Venisse Abanilla Caldwell University, Class of 2022 Major: Health Science

Sudeep Khadka Caldwell University, Class of 2021 Major: Biology

Faculty Darryl Aucoin, Ph.D., Assistant Professor of Chemistry

Advisor: Department of Natural Sciences Faculty Agnes T. Berki, Ph.D., Associate Professor

Advisor: Department of Natural Sciences

Plastic is an inspiring innovation that has helped us to make our life better, safer, and easier. Plastics are used

worldwide. They are in cell phones, household products, televisions, computers, and all electronic products that have

made modern life possible. The maximum use of plastics without proper management techniques is leading to the

accumulation of plastic debris in the environment, resulting in plastic pollution. Several preventive measures, like

recycling, regulation of plastic usage and burning are taken in order to reduce plastic pollution. However, these

measures require extensive economic resources and utilize considerable amounts of energy. In order to increase

efficiency and promote sustainable energy, a novel approach has been studied to utilize microorganisms to reduce

the population of plastic via biodegradation. Biodegradation of plastic requires relatively less economic resources and

has minimal environmental side effects, if any. We proposed to study the varied environmental conditions that

optimize the growth of Rhodococcus ruber, a polyethylene plastic digesting bacterium. To achieve this goal, the

bacterium was sequentially subjected to varied temperatures, pH, and media type. The efficiency at which the plastic

was degraded was indicated by the weight loss, and depicted how well the bacteria flourished in varied temperatures

and pH. In the first test, the plastic tubes with bacteria were treated at three different temperatures, 26°C, 30°C, and

34°C. There was a significant decrease in the weight of plastic tubes at 30°C. Secondly, the bacteria were grown in

three different pH, 6.5, 7.8 and 8.4. The results show the most significant decrease in the weight of plastic tubes was

at a pH of 7.8. These results suggest 30°C and pH of 7.8 as the optimal temperature and pH for this bacterium. Finally,

we measured various mixtures of two separate media, nutrient broth and synthetic medium, to consider which

mixture would optimize the growth of the bacterium. The bacterium grew best in the nutrient broth and in the 2:3

and 4:1 mixture of nutrient broth and synthetic media. These significant results can be used for a mass biodegradation

of polyethylene by R. ruber at 30°C, pH of 7.8, and in three different media to reduce plastic pollution.

PROJECT

1

Biodegradation of Polyethylene by Rhodoccocus ruber:

Optimizing Growth ConditionsVenisse Abanilla,Sudeep Khadka, Dr. Agnes Berki, Dr. Darryl Aucoin

Independent Scientific Research Project

Caldwell University

INTRODUCTION

• Polyethylene plastic & Polystyrene plastic

• Biodegradation

• Rhodoccocus ruber

PURPOSE

To determine the best growth conditions of R. ruber for polyethylene and polystyrene

degradation:

pH, Temperature, and Medium composition

METHODS

• Preparation of media and compositions

• Sterile culture of bacteria

• Temperatures, 26°C, 30°C, 34°C

• pH, 6.5, 7.8 and 8.4

• Measure degradation

Figure 5. Nutrient media B1, compositions B2-B9, and synthetic media B10

Figure 6. Polystyrene tubes after vacuum oven

BIODEGRADATION OF POLYETHYLENE BY RHODOCCOCUS RUBER:

OPTIMIZING GROWTH CONDITIONS

Venisse Abanilla, Sudeep Khadka and Darryl S. Aucoin, Ph.D, Agnes T. Berki, Ph.D.

Department of Natural and Physical Sciences, Caldwell University, Caldwell, New Jersey 07006

POLYETHYLENE PLASTIC

• Most popular plastic

• Water bottles, grocery bags, electronic

products

• Current solution for plastic pollution:

recycling

BIODEGRADATION

• Decomposition of organic material using

microorganisms

• Bacteria and fungi

BACTERIA

• Rhodoccocus ruber

Figure 1

• Abundant in soil,

water, marine

environments

• Degrades

polyethylene plastic

To determine the best growth conditions of R.

ruber for polyethylene and polystyrene

degradation

• pH

• Temperature

• Medium composition

INTRODUCTION

PURPOSE

RESULTS

• The best growth of R. ruber and

maximum degradation of plastic was

at the temperature of 30°C• The best growth of R. ruber and

maximum degradation of plastic was

at the pH of 7.8

• The best media composition for

growth of R. ruber was the 80%

nutrient media

CONCLUSIONS

MATERIALS

• Rhodoccocus ruber

• Polystyrene tubes

• Polyethylene plastic

• Nutrient Media (BBL® Nutrient Broth)

• Synthetic Media • 26°C, 30°C, 34°C Incubator

• Vacuum oven

• Preparation of nutrient media, synthetic media, and compositions

Figure 5.

• Sterile culture of bacteria

• Grow bacteria in all media compositions at three different temperatures, 26°C, 30°C, 34°C in polystyrene tubes with

polyethylene plastic pieces

• Grow bacteria in B4 medium at three different pH, 6.5, 7.8

and 8.4 in polystyrene tubes with polyethylene plastic pieces

• Measure degradation by weight loss of plastics using vacuum

oven and scale Figure 6.

ACKNOWLEDGEMENTS

Lois Ayim

Chennelle Lawrence

Christabel Osei-Du

Caldwell University

Independent College Fund of New Jersey

• Treat R. ruber with different plastics other

than polyethylene and polystyrene

• Test in lower pH and temperature

• More precise measurements

• Increase sample size

METHODS

FURTHER RESEARCH

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PE #

PE in Incubation Temperature

26°C

30°C

34°C

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PS Tube #

Media pH in PS Tubes

pH 6.5

pH 7.8

pH8.4

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Incubation Temperature in PS Tubes

26°C

30°C

34°C

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PE in Media pH

pH 6.5

pH7.8

pH8.4

0.108

0.025

0.048

0.042

0.086

0.064

0.082

0.1

0.126

0.093

0

0.02

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0 10 20 30 40 50 60 70 80 90 100

Ab

so

rba

nce

at 6

00

nm

% BBL Nutrient Broth in Synthetic Media

Absorbance Curve of R. ruber

0.004

0.187

0.2570.263

0.302

0.349

0.379

0.416

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0

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0 10 20 30 40 50 60 70 80 90 100A

bso

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at 6

00

nm

% BBL Nutrient Broth in Synthetic Media

Absorbance Curve of E. coliFigure 1. Rhodoccocus ruber

Figure 5. Nutrient media B1, compositions B2-B9,

and synthetic media B10

Figure 6. Polystyrene tubes after vacuum oven

Temperature Tubes

26°C 30°C 34°C 26°C v 30 26

p-Value 0.010573 0.979645 0.464697 0.112781 0.946409 0.00122641

pH Tubes

6.5 7.8 8.4 6.5 vs 7.8 7.8 vs 8.4 6.5 vs 8.4

p-Value 0.427406 0.156038 0.3204 0.517184 0.991219 0.61617

Table 1. T-test and Standard Deviation of Temperature Tubes and pH Tubes

Figure 2.1 Treatment of polyethylene at various temperature Figure 2.2 Treatment of polystyrene at various temperature

Figure 3.1 Treatment of polyethylene at different pH media Figure 3.2 Treatment of polystyrene at different pH media

Figure 4.1 Absorbance curve of R. ruber Figure 4.2 Absorbance curve of E. coli

RESULTS

● Optimal Temperature: Polyethylene (26°C) and

Polystyrene (30°C)

● Optimal pH: Polyethylene (7.8) and Polystyrene

(undetermined)

● Optimal media for R. ruber: 80% of BBL nutrient

broth in synthetic media

CONCLUSION

● Significant degradation of polyethylene is

observed at 26°C with pH environment of 7.8.

FURTHER RESEARCH

● Study of all three obtained variables in a single

system to observe the efficiency of R. ruber in

biodegradation of polyethylene.

ACKNOWLEDGEMENTS

Caldwell University (Natural and Science Department)

Independent College Fund of New Jersey