Microbially Enhanced Oil Recovery by Sequential Injection of Light Hydrocarbon and Nitrate in Low- And

High-Pressure Bioreactors
Gassara et al., 2015

INTRODUCTION
 Extracting more oil from reservoirs through EOR methods is a major challenge to the oil industry
 oil-soluble, low molecular weight MEOR substrates, like toluene or heptane, could be
advantageous, as these bind to oil preventing them from being lost during continued water
injection.
 Injection of nitrate, a high-potential electron acceptor for heterotrophic nitrate-reducing bacteria
(hNRB) in oil fields use alkylbenzenes, especially toluene, as the preferred electron donor.
 Current study explores whether increasing the concentration of toluene or of toluene and
heptane in MHGC oil increases nitrate reduction and the associated production of heavy oil.
MATERIALS AND METHODS
 Toluene-Oxidizing Nitrate-Reducing Enrichment Cultures:
- Heavy oil from the MHGC field near Medicine Hat, Alberta, Canada with an American
Petroleum Institute (API) gravity of 16° and a viscosity of 3400 cP at 20 °C was used
- field is subject to nitrate injection to limit microbial H2S production and produced waters
contain a significant fraction of the hNRB Thauera.
- MHGC oil (1 mL) was amended with up to 570 mM of toluene to reduce up to 80 mM of nitrate
in 47.5 mL of aqueous phase.
- Samples from enrichment cultures were taken with N2−CO2 flushed syringes to measure
nitrate and nitrite concentrations with high performance liquid chromatography
 Low pressure bioreactors:
- Concentrations of nitrate and nitrite in the effluent aqueous phase and of toluene in the
effluent oil phase were measured by HPLC and GC-MS, respectively
 High Pressure Bioreactor:
- Effluent fractions were collected to determine produced oil and its concentration of toluene,
as well as the aqueous concentrations of nitrate and nitrite.
 Increasing the Toluene Concentration of ROIP by Injecting Aqueous Toluene:
- Bioreactors were injected with CSBK containing up to 3 mM of dissolved toluene
- Increased concentrations of toluene in produced or residual oil were determined from the
OD600 of the DCM extracts and their toluene concentration as determined by GC-MS
 Quantification of Toluene in Oil by Gas Chromatography−mass Spectrometry (GC-MS):
- After measuring the oil content of the produced oil−water mixtures, 1 mL of each oil−DCM
phase was amended with 1 μL of internal standard
- The concentration of toluene in MHGC oil was calculated from its peak area relative to that
of the internal standard
 Chemical and Physical Analyses: The bioreactor effluent was analyzed for the chemical
parameters of emulsification index and emulsification index % and the physical parameters of
pressure difference (ΔP) between the inlet and the outlet, flow rate through the porous medium
q, permeability K (Darcy), cross-section area A, path length L (cm) viscosity η (cP) by using the
Darcy’s Law equation.
 Microbial Community Analysis:
RESULTS
 MEOR with Oil-Soluble Electron Donors in Low-Pressure Bioreactors:
With high toluene or high toluene and heptane and 80 mM nitrate was 23.0 ± 3.9% of ROIP (N =
5), whereas the average was 6.3 ± 0.8% of ROIP (N = 5) with 0 mM nitrate

 MEOR with Aqueous Electrons Donors in LowPressure Bioreactors:

In the presence of sufficient molasses, glucose or acetate to reduce 80 mM of nitrate, the average
production was 25.0 ± 11.1% of ROIP (N = 5), similar to that obtained with high toluene or high
toluene and heptane.

 Toluene- and Nitrate-Mediated MEOR in HighPressure Bioreactors

- Toluene oxidation was coupled to nitrate reduction with 10−17 mM of nitrate and 2−5 mM
nitrite remaining
- In the absence of nitrate no toluene oxidation was observed

 Retention of Aqueous Toluene Injected into Oil Containing Bioreactors

- The distribution of toluene between oil and aqueous phases is governed by the partition
coefficient π = Co/Cw, where Co and Cw are the equilibrium concentrations of toluene in the
oil phase and in water, respectively
- π of toluene for MHGC oil =17800 ± 3000, - π of toluene for water= 1380 ± 420; indicating
that equilibrium had not yet been reached under these conditions
- Nevertheless, these values indicate that if aqueous toluene is injected slowly most would be
expected to be retained by the oil.
- because injected aqueous toluene is transferred from the aqueous phase into a viscous oil
phase where diffusion is slow, its concentration in volume elements lining aqueous channels
may be higher than the calculated average Co.

 Sequential Injection of Toluene and Nitrate in Low Pressure Bioreactors

- sequential injection of toluene and nitrate gave an additional 17.6 ± 7.0% of ROIP
- sequential injection of toluene also created low concentrations of nitrate in effluents
- In stage 1 of oil production, all the bioreactors regardless of sequential injection, underwent
large pressure drop
- In stage 2 of incubation, pressure rise was observed for BioX6 that underwent the sequential
injection. To maintain a constant flow rate, this rise in pressure was equivalent to a decrease
in area A and increase in viscosity.
 Sequential Injection of Toluene and Nitrate in High Pressure Bioreactors:
Similar results observed.
 Microbial Community Analysis
- Thauera dominated the community grown with toluene, as found in previous enrichment
cultures and in the field
DISCUSSION
 additional oil can be produced from heavy oil-containing bioreactors in which hNRB activity is
stimulated by the presence of added or injected oil-soluble electron donor (e.g., toluene) and
aqueous nitrate.
 ability of bacteria to produce biosurfactants, decreasing interfacial tension of oil held in small rock
pores
 Blockage of preferred flow paths by in situ production of biomass and exopolysaccharide or other
biopolymers is also thought to be an important MEOR mechanism
 Increased oil production through sequential injection of light hydrocarbon and nitrate to activate
hNRB is a novel MEOR technology
 HN-MEOR has the potential to be profitable