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renewable energies [13] and support agro-industry within various en
ergy, economic, and geographical settings [14], so that more corporate
benefits should be obtained. Recycling various materials and using
renewable energy would contribute to the success [15]. Several inter
national organizations are committed to developing networks that guide
renewable energy policies, such as the Organization for Economic
Cooperation and Development (OECD), the European Union (EU), the
Slovakia Market Overview [16], and the International Energy Agency
(IEA) [17]. Bioenergy represents a replacement for fossil fuels in the
short- and medium-term, and contributes to mitigating greenhouse gas
emissions [18].
Production of biofuels has increased worldwide in emerging econo
mies, featuring productive approaches, which take advantage of mi
croorganisms [19], enzymes, incubation conditions, and engineering
schemes for efficient depolymerization [20]. Other results in biomass
experiments demonstrate that sodium carbonate pretreatment consid
erably improves ethanol concentration, with high levels of solids and
low enzyme loads [21]. Da Costa, Pereira, and Gomes [22] pointed out
that biorefinery based on residual and agro-industrial biomass generates
a variety of molecules. Consequently, in addition to their positive effects
as an alternative and ecological energy source [23], biorefineries can
enable industrial facilities to reduce their impact on public health and
the environment [24].
Our review paper addresses the study of bioethanol, and we will refer
to bioethanol as a biofuel. The terms ethanol and bioethanol are often
used alternatively. While ethanol is manufactured from petroleum by
hydration of ethylene, bioethanol is produced from existing or renew
able biomass. Ethanol is also known as petroleum-derived ethanol, and
signicant amounts of ethanol made in the world are derived from pe
troleum, while bioethanol represents renewable ethanol. Chemical
composition of petroleum-derived ethanol and bioethanol is the same.
Nevertheless, they are isotopically different. Bioethanol contains 14
C, in
the same ration as in its feedstocks, while there is no 14
C content
petroleum-derived ethanol given that all 14
C decays over time.
In this context, the main objective of this review paper is analyzes
perspectives of manufacturing bioethanol via biotechnology, using
sustainable management. Two categories are emphasized: (i) agro-
industrial valuation, which represents alternative methods for bio
ethanol production, and (ii) Agro-industrial Techniques and Infrastruc
ture Analysis. Our review paper examines in an updated manner one
crucial research question: the state of knowledge on bioethanol. We
report alternative methods of bioethanol production and address the
technology and industrial infrastructure used within a sustainable
scenario. Discussed are the gaps in each examined group. To answer
these questions, scientific literature was screened and evaluated sys
tematically and organized into two categories, as described below
(section 2).
The study is divided into seven sections. Section 1, The complete
introduction that relates the different indications of the growing need
for alternative energy, alternative methods of biofuel production and the
entire scenario of sustainable industrial production, section 2, presents a
general literature review, highlighting two categories main. Section 3
includes the methodological reference framework used to develop the
systemic review. Section 4 considers the alternatives for the production
of bioethanol from its advances by generation. Section 5 highlights
alternative industrial methods, such as biorefineries, the importance of
sustainable production processes and the importance of bioethanol
production from an energy perspective for the leading countries in
production, while section 6 directs us to recognize the future of bio
ethanol production. It ends with the main conclusions of the review
study in section 7.
2. General literature review
2.1. Agro-industrial valuation: Alternative methods for bioethanol
production
Agribusiness generates about 330 million metric tons of biomass
waste per year [25]. Therefore, the corporate world values renewable
energy sources to produce environmentally sustainable biofuels [26]. In
this framework, chemical processes carried out in the agro-industrial
production sector include clean technologies, such as bioprocessing,
pyrolysis, gasification, and Fischer-Tropsch synthesis, the latter being
used in a variety of processes for converting biomass to liquid [27]. Also,
chemical catalysis in Guerbet reaction yields molecules of great
complexity [28].
Aro [29] highlighted similar implications, when using selected plant
materials and evaluating the presence of microorganisms [30–32].
Agro-industrial biodiesel processing utilizes as raw materials oils plants
and microalgal biomass [33,34]. On their turn, oils are obtained by
distillation, pyrolysis, gasification, or catalytic processes. Hence,
agro-industrial valorization of the waste biomass represents a priority in
biofuel production at this scale [35].
A complementary study by Mendes et al. [36] identfied the role of
hydrolysis and esterification reactions in treatments with filamentous
fungi. For this, appropriate fungi were previously isolated from leaves,
and decomposed in aqueous environment. Lipases offer additional value
via their transesterification reactions. Assessed were the ethanolysis of
native oils in Brazilian Amazonian plants, such as andiroba (Carapa
guianensis), babassu (Orbignya sp.), Jatropha (Jatropha curcas), and palm
(Elaeis sp.) [37].
Escobar et al. [35] managed to isolate 1016 lipolytic microorganisms
that were analyzed by colorimetric assay. For further characterization,
the authors selected 30 initial lipolytic strains. Next, they performed
phylogenetic analysis and quantified the production of biofuels by a gas
chromatography. Agro-industrial wastes have proven rich in sugars,
minerals, and proteins [38–40]. Owing to the potential of such wastes to
promote efficient growth of microorganisms, they should be considered
as alternative raw materials with high added value for reuse [41].
2.2. Agro-industrial Techniques and Infrastructure Analysis
Technical capabilities and infrastructure in the agro-industrial pro
duction of biofuels, along with the relevant technologies and regula
tions, represent a continuous challenge [42], as one should also comply
with energy security and environmental sustainability [43]. To accom
plish these requirements, various approaches are adopted, which
consider both market dynamics and economic risks [44]. Benefits ob
tained in biofuel production depend on the raw material, the conversion
List of abbreviations
C. shehatae Candida shehatae
E. coli Escherichia coli
HRAP High raceway algal pond
1G First generation
2G Second generation
3G Third generation
LHC Lignin-hemicellulose-cellulose
CSLF Cellulose solvent-based lignocellulose fractionation
CbU/g Cellobiase unit per g
FPU/g Filter-paper units per g
EFB Empty fruit bunch
IEA Agency Energy International
SSF Simultaneous saccharification and fermentation
SHF Separate hydrolysis and fermentation
S. cerevisiae Saccharomyces cerevisiae
OECD Economic Cooperation and Development
EU European Union
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3. Renewable and Sustainable Energy Reviews 160 (2022) 112260
3
route, and the local context [45]. Also, transportation costs need to be
considered [46]. This productive system should, however, be inter
preted not only from the standpoint of technology and sustainability
[47,48]. Controversies typically arise from the global impact of national
biofuel systems, food security problems, environmental concerns, limi
tations of local territories, technologies, and economies [49].
Effective technical management of productivity and profitability in
biofuel production requires intelligent monitoring and control tools
[50]. Given that new technological discoveries are made and better
agro-industrial appropriation techniques evolve one must pay attention
to raw materials, processing, conversion efficiency, manufacturing of
co-products, and sustainability [51].
All the above include critical gaps and contain a series of factors that
should to be related to uncertain economic processes and costs [52].
Cost increase is caused mainly by biomass mobilization over more
considerable distances or failures in industrial quality control [53]. By
contrast, cost reduction can occur as a result of cutting back in size [54].
Net profit generation from biofuel production would be triggered by
adopting new agro-industrial policies [55]. Likewise, production could
be increased by the in-depth study and better understanding of value
chains and by implementing complex adaptive systems in small-scale
biofuel production [56,57].
3. Methodological framework and study design
According to Beltrán’s recommendation [58], for a systematic liter
ature review (SLR ) information should be retrieved by articulating one
straightforward question. Next, one should collect all available evidence
for answering the question [59]. Therefore, the SLR should follow the
steps listed below [60]: (1) create the research question and objective,
(2) locate scientific articles, (3) select the most relevant papers for in
clusion, (4) evaluate the quality of primary studies, (5) extract data, and
(6) analyze data.
Our research question referred to the state of knowledge on bio
ethanol production from an industrial and biotechnological manage
ment perspective. Two categories were considered: (i) agro-industrial
valuation: alternative methods for bioethanol production, and (ii) Agro-
industrial Techniques and Infrastructure Analysis. This approach
allowed to establish the following objectives of the research: (1) eval
uate the agro-industrial methods in bioethanol production and (2) un
derstand the need for technological advances and agribusiness
infrastructures for the production of bioethanol in a sustainable manner.
For creating our database we considered journals indexed by Scopus
and Web of Science. Explored time frame spanned from year
2010–2021. Our study design was based on a validated methodology
[61]. We adopted a boolean type search strategy, using the following
keywords: solid, liquid, and gaseous bioethanol; agro-industrial valori
zation of bioethanol processes; and techniques in agro-industrial
infrastructure.
An interpretive synthesis using a contingent or sequential review of
the information was done, which addressed the main topics of reviewed
papers [62]. Data analysis was performed based on information ac
quired from each established category, with its corresponding
subcategories.
4. Agro-industrial valuation: Alternative methods for bioethanol
production
In this category, the main features, which describe each of the sub
categories are listed below.
4.1. Raw edible substances, biomass, and waste
While non-renewable sources of fossil fuels are dependent on econ
omy, biomass is valued for its facile processing and for offering oppor
tunities toward desirable sustainable development. Consequently,
biomass represents a practical and adaptable alternative for the agri
business market [63].
Jeguirim and Limousy [64] proposed the agro-industrial use of
by-products and waste streams. Produced bioethanol has been catego
rized into three generations, as displayed in Fig. 1.
Manufaturing of first-generation (1G) bioethanol affords high yields
[65], when using as raw materials cane sugar, corn, and beet sugar. In
the meanwhile, bioethanol extraction methods have evolved techno
logically into new industrial processing methods that utilize diverse
sources of cellulosic raw materials including wood, grass, and other
organic materials [66].
In present, most bioethanol produced worldwide belongs to first-
generation, being obtained by fermentation from sugar-based edible
raw substances, usually juice from sugarcane, or starchy biomass, e.g.,
potato, corn, grains, and seeds [67]. A simple step is required to produce
1G bioethanol; feedstocks like sugarcane juice and molasses need a
simple fermentation process, while starchy biomass necessitates hy
drolysis followed by the fermentation process [68]. Because of the
absence of solid particles in the feedstock (when processing sugarcane
juice and molasses), after completing the fermentation process, yeast
can be separated and recycled, i.e., reused in the next batch. Eventually,
yeast recycling reduces the production cost of bioethanol and the pro
cessing time of fermentation [29]. Nonetheless, starch-based feedstocks
require liquefaction and saccharification steps to complete the hydro
lysis. Depending on the end products in corn processing one commonly
use two types of liquefaction processes. Dry-milling liquefaction is
suitable for small-scale industries only, aiming for bioethanol produc
tion as the final product.
In contrast, the wet-milling process of liquefaction is popular for
large scale industries, which target along with bioethanol the production
of its co-products, as well, such as corn syrup, glucose syrup, and
dextrose syrup [29]. A schematic representation of bioethanol produc
tion from sugar-based and starch-based biomass is displayed in Fig. 2. It
reveals that various feedstocks undergo similar fermentation and bio
ethanol recovery processes; nonetheless, there are several approaches to
obtain different fermentable sugars and to prepare various co-products.
Overall, the crops used as feedstocks for bioethanol will result in food
versus fuel implications.
Production of second-generation bioethanol takes advantage of a
large variety of non-edible agricultural and industrial lignocellulosic
wastes, including rice husk, wheat straw, corn stalks, olive pomace,
bagasse, coconut husk, and paper pulp industries waste, or even fruit
peels [69–74]. A list of lignocellulosic wastes and production of bio
ethanol is compiled in Table 1.
Cellulose in lignocellulosic materials is not suitable for direct hy
drolysis because of the complex structure of lignin-hemicellulose-
cellulose (LHC) (Fig. 3). Hence, it is necessary to remove the envelope
of lignin and hemicellulose surrounding the cellulose, which requires an
extra step, commonly known as pretreatment. Different sources of
lignocellulosic biomass have variations in LHC composition. In addition
to LHC differences, various sources of lignocellulosic biomass contain
dissimilar secondary metabolites, such as waxes, lipids, tannins, ter
penes, alkaloids, and resins. Secondary metabolites in plants are mainly
for defense purposes, helping them to survive. During enzymatic hy
drolysis and fermentation processes of cellulose extracted from ligno
cellulosic wastes, the presence of secondary metabolites deactivates the
enzyme or kills the microorganisms that promote the fermentation
process. Therefore, the composition of wastes from agricultural biomass
must be analyzed prior to using them as raw materials for bioethanol
production.
In general, during the pretreatment step, most of the secondary
metabolites are removed from the biomass. Bensah et al. [93] success
fully enhanced bioethanol production by eliminating cuticular and
epicuticular waxes from wheat straw waste; for this, plasma-assisted
pretreatment was employed. Liu et al. [94] de-waxed palm oil-empty
fruit bunch waste by Soxhlet extraction, using a solvent mixture of
J.R. Melendez et al.
4. Renewable and Sustainable Energy Reviews 160 (2022) 112260
4
toluene/ethanol (2:1, v/v). To remove wax, Kristensen et al. [95] treated
wheat straw waste taking advantage of a hydrothermal process. High
value terpenes, such as citral and geraniol were extracted from the
lignocellulosic waste of lemongrass and palmarosa, by steam distilla
tion. Terpene removal was necessary, as it would inhibit microbial ac
tivity of Saccharomyces cerevisiae during fermentation [96].
Citrus peel represents a suitable lignocellulosic waste for bioethanol
production owing to its low lignin and high fermentable sugar content.
Its D-limonene terpene hinders, however, the growth of yeast over the
fermentation process and lowers the amount of produced bioethanol.
Reported procedures include steam distillation [97–99], acid-catalyzed
assisted steam explosion [100], hydrothermal process [101], and
auto-hydrolysis [102]. All these techniques were intended to recover
limonene from citrus fruit peel wastes, so that microbial activity during
fermentation can be resumed. Peretz and co-workers [103] removed
tannic acid and lignin simultaneously by ozonation. Lignin hinders
enzymatic hydrolysis and fermentation, therefore, removing lignin from
biomass prior to hydrolysis of cellulose is mandatory.
Lignocellulosic processing industry considered lignin as a waste,
although high-quality lignin in abundance should be used as a
biodegradable polymer [104]. This is why, bioethanol production and
concurrent recovery of lignin, as a biodegradable material, represent a
waste-to-worth paradigm.
Other production alternatives are focused on industrial manage
ment, which considers raw materials that provide high yields [105]. In
this approach a combination of first-generation methods are use, based
on the extraction of sugar cane juice, and second-generation technolo
gies, which use raw materials derived from cellulose or lignocellulose
biomass [106]. Tapia Carpio and Simone de Souza [107] suggested that
the technological integration of 1G and 2G bioethanol production can
reduce costs of operation and investment risks.
Third-generation bioethanol is produced from microalgae, which are
primarily photoautotrophic in nature. Several species can undergo a
metabolic shift to change their mode from photoautotroph to hetero
trophs, when environmental conditions change. This allows microalgae
to withstand extreme conditions. They can accumulate substantial
amounts of carbohydrates in starch, cellulose, hexoses, and pentoses,
which are being converted into fermentable sugars to produce bio
ethanol. One should also consider that different species of microalgae
accumulate different amounts of carbohydrates (see Table 2).
Fig. 1. Three generations of bioethanol production based on the feedstock. Developed by the authors based on reference [64].
Fig. 2. Schematic diagram comparing first-generation bioethanol production from (A) sugar based and (B) starch based feedstocks. Developed by the authors based
on reference [29].
J.R. Melendez et al.
5. Renewable and Sustainable Energy Reviews 160 (2022) 112260
5
Table 1
Bioethanol production from lignocellulosic agricultural and industrial wastes.
Agricultural wastes Technical Descriptions Source
Wheat straw 37.0 g/L bioethanol obtained by subcritical
water pretreatment (extraction at 220.5 ◦
C;
extraction time, 22.0 min) combined with
separate high solid (15%) hydrolysis and
fermentation (SHF).
[75]
Whole plant cassava Bioethanol obtained by hydrothermal
pretreatment (180 ◦
C; 2 MPa; 60 min)
followed by fermentation of integrated
cellulosic C5 sugar and starch from whole
plant cassava, in a simultaneous
saccharification and fermentation method
(SSF).
[76]
Wheat and rye stillages Microwave-assisted pretreatment with
dilute acid of wheat and rye stillages to
produce >156 mg/g glucose at microwave
power of 300 W (15 min, 54 PSI in 24 h
process), while after 48 h of fermentation
using S. cerevisiae, 20 g/L of bioethanol was
obtained.
[77]
Cotton stalk Cotton stalk was pretreated in organosolv
and hydrothermal processes, followed by
pre-hydrolysis using 80 FPU/g cellulose (at
50 ◦
C, at pH 5.0, for 6 h). 15 mg yeast per
gram of dry pretreated cotton stalk allowed
to carry out the fermentation process at
30 ◦
C with an initial pH 5.0. By this, 47.0 g/
L bioethanol were obtained.
[78]
Sugarcane bagasse Sugarcane bagasse was pretreated by
hydrodynamic cavitation to assist alkaline-
hydrogen under optimized conditions, e.g.,
0.29 M of NaOH, and 0.78% (v/v) of H2O2
(9.95 min process time at 3 bar inlet
pressure). Doing so, 95.4% digestion of the
cellulosic fraction was achieved. Cellulase
enzyme was used for hydrolysis, followed
by fermentation with Scheffersomyces stipitis
NRRL-Y7124. Hence, 31.50 g/L of
bioethanol were produced.
[79]
Eucalyptus biomass Alkaline extrusion pretreatment at 150 ◦
C
allowed to obtain the highest glucan and
xylan conversion in bioethanol production
via enzymatic hydrolysis, namely close to
40% and 70% yields, respectively.
[80]
Sugar cane bagasse For pretreatment, 5% Na2CO3 solution was
used, at 140 ◦
C in 1 h. During hydrolysis
97.6% glucose was obtained. Next,
fermentation at 37 ◦
C for 72 h produced
7.27 g/L bioethanol.
[81]
Olive pruning Waste biomass was soaked separately in
aqueous 1%, wt./wt., H2SO4 solution for 30
min, and then loaded into the steam
explosion reactor. For pretreatment, the
mixtures were heated with steam at 195 ◦
C
for 10 min. Next, the biomass was processed
by SSF method, using for hydrolysis, and for
the fermentation a cellulolytic enzyme
cocktail Cellic CTec2 and high-ethanol-
tolerant industrial Saccharomyces cerevisiae
strain “Ethanol Red”. Highest bioethanol
concentrations reached were of 47.8 g/L for
almond shells, 41.1 g/L for olive pruning,
and 21.4 g/L for vineyard pruning.
[82]
Vineyard pruning
Almond shells
Jerusalem artichoke
stalks
5% Nitric acid was used for pretreatment,
followed by enzymatic hydrolysis with
Cellic CTec2, a blend of cellulases,
β-glucosidases, and hemicellulases. Then,
fermentation was performed with
Saccharomyces cerevisiae strains. The
concentration of bioethanol produced
without solid residue was 3 times greater
(1.5 g/L) than in the presence of solid
residue (0.5 g/L).
[83]
Corncob Pretreated corncob was fermented with
Spathaspora passalidarum U1-58, which
[84]
Table 1 (continued)
Agricultural wastes Technical Descriptions Source
utilized hexoses and pentoses. Two
approaches, SHF and SSF were compared.
SSF afforded higher yield of bioethanol than
SHF, which were reported after 96 h as
42.46 g/L and 53.24 g/L, respectively.
Potato peels wastes 0.5% HCl was used in the acid hydrolysis
process, followed by fermentation with
commercial and genetically modified
S. cerevisiae. 2 g/L S. cerevisiae has proven
the optimum concentration of inoculum for
fermentation, yielding 2.83% and 2.64%
bioethanol by commercial and genetically
modified S. cerevisiae, respectively, within
3–4 days.
[85]
Green coconut husk
fibers
Alkaline method for pretreatment,
enzymatic hydrolysis, and S. cerevisiae
fermentation were used. Bioethanol
conversion efficiency was as high as 59.6%
of the fermentable sugars.
[71]
Extracted Olive Pomace Extracted olive pomace was pretreated with
dilute HCl and followed by SHF and SSF
processes, covering enzymatic hydrolysis
and fermentation. SSF showed better
fermentation yield (0.46 g/g) than SHF
(0.36 g/g) in 72 h process.
[72]
Empty fruit bunch SHF and SSF methods were compared to get
a better yield to bioethanol. The empty fruit
bunch was pretreated with 10% NaOH (aq)
at 150 ◦
C for 30 min. 4.74% of bioethanol in
72 h and 6.05% of bioethanol in 24 h
fermentation, respectively, were obtained
by SHF and SSF processes.
[86]
Empty fruit bunch 1% NaOH sol was used for pretreatment,
which reduced 90.3% lignin. Obtained
cellulose was hydrolyzed by adding
xylanase and cellulase at pH 6, for 6 da,
which released 19.3–20.6% product. Next,
fermentation with S. cerevisae proceeded in
two days, yielding 540–655 mL/3.82–4.63
kg EFB at the pilot scale.
[87]
Eucalyptus wood In a bench-scale study of bioethanol
production from eucalyptus, wood was
pretreated in a hydrothermal process
(150 ◦
C, for 4 h). Cellulase (20 FPU/g
substrate) was used for hydrolysis and
incubated at 50 ◦
C for 72 h. SHF approach
was used with S. cerevisiae strain for
fermentation. 53.5 g/L of bioethanol was
produced in 72 h.
[88]
Rice straw Maximum glucose yield of 93.6% was
obtained using CO2-incorporated ammonia
explosion pretreatment, under optimized
conditions: 14.3% ammonia, 2.2 MPa CO2,
at 165.1 ◦
C for 69.8 min residence time.
97% bioethanol were obtained by
simultaneous saccharification and
fermentation method (SSF).
[89]
Paper shredder scrap No pretreatment step was reported. Scrap
was hydrolyzed by two methods: (i) direct
enzyme hydrolysis and (ii) sulfuric acid
treatment followed by enzymatic
hydrolysis. Direct enzyme hydrolysis
provided a better yield of glucose (750 mg/
g). A novel Ethanol Trap System was used to
increase bioethanol production. 12% w/v of
bioethanol was obtained by fermentation
with marine strain Saccharomyces cerevisiae
C-19.
[90]
Miscanthus sp. (Plants
from grass family)
Biomass was pretreated with NaOH (at
145.29 ◦
C for 28.97 min, with 1.49 M
aqueous NaOH solution) followed by
enzymatic hydrolysis (50 FPU/g cellulase
and 30 CbU/g β-glucosidase), which
provided 83.9% glucose conversion.
Maximum bioethanol yield (59.2 g/L) was
obtained in fermentation with S. cerevisiae.
[91]
(continued on next page)
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6. Renewable and Sustainable Energy Reviews 160 (2022) 112260
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Microalgae can easily be cultured on a large scale in HRAP (high
raceway algal pond). The process of producing bioethanol from micro
algal biomass is the same as for cellulosic materials, including pre-
treatment of the biomass, hydrolysis, fermentation, and product recov
ery (see Fig. 4). Nonetheless, there are plenty of different options to
utilize microalgal biomass for other purposes, such as bio-fertilizers,
animal feeds, nutraceuticals, etc.
4.2. Pretreatment and hydrolysis of carbohydrates
Typically, pretreatment is an expensive step, which represents
almost 40% of the total production cost [109]; concomitantly, excessive
use of chemicals is indeed environmentally unfriendly. Biological pre
treatment could be a better option, and extensive research was per
formed to degrade the complex structure of LHC and for liberating
cellulose by using white rot, brown rot, and soft rot fungi [110,111].
Zhang et al. [112] successfully pretreated lignocellulosic bamboo ma
terial using white-rot fungi. Though biological pretreatment is envi
ronmentally friendly and works under mild conditions, the use of
microbes for pretreatment is still not the preferred choice for bioethanol
production, because of its poor performance (it releases less cellulose),
low rate, and multiple products formed (lignin and phenolic com
pounds), which inhibit the microbial activity [113]. Different detoxifi
cation processes are being developed, which aim to reduce the
concentration of the inhibitor. Unfortunately, this involves extra costs.
Inhibitor-resistance or acclimatized microbial strains are being
investigated to overcome the unwanted effects of inhibitors [114,115].
In earlier literature (beyond the scope of this review), there were
handful reports on the use of biological pretreatments to remove lignin
and to expose cellulose. These processes require 4–6 weeks, depending
on the type of biomass wastes and microorganisms [116–120]. Hence, it
is more practical to adopt physical and chemical methods for pretreat
ment, rather than biological procedures.
After pretreatment, hydrolysis of carbohydrates (cellulose and
hemicelluloses) represents the second crucial step in bioethanol pro
duction. There are several methods (both chemical and biological) to
hydrolyze carbohydrates; nonetheless, biological methods are more
appropriate to break down cellulose and hemicellulose into simple
monomers. There are some advantages and limitations associated with
both biological and chemical hydrolysis, as shown in Table 3. Since this
review focuses on environmentally friendly methods of bioethanol
preparation, we discuss in detail the only biological method reported, so
far. Hydrolysis of carbohydrates is also known as saccharification, where
fermentable sugars are produced. In biological methods, extracted en
zymes are directly used to produce fermentable sugars.
A large portion of hemicellulose can be removed in a successful
pretreatment process, leaving behind mostly cellulose for hydrolysis or
saccharification. Within cellulose, amorphous sites of cellulose are more
susceptible to enzymatic attack than its crystalline sites. The high con
centration of glucose generated from amorphous sites causes enzyme
inhibition and, eventually, prevents saccharification of crystalline cel
lulose. This issue can be overcome by a one stage hybrid process,
commonly termed as simultaneous saccharification and fermentation.
Cellulose and hemicellulose are hydrolyzed by cellulases and hemi
cellulases enzyme-producing glucose and pentoses (xylose and
Table 1 (continued)
Agricultural wastes Technical Descriptions Source
Miscanthus (Illinois
clone)
Pretreatment was done with 72% w/w
H2SO4 at 30 ◦
C for 60 min to extract
cellulose, known as Cellulose solvent-based
lignocellulose fractionation (CSLF)
pretreatment. Extracted cellulose was
hydrolyzed with cellulase. Miscanthus
(Illinois), giant reed, miscanthus (Q42641),
elephantgrass, and sugar cane yielded a
greater amount of glucose per gram of
biomass, which ranged from 0.290 to 0.331
g/g, as compared to rice husk and soybean,
which provided 0.181 g/g and 0.186 g/g,
respectively. The fermentation process was
performed with self-flocculating yeast strain
(SPSC01), which produced <0.068 g/g for
rice husk and soybean litter.
[92]
Giant reed
Miscanthus (Q42641)
Elephantgrass
Sugar cane
Rice husk
Soybean litter
Fig. 3. Schematic diagram of the complex structure of lignin-hemicellulose-cellulose.
Table 2
% Carbohydrate of the dry weight of microalgal biomass in different species
[108].
Microalgal Species % Carbohydrate of dry wt. biomass
Anabaena cylindrical 25–30
Chlorella pyrenoidosa 26
Chlamydomonas rheinhardii 17
Chlorella vulgaris 12–17
Dunaliella bioculata 4
Dunaliella salina 32
Euglena gracilis 14–18
Porphyridium cruentum 40–57
Prymnesium parvum 25–33
Scenedesmus dimorphus 21–52
Scenedesmus obliquus 10–17
Spirogyra sp. 33–64
Spirulina maxima 13–16
Spirulina platensis 8–14
Synechoccus sp. 15
Tetraselmis maculate 15
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arabinose) and hexoses (glucose, galactose, and mannose), respectively
[110]. These enzymes are highly specific and operate under mild con
ditions (at 45–50 ◦
C and pH 4.8) [121]. Three main groups of cellulases
are involved in the hydrolysis of cellulose: endoglucanase, exoglucanase
(cellobiohydrolase), and β-glucosidase. Endoglucanase breaks
non-covalent bonds present in amorphous cellulose, while exoglucanase
hydrolyzes the free end of the cellulose chain in the crystalline region
into disaccharides. β-Glucosidase hydrolyzes disaccharides into glucose
[122,123]. Hemicelluloses are present in nature in complex and het
erogeneous forms; hence, many hemicellulases are available to hydro
lyze the polymer’s side groups and main backbone.
Xylanases, β-xylosidase, α-arabinofuranosidase, and α-glucuronidase
should also be considered standard classes of hemicellulases to convert
hemicelluloses into monosaccharides [124,125]. Gao et al. [124]
demonstrated that it is vital to optimize the ratio of cellulases and
hemicellulases for achieving a high yield of glucose and xylose, while
decreasing the total loading of enzymes. Pretreated biomass waste of
corn stover hydrolyzed by the optimal mass ratio of xylanases and cel
lulases (25:75) can produce up to 20% xylose.
Both bacteria and fungi can generate cellulases and hemicellulases.
Acetovibrio, Bacteriodes, Bacillus, Clostridium, Cellulomonas, Erwinia,
Microbispora, Ruminococcus, Streptomyces, and Thermomonospora are
cellulases producing bacteria, while Aspergillus niger, Trichoderma,
Penicillium, Fusarium, Phanerochaete, Humicola, and Schizophillum sp. are
cellulases producing fungi. However, Trichoderma reesei, Trichoderma
longibrachiatum, and Trichoderma viride are a few major filamentous
fungi, famous for production of cellulases and hemicellulases. Clos
tridium thermocellum, Geobacillus thermocellum, Geobacillus stear
othermophilus, and Dictyoglomus turgidum are a few widely known
hemicellulases that produce bacteria [124].
On their turn, Trichoderma resei and Aspergillus niger produce an
almost complete range of enzymes needed to hydrolyze cellulose and
hemicellulose [70,110]. Despite numerous advantages of
enzymes-based hydrolysis, the high cost of enzyme production is the
main reason for not being used in bioethanol production. Dos Santos
et al. [122] estimated that the cost of enzymes alone represents almost
20% of the total production cost of bioethanol production. To overcome
this inconvenience, several factors are optimized for cost-effective bio
ethanol production, including pH, temperature, substrate concentration,
enzymes loading, mixing rate, and the addition of surfactants. All these
parameters can increase the yield of monosaccharides obtained in the
hydrolysis process [110].
Final products of hydrolysis of cellulose and hemicellulose are
fermentable sugars like hexoses and pentoses, which are the most suit
able substrates for bioethanol production by fermentation. The latter is
an anaerobic process, which can be metabolized by several microor
ganisms, as shown in equations (a) and (b) below [70,126].
C6H12O6 → 2 C2H5OH + 2 CO2 (a)
3 C5H10O5 → 5 C2H5OH + 5 CO2 (b)
4.3. Microorganisms for fermentation
In the meantime, the selection of microorganisms for fermentation
depends on the composition of fermentable sugars. Fungi Saccharomyces
cerevisiae and bacteria Zymomonas mobilis are the most commonly used
microorganisms in bioethanol production. Both microbes enabale high
ethanol yields, high ethanol tolerance, and ferment a wide range of
hexoses and disaccharides. Z. mobilis is considered superior as it pro
duces less biomass [70,127,128], but none of them is capable to ferment
pentoses [129]. Dos Santos et al. [122] reported that most of the
lignocellulosic biomass contains up to 25% pentoses, mainly xylose. An
edible strain of filamentous fungi Neurospora intermedia can assimilate
pentoses [130]. Saini and co-workers [131] mentioned several yeasts,
Fig. 4. Process flow diagram of bioethanol production from microalgal biomass. Developed by the authors based on reference [151].
Table 3
Advantages and limitations of biological and chemical hydrolysis.
Biological hydrolysis Chemical hydrolysis
Low utility cost High utility cost
45–50 ◦
C temperature Low to high temperatures
Conducted under mild conditions Conducted under strong/severe
conditions
Does not cause corrosion Causes corrosion of equipment
Has better yields than chemical
hydrolysis
Yields are lesser than for biological or
enzymatic hydrolysis
Costly manufacturing of enzymes drives
the overall cost of enzymatic
hydrolysis, and ultimately the overall
bioethanol production cost
Large amounts of acid are needed, the
formation of inhibitors, which affect
fermentation and environmental issues
are disadvantageous
Slow reaction Fast reaction
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8
including Candida shehatae, Scheffersomyces stipitis, and Pachysolan tan
nophilus, which ferment pentoses even in the presence of hexoses.
However, on the industrial scale S. stipitis is the most commonly used
yeast for high yield bioethanol production [129,132]. Hence, the se
lection of fermenting microbes is a key step to produce bioethanol at
high yield.
Goncalves et al. [133] reported a recombinant S. cerevisiae strain
with an integrative plasmid to over-express the genes for producing
xylose metabolic enzymes. The recombinant strain of S. cerevisiae suc
cessfully co-ferments xylose and glucose under anaerobic conditions.
Currently, genetically engineered S. cerevisiae strains, which facilitate
co-fermentation, are widely available for manufacturing bioethanol.
Recombinant bacteria Zymomonas mobilis assist pentose fermentation by
introducing a xylose-metabolizing pathway from E. coli [134].
Sharifyazd and Karimi [135] demonstrated that Filamentous fungi
Mucor indicus white-rot basidiomycete Trametes versicolor [136] and
brown rot fungus Neolentinus lepideus [137] are capable of fermenting
both hexoses and pentoses. Contributions by Sarkar et al. [110] esti
mated sequential fermentation with two different microbes, in different
periods. Unfortunately, their results were not relevant. Initially, the
fermenter was exposed to S. cerevisiae for fermenting hexoses, and later
the fermenter was fed with C. shehatae, which is a pentose assimilator.
Wood rot fungus Schizophyllum commune is capable to produce
lignocellulolytic, cellulolytic, and xylanolytic enzymes, which constitute
a relatively complete set of enzymes. They enable the complete tasks of
bioethanol production, i.e., cellulose production, hydrolysis, and
fermentation of hexoses and pentoses [138,139]. Kumar and Kumar
[140] highlighted the metabolic diversity of different microorganisms
that allow bioethanol production. Similarly, Barnard et al. [141]
analyzed the effects of technological processes and identified organic
products: bioethanol, biodiesel, biobutanol, and biogas. Stroparo et al.
[142] evaluated the use of fungi that produce hydrolytic enzymes in
agro-industrial wastes and trigger a higher production level.
4.4. Efficiency and environment
Gumisiriza et al. [143] proposed an approach, which combines
different biomass conversion technologies. In the meanwhile, they
evaluated the socio-environmental impacts of gaseous biofuel genera
tion [144]. Efficiency and productivity from microalgae (third-genera
tion production) are also part of this scenario. Appropriate management
of agro-industries is considered indispensable [145,146].
Rapid industrial development and technological demand caused the
depletion of fossil fuels, along with air pollution, climate change, and
ecological disturbance. All these have strengthened the committment of
scientists to develop sustainable, renewable, and eco-friendly alterna
tive energy sources. Many developed and developing nations are now
engaged in research to produce the necessary quantities of biofuels and
to develop efficient technologies, which would allow to depend less on
fossil fuels. In addition, such technologies would offer a better chance to
repair the damaged components of the ecosystem [147]. It is anticipated
that when a nation produces biofuels from agricultural wastes, this will
stimulate agricultural production. Biodiesel, bioethanol, and biogases
are the most common biofuels made from industrial and agricultural
biomass wastes [148].
4.5. Feedstock categories for bioethanol production
Feedstocks for bioethanol production are categorized into three
biomass types or generations. First-generation bioethanol or biofuels
produced from sucrose and starch-containing feedstocks (e.g., sugar
cane, sugar beet, sweet sorghum, corn, barley, potato/potato wastes, or
vegetable wastes), while second-generation bioethanol or biofuels are
being made from lignocellulosic biomass (e.g., straw, grass, and wood).
Regrettably, lignin is highly recalcitrant, therefore, it is not involved in
bioethanol production; its only use is to generate energy for the process
via combustion [149]. In Sections 4.1. and 4.2. we have discussed bio
ethanol production from sugar-based, starch-based, and
lignocellulose-based feedstocks. Third-generation bioethanol or biofuels
are produced from microalgae. Table 4 summarizes merits, drawbacks,
and challenges related to 1G, 2G, and 3G bioethanol generations.
Because of the absence of lignin, their short life cycle, relatively high
growth rate, cost-efficient cultivation, and their bioremediation capa
bility, microalgae are excellent feedstock for bioethanol production. In
addition, some of the microalgae can accumulate a greater amount of
lipids, as secondary metabolites, which can be used to produce biodiesel
[150]. Kim et al. [147] cultivated microalgae Chlamydomonas sp, which
were utilized to extract lipids for biodiesel production, while the resid
ual biomass was pretreated for bioethanol production. In this approach,
microalgae exhibit capability for bioremediation, hence, they offer
additional environmental benefits.
Hena et al. [151] suggested anaerobic digestion of biomass of
Chlorella vulgaris to produce bio-fuel, which was cultivated in waste
water during its treatment. Chlorella vulgaris was able to remove recal
citrant pharmaceuticals and metronidazole from wastewater.
Thornley and Gilbert [152] demonstrated the usefulness of
combining technological alternatives for efficient biofuel production, so
that environmental benefits would be maximized. Climate change is the
current topic of concern, which can be amended by replacing fossil fuels
with a renewable energy source, which would cause fewer environ
mental and social concerns.
Several advantages of using different plant species intended for
biofuel manufacturing were identified, including the use of sugarcane,
which enables high photosynthetic yields and productivity in biofuel
manufacturing, while also reducing greenhouse gas emissions relative to
fossils fuels.
5. Agro-industrial Techniques and Infrastructure Analysis
5.1. Biorefineries
According toy Berry [153], biological engineering allows researchers
to boost production using biofuel production innovation, rather than
adopting existing approaches. Similarly, Branco, Serafim, and Xavier
[70] emphasized that biorefineries could be integrated into existing
plants of pulp and paper industry via exploiting the high level of tech
nology, infrastructure, and logistics.
Black liquors represent the waste of paper and pulp industries. Black
liquor from Kraft process of wood contains hemicelluloses, some cellu
lose, and almost 90% of the lignin from wood, solidified by evaporation
and subsequent combustion to generate energy, while Kraft pulp is used
for paper production [154]. Unlike the Kraft process, the sulfite process
is not suitable for all kinds of wood. Sulfite process is preferred for
softwood like fir, hemlock, and spruce wood [155]. The sulfite process
removes hemicelluloses and lignin from wood. In addition, it hydrolyzes
the polysaccharides of hemicelluloses and cellulose contained in wood;
therefore, besides lignin, sulfite liquors contain hexoses and pentoses
[156].
Composition of sulfite liquors is different for different kinds of
woods. Hexoses dominate sulfite liquors originating from softwoods,
while hardwoods release more pentoses into the sulfite liquor [157].
Interestingly, sulfite liquors represent a waste for paper industries, but
they contain a load of fermentable sugars, such as arabinose, xylose,
mannose, galactose, and glucose, which are valuable feedstocks for
bioethanol production. Portugal-Nunes et al. [158] estimated that 90
billion L/year of sulfite liquor are produced, which are not appropriate
for direct disposal in the environment, given their high biological oxy
gen demand. This is why, using sulfite liquors from paper pulp industries
as the feedstock represents an excellent option for bioethanol production
[70].
Portugal-Nunes and co-workers [158] fermented sulfite liquor of
Eucalyptus globulus (hardwood), using immobilized cells of
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Scheffersomyces stipitis. Harner et al. [159] studied the tolerance of
genetically modified Pachysolen tannophilus for fermenting sulfite liquor
of hardwood. Henriques et al. [160] performed a two-stage aera
tion/fermentation process to improve bioethanol production from
hardwood sulfite liquor. Pereira et al. [157] reported 2.4 g/L bioethanol
made from sulfite liquor of Eucalyptus globulus, using Scheffersomyces
stipitis for fermentation. S. cerevisiae strains were employed to produce
bioethanol from softwood sulfite liquor [161,162]. Studied was the ef
ficiency of S. stipitis for fermenting softwood sulfite liquor [163]. Sulfite
liquor of mixed 80% softwood (spruce) and 20% hardwood (beech),
containing hexoses and pentoses, was fermented by S. cerevisiae,
securing a yield in the range of 0.31–0.44 g/g [164]. S. cerevisiae is a
yeast, which ferments hexoses; therefore, it is mainly used for softwood
sulfite, whereas Scheffersomyces stipitis is a pentose fermenting yeast,
found more suitable for hardwood sulfite liquors.
Pulp and paper mill sludge is a waste containing organic residues,
generated from paper and pulp industries after using a considerable
amount of water as the reaction medium and for washing. Cellulose fi
bers and lignin are the main components of the organic residues of pulp
and paper mill sludge; however, lignin is present only at very low con
centrations. Disposal of pulp and paper mill sludge is not an easy task,
since it needs to be thickened before landfilled, which is an energy-
consuming process. In addition, leaking hazardous substances into the
environment is a threat, because of its large volume. Several scientists
consider that bioethanol production from pulp and paper mill sludge is
the best option [165,166].
Pulp and paper mill sludges do not need multiple steps to generate
bioethanol, since they already contain cellulose and a negligible amount
of lignin. Hydrolysis and fermentation are the only steps prior to bio
ethanol extraction [167,168]. Simultaneous saccharification and
fermentation are more common to manudacturing bioethanol from the
same sludge [166,169]. In bioethanol production from the same raw
material Mendes et al. [166] used S. cerevisiae. Peng and Chen [170]
Table 4
Summary of merits, drawbacks, and challenges related to 1G, 2G, and 3G bio
ethanol generations.
Bioethanol
Generation
Merit Drawbacks Challenges
1G Expansion in
agriculture sector.
Increase the food
price
Bioethanol is
renew-able energy;
however, it is still
not even
comparable to
petro-leum due to
high production
cost and using food
feedstocks.
Enhancing the
socio-economic
conditions of local
farmers.
Overloads the irri-
gation system.
Promote agriculture
for food and energy.
Requires large size
of land.
Encourage
transforming bar-
ren land into fertile
ground.
Demands high
amounts of energy
during production.
Increase
employment.
Insignificant
contribution to GHG
fall.
Production of co-
products during
fermentation
process e.g.,
fertilizer, fodder
and feedstocks for
biogas plants.
Fibrous residue
(bagasse) can be
used for heat
production.
2G Utilizes non-food
ligno-cellulosic
material.
Does not produce at
fully
commercialized
scale.
Though the price of
the feedstocks of
2G is significantly
less than 1G,
however, feed-
stocks of 2G are
com-plex in nature
hence their
conversion into
bioethanol requires
many steps, which
is not cost effective.
One should develop
an efficient process
to produce cost-
effective
bioethanol from 2G
feedstocks
lignocellu-losic
biomass.
Produces less GHG
comparing to 1G
during production.
High production
cost.
No competition for
land with
agricultural field.
Alteration in
agriculture and
forestry sectors.
Does not affect the
food prices.
3G The growth yield of
feedstocks i.e.,
microalgae is higher
than the growth
yield of 1G and 2G
feedstocks.
Nutrients are
required in culture
media.
Harvesting micro-
algal biomass is the
grea-test challenge
of 3G bioethanol
production.
Harvesting micro-
algal biomass
represents 20–30%
of the total cost of
the bioethanol
production.
Produces 30 times
more energy per
acre than land
crops.
Harvesting microal-
gal biomass consti-
tutes an expensive
process.
Feedstocks can
grow in different
modes of culti-
vation, while the
feedstocks of 1G
and 2G can only
grow in autotrophic
mode.
Table 4 (continued)
Bioethanol
Generation
Merit Drawbacks Challenges
Its feedstocks have
shorter harvesting
life cycle as
compared to 1G and
2G.
Relative to 1G and
2G, the 3G
feedstocks biofix
atmos-pheric CO2 at
higher rate.
Water consumption
rate in producing
feedstocks is lower
for 3G than for 1G
and 2G.
Microalgae can
grow in different
kinds of water, in-
cluding saline,
brackish water, or
coastal seawater,
hence their
cultivation does not
affect agriculture
adver-sely.
Cultivation does not
require pesticides,
like for 1G and 2G.
Microalgal biomass
con-tains no lignin,
hence the process of
bioethanol
production is
simpler than for 2G.
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applied separate hydrolysis and fermentation steps of pulp and paper
mill sludge. As in most cases, fermentation was enabled by S. cerevisiae.
Biorefinery models rely on various biomass raw materials, such as
lignocellulose, algae, and numerous types of wastes, and integrate them
in the process of 1st/2nd generation technology [171]. In a simulta
neous process of flow of sugarcane juice and lignocellulosic fractions
biofuels are being produced [67]. Also, multiple bioenergy products can
be obtained via relevant conversion technologies [172]. Nevertheless,
lignocellulose-based processes for the production of value-added prod
ucts still represent the bottleneck of viability.
On the basis of their development, biorefineries belong to two cat
egories: (i) bottom-up bio-refineries and (ii) top-down biorefineries. The
bottom-up approach is characterized by expanding the facilities to
produce a vast range of products, which aim to maximize the utilization
of raw materials. In comparison, the top-down approach is characterized
by the use of all kinds of agricultural wastes to produce value-added
products. An example of bottom-up biorefinery uses corn and wheat
as feedstocks to produce starch. Eventually, its product portfolio can be
expanded exploiting available technologies e.g., for starch derivatives
(glucose syrup, hydrolysates, maltodextrin, cyclodextrin, and distarch
phosphate). Lenzinger Berichte in Austria, Borregaard in Norway, and
BioHub in France use the bottom-up approach. At the same time, Green
biorefinery and Utzenaich in Austria, DONG Energy, Biogasol, Estibio
and Haldor Topsoe in Denmark, Chemtex in Italy, Biowert and Leuna in
Germany, and Microbiogen in Australia are top-down biorefineries,
producing a renewable source of energy.
Modern bio-refineries operate on a multifunctional concept for a
complete utilization of agricultural waste biomass for bioethanol pro
duction. More importantly, the agro waste generating industries should
integrate bio-refineries to cut down the cost of waste disposal, and to use
the waste as a feedstock to produce eco-friendly renewable green en
ergy, such as bioethanol. This can be done utilizing Kraft liquor or sulfite
liquor and solid biofuel from waste lignin.
One can state that besides paper industries, agro-industry should
integrate bio-refineries to use their agricultural waste into value-added
bioethanol, while generating income and maintaining a sustainable
environment.
Other studies conclude that the key toward a convenient outcome
would be the biorefining of all major lignocellulose polymers: cellulose,
hemicellulose, and lignin [173]. These processes allow lignocellulosic
biomass to produce biochemicals and biofuels in a sugar platform.
Indeed, there are high costs of operation and expenses related to
installing equipment suitable to carry out the pretreatment processes, i.
e., the conditioning of hydrolyzate.
Development of biorefineries can also be considered. Callegari et al.
[174] descibed the use of wear mill for pretreatment. Simple biofuel
processes do not require washing, detoxification, and solvent recovery.
Likewise, Dragone et al. [175] report that technological innovations
bring biorefineries to a new status of “advanced biorefineries,” owing to
the use of biomass components other than carbohydrates, such as pro
teins, acetic acid, and lignin. Enzymes are produced in situ, and new
biorefinery designs are implemented, which integrate heat in the system
to intensify the production of cellulosic ethanol from lignocellulosic
biomass [176].
Hashemi, Mirmohamadsadeghi, and Karimi [177] evaluated the
development of biorefineries based on the processing of safflower,
composed of straw (79.6% by weight) and oilseeds (20.4% by weight).
These are important raw materials for the development of multiple
biofuels, including bioethanol, biogas, and biodiesel. Authors’ results
can be summed up as each kg of safflower plant producing 97.2 g of
ethanol, 22.4 L of methane, and 46.6 g of biodiesel equivalent to 0.168 L
of gasoline per kg of safflower.
5.2. Sustainable technology
Wasiak [65] believes that the right choice of agricultural plants,
tillage, and conversion technologies in one unit of arable land area,
dedicated to an energy plantation, may provide energy and manage
sustainable operations. Concerning the technical limitations, forest
farming systems, and high expectations, biomass conversion technolo
gies are called to reduce concurrent limits to the global food deficit and
minimize this type of competition [178].
Conditions of agro-industrial infrastructure require technological
improvements to achieve sustainable innovation in plants. This is where
investments, markets, and government incentives affect decision-
makers’ performance in producing biofuels [179]. One should consider
economic, environmental, and social factors; therefore, certification
efforts must be adopted to alleviate the problems in the agro-industrial
benchmarks against the country’s economic risks [178].
Bioethanol production via microbial fermentation [180] represents
an example of this approach. Specific responsibilities in producing
biofuels at this scale need to account for possible conflicts related to
technological investment, infrastructure, and ecological care [181] and
the effects of sustainability, biodiversity, and impacts of social, political,
economic, and environmental development. Additionally, the scenario
must consider that sustainability of biofuels depends mainly on the
ability to maintain the supply chain of the initial biomass. Studies by
Kargbo et al. [182] revealed that the second generation raw materials
are more sustainable than first generation materials. Second-generation
biofuels possess a greater potential to reduce greenhouse gas emissions
(50–100%) relative to first-generation biofuels (50–90%). Nevertheless,
when using second-generation raw materials, production costs are twice
greater than of fossil fuels.
Most studies focus on the conflict between biofuel production and
global food security [183], implying the priority of permanent assess
ments in agribusiness and in quality certification policies. It adds a third
space to the familiar combination of government and industry: the social
dimension, to account for the law and appropriation of arable land for
farmers in ecosystem control [184].
According to Mohr and Raman [185], sustainable nature refers to
second-generation bioethanol (2G), which has been questioned from a
social standpoint, given that farmers who cultivate the land believe that
biofuel production is harmful to the environment [186]. On the opposite
end, Guatemala has exported bioethanol mainly to European markets,
and the country’s production has been certified as sustainable. Negative
impacts of “agrofuels” are still debated, particularly in the case of
marginalized communities [187].
These social meanings also relate to the production of biofuels with
plant materials and agree with protecting the environment, reducing
greenhouse gas emissions, and getting in compliance with ethical con
siderations, primarily in not violating human rights [188–190]. Lo SLY
et al. [191] recognize that energy derived from biomass provides ben
efits, including lower emission of greenhouse gases and reduction of
wastes. For commercializing the entire energy generated from biomass,
adequate management of the supply chain is needed. The latter would
guarantee that the logistics processes do not affect the economic
viability of biomass use.
In terms of energy security improvements, Foust et al. [192]
demonstrated that the most significant impediment to energy security
was the lack of global food access. Energy policies all over the world
assume commitments to maintaining environmental balance [193],
often by adjusting ethical responsibilities and by imposing certifications.
All these secure the necessary technologies and infrastructure to address
biofuel production respectfully and cleanly [194], such that they also
adhere to existing policies. By this, the gap between the ethical and the
environmentally sustainable goals of agro-industrial or chemical com
panies producing biofuels [195] ponders care, control, and
decision-making against large-scale conversion of biomass. By these
measures production becomes sustainable and ethically responsible to
its stakeholders. In addition to considering life-cycle assessment (LCA)
of sustainable technologies all measures mentioned above represent
important decision-making tools, being used by many to examine and
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define the environmental impacts, energy consumption, and economic
feasibility of different methods or pathways in bioethanol production,
which meet the challenges of future research and innovation. LCA does
not indicate which method is beneficial, but it notifies the viable ad
justments among different bioethanol manufacturing methods or path
ways. For example, renewable feedstocks for bioethanol production save
the non-renewable energy resources and mitigate the GHG, however,
they cause acidification and eutrophication of soil and water, respec
tively [196]. Khoshnevisan et al. [197] compared steam explosion and
N-methylmorpholine-N-oxide the pretreatment processes from the life
cycle perspective using pinewood. It was revealed that N-methyl
morpholine-N-oxide is environmentally friendlier than steam explosion.
5.3. Global scenario of sustainable production of bioethanol
The future of bioethanol production contemplates a complex sce
nario determined by the growth of the world population, and high de
mand for energy, in addition to new requirements for policies for the
certification of biofuels such as Bioethanol [198]. Meanwhile, techno
logical advances and industrial methods to improve production perfor
mance are part of this global trend, and this can promote energy
production’s economical and efficient development based on sustain
able mechanisms [199].
Falck-Zepeda et al. [200] (2011) focused on the potential for biofuel
production in the Latin American and Caribbean regions. A combination
of a global model for the agricultural sector representing energy demand
and trade in biofuel products is proposed. While the data provided by
the OECD/FAO [201], estimate that world production of Bioethanol
should go from 120 Mml (billion litres) in 2016 to 137 Mml in 2026,
data shown in Table 5. By 2026, it is estimated that world production of
bioethanol 1G will distribute 55% of the production based on corn and
35% to sugar crops. Within this distribution, biofuels based on bagasse
(residue) from 2G production processes will have a low share due to the
lack of investment in research and development (R&D).
This situation could be maximized by generating greater interest and
more significant investment from biomass-based industry players and
new research that includes a comprehensive techno-economic analysis
[202]. It is essential to point out that 60% of this increase is expected to
originate in Brazil, mainly for consumption. The United States, China
and Thailand are also expected to exceed 14%, 11% and 8%, respec
tively, on the 17 Mml (billion litres) of expected growth. According to
OECD/FAO [201], the increase of 17 Mml will be achieved at the
expense of producing raw materials such as corn, sugar cane and other
crops.
In this sustainable and global scenario, some situations put pressure
on alternative energies that have a lower impact on the seven environ
mental footprints. The environmental impact caused in the European
Union by the transport sector, which represented 24% of greenhouse gas
(GHG) emissions, 33% of final energy consumption in the European
Union in 2015 [203], which promoted low-emission mobility strategies
with the replacement of fossil fuels, accelerating the deployment of
low-emission alternative fuels [204].
In this context, the sustained advances of the countries with the most
significant potential for the production of Bioethanol, such as China
(mainland), the US, India and Brazil, are projected towards the use of
waste for the production of 2G bioethanol. In terms of alternative en
ergy, the results by Holmatov et al. [205] express that the global net
production of lignocellulosic Bioethanol ranges between 7.1 and 34.0 EJ
(Exajoule) per year, replacing between 7% and 31% of petroleum
products used in the transport sector, which generating relative emis
sions savings of 338 megatonnes (Mt; 70%) to 1836 Mt (79%) of the
world. On the other hand, it was determined that the environmental
carbon footprint produced varies according to the crops. The study by
Holmatov et al. [205] estimated that the land, water and carbon foot
prints of net Bioethanol vary between potentials, countries/territories
and feedstocks by 28–44 g CO2 equivalent MJ− 1 (MJ = Megajoule).
Other results presented by Brandão et al. [206] established that
Bioethanol trajectories show lower climate change impact overall
compared to the fossil fuel benchmark, but higher than the minimum
greenhouse gas (GHG) emission savings of 33gCO2-eq/MJ (>65%),
established in the RED (European Commission’s Renewable Energy
Directive). In short, the sustainable production of Bioethanol can be
considered a viable alternative to reduce the impact on environmental
footprints. However, more studies will still be needed to approach a
more detailed result.
6. Future of bioethanol
The general future of biofuel production envisages a complex sce
nario determined by the global population growth and new re
quirements for certification policies in different countries [207].
Technological advances and industrial methods capable of improving
production performance are part of this overall trend. Another impor
tant factor in business management is to execute comprehensive de
cisions for the sustainable production of biofuels and to adopt
technological advances and novel infrastructures in agribusiness to
produce biofuels [208,209].
Leadership of public and private companies should make well
informed and firm decisions to develop different future applications of
biofuels. Current trends are focused on evolving and implementing
transport systems based on biofuels originating from biomass [210]. In
addition, one needs to improve the techniques, procedures, and con
version of biomass by biochemical and thermochemical methods [211].
Expected for the future is a cleaner energy from biofuels, such as 2G and
3G ethanol [212].
The agribusiness sector is committed to engage in the adaptation of
its industrial infrastructure. This is enabled by technological innovation
and the adoption of complex bio-production methods, such as enzyme
dosage, which will increase sustained production of bioethanol in sec
ond and third-generation plants. As a result, one can promote efficient
economic development of energy production via sustainable mecha
nisms [213] and production from agricultural wastes [214,215].
One should also implement business management practices that
involve efficient employee participation in technological processes
[216,217]. Fig. 5 summarizes two general processes of bioethanol pro
duction in 1G and 2G pathways, which have served as the basis for
future ambitious new methods.
Table 5
Bioethanol world projection.
Bioethanol Production 2016* 2017 2018 2019 2020 2021 2022 2023 2024 2025 2026
Average world production** 120 123.7 126.8 128.4 130.7 131.5 132.8 133.7 134.7 135.8 136.7
from the corn 68.2 71.9 73.7 73.9 74.7 74.5 74.5 74.3 74.2 74.0 73.7
from the sugar cane 27.9 29.5 30.2 31.1 31.9 32.5 33.3 33.7 34.3 34.9 35.5
overall consumption 117.2 124.6 127.0 128.8 130.8 131.8 133.0 134.0 134.9 136.0 136.9
consumption for fuel 96.2 103.1 105.4 107.0 108.9 109.6 110.7 111.4 112.1 113.0 113.6
Average production for 2016*; ** Average world production in Mml (billion litres). Data adapted from [201].
J.R. Melendez et al.
12. Renewable and Sustainable Energy Reviews 160 (2022) 112260
12
7. Conclusions and future perspective
Promotion of biofuels, specifically of bioethanol in the industrial or
agro-industrial sector, attracts increasing interest in process quality
certifications, which secure the effectiveness of alternative energy
sources. Nevertheless, the main gaps of biofuel production go beyond
the impact of not using proper technology or not relying on its devel
opment. The industrial sector must promote and adopt the innovation of
methods, equipment, and infrastructure to reach new levels of bio
ethanol production efficiently and sustainably, harmonized with society
and the environment. Additionally, the technical advances and agri
business infrastructures available to produce biofuels need a continuous
review of sociopolitical and economic factors to promote second-
generation bioethanol. By considering these requirements, the main
results of our review pertain the agro-industrial valorization of bio
ethanol production processes, such that one minimizes the integral im
pacts throughout the production cycle. This is accomplished by
promoting sustainable designs, which consider economic, environ
mental, and social factors together with biofuel production policies.
They also stimulate the feasibility of integrating biofuels into the current
liquid fuel infrastructure and third-generation materials derived from
algae biomass.
To promote the optimal development of biofuel production, exten
sive research and government investments are needed. Governments
must provide appropriate infrastructure, including port facilities and
roads, to connect feedstock producers and biorefineries. Bioethanol
production needs to be stimulated by tax cuts and subsidies offered to
biofuel producers. Additionally, biofuels crops may compete with food
or arable land sectors, making them unsustainable unless there is a well-
established agricultural sector, which guarantees food self-sufficiency.
Funding sources
This research did not receive any specific grant from funding
agencies in the public, commercial, or not-for-profit sectors.
CRediT authorship contribution statement
Jesus R. Melendez: Conceptualization, Methodology, Investigation,
Writing – original draft, Supervision. Bence Mátyás: Investigation,
Validation, Writing – review & editing. Sufia Hena: Conceptualization,
Investigation, Validation, Writing – original draft. Daniel A. Lowy:
Investigation, Validation, Writing – review & editing. Ahmed El Salous:
Writing – review & editing.
Declaration of competing interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Acknowledgements
The authors acknowledge the support received by the Universities
from their policies to promote international research networks. Special
thanks to the Catholic University of Santiago of Guayaquil and the
Research Group of Applied Plant Glycobiology, Dama Research Center
limited.
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J.R. Melendez et al.
![Renewable and Sustainable Energy Reviews 160 (2022) 112260
Available online 19 February 2022
1364-0321/© 2022 Published by Elsevier Ltd.
Perspectives in the production of bioethanol: A review of sustainable
methods, technologies, and bioprocesses
Jesus R. Melendez a,b,c,d,*
, Bence Mátyás e
, Sufia Hena f
, Daniel A. Lowy e,g
, Ahmed El Salous h
a
Faculty of Technical Education for Development, Catholic University of Santiago of Guayaquil, Guayaquil, 090615, Ecuador
b
Postdoctoral Program, Universidad Yacambú, Cabudare, 3023, Venezuela
c
Antonio José de Sucre National Polytechnic Experimental University, Barquisimeto, 3001, Venezuela
d
Universidad Nacional Experimental de los Llanos Occidentales Ezequiel Zamora, San Carlos, 2201, Venezuela
e
Genesis Sustainable Future, Ltd., 33 Rákóczi St., B-A-Z, Sárospatak, H-3950, Hungary
f
Department of Chemical Engineering, Curtin University, Bentley WA, 6102, Australia
g
Department of Mathematics, Sciences, Technologies and Business, Northern Virginia Community College, 5000 Dawes Avenue, Alexandria, VA, 22311, USA
h
Dirección de investigación, Universidad Agraria del Ecuador, Guayaquil, 090104, Ecuador
A R T I C L E I N F O
Keywords:
Bioethanol
Biomass
Biorefineries
Chemical processes
Industrial wastes
And sustainability
A B S T R A C T
The main objective of this review paper is analyzes perspectives of manufacturing bioethanol via biotechnology,
using sustainable management. Two categories are emphasized: (i) agro-industrial valuation, which represents
alternative methods for bioethanol production, and (ii) Agro-industrial Techniques and Infrastructure Analysis.
Future of biofuels is addressed in terms of applicability and production methods. Authors perform a systematic
literature review to characterize the present state of knowledge on ethanol production. The study is conducted
from a decision-making standpoint, so that they identify several problems (subcategories) in bioethanol pro
duction. The section on technology and industrial infrastructure focuses on advances in biorefineries, revealing
technical and economic limitations, and also the gap between sustainable production policies and environmental
impact. The second part of the paper presents the evolution of the industrial sector toward technological
innovation and adoption of complex bio-production methods for clean energy production. This review mostly
covers results reported over the past decade, with a special emphasis on papers published recently, from 2010 to
2021, therefore, it can serve for extended future research that would address additional specific categories. One
concludes that the industrial sector must promote bioethanol production efficiently and sustainably, harmonized
with the society and the environment. Sustainable production points to alternative energy sources based on
cellulosic materials and third generation materials derived from algal biomass.
1. Introduction
Development of bioethanol production is an essential strategy to
assure energy security. Therefore, adoption of bioethanol
manufacturing is being considered in many countries, though food se
curity policies limit its implementation [1]. Bioethanol obtained from
alternative biomass routes has offered important benefits by reducing
greenhouse gas emissions. In the meanwhile, it triggered the develop
ment of technologically suitable, sustainable industries for the produc
tion of bioethanol from sugar or biomass [2,3].
In present, the global agro-producer scenario and the experience
earned in agricultural production triggered innovation in agricultural
production [4–6], including education toward increasing agribusiness
efficiency in rural areas [7].
Similarly, global production of renewable energy stimulated coun
tries to close agreements and strategic alliances, which added value to
productive alternatives and imposed international responsibility [8–10].
In a broad context, the introduction and development of eco-friendly
and cost-effective technologies is in great demand and can be
addressed via circular economy approach [11]. When utilized in bio
refineries, such processes may encompass the production of bioethanol
from first-to third generation biofuels. International agreements and
alliances use sustainable development strategies to contribute to the
replacement of fossil fuels.
Consequently, new industrial developments must adhere to alter
native energy sources that will limit the use of oil, gas, coal, and wood
[12]. This scenario resulted in government policies, which promote
* Corresponding author. Faculty of Technical Education for Development, Catholic University of Santiago of Guayaquil, Guayaquil, 090615, Ecuador.
E-mail address: jesus.melendez@cu.ucsg.edu.ec (J.R. Melendez).
Contents lists available at ScienceDirect
Renewable and Sustainable Energy Reviews
journal homepage: www.elsevier.com/locate/rser
https://doi.org/10.1016/j.rser.2022.112260
Received 8 November 2021; Received in revised form 26 January 2022; Accepted 9 February 2022](https://image.slidesharecdn.com/1-s2-230409232833-6d4bccc7/85/1-s2-0-S1364032122001812-main-pdf-1-320.jpg)
![Renewable and Sustainable Energy Reviews 160 (2022) 112260
2
renewable energies [13] and support agro-industry within various en
ergy, economic, and geographical settings [14], so that more corporate
benefits should be obtained. Recycling various materials and using
renewable energy would contribute to the success [15]. Several inter
national organizations are committed to developing networks that guide
renewable energy policies, such as the Organization for Economic
Cooperation and Development (OECD), the European Union (EU), the
Slovakia Market Overview [16], and the International Energy Agency
(IEA) [17]. Bioenergy represents a replacement for fossil fuels in the
short- and medium-term, and contributes to mitigating greenhouse gas
emissions [18].
Production of biofuels has increased worldwide in emerging econo
mies, featuring productive approaches, which take advantage of mi
croorganisms [19], enzymes, incubation conditions, and engineering
schemes for efficient depolymerization [20]. Other results in biomass
experiments demonstrate that sodium carbonate pretreatment consid
erably improves ethanol concentration, with high levels of solids and
low enzyme loads [21]. Da Costa, Pereira, and Gomes [22] pointed out
that biorefinery based on residual and agro-industrial biomass generates
a variety of molecules. Consequently, in addition to their positive effects
as an alternative and ecological energy source [23], biorefineries can
enable industrial facilities to reduce their impact on public health and
the environment [24].
Our review paper addresses the study of bioethanol, and we will refer
to bioethanol as a biofuel. The terms ethanol and bioethanol are often
used alternatively. While ethanol is manufactured from petroleum by
hydration of ethylene, bioethanol is produced from existing or renew
able biomass. Ethanol is also known as petroleum-derived ethanol, and
signicant amounts of ethanol made in the world are derived from pe
troleum, while bioethanol represents renewable ethanol. Chemical
composition of petroleum-derived ethanol and bioethanol is the same.
Nevertheless, they are isotopically different. Bioethanol contains 14
C, in
the same ration as in its feedstocks, while there is no 14
C content
petroleum-derived ethanol given that all 14
C decays over time.
In this context, the main objective of this review paper is analyzes
perspectives of manufacturing bioethanol via biotechnology, using
sustainable management. Two categories are emphasized: (i) agro-
industrial valuation, which represents alternative methods for bio
ethanol production, and (ii) Agro-industrial Techniques and Infrastruc
ture Analysis. Our review paper examines in an updated manner one
crucial research question: the state of knowledge on bioethanol. We
report alternative methods of bioethanol production and address the
technology and industrial infrastructure used within a sustainable
scenario. Discussed are the gaps in each examined group. To answer
these questions, scientific literature was screened and evaluated sys
tematically and organized into two categories, as described below
(section 2).
The study is divided into seven sections. Section 1, The complete
introduction that relates the different indications of the growing need
for alternative energy, alternative methods of biofuel production and the
entire scenario of sustainable industrial production, section 2, presents a
general literature review, highlighting two categories main. Section 3
includes the methodological reference framework used to develop the
systemic review. Section 4 considers the alternatives for the production
of bioethanol from its advances by generation. Section 5 highlights
alternative industrial methods, such as biorefineries, the importance of
sustainable production processes and the importance of bioethanol
production from an energy perspective for the leading countries in
production, while section 6 directs us to recognize the future of bio
ethanol production. It ends with the main conclusions of the review
study in section 7.
2. General literature review
2.1. Agro-industrial valuation: Alternative methods for bioethanol
production
Agribusiness generates about 330 million metric tons of biomass
waste per year [25]. Therefore, the corporate world values renewable
energy sources to produce environmentally sustainable biofuels [26]. In
this framework, chemical processes carried out in the agro-industrial
production sector include clean technologies, such as bioprocessing,
pyrolysis, gasification, and Fischer-Tropsch synthesis, the latter being
used in a variety of processes for converting biomass to liquid [27]. Also,
chemical catalysis in Guerbet reaction yields molecules of great
complexity [28].
Aro [29] highlighted similar implications, when using selected plant
materials and evaluating the presence of microorganisms [30–32].
Agro-industrial biodiesel processing utilizes as raw materials oils plants
and microalgal biomass [33,34]. On their turn, oils are obtained by
distillation, pyrolysis, gasification, or catalytic processes. Hence,
agro-industrial valorization of the waste biomass represents a priority in
biofuel production at this scale [35].
A complementary study by Mendes et al. [36] identfied the role of
hydrolysis and esterification reactions in treatments with filamentous
fungi. For this, appropriate fungi were previously isolated from leaves,
and decomposed in aqueous environment. Lipases offer additional value
via their transesterification reactions. Assessed were the ethanolysis of
native oils in Brazilian Amazonian plants, such as andiroba (Carapa
guianensis), babassu (Orbignya sp.), Jatropha (Jatropha curcas), and palm
(Elaeis sp.) [37].
Escobar et al. [35] managed to isolate 1016 lipolytic microorganisms
that were analyzed by colorimetric assay. For further characterization,
the authors selected 30 initial lipolytic strains. Next, they performed
phylogenetic analysis and quantified the production of biofuels by a gas
chromatography. Agro-industrial wastes have proven rich in sugars,
minerals, and proteins [38–40]. Owing to the potential of such wastes to
promote efficient growth of microorganisms, they should be considered
as alternative raw materials with high added value for reuse [41].
2.2. Agro-industrial Techniques and Infrastructure Analysis
Technical capabilities and infrastructure in the agro-industrial pro
duction of biofuels, along with the relevant technologies and regula
tions, represent a continuous challenge [42], as one should also comply
with energy security and environmental sustainability [43]. To accom
plish these requirements, various approaches are adopted, which
consider both market dynamics and economic risks [44]. Benefits ob
tained in biofuel production depend on the raw material, the conversion
List of abbreviations
C. shehatae Candida shehatae
E. coli Escherichia coli
HRAP High raceway algal pond
1G First generation
2G Second generation
3G Third generation
LHC Lignin-hemicellulose-cellulose
CSLF Cellulose solvent-based lignocellulose fractionation
CbU/g Cellobiase unit per g
FPU/g Filter-paper units per g
EFB Empty fruit bunch
IEA Agency Energy International
SSF Simultaneous saccharification and fermentation
SHF Separate hydrolysis and fermentation
S. cerevisiae Saccharomyces cerevisiae
OECD Economic Cooperation and Development
EU European Union
J.R. Melendez et al.](data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7)