ISOLATION OF POLYETHYLENE PLASTIC DEGRADING-BACTERIA
Dede Mahdiyah1*‡, Elpawati1, Bayu Hari Mukti2 1
2
UIN Syarif Hidayatullah Jakarta
Pendidikan Biologi, STKIP Banjarmasin, South Borneo
*correspondence author: Telepon: 0511-3268105, Fax: 0511-3270134, E-mail:
[email protected] ‡
Current author: Academy of Midwifery Sari Mulia Banjarmasin, South Borneo
Abstract Plastic polyethylene waste accumulating in the environment are posing an ever increasing ecological threat. Bacteria was isolated used NA medium and TSA medium were incubated for five days. Isolates were resulted towards analyzed 1 month of incubation in liquid culture method at 37 oC with agitation 130 rpm. The biodegradation of polyethylene was resulted by isolate of 22 TSB (used medium Tryptic Soy Broth) were 17% in 1 month. Microbial counts in the degrading materials were recorded up 3.08 x 106. The microbial species found associated with the degrading materials were identified as Gram positive (shape rod, have no spore). Key Words: bacteria, biodegradation, polyethylene, plastic. Introduction During the past 3-decades, plastic materials have been increasingly used in food clothing, shelter, transportation, construction, medical, and recreation industries. Plastics are advantageous as they are strong, light-weighted, and durable. However, they are disadvantageous as they are resistant to biodegradation, leading to pollution, harmful to the natural environment. Increasing environmental pollution and waste that can not be renewed and degrade it encourages research and studies in the field of biosynthetic and biodegradation material. One of the waste that can not be destroyed is plastic waste, which is a type of polyethylene plastic.
Low density polyethylene is one of the major sources of environmental pollution. Polyethylene is a polymer made of long chains of ethylene monomers. The use of polyethylene growing worldwide at a rate of 12% per year and about 140 million tons of synthetic polymers are produced worldwide each year. With such a large amount of polyethylene gets accumulated in the environment, generating plastic waste ecological problems are needed thousands of years to efficiently degradation (Usha et al. 2011). Microorganisms can degrade plastic over 90 genera, from bacteria and fungi, among them; Bacillus megaterium, Pseudomonas sp., Azotobacter, Ralstonia eutropha, Halomonas sp., etc. (Chee et al. 2010). Plastic degradation by microbes due to the activity of certain enzymes that cause cleavage of the polymer chains into monomers and oligomers. Plastic that has been enzymatically broken down further absorbed by the microbial cells to be metabolized. Aerobic metabolism produces carbon dioxide and water. Instead of anaerobic metabolism produces carbon dioxide, water, and methane as end products (Usha et al. 2011). This study aims to isolate the bacteria from waste polyethylene plastics that can degrade polyethylene plastic. Materials and Methods Isolation of Microorganisms degrading Plastics from Waste Plastics Polyethylene (Gupta et al. 2010) Polyethylene plastic waste in the capture of landfill Kampung Utan Ciputat washed with sterile distilled water and then cut with sterile scissors. Then inoculated onto media containing 100 ml of sterile distilled water in a 250 mL erlenmeyer. In doing serial dilutions from 10-1 to 10-6 with sterile 0.9% NaCl. Three recently taken 0.1 mL dilution was inoculated into media for NA and TSA. Then spread the rod L and then incubated in an incubator at 37 ° C for seven days. After seven days of incubation observed colony growth. Colonies were grown in petri dishes scratch media to NA and TSA back then incubated at 37 ° C for 7 days. Colonies that grew were then purified to agar slant NA and TSA for further testing.
Determination of Degradation of Polyethylene Plastic Waste (Kathiresan 2003) Weigh polyethylene plastic (initial weight) and then washed with sterile distilled water and sprayed with 70% alcohol. Plastic is inserted into the 100 mL erlenmeyer containing NB and TSB media as much as 50 mL aseptically. So as much as 2 loops inoculated bacterial isolates to the media. Then incubated in an incubator shaker at room temperature, with agitation of 130 rpm for a month. Polyethylene plastic that has been incubated for a month, washed with sterile distilled water and then sprayed with alcohol dried aired then weighed (final weight). Determination of the percentage of degradation of polyethylene plastic by bacteria by: %Degradation = 1-final weight x 100% initial weight Physiological and Biochemistry Test degrading microorganisms Polyethylene Plastic Potentially degrading bacteria tested physiological polyethylene plastic, the morphology, color, texture, Gram staining and spore staining. Results Acquired as much as 25 isolates of the isolated microorganisms were isolated from plastic waste polyethylene derived from landfill in Kampung Utan Ciputat NA and TSA media. The 25 isolates was purified in order to tilt to the next test (Figure 1). As many as 25 isolates obtained polyethylene plastic degradation determined by incubation for 1 month in an incubator shaker at 130 rpm agitation in a 37 ° C temperature conditions. Acquired eight isolates of potentially degrading polyethylene plastic with different percentages. The eight isolates, namely (1 isolates in TSB media, isolates 6 on TSB media, isolates 11 at TSB media, isolates 22 on TSB media, isolates 17 on NB medium, isolates 15 in NB media, isolates 30 on NB medium, and isolates 21 on NB medium). Of the eight isolates had percentages of different degradation and the highest degradation obtained by isolate of 22 TSB at 17%, then the other isolates have the capacity degradation of plastics: isolate of 1 TSB (6%), isolate of 6 TSB (12% ),
isolate of 11 TSB (4%), isolate of 17 NB (4%), isolate of 15 NB (5%), isolate of 30 NB (6%), and isolate of 21 NB (5%) (Figure 2). The eight potential degrading isolates were characterized morphologically polyethylene plastic, shape, Gram staining and spore staining. Of these eight isolates including Gram negative 1 TSB spherical shape, have no spore. Six isolates including Gram-negative in TSB media and round shape. Isolate of 11 TSB including Gram-negative, round shape and have no spore. Isolate of 22 TSB including Gram-positive, rod-shaped and have no spore. Isolate of 15 NB including Gramnegative bacteria are round. Isolate of 21 TSB including Gram-positive rod-shaped bacterium and have no spore. Isolate of 30 NB including Gram-negative bacteria round shape and isolate of 17 NB including Gram negative spherical shape (Figure 3). Discussion 22 isolates using TSB medium isolated from polyethylene plastic waste polyethylene plastics successfully degrade by 17% under anaerobic conditions. Anaerobic Biodegradation is an important component of natural attenuation of contaminants at many hazardous waste sites. This is the breakdown of organic contaminants by microorganisms when oxygen is absent. Some anaerobic bacteria use nitrate, sulfate, iron, manganese, and carbon dioxide as an electron acceptor, and organic chemicals break down into smaller compounds (Priyanka 2011). Phanerochaete chrysosporium has been found to degrade LDPE blended with starch incubated in soil.
P.
chrysosporium was able to degrade LDPE more than 50%, then P. chrysosporium grown on media other than soil degradation occurs only 12% (Orhan and Buyukgungor 2000). Polyethylene has a high molecular weight can be degraded by fungi that degrade lignin under conditions of limited nitrogen and carbon limited and by manganese peroxidase. Fungi such as Mucor rouxii NRRL 1835 and Aspergillius flavus and some Streptomyces strains capable of degrading polyethylene containing 6% starch (Premraj and Mukesh 2005). Isolate of 22 TSB has a physiological characteristic that is Gram-positive, rod shape, and has no spores. The ability of isolates 22 TSB in degrading polyethylene in the presence of an
enzyme stimulator that can accelerate catalytic reaction substrates, because TSB isolates grew well in the higher alkanes. Polyethylene is also relatively firmly linked to the bacterial surface, increased hydrophobicity, and mediates adhesion of bacteria on the surface of the substrate and passive transport of substrate molecules. Percentage of polyethylene degradation by bacterial isolates 22 TSB (under shaker culture) is quite high in the incubation period of one month (17%) were anaerobic in isolation from polyethylene plastic waste in Kampung Utan Ciputat. The bacteria are able to grow well on alkanes and able to utilize oxidized PE as a substrate similar chemical structure, it also can produce strain biosurfactant required for the mobilization of soluble hydrophobic substrate molecules. Biodegradation of plastics take an active part in different soil conditions according to the nature of the bacteria, because the microorganisms responsible for the degradation different from each other and have optimal growth of bacteria in the soil. It is also based on research Katherisan (2003) these bacteria Pseudomonas sp. able to degrade the plastic by 8.16% and was able to degrade the polythene by 20.54% within one month incubation anaerobically. While this type of fungi Aspergillus Glaucus able to degrade the plastic by 7.26% and was able to degrade the polythene by 28.80% within one month incubation anaerobically. From the results of the degradation of polythene faster and easier than plastic degradation. Polythene and plastics are two polymers with wide application, both are recalcitrant and thus remain inert to degradation and damage that leads to accumulation in the environment, and create serious environmental problems. Therefore, further research is needed to prevent environmental damage caused by plastic and polythene waste contamination. Have been isolated and characterized bacteria that can degrade polyethylene plastic derived from polyethylene insulation plastic waste by 25 isolates of bacteria. Isolates that have a high percentage of degradation (17%) isolates were 22 TSB. Isolate is a Gram positive rod shape and do not have flagella and have no spores.
Acknowledgements We are very grateful to State Islamic University Syarif hidayatullah Jakarta, which has funded this research through funding DIPA.
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Figure 1 isolate bacterial isolation results of polyethylene plastic waste with NA and TSA media. 30 % Degradation
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Figure 2. Percentage degradation of polyethylene plastic by bacteria isolated from polyethylene plastic waste.
Figure 3. characterization of polyethylene plastic degrading bacteria
