Defensive Ferments of the Animal Organism/Methods in Use/The Optical Method
II. The Optical Method.
The principle of the Method.—The optical method enables us to demonstrate alterations in optically active substrates by a determination, with the aid of a polariscope, of changes in their angle of rotation.
The aim of the optical method is, in principle, exactly the same as that of the dialysation process. In the latter we determine the transformation of a colloid into a diffusible crystalloid. This transformation is the result of a hydrolytic decomposition. In the optical method we start, for purely technical reasons, not with albumen, but with peptone produced from the latter. We cannot use albumen, because it would prevent us determining the angle of deviation of the substrate-serum mixture. It would either give rise to precipitates, or else render the mixture so heterogeneous, that slight changes of rotation would be very difficult to follow. When using the optical method, we allow the decomposition, produced by the ferments present in the serum, to set in later, than in the dialysation process. We remove part of the decomposition from the influence of the ferment, when we convert albumen into peptone in the test-tube. It must be our aim to maintain the peptone mixture in as high a molecular state as is possible, as experience has shown, that decomposites of too low molecularity are not attacked by some kinds of serum, which decompose more highly molecular peptones. It is very clearly shown, in this connection, that the conception of the unity of the proteolytic ferments does not correspond at all with the reality. There is not the slightest doubt, that different ferments exist for different stages of decomposition. The principal problem, in the application of the optical method to biological questions, was the elaboration of a method of dealing with highly molecular peptones, which are very closely related to the albumens.
The Application of the Optical Method.—This is very simple. 1 c.c. of serum, absolutely free from hæmoglobin, is placed in a test-tube. It must not contain any form-elements, and must be sterile. To this is added 1 c.c. of a 5 to 10 per cent. solution of peptone, prepared from the organ in question. Of course, peptones may also be prepared from bacilli, or else from certain proteins. The serum is mixed with the peptone solution and poured into a polarization tube, of a capacity of 2 c.c., and the angle of rotation of the mixture, at a temperature of 37° C., is determined. The deviations are noted at certain intervals. If there is no change in the deviation, then we conclude that no decomposition has taken place. Should there be an alteration in the rotation after some time, then we must infer a fermentative decomposition, such as has been demonstrated by special experiments with ferment solutions.
We shall now give a description of the preparation of the peptone.