Polymer Degradation and Stability xxx (2013) 1e9

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Polymer Degradation and Stability

journal homepage: www.elsevier.com/locate/polydegstab

The influence of biotic and abiotic factors on the rate of degradation of

poly(lactic) acid (PLA) coupons buried in compost and soil
Mehlika Karamanlioglu, Geoffrey D. Robson*
Faculty of Life Sciences, Oxford Road, Michael Smith Building, University of Manchester, Manchester M13 9PT, UK

a r t i c l e i n f o a b s t r a c t

Article history: Poly(lactic) acid (PLA) is a compostable biopolymer and has been commercialised for the for the
Received 27 May 2013 manufacture of short-shelf life products. As a result, increasing amounts of PLA are entering waste
Received in revised form management systems and the environment; however, the degradation mechanism is unclear. While
28 June 2013
hydrolysis of the polymer occurs abiotically at elevated temperature in the presence of water, potential
Accepted 3 July 2013
catalytic role for microbes in this process is yet to be established. In this study, we examined the
Available online xxx
degradation of PLA coupons from commercial packaging at a range of temperatures (25
, 37
, 45
, 50

and 55
C) in soil and compost and compared with the degradation rates in sterile aqueous conditions by
Keywords:
Poly(lactic) acid
measuring loss of tensile strength and molecular weight (Mw). In addition, in order to assess the possible
PLA influence of abiotic soluble factors in compost and soil on degradation of PLA, degradation rates in
Degradation microorganism-rich compost and soil were compared with sterile compost and soil extract at 50
C.
Compost Temperature was determined to be the key parameter in PLA degradation and degradation rates in
microorganism-rich compost and soil were faster than in sterile water at temperatures 45
and 50
C
determined by tensile strength and Mw loss. Furthermore, all tensile strength was lost faster after 30 and
36 days in microorganism-rich compost and soil, respectively, than in sterile compost and soil extract, 57
and 54 days, respectively at 50
C. Significantly more Mw, 68% and 64%, was lost in compost and soil,
respectively than in compost extract, Mw, 53%; and in soil extract, 57%. Therefore, degradation rates were
faster in microorganism-rich compost and soil than in sterile compost and soil extract, which contained
the abiotic soluble factors of compost and soil at 50
C. These comparative studies support a direct role
for microorganisms in PLA degradation at elevated temperatures in humid environments. No change in
tensile strength or Mw was observed either 25
or 37
C after 1 year suggesting that accumulation of PLA
in the environment may cause future pollution issues.
Ó 2013 Elsevier Ltd. All rights reserved.

1. Introduction conventional plastics are derived from petrochemicals and their

Conventional plastics such as polyethylene terephthalate (PET),
polyvinylchloride (PVC), polyethylene (PE), polypropylene (PP),
polystyrene (PS) and polyamide (PA) have been used for decades in
a diverse range of applications as they are easily processed,
available in large quantities, are inexpensive and have favourable
mechanical characteristics [1e9]. However, many are poorly
degraded in the environment causing widespread environmental
pollution and putting pressure on diminishing landfill capacity
[2,9e14]. In many instances, recycling has not been developed
sufficiently to keep up with increasing amounts of plastic
consumption and demand [7]. In addition, the vast majority of
* Corresponding author. Tel.: þ44 0 1612755048.
E-mail address: geoff.robson@manchester.ac.uk (G.D. Robson).
manufacture is dependent on non-renewable fossil fuels
[2,4,15,16]. Consequently, research has focused on the develop-
ment of novel plastics derived from renewable resources that are
susceptible to biodegradation in the environment [16e18]. One
such plastic is poly(lactic acid) (PLA), a synthetic aliphatic poly-
ester with a hydrolysable backbone that undergoes chemical
hydrolysis at elevated temperatures in the presence of water and
uses starch as a renewable feedstock [19e23]. Lactic acid, the
precursor of PLA, is predominantly obtained by bacterial fermen-
tation of starch [24e26]. PLA has many favourable properties; it is
easy to fabricate, is not toxic or carcinogenetic, is biocompatible,
has high mechanical strength and is compostable [4,18e21,24,27e
32]. Originally, PLA was restricted to medical applications
[19,33,34] but in recent years has been commercialised for the
manufacture of compostable food-packaging films, bags, con-
tainers and fibres [20,24,25,28].

0141-3910/$ e see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.polymdegradstab.2013.07.004

Please cite this article in press as: Karamanlioglu M, Robson GD, The influence of biotic and abiotic factors on the rate of degradation of
poly(lactic) acid (PLA) coupons buried in compost and soil, Polymer Degradation and Stability (2013), http://dx.doi.org/10.1016/
j.polymdegradstab.2013.07.004
2 M. Karamanlioglu, G.D. Robson / Polymer Degradation and Stability xxx (2013) 1e9
Degradation of PLA under composting conditions by chemical
hydrolysis is well established, however, the role of microorganisms
in PLA degradation in the environment is still unclear. The most
accepted mechanism of PLA degradation involves a two-step
mechanism involving chemical hydrolysis in the presence of wa-
ter at elevated temperatures followed by microbial degradation in
which microorganisms mineralize polymer break-down products
generating carbon dioxide under aerobic conditions and methane
under anaerobic conditions [24,26,28,29,31,35e39]. It has been
reported that microorganisms do not enhance PLA degradation and
that polymer cleavage proceeds solely through abiotic hydrolysis of
ester linkages in the presence or absence of microorganisms [40].
However, other studies provide evidence suggesting that microbial
enzymes exist that are capable of directly degrading high molecular
weight PLA [41e43] and Sangwan and Wu [36] highlighted the lack
of clarity on the potential role of microorganisms during PLA
degradation.
In this study we investigated the degradation of PLA under
different environmental conditions to determine the relative
importance of microbiological and chemical factors in PLA degra-
dation by comparing the rate of tensile strength loss and molecular
weight decrease in microorganism-rich compost and soil and in
sterile water at 25
, 37
, 45
, 50
and 55
C. Furthermore, the in-
fluence of abiotic soluble factors in compost and soil on degrada-
tion of PLA was assessed by comparing degradation rates in
microorganism-rich compost and soil with sterile compost and
soil extract at 50
C.
2. Materials and methods
2.1. Polylactic acid (PLA)
High molecular weight PLA (Mw 159,000  5800 g/mol, deter-
mined in this study) was obtained in the form of precast food
containers produced from semi-crystalline PLA resins (IngeoÔ
Biopolymer, Grade 2003D), manufactured by NatureWorks LLC
(USA) and distributed through VegWare, Edinburgh, UK. According
to the supplier, PLA resins contained 4% D-lactic acid isomer, a
density of 1.24 g/cm3, melting temperature of 145e170
C, glass
transition temperature (Tg) of 55e58
C, and crystallinity of 35%.
2.2. Compost and soil source
Commercial soil was purchased from Online Turf, Lancashire, UK
and mature green compost from The Compost Shop, UK. Compost
and soil were sieved through a 2 mm mesh to remove particles
>2 mm prior to use and stored in sealed containers at room
temperature.
2.3. Preparation of compost and soil extracts
Compost or soil extracts were prepared according to a modified
protocol of Alef [44] by mixing 1 kg of dried compost or soil with 1 l
of tap water and left at room temperature for 1 h with occasional
shaking. The supernatant was filtered through J-cloth two times to
remove large particles and then through filter paper (Whatman No
6, UK). The supernatant was finally filter sterilized through a
0.22 mm membrane (Sterilin, Steritop, Merck, Germany) with a
vacuum pump and extracts stored at 4
C until use.
2.4. Determination of the water holding capacity (WHC) and
moisture content of compost and soil
The water holding capacity (WHC) of compost or soil was
determined by a modified protocol of Forster [45] by filling a
Please cite this article in press as: Karamanlioglu M, Robson GD, The influence of biotic and abiotic factors on the rate of degradation of
poly(lactic) acid (PLA) coupons buried in compost and soil, Polymer Degradation and Stability (2013), http://dx.doi.org/10.1016/
j.polymdegradstab.2013.07.004
strainer with compost or soil and saturating with distilled water
before allowing excess water to drain. The saturated compost or
soil was weighed on filter paper previously dried to constant
weight to give the saturated weight (wsat). The sample was then
dried in an oven at 105
C to constant weight to determine the
dry weight, (wdry). The WHC was calculated according to Eq. (1).
In order to measure moisture content, compost or soil samples
were placed onto filter papers previously dried to constant
weight and their weights were measured to give the initial
weight (wini). Samples were dried in an oven at 105
C until
constant weight to give the final weight (wfinal). The moisture
content of the compost or soil was calculated according to Eq.
(2).
wsat  wdry
WHCð%Þ ¼  100 (1)
wsat
wini  wfinal
Moisture contentð%Þ ¼  100 (2)
wini
2.5. pH determination of compost and soil
The method used by Forster [45] was adapted to measure pH of
compost and soil. Air-dried and sieved (<2 mm) compost and soil
samples were mixed with distilled water (1:2), left at room tem-
perature for 1 h and the pH value of the supernatant was measured
with a pH meter (Metler, Toledo, UK).
2.6. Compost and soil burial of PLA coupons
PLA coupons (7  3  0.02 cm and 2  2  0.02 cm) were cut
from PLA delicatessen container lids, surface sterilized with 70% (v/
v) ethanol, allowed to air dry and buried in rectangular plastic boxes
(3 l) containing compost or soil to a depth of ca. 7 cm and incubated
at 25
, 37
, 45
, 50
or 55
C. PLA coupons (7  3  0.02 cm) were
periodically recovered for tensile strength measurements and
molecular weight determination; PLA coupons of 2  2  0.02 cm
dimensions were used for weight loss determination. Containers
were weighed weekly and water loss through evaporation adjusted
by the addition of sterile water.
2.7. Incubation of PLA coupons in sterile water, compost and soil
extract
PLA coupons (7  3  0.02 cm) were placed in sterile glass slide
staining jars containing 100 ml sterile water, and incubated at 25
,
37
, 45
, 50
and 55
C. Coupons in sterile soil and compost extracts
were incubated only at 50
C. Each PLA coupon was placed between
sterile glass slides to prevent deformation of PLA during incubation.
PLA coupons were collected from 3 jars for tensile strength mea-
surements at each temperature and in addition at 50
C for gel
permeation chromatography (GPC) analysis. When coupons were
removed, 50 ml of fluid were plated onto PDA and LB agar to check
for sterility. pH was measured in separate jars where no PLA cou-
pons were removed during the course of the experiment to
determine lactic acid release. Staining jars containing sterile water,
compost or soil extract without PLA coupons were used as controls
to monitor changes in pH.
2.8. Weight loss measurements of PLA coupons
To determine weight loss of PLA coupons, 2  2  0.02 cm
coupons were recovered, washed briefly with Trigene followed by
distilled water to remove loosely adhered compost and soil parti-
cles, sterilized by dipping in 70% (v/v) ethanol and dried at 45
C to
 M. Karamanlioglu, G.D. Robson / Polymer Degradation and Stability xxx (2013) 1e9
constant weight. Weight loss of the coupons was calculated ac-
cording to Eq. (3) [46].
mini  mdry
Weight lossð%Þ ¼  100
mini
where mini is the initial weight of the PLA coupon before burial and
mdry is the dry weight of the coupon after burial.
2.9. Determination of PLA tensile strength
Degradation of PLA coupons was assessed by measuring their
tensile strength loss (MPa). Triplicate PLA coupons were recovered
from compost, soil and sterile water at 25
and 37
C monthly, 45
C
weekly, 50
and 55
C every 3 days. Three PLA coupons were also
recovered from sterile compost and soil extract at 50
C every 3
days. PLA coupons were collected from 25
to 37
C for a year; and
from 45 to 55
C until no intact PLA coupons could be recovered.
Dumb-bell shapes (width 3.8e4.3 mm, thickness 0.2e0.3 mm,
space gauge 1.7 mm with a total length of 5 mm) were cut from the
coupons with a manual press (Wallace Test Equipment, UK) and
tensile strength determined using a T-Series Tensile Test Machine
(Tinius Olsen Ltd, UK) supported with QMAT Professional software.
The load cell was 1 kN with a crosshead speed of 10 mm/min.
Control PLA coupons were stored in dry conditions in sealed bags.
2.10. Gel permeation chromatography (GPC)
GPC was used to determine changes in the weigh average mo-
lecular weight (Mw) of PLA coupons. Molecular weight distribution
was determined on coupons buried in compost and soil; and
incubated in sterile water at 25
and 37
C after 1 year of burial.
Molecular weight of PLA coupons recovered from compost, soil,
extracts and sterile water at 50
C were assessed after 1 month. GPC
was conducted in University of Manchester in School of Chemistry.
Samples from coupons were dissolved in tetrahydrofuran, THF,
(HPLC grade, Sigma) to give a final concentration of PLA of 0.1% (w/
v) and filtered through a 0.45 mm filter to remove any small parti-
cles. 100 ml of sample was injected onto a 500 Å column (length
30 cm, column temperature 35
C), and eluted with a flow rate of
1 ml/min. Columns were calibrated using polystyrene standards
(Polymer Laboratories) and data analysed using OmniSEC 354
software.
2.11. Statistical analyses
Analysis of variance (one way ANOVA), Tukey’s HSD (honestly
significant difference) test on SPSS version 15 were used to evaluate
statistical significance of any change in tensile strength and Mw
over time with the significance threshold set at P value <0.05.
3. Results
3.1. Soil and compost analysis
The water holding capacity (WHC) and initial moisture content
of compost and soil were 65.5  2.5% and 44.4  1.2%; and 30e35%
and 20e25%, respectively. The initial pH of compost and soil were
determined as 7.2 and 7.1, respectively.
3.2. Changes in physical appearance of PLA coupons buried in soil
or compost
To determine any physical changes to the PLA coupons buried
in soil or compost, coupons were removed periodically and
Please cite this article in press as: Karamanlioglu M, Robson GD, The influence of biotic and abiotic factors on the rate of degradation of
poly(lactic) acid (PLA) coupons buried in compost and soil, Polymer Degradation and Stability (2013), http://dx.doi.org/10.1016/
j.polymdegradstab.2013.07.004
3
examined both visually (Fig. 1) and under light microscopy
(Fig. 2). When the PLA coupons were buried at 45
, 50
and 55
C,
coupons which were initially transparent became increasingly
opaque and white in appearance. However, PLA coupons buried
(3)
in soil or compost at 25
C remained transparent even after
burial for one year. Coupons buried in soil or compost at 37
C
became partly opaque at the end of one year and coupons
recovered from compost at 37
C had visible signs of fungal
growth on the surface which was confirmed by the presence of
extensive fungal hyphae revealed by light microscopy (Fig. 2B).
Films recovered from temperatures 37
C in both compost and
soil also showed signs of perforations, fungal growth and phys-
ical disintegration.
3.3. The influence of soil and compost burial and sterile water
incubation on the disintegration of PLA
Fig. 3 shows the effect of soil and compost burial and sterile
water incubation on the tensile strength of PLA coupons at
different incubation temperatures. At 25
C, no significant
(P > 0.05) reduction in tensile strength was observed under any
condition for a period of up to 12 months. Likewise, there was no
significant change (P > 0.05) in tensile strength for PLA coupons
buried for 12 months in soil at 37
C compared to unburied
control. However, tensile strength of coupons buried in compost
at 37
C began to decrease after 8 months and were significantly
(P < 0.05) lower after 11 months compared to coupons buried in
soil and control coupons. Tensile strength of coupons in sterile
water gradually declined after 10 months and was significantly
lower after 11 months at 37
C compared to the unburied control
(P < 0.05). Tensile strength loss was similar after 12 months for
coupons buried in compost and suspended in sterile water. At
45
C, PLA coupons buried in compost and suspended in sterile
water showed a significant decrease (P < 0.05) in tensile strength
compared to the unburied control after 3 weeks incubation and
continued to decline until total loss in tensile strength was
observed after 9 and 11 weeks, respectively. Loss in tensile
strength in PLA coupons buried in compost was greater at weeks
4e7 compared to coupons in soil and sterile water. A significant
loss (P < 0.05) in tensile strength of PLA buried in soil was only
first observed after 6 weeks compared to the unburied control
with total loss occurring after 10 weeks. Tensile strength of PLA
in soil was significantly (P < 0.05) lower than in sterile water
after 9 weeks of incubation at 45
C. When PLA coupons were
incubated at 50
C, tensile strength of coupons in compost were
significantly lower (P < 0.05) than the soil, water recovered
coupons and unburied controls after 15 days and continued to
decline until total loss in tensile strength was observed after 30
days. Tensile strength of coupons buried in soil did not decline
until after 30 days and tensile strength was lost after 36 days. By
contrast, tensile strength of PLA suspended in sterile water
declined more slowly. Although significant tensile strength was
observed after 21 days compared to control, the total tensile
strength was lost after 48 days. The greatest rate of decrease in
tensile strength was observed when PLA coupons were buried in
soil or compost, or suspended in sterile water at 55
C. PLA
coupons were in too small pieces for measurable tensile strength
after 9 days.
To further monitor polymer hydrolysis, the pH of the sterile
water in the presence and absence of PLA was also monitored for
a month at 25e55
C (Fig. 4.). The end stage of PLA hydrolysis
results in the release of lactic acid and a fall in pH. No reduction
in pH was observed at either 25
C or 37
C in the presence of
PLA over a 12 month period, indicating hydrolysis had not pro-
ceeded to the point of lactic acid release after one year. However,
4 M. Karamanlioglu, G.D. Robson / Polymer Degradation and Stability xxx (2013) 1e9

Fig. 1. Physical changes to PLA coupons buried in soil and compost. (A) before burial; (B) after 1 year burial at 25
C in compost; (C) after 1 year burial at 37
C in soil; (D) after 1 year
burial at 37
C in compost; (E) after 10 weeks of burial at 45
C in soil; (F) after 10 weeks of burial at 45
C in compost; (G) after 2 months of burial at 50
C in soil; (H) after 2 months
of burial at 50
C in compost; (I) after 1 month of burial at 55
C in soil; (J) after 1 month of burial at 55
C in compost. Scale bar represents 2 cm.

a large reduction in pH indicating lactic acid release was
observed at 45
C and 50
C occurring between 14 and 16
weeks; and 11e13 weeks, respectively after measurable tensile
strength had been lost (Fig. 3). A rapid decline in pH was
observed at 55
C in sterile water where lactic acid release was
observed between 15 and 18 days after complete loss of tensile
strength.
3.4. Influence of compost and soil on weight loss of PLA coupons
Fig. 5 shows weight loss of PLA coupons (2  2  0.02 cm) at 37
,
45
and 50
C as a measure of PLA degradation rates in soil and
compost independent of tensile strength. PLA coupons buried in
soil showed no significant weight loss at either 25
C or 37
C over
the course of 12 months. Similarly, no weight loss occurred at 25
C

Fig. 2. Light microscopy of PLA coupons. (A) unburied (B) buried in compost at 37
C for 1 year and showing extensive hyphal growth. Scale bar represents 100 mm.

Please cite this article in press as: Karamanlioglu M, Robson GD, The influence of biotic and abiotic factors on the rate of degradation of
poly(lactic) acid (PLA) coupons buried in compost and soil, Polymer Degradation and Stability (2013), http://dx.doi.org/10.1016/
j.polymdegradstab.2013.07.004
 M. Karamanlioglu, G.D. Robson / Polymer Degradation and Stability xxx (2013) 1e9 5

100 weight loss of 21  3% (P < 0.05) compared to the control after 3

90 (A) weeks and weight loss further increased to a maximum of 57  4%
Tensile strength (MPa)

80 after 9 weeks. Coupons recovered from soil had a significant weight

70 loss of 40  3% (P < 0.05), after 8 weeks of burial. Weight loss of
60 coupons buried in compost was significantly higher (P < 0.05) than
50 coupons buried in soil for 6 weeks. After 7 weeks, weight loss be-
40 tween coupons buried in compost and soil was not significantly
30 different (P > 0.05). When PLA was incubated at 50
C in compost,
20 weight loss of 32  5% was observed after 3 weeks and increased to
10 44  6% after 5 weeks. In contrast, no significant decrease was
0 observed in soil-recovered coupons at 50
C. At 55
C, no loss in
100 weight was observed in PLA coupons buried in either soil or
(B)
Tensile strength (MPa)

90 compost, however, by the third week, samples had physically dis-

80 integrated (data not shown). Likewise, when PLA coupons were
70 buried in soil and/or compost at 45
C and 50
C, samples for weight
60
50 loss determination could not be collected after 9 weeks and 6
40 weeks, respectively due to physical disintegration of the samples.
30
20 3.5. Influence of soluble soil and compost factors on PLA
10
0 degradation
0 1 2 3 4 5 6 7 8 9 10 11 12
100 In order to determine the relative importance of microbial and
(C)
Tensile strength (MPa)

90 Time (months) abiotic factors in PLA degradation further, tensile strength was used
80 to compare PLA degradation of coupons buried in soil or compost at
70 50
C with coupons in sterile soil or compost extracts and in sterile
60 water (Fig. 6) to establish if any soluble factors present in soil or
50 compost could account for higher degradation rates observed when
40 PLA coupons were buried in soil or compost. The loss of tensile
30
strength of PLA coupons buried in compost was significantly
20
greater than in coupons in sterile compost extract (P < 0.05).
10
Tensile strength of PLA coupons could no longer be determined
0
0 1 2 3 4 5 6 7 8 9 10 11 after 30 days when buried in compost as compared to 57 days in
Time (weeks) compost extract. Similarly, tensile strength of PLA coupons could
100 (D) not be determined after 36 days when buried in soil as compared to
Tensile strength (MPa)

90 54 days in soil extract. There was no significant difference in the

80 rate of hydrolysis in sterile compost extract or soil extract
70
(P > 0.05), however, loss of tensile strength was significantly
60
greater in sterile water compared to either soil or compost extract
50
40 (P < 0.05).
30 Fig. 7 represents the pH values monitored over 15 weeks to
20 further investigate differences in the degradation of PLA between
10 sterile water, soil and compost extracts. Reductions in pH values
0 were observed after 11 weeks in all cases and were of a similar
0 3 6 9 12 15 18 21 24 27 30 33 36 39 42 45 48 51 54 57 60 magnitude.
100 (E) Time (days)
Tensile strength (MPa)

90 3.6. Effect of soil and compost burial; and sterile water, compost
80 and soil extract incubation on the molecular weight of PLA
70
60
50 To confirm that changes in tensile strength of PLA coupons were
40 the result of polymer degradation, the weight average molecular
30 weights (Mw) of PLA coupons incubated in compost, soil and sterile
20 water at 25
and 37
; and in soil, compost, sterile soil and compost
10
extracts and water at 50
C were measured (Table 1). The molecular
0
0 3 6 9 12 15 18 21 24 27 30 weight of PLA coupons incubated at 50
C was determined after 4
Time (days) weeks and after 1 year for coupons incubated at 25
C and 37
C.
The Mw of PLA did not change after a year when PLA was buried in
Fig. 3. Tensile strength loss (MPa) of PLA coupons. PLA coupons at (A) 25
C and (B) either compost or soil at 25
C compared to control PLA coupons of
37
C for a year; (C) 45
C for 11 weeks (D) 50
C for 60 days (E) 55
C for 30 days were one year, however, significantly decreased when compared to
buried in soil (–,–) or compost (:); or incubated in sterile water (C) or kept under
dry conditions (B). Data represents the means of 3 replicates  standard error of
initial molecular weight. Significant decrease was also observed in
mean. PLA incubated in sterile water compared to initial molecular weight
and the unburied control. When incubated at 37
C in compost, soil
when PLA was buried in compost (data not shown), however, at and sterile water, Mw significantly decreased at the end of one year
37
C, insignificant weight loss in compost was observed after 4 compared to initial molecular weight and the unburied control
months and ca. 20% weight loss was observed after 12 months of (P < 0.05). The Mw of PLA recovered from compost, soil and sterile
burial. At 45
C, PLA coupons buried in compost had a significant water was not significantly different from each other at 37
C. The

Please cite this article in press as: Karamanlioglu M, Robson GD, The influence of biotic and abiotic factors on the rate of degradation of
poly(lactic) acid (PLA) coupons buried in compost and soil, Polymer Degradation and Stability (2013), http://dx.doi.org/10.1016/
j.polymdegradstab.2013.07.004
6 M. Karamanlioglu, G.D. Robson / Polymer Degradation and Stability xxx (2013) 1e9

12 Mw loss was faster at 50
C as Mw significantly decreased at 50
C

10
(A) in all samples after 4 weeks of either burial in compost and soil; or
incubation in sterile water, sterile compost or sterile soil extract
8 compared to the unburied control or the initial Mw (P < 0.05). PLA
buried in compost and soil had significantly lower Mw (51,600 and
6
pH

57,400 g/mol, respectively) than PLA incubated in sterile water

(67,900 g/mol) after 4 weeks of incubation. Compost-recovered PLA
4
had significantly lower Mw than compost extract-recovered PLA
2 (73,400 g/mol). Likewise, Mw of soil-recovered PLA was signifi-
cantly lower than PLA incubated in soil extract, 67,400 g/mol. There
0 was not a significant difference between Mw of PLA recovered from
12
(B) compost extract and soil extract.
10
8 4. Discussion
6
pH

Previously, several studies on PLA degradation suggested that

4 abiotic chemical hydrolysis at elevated temperatures in the pres-
2 ence of water was primarily responsible for PLA polymer cleavage
and degradation with microbes only acting on released products
0
[24,26,28,29,31,35e40] while other studies suggested a direct role
0 1 2 3 4 5 6 7 8 9 10 11 12 for microbes in depolymerisation [41e43]. In order to investigate
Time (months) the potential role of microbes in PLA degradation, PLA coupons
12 were incubated at temperatures between 25
and 55
C in sterile
(C) water and the rate of loss of tensile strength compared to the rate of
10 loss when buried in microorganism-rich soil and compost at the
same temperature range. Degradation was found to be faster in
8
microorganism-rich environment than abiotic media at 45
and
6 50
C. Degradation rates were similar at 25
and 37
C in biotic and
pH

abiotic systems; however, at elevated temperatures, 45
and 50
C,

4 degradation was shown to be faster in microorganism-rich
2 compost and soil than in sterile water suggesting that the pres-
ence of microbes increased degradation.
0 When incubated at 45
, 50
and 55
C in both compost, soil and
12 sterile water, loss of tensile strength was observed with the greatest
(D) rate as temperature increased indicating that temperature is a key
10 parameter for PLA degradation in humid conditions as has been
reported previously [24,26,28,31,39,40,47]. High molecular weight
8 PLA has a glass transition temperature (Tg), within the range 55e
62
C [26,31,49] and the rate of degradation of PLA is much greater
6
pH

above its Tg than below because polymer chains become more

4 flexible and water absorption increases accelerating hydrolysis
[24,26,31,40,48,49] and microbial attachment [49]. In this study at
2 Tg of PLA, 55
C, tensile strength was lost the fastest, after 9e12
days of incubation in compost, soil and in sterile water. Since
0
disintegration at 55
C was quick, there was limited growth of
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 microorganisms on the surface of PLA, therefore, role of abiotic and
Time (weeks) biotic factors were not observed. Degradation was slower at 50
C;
however, tensile strength was lost faster in microorganism rich
12
(E) compost and soil than sterile water as tensile strength was lost after
30, 36 and 48 days at 50
C in compost, soil and in sterile water,
10
respectively. Likewise, tensile strength was lost faster in compost
8 and soil than sterile water at 45
C; however at a slower rate than
50
C. Tensile strength of PLA incubated in compost and soil
6
pH

was significantly less after 7 and 9 weeks of incubation, respectively

than PLA incubated in sterile water and all tensile strength was
4 lost after 9, 10 and 11 weeks in compost, soil and sterile water,
respectively. It has been suggested previously that after abiotic
2
hydrolysis of PLA, microorganisms can subsequently assimilate the
0 released products [24]. However, in this study disintegration of PLA
0 3 6 9 12 15 18 21 24 27 30 was accelerated when biotic factors were involved. At lower
Time (days)

Fig. 4. pH changes in sterile water in the presence of PLA (-) and absence (,) of PLA. for 18 weeks (E) 55
C for 30 days. Data represents the means of 3
Coupons incubated at (A) 25
C and (B) 37
C over 12 months; (C) 45
C and (D) 50
C replicates  standard error of mean.

Please cite this article in press as: Karamanlioglu M, Robson GD, The influence of biotic and abiotic factors on the rate of degradation of
poly(lactic) acid (PLA) coupons buried in compost and soil, Polymer Degradation and Stability (2013), http://dx.doi.org/10.1016/
j.polymdegradstab.2013.07.004
 M. Karamanlioglu, G.D. Robson / Polymer Degradation and Stability xxx (2013) 1e9 7

100
70 (A) 90
(A)
80

Tensile strength (MPa)

70
60
60
50 Control

50 40
30 50ºC, compost
Weight loss (%)

20
40 10 50ºC, compost
0
extract

30 100
90 (B)
80
20

Tensile strength (MPa)

70
60

10 50 Control

40
30 50ºC, soil
0 20
2 4 6 8 10 12 10
50ºC, soil extract

Time (months) 0
100
70 (B) 90
(C)
80
60
Tensile strength (MPa)

70

50 60
Weight loss (%)

50 Control

40 40

30
50ºC, sterile
30 20 water

10
20
0
0 3 6 9 12 15 18 21 24 27 30 33 36 39 42 45 48 51 54 57 60
10 Time (days)

Fig. 6. Comparison of the rate of degradation of PLA coupons at 50
C in (A) compost

0 and sterile compost extract (B) soil and sterile soil extract (C) sterile water for up to 60
1 2 3 4 5 6 7 8 9 days. Control coupons were incubated in dry conditions. Data represents the means of
3 replicates  standard error of mean.
Time (weeks)
70
temperatures, 25
and 37
C, the effect of microorganisms on PLA
(C) degradation was not observed and 53% of tensile strength was lost
60 in sterile water and in compost at 37
C in one year, whereas in soil
only a 10% loss in tensile strength was observed due to a lower
50 moisture content in the soil. In accordance with this study, PLA
Weight loss (%)

samples incubated at 37
C in PBS lost 52% of their initial strength

40 after 44 weeks [50]. No loss of tensile strength was observed when
PLA was incubated at 25
C for a year in compost, soil or sterile
30 water. In another study [51], high molecular weight PLA buried in
soil for 1 year with temperatures ranging from 0 to 30
C also
20 showed no significant reduction in tensile strength.
In agreement with the tensile strength loss rate, the elevated
temperature, 50
C, gave the greatest rate of reduction in Mw when
10
compared with 25
and 37
C due to Tg of PLA and Mw of PLA
decreased at a faster rate in microorganism rich compost and soil
0
than sterile water at 50
C. Mw decreased 68%, 64% and 57% in
1 2 3 4 5 6 compost, soil and sterile water at 50
C after 4 weeks of incubation.
No change in Mw was observed in PLA after a year of incubation in
Time (weeks)
compost, soil and sterile water at 25
or 37
C.
Fig. 5. Weight loss of films buried in compost (-) and soil (,) at (A) 37
C for 1 year Depolymerization and hydrolysis of PLA has been reported to be
(B) 45
C for 9 weeks (C) 50
C for 6 weeks. Data represents the means of 3 accompanied by lactic acid generation and a decrease in pH
replicates  standard error of mean. [40,52,53]. Therefore pH was also measured to assess PLA

Please cite this article in press as: Karamanlioglu M, Robson GD, The influence of biotic and abiotic factors on the rate of degradation of
poly(lactic) acid (PLA) coupons buried in compost and soil, Polymer Degradation and Stability (2013), http://dx.doi.org/10.1016/
j.polymdegradstab.2013.07.004
8 M. Karamanlioglu, G.D. Robson / Polymer Degradation and Stability xxx (2013) 1e9

10 Table 1
9
(A) Molecular weight of PLA coupons kept in different degradation media at different
temperatures over time.
8
Control compost Conditions PLA coupons stored Average Mwa (g/mol)
7
extract, no PLA Control initial 159,300  5800
6
50ºC, compost Control final (1 year) 134,000  600b
5
pH

extract 25
C buried in compost (1 year) 125,200  4,200b

4 25
C buried in soil (1 year) 130,200  2,200b
25
C in sterile water (1 year) 100,700  3,100b
3
37
C buried in compost (1 year) 69,900  1,000b
2 37
C buried in soil (1 year) 68,900  1,200b
1 37
C in sterile water (1 year) 63,300  3138b
Control (4 weeks) 155,500  2700
100 (B) 50
C buried in compost (4 weeks) 51,600  1,300b
9 50
C in sterile compost extract (4 weeks) 73,400  400b
50
C buried in soil (4 weeks) 57,400  630b
8
50
C in sterile soil extract (4 weeks) 67,400  1,400b
7 50
C in sterile water (4 weeks) 67,900  1,500b
Control soil
6 extract, no PLA Data represents the means of 3 replicates  standard error of mean.
a
5 Weight average molecular weight.
pH

50ºC, soil b
Significant decrease in Mw compared to initial Mw.
4 extract

3
Compost buried PLA showed more physical change, disintegra-
2
tion and greater weight loss at temperatures of 37
, 45
and 50
C.
1 Moisture content of the soil was lower than compost at the same
0 temperature due to different WHC values of compost and soil,
10 (C) 65.5  2.5% and 44.4  1.2%, respectively. When tensile strength
9
loss of PLA was compared in compost and soil at 37
, 45
and 50
C,
8
compost-buried coupons disintegrated faster than soil-buried ones
7 Control sterile
water, no PLA
suggesting necessity of water for degradation as water decreases Tg
6 of PLA over time; increasing hydrolysis [24,31,40,48,49] and mi-
5 50ºC, sterile crobial attachment [49]. It also suggests physical and nutritional
pH

water structure of compost encouraged more mesophilic and/or ther-

4
3
2
1 compost and soil was compared with sterile compost and soil ex-
0 tracts where only hydrolysis took place with all the soluble com-
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
Time (weeks)
Fig. 7. pH changes in (A) sterile compost extract (B) sterile soil extract (C) sterile water
in the presence of PLA ( ) and absence ( ) of PLA coupons incubated at 50
C over 30
days. Data represents the means of 3 replicates  standard error of mean.
degradation. Tensile strength was lost after 11 and 8 weeks in sterile
water at 45
and 50
C, respectively and after 9tdays at 55
C. Lactic
acid release was observed at 45
, 50
and 55
C approximately on the
15th week, 11th week and 15th day, respectively in sterile water.
Lactic acid release in the sterile extracts was in agreement with the
loss of tensile strength when assessing degradation. At 50
C, tensile
strength was lost approximately in 8 weeks in compost and soil
extract and lactic acid release was observed after 11 weeks in
compost and soil extract. Complete tensile strength loss was
observed before the lactic acid release as cleavage of PLA chains
caused the release of lactic acid monomers and a decrease in pH.
When weight loss was assessed to compare the PLA degradation
activity of microorganisms in compost and soil, more weight loss in
compost-buried PLA was observed at temperatures 37
C. Studies
also suggest a low weight loss of PLA over time and that weight loss
is less at lower temperatures [51,54,55]. In the present study,
weight loss was not observed as much as tensile strength and
molecular weight loss. It is suggested that in semi-crystalline PLA
when amorphous regions are reduced, molecular weight signifi-
cantly decreases and crystalline regions start to be reduced causing
mechanical strength loss and weight loss [50,56]. Therefore, a sig-
nificant molecular weight loss is necessary for a significant tensile
strength loss and weight loss.
mophilic PLA degraders.
In order to further assess the importance of microorganisms and
abiotic factors in PLA degradation mechanism, PLA degradation in
ponents of compost and soil except for the microorganisms at 50
C.
Rate of degradation in compost extract was significantly slower than
in compost and it was also found to be slower in soil extract than soil
(P < 0.05). All tensile strength was lost in 30 days in compost but in
compost extract it was lost in 57 days. PLA was degraded in 36 days
in soil and in 54 days in soil extract. In compost, significantly more
Mw, 68%, was lost than in compost extract, Mw, 53%. Likewise, in
soil, 64% Mw was lost which was significantly more than Mw loss in
soil extract, 57%. In a previous study [40] it was found that there
was no evidence for the enhanced degradation of high molecular
weight PLA in the presence of microorganisms as no significant
degradation rate difference was observed between abiotic and biotic
systems when PLA was incubated at 40
, 50
and 60
C [40].
Therefore, it is widely accepted that PLA degradation involves a two-
step mechanism with a chemical hydrolysis in the presence of water
and microorganisms utilising the products of PLA degradation
[24,26,28,29,31,35e39]. However, in this current study, loss of ten-
sile strength and decline in the Mw of PLA in microorganism-rich
compost and soil were greater than chemical hydrolysis in sterile
water, compost and soil extract at elevated temperatures indicating
microorganisms can directly enhance PLA degradation. In addition
to potential enzymatic contributions to enhanced PLA degradation,
physical effects of growth by microbes on surfaces are well docu-
mented. For example, the growth of fungal hyphae and bacteria can
cause disruption of surfaces such as stone and marble by penetrating
and growing in pits and fissures on the surface [57,58].
In conclusion, this study suggests that microbes can directly
enhance degradation of high Mw PLA over and above chemical
hydrolysis at elevated temperatures and that microorganisms in

Please cite this article in press as: Karamanlioglu M, Robson GD, The influence of biotic and abiotic factors on the rate of degradation of
poly(lactic) acid (PLA) coupons buried in compost and soil, Polymer Degradation and Stability (2013), http://dx.doi.org/10.1016/
j.polymdegradstab.2013.07.004
 M. Karamanlioglu, G.D. Robson / Polymer Degradation and Stability xxx (2013) 1e9
compost are more active in this process compared to soil. However,
little or no degradation appears to occur at lower temperatures
(25
C and 37
C) over 12 months and PLA accumulating in the
environment may therefore cause environmental pollution prob-
lems. Further studies are required to identify the microbes involved
in enhancing PLA degradation and the impact of PLA degradation
on composting systems.
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