The fermentability or non-fermentability of a given sugar is a question which must be constantly decided in the classification of bacteria. Because of the number of sugars and often the larger number of cultures involved, the bacteriologist usually resorts to indirect methods to determine his decision. Turbidity of the media, gas or acid production are the usual criteria taken to indicate fermentation. Everyone realizes that these indirect methods are uncertain and often misleading. Some bacteria produce little or no cloudiness in the media, large numbers produce such small quantities of gas that it cannot be detected except by very refined methods, and others convert most of the sugar into neutral products, such as ethyl alcohol, mannitol, acetone, etc. (Stiles, Peterson and Fred, 1925; Reilly, Hickenbottom, Henley and Thaysen, 1920). In many instances fermentations are best made with an excess of CaCO8 and in such cases gas and acid production are excluded as a means of determining fermentability. In a recent study of anaerobic bacteria Brown (1925) emphasizes the difficulty of determining the fermentability of sugars by indirect methods. A direct method of measuring the sugar destroyed is much to be preferred. It gives a definite and accurate measure of the fermentability of the sugar. Such a method to be widely used must be quick and fairly accurate. The gravimetric methods
Lactic acid has been made from such materials as corn, potatoes, molasses, sugar beets, glucose, and whey by fermentation with Lactobacillus casei, Lactobacillus bulgaricus, Streptococcus lactis, Laotobacillus detbriickii, or other organisms (See references at end of paper). The manufacturer's choice of raw material depends upon his location, equipment, the price of the carbohydrate, and the quality of the product desired. Choice of culture depends upon the raw material and the conditions of fermentation selected.Of the widely available, directly fermentable raw materials, molasses is cheap, and L. delbrickii is perhaps the most useful culture for the transformation of its sugar to lactic acid. We have chosen this combination and determined the conditions producing the largest yield in the shortest time.
EXPERIMENTAL PROCEDUREThe cultures, obtained from distiller's malt, were maintained on a 10 per cent malt mash, with transfers made every three or four days. Titratable acidity of this seed mash should be the equivalent of at least 15 cc. of 0.1 N acid per 10 cc. Crude cultures appeared to give as good results as pure cultures.The inoculant for the final mashes was grown for 24 hours in molasses mash containing 5 per cent sugar, with half the usual CaCO3 and a slight excess of nutrients over those of the final mashes. The latter were seeded with 3 per cent of the inoculant.Eight liters of final mash was fermented in a 9.5 liter Pyrex 149 H. R. STILES AND L. M. PRUESS bottle equipped with a mechanical agitator, and also with a tube for moderate aeration of the fermenting mash. The theoretical amount of calcium carbonate for the complete neutralization of a 95 per cent yield of lactic acid was added at the time the mash was made up. AUl mashes were-steamed for 90 minutes.It was found that an incubation temperature of 5052oC. was as high as could be used for good yields. There was no apparent trouble with infection in this range.
ANALYTICAL METHODSAcids were determined by acidifying 300 cc. of the fermented mash with 10 cc. of concentrated sulfuric acid. After the sample had cooled and the calcium sulfate had settled, a 5-cc. clear, top sample was taken and continuously extracted with ethyl ether for 5 to 7 hours. This extraction removed both the lactic and volatile acids, which were titrated with 0.
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