Effect of the Opuntia (ficus-indica) cactus mucilage extract in reducing sediment and
bacteria contamination from water.
Kathy Tavakoli
Department of Biological Sciences
Saddleback College
Mission Viejo, California 92692
Although nearly all newly derived water purification methods have improved the water
quality in developing countries, few have been accepted and maintained for long-term use.
Field studies indicate that the most beneficial methods use indigenous resources, as they are
both accessible and accepted by communities they help. In an effort to implement a
material that will meet community needs, three fractions of mucilage gum were extracted
from the Opuntia ficus-indica cactus and tested as flocculation agents against sediment and
bacterial contamination. Column tests containing suspensions of the sediment kaolin
exhibited particle flocculation and settling rates up to 15.0 cm/min with mucilage versus
control settling rates of 1.0 cm/min (p=6.94 x 10-5 ANOVA). Escherichia coli tests displayed
flocculation and improved settling times with mucilage concentrations lower than 5 l and
removal rates between 96 and 91% were observed for high bacteria concentration tests
(>108 cells/ml) (p = 9.5 x 10-4 ANOVA). These results indicate the cactus mucilage extract
can reduce sediment and bacteria from water. This natural material not only displays
water purification abilities, but it is also affordable, renewable and readily available.
Introduction
The United Nations has estimated
that 1.1 billion people lack access to potable
water (Bugbee and Reigel 2008). With so
many people living on the brink of illness, a
great deal of attention has been drawn to
designing and implementing new and
innovative methods of water purification,
particularly in developing countries.
Gradually, the goal of bringing safe water to
the world has developed into a series of
goals
from
educating
(about
the
problem….about the methods..) to finding a
method of purification that will be culturally
accepted and sustainable (Bugbee and
Reigel 2008). In an attempt to circumvent
problems associated with implementing
purification methods based solely on
technology, the Opuntia ficus-indica cactus
(also known as the Nopal or Prickly Pear)
indigenous to Mexico is being tested as a
flocculating
agent
for
waterborne
contaminant removal. Through (a) simple
extraction processes, two fractions of
mucilage gum can be obtained from fresh
cut Opuntia ficus- indica pads including a
Gelling Extract (GE) and Non-Gelling
Extract (NE) (Cook, 2000). Mucilage
consists of up to 55 sugars, mainly
arabinose, galactose, rhamnose, xylose,
glucose, and uronic acids, the percentage of
which varies with mucilage type (Edward
2010). Literature has indicated that these
extracts, particularly the GE, undergo
property alterations including viscosity
changes in the presence of diatomic ions
such as Ca2+ (Edward 2010). (A) University
of Florida scientific team demonstrated that
cactus mucilage is an effective tool for
separating sediments (clay and mud
particles) represented by kaolin, from
deionized
(DI)
water
suspensions.
Buttice (2009) observed,(delete) kaolin
settling rates observed in columns treated
with mucilage and aluminum sulfate
(Al2(SO4)3), a common commercially used
flocculent, were compared and mucilage
was concluded to induce a greater increase
in settling rate (Young 2011). Related tests
also indicate that mucilage extracted from
the Opuntia spp. acts as an efficient
coagulant in surrogate turbid water (Young
2011). In this work, the efficiency of
mucilage to remove kaolin suspended in DIWater was evaluated. Aside from sediment
intrusion, another common problem
associated with drinking water has been
bacterial contamination. Even in more
developed countries where purification and
distribution systems are technologically
advanced and water is closely monitored,
cases of waterborne illnesses caused by
bacteria are occasionally observed (Cook,
2000). In developing countries, issues with
bacterial contamination are more severe.
Escherichia coli, a Gram-positive, sporeforming, non-pathogenic, soil-dwelling rod
(∼1 by 3 μm), was chosen to study bacterial
aggregation with mucilage for its ease of use
and potential as a possible surrogate for
waterborne bacteria with similar structural
characteristics. Data on its effects could
impact the status of this widely used product
and may have influence in water purification
industries.
Material and Method
stir-hot-plate for 30 min. The pH was
adjusted to 2 using 12 mL of 0.1 M HCl
solution,(.)
(T)the
mixture
(was)
centrifuged,(delete) and the supernatant
discarded. The precipitate was re-suspended
in DI water,(delete) and the pH was
increased to 8 using 5 mL of 1 M NaOH.
This suspension was filtered via vacuum
filtration using #41 Whatman filter. The
portion of the original suspension that was
reserved for extraction of NE was mixed
with 200 mL of 1 M NaCl solution and
filtered using #41 Whatman filter and a
vacuum filtration system. The filtrates of
both the NE and GE were separately mixed
with acetone (1:1 v/v), spread upon watchglass dishes and left overnight for(to)
allowing(allow) water to evaporate. The
recovered mucilage was removed and
washed in isopropanol (1:1 v/v), and some
NE (v/v) were mixed with GE mucilage to
make CG (combined- gelling). The mucilage
that spread upon watch-glass dishes,(delete)
dried,(delete) (and was) ground using mortar
and pestle(.) and some of them stored for
further
experiments.(delete)
Unused
mucilage suspension was stored in the
Pads were obtained from Opuntia
ficus-indica
cactus,
originally(delete)
purchased from Crown Valley Market,
located in Crown valley Parkway, Mission
Viejo. This method and procedure is
replicate of the project conducted by
University of Southern Florida, Buttice
(2009) study. The 3 cactus pads (average
weight 252 g) were diced, heated with
Nuova stir-hot-plate for 20 minutes at 85
o
C,(delete) (water bath)(,) and then the
solution was naturalized to pH 7-7.5 with 4
mL of 1 M NaOH. The pads were liquidized
by Braun blender with high speed (range 4)
for 5 minutes. The pH (was) adjusted to 7
using approximately 4 mL of 1 M NaOH
solution and the solid separated using Clay
Adams Compact II Centrifuge and 12 - test
tube placed (6 test tubes at a time) in there
with(at) 3200-RPM speed(delete) for 10
minutes. The precipitate was removed and
used in the extraction of GE and the
supernatant was reserved for the acquisition
of NE. In precipitate, a 50 mM NaOH with
0.75% (w/w) was added until the precipitate
was covered and then stirred with Nuova
refrigerator for future use (and dried
mucilage was stored for future experiments).
The two distinct fractions of mucilage gum
were
studied
for
their
different
characteristics and removal abilities. Simple
test tubes were used to evaluate the
flocculation effect of the mucilage. The test
tube contents (mucilage and DI- water) were
mixed to 10 mL in 15 mL centrifuge tubes,
vortexed and poured in to column arrays,
which were observed over a period of time.
Kaolin (hydrated aluminum silicate)
was ordered online from “Allied Products
Wholesale” via “eBay”. The suspension
with final concentration 25g/500 L were set
up in the Di-water at least 24 h prior to the
run of the experiment to allow kaolin
particles thorough hydration time. In test
tubes tests(delete) of specific concentrations,
kaolin has been observed to from(form) a
clear interface which was read every minute
fro(for) up (to) 60 min, and the column
marker at the interface was recorded for
ate(??) plotting. These plots were truncated
where compression in the column began and
settling rate in cm/min was obtained.
The Escherichia coli were cultured
overnight, with final column cell count 108
cell/mL. The final concentration was
determined with direct counting chamber
and Hemocytometer plate. 8 dilution tubes
were taken, each containing 9.0 mL of
sterile saline. Aseptically diluted 1.0 mL of
a sample of E. coli. Dilution was repeated
for total 10 final tubes each containing 108
concentrations. The final and initial
concentration
read
by
Marienfeld
Hemocytometer plate and Nikon microscope
(For accurate result, count ten (10) of the
1/16 sq. mm squares. Calculate the average
value and multiply by 1.6 X 105 to
determine the number of cells per milliliter).
E.coli did not from a visible interface while
settling. The time when flocs began to
appear as small white flecks in the otherwise
turbid water time that the flocs ceased to fall
was recorded. Box plot were used to
represent settling times, where the bottom of
the box indicates the time when flocs were
no longer falling in the column. The dotted
lines represent the start (lower line) and
completion time (upper line) of the control
columns containing only E.coli. Once the
flocs had completed their descent, a 1 mL
sample was taken from the top of the
column and final cell counts were evaluated
using hemocytometer plate and microscope
microscope.
The
resulting
bacteria
concentration was compared to the initial
and final concentration and a removal rate
was
obtained
base
on
clarity.
Result
Kaolin Removal with Cactus
Mucilage. Kaolin suspension in DI water
was treated with 0, 2 l, 5 l and 10 l
concentrations for each GE (gelling
mucilage), NE (non gelling mucilage) and
CG (combined gelling mucilage), which is
demonstrated two characteristic of the
mucilage
induced
settling
(n=10).
Frist(First) it was observed that as mucilage
concentration increased, for all NE, GE, and
CG, so did the removing(removal) rate of
the kaolin (Figure 2). Initially (without any
mucilage), the relationship between
concentration and settling rate appeared to
be linear (Figure 1). (settling rate of figure 3
is linear too… figure 1 indicates no change)
Kaolin Settling Rate (cm/min)
Figure 1. The effect of the control solution (nonmucilage) on the settling rate of kaolin (50 g/L)
suspended in DI water.
1.2
1
0.8
0.6
0.4
0.2
0
0
5
10
15
Concentration (ppm)
9.2
9.0
8.8
8.6
8.4
8.2
8.0
NE (2 uL)
NE (5 uL)
GE (2uL)
GE (5uL)
NE (10 uL)
Kaolin Settling Rate
(cm/min)
8.0
7.5
7.0
6.5
Kaolin Settling Rate
(cm/min)
6.0
GE (10 uL)
that any differences observed among the
treated suspensions were primarily due to
ion interaction with the mucilage. Base on
the result, it can be indicate(d) the mucilage
extract did reduce the kaolin from water.
There was significant statistically difference
between GE, NE, and CE mucilage in
removing kaolin (p = 6.94 x 10-5 ANOVA).
GE
mucilage
observed
92%
removable(removal of) kaolin sediment
from water in 60 min (15 cm initial to 1.2
cm final), which became a highest
removable(removal from) solution, and after
CG for 86% (15 cm initial to 2.1 cm final),
and NE (15 cm initial to 3.5 cm final) with
79%, each remove kaolin from water
(Figure 3).
9.0
16.0
8.8
14.0
8.6
8.4
8.2
8.0
CG (2 uL)
CG (5 uL)
CG (10 uL)
Figure 2. The effect of the mucilage concentration,
for all NE, GE, and CG, on the settling rate of kaolin
(50 g/L) suspended in DI water. As the concentration
increase, settling rate of kaolin decrease (reducing
rate increase).
Second, it was observed that with no
mucilage treatment, all kaolin suspension
settled at rate close to 15 cm/min, indicating
Kaolin Settling Rate (cm/min)
Kaolin Settling Rate
(cm/min)
9.4
12.0
Average CG(cm/min)
10.0
Average GE (cm/min)
Contorl cm
Average NE (cm/min)
8.0
6.0
4.0
2.0
0.0
0
20
40
60
Time ( min )
Figure 3. Comparing the average Kaolin settling rate
(cm/min) with removing sediments time in each 1
minute, for 60 minutes, with respect to the GE, NE,
CG and control (non-gelling) solutions (n=10). GE
mucilage observed most removable kaolin sediment
from water in 60 minutes (15 cm initial to 1.2 cm
final). There was significant statistically difference
between GE, NE, and CE mucilage in removing
kaolin (p = 6.94 x 10-5 ANOVA).
Escherichia coli Removal with Cactus
Mucilage. Arrange(ment) of mucilage
concentration were tested to evaluate the
effect of mucilage on the settling time. The
settling rate of E. coli with 5 l NE, GE, and
CG were obtained from columns
contacting(containing??) steamed water and
108 (cell/min) diluted E. coli (n =10). From
comparing the initial counting E. coli cells
and final counting, indicated that mucilage
extract did remove E. coli from water. The
difference in initial and final concentration
shows that there is statistically difference
between NE, GE, and CG solutions in
reducing E. coli from water (p = 9.5 x 10-4
ANOVA). Difference concentration in GE
was more 0.29 (cell/min), which indicated
the most removable(removal of) E. coli from
water. At GE concentration of 5 l, signs of
flocculation (14:95 min:sec) occurred faster
than NE (17:34 min:sec) and CG (23:35
min:sec)
mucilage.
Bacteria
removable(removal) rate associated with 5
l GE, NE, and CG were determined to be
96.43% , 94.35%. and 91.24% (Figure 4).
Difference in bacteria concentartion
0.35
0.30
0.25
0.20
0.15
0.10
0.05
0.00
NE (5 uL)
GE (5 uL)
CG (5 uL)
Figure 4. The effects of the mucilage extracts (NE,
GE, CG) to the difference concentration of E. coli
(cell/min) (initial - Final) (n=10). Difference
concentration in GE was more 0.29 (cell/min), which
indicated the most removable(removal of) E. coli
from water. The difference in initial and final
concentration shows that there is statistically
difference between NE, GE, and CG solutions in
reducing E. coli from water (p = 9.5 x 10-4 ANOVA).
Error bars indicate mean ± SEM.
Discussion
An important difference between the
mucilage‘s ability to aggregate kaolin
particle compared to bacteria was that there
existed a concentration where the bacteria
was that(delete or reword) reacted to the
mucilage in positive manner. In columns
contacting kaolin, increases in mucilage
concentration resulted in higher settling
rate,(delete) but no concentrations were
observed to restrict settling as seen in
suspension of E. coli. Also, in columns
containing kaolin, the GE appeared to cause
larger increase in settling rate while the NE
seems to work slightly better as a treatment
for E. coli suspension (be)cause the flocs
that formed appeared as organized, stable,
and larger as those formed in GE, and CG.
The indicated results are equivalent to the
University of Florida studies (2009) studies,
(which) observed that small amount(s) of
mucilage that(delete) is required for
contaminant removal from surrogate waters.
Although in this research, as diatomic ions
are known to affect both mucilage and
promote cell aggregation, CaCl2 was studied
in conjunction and compared with mucilage
as a bacteria removal method in three
different kind of water (Hard/Soft/DI),
which each contain different ions and
Bacillus cereus was tested to displayed
flocculation and improved settling times.
The experiment that(delete) performed in(at)
Saddleback College was base(d) on the
specific sediment (kaolin) and bacteria
(Escherichia
coli),
with
different
concentration in mucilage for kaolin and
same concentration but repeated three times
for bacteria (more accurate result). In
addition
in(delete)
Young
(2011)
demonstrated the use of Non-Gelling (NE)
and Gelling (GE) Extract; mucilage
fractions for removal of sediment in DI
water.(consider rewording last sentence) He
concluded that both mucilage fractions act
faster in sediment removal than the controls
containing no flocculating agent, and
solutions treated with the commonly used
Alum. He suggested the combined gelling
solution for future experimentations. In
Saddleback College experiment, the NE, GE
were testing in addition of CG as new
experiment and the results are new, since it
could find in another research experiment
for result comparison. The result(s)
that(delete) observed here indicated (that)
there is significant difference in effect of the
mucilage extract (NE, GE, and CG ) in
reducing bacteria and kaolin from water.
The result(s) observed the GE solution had
removed more kaolin and bacteria in less
time from water;(.) (C)compare(d) to NE
removed less kaolin and take(took) much
long(er) to (form)flocs. In (the) CG result(s),
it appear(ed) to have (a) removing settling
rate and flocculation time in between GE
and NG,(.) from this(these) results, (i)It
appears the CG solution have(has)
characteristics from both gelling and nongelling, which is part that we observed in
non-experimental cactus pad. Although
these removal rates are high, the level of
bacteria remaining in the columns renders
the water still unsafe to drink. This is due to
the initial cell concentration (108 cell/mL),
which would not be typically observed in
the real world,(delete) but was used to
obtain a visual indicator of flocculation and
removal. Future experimentation will
involve optimizing parameters for bacteria
removal at lower level of contamination.
The results discussed in this work
demonstrate the potential of mucilage
extracted from the Opuntia ficus-indica as
flocculation agent for sediment and bacteria
contamination in ion rich water. The cactus’
prevalence, affordability, and cultural
acceptance make it an attractive natural
material
fro(for)
water
purification
technologies that could be beneficial in
Mexico
and
around
world.
The
difference(different) mucilage fractions have
been observed to provide diversity
both(delete) in their structural features as
well as in their reactions to the natural ion
concentration in water they are treating.
Literature Cited (remove “–“ and indent
after first line)
water using mucilage extracted from the
Opuntia ficus- indica cactus. Graduate
School Theses and Dissertations.pp 1-9.
- Bugbee, R.E. and A. Reigel. 2008. The
cactus moth, Melitara dentata (Grote), and
its effect on Opuntia macrorhiza in western
Kansas. pp 1-94.
- Buttice, Audrey Lynn. 2009. Reducing
sediment and bacterial contamination in
Acknowledgements
I would like to acknowledge the generous
support received from Professor Teh who
supplied the necessary equipment and
guidance to make the research happen. In
addition, we thank Saddleback College
Biological Sciences Department for
allowing us to use their facilities in the
study.
- Larsen, N.; Nissen, P.; Willatts. 2007.
The effect of calcium ions on adhesion and
competitive exclusion of Lactobacillus.ssp
and E. coli O138. Int. J. Food Microbiol. ,
114, 113–119.
- Lauenroth, William K.; Dougherty, R. L.
and Singh, J. S. 2009. Precipitation Event
Size Controls on Long-Term Abundance of
Opuntia Polyacantha (Plains Prickly-Pear)
in Great Plains Grasslands. Great Plains
Research: A Journal of Natural and Social
Sciences. pp 996.
- Edward, Lin. 2010. Removal of Sediment
and Bacteria from Water Using Green
Chemistry, Environ. Sci. Technol. 44 (9). pp
3514-3519.
- Grant, V. and K.A. Grant. 2010.
Systematics of the Opuntia phaeacantha
group in Texas. Bot. Gaz. Opuntia
lindheimeri group. Bot. Gaz. pp 1-18.
- Natural Standard Monograph. 2013.
Nopal(Opuntia) The Authority on
Integrative Medicine,accessed 12 FEB 2011.
- Young, N. 2011. The medicago genome
provides insight into the evolution of
rhizobial symbioses. Nature Magazine . 480.
pp 520–524.
Review Form
Department of Biological Sciences
Saddleback College, Mission Viejo, CA 92692
Author (s):________Tavakoli, Kathy_____________________________
Title:___
Effect of the Opuntia (ficus-indica) cactus mucilage extract in reducing sediment and
bacteria contamination from water.
Summary
Summarize the paper succinctly and dispassionately. Do not criticize here, just show that you understood the paper.
A substantial number of people lack access to clean drinking water. Turbidity of
water has a direct relationship with the likelihood of causing disease. Flocculation
decreases turbidity. Flocculation of water can be achieved with local naturally occurring
substance and can be on par with, if not better (more easily obtained and implemented on a
local level) than Aluminum Sulfate (A very common flocculant). This experiment compared
Opuntia ficus-indica mucilage of different types (gelling, non-gelling, and mixed) to
determine which one would be most effective (fastest settling rate and reducing
concentration E coli. rate).
There was no statistically significant difference in E coli. flocculation rates among
the different mucilages. While E coli. numbers were reduced (settled), there were still
sufficient numbers remaining afloat to warrant further purification (which is to be
expected with flocculation). For all three mucilages tested, the greater the concentration of
flocculant, the greater the floc formation rate in kaolin. The Gel forming extract had a
statisitically significantly faster rate of kaolin flocculation rate.
Part of what makes the mucilage an attractive option is their possibility of being
culturally accepted and implemented. Some people may have a suspicious knee-jerk
reaction to adding chemicals (alum.) to their water or these flocculants may not see
distribution on a broad enough basis. Mucilage is widely available, cheap, and easy to make.
Additionally it can potentially be grown and replenished. Even in settings where other
flocculants are available it can be used to supplement these processes.
General Comments
Generally explain the paper’s strengths and weaknesses and whether they are serious, or important to our
current state of knowledge.
The topic was interesting and very relevant to biology and current events.
Flocculation is an initial step in many processes of water treatment worldwide and
although it does not yield immediately drinkable water it provides vast improvement in
water quality. Access to clean water will likely become a more visible issue in future years
due in part to continued worldwide population growth (although growth rate is slowing it
is still growing) and variances in traditional water resources due to climatic change.
Finding new and innovative means of water purification can improve how we use life’s
most precious resource (excluding oxygen for humans). I would say that this experiment
was designed and executed relatively well (it seems like larger sample sizes could have
been relatively easily run but I do not know if that would have changed much). It is
interesting that the different mucillages had slightly different properties. Optimizing
mucillages to real-world scenarios would be a needed future step (as noted) to make the
process as economical as possible and therefore more likely to be implemented at a larger
level. Other than some minor typographical and formatting issues, it is a good paper.
Technical Criticism
Review technical issues, organization and clarity. Provide a table of typographical errors, grammatical errors,
and minor textual problems. It's not the reviewer's job to copy Edit the paper, mark the manuscript.

This paper was a final version
This paper was a rough draft
Suggested changes or notes indicated with this color (in parenthesis I added)
outside parenthesis was original text. Take ‘em or leave ‘em; they are just
suggestions I could be mistaken.
Figure 3 could have been represented with lines (or smaller points) and been more
readable.
Figure 4 y-axis (cell/min label). It does say this in the figure caption however I think it
would add to readability of image by itself.
Figure 4 Control? Comparison to water not treated with flocculant did it settle at all?
Weird/awkward spacing of columns which made it difficult to read, particularly at the
Top of page 2, Bottom of page 2, top of page 3. Columns should go top to bottom,
left to right. I suspect hat this was probably some weird error from changing
formats??
Recommendation
 This paper should be published as is
This paper should be published with revision
 This paper should not be published