Recent advances in bio-based multi-products of agricultural Jerusalem artichoke resources

15 Pages·2017·2.98 MB·English

by  Yibin Qiu

Qiu et al. Biotechnol Biofuels (2018) 11:151 Biotechnology for Biofuels
https://doi.org/10.1186/s13068-018-1152-6
REVIEW Open Access
Recent advances in bio-based
multi-products of agricultural Jerusalem
artichoke resources
Yibin Qiu1,3†, Peng Lei2†, Yatao Zhang1, Yuanyuan Sha1,3, Yijing Zhan1,3, Zongqi Xu3, Sha Li1,3, Hong Xu1,3*
and Pingkai Ouyang1,3
Abstract
The Jerusalem artichoke is a perennial plant that belongs to the sunflower family. As a non-grain crop, Jerusalem
artichoke possesses a number of desirable characteristics that make it a valuable feedstock for biorefinery, such as
inulin content, rapid growth, strong adaptability, and high yields. This review provides a comprehensive introduction
to renewable Jerusalem artichoke-based biomass resources and recent advances in bio-based product conversion.
Furthermore, we discuss the latest in the development of inulinase-producing microorganisms and enhanced inulin
hydrolysis capacity of microbes by genetic engineering, which lead to a more cost-effective Jerusalem artichoke
biorefinery. The review is aimed at promoting Jerusalem artichoke industry and new prospects for higher value-
added production.
Keywords: Jerusalem artichoke, Inulin, Inulinase, Non-grain fermentation, Biorefinery
Background In addition to its unique value in ecological environ-
Concerns over fossil fuels depletion and environmental ment improvement, inulin-rich Jerusalem artichoke
protection have attracted increasing worldwide inter- serves as an important raw material in the food, chemi-
est in the development and utilization of sustainable and cal, and pharmaceutical industries (Fig.  1) [5, 6]. A
taintless energy resources. A sunflower species native to promising candidate, Jerusalem artichoke exhibits many
North America, Helianthus tuberosus L., also known as advantages for biorefinery. Compared with traditional
the Jerusalem artichoke, is a potential source of renew- grain crops, the Jerusalem artichoke produces large
able energy [1]. Meanwhile, the increasing use of chemi- amounts of biomass and can be harvested three times
cal fertilizers and development of irrigation agriculture a year [7]. Furthermore, the inulin found in Jerusalem
have led to the constant expansion of secondary saliniza- artichoke tubers can be easily processed using available
tion land area [2]. Because of more tolerance to harsher technologies, and can be suitable as a substrate for biore-
conditions than that of most commercial crops, Jerusa- finery. More importantly, the non-food utilization of this
lem artichoke was selected and developed for large-scale biofuel crop does not compete for arable land with grain
cultivation in saline–alkaline soils or coastal shoals [3, 4]. crops cultivated for food production [8]. This review aims
Therefore, the establishment of this cover crop in non- to provide a comprehensive overview on the characteris-
cultivatable land can reap both ecological and economic tics of Jerusalem artichoke and the pretreatment process
benefits. for biorefinery, as well as a description of inulinase-pro-
ducing microorganisms and inulinase catalytic mecha-
nisms that will provide fundamental knowledge for inulin
*Correspondence: [email protected] biorefinery studies. Furthermore, recent advances in
†Yibin Qiu and Peng Lei contributed equally to this work
Jerusalem artichoke research and development of bio-
1 College of Food Science and Light Industry, Nanjing Tech University,
Nanjing 211816, China products derived from these artichokes are summarized.
Full list of author information is available at the end of the article
© The Author(s) 2018. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License
(http://creat iveco mmons .org/licen ses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium,
provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license,
and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creat iveco mmons .org/
publi cdoma in/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
Qiu et al. Biotechnol Biofuels (2018) 11:151 Page 2 of 15
Fig. 1 Schematic diagram of the biorefinery of Jerusalem artichoke biomass
Finally, we discuss the prospects of energy crops in future Jerusalem artichoke components
biological manufacturing. Jerusalem artichoke is a semi-wild plant resource with
a great potential for development. Jerusalem artichoke
Overview of the Jerusalem artichoke resource tubers, which are mainly used during processing com-
Jerusalem artichoke features prise 75.88% water, 12.46% total sugar, 1.58% total protein
(w/w), 1.77% total fat, and other microelements as well
Jerusalem artichoke (Helianthus tuberosus) is a perennial
as vitamins [17]. Among them, inulin accounted for 80%
herbaceous plant that belongs to the Compositae family
of the carbohydrates found in tubers (Table 1) [14, 16].
[9, 10]. Due to its strong adaptability, Jerusalem artichoke
Generally, differences among cultivars, harvest periods,
is widely distributed in China as a cultivated crop [11].
production conditions, postharvest storage, and process-
Morphologically, the Jerusalem artichoke is large, gan-
ing methods result in variations. As a main component of
gly, and highly branched with yellow flower heads. The
tubers, inulin is a storage polysaccharide found in many
young stems are stout and can grow 100–300 cm tall. It
plants. The inulin mostly consists of d-fructose bonded
has opposite leaves on the lower part of the stem that are
roughly 10–20 cm long and 5–10 cm in width, and are
believed to be important for tuber yields [12]. The Jerusa-
lem artichoke has a hairy and fibrous root system which Table 1 Chemical composition of  Jerusalem artichoke.
can grow as long as 127 cm, showing strong environ- Data from Kaldy et al. [17]
mental tolerance and effective acquisition of soil nutri-
Ingredient Content
ents [13]. The tubers are irregularly spherical or spindle (% of fresh
shaped and vary in color from pale brown to white, weight)
red, or purple [14]. As an easily grown crop, it endures
Water 75.88
a wider range of temperatures and can tolerate pH lev-
Dry mass 24.12
els ranging from 4.5 to 8.2 [13]. In addition, cultivation
Total sugar 12.46
of Jerusalem artichoke does not need too much manage-
Total protein 1.58
ment and protection due to its strong resistance to pests
Total fat 1.77
and plant diseases [15]. It can be harvested annually after
P, K, Ca, Mg, Fe 0.50
planting, leading to high yields of fresh weight (90 t) per
Vitamin A (I.U. in fresh weight) 37
hectare that can be used as a sustainable biomass feed-
Vitamin C (mg/100 g fresh weight) 0.82
stock for biorefinery [16].
Qiu et al. Biotechnol Biofuels (2018) 11:151 Page 3 of 15
by (2 1) β-linkages that are terminated by d-glucose tubers pretreatment and showed that acid and inulinase
→
molecules bonded to fructose by (2 1) α-bonds. The hydrolysis produced 178.8 and 68.8 g reducing sugars per
→
degree of polymerization (DP) of standard inulin ranges kg wet matter of tuber for 1 h, respectively [26]. Com-
from 2 to 60 and the average molecular weight is approxi- pared with acid hydrolysis, enzymatic hydrolysis has
mately 5500 Da [18]. Besides the presence of abundant mild reaction, less by-products, no pigmentation, and is
fermentable polysaccharides, the Jerusalem artichoke is easier to separate and refine. Furthermore, the unique
rich in amino acids, B vitamins, and a variety of minerals advantage of enzymatic method is that it can simultane-
that are advantageous for multiproduct biorefineries. ously combine hydrolysis and fermentation in Jerusalem
artichoke biorefinery, which shorten the overall time and
Jerusalem artichoke pretreatment improve the productivity [27]. In addition, some micro-
An efficient inulin extraction process is important to organisms can directly produce inulinases to hydrolyze
ensure the successful utilization of Jerusalem artichoke inulin and avoid the addition of enzymes during refining,
in many fields. Fresh tubers are washed, dried, and which greatly reduces the cost [28, 29]. With the popular-
ground into a crude powder using a crushing machine ity of green production technology, the use of enzymatic
over a 40-mesh screen. Due to the high solubility of inu- degradation of inulin has become a major trend in the
lin in hot water, extracting inulin from the powder only industry. After hydrolysis, the composition of Jerusalem
requires a straightforward hot water extraction tech- artichoke hydrolysate was determined using the method
nique. Wei et al. optimized the conditions by using a previously described by Kaldy et al. [17].
neutral pH for 20 min at 76.65 °C and solvent:solid ratios
Microbial inulinases for biorefinery
of 10.56:1 (v/w); an inulin extraction yield of 83.6% was
obtained [19]. Yi et al. extracted inulin from fresh, frozen, During the process of Jerusalem artichoke biorefining,
or dry tubers through hot water extraction, and showed degradation of inulin into fermentable sugars for micro-
that the highest yield of 93% was obtained from Jerusa- bial utilization is the key step. Inulin is mainly broken
lem artichoke powder [20]. When Jerusalem artichoke is down through acid hydrolysis and enzymatic hydroly-
used in industrial biorefinery, the inulin extract is firstly sis into fermentable sugars for biorefinery. Compared
degraded into fermentable sugars for microbial utiliza- with acid hydrolysis of inulin, inulinase hydrolysis has
tion through acid or enzyme hydrolysis. Acid hydrolysis the advantages of less by-products, no pigmentation,
is an early method for inulin hydrolysis. The acid hydrol- and is easier to separate and refine, which is more suit-
ysis of inulin has been investigated utilizing mainly sul- able for fermentation production. As a type of furan
furic or hydrochloric acid. Nasab et al. investigated the fructose hydrolase, inulinase can act on the β-2,1 glyco-
effects of pH, temperature, and time on acid hydrolysis sidic bond, converting inulin into glucose, fructose, or
of inulin and the maximum amount of inulin hydrolysis inulooligosaccharides [30]. The main commercial source
(> 90%) was obtained at the pH < 2, temperature > 90 °C of inulinase is microorganisms due to the ease of large-
and the time of 1 h [21]. Xu et al. obtained 92.3% of the scale cultivation and their high inulin hydrolysis activity.
total sugars (20.2 0.1 g/L of glucose and 87.5 0.3 g/L At present, inulinase-producing microorganisms can be
± ±
of fructose) from 120 g/L inulin hydrolyze with 5% (w/w) screened using a plate assay as previously described by
HCl at 60 °C for 1 h [22]. Therefore, acid hydrolysis of Castro et al. [31]. Table 2 lists the high inulinase activ-
inulin has lower cost and proceeds faster. However, acid ity of different inulinase-producing microorganisms. To
hydrolysis results in colored by-products and inhibitors isolate inulinase overproducers, a number of inulinase-
that interfere with microbial growth [23]. And nowadays, producing strains have been reported. Kango et al. iso-
more and more attentions are paid to the development lated a new strain, Aspergillus niger NK-126, and used
of green production technology, and acid hydrolysis is dandelion tap root extract as the medium substrate for
not fit with this idea. So, researchers begin to focus on inulinase production; inulinase activity reached a final
the enzyme hydrolysis. Inulinase is the most commonly concentration of 55.0 U/mL after 96 h of fermentation
used for inulin hydrolysis. Sarchami et  al. optimized [32]. Ge et al. obtained the mutant strain A. niger SL-09
the enzymatic hydrolysis of inulin from Jerusalem arti- by using UV light and LiCl, whose inulinase 160 activ-
choke tubers and the 94.5% of the inulin conversion was ity increased nearly twofold (150 U/mL). For the A. niger
achieved at 48 °C for 48 h with the inulinase loading of SL-09 strain, the culture medium was further optimized
10 units g/inulin [24]. Singh et al. used immobilized yeast by using sucrose ester that can act as an effective inducer
inulinase for the hydrolysis of inulin in a batch system of inulinase synthesis; the highest inulinase activity
and maximum hydrolysis of inulin 84.5% was observed reached 230 U/mL after 96 h of shake flask fermentation
at 125 rpm after 4 h [25]. Szambelan et al. compared the at 30 °C with 140 rpm [33]. Moreover, yeast strains were
acid and enzymatic hydrolysis in Jerusalem artichoke also proven as another predominant inulinase producer.
Qiu et al. Biotechnol Biofuels (2018) 11:151 Page 4 of 15
s
e
c
n
Refere [49] [32, 44] [33] [61] [62] [63, 64] [65] [54] [34, 47] [66, 67] [68] [69, 70] [38, 71] [72] [73] [74, 75] [39] [76]
Effects of metal ions 2Activated by K and Cu; ++2inhibited 5 mM Hg +2and Fe+2; inhib-Activated by Cu+2ited by Fe+ 2Inhibited by Ag Fe and ++3 Al+ Inhibited by Ag and Hg++ 2 and Activated by Mn+22 Ca; inhibited by Hg ++2and Ag+22, Mg Activated by Mn++2and Ca; inhibited by +222 Zn, Cu and Fe+++ 2Activated by Ca, K, ++ Na, Fe2 and Cu2 ; +++22inhibited by Mg, Hg, ++and Ag+ 22Activated by Zn, Fe, ++2and Mg; inhibited by +22 Co, Cu++
g
m
n/
mi g
Vm 43.47 10/min× 402.1 53.1 µmol/± 15.72 mg/(mL s) 0.008502 mg/min 21.69 μmol/min/m
Kinetic characteristics Km 1.25 mM 5.3 1.1 mM± Exo-I: 43.1 mg/mLExo-II: 31.5 mg/mLExo-III: 25.3 mg/mLEndo-I: 14.8 mg/mL Endo-II: 25.6 mg/mL 0.20 mM at 40 °C and pH 5.0 10.63 mg/mL 20.0602 mg/mL 1.72 mM
ms and inulinase activity Enzyme property Mr (kDa)Optima T (°C)pH 62606.0 65555.0 454.0 and 4.8 604.5 Exo-I: 7045Exo-I: 4.5Exo-II: 40Exo-II: 4.5Exo-III: 46Exo-III: 4.5Endo-I: 34Endo-I: 5.0Endo-II: 31Exo-II: 5.0 68505.2 505.5 505.5 59 60.0505 57.6606 605.5 56256.0 75507.5 704.5
s y
g microorgani Enzyme activit 52.3 U/mL 230 U/mL 11 U/mL 53.1 U/mL 205.6302 U/gds 25.01 U/mL 9.9 U/mL 55.4 U/mL 409.8 U/gds 121 U/mL 85.0 1.1 U/mL± 127.7 U/mL 0.552 U/mL 0.45 U/mL 10.8 U/mL 135.2 U/mL 28.67 U/mL
ent inulinase-producin Strains Aspergillus fumigatus Aspergillus niger NK-126 Aspergillus niger SL-09 Aspergillus niveus 4128URM Aspergillus ficuum SK004 Aspergillus ficuum JNSP5-06 Aspergillus tritici Penicillium sp. TN-88 Kluyveromyces marxianus YS-1 Kluyveromyces sp. S120 K. marxianus ATCC 16045 Cryptococcus aureus G7a P. guilliermondii M-30 Streptomyces sp. GNDU-1 Paenibacillus polymyxa ZJ-9 Arthrobacter sp. S37 Bacillus smithii T7 Bacillus safensis AS-08
r
Diffe Type Exo- Exo-Endo- Exo- Exo- Exo- Exo-Endo- Endo- Endo- Exo- Exo- Exo- Exo- Exo- Exo- Exo- Endo- Endo- Endo-
2
Table Source Molds Yeasts Bacteria
Qiu et al. Biotechnol Biofuels (2018) 11:151 Page 5 of 15
Kluyveromyces marxianus is the most common inulinase Bacillus smithii T7 were optimized, and inulinase activity
producer that has been studied extensively. A recently increased to 135.2 U/mL after incubating for 72 h [39].
isolated strain, K. marxianus YS-1, was researched for This is the highest reported inulinase activity produced
its inulinase production [34]. After condition optimi- by bacteria thus far.
zation, maximum inulinase activity reached 55.4  U/ Based on the differential hydrolysis of glycosidic
mL after 60 h at an agitation rate of 200 rpm and aera- bonds, inulinase can be divided into exo-inulinases (E.C.
tion of 0.75 vvm when using 2% of the inulin extracted 3.8.1.80) or endo-inulinases (E.C. 3.2.1.7). Exo-inulinase
from dahlia tubers. K. marxianus ATCC 16045 was also functions by removing individual fructose from the non-
reported to produce a large amount of inulinase (121 U/ reducing end of inulin. Endo-inulinase can randomly
mL) and was proposed as a future inulinase overproducer break the β-2,1 glycosidic bonds in inulin, and can be
in yeasts [35]. Additionally, some marine yeast were also used to produce inulotrioses (F3), inulotetraoses (F4), and
found to efficiently synthesize inulinase. After screen- other IOSs that can be designated as GFn or Fn, where G
ing over 300 marine yeast strains from different marine and F stand for glucose and fructose, respectively (Fig. 2)
environments, Cryptococcus aureus G7a, a yeast found in [40]. The classical method of inulinase activity determi-
sediments from the South China Sea, was found to pro- nation is the Nelson–Somogyi colorimetric method [41].
duce the largest amount of extracellular inulinase. Under
optimal conditions, over 85.0 U/mL of inulinase activity Catalytic properties and mechanism of exo‑inulinase
was produced within 42 h of shake flask fermentation The exo-inulinase gene, INUI, was firstly cloned by
[36]. Fang et al. selected the yeast strain, Pichia guillier- Laloux et al. from K. marxianus ATCC 12424, and it had
mondii, from marine algae. Using optimal culture con- an ORF of 1668 bp and 555 encoded amino acids [42].
ditions, inulinase activity reached 61.5 0.4 U/mL after Then Wen et al. reported on the gene encoding exo-inuli-
±
48 h [37]. To further increase inulinase activity of the nase from K. cicerisporus CBS4857, and found that the
strain, mutant strain M-30, with a high inulinase activity gene structure contains an ORF of 1665 bp that encodes
(115.0 1.1 U/mL), was obtained after mutagenesis by 555 amino acids, including a signal peptide composed of
±
UV and LiCl. When using response surface methodology 23 amino acids [43]. The properties of inulinases from
(RSM) to optimize composition and conditions of the different microorganisms are summarized in Table  2.
medium, inulinase activity of the M-30 strain ultimately The molecular weight of exo-inulinase ranges from 40.0
reached 127.7 0.6 U/mL [38]. Bacteria are important to 256.0 kDa. As reported, most of the exo-inulinases
±
microorganisms in biorefinery and the inulinases they derived from fungi have a greater molecular weight than
produce are stable at a wide range of pH values and 50.0 kDa, such as C. aureus G7a (60.0 kDa), P. guillier-
higher temperatures, indicating application value. A ther- mondii strain1 (50.0 kDa), and A. niger 12 (65.0 kDa) [36,
mophilic inulinase-producing strain was isolated from 37, 44]. The optimal pH ranges were 5.0–6.5 for yeast
different soil samples using inulin as the sole carbon exo-inulinase, 4.0–6.0 for mold exo-inulinase, and 4.5–
source and culturing at 50 °C; fermentation conditions of 7.5 for bacterial exo-inulinase. For industry application
Fig. 2 Two modes of action on inulin by inulinases
Qiu et al. Biotechnol Biofuels (2018) 11:151 Page 6 of 15
and increasing solubility of inulin, thermal stability would
be an advantageous property of inulinase. The opti-
mal temperature of inulinase produced by yeast is usu-
ally 50 °C [45–47], while it was 60 °C for exo-inulinases
from Aspergillus ficuum JNSP5-06 [48] and Aspergillus.
fumigatus [49]. In addition, the optimal temperature for
exo-inulinases produced from B. polymyxa MGL21 is
35 °C [50]. The results indicate that optimal temperatures
of inulinases produced by various microorganisms are
different.
At present, most of works on inulinase are focused
on screening of inulinase overproducers and extraction
and purification of the enzyme. There are only a hand-
ful of studies on the mechanisms of inulinase. The crystal
knot of exo-inulinase derived from Aspergillus awamori
was determined for the first time by X-ray diffraction by
Nagem et al. [51]. As a member of the G32 family of gly-
cosyl hydrolases, exo-inulinase consists of two domains,
an N-terminal comprising 353 amino acid residues
(Phe20 to Gln372) and a C-terminal with 156 amino acid
residues (Arg382 to Asn537). The N-terminal domain is
the catalytic region of exo-inulinases and is composed
of 5 leaf-like helical fold structures. In the conserved
sequence 38-WMNDPNG-44 [motif A], the Asp41 resi-
due acts as a nucleophile to attack the substrate, forming
the intermediate between enzyme and substrate, and in
the sequence 241-ECPGL-245 [motif F], Glu241 func-
tions as a general acid/base catalyst that constitutes the
active site of the enzyme (Fig. 3a). Arg188 and Arg189 of Fig. 3 a Stereoview of the catalytic site of exo-inulinase from
the E motif (RDPKV) participate in the recognition and Aspergillus awamori. b Structural rearrangements in the EnIA putative
catalytic site of endo-inulinase from Arthrobacter sp. S37
binding of substrates. Moreover, Goosen et al. studied
the C-terminal conserved G motif (SVEVF) of A. niger
exo-inulinases and found that when Ser469 was replaced
with threonine residues, hydrolysis activity of the mutant high yields of inulotrioses (70.3  mM), inulotetraoses
in forming sucrose, inulin, and levan, was decreased. (38.8 mM), and inulopentaoses (3.5 mM) [54]. In addi-
When G motif Ser469 was replaced with the hydrophobic tion, bacterial endo-inulinases were also studied; Li et al.
valine residue, the enzyme was completely inactivated. reported that the Arthrobacter sp. S37 strain can produce
The results showed that the G motif is responsible for endo-inulinase EnIA (2439 bp). The molecular weight
hydrolysis of fructans and sucrose in the inulinase cata- is 75.0 kDa, and a pH of 7.5 and a temperature of 50 °C
lytic process [52]. are the optimal conditions for this purified enzyme.
The main products of inulin hydrolysis are inulotrioses,
inulotetraoses, and inulopentaoses [55]. A novel endo-
Catalytic properties and mechanism of endo‑inulinase
inulinase-producing strain, Streptomyces rochei E87,
Due to the health benefits of fructooligosaccharides,
was identified and found to degrade inulin into inulo-
the utilization of endo-inulinase to produce fructoo-
triose as the main product with a yield of 71% [56]. In
ligosaccharides from inulin is increasing. An endo-type
addition, Xanthomonas sp. was also implicated in inulin
inulinase containing 555 amino acids was purified from
hydrolysis, with the inulooligosaccharide conversion rate
Penicillium sp. TN-88 [53]. The enzyme is 68.0 kDa with
of 93% at pH 7, and DP 5 oligosaccharides [56, 57]. In
a high inulinase activity at pH 5.2 and 50 °C. Hydrolysis ≥
contrast to exo-inulinases (Fig. 4), the Glu (E) residue in
rates reached 70% after 72 h and the product was mainly
the motif A (WMNEPNG) of endo-inulinases has a key
composed of F3. Mutanda et al. used the purified endo-
role in the catalytic activity of the enzyme, while the con-
inulinase from A. niger for fructooligosaccharides pro-
served region (SVEVF) is important in anchoring mac-
duction. Molecular mass of the enzyme is 68.1 kDa and
romolecular glycans and attacking long chain inulin or
can act on inulin substrates at pH 6 and 60 °C, producing
Qiu et al. Biotechnol Biofuels (2018) 11:151 Page 7 of 15
Fig. 4 Sequence alignment of inulinase representative members
Qiu et al. Biotechnol Biofuels (2018) 11:151 Page 8 of 15
glycans (Fig. 3b) [58, 59]. Kim et al. studied the functional from Aspergillus ficuum and achieved FOSs (DP 2–8)
=
and catalytic mechanisms of endo-inulinases derived production of 80% using the Jerusalem artichoke juice
from Arthrobacter sp. S37. Three conserved amino acid as substrate after 72 h [85]. A newly discovered producer
residues (Glu323, Asp460, and Glu519) from 319-WMN- Aspergillus trtici BGPUP6 with 25.01 U/mL of endo-
DEPNGL-327 [motif A], 459-RDF-461 [motif E], and inulinase was considered as a candidate for FOSs produc-
519-ECMP-522 [motif F], respectively, were modified by tion [65]. Due to the low endo-inulinase activity by native
site-directed mutagenesis. The kcat values of E323A and microorganisms, recombinant endo-inulinases have been
E519A mutants decreased, but there was no variation in also used to produce FOSs from inulin in recent years.
Km, consistent with their putative roles as nucleophiles Yun et al. first cloned the endo-inulinase gene, INU1,
and acid–base catalysts. On the other hand, the D460A from Pseudomonas sp. and expressed constitutively in
mutant was completely inactive. These results proved Escherichia coli HB101 for FOSs production [86]. A yield
that Glu323, Asp460, and Glu519 are essential amino of 78% was obtained by using immobilized cells at 50 °C
acids for enzyme activity [60]. and continuously operated for 17 days without endo-
inulinase loss. E. coli BL21 (DE3) was also used to express
Research advances on the biorefinery potential
the endo-inulinase isolated from Aspergillus ficuum; the
of Jerusalem artichoke
highest endo-inulinase enzyme activity (75.22  U/mg)
Fructooligosaccharides
was obtained and an IOS yield of 94.41% was achieved
Fructooligosaccharides have been classified as prebiot- at 55 °C and pH 4.6 for 24 h [87]. Although a high FOSs
ics due to their bifidogenic nature and health-promot- conversion rate was achieved in an E. coli expression sys-
ing properties when consumed in sufficient amounts as tem, the system’s lack of safety greatly limits its applica-
recommended by health practitioners [77, 78]. From an tion in the food industry. Kim et al. cloned INU1 from
industrial point of view, FOSs are mainly enzymatically Pseudomonas mucidolens, after which the expressed
manufactured from sucrose using fructosyltransferases endo-inulinases in Saccharomyces cerevisiae cells hydro-
(DP < 4) or by controlled enzymatic hydrolysis of inulin lyzed inulin with an IOS yield of 71.2% after 30 h; the
(DP < 9) [79]; however, the former process is complex as IOSs mainly consisted of inulotetraoses [88]. On the
several reactions occur and is inevitably accompanied by other hand, a high yield of FOS production (91.3%) was
a residual of sucrose, that can lead to food-related obe- obtained from recombinant Pichia pastoris expressing
sity [80]. By contrast, the production of FOSs from inulin the endo-inulinase gene (EnInu) from A. niger [89]. As an
using endo-inulinases is a single-step process. Moreo- important value-added product from Jerusalem artichoke
ver, FOSs produced from inulin contain longer chains material, FOSs as function food have attracted extensive
(DP = 2–9) that have enhanced physiological activity attention. By recombinant expressions of endo-inulinase,
than those synthesized from sucrose (DP = 2–4). Recent a handful of reports on the FOSs production with high
advances in industrial enzymology have made the large- yields have been presented above which exhibits great
scale production of FOSs from inulin possible. potential for industrial applications, and developments
It has been reported that many filamentous fungi in separation of by product (glucose, sucrose) and impu-
and bacterial strains can produce endo-inulinases [81]. rity for high pure FOSs product is the need for future
Endo-inulinase from pseudomonas sp. was used for work. To simplify the IOS production process, high-
FOSs (DP = 2–7) production with a yield of 75.6% [82]. efficiency immobilization technology, novel reactors for
At the same time, Yun et al. reported FOSs yields of 83% biocatalysts should be explored to scale up to industrial
from inulin by using immobilized endo-inulinase from production.
Pseudomonas sp. The immobilized endo-inulinase reac-
tor operated for 15 days for FOSs production without a Ethanol
significant loss in enzyme activity which indicated the Ethanol is not only an important raw material in the
potential of industrial application [83]. Batch-wise pro- chemical synthesis industry, but it is also the most prom-
duction of FOSs from pure inulin was performed using ising biofuel. Among the biorefinery products of Jeru-
partially purified endo-inulinases from Xanthomonas salem artichoke, ethanol has been studied extensively.
sp. obtaining a yield of 86% and DP = 5–6 of the main However, the most commonly used alcohol production
product [57]. Cho et al. developed a dual endo-inulinase strains in the industry, S. cerevisiae and Zymomonas
system from Xanthomonas sp. and Pseudomonas sp. mobilis, cannot effectively utilize inulin to produce etha-
for the maximum FOSs yields of 92% from pure inulin. nol, which severely restricts the industrialization of etha-
In addition to FOSs production from inulin by bacte- nol production of inulin-based biomass [90]. Traditional
rial endo-inulinases, molds are also important sources ethanol production processes firstly convert inulin to
of endo-inulinase [84]. Jin et al. purified endo-inulinase fructose via acid hydrolysis, and then reuse the microbes
Qiu et al. Biotechnol Biofuels (2018) 11:151 Page 9 of 15
to ferment ethanol. Onsoy et al. used the acid hydro- ethanol. Ethanol fermentation from 200 g/L inulin and
lysates of Jerusalem artichoke juices as the media for 250 g/L raw Jerusalem artichoke tuber powder resulted
ethanol fermentation by Z. mobilis, which led to consist- in productivity of 2.44 and 3.13 g/L/h, respectively [97].
ent ethanol yields (0.45 g/g) and a conversion efficiency Actually, in addition to building engineering strains,
of 83.19% of theoretical value [91]. The cost of inulin acid some yeast genus, such as Kluyveromyces fragilis and
hydrolysis technology for ethanol production is low, but Kluyveromyces marxianus can both produce inulinase
the process is relatively complicated as hydrolysates often and ethanol. Rosa et al. selected K. marxianus to utilize
contain HMF, a microbial growth inhibitor, and is not extracted juice of Jerusalem artichoke tubers for ethanol
environment friendly [92]. Therefore, researchers begin production, the production of 12.8% (v/v) of ethanol in
to use microbes or enzymes to assist with S. cerevisiae 70 h with the consumption of 95% of initial sugars, and
in producing ethanol directly from inulin. Zhang et al. an ethanol yield 77% of the theoretical maximum were
used the recombinant inulinase produced by P. pastoris, achieved [98]. Yuan et al. studied ethanol fermentation
X-33/pPICZaA-INU1, to hydrolyze inulin and ethanol of K. marxianus ATCC8554 using inulin as substrate and
fermentation by S. cerevisiae sp. W0. The total sugar uti- the highest ethanol yield of 91.5% of the theoretical value
lization rate after 120 h of fermentation was 98.9%, and was achieved [99]. Above all, we believe that the use of
an ethanol yield of 0.384 g/g inulin was obtained [93]. Jerusalem artichoke in ethanol fermentation will become
The study further proved that inulinase pretreatment more economical and practical.
of inulin products for ethanol fermentation is feasible.
Although inulinase-assisted S. cerevisiae ethanol pro- Biodiesel
duction is an environmentally friendly technology, the Compared with bioethanol, biodiesel has a higher heat-
formed fructose inhibits inulinase secretion. Therefore, ing value and lower water absorption, and can be used
to relieve product inhibition, researchers have developed directly in vehicles without engine modification [100].
a co-fermentation technology on ethanol production Cheng et al. determined the feasibility of Jerusalem arti-
combined with inulinase-producing microorganisms. choke tubers as feedstock for biodiesel preparation,
Through mutagenesis, Ge and Zhang obtained an inuli- expecting to reduce the cost of microalgal cultivation
nase-producing strain, A. niger SL-09, and co-cultured it for bio-oil and biodiesel production [101]. In their study,
with the highly ethanol-tolerant S. cerevisiae Z-06. After Jerusalem artichoke hydrolysate was used as carbon
48 h of Jerusalem artichoke fermentation, the utiliza- sources for lipids accumulation by Chlorella protothe-
tion rate of inulin was 98%, and the ethanol concentra- coides. After 4-day scale cultivation, the lipid concentra-
tion reached 19.6% (v/v) [94]. Therefore, there is a need tion of 44% by dry mass was extracted and turned into
to further optimize microbial strains and the process, biodiesel by transesterification. The biodiesel contained
by improving microbial fermentation co-culture param- oleic acid methyl ester, linoleic acid methyl ester, and
eters to maximize efficiency of ethanol production from cetane acid methyl ester as the main components. Zhao
inulin. Since the genome of S. cerevisiae is well studied et al. also reported on lipid production from Jerusalem
and genetic manipulation techniques are more advanced, artichoke tubers using the oleaginous yeast, Rhodosporid-
more researchers are focusing on the use of genetic engi- ium toruloides Y4. The lipid titer of 39.6 g/L and cellular
neering strategies to obtain a recombinant Saccharomy- lipid content of 56.5% (w/w) were obtained when Jeru-
ces cerevisiae with inulinase that can directly produce salem artichoke hydrolysates were fed [102]. Sung et al.
ethanol from inulin. Tong et al. expressed the INU1 gene used a lab yeast strain of Cryptococcus sp. and achieved
from marine-derived P. guilliermondii and the recombi- lipid productivity of 1.73 g/L/d after optimization [103].
nant Saccharomyces sp. W0 was able to produce 34.2 U/ These studies suggest the feasibility of an alternative
mL of extracellular inulinase activity in 72 h. During 2 L method of producing biodiesel from Jerusalem artichoke
fermentation, 14.9% (v/v) of ethanol was obtained with tubers using microalgal cultivation, where a cost reduc-
the conversion efficiency of 99.5% from inulin to ethanol tion of carbon source feed in algal oil production can be
[95]. Yuan et al. cloned the exo-inulinase gene from Can- expected.
dida kutaonensis and expressed in S. cerevisiae for the
improvement of inulin utilization, and the recombinant 2,3‑Butanediol
S. cerevisiae was able to produce high ethanol yields from 2,3-Butanediol (2,3-BD) is an attractive chemical which
both inulin and Jerusalem artichoke tuber flour [96]. A can be used as a starting material for the manufacture
natural engineering S. cerevisiae engineered with rational of bulk chemicals such as 1,3-butadiene, methyl ethyl
strategies such as co-expressing exo- and endo-inulinase ketone, and so on [104]. As an important platform chem-
gene, inactivated proteases between haploid and dip- ical, the production of 2,3-BD is mainly by chemical or
loid was investigated for inulin utilization to produce biotechnological methods. With the depletion of crude
Qiu et al. Biotechnol Biofuels (2018) 11:151 Page 10 of 15
oil, biotechnological production of 2,3-BD has received Lactic acid
more and more attention [105]. Recently, several strains As an important platform chemical, lactic acid has been
of bacteria and fungi are considered to produce 2,3-BD, widely used in the many fields such as food, pharma-
including Aeromonas hydrophila, Klebsiella oxytoca, ceutical, and chemical industries. In order to construct
Pseudomonas hydrophila, Trichoderma harzianum, K. a cheap and energy-efficient process for lactic acid pro-
pneumoniae, and B. polymyxa [106]. From an economic duction, a variety of work has been undertaken for inex-
point of view, Fages et al. were the first to utilize Jerusa- pensive raw substrates. As early as 1942, Andersen and
lem artichoke tubers as the cheap raw material on effi- Greaves utilized Jerusalem artichoke tubers to produce
cient 2,3-BD production with B. polymyxa ATCC 12321. d-lactic acid [111]; however, by the 21st century, more
By optimizing k a profile, 44 g/L of 2,3-BD with a pro- researchers have begun to pay attention to the produc-
L
ductivity of 0.79 g/L/h was obtained during batch culture tion of l-lactic acid using Jerusalem artichokes. Ge et al.
[107]. Unfortunately, since then, little attention was paid creatively used a mixed-culture fermentation process,
to the microbial production of 2,3-BD from Jerusalem where A. niger SL-09 and Lactobacillus sp. G-02 were
artichokes. Until 2009, separate hydrolysis and fermen- used to directly form l-lactic acid from Jerusalem arti-
tation (SHF) and simultaneous saccharification and fer- choke tubers. The inoculation of l-lactic acid-producing
mentation (SSF) were successfully introduced to 2,3-BD strain Lactobacillus sp. G-02 greatly enhanced inulinase
production by K. pneumoniae from Jerusalem artichoke and invertase from A. niger SL-09, leading to the high-
tubers. In batch SHF process, the high concentration of est l-lactic acid concentration of 120.5 g/L with a high
fructose caused substrate inhibition for growth of cells conversion efficiency of 94.5% after 36 h of fed-batch
and target products accumulation, leading to a low 2,3- fermentation [112]. In their study, inulinase activity was
BD productivity of 1.08 g/L/h while SSF, which was pref- subjected to product inhibition in SSF, whereas fermenta-
erable for 2,3-BD production with high concentration tion activity of Lactobacillus casei G-02 was subjected to
(84.03 g/L) and productivity (2.1 g/L h) in fed-batch pro- substrate inhibition. To further enhance lactic acid pro-
cess [108]. Around the same time, Li et al. also reported ductivity, Ge et al. supplemented the media with sodium
that fed-batch SSF using K. pneumoniae was successfully citrate to maximize the specific growth and fructose con-
performed, and 80.5 g/L of target products (2,3-butan- sumption rates of L. casei G-02. As a result, a l-lactic
ediol and acetoin) were obtained by a stage-shift aera- acid yield of 141.5 g/L after 30 h was obtained by sup-
tion strategy after 68 h [109]. Li et al. also reported that plement of 10 g/L sodium citrate when inoculated a 10%
fed-batch SSF using K. pneumoniae was successfully volume of L. casei G-02 [113]. Shi et al. provided a way of
performed, and 80.5 g/L of target products (2,3-butan- producing l-lactic acid by immobilized cells. A fibrous-
ediol and acetoin) were obtained by a stage-shift aeration bed bioreactor was used to immobilized Lactococcus lac-
strategy after 68 h and used the thermophilic Bacillus tis cells for l-lactic acid production from hydrolysates of
licheniformis ATCC 14580 for 2,3-BD production from Jerusalem artichoke. The maximum l-lactic acid concen-
inulin, with the yield of 103.0 g/L in 30 h by fed-batch tration of 142 g/L was obtained in the fed-batch fermen-
SSF [105]. This is the highest 2,3-BD production reported tation [114]. Choi et al. found Lactobacillus paracasei
by utilizing inulin as the substrate nowadays. However, KCTC13169 could more efficiently ferment Jerusalem
all the above studies needed separate processes to hydro- artichoke tubers than other Lactobacillus spp. 92.5 g/L
lyze inulin of Jerusalem artichoke or exogenous inulinase of lactic acid was achieved by a direct fermentation from
added for 2,3-BD production. On the other hand, some Jerusalem artichoke extracts at 111.6 g/L of sugar con-
2,3-BD producers of Paenibacillus polymyxa were proven tent without acidic or enzymatic inulin hydrolysis. The
capable of fermenting inulin without previous hydroly- conversion efficiency of inulin to lactic acid reached 98%
sis. Gao et al. developed a one-step fermentation tech- of theoretical yield [115]. Recently, Wang et al. man-
nique of raw inulin extracts from Jerusalem artichoke aged to obtain high-optical purity of l-lactate from
tubers by P. polymyxa ZJ-9 to produce (R,R)-2,3-BD. hydrolysates of Jerusalem artichoke powder by using
Under optimal conditions, the concentration of obtained a thermophilic bacterium, Bacillus coagulans XZL4; a
(R,R)-2,3-BD reached 36.92 g/L, at more than 98% optical concentration of 134 g/L of l-lactic acid was obtained
purity [110]. This process greatly decreased the cost and and the optical purity of the final product was 99% [116].
facilitated its practical application in 2,3-BD production. With further research, utilizing Jerusalem artichokes for
By genetic engineering technologies, the future work on lactic acid production will become more significant for
manufacturing and modifying the industrial strains could industrialization.
have economic benefit on (R,R)-2,3-BD production.