Wastewater Treatment Jae K Jim Park Department of

Wastewater Treatment Jae K. (Jim) Park Department of Civil and Environmental Engineering University of Wisconsin Madison 1

Microorganisms – Classifications Energy l Solar radiation: Photo synthetic autotrophs, e. g. , algae l Organics: Heterotrophs, e. g. , activated sludge biomass, denitrifiers, etc. l Inorganics: Chemoautotrophs, e. g. , nitrifiers Oxygen use l Obligate (strict): use only one condition for growth l Facultative: use either dissolved oxygen or chemically derived oxygen (from nitrate, sulfate or carbonate) for respiration and use organic materials for energy and growth Temperature l Psychrophiles: < 20°C. , opt. 13°C l Mesophiles: 20~45°C, opt. 35°C l Thermophiles: 45~60°C, opt. 55°C 2

Organic Matter Energy for Mircoorganisms l Carbonaceous Energy: Carbon as energy source ØHeterotrophs l Nitrogenous Energy: Nitrogen as energy source ØChemoautotrophs 3

Energy Measurement (1) l Theoretical Oxygen Demand (Th. OD) l Chemical Oxygen Demand (COD) l Biochemical (Biological) Oxygen Demand (BOD) ü Carbonaceous BOD (C) ü Nitrogenous BOD (N) l Total Organic Carbon (TOC) 4

Energy Measurement (2) Theoretical Oxygen Demand (Th. OD) 1. Carbonaceous demand: C CO 2; N NH 3 2. Nitrogenous demand: NH 3 HNO 2; HNO 2 HNO 3 3. Th. OD = O 2 req. in steps 1& 2 Ex. Glycine (10 mg/L) [CH 2(NH 2)COOH] (MW = 75 g/mol) 1. Carbonaceous demand CH 2(NH 2)COOH + 1. 5 O 2 2 CO 2 + H 2 O + NH 3 2. Nitrogenous demand NH 3 + 1. 5 O 2 HNO 2 + H 2 O; HNO 2 + 0. 5 O 2 HNO 3 3. Th. OD = [1. 5 + (1. 5+0. 5)] mol O 2/mol glycine = 3. 5 × 32 g O 2/mol = 112 75 g/mol = 1. 49 g O 2/g glycine Thus, Th. OD = 1. 49 x 10 mg/L = 14. 9 mg/L Cannot be used if chemical composition is not known. l 5

Energy Measurement (3) l Chemical Oxygen Demand (COD) O 2 req. for oxidation of organics ü Oxidize carbonaceous matter with a strong oxidant (e. g. , hot dichromate sol. with sulfuric acid) ü Heat Catalyst silver sulfate Dichromate H 2 SO 4 Reduction of O 2 4 e + 4 H+ + O 2 2 H 2 O 1 mole of O 2 (32 g) 4 e equivalents 1 g COD 1 g O 2 1/8 electron equiv. ü NH 3 not oxidized (carbonaceous energy only) ü Aromatic hydrocarbons (benzene and toluene) and pyridines are not oxidized ü 6

Domestic Wastewater COD Fractionation Influent COD (Sti) 100% Biodegradable COD (Sbi) Unbiodegradable COD (Sui) ~80% ~20% Sol. readily biodegradable COD (Sbsi) ~20% Partic. slowly biodegradable COD (Sbpi) ~60% Soluble unbiodeg. COD (Susii) ~7% Particulate unbiodeg. COD (Supi) ~13% 7

Energy Measurement (4) l Biochemical (Biological) Oxygen Demand (BOD) ü O 2 required for microbial decomposition l Oxygen consumption by microorganisms DO consumed, mg/L BODu Nitrogenous energy BOD 5 Carbonaceous energy 5 Inadequate to assess the electron donor capacity; after 5 days, still some biodegradable matters exist. Time, days ~30 8

Energy Measurement (5) l Biochemical (Biological) Oxygen Demand (BOD) Carbonaceous BOD: aerobic heterotrophs v Decompose organic molecules to minerals (CO 2) and residues v Obtain their cell carbon from the organic material ü Nitrogenous BOD: obligate aerobic chemoautotrophs v Characteristics of nitrifiers (chemoautotrophs) DO < 2 mg/L action slow DO < 0. 5 mg/L action ceases Optimum p. H: 8. 0; p. H < 7. 2: slows down More sensitive than heterotrophs to toxins Slow growers (longer sludge age required) ü 9

Inert Organic Matter l Measured with COD l Not biodegraded, thus not measured with BOD 5 l Polymerized waste product l Inert material from lysed cells l Refractory organics: humic acid (M. W. – 5, 000~100, 000); fulvic acid (2, 000~10, 000) l Certain high M. W. carbohydrates alone or in combination with humic material are resistant to microbial attack. l High M. W. carbohydrates are excreted at the end of the logarithmic growth phase and help forming flocs by bridging of bacterial cells. 10

Acclimated Culture l Selection of populations by controlling environmental factors to encourage only the desired species. l An increase in the biodegradation rate of a chemical after exposure of the microbial community to the chemical for some period of time. 11

Example of Acclimation Lag phase Acclimation 25 Result of acclimation 20 Biomass conc. , mg/L CO 2 production, vol. Concentration, mg/L Chemical concentration 15 10 CO 2 production 5 Microbial biomass 0 0 5 10 15 20 Days 25 30 35 40 12

Example Acclimation Day 1 Day 2 Day 3 NBW RBW Feed ratio RBW NBW l Hazardous Industrial Wastewater Day 4 Biological Treatment Day 5 RBW: Readily biodegradable wastewater, e. g. , glucose, methanol, domestic wastewater, etc. NBW: Not readily biodegradable wastewater, e. g. , industrial wastewater, hazardous wastewater, polychlorinated biphenyls (PCBs), pentachlorophenol, etc. 13

COD, mg/L Influence of Acclimated Biomass on COD of Treated Wastewaters BOD 5 equivalents Total COD – BOD 5 Not readily Biodegradable COD Untreated Raw Non biodegradable COD Treated with unacclimated biomass Treated with acclimated biomass BOD: Not affected by acclimation COD: Significantly affected by acclimation 14

BOD, CODCr, CODMn, TOC Organic matter Biodegradable Unbiodegradable TOC BOD 5 CODCr Cl-, H 2 S CODMn Cl-, H 2 S Nitrification 15

Energy Measurement (6) l Total Organic Carbon (TOC) Oxidize in a combustion chamber with O 2 ü Easy to measure O 2 + 4 H+ + 4 e = 2 H 2 O ü Glucose, C 6 H 12 O 6 (M. W. = 180) C 6 H 12 O 6 + 6 O 2 6 CO 2 + 6 H 2 O 6 moles O 2 6 4 = 24 e 24/6 = 4 e available per unit organic C Ex. 100 mg/L of glucose: TOC and COD = ? TOC: (6 12)/180 100 = 40 mg/L C COD: (6 32)/180 100 = 107 mg/L O Glycerol, C 3 H 8 O 3 (M. W. = 92) C 3 H 8 O 3 + 7/2 O 2 3 CO 2 + 4 H 2 O 7/2 moles O 2 7/2 4 = 14 e 14/3 = 4. 67 e available per unit organic C Ex. 100 mg/L of glycerol: TOC and COD = ? TOC: (3 12)/92 100 = 39 mg/L C COD: (3. 5 32)/92 100 = 122 mg/L O • TOC values are very similar for both glucose and glycerol; however, COD values are quite different. • Thus, waste specific; cannot apply the result to other WWTPs. • Good as an operational tool with previous historical data. Similar Different 16

Energy Measurement (7) BOD 5/COD ratio: a good indicator for biodegradability of a specific wastewater l Domestic wastewater ü BOD 5/COD 0. 4 ~ 0. 8 ü BOD 5/TOC 1. 0 ~ 1. 6 l BOD 5/COD 0. 6: can be decomposed completely, biological treatment feasible l BOD 5/COD 0. 2: cannot be decomposed easily, chemical or physical treatment desired l BOD 5/COD 0: has toxic materials l 17

TOC Analyzer l. Measure the amount of total organic carbon present in a liquid sample; l. Convert inorganic carbon in the sample to CO 2 after adding acid and strip CO 2 by a sparge carrier gas; l. Oxidize organic carbon by either combustion, UV persulfate oxidation, ozone promoted, or UV fluorescence; and l. Measure CO 2 stripped using the conductivity or non dispersive infrared (NDIR) detection system. On-Line TOC Analyzer: a reagentless analyzer designed for continuous monitoring of organics. 18

Use of BOD 5, COD, and TOC BOD 5 l Good for regulating organic loading to a receiving water body for DO depletion by heterotrophs l Not good for design since some organics biodegrade slowly or after acclimation COD l Not good for regulation since it does not reflect true organic loading impact to aqua systems l Good for design if the input and output within a biological system is monitored; true energy count for carbonaceous energy only TOC l Good for operating a wastewater treatment plant due to real time monitoring capability l Values cannot be transferred to other wastewater due to specificity of carbon in the wastewater in terms of electro donor capability 19

Priority Pollutants l Designated by EPA in 1979 l A list of 126 specific pollutants that includes 14 heavy metals and 112 specific organic chemicals l Heavy Metals (Total and Dissolved): heavy, dense, metallic elements that occur only at trace levels in water, but are very toxic and tend to accumulate l Pesticides: DDT, Aldrin, Chlordane, Endosulfan, Endrin, Heptachlor, and Diazinon l Polycyclic Aromatic Hydrocarbons (PAHs): naphthalene, anthracene, pyrene, and benzo(a)pyrene l Polychlorinated biphenyls (PCBs): organic chemicals that formerly had widespread use in electrical transformers and hydraulic equipment 20

Fats, Oils, and Grease (FOG) l Sewage backups and overflows can lead to costly clean ups and repairs, as well as public health concerns. l Many utilities acknowledge fat, oil, and grease (FOG) as the main cause of sewer clogging. l EPA estimated that utilities spend on average $33, 000 per mile of sewer per year on capital project and $8, 000 per mile for O&M (2004). l The capital investment in wastewater infrastructure is over $13 billion annually (EPA, 2002). l Local government and utilities pay up to 90% of capital expenditures on wastewater infrastructure (AMSA and WEF, 1999). 21

Causes of Sanitary Sewer Overflow Mechanical or power failures (11%) Line breaks Misc. (5%) (10%) Blockages (48%) Wet weather I/I (26%) EPA, 2004 22

Causes of Sewer Clogging Roots and FOG (4%) FOG (47%) Roots (22%) Grit, rock, and other debris (27%) EPA, 2004 23

Sewer Clogging FOG Wastewater 24

Consequences of Sewer Clogging Sewer overflow 25

Consequences of Sewer Clogging Odor H 2 S + 2 O 2 H 2 SO 4 Bacteria Crown corrosion of concrete pipes 26

Consequences of Sewer Clogging Odor 27

Trans Fatty Acids (TFA) l Created in an industrial process that adds hydrogen to liquid vegetable oils to make them more solid l Easy to use, inexpensive to produce, and last a long time l Give foods a desirable taste and texture l Use trans fats to deep fry foods because oils with trans fats can be used many times in commercial fryers l Raise bad LDL (low density lipoproteins ) cholesterol levels and lower your good HDL (high density lipoproteins) cholesterol levels 28

Use of Zero Trans Fatty Acids l Inefficient removal in conventional grease removal systems l Potential foaming in wastewater treatment plant aeration basins l No knowledge on the fate of zero trans fatty acids in sewers and wastewater treatment plants 29

Prevention of Sewer Clogging (1) l Grease trap or interceptors, exhaust hood filters, and floor mats l Proprietary grease removal devices 30

Prevention of Sewer Clogging (2) l Chemicals and additives (emulsifiers, detergents or caustic substances) that claim to dissolve grease ü Prohibited for use as an additive because these substances reduce the efficiency of the interceptor or trap l Best Management Practices (BMP) during daily operations to keep FOG out of drains leading to the sewer l Enzymes ü Prohibited as additives due to the same effect as emulsifiers l Microorganisms ü Not prohibited as an additive l Education 31

Nitrogen l Main species üOrganic nitrogen üNH 4+: Ionized ammonia, nutrient to algae üNH 3: Free (unionized) ammonia, toxic to fish üNO 2 : Intermediate byproduct of nitirification, < 1 mg/L, causes the hemoglobin in the blood to change to methemoglobin, cause methemoglobinemia (‘blue baby’ syndrome) üNO 3 : Final product of nitrification, undeveloped digestive tracts of an infant possess bacteria that convert nitrate into nitrite, < 10 mg/L 32

Nitrogen Transformation in Biological Treatment Processes Organic nitrogen (proteins, urea, etc. ) Nitrification Bacterial decomposition and hydrolysis Ammonia nitrogen Assimilation Organic nitrogen (NH 3 N) (bacterial cells) Lysis and autooxidation O Organic nitrogen (net growth) 2 Nitrite (NO 2 ) O 2 Nitrate (NO 3 ) Denitrification Organic carbon (substrate) Nitrogen gas (N 2) 33

Subdivision of Total Influent TKN (Nti) 100% NH 3 & NH 4 (Nai) Organically bound N (Nti Nai) ~75% ~25% Unbiodegrad. soluble N (Nui) ~3% Unbiodegrad. Particulate N (Npi) ~10% Biodegrad. N (Nai) ~12% Total Kjeldahl Nitrogen (TKN): sum of organic nitrogen, ammonia (NH 3), and ammonium (NH 4+) in biological wastewater treatment 34

Free Ammonia (Unionized, NH 3) and Ionized (NH 4+) Ammonia More toxic to fish p. H eff ect Temperature effect Ammonia not regulated in winter More toxic to fish 35

Nitrification: Chemoautotrophs (1) l Nitrification: Conversion from ammonia to NO 2 / NO 3 Nitrosomonas + NH 4 + 1. 5 O 2 NO 2 + H 2 O + 2 H+ + New biomass Nitrobactor NO 2 + 0. 5 O 2 NO 3 + New biomass l Oxygen demand l Total oxygen demand for nitrification: 4. 57 g O/g N 36

Nitrification: Chemoautotrophs (2) l CO 2 (carbonate): carbon source l Ammonia: energy transfer source in a non assimilative way so only a small amount of biomass (sludge) is produced Nitrosomonas + NH 4 + 1. 5 O 2 NO 2 + H 2 O + 2 H+ + New biomass H+ +CO 32 → HCO 3 ; H+ +HCO 3 → H 2 CO 3 l Alkalinity ü 2 H+ 1 mol alkalinity [Ca. CO 3 (40+12+16× 3=100 g/mol)] ü 100 g Alk/14 g N = 7. 14 g Alk consumed/g N nitrified 37

Nitrification: Chemoautotrophs (3) Example l Influent TKN = 42 mg N/L; Effluent TKN 2 mg/L l Alkalinity = 200 mg/L as Ca. CO 3 Oxygen demand? l 4. 57 g O/g N × (42 – 2) mg N/L = 182. 8 mg O/L Alkalinity after nitrification? l 7. 14 g Alk/g N × (42 – 2) mg N/L = 285. 6 mg/L as Ca. CO 3 l Unless additional alkalinity (Ca. O, Na 2 CO 3, Na. OH, etc. ) is added, nitrification will stop (see the next slide). l Since the influent is 200 mg/L, 85. 6 mg/L + 10~15 mg/L (residual) = 95. 6~100. 6 mg/L as Ca. CO 3 required 38

Effect of p. H on Nitrification Operational range Nitrifiers: very sensitive to p. H Thus, buffer capacity (alkalinity) of wastewater important 39

Denitrification: Heterotrophs (1) 2 NO 3 + 10 e + 12 H+ → N 2 + 6 H 2 O O 2 + 4 e + 4 H+ → 2 H 2 O 40

Denitrification : Heterotrophs (2) l ~1 mol of H+ is recovered from denitrification l Thus, Alk of 3. 57 g/g N recovered l For low alkalinity water, denitrification is recommended. l Denitrification conditions üNo O 2 üReadily biodegradable soluble substrate (COD) l For complete removal of nitrogen species from wastewater: nitrification followed by denitrification 41

Substrate Requirement for Denitrification 0. 67 0. 33 CODutilized = CODbiomass + O 2 utilized = fcv X + O 2 utilized = YCOD CODutilized + O 2 utilized fcv = COD/VSS = CODbiomass/ X (mg COD/mg VSS) YCOD = CODbiomass/ CODutilized (mg COD/mg COD) O 2 utilized = (1 YCOD) CODutilized Biomass empirical YCOD = fcv Yh (mg VSS/mg COD) formula C 5 H 7 O 2 N + 5 O 2 5 CO 2 + 2 H 2 O + NH 3 (5 × 16 × 2 g) (1 × 113 g) = 1. 42 mg COD/mg VSS COD/VSS = 1. 42 mg COD/mg VSS O 2 = (1 fcv. Yh) CODutilized 0. 67 Nitrate consumption per mg COD utilized 2. 86 mg O 2/mg NO 3 N (1 1. 42· 0. 47) mg O 2/mg COD = 8. 6 mg COD req. /mg NO 3 N denitrified 42

COD Requirement for Denitrification Example l CODinf = 400 mg/L, CODeff = 50 mg/L, TKNinf = 55 mg N/L, TKNeff = 5 mg N/L, Q = 10 MGD l COD (methanol) required for denitrification? Solution l (55 5) mg N/L × 8. 6 mg COD/mg N = 430 mg COD/L req. l [430 (400 50)] mg/L = 80 mg COD/L req. l Methanol (CH 3 OH) (MW = 32 g/mol) l CH 3 OH + 1. 5 O 2 → CO 2 + 2 H 2 O l 80 mg COD/L× 10 MGD 1. 5× 16× 2 g COD/32 g Me. OH = 202 kg/day = 445 lb of Me. OH/day 43

Phosphorus Source: human body waste, food waste, various household detergents Subdivision of Total Influent P Influent TP (Pti) 100% Sol. PO 4 (P 70 ~ 90% si) Organically bound P (Pti Pbi) 10 ~ 20% in the activated sludge process 10 ~ 30% 44

Forms of Phosphate mg/L Old Now Forms 5 4 Orthophosphate 3 0 Tripolyphosphate (detergents) 1 0 Pyrophosphate (breakdown of tri P) 1 1 Organic phosphates 0 ? Hexametaphosphate (corrosion inhibitor) 10 5 Total Why? Ban of phosphate based detergents 45

Phosphorus in Wastewater l Addition to water üCorrosion (and scale) control in drinking water üIndustrial water softening üBoiler waters üCleaning compounds l Sewage ü 1. 2 lb/capita/yr from human and food waste 46

Mechanisms of Polyphosphate. Accumulating Organisms (PAOs) Short chain fatty acids (SCFAs) Organic substrate New Cell (Acetate) NADH, ATP PHA Glycogen ATP Facultative microbes ATP PHA Poly-P PAOs PO 4 Anaerobic condition PHA: Polyhydroxyalkanoates Glycogen 3 - PAOs PO 4 3 - Aerobic condition 47

Observations in Biological Phosphorus Removal (BPR) Systems AN O Ortho-P Bulk Liquid Acetate PHA Glycogen Biomass Poly-P PHA: Polyhydroxyalkanoates Reaction Time 48

BPR Mechanism Aerobic mg/L Anaerobic Ortho-P Acetate Poly β hydroxybutyrate (PHB) (Storage) Biomass Poly P Time Biomass PHB Poly P 49

Anaerobic/Oxic Process Readily biodegradable soluble COD Vital for P Uptake Excess sludge Better SRT control SRT: Solid retention time, sludge age, or mean cell residence time (MCRT); total biomass in the system/biomass wasted/loss 50

Whole Effluent Toxicity Bioassay 51

Bioassay l Part of whole effluent toxicity (WET) tests for NPDES permit l Use of a biological organism to test for chemical toxicity Mysidopsis bahia, female, approx. 6 mm in length Ceriodaphnia dubia 52

Use of Toxicity Testing in Water Quality Based Toxics Control l To characterize and measure the aggregate toxicity of an effluent or ambient waters l To measure compliance with whole effluent toxicity limits l As an investigative tool and to measure progress in a toxicity reduction program l As an ambient instream measure of toxicity to identify pollution sources 53

Bioassay l Tested sample: most commonly, effluent from industrial or municipal wastewater discharges l Sample holding time: max. 36 hrs stored at 4°C l Test organisms üCeriodaphnia dubia (water flea) üPimephales promelas (fathead minnow) üCyprinella leedsi (bannerfin shiner) üMysidopsis bahia (mysid shrimp) üMenidia beryllina (tidewater silverside) l Acute test: 24, 48, or 96 hrs (species specific) l Chronic test (short term): 4~10 (7) days 54

Rules for Conducting Toxicity Tests l 40 CFR 136. 3 Table 1 A (List of Approved Biological Methods) l Effective November 15, 1995 l Amended November 19, 2002 and effective December 19, 2002 l Methods must be followed as they are written

Incorporate by Reference l Methods for Measuring the Acute Toxicity of Effluents to Freshwater and Marine Organisms. 5 th Edition, USEPA, Office of Water, October 2002, EPA 821 R 02 012 l Short-term Methods for Estimating the Chronic Toxicity of Effluents and Receiving Waters to Freshwater Organisms. 4 th Edition, USEPA, Office of Water, October 2002, EPA 821 R 02 013 l Short-term Methods for Estimating the Chronic Toxicity of Effluents and Receiving Waters to Marine and Estuarine Organisms. 3 rd Edition. USEPA, Office of Water, October 2002, EPA 821 R 02 014 56

USEPA Methods Documents l Health and safety l Quality assurance l Facilities, equipment and supplies l Test organisms and culture methods l Dilution water l Effluent sampling and handling l Endpoints and data analysis l Individual test methods l Report preparation and test review 57

Test Types l Acute and Short term Chronic Tests üStatic non renewal üStatic renewal üFlow through l Test Species dependent l Use dependent 58

Test Design l 5 Concentrations + Control üSerial dilution’s of effluent and “control water” (also termed “dilution water”) üDilution series of 0. 5 or greater üSingle concentration test l Replicates l Randomization (organisms/chambers)

Acute Toxicity Tests l Test Procedures ü 96 hours or less (species specific) üMortality is the measured endpoint üFor daphnia mortality determined by immobilization l Advantages üLess expensive and time consuming than chronic üEndpoint is easy to quantify l Disadvantages üIndicates only lethal concentrations üOnly the effects of fast acting chemicals are exhibited 60

Acute Test Acceptability Criteria l Minimum control survival at least 90% l Temperature maintained at 20± 1 o C l Maximum test organism age at start: ü 14 days for fish ü 5 days for Mysid shrimp ü 24 hours for daphnids (Ceriodaphnia dubia and Daphnia magna) 61

Short-term Chronic Toxicity Tests l Test Procedures üTypically 4 10 days üMortality, growth, fecundity, reproduction l Advantages üMore sensitive than acute, assess parameters other than lethality üMay better reflect real world l Limitations üMore costly and time intensive than acute üMore sensitive to low level contamination 62

Chronic Test Acceptability Criteria l Minimum control survival 80% l Minimum control dry weight (average): ü 0. 25 mg for fish ü 0. 20 mg for Mysid shrimp l Minimum of 15 young (average) for control C. dubia l Temperature maintained @ 25 +/ 1 o C l Maximum test organism age at start: ü 48 hours for fish ü 7 days for Mysid shrimp ü 24 hours for daphnids 63

Selection of Dilution Water l May be either a standard laboratory water or the receiving water l Choice of water is dependent on the objectives of the test üAbsolute toxicity use standard water üEstimate of toxicity in uncontaminated receiving water, use receiving water üContaminated receiving water, use laboratory water 64

Acute Test Endpoints l LC 50 Concentration of effluent that is lethal to 50 percent of the exposed organisms at a specific time of observation (e. g. 96 hr LC 50), (expressed as % effluent) l NOAEC No Observed Adverse Effect Concentration üLowest concentration at which survival is not significantly different from the control üalways set equal to 100% effluent l EC - Effect Concentration 65

Test Data l Typical dose response where mortality increases as the concentration of effluent in the mixture increases. l LC 50 would be somewhere between 25% effluent and 50% effluent. Control 0% Mortality 6. 25 % Effluent 12. 5 % Effluent 0% mortality 20 % Mortality 25. 0% Effluent 50. 0% Effluent 100. 0% Effluent 40% Mortality 80% Mortality 100% Mortality 66

Chronic Test Endpoints l IC 25 - Inhibition Concentration of effluent which has an inhibitory effect on 25% of the test organisms for the monitored effect, as compared to the control (expressed as % effluent). l NOEC - No Observable Effect Concentration Highest concentration of effluent tested which shows no statistically significant effect on the organisms as compared to the control (expressed as % effluent). 67

Toxicity Values l NPDES permits in the past used a “no observable effect concentration” (NOEC) to measure chronic toxicity and a 96 hour lethal concentration 50 (LC 50) to measure acute toxicity. l Permits are now being issued with an inhibition concentration 25% (IC 25). Limit: more stringent LC 50, IC 25, NOAEC LC 50, IC 25 Toxicity 68

Toxic Units (TU’s) l Reciprocal of the fractional LC 50, NOEC, IC 25 value l Calculated by dividing the value into 100 üTUa = 100/LC 50 üTUc = 100/IC 25 69

Methodology for Setting Limits (1) l IC 25 is a calculation based on the design flow of the POTW and the seven day low flow over 10 years in the receiving stream (7 Q 10) as follows: IC 25 = design flow/(7 Q 10 + design flow) × 100 Example: l The low for the receiving stream (7 Q 10) is 23 MGD. The design flow for the POTW is 4 MGD. IC 25 = 4/(23+4) × 100 IC 25 = 14. 8% l The POTW demonstrates toxicity if the test value is less than or equal to the calculated value of 14. 8%. This constitutes a violation of the NPDES permit. 70

Methodology for Setting Limits (2) l A serial dilution that the laboratory: 59. 2, 29. 6, 14. 8, 7. 4, 3. 7 and a control with 0% effluent. l Toxicity is demonstrated if there is a statistical significant difference in any dilution from the control set. l The difference can be in any of the three parameters: survival, reproduction, or growth. l In the example, the effluent fails if toxicity appears in the 14. 8% or 7. 4% or 3. 7% dilutions. 71

Redox Reaction Organic molecule (C+H+O+N+S+P) (wastewater, hazardous chemicals, etc. ) Electron donor Oxidized CO 2, H+, and e Reduced Electron A molecule acceptor 72

Aerobic Condition O l Aerobic respiration l O 2 present l Electron acceptor: O 2 (→ H 2 O) l Good for large volumes of dilute wastewater (< 500 mg BOD 5/L) l High growth rates, thus high sludge production (0. 3~1 lb VSS/lb BOD 5) l Produce a more stable end product 73

Anoxic Condition AX l Anaerobic respiration (denitrification) l No dissolved oxygen l NO 3 and NO 2 present l Electron acceptor: NO 2 and NO 3 (→ N 2 + H 2 O) l Relatively high sludge production l Should be avoided in the clarifier 74

Anaerobic Condition l l l AN Fermentation No O 2, NO 3 , NO 2 , or SO 42 present Electron acceptor: endogenously generated by the microorganism Good for concentrated wastes (> 1000 mg BOD 5/L) Low sludge production Complex organic compounds Low molecular weight fatty acids CH 4, CO 2, and H 2 O 75

Microbial Classification Energy Carbon CO 2 Chemical Chemoautotroph H 2 bacteria Sulfur bacteria Nitrifiers Iron bacteria Electron donor H 2, S, H 2 S, Fe 2+, NH 3, NO 3 Photoautotroph Light Green plants Algae Purple bacteria Green bacteria Electron donor H 2 O, S, H 2 S Organics Chemoheterotroph Animals Most bacteria Fungi Protozoa Organics (reduced) Photoheterotroph Few algae Cyanobactor Some purple & green bacteria Organics (reduced) 76

Oxidation-Reduction Potential (ORP) ORP m. V +300 +200 Process 1 2 3 +100 Electron acceptors Conditions O 2 Oxic or aerobic NO 3 Anoxic 4 0 100 200 300 400 5 SO 42 6 7 8 Carbonaceous organics 1. Organic carbon oxidation 4. Denitrification 2. Polyphosphate release 5. Polyphosphate uptake 3. Nitrification 6. Sulfide formation Fermentative anaerobic 7. Acid formation 8. Methane formation 77

Three Origins of Life Phylogenetic Tree Bacteria Origin of life Archaea Eukarya The phylogenetic tree shows that the Eukarya are more closely related to the Archaea than they are to the Bacteria. PROCARYOTA 78

Archaea l A group of single celled microorganisms l Requires neither sunlight for photosynthesis as do plants, nor oxygen. l Absorbs CO 2, N 2, or H 2 S and gives off methane gas as a waste product. 79

Bacteria l Single celled microorganisms which can exist either as independent (free living) organisms or as parasites, typically 0. 5– 5. 0 µm length Classification l Shape l Ability to form spores l Method of energy production E. Coli (glycolysis for anaerobes, cellular respiration for aerobes) l Nutritional requirements l Reaction to the Gram stain 80