1. TTrreeaattmmeenntt ooff IInndduussttrriiaall WWaassttee WWaatteerr::BBiioollooggiiccaall RReemmeeddiiaattiioonn ooff CCyyaanniiddeessDR. CHANDRAJIT BALOMAJUMDERPROFESSORDEPARTMENT OF CHEMICAL ENGINEERINGINDIAN INSTITUTE OF TECHNOLOGY ROORKEE, INDIA

2. CCYYAANNIIDDEE Carbon-nitrogen radical and refers to all CN (-CN) groups thatcan be determined analytically as the cyanide ion, CN- Highly toxic Found in a wide variety of organic and inorganic compounds Produced by certain bacteria, fungi, and algae Important form of nitrogen for microorganisms, fungi and plants Used in various industrial applications Produced as wastes and emissions from industries in largequantities 3. CCLLAASSSSIIFFIICCAATTIIOONN OOFF CCYYAANNIIDDEE CCOOMMPPOOUUNNDDSSTypes of cyanide Examples RemarksK Ionize easily, 2Zn(CN)4, K2Cd(CN)Weak 4K Ionize moderately 2Cu(CN)3, K2Ni(CN)4ModeratelystrongK Dont ionize easily, very stable 2Fe(CN)6, K3Co(CN)Strong 6Inorganic SCN-, CNO- Cyanate unstableAcetonitrile, Acylonitrile, StableOrganic Aliphatic Adiponitrile, Propionitrile(Nitriles)Aromatic Benzonitrile StableComplexZn(Cn)2, Cd(CN)2Insoluble , AgCNIonize in aqueous solution atlow concentration and mostlypresent as HCN below pH 8NaCN, KCN, Ca(CN)Soluble 2SimpleEquilibrium depends on pH,HCN, CN (pKa, 9.2 at 25oC) Free - 4. STABILITY AND TTOOXXIICCIITTYY OOFF CCYYAANNIIDDEESS The stability of cyanide salts and complexes are pH dependent Toxicity depends on their chemical form stability bioavailability to the exposed microbes and animals Cyanide compounds remain in the aqueous phase and formcomplexes with other metallic contaminants Metal-cyanide complexes are much less toxic than freecyanide, but their dissociation releases toxic free cyanide aswell as the metal cations 5. TTooxxiicciittyy lleevveell ooff VVaarriioouuss CCyyaanniiddee CCoommppoouunnddssLD50 Physical TLVformCompoundHydrogen cyanide (HCN) Gas 5 mg/m3 1 mg/kg humanPotassium Cyanide (KCN) Solid 5 mg/m3 10 mg/kg rat, 2.85 mg/kg humanSodium cyanide (NaCN) Solid 5 mg/m3Cyanogen chloride (CNCl) Gas 0.3 ppm6.44 mg/kg rat,2.85 mg/kg humanSodium Cyanate (NaCNO) Solid 260 mg/kg micePotassium Cyanate (KCNO) Solid 320 mg/kg micePotassium ferricyanide Solid 1600 mg/kg rat(K2 [Fe(CN)6])TLV threshold limit value is the time-weighted average concentration for an 8-hour workday and 40-hourworkweek to which a worker may be repeatedly exposed without adverse effect.LD50 lethal dose to 50% of a specified population. 6. EFFECT OOFF CCYYAANNIIDDEE CCOOMMPPOOUUNNDDSS Exert direct toxicity to fish and higher forms of life includinghuman (acute and chronic effects) Acute effect of cyanide include rapid breathing Gasping Arrhythmia Coma eventual death Chronic effects include acute effects neurological changes impairment of thyroid function demyelization of nerve fibers 7. Potent inhibitors of cellular metabolism - causing problem insewage treatment The biological activity of microorganisms that digest sewageand sludge is lost at 0.3 mg/l cyanide Binding of cyanide to metallic cofactor cause inhibition toactivities of metalo-enzymes Free cyanides inhibit cytochrome oxidase and suppressaerobic respiration Free cyanide found to be highly toxic to methanogenic bacteria,causing problem for anaerobic degradation of cyanide 8. INDUSTRIES USE CYANIDE ARE Electroplating Mining (extraction of Gold, Silver etc.) Case Hardening Automobile Manufacturing Printed Circuit Board Manufacturing Steel and Cock plant Petrochemical Refining Synthesis of Organic Chemical and Synthetic Fibers Paint, Ink formulation and Plastics Aluminum Works Explosives manufacture Pesticides etc. 9. Malononitrile(lubricating oiladditive)Propionitrile(solvent)PetroleumFumigant poison Cyanogen,gas,Zinc cyanideCopper cyanideCalcium cyanideHydrogen cyanideAmmoniumthiocyanate(pesticides)parasiticideCyanogenbromideinsecticides,Cvanogenchloridepesticides,Sodium cyanide,MalononitrileCyanogen bromideBarium cyanideCalcium cyanideFerrocyanide (usedas a flotation agentfor copper andlead/zincseparation)Ammonium MiningthiocyanatePotassium- or sodium- Herbicidescyanide(degreasing)Propionitrile (solvent,dielectric fluid)Nickel cyanideSilver cyanideBarium cyanideZinc cyanideCopper cyanideHydrogen cyanideCyanogen chloride (metalcleaner)Mercuric potassiumcyanide (mirrormanufacturing)ElectroplatingPotassiumferrocyanideAdhesives Ammonium thiocyanate Cement stabiliser Calcium cyanide Fire retardantPrimary cyanidecompounds used inthe processPrimary cyanide Industrycompounds usedin the processPrimary cyanide Industrycompounds used in theprocessIndustryUse of cyanide compounds in various industries 10. Primary cyanidecompounds used inthe processFerricyanideFerrocyanidePropionitrileAmmoniumthiocyanate(ingredient inantibioticpreparations)AmmoniumthiocyanatePotassiumferrocyanidePrimary cyanide Industrycompounds usedin the processPharmaceuticals(includesantibiotics, andnonprescriptionprescriptionsteroids, drugs)Rocket and missilepropellantCyanogenWineFerricyanide,FerrocyanideFerricferrocyanide(Prussian blue,MalononitrileMercuric cyanide(germicidal soap)Copper cyanide(marine paint)SodiumferrocyanideFerricferrocyanide(Prussian blue,PotassiumferrocyanidePrimary cyanide Industrycompounds used in theprocessPigments, paints,dyes, ink,personalcare productsFerricyanide bleachMercuric cyanideHydrogen cyanideMalononitrile Adiponitrile Road salt(intermediate in themanufacture of nylon)Cyanogen bromideCyanogen chlorideHydrogen cyanide(production of nylonand other syntheticfibers and resins)Ammonium thiocyanate(improve thestrength of silks)IndustryPhotographySynthetic fiberacrylic fiber,nylon,syntheticrubberSource: Data from MPI, Final Technical Memorandum: Summary of cyanide investiation a SRWTP and preliminary conclusions andreconmiendations, report by Malcolm Pirnie. Inc. Emeryville, CA to the Sacramento Regional County Sanitation District,Sacramento Reg:c: Wastewater Treatment Plant, Regulatory Compliance Group, Sacramento, CA, 2004. 11. Cyanide levels in wastewatersIndustrial waste water source Simple Complex Total % Simplec Ref. NoCoke-oven waste water 54.8b [5]Coke plant 100-1000 [6]Coke plant 1.6-6.0Coke plant 0.1-0.6Coke plant 0.1-0.725.4a 25.7 a 1.17Coke plant 0.3Coke oven plant 10-150a [7]Coke plant waste 10-38.1 [8]Coke plant waste 91-110 [9]Coke plant waste streams [10]Coke oven liquor 0-8Decantation tank 8Final cooler condensate 196Benzole separator 2736Oil generation plant separator 104Spent limed liquor 4Coke plant ammonia liquor 2-44.5 [11]Coke plant ammonia liquor 20-60 [9]Coal conversion (synthane) 1-6 [9]Coal conversion wastes 2-30 [9] 12. Industrial waste water source Simple Complex Total % Simplec Ref. No0.0-0.2a 0.03-0.27a 26-100 [12]Electroplating plants 0.03-0.07Electroplating plants 0.01-14.24b [13]Electroplating plants/PCB plants 3.0-59.0b [14]Electroplating plants 3.6-6.6 [15]Plating rinse 0.3-4 [8]32.5 [8]25 [9]60-80 [16]30-50 [17]1.4-256 [8]Plating industries(rising waste) 1.4-256 b [18]Plating industries (plating bath) 4000-100 000b [12]Plating bath 30000 [19]Plating bath 45000-100000 [20]Plating bath [11,21,22]Brass 16000-48000Bronze 40000-50000Cadmium 20000-67000Copper 15000-67000Silver 12000-60000Tin-zinc 40000-50000Zinc 4000-64000Alkaline cleaning bath 4000-8000 [17] 13. Industrial waste water source Simple Complex Total % Simplec Ref. NoBlast furnace scrubber water [9]Steel making segment (range) 0.2-1.4Steel making segment (max) 2.4Blast furnace gas wash 48.5Steel mill Coke plant liquor 7.5-39.6 [9]Color film bleaching process 71 [23,24]Paint and ink formulation 0-2 [9]Bright dip 15-20 [17]10.4-50.9a 10.4-50.9a 0-0.3 [25]Chemical industry 0-0.03Gold ore extraction 18.2-22.3 [9]Explosives manufacture 0-2.6 [9]2.25a 2.25a 0 [12]Oil refinery 0.0Petroleum refining 0-1.5 [9]a Includes thiocyanate.b Unspecified whether thiocyanate included.c % of total cyanide concentration 14. Standards for cyanide level in water and wastewater:US-health service cites 0.01mg/l as guideline and 0.2 mg/l aspermissible limit for cyanide in effluentMinimal national standard (MINAS) for cyanide in effluent - 0.2mg/l in IndiaU.S.EPA standard for drinking and aquatic-biota watersregarding total cyanide are 200 and 50 ppb respectivelyLimit for cyanide In Mexico is 0.2 mg/lGerman and Swiss regulations have set limit of 0.01 mg/l forcyanide for surface water and 0.5 mg/l for sewers 15. CYANIDE REMOVAL TTEECCHHNNOOLLOOGGIIEESS Alkaline-chlorination-oxidation Hydrogen peroxide oxidation Ozonation Electrolytic oxidation Ion exchange Acidification AVR Process Lime-sulfur method Reverse osmosis Activated carbon adsorption Caros Acid method Thermal hydrolysis INCO Process (by SO2/Air) Biological Treatments 16. METHODS USED IN TTHHEE PPRREESSEENNTT SSTTUUDDYYAdsorptionBiodegradationSimultaneous Adsorption and Biodegradation 17. AADDSSOORRPPTTIIOONN Effective method for cyanide removal Used mostly as a polishing process Cyanide species exhibit surface reactivity with certain mineralsolids and with activated carbon Granular/Powered activated carbon is the most widely usedadsorbent for cyanide complexes Activated carbon particle has a porous structure consisting of anetwork of inter connected microspores and macrospores thatprovide a good capacity for the adsorption due to its highsurface area 18. BBIIOODDEEGGRRAADDAATTIIOONN Natural approach Relatively inexpensive No chemical handling equipment or expensive control needed Cost is fixed with greater volumes of waste also Biomass can be activated by aeration Can treat cyanides without generating another waste stream No toxic byproducts, hence environmental friendly Used as nitrogen or both carbon and nitrogen source bymicrobes Microbes convert cyanide enzymatically to ammonia, which isreadily assimilated into cellular nitrogen 19. Biodegradation can be possible under both anaerobic andaerobic conditions Aerobic conditions are maintained due to the toxicity of cyanidesto methanogenic bacteria under anaerobic conditions Attach growth processes have better toxicity tolerance thansuspended growth Various strains of microorganisms have been found for thedegradation of cyanide and metal cyanides Biodegradation occurs through specific enzymes and pathways 20. Microbes for Cyanide biodegradation studiesP.putidaFusarium solaniP.fluorescens immobilized on Calcium AlginateP.fluorescens in presence of Glucose (conc=.465g/l)Klebsiella oxytocaMixed culture of bacteriaP.putida immobilized on sodium alginateFusarium oxysporum & F solaniP.putida immobilized on Ultrafiltration membranesCitrobacter sp , Pseudomonas spFusarium oxysporum immobilized on Sodium alginate,Methylobacterium spStrains of Trichoderma spp 21. Microbes for Cyanide biodegradation studiesP.fluorescens immobilized on zeoliteBurkholderia Capcia stain C-3B. stearothermophilus NCA 1503Mix of (1) F.Solani T.polysporum, (2)F.oxyspoum, Scytalidiumthemophilum, Pencillium miczynskiGranular CyanidaseBacillus magateriumE.Coli BCN6Stemphilium lotiPseudomonas fluorenscens NCIB11764 (CN) Pseudomonasgr (phenol)S. loti, G. SorgiPseudomonas putida BCN3Bacillus pumilis (clay,purchage, filamentous develp)Pseudomonas Acidovorans 22. PATHWAYS FOR BIODEGRADATION OF CYANIDE AND THIOCYANATEHydrolytic reactionsCyanide hydrataseHCN + H2O HCONH2CyanidaseHCN + 2H2O HCOOHNitrile hydrataseR-CN + H2O R-CONH2NitrilaseR-CN + 2H2O R-COOHOxidative reactionsCyanide monoxygenaseHCN + O2 + H+ + NAD(P)H HOCN + NAD(P)+ + H2OCyanide dioxygenaseHCN + O2 + 2H+ + NAD(P)H CO2 + NH3 + NAD(P)+Reductive reactionsHCN + 2H++ 2e CH=NH + H2O CH2=OCH2=NH + 2H+ + 2e CH3-NH + 2H+ + 2e CH4 + NH3Substitution/Transfer reactionsCyanoalanine synthaseCysteine + CN -cyanoalanine + HS2OAS + CN -cyanoalanine + CH3COOThiosulfate:cyanide sulfurtransferaseCN + SO2 SCN + SO2233Thiocyanate biodegradationCarbonyl pathway (thiocyanate hydrolase)SCN + 2H2O COS + NH3 + OHCyanate pathway (cyanase)SCN + 3H2O + 2O2 CNO + HS HS + 2O2 SO42 + H+CNO + 3H+ + HCO3 NH4+ + 2CO2The general categories of chemical reactions responsible for the biodegradation of cyanide and thiocyanate. For the hydrolytic reactioninvolving nitriles, R represents either an aliphatic or aromatic group. The substitution/transfer reaction catalyzed by cyanoalanine synthasecan also use O-acetylserine (OAS) as a substrate. The cyanate formed by cyanide monoxygenase is converted to NH4 and CO2 by thesame pathway as the cyanate from thiocyanate. The reductive pathway is derived from the action of nitrogenase and the productsresulting from the transfer of pairs of electrons. 23. SIMULTANEOUS AADDSSOORRPPTTIIOONN AANNDDBBIIOODDEEGGRRAADDAATTIIOONN ((SSAABB)) Adsorption onto adsorbent reduces the inhibitory effect of thecyanides for microbial mass Presence of activated carbon increases liquid-solid surfaces, onwhichmicrobial cellsenzymesorganic materialsoxygenare adsorbed providing an enriched environment for microbialmetabolism Activated carbon can be partially regenerated bymicroorganisms while the carbon bed is in operation 24. Carbon adsorption capacity, controlled by the bioregeneration,highly increased Carbon adsorption column cycle prolonged as compared to pureadsorption system alone Stable performance of the combined process during peak loadbecause the reserve of adsorption capacity due tobioregeneration The adsorbed cyanides desorbed back in to the biofilm and alsothrough it into the liquid phase and become accessible to themicrobial degradation Both processes in one unit results in a better removal andprocess performance SAB process has been utilized for treatment of lots ofsubstances, but fate of cyanide removal is not established 25. OBJECTIVE OF PRESENTWORK 26. Effect of process parameters such as: pH Temperature Contact time Initial concentration of cyanideon removal of Sodium, Zinc and Iron Cyanide fromaqueous solutions by Adsorption Biodegradation Simultaneous adsorption and biodegradation 27. MMAATTEERRIIAALLSS AANNDDMMEETTHHOODDSS 28. Sodium Cyanide [NaCN] Simple alkali salt of cyanide Highly soluble in water and dissociate to release CN- Highly toxic due to ionization in aqueous solutions Produced largely from Electroplating (for degreasing) andMining industries Stock solution of 1 g/L was prepared by dissolving 1.88 g/LNaCNin Milli-Q water (Q-H2O, Millipore Corp. with resistivity of18.2Mcm) 29. Zinc cyanide [Zn(CN)42- ] Weakly stable complexes of metal cyanide Classified as weak-acid dissociable (WAD) as they are easilydissolved under mildly acidic conditions (pH = 4 - 6) Dissociation to release free cyanide Produced from Electroplating, Pesticide industries Stock solution of 1 g/L was prepared by mixing3.57 g/L of autoclaved Zinc sulphate salt solutionsand 3.25 g/L of filter-sterilized KCN solutionin Milli-Q water (Q-H2O, Millipore Corp. with resistivity of18.2Mcm) to obtain K2Zn(CN)4 30. Ferro cyanide [Fe(CN)64- ] Highly stable complex Dissociate in highly acidic condition (pH