Environmental Engineering – 20-CE-92

Experiment No. 1
TO DETERMINE THE TOTAL COLIFORM & FECAL COLIFORM BY
MEMBRANE FILTRATION TECHNIQUE METHOD
Theory

 Coliform

Coliforms are a group of bacteria that are commonly found in the environment, including in soil,
water, and on plants. The presence of coliform bacteria in water or food can indicate that it may be
contaminated with other harmful bacteria, such as E. coli.

 Fecal Coliform

Fecal coliforms are a specific subgroup of coliform bacteria that are found in the feces of warm-
blooded animals, including humans. The presence of fecal coliforms in water indicates that it may be
contaminated with human or animal fecal matter, which can contain bacteria, viruses, and parasites.

 Membrane Filtration Method

Membrane filtration is a laboratory technique used for the detection and enumeration of bacteria in
liquid samples by passing a known volume of the sample through membrane filter with a defined
pore size, which retains the bacteria on the filter surface.

Apparatus

1. Autoclave
2. Glassware
3. Incubator
4. Petri Dish
5. Membrane Filter Paper
6. Absorbent Pad
7. Filtration Assembly
8. Forceps
Chemicals

1. M.Coli Blue 24 Broth (For E.Coli)

2. Membrane Fecal Coliform - MFC (Agar Broth – For Fecal Coliform)
Procedure

1. First, sterilize the glassware using an autoclave and collect 100 ml of the sample in a wash bottle.
2. Use sterilized forceps to place a sterile absorbent pad in a sterile petri dish.

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 Environmental Engineering – 20-CE-92

3. Filter 100 ml of the provided control and sample water through a filter.
4. Place the filter on a filtration apparatus, pour the water inside, and start the vacuum pump.
5. Carefully remove the filter with sterilized forceps and place it in the center of the petri dish.
6. Apply 2 ml of both MFC agar broth and M coli broth to the filter paper to allow bacterial growth.
7. Incubate the petri dish at 35°C for 24 hours to ensure the growth of bacteria.
8. After 24 hours, take out the petri dish from the incubator and count the colonies of bacteria
formed. Use a formula to analyze the results.

Minor Pink Spot, observed under microscope.

Calculation

CFU per 100 ml = (No of colonies counted / sample volume filtered in mL) x 100

Colonies formed =1
Volume of sample filtered = 100 ml CFU
per 100 ml = (1/ 100) x 100

= 1 CFU per 100ml

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