FINAL PROJECT REPORT

Algae Based Wastewater Treatment
AISSMS COE, PUNE. Page 1
ABSTRACT
In 1992 World Health Organization studies claimed to have reported that out of
India's 3,119 towns and cities, just 209 have partial sewage treatment facilities, and
only 8 have full wastewater treatment facilities. According to another 2005 report,
sewage discharged from cities and towns is the predominant cause of water pollution
in India. Investment is needed to bridge the gap between 29000 million litres per day
of sewage India generates, and a treatment capacity of mere 6000 million litres per
day. A large number of Indian rivers are severely polluted due to disposal of domestic
waste.
The wastewater is directly discharged without treatment into the water bodies are
causing environmental problems also affecting the health of human beings and it will
create the environmental inbalance in aquatic life .Therefore the necessity is arises to
remove the nutrients and other water pollutant parameters.
For removing nutrients from the waste water would be helpful for growing the aquatic
plant as well as being helpful for removing water pollutants. Therefore this study is
highly emphasizes on biological treatment over conventional treatment and focuses a
short review on the current scenario in the cultivation of microalgae in wastewater for
nutrient removal.
The objective of this study is to evaluate the effect of Chlorella
species(C.Vulgaries) and Scenedesmus (S.Quadriquada) species on wastewater
collected from the primary treatment process of Municipal Wastewater Treatment
Plant (MWTP). . The suitability of algae cultivation over a conventional wastewater
treatment process flow can be checked by removing nitrates, phosphates, chemical
oxygen demand (COD),Biological oxygen demand(BOD) etc. from the wastewater
In future the removed nutrients can be utilized for cattle feed ,fertilizers and
biodiesel production.

CHAPTER – 1
INTRODUCTION

CHAPTER 1
INTRODUCTION
1.1 introduction to wastewater
Wastewater is a general term used to represent the water with poor quality that
contains more amounts of pollutants and microbes. If wastewater is discharged into the
nearby water bodies, it can cause serious environmental and health problems to human
beings. Wastewater treatment is an important measure to reduce the pollutant and
other contaminants present in wastewater. The first step in wastewater treatment
method is primary treatment which removes the solids, oil, and grease from
wastewater. Secondary treatment or biological treatment is the second step, which
exploits microorganisms to eliminate the chemicals present in wastewater. Final step is
the tertiary treatment; which eliminates the microbes from wastewater before
discharging into the river.
1.2 Types of wastewater
o Domestic Waste Water
o Industrial Waste Water
o Storm Water
1.3 Source of wastewater
o Domestic waste water is generated from domestic as well as
commercial areas.
o Industrial waste water is generated from industrial areas and factories.
o Storm water is nothing but the run – off resulting from the rain storms.

1.4 Treatment of wastewater by conventional method
Fig no .1.1
1.5 Fit falls of conventional treatment
The major disadvantages associated with current wastewater treatment practices are:
 Many wastewater treatment processes generate large amounts of sludge that
must be sent off-site for disposal. Handling and disposal of this sludge is
typically the largest single cost component in the operation of a wastewater
treatment plant.
PRELIMINARY
• Removal of wastewater costituents such as rags,sticks,flotables,grit and
grease and they cause maintainance or operational problems
PRIMARY
• Removal of suspended solids and organic matter or enhanced removed
typically accomplished by chemical addition or filteration.
SECONDARY
• Removal of biodegradable organic matter and suspended solids .
Removal of nutrients such as nitrogen, phosphours or both may be
coupled here.
TERTIARY
• Removal of residual suspended solids after secondary treatment usually
by granular medium filtration. Disinfection is also typically a part of this
step.Nutrient removal is often included in this definition.

 Most wastewater treatment processes cannot effectively respond to diurnal,
seasonal, or long-term variations in the composition of wastewater. A
treatment process that may be effective in treating wastewater during one time
of the year may not be as effective at treating wastewater during another time
of the year.
 High energy requirements will make many wastewater treatment methods
unsuitable for low per-capita energy consumption countries.
 High operation and maintenance requirements, including production of large
volumes of sludge (solid waste material), make them economically unviable
for many regions.
1.6 Introduction to algae
Definition of algae
Algae are a large and diverse group of simple aquatic organisms ranging from
unicellular to multi-cellular forms and they mainly grow based on the photosynthesis
mechanism, just like the plants.
Figure 1.2

Figure 1.3
Algae and Wastewater

1.7 Algae-based Wastewater Treatment vs. Conventional Methods
Using algae for wastewater treatment offers some interesting advantages over
conventional wastewater treatment.
Advantages of algae based wastewater treatment -
 Cost effective
 Low energy requirement
 Reductions in sludge formation
 GHG emission reduction
 Production of useful algal biomass
1. Cost Effective - It is more cost effective way to remove biochemical oxygen
demand, pathogens, phosphorus and nitrogen than activated sludge process and other
secondary treatment processes (Green et al., 1996).
2. Low Energy Requirements - Traditional wastewater treatment processes involve
the high energy costs of mechanical aeration to provide oxygen to aerobic bacteria to
consume the organic compounds in the wastewater, whereas in algae based wastewater
treatment, algae provides the oxygen for aerobic bacteria. Aeration is an energy
intensive process, accounting for 45 to 75% of a wastewater treatment plant’s total
energy costs. Algae provide an efficient way to consume nutrients and provide the
aerobic bacteria with the needed oxygen through photosynthesis. Roughly one kg of
BOD removed in an activated sludge process requires one kWh of electricity for
aeration, which produces one kg of fossil CO2 from power generation (Oswald, 2003).
By contrast, one kg of BOD removed by photosynthetic oxygenation requires no
energy inputs and produces enough algal biomass to generate methane that can
produce one kWh of electric power (Oswald, 2003).

3. Reductions in Sludge Formation - In conventional wastewater treatment systems
the main aim is to minimize or eliminate the sludge. Industrial effluents are
conventionally treated using a variety of hazardous chemicals for pH correction,
sludge removal, colour removal and odour removal. Extensive use of chemicals for
effluent treatment results in huge amounts of sludge which forms the so called
hazardous solid waste generated by the industry and finally disposed by depositing
them in landfills. In algae wastewater treatment facilities, the resulting sludge with
algal biomass is energy rich which can be further processed to make biofuel or other
valuable products such as fertilizers. Algal technology avoids use of chemicals and the
whole process of effluent treatment is simplified. There is considerable reduction in
sludge formation.
4. The GHG Emission Reduction – The US Environmental Protection Agency (EPA)
has specifically identified conventional wastewater treatment plants as major
contributors to greenhouse gases. Algae based wastewater treatment also releases CO2
but the algae consume more CO2 while growing than that is being released by the
plant, this makes the entire system carbon negative.
5. Production of Useful Algal Biomass – The resulting algae biomass is a source of
useful products such as biodiesel. Previous research in the early 1990’s by the
National Renewable Energy Laboratory (NREL) showed that under controlled
conditions algae are capable of producing 40 times the amount of oil for biodiesel per
unit area of land, compared to terrestrial oilseed crops such as soy and canola.

1.8 Algae based wastewater treatment
Figure 1.4
1.9 Objectives of project
 It aims to check the efficiency of microalgae strains in removal inorganic
nutrient to prevent further deterioration of wastewater quality of domestic
wastewater.
 Present investigation focuses on the bioremediation of wastewater by
developing culture of C. vulgaris and S. quadricauda microbes.
Primary TreatmentWastewater
Secondary Treatment
or
Conventional
system
Algae based system
Tertiary treatment Final effluent

 To study the role of microalgae in wastewater, the following protocols are
used-
 Wastewater treated with culture of C. vulgaris and S. quadricauda microbes,
and compare with conventional method.
 Samples are periodically (every 5th day) analyzed for physico-chemical
parameters such as pH, phosphate, nitrate, BOD and COD etc. using standard
method.
1.10 Scope of the project
 The world is facing problems with a wide variety of pollutants and
contaminates from various developmental activities. The population explosion
in the world has resulted in an increase in the area of polluted water.
 The concern on the quantity and quality of waste generated and discharged into
natural water bodies has recently indicated the need for different strategies to
address water quality challenges in the regions.
 Bioremediation uses naturally occurring microorganisms and other aspects of
the natural environment to treat wastewater of its nutrients. Bioremediation can
prove less expensive than other technologies that are used for cleanup of
hazardous waste.
 Algae are universally acknowledged as playing a very important role in natural
water purification process.
 Thus, the use of microalgae for removal of nutrients from different wastes has
been described by a number of authors.

CHAPTER - 2
LITERATURE REVIEW

CHAPTER 2
LITERATURE REVIEW
2.1 Present scenario
The world is facing problems with a wide variety of pollutants and contaminates from
various developmental activities. The population explosion in the world has resulted in
an increase in the area of polluted water. The concern on the quantity and quality of
waste generated and discharged into natural water bodies has recently indicated the
need for different strategies to address water quality challenges in the regions.
Bioremediation uses naturally occurring microorganisms and other aspects of the
natural environment to treat wastewater of its nutrients. Bioremediation can prove less
expensive than other technologies that are used for cleanup of hazardous waste.
Algae are universally acknowledged as playing a very important role in natural water
purification process.
Thus, the use of microalgae for removal of nutrients from different wastes has been
described by a number of authors such as (Benemann et al., 1977; Gupta and Rao,
1980; Williams, 1981; Kunikane et al., 1984; Senegar and Sharma, 1987; Tam and
Wong, 1989; Gantar et al., 1991; De la Noue, 1992; De-Bashan et al., 2002; Queiroz
et al., 2007; Rao et al., 2011, s.k.birdi, Garg).
2.1.1 Research& Updates
 During the U.S. Department of Energy’s Aquatic Species Programme (ASP), it
was found that for the algae remediation of wastewater, energy outputs were
twice the energy inputs, based on digester gas production and requirements for
pumping the wastewater, mixing the ponds, etc. The overall economics were
very favorable because of the wastewater treatment credits.

 In April 2009, NASA scientists have proposed a project called “Sustainable
Energy for Spaceship Earth.” NASA uses large plastic bags in the ocean, and
fill it with wastewater. The algae use wastewater and solar energy to grow, and
in the process of growing they clean up the sewage. The bag will be made of
semi-permeable membranes that allow fresh water to flow out into the ocean,
while retaining the algae and nutrients. The membranes are called “forward-
osmosis membranes.” NASA is testing these membranes for recycling dirty
water on future long-duration space missions. This project called as OMEGA
(Offshore Membrane Enclosures for Growing Algae) has gained significant
attention recently. OMEGA process aims to investigate the technical feasibility
of a unique floating algae cultivation system and prepare the way for
commercial applications for the production of algae fuels.
 In January 2012, researchers at the California Polytechnic State University
launched a pilot project to test the viability of using algae to treat wastewater.
Nine algae-rich ponds that circulate the waste water are employed to treat the
polluted water. Fueled by sunlight, the algae feed on pollutants in the
wastewater and results in cleaner water and an increased volume of oil-rich
algae that can be converted to products such as liquid biofuel or fertilizer. The
project is funded by a $250,000 grant from the California Energy Commission.

2.2 Case Studies referred are as follows -
Microalgae Cultivation in Wastewater for Nutrient Removal by S. Sriram and R.
Seenivasan
Based on this review, it was concluded that the microalgae alone cannot efficiently
remove the nutrients from the wastewater. The microalgae growth - promoting
bacteria (MGPB), starvation and dilution of the wastewater are the different ways used
to enhance microalgae nutrient removal rate. Microalgae cultivated in the wastewater
can be used for the biodiesel production and as feed for animals. This dual process
(microalgae cultivation in effluent coupled with biodiesel production) has several
advantages such as less cost and less energy input for biodiesel production, and less
greenhouse gas emission during biodiesel production.
Wastewater treatment with microalgae – a literature review by Karin Larsdotter,
Environmental Microbiology, School of Biotechnology, KTH, AlbaNova
University Center, 106 91 Stockholm.
Microalgae can be used for tertiary treatment of wastewater due to their capacity to
assimilate nutrients. The pH increase which is mediated by the growing algae also
induces phosphorus precipitation and ammonia stripping to the air, and may in
addition act disinfecting on the wastewater. Domestic wastewater is ideal for algal
growth since it contains high concentrations of all necessary nutrients. The growth
limiting factor is rather light, especially at higher latitudes. The most important
operational factors for successful wastewater treatment with microalgae are depth,
turbulence and hydraulic retention time.

Purification of waste water using Algal species by Deviram GVNS*,
Pradeep K.V and R Gyana Prasuna Department of Microbiology, Gitam
Institute of Science, GITAM University,Visakhapatnam, India.
The study speculates that there is unlimited scope for using such potential
strains for enhanced activity towards rapid treatment and probable recycling of
waste water for useful purposes.
Evaluating algal growth at different temperatures By Keelin Owen
Cassidy University of Kentucky.
 As this study mentioned, carbon dioxide emission might be the cause of global
warming, and one way to reduce the emission is by algae. Like all living
things, algae needs the correct environment in order for it to perform at its best,
and, for this case, capturing carbon dioxide. From this study, the optimum
temperature for the algae growth was found and a heat transfer model was
developed to see how the temperature of the greenhouse would affect algae
growth.
 In this study, the growth of algae was measured at different temperatures,
showing that as temperature rises the algal growth will increase, reach an
optimum, and then decrease. This type of growth pattern was observed for
Chlorella and Scenedesmus grown on M-8 and urea growth media. The
temperatures tested were 25, 30, and 35°C, where 30°C was considered as an
optimum for both strains. The growth rate was 0.0191 and 0.0235 1/hr for
Scenedesmus grown on urea and M-8 and 0.0292 and 0.017 1/hr for Chlorella
grown on urea and M-8. Chlorella had the best growth rate of 0.0292 1/hr
while grown on urea growth media; however, other studies (Converti et al.
(2009) and Bajguz (2009)) have said it will not grow very well with
temperatures above 30°C. Scenedesmus' growth was more consistent and
favors temperatures ranging from 20-40°C.

 The further testing proved Scenedesmus' optimum temperature is 27°C with a
growth rate of 0.0284 1/hr. The test also proved the growth rate was
statistically different from the other temperatures.
2.3 General advice on operation
Domestic wastewater is very nutrient rich, and basically all nutrients needed for algal
growth are present. The factors limiting algal growth, and hence treatment efficiency
is therefore more likely to be light and carbon. Light is the most important parameter
to optimise, and hence culture depth and turbulence are vital for good performance. To
avoid temperature limitation in northern climate, greenhouses would be recommended
in order to have functioning treatment during longer periods than just during summer.
To increase the performance during winter, artificial light may also be needed,
however that demand extra costs for energy.
The easiest way to start a microalgal wastewater treatment process is to inoculate with
water containing a large variety of algae, e.g. water from outdoor ponds. This will
create a mixture of algae and other organisms, where the best suited species will grow
fastest and dominate the treatment step. Other microalgae will also be introduced
eventually, partly from the wastewater itself, partly from algal particles in the air dust.
This approach requires less supervision and operation than if a particular algae is
chosen to be cultivated for any purpose. A drawback is that the fastest growing
microalgae are most often unicellular green algae (Chlorophyceae) which are
difficult to harvest.
Depth
Depths of between 15 and 50 cm are generally recommended. During winter, however,
shallower depths than 20 cm should not be used to account for the decreased incident
light intensity.

Hydraulic retention time
The hydraulic retention time (HRT) should be long enough to prevent the treatment
step from wash-out effects, i.e. it should not be shorter than the minimum generation
time of the algae (i.e. the dilution rate should not exceed the maximum algal growth
rate, μmax). On the other hand, too long HRT allows the algae to grow slower due to
nutrient limitation and increased internal shading, and should also be avoided. The
effluent concentrations of nitrogen and phosphorus will, on the other hand, be lower at
longer HRTs. Between 2 and 7 days HRT are common in microalgal wastewater
treatment. During winter, longer retention times would probably be necessary than
during summer as a result of the lower growth rate.
Figure 2.1 – Treatment Process.

CHAPTER - 3
METHODOLOGY
AND
INVESTIGATIONS

CHAPTER 3
METHODOLOGY AND INVESTIGATIONS
3.1 Methods of waste water treatment –
Activated sludge process-conventional treatment units –
1 Inlet chamber - in the inlet chamber we receive the sewage.
2 fine screen chamber - fine mechanical screen will help to screen physical
material up to size of 6 mm.
3 grit chamber - in this chamber grit that is heavy suspended solids like fine
sand, wooden particles etc. will settle down. Central rotary arm collects settle
material and move towards grit clarifier.
4 grit clarifier – it will help to remove settle material from grit chamber to
container for dispose as garbage.
5 Primary clarifier - these are circular tanks which help to settle suspended
solids from sewage at bottom.
6 Aeration tank – aeration tank is rectangular tanks containing surface
aerators. These aerators dissolve atmospheric oxygen in to the sewage. In
aeration tank activated micro-organisms are available for degradation of
organic matter which is present in sewage. Dissolved oxygen is required for
proper growth of micro-organism. Overflow of aeration tank is given to
secondary clarifier.
7 Secondary clarifier - Treated sewage from aeration tank contains activated
sludge which is settled in bottom of secondary clarifier.settle activated sludge
again pumped to aeration tank to maintain proper micro-organism strength.
Overflow of secondary clarifier is given to chlorine contact tank by channel.
8 Chlorine contact tank- treated sewage collected from secondary clarifier
chlorinate into tank with help of liquid chlorine and disposed into river.
9 primary and excess sludge slump – settled sludge collected from primary
clarifier and excess activated sludge from secondary clarifier into primary
sump. sump having pumps which are pumping these sludge to thickner for
further sludge treatment.

10 thickener – sludge generated from primary clarifier and secondary clarifier
and secondary clarifier is pumped to thickener for thickening the same.
Thickened sludge shall be conveyed to sludge digester.
11 anaerobic digester- anaerobic sludge digester shall be provided to digest
sludge from sludge thickener. The sludge from the thickener unit is pumped to
the digester. The organic material in sludge under anaerobic condition is
biologically converted to methane carbon dioxide. The stabilized sludge from
the digester is taken to the centrifuge.
12 centrifuge – digested sludge from bottom of digester is pumped to
centrifuge is collected at bottom and used as manure for agriculture purpose.
3.2 Materials and Methods
3.2.1 Microorganism Selection
Figure 3.1

The algal species used in this study were taken from Botany Department ,University
Of Pune.
Chlorella vulgaris and Scenedesmus quadricauda were used as test organisms for the
treatment of domestic wastewater.
Chlorella vulgaris shows great potential for capturing carbon dioxide. It will grow at
a fast rate (0.6 g/L day) and tolerate 10-15% carbon dioxide. Chlorella vulgaris can
also grow in extreme environments, high temperatures of 30-35°C and acidic
environments such as a pH of 3. When it comes to flue gas, it can tolerate up to 200
ppm of NOx and 50 ppm of SOx.
Once the algae is used for carbon dioxide consumption, it can be used in a secondary
process or product such as animal feed. For secondary processes, Chlorella vulgaris
has a high percent of proteins, minerals, and vitamins .
In sewage treatment plants, Scenedesmus takes up CO2 and provides oxygen to
bacteria as it breaks down organic matter. Hence, Scenedesmus is an attractive
candidate for CO2 mitigation with flue gas because it can tolerate being grown in
wastewater. The rate of daily carbon dioxide consumption is 28.08% at a 6% carbon
dioxide level. The temperature in which Scenedesmus will grow ranges from 10 to
40°C .
3.3 Requirements for growth of algae
Carbon and nutrients
Algae are autotrophs, i.e. they can synthesise organic molecules themselves from
inorganic nutrients. A stoichiometric formula for the most common elements in an
average algal cell is C106H181O45N16P, and the elements should be present in these
proportions in the medium for optimal growth. High ratios between nitrogen and
phosphorus, about 30:1, suggest P-limitation, whereas low ratios of about 5:1 suggest
N-limitation. According to the ratios most often found in wastewater, phosphorus is
rarely limiting algal growth, but nitrogen may be. Though, since wastewater often

exposes the algae to nutrient concentrations of up to three orders of magnitude higher
than under natural conditions, growth is more likely limited by carbon and light . The
rate at which an algal cell takes up a specific nutrient depends on the difference
between the concentration inside and outside the cell, and also on the diffusion rates
through the cell wall. The thickness of the unstirred layer of water just outside the cell
wall also plays a role, where thicker layers give slower diffusion rates. To avoid such
thick boundary layers in order to enhance mass transfer rates of nutrients and
metabolites, turbulence in the water is essential.
Carbon
Microalgae assimilate inorganic carbon in the photosynthesis. Solar energy is
converted to chemical energy with oxygen (O2) as a by-product, and in a second step
the chemical energy is used to assimilate carbon dioxide (CO2) and convert it to
sugars. The overall stoichiometric formula for photosynthesis is:
6 H2O + 6 CO2 + light ⇒ 6H12O6 + 6 O2
The inorganic carbon species normally used by microalgae are CO2 and HCO3 –, the
latter requiring the enzyme carbonic anhydrase to convert it to CO2 . Beside these,
some algal species are able to use organic carbon sources as well, such as organic
acids, sugars, acetate or glycerol. This heterotrophic metabolism is probably
significant in waste loaded ponds, where the standing crops of algae can be very high
and consequently exhausted on carbon dioxide . Some studies have indicated that
about 25–50 % of the algal carbon in high rate algal ponds is derived from
heterotrophic utilisation of organic carbon . The organic carbon sources can be
assimilated either chemo- or photoheterotrophically . In the first case, the organic
substrate is used both as the source of energy (through respiration) and as carbon
source, while in the second case, light is the energy source. In several algal species, the
mode of carbon nutrition can be shifted from autotrophy to heterotrophy when the
carbon source is changed; this is the case with e.g. the green algae Chlorella and
Scenedesmus.

Nitrogen
Besides carbon, nitrogen is the second most important nutrient to microalgae since it
may comprise more than 10 % of the biomass . Nitrogen exists in many forms, and the
most common nitrogen compounds assimilated by microalgae are ammonium (NH4 +)
and nitrate (NO3 –) . The preferred compound is ammonium, and when this is
available, no alternative nitrogen sources will be assimilated . However, ammonium
concentrations higher than 20 mg NH4 +-N per litre are not recommended due to
ammonia toxicity. In addition to these nitrogen compounds, urea (CO(NH2)2) and
nitrite (NO2 –) can be used as nitrogen sources. However, the toxicity of nitrite at
higher concentrations makes it less convenient . Cyanobacteria are also able to
assimilate the amino acids arginine, glutamine and asparagine and some species can
fix nitrogen gas (N2) . Of all nitrogen sources, this nitrogen fixation is the most energy
demanding and only occurs in some cyanobacteria when no other nitrogen compounds
are available in sufficient amounts . Several microalgae can take up nitrogen in excess
of the immediate metabolic needs, so called luxury consumption. This can be used
later in the case of nitrogen starvation.
Phosphorus
Phosphorus is another macro-nutrient essential for growth, which is taken up by algae
as inorganic orthophosphate (PO4). The uptake of orthophosphate is an active process
that requires energy. Organic phosphates can be converted to orthophosphates by
phosphate at the cell surface, and this occurs especially when inorganic phosphate is in
short supply. Microalgae are able to assimilate phosphorus in excess, which is stored
within the cells in the form of polyphosphate (volutin) granules. These reserves can be
sufficient for prolonged growth in the absence of available phosphorus. The growth
rate of an algae may therefore not respond at once to changes in the external
concentration of phosphorus, in opposite tothe immediate responses to temperature
and light.

Temperature
Increased temperature enhances algal growth until an optimum temperature is reached.
Further increase in temperature leads to a rapid decline in growth rate. Overheating of
algal cultures is a problem especially in humid climates where evaporation is inhibited,
but in Sweden, the problem is rather growth limitation caused by low temperatures if
cultivating outdoors. At low temperatures, microalgae easily get photo inhibited by
high light intensities. This sensitivity to bright light at low temperatures may pose an
operational constraint on outdoor wastewater treatment in cold climate. At
temperatures near optimum for growth, microalgae can better tolerate high light
intensities before getting inhibited . Generally, temperatures around 15–25ºC seems to
suit most algal species, even those which are adapted to growth at colder temperatures.
To enable higher temperatures in algal cultures, greenhouses may be a solution at
higher latitudes.
pH
Microalgal growth rate and species composition may also be affected by pH. As an
example, Fontes et al (1987) found that optimal productivity of the cyanobacterium
Anabaena variabilis were obtained at pH 8.2–8.4, being slightly lower at 7.4–7.8,
decreasing significantly above pH 9, and at pH 9.7–9.9 the cells were unable to thrive.
However, many algal species accept higher Ph values than that. In algal cultures, pH
usually increases due to the photosynthetic CO2 assimilation . pH values above 10 is
not uncommon when no CO2 is supplied , and pH can reach 11 or more if CO2 is
limiting and bicarbonate is used as a carbon source.
Nitrogen absorption by the algae also affects pH in the medium. Assimilation of
nitrate ions tend to raise the pH, but if ammonia is used as nitrogen source, the pH of
the medium may decrease to as low as 3, which is too acid to support growth.

3.4 Cultivation methods
For commercial cultivation of algae, shallow raceway ponds and circular ponds with a
rotating arm to mix the cultures are usually used. The raceway pond is set in a
meandering configuration with paddle wheel mixers that exert low shearing forces.
For wastewater treatment, facultative ponds and high rate algal ponds (HRAP) are the
most commonly used. A facultative pond is usually deeper than one meter, has algae
growing in the surface water layers and is anoxic near the bottom. An HRAP, on the
other hand, is usually less than a meter deep, is continuously mixed by gentle stirring
and is aerobic throughout its volume. In HRAPs, microalgae supply oxygen to
heterotrophic bacteria, and the nutrients in the wastewater are converted into algal and
bacterial biomass. Like in facultative ponds, the raised pH causes ammonia stripping
and phosphate precipitation, and most studies about the role of algae in HRAPs point
out that this indirect nutrient removal is often more important than direct uptake. The
denitrification that occurs in facultative ponds should be considered negligible in an
HRAP though, because of the aerobic environment . According to Oswald (1988),
properly designed and operated HRAPs are capable of removing more than 90 % of
the biochemical oxygen demand (BOD) and up to 80 % of the nitrogen and
phosphorus.
Figure 3.2 – Open Pond system.

3.5 Closed photobioreactors
Closed photobioreactors can be grouped into two major classes: covered raceways and
tubular reactors . Closed photobioreactors usually have better light penetrating
characteristics than open ponds; the light path is usually less than 30 mm, which make
it possible to sustain high biomass and productivity with less retention time than is
possible in ponds . However, since they are more technically complicated, often need
expert personnel and require more energy than open systems; the operating cost is
higher . By using transparent pipes for cultivation, the internal shadowing effect
between the algae is minimised, and the cells can be illuminated from more than one
direction. The light refraction will create shaded areas in the tubes though, and
sufficient turbulence is therefore needed to provide all cells with light. Tubular
reactors can be placed vertically or horizontally, and be constructed of several
materials, rigid or soft. In a vertical column reactor, aeration and agitation can be
provided by injection of CO2-enriched air at the bottom of the column . A drawback,
however, is that these reactors are more or less parallel to the sun’s rays and a
substantial amount of solar energy is thus reflected in the summer.
Figure 3.3 - Closed Photo Bioreactors

3.6 Experimental Set-up
To study the role of microalgae in wastewater treatment, the following method was
employed,
 Wastewater treated with culture of C. vulgaris and S. quadricauda; and
 Wastewater treated without culture of C. vulgaris and S. quadricauda
2 ml of uniform suspension of C. vulgaris and S. quadricauda as initial inoculums (9
days old culture) in each flask containing 200 ml wastewater sample. The initial total
count of the C. vulgaris and S. quadricauda were 7.32×104 cell/ml and 3.46×104
cell/ml respectively. The experiment was conducted under controlled conditions
(Temp 27 ± 2º C) for a total duration of 20 days. Samples were periodically (every 5th
day) analyzed for physico-chemical parameters such as pH, phosphate, nitrate, BOD
and COD using standard methods.
Figure 3.4

3.7 Investigation - Lab work
3.7.1 Inlet readings of wastewater of rainy seasons
Table no. 3.1
CHARACTERISTS
OF
WASTE WATER
SAMPE
NO.
1
(1/7/2013)
SAMPE
NO.
2
(6/7/2013)
SAMPE
NO.
3
(11/7/2013)
SAMPE
NO.
4
(22/7/2013)
pH 7.24 7.18 7.27 7.20
DO 5.1 5.08 5.3 5.1
TSS 140 148 128 152
COD 216 202 230 216
BOD 110 100 115 123

3.7.2 Inlet readings of wastewater of winter season
Table no. 3.2
CHARACTERISIS
OF
WASTE WATER
SAMPLE
NO.
1
(2/12/2013)
SAMPLE
NO.
2
(7/12/201)
SAMPLE
NO.
3
(17/12/2013)
SAMPLE
NO.
4
(27/12/201)
PH 7.18 7.20 7.18 7.30
DO 4.60 4.30 4.60 4.15
TSS 158 170 165 170
COD 260 304 256 240
BOD 120 135 120 125

3.7.3 Inlet readings of wastewater of summer season
Table no. 3.3
CHARACTERISTCS
OF
WASTE WATER
SAMPLE
NO.
1
(1/2/2014)
SAMPLE
NO.
2
(6/2/2014)
SAMPLE
NO.
3
(17/2/2014)
SAMPLE
NO.
4
(14/03/2014)
pH 7.2 7.4 7.14 7.18
DO 2.50 2.35 2.20 2
TSS 188 166 174 172
COD 340 220 304 250
BOD 145 110 130 125
NITRATES 32.2 30.1 32.5 33.4
PHOSPHATES 4.0 4.2 4.15 3.99

CHAPTER 4
RESULTS

CHAPTER 4
RESULTS, DISCUSSION & CONCLUSION
4.1 ANALYSIS OF pH OF WASTEWATER USING C.VULGARIS AND
S.QUADRICAUDA
Table no. 4.1
DAYS S.QUARICAUDA C.VULGARIS
5 th 7.72 7.64
10 th 7.75 8.40
15th
7.77 7.76
20th
7.75 8.10

Analysis of ph of wastewater using c.vulgaris and s.quadricauda
Graph 4.1
Comment
 Mostly ranges between 7 to 9 but optimum is 8.2 to 8.7. In this, aerating and
mixing is necessary. Increased pH is controlled by addition of CO2.
 The nutrient removal is basically an effect of assimilation of nutrients as the
algae grow, but other nutrient stripping phenomena also occur, e.g. ammonia
volatilisation and phosphorus precipitation as a result of the high pH induced
by the algae.
 In algal cultures, pH usually increases due to the photosynthetic CO2
assimilation.
 In order to avoid extreme pH values, turbulence can promote the gas exchange
between water and air which in turn regulates pH somewhat in the water.
7.2
7.4
7.6
7.8
8
8.2
8.4
8.6
5 10 15 20
s qudriquada
c. vulgaris
X AXIS – Days Y AXIS – pH

4.2 ANALYSIS OF DO OF WASTEWATER
Table no. 4.2
Graph 4.2
0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
5 10 15 20
s. quadriquada
c. vulgaris
Incubation period in
days
DO increased by
s.quadricauda
DO increased by c.vulgaris
5 2.28 2.24
10 3.48 3.26
15 3.94 3.77
20 4.20 4.17
X axis – Days Y axis - DO

4.3 ANALYSIS OF COD OF WASTEWATER
Table no. 4.3
Incubation period in
days
% removal by s.quadricauda % removal by c.vulgaris
5 37 48
10 51 72
15 70 80
20 79 89

Analysis of COD of wastewater
Graph 4.3
Comment
 This graph shows that C. Vulgaris is more sustainable for chemical reactions
than S. Quadriquada as it grows faster than S. Quadriquada.
 C. vulgaris removes maximum COD content due to metabolism of
endogeneous bacteria.
 It is highly resistive to bacterial conditions and thus grows well in waste
stream.
0
10
20
30
40
50
60
70
80
90
100
5 10 15 20
% removal by s. quadriquada
% removal by c. vulgaris
X AXIS – Days Y AXIS - % Removal

4.4 ANALYSIS OF BOD OF WASTEWATER
Table no. 4.4
Incubation period in days % removal by s. quadricauda % removal by c.vulgaris
5 19 28
10 38 53
15 90 71
20 95 90

Analysis of BOD of wastewater
s
Graph 4.4
Comment
This graph shows that C. Vulgaris removes BOD at initial stage and at later stage S.
Quadriquada removes BOD efficiently.
0
10
20
30
40
50
60
70
80
90
100
5 10 15 20

Figure 5: Removal % of BOD and COD of Wastewater Using C. vulgaris and S.
quadricauda
Graph 4.5

4.5 ANALYSIS OF NITRATES
Table no. 4.5
Incubation period in days % removal by s. quadricauda % removal by c. vulgaris
5 28 43
10 50 62
15 70 79
20 80 90

Analysis of nitrates
Graph 4.6
Comment
C. Vulgaris has higher cell densities within the beads than S. Quadriquada as C.
Vulgaris grows faster and gets converted into multi- cellular organisms and this
indicates that it consumes more nitrates than S. Quadriquada.
0
10
20
30
40
50
60
70
80
90
100
5 10 15 20

4.6 ANALYSIS OF PHOSPHATES
Table no. 4.6
Incubation period in days % removal by s. quadricauda % removal by c. vulgaris
5 32 19
10 48 42
15 82 63
20 88 80

Analysis of phosphates
Graph 4.7
Comment
This graph shows that S. Quadriquada removes more phosphate content than
C.Vulgaris.
0
10
20
30
40
50
60
70
80
90
100
5 10 15 20
% removal by s. quadricauda

Figure 8: Removal % of Nitrate and Phosphate of Wastewater Using C. vulgaris
and S. quadricauda
Graph 4.8

CHAPTER 5
RESULT AND DISCUSSIONS

5.1 Results and discussions
 All physico-chemical parameters are quantified for 5th, 10th, 15th and 20th
days, respectively. The initial pH of wastewater was 7.41 ± 0.10 .
 When the wastewater is treated with C. vulgaris and S. quadricauda then the
pH increases as compared to control.
 BOD and COD levels of treated effluent are reduced significantly. The BOD is
an indicator measurement of substances that can be degraded biologically,
consuming dissolved oxygen in the treatment upto 20th days.
 The BOD level is reduced to 70.91 % by C. vulgaris and 89.21 % by S.
quadricauda up to 20th day.
 In this project, the COD level is reduced to 80.64% and 70.97% by C. vulgaris
and S. quadricauda upto 20th day respectively. C. vulgaris showed the best
removal capacity of COD from wastewater.
 C. vulgaris induced progressive reduction in both BOD and COD values of the
effluent and this could be attributed to the high algae growth rate and intense
photosynthetic activity. It is also observed that Scendesmus sp. showed high
removal efficiency for inorganic nutrients from domestic effluents. In the same
experiment, removal of nitrate using C. vulgaris and S. quadricauda from
wastewater is determined. Removal of nitrate from wastewater is 78.08% and
70.32% when treated with C. vulgaris and S. quadricauda upto 15th day.
 C. vulgaris shows best reduction capacity of nitrate from wastewater than
S.quadricauda.
 High levels of nitrogenous compounds in wastewater can be effectively
removed only by algae.

 In the present study C. vulgaris removed 62.73% of phosphate in wastewater
during the 15th days, while the maximum capability of removal is 79.66% on
20th day of experiment.
 Such a high percentage of removal is found. These results similar to that
reported 81.34 % for S. quadricauda during 15th day who concluded that
Chlorella and Scenedesmus were the most efficient algal strains to eliminate
phosphate from municipal waste.
 Phosphate was efficiently removed from the wastewater by S. quadricauda
within 15th days. The wastewater treatment using S. quadricauda found higher
removal rates of phosphate.
 Phosphate removal by C. vulgaris during remediation is due to the utilization
of phosphorus for growth. C. vulgaris removed 58.7% of phosphate in
wastewater while the maximum capability of removal was 91.9% on 20th day
of experiment while such a high percentage of removal was found 80.0 % for
S. quadricauda during the 15th day.
 DO of the wastewater increases from 2 mg/l to 4.17 mg/l by C. Vulgaris and
4.20 mg/l by S. Quadriquada as algae gives out oxygen during photosynthesis.

CONVENTIONAL METHOD VS ALGAE BASED ( S. QUADRIQUADA )
WASTEWATER TREATMENT
Graph 5.1
Comment
Comparing the natural conditions with the artificial conditions at a given period of
time, algae treatment shows better results than conventional treatment.
0
10
20
30
40
50
60
70
80
90
100
BOD COD Nitrate Phosphate
s. quadriquada
conventional
X axis - Parameters Y axis - % Removal

CONVENTIONAL METHOD VS ALGAE BASED ( C. VULGARIS )
WASTEWATER TREATMENT
Graph 5.2
Comment
Comparing the natural conditions with the artificial conditions at a given period of
time, algae treatment shows better results than conventional treatment.
0
10
20
30
40
50
60
70
80
90
100
c. vulgaris
conventional method

CONVENTIONAL METHOD VS ALGAE BASED WASTEWATER
TREATMENT
Graph 5.3
0
10
20
30
40
50
60
70
80
90
100
s. quadriquada
c. vulgaris
conventional

Calculation of F/M Ratio In Flask :-
Wastewater Flow(Q) = 0.001 m3
Volume of flask(V) = 0.001 m3
Inffluent BOD (Y0) =125 mg/litre
Effluent BOD = 6.25 mg/litre
Mixed liquor suspended solids(Xt) = 2150 mg/litre
F= Mass of BOD applied to aeration system=Q X Y0
=0.001 X125 gm/day
=0.001 X 125/1000 kg/day
=1.25 X 10 -4 kg/day
M=Mass of MLSS=V X XT = 0.001 m3 X 2150 mg/litre
= 0.001 X 2150/1000
= 2.15 X 10 -3 Kg
F/M RATIO = 1.25 X 10 -4/ 2.15 X 10-3=58.13 X 10 -3S

CHAPTER – 6
COST ANALYSIS

6.1 Cost Analysis of Conventional method of treatment of sewage:
 Let us consider conventional plant of treatment as Activated Sludge process
(ASP).
 Cost of construction of ASP plant = 70 lakh per MLD.
 For 1 MLD plant area required = 1 acre.
 Considering cost of land = 20 lakh per acre.
 Cost of land required for 1 MLD plant = 20 lakh.
Therefore Capital Investment required for setting up the Activated Sludge Process
treatment plant = Cost of construction + Cost of land required
= 70,00,000 + 20,00,000
= Rs 90,00,000 /-
According to DSR, operational and maintenance cost including electricity cost, labour
cost, repair and maintenance cost and depreciation cost is worked out as Rs 12 per
1000 litres of sewage.
𝐘𝐞𝐚𝐫𝐥𝐲 𝐂𝐨𝐬𝐭 𝐨𝐟 𝐨𝐩𝐞𝐫𝐚𝐭𝐢𝐨𝐧 𝐚𝐧𝐝 𝐦𝐚𝐢𝐧𝐭𝐞𝐧𝐚𝐧𝐜𝐞 𝐟𝐨𝐫 𝟏 𝐌𝐋𝐃 𝐩𝐥𝐚𝐧𝐭
= 𝟏𝟐 𝐱 𝟏𝟎𝟎𝟎𝟎𝟎𝟎 𝐱
𝟑𝟔𝟓
𝟏𝟎𝟎𝟎
= 𝐑𝐬 𝟒𝟑, 𝟖𝟎, 𝟎𝟎𝟎 /−

6.2 Cost Analysis of Algae based wastewater treatment plant:
 Cost Of algal Strain : Rs 575 per 50 ml
 For 2 ml algal strain cost required is Rs.115 /-
 Cost of Urea :Rs 350 per 25 kg
 For 2 gm of Urea : Rs 0.028 /-
 Electricity Cost 1% of treatment cost: Rs 1.15/-
 Total cost of treatment per litre of water =
Cost required for 2 ml algal strain+cost required
for 2 gm of urea+ Electricity cost
=Rs 115+ Rs 0.028+Rs 1.15
=Rs 116.17/-

CHAPTER - 7
CONCLUSION

CONCLUSION
In this project, it is seen that the growth rate of Chlorella vulgaris and
Scenedesmus quadricauda in the wastewater increases by reducing the rate of different
pollutants.It is observed that Chlorella vulgaris removes more nitrates and COD than
Scenedesmus quadricauda .while Scenedesmus quadricauda shows best result for
BOD and phosphate removal. Unicellular green algae such as Chlorella and
Scenedesmus have been widely used in wastewater treatment as they have fast growth
rates and high nutrient removal capabilities.
Therefore, it is found that the remediation using Chlorella vulgaris and
Scenedesmus quadricauda of wastewater provides an effective and environmentally
acceptable option for wastewater remediation, which is not only recycles valuable
nutrients but also improves wastewater quality.
This project also concludes that the algae treatment is more efficient for small scale
treatment in rural areas or communities than conventional methods.
In future the removed nutrients can be utilized for cattle feed ,fertilizers and biodiesel
production.

CHAPTER 8
FUTURE SCOPE

FUTURE SCOPE
Microalgae cultivated in the wastewater can be used for the
biodiesel production and as feed for animals. This dual process (microalgae
cultivation in effluent coupled with biodiesel production) has several
advantages such as less cost and less energy input for biodiesel production, and
less greenhouse gas emission during biodiesel production.
The present study is speculated that there is unlimited scope for using such
potential algae strains for enhanced activity towards rapid treatment and
probable recycling of waste water for useful purposes.

CHAPTER 8
REFERENCES
REFERENCES
o MENG Rui, HE Lian-sheng Study on Purifying the Deteriorated
Aquaculture Water with Bacteria-alga System International Conference
on Agricultural and Biosystems Engineering 2011 Advances in
Biomedical Engineering Vols. 1-2
o Erick griffiths, utah university, biological department algalbiomass
from wastewater for biodiesel production Evaluating algal growth at

different temperatures
o Karin Larsdotter, Environmental Microbiology, School of
Biotechnology, KTH, AlbaNova University Center, 106 91 Stockholm
Wastewater treatment with microalgae – a literature review
o Deviram GVNS*, Pradeep K.V and R Gyana Prasuna Department of
Microbiology, Gitam Institute of Science, GITAM University,
Visakhapatnam, India Purification of waste water using Algal species
Pelagia Research Library European Journal of Experimental Biology,
2011.
o S. Sriram and R. Seenivasan School of Bio Sciences and Technology,
VIT University, Vellore – 632014, India Microalgae Cultivation in
Wastewater for Nutrient Removal, Journal of Algal Biomass Utilization
J. Algal Biomass Utln. 2012.
o Sewage treatment and air pollution engineering by S.K.Garg.
o American Public Health Association (APHA)

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