Journal of Water Process Engineering 7 (2015) 306–313

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Journal of Water Process Engineering

journal homepage: www.elsevier.com/locate/jwpe

Bioflocculants extraction from Cactaceae and their application in

treatment of water and wastewater
M. Belbahloul ∗ , A. Zouhri, A. Anouar
Laboratoire de chimie Appliquée et Environnement, Faculté des Sciences et Techniques Université Hassan 1er B.P. 577, Settat, Morocco

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

Article history: Effect of extraction conditions, including solvent types and extraction temperatures on the power floc-
Received 15 July 2014 culant of the mucilage (MP), and water (WP), acid (AP), chelating (CP) and alkali-pectin (PHEX) extracted
Received in revised form 12 July 2015 from the cladodes of Opuntia Ficus Indica (OFI) was evaluated. Two protocols (protocol 1 at 90 ◦ C and
Accepted 14 July 2015
protocol 2 at ambient temperature) were used with different combination and temperature for the extrac-
Available online 27 August 2015
tion of the foregoing. It was found that AP, PHEX and CP extracted by high temperature showed higher
yield than those extracted at room temperature except water extract. Flocculant power was evaluated
Keywords:
for sets of products on the elimination of materials in suspension for three different pH values. The PHEX
Cactus
Coagulation extracted at ambient temperature can act as a coagulant and flocculant together for turbid water.
Flocculation © 2015 Elsevier Ltd. All rights reserved.
Turbidity
Water treatment

1. Introduction from the cactaceae family; specially mucilage and pectins, have
been used empirically to modify the rheological properties of some
Turbidity, which is caused by fine suspended particles of clay, products[4]; they can modulate rheological properties of foods, and
silt, organic and inorganic matter, plankton and other microscopic are generally used as food thickeners, texture modifier, stabilizers
organisms, is an important indicator to evaluate the safe use of and emulsifiers for various applications.
water for human consumption [1]. Turbidities less than 5 Neph- Opuntia Ficus Indica (Cactaceae) is cultivated in Morocco for fruit
elometer Turbidity Units (NTU) are considered to be ‘safe’ by most production. The production of mucilages is characteristic of mem-
consumers [2]. bers of the Cactaceae family [5]. The chemical composition of O.
However, the World Health Organization (WHO) unofficially ficus indica mucilage from cladodes, commonly named nopal, has
considers 0.1 NTU to be the maximum turbidity allowed for dis- been the subject of various studies. Amin et al. [6] found that the
infection [2], because suspended particles promote the growth mucilage was a neutral polysaccharide that contained arabinose,
of microorganisms in water and therefore decrease the effective- rhamnose, galactose and xylose. According to Trachtenberg and
ness of the disinfection procedure [3]. Thus, turbidity reduction Mayer [7]; the mucilage was a polysaccharide, which contained
can vastly improve the effectiveness of disinfection methods. The 10% uronic acids, arabinose, galactose, rhamnose and xylose. The
use of natural environmentally benign agents in the treatment of mucilage of OFI is a thick, gummy substance and is what provides
water is rapidly gaining interest due to their inherently renew- the cacti’s natural ability to store large amounts of water. When in
able character and low toxicity. Scientists, governments, the media, water, the mucilage swells, producing unique surface-active prop-
and individuals worldwide recognize the benefits of green tech- erties seen in many natural gums, giving the mucilage a suspected
nology. There is a tremendous push towards developing new green ability to precipitate particles and ions from aqueous solutions.
technologies in order to minimize environmental changes. Polysac- The mucilage is extracted from the pads of the cactus. Diced nopal
charides from plant extracts are an interesting source of additives cladodes have been used for centuries in Latin America as a prim-
for several industries, in particular for the environment. These itive technology for the rapid flocculation of turbid natural spring
polymers have the advantage of being regarded as totally natu- waters, but a scientific baseline has never been provided for this
ral for many consumers. Many of these polysaccharides, like those observed phenomenon [8].
Pectins are a family of complex polysaccharides (heteropolysac-
charides) which comprise a diversity of carbohydrate residues.
∗ Corresponding author. Similarly, the majority of other plant polysaccharides, the pectins,
E-mail address: belbahloulmounir@yahoo.fr (M. Belbahloul). are polydisperse in the composition and molecular size, and they

http://dx.doi.org/10.1016/j.jwpe.2015.07.002
2214-7144/© 2015 Elsevier Ltd. All rights reserved.
 M. Belbahloul et al. / Journal of Water Process Engineering 7 (2015) 306–313 307

are heterogeneous with respect to the chemical structure and materials, including clay, silt, or mineral particles from soils, or from
molecular weight. Its composition varies with the source and the natural organic matter created by the decay of vegetation.
conditions of the extraction, position, and other ambient factors
[9]. Pectin is a complex anionic polysaccharide composed of ß-1, 2.2. Methods
4-linked D-galacturonic acid residues, wherein the uronic acid car-
boxyls are either fully (HMP, high methoxyl pectin, DE > 50%) or 2.2.1. Mucilage and pectins extraction
partially (LMP, low methoxyl pectin, DE < 50%) methyl esterified. Prior to pectin extraction, the fresh cactus pads were cleaned to
Like alginates, low methoxyl pectins, form gel with divalent cations remove thorns and cut into small pieces (2 × 0.1 cm) with a kitchen
[10]. Various studies have shown that parts of plants and extraction knife. Cactus pieces were heated in water at 85 ◦ C for 20 min to inac-
conditions significantly influenced the yield and physicochemi- tivate enzymes and left to cool to ambient temperature; neutralized
cal characteristics of plant polysaccharides, such as the chemical to pH 7.5 from initial pH 4.0 in order to induce de-esterification
compositions, molecular weight distribution, and would affect the of methoxyl groups and filtered through a cloth filter to extract
rheological characteristics such as thickening and gelling proper- mucilage as much as possible [16]. Pectic polysaccharides were
ties, as well as surface activity such as emulsifying capability and sequentially extracted from cladodes according to two protocols;
emulsification stability [11]. Extensively used as a gelling and thick-
ening agent, pectin has been industrially obtained under acidic • Protocol 1
conditions with oxalic [12], hydrochloric [13], and sulphuric acid
[14]. Although the use of strong acids provides high extraction yield Pectins were extracted according to Zhang et al. [17] with
and time-saving advantages, it can cause serious environmental slight modification. The Pectic polysaccharides were sequentially
problems such as the disposal of acidic wastewater and also play a extracted from cladodes residue (R1) by water (2 × 2 h at 85 ◦ C)
negative role in consumer preference. Alternative treatments have [18], aqueous solution of 0.5% oxalate (0.25% oxalic acid + 0.25%
been therefore taken into account for minimizing the use of detri- ammonium oxalate) [18](2 × 2 h at 85 ◦ C), then the solid residue
mental chemicals during pectin extraction. R3 was suspended in a NaOH 50 mM sodium hexametaphosphate
Many chemical flocculants, including polyaluminum chloride 7.5 g/L solution (pH 12.0) stirred continuously (2 × 1 h at 85 ◦ C) and
(PAC), ferric chloride, and polyacrylamide (PAM), have been widely finally the residue R4 were extracted with HCl 0.05 N (2 × 2 h at
used in wastewater treatment in step of flocculation for removing 85 ◦ C) [18]. All extracts were separated from the residues by fil-
suspended solids (SS), although there are concerns about the tox- tering through a nylon cloth [18], and the pectin was precipitated
icity of these chemicals for recovering organics, especially in the by centrifugation (30 min, 4500 rpm) with 2 volumes of ethanol
food and fermentation industries. Since, bioflocculants can be non- resulting in a Water-Soluble Pectin (WP1 ), Chelating-Soluble Pectin
toxic, harmless and without secondary pollution, they have a great (CP1 ), Sodium Hexametaphosphate-Soluble Pectin (PHEX1 ) and
potential for use in those industries. However, there is evidence Acid-Soluble Pectin (AP1 ), respectively. The extraction scheme is
of health problems including Alzheimer’s disease [15] caused by given in Fig. 1a.
aluminum salts, as well as the formation of neurotoxic and carcino-
genic acrylamide monomers that are harmful to humans and the • Protocol 2
environment. The dosage of flocculants significantly increases with
the deterioration of coagulation effect at low temperatures, giving The residue R1 was extracted twice by bidistilled water, 0.5%
rise to a potential safety hazard. In view of these concerns, biofloc- solutions oxalate, NaOH 50 mM sodium hexametaphosphate 7.5 g/L
culants are attractive alternatives to existing synthetic flocculants solution (pH 12.0) (2 × 1 h) and pectin was recovered according to
such as PAC, PAM, pectin and mucilage. This study seeks to uncover Cardenas et al. [16] and finally HCl 0.05 N at ambient temperature.
an innovative new low-cost technology based on the mucilage and The pectic extracts were separated from the residues using a nylon
pectins extracted at 90 ◦ C and at ambient temperature, and with cloth filter [18], and the pectins other than that extracted with
different solvent from a common cactus to be implemented for sodium hexametaphosphate was coagulated with two volumes of
turbidity reduction in water with lime as coagulant, the elimina- ethanol by centrifugation (30 min, 4500 rpm), the pellets obtained
tion power of sets of fives products extracted, was compared for corresponds respectively to the WP2 , CP2 , PHEX2 and AP2 (Fig. 1b).
both protocols and at three pH values; 3.0, 8.0 and 11.5. The results
of this study indicate that this technology may have the poten- 2.2.2. Pectin and mucilage purification
tial to be implemented in large-scale water treatment systems, Mucilage and Pectins coagulates were washed several times
particularly when considering the trend towards ‘green’ chemistry with 50, 70, 85% ethanol and finally with pure ethanol before dry-
solutions. ing at 50 ◦ C overnight. The extraction yield of pectin production was
calculated by dividing the dry weight of pectin by the dry weight
of starting cladodes [19].
2. Materials and methods
2.2.3. Optimization of pH and flocculants dosage
2.1. Materials The optimum dosage of the natural flocculants can be evaluated
by using Jar test apparatus VELP Scientifica JLT6, equipped with six
A batch of fresh cladodes of OFI, was obtained from a plantation mixing posts. The collected soil was used as a model suspension.
in Berrechid (33◦ 15 36 N and 7◦ 36 0 W), Morocco, and stored at Two hundred grams of soil were dispersed in 1 L of deionized water
4 ◦ C until mucilage and pectins extraction (period not exceeding 10 in a high-speed blender to prepare a stock solution. The synthetic
days). Harvesting of cladodes was carried out in December 2013. raw water was prepared by mixing various volumes of a stock solu-
The chemicals used in the experiment were of analytical grade tion with distilled water, for five hours rapid mixing and then set
and were procured from the Sigma–Aldrich, Fluka, and Merck. Bi- to stand for 30 min. Second, the 600 ml supernatant from the high
distilled water was used throughout. turbidity synthesized raw water was diluted into distilled water.
Natural soil was used to prepare synthetic turbid water, the sam- The mixture was diluted to 1000 NTU, the various parameters ana-
ples loaded with suspended solids are obtained from a dry lake that lyzed in this study were Turbidity and pH. Turbidity measurements
lies in the region of Sahel province of Berrechid (33◦ 20 51 N and were conducted using digital turbidity Meter (HANNA LP2000-11).
7◦ 49 52 W), turbidity may be created from a wide variety of eroded The pH was measured using a digital pH multi-parameter (Consort
308 M. Belbahloul et al. / Journal of Water Process Engineering 7 (2015) 306–313

(a)

(b)

Fig. 1. (a) Extraction of mucilage and pectic polysaccharides from cactus cladode at 85 ◦ C (i = 1). (b) Extraction of mucilage and pectic polysaccharides from cactus cladode at
ambient temperature (i = 2). Where: MP = Mucilage Pads, APi = Acid-soluble Pectin, WPi = Water-soluble Pectin, CPi = Chelating-soluble Pectin, PHEXi = Hexametaphosphate-
Soluble Pectin and R = Residue.

C3050). The pH of the synthesized water was adjusted using H2 SO4 dosage of flocculants. Instantaneous variations in turbidity were
(0.1N) or lime (also used as natural coagulant) in the pH range of monitored over the 60 min of flocculation. Data were processed
3.0–11.5 (pH 11.5 is the optimum flocculation when using the juice using Microsoft Excel, and the curve of turbidity vs. flocculation
of the whole cladode for treatment of the same sample). To perform time was plotted.
flocculation and sedimentation tests, a known volume of 0.2 wt.%
polymer solutions (mucilage or pectin) was added in a single step 3. Results and discussion
to 500 ml of well-mixed slurry using a pipette and placed under jar
test apparatus. The apparatus was switched on and the speed of 3.1. Effect of pH on flocculation of natural clay using cactus
mixing was set at 160 rpm (coagulation). The required dose of pre- extracts
viously prepared flocculant was added simultaneously and rapidly
mixed for a minute. The speed of mixing was reduced to 30 rpm Tests were conducted as described, at pH values of 3.0, 8.0 and
and allowed for slow mixing for 60 min for flocculation. Then, the 11.5 using flocculants concentration of 0.2 wt.%. The results are pre-
supernatant is pipetted out from the each beaker to record the final sented in Fig. 2. From the figure, it is seen that at pH 3.0, only
turbidity. The procedure was repeated to evaluate pH optimum by PHEX2 and CP2 produces appreciable reduction of turbidity higher
varying the pH of synthetic water as 3.0, 8.0 and 11.5 at constant than 90%, the maximum removal observed was 94.68 and 91.48%
 M. Belbahloul et al. / Journal of Water Process Engineering 7 (2015) 306–313 309

Fig. 2. Effect of pH on removal of turbidity of natural clay using cactus extracts (0.2 wt.%) as flocculants, Initial turbidity = 1000 NTU. (a) residual turbidity at pH 3.0, (b)
turbidity removal at pH 3.0, (c) residual turbidity at pH 8.0, (d) turbidity removal at pH 8.0; (e) residual turbidity at pH 11.5 and (f) turbidity removal at pH 11.5.

with residual turbidities of 4.0 and 6.0 NTU respectively, while CP1 with MP are very coarse and settled almost completely in less than
increases turbidity of our samples against the standard and increase 15 min. Residual turbidity of 37, 33 and 10 NTU were obtained for
the turbidity of more than 50% (see Fig. 2(a) and (b)). The maxi- PHEX1, AP1 and AP2, respectively at pH 11.5, with suppression of
mum turbidity removal was found to be 98.75% and 93.92% when the maximum turbidity of 93.47, 94.90 and 98.14%, respectively.
treated with PHEX2 and CP2 , with the final turbidity being less than (see Fig. 2(e) and (f))
or equal to 3 NTU and 21 NTU, respectively for pH 8.0 as shown in
Fig. 2(c) and (d). The maximum efficiency is observed at pH 11.5 for 3.2. Determination of optimum dosage of cactus extracts
almost all extracts more than 98% removal of turbidity is achieved
at this pH and the residual turbidity drops below 5 NTU for MP after As maximum efficiency of flocculation was around pH 11.5,
10 min of flocculation and can achieve 99.08%, while WP1 reduce experiments were conducted by varying the dosage of all extracts
turbidity with removal ranging from 98.41 to 99.55% and the resid- at this pH. Tests were run using cactus extracts ranging from 0.1
ual turbidity was found between 2 and 8 NTU and the flocs obtained to 0.6 wt.%. The results are presented in Fig. 3. The lowest dosage
310 M. Belbahloul et al. / Journal of Water Process Engineering 7 (2015) 306–313

Fig. 3. Effect of varying mucilage and pectins dosages on the removal of turbidity due to river silt in water having initial turbidities of 1000 NTU at pH 11.5.

with maximal efficiency was found to be 0.2 Wt.% for MP (99.28%), this dosage, turbidity increases, probably due to destabilization
WP1 (99.64%), AP1 (94.91%), and AP2 (97.99%). While the opti- of suspended particles. Again, in the present study, it is observed
mum dosage for PHEX1 (94.27%) and WP2 (98.30%) was found that irrespective of initial turbidity, application of 0.2–0.3Wt.%
0.3 Wt.% and 0.6 Wt.% for CP1 (86.42%), and 0.5 Wt.% for PHEX2 of MP or WP1 leaves a residual turbidity of only less than 4
(98.3%) and finally 0.1 Wt.% for CP2 (98.54). Above these dosages, NTU as can be seen from Fig. 4(a) and (b), respectively. It was
the suspension showed a tendency to destabilize. These results also observed that at higher initial turbidity of 1000 NTU, flocs
indicate that the effectiveness of cactus extracts in removing tur- appear rapidly and grow to a larger size with MP. The flocs are
bidity due to the soil river is highly dependent on flocculants fibrous and stick to the stirrer or form large entangled mass resem-
concentration. Irrespective of initial turbidity, 0.2 Wt.% of MP and bling cobwebs, probably indicating that the bridging mechanism is
WP1 lowers turbidity below 3 and 1 NTU, respectively. Beyond operating.
 M. Belbahloul et al. / Journal of Water Process Engineering 7 (2015) 306–313 311

Fig. 4. optimization of two water soluble extract dosage (0.1–0.6 wt.%) on removal of turbidity at pH 11.5. (a) residual turbidity of MP and (b) residual turbidity of WP1 .

3.3. Effect of a new PHEX2 extract on turbidity removal without rise to a significant chlorine demand [2]. All the residual turbidi-
coagulant ties satisfies WHO’s limit (5 NTU) for drinking water. It was clear
from the present study that the PHEX2 showed a good flocculating
In coagulation process, rapid mixing is used to spread out the activity for synthetic turbid water. However PHEX1 had a maxi-
coagulant throughout the turbid water. In flocculation process, mum removal near 70% and with residual turbidity about 100 NTU.
slow mixing is a key part to get most favorable performance. Ade- New technique has considerably reduced the lime dose used. Also
quate time must be provided to allow production of particles of sludge produced was less voluminous and readily biodegradable.
sufficiently large size to permit their efficient removal in sedimen- These results agree well with those obtained by Nkurunziza et al.
tation process [20]. In the present study, pectin PHEX2 was added [21], Narasiah et al. [22] and Bina et al. [23] for various natural
to turbid water with rapid mixing for 5 min, followed by gentle flocculants.
mixing (flocculation) for 60 min and the residual turbidities were The cactus pectin extract afforded by the alkaline process,
measured. Fig. 5 demonstrates the performance of conventional referred here as PHEX2 represents 0.82% of the fresh tissue weight
process with coagulant-flocculant and a new technique with PHEX2 (Table 1). Even when some soluble pectin may have been lost
alone. Excellent results were observed using this new technique. during the first aqueous extraction treatment intended to remove
Residual turbidities at 35 min were observed to be 3 NTU with mucilage, the yield obtained for the extraction process was con-
percentage removal of 98.5%, and at 40 min were 2 NTU with per- siderably greater than that documented by Cardenas et al. [16],
centage removal of 98.8% for the high turbid water Fig. 5(a) and 0.6% and for pectin extracted from prickly pear peel, 0.12% [24].
(b), high levels of turbidity can protect microorganisms from the The contents of PHEX1 (0.98%), PHEX2 (0.82%) and CP1 (0.66%)
effects of disinfection, stimulate the growth of bacteria and give are much higher than that of WP1 (0.05%), AP1 (0.24%), WP2
312 M. Belbahloul et al. / Journal of Water Process Engineering 7 (2015) 306–313

Fig. 5. Effect of PHEX2 on turbidity removal with coagulant (pH 11.5) and alone (pH 8.0). (a) Turbidity removal of clay by PHEX2 , (b) residual turbidity with PHEX2 .

Table 1 3.4. The extracts used as flocculating agents are environmentally

Percentage yield of mucilage and pectins (g/100 g fresh weight) extracted from the
harmless
cactus pads under various conditions.

Cactus extracts Yield (%) According to the results obtained in this study, we find that MP,
Mucilage 0.26 WP1 and PHEX2 give better results compared to other pectin’s fam-
Water Soluble Pectin1 0.05 ilies. It was also found that PHEX2 did not affect the pH of the
Acid- Soluble Pectin1 0.24 solutions. As already mentioned in the extraction protocols, MP
Sodium Hexametaphosphate-Soluble Pectin1 0.98
and WP1 are extracted only by distilled water, unlike other types
Chelating- Soluble Pectin1 0.66
Water soluble pectin2 0.13 of pectins already discussed above. However, the use of PHEX2
Acid-soluble pectin2 0.14 in water treatment raises the question about his respect for the
Sodium Hexametaphosphate-Soluble Pectin2 0.82 environment, which suggests another study to unveil its effects.
Chelating-Soluble Pectin2 0.24
The efficiency of MP, WP1 and PHEX2 was compared with
two other industrial flocculants (an anionic and nonionic) for the
removal of turbidity at optimum conditions for each was shown
(0.13%), AP2 (0.14%) and CP2 (0.24%), together with total yields in Table 2. It was found that the anionic flocculant reduced maxi-
of pectins obtained at high temperature (1.93%) are higher than mum turbidity among nonionic flocculant. It reduced up to 99.3%
those obtained at room temperature (1.33%). The mucilage yield for highly turbid water at pH 11.5 which is almost as same as the
expressed with respect to fresh weight (0.26%), was higher than reduction capacity of MP, WP1 and PHEX2 . So, the nonionic floc-
the 0.07% obtained by Cardenas et al. [25] and lower than 1.5% culant have a lowest reduction of turbidity among all flocculants
obtained by Saenz et al. [26]. However, the climatic conditions at studied, it had a residual turbidity of 12 NTU with a maximum
the time of the pads collection (winter, cold and rain), may justify removal of 87%. Renuka et Jadhav [28] have used Moringa Oleifera
this high value, since the same authors indicate that such values (Drum sticks), Okra gum, and the mucilage isolated from the dry
could increase under favorable hydrated conditions, especially in flowers of C. Procera as flocculent for the treatment of turbid water
the cells of the parenchyma where the water was stored [26] and samples containing synthetic turbidity caused by clay materials of
contents of mucilage and pectins show a random variation with initial turbidity of 500 NTU. The maximum turbidity removal effi-
cladodes age [27]. ciency was observed for Moringa Oleifera with 92% and a residual
 M. Belbahloul et al. / Journal of Water Process Engineering 7 (2015) 306–313 313

Table 2
Residual turbidity and percentage turbidity removal with different flocculants.

Product Mucilage Water Soluble Pectin1 Sodium Hexametaphosphate-Soluble Pectin2 Anionic Flocculant Nonionic Flocculant

Residual turbidity (NTU) 4 3 2 2 12

% of turbidity removal 99.28 99.64 98.8 99.3 87

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