Strain capable of degrading polypropylene plastic, microbial inoculant and application

Abstract

The present application relates to the technical field of microorganisms, and particularly relates to a strain capable of degrading polypropylene plastic, a microbial inoculum and application Pseudomonas aeruginosa Strain PP01, preserved in the Guangdong Microbial Culture Collection Center on May 26, 2025, with a preservation number of GDMCC No: 66399. The strain has high degradation efficiency, and the degradation rate of the strain on polypropylene plastic reaches 15.71% in 20 days of culture time, i.e. 0.79% per day. Moreover, the strain is suitable for a wide range of environments. Pseudomonas aeruginosa Strain PP01 can be cultured in aerobic and anoxic environments, and can grow in an inorganic salt culture medium with polypropylene plastic as the only carbon source, which has advantages in practical application.

Description

Technical Field

[0001] This invention relates to the field of microbial technology, specifically to a strain of microorganisms, a microbial agent, and its application that can degrade polypropylene plastic. Background Technology

[0002] Polypropylene (PP) has consistently been the most produced thermoplastic polymer, accounting for 17% of global plastic production. PP possesses properties such as chemical resistance, elasticity, toughness, fatigue resistance, insulation, and light transmittance. Due to these advantages, PP is widely used in the packaging industry, electrical equipment manufacturing, home appliances, the automotive industry, and building materials. PP takes approximately 450 years to decompose naturally. Like other types of plastics, its primary fate is to end up in landfills or incinerate. In the former case, due to its persistence in the environment, it may undergo fragmentation, becoming a source of microplastics and having several related consequences for human health, such as inflammation, genotoxicity, and apoptosis. In the latter case, PP releases hazardous pollutants such as dioxins when incinerated. Traditional plastic disposal methods are difficult to guarantee effectiveness, while biodegradation holds promise as a key pathway.

[0003] Previous researchers have isolated various bacterial strains capable of degrading polypropylene (PP) from the environment. For example, *Pseudomonas aeruginosa* (PP) isolated from topsoil contaminated with wastewater... Pseudomonas aeruginosa After one month of incubation in nutrient broth medium, the weight loss of PP was 5.37%, or 0.18% / day. Two bacterial strains isolated from mangrove sediments belong to Bacillus subtilis (…). Bacillus subtilis ) and Rhodococcus ( Rhodococcus ruber After 40 days of incubation, the degradation efficiencies of PP were 4.00% (0.10% / d) and 6.40% (0.16% / d), respectively. Lysine-containing Bacillus bacteria isolated from soil trenches degraded 4.00% (0.15% / d) of PP within 26 days. An oligotrophic monotypic bacterium was isolated and identified from a municipal solid waste dump. Stenotrophomonas panacihumi PA3-2, after a 90-day incubation period at 37°C, degraded 20.30% (0.23% / day) and 12.70% (0.14% / day) of low-molecular-weight and high-molecular-weight polypropylene, respectively. Two Bacillus subspecies of Bacillus thuringiensis, P6 and P8, isolated from compost, showed a PP decomposition efficiency of 10.00% (0.67% / day) after 15 days of incubation. Antarctic soil bacteria - Pseudomonas sp. ADL15 and RhodococcusAfter 40 days of incubation with sp. ADL36 and PP microplastics, the percentage weight loss of PP microplastics was 17.30% (0.43% / day) and 7.30% (0.18% / day), respectively. Using PP as the sole carbon source, two Pseudomonas aeruginosa strains, WZH-4 and WGH-6, were isolated, causing PP mass losses of 9.35% (0.23% / day) and 17.20% (0.43% / day), respectively, within 40 days.

[0004] However, these isolated strains capable of degrading PP suffer from low degradation efficiency. Summary of the Invention

[0005] To address the above problems, this invention provides a strain of microorganisms, a microbial agent, and its application that can degrade polypropylene plastic.

[0006] This invention is achieved through the following technical solution:

[0007] A strain capable of degrading polypropylene plastic, said strain is Pseudomonas aeruginosa StrainPP01 was deposited at the Guangdong Provincial Center for Microbial Culture Collection on May 26, 2025, with accession number GDMCC No: 66399.

[0008] A microbial inoculant, the preparation method of which is as follows:

[0009] The strain was inoculated into a culture medium suitable for bacteria and cultured, and the bacterial cells were collected by centrifugation.

[0010] The collected bacterial cells are mixed with a carrier to prepare a microbial inoculum; the mass ratio of bacterial cells to carrier is 1:4~6.

[0011] Preferably, the number of live bacteria in the microbial agent is 10. 8 CFU~10 10 CFU.

[0012] Preferably, the culture temperature of the strain is 25℃~30℃, the rotation speed is 180rpm~200rpm, and the culture time is 20d.

[0013] Preferably, the carrier is straw powder or biochar; the particle size of the straw powder is 30μm~75μm; the particle size of the biochar is 30μm~75μm.

[0014] Preferably, the culture medium is LB medium, or the culture medium formulation is: 1000 mL deionized water, 5.1 g to 5.3 g K₂HPO₄, 3.6 g to 3.8 g KH₂PO₄, 0.9 g to 1.1 g Na₂SO₄, 0.1 g to 0.3 g MgSO₄·7H₂O, 1.9 g to 2.1 g NH₄Cl, 1 mL metal ion solution, and 6 g to 10 g polypropylene plastic powder; wherein the metal ion solution formulation is: 1000 mL L of deionized water, 0.2g~0.4g of FeCl2·4H2O, 0.037g~0.039g of CoCl2·6H2O, 0.01g~0.03g of MnCl2·4H2O, 0.013g~0.015g of ZnCl2, 0.0123g~0.0125g of H3BO3, 0.03g~0.05g of Na2MoO4·2H2O, and 0.0033g~0.0035g of CuCl2·2H2O.

[0015] Preferably, the particle size of the polypropylene plastic powder is 87μm~89μm.

[0016] The application of the aforementioned microbial agent in the degradation of polypropylene plastics.

[0017] Preferably, the microbial agent is sprayed onto the polypropylene plastic waste at a rate of 1 kg to 1.5 kg of microbial agent per cubic meter of polypropylene plastic waste.

[0018] Compared with the prior art, the present invention has the following beneficial effects:

[0019] This application provides a strain capable of degrading polypropylene plastics. This strain exhibits high degradation efficiency, achieving a degradation rate of 15.71% for polypropylene plastics during a 20-day culture period, or 0.79% degradation per day. Furthermore, this strain is adaptable to a wide range of environments. Pseudomonas aeruginosa Strain PP01 can be cultured in both aerobic and anaerobic environments. It can grow in inorganic salt culture medium with polypropylene plastic as the sole carbon source, indicating that the strain has a complete polypropylene metabolic pathway. It can complete the degradation and transformation of polypropylene without the need for additional organic nutrients containing carbon sources. This not only significantly reduces the application cost, but also shows significant application potential in complex environments, especially in nutrient-deficient contaminated sites. Therefore, it has advantages in practical applications.

[0020] Furthermore, this strain has the potential for expanded and in-depth applications. Isolated from the intestines of the white-spotted flower beetle larvae, which can feed on polypropylene plastic, this strain can be utilized... Pseudomonas aeruginosaThe dual degradation process of StrainPP01 and its host makes the degradation of polypropylene plastic more efficient. First, the mechanical force applied by the larvae's chewing breaks the plastic into smaller fragments, transforming it into small particles with a chemically modified surface. Then, the degradation process is further enhanced by... Pseudomonas aeruginosa In vitro fermentation of Strain PP01 degrades polypropylene plastic.

[0021] Information on the preservation of biological materials

[0022] Pseudomonas aeruginosa ( Pseudomonas aeruginosa Named Pseudomonas aeruginosa Strain PP01, taxonomic name is Pseudomonas aeruginosa, It was deposited on May 26, 2025 at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No: 66399, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou. Attached Figure Description

[0023] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0024] Figure 1 The image shows the result of the invention that the white-spotted beetle can feed solely on polypropylene plastic; Figure 1 In the figures, A represents polypropylene plastic without 3rd instar larvae of the white-spotted flower beetle, left to stand for 3 days; B represents polypropylene plastic with 3rd instar larvae of the white-spotted flower beetle placed inside and reared for 3 days.

[0025] Figure 2 This invention describes a single colony screened from the intestines of the white-spotted flower beetle. Pseudomonas aeruginosa Results of Strain PP01 degrading polypropylene plastic; Figure 2 In the image, A shows a control group of polypropylene plastic; B shows a polypropylene plastic with a single colony added. Pseudomonas aeruginosa A display image of Strain PP01 degradable polypropylene plastic.

[0026] Figure 3 This is a phylogenetic analysis result diagram of the present invention; the maximum likelihood tree sequence of the 16S rRNA comes from B32 ( Pseudomonas aeruginosa Strain PP01) P. aeruginosa S2QPS8 (HQ844502.1, Pseudomonas aeruginosa strain S2QPS8), P. aeruginosaALK318 (KC456533.1, Pseudomonas aeruginosa strain ALK318), P. aeruginosa 9 (MN911373.1, Pseudomonas aeruginosa strain 9) P. aeruginosa SSRP6 (OR352454.1, Pseudomonas aeruginosa strain SSRP6), P. aeruginosa ALK320 (KC456535.1, Pseudomonas aeruginosa strain ALK320), B. subtilis s3e(JX274662.1, Bacillus subtilis strains3e), A. refrigerator WB1(NR_178633.1, Acinetobacter refrigeratoris (strainWB1), the numbers on the branches of the phylogenetic tree are expansion values, and the scale bar represents the differences in the base sequence. Detailed Implementation

[0027] To facilitate understanding of the present invention, a more comprehensive description is provided below, along with preferred embodiments. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein. Rather, these embodiments are provided to provide a thorough and complete understanding of the disclosure of the present invention.

[0028] Unless otherwise defined, all technical and scientific terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. The terminology used in this invention and in its specification is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention.

[0029] The beneficial effects of the present invention will be illustrated below through specific embodiments.

[0030] The polypropylene plastic powder used in this invention was purchased from Maclean, with a molecular weight of 42.08M and a particle size of 87μm~89μm. The straw powder was wheat straw powder, purchased from Taobao's Carbon Road, with a particle size of 30μm~75μm; the biochar was straw biochar, purchased from Taobao's Carbon Road, with a particle size of 30μm~75μm.

[0031] Example 1

[0032] Pseudomonas aeruginosa ( Pseudomonas aeruginosa Screening of Strain PP01.

[0033] The third instar larvae of the white-spotted scarab beetle were obtained from the Institute of Pig Breeding and Animal Nutrition, Yunnan Academy of Animal Science and Veterinary Medicine. In terms of artificial rearing, the third instar larvae were first placed in an artificial rearing glass bottle, 9.4 cm high and 6.8 cm in diameter, containing a 6 cm thick layer of polypropylene plastic powder. The bottle was then placed in an artificial climate chamber with environmental conditions set as a dark environment, 28°C, and 40% relative humidity. Figure 1 As shown. After 3 days of feeding, the worms were anesthetized on an ice plate. To remove surface bacterial contamination, the worms were sterilized with 75% ethanol for 90 seconds, then with 1% sodium hypochlorite for 1 minute, and finally washed five times with sterile double-distilled water. Under aseptic conditions, the entire intestine was dissected and placed in a sterile mortar. Sterile water was added, and the intestine was lightly ground with a sterile grinding rod. All the broken tissue was added to 40 mL of LB liquid medium and cultured at 30°C and 200 rpm for 1 day to obtain the enriched total bacterial solution. The total bacterial solution was transferred at a ratio of 1 / 10 to LB liquid medium containing 10 g / L polypropylene and acclimated at 30°C and 200 rpm for 1 day. The pre-concentrated bacterial solution was added at a ratio of 1 / 10 to inorganic salt liquid medium with 10 g / L polypropylene powder as the sole carbon source and acclimated at 30°C and 200 rpm for 2 days. Take 1 mL of the above bacterial suspension and spread it onto an inorganic salt solid medium containing 10 g / L polypropylene powder as the sole carbon source. Incubate at 30°C upside down for 1 day. Pick a single colony with a sterile inoculation loop and streak it onto an inorganic salt solid medium containing polypropylene powder using the continuous streak method. Incubate at 30°C upside down for 1 day. Pick a single colony with a 10 μL sterile pipette tip and place it into an inorganic salt liquid medium containing 10 g / L polypropylene powder as the sole carbon source. Incubate at 30°C and 200 rpm for 20 days. Figure 2 As shown.

[0034] Example 2

[0035] Pseudomonas aeruginosa ( Pseudomonas aeruginosa Identification of Strain PP01.

[0036] To determine the genus of this strain, its 16S rRNA sequence was amplified using primer 27F (nucleotide sequence shown in SEQ ID NO. 1: 5ʹ-AGAGTTTGATCMTGGCTCAG-3ʹ) and primer 1492R (nucleotide sequence shown in SEQ ID NO. 2: 5ʹ-TACGGYTACCTTGTTACGACTT-3ʹ). Sequencing was performed after PCR amplification, and a phylogenetic tree was constructed. The results are shown below. Figure 3 As shown, the results indicate that the newly isolated strain is *Pseudomonas aeruginosa*, named... Pseudomonas aeruginosa Strain PP01.

[0037] Example 3

[0038] Pseudomonas aeruginosa ( Pseudomonas aeruginosa Determination of the degradation efficiency of PP by Strain PP01.

[0039] 100 μL of bacterial culture was added to an inorganic salt liquid culture medium with 10 g / L polypropylene powder as the sole carbon source and cultured at 30 °C and 200 rpm for 20 days, with three biological replicates. The degradation efficiency of the polypropylene plastic was then determined. The specific steps were as follows: After culture, the bacterial culture was passed through a Bioharp BS-40-CS cell filter with a pore size of 40 μm to collect the plastic, which was then treated overnight with 2% sodium dodecyl sulfate (SDS) to remove bacteria adhering to the plastic surface. The plastic was then thoroughly washed twice with deionized water and dried in an oven at 60 °C for 1 day. The weight loss of the plastic was then measured. The formula for the plastic degradation efficiency is as follows:

[0040] Plastic degradation efficiency = 100% × (initial weight - remaining weight) / initial weight × 100%.

[0041] The results showed that under conditions of 30℃ and 200rpm, single colony- Pseudomonas aeruginosa StrainPP01 can be grown in an inorganic salt liquid medium with 10 g / L polypropylene powder as the sole carbon source, and its degradation efficiency reaches 0.79% / d.

[0042] Example 4

[0043] Pseudomonas aeruginosa ( Pseudomonas aeruginosa Practical applications of Strain PP01 in the degradation of polypropylene plastics.

[0044] In municipal solid waste landfills, areas rich in plastic waste, such as polypropylene plastic packaging material accumulation zones, are selected to construct in-situ biodegradation systems. The specific steps are as follows:

[0045] Preparation of inoculum:

[0046] Filter Pseudomonas aeruginosa The Strain PP01 strain was cultured in a medium at 30°C and 200 rpm for 48 hours, and the cells were collected by centrifugation.

[0047] The culture medium is formulated as follows: 1000 mL of deionized water, 5.2 g of K2HPO4, 3.7 g of KH2PO4, 1.0 g of Na2SO4, 0.2 g of MgSO4·7H2O, 2.0 g of NH4Cl, 1 mL of metal ion solution and 10 g of polypropylene plastic powder.

[0048] The metal ion solution formulation is as follows: 1000 mL of deionized water, 0.3 g of FeCl2·4H2O, 0.038 g of CoCl2·6H2O, 0.02 g of MnCl2·4H2O, 0.014 g of ZnCl2, 0.0124 g of H3BO3, 0.04 g of Na2MoO4·2H2O, and 0.0034 g of CuCl2·2H2O.

[0049] The polypropylene plastic powder has a particle size of 88 μm.

[0050] The bacterial cells and carrier were mixed at a mass ratio of 1:5 to prepare a solid microbial inoculum with a viable count of 10-1. 8 CFU. The carrier is wheat straw powder with a particle size of 45 μm.

[0051] Example 5

[0052] Pseudomonas aeruginosa ( Pseudomonas aeruginosa Practical applications of Strain PP01 in the degradation of polypropylene plastics.

[0053] In municipal solid waste landfills, areas rich in plastic waste, such as polypropylene plastic packaging material accumulation zones, are selected to construct in-situ biodegradation systems. The specific steps are as follows:

[0054] Preparation of inoculum:

[0055] Filter Pseudomonas aeruginosa The Strain PP01 strain was cultured in a medium at 25°C and 200 rpm for 48 hours, and the cells were collected by centrifugation.

[0056] The culture medium is formulated as follows: 1000 mL of deionized water, 5.1 g of K2HPO4, 3.6 g of KH2PO4, 0.9 g of Na2SO4, 0.1 g of MgSO4·7H2O, 1.9 g of NH4Cl, 1 mL of metal ion solution and 6 g of polypropylene plastic powder.

[0057] The metal ion solution formulation is as follows: 1000 mL of deionized water, 0.2 g of FeCl2·4H2O, 0.037 g of CoCl2·6H2O, 0.01 g of MnCl2·4H2O, 0.013 g of ZnCl2, 0.0123 g of H3BO3, 0.03 g of Na2MoO4·2H2O, and 0.0033 g of CuCl2·2H2O.

[0058] The particle size of the polypropylene plastic powder is 87 μm.

[0059] The bacterial cells and carrier were mixed at a mass ratio of 1:4 to prepare a solid microbial inoculum with a viable count of 10-1. 9 CFU. The carrier is straw biochar with a particle size of 45 μm.

[0060] Example 6

[0061] Pseudomonas aeruginosa ( Pseudomonas aeruginosa Practical applications of Strain PP01 in the degradation of polypropylene plastics.

[0062] In municipal solid waste landfills, areas rich in plastic waste, such as polypropylene plastic packaging material accumulation zones, are selected to construct in-situ biodegradation systems. The specific steps are as follows:

[0063] Preparation of inoculum:

[0064] Filter Pseudomonas aeruginosa The Strain PP01 strain was cultured in a medium at 30°C and 200 rpm for 48 hours, and the cells were collected by centrifugation.

[0065] The culture medium is formulated as follows: 1000 mL of deionized water, 5.3 g of K2HPO4, 3.8 g of KH2PO4, 1.1 g of Na2SO4, 0.3 g of MgSO4·7H2O, 2.1 g of NH4Cl, 1 mL of metal ion solution and 8 g of polypropylene plastic powder.

[0066] The metal ion solution formulation is as follows: 1000 mL of deionized water, 0.4 g of FeCl2·4H2O, 0.039 g of CoCl2·6H2O, 0.03 g of MnCl2·4H2O, 0.015 g of ZnCl2, 0.0125 g of H3BO3, 0.05 g of Na2MoO4·2H2O, and 0.0035 g of CuCl2·2H2O.

[0067] The particle size of the polypropylene plastic powder is 89 μm.

[0068] The bacterial cells and carrier were mixed at a mass ratio of 1:6 to prepare a solid microbial inoculum with a viable count of 10-1. 10 CFU. The carrier is wheat straw powder with a particle size of 30 μm.

[0069] Example 7

[0070] The wheat straw powder has a particle size of 75 μm, and the remaining steps are exactly the same as in Example 4.

[0071] Example 8

[0072] The straw biochar particle size was 30 μm, and the remaining steps were exactly the same as in Example 5.

[0073] Example 9

[0074] The straw biochar particle size was 75 μm, and the remaining steps were exactly the same as in Example 5.

[0075] Example 10

[0076] Filter Pseudomonas aeruginosa The Strain PP01 strain was cultured on LB medium, and the remaining steps were exactly the same as in Example 4.

[0077] Example 11

[0078] Filter Pseudomonas aeruginosa The Strain PP01 strain was cultured on LB medium, and the remaining steps were exactly the same as in Example 5.

[0079] Example 12

[0080] Filter Pseudomonas aeruginosa The Strain PP01 strain was cultured on LB medium, and the remaining steps were exactly the same as in Example 6.

[0081] On-site handling:

[0082] In the plastic-rich area of ​​the landfill, a degradation trench with a depth of 1m and a width of 2m was excavated, and a breathable, impermeable membrane was laid at the bottom of the trench. Polypropylene plastic waste, such as plastic bags and packaging films, was broken into 5×5cm fragments and spread evenly in the trench, with each layer approximately 20cm thick. The microbial agent prepared in Example 4 was sprinkled between layers at a rate of 1kg of microbial agent / m³. 3 Polypropylene plastic waste. Cover with 30cm thick humus soil to retain heat and moisture, and install temperature and humidity sensors to maintain 30℃ and 60% moisture content.

[0083] Five PP samples (10g each) were randomly selected from the trench every 30 days. After cleaning and drying according to the method in Example 3, they were weighed, and the cumulative weight loss rate was calculated. After 180 days of treatment, the average weight loss rate of polypropylene plastic waste reached 12.6%, equivalent to 0.07% / day, with obvious pores and cracks appearing on the surface. The COD and microplastic content in the landfill leachate were reduced by more than 35% compared to the untreated area, indicating that the microbial agent can synergistically reduce secondary pollution.

[0084] It should be noted that the average weight loss rate of polypropylene plastic waste treated on-site using the microbial agents prepared in Examples 5 to 12 was between 12.4% and 12.8%, with obvious pores and cracks appearing on the surface. The chemical oxygen demand (COD) and microplastic content in the landfill leachate were reduced by more than 35% compared to the untreated area, indicating that the microbial agents can synergistically reduce secondary pollution. Applying 1 kg to 1.5 kg of microbial agent per cubic meter of polypropylene plastic waste was sufficient to degrade the polypropylene plastic waste.

[0085] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0086] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are specific and detailed, they should not be construed as limiting the scope of the invention. Those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these all fall within the scope of protection of the present invention. Therefore, the scope of protection of this invention should be determined by the appended claims.

Claims

1. The application of a bacterial strain and its inoculant in the degradation of polypropylene plastics, characterized in that, The strain is *Pseudomonas aeruginosa* (… Pseudomonas aeruginosa strain PP01 was deposited at the Guangdong Provincial Center for Microbial Culture Collection on May 26, 2025, with accession number GDMCC No: 66399; The preparation method of the bacterial agent is as follows: The strain was inoculated into a culture medium suitable for bacteria and cultured, and the bacterial cells were collected by centrifugation; The collected bacterial cells are mixed with a carrier to prepare a bacterial agent; the mass ratio of bacterial cells to carrier is 1:4~6.

2. The application as described in claim 1, characterized in that, The number of live bacteria in the bacterial agent is 10. 8 CFU~10 10 CFU.

3. The application as described in claim 1, characterized in that, The strain was cultured at a temperature of 25℃~30℃, a rotation speed of 180rpm~200rpm, and a culture time of 20 days.

4. The application as described in claim 1, characterized in that, The carrier is straw powder or biochar; the particle size of the straw powder is 30μm~75μm; the particle size of the biochar is 30μm~75μm.

5. The application as described in claim 1, characterized in that, The culture medium is LB medium, or the culture medium formula is: 1000 mL deionized water, 5.1 g to 5.3 g K₂HPO₄, 3.6 g to 3.8 g KH₂PO₄, 0.9 g to 1.1 g Na₂SO₄, 0.1 g to 0.3 g MgSO₄·7H₂O, 1.9 g to 2.1 g NH₄Cl, 1 mL metal ion solution, and 6 g to 10 g polypropylene plastic powder; the metal ion solution formula is: 1000 mL... Deionized water, 0.2g~0.4g FeCl2·4H2O, 0.037g~0.039g CoCl2·6H2O, 0.01g~0.03g MnCl2·4H2O, 0.013g~0.015g ZnCl2, 0.0123g~0.0125g H3BO3, 0.03g~0.05g Na2MoO4·2H2O, and 0.0033g~0.0035g CuCl2·2H2O.

6. The application as described in claim 5, characterized in that, The particle size of the polypropylene plastic powder is 87μm~89μm.

7. The application as described in claim 1, characterized in that, The microbial agent is sprayed onto polypropylene plastic waste at a rate of 1 kg to 1.5 kg of microbial agent per cubic meter of polypropylene plastic waste.

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