Dr v sivasubramanian


Director, Phycospectrum Environmental Research Centre (PERC), 52A, AK Block, 7th Main Road, Anna Nagar, Chennai 600040, India and Executive Editor, Journal of Algal Biomass Utilization (JABU). Secretary, Indian Biomass Association (IBA). Assistant editor, PHYKOS. Member of Editorial Board, Seaweed research and utilization Journal.. BEST REVIEWER AWARD 2014 - Bioresource Technology Journal, Former Director, Vivekananda Institute of Algal technology (VIAT- 2006 - 2012). Former Professor and Head, Plant Biotechnology, RKM Vivekananda College, Chennai. Member of expert committee of Ministry of New and Renewable Energy (MNRE), New Delhi, on Second generation bio-fuels. Member of Monitoring Committee on Algal Biofuels, Council of Scientific and Industrial Research (CSIR), Member of project advisory and monitoring committee (PAMC) on CO2 sequestration projects of Department of Science and Technology (DST) and member of monitoring committee in projects on algal biofuels of Department of Biotechnology (DBT), New Delhi. member of Scientific Advisory Committee (SAC),DBT-IOC Centre for Advanced Bio-Energy Research, Faridabad. Member of international advisors of the Universidad del Norte Biotechnology Research Group - Biotechnology Research Group and the Group Rational Use of Energy and Environment Preservation, UREMA of Uni Norte. Member of the monitoring committee Technology Development Board (TDB) of DST.

Contact details: Phycospectrum, 52A, A K Block, 7th Main Road, Anna Nagar, Chennai 600040. Mobile: +91 9677144453: Email vsivasubramanian@gmail.com,


Since before the beginning of the century, we have dumped enormous amounts of waste products into the environment following the principle of ‘out of sight, out of mind”. Until World War II these waste products and their effects went largely unnoticed. However after WWII, the severity of pollution problems, resulting from careless waste disposal, has steadily increased. Today more than ever the general public is aware of these problems. Historically, sources of waste products have been industrial and agricultural. Industries generate waste during processing of coal and oil and also nuclear energy production. The wastes generated are buried and as a result contaminants have migrated through the soil into groundwater supplies (Point Source). Pesticides and fertilizers used in agriculture (Non-point source) also have contaminated the ground water. Microbial interactions with organic pollutants can be harnessed to help prevent contamination and cleanup of such sites.

The breakdown of organic contaminants that occurs due to microbial activity (Biodegradation). A series of biological degradation steps or a pathway ultimately result in the oxidation of the parent compound. Complete biodegradation or mineralization involves oxidation of the parent compound to form carbon-dioxide and water. For the microbes to act the contaminant should be available in water (Bioavailability) in soluble form. The contaminants with limited solubility or that are strongly sorbed to soil or sediments are not available and biodegradation.

Bioremediation is the process of using microorganisms to transform hazardous chemical compounds (most of them are organic compounds) in a contaminated soil, sediment to non-hazardous end products. Almost all-organic compounds and some of the inorganic compounds can be degraded biologically if sufficient time and proper physical and chemical conditions are provided. Two basic methods are available for obtaining the microorganisms necessary to initiate bioremediation: 1) Bio-augmentation, which involves the addition of adapted genetically coded toxicant degrading microorganisms 2) Bio-stimulation, which involves the injection of the necessary nutrients to stimulate the growth of indigenous microorganisms

Removal of inorganic materials by microorganisms may be achieved when the organic compounds are utilized as electron donors by the microorganism. In addition, metals may be removed from environment by precipitation during biological activity and also by bioaccumulation. The effectiveness of bioremediation of hazardous compounds is mainly influenced by the degradability and the toxicity of the compounds, which are present in the soil or sediments. Based on these parameters, the chemical compounds are broadly categorized as 1) Degradable and nontoxic 2) Degradable and toxic 3) Non-degradable and toxic and 4) Non-degradable and nontoxic

Goals of bioremediation:

The main goals of bioremediation can be 1) Enhancing the rate and extent of biodegradation of the pollutants in consideration 2) Utilizing or developing microorganisms that is capable of surviving the toxic effects of the pollutants and 3) utilizing the microorganisms in such a way that the products of the degradation process are not toxic.

Advantages and applications:
All the current technologies that are available for treatment of waste have lots of limitations. Most of them are not cost effective and are inappropriate for the in situ treatment. Some of them are not effective in treating a complex array of different pollutants. Biological treatment appears to offer solution to these limitations. The basic advantage of bioremediation are 1) Biodegradation is a “natural” process in which naturally occurring microorganisms are used for treatment of wastes 2) The residues or the by-products of biological processes (CO2, H2O) are usually geochemically cycled in the environment as harmless products 3) Microorganisms such as bacteria and fungi have a wide range of abilities to metabolize different chemicals 4) the technologies are developed to utilize and improve native microorganisms that have been demonstrated to degrade the pollutants on site. In this case, adding nutrients or other amendments to the site can accelerate the rate of microbial activity. In other cases, microorganism known to metabolize the pollutants can be introduced and supplemented to improve biodegradation 5) For in situ treatment of soils, sludge, and ground water, bioremediation is less expensive and less disruptive than options frequently used for treatment, such as excavation followed by incineration or land filling. Today, bioremediation is widely applied in the treatment of contaminated water, soil, sludge, and sediments. Bioremediation is the best method for remediation of the long chain molecular organic compounds, hazardous waste and toxicity chemical.

Key factors – critical to successful application of bioremediation:
Environmental conditions, contaminant and nutrient availability and the presence of degrading microorganisms are the most important factors critical to successful application of bioremediation. If biodegradation does not occur, the first thing that must be done is to isolate the factor limiting bioremediation. Initial laboratory tests using soil or water from a polluted site can usually determine whether degrading organisms are present and whether there is an obvious environmental factor that limits bioremediation for ex. extremely low pH or lack of nitrogen and /or phosphorus. Low bioavailability due to sorption and aging is another factor that can limit bioremediation.

Metal pollutants
Metal pollution is a global concern and the levels of metals in all environments, water, air and soil, are increasing, in some cases to toxic levels, with contributions from a wide variety of industrial and domestic sources. Metal availability is strongly dependant on environmental components, such as pH, redox and organic content and soluble and bio-available metals. Metals in the environment can be divided into two classes: 1) bio-available (soluble, non sorbed and mobile) 2) non bio-available (precipitated, complexed, sorbed and no mobile). According to Becker (1983, 1994) utilizing planktonic algae with a high potential to absorb heavy metals for the removal of residual metals from waste waters including the separation of the metal-saturated algae from medium is an economic method resulting in high quality reusable effluent water and valuable biomass which could be used for different purposes (production of biogas, fertilizer, fodder etc.)

Micro algae and wastewater treatment

Micro algae are small photosynthetic microorganisms, adapted to almost any possible environment. Phycoremediation may be defined in a broad sense as the use of macro algae or micro algae for the removal or biotransformation of pollutants, including nutrients and xenobiotics from wastewater and CO2 from waste air (Olguín, 2003). This field has evolved from the early work done by Oswald and Gotaas (1957) for the use of micro algae for tertiary treatment of municipal wastewater to many other applications in which micro algae and macro algae are cultivated and utilized for specific bioremediation needs. Thus, phycoremediation comprises several applications: (a) nutrient removal from municipal wastewater and effluents rich in organic matter; (b) nutrient and xenobiotic compounds removal with the aid of algae-based biosorbents; (c) treatment of acidic and metal wastewaters. The use of micro algae for the treatment of municipal wastewater has been subject of research and development for several decades. The result of such effort is that some commercial technologies and processes are available in the market such as the Advanced Integrated Wastewater Pond Systems (AIWPS) Technology commercialized by Oswald and Green, LLC, in the United States.

Three types of ponds are employed in wastewater treatment; 1) anaerobic, 2) facultative and 3) aerobic. Anaerobic ponds are several meters deep; they are free of dissolved oxygen and have high BOD removal rates. The facultative ponds exhibit aerobic conditions on the surface due to photosynthetic oxygen production by algae and anaerobic conditions in the bottom layers and are the most common form of oxidation ponds. Aerobic ponds, also known as high-rate ponds, are shallow and completely oxygenated throughout (Oswald, 1978)
According to Becker (1983) the efficiency of the concept will be determined principally by the following parameters:
1. Growth rate of algae
2. Metal concentration factor attained by the alga
3. Concentration of heavy metal in the medium
4. Desired percentage of metal removal from the medium
5. Metal recovery in relation to capital and operating costs

a. High-rate algal ponds (HRAP)
Nutrient removal with the aid of algae compares very favorable to other conventional technologies (Muthukumaran et al 2005; De la Noue et al., 1992). In contrast to facultative ponds, HRAPs are designed to promote algae growth. They are shallow (0.3-0.6 m) in order to allow maximum light penetration. They can operate at short hydraulic retention time (HRT) in the range of 4 to 10 days depending on climatic conditions reducing the required surface area. Continuous mixing is provided to keep the cells in suspension and to expose them periodically to light. The most common design that has proven successful at large scale is the single loop paddlewheel mixed. Due to energy cost dependence on velocity, most ponds have been operated at velocities from l0 to 30 cm/ sec (Dodd, 1986). More recently, a special flow pattern was introduced to improve the efficiency of this type of ponds (Mihalyfalvy et al., 1998).
. HRAPs are by far the most cost-effective reactors available for liquid waste management and for efficient capture of solar energy (Oswald, 1995). The AIWPS system designed by Oswald , LLC, consumes from O to 0.57 kWh/kg BOD removed. In contrast, mechanical aerated ponds consume a much higher amount of energy in the range of 0.80 to 6.41 kWh per kg of BOD removed.
b. Cell immobilization
In phycoremediation, harvesting of micro algae is the costliest step and to avoid excessive harvesting cost, a technique has been developed by various research groups working in the field of cell immobilization. In depth discussion of the various immobilization methods and their applications is already available (Wilde and Benemann, 1993; Garbisu et al., 2000; Mallick, 2002). Green algae and cyanobacteria have been successfully immobilized for the removal of nutrients. It is worth noticing that chitosan and polyvinyl foams are low-cost polymers with a long-term performance. On the other, although each one of these particular immobilization systems have been found to be successful at the lab level, some matrices and immobilization techniques would have some limitations at the industrial level (Levy and Shoseyov, 2002). For example, hollow fibers are expensive (Kang et al., 1990). As for techniques, loss of cell viability has been observed when using covalent immobilization (Jirku, 1999), and mass transfer limitations at a high degree have been encountered while using cell entrapment methods (Pilkington et al., 1998). Thus, it is recommended to choose matrices with physical and chemical resistance or long-term use and to avoid complex immobilization method

c. Use of strains with special attributes
Nutrient removal has been shown to be more efficient by using algae strains with special attributes. Such special attributes include tolerance to extreme temperatures, chemical composition with predominance of high added value products, a quick sedimentation behavior, or a capacity for growing mixotrophically. A Phormidium strain capable of removing nutrients more efficiently than a community of green algae below 10°C was iso1ated from polar environments (Tang et al., 1997). The authors suggested that this strain was appropriate for wastewater treatment in cold climates during spring and autumn.
On the other hand, Talbot and de la Noue (1993) reported that Phormidium bohneri was a
good candidate for treating wastewater at high temperatures (around 30°C); additional1y,
such strain had a quick sedimentation behavior. Spirulina (Arthrospira) is one of the most favoured micro algae for wastewater treatment (Laliberté et al., 1997). The advantages of using Spirulina are as follows (Olguín et al., 2003): (1) capacity to floccu1ate makes harvesting easier and cheaper than for other microa1gae( Mohn, 1988); (2) the biomass with the highest possible protein content (60-70% dry weight) when grown under conditions avoiding nitrogen limitation (Ciferri, 1983); (3) used successfu11y as a feed supp1ement for mammals( Becker, 1994) and fish larvae (Belay et al., 1993); (4) high content of high added va1ue compounds such as polyunsaturated fatty acids (Olguín et al., 2001), which have been reported to have therapeutic effects in humans (Belay et al., 1993); (5) biomass enriched in polysaccharides may be utilized as a very efficient bio-adsorbent for heavy metals (Hernández and Olguín, 2002); (6) ability to grow at high pH values reduces contamination by other species (Olguín et al.,1997; Olguin, 2000); (7) some strains grow at a very high ammonia-nitrogen concentration (130 mg 1- 1) and (8) ability of some strains to grow under heterotrophic and mixotrophic conditions (Márquez et al., 1993, 1995). The use of algae for improving water quality (pH, dissolved oxygen, suspended solids etc) and removal of nutrients and metals from eutrophic or contaminated water has been increasing over the past few decades (Oswald, 1988). High rates of nutrient removal and algal production have been measured with monocultures of cyanobacteria such as Spirulina (Lincoln et al., 1996; Olguin et al 1997) and Phormidium (Blier et al 1996) grown on manure effluent from diary and swine operations.

Mulbry et al (2005) working on recycling of manure nutrients reported the use of algal biomass from dairy manure treatment as a slow release fertilizer. During storage and land application of manure effluents, large amount of N are lost to the atmosphere due to volatilization of ammonia. Instead, crops of algae are grown on the N and P present in the manure and convert manure N and P into algal biomass. Algal biomass generated is used for soil conditioning and as a slow release fertilizer. Westhead et al (2003) worked on production and nutrient removal by periphyton grown under anaerobically digested flushed dairy manure. They suggested growing algae to scrub nutrients from manure as an alternative to the current practice of land application and provide utilizable algal biomass. Kirkwood et al (2003) working on the physiological characteristics of cyanobacteria (Phormidium, Pseudanabaena) in pulp and paper waste-treatment systems reported that the pulp and paper industry depend on secondary (biological) waste-treatment system to treat highly concentrated organic waste. The wastewater contains hundreds of wood extractives (eg resin acids) and chlorinated organic compounds (eg Chlorphenicols) of environmental concern and consequently, pulp and paper wastewater is toxic and requires microbial mineralization to reduce effluent toxicity. ASBs (aerated stabilization basins and activated sludge systems (ASs) used by pulp industry are designed to support high densities of heterotrphic bacteria Both ASBs and ASs are open-air facilities. The ubiquitous presence of cyanobacteria (Phormidium, Pseudanabaena) observed by Kirkwood et al made them to suggest the utilization of these algae for wastewater treatment systems. Although all cyanobacteria are photoautotrophic, many can utilize simple organic carbon (DOC) compounds for heterotrophic growth or for mixotrophic growth in light. Cyanobacteria also have the potential for catabolism of the contaminants in wastewater.

Halogenated compounds represent one of the most predominant environmental pollutants due to their widespread usage as biocides, fungicides, disinfectants, solvents and other industrial chemicals. Biodegradation of chlorinated phenols has been studied with pure and mixed bacterial cultures. Only a few algae like Chlorella sp were found to decolourize certain azo dyes and use them as carbon and nitrogen sources (Jinqi and Houtian, 1992). Phenol degradation by Ochromonas danica was reported by Semple and Cain (1996). Luther (1990) has reported that the alga Scenedesmus obliquus was able to utilize naphthalenesulphonic acids as a source of sulphur for their biomass, releasing the carbon ring into the medium. Pentachlorophenol has been reported to be degraded by Chlorella sp. Lima et al (2004) reported micro algae isolated from a waste discharge container fed with several aromatic pollutants were able to remove ?-chlorophenol and ?- nitrophenol under different photo-regimes. The study of the role of micro algae in biodegradation systems is scarcely reported (Lima et al, 2004). A common approach used to treat organic compounds is a combination of biodegradation and adsorption processes. Adsorbing material, like zeolite or activated carbon, may be added to a biological process in order to improve the overall performance of the system and to increase the removal of the most recalcitrant organic material from wastewater.

Sanchez et al (2001) grew a mixotrpohic culture of Chlorella pyrenoidosa with olive-mill wastewater as a nutrient medium. Olive-mill wastewater normally contains 1) Vegetable water from the fruit 2) Water from the process and 3) Water from industrial installation (cleaning, sewage waters etc). This can support a luxuriant growth of algae due to the presence of carbohydrates and mineral salts. So, waste currently causing grave environmental problems can be used to grow algae. The biomass generated can render valuable biomolecules, such as pigments and fatty acids.

Micro algae at wastewater pond treatment in cold climate:
Cold climate is normally not favourable for phycoremediation due to the poor light availability and low temperature. But still many workers in the past have successfully demonstrated the feasibility of using certain micro algae with special attributes to treat wastewater. Erik Gronlund (2004) working with wastewater treatment in Sweden discusses the following stages:
1. Primary – easily settled materials
2. Secondary – oxidize organic material present in the wastewater – stabilization
3. Tertiary – removal of solved nutrients
4. Quaternary – removal of refractory and toxic organics
5. Quinary – removal of heavy metals, organic compounds and soluble materials
Erik Gronlund used Coelastrum, Chlamydomonas, Chlorella, Selenastrum, Scenedesmus and Micractinium pusillum in HRAs with artificial illumination and covered with green house plastic. Relatively few investigations have been done in cold climates. Tang et al (1997) compared polar cyanobacteria and chlorococcales for tertiary wastewater treatment. Phormidium sp was considered more promising. The cyanobacteria are preferred due to their flocculation properties, active growth and nitrate and phosphate uptake at low temperatures. Craggs et al(1995) showed in Scotland the possibilities of treating wastewater by diluting it with seawater and let marine micro algae (Phaeodactylum tricornutum) assimilate the nutrients.

d. Starved and activated algae

De La Noue et al (1980) showed that nitrogen uptake could be increased if the microalgae were preconditioned by starvation. These hyperconcentrated algal cultures, called ‘activated algae’ were shown to decrease the land and space requirements for micro algal treatment of wastewaters. This process removed nitrogen and phosphorus within very short period if time i.e., less than one hour. (Levoie and De la Noue, 1985). There is evidence that production of micro algae, given proper conditions, may be high enough even during colder periods to be of interest for wastewater treatment. However, this is to be verified under the actual local environmental conditions, since many strongly variable factors are involved when defining micro algal growth and species composition.

When phycoremediation is a success in cold climates where light and temperature, the two most essential factors for better remediation, are not available at optimum levels and definitely it can work very well in tropical conditions where sunlight and heat is available almost through out the year.

Our research group (Unit of Algal Physiology & Biotechnology, RKM Vivekananda College, Chennai) working with wastewater treatment system in SNAP Natural and Alginate Products , Ranipet have isolated a cyanobacterium, Chroococcus turgidus, responsible for increase in pH and remediation of acidic effluent generated by the industry. We have identified, isolated and experimentally proved the micro alga responsible for efficient remediation of the effluent. Chroococcus sp has other special qualities.
1. It can grow in a wide range of salinities.
2. Can tolerate a wide range of pH
3. Tolerance to high temperature
4. Capacity to degrade phenols
5. Tolerance to detergents
6. Fix nitrogen
and many more……
The acidic effluent form this industry is being treated in a number of shallow tanks (HRAPs) using micro algal technology to increase pH and to reduce salt levels. Our project with SNAP is in progress. Work is underway to treat effluent using sloped pond and the results are encouraging (unpublished).

Stahl India Chemicals Pvt Ltd., Ranipet, is one of the world’s leading suppliers of leather processing products The effluent is collected in a tank, aerated and sent to filter press to remove solids and the water is sent to aeration tank and used to water plants and grasses in STAHL. Tons of solid waste had been accumulated for the last 5 years.
The effluent has residual pigments, chemicals and protein and heavy metals such as copper, zinc, chromium, cadmium, lead and nickel. Our team made laboratory trials using micro algae, including Chlorella, Oscillatoria and Phormidium isolated from the effluent along with Scenedesmus, Chroococcus and Spirulina which grew very well in raw effluent. Preliminary field trials were conducted with Chlorella, Scenedesmus and Spirulina. Based on the results it was decided to use the raw effluent as such without any prior anaerobic digestion in further field trial experiments. Micro algae could effectively remove heavy metals including the most toxic lead, within 5 days of algal growth. Our results also suggest the possibility of some metals disappearing from the effluent either by getting converted by the algae or possibly shed as nanoparticles. The aspect requires further investigation.

In a recent article Venkataraman (2005) observed “the production technologies of micro algae and biochemicals of high value from them have not been substantial. The reasons are many and reflect the ethos of industry and inability to adopt unconventional technologies. Control of algal growth in unwanted regions viz., ponds, lakes, industrial pipelines, swimming pool which has a huge market has never been addressed. India has failed to introduce anything special and saleable in this area of algal technology as in the west. This is a paradox considering that most part of India with plentiful sunshine is really well suited for algal production almost year around”. Wastewater grown micro algal products with commercial value are few today. The most widespread use is the photosynthetic oxygenation. There is also biogas production and fertilizer use of sludge partly made up of micro algae. In biotechnological micro algal production there are many products from micro algae at a research scale and a few at a commercial scale. These could also be produced from wastewater, with benefits on the economic side, but constraints like hygienic aspects, unwanted contaminants and in many countries probably some concern regarding the use of human excretion products to produce consumables. Possible products are hydrogen gas, feed for aquaculture and in feedlots, advanced fertilizers and biochemical constituents. Only about one per thousand of the world algal species are sufficiently explored concerning biochemical content and in India the fraction could be still lower. It is high time that scientists and industry people should work together to find eco-friendly and cost effective solutions using micro algal technology to most of the problems caused by effluents in India now.


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