CIVL 407 Lab Manual

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

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Course Description
(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:

kvlo@interchange.ubc.ca scharper@civil.ubc.ca barathird@yahoo.co.uk parvez@interchange.ubc.ca

(604) 822-4880 (604) 822-4397

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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
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September 2007 CIVL 407 Lab M anual

2. Pipettes must be placed with tips pointing up into the pipet basket supplied. Most pipettes are calibrated “to deliver” or TD and should never be blown out. Clean-up: You must leave your work station as you found it . WHMIS: Workplace Hazardous Materials Information System is a government regulation. Pipettes/ graduated cylinders/ beakers/ flasks: 5) 6) 7) 8) 9) 10) Volumetric pipettes are the most accurate way to measure volume. 25.0. but if you do please ask for assistance. 1. eye protection. Safety Equipment: Eye Wash Station. Spills: Wipe up all spills immediately. calculators. Be aware that handling your pens.2 First Session: Laboratory Safety and Orientation 1) 2) 3) 4) NO Food or drink. Some pipettes may be “to 3 September 2007 CIVL 407 Lab M anual . Personal Protective Equipment: Lab coat (long). They are calibrated by weighing the volume of distilled water that will flow from them by gravity. 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. if there is room.5.. You must be informed of the hazards of a substance before using it. Do not put lids and small items onto racks that may slip through to the filthy drip tray. If you cut yourself. 20. Wearing lab coats out of the lab is forbidden.0 ml. They are available in 0..clean and dry. Remember to empty and rinse burets as well.up to 10. Sanitation: antibiotic soap for hand washing. Broken Glass: Be careful not to break glass. Safety Shower. please get attendant help. gloves as required.0. MSDS: is a supplier information sheet that must be available for each product and describe the hazards and necessary handling requisites. DO NOT PUT BROKEN GLASS INTO THE REGULAR GARBAGE CONTAINER. 15. 50 and 100 ml. no matter how minor. wash down and dry the bench. Hygiene: At times you will be working with sewage. Please take your labcoat away in a plastic bag. disinfectant for bench surfaces. with the tip against the side of the receiving vessel. A small amount of liquid always remains in the tip and must not be blown out. No sandals or open-toe or heel shoes. packs. These should be used whenever standard solutions are measured or when upmost accuracy is required. 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. Please ask for assistance if chemicals are spilled. etc.

2 & 4 L Beakers: are cylindrical in shape with straight sides and flat bottoms. LISTEN AND MAKE NOTE OF VERBAL AND CHALK BOARD INSTRUCTIONS. Use the size of cylinder CLOSEST TO THE VOLUME YOU WISH to measure for best accuracy. 4 September 2007 CIVL 407 Lab M anual . do not use a 1 L graduated cylinder. not wastefully as quantities are limited. Erle n m e y e r or conical flask markings are not accurate. Make sure that you label beakers and flasks to avoid confusion and waste. Chemical residues can be splashed in you face. They are useful for measuring samples 20 mls . especially at smaller volumes. Flasks: Vo lu m e tric flasks are very accurate and should be used when diluting standards (together with volumetric pipettes). markings are not accurate at all . They are containers best suited for “swirling” liquid ie to mix reagents with samples in a colourimetric determination (Lab 4 or 6). 1. please rinse out immediately and set aside to dry. If liquid should enter a bulb or pump. 13) Stir bars: Please remove from containers before pouring contents down the drain. YOU WILL USUALLY NEED THE INFORMATION GIVEN TO COMPLETE YOUR LAB WRITE-UPS. Used to hold/transfer approximate volumes of samples. 12) Cup sinks: Please do not use these sinks for the disposal or delivery of liquids.pay attention to the markings. Even the force of air from the air taps can be dangerous because it can contain grit or water. 14) Glassware/supplies: Each station has a drawer where some glassware and other useful tools will be kept. 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. Undetected gas leaks can be deadly. 500 mls.. They are calibrated the same way as TD except that the remaining drop is blown into the receiving vessel. NEVER mouth pipette. 11) Gas/air/vacuum taps: Do not touch these unless instructed to do so.1000 mls. They are usually identified by a double etched or coloured band at the top of the pipette. 10 mls. chemicals or for titrations. 25 mls. Cylinders are available: 5 mls. A demonstration will be given. 250 mls. Do not squish bulbs in the direction of another person. 50 mls.deliver with Blow-Out”. Ie If you want to measure 20 mls.merely approximations. Some are blow-out. They can be unpredictable in force and are close to eye level. Pipette bulbs or pumps must be used and great care must be taken not to contaminate bulbs. Additional supplies are available in the supply room (around the corner). Chemicals will be put out as required for each lab and should be used sparingly. Graduated cylinders: Not as accurate as pipettes. Pipettes are in drawers indicated. 100 mls. some are not meant to drain completely .

Required [1] Required [2] Brief discussion required [2] Required Required [4] 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. Sufficient detail should be provided so that what was done could be understood and the experiment could be repeated from the procedure reported. 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. Required [2] Required [2] Required [5] Required [2] Required [6] Required [1] Required [10] [20] 5 September 2007 CIVL 407 Lab M anual . Total Mark L ONG R EPORT L AB 7 Required Required Required Include a brief description of the procedure.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. 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. 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. The short report is intended to be a concise yet complete laboratory report whereas the long report is more elaborate.

graduated cylinders. Do not wear lab coats outside of lab and do not take away from lab unless stored in a plastic bag.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. samples of raw sewage. if necessary. keep hands and pencils away from your mouth and wash hands frequently. Alternatively a place will be provided in the lab to store your coat. wipe up all spills and wash with disinfectant. settle for 15 minutes longer. gently dislodge any solids that have clung to the sides using a stirring rod. tin dishes. and record the volume of settleable solids. final effluent. Do not include any surface floating material as settled solids. balance. If large pockets of liquid form between the particles of settled matter. oven muffle furnace. distilled water bottles. MATERIALS: -Imhoff cones. 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. Settle for 45 minutes. A. to measure sewage treatment plant efficiency in removing residue. 2. sludge. 6 September 2007 CIVL 407 Lab M anual . evaporating dishes. subtract an estimated volume from the measured volume of matter. Mix the samples thoroughly and fill individually marked Imhoff cones to the 1 litre mark. Use pipette bulbs. desiccators. A 1 L graduated cylinder may be used in place of the Imhoff cone for the sludge sample. SETTLEABLE SOLIDS 1..

The loss in weight represents the volatile solids lost from the originally measured sample volume. SUSPENDED SOLIDS . 6. if instructed.done for you.] Rinse the graduated cylinder with small amounts of distilled water and add to filter.so be quick. the dish will be transferred to a desiccator. Put the dish in the 103-105 °C oven for drying. 3.] Using very small amounts of distilled water. [Hint: pour out small portions (say 25 mls at a time for Effl. Once dishes are at room temperature they can be re-weighed. Wet filter with a small volume of distilled water to seat it. 10 mls for Raw & 5 mls for sludge) of well-mixed sample and filter entire portion before pouring another portion.[Note: Sludge Volume Index is the volume occupied by 1 gram of sludge after 30 minutes of settling. [Notes: to allow for rinsing and to avoid spillage when transferring dish to oven don't pour more than 40 mls]. when filtering slows significantly do not add more.not beakers.]. TOTAL SOLIDS and TOTAL VOLATILE AND FIXED SOLIDS AT 550o C 1. dried and fired). Conversely. C. This is a different test to the one we are doing using Imhoff cones and should not be confused.TOTAL AND VOLATILE 1. the solids remaining represent the fixed solids.[Note: it may be harder to wash out solids if you allow them to settle before pouring into the dish . The samples will be fired at 550° C for one hour. stir the beaker contents and then rapidly (so that it doesn't settle) measure a volume of sample into the appropriate graduated cylinder. Carefully remove filter from filtration apparatus and transfer it back to the aluminum dish. . sample source and sample volume. rapidly pour sample into a 50 ml graduated cylinder to approximately equal 35-40 mls (if using 50 ml dishes). adding the rinsings to the dish. After drying overnight. immediately after stirring the beaker. Obtain the gross weight of the dish [Note: it should be room temperature before weighing. Record the total volume filtered. 2. SVI= settled sludge volume (ml/L) x 1000/suspended solids (mg/L)] B. position the filter and begin suction (vacuum tap). Subtract the tare weight to obtain the total solids weight and calculate the mg/L value. pre-fired and desiccated . pour about 500 mls (for Part B & Part C) into a beaker and then. 4. rinse the cylinder. 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 . Mix the sample in the carboy thoroughly. Assemble filtering apparatus. 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. tare weight. prior to session). Using the sample poured out for Part B (for consistency). The furnace will then be allowed to cool significantly before dishes are removed to a desiccator. Obtain the tare weight of a properly prepared porcelain evaporating dish (ie one that has been washed. The dish containing the dried total solids can now be placed in a muffle furnace or. Do not write on crucibles as high temperatures during the next steps will burn writing off. Quickly record the exact volume and pour the sample into the dish. Suggested portions: 100-200 ml of final effluent. Make sure that you have recorded all the pertinent details: Dish #. dried. Pinch sides of dish in a bit to protect the filter from oven drafts. 2. on a cart in preparation for staff to do so when all are ready. 5. Calculate the mg/L volatile and fixed solids. 3. Obtain the tare weights of three aluminum dishes each containing a glass fibre filter (already pre-washed.

6. If sludge is used in Part A. Calculate as the mg/L total suspended solids. where the volatile solids are reduced from 65% to 40%.) a) a bacterium b) sodium chloride c) sugar d) clay 8 September 2007 CIVL 407 Lab M anual .e. What two techniques can be used to overcome or correct for the loss of material from filter pads when firing at 550 oC? 4. compute the sludge volume index (SVI). if any. Classify each of the following into its proper solids category(s) i. 3. From the above determinations it will now be possible to obtain a value for the dissolved solids present in the sample. cool and weigh. Lab personnel will perform the timed-firing after the class. If all of the volatile solids reduced is given off as gas and if the specific gravity of the volatile solids is 1. A raw sewage sludge goes through an anaerobic digestion process. DO NOT DISCARD! The gain in weight represents the total suspended solids of the sample. 7. Discuss the meaning and significance of this index. What major types of solids are removed in primary treatment? in secondary treatment? 2. overnight).5. % reduction in settleable matter _________ % reduction in total solids _________ % reduction in volatile solids _________ % reduction in fixed solids _________ % reduction in suspended solids _________ % reduction volatile susp. For the time being. solids _________ % reduction fixed suspended solids _________ % reduction dissolved solids _________ Lab 1 Questions 1. 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. They must be allowed to cool completely before re-weighing them to determine the loss on ignition or volatile suspended solids. Enter all data in the tables on following page. 6. volatile. 8. 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).3 and fixed solids 2. Calculate the mg/L total volatile solids and total fixed suspended solids. fixed (use more than one adjective to describe each substance. Transfer dish to a desiccator. After firing the dishes will be moved to a desiccator. suspended. Please return tomorrow to obtain final weight. Your hands may have moisture and natural and/or applied oil/lotion on them. Calculate the percent solids reduction through the treatment plant. what is the percentage reduction in solids. dissolved.50. What. place the dishes on a cart designated by the instructor.

Suspended Solids Tin dish marking (do not use ink .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 .oven dry (G) Dish tare weight (G) Total solids (G) Total solids (mg/L) Gross weight . Settleable Matter After 1 hour (mL/L) B. Total Solids Dish marking (do not use ink .use embossed mark) Sample volume (ml) Gross weight .use permanent mark on dish) Sample volume (mL) Gross weight .fired (G) Loss on ignition (G) Volatile solids (mg/L) Fixed solids (mg/L) Percentage fixed residue C.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.

etc. as well as handling samples probably containing pathogenic bacteria. potassium hydrogen phthalate standards: 800. etc out of your mouth. OBJECT: -to familiarize you with the commonest tests run for assessing treatment plant efficiency and pollution loading to receiving waters. 200. COD . Digests from either method can be titrated or read colourimetrically. pencils.Closed Reflux. homogenization of samples containing suspended solids may be necessary in order to get a representative sample and to improve reproducibility. Keep fingers. 400. 10 September 2007 CIVL 407 Lab M anual . Hach block digester. D O a n d B O D 5 WARNING You will be using concentrated acid and other corrosive and toxic chemicals. otherwise reagents will run to the bulb end and make pipetting difficult and contamination likely. NEVER pour water into acid. pipettes. Materials: Raw sewage and final effluent samples. and to demonstrate the use of dissolved oxygen probes. Wipe up all spills immediately.La b o ra to ry 2: CO D . Always use and store pipettes with the top higher than the tip. I. 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. to illustrate the shortcomings of these tests. Wash hands frequently with soap. 100 and 50 mg/L as COD. spectrophotometer. The closed reflux method is more economical but because a small sample volume (2 mls) is used. vials with pre-measured reagents.

the azide modification of the iodometric method) and a dissolved oxygen probe and meter. manganous sulfate. 200. II. If sample is known to be outside the standard range (above 800 mg/L). a smaller aliquot (making volume up to 2 mls with distilled water) or 2 mls of a pre-diluted sample can be used. which is set at 600 nm to read absorbance of light by the sample. DISSOLVED OXYGEN (DO) Materials: Raw sewage and final effluent samples. [Note: it should look homogenous and be careful it will be very hot!] 3. Remove the vials and allow them to cool to room temperature (use cold water to speed up if necessary). burette stand with burettes. 100. in which case. 50 and 0 COD as mg O 2 /L. The instructor will demonstrate the use of the spectrophotometer. Raw Sewage (Infl) and/or Effluent (Effl) in the upper 1/4 of the tube using permanent markers. a volumetric correction must be made to the value obtained from the curve. 4. aerated dilution water. except without mercuric sulfate). 6. 5. well-mixed samples in the 25-position digester. The four liquids are cold tap water.noting the location of your well-labelled. absorbance versus concentration and calculate the COD in samples. The vials should be clean and dry on the outside. raw sewage and final effluent. 400. alkaline-iodide-azide reagent. 10-ml graduated pipettes. Read your samples and the set of COD standards: 800. BOD bottles.025 N sodium thiosulphate. 0.Procedure: 1. 11 September 2007 CIVL 407 Lab M anual . Take vials to fumehood and place in block digester . before the lab. At your station you will find four vials with pre-measured solutions containing potassium dichromate and sulfuric acid (prepared according to standard methods. starch indicator. The probe has been previously calibrated. into the appropriate tubes and screw cap on snugly. Note: You are going to measure DO on four different liquids using two different methods. Put identifying labels on each tube ie your initials. 500-ml Erlenmeyer flasks. Prepare calibration curve. and are at the spectrophotometer. Pipette 2 mls of sample (using volumetric pipettes unless particulate is present. concentrated sulphuric acid. DO NOT ADJUST THE SETTINGS ON THE METER. 2. 250-ml graduated cylinder. You will use a chemical titration (Winkler . Allow samples to digest for 30 minutes (normally this would be 2 hours). thermometers. dilution water. use wide mouth graduated pipettes) in duplicate. Standards have been digested for you. Make sure sample is well mixed with the acid contents of the vial. If less than two mls of sample or diluted sample was used.

025 N Na 2 S2 O 3 until only a faint yellow or "straw" colour remains. 4. Fill a BOD bottle with each of the four samples (or use ones saved from A ). tip the liquid out of the space around the stopper (caution -it may be corrosive) and invert several times. The chemical floc should completely dissolve.A. For the cold tap water sample. To the same bottles. right to the top and place stopper on bottle. (Fill carefully. Determine DO. Fill four BOD bottles with the four samples (cold tap water. When the floc has settled the second time to about 1/3 bottle volume. 2. Determine the percent saturation for each sample. raw sewage or influent and final effluent). as demonstrated. Tip out the liquid from the area around the stopper and invert several times to mix thoroughly. Note: if solution already seems pale yellow add starch right away. 8. Add enough starch solution to give a strong blue colour (one dropper full) and continue to titrate. Stopper the bottles without trapping any air bubbles. Set back in sink. Use the specially marked volumetric flask to dispense the 201 ml. Instructions on the use of the probe will be given in the lab. 1. taking care not to entrain additional air. 2. add 1 ml of concentrated H 2 SO 4 and quickly restopper. to thoroughly mix contents. until the first complete disappearance of the blue colour. 9. 1. using the DO probe and meter. for each of the four samples. add 1 ml of alkaline-iodide-azide solution the same way. remove the stopper. (Hint: if you measured very low DO level with the meter. WINKLER TITRATION (Azide modification) NOTE: Use only the droppers attached to the reagent bottles for dispensing into samples. (Note: biological solids originally present in the sample will not dissolve.) Transfer 201 ml of each bottle (do one at a time) to a 500 ml conical flask. Allow the floc to settle to about 1/3 the bottle volume. Do not mix droppers and do not allow tap water into the reagent bottles. Titrate this volume with 0. . (Note: DO meter temperature is usually not accurate. 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. Take each stoppered bottle. invert several times again and allow to settle again. 12 September 2007 CIVL 407 Lab M anual 3. then the same sample will require little or no titre and may not turn blue when starch is added . aerated dilution water. submerge the hose in the bottle and allow the water to overflow for 15-30 seconds). 7. again quickly replacing the lid.) You can save these bottles for B. DISSOLVED OXYGEN PROBE NOTE: DO NOT add any reagents to the samples used for the probe method. 5. The DO concentration in mg/L is equal to the number of ml of titre as long as 201 ml is titrated. Neglect any reappearance of the blue colour. Place the bottles in the sink and one at a time remove the stopper.think about it!) Complete all four titrations. 6. Record the volume of titre. dropwise. B. Record the temperature of each sample using the DO meter or a thermometer.

Raw Sewage Final Effluent Tap Water Dil’n Water 13 September 2007 CIVL 407 Lab M anual . Draw a straight line between the water temperature and the mg/l of dissolved oxygen.Dissolved Oxygen Data Sample/Data Bottle # Sample Volume (mls) Initial Buret reading Final Buret reading Net Titre (mls) DO conc. 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. use the saturation chart above. (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. (mg/L) DO conc. Pair up the mg/l of dissolved oxygen you measured and the temperature of the water in degrees C. The percent saturation is the value where the line intercepts the saturation scale.

ie Rinse the filling tube if it has been in the sample. Be sure to record the bottle numbers and sample volumes on the following table . showing sample calculations. 3. Dissolved oxygen meter and probe. Note: Each group should have 9 BOD bottles in the incubator.do not write on the bottles and do not use the numbers on the lids. These caps are designed to prevent the evaporation of the liquid around the stopper. Materials: Raw sewage and final effluent samples. Calculate the 5-day (or 6-day if unavoidable) BOD. being sure to exclude any air bubbles that might be trapped under the stopper by adding dilution water. In addition. Be sure to note this in your lab write-up. 14 September 2007 CIVL 407 Lab M anual . Stopper the BOD bottles. The blank is used to check that the dilution water has negligible BOD. graduated cylinders. Place all bottle sets in the 20 o C incubator for five days (or 6 for Tues. but if there is a significant BOD then it will have to be subtracted before the dilution factor is applied in the calculation. 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). groups). 5. Because you want to be sure that the dilution water has no BOD. If the dilution water BOD is greater than 1 ppm. 10-ml graduated pipettes. it may indicate that probe may not have been calibrated properly or the dilution water had been contaminated. Measure the Initial DO using a calibrated probe and meter. carefully measure the waste water samples into BOD bottles. thereby maintaining a water seal for each bottle. dilution water. Using the volumes and samples provided by the instructor. 4.if not add a some dilution water (or distilled water) to the well. be careful not to contaminate it with sample. 6. BIOCHEMICAL OXYGEN DEMAND Note: Because of the logistics of organization some groups may have 6 day BOD 5 instead of the usual 5 day. fill one BOD bottle with DILUTION WATER alone. After five (or six)days. Place plastic caps over all bottles. All bottles should have liquid in the well around the stopper . Carefully top up the BOD bottles with DILUTION WATER. There should be negligible BOD in the Dilution water. remove bottles and measure the DO.III. preferably using the same meter and probe used to get IDO reading. Measure the Initial DO of the blank using the calibrated probe and meter. 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. to bring the levels above the ground glass neck of the bottle. 1.

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. mg/L = DO of diluted sample after 5 d incubation at 20 oC. = = = = decimal volumetric fraction of sample used DO of seed control before incubation. and ratio of seed water in diluted sample to seed in seed control= (%seed in diluted sample) / (%seed in seed control). 15 September 2007 CIVL 407 Lab M anual . mg/L DO of seed control after incubation. DO of diluted sample immediately after preparation. mg/L.

17. Diln. If an industrial waste has a COD of 450 mg/L. 2) 3) 4) Show on a graph the approximate dissolved oxygen sag curves in a river receiving two wastes. 1 Raw Sewage. One waste has a K value of 0. the other 0. Diln. what would you expect its I) BOD 5 and ii) BOD L to be.) Why is your COD result different from the BOD result? 5.Summary Raw Sewage. 1 Final Effluent. Diln. if at all possible. 6) 7) 8) 16 September 2007 CIVL 407 Lab M anual . Diln. Calculate theoretical oxygen demand for 300 mg/L of methyl alcohol. open reflux. titrimetric method.25. 2 Final Effluent. 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. Both wastes have equal BOD 5 temperature and volume. 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.

Wash hands with soap before leaving. 35oC incubator. 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 . filtering apparatus. lab coats.leave it here or take away in a plastic bag to launder. 35 oC incubator. EC tubes and/or Hach A1 tubes. eye protection. OBJECT: -to demonstrate different methods for the determination of the coliform group of organisms and to acquaint you with bacteriological procedures. T h e re f o re . plates. HACH P/A Broth with MUG. membrane filters.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. LES Endo agar plates. Membrane filter method: dilution tubes. Demonstration only SAMPLE: You will do either a raw sewage sample or a final effluent sample.. Get raw data from another group and include in your report. etc.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. away from your mouth. sterile pipets. not both. Demonstration only. Students will do only sections marked ? MATERIALS: Multiple tube method: dilution tubes. sterile pipettes. Wipe lab benches with disinfectant before and after each lab. Heterotrophic plate count: dilution tubes. wipe up all spills and disinfect the area of the spill. lauryl tryptose broth tubes. sterile dilution water. Quebec colony counter. DO NOT MOUTH PIPETTE. 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 . agar tubes. pencils. Do not wear lab coat out of lab . Keep hands.

EC or A-1 tubes positives: 1. It is used as a faster alternative to the LTB/EC procedure for enumerating faecal coliforms in water. Parallel positive BGLB broth cultures with negative EC broth cultures indicate the presence of nonfaecal coliforms. Consider positive EC broths incubated at the elevated temperature (44. All tubes have been examined for gas production. ? B. Streak plates in a manner to insure presence of some discrete colonies. Hach’s Most Probable Number Method 8368. 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. 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. Another series of tubes are inoculated from these positive lauryl tryptose tubes. (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. as demonstrated. Heterotrophic Plate Count . MUG produces a fluorogenic product when hydrolyzed by an enzyme specific to E. Membrane Filter Technique: Serial dilutions of sample are collected on membrane filters and placed on M-Endo media.COLI in 4 to 24 hours. using a ready-to-use P/A Broth with MUG. 18 September 2007 CIVL 407 Lab M anual . The broth will turn yellow indicating total coliform and fluoresce in the presence of E. Using aseptic technique. Bacterial numbers are calculated from the numbers of positive tubes. Microscope Slides (Completed Phase of Multiple-Tube Fermentation): Single colonies arising from single cells are prepared and stained for microscope examination. 2. Colonies that are pink to dark red with a golden green metallic sheen are counted after incubation 20-22 hrs at 35oC. is USEPA-accepted for testing non-potable waters. A-1 Medium.7oC for 24 hours (confirmed test for faecal coliforms). streak one LES Endo agar plate from each tube of BGLB broth showing gas from the highest dilution and the second highest dilution. D. ? C. Note: these methods will be demonstrated but data must be recorded.Pour Plate Method: Serial dilutions of samples mixed with agar and incubated for 48 hrs at 35oC. Gas-positive Hach A-1 tubes indicate the presence of faecal coliform. 3. Use Table 2 on page 27. 1.coli. Demonstration. 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). Presence/Absence Testing: HACH technique to simultaneously screen for total coliform and E. Express as Total coliforms. Calculate the MPN value from the number of positives using the Table 1.coli. Gas formation indicates the possible presence of coliform organisms.5 oC) as a positive completed test response for both Total and faecal coliform. The number of colonies counted directly. Results will be provided for the demonstration set.A.) Methods: ? Recording BGLB. 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.

choose appropriate dilutions for counting.1. The typical coliform colony has a pink to dark-red colour with a golden green metallic surface sheen. The filtration apparatus has been sterilized by autoclaving. ?B. Pipette about 10 ml of sterile dilution water over the surface of the filter with the suction valve still closed. or colourless and are considered to be non-coliforms. Verification is usually done by a test for lactose fermentation. -Note: “typical” colonies count as coliforms in this procedure. Inoculating. prepare one set of six dilution tubes following the instructions given under Multiple Tube Fermentation. The sheen area may vary in size from a small pinhead to complete coverage of the colony surface. from one edge. 4. Incubate plates (inverted) at 35 oC for 24 ± 2 hr. transfer 1 ml of sample from dilution tube # 6 (most dilute for Raw sewage) or #5 (for Effluent) to an empty (large) Petrie dish. Dilution tubes. then immediately replace the cover. Mount apparatus on suction flask with valve sideways to block suction. coliform bacteria are Gram-negative (pink) rods (sometimes coccus-like).applying gentle pressure. therefore the results table should specify #coliform colonies/100 ml. 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. Colonies that lack sheen may be pink. Membrane Filter Technique -Total Coliform 1. Filtering. 2. Pour the full volume of dilution tube # 6 onto the filter. Using a sterile pipette. Counting.2. ?C. removing the cover just enough to insert pipette tip and dispense sample. Gram staining technique can be also be used. red. Flame loop between second and third quadrants to improve colony isolation. Use dishes with 20 to 80 coliform colonies and not more than 200 of all types. 5. Using sterile forceps. Atypical coliform colonies can be dark red or nucleated without sheen. After 20-22 hr. Close valve and rinse the funnel with three aliquots (portions) of sterile dilution water opening and closing valve after each aliquot. Do not push down on the filter except on the outside edge and use only sterile forceps . open the valve slowly and allow all the liquid to be pulled through. white. In general. Heterotrophic Plate Count . 19 September 2007 CIVL 407 Lab M anual . Be sure dishes are labelled with your name and dilutions. with a motion that excludes air as it comes in contact with the agar. When filtering is complete and the valve is closed. If you haven’t already..Pour Plate Method 1. A high count of non-coliform colonies may interfere with the maximum development of coliforms. The sheen area may vary in size from a small pinhead to complete coverage of the colony surface. Place the filter on a small LES Endo agar plate carefully. 3. Incubate inverted Petrie dishes for 20-22 hr at 35 oC. Repeat for the rest of the dilution tubes to # 2. Dilution tubes. place a sterile membrane filter (grid side up) onto the filtration apparatus. 2. Prepare Gram Stain slides for examination under a microscope. Prepare one set of six dilution tubes as described in Section A. remove filter using flamesterilized forceps (allow forceps to cool or they will damage the filter). Verification of both types of colonies is advisable as some typical sheen colonies may be noncoliform and some atypical may be coliform.

Flood the slide with fresh alcohol-acetone solution to act for 30 seconds.too hot for a baby . 2. on the bench. a. if fewer than 10/cm 2. Test temperature on your wrist . Multiply the number by 5 to compute estimated colonies per plate (66 cm 2 ). d. 4. Wash slide in tap water. Let them cool down from the 60C that they are kept at but not enough to solidify. Presence/Absence Testing . D. Wash off stain with water and then with iodine solution (Lugol’s). Repeat with dilutions 5 to 2 (for Raw) and 4-1 (for Effluent). select the appropriate dilution to count. Slide preparation: With a flame sterilized inoculating loop.coli). Resterilise the loop and remove one colony from the surface of the Les Endo streaked plate (prepared previously). A hand counter is also available. Flood slide with crystal violet solution and allow to act for 30 seconds. ?3. Label the slide using a grease pencil (T or A). Instead. Allow the agar to solidify in the dish before inverting. If necessary. After 48 hr incubation. Spread the colony in the drop of water so that you get a uniform dispersion over an area about the size of a dime. Do this for a typical and an atypical colony. Repeat for the rest of the dilutions. Take 5 tubes of melted agar from the incubator. marking (on edge of dish so as not to impede counting) and incubating for 48 hr at 35 oC. 20 September 2007 CIVL 407 Lab M anual . place 1 loopful of sterile dilution water in the centre of a slide.too hot for a bacterium. average and multiply by 57 (for plastic Petrie dish). Examine for yellow colour (total coliforms) and fluorescence (E. refer to Standard Methods for more instruction. Preferably select seven consecutive squares horizontally and six consecutive squares vertically. to mix the sample with the agar. e. count colonies in 13 squares (of the colony counter) of the most dilute plate having representative colony distribution.3.d e m o n s tra tio n -s ta rte d a h e a d o f la b 1. f. Air dry the slide high above the Bunsen burner and fix by passing slide briefly through a flame. Counting. Microscope Slides: 1. Allow iodine to act for 30 seconds. use the plate(s) that will give from 30 to 300 colonies/plate. Slide the cover of a Petrie dish back just far enough to pour the agar. If more than 10/cm 2. Work quickly or the agar will solidify in the tube before you get a chance to pour it. count four representative squares. Add 100 mls of sample directly to pre-measured medium provided. The aim of using several dilutions is to have at least one dilution giving colony counts between these limits. 5. Incubate at 35 oC for 24-48 hours. Compute bacterial density and report as “colony-forming units” (CFU) per ml. 6. Use the Quebec colony counter to aid in counting and a grease pencil to mark off colonies counted. b. E. Do not report as “too numerous to count” if all plates exceed 300 colonies. Record results of previously prepared test. ?4. pour and replace cover. 2. Drain off excess iodine and wash freely with water and then alcohol-acetone decolourising solution. Ideally. Gently swirl the dish. c. Slide staining.

Come in and count colonies on LES Endo Membrane Filter dishes. . . usually motile. . Wash in water. After application of the counterstain. length to width ratio. blot and then dry in air high above the Bunsen burner (so as not to burn away your efforts). h. Day 3. or in short chains. Cells occur singly. concave). coliform are Gram-negative (pink).Membrane filter prepared dilutions.Prepare dilutions for Heterotrophic Plate Count and Membrane Filter technique.3 µm in size. Examine the prepared slides . transparency. odour. Gram negative organisms are visualized (pink). c. .Prepare and incubate Heterotrophic (Pour) Plates using dilutions above. Day 2.counts will be provided on the board. in pairs. 21 September 2007 CIVL 407 Lab M anual .Examine prepared slides.Record positives in all Multiple Tube Fermentation inoculations: LTB. plump rods. Microscopic examination.Come in and count Heterotrophic Plate colonies after 48 hours. dull. cocci or rods. gram positive or negative. Apply fuchsin or safranin (counter)stain for 30 seconds. BGLB and EC and AI tubes . metallic. pairs chains). smooth. rough. In general. cell groupings (single. a. Note: in the staining process: before decolourisation all organisms appear Gram positive (blue). and on the slide: typical or atypical. b. sometimes coccus-like and 0.g. . . after decolourisation Gram negative organisms are no longer visible.Record Presence/absence testing results of Hach Prepared Media bottles. colour. Description of bacteria: Describe what you see on the plates: colonial morphology of each type of colony seen ie surface (shiny. short. place on LES Endo agar dishes and incubate.5 . . Schedule Summary Day 1. ?3. . Plates ready on a weekend will be put in cold room until Monday.do not turn knobs except the positioning ones. nonsporing.

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 .pour plate method. Multiply the MPN index by the dilution factor from the series used when dilutions other than 1/1. 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 . 1/ 10 and 1/100 are used..

Most Probably Number (MPN) When more than three dilutions are used in a decimal series of dilutions. They are in concentrated form and merely require the addition of 100 ml of sample (or diluted sample) to the media and incubation. A long-wave UV light is required to detect fluorescence. 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.coli.coli. E. Estimation of Bacterial Density . use the results from only three of these in computing the MPN. an enzyme specific to E. MF results. 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. Both media meet USEPA guidelines for testing of total coliforms in drinking water. Use the results at these three volumes in computing the MPN index. 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. The number in the numerator represents positive tubes. MUG (4methylumbelliferyl-$-D-glucuronide) can be used to detect E.01 ml 2/5 2/5 0/5 0.1 ml 5/5 4/5 1/5 0.coli will fluoresce as early as 4-24 hours. the significant dilution results are in boldface. you can simultaneously detect total coliforms and E. To select the three dilutions to be used in determining the MPN index. that in the denominator. If zero total coliform is the maximum contamination goal. the total tubes planted. With MUG reagent added to P/A Broth.coli because it produces a fluorogenic product when hydrolyzed by glucuronidase. it is not necessary to enumerate.Discussion: -sources of error? -compare MPN.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 . In the examples given below.

0. 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 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 .Table 1 .1 mL) Comb.

MPN Table 2 25 September 2007 CIVL 407 Lab M anual .Hach .

htm Lab 5 Questions: 1) Compare the advantages and disadvantages of the multiple tube fermentation and membrane filter techniques.http://www.D.org/microbelibrary/files/ccImages/Articleimages/keen/Gram stainkeen.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.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. 2) 3) 26 September 2007 CIVL 407 Lab M anual .hach.microbelibrary. -The Use of Indicator Organisms to Assess Public Water Safety . What are the important pathogens transmitted by water? How is quality control achieved in microbiological analysis.

standard chloride solution (0. indicator. NaHCO 3. standard Hg(NO 3)2. immediately. standards. beakers. If chemicals are spilled alert instructors. beakers.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. buffer. indicator-acidifier reagent. spectrophotometer. Hint: optimum ranges will coincide with the standards provided or will be indicated within the pertinent section. Use personal protective equipment: gloves. volumetric pipettes. sample. conc. graduated cylinders. semi-log graph paper. 125 ml Erlenmeyer flasks. HNO 3. reference electrode (double junction). Mercuric thiocyanate is very toxic. That means you will probably have to dilute the sample by the most accurate means available. MATERIALS: -millivolt meter. burettes. known addition and calibration techniques and colourimetric and potentiometric titrations. goggles or glasses and lab coats. 27 September 2007 CIVL 407 Lab M anual . Note and consider carefully: Different methods of analysis have different optimum ranges. chloride specific ion electrode. OBJECT: -to acquaint you with the use of specific ion electrodes. to illustrate the differences in operation and accuracy between instrumental methods and wet chemical methods. pH meter with double-junction electrode and silver billet electrode. 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. magnetic stirrer. standard AgNO 3 solution.5 mg/L).

3. place the lowest standard on the stirrer and lower the electrodes into it. where: E1 = E2 = E = Vo = Vs = A = S = potential of sample (mV) potential of sample plus stock addition potential change = E 2 . rinsing and blotting dry the electrodes between each. Unless already done by a previous group. 2.SPECIFIC ION ELECTRODE Part One: Preparation and reading standards and samples 1. stir and obtain a reading for it. Determine the concentration of chloride in the sample. 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 ). 6.in sample = mg/L Cl. As probe tends to drift for some time it may be expedient to select a “standard time to read” (i. labelled beakers.E 1 volume of sample volume of stock solution added mg Cl. Repeat for rest of standards. 5.added to the sample "slope" = change in potential over 10 fold change in concentration of standard (use data from Step 2). added 2 mls of ISA. pour out 100 mls of each standard into separate. Calculate the chloride concentration. mg Cl. 4. plot the millivolt readings (linear axis) against the concentration of the standards (log axis) to produce a calibration curve. Turn on stirrer and while stirring record the millivolt (mv) reading when the reading appears to be stable. CHLORIDE . Unless already done by a previous group (values written on the board).= ____________________ 28 September 2007 CIVL 407 Lab M anual .I. Standards of 1000. Measure 100 mls of sample. 2 minutes) and use it for all standards and samples. Using semi-log paper or software equivalent. 100 and 10 ppm (mg/L) have been prepared for you and are at the meter.e. Add 2 mls of IONIC STRENGTH ADJUSTMENT (ISA) buffer to each using the plastic dropper (filled to the mark).

insert in the spectrometer and record the absorbance displayed on the meter.they can’t all be at 10 minutes at once. it should be rerun from the start after diluting. 4. using the dispenser provided. the 1 ml of the indicator-acidifier reagent will adjust the pH to 2. and Hg ++ (excess) + DPC º violet colour 3. 3.014N Hg(NO 3) 2 to a definite purple end-point. Add 1 ml (use pipette to measure accurately) of indicator-acidifier reagent to sample. 4. Titrate with 0. Using a 25 ml graduated cylinder. (No further adjustment is required after the blank or distilled water has been set to zero.. CHLORIDE . after a few more minutes.BY COLOURIMETRIC METHOD The principle of this method is based on the following formula: 2Cl. use a plastic dropper to transfer blank into a spectrophotometer cuvette (do not fill more than 3/4 ). Pour 100 ml of sample (or a portion diluted such that the concentration is less than 100 mg/L) into a 250 ml beaker. a pH of greater than 3. discard and refill at least twice to rinse cuvette. If sample is above the highest standard.º HgCl2 1.] After 10 minutes has passed (since the combined reagent was added to the blank). Hg ++ + 2Cl. the excess mercuric ions combine with diphenylcarbazone (DPC) to produce a violet colour (the end-point). pure blue. Determine the chloride concentration in the sample from the calibration curve.0 ppm). 2.+ Fe+++ º Fe(SCN) ++ (Reddish brown) 1.0 and 8. zero instrument on distilled water and obtain an absorbance value for the blank.ions have been bound.II. 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). 2.1.) Every two or three minutes thereafter you will rinse the cuvette with the next standard or sample. 5.8. MERCURIC NITRATE METHOD FOR CHLORIDE DETERMINATION The principle of this method is as follows: Cl. III. 6. fill.) Mix thoroughly by gently swirling the flask.+ Hg(SCN)2 º HgCl2 + 2SCN SCN . Near the end-point.0. [Note: this delay between additions is to allow you time to measure absorbance of each at the 10 minute time required . For most samples. Wait two or three minutes and then add the reagent to the first (lowest) standard. (Alternatively.) Prepare a calibration curve by plotting the absorbance readings versus the concentration of chloride in the standards. The colour of the solution should be green-blue at this point. (Light green indicates a pH of less than 2. measure 25 mls of “zero” standard or “blank” (halide-free distilled water). add 5 ml of the “combined reagent” (containing ferric alum and mercuric thiocyanate solutions) to the blank first.ions in the sample combine with added mercuric ions forming a soluble but virtually undissociate-able complex. the three standards (2. which then must be subtracted from the standards and sample. (Use extreme caution as this is a corrosive & toxic chemical. 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 . After all Cl. Marking the time (T=0). and the sample (diluted if necessary) into separate and labelled 125 ml Erlenmeyer flasks (5 in total).5 ± 0. 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.0.

large increments (1-2 mls) may be added.4. The end-point occurs at the greatest mV change per unit addition of AgNO 3. Continue the titration about 5 ml past the end-point. CHLORIDE BY POTENTIOMETRIC TITRATION Note: For this part form two groups comprised of 1 member from each station. 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). smaller increments (0. This way each station will have data for each part. in increments.. 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. Part one: Standard 1. where: A = ml titrant for sample B = ml titrant for blank N = normality of titrant IV.2 ml or drop-wise) should be added. At the start. Nitric acid is in the sink and is very corrosive. Calculate the chloride concentration: 5. Calculate the Normality of the AgNO 3 using the following equation: 30 September 2007 CIVL 407 Lab M anual . as the end-point of the reaction is approached (ª mV/ml increases). Begin titrating.if one is necessary. Take care that stirrer is not hitting the electrodes. Start the stirrer and set the millivolt reading to zero or record initial mv reading [Note: Some meters can not be set to zero]. Plot a differential titration curve to determine the exact end-point. 3. 6. 2. Don’t forget to add the indicator. with standard AgNO 3. and add 2. drops of the purple end-point. place beaker on stir plate and lower electrodes into the solution.0 ml (use volumetric pipette) of standard chloride solution in a 250 ml beaker. Rotate the work within the group so that different pairs are doing the titrating and recording of numbers. dilute to about 100 ml with distilled water. then. until past the endpoint when larger increments can be used again (ª mV/ml decreases).1 to 0.0 ml of concentrated HNO 3 (use plastic dropper). recording the volume and millivolt reading for each increment. Immerse stir bar. 4. 5. A reference set of data will be provided for comparison. The titre from this step must be subtracted from the titre for the sample.

c. 4. 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 . if dilution is necessary into a 250 ml beaker (use a graduated cylinder to measure). b. Measure 100 ml of sample or portion made up to 100 ml. (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. 2. in this case. Calculate the chloride concentration in the sample using the following equation: 3. be careful to use the average value of ml of titrant on the abscissa. Millivolt reading versus ml of titrant. Millivolts per ml per ml versus ml of titrant.) Plot three curves for this titration: a. Millivolts per ml versus ml of titrant. Add 2 ml of HNO 3 and proceed as described in Part One.Part Two: Sample 1. In plotting b and c. It is not. which are merely first and second derivatives.

Chloride .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. 32 September 2007 CIVL 407 Lab M anual .

3 mg/L? Outline the advantages and disadvantages of all the methods used in this lab to measure chloride concentration. SPECIFIC ION ELECTRODE # All halides interfere (the electrode is a halide electrode). and nitrite interfere. thiosulphate. # Mercury must be absent from samples. 33 September 2007 CIVL 407 Lab M anual . in itself. strongly reducing solutions may form a surface layer of silver. 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. 2. KNOWN (STANDARD) ADDITION TECHNIQUE # The known addition techniques will only correct for certain types of interferences. causing electrode malfunction. # Pretreatment of environmental samples usually required. PRETREATMENT TECHNIQUES # There are no "standard" pretreatment techniques. 3. MERCURIC NITRATE TITRATION METHOD # Iodide. and sulfite ions interfere when concentration greater than 10 mg/L. # Sample pH may need adjustment beyond the capacity of indicator-acidifier reagent. known addition cannot correct for any ions that test as chloride ion. # Environmental samples usually require pretreatment. # In practice. requires knowledge of the compounds in the sample which are likely to interfere. fluoride and bromide titrate as chloride. Lab 3 Questions 1. COLOURIMETRIC METHOD # Bromide. fluoride. # Colour in the sample may interfere in the absorbance measurement. ferric. # Colour may obscure or interfere with end-point determination. hydrazine. can you see any advantage in suing the first or second derivative curve? Discuss. cyanide. # Chromate. In the above tests. ferric ion. # Other interferences: ferricyanide. # High levels of ions which form very insoluble salts of silver may deposit a layer of salt on the electrode membrane. This is a complex. timeconsuming process requiring a sound knowledge of chemistry and extensive experience working with environmental samples. specific ion electrodes are rarely usable in environmental samples. From your chloride results. Also. chromate. Each sample type requires development of an appropriate method which.INTERFERENCES IN CHLORIDE DETERMINATIONS POTENTIOMETRIC METHOD # Iodide. # Environmental samples usually require complex pre-treatment. iodide. fluoride and bromide titrate as chloride. It can only correct for substances that enhance or suppress the test response proportional to the amount of chloride ion present. dichromate.

PREPARATION OF STOCK CHLORINE SOLUTION (Iodometric method . pipettes.N-diethyl-p-phenylenediamine (DPD) indicator solution. Make up to the 250 ml and invert flask several times to mix. Materials: -600 ml beakers.025 N sodium thiosulphate solution (use a volumetric pipette). 2.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. I. 3. 1995.025 N sodium thiosulphate. about 50 ml of chlorine-free water (distilled water will do). Rinse a burette and fill it with this stock chlorine solution. potassium iodide (KI) crystals. 250 ml Erlenmeyer flasks. 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.. chlorine demand and break-point chlorination. (concentrated) glacial acetic acid. burettes and stands. clean up all spills with a brush into a container and wash crystals down the drain. chlorine free water. standard ferrous ammonium sulphate (FAS) solution. Bleach/chlorine solutions are strong oxidants. and exactly 10 ml of 0. to illustrate the concepts of free and combined chlorine residuals. gloves and lab coats.total chlorine residual) 1. Take a 250 ml Erlenmeyer flask and add about 1 gram of potassium iodide. 0. add 1 ml of starch solution. OBJECT: -to acquaint you with disinfection of water supplies and to demonstrate techniques for the determination of chlorine residuals. starch indicator. allow to mix again. pages 4-56 to 4-57 or older/newer editions. Titrate rapidly with the stock chlorine solution (in burette) until the appearance of a constant 34 September 2007 CIVL 407 Lab M anual . 250 ml volumetric flasks. REFERENCES: Standard Methods 19th ed. Wipe up all liquid spills immediately. When weighing chemicals at the balance. N. Personal protective equipment required at all times when working in the lab are: eye protection. 100 ml graduated cylinder. about 5 ml of glacial acetic acid (use a graduated cylinder). Mix well (on stir plate with stir bar). bleach or hypochlorite solution. phosphate buffer solution.

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). Let stir for 2 min and then continue titrating until red colour (if any) is discharged (Reading C).) Free and combined chlorine or total chlorine: To obtain total chlorine in one reading. calculate the chlorine concentration in the stock solution (See Data and Results section for formula). pp 4-43. From the amount of chlorine solution used. 18th edition. c) Dichloramine: Add about 1 g KI (to the same sample) and mix to dissolve. After 15 minutes of contact time. 2. 4. These 6 flasks can be prepared at once to make them ready for the next step. b) Monochloramine: Add two drops of KI solution (to the same sample) and mix. Work in groups of three or four for this part. Let stand 2 minutes more. add full amount of KI at the start (ie 2 drops KI solution and 1 g of KI crystals) to a fresh sample.COORDINATE! IIa. (*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. 3.A Monochloramine= B-A Dichloramine= C-B For interferences.Breakpoint Chlorination using Free and Combined Chlorine Residuals Values 1.).purplish-blue colour. let stand for two minutes and titrate until the red colour (if present) is discharged. 35 September 2007 CIVL 407 Lab M anual . It is likely that the highest dose will require dilution. a) Free chlorine: titrate rapidly with standard FAS (Ferrous ammonium sulfate) titrant until the red colour is discharged. refilling burette. Continue titrating until red colour (if any) is discharged once again (Reading B). 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). Record the burette reading (mls) = Reading A. if colour drifts back (indicating an incomplete reaction). let stand 2 min more if colour drifts back indicating incomplete reaction. For dichloramine concentrations greater than 1 mg/L. 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. Go to c. 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. Record the burette reading (Reading C). Total chlorine (Free plus Combined) = Reading C Free residual chlorine = Reading A Combined residual chlorine (dichloramine plus monochloramine) = Reading C . IIb. d) Alternatively: (Not to be done in this lab. Method 4500-Cl F. titrate to colourless again. as there is lots to do . Place 5 ml of phosphate buffer solution and 5 ml of DPD solution in a 250 ml conical (Erlenmeyer) flask and mix. 2. Determination of Free and Combined Chlorine by the DPD Method 1. 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). Add 100 ml of sample or diluted* sample using a 100 ml graduated cylinder and mix again. mix to dissolve. etc. please refer to Standard Methods.

free and combined residual chlorine for each applied chlorine dose. 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. Make a separate graph for the 15 minute and 1 hour contact time. If this is true.e. FAS titrant concentration is 1 ml = 100 µg Cl as Cl2 a. The rate of kill of bacteria by chlorination follows first-order reaction kinetics. what percentage of bacteria would be killed in 10 minutes at a chlorine residual of 0. Why is it important to determine chlorine residuals in domestic water supplies? 2. Given: HOCl W H + + OClK eqv = 2. Determine the concentration of total.DATA AND RESULTS I. Make a plot of the three residual chlorine components against the applied chlorine dosage. II. a. how many coliforms would remain after a chlorine contact time of 10 minutes? 20 minutes? 36 September 2007 CIVL 407 Lab M anual . Under what conditions is the practice of break point chlorination necessary? 4. 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. If a sewage sample contains 10 x 10 6 coliforms/100 ml. c. d. 250) then convert mg/L to g/100 mls = %.7 x 10-8 at 25 o C % HOCl = 27 What is the pH of the solution? 3.5 mg/L if 50% are killed in 1½ minutes at this concentration? b. b. 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.

(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 .15 MINUTE CONTACT TIME BEAKER NUMBER Vol. of Cl Soln.

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. of FAS to endpoint B (mls) (includes A) Vol. (mls) (Stock=_____mg/L Cl as Cl2) Applied Cl Dosage (mg/L) Vol.60 MINUTE CONTACT TIME BEAKER NUMBER Vol. of Cl Soln.

Ac id ity . hardness indicators. metres and calibrated glassware are available in limited quantities and are very expensive to replace. Calibrate the pH meter by following the procedure outlined on the instruction sheet beside the meter unless told otherwise. T u rb id ity WARNING Please use caution when handling all chemicals. 3. Use care when using lab equipment. MATERIALS: -pH meter. 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. pH test paper and other water quality assessment equipment. standard acid. 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. Rinse probe when you are finished and leave immersed in pH 7 buffer for storage. if above 7. standard base. DETERMINATION OF pH 1.) Record value in following table. OBJECT: -to familiarize you with the most common water treatment tests. turn on stirrer and obtain a stable reading. After calibration is complete. buffer solution. magnetic stirrer. colour comparator. Normally. Always wear personal protective equipment.) 2. p H. beakers. then use buffers 4 and 7 to calibrate. 39 September 2007 CIVL 407 Lab M anual . Probes. to acquaint you with the use of pH metres. place a beaker containing the sample and stir-bar on the stir-plate. 4. rinse & dry probe. graduated cylinders. particularly those labelled with specific hazards. metacresol purple. bromcresol green. the two buffers that are chosen for calibration will bracket your sample ie if sample is less than 7.Lab o rato ry 6: Alkalin ity . lower probe. 1 N NaOH. water samples. Co lo u r. I. burettes and stand.(Do not allow the stir bar to hit the probe. EDTA. Hard n e s s . flasks. turbidimeters. then use buffers 7 and 10. bromphenol blue. (Take care not to contaminate paper in case. phenolphthalein. pH test paper.

Rinse and fill a burette with standard EDTA solution and record the initial burette reading.1 N sodium thiosulphate to the sample. 2. titrate until just colourless. Add 1 drop of sodium thiosulphate solution. Pour into a 250 ml Erlenmeyer flask. Titrate slowly with EDTA. it may be necessary to repeat the titration with the sample diluted 1 to 1 (again) with distilled water. A pH much above 10 promotes CaCO 3 precipitate. 2. Add stir-bar.. while stirring. Measure 100 ml of water sample into a clean. Add the last few drops at 3-5 second intervals so that you do not "overshoot" the end-point. (BG or MO) 6. TOTAL HARDNESS DETERMINATION . 3. Apply dilution factor to obtain final result. add 90% of the titrant to the sample before adjusting the pH with buffer. 3. IV. Record the volume of titrant required in the following table. Complete the titration within 5 minutes to minimize the tendency toward CaCO 3 precipitation. until the reddish tinge disappears from the solution and a pure blue remains. Record the burette reading in the following table. add 1 drop of 0.5).EDTA TITRIMETRIC METHOD 1. sample-rinsed graduated cylinder and transfer to an Erlenmeyer flask. Alternatively. 5. III.II. White paper placed under the flask will facilitate this determination. Record burette reading (P). DETERMINATION OF ALKALINITY 1. (If solution is clear after addition of P. DETERMINATION OF ACIDITY 1. 4. Rinse and fill another burette with standard base solution and label it. If a ppt does form within the 5 minutes. Measure out 25 mls of sample using a graduated cylinder and dilute to 50 ml with distilled water. Add a full dropper of bromphenol blue indicator and titrate until the colour changes from yellow to blue ("MO" Acidity). Measure out a new sample and add 1 drop of sodium thiosulphate and a full dropper of phenolphthalein. 40 September 2007 CIVL 407 Lab M anual ... Titrate to the first uniform pink colour (colour stays). 2. If sample is highly alkaline (titration goes over one burette full) dilute the sample and titrate again. 3. 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. Measure 100 ml of sample (with minimum agitation or exposure to air) using a graduated cylinder and transfer carefully to an Erlenmeyer flask. In order to remove any free residual chlorine (which interferes with the indicator colour response). (If the solution is already blue after indicator. The buffer elevates the pH to 10.?) 4. 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. Add a stir-bar. Rinse and fill a labelled burette with standard acid solution... Add a full dropper of phenolphthalein (P) indicator and if the solution stays pink or red. if the approximate hardness is known (by unsuccessful attempts).think what that means!) 4.

Fill a sample cell to the line (about 15 mL). 4. Refill the burette with EDTA titrant and note the initial reading. Measure out 50 ml of sample into an Erlenmeyer flask and add 2. TURBIDITY . 4. Select automatic range by pressing the RANGE key. Pour the supernatant into one of the cells (to the etched mark) and fill another with distilled water. Select signal averaging mode by pressing the SIGNAL AVERAGE key.BY HACH 2100P Turbidity Meter. Demonstration of Jackson Candle and Hellige PART A: Hach Turbidimeter 1. 4. 5. V.. Filter about 20 mls of sample using a filter funnel and stand and the pre-fluted filter paper provided 2. Press: I/O.TRUE AND APPARENT PART A: TRUE COLOUR . to determine the apparent colour (measures the influence of turbidity on the reading).. 3.EDTA TITRIMETRIC METHOD 1. Record all colour data in the following table. with continuous stirring to a pure blue end-point. 2.0 ml of 1 N NaOH solution (use 3 droppers full). Use signal average mode if 41 September 2007 CIVL 407 Lab M anual . Use the technique outlined in Part A. Add one aliquot of the Calcium Hardness Indicator and titrate with EDTA slowly. Place the cell containing sample in it and press Read. Record the final burette reading in the following table. The display will show SIG AVG when the instrument is using signal averaging. 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. CALCIUM HARDNESS DETERMINATION . to mg/L platinum as chloroplatinate) PART B: APPARENT COLOUR 1. Wipe the cell with a soft. Add a stir-bar and stir to mix. The reading might be higher so if you had to dilute in Part A dilute for Part B. The instrument will turn on but will turn off automatically after a short time so you should be ready. 2. COLOUR . except on an unfiltered sample. 3. Values are reported as APHA Colour Units (eqv. The display will show AUTO RNG when the instrument is in automatic range. lint-free cloth to remove water spots and fingerprints. Place the cell containing water into the spectrophotometer and press Zero. .Enter the stored program number for true colour=120 1. The instrument may suggest diluting if over range..HACH Spectrophotometer. taking care to handle the sample cell by the top. 2. Cap the cell. Record the final burette reading in the following table. VII. or a volume sufficient to produce a pH of 13-14 (designed to precipitate the magnesium as a hydroxide). Close the lid. VI.5.

Wipe the outside of the tube to dry it and place it in the turbidimeter (in the circular groove of the mirror unit). Note that there are several different scales on the side of the tube. Carefully fill the 20 mm viewing depth tube to the mark with sample. (This allows you to make a more gradual and accurate approach to the end-point.NTU. 2. Press: READ The display will show . 16th edition. PART B: Hellige Turbidimeter . 3. Pinch all excess charred wick with fingers before lighting or soot may be deposited on the glass tubes. 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. record the number and refer to the appropriate graph to determine the turbidity as mg/L SiO 2. obscuring vision. Lower the plunger into the liquid making sure no bubbles are trapped under the plunger and the top is dry. that the filter selector shows "none" and that bulb B is in use. Be sure that the calibrated tubes are clean before adding samples to the tubes. Take the reading where the dark central part first merges with the surrounding field. Record the turbidity after the lamp symbol turns off. 6.) Remove the glass tube from the holder and read the turbidity at the meniscus of the sample. then the turbidity in NTU..the sample causes a noisy signal (display changes constantly). If you take too long. 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.Demonstration only 1. Pour sample into the tube in small increments until you can no longer distinguish the shape of the candle flame.. the turbidity may start to settle and condense on the bottom of the tube. Read the scale on the dial.T. 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.adding liquid to a hot tube will cause it to shatter.. 42 September 2007 CIVL 407 Lab M anual . See Standard Methods. page 272-273.s.demonstration only. Record value in following table as J.. Note that the rectangular door mirror is closed. DO NOT PLACE AN EMPTY TUBE IN THE TUBE HOLDER WITH THE CANDLE LIT .U. PART C: Jackson Candle Turbidity . 1.. 2. 3. Close the door and switch on the light. Try to be quick and not burn the candles more than a few minutes at a time. T = Total alkalinity.

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.O." Alkalinity end-point (mls) Initial burette reading Net P.Sample volume "M.DATA Sample 1 pH . Alkalinity end-point reading (mls) __________ "M. Acidity end-point reading (mls) Initial burette reading Net "M." Alkalinity titre (mls) ACIDITY.O. Alkalinity titre (mls) Net "M.O.Initial pH by test paper Initial pH by pH meter ALKALINITY.O." 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 .

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 1.00 ml EDTA titrant at the calcium indicator endpoint = 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.

Alkalinity as OH .RESULTS (sample) P." Alkalinity as CaCO 3 (mg/L) OH .APHA Colour Units: True Apparent Turbidity .U. ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ ______________________________________ 45 September 2007 CIVL 407 Lab M anual .O.Hach .Alkalinity as HCO 3 .Alkalinity as CaCO 3 (mg/L) OH .ion (mg/L) "M.O. Alkalinity as CaCO 3 (mg/L) "M." 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 .T.ion (mg/L) CO 3 -2 Alkalinity as CO 3 -2 ion (mg/L) HCO 3.Units= N.Alkalinity as CaCO 3 (mg/L) CO 3 -2 Alkalinity as CaCO 3 (mg/L) HCO 3.

46 September 2007 CIVL 407 Lab M anual . What sanitary significance does colour have? 4.Lab 6 Questions 1. 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.7 x 10 -11 and K w = 10 -14. Does this agree with the results from Part II of this lab? Explain. given K 2 = 4. What errors can occur in the calcium hardness determination? Why? 3. Is this water suitable for a domestic water supply? Why? 2. non-carbonate and total hardness of this water in mg/L as CaCO 3 ? in meq/L? Is the analysis complete? Why? 6. calculate the carbonate alkalinity in mg/L as CaCO 3 for the water sample analyzed during this lab. 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. From equations (17-12) and (17-13) in Sawyer and McCarty.

47 September 2007 CIVL 407 Lab M anual . No. soda ash solution (Na 2CO 3) and lime (Ca(OH)2) or sodium hydroxide (NaOH). In the second lab you will use the previously determined dosages to coagulate and soften the sample and then re-analyse the resultant water. You should have with you the part of Exercise 7 that gives instructions for the various tests. MATERIALS: -Jar Test Apparatus. Several lime and soda ash dosages will also be selected for determining the efficiency of water softening. 7 (except Jackson Candle and Hellige Turbidimeter).6. OBJECT: -to illustrate the principles of coagulation and water softening. 1000 ml beakers. to be conducted over two lab periods. This is a two-step experiment. 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. Please try to coordinate your work with the rest of the class. The optimum alum dosage for coagulation of the sample will then be chosen. and all of the materials from Lab.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. alum solution (Al2 (SO 4 )3 @18H 2 O). No.for the next session (b).

alkalinity. medium. Stop stirrer and allow floc to settle. Pour six 1000 ml aliquots of sample into 1000 ml beakers and place on the jar test stirrer apparatus. if available. Record observations on settling rate. floc volume. decant the supernatant liquid carefully into another set of beakers (try not to stir up what has settled). Make sure it is centred. Lift paddles out of beakers and secure. but because of equipment limitations. drop of 0. 5. 3.1 N sodium thiosulphate 48 September 2007 CIVL 407 Lab M anual . and fill in the table on the board. In the lecture. hardness (total and calcium). 6. moderate. Stop stirrer. turbidity and colour. alkalinity. 1. Discard the floc and clean beakers in preparation for a second decanting in Step 4. Add the designated lime (or NaOH) and soda ash additions as quickly as possible to the decanted samples. small.. Record relative floc sizes (pin-point. Complete the coagulation summary sheet during the lab period. Brief Methods Summary for Analyses Alkalinity: @ 100 ml of sample. 7. Carefully decant into a graduated cylinder 800 mls of the supernatant liquid and pour into fresh beakers. Make relative estimates of floc sizes. if time permits. 2. 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). Analyze the supernates for pH. position them under stirrers. Place beakers on the jar tester apparatus. hazy. clear. decant the supernatant liquid carefully into a clean set of beaker (try not to stir up what has settled) and discard the floc. acidity. use indicators as specified in Lab 6. Observe and make notes describing floc formation and note the time of appearance. clarity of supernatant liquid (very clear. Stir at 100 rpm for 1 minute. 3. A pH meter could be used for the alkalinity and acidity determinations.work in 2 or 3 teams to assess a total of 6 dosages for each team. true colour and turbidity (Hach). Stop stirring. you will discuss the data and select the appropriate alum dosage to be used by all groups in the water softening step next week. acidity. slow). large). After the flocs have settled (allow 15 min).5 to 1 cm off the bottom of the beaker. Add the assigned alum dosages as quickly as possible and then start the stirrer. Stir at 100 rpm for 1 minute. 1. 5. Stir at 100 rpm for 1 minute and at 20 rpm for 10 minutes. hardness (total and calcium). and settling characteristics of the floc (rapid. After the floc has settled (allow 15 min). not a beaker ) and transfer to 1000 ml beakers. 2.Part A:. Reduce the stirring rate to about 20 rpm for 10 minutes. 4. (NB dosage now based on 800 ml sample size). Determine which dosages were most appropriate for this sample. Measure out six 1000 ml aliquots of sample (using graduated cylinder to measure. 6. 4. You may need to use blocks to position the paddle 0. 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. COAGULATION USING ALUM . cloudy). times of appearance and descriptions and record. and clarity of supernatant liquid. Add the appropriate alum dosage and immediately start stirring. Part B: WATER SOFTENING PROCEDURE (to be done the following week) again work as teams of 2 or 3. Analyze these supernates plus a sample of raw water for pH.

Softening 1.02 N base with bromphenol blue to blue . show by calculation the theoretical amount of lime and soda ash required for excess lime/soda ash softening. 2. 3. Compare removal of hardness components under the different doses . @ 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 .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 ("methyl orange" acidity) and with phenolphthalein (using a fresh sample) to pink .pH 8.pH 8. Based on your chemical analysis of the raw water.pH 4. @ calculation .pure blue).5. 4.pH 3.000/mls sample Acidity: @ 100 ml of sample.use graphical presentation. 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.@ titrate with 0. 2. Coagulation 1. What treatment would you recommend to prepare this water for human consumption? Why? 49 September 2007 CIVL 407 Lab M anual .1 N sodium thiosulphate @ titrate with 0. Compare your experimental softening results to the theoretical results based on solubility equations or mass balance equations.3 and with bromcresol green (continuing with the same sample) to yellow . drop of 0. @ calculation: Ca hardness (mg/L CaCO 3 )= mls titre x 1000/mls sample Lab 7 Questions A. Analyze the response of colour and turbidity removal as a function of alum dose.alkalinity (mg/L CaCO 3)= mls titre x N acid x 50.02 N acid with phenolphthalein to colourless .3 ("phenolphthalein" or total acidity) @ calculation .acidity (mg/L CaCO 3)= mls titre x N base x 50.pure blue).

5 “P” Net titre to pH 4.5 “TOT” Total Hardness sample vol Net titre Ca Hardness .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.sample vol ml Net titre to pH 3.7 “MO” Net titre to pH 8.5 “TOT” Acidity .sample vol Net titre Notes: 50 September 2007 CIVL 407 Lab M anual .

A.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.U.H.P.) (filtered) Turbidity (N.P.) unfiltered Notes: 51 September 2007 CIVL 407 Lab M anual .A.H.T.) True Colour (A.

5 Acidity .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 8.7 Net (Total) titre pH 8.5 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 .sample vol ml Net titre to pH 3.

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.P.) Note:* Total Alkalinity must include Phenolphthalein Alkalinity just as Total Acidity must include “MO” Acidity.) (Filtered) Turbidity (N.P.T. Calculation 53 September 2007 CIVL 407 Lab M anual ** By .H.) True Colour (A.A. -mg/L CaCO 3 Apparent Colour (A.A. -mg/L CaCO 3 **Non-Carb Hard -mg/L CaCO 3 **Excess Alk.H.Coagulation and Softening Summary Sheet Data Lime/NaOH mg/L Soda Ash mg/L Alum mg/L pH P.U.

.Chlorine Demand . . . . . . . . . . . Chemical Requirements for Water Softening . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 96 21. Alkalinity. . Acidity.Levels and Water Use . . . . . . Normal Solutions and Equivalent Weights . . . 102 23. . . . . . 117 29. . . . . . . . . . . . . . . . . . . . . . . .pdf 1. . . . . . . . Periodic table . . . . . . . . . . . . . . Alkalinity Species as a Function of pH . . . . . . . . . . . . . . . . . . . . . . Residues . DO. . . . . Solids Removal in Wastewater Treatment . . . . . . 72 11. . Extra tables for Chlorine Lab . 107 26. . . . . . . . . . . . . . . . . . . . . . . . . Colour. 64 5. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60 4. . . . . . . . . . . . . Milliequivalent and mg/l as CaCO 3 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Equilibrium Relationships of Chlorine . . . . . . . . . . . . . . . . . . . Known Addition Technique . . . . Water Softening . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 110 27. . . . . . . . . . . . . . . . Turbidity. . . . . . . . . . . . . . . . . . . . . . . . . . . Typical Problems for Acid-Base and General Chemistry . . 118 54 September 2007 CIVL 407 Lab M anual . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 92 19. . . . . . . . . . . . Introduction . . . . Bacteriological Examination . . . . . . . . . . . . . . . . . . . . . . . . . . . 104 24. . . . Standard Solutions. . . . .COD . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69 8. . . 91 18. . . . . . . . . . . pH . Reading and Reference Material . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73 12. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 94 20. . . BOD. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 82 15. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Hach Absorption Method . . . . COD . . . . . . . . . . 67 6. . 70 9. . . . . . . . . . . . . . 112 28. . . . . . . . . . . . . . . . . Chloride Analytical Methods . . . .4 Appendices see: CIVL407Manual_CourseNotes. . . . . . . . . . Bar Diagram Method for Water Softening Problems . . . . . . . . . . . . . 100 22. . . . . . . . . . . . . . . . . . . . . . . . . Chlorine . Titrations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Faecal Coliforms . . . . . . . 57 3. 105 25. . . . . . . . . . . . . 71 10. . . . . . . . 83 16. . . . . . . . . . . 84 17. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Balancing an Oxidation Reduction Equation . . . . Hardness Complexes . . . . . . . . . . . . . 55 2. . . . . . . . . . . . . Concepts and Definitions . . . . . . . . . . . . . . . . . . . . . . . . Coagulation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Bacteriology and Water-related diseases . . . . . . . . . . . . . . . . . . . . . . . Hardness. . Instructions for Laboratory Report Writing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80 14. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 74 13. . . . . . . . . . . . . 68 7.Additional Notes . . . . . . . . . . . . . . . . .

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