266 
The experiment Gonducted under the direction of Doctor Penning- 
ton follows: 
While the temperature of 0° C. is ordinarily accepted as that at which bac- 
terial life is either quiescent or annihilated, the making of ice cream in a freez- 
ing mixture of ice and salt reduces the substances to a temperature of from 
— 10° C. to — 20° C. To test the action of this temperature on the very rich 
bacterial flora ordinarily occuring in commercial ice creams, samples were 
obtained from various sources and maintained for several days at a tem- 
perature varying at from — 10° C. to — 20° C. It has been found by inquiry and 
observation that ice cream may be kept by a manufactory or more likely by 
the retail dealer for a week or ten days. It is of course kept for greater 
lengths of time when provisioning ocean liners or stored for some particular 
purpose, but this is rather the exception. It was deemed advisable, therefore, 
to limit this preliminary investigation to the period which is ordinarily that 
of commerce. 
The samples of ice cream tested were purchased in open market or sent 
directly from the manufacturer, who had no knowledge of the purpose for 
which they were intended. When coming from the manufacturer they were 
packed in the usual tin ice-cream storage can, set in an ice and salt mixture. 
When purchased from restaurants, confectioners, etc., the sample was obtained 
in a sterile wide-mouthed glass jar, tightly capped, and was immediately 
packed down in ice and salt. All the samples to facilitate keeping were placed 
in a room in the cold-storage warehouse, where the temperature was slightly 
below freezing. Here they were inspected daily, ice and salt added as required, 
and samples for study removed with sterile glass spatulas. 
For the determination of the total number of organisms, approximately 
1 cubic centimeter of the cream, which was melted as promptly as j)ossible 
after reaching the laboratory, was weighed in a tared, sterile weighing bottle, 
made up to 10 cubic centimeters with sterile water, and from this mixture 
were prepared appropriate dilutions for the counting of organisms. 
The plating was done on litmus lactose agar, half the plates of each sample 
being allowed to develop colonies at a temperature of 37° C., and the other 
half placed in the refrigerator, running from 18° C. to 20° C. It has been 
found necessary to use both these temperatures if an accurate idea is to be 
obtained of the changes undergone by organisms when submitted to continuous 
low temperatures, since there is apparently a dying off of certain groups of 
organisms in the early stages of storage, and the gradual increase of other 
organisms, which seem ultimately to thrive under what are usually conditions 
fatal to growth. 
The results obtained in the study of 8 samples of ice cream are tabulated 
as Series I, II, and III, of Table I. Series I extended over sixty-six hours; 
Seres II covered a total of one hundred and ninety-two hours, and Series III a 
total of two hundred and sixteen hours. 
As with all experiments where the bacterial flora is of as varied a character 
as that found in commercial cream, there is observed in this work a consid- 
erable variation in the behavior of different samples. Generally the tendency 
is to show a decrease for at least several days in a number of organisms 
developing at body heat, though occasionally these organisms persist and thrive 
at low temperatures. The organisms developing at the temperature of the 
refrigerator show usually a period of decrease which may last several days, to 
be followed later by a very pronounced rise. Sometimes the killing off of the 
organisms is very slight, their numbers remaining almost stationary or making 
a continuous upward curve. 
