UNIVERSITY OF CATANIA
Department of Agriculture, Food and Environment
Master of Science
Agricultural Science and Technology
EFFECTS OF THERMAL AND LIGHT CONDITIONS ON
EDIBLE SPROUTS YIELD AND FUNCTIONAL TRAITS OF
THREE VEGETABLE SPECIES
Federico Incardona
Supervisor: Prof. Rosario Mauro
______________________________________________________________________
Accademic Year 2021-2022
1
SOMMARIO
1.
INTRODUCTION .................................................................................................. 5
2.
NEW FRESH CONSUPTION VEGETABLE CATEGORIES: SPROUTS,
MICROGREENS AND BABY-LEAF .......................................................................... 7
2.1
3.
Edible sprouts: main characteristics .................................................................. 7
2.1.1
Most exploited species and neglected species .......................................... 10
2.1.2
Sprouts production.................................................................................... 13
2.2
A look on the other micro-scale vegetables: microgreens and baby-leaf ....... 15
2.3
Historical background of edible sprouts .......................................................... 16
NUTRITIONAL AND NUTRACEUTICAL PROPRIETIES OF EDIBLE
SPROUTS ...................................................................................................................... 18
3.1
Germination process and variation of seed chemical content .......................... 21
3.1.1
Content of structural carbohydrates .......................................................... 23
3.1.2
Content of non-structural carbohydrates .................................................. 24
3.1.3
Proteins content ........................................................................................ 24
3.1.4
Lipids content ........................................................................................... 26
3.1.5
Minerals content ....................................................................................... 27
3.1.6
Antioxidants content ................................................................................. 27
3.2 Antinutrients and allergens ................................................................................... 28
3.3 Quality sprouts and food safeness ........................................................................ 30
4. BIOLOGICAL TRAITS OF EDIBLE SPROUTS ................................................ 32
4.1 Leek (Allium porrum L.) ...................................................................................... 32
2
4.2 Fennel (Foeniculum vulgare Miller var. azoricum) ............................................. 33
4.3 Cucumber (Cucumis sativus L.) ........................................................................... 34
5. DIFFERENT USE TYPOLOGIES FOR SPROUTS ............................................ 36
6. AIM OF THE THESIS ............................................................................................ 39
7. MATERIALS AND METHODS ............................................................................. 42
7.1 Plant materials, seeds germination, light and thermal conditions ........................ 42
7.2 Determination of the fresh weight and dry matter content ................................... 43
7.3 Sprouts growth parameters ................................................................................... 43
7.4 Determination of the antioxidant capacity (DPPH) .............................................. 44
7.5 Determination of Total Carotenoids Concentration (TCC) .................................. 45
7.6 Determination of the Total Polyphenol Content (TPC) ....................................... 45
7.7 Statistical procedures ............................................................................................ 46
8. RESULTS .................................................................................................................. 47
8.1
Sprouts Biomorphometric Characteristics ....................................................... 47
8.1.1
Leek .......................................................................................................... 47
8.1.2
Fennel ....................................................................................................... 48
8.1.3
Cucumber ................................................................................................. 48
8.2
Sprouts Biochemical Characteristics ............................................................... 49
8.2.1
Leek .......................................................................................................... 49
8.2.2
Fennel ....................................................................................................... 50
8.2.3
Cucumber ................................................................................................. 51
9. CONCLUSIONS ....................................................................................................... 52
3
TABLES ........................................................................................................................ 53
REFERENCES ............................................................................................................. 59
4
1. INTRODUCTION
The consumption and production of edible sprouts is increasing across the world. This
trend is explained by the high functional traits of edible sprouts in terms of health benefits,
ease to grow them and the low costs which are involved in the process of growing sprouts
(Marra et all., 2000). The entire sprouts growing cycle is accomplished in few days and
it is possible to carry on continuous growing cycle to produce a fresh edible product
almost for all the year. From a biological point of view, sprouts represent the very first
stage of a plant growth, which starts straight from the seed germination, producing a stem
and cotyledons, the edible part. The edible part includes the germinated seeds and the root
system as well (Renna, M., et al., 2017). Due to the early stage of growth, edible sprouts
hold a high quantity and quality of available nutritional and nutraceutical elements and as
people are becoming more conscious of the need to maintain a healthy style of living, the
interest of consumers is going towards products which allow to make nutrition functional
(Di Gioia, F., et al., 2015). The ready-to-use properties and the tasteful of edible sprouts
meet these needs. Recently, there has been interest in food demand more oriented towards
diets rich in fruits and vegetables, characterized by a high content of bioactive molecules
and, besides sprouts, other new fresh vegetable products such as microgreens and babyleaf are becoming popular among consumers (Galieni, A., et al., 2020). For each of these
products several ambiguous definitions occur, lead often, to misunderstanding and
deprive specialists from the specific terminology, so it seems helpful to understand the
differences between these new fresh vegetable categories. Nevertheless, sprouts and
microgreens are considered innovative culinary ingredients. They are used as additions
to sandwiches, salads, soups, desserts and drinks, and their popularity is due to their
delicate texture, unique colours, and high palatability, making them useful in the culinary
industry (Renna, M., et al., 2017). Edible sprouts include products from many different
5
vegetable species, such as alfalfa (Medicago sativa L.), legumes, cabbage, broccoli and
others Brassicaceae species or buckwheat (Fagopyrum esculentum), oats (Avena sativa
L.) and other Graminaceae species. In the recent time, even some other horticultural
vegetable species such as watermelon, cucumber, carrot, chard, lettuce, raddish, onion,
leek and Alliaceae species, has been used to produce sprouts from germinated grains
(Ebert, A.W., 2022).
6
2. NEW FRESH CONSUPTION VEGETABLE
CATEGORIES: SPROUTS, MICROGREENS AND
BABY-LEAF
Today, the different branches of economics are changing, especially after the outbreak of
COVID-19 pandemic, and even the agricultural sector is not left behind this kind of
switch. The food production system must work to face new challenges for the next years,
such as climate change, the massive growth of the human population and the need to
develop a sustainable industry of food production. Therefore, the goal, is also to develop
new categories of fresh vegetable which are easy to produce, have a low carbon footprint
and at the same time guarantee the presence of good compounds for human health. In this
scenario, the production of edible sprouts fits in all this goals. With society’s growing
interest in healthy eating and lifestyles, e.g., the Slow Food movement and the promotion
of novel and superfoods, the interest in fresh, ready-to-eat
functional and nutraceutical food has been on the rise in recent decades. In this context,
microscale vegetables like sprouted seeds and microgreens, are becoming increasingly
popular worldwide as fresh, ready-to-eat functional and nutraceutical food.
They have great potential to diversify and enhance the human diet and address nutrient
deficiencies due to their high content of phytochemicals (Galieni, A., et al., 2020).
2.1 Edible sprouts: main characteristics
According to the definition which is given by the American Association of Cereal
Chemists (AACC), ‘’Sprouts are the products obtained from the germination of seeds and
their development in water or another medium, harvested before the development of the
first true leaves and which is intended to be eaten whole, including the seed’’ (AACC
7
International, 2000). This is also the definition which is given by the European
Commission regulation (No. 208/2013). Sprouts are commonly grown in the dark, under
high relative humidity. They are harvested when the cotyledons are still under-developed
and true leaves have not begun to emerge, usually after 3–5 days from seed hydration. So
far, sprouts have been more often than microgreens recognized as a wellness and healthpromoting foods, widely recommended by dietitians due to their high content of nutrients
and bioactive compounds, such as flavonoids, hydroxycinnamic acids, vitamins and
glucosinolates, minerals, and carotenoids (Kyriacou, M.C., et al., 2017).
There are some important agronomic aspects which must be considered for commercial
scale producer in order to accomplish a successful sprouts production cycle: the key factor
is to choose those species which are strictly related to seeds availability and quality in
terms of homogeneous germination rate and in term of sanitary safeness. Seeds shouldn’t
have special needs in terms of germination conditions, like temperature or dormancy, in
order to achieve multiple growing cycle across all the year. Moreover, it is important to
guarantee the availability of low-cost seeds, and another critical aspect is related to the
shelf life of germinated sprouts (Di Gioia, F., et al., 2015).
The production process starts with germination, which follow the soaking phase of the
seeds. Germination is a complex stage of plant ontogenesis, involving growth initiation
but not comprising final growth processes and maturation (Baenas, et al., 2017).
Germinating seeds could contain from two to ten times more phytochemicals as compared
with commercial adult plants. This content depends on the species, cultivar,
environmental conditions and the time of germination, storage, and processing (Choe, U.,
et al., 2018).
According to the used species, sprouts, can be harvested from 7 to 25-28 days after
germination, when the cotyledonal leaves are completely formed (Di Gioia, F., et al.,
8
2015) and the edible part of sprouts consists of the stem, the cotyledonal leaves, and
sometimes, even the early formed true leaves. The teguments of the germinated seeds are
an edible part of the sprouts too, especially if they are tiny, soft, and it is not uncommon
to find them still attached to the stem. The cultivated species which are used for sprouts
production, but also for micro-greens, can be collected in four different groups, which
are: legumes, cereals or pseudo-cereal like buckwheat, oil crops and horticultural species
(Cacciola, G., 2014). Despite the diversity, almost all the market is interested on legumes
sprouts. In the sprout’s world, some terms like ‘’soy sprouts’’ are often used in a wrong
way, generically referring to all the sprouts, even if they come from other species which
aren’t soy. Indeed, also sprouts from Green or Red Mung, Green Azuki and yellow soy,
have in common the ‘’soy sprouts’’ appellative. Only yellow soy can be considered as a
Soy sprout, but it’s not uncommon to find product on the market which are called ‘’soy
sprouts’’ even if they don’t come from soy. This kind of confusion is due to the low levels
of knowledge, in addition to the translation difficulty, which gives many problems,
considering that legumes are species which came from the Oriental part of the world,
where the different cultures and populations provide different terms or words to identify
soy plant (Cacciola, G., 2014).
9
Table 1 - Suitable species for the production of sprouts (Source: La Malfa e Bianco, 2006; Cacciola, 2014; USDA,
2016).
Sprouts cultivated species
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
Garlic (Allium sativum L.),
Dill (Anethum graveolens L.),
Peanut (Arachis hypogaea L.),
Red Azuki (Vigna angularis L.),
Green Azuki (Vigna radiata L.),
Basil (Ocinum basilicum L.),
Chard (Beta vulgaris L.),
Carrot (Daucus carota L.),
Cauliflower (Brassica oleracea L.
Gruppo botrytis),
Cavolo broccolo (Brassica
oleracea L. Gruppo italica),
Cavolo cappuccio (Brassica
oleracea L. Gruppo capitata),
Cavolo da foglia (Brassica
oleracea L. gruppo acephala),
Brussels Sprouts (Brassica
oleracea L. Gruppo gemmifera),
Chickpea (Cicer arietinum L.),
Chicory (Cichorium intybus L.),
Turnip Top (Brassica oleracea L.
Gruppo broccoletto),
Onion (Allium cepa L.),
Rapeseed(Brassica napus L.),
Watercress (Lepidium sativum L.),
Alfalfa (Medicago sativa L.),
Green bean (Phaseolus vulgaris
L.),
Cowpea (Vigna unguiculata [L.].
Walp. subsp. unguiculata L.
(Walp.),
Emmer (Triticum dicoccon L.),
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
•
Broad bean (Vicia faba L.),
Fenugreek (Trigonella foenumgraecum L.),
Fennel (Foeniculum vulagare
Mill.),
Buckwheat (Polygonum
fagopyrum L.),
Wheat (Triticum ssp.),
Sunflower (Helianthus annuus
L.),
Endive (Cichorium endivia L.),
Lattuce (Lactuca sativa L.),
Lentis (Lens culinaris Medic.),
Linen (Linum usitatissimum L.),
Millet (Panicum miliaceum L.),
Barley (Hordeum vulgare L.),
Pea (Pisum sativum L.),
Leek (Allium porrum L.),
Quinoa (Chenopodium quinoa
Willd.),
Red Raddish (Cichorium intybus
L. Gruppo rubifolium),
Turnip (Brassica rapa L.
Gruppo rapa),
Radish (Raphanus sativus L.),
Rice Oryza sativa L.),
Rucola (Eruca vesicaria L.
Cav.),
Sesame (Sesamum indicum L.),
Soy (Glycine max L.),
Spinach (Spinacia oleracea L.),
Clover (Trifolium ssp.),
Pumpkin (Cucurbita moshata
Duch.).
2.1.1 Most exploited species and neglected species
Plant species in the Poaceae, Brassicaceae, and Fabaceae families are the most exploited
for sprouting purposes and therefore the most reported by the scientific literature. For
these species, research has provided a lot of information on the nutritional traits of sprouts
10
and microgreens. Other cultivated species are used for sprouting, being appreciated for
several peculiar traits: vivid colours (i.e., red for red basil; green for spinach), intense
smells (i.e., aromatic herbs), pleasant textures (i.e., regular for Asteraceae; juicy for
sunflower and beet; crunchy for celery), and variable tastes (i.e., regular for Asteraceae;
slightly sour for beet; bitter for Cucurbitaceae) (Di Gioia, F., et al., 2017) (Renna, M., et
al., 2017). In this regard, the definition and quantification of organoleptic traits represent
a subject still unexplored, even for the species commonly used for sprouting. A first
preliminary work on this has been carried out by Bianchi et al. (Bianchi, G., et al., 2019)
to analyse taste quality traits and volatile profile in wheatgrass and Triticum spp. sprouts.
Other grains, such as amaranth, quinoa, and buckwheat have been also studied in detail
and appreciated as gluten free products. Several other cultivated species have been studied
occasionally, for example hemp, whose sprouts are destined to become very popular
(Frassinetti, S., et al., 2018).
The results of Bianchi, G. et al work indicate the similarity of the sensory related traits
among Triticum genotypes. Based on Soluble Solid Content and Treatable acidity values,
the palatability of sprouts seems higher than that of wheatgrass. The gas chromatographyolfactometry profiles of wheat sprouts indicate the presence of odor-active compounds
which provide notes such as “earthy and mushroom” in addition to the expected “green”
scent. These characteristics suggest the consumption of wheat sprouts and wheatgrass as
ingredient in vegetable-based products, in order to enhance the taste complexity while
providing valuable phytochemicals, besides the common use as juice (Bianchi, G., et al.,
2019). Other kind of opportunities and research challenges can be found in underexplored
species
for
sprouting
like
voluntary
species,
wild
relatives,
ancestors,
neglected/local/accessions of cultivated species, and even fruit tree species. All of these
species would be very interesting for sprouting purposes because they are supposed to
11
have a higher phytochemical content compared to cultivated species (Kumar, J., et al.,
2020) (Harouna, D.V., et al., 2018). Those species are characterized by high rusticity and
tolerance to extreme environmental conditions that can cope with the negative effects of
climate changes, thereby representing a potential contribution to world food security.
Their resilience (adaptability) is often due to biochemical mechanisms involving higher
contents of secondary metabolites (Montevecchi, G., et al., 2019). This also contributes
to the high seedling vigour and fast vegetative growth, which warrants competitive
adaptability in early growth stages (Gaba, S., et al., 2017). For these reasons, sprouts from
these species are expected to have a very high nutritional value. Some wild species were
proposed and studied for micro-scale vegetables production and many other might be
considered for sprouting: Galium aparine L., Convolvulus arvensis L., Solanum nigrum
L. and Papaver rhoeas L. (Abbas, M.N., et al., 2012) (Bano, Y., et al., 2019) (Senila, L.,
et al., 2020). Of course, all off these species, especially the wild ones, will be effectively
suitable for sprouting only after solving limitations related to low grain yields, their
dispersal at ripening, and seed dormancy (Gaba, S., et al., 2017). Since many wild species
can be considered weeds in the agricultural ecosystems, these studies might also provide
information for weed science and management.
Seeds of fruit tree species often represent an important by-product of juice/jam food
industries, as well as a relevant source of phytochemicals (Villacís-Chiriboga, J., et al.,
2020). In the last decade, many utilizations have been proposed to add value to these
agricultural by-products, including the possibility to use them for nutritional purposes
(Villacís-Chiriboga, J., et al., 2020) (González-Hidalgo, I., et al., 2012). The direct
consumption of seeds is not possible in many cases, since they are coriaceous, and/or
bitter and astringent. Several processing methods have been proposed for extracting
phytochemicals from fruit seeds, while sprouting is a relatively new option, which would
12
help producing high value food while reducing wastes. Until now, fruit seeds sprout use
has been studied only for grapeseed (Weidner, S., et al., 2011), pomegranate (Falcinelli,
B., et al., 2017), olive (Falcinelli, B., et al., 2018) , and Citrus species (Falcinelli, B., et
al., 2020). In most cases, the increase in phytochemical content and antioxidant activity
observed in sprouts in relation to seeds were found to be much higher (e.g., 30-fold for
polyphenols and 90-fold for antioxidant activity in pomegranate) than in herbaceous
species, thus advocating further research. However, among these species, only
pomegranate showed acceptable germination rate and time, and a suitable consistency
and taste to be actually used for the production of edible sprouts. In the other cases,
authors realized that sprouts might better be used for extraction of food additives,
cosmetics, and pharmaceuticals. The limitations related to seed dormancy and low
germination performances appear hard to overcome in some species. However,
considering the large amount of seeds waste obtained from processing, even low
germination percentages could be still suitable for sprouting purposes (Falcinelli, B., et
al., 2020).
2.1.2 Sprouts production
Sprouts production is not only related to commercial scale; indeed, home growing is also
a common low-cost way to produce sprouts. In both cases, what is important, is the use
of Good Agricultural Practice (GAP), which start with the sanification of seeds, which is
achieved by using antimycotic and antibacterial substances like Sodium hypochlorite or
Calcium (Foood and Drugs Administration, 1999). The next step, after sanification, is a
pre-germination treatment which consist of a seeds water soaking for 20 minutes up to 12
hours, according to the seeds dimension and weight. For what concern soaking, it is
important to take care of the duration of this step and while a too short soaking could lead
to a germination rate drop, a too long soaking, could easily lead to unwanted fermentation
13
(Foood and Drugs Administration, 1999). Once imbibition is carried out, the excess of
water is removed to ensure the right humidity condition during the germination process.
Also, water temperature, should not goes under 22-20°C, in order to avoid slowing down
the germination time of grains (Price, T.V., 1988). While germination occurs, the
presence of light is not required, especially for etiolated sprouts, so, generally,
germination doesn’t need any light source. However, it has been observed that some
presence of light could bring to positive effect, increasing the length and diameter of
sprouts hypocotyl or even weight and texture, but it is also important to consider that the
presence of light could bring sprouts to become too much green, or too much tall or thin
(Price, T.V., 1988). Once the sprouting process is accomplished, a phase of ‘’freezing’’
occur, which is intended to stop the process of germination and to increase the shelf life
of sprouts (Cacciola, G., 2014).
Home-made sprouts production can be achieved by using different containers which
could be like simple bowl-shaped container, glass jar, or even last generation germinator.
The difference between these tools is just related to how much grains or seeds con be
sprouted inside them. The ideal container should make possible to easily drain water from
it by putting it upside down or with the help of holes located just at the bottom of the
container. Different kind of containers and germinator are available in the market: jar
with perforated lid, tray boxes and some models even include automatic sprinkler which
waters grains. Glass jar are the most used in sprouts home production because they allow
to easily drain water by using perforated lid and can also become the final container for
sprouts storage after germination occurred (Cacciola, G., 2014).
14
2.2 A look on the other micro-scale vegetables: microgreens and
baby-leaf
Microgreens, as well as Sprouts, can be considered as a functional food, capable to hold
and guarantee all the necessary nutrients for a correct daily diet. Microgreens contain
bioactive compounds which are proven to give some positive actions to the human
organism, and their use is related to a drastic risky disease reducing (DI Gioia, F., et al.,
2015) (Xiao, Z., et al., 2012). If compared to the normal vegetable commercial stage of
harvesting, microgreens show a much more higher antioxidant substances content (Xiao,
Z., et al., 2012) and furthermore, the fresh consumption of sprouts, microgreens and babyleaf, allow to preserve nutrients, vitamins and thermolabile antioxidants from loss by the
heating, if compared to the traditional cooked vegetables (DI Gioia, F., et al., 2015).
Microgreens defined as tender, immature greens, are larger than sprouts, but smaller than
baby vegetables or greens. They have a central stem with two fully developed, nonsenescent cotyledon leaves and mostly one pair of small true leaves (Di Gioia, F., et al.,
2015). The stem, the cotyledons, and the first true leaves are the edible part of
microgreens. Microgreens have been produced in Southern California since the 1980
(Zhang, Y., et al., 2021) and have, since then, gained popularity due to their vivid colours
(like red and purple), delicate textures, and flavour enhancing properties. They are used
as garnishes in salads, sandwiches, soups, appetizers, desserts, and drinks and are highly
appreciated because of their nutritional benefits (Ebert, A.W., 2015) (Galieni, A., et al.,
2020). As well as sprouts, the commercial and home-grown production of microgreens
and baby-leaf comprises several similar aspects like selecting the appropriate species and
growing systems, the phytosanitary quality of seeds and germination, the harvesting and
post-harvest storage practices but also some differences, like the need to use a medium,
which is not intended to be used in sprouts production or the need of irrigation and
15
fertilization. (Di Gioia, F., et al., 2015). By contrast, “baby leaf” vegetables are grown in
the presence of light, either in soil or soilless systems, have a longer growth cycle (20–40
days), usually require the use of fertilizers and agrochemicals and are harvested after the
development of the true leaves (Di Gioia, F., et al., 2017).
Table 2 - Differences among sprouts, microgreens, and baby leaf vegetables (Modified from: Di Gioia et al., 2015)
GROWING CYCLE
SPROUTS
MICRO-GREENS
BABYLEAF
4-10 days
7-28 days
20-40 days
Sprout, roots, seed
Sprout with cotyledons
and first true leaves
Sprout with cotyledons
and true leaves
Without any medium
Soilless / on substrate
Soil / soilless / other
medium
Absence / presence of
light
Light / no light
Presence of light
Not necessary
Not necessary
Necessary
Before cotyledons
formation
Between cotyledon and
first true leaves
Between first true
leaves and 8th leaf
Without cutting
With cutting
/ Without cutting
With cutting
LENGTH
EDIBLE PORTION
PRODUCTION
SYSTEM
LIGHT CONDITION
PESTICIDE USE
PLANT STAGE FOR
HARVEST
HARVESTING
METHOD
2.3 Historical background of edible sprouts
Medicinally and nutritionally, sprouts have a long history. It has been written that the
ancient Chinese physicians recognized and prescribed sprouts for curing many disorders
over 5,000 years ago. Sprouts have continued to be a main staple in the diets of other
populations like Americans of Asian descent, but it took centuries for the West to fully
realize its nutrition merits. The credit for discovering the value of sprouted seeds
traditionally goes to the Chinese. Chinese people used to carry mung beans on their oceangoing ships, sprouting them throughout their voyages and then consuming them in a
16
sufficient quantity to prevent scurvy. The Chinese instinctively knew that an important
factor was missing in their diet during the long travels they carried out, that substance
was vitamin C (International Sprouts Growers Association, 2022).
Today, is still difficult to say when sprouts first showed up as medicine or food in the
Chinese culture. Some proofs say they probably first came into medicinal use during the
Han dynasty (Huang, 2000). Soy sprouts are mentioned twice in the volume which is
widely reputed to be the first Chinese compendium of pharmacopoeia (Bencao), which
author(s) are unknown. The book contains instructions on how to prescribe, administer,
and process 365 drugs that are divided according to their strength and toxicity into three
categories: superior, common, and inferior. Soybeans sprouts are mentioned twice among
the “common” drugs. The use of soybean sprouts as food did not become popular in China
until the Sung dynasty (A.D. 960- 1127), about a thousand years later then the Han
dynasty. During the Sung dynasty, when bean sprouts became popular in China, mung
beans were most widely used for sprouting, but soybean sprouts were also used as food
at that time (Shurtleff, W. & Aoyagi, A., 2003).
Anyway, there are many example of the different cultures using edible sprouts in different
ways and recipes among the history, for example, in India, rice and lentils are fermented
for at least two days before they are prepared as ‘’idli’’ and ‘’dosas’’; in Africa the natives
soak coarsely ground corn overnight before adding it to soups and stews and they ferment
corn or millet for several days to produce a sour porridge called ‘’ogi’’. A similar dish
made from oats was traditional among the Welsh; in some Oriental and Latin American
countries rice receives a long fermentation before it is prepared. Ethiopians make their
distinctive ‘’injera’’ bread by fermenting a grain called teff for several days. All these
typical foods and recipes are witness of how sprouted seeds were diffused in many
different country’s cultures and in the different époques of the human history.
17
3. NUTRITIONAL AND NUTRACEUTICAL
PROPRIETIES OF EDIBLE SPROUTS
Numerous studies have been conducted to investigate the role of sprouts as functional
foods or nutraceutical and while functional foods generally imply health benefits in
addition to nutritional value, the nutraceutical value tend to denote a food (or a part of it)
with an impact on prevention and/or treatment of a specific disease or disorder (Brower,
V., 1998). Sprouts are a good fit for nutrition research, due to their wealth of
phytochemical content combined with reduced antinutrient levels and the expression of
distinct kind of secondary metabolites that are subject to modes of expression which may
largely diverge from patterns and profiles displayed in seeds and mature plants.
From a biological point of view, the very first stage of the plant formation is represented
by the process of germination which leads to the plant tissue formation. In this stage,
seeds start to produce some enzymes which convert the nutritive substances, which are
stored within the seeds in a macromolecular form, in simpler molecules and in energetic
molecules. Those simpler substances are a fundamental key factor in the first stage of
plant development and are used when the plant is not able to run photosynthesis yet.
Sprouts are often defined as ‘’biogenic foods’’, an appellative which highlight the plenty
molecules content with nutritional value such as vitamins, minerals, proteins and many
other components like carbohydrates or lipid (according to the species), with a beneficial
action to human health (Helweg, R., 2011). The enzymatic activation and the hydrolysis
of macromolecules in simpler ones, act like a sort of pre-digestion, which break proteins
in oligopeptides and amino acids; amides and oligosaccharides in simple sugar and lipids
into glycerol and fat acids (MARTON, M., et al., 2010). Furthermore, some research
show how mineral salts and oligo elements can be more easily absorbed by the human
organism thanks to the chelating action which is drove by amino acids and for the
18
possibility to downgrade the phytic acid through the phytase enzyme action (Van Hofsten,
B., 1979). Some interesting fact which has been noticed, is how the vitamins content in
sprouted seeds, is able to grow in just few days of germination (3-5 days), indeed, from
the very start of seed germination process, in just few days, the content of Ascorbic Acid
in sprouted Mung beans grow by eight times if compared to the content of dried beans.
Even the growth of the B group vitamins in some cereal such as wheat, has increased by
six times during the germination process (Cacciola, G., 2014). Some other sprouts, like
soy sprouts, are characterized by a high protein content, which can go up to 13,09 g for a
100 g of soy sprouted seeds, a similar quantity which can be found in eggs, making
sprouts a ‘’fit food’’ and a replaceable product to other high protein content foods. On the
other side, some human health beneficial elements are noticeable only in some sprout
species. Isolflavone is one of these substances. Isoflavones is related to some preventive
actions for a wide range of diseases, like neoplasm, cardiovascular diseases or
osteoporosis and in some species, like Glycine max L., it is possible to find it in high
concentrations, otherwise its content in other legumes species is markedly lower
(NAKAMURA, Y., et al., 2001).
19
Table 3- Water, energy, and nutrients content (on 100 g of fresh products) in some species of sprouts (Source: U.S
Department of Agriculture, 2016).
VEGETABLE
Water
Energy
Proteins
Lipids
Tot Carb
Fibers
Ashes
SPECIES
(g)
(kcal)
(g)
(g)
(g)
(g)
(g)
Brassica oleracea L.
86
43
3,38
0,3
8,59
3,8
1,37
Raphanus sativus L.
90,1
43
3,81
2,53
3,6
2,35
0,53
69
122
13,09
6,7
9,57
1,1
1,59
Lens culinaris L.
67,3
106
8,69
0,55
22,14
5,77
1
Medicago sativa L.
92,8
23
3,99
0,69
2,1
1,9
0,4
Phaseolus vulgaris L.
79,1
67
6,15
0,7
13,5
3,82
0,95
Pisum sativum L.
62,3
124
8,8
0,68
27,11
6,44
1,14
Vigna radiata L.
90,4
30
3,04
0,18
5,94
1,8
0,44
Vigna unguiculata L.
81,3
62
5,25
0,9
11,6
3,38
0,95
Glycine max L.
Walp. subspe
The assessment of biological and nutraceutical effects of sprouts on human health is
typically based on studies involving cellular systems representing given phenotypic
trait(s) with preclinical studies in relevant animal models and, ultimately, human clinical
trials. Cell models generally serve as the biological systems for investigating phenotypic
effects, or mechanistically identifying biologically relevant targets and related molecular
networks. By way of example, this model was applied to show a protective effect by
Brassica oleracea L. sprouts against a human model of gut inflammation (Ferruzza, S.,
et al., 2016). It has been reported that Brassica oleracea L. sprouts extract, reduces the
level of blood glucose in the body. Some others result indicated that Brassica family
sprouts have shown an anti-hyperglycaemic activity. The hypolipidemic activity has also
been observed in bean sprouts (Mendoza-S´anchez, M. & P´erez-Ramírez, I. F.,, 2019).
Sprouts have been also found to contain some antibacterial activity. For instance, broccoli
and pea sprouts have shown the antibacterial activity against Helicobacter pylori, which
is associated with gastric cancer (Ho, C. Y.,, et al., 2006). Different research have
highlighted how sprouts play a significant role in the prevention of various types of
20
cancers due to the presence of biologically active compounds. Literature has reported the
significant antigenotoxic effect of sprouts against DNA damage (Ki, H. H., et al., 2016);
others researches show how Brassica and vegetable sprouts help minimize the risks of
lung and colorectal cancer (Gawlik-Dziki, U., et al., 2012). Epidemiological evidence,
has shown that consumption of broccoli sprouts reduce the risk of different types of
cancers and chronic degenerative diseases (Kensler, T. W., et al., 2012) and recently it
has also been observed that cowpea sprouts decrease cell proliferation and increase the
anti-colorectal cancer action (Teixeira-Guedes, C. I., et al., 2012). All of this evidence,
prove how sprouts can be an ally to prevent different form of risky disorders and diseases
and how they can play a primary role in the human nutrition and prevention.
3.1 Germination process and variation of seed chemical content
The germination process involves events that begin with the uptake of water by the
quiescent dry seed and terminate with the elongation of the embryo axis, usually the
radicle, which extends to penetrate the structures that surround it. Some authors define
the step of radicle penetration as the accomplishment of the germination phase. The
subsequent mobilization of the major storage reserves is associated with the growth of
seedling. The engine that runs this process is represented by some biochemical and
physical events which includes a rapid start of the metabolic activity, the weakening of
the quiescent seeds, the activation of gene transcription, the biogenesis of organelles and
others (Bewley, J.D., 1997 ).
21
Figure 1- Levels of seed water uptake during germination time (Bewley, J.D., 1997).
The uptake of water by the dry quiescent seed is a triphasic process which starts with a
rapid initial uptake (phase I) followed by a plateau phase (phase II). A further increase in
water uptake occurs only after germination is completed, as the embryonic axes elongate,
moving the germination through the phase III. Until dormant seeds do not complete
germination, they cannot enter in the phase III. The phase I is characterized by a rapid
imbibition of water by the dry seeds which go forward until all the matrices and cell
contents are fully hydrated. The influx of water into the cells of dry seeds during phase I
results in temporary structural perturbations, particularly to membranes, which lead to an
immediate and rapid leakage of solutes and low molecular weight metabolites into the
surrounding imbibition solution. Within a short time of rehydration, the membranes
return to their more stable configuration, at which time solute leakage is reduced. One of
the first changes upon imbibition is the resumption of respiratory activity, which can be
detected within minutes. After an initial increase in oxygen consumption, the rate declines
until the radicle penetrates the surrounding structures (Bewley, J.D. & Black, M., 1994).
With few exceptions, radicle extension through the structures surrounding the embryo is
the event that terminates germination and marks the start of seedling growth. In the phase
II, the uptake of water is drastically reduced (plateau phase), but it’s possible to recognize
22
a strong metabolic reactivation. The structures and enzymes necessary for this initial
restoring of metabolic activity are generally assumed to be present within the dry seed,
having survived, at least partially intact, the desiccation phase that terminates seed
maturation. Reintroduction of water during imbibition is sufficient for metabolic
activities to resume. The increase in water uptake associated with Phase III, leads to the
cell elongation and toward the completion of germination (Bewley, J.D., 1997 ). The
increase in water uptake, which is associated with the Phase III, result in a rapidly restore
of the seed metabolic activity, including remobilization, degradation, and accumulation,
which imply important biochemical, nutritional and sensorial changes in the edible
sprouts (Dziki, D., et al., 2015). The outcoming primary and secondary metabolites
produce differential biological health effects when consumed if compared with nongerminated seeds (Di Gioia, F., et al., 2017), (Nelson, K., et al., 2013).
3.1.1 Content of structural carbohydrates
The alteration of grain carbohydrates has been particularly studied thanks to the interest
to one of the most discussed topic, which is involved in the stage of germination, that is
to say the mobilization of complex polymers, such as starch. In germinating grains, the
amylases catalyse the hydrolysis of starch, stored as amylose and amylopectin, to simple
sugars, i.e., the reducing sugars glucose and maltose and for a minor quantity to the nonreducing sugar sucrose, resulting in a higher digestibility (Chung, et al., 2006). However,
in germinated seeds it is possible to observe a different behaviour for what concern the
sugar content and profile, which are related to the different species.
Some of them, like rice, sorghum and millet tend to store more maltose than glucose,
other species like buckwheat are prone to accumulate plenty quantity of only glucose
(Agu, R.C., et al., 2012). Different effects have been reported in response to germination
23
time, with sucrose as the dominant source of carbohydrates during early wheat
germination phase and glucose and maltose during the later stages (Aoki, N., et al., 2016).
3.1.2 Content of non-structural carbohydrates
Dietary fibers represent an important component of the whole grain. Cellulose,
hemicellulose and lignans are water insoluble fibers, while b-glucans and arabinoxylans
(AXs) are grouped as water-soluble dietary fibers. Barley and oats are particularly rich in
b-glucans, as well as sorghum and millet, while other cereals species contain only lower
amounts. The effect of germination on dietary fiber content of sprouted grains is often
inconsistent and strictly depends on fiber fraction, germination time and genotypes. In
germinated wheat the increase in total dietary fibres seemed appreciable at 196 h of
incubation, whilst within the first 48 h of germination time, the fiber content even
decreased. No significant effects were recorded for barley when sprouted for 72 h. In rice,
an increase in total dietary fiber after malting has been observed. In sprouted barley and
oats, the hydrolytic activity of endogenous b-glucanases determined a significant
reduction in b-glucans. The extent of changes in arabinoxylans (AXs) content during
germination has garnered much less attention than b-glucan (Paolo, B., et al., 2019). AXs
are non-starch polysaccharides, found as cell wall constituents. In general, the total AXs
content is not significantly affected by sprouting. However, significant lower content in
AXs has been associated with the germination of oats and rye; during malting of barley,
Han, J. et. al, observed a loss up to 50% of total AXs content (Han, J-Y., 2000).
3.1.3 Proteins content
Cereal proteins are stored in the whole grain and are classified according to their solubility
into albumins (water-soluble), globulins (salt-soluble), glutelins (alkali-soluble), and
24
prolamins (alcohol-soluble). During grain germination, the storage proteins are
hydrolyzed into peptides and amino acids by proteolytic enzymes after 2–3 days from
imbibition, thereby increasing nutrient bioavailability. Some vegetable species like
triticale, barley, rye, oats, and wheat, reduce their content of proteins, during the sprouting
process, whilst some other species did the opposite, studies found. Several authors have
reported an increase in crude proteins in barley, waxy wheat, brown rice and oats.
However, the protein content is strongly influenced by the balance between protein
degradation and protein biosynthesis during germination. Besides an increase in amino
acids contents, significant alterations in free amino acid composition have been observed.
In particular, germinated whole grains contain higher quantities of essential amino acids,
which are a fundamental compound involved in the process of protein production in the
human body. Grain type and germination time have the greatest influence on the amino
acid composition. In waxy wheat, the essential amino acids isoleucine, leucine,
phenylalanine and valine reached the maximum levels after 36 h of germination, while
other essential amino acids (i.e., threonine and methionine) were at the highest level after
24 and 48 h of germination (Paolo, B., et al., 2019).
g-Aminobutyric acid (GABA) is characterized by a rapid increase after sprouting and
several authors report it on studies conducted on wheat and barley. GABA is a fourcarbon non-protein amino acid that is produced primarily by the a-decarboxylation of Lglutamic acid, catalysed by glutamate decarboxylase (GAD). It acts as the main inhibitory
neurotransmitter in the mammalian cortex. Brown rice has reached out a 4-5 times of
GABA increase during germination and even an increase up to 8-12 times occurred after
3-4 days of sprouting with a temperature of 27 to 35°C. (Paolo, B., et al., 2019). However,
besides those results, most of the proteins and amino acid behaviour occurring during
25
germination, is related to the environmental condition and varieties which has been used
(Paolo, B., et al., 2019).
3.1.4 Lipids content
Within the cereal grain, the lipids content is strongly present in the living tissue of the
embryo, scutellum and aleurone, as the form of oil (triacylglycerols, TAG) (Paolo, B., et
al., 2019). It is well known that some kind of lipids, have a crucial importance in the
health of the human being. Indeed, some essential fat acid (EFA) cannot be synthetized
by the human body, so they’re supposed to be introduced through the food diet and while
it is possible to find an abundant content of EFA in many fruits tree seeds such us walnut,
almond or hazel, also sprouted seeds can be considered a source of EFA. However, lentil
and radish linoleic acid and α-linoleic acid content has gained an increase when sprouted
if compared to the content of the dry seed, studies found (Márton, M., et al., 2010).
26
Table 4- Fatty acid composition of lentil seed and lentil sprout (M. Márton, et al 2010)
Fatty Acids
Undecanic acid
Lauric acid
Tridecanonic acid
Myristic acid
Pentadecanonic acid
Palmitic acid
Stearic acid
Oleic acid
Linoleic acid
Arachidic acid
Eicosenoic acid
α-linoleic acid
Behenic acid
Lentil seeds fatty
acids methyl ester
(%)
0.4
0.2
0.2
1.1
0.4
26.2
1.6
14.0
19.4
0.3
0.3
3.3
1.2
Lentil sprouts on
3rd day fatty acids
methyl ester (%)
0.6
0.2
0.4
1.1
0.7
27.0
2.2
9.3
27.4
0.5
0.4
4.7
1.6
3.1.5 Minerals content
During the germination process, while the phytic acid content decrease, the content and
the availability of some minerals increase, and it’s the case of oats, barley, and wheat. For
these species it has been observed a markable major content of Mg after sprouting. In
corn an odd behaviour occurred: the content of the main macro-elements (Na, K, Mg, Ca,
P) tended to decrease after 2 days of germination and then to increase up once reached
the 6th day of germination. Even other minerals such as Fe, Zn, Mn, Cu and Co showed a
clear upward trend over germination time (Paolo, B., et al., 2019).
3.1.6 Antioxidants content
Whole grains contain high concentrations of antioxidants, such as polyphenols,
carotenoids, ascorbic acid, and tocopherols, which balance oxidative damages of seedling
cell components. Polyphenols are natural antioxidant which can be found with abundance
in plants, which is well knew to prevent from the substance’s oxidation. Their role in
27
human health is associated to a preventive action from the oxidation of lipoproteins and
elimination of free radicals, moreover, positive biomedical effects on a cardiovascular
level have been noticed. Polyphenols also play a role in the prevention of disorders and
diseases caused by the effect of senescence and act as a substance that prevent cancer
(Paolo, B., et al., 2019). Generally, germination leads to a mild increase of the total
polyphenols content with the increase of the free phenolic acid fraction and a decrease of
the bound one, as sprouting goes on, as observed in 2-day old wheat, grown under
controlled conditions. The content of different antioxidants in different vegetable sprouts
have shown a similar pattern of variation, characterized by a decreasing at the first stages
of germination and then increasing again on the 6-8 day of sprouting. In buckwheat, the
main flavonoid such as rutin and quercetin, increased their content within 8 days. Also,
other antioxidants such as vitamin C and β-carotene, in wheat, and tocopherol in rice,
showed an increase in their content, if compared to the whole grain (Paolo, B., et al.,
2019).
3.2 Antinutrients and allergens
Antinutrients in plant-based foods, have been for years a subject of much interest in
human health and malnutrition. Antinutritional factors are defined as compounds or
substances, which negatively interfere with the absorption of other nutrients in the diet,
causing reduced nutrient intake, digestion, and utilization, and the occurrence of adverse
effects. In general, the germination process is known to decrease the concentration of
antinutrients in the resulting sprouts, lowering the levels of tannins, phytic acid, and
trypsin. Since the biochemical characteristics of sprouted seeds are strictly related to
sprouting conditions, different techniques, alone or in combination, could be performed
to easily impact antinutritional levels. Some studies used blue and red light to reduce the
28
phytic acid content and soon genetic improvement will produce selected varieties for a
low level of antinutrients or toxins (Khattak, A.B., et al., 2017).
Phytate is highly concentrated in several food items derived from plants; it represents the
major storage form of phosphorus in mature grains and legumes. However, in human
beings the insufficient endogenous intestinal phytase limits phosphorus utilization so
phytate also negatively impacts the bioavailability of mineral ions such as Zn2+, Fe2+/3+,
Ca2+, Mg2+, Mn2+ and Cu2+ since it is characterized by a strong chelation affinity with
cations and is therefore considered an antinutritional factor.
The phytase activity tends to increase during germination; in barley, has been observed a
very low phytase activity at the beginning of sprouting which, in the first couple of days,
increased about 8 times. However, the concentration of phytase in whole grains greatly
varies among cereal species, with rye having the highest values and oats the lowest ones.
However, it has also been observed that phytate content diminishes to a different extent
during germination of brown rice over a period of 12–72 h leaded to a reduction of the
60% in phytate content, whereas in 4-day old sorghum seedlings, a degradation of up to
87% of phytate was observed. Also, brown rice, barley, pearl millet, corn, sorghum and
wheat have performed a decreasing of phytate content along the phase of germination. As
phytate content decreases, bioavailability of phosphorus and minerals increases (Paolo,
B., et al., 2019). The allergenic potential of sprouts and its management is an issue of
major concern in food science as well. Allergic reactions are generally mediated by
proteins that act as antibodies. During sprouting, a deep modification of proteins profile
occurs, thus potentially, the concentration of allergenic storage proteins is reduced
(Khattak, A.B., et al., 2017).
In some cases, the consumption of germinated seeds does not cause an allergic reaction
as severe as that associated to raw seeds, but sprouts can manifest unexpected cross-
29
reactions in sensitive individuals. In mung bean sprouts Vig r 6 and Vig r 1 proteins can
activate basophils high concentrations, and cause the same allergic reaction caused by a
protein responsible for pollen allergy, namely Bet v 1. Cross-reactions can also underlie
allergic conditions, as shown for peanuts and sprouts of different Leguminosae species
(Galieni, A., et al., 2020).
3.3 Quality sprouts and food safeness
Quality standard is not a static concept and as consumers change their needs in lifestyle
and tastes, the definition of quality may be change too. By definition, quality can be
described as ‘’the characteristic sum of a subject which are involved in the satisfaction of
needs that can be expressed, implied or mandatory’’. According to this definition, the
concept of quality implies objective aspects which are related to the subject intrinsic
characteristics and subjective aspects as well, which are bounded with the specific needs
of the consumer (Santamaria, P., et al., 2009). For what concern sprouts, some of the most
important quality traits are related to the length and thickness of the hypocotyl, the
etiolation and other aspects which are referred to the roots system as well as cotyledons.
An ideal sprout, according to some authors, is characterized by a hypocotyl thickness for
at least of 2 mm and a length which must be more than 5 cm, while the length of the roots
can be overlooked. The set of organoleptic traits which can be detected through the taste,
smell and texture are without any doubt another primarily factor which influences the
quality consumer perception and even the appealability (Renna, M., et al., 2017). The
shape, the colour and in general the visual appearance, are some of the aspects which
more impact consumers and their propensity to purchase the product. Another
commercial characteristic related to the visual appearance is related to the sprouts ability
30
to maintain the cellular turgor along the shelf life, which allow to guarantee a good taste
and consistency when consumed (Renna, M., et al., 2017).
It is important to underline that sprouts are a fresh-cut product, which are supposed to be
consumed without any form of baking or with other temperature treatment, thus, another
fundamental aspect is the health and hygiene standards, which must ensure the final
product to be free from bacterial or other micro-organism contamination who could lead
to intoxication. The health and hygiene properties seriously impact and define the
complex of the quality traits of sprouts. The European food safety authority (EFSA)
reported a scientific opinion about the foodborne risks related to some bacterial organism
like Escherichia coli and even other pathogens as Salmonella, which could be commonly
detected in germinated grains. Both pathogens, are capable to produce toxin which cause
alimental intoxication in human. In Escherichia coli the Shiga (STEC) toxin is
responsible for causing the alimental intoxication. The high relative humidity and
temperature conditions which characterized the germination process, represent a good
environment for those pathogens who are located on the external surfaces of the dry seeds,
who can easily spread and become a serious health risk associated with sprouts
consumption (Renna, M., et al., 2017).
31
4. BIOLOGICAL TRAITS OF EDIBLE SPROUTS
In the following chapter the environmental needs and some botanical characteristic of the
species which have been used during the experimental tests, will be displayed. The tests
have been conducted with four different macro-thermal species: leek (Allium porrum L.),
fennel (Foeniculum vulgare Miller var. azoricum) and cucumber (Cucumis sativus L).
4.1 Leek (Allium porrum L.)
Despite there aren’t any certain information about Allium porrum L. origin area, many
researchers agree with saying that leek is actually a Mediterranean native species, already
known and used as vegetable by those population. Leek is a biennial herbaceous plant
which belong to the Liliaceae family. The lower part of the plant is characterized by a
stem from which numerous tiny roots depart, while the upper part hosts the opposite,
linear, sheathed, and acuminated leaves, which is the edible part of the plant. The stem
gets taller and thicker on the second year of growth, and it end with the inflorescence.
Leek bulb is not so thick and neither pronounced, the lower part has a white colour and
consists of some really tiny, wrapped leaves, while it can reach even 20-30 cm of length.
The typology of flower is called Umbrella, it consists of many little flowers which can be
from a white, to a pink or lilac colour. The fruit is a trine trilocular boll, which hosts a
plenty amount of black angular and a little bit flattened seeds. 1000 seeds weight from
2,5 up to 3 gr, while 1 gr of seeds has on average 350-400 seeds. As onion, even leek can
tolerate low temperature, even of 0°C, but for an optimal germination the required
temperature is about 20°C (Bianchi, V. & Pimpini, F., 1990).
32
4.2 Fennel (Foeniculum vulgare Miller var. azoricum)
Fennel is a native species from the Mediterranean basin. Some wild varieties were also
known in the Greek-Roman epoque and in some coastal regions of Italy is still possible
to find it as a wild species. The name come from the old Latin appellative ‘’foeniculum’’
which refers to the typical hairy leaf’s appearance or, perhaps, to the yellowish colour
which the plant assumes when it dries out. Some witness, report that some fennel varieties
that are still used today, were already cultivated in the XI century in Italy, and still today,
Italy has got the primate for the most production of the world of this species. Fennel is a
biennial plant with a brownish or yellowish taproot which can reach 4 cm of thickness,
who hosts numerous lateral radicles. The entire plant has a typical smell which can be
noticed especially when the seeds or the plant get older or become dry. The stem can
reach the length of 150 cm with a 2 cm of thickness at the base, branched and crossed
along all of its length. The pinnatisect leaves are completely divided into lacine threadlike leaves marked, with a huge stalk which end with amplessicaule sheath. Together, the
stem and the sheath, form the edible part which is a fake-bulb also called ‘’grumolo’’.
The Umbrella, a composed flower, which is pentameric, is collected on the apex of the
central and lateral stem and while the main central stem can host even 70 to 80 flowers,
the secondary stems generally produce 30 or 40 flowers. The fruit, or seeds, if we refer
to the commercial name, is a concave schizocarp fruit, glabrous, oblong, oval and
elliptical, with a variable length of 4 to 10 cm, with a yellowish or brownish colour. It
consists of two welded mericarps with a central axis which is called carpophore, while
the pericarp is formed by the mesocarp, the endocarp and the fibrovascular bundles
(Bianchi, V. & Pimpini, F., 1990). Fennel lives in mild climates with optimal temperature
for its growth between 15-20°C. It doesn’t tolerate too low temperature, and a stunted
growth can be observed when the thermic levels are lower than 4-5°C. Temperatures of
33
0-2°C, especially if an extended period occurs, can even cause tissues alteration. Some
experimental trials showed how germination can be carried out more rapidly if different
temperatures are alternated during the day; in particular, 8 hours with 29°C of temperature
followed by 16 hours at 22°C can accelerate the germination process (Bianchi, V. &
Pimpini, F., 1990).
4.3 Cucumber (Cucumis sativus L.)
Cucumber origin centre is probably related to the Himalaya Mountain range, while a
secondary origin centre is located in China, but it’s quite right to affirm that this species
is originated, and in a first moment diffused, all across the Oriental countries. For
instance, in India, where it is still possible to find the wild species C. hardwickii Royle,
Cucumber is a species which has been grown for at least 3000 years. According to the
destination use of the product, different kind of varieties can be distinguished. Some of
them are consumed as a fresh product or for slicing purpose, while others are stored in
oil, and are commonly called pickle. Cucumber belongs to Cucurbitaceae family, and it
runs an annual biological cycle of growing. At a first moment, Cucumber grows a taproot
root system which can explore the soil until 80-90 cm, while later developing a quite
superficial fibrous root system. The herbaceous stem is angular in its section, with a rough
surface full of hard hairs, the stem can be more or less branched, creeping and sometimes
climbing if attached to some support device. With an alternated pattern, leaves depart
from each node of the plant and other secondary branches can develop from the axils of
the leaf. Leaves are always five-lobed leaves, big in their dimension, pendulous, with a
little and long stalk rich in hairs. Cucumber shows different expression of its sexuality.
Some varieties are monoecious with both male and female flowers, which are separated
but on the same plant, while some other plants have only male flowers (andromonoecious)
34
or female flowers. It is also common to find hermaphrodite varieties. Female flowers are
yellow in colour with a little and tubular calyx surrounded by a pentameric corolla, while
male flowers are short peduncles which hosts five stamens, and they are gathered in
groups of three or five flowers. The Cucumber fruit is called Peponide, a fruit which has
an elliptical and elongated form with a 10 to 40 cm of length, circular in its section with
a diameter of 4-6 cm.
The fruit is harvested when is still immature, the edible part consists of a striped
epidermis, uniform in colour which can varies from white to dark green or yellowyellowish when it’s full ripe. Also, the fruit epidermis can be glabrous or rough due to the
presence of trichomes. The mesocarp tissues are white-greenish, aqueous, crunchies and
sometimes slightly bitter due to the presence of bitter substances like cucurbitacine-C.
The endocarp is formed by a reduced placenta which hosts several seeds disposed in a
radial position, in rows of twelve with the hilum facing the mesocarp. Seeds are plenty,
oval in their form, slightly flattened, white hey in colour with e variable length between
0,6 to 1 cm. 1000 seeds weight about 20-30 gr and 1 gram of seeds has on average from
30 to 50 seeds (Bianchi, V. & Pimpini, F., 1990).
Cucumber is a species with high thermal needs, whit some minimum threshold thermal
levels for germination and growth which is around 15°C. Optimal levels of temperature
are located between 25-30°C. At this temperature, emergency can be carried out in 3 to 6
days, while stunted growth can affect the plant if temperatures of 11-12°C occur. For the
rest of the growing cycle, cucumber optimal temperatures are located between 18-20°C
during the night and 24-28°C during the day, with high need even in relative humidity
which should not go under the threshold of 50-70% (Bianchi, V. & Pimpini, F., 1990).
35
5. DIFFERENT USE TYPOLOGIES FOR SPROUTS
Fresh consumption is the main way to consume sprouts. This kind of use allow to
preserve the nutritional and nutraceutical value of this product. It has been told that a fruit
and vegetable-based nutrition is fundamental to preserve the human health, and sprouts
can be considered as a ‘’novel food’’ which can enhance people possibilities to consume
fresh product which contain plenty substances with a positive impact on health such as
vitamins, polyphenols, minerals, and others. The European Union’s (EU’s) Novel Foods
Regulation (EC) No 258/97 is applied to foods and food ingredients that have not been
used for human consumption to a significant degree within the European Community
before 15 May 1997. 'Novel Food' can be newly developed, innovative food, food
produced using new technologies and production processes, as well as food which is or
has been traditionally eaten outside of the EU. Examples of Novel Food include new
sources of K-vitamin (menaquinone), extracts from existing food (Antarctic Krill oil rich
in phospholipids from Euphausia superba), agricultural products from third countries
(chia seeds, noni fruit juice) or food made by new production processes like UV-treated
food (milk, bread, mushrooms, and yeast). As the European regulation said, even
processed food like snacks, juice or fermented drinks, bakery products or pro-biotic
beverage obtained by sprouts can be considered as a novel food (European Commission,
2022).
Beside sprouts nutrition purposes, like fresh consumption or processed food, other
categories of sprout’s utilization have been investigated. One of this, includes drug use.
The biological and pharmacological activities of whole sprouts or their extracts have been
extensively investigated at different levels, including preclinical models, and human
studies cantered on multiple conditions. Despite such a wealth of information, sprouts
were never systematically employed to generate natural product collections aimed at
36
selecting bioactive entities applicable to medicinal and chemistry programs, indeed, this
scenario, seems to be a great purpose for further research and new opportunities of
employment for sprouts. Sprouts can grow quickly, under consistent and controlled
conditions within relatively small footprints and confined layouts, and potentially scaledup for industrial production, furthermore the opportunity to obtain natural products from
sprouts, produced via local greenhouse arrangements, offers environmental and ethical
advantages, like a more sustainable approaches to drug discovery and production. It is
well known that plants can be considered a sort of chemical factory and the same chemical
diversity can be observed also in sprouts. Sprouts diversity and consistency of chemical
compounds may be enhanced as the result of the influence that genetic and environmental
factors, and agronomic setups, which are involved in a drug-based production industries,
thus, sprouts appear to be a good candidate for the OSMAC concept (One Strain Many
Compounds), which refers to the rational, systematic modification of culture conditions
aimed at maximizing the range of secondary metabolites produced in a given system.
Sprouts contain significant amounts of phytochemicals, which have historically been
studied in human clinical trials. In this regard, phenolics (and, particularly, flavonoids)
and the breakdown products of glucosinolates, isothiocyanates, and indoles, have been
investigated, either singly or combined with other treatments (Galieni, A., et al., 2020).
Although many of these chemicals are very well known and have been comprehensively
reviewed, one must note that phenolics account for more than 8000 widely and
differentially distributed entities and glucosinolates are represented by at least 137
different structures, subject as well to genetic and environmental variability. Such
extraordinary diversity, combined with a multi-target chemistry shown by polyphenols,
flavonoids, and isothiocyanates, may represent novel and significant opportunities to
explore, and possibly extend, druggable space. An example of such opportunities is
37
illustrated by sulforaphane, an isothiocyanate resulting from the hydrolysis of its
glucosinolate precursor, glucoraphanin, which is for about 10 up to 100 times more
abundant in broccoli sprouts than in the mature plant (Galieni, A., et al., 2020).
Despite a low amount of literature available, another recent utilization involving sprouts
is for animal feeding purpose. The use of sprouts to supplement animal feeding can be
aimed at maintaining or enhancing animal health as well as transferring phytochemicals
to livestock products (milk, meat, eggs), which are subsequently ingested by humans.
Through this approach, it is possible to bypass the risks to human health posed by
microbiological contamination of raw sprouts. Moreover, sprouts can be integrated into
feed rations in seasons when fodder production is limited. As regards supplementation
levels, studies are needed to evaluate their use in the form of dried powder or pellet
(Galieni, A., et al., 2020). Mattioli et al. demonstrated that drying, in particular freezedrying, did not compromise the bioactive compound contents of alfalfa and flax sprouts,
indeed, early research aimed at studying nutritional properties of rabbit meat was carried
out by researchers, who used alfalfa and flax sprouts and barley sprouts. Both studies
demonstrated an increase in rabbit meat quality. Others research have shown the potential
use of sprouts and microgreens to supplement the diet of pets. Pets tend to be considered
family members, thereby inducing owners to search for healthy and nutraceutical foods,
both for disease prevention and improving quality of life, but further research is needed
in all pet species using multiple varieties of sprouts. Another possible rising interesting
use for sprouts regards feed insects. The sector of edible insects is rapidly expanding and
shows wide research possibilities, including in relation to rearing (Galieni, A., et al.,
2020).
38
6. AIM OF THE THESIS
Sprouts popularity is increasing across the world as research and many studies have
recognized them as a functional food for their nutritional and nutraceutical properties (DI
Gioia, F., et al., 2015), and as a functional fresh vegetable product for the possibility to
grow them without any particular difficulty and with low investments (Marra et all.,
2000). Indeed, sprouts can be produced quickly, easily, and cost-effectively due to simple
requirements for equipment and supplies, in a developmental process which can be
carried out from a few days to approximately two weeks. At the same time, agricultural
growing practices are also evolving toward more sustainable systems, in contrast to
climate change, with the main aim of reduce inputs without reducing the yield and the
quality of the products. In this context, the production of sprouts fit all this goals thanks
to their low requirements which allow to avoid the use of agrochemicals and fertilizers.
Those conditions, suggest a unique opportunity for industrial scalability, coupled with the
prospect for consumers to independently access food with proven or purported nutritional
benefits (Renna, M., et al., 2017). Today, people are becoming increasingly conscious of
the need to adopt a healthy style of living and their preferences of consume are going
toward diets based on fresh fruits and vegetable products, which are characterized by a
high content of bioactive molecules with a positive impact on human health. Like
commons vegetable that are daily consumed, also sprouts are known to possess such
bioactive compounds with antioxidant and antiviral activity, which act as an immune
system activity stimulant, with antidiabetic activity and many other functions associated
to their consume for human and even animals (Saha, M.A, et al., 2016). Polyphenols are
one of the most important groups of bioactive molecules which can be found with
abundance in plants. They are characterized by a prime role into preventing oxidation of
lipoproteins and elimination of free radicals, acting as a preventive cure for all the
39
diseases or disorders which are associated with aging. Some of the most abundant
polyphenols which can be found in plants and sprouts are flavonoids, such as rutin and
quercetin; carotenoids, like vitamin C and β-carotene, vitamins such as tocopherol and
many others (Paolo, B., et al., 2019). For instance, sprouted buckwheat is well-known for
its antioxidant, antihypocolesterolemic and neuroprotective functions (Bestida, J.A.G &
Zielinski, H, 2015), while red cabbage and broccoli sprouts are popular brassica vegetable
that exhibit antimicrobial, anticancer as well as anti-obesity properties (Saha, M.A, et al.,
2016). Many factor, such as the time of germination, the light vs dark conditions, the
environmental temperatures and even the seeds quality and sanitary safety can influence
the physiological process and biosynthetic pathways in sprouts, leading to different
expression for what concern the quality and quantity of secondary metabolites
(Mastropasqua, L., et al., 2020). Plant species in the Poaceae, Brassicaceae, and
Fabaceae families are the most exploited for sprouting purposes and therefore most
reported by the scientific literature. For these species, research has provided a lot of
information on the nutritional traits of sprouts (Renna, M., et al., 2017), while other
species like horticultural species, fruit tree species and wild species, are appreciated but
relatively used and studied for sprouting purpose, enough to be considered as underused
or neglected species (Galieni, A., et al., 2020). According to the different ranges of
germination temperatures (22-24-26°C) and light intensity (of 0 µmoL/m²/s and 93
µmoL/m²/s), which have been reproduced in a controlled environment (a germination
chamber), the fresh wight of sprouts, the quantities of chlorophyll pigments (both A and
B), the antioxidant power and the total phenolic content were evaluated. The chosen
horticultural species, are some which are diffused in the Mediterranean basin, therefore
characterized by high thermal needs for their germination and successive growth.
40
Those species have been less or not even used for sprouting purposes and are represented
by cucumber (Cucumis sativus L.), onion (Allium cepa L.), leek (Allium porrum L.) and
fennel (Foeniculum vulgare Miller var. azoricum).
41
7. MATERIALS AND METHODS
7.1 Plant materials, seeds germination, light and thermal conditions
Different non-professional brand of seeds of onion (Allium cepa L.), fennel (Foeniculum
vulgare Miller), leek (Allium porrum L.) and cucumber (Cucumis sativus L.) were
purchased from different local stores. As a first operation, 150 seeds were weighted in
order to distribute them on each Petri dish. Then, all the seeds were sanitized with a 5%
of sodium hypochlorite solution for 10 minutes and then washed carefully for another 5
minutes in distilled water to eliminate any trace of the sanitizer solution. After washing,
the seeds were left to dry on a tray with absorbent paper for 15 minutes. After that, 150
seeds of every species were divided into a 90 mm Petri dish. For every species, six
replicates were produced. Each Petri dish hosted two layers of filter paper with 90 mm of
diameter which were moistened with 3 ml of distilled water in order to hydrate the seeds.
Then, the seeds were incubated in a plant growth chamber (95 x 80 x 100 cm) with
different temperature according to the different experimental testing (22, 24 and 26 °C)
and different light intensity (0 and 93 µmoL m² s-1) with a 24h photoperiod. For the light
intensity level of 0 µmoL m² s-1, the lamps were switched off. The seeds were moistened
every day or as needed with different quantities of water according to the seeds water
consumption. Seedlings were collected according to the used temperature, with a
maximum period of time of 14 days given to the seed to allow germination.
42
Figure 2 – Onion sprouts collected at the 11th day of germination at 24°C in darkness conditions.
7.2 Determination of the fresh weight and dry matter content
The dry matter was determined after collecting the sprouted seeds from the growing
chamber. Sprouts were weighted (fresh matter) after the accomplishment of germination
as quickly as possible in order to limit the losses through evaporation and then then freezedried until reaching a constant weight (approximately for 24-48 h). The dry matter content
(g) was expressed as a percentage of the ratio of constant weight following drying and
the initial fresh matter.
7.3 Sprouts growth parameters
At the end of the cycle, the sprouts have been weighed and counted as well as the
ungerminated seeds. From the ratio between the seeds germinated and the total seed used
the percentage of germination was calculated (germinability %). In the same way, at the
43
end of the cycle, the sprouts were counted and weighed in order to obtain the fresh weight
per 1000 sprouts. The germination energy was evaluated according to the ISTA (2006)
standard. The seeds that sprouted were counted each day from the second to the last day
following positioning in Petri dishes. For each batch of 50 seeds, the seeds that
germinated normally were counted, taking into account the initial and final counts. The
germinative energy, expressed as days of average germination time (TMG) was
calculated by summing the product of seeds germinated in each day by the number of
days taken to germinate and, dividing this sum by the total number of seeds germinated.
Figure 3 - Cucumber sprouts collected at the 7th day of germination at 26°C in a dark environment.
7.4 Determination of the antioxidant capacity (DPPH)
The DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging activity of sprouts extract
was determined according to Brand-Williams et al. (1995). Three-hundred mg of
lyophilized sprouts powder were mixed with 5 mL methanol (80%) and vortexed for 1
minute. Samples were then submitted to 10 minutes of ultra-sonic bath (below 10 °C) and
centrifuged for 15 minutes at 4000 g and 6 °C. For the reaction, 150 µL of supernatant
44
were mixed with 1350 µL of DPPH solution (150 µmol), then samples were vigorously
agitated and placed in the dark for 30 minutes. The decrease in absorbance of methanolic
solution of DPPH was read at 515 nm and DPPH values were obtained from a straight
line (r = 0.999***) obtained by change in absorbance against different Trolox
concentrations; the results were expressed as µmol Trolox equivalents (TE) g-1 DW.
7.5 Determination of Total Carotenoids Concentration (TCC)
Total carotenoids concentration (TCC, including xanthophylls) in fruits was determined
according to Lichtenthaler and Wellburn (1983), with slight modifications. Threehundred mg of lyophilized tomato powder were extracted with 5 mL acetone (80%),
vortexed for 1 minute, then left overnight in the dark at 10 °C. After that, samples were
sonicated for 10 minutes through an ultrasonic bath (FALC INSTRUMENTS) (below 10
°C) and centrifuged on a Neya 10r professional (REMI) centrifuge for 10 minutes (5000
g at 6 °C). Samples were read using acetone 80% as blank. Readings were done at 470,
649 and 665 nm and the absorbance values (A) were substituted in the following
equations:
-
Chlorophyll A= 12,25 x ABS663.6 – 2,55 x ABS646.6 (µg/ml)
-
Chlorophyll B= 20,31 x ABS646.6 – 2,55 x ABS663.6 (µg/ml)
-
Carotenoids:
((1000 x ABS470) − (1,82 x (chlor. a)) – (85,02 x (chlor. b))
198
(µg/ml).
7.6 Determination of the Total Polyphenol Content (TPC)
For the extraction of TPC, the method proposed by Nunzia Cicco et al. (2008) was
followed. 0.03 g of freeze-dried sample flour was mixed with 2 mL of 80% methanol in
an Eppendorf tube. This mixture was stirred for 15 minutes in a warm bath in darkness at
70 °C using a WBMF 24 (FALC INSTRUMENTS) then it was centrifuged at 4000 g for
45
5 min in a Neya 10R professional (REMI) and the extract was transferred to another
Eppendorf tube. Then, 50 µL of supernatant were mixed with 50 µL of methanol with 0.7
ml of Folin-C (1:10).
The mixture was vortexed and keep 5 min in dark to make the reaction happen and
following, 0.7ml of Na2CO3 (6%) were added. The mix was vortexed again and in
incubated for 60 minutes in the darkness.
The absorption was checked at 765 nm, and results were expressed as mg Gallic acid g-1
of fresh weight (FW).
7.7 Statistical procedures
All collected and calculated data were firstly subjected to Shapiro-Wilk’s and Levene’s
test, in order to check for normal distribution and homoscedasticity, respectively, then to
a factorial ‘Temperature level × Light level’ analysis of variance (ANOVA), according
to the experimental layout adopted in the field. Percentage data were Bliss’ transformed
before the ANOVA (untransformed data are reported and discussed) whereas multiple
mean comparisons were performed through the least significant difference (LSD) test (at
least for P ≤ 0.05). All calculations were performed using Excel version 2016 (Microsoft
Corporation, Redmond, WA) and Minitab version 16.1.1 (Minitab Inc., State College,
PA, USA)
46
8. RESULTS
8.1Sprouts Biomorphometric Characteristics
8.1.1 Leek
Among the main factors, leek germinability (%) showed no difference among different
temperature adopted, while a significant difference turned out between the L0 and L90
light level, with the first did register significantly higher value (64.4% and 62.0%,
respectively) (Table 1). As regards the average germination time (TMG) in leek, there
was a significant difference between the three temperatures used, with higher values at
the temperature of 22 °C and 24 °C degrees (6.5 days for both), while a significantly
lower number of days was recorded at 26 °C (6.0 days). In this case a lower number of
days for germination is a positive index, and here the lowest value was recorded at 26 °C
(no optimal temperature). As regards the effect of the light treatment, a significant
difference was recorded between L0 and L90 with higher values at L0 (6.5 and 6.2 days,
respectively) (Table 1). The fresh weight of 1000 sprouts of leek was significantly
influenced both by the temperatures adopted and by the light treatments. Specifically,
among the three thermal regimes used, the highest value was recorded at 24 °C (19.4 g),
while the lowest value was recorded at 26 °C (5.1 g). The effect of the light treatment was
also significantly different, showing the highest value at L0 and the lowest value at L90
(15.8 and 12.1 g, respectively) (Table 1). Among the main factors, in leek the dry matter
(DM) content was significantly affected by temperature, the higher values were recorded
at 26 °C and 22 °C (30.0 and 22.0 g, respectively) while the lower value was recorded at
24 °C, which represents the optimum (15.4 g). L90 recorded the highest significantly
value between the light intensities, equal to 25.1 g.
47
8.1.2 Fennel
The percentage of germination of fennel was significantly influenced by both the
temperatures and the light treatments applied. The highest values for germinability were
recorded respectively at 22 °C and 24 °C, while the lowest value was recorded at 26 °C
(79.1%, 76.5% and 74.5%, respectively) (Table 2). The fennel % of germination was not
affected by the light treatments (Table 2). The average germination time of fennel was
significantly influenced by the thermal regimes adopted, in particular the highest value
was recorded at 22 °C, the average at 24 °C and the lowest at 26 °C (6.2, 6.0 and 5.7 days,
respectively). As for the light treatments, L0 showed a significantly higher value than the
L90 (6.1 and 5.9 days, respectively) (Table 2). The fresh weight per 1000 sprouts recorded
the significantly higher value at 26 °C (21.2 g) while there were no differences between
22 and 24°C. As regards the effect of the light treatments, the highest value was recorded
at L0 equal to 19.6 g while the significantly lower value of 16.2 g was recorded at L90
(Table 2). The percentage of dry matter (DM%) in fennel was not significantly influenced
by the thermal regimes adopted, while a significant difference was reported in response
to light treatments. In particular, the significantly higher value for percentage of dry
matter was recorded at L90, equal to 17.9% (Table 2).
8.1.3 Cucumber
The percentage of germinability of cucumber sprouts was significantly influenced by the
thermal regimes applied but not by the applied light treatments. The percentage of
germinability with the significantly higher value was recorded at 24 °C (89.0%), while a
no difference between the values recorded at 22 °C and 26 °C (73.0% and 74.8%,
respectively) (Table 3). As regards the average germination time for the cucumber, the
significantly higher value was recorded at 24 °C (2.2 days), while the lower values at 22
48
°C and 24 °C, respectively. The mean germination time was not significantly influenced
by applied light regimes (Table 3). The fresh weight per 1000 sprouts was not
significantly influenced by the light regimes applied, while it was significantly influenced
by the thermal regimes. Specifically, the highest value was reported at 22 °C (105 g),
while the lowest value was instead reported at 24 °C (66 g) (Table 3). Similarly to the
fresh weight per 1000 sprouts, the dry matter content for the cucumber was not
significantly influenced by the light regimes applied. However, the dry matter content
was significantly influenced by the different thermal regimes, with the highest value
recorded at 26 °C (29.2%) (Table 3).
8.2Sprouts Biochemical Characteristics
8.2.1 Leek
For leeks sprouts there was a significant effect of temperature levels applied. The highest
value of total phenols content was found at 24 °C (306 mg GAE kg-1 FW), at the optimum
for the species, while the significantly lower value was instead recorded at 26 °C (94 (mg
GAE kg-1 FW). The different light conditions also had a significant influence on the total
content of phenols, specifically the highest value was detected in dark conditions (235
mg GAE kg-1 FW) (Table 4). Similar to the content in total phenols, the DPPH assay was
significantly affected by light treatment with the highest value recorded in dark
conditions, L0 (13% of inhibition) than at L90 (12% of inhibition). As regards the effect
of the different thermal regimes, the highest value was recorded at 26 °C (18% of
inhibition) while the lowest value was recorded at 22 °C (8% of inhibition) (Table 4).
As regards the effect of light regimes on chlorophyll A, chlorophyll B and total
carotenoids content it was possible to find a common trend. It is well known that light is
necessary for the synthesis of this of these compounds and, as direct a consequence, the
49
highest values for chlorophyll A, chlorophyll B and total carotenoid content were
recorded at L90 (221,71 and 69 mg kg-1 FW, respectively) than a L0 (134,49 and 32 mg
kg-1 FW, respectively) (Table 4). As regards the effect of the thermal regimes applied, the
highest value for chlorophyll A was recorded at 26 °C (208 mg kg-1 FW) and the lowest
at 22 °C (144 mg kg-1 FW). As regards chlorophyll B, the statistically higher value was
recorded at 26 °C (121 mg kg-1 FW).), while for total carotenoids content it was recorded
at 24 °C (69 mg kg-1 FW).
8.2.2 Fennel
The total content of phenols for fennel was significantly influenced by the thermal
regimes used. Specifically, the highest value was recorded at 24 °C (679 mg GAE kg-1
FW) at the optimum for the species. The different light regimes applied, also, significantly
influenced the total content of phenols in fennel, with the highest value recorded in dark
conditions, equal to 680 mg GAE kg-1 FW (Table 5). Similarly to the total content in
phenols, for the DPPH test the highest value was recorded at 24 °C, in correspondence
to the optimum of germination for this species, equal to 37 mg GAE kg-1 FW. Conversely,
the opposite trend was recorded As regards the effect of the light regime, the significantly
highest value was recorded in lighting conditions L90 than in dark conditions (31 vs 26
mg GAE kg-1 FW, respectively) (Table 5).
As previously reported for leek, in fennel sprouts the chlorophyll A, the chlorophyll B
and the total carotenoids content in relation to the light regimes applied, were significantly
higher at L90 than in dark conditions (164, 44 and 77 mg kg-1 FW at L90, and 79,25 and
23 mg kg-1 FW at L0, respectively). As for the temperature effect, when chlorophyll A
was concerned, the significantly higher value was recorded at 22 °C as well as for total
carotenoids content (155 and 57 mg kg-1 FW, respectively). As regards the chlorophyll B
50
content of fennel sprouts, the highest value was recorded at 26 °C (52 57 mg kg-1 FW),
while the lowest values, not significantly different from each other, were recorded at 22
°C and 24 °C (Table 5).
8.2.3 Cucumber
In cucumber sprouts the total content of phenols and the DPPH assay were significantly
influenced by the thermal regimes, with the highest value being recorded at the optimum,
i.e. 24 °C for both (229 mg GAE kg-1 FW and 7.3% of inhibition). As regards the effect
of light on the total phenol content, there were no statistically differences between the
two regimens used, whereas the DPPH assay was concerned, the highest significative
value was recorded under lighting conditions, L90 equal to 6.3% of inhibition (Table 6).
Mirroring the previously exposed trends for leek and fennel sprouts, the cucumber sprouts
showed a pattern for chlorophyll A, chlorophyll B and total carotenoids content. In detail,
regarding the effect of thermal regimes on these compounds, the significantly higher
values have always been recorded at 24 °C, that is in correspondence with the optimum
for the species, for chlorophyll A, chlorophyll B and total carotenoids content (286,171
and 135 mg kg-1 FW, respectively). Similarly, as regards the effect of the light regimes
used, the highest values have always been recorded under present light conditions, L90
(244, 143 and 109 mg kg-1 FW, respectively) than at L0 (150, 78 and 37 mg kg-1 FW,
respectively) (Table 6).
51
9. CONCLUSIONS
The use of sprouts in diets is closely related to the nutritional and nutraceutical benefits
derived from their consumption. The beneficial effects exerted by germination, produce
changes in the chemical composition of raw seeds, which is usually reflected in an
increase in the antioxidant capacity, the chlorophyll content and in the total amount of
polyphenols. It is well known that all these properties can be influenced by several
conditions, including the environment of growth, in particular light and temperature.
Consequently, different growth conditions can produce changes in the quality of the
sprouts. In this study, as a consequence of light treatments (0 or 93 µmoL/m²/s) and
according to the different temperature applied (22°C - 24°C - 26°C) during the sprouts
growing cycle, the quality-related parameters were differentially influenced in the three
different vegetable species, characterized by mid-high thermal needs as above reported.
However, the investigation of the specific growth conditions capable to enhance sprouts'
nutritional and nutraceutical quality of the different vegetable species must be more
investigated, in order to increase the production of sprouts and the subsequent
consumption by those people with specific nutritional needs or by those conscious
consumers who understand the nutritional value of sprouts for the health.
52
TABLES
Table 1. Biomorphometric characteristics of leek sprouts 'De Carentan' in relation to the
factors under study. Different letters within each factor indicate significance on Fisher's
LSD test (P < 0.05). L0: 0 µmol m-2 s-1; L90: 90 µmol m-2 s-1; TMG: average
germination time; FW: fresh weight; DM: Dry matter. NS: not significant
Variable
Temperature L0
L90
Mean
LSD interaction
(P < 0.05)
Germinability
22 °C
64.7
52.0
58.3 a
(%)
24 °C
67.2
50.8
59.0 a
26 °C
61.3
53.1
57.2 a
Mean
64.4 a
52 b
TMG
22 °C
6.6
6.4
6.5 a
(days)
24 °C
6.9
6.2
6.5 a
26 °C
6.0
5.9
6.0 b
Mean
6.5 a
6.2 b
FW 1000 sprouts
22 °C
18.4
15.9
17.2 b
(g)
24 °C
23.0
15.9
19.4 a
26 °C
5.9
4.4
5.1 c
Mean
15.8 a
12.1 b
DM
22 °C
17.9
26.1
22.0 b
(%)
24 °C
12.4
18.4
15.4 c
26 °C
29.2
30.9
30.0 a
Mean
19.8 b
25.1 a
NS
0.2
2.2
2.1
53
Table 2. Biomorphometric characteristics of fennel sprouts 'Romanesco' in relation to the
factors under study. Different letters within each factor indicate significance on Fisher's
LSD test (P < 0.05). L0: 0 µmol m-2 s-1; L90: 90 µmol m-2 s-1; TMG: average
germination time; FW: fresh weight; DM: Dry matter. NS: not significant.
Variable
Temperature L0
L90
Mean
LSD interaction
(P < 0.05)
Germinability
22 °C
82.8
75.4
79.1 a
(%)
24 °C
73.8
79.1
76.5 a
26 °C
75.5
73.4
74.5 b
Mean
77.4 a
76.0 a
TMG
22 °C
6.4
6.0
6.2 a
(days)
24 °C
6.1
6.0
6.0 b
26 °C
5.8
5.6
5.7 c
Mean
6.1 a
5.9 b
FW 1000 sprouts
22 °C
17.8
15.7
16.7 b
(g)
24 °C
18.7
13.0
15.9 b
26 °C
22.4
20.1
21.2 a
Mean
19.6 a
16.2 b
DM
22 °C
12.9
18.3
15.6 a
(%)
24 °C
11.4
17.7
14.6 a
26 °C
12.4
17.7
15.0 a
Mean
12.2 b
17.9 a
3.4
0.2
1.8
NS
54
Table 3. Biomorphometric characteristics of cucumber sprouts 'Marketmore' in relation
to the factors under study. Different letters within each factor indicate significance on
Fisher's LSD test (P < 0.05). L0: 0 µmol m-2 s-1; L90: 90 µmol m-2 s-1; TMG: average
germination time; FW: fresh weight; DM: Dry matter. NS: not significant.
Variable
Temperature
L0
L90
Mean
LSD interaction
(P < 0.05)
Germinability
22 °C
70.2
75.7
73.0 b
(%)
24 °C
89.0
89.0
89.0 a
26 °C
72.1
77.4
74.8 b
Mean
77.1 a
80.7 a
TMG
22 °C
2.0
2.1
2.1 b
(days)
24 °C
2.2
2.2
2.2 a
26 °C
2.1
2.0
2.0 c
Mean
2.1 a
2.1 a
FW 1000 sprouts
22 °C
124.3
86.3
105 a
(g)
24 °C
63.0
69.6
66 c
26 °C
90.0
88.3
89 b
Mean
92 a
81 b
DM
22 °C
19.5
27.3
23.4 b
(%)
24 °C
24.1
20.6
22.4 b
26 °C
28.2
30.1
29.2 a
Mean
23.9 a
26.0 a
6.0
0.1
16
5.1
55
Table 4. Biochemical characteristics of leek sprouts 'De Carentan' in relation to the factors
under study. Different letters within each factor indicate significance on Fisher's LSD test
(P < 0.05). L0: 0 µmol m-2 s-1; L90: 90 µmol m-2 s-1; GAE: gallic acid equivalents; FP:
fresh weight; NS: not significant.
Variable
Temperature L0
L90
Mean
LSD interaction
(P < 0.05)
Total Phenols content
22 °C
217
135
176 b
(mg GAE kg-1 FW)
24 °C
381
232
306 a
26 °C
106
83
94 c
Mean
235 a
150 b
Chlorophyll A
22 °C
111
178
144 c
(mg kg-1 FW)
24 °C
122
239
180 b
26 °C
169
247
208 a
Mean
134 b
221 a
Chlorophyll B
22 °C
22
33
27 b
(mg kg-1 FW)
24 °C
21
40
30 b
26 °C
103
139
121 a
Mean
49 b
71 a
Total carotenoids
22 °C
27
45
36 b
(mg kg-1 FW)
24 °C
39
98
69 a
26 °C
31
64
47 b
Mean
32 b
69 a
DPPH
22 °C
8
8
8c
(% inhibition)
24 °C
9
12
11 b
26 °C
20
17
18 a
Mean
13.0 a
12.0 a
24
42
15
20
2
56
Table 5. Biochemical characteristics of fennel sprouts 'Romanesco' in relation to the
factors under study. Different letters within each factor indicate significance on Fisher's
LSD test (P < 0.05). L0: 0 µmol m-2 s-1; L90: 90 µmol m-2 s-1; GAE: gallic acid
equivalents; FP: fresh weight; NS: not significant.
Variable
Temperature
L0
L90
Mean
LSD interaction
(P < 0.05)
Total Phenols content
22 °C
592
431
512 b
(mg GAE kg-1 FW)
24 °C
775
582
679 a
26 °C
672
421
546 b
Mean
680 a
478 b
Chlorophyll A
22 °C
100
210
155 a
(mg kg-1 FW)
24 °C
62
160
111 b
26 °C
75
123
99 c
Mean
79 b
164 a
Chlorophyll B
22 °C
22
38
30 b
(mg kg-1 FW)
24 °C
14
30
22 b
26 °C
41
64
52 a
Mean
25 b
44 a
Total carotenoids
22 °C
32
81
57 a
(mg kg-1 FW)
24 °C
19
53
36 b
26 °C
20
36
28 b
Mean
23 b
57 a
DPPH
22 °C
22
26
24 b
(% inhibition)
24 °C
31
42
37 a
26 °C
25
24
25 b
Mean
26 b
31 a
87
10
14
11
5
57
Table 6. Biochemical characteristics of cucumber sprouts 'Marketmore' in relation to the
factors under study. Different letters within each factor indicate significance on Fisher's
LSD test (P < 0.05). L0: 0 µmol m-2 s-1; L90: 90 µmol m-2 s-1; GAE: gallic acid
equivalents; FP: fresh weight; NS: not significant.
Variable
Temperature L0
L90
Mean
LSD interaction
(P < 0.05)
Total Phenols content
22 °C
186
129
157 b
(mg GAE kg-1 FW)
24 °C
187
271
229 a
26 °C
108
95
102 c
Mean
160 a
165 a
Chlorophyll A
22 °C
117
93
105 c
(mg kg-1 FW)
24 °C
201
371
286 a
26 °C
133
268
201 b
Mean
150 b
244 a
Chlorophyll B
22 °C
56
47
51 c
(mg kg-1 FW)
24 °C
122
219
171 a
26 °C
55
163
109 b
Mean
78 b
143 a
Total carotenoids
22 °C
28
26
27 b
(mg kg-1 FW)
24 °C
55
215
135 a
26 °C
28
84
56 b
Mean
37 b
109 a
DPPH
22 °C
5
7
5.7 b
(% inhibition)
24 °C
6
9
7.3 a
26 °C
4
3
3.5 c
Mean
4.7 b
6.3 a
40
57
37
46
1.4
58
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