with these amino acid-containing polyanhydrides.
We gratefully acknowledge Andrea Staubli and Dr. Robert S. Laager, Depafiment of Chemical Engineering, Massachusetts Institute of Tech-
nology, Cambridge, MA, for providing us with
several PTIASA also thank
samples and encouragement.
Elizabeth
Feliir
for
her excellent
WC sec-
retarial assistance. Finally, we would like to thank the NSF-EPSCoR Program for funding this praject through grant RII-8610677.
4
R. Lund, D. Lcong, K, Fokman, ,. Co”lrclled ~C,EBECof macmm~!rx~,~s: Biological studies, 1. CanLanger.
trolled Release, 2 (1985) 331-341. Leon& KW, Bran. BC. Langw. R. Bmerodible polyanhydridcs as drwpcarner matrices. 1. Characlcrization, degradation. and release characteristics, J. Biom~ul. Mater. as.. 19 (1985,94,-955. 6 Langrr. R, Emmaterials in conlrollcd drug delivery: NFW per~paclwer from biolecbnological advances. PharmaCeUticQ,Technology, 13 ( Ron, E. Langer. R. Hydrolytically degradable ammo acld containing polymers, J. Am. Chcm. Sot.. (1950) 4419-4424. 8 Rosen, HB, Chang. J, Wnek, GE, Linhardl. RJ, Langcr, R, B,oe,od,ble polyanhydrides far controlled dNg deli”cry, Biomaterialr,4 (1983) 131-133.
5
7 S,au~4,
i1980) 51-57. 2
Leon& KW, D’Amore. P, Marlcua. M. Langer, R, Bioeredible poiyanbydridcs 8s drug-tamer matrices. II. Biocompatibibty and chemical rcac~irity, I. Bnmcd. Maler. Res..20(1986~51-64. 3 Mathtowtz. E, Luger, R. Polyanhydridr microrpheres ar drUg tamers. I. Hobmcll micrxncapsulatmn, 3. Conlrolled Rclcasc, 5 (1987) J-22.
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