Requirements: test tubes, cotton plugs, 10 ml pipette, 1ml pipette, distilled water.
Testing sample: provided sample of the milk.
1. Methylene blue solution prepared by dissolving 1 mg of methylene blue in 25 ml of distilled water.
2. Three test tubes were marked as A, B, C and 10 ml of raw milk sample was taken in test tube A, B, C respectively.
3. 10 ml of methykene blue solution was added to each of samples and was mixed well.
4. The tubes were incubated in hot water bath at 37.C for 3-6 hours.
5. The test tubes were observed for every 30 minutes for the colour change of the sample from blue to the white colour.
6. The time required for decolouration was recorded
|Sample||Time of decolouration|
The milk sample can be classified as per their decolouration time.
|Sl. No.||MBRT time||Classification of quality||Approx no. Of bacteria per ml of sample|
|1||1-30 minutes||Very poor quality||>2 x 1027|
|2||31-120 minutes||Poor quality||>4 x 106|
|3||121-60 minutes||Fair quality||>5 x 102|
|4||>360 minutes||Good quality||<5 x 102|
Theory: milk is balanced diet and it contains carbohydrate, fat, minerals, vitamins and proteins. Due to its high nutrient value the milk is often susceptible for the microbial growth due to contamination. Contaminated milk sample when consumed cam spread disease like diphtheria, typhoid etc......
It is therefore essential to determine the microbial quality of milk before processing for consumption.
Rational behind technique:
The bacteria present in the milk utilise the O2 and produce a reduced environment. Methylene blue is a colour sensitive chemical which is blue in oxidised start and colourless in reduced state. The speed of colour disappearance of MBRT is proportional to the number of bacterial present and taken as an indication of bacterial load in the milk.
Methylene blue-----reduction----------> methylene blue
(Oxidised state) (Reduced state)