INSECTIVOROUS PIvANTS. 149 



chloride). Fibrin was dissolved in 4 hours or less. Hydrocyanic acid, which according to Fiechter 

 (Basel, 1875) does not injure enzymes while it arrests the actions of, and even kills, yeasts and bacteria, 

 was used with equally favorable results. Fibrin (50 mgs.) was digested in 1.5 hours in a solution 

 made up as follows : 5 cc. pitcher liquid, 5 cc. of 2 per cent solution HCN, and one drop of strong 

 hydrochloric acid. 



These results seemed to exclude the hypothesis of bacterial action, yet Vines thought best to 

 strengthen his position by experiments on the other side; i. e., to use conditions favorable to bacterial 

 growth, so said, but fatal to enzymes. To this end he treated pitcher liquid, at a temperature of 

 35° to 40° C, with alkalies in various strengths aud for various periods, and after neutralizing it, 

 tested its digestive activity. He found that the activity of the liquid was destroyed by treatment with 

 I per cent sodium hydrate for i hour, and by treatment with 5 per cent sodium carbonate for 3 hours. 

 These alkaUes produced a precipitate in the pitcher liquid. 



A study of the relation of digestive activity to the amount of free acid present in the liquid gave 

 the following results: Tubes A, B, C, and D, containing respectively i per cent, 0.5 per cent, 0.25 

 per cent, and 0.125 per cent of hydrochloric acid, kept in the incubator, digested i gram of fibrin in 

 the order of rapidity of C, B, A, D, that in D not disappearing within 2 hoiurs while C required 

 only half an hour. 



The author succeeded in removing from the pitcher liquid a substance with which a new digestive 

 liquid could be prepared. This he did by obtaining a bulky precipitate with absolute alcohol, phos- 

 phoric acid, and lime water, the liquid being finally neutralized with ammonium carbonate. This 

 precipitate was shaken up with a 0.25 per cent solution of HCl, and the liquid filtered. In this 

 liquid rapid digestion of fibrin took place with good biuret-reaction. A month later the same success 

 was obtained by using some of the precipitate which had been kept dry in the presence of chloroform 

 vapor. 



The liquid of unopened pitchers was found to be generally acid, that of opened ones, either acid 

 or neutral. The pitcher liquid is quite inert in the absence of acid. Only a minute quantity of 

 proteid was present, nor was there evidence of the presence of zymogen. Hence he thinks that either 

 the enzym is not a proteid, or if it is, is present in extremely minute quantity, though it is difficult 

 to accept the second view because of the remarkable digestive activity of the liquid. 



Active glycerin extracts were prepared, some with, others without previous treatment of the 

 pitcher-material with absolute alcohol. The extract from untreated material was the more active 

 in the digestion of fibrin. These pture glycerin extracts did not retain their activity for a prolonged 

 period but one was active at the end of 2 months. Only relatively young pitchers yielded this extract. 

 This indicates that enzym secretion ceases some time before the pitchers show signs of withering. 



The presence of a digestive agent in the plant tissues affords collateral evidence that this agent 

 is not a bacterium but an enzym. Vines thinks it highly improbable that any bacterium would 

 retain its vitality after two months sojourn in pure gylcerin.* His observations led him to differ 

 with Gorup-Besanez who described the products of digestion as peptones. He finds the main prod- 

 uct to be an albumose, allied to deutro-albumose, and failed to detect the presence of a true peptone. 

 He says: 



"The only certain proof that a liquid contains true peptones Ues in the fact that such a liquid 

 continues to give proteid reactions after sattu-ation with ammonium sulphate, since this neutral salt 

 precipitates all proteids except true peptones; but this test was not applied by Gorup-Besanez in any 

 of the above-mentioned researches. As I have already pointed out, I have invariably found that a 

 solution of the products of digestion by pitcher-liquid gives no proteid reaction after saturation with 

 ammonium sulphate, all the proteids present having been precipitated by the salt." 



No readily dialysable proteid was demonstrated. The nature of the ultimate product of digestion 

 was not determined with complete success, though Vines thinks it may be leucin or some allied body. 



He lays stress on the fact that the liquid of unopened pitchers of N. mastersiana is distinctly 

 acid, a fact which controverts the idea that the secretion of acid is the result of stimulation. 



In 1898 Vines published a paper on the proteolytic enzyme of Nepenthes in which he 

 recorded his observations on the effects of exposure to high temperatures, of treatment 

 with alkalies, and of filtration of the pitcher-liquid, upon its digestive activity. 



The method of experiment with heat was to maintain the liquid for a given time at 

 the required temperature, and then to institute a digestion experiment, adding fibrin and 

 the necessary acid; in nearly every case there was a control digestion experiment with 

 unheated liquids. The results were as follows : 



*The writer being very sceptical instituted some experiments and found the spores of the hay bacillus alive in pure 

 glycerin in one case after 19 months, and in another case after 10 months. This spore bearing material was put into 

 the glycerin in cotton plugged test tubes and tested for viability every few months, always with positive results. 



