Cells of Streptococcus lactis subsp. diacetylactis DRC3 which had been grown on lactose in the presence of citrate were unable to form acetoin and diacetyl (AD) from citrate when suspended in succinate buffer, pH 4.4. Inclusion of both a source of energy and nitrogen in the buffer was necessary for AD formation. Concentrations of AD and D at pH 4.4 were about five and three times the concentrations at pH 6.4. The amounts of AD and D from citrate were about eight and two times those from pyruvate, calculated on a molar basis. It appears that D, at least, is formed not only from pyruvate arising during citrate cleavage, but also from acetyl-CoA resulting from a probable citrate breakdown in a reversible reaction of a citrate synthetase. The rate of AD formation, under optimum conditions, was 0.047 μmoles mg (dry wt−1) cells ml−1 min−1. Pyruvate-grown cells produced little AD from pyruvate. AD production was inhibited partly by acetate and completely by acetaldehyde. Cetylpyridinium chloride at a concentration higher than μg ml−1 suppressed AD production from citrate because of the absence of interfering compounds normally present in milk.

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