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A Comparative Study on Biodegradation of selected Monomers: Effect of Molecular Structure

A Comparative Study on Biodegradation of selected Monomers: Effect of Molecular Structure. Jongtai Jung (Professor/Ph. D). Major of Environmental Engineering College of Urban Science , University of Incheon. Introduction. The biological breakdown and degradation of both

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A Comparative Study on Biodegradation of selected Monomers: Effect of Molecular Structure

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  1. A Comparative Study on Biodegradation of selected Monomers: Effect of Molecular Structure • Jongtai Jung • (Professor/Ph. D) • Major of Environmental Engineering • College of Urban Science, University of Incheon

  2. Introduction • The biological breakdown and degradation of both • synthetic and natural monomers and polymers is • becoming increasingly important for a broad spectrum • of application. • The factors that determine biodegradability of a polymer • are the linkage among its monomers, molecular weight of • the polymer, it's origin(synthetic or natural), and • molecular structure of the monomer. • A systematic study of the degradation of monomers may • be helpful in understanding of the degradability of • polymers

  3. The objective of this work • To study biodegradation of monomeric units • such as β-hydroxybutyric acid(HBA), MMA(metyl • metacrylate) and styrene using an immobilized • activated sludge, • and to compare their relative degradation rates • as function of their molecular structure.

  4. Table 1. Properties of Selected Monomers

  5. Fig 1. Experimental set-up of the bioreactor

  6. Experimental Set-up(1) 1) Recirculation flow-type bioreactor, - Reactor size : 6.4 cm in diameter 20 cm in length.. 2) Reservoir - Reservoir size : 11.4 cm in diameter 25.4 cm in length 3) Total reaction volume - 2 liters including the reservoir.

  7. Experimental Set-up(2) 1) Culture medium - 100 ppm MgCl2, - 0.5 ppm FeCl3 - 10 ppm MgSO4, - 10 ppm K2PO4 • Oxygen supply - Air (1.5 liter/min for HBA,) - H2O2 (3% v/v for Styrene, MMA) • Recirculation flow rate : 325ml/min. • Chosen Substrate - MMA(Methyl methacylate), Styrene - β-hydroxybutyric acid(HBA),

  8. Microorganism • Activated sludge(Mixed microbial population) • from Waste water treatment plant • 100 g alginate-immobilized activated sludge • How to immobilize • - Distilled water • - Concentrated sludge(50 mg dry biomass/ g of pallet) • - 0.5% sodium chloride • - 1% sodium alginate • - 0.1 mol/liter CaCl2 • - Distilled water and Conc. Pellets in a ratio 5:2 mixed • with NaCl and Sodium Alginate in a blender • - The homogeneous cell suspension was then extruded • using a syringe pump into CaCl2 solution to obtain the • immobilized bacterial beads

  9. Analytical Methods • Dissolved Oxygen concentration : • - Clark-type dissolved oxygen probe • -Chart recorder. • 2) MMA, Styrene Concentration : • - Perkin Elmer8500 Gas Chromatograph, • -Detector : FID • 3) HBA Concentration • - Perkin Elmer HPLC • - Diode array detector at a wavelength of 215 nm • pH : • - pH probe (Orion Cat. No. 91-04) • - pH meter (Corning Model 250, NY)

  10. Experimental Methods - Experiments were conducted by spiking the reservoir • with the compound of interest to a predetermined • concentration • Samples from the reservoir were then analyzed • periodically to determine the rate of biodegradation • after accounting for abiotic losses • Control experiment were performed with only alginate • beads(without the microbes) under identical • experimental conditions to determine abiotic losses • due to adsorption, volatilization, and chemical reactions

  11. Parameters to be monitored • Rate of oxygen consumption • (nmol/min∙ml) • 2) pH • 3) Rate of phenol biodegradation(ppm/hr)

  12. Results and discussions

  13. 1) Biodegradation of styrene(1) • Figure 2 shows biodegradation of styrene, as well as • abioticremoval of styrene from the reactor. • The data indicate that initially the concentration of • styrene drops rapidly due to its adsorption on the • alginate beads. • This is shown by the line AB and AD. Later, the drop • in concentration is gradual in both cases, but relatively • higher when biomass is present (see line BC and DE). • Line BC indicates losses due to volatilization while • line DE indicates removal of styrene by both biotic • and abiotic losses.

  14. Fig 2. Biodegradation of Styrene

  15. 1) Biodegradation of styrene(2) • Asimilar trend is observed in successive spikes of • styrene to the reactor, which suggests that the absorbed • styrene from the beadsslowly desorbed back into the • solution and was subsequentlydegraded. • As seen in Figure 2, starting from a concentration of • 75 ppm, 27% of styrene is very quickly adsorbed • on the alginate beads within 2 hours in both cases • (see curves AB andAD). • After equilibrium is reached, a comparison of the two • removal rates (see curves BC and DE) shows that • about 20% of styrene islost by abiotic means and • the rest 80% is degraded biologically.

  16. 1) Biodegradation of styrene(3) • The biodegradation rate is 3.3 ppm/hr. • Here, it is assumed that the initially adsorbed styrene is • eventually desorbed, and degraded. • It needs to be addressed that hydrogen peroxide has a • short half-life in presence of the enzyme catalase. • The possibility of a chemical reaction between • hydrogen andthe monomers(styrene or MMA) is less • likely due to the verylow H2O2concentrations in the • presence of catalase.

  17. Fig 3. Substrate Dependent Oxygen Consumption • △△△ control for HB sodium salt • ▲▲▲ substrate dependent O2 consumption for HB sodium salt • □□□ control for styrene • ■■■ substrate dependent O2 consumption for styrene • ○○○ control for MMA • ●●● substrate dependent O2 consumption for MMA

  18. 1) Biodegradation of styrene(4) • Figure 3 shows oxygen uptake rates in the presence • and absence of substrates. • During every batch experiment, the oxygen uptake rate • is measured before and after the injection of the • substrate. • As shown by the two sets of intersecting lines • representing two styrene injections, bio-oxidation rates • in the presence of styrene are higher than that without • styrene. • This is defined as substrate dependent oxygen uptake, • and is a tool used to determine biodegradability based • on response to oxygen uptake.

  19. 1) Biodegradation of styrene(5) • Figure 4 also shows the variation in pH during • biodegradation of MMA. Starting from 6.95, • the pH drops to 6.01. • Again a drop in pH suggests mineralization of MMA • to acids, and eventually to carbon dioxide.

  20. Fig 4. pH Variation during Biodegradation

  21. 2) Biodegradation of MMA(1) • Control experiments were also performed with • hydrogen peroxide, and without biomass to account • for abiotic losses of MMA. • As seen in figure 5, over a period 8hrs 11% of MMA • was lost by abiotic means which means that 89% of • the total removal in biological treatment experiments • can be accounted as due to biodegradation • The abiotic losses are primarily due to removal of • MMA by volatilization from the reservoir.

  22. Fig 5. Biodegradation of MMA

  23. 2) Biodegradation of MMA(2) • Once again it may be assumed that losses due to • chemical reaction with hydrogen peroxide are negligible. • Unlike styrene, MMA was neither significantly • adsorbed on alginate beads nor volatilized. • The biodegradation rate for MMA at 75 ppm starting • concentration is 9.3 ppm/hr. • Figure 6 shows performance of the bioreactor when • challenged with multiple injection of MMA on a • continuous basis. • Concentrations as high as 550ppm MMA are treated, • and the biodegradation ability of the microbes remains • consistent.

  24. Fig 6. Biodegradation of MMA during Multiple Injection

  25. 2) Biodegradation of MMA(3) • Figure 3 also shows oxygen uptake rate for MMA. • Once again the oxygen uptake rates are higher in a • presence of MMA than that in the absence of MMA, • indicating substrate dependent oxygen uptake. • Figure 4 also shows the variation in pH during • biodegradation of MMA. Starting from 6.95, the pH • drops to 6.01. • Again a drop in pH suggests mineralization of MMA to • acid, and eventually to carbon dioxide

  26. 3) Biodegradation of HBA(1) • Biodegradation of hydroxybutyricacid was studied • by injecting its sodium salt in the reactor. • Control experiments were done without biomass • to account for any abiotic losses of HBA as before. • Over a period of 5 hours essentially no HBA was either • adsorbed to the alginate beads or stripped from the • system. • Biodegradation experiments were conducted at 75 ppm • starting concentrations of HBA. As seen in Figure 7, • the biodegradation rate is 15ppm/hr.

  27. Fig 7. Biodegradation of HBA

  28. 3) Biodegradation of HBA(2) • Figure 4 shows the variation in pH during the course of • HBA biodegradation. • Unlike that observed for both styrene and MMA, • the pH in this case shows an increasing trend. • Starting from 6.85, the pH increased to 8.5. • Since the substrate is the sodium salt of HBA, • its mineralization would result in the formation of • either NaHCO3 or Na2CO3, and consequently cause • a rise in pH. • We have subsequently confirmed this phenomenon • with other acids and their salts.

  29. 4) Comparison of Parameters(1) • Results from these experiments indicate that styrene, • MMA, and HBA can all be biodegraded using • activated sludge. • A comparison of maximum biodegradation rates • (3.3,9.3 and 15 ppm/hr for styrene, MMA, and HBA • respectively) indicates the scale of difficulty in • biodegrading the above monomers. • The biodegradation trend reflected by these rates is • analogous to the observed trend in oxygen • consumption rates and pH during their respective • biodegradation experiments( see Figure 3,4).

  30. 4) Comparison of Parameters(2) • Table 2 below summarizes the response as measured • in terms of oxygen uptake when the microbes are • challenged with monomers during five different tests. • It is observed that the response from baseline oxygen • uptake is higher for HBA followed by MMA and • styrene. • The results indicate the natural relative degradability • of these monomers in the environment.

  31. 4) Comparison of Parameters(3) • It is clearly seen that styrene which is a benzene • derivativeis relatively difficult to degrade because • the degradation mechanism comprises of a formidable • ring opening step. • Both MMA and HBA are straight chain compounds, • and they can be biodegraded relatively easily. • A comparatively high biodegradation rate for HBA is • understandable because the monomer does not have • double bond in its chemical structure.

  32. Table 2. Bio-oxidation Response of Monomers A: Baseline oxidation rate without substrate( nmole O2/ml ∙min.) B: Initial oxidation rate with substrate ( nmole O2/ml ∙min.) C: Ratio of B to A

  33. Conclusions(1) • Many factors determine the biodegradability of any • monomer as mentioned earlier. In this study only one • factor (monomer structure) has been investigated. • Although final conclusions can only be drawn after • all the factors have been studied, this study in monomer • biodegradation. • If other factors were not limiting, then based on these • results we can say that HBA may be more easily • degraded than MMA,which is comparatively easy to • degrade than styrene.

  34. Conclusions(2) • HBA which has straight chain without double bond can • be used as a reference to assess relative degradability • of other monomers. • On a scale of 1 to 100, it can be said that HBA has a • degradabilitynumber of 100, styrene of 22, and MMA of • 62. • The higher the number, The higher is the chance to • degrade the monomer naturally.

  35. Thank you very much for listening

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