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Assessment of biodegradability of plastics under simulated composting conditions in a laboratory test system
International
Biodeterioration
& Biodegradation
(1996) 85-92
Copyright 0 1996 Elsevier Science Limited Printed in Great Britain. All rights reserved 0964-8305/96 $15.00 + 0.00 0964-8305(95)00089-5
ELSEVIER
Assessment of Biodegradability of Plastics under Simulated Cornposting Conditions in a Laboratory Test System Andreas Starnecker’ & Michael Menner Fraunhofer-Institute for Food Process Engineering and Packaging, Steinerstr. 15, 81369 Munich, Germany
An automated laboratory-scale test system was developed for measuring the aerobic biodegradability of degradable plastics under simulated composting conditions. Biodegradation was monitored by measuring microbial carbon dioxide formation and oxygen consumption. Completeness of biodegradation was assessed in an aquatic test by conducting a carbon mass balance. The percentage of plastic carbon degraded to carbon dioxide, biomass and watersoluble byproducts were determined. The rate of biodegradation under simulated composting conditions was measured in a fixed-bed system with mature compost. A time-dependent temperature profile was applied to simulate the natural selfheating of a composting process. The aquatic test was conducted at a constant temperature of 30°C as well as with a temperature profile. The rate of biodegradation was significantly higher in the aqueous environment. Equal degrees of mineralization were reached in the aquatic and the fixed-bed system only if the same temperature profile was applied. Conducting the aquatic test at a constant temperature of 30°C less microbial carbon dioxide formation was observed. However, a carbon mass balance revealed that taking into consideration the portion of the plastic’s carbon incorporated into biomass, a similar degree of biodegradation was reached. Consequently, the measurement of microbial carbon dioxide production is not sufficient to assess the extent of biodegradation of plastics. In the heterogeneous matrix ‘compost’, it is not feasible to assess the completeness of biodegradation due to limited possibilities to analyze degradation intermediates and biomass growth. Therefor& a new fixed-bed system with an inert, carbon-free packing material was developed. The inert material was inoculated with an aqueous eluate from compost. First results showed biodegradation rates similar to a compost environment. Copyright 0 1996 Elsevier Science Limited.
admission criteria for input In Germany, materials of an industrial composting process are specified in the German federal states guidelines for compost (LAGA M 10, 1995). Only input materials which are completely biodegradable are accepted for cornposting. Non native-organic materials have to pass a suitability test. Figure 1 shows the current test procedure for the assessment of compostability as proposed by the working committee ‘Biodegradable Plastics’ FNK 103.3 of the German Institute for Normification (DIN) (Schroeter et al., 1994). First, there is a basic chemical product analysis to identify pollutants (e.g. heavy metals, polychlorinated aromatics) or non-biodegradable components of the plastic. Second, two laboratoryscale tests for the assessment of complete aerobic
INTRODUCTION Composting represents an increasingly important route of disposal for the organic fraction of municipal solid waste. In the future also, materials that are not of native-organic origin are planned as input materials for composting facilities. One important group of those man-made materials are biodegradable plastics. Unlike most of the can be commodity plastics, these polymers degraded and utilized as a source of .carbon and energy by a variety of microorganisms, if they get into an environment favourable for microbial activity (e.g. compost, soil, waste water). ‘Paper presented at the fourth meeting of the Biodegradable Plastics Group, International Biodeterioration Research Group, Winchester, UK, May 3-5, 1995. 85
86
A. Starnecker, M. Menner
Laboratory-scale
Pilot plant-scale
+Lzzq field-proven model experiment
* Field testing
Fig. 1. Test procedure for the assessment of compostability
biodegradability are proposed: an aquatic screening test and a solid-state compost simulation test. Existing biodegradability lab-tests for plastics (e.g. ASTM 5209-92, ASTM 5338-92) are currently being modified to experimentally prove completeness of biodegradation. The next two steps of the test procedure examine the suitability of the plastic material for cornposting. This is assessed under controlled conditions in pilot plant-scale cornposting units and under practical conditions in technical cornposting plants (Streff et al., 1994). Finally, utilization properties and ecotoxicology of the compost obtained by adding a certain percentage of the plastic as input material are investigated. Ultimate biodegradation Microbial degradation of plastics is initiated by the secretion of enzymes, which cause a chain cleavage of the polymer into oligomers or monomers, respectively. These water-soluble enzymatic cleavage products are absorbed into the microbial cell where they are metabolized. End-products of the aerobic metabolism are basically carbon dioxide and water. A portion of the plastic’s carbon is utilized for the build-up of new biomass. According to the definition of the DIN, a plastic is called biodegradable if all of its organic components undergo a total biodegradation to naturally occurring metabolic end-products (Pantke, 1994). This process is called ultimate
(draft proposal DIN FNK 103.3).
biodegradation (Gilbert & Watson, 1977). Biomass itself eventually will be degraded to carbon dioxide. However, over the usual time scale of a biodegradation test, complete mineralization to carbon dioxide generally will not be observed. Therefore, the DIN proposes to conduct a carbon mass balance to assess ultimate biodegradation C plastic
-
Cc02
+ Cbiomass
+ Cresidual
+ Csoluble
polymeric material
The carbon content of the plastic can, in principle, either be degraded to carbon dioxide (C&J, or water-soluble intermediates biomass (Cbiomass) soluble). The percentage of residual, non-biode(C graded material can be calculated from the difference to the initial plastic carbon (Cplastic). The carbon content of the plastic can either be derived from its chemical structure or from experimental (elemental) analysis. Ultimate biodegradation is reached if 100% of the plastic’s carbon is degraded to carbon dioxide and biomass. Currently, a carbon mass balance can only be conducted in an aquatic system where the plastic is the sole source of carbon. In compost - a heterogeneous matrix with various carbon sources - the build-up of new biomass and the formation of degradation intermediates cannot be determined with sufficient accuracy. As a consequence, complete biodegradation in compost only can be guaranteed if 100% mineralization of the plastic’s carbon to carbon dioxide is measured (CcoZ = Cpiastic).
Assessment of biodegradability of plastics
Automated laboratory test system
At the Fraunhofer-Institute for Food Process Engineering and Packaging (FhILV) an automated laboratory test system was developed to measure the aerobic biodegradability of plastics (Fig. 2). Biodegradation tests are conducted in an aquatic system as well as in a solid-state compost environment. The biodegradation process is continuously monitored by measuring the microbial carbon dioxide formation and oxygen consumption. Currently, the test system consists of 16 bioreactors with a capacity of 1.5 1 each for the aquatic test and 51 each for the fixed-bed test, respectively. The bioreactors are temperaturecontrolled to simulate the natural self-heating of a composting process. The aeration rate of each reactor is automatically controlled based on the measured oxygen concentration to avoid anaerobic conditions in the bioreactors. In standard aquatic biodegradation tests aeration is performed with air free of CO2 (e.g. ASTM 5209-92). However, the absence of carbon dioxide in the test medium in some cases may delay the onset of biodegradation (Pitter & Chudoba, 1990; Gilbert & Watson, 1977). Therefore, aeration of the bioreactors is performed with air of ambient C02-concentration.
MATERIAL
AND METHODS
Poly-e-caprolacton (PCL) (Tone@ P-767, Union Carbide) was chosen as a test polymer. PCL is a synthetic aliphatic homopolyester with a molecular weight of Mn = 43 OOODa and a melting temperature of 60°C. PCL is used frequently in the
81
preparation of biodegradable polymer blends due to its compatibility with other polymers. PCL was applied as a powder which was separated into different particle size fractions by sieving to investigate the influence of particle size on the biodegradation rate. As a second test material, a plastic bag made from a blend of corn starch and synthetic polymers was investigated. This bag is used for the collection of bio-waste from households and is supposed to be composted together with the bio-waste. Mature compost from an industrial composting plant was used for the fixed-bed system. The compost had a volatile solids content of 50% (w/w) of dry solids, a C/N-ratio of 12 and a pH of 7.6. The percentage of carbon and nitrogen in the compost was measured with a total organic carbon analyzer (Model Coulomat 702, Stroehlein Instr., Germany) and a total nitrogen analyzer (Model Macro N, Foss-Heraeus, Germany). The moisture content of the compost was adjusted to 50% (w/w). For the aquatic test, a mineral salt solution was used that provided all essential nutrients for microbial growth, but no source of carbon (Table 1). The medium was inoculated with an aqueous eluate from the above mentioned mature compost. The inoculum represented a broad mixed microbial flora specific for compost. For the aquatic test and the fixed-bed test, a time-dependent temperature profile according to ASTM 5338-92 was applied simulating the natural self-heating of a composting process (Table 2). Additionally, the aquatic test was conducted at a constant temperature of 30°C. In the aquatic system a carbon mass balance was performed. Biomass growth was assessed by measuring the intracellular biomass protein
Biodegradable plastic (powder, film) Mineral salts solution Mixed microbial population (eluate from compost)
Biodegradable plastic
Fig. 2. Automated
laboratory
test system for measuring
biodegradability
of plastics.
88
A. Starnecker, M. Menner
according to a modified method of Lowry (Sperandio & Puechner, 1993). An average protein and carbon content of 50% (w/w) of the cell dry Table 1. Composition
mass was assumed (Schlegel, 1992). Water-soluble intermediates were determined by measuring the concentration of dissolved organic carbon (DOC) with a total organic carbon analyzer (Model TOC500, Shimadzu Corp., Japan) after filtration through a 0.2 pm membrane filter.
of the Mineral Salt Solution
Component
Concentration
NazHP04 * 2 HZ0 KH2P04
[g 1-‘I
6.97 3.75 4.0 0.2 0.13 0.002 7 0.001
(m)2so4
MgS04 * 7 H20 CaC12 * 2 H20 FeCls * 6 HZ0 Trace elements solution (Drews, 1968)
RESULTS
Carbon balance
Figure 3 shows the time course of microbial C02formation and carbon mineralization during biodegradation of PCL in the aquatic system. A microbial adaption (lag-) phase of approximately one day was observed followed by an exponential phase of microbial CO*-formation. The CO*evolution rate reached a maximum after two days of incubation. After eleven days, 80% of the polymer carbon was degraded to carbon dioxide. After a duration of 2, 3, 4 and 9 days, a carbon mass balance was performed. Figure 4 reveals that
Table 2. Time-dependent Temperature Profile for the Fixedbed Test and the Aquatic Test Time [days]
Temperature
O-l l-5 5-28 2845
AND DISCUSSION
[“Cl
35 58
100 /
-- 90
;:I’;:;
d
I,2 --
2 6 t
I-0.9 --
E & e
0,6 --
8
0,4 -0,2 --
~,~
~.
9
10
01 0
1
2
3
4
5
6
7
8
11
Time I d
Fig. 3. Microbial
COz-formation
c .o 2 'C
rate and carbon mineralization in the aquatic test Polymer: PCL powder, 18&200pm, concentration; 1.Og l-l, temperature: 30°C.
76
W C-residual polymer
60
(calculated) q C-soluble
g66
0 C-biomass m c-co2
546 e J
30
2 days
Fig. 4. Carbon distribution
3 days
4 days
during microbial degradation
9 days
of PCL in the aquatic system.
test
Assessment of biodegradability of plastics
a significant portion of the plastic’s carbon was incorporated into biomass. After 3 days biomass carbon reached a maximum of 38% of the initial polymer carbon. Summing up the percentage of carbon dioxide and biomass carbon a biodegradation degree of approximately 80% is reached already after 3 days. After 9 days 78% of the plastic was mineralized to carbon dioxide, 13% was incorporated into biomass and 9% was measured as water-soluble metabolites. From the carbon balance it is apparent that after 9 days no more residual polymeric carbon was left.
(increasing surface area) of the polyester powder. However, the degree of mineralization after 20 days was similar for all particle size fractions. Consequently, a comparison of the degradation rate of different biodegradable plastics is only valid if the test specimens have similar geometry (powder, film) and surface area. Comparison of compost simulation test and aquatic test Figures 6-8 show the results of a biodegradation test for a starch-based plastic bag in compost in comparison with the aquatic test. Figure 6 is a plot of the microbial carbon dioxide production in the compost system as a function of time. The addition of the test material caused a significant increase in the microbial respiration rate, compared to the blank compost assay without test material. The
Influence of particle size on biodegradation rate Figure 5 shows the rate of biodegradation for different particle-size fractions of PCL powder in the aquatic test. The biodegradation rate increased significantly with decreasing particle size
0
2
4
6
8
10 Time
Fig. 5. Biodegradation
12
I
14
16
18
20
d
of different particle size fractions of PCL powder in the aquatic system.
20 Time
Fig. 6. Time course of C02-formation
89
25 I d
in the compost simulation test (test material; starch-based plastic bag, film thickness: 50 pm) (results are mean values from triple assays).
A. Starnecker, M. Menner
90
0
1
0
10
5
The
I
45
40
35
30
25
20
15
d
Fig. 7. Carbon mineralization in the compost system with temperature profile in comparison to the aquatic system with tempera ture profile and constant temperature of 30°C (Test material: starch-based plastic bag, film thickness: 50 pm).
temperature profile
constant temperature ( T = 30°C)
Fig. 8. Influence of temperature
( T = 36-66~60-3SC )
on the carbon balance in the aquatic system after a test duration of 45 days.
80
10 0 0
5
10
15
20 Time
Fig. 9. Biodegradation
30
25 I
35
40
45
d
in compost in comparison with a carbon-free fixed-bed (Test material: starch-based plastic bag, 50 pm).
Assessment
of biodegradability
influence of the temperature profile on the microbial respiration rate can be seen clearly. COZ-production due to microbial degradation of the test material was determined from the difference in CO*-formation to the blank compost assay. Figure 7 shows the time course of carbon mineralization in compost in comparison to the aquatic system. After 45 days, 68% of the plastic’s carbon was degraded to carbon dioxide in the compost environment. An equal degree of mineralization was reached in the aquatic test if the same temperature profile was applied. However, the biodegradation rate was significantly higher in the aquatic system. A carbon mineralization of nearly 70% was measured already after 15 days, whereas it took more than double of the time in the compost system to reach the same degree of mineralization. Conducting the aquatic test at a constant temperature of 30°C a reduced rate of biodegradation was observed compared to the aquatic system with time-dependent temperature profile. Only 62% of the plastic’s carbon was mineralized after 45 days instead of 70% when applying the temperature profile (Fig. 7). A carbon balance revealed that the ratio of microbial carbon dioxide formation to biomass growth was dependent on temperature (Fig. 8). After 45 days less than half of the biomass was detected in the aquatic system with temperature profile compared to the assay with constant temperature. However, the degree of biodegradation - combining carbon dioxide and biomass estimations - amounted to a similar value of 80 or 77%, respectively. According to Fig. 8, approximately 20% of the plastic’s carbon was still not degraded after 45 days. Further investigations are under way to find out whether this portion of the plastic will be biodegraded at extended test duration. The results of this experiment confirm that an assessment of plastic biodegradation only based on the measurement of microbial carbon dioxide formation is not sufficient. In order to assess the completeness of biodegradation the portion of the plastic’s carbon which is incorporated into new biomass has to be determined. Fixed-bed test with inert packing material As already mentioned, ultimate biodegradation in compost can only be measured if 100% mineralization of the plastic’s carbon to carbon dioxide occurs. So far, however, it has not been investi-
of plastics
91
gated whether a part of the plastic’s carbon may be incorporated into biologically-stable humic substances. Consequently, 100% mineralization could not be measured even for completely biodegradable materials. On the other hand, it is reported that background C02-evolution of compost or soil may be enhanced by the addition of a biodegradable material (Sharabi & Bartha, 1993). Carbon dioxide formation may be measured that does not result from microbial degradation of the test material. Therefore, at our institute a new lab-test was developed, which replaces the heterogeneous matrix ‘compost’ by a biologically inert, carbon-free material. Screening tests with different, porous materials indicated that environmental conditions similar to compost could be created with those materials (e.g. water activity, porosity, storage of nutrients). Figure 9 shows the time course of microbial degradation of a starch-based plastic bag in a tixed-bed with inert packing material in comparison to compost. The inert packing material was inoculated with an aqueous eluate from compost. Figure 9 demonstrates that similar biodegradation rates could be observed compared to the compost system. Since the plastic is the sole source of carbon in the fixed-bed, it will be possible to conduct a carbon mass balance for the assessment of ultimate biodegradation. REFERENCES American Society for Testing and Materials (ASTM) (1992). Standard Test Method for Determining the Aerobic Biodegradation of Plastic Materials in the Presence of Municipal Sewage Sludge. ASTM D5209-92, Philadelphia,
USA. American Society for Testing and Materials (ASTM) (1992). Standard Test Method for Determining the Aerobic Biodegradation of Plastic Materials under Controlled composting Conditions. ASTM D5338-92, Philadelphia,
USA. Drews, G. (1968). Mikrobiologisches Praktikum fur Naturwissenschaftler. Springer, Berlin. Gilbert, P. A. & Watson, G. K. (1977). Biodegradability testing and its relevance to environmental acceptability. Tenside Detergents, 14, 171-177.
Llnderarbeitsgemeinschaft Abfall (LAGA) Guideline M 10 (1995). Qualitdtskriterien und Anwendungsempfehlungen fur Kompost (‘Quality criteria and recommendations for the use of compost’). Pantke, M. (1994). Biologisch abbaubare Kunststoffe Begriffsbestimmung und Normung von Prtifverfahren. Kunststoffe, 84, 1090. Pitter, P. & Chudoba, J. (1990). Biodegradability of Organic Substances in the Aquatic Environment. CRC Press, Boca Raton, FL. Schlegel, H. G. (1992). Allgemeine Mikrobiologie. Georg Thieme, Stuttgart. Schroeter, J., DeWilde, B., Gottschall, R., Koch, R. &
92
A. Starnecker, M. Menner
Starnecker, A. (1994). Assessment of compostability of polymeric materials. Draft proposal DIN FNK 103.3. Sharabi, N. E. & Bartha, R. (1993). Testing of some assumptions about biodegradability in soil as measured by carbon dioxide evolution. Appl. Environ. Microbial., 59, 1201-1205. Sperandio, A. & Ptichner, P. (1993). Bestimmung der
Gesamtproteine als Biomasse-Parameter in wll3rigen Kulturen und auf Tragermaterialien aus Bio-Reaktoren. Wasser, Abwasser., 134,482-485.
Streff, L., Gottschall, R., Bidlingmaier, W. & Vogtmann, H. (1994). Biodegradable plastics in cornposting. Conference papers, Rocky Mountain Conf. of Composting, Denver (in press).
Biodeterioration
& Biodegradation
(1996) 85-92
Copyright 0 1996 Elsevier Science Limited Printed in Great Britain. All rights reserved 0964-8305/96 $15.00 + 0.00 0964-8305(95)00089-5
ELSEVIER
Assessment of Biodegradability of Plastics under Simulated Cornposting Conditions in a Laboratory Test System Andreas Starnecker’ & Michael Menner Fraunhofer-Institute for Food Process Engineering and Packaging, Steinerstr. 15, 81369 Munich, Germany
An automated laboratory-scale test system was developed for measuring the aerobic biodegradability of degradable plastics under simulated composting conditions. Biodegradation was monitored by measuring microbial carbon dioxide formation and oxygen consumption. Completeness of biodegradation was assessed in an aquatic test by conducting a carbon mass balance. The percentage of plastic carbon degraded to carbon dioxide, biomass and watersoluble byproducts were determined. The rate of biodegradation under simulated composting conditions was measured in a fixed-bed system with mature compost. A time-dependent temperature profile was applied to simulate the natural selfheating of a composting process. The aquatic test was conducted at a constant temperature of 30°C as well as with a temperature profile. The rate of biodegradation was significantly higher in the aqueous environment. Equal degrees of mineralization were reached in the aquatic and the fixed-bed system only if the same temperature profile was applied. Conducting the aquatic test at a constant temperature of 30°C less microbial carbon dioxide formation was observed. However, a carbon mass balance revealed that taking into consideration the portion of the plastic’s carbon incorporated into biomass, a similar degree of biodegradation was reached. Consequently, the measurement of microbial carbon dioxide production is not sufficient to assess the extent of biodegradation of plastics. In the heterogeneous matrix ‘compost’, it is not feasible to assess the completeness of biodegradation due to limited possibilities to analyze degradation intermediates and biomass growth. Therefor& a new fixed-bed system with an inert, carbon-free packing material was developed. The inert material was inoculated with an aqueous eluate from compost. First results showed biodegradation rates similar to a compost environment. Copyright 0 1996 Elsevier Science Limited.
admission criteria for input In Germany, materials of an industrial composting process are specified in the German federal states guidelines for compost (LAGA M 10, 1995). Only input materials which are completely biodegradable are accepted for cornposting. Non native-organic materials have to pass a suitability test. Figure 1 shows the current test procedure for the assessment of compostability as proposed by the working committee ‘Biodegradable Plastics’ FNK 103.3 of the German Institute for Normification (DIN) (Schroeter et al., 1994). First, there is a basic chemical product analysis to identify pollutants (e.g. heavy metals, polychlorinated aromatics) or non-biodegradable components of the plastic. Second, two laboratoryscale tests for the assessment of complete aerobic
INTRODUCTION Composting represents an increasingly important route of disposal for the organic fraction of municipal solid waste. In the future also, materials that are not of native-organic origin are planned as input materials for composting facilities. One important group of those man-made materials are biodegradable plastics. Unlike most of the can be commodity plastics, these polymers degraded and utilized as a source of .carbon and energy by a variety of microorganisms, if they get into an environment favourable for microbial activity (e.g. compost, soil, waste water). ‘Paper presented at the fourth meeting of the Biodegradable Plastics Group, International Biodeterioration Research Group, Winchester, UK, May 3-5, 1995. 85
86
A. Starnecker, M. Menner
Laboratory-scale
Pilot plant-scale
+Lzzq field-proven model experiment
* Field testing
Fig. 1. Test procedure for the assessment of compostability
biodegradability are proposed: an aquatic screening test and a solid-state compost simulation test. Existing biodegradability lab-tests for plastics (e.g. ASTM 5209-92, ASTM 5338-92) are currently being modified to experimentally prove completeness of biodegradation. The next two steps of the test procedure examine the suitability of the plastic material for cornposting. This is assessed under controlled conditions in pilot plant-scale cornposting units and under practical conditions in technical cornposting plants (Streff et al., 1994). Finally, utilization properties and ecotoxicology of the compost obtained by adding a certain percentage of the plastic as input material are investigated. Ultimate biodegradation Microbial degradation of plastics is initiated by the secretion of enzymes, which cause a chain cleavage of the polymer into oligomers or monomers, respectively. These water-soluble enzymatic cleavage products are absorbed into the microbial cell where they are metabolized. End-products of the aerobic metabolism are basically carbon dioxide and water. A portion of the plastic’s carbon is utilized for the build-up of new biomass. According to the definition of the DIN, a plastic is called biodegradable if all of its organic components undergo a total biodegradation to naturally occurring metabolic end-products (Pantke, 1994). This process is called ultimate
(draft proposal DIN FNK 103.3).
biodegradation (Gilbert & Watson, 1977). Biomass itself eventually will be degraded to carbon dioxide. However, over the usual time scale of a biodegradation test, complete mineralization to carbon dioxide generally will not be observed. Therefore, the DIN proposes to conduct a carbon mass balance to assess ultimate biodegradation C plastic
-
Cc02
+ Cbiomass
+ Cresidual
+ Csoluble
polymeric material
The carbon content of the plastic can, in principle, either be degraded to carbon dioxide (C&J, or water-soluble intermediates biomass (Cbiomass) soluble). The percentage of residual, non-biode(C graded material can be calculated from the difference to the initial plastic carbon (Cplastic). The carbon content of the plastic can either be derived from its chemical structure or from experimental (elemental) analysis. Ultimate biodegradation is reached if 100% of the plastic’s carbon is degraded to carbon dioxide and biomass. Currently, a carbon mass balance can only be conducted in an aquatic system where the plastic is the sole source of carbon. In compost - a heterogeneous matrix with various carbon sources - the build-up of new biomass and the formation of degradation intermediates cannot be determined with sufficient accuracy. As a consequence, complete biodegradation in compost only can be guaranteed if 100% mineralization of the plastic’s carbon to carbon dioxide is measured (CcoZ = Cpiastic).
Assessment of biodegradability of plastics
Automated laboratory test system
At the Fraunhofer-Institute for Food Process Engineering and Packaging (FhILV) an automated laboratory test system was developed to measure the aerobic biodegradability of plastics (Fig. 2). Biodegradation tests are conducted in an aquatic system as well as in a solid-state compost environment. The biodegradation process is continuously monitored by measuring the microbial carbon dioxide formation and oxygen consumption. Currently, the test system consists of 16 bioreactors with a capacity of 1.5 1 each for the aquatic test and 51 each for the fixed-bed test, respectively. The bioreactors are temperaturecontrolled to simulate the natural self-heating of a composting process. The aeration rate of each reactor is automatically controlled based on the measured oxygen concentration to avoid anaerobic conditions in the bioreactors. In standard aquatic biodegradation tests aeration is performed with air free of CO2 (e.g. ASTM 5209-92). However, the absence of carbon dioxide in the test medium in some cases may delay the onset of biodegradation (Pitter & Chudoba, 1990; Gilbert & Watson, 1977). Therefore, aeration of the bioreactors is performed with air of ambient C02-concentration.
MATERIAL
AND METHODS
Poly-e-caprolacton (PCL) (Tone@ P-767, Union Carbide) was chosen as a test polymer. PCL is a synthetic aliphatic homopolyester with a molecular weight of Mn = 43 OOODa and a melting temperature of 60°C. PCL is used frequently in the
81
preparation of biodegradable polymer blends due to its compatibility with other polymers. PCL was applied as a powder which was separated into different particle size fractions by sieving to investigate the influence of particle size on the biodegradation rate. As a second test material, a plastic bag made from a blend of corn starch and synthetic polymers was investigated. This bag is used for the collection of bio-waste from households and is supposed to be composted together with the bio-waste. Mature compost from an industrial composting plant was used for the fixed-bed system. The compost had a volatile solids content of 50% (w/w) of dry solids, a C/N-ratio of 12 and a pH of 7.6. The percentage of carbon and nitrogen in the compost was measured with a total organic carbon analyzer (Model Coulomat 702, Stroehlein Instr., Germany) and a total nitrogen analyzer (Model Macro N, Foss-Heraeus, Germany). The moisture content of the compost was adjusted to 50% (w/w). For the aquatic test, a mineral salt solution was used that provided all essential nutrients for microbial growth, but no source of carbon (Table 1). The medium was inoculated with an aqueous eluate from the above mentioned mature compost. The inoculum represented a broad mixed microbial flora specific for compost. For the aquatic test and the fixed-bed test, a time-dependent temperature profile according to ASTM 5338-92 was applied simulating the natural self-heating of a composting process (Table 2). Additionally, the aquatic test was conducted at a constant temperature of 30°C. In the aquatic system a carbon mass balance was performed. Biomass growth was assessed by measuring the intracellular biomass protein
Biodegradable plastic (powder, film) Mineral salts solution Mixed microbial population (eluate from compost)
Biodegradable plastic
Fig. 2. Automated
laboratory
test system for measuring
biodegradability
of plastics.
88
A. Starnecker, M. Menner
according to a modified method of Lowry (Sperandio & Puechner, 1993). An average protein and carbon content of 50% (w/w) of the cell dry Table 1. Composition
mass was assumed (Schlegel, 1992). Water-soluble intermediates were determined by measuring the concentration of dissolved organic carbon (DOC) with a total organic carbon analyzer (Model TOC500, Shimadzu Corp., Japan) after filtration through a 0.2 pm membrane filter.
of the Mineral Salt Solution
Component
Concentration
NazHP04 * 2 HZ0 KH2P04
[g 1-‘I
6.97 3.75 4.0 0.2 0.13 0.002 7 0.001
(m)2so4
MgS04 * 7 H20 CaC12 * 2 H20 FeCls * 6 HZ0 Trace elements solution (Drews, 1968)
RESULTS
Carbon balance
Figure 3 shows the time course of microbial C02formation and carbon mineralization during biodegradation of PCL in the aquatic system. A microbial adaption (lag-) phase of approximately one day was observed followed by an exponential phase of microbial CO*-formation. The CO*evolution rate reached a maximum after two days of incubation. After eleven days, 80% of the polymer carbon was degraded to carbon dioxide. After a duration of 2, 3, 4 and 9 days, a carbon mass balance was performed. Figure 4 reveals that
Table 2. Time-dependent Temperature Profile for the Fixedbed Test and the Aquatic Test Time [days]
Temperature
O-l l-5 5-28 2845
AND DISCUSSION
[“Cl
35 58
100 /
-- 90
;:I’;:;
d
I,2 --
2 6 t
I-0.9 --
E & e
0,6 --
8
0,4 -0,2 --
~,~
~.
9
10
01 0
1
2
3
4
5
6
7
8
11
Time I d
Fig. 3. Microbial
COz-formation
c .o 2 'C
rate and carbon mineralization in the aquatic test Polymer: PCL powder, 18&200pm, concentration; 1.Og l-l, temperature: 30°C.
76
W C-residual polymer
60
(calculated) q C-soluble
g66
0 C-biomass m c-co2
546 e J
30
2 days
Fig. 4. Carbon distribution
3 days
4 days
during microbial degradation
9 days
of PCL in the aquatic system.
test
Assessment of biodegradability of plastics
a significant portion of the plastic’s carbon was incorporated into biomass. After 3 days biomass carbon reached a maximum of 38% of the initial polymer carbon. Summing up the percentage of carbon dioxide and biomass carbon a biodegradation degree of approximately 80% is reached already after 3 days. After 9 days 78% of the plastic was mineralized to carbon dioxide, 13% was incorporated into biomass and 9% was measured as water-soluble metabolites. From the carbon balance it is apparent that after 9 days no more residual polymeric carbon was left.
(increasing surface area) of the polyester powder. However, the degree of mineralization after 20 days was similar for all particle size fractions. Consequently, a comparison of the degradation rate of different biodegradable plastics is only valid if the test specimens have similar geometry (powder, film) and surface area. Comparison of compost simulation test and aquatic test Figures 6-8 show the results of a biodegradation test for a starch-based plastic bag in compost in comparison with the aquatic test. Figure 6 is a plot of the microbial carbon dioxide production in the compost system as a function of time. The addition of the test material caused a significant increase in the microbial respiration rate, compared to the blank compost assay without test material. The
Influence of particle size on biodegradation rate Figure 5 shows the rate of biodegradation for different particle-size fractions of PCL powder in the aquatic test. The biodegradation rate increased significantly with decreasing particle size
0
2
4
6
8
10 Time
Fig. 5. Biodegradation
12
I
14
16
18
20
d
of different particle size fractions of PCL powder in the aquatic system.
20 Time
Fig. 6. Time course of C02-formation
89
25 I d
in the compost simulation test (test material; starch-based plastic bag, film thickness: 50 pm) (results are mean values from triple assays).
A. Starnecker, M. Menner
90
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Fig. 7. Carbon mineralization in the compost system with temperature profile in comparison to the aquatic system with tempera ture profile and constant temperature of 30°C (Test material: starch-based plastic bag, film thickness: 50 pm).
temperature profile
constant temperature ( T = 30°C)
Fig. 8. Influence of temperature
( T = 36-66~60-3SC )
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influence of the temperature profile on the microbial respiration rate can be seen clearly. COZ-production due to microbial degradation of the test material was determined from the difference in CO*-formation to the blank compost assay. Figure 7 shows the time course of carbon mineralization in compost in comparison to the aquatic system. After 45 days, 68% of the plastic’s carbon was degraded to carbon dioxide in the compost environment. An equal degree of mineralization was reached in the aquatic test if the same temperature profile was applied. However, the biodegradation rate was significantly higher in the aquatic system. A carbon mineralization of nearly 70% was measured already after 15 days, whereas it took more than double of the time in the compost system to reach the same degree of mineralization. Conducting the aquatic test at a constant temperature of 30°C a reduced rate of biodegradation was observed compared to the aquatic system with time-dependent temperature profile. Only 62% of the plastic’s carbon was mineralized after 45 days instead of 70% when applying the temperature profile (Fig. 7). A carbon balance revealed that the ratio of microbial carbon dioxide formation to biomass growth was dependent on temperature (Fig. 8). After 45 days less than half of the biomass was detected in the aquatic system with temperature profile compared to the assay with constant temperature. However, the degree of biodegradation - combining carbon dioxide and biomass estimations - amounted to a similar value of 80 or 77%, respectively. According to Fig. 8, approximately 20% of the plastic’s carbon was still not degraded after 45 days. Further investigations are under way to find out whether this portion of the plastic will be biodegraded at extended test duration. The results of this experiment confirm that an assessment of plastic biodegradation only based on the measurement of microbial carbon dioxide formation is not sufficient. In order to assess the completeness of biodegradation the portion of the plastic’s carbon which is incorporated into new biomass has to be determined. Fixed-bed test with inert packing material As already mentioned, ultimate biodegradation in compost can only be measured if 100% mineralization of the plastic’s carbon to carbon dioxide occurs. So far, however, it has not been investi-
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gated whether a part of the plastic’s carbon may be incorporated into biologically-stable humic substances. Consequently, 100% mineralization could not be measured even for completely biodegradable materials. On the other hand, it is reported that background C02-evolution of compost or soil may be enhanced by the addition of a biodegradable material (Sharabi & Bartha, 1993). Carbon dioxide formation may be measured that does not result from microbial degradation of the test material. Therefore, at our institute a new lab-test was developed, which replaces the heterogeneous matrix ‘compost’ by a biologically inert, carbon-free material. Screening tests with different, porous materials indicated that environmental conditions similar to compost could be created with those materials (e.g. water activity, porosity, storage of nutrients). Figure 9 shows the time course of microbial degradation of a starch-based plastic bag in a tixed-bed with inert packing material in comparison to compost. The inert packing material was inoculated with an aqueous eluate from compost. Figure 9 demonstrates that similar biodegradation rates could be observed compared to the compost system. Since the plastic is the sole source of carbon in the fixed-bed, it will be possible to conduct a carbon mass balance for the assessment of ultimate biodegradation. REFERENCES American Society for Testing and Materials (ASTM) (1992). Standard Test Method for Determining the Aerobic Biodegradation of Plastic Materials in the Presence of Municipal Sewage Sludge. ASTM D5209-92, Philadelphia,
USA. American Society for Testing and Materials (ASTM) (1992). Standard Test Method for Determining the Aerobic Biodegradation of Plastic Materials under Controlled composting Conditions. ASTM D5338-92, Philadelphia,
USA. Drews, G. (1968). Mikrobiologisches Praktikum fur Naturwissenschaftler. Springer, Berlin. Gilbert, P. A. & Watson, G. K. (1977). Biodegradability testing and its relevance to environmental acceptability. Tenside Detergents, 14, 171-177.
Llnderarbeitsgemeinschaft Abfall (LAGA) Guideline M 10 (1995). Qualitdtskriterien und Anwendungsempfehlungen fur Kompost (‘Quality criteria and recommendations for the use of compost’). Pantke, M. (1994). Biologisch abbaubare Kunststoffe Begriffsbestimmung und Normung von Prtifverfahren. Kunststoffe, 84, 1090. Pitter, P. & Chudoba, J. (1990). Biodegradability of Organic Substances in the Aquatic Environment. CRC Press, Boca Raton, FL. Schlegel, H. G. (1992). Allgemeine Mikrobiologie. Georg Thieme, Stuttgart. Schroeter, J., DeWilde, B., Gottschall, R., Koch, R. &
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Starnecker, A. (1994). Assessment of compostability of polymeric materials. Draft proposal DIN FNK 103.3. Sharabi, N. E. & Bartha, R. (1993). Testing of some assumptions about biodegradability in soil as measured by carbon dioxide evolution. Appl. Environ. Microbial., 59, 1201-1205. Sperandio, A. & Ptichner, P. (1993). Bestimmung der
Gesamtproteine als Biomasse-Parameter in wll3rigen Kulturen und auf Tragermaterialien aus Bio-Reaktoren. Wasser, Abwasser., 134,482-485.
Streff, L., Gottschall, R., Bidlingmaier, W. & Vogtmann, H. (1994). Biodegradable plastics in cornposting. Conference papers, Rocky Mountain Conf. of Composting, Denver (in press).