2490
Proceedings of the 18
th
International Conference on Soil Mechanics and Geotechnical Engineering, Paris 2013
Portland cement through either direct substitution or
complementary use of MICP could contribute considerably
towards reduction in CO
2
emissions. Research suggests that
cementation using MICP can address a number of important
geotechnical problems in granular soils, including slope
stability, erosion and scour, under-seepage of levees, the
bearing capacity of shallow foundations, tunneling, and seismic
settlement and liquefaction (Dejong et al. 2010, Harkes et al.
2010, Kavazanjian and Karatas 2008, van Paassen et al. 2010).
1.3
Ureolytic MICP
MICP attempts to create a cemented soil mass by precipitating
calcium carbonate from the pore fluid such to form cementation
bonds at the interparticle contacts (van Paassen et al. 2010,
DeJong et al. 2006). Karatas et al. (2008) have identified several
mechanisms for MICP. The MICP mechanism that has garnered
the most attention and is most advanced in terms of
development is ureolytic hydrolysis, or ureolysis (Chou et al.
2011, DeJong et al. 2006, van Paassen et al. 2010, Whiffin et al.
2007). Ureolytic MICP has typically been accomplished using a
technique best described as biogrouting (Harkes et al. 2010, van
Paassen et al. 2010), wherein bacteria and nutrients are mixed in
a tank ex-situ and then injected into the soil followed by a
fixation fluid to foster microbial attachment to soil particles and,
finally, by a calcium-laden cementation fluid. Ureolytic MICP
by stimulation of indigenous bacteria has also been reported in
the literature (Burbank et al. 2012).
1.4
Agricultural Urease
Urease is a widely occurring hexameric protein found in many
microorganisms, higher order plants, and some invertebrates.
The enzyme is approximately 12 nm in dimension (Blakely &
Zerner 1984). The small size of a solubilized urease enzyme
affords it a distinct advantage over carbonate cementation
methods that employ ureolytic microbes in cases that require
penetration into very small pore spaces as nearly all known
bacteria are greater than 300 nm in diameter, with the majority
in the range of 500-5000 nm. Several families of common
plants are very rich in urease, including some varieties of beans,
melons and squash, and the pine family (Das et al. 2002).
Extraction of urease enzyme from most urease containing plants
has been shown to be very simple (Srivastava et al. 2001) and
the enzyme is readily available from laboratory suppliers.
It is well-established that urease can occur as both an intra-
and extra-cellular enzyme (Ciurli et al. 1996, Marzadori et al.
1998). Free soil urease (i.e. urease not bound to any living
organism), generally derived from dead and decaying
microorganisms and possibly from plant sources, readily occurs
apart from the host microorganism and, upon absorptive
association with soil particles, can persist for long periods of
time without degradation or loss of function (Pettit et al. 1976).
By contrast, exogenously added urease (i.e. urease added as a
free enzyme) has a limited lifespan and its activity and function
decrease with time (Marzadori et al. 1998, Pettit et al. 1976).
This limited lifespan is potentially advantageous in some
engineering applications as the enzyme can naturally degrade
thereby eliminating long term impacts to the ecosystem.
2. METHODS
2.1
Ottawa 20-30 Sand
Laboratory column tests were conducted using plant derived
urease to induce CaCO
3
precipitation in Ottawa 20-30 sand
These tests were carried out in 6”x 2” (152 mm x 51 mm)
acrylic tubes and membrane-lined 2.8” x 6” (71 mm x 152 mm)
split molds (for creating specimens for triaxial testing). Three
acrylic tubes and two columns for triaxial testing were filled
with 20-30 Ottawa silica sand (mean grain size 0.6 mm,
coefficient of uniformity 1.1) and treated as follows: tube #1:
the sand was dry pluviated via funnel at ≈3” (76 mm) drop
height and then received 5 applications of a cementation
solution containing urea and calcium chloride mixed with
1.4g/L enzyme (total solution volume ≈ 300 ml); tube #2: sand
was added in same manner as tube #1 and then received 2
applications (≈ 150 ml total) of the same cementation solution
mixed with 1.4g/L enzyme; tube #3: the lower-third of tube was
filled with sand and dry enzyme (≈ 3g), the remainder of the
tube contained dry pluviated sand without enzyme, and the tube
then received 2 applications (≈ 150 ml) of the cementation fluid
with no enzyme added. The cementation fluid composition was
based upon stochiometry and experience with microbial urease
cementation, e.g. DeJong et al. (2007), Whiffin et al. (2008).
Approximately 100 mL of a pH=7.8 solution containing 383
mM urea (reagent grade, Sigma-Aldrich), 272 mM CaCl
2
-2H
2
O
(laboratory grade, Alfa Aesar) was used for the first application
in each acrylic tube. Subsequent applications employed
approximately 50 mL of a pH=7.6 solution containing 416 mM
urea and 289 mM CaCl
2
-2H
2
O. Solution concentrations, while
variable, were formulated within a reasonably similar range as a
matter of convenience. In each application, the cementation
fluid was poured into the top of the acrylic tube with the bottom
closed off. The cementation fluid was allowed to stand, loosely
covered, in the acrylic tube for at least 24 hours and then
drained out the bottom of the cylinder. The next application
followed immediately after drainage was complete. Drainage
was accomplished by puncturing the base of the cylinder with a
20-gauge needle. When drainage was complete, the needle was
removed and the puncture was plugged with a dab of silicone.
Occasionally, the needle became plugged and an additional
needle was inserted through the base. The triaxial columns
were filled with sand in the same manner as tube 1 and then
received 2 applications (each application ≈ 250 ml) of
cementation solution with 1.4g/L enzyme.
In each application of cementation fluid, the fluid was
added until it rose to approximately ½-inch (12-mm) above the
soil line. After 2 applications, tubes #2 and #3 were allowed to
air dry for several days and then analyzed. Experimentation
with tube #1 was continued for several more days as three more
batches of cementation fluid were applied. The last 2
applications of cementation fluid were allowed to slowly drain
through the needle in the base immediately after application
rather than sit for 24 hours (drainage rate ≈10-25ml/hour). The
triaxial columns were allowed to stand for at least a week after
the second cementation fluid application and then drained.
After drainage was complete, the triaxial columns were
moved to a triaxial testing device. After draining the specimens
from the acrylic tubes and after the completion of the triaxial
tests, all samples were triple washed with de-ionized water.
Tubes #2 and #3 were separated in 3 layers, while tube #1 was
separated into six layers (for better resolution). Each layer from
the specimens in the acrylic tubes and the entire mass of the
triaxial specimens were acid washed to determine CaCO
3
content by oven drying for 48 hours, weighing, digesting with
warm 1M HCl, washing, drying, and reweighing to determine
carbonate mineral content.
Several of the cemented specimens were analyzed for
mineral identification using X-Ray Diffraction (XRD). Samples
were ground in an agate mortar and pestle and powdered onto a
standard glass slide for analysis. Scanning electron microscopy
(SEM) imaging was performed on intact cemented chunks of
material with an Agilent 8500 Low-Voltage SEM (LV-SEM). A
LV-SEM is a field emission scanning electron microscope
capable of imaging insulating materials, such as organic and
biological substances without the need for a metal coating and
without causing radiation damage to samples.
2.2
Ottawa F-60 Sand
A triaxial column was prepared using Ottawa F-60 silica sand
(mean grain size 0.275 mm, coefficient of uniformity 1.74) to
