1908. | Studies on Enzyme Action. 829 
Ultimately we were led to devise a method which not only renders possible the 
estimation of hydrogen cyanide with a fair approximation to accuracy, but also permits 
of a considerable number of determinations being taken in hand in rapid succession—a 
matter of some importance. The solution (10 ¢.c.) in which hydrolysis has taken place is 
added to a decinormal solution of silver nitrate (20 cc.) previously mixed with an equal 
bulk of a decinormal solution of sodium acetate. The mixture is then heated (5—10 mins.) 
on a water bath. A bulky precipitate of silver cyanide is thus formed, which may be set 
aside until it is convenient to complete the analysis. The precipitate, having been filtered 
off and washed with boiling water, is transferred to a beaker with the aid of about 50 c.c. 
of water ; it is then dissolved in 5 c.c. of strong ammonia, the precipitate adhering to the 
filter being removed by means of 5 c.c. of ammonia, diluted with 25 ¢.c. of water, and the 
beaker in which precipitation is effected also washed out with 5 cc. ammonia and 
30—40 c.c. of water. The combined solutions are mixed with 4 c.c. of a normal solution 
of sodium chloride and poured through a thistle funnel into a round-bottomed Jena flask 
containing a solution of 25 grammes of tartaric acid. The liquid is raised to the boiling- 
point and distilled during 15 mins. in a current of steam, the distillate being received in 
a flask containing a slight excess of potash. The solution is then titrated With a solution 
of silver nitrate. A small amount of benzoic acid often distils over, but this does not 
appear to interfere with the titration. 
To determine the amount of glucose produced during hydrolysis, the standard method 
described by Brown, Morris and Millar,* involving the use of Fehling’s solution, was 
adopted, as it was found that neither amygdalin nor hydrogen cyanide had any reducing 
action on Fehling’s solution and that emulsin had a very slight action, the effect of which 
was determined and allowed for. The procedure is as follows :— 
10 c.c. (or 20 c.c. at the end of the first hour) of the liquid in which the action has been 
stopped by adding a drop of sulphuric acid are introduced into a Wurtz flask and the 
sulphuric acid neutralised by adding a previously determined quantity of N/5 potash ; the 
liquid is then diluted to about 125 c.c. and distilled by passing in steam during about 
30 minutes. The liquid is next rinsed into an evaporating basin, evaporated to a small 
bulk, transferred to a 50-c.c. flask, cooled and diluted to 50 ¢.c. The required quantities 
(25 e.c.) of each of the two Fehling’s solutions are mixed in a 400-c.c. wide Jena glass 
beaker with such a quantity of water that addition of the sugar solution will make the 
volume up to 100 c.c.; this beaker is heated in a boiling-water bath during 5 mins., then 
removed and 25, 15 or 10 cc. (according to its strength) of the solution in which glucose 
is to be determined is added. The beaker is then replaced in the water bath, covered 
with a clock-glass and heated during exactly 12 mins. The precipitated cuprous oxide is 
collected on an asbestos filter in a Soxhlet tube, washed with about 200 c.c. of boiling 
water, then with 10 c.c. of alcohol and dried in a water oven. The oxide is oxidised by 
heating in a current of air and then reduced to copper by heating in a current of purified 
dry hydrogen, cooled and weighed. The initial oxidation is necessary, in order to prevent 
volatilisation of part of the copper, which occurs when the cuprous oxide is heated directly 
in a current of hydrogen, probably owing to the presence of organic matter. 
’ 
Using Fischer’s glucoside as test material, which is resolved simply into 
glucose, benzaldehyde and hydrogen cyanide by emulsin, the amounts of 
hydrogen cyanide found have been uniformly low—on the average from 
one to two units below the calculated percentage values—so that the process 
is certainly open to improvement and simplification ; we hope to improve it, 
In numerous experiments in which the proportions of reducing sugar and 
* “Chem. Soc. Trans., 1897, vol. 71, p. 281. 
