Degradation and depolymerization of plastic waste by local bacterial isolates and

bubble column reactor

Amal A. Hussein, Mohammed Alzuhairi, and Noor H. Aljanabi

Citation: AIP Conference Proceedings 1968, 030081 (2018); doi: 10.1063/1.5039268

View online: https://doi.org/10.1063/1.5039268
View Table of Contents: http://aip.scitation.org/toc/apc/1968/1
Published by the American Institute of Physics
 Degradation and Depolymerization of Plastic Waste by
Local Bacterial Isolates and Bubble Column Reactor
Amal A. Hussein1,a), Mohammed Alzuhairi2,b) and Noor H. Aljanabi1
1
Department of Applied Science, University of Technology – Baghdad, Iraq.
2
Department of Material Engineering, University of Technology-Baghdad, Iraq.
b)
Corresponding author: dr.alzuhairi@gmail.com
a)
amelali71@yahoo.com

Abstract. Accumulation of plastics, especially Polyethylene terephthalate (PET), is an ever increasing ecological threat
due to its excessive usage in everyday human life. Nowadays, there are many methods to get rid of plastic wastes
including burning, recycling and burying. However, these methods are not very active since their long period, anaerobic
conditions that increase the rate of toxic materials released into the environment. This work aims to study the biological
degradation of PET microorganism isolated from soil sample. Thirty eight (38) bacterial isolates were isolated from ten
soil and plastic waste sample collected from four different waste disposal sites in Baghdad city during different periods
between December 2016 and March 2017. Isolation was performed using enrichment culture method (flasks method) by
culturing the soil samples in flasks with MSM medium where there is no carbon source only PET. Results showed that
Al-Za’farania sample gave a higher number of isolates (13 isolates), while other samples gave less number of isolates.
Screening was performed depending on their ability to grow in liquid MSM which contains PET powder and pieces and
change the color of the PET-emulsified liquid medium as well as their ability to form the clear zone on PET-MSM agar.
The results showed that NH-D-1 isolate has the higher ability to degrade DPET and PET pieces. According to
morphological, biochemical characterization and Vitek-2 technique, the most active isolate was identified as
Acinetobacter baumannii.

Keywords: Polyethylene terephthalate pieces, plastic wastes, Acinetobacter baumannii, biodegradation.

INTRODUCTION
Plastic is non-biodegradable, powerful, potent, light weight, moisture resistant polymer of carbon, hydrogen,
sulphur, nitrogen and other organic and inorganic elements made of fossil fuel which is a non-renewable
source [1]. Depending on the type of their chemical reaction, plastics are classified into thermoplastics
(undergo strong molecular motion when heated, which causes them to soften and harden when cooled, and
repeated heating and cooling allows them to be molded into a variety of different shapes) and thermoset
(They are solidified after being melted by heating). The process of changing from the liquid state to the solid
state is irreversible). Based on the chemical structure, they are classified to polystyrene (PS), polyethylene
terephthalate (PET), polypropylene (PP), low density polyethylene (LDPE), high density polyethylene
(HDPE), and polycarbonate. It is clear that in the last decade’s plastics become very important. Nowadays,
most of products are either made of plastic or manufactured in plastic containing equipment. Large quantities
of plastics are unfortunately being treated as wastes after they have served their main purpose [2]. In general,
plastics like: polyethylene (PE), polystyrene (PS), polyethylene terephthalate (PET), polypropylene (PP), and
polyvinyl chloride (PVC), are largely used in packaging and industries that needs to be recycled. Polyethylene
terephthalate PET is one of the most interesting thermoplastic materials that belong to PS group. There are
used in most industries such as films, packaging, and textile fibers

MATERIALS AND METHODS

Polyethylene terphthalate (pet)
The polyethylene terephthalate (PET) pieces used in this study were obtained from local water bottles. PET
was cut into small pieces of about (2cm×2cm).

Technologies and Materials for Renewable Energy, Environment and Sustainability

AIP Conf. Proc. 1968, 030081-1–030081-11; https://doi.org/10.1063/1.5039268
Published by AIP Publishing. 978-0-7354-1675-8/$30.00

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 Preparation of pet powder
As a first step, 100 gm of PET pieces mixed with 116 ml of ethylene glycol (EG) (4:1 EG, PET molar
ratio) with 0.05% Nano-Magnesium Oxide as a catalyst depending on the weight of PET at EG boiling point
for 40 min until fully depolymerization. Treatment with heat involve complete condensation using refluxing
unit in bubble column reactor ( zero material loss ) using glass condenser cooling with water, then separate
unreacted ethylene glycol out of mixture leaving the final form of PET as in the fig. (1,2) [3]:

(a) (b)

FIGURE 1. Chemical conversion of plastic bottles into powder. (a): plastic bottles pieces; (b): the final form of PET
powder after washing.

FIGURE 2. Bubble column reactor

Pet degrading bacterial isolates

The bacterial isolate has been isolated from Al-Za,farania site which is contaminated with plastic waste and
selected according from previous study and this isolate was identified as an Acinetobacter baumannii. [4].

Biodegradation assay of polyethylene by bacterial isolate

To determine the ability of the selected isolate to degrade PET, 25 milliliter of mineral salt medium (MSM)
was dispensed in Erlenmeyer flasks (100ml) and autoclaved. After sterilization, PET pieces (2cm×3cm) were
weighed accurately and placed in the sterile flasks as a substrate and sole source of carbon, the flasks were
inoculated with 1ml v/v 18hrs. old (OD = 0.5) of Acinetobacter baumannii with control (medium without

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inoculation). After 28days in aseptic conditions, plastic samples were removed from broth and were placed
on filter papers for drying [5]. The degradation ability of these bacteria was determined by monitoring the
Percentage loss in weight, FTIR spectrum for powder and piece. The change in topography of pieces surface
was determine.

RESULTS AND DISCUSSION

Very large amounts of PET bottles are now thrown away in the environment; therefore, industrial
companies are trying to exploit these wastes in different manufacturing processes by converting it into
powder in order to facilitate the mixing of PET with other industrial materials.

The conversion process was done by mixing PET pieces with EG and MgO Nano-catalyst which is an
efficient catalyst to reduce the time of chemical reaction [6].

FTIR spectrum shown in fig. (3) explains the difference between plastic bottles before and after the chemical
treatment:

(a)

(b)

FIGURE 3. FTIR spectrum for PET (a) before treatment (b) after treatment

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PET pieces were also tested for their degradation using the active bacteria, after incubation at 37ºC for 90
days. The pieces were weighed and their chemical structure was measured with FTIR spectrum, while the
surface changes have been checked with SEM.

TABLE 1. Weight measurement of PET piece before and after culturing.

No. Isolate PET piece weight (g) Wt. of PET Percentage of
symbol Before treatment After degraded degraded PET
treatment

1 Contro 0.201 0.200 0.001 0.497%

2 NH-D-1 0.201 0.145 0.055 27.363%

From the result above, NH-D-1 showed a great ability to degrade PET pieces in MSM medium which may
be represented as weight loss due to the enzymatic mechanism of bacteria to breakdown the polymer.
Depending on the results obtained from screening, NH-D-1 was select as a strongest and active isolate for
PET powder and pieces biodegradation on MSM medium as a sole source of carbon.

The pH of liquid emulsifying medium was measured for determination of pH changes to make sure of the
metabolic activity of the bacterial isolates in supplemented medium, because the metabolism of microbial
cells can also significantly assist the proof of degradation. Table (2) shows the variation in pH of the medium
during this biodegradation study, as below:

TABLE 2. Change in pH values of liquid MSM with PET powder.

No. Isolate pH value

1 Control 7.0
2 NH-D-1 5.6

The isolate NH-D-1 showed a significant decline in pH value of Inoculated medium as compared with
control. There are no results for similar researches.

Microorganisms secrete a variety of enzymes into the media, which initiate the breakdown of the polymers.
Two types of enzymes are involved in the process, namely intracellular and extracellular depolymerizes.
Exoenzymes from microorganism first breakdown the complex polymers giving short chains or monomers
that are small enough to permeate through the cell walls to be utilized as carbon and energy sources by a
process of depolymerization [7].

Analysis of the changing in pet functional units using ftir:

The FTIR was used to test the spectral changes of PET pieces or by measuring the intensity of PET bands
before and after biological treatment. The change was observed in the 2906 cm-1band intensity which
represents the functional group (C-H) [8].

A.baumannii has showed a great increase in transmission value of C-H band of the plastic piece, with
and obvious decrease in the absorption value. The substances result from that breaking down can be entirely
utilized by other microorganisms; otherwise it will be an environmental and health risk [9].

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FIGURE 4. FTIR spectrum of PET piece biologically untreated.(Control).

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 FIGURE 5. FTIR spectrum of PET piece after biological treated

FTIR was also used to test the biodegradability of PET powder in liquid MSM as below:

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FIGURE 6. FTIR spectrum for PET powder in liquid MSM without biological treatment (Control).

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 FIGURE 7. FTIR spectrum for PET powder in liquid MSM after treated with A.baumannii

SEM Spectroscopy
The changes in PET surface were illustrated using SEM. The untreated PET piece (control) showed no cracks,
pits or any attached particles on the surface fig. (8).

Before washing, the PET treated with A.baumannii showed many particles attached to the plastic surface
with no obvious pits, fig. (9).

Meanwhile washed piece wasn’t having any attached particle but clear pit showed up as seen in fig. (10).

Nakkabi et al., [10] used SEM technique to monitor the changes in PET surface which appeared as cracks
when treated with Bacillus subtilis. Sharon et al., [11] reported “Degradation impact by microbes on the

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crystalline structure and presence of microbes inside the polyethylene terephthalate were seen in scanning
electron microscopy (SEM) micrographs”

FIGURE 8. SEM untreated PET piece (Control).

FIGURE 9. SEM of the active isolate settling on the surface of PET piece (before washing).

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 FIGURE 10. SEM of the surface of PET piece affected by bacterial activity (after washing).

SEM was used to see the level of scission and attachment of the microbes on the surface of the polythene
before and after the microbial attack [12]. The biodegradation of polyethylene was evidenced through
formation of cavities on the surface of polyethylene and structural changes like erosion on the surface of PET
pieces were observed. Furthermore, polyesters when buried in the soil elicited large number of holes, cavities
and pinholes, and attributed it to the attack of microorganisms under soil environment.

CONCLUSION
Depending on the results obtained from this study, the conclusions may be given:

1. Widespread studies on the degradation of plastics have been carried out in order to overcome the
environmental problems associated with synthetic plastic waste. Biodegradation of plastics by
microorganisms seems to be the most effective process to fight against plastic waste.

2. Soil samples from landfill dump of plastic waste considered an excellent source for polymer
degrading bacteria.

3. PET can be degraded by microorganisms (biodegradation) like Acinetobacter baumannii, present

in soil contains plastic waste. The method used in the present experiment is cost effective, easy to perform,
and environmentally friendly.

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