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Saffron: Its Phytochemistry, Developmental

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DOI: 10.1021/acs.jafc.5b03194

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Saron: Its Phytochemistry, Developmental Processes, and

Biotechnological Prospects
Oussama Ahrazem,, Angela Rubio-Moraga, Sergio G. Nebauer, Rosa Victoria Molina,
and Lourdes Gomez-Gomez*,

Instituto Botanico, Departamento de Ciencia y Tecnolog a Agroforestal y Genetica, Facultad de Farmacia, Universidad de Castilla-La
Mancha, Campus Universitario s/n, 02071 Albacete, Spain

Fundacion Parque Cient co y Tecnologico de Castilla-La Mancha, Campus Universitario s/n, 02071 Albacete, Spain

Departamento de Biolog a Vegetal, Universidad Politecnica de Valencia, 46071 Valencia, Spain

ABSTRACT: The present state of knowledge concerning developmental processes and the secondary metabolism of saron,
Crocus sativus L. (Iridaceae), along with the genes involved in these processes so far known, is reviewed. Flowers and corms
constitute the most valuable parts of saron. Corm and ower development are two key aspects to be studied in saron to
increase the yield and quality of the spice, to raise its reproductive rate, and to implement new production systems. Important
knowledge about the physiology of owering and vegetative growth has been acquired in recent years, but there is still only
limited information on molecular mechanisms controlling these processes. Although some genes involved in ower formation
and meristem transition in other species have been isolated in saron, the role of these genes in this species awaits further
progress. Also, genes related with the synthesis pathway of abscisic acid and strigolactones, growth regulators related with bud
endodormancy and apical dominance (paradormancy), have been isolated. However, the in-depth understanding of these
processes as well as of corm development is far from being achieved. By contrast, saron phytochemicals have been widely
studied. The dierent ower tissues and the corm have been proved to be an important source of phytochemicals with
pharmacological properties. The biotechnological prospects for saron are here reviewed on the basis of the discovery of the
enzymes involved in key aspects of saron secondary metabolism, and we also analyze the possibility of transferring current
knowledge about owering and vegetative propagation in model species to the Crocus genus.
KEYWORDS: apocarotenoids, corm development, owering, avonoids, saron, saponins, stigmas, genes

INTRODUCTION
The genus Crocus comprises around 85100 species, primarily
distributed in the MediterraneanEurope and western Asia.
Turkey and the Balkan peninsula accumulate the major number
of species and subspecies. Greece alone contributes to ca. 40% of
the worlds wild Crocus diversity,1 whereas a total of 32 species
(18 of them being endemic) are included in Turkey ora.2
Several countries also have representatives of some Crocus
species: Italy (10 species), Spain (6 species), Hungary (6
species), among others. The genus Crocus is divided into two
subgenera:3 the subgenus Crocus, which includes all species
except one, and C. banaticus, which is the unique member of the
subgenus Crociris. The subgenus Crocus is further divided into
two sections: section Crocus and section Nudiscapus. This
subdivision of the genus is based on morphological and
cytological characters,3 as well as genetic analysis.4 Crocus sativus Figure 1. Most valuable parts of Crocus sativus: (A) saron owers; (B)
belongs to the series Crocus, commonly known as saron. Within corms; (C) stigma.
this series the plants have nely brous corm tunics, autumnal
owers, white or unmarked transparent membranous accid Historical data suggest that saron is a very ancient cultivation
bracts usually not closely sheathing the perianth-tube, yellow dating back from 2500 to 1500 BCE.6 The place of origin,
anthers, and three style branches, which are usually red due to the although not clear, is probably Greece, Iran, or Asia Minor, later
accumulation of crocins5 and often expended at the apex, entire
or at most mbriate.3 Received: June 30, 2015
C. sativus L. (Figure 1), a triploid geophyte, is a sterile plant Revised: September 25, 2015
propagated by corms and adapted to overcome the dry dormant Accepted: September 28, 2015
period of summer in the form of an underground corm.3 Published: September 28, 2015

2015 American Chemical Society 8751 DOI: 10.1021/acs.jafc.5b03194

J. Agric. Food Chem. 2015, 63, 87518764
Journal of Agricultural and Food Chemistry
spreading to India, China, the Mediterranean basin, and eastern
Europe.7
Saron is widely used mainly as a spice and as a coloring and
avoring agent in both the agro-food and cosmetic industries.
The saron spice is a complex mixture of volatile and nonvolatile
compounds and carotenoid derivatives, which all contribute to
the overall aroma and avor of the spice.8,9 In addition to its
color, taste, and aroma, saron displays a variety of health
benets.10 Saron shows analgesic and sedative properties11 and
is a potential anticancer agent.12,13 Furthermore, saron is
benecial for the treatment of mental disorders such as
depression14 and dementia.15
AGRONOMICAL IMPROVEMENT OF SAFFRON
PRODUCTION
The ideal condition for improving crop yield would be the
optimization of all metabolic cues together with the environ-
mental conditions where the crop develops.16 For such a
purpose, the rates of all important processes should be optimized
and also their interactions and duration, which are generally
determined by genetic mechanisms often inuenced by the
environment. Cultivation of saron occurs in temperate regions.
The life cycle of saron is adapted to the climate of the
Mediterranean regions and is quite similar in all producing
countries, but with wide dierences in the timing of events.17 In
the Mediterranean region, saron owers in autumn, after the
formation of replacement corms at the base of the shoots is
initiated. At the beginning of the dry season (AprilMay), the
leaves senesce and wither, and the corms go into dormancy. The
transition from the vegetative to the reproductive stage occurs
before summer in the apex of the buds of underground corms,
which do not have roots or foliage leaves.17 Not all of the newly
developed corms are able to produce owers. Flower formation is
directly related to corm size.18 Therefore, the adequate
production of corms is extremely important to guarantee ower
production.
During the past decades, the area traditionally dedicated to
saron cultivation in many European countries has fallen
dramatically,19,20 mainly due to the intensive manual labor
required for saron harvest and processing (0.252.5 million
owers ha1). However, in recent years this tendency has
reversed, and saron production in Europe and other developed
countries outside Eurasia, for example, in Tasmania (Australia),
is increasing, with a specic emphasis on the production of high-
quality saron. A technological breakthrough to increase the
quality of saron cultivation would entail the cultivation under
controlled conditions in containers to facilitate the mechaniza-
tion of ower harvesting and stigma separation.21 To achieve this,
the owering season must be extended as much as possible to
maximize the use of harvesting installations and thus reduce
overall installation and running costs. Thus, an in-depth
knowledge about the regulation of the owering process could
help to increase the yield of the spice.
Another important factor that limits the areas in which saron
is cultivated is the diculty in obtaining high-quality propagation
material with guaranteed levels of purity, homogeneity, and
health. It is a slow-growing species from which three to four
daughter corms per mother corm are obtained each season under
natural eld conditions. Low multiplication and fungal diseases
of corms reduce the productivity and quality, thereby restraining
the availability of planting material and the widespread use of
improved genotypes obtained in the future.
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Every sprouting bud in the mother corm has the potential to
produce a new corm, but saron corms show only a few
sprouting apical buds plus many axillary dormant buds.
Furthermore, there is a heterogeneity in bud emergence and
development, which results in heterogeneous ower emer-
gence.17 This process must therefore be controlled to establish
industrial production. In this context, environmental and genetic
factors aecting corm development, dormancy, and sprouting, as
well as the identication of environmental signals and the genetic
and regulatory mechanisms that promote or inhibit owering in
saron, constitute the most relevant processes to be elucidated
for implementing a technological breakthrough and achieving
agronomical improvements in this crop.
WHAT IS KNOWN ABOUT FLOWERING AND CORM
DEVELOPMENT IN SAFFRON?
Saron Flowering. The transition from the vegetative to the
reproductive stage in saron occurs early in July when the apex
becomes dome shaped and increases in size. This process is
followed by the formation of the leaf primordia. By mid July the
sheathing leaves begin to grow relatively quickly. Then follows
the bract primordia stage, the formation of stamen primordia, the
initiation of the perianth, and the formation of gynoecium. All of
the ower parts are already dierentiated by the end of August
(Figure 2). High temperatures are required to release bud
Figure 2. Saron ower development during summer: (A) resting bud
at the end of June; (B) stamen formation in early-mid August; (C) rst
whorl of tepals is already initiated in the ower at mid-late August; (D)
gynoecium has reached half of the length of the stamens in mid
September. Scale bars in panels AC = 0.5 mm. Scale bar in panel D = 1
mm.
dormancy and for ower initiation, which is optimal between 23
and 27 C21 and occurs during early spring to mid summer,
depending on location.2224 The release of bud dormancy could
be accelerated by curing the corms at 30 C for a short time.
Storage of corms at 2 C after ower initiation results in a time-
dependent abortion of those owers already initiated. Corms
stored in the cold before ower initiation will form owers when
incubated after storage at 2125 C. When corms are maintained
for a long time at very low temperatures (110 C), the
meristem does not give rise to ower primordia but to a new
DOI: 10.1021/acs.jafc.5b03194
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Journal of Agricultural and Food Chemistry
Figure 3. Development of saron daughter corms (DC) in the eld (Valencia, Spain): relationship between mother corm (MC) and daughter corm
sizes (owering took place in November): (A) in early December; (B) by the end of January; (C) in mid February; (D) by the end of February; (E) by
the end of April.
corm.19 Root development and ower emergence occur in late
autumn at a markedly lower temperature, in the range of 1517
C.21,25 This contrasts with spring-owering bulbous species,
which require a warmcoldwarm temperature sequence to
ower.26 The research carried out so far on saron has produced
extensive knowledge related to the inuence of temperature on
ower induction and owering and the extension of the
harvesting period over several months,17,21,22 but this strategy
has reached its peak.
Few works27,28 have addressed the role of hormones on
owering. In these works owering development is related to GA
activity. The involvement of other environmental or endogenous
factors has not been studied, and the molecular mechanisms
controlling these processes are largely unknown. The CEN/
TFL1-like gene (CENTRORADIALIS/TERMINAL FLOWER1)
has been cloned and characterized in saron.29 CsatCEN/TFL1
transcripts were detected in corms and ower organs but not in
leaves, and their role in saron owering control is still unclear.
Three FLOWERING LOCUS T (FT)-like genes, designated
CsatFT1-like, CsatFT2-like, and CsatFT3-like, have been
isolated.30 The expression analysis indicated dierences in the
expression of the three FT-like genes in dierent organs.
However, the expression has been measured in owers already
developed and in immature owers, but not in the run-up to
oral transition. Hence, the role of the FT-like genes awaits
further progress.
In relation to the molecular mechanisms controlling ower
development in cultivated Crocus, dierent full-length cDNA
sequences encoding MADS-box transcription factor proteins
involved in ower formation have been cloned and characterized.
Homologues of the major ABC and E types from Crocus have
been isolated and named Csat AP1/FUL,31 CsatAP2,29 CsatAP3
and CsatPI,32 and CsatAG33 and CsatSEP3.34 However, the
expression analysis did not always show consistent results with
the ABC model, and a functional understanding has to be
claried. For example, the three isolated CsatAP1 genes were
expressed in leaves, as well as in the three mature ower parts:
tepals, stamen, and carpel. Furthermore, an expanded expression
8753
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of class B genes in whorls 1 and 4 was also detected. The
expression of class B genes into whorl 1 ts the modied ABC
model for monocots; however, the additional expression is not
consistent with this model.
Corm Development. There is limited knowledge about the
physiology of vegetative development of saron (Figure 3). The
relative contribution of dierent carbon sources (leaf photo-
synthetic activity and mother corm reserves) to the growth of
vegetative organs and the inuence of internal and external
factors on the photosynthetic processes sustaining corm growth
have been studied.19 After owering, most of the remaining
reserves in the mother corm are depleted during root and leaf
development. Once these organs reach their maximum size, the
growth of replacement corms is initiated, and this development
primarily relies on photosynthesis. During this vegetative
development, the reserves from the mother corm contribute to
only 10% of the biomass. The photosynthetic rate is consistently
high and constant (26 mol m2 s1) throughout the year, as has
also been reported in C. vernus.35,36 Plants with larger corms
show reduced photosynthetic rate, although a signicantly higher
total leaf area is present.19 A sink capacity limitation arises during
growth of replacement corms because the increase of biomass is
limited before total leaf senescence. A reduction in the demand of
photoassimilates can lead to sugar accumulation in leaves, which
in turn can promote leaf senescence.37 The studies carried out in
C. vernus, a spring ephemeral, support the hypothesis that corm
sink strength is the most important factor determining leaf
duration.35,38 Dierences in sink strength have been associated
with dierential carbon partitioning between sugar forms in the
newly formed corms. A higher availability in hexoses at the
beginning of the development of the daughter corms would aect
cell division and elongation and, thus, higher corm size that
allowed prolonged sink activity.36 However, an in-depth
knowledge about the environmental and genetic factors
controlling the nal size and the sink strength of the saron
corm still seems far from being achieved.
Sprouting Control in Saron. Sprouting is another
important aspect in saron research. As stated previously,
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every sprouted bud produces a corm and factors aecting apical
dominance (para-dormancy) are highly important for corm
production. However, the control of dormancy (endo-
dormancy) is also crucial in solving the many problems
associated with storage and distribution of corms and for
extending the owering and harvesting periods. Physiological
and molecular studies support the concept that the mechanisms
of owering induction and dormancy are interrelated.39 It has
been suggested that dormancy-associated MADS-box (DAM)
genes induced by cold temperatures in leafy spurge negatively
regulates FT or similar genes, which regulate growth cessation
and dormancy induction. Most of these studies were performed
on species showing winter dormancy, where dormancy was
regulated by low temperature and a short photoperiod. However,
saron shows summer dormancy, which is common in typical
bulbous species. This plant strategy has been correlated with
superior survival and high persistence under severe drought
conditions. Summer dormancy is therefore of great interest
because intense drought and heat are increasing due to climate
change and may aect plant persistence. However, this type of
dormancy is relatively uncharacterized at a molecular level.
Recently, the characterization of summer dormancy in the
bulbous species Narcissus tazetta and the role of FTs in summer
dormancy has been reported.40 In saron, FT genes and CEN/
TFL1-like genes that could be related with the dormancy
pathway have been isolated.30
Dormancy interacts at some level with the mechanisms
involved in cell cycle regulation, which also involves hormonal
signals.39,41 Studies of the hormonal regulation of dormancy in
geophytes are abundant.42 Clear roles for abscisic acid (ABA),
such as inducing dormancy, have been revealed. ABA has been
recognized to control dormancy in the corm tissue of C. sativus,
where it declines at the time of dormancy release.28 It has been
proposed that ABA is synthesized from carotenoids (C40) in
plants, and three genes that participate in ABA biosynthesis have
been isolated and characterized in several plant species. They
encode zeaxanthin epoxidase (ZEP43), 9-cis-epoxycarotenoid
dioxygenase (NCED44), and abscisic aldehyde oxidase (AAO45).
ZEP catalyzes the epoxidation of zeaxanthin to produce
epoxycarotenoid; NCED catalyzes the cleavage reaction of
epoxycarotenoids to produce xanthoxin (the rst C15
intermediate) and is considered to be the regulatory enzyme of
ABA biosynthesis; and AAO catalyzes the nal step of ABA
biosynthesis, which converts ABA aldehyde to ABA. Homologue
genes for ZEP, AAO, and NCED have been identied in saron.
However, only one NCED gene, CsNCED, has been studied in
detail.46 CsNCED is expressed in oral and corm tissues. In
stigmas, CsNCED transcript abundance closely mirrors ABA
content with a maxima a day after anthesis, and it has been
suggested that the ABA levels in the stigma could be related to
the initiation of the senescent process in this tissue.47 During
corm development, zeaxanthin levels are correlated with the
increase in CsNCED expression, suggesting a coordinate
regulation of the carotenoid and ABA biosynthetic pathways
during corm dormancy.46
Mature saron corms usually show one to three apical
dominant buds, which will sprout in the following season plus
many axillary dormant buds.22 Each axillary bud has the same
developmental potential as the primary shoot apical meristem in
its ability to produce a growing shoot axis (Figure 4). However,
axillary buds enter a dormant state after forming only a few
leaves. Plant hormones are major players in the control of axillary
bud outgrowth. In particular, auxin, strigolactone, and cytokinin
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Review
Figure 4. Apical dominance in saron: (A) corm with the main bud
sprouting and dormant axillary buds; (B) axillary bud growth after
removal of the main bud.
are involved in this control.48 In saron, auxin is supplied from
the apical bud and, together with the strigolactones (SL),
represses axillary bud outgrowth. By contrast, the cytokinins,
produced in the adventitious roots, directly induce the sprout of
the axillary buds.49 Gibberellins (GAs) seem to be involved in
apical sprout growth after dormancy break, but not in dormancy
maintenance or release.28
SLs are derived from the oxidative cleavage of carotenoids and
have been implicated in many processes including root growth,
root hair and stem elongation, lateral root formation,
adventitious rooting, secondary growth, leaf expansion, leaf
senescence, and drought and salinity responses but most
prominently in shoot branching.48 SLs seem to act systemically
to dampen the polar auxin transport stream, which enhances
competition among buds for a common auxin sink.50 To date,
four loci involved in SL biosynthesis and three loci involved in SL
signaling have been identied.5153 The rst steps of SL
biosynthesis involve isomerization by the plastidial-isomerase,
followed by the sequential cleavage activity of CCD7 and CCD8
carotenoid cleavage dioxygenases. MAX1 encodes a cytP450
predicted to act downstream from CCD7 and CCD8 and is
required for synthesis of active SL. Among all of the identied
components of the strigolactone biosynthetic pathway, CsCCD7
and CsCCD8 genes have been recently isolated and studied in
saron.54 SL production related to bud dormancy in saron
seems to be controlled at the CsCCD8 level. In addition,
CsCCD8 seems to play an important role in the development of
the vasculature of the axillary buds in saron. Recent evidence
shows that SLs positively regulate cambial activity, and this
function is conserved among species. Saron is a monocot
without cambial activity. However, given the relationship
between cambial and procambial cells, it can be postulated that
SLs could also positively regulate procambium activity and the
development of vascular tissues in growing buds of monocots.
THE PHYTOCHEMISTRY OF SAFFRON:
COMPOUNDS IDENTIFIED FROM SAFFRON
FLOWERS AND CORMS
Apocarotenoids in Saron Flowers, Biosynthesis, and
Properties. Apocarotenoids are the products of the oxidative
cleavage of carotenoids by specic carotenoid cleavage oxy-
genases (CCDs) that recognize and specically cleave one or two
double bonds.55 If sucient numbers of conjugated double
bonds are maintained in the cleavage products they can show
coloration, like their parent carotenoids. This is the case of the
crocetin molecule (8,8-diapo-8,8-carotene-dioic acid) (Figure
5), widely used as a colorant in foods and cosmetics. The crocetin
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Figure 5. Pathway for crocin and picrocrocin biosynthesis in saron stigmas.
molecule is further modied by the activity of glucosyltrans-
ferases,56,57 which add dierent numbers of glucose molecules to
produce characteristic crocins, these being the major compo-
nents of the stigmas of saron and which confer solubility.58,59
Crocins are widely present in the species of the genus Crocus,60
but have been also identied in Buddleja ocinalis,61 in Jacquinia
angustifolia,62 in Coleus forskolii,63 in Nyctanthes abor-tristis,64 in
the fruits of Artocarpus heterophyllus,65 Gardenia jasminoides,66
and even in the cyanobacterium Microcystis.67
Crocetin is formed from the enzymatic cleavage of zeaxanthin
on the 78/78 double bonds (Figure 6).68,69 The resulting
products are crocetindial and hydroxyl--cyclocitral, which is
further glucosylated to form picrocrocin (-D-glucopyranoside-
hydroxyl--cyclocitral), the degradation product of which is the
odor-active safranal.69 Carotenoid biosynthesis, cleavage activ-
ities, and the expression of corresponding genes have been
studied during the development of the stigma of C. sativus. A total
of ve CCD genes belonging to two classes of cleavage enzymes
were identied from C. sativus.68,70,71 Heterologous expression of
CsCCD1a, CsCCD1b, CsCCD4a, CsCCD4b, and CsCCD4c
sequences reduced accumulation of -carotene in Escherichia coli
strains engineered for the accumulation of this carotenoid.68,72
As for volatile cleavage products produced in these strains, only
C13 -ionone could be detected, but no C10 apocarotenoids,
whereas a 910/910 cleavage activity for both types of
enzymes is predicted (Figure 6). Analysis of stigma volatiles in
dierent stages of ower development indicated high production
of -ionone shortly before and at anthesis.68,73 In the plant,
CsCCD1a and CsCCD4a/b dier if their subcellular location is
cytosolic or plastidial, respectively.68 CsCCD4a sequences, most
likely constituting an allelic variant of CsCCD4b (98% of
identity) turned out to be identical to a Crocus ZCD sequence,70
albeit CsZCD is a truncated sequence compared to CsCCD4a,
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with 200 fewer amino acids. For CsZCD a 78/78 cleavage
activity in vitro was reported.70 However, CsZCD lacks
important residues and domains for a dioxygenase activity,74
and no activity was found for this CsZCD in later trials.68,75
Mining of an EST database from C. sativus stigmas has provided
further information on potential saron biosynthetic genes but
no candidate for a novel CCD with 78/78 cleavage
specicity.76 Therefore, the previously isolated enzymes were
analyzed against a battery of dierent carotenoids, including
zeaxanthin and lutein,77 and an enzyme, CsCCD2, with a
conrmed specicity for the 78/78 double bonds was
identied, showing 97% identity with the initially identied
CsCCD1b68 (Figure 6A). On the basis of the in vitro results,
lutein and zeaxanthin, but not -carotene, are good substrates for
CsCCD2, and the availability or accessibility to the enzyme
would determine the specicity. Accumulation of apocarotenoids
in saron is stigma specic and appears to be developmentally
regulated.73 Higher levels of saron apocarotenoids increase as
stigmas develop and reach their peak in the red stage,73 and it is
likely that the genes involved in their biosynthesis may be
stimulated during the process and enhanced in the stigma tissue.
CsCCD1b and CsCCD2 expressions are limited to the stigma
tissue and detected in the earlier developmental stages, when
crocetin accumulation takes place.68 Analysis of saron EST
collections obtained from the stigma at dierent developmental
stages revealed that CsCCD2 ESTs were more highly represented
in libraries obtained from early stages.77 Recently, homologues of
CsCCD2 have been identied in spring Crocus species, which
accumulate crocins in stigmas and tepals.78 All together, the
CCD2 enzymes constitute a new CCD subfamily with no
homologues identied in other plant species so far.
Apocarotenoid Properties. The stigmas of C. sativus have
been used in folk medicine to alleviate dierent health
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Figure 6. Carotenoid cleavage dioxygenases identied in the stigma of saron: (A) phylogenetic relationships among the isolated CsCCDs; (B) cleavage
activity for the isolated CsCCDs enzymes.
problems.79 Recent pharmacological studies have demonstrated
that crude extracts from saron and puried chemicals possess
antitumor eects, display anti-inammatory properties, and
counteract atherosclerosis and hepatic damage.11,12 In studies on
animal models, it has been shown that crocetin has several
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pharmacological properties, including antioxidant,80,81 anti-
inammatory,82 antiatherosclerotic,83 insulin resistance improve-
ment,84 attenuation of physical fatigue,85 and sleep.86 Crocetin
also alters the growth of cancer cells by inhibiting replication,
inducing apoptosis, and enhancing the antioxidative system.87
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Journal of Agricultural and Food Chemistry
Crocetin shows neuroprotective eects.88,89 Crocetin inhibits in
vitro amyloid- aggregation,90 and a clinical pilot study revealed
signicant improvements in cognition after the treatment of
mild-to-moderate Alzheimers disease patients with crocetin.91
Crocins had protective eects on neuronal injury,92 and
attenuated the symptoms of obsessive compulsive disorder, a
common psychiatric disorder dened by the presence of
obsessive thoughts and repetitive compulsive actions.93 Safranal,
the main constituent of the volatile oil fraction, attenuated
cerebral ischemia94 and retinal degeneration.95,96
Flavonoids in Saron Flowers. Flavonoids are among the
best-characterized plant secondary metabolites in terms of
chemistry, coloration mechanism, biochemistry, genetics, and
molecular biology. With a basic structure of C6C3C6, they
are widely distributed among land plants. Flavonoids are the
result of the contribution of two pathways, the avonoid branch
of the phenylpropanoid and the acetatemalonate metabolic
pathway. Although the central pathway for avonoid biosyn-
thesis is conserved in plants, depending on the species, a group of
enzymes, such as isomerases, reductases, hydroxylases, and
several Fe2+/2-oxoglutarate-dependent dioxygenases, modify the
basic avonoid skeleton, leading to the dierent avonoid
subclasses.97 Finally, transferases modify the avonoid backbone
with sugars, methyl groups, and/or acyl moieties, modulating the
physiological activity of the resulting avonoid by altering its
solubility, reactivity, and interaction with cellular targets.98 In C.
sativus, avonols are usually glycosylated at their 3-OH, 7-OH
and 4-OH positions,8,99,100 producing a complex pattern of
avonols. In C. sativus stigmas three main glucosides of
kaempferol have been identied: kaempferol 7-O-sophoroside,
kaempferol 3-O-sophoroside-7-O-glucopyranoside, and kaemp-
ferol 3,7,4-triglucoside. By contrast, 21 dierent glycosides of
isorhamnetin, kaempferol, myricetin, naringenin, quercetin,
tamarixetin, and taxifolin have been identied in tepals,101
kaempferol and kaempferol glycosides being the most dominant
class of avonoids in this tissue (70 and 90% of the total content
of avonoids) followed by quercetin glycosides (510%).102 In
pollen kaempferide, a methylated kaempferol and the
isorhamnetin glycosides isorhamnetin-3,4-diglucoside, isorham-
netin-3-O-robinobioside, and isorhamnetin-3--D-glucoside are
detected.100 In addition, the anthocyanins delphinidin, petuni-
din, and malvidin have been identied in tepals, with dierent
sugar substitutions,102 and are responsible for their blue color.
Transcriptome analysis of saron stigmas has allowed the
identication of genes encoding for key enzymes of the avonol
pathway, and putative UDP-glycosyltransferase enzymes have
been identied (data not shown). To date three genes encoding
avonol glycosyltransferases have been characterized in
saron.99,103 The saron CsGT45 (UGT75P1) protein showed
specicity toward avonoid aglycones and was found to be active
on the C-7 position of kaempferol, kaempferol 7-O-sophoroside
being the most abundant kaempferol glucoside present in the
stigma tissue. The UGT707B1 enzyme was recently charac-
terized as a avonol-3-O-glucoside: 2-glucosyltransferase. The
expression of UGT707B1 in the oral organs of C. sativus allowed
the accumulation of a specic set of avonols that inuence the
transport of the plant hormone auxin and could be responsible
for the characteristic morphology of the stigma.103 The last UGT
characterized in saron involved in avonoid glucosylation is
UGT703B1.104 This enzyme recognizes isorhamnetin and
kaempferol as substrates in vitro. In addition, UGT703B1
expression was found to be highly correlated with the presence of
kaempferol 7-O-biglucoside-3-O--glucoside and isorhamnetin-
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3,7-O-diglucoside. These avonols are present in the owers of
Crocus sp. from the series Crocus, but absent in Crocus species
from other series, and could be used for quimiotaxonomic
purposes.
Flavonoid Properties. In plants, avonoids have many
diverse functions including UV protection, defense, allelopathy,
ower coloring to attract pollinators, auxin transport inhibition,
plantmicroorganism communication, and regulation of reactive
oxygen species, and in many species they are required for pollen
viability.105 A number of biological processes, such as signal
transduction, transcriptional regulation, and cell-to-cell commu-
nication, are also inuenced by avonoids. In addition, they play
crucial roles in human nutrition, and many therapeutic benets of
avonoids are known in animal systems. Flavonoids have
antioxidant, anti-inammatory, antitumor, antiproliferative,
cardioprotective, and pro-apoptotic activities.106109
The main avonoid present in saron tepals is kaempferol-3-
O--D-glucopyranosyl-(12)--D-glucopyranoside (kaempferol
3-O--sophoroside).103 Interestingly, this glucosylated avonoid
is not present in all Crocus species, and its presence is relegated to
a few series, including the Crocus series. Kaempferol 3-O--
sophoroside has been reported to have analgesic activity.110
Furthermore, kaempferol 3-O--sophoroside inhibits the
vascular endothelial inammatory responses and is considered
a promising target for the treatment of vascular diseases such as
atherosclerosis, shock, heart attack, and sepsis.111,112 The
identication in saron of the rst UGT involved in the
formation of this glucosylated avonoid opens new possibilities
for the production of this compound in other crop species or in
other heterologous systems.
SAPONINS IN SAFFRON CORMS
Saponin Biosynthesis. Saponins are a widespread group of
terpenoids. Saponins are important plant defense compounds,
which are determinant for disease resistance due to their
antimicrobial, fungicidal, allelopathic, and insecticidal activ-
ities.113 Humans exploit saponins for their medicinal properties
and antimicrobial activity. The composition of saponins depends
on the genetic background of the plant material, the tissue type,
and the age and physiological state of the plant as well as
environmental factors.114 The name saponin is derived from
sapo, soap in Latin, because the surfactant properties produce
soap-like foams in aqueous solution. This trait is the result of the
amphiphilic nature of saponins due to linkage of the lipophilic
sapogenin to hydrophilic saccharide side chains.
According to the chemical nature of the aglycone (known as
sapogenin), the saponins are divided into steroidal and
triterpenoid saponins. Steroidal saponins are produced especially
in some families of the monocotyledons, for example, Agavaceae,
Alliaceae, Asparagaceae, Convallariaceae, Dioscoreaceae, Lilia-
ceae, Smilacaceae, and Trilliacea. In dicotyledons, steroidal
saponins are detected only in a few families, for example,
Fabaceae, Solanaceae, and Scrophulariaceae.115 Cholesterol is
considered the starting molecule for the biosynthesis of steroidal
saponins.116 The rst committed step in the biosynthesis of
triterpenoid saponins and phytosterols is the cyclization of 2,3-
oxidosqualene by oxidosqualene cyclases (OSGs). The high
number of possibilities for establishing dierent internal linkages
during cyclization gives rise to a vast array of dierent triterpene
skeletons.117 Following formation of basal sapogenin backbone
structures mediated by OSCs, various modications take place
prior to glycosylation, the most common being the addition of
small functional groups such as aldehyde, hydroxyl, keto, and
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Journal of Agricultural and Food Chemistry
Figure 7. Structures of the triterpenic saponins, azafrines 1 and 2, identied in the corm.
carboxyl moieties at dierent positions of the backbone, most of
them introduced by cytochromes P450 enzymes.118 Saponin
glycosylation involves sequential activity of dierent UGTs.98
Typical triterpenoid saponin glycosylation patterns consist of
oligomeric sugar chains, two to ve monosaccharide units, which
are most often linked at position C3 and/or C28.
In saron, two triterpenic saponins have been identied in the
corm, azafrines 1 and 2119 (Figure 7). Azafrine 1 contains 3-O--
D-glucopyranosiduronic acid echinocystic acid as prosapogenin,
and azafrine 2 contains 3-O--D-galactopyranosiduronic acid
echinocystic acid as prosapogenin. Both prosapogenins are
substituted at the C28 position with a pentasaccharide consisting
of -D-galactopyranosyl-(12)--L-arabinopyranosyl-(12)-
[-D-xylopyranosyl-(14)]--L-rhamnopyranosyl-(12)--D-
fucopyranose, and a 4-O-di--L-rhamnopyranosyl-3,16-dihy-
droxy-10-oxo-hexadecanoyl group is linked to C-4 of the -D-
fucopyranosyl residue. Azafrines 1 and 2 are localized in the
external part of the corms, suggesting their involvement in plant
defense.119,120
Saponin Properties. Saponins or saponin-containing plant
extracts are exploited by the pharmaceutical industry. Several
saponins, including the ones identied in saron, exhibit
pharmacological activities and thus attract attention as a target
for drug discovery. Saponins are valuable adjuvants, and the rst
saponin-based vaccines have been introduced commercially.121
In addition, a recent study demonstrates the potential use of
azafrines 1 and 2 in human immunotherapeutic vaccines,122 and
they can also be used for further industrial applications, for
instance, preservatives, avor modiers, and agents for removal
of cholesterol from dairy products123
PROSPECTS FOR IMPROVING AND INNOVATING
SAFFRON PRODUCTION: THE ROLE OF GENOMIC
TOOLS AND BIOTECHNOLOGICAL APPLICATIONS
All over the world saron is known as one cultivar. The
continuous vegetative cultivation of saron oers certain
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advantages in terms of preservation of the genetic characteristics
of the plant, but makes any genetic improvement dicult.
Variability studies with saron from dierent countries and
locations showed the absence of or very low variability among
entries.124126 This insucient genetic variability makes the use
of this plant dicult for selection programs,127,128 whereas the
induction of genetic variability through mutagenesis has been
addressed using corms at dierent developmental stages. Even
though mutations have been identied as resulting from
experimental as well as natural mutagenesis,129,130 these are
not maintained because they are not heritable.
In C. sativus, many experiments have been performed to obtain
seeds131 and in vitro regenerated plants132 without success. As an
alternative, researchers have been trying to elucidate how the
triploid species was generated to identify the progenitors or at
least the closest relatives of C. sativus. An autotriploid origin was
initially suggested for saron. However, an allotriploid origin
from two wild species has been proposed in dierent
papers,102,133,134 C. cartwrightianus being one of the accepted
progenitors.5,34,135 Nevertheless, no such eort has been
undertaken to exploit the closet relatives and hybrid generation.
A rst EST collection from saron was developed in 200776
comprising 6603 high-quality ESTs from a saron mature stigma
cDNA. The rapid evolution of next-generation DNA sequencing
technologies has allowed the analysis of the transcriptome of six
dierent developmental stages from saron stigma (unpublished
results). This will allow deciphering key players involved in one
of the most important processes in saron from the
biotechnological point of view such as apocarotenoid metabo-
lism.
However, it is necessary to point out that one of the most
important problems in applying modern biotechnological
approaches to saron and geophytes in general is the lack of
high-frequency transformation systems. Further eorts should be
made to work in that direction.
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Journal of Agricultural and Food Chemistry
Secondary Metabolism and Biotechnological Applica-
tions. The current state of the art regarding secondary
metabolism in saron has been discussed above. The
identication of the key enzymes involved in apocarotenoid
formation represents a very important tool for future
biotechnological applications in other crop species where the
substrates for crocetin formation are present. This is the case of
maize and tomato, where introduction of these enzymes will
provide intense red colors and higher nutritional quality, due to
the higher bioavailability of the saron apocarotenoids compared
with carotenoids.136 In addition, rice and tobacco are also good
candidates to increase the production of crocins, providing added
value to the crop, in the case of rice, or using the plant as a
cheaper way to produce crocetin in sucient quantities for
dierent industrial applications. In this regard, it is also important
to consider other Crocus species as alternative sources of saron
apocarotenoids, paying special attention to those that accumulate
crocins in both stigma and tepals,60 providing an attractive crop
from an economic standpoint, due to the reduction of processing
costs.
With regard to corm saponins, the genetic machinery required
for the elaboration of this important family of plant secondary
metabolites is as yet largely uncharacterized in saron, despite
the considerable commercial interest in this important group of
natural products. The development of transcriptomes from the
corm combined with metabolomic analyses will allow the
identication of key enzymes involved in their biosynthesis.
Nevertheless, still little is known about the genetics that
control quantitatively and qualitatively the biosynthesis of these
secondary metabolites in saron and its allies. In addition,
knowledge of the regulation of these pathways is practically
nonexistent, and such knowledge is of crucial importance to
bypass the possibilities of low product yield of these secondary
metabolites in other plants or plant cell cultures.
Control of Flower Development. Understanding ower-
ing regulation at the molecular level in saron, and also in other
spring and autumn Crocus, would provide the potential for more
accurately controlling both owering time and the development
of transcriptional markers for ower initiation to increase genetic
variation of saron for owering time from spring Crocus using
biotechnological and omics tools.
Although several key owering genes (FT, CEN/TFL1, and
AP1) have been found in saron, understanding oral inductive
pathways in Crocus requires not only the identication of some
possible regulator genes but a precise knowledge of their
expression in the dierent developmental stages and their control
by environmental or internal factors. To date, we are far from
achieving this knowledge. Limited information is available on the
ower transition of bulbous species, which require high
temperature for ower dierentiation in species such as saron.
The results obtained by Li et al.137 working on Narcissus tazetta, a
bulbous species that requires high temperature for ower
dierentiation, showed that Narcissus FLOWERING LOCUS
T1 (NFT1) transcripts were abundant during ower initiation in
mature bulbs and were up-regulated by high temperature,
suggesting that NFT1 possibly takes part in ower transition
control in response to this factor. Given the similarities in the
oral induction process of saron and narcissus, the same up-
regulation of FT by high temperature could be expected to
happen in saron, although this needs to be conrmed.
Also, the role of CsatCEN/TFL1 needs to be claried.
Although the involvement of TFL1 genes in oral transition
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and axillary meristem identity has been demonstrated in other
monocot species,138 their role in saron is not clear.
As previously mentioned, treating dry saron corms with GA
in JuneJuly accelerated ower formation and promoted the
formation of additional ower buds from undierentiated
meristems. Furthermore, some observations point out that
GAs and nutrients act through a common signaling pathway to
promote owering.139 This pathway controlling owering in
response to gibberellin and nutrients is highly recommended to
be studied in saron. Because ower dierentiation takes place in
summer when the corm is located underground with no foliage
or roots, the involvement of the photoperiod and vernalization
pathways seems unlikely, but they should not be entirely ruled
out. Another important aspect is related to the decision to bloom,
which requires lower temperatures than for ower induction.
The genetic control of this process has not yet been studied.
Understanding ower induction in saron could reveal ways to
extend the harvesting period, but the introduction of genetic
variation from other Crocus species by means of biotechnological
tools should also be considered. Furthermore, the development
of new genotypes from C. cartwrightianus should not be excluded.
There is a high variability for owering time in the Crocus genus.
This fact enables us to establish a research line aimed at acquiring
in-depth knowledge about the dierences in the owering
processes at the molecular level between autumn and spring
Crocus species and making use of this knowledge to extend the
saron owering period.
Another interesting biotechnological prospect has been
pointed out by Tsaftaris and co-workers.33 It would be desirable
to have mutants without stamen formation in saron owers,
because removal of stamens and separation of stigmas by hand is
very labor intensive, leading to high costs. More interesting
would be the transformation of stamens to carpel, which should
increase saron production in a single ower. However, it is not
at all clear that saron production would be doubled. The
identication of saron ower development genes that has been
already carried out, as mentioned above, will facilitate the
achievement of these objectives, and the usefulness of these
materials can be tested.
Control of Saron Sprouting. The understanding and
control of dormancy are critical for eectively managing crop
production, commercial handling, and storage. Control of endo-
dormancy in saron store corms is a requirement for staggering
the production, and reliable markers for endo-dormancy in
saron and other geophytes are one of the most important issues
in horticultural practice. The saron genes ZEP, AAO, NCED,
and dormancy-associated MADS-box (DAM) genes could be
used as markers to determine the exact time at which
environmental or internal factors trigger dormancy. It is evident
that the application of molecular techniques is indispensable for
clarifying the precise mechanism(s) of dormancy induction and
release in bulbous species, and the establishment of such a model
system is crucial for further research on dormancy of bulbous
species.42
Another important aspect of dormancy regulation in the
Crocus genus is the dierential response of various plant organs to
environmental signals as has been pointed out by Okubo.42 The
genus includes both synanthous and hysteranthous species. In
both types, ower initiation occurs in summer, and the initiation
orders of the leaves and owers are similar. However, a time
interval takes place between leaf elongation and anthesis in
hysteranthous species, when compared to synanthous species
(Figure 8). Hence, dierent organs might vary in their dormancy
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Journal of Agricultural and Food Chemistry
Figure 8. Crocus genus includes both (A) synanthous (e.g., Crocus
sativus) and (B) hysteranthous (e.g., Crocus nudif lorus) species.
requirements, and the process might be controlled by both
external and internal signals. The absence of leaves at owering
time is of great interest in saron collection to make harvest work
easier, and the understanding of that dierential response of
leaves and owers in hysteranthous species takes on great
importance.
The rate of propagation in saron is directly related to axillary
meristem formation and outgrowth, and it has been shown that
there is a control exerted by the main shoot apex of the corm over
the outgrowth of the lateral buds mediated by a hormonal
balance. Auxin and strigolactones inhibit bud outgrowth, and
cytokinins promote it. A bud-specic gene that promotes bud
arrest could be the key element to integrate the bud outgrowth
pathway.140 Such a gene exists in maize (Teosinte branched
1TB1),141 and there is evidence that the gene BRANCHED1
(BRC1) acts as an integrator of branching signals within axillary
buds of Arabidopsis.142 BCR1 is up-regulated by SLs and down-
regulated by cytokinins. Because outgrowth of axillary buds
seems to be the major limiting factor in vegetative propagation of
bulbs and corms, the SL signaling pathway and BRC1-like genes
are important targets of choice to study and optimize vegetative
propagation.140
Improvement of Corm Growth. The manipulation of
assimilate partitioning and allocation within the plant is a key
factor in the improvement of corm growth. This goal could be
achieved through biotechnological approaches based on the
identication of the involved genes and regulatory pathways.
Photosynthesis eciency can be improved through dierent
targets. In relation to light reactions, it could be achieved by
increasing the eciency of light capture, energy transduction,
and photoprotective processes, and for carbon xation reactions
by reducing limitations in the diusion of CO2 to the site of
carboxylation (e.g., mesophyll conductance), in the capacity for
carboxylation/oxygenation of Rubisco, and in the regeneration of
RuBP.143 In addition, other processes involving carbon
metabolism, such as starch and sucrose synthesis, photo-
respiration, or respiration, also determine net photoassimilate
production.144
Despite the potential of biotechnological approaches in
optimizing the photosynthetic process, the study of natural
genetic variation on photosynthesis within species and between
related species is a valuable undertaking.145 A survey performed
on 60 dierent accessions of C. sativus in the eld revealed
signicant dierences in the photosynthetic rate and in the
quantum eciency of photosystem II (unpublished data).
Research is currently being conducted to characterize the
sources of variation within the photosynthetic process and to
assess the relationship to biomass production in saron and
related Crocus species. This would provide insight into the
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selective forces that act on this trait and extend our under-
standing of the regulation of photosynthesis, ultimately
increasing the productivity of crops.
Several experimental manipulations suggest that photosyn-
thesis and sink utilization of carbohydrates are closely
coordinated.146 The regulation of carbon partitioning between
sources and sinks has a great impact on plant growth and yield
and is directly linked to the mechanisms of assimilate transport
and allocation. Depending on the main sugar transported and the
phloem loading/unloading mechanisms working, dierent
transporters can be involved (e.g., SWEET, H+/sucrose
symporters). The biotechnological manipulation of the transport
systems has been reported to positively alter carbon and also
nitrogen partitioning and consequently improve crop yields,
although pleiotropic eects limiting these gains can also occur.147
However, at this time no information is available about sugar
transport in Crocus.
The growth capacity of the sinks is a measure of its strength
and depends on sink size and activity. Sink size includes cell
number and cell size and is potentially regulated by hormones
(gibberellins and cytokinins) along with the supply of assimilates.
Bigger corms are formed when photoassimilate availability is
higher.19 Despite the genetic basis, corm size has been proposed
to be inuenced in Crocus by the content and type of soluble
sugar content at earlier stages of corm development.36 Sink
activity includes multiple components and key enzymes of
carbohydrate metabolism and storage, thus maintaining an
assimilate gradient and transport between source and sink.148
Sucrose is the main transported sugar in many species, and
several key enzymes are involved in carbon partitioning. Cell wall
and vacuole invertases (INV) in apoplastic unloaders have been
related to sink strength, although other activities such as sucrose
phosphate phosphatase (SPP), sucrose synthase (SS), or sucrose
phosphate synthase (SPS) may also be involved. In addition, nal
storage products, for example, starch, are the ultimate sinks, and
genes involved in their metabolism also inuence sink
strength.149
The integration of carbon assimilation and growth of organs is
highly regulated, and hormonal regulation of source-sink
relations has been well stated, although many aspects are not
fully understood.148,150 Another interesting approach would be
the use of transcription factors to alter the activity of several
genes involved in a given process. Several studies in recent years
point out this possibility to enhance photosynthesis and biomass
production.150
CONCLUSIONS
On the basis of this review, we conclude that the molecular and
physiological approaches to analyzing and addressing saron
production processes and the exploitation of their secondary
metabolism products remain largely unexplored. It is essential to
integrate crop physiology and genomic approaches to provide a
useful framework in which synergies can be explored between
technology and systems-oriented approaches. This review
provides a starting point for key aspects in saron biotechnology.
AUTHOR INFORMATION
Corresponding Author
*(L.G.-G.) E-mail: Marialourdes.gomez@uclm.es. Phone: +34
967599200.
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J. Agric. Food Chem. 2015, 63, 87518764
Journal of Agricultural and Food Chemistry
Funding
This work was supported by the Spanish Ministerio de Econom a
y Competitividad (BIO2013-44239-R).
Notes
The authors declare no competing nancial interest.
REFERENCES
(1) Tsoktouridis, G.; Krigas, N.; Karamplianis, T.; Constantinidis, T.;
Maloupa, E. In Genetic Dierences among Wild Greek Crocus Taxa and
Cultivated Saron (Crocus sativus L.), 3rd International Symposium on
Saron: Forthcoming Challenges in Cultivation Research and
Economics, Kozani, Greece; ISHS: Leuven, Belgium, 2009; p 37.
(2) CoskunK, F.; Selvi, S.; Satil, F. Phylogenetic relationships of some
Turkish Crocus (Iridaceae) taxa based on morphological and anatomical
characters. Turk J. Bot 2010, 34, 171178.
(3) Mathew, B. The Crocus a Revision of the Genus Crocus; Timber
Press: Portland, OR, USA, 1982.
(4) Seberg, O.; Petersen, G. How many loci does it take to DNA
barcode a crocus? PLoS One 2009, 4, e4598.
(5) Castillo, R.; Fernandez, J. A.; Gomez-Gomez, L. Implications of
carotenoid biosynthetic genes in apocarotenoid formation during the
stigma development of Crocus sativus and its closer relatives. Plant
Physiol. 2005, 139, 674689.
(6) Negbi, M. Saron cultivation: past, present and future prospects In
Saron. Crocus sativus L.; Negbi, M., Ed.; Harwood Academic Publishers:
Reading, UK, 1999; pp 118.
(7) Srivastava, R.; Ahmed, H.; Dixit, R. K.; Dharamveer; Saraf, S. A.
Crocus sativus L.: a comprehensive review. Pharmacogn. Rev. 2010, 4,
200208.
(8) Tarantilis, P. A.; Tsoupras, G.; Polissiou, M. Determination of
saffron (Crocus sativus L.) components in crude plant extract using high-
performance liquid chromatography-UV-visible photodiode-array
detection-mass spectrometry. J. Chromatogr A 1995, 699, 107118.
(9) Winterhalter, P.; Straubinger, M. Saffron-renewed interest in an
ancient spice. Food Rev. Int. 2000, 16, 3959.
(10) Bathaie, S. Z.; Mousavi, S. Z. New applications and mechanisms of
action of saffron and its important ingredients. Crit. Rev. Food Sci. Nutr.
2010, 50, 761786.
(11) Abdullaev, F. I. Biological effects of saffron. Biofactors 1993, 4,
8386.
(12) Abdullaev, F. I.; Espinosa-Aguirre, J. J. Biomedical properties of
saffron and its potential use in cancer therapy and chemoprevention
trials. Cancer Detect. Prev. 2004, 28, 426432.
(13) Feizzadeh, B.; Afshari, J. T.; Rakhshandeh, H.; Rahimi, A.; Brook,
A.; Doosti, H. Cytotoxic effect of saffron stigma aqueous extract on
human transitional cell carcinoma and mouse fibroblast. Urol. J. 2008, 5,
161167.
(14) Schmidt, M.; Betti, G.; Hensel, A. Saffron in phytotherapy:
pharmacology and clinical uses. Wien. Med. Wochenschr. 2007, 157,
315319.
(15) Howes, M. J.; Perry, E. The role of phytochemicals in the
treatment and prevention of dementia. Drugs Aging 2011, 28, 439468.
(16) Rossi, M.; Bermudez, L.; Carrari, F. Crop yield: challenges from a
metabolic perspective. Curr. Opin. Plant Biol. 2015, 25, 7989.
(17) Molina, R. V.; Valero, M.; Navarro, Y.; Guardiola, J. L.; Garca-
Luis, A. Temperature effects on flower formation in saffron (Crocus
sativus L.). Sci. Hortic. 2005, 103, 361379.
(18) Negb, M.; Dagan, B.; Dror, A.; Basker, D. Growth, flowering,
vegetative reproduction, and dormancy in the saffron crocus (Crocus-
sativus L). Isr. J. Bot. 1989, 38, 95113.
(19) Renau-Morata, B.; Nebauer, S. G.; Sanchez, M.; Molina, R. V.
Effect of corm size, water stress and cultivation conditions on
photosynthesis and biomass partitioning during the vegetative growth
of saffron (Crocus sativus L.). Ind. Crops Prod. 2012, 39, 4046.
(20) de Juan, J. A.; Corcoles, H. L.; Munoz, R. M.; Picornell, M. R.
Yield and yield components of saffron under different cropping systems.
Ind. Crops Prod. 2009, 30, 212219.
8761
Review
(21) Molina, R. V.; Valero, M.; Navarro, Y.; Garcia-Luis, A.; Guardiola,
J. L. Low temperature storage of corms extends the flowering season of
saffron (Crocus sativus L.). J. Hortic. Sci. Biotechnol. 2005, 80, 319326.
(22) Molina, R.; Valero, M.; Navarro, Y.; Garcia-Luis, A.; Guardiola, J.
L. The effect of time of corm lifting and duration of incubation at
inductive temperature on flowering in the saffron plant (Crocus sativus
L.). Sci. Hortic. 2004, 103, 7991.
(23) Milyaeva, E. L.; Azizbekova, N. S. Cytophysiological changes in
course of development of stem apices of saffron crocus. Soviet Plant
Physiol. 1978, 25, 227233.
(24) Koul, K. K.; Farooq, S. Growth and differentiation in the shoot
apical meristem of saffron plant (Crocus sativus L.). J. Indian Bot. Soc.
1982, 63, 153160.
(25) Plessner, O.; Negbi, M.; Ziv, M.; Basber, D. Effects of temperature
on the flowering of the saffron crocus (Crocus sativus L.): induction of
hysteranthy. Isr. J. Bot. 1989, 38, 7.
(26) Wilkins, H. F. Crocus vernus, Crocus sativus. In Handbook of
Flowering; Halevy, A. H., Ed.; CRC Press: Boca Raton, FL, USA, 1985;
Vol. 2, pp 350355.
(27) Azizbekova, N. S. H.; Milyaeva, E. L.; Lobova, N. V.; Chailakhyan,
M. K. H. Effects of gibberellin and kinetin on formation of flower organs
in saffron crocus. Fiziol. Rast 1978, 25, 6.
(28) Farooq, S.; Koul, K. K. Changes in gibberellin-like activity in
corms of saffron plant (Crocus sativus L.) during dormancy and
sprouting. Biochem. Physiol. Pflanz. 1983, 178, 685689.
(29) Tsaftaris, A.; Pasentsis, K.; Kalivas, A.; Michailidou, S.; Madesis,
P.; Argiriou, A. Isolation of a CENTRORADIALIS/TERMINAL
FLOWER1 homolog in saffron (Crocus sativus L.): characterization
and expression analysis. Mol. Biol. Rep. 2012, 39, 78997910.
(30) Tsaftaris, A.; Pasentsis, K.; Argiriou, A. Cloning and character-
ization of FLOWERING LOCUS T-like genes from the perennial
geophyte saffron crocus (Crocus sativus). Plant Mol. Biol. Rep. 2013, 31,
15581568.
(31) Tsaftaris, A.; Pasentsis, K.; Iliopoulos, I.; Polidoros, A. N. Isolation
of three homologous AP1-like MADS-box genes in crocus (Crocus
sativus L.) and characterization of their expression. Plant Sci. 2004, 166,
12351243.
(32) Tsaftaris, A. S.; Polidoros, A. N.; Pasentsis, K.; Kalivas, A. Cloning,
structural characterization, and phylogenetic analysis of flower MADS-
box genes from crocus (Crocus sativus L.). Sci. World J. 2007, 7, 1047
1062.
(33) Tsaftaris, A. S.; Pasentsis, K.; Polidoros, A. N. Isolation of a
differentially spliced C-type flower specific AG-like MADS-box gene
from crocus (Crocus sativus) and characterization of its expression. Biol.
Plant. 2005, 49, 499504.
(34) Tsaftaris, A.; Pasentsis, K.; Makris, A.; Darzentas, N.; Polidoros,
A.; Kalivas, A.; Argiriou, A. The study of the E-class SEPALLATA3-like
MADS-box genes in wild-type and mutant flowers of cultivated saffron
crocus (Crocus sativus L.) and its putative progenitors. J. Plant Physiol.
2011, 168, 16751684.
(35) Badri, M. A.; Minchin, P. E. H.; Lapointe, L. Effects of
temperature on the growth of spring ephemerals: Crocus vernus (L.) Hill.
Physiol. Plant. 2007, 130, 6776.
(36) Lundmark, M.; Vaughan, H.; Lapointe, L. Low temperature
maximizes growth of Crocus vernus (L.) Hill via changes in carbon
partitioning and corm development. J. Exp. Bot. 2009, 60, 22032213.
(37) Wingler, A.; Masclaux-Daubresse, C.; Fischer, A. M. Sugars,
senescence, and ageing in plants and heterotrophic organisms. J. Exp.
Bot. 2009, 60, 10631066.
(38) Lapointe, L. How phenology influences physiology in deciduous
forest spring ephemerals. Physiol. Plant. 2001, 113, 151157.
(39) Horvath, D. Common mechanisms regulate flowering and
dormancy. Plant Sci. 2009, 177, 523531.
(40) Feng, Y.; Zhu, L.; Pan, T.; Guo, Z.; Zhong, X.; Ding, A.; Pan, D.
Characterization of summer dormancy in Narcissus tazetta var. chinensis
and the role of NtFTs in summer dormancy and flower differentiation.
Sci. Hortic. 2015, 183, 109117.
DOI: 10.1021/acs.jafc.5b03194
J. Agric. Food Chem. 2015, 63, 87518764
Journal of Agricultural and Food Chemistry
(41) Horvath, D. P.; Anderson, J. V.; Chao, W. S.; Foley, M. E.
Knowing when to grow: signals regulating bud dormancy. Trends Plant
Sci. 2003, 8, 53450.
(42) Okubo, H. Dormancy. In Ornamental Geophytes: From Basic
Science to Sustainable Production;Okubo, R. K. a. H., Ed.;CRC Press:
Boca Raton, FL, USA, 2013; pp 233260.
(43) Marin, E.; Nussaume, L.; Quesada, A.; Gonneau, M.; Sotta, B.;
Hugueney, P.; Frey, A.; Marion-Poll, A. Molecular identification of
zeaxanthin epoxidase of Nicotiana plumbaginifolia, a gene involved in
abscisic acid biosynthesis and corresponding to the ABA locus of
Arabidopsis thaliana. EMBO J. 1996, 15, 23312342.
(44) Schwartz, S. H.; Tan, B. C.; Gage, D. A.; Zeevaart, J. A.; McCarty,
D. R. Specific oxidative cleavage of carotenoids by VP14 of maize. Science
1997, 276, 18721874.
(45) Seo, M.; Koiwai, H.; Akaba, S.; Komano, T.; Oritani, T.; Kamiya,
Y.; Koshiba, T. Abscisic aldehyde oxidase in leaves of Arabidopsis
thaliana. Plant J. 2000, 23, 481488.
(46) Ahrazem, O.; Rubio-Moraga, A.; Trapero, A.; Gomez-Gomez, L.
Developmental and stress regulation of gene expression for a 9-cis-
epoxycarotenoid dioxygenase, CstNCED, isolated from Crocus sativus
stigmas. J. Exp. Bot. 2012, 63, 681694.
(47) Rubio-Moraga, A.; Trapero, A.; Ahrazem, O.; Gomez-Gomez, L.
Crocins transport in Crocus sativus: the long road from a senescent
stigma to a newborn corm. Phytochemistry 2010, 71, 15061513.
(48) Cheng, X.; Ruyter-Spira, C.; Bouwmeester, H. The interaction
between strigolactones and other plant hormones in the regulation of
plant development. Front. Plant Sci. 2013, 4, 199.
(49) Rubio-Moraga, A.; Ahrazem, O.; Perez-Clemente, R. M.; Gomez-
Cadenas, A.; Yoneyama, K.; Lopez-Raez, J. A.; Molina, R. V.; Gomez-
Gomez, L. Apical dominance in saffron and the involvement of the
branching enzymes CCD7 and CCD8 in the control of bud sprouting.
BMC Plant Biol. 2014, 14, 171.
(50) Domagalska, M. A.; Leyser, O. Signal integration in the control of
shoot branching. Nat. Rev. Mol. Cell Biol. 2011, 12, 211221.
(51) Zhao, J.; Wang, T.; Wang, M.; Liu, Y.; Yuan, S.; Gao, Y.; Yin, L.;
Sun, W.; Peng, L.; Zhang, W.; Wan, J.; Li, X. Dwarf3 participates in an
SCF complex and associates with Dwarf14 to suppress rice shoot
branching. Plant Cell Physiol. 2014, 55, 1096.
(52) Waldie, T.; McCulloch, H.; Leyser, O. Strigolactones and the
control of plant development: lessons from shoot branching. Plant J.
2014, 79, 607.
(53) Jiang, L.; Liu, X.; Xiong, G.; Liu, H.; Chen, F.; Wang, L.; Meng, X.;
Liu, G.; Yu, H.; Yuan, Y.; Yi, W.; Zhao, L.; Ma, H.; He, Y.; Wu, Z.;
Melcher, K.; Qian, Q.; Xu, H. E.; Wang, Y.; Li, J. DWARF 53 acts as a
repressor of strigolactone signalling in rice. Nature 2013, 504, 401405.
(54) Rubio-Moraga, A.; Ahrazem, O.; Perez-Clemente, R. M.; Gomez-
Cadenas, A.; Yoneyama, K.; Lopez-Raez, J. A.; Molina, R. V.; Gomez-
Gomez, L. Apical dominance in saffron and the involvement of the
branching enzymes CCD7 and CCD8 in the control of bud sprouting.
BMC Plant Biol. 2014, 14, 171.
(55) Walter, M. H.; Floss, D. S.; Strack, D. Apocarotenoids: hormones,
mycorrhizal metabolites and aroma volatiles. Planta 2010, 232, 117.
(56) Moraga, A. R.; Nohales, P. F.; Perez, J. A.; Gomez-Gomez, L.
Glucosylation of the saffron apocarotenoid crocetin by a glucosyl-
transferase isolated from Crocus sativus stigmas. Planta 2004, 219, 955
966.
(57) Dufresne, C.; Cormier, F.; Dorion, S. In vitro formation of
crocetin glucosyl esters by Crocus sativus callus extract. Planta Med.
1997, 63, 150153.
(58) Kuhn, R.; Winterstein, A. U ber einen lichtempfindlichen Carotin-
Farbstoff aus Safran. Ber. Dtsch. Chem. Ges. B 1933, 66, 209214.
(59) Bouillon-Lagrange, E.-J. B. Analyse du Safran; 1811.
(60) Rubio Moraga, A.; Ahrazem, O.; Rambla, J. L.; Granell, A.; Gomez
Gomez, L. Crocins with high levels of sugar conjugation contribute to
the yellow colours of early-spring flowering crocus tepals. PLoS One
2013, 8, e71946.
(61) Liao, Y. H.; Houghton, P. J.; Hoult, J. R. Novel and known
constituents from Buddleja species and their activity against leukocyte
eicosanoid generation. J. Nat. Prod. 1999, 62, 12411245.
8762
Review
(62) Eugster, C. H.; Hurlimann, H.; Leuenberger, H. J. Crocetindial-
dehyd und crocetinhalbaldehyd als blutenfarbstoffe von Jacquinia
angustifolia. Helv. Chim. Acta 1969, 52, 806807.
(63) Tandon, J.; Katti, S.; Ruedi, P.; Eugster, C. Crocetin-dialdehyde
from Coleus forskohlii. Helv. Chim. Acta 1979, 62, 27062707.
(64) Gadgoli, C.; Shelke, S. Crocetin from the tubular calyx of
Nyctanthes arbor-tristis. Nat. Prod. Res. 2010, 24, 16101615.
(65) Priyadarshani, A. M. B.; Jansz, E.; Peiris, H. Studies on the
carotenoids of jakfruit (Artocarpus heterophyllus Lam.) from Matale and
Kurunegala districts. J. Natl. Sci. Found. Sri Lanka 2007, 35, 259262.
(66) Pfister, S.; Steck, A.; Pfander, H. Isolation and structure
elucidation of carotenoid glycoslyesters on gardenia fruits (Gardenia
jasminoides) and saffron (Crocus sativus). J. Agric. Food Chem. 1996, 44,
26122615.
(67) Juttner, F.; Hoflacher, B. Evidence of -carotene 7,8(7,8)
oxygenase (-cyclocitral, crocetindial generating) in Microcystis. Arch.
Microbiol. 1985, 141, 337343.
(68) Rubio, A.; Rambla, J. L.; Santaella, M.; Gomez, M. D.; Orzaez, D.;
Granell, A.; Gomez-Gomez, L. Cytosolic and plastoglobule-targeted
carotenoid dioxygenases from Crocus sativus are both involved in -
ionone release. J. Biol. Chem. 2008, 283, 2481624825.
(69) Pfander, H.; Schurtenberger, H. Biosynthesis of C20-carotenoids
in Crocus sativus. Phytochemistry 1982, 21, 10391042.
(70) Bouvier, F.; Suire, C.; Mutterer, J.; Camara, B. Oxidative
remodeling of chromoplast carotenoids: identification of the carotenoid
dioxygenase CsCCD and CsZCD genes involved in Crocus secondary
metabolite biogenesis. Plant Cell 2003, 15, 4762.
(71) Ahrazem, O.; Trapero, A.; Gomez, M. D.; Rubio-Moraga, A.;
Gomez-Gomez, L. Genomic analysis and gene structure of the plant
carotenoid dioxygenase 4 family: a deeper study in Crocus sativus and its
allies. Genomics 2010, 96, 239250.
(72) Rubio-Moraga, A.; Rambla, J. L.; Fernandez-de-Carmen, A.;
Trapero-Mozos, A.; Ahrazem, O.; Orzaez, D.; Granell, A.; Gomez-
Gomez, L. New target carotenoids for CCD4 enzymes are revealed with
the characterization of a novel stress-induced carotenoid cleavage
dioxygenase gene from Crocus sativus. Plant Mol. Biol. 2014, 86, 555
569.
(73) Moraga, A. R.; Rambla, J. L.; Ahrazem, O.; Granell, A.; Gomez-
Gomez, L. Metabolite and target transcript analyses during Crocus
sativus stigma development. Phytochemistry 2009, 70, 10091016.
(74) Kloer, D. P.; Schulz, G. E. Structural and biological aspects of
carotenoid cleavage. Cell. Mol. Life Sci. 2006, 63, 22912303.
(75) Sergeant, M. J.; Li, J. J.; Fox, C.; Brookbank, N.; Rea, D.; Bugg, T.
D.; Thompson, A. J. Selective inhibition of carotenoid cleavage
dioxygenases: phenotypic effects on shoot branching. J. Biol. Chem.
2009, 284, 52575264.
(76) DAgostino, N.; Pizzichini, D.; Chiusano, M. L.; Giuliano, G. An
EST database from saffron stigmas. BMC Plant Biol. 2007, 7, 53.
(77) Frusciante, S.; Diretto, G.; Bruno, M.; Ferrante, P.; Pietrella, M.;
Prado-Cabrero, A.; Rubio-Moraga, A.; Beyer, P.; Gomez-Gomez, L.; Al-
Babili, S.; Giuliano, G. Novel carotenoid cleavage dioxygenase catalyzes
the first dedicated step in saffron crocin biosynthesis. Proc. Natl. Acad.
Sci. U. S. A. 2014, 111, 1224612251.
(78) Ahrazem, O.; Rubio-Moraga, A.; Berman, J.; Capell, T.; Christou,
P.; Zhu, C.; Gomez-Gomez, L. The carotenoid cleavage dioxygenase
CCD2 catalyzing the synthesis of crocetin in spring crocuses and saffron
is a plastidial enzyme. New Phytol. 2015, 13609.
(79) Rios, J. L.; Recio, M. C.; Ginger, R. M.; Manz, S. An update review
of saffron and its active constituents. Phytother. Res. 1996, 10, 189.
(80) Tseng, T. H.; Chu, C. Y.; Huang, J. M.; Shiow, S. J.; Wang, C. J.
Crocetin protects against oxidative damage in rat primary hepatocytes.
Cancer Lett. 1995, 97, 6167.
(81) Kanakis, C. D.; Tarantilis, P. A.; Tajmir-Riahi, H. A.; Polissiou, M.
G. Crocetin, dimethylcrocetin, and safranal bind human serum albumin:
stability and antioxidative properties. J. Agric. Food Chem. 2007, 55,
970977.
(82) Kazi, H. A.; Qian, Z. Crocetin reduces TNBS-induced
experimental colitis in mice by downregulation of NFkB. Saudi J.
Gastroenterol. 2009, 15, 181187.
DOI: 10.1021/acs.jafc.5b03194
J. Agric. Food Chem. 2015, 63, 87518764
Journal of Agricultural and Food Chemistry
(83) Zheng, S.; Qian, Z.; Sheng, L.; Wen, N. Crocetin attenuates
atherosclerosis in hyperlipidemic rabbits through inhibition of LDL
oxidation. J. Cardiovasc. Pharmacol. 2006, 47, 7076.
(84) Sheng, L.; Qian, Z.; Shi, Y.; Yang, L.; Xi, L.; Zhao, B.; Xu, X.; Ji, H.
Crocetin improves the insulin resistance induced by high-fat diet in rats.
Br. J. Pharmacol. 2008, 154, 10161024.
(85) Mizuma, H.; Tanaka, M.; Nozaki, S.; Mizuno, K.; Tahara, T.;
Ataka, S.; Sugino, T.; Shirai, T.; Kajimoto, Y.; Kuratsune, H.; Kajimoto,
O.; Watanabe, Y. Daily oral administration of crocetin attenuates
physical fatigue in human subjects. Nutr. Res. (N. Y., NY, U. S.) 2009, 29,
145150.
(86) Kuratsune, H.; Umigai, N.; Takeno, R.; Kajimoto, Y.; Nakano, T.
Effect of crocetin from Gardenia jasminoides Ellis on sleep: a pilot study.
Phytomedicine 2010, 17, 840843.
(87) Gutheil, W. G.; Reed, G.; Ray, A.; Anant, S.; Dhar, A. Crocetin: an
agent derived from saffron for prevention and therapy for cancer. Curr.
Pharm. Biotechnol. 2012, 13, 173179.
(88) Ahmad, A. S.; Ansari, M. A.; Ahmad, M.; Saleem, S.; Yousuf, S.;
Hoda, M. N.; Islam, F. Neuroprotection by crocetin in a hemi-
parkinsonian rat model. Pharmacol., Biochem. Behav. 2005, 81, 805813.
(89) Nam, K. N.; Park, Y. M.; Jung, H. J.; Lee, J. Y.; Min, B. D.; Park, S.
U.; Jung, W. S.; Cho, K. H.; Park, J. H.; Kang, I.; Hong, J. W.; Lee, E. H.
Anti-inflammatory effects of crocin and crocetin in rat brain microglial
cells. Eur. J. Pharmacol. 2010, 648, 110116.
(90) Papandreou, M. A.; Kanakis, C. D.; Polissiou, M. G.;
Efthimiopoulos, S.; Cordopatis, P.; Margarity, M.; Lamari, F. N.
Inhibitory activity on amyloid-beta aggregation and antioxidant
properties of Crocus sativus stigmas extract and its crocin constituents.
J. Agric. Food Chem. 2006, 54, 87628768.
(91) Akhondzadeh, S.; Sabet, M. S.; Harirchian, M. H.; Togha, M.;
Cheraghmakani, H.; Razeghi, S.; Hejazi, S.; Yousefi, M. H.; Alimardani,
R.; Jamshidi, A.; Zare, F.; Moradi, A. Saffron in the treatment of patients
with mild to moderate Alzheimers disease: a 16-week, randomized and
placebo-controlled trial. J. Clin. Pharm. Ther. 2010, 35, 581588.
(92) Ochiai, T.; Shimeno, H.; Mishima, K.; Iwasaki, K.; Fujiwara, M.;
Tanaka, H.; Shoyama, Y.; Toda, A.; Eyanagi, R.; Soeda, S. Protective
effects of carotenoids from saffron on neuronal injury in vitro and in
vivo. Biochim. Biophys. Acta, Gen. Subj. 2007, 1770, 578584.
(93) Georgiadou, G.; Tarantilis, P. A.; Pitsikas, N. Effects of the active
constituents of Crocus sativus L., crocins, in an animal model of
obsessive-compulsive disorder. Neurosci. Lett. 2012, 528, 2730.
(94) Hosseinzadeh, H.; Sadeghnia, H. R. Safranal, a constituent of
Crocus sativus (saffron), attenuated cerebral ischemia induced oxidative
damage in rat hippocampus. J. Pharm. Pharm. Sci. 2005, 8, 394399.
(95) Fernandez-Sanchez, L.; Lax, P.; Esquiva, G.; Martn-Nieto, J.;
Pinilla, I.; Cuenca, N. Safranal, a saffron constituent, attenuates retinal
degeneration in P23H rats. PLoS One 2012, 7, e43074.
(96) Hooshmandi, Z.; Rohani, A. H.; Eidi, A.; Fatahi, Z.; Golmanesh,
L.; Sahraei, H. Reduction of metabolic and behavioral signs of acute
stress in male Wistar rats by saffron water extract and its constituent
safranal. Pharm. Biol. 2011, 49, 947.
(97) Winkel-Shirley, B. Flavonoid biosynthesis. A colorful model for
genetics, biochemistry, cell biology, and biotechnology. Plant Physiol.
2001, 126, 485493.
(98) Bowles, D.; Lim, E. K.; Poppenberger, B.; Vaistij, F. E.
Glycosyltransferases of lipophilic small molecules. Annu. Rev. Plant
Biol. 2006, 57, 567597.
(99) Moraga, A. R.; Mozos, A. T.; Ahrazem, O.; Gomez-Gomez, L.
Cloning and characterization of a glucosyltransferase from Crocus sativus
stigmas involved in flavonoid glucosylation. BMC Plant Biol. 2009, 9,
109.
(100) Li, C. Y.; Wu, T. S. Constituents of the pollen of Crocus sativus L.
and their tyrosinase inhibitory activity. Chem. Pharm. Bull. 2002, 50,
13051309.
(101) Termentzi, A.; Kokkalou, E. LC-DAD-MS (ESI+) analysis and
antioxidant capacity of crocus sativus petal extracts. Planta Med. 2008,
74, 573581.
8763
Review
(102) Nrbk, R.; Brandt, K.; Kvist Nielsen, J.; rgaard, M.; Jacobsen,
N. Flower pigment composition of Crocus species and cultivars used for
a chemotaxonomic investigation. Biochem. Syst. Ecol. 2002, 30, 763.
(103) Trapero-Mozos, A.; Ahrazem, O.; Rubio-Moraga, A.; Jimeno, M.
L.; Gomez, M. D.; Gomez-Gomez, L. Characterization of a
glucosyltransferase enzyme involved in the formation of kaempferol
and quercetin sophorosides in Crocus sativus. Plant Physiol. 2012, 159,
1335.
(104) Ahrazem, O.; Rubio-Moraga, A.; Mozos, A. T.; Gomez-Gomez,
M. L. Genomic organization of a UDP-glucosyltransferase gene
determines differential accumulation of specific flavonoid glucosides
in tepals. Plant Cell, Tissue Organ Cult. 2014, 119, 227.
(105) Buer, C. S.; Imin, N.; Djordjevic, M. A. Flavonoids: new roles for
old molecules. J. Integr. Plant Biol. 2010, 52, 98111.
(106) Williams, R. J.; Spencer, J. P.; Rice-Evans, C. Flavonoids:
antioxidants or signalling molecules? Free Radical Biol. Med. 2004, 36,
838849.
(107) Gutierrez-Merino, C.; Lopez-Sanchez, C.; Lagoa, R.; Samhan-
Arias, A. K.; Bueno, C.; Garcia-Martinez, V. Neuroprotective actions of
flavonoids. Curr. Med. Chem. 2011, 18, 11951212.
(108) Rendeiro, C.; Guerreiro, J. D.; Williams, C. M.; Spencer, J. P.
Flavonoids as modulators of memory and learning: molecular
interactions resulting in behavioural effects. Proc. Nutr. Soc. 2012, 71,
246262.
(109) Mulvihill, E. E.; Huff, M. W. Antiatherogenic properties of
flavonoids: implications for cardiovascular health. Can. J. Cardiol. 2010,
26 (Suppl.A), 17A21A.
(110) Palanichamy, S.; Nagarajan, S. Analgesic activity of Cassia alata
leaf extract and kaempferol 3-O-sophoroside. J. Ethnopharmacol. 1990,
29, 7378.
(111) Kim, T. H.; Ku, S. K.; Bae, J. S. Inhibitory effects of kaempferol-3-
O-sophoroside on HMGB1-mediated proinflammatory responses. Food
Chem. Toxicol. 2012, 50, 11181123.
(112) Kim, T. H.; Ku, S. K.; Lee, I. C.; Bae, J. S. Anti-inflammatory
effects of kaempferol-3-O-sophoroside in human endothelial cells.
Inflamm. Res. 2012, 61, 217224.
(113) Sparg, S. G.; Light, M. E.; van Staden, J. Biological activities and
distribution of plant saponins. J. Ethnopharmacol. 2004, 94, 219243.
(114) Haralampidis, K.; Trojanowska, M.; Osbourn, A. E. Biosynthesis
of triterpenoid saponins in plants. Adv. Biochem. Eng. Biotechnol 2002,
75, 3149.
(115) Voigt, G.; Hiller, K. Advances in the chemistry and biology of the
steroid saponins. Sci. Pharm. 1987, 55, 201222.
(116) Vincken, J. P.; Heng, L.; de Groot, A.; Gruppen, H. Saponins,
classification and occurrence in the plant kingdom. Phytochemistry 2007,
68, 275297.
(117) Xu, R.; Fazio, G. C.; Matsuda, S. P. On the origins of triterpenoid
skeletal diversity. Phytochemistry 2004, 65, 261291.
(118) Augustin, J. M.; Kuzina, V.; Andersen, S. B.; Bak, S. Molecular
activities, biosynthesis and evolution of triterpenoid saponins.
Phytochemistry 2011, 72, 435457.
(119) Rubio-Moraga, A.; Gerwig, G. J.; Castro-Daz, N.; Jimeno, M. L.;
Escribano, J.; Fernandez, J. A.; Kamerling, J. P. Triterpenoid saponins
from corms of Crocus sativus: localization, extraction and character-
ization. Ind. Crops Prod. 2011, 34, 1401.
(120) Rubio-Moraga, A.; Gomez-Gomez, L.; Trapero, A.; Castro-Daz,
N.; Ahrazem, O. Saffron corm as a natural source of fungicides: the role
of saponins in the underground. Ind. Crops Prod. 2013, 49, 915.
(121) Adams, M. M.; Damani, P.; Perl, N. R.; Won, A.; Hong, F.;
Livingston, P. O.; Ragupathi, G.; Gin, D. Y. Design and synthesis of
potent Quillaja saponin vaccine adjuvants. J. Am. Chem. Soc. 2010, 132,
19391945.
(122) Castro-Diaz, N.; Salaun, B.; Perret, R.; Sierro, S.; Romero, J. F.;
Fernandez, J. A.; Rubio-Moraga, A.; Romero, P. Saponins from the
Spanish saffron Crocus sativus are efficient adjuvants for protein-based
vaccines. Vaccine 2012, 30, 388397.
(123) Gu cl u -U stu ndag, O .; Mazza, G. Saponins: properties,
applications and processing. Crit. Rev. Food Sci. Nutr. 2007, 47, 231.
DOI: 10.1021/acs.jafc.5b03194
J. Agric. Food Chem. 2015, 63, 87518764
 Journal of Agricultural and Food Chemistry Review

(124) Grilli-Caiola, M. G.; Caputo, P.; Zanier, R. RAPD analysis in (146) Ainsworth, E. A.; Yendrek, C. R.; Skoneczka, J. A.; Long, S. P.
Crocus sativus L. accessions and related Crocus species. Biol. Plant. 2004, Accelerating yield potential in soybean: potential targets for
48, 375. biotechnological improvement. Plant, Cell Environ. 2012, 35, 3852.
(125) Rubio-Moraga, A.; Castillo-Lopez, R.; Gomez-Gomez, L.; (147) Yadav, U. P.; Ayre, B. G.; Bush, D. R. Transgenic approaches to
Ahrazem, O. Saffron is a monomorphic species as revealed by RAPD, altering carbon and nitrogen partitioning in whole plants: assessing the
ISSR and microsatellite analyses. BMC Res. Notes 2009, 2, 189. potential to improve crop yields and nutritional quality. Front. Plant Sci.
(126) Nemati, Z.; Mohsen, M.; Majidian, P.; Zeinalabedini, M.; 2015, 6, 275.
Pirseyedi, S. M.; Bahadori, M. Saffron (Crocus sativus L.), a (148) Albacete, A. A.; Martinez-Andujar, C.; Perez-Alfocea, F.
monomorphic or polymorphic species? Span. J. Agric. Res. 2014, 12, 753. Hormonal and metabolic regulation of source-sink relations under
(127) Agayev, Y.; Shakib, A.; Soheilivand, S.; Fathi, M. In Breeding of salinity and drought: from plant survival to crop yield stability.
Saron (Crocus sativus): Possibilities and Problems, II International Biotechnol. Adv. 2014, 32, 1230.
(149) Bihmidine, S.; Hunter, C. T., 3rd; Johns, C. E.; Koch, K. E.;
Symposium on Saron Biology and Technology 739; ISHS: Leuven,
Braun, D. M. Regulation of assimilate import into sink organs: update on
Belgium, 2006; pp 203207.10.17660/ActaHortic.2007.739.25
molecular drivers of sink strength. Front. Plant Sci. 2013, 4, 177.
(128) Mir, J.; Ahmed, N.; Singh, D.; Khan, M.; Zaffer, S.; Shafi, W.
(150) Saibo, N. J.; Lourenco, T.; Oliveira, M. M. Transcription factors
Breeding and biotechnological opportunities in saffron crop improve- and regulation of photosynthetic and related metabolism under
ment. Afr. J. Agric. Res. 2015, 10, 970974. environmental stresses. Ann. Bot. 2009, 103, 609623.
(129) Khan, M. A.; Nagoo, S.; Naseer, S.; Nehvi, F. A.; Zargar, S. M.
Induced mutation as a tool for improving corm multiplication in saffron
(Crocus sativus L.). J. Phytol. 2011, 3, 810.
(130) Dar, S.; Makhdoomi, M.; Allie, B.; Mir, Z.; Nehvi, F.; Wani, S. In
Biological Interventions for Enhancing Saron Productivity in Kashmir, II
International Symposium on Saron Biology and Technology 739;
ISHS: Leuven, Belgium, 2006; pp 253210.17660/Acta-
Hortic.2007.739.1.
(131) Grilli-Caiola, M.; Canini, A. Saffron reproductive biology. Acta
Hortic. 2004, 650, 2537.
(132) Souret, F.; Weathers, P. Crocus sativus L. (saffron): cultivation, in
vitro culture, secondary metabolite production and phytopharmacog-
nosy. J. Herbs, Spices Med. Plants 2000, 6, 99116.
(133) Agayev, Y. M. New features in karyotype structure and origin of
saffron, Crocus sativus L. Cytologia 2002, 67, 245252.
(134) Grilli-Caiola, M.; Canini, A. Looking for sarons (Crocus sativus
L.) parents. In Functional Plant Science and Biotechnology, Saron Special
Issue; Husaini, A. M., Ed.; Global Science Books: London, UK, 2010;
Vol. 4, pp 114.
(135) Grilli-Caiola, M.; Leonardi, D.; Canini, A. Seed structure in
Crocus sativus L. , C. cartwrightianus Herb., C. thomasii Ten., and C.
hadriaticus Herb. at SEM. Plant Syst. Evol. 2010, 285, 111120.
(136) Abourashed, E. A. Bioavailability of plant-derived antioxidants.
Antioxidants 2013, 2, 309325.
(137) Li, X. F.; Jia, L. Y.; Xu, J.; Deng, X. J.; Wang, Y.; Zhang, W.;
Zhang, X. P.; Fang, Q.; Zhang, D. M.; Sun, Y.; Xu, L. FT-like NFT1 gene
may play a role in flower transition induced by heat accumulation in
Narcissus tazetta var. chinensis. Plant Cell Physiol. 2013, 54, 270281.
(138) Jensen, C. S.; Salchert, K.; Nielsen, K. K. A TERMINAL
FLOWER1-like gene from perennial ryegrass involved in floral
transition and axillary meristem identity. Plant Physiol. 2001, 125,
15171528.
(139) Eriksson, S.; Bohlenius, H.; Moritz, T.; Nilsson, O. GA4 is the
active gibberellin in the regulation of LEAFY transcription and
Arabidopsis floral initiation. Plant Cell 2006, 18, 21722181.
(140) Leeggangers, H. A.; Moreno-Pachon, N.; Gude, H.; Immink, R.
G. Transfer of knowledge about flowering and vegetative propagation
from model species to bulbous plants. Int. J. Dev. Biol. 2013, 57, 611
620.
(141) Doebley, J.; Stec, A.; Hubbard, L. The evolution of apical
dominance in maize. Nature 1997, 386, 4858.
(142) Aguilar-Martinez, J. A.; Poza-Carrion, C.; Cubas, P. Arabidopsis
BRANCHED1 acts as an integrator of branching signals within axillary
buds. Plant Cell 2007, 19, 458472.
(143) Long, S. P.; Marshall-Colon, A.; Zhu, X. G. Meeting the global
food demand of the future by engineering crop photosynthesis and yield
potential. Cell 2015, 161, 5666.
(144) Zhu, X. G.; Long, S. P.; Ort, D. R. Improving photosynthetic
efficiency for greater yield. Annu. Rev. Plant Biol. 2010, 61, 235261.
(145) Flood, P. J.; Harbinson, J.; Aarts, M. G. Natural genetic variation
in plant photosynthesis. Trends Plant Sci. 2011, 16, 327335.

8764 DOI: 10.1021/acs.jafc.5b03194

J. Agric. Food Chem. 2015, 63, 87518764

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