The Effects Of Chlorine On Harmful Bacteria By: Soh Yee Kiat (1P1), Justin Soh (1P1), Leow Shawn Tao (1A4), Sow Jeng Wei (1O2). Group ID: 1-010 A Category 1 Experimental Project (IS)
Table of Contents • Information About Chlorine • Materials Used • Information About Bacteria • Variables • Methodology • Data Collection and Comparison of Results • Conclusion
Rationale Since 1920, chlorine water has been commonly used to disinfect water supplies and swimming pools. During this period of 92 years, these bacteria might have adapted to chlorine, thus scientists have recently doubted chlorine’s effectiveness in killing bacteria.
What is CHLORINE? • Chlorine is a rather abundant halogen with an atomic number of 17 and a symbol of Cl. • It was first discovered by the Swedish scientist Carl Wilhelm Scheele.
Uses and effects of CHLORINE • It can be deadly even at an amount of 430 parts per million(ppm) and exposure of 0.2ppm already affects the human body. • Overexposure to chlorine can cause temporary inflammation of the eyes and hyper-excretion of mucus. • Chlorine water is commonly used as a disinfectant in swimming pools.
HYPOTHESIS • Chlorine water is ineffective in killing common harmful bacteria.
Materials Bacteria Equipment • E.coli • 20 Agar Plates • Sterile Swabs • Microscope • Chlorine Water (Collected from HCI swimming pool) • Pipette
What about E.coli? • It is a type of bacterium that is commonly found in the lower intestines of warm blooded organisms and is common cause of food poisoning in humans. • It is generally transmitted through consumption of contaminated food. E.coli
Content • Water Collection • Broth Preparation • Agar Preparation • Agar Spreading • Overnight Culture Preparation • Observations Methodology
Methodology (Water Collection) • 1. A measuring cylinder was used to collect 30ml of water at the timings 7.50am, 2.30pm and 5.00pm. • 2. Water collection took place on the day before the bacteria were spread. The water was then placed in the lab’s refrigerator.
Methodology (Broth) • 1. Using a weighing machine (to 2 decimal places), we measured precisely 3.25 grams of nutrient broth and placed it in a 250ml sterilised bottle. • 2. Using a measuring cylinder of maximum volume 500ml, we measured 250ml of de-ionized water and carefully poured the distilled water into the bottle before thoroughly and carefully stirring the mixture.
Methodology (Broth) • 3. After a clear, pale yellow solution was formed, the broth was placed in the lab refrigerator. This methodology was carried out a second time to prepare another 250ml of nutrient broth.
Methodology (Agar) • 1. Using a weighing machine (to 2 decimal places), we measured precisely 14 grams of nutrient agar before placing the agar inside a 500ml bottle. • 2. Secondly, using a 1 litre measuring cylinder, we collected 500ml of de-ionized water and poured the water into the bottle before thoroughly mixing the mixture until a cloudy suspension was formed. This agar was then placed in the fridge for autoclaving the next day. • 3. This procedure was repeated once for another 500ml bottle of agar.
Methodology (Bacteria) • 1. To prepare for the overnight culture, we borrowed a plate of E.coli from the SRC and using a bacteria streaking rod, we displaced some bacteria onto a spare agar dish of our own. • 2. This procedure was carried out in the bio-safety cabinet to prevent contamination.
Methodology (Agar Spreading) • 1. Using the recently autoclaved agar, we displaced the agar onto Petri dishes to a height of about ½ the agar plates. It was then left to solidify for around 15 minutes. • 2. Then, water vapour found on the agar plates were wiped off with sterile wipes to prevent the water from interrupting with our experiment. All these steps were carried out in the biosafety cabinet.
Methodology (Overnight Culture) • 1. Using autoclaved nutrient broth prepared on the day before, we poured 10ml of nutrient broth into 4 centrifuge tubes. • 2. Then, using a bacteria streaking rod, bacteria that was prepared a day earlier was scraped and placed into the broth in the centrifuge tubes.
Methodology (Overnight Culture) • 3. To minimise the risk of contamination, we flamed the streaking rod, the broth bottle and the centrifuge tubes both before and after usage. • 4. The centrifuge tubes were placed in the shaking incubator for 1 day in preparation for dilution and spreading.
Methodology (Bacterial Spreading) • 1. Firstly, our agar plates were labeled as accordingly to the type of water that they will be filled with, and the bacteria. Each type of water has 3 prototypes for triplication. Also, 3 agar plates with only bacteria and sterile water was set up. • (E.g.: First plate of7.50am water with E.coli: 7.50am,E.coli,1) • (E.g.: Second Control: E.coli, Sterile Water,2)
Methodology (Bacterial Spreading) • 2. Next, according to the labeling on our agar plates, we carried out our experiment in the bio-safety cabinets. Using micropipettes of the range 0.5-10 microlitres, we pipetted 10 microlitres of bacteria together with 10 microlitres of chlorine water of their respective timings. The controls were done last, with sterile water instead of chlorine water.
Methodology (Bacterial Spreading) • 3. Flaming was carried out after each use of the centrifuge tubes and we used a sterilised and flames bacteria spreader to spread the bacteria after each filling. • 4. To let the bacteria set in appropriately, Parafilm was used to securely tape up the agar dishes and prevent contamination. The bacteria was then left in an incubator at 36.00 Celsius for 1 day.
Bibliography • http://www.medicalnewstoday.com/articles/68511.php • http://natsci.edgewood.edu/wingra/wingra_bacteria.htm • http://www.cbc.ca/news/health/story/2009/07/02/f-ecoli-recall-food-safety.html • http://www.sciencebuddies.org/science-fair-projects/project_ideas/MicroBio_p013.shtml • http://proquest.umi.com.libproxy.nlb.gov.sg/pqdweb?index=9&did=2037658811&SrchMode=1&sid=2&Fmt=6&VInst=PROD&VType=PQD&RQT=309&VName=PQD&TS=1332762445&clientId=13402
Bibliography(2) • http://biofilmbook.hypertextbookshop.com • Usborne Encyclopedia Of Science • Usborne Mini-Encyclopedia: All About Chlorine • Usborne Information- All About Bacteria