Determination of Phenol Coefficient of Disinfectants

There are many liquid chemicals that are used for cleaning of materials to lower the number of microorganisms. Such chemicals are called disinfectants e.g. lysol, phenol, hypochlorite (bleach), etc. However, the effectiveness of different disinfectants varies against a given microorganism. Therefore, phenol coefficient of these disinfectants by using a standard bacterial culture (e.g. Staphylococcus aureus) is determined. Hence, effectiveness of phenolics (phenol and its derivatives) is determined by comparing with phenol. The phenolic compounds kill bacteria by inactivating plasma membrane and enzymes, and denaturing the protein. The phenol coefficient is determined only of such chemicals which are bactericidal (but not bacteriostatic) in nature

Requirements

• Phenol dilution (1:10, 1:20, 1:30........1:90, 1:100) ,  
• Phenol derivative's dilution (1:100, 1:150, 1:200, 1:400, 1:450, 1:500) ,   
• Sterile nutrient broth tubes (20),
• Broth culture of Staphylococcus aureus, 
• Sterile pipette (1ml),   
• Inoculation loop, 
• Bunsen burner, 
• Test tube stand,

Procedure

1. Take the test tube stands each for phenol and phenolic compound to be tested.
2. Place test tubes of each concentration of each phenol and phenolic compound separately in the test tube stand.
3. Similarly, take equal number of sterile nutrient broth tubes (in other test tube stand) for each dilution of phenol and phenolic compounds. Label them according to dilution of disinfectant(s). Take one sterile nutrient broth tube as control.
4. After 5, 10 and 15 minutes of intervals, transfer aseptically one loopful bacterial culture from each nutrient broth tube appropriately labelled with the same dilution.
5. Similarly, also transfer one loopful fresh bacterial culture in control broth tube.
6. Incubate all the broth tubes inoculated with Staphylococcus aureus at 37°C for 48 hours. 

Phenol coefficient of the test chemicals = Reciprocal of the test chemical dilution marked/Reciprocal of the phenol dilution marked

                                  Suppose, the test chemical shows no growth at 10 minutes in 1:400 dilution, but growth at 5 minutes, and phenol shows no growth at 10 minutes in 1:80 dilution but no growth at 5 minutes, the phenol coefficient of test chemicals will be as below:

Phenol coefficient of test chemical= 1/400 divided by1/80 = 5,

Results

Observe the growth of Staphylococcus aureus in all the broth tubes treated with phenolic com- pounds and arrange the result as given in Table. Put mark on the highest dilution of phenol and the phenolic compounds tested that killed the bacteria in 10 minutes but not in 5 minutes and calcu- late the phenol coefficient as given below:

Disinfectant chemicals	DILUTION	Exposure time (min)
		5	10	15
Phenol	1:10-1:60	-	-	-
	1:70	-	-	-
	1:80	+	-	-
	1:90	+	+	-
	1:100	+	+	-
Phenolic derivative	1:100-1:350	-	-	-
	1:400	+	-	-
	1:450	+	+	-
	1:500	+	+	-

Table:- Effect of phenol and phenol derivatives on Staphylococcus aureus treated for different time intervals.