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Procedia Engineering 00 (2017) 000–000
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Procedia Engineering 200 (2017) 428–433

3rd International Conference on Natural Fibers: Advanced Materials for a Greener World, ICNF
2017, 21-23 June 2017, Braga, Portugal

Production and characterization of cellulose nanoparticles from

nopal waste by means of high impact milling
Marin-Bustamante, M. Q.1 ; Chanona-Pérez, J. J.2*; Güemes-Vera, N3 . Cásarez-Santiago R.4
PereaFlores, M. J5 . ; Arzate-Vázquez, I6 . ; Calderón-Domínguez, G7 .
1, 2, 4, 7
Instituto Politécnico Nacional-Escuela Nacional de Ciencias Biológicas Unidad Profesional Adolfo López Mateos, Av. Wilfrido Massieu
s/n. Instituto Politécnico Nacional, 07738 México, CDMX.
3
Instituto de Ciencias Agropecuarias-Universidad Autónoma de Hidalgo-CICyTA, Av. Universidad s/n Rancho Universitario C.P. 43600.
Tulancingo, Hidalgo, México.
5, 6
Centro de Nanociencias y Micro y Nanotecnologías Instituto Politécnico Nacional (IPN), Av. Luis Enrique Erro S/N, Unidad Profesional
Adolfo López Mateos, Zacatenco, Delegación Gustavo A. Madero, C.P. 07738, México, CDMX.

Abstract

Apart from its dietary use, Opuntia ficus-indica (OFI), also known as nopal, is widely used in diverse non-food
areas given its multiple properties and health benefits. In Mexico City, each year de-thorning process of nopal
generates around 40,000 tons of waste. This waste contains a huge amount of hemicellulose and cellulose that can
be used as a new biodegradable nanocomposite. Therefore, the aim of this work was to purify nopal thorns to obtain
cellulose nanoparticles by high impact milling or high-energy ball milling. Confocal laser scanning microscopy
(CLSM) showed that thorns have two main polysaccharides arranged in parallel fibers: cellulose and lignin. Size of
the obtained cellulose nanoparticles were ranged from 24 to 122 nm. Crystallinity, as well as the type of cellulose
obtained, were analyzed by means of XRD to evaluate its potential use as nanocomposite.

Click here and insert your abstract text.

© 2017 The Authors. Published by Elsevier Ltd.
Peer-review under responsibility of the scientific committee of the 3rd International Conference on Natural Fibers: Advanced
Materials for a Greener World.

Keywords: nanoparticles, mechanical milling, nopal wastes, cellulose.

* Corresponding author. Tel.: +52 01 5539999805.

E-mail address: jorge_chanona@hotmail.com. (Chanona-Pérez, J. J.).

1877-7058 © 2017 The Authors. Published by Elsevier Ltd.

Peer-review under responsibility of the scientific committee of the 3rd International Conference on Natural Fibers: Advanced Materials for a
Greener World.

1877-7058 © 2017 The Authors. Published by Elsevier Ltd.

Peer-review under responsibility of the scientific committee of the 3rd International Conference on Natural Fibers: Advanced
Materials for a Greener World
10.1016/j.proeng.2017.07.060
 Marin-Bustamante et al. / Procedia Engineering 200 (2017) 428–433 429
Marin-Bustamante, M. Q et, al / Procedia Engineering 00 (2017) 000–000 2

1. Introduction

Food wastes have been considered an important issue due to the environmental problems that they generate.
Nowadays, the search of environment-friendly sources, as well as the use of non-timber agro-industrial wastes, for
obtaining lignocellulosic materials, can be considered as potential alternatives to obtain cellulose without affecting
forests (Sharma and Varma 2014).
Cactus (Opuntia ficus-indica: OFI) is an endemic cactaceae from America that develops in arid and semiarid
lands (Habibi et al., 2008). Plant structure is formed by modified flattened stems known as cladodes. Cladodes are
covered with thorns and multicellular hairs called trichomes that form the cactaceae characteristic areole (Vignon, et
al., 2004). OFI thorns can have different sizes, while cladodes usually present different green tones and sometimes
reddish or purple tones (Guevara, et al., 2010).
Young OFI cladode (nopal) is consumed as a vegetable at different stages of maturation (30 to 110 days) and has
proved to be an important food with bioactive properties in both human and animal diets. Given its multiple
properties, and its use as a cosmetic or as a medicinal plant, nopal has been widely studied (Ammar, et al., 2015).
On the other hand, while is true that OFI has a high fiber content, that can be considered as an ecological source of
cellulose, presence of lignin and hemicellulose microfibrils arrangements can reduce accessibility to cellulose
microfibrils, making difficult the purification of cellulose.
Therefore, delignification method is a critical step to successfully separate cellulose from other components.
Nevertheless, variations on contents and types of lignin and hemicellulose affect separation performance, obtaining
costs, and purity of the resulting cellulose (Sharma and Varma 2014). On the other hand, cellulose nanoparticles
(CNP) have generated scientific interest given their availability, low cost, high biodegradability, high strength,
among other characteristics.
However, dimensions of CNP depend on several factors that include the source from where the cellulose was
obtained and the precise preparation conditions (Hubbe et al., 2008). Currently, organic nanoparticles are being
synthesized due to its importance in diverse areas such as in food and pharmaceutical industries, in packaging
technologies, as components of paints, etc. (Cruz-Estrada et al., 2005; Kallioranta, 2012).
Furthermore, cellulose nanofibers are being used as reinforcement material in electronic devices, medicines, and
many other products, given their functionality, easy synthesis, properties, biodegradability, and biocompatibility.
However, the use of spherical CNP has not widely been applied in the industry at present (Qing et al., 2012). Hence,
the aim of this work was to obtain CNP from nopal thorns that could be used as reinforcement material in diverse
industries.

Nomenclature

CNP Cellulose nanoparticles

CR Cellulose reference
OFI Opuntia ficus-indica cactus
IAM Amorphous region
I200 Crystalline region

2. Materials and procedures

2.1 Plant material

OFI thorns were removed from nopal wastes from a recollection center located in Milpa Alta, Mexico City.
Removed thorns were dried at 35 °C and sieved (Siave Shaker RX-86 US) to eliminate any trace of cladodes.
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Marin-Bustamante, M. Q et, al / Procedia Engineering 00 (2017) 000–000 3

2.2 Confocal laser scanning microscopy (CLSM)

Dried and sieved thorns were cut in a cryostat (Leica, CM1850, Germany) to obtain slices of 20 μm of thickness.
Sliced thorns were stained during 5 minutes with calcofluor white M2R (fluorescent brightener 28F3543, Sigma,
USA) at 0.01% to determine the presence of cellulose. Stained thorns were dried at room temperature and observed
at 405 nm by using a diode laser (CW/pulsed) coupled on a CLSM (Carl Zeiss, LSM 710, Germany). On the other
hand, lignin presence was determined at 488 nm by using an Argon laser coupled on the same CLSM. No staining
was performed to determine lignin as this compound exhibits autofluorescence at the wavelength of analysis.

2.3 Nanoparticles obtaining

Material delignification was performed per Milox method. A sample amount of 2 g was placed in an 80-mL ball
flask to prepare a 1:40 solution (w/v). Solution was prepared with formic acid (64 mL), hydrogen peroxide (4 mL)
and distilled water (12 mL), and heated at 80 °C for 180 minutes under constant stirring. Delignified cellulose was
repeatedly rinsed with distilled water as reported by Hernández-Hernández et, al (2016), and pulverized in a high-
energy ball mill (Fritsch, Planetary ball mill model Pulverisette 7, Germany) to obtain CNP. Milling process used
agate bowls of 45 mL and 5 mm balls, while operating conditions were selected following the recommendations of
Neri-Torres et, al (2016) and Abbaszadeh et, al (2016).

2.4 Scanning electron microscopy (SEM)

Scanning electron microscopy (SEM Jeol Field Emission 6701F) was used to determine the size of CNP. Particle
size distribution and descriptive statistics were obtained by using SigmaPlot v 12.5 (Systat Software, Inc. USA).

2.5 X-Ray diffraction (XRD)

XRD (Miniflex 600 Rigaku Japan) was used to obtain the diffractograms of CNP and of cotton microcrystalline
cellulose (MCC, Aldrich, USA) that were employed for the later determination of the respective crystallinity
indexes (XCR). MCC was used as reference given its high crystallinity value. To obtain the diffractograms, samples
were placed at 45° with respect to the primary beams while operating conditions were: 0-60° of scanning angle (2θ),
300 s of step time, 0.05 s of step size, and 40 kV of power.
Percentage of XCR was estimated through Eq. 1 (Segal et al., 1959) where I200 (2θ= 22.7°) represents the
crystalline and amorphous regions, while IAM (IAM 2θ = 18°) represents only the amorphous one.

𝐼𝐼200 − 𝐼𝐼𝐴𝐴𝐴𝐴
𝑋𝑋𝐶𝐶𝐶𝐶 = × 100 (1)
𝐼𝐼200

3. Results and discussion

Figures 1A-1C show CLSM images of nopal thorns in longitudinal cuts along with the distribution of both
cellulose (blue) and lignin (green). Cellulose presented a poor fluorescence intensity while lignin autofluoresence
was very intense. Complex cellulose-lignin network (Figure 1C) showed an ordered arrangement along the thorns.
On the other hand, Figure 2 shows a SEM image of CNP obtained by Milox extraction and high-energy ball
milling. SEM analysis demonstrated that it is possible to generate particles of nanometric size (ranged from 24 to
122 ± 29 nm) and irregular shape.
Additionally, XRD showed that CNP peaks (Figures 3B and 3D) were wider than those of cotton cellulose
(Figure 3A) and of unpurified material (Figure 3C). This difference on the shape of the peaks could indicate a great
content of amorphous material on CNP (Thygesen et al., 2005). Liitiä et al. (2003) reported that a high breakup
degree of cellulose chains could release a large amount of free C4. Complementary, Thygesen et al., (2005) mention
 Marin-Bustamante et al. / Procedia Engineering 200 (2017) 428–433 431
Marin-Bustamante, M. Q et, al / Procedia Engineering 00 (2017) 000–000 4

that peaks (110’), (110), and (200) are associated with crystalline diameter on a perpendicular direction to the axis of
the cellulose fibers. Furthermore, coincidence between reference peak (Figure 3A) and CNP peaks (Figures 3B and
3D) confirms that cellulose is the main component of these nanoparticles.

Figure 1. CLSM images of longitudinal-sections of nopal thorns. Cellulose stained with calcofluor A), autofluorescence of lignin B), and
distribution of cellulose (blue) and lignin (green) in the thorn C).

Figure 2. SEM image of cellulose nanoparticles.

 Marin-Bustamante, M. Q et, al / Procedia Engineering 00 (2017) 000–000 5

432 Marin-Bustamante et al. / Procedia Engineering 200 (2017) 428–433

A
A

Figure 3. XRD patterns of cellulose samples. Reference cotton cellulose A), purified nanoparticles B), unpurified non-milled material C), and
non-delignified milled CNP D).

Finally, it was found that high values of XCR were associated to purified CNP and non-milled samples. Table 1
shows XCR for each sample.

Table 1: Crystallinity index

Sample Crystallinity index

Reference A 49.88%
Purified nanoparticles B 33.81%
Unpurified and without milled material C 46.79 %
Unpurified nanoparticles D 30.42%

4. Conclusions

This study showed the possibility to obtain cellulose from nopal thorns, that could allow the reuse of cactus waste.
High–energy ball milling demonstrated to be a simple and efficient technology to produce nanomaterials of
biological origin.

Acknowledgements

Authors thank financial grant provided through projects IPN-SIP 20170232 and CONACyT 239899, and by
COFAA-IPN Mexico.
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Marin-Bustamante, M. Q et, al / Procedia Engineering 00 (2017) 000–000 6

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