2. When bacteria were suspended in olive oil or in pus, chlorlyptus showed marked germicidal action.
3. Chlorlyptus can be injected into the peritoneum or the pleural cavities of guinea-pigs in the proportion of 1 c.c. per 400 gm. of body weight without detriment to the animal.
4. Chlorlyptus in 5 per cent. oil solution (taking Clause 3 as comparison) can perhaps be injected in man as an antiseptic agent when there is a walled-in abscess in the peritoneum or pleural cavity where there is drainage, in the proportion of 0.5 to 1 c.c. per pound of body weight with good result.
REPORT ON THE GERMICIDAL ACTION OF CHLORLYPTUS ON PATHOGENIC BACTERIA IN VITRO AND IN VIVO
Experiment 1.—The germicidal action of eucalyptus oil.—Typhoid bacillus was destroyed in less than five minutes when exposed to the action of a 5 per cent. suspension of oil of eucalyptus. The exposure for four hours in a 5 per cent. suspension of chlorlyptus in paraffin oil was without effect on typhoid bacillus. It requires an exposure of two to four hours in a 10 per cent. suspension of chlorlyptus in paraffin oil to destroy typhoid bacillus.
Experiment 2.—Bacilliary action of chlorlyptus on the growth of pathogenic bacteria.—Typhoid and anthrax bacilli were selected for the experiment. Two series of five tubes each were made. The culture medium used was nutrient bouillon. Chlorlyptus was added in the following proportions: Tube 1, 1:10; Tube 2, 1:100; Tube 3, 1:1,000; Tube 4, 1:10,000, and Tube 5, 1:100,000. One series was inoculated with typhoid bacillus. All tubes were incubated for three days at 37 C.
Chlorlyptus inhibited the growth of typhoid bacillus when added to the bouillon in the proportions of 1:10. The growth of anthrax bacillus was inhibited by chlorlyptus when it was added in the proportions of 1:10, 1:100 and 1:1,000, as shown in the accompanying table. [The table was not submitted.—Ed.] In one instance the growth was markedly inhibited by chlorlyptus when added in the proportion of 1:10,000.
Experiment 3.—Germicidal action of chlorlyptus on typhoid bacillus.—Bouillon cultures of typhoid bacillus forty-eight hours old, and a suspension of forty-eight-hour agar cultures of typhoid bacillus in sterile salt solution were used for the experiment. Chlorlyptus was added in the proportion of 1:1,000; 1:1,500; 1:100; 2 per cent.; 3 per cent.; 4 per cent.; 5 per cent. and 10 per cent., respectively.
Inoculations were made in trypsinized peptone bouillon after the addition of chlorlyptus at different intervals, namely: at once, after five minutes, after ten minutes, after fifteen minutes, after thirty minutes, after one hour and after two hours, and tubes incubated at 37 C. for forty-eight hours.
Result: Growth was shown in all tubes except those in which chlorlyptus was added in the proportion of 10 per cent. and after the action of the antiseptic for two hours or longer.
Experiment 4.—Inhibitory action of chlorlyptus in the growth of typhoid bacillus.—Chlorlyptus was added to sterile bouillon in the proportion of 1:100, 1:1,000, 1:10,000 and 1:100,000, and incubated for forty-eight hours at 37 C. to eliminate any possible contamination of the bouillon during the manipulations. All tubes were found sterile and inoculated with typhoid bacillus.
Result: All tubes were found sterile again after being inoculated with typhoid bacillus and incubated at 37 C. for forty-eight hours, which shows chlorlyptus inhibited and the growth of typhoid bacillus in bouillon when this antiseptic was added in the proportions of 1:100 to 1:100,000.
Remarks: In another experiment made, chlorlyptus showed a weaker inhibitory action on the growth of typhoid bacillus.
Experiment 5.—Germicidal action of carbolic acid.—The technic was the same as that outlined in Experiment 1. except that carbolic acid was used instead of chlorlyptus.
Result: Carbolic acid showed a distinct germicidal action on typhoid bacillus in the proportions of 1 per cent. in ten minutes.
Experiment 6.—Action of nitrogen gas on the growth of typhoid bacillus in bouillon and nutrient agar when chlorlyptus was added to this culture medium.—Chlorlyptus was added to the bouillon in the proportions of 1:100, 1:1,000, 1:10,000 and 1:100,000, as outlined in Experiment 2; also to agar kept melted at 45 C. Tubes were inoculated with typhoid bacillus; plates were made of the inoculated agar tubes; all plates and tubes were incubated at 37 C. for forty-eight hours in an atmosphere of nitrogen gas.
Duplicate experiments were made with cultures of typhoid bacillus as above in bouillon and agar plates containing the same amount of chlorlyptus and incubated at 37 C. in ordinary atmosphere as control.
Result: Nitrogen gas did not show any appreciable increase of the germicidal action of typhoid bacillus when grown in medium containing chlorlyptus. Growth was about the same in cultures supplied with nitrogen gas as in those growing in ordinary atmosphere.