A Process for Obtaining Strong Tuberculosis Toxins from Tubercle Bacilli.
Year: 1898

page_text: Date of Application, 5th Mar., 1898
Complete Specification Left, 5th Dec., 1898-Accepted, 4th Mar., 1899
DE BOTICAIIUN.
A Process for Obtaining Strong Tuberculosis Toxins from
Tubercle Bacilli.
I, OLIVER IMRAY, of Birkbeck Bank Chambers, Southampton Buildings,
in the County of London, Chartered Patent Agent, do hereby declare the nature
of this invention (as communicated to me from abroad by The Farbwerke
vormals Meister Lucius and Brüning of Hoechst a/Main, in the Empire of
Germany, Chemical Manufacturers) to be as follows:
The tuberculosis toxins hitherto known by the name of "Tuberculin" have
been obtained by cultivating bacilli on a nourishing fluid and by then sub-
jecting the culture-fluid or the bacilli themselves to an appropriate treatment.
The fluid, for instance, was simply filtered off and concentrated, if requisite,
by evaporation or reduced together with the bacilli to a small volume by evapora-
tion, whereupon the bacilli were removed by filtration (Koch).
A concentration of the toxin of tuberculosis has also been obtained by using
precipitating agents, such for instance as alcohol, which separate the effective
substance (Koch). Effective substances have also been obtained from the bacilli
freed from the culture-fluid by trituration of the dried bacilli, addition of water
and treatment in a powerful centrifugal apparatus (New tuberculin by Koch).
A liquid moreover has been obtained from the pressure of the damp bacilli
triturated with arenaceous quartz, which is said to contain specifically effective
elements (Buchner's plasmin of tuberculosis). Besides, bacilli have been ex-
tracted with weak alkaline solvents (soda) and also bacilli previously freed from
their fat with ether, chloroform, bisulphuret of carbon &c., have been employed
or the above methods have been combined in order to withdraw from the tubercle-
bacilli more or less of the poison of tuberculosis which they contain.
Quantitative calculations of the poison originally contained in the bacilli and
of the poison remaining after applying the above methods of extraction have
however shewn my foreign correspondents that the residues hitherto considered
useless still contain a great deal of toxin of the tuberculosis and that in making
use of these residues, a strong toxin of tuberculosis hitherto unknown may be
obtained, by heating the already extracted, triturated tubercle bacilli, deprived
of fat, with water, aqueous glycerine, solutions of salt &c. to about 150° C.,
air being excluded.
The preparations free from glycerine in a dry state, hitherto known, obtained
either from the fluid of tuberculosis or from the body substance of tubercle
bacilli, have at their best a toxic value of infallibly killing three sound guinea-
pigs with the quantity of poison, contained in 1 gramme whilst preparations
containing glycerine are much less poisonous.
My foreign correspondents have, on the other hand, obtained preparations from
tubercle bacilli already subjected to the known extraction methods, of which
1 gramme contains the deadly dose for not less than 50 sound guinea-pigs. This
result may perhaps be still better ilustrated when it is recollected that fresh

page_text: bacilli free from glycerine and dried bacilli do not quite contain in 1 gramme
the requisite deadly dose for 3 sound guinea-pigs.
It evidently follows from these statistics that the stronger toxin must represent
an element different from the principal mass of the body substance.
The tubercle bacilli must therefore contain besides the specific toxin of tuber-
culosis a great quantity of non-poisonous or less poisonous substance.
That the stronger toxin of tuberculosis is particularly suitable for the purpose
of giving, immunity has been proved by numerous experiments with large and
small animals.
The method of obtaining it is illustrated by the following example:-
The tubercle bacilli are freed from the easily extracted elements for which the
glycerine extraction and the soda extraction are mostly used. The bacilli are
then washed with distilled water, precipitated with alcohol, dried and freed from
their fat with ether, whereupon they are again dried and reduced by trituration.
The triturated bacilli are then ground, when freed from fat, with a glycerine
solution of 4 per cent. in the proportion of from 1 to 10. Sometimes instead of
the glycerine solution ordinary water, salt solutions &c, have been used.
The pulpy mass, obtained after trituration, is heated for 1 hour to 150° C.
in a closed vessel previously freed from air. The mass still hot is then treated
in a powerful centrifugal apparatus and the fluid thus obtained serves finally
for obtaining a stronger toxin of tuberculosis, amounting on the average to
50 per cent. of the whole quantity of poison originally contained in the treated
tubercle bacilli. Even then the contents of the residual mass of bacilli is not
exhausted of valuable toxin. By repeating the whole process a yield may be
obtained, which, though less abundant, is still sufficiently remunerative.
The liquid toxin of tuberculosis thus obtained is not very stable and it is
therefore transformed by way of precipitation with alcohol into a solid form,
in which it may be kept unchanged for an indefinite time.
Dated this 5th day of March 1898.
ABEL & IMRAY,
Agents for the Applicant.
A Process for Obtaining Strong Tuberculosis Toxins from
Tubercle Bacilli.
I, OLIVER IMRAY, of Birkbeck Bank Chambers, Southampton Buildings,
in the County of London, Chartered Patent Agent, do hereby declare the nature
of this invention (as communicated to me from abroad by The Farbwerke
vormals Meister Lucius and Brüning of Hoechst a/Main, in the Empire of
Germany, Chemical Manufacturers) and in what manner the same is to be per-
formed, to be particularly described and ascertained in and by the following
statement:-
The tuberculosis toxins hitherto known by the name of "Tuberculin" have
been obtained by cultivating bacilli on a nourishing fluid and by then sub-
jecting the culture-fluid or the bacilli themselves to an appropriate treatment.
The fluid, for instance, was simply filtered off and concentrated, if requisite,
by evaporation or reduced together with the bacilli to a small volume by evapora-
tion, whereupon the bacilli were removed by filtration (Koch).
A concentration of the toxin of tuberculosis has also been obtained by using
precipitating agents, such for instance as alcohol, which separate the effective

page_text: substance (Koch). Effective substances have also been obtained from the bacilli
freed from the culture-fluid by trituration of the dried bacilli, addition of water
and treatment in a powerful centrifugal apparatus (New tuberculin by Koch)..
A liquid moreover has been obtained from the pressure of the damp bacilli
triturated with arenaceous quartz, which is said to contain specifically effective
elements (Buchner's plasmin of tuberculos). Besides, bacilli have been ex-
tracted with weak alkaline solvents (soda) and also bacilli previously freed from
their fat with ether chloroform, bisulphuret of carbon &c., have been employed
or the above methods have been combined in order to withdraw from the tubercle-
bacilli more or less of the poison of tuberculosis which they contain.
Quantitative calculations of the poison originally contained in the bacilli and
of the poison remaining after applying the above methods of extraction have
however shewn my foreign correspondents that the residues hitherto considered
useless still contain a great deal of toxin of the tuberculosis and that in making
use of these residues, a strong toxin of tuberculosis hitherto unknown may be
obtained, by heating the already extracted triturated tubercle bacilli, deprived
of fat, with water, aqueous glycerine, solutions of salt &c., to about 150° C.,
air being excluded.
The preparations free from glycerine in a dry state, hitherto known, obtained
either from the fluid of tuberculosis or from the body substance of tubercle
bacilli, have at their best a toxic value of infallibly killing three sound guinea-
pigs with the quantity of poison contained in 1 gramme, whilst preparations
containing glycerine are much less poisonous.
My foreign correspondents have, on the other hand, obtained preparations from
tubercle bacilli already subjected to the known extraction methods, of which
1 gramme contains the deadly dose for not less than 50 sound guinea-pigs. This
result may perhaps be still better illustrated when it is recollected that fresh
bacilli free from glycerine and dried bacilli do not quite contain in 1 gramme
the requisite deadly dose for 3 sound guinea-pigs.
It evidently follows from these statistics that the stronger toxin must represent
an element different from the principal mass of the body substance.
The tubercle bacilli therefore contain besides the specific toxin of tuber-
culosis a great quantity of non-poisonous or less poisonous substance.
That the stronger toxin of tuberculosis is particularly suitable for the purpose
of giving immunity has been proved by numerous experiments with large and
small animals.
The method of obtaining it is illustrated by the following example.
The tubercle bacilli are freed from the easily extracted elements for which the
glycerine extraction and the soda extraction are mostly used. The bacilli are
then washed with distilled water, precipitated with alcohol, dried and freed from
their fat with ether, whereupon they are again dried and reduced by trituration.
The triturated bacilli are then ground when freed from fat, with a glycerine
solution of 4 per cent. strength in the proportion of about 1 gramme bacilli to
10 cubic centimetres of solution. Sometimes instead of the glycerine solution
ordinary water, salt solutions &c. have been used, such as conimon salt, soda,
potash or their carbonates not exceeding 1'5 per cent.
The pulpy mass, obtained after trituration, is heated for 1 hour to 150° C.
in a closed vessel previously freed from air. The mass still hot is then treated
in a powerful centrifugal apparatus and the fluid thus obtained serves finally
for obtaining a strong toxin of tuberculosis, amounting on the average to
50 per cent. of the whole quantity of poison originally contained in the treated
tubercle bacilli. Even then the contents of the residual mass of bacilli is not
exhausted of valuable toxin. By repeating the whole process a yield may be
obtained, which, though less abundant, is still sufficiently remunerative.
The liquid toxin of tuberculosis thus obtained is not very stable and it is
therefore transformed by way of precipitation with alcohol into a solid form,
in which it may be kept unchanged for an indefinite time.

page_text: Having now particularly described and ascertained the nature of this inven
tion and in what manner the same is to be performed I declare that what I
claim is:
A process for obtaining strong tuberculosis toxins by treating, substantially as
desribed, with solvents in vacuo at a temperature above 100° C., tubercle bacilli
extracted, freed from fat and triturated according to the methods hitherto em-
ployed (for instance for obtaining "Old-tuberculin" and tuberculoplasmin).
Dated this 5th day of December 1898.
ABEL & IMRAY,
Agents for the Applicant.