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)
Slide 2
Table of Contents Information About Chlorine Materials Used
Information About Bacteria Variables Methodology Data Collection
and Comparison of Results Conclusion
Slide 3
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 chlorines effectiveness in killing
bacteria.
Slide 4
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.
Slide 5
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.
Slide 6
HCI Pool Information InformationSize (metres)Chlorine Water
Added (mass) Value50x25x1.4Approximately 1kg
Slide 7
HYPOTHESIS Chlorine water is ineffective in killing common
harmful bacteria.
Slide 8
Slide 9
Materials Bacteria Equipment E.coli Agar Plates Sterile Swabs
Microscope Chlorine Water (Collected from HCI swimming pool)
Pipette Centrifuge Tubes Nutrient Agar Nutrient Broth Measuring
Cylinder
Slide 10
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
Slide 11
VARIABLES ControlledDependent I ndependent Variable Volume of
Chlorine water used Colonies of bacteria left after the experiment
Type of bacterium used Amount of bacteria used The time in which
the chlorine water is collected Area of collection of water Volume
of Silver Nitrate Used
Slide 12
Slide 13
Content Water Collection Broth Preparation Agar Preparation
Agar Spreading Overnight Culture Preparation Observations
Methodology
Slide 14
Slide 15
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 labs refrigerator.
Slide 16
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.
Slide 17
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.
Slide 18
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.
Slide 19
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.
Slide 20
Slide 21
Methodology (Agar Spreading) 1. Using the recently autoclaved
agar, we displaced the agar onto Petri dishes to a height of about
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.
Slide 22
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.
Slide 23
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.
Slide 24
Slide 25
Methodology (Dilution) 1. Using the overnight culture, we
started with dilution. Firstly, our aim was to reach 1 x 10- and 1
x 10- of bacterial dilution. Using 6 centrifuge tubes, we filled
each centrifuge tube with 9ml of autoclaved nutrient broth using a
pipette gun and pipette. 2. Using a micropipette, we displaced 1ml
of the bacterial broth from the overnight culture into a centrifuge
tube labeled 1 x 10-.
Slide 26
Methodology (Dilution) 3. By repeating step 2, we displaced 1ml
of broth from the centrifuge tube containing 1 x 10- into a
centrifuge tube labeled 1 x 10-, and continued this procedure with
the tubes labeled 1 x 10-, 1 x 10- , 1 x 10- and 1 x 10- . 4. After
this method of dilution had been carried out in the biosafety
cabinet with proper flaming, we used the tubes with the smallest
and second smallest concentration of bacteria for use in the
bacterial spreading.
Slide 27
Methodology (Bacterial Spreading) 1. Firstly, our agar plates
were labeled as accordingly to the type of water that they will be
filled with, the bacteria and the dilution used. 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 with 1 x 10- concentration:7.50am,E.coli, 1 x 10-
,1) (E.g.: Second Control with 1 x 10- : E.coli, Sterile Water, 1 x
10- ,2)
Slide 28
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.
Slide 29
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.
Slide 30
Slide 31
7.50am Water Water SampleNumber of colonies 7.50am, E.coli, 1
7.50am, E.coli, 2 7.50am, E.coli, 3 Control
Slide 32
2.30pm Water Water SampleNumber of colonies 2.30pm, E.coli, 1
2.30pm, E.coli, 2 2.30pm, E.coli, 3 Control
Slide 33
5.00pm Water Water SampleNumber of colonies 5.00pm, E.coli, 1
5.00pm, E.coli, 2 5.00pm, E.coli, 3 Control
Slide 34
Slide 35
Silver Nanoparticles These are particles of mainly silver oxide
ranging from 1 to 100 nanometres in length. They are scientifically
proven to possess strong antibacterial properties.
Slide 36
Problems 1/9 of people are allergic to the nickel present in
silver nanoparticles. Scientists have conducted a test in which the
silver nanoparticles have actually caused severe developmental
defects in zebrafish. Overexposure might cause degeneration of body
parts due to the strong antibacterial properties.
Slide 37
Slide 38
Slide 39
Slide 40
Timeline MonthsFebMarchAprilMayJune Events:PlanningMentorship
Approved PrelimsCollection Of Data Conduction Of 1 st Experiment
Project Confirmation First Project Meeting First SRC Meeting
PlanningConduction of 2 nd Experiment Group Formation PowerPoint
Prelim Creation Final Experiment Conduction
Slide 41
Timeline MonthJulyAugustSeptemberOctober EventFinalisation of
Project Conduction of Silver Nanoparticles Experiments Full
Completion of Project Grand Finals Semi-FinalsRepetition of
S.Particles Experiments Finals Silver Nanoparticles Planning
Bibliography(2) http://biofilmbook.hypertextbookshop.com
Usborne Encyclopedia Of Science Usborne Mini-Encyclopedia: All
About Chlorine Usborne Information- All About Bacteria
http://www.charpan.com/antibacterial-silver- nanoparticles
http://www.charpan.com/antibacterial-silver- nanoparticles
http://www.nanowerk.com/spotlight/spotid=5966.ph p
http://www.nanowerk.com/spotlight/spotid=5966.ph p