SULPHIDE PRODUCING ORGANISMS 
421 
no longer become blackened. In such cases the medium has become markedly acid 
to litmus. 
Egg albumen and blood serum were proved by Zorkendorfer and Petri and 
Maassen, to be favourable for the production of hydrogen sulphide by bacteria. 
Appel has obtained the formation in sterile urine, after the addition or flowers of 
sulphur. 
B. Reaction 
The reaction of the media was discovered to be a matter of considerable 
importance in the interest of sulphide formation. In order to ascertain the most 
favourable point of reaction, different degrees of alkalinity and acidity were selected 
in two per cent, peptone water, with phenolphthalein as indicator, as follows : -6,-3, 
o, + 3, +6, +8, +10, +12, +14, +16, +18, 4-20, +24, + 27, +30. 
It was observed that the organisms selected certain points at which the blacken- 
ing occurred most rapidly and deeply. These points ranged, as a rule, from + 10 
to + 16, differing with different organisms and, indeed, varying slightly from time 
to time with the same organism. 
The importance of reaction in the cultivation of bacteria is well known. But 
it is of yet greater importance for the formation of sulphide ; for a peptone tube may 
be very cloudy with the growth of a strong sulphide builder, in a too acid or too 
alkaline medium, with little or no blackening of the fluid or lead paper. Not 
infrequently, however, with the lapse of days, sometimes weeks, the medium seems 
to be so altered that sulphide formation commences. One has to consider also the 
action of acidity in preventing the precipitation of the sulphide of iron ; for while, 
say, + 12 shows marked blackness of paper and fluid, a considerably higher degree 
of acidity may present as much blackness on the paper, but the fluid be almost or 
entirely free. 
In ten per cent, peptone water the reaction is of much less consequence for the 
strong sulphide formers. The additional energy imparted to their activity in this 
medium evidently enables them to overcome the retarding influence of unfavourable 
reaction. It must not, however, be neglected even in ten per cent, peptone, 
especially in the case of weaker sulphide builders. 
A note on the indicators used may not be out of place here. The end reaction 
of phenolphthalein is very delicate in peptone solution. A two per cent, solution 
of peptone in tap water is acid to phenolphthalein to the extent of about 10 c.cm. 
normal acid per litre, and slightly alkaline to litmus ; a ten per cent, solution is acid 
to the extent of about 45 c.cm. normal acid per litre to phenolphthalein, but of 
nearly the same alkalinity to litmus as the two per cent, solution. Some substances 
in peptone .evidently respond very differently to the two indicators ; and neither of 
them appears reliable for the standardizing of these two solutions at a common point 
b 1 
