Biodegradation of Synthetic Polymers Containing Ester Bonds

Chapter 12

Downloaded by UNIV OF CALIFORNIA SAN DIEGO on January 5, 2016 | http://pubs.acs.org Publication Date: July 13, 1990 | doi: 10.1021/bk-1990-0433.ch012

Biodegradation of Synthetic Polymers Containing Ester Bonds Yutaka Tokiwa, Tadanao Ando, Tomoo Suzuki, and Kiyoshi Takeda Fermentation Research Institute, 1-1-3, Higashi, Tsukuba, Ibaraki, Japan 305 Oligomers of synthetic polymers are biodegradable, whereas only few polymers are biodegradable. Polyethylene adipate with Mn 3000 and polycaprolactone with Mn 25,000 were completely degraded by Penicillium spp. Aliphatic and alicyclic polyesters, ester type polyurethanes (I), copolyamide-esters (II) and copolyesters (III) of aliphatic and aromatic polyesters were hydrolyzed by lipases from various microorganisms and hog pancreas, and hog liver esterase. When aliphatic polyesters were used as enzyme substrates, it was found that their melting points (Tm) had a effect, in addition to their chemical structure, on biodegradability. It was assumed that the hydrogen bonds in the I, II chains and aromatic ring in the III chains, which were related to high Tm of I, II and III, influenced their biodegradation by lipases. A v a r i e t y of low-molecular-weight a r t i f i c i a l synthetic compounds are biodegradable, whereas only few synthetic polymers are biodegradable. Among synthetic polymers, a l i p h a t i c polyesters are generally known to be susceptible to b i o l o g i c a l attack (1-5). We report here that polyethylene adipate (PEA) and polycaprolactone (PCL) were degraded by Pénicillium spp., and a l i p h a t i c and a l i c y c l i c polyesters,ester type polyurethanes, copolyesters composed of a l i p h a t i c and aromatic polyester (CPE) and copolyamide-esters (CPAE) were hydrolyzed by several lipases and an esterase. Concerning these water-insoluble condensation polymers, we noted that the melting points (Tm) had a effect on biodegradability. Materials and Methods Materials. PCL and polypropiolactone (PPL) were prepared by r i n g opening polymerization of ε -caprolactone (&.) and $ -propiolactone respectively i n benzene i n a nitrogen atmosphere at 60 °C with a d i -

0097-^156/90/0433-0136$06.00/0 © 1990 American Chemical Society

In Agricultural and Synthetic Polymers; Glass, J. Edward, et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1990.


12. TOKIWAETAL.

Biodégradation of Synthetic Polymers with Ester Bonds

ethylzinc-water catalyst system. Poly-DL- 3 -methylpropiolactone (Mn: 8 1 9 0 ; Tm:16T-1T1 °C; poly-DL- 3 -hydroxybutyrate) was made from DL3 - methylpropiolactone by the method of Yamashita et al.(j.) with a triethylaluminum-water catalyst system. Poly-D- 3 - methylpropiolactone (PHB) produced by Alcaligenes eutrophus was kindly supplied by ICI. PCL-diols (Mn: 530, 1250, 2000, 3000) were purchased from A l d r i c h Chemical. Other saturated a l i p h a t i c polyesters were synthesized by a melt polycondensation technique (8.) > and unsaturated polyesters were synthesized by high temperature solution polycondensation (9.). A l l a l i c y c l i c and aromatic polyesters were from Nihon Chromato except polyethylene terephthalate (PET) from Asahikasei, p o l y t e t r a methylene terephthalate (PBT) from Union Carbide, copolyester of PET and polycyclohexylenedimethyl succinate (PETG) from Eastman Chemical Products. A l l diisocyanates were from Tokyo Chemical. Polyurethanes were synthesized by a solution polycondensation method (10). CPE and CPAE were synthesized by the t r a n s e s t e r i f i c a t i o n reaction ( l l ) and the amide-ester interchange reaction (12) respectively. A l l nylons were from A l d r i c h Chemical except nylon 6 ( (η) =1.30 d l / g i n m-cresol at 2 5 °C) from Toyokasei and nylon 11 and nylon 12 from Nihon L i l s a n . Achromobacter sp. and Candida cylindracea lipases (Meito Sangyo) were p u r i f i e d by g e l f i l t r a t i o n on Sephadex G-100 (2.6 χ 7 9 cm) from crude preparations. U l t r a c e n t r i f u g a l l y homogeneous preparations of Geotrichum candidum and Rhizopus delemar lipases were from Seikagaku Kogyo and p a r t i a l l y p u r i f i e d preparations of arrhizus lipase and hog l i v e r esterase were from Boehringer Mannheim Yamanouchi. A par t i a l l y p u r i f i e d preparation of hog pancreas lipase was obtained from Worthington Biochemicals. One unit of enzyme l i b e r a t e d one μ mole of f a t t y acid from o l i v e o i l per min at pH 7.0 and 37 °C. Culture. Fungi were cultured on a rotary shaker at l80 rpm at 30 °C. The composition of cuture medium was as follows; PEA (or PCL), 1.0 g; (NH ) S0i , 1.0 g; K^PO^, 0.2 g; I^HPO^, 1.6 g; MgS0^7H 0, 0.2 g; NaCl, 0.1 g; CaCl -2H 0, 0.02 g; FeSO^-71^0, 0.01 g; Na^oO^·2H 0, 0.5 mg; Na^O^ · 2H 0, 0.5 mg; MnSO^, 0.5 mg i n one l i t e r of d i s t i l l e d water. pH was adjusted to 7.2. After s t e r i l i z a t i o n , PEA (or PCL) i n the culture medium was dispersed by shaking. The p a r t i c l e size of PEA and PCL i n the medium was about 0.1-3 mm, 1-5 mm respectively. i|

2

|

2

2

2

2

2

Assay of Enzymatic Hydrolysis of Synthetic S o l i d Polymers. Hydrol y s i s of s o l i d polymers was measured by the rate of t h e i r s o l u b i l i z a t i o n , and the measurement process does not necessarily involve com plete hydrolysis into the constituent parts. The rate was determined by measuring the water-soluble t o t a l organic carbon (T0C) concentra t i o n at 30 °C i n the reaction mixture using a Beckman T0C analyzer (Model 915-B). In the substrate and enzyme controls, enzyme or sub strate was omitted from the reaction mixture. Determination of Molecular Weight. The number average molecular weight (Mn) was measured by the vapor pressure equilibrium method with a Hitachi Molecular Weight apparatus (Model 117). Measurement of Melting Point. Melting points of polyesters (Tm) were measured using a Yanaco Micro Melting Point apparatus (Model MP-S3).


137

AGRICULTURAL AND SYNTHETIC POLYMERS

138


D e t e r m i n a t i o n o f C a r b o x y l i c Group T e r m i n a l s . The c a r b o x y l i c group t e r m i n a l s were determined by a l k a l i t i t r a t i o n (13.). A f t e r 200 mg o f the sample was d i s s o l v e d i n 7.5 ml o f b e n z y l a l c o h o l i n a n i t r o g e n atmosphere at 110 °C, the s o l u t i o n was t i t r a t e d w i t h 0.05N potas^'urn h y d r o x i d e u s i n g p h e n o l p h t h a l e i n as an i n d i c a t o r . A b l a n k t e s t wab c a r r i e d out by t h e same method but o m i t t i n g t h e sample. E s t i m a t i o n o f M o l e c u l a r Weight D i s t r i b u t i o n o f Polyamide B l o c k s . The m o l e c u l a r weight d i s t r i b u t i o n o f t h e polyamide b l o c k s was e s t i m a t e d by g e l permeation chromatography (GPC) u s i n g two i n s t r u m e n t s , model HLC-802R (Toyo Soda I n d u s t r y Co., L t d . ) and model GPC-2U1+ (Waters A s s o c i a t e s , I n c . ) . P o l y s t y r e n e and n y l o n o l i g o m e r were used as s t a n d ards. R e s u l t s and D i s c u s s i o n D e g r a d a t i o n o f A l i p h a t i c P o l y e s t e r s by F u n g i . P o l y e t h y l e n e a d i p a t e (PEA) w i t h Wi 3000 was almost c o m p l e t e l y degraded by Pénicillium sp. s t r a i n l U - 3 i s o l a t e d from s o i l o f a f a c t o r y p r o d u c i n g p o l y u r e t h a n e ( F i g u r e l ) . As s t r a i n l U - 3 grew, PEA was r a p i d l y degraded and d i s a p peared i n 120 hours. As shown i n F i g u r e 1, w a t e r - s o l u b l e TOC were found i n t h e c u l t u r e medium. A d i p i c a c i d and e t h y l e n e g y l c o l were d e t e c t e d i n t h e c u l t u r e f l u i d as d e g r a d a t i o n p r o d u c t s . Furthermore the s t r a i n a s s i m i l a t e d some o t h e r a l i p h a t i c p o l y e s t e r s . F i g u r e 2 i l l u s t r a t e s t h e t i m e course o f p o l y c a p r o l a c t o n e (PCL; Mn 2 5 - 0 0 0 ) d e g r a d a t i o n by Pénicillium sp. s t r a i n 2 6 - 1 i s o l a t e d from s o i l . The i s o l a t e almost c o m p l e t e l y degraded PCL i n 12 days. Among d e g r a d a t i o n p r o d u c t s o f PCL, ε- h y d r o x y c a p r o i c a c i d was d e t e c t e d . The fungus a s s i m i l a t e d v a r i o u s p o l y e s t e r s . I n g e n e r a l , a s s i m i l a t i o n o f a l i p h a t i c p o l y e s t e r s by t h e fungus was b e t t e r the g r e a t e r the num b e r o f carbon atoms between t h e e s t e r bonds. P o l y e s t e r s w i t h s i d e c h a i n s were g e n e r a l l y l e s s a s s i m i l a t e d t h a n w i t h o u t s i d e c h a i n s . The fungus a l s o a s s i m i l a t e d u n s a t u r a t e d a l i p h a t i c p o l y e s t e r s , but h a r d l y a s s i m i l a t e d a l i c y c l i c and a r o m a t i c p o l y e s t e r s . P o l y e s t e r - D e g r a d i n g Enzyme. A p o l y e s t e r - d e g r a d i n g enzyme from Pénic i l l i u m sp. s t r a i n 1^-3 was p u r i f i e d i n t o a homogeneous s t a t e u l t r a c e n t r i f u g a l l y and e l e c t r o p h o r e t i c a l l y (ih). I n s u b s t r a t e s p e c i f i c i t y , t h i s enzyme degraded v a r i o u s k i n d s o f s a t u r a t e d and u n s a t u r a t e d a l i p h a t i c p o l y e s t e r s and p o l y c y c l o h e x y l e n e d i m e t h y l a d i p a t e as a l i c y c l i c p o l y e s t e r but not a r o m a t i c p o l y e s t e r s . A k i n d o f t e r m i n a l groups, such as h y d r o x y , h e x a h y d r o p h t h a l i c a c i d and h e x a h y d r o p h t h a l i c a c i d g l y c i d i l e s t e r t e r m i n a l d i d not a f f e c t so much on t h e enzyme a c t i v i t y . The enzyme f u r t h e r h y d r o l y z e d v a r i o u s p l a n t o i l s , t r i g l y c e r i d e s and m e t h y l e s t e r s o f f a t t y a c i d . T h e r e f o r e t h e p o l y e s t e r - d e g r a d i n g enzyme was assumed t o be a k i n d o f l i p a s e . H y d r o l y s i s o f P o l y e s t e r by L i p a s e . A l i p h a t i c p o l y e s t e r , PEA and PCL were h y d r o l y z e d by l i p a s e s from Achromobacter s p . , C. c y l i n d r a c e a , G. candidum, R. a r r h i z u s , R. delemar, hog pancreas and hog l i v e r e s t e r a s e ( 1 5 ) . E s p e c i a l l y R. a r r h i z u s and R. delemar l i p a s e s were found capable o f h y d r o l y z i n g v a r i o u s k i n d s o f p o l y e s t e r s (Table I ) .




12. TOKIWA ET AL.

0

100 200 C u l t u r e t i m e (h)

300

F i g u r e 2. Time course o f PCL d e g r a d a t i o n by s t r a i n 2 6 - 1 . The s t r a i n was c u l t u r e d a t 30 °C. Growth ( O ) , PCL ( · ) , w a t e r - s o l u b l e T0C ( Δ ) and pH ( • ) a r e shown.


139

AGRICULTURAL A N D SYNTHETIC POLYMERS

140

Table I. Hydrolysis o f Polyesters by R. delemar and R. arrhizus Lipases


Polyester Polyethylene adipate Polyethylene suberate Polyethylene azelate Polyethylene sebacate Polyethylene decamethylate Polytetramethylene succinate Polytetramethylene adipate Polytetramethylene sebacate Polyhexamethylene sebacate Poly-2,2-dimethyltrimethylene succinate Poly-2,2-dimethyltrimethylene adipate Polyglycolide Copolyester o f g l y c o l i d e and lactide(molar r a t i o , 92:8) Polypropiolactone Poly-DL- 3-methylpropiolactone Poly-D - B-methylpropiolactone (PHB) Polycaprolactone (PCL) Poly-cis-2-butene adipate Poly-cis-2-butene sebacate Poly-trans-2-butene sebacate Poly-2-butyne sebacate Polyhexamethylene fumarate Poly-c i s-2-butene fumarate Polytetramethylcyclobutane succinate Polycyclohexylenedimethyl succinate Polycyclohexylenedimethyl adipate Polytetramethylene terephthalate Polyethylene tetrachlorophthalate Poly-2,2-dimethyltrimethylene isophthalate Poly-p-hydroxybenzoate Poly-3,5-dimethyl-p-hydroxybenzoate Poly-U-ethoxy-3,5-dimethylbenzoate

Mn

Tm (°C)

2720 U05U U510 1570 I6l0

U8.5 6U.5 52.1 7*+.5 86.0 117 72.0 65.8 7u:o 76.5

k2k0

1790 2kh0

5820 2370 2020

TOC formed by l i p a s e s (ppm) Powder size R. delemar R.arrhi:

836Ο 1020 3080 550 180 150 3360 980 38Ο 2U0

9290 I62O 3770 98Ο 2U0 210 2900 3300 ll60 50

22k0

10 0 310 58Ο

150 0 0 l600 60 0 3610 550

300

3^30

1190 910 0

Ε

3U0 67Ο 35 30 0

C Β Β A Β A Β A Β A

3UU0

36.5 226-23U 200-210 95.0 167-171 175-181 59.0 56.9-59.8 60.8-62.5 57.0-59.0 61.9-63.0 113-117 300 63.5-8U.O

3910

123-130

Β

130

120

3250

108-llU

Β

200

160

230-2U0

C

0

0

I67O 78.0-8U.2

A

0

0

66.5-75.0 300 300

A

0

A

0 0 0

259-267

A

0

U270 8190 25000 67UO 2700 6190 3560 U930

Ε Ε Ε A Ε A A C Β A C A Β

A

hQ

0

The p a r t i c l e size o f each polyester powder was ranked A, B, C, D, or E, c o r r e sponding r e s p e c t i v e l y t o roughly l e s s than 0.25 mm, l e s s than 0.50 mm, l e s s than 1.0 mm, 0.25-1.5 mm, 0.25-3 mm. Each r e a c t i o n mixture contained kOO u mol of phosphate b u f f e r (pH 7.0), 1 mg o f surfactant P l y s u r f A210G, 300 mg o f the polyester powder and 60 μ g o f R. arrhizus l i p a s e (or 300 μg o f R. delemar l i p a s e ) i n a t o t a l volume of 10.0 ml. In the case of R. delemar l i p a s e , sur factant was omitted and pH of phosphate b u f f e r was 6.0. In the substrate and enzyme c o n t r o l s , enzyme or substrate was omitted from the r e a c t i o n mixture. The r e a c t i o n mixtures were incubated at 30 °C f o r l 6 hours.


12. TOKIWA ET AL.



E f f e c t of the P a r t i c l e Size of PEA Powders on the Hydrolysis by R. delemar Lipase. In the case of PEA, small p a r t i c l e s were hydrolyzed better than large ones as shown i n Figure 3. So i t was assumed that the enzymatic hydrolysis depends on the amounts of surface area of polyester powders. Effect of Molecular Weight of Polyester on the Hydrolysis by Rhizopus l i p a s e . Using three kinds of polyesters, PCL-diol ( I ) , polyhexamethylene adipate ( I I ) , and a copolyester (III) made from 1,6-hexamethylenediol and a 70:30 molar r a t i o mixture of ε- caprolactone and adipic a c i d , the e f f e c t s of the Mn_ of polyester on the hydrolysis by l i p a s e were examined (Figure k). Mn did not a f f e c t the rates of hydrolysis by R. arrhizus and R^ delemar lipases when Mh was more than about UOOO. This would indicate these lipases randomly s p i l t s ester bonds i n p o l mer chains. In contrast, when Mh was less than about k000^ the rates of the enzymatic hydrolysis were faster with the smaller Mn of poly esters. This corresponded to the fact that Tm was lower with the smaller Mn of polyesters. The rates of hydrolysis of copolyester III by both lipases were much higher than those of homopolymers I and I I . I l l was c r y s t a l l i n e , but showed lower Tm than the homopolymers. This would show that III have l e s s order and more amorphous regions than the homopolymers do. E f f e c t of ε- Caprolactone and Adipic Acid Molar Ratio for Copolyes t e r I I I on the Hydrolysis by R. delemar Lipase. The hydrolysis of various copolymers by R. delemar l i p a s e was examined to see whether there was an optimum chemical structure or not. Mn of those copolyesters was selected from 17^0 to 2220, to diminish the e f f e c t of molec u l a r weight. Optimum molar r a t i o of ε- caprolactone and adipic a c i d was about from 90:10 to 70:30 (Figure 5). The Tm at the optimum molar r a t i o was the lowest of a l l . So i t seemed that the existence of o p t i mum molar r a t i o came from the lowest Tm, which would show the most amorphous material, rather than the optimum chemical structure. Relationship Between Tm and the Biodegradability of Polyester by L i pases. The r e l a t i o n s h i p between Tm and the biodegradability of satu rated a l i p h a t i c polyester i s shown i n Figure 6. For the same series polyesters, the b i o d e g r a d a b i l i t i e s decreased with increasing Tm. In general, Tm was represented by the following formula: Tm = ΔΗ/Δ S where ΔΗ i s the change of enthalpy i n melting and Δ S i s the change of entropy i n melting. It i s known that the interactions among poly mer chains mainly a f f e c t the Δ Η value and that the i n t e r n a l r o t a t i o n energies corresponding to the r i g i d i t y ( f l e x i b i l i t y ) of the p o l ymer molecule remarkably a f f e c t the Δ S value. The high Tm of a l i phatic polyamide (nylon) i s caused by the large Δ Η value based on the hydrogen bonds among polymers chains. Nylon i s not biodegradable though nylon oligomer i s biodegradable. On the other hand, the high Tm of aromatic polyester i s caused by the small Δ S value with i n crease i n r i g i d i t y of the polymer molecule based on an aromatic r i n g . Aromatic polyester i s not biodegradable. Hydrolysis of Polyurethanes by Lipase. E f f e c t s of Mn of PCL-diol moiety on the hydrolysis of polyurethanes, which were composed of


141



142

2

6

h

Reaction

8 10 time(h)

F i g u r e 3. E f f e c t o f t h e p a r t i c l e s i z e o f PEA powders on t h e hy d r o l y s i s by R. delemar l i p a s e . R e a c t i o n m i x t u r e s were i n c u b a t e d at 30 °C. P a r t i c l e s i z e : - Ο - , 0-0.25 mm; 0-1.00 mm, - · - ,

0.25-1.00

mm.

-ο 6 0 ^50 1+0 H

h-

H

h

R. a r r h i z u s

2

k

6

8

10 12

M o l e c u l a r weight(Mn) x l O ^ F i g u r e U. E f f e c t s o f m o l e c u l a r weight o f p o l y e s t e r on t h e h y d r o l y s i s by l i p a s e s . Three k i n d s o f p o l y e s t e r s were used: P C L - d i o l ( • ), polyhexamethylene a d i p a t e ( Ο ) and t h e i r c o p o l y e s t e r ( · ). The dashed l i n e shows t h e r e s u l t when one t e n t h enzyme c o n c e n t r a t i o n was used.



12.

TOKIWAETAL.

Biodégradation ofSynthetic Polymers with Ester Bonds

60

0

20 Adipic

kO 6 0 8 0 1 0 0 a c i d {mol%)

F i g u r e 5. E f f e c t o f ε-caprolactone and a d i p i c a c i d molar r a t i o f o r copolymers made from 1,6-hexamethylenediol and a m i x t u r e o f ε - c a p r o l a c t o n e and a d i p i c a c i d on t h e h y d r o l y s i s by R. delemar l i p a s e . Four o r d e r s numbers i n t h i s f i g u r e showed Mn o f each p o l y e s t e r . Β and C i n t h i s f i g u r e showed t h e p a r t i c l e s i z e o f each p o l y e s t e r as same as T a b l e I .


143



144

10 fib

'

»

ft a)

~ 8 Β ft ft

«

r

Γ #1

τ

• PEA

1

1

1

r c)

b)

•PEA

m ο

PEA

PEAz

ο

«Η

ο ο

Eh

D

$èsE Ο

PBA PPL • PHSE PBS

PEPe

120

60

8 0 1 0 0 120 Tm

(°C)

Figure 6. Relationship between Tm and the biodegradability of polyesters by R. delemar (a) and R. arrhizus (b) l i p a s e s , and PEAdegrading enzyme from Pénicillium sp. s t r a i n lk-3 ( c ) . PESu: polyethylene suberate; PEAz: polyethylene azelate; PESE: polyethylene sebacate; PEDe: polyethylene decamethylate; PBS: polytetramethylene succinate; PBA: polytetramethylene adipate; PBSE: p o l y t e t r a methylene sebacate; PHSE: polyhexamethylene sebacate; PPL: polypropiolactone.


12.

TOKIWAETAL.


P C L - d i o l and d i p h e n y l m e t h a n e - U k ' - d i i s o c y a n a t e (MDI), "by R. delemar l i p a s e were examined. These p o l y u r e t h a n e s have b o t h the hydrogen bonds among polymer c h a i n s and aromatic r i n g s i n t h e polymer molec u l e s . R. delemar l i p a s e c o u l d h y d r o l y z e t h e p o l y u r e t h a n e s though t h e r a t e o f h y d r o l y s i s toward p o l y u r e t h a n e s d e c r e a s e d as compared t o t h a t toward P C L - d i o l . The r a t e o f h y d r o l y s i s d e c r e a s e d w i t h d e c r e a s i n g t h e Mn o f PCL-moiety o f p o l y u r e t h a n e s ( F i g u r e 7 ) . The e f f e c t s o f c h e m i c a l s t r u c t u r e o f d i i s o c y a n a t e component on t h e h y d r o l y s i s o f p o l y u r e t h a n e s by R. delemar l i p a s e were examined ( F i g u r e 8 ) . The r a t e s o f h y d r o l y s i s o f t h e p o l y u r e t h a n e s c o n t a i n i n g MDI o r t o l y l e n e - 2 , U - d i i s o c y a n a t e (TDI) were s m a l l e r t h a n t h a t o f t h e p o l y u r e t h a n e c o n t a i n i n g 1 , 6 - h e x a m e t h y l e n e - d i i s o c y a n a t e (HDI). Thus i t was assumed t h a t the r i g i d i t y o f t h e p o l y u r e t h a n e molec u l e s based on t h e aromatic r i n g s , r a t h e r t h a n t h e hydrogen bonds among t h e p o l y u r e t h a n e c h a i n s , would i n f l u e n c e d t h e i r b i o d e g r a d a b i l i t y by R. delemar l i p a s e .


9

H y d r o l y s i s o f C o p o l y e s t e r s (CPEs) C o n t a i n i n g Aromatic and A l i p h a t i c E s t e r B l o c k s by L i p a s e ( l 6 ) . CPEs were s y n t h e s i z e d by t h e t r a n s e s t e r i f i c a t i o n r e a c t i o n between aromatic p o l y e s t e r s (PET, PBT, PETG, PEIP) and a l i p h a t i c p o l y e s t e r (PCL). The s u s c e p t i b i l i t y o f CPEs t o h y d r o l y s i s by R. delemar l i p a s e dropped o f f r a p i d l y d u r i n g t h e i n i t i a l stage o f t h e t r a n s e s t e r i f i c a t i o n r e a c t i o n and i n c r e a s e g r a d u a l l y as the r e a c t i o n proceeded. The s u s c e p t i b i l i t y t o h y d r o l y s i s dec r e a s e d w i t h i n c r e a s e i n a r o m a t i c p o l y e s t e r content ( F i g u r e 9 ) . The s u s c e p t i b i l i t y t o h y d r o l y s i s by t h e l i p a s e o f CPEs composed o f PCL and p o l y e t h y l e n e i s o p h t h a l a t e ( P E I P ) , t h e l a t t e r b e i n g used as a low Tm ( 1 0 3 °C) a r o m a t i c p o l y e s t e r , were g r e a t e r t h a n t h o s e o f o t h e r CPEs as shown i n F i g u r e 9. I t was assumed t h a t t h e r i g i d i t y o f t h e aromati c r i n g i n t h e CPE c h a i n s i n f l u e n c e d t h e i r b i o d e g r a d a b i l i t y by t h i s lipase. H y d r o l y s i s o f C o p o l y a m i d e - e s t e r s (CPAEs) by L i p a s e ( 1 £ ) . CPAEs were s y n t h e s i z e d by t h e a m i d e - e s t e r i n t e r c h a n g e r e a c t i o n between polyamide and p o l y e s t e r . The l e n g t h o f the polyamide b l o c k s was measured a f t e r h y d r o l y s i s o f e s t e r bonds i n CPAE by a l k a l i at 30 °C. The i n f r a r e d s p e c t r a a f t e r h y d r o l y z i n g e s t e r bonds on CPAEs showed t h a t t h e e s t e r bonds were almost c o m p l e t e l y removed. The m o l e c u l a r weight d i s t r i b u t i o n o f polyamide b l o c k s was examined by GPC (Table I I ) . The f o l l o w ing samples were used: CPAE-1 ( r e a c t i o n t i m e f o r s y n t h e s i s , 1 h r ) and CPAE-2 ( r e a c t i o n t i m e , h h r ) composed o f n y l o n 6 and PCL a t a 50/50 molar r a t i o , CPAE-3 ( r e a c t i o n t i m e , 1 h r ) and CPAE-U ( r e a c t i o n t i m e , h h r ) composed o f n y l o n 12 and PCL a t a 50/50 molar r a t i o , CPAE-5 ( r e a c t i o n t i m e , k h r ) composed o f n y l o n 6 and PCL at a 20/80 molar r a t i o , and CPAE-6 ( r e a c t i o n t i m e , h h r ) composed o f n y l o n 12 and PCL at a 20/80 molar r a t i o . I n a d d i t i o n , t h e Mn o f polyamide b l o c k s o f Table I I .

P o l y m e r i z a t i o n Degree o f t h e Main Component o f t h e Polyamide B l o c k s o f CPAEs 1-6

S t a n d a r d f o r GPC Polystyrene Each n y l o n o l i g o m e r

CPAE-1 CPAE-2 CPAE-3 CPAE-1+ CPAE-5 CPAE-6 9k 32 16 53

9-10

2-6

7-8


2-U

145

146



200

2 k 6 Reaction time (h)

0

F i g u r e 7. E f f e c t s o f m o l e c u l a r weight o f P C L - d i o l p a r t s on t h e h y d r o l y s i s o f p o l y u r e t h a n e s by R. delemar l i p a s e . Each r e a c t i o n m i x t u r e f o r b i o d e g r a d a b i l i t y assay c o n t a i n e d 15.6-37.2 mg o f p o l yurethane f i l m (lH.l+-27.1 mg as p o l y e s t e r m o i e t y ) on t h e c o v e r g l a s s (3.2 cm^) i n a t o t a l volume o f 10 ml. I n t h i s _ c o n d i t i o n , no e f f e c t o f amount o f p o l y u r e t h a n e was observed. (Mn)s o f PCLd i o l s p a r t s o f p o l y u r e t h a n e s I , I I , I I I and I V were 530, 1250, 2000, 3000 r e s p e c t i v e l y . The dashed l i n e s show P C L - d i o l (Mn 2000) ( • ) and P C L - d i o l (Mn 3000) ( · ).

A0-PÇL-0C0NH-R-NH0C-4 -0n

100

h 2000

-(CH ) 2

6

50

1 + 6

8

Reaction time (h) F i g u r e 8. E f f e c t s o f c h e m i c a l s t r u c t u r e o f d i i s o c y a n a t e component on t h e h y d r o l y s i s o f p o l y u r e t h a n e s by R. delemar l i p a s e . HDI ( · ) , TDI ( • ) o r MDI ( A ) was_used as a d i i s o c y a n a t e o f t h e p o l y u r e t h a n e c o n t a i n i n g P C L - d i o l (Mn 2000). Assay c o n d i t i o n s a r e t h e same as i n t h e case o f F i g u r e 7.


12. TOKIWA ET AL.

Biodégradation of Synthetic Polymers with Ester Bonds 147 —τ

100

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1

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I

80

Γ

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1

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20

0

1

2 0 1+0 60 0 2 0 1+0 60 PETG (mol?)

ΡΒΤ (mol?)

0 20

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60

PEIP (mol?)

F i g u r e 9. E f f e c t o f molar r a t i o o f PCL and a r o m a t i c p o l y e s t e r on t h e b i o d e g r a d a b i l i t y o f CPE by R. dememar l i p a s e , ( a ) , ( b ) , and (c) i n d i c a t e PCL-PETG, PCL-PBT, and PCL-PEIP systems, r e s p e c t i v e l y . Each r e a c t i o n m i x t u r e f o r b i o d e g r a d a b i l i t y assay c o n t a i n e d CPE powder o r i t s f i l m s ( 2 0 mg as p o l y e s t e r m o i e t y ) i n a t o t a l volume o f 1.0 m l . R e a c t i o n m i x t u r e s were i n c u b a t e d a t 37 °C f o r 16 h o u r s . F o r m a t i o n o f t h e w a t e r - s o l u b l e T0C was i n p r o p o r t i o n t o s u b s t r a t e amounts (up t o 50 mg as PCL m o i e t y ) i n t h i s r e a c t i o n system. (Reproduced from Reference 1 6 . C o p y r i g h t 1 9 8 1 John Wiley. )

20 1+0 Nylon

(mol?)

F i g u r e 1 0 . E f f e c t o f molar r a t i o o f PCL and n y l o n on t h e b i o d e g r a d a b i l i t y o f CPAE by R. delemar l i p a s e . The r e a c t i o n time f o r each CPAE s y n t h e s i s was 1+ h o u r s . The b a s i c s t r u c t u r e s o f n y l o n were o f two t y p e s . One was -{NH(CH ) C0J ( l e f t ) ; t h e o t h e r was -jNH(CH )6NHC0(CH ) C0j- ( r i g h t ) . L e f t : n y l o n 6 (O ); n y l o n 1 1 ( A ) ; n y l o n 12 (• ) ; r i g h t : n y l o n 6,6 (O ); n y l o n 6,9 ( ) > n y l o n 6,12 ( • ) . Assay c o n d i t i o n s a r e t h e same as i n t h e case o f F i g u r e 9 except CPAE was used i n s t e a d o f CPE. (Reproduced from R e f e r ence 1 7 . C o p y r i g h t 1 9 7 9 John W i l e y & Sons,, I n c . ) 2

2

2

n

n

m

m

Δ

American Chemical Society Library 1155 15th St., N.W.

In Agricultural and Synthetic Polymers; Glass, J. Edward, et al.; Washington, DJG. 20036 ACS Symposium Series; American Chemical Society: Washington, DC, 1990.



148

CPAEs 1-6, as estimated by end-group assay, were 3750, 1030, 2 Ô 7 0 , 8 5 Ο , 8 3 0 , and 4 8 0 , respectively. Nylon oligomers with low molecular weight are expected to be biodegradable. The s u s c e p t i b i l i t y of CPAEs t o hydrolysis by R. delemar lipase decreased with the shortening of the polyamide blocks and with i n creasing polyamide content (Figure 1 0 ) . The simple blends of nylon and PCL at 2 7 0 °C f o r 1 0 min retained high biodegradability of PCL. So i t was assumed that the amount and d i s t r i b u t i o n of hydrogen bonds, based on the amide bonds, i n the CPAE chains influenced t h e i r b i o degradability by t h i s l i p a s e . The new biodegradable synthetic polymer, CPAE can be formed into any desirable shape. A transparent t h i n f i l m (about 0.02 mm t h i c k ness) was made from CPAE. It would be very important that various types of interactions among macromolecular chains, which are related to Tm, are taken into consideration when designing the biodegradable s o l i d polymers.

Literature Cited 1. 2.

Darby, R. T.; Kaplan, A. M. Appl. Microbiol. 1968, 16, 900. Potts, J. E.; Clendinning, R. Α.; Ackart, W. B.; Niegisch, W.D. Am. Chem. Soc., Polymer Preprints 1972, 1 3 , 629. 3. Fields, R. D.; Rodriguez, F.; Finn, R. K. J. Appl. Polym. Sci. 1974, 1 8 , 3571. 4. Diamond, M. J.; Freedman, B.; Garibaldi, J. A. Int. Biodetn. Bull. 1975, 11, 127. 5. Huang, S. J . ; Bell, J. P.; Knox, J. R.; Atwood, H.; Bansleben, D.; Bitritto, M.; Borghard, W.; Chapin, T.; Leong, K. W.; Natarjan, K.; Nepumuceno, J.; Roby, M.; Soboslai, J.; Shoemaker, N. Proc. 3rd Int. Biodegradation Symp., 1976.p731. 6. Lunberg, R. D.; Koleske, J. V.; Wischmann, Κ. B. J. Polym. Sci. 1969, A-l, 7, 2915. 7. Yamashita, Y.; Tuda, T.; Ishikawa, Y.; Miura, S. J. Chem. Soc. Jpn, Ind. Chem. Sec. 1963,66,110. 8. Carothers, W. H.; Arvin, J. A. J. Am. Chem. Soc. 1929, 5 1 , 2560. 9. Batzer, H.; Holtschmidt, H.; Wiloth, F.; Mohr, B. Makromol. Chem. 1951, 7 , 8 2 . 10. Marvel, C. S.; Johnson, J. H. J. Am. Chem. Soc. 1950, 7 2 , 1674. 11. Charch, W. H.; Shivers, J. C. Text. Res. J. 1 9 5 9 , 2 9 , 538. 12. Kiyotsukuri, T.; Takada, K.; Imamura, R. Chem. High Polym. Jpn. 1970, 2 7 , 410. 13. Waltz, J. E.; Tayler, G. B. Anal. Chem. 1947,19,448. 14. Tokiwa, Y.; Suzuki, T. Agric. Biol. Chem. 1977,41,2 6 5 . 15. Tokiwa, Y.; Suzuki, T. Nature 1977, 2 7 0 , 76. 16. Tokiwa, Y.; Suzuki, T. J. Appl. Polym. Sci. 1981, 2 6 , 441. 17. Tokiwa, Y.; Suzuki, T.; Ando, T. J. Appl. Polym. Sci. 1979, 24, 701. RECEIVED

January 8, 1990


Biodegradation of Synthetic Polymers Containing Ester Bonds

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