Biological Desulfurization

Murray Gray
Department of Chemical and Materials
Engineering
Phillip M Fedorak and Julia M Foght
Department of Biological Sciences
University of Alberta, Edmonton, CANADA

1
Biodesulfurization vs. Bioremediation

Biodesulfurization - the selective removal

of sulfur from organosulfur compounds,
without the removal of carbon atoms.
 Retain the fuel value.

Bioremediation - the complete mineraliz-

ation of organosulfur compounds.
 All bonds are broken.
 CO2, H2O, and SO4= are products.
 Reduce toxicity.
2
Microbial Requirements

 Temperatures < 80 oC
Higher temperature organisms are known but only at
extremely high pressure
 Water all microbes live in aqueous phase
 Energy source
 Biochemical raw materials C, N, P, S, Fe
 Terminal electron acceptor for respiration

Topsoe Catalysis
Forum 2007 Slide 3
Nutrients for microbes

 Macronutrients make up most of cell

Carbon source = petroleum hydrocarbons,
sugars, lipids, proteins, carbon dioxide,
methane
Nitrogen source = nitrate, nitrogen, organic
nitrogen compounds
 Micronutrients are needed in small
amounts
Range of elements P, S, Fe, Ca, Mg..

Topsoe Catalysis
Forum 2007 Slide 4
 Aerobic Metabolism
Carbon
Dioxide

Initial Characteristic
Oxygenase Degradation Central
Attack Pathways Metabolism

Contaminant Detoxification?
Nutrient scavenging?
(Nitrogen, Sulfur)

Oxidized Metabolites
Topsoe Catalysis
Forum 2007 Slide 5
Intracellular vs Extracellular

 Large molecules are attacked outside cells =

extracellular
Fungi attack cellulose and lignin
Bacteria attack proteins and lipids
Requires excretion of enzymes, loss of energy
 Small molecules are attacked inside cells =
intracellular
Cells conserve valuable enzymes

Topsoe Catalysis
Forum 2007 Slide 6
Bacterial transport

 Most substrates (carbon source, energy

source) must enter the cell for processing
 Cell wall controls ion transport
 Small molecules can diffuse across (e.g.
oxygen)
 Large molecules cannot enter

Topsoe Catalysis
Forum 2007 Slide 7
Aqueous solubility
Compound Water solubility (mg/L)
Benzene 1750
Naphthalene 31.0
Phenanthrene 1.1
Anthracene 0.05
Pyrene 0.13
Chrysene 0.002
C10 (decane) 0.052
C16 (hexadecane) 0.00002

chemical or bio- surfactants increase apparent solubility

Topsoe Catalysis
Forum 2007 Slide 8
 Dealing with low solubility, high Kow

 Hydrophilic cells (e.g. Gram

negative)
Biodegradation is limited by mass
transport from hydrophobic phase to cell
interior
 Hydrophobic cells (e.g. Gram
positive)
Biodegradation is limited by association
with hydrophobic phase (droplet or
crystal)
Biodegradation is limited by surface area
and influenced by Kow Topsoe Catalysis
Forum 2007 Slide 9
Hydrophobic cell surfaces

Rhodococcus sp. S1 Rhodococcus sp. 20S-E1c

anthracene crystal
hexadecane droplet

M. Pickard (1996) Univ. of Alberta L. Dorobantu (2004) Univ. of Alberta

Topsoe Catalysis
Forum 2007 Slide 10
Emuslification by Acinetobacter
- bacterial attachment and emulsification
- changes in interfacial tension and droplet
size

6h

12 h

Baldi et al., AEM 65:2041-2048 (1999)

28 h Topsoe Catalysis
Forum 2007 Slide 11
 Overcoming low substrate
solubility
 Bacterial uptake mechanisms:
5
Passive uptake: Very fluoranthene

Log partition coefficient

4 anthracene
rapid and related to Kow naphthalene 3 phenanthrene
e.g. Pseudomonas fluorescens LP6a 2
T. Bugg (2000) Appl. Environ.Microbiol. 66:5387 toluene
1

0
2 2.5 3 3.5 4 4.5 5 5.5

Log Kow

Active uptake: specific, saturable uptake

in addition to passive uptake:
e.g., Mycobacterium sp. RJGII-135 and phenanthrene
N. Miyata (2004) Appl. Environ. Microbiol. 70:363 Topsoe Catalysis
Forum 2007 Slide 12
Dibenzothiophene:
Biodesulfurization
+ SO4 2-
S
HO
2-Hydroxybiphenyl Sulfate

Mineralization
+ 15.5 O 2 12 CO 2 + 3 H 2 O + H 2 SO4
S

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Three known pathways for the
aerobic metabolism of DBT

4S pathway conserves carbon backbone

van Afferden pathway

Kodama pathway

Anaerobic metabolism is not known

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4S pathway for biodesulfurization
Repressed by Sulfate

S S S

O O O

Sulfoxide Sulfone

2-
+ SO 3 2 - SO 4

HO SO2 H HO Sulfate
2'-Hydroxybiphenyl- 2-Hydroxybiphenyl
2-sulfinic acid

- Multiple enzyme steps so only intracellular

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Energy Biosystems
Biodesulfurization Process

 Desulfurization of diesel fuel

 Rhodococcus erythropolis IGTS8 gave 4S
pathway
 Genetically modified to enhance activity
 Operated at ambient temperature and
pressure
 Extensive pilot testing

Topsoe Catalysis
Forum 2007 Slide 16
Biodesulfurization process
Vent gas

Oil Phase
Separator Diesel
Cleanup
Product

Waste
Stage 2 Biomass
Separator

Water +
Multi-Stage Water
Nutrients Bioreactor Cleanup

Catalyst Byproduct
Recycle Recovery Sulfinate
Air
Byproduct
Biocatalyst
Production
and Regeneration
Waste
Diesel fuel Water
Water recycle
Topsoe Catalysis
Forum 2007 Slide 17
EBS Separation Unit

Yu et al., US Patent 5,772,901 (1998) Topsoe Catalysis

Forum 2007 Slide 18
Selectivity of Enzymes

Enzymes can be extremely selective

Desulfurization enzymes from IGTS8 give some
activity for other sulfur heterocycles and non-
thiophenic sulfur
 Best activity for dibenzothiophene
 Less active for alkyl dibenzothiophenes and dialkyl
dibenzothiophenes

Each species of bacteria is more selective than
NiMoS catalysts
 Limited range of boiling point
 Limited range of sulfur species

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Enzymatic desulfurization

 Proposed by Vasquez-Duhalt et al. as

follows: Peroxidase enzyme

+ H2O2

S S
O

 Sulfoxide has higher boiling point

 Remove desulfurized fraction by
distillation
Topsoe Catalysis
Forum 2007 Slide 20
Viscosity of heavy oil

Viscosity is correlated
S to the molecular
weight of the oil

Cleavage of aliphatic
S
sulfide bridges may
S
reduce the size of
O large oil molecules
NH  Should contribute to
S Reaction viscosity reduction
points?

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Research objectives

Isolate bacteria active towards model
compounds with aliphatic sulfide linkages

Characterize activity of isolates towards

model compounds

Assess suitability of isolates for

bio-upgrading

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Model compound synthesized
F F
F F
S
F F F F
F F
bis-(3-Pentafluorophenylpropyl) sulfide (PFPS)

Rhodococcus sp. strain JVH1 degrades

PFPS as sole sulfur source

Does not oxidize or desulfurize
dibenzothiophene
Van Hamme et al. (2004) Appl. Environ. Microbiol. 70:1487
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PFPS desulfurization pathway
F F F F
F F F F
S S
F F F F F F O F F
F F F F
Sulfide Sulfoxide

F F F
F F F Sulfinic
S SO2H
F F O O F F F F acid
F F F
Sulfone [SO32-] [SO42-]
F
F
OH
F F
F Alcohol
Van Hamme et al. (2004) Appl. Environ. Microbiol. 70:1487
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Effect of JVH1 on Viscosity
Sample Viscosity, Pa.s Sulfur, wt%
at 25 oC
Crude Oil 1.60.1 2.880.02

Filtered oil 19.60.1 3.230.02

Sterile controls 51.61.5 3.380.01

Live cells 78.52.6 3.460.01

Killed cells 69.63.8 3.380.01

Lloydminster heavy oil, incubated 17 days with JVH1

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Observations

No evidence for bacterial reduction in
viscosity

Both live cells and killed cells increased
viscosity compared to control

Cells were active against test compounds
in presence of crude oil (positive control),
but no significant activity against sulfur in
heavy fractions

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General issues with bio-processing
of crude oil
 Intracellular processes face major
challenges
Limited effectiveness for large molecules
Maintenance of active biomass
Recovery of biomass from oil/water mixtures
 Accumulation of inhibitory metabolites and
abiotic dead-end products
 Mixed substrates: competitive inhibition of
enzymes; sequential degradation of
substrates Topsoe Catalysis
Forum 2007 Slide 27
Hydrothermal vent communities

 Black smokers
provide chemical
energy and
terminal electron
acceptors
 Support complex
ecosystems

Topsoe Catalysis
Forum 2007 Slide 28
Seafloor anaerobic community

Topsoe Catalysis
Forum 2007 Slide 29
Shell-Paques Biological Process
(Conceptual flowsheet)

Soda solution containing hydrogen sulfide-oxidizing bacteria

Process suitable for 0.5-30 t/d (GPSA)
Topsoe Catalysis
Forum 2007 Slide 30