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Prepared for Dept. Of Civil Engineering by Susan C.M. Harper September 2007
Table of Contents
Course Description . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 Schedule . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 Station/Drawer Supply List (if supplies are missing, please report) . . . . . 2 First Session: Laboratory Safety and Orientation . . . . . . . . . . . . . . . . . . . . 3
L ABORATORY R EPORT W RITE U P G UIDELINES . . . . . . . . . . . 5
Laboratory Exercises . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 Laboratory 1: Solids Determination . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 Laboratory 2: COD, DO and BOD 5 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10 Laboratory 3: Bacteriological Examination of Sewage . . . . . . . . . . . . . . . 17 Laboratory 4: Chloride Determination . . . . . . . . . . . . . . . . . . . . . . . . . . . 27 Laboratory 5: Chlorine and Chlorine Demand . . . . . . . . . . . . . . . . . . . . . 34 Laboratory 6: Alkalinity, Acidity, pH, Hardness, Colour, Turbidity . . . . 39 Laboratory 7a: Coagulation and 7b: Coagulation & Softening . . . . . . . . . 47
Appendices see: CIVL407Manual_CourseNotes.pdf . . . . . . . . . . . . . . . . . 54
(Environmental Laboratory Analysis - 1-3-0, 0-0-0) Testing procedures used in water quality studies and in the operation of water and wastewater treatment plants. Prerequisite Chem 151. Sc h e d u le Lecture CEME1204: Laboratory CEME1301: Wednesday 1:00 pm 1) Tuesday (time to be arranged by consensus) 2) Wednesday 2:00 to 5:00 pm 3) Thursday (time to be arranged by consensus) 4) Friday (time to be arranged by consensus) (Note: Lab Session must have at least 4 students to run)
Subject No Lab No Lab Laboratory Safety & Orientation (~1 hr) #1 Solids determination #2 Dissolved oxygen, COD, BOD No lab - Thanksgiving week - catch up #3 Bacteriological Examination of waste water #4 Chloride #5 Chlorine demand #6 Acidity, alkalinity, hardness, colour, turbidity No lab - Remembrance day - catch up #7a Coagulation #7b Coagulation and Softening
Date 2007 Sep. 4 - 7 Sep. 11 - 14 Sep. 19 Sep. 25 - 28 Oct 2 - 5 Oct 9 - 12 Oct. 16 - 19 Oct. 23 - 26 Oct. 30 - Nov.2 Nov. 6 - 9 Nov. 13 - 16 Nov. 20- 23 Nov. 27 - 30
Lecturer: Lab instructor: T.A.s: Lab: Marker:
Victor Lo, email: Susan Harper, email: Margaret Parsotan, email: Kazi Parvez Fattah, email:
email@example.com firstname.lastname@example.org email@example.com firstname.lastname@example.org
(604) 822-4880 (604) 822-4397
September 2007 CIVL 407 Lab M anual
Sta tio n /D ra w e r Su p p ly Lis t (if s u p p lie s a re m is s in g , p le a s e re p o rt)
The following items will be maintained in each stations supply drawer or at station on bench when required: 2 pair safety glasses 2 permanent marker pens 2 stir bars and a bar retriever 1 stir plate 2 buret funnels 4 30 ml beakers 4 50 ml beakers 2 100 ml beakers 2 150 ml beakers 2 250 ml beakers 2 1 L plastic beakers 2 600 ml plastic beakers 2 400 ml plastic beakers 2 10 ml grad cylinders 2 25 ml grad cylinders 2 50 ml grad cylinders 2 100 ml grad cylinders 1 buret stand 1 double buret clamp 2 50 ml burets 2 25 ml burets 2 10 ml burets 0-14 pH strips Thermometer 2 Stir stick or spatula Filter funnel (Nalgene magnetic) Tweezers (Millipore or Gelman for membrane filters)) On benches each row: Latex gloves- small, medium and large Non-latex gloves (keep on reserve for latex sensitive persons only) Filter racks with funnels #4 Whatman filter paper or equivalent Vacuum flasks and hoses 8 1 L grad cylinders 8 3 L plastic beakers
September 2007 CIVL 407 Lab M anual
Safety Shower..0. Spills: Wipe up all spills immediately. Safety Equipment: Eye Wash Station. Please take your labcoat away in a plastic bag. You must be informed of the hazards of a substance before using it. please get attendant help. Please ask for assistance if chemicals are spilled. A small amount of liquid always remains in the tip and must not be blown out.clean and dry. Sanitation: antibiotic soap for hand washing. WHMIS: Workplace Hazardous Materials Information System is a government regulation. with the tip against the side of the receiving vessel. No sandals or open-toe or heel shoes. 1. etc. DO NOT PUT BROKEN GLASS INTO THE REGULAR GARBAGE CONTAINER. Remember to empty and rinse burets as well. calculators.up to 10. Some pipettes may be “to 3 September 2007 CIVL 407 Lab M anual . MSDS: is a supplier information sheet that must be available for each product and describe the hazards and necessary handling requisites. no matter how minor. Putting pens in your mouth that may have been on the bench or handled with gloves could be hazardous. with contaminated gloves will contaminate those items.0 ml. They are calibrated by weighing the volume of distilled water that will flow from them by gravity. 15. if there is room. Clean-up: You must leave your work station as you found it . Pipettes must be placed with tips pointing up into the pipet basket supplied. Do not put lids and small items onto racks that may slip through to the filthy drip tray. Personal Protective Equipment: Lab coat (long). but if you do please ask for assistance. Be aware that handling your pens. If you cut yourself. Most pipettes are calibrated “to deliver” or TD and should never be blown out. wash down and dry the bench. Wearing lab coats out of the lab is forbidden.5. packs. 25.2 First Session: Laboratory Safety and Orientation 1) 2) 3) 4) NO Food or drink.. eye protection. disinfectant for bench surfaces. gloves as required. 50 and 100 ml.0. Hygiene: At times you will be working with sewage. All glassware must be rinsed well (at least 5 times with tap water) and put on paper towels at your station or on drying racks. 20. These should be used whenever standard solutions are measured or when upmost accuracy is required. Pipettes/ graduated cylinders/ beakers/ flasks: 5) 6) 7) 8) 9) 10) Volumetric pipettes are the most accurate way to measure volume. 2. Broken Glass: Be careful not to break glass. They are available in 0.
Ie If you want to measure 20 mls. Graduated or “measuring” pipettes are not as accurate as volumetric (bulb type) pipettes and the one having the maximum capacity closest to the volume required is the most accurate one to select. Chemical residues can be splashed in you face.deliver with Blow-Out”.. 50 mls. 13) Stir bars: Please remove from containers before pouring contents down the drain. Chemicals will be put out as required for each lab and should be used sparingly. Cylinders are available: 5 mls. Use the size of cylinder CLOSEST TO THE VOLUME YOU WISH to measure for best accuracy. 4 September 2007 CIVL 407 Lab M anual . 250 mls. YOU WILL USUALLY NEED THE INFORMATION GIVEN TO COMPLETE YOUR LAB WRITE-UPS. They can be unpredictable in force and are close to eye level. 11) Gas/air/vacuum taps: Do not touch these unless instructed to do so. They are useful for measuring samples 20 mls . 10 mls. do not use a 1 L graduated cylinder. Even the force of air from the air taps can be dangerous because it can contain grit or water. Undetected gas leaks can be deadly. If liquid should enter a bulb or pump.pay attention to the markings. chemicals or for titrations. Used to hold/transfer approximate volumes of samples. 500 mls. Erle n m e y e r or conical flask markings are not accurate. Pipette bulbs or pumps must be used and great care must be taken not to contaminate bulbs. They are usually identified by a double etched or coloured band at the top of the pipette. 2 & 4 L Beakers: are cylindrical in shape with straight sides and flat bottoms. 12) Cup sinks: Please do not use these sinks for the disposal or delivery of liquids. Flasks: Vo lu m e tric flasks are very accurate and should be used when diluting standards (together with volumetric pipettes).merely approximations. 25 mls. 100 mls. Additional supplies are available in the supply room (around the corner). Some are blow-out. A demonstration will be given. Pipettes are in drawers indicated. not wastefully as quantities are limited.1000 mls. Graduated cylinders: Not as accurate as pipettes. 1. NEVER mouth pipette. some are not meant to drain completely . LISTEN AND MAKE NOTE OF VERBAL AND CHALK BOARD INSTRUCTIONS. markings are not accurate at all . 14) Glassware/supplies: Each station has a drawer where some glassware and other useful tools will be kept. Make sure that you label beakers and flasks to avoid confusion and waste. Do not squish bulbs in the direction of another person. please rinse out immediately and set aside to dry. especially at smaller volumes. They are calibrated the same way as TD except that the remaining drop is blown into the receiving vessel. They are containers best suited for “swirling” liquid ie to mix reagents with samples in a colourimetric determination (Lab 4 or 6).
R EPORT E LEMENTS Title page Objective Materials and Method S HORT R EPORT L ABS 1 TO 6 Required Required Details are not required. State that the procedures were in accordance with CIVL 407 Lab Manual and note any deviations to the manual’s procedure. The mark allocation for some sections for both reporting format is indicated in parenthesis in the table below. The short report is intended to be a concise yet complete laboratory report whereas the long report is more elaborate.L ABORATORY R EPORT W RITE U P G UIDELINES Guidelines for laboratory report write up are also given in the CIVL407 Manual Course Notes and summarized here. Please note that the marking outline is intended only as a guideline to assist in report write-up and that marks would also be allocated for overall presentation and clarity of the report. Sufficient detail should be provided so that what was done could be understood and the experiment could be repeated from the procedure reported. Required  Required  Brief discussion required  Required Required  Required Observations and Results Sample Calculations Discussion Including sources of error Conclusions Questions and Answers Other Students’ Data and Results References Presented in an acceptable format. Total Mark L ONG R EPORT L AB 7 Required Required Required Include a brief description of the procedure. Required  Required  Required  Required  Required  Required  Required   5 September 2007 CIVL 407 Lab M anual .
and record the volume of settleable solids. Do not include any surface floating material as settled solids. oven muffle furnace. tin dishes. MATERIALS: -Imhoff cones. 2.. keep hands and pencils away from your mouth and wash hands frequently. settle for 15 minutes longer. graduated cylinders. desiccators. wipe up all spills and wash with disinfectant. distilled water bottles. If large pockets of liquid form between the particles of settled matter. sludge. Do not wear lab coats outside of lab and do not take away from lab unless stored in a plastic bag. Settle for 45 minutes. Use pipette bulbs. subtract an estimated volume from the measured volume of matter. final effluent. Mix the samples thoroughly and fill individually marked Imhoff cones to the 1 litre mark. SETTLEABLE SOLIDS 1. to measure sewage treatment plant efficiency in removing residue. Alternatively a place will be provided in the lab to store your coat. 6 September 2007 CIVL 407 Lab M anual . samples of raw sewage.3 Laboratory Exercises La b o ra to ry 1: So lid s D e te rm in a tio n WARNING You will be handling sewage samples which may contain pathogenic bacteria. A 1 L graduated cylinder may be used in place of the Imhoff cone for the sludge sample. A. gently dislodge any solids that have clung to the sides using a stirring rod. if necessary. evaporating dishes. OBJECT: -to become familiar with a number of the most common sewage treatment plant tests and to illustrate some of the difficulties in performing these tests. balance.
Once dishes are at room temperature they can be re-weighed. [Notes: to allow for rinsing and to avoid spillage when transferring dish to oven don't pour more than 40 mls].[Note: it may be harder to wash out solids if you allow them to settle before pouring into the dish . rinse the cylinder. Wet filter with a small volume of distilled water to seat it.TOTAL AND VOLATILE 1.not beakers. The furnace will then be allowed to cool significantly before dishes are removed to a desiccator. 6. Obtain the tare weights of three aluminum dishes each containing a glass fibre filter (already pre-washed. the dish will be transferred to a desiccator. Put the dish in the 103-105 °C oven for drying. when filtering slows significantly do not add more. 25-50 mls of raw sewage and 5-10 ml of sludge u s in g g ra d u a te d c y lin d e rs to measure . on a cart in preparation for staff to do so when all are ready. Obtain the gross weight of the dish [Note: it should be room temperature before weighing. 2. pre-fired and desiccated . Conversely. Using the sample poured out for Part B (for consistency). immediately after stirring the beaker. The dish containing the dried total solids can now be placed in a muffle furnace or. pour about 500 mls (for Part B & Part C) into a beaker and then. Pinch sides of dish in a bit to protect the filter from oven drafts. Quickly record the exact volume and pour the sample into the dish. . [Hint: pour out small portions (say 25 mls at a time for Effl. stir the beaker contents and then rapidly (so that it doesn't settle) measure a volume of sample into the appropriate graduated cylinder.] Rinse the graduated cylinder with small amounts of distilled water and add to filter. After drying overnight. Subtract the tare weight to obtain the total solids weight and calculate the mg/L value. 3. This is a different test to the one we are doing using Imhoff cones and should not be confused. The loss in weight represents the volatile solids lost from the originally measured sample volume. Assemble filtering apparatus. 4. Suggested portions: 100-200 ml of final effluent. tare weight. 10 mls for Raw & 5 mls for sludge) of well-mixed sample and filter entire portion before pouring another portion. SUSPENDED SOLIDS . position the filter and begin suction (vacuum tap).[Note: Sludge Volume Index is the volume occupied by 1 gram of sludge after 30 minutes of settling. adding the rinsings to the dish. Record the total volume filtered. dried. prior to session). Do not write on crucibles as high temperatures during the next steps will burn writing off.].done for you. Obtain the tare weight of a properly prepared porcelain evaporating dish (ie one that has been washed.] Using very small amounts of distilled water. SVI= settled sludge volume (ml/L) x 1000/suspended solids (mg/L)] B. 3.so be quick. The samples will be fired at 550° C for one hour. Place the aluminum dish into the 103°C oven to dry for at least one hour (or leave drying 7 September 2007 CIVL 407 Lab M anual 4. TOTAL SOLIDS and TOTAL VOLATILE AND FIXED SOLIDS AT 550o C 1. Mix the sample in the carboy thoroughly. dried and fired). sample source and sample volume. C. the solids remaining represent the fixed solids. Calculate the mg/L volatile and fixed solids. 2. Carefully remove filter from filtration apparatus and transfer it back to the aluminum dish. if instructed. 5. rapidly pour sample into a 50 ml graduated cylinder to approximately equal 35-40 mls (if using 50 ml dishes). Make sure that you have recorded all the pertinent details: Dish #.
3. What major types of solids are removed in primary treatment? in secondary treatment? 2. compute the sludge volume index (SVI). 7. For the time being. overnight). After firing the dishes will be moved to a desiccator. Please return tomorrow to obtain final weight. Lab personnel will perform the timed-firing after the class. What. Your hands may have moisture and natural and/or applied oil/lotion on them. dissolved.50. Calculate the percent solids reduction through the treatment plant. cool and weigh. Classify each of the following into its proper solids category(s) i. Discuss the meaning and significance of this index. Calculate as the mg/L total suspended solids. From the above determinations it will now be possible to obtain a value for the dissolved solids present in the sample. 6. 6. % reduction in settleable matter _________ % reduction in total solids _________ % reduction in volatile solids _________ % reduction in fixed solids _________ % reduction in suspended solids _________ % reduction volatile susp. DO NOT DISCARD! The gain in weight represents the total suspended solids of the sample.e. If all of the volatile solids reduced is given off as gas and if the specific gravity of the volatile solids is 1. 8. suspended. volatile. solids _________ % reduction fixed suspended solids _________ % reduction dissolved solids _________ Lab 1 Questions 1. What two techniques can be used to overcome or correct for the loss of material from filter pads when firing at 550 oC? 4. The aluminum dish and filter (from #4) must now be fired in a muffle furnace at 550°C for at least 30 minutes (or until a constant weight is achieved). fixed (use more than one adjective to describe each substance. Transfer dish to a desiccator. They must be allowed to cool completely before re-weighing them to determine the loss on ignition or volatile suspended solids. If sludge is used in Part A. place the dishes on a cart designated by the instructor. if any. Calculate the mg/L total volatile solids and total fixed suspended solids.5. where the volatile solids are reduced from 65% to 40%. Enter all data in the tables on following page. affect would there be to the TSS and TVSS results if your bare hands were used to handle a tin dish a) prior to obtaining the initial tare weight and b) after obtaining the initial tare weight? 5.) a) a bacterium b) sodium chloride c) sugar d) clay 8 September 2007 CIVL 407 Lab M anual . what is the percentage reduction in solids. A raw sewage sludge goes through an anaerobic digestion process.3 and fixed solids 2.
use embossed mark) Sample volume (ml) Gross weight . Suspended Solids Tin dish marking (do not use ink . Settleable Matter After 1 hour (mL/L) B.oven dry (G) Tin dish with filter tare weight (G) Total suspended solids (G) Total suspended solids (mg/L) Gross weight .DATA AND RESULTS Raw Sewage Aerobic Sludge A.fired (G) Loss on ignition (G) Volatile solids (mg/L) Fixed solids (mg/L) Percentage fixed residue C.oven dry (G) Dish tare weight (G) Total solids (G) Total solids (mg/L) Gross weight .use permanent mark on dish) Sample volume (mL) Gross weight .fired (G) Loss on ignition (G) Volatile suspended solids (mg/L) Fixed suspended solids (mg/L) Dissolved solids (mg/L) 9 September 2007 CIVL 407 Lab M anual Final Effluent . Total Solids Dish marking (do not use ink .
Closed Reflux. pencils. OBJECT: -to familiarize you with the commonest tests run for assessing treatment plant efficiency and pollution loading to receiving waters. vials with pre-measured reagents. spectrophotometer. otherwise reagents will run to the bulb end and make pipetting difficult and contamination likely.La b o ra to ry 2: CO D . NEVER pour water into acid. Hach block digester. The closed reflux method is more economical but because a small sample volume (2 mls) is used. COD . D O a n d B O D 5 WARNING You will be using concentrated acid and other corrosive and toxic chemicals. to illustrate the shortcomings of these tests. and to demonstrate the use of dissolved oxygen probes. Wipe up all spills immediately. Materials: Raw sewage and final effluent samples. Keep fingers. potassium hydrogen phthalate standards: 800. as well as handling samples probably containing pathogenic bacteria. Always use and store pipettes with the top higher than the tip. homogenization of samples containing suspended solids may be necessary in order to get a representative sample and to improve reproducibility. etc out of your mouth. 100 and 50 mg/L as COD. 10 September 2007 CIVL 407 Lab M anual . etc. 400. Digests from either method can be titrated or read colourimetrically. Wash hands frequently with soap. Colourimetric Method NOTE: Open reflux method is the most accurate means of determining COD because a large sample size is used (20 mls). wash/rinse bench tops with water/disinfectant. pipettes. 200. I.
400. thermometers. raw sewage and final effluent. 11 September 2007 CIVL 407 Lab M anual . into the appropriate tubes and screw cap on snugly.the azide modification of the iodometric method) and a dissolved oxygen probe and meter. Prepare calibration curve. well-mixed samples in the 25-position digester. Pipette 2 mls of sample (using volumetric pipettes unless particulate is present. 2. Note: You are going to measure DO on four different liquids using two different methods. Make sure sample is well mixed with the acid contents of the vial. The instructor will demonstrate the use of the spectrophotometer. 10-ml graduated pipettes. Remove the vials and allow them to cool to room temperature (use cold water to speed up if necessary). Raw Sewage (Infl) and/or Effluent (Effl) in the upper 1/4 of the tube using permanent markers. [Note: it should look homogenous and be careful it will be very hot!] 3. concentrated sulphuric acid. which is set at 600 nm to read absorbance of light by the sample. Allow samples to digest for 30 minutes (normally this would be 2 hours). 5. burette stand with burettes. BOD bottles. The vials should be clean and dry on the outside. use wide mouth graduated pipettes) in duplicate. 6. a volumetric correction must be made to the value obtained from the curve. absorbance versus concentration and calculate the COD in samples. DO NOT ADJUST THE SETTINGS ON THE METER. alkaline-iodide-azide reagent. If sample is known to be outside the standard range (above 800 mg/L). The four liquids are cold tap water. and are at the spectrophotometer. Read your samples and the set of COD standards: 800. manganous sulfate. dilution water. DISSOLVED OXYGEN (DO) Materials: Raw sewage and final effluent samples. 50 and 0 COD as mg O 2 /L. in which case. Standards have been digested for you. 500-ml Erlenmeyer flasks. At your station you will find four vials with pre-measured solutions containing potassium dichromate and sulfuric acid (prepared according to standard methods. You will use a chemical titration (Winkler . except without mercuric sulfate).noting the location of your well-labelled. 100. 250-ml graduated cylinder. Put identifying labels on each tube ie your initials. 4. II. 0.025 N sodium thiosulphate. 200. Take vials to fumehood and place in block digester . a smaller aliquot (making volume up to 2 mls with distilled water) or 2 mls of a pre-diluted sample can be used. If less than two mls of sample or diluted sample was used. aerated dilution water. before the lab. The probe has been previously calibrated.Procedure: 1. starch indicator.
then the same sample will require little or no titre and may not turn blue when starch is added . . 1. Determine the percent saturation for each sample. Take each stoppered bottle. WINKLER TITRATION (Azide modification) NOTE: Use only the droppers attached to the reagent bottles for dispensing into samples. dropwise. add 1 ml of alkaline-iodide-azide solution the same way. 4. right to the top and place stopper on bottle. 1. 8. aerated dilution water. until the first complete disappearance of the blue colour. 2.think about it!) Complete all four titrations. invert several times again and allow to settle again. add 1 ml of concentrated H 2 SO 4 and quickly restopper. 5. Add enough starch solution to give a strong blue colour (one dropper full) and continue to titrate. 9.A. 7. The DO concentration in mg/L is equal to the number of ml of titre as long as 201 ml is titrated. Tip out the liquid from the area around the stopper and invert several times to mix thoroughly. Determine DO. DISSOLVED OXYGEN PROBE NOTE: DO NOT add any reagents to the samples used for the probe method. Record the temperature of each sample using the DO meter or a thermometer. Fill a BOD bottle with each of the four samples (or use ones saved from A ). Fill four BOD bottles with the four samples (cold tap water. To the same bottles. Place the bottles in the sink and one at a time remove the stopper. submerge the hose in the bottle and allow the water to overflow for 15-30 seconds). again quickly replacing the lid. The chemical floc should completely dissolve. Neglect any reappearance of the blue colour. Use the specially marked volumetric flask to dispense the 201 ml. Instructions on the use of the probe will be given in the lab. B. for each of the four samples. (Note: DO meter temperature is usually not accurate. Set back in sink. tip the liquid out of the space around the stopper (caution -it may be corrosive) and invert several times. (Fill carefully. (Hint: if you measured very low DO level with the meter.) Transfer 201 ml of each bottle (do one at a time) to a 500 ml conical flask. to thoroughly mix contents. 2. Allow the floc to settle to about 1/3 the bottle volume. (Note: biological solids originally present in the sample will not dissolve. Titrate this volume with 0.) You can save these bottles for B. Stopper the bottles without trapping any air bubbles.025 N Na 2 S2 O 3 until only a faint yellow or "straw" colour remains. 6. Note: if solution already seems pale yellow add starch right away. 12 September 2007 CIVL 407 Lab M anual 3. taking care not to entrain additional air. raw sewage or influent and final effluent). as demonstrated. remove the stopper. For the cold tap water sample. Do not mix droppers and do not allow tap water into the reagent bottles. Record the volume of titre. add 1 (one) ml of manganous sulfate reagent using plastic dropper provided (keeping the dropper tip just below the surface) and quickly replace the lid. using the DO probe and meter. When the floc has settled the second time to about 1/3 bottle volume.
Pair up the mg/l of dissolved oxygen you measured and the temperature of the water in degrees C.Dissolved Oxygen Data Sample/Data Bottle # Sample Volume (mls) Initial Buret reading Final Buret reading Net Titre (mls) DO conc. use the saturation chart above. Draw a straight line between the water temperature and the mg/l of dissolved oxygen. Note that this nomogram assumes that the water is at sea level The saturation value can also vary slightly depending on barometric pressure with lower values expected when a storm front moves through as compared to bright and sunny "high pressure" days. Raw Sewage Final Effluent Tap Water Dil’n Water 13 September 2007 CIVL 407 Lab M anual . The percent saturation is the value where the line intercepts the saturation scale. (mg/L) DO conc. (Probe) Temp (oC) (thermometer) Saturation concentration Percent saturation DETERMINING PERCENT SATURATION THE "QUICK AND EASY" METHOD For a quick and easy determination of the percent saturation value for dissolved oxygen at a given temperature.
The blank is used to check that the dilution water has negligible BOD. Place all bottle sets in the 20 o C incubator for five days (or 6 for Tues. it may indicate that probe may not have been calibrated properly or the dilution water had been contaminated. thereby maintaining a water seal for each bottle. Measure the Initial DO using a calibrated probe and meter. Note: Each group should have 9 BOD bottles in the incubator. Because you want to be sure that the dilution water has no BOD. Try not to aerate the contents by keeping the filling tube below the surface of the liquid in the bottle or carefully running the water down the side of the bottle. BOD bottles. dilution water. If the dilution water BOD is greater than 1 ppm. remove bottles and measure the DO. 6. 3.III. Carefully top up the BOD bottles with DILUTION WATER. showing sample calculations. ie Rinse the filling tube if it has been in the sample. 1. These caps are designed to prevent the evaporation of the liquid around the stopper. There should be negligible BOD in the Dilution water. but if there is a significant BOD then it will have to be subtracted before the dilution factor is applied in the calculation. Place plastic caps over all bottles. All bottles should have liquid in the well around the stopper . BIOCHEMICAL OXYGEN DEMAND Note: Because of the logistics of organization some groups may have 6 day BOD 5 instead of the usual 5 day. 4. Measure the Initial DO of the blank using the calibrated probe and meter. graduated cylinders. be careful not to contaminate it with sample. Be sure to record the bottle numbers and sample volumes on the following table . 10-ml graduated pipettes. to bring the levels above the ground glass neck of the bottle.do not write on the bottles and do not use the numbers on the lids. Using the volumes and samples provided by the instructor. 14 September 2007 CIVL 407 Lab M anual . groups). Be sure to note this in your lab write-up.if not add a some dilution water (or distilled water) to the well. carefully measure the waste water samples into BOD bottles. 5. being sure to exclude any air bubbles that might be trapped under the stopper by adding dilution water. In addition. Stopper the BOD bottles. Two dilutions will be made for each sample and each dilution will be in duplicate for a total of eight bottles (plus one bottle for the blank mentioned in Step 4). Dissolved oxygen meter and probe. preferably using the same meter and probe used to get IDO reading. Calculate the 5-day (or 6-day if unavoidable) BOD. After five (or six)days. Materials: Raw sewage and final effluent samples. fill one BOD bottle with DILUTION WATER alone.
mg/L = DO of diluted sample after 5 d incubation at 20 oC. and ratio of seed water in diluted sample to seed in seed control= (%seed in diluted sample) / (%seed in seed control).BIOCHEMICAL OXYGEN DEMAND (BOD) DATA Sample source: ________________________________ Raw Sewage Bottle number (not lid #) Sample volume (ml) Initial DO (mg/L) Final DO (mg/L) Final Effluent Bottle number (not lid #) Sample volume (ml) Initial DO (mg/L) Final DO (mg/L) Dilution water Bottle number (not lid #) Sample volume (ml) Initial DO (mg/L) Final DO (mg/L) _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ _____ BOD Calculation When dilution water is not seeded (our water is not seeded for this exercise): When seeded water is used: where: D 1 = D2 P B1 B2 f mg/L. DO of diluted sample immediately after preparation. mg/L DO of seed control after incubation. = = = = decimal volumetric fraction of sample used DO of seed control before incubation. 15 September 2007 CIVL 407 Lab M anual . mg/L.
One waste has a K value of 0. open reflux. Both wastes have equal BOD 5 temperature and volume. titrimetric method. the other 0. If an industrial waste has a COD of 450 mg/L. 2 Final Effluent.17. 2 ________ ________ ________ ________ ________ ________ ________ ________ Average ________ Average ___________ ________ Percentage BOD reduction through treatment plant Lab 2 Questions 1) At what relative times should influent and effluent samples be taken from a sewage treatment plant in order to determine true % removal efficiency? Give two reasons why samples for dissolved oxygen should be “fixed” in the field. Diln. 2) 3) 4) Show on a graph the approximate dissolved oxygen sag curves in a river receiving two wastes. 1 Final Effluent. Diln. What interference does the Azide Modification of the Winkler Dissolved Oxygen method overcome and why is it the most common method used for domestic sewage? Why is a blank titrated in the COD test? (Refer to standard methods. 6) 7) 8) 16 September 2007 CIVL 407 Lab M anual .Summary Raw Sewage.) Why is your COD result different from the BOD result? 5. Diln. 1 Raw Sewage. Diln. Calculate theoretical oxygen demand for 300 mg/L of methyl alcohol. if at all possible. what would you expect its I) BOD 5 and ii) BOD L to be.25.
Keep hands.. LES Endo agar plates. EC tubes and/or Hach A1 tubes. Heterotrophic plate count: dilution tubes. lab coats.b e c a u s e o f lo g is tic s it is n o t p o s s ib le f o r y o u to p e rf o rm th e la b a s p re v io u s ly w ritte n . Brilliant Green Lactose Broth tubes. etc. pencils. agar tubes. F iv e d if f e re n t p ro c e d u re s w ill b e s tu d ie d . sterile pipettes.La b o ra to ry 3: B a c te rio lo g ic a l Exa m in a tio n o f Se w a g e WARNING As you are handling and pipetting samples probably containing pathogenic organisms please observe the following rules: Wear gloves. Wipe lab benches with disinfectant before and after each lab. wipe up all spills and disinfect the area of the spill. y o u w ill d o th o s e m a rke d b y ? only 17 September 2007 CIVL 407 Lab M anual . HACH P/A Broth with MUG. lauryl tryptose broth tubes. Wash hands with soap before leaving. 35 oC incubator. not both. sterile dilution water. Demonstration only SAMPLE: You will do either a raw sewage sample or a final effluent sample. OBJECT: -to demonstrate different methods for the determination of the coliform group of organisms and to acquaint you with bacteriological procedures. DO NOT MOUTH PIPETTE. 35oC incubator. membrane filters.leave it here or take away in a plastic bag to launder. filtering apparatus. Quebec colony counter. Get raw data from another group and include in your report. sterile pipets. Students will do only sections marked ? MATERIALS: Multiple tube method: dilution tubes. T h e re f o re . Do not wear lab coat out of lab . eye protection. Demonstration only. plates. Membrane filter method: dilution tubes. away from your mouth.
) Methods: ? Recording BGLB. MUG produces a fluorogenic product when hydrolyzed by an enzyme specific to E. Microscope Slides (Completed Phase of Multiple-Tube Fermentation): Single colonies arising from single cells are prepared and stained for microscope examination. (Single colonies are obtained by streaking LES Endo agar plates with an inoculum from positive BGLB or EC broth tubes and incubating for 24 hours. 18 September 2007 CIVL 407 Lab M anual . Formation of gas in any amount in the inverted vial of the BGLB broth fermentation tube at any time within 48 hr ±3 hr constitutes a positive confirmed phase. Bacterial numbers are calculated from the numbers of positive tubes.Pour Plate Method: Serial dilutions of samples mixed with agar and incubated for 48 hrs at 35oC. Another series of tubes are inoculated from these positive lauryl tryptose tubes. EC or A-1 tubes positives: 1.A. All tubes have been examined for gas production. using a ready-to-use P/A Broth with MUG. Note: these methods will be demonstrated but data must be recorded.coli. Streak plates in a manner to insure presence of some discrete colonies. Hach’s Most Probable Number Method 8368. 1. The broth will turn yellow indicating total coliform and fluoresce in the presence of E. Consider positive EC broths incubated at the elevated temperature (44. The number of colonies counted directly. It is used as a faster alternative to the LTB/EC procedure for enumerating faecal coliforms in water. Gas formation indicates the possible presence of coliform organisms. Using aseptic technique. A-1 Medium. streak one LES Endo agar plate from each tube of BGLB broth showing gas from the highest dilution and the second highest dilution. Gas-positive Hach A-1 tubes indicate the presence of faecal coliform. as demonstrated.COLI in 4 to 24 hours.coli. Completed Phase LES Endo agar Plates: A p la te w ill b e a v a ila b le to lo o k a t. E. Brilliant green lactose bile (BGLB) tubes are inoculated and incubated at 35oC for 48 hours (confirmed test for total coliforms) and EC tubes are inoculated and incubated at 44. Colonies that are pink to dark red with a golden green metallic sheen are counted after incubation 20-22 hrs at 35oC. Express as Total coliforms. ? B. Results will be provided for the demonstration set. Use Table 2 on page 27. Presence/Absence Testing: HACH technique to simultaneously screen for total coliform and E. Calculate the MPN value from the number of positives using the Table 1.5 oC) as a positive completed test response for both Total and faecal coliform. Membrane Filter Technique: Serial dilutions of sample are collected on membrane filters and placed on M-Endo media. D. Parallel positive BGLB broth cultures with negative EC broth cultures indicate the presence of nonfaecal coliforms.7oC for 24 hours (confirmed test for faecal coliforms). Multiple-tube Fermentation or Most Probable Number: Serial dilutions of sample are incubated in lauryl tryptose broth at 35oC for 48 hours (swirl after 24 hrs). is USEPA-accepted for testing non-potable waters. 2. Demonstration. Heterotrophic Plate Count . 3. ? C.
Colonies that lack sheen may be pink. or colourless and are considered to be non-coliforms. removing the cover just enough to insert pipette tip and dispense sample. therefore the results table should specify #coliform colonies/100 ml. Atypical coliform colonies can be dark red or nucleated without sheen. Dilution tubes. The filtration apparatus has been sterilized by autoclaving. 4. Verification is usually done by a test for lactose fermentation. Use dishes with 20 to 80 coliform colonies and not more than 200 of all types. place a sterile membrane filter (grid side up) onto the filtration apparatus. choose appropriate dilutions for counting. Counting. Place the filter on a small LES Endo agar plate carefully. Inoculating. Close valve and rinse the funnel with three aliquots (portions) of sterile dilution water opening and closing valve after each aliquot. Pipette about 10 ml of sterile dilution water over the surface of the filter with the suction valve still closed. Flame loop between second and third quadrants to improve colony isolation. After 20-22 hr. 2. 2. If you haven’t already.applying gentle pressure. The total count of colonies (coliform and noncoliform) on Endo-type medium has no consistent relationship to the total number of bacteria present in the original sample. with a motion that excludes air as it comes in contact with the agar. then immediately replace the cover. Prepare Gram Stain slides for examination under a microscope.Pour Plate Method 1. Be sure dishes are labelled with your name and dilutions. open the valve slowly and allow all the liquid to be pulled through. 3. red. from one edge. Repeat for the rest of the dilution tubes to # 2. 19 September 2007 CIVL 407 Lab M anual . ?C. Incubate inverted Petrie dishes for 20-22 hr at 35 oC. 5. Gram staining technique can be also be used.2. Verification of both types of colonies is advisable as some typical sheen colonies may be noncoliform and some atypical may be coliform. The sheen area may vary in size from a small pinhead to complete coverage of the colony surface. The typical coliform colony has a pink to dark-red colour with a golden green metallic surface sheen. Incubate plates (inverted) at 35 oC for 24 ± 2 hr. Using a sterile pipette. Membrane Filter Technique -Total Coliform 1.. Pour the full volume of dilution tube # 6 onto the filter. In general. prepare one set of six dilution tubes following the instructions given under Multiple Tube Fermentation. transfer 1 ml of sample from dilution tube # 6 (most dilute for Raw sewage) or #5 (for Effluent) to an empty (large) Petrie dish. Filtering. Heterotrophic Plate Count . -Note: “typical” colonies count as coliforms in this procedure. A high count of non-coliform colonies may interfere with the maximum development of coliforms. Dilution tubes. The sheen area may vary in size from a small pinhead to complete coverage of the colony surface. Mount apparatus on suction flask with valve sideways to block suction. coliform bacteria are Gram-negative (pink) rods (sometimes coccus-like). ?B.1. white. Prepare one set of six dilution tubes as described in Section A. When filtering is complete and the valve is closed. Using sterile forceps. Do not push down on the filter except on the outside edge and use only sterile forceps . remove filter using flamesterilized forceps (allow forceps to cool or they will damage the filter).
2. Do this for a typical and an atypical colony. Flood the slide with fresh alcohol-acetone solution to act for 30 seconds. if fewer than 10/cm 2. Take 5 tubes of melted agar from the incubator. select the appropriate dilution to count.d e m o n s tra tio n -s ta rte d a h e a d o f la b 1. Allow the agar to solidify in the dish before inverting. If more than 10/cm 2.too hot for a bacterium. Incubate at 35 oC for 24-48 hours. 2. E. place 1 loopful of sterile dilution water in the centre of a slide. marking (on edge of dish so as not to impede counting) and incubating for 48 hr at 35 oC. Flood slide with crystal violet solution and allow to act for 30 seconds. Repeat for the rest of the dilutions. Examine for yellow colour (total coliforms) and fluorescence (E. Drain off excess iodine and wash freely with water and then alcohol-acetone decolourising solution. on the bench. 20 September 2007 CIVL 407 Lab M anual . Air dry the slide high above the Bunsen burner and fix by passing slide briefly through a flame. Do not report as “too numerous to count” if all plates exceed 300 colonies. ?3. Test temperature on your wrist . If necessary. count four representative squares. After 48 hr incubation. Resterilise the loop and remove one colony from the surface of the Les Endo streaked plate (prepared previously). f. Spread the colony in the drop of water so that you get a uniform dispersion over an area about the size of a dime. 4. a. c. Wash off stain with water and then with iodine solution (Lugol’s). d. Work quickly or the agar will solidify in the tube before you get a chance to pour it. Presence/Absence Testing . Label the slide using a grease pencil (T or A). Slide preparation: With a flame sterilized inoculating loop. pour and replace cover. average and multiply by 57 (for plastic Petrie dish). Multiply the number by 5 to compute estimated colonies per plate (66 cm 2 ). use the plate(s) that will give from 30 to 300 colonies/plate. Slide staining. e. The aim of using several dilutions is to have at least one dilution giving colony counts between these limits. Allow iodine to act for 30 seconds. Use the Quebec colony counter to aid in counting and a grease pencil to mark off colonies counted. ?4. count colonies in 13 squares (of the colony counter) of the most dilute plate having representative colony distribution. b. Add 100 mls of sample directly to pre-measured medium provided. Instead. Gently swirl the dish. D. A hand counter is also available. Ideally. Record results of previously prepared test. Slide the cover of a Petrie dish back just far enough to pour the agar. Repeat with dilutions 5 to 2 (for Raw) and 4-1 (for Effluent). Compute bacterial density and report as “colony-forming units” (CFU) per ml.3. 5.coli). to mix the sample with the agar. refer to Standard Methods for more instruction. Let them cool down from the 60C that they are kept at but not enough to solidify. Preferably select seven consecutive squares horizontally and six consecutive squares vertically.too hot for a baby . Microscope Slides: 1. 6. Wash slide in tap water. Counting.
Wash in water. and on the slide: typical or atypical.Record positives in all Multiple Tube Fermentation inoculations: LTB. In general.Come in and count colonies on LES Endo Membrane Filter dishes. . coliform are Gram-negative (pink). a. concave). usually motile. smooth.counts will be provided on the board. colour.Membrane filter prepared dilutions.Record Presence/absence testing results of Hach Prepared Media bottles. length to width ratio. blot and then dry in air high above the Bunsen burner (so as not to burn away your efforts). after decolourisation Gram negative organisms are no longer visible. After application of the counterstain. . h.3 µm in size.g. Note: in the staining process: before decolourisation all organisms appear Gram positive (blue).Examine prepared slides. sometimes coccus-like and 0. ?3.Prepare and incubate Heterotrophic (Pour) Plates using dilutions above. BGLB and EC and AI tubes . odour. Plates ready on a weekend will be put in cold room until Monday. place on LES Endo agar dishes and incubate. short. Schedule Summary Day 1. or in short chains. . cell groupings (single. Cells occur singly. . plump rods. Microscopic examination. cocci or rods.Come in and count Heterotrophic Plate colonies after 48 hours. Examine the prepared slides . Day 2. Gram negative organisms are visualized (pink). Apply fuchsin or safranin (counter)stain for 30 seconds. . metallic. Description of bacteria: Describe what you see on the plates: colonial morphology of each type of colony seen ie surface (shiny.5 . transparency. 21 September 2007 CIVL 407 Lab M anual . b.do not turn knobs except the positioning ones. nonsporing. pairs chains). rough. . gram positive or negative.Prepare dilutions for Heterotrophic Plate Count and Membrane Filter technique. in pairs. c. . Day 3. dull. .
Multiply the MPN index by the dilution factor from the series used when dilutions other than 1/1.. 1/ 10 and 1/100 are used.pour plate method. 35 oC/48 hr Tube # 1 2 3 Mls of inoculum Infl Effl Plate count Infl Effl *CFU/ml Infl Effl *CFU= colony-forming units Membrane Filter Technique Tube # Dilution (mls) Infl Effl Coliform Colonies Infl Effl #Colonies/100 mls Infl Effl 1 2 3 4 5 6 4 5 6 22 September 2007 CIVL 407 Lab M anual .Bacteriological Examination Data Multiple Tube Fermentation : indicate the number of positives for each dilution Tube # Mls of sample/diln tube LTB 24 hr (+ or -) BGLB 48 hr (+ or -) EC 24 hr (+ or -) AI 24 hr (+ or -) Infl Effl Infl Effl Infl Effl Infl Effl Infl Effl From MPN Index table find MPN Index per 100 ml. 1 2 3 4 5 6 Heterotrophic Plate Count .
E. Do they agree? Why/why not? What is expected? What is a heterotroph? -were the reductions through the sewage treatment plant as expected? Additional notes Presence/Absence (P/A) Testing: Media chosen may be Presence/Absence broth (P/A) or Lauryl Tryptose broth with Bromcresol Purple broth (LT/BCP) to indicate acid formation. Do they agree? Why/why not? -compare MPN/MF with Heterotrophic Plate Count.01 ml 2/5 2/5 0/5 0.001 ml 0/5 0/5 0/5 Combination of positives 5-2-0 5-4-2 0-1-0 MPN Index /100 ml 5000 2200 20 23 September 2007 CIVL 407 Lab M anual .coli. A long-wave UV light is required to detect fluorescence. an enzyme specific to E. With MUG reagent added to P/A Broth.Discussion: -sources of error? -compare MPN. the total tubes planted.coli will fluoresce as early as 4-24 hours. choose the highest dilution that gives positive results in all five portions tested (no lower dilution giving any negative results) and the two next succeeding higher dilutions. If zero total coliform is the maximum contamination goal.coli because it produces a fluorogenic product when hydrolyzed by glucuronidase. In the examples given below. The number in the numerator represents positive tubes. the combination of positives simply represents the total number of positive tubes per dilution: Example a b c 1 ml 5/5 5/5 0/5 0.1 ml 5/5 4/5 1/5 0. that in the denominator. you can simultaneously detect total coliforms and E. They are in concentrated form and merely require the addition of 100 ml of sample (or diluted sample) to the media and incubation. Estimation of Bacterial Density . use the results from only three of these in computing the MPN. it is not necessary to enumerate. MF results.coli. Both media meet USEPA guidelines for testing of total coliforms in drinking water. the significant dilution results are in boldface. To select the three dilutions to be used in determining the MPN index. Use the results at these three volumes in computing the MPN index. MUG (4methylumbelliferyl-$-D-glucuronide) can be used to detect E.Most Probably Number (MPN) When more than three dilutions are used in a decimal series of dilutions.
0 mL.MPN Index and 95% Confidence Limits for Various Combinations of Positive Results when 5 Tubes are used per Dilution (10 mL. Of Positives MPN Index/ 100 ml 22 26 27 33 34 23 30 40 30 50 60 50 70 90 80 110 140 170 130 170 220 280 350 240 300 500 900 1600 95% Confidence Limits Upper Lower <2 2 2 4 2 4 4 6 6 4 7 9 9 12 8 11 11 14 14 17 13 17 17 21 26 1 10 13 11 15 15 18 18 17 20 24 25 29 24 29 29 35 35 40 38 45 46 55 63 4-2-0 4-2-1 4-3-0 4-3-1 4-4-0 5-0-0 5-0-1 5-0-2 5-1-0 5-1-1 5-1-2 5-2-0 5-2-1 5-2-2 5-3-0 5-3-1 5-3-2 5-3-3 5-4-0 5-4-1 5-4-2 5-4-3 5-4-4 5-5-0 5-5-1 5-5-2 5-5-3 5-5-4 5-5-5 9 12 12 15 16 9 10 20 10 20 30 20 30 40 30 40 60 80 50 70 100 120 160 100 100 200 300 600 -— 56 65 67 77 80 86 110 140 120 150 180 170 210 250 250 300 360 410 390 480 580 690 820 940 1300 2000 2900 5300 ---- $1600 24 September 2007 CIVL 407 Lab M anual .1 mL) Comb. 1. Of Positiv es 0-0-0 0-0-1 0-1-0 0-2-0 1-0-0 1-0-1 1-1-0 1-1-1 1-2-0 2-0-0 2-0-1 2-1-1 2-2-0 2-3-0 3-0-0 3-0-1 3-1-0 3-1-1 3-2-0 3-2-1 4-0-0 4-0-1 4-1-0 4-1-1 4-1-2 MPN Index/ 100 ml 95% Confidence Limits Lower 1 1 1 1 1 1 2 2 1 2 3 3 5 3 4 4 6 6 7 5 7 7 9 12 Upper Comb. 0.Table 1 .
MPN Table 2 25 September 2007 CIVL 407 Lab M anual .Hach .
-The Use of Indicator Organisms to Assess Public Water Safety .microbelibrary. What are the important pathogens transmitted by water? How is quality control achieved in microbiological analysis.Further information can be found in the laboratory library (do not remove without permission): Standard Methods: -Part 9000 Microbiological Examination HACH publications:-Catalogue listing testing methods with descriptions.com/ ¸Information Central/Learning Library/Microbiological Testing ASTM: (STP635) D. Mara: -Bacterial Indicators/ Health Hazards Associated with Water -Bacteriology for Sanitary Engineers Microbiology 321: -Manual of Microbiological Techniques Website: http://www.D.hach. 2) 3) 26 September 2007 CIVL 407 Lab M anual .htm Lab 5 Questions: 1) Compare the advantages and disadvantages of the multiple tube fermentation and membrane filter techniques.org/microbelibrary/files/ccImages/Articleimages/keen/Gram stainkeen.http://www.
5 mg/L). magnetic stirrer. goggles or glasses and lab coats. standard Hg(NO 3)2.La b o ra to ry 4: Ch lo rid e D e te rm in a tio n WARNING You will be using concentrated acid and other very dangerous chemicals. Hint: optimum ranges will coincide with the standards provided or will be indicated within the pertinent section. HNO 3. sample. reference electrode (double junction). If chemicals are spilled alert instructors. chloride specific ion electrode. Mercuric thiocyanate is very toxic. You will be told the approximate concentration in the sample and it is up to you to make sure that the sample will fit within the specified ranges. indicator-acidifier reagent. volumetric pipettes. beakers. standard chloride solution (0. immediately. 27 September 2007 CIVL 407 Lab M anual . to illustrate the differences in operation and accuracy between instrumental methods and wet chemical methods. semi-log graph paper. buffer. standards. known addition and calibration techniques and colourimetric and potentiometric titrations. graduated cylinders. beakers. spectrophotometer. 125 ml Erlenmeyer flasks. burettes. Note and consider carefully: Different methods of analysis have different optimum ranges. pH meter with double-junction electrode and silver billet electrode. indicator. Use personal protective equipment: gloves. That means you will probably have to dilute the sample by the most accurate means available. MATERIALS: -millivolt meter. standard AgNO 3 solution. conc. NaHCO 3. OBJECT: -to acquaint you with the use of specific ion electrodes.
SPECIFIC ION ELECTRODE Part One: Preparation and reading standards and samples 1.in sample = mg/L Cl. Method of Standard Addition :Add 5 mls of the stock (4 mg/ml) chloride solution (V s) to the sample measured in Step 3 (E 1) and take a new reading after value is stable (E 2 ). added 2 mls of ISA. 2 minutes) and use it for all standards and samples.e. As probe tends to drift for some time it may be expedient to select a “standard time to read” (i. where: E1 = E2 = E = Vo = Vs = A = S = potential of sample (mV) potential of sample plus stock addition potential change = E 2 . 2. plot the millivolt readings (linear axis) against the concentration of the standards (log axis) to produce a calibration curve. CHLORIDE . stir and obtain a reading for it. 100 and 10 ppm (mg/L) have been prepared for you and are at the meter. Calculate the chloride concentration. rinsing and blotting dry the electrodes between each. Unless already done by a previous group (values written on the board).= ____________________ 28 September 2007 CIVL 407 Lab M anual . place the lowest standard on the stirrer and lower the electrodes into it. Standards of 1000. 3. 5. 6. Measure 100 mls of sample. mg Cl. Determine the concentration of chloride in the sample.E 1 volume of sample volume of stock solution added mg Cl.I. Repeat for rest of standards.added to the sample "slope" = change in potential over 10 fold change in concentration of standard (use data from Step 2). labelled beakers. Add 2 mls of IONIC STRENGTH ADJUSTMENT (ISA) buffer to each using the plastic dropper (filled to the mark). Turn on stirrer and while stirring record the millivolt (mv) reading when the reading appears to be stable. pour out 100 mls of each standard into separate. 4. Unless already done by a previous group. Using semi-log paper or software equivalent.
discard and refill at least twice to rinse cuvette. If sample is above the highest standard. 4.0.ions have been bound.1. which then must be subtracted from the standards and sample. the 1 ml of the indicator-acidifier reagent will adjust the pH to 2. 5. add the reagent to the second standard and so on for the rest of the standards and sample (which may need to have been diluted).ions in the sample combine with added mercuric ions forming a soluble but virtually undissociate-able complex. (Light green indicates a pH of less than 2. add 5 ml of the “combined reagent” (containing ferric alum and mercuric thiocyanate solutions) to the blank first. (Use extreme caution as this is a corrosive & toxic chemical. after a few more minutes.5 ± 0. the solution should be green-blue and will turn to pure blue (no green) within a few 29 September 2007 CIVL 407 Lab M anual . (No further adjustment is required after the blank or distilled water has been set to zero. fill. (Alternatively. and the sample (diluted if necessary) into separate and labelled 125 ml Erlenmeyer flasks (5 in total).0 ppm). using the dispenser provided.BY COLOURIMETRIC METHOD The principle of this method is based on the following formula: 2Cl.014N Hg(NO 3) 2 to a definite purple end-point.] After 10 minutes has passed (since the combined reagent was added to the blank).. it should be rerun from the start after diluting. insert in the spectrometer and record the absorbance displayed on the meter. The colour of the solution should be green-blue at this point. For most samples.II.+ Fe+++ º Fe(SCN) ++ (Reddish brown) 1.they can’t all be at 10 minutes at once.+ Hg(SCN)2 º HgCl2 + 2SCN SCN . 3. III. Using a 25 ml graduated cylinder. and Hg ++ (excess) + DPC º violet colour 3. use a plastic dropper to transfer blank into a spectrophotometer cuvette (do not fill more than 3/4 ).) Every two or three minutes thereafter you will rinse the cuvette with the next standard or sample.) Prepare a calibration curve by plotting the absorbance readings versus the concentration of chloride in the standards. 6. Determine the chloride concentration in the sample from the calibration curve. Marking the time (T=0). [Note: this delay between additions is to allow you time to measure absorbance of each at the 10 minute time required . Insert cuvette into the spectrophotometer (wavelength must be set at 463 nm) and press “zero” or “ref set” (depending on model) to make the meter read zero. the excess mercuric ions combine with diphenylcarbazone (DPC) to produce a violet colour (the end-point). Near the end-point. MERCURIC NITRATE METHOD FOR CHLORIDE DETERMINATION The principle of this method is as follows: Cl. After all Cl. 2. zero instrument on distilled water and obtain an absorbance value for the blank.0. Pour 100 ml of sample (or a portion diluted such that the concentration is less than 100 mg/L) into a 250 ml beaker.0 and 8.8. Add 1 ml (use pipette to measure accurately) of indicator-acidifier reagent to sample. 4. pure blue. measure 25 mls of “zero” standard or “blank” (halide-free distilled water). the three standards (2. 2. Titrate with 0.º HgCl2 1. Wait two or three minutes and then add the reagent to the first (lowest) standard. a pH of greater than 3. CHLORIDE .) Mix thoroughly by gently swirling the flask. Hg ++ + 2Cl.
drops of the purple end-point. Part one: Standard 1. The titre from this step must be subtracted from the titre for the sample. dilute to about 100 ml with distilled water.if one is necessary. 4. Rotate the work within the group so that different pairs are doing the titrating and recording of numbers. Determine the blank by titrating 100 ml of distilled water containing 10 mg of NaHCO 3 which serves to provide some alkalinity to the blank (to make it similar to the sample) and provides a correction for alkalinity and reagents . Place 10. Nitric acid is in the sink and is very corrosive. with standard AgNO 3.2 ml or drop-wise) should be added.1 to 0. This way each station will have data for each part. Start the stirrer and set the millivolt reading to zero or record initial mv reading [Note: Some meters can not be set to zero]. until past the endpoint when larger increments can be used again (ª mV/ml decreases). in increments. 3. and add 2. CHLORIDE BY POTENTIOMETRIC TITRATION Note: For this part form two groups comprised of 1 member from each station. A reference set of data will be provided for comparison. recording the volume and millivolt reading for each increment. Don’t forget to add the indicator.0 ml of concentrated HNO 3 (use plastic dropper). Calculate the chloride concentration: 5.0 ml (use volumetric pipette) of standard chloride solution in a 250 ml beaker. Take care that stirrer is not hitting the electrodes. where: A = ml titrant for sample B = ml titrant for blank N = normality of titrant IV.4. large increments (1-2 mls) may be added. 2. 6. The first group will do the standard (part one) in duplicate or triplicate depending on the size of the group and the other will do the sample (part two) in duplicate (or triplicate). place beaker on stir plate and lower electrodes into the solution.. Calculate the Normality of the AgNO 3 using the following equation: 30 September 2007 CIVL 407 Lab M anual . Immerse stir bar. The end-point occurs at the greatest mV change per unit addition of AgNO 3. Begin titrating. Continue the titration about 5 ml past the end-point. Plot a differential titration curve to determine the exact end-point. At the start. smaller increments (0.. as the end-point of the reaction is approached (ª mV/ml increases). then. 5.
(If pH of the sample were quite high it would be necessary to first adjust the pH to about 4 and then add 2 ml HNO 3. c. Where: A = ml AgNO 3 B = ml Blank N = normality of titrant (AgNO 3) D = ml of sample ml From graph a b c mg/L Cl- S))))))))Q S))))))))Q S))))))))Q S))))))))Q S))))))))Q S))))))))Q 31 September 2007 CIVL 407 Lab M anual . Millivolt reading versus ml of titrant. which are merely first and second derivatives.) Plot three curves for this titration: a. Millivolts per ml per ml versus ml of titrant. if dilution is necessary into a 250 ml beaker (use a graduated cylinder to measure). In plotting b and c. b. be careful to use the average value of ml of titrant on the abscissa. It is not. Millivolts per ml versus ml of titrant. in this case. Measure 100 ml of sample or portion made up to 100 ml. Calculate the chloride concentration in the sample using the following equation: 3.Part Two: Sample 1. 4. 2. Add 2 ml of HNO 3 and proceed as described in Part One.
32 September 2007 CIVL 407 Lab M anual .Potentiometric Titration RAW DATA A Volume AgNO 3 Added (Vol) (ml) B Meter Reading (E) (mV) C FIRST DERIVATIVE D SECOND DERIVATIVE F G H ª (Vol) (ml) ª (E) (mV) E ave (AgNO 3 ) Volume (ml) ª (E)/ ª (Vol) (mV/ml) ª ( ª E/ ª Vol) (mV/ml) ª [Ave(Vol)] (ml) I Ave of Ave(Vol) (ml) J ª( ª E/ ª Vol) /ml (mV/ml/ml ) Extra pages are at the back of this lab manual in appendix.Chloride .
PRETREATMENT TECHNIQUES # There are no "standard" pretreatment techniques.INTERFERENCES IN CHLORIDE DETERMINATIONS POTENTIOMETRIC METHOD # Iodide. cyanide. fluoride and bromide titrate as chloride. Each sample type requires development of an appropriate method which. in itself. Also. In the above tests. KNOWN (STANDARD) ADDITION TECHNIQUE # The known addition techniques will only correct for certain types of interferences. # Pretreatment of environmental samples usually required. # Mercury must be absent from samples. # Sample pH may need adjustment beyond the capacity of indicator-acidifier reagent. ferric. known addition cannot correct for any ions that test as chloride ion. dichromate. fluoride. hydrazine. # Colour in the sample may interfere in the absorbance measurement. Lab 3 Questions 1. 33 September 2007 CIVL 407 Lab M anual . # Environmental samples usually require pretreatment. # Other interferences: ferricyanide. It can only correct for substances that enhance or suppress the test response proportional to the amount of chloride ion present. This is a complex. timeconsuming process requiring a sound knowledge of chemistry and extensive experience working with environmental samples. # High levels of ions which form very insoluble salts of silver may deposit a layer of salt on the electrode membrane. 3. SPECIFIC ION ELECTRODE # All halides interfere (the electrode is a halide electrode). # In practice. fluoride and bromide titrate as chloride. # Environmental samples usually require complex pre-treatment. From your chloride results. causing electrode malfunction. ferric ion. chromate. thiosulphate. specific ion electrodes are rarely usable in environmental samples. requires knowledge of the compounds in the sample which are likely to interfere. and nitrite interfere. MERCURIC NITRATE TITRATION METHOD # Iodide. 2. COLOURIMETRIC METHOD # Bromide. and sulfite ions interfere when concentration greater than 10 mg/L. can you see any advantage in suing the first or second derivative curve? Discuss. strongly reducing solutions may form a surface layer of silver.3 mg/L? Outline the advantages and disadvantages of all the methods used in this lab to measure chloride concentration. # Chromate. iodide. In the Mohr method for chlorides (described in Sawyer and McCarty) what would the equilibrium concentration of silver ions be (in mg/L) when the chloride concentration has been reduced to 0. # Colour may obscure or interfere with end-point determination.
Materials: -600 ml beakers. (concentrated) glacial acetic acid. to illustrate the concepts of free and combined chlorine residuals. clean up all spills with a brush into a container and wash crystals down the drain. 250 ml Erlenmeyer flasks. potassium iodide (KI) crystals. Titrate rapidly with the stock chlorine solution (in burette) until the appearance of a constant 34 September 2007 CIVL 407 Lab M anual .total chlorine residual) 1. REFERENCES: Standard Methods 19th ed. Mix well (on stir plate with stir bar).. OBJECT: -to acquaint you with disinfection of water supplies and to demonstrate techniques for the determination of chlorine residuals.025 N sodium thiosulphate.N-diethyl-p-phenylenediamine (DPD) indicator solution. burettes and stands. When weighing chemicals at the balance. about 50 ml of chlorine-free water (distilled water will do). pages 4-56 to 4-57 or older/newer editions. 250 ml volumetric flasks. chlorine demand and break-point chlorination. gloves and lab coats. and exactly 10 ml of 0. N. Take a 250 ml Erlenmeyer flask and add about 1 gram of potassium iodide. Wipe up all liquid spills immediately. Rinse a burette and fill it with this stock chlorine solution. allow to mix again. phosphate buffer solution. PREPARATION OF STOCK CHLORINE SOLUTION (Iodometric method . Make up to the 250 ml and invert flask several times to mix. Personal protective equipment required at all times when working in the lab are: eye protection. 1995. 3. I.La b o ra to ry 5: Ch lo rin e a n d Ch lo rin e D e m a n d WARNING Glacial acetic acid is very corrosive. Bleach/chlorine solutions are strong oxidants. add 1 ml of starch solution.025 N sodium thiosulphate solution (use a volumetric pipette). standard ferrous ammonium sulphate (FAS) solution. Prepare a stock solution of strong chlorine water by adding 5 mls of bleach solution (sodium hypochlorite) to about 200 mls of water in a 250 ml volumetric flask. bleach or hypochlorite solution. pipettes. 0. 2. starch indicator. about 5 ml of glacial acetic acid (use a graduated cylinder). 100 ml graduated cylinder. chlorine free water.
Place 5 ml of phosphate buffer solution and 5 ml of DPD solution in a 250 ml conical (Erlenmeyer) flask and mix. as there is lots to do . a) Free chlorine: titrate rapidly with standard FAS (Ferrous ammonium sulfate) titrant until the red colour is discharged. 4. 2. titrate to colourless again. 35 September 2007 CIVL 407 Lab M anual . Method 4500-Cl F. mix to dissolve.A Monochloramine= B-A Dichloramine= C-B For interferences. Work in groups of three or four for this part.Breakpoint Chlorination using Free and Combined Chlorine Residuals Values 1. d) Alternatively: (Not to be done in this lab.) Free and combined chlorine or total chlorine: To obtain total chlorine in one reading.). c) Dichloramine: Add about 1 g KI (to the same sample) and mix to dissolve. b) Monochloramine: Add two drops of KI solution (to the same sample) and mix. For dichloramine concentrations greater than 1 mg/L. IIb. calculate the chlorine concentration in the stock solution (See Data and Results section for formula). Continue titrating until red colour (if any) is discharged once again (Reading B). 3. From the amount of chlorine solution used. 18th edition. 2. add full amount of KI at the start (ie 2 drops KI solution and 1 g of KI crystals) to a fresh sample. let stand for two minutes and titrate until the red colour (if present) is discharged.COORDINATE! IIa. Allow at least 5 minutes of time to elapse between applications of chlorine doses (use the buret containing diluted chlorine solution from Part I above) to successive beakers in order to allow enough time for titrating the free and combined chlorine residuals. Mix usual volumes of buffer reagent and DPD indicator solution with distilled water b e fo re adding sufficient sample to bring total volume to 100 ml or the test will not work. (*NOTE: If total chlorine exceeds 5 mg/L use a smaller sample and dilute to a total volume of 100 mls with chlorine-free water. etc. Sequentially add the necessary volumes of chlorine stock solution to each beaker at the appropriate time spacing to provide the applied dosages given by the instructor (see board). After 15 minutes of contact time. Determination of Free and Combined Chlorine by the DPD Method 1. Let stand 2 minutes more. Add 100 ml of sample or diluted* sample using a 100 ml graduated cylinder and mix again. pp 4-43. These 6 flasks can be prepared at once to make them ready for the next step. refilling burette. Set up a numbered row of six 600 ml beakers each containing 500 mls of the sample of water provided (it has been prepared using Ammonium chloride and Glutamic acid). It is likely that the highest dose will require dilution. Record the burette reading (mls) = Reading A. if colour drifts back (indicating an incomplete reaction). Go to c. let stand 2 min more if colour drifts back indicating incomplete reaction. Total chlorine (Free plus Combined) = Reading C Free residual chlorine = Reading A Combined residual chlorine (dichloramine plus monochloramine) = Reading C . Record the burette reading (Reading C). and again after 1 hour of contact time determine the free and combined chlorine (see below) in 100 ml samples taken from each of the beakers at the appropriate time spacing (15 minutes and 60 minutes after the addition of the chlorine dose to each beaker).purplish-blue colour. please refer to Standard Methods. Let stir for 2 min and then continue titrating until red colour (if any) is discharged (Reading C).
Standardization of chlorine stock solution: Volume of sodium thiosulphate used Normality of sodium thiosulphate used Volume of stock chlorine solution Cl concentration of stock solution = = = = ______ mls (A) ______ (N) ______ mls (B) = _________mg/L Cl as Cl2* *To determine bleach concentration as percent you must multiply first by your dilution (i. b. free and combined residual chlorine for each applied chlorine dose. Under what conditions is the practice of break point chlorination necessary? 4. Why is it important to determine chlorine residuals in domestic water supplies? 2. How would you expect the forms of residual chlorine and their speed of formation to change as a function of 1) temperature and 2) pH? Lab 5 Questions 1. d. If a sewage sample contains 10 x 10 6 coliforms/100 ml. c. Make a separate graph for the 15 minute and 1 hour contact time. Make a plot of the three residual chlorine components against the applied chlorine dosage. II.5 mg/L if 50% are killed in 1½ minutes at this concentration? b. Given: HOCl W H + + OClK eqv = 2.DATA AND RESULTS I. 250) then convert mg/L to g/100 mls = %. If this is true. a. The rate of kill of bacteria by chlorination follows first-order reaction kinetics. FAS titrant concentration is 1 ml = 100 µg Cl as Cl2 a.e. what percentage of bacteria would be killed in 10 minutes at a chlorine residual of 0. Guaranteed analysis is normally printed on the container and is usually 5-6%. Discuss your results as a function of the residual forms of chlorine present at different chlorine dosages and at different contact times. Determine the concentration of total.7 x 10-8 at 25 o C % HOCl = 27 What is the pH of the solution? 3. how many coliforms would remain after a chlorine contact time of 10 minutes? 20 minutes? 36 September 2007 CIVL 407 Lab M anual .
15 MINUTE CONTACT TIME BEAKER NUMBER Vol. of Cl Soln. (mls) (Stock=_____mg/L Cl as Cl2) Applied Cl dosage (mg/L) Volume of FAS to endpoint A Volume of FAS to endpoint B (includes A) Volume of FAS to endpoint C (includes A & B) Free chlorine (mg/L) (A) Monochloramine (mg/L) (B-A) Dichloramine (mg/L) (C-B) Combined residual chlorine (mg/L) (C-A) Total chlorine (mg/L) © Time of Cl addition Time of Residual measurement 1 2 3 4 5 6 37 September 2007 CIVL 407 Lab M anual .
(mls) (Stock=_____mg/L Cl as Cl2) Applied Cl Dosage (mg/L) Vol. of FAS to endpoint C (mls) Free chlorine (mg/L) (A) Monochloramine (mg/L) (B-A) Dichloramine (mg/L) (C-B) Combined residual chlorine (mg/L) (C-A) Total chlorine (mg/L) © Time of Cl addition Time of residual measurement 1 2 3 4 5 6 (includes A+B) 38 September 2007 CIVL 407 Lab M anual . of FAS to endpoint A (mls) Vol.60 MINUTE CONTACT TIME BEAKER NUMBER Vol. of FAS to endpoint B (mls) (includes A) Vol. of Cl Soln.
turn on stirrer and obtain a stable reading. if above 7.) 2. pH test paper. standard base. magnetic stirrer. burettes and stand. Hard n e s s . turbidimeters. 39 September 2007 CIVL 407 Lab M anual . (Take care not to contaminate paper in case.Lab o rato ry 6: Alkalin ity . I. particularly those labelled with specific hazards.(Do not allow the stir bar to hit the probe. Probes. phenolphthalein. Check the pH of a small aliquot of the sample by wetting a pH test paper and comparing the colour combinations to those shown on the case. metacresol purple. then use buffers 7 and 10. place a beaker containing the sample and stir-bar on the stir-plate. Co lo u r. DETERMINATION OF pH 1. water samples. pH test paper and other water quality assessment equipment. lower probe. Use care when using lab equipment. bromphenol blue. rinse & dry probe. Calibrate the pH meter by following the procedure outlined on the instruction sheet beside the meter unless told otherwise. beakers. 3. Normally. EDTA. After calibration is complete. then use buffers 4 and 7 to calibrate. buffer solution. metres and calibrated glassware are available in limited quantities and are very expensive to replace. Rinse probe when you are finished and leave immersed in pH 7 buffer for storage. MATERIALS: -pH meter. p H. T u rb id ity WARNING Please use caution when handling all chemicals. hardness indicators. colour comparator. Always wear personal protective equipment. standard acid. 4. Be sure that buffers are being stirred when you perform the calibration and that when you change buffers you thoroughly rinse the probe into a waste beaker and blot dry so that you do not contaminate other buffers or your samples. graduated cylinders. flasks.) Record value in following table. OBJECT: -to familiarize you with the most common water treatment tests. bromcresol green. the two buffers that are chosen for calibration will bracket your sample ie if sample is less than 7. Ac id ity . 1 N NaOH. to acquaint you with the use of pH metres.
The buffer elevates the pH to 10. if the approximate hardness is known (by unsuccessful attempts). 3. 2. Titrate to the first uniform pink colour (colour stays). 40 September 2007 CIVL 407 Lab M anual . (If the solution is already blue after indicator. Apply dilution factor to obtain final result. until the reddish tinge disappears from the solution and a pure blue remains. Add a full dropper of bromphenol blue indicator and titrate until the colour changes from yellow to blue ("MO" Acidity). In order to remove any free residual chlorine (which interferes with the indicator colour response). 5. 3. Measure out a new sample and add 1 drop of sodium thiosulphate and a full dropper of phenolphthalein.. DETERMINATION OF ACIDITY 1. Rinse and fill another burette with standard base solution and label it. Add 1 drop of sodium thiosulphate solution. Add 1-2 ml of buffer solution using dropper bottle (3 full droppers) and one aliquot of Total Hardness Indicator using the special dispenser on the chemical bottle.think what that means!) 4. TOTAL HARDNESS DETERMINATION . White paper placed under the flask will facilitate this determination. add 90% of the titrant to the sample before adjusting the pH with buffer. Measure 100 ml of water sample into a clean. DETERMINATION OF ALKALINITY 1..II. If sample is highly alkaline (titration goes over one burette full) dilute the sample and titrate again.?) 4.5). add 1 drop of 0. 4. 2. 3. If a ppt does form within the 5 minutes.. (BG or MO) 6. Add a dropper full of bromcresol green (BG= MO) to this same water sample and continue the titration until the colour changes from blue to yellow (at pH 4.1 N sodium thiosulphate to the sample. IV. titrate until just colourless. Add a full dropper of phenolphthalein (P) indicator and if the solution stays pink or red. Add a stir-bar. (If solution is clear after addition of P. Record burette reading (P). Rinse and fill a labelled burette with standard acid solution. III. Pour into a 250 ml Erlenmeyer flask. Rinse and fill a burette with standard EDTA solution and record the initial burette reading. Add stir-bar. it may be necessary to repeat the titration with the sample diluted 1 to 1 (again) with distilled water. Record the volume of titrant required in the following table. Titrate slowly with EDTA. Measure 100 ml of sample (with minimum agitation or exposure to air) using a graduated cylinder and transfer carefully to an Erlenmeyer flask. while stirring. sample-rinsed graduated cylinder and transfer to an Erlenmeyer flask.. Complete the titration within 5 minutes to minimize the tendency toward CaCO 3 precipitation. 2.EDTA TITRIMETRIC METHOD 1.. A pH much above 10 promotes CaCO 3 precipitate. Record the burette reading in the following table. Measure out 25 mls of sample using a graduated cylinder and dilute to 50 ml with distilled water. Alternatively. Add the last few drops at 3-5 second intervals so that you do not "overshoot" the end-point.
EDTA TITRIMETRIC METHOD 1. Select signal averaging mode by pressing the SIGNAL AVERAGE key. The display will show SIG AVG when the instrument is using signal averaging. Use signal average mode if 41 September 2007 CIVL 407 Lab M anual . Values are reported as APHA Colour Units (eqv. Measure out 50 ml of sample into an Erlenmeyer flask and add 2. VI. . except on an unfiltered sample. taking care to handle the sample cell by the top.HACH Spectrophotometer. with continuous stirring to a pure blue end-point. 2. 4. TURBIDITY .0 ml of 1 N NaOH solution (use 3 droppers full).Enter the stored program number for true colour=120 1. Cap the cell. 4. 2. or a volume sufficient to produce a pH of 13-14 (designed to precipitate the magnesium as a hydroxide)... Place the cell containing sample in it and press Read. 3. Insert the sample cell in the instrument cell compartment so the diamond or orientation mark aligns with the raised orientation mark in front of the cell compartment. Close the lid. Record the final burette reading in the following table. Add one aliquot of the Calcium Hardness Indicator and titrate with EDTA slowly. 3. Record the final burette reading in the following table.5. Pour the supernatant into one of the cells (to the etched mark) and fill another with distilled water. Fill a sample cell to the line (about 15 mL). V. 5. Wipe the cell with a soft.TRUE AND APPARENT PART A: TRUE COLOUR . Select automatic range by pressing the RANGE key.BY HACH 2100P Turbidity Meter. Use the technique outlined in Part A. The instrument will turn on but will turn off automatically after a short time so you should be ready. COLOUR . Add a stir-bar and stir to mix. 3. 2. The instrument may suggest diluting if over range. 4. Place the cell containing water into the spectrophotometer and press Zero. to determine the apparent colour (measures the influence of turbidity on the reading). VII.. Demonstration of Jackson Candle and Hellige PART A: Hach Turbidimeter 1. lint-free cloth to remove water spots and fingerprints. Filter about 20 mls of sample using a filter funnel and stand and the pre-fluted filter paper provided 2. The display will show AUTO RNG when the instrument is in automatic range. Press: I/O. The reading might be higher so if you had to dilute in Part A dilute for Part B. Refill the burette with EDTA titrant and note the initial reading. CALCIUM HARDNESS DETERMINATION . Record all colour data in the following table. to mg/L platinum as chloroplatinate) PART B: APPARENT COLOUR 1.
2.adding liquid to a hot tube will cause it to shatter. Close the door and switch on the light. page 272-273. then the turbidity in NTU. PART C: Jackson Candle Turbidity . Press: READ The display will show . 3. Read the scale on the dial. 16th edition. 42 September 2007 CIVL 407 Lab M anual .. record the number and refer to the appropriate graph to determine the turbidity as mg/L SiO 2. If you take too long. 2. obscuring vision.U. Carefully fill the 20 mm viewing depth tube to the mark with sample..Demonstration only 1. Take the reading where the dark central part first merges with the surrounding field. Remove about 10 mls of sample by pipette and slowly dispense this aliquot back into the tube until the shape of the flame is no longer distinguishable. DO NOT PLACE AN EMPTY TUBE IN THE TUBE HOLDER WITH THE CANDLE LIT .. Try to be quick and not burn the candles more than a few minutes at a time. (This allows you to make a more gradual and accurate approach to the end-point.NTU.T.. Result of Titration Hydroxide Alkalinity as CaCO 3 Carbonate Alkalinity as CaCO 3 Bicarbonate Concentration as CaCO 3 T T-2P 0 0 0 P=0 0 0 P<1/2T 0 2P P=1/2T 0 2P P>1/2T 2P-T 2(T-P) P=T T 0 Where: P = Phenolphthalein alkalinity.s. Note that there are several different scales on the side of the tube. Record the turbidity after the lamp symbol turns off. the turbidity may start to settle and condense on the bottom of the tube. Wipe the outside of the tube to dry it and place it in the turbidimeter (in the circular groove of the mirror unit).demonstration only. Lower the plunger into the liquid making sure no bubbles are trapped under the plunger and the top is dry. 1.. that the filter selector shows "none" and that bulb B is in use. 3. Pinch all excess charred wick with fingers before lighting or soot may be deposited on the glass tubes. Note that the rectangular door mirror is closed. Be sure that the calibrated tubes are clean before adding samples to the tubes. See Standard Methods.the sample causes a noisy signal (display changes constantly).) Remove the glass tube from the holder and read the turbidity at the meniscus of the sample. 6. Pour sample into the tube in small increments until you can no longer distinguish the shape of the candle flame. T = Total alkalinity. Immediately balance the light intensity of the central spot with the surrounding observation field by turning the dial on the right-hand side of the instrument. PART B: Hellige Turbidimeter . Record value in following table as J.
Sample volume "M.Sample volume Initial burette reading P." titre (mls) Net Total acidity titre (mls) Net CO 2 Acidity titre (mls) Final burette reading (mls) Initial burette reading (mls) Net titre (mls) Total Hardness as mg/L CaCO 3 CALCIUM Sample volume (mls) Final burette reading (mls) Initial burette reading (mls) Net titre (mls) Calcium as mg Ca ++ /L __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ __________ ___________ ___________ ___________ ___________ ___________ ___________ ___________ ___________ Sample 2 ___________ ___________ P.DATA Sample 1 pH .O. Alkalinity titre (mls) Net "M." Alkalinity titre (mls) ACIDITY." Alkalinity end-point (mls) Initial burette reading Net P.O. Acidity end-point reading (mls) Initial burette reading Net "M." Acidity end-point reading (mls)__________ TOTAL Hardness Sample volume (mls)__________ Calcium hardness as mg CaCO 3/L __________ 43 September 2007 CIVL 407 Lab M anual . Alkalinity end-point reading (mls) __________ "M.O.O.Initial pH by test paper Initial pH by pH meter ALKALINITY.
0 Alkalinity: where: A= N= ml standard acid used normality of standard acid Acidity: where:A = N= ml standard base used normality of standard base 44 September 2007 CIVL 407 Lab M anual .CALCULATIONS Calcium: Calcium Hardness: where: A= B= ml titrant for sample mg CaCO 3 equivalent to 1.0 Total Hardness (EDTA): where: A= B= ml titrant for sample mg CaCO 3 equivalent to 1.00 ml EDTA titrant at the calcium indicator endpoint = 1.00 ml EDTA titrant at the calcium indicator endpoint = 1.
" Acidity as CaCO 3 (mg/L) Total Acidity as CaCO 3 (mg/L) Free carbon dioxide as CO 2 (mg/L) Total Acidity as CO 2 (mg/L) Ca Hardness as mg Ca +2 (mg/L) Ca Hardness as CaCO 3 (mg/L) Mg Hardness as CaCO 3 (mg/L) Mg Hardness as Mg+2 (mg/L) Carbonate Hardness as CaCO 3 (mg/L) Non-carbonate Hardness as CaCO 3 (mg/L) Excess alkalinity as CaCO 3 (mg/L) Colour .ion (mg/L) CO 3 -2 Alkalinity as CO 3 -2 ion (mg/L) HCO 3.Alkalinity as CaCO 3 (mg/L) CO 3 -2 Alkalinity as CaCO 3 (mg/L) HCO 3.Units= N.ion (mg/L) "M.O." Alkalinity as CaCO 3 (mg/L) OH .T.Alkalinity as HCO 3 .Hach .Alkalinity as CaCO 3 (mg/L) OH . ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ 45 September 2007 CIVL 407 Lab M anual .APHA Colour Units: True Apparent Turbidity .RESULTS (sample) P.U.Alkalinity as OH .O. Alkalinity as CaCO 3 (mg/L) "M.
non-carbonate and total hardness of this water in mg/L as CaCO 3 ? in meq/L? Is the analysis complete? Why? 6.Lab 6 Questions 1. 46 September 2007 CIVL 407 Lab M anual . A water has the following analysis: Na + K+ Ca +2 15 mg/L 33 mg/L 12 mg/L ClSO 4-2 HCO 3 70 mg/L Mg+2 18 mg/L 42 mg/L 8 mg/L What is the carbonate. calculate the carbonate alkalinity in mg/L as CaCO 3 for the water sample analyzed during this lab. Is this water suitable for a domestic water supply? Why? 2. From equations (17-12) and (17-13) in Sawyer and McCarty.7 x 10 -11 and K w = 10 -14. given K 2 = 4. What sanitary significance does colour have? 4. Note: You should bring a copy of this lab to the next two sessions as you will need the same instructions to perform the same tests in the Coagulation and Softening Labs. Does this agree with the results from Part II of this lab? Explain. What errors can occur in the calcium hardness determination? Why? 3.
No. Several lime and soda ash dosages will also be selected for determining the efficiency of water softening. You should have with you the part of Exercise 7 that gives instructions for the various tests. In the second lab you will use the previously determined dosages to coagulate and soften the sample and then re-analyse the resultant water. No. The first step will be to analyse the raw water sample and the supernate from a number of alum dosages (to be announced in the class) using the techniques learned in Lab. 47 September 2007 CIVL 407 Lab M anual . and all of the materials from Lab. alum solution (Al2 (SO 4 )3 @18H 2 O). 1000 ml beakers.6.for the next session (b). soda ash solution (Na 2CO 3) and lime (Ca(OH)2) or sodium hydroxide (NaOH). MATERIALS: -Jar Test Apparatus. OBJECT: -to illustrate the principles of coagulation and water softening. to be conducted over two lab periods.Lab o rato ry 7a: Co ag u latio n an d 7b : Co ag u latio n & So fte n in g ATTENTION This lab will require patience and cooperation from all of you as we have only two jar testers with 6 paddle sites each. Please try to coordinate your work with the rest of the class. 7 (except Jackson Candle and Hellige Turbidimeter). This is a two-step experiment. The optimum alum dosage for coagulation of the sample will then be chosen.
You will also determine the optimum lime and soda ash dosages to soften this water and select a range of dosages surrounding this optimum for investigation next week. Stop stirrer. acidity. 7. 5.Part A:. not a beaker ) and transfer to 1000 ml beakers. small. cloudy). Stop stirring. Add the assigned alum dosages as quickly as possible and then start the stirrer. You may need to use blocks to position the paddle 0. After the flocs have settled (allow 15 min). Analyze these supernates plus a sample of raw water for pH. Add the designated lime (or NaOH) and soda ash additions as quickly as possible to the decanted samples. true colour and turbidity (Hach). Complete the coagulation summary sheet during the lab period. 6.work in 2 or 3 teams to assess a total of 6 dosages for each team. In the lecture. Determine which dosages were most appropriate for this sample. 2. hardness (total and calcium). 4. acidity. hardness (total and calcium). position them under stirrers. Lift paddles out of beakers and secure. hazy. clear.. then reduce stirring rate to about 20 rpm for 10 minutes (just fast enough to keep the floc suspended but not so fast that the floc is sheared). Stir at 100 rpm for 1 minute and at 20 rpm for 10 minutes. Pour six 1000 ml aliquots of sample into 1000 ml beakers and place on the jar test stirrer apparatus. times of appearance and descriptions and record. Make sure it is centred. 1. if available. 3. moderate. Record relative floc sizes (pin-point. 5. Discard the floc and clean beakers in preparation for a second decanting in Step 4. Record observations on settling rate. clarity of supernatant liquid (very clear. large). and fill in the table on the board. COAGULATION USING ALUM . Stir at 100 rpm for 1 minute. 4. but because of equipment limitations. use indicators as specified in Lab 6. alkalinity. Stop stirrer and allow floc to settle. 6. you will discuss the data and select the appropriate alum dosage to be used by all groups in the water softening step next week. A pH meter could be used for the alkalinity and acidity determinations. medium. Stir at 100 rpm for 1 minute. floc volume. Observe and make notes describing floc formation and note the time of appearance. if time permits. turbidity and colour. drop of 0. Measure out six 1000 ml aliquots of sample (using graduated cylinder to measure. After the floc has settled (allow 15 min). decant the supernatant liquid carefully into another set of beakers (try not to stir up what has settled). (NB dosage now based on 800 ml sample size). Analyze the supernates for pH. Brief Methods Summary for Analyses Alkalinity: @ 100 ml of sample. decant the supernatant liquid carefully into a clean set of beaker (try not to stir up what has settled) and discard the floc. Reduce the stirring rate to about 20 rpm for 10 minutes. alkalinity. Carefully decant into a graduated cylinder 800 mls of the supernatant liquid and pour into fresh beakers. Place beakers on the jar tester apparatus. 3. and clarity of supernatant liquid. Part B: WATER SOFTENING PROCEDURE (to be done the following week) again work as teams of 2 or 3. 2. and settling characteristics of the floc (rapid. slow).1 N sodium thiosulphate 48 September 2007 CIVL 407 Lab M anual . Add the appropriate alum dosage and immediately start stirring. Make relative estimates of floc sizes.5 to 1 cm off the bottom of the beaker. 1.
@ calculation: Ca hardness (mg/L CaCO 3 )= mls titre x 1000/mls sample Lab 7 Questions A.@ titrate with 0.pH 3. Coagulation 1. What treatment would you recommend to prepare this water for human consumption? Why? 49 September 2007 CIVL 407 Lab M anual . 3. Compare removal of hardness components under the different doses .3 ("phenolphthalein" or total acidity) @ calculation . Based on your chemical analysis of the raw water. How does the consumption of alkalinity by alum compare with theoretical calculation based on balanced equations? How much extra soda ash should you add to compensate for the alkalinity consumption by 70 mg/L alum? B.pH 8.alkalinity (mg/L CaCO 3)= mls titre x N acid x 50.1 N sodium thiosulphate @ titrate with 0.acidity (mg/L CaCO 3)= mls titre x N base x 50. @ calculation: hardness (mg/L CaCO 3 )= mls titre x 1000/mls sample Calcium Hardness: @ 50 ml sample @ add 1 ml 1 N NaOH (or to pH 13) and Ca Hardness Indicator (Hydroxy Naphthol Blue) @ titrate quickly with EDTA to blue end-point (no pink/purple tinge .3 and with bromcresol green (continuing with the same sample) to yellow . Softening 1.02 N base with bromphenol blue to blue .pH 4.02 N acid with phenolphthalein to colourless .use graphical presentation.pure blue).000/mls sample Acidity: @ 100 ml of sample. 2.5. Compare your experimental softening results to the theoretical results based on solubility equations or mass balance equations.7 ("methyl orange" acidity) and with phenolphthalein (using a fresh sample) to pink .pH 8. @ calculation . 2. 4. Analyze the response of colour and turbidity removal as a function of alum dose. show by calculation the theoretical amount of lime and soda ash required for excess lime/soda ash softening. drop of 0.pure blue).000/mls of sample Total Hardness: @ 25 ml sample diluted to 50 ml @ add 1 ml buffer and Total Hardness Indicator (Calmagite) @ titrate quickly with EDTA to blue end-point (no pink/purple tinge .
7 “MO” Net titre to pH 8.5 “TOT” Total Hardness sample vol Net titre Ca Hardness .sample vol Net titre Notes: 50 September 2007 CIVL 407 Lab M anual .5 “P” Net titre to pH 4.sample vol ml Net titre to pH 3.Coagulation Raw Data Sheet DATA Raw Sample Alum mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L pH Alkalinity-sample vol ml Net titre to pH 8.5 “TOT” Acidity .
) (filtered) Turbidity (N.) unfiltered Notes: 51 September 2007 CIVL 407 Lab M anual .) True Colour (A.H.A.T.Coagulation Summary Sheet DATA Raw Sample Alum mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L mg/L pH Floc size Clarity Floc settling rate Alkalinity (mg/L CaCO 3 ) “P” Alkalinity (mg/L CaCO 3 ) “TOT” Acidity (mg/L CaCO 3 ) “MO” Acidity (mg/L CaCO 3 ) “TOT” Total Hardness (mg/L CaCO 3 ) Ca Hardness (mg/L CaCO 3 ) Apparent Colour (A.P.P.A.U.H.
Coagulation and Softening Raw Data D ATA Lime/NaOH mg/L Soda Ash mg/L Alum mg/L pH Alkalinity-sample vol ml Net titre to pH 8.sample vol ml Net titre to pH 3.5 Total Hardness sample vol Net titre Ca Hardness .5 Net (Total) titre pH 4.sample vol Net titre Notes: 52 September 2007 CIVL 407 Lab M anual .7 Net (Total) titre pH 8.5 Acidity .
-mg/L CaCO 3 Apparent Colour (A.H. -mg/L CaCO 3 **Non-Carb Hard -mg/L CaCO 3 **Excess Alk.Alkalinity -mg/L CaCO 3 *Total Alkalinity-mg/L CaCO 3 “MO” Acidity -mg/L CaCO 3 *Total Acidity -mg/L CaCO 3 Total Hardness -mg/L CaCO 3 Ca Hardness -mg/L CaCO 3 Mg Hardness -mg/L CaCO 3 **Carbonate Hard.H.P.P. Calculation 53 September 2007 CIVL 407 Lab M anual ** By .A.U.) Note:* Total Alkalinity must include Phenolphthalein Alkalinity just as Total Acidity must include “MO” Acidity.A.T.) True Colour (A.Coagulation and Softening Summary Sheet Data Lime/NaOH mg/L Soda Ash mg/L Alum mg/L pH P.) (Filtered) Turbidity (N.
. . . 91 18. . . . . . . . . . . . . . . Acidity. . . . . . . . . . . . . . . . . . . . . . . . . . . BOD. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Alkalinity. . . . Milliequivalent and mg/l as CaCO 3 . . . .4 Appendices see: CIVL407Manual_CourseNotes. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 112 28. . . . . . . . . . . . . . . . . . . . . . . . . . . . 105 25. . . . Hach Absorption Method . 92 19. . . . Chlorine . . . . . . . . . . . . . . . . 83 16. . . . . . . . . . . . . . 72 11. . . . . . . . . . . . . . . . . . . . 102 23. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Bar Diagram Method for Water Softening Problems . . . . . . . . . . 107 26. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84 17. . . . . . . . . . . . . . . . . . . . . . . 74 13. . . . . . . . . . . . . . . . . . 96 21. . . . . . 100 22. . . Titrations . . . . . . . . . . . . . . . . . . . Alkalinity Species as a Function of pH . . . . . . . . . . . . . . . 80 14. . . . . . . . . . . . . Reading and Reference Material . . . . . . . . . . . . . . . . . . . . 57 3. . . . . . . . . . . . . Periodic table . . . . . . . . . . . . . . . . . . . . . . . . . . . . Bacteriological Examination . . . . . . 69 8. . . . . . . . . . . . 104 24. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Hardness Complexes . . . 73 12. . . . . . . . . . . . . . . . . . . . . . pH . 117 29. . . . . . . . . . . . . . . . Standard Solutions. . . . Hardness. . . . . Normal Solutions and Equivalent Weights . . . . . . . . . . Water Softening . . . . . . . . . . . . . . . . . .COD . . . . . . 55 2. . . . . . . . . . Balancing an Oxidation Reduction Equation . . . . . . . . . . . . . . . . . . . . 71 10. . . . . Chemical Requirements for Water Softening . . . . . . . . . Faecal Coliforms . . . . . . . . . . . . . . . . . . . 118 54 September 2007 CIVL 407 Lab M anual . . . . Concepts and Definitions . . . . . . . . . . . . . . .pdf 1. . . . Instructions for Laboratory Report Writing . . . . . . . Chloride Analytical Methods . 94 20. . . . . . . . . . . .Chlorine Demand . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Solids Removal in Wastewater Treatment . . . . . . . . . . . . . . . . 82 15. . . . Known Addition Technique . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68 7. . . . Introduction . . . . . . . . . . . . . . . . . . . . .Levels and Water Use . . . . . . . . . . . . Extra tables for Chlorine Lab . . COD . 70 9. . Equilibrium Relationships of Chlorine . . 60 4. . . Coagulation . . . . . . . . . . Colour. . . . . . . 64 5. . . . . . . . . . . . 110 27. . . . . . . . . . . . . . . . . . . . . . . . . . . . Typical Problems for Acid-Base and General Chemistry . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .Additional Notes . . . . . . . DO. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Bacteriology and Water-related diseases . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Turbidity. . . . . . . . . . . . . . . . . . . . . . . . . . . . Residues . . . . . . . . . 67 6. . . . . . . . . . . . . . . . . . . . .
This action might not be possible to undo. Are you sure you want to continue?
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