6. Solubility of Edestin in Sulphuric Acid.

Preparation 23, which I have shown on page 51 to be nearly pure edestin sulphate, required nearly ten times as much decinormal sulphuric acid for solution as edestin chloride required of decinormal hydrochloric acid. It was, therefore, necessary to use a decinormal acid in carrying out experiments similar to those just described, but in the following table the amount of acid is stated in terms of a centinormal solution, in order to avoid possible confusion on comparing the figures with those of the preceding tables. This preparation had an original acidity equal to 10 cc. of a centinormal solution per gram and, as none of it dissolved in water, this amount of acid is added to that applied to the preparation, so that all the acid present with which the edestin could react is here taken into account.

TABLE XVIII.-EDESTIN SULPHATE DISSOLVED BY CENTINORMAL SULPHURIC ACID.

Amount of Edestin Sulphate Dissolved Per cc. of Centinormal H2SO

Solution.

No. 23. 0.0000 0.0035

0.0058 0.0109 0.0125

0.0000

0.0130

0.0026 0.0076 0.0118 0.0139 0.0145 0.0130

Acidity of the Solution of Edestin Sulphate in cc.

Acidity of the Solution Per Gram of Dissolved Edestin in cc.

From these figures it is plain that a much larger quantity of sulphuric acid is required to dissolve a given amount of edestin than of hydrochloric acid. The edestin sulphate corresponding to the bichloride is insoluble in water. Whether the soluble compound formed with a sufficient quantity of sulphuric acid is a salt of edestin or whether a hydrolytic derivative of edestin is first produced which forms soluble compounds with the larger quantity of sulphuric acid, was not ascertained, but the behavior of edestin with hydrochloric acid would indicate this to be the case.

7.

The Solubility of Edestin in Phosphoric and Acetic Acids. Phosphoric acid reacts with edestin as a monobasic acid, in accordance with its dissociation into the ions H and H,PO,.

One gram of the air-dry preparation, suspended in 6 cc. of water, was completely dissolved when treated with 14 cc. of a centinormal solution of phosphoric acid. With 13 cc., 0.8920 gram was dissolved, which is very nearly the calculated quantity, namely, 0.9230 gram.

Acetic acid likewise reacts with almost the calculated amount of edestin, since I found that 13 cc. of a centinormal solution dissolved 0.8804 gram.

Both these acids dissolve somewhat more edestin than does an equivalent quantity of hydrochloric acid, 13 cc. of a centinormal solution of which dissolved about 0.7770 gram. This difference appears to be due to the formation of different proportions of the more basic edestan, as the following experiments indicate. To each of 3 gram portions of edestin suspended in 6 cc. of water, were respectively added 14 cc. of a centinormal solution of each of these acids. After standing for about two hours at 25°, the acid in each was exactly neutralized, and an equal volume of 20 per cent. sodium chloride solution was added. The amount of the insoluble edestan present in the portion with hydrochloric acid was 0.1786 gram, in that with phosphoric acid o. 1484 gram, and in that with acetic acid 0.0565 gram. These results are approximately in accord with the degree of ionization of these acids and appear to explain the relative incompleteness of the reactions with the respective acids.

8.

The Solubility of Edestin in Nitric Acid.

Nitric acid dissolves edestin chloride in nearly the same proportion as does hydrochloric acid, but a larger quantity of the former acid is required to dissolve the neutral edestin at about 20° than of the latter. At 35° one air-dry gram of neutral edestin, equal to 0.9300 gram dried at 110°, was completely dissolved by 14 cc. of centinormal nitric acid, but by 12 cc. at this temperature much remained undissolved. At 20°, I gram was completely dissolved by 20 cc., all but a very few milligrams by 19 cc., while with 18 cc. much remained undissolved.

This quantity of acid is in such close agreement with that required for the formation of a trinitrate that it strongly suggests

that such is formed, but it is not safe to assume this without other evidence to confirm it.

That a compound with 2 molecules of nitric acid should exist which is more soluble in warm than in cold water, is in harmony with the known behavior of this acid with proteoses, some other protein substances and the histons.

A more extended study of this question is necessary, and I shall take it up as soon as possible.

III. COMPOUNDS of EDESTIN WITH ALKALIES.

It is well known that protein substances react with alkalies as well as with acids, in which respect they closely resemble the purin bases, which, as pronounced bases, form salts with acids. and are also able, like weak acids, to form with bases definite compounds, of which the silver salt is one of the best known, since it is used for the separation of these bodies from their ammoniacal solution.

Many unsuccessful attempts have been made to obtain definite compounds of the proteins with bases, especially with the heavy metals. The chief reason that these failed probably lies in the fact that the small quantity of acid which these substances still contain when their solutions are made neutral to litmus has been overlooked, and also to the fact that salts of the heavy metals are hydrolytically dissociated to such an extent as to make it difficult or impossible to maintain suitable conditions for the formation of definite metallic compounds with the proteins.

The experiments next to be described show that edestin enters into definite reaction with potassium and sodium.

1. Solubility of Edestin in Sodium Hydroxide Solution. Preparation 31, which was strictly neutral to phenolphthalein, was used to determine the solubility of edestin with definite quantities of sodium hydroxide, in a manner similar to that employed in determining the solubility of edestin with definite quantities of hydrochloric acid.

Gram portions of the air-dry preparation were suspended, in glass-stoppered bottles, in enough water, free from carbonic acid, to make 20 cc. with the alkali solution to be afterwards added. To the first, 2 cc., to the second, 3 cc., and so on up to 7 cc., of centinormal sodium hydroxide solution were added. After agitating frequently for an hour, the solutions were allowed to stand

at rest for another hour, during which time the undissolved matter settled, leaving the solution nearly clear. From each portion 10 cc. were drawn out with a pipette, evaporated to dryness, and the residues dried to constant weight at 110°. The amount dissolved by each quantity of alkali added is shown in the following table:

Table XIX.-AMOUNT OF EDESTIN 31 DISSOLVED BY A CENTINORMAL SOLUTION OF SODIUM HYDROXIDE.

The amount, in grams, of edestin dissolved per cubic centimeter with each quantity of alkali added, was the following: TABLE XX.-AMOUNT OF EDESTIN 31 DISSOLVED PER CC. OF CENTINORMAL SODIUM HYDROXIDE.

NaOH

2 CC.

3 cc.

0.0622

0.0855

4 CC. 0.1090

5 cc. 0.1217

6 cc.

0.1306

7 cc.

100

0.1283 gram dissolved. These figures show that the amount dissolved per cubic centimeter steadily rises, until with 6 and 7 cc. it reaches a maximum. This is due to the difficulty with which all of the soluble sodium edestin is separated from the relatively large quantity of edestin remaining undissolved in those portions to which the smaller amounts of alkali had been added, since the great extent of surface presented by the fine crystalline powder strongly adsorbs the soluble sodium edestin and also, to the indiffusibility of the substance, since any sodium edestin formed within the solid particles is removed with difficulty. With increasing quantities of alkali the proportion of undissolved edestin diminishes and the proportion dissolved per cubic centimeter correspondingly increases until it reaches a quantity but little less than that calculated for a complete reaction between one molecule of edestin and one of sodium hydroxide.

Another series of gram portions of preparation 30, 1 gram of which had an acidity requiring 2 cc. of centinormal sodium hydroxide solution for neutralization to phenolphthalein, was treated in the same way as the preceding and the following amounts were found to be dissolved:

TABLE XXI.-AMOUNT OF EDESTIN 30 DISSOLVED BY CENTINORMAL SODIUM HYDROXIDE SOLUTION. 4 CC. 6 cc. 7 CC.

I CC.

2 CC.

3 cc.

5 cc.

NaOH

8 cc.

9 cc.

100

0.0490 0.0940 0.1636 0.3208 0.4800 0.6060 0.7120 0.8188 0.8782 dissolved. gram

In this case the effect of the carbonic acid contained in this preparation is plainly manifested, since 9 cc. of the alkali were required to dissolve the same amount that 7 cc. dissolved of the perfectly neutral preparation 31. The small quantity appearing

to be dissolved by 1 and 2 cc. consisted mostly of suspended matter unavoidably drawn off with the solution. The agreement between these results and those obtained in the first experiment is best shown by the solubility curves given in Plate II.

2. Solubility of Edestin in Potassium Hydroxide Solution. Preparation 28, 1 gram of which required 2 cc. of centinormal alkali for neutralization to phenolphthalein, was treated with a centinormal solution of potassium hydroxide in the way described for experiments with sodium hydroxide. Table XXII shows the weight in grams dissolved by the several quantities of alkali.

TABLE XXII.-AMOUNT OF EDESTIN 28 DISSOLVED BY CENTINORMAL POTASSIUM HYDROXIDE SOLUTION.

0.0000 0.0150 0.0938 0.1944 0.3294 0.4772 0.7592 0.8500 gram dissolved.

The amount dissolved per cubic centimeter above 2 cc. was as follows:

TABLE XXIII.—AMOUNT Dissolved Per cc. of CenTINORMAL POTASSIUM HYDROXIDE.

KOH

3 cc.

4 CC.

0.0938

0.0972

5 cc. 0.1098

6 cc. 0.1193.

0.1518

100

0.1417 gram dissolved. In this, as in the experiments with sodium hydroxide, the proportion of dissolved edestin increased as the proportion of undissolved edestin diminished, the amount dissolved by 8 cc. being nearly equal to that calculated for a complete reaction between equal numbers of molecules of each substance. The somewhat higher figure found for 7 cc. is doubtless due to a slight error of manipulation, as indicated by the rise in the curve given in Plate II, showing the results of this experiment.

Another similar series of gram portions of edestin 30, treated with centinormal potassium hydroxide in the same way as 28 had been treated, gave the following results :

TABLE XXIV.-AMOUNT OF EDESTIN 30 DISSOLVED BY CENTINORMAL POTASSIUM HYDROXIDE SOLUTION.