Received 21 March 2002; received in revised form 26 June 2002; accepted 17 July 2002

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1 European Polymer Journal 39 (2003) The biodegradation of poly-b-(hydroxybutyrate), poly-b-(hydroxybutyrate-co-b-valerate) and poly(e-caprolactone) in compost derived from municipal solid waste D.S. Rosa a, *, R.P. Filho a,b, Q.S.H. Chui a, M.R. Calil a, C.G.F. Guedes a a Laboratorio de Polımeros Biodegradaveis e Solucß~oes Ambientais UAACET, Universidade S~ao Francisco, Rua Alexandre Rodrigues Barbosa, no 45 Centro, CEP Itatiba, SP, Brazil b Pontifıcia Universidade Catolica de Campinas, Rodovia D. Pedro I, km 136, CEP Campinas, SP, Brazil Received 21 March 2002; received in revised form 26 June 2002; accepted 17 July 2002 Abstract Understanding the behavior of polymeric materials, particularly their biodegradation, is fundamental for solving problems in the management of environmental residues. In this work, we used a monitoring system based on an aerobic biodegradation technique known as the Sturm test to investigate the biodegradation of poly-b-(hydroxybutyrate), polyb-(hydroxybutyrate-co-b-valerate) and poly(e-caprolactone), in compost derived from municipal solid waste. The thermal analysis of these polymers was done using differential scanning calorimeter. The melting temperature and crystallinity were also determined. The results showed that poly-b-(hydroxybutyrate) degraded faster than the other two polymers, probably because the chemical structure of this polymer made attack by microorganisms easier. Ó 2002 Elsevier Science Ltd. All rights reserved. Keywords: Biodegradation; Sturm test; Compostage soil; Polyester 1. Introduction Polymers are widely used in the manufacturing of products such as bottles, bags, toothbrushes, tires and supports for electronic components. Most polymers are extremely durable, requiring more than 100 years for their degradation. This durability poses a serious environmental problem because of the large amount of waste produced, especially in urban centers [1]. One solution proposed for the management of plastic waste is the use of biodegradable products [1]. Biodegradation is the breakdown of materials through the * Corresponding author. Tel.: ; fax: address: derval@saofrancisco.edu.br (D.S. Rosa). action of living organisms. As defined by ASTM D [2], biodegradable polymers are polymers whose degradation occurs initially through the action of microorganisms such as bacteria, fungi and algae [3]. The most common products of such degradation are CO 2, CH 4, microbial cellular components and other products. In recent years, there has been an increasing interest in biodegradable polymers. However, the high cost of producing such polymers compared to conventional plastics is still one of the major problems to be solved. Most high molecular weight biodegradable polymers are polyesters that contain functional ester groups in their structures, which makes them more susceptible to attack and hydrolysis by fungi. Among biodegradable polymers, the best known is poly-b-(hydroxybutyrate) (PHB) which is produced on a large scale through bacterial fermentation [4] /02/$ - see front matter Ó 2002 Elsevier Science Ltd. All rights reserved. PII: S (02)00215-X

2 234 D.S. Rosa et al. / European Polymer Journal 39 (2003) Poly(e-caprolactone) (PCL) has also been studied as a substrate for biodegradation and as a matrix for the controlled release of drugs [5,6]. PCL is generally prepared from the ring-opening polymerization of e-caprolactone [7]. A third biodegradable polymer, Polyb-(hydroxybutyrate-co-b-valerate) (PHB-V), is a copolymer of hydroxybutyrate and random hydroxyvalerate segments. Both PHB-V and PHB are produced by bacteria. Variations in the concentration of valerate in the copolymer may lead to degradation times that range from some weeks to many years [8]. The chemical structures of PHB, PCL and PHB-V are shown in Fig. 1. Apart from the chemical structure of the polymers, other factors, such as their morphology and crystallinity, can influence the rate of degradation [9]. The ASTM has proposed several methods for analyzing and monitoring polymer biodegradation [10,11], including the determination of aerobic plastic biodegradation and plastic exposure to a simulated soil environment [3]. The methods proposed by the ASTM for assessing aerobic biodegradation are based on the Sturm test [12]. This assay is considered as the most reliable for evaluating polymer biodegradability in an active microbial environment. Appropriate environments for evaluating polymer biodegradation include activated slime and composted soil [1]. The production of CO 2 during polymer biodegradation is an important parameter for this process since the rate of CO 2 formulation corresponds to the rate of polymer biodegradation. In order to understand the behavior of PHB, PHB-V and PCL better, we have evaluated the biodegradation of these polymers in organic compounds. The melting temperature and crystallinity of these polymers were determined using differential scanning calorimetry (DSC). 2. Experimental 2.1. Materials (a) PHB was supplied by Copersucar (Cooperativa de Produtores de Cana, Acßucar e Alcool do Estado de S~ao Paulo, Brazil) and had a weight average molecular weight (M w ) of 250,000 with 0.2% nitrogen and 0.66% ash (both w/w). PHB was supplied in powder form and was sieved to obtain PHB with a size equivalent to 10 mesh. (b) PHB-V was also supplied by Copersucar and had a weight average molecular weight (M w ) of 150,000 with 0.09% nitrogen and 0.27% ash (both w/w). PHB-V was supplied in powder form and was sieved to obtain PHB-V with a size equivalent to 10 mesh. (c) PCL was provided by Union Chemical Carbide Ltd. (P-767) (Cubat~ao, SP, Brazil). The melting index was 1:9 0:3 g/10 min (ASTM D-1238), with a density of 1140 kg/m 3 and a weight average molecular weight (M w ) of 80,000. PCL was supplied in pellet form and ground using a model TE-625 mill (Tecnal Equipamentos para Laboratorios Ltda., Piracicaba, SP, Brazil), to provide PCL with a size equivalent to 10 mesh. A grain size (10 mesh) was chosen because the area of contact between the microorganisms and the polymers did not exceed the volumetric ratio (compost inoculum/ polymer) established by ASTM D-5338 [11] Composting of municipal solid waste The composted organic matter used as inoculator was supplied by the Araraquara composting plant (Araraquara, SP, Brazil). The sample used contained 18.7% organic matter, 10.4% total organic carbon, 5.9% humidity (110 C) and a ph of 8.6. This content was determined as described in ASTM D-4129 and ASTM D-1293 [13,14] Biodegradation test Fig. 1. Chemical structure of (a) PHB, (b) PCL and (c) PHB-V. To monitor CO 2 production in composted organic matter, four identical systems were assembled (Fig. 2), one for each polymer analyzed and one control to monitor the amount of CO 2 produced by organic material contained in the humus [10,11]. In each of the first three assemblages, a sample of the desired polymer was placed in recipient B (reactor) and immersed in the composted organic matter in the follow proportion: 600 g of humus, 100 g of polymer and 700 ml of distilled

3 D.S. Rosa et al. / European Polymer Journal 39 (2003) Fig. 2. Scheme for monitoring CO 2 production. water. The biodegradation was considered volumetric since the polymers used were in powder form, making it easier for microorganisms to act on them. The reactor was maintained at 58 2 C, as recommended by ASTM D [11] and the temperature of the mixture was monitored at 24 h intervals. If necessary, heating adjustments were made. To avoid contamination by CO 2 from the compressor, an aqueous solution of 0.03 M Ba(OH) 2 was added to recipient A (Fig. 2) to remove trace amounts of this gas. The CO 2 generated from the biodegradation of the polymer in recipient B was collected in recipient C, which also contained Ba(OH) 2 solution, thus resulting in barium carbonate (BaCO 3 ). This solution was prepared by adding 250 ml of distilled water and 2 12 g of barium hydroxide [Ba(OH) 2 ] to recipient C, depending on the speed of polymer biodegradation and, hence, the amount of CO 2 produced. The system was monitored every 24 h for about 54 days. The amount of CO 2 produced during biodegradation and collected in recipient C was determined by titration with 0.05 M HCl. The crystallinity was determined by using the following heat of fusion values for 100% crystalline materials: DH 0 PHB ¼ 146,000 J/kg, DH 0 PCL ¼ 136,100 J/kg and DH 0 PHB-V ¼ 146,000 J/kg. Note that the DH 0 for PHB-V was assumed to be the same as that for PHB [15]. 3. Results and discussions 3.1. Degradation test As shown in Fig. 3, PHB was degraded faster than the other polymers, with most degradation occurring between the 27th and 40th days of monitoring CO 2 production (2.72 g/day) (Table 1); PHB-V had the slowest degradation rate (practically zero) Thermal analysis Thermal analyses of PHB, PHB-V and PCL were done using a model 204 TASC 414/3A differential scanning calorimeter (DSC) (Netzsch-Ger atebau GmbH, Bavaria, Germany) under a nitrogen atmosphere, at a heating rate of 10 C min 1. Two heating cycles were used for each polymer. PHB and PHB-V were first heated from room temperature to 180 C to eliminate the thermal history of the samples, and then cooled to room temperature and immediately reheated to 200 C. For PCL, the temperatures used were 80 and 100 C, respectively. The second scan was done using the same heating rate as the first. All DSC experiments were done in duplicate and the thermograms shown refer to the second heating. Fig. 3. Mass of CO 2 collected as a function of time in the biodegradation tests. Table 1 Biodegradation rate of PHB, PCL and PHB-V Period (days) Speed of degradation (g/day) PHB PCL PHB-V ) )0.08

4 236 D.S. Rosa et al. / European Polymer Journal 39 (2003) PHB showed three different phases of degradation. In the first phase (0 26 days), the rate of CO 2 production was slow, in the second phase (27 40 days) this rate of production increased markedly, and in the third phase (41 54 days), CO 2 production stabilized. In the first phase, CO 2 production was detected only from the 13th day of monitoring onwards for the three polymers studied, probably because of the initial difficulty in breaking the polymer chain. Indeed, the chemical structure and morphology of these polymers can make the hydrolytic cleavage of ester linkages by microorganisms difficult, such that, as long as their molecular weight remains high, the plastics remain relatively immune to microbial attack [7]. According to Pitt [16], in this initial phase there is a small amount of random hydrolytic cleavage of ester linkages which decreases the molecular weight and produces some changes in the moleculeõs mechanical properties and morphology, but no weight loss. In this case, the cleavage is non-enzymatic and occurs within the polymer bulk. Such cleavage is not enough to produce CO 2. Following this initial phase, the rate of polymer degradation increases, indicating a greater susceptibility to degradation. There is now an exponential increase in the rate of carbon dioxide production through scission of the polymeric chains to form smaller fragments that are more easily biodegraded. Chemical degradation may decrease the molecular weight to the point where microbial attack can proceed [7]. This phase involves measurable weight loss in addition to chain cleavage that begins when the molecular weight of the polymer decreases to a point where scission produces fragments small enough to diffuse from the polymer bulk [16]. As show here, these fragments can produce carbon dioxide. The third phase is characterized by stabilization of the values since there is no CO 2 production, thus indicating the total degradation of PHB. Although PHB had a higher average molecular weight than the other polymers, this did not interfere with its biodegradation, probably because the chemical structure of the polymer made attack by microorganisms easier, i.e. a hydrolysis reaction leading to the production of an acid and an alcohol. The rate of CO 2 production for PCL was very low when compared to those for PHB and PHB-V, and probably reflected the strong interatomic forces holding the atoms together in this moleculeõs chemical structure [17] which hindered scission of the polymeric chains and consequently limited biodegradation. Unexpectedly, PHB-V showed no degradation during this monitoring (Fig. 3), probably because the ester hydrolysis responsible for the formation of acid fragments changed the soil ph, making attack by microorganisms difficult, as discussed in Section 3.3. Attack by molecular water was probably more difficult because this polymer had more ramifications, which would cause steric hinderance [8]. The negative results for PHB-V shown in Fig. 3 reflect experimental variation in the difference between the CO 2 production of control and PHB-V samples, as described in Section Thermal analysis The DSC curves for PHB, PHB-V and PCL are shown in Fig. 4. Table 2 summarizes the melting temperatures, the melting heat and the crystallinity of the polymers. PHB-V showed the lowest crystallinity values and PHB, the highest. The DSC curve for PHB (Fig. 4) had a short shoulder at low temperature, which possibly reflected the lower amount of low molecular weight polymer. Although this polymer had higher crystallinity values, the biodegradation test showed that PHB degraded before the other polymers, probably because it was more easily hydrolyzed. There was no correlation between the extent of biodegradation and the crystallinity of the polymers since PHB-V, which had lower crystallinity than the other polymers, also showed almost no biodegradation during the period of CO 2 monitoring. The crystallinity values showed that the PHB-V chains had greater difficulty in orientation than PHB and PCL because of the branched structure of PHB-V. Fig. 4. DSC curves for PHB, PHB-V and PCL. Table 2 Melting temperature, melting heat and crystallinity for PHB, PHB-V and PCL Polymer Melting temperature ( C) Melting heat (J/kg) PHB PCL PHB-V Crystallinity (%)

5 D.S. Rosa et al. / European Polymer Journal 39 (2003) Table 3 ph values after biodegradation study Polymer ph PHB 8.2 PCL 8.0 PHB-V 5.1 Control 8.6 The existence of many branches could make the hydrolysis of this material difficult. PCL had a crystallinity and biodegradation rate that was intermediate to those of the other two polymers Effect of biodegradation on soil ph The ph of the soil compostage was measured at the end of the experiment (54 days) (Table 3). The final ph of the soil compostage containing PHB and PCL was the same as the initial ph (Table 3). However, the ph of soil compostage containing PHB-V decreased from 8.6 to 5.1. The variation in ph probably reflected the accumulation of organic acids through the degradation of organic material already contained in the humus [8,14]. In this case, the low ph (5.1) must have inhibited the activity of microorganisms (enzyme secretion and/or release) responsible for biodegradation and would not have favored hydrolysis of the ester linkage necessary for biodegradation. 4. Conclusions The system used here to monitor the biodegradation of polymers proved viable and provided information on the overall rate of degradation. PHB was biodegraded faster than the other polymers; no CO 2 production was registered for PHB-V. The biodegradation of PHB occurred in three phases that differed in their speed. The first phase involved hydrolysis of the polymer, with attack by water molecules and consequent polymer scission, and the formation of polymer fragments. The second phase involved action by microorganisms, with enzymatic attack on the fragments generated in the first phase; the third phase was characterized by stabilization of the values, indicating total biodegradation. The decrease in the ph of soil compostage containing PHB-V possibly reflected the lack of degradation of the acids in the first phase of degradation involving ester hydrolysis (as described for PHB). No correlation was observed between crystallinity and the biodegradation of these polymers. The low biodegradation of PCL was attributable primarily to the structure formed by CH 2 groups, which have a large bonding force, and to the absence of tertiary carbons. Acknowledgements The authors thank FAPESP for financial support (process 99/ ) and the companies Union Carbide of Brazil Ltd. and PHB of Brazil for supplying raw materials. The authors also thank the Laboratorio de Caracterizacß~ao de Materiais (LCAM), Universidade S~ao Francisco, for use of their thermal analysis equipment. References [1] Huang SJ. In: Eastmond GC, Ledwith A, Russo S, Sigwalt P, editors. Comprehensive polymer science. Oxford: Pergamon Press; p [2] ASTM Designation D Standard Terminology Relating to Plastics, 2000;8(01): [3] Raghavan D. Characterization of biodegradable plastics. Polym Plast Technol Eng 1995;34(1): [4] Chiellini E, Solaro R. Biodegradable polymeric materials. Adv Mater 1996;8(4): [5] Pitt CG, Marks TA, Schindler A. In: Baker R, editor. Biodegradable drug delivery systems based upon aliphatic polyesters: Application to contraceptives and narcotic antagonists. New York: Academic Press; p [6] Pitt CG et al. The degradation of poly(e-caprolactone) in vivo. J Appl Polym Sci 1981;26: [7] Chandra R, Rustgi R. Biodegradable polymers. Progr Polym Sci 1998;23: [8] Scott G. Introduction to the abiotic degradation of carbon chain polymers. In: Scott G, Gilead D, editors. Degradable polymers. London: Chapman & Hall; p [9] Stevens MP. In: Polymer chemistry an introduction. New York: Oxford University Press; p [10] ASTM Designation D Standard Specification for Compostable Plastics,2000:8(03); [11] ASTM Designation D e. Standard Test Method for Determining Aerobic Biodegradation of Plastic Materials under Controlled Composting Conditions, 2000:8(03); [12] Rosa DS, Penteado DF, Calil MR. Biodegradabilidade de PCL e PHB em pool de microorganismos. Polımeros Ci^encia e Tecnologia 2001;11(2):82 8. [13] ASTM Designation D (1993) e1. Standard Test Method for Total and Organic Carbon in Water by High Temperature Oxidation and by Coulometric Detection, 1999:11(02); [14] ASTM Designation D Test Methods for ph of Water, 1999:11(01); [15] L upke T, Radusch HJ, Metzner K. Solid-state processing of PHB-powders. Macromol Symp 1998;127: [16] Pitt CG. Non-microbial degradation of polyesters. In: Vert M et al., editors. Mechanisms and modifications. Cambridge: The Royal Society of Chemistry; p [17] Callister Jr WD. In: Ci^encia e engenharia de materiais: uma introducß~ao. Rio de Janeiro: LTC SA; p