Heavy Metals Removal Using Immobilized Algae Biology Essay


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The objective of this work is to investigate the technical feasibility of using immobilized inactive algal biomass for the removal of heavy metals from aqueous solutions. Factors affect the selection of metals for biosorption studies will be mentioned. Comparison between The methods of metal removal from aqueous streams will also be investigated. The principal techniques of immobilization also will be discussed and comparison between them to choose one for a future work will be made.


According to the American College Dictionary, pollution is defined as:  to make foul or unclean; dirty. Water pollution occurs when a body of water is adversely affected due to the addition of large amounts of materials to the water.  When it is unfit for its intended use, water is considered polluted.  Many types of water pollutants exist such as sewage, fertilizers contain nutrients, such as nitrates and phosphates, and heavy metals.

Pollution of water by heavy metals has become a serious problem in some industrialized

countries (Inthorn et al., 1996).

Heavy metals enter the environment through wastewater streams from industrial processes such as electroplating, plastics manufacturing, mining and metallurgical processes (Yu and Kaewsarn, 1999).Heavy metal pollution of waterbodies due to indiscriminate disposal of industrial and domestic wastesthreatens all kinds of inhabiting organisms (De Filippis and Pallaghy, 1994; Nagase et al., 1997). Therefore, it is necessary to alleviate heavy metal burden of wastewaters before discharging them into waterways. At present, a number of different technologies exist for treating heavy metals bearing streams, such as chemical precipitation, adsorption, solvent extraction, ion exchange and membrane separation (Eccles, 1999). However, these methods have several disadvantages, such as incomplete metal removal, expensive equipment and

monitoring system requirements, high reagent or energy requirements and generation of toxic sludge or other waste products that require disposal.

Further, they may be ineffective or extremely expensive when metal concentration in wastewater is in the range 10-100 mg l-1 (Mehata and Gaur, 2005). The use of biological processes for the treatment of metal enriched wastewaters can overcome some of the limitations of physical and chemical treatments and provide a means for cost- effective removal of metals. A great deal of interest has recently been generated using different kinds of inexpensive biomass for adsorbing and removing heavy metals from wastewater (Volesky and Holan, 1995). In this context, accumulation of metals by microorganisms, including algae, has been known for a few decades, but has received increased attention only in recent years because of its potential for application in environmental protection or recovery of precious or strategic metals (Tsezos, 1985, 1986; Volesky, 1987; Malik, 2004). Algal biomass (continuous process) cannot be used directly in a standard sorption process. It is generally of small particle size and low strength and density, which can limit the choice of a suitable reactor and make biomass or effluentseparation difficult (Tsezos, 1986). Immobilized biomass(batch process) has the potential to provide asimple technology to remove and recover heavy metals from wastewater, and is suitable for reuse (Rai and Mallick, 1992). This study aimed to investigate the removal and recovery of heavy metals immobilized alginate beads (with live or dead cells or blank beads).

Chapter 1: water treatment


   With two thirds of the earth's surface covered by water and the human body consisting of 75 percent of it, it is evidently clear that water is one of the prime elements responsible for life on earth. Water circulates through the land just as it does through the human body, transporting, dissolving, replenishing nutrients and organic matter, while carrying away waste material. Further in the body, it regulates the activities of fluids, tissues, cells, lymph, blood and glandular secretions.

       An average adult body contains 42 litres of water and with just a small loss of 2.7 litres he or she can suffer from dehydration, displaying symptoms of irritability, fatigue, nervousness, dizziness, weakness, headaches and consequently reach a state of pathology.

1.2 Water pollution

Water pollution is the contamination of water bodies (e.g. lakes, rivers, oceans, groundwater).Water pollution affects plants and organisms living in these bodies of water; and, in almost all cases the effect is damaging either to individual species and populations, but also to the natural biological communities.

Water pollution occurs when pollutants are discharged directly or indirectly into water bodies without adequate treatment to remove harmful compounds.

Water pollution is a major problem in the global context. It has been suggested that it is the leading worldwide cause of deaths and diseases,[1][2] and that it accounts for the deaths of more than 14,000 people .

1.2.1- Types of water pollution:

1.chemical pollution:

Total dissolved solids.

Nutrients (nitrates &phosphorous).

Toxic organics.

Heavy metals.

2. physical pollution:

Thermal pollution.

Flouting debris.


3.Biological pollution:

Fecal coliforms.


1.3 Heavy metals

The term heavy metal refers to any metallic chemical element that has a relatively high density and is toxic or poisonous at low concentrations.

Heavy metals are natural components of the Earths crust. They cannot be degraded or destroyed ,to a small extent they enter our bodies via food ,drinking -water and air.

At higher concentrations they can lead to poisoning. Heavy metals poisoning could result, for instance, from drinking-water contaminations, high ambient air concentrations near emission sources, or intake via the food chain.

Heavy metals enter the environment through wastewater streams from industrial processes such as electroplating, plastics manufacturing, mining and metallurgical processes (Yu and Kaewsarn, 1999).Heavy metal pollution of water bodies due to indiscriminate disposal of industrial and domestic wastes threatens all kinds of inhabiting organisms (De Filippis and Pallaghy, 1994; Nagase et al., 1997).

In spite of the ever-growing number of toxic metal contaminated sites, the most commonly used methods dealing with heavy metal pollution are extremely costly process. Therefore in the recent years new and cheap method of water treatment is the concern.

1.3.1 Sources of discharge of metals

Lead: present in petrol-based materials and many other industrial facilities

(Sag and Kutsal 1997).

Chromium industrial operations including chrome plating, petroleum refining,

leather, tanning, wood preserving, textile manufacturing and pulp

processing. It exists in both hexavalent and trivalent forms.

Iron: Iron and steel units, electroplating industries and galvanizing units.

Zinc: widely used in industry to make paint, rubber, dye, wood

preservatives, and ointments and electroplating industries.

Nickel: galvanized, paint and powder batteries processing units

1.3.2 Harmful effects

Metals Health Risks:

Iron: The lack of iron in the diet may lead to iron-deficient anemia, Fatigue,

Weakness, Drowsiness, Pallor (paleness), Cold extremities, Brittle nails, Loss of appetite, Constipation, Headaches, Irritability, Difficulty concentrating, Depression, Loss of libido, Tinnitus (ringing in the7 ears), Spots before eyes, Bizarre behavior, Gastrointestinal complaints, Cessation of menstruation, Jaundice.

Chromium: Irritant, nausea and vomiting, carcinogen (Oxidation state of +6), Low-level exposure can irritate the skin and cause ulceration. Long-term

exposure can cause kidney and liver damage, and damage too circulatory and nerve tissue.

Zinc : Nausea and vomiting. Zinc combines with other elements to form zinc

compounds; common zinc compounds found at hazardous waste sites

include zinc chloride, zinc oxide, zinc sulfate, zinc phosphide, zinc

cyanide, and zinc sulfide.

Lead: Damage to nervous system, circulatory system, blood forming system, reproductive system, gastrointestinal tract and kidney Lead is known for its harmful affect on the living world, enters the organism by

breathing, swallowing, or absorption through the skin. The greatest

danger from lead comes from its tendency to accumulate in the

human organism. The central nervous system is most sensitive to the

effects of lead.

Nickel: Short-term overexposure to nickel is not known to cause any health problems, but long-term exposure can cause decreased body weight,

heart and liver damage, and skin irritation. The EPA does not currently regulate nickel levels in drinking water.

The health risks of heavy metal ingestion are widely ranging. Some metals causes physical discomfort while others may cause life-threatening illnesses, damage to vital body system, or other damage. In many instances, the effects of heavy metals on human are not well understood or documented.

1.3.3 Heavy metal toxicity

Metals and their "free radicals" are highly reactive attacking other cellular structures.

The ability of metals to disrupt the function of essential biological molecules, such as protein, enzyme and DNA is major cause of their toxicity. Displacement of certain metals essential for cell by a similar metal is another cause of toxicity.

For example cadmium can substitute for the essential metal zinc in certain protein that require zinc for their structure and function. The alteration in protein can lead to toxic consequences. In the same way, lead can substitute for calcium in bone and in other sites where calcium is required.

1.4 Methods of water treatment:

The treatment of water containing heavy metals is an area in which much research has bean performed.

The technology is currently in place to remove heavy metals from water to safe levels.

The method of metal removal chosen can depend on several factors. As economics is always a prime concern in industry, cost is one of the biggest factors for any case. Other variables may include such factors as waste location, other contaminants present, volume of water to be treated, and type of metal being removed. Sometimes it is possible to find a relatively inexpensive solution, and other cases may require substantial expenditures to clean the heavy metals from the water.

The commonly used procedures for removing metal ions from aqueous streams include chemical precipitation, lime coagulation, ion exchange, reverse osmosis and solvent extraction (Rich and Cherry, 1987). The process description of each method is presented below.

1.4.1 Reverse Osmosis: It is a simple process in which heavy metals are separated by a semi-permeable membrane at a pressure greater than osmotic pressure caused by the dissolved solids in wastewater. The disadvantage of this method is that it is expensive.

1.4.2 Electrodialysis: In this process, the ionic components (heavy metals) are separated through the use of semi-permeable ion­selective membranes. Application of an electrical potential between the two electrodes causes a migration of cations and anions towards respective electrodes. Because of the alternate spacing of cation and anion permeable membranes, cells of concentrated and dilute salts are formed. The disadvantage is the formation of metal hydroxides, which clog the membrane.

1.4.3 Ultrafiltration: They are pressure driven membrane operations that use porous membranes for the removal of heavy metals. The main disadvantage of this process is the generation of sludge.

1.4.4 Ion-exchange: In this process, metal ions from dilute solutions are exchanged with ions held by electrostatic forces on the exchange resin. The disadvantages include: high cost and partial removal of certain ions.

1.4.5 Chemical Precipitation: Precipitation of metals is achieved by the addition of coagulants such as alum, lime, iron salts and other organic polymers. The large amount of sludge containing toxic compounds produced during the process is the main disadvantage.

1.4.6 Phytoremediation: Phytoremediation is the use of certain plants to clean up soil, sediment, and water contaminated with metals. The disadvantages include that it takes a long time for removal of metals and the regeneration of the plant for further biosorption is difficult.

But it is also an ecologically friendly, solar-energy driven clean-up technology,based on the concept of using to cleanse nature.

The objective of this project is to study the immobilization algae to reduce the heavy metals pollution by using the phytoremediation technology.


This technology will be discussed in the next chapter.

Chapter 2: Literature survey:

1) Norton et aI. 2003, used dewatered waste activated sludge from a sewage treatment plant for the biosorption of zinc from aqueous solutions. The adsorption capacity was determined to be 0.564 mM/g of biosolids. The use of biosolids for zinc adsorption was favourable compared to the bioadsorption rate of 0.299 mM/g by the seaweed Durvillea potatorum (Aderhold et aI. 1996). Keskinkan et al. 2003 studied the adsorption characteristics of copper, zinc and lead on submerged aquatic plant Myriophyllum spicatum. The adsorption capacities were 46.69 mg/g for lead, 15.59 mg/g for zinc and 10.37 mg/g for copper.

2) Pagnanelli, et al 2002 have carried out a preliminary study on the 'Use of oli ve mill residues as heavy metal sorbent material The results revealed that copper was maximally adsorbed in the range of 5.0 to 13.5 mg/g under different operating conditions.

3)The simultaneous biosorption capacity of copper, cadmium and zinc on dried activated sludge (Hammaini et al. 2003) were 0.32 mmoI/g for metal system such as Cu­Cd; 0.29 mmoI/g for Cu-Zn and 0.32 mmoI/g for Cd-Zn. The results showed that the biomass had a net preference for copper followed by cadmium and zinc.

2.1. Immobilization methods of P. chrysosporium

2.1.1. Immobilization in Ca-alginate

Varying quantities of biomass (0.8-3%, w/v) were suspended in a 2% (w/v) Na-alginate solution and stirred. The mixture was then dropped into a 0.2 M CaCl2 solution, and the drops of alginate-biomass mixture were later gelled into beads with a diameter of 4.0 ± 0.2 mm. The Ca-alginate immobilized P. chrysosporium beads were stored in 0.2 M CaCl2 solution at 4 °C for 4 h to cure. The beads were rinsed twice with distilled water and stored at 4 °C prior to use. For blank Ca-alginate beads, similar procedures were used but without fungal biomass. Na-alginate solutions with different concentrations were also prepared to form the Ca-alginate immobilized fungal beads and blank Ca-alginate ones.

2.1.2. Immobilization in Ca-alginate-PVA

A Na-alginate-PVA mixture was prepared with different concentrations of PVA (1-6%, w/v) and a constant concentration of Na-alginate (2%). The biomass of 1.25% (w/v) was added in the mixture above and stirred. Ca-alginate-PVA immobilized fungal beads and blank beads were prepared using the same procedure for Ca-alginate immobilized fungal beads.

2.1.3. Immobilization in pectin

For biomass immobilization, 2%, 3%, and 4% (w/v) pectin solutions were prepared and the required dose of the biomass (1.25%, w/v) was mixed with the pectin solution. The mixture was then dropped into 0.2 M CaCl2 solution for polymerization. The resultant beads were cured in 0.2 M CaCl2 solution at 4 °C for 6 h. Blank beads were prepared with the same procedure for the immobilized biomass beads but without fungal biomass. All beads were rinsed thoroughly with distilled water and stored at 4 °C prior to use.

2.2 . Biosorption studies

The biosorption of 2,4-DCP onto the blank beads, free, and immobilized fungal biomass was investigated in batch experiments. The stock solution of 2,4-DCP at 100 mg/l was prepared using distilled water, and all solutions used in tests were prepared by appropriately diluting the stock solution to a pre-determined concentration. For comparison, 0.5 g free biomass (dry weight), immobilized fungal beads containing 0.5 g biomass and blank beads were respectively mixed with 100 ml of 2,4-DCP solution with a known initial concentration at natural pH of 5.0 in a 250-ml glass Erlenmeyer flask. Flasks were agitated on a shaker at 180 rpm and 25 °C. Samples were taken from the solution at given time intervals and analyzed for 2,4-DCP concentration as described below.

For isotherm studies, the blank beads, free, and immobilized fungal biomass were respectively put into 2,4-DCP solutions with initial concentrations from 10.16 mg/l to 81.38 mg/l. All biosorption experiments were carried out at pH 5.0, which was found to be appropriate for biosorption experiments in our previous work (Wu et al., 2005). The amount of adsorbed 2,4-DCP was calculated from the following equation:


where qe is the equilibrium uptake (mg/g), C0 is the initial 2,4-DCP concentration (mg/l), Ce is the equilibrium 2,4-DCP concentration (mg/l), V is the volume of the solution (1) and w is the quantity of the adsorbent (g).

2.3. Desorption studies

To recover the 2,4-DCP adsorbed from the blank beads, free, and immobilized fungal biomass, the 2,4-DCP-laden adsorbents were soaked in distilled water with continuously stirring at 180 rpm and 25 °C for 90 min. In order to obtain the optimum S/L ratio for the desorption, the 2,4-DCP-laden adsorbents was eluted with distilled water of 30 ml (S/L = 38.0), 40 ml (S/L = 28.5), 60 ml (S/L = 19.0), 80 ml (S/L = 14.3), 100 ml (S/L = 11.4), and 140 ml (S/L = 8.1), respectively. Desorption efficiency was calculated from the amount of 2,4-DCP adsorbed onto the beads and the final amount of 2,4-DCP desorbed into the eluant.

2.4. Biosorption/desorption cycles

In order to assess the reusability of the Ca-alginate immobilized fungal beads, experiments of five successive adsorption/desorption cycles were carried out using the same biosorbent with distilled water. The efficiencies for re-adsorption of 2,4-DCP in the repeated adsorption/desorption cycles were compared. The mass loss after consecutive adsorption/desorption cycles was also compared among the blank Ca-alginate beads, free, and immobilized fungal biomass.


3.1 Adsorption

The use of solids for removing substances from either gaseous or liquid solutions has been widely used. This process, known as adsorption, involves nothing more than the preferential partitioning of substances from the gaseous or liquid phase onto the surface of a solid substrate. Adsorption phenomena are operative in most natural physical, biological, and chemical systems, and adsorption operations employing solids such as activated carbon and synthetic resins are used widely in industrial applications and for purification of waters and wastewaters.

The process of adsorption involves separation of a substance from one phase accompanied by its accumulation or concentration at the surface of another. The adsorbing phase is the adsorbent, and the material concentrated or adsorbed at the surface of that phase is the adsorbate. Adsorption is thus different from absorption, a process in which material transferred from one phase to another (e.g. liquid) interpenetrates the second phase to form a "solution". The term sorption is a general expression encompassing both processes.

Activated carbon is the most widely and effectively used adsorbent. A typical activated carbon particle, whether in a powdered or granular form, has a porous structure consisting of a network of interconnected macropores, mesopores, and micropores that provide a good capacity for the adsorption of organic molecules due to its high surface area. The surface chemistry of activated carbon and the chemical characteristics of adsorbate, such as polarity, ionic nature, functional groups and solubility determine the nature of bonding mechanisms as well as the extent and strength of adsorption. A variety of physiochemical mechanisms/forces, such as Van der Waals, H-binding, dipole-dipole interactions, ion exchange, covalent bonding, cation bridging and water bridging, can be responsible for adsorption of organic compounds in activated carbon (Aksu and Yener, 2001).

In spite of these characteristics, activated carbon suffers from a number of disadvantages (Aksu and Yener, 2001):

It is quite expensive and the higher the quality, the greater the cost.

Both chemical and thermal regeneration of spent carbon is expensive.

Impractical on a large scale and produces additional effluent and results in considerable loss of the adsorbent.

Thus, the research has been active to find alternative and yet efficient sorbents. These adsorbents should have the following properties: the ability to reduce the concentration of pollutants below the acceptable limits, high adsorption capacity and long lifetime. Biosorbents, which are sorbents of biological origin, have proved to be good sorbents for many different pollutants.

3.2 Biosorption

The search for new technologies involving the removal of toxic metals from wastewaters has directed attention to biosorption, based on metal binding capacities of various biological materials. Biosorption can be defined as the ability of biological materials to accumulate heavy metals from wastewater through metabolically mediated or physico-chemical pathways of uptake (Fourest and Roux, 1992). Algae, bacteria and fungi and yeasts have proved to be potential metal biosorbents (Volesky, 1986). The major advantages of biosorption over conventional treatment methods include (Kratochvil and Volesky, 1998 a):

•  Low cost;

•  High efficiency;

•  Minimisation of chemical and lor biological sludge;

• No additional nutrient requirement;

•  Regeneration of biosorbent; and

•  Possibility of metal recovery.

The biosorption process involves a solid phase (sorbent or biosorbent; biological material) and a liquid phase (solvent, normally water) containing a dissolved species to be sorbed (sorbate, metal ions). Due to higher affinity of the sorbent for the sorbate species, the latter is attracted and bound there by different mechanisms. The process continues till equilibrium is established between the amount of solid-bound sorbate species and its portion remaining in the solution. The degree of sorbent affinity for the sorbate determines its distribution between the solid and liquid phases.

3.2.1. Biosorbent material:

Strong biosorbent behaviour of certain micro-organisms towards metallic ions is a function of the chemical make-up of the microbial cells. This type of biosorbent consists of dead and metabolically inactive cells.

Some types of biosorbents would be broad range, binding and collecting the majority of heavy metals with no specific activity, while others are specific for certain metals. Some laboratories have used easily available biomass whereas others have isolated specific strains of microorganisms and some have also processed the existing raw biomass to a certain degree to improve their biosorption properties;

3.2.2. Biomass Sources

Three major sources of biomass can be readily identified:

Waste biomass from industrial large scale fermentations (e.g. from antibiotics, enzyme, organic acid production processes, etc.) (Chubar et al., 2004, Aksu and Yener, 2001; Aksu and Gonen 2004; Aksu and Akpinar, 2001). Basically, industrial biomass comes in the form of amorphous "mud" and requires different types of more or less sophisticated processing into granules of desirable physio-chemical properties before it could be considered as biosorbent (Volesky, 2003).

Microorganisms: A wide variety of microorganisms (both living and nonviable) have been found to be capable of sequestering trace levels of metal ions from dilute aqueous solutions. The nonviable forms have been proposed as potential sorbents, since these are essentially dead materials, which require no nutrition to maintain biomass. Problems associated with metallic toxicity in living biomass and the need to provide suitable growth media also do not arise. Indeed, many early studies have shown that nonliving biomass may be even more effective than living cells. One of the most promising types of biosorbents is marine algae biomass (seaweeds), in view of their high uptake capacity as well as the ready abundance of the biomass in many parts of the world's ocean (Sheng et.al., 2004). Algae, bacteria, fungi, and yeasts have proved to be potential metal sorbents (Veglio and Beolchini, 1997). Many studies have demonstrated the efficiency of metals and organics removal by microbial biomass under a range of physical and chemical conditions (Rao and Viraraghavan, 2002; Denzili et al., 2004; Feng and Aldrich, 2004; Arica et al., 2004; Abu Al-Rub et al., 2004; Pagnanelli et al., 2001; Ibanez and Umetsu, 2004).

Agricultural wastes: Various agricultural products and by-products have been investigated to remove dyes from aqueous solutions. These include cotton waste, rice husk, bark (Mckay et al., 1999), sugar industry mud (Magdy and Daifullah, 1998), peat (Ho et al., 2002), tree fern (Ho et al., 2005), olive pomace (Pagnanelli et al., 2003).

3.3 Biosorption experiments:

Recent biosorption experiments have focused attention on waste materials, which are by-products or the waste materials from large-scale industrial operations. For e.g. the waste mycelia available from fermentation processes, olive mill solid residues (Pagnanelli, et al 2002), activated sludge from sewage treatment plants (Hammaini et aI. 2003), biosolids (Norton et al 2003), aquatic macrophytes (Keskinkan et aI. 2003), etc.

Another inexpensive source of biomass where it is available in copious quantities is in oceans as seaweeds, representing many different types of marine macro-algae. However most of the contributions studying the uptake of toxic metals by live marine and to a lesser extent freshwater algae focused on the toxicological aspects, metal accumulation, and pollution indicators by live, metabolically active biomass. Focus on the technological aspects of metal removal by algal biomass has been rare.

Although abundant natural materials of cellulosic nature have been suggested as biosorbents, very less work has been actually done in that respect.

3.4. Choice of metal for biosorption process:

The appropriate selection of metals for biosorption studies is dependent on the angle of interest and the impact of different metals, on the basis of which they would be divided into four major categories: (i) toxic heavy metals (ii) strategic metals (iii) precious metals and (iv) radio nuclides. In terms of environmental threats, it is mainly categories (i) and (iv) that are of interest for removal from the environment and/or from point source effluent discharges.

Apart from toxicological criteria, the interest in specific metals may also be based on how representative their behaviour may be in terms of eventual generalization of results of studying their biosorbent uptake. The toxicity and interesting solution chemistry of elements such as chromium, arsenic and selenium make them interesting to study. Strategic and precious metals though not environmentally threatening are important from their recovery point of view.

3.5. Biosorption by immobilized cells:

Microbial biomass consists of small particles with low density, poor mechanical strength and little rigidity. The immobilization of the biomass in solid structures Qeates a material with the right size, mechanical strength and rigidity and porosity necessary for metal accumulation. Immobilisation can also yield beads and granules that can be stripped of metals, reactivated and re­used in a manner similar to ion exchange resins and activated carbon.

Chapter 4.Immobilization

Cell immobilization is an attractive technique to fix and retain biomass on suitable natural or synthetic materials support for a range of physical and biochemical unit operations (Abu Al. Rub et al., 2004). Immobilization of the biomass in solid structures creates a material with the right size, mechanical strength, rigidity and porosity necessary for use in unit operations typical of chemical engineering (Veglio and Beolchini, 1997).

The main advantages of this technique include:

Improved biomass performance and biosorption capacity (Aksu and Gonen, 2004).

Increase mechanical strength (Aksu and Gonen, 2004).

Facilitate separation of biomass from pollutant bearing solution (Aksu and Gonen, 2004).

Immobilization can overcome processing problems arising from using powder biomass which in most cases has low density and strength (Abu Al. Rub et al., 2004).

4.1. The principal techniques of immobilization

Various applications are available for biomass immobilization that are available in literature for the application of biosorption are based on adsorption on inert supports, on entrapment in polymeric matrix, on covalent bonds in vector compounds, or on cell cross-linking.

4.1.1. Adsorption on inert supports:

Support materials are introduced prior to sterilization and inoculation with starter culture and are left inside the continuous culture for a period oftime, after which a film of microorganisms is apparent on the support surfaces. This technique has been used by Zhou and Kiff, 1991 for the immobilization of Rhizopus arrhizus fungal biomass in reticulated foam biomass support particles; Macaskie et al. 1987, immobilised the bacterium Citrobacter sp. by this technique. Scott and Karanjakar 1992, used activated carbon as a support for Enterobacter aerogens biofilm. Bai and Abraham, 2003 immobilized Rhizopus nigricans on polyurethane foam cubes and coconut fibres.

4.1.2. Entrapment in polymeric matrices:

The polymers used are calcium alginate (Babu et al. 1993, Costa and Leite, 1991, Peng and Koon, 1993, Gulay Bayramoglu et al. 2002), polyacrylamide (Macaskie et aI., 1987, Michel et al. 1986, Sakaguchi and Nakajima et al. 1991, Wong and Kwok, 1992), polysulfone (Jeffers et al. 1991, Bai and Abraham, 2003) and polyethylenimine (Brierley and Brierley, 1993). The materials obtained from immobilization in calcium alginate and polyacrylamide are in the form of gel particles. Those obtained from immobilization in polysulfone and polyethyleneimine are the strongest.

4.1.3. Covalent bonds to vector compounds:

The most common vector compound (carrier) is silica gel. The material obtained is in the form of gel particles. This technique is mainly used for algal immobilization (Holan et al. 1993, Mah:!mn and Holocombe, 1992).

4.1.4. Cross-linking:

The addition of the cross-linker leads to the formation of stable cellular aggregates. This technique was found useful for the immobilization of algae (Holan et al. 1993). The most common cross linkers are: formaldehyde, glutaric dialdehyde, divinylsulfone and formaldehyde - urea mixtures.

4.2. Desorption:

If the biosorption process were to be used as an alternative to the wastewater treatment scheme, the regeneration of the biosorbent may be crucially important for keeping the process costs down and in opening the possibility of recovering the metals extracted from the liquid phase. For this purpose it is desirable to desorb the sorbed metals and to regenerate the biosorbent material for another cycle of application.

The desorption process should:

•  yield the metals in a concentrated form;

•  restore the biosorbent to close to the original condition for effective reuse with undiminished metal uptake and

•  no physical changes or damage to the biosorbent.

4.3. Supporters

A variety of inert supports has been used to immobilize biomaterials either by adsorption or physical entrapment. Silica gel, an inert and efficient support for microorganisms has been used to immobilize Stichococcus bacillaris for Pb preconcentration and determination by flame atomic absorption spectrometry. This algae-silica material was also used to simultaneous preconcentrate Cu, Cd, Pb, and Zn in simulated riverine water, brine and seawater solutions . Pilayella littoralis, a filamentous free-living brown alga has been previously investigated by Carrilho and Gilbert. The authors describe a series of experiments designated to determine the potential of dead biomass from the marine alga P. littoralis for biosorption of metal from solution in batch systems. The effect of pH on metal uptake and the kinetic of metal sorption were assessed. Metals were bound to the algae within the first 5 min of exposure at pH 5.5 and were efficiently desorbed with 0.12 mol l−1 HCl. In a recent work, Carrilho et al. proposed some procedures to characterize metal binding sites on P.

littoralis using nuclear magnetic resonance (NMR) spectroscopy and Fourier transformed infrared spectrometry (FTIR). The results provided information on the type of functional groups responsible for metal uptake such as carboxylates, ethers, amines and hydroxyls. Metal interaction with this alga and sorption sites competition among metals were assessed by 27Al and 113Cd NMR.

The present work proposes the use of this new P. littoralis-based material for preconcentration in trace metal analysis. Unlike our previous work, biosorption is assessed in lake water samples

4.4. The algae immobilization procedure

Algae were immobilized on silica gel based on the procedure previously reported by Mahan and Holcombe. Samples of 40 mg of clean powdered algae (approximately 50 μm particle size) and 100 mg of silica gel were dried at 80 °C for 20 min, in separate porcelain crucibles, and then mixed. An algae-silica paste was formed by adding a few drops of deionized water to the mixture and blending. The paste was dried at 80 °C for 20 min. The procedure of wetting and drying the algae-silica material was repeated 3-5 times to achieve better immobilization of the alga on the silica. The oven-dried algae-silica matrix was sieved in a plastic strainer to discard free algae not immobilized on the silica and some of the uncovered silica. The silica-immobilized algae were packed into the column and 0.12 mol l−1 HCl solution was peristaltically pumped through at 0.9 ml min−1 for approximately 20 min. Following, the column was conditioned by running through 5 mmol l−1 CH3COOK buffer solution at pH 5.5. A silica column containing no algae was tested with every alga-silica column in order to evaluate possible metal sorption by the silica alone.

Chapter 5: future work

This study proved the technical feasibility of using immobilized inactive algal biomass for the removal of heavy metals from aqueous solutions.

By comparing the principal techniques of immobilization we decided to

use covalent bond to vector compounds technique for biomass immobilization for the application of biosorption to remove a heavy metal .This technique is mainly used for algal immobilization .The selected heavy metal is zinc and we will use silica gel as a carrier.

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