Feasibility of Using Microalgae for Biocement Production through Biocementation

Abstract

The invention of microorganism’s involvement in carbonate precipitation, has lead the exploration of this process in the field of construction engineering. Biocement is a product innovation from developing bioprocess technology called biocementation. Biocement refers to a CaCO3 deposit that formed due to microorganism activity in the system rich of calcium ion. The primary role of microorganism in carbonate precipitation is mainly due to their ability to create an alkaline environment (high pH and DIC increase) through their various physiological activities. Three main groups of microorganism that can induce the carbonate precipitation: (i) photosynthetic microorganism such as cyanobacteria and microalgae; (ii) sulphate reducing bacteria; and (iii) some species of microorganism involved in nitrogen cycle. Microalgae are photosynthetic microorganism and utilize urea using urease or urea amidolyase enzyme, based on that it is possible to use microalgae as media to produce biocement through biocementation. This paper overviews biocement in general, biocementation, type of microorganism and their pathways in inducing carbonate precipitation and the prospect of microalgae to be used in biocement production.

Full text

Research Article Open Access
Bioprocessing & Biotechniques
Ariyanti et al., J Bioproces Biotechniq 2012, 2:1
http://dx.doi.org/10.4172/2155-9821.1000111
Volume 2 • Issue 1 • 1000111
J Bioproces Biotechniq
ISSN:2155-9821 JBPBT, an open access journal
Keywords: Biocement; Biocementation; Microalgae; CaCO3 precipi-
tation
Introduction
Construction engineering consumes a large amount of materials
from non-renewable resources, which most of the materials contrib-
ute CO2 emission to the air at their production or application stage.
Technology development related to the construction material and their
production is necessary, in order to maintain the sustainability and to
reduce the production of CO2 emission. e evidence of microorgan-
ism involvement in carbonate precipitation, has lead the development
of bioprocess technology in the eld of construction material [1,2].
e precipitation of calcium carbonate (CaCO3) may be performed
due to microorganism activity and it produces massive limestone or
small crystal forms [3]. ese deposit of calcium carbonate known as
biocement or microbial induced carbonate precipitation (MICP) [3,4].
Biocement has many advantages compared to an ordinary cement,
such as: the production process is slightly dierent with sandstone pro-
duction, biocement need a much shorter time; it is suitable for in-situ
process; raw material of biocement are produced at low temperature,
more ecient compared to an ordinary cement which used tempera-
ture up to 1500C in production process; biocement can be used as eco-
construction material since it consume less energy and less CO2 emis-
sion in the production process rather than other ordinary cement [3,5].
Recently, research and study of biocement production through bio-
cementation still focused to the nitrogen cycle mechanism using urease
enzyme producing bacteria [3-7]. While research using microalgae as
media for biocementation still lack in literature, in fact microalgae have
a great potency for the objective of biocementation. Overview of bio-
cement, biocementation, type of microorganism, mechanism type and
feasibility of microalgae as media for biocement production will briey
described throughout this paper.
Microbial Induced Carbonate Precipitation (MCIP)
Calcium carbonate (CaCO3) precipitation is a common phenome-
non found in nature such as marine water, freshwater, and soils [1,6,8].
is precipitation is governed by four key factors: (i) the calcium (Ca2+)
concentration, (ii) the concentration of dissolved inorganic carbon
(DIC), (iii) the pH (pK2 (CO) = 10.3 at 25°C) and (iv) the availabil-
ity of nucleation sites [1,9]. Numerous species of microorganism have
been detected previously and assumed to be associated with natural
carbonate precipitates from diverse environments. e primary role of
microorganism in carbonate precipitation is mainly due to their abil-
ity to create an alkaline environment (high pH and [DIC] increase)
through their various physiological activities [1,6].
ere are three main groups of microorganism that can induce
the carbonate precipitation: (i) photosynthetic microorganism such as
cyanobacteria and microalgae; (ii) sulphate reducing bacteria; and (iii)
some species of microorganism involved in nitrogen cycle [1,6,7]. e
most common MCIP phenomena appeared in aquatic environments is
caused by photosynthetic microorganisms [7,10]. Photosynthetic mi-
croorganisms use CO2 in their metabolic process (equation 1) which
is in equilibrium with HCO3- and CO3
2- as described in equation 2.
Carbon dioxide consumed by photosynthetic microorganisms shi the
equilibrium and resulting the increment of pH (equation 3) [7]. When
this reaction occurs in the present of calcium ion in the system, calcium
carbonate is produced as described at chemical reaction in equation 4
[6].
CO2+ H2O  (CH2O) + O2 (1)
2HCO3- ↔ CO2 + CO3
2- + H2O (2)
CO3
2- + H2O ↔ HCO3-+ OH- (3)
Ca2+ + HCO3-+ OH- ↔ CaCO3+ 2H2O (4)
*Corresponding author: Dessy Ariyanti, Department of Chemical Engineering,
Faculty of Engineering, Diponegoro University, Prof Soedarto, SH Kampus Tembal-
ang, Semarang, Indonesia, Tel: +62-24-7460058; Fax: +62-24-76480675; E-mail:
dessy@undip.ac.id
Received December 24, 2011; Accepted January 19, 2012; Published January
22, 2012
Citation: Ariyanti D, Handayani NA, Hadiyanto (2012) Feasibility of Using
Microalgae for Biocement Production through Biocementation. J Bioprocess
Biotechniq 2:111 doi: 10.4172/2155-9821.1000111
Copyright: © 2012 Ariyanti D, et al. This is an open-access article distributed under
the terms of the Creative Commons Attribution License, which permits unrestricted
use, distribution, and reproduction in any medium, provided the original author and
source are credited.
Abstract
The invention of microorganism’s involvement in carbonate precipitation, has lead the exploration of this process
in the eld of construction engineering. Biocement is a product innovation from developing bioprocess technology
called biocementation. Biocement refers to a CaCO3 deposit that formed due to microorganism activity in the system
rich of calcium ion. The primary role of microorganism in carbonate precipitation is mainly due to their ability to create
an alkaline environment (high pH and DIC increase) through their various physiological activities. Three main groups
of microorganism that can induce the carbonate precipitation: (i) photosynthetic microorganism such as cyanobacteria
and microalgae; (ii) sulphate reducing bacteria; and (iii) some species of microorganism involved in nitrogen cycle.
Microalgae are photosynthetic microorganism and utilize urea using urease or urea amidolyase enzyme, based on
that it is possible to use microalgae as media to produce biocement through biocementation. This paper overviews
biocement in general, biocementation, type of microorganism and their pathways in inducing carbonate precipitation
and the prospect of microalgae to be used in biocement production.
Feasibility of Using Microalgae for Biocement Production through
Biocementation
Dessy Ariyanti*, Noer Abyor Handayani and Hadiyanto
Department of Chemical Engineering, Faculty of Engineering, Diponegoro University, Prof Soedarto, SH Kampus Tembalang, Semarang, Indonesia

Citation: Ariyanti D, Handayani NA, Hadiyanto (2012) Feasibility of Using Microalgae for Biocement Production through Biocementation. J Bioprocess
Biotechniq 2:111 doi: 10.4172/2155-9821.1000111
Page 2 of 4
J Bioproces Biotechniq
ISSN:2155-9821 JBPBT, an open access journal Volume 2 • Issue 1 • 1000111
e precipitation of calcite (CaCO3) can also be induced by het-
erotrophic organism. is microorganism produces carbonate or bi-
carbonate and modied the system so that the carbonate precipitation
may occur [1]. Abiotic dissolution of gypsum (CaSO4.H2O) (equation
5) causes system rich of sulphate and calcium ion. In the presence of
organic matter and the absence of oxygen, sulphate reducing bacteria
(SRB) can reduce sulphate to H2S and HCO3- as described in equation
6 [1,7]. When the H2S degasses from the environment, pH of system
will increase and the precipitation of calcium carbonate will occur [1].
CaSO4.H2O  Ca2++ SO4
2-+ 2H2O (5)
2(CH2O) + SO4
2- HS- + HCO3
-+ CO2 + H2O (6)
Currently, urease enzyme activity in most of microorganism me-
tabolism process has been used as a tool to induce the precipitation of
calcium carbonate [11,12]. e hydrolysis of urea by urease enzyme in
heterotrophic microorganism will produce carbonate ion and ammo-
nium. is mechanism will result system with higher pH and rich of
carbonate ion [12]. One mole of urea hydrolysed intracellularly to one
mole ammonia and one mole carbamate (equation 7), which spontane-
ously hydrolysed to one mole ammonia and one mole carbonic acid
(equation 8). Ammonia and carbamate subsequently equilibrate in wa-
ter to form bicarbonate and 2 moles of ammonium and hydroxide ions
as described in equation 9 and 10 [2].
CO(NH2)2 +H2O H2COOH + NH3 (7)
NH2COOH + H2O  NH3 + H2CO3 (8)
2NH3 + 2H2O  2NH4
+ + 2OH- (9)
2OH- + H2CO3  CO3
2- + 2H2O (10)
Total reaction:
CO(NH2)2 + 2H2O  2NH4
+ + CO3
2- (11)
e presence of calcium ion in the system will lead to the calci-
um carbonate precipitation once a certain level of supersaturation is
reached. e calcium carbonate precipitation mechanism induced by
urease enzyme activity illustrated in gure 1.
Calcium ions in the solution are attracted to microorganism cell
wall due to the negative charge of the latter. Aer the addition of urea to
the system, microorganism convert urea to dissolved inorganic carbon
(DIC) and ammonium (AMM) and released it to the environment (A).
e presence of calcium ion cause the supersaturation condition and
precipitation of calcium carbonate in microorganism cell wall (B). Af-
ter a while, the whole cell becomes encapsulated by calcium carbonate
precipitate (C). As whole cell encapsulated, nutrient transfer becomes
limited and resulting in cell death. Image (D) shows the imprints of
microorganism cell involved in carbonate precipitation [6].
Biocementation
Biocementation is a process to produce binding material (bioce-
ment) based on microbial induced carbonate precipitation (MICP)
mechanism. is process can be applied in many elds such as con-
struction, petroleum, erosion control, and environment. Application
in construction eld include wall and building coating method, soil
strengthening and stabilizing, and sand stabilizing in earthquake prone
zone [2].
In application, the precipitation of calcium carbonate (biocement)
is combined with other supporting material such as sand. e patented
method of producing biocement can be seen in gure 2 [7,4].
Biocementation illustrated in gure 2 uses heterotroph bacteria
Bacillus pasteurii with urea hydrolysis mechanism. e cementation
process occurs in pipe columns lled with commercial sand contained
silica. Urea/calcium solution and bacteria solution were mixed imme-
diately and put in the pressurized vessel to be injected to the sand core
in pipe column for several time until the sand core fully saturated. Bio-
cementation takes about 24 hours to complete the reaction, aer that
the biocement were dried in temperature of 60˚C [7].
Biocementation were also developed in the process of biological
mortar production, crack in concrete remediation and production of
bacterial concrete [2,9]. Table 1 shows overview of various construc-
tion materials made from biocementation.
In general, mortar refers to “ready to use” binder material con-
tained a binder, and sand or aggregate. Biological mortar consists of
three main components such as limestone powder, nutrient and bacte-
rial paste [2]. Biocementation applied in concrete ri remediation and
the production of bacterial concrete has been investigated (Santhosh
et al. [13]). Specimen of crack in concrete lled with biocement shows
the signicant increment of strength and stiness value compared with
specimen without biocement [13].
eoretically, calcium carbonate precipitation occur in nature fol-
lowing several process such as: (i) abiotic chemical precipitation from
saturated solution due to evaporation, temperature increase and/or
pressure decrease; (ii) production of external and internal skeleton by
eukaryotes; (iii) CO2 pressure derivation under eect of autotrophic
processes (photosynthesis, methanogenesis); (iv) fungal mediation;
(A) (B) (C) (D)
Ca
Ca
Ca
Ca
Ca
Ca
Ca
DIC
UREA
AMM
2 µm
Figure 1: Illustration of calcium carbonate precipitation mechanism induced by urease enzyme activity in microorganism [6].

Citation: Ariyanti D, Handayani NA, Hadiyanto (2012) Feasibility of Using Microalgae for Biocement Production through Biocementation. J Bioprocess
Biotechniq 2:111 doi: 10.4172/2155-9821.1000111
Page 3 of 4
J Bioproces Biotechniq
ISSN:2155-9821 JBPBT, an open access journal Volume 2 • Issue 1 • 1000111
(v) heterotrophic bacterial mediation [1]. Most of the mentioned pro-
cesses above are mediated by microorganism. Both photosynthetic and
heterotrophic microorganisms have natural ability to induce the pre-
cipitation of calcium carbonate. ere are large amount of microor-
ganism in many type of species spreads throughout the world. Table 2
shows several species which is already investigated as media in calcium
carbonate precipitation [6].
In biocementation, microorganism that used as media should meet
the specic requirement, since the process create a high pH in the en-
vironment and involving high concentration of calcium ion. For ex-
ample, in biocementation based on urea hydrolysis, the process will
produce high concentration of ammonium and not all type of micro-
organism can survive in such condition. Based on that, the selected of
microorganism should meet the criteria such as: (i) have a high urease
enzyme activity; (ii) ammonium and calcium ion tolerable; (iii) not
pathogenic [7].
Feasibility of Using Microalgae in Biocementation
Microalgae are a promising media to be used in biocementation,
due to its photosynthetic metabolism. Algae’s species like Spirulina,
Arthrospira plantensis (Cyanophyta), Chlorella vulgaris (Chlorophyta),
Dunaliella salina, Haematococcus pluvialis, Muriellopsis sp., Porphyrid-
ium cruentum (Rhodophyta) basically are autotrophic microorganisms
that live through photosynthetic process [14-16].
Experiment of nine green algae, a diatom and three cyanobacteria
were shown to precipitate CaCO3 in batch culture, where grown in the
light in a hard water medium containing 68 mg L−1 soluble calcium.
e composition of the medium was based on that found in natural
marine hard water where precipitation of CaCO3 within algal biolms
occurred. Deposition occurred as a direct result of photosynthesis
which caused an increase in the pH of the medium. Once a critical pH
had been reached, typically approximately pH 9.0, precipitation be-
gan evidenced by a fall in the concentration of soluble calcium in the
medium [17]. In other experiment, Synechococcus cyanobacteria, the
eukaryotic Mychonastes sp., and Chlorella sp., were found to induce the
precipitation of CaCO3 [18]. In all experiments the precipitation pro-
cess developed in three stages: (1) a pH-dri period, (2) the actual pre-
cipitation reaction, and (3) an equilibration phase. e time intervals
of the stages as well as the concentration changes found in the work
were comparable to the results of other experimental studies on CaCO3
precipitation by algae as shown in table 3 and gure 3 [18].
Several types of microalgae also use urea hydrolysis mechanism to
full the needs of nitrogen. For example, Chorella sp utilizes urea as a
nitrogen source; urea is hydrolysed by urease or urea amidolyase en-
zyme to produce ammonia and bicarbonate [19]. e activity of urease
enzyme also can induce the precipitation of calcium carbonate [11,12].
Figure 2: Injection method of cementation liquid (contain calcium/urea solu-
tion and bacterial cell) in biocementation [4,7].
(a)
(b)
Acc.v Spot Magn Det WD Exp
20.0kV 4.0 250x SE 10.2 599 EAWAG
Acc.v Spot Magn WD
12.0kV 2.0 12000x 100 FAWAG
10µm
2µm
Figure 3: (a) SEM photograph of carbonate precipitates in presence of eu-
karyotic picoplankton, holes in the carbonate structure correspond to pico-
plankton cells and (b) picocyanobacteria [18].
Application Microorganism Metabolism Solution Reference
Biological
mortar
Bacillus cereus oxidative
deamination of
amino acids
Growth media
(peptone, extract
yeast, KNO3, NaCl)
+ CaCl2.2H2O, Acti-
cal, Natamycine
[2]
Crack in
concrete
remediation
Bacillus pasteurii
Bacillus sphaeri-
cus
Hydrolysis of
urea
Hydrolysis of
urea
Nutrient broth, urea,
CaCl2.2H2O, NH4Cl,
NaHCO3
Extract yeast, urea,
CaCl2.2H2O
[13]
[9]
Bacterial
concrete
Bacillus pasteurii Hydrolysis of
urea
Nutrient broth, urea,
CaCl2.2H2O, NH4Cl,
NaHCO3
[13]
Table 1: Overview of various construction materials made from biocementation.

Citation: Ariyanti D, Handayani NA, Hadiyanto (2012) Feasibility of Using Microalgae for Biocement Production through Biocementation. J Bioprocess
Biotechniq 2:111 doi: 10.4172/2155-9821.1000111
Page 4 of 4
J Bioproces Biotechniq
ISSN:2155-9821 JBPBT, an open access journal Volume 2 • Issue 1 • 1000111
ere are some advantages of using microalgae as media for bioce-
ment production. Microalgae are type of renewable resources that eas-
ily cultivated rather than other type of microbe such as bacteria which
already proved to be used in biocementation, so that its availability as
raw material can be maintained properly. It’s easy to grow especially in
tropical area, where many non-agricultural landlls can be utilized as
a raceway pond for microalgae cultivation. Tropical country also has a
good temperature and water with high mineral contained which is very
suitable for microalgae cultivation [15]. Another advantage is that the
biocement production using microalgae can reduce the CO2 emission,
which produced in conventional cement production [5,3].
Based on table 3, the microalga is able to precipitate calcite very
eectively within a couple days [18], while using bacteria such as Spo-
rosarcina pasteurii is able to precipitate calcite under certain condition
within 24 hours. But yet the exact data of experiment and literature still
lack for the microalgae carbonate precipitation.
Future Challenge
Biocement is product innovation in material eld that can be pro-
duce naturally using microorganism such as bacteria and microalgae.
Microalgae have a great potential to be developed as media for bio-
cement production through biocementation. Microalgae metabolism
activity such as photosynthesize and hydrolysing urea can create the
alkaline environment (pH and DIC elevation), so that calcium carbon-
ate precipitation occurs in the presence of calcium ion in the system.
On the other side, microalgae also part of renewable resource that
is easily cultivated especially in tropical area, so that its availability as
raw material can be maintain properly. Further basic research needs to
be done, primary to the theme related to suitable type of microalgae,
mechanism used in biocement production through biocementation,
the kinetics of process, and also the optimum condition to produce
good quality of biocement.
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Type of microorganism System Chrystal type Reference
Photosynthetic organism
Synechococcus GL24
Chlorella
Meromictic lake
Lurcene Lake Calcite CaCO3)
Calcite (CaCO3)
[21]
[18]
Sulfate reducing bacteria
Isolate SRB LVform6 Anoxic hypersaline lagoon Dolomite (Ca(Mg) CO3) -
Nitrogen cycle
Bacillus pasteurii
Bacillus cereus
Urea degradation in synthetic medium
Ammonication and nitrate reduction
Calcite (CaCO3)
Calcite (CaCO3)
[10]
[1]
Table 2: Several species which already investigated as media in calcium carbonate precipitation [6].
Experiment Cell abundance [103cells.ml-1]Chlorophyll [μg.l-1] pH drift time [h] pH at Start of prec. Length of prec. [h] % of Ca2+ precipitation
Mychonastes sp. (1) 13.2 142 45 9.05 50 41
Mychonastes sp. (2) 22.9 448 18 9.20 30 34
Chlorella sp. (1) 6.85 222 25 9.00 10 26
Chlorella sp. (2) 8.71 379 11 8.95 4 29
Synechococcus (1) 33.4 130 40 8.95 40 13
Synechococcus (2) 94.1 324 30 9.05 8 32
Table 3: Precipitation experiments of CaCO3 induced by several types of algae [18].