ArticlePDF Available

Ecotoxicity Evaluation of the Biodegradable Polymers PLA, PBAT and its Blends Using Allium cepa as Test Organism

Authors:

Abstract and Figures

Biodegradable polymers are considered a feasible option to minimize the environment impacts of high disposal of solid waste. Nevertheless, environmental safety of these materials is a few explored issue. In this context, this study evaluated ecotoxicological effects in soil of the biodegradable materials poly(lactic acid)-PLA, poly(butylene adipate co-terephthalate)-PBAT and their blends compatibilized with a chain extender. The tool used for this analysis was the bioassay with Allium cepa as test organism. The studied materials were not phytotoxic, cytotoxic, genotoxic nor mutagenic for meristematic cells of A. cepa.
This content is subject to copyright. Terms and conditions apply.
Vol.:(0123456789)
1 3
J Polym Environ
DOI 10.1007/s10924-017-0990-9
ORIGINAL PAPER
Ecotoxicity Evaluation oftheBiodegradable Polymers PLA,
PBAT andits Blends Using Allium cepa asTest Organism
PaulaA.Palsikowski1· MatheusM.Roberto2· LaisR.D.Sommaggio2·
PatríciaM.S.Souza1· AnaR.Morales1,3· MariaA.Marin‑Morales2
© Springer Science+Business Media New York 2017
polymeric packages), the eventual risks associated to poly-
mers and its by-products should be assessed.
Most of the biodegradable polymers belong to the poly-
ester group, since ester groups are susceptible to hydroly-
sis, favouring the formation of low molecular weight sub-
stances that can be assimilated by microorganisms and
converted into water, carbon dioxide, biomass and degra-
dation products [1]. Poly(butylene adipate-co-terephtalate)
(PBAT) and Poly(lactic acid) (PLA) are examples of biode-
gradable polyesters with potential agriculture applications
as soil covering (mulching) [2, 3].
On 1990s, BASF launched the Ecoflex®, a PBAT. This is
a fossil-based polyester proposed for different applications
as agricultural films and packaging [4]. It has also been
used in blends with another polyester, PLA (poly lactic
acid), aiming to overcome various drawbacks of PLA such
as its brittleness and processability limitations [5].
In order to evaluate ecotoxicity related to biodegradable
polymers it is important to define timing for assessment. A
plastic material can be safe before biodegradation, but may
be toxic during degradation. Besides, suitable and sensitive
test methods should be considered [6].
Witt et al. identified by gas chromatograph/mass spec-
troscopy (GC-MS) the by-products of PBAT degradation
by the actynomycete Thermomonospora fusca. The solu-
tion with intermediates of Ecoflex degradation (1,4-butane-
diol, adipic acid and terephtalic acid) were tested by Daph-
nia magna and Photobacterium phosphoreum. In this study
no significant toxicological effects were observed [7].
Besides, Ecoflex has been tested for following toxi-
cological assays: terrestrial plant toxicity (OECD 208),
earthworm toxicity (OECD 207), primary skin irritation
rabbit (OECD 404), primary irritations of the mucus mem-
brane rabbit (OECD 205), guinea pig (OECD 406), LD50
rat (OECD 423) and Ames test (OECD 471). The tests
Abstract Biodegradable polymers are considered a feasi-
ble option to minimize the environment impacts of high dis-
posal of solid waste. Nevertheless, environmental safety of
these materials is a few explored issue. In this context, this
study evaluated ecotoxicological effects in soil of the bio-
degradable materials poly(lactic acid)-PLA, poly(butylene
adipate co-terephthalate)-PBAT and their blends compati-
bilized with a chain extender. The tool used for this analy-
sis was the bioassay with Allium cepa as test organism. The
studied materials were not phytotoxic, cytotoxic, genotoxic
nor mutagenic for meristematic cells of A. cepa.
Keywords Biodegradable polymers· Ecotoxicity·
Bioassay· Allium cepa
Introduction
Biodegradable polymers have been widely studied for new
applications. The contribution related to waste reduction is
notorious, but since these new materials are proposed to be
disposed directly on the environment (mulch films used for
soil covering in agriculture) or indirectly (composting of
* Ana R. Morales
morales@feq.unicamp.br
1 Department ofMaterials Engineering andBioprocess, School
ofChemical Engineering, State University ofCampinas
(UNICAMP), Campinas, SP, Brazil
2 Department ofBiology, Institute ofBiosciences, UNESP—
Univ Estadual Paulista, RioClaro, Brazil
3 Albert Einstein Avenue, 500-Cidade Universitária,
13083-852Campinas, SP, Brazil
J Polym Environ
1 3
evaluated ecotoxicological effects of degradation interme-
diates as well as the product safety during the use phase
and no adverse effects were verified [4].
The majority of published studies related to ecotoxic-
ity of biodegradable polymers are focused on germination
assays on plants such as: radish, rice and beans [810].
According to ASTM D6400 [11], the standard related to
certification of compostable plastics, the ecotoxicologi-
cal test recommended to access terrestrial safety is evalua-
tion of germination rate and plant biomass, using the guide
OECD 208 for two different plant species.
In this context it is important to consider that some plant
species are capable of offering a more detailed ecotoxico-
logical information as cytotoxic, genotoxic and mutagenic
effects associated to polymers degradation process [12].
The Allium cepa test has been applied for environmental
monitoring to detect different classes of pollutants and also
complex mixtures, such as water and soil samples from
contaminated areas [13].
Souza etal. [12] reported that A. cepa could be an effi-
cient tool to study ecotoxicity related to degradation of
pure PLA and nanocomposites with in composting process,
including quantifications of mitotic index, chromosomal
aberrations and micronucleus. In this study the compost
samples after degradation of the polymer did not presented
mutagenic effects and the types of the observed chromo-
somal aberrations indicated a possible genotoxic effect of
the materials, which may be related to an aneugenic action
of PLA degradation products.
The mitotic index (MI), characterized by the ratio of
dividing cells and total cells observed, could be used as a
parameter for evaluating the cytotoxicity of various agents
[13].
The term genotoxic is used to describe an agent capa-
ble of promoting DNA damage. Chromosomal aberrations
as C-metaphases, polyploid metaphases, metaphases with
adherences, metaphases with chromosomal losses, ana-
phases and telophases with delays, losses, chromosomal
bridges and nucleus abnormalities are examples of altera-
tions which can be considered genotoxic evidences [13,
14].
The micronuclei have been considered by many authors
as the most effective and simplest endpoint to analyze
mutagenic effect induced by chemicals. A mutation is
defined as a change in the DNA sequence that leads to her-
itable genetic changes. The micronucleus appears in daugh-
ter cells as a result of damage induced in parental cells [15].
This paper will present recent results from A. cepa bioas-
say applied to evaluate cytotoxic, genotoxic and mutagenic
effects in soil after degradation of different biodegradable
polymeric materials. The tested polymers were poly(lactic
acid)—PLA, poly(butylene adipate-co-terephthalate)-
PBAT and their blends. These materials were previously
studied in terms of thermal and mechanical behavior [16]
and biodegradation in soil [17].
Experimental Procedure
Materials
The preparation of polymers and blends samples were
made using the following materials: poly(butylene adipate-
co-terephtalate)-PBAT, grade Ecoflex F BX 7011 (BASF);
poly(lactic acid)-PLA, grade 4042D (Natureworks) and the
chain extender- CE, grade Joncryl ADR-4368 (BASF).
The blend nominated 25/75 was prepared using 25% by
weight (wt.) of PLA and 75% wt of PBAT. The blend 75/25
was prepared with 75% wt. of PLA and 25% wt of PBAT.
For all the samples it was added at 1 pcr (parts per 100
parts of resin). Notations and compositions of the studied
materials are detailed in Table1.
The samples were prepared previously by Kuchnier
[16] in a torque rheometer HAAKE Rheomix 600P during
5min, at 120rpm and 180 °C. After that, samples with a
thickness of 0.3mm were obtained in a press Labtech Engi-
neering Co. Ltd., model LP20B at a constant pressure of
800psi, and temperatures of 120 °C (PBAT), 160 °C (PLA)
and 150 °C (blends).
Characteristic andSample Preparation
The samples were submitted to disintegration in soil.
According to Innocenti [18], in order to have a good chance
to detect possible negative effects, it is advisable to apply
high initial concentrations of the polymer under study. A
‘high’ concentration is considered to be one that is at least
1–2 order of magnitude greater than the normal dose used
in real applications. So, if at high concentrations there is no
effect observed, the environmental risk at normal doses is
negligible.
In this work the concentration of each material in soil
was calculated according to Innocenti [18], based on the
sample thickness and considering the value of 1g/cm3 for
soil apparent density in São Paulo State [19]. Table2 shows
the thickness, density and mass of each sample used in a
mixture with 200g of soil (suitable for the bioassay test).
Table 1 Compositions of samples (PLA, PBAT and blends)
Sample name PLA (wt%) PBAT (wt%) CE (phr)
PLA 100 0 1
PBAT 0 100 1
75/25 75 25 1
25/75 25 75 1
J Polym Environ
1 3
The plastic samples with a thickness of 0.3mm were cut
into square pieces of 1cm2. The mass reported in Table 2
of each material was mixed with 200g of the environment
matrix, consisted of different soils collected in December of
2013 in the campus of State University of Campinas (São
Paulo State- Brazil), in a house yard also located in Campi-
nas and a garden located in the city of São Jorge d’Oeste
(Paraná State-Brazil). The mixture was done following an
recommendation of ASTM D5988 [20] for biodegradation
tests of plastics, that indicates to make a laboratory mixture
of equal parts (by weight) of soil samples obtained from at
least three diverse locations in order to maximize biodiver-
sity of microorganisms.
The characterization of this mixture of soils is presented
in Table3. The determination of soil moisture was made
at 105 °C for 24 h. The water holding capacity was deter-
mined according to the methodology described by Casado
[21]. The parameters of organic matter, nitrogen and pH
were quantified according to the methods described by
MAPA [22]. The result for organic carbon was obtained by
dividing the result of organic matter for 1.8, as proposed by
Kiehl [23].
Each mixture of soil and samples was added in a 1L
vessel. In a separated vessel it was added just 200g of pure
soil. The vials were kept at room temperature for 6months.
After this period it was observed a low level of disintegra-
tion. In order to promote degradation process the vessels
were placed in an oven at controlled temperature (60 °C)
for 2months until the disintegration of samples. The higher
temperature was chosen to be close of the PLA’s Tg in
order to promote hydrolytic degradation since it increases
free volume in polymeric materials, enhancing the chain
mobility and water diffusion [24]. During degradation
period, aeration was promoted by turning the soil periodi-
cally (every 2days).
After this period the aqueous extract of soil samples
were obtained based on the standard ABNT 10006-04 [25]
by mixing 200g of each sample (dry weight) and 800mL
of water. The moisture after disintegration process in soil
was calculated at 105 °C by 24 h. The moisture, the wet
mass and the quantity of added water in each sample for
preparing the solubilized are shown in Table4.
After stirring the mixtures of water/soil for 5 min, the
vessels were covered with PVC film and kept on standing
for a period of 7days at 22 °C. The supernatant was filtered
using a membrane with 0.45µm of porosity.
Bioassay withAllium cepa Organism
Seeds of A. cepa from variety Baia Periforme were
arranged in Petri dishes lined with filter paper (100 seeds
per plate, two plates per treatment). The seeds were sub-
jected to germination with samples of aqueous extract in
incubator at 22 ± 2 °C. A negative control (NC) was per-
formed with ultrapure water. Also, a positive control was
performed by using methylmethane sulfonate (MMS) at a
concentration of 4 × 10−4M, which has a renowned clasto-
genic action [26], and the trifluralin herbicide (TRF) at the
concentration of 0.84ppm, which has an aneugenic action
already described [14]. The bioassay was performed based
in two independent exposures in two different batches, each
one with two replicates per treatment.
After reaching about 1.5 cm in length, the roots were
collected and fixed with Carnoy solution (ethanol/acetic
acid—3:1—v/v) for 6 h at room temperature. After this
time, the fixative was replaced by a recently prepared solu-
tion. The roots were stored at 4 °C until the slides prepa-
ration. The test with meristematic cells of A. cepa was
performed based on the protocol established by Grant
[27] with some modifications. The fixed roots were sub-
jected to the Feulgen reactive (Schiff’s solution), which
reacts specifically with DNA. For the preparation of the
slides, the meristems were placed on slides containing a
drop of acetic carmine (2%), covered with coverslips, and
Table 2 Thickness, density and mass of polymeric samples used in
ecotoxicity test
Material PLA 75/25 25/75 PBAT
Thickness (cm) 0.30 0.32 0.28 0.30
Density (g/cm3) 1.24 1.25 1.26 1.27
Mass (g) 20.9 21.70 19.3 20.6
Table 3 Soil characterization
Parameters Results
Moisture (%) 45.1
Water holding capacity (%) 75.2
Organic matter (g/dm3) 45.0
Organic carbon (g/dm3) 40.0
Nitrogen (g/Kg) 2.7
C/N 14.8
Table 4 Data quantified for the preparation of solubilized
Soil sample Moisture (%) Wet mass (g) Volume of dis-
tilled added water
(mL)
PLA 47 378.1 621.8
PBAT 46 374.0 626.0
75/25 48 381.7 618.3
25/75 50 396.3 603.7
Solo 44 356.3 643.7
J Polym Environ
1 3
gently squashed. After the removal of the coverslips in liq-
uid nitrogen, the permanent slides were prepared with syn-
thetic resin.
The analysis of slides was performed in a light micro-
scope (Nikon, model Eclipse E200), using a 100× objec-
tive. Around 5000 cells were counted per treatment, being
500 cells counted per slide and five slides evaluated for
each replicate.
The phytotoxicity was analyzed by Germination Index
(GI), calculated according to Eq.1:
The cytotoxicity was assessed by Mitotic Index (MI) cal-
culation, acquired by the ratio between the total number of
cells on division (counting cells in different mitosis phases:
prophase, metaphase, anaphase and telophase) and the total
number of observed cells, according to Eq.2.
The genotoxicity was accessed by the Chromosomal
Alterations Index (CAI), calculated according to Eq. 3,
considering the number of cells carrying chromosomal
aberrations (adherence, polyploidy, loss, C-methaphase,
multipolarity, bridge and loss) and nuclear abnormalities
(binucleated cell, trinucleated cell and lobulated nucleus).
The mutagenicity endpoint was based on the presence
of micronucleated cells and the Mutagenicity Index (MutI)
was calculated according to Eq.4.
The statistical analysis was performed using the software
BioEstat 5.3 (Mamirauá Institute, Brazil). The parameters
GI and MI were evaluated by ANOVA-One Way (p < 0.05),
since the results presented a normal distribution. The CAI
and MutI did not presented normal distribution and, in this
case, the results were evaluated by Kruskal–Wallis method
(p < 0.05).
Results andDiscussion
The results showed that, overall, there was no significant
germination inhibition by the samples, when they were
compared with the negative control. Exception occurred
for the treatments with PLA and MMS, but it happened
just for one of the batches of experiment for each treatment
(Fig.1).
(1)
GI
=
total number of germinated seeds
total number of exposed seeds
×
100
(2)
MI
=
total number of cells on division
total number of observed cells
×
100
(3)
CAI
=
total number of altered cells
total number of observed cells
×
100
(4)
Mult
=
total number of altered cells
total number of observed cells
×
100
The A. cepa species is one of the recommended organ-
isms by OECD-208 [28] for ecotoxicity evaluations. Nev-
ertheless in the field of biodegradable polymers it was not
found ecotoxicological studies in the literature reporting
this test organism.
Rudeekit etal. [10] evaluated blends of PLA and starch
after composting based on OECD-208 [28], where the
parameters quantified were: rate of germination and growth
of plants Oryza sativa (monocotyledonae) and Vigna
radiata (dicotyledonae).
Mitelut and Popa [9] studied the degree of toxicity of
PLA/PBAT/lignin blends after composting using the ger-
mination of Raphanus sativus as a parameter, following the
methodology proposed by Gariglio etal. [29].
This shortage of information highlights the importance
of ecotoxicology studies for biodegradable polymers,
which could consider different test organisms in addition
to the different environments where these materials can be
disposed off after their useful life (soil, composting, land-
fills). Therefore, the A. cepa organism is able to support
analysis in this relevant field.
None of the treatments presented Mitotic Index (MI) sig-
nificantly different from the negative control. So, the mate-
rials PLA, PBAT and its blends (25/75 and 75/25) showed
to be not cytotoxic for A. cepa organism (Fig.2).
The Chromosomal Alterations Index (CAI) showed
significant values for the positive controls with trifluralin
(TRF) and methylmethane sulfonate (MMS) (Fig.3).
Chromosomal aberrations are based on changes in the
structure or number of chromosomes of cells, which can
occur spontaneously or as a result of action of chemical or
physical agents. Changes in chromosome structure may be
induced by inhibition of DNA synthesis, by replication of
the modified DNA, or by breaks in the DNA strand, char-
acterizing a so-called clastogenic effect. The occurrence
of micronucleus is result of genotoxic events that were not
repaired or repaired wrongly in the parental cell, resulting
Fig. 1 Germination index for PLA, PBAT and its blends (*Statisti-
cally significant, p < 0.05—ANOVA-One Way)
J Polym Environ
1 3
in chromosomal breaks that form the micronuclei. The
changes in the number of chromosomes, as observed in
aneuploid and polyploid cells, may be a result from abnor-
mal segregation of chromosomes during the cell division,
that occur spontaneously or by the influence of aneugenic
agents. The MNs can still be formed by action of the toxic
agent on proteins or cytoplasmic structures, such as mitotic
spindles, which result in losses of whole chromosomes and
consequently in the formation of MNs in daughter cells
[13].
Leme and Marin-Morales [13] state that the A. cepa test
is an excellent indicator of genotoxicity and mutagenicity
promoted by toxic agents. The presence of chromosomal
aberrations and micronuclei in A. cepa cells is a signal that
the agent has genotoxic or mutagenic potential respectively,
which supports its applicability to detect both classes of
damages, being this assay recommended to assess the
effects of environmental chemicals or pollutants.
According to Fernandes etal. [30] the 0.84ppm concen-
tration of herbicide can be regarded as “diagnostic concen-
tration” for genotoxicity, since it causes high rates of chro-
mosomal aberrations and nuclear abnormalities, indicating
an aneugenic effect. The other positive control, MMS, also
presented significant values of CAI related to negative con-
trol. It is also expected considering its ability to induce
chromosomal aberrations [26].
Among the tested materials, one of the experiments pre-
sented a significant CAI in comparison to negative control.
It happened for the blend 25/75, but this behavior was not
observed for the second experimental batch. Palsikowski
etal. [17] monitored the number-average molecular weight
(Mn) of these materials during degradation in soil at dif-
ferent periods (0, 60, 120, 180, 240, 300 and 360days) at
room temperature. It was observed that for all the periods
the pure PBAT and the blend 25/75 presented the lowest
values of Mn. This parameter is sensible to low molecular
weight species in the sample. Therefore, it is possible to
infer that the solubilized samples from soils after degrada-
tion of PBAT and 25/75 may contain degradation products
at higher concentrations than other materials evaluated.
Palsikowski et al. [17] also studied the mineraliza-
tion rate of all the materials in soil during 120 days. The
mineralization after this period for PBAT and 25/75 was
similar—16 and 11%, respectively. So, the main difference
between the both solubilized could be that 25/75 would
have more degradation products derived from PLA. There-
fore, the significant result of CAI for one of experiments
with the blend 25/75 may be associated to PLA degrada-
tion products. Souza etal. [12] also observed values of CAI
statistically significant associated to degradation of PLA in
compost environment using A. cepa as test organism. Since
in the present study this result was not observed in the sec-
ond experiment, new assays are necessary to consolidate
and better clarify this behavior. The frequencies of altera-
tions observed among the different treatments are presented
on Table 5. Figure 4 shows A. cepa cells with different
chromosomal aberrations.
At least one of the experiments with the solubilized
samples tested presented aberrations as chromosomal
breakages and nuclear bud. The breaks are promoted by
clastogenic effect and can be observed at higher frequen-
cies in the case of MMS, a compound that is known for its
clastogenic action [23].
The treatments with PLA or PBAT induced aberration
as C-metaphase. This aberration is possibly reversible [31]
but if no repairs occur, polyploid cells may be formed.
Among the nuclear alterations, the most frequent was
nuclear bud, that can be a consequence of breaks, bridges
and delays, since in these cases the chromosomes or frag-
ments can not be incorporated to the main nucleus of the
cell [32].
Fig. 2 Results for Mitotic Index (*Statistically significant, p < 0.05—
ANOVA-One Way)
Fig. 3 Results for Chromosomal Alterations Index (*Statistically sig-
nificant, p < 0.05—Kruskal–Wallis)
J Polym Environ
1 3
Lobulated nuclei were also observed. Nevertheless, sig-
nificant results were observed only for trifluralin. This aber-
ration may be formed by chromosomal bridges in multipo-
lar anaphases, which can be involved by a nuclear envelope
[33]. The presence of lobulated nucleus can indicate cell
death, since these alterations are not observed on F1 cells
of A. cepa organism [34].
The unique experiment that presented chromosomal
adherence was the one related to PBAT. This kind of altera-
tion indicates toxic effect of the material, which can pro-
mote irreversible damages to cells, including cell death [34,
35].
The Mutagenicity Index (MutI) results are showed in
Fig.5.
Statistically significant values were obtained for MutI in
the case of MMS, due to the higher frequency of chromo-
somal breakages and micronucleus that can be attributed to
clastogenic effect of the substance. This behavior indicates
an adequate response of the system, since this substance is
used as a positive control for mutagenicity [36].
The treatments conducted with trifluralin also showed
statistically significant values for MutI. It may be associ-
ated with its aneugenic action because the inactivation of
mitotic fuse prevents the migration of chromosomes to the
poles of the cell, which once dispersed in the cytoplasm,
leads to losses that can induce micronucleus formation
[14].
The treatments with PLA, PBAT and its blends did not
show statistically significant values of MutI, so none of
these materials presented mutagenic effects on A. cepa
species.
Conclusion
The bioassay with the test organism A. cepa was a sensi-
tive tool for assessment of ecotoxicity of soil samples after
PBAT, PLA and its blends disintegration.
In general the studied materials did not present any cyto-
toxic, genotoxic and mutagenic effects on meristematic
cells of A. cepa. An exception occurred for the blend 25/75,
in which one of the experiments presented chromosomal
aberration index statistically significant related to negative
control. Although it was not observed in the experiment
repetition (Exp. 2), it is not negligible. New experiments
are recommended for all materials, in order to consolidate
the information about ecotoxicological effects from deg-
radation products of polymeric materials. In a following
study it is recommended to include analysis on cells from
F1 region, which are cells derived from mitotic division
Table 5 Chromosomal alterations evaluated in Allium cepa cells after germination in different samples and blends of PLA and PBAT (aver-
age ± standard deviation—%)
Alterations Experiment CN MMS TRIF SOIL PLA 75/25 25/75 PBAT
Chromosomal break 1 0.00 ± 0.00 0.26 ± 0.20 0.06 ± 0.09 0.02 ± 0.05 0.09 ± 0.16 0.06 ± 0.09 0.07 ± 0.16 0.04 ± 0.08
20.09 ± 0.12 0.19 ± 0.20 0.05 ± 0.12 0.00 ± 0.00 0.04 ± 0.12 0.02 ± 0.06 0.00 ± 0.00 0.04 ± 0.08
Nuclear bud 1 0.00 ± 0.00 0.36 ± 0.47 0.95 ± 0.49 0.06 ± 0.09 0.36 ± 0.34 0.26 ± 0.22 0.39 ± 0.28 0.19 ± 0.13
20.07 ± 0.23 0.81 ± 0.60 0.71 ± 0.45 0.00 ± 0.00 0.02 ± 0.06 0.04 ± 0.08 0.00 ± 0.00 0.06 ± 0.18
Chromosomal adherence 1 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.04 ± 0.08
20.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
Chromosomal bridge 1 0.00 ± 0.00 0.02 ± 0.06 0.11 ± 0.15 0.02 ± 0.06 0.04 ± 0.13 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
20.00 ± 0.00 0.04 ± 0.12 0.09 ± 0.13 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.02 ± 0.06
Chromosomal loss 1 0.00 ± 0.00 0.27 ± 0.34 0.02 ± 0.06 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
20.02 ± 0.06 0.07 ± 0.18 0.17 ± 0.14 0.02 ± 0.06 0.00 ± 0.00 0.02 ± 0.06 0.00 ± 0.00 0.00 ± 0.00
C-metaphase 1 0.00 ± 0.00 0.02 ± 0.05 0.00 ± 0.00 0.00 ± 0.00 0.02 ± 0.06 0.00 ± 0.00 0.00 ± 0.00 0.02 ± 0.06
20.00 ± 0.00 0.02 ± 0.06 0.33 ± 0.40 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
Multipolar anaphase 1 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.02 ± 0.06 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
20.00 ± 0.00 0.00 ± 0.00 0.17 ± 0.19 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
Binucleated cell 1 0.00 ± 0.00 0.00 ± 0.00 0.73 ± 0.77 0.06 ± 0.12 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
20.00 ± 0.00 0.00 ± 0.00 0.07 ± 0.18 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
Trinucleated cell 1 0.00 ± 0.00 0.00 ± 0.00 0.08 ± 0.13 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.02 ± 0.06
20.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
Polynucleated cell 1 0.00 ± 0.00 0.00 ± 0.00 0.04 ± 0.12 0.02 ± 0.06 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
20.00 ± 0.00 0.00 ± 0.00 0.04 ± 0.12 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
Lobulated cell 10.00 ± 0.00 0.11 ± 0.20 2.27 ± 2.08 0.02 ± 0.06 0.02 ± 0.06 0.02 ± 0.06 0.00 ± 0.00 0.09 ± 0.17
20.00 ± 0.00 0.02 ± 0.06 1.56 ± 1.32 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00 0.00 ± 0.00
J Polym Environ
1 3
of meristematic cells and could prove the permanence of
damages. Besides, for a better understanding of ecological
impact related to biodegradable materials it is important
to consider new researches with different test organisms
(plants, algae, annelids, bacteria, human cells). Also, par-
allel chemical analysis would permit to detail degradation
process and to indicate the byproducts in solubilized sam-
ples from soil.
Acknowledgements The authors would like to thank you the Foun-
dation for Research of the State of São Paulo - FAPESP (Process
2014/09883-5) and National Council of Scientific andTechnological
Development CNPQ for the financial support.
References
1. Rizzareli P, Carroccio S (2014) Anal Chim Acta 808:18
2. Jandas PJ, Mohant S, Nayak SK (2013) Ind Eng Chem Res
52:17714
3. Kijchavengkul T, Auras R, Rubino M, Selke S, Ngouajio M, Fer-
nandez RT (2011) Polym Degrad Stab 96:1919
4. Siegenthaler KO, Künkel A, Skupin G, Yamamoto M (2012)
Ecoflex and Ecovio: biodegradable, performance-enabling plas-
tics. In: Abe A, Albertsson A-C, Dusek K, Genzer J, Jeu WH,
Kobayashi S, Lee K-S, Leibler L, Long TE, Manners I, Möller
M, Terentjev EM, Vicent M, Voit B, Wegner G, Wiesner U (eds)
Advances in polymer science 245. Springer, Berlin, pp91–136
5. Nofar M, Heuzey MC, Carreau PJ, Kamal MR, Randall J (2016)
J Rheol 60:637
6. Innocenti FD (2014) Front Microbiol 5:475
Fig. 4 Chromosomal aberrations observed in Allium cepa meris-
tematic cells: a normal interphase; b normal prophase; c normal
metaphase; d normal anaphase; e normal telophase; f interphase
with nuclear bud; g micronucleated cell in interphase; h micronu-
cleated and polyploid cell in prophase; i C-metaphase; j metaphase
with adherence; k polyploid metaphase; l micronucleated cell in
metaphase; m anaphase cell with chromosomal bridge; n multipolar
anaphase; o telophase with chromosomal breakage; p telophase with
chromosomal loss; q binucleated and lobulated cell
Fig. 5 Results for Mutagenicity Index (*Statistically significant,
p < 0.05—Kruskal–Wallis)
J Polym Environ
1 3
7. Witt U, Einig T, Yamamote M, Kleeberg I, Decwer WD, Müller
RJ (2001) Chemosphere 44:289
8. César MEF, Mariani PDSC, Innocentini-Mei LH, Cardoso EJBN
(2009) Polym Testing 28:680
9. Mitelut C, Popa ME (2011) Rom Biotechnol Lett 16:121
10. Rudeekit Y, Siriyota P, Intaraksa P, Chaiwutthinan P (2012) Adv
Mater Res 506:323
11. ASTM D6400-12. Standard specification for labeling of plastics
designed to be aerobically composted in municipal or industrial
facilities
12. Souza PMS, Corroqué NA, Morales AR, Marin-Morales MA,
Mei LH (2013) J Polym Environ 21:1052
13. Leme M, Marin-Morales MA (2009) Mutat Res 682:71
14. Fernandes TCC, Mazzeo DEC, Marin-Morales MA (2007) Pes-
tic Biochem Physiol 88:252
15. Serres FJ (1978) Environ Health Perspect 27:3
16. Kuchnier CN (2014) Estudo do efeito de aditivo extensor de
cadeia multifuncional em blendas de PLA/PBAT. Dissertação
de Mestrado em Engenharia Química, Faculdade de Engenharia
Química—UNICAMP, Campinas, SP
17. Palsikowski PA, Kuchnier CN, Pinheiro IF, Morales AR (2017)
Biodegradation in soil of PLA/PBAT blends compatibilized with
chain extender. J Polym Environ 1–12
18. Innocenti FD (2005) In: Bastioli C (ed) Handbook of biodegrad-
able polymeric materials and their applications. American Scien-
tifica Publishers, Valencia
19. Setzer J (1941) Bragantia 1:255
20. ASTM D5988-12. Standard test method for determing aerobic
biodegradation of plastic materials in soil
21. Casado EB (2009) Desenvolvimento e caracterização de blen-
das de poliéster sintético biodegradável com proteína de soja e
estudo de biodegradação em solo. Dissertação de Mestrado em
Engenharia Química—Departamento de Tecnologia de Materi-
ais, Faculdade de Engenharia Química—UNICAMP, Campinas,
SP
22. MAPA—Ministério da Agricultura, Pecuária e Abastecimento
(2007) Instrução Normativa No. 28: Manual de Métodos Analíti-
cos Oficiais para Fertilizantes Minerais, Orgânicos, Organo-min-
erais e Corretivos
23. Kiehl JE (1985) Fertilizantes orgânicos. Agronômica Ceres,
Piracicaba
24. Xu H, Yang X, Xie L, Hakkarainen M (2016) Biomacromole-
cules 17:985
25. ABNT—Associação Brasileira de Normas Técnicas (2004) NBR
10.006: procedimento para obtenção de extrato solubilizado de
resíduos sólidos. ABNT, Rio de Janeiro
26. Rank J, Nielsen MH (1997) Mutat Res 390:121
27. Grant WF (1982) Mutat Res 99:273
28. OECD 208 (2006) Terrestrial plant test: seedling emergence and
seedling growth test
29. Gariglio NF, Buyati MA, Pillati RA, Russia DEG, Acosta MR
(2002) NZJ Crop Hortic Sci 30:135
30. Fernandes TCC, Mazzeo DEC, Marin-Morales MA (2009) Eco-
toxicol Environ Saf 72:1680
31. Odeigah PGC, Nurudeen O, Amund OO (1997) Heredita
126:161
32. Serrano-Garcia L, Monteiro-Montoya R (2001) Environ Mol
Mutagen 38:38
33. Fernandes TCC (2005) Investigação dos efeitos tóxicos,
mutagênicos e genotóxicos do herbicida trifluralina, utilizando
Allium cepa e Oreochromis niloticus como sistemas-teste. Dis-
sertação de Mestrado em Biologia Celular e Molecular—Insti-
tuto de Biociências—UNESP, Rio Claro, SP
34. Leme DM, Agnelis DF, Marin-Morales MA (2008) Aquat Toxi-
col 88:214
35. Mazzeo DEC, Fernandes TCC, Marin-Morales MA (2011) Che-
mosphere 85:13
36. Rank J, Nielsen MH (1998) Mutat Res 418:113
... Existing research on bioplastics indicates a wide range of efects, from no adverse efect to efects on organism growth, reproduction, and membrane damage (Shruti & Kutralam-Muniasamy 2019;Palsikowski et al. 2018;Zuo et al. 2019), however, the majority of these studies utilize high concentrations that generally far exceed environmental relevance. No adverse efect reported is most common, though some studies do report adverse efects to organisms. ...
... These include studies on Allium sepia, Daphnia magna, and Chlamydomonas reinhardtii, with A. sepia experiencing potential cytotoxic and genotoxic efects upon exposure to PLA at treatments of either 50g or 300g per litre of fltered water for 76 days, and D. magna and C. reinhardtii exposed to PHB nanoplastics exhibit reduced growth and severe membrane damage after a 2-day exposure (Shruti & Kutralam-Muniasamy 2019;Napper & Thompson 2019;Palsikowski et al. 2018). Plants experience a change in soil chemistry, causing them to distance further from one another, therefore decreasing density (Shruti & Kutralam-Muniasamy 2019;Napper & Thompson 2019). ...
... BDFs could be left in agricultural fields after use and then degraded into CO 2 and H 2 O by soil microorganisms (Bandopadhyay et al., 2018;Bettas Ardisson et al., 2014). However, the total degradation of BDFs in farmland conditions is rarely observed (Li et al., 2014;Palsikowski et al., 2017b). In addition, Sintim and Flury (2017) expressed their concerns about the toxicity of biodegradable material and indicated that "out-of-sight does not mean they are safe". ...
... However, Whitacre (2014) proposed that current biodegradable materials available on the market were more prone to break down into smaller particles than actually biodegrade, which in turn would generate more bio-microplastics. The toxicity and ecological effects of these biomicroplastics in soil-plant systems remained unclear (Palsikowski et al., 2017b;Sintim and Flury, 2017). As a result, we decided to incorporate these biodegradable microplastics into our study and thus, two types of microplastics were examined. ...
... PBAT and LDPE have been reported as safe and nontoxic plastics ( Anon 2007 ;Palsikowski et al., 2018 ). Palsikowski et al. ( Palsikowski et al., 2018 ) report that PBAT films do not show any cytotoxic, genotoxic, and mutagenic effects on meristematic cells of Allium cepa. ...
... PBAT and LDPE have been reported as safe and nontoxic plastics ( Anon 2007 ;Palsikowski et al., 2018 ). Palsikowski et al. ( Palsikowski et al., 2018 ) report that PBAT films do not show any cytotoxic, genotoxic, and mutagenic effects on meristematic cells of Allium cepa. However, after exposure to UV radiation, the small organic low molecular weight molecules, including carboxylic acids, are formed due to the photooxidation of PBAT, and these molecules are toxic. ...
Article
Full-text available
Biodegradable polymers have been regarded as a promising solution to tackle the pollutions caused by the wide use of conventional polymers. However, during the biodegradation process, the material fragmentation leads to microplastics. In this work, the formation of microplastics from biodegradable poly (butylene adipate-co-terephthalate) (PBAT) in different aquatic environments was investigated and compared with the common non-biodegradable low-density polyethylene (LDPE). The results showed that a much larger quantity of plastic fragments/particles were formed in all aquatic environments from PBAT than from LDPE. In addition, UV-A pretreatment, simulating the exposure to sunlight, increased the rate of PBAT microplastic formation significantly. The size distribution and shapes of the formed microplastics were systematically studied, along with changes in the polymer physicochemical properties such as molecular weight, thermal stability, crystallinity, and mechanical properties, to reveal the formation process of microplastics. This study shows that the microplastic risk from biodegradable polymers is high and needs to be further evaluated with regards to longer timeframes, the biological fate of intermediate products, and final products in freshwater, estuarine and seawater natural habitats. Especially, considering that these microplastics may have good biodegradability in warmer 20 – 25 degree water but will most likely be highly persistent in the world's cold deep seas.
... Nevertheless, a limited number of studies have addressed the toxicity of those coming from biodegradable plastics, the materials expected to replace traditional oil-based plastics for a wide variety of applications (Green et al., 2016(Green et al., , 2021Palsikowski et al., 2018;Yokota and Mehlrose, 2020;Zhang et al., 2021;Zhuikov et al., 2021;Zimmermann et al., 2020). To our knowledge, there is only one previous study that tested secondary NPs from a biodegradable polymer , but there is a complete lack of studies regarding the toxicity of PCL byproducts in aquatic environments. ...
Article
Full-text available
Bioplastics are thought as a safe substitute of non-biodegradable polymers. However, once released in the environment, biodegradation may be very slow, and they also suffer abiotic fragmentation processes, which may give rise to different fractions of polymer sizes. We present novel data on abiotic hydrolytic degradation of polycaprolactone (PCL), tracking the presence of by-products during 132 days by combining different physicochemical techniques. During the study a considerable amount of two small size plastic fractions were found (up to ∼ 6 mg of PCL by-product/g of PCL beads after 132 days of degradation); and classified as submicron-plastics (sMPs) from 1 μm to 100 nm and nanoplastics (NPs, <100 nm) as well as oligomers. The potential toxicity of the smallest fractions, PCL by-products < 100 nm (PCL-NPs + PCL oligomers) and the PCL oligomers single fraction, was tested on two ecologically relevant aquatic primary producers: the heterocystous filamentous nitrogen-fixing cyanobacterium Anabaena sp. PCC 7120, and the unicellular cyanobacterium Synechococcus sp. PCC 7942. Upon exposure to both, single and combined fractions, Reactive Oxygen Species (ROS) overproduction, intracellular pH and metabolic activity alterations were observed in both organisms, whilst membrane potential and morphological damages were only observed upon PCL-NPs + PCL oligomers exposure. Notably both PCL by-products fractions inhibited nitrogen fixation in Anabaena, which may be clearly detrimental for the aquatic trophic chain. As conclusion, fragmentation of bioplastics may render a continuous production of secondary nanoplastics as well as oligomers that might be toxic to the surrounding biota; both PCL-NPs and PCL oligomers, but largely the nanoparticulate fraction, were harmful for the two aquatic primary producers. Efforts should be made to thoroughly understand the fragmentation of bioplastics and the toxicity of the smallest fractions resulting from that degradation.
... The chromosome breakage or fusion, formation of dicentric chromosomes, cross-links between proteins and chromosomes, unequal chromatid exchange, or stickiness could be the responsible of anaphase bridge (Fig. 2d, Feretti et al. 2007: Dutta et al. 2018. Atypical scattering of chromosomes during replication or chromosome segregation may lead to polyploidy (Fig. 2e, Nefic et al. 2013;Palsikowski et al. 2018). Unlike our results, clopyralid did not induce CAs in Chinese hamster lung cells (Wang et al. 2012) and in mammalian bone morrow cells (Ilyushina et al. 2019). ...
Article
Full-text available
Clopyralid is one of the synthetic pyridine-carboxylate auxin herbicides and used to control perennial and annual broadleaf weeds in wheat, sugar beets, canola, etc. In this study, dose-dependent cytotoxicity and genotoxicity of clopyralid at different concentrations (25, 50, and 100 μg/mL) have been evaluated on the Allium cepa roots. The evaluation has been performed at macroscopic (root growth) and microscopic levels [mitotic index (MI), chromosome aberrations (CAs) in ana-telophase cells, and DNA damage] using root growth inhibition, Allium ana-telophase, and comet tests. The percentage of root growth inhibition and concentration of reducing root growth by 50% (EC50) of clopyralid were determined compared with the negative control by using various concentrations of clopyralid (6.25–1000 μg/L). The 96 h EC50 of clopyralid was recorded as 50 μg/L. The gradual decrease in root growth and the MI reveals the cytotoxic effects of clopyralid. All the tested concentrations of clopyralid induced total CAs (polyploidy, stickiness, anaphase bridges, chromosome laggards, and disturbed ana-telophase) and DNA damage dose and time dependently. These results confirm the cytotoxic and genotoxic effects of clopyralid on non-target organism.
... This practice emphasizes the need for clear answers about the ecotoxicological effects of PLA. Currently, few papers can provide much information about the biodegradation and ecotoxicology of PLA in soil (Palsikowski et al., 2018;Adhikari et al., 2016). As a result, the effects of PLA microplastics on marine biota remains largely unexplored. ...
Article
The substitution of petrochemical plastics by bio-based and biodegradable plastics are in need of an evaluation for the potential toxic impacts that they can have on marine wildlife. This study aims to assess the toxicological effects of polylactic acid microparticles at two concentrations, 10 and 100 μg/L, during 8 days on the blue mussel, Mytilus edulis. No significant oxidative stress (catalase, glutathione-S-transferase and superoxide dismutase activities), neurotoxicity (acetylcholinesterase), or immunotoxicity (lysosomal membrane stability and acid phosphatase activity) were detectable. The multivariate analysis of metabolomic data allowed us to differentiate the individuals according to the exposure. From the loading plot of OPLS-DA, 48 ions down-regulated in the individuals exposed to microplastics. They were identified based on HRMS data as glycerophospholipids.
Article
Full-text available
The emergence of the SARS‐CoV‐2 pandemic and airborne particulate matter (PM) pollution has led to remarkably high demand for face masks. However, conventional respirators are intended for single use and made from nondegradable materials, causing serious concern for a plastic‐waste environmental crisis. Furthermore, these facemasks are weakened in humid conditions and difficult to decontaminate. Herein, a reusable, self‐sustaining, highly effective, and humidity‐resistant air filtration membrane with excellent particle‐removal efficiency is reported, based on highly controllable and stable piezoelectric electrospun poly (l‐lactic acid) (PLLA) nanofibers. The PLLA filter possesses a high filtration efficiency (>99% for PM 2.5 and >91% for PM 1.0) while providing a favorable pressure drop (≈91 Pa at normal breathing rate) for human breathing due to the piezoelectric charge naturally activated by respiration through the mask. The filter has a long, stable filtration performance and good humidity resistance, demonstrated by a minimal declination in the filtration performance of the nanofiber membrane after moisture exposure. The PLLA filter is reusable via common sterilization tools (i.e., an ultrasonic cleaning bath, autoclave, or microwave). Moreover, a prototype of a completely biodegradable PLLA nanofiber‐based facemask is fabricated and shown to decompose within 5 weeks in an accelerated degradation environment. The piezoelectric nanofibers of poly (l‐lactic acid) (PLLA) are employed to fabricate a reusable, moisture‐resistant, and highly effective facemask filter with long‐term biodegradability. The PLLA filter could offer an eco‐friendly solution to preventing the transmission of highly infectious viruses and resolving the environmental crisis caused by the massive use of current permanent plastic facemask filters.
Thesis
En réponse aux problématiques posées par la persistance des matériaux polymères conventionnels tels que les polyoléfines dans l’environnement, ainsi que par l’utilisation de ressources fossiles, de nombreuses alternatives biosourcées et/ou biodégradables apparaissent sur le marché. Dans une démarche d’éco-conception, lors de la formulation d’un nouveau matériau pour en moduler les performances, les propriétés d’usage, la mise en oeuvre, mais aussi la fin de vie doivent être considérées dans le but de proposer des compromis visant à réduire l’empreinte environnementale globale. La thèse propose une méthodologie visant à guider l’anticipation de la fin de vie dès l’étape de conception, en particulier en vue de valorisations organiques par traitement biologique. Cette méthodologie mise en place vise à favoriser le développement de matériaux à fin de vie contrôlée et à identifier les paramètres influençant leur dégradation. Un ensemble d’études permettant d’évaluer l’hydrolysabilité abiotique et enzymatique, la biodégradabilité dans des boues de station d’épuration, ainsi que la fragmentation dans des milieux scénarisés, ont été mis en place, à l’aide de séries de films biodégradables. Il s’agit de matériaux à base de polyesters : poly(butylène succinate) ou poly(butylène succinate-co-adipate), mélangés avec de la lignine, de l’acide polylactique ou de l’amidon. Afin d’assurer la compatibilité de ces mélanges, des liquides ioniques et solvants eutectiques ont été employés. L’impact de ces additifs sur les propriétés de biodégradabilité des matériaux polymères a été peu étudié dans la littérature. La thèse permet dans une certaine mesure d’établir un lien entre formulation et dégradabilité, grâce à la caractérisation des échantillons à l’état initial et au suivi de leurs propriétés après dégradation, et de proposer des pistes visant à la conception de matériaux à dégradabilité contrôlée.
Article
The fungi are becoming the distinguished organisms utilized in the biological synthesis of metallic nanoparticles because of their metal bioaccumulation ability. Addressed herein, the extracellular synthesis of silver nanoparticles (AgNPs) was carried out by using the cell‐free filtrate of Penicillium toxicarium KJ173540.1. P. toxicarium was locally isolated and identified using both classical and molecular methods according to ribosomal internal transcribed spacer area of 18S rDNA. The optimum conditions for the AgNPs synthesis were found as 0.25 mM AgNO3 concentrations with pH 12 values at 45°C after 64 hr incubation in dark. Biosynthesized AgNPs were characterized via microscopic and spectroscopic techniques such as transmission electron microscopy, scanning electron microscopy, energy dispersive X‐ray analysis, Fourier transform infrared spectrophotometer, and ultraviolet–visible spectroscopy. Zetasizer measurements presented that the high negative potential value (−18.1 mV) and PDI (0.495) supported the excellent colloidal nature of AgNPs with long‐range stability and high dispersity. AgNPs exhibited cyto–genotoxicity in Allium cepa root meristem cells by decreasing mitotic index and increasing chromosome aberrations in a dose‐dependent manner. Then, 100 and 50% concentration of biosynthesized AgNPs showed antibacterial activity on Staphylococcus aureus and Bacillus subtilis. A decreasing biofilm formation of Pseudomonas aeruginosa 80.69, 48.32, and 28.41% was also observed at 100, 50, and 25% of mycosynthesized AgNP, respectively.
Preprint
Full-text available
Clopyralid is a one of the synthetic pyridine-carboxylate auxin herbicides and used to control perennial and annual broadleaf weeds in wheat, sugar beets and canola etc. In this study, dose dependent cytotoxicity and genotoxicity of clopyralid at different concentrations (25, 50, and 100 µg/mL) on the Allium cepa roots were evaluated at macroscopic (root growth) and microscopic levels (Mitotic index (MI), chromosome aberrations (CAs) in ana-telophase cells and DNA damage) using root growth inhibition, Allium ana-telophase and comet tests. The percentage root growth inhibition and concentration reducing root growth by 50% (EC 50 ) of clopyralid in relation to the negative control were determined by using various concentrations of clopyralid (6.25–1000 µg/L). The 96 h EC 50 of clopyralid was recorded as 50 µg/L. The gradual decrease in root growth and the MI reveals the cytotoxic effects of clopyralid. All the tested concentrations of clopyralid induced total CAs (polyploidy, stickiness, anaphase bridges, chromosome laggards, and disturbed ana-telophase) and DNA damage dose and time dependently. This study confirmed cytotoxic and genotoxic effects of clopyralid on non-target organism.
Article
Full-text available
This paper presents a study of biodegradation, in soil, of samples of poly(butylene adipate-co-terephthalate)(PBAT), poly(lactic acid) (PLA) and blends of these materials prepared in torque rheometer with the addition of a chain extender. Film samples of these materials were buried in soil under controlled laboratory conditions. The degraded samples were regularly taken from soil and analyzed by visual inspection, size exclusion chromatography, differential scanning calorimetry and infrared spectroscopy. Respirometry biodegradation tests were conducted to assess samples mineralization degree. Blends showed higher degree of crystallinity compared to pure polymers. Crystallinity degree enhanced during the biodegradation process in all samples, being able to causing the samples to degrade slowly. The study showed the great complexity of the biodegradation process of PLA and PBAT blends when compatibilized with a chain extender. The biodegradation rate showed different results depending on the characteristic applied to evaluate it: visual, molecular weight or mineralization. The chain extender had strong influence in PBAT and blends degradation, slowing the process as observed by the variation of molecular weight and carbonyl index. Blends showed an intermediate behavior compared to the original polymers.
Article
Full-text available
The origin of hydrolysis-induced nanofibrillation and crystallization, at the molecular level, was revealed by mapping the conformational ordering during long-term hydrolytic degradation of initially amorphous poly(lactic acid) (PLA)-a representative model for degradable aliphatic polyesters generally displaying strong interplay between crystallization and hydrolytic erosion. The conformational regularization of chain segments was essentially the main driving force for the morphological evolution of PLA during hydrolytic degradation. For hydrolysis at 37 °C, no significant structural variations were observed due to the immobilization of "frozen" PLA chains. In contrast, conformational ordering in PLA was immediately triggered during hydrolysis at 60 °C and was responsible for the transition from random coils to disordered trans, and further to quasi-crystalline nanospheres. On the surfaces, the head-by-head absorption and joining of neighboring nanospheres led to nanofibrillar assemblies following a "gluttonous snake"-like manner. The length and density of nanofibers formed were in close relation to the hydrolytic evolution, both of which showed a direct rise in the initial 60 days and then a gradual decline. In the interior, presumably the high surface energy of the nanospheres allowed for the preferential anchoring and packing of conformationally ordered chains into lamellae. In accordance with the well-established hypothesis, the amorphous regions were attacked prior to the erosion of crystalline entities, causing a rapid increase of crystallinity during the initial 30 days followed by a gradual fall until 90 days. In addition to the illustration of hydrolysis-induced variations of crystallinity, our proposed model elucidates the formation of spherulitic nuclei featuring an extremely wide distribution of diameters ranging from several nanometers to over 5 micrometers, as well as the lower resistance to hydrolysis observed for the primary nuclei. Our work fuels the interest in controlling nanofibrillation mechanism during hydrolysis of PLA, opening up possibilities for straightforward nanofiber formation.
Article
Full-text available
In this study, nanocomposites of PLA and organoclays Cloisite 20A and Cloisite 30B were prepared by the melt intercalation method and the obtained samples were characterized by transmission electron microscopy (TEM). Since composting is an important proposal to the final disposal of biopolymers, the influence of clays on the hydrolytic degradation process of PLA was evaluated by visual analysis and monitoring of molecular weight after periods of 15 and 30 days of degradation in compost. After degradation of the materials in composting environment, the evaluation of cytotoxic, genotoxic and mutagenic effects of compost aqueous extract was carried out using a bioassay with Allium cepa as test organism. The TEM micrographs permitted the observation of different levels of dispersion, including exfoliated regions. In the evaluation of hydrolytic degradation it was noted that the presence of organoclays can decrease the rate of degradation possibly due to the barrier effect of clay layers and/or the higher degree of crystallinity in the nanocomposite samples. Nevertheless, even in the case of nanocomposites, the molecular weight reduction was significant, indicating that the composting process is favorable to the chain scission of PLA in studied materials. In the analysis performed by the bioassay using A. cepa as test organism, it was found that after degradation of the PLA and its nanocomposites the aqueous extract of compost samples induced a decreasing in the mitotic index and an increasing in the induction of chromosomal abnormalities. These results were statistically significant in relation to the negative control (distilled water). By comparing the results obtained for the nanocomposites in relative to pure polymer, there were no statistically significant differences. The types of the observed chromosomal aberrations indicated a possible genotoxic effect of the materials, which may be related to an aneugenic action of PLA degradation products.
Article
Full-text available
Willow (Salix sp.) sawdust (WS) is used as a component of growing media in horticultural crops and potted plant production in Santa Fe, Argentina. We evaluated the use of the germination bioassay as an indicator of WS maturity/phytotox‐icity by comparing bioassays results with dry matter production of plants growing in the same substrate. Lettuce (Lactuca sativa L.) seeds were sown on filter paper moistened with extracts of WS composted for different times. Germinated seeds were counted (G) and the radicle growth (L) measured. Germination index (Gi) = G/G0 × L/L0 × 100, where G0 and L0 are values obtained using distilled water (control). The global germination index (GI) was the Gi average of the 50 and 75% extract dilution. GI of lettuce increased from 5% in the non‐composted WS to 93.3% in the WS composted for 40 days. GI did not change significantly after 40 days of composting. Calendula dry matter production increased from 8 g/plant in non‐composted WS to 17.1 g/plant in WS composted for 40 or 60 days. The addition of nitrogen and pH corrector did not affect GI of the WS growth media but they both increased calendula dry matter production to 21.1–22.7 g/plant when WS composted for 40 or 60 days was used as a growth medium.
Article
Blends containing 75 wt. % of an amorphous polylactide (PLA) with two different molecular weights and 25 wt. % of a poly[(butylene adipate)-co-terephthalate] (PBAT) were prepared using either a Brabender batch mixer or a twin-screw extruder. These compounds were selected because blending PLA with PBAT can overcome various drawbacks of PLA such as its brittleness and processability limitations. In this study, we investigated the effects of varying the molecular weight of the PLA matrix and of two different mixing processes on the blend morphology and, further, on droplet coalescence during shearing. The rheological properties of these blends were investigated and the interfacial properties were analyzed using the Palierne emulsion model. Droplet coalescence was investigated by applying shear flows of 0.05 and 0.20 s−1 at a fixed strain of 60. Subsequently, small amplitude oscillatory shear tests were conducted to investigate changes in the viscoelastic properties. The morphology of the blends was also examined using scanning electron microscope (SEM) micrographs. It was observed that the PBAT droplets were much smaller when twin-screw extrusion was used for the blend preparation. Shearing at 0.05 s−1 induced significant droplet coalescence in all blends, but coalescence and changes in the viscoelastic properties were much more pronounced for the PLA-PBAT blend based on a lower molecular weight PLA. The viscoelastic responses were also somehow affected by the thermal degradation of the PLA matrix during the experiments.
Article
This paper revealed the compostability of poly (lactic acid) (PLA) and PLA/starch blends with various amounts of starch contents. The results showed that the ultimate aerobic biodegradation under controlled composting conditions of PLA and PLA/starch with 30, 50 and 70 wt% starch contents were 83.43%, 84.28%, 88.04% and 95.83%, respectively. Under the same testing conditions, the biodegradation of cellulose, as a positive material, was 84.89%. In the disintegration testing, the tested materials were completely biodegraded and no residuals were observed through visual inspection after 30 days. In ecotoxicity test, the rate of germination and plant growth of monocotyledon and dicotyledon on the resulting compost were no significant different when compared to blank compost under. It can be concluded that the PLA and PLA/starch blends were clearly safe for the ecosystem. Furthermore, these materials were biodegradable and compostable materials as they pass all requirements of ISO 17088.
Article
Estadual Paulista, como parte dos requisitos para obtenção do título de Mestre em Ciências Biológicas (Biologia Celular e Molecular).
Article
Agricultural mulch film of poly(lactic acid) (PLA) has been prepared under industrial conditions by the extrusion blown film method with modified properties using poly(hydroxibutyrate) (PHB) and reactive compatibilizer maleic anhydride (MA). Processing parameters and blend composition have been optimized based upon processability and mechanical properties of the final materials. Because PLA is a biopolymer, evaluation of service life period of the mulch film is very much important. Sustainability of the film has been analyzed by keeping the films in a weatherometer, which can create accelerated weather conditions, followed by mechanical testing at regular intervals. Similarly, variation in compostability has been analyzed as per the American standard for test method, ASTM D 5988, using vermi-compost. In addition, specific microbial action on the mulch films also has been analyzed using bacteria Berkholdaria cepacia (B. cepacia), which is selective in particular toward PLA degradation and in mixed fungal inoculums.
Article
In the last decades, the solid-waste management related to the extensively growing production of plastic materials, in concert with their durability, have stimulated increasing interest in biodegradable polymers. At present, a variety of biodegradable polymers has already been introduced onto the market and can now be competitive with non biodegradable thermoplastics in different fields (packaging, biomedical, textile, etc.). However, a significant economical effort is still directed in tailoring structural properties in order to further broaden the range of applications without impairing biodegradation. Improving the performance of biodegradable materials requires a good characterization of both physico-chemical and mechanical parameters. Polymer analysis can involve many different features including detailed characterization of chemical structures and compositions as well as average molecular mass determination. It is of outstanding importance in troubleshooting of a polymer manufacturing process and for quality control, especially in biomedical applications. This review describes recent trends in the structural characterization of biodegradable materials by modern mass spectrometry (MS). It provides an overview of the analytical tools used to evaluate their degradation. Several successful applications of MALDI-TOF MS (matrix assisted laser desorption ionization time of flight) and ESI MS (electrospray mass spectrometry) for the determination of the structural architecture of biodegradable macromolecules, including their topology, composition, chemical structure of the end groups have been reported. However, MS methodologies have been recently applied to evaluate the biodegradation of polymeric materials. ESI MS represents the most useful technique for characterizing water-soluble polymers possessing different end group structures, with the advantage of being easily interfaced with solution-based separation techniques such as high-performance liquid chromatography (HPLC).