salt.

The (H) concentrations observed here are practically all due to the dissociation of the inorganic acid, as the dissociation of the glutaminic acid is very low. The residual concentrations of similar solutions, standing over egg albumin, were found to be very trifling in a number of experiments similar to those carried out for free hydrochloric acid, to be referred to below.

10 cc. with phenolphthalein require 1'7 cc.
=155 mgrms. HCl for whole.

This result shows that while some of the acid comes

through combined with protein, some must be considered as existing in the form of free acid. This amounts to about 20 per cent of the whole acid in the substance shaken up with water.

In the presence of pepsin the mixture undergoes rather a slow digestion, and the amount of hydrochloric acid now found in the filtrate is increased, but not greatly, as the digestion is incomplete. This is shown by Table VIII., where the effect of adding more protein is also brought out. In these experiments 2 grms. of the powder and 50 mgrms. of pepsin were added to 100 cc. of water and digested through six hours at 40°. In some of the cases the coagulum from egg powder with 70 per cent of real protein, and fibrin with 29 per cent of real protein, was added. The results were as follows:

Total

N.

Glut. N.

5

ΙΟ

15

0'0196 0.0096 0'0711 0.0192 O'1211 0.0288

Ο ΟΙΟΟ

0'0519

0'0923

20

lost

0'0384

E

25

0.1765

0.0480 0.1285

0.8051

0.8244

III. The Behaviour of Protein Hydrochloride. In the foregoing we have seen the behaviour of hydrochloric acid combined with small groups comparable to the component structures of the protein molecule itself. The acid unites readily with the complex protein, as it does with the amino acid derivatives, and the question of its physiological action here now comes up. In the therapeutic use of hydrochloric acid it is usually administered in the form of diluted solutions of the free acid, and the only important objection to this is the strongly acid taste. To overcome this objection such bodies as the betain hydrochloride have found favour, and it has been shown above that a marked digestive activity is actually present with them. Combinations of proteins and hydrochloric acid have also come into use in recent years, and for these claims of acid strength are made. The experiments to be given below are intended to throw light on this point. In the investigations I used a form of acid albumin obtained by adding hydrochloric acid to egg albumin and evaporating to dryness at a low temperature. The dry substance was powdered and mixed with enough more albumin powder to bring the HCl content to exactly 5 per cent. The product has a strong acid taste, and is only partly soluble in water. The nitrogen content was found to be 12:25 per cent, but when the product is shaken up in water the amount of nitrogen dissolved is not large. Two experiments illustrate this. Two portions of 2 grms. were shaken up with 100 cc. of water and incu. bated at 40° through six hours. To one of these portions 50 mgrms. of killed pepsin had been added. At the end of the time the liquids were filtered and on the filtrates nitrogen determinations were made :

The protein in the 2 grms. of the powder used above amounts to about 1.53 grms. Slightly over 60 per cent of this digests when no more protein is present, but the addition of either egg or fibrin brings the digested amount down to a much lower figure. With increasing amounts of added protein the digested fraction progressively diminishes. This is undoubtedly because of the binding of the hydrochloric acid. In similar experiments with other amounts of protein added no greater weight of protein was digested. It appears, therefore, that acid held in this way is not capable of insuring an active digestion, even of its own protein.

The concentration of the dissociated hydrochloric acid is very low in the mixture of the powder with water. This was observed with the mixture made by adding 10 grms. of the powder to water enough to make 250 cc. of solution. The mixture was well shaken through an hour and allowed to stand over night. The acid strength was determined in a cell of the Hamburger type against o'1 N HCl. The result was found, CH = 000726. The insoluble residue was again shaken with water and allowed to stand. The supernatant liquid gave a lower acid value than before. This operation was repeated twice. The results of the four trials were as follows, showing for the (H) concentrations—

First water, o'00726; second water, o'00517; third water, 0.00483; fourth water, o'00352.

Assuming the hydrochloric acid to be all split off in the first water treatment the concentration should be 0.0548, or with 95 per cent dissociated 0.05296. The amount o free acid available for activation of pepsin is therefore but a fraction of that split off from the betain hydrochloride. A preparation of this character is sold under the name of oxyntin as a hydrochloric acid substitute.

This holding of the HCl by protein is shown easily in another way, using moist freshly coagulated egg albumin as a substratum. I'made a series of experiments in which different amounts of coagulated egg were mixed with 150 cc. of N/15 HCl, and either digested through a certain time or allowed to stand some hours before testing. In

CHEMICAL NEWS Aug. 6, 1915

In A, B, C, D, and G the digestions were carried through four hours at 50°, with the addition of 50 mgrms. of pepsin. In A and B practically all of the protein went into solution before the end of the period. In C some was left which held a little of the digesting liquid, and in D the amount was still greater. In G the portion digested was apparently very slight, and the volume of liquid which ran through the filter was much less. The weights of actual protein and acid added, expressed in percentages of the protein weight, are as follows:

In A an appreciable amount of free acid is found in the liquid after digestion, while in E, of the same original strength but not digested, the free acid concentration is nearly twice as great. In C there is scarcely enough acid for proper digestion, while in D there was an appreciable amount of albumin left. In G the acid concentration was far too low for digestion, as most of the egg remained undissolved. What passed through the filter was merely acid albumin. In E and F there was, of course, no digestion and very little solution; the amounts of free acid were consequently higher here.

After making the potential tests some of the liquids were returned to the original filtrates and mixed with wash water sufficient to bring all the volumes up to 250 cc. In this way practically everything soluble was washed out of the filters. 25 cc. portions of the filtrates were titrated with o'r N alkali and with o'I N silver nitrate, after separation of albumin, with the following results :TABLE XII.

NaOH with NaOH with NaOH

Per cent of formal AgNO3 HCI titration. titration. recovered.

methyl

orange.

phenolphthalein.

71

that the weak acid concentrations were probably not sufficient to prevent a slight bacterial action between the time of digestion and the titration. But the results of the silver nitrate determinations are interesting. If all the hydrochloric acid had passed into the filtrates the amount of silver nitrate required would have been exactly Io cc. in each case. But there was always a loss depending on the amount of acid held by the undigested protein. By prolonged washing all of this acid could have been dissociated and washed out. Such acid cannot be considered as mechanically retained, but is doubtless chemically combined. In Sample G two-thirds of the acid is so held, and here the protein residue is very large. For the actual digestion of protein by pepsin and acid it is not necessary that the amount of free acid should be large, but there must be some excess if the digestion is to be at all rapid. This slight excess may probably come in some cases as a result of dissociation. In Sample C we have about the limit of digestive action, but with the same amount of acid and double the protein there is practically no digestion in G.

These experiments are sufficient to show that combinations of protein and HCl, with not more than 5 or 6 per cent of the latter present, can have no value as digestive agents. This amount of acid is scarcely sufficient to permit the digestion of the protein itself, to say nothing of the digestion of added protein. In this respect the protein acid combinations are not comparable with the combinations with amino acids. In Experiment A, where the digestion is rather rapid and is practically completed in two hours, the (H) concentration was found to be PH 1696. It is interesting to note that this value is between the limits found by Soerensen (Biochem. Zeit., 1909, xxi., 297) for the optimum digestion of egg albumin, as measured by a quite distinct process, viz., the determination of the amount of nitrogen in the digestion filtrate which may not be precipitated by stannous chloride. His optimum is given at PH 163. It is evident, however, that any marked increase of albumin in A would lead to a decided decrease in the digestive activity.

In these experiments I have received valuable assistance from Miss Mary Hull, to whom my thanks are due.

Resumé.

It has been shown in this paper that the hydrochlorides of betain and glutaminic acid dissociate in aqueous solution to sufficient extent to permit the acid to aid pepsin in the rapid digestion of egg albumin or fibrin. This behaviour is probably typical of amino acid combinations in general. In the case of the betain salt the action is almost equal to that of dilute hydrochloric acid of the same gross concentration. With glutaminic acid hydrochloride the action is somewhat slower, but still marked. In either case, in the mixture of coagulated protein and the hydrochloride, a part of the HCl will leave the latter and become attached to the protein.

Mixtures made by combining hydrochloric acid with protein, in a sense analogous to the hydrochlorides of amino acids, are physiologically much less active. Such mixtures hold scarcely enough acid to digest themselves perfectly. If further amounts of protein are added with pepsin digestion becomes very slow. When the protein and hydrochloric acid are so related as to bring the (H) concentration down to PH = 2.96 the rate of digestion is slow. This is the case when the weight of the acid is about 3.5 per cent of the weight of the egg albumin and 150 cc. of N/15 acid is the liquid volume.

On the other hand, when the weight of the acid in 150 cc. of N/15 HCl is about 10 per cent of the weight of the albumin, and the hydrogen concentration of the supernatant liquid is about PH 169, we have very rapid digestion. This appears to be near the maximum of activity. We find all degrees of digestive activity between these limits. Dry preparations of protein and hydrochloric acid about midway between these limits cease to be physiologically active.

WELSH NOTES.

SIR CLIFFORD CORY on July 26 formally opened the new Metallurgical Buildings of the University College of South Wales and Monmouthshire, at Cardiff, which have been erected at a cost of some £3000. For the past seventeen years Prof. Read, the head of the metallurgical department of the University College, has been conducting the work of this important section of education considerably handicapped by want of proper accommodation and appliances. Despite these drawbacks, however, he has achieved valuable results, both in the way of routine education and original research, acquiring for the metallurgical school in Cardiff a position only second to that of the similar institution at Sheffield. Owing, however, to the influx of students from the Treforest School of Mines, who now take the metallurgical course in the study of fuel at the University College, an increased accommodation has become absolutely necessary.

The new buildings consist principally of a series of laboratories, the largest of which is that devoted to assay work. This is fitted with a wind crucible furnace and with all the necessary appliances for assaying. There are laboratories for metallography and for photomicroscopy, the latter with a dark room adjoining. The rest of the buildings include a big balance room, a reading room, a lecture theatre, the professor's private room, and usual offices. An important gift of the Welsh coal owners is that of a large new engine which has been installed in the engineering department for the purpose of conducting efficiency tests.

Lord Pontypridd presided at the opening function, and remarked in the course of his address that they were removing the reproach that had been levelled at the backwardness of their technical education, and in future, he thought, there would be little room for complaint. Lengthy speeches were also made by Sir Clifford Cory (in declaring the building open), Principal Griffiths, and Prof. Read. CHEMICAL NOTICES FROM FOREIGN SOURCES.

Bulletin de la Société Chimique de France.

Vol. xvii.-xviii., No. 7, 1915.

Formation of Aspirin.-D. E. Tsakalotos and S. Horsch. The authors have studied the formation of aspirin (acetyl salicylic acid) from acetic anhydride and salicylic acid as a function of the time and temperature. The equation is

C6H4OHCOOH+(CH3CO)2 =

+

+

C6H4O(CH3CO).COOH + CH3COOH. The products on the left-hand side of the equation when dissolved in water give three molecules of acid (one of salicylic acid and two of acetic acid), while those on the left-hand side give only two (one of acetyl salicylic acid and one of acetic acid). Thus, the transformation is 3(H) → 2(H). In the experiments a weighed quantity (about one molecule) of salicylic acid dissolved in benzene was put into a flask and a given weight of acetic anhydride (about one molecule) added. The variation of the acidity was determined by means of a N/50 solution of barium hydrate, using phenolphthalein as an indicator, 5 cc. of the benzene solution being removed from the flasks at different intervals of time and titrated. The temperatures at which the reaction was studied were 25°, 30°, and 50°. From the results obtained it was found that the velocity of the reaction is that of a reaction of the second order. The temperature coefficient for 10° 22, which agrees with van't Hoff's law, according to which an increase of temperature of 10° increases the velocity of the reaction two to three times. The acceleration of the reaction with the temperature has a limit, for at about 90° the aspirin formed begins to decompose with an appreciable velocity into acetic anhydride and salicylo-salicylic acid.

Nitration of Dimethyl- m - phenetidine.-Frédéric Reverdin. It is often found that aromatic compounds containing one or more ethoxy or ethyl groups behave differently from the analogous compounds containing one or more methoxy or methyl groups. Thus, when diethylp-aminobenzoic acid or its ethyl ether is nitrated, a mononitro derivative of monoethylamino benzoic acid or its ether is formed, one of the alcoholic radicles united to the nitrogen being eliminated, while in the same circumstances the corresponding dimethyl acid or its methyl ether gives a mononitro derivative in which the two alcoholic residues remain attached to the nitrogen. Similarly, p-phenetidine when nitrated behaves differently from p-anisidine, the ethyl residue in the dinitro compound of the former being less firmly attached to the oxygen than the methyl group in the corresponding dinitro-p-anisidine. The ethyl group of diethylaniline also gives this base characteristics which differ, from the point of view of the reactions it undergoes, from those of dimethylaniline. In order to add to these results the author has studied the nitration of dimethyl-mphenetidine, and will describe the analogous researches on diethyl-m phenetidine in a later report. When nitric acid is added to a cooled mixture of the dimethyl base and acetic acid the temperature hardly rises, and no nitration occurs. If, however, the mixture is heated gently on the waterbath the liquid turns reddish brown, and nitrous vapours begin to be formed at 70'. When the liquid is poured into water a more or less resinous precipitate is formed, which, after some hours, is transformed into yellow flakes. This substance is dinitro-4.6 nitrosomethylamino-3-ethoxyI-benzene (m. p. 113-114°). If in this nitration instead of heating on the water-bath the product of the reaction is allowed to stand at the ordinary temperature, the liquid gradually turns brown, and on pouring into water a canary-yellow precipitate is obtained. From it the above product of melting-point 113-114° can be separated, and also dinitro-4.6-dimethylamino-3-ethoxy-1-benzene of melting-point 1720. When the nitrosamine derivative (m. p. 113-114°) in solution in acetic anhydride is treated with nitric acid at the temperature of the water-bath the corresponding nitramine (m. p. 137-138°) is obtained. If it is boiled with hydrochloric acid it is transformed into dinitro-4.6-monomethylamino-3-ethoxy-1-benzene. If the nitration of dimethyl-m-phenetidine is compared with that of dimethyl-m-anisidine it will be seen that the same series of nitrated compounds is obtained, but the reactions begin at a higher temperature.

MISCELLANEOUS.

Institute of Chemistry.-Past List, June-July (1915) Examinations.-The results of the examinations of the institute of Chemistry recently held in London, Glasgow, and Dublin have now been published. Of eleven candidates who presented themselves for the Intermediate Examination, seven passed :-C. E. Corfield; A. Hancock; T. Hopkins, B.Sc. (Lond. and Wales); J. McLeod; J. Ogilvie, B.Sc. (Edin.); Agnes Shore; and A. Stewart, B.Sc. (Glas.). In the Examination in General Chemistry for the Associateship (A.I.C.), E. R. Taylor, A.R.S.M. (Lond.), passed. Of twenty-four candidates who presented themselves for the Final (A.I.C.) Examination, sixteen passed: - In the Branch of Mineral Chemistry-E. Arundel, B.Sc. (Lond.). In the Branch of Organic Chemistry— G. M. Bennett, B.A. (Cantab. and Lond.), B.Sc. (Lond.); A. J. Boyd; R. C. Denington; H. Hepworth, B.Sc. (Lond.); I. Hopper, A.R.C.S.I.; G. L. Hutchison, R.Sc. (Lond.); C. H. Lumsden, B.Sc. (Lond.); J. W. Porter, A.R.C.S.I.; and S. H. Tucker, B.Sc., A.R.C.S. (Lond.). In the Branch of the Chemistry (and Microscopy) of Food and Drugs, Fertilisers and Feeding Stuffs, Soils, and Water- G. A. Bracewell; P. Cheng, B.Sc. (Birm.); A. O. Jones; C. H. Manley, B.A. (Oxon); C. C. Roberts, M.A. (Cantab.); and S. Emsley, B.Sc. (Vict.).