Cement and Concrete Research 56 (2014) 139–152

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Cement and Concrete Research

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Self-healing concrete by use of microencapsulated bacterial spores

J.Y. Wang a,b, H. Soens c, W. Verstraete b, N. De Belie a,⁎
a
Magnel Laboratory for Concrete Research, Faculty of Engineering and Architecture, Ghent University, TechnologieparkZwijnaarde 904, B-9052 Ghent, Belgium
b
Laboratory of Microbial Ecology and Technology (LabMET), Faculty of Bioscience Engineering, Ghent University, Coupure Links 653, B-9000 Ghent, Belgium
c
Devan Chemicals NV, Klein Frankrijk 18, 9600 Ronse, Belgium

a r t i c l e i n f o a b s t r a c t

Article history: Microcapsules were applied to encapsulate bacterial spores for self-healing concrete. The viability of encapsulated
Received 27 March 2013 spores and the influence of microcapsules on mortar specimens were investigated first. Breakage of the microcap-
Accepted 27 November 2013 sules upon cracking was verified by Scanning Electron Microscopy. Self-healing capacity was evaluated by crack
healing ratio and the water permeability. The results indicated that the healing ratio in the specimens with bio-
Keywords:
microcapsules was higher (48%–80%) than in those without bacteria (18%–50%). The maximum crack width healed
Crack (B)
Water permeability (C)
in the specimens of the bacteria series was 970 μm, about 4 times that of the non-bacteria series (max 250 μm). The
Organic materials (D) (microcapsules) overall water permeability in the bacteria series was about 10 times lower than that in non-bacteria series. Wet–dry
Microbial CaCO3 (D) cycles were found to stimulate self-healing in mortar specimens with encapsulated bacteria. No self-healing was
Self-healing observed in all specimens stored at 95%RH, indicating that the presence of liquid water is an essential component
for self-healing.
© 2013 Elsevier Ltd. All rights reserved.

1. Introduction Due to the harsh environment inside the concrete, encapsulation or

Self-healing systems with microencapsulated healing agents, have
been developed mainly in polymers and composites [1–4]. The research
on the application of microcapsules into concrete to obtain a self-healing
capacity was started recently. Yang et al. [5] showed that the addition of
microencapsulated methylmethacrylate based healing agent into carbon
microfiber-reinforced mortar can greatly improve the crack resistance
and toughness under fatigue loading. This demonstrated that the con-
cept of using microcapsules as carriers for self-healing agents in concrete
was feasible and very promising.
Microbial CaCO3 is regarded as environmentally friendly and eco-
nomical material which has a promising potential for a wide range of
engineering applications, for instance, consolidation and protection of
concrete and stone surfaces, repair of defects and flaws (specifically,
cracks), cementation or consolidation of loose particles (specifically,
soil and sands), etc. [6–13]. The basic principle of applying microbial
CaCO3 for self-healing concrete is that bacteria and other relevant
agents are added into the concrete matrix during casting. When cracks
appear, bacteria around the crack surfaces will be activated (by moisture,
O2, etc.) and precipitate CaCO3 to heal the cracks. In this study, microbial
CaCO3 was precipitated by Bacillus sphaericus through urease catalyzed
urea hydrolysis [14].
⁎ Corresponding author at: Magnel Laboratory for Concrete Research, Faculty of
Engineering and Architecture, Ghent University, TechnologieparkZwijnaarde 904,
B-9052 Ghent, Belgium. Tel.: + 32 9 264 59 22; fax: + 32 9 264 58 45.
E-mail address: nele.debelie@ugent.be (N. De Belie).
immobilization of bacteria in a protective carrier before adding them to
the concrete is preferable [15,16]. A self-healing system by using porous
expanded clay particles for immobilization has been described [17,18].
Healing superiority in the specimens with bio-agents was observed
after 40 days; the maximum crack width healed reached 0.46 mm,
which was almost two times that in the reference specimens. In our ear-
lier work, we developed a system in which glass capillaries were used to
encapsulate bacteria and filling materials (polyurethane or silica sol gel)
and found that the water permeability in the bacterial series was 2–3
orders of magnitude lower than the non-bacteria series [11]. Addition-
ally, diatomaceous earth has also been explored to immobilize bacteria
for self-healing. Cracks of a width of 0.17 mm were completely healed
in the specimens in which diatomaceous earth immobilized bacteria
was incorporated, while no crack healing was observed in the non-
bacterial series [19]. In these experiments, the cracks were created
in the mortar specimens at the age of 14 days, and the healing efficiency
was analyzed after a healing period of 40 days.
In this study, microcapsules were used as bacterial carriers. The mi-
crocapsules were resistant to the high pH of concrete and humidity sen-
sitive. They are flexible under high humidity (like in water) and become
brittle at low humidity. This means that the capsules can withstand the
mixing process and are easily broken when cracks appear. Upon crack-
ing, microcapsules in crack zone will break. In the presence of water in
the cracks, spores in the broken capsules can germinate and precipitate
CaCO3 to heal the cracks. The aim of this research was to demonstrate
the feasibility of using microencapsulated spores to self-heal cracks in
a cementitious matrix. The food of bacterial spores (yeast extract) and
the deposition agents including urea and Ca source (Ca-nitrate) were

0008-8846/$ – see front matter © 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.cemconres.2013.11.009
140 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152
incorporated together with the microcapsules during the mixing
process.
2. Materials and methods
2.1. Bacterial strain
Bacillus sphaericus LMG 22557 (Belgian coordinated collection of mi-
croorganisms, Ghent) was used in this study. B. sphaericus spores were
produced in the liquid minimal basal salts (MBS) medium [20]. The
MBS medium was autoclaved at 120 °C for 20 min before use. Mature
spores were transferred as inoculum (1%) into MBS medium. The cultures
were incubated (28 °C, 100 rpm) for 28 days. Subsequently, they were
subjected to pasteurization (80 °C for 20 min, then 5 min in ice-cold
water) to minimize the vegetative cells. Spores were then harvested by
centrifuging the culture (7000 rpm, 4 °C, Eppendorf MiniSpin, Hamburg,
Germany) for 7 min. The supernatant was removed and the paste of
spores was subjected to vacuum drying for 3 days under room tempera-
ture. Dry paste of the spores was then ground in a cement mill (Aurec S.A,
Bruxelles) for 5 s to obtain fine powders for encapsulation.
2.2. Microencapsulation of the bacterial spores
Bacterial spores were encapsulated following a patented poly-
condensation reaction based microencapsulation process [21].
The microcapsule is melamine based, and contains inert substance to
protect the spores. The size is about 5 μm. The final product is an emul-
sion consisting of microcapsules and water. The concentration of the
spores in the microcapsules was around 109 cells/g microcapsules
(dry weight). The dry matter content was 38.9% and 51.7% of the emul-
sion containing the microcapsules with and without bacterial spores,
respectively.
2.3. Viability of the microencapsulated spores —Proof of concept 1
The viability of the spores after immobilization into microcapsules
was investigated. In order to investigate whether the spores inside the
microcapsules were still viable and whether the microcapsules can really
protect the spores from being activated before they were broken, a two-
step test (Test 1 and Test 2) was performed. Urea decomposition by
intact and broken microcapsules (loaded with bacterial spores) was
examined. The viability of the spores includes the germination into
vegetative cells (active) and the revival of the ureolytic activity. There-
fore, the amount of urea decomposed by the spores can be used as an
index to evaluate the viability of the spores.
In a first test (Test 1), 2 mL emulsion with intact microcapsules
(with or without bacterial spores loaded) was added to 100 mL sterile
YU medium (consisting of 20 g/L yeast extract and 20 g/L urea). The
amount of urea decomposed in YU medium was measured.
In a second test (Test 2), the viability of the spores from broken mi-
crocapsules was investigated. The shell of the capsules is impermeable
for spores, which means that spores cannot leak out from the capsules.
If the nutrients in the YU medium cannot penetrate through the shell,
spores will not germinate and decompose urea. Microcapsules in emul-
sion (5 mL) were first dried and then were ground in a mortar to break
the capsules. The broken capsules (from 2 to 2.5 mL original emulsion)
were added to 100 mL YU medium. The amount of urea decomposed
was determined.
The above-mentioned experiments were performed in duplicate
(n = 2). The amount of urea decomposed was calculated based on
the total ammonium nitrogen (TAN) measured in the medium [22].
One mole of urea (CO(NH2)2) produces 2 mol of NH+ 4 . The amount of
NH+ 4 can thus indicate the amount of urea decomposed.
2.4. Microcapsules in mortar specimens —proof of concept 2
2.4.1. Influence of the microcapsules on the mechanical properties of mortar
Mortar specimens (40 mm × 40 mm × 160 mm, n = 3) with
different amounts of microcapsules (without bacteria) incorporated,
were cast to investigate the influence of microcapsules on the mechani-
cal properties of mortar specimens. The specimens were made with a
water-to-cement ratio of 0.5 and a sand-to-cement ratio of 3 by using
Ordinary Portland Cement (CEM I 52.5N), sand (DIN EN 196-1 Standard-
ized sand) and tap water. The amount of the microcapsule emulsion
added was based on the microcapsule dry weight content as 0% (R),
1%, 2%, 3%, 4% and 5% of cement (by weight). The microcapsule emulsion
was first mixed with water. Then the mortar mixture was prepared ac-
cording to the standard NBN EN 196-1 [23]. After casting, all molds
were put in an air-conditioned room (20 °C, N95%RH). The specimens
were de-molded after 48 h (24 h for R) and were placed in the same
air-conditioned room. After 28 days, strength was measured according
to the standard NBN EN 196-1 [23].
2.4.2. Survival of the microcapsules during the mortar mixing process
Light microscopy (Leica S8AP0, Switzerland) was used to visualize
the microcapsules before and after they were mixed with mortar
paste. After the mortar paste (containing 1% microcapsules) was ready,
a glass slide was inserted into the paste and was then pulled out. Imme-
diately, the glass slide was subjected to light microscopy to observe the
thin layer of the mortar paste with the remaining microcapsules.
2.4.3. Breakage of the microcapsules upon cracking
Breakage of the microcapsules upon cracking is the prerequisite for
self-healing behavior caused by bacteria. The fracture surfaces of the
specimens (with 1% microcapsules embedded) generated during the
tensile test were subjected to Scanning Electron Microscopy (SEM, FEI
QUANTA 200F) analysis to verify the breakage of the microcapsules.
Dry samples were gold coated by a Baltec SCD030 Sputter Coater before
the SEM examination.
2.4.4. Influence of the microcapsules and nutrients on cement hydration
Except for microcapsules, nutrients that were needed for bio-
precipitation, including the food for bacteria (yeast extract, YE) and depo-
sition agents (urea and Ca-nitrate) were also incorporated. To investigate
the influence of these additives on hydration, hydration heat production
was measured, which can be used as an indicator for hydration degree.
Seven kinds of cement paste mixtures of the same water to cement
ratio (0.5) were made: a reference cement paste R, and cement pastes
with urea (4%), YE (0.35%), YE (0.85%), Ca-nitrate (8%), μ-capsules (3%),
μ-capsules (5%), respectively. The dosage of the additives was versus
cement weight. The hydration heat production (at 20 °C) was deter-
mined by a TAM AIR isothermal heat conduction calorimeter [24].
2.5. Applying microencapsulated spores for self-healing cracks
2.5.1. Preparation of the specimens
Six series of specimens were made and the composition of each se-
ries is shown in Table 1. Group R are the specimens without any addi-
tions. Group N are the specimens with all nutrients needed for bio-
precipitation. Group C are the specimens with the microcapsules (3%)
without bacterial spores. Group NC are the specimens with nutrients
and microcapsules (3%, no bacterial spores loaded). Group NCS3% and
NCS5% are the specimens with nutrients and microencapsulated bacte-
rial spores (3% and 5%, respectively).
In each series, three types of specimens were made, 5 long rein-
forced prisms, 3 normal prisms and 20 cylinders, as summarized in
Table 2. After casting, the molds were placed in the air-conditioned
room (20 °C, N95%RH). The specimens were de-molded after 48 h
(24 h for R) and were placed in the same air-conditioned room until
the time of testing.
 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152 141

Table 1 surface water. The test was done in an air-conditioned room (20 °C,
Composition of the specimens in each series. 60%RH). The water absorption coefficient k (g.cm−2.h−1/2) was deter-
Group Cement Sand Water Nutrients Microcapsule Dry weight of Bacterial mined by Eq. (1).
(g) (g) (g) (g) emulsion (g) microcapsules spores
pffiffi
(g) Q =S ¼ k t ð1Þ
R 450 1350 225 0 0 0 N
N 450 1350 214 57.8 0 0 N Q is the weight of water absorbed at different time intervals (g); S is the
C 450 1350 212.4 0 26.1 13.5 N
area in contact with water (cm2); k is the slope of a plot of water
NC 450 1350 201.4 57.8 26.1 13.5 N
NCS3% 450 1350 192.8 57.8 34.7 13.5 Y
absorbed per square centimeter in function of the square root of time.
NCS5% 450 1350 178.7 57.8 57.8 22.5 Y A vacuum saturation test was also performed. The specimens were
dried in an oven at 60 °C until the weight changes were less than 0.1%
Nutrients included yeast extract, urea and Ca(NO3)2.4H2O. The addition ratio was 0.85%,
4% and 8% of cement by weight. Ca(NO3)2.4H2O contains 30.5 wt.% water. Therefore the at 24 h intervals. The completely dry specimens were then subjected
amount of the mixing water was reduced by the amount of the water in Ca(NO3)2.4H2O to the vacuum saturation test (NBN B 24-213) [26]. The specimens in
and the amount of the water in the microcapsule emulsion. The last column shows dry state were placed in a container and were subjected to vacuum for
whether bacteria were present (Y) or not (N).
3 h and then de-ionized water was added into the container till the
cylinders were completely immersed. The vacuum was maintained
during water addition and lasted 1 h more at constant water level.
2.5.2. Creation of cracks and incubation conditions After the vacuum was stopped, the specimens were kept submersed
28 days after casting, the long reinforced prisms were subjected to a for another 12 h. The final weight of the water saturated specimens
tensile test to create multiple cracks. A uniaxial tensile load was applied was measured. The saturated water absorption was calculated by
to the specimen at a speed of 0.01 mm/s under stroke control. The loading Eq. (2).
was stopped at the point where the average crack width in the specimen
reached 150 μm. The details about determination of stopping point can be Ws ¼ ðWw −Wd Þ=Wd  100% ð2Þ
seen in the supplementary file.
The cylinders were also subjected to a splitting test (Walter&Bai,
250/50, Switzerland) to make cracks after 28 days. The crack width
was controlled by the value of the crack opening measured by the Ws saturated water absorption ratio
attached LVDT. The final crack width in the cylinders was about 0.20– Wd dry weight of the specimen
0.22 mm. Ww wet weight of the water saturated specimen
The cracked specimens were subjected to five incubation conditions:
1) 20 °C, N95%RH; 2) immersion in water; 3) immersion in the deposi-
tion medium (DM); 4) wet–dry cycles with water; 5) wet–dry cycles 2.5.4. Mercury intrusion porosimetry (MIP) test
with DM. DM was composed of 0.2 M urea and 0.2 M Ca(NO3)2. During Pore properties of the specimens were investigated by MIP (PACAL
one wet–dry cycle, the specimens were immersed in water/DM for 16 h 140+440 Series, Thermo Fisher Scientific). The freeze-dried samples
and were then exposed to air for 8 h. The incubation of 2), 3), 4) and 5) (around 5 g) were added to the dilatometer (sample container),
was performed in an air-conditioned room (20 °C, 60%RH). which was then placed in the specific position of the MIP instrument 1
(PACAL 140 Series). The pressure applied was from 0 to 200 kPa. Subse-
2.5.3. Capillary water absorption and vacuum saturation of the quently, the dilatometer was transferred into MIP instrument 2 (PACAL
mortar specimens 440 Series) and was subjected to a high pressure from 0.1 MPa to
After 3 months, two of the three normal prisms (40 mm × 40 mm × 400 MPa, following again the procedure of mercury intrusion and
160 mm) were subjected to the standard strength test [23] to investi- extrusion. Porosity and pore size distribution were obtained.
gate the later age strength of the specimens with additions. The third
prism was cut into 3 cubes (40 mm × 40 mm × 40 mm) for the water 2.5.5. Evaluation of the self-healing efficiency
absorption test (based on the RILEM 25 PEM (II-6) [25]). The cubes Since the self-healing by the aid of microorganisms is mainly due to
were put in the 40 °C oven until the mass changes were less than 0.1% the microbial precipitation of calcium carbonate, measurement of the
at 24 h intervals The four sides of the cubes adjacent to the cut surface healed part of the crack by the deposition and the subsequent regain
were then wrapped in an aluminum tape to prevent water evaporation of water tightness can directly indicate the healing efficiency.
through the sides during the water absorption test. The cubes were
brought into a water bath with a water level of 10 ± 1 mm and the 2.5.5.1. Crack filling. Crack filling was visualized by light microscope
cut surface facing downwards. At regular time intervals, the cubes (Leica S8 APO, Switzerland). The filling efficiency was evaluated by the
were taken out from the water bath and weighed after removing the healing ratio (r), which was the ratio between healed crack area and

Table 2
Information on the specimens prepared in each series.

Shape Size Number Test performed

Type I 30 mm × 30 mm × 360 mm (Reinforcement inside: 5 Multiple-cracking: monitor crack healing

D = 6 mm, L = 660 mm) by light microscopy

Type II 40 mm × 40 mm × 160 mm 3 Two for strength test;

one for water absorption test

Type III D = 78 mm, h = 22 mm (with two steel fibers inside) 20 Water permeability test
142 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152

initial crack area. The initial images of the cracks in the specimens were
taken immediately after multiple cracking. During incubation, the spec-
imens were subjected to light microscopy every week in the first month
and at the end of the second month. The values of the initial and final
crack area in the images were determined by a Leica image analysis
program. The morphology of bio-CaCO3 precipitated in the cracks was
studied by SEM analysis.

2.5.5.2. Water permeability test. The cracked cylinders were in different

incubation conditions for 8 weeks and were subjected to the water per-
meability test. The cylinders stored under the condition of 95%RH were
not subjected to the water permeability test because no visual healing
occurred. The detailed description of the test process and the calculation
Fig. 2. Strength properties of the specimens (with different additions of microcap-
of the water permeability coefficient (k) can be seen in the previous
sules, 0% (R), 1%, 2%, 3%, 4% and 5% of cement by mass, respectively) at the age of 28 days.
research [11].

2.6. Statistical analysis

Statistical analysis was done with the software SPSS 19.0. A one-way
ANOVA was used to compare the average values. Significant differences
between multiple average values were determined according to a
Student–Newman–Keuls test. The level of significance was 0.05. In
the following graphs and tables, the average values and the standard
deviations are shown.
3. Results
3.1. Proof of concept
3.1.1. Viability of the microencapsulated spores
The viability of the spores after being immobilized into microcap-
sules was evaluated by using the amount of decomposed urea as an
index. A limited amount of urea was decomposed in YU medium with
the addition of intact bio-microcapsules, around 3 g/L after 1 day; no
obvious increase occurred from 1 day to 3 days (Fig. 1). While in the
series of the broken bio-microcapsules, the urea was completely
decomposed after 3 days though only small amount (around 3–4 g/L)
of urea was decomposed in the first day. It should be noted that there
was already small amount of urea decomposed (about 1–2 g/L) in the
medium before the addition of microcapsules. This is due to the heating
process during sterilization of the medium. In addition, during the en-
capsulation process, a limited amount of spores were not encapsulated
into capsules but stayed in the emulsion. These free spores germinated
and decomposed a limited amount of urea in the first day. The spores
inside the capsules could only start to germinate when the capsules
were broken and hence they could reach the nutrients. Spores need
time to transform from dormant state to activate state. Therefore, only
25
1d
3d
Urea decomposed (g/L)
after 3 days, the urea was completely decomposed. According to the
above results, it can be concluded that the spores were still viable
after immobilization into the microcapsules. Furthermore, only when
the capsules are broken, the spores can germinate and decompose urea.
No urea was decomposed in the series of the microcapsules (intact
and broken) without spores loaded (data not shown).
3.1.2. Influence of the microcapsules on the mechanical properties of mortar
The addition of microcapsules did not significantly affect the volume
density (data not shown) but had a negative effect on the mechanical
properties of the specimens. As shown in Fig. 2, the tensile strength
was decreased significantly (at a confidence level of 0.05) only for a
dosage of microcapsules higher than 3%.
The addition of microcapsules had a more negative effect on the
compressive strength of the specimens. As shown in Fig. 2, the compres-
sive strength was dramatically decreased by 15% to 34% with increasing
addition of 1% to 5% microcapsules.
The mechanical properties of the specimens with both microcap-
sules and nutrients were investigated to see the combined effect of the
bio-healing agents. In this case, the strength at a later age (90 days)
was tested. The results are shown in Fig. 3. According to the statistical
analysis, no significant difference was observed in tensile strength of
the specimens in different series. Regarding the compressive strength,
the minimum decrease was noticed in the series of N, around 9%.
Other series with microcapsules showed distinct decrease, varying
from 22% (NC) to 47% (NCS5%). It can be seen that the addition of
nutrients had less effect on the long-term strength than the addition
of microcapsules. The decrease of the strength was therefore mainly
due to the addition of microcapsules.
3.1.3. Survival of the microcapsules during mortar mixing
The microcapsule emulsion was first mixed with water; the distribu-
tion of the microcapsules in water is shown in Fig. 4(a). It can be seen

20

15

10

0
Bacteria loaded Bacteria loaded
microcapsules (unbroken) microcapsules (broken)

Fig. 1. Urea decomposed in YU medium after the addition of the bio-microcapsules. Fig. 3. Mechanical properties of the specimens at the age of 3 months.
 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152 143

Fig. 4. Microcapsules (without spores) in the emulsion (a) and mixed with mortar paste (b).

that the microcapsules have a regular round shape. The microcapsule
solution was then mixed with other components of mortar; the image
of the final mortar paste is shown in Fig. 4(b). It was noticed that
quite some light round dots adhered to the cement or sand particles
and distributed all over the mortar paste. Comparing the images of the
microcapsules before and after being mixed in mortar, it can be deduced
that these dots were those unbroken microcapsules.
3.1.4. Breakage of the microcapsules upon cracking
The microcapsules were broken under tensile force. As shown in
Fig. 5, at the fracture surface, many microcapsules were broken and
capsule shells still remained in the matrix, which indicated a good
bond strength between microcapsules and the mortar matrix. It is also
seen from Fig. 5 that the size of the microcapsules is in the range of
2–5 μm.
Fig. 5. Fracture surface of the mortar specimen with microcapsules (3%) embedded
(a: 5000 ×; b: 20,000 ×).
3.2. Influence of the microcapsules on water absorption of the
mortar specimens
Although the addition of nutrients and microcapsules had a negative
effect on the mechanical properties, they had a positive effect on
decreasing capillary water absorption of the specimens. Compared
with the R series, the decrease of water absorption in the specimens
15 400
Cumulative heat production (J/g)

12
Heat production rate (J/gh)

300
3.1.5. Influence of the microcapsules and nutrients on cement hydration
The heat production rate and cumulative heat production of each R
9
kind of cement pastes are shown in Fig. 6. It can be seen that the urea(4%)
YE(0.35%)
Ca-nitrate can accelerate cement hydration. 0.35% YE, urea and 3% YE(0.85%)
200
μ-capsules delayed somewhat the appearance of the second hydra- 6 Ca-nitrate(8%)

tion peak. When the dosage of YE reached 0.85% and the dosage of µ-C3%

μ-capsules reached 5%, more delay happened. However, it was found

µ-C5%
100
that the cumulative heat production after 7 days was quite similar for 3
R, urea(4%), YE(0.35%), μ-capsule 3% and μ-capsule 5%: in the range of
355–364 J/g. The YE(0.85%) had somewhat lower heat production of
342 J/g, indicating a lower hydration degree. It was also noticed that 0 0
0 1440 2880 4320 5760 7200 8640 10080
Ca-nitrate not only accelerated hydration but also had a positive effect Time (min)
on the hydration degree. The heat production after 7 days was about
397 J/g, which was about 9% higher than for the R. Fig. 6. Influence of different additives on the isothermal heat production (20 °C).
144
0.3
0.25
Water absorbed (g/cm2)
0.2
0.15
0.1
0.05
0
0 12 24 36 48
Time (h)
Fig. 7. Water absorption of the specimens with or without microcapsules.
with only nutrients (series N), with nutrients and pure microcapsules
(series C and NC), with nutrients and bacteria loaded microcapsules
was around 14%, 42% and 48%, respectively, after 72 h (Fig. 7). The series
with microcapsules, which were loaded with or without bacteria, all
showed a large decrease in the amount of water absorption (more
than 40%) and in the water absorption rate. The specimens with bio-
microcapsules (with encapsulated spores) showed slightly lower water
absorption than the ones with pure microcapsules. No significant differ-
ence was observed between series C and NC, and between NCS3% and
NCS5%.
The result of saturated water absorption is summarized in Table 3.
The R series had the highest water absorption and the specimens with
nutrients and microencapsulated bacteria showed the lowest water
absorption. No significant difference was noticed between the series C
and NC, and between series NCS3% and NCS5%. Series N had a slightly
lower water absorption than Series R. Water absorption after vacuum
saturation by the specimens with bio-microcapsules was also decreased
in the range of 20% to 30%.
3.3. Pore properties of the specimens with and without microcapsules
Fig. 8 shows the cumulated mercury intrusion into the samples as a
function of the pressure applied. It is shown that graphs of R and N have
a similar steep part and the graphs of NC, NCS3% and NCS5% are quite
close to each other. The ultimate amount of intruded mercury was
almost the same in these series, except for NCS5%, in which an extra
abrupt increase of mercury intrusion occurred at the pressure around
350 MPa, resulting in a higher amount of mercury intrusion. The sample
of series C had much more mercury intruded than the samples in other
series and the intrusion curve was quite different. The ultimate mercury
intruded per unit mass directly relates with the porosity of the matrix.
Therefore, it can be concluded that the specimens in series C had higher
porosity than the ones in other series, which had almost the same
porosity.
The specimens with microcapsules incorporated (with and without
bacteria) had different pore size diameter distribution compared to
the ones without microcapsules. In series R and N, most of the pores
were around 0.1 μm (the peaks in Fig. 9(a) and (b)). The sample in
Table 3
Saturated water absorption of the specimens in different series.
R N
Saturated water absorption Ww (%) 5.54 ± 0.14
J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152
series N also had some larger pores around 0.3–0.4 μm. Differently,
the sample in series C had more heterogeneous pore size distribution,
with a large amount of big pores around 2 μm and small pores (about
0.02 μm). Samples in series NC, NCS3% and NC5% had a similar pore
size distribution: most of the pores were in the range of 0.04–0.05 μm.
There was also a small peak at the position of pore size 1–2 μm, which
indicated that there were some large pores (1–2 μm) in these samples,
similarly with the ones in series C. Furthermore, nano-pores with a size
around 4–5 nm were found in NC and NCS5%.
In summary, there are two main categories of pores in the samples
R NC with microcapsules, i.e., large pores (around 1–2 μm) and small pores
(0.02–0.05 μm). The samples in R and N had a more homogeneous
N NCS3%
pore size distribution: the pore size was mainly in the range of
C NCS5% 0.1–0.3 μm.
60 72 84
3.4. Self-healing efficiency
3.4.1. Quantification of self-healing efficiency by microscopy
The development of the crack healing process can be seen from the
microscopic images of cracks taken at certain time intervals (0 day,
1 week, 2 weeks, etc). An example is shown in Fig. 10. It was clearly
observed that the crack area gradually decreased as time went on. By
3 weeks, the crack was almost completely healed.
The cumulative healed crack area in each specimen after 8 weeks
can be calculated based on its total initial and total final crack area,
which is shown in Fig. 11. Although the same methodology was applied
to create cracks, the cracking behavior was different due to different
mechanical properties of the specimens. Crack numbers varied on the
specimens and the crack widths varied from 0.05 mm to 1 mm,
resulting in different initial crack area.
As shown in Fig. 11, the crack area was decreased after 8 weeks in all
specimens except those incubated in the air-conditioned room with 95%
RH (20 °C), in which no obvious healing was visualized under light mi-
croscopy. It was also noticed that the crack area in all specimens was not
completely healed. The minimum value of the unhealed crack area was
around 20 mm2, in the specimen of series NCS3%, which was subjected
to wet–dry cycles. Crack healing ratio gave an overall estimation of the
healing efficiency in the specimens (Fig. 12).
Crack healing was observed in all specimens except the ones stored
at 95% RH. Without the presence of liquid water, the healing ratio in all
specimens was zero. The specimens subjected to full submersion or
wet–dry cycles, all showed certain amount of crack healing. The speci-
mens without microencapsulated bacteria (series R, N, C and NC) had
a healing ratio of 18% to 50%. No significant difference in the overall
healing ratio among different series (R, N, C and NC). Yet, the specific
healing ratio of each specimen in the same series varied when subjected
to different incubation conditions. In the R series, higher healing ratios
were observed in the specimens which were subjected to the conditions
with the deposition medium. In the series N, C and NC, higher healing
ratios were obtained when water was used for submersion. When
using water as the solution, the healing ratios were higher in the speci-
mens (except series C) under full immersion than in wet–dry cycles,
while using the deposition medium as the solution, the opposite result
was observed. The healing ratio of the specimens subjected to the con-
ditions using water as the solution was higher than those using medium
as the solution.
The specimens with microencapsulated bacteria had a much higher
healing ratio, which ranged from 48% to 80%. In view of the overall
C NC NCS3% NCS5%
5.32 ± 0.19 5.22 ± 0.17 5.19 ± 0.21 4.29 ± 0.25 3.85 ± 0.15
 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152 145

0,07

0,06

0,05

Cumulated mercury
volume (ml/g)
R
0,04
N
C
0,03
NC

0,02 NCS3%
NCS5%

0,01

0
0,001 0,01 0,1 1 10 100 1000
Pressure (MPa)

Fig. 8. Mercury intrusion into different samples.

healing ratios, there was no significant difference between the series of the solution. It was noticed that the largest amount of healed crack
NCS3% and NCS5%. The highest value (80%) was obtained in the speci- area (about 80 mm2) was also obtained in the same specimen. The
men of NCS3%, which was subjected to wet–dry cycles with water as healing ratios, in the specimens subjected to the conditions of wet–

a) R b) N
200 200
180 180
160 160
140 140
dV/dlogD

dV/dlogD

120 120
100 100
80 80
60 60
40 40
20 20
0 0
1000 100 10 1 0.1 0.01 0.001 1000 100 10 1 0.1 0.01 0.001
Pore size (um) Pore size (um)

c) C d) NC
200 200
180 180
160 160
140 140
dV/dlogD
dV/dlogD

120 120
100 100
80 80
60 60
40 40
20 20
0 0
1000 100 10 1 0.1 0.01 0.001 1000 100 10 1 0.1 0.01 0.001
Pore size (um) Pore size (um)

e) NCS3% f) NCS5%
200 200
180 180
160 160
140 140
dV/dlogD

dV/dlogD

120 120
100 100
80 80
60 60
40 40
20 20
0 0
1000 100 10 1 0.1 0.01 0.001 1000 100 10 1 0.1 0.01 0.001
Pore size (um) Pore size (um)

Fig. 9. Pore size distribution in the samples from different series of specimens.
146 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152
0d
2 weeks
Fig. 10. An example of a crack healing process (specimen in series NCS5% in ‘wd + water’).
dry cycles with water, were higher than those stored at other incubation
conditions. The specimens that were kept immersed in the deposition
medium had lower healing ratios compared with those in water.
It can be seen that the overall healing ratios of the specimens in
bacterial series were about 30% higher than those non-bacterial series.
This improved healing efficiency was most probably due to the bacterially
induced carbonate precipitation.
Table 4 summarizes the values of the maximum crack widths healed
in different specimens. The specific microscopy images showing the
maximum healed crack widths can be seen in the supplementary file.
It can be seen that the specimens of bacterial series had much wider
cracks healed than for the reference ones.
Some CaCO3 precipitation (verified by EDS) in the cracks was found
with bacterial indents on the surface (Fig. 13(a)). Some remains of
bacteria were also found in between the particles (indicated by the
white arrow in Fig. 13(b)).
3.4.2. Water permeability
The water permeability of the cracked cylinders after the incubation
under different conditions for 8 weeks is shown in Fig. 14. It can be seen
that during the 30 day testing period, the water permeability coefficient
k gradually reached a stable value. There was only a slight difference
between the values at 0 day and 30 days in most cylinders, which indi-
cated that no additional healing occurred during the testing period. In
the series of R, N and C, the k values at 0 day were all in the range of
10−6 to 10− 5 m/s. The series NC had slightly lower initial k values,
around 10−6 to 4.7*10−6 m/s. In the series NCS3% and NCS5%, the initial
k values varied from 2.2*10 − 7 to 4.8*10 − 6 m/s, and 2.0*10 − 7 to
2.0*10− 6 m/s, respectively. The specimens subjected to wet–dry cycles
with water had the lowest water permeability (the blue dash-dotted
curves in chart NCS3% and NCS5%). It was found that the k values after
15 days were almost stable. In this study, the final water permeability
of the specimens was represented by the average k-value from days 15
to 30 (Fig. 15).
In the series R, the cylinders which were subjected to the incubation
with DM, both full immersion and wet–dry cycles, had lower water
1 week
3 weeks
permeability than those in the conditions with water. In series N, the
cylinders in ‘wd + water’ had a lower water permeability than the
ones in ‘wd + medium’ and the one in ‘in water’. In the series C, the
lowest water permeability was observed in the specimens in the condi-
tion of ‘in medium’. Compared with the series R, N and C, the series NC
had a generally lower water permeability. In the series NCS3%, speci-
mens had a large variation of k-values. The lowest water permeability
was observed in the specimens in the condition of ‘wd + water’. Com-
pared with the series NCS3%, specimens in NCS5% had less variation in
k-values within the replicates. As shown in Fig. 14, most of the
k-values were in the zone of 10− 6 and 10 − 7 m/s. Therefore, the
overall final water permeability of the specimens in series NCS5%
was relatively lower compared with the ones in other series. As in
the series NC and NCS3%, the lowest k-values were observed in the
specimens subjected to wet–dry cycles with water, in the range of
6.4*10− 8 m/s to 1.6*10− 7 m/s.
4. Discussion
4.1. Influence of the addition of microcapsules on the properties of
mortar specimens
To engineer the self-healing capacity inside the mortar specimens by
the aid of bacteria, nutrients needed to be incorporated inside the speci-
mens. The best option would be to immobilize spores and the relevant
nutrients in one capsule, which could greatly facilitate spores germina-
tion under suitable conditions (water/high humidity, O2) since they are
easily reachable for spores. However, due to the technical difficulties of
encapsulating water soluble agents into microcapsules, this option was
not feasible for the moment. Therefore, the nutrients were added to the
matrix by mixing together with the mortar paste. The TAM air test results
show that Ca-nitrate has a positive effect on cement hydration while
yeast extract delays hydration and decreases hydration degree if the
dosage is higher than 0.85%. Similar results were also reported by
Bolobova and Kondrashchenko [27]. Small dosages of carbohydrates or
amino acids can enhance the plasticizing effect and accelerate hardening
 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152 147

a) R b) N
160 Initial crack area 160 Initial crack area
Final crack area Final crack area
140 140

Crack area (mm2)

Crack area (mm2)

120 120
100 100
80 80
60 60
40 40
20 20
0 0
95% RH in water in medium wd+water wd+medium 95% RH in water in medium wd+water wd+medium

c) C d) NC
160 Initial crack area 160 Initial crack area
Final crack area Final crack area
140 140
Crack area (mm2)

Crack area (mm2)

120 120
100 100
80 80
60 60
40 40
20 20
0 0
95% RH in water in medium wd+water wd+medium 95% RH in water in medium wd+water wd+medium

e) NCS3% f) NCS5%
Initial crack area Initial crack area
160 160
Final crack area Final crack area
140 140
Crack area (mm2)

Crack area (mm2)

120 120
100 100
80 80
60 60
40 40
20 20
0 0
95% RH in water in medium wd+water wd+medium 95% RH in water in medium wd+water wd+medium

Fig. 11. Initial and final (after 8 weeks) crack area in the specimens of different series (a–f) under different incubation conditions.

simultaneously. However, if the dosage is larger than 0.5%, inhibition of
cement hydration will happen. The inhibition effect from organic com-
pounds is mainly because they are easily be absorbed on the surface of
mineral particles, thereby screening the contact of cement with water,
and hence decrease cement hydration. Therefore, the dosage of yeast
extract used in this research is 0.85% of cement by mass. The limited neg-
ative effect on the hydration degree from YE can be compensated by the
positive effect from Ca-nitrate.
The addition of microcapsules had dual effects on the mortar matrix.
On the one hand, it had a negative effect on the compressive strength.
100%
80%
Healing ratio
On the other hand, the addition of microcapsules decreased the water
absorption. Strength is influenced by the composition and the micro-
structure of the specimen. Factors such as water to cement ratio, aggre-
gates grading, age and curing condition, admixtures, can influence
strength because they will influence the hydration degree and micro-
structure of the specimen. The more hydration, the more pore space
will be taken by solid phase, hence less porosity and higher strength
will result. Therefore, microstructure is the ‘root’ factor influencing
strength. In this study, the specimens were made with the same cement,
sand and water, using the same w/c ratio and sand to cement ratio. The
only difference was that extra healing agents were added in some of the
mixes, which consisted of nutrients and microencapsulated bacteria.
The strength decreased after the specimens were incorporated with
these agents, which means the microstructure was changed. The changes
in microstructure could be due to a different hydration degree, or due to
different arrangement (or distribution) of the components, or both. Ac-

95% RH
60% cording to the TAM air test results, it can be deduced that the hydration
in water
in medium
40%
wd+water
Table 4
wd+medium
20% Maximum crack widths healed in the specimens of series R and NCS5%.

In water (μm) In medium (μm) wd + water wd + medium

0% (μm) (μm)
R N C NC NCS3% NCS5%
R 240–250 170 75 220–230
NCS5% 850–970 330–400 540–600 280–290
Fig. 12. Healing ratio in each specimen under different incubation conditions.
148 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152
a with microcapsules had a lower strength than the reference ones.
b Furthermore, some functional additives such as water reducing admix-
Fig. 13. Precipitation in the cracks of the specimen NCS5% under the condition of
‘wd + water’ (a: CaCO3 precipitation with bacterial traces on the surface; b: bacterial
remains in between).
degree after 28 days in the specimens was similar. Therefore, the micro-
structure was modified mainly due to the presence of microcapsules.
In view of the water absorption results, it can be concluded that the
open porosity was decreased after the addition of microcapsules. How-
ever, based on the cumulative mercury intrusion (Fig. 8), this is not the
case. The specimens with microcapsules had similar or even a bit higher
amount mercury intruded, which indicated a similar or a bit higher
porosity. These conflicting results were due to the breakage of the
microcapsules under high pressure during mercury intrusion. After
the microcapsules broke, more space was released (that is the reason
why more small pores (b 0.05 μm) appeared), therefore MIP results
exhibited a higher porosity compared with results obtained from satu-
rated water absorption (original porosity). In addition, the space which
is taken by a microcapsule can still be regarded as a pore or weak point
during strength testing, since it cannot withstand the load from com-
pression (while during the process of water absorption and water
saturation, the microcapsules will stay intact and block the pores). This
is one reason why the lower porosity did not contribute to a higher
strength in the specimens with microcapsules.
Strength is not only influenced by porosity, but also by pore size
distribution, and pore geometry (connectivity, tortuosity). Although
microcapsules took up some pore space, they also caused large pores
of 1–2 μm (see Fig. 9). In general, a pore size of b20 nm is classified as
harmless, while 20–50 nm is thought to be less harmful and 50–
200 nm is harmful. A pore size larger than 200 nm (0.2 μm) is said to
be extremely harmful [28]. This is another reason why the specimens
In conclusion, the addition of microcapsules had no obvious effect on
hydration degree but mainly modified the pore structure of the speci-
mens. They decreased the total pore space resulting in decreased water
absorption. Yet they were weak points (much lower strength compared
with cementitious materials) in the matrix and created more large pores
resulting in a decreased strength. Therefore, the mechanical properties,
especially the compressive strength, were greatly decreased if the addi-
tion amount was higher than 3%.
It is important to investigate the combined effect of the microcap-
sules and nutrients on the mechanical properties of the specimens.
The experimental results showed that more decrease (around 33% to
47%) in the compressive strength occurred with the addition of both
nutrients and microcapsules. Such a drastic decrease of the compressive
strength is not acceptable in practical cases for economic and safety
reasons. Therefore, it is suggested to restrict the addition of bio-
microcapsules to 3% maximum. The compressive strength of NCS3%
of about 55 N/mm 2 is acceptable for many practical applications.
tures can be used to increase the mechanical strength of the concrete in
practical situations.
4.2. Self-healing properties in mortar specimens
4.2.1. Autogeneous healing in the control series
The crack healing ratio in the specimens without microencapsulated
spores was in the range of 18% to 50% in the specimens after 8 weeks
(56 days) under different incubation conditions. The healing that oc-
curred was due to autogenic sources. Cementitious materials have a cer-
tain capacity of autogenous healing depending on the composition of
the matrix and the environment they are exposed to [29,30]. In general,
two main mechanisms are responsible for the autogenous healing:
secondary hydration of the unhydrated cement particles and precipita-
tion of calcium carbonate. Besides that, the swelling of the hydration
products also contributes to the decrease of the crack area.
In early age concrete and high strength concrete, there are still many
unhydrated cement particles in the matrix. Therefore, when there is
water available in the cracks, the unhydrated cement along the crack
wall would start hydration again. The formed products could heal the
cracks. The healing efficiency from this source is greatly dependent on
the age of the concrete, the water to cement ratio (w/c) and the avail-
able water in the cracks. Autogeneous healing from the precipitation
of calcium carbonate could happen in any type of concrete. The CO2
from atmosphere dissolves in water and generates CO2− 3 in the alkaline
environment (the pH of concrete is around 12.5–13). The free dissolved
Ca2 + (from the crack surface) reacts with CO23 − and CaCO3 can be
formed.
The specimens in this study had a w/c of 0.5 and were cracked at the
age of 28 days. Theoretically, complete hydration could be obtained if
the w/c is larger than 0.42 (1 g cement binds about 0.23 g water and
physically absorbs 0.19 g water) in a sealed environment. However, in
practical cases, full hydration of OPC is not possible due to the non-
uniformly distributed water and limited available space for the develop-
ment of hydration products [24]. It has been reported that the hydration
degree of OPC at the w/c of 0.5 is about 74% after 28 months [31]. There-
fore, in our test, the specimens still had unhydrated cement inside the
matrix (at least 26% of the initial amount of cement) at the cracking
age (28 days).
The healing was also due to the precipitation of calcium carbonate.
The amount of CaCO3 precipitation depends on the amount of CO23 −
formed and Ca2 + dissolved, which relates with the concentration of
CO2, the pH in the surroundings and the composition of the matrix.
Ca2 + can be available from Ca(NO3)2, from Ca(OH)2, or from un-
hydrated cement particles. The specimens with the addition of nutrients
had more crack healing under the conditions of ‘in water’ (47%) and
 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152 149

0 5 10 15 20 25 30 0 5 10 15 20 25 30
1.00E-05 1.00E-05

1.00E-06 1.00E-06

1.00E-07 1.00E-07

R N
1.00E-08 1.00E-08

0 5 10 15 20 25 30 0 5 10 15 20 25 30
1.00E-05 1.00E-05

1.00E-06 1.00E-06

1.00E-07 1.00E-07

C NC
1.00E-08 1.00E-08

0 5 10 15 20 25 30 0 5 10 15 20 25 30
1.00E-05 1.00E-05

1.00E-06
1.00E-06

1.00E-07

1.00E-07
1.00E-08

1.00E-09
NCS3% 1.00E-08
NCS5%

In water: ; In medium: ; wd+water: ; wd+medium:

Y-axis: k-value (m/s); X-axis: Time (d)

Fig. 14. Water permeability of the specimens during the test periods.

‘wd + water’ (38%), because of more released Ca2+ (due to Ca(NO3)2)
compared with the R series (30% and 18%, respectively). However,
under the conditions of ‘in medium’ and ‘wd + medium’, the speci-
mens in the series R had more crack area healed (34% and 46%, respec-
tively) than that in the series N (25% and 38%, respectively). Similarly,
series C had higher healing ratios than NC. This is attributed to the fact
that although DM can provide extra Ca2 +, the pH of the medium
(pH = 5.5–6) was lower than for plain water (pH = 6.8–7.2). A higher
pH is more beneficial for the dissolution of CO2 and the subsequent
transformation of CO2 to CO2− 3 , and hence CaCO3 formation. The addi-
tion of microcapsules had an ambiguous effect on autogeneous healing.
Under the conditions of ‘in water’ and ‘wd + water’, the healing effi-
ciency in the microcapsule series was higher than that of the series R.
The healed crack area in the microcapsule series was around 1.6–2.1
times (‘in water’) and 2.7–3.1 times (‘wd + water’) of those in the R
series. The opposite result was obtained under the condition of
‘wd + medium’. Theoretically, microcapsules occupied some space in
the crack wall which would decrease the area of the exposed cementi-
tious matrix, and hence, less Ca2+ could be released. However, on the
other hand, the large pores in the matrix due to the addition of microcap-
sules (based on MIP results) might facilitate the solubilization of Ca2+
and the penetration of CO2−
3 . However, for the specimens with multiple
cracks of the widths from 0.05 mm to 1 mm, cracks, instead of matrix
permeability, became the main issue for water penetration. Since all
specimens showed multiple cracks, with similar crack width ranges
and crack numbers, the permeability in these cracked specimens should
be of the same magnitude. Therefore, the release of Ca2+ mainly depends
on the Ca source and storage condition (with sufficient water or not).
150 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152

4.2.2. Self-healing efficiency by microencapsulated bacteria
The viability test results demonstrated that the spores, after encap-
sulation into microcapsules, can still germinate and revive the ureolytic
activity to decompose urea. The prerequisite is that the microcapsules
need to be broken. Then the spores inside can start to work because
the impermeable capsule shell is no longer intact. In the unbroken cap-
sules, the spores stay dormant since no activators (water, nutrients and
oxygen) are reachable. The cracking triggered the breakage of the
microcapsules and the increased self-healing efficiency was then con-
tributed by the microencapsulated bacteria. When there was water
available in the surroundings, the nutrients embedded in the crack
zone were released and became available for the spores in the broken
capsules. The spores then started to germinate and revive the ureolytic
activity. Urea, released from the crack zone or dissolved in the deposi-
tion medium (if present), was decomposed into CO2− 3 and NH3/NH+ 4
by the germinated bacteria (catalyzed by bacterial urease) under alkaline
pH. When CO2− 3 ions met with Ca2+ ions in the surroundings, CaCO3
2+
formed. The Ca source was derived from the matrix itself, the added
Ca(NO3)2 and the DM (if present). During the incubation period, it was
noticed that the white precipitation existed not only in the cracks, but
also on the surface of the specimens, in the surface pores of the specimens
and on the wall of the incubation containers. But most of the precipitation
was concentrated in the cracks
No significant difference in healing efficiency was observed between
NCS3% and NCS5%. Specimens in series NCS3% had slightly higher crack
healing ratios while the ones in NCS5% had an overall lower water per-
meability. With an increased addition of microencapsulated bacteria,
the healing efficiency was not increased proportionally. This is probably
due to the inert substance that is encapsulated together with the bacteria,
which may give a water-proofing effect to the crack surfaces after release.
This water-proofing effect could slow down the penetration of water into
the matrix around the crack zone to dissolve the nutrients, and the trans-
port of dissolved nutrients to the spores. Hence, with large amount of
broken microcapsules, the increased water-repellent effect may make it
difficult for the spores to reach enough nutrients to germinate and revive
the ureolytic activity. For future optimization of the dosage of the micro-
capsules used, except for mechanical properties, the barrier effect on the
diffusion of water and nutrients due to the inert substance on the crack
surface should also be considered.
4.2.3. Influence of incubation conditions
The incubation condition has great influence on the self-healing effi-
ciency, both on the autogenous healing in non-bacteria series, and on
the engineered self-healing in bacteria series.
Water is the essential element for autogenous healing. It participates
in all relevant reaction processes, like secondary hydration of unhydrated
cement, precipitation of CaCO3 and swelling of hydration products. With-
out water, all these reactions would not happen, and hence, healing
would not occur. Therefore, the specimens, stored in the conditions of
95%RH in which no free water is present in the surroundings, had no
crack area healed, including the ones with microencapsulated bacteria.
This is because bacterial spores also need water to revive and the nutri-
ents in the matrix need to dissolve in water to become available for

in water in medium wd + water wd + medium in water in medium wd + water wd + medium

1.00E-05 1.00E-05

1.00E-06 1.00E-06

1.00E-07 1.00E-07

1.00E-08 1.00E-08

R N
1.00E-09 1.00E-09

in water in medium wd + water wd + medium in water in medium wd + water wd + medium

1.00E-05 1.00E-05

1.00E-06 1.00E-06

1.00E-07 1.00E-07

1.00E-08 1.00E-08

C NC
1.00E-09 1.00E-09

in water in medium wd + water wd + medium in water in medium wd + water wd + medium

1.00E-05 1.00E-05

1.00E-06 1.00E-06

1.00E-07 1.00E-07

1.00E-08 1.00E-08

NCS3% NCS5%
1.00E-09 1.00E-09

Fig. 15. Final water permeability coefficients (k-values) of the specimens in each series (three black dots represent three replicates for each condition).
 J.Y. Wang et al. / Cement and Concrete Research 56 (2014) 139–152
bacteria. Thus, bio-CaCO3 cannot precipitate if no water is present in the
crack zone.
Theoretically, the DM containing 0.2 M urea and 0.2 M Ca(NO3)2 can
provide extra Ca2+ for CaCO3 precipitation and more urea can be avail-
able for bacteria, hence should result in an increased healing efficiency.
However, no significant obvious beneficial effect was obtained when
the specimens were subjected to the conditions with the DM. During
the incubation stage, Ca2+ releases during the dissolution of Ca(OH)2
(CH) in the matrix. CH has very low solubility (1.7 g/L (0.02 M) at
20 °C). The dissolution of CH follows the equilibrium which is shown
in Eq. (3).
2þ
CaðOHÞ2 ↔Ca þ 2OH
The reaction will shift to the left side if the concentration of Ca2+ is
higher than 0.02 M in the solution. When the reaction shifts to the left
side, the amount of OH− will be decreased, and hence, the pH decreases.
For the sequestration of CO2, transformation from CO2 to CO23 −, and
CaCO3 precipitation, an alkaline environment is preferable. If the speci-
mens were kept fully immersed in the DM, although more Ca2+ would
be available, the pH condition will restrict the increase of the CaCO3
precipitation. Therefore, no obvious enhanced healing efficiency was
obtained when using DM as the immersion solution. Due to the pH in-
crease during urea decomposition, the specimens in the bacteria series
had a higher healing efficiency than the ones without bacteria, under
the condition with the DM. Furthermore, Ca2+ will also influence the
germination of bacterial spores and the subsequent ureolytic activity
[32,33]. Therefore, in future research, the optimization of the incubation
solution is needed. The solution should contain a very low concentra-
tion of Ca2+ (b0.02 M).
In many practical cases, no full submersion occurs, while wet–dry
cycles may be present because of subsequent rainy and dry periods, or
because the structures are present in tidal zones. In this study, it was
found that among the specimens with microencapsulated bacteria, the
ones subjected to wet–dry cycles with water had the highest healing
efficiency, both for the aspects of healed crack area, water permeability
and maximum crack width healed. During the wet stage, the specimens
absorbed enough water which can keep the matrix in wet state during
the dry stage. When the specimens were exposed to the atmosphere,
more oxygen becomes available for the bacteria than in the case of con-
tinuous immersion. Furthermore, less immersion could also decrease
the leakage of the nutrients into the incubation solution and the escape
of the bacteria from the crack surfaces. An optimal wet–dry cycle can
promote the diffusion of the nutrients from the internal matrix to the
surficial cracking zone without leaching too much to the bulk solution
and can keep the specimens with sufficient available water for bacterial
activities during the dry state.
5. Conclusions
The enhanced self-healing efficiency in cracked specimens contrib-
uted by microencapsulated bacterial spores was demonstrated based
on the experimental results from light microscopy and the water per-
meability test. The specimens with bacteria had much higher crack
healing ratio compared to the ones without bacteria: 18% to 50% of
crack area healed in non-bacteria series (R, N, C, NC) and 48% to 80%
of crack area was healed in the bacterial series (NCS3% and NCS5%).
The maximum crack width healed in the specimens of the bacteria
series was 970 μm, which was much wider (about 4 times) than that
in the specimens of non-bacteria series (max 250 μm). The overall
water permeability in the bacteria series was also lower than that in
non-bacteria series. The average k-value was decreased around 10
times. Wet–dry cycle treatment is the best incubation condition for
the specimens with bacteria. It was noticed that free water is the essen-
tial component to obtain significant amount of self-healing, both for the
specimens with and without encapsulated bacteria. Further optimization
151
of the dosage of the microcapsules, the composition and pH of the incu-
bation solution and the time interval between wet and dry stage is
needed to improve the healing efficiency. Although it will often not be
possible to change the latter two parameters in a practical situation,
extra supplementary techniques could be applied, such as surface cover-
age by moisture retaining materials, to prolong the wet stage beneficial
for self-healing.
Acknowledgments
The financial support from the Research Foundation Flanders (FWO-
Vlaanderen, grant no. G.0157.08) and Ghent University (a BOF grant) is
ð3Þ
gratefully acknowledged.
Appendix A. Supplementary data
Supplementary data to this article can be found online at http://dx.
doi.org/10.1016/j.cemconres.2013.11.009.
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