Pineapple crown delignification using low-cost ionic liquid based on ethanolamine and organic acids

Carbohydrate Polymers 206 (2019) 302–308

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Carbohydrate Polymers
journal homepage: www.elsevier.com/locate/carbpol

Pineapple crown delignification using low-cost ionic liquid based on
ethanolamine and organic acids

T

Rita de C.M. Mirandaa,b, Jaci Vilanova Netaa, Luiz Fernando Romanholo Ferreiraa,c,
Walter Alves Gomes Júniord, Carina Soares do Nascimentod, Edelvio de B. Gomesd,
⁎
Silvana Mattedie, Cleide M.F. Soaresa,c, Álvaro S. Limaa,c,
a

UNIT, Universidade Tiradentes, Av. Murilo Dantas, 300, Farolândia, 49032-490, Aracaju, SE, Brazil
Uniceuma, Mestrado em Meio Ambiente, Renascenỗa, 65075-120, Sóo Luớs, MA, Brazil
c
ITP, Instituto de Tecnologia e Pesquisa, Av. Murilo Dantas, 300-Prédio do ITP, Farolândia, 49032-490, Aracaju, SE, Brazil
d
IFBA, Instituto Federal da Bahia, Campus Salvador, Departamento de Tecnologia em Saúde e Biologia, Rua Emídio dos Santos, s/n - Barbalho, 40301-015, Salvador, BA,
Brazil
e
UFBA, Universidade Federal da Bahia, Escola Politécnica, Departamento de Engenharia, Rua Aristides Novis 2, Federaỗóo, 40210-630, Salvador, BA, Brazil
b

A R T I C LE I N FO

A B S T R A C T

Keywords:
Ionic liquid
Biomass
Pineapple
Lignocellulose
Treatment

Pineapple fibre was treated with protic ionic liquids (PILs) and the effects on the structure, composition, and
properties of the fibres were evaluated. Treatment with PILs efficiently exposed the fibre surface, as confirmed
by scanning electron microscopy. The chemical composition analysis revealed reductions in the lignin and
hemicellulose contents in the treated fibres, promoting exposure of cellulose. The results correlated with the
crystallinity index, which was greater in the treated fibres compared with that in the untreated fibres. The
generated residue from the treatment of fibres with PIL (1%, v/v) showed lower levels of toxic compounds,
demonstrating the advantages of this treatment over conventional biomass treatments.

1. Introduction
The treatment of lignocellulosic biomass is fundamental in the
economy (Tan et al., 2009; Yuan, Xu, & Sun, 2013) and characterized as
an expensive step in conversion of biomass. The main basic treatment
based on physical (mechanical comminution, pyrolysis, steam pretreatment and steam explosion), chemical (acid pretreatment, alkali
pretreatment and sufur dioxide), biological (microbial and enzymatic)
or hydrid tion, microwave irradiation, ammonia fibre explosion and
liquid hot water (Singh, Shukla, Tiwari, & Srivastava, 2014; Fu, Mazza,
& Tamaki, 2010).
Recently, several studies on the efficiency of delignification of
aprotic ionic liquids (AILs) were performed (Brandt, Gräsvik, Halletta,
& Welton, 2013; Pinkert, Dagmar, Goeke, Marsh, & Pang, 2011; Sun
et al., 2009). Their dissolution properties are responsible for the appropriate choice of the cations and/or anions of AILs, moreover the

environmentally friendly properties has also an importante role in this
process (Anugwom et al., 2014). The short alkyl chain AILs are preferred, and the most traditionally used is imidazolium acetate due to the
highest solubility of lignin (Zakrzewska, Bogel-Łukasik, & BogelŁukasik, 2010). However, there are many drawbacks, such as high
⁎

process temperature (> 100 °C), low efficiency (< 50%), solid waste
accumulation, high viscosity recovery difficulties and high cost (Rashid,
Kait, Regupathi, & Murugesan, 2016).
Alternately, the literature has been working with protic ionic liquids
(PILs) because they are cheap and easily synthesized (Achinivu,
Howard, Li, Gracz, & Henderson, 2014). PILs are salts formed from acid
/ base reactions (MacFarlane, Pringle, Johansson, Forsyth, & Forsyth,
2006) at mild temperature (< 100 °C) as an alternative to conventional
lignin removal methods (Achinivu et al., 2014). The mechanism of
action of LPIs is based on the solubility of lignin.
In the last decade, the studies focused on the dissolution of biogenic
polymers and demonstrated the great potential of PILs (Swatloski,
Spear, & Holbrey, 2002). The ability of PILs to solubilize lignin and
carbohydrates depends on the fact that these compounds act on the
complex of bonds formed by the rupture of the lignocellulosic complex
(Singh, Simmons, & Vogel, 2009; Swatloski et al., 2002). The action of
PILs on lignin, promoting the cleavage of βeOe4 bonds between 130
and 200 °C (Cox & Ekerdt, 2012, 2013; Jia, Cox, Guo, Zhang, & Ekerdt,
2010; Long, Li, Guo, Wang, & Zhang, 2013).
Therefore, this work hypothesized that the PILs could remove the
residual lignin from pineapple crown as biomass. In this study, the

Corresponding author at: UNIT, Universidade Tiradentes, Av. Murilo Dantas, 300, Farolândia, 49032-490, Aracaju, SE, Brazil.
E-mail address: (Á.S. Lima).

/>Received 1 June 2018; Received in revised form 29 October 2018; Accepted 30 October 2018
Available online 31 October 2018
0144-8617/ © 2018 Elsevier Ltd. All rights reserved.


Carbohydrate Polymers 206 (2019) 302–308

R. de C.M. Miranda et al.

2.3. Structural characterization

selected ionic liquids are formed by organic acids with small alkyl chain
(acetic, propionic, butyric and pentanoic acid) due to their better delignification capacity; and by amines with different substitutions of the
monoethanolamine hydrogen, producing amines more hydrophilic (2hydroxyethyl - diethanolamine) or more hydrophobic (methyl – methyl-monoethanolamine). In this way the set of ionic liquids depicts
different properties, low toxicity and low cost (Oliveira et al., 2016;
Ventura et al., 2012).

Amorphous and crystalline regions of the samples were characterized by Energy Dispersive X-ray diffraction spectroscopy (XRD,
Shimadzu Corp., model XRD-6000), with a CuK radiation source, a
voltage of 40 kV, and a current of 40 mA. Scans were taken over a range
of 2θ values of 10–90° at a scan rate of 0.05° min−1. The degree of
crystallinity was evaluated using the crystallinity index, calculated according to the empirical model (Eq. 1) of Segal, Creely, Martins, and
Anndre Conrad (1959).

2. Material and methods

CrI =
2.1. Materials

2.4. Ionic liquid treatment

The methodology for biomass delignification using PILs was developed by Varanasi et al. (2012). Biomass (300 mg) was mixed with
9.7 mL of PIL at room temperature. The solution was then heated to
100 °C in a water bath for 1 h. After the treatment, samples were
thoroughly mixed and 35 mL of hot water was added to the sample to
precipitate the dissolved biomass. The mixture of PIL, water, and biomass was centrifuged to separate the solid (recovered biomass) and
liquid (PIL and water) phases. The recovered biomass was washed four
times with hot water to remove excess PIL.
2.5. Chemical characterization
Untreated and treated biomass samples were analysed according to
the protocols of the National Renewable Energy Laboratory (NREL)
(Sluiter et al., 2006, 2012). Before the analysis, 5 g of the sample were
extracted in two consecutive steps with water and ethanol using 250 mL
of solvent for 2 h. After extraction, 300 mg of the samples were hydrolysed with 3 mL of sulphuric acid at 30 °C for 1 h. The reaction
mixture was diluted to 4% (by weight) with water and autoclaved at
121 °C for 1 h. The liquid was then analysed for its sugar content using
ultrafast high-pressure liquid chromatography (UFLC) on a ShimadzuProminence liquid chromatograph with a refractive index detector
(RID-10 A). The concentrations of monomeric sugars in the soluble
fraction were determined using a Supelcogel-Pb column (30 cm × 7.8
mm, 9 μm, equipped with a pre-column) at 85 °C and an elution rate of
0.6 mL min−1, using ultrapure water as the mobile phase. The concentrations of sugars derived from the hydrolysis of cellulose and
hemicellulose were determined from calibration curves generated using
standard solutions (Sluiter et al., 2012).
The acetyl groups were determined using high-performance liquid
chromatography (HPLC) with the same system as above but with a BioRad HPX-87H column at 45 °C. The mobile phase was 5 mM H2SO4, at a
flow rate of 0.5 mL min−1. The solids were dried to constant weight at
105 °C and considered insoluble lignin (IL). The soluble lignin (SL)
concentration in the filtrate was determined based on UV spectrophotometry at 280 nm.
Total lignin content in the untreated and treated samples was
measured by the acetyl bromide method, according to Sluiter et al.
(2012), with modifications. Pineapple powder (5 mg) was treated with

25 wt% acetyl bromide in glacial acetic acid (0.2 mL). The tubes were
sealed and incubated at 50 °C for 2 h with shaking at 500 rpm on a
thermomixer. After digestion, the solutions were diluted with three
volumes of acetic acid (0.6 mL), and then 0.1 mL aliquots were

2.2. Morphological characterization
Photomicrographs of the untreated and treated biomass samples
were obtained using scanning electron microscopy (SEM) (JEOL model
JSM5310) by detecting secondary electrons after depositing the sample
on a gold substrate.

Table 1
Chemical structure of designed ionic liquid cations and anions.

2-hydroxyethylammonium

Anion

Acetate

bis(2-hydroxyethyl)ammonium

Propionate

N-methyl-2-hydroxyethylammonium

Butyrate

(1)

where: CrI = crystallinity index (%); I(002) = diffraction peak intensity
of the crystalline material that is close to 2θ = 22°; I(am) = diffraction
peak intensity of the amorphous material that is close to 2θ = 18°.
The values found after the calculation of the crystallinity index are
relative values, assuming that the value for microcrystalline cellulose
(MCC) is 100%.

In the present study, 12 PILs were used: 2-hydroxyethylammonium
acetate (2HEAA), 2-hydroxyethylammonium propionate (2HEAPr), 2hydroxyethylammonium butyrate (2HEAB), 2-hydroxyethylammonium
pentanoate (2HEAP), bis(2-hydroxyethyl)ammonium acetate (BHEAA),
bis(2-hydroxyethyl)ammonium propionate (BHEAPr), bis(2-hydroxyethyl)ammonium butyrate (BHEAB), bis(2-hydroxyethyl)ammonium
pentanoate (BHEAP), N-methyl-2-hydroxyethylammonium acetate (m2HEAA), N-methyl-2-hydroxyethylammonium propionate (m-2HEAPr),
N-methyl-2-hydroxyethylammonium butyrate (m-2HEAB), and N-methyl-2-hydroxyethylammonium pentanoate (m-2HEAP). Their cation
and anion chemical structures are depicted in Table 1.
The PILs were synthesized by reacting equimolar amounts of amine
and the respective organic acids, according to Alvarez, Mattedi, MartinPastor, Aznar, and Iglesias (2010). During the course of the experiments, the purities of the solvents were monitored by their density and
speed of sound measurements.
Pineapple crown samples (Ananas comosus) were used as the raw
material, which were obtained from a local market in Aracaju SE,
Brazil. Pineapple crown were washed, cut, and dried at 60 °C for 48 h.
The dried biomass was milled through a 32–60 mesh sieve.

Cation

I(002) − I(am)
I(002)

Pentanoate

303

304

35.7 ± 0.9
26.6 ± 1.3
10.6 ± 0.3
8.9 ± 1.1
3.9 ± 0.1
3.2 ± 0.6
1.34 ± 0.03
21.9 ± 0.6
3.92 ± 0.02
0.09 ± 0.05
89.55
34.9 ± 0.9
29.5 ± 1.3
10.6 ± 0.3
8.9 ± 1.1
5.6 ± 0.1
4.4 ± 0.6
2.04 ± 0.03
21.9 ± 0,6
3.92 ± 0.02
1.09 ± 0.05
91.75
35.8 ± 0.9
26.1 ± 1.3
10.6 ± 0.3
8.9 ± 1.1

3.9 ± 0.1
2.7 ± 0.6
1.12 ± 0.03
21.9 ± 0.6
3.91 ± 0.02
2.09 ± 0.05
90.92
35.9 ± 0.9
30.0 ± 1.3
11.8 ± 0.3
10.6 ± 1.1
4.7 ± 0.1
2.9 ± 0.6
1.14 ± 0.03
19.9 ± 0.6
2.72 ± 0.02
1.29 ± 0.05
91.35
35.6 ± 0.9
30.7 ± 1.3
11.3 ± 0.3
10.9 ± 1.3
3.6 ± 0.1
1.9 ± 0.6
1.24 ± 0.03
17.9 ± 0.6
3.91 ± 0.02
0.09 ± 0.05
92.4
38.6 ± 0.9

33.3 ± 1.3
12.6 ± 0.3
13.9 ± 1.1
4.8 ± 0.1
1.9 ± 0.6
0.91 ± 0.03
18.9 ± 0.6
1.12 ± 0.02
1.09 ± 0.05
91.86
35.6 ± 0.9
29.9 ± 1.3
13.8 ± 0.3
8.9 ± 1.1
4.1 ± 0.1
3.1 ± 0.6
1.32 ± 0.03
19.9 ± 0.6
2.92 ± 0.02
1.22 ± 0.05
90.8
36.6 ± 0.9
32.1 ± 1.3
13.6 ± 0.3
11.9 ± 1.1
4.6 ± 0.1
2.6 ± 0.6
1.25 ± 0.03
17.9 ± 0.6
1.91 ± 0.02

1.09 ± 0.05
90.4

BHEAA
2HEAP
2HEAB
2HEAPr
2HEAA
H2O
Untreated

Samples
Constituent

Table 2
The chemical characteristics of the untreated biomass and that subjected to treatment with protic ionic liquid.

BHEAPr

Although there are some studies on the solubility of lignin in ionic
liquids, little is known about the potential of these compounds to treat
biomass. Chemical characterizations performed before and after the
treatment with the ionic liquid are important to discern what changes
occur in the biomass as a result of treatment. The chemical compositions of the untreated and PIL-treated biomass samples are shown in
Table 2.
The composition of the lignocellulosic material in the untreated
biomass was 34.6% and 25.4% cellulose and hemicellulose, respectively, and 5.14% total lignin. In the untreated biomass, the total lignin
content was significantly lower than the other components of the lignocellulosic portion. After treatment with PILs, the quantities of cellulose and hemicellulose increased, while amount of lignin decreased.
Hemicellulosic sugars represented 15.43 ± 1.77% of the raw material,
with xylose as the main sugar (67%). The cellulose (as glucose) and

lignin content (39.97 ± 0.95% and 17.83 ± 0.05%, respectively)
were similar to those in other studies using sunflower (Díaz, Cara, Ruiz,
Pérez-Bonilla, & Castro, 2011; Pandey & Pitman, 2004; Ruiz et al.,
2013).
According to Mohanty, Misra, and Drzal (2001) increases in cellulose and hemicellulose fractions after treatment results from the removal of additional materials such as waxes and plant gums. Moreover,
there was an increase in the quantity of cellulose in the biomass treated
with the 2HEAPr (39.3%) compared with the biomass treated with
BHEAPr (38.6%). The amount of hemicellulose in the biomass was
higher after treatment with m-BHEAP (34.9%) than after treatment
with 2HEAPr.
Xylose was the predominant sugar in all treated and untreated
biomass samples. The amount of xylose extracted from the untreated
biomass was 21.9%, which decreased in biomass treated with PILs.
Biomass treated with HEAA showed lowest amount of xylose (12.9%).
The concentration of ash and acetyl groups also decreased after PIL
treatment. Ash and acetyl groups were present at 3.9% and 2.09%,
respectively, in the untreated biomass while the concentrations in
biomass treated with the BHEAPr was 1.1% and in that treated with IL
2HEAB, BHEAB and m-2-HEAB was about 12.09%.
Similar to the results of the present study, Singh et al. (2009) observed changes in the biomass composition of switchgrass after treatment with PIL 1-ethyl-3-methylimidazolium acetate ([C2mim]OAc).

37.9 ± 0.9
27.3 ± 1.3
12.8 ± 0.2
9.1 ± 0.1
4.6 ± 0.1
0.8 ± 0.05
1.14 ± 0.03
15.9 ± 0.6
3.31 ± 0.02

1.09 ± 0.01
86.64

3.1. Chemical characterization of biomass

39.3 ± 0.2
27.0 ± 0.3
11.6 ± 0.3
9.9 ± 1.1
4.6 ± 0.1
0.9 ± 0.6
1.04 ± 0.03
19.9 ± 0.6
1.31 ± 0.02
1.92 ± 0.05
90.47

BHEAB

The characterization of pineapple fibres before and after treatment
with protic ionic liquids for delignification is important because the
changes caused by PIL treatment affect the success of potential applications of PILs for biomass recovery.

37.8 ± 0.3
29.0 ± 1.3
10.6 ± 0.3
9.9 ± 0.3
6.6 ± 0.1
3.9 ± 0.2
2.14 ± 0.03

18.9 ± 0.6
1.91 ± 0.02
2.09 ± 0.05
91.84

BHEAP

3. Result and discussion

35.6 ± 0.5
27.7 ± 0.8
14.6 ± 0.5
8.2 ± 0.9
4.6 ± 0.3
0.3 ± 0.02
5.14 ± 0.03
21.9 ± 0.6
3.82 ± 0.04
1.19 ± 0.08

m-2HEAA

Furfural and hydroxymethylfurfural concentrations were analysed
using HPLC equipped with a SPD-M20 A Diode-array detector; the separation was performed using a LiChrospher 100 RP-18 (125 × 4 mm,
5 μm) column (Hewlett-Packard), operating at 25 °C, with acetonitrile/
water as an eluent at a flow rate of 0.5 mL min−1.

34.6 ± 0.9
25.4 ± 1.3
10.6 ± 0.3

8.9 ± 1.1
3.6 ± 0.1
1.9 ± 0.6
5.14 ± 0.03
21.9 ± 0.6
3.91 ± 0.02
2.09 ± 0.05
93.04

2.6. Quantification of toxic compounds

Cellulose
Hemicellulose
Xylose
Galactose
Arabinose
Mannose
Total Lignin
Extractive
Ash
Acetyl Group
Total

m-2HEAPr

m-2HEAB

m-2HEAP

transferred to 15 mL centrifuge tubes and 0.5 mL acetic acid was added.

The solutions were mixed well and 0.3 M sodium hydroxide (0.3 mL)
and 0.5 M hydroxylamine hydrochloride (0.1 mL) were added. The final
volume adjusted to 2 mL with acetic acid. The UV spectra of the solutions were measured against a blank prepared using the same method,
excluding the biomass. The lignin content was determined with the
absorbance at 280 nm and a mass extinction coefficient of 15.02 L
g−1 cm−1 (standard lignin) according to an established method (Arora
et al., 2010.

35.9 ± 0.9
34.9 ± 1.3
12.6 ± 0.3
11.4 ± 1.1
5.6 ± 0.1
4.9 ± 0.6
2.22 ± 0.03
17.9 ± 0.6
2.09 ± 0.02
1.07 ± 0.05
94.08

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R. de C.M. Miranda et al.


Carbohydrate Polymers 206 (2019) 302–308

R. de C.M. Miranda et al.

appearance were observed and the exposed pits emphasize the removal

of lignin by the PILs. In addition, PIL treatment preserved the cellulose
structure because of contact with the surface.
Fig. 1F–I shows the profile of fibres after treatment with ionic liquids BHEAA, BHEAPr, BHEAB, and BHEAP. The fibres are well preserved but show the presence of pits. In such treated fibres, conserved
cellulose is observed as rod-shaped fibrous structures, suggesting the
presence of pectin.
Fig. 1J–M depicts the profiles of the fibres after treatment with ionic
liquids m-2HEAA, m-2HEAPr, m-2HEAB, and m-2HEAP. Unlike previous treatments, the cellulose fibres do not appear to be preserved,
although pits are present. The fibres do not appear to be stretched, and
their appearance is not preserved compared with pre-treatment, suggesting an aggressive treatment compared with the PILs used in
Figs. 1J–M. The damage to the fibre denotes the presence of large
amounts of pectin, which may hinder the absorption of the PIL and the
removal of lignin for good cellulose exposure.
Brígida et al. (2011) treated coconut fibres with sodium hydroxide
and observed pits and fibre disorganization. Auxenfans et al. (2014)
reported a complex fibre organization characterized by a highly fibrillated morphology in untreated sawdust oak.
Treatment with the 12 PILs altered the organization of the fibres
inside the samples, resulting in a more irregular and porous texture.
Thus, these data suggest a strong change in the organization of primary
particles without any noticeable changes in their specific surface area.
These changes in texture create a large volume available among the
primary wood grains and therefore should improve the accessibility of
enzymes.

The authors reported lignin, cellulose and hemicellulose contents of
about 27%, 36%, and 37%, respectively, before treatment, while after
PIL treatment, the concentrations reached 27%, 34% and 39%, respectively. This could be explained by the fact that, after treatment with
the ionic liquid, the fibres become more exposed and porous.
Brớgida, Calado, Gonỗalves, and Coelho (2011) treated coconut bres with three different chemicals (NaOCl, NaOCl/NaOH, and H2O2)
and cellulose recovery increased to 62.77% when treated with NaOCl/
NaOH, compared with the 45.93% recovered from untreated fibre. The

authors attributed this to the partial removal of hemicellulose, which
was confirmed by the disintegration of the biomass. Perez-Pimienta
et al. (2015) used the NREL methodology to characterize agave bagasse
pre-treated with [C2mim]OAc and observed increased cellulose and
hemicellulose contents, and decreased lignin contents. The same profile
was observed for ash, with a higher content in the treated biomass.

3.2. Morphological characterization of the biomass
Scanning electron microscopy (SEM) was used to reveal morphological differences before and after biomass treatment. We evaluated
the changes in cell wall morphology after the application of PILs in a
similar way to previous microscopic studies (Sun, Li, Xue, Simmons, &
Singh, 2013). SEM images of pineapple fibres before and after PIL
treatment are shown in Fig. 1A–M. Fig. 1A shows micrographs of the
untreated fibres with a preserved structure without pores or pits. The
"pits" observed in Figs. 1B-M indicate the removal of lignin and cellulose exposure (Pereira, Voorwald, Cioffi, & Pereira, 2012).
Fig. 1B–E depicts SEM images of the fibre treated with the ionic
liquids 2HEAA, 2HEAPr, 2HEAB, and 2HEAP, which indicate the effectiveness of treatments. Elongated structures with a fibrous

Fig. 1. Micromorphological aspect of pineapple waste fibre without treatment (A) and after treatment with protic ionic liquids 2HEAA (B), 2HEAPr (C), 2HEAB (D),
2HEAP (E), BHEAA (F), BHEAPr (G), BHEAB (H) and BHEAP. (I), m-2HEAA (J), m-2HEAPr (K), m-2HEAB (L), and m-2HEAP (M).
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R. de C.M. Miranda et al.

Fig. 2. X-ray diffraction data of untreated biomass, microcrystalline cellulose (MCC

a−c

), and biomass treated with the protic ionic liquids

(d–o)

.

by the material treated with 2HEAPr (68%) and the ionic liquid BHEAB
(66%). According to George et al. (2015), an increase in the CrI is often
indicative of hemicellulose and/or lignin removal.
Enzinne, Reagan, Guoquing, Hanna, and Wesley (2014) carried out
recyclability experiments with cellulose in PILs and confirmed that the
recovered cellulose largely maintains its cellulose-I crystal structure
because of the low solubility of cellulose in the PILs.
Similar results were obtained by Zhang et al. (2014), who reported
an increase in the CrI of switchgrass, corn stove, and rice husk cellulose
after treating the biomass with PIL 1-butyl-3-methylimidazolium
acetate ([C4mim]OAc). In the present study, we used Energy Dispersive
X- Ray diffraction Spectroscopy (XRD) to determine the CrI of the cellulosic structure after treatment. After treatment of lignocellulosic
biomass of agave bagasse, Perez-Pimienta et al. (2015) reported that
cellulose I and II became more dissolved and hence more amorphous,
reducing the crystallinity,

3.3. Crystal index
The determination of the degree of crystallinity is important to
understand the behaviour of cellulosic materials, because these materials possess crystalline and amorphous regions. Crystallinity determination enables the observation of changes that occur in the structure of
the cellulosic material, both in the crystalline and amorphous region
(Pereira et al., 2012). X-ray diffraction analysis of untreated cellulose
and microcrystalline samples treated with PILs are shown in Fig. 2. The
samples treated with ionic liquids displayed peaks in the diffractograms

in the region 10° ≤ 2θ ≤ 20° and regions 18° ≤ 2θ ≤ 20°. These peaks
after the treatment with ionic liquids were similar to microcrystalline
cellulose as all of them had a peak at 2θ = 22.1°. This peak probably
indicates the distance between hydrogen-bonded sheets in cellulose I,
as reported previously. After treatment with PILs, the crystallinity of
the samples increased when compared to untreated samples, even
though the peaks at approximately 38, 44, 65, and 78° do not represent
cellulose, and are probably characteristics of lignocellulosic samples,
because the profile is different from the MCC sample and similar to that
of the untreated sample.
Diffraction data were used to determine the Crystallinity Index (CrI)
using Eq. 1, and the results are shown in Fig. 3. The highest CrI was
observed after treatment with the ionic liquid BHEAPr (70%), followed

3.4. Cellulose, Hemicellulose, lignin, and toxic compounds quantification
The cellulose, hemicellulose, and total lignin removed from the
lignocellulosic material by treatment with the PILs are shown in Fig. 4.
PILs were demonstrated to be efficient in the removal of lignin and
hemicellulose, while preserving the pulp, and thus have the potential to
treat lignocellulosic biomass.
Among the 12 PILs tested, the highest efficiency was demonstrated
by 2HEAPr, (lignin and hemicellulose removals of 92% and 48%, respectively). The PIL m-2HEAPr removed 89% of the lignin, 34% of the
hemicellulose, and only 0.9% of the cellulose. The PIL BHEAA, removed
83% of the lignin, 33% of the hemicellulose, and 1.2% of the cellulose.
While 2HEAPr, m-2HEAPr, and BHEAA showed the best performances for delignification, all the other tested PILs showed similar
trends when treating lignocellulosic biomass from pineapple crowns.
All PILs removed almost all the lignin and hemicellulose without interfering with the cellulose content. In Fig. 4, higher lignin removal was
observed for PILs with the propionate anion.
The calculated octanol-water partition coefficients (log P values) for
the cations are −1.32, −1.57, and −0.88 for ethanolamine, diethanolamine, and methylmonoethanolamine, respectively. Considering the

entire PIL compound, when the filaments are under the compound it is
more hydrophilic. For the larger chains, the hydrophobicity of the alkyl

Fig. 3. Crystallinity index of microcrystalline cellulose (MCC), the untreated
biomass, and biomass after protic ionic liquids treatment.
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R. de C.M. Miranda et al.

Fig. 4. Cellulose, hemicellulose, and lignin removed from the lignocellulosic material after treatment with the protic ionic liquids.

reacted with water to produce guaiacol at 150 °C. Achinivu et al. (2014)
developed a lignin extraction method from lignocellulosic biomass
using PIL, and observed a positive correlation between xylan solubility
in the IL and fibre disruption/penetration.
Studies have explored the effect of cations on the action of PILs.
George et al. (2015) studied the effect of protic and ionic liquids on
saccharification and reported that treatment with diethyl-, triethyl-,
and diisopropylammonium ILs resulted in higher saccharification yields
and similar performances. It appears that the overall trend in that work
was that the addition of eOH groups to the cation reduced the hydrolysis yield and an increase in the number of alkyl chains increased
the yield of enzymatic hydrolysis. The performance of the IL diisopropylammonium indicated that steric effects do not play a role in the
hydrolysis efficiency, at least for cations with short alkyl chain lengths
(n = 2–3). Rocha, Costa, and Aznar (2014) used BHEAA to pretreat
sugarcane bagasse for enzymatic hydrolysis, with promising results.
A problem in current lignocellulosic refineries is that conventional
treatments use organic solvents and buffers (basic and acid), during

which toxic intermediates are generated, such as hydroxymethylfurfural (HMF) and furfural (MF). In the present study, MF and
HMF levels were measured after treatment of the biomass with the PIL
to indirectly assess the toxicity of the process.
As shown in Fig. 5, the yields of HMF and MF during treatment of
biomass with PILs were low. Treatment with the PILs produced the
following yields of MF: 2HEAA - 5%; 2HEAP - 4%; BHEAP - 8%; m2HEAA - 1%; m-2HEAPr - 5%; and m-2HEAP - 6%. HMF was produced
by 2HEAA (2%) and BHEAP (2%).
Brandt et al. (2011) reported that the concentration of hemicellulose decreased as a result of treatment, suggesting the conversion
of carbohydrate monomers to furfurals. The production of MF and HMF
shown in Fig. 5 is expected, as high temperatures were used in the
present study (120 °C).
Sharaf, Mehrez, and Naggar, (2018) studied the preparation of bee
honey extracts using cellulose nanofibres as the immobilizing agent.
The authors reported eco-friendly methods for extracting honey, stating
that they obtained good results using ultrasound, soxhlet, and magnetic
stirring for propolis extraction. The authors further stated that the extract in the cellulose nanofibre was prepared using an environmentally
friendly solvent. El-Naggar et al. (2018), using microcrystals of cellulose for the elaboration of nanogees with the aim of using them for a
heavy grating. The authors state that they are more efficient for use as a
continuous process.

Fig. 5. Percentage of intermediate toxic compounds produced during the
treatment of lignocellulosic biomass with protic ionic liquids.

chain is predominant. Among all the PIL, BHEAPr showed highest lignin
solubilisation because of intermediate solubility.
Cellulose did not seem change when the biomass was treatment
with the PILs. Fig. 5 shows that 2HEAA had the greatest solubility for
cellulose (5%). According to Brandt et al. (2013), lengthening the alkyl
chains of the cation progressively reduces cellulose solubility.
The ionic liquid behaviour has been explained by Pu, Jiang, and

Ragauskas (2007), who showed that lignin is more soluble in the PILs
than in AILs because of the higher affinity of lignin for the protic ILs.
This is the results of the nature of the anion synthesized by proton
transfer between an equimolar mixture of a Brønsted acid and Brønsted
base. Cox and Ekerdt (2012) suggested that acidic ILs are successful at
breaking down lignin model compounds by hydrolysing the β-O- 4
ether bond; while the acidic environment of the IL catalyses the hydrolysis reaction. The anions have a significant effect on the yield and
the observed intermediates. Brandt et al. (2013) reported that solubility
seems to be strongly affected by the choice of anion, although hydrogen-bond basicity does not need to be as high as that for cellulose;
some intermediate-chain basic ionic liquids seem to be better solvents
for lignin than their basic relatives with more hydrogen-bonds. The
authors also emphasized that a protic cation failed to solubilise cellulose in many cases because of strong interactions between cations and
anions. For cellulose solubility, the cation should be based on a strong
base and a weak carboxylic acid, such as acetic acid or propionic acid.
Perez-Pimienta et al. (2015) reported that the differences in delignification efficiencies during pretreatment of agave biomass with
ionic liquids could be attributed to specific interactions of ionic liquids
with biomass factors (cation, anion, temperature, and time), and the
extent and degree of recalcitrance of the biomass factors (age, method
of harvesting, drying point, and storage conditions). Jia et al. (2010)
reported that the in process of cleavage of βeOe4 bonds of lignin
model, compounds that conserved 70% of the βeOe4 bonds of both
guaiacylglycerol-β-guaiacyl ether and veratrylglycerol-β-guaiacyl ether

4. Conclusion
Treatment of lignocellulosic fibres with ionic liquids was effective to
remove lignin and hemicellulose by exposing the cellulose, thereby
increasing the surface area of the fibres and providing free hydroxy
groups. The presence of cellulose increases the potential use of this fibre
307

Carbohydrate Polymers 206 (2019) 302–308

R. de C.M. Miranda et al.

when free OH is crucial. Small amounts of furfural and hydroxymethylfurfural were produced, demonstrating the low toxicity of
PILs. The PIL BHEAPr showed the greatest efficacy (90%), maintaining
the pulp, as observed in the morphological analysis and the calculation
of the crystallization rate. One of the objectives of delignification is the
removal of lignin present in the cells, to obtain inputs that could be
used in industry, such as in biofuels. In this context, the treatment of the
pineapple fibre (crown) to remove lignin using PILs proved effective to
obtaining a support for enzymatic immobilization by preserving the
cellulose. Thus the PIL-treated fibre could be used in industry for the
immobilization of enzymes, among which lipase is used as a catalyst in
the transesterification process for the production of bio-diesel.

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Declarations of interest
None.
Acknowledgments

The authors are grateful financial support from Conselho Nacional
de Desenvolvimento Cientớco e Tecnolúgico CNPq, Fundaỗóo de
Amparo a Pesquisa e Inovaỗóo Tecnolúgica do Estado de Sergipe
FAPITEC, and Coordenaỗóo de Aperfeiỗoamento de Pessoal de Nớvel
Superior CAPES for the scholarship of R.C.M. Miranda and Á. S. Lima.
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