Download 1. INTRODUCTION The dried cells of

1. INTRODUCTION
The dried cells of microorganisms such as bacteria, fungi, yeasts
and algae that are grown in large-scale culture systems as proteins, for
human or animal consumption are collectively known as single cell
proteins (Sasson, 1997). Single cell proteins (SCP) are characterized by
fast growth rate, high protein content (43-85%) compared to field crops,
require less water, land and independent of climate, grow on wastewater,
can be genetically modified for desirable characters such as amino acid
composition and temperature tolerance (Tri-Panji and Suharyanto.
2001). Microorganisms used to produce biomass and the choice of a
microorganism depends on numerous criteria, the most important of
which
is
the
nature
of
the
raw
material
available.
The
ideal
microorganism should possess the following technological characteristics
like
high
specific
growth
rate,
biomass
yield,
low
nutritional
requirements, ability to develop high cell density, stability during
multiplication, capacity for genetic modification and good tolerance to
temperature and pH. In addition, it should have a balanced protein and
lipid composition. It must have a low nucleic acid content, good
digestibility and to be non-toxic.
Nowadays, food industries need new food ingredients obtained
from
natural
sources
and
nutraceuticals. Microalgae
developed
now
combine
novel
the
functional
traditional
foods
and
or
new
biotechnologies where microalgal biomass can be used as a source of
proteins, biochemicals, lipids, polysaccharides and colorants (Athukorala
et al., 2006). Among microalgal species, Spirulina utilized as food in the
area around Lake Chad for long time ago and marketed as a healthy
aliment in the United States and Japan (Borowizka, 1998). Spirulina
(Arthrospira) is a blue-green algae found in alkaline Lakes around the
world. The name “Spirulina” is derived from the Latin word for “helix” or
“spiral”, referring to the physical structure of the organism. It is motile
multicellular filamentous blue-green algae and reproduces by binary
fission. Spirulina is a non nitrogen-fixing blue-green alga and cell wall
made of mucopolysaccharide its soft and easily digestible nature, which
makes it safe for human consumption. Spirulina is capable of growing in
high alkalinity with the presence of carbonate, bicarbonates and
inorganic nitrogen (Aiba and Ogawa, 1977; Yang et al., 2010). The ability
of Spirulina to grow in hot and alkaline environments ensures its
hygienic status, as no other organisms can survive to pollute the waters
in which this alga thrives. Spirulina is one of the cleanest, most naturally
sterile foods found in nature. It has been used as feed for fish, poultry
and farm animals (Tragut et al., 1995; Abdulqader et al., 2000).
Microalgal species are an important alternative material to extract
natural antioxidant compounds to delay or prevent the oxidative damage
caused by Reactive Oxygen Species (ROS) molecules (El-Baz et al., 2002).
Spirulina has reported to prevent oxidative damage and hence can
indirectly reduce cancer formation in human body. In this respect, the
increased consumption of foods characterized by free radical scavenging
activity, leads up to a doubling of protection against many common types
of cancer formation (Cooke et al., 2002; Romay et al., 2003; Anbarasan et
al., 2011).
The United Nations world food conference declared Spirulina as
“the best for tomorrow” and it is gaining popularity in recent years as a
food supplement (Kapoor and Mehta, 1993). Arthrospira (Spirulina) is an
economically
important
filamentous
cyanobacterium.
The
annual
production of the microalga is about 10,000 tons which makes it the
largest microalgal cultivation industry in the world (Zhang and Skolnick,
2005). The Spirulina platensis was recognized a “wonderful food for
health” since it contain high proteins (55-70%), (Umesh and Sheshagiri,
1984; Sanchez and Bernal, 2006), bioactive compounds such as,
essential
fatty
acids
(4-7%)
like,
linolenic
and
DŽ-Linolenic
acid
(Borowitzka, 1998; Othes and Pire, 2001; Sanchez and Bernal, 2006;
Kumar et al., 2012), vitamins like, provitamin A, vitamin B complex, B1
(thiamine), B2 (riboflavin), B3 (nicotinamide), B6 (pyridoxine), B9 (folic
acid), B12 (cyanocobalamin), vitamin C, vitamin D
and vitamin E
(Richmond, 1992; Belay, 1997), bio-pigments like phycocyanin and
chlorophyll-a (Achmadi and Tri-Panji, 2000; Manojkumar et al., 2011)
and antioxidant compounds, carotenoids (Cohen, 1997; Kawata et al.,
2004; Chen et al., 2006). Based on these effects, Spirulina was approved
in Russia as “medicine food” for treating radiation sickness (Becker,
1984)
The large-scale production of Spirulina biomass depends on many
factors, the most important of which are nutrient availability, pH,
temperature and light. These factors can influence the growth of
Spirulina and the composition of the biomass produced by causing
changes in metabolism, which considerably modify the time course of the
accumulation of the main biomass components.
In 1950, the United States and Japan began the experimental
cultivations of Spirulina investigated its chemical composition and
industrial applications. The commercial production of Spirulina started in
the early 1960s in Japan, in the early 1970s at Lake Texcoco, Mexico
and the third major microalgae industry was established in Australia in
1986. Currently, the commercial production of Spirulina is mainly
oriented towards the health food market, utilizing a chemically defined
medium (Belay et al., 1993). Spirulina is marketed and consumed in
Germany, Brazil, Chile, Spain, France, Canada, Belgium, Egypt, United
States, Ireland, Argentina, Philippines, India, Africa, and other countries,
where public administration, sanitary organizations and associations
have approved for human consumption. The first commercial plant for
cultivation of Spirulina in seawater was set up at the Tropic Sanya area
in 1992. The total area of seawater Spirulina cultivation is about 40000
m2 for production capacity of above 80 tons. Commercial products of
seawater Spirulina including algal powder, tablet, natural blue colorant
phycocyanin and highly purified phycobiliproteins were developed. The
former two products are already in the market (Wu et al., 1993; Wu and
Xiang, 1996; Xiang et al., 1995).
Lower cost of production by the use of a low-cost medium could be
a key factor in developing a competitive process for the production of
Spirulina as a feed and as a source of added-value products.
Development of seawater medium for culture of Spirulina is inevitable for
propagating outdoor cultivation of these cyanobacteria in many tropical
arid areas, where climatic conditions are favourable but freshwater is
scarce (Materassi et al., 1980). The use of seawater as an alternative
medium, after being pretreated (Faucher et al., 1979) or after being
supplemented
with
specific
nutrients
(Materassi
et
al., 1980)
or
supplemented with animal waste (Olguin et al., 1994) under laboratory
conditions or in outdoor raceways would reduce the cost of the
production of Spirulina (Tredici et al., 1986; Wu et al., 1993).
The advantages of Spirulina cultivation in seawater medium
(compared to freshwater medium) are (1) lower fertilizer cost, (2) the
medium after harvest can be repeatly used, (3) the pH of the seawater
medium constantly remains at 9- 10 and need not be adjusted, (4)
seawater
culture
is
not
easily
polluted
by
heavy
metals
and
contaminants. Hence, the both growth and quality of S. platensis were
improved in seawater medium (Wu et al., 1993).
The aim of the present study is to develop multi-stress tolerant
mutant Spirulina platensis strain for mass cultivation in low cost
seawater enriched medium. The study was therefore contemplated with
the following objectives.
1. Isolation and characterization of Spirulina isolates from aquatic
habitats.
2. Studies on the effect of temperature, pH, light intensity and
salinity on the growth of Spirulina platensis.
3. Collection and physico-chemical characterization of seawater, to
formulate new low-cost seawater enriched medium with various
nutrients.
4. Improvement of stress tolerant Spirulina platensis strains using
chemical mutagen, Ethyl Methane Sulfonate (EMS) and selection of
multi stress (salinity and temperature) tolerant mutant strains.
5. Studies on growth parameters and biochemical (protein, lipid,
chlorophyll-a, total phycobiliproteins and carotenoid) contents of
mutant strains compared to wild strains in the seawater medium
under laboratory condition.
6. Studies on identification of biochemical constituents in mutant
Spirulina platensis strains by TLC, HPLC methods and their
antioxidant activities.
7. Mass production of multi-stress tolerant Spirulina platensis
mutant strain in seawater enriched medium during summer
season in outdoor artificial cement tank.