Radiation effect on polyesters

Radiat. Phys. Chem. Vol.46, No. 2, pp. 233-238, 1995 Copyright (~, 1995ElsevierScienceLtd 0969-806X(95)00018-6 Printed in Great Britain.All rights reserved 0969-806X/95 $9,50+ 0,00

Pergamon

RADIATION EFFECT ON POLYESTERS HIROSHI MITOMO, I YUHEI WATANABE, 2 F U M I O YOSHII 2 and KEIZO M A K U U C H F ~Department of Biological and Chemical Engineering, Faculty of Engineering, Gunma University, Kiryu, Gunma 376, Japan and :Japan Atomic Energy Research Institute, Takasaki Radiation Chemistry Research Establishment, 1233 Watanuki, Takasaki, Gunma 370-12, Japan Abstraet--Poly(3-hydroxybutyrate)(PHB) and its copolymer poly(3-hydroxybutyrate-3-hydroxyvalerate) [P(HB-HV)] were irradiated with ?-rays in air or vacuum. Polymer chain scission occurred and resulted in depression of melting points (Tin), glass-transition temperatures (Tg) and number-average molecular weight (-~n). Decrease in A~, of the sample irradiated in vacuum was smaller than that irradiated in air, implying introduction of crosslinking. The Tmand T~of samples irradiated in air were inversely proportional to AT,. Their biodegradabilitywas clearly promoted with decreasing 2Q.. Radiation grafting of methyl methacrylate (MMA) or 2-hydroxyethyl methacrylate (HEMA) was carried out by in-source polymerization. Degree of grafting (X~) increased as irradiation dose increased and leveled off around 5 kGy. The Xg of PHB grafted was lower than that of P(HB-HV) because of higher crystallinity of the former. Crosslinking between the grafted PMMA chains was easily formed. Biodegradability of both polymers steeply decreased by introduction of MMA grafting, while that of polymers grafted with HEMA increased at first because of improvement of wettability then steeply decreased with increasing Xg of HEMA.

copolymer are thermoplastic and biocompatible materials (Abe et al., 1992), In the present study, the radiation-induced degradation of PHB and its copolymer was investigated and the changes in their properties and biodegradability were discussed. Radiation-induced graft polymerization of methyl methacrylate (MMA) or 2-hydroxyethyl methacrylate (HEMA) onto these polymers was carried out and improvement of their properties was studied. Changes in the properties and biodegradability were compared with the degree of grafting.

INTRODUCTION Poly(3-hydroxybutyrate) (PHB) and its copolymer poly(3-hydroxybutyrate-3-hydroxyvalerate) [P(HBHV)] are microbial and biodegradable polyesters produced by many types of bacteria (Doi et al., 1988). Radiation-induced degradation has been investigated for common polymers, e.g. polyethylene (Geetha et al., 1988) and polypropylene (Kagiya et al., 1985). Recently N M R study on structural changes in butyl rubber after 7-rays irradiation has been reported, where the yields of main-chain scission and crosslinking were estimated and identified (Hill et al., 1992). Radiation grafting of hydrophilic monomers onto many polymers, e.g. polyethylene (Ishigaki et al., 1982), polypropylene (Mukherjee et al,, 1985; Gupta et al., 1990) and poly(4-methylpentene-1) (Soebianto et al., 1987) has been studied mainly for biomedical applications. It is worth improving their properties by radiation grafting for wider usage though PHB and its

EXPERIMENTAL Materials

PHB and P(HB-HV) containing 20 and 24 mol.% HV (hereinafter abbreviated as 20 M and 24 M samples), which were isolated from Alcaligenes eutrophus, were purchased from Aldrich Chemical Co. Irradiation

HB

U!:3

PHB and 20 M samples were sealed in glass ampules with the atmosphere of dried air or vacuum, and then irradiated with 6°Co ?-rays at a dose rate of 10 kGy/h for various hours at 25°C.

I-

HV

Jr

Scheme 1 233

Graft polymer&ation

PHB and 24 M samples immersed with M M A (bulk and 20 vol.% M M A in CH3OH) or HEMA (10 vol.% in CH3OH) monomer in vacuum glass ampules at room temperature were directly irradiated with 6°Co

Hiroshi Mitomo et al.

234

y-rays at a dose rate of 1 kGy/h for various periods of time (called as in-source polymerization) at 25°C. For comparison, both samples were preirradiated with 5 kGy in vacuum at -78°C. The MMA or HEMA monomer solution was introduced to the irradiated polymers and graft polymerization was carried out under nitrogen gas atmosphere for various periods of time at a constant temperature (hereafter called post polymerization). Grafted PHB was Soxhlet extracted with acetone for 3 days, while grafted 24 M sample was soaked with fresh acetone at room temperature for a week so as to remove any MMA monomer and adhering homopolymer. Both samples grafted with HEMA were Soxhlet extracted with methanol for 3 days and dried under vacuum at 35°C to constant weight. Degree of grafting X~ (%) was determined by the percent increase of weight based on the original sample weight W~ (g), using equation (1) x~ = [ ( w ~ - w,)/wi] x 100

(1)

where Wg is the weight of sample after grafting (g).

1000

~

3000 ~

-

20

M

2000 I

I 80

I

( 100

I

I 120

Temperature

I

I

I

140

J

160

I

I

180

(°C)

Fig. 1. DSC heating curves of PHB and 20 M samples irradiated with various doses in air.

Analytical procedures Melting point Tm and glass transition temperature T~ were measured with a Perkin Elmer Model DSC-7 differential scanning calorimeter (DSC) at a heating rate of 10°C/min under helium or nitrogen atmosphere. The melting peak temperature was calibrated with high-purity standards. Gel permeation chromatography (GPC) was carried out with an HLC-802A high performance liquid chromatograph (Tosoh Co., Ltd) at 38°C. The eluent was chloroform with a flow rate of I ml/min. The 37, was calibrated using polystyrene standards and radiation-degradated PHB samples of six different 37, values (4.29 x 103 to 5.64 x 105) evaluated by GPC and a low-angle laser light scattering system.

Enzymatic degradation The enzymatic degradation of polymers by the extracellular PHB depolymerase (kindly offered by Professor T. Saito, Kanagawa University) purified from A.faecalis T1 was carried out at 37°C in 0.1 M phosphate buffer (pH 7.4). The films (initial weights, 11-13 mg; initial film dimensions, 10 x 10 × 0.1 mm) melt-quenched or solvent-casted from chloroform solution were immersed with 1 ml of the buffer. The reaction was started by the addition 32pl of an aqueous solution of PHB depolymerase (8/~g). The reaction solution was incubated at 37°C with shaking. The samples were periodically removed, washed with warm water, and dried to constant weight in vacuo. Weight loss of the film (X~) was calculated by the percent decrease of weight based on the original film weight W~ (g), using equation (2). Xd(%) = [(W~ - Wa)/W,] × 100

zoo

(2)

where Wa is the weight of sample after enzymatic degradation (g).

RESULT AND DISCUSSION

The T,,, Tg and M, of radiation degradated PHB and 20 M samples Typical DSC heating curves of PHB and 20 M samples irradiated in air with various irradiation doses are shown in Fig. 1. The peak temperature of PHB steeply decreased and the peak decreased in its area by degrees as the irradiation dose increased except for the sample irradiated with 50 kGy, which showed slight increase (ca 5%) in peak area. The sample irradiated at the dose above 50 kGy showed a doublet melting peak composed of a main peak and small peak appearing at temperatures 10-20°C lower, which implies that the further degradation proceeded heterogeneously in crystalline regions. The curves of 20 M sample showed similar variation to those of PHB. Melting peak area of 20 M sample was smaller than that of PHB, reflecting its low crystallinity and slow crystallization rate (Mitomo et al., 1987). Figure 2 shows plots of Tmand 37, of PHB and 20 M samples irradiated in air and vacuum. The Tm values of both samples irradiated in air decreased almost parallel, while those in vacuum decreased gently as the dose increased. This implies that destruction in crystalline region by chain scission proceeds gently rather than that in air because of absence of oxygen and introduction of crosslinking. The 37, values of both sample significantly decreased at first, then decreased more slowly as the dose increased. At lower doses (0-100 kGy), the main chain scission mainly occurred in the folded chain regions of crystal surface and then it began to take place even within the crystalline regions at the dose above 100 kGy. The 37,

Radiation effect of polyesters

235

200

80

15£ ¢

,

lOO

~ 50

i

i

I

I

40

""

t

10~ 0 0

200

500

1000

Irradiation dose (kGy) 104

. . . . - : : I t : ..... ~t 103 0

1000

2000

3000

Irradiationdose (kGy) Fig. 2. Plots of Tmand/~, of irradiated PHB (O, O) and 20 M (A, &) samples against the irradiation dose. The solid and dashed lines show those irradiated in air and vacuum, respectively.

values of both samples irradiated in vacuum decreased more slowly than those irradiated in air. Figure 3 shows plots of Tm and T~ of irradiated samples against h,7~. Linear relationships of them are

o

200

150

15(

100

100

50

E

Fig. 4. Plots of weight loss of the irradiated samples by biodegradation against the irradiation dose. PHB irradiated in air (O) and vacuum (0); 20 M in air (A) and vacuum (A). Solid and broken lines show the treatment time of 8 and 16 h, respectively. observed for both samples irradiated in air just similar as reported for polystyrene considering the free volume (Fox and Flory, 1950), however, such relations are not observed any more for them irradiated in vacuum because of introduction of crosslinking between the polymer chains.

Enzymatic degradation o f radiation degradated PHB and 20 M samples Figure 4 shows weight loss curves of the irradiated films by enzymatic degradation. The films used were obtained by melt-quenching. Weight loss values of PHB were 15 and 29%, whereas those of 20 M were 11 and 21% after treated for 8 and 16 h, respectively, The PHB films show faster rate of degradation than 20 M irradiated with the same dose. Both PHB and 20 M films irradiated with higher doses than 120 and 210kGy were completely degradated after the treatment time of 16 h, respectively. Whereas weight loss values of both samples irradiated in vacuum were ca 50% at the same conditions and levelled off at the dose of 1000kGy, while that irradiated in air continued to increase. Therefore, it can be said that effect of crosslinking on biodegradability is considerably large. Comparing weight loss curves of both samples with kTncurves in Fig. 2, it can be roughly said that the weight loss is inversely proportional to logarithmic value of A~,. In other words, biodegradation is clearly promoted with decreasing ~S¢o.

Radiation grafting with M M A and H E M A onto PHB and 24 M samples

-20 I

s 1/~' n (XI0

i0 4)

Fig. 3. Plots of Tmand Ts of the irradiated PHB and 20 M samples against 1/ATe.Samples and symbols in Fig. 3 are the same as those in Fig. 2.

Typical DSC heating curves of PHB and 24 M samples grafted with MMA are shown in Fig. 5. The Tm values slightly decreased and peak area decreased significantlywith introduction of MMA grafting. DSC curve of second run were shown as dashed lines, which displayed that MMA grafting hardly prevented from

236

Hiroshi Mitomo et al. Table I. Meltingpoints(Tin)and enthalpiesof fusion(AHm)of PHB and 24 M samplesgrafted with MMA Dose T~ AHm (AH,~) .... X~ Sample (kGy) (C) (J/g) (J/g) (%) PHB 0 176.8 86.1 86.1 0 0.5 174.3 78.0 87.4 12 I 173.3 73.7 86.2 17 2 172.5 66.9 82.3 23 3 171.8 62.8 79.8 27 24 M 0 123.5 64.2 64.2 0 0.5 122.5 52.4 60.3 13 I 120 46.6 58.5 24 2 116 33.4 51.9 54 3 111.5 21.3 39.8 87

PHB O

LU

3 kGy . . . . . . . . . . . . . . . . . . . . . . .

~

4M 1 kGy

~

3kGy I

t

I

i

I

i

I

lO0

60

i

i

i

I

,

140

i

i

180

Temperature (°C)

Fig. 5. DSC heating curves of PHB and 24 M samples grafted with MMA by in-source polymerization. Simultaneous radiation doses are indicated.

recrystallizing of PHB but prevented from that of 24 M sample. This implies that M M A grafting onto 24 M sample was introduced not only in amorphous but also in crystalline regions because of disordered PHB crystal lattice by occlusion of the HV units (Mitomo e t al., 1987), Figure 6 shows plots of degree of grafting (Xs) with M M A (bulk and 20 vol.% M M A in CH3OH) onto PHB and 24 M samples against the irradiation dose. The Xg of 24 M sample, which has lower crystallinity than PHB, increased far faster than that of PHB. This is explained that M M A grafting was mainly

introduced in amorphous part or distorted crystal lattice regions. Both samples showed slight increase in Xg when used with 20% M M A in CH3OH, which is explained that the M M A solution became less viscous by dilution and improved on monomer supply during polymerization. Figure 6 also plots of Xg of H E M A onto both samples as dashed lines. The monomer solution was 10% H E M A in CH3OH. Very similar to the case of M M A grafting, Xs of 24 M sample increased faster than that of PHB. This is similar to the result of polypropylene grafted with H E M A (Gupta e t al., 1990), where Xg sharply increased as the dose increased and levelled off to c a 75% at 3 kGy. The Tm enthalpies of melting (AHm) and Tg for PHB and 24 M samples grafted with M M A are listed in Table 1, The decrease in T~ of 24 M was larger than that of PHB, reflecting the introduction of M M A grafting in crystalline regions in addition to amorphous regions. The AHm of PHB decreased from 86 to 63 J/g, however, this decrease is due to the relative decrease in PHB content in the grafted sample. Therefore, AHm may be corrected by the weight fraction of PHB and 24 M polymers in the grafted samples according to the following equation: Corrected enthalpy of melting (AHm)..... = AHm(1 + Xg) (3)

lOO

ID

N so D

I

I

I

I

1

2

3

4

5

Dose (kGy)

Fig. 6. Plots of X~with MMA (bulk) onto PHB (O) and 24 M (I--I)samples and X8 with 20% MMA in CH~OH onto PHB (O) and 24 M (11) against the irradiation dose. Dashed lines show plots of Xs with 10% HEMA in CH3OH onto PHB (A) and 24 M (&) samples.

where Xg is the degree of grafting. The (AHm)..... of PHB remained almost unchanged, implying that M M A grafting was hardly introduced in crystalline regions, whereas (AHm)..... obtained for 24 M sample decreased significantly, implying that the crystalline regions were considerably destructed by M M A grafting. The Tg values of PHB and 24 M were 2.5 and -5.5°C, respectively, which shifted up to 8~C higher with increasing X~ value. Since Tg of P M M A appeared at a broad temperature range around 80'C, it is natural that Tg of the grafted polymers increased with increasing Xg. The Mo values of PHB and 24 M samples were 2.84 × 105 and 2.09 × 105, respectively, which increased up to c a 20 × l0 s after M M A grafting. From TG measurement, grafted P M M A homopolymer could obtain by heating up to ca 300°C, because the both polyesters completely decomposed at 280:'C,

Radiation effect of polyesters

237 CONCLUSION

8o |

O, • : PHB

L

""

60(

~.11

24M

,

,.

-

~

',

40

",

"',"a 20

I

0

__

50

__

I

100

Degree of grafting (%)

Fig. 7. Weight loss values of PHB (O) and 24 M (I--1)samples grafted with MMA (solid line) and those of PHB ( 0 ) and 24 M ( I ) grafted with HEMA (dashed line) obtained by enzymatic degradation test.

while P M M A decomposed at a higher range of 300-420"C. The M,, of grafted P M M A thus obtained was around 15 × 105, while that of grafted P M M A obtained from the samples grafted by post polymerization was around 3 × 105. This suggests that crosslinking between grafted P M M A or polyester chains was introduced by in-source polymerization.

The chain scission occurred within amorphous regions at lower irradiation doses (,,~ 100 kGy), and proceeded farther to crystalline regions at higher doses (above 100 kGy). The T,, and Ts of the samples irradiated in air were inversely proportional to .~,, while the samples irradiated in vacuum no longer showed linear relations because of simultaneous formation of crosslinking. Rate of biodegradation increased as the dose increased, though rate of them irradiated in vacuum became slower than that irradiated in air because of introduction of crosslinking. Degree of grafting X~ of M M A onto PH B was lower than that onto 24 M sample. This is explained since the former had higher crystallinity and grafting was mainly introduced to amorphous regions. The h4, of P M M A grafts introduced by post polymerization is considered to be comparable to that of trunk polymers, while/Q, of P M M A grafted by in-source polymerization was far larger reflecting introduction of crosslinking between grafted P M M A chains. Biodegradability decreased by introduction of M M A grafting, while that increased at first by introduction of hydrophilic H E M A grafting because of improvement of wettability then decreased almost parallel to that of the sample grafted with MMA. REFERENCES

Enz)'matic degradation o f the grafted P H B and 24 M samples

Figure 7 shows enzymatic degradation profiles on these samples grafted by in-source and post polymerization methods. The samples used were solvent-casted films. The weight loss of PHB (60%) was larger than that of 24 M sample (46%)just similar to values in Fig. 4, implying that the introduction of the HV component causes slight decrease in biodegradability (Doi et al., 1990). The weight loss of PHB and 24 M samples grafted with M M A steeply decreased to 16 and 13 % at Xg of ca 50%, respectively, and both leveled off to ca 10% at the higher value of Xg. It is considered that this enzymatic degradation process is depolymerization process from the polymer chain end (Doi et al., 1990), therefore, this is strongly affected by grafting or crosslinking. Whereas, the weight loss values of both samples grafted with H E M A increased clearly at Xg of ca 10-20% and decreased almost parallel to those of the samples grafted with M M A and levelled off to ca 20%. This is explained that the samples were modified to be hydrophilic and improved on wettability with the enzyme solution by introduction of H E M A grafting nevertheless P(HEMA) per se showed a poor biodegradability. The more predominant increase in biodegradability was observed in these polymers grafted with more hydrophilic monomer such as acrylic acid, whose results will be reported elsewhere.

Abe H., Yamamoto Y. and Doi Y. (1992) Preparation of poly(3-hydroxybutyrate) microspheres containing lastet of an anticancer drug and its application to drug delivery system. Kobunshi Ronbunshu 49, 61. Doi Y., Tamaki A., Nakamura M. and Soga K. (1988) Production of copolyesters of 3-hydroxybutyrate and 3-hydroxyvalerate by Alcaligenes eutrophus from butyric and pentanoic acids. Appl. Microbiol. Biotechnol. 28, 330. Doi Y., Kanesawa Y., Kunioka M. and Saito T. (1990) Biodegradation of microbial copolyesters: poly(3hydroxybutyrate-co-3-hydroxyvalerate) and poly(3-hydroxybutyrate-co-4-hydroxybutyrate). Macromolecules 23, 26. Fox T. G. and Fiery P. J. (1950) Second-order transition temperatures and related properties of polystyrene. I. Influence of molecular weight. J. Appl. Phys. 21, 581. Geetha R., Torikai A., Yoshida S., Nagaya S., Shirakawa H. and Fueki K. (1988) Radiation-induced degradation of polyethylene: effects of processing and density on the chemical changes and mechanical properties. Polym. Deg. Stab. 23, 91. Gupta B. D., Tyagi P. K., Ray A. R. and Singh H. (1990) Radiation-induced grafting of 2-hydroxyethyl methacrylate onto polypropylene for biomedical applications. I. Effect of synthesis conditions. J. Macromol. Sci.-Chem. A27, 831. Hill D. J. T., O'Donnell J. H., Senake Perera and Pomery P. J. (1992) Determination of scission and crosslinking in gamma irradiated butyl rubber. Radiat. Phys. Chem. 40, 127. lshigaki I., Sugo T., Senoo K., Okada T., Okamoto J. and Machi S. (1982) Graft polymerization of acrylic acid onto polyethylene film by preirradiation method. 1. Effects of preirradiation dose, monomer concentration, reaction temperature, and film thickness. J. Appl. PoO'm. Sci. 27, 1033.

238

Hiroshi Mitomo et al.

Kagiya T., Nishimoto S., Watanabe Y. and Kato M. (1985) Importance of the amorphous fraction ofpolypropylene in the resistance to radiation-induced oxidative degradation. Polym. Deg. Stab. 12, 261. Mitomo H., Barham P. J. and Keller A. (1987) Crystallization and morphology of poly(fl-hydroxybutyrate) and its copolymer. Polymer J. 19, 1241. Mukherjee A. K. and Gupta B. D. (1985) Radiation-induced graft copolymerization of methacrylic acid onto

polypropylene fibers. I. Effect of synthesis conditions. J. AppL Po(vm. Sci. 30, 2643. Soebianto Y. S., Yoshii F., Makuuchi K. and lshigaki I. (1987) Radiation grafting of hydrophilic monomers onto poly(4-methylpentene-1). I, Grafting of acrylic acid, Angew. Macromol. Chem. 149, 87; II. Grafting of long chain monomers and physical properties of the grafted films, ibid. 152, 159.