In these experiments of De la Croix, however, the nature of the bacteria experimented on was not determined, and there might be a mixture of several sorts. Koch has therefore sought to ascertain the action of disinfectants upon definite forms of microzymes by cultivating them in pure crops before applying the disinfectant. Those which he has chiefly experimented on are the red micrococcus prodigiosus, the bacteria of blue pus, and the bacillus anthracis.

The first two do not form spores, and are easily destroyed by disinfectants. The bacillus anthracis forms spores, and was therefore employed to test the action of disinfectants upon them.

Mode Of Experimenting- On The Action Of Drugs On Reproduction Of Bacilli

In order to avoid admixture with other species, Koch cultivated the first two on slices of potato, instead of in a solution. Upon one piece of potato the unaltered microzymes were sown (control specimen), and upon the others similar microzymes which had been exposed to the action of disinfectants. If the microzymes had been destroyed by the disinfectants, no result occurred, but if not, then a crop was obtained which, in comparison with the control specimen, was more or less abundant, according as the action of the disinfectant had been less or more complete.

For the cultivation of the anthrax bacillus, Koch used as a soil gelatine mixed with some other nutritive substance, usually meat infusion and peptone sterilised and spread upon a slip of purified glass, and exposed to such a heat as just to solidify it. Koch did not use his solidified blood-serum in these experiments. This could be placed under the microscope, and the growth of bacilli observed from day to day. Middle-sized test-tubes were then partially filled with the disinfecting solutions, and silk threads, steeped in a fluid containing bacilli and then dried, were placed in them; from time to time a thread was removed from the tubes by means of a previously heated platinum wire and placed on the slide, which was then subjected to microscopical observation. In this way it was easy to determine what strength of solution, and what time of exposure to its action, were required to destroy the spores.

The results of experiments made in this way with carbolic acid were very surprising. It was to be expected that carbolic acid would readily destroy the spores, but this was not the case. A 1 per cent. watery solution had almost no action upon them even after they had been exposed to it for 15 days; 2 per cent. slightly retarded their growth, but it did nothing more; 3 per cent. killed the spores in 7 days; 4 per cent. in 3 days; and 5 per cent. in 1 day.

This comparatively slight action of carbolic acid on spores and the long time that it requires to destroy them show that it cannot be relied upon as a universal disinfectant. But it has nevertheless great power in destroying microzymes which have not formed spores.

The fresh blood of an animal which has died from anthrax contains only bacilli and no spores. When it is mixed with its own bulk of a 1 per cent. solution of carbolic acid, it can very soon afterwards be injected into an animal without producing any marked symptoms. A 1/2 per cent. solution will not do this, so that the limit lies between .5 and .25 per cent. of the mixture (v. p. 97).

The action of carbolic acid on other fully-developed microzymes, or on the spores, is almost the same as on the anthrax bacilli.

The following table gives the result of Koch's experiments with other substances, the figures indicating the number of days during which the spores had been submitted to the action of the antiseptic previous to cultivation. The black-faced figures indicate that the spores were destroyed, and their germination prevented by exposure to the disinfectant for that number of days; a * indicates that their vitality was diminished, and that the crop from them was scanty; a + indicates that their growth was retarded; *+ that it was both scanty and retarded. The disinfectants are divided into three groups. The first contains the group of fluids; the second of solutions in water; and the third of solutions in alcohol, ether, or oil.

Group I. - Fluids.

Distilled water ....

7

15

20

35

90

Alcohol (absolute) . . .

1

3

5

10

12

20

30

40

50

65

110

Alcohol (1 to 1 of water) . . .

1

3

20

30

40

50

65

110

Alcohol (1 to 2 of water) . . .

1

3

20

30

40

50

65

110

Ether . . . . . .

1

5

8*

30

Acetone . . . . . .

2

5*

Glycerine . . . . .

1

3

10

20

30

40

50

65

110

Butyric acid ....

1

5

French salad oil ...

5

30

90

Bisulphide of carbon .

1

5

10

20

Chloroform ....

1

3

10

20

100

Benzol . . . . . .

1

5

10

20

Petroleum ether . . . .

1

5

Turpentine oil . . . . .

1*

5

10

Group II. - Solutions In Water.

Chlorine water (freshly made) ....

1

5

Bromine (2 p. c. in water) ....

1

5

Iodine water (1 in 7,000) . . . . . .

1

Hydrochloric acid (2 p. c. in water) .

1

5

10

Ammonia .......

1

5

10

Ammonium chloride (5 p. c. in water) . . .

1

5

10

25

Common salt (saturated solution) . . . .

1

5

10

20

0

Calcium chloride (saturatedsolution). . . .

1

5

20

40

Barium chloride (5 p. c. in water) . . .

5

10

45

100

Ferric chloride (5 p. c. in water) . . .

2+

6

Potassium bromide (5 p. c. in water) . . . .

5

10

25

Potassium iodide (5 p. c. in water) . . . .

5

10

25

80

Corrosive sublimate (1 p. c. in water) . . .

1

2

Arsenic (1 p. c. in water) . . . . . .

1

6

10

Lime water . . . . . . .

5

10

15*+

20*+

Chloride of lime (5 p. c. in water) . . . .

1+

2*+

5

Sulphuric acid (1 p. c. in water) . . . .

1

3

10*

20*

Zinc sulphate (5 p. c. in water) . . .

1

5*

10*

Copper sulphate (5 p. c. in water) . . .

1

5*

10*

Ferrous sulphate (5 p. c. in water) . . . .

2

6

Sulphate of aluminium (5 p. c. in water) . . .

1

5

12

Alum (4 p. c. in water) . . . . . . . . .

1

5

12

Potassium chrornate (5 p. c. in water) . . . .

1

2

Potassium bichromate (5 p. c. in water) . . . . .

1

2

Chrome alum (5 p. c. in water) . . . . . . .

1

2

Chromic acid (1 p. c. in water) . . . . . .

1

2

Potassium permanganate (5 p. c. in water) . . . .

1

Do. do. (1 p. c. in water) . . .

1

2

Potassium chlorate (5 p. c. in water) . . .

2

6

Osmic acid (1 p. c. in water) ....

1

Boracic acid (5 p. c. in water) not quite dissolved

1

2

6+

10+

Borax (5 p. c. in water) . . . . . . . .

5

10

15

Sulphuretted hydrogen water ....

1

5*

Ammonium sulphide .....

1

2

5

Oil of mustard with water ....

1

5

10*

Formic acid (sp. gr. 1.120) ....

1

2

4

10

Acetic acid (5 p. c. in water) ....

1

5

Potassium acetate (saturated solution) . . . .

1

4

10

Lead acetate (5 p. c. in water) ....

1

5

12

Soft (potash) soap (2 p. c. in water) .

1

5

12

Lactic acid (5 p. c. in water) ....

1

2

5

Tannin (5 p. c. in water) . . . . .

1

5

10

Trimethylamine (5 p. c. in water) . . . . .

1

5

12

Chloropicrin (5 p. c. in water) ....

1

2

6

12

Benzoic acid (saturated solution in water. . . .

1

5

10

45

90

Benzoate of sodium (5 p. c. in water) . . . .

1

2

5

10

Cinnamic acid (2 p. c. in water 60 and alcohol 40 parts) . . . . . . .

1

3

5

10

Indol (in excess in water) . . . . .

1

5

10

25

80

Skatol (in excess in water) ....

1

5

10

25

80

Leucin (1/2 p. c. in water) . . . . .

1

5

10

Quinine (2 p. c. in water and 40 alcohol 60 parts)

l*+

5*+

Quinine (1 p. c. in water with HC1) . . . .

1

5

10

Group III. - Solutions In Alcohol, Or Ether, Or Oil

Iodine (1 p. c. in alcohol) ....

1*

2*

Valerianic acid (5 p. c. in ether)

1

5

Palmitic acid (5 p. c. in ether) ....

1

5

Stearic acid (5 p. c. in ether) ....

1

5

Oleic acid (5 p. c. in ether) ....

1

5

Xylol (5 p. c. in alcohol) . . . . .

1

5

30

50

90

Thymol (5 p. c. in alcohol) ....

1

6

10

15

Salicylic acid (5 p. c. in alcohol)

1

6

10

15

Salicylic acid (2 p. c. in oil) ....

5

10

20

80

Oleum animale (Dippel's oil, 5 p. c. in alcohol)

1

5

12

Oleum menthae piperitae (5 p. c. in alcohol) . .

1

5

12

From this table it appears that the ordinary method of separating between formed and unformed ferments by precipitation with alcohol and solution in glycerine cannot be relied upon as a trustworthy means of separating them, since neither alcohol nor glycerine destroys the activity of formed ferments.

It is remarkable that ether and turpentine oil, which are both ozone carriers, should have such a marked action in comparison with other fluids. This is in harmony with some recent observations of Paul Bert and Regnard, who found that oxygenated water in sufficient quantity destroys the bacteria of anthrax.

The spores of anthrax bacilli resist in an extraordinary way the action of certain substances which usually are fatal to life, as hydrochloric acid (2 per cent.), salicylic acid (1 per cent.), concentrated solutions of chloride of sodium, chloride of calcium, metallic solutions, borax, boric acid, chloride of potassium, benzoic acid, benzoate of sodium, cinnamic acid, and quinine.