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Indian Journal of Geo Marine Sciences Vol. 46 (09), September 2017, pp. 1731-1742 Review Paper Cultivation and harvesting of micro-algae for bio-fuel Production A review R. Dineshkumar, R. Narendran & P. Sampathkumar* Centre of Advanced Study in Marine Biology, Faculty of Marine Sciences Annamalai University, Parangipettai-608 502, India. [Email: [email protected]] Received 08 December 2015 ; revised 30 October 2016 Although micro-algal biomass can be ‘energy rich’, the growth of algae in dilute suspension at around 0.02–0.05 % dry solids poses considerable challenges in achieving a viable energy balance in micro-algal biofuel process operations. Similarity in density of the algal cells to the growth medium, the negative surface charge on the algae and the algal growth rates which require frequent harvesting compared to terrestrial plants. Micro-algae are harvested by a number flocculation, flotation, centrifugation and filtration or a combination of any of these. This paper reviews the various methods of harvesting and dewatering micro-algae for the production of biofuel. There appears to be no one method or combination of harvesting methods suited to all micro-algae and harvesting method will have a considerable influence on the design and operation of both upstream and downstream processes in an overall micro-algal biofuel production process. [Keywords: Microalgae, biodiesel, transestrification, renewable energy] Introduction Deficiency of fossil fuels may result in global famine as technological development, including modern agriculture advance, is predominantly petrol dependent 1 . Renewable fuel sources are permanently searched for, especially ones suitable for transportation. One of the resources is products originating from living organisms (animals, plants and micro- organisms). Fuels from these sources are called biofuels. The review explores the opportunities for energy products encompassing both fresh and marine habit microalgae. This paper also discusses the variety of algal resources and their environment along with the mass production systems that have been demonstrated for use, as well as algae mass cultivation. Biodiesel The choice of raw material is a critical factor contributing to the final cost of biodiesel and accounts for 50-85% of the total cost of the fuel. Therefore, to minimize the cost of this biofuel, it is important to assess the raw material in terms of yield, quality, and the utilization of the by-products 2 . A positive aspect of the production of biodiesel from microalgae is the area of land needed for production. For example, to supply 50% of the fuel used by the transportation sector using palm oil, which is derived from a plant with a high oil yield per hectare, would require 24% of the total agricultural area available in the country. In contrast, if the oil from microalgae grown in

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Page 1: Cultivation and harvesting of micro-algae for bio-fuel ...nopr.niscair.res.in/bitstream/123456789/42614/1/IJMS 46(9) 1731... · Cultivation and harvesting of micro-algae for bio-fuel

Indian Journal of Geo Marine Sciences

Vol. 46 (09), September 2017, pp. 1731-1742

Review Paper

Cultivation and harvesting of micro-algae for bio-fuel Production –

A review

R. Dineshkumar, R. Narendran & P. Sampathkumar*

Centre of Advanced Study in Marine Biology, Faculty of Marine Sciences Annamalai University, Parangipettai-608 502, India.

[Email: [email protected]]

Received 08 December 2015 ; revised 30 October 2016

Although micro-algal biomass can be ‘energy rich’, the growth of algae in dilute suspension at around 0.02–0.05 % dry

solids poses considerable challenges in achieving a viable energy balance in micro-algal biofuel process operations. Similarity in

density of the algal cells to the growth medium, the negative surface charge on the algae and the algal growth rates which require

frequent harvesting compared to terrestrial plants. Micro-algae are harvested by a number flocculation, flotation, centrifugation

and filtration or a combination of any of these. This paper reviews the various methods of harvesting and dewatering micro-algae

for the production of biofuel. There appears to be no one method or combination of harvesting methods suited to all micro-algae

and harvesting method will have a considerable influence on the design and operation of both upstream and downstream

processes in an overall micro-algal biofuel production process.

[Keywords: Microalgae, biodiesel, transestrification, renewable energy]

Introduction

Deficiency of fossil fuels may result in

global famine as technological development,

including modern agriculture advance, is

predominantly petrol dependent1. Renewable

fuel sources are permanently searched for,

especially ones suitable for transportation. One

of the resources is products originating from

living organisms (animals, plants and micro-

organisms). Fuels from these sources are called

biofuels. The review explores the opportunities

for energy products encompassing both fresh

and marine habit microalgae. This paper also

discusses the variety of algal resources and their

environment along with the mass production

systems that have been demonstrated for use, as

well as algae mass cultivation.

Biodiesel

The choice of raw material is a critical

factor contributing to the final cost of biodiesel

and accounts for 50-85% of the total cost of the

fuel. Therefore, to minimize the cost of this

biofuel, it is important to assess the raw material

in terms of yield, quality, and the utilization of

the by-products2. A positive aspect of the

production of biodiesel from microalgae is the

area of land needed for production. For example,

to supply 50% of the fuel used by the

transportation sector using palm oil, which is

derived from a plant with a high oil yield per

hectare, would require 24% of the total

agricultural area available in the country. In

contrast, if the oil from microalgae grown in

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INDIAN J. MAR. SCI., VOL. 46, NO. 09, SEPTEMBER 2017

photo bioreactors was used, it would require

only 1-3% of the total cultivation area3.

Biochemical composition of the algal

biomass can be manipulated through variations

in the growth conditions, which can significantly

alter the oil content and composition of the

micro organism. Biodiesel produced from

microalgae has a fatty acid composition (14 to

22 carbon atoms) that is similar to the vegetable

oils used for biodiesel production4. Biodiesel

produced from microalgae contains unsaturated

fatty acid and is composed of a mixture of

microalgae genera, Chlorella sp., Euglena sp.,

Spirogyra sp., Scenedesmus sp., Desmodesmus

sp., Pseudokirchneriella sp., Phormidium sp.

(cyanobacteria), and Nitzschia it can exhibit

various fatty acid profiles.

Introduction of algae as biodiesel feedstock

Microalgae are prokaryotic or

eukaryotic photosynthetic microorganisms that

can grow rapidly and live in extreme alkaline

conditions due to their unicellular or simple

multicellular structure. Examples of prokaryotic

microalgae are Cyanobacteria (Cyanophyceae)

and eukaryotic microalgae are green algae

(Chlorophyta) and diatoms (Bacillariophyta)5.

They are, not just aquatic but also terrestrial,

representing a big variety of species living in a

wide range of environmental conditions. It is

estimated that more than 50,000 species exist,

but only a limited number, of around 30,000,

have been studied and analyzed. Previous

studies reported that, in order to not compete

with edible vegetable oils, the low cost and

profitable biodiesel should be produced from

low cost feed stocks such as non edible oils,

used frying oils, animal fats, soap stocks and

greases6.

However, the available quantities of

waste oils and animal fats are not enough to

match the demands for biodiesel. Thus transition

to second generation biofuels, such as

microalgae, can also contribute to a reduction in

land requirements due to their presumed higher

energy yields per hectare as well as to their non

requirement of agricultural land. Additionally,

biodiesel needs to have lower environmental

impacts and ensure the same level of

performance of existing fuels. It is still on its

infancy despite it’s the growing interest. A large

investment in research and development and

correct policies and strategies are still needed,

for all stages of the biofuels value chain, from

raw materials production to delivery and final

consumption. Among the various possibilities,

currently being investigated and implemented at

pilot scale or even at industrial scale concerns

potential feed stocks, as well as the potential of

the microalgae. Since microalgae cultivation is

not directly linked to human consumption, they

should have a low space requirement for its

production.7 have described that the microalgae

reproduce using photosynthesis to convert sun

energy into chemical energy, completing an

entire growth cycle every few days.

Moreover, they can grow almost

anywhere, requiring sunlight and some simple

nutrients, although the growth rates can be

accelerated by the addition of specific nutrients

and sufficient aeration8; Different microalgae

species can be adapted to live in a variety of

environmental conditions. Thus, it is possible to

find species best suited to local environments or

specific growth characteristics, which is not

possible to do with other current biodiesel feed

stocks (e.g. soybean, rapeseed, sunflower and

palm oil). They have much higher growth rates

and productivity when compared to conventional

forestry, agricultural crops, and other aquatic

plants, requiring much less land area than other

biodiesel feed stocks of agricultural origin, up to

49 or 132 times less when compared to rapeseed

or soybean crops, for a 30% (w/w) of oil content

in algae biomass 9. Therefore, the competition

for arable soil with other crops, in particular for

human consumption, is greatly reduced by

microalgae cultivation. 10

suggested that genetics

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DINESHKUMAR et al.: CULTIVATION AND HARVESTING OF MICRO-ALGAE– A REVIEW

is one of the factors that affect the microalgae

lipid and biomass productivity. Thus, it is

important to choose species that have the

potential for commercialization. Chlorella

achieved the highest average lipid and biomass

yield among the tested algae, specifically

Chlorella emersonii and C. protothecoides.

Although C. vulgaris and C. minutissima are

capable of producing high lipid content

however, the triglycerides content is low, thus

these species are inefficient as biodiesel feed

stocks 11

.

Microalgae can provide feedstock for

several different types of renewable fuels such

as biodiesel, methane, hydrogen, ethanol, etc.

Algae biodiesel contains no sulfur and performs

as well as petroleum diesel, while reducing

emissions of particulate matter, CO,

hydrocarbons, and SOx. 10,12

reported that for the

past 50 years, extensive research has been

performed on microalgae and how they can be

used in a wide variety of processes to

manufacture many practical and economically

important products. The first large scale culture

of microalgae started in the early 1960s in Japan

by Nihon Chlorella with the culture of

Chlorella. Interest in using microalgae for

renewable energy increased in 1970s during the

first oil crisis (Fig 1).

Fig. 1: Potential fuels from microalgae.

Isolation of algae

2Studied the isolation of pure culture of

green algae from the soil. 13

obtained pure culture

of algae by washing, centrifugation by

antibiotics by dilution and plating by UV

irradiation or by combination of all these. There

are four major techniques for obtaining unialgal

isolates are streaking, spreading, serial dilution

and single cell isolate. Studied the Nile red

staining is used to screen the microalgae for the

presence of lipids droplets in the cells14

. Niles

red can be applied to algal cell and it does not

dissolve in the lipids. 15

Stated that the first to

isolate free living Chlorella and Scenedesmus in

allegedly pure culture of other algae, including

Cyanobacteria and Diatoms.

Cyanobacteria and other algae have a

wide range of pigmented compounds, including

carotenoids, chlorophyll and phycobiliproteins

which consist of timer with two subunits (a, b)

of pigmented poly peptide16

. Some species of

microalgae such as Spirulina standout in this

context by presenting biomass with excellent

nutritional characteristics. The most group of

algae are obligatory in other word they are

entirely depended on their photosynthetic

apparatus for their metabolic needs using

sunlight as energy source and CO2 as carbon

source to produce the carbohydrate and ATP17

.

Spirulina is a spiral filament generally found in

fresh water consists of 60%-70% protein in dry

weight. Protein elements consist of 18 types of

amino acid, several vitamins such as vitamin A,

B, E and K, minerals and fatty acids necessary to

the body. Blue green algae S.maxima were

maintained in the zarrouk medium a standard

synthetic medium, containing 2.5 g/l sodium

nitrate as the nitrogen source (Zerrouk. 1966).

Scenedesmus is one of the most

common freshwater genera; however, the

extremely diverse morphologies found among

species make identification and understanding of

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INDIAN J. MAR. SCI., VOL. 46, NO. 09, SEPTEMBER 2017

their mechanisms difficult. While most species

are found across the world, certain species exist

only in local populations such as S. intermedius

and S. serratus. The formation of colonies as

opposed to unicellularity is dependent on a

number of factors. A higher proportion of

unicellular organisms were found at high light

intensities and high temperatures, suggesting

that at higher growth rates these organisms

prefer to be non-colonized (Fig 2).

Fig. 2: Taxonomy of some algae species potentially useful

for biodiesel production.

Growth Requirements of Microalgae

The biochemical composition of

microalgae can be affected by such factors as

growth rate, environmental conditions, and life

cycle18

. Microalgal growth and chemical

composition are mainly controlled by light,

temperature, available carbon dioxide, pH, and

nutrients (Other factors, such as salinity, can be

of vital importance to some species. Growth

medium must provide sufficient nutrients for

microalgal growth. 19

found that the quality and

quantity of lipids within the cells can vary as a

result of changes in growth conditions

(temperature and light intensity) or nutrient

media characteristics (concentration of nitrogen,

phosphates and iron). 20

reported that while

many microalgae strains naturally have high

lipid content (20-70% on dry weight basis), it is

possible to increase the concentration by

optimizing the growth determining factor such

as the control of nitrogen level light intensity,

temperature and salinity21

, CO2 concentration

and harvesting procedure22

had studied that not

only organic carbon or substrate (a carbon

source such as sugars, proteins and fats),

vitamins, salts and other nutrients (nitrogen and

phosphorous) are vital for algal growth, but also

equilibrium between operational parameters

(oxygen, CO2, pH, temperature, light intensity,

product and by product removal). CO2 is also

involved in the regulation of pH and carbon

balance. Some algae are capable of growing in

extreme conditions of pH. Studies revealed the

effect of changes in pH alters on lipid

metabolism in non-extremophiles. Alkaline pH

stress resulted in TAG accumulation and a

decrease in membrane lipid in Chlorella spp.

Light

The use of monocultures is required in various

microalgal applications and controlled

cultivation systems. This requirement has

favored the development of closed photo

bioreactors. Efficient utilization of light is one of

the major challenges in microalgal

biotechnology, especially when an increase in

the biomass yield is desired22

. It has been

recognized that environmental factors such as

temperature, light, nutrients, affect the lipid

composition of algae23

. Some researchers used

continuous light for culturing

Botryococcusbraunii, 24

as well as Chlorella sp. 25

other researchers applied dark and light

photoperiod for growing Chlorella sp. To

provide dark and light photoperiod in the

system, air lift bioreactor stirred tank photo

bioreactor and other types or photo bioreactor

were employed.

pH

The pH of the culture medium is one of

the important factors in algal cultivation. pH of

medium for algae growth is usually neutral or

slightly acidic, mainly to avoid precipitation of

several major elements. The algae exhibit a clear

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DINESHKUMAR et al.: CULTIVATION AND HARVESTING OF MICRO-ALGAE– A REVIEW

dependency on pH of the growth medium and

different species vary greatly in their response to

pH26

. The pH of algal cultures can be influenced

by various factors such as composition and

buffering capacity of the medium, amount of

CO2 dissolved temperature (which controls CO2

solubility) and metabolic activity of the algal

cells. pH increases during the day to values up to

10, mainly because of the depletion of the anions

NO3- and of CO2 formed in the medium and the

excretion of OH- ions

27. One of the common

effects observed in culture media having pH

values above 9, is the precipitation of several

calcium salts, i.e. carbonates, phosphates and

sulphates, leading to nutrient deficiencies and

growth retardations or even algal flocculation,

induced by the precipitating minerals. Optimum

pH for most cultured algal species ranges

between 7 and 9, with the optimum pH being

8.2-8.7. Complete culture collapse due to the

disruption of many cellular processes can result

from failure to maintain an acceptable pH. In the

case of high intensity algal culture, the addition

of carbon dioxide allows correction for

increased pH, which may reach limiting values

of pH during algal growth 28

.

Temperature

The optimum temperature to grow

Scenedesmus sp. is between 20-40°C (Sanchez

et al., 2008). Christov et al. (2001) studied

Scenedesmus sp. at temperatures of 15 to 36°C

and found at lower temperatures, the chlorophyll

and protein levels were reduced, while levels of

carotenoids, saccharides, and lipid were

increased. They also observed an increase of

30% of the sugars and lipids at extreme

temperatures (36°C).

Scientists are looking for algae that

show good thermo stability at high temperatures.

They have interest in the effect of growth

temperature on chemical and fatty acid

compositions of tropical microalgae

species29

.The effect of temperature exerts on

biochemical reactions and how it affects the

biochemical composition of algae, makes

temperature one of the most important

environmental factors 20

.

Nutrients

Nutrients are inorganic or organic

compounds other than carbon dioxide and water,

used for growth whose presence in the cell is

necessary for cellular function. Some algae

require specific organic compounds synthesized

by other organisms. However, many algae

require only inorganic nutrients, and it is likely

that these algae could be used as feedstock for

biomass fuel production. Limiting nutrients to

algae are nitrogen, phosphorus, silica (for

diatoms) and iron 30

.

Carbon sources

As for microalgae grown on the

heterotrophic mode, experiments using diverse

sources such as molasses, acetic acid and

hydrocarbons (such as n-heptadecane) have been

evaluated and shown to work at different

concentrations. To determine the availability of

using microalgae as carbon di-oxide

sequestration option, CO2 fixation rates of the

test microalgae were studied. The maximum

CO2 fixation rate was shown by mixed algae

sample which could be the combined result of

all the algae species present in the sample 31

.

The increased lipid accumulation has

been observed during the stationary stages of

growth, which is in accordance with the earlier

studies 32

. Nitrogen limitation result in lower

photosynthetic carbon fixation for protein

synthesis evident from the carbohydrate to

amide ratio, consequently stocking carbon in the

form of lipid or carbohydrates, depending on the

species 33

.

Algae growths in waste water treatment

ponds contribute mainly through dissolved

oxygen production and nutrient assimilation and

there are already being used by many waste

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INDIAN J. MAR. SCI., VOL. 46, NO. 09, SEPTEMBER 2017

water facilities as bioremediation agent. In waste

water stabilization ponds, the algae produce

oxygen from water as a by- product of

photosynthesis. This oxygen is used by the

bacteria and they bio-oxidize the organic

compounds in the waste water. An end product,

carbon di oxide is fixed into cell carbon by the

algae during photosynthesis34

. Mixotrophy is

growth in which CO2 and organic carbon are

assimilated simultaneously and hence, both

respiratory and photosynthetic metabolism have

to operate concurrently 35

.

Nitrogen

After carbon, nitrogen (N) is the most

important nutrient contributing to the production

of biomass. The nitrogen content in the

microalgae biomass can range from 1% to more

than 10% and depending on its supply and

availability, it can vary between different groups

(e.g. low in diatoms) and within a particular

species 36

. Discoloration of the cells is a frequent

response to nitrogen limitation due to a decrease

in chlorophyll content and an increase in

carotenoids, as well as the accumulation of

organic carbon compounds such as

polysaccharides and certain oils like

polyunsaturated fatty acids (PUFAs)37

. Nitrogen

is mostly supplied as nitrate, but often ammonia

and urea are used, both displaying similar

growth rates 38

.

Many microorganisms tend to prefer

ammonia as a nitrogen source, and the

assimilation of either nitrate or ammonia is said

to be related to the pH of the growth media 39

. A

drop on pH can be observed when ammonia is

used as the only source of N, especially during

active growth owing to the release of H+ ions.

An increase in pH is observed when nitrate is

used as the only N source 40

. Ammonia loss due

to volatilization is an important factor to be

considered when deciding whether to supply

either one.

Urea can be considered another

attractive source of nitrogen 41

reported that the

microalgae fed with a combination of urea and

sodium nitrate had the highest ash-free dry

biomass content with a yield of 4.15 ± 0.38 g

L⁻1. The urea molecule contains a carbon atom

as well as two nitrogen atoms. This carbon atom

is released as CO2 when urea is utilized and

presumably, it is available for photosynthetic

assimilation. Therefore, urea has the potential of

providing both the nitrogen and 1.5 to 10 % of

the carbon requirement10

reported the

heterotrophic cultivation of Chlorella

protothecoides in various N sources for lipid

production.

The lipid content, lipid class

composition and the proportion of the various

fatty acids in a microalgae vary according to the

environmental or culturing variables such as

light intensity growth phase photoperiod and

nitrogen concentration. Most critical nutrient

affecting lipid metabolism in microalgae is

nitrogen limitation42

, nitrogen limitation leads to

a decrease in protein content in both freshwater

algae and diatom. Cell size for nutrient limited

cultures were significantly smaller than the non

–limited cells43

. Previous studies have

demonstrated that lipid content in some

microalgae increase during different cultivation

conditions such as nitrogen deprivation.

Phosphorus

Phosphorus is essential for growth in many

cellular processes such as energy transfer and

during the biosynthesis of nucleic acids.

Orthophosphate is the preferred form in which it

is provided to algae and its uptake is said to be

energy dependant 44

. In spite of the fact that

algal biomass contains less than 1% phosphorus

(P), it usually becomes one of the most

important growths limiting factors in algal

culture 45

. This happens because P binds easily

to other ions (e.g. carbonate and iron) resulting

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DINESHKUMAR et al.: CULTIVATION AND HARVESTING OF MICRO-ALGAE– A REVIEW

in its precipitation. Insolubility of the resulting

phosphate makes this essential nutrient

unavailable for algal uptake46

. Supply of P also

influences the composition of the produced

biomass. Similar effects to the ones obtained in

algae grown under nitrogen starvation, such as

the tendency to accumulate large amount of

lipids, with a decreased amount of proteins,

chlorophyll and nucleic acids content have been

reported on phosphate deficient cultures47, 48

,

reported that the very high concentration of cell

lipid tend to be coupled with low biomass

particularly induced by environmental stress

such as N and P limitation.

Other macro and micronutrients

Sulfur, potassium, sodium, iron,

magnesium, calcium and trace elements like

boron, copper, manganese, zinc, molybdenum,

cobalt, vanadium, and selenium are also

important in algal nutrition 49

. Silicon, present in

the cell walls of many algal groups especially in

diatoms, is an important component where it

constitutes an essential nutrient for their growth

and production. Silicon limitation, which is

prone to happen, can also lead to the

accumulation of secondary metabolites, such as

lipids. Experiments conducted with silicon-

deficient cells of Cyclotella cryptic (a diatom

species that accumulates lipids under non-

growing conditions) indicated that lipid

accumulation occurs as a function of both

increased partitioning of newly photo-

assimilated carbon into lipids and slow

conversion of non-lipid compounds.

Utilization of sewage by algal sp

It was reported that the algal growth

rates and nutrient uptake were found to be high

and equivalent to value from algae grown on

municipal wastewater51

. It is defined that

Chlorella minutissima which was identified in

waste water oxidation ponds in India52

. Waste

water was shown to be low enough in toxins and

had enough P and N to support algal growth of

microalgae, Chlorella saccharophila able to

grow particularly well on the untreated waste

water50

. They also reported that the potential for

some industrial waste waters in providing

resources for the generation of significant algal

biomass came from the analysis of waste water

from carpet mill effluent. The earlier researcher

suggested that a more practical approach for

algae biodiesel production is to utilize

wastewater for algae propagation 34

.

The use of waste water for algal and

biofuel production would ensure sustainability in

terms of (a) biofuel production (b) nutrient

removal and remediation of wastewater and (c)

mitigation of GHGs. Thus, the development of

renewable clean algal biofuel fosters sustain-

ability and helps in maintaining the

environmental quality. Waste water which is

produced from various industries can also be a

source of feed for the algae to grow and hence,

play a dual faction i.e., nutrient removal and

biofuel production. The effluent from the

secondary treatment unit of domestic waste

water treatment plant is used for large scale

cultivation of microalgae for biofuel production.

The best suitable method for cultivation of algae

in the waste water is open pond way reactor for

biofuel production 23

.

The effluent from the secondary

treatment unit of domestic waste water treatment

plant is used for large scale cultivation of

microalgae for biofuel production and observed

the inhibition in the growth of microalgae at the

initial stage due the presence of high nutrient

and the other factors present in sewage as

nutrient medium. Various sp of Chlorella and

Scenedesmus can provide very high (>80%) and

many cases almost complete removal of

ammonia, Nitrate and total P from secondary

treatment waste water indicating the potential of

microalgae for tertiary sewage treatment. Many

of these experiments were performed under

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INDIAN J. MAR. SCI., VOL. 46, NO. 09, SEPTEMBER 2017

laboratory based batch culture condition with the

microalgae showing high growth rates over the

batch growth period. Some unicellular green

microalgae species are particularly tolerant to

sewage effluent condition; most notably those of

the Chlorella and Scenedesmus genus and so

most studies have examined the growth of these

species52

. Microalgae are effectively able to

grow in wastewater conditions through their

ability to utilize abundant organic carbon and

inorganic N and P in the waste water.

It is stated that the Chlorella vulgaris

immobilized in alginate pellets placed in a

fluidized bed column could effectively remove

approximately 80% of the ammonia content and

70% of the total P contentment in a sewage

culture35

. It is reported that Scenedesmus

immobilized in alginate sheets could effectively

remove ammonia and orthophosphate from

secondary treated effluent54

. Algae species

Chlorella was widely applied for waste water

treatment and had proven abilities of removing

N, P and chemical oxygen demand (COD) with

different retention times ranging from 10 to 40

days. Further same researches reported that 55%

P removal from agro industrial waste water by

Chlorella. 45

Stated that the algae based

treatments have been found to be as efficient at

removing P from waste water as compared to

chemical treatment. 55

Studied the Chlorella

kesslerii able to uptake only 8-20% of P under

the light and dark cycle for PO4 concentration of

10mg. 56

Reported that recovered N and P rich

algal biomass can be used as a low cost fertilizer

or as animal feed. Activated algal process to

treat waste water and found that it was able to

remove 80-88% of BOD, 70-82% of COD, 60-

70% of N and 50-60% of P with a retention

period of 15 days56,57

reported that the growth

potential and the N and P scavenging ability was

examined in micro algal strains grown under

secondary treated sewage effluent.

Collection of biomass

Harvesting, thickening and dewatering is a key

step in the production of algal biofuel is

harvesting and dewatering of the algal biomass

before extraction. The high cost of harvesting

and dewatering presents major challenges to the

development of commercially viable

microalgae-based biofuel. It is reported that for a

process to be suitable for very large volumes of

biomass 47

. As microalgae vary greatly in those

properties, which affect harvesting processes

(e.g., size, surface charge, resistance of cell to

breakage, compressibility etc.) between species

and with growth phase the biomass recovery

processes must be tailored to the species of

microalgae and the growth system. As very large

amounts of water are used in microalgae culture

the medium which still contains nutrients, must

also be able to be recycled after cell harvesting.

The harvesting, thickening and

dewatering of microalgae cultures have been

extensively reviewed 18, 29, 36

. Key properties of

microalgae which influence their separation are:

(a) shape: [rods, spheres or chains or filaments],

(b) size: [generally between 2 and 30 μm], (c)

specific weight: [1.05–1.1], (d) surface charge:

[usually negative]. Microalgal cultures to be

harvested usually contain between 0.2 to 2 g L−1

solids and for lipid extraction a concentration of

at least 20 g L−1

solids is required. Filamentous

algae such as Spirulina can be harvested by

filtration, but almost all of the algae under

consideration as a source of biofuels (e.g.,

Nannochloropsis or Chlorella) are unicellular

and too small for effective filtration.

Centrifugation is very energy intensive (Mohn,

1988) and not practical for the large volumes for

algal biofuel production. Sedimentation is also a

possibility, but it is time-consuming .

The most commonly considered

processes are flocculation followed by flotation

or by settling as the first step. Flocculation is the

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DINESHKUMAR et al.: CULTIVATION AND HARVESTING OF MICRO-ALGAE– A REVIEW

first stage in the bulk harvesting process to

aggregate the cells, so increasing their effective

particle size and thus easing subsequent

centrifugation, filtration or sedimentation steps.

Flocculation can be achieved by the use of

inorganic flocculants such as alum 19

or organic

flocculants such as or starch, although the cost

of these flocculants is substantial. The flocculant

used must be compatible with the need to

recycle the water back to the growth system

without complex pretreatment of this recycled

water.

Lipid extraction

Lipids are mostly hydrophobic

molecules (e.g., neutral lipids) which interact

with relatively non-polar solvents such as ethyl

ether, chloroform and benzene, while membrane

associated polar lipids require polar solvents

such as ethanol and methanol to disrupt the

hydrogen bonding and electrostatic forces

between the lipids and proteins 59

studied that

the most common laboratory methods for lipid

extraction are the Soxhlet extraction (usually

with n-hexane as solvent), and the Folch, Bligh

and Dyer methods which use chloroform and

methanol in varying ratios as solvents. Because

of the toxicity of methanol and chloroform and

the fact that chloroform extracts more than just

saponifiable lipids (e.g., pigments and other

lipids and non-lipid contaminants), methods

using other alcohols such as ethanol, 1-butanol

and isopropanol are being developed to replace

the methanol. Other combinations of co-solvents

also have been proposed for the extractions of

lipids from microalgae are hexane/ethanol, and

hexane/ isopropanol. 60

It is also studied that the

hexane system, the hexane and alcohol will

readily separate into two separate phases when

water is added, thereby improving downstream

separations. Furthermore, the combination of

ethanol and hexane at a 5:1 v/v ratio in a 2-step

extraction procedure reduces the solvent

requirement for the same amount of biomass by

approximately 10-times compared to the

chloroform/methanol extraction procedure.

Conclusion

In spite of many advantages of biodiesel

production from algae, there are a lot of

limitations blocking its real competition with

petroleum derived diesel. Production costs seem

the most substantial problem. Out of all

recognized methods of microalgae production,

the culture in photo bioreactors seems the most

favorable for biodiesel production, however the

costs are discouraging. Investments cost of

whole plant is one of the major limiting factors.

It seems that small modular systems and outdoor

Raceway pond cultivation system may be an

attractive solution with positive economical

rationale. Algae production plant should also be

integrated with other existing installation like

sewage treatment, biogas and power

cogeneration plant. Co-localization these

production processes results in lowered

operating and investments costs. Also, such a

waste treatment and the biofuel-energy centre

generate a number of additional by-products that

can be sold on or used for self-consumed.

The most energy-efficient method of

harvesting or of producing feasible energy from

micro-algae may depend on harvesting and oil

extraction method. Major challenge of

commercializing micro-algal biofuel has a

considerable influence on the design and

operation of both upstream and downstream

processes in an overall micro-algal biofuel

production process.

Algae derived biodiesel is ecofriendly

which in the case of petroleum shortage may

replace petro diesel. However, the biodiesal

derived from algae is very cost-effective.

Presences of alternative technologies of fuels

production are optimistic in the face of

discussion about petroleum stock out.

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INDIAN J. MAR. SCI., VOL. 46, NO. 09, SEPTEMBER 2017

Acknowledgements

Authors are thankful to Dean and

Director, Centre of Advanced Study in Marine

Biology, Faculty of Marine Sciences, Annamalai

University, Parangipettai, India for providing

necessary lab facilities. The first author is

grateful to the University Grants Commission –

Rajiv Gandhi National Fellowship, Government

of India for the financial assistance (Name of the

Awardee: R. Dineshkumar) with UGC Circular

No. F1-17.1/2015-16/RGNF-2015-17-SC-TAM-

4190 in January, 2016.

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