Biodegradation of Lactic-Glycolic Acid Oligomers - ACS Symposium

Chapter 19

Biodegradation of Lactic-Glycolic Acid Oligomers 1

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Nuo Wang , Jin Song Qiu , and Xue Shen Wu

Downloaded by UNIV OF AUCKLAND on December 16, 2015 | http://pubs.acs.org Publication Date: September 24, 1998 | doi: 10.1021/bk-1998-0709.ch019

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Division of Pharmaceutics and Industrial Pharmacy, Arnold and Marie Schwartz College of Pharmacy and Health Sciences, Long Island University, Brooklyn, NY 11201 Herman F. Mark Polymer Research Institute, Polytechnic University, Brooklyn, NY 11201 2

The biodegradation of lactic/glycolic acid (LGA) oligomers having different composition and molecular weight has been investigated. The results of this investigation show that composition and molecular weight of LGA oligomers and pH of incubating media all affect biodegradation of LGA oligomers. A higher content of glycolic acid residue in an LGA oligomer results in a higher biodegradation rate of the oligomer. For oligomers having the same composition and in a constant pH medium, the weight loss of a higher molecular weight LGA oligomer is slower than that of a lower molecular weight counterpart. The weight loss of the LGA oligomers is faster in an alkaline incubating medium (pH 9.4) than that in a neutral incubating medium (pH 7.4). When distilled water is used as an incubating medium, the pH of the incubating medium drops rapidly along with the biodegradation of the LGA oligomers. A three -step biodegradation mechanism is proposed for the LGA oligomers. These three steps are hydration, matrix bulk degradation, and surface erosion-controlled solubilization. The controllable biodegradation properties of the LGA oligomers have useful applications to controlled -release drug delivery.

Lactic/Glycolic acid (LGA) polymers, including poly(lactic acid), poly(glycolic acid) as well as their co-polymers are one of the most promising biomaterials in the medical and pharmaceutical area because of not only their outstanding biocompatibility but also their controllable or programmable biodegradation behavior. The biodegradation products of these polymers are lactic and/or glycolic acid that are (is) readily metabolized and eliminated from the body. Many researchers (1-7) have investigated these polymers in the molecular weight range from 5,000 to 320,000. Biodegradation of these polymers is believed to undergo bulk hydrolysis throughout the polymer matrix (8). The in vitro and in vivo experiments reveal that many factors may influence the biodegradation of LGA polymers. These factors include polymer composition (ratio of lactic to glycolic acid in a copolymer) (9), molecular weight of polymers (10, 11), pH

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©1998 American Chemical Society

In Tailored Polymeric Materials for Controlled Delivery Systems; McCulloch, I., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1998.

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of an incubating medium (12, 13), buffer effect (13, 14), and ionic strength of an incubating medium (13). However, little data are available for biodegradation of low molecular weight lactic/glycolic acid polymers or LGA oligomers. The biodegradation mechanism, kinetics, and factors affecting the biodegradation of LGA oligomers are still not well studied. To clearly understand the behavior of LGA oligomers in an aqueous medium, we investigate the in vitro biodegradation mechanism, kinetics, and factors affecting the biodegradation of LGA oligomers. The factors affecting the biodegradation are investigated with regards to oligomer composition, molecular weight of oligomers, and pH of an incubating medium. Experimental Materials. LGA oligomers having three different compositions were synthesized and characterized in our laboratory and previously reported (15). The three compositions of the LGA oligomers were LGA046/54 having molar ratio of lactic to glycolic acid moiety of 46 to 54 (Mn: 990), LGA065/35 having that of 65 to 35 (Mn: 912), and LGA072/28 having that of 72 to 28 (Mn: 1317 and 3025). The variation of the number average molecular weight of LGA072/28 was generated by varying the polymerization time. The oligomers were ground into small pieces having approximately 5 mm in diameter before use. All other chemicals were purchased from Spectrum Chemical Manufacturing Corp. (Gardena, CA). Investigation of Composition Effect on Biodegradation of the LGA Oligomers. Three oligomer samples with different compositions, LGA046/54, LGA065/35, and LGA072/28, were used in this experiment. For each composition, ten oligomer samples (0.5 g for each sample) were weighed and placed in ten test tubes. All test tubes containing the oligomers were incubated in a phosphate buffer (0.2 M, pH 7.4) and placed in a 37 °C water bath shaking at 30 rpm. At predetermined time intervals, one test tube in each sample group was taken out of the water bath and centrifiiged for 10 min at lOOOxg. The supernatant was discarded and the test tubes were dried in vacuo to a constant weight at 40 °C. The weight of the test tubes was recorded after drying. The weight loss of the LGA oligomers was calculated using the following equation: Weight Loss (%) = (W -W )/W = 1 - W /W =1 - (W - W )/W Q

R

0

R

0

T

E

0

(1)

where W is the original weight of an oligomer, W is the remaining weight of an oligomer after incubation or biodegradation, W T is the weight of a test tube plus the remaining oligomer, and W E is the weight of the test tube, respectively. The biodegradation profile for each oligomer was obtained by plotting percent weight loss of the oligomer versus time. 0

R

Assay of Molecular Weight Effect on Biodegradation of the LGA Oligomers. LGA072/28 having molecular weight of 1,317 and 3,025 were used in this experiment. Ten oligomer samples for each molecular weight were weighed (0.5 g for each sample)


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and placed in test tubes with known weight. All test tubes were incubated in a phosphate buffer (0.2 M, pH 7.4) and placed in a 37 °C water bath shaking at 30 rpm. At predetermined time intervals, one test tube in each molecular weight sample was taken out of the water bath and processed following the same procedures described in the section of "Investigation of Composition Effect on Biodegradation of the LGA Oligomers". Measurement of pH Change of the Incubating Medium during Biodegradation of the LGA Oligomers. The LGA oligomers with the three different compositions (LGA046/54, LGA065/35, and LGA072/28) were incubated in distilled water separately in three test tubes. The test tubes were shaken at 30 rpm in a 37 °C water bath. At predetermined time intervals, the pH of the incubating medium in each sample was measured and recorded. The pH of the incubating medium was then plotted versus incubating time. Study of pH Effect on Biodegradation of the LGA Oligomers. The effect of pH on biodegradation of the LGA oligomers was determined by comparing the biodegradation of LGA oligomers in two different pH conditions. The LGA072/28 having a number average molecular weight of 3,025 was chosen for this study. The oligomer was divided into two sample groups. Each sample group has ten oligomer samples (0.5 g oligomer for each sample). Each oligomer sample was placed in a test tube. One group of the samples was incubated in a phosphate buffer (200 mM, pH 7.4) and the other group was incubated in a Na2B O7«10 H 0 buffered solution (100 mM, pH 9.4). These two buffer solutions had the capability to maintain a constant pH throughout the experiments. All samples were placed in a 37 °C water bath shaking at 30 rpm. At predetermined time intervals, one test tube in each sample group was taken out of the water bath and processed following the same procedures described in the section of "Investigation of Composition Effect on Biodegradation of the LGA Oligomers". 2

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Results and Discussion Composition Effect on Oligomer Biodegradation. The changeable biodegradation rate by changing the composition is one of the advantages of LGA polymers/oligomers. The ratio of lactic to glycolic acid moiety determines the hydrophobicity of the LGA polymers/oligomers (16). Lactic acid moiety is more hydrophobic than glycolic acid moiety because of the pendant methyl group in the lactic acid residue. The hydrophobicity controls the water accessibility to the LGA polymer/oligomer matrix and further influences the biodegradation of the LGA polymers/oligomers. By changing the composition of LGA polymers/oligomers, desirable biodegradation duration can be obtained, which can be used to control the release time of a drug delivery system made from an LGA polymer/oligomer. This programmable biodegradation characteristic completely applies to LGA oligomers. Although an LGA oligomer is more hydrophilic than its high molecular weight counterpart, the composition effect on the biodegradation is still clearly observed as shown in Figure 1. Figure 1 shows the weight loss of the LGA oligomers having varying compositions versus time. As shown in the figure, the weight loss of all three oligomers has a linear dependence on time after a



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hydration period. The rate of weight loss can be observed from the slope of the linear part of the data points. As one can see, the rate of weight loss is influenced by the composition of the LGA oligomer. An increase of glycolic acid residue in the oligomers accelerates the rate of weight loss. The LGA046/54 has the highest rate of weight loss while the LGA072/28 has the lowest rate of weight loss. This property can be used to control the drug release duration of a drug delivery system prepared from the LGA oligomers. Molecular Weight Effect on Oligomer Biodegradation. The molecular weight of LGA polymers/oligomers affects their biodegradation mainly through influencing their hydrophobicity (16). The molecular weight of LGA polymers/oligomers determines the number of carboxylic end groups in the polymer/oligomer matrix. For the same amount of material, a low molecular weight LGA polymer/oligomer contains more carboxylic end groups per unit weight than a high molecular weight counterpart. Therefore, a low molecular weight LGA polymer/oligomer is more hydrophilic than its high molecular weight counterpart and has a faster biodegradation rate. Figure 2 shows the weight loss of two LGA oligomers having the same composition but different molecular weights. These two oligomers have a similar hydration period of 6 days. But the oligomer having high molecular weight has lower biodegradation rate than that having lower molecular weight. In addition to the hydrophilicity of the LGA oligomers, another parameter frequently influenced by molecular weight is glass transition temperature (T ). The T of high molecular weight LGA polymers is usually higher than the physiological temperature, i.e. 37 °C (17). However, the T of LGA oligomers is lower than the physiological temperature (15). At 37 °C, an LGA oligomer is in a rubbery state and can absorb much more water than a LGA polymer in a glassy state, which may be one reason that LGA oligomers biodegrade much faster than high molecular weight LGA polymers (17). g

g

pH Effect on Oligomer Biodegradation. It is reported that LGA polymers/oligomers degrade in an aqueous medium following the mechanism of ester hydrolysis (14,16). The hydrolysis of a polymer/oligomer increases the number of carboxylic end groups and decreases the molecular weight of the polymer/oligomer. The polymer/oligomer eventually degrades to free acids, lactic and/or glycolic acid. As the number of the carboxylic end groups increases, protons or hydronium ions are released to the incubating medium and, if the incubating medium is not buffered, the pH of the incubating medium is lowered. During the biodegradation of the LGA oligomers, this lowering of pH of the incubating medium can be clearly observed. Figure 3 shows the pH change of the incubating medium with time for the three oligomers (LGA046/54, LGA065/35, and LGA072/28). As shown in the figure, the pH of the incubating medium drops rapidly with time and finally reaches the value of approximately 2. This quick pH drop of the incubating medium suggests that a large amount of protons or hydronium ions have been produced during the biodegradation of the oligomers, which suggests that the LGA oligomers have a fast biodegradation rate. In contrast to the experiment shown in Figure 3, if the pH of the incubating medium is controlled or the incubating medium is buffered, one can see the effect of pH In Tailored Polymeric Materials for Controlled Delivery Systems; McCulloch, I., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1998.

g


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Figure 3 Change of pH of the incubating medium during the biodegradation of the LGA oligomers of three different compositions.



249 of the incubating medium on biodegradation of the LGA oligomers (Figure 4). Figure 4 shows that the biodegradation of the oligomer is faster at pH 9.4 than at pH 7.4. The percent weight loss of the oligomer in the basic medium (pH 9.4) is 56% after 14 days of incubation, while that of the same oligomer in the neutral medium (pH 7.4) is only 42%. The pH influence on biodegradation of LGA polymers has been studied by some researchers (6, 10, 11, 18, 19). The hydrolysis of LGA polymers/oligomers can be catalyzed by an acid or a base as shown in Figure 5. In the basic pH range (pH 7.49.4), the base catalysis may be dominant. It is known that the hydrolysis of a carboxylic ester is an equilibrium reaction (20). Therefore, a basic incubating medium favors shifting the reaction of the biodegradation of LGA polymers/oligomers (Figure 5) towards the right because one of the hydrolysis products of the LGA polymers/oligomers hasfreecarboxylic group. Thefreecarboxylic group is neutralized by the alkaline buffer. The pH 9.4 buffer can neutralize more free carboxylic groups than the pH 7.4 buffer. Therefore, the biodegradation of the LGA oligomers in pH 9.4 buffer is faster than in pH 7.4 buffer. Proposed Mechanism for Oligomer Biodegradation. A lag time can be observed in the weight loss of the LGA oligomers as shown in Figure 2 and 4. Significant weight loss starts 6 days after the incubation. After the 6-day lag time, a massive weight loss starts, having a linear relationship to the incubating time. This biodegradation behavior suggests a different biodegradation mechanism for the LGA oligomers from their high molecular weight counterpart. The biodegradation of LGA polymers follows a fourstep process: hydration, initial degradation, further degradation and solubilization (16). For the LGA oligomers, only three biodegradation steps were observed as schemed in Figure 6. The first step is the hydration of LGA oligomers. In this period, the LGA oligomer matrix absorbs water and swells. There is no weight loss occurring but the pH of the incubating medium starts declining due to the release of protons from the carboxylic end group of the oligomer chain in the oligomeric matrix to the incubating medium (as shown in Figure 3 between 0-2 days). After the hydration period, the biodegradation starts all over the hydrated matrix, which is believed to undergo bulk degradation. This period is characterized by a rapid decline of the pH of the incubating medium and a slight weight loss of the oligomer matrix (as shown in Figure 3 between 2-6 days). After this bulk degradation period, a massive weight loss period takes over and follows a linear dependency with respect to incubating time. This linear weight loss period may suggest a surface erosion-controlled solubilization mechanism. This solubilization mechanism may be speculated as follows. After the bulk degradation period, the pH inside the oligomer matrix may have approached to an acidic value. This acidic micro-environment may retard the hydrolysis or biodegradation inside the oligomeric matrix. The surface of the oligomeric matrix is still exposed to the buffered incubating medium that has a basic pH. Therefore, the matrix surface undergoes biodegradation at a much faster rate than the matrix core. This difference of biodegradation rate between the matrix surface and the matrix core brings about a surface erosion-controlled solubilization of the LGA oligomer after the stage of bulk degradation. The proposed surface erosion-controlled solubilization mechanism may be characterized by a slower rate of pH decrease after 8 days of incubation (see Figure 3). This surface erosion results in a linear relationship of weight loss with time. In Tailored Polymeric Materials for Controlled Delivery Systems; McCulloch, I., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1998.


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R O I II -CH-C-0

R OH I II -CH-C-O-

R

O

E

-CH-C-0-OH

HO-+H*

(a)

R O O/'h-OH ? H I II -CH-C + 0 • - C H - C + HO- + O H

G

9

OH

E

(b)

OH

Figure 5 Hypothetical mechanisms of hydrolysis of LGA polymers/oligomers. (a) at pH < 7; (b) at pH > 7.

HYDRATION

MATRIX B U L K DEGRADATION

SURFACE EROSION-CONTROLLED SOLUBILIZATION

Figure 6. Scheme of the three steps proposed for the biodegradation of LGA oligomers.


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Conclusions The factors affecting the biodegradation of the LGA oligomers have been separately investigated. The experimental data show that the biodegradation of LGA oligomers is influenced by the composition and the molecular weight of the LGA oligomers, and the pH of the incubating medium. A higher content of glycolic acid residue in an LGA oligomer results in a higher biodegradation rate of the oligomer. In a constant pH medium and for the oligomers having the same composition, the weight loss of a higher molecular weight LGA oligomer is slower than that of a lower molecular weight counterpart. The weight loss of the LGA oligomers is faster in an alkaline incubating medium (pH 9.4) than that in a neutral incubating medium (pH 7.4). When distilled water is used as an incubating medium, the pH of the incubating medium drops rapidly along with the biodegradation of the LGA oligomers. A three-step biodegradation mechanism is proposed for the LGA oligomers. These three steps are hydration, matrix bulk degradation, and surface erosion-controlled solubilization. The controllable biodegradation properties of the LGA oligomers have useful applications to controlledrelease drug delivery. Literature Cited 1. Park, T. G. J. Controlled Release 1994, 30, 161-173. 2. Visscher, G.E.; Robison, R.L.; Maulding, H.V.; Fong, J.W.; Pearson, J.E. and Argentieri, G.J. J.Biomed.Mater. Res. 1986, 20, 667-676. 3. Bodmer, D.; Kissel, T. and Traechslin, E. J. Controlled Release 1992, 21, 129-138. 4. Spenlehauer, G.; Vert, M.; Benoit, J.P. and Boddaert, A. Biomaterials 1989, 10, 557-563. 5 Wang,. H.T.; Palmer, H.; Linhardt, R.J.; Flanagan, D.R. and Schmitt, E. Biomaterials 1990, 11, 679-685. 6. Ginde, R.M. and Gupta, R.K. J. Appl. Polym. Sci. 1987, 33, 2411-2429. 7. Zhu, J.H.; Shen, Z.R.; Wu, L.T. and Yang, S.L. J. Appl. Polym. Sci. 1991, 43, 2099-2106. 8. Thies, C. and Bissery, M.C. In Biomedical Applications of Microencapsulation; Lim, F. Ed.; CRC Press: Boca Raton, FL, 1984, pp. 53-74. 9. Kaetsu, I.; Yoshida, M.; Asano, M.; Yamanaka, H.; Imai, K.; Yuasa, H.; Mashimo, T.; Suzuki, K.; Katakai, R. and Oya, M. J. Controlled Release 1987, 6, 249-263. 10. Asano, M.; Fukuzaki, H.; Yoshida, M.; Kumakura, M.; Mashimo, T.; Yuasa, H.; Imai, K. and Yamanaka, H. Drug Design and Delivery 1990, 5, 301-320. 11. Huffman, K.R. and Casey, D.J. J. Polym. Sci. Polym.: Chem. Ed. 1985, 23, 19391954. 12. Chu, C.C. J.Biomed.Mater. Res. 1981, 15, 795-804. 13. Makino, K.; Ohshima, H. and Kondo, T. J. Microencapsulation 1986, 3, 203-212. 14. Chu, C.C. J. Biomed. Mater. Res. 1981, 15, 19-27. 15. Wang, N.; Wu, X.S.; Upton, H.L.; Donahue, E. and Siddiqui, A. J. Biomat. Sci.: Polym. Ed., 1997, 8, 905-917. 16. Wu, X.S. In Encyclopedic Handbook of Biomaterials and Bioengineering, Part A: Materials, Wise, D.L.; Trantolo, D. J.; Altobelli, D. E.; Yaszemski, M.J.; Gresser, J. D. and Schwartz, E. R., Ed.; Marcel Dekker: New York, NY, 1995, pp. 1015-1054. 17. Vert, M., Li, S. and Garreau, H., J. Controlled Release 1991, 16, 15-26.


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18. Kishida, A.; Yoshioka, S.; Takeda, Y. and Uchiyama, M. Chem.Pharm.Bull. 1989, 37, 1954-1956. 19. Williams, D. F. J. Biomed. Mater. Res. 1980, 14, 329-338. 20. March, J. Advanced Organic Chemistry, Fourth Edition, John Wiley & Sons: New York, NY, 1992; p. 378.


Biodegradation of Lactic-Glycolic Acid Oligomers - ACS Symposium

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