Tuning enzyme concentration and particle size for LDPE biodegradation using lipase and laccase systems

Abstract

The persistent accumulation of low-density polyethylene (LDPE) waste in the environment has necessitated the exploration of eco-friendly degradation methods. This study aimed to degrade LDPE films using lipase (Lip) and laccase (Lac) enzymes obtained from Aspergillus flavus. The effects of enzyme concentration and LDPE particle size on the degradation rate were examined. LDPE samples were prepared in three particle sizes: 0.5, 1, and 2 cm. These samples were incubated with Lip, Lac, and a combination of both enzymes (Lip-Lac) at two concentration levels: 50% and 100% (v/v). The degradation process or extent of degradation was monitored over 10 and 30 days by analyzing percentage weight loss and observing surface morphology using scanning electron microscopy (SEM). Results indicated that the highest degradation occurred in the Lip-Lac system with 0.5 cm particles at 100% enzyme concentration, yielding a weight loss of 23.81% after 30 days, thereby suggesting that the blend performed better than the single enzyme system. SEM analysis confirmed extensive surface erosion and cracking in smaller particles treated at higher enzyme concentrations. This study also demonstrated that both enzyme concentration and LDPE particle size significantly influence biodegradation efficiency. Taken together, the bifunctional enzyme system is an efficient treatment method for enhancing the degradation process of plastics such as LDPE.

Keywords

LDPE plastic / Enzymatic degradation / Aspergillus flavus / Lipase / Laccase

Funding

None.

Conflict of interest

The authors declare that they have no competing interests.

References

[1] Desidery L, Lanotte M. Polymers and plastics:Types, properties, and manufacturing. In: Giustozzi F, Nizamuddin S,editors. Plastic Waste for Sustainable Asphalt Roads. Woodhead Publishing Series in Civil and Structural Engineering. Delhi: Woodhead Publishing; 2022. p. 3-28. doi: 10.1016/B978-0-323-85789-5.00001-0

[2] Ho BT, Roberts TK, Lucas S. An overview on biodegradation of polystyrene and modified polystyrene: The microbial approach. Crit Rev Biotechnol. 2018; 38(2):308-20. doi: 10.1080/07388551.2017.1355293

[3] Restrepo-Flórez JM, Bassi A, Thompson MR. Microbial degradation and deterioration of polyethylene - a review. Int Biodeterior Biodegrad. 2014; 88:83-90. doi: 10.1016/j.ibiod.2013.12.014

[4] Loredo-Treviño A, Gutiérrez-Sánchez G, Rodríguez- Herrera R, Aguilar CN. Microbial enzymes involved in polyurethane biodegradation: A review. J Polym Environ Polym Degrad. 2012; 20:258-265. doi: 10.1007/s10924-011-0390-5

[5] Fujisawa M, Hirai H, Nishida T. Degradation of polyethylene and nylon-66 by the laccase-mediator system. J Polym Environ. 2001; 9:103-108. doi: 10.1023/A:1020472426516

[6] Anuar NFSK, Huyop F, Ur-Rehman G, et al. An overview into polyethylene terephthalate (PET) hydrolases and efforts in tailoring enzymes for improved plastic degradation. Int J Mol Sci. 2022; 23(20):12644. doi: 10.3390/ijms232012644

[7] Pires JP, Miranda GM, De Souza GL, et al. Investigation of degradation of polypropylene in soil using an enzymatic additive. Iran Polym J. 2019; 28:1045-1055. doi: 10.1007/s13726-019-00766-8

[8] Khandare SD, Chaudhary DR, Jha B. Isolation and purification of esterase enzyme from marine bacteria associated with biodegradation of polyvinyl chloride (PVC). Biodegradation. 2025; 36(4):1-20. doi: 10.1007/s10532-024-10101-5

[9] Lintsen H, Hollestelle M, Hölsgens R.The Plastics Revolution. How the Netherlands Became a Global Player in Plastics. In: Gales B, Homburg E, Joosten J, Vikas Sonak MA,editors. Dutch Polymer Institute (DPI), the Netherlands Organisation for Scientific Research (NWO) and Eindhoven University of Technology; 2017. p. 1-18. Available from: https://www.polymers.nl/_asset/_public/news/first-instalment-the-plastics-revolution.pdf

[10] Pilapitiya PGCNT, Ratnayake AS. The world of plastic waste: A review. Clean Mater. 2024; 11:100220. doi: 10.1016/j.clema.2024.100220

[11] Ritchie H, Samborska V, Roser M. Our World in Data; 2023. Available from: https://ourworldindata.org/plastic-pollution [Last accessed on 2025 Jul 16.

[12] Das MP, Kumar S. Microbial deterioration of low density polyethylene by Aspergillus and Fusarium sp. Int J ChemTech Res. 2014; 6(1):299-305.

[13] Hurley R, Horton A, Lusher A, Nizzetto L. Plastic waste in the terrestrial environment. In: Letcher TM,editor. Plastic Waste and Recycling: Environmental Impact, Societal Issues, Prevention, and Solutions. United States: Academic Press (AP); 2020. p. 163-93. doi: 10.1016/B978-0-12-817880-5.00007-4

[14] Jambeck JR, Geyer R, Wilcox C, et al. Plastic waste inputs from land into the ocean. Science. 2015; 347(6223):768-771. doi: 10.1126/science.1260352

[15] Law KL, Starr N, Siegler TR, Jambeck JR, Mallos NJ, Leonard GH. The United States’ contribution of plastic waste to land and ocean. Sci Adv. 2020; 6(44):eabd0288. doi: 10.1126/sciadv.abd0288

[16] Palansooriya KN, Shi L, Sarkar B, et al. Effect of LDPE microplastics on chemical properties and microbial communities in soil. Soil Use Manag. 2022; 38(3):1481-192. doi: 10.1111/sum.12808

[17] Gajendiran A, Subramani S, Abraham J. Effect of Aspergillus versicolor strain JASS 1 on low density polyethylene degradation. In: 14th ICSET-2017. IOP Publishing Ltd.; 2017. p. 1-7. doi: 10.1088/1757-899X/263/2/022038

[18] Das MP, Kumar S. Influence of cell surface hydrophobicity in colonization and biofilm formation on LDPE biodegradation. Int J Pharm Pharm Sci. 2013; 5(4):690-694.

[19] Abraham J, Ghosh E, Mukherjee P, Gajendiran A. Microbial degradation of low density polyethylene. Environ Prog Sustain Energy. 2017; 36(1):147-154. doi: 10.1002/ep.12467

[20] Lu X, Zhang J, Zhang C, Han Y. Low-density polyethylene (LDPE) surface with a wettability gradient by tuning its microstructures. Macro Mol Rapid Commun. 2005; 26(8):637-642. doi: 10.1002/marc.200400626

[21] Rungswang W, Narkchamnan K, Petcharat N, Thitisak B, Pathaweeisariyakul T. Primitive structure and its morphology for describing highly branched structure of low-density polyethylene. Polym Bull. 2017; 74:3229-3242. doi: 10.1007/s00289-016-1893-y

[22] Sen SK, Raut S. Microbial degradation of low density polyethylene (LDPE): A review. J Environ Chem Eng. 2015; 3(1):462-473. doi: 10.1016/j.jece.2015.01.003

[23] Anuar SZK, Nordin AH, Husna SMN, et al. Recent advances in recycling and upcycling of hazardous plastic waste: A review. J Environ Manage. 2025; 380:124867. doi: 10.1016/j.jenvman.2025.124867

[24] Ray SS, Singh R, Ganesapillai M, Ahn YH. Plastic pollution management-innovative solutions for plastic waste. In: Surampalli RY, Zhang TC, Al-Hashimi BM, et al., editors. Microplastics in the Environment: Fate, Impacts, Removal, and Management. Hoboken: Wiley Online Library; 2025. doi: 10.1002/9781394251100.ch14

[25] Viljakainen VR, Hug LA. New approaches for the characterization of plastic-associated microbial communities and the discovery of plastic-degrading microorganisms and enzymes. Comput Struct Biotechnol J. 2021; 19:6191-6200. doi: 10.1016/j.csbj.2021.11.023

[26] Bhardwaj H, Gupta R, Tiwari A. Communities of microbial enzymes associated with biodegradation of plastics. J Polym Environ. 2013; 21:575-579. doi: 10.1007/s10924-012-0456-z

[27] Roohi, Bano K, Kuddus M, et al. Microbial enzymatic degradation of biodegradable plastics. Curr Pharm Biotechnol. 2017; 18(5):429-440. doi: 10.2174/1389201018666170523165742

[28] Temporiti MEE, Nicola L, Nielsen E, Tosi S. Fungal enzymes involved in plastics biodegradation. Microorganisms. 2022; 10(6):1180. doi: 10.3390/microorganisms10061180

[29] Danso D, Chow J, Streit WR. Plastics: Environmental and biotechnological perspectives on microbial degradation. Appl Environ Microbiol. 2019; 85(19):1-14. doi: 10.1128/aem.01095-19

[30] Safdar A, Ismail F, Imran M. Biodegradation of synthetic plastics by the extracellular lipase of Aspergillus niger. Environ Adv. 2024; 17:100563. doi: 10.1016/j.envadv.2024.100563

[31] Tournier V, Topham CM, Gilles A, et al. An engineered PET depolymerase to break down and recycle plastic bottles. Nature. 2020; 580:216-219. doi: 10.1038/s41586-020-2149-4

[32] Maity W, Maity S, Bera S, Roy A. Emerging roles of PETase and MHETase in the biodegradation of plastic wastes. Appl Biochem Biotechnol. 2021; 193:2699-2716. doi: 10.1007/s12010-021-03562-4

[33] Buchholz PCF, Feuerriegel G, Zhang H, et al. Plastics degradation by hydrolytic enzymes: The plastics-activeenzymes database-PAZy. Proteins. 2022; 90(7):1443-1456. doi: 10.1002/prot.26325

[34] Huaman SMM, Nicholson JH, Brogan APS. A general route to retooling hydrolytic enzymes toward plastic degradation. Cell Rep Phys Sci. 2024; 5(2):1-13. doi: 10.1016/j.xcrp.2024.101783

[35] Ehrhardt T, Rothenberg G.The hydrogen economy fairytale. Green Chem. 2025; 27:6690-6698. doi: 10.1039/d5gc00946d

[36] Yalwaji B, John-Nwagwu HO, Sogbanmu TO. Plastic pollution in the environment in Nigeria: A rapid systematic review of the sources, distribution, research gaps and policy needs. Sci Afr. 2022; 16:e01220. doi: 10.1016/j.sciaf.2022.e01220

[37] Dumbili E, Henderson L.The challenge of plastic pollution in Nigeria. In: Letcher TM, editor. Plastic Waste and Recycling: Environmental Impact, Societal Issues, Prevention, and Solutions. United States: Academic Press; 2020. p. 569-583. doi: 10.1016/B978-0-12-817880-5.00022-0

[38] Duru RU, Ikpeama EE, Ibekwe JA. Challenges and prospects of plastic waste management in Nigeria. Waste Dispos Sustain Energy. 2019; 1:117-126. doi: 10.1007/s42768-019-00010-2

[39] Olanrewaju OO, Oyebade AD. Environmental Menace of Plastic Waste in Nigeria:Challenges, Policies and Technological Efforts. In: World Environmental Conservation Conference; 2019. p. 322-333. Available from: https://www.researchgate.net/publication/335989265 [Last accessed on 2025 Jun 15].

[40] Aidonojie PA, Afolabi MT, Obieshi E, Adeyemi-Balogun MJ, Wakili SA. Breaking legal and socio-economic challenges to plastic waste regulation in Nigeria: Lessons learned from Singapore. Yustisia. 2024; 13(1):64-88. doi: 10.20961/yustisia.v13i1.78388

[41] Ezeudu OB, Tenebe IT, Ujah C. Status of production, consumption, and end-of-life waste management of plastic and plastic products in Nigeria: Prospects for circular plastics economy. Sustainability. 2024; 16:7900. doi: 10.3390/su16187900

[42] Nyakuma BB, Ivase TJP. Emerging trends in sustainable treatment and valorisation technologies for plastic wastes in Nigeria: A concise review. Remediat Treat. 2021; 40(5):e13660. doi: 10.1002/ep.13660

[43] Kehinde O, Ramonu OJ, Babaremu KO, Justin LD. Plastic wastes: Environmental hazard and instrument for wealth creation in Nigeria. Heliyon. 2020; 6(10):e05131. doi: 10.1016/j.heliyon.2020.e05131

[44] Adekomaya O, Ojo K. Adaptation of plastic waste to energy development in Lagos: An overview assessment. Niger J Technol. 2016; 35(2):778-784. doi: 10.4314/njt.v35i1.12

[45] Raji SA, Abdulkareem SA, Ibrahim AO. Effects of reprocessed polyethylene water sachets (PWS) on strength and permeability of laterized concrete. Centrepoint. 2009; 16:24-36.

[46] Nwaigwe DN, Sulymon NA, Bello T, Amiara CA. An investigation into the properties of concrete containing polyethylene (Pure-water sachet waste). Int J Eng Trends Technol. 2019; 67(8):73-77. doi: 10.14445/22315381/ijett-v67i8p212

[47] Liu F, Zhang N, Shang Y, Yao M, Ding M, Yuan Y. Construction of Yarrowia lipolytica for degradation of low-density polyethylene. Process Saf Environ Prot. 2025; 195:106818. doi: 10.1016/j.psep.2025.106818

[48] Mazaheri H, Nazeri S. Biodegradation and detoxification of low-density polyethylene (LDPE) by Stenotrophomonas sp. and Alcaligenaceae bacterium. Bull Environ Contam Toxicol. 2024; 112:19. doi: 10.1007/s00128-023-03836-z

[49] Khandare SD, Chaudhary DR, Jha B. Marine bacterial biodegradation of low-density polyethylene (LDPE) plastic. Biodegradation. 2021; 32:127-143. doi: 10.1007/s10532-021-09927-0

[50] DSouza GC, Sheriff RS, Ullanat V, et al. Fungal biodegradation of low-density polyethylene using consortium of Aspergillus species under controlled conditions. Heliyon. 2021; 7(5):e07008. doi: 10.1016/j.heliyon.2021.e07008

[51] Kyaw BM, Champakalakshmi R, Sakharkar MK, Lim CS, Sakharkar KR. Biodegradation of low density polythene (LDPE) by Pseudomonas species. Indian J Microbiol. 2012; 52:411-419. doi: 10.1007/s12088-012-0250-6

[52] Acharya T, Hare J.Sabouraud Agar and other fungal growth media. In: Gupta VK, Tuohy M, editors. Laboratory Protocols in Fungal Biology. Cham: Springer; 2022. p. 69-86. doi: 10.1007/978-3-030-83749-5_2

[53] Khan S, Ali SA, Ali AS. Biodegradation of low density polyethylene (LDPE) by mesophilic fungus “Penicillium citrinum” isolated from soils of plastic waste dump yard, Bhopal, India. Environ Technol. 2022; 44:2300-2314. doi: 10.1080/09593330.2022.2027025

[54] Nouroozi RV, Noroozi MV, Ahmadizadeh M. Determination of protein concentration using bradford microplate protein quantification assay. Int Electron J Med. 2015; 4(1):11-17. doi: 10.31661/iejm158

[55] Ren L, Men L, Zhang Z, et al. Biodegradation of polyethylene by Enterobacter sp. D1 from the guts of wax moth Galleria mellonella. Int J Environ Res Public Health. 2019; 16(11):1941. doi: 10.3390/ijerph16111941

[56] Yang WK, Gong Z, Wang BT, et al. Biodegradation of low-density polyethylene by mixed fungi composed of Alternaria sp. and Trametes sp. isolated from landfill sites. BMC Microbiol. 2024; 24:321. doi: 10.1186/s12866-024-03477-0

[57] Zahra S, Abbas SS, Mahsa MT, Mohsen N. Biodegradation of low-density polyethylene (LDPE) by isolated fungi in solid waste medium. Waste Manag. 2010; 30(3):396-401. doi: 10.1016/j.wasman.2009.09.027

[58] Kunlere IO, Fagade OE, Nwadike BI. Biodegradation of low density polyethylene (LDPE) by certain indigenous bacteria and fungi. Int J Environ Stud. 2019; 76(3):428-40. doi: 10.1080/00207233.2019.1579586

[59] Yao Z, Seong HJ, Jang YS. Degradation of low density polyethylene by Bacillus species. Appl Biol Chem. 2022; 65:84. doi: 10.1186/s13765-022-00753-3

[60] Harrat R, Bourzama G, Burgaud G, Coton E, Bourezgui A, Soumati B. Assessing the biodegradation of low-density polyethylene films by Candida tropicalis SLNEA04 and Rhodotorula mucilaginosa SLNEA05. Diversity. 2024; 16(12):759. doi: 10.3390/d16120759

[61] Awasthi S, Srivastava N, Singh T, Tiwary D, Mishra PK.Biodegradation of thermally treated low density polyethylene by fungus Rhizopus oryzae NS 5. 3 Biotech. 2017; 7:73. doi: 10.1007/S13205-017-0699-4

[62] Gong Z, Jin L, Yu X, et al. Biodegradation of low density polyethylene by the fungus Cladosporium sp. recovered from a landfill site. J Fungi. 2023; 9(6):605. doi: 10.3390/jof9060605

[63] Samanta S, Datta D, Halder G. Biodegradation efficacy of soil inherent novel sp. Bacillus tropicus (MK318648) onto low density polyethylene matrix. J Polym Res. 2020; 27:327. doi: 10.1007/s10965-020-02296-x

[64] Azevedo HS, Reis RL. Understanding the enzymatic degradation of biodegradable polymers and strategies to control their degradation rate. In: Reis RL, Román JS,editors. Biodegradable Systems in Tissue Engineering and Regenerative Medicine. 1st ed. Boca Raton: CRC Press LLC.; 2005. p. 177-201. Available from: https://repositorium.uminho.pt/bitstream/1822/14150/1/file.pdf [Last accessed on 2025 Jun 13].

[65] Tao X, Ouyang H, Zhou A, et al. Polyethylene degradation by a Rhodococcous strain isolated from naturally weathered plastic waste enrichment. Environ Sci Technol. 2023; 57(37):13901-13911. doi: 10.1021/acs.est.3c03778

[66] Jun Y, Xiangyan W, Yu Y, Yixiao Y. Method of Degrading Polyethylene by Extracellular Laccase of Bacillus. Patents no:CN201410225175.2A; 2017. Available from: https://patents.google.com/patent/cn103980535a/en [Last accessed on 2025 Jun 14].

[67] Das MP, Kumar S. An approach to low-density polyethylene biodegradation by Bacillus amyloliquefaciens. 3 Biotech. 2015; 5:81-86. doi: 10.1007/s13205-014-0205-1

[68] Mumtaz T, Khan MR, Hassan MA. Study of environmental biodegradation of LDPE films in soil using optical and scanning electron microscopy. Micron. 2010; 41(5):430-438. doi: 10.1016/j.micron.2010.02.008

[69] González-Márquez A, Andrade-Alvarado AD, González- Mota R, Sánchez C. Enhanced degradation of phototreated recycled and unused low-density polyethylene films by Pleurotus ostreatus. World J Microbiol Biotechnol. 2024; 40:309. doi: 10.1007/s11274-024-04116-6

[70] Dey AS, Bose H, Mohapatra B, Sar P. Biodegradation of unpretreated low-density polyethylene (LDPE) by Stenotrophomonas sp. and Achromobacter sp., isolated from waste dumpsite and drilling fluid. Front Microbiol. 2020; 11:603210. doi: 10.3389/fmicb.2020.603210

[71] Yao C, Xia W, Dou M, Du Y, Wu J. Oxidative degradation of UV-irradiated polyethylene by laccase-mediator system. J Hazard Mater. 2022; 440:129709. doi: 10.1016/j.jhazmat.2022.129709

[72] Sowmya HV, Ramalingappa B, Krishnappa M, Thippeswamy B. Degradation of polyethylene by Trichoderma harzianum-SEM, FTIR, and NMR analyses. Environ Monit Assess. 2014; 186(10):6577-6586. doi: 10.1007/s10661-014-3875-6

[73] Sanniyasi E, Gopal RK, Gunasekar DK, Raj PP. Biodegradation of low-density polyethylene (LDPE) sheet by microalga, Uronema africanum Borge. Sci Rep. 2021; 11:17233. doi: 10.1038/s41598-021-96315-6

[74] Jones DL, Nguyen C, Finlay RD. Carbon flow in the rhizosphere: Carbon trading at the soil-root interface. Plant Soil. 2009; 321:5-33. doi: 10.1007/s11104-009-9925-0

[75] Azeko ST, Etuk-Udo GA, Odusanya OS, Malatesta K, Anuku N, Soboyejo WO. Biodegradation of linear low density polyethylene by Serratia marcescens subsp. marcescens and its cell free extracts. Waste Biomass Valorization. 2015; 6(6):1047-10457. doi: 10.1007/s12649-015-9421-0