Studies on Oxidation-Reduction in Milk: The Methylene Blue Reduction Test

Abstract

Summary The mechanism of reduction is discussed. The work reported in this paper tends to confirm the theory of Barthel that the disappearance of methylene blue in milk takes place in two stages, viz.: 1.The removal of the dissolved oxygen by bacteria. 2.The reduction of the dye by constituents of the milk. The time taken for the first stage may be long. The time taken for the second stage is usually short. The oxygen relationship is of great importance. We were unable to find that a preliminary shaking of the milk materially affects reduction times. Evidence is presented that milks produced normally are in approximate oxygen equilibrium with the atmosphere. In milks to which this test is applicable, the variations in pH are insufficient to cause a measurable difference in reduction times. The importance of the role played by the leucocytes is not known. The plate counts of 38 samples of milk taken at the moment of reduction varied from 3.5million to 45million with an average of 22million. It is probable that the two most important sources of inaccuracy are :- 1.The different rates of oxygen uptake of different species of bacteria. 2.The sweeping of bacteria out of the body of the milk by the rising butterfat during the operation of the test. It is probable that the inaccuracies in this test follow the growth curve of the bacteria. Therefore, we do not consider the test reasonably accurate after the 5½-hour period as laid down in Standard Methods of Milk Analysis. Since the methylene blue is, indirectly, an indicator for oxygen, different concentrations of the dye, within limits defined in a previous paper, give identical reduction times. A 10-cc. dipper is, therefore, sufficiently accurate for use in measuring samples for this test. It is sufficient that this dipper be thoroughly rinsed in the incoming milk or in water between each sample, if times are not to be read after the 5½-hour period. The use of any water fit for drinking purposes is warranted as the solvent for the dye. The methylene blue reduction test is as accurate a measure of the keeping quality of milk as any method yet available. It will divide milks into three or four classes with reasonable accuracy, as will any of a number of tests for milk quality. It is inexpensive and as nearly fool-proof as any method for this purpose available to the dairy bacteriologist. Methylene blue reduction times should not be reported in terms of the plate count.