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In the last few decades,environmental pollution caused by various hazardous materials especially the heavy metals has been a topic of management and has been monitored closely. Removal of heavy metals from soil and water around industrial plants has been a major challenge. For the removal of theseheavy metals from liquid wastes various methods have been adopted.Some of them includes precipitation,evaporation,membrane process etc.These methods have however several disadvantages like unpredictable metal ion removal,requirement of huge amount of reagent, disposal of toxic wastes and sludge etc.Biosorption is a process, which involves a biotechnological innovation,advancement anddevelopment of a cost effective method for the removal of heavy metals.The present article contains the overview of the biosorption work carried outon bacteria,which could serve as an economical means and eco-friendly method of toxic metal removal. Present work was done as a solution to various environmental threats caused by heavy metals .Analysis of results conferred that the bacterial species (S.aureus and E. coli) have the ability to biosorb heavy metals.
In present era heavymetal has posed serious and deleterious impact on the environment. Presence of even trace amounts ofheavy metals can be prove to be toxic to both flora and fauna. Development of huge industrial sectors like mining,energy and fuel producing etc has resulted in the release of wastes containing metals into the environment.These metals are dumped either directly or indirectly in open area,thus causing serious environmental pollution.This affects human life and natural vegetation..Numerous methods are now presently used for the removal of heavy metal ions from aqueous wastes(chemical precipitation, ion exchange,electro chemical treatment, membrane technologies,adsorption on activated carbon etc.). Some techniques like chemical precipitation or electrochemical treatments etc. are ineffective, especially at low metal concentrations.Moreover such treatments producesa large amounts of sludge which are undisposable andhence causes treatment problems.Besides,other techniques likes Ion exchange,membrane technologies etc. are effective but are economically les favourable.Therefore,the search for a new cost effective technologies and eco-friendly approach for heavy metal removal has led to development of biosorption.The main principle of biosorption is that, this process ismainly based upon the metal binding capacities of various biological materials(G.M.Gadd,1990).Biosorption can be defined as the ability of any biological materials(Algae,bacteria,fungi and yeasts ) to accumulate or absorb heavy metals from waste,either through metabolic activity or by any physico-chemical pathways.Biological species like Algae,fungi,bacteria and yeasts etc. have proved to be a potential metal biosorbents (F.Veglio et al., 1997). The actual process of biosorption involves two agents- a solid phase(biosorbent/biological material) and a liquid phase (solvent usually water)whichcontains ametals which has to be adsorbed.. The very high affinity of the sorbent for the sorbet speciesresults in the binding of metals to the biosorbent species.This binding follows different mechanisms. This binding process continues untilan equilibrium is reached between the amount of sorbed and its quantity present in the solution.The chemical affinityof biosorbentmaterial for the metal salts, determines their distribution.This ability of these microorganisms for the accumulation of metal depends mainly on the chemical and biological composition of cells.Some microbial species can accumulate wide variety of heavy metals while some are specific only towards certain heavy metal(pagnamelli,et al.2002).The commonly used biosorbents includes some waste mycelia (fermentation) ,solid mill residues ,activated sludge from sewage plants(Hammaini et al.2003),biosolids and aquatic algae (keskinkan et al.2003).
Thegeneral mechanism of biosorption can also be classified according to cell'smetabolism. These are 'metabolism-dependent' and 'metabolism independent' mechanism.The metals which are removed from metal solution are found either outside the cell(extra-cellular accumulation/precipitation) or inside the surface of cell. (surface sorption/precipitation) or within the cell(intracellular accumulation).
Intracellular accumulationwill occur when the heavy metal is transported across the or through the cell's membrane. This sort of process is dependent upon the cell's metabolic activity.Normallyviable cells and living cells are involved in such type of biosorption process.These type of processis generally associated with the cell's(microbial species) active defense mechanism .In case ofmetabolism independent biosorption,the uptake of heavy metal is mainly achieved by various physico-chemical interaction which occurs between the heavy metals and the various functional groups present on cell'ssurface.Chemical sorption, Physical adsorption, ion exchangeetc. are examples of the non metabolism dependent biosorption.(P.King,et al.,2007).Biosorption is generally a two-step process:
1. Passive biosorption: This is actually a metabolism independent process which uses either one or a combination of the following processes:
• ion exchange
• physical adsorption
• Inorganic micro precipitation
It involves dynamic equilibrium of a reversible adsorption-desorption mechanism. The metal ions bounded onto the surface of biosorbent can be eluted with the aid of other metal ions, or organic acids or chelating agents. Various chemical functional groups present onto the surface of various biosorbent facilitates the above mentioned processes.Some such functional groups includes the acetamido groups present in chitin, polysaccharides present in fungi, amino or phosphate groups present in nucleic acids; amide, amino,etc,present in proteins.
2. Active biosorption: During thisphase of biosorption, the metal ions will penetrate within the cell's membrane and thus enters into cell.
On the basis of position or location of metal sorbed,bio sorption are of following types:-
Extra cellular metal accumulation or precipitation
Intracellular metal accumulation
Cell- surface sorption or precipitation.
Various factors influences the processof biosorption.The optimum temperature required for the biosorption process ranges from 25-30o C. pH is also the most vital parameter which influences.This is because,pH affects solution chemistry of metals,or the functional groups present in biomass and their competition for metallic ions. The biomass concentration present in solution also influences the specific uptake rate,for lower biomass concentration,there is average increase in uptake rate.This is because, any increase in concentration of biomass will lead to the interference between various binding sites. This hypothesis This phenomenomena was attributed to decrease in the concentration of metal or their shortage in solution. Henceforth this factor has to be noted while using any microbial species as biosorbent.Also the presence of various multiple metal ions in a solution will also affect the biosorption mechanism.
In our present work, we worked with bacterial species, where the bio sorption is mediated by the metabolism independent mechanism .Here the metal uptake was mainly by various physical- chemical interactions between the sorbed metal and the functional groups present on cell surface of microorganism.. This processis based on various physical adsorption,chemical sorption,or ion exchange process which are non metabolism dependent.The cell walls aremainlymade of polysaccharides,lipids or proteins.These have various metal binding sites.(carboxyl groups,amino groups etc.)
In physical adsorption process,biosorption occursmainly due to the Vander Waal's forces of attraction. In these processes, the metals can be adsorbed by dead /non-viable biomaas.The biosorption is mainly by the electrostatic interactions occurring between metals present in solutions and the walls of microbial cells. The ion -exchange process mainly depends upon thecomposition of cell wall.Themicrobial cell wall mainly contains polysaccharides,lipids etc.The metal ions gets counter exchanged with the ions of the polysaccharides and hence results in uptake of heavy metals.
In complexation mechanism, themetal removal occurs by a complexation. This occurs mainly due to interaction between various metals and the functional groups present in cell's membrane.This mechanism was found in Cu, Mg accumulation by various strains of Pseudomonas sp.
In our work have considered the initial metal concentration and biomass concentration. The biomass concentration was found to influence metal uptake rate.
MATERIALS AND METHODS
Preparation of Biomass
Two bacterial species (E. coli, S. aureus)were collected from laboratoryand were inoculated separately in freshly prepared nutrient broth. Two ml. of the overnight grown culture was inoculated in the newly prepared nutrient broth separately. O.D. of the inoculated broths was measured immediately after the inoculation .It was found to be 0. O.D values were measured at different intervals of time for both the species.E. coli grown medium was collected at O.D. values of0.23 and 0.5.whereas S. aureus was collected at O.D. corresponding to 0.3 and 0.6.Culture collected and then centrifuged at 6000rpm for 10 min .The pellets of different bacterial species were then mixed or dissolved into the freshly prepared metal stock solutions.
Stock solutions of metal ions
Stock solution of two different metals namely Ni and Cr. wasprepared in deionized water. The two metals were dissolved to make different concentration of (50mg/100ml,100mg/100ml and150mg/100ml ).These solutions were then stored in bottles (Volesky,2003)
Solutions in a fixed volume(100ml) with varying concentrations of Cr and Ni salts were mixed or dissolved with two different quantity of biomass,in a conical flask. These flasks were then kept on rotating shaker.At different time intervals (2hrs.,24 hrs, and 48 hrs.) 1 ml samples were collected for metal analysis.
Different concentrations of the heavy metals in the solutions were determined at different time intervals (2hr, 24hr,48hr) by using Coleman 395D- UV-Visible spectrophotometer(Japan).The standard graph was plotted forNi and Cr by using dimethyl glyoxime and potassium dichromate . O.D. of Ni(II) was taken at 530nm (Padmavathy et al.,2003) and O.D.of Cr(III) at 440nm (Vogel et al.2000).
RESULT AND DISCUSSION:
Fig.1Percentage Removal of Ni using E. coli at three time intervals when exposed to three different amounts of metal.(low biomass)
Fig.2 Percentage Removal of Cr using E.coli at three time intervals when exposed to three different amounts of metal.( High biomass)
Fig.3.Percentage of Cr using E. coli at three time intervals when exposed to three different amounts of metal (Low biomass)
Fig.4 Removal of Ni using E.coli at three time intervals when exposed to three different amounts of metal (High biomass).
Fig.5 Percentage Removal of Ni using S.aureus at three time intervals when exposed to three different amounts of metal (low biomass).
Fig.6percentage removal of Ni using S.aureus at three time interval when exposed to three different amount of metal (High biomass)
Fig.7 Percentage removal of Cr using S.aureus atthree time interval when exposed to three different amount of metal (Low biomass)
Fig.8 Percentage removal of Cr using S.aureus at different time intervals when exposed to three different amount of metal(High biomass)
Fig.9 Percentage Removal of Ni and Cr using E.coli at three time intervals when exposed to three different amounts of metal(High biomass)
Fig.10Percentage Removal of Ni and Cr usingE. coliat three time intervals when exposed to three different amounts of metal (Low biomass).
Fig.11 PercentageRemoval of Ni and Cr using E.coli at three time intervals when exposed to three different amounts of metal(High biomass).
Fig.12Percentage Removal of Cr using E .coliandS. aureusat three time intervals when exposed to three different amounts of metal (Low biomass).
Fig.13 PercentageRemoval of Cr using E.coli and S.aureus at three time intervals when exposed to three different amounts of metal (High biomass)
Fig.14 Percentage Removal of Ni using E.coli and S.aureus at three time intervals when exposed to three different amounts of metal( Low biomass).
Fig.15 Percentage Removal of Ni using E. coli and S. aureusat three time intervals when exposed to three different amounts of metal (High biomass).
RESULT AND DISCUSSION:
In fig.1 it is seen that the percent removal of Ni increases with increase in metal concentration, for low biomass of E. coli. For high biomass of E. Coli ,the overall percent removal increased with increase in metal concentration.(fig.4).(Jamil and Yusoff, 2000).In case of Cr. It was observed that percent removal was almost constant for low biomass (fig.3) and for high biomass, the removal percent, with increase in metal concentration. Such decline in percent removal rate is mainly due to saturation of adsorption sites, which occurs with an increase in time duration.
In case of S. aureus , it was observed that, for low biomass, with increase in time duration and increase in metal concentration, the percent removal rate for Ni slightly decreased. But overall removal rate percent was constant (fig.5). For higher O.D.it was observed that percent removal increased with increase in metal concentration and time duration (fig.6).This was mainly because the surface saturation of the biomass is also dependent upon the metal's initial concentration.(Jamil and Yusoff, 2000).
In case of Cr., it was found that overall percent removal rate increased with increase in metal concentration and time duration for low biomass of S. aureus whereas, for high biomass it was found that similar trend was observed.(fig.8).
High biomass of E. coli species were found to be more efficient in Ni removal than Cr.(fig.9) and its removal efficiency increased with increase in time and metal concentration. But for Cr., it was found that removal efficiency decreased overall. Similar trends were observed for low biomass of E. coli (fig.11)
E. coli is more efficient in removing Cr at 50mg (fig.12) however the efficiency decreased with increase in metal concentration. This occured mainly because the surface saturation of the biomass is also dependent upon the initial metal's concentration.
At high biomass it was found that E. coli at 50 mg. and S. aureus at 150 mg was found to be more efficient (fig.16).in removal of Ni. This was because biosorption also depends uponthe initial metal concentration. For low biomass, E. coli was more efficient in metal removal (fig.15)
S. aureus was found to be more efficient in removal of Cr. at 100 mg.(fig.14). Similar trends were observed at low biomass.
From the overall result,it was conferred that the percent removal of heavy metals by the bacterial sp(E. coli and S.aureus) increased with the increase in time duration and metal concentration.This occurs mainly due to surface saturation of biosorbent.(Jamil and Yusoff, 2000)(fig.1,3,4,5,6,7,8,9)
In fig.(3,5,7,8,15) it was found that the metal removal rate decreased with increasing metal concentration. Such reduction in % removal rate is mainly due to saturation of adsorption sites.(R.Avavindhan et.al;2009)
In fig(11 ,12) % removal rate decreased with increasing time duration the occurred mainly because of saturation of site of accumulation for metal ion on bacterial cell surface.In the work of Ilhan et.al; he founded that Staphylococcus species where more potent for Cr removal than Ni.In our work it was found that on an average S. aureus were more potent for Cr removal (Fig:13).
Rabin et al; also reported the biosorption of Cr by gram positive bacteria ,he found that these bacteria were more capable for Cr bioabsorption.In our work it was reveal that S. aureus exhibited more potent for Cr removal.(Fig:9 ,11).Ozdemir et.al.,reported that gram negative bacteria were more tolerant for Cr than Ni. In our work it was found that average tolerance for Ni was similar for both species E.coli and S.aureus. It is due to potential binding sites present in membrane.Huang and Huang (1996) found that Ni removal efficiency for by A.oryzae was enhanced by pretreatment with Hcl04,but in our work Ni removal efficiency was only 50%.In the work of A.J.Morais,Barros,S.Prasad(2005)average removal of 83.4% Ni removal was found but in our work only 605 Ni were removed.This was due to varying environmental parameters.
Various research over past few decades have provided a better and effective understanding of the metal bio sorption by certain potential bio sorbents. Various eco-friendly and cost effective biosorbent materials also includes the waste biomasses. This process has a major advantage in decontamination of heavy metals from industrial effluents. Application aspects of bio sorption are being aimed at bio sorption process optimization. Various mathematical models are used in biosorption process. For further future experimental works in this aspect and to predict the efficiency of performance of biosorption process, various strategies have been developed to analyse its performance under various operating conditions, to exhibit more usage of biosorption process.