Environ. Sci. Technol. 2008, 42, 7696–7701
Up-Cycling of PET (Polyethylene Terephthalate) to the Biodegradable Plastic PHA (Polyhydroxyalkanoate) SHANE T. KENNY,† JASMINA NIKODINOVIC RUNIC,† WALTER KAMINSKY,‡ TREVOR WOODS,§ RAMESH P. BABU,§ CHRIS M. KEELY,§ WERNER BLAU,§ AND K E V I N E . O ’ C O N N O R * ,† School of Biomolecular and Biomedical Sciences, Centre for Synthesis and Chemical Biology, Conway Institute for Biomolecular and Biomedical Research, Ardmore House, National University of Ireland, University College Dublin, Belfield, Dublin 4, Republic of Ireland, Institute for Technical and Macromolecular Chemistry, University of Hamburg, Bundesstrasse 45, 20146 Hamburg, Germany, and Materials Ireland Polymer Research Center, School of Physics, Trinity College, University of Dublin, Dublin-2, Ireland
Received April 11, 2008. Revised manuscript received August 1, 2008. Accepted August 6, 2008.
The conversion of the petrochemical polymer polyethylene terephthalate (PET) to a biodegradable plastic polyhydroxyalkanoate (PHA) is described here. PET was pyrolised at 450 °C resulting in the production of a solid, liquid, and gaseous fraction. The liquid and gaseous fractions were burnt for energy recovery, whereas the solid fraction terephthalic acid (TA) was used as the feedstock for bacterial production of PHA. Strains previously reported to grow on TA were unable to accumulate PHA. We therefore isolated bacteria from soil exposed to PET granules at a PET bottle processing plant. From the 32 strains isolated, three strains capable of accumulation of medium chain length PHA (mclPHA) from TA as a sole source of carbon and energy were selected for further study. These isolates were identified using 16S rDNA techniques as P. putida (GO16), P. putida (GO19), and P. frederiksbergensis (GO23). P. putida GO16 and GO19 accumulate PHA composed predominantly of a 3-hydroxydecanoic acid monomer while P. frederiksbergensis GO23 accumulates 3-hydroxydecanoic acid as the predominant monomer with increased amounts of 3-hydroxydodecanoic acid and 3-hydroxydodecenoic acid compared to the other two strains. PHA was detected in all three strains when nitrogen depleted below detectable levels in the growth medium. Strains GO16 and GO19 accumulate PHA at a maximal rate of approximately 8.4 mg PHA/l/h for 12 h before the rate of PHA accumulation decreased dramatically. Strain GO23 accumulates PHA at a lower maximal rate of 4.4 mg PHA/l/h but there was no slow down in the rate of PHA accumulation over time. Each of the PHA polymers is a thermoplastic with the onset of thermal degradation occurring around 308 °C with the complete degradation occurring by * Corresponding author phone: +353 1 716 1307; fax +353 1 716 1183; e-mail:
[email protected]. † University College Dublin. ‡ University of Hamburg. § Trinity College. 7696
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370 °C. The molecular weight ranged from 74 to 123 kDa. X-ray diffraction indicated crystallinity of the order of 18-31%. Thermal analysis shows a low glass transition (-53 °C) with a broad melting endotherm between 0 and 45 °C.
Introduction PET is one of many petrochemical based plastics that contribute greatly to the convenience of everyday life. Best known for its use in plastic bottles it is produced on a multimillion tonne scale worldwide. Like other petrochemical plastics, the success of PET as a convenience bulk commodity polymer has led to post consumer PET products becoming a major waste problem. Greater than 5400 million lbs (2400 million kg) of PET bottles were on shelves in the U.S. in 2006 with only 23.5% of these bottles being recycled, and thus the vast majority of PET bottles end up in the landfill (1). This occurs despite a variety of recycling technologies being available such as mechanical grinding for use in the fiber industry and reprocessing for food contact usage. The current technologies recycle PET to a low value product and thus factors such as the high relative cost of sorting and the low value of the downstream product contribute to the poor recycling rates for PET. The conversion of PET to a high value product (upcycling) should lead to higher levels of PET recycling. The thermal treatment of PET in the absence of air (pyrolysis) generates terephthalic acid (TA) as the major product (Table 1). The TA generated is potentially a feedstock for the microbial synthesis of the value added biodegradable polymer polyhydroxyalkanoate (PHA). Consequently we are investigating the conversion of PET to this desirable high value material. We have previously reported the conversion of polystyrene to the biodegradable plastic PHA using a two step chemobiotechnological process which involves pyrolysis of polystyrene to styrene oil and subsequent feeding of that oil to bacteria that can utilize it as a carbon source to make a biodegradable carbon-based plastic (2). The conversion of PET to a biodegradable plastic has never been demonstrated and very few bacteria are known to degrade TA. Of those that degrade TA, none make PHA and thus we have searched a PET granule exposed soil to isolate new terephthalic acid degrading bacteria capable of accumulating PHA. PHA is the general term for a range of diverse polymers that consist of polyesters of (R)-3-hydroxyalkanoic acids. These polymers are accumulated by bacteria as intracellular carbon storage materials. It has been shown PHA accumulation occurs in response to a range of environmental stress factors such as inorganic nutrient limitation (3-5). The substrates supplied to bacteria to accumulate PHA can be divided into two groups: (1) PHA related substrates (fatty acids) that resemble the monomers that make up PHA (i.e., alkanoic acids, fatty acids) and (2) PHA unrelated substrates (i.e glucose, TA, etc.). There are two classifications of PHA based on the length of the monomer chains, short chain length PHA (sclPHA) which have monomers of 3-5 carbons long and medium chain length PHA (mclPHA) which have monomers of 6-14 carbons long (5). This variance in monomer chain length gives rise to varying properties in the polymer with sclPHA being more rigid and brittle than the elastomeric mclPHA (6). These biopolymers are of interest due to a broad range of applications and the fact that they are completely biodegradable (3, 7, 8). The cost of PHA production through fermentation is inextricably linked to the cost of the starting substrate. We investigate here the use of an easily sourced and inexpensive 10.1021/es801010e CCC: $40.75
2008 American Chemical Society
Published on Web 09/12/2008
post consumer petrochemical plastic waste as a feedstock for the synthesis of PHA by bacteria.
Experimental Section Hydrolytic Pyrolysis of PET. Virgin PET Krupp-Formoplast was supplied to a laboratory scale pyrolysis plant as previously described (9-11) at a feed rate of approximately 1 kg/h. The electrically heated fluidized bed had a diameter of 130 mm. Nine kg of quartz sand with diameters between 0.3 and 0.5 mm gave a height of 480 mm in the fluidized bed, which was maintained at a temperature of 450 °C. PET entered the fluidized bed reactor via a screw conveyor. The hot pyrolysis products passed a cyclone to be cleaned by small amounts of fillers and then were cooled down by mixing up with cold water to room temperature in a precipitator (desublimator). In the precipitator the terephthalic acid and some other solids (Table 1) were desublimated to generate a white powder. The sand bed was fluidized by steam with a flow rate of 2.5 kg/h. The solids were analyzed by HPLC-MS-system (HP 1100, column Multospher 100) using a diode array detector at 220 nm. The gas and oil fractions were characterized by gas chromatography (GC-FID, HP 5890 Machery & Nagel SE 52) and GC-MS (Fisons Instruments VG 70 SE, Machery & Nagel SE 52). Bacterial Growth Medium. The minimal mineral salts media E2 was prepared as previously described (12) and used as the media supplemented with sodium terephthalate as the sole source of carbon and energy for all culture techniques discussed in this work. The sodium terephthalate was prepared by taking the solid fraction of PET pyrolysis and dissolving in sodium hydroxide (Sigma). In addition, benzoic acid (2.4 g/L) was also used as a carbon source. Isolation of Bacteria from Soil. One kg of soil sample was collected from PET exposed soil at an industrial site used to mold PET granules to PET products. The granules were present in the soil adjacent to the factory setting. We reasoned that leaching of TA from PET may occur and that this soil would be a good source of TA degrading bacteria. The soil was sieved under aseptic conditions to a particle size of 5 mm, and then 10 g of soil were added to 90 mL of sterile Ringers solution (Sigma), this was vortexed for 5 min to homogenize the sample. Serial dilutions were performed to obtain a 10-5 dilution of the original soil sample. The serial dilutions were spread plated on solid E2 media containing 1.1 g/L of sodium terephthalate as the sole source of carbon and energy. Plates were incubated at 30 °C for 48 h. Various isolates were selected by visual differentiation of contrasting colony morphology. These isolates were then assessed for the ability to accumulate PHA. Growth Conditions for PHA Accumulation. In addition to microorganisms isolated from soil some bacterial strains, known to utilize TA as a sole source of carbon and energy, were tested for PHA accumulation. Both soil isolates and known TA degraders were grown in shake flask experiments, where each strain was grown in a 250 mL Erlenmeyer flask containing 50 mL E2 medium (4.2 g/L of TA) at 30 °C with shaking at 200 rpm. To screen for organisms capable of PHA accumulation the inorganic nitrogen source sodium ammonium phosphate (NaNH4HPO4.4H2O) was limited to 1 g/L (67 mg nitrogen/L). PHA Screening and Composition Analysis. Thirty-two soil isolates and three commercially obtained strains were grown in shaken flasks for 48 h and tested for PHA accumulation as previously described (13). The samples were analyzed on an Agilent 6890N series GC fitted with a 30 m × 0.25 mm × 0.25 µm HP-1 column (Hewlett-Packard) using a split mode (split ratio 10:1). The oven method employed was 60 °C for 2 min, increasing by 5 °C/min to 200 °C and holding for 1 min. For peak identification, PHA standards from P. putida CA-3 (14) and (R)-3-hydroxydodecanoic acid
(3HDDA) (Sigma) were used. PHA monomer determination was confirmed using an Agilent 6890N GC fitted with a 5973 series inert mass spectrophotometer, a HP-1 column (12 m × 0.2 mm × 0.33 µm) (Hewlett-Packard) was used with an oven method of 50 °C for 3 min, increasing by 10 °C/min to 250 °C and holding for 1 min. Nitrogen Determination Assay. The concentration of nitrogen in the growth media was monitored over time using the previously described method of Scheiner (15). Determination of Terephthalic Acid Utilization during Growth. The concentration of TA in the media was monitored by taking 1 mL samples from the culture flask at various time points and centrifuging the samples at 14 000g for 2 min. The supernatant was retained, filtered, and analyzed by HPLC. In order to analyze the sample and maintain a linear relationship between peak area on the HPLC chromatograph and TA concentration, samples had to be diluted so that the concentration of TA in the final preparation did not exceed 0.63 g/L. An Agilent 1100series HPLC using a C18 ODS Hypersil column (125 × 3 mm, particle size 5 µm) (Thermo) was used, and samples were isocratically eluted using 0.2% formic acid and acetonitrile (ratio 80:20, respectively) at a flow rate of 0.5 mL/min and read on a UV-vis detector at 230nm. The TA retention time under the above conditions was 3 min. Nuclear Magnetic Resonance (NMR). Solution NMR were recorded on a Bruker DPX400 with 1H at 400.13 MHz and 13C at 100.62 MHz. The solvent chloroform-d and tetramethylsilane (TMS) were used as internal references for chemical shifts in 13C and 1H NMR, respectively. 13C NMR spectra were recorded with proton-decoupling. Typically 2200 transients were accumulated. Spectrometer peak areas were obtained directly by standard signal integration. Thermal Analysis. Differential scanning calorimetry (DSC) was performed with Perkin-Elmer Pyris Diamond calorimeter calibrated to Indium standards. The samples weighing 7-8 mg were encapsulated in hermetically sealed aluminum pans and heated from -70 to 100 °C at a rate of 10 °C/min. To determine the glass transition temperature (Tg) the samples were held at 100 °C for 1 min and rapidly quenched to -70 °C. The samples were then reheated from -70 to 100 °C at 10 °C/min to determine the melting temperature (Tm) and Tg. The Tm was taken at the peak of the melting endotherm, while the Tg was taken as the mid point of heat capacity change, respectively. Thermogravimetric Analysis (TGA). To determine the thermal stability and decomposition profile of the samples, TGA was carried out on a Perkin-Elmer Pyris 1 thermogravimetric analyzer calibrated using Nickel and Iron standards. Each sample was weighed to ca. Seven mg and placed in a platinum pan and heated from 30 to 700 °C at the heating rate of 10 °C/min under an air atmosphere. Dynamic Mechanical Analysis (DMA). DMA was carried out on a Perkin-Elmer mechanical analyzer. Dynamic measurements were made in extension mode on clamped film samples with dimensions of 5 × 2.8 × 0.5 mm. The experiments were performed under nitrogen atmosphere at a temperature range of -100 to 50 °C at a heating rate of 2 °C/min and frequency of 0.1, 1, and 10 Hz. The Tg was identified by the sharp drop in storage modulus and the corresponding the peak in the loss modulus (16). DMA glass transition temperature is frequency dependent and detectable at higher temperature compared to the quasistatic DSC data. The temperature at the maximum point of the loss modulus (E′′) was taken as the measure of the glass transition temperature. Gel Permeation Chromatography. Molecular weight distribution were obtained by gel permeation chromatography (GPC) using PL gel 5 mm mixed-C +PL gel column (Perkin-Elmer) with PELV 290 UV-vis detector set at 254 VOL. 42, NO. 20, 2008 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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TABLE 1. Hydrolytic Pyrolysis of PET at 450°C product composition
weight percentage (%)
solids terephthalic acid oligomers benzoic acid others oil ethyleneglycol acetic aldehyde others gases co2 co hydrogen ethene others
77 51.0 20.0 1.0 5.0 6.3 0.75 5.10 0.45 18 13.0 3.5 0.18 1.0 0.34
nm. Spectroscopic grade chloroform was used as the eluent at flow rate of 1.0 mL/min. Sample concentration of 1% (w/ v) and injection volumes of 500 µL were used. A molecular weight calibration curve was generated with polystyrene standards with low polydispersity using the Turbochrom 4.0 software. X-ray Diffraction (XRD) Analysis. XRD was performed at room temperature and diffraction patterns were collected on a Siemens D500 diffractometer fitted with a Cu-KR radiation source. The X-ray beam was Cµ-KR (λ ) 0.1514 nm) radiation operated at 40 KV and 30 mA. Data was obtained from 2-60 °C(2θ) at a scanning speed of 0.1 °C/ min. 16S rDNA Identification. Three strains capable of accumulating PHA with TA as the sole carbon and energy source were selected and identified by sequence analysis of 16S rRNA genes. The genomic DNA of each bacterium was extracted as previously described (17). The 16S rRNA genes were amplified by PCR using primers 27F (agagtttgatcmtggctcag) and 1392R (acgggcggtgtgtgtrc) (18) and the sequences were determined by GATC-Biotech, Germany. The resulting sequences were compared to known sequences in the NCBI GenBank database by BLAST program (19).
Results and Discussion Polyethyleneterephthalate Pyrolysis. The pyrolysis of PET resulted in the generation of a solid, liquid and gaseous fraction (Table 1). 72% w/v of the terephthalic acid present in PET is recovered as monomeric TA (solid fraction). Oligomers of terephthalic acid make up almost 26% of the solid fraction. The addition of the solid fraction of PET pyrolysis to a solution of sodium hydroxide resulted in the hydrolysis of the oligomers and increased the proportion of TA making up the solid fraction to 97% w/w. The pyrolysis liquid fraction made up 6.3% w/v of the total weight of the pyrolysis products and contained predominantly acetic aldehyde and minor amounts of ethylene glycol. The gaseous fraction made up 18% of the pyrolysis products and contained predominantly CO2. The liquid and gaseous fraction were burned to provide energy for the pyrolysis of PET while the solid is desublimated, collected, and used as a feedstock for biodegradable plastic synthesis by bacteria. Isolation and Identification of PHA Accumulating TA Degraders from Soil. Three bacteria, known to utilize TA as the sole source of carbon and energy, i.e. Comamonas testosteroni YZW-D, C. testosteroni T-2, and C. testosteroni PSB-4 (20-24) were tested for their ability to accumulate PHA. However, these strains were incapable of PHA accumulation from TA derived from PET or commercially available sodium terephthalic acid (control). 7698
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TABLE 2. Identification and Comparison of Bacterial Isolates, From a Pet Exposed Soil, Capable of Growth and PHA Accumulation with TA As the Sole Source of Carbon and Energy
isolate
GenBank accession number
reference strain
GO16 DQ133506 Pseudomonas putida GO19 AY512611 Pseudomonas putida Pseudomonas GO23 AJ249382 frederiksbergensis
% % homology coverage 99 99
97 99
99
99
Since these strains were unable to synthesize PHA, screening for TA degrading strains from soil was performed. TA degrading strains were isolated from soil that had been exposed to PET granules at a PET processing factory in Ireland. From the PET exposed soil, 32 colonies with different morphologies growing on E2 media with TA as the sole source of carbon and energy were selected for further study. All of these isolates were screened for PHA accumulation after growth in shake flasks with limited nitrogen (to stimulate PHA production) and with TA as the sole source of carbon and energy. Of the 32 isolates screened, only three accumulated detectable levels of PHA. These three organisms were identified using 16S rDNA techniques. All three strains shared 99% homology with known Pseudomonas species (Table 2). Two of the three Pseudomonas strains are from the species putida, which are known to degrade a wide variety of aromatic compounds and more recently accumulate aliphatic mcl-PHA from aromatic compounds such as styrene and phenylacetic acid (14, 25). The other strain Pseudomonas frederiksbergensis GO23 is from a species reported to degrade the aromatic hydrocarbon phenanthrene (26). However this is the first report of any strain from this species accumulating PHA and any microorganism accumulating PHA from TA. Conversion of PET Derived Sodium Terephthalate to PHA in Shake Flask Experiments. All three strains accumulated PHA to between 23 and 27% of the total cell dry weight when supplied with TA either from a commercial source or from TA derived from the pyrolysis of PET. We used commercially available TA as a comparison to PET derived TA. PHA levels and composition were identical from both sources. PHA accumulation by each of the three strains was monitored over time to determine when the onset of PHA occurred as well as the time course for PHA production. All three organisms were grown in shake flasks under the nitrogen limited conditions with 4.2 g/L of sodium terephthalate (generated by PET pyrolysis). Nitrogen concentration (as ammonium), TA concentration, cell dry weight and quantity of PHA accumulated (Figure 1) were monitored. All three bacteria had similar growth patterns, they showed a long lag period in growth (of between 8 and 12 h) which coincided with a lag in TA utilization, despite being grown in precultures overnight on TA (4.2 g/L). During the exponential phase of growth strains GO16, GO19, and GO23 consumed TA at 0.135 g/L/h, 0.157 g/L/h, and 0.121 g/L/h, respectively, and had specific growth rates of 0.04 h-1, 0.043 h-1, and 0.049 h-1. All three strains consumed TA fully within the same period of time (Figure 1). While GO23 had the lowest rate of TA utilization during the exponential phase of growth it achieved the highest final cell dry weight and the highest growth yield from TA. While benzoic acid is a minor component of the solid fraction of the PET pyrolysis product, all three strains were capable of utilizing it as a sole carbon and energy source and thus it was also utilized during growth of the bacteria when the pyrolysis product was used as the substrate (data not shown). All three strains were also able
FIGURE 1. PHA accumulation by (A) P. putida GO16, (B) P. putida GO19, and (C) P. frederiksbergensis GO23 in shake flask containing growth medium consisting of 4.202 g/L of sodium terephthalate and 67 mg/L of nitrogen at 30 °C. CDW g/L (]), PHA accumulation g/l (∆), TA concentration (∆) and nitrogen concentration g/L (•) supplied as sodium ammonium phosphate were all monitored over a 48 h period. All data shown is the average of at least three independent determinations. to accumulate mclPHA when grown on benzoic acid (2.4 g/L) under nitrogen limited conditions to 20-27% of CDW. Once the nitrogen concentration in the growth medium was depleted below 15 mg/L at approximately 13 h, the onset of PHA accumulation occurred in strains GO16 and GO19, whereas PHA accumulation in GO23 began earlier. A comparison of the rate of TA utilization (g/l/h) and PHA accumulation (g/l/h) when PHA accumulation was occurring maximally indicates that strain GO19 was the most efficient at converting TA to PHA during this time period at 8.4 mg PHA/L/h. Strain GO23 appeared to accumulate PHA at a lower rate of 4.4 mg PHA/l/h but maintained this rate of PHA accumulation for a longer period of time compared to strains GO16 and GO19 (Figure 1). Thus process development and optimization of PHA production will be strain dependent. PHA Composition. The 1H NMR spectra for each PHA derived for strains GO16, GO19, and GO23 were established (Figure 2a). Peak assignments were typical of medium chain length PHA derivates (27-30). Terminal CH3 protons were detected by the strong resonance at 0.86 ppm. Another strong signal from the methylene hydrogens associated with saturated side chains occured at 1.25 ppm, whereas those methylenes associated with double bonds were detected at 2.0 ppm. The resonance at 1.56 ppm was associated with the methylene protons on C4. Methine protons were assigned to the quadruplet resonance located at 5.2 ppm. Finally the weak signal at 5.5 ppm was assigned to side-chains with sCHdCHs moieties.
FIGURE 2. NMR spectra of the mclPHA isolated from P. putida GO19 recorded at 20°C in CDCl3 (a) 400 MHz 1H spectrum (b) 13C spectrum.
TABLE 3. Composition of PHA Accumulated from Terephthalic Acid bacterial strain P. putida GO16 P. putida GO19 P. frederiksbergensis GO23
PHA (% CDW) 3HH 3HO 3HD 3HDDA 3HDDE 27 23
1 1
21 23
48 45
14 14
16 17
24
1
14
42
22
21
3HH ) 3-hydroxyhexanoic acid, 3HO ) 3-hydroxyoctanoic acid, 3HD ) 3-hydroxydecanoic acid, 3HDDA ) 3-hydroxydodecanoic acid, 3HDDE ) 3-hydroxydodecenoic acid.
The 13C spectrum for PHA produced from GO19 is illustrated (Figure 2b). Chemical shift assignments are prominently associated with medium chain length monomer structural units of 3-hydroxydodecenoic acid (3HDDE), 3-hydroxydecanoic acid (3HD), and 3-hydroxyoctanoic acid (3HO). The percentage of unsaturated side chains was estimated to be on the order of 5% by comparison of the methine signal intensities (134-123 ppm) to those of the methylene groups (10-40 ppm). The signal at 18.2 ppm is less than 1% of the total methylene group intensities and was previously assigned to 3-hydroxyhexanoic acid (3HH) (30). GCMS analysis of the PHA samples confirmed the presence of 3HH, 3HO, 3HD, 3HDDA, and 3HDDE with 3HD as the predominant monomer (Table 3). PHA from strain GO23 contained a high proportion of 3HDDA compared to PHA from strains GO16 and GO19. GCMS analysis indicated a higher proportion of 3HDDE monomer than estimated by 13C NMR. PHA standards from P. putida CA-3, commercially available 3HDDA, and a GCMS library (Agilent) were used to determine peak identity. VOL. 42, NO. 20, 2008 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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TABLE 4. Properties of PHA Polymer Extracted from P. putida GO16, P. putida GO19, and P. frederiksbergensis GO23 ∆ Tm TG strain Hm (°C) (°C)
MW
MN
PD
GO 16 12 35 -53 7.4 × 104 3.7 × 104 1.97 GO 19 10 34 -53 12.3 × 104 5.2 × 104 2.37 GO 23 11 35 -53 9.3 × 104 4.4 × 104 2.10
% crystallinity 26.8 18.7 31.1
While the sequence homology of 16S rDNA indicated a strong similarity between these bacteria often closely related species have differing PHA accumulation abilities (25). The PHA from P. putida GO23 contained a higher proportion of 3HDDA (22%) and 3HDDE (21%) compared to PHA from the other two strains (Table 3). It has been documented that the monomer composition of the PHA dictates the polymer properties (31) and while the difference between the GO23 polymer and the other two polymers presented in this study may appear small, the PHA polymer isolated from P. frederiksbergensis GO23 was physically different, showing an increased tackiness and malleability at room temperature compared to plastic from GO16 and GO19. PHA Properties. GPC analysis showed the polymers tested in this study ranged in molecular weight (Mw) from 74 kDa to 123 kDa (Table 4). The molecular weight distribution (Mw/ Mn) of the PHAs ranged from 1.9 to 2.4 (Table 4) and these values are typical for mclPHAs (32). The DSC analysis shows that PHA polymers produced in this study are partially crystalline, as evidenced by the presence of a melting peak. Previous reports have shown that short chain PHA produced by P. oleovorans from hexanoate and heptanoate did not have a clear melting peak, whereas PHA produced from longer-chain alkanoates exhibited melting endotherms (28). All three polymers produced with different bacterial strains showed similar Tg, with slight changes in their Tm and ∆Hm values (Table 4). X-ray diffraction (XRD) of cast films was used to calculate the crystallinity of the polymers (33). The strong diffraction peaks were located at the 2θ ) 19.58, 21.38, and 19.38 for PHA samples produced form GO16, GO19, and GO23, respectively. The calculated crystallinity values are between 26.8 and 31.1% for GO16, GO19, and GO23. The presence of higher amounts of 3HDDA and 3HDDE monomers leading to a higher percentage crystallinity is in keeping with previous reports (34). All three PHA products had similar thermal degradation patterns. Peak degradation maximum occurred at ca. 308 °C with a high temperature shoulder most evident at ca. 350 °C in the differential thermograph. Polymer degradation was completed by 370 °C with all residual carbonaceous materials produced during thermal degradation being burnt at about 600 °C. Thus three thermoplastics were generated through the chemo-biotecnological processing of PET. While certain avenues for PET recycling exist, a technology that can offer a clean and cost-effective way of converting PET to a high value polymer will generate a niche market in PET recycling. Pyrolysis offers a means of streaming this waste as a carbon feedstock for the production of PHA, a biodegradable polymer with broad uses encompassing biomedical as well as packing applications (35-37).
Acknowledgments This project has been funded under a grant from the Environmental Protection Agency of Ireland (ERTDI 2005ET-LS-9-M3), we thank Dr Zysltra for the provision of strain C. testosteroni YZW-D.
Literature Cited (1) NAPCOR 2006 Report on Post Consumer PET Container Recycling Activity; NAPCOR: Sonoma, CA, 2006. 7700
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