4.3 Biological Recovery and/or Treatment of Heavy Metals 4.3.1 Background Heavy metals are among the contaminant classifications receiving the greatest scrutiny in waste minimization pro
Trang 1of the innovations that have demonstrated the capability of biotechnology to convert a useless waste stream into a viable product.
4.2 Isolated Enzymes and Biocatalysts
While microbial processing may offer effective decomposition of organic pounds, the practice of microbial degradation on a large scale can be complicated
com-by a few inherent limitations (pH sensitivity, nutrient amendments, population dynamics, etc.), depending on the waste stream to be treated In addition, compounds such as organic solvents often present great toxicity toward microor- ganisms Above all of these, most of the recalcitrant organic contaminants usually possess minimal solubility in the aqueous phase, where the microorganisms are hosted and considered the most active Recent advances in biocatalysis have demonstrated that it is feasible to carry out biotransformations in nearly pure organic media (180–182).
Enzymes are biocatalysts, or proteins secreted by microorganisms to erate the rate of a specific biochemical reaction without being consumed in the reaction The key environmental parameters critical to microorganisms must be
accel-in place for producaccel-ing the enzymes from the accel-intact microbial cells The difference between normal microbial degradation and biocatalysis occurs once the enzymes have been harvested After harvesting, the enzymes do not require subsequent nutrient, TEA, or energy source amendments As with intact cells, the enzymes can either be utilized in a free suspension or immobilized on a support Im- mobilized enzymes often show improved stability during variations in environ- mental conditions over free enzymes For example, when laccase was immobilized for treating elevated concentrations of phenol, it maintained over 80% of its activity when subjected to changes in pH, temperature, and storage conditions (183).
Isolated enzymes can also afford much faster reactions, which are usually several orders of magnitude higher than those resulting from traditional microbial process (184) The increase in reaction time has been attributed to the absence of competing processes that are present with the natural metabolism of intact organisms Since the competing processes are absent, enzymes possess a greater specificity toward a particular compound An example of an enzyme’s ability to treat recalcitrant compounds has been demonstrated by the textile industry.
Lacasse is the primary extracellular enzyme produced by Trametes color Lignin peroxidase is one of the many enzymes secreted by P chrysospor- ium Both of these enzymes have been used for the decolorization of the synthetic
versi-dyes contained in textile effluents (20) Utilization of these isolated enzymes yielded greater remediation efficiencies in less time and was independent of microbial growth rates Decolorizations of 80% and over 90% were achieved for anthraquinone dyes with lignin peroxidase and lacasse (20,185) Lacasse was also
Trang 2capable of degrading the reaction by-products present in the textile effluent Lacasse and lignin peroxidase were not the only enzymes that have been used for decolorizing textile effluents Horseradish peroxidases and manganese peroxi- dases have also shown success for decolorizing effluents from the textile and paper industries (186–189).
As stated earlier, enzymes can also possess greater affinity toward a particular compound Therefore, utilization of enzymes as biocatalysts is one of the processes that enable the selective removal of unwanted chemicals in product streams (190,191) For example, researchers have recently utilized enzymes to minimize waste at a cattle dipping facility The cattle dipping liquid must be discarded when potasan, a dechlorination by-product, accumulates to a specified level Due to the high degree of chlorination, the waste cannot be reused Grice
et al (192) employed parathion hydrolase to selectively hydrolyze the potasan Since the potasan concentration was reduced, the use of the dipping liquid was extended, thereby decreasing the amount of waste generated.
Phenol and substituted phenolics are among the constituents present in most industrial wastewaters Due to their prominence in the waste streams and associ- ated toxicity, phenolics have been identified for selective removal via enzymes.
One approach is to use tyrosinase to convert phenol to o-quinones, which are then
easily adsorbed (193) This two-step process was effective even in the presence
of microbial inhibitors When the tyrosine was not immobilized, the remediation efficiency was decreased Other illustrations of treating phenolic wastewaters include the treatment of 4-chlorophenol with horseradish peroxidase (194), pentachlorophenol with horseradish peroxidase (195), and various phenol substi- tutions with immobilized peroxidases (196,197).
4.3 Biological Recovery and/or Treatment of Heavy Metals
4.3.1 Background
Heavy metals are among the contaminant classifications receiving the greatest scrutiny in waste minimization programs These compounds are present in normal municipal wastewater and in various industrial effluents such as those of elec- troplating and metal finishing Not all heavy metals pose a threat to the microor- ganisms used in treatment operations For instance, Fe, Mo, and Mn are important trace elements with very low toxicity, while Zn, Ni, Cu, V, Co, W, and Cr are classified as toxic elements Although toxic, Zn, Ni, and Cu have moderate importance as trace elements As, Ag, Cd, Hg, and Pb have a limited beneficial function to most microorganisms (198).
Activated sludge consortiums and other microbial processes can tolerate most heavy metals at very low concentrations As heavy metal concentration increases, the metals can become harmful to the bacteria (nontoxic), achieve toxicity status, or pass through the system unaltered All three of these scenarios
Trang 3pose a serious threat to microbial and human health When metal toxicity becomes too great, the microbes that are required for degrading the organic compounds perish If metals such as Cu, Cr, and Zn pass through the system and are ingested, chronic human health disorders will arise (199) Recently, scientists have found that certain microorganisms either possess an inherent resistance to heavy metals or can develop the resistance through changing their internal metabolism.
Metal-resistant microbes have potential applications to facilitate a nology process that occurs in the presence of, but does not require, heavy metals Bacteria that can resist toxic metal concentrations could be utilized to convert the organic constituent present into a readily usable or disposable form Microorga- nisms that can do more than simply tolerate elevated metal concentration can
biotech-be used in the recovery of metals or bioremediation of contaminated media (198,200–202) The microbial processes that remediate or recover heavy met- als include leaching (biological solubilization), precipitation, sequestration, and biosorption (203–206) Biosorption has been the most widely studied aspect for waste minimization activities.
4.3.2 Biosorption
Donmez et al (207) defined biosorption as the “accumulation and concentration
of contaminants from aqueous solutions via biological materials to facilitate recovery and/or acceptable disposal of the target contaminant.” This definition can be expanded to depict the difference between active and passive biosorption Active biosorption entails passage across the cell membrane and participation in the metabolic cycle Passive sorption is the entrapment of heavy metal ions in the cellular structure and subsequent sorption onto the cell binding sites (208) Live biomass involves both active and passive sorption, whereas inactive cells entail only the passive mode.
Bioadsorbent materials encompass a broad range of biomass sources, including cyanobacteria, algae, fungi, bacteria, yeasts, and filamentous microbes The biomass can be cultivated specifically for metal sorption, or a waste biomass can be utilized Table 5 includes a brief compilation of the microorganisms studied for the biosorption of heavy metals via passive sorption with dead biomass Although the research contained in Table 5 focused on the sorption of dead biomass, live cells can also be used.
Both live and inactivated (dead) biomass possess interesting metal-binding capacities due to the high content of functional groups contained in their cell walls (39,222) A few differences have been exhibited between the use of live and dead biomass In general, live biomass can accumulate more metal ions per unit cell weight Live cells, if processed correctly, can be reused almost indefinitely For
instance, live Candida sp can sorb (per gram of live biomass) 17.23 mg, 10.37
mg, and 3.2 mg of Cd, Cu, and Ni, respectively (212) The sorption capacity was
Trang 4reduced by over 35% of each metal when inactive cells were used Unfortunately, active sorption with live cells usually takes longer and poses stricter environmen- tal controls than the use of inactivated biomass (223) Moreover, the use of dead biomass from pharmaceutical or other industrial operations is a waste minimiza- tion technique in itself (224) It converts a previously useless waste stream into a viable process step.
Also, as indicated in Table 5, sorption selectivity and specificity will depend
on the source of biomass used Research conducted by Kanosh and El-Shafei (214) showed that fungi had a metal selectivity of Ni >> Mn > Pb > Co > Cu >>
Zn >> Al, whereas bacterial strain selectivity was Pb > Mn > Ni > Zn >> Al > Li
> Cu > Co Isotherm studies with brewer’s yeast (Saccharomyces cerevisiae )
resulted in a sorption order of Pb > Cu > Cd ≈ Zn > Ni (225) When Candida sp.
is used as the source of live biomass, selectivity order changes for three of the metals, to Cd > Cu > Ni The differences in degree of sorption and specificity are attributed to the inherent differences in cellular metabolism and functional groups adhered to the cell walls.
TABLE 5 Examples of a Few of the Yeast, Bacteria, Algae, and Fungi Usedfor the Biosorption of Heavy Metals from Waste Effluents (metals listed inorder of selectivity)
Species Heavy metal ReferencesYeast:
Saccharomyces cerevisiae Cu, Cr(VI), Cd, Ni, Zn 209–211
Trang 5It is important to note that the success of metal biosorption has led researchers to investigate this technique for other waste compounds For example, biosorption has also received a cursory investigation for the decolorization of dye
effluents The yeast strain Kluyveromyces marxianus IMB3 was used to treat
effluents containing azo and diazo dyes The biomass was able to adsorb over 68
mg dye/mg biomass during the 3-day study (226).
4.3.3 Heavy Metal Recovery
Microbial immobilization of heavy metals is used more for the recovery of specific metal ions The ability of the microbes to immobilize the metals is an extension of its normal survival mechanism Initially the microbes capable of immobilization were studied for their ability to transform the compound into a less toxic form (227) The first studies documented the ability of bacteria to change the valence state of soluble chromium from 6 to 3 When the experiment was conducted over a longer period of time, other bacteria were able to facilitate the precipitation of the metals (228) Precipitation occurred as a result of the microbial excretion of organic acids, enzymes, and polymeric substance When the metals have been precipitated, they no longer pose a threat to the microorgan- isms and enable easy recovery.
Biosorption can also be used for the recovery of metals However, the procedure is often more involved and costly than microbial immobilization Recovery from dead biomass entails the digestion of the cellular material, followed by the separation of the different metal ions by chemical or electrolytic methods Washing live cells with different electrolytic and buffer solutions can cause the elution of the bound metal Again, separation and purification of the different metals in the eluted solution can be achieved by traditional chemical or electrolytic methods.
4.4 Minimization of Biomass
As stated in the activated sludge section, part of the biomass (i.e., sludge) is recycled and the remainder is wasted In the past, the wasted biomass was either used as a nutrient amendment for farm lands, compost supplement, or disposed
of in a landfill However, depending on the waste being treated, the degradation by-products and inorganics that have sorbed to the suspended flocs may cause the wasted biomass to be classified as a hazardous waste Even when not classified
as hazardous, changing legislation and rising processing and disposal costs have led to studies that focus on biomass growth to decrease the amount of excess microbial matter (229,230).
The U.S Air Force developed a two-step process to decrease the volume
of biomass disposed of First, the wasted biomass and suspended solids were solublized via a hot acid hydrolysis step After cooling, the solublized material
Trang 6was recycled through the normal activated sludge treatment area Utilization of this process has resulted in a 90% reduction of the hazardous biological solid waste disposed of in controlled landfills (231).
A second innovation was to eliminate excess sludge production by treating
it in the aeration tank The simultaneous activated sludge treatment and sludge reduction was brought about by the addition of ozone In some facilities, ozone
is used to supply the necessary oxygen to the microbial population For this application, care has to be taken to ensure that too much ozone is not added, so that normal chemical oxidation reactions do not occur The phenomenon led engineers to investigate ozonation for the reduction of excess biomass Control- ling the ozone dosage to 0.05 g per gram of suspended solids with a recycle ration
of 0.3 reduced the excess sludge production to essentially zero (232) Although successful, this technique is not fully utilized due to the extensive monitoring and controls needed to ensure that the required population is kept viable and not seared.
Other approaches have encompassed increases in process temperatures or extending the aeration step Unfortunately, neither avenue is economical or particularly effective (233) Perhaps a more efficient and controlled approach would be the direct manipulation of the microbial population As stated earlier, bacterial populations will shift and alter their growth based on the available carbon, energy, and TEA sources If the metabolic pathway is successfully uncoupled, substrate (i.e., contaminant) catabolism will continue, while biomass anabolism is restricted Most microorganisms must satisfy their maintenance energy requirements before reproduction, so excess biomass formation will be reduced (234).
Any easy way to uncouple metabolic pathways and cause a population shift
is to either switch the respiration mode from aerobic to anaerobic or add chemicals to deter growth rate (235,236) For most activated sludge processes, switching the respiration mode would be detrimental to the entire process This approach is recommended only for the facultative anaerobes found in anaerobic digestors (237,238) Chemical additions, however, have proven to be quite successful at minimizing excess biomass formation without jeopardizing the overall process efficiency.
One study utilized nitrate as the nitrogen source instead of ammonia This was a very cost-effective, easily adaptable method that decreased excess biomass generation by over 70% (239) A more unique means of manipulating biomass formation was the introduction of nickel to the raw influent When nickel was added at a concentration of 0.5 mg/liter, the observed cell yield was reduced by approximately 65% (240) Unfortunately, a better biomass reduction could not be achieved If too high a nickel concentration was implemented, steady state could not be attained.
Trang 74.5 Innovative Bioreactors
Manipulation of reactor design can decrease excess biomass formation, increase oxygen transfer, decrease shear stress, enhance contact time, enable treatment of recalcitrant compounds, and improve overall treatment efficiencies (127,229) The incorporation of baffles to bioreactors is known to enhance mixing and decrease shear (241) Although effective, the use of baffles is not considered a new technique One of the most innovative and versatile developments is the use
of membranes Membranes can facilitate a multitude of different scenarios, one
of which employs extractive membranes.
In general, extractive membranes are permeable to organics and virtually impermeable to water and heavy metals, thereby facilitating the biological treatment of the organics contained in mixed waste (198,242,243) Extractive membranes can be modified to allow the selective transfer of specific pollutants into an area where a specialized strain of bacteria will utilize the pollutant as the sole carbon source Brookes and Livingston (112) used this approach to demon- strate the selective degradation of 3,4-dichloroanaline in wastewater A similar methodology was used by Inguva et al (244) to treat TCE and 1,2-dichloroethane contaminated wastewater A sequence of such membranes can be employed
in conjunction with the appropriate bio-areas containing different isms for facilitating the treatment of high-molecular-weight compounds and/or mixed wastes.
microorgan-Traditional activated sludge system efficiencies have been compared to those obtained by membrane bioreactors for high-molecular-weight compounds The membrane bioreactor removed 99% of the chemicals, while the activated sludge system treated only 94% (245) Although this may not appear to be a significant difference, the membrane bioreactor was also highly effective in degrading the degradation by-products, whereas the activated sludge system was not This was due largely to the enhancement of the biomass’ viability by increasing the oxygen transfer when the membrane was in place (115,246) Membrane versatility also allows for high-strength wastes to be treated in both plug flow and completely mixed conditions (247,248).
The successful implementation of membranes is not limited to aerobic systems Previously, ultrastrength wastewater was considered treatable only by a two-stage anaerobic process Membrane anaerobic reactors enable an effective, single-phase treatment to be employed (38) Developments with hollow-fiber membranes have resulted in the simultaneous aerobic-anaerobic treatment of chlorinated compounds (249,250).
Hollow-fiber membranes have also been used as the support system for immobilized enzymes The membranes provide a very large surface area for the immobilized enzymes As with other enzyme applications, the reaction times and subsequent remediation efficiencies are greater than those typically achieved with
Trang 8intact cells This approach has been used for wastewater denitrification, heavy metal recovery, and treatment of chlorinated compounds (251–255).
5 CONCLUSIONS
As shown by the brief examples presented here, biotechnology has demonstrated applicability in all areas of waste minimization—material substitution, selective removal, recycle/reuse of intermediates, end-of-pipe treatment, and source reduc- tion These demonstrations have included traditional processes such as activated sludge and trickling filters to innovative techniques involving heavy metal biosorption and immobilized enzymes With continued advances in analytical and technological expertise, effluent constraints will promulgate to become more stringent Biotechnology will continue to rise to the challenge As scientists and engineers strive to learn more about microorganisms and their metabolic path- ways, they will be able to convince Mother Nature to do as they wish Natural approaches (i.e., green chemistry) are more effective and acceptable Further- more, if complete mineralization of waste effluents is achieved, or successful biosubstitutions are made, a secondary stream will not have to undergo subse- quent treatment as with other technologies, thereby eliminating the need for waste minimization.
i influent condition
k maximum specific substrate utilization rate, d–1
kd microbial decay coeffient, d–1
K Monod saturation constant, g/m3
m,n emperical constants in Eckenfelder equation
Ns number of stages (RBC units) used OLR organic loading rate
Trang 9T water temperature, ˚C TEA terminal electron acceptor
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