While such compounds like asparagine, e. g., represent plant food, tyrosin is considered as being hurtful to plants. Whereas leucine and similar compounds, in the presence of nitrates, can give rise to the process of dentrification, somuch so that, under favorable conditions, leucine is thus quantitatively converted into leucinic acid, according to the equation:

>CH-CH2-CH.NH2-COOH+NOOH = H2O+N2+

other bodies with different chemical structure cannot initiate that process. Important as is the knowledge of the organic matter or humus in the soil, our present scope of information is extremely limited, to say the least. This is very well illustrated by the fact that out of 3794 pages in which Beilstein3 treats of organic chemistry, but two pages are devoted to humus and humin substances.

PLAN OF THIS INVESTIGATION

Whereas the identification of one or another organic nitrogenous compound in the soil is of certain interest, yet of greater importance, it seems to us, is to demonstrate what kind of bodies occur in predominant proportions. In a previous article1 the writer has demonstrated that the organic nitrogen in Michigan peat soils is made up of acid amides, monamino acids and diamino acids. What is true of peat soils may not necessarily hold good for other soil types, for a number of reasons. While peat soils are exceedingly rich in humus and nitrogen, other soil types usually contain a small amount of both of them. Whereas the humification process in peat soils, because they are saturated and oftentimes covered with water, is essentially one of putrefaction taking place in the absence of air, the humification in other soils is one of eremacausis inasmuch as it occurs chiefly in the presence of air.

Further, it will be well to recollect that the method applied for the separation of the organic nitrogenous bodies in peat soils was essentially the Hausmann-Osborne method used in the protein chemistry. However, the very assumption that organic nitrogenous bodies occurring in soils can be separated

3See Beilstein's Handbuch der Organischen Chemie, third edition, first volume, pages 1108 and 1109.

4 See Organic Nitrogenous Compounds in Peat Sails, Journal of the American Chemical Society, 32, 396. 5See, The Utilization of Peat, by Samuel L. Jodidi,Journal of the Americam Peat Society 2, (April, 1909).

16Ztschr. physiol. Chem., 27, 95 (1899); 29, 136 (1900); also Journal of the American Chemical Society, 25, 3231(1903).

by the same method as applied for protein decomposition products is of a somewhat hypothetical nature and needs direct verification, especially in view of the fact that phosphotungstic acid throws down not only diamino acids, but also peptones, purine derivatives, alkaloids and other compounds.

The nature of the organic nitrogen in soils being of considerable general interest as well as of paramount importance to agriculture, it may be worth while briefly to show what considerations have led the writer to the above assumption which, it may be stated here, has proved to be correct. Starting from the idea that soil organic matter or humus is the result of decayed plants all of which contain a large if not predominant portion of their nitrogen in the shape of proteins, it was but natural to assume the existence in the soil of some disintegration products of proteins. Now, protein bodies can be split into their constituent parts either by chemical means (e. g., mineral acids or alkalies), or through the activity of certain microorganisms, or through the agency of enzymes. Of these means the first mentioned is out of the question inasmuch as even peat soils-the most acid soils known-usually show but a slight acidity largely due to the presence of weak organic acids and acid salts On the other hand decomposition of protein bodies in the soil can and undoubtedly does take place through bacterial activity or the agency of enzymes widely distributed in the vegtable kingdom. Furthermore, disintegration in the soil of protein holding materials into simpler molecules can be accomplished also in a purely chemical way. A. E. Taylor,' e. g., has shown that leucin can be recovered from a sterile suspension of casein in pure water; he was also able to recover arginin from a solution of protamin sulphate in pure water, after the lapse of about one year. These facts assume considerable signficance in processes taking place for a considerable length of time, such as in the soil. While, however, partial hydrolysis yields albumoses, peptones or polypeptides-viewed in the light of the brilliant researches by Emil Fischer-total hydrolysis leads to the simplest molecules (acid amides and amino acids) out of which the protein molecule is built up. Schematically the degradation of the protein bodies can be represented in the following way: Proteins→ Albumoses Peptones (Polypeptides) → Amino Acids and Acid Amides → Ammonia → Nitrites Nitrates. The motive force in these processes is the higher chemical potential of the protein as against albumoses and the other phases.

7A. E. Taylor, "On Fermentation," Univ. Calif. Publ. Pathol., 1 (1907), page 223.

8 Untersuchungen uber Aminosauren, Polypeptide und Proteine, Berlin, 1906.

From the foregoing it follows that theoretically the formation in the soil of smaller molecules (amino acids, etc.) out of the various protein bodies contained in decaying vegetable materials seems to be altogether possible and even probable. Nevertheless such deductions as these must be verified by direct experiments as presented in this bulletin, because of the existence in the soil of a variety of conditions which may more or less interfere with protein hydrolysis or mask it.

DISCUSSION OF THE PROBLEMS INVOLVED.

While details will be found in the experimental part, a very brief outline of the questions involved may be given here. The hydrochloric acid extract of the soil was evaporated practically to dryness and distilled with cream of magnesia. The distillate contained, in the form of ammonia, all the nitrogen corresponding to the amides present in the soil. The residue from distillation with magnesia was extracted with water, and the extract, after acidultating with sulphuric acid, treated with PTA. The precipitate contained the diamino acids and the filtrate from that precipitate represented the monamino acids.

Now, in the case of protein bodies proper the distillation of the acid-treated protein with cream of magnesia gives pure ammonia. This, however, may not necessarily hold true for soils, recalling that some protein bodies, through decay yield organic bases. Dimethylamine,10 for instance, is formed through putrefaction of fish, trimethylamine" through putrefaction of wheat flour and fish. Under certain conditions, decay of organic materials furnishes the bases putrescin and cadaverin. Putrescin can also be formed from arginin12 and ornithin,13 through bacterial activity, just as cadaverin can result from lysin.14 These processes can be expressed by the following equations:

NH2

1

NH2

NH2+

NH-C-NH-CH2-CH2-CH2-CH-COOH+H2O=OC<NH2

Arginin

Urea

9The abbreviation "PTA" will stand for the words: phosphotungstic ac

10 Ber. d. Chem. Ges. 18, 87.

11 Jahresbericht uber die Fortschritte der Chemie, 1858, 231.

12Ztschr. physiol. Chem., 43, 338 (1904-5).

13Ztschr. physiol, Chem., 29, 334 (1900).

14Ibid.

NH,-CH-CH,-CH,-CH.NH,-COOH (Ornithin);
NH2-CH2-CH2-CH2-CH.NH2-COOH = CO2+

Ornithin

NH2-CH2-CH2-CH2-CH2-NH2;

Putrescin

NH2-CH2-CH2-CH2-CH2-CH-NH2-COOH = CO2+

Lysin

NH2-CH2-CH2-CH2-CH2-CH2-NH2

Cadaverin

Hence, it is not out of the question that the distillation of the acid-treated soil with magnesia may yield in addition to ammnoia, also organic bases. If the distillate contains ammonia only, then it must, when saturated with hydrochloric acid and evaporated to dryness, yield a salt which heated in a test tube will not melt nor be charred, but will sublime and condense on the cooler parts of the tube. The same salt when treated with platinic chloride must yield a compound the analysis of which is bound to lead to the formula PtCl, 2NH Cl. This latter salt when decomposed with hydrogen sulphide must contain, in the filtrate from platinum sulphide, pure ammonium chloride. All of these operations were actually made with positive result thus showing the presence of ammonia only.

Further, if the PTA precipitate actually contains diamino acids we logically must expect them to show all the precipitation reactions which diamino acids generally display. Moreover, inasmuch as protein hydrolysis leads to a amino acids of the general type R.CH NH, COOH all of which, with the exception of glycocoll, contain an asymmetric C-atom, both the diamino acids in the precipitate and the monamino acids in the filtrate, if they be such compounds, must in accordance with the fruitful theory of Van't Hoff15 and Le Bel be optically active (dextrotatory or levorotatory). Furthermore, both must show the presence in their molecules of amino-groups as well as of the carboxyl-group. The presence of two amino groups in the diamino acids must cause them to show a dis tinctly alkaline reaction, while the monamino acids in which one NH-group is neutralized by one COOH-group must show neutral or slightly acid reaction. An exception to the latter rule are aspartic acid, COOH CH, CHNH, COOH, and glutaminic acid, COOH CH, CH, CHNH, COOH, containing in their molecule one NH-group and two COOH-groups.

2

And if the NH-groups of the amino acids are fixed, e. g., converted into the-N-CH, group by treating with formalde

15 Van't Hoff, Lagerung der Atome im Raume, Braunschweig, 1894, also Ber. d. Chem Ges., 18, 2277; 20, Ref., 448, 21, 265, etc.

16

hyde, then the COOH group must come into play by causing acid reaction, etc. All of the above mentioned experiments and reactions, having as their object the demonstration of the presence of acid amides and amino acids in the soil extracts, have actually been carried out with positive results and are given in the experimental part of this bulletin.

EXPERIMENTAL

In order to find out the nature of soil humus formed from various organic materials under field conditions, the experiment station field, located on the Wisconsin Drift, was divided into a number of plots of 1/10 or 1/20 of an acre each. The various plots were treated with materials like manure, hay, straw, etc., which on cultivated fields actually represent the principal sources for humus formation. For the purpose of rendering the sources of nitrogen in each plot easily reviewable it was thought best to put down the data in question in the form of a table.

TABLE I-THE SOURCES OF NITROGEN IN THE VARIOUS