T H E JOURNAL OF I N D U S T R I A L A N D ENGINEERING CHEMISTRY
Nov., 1916
observed, b u t not more t h a n would be obtained on aqueous solutions of sucrose. It is only necessary t o give one out of several series of tests. Meat extract solutions containing about 4 per cent of total sugar, mostly reducing, were each treated with 4 0 mg. sucrose (about j per cent) and allowed t o stand a few minutes before neutralization. Reducing sugar was then determined on this solution a n d also on a control solution identical except t h a t no sucrose was added. A blank test on 40 mg. cane sugar showed reduction equivalent t o 0 . 2 1 per cent sugar on t h e amount of sample used. The extract solutions gave t h e following results (percentages) : Time of standing Minutes 1
Sucrose present 5.27
42
4.94 5.00
RIDUCING SUGAR FOUXD I n (control I n sucrose solution Increase 3.95 4.16 0.21 4 . 2 28
4.41 4.35
3 4.92 4.30 4.46 Reduction of aqueous sucrose solution. . . . . . .
~:;~]Av.O.16
.. ,
0.16 0.21
The temperature of t h e room during this experiment was about 2 5 ' C., which was favorable t o inversion. The above figures show t h a t t h e estimation of reducing sugar in t h e presence of sucrose is quite practicable by this method, t h e increased reduction being within the limits of error of ordinary sugar methods. It should be remembered also t h a t t h e solution is acid before t h e addition of hydrochloric acid, so t h a t although inversion is slow, in t h e presence of sucrose t h e whole process should be carried through without too much delay. After t h e addition of t h e hydrochloric acid the filtrate should be neutralized immediately. Where only total invert sugar is &aired, t h e solutions do not change for several days. Clarification becomes more complete as t h e solution stands. It is believed t h a t t h e methods outlined in this paper offer accurate means of determining reducing sugars in meat extract before a n d after inversion. S U X MAR Y
I-Clarification of meat extract solutions for estimation of sugar by Fehling's solution is best accomplished by a n excess of picric and phosphotungstic acids, followed by a minimum amount of hydrochloric acid. 11-In t h e presence of sucrose, reducing sugar may be determined within 0 . I or 0 . 2 per cent, provided proper precautions are observed. 111-Total reducing sugar may be determined within 0.1per cent.
u. s.
M E A T INSPECTIOK
LABORATORY
KANSASCITY, K A N ~ A S
LIGNOCERIC ACID FROM ROTTEN OAK WOOD By M. X. SULLIVAN Received July 3, 1916
I n t h e study of the origin of organic soil constituents a chemical study of rotten wood was undertaken in these laboratories in t h e year 1908. A large amount of dead oak wood was collected from a woody section near Washington, D. C. Since vanillin had long been reported1 as a constituent of wood, the ground-up wood was extracted with alcohol, the alcohol extract freed Singer, Monatsh , 3 (1882), 395.
1027
from alcohol a n d t h e residue treated with ether. F r o m the ether extract, by the customary methods of testing for aldehydes, a small amount of a n aldehydecontaining resin of a vanillin odor was obtained which gave t h e color reactions of vanillin. Since vanillin is sublimable, about 2 0 0 g. of wood were heated between large watch-glasses t o obtain sublimate matter, and though vanillin was subsequently found in wood and other vegetable material by appropriate methods,l practically all the material sublimed from t h e wood was insoluble in hot water and was of a f a t t y nature. To obtain a large amount of this sublimable f a t t y matter, about 2 0 lbs. of rotten wood were placed in a retort, connected with a Liebig condenser, and subjected t o heat. As a result of regulated heating with Bunsen burners a steady stream of material varying from tarry matter t o a yellowish white crystalline solid distilled over through t h e condenser. The material obtained was soluble in hot alcohol. On cooling, crystalline matter separated out of the alcohol. On filtering while t h e alcohol was still warm and washing with cold alcohol, most of t h e coloring matter went into t h e filtrate. The crystalline matter was then placed on a porous plate and treated with cold petroleum ether until colorless. The purified substance recrystallized from alcohol melted a t 737 5 ' . The compound was soluble in ether, hot alcohol, warm petroleum ether, a n d was insoluble in water. Subsequently, b y much t h e same method, lignoceric acid2 was obtained from peat soil. Lignoceric acid has t h e elementary composition of C, 78.2; H, 13.0; 0, 8.8, As determined by Dr. E. C. Shorey at t h e time he was working on t h e lignoceric acid of peat soil, t h e elementary composition of t h e substance obtained by me from wood was C, 7 8 . 6 ; H , 1 3 . 3 5 ; 0, 8 . 5 . I t was thought a t t h e time (1909) t h a t t h e substance from t h e wood might be identical with lignoceric acid which Hell3 found in beech wood tar. The elementary composition and melting point, however, were suggestive t o me of cerotic acid from beeswax, which has a percentage composition of C, 78.78; H, 1 3 . 1 3 ; and melts a t 78'. No further work was done on t h e identification of t h e compound from t h e rotten wood until very recently, when t h e question of t h e identity of the wood substance was revived by t h e finding of cerebrosides4 in mold from soil and b y t h e work of Levene and West, who found t h a t lignoceric acid is a n oxidation product of cerebronic acid derived from cerebrosides of brains b y hydrolysis. Lignoceric acid forms a lithium salt insoluble in hot methyl alcohol. Accordingly, the material from t h e wood was dissolved in methyl alcohol and the resulting solution was treated with a methyl alcohol solution of lithium acetate as long as a precipitate formed. I n this way t h e material collected from t h e Sullivan, THIS JOURNAL, 6 (1914), 919. 0. Schreiner and E. C. Shorey, U. S. Dept. Agr., Bureau of Soils, B U Z Z . 74 (1910). 3 Ber., 13 (1880), 1709. 4 Sullivan, Science, 38 (1913), 678. 6 J. Riol. Chem., 15 (1913), 193. 1
2
T H E JOCRNAI, OF I N D U S T R I A L A X D ESGINEERING C H E X I S T R Y
1028
dry distillation of t h e wood was divided into two portions, t h e one, the major portion with a lithium salt insoluble in hot methyl alcohol; t h e other, small in amount, with a lithium salt soluble in methyl alcohol. The acids were liberated b y boiling with dilute sulfuric acid and extracting t h e cooled mixture with ether. Recrystallized from alcohol, the respective acids melted a t 78 and 8 ; " . T h e acid from the lithium salt insoluble in methyl alcohol, melting a t 7 8 ' , was further purified and t h e melting point rose t o 80-82'. This latter acid, presumably lignoceric acid, was compared with cerotic acid made from beeswax according t o t h e method employed. b y Marie.* T h e acid obtained from t h e wax melted a t 78-79'; .mixed with t h e acid from rotten wood melting a t So', t h e melting point of t h e mixture was 72-74'. T h e lowering of t h e melting point of t h e mixture shows t h a t the acid from t h e wood is not identical with cerotic acid. T h e wood acid mixed with lignoceric acid from peat soil gave a mixture melting at 79-81', so these acids are probably identical. Lignoceric acid forms a lead salt melting a t I I ~ ' , and a lithium salt decomposing about 190-195 0.3 The acid from t h e rotten wood gave a lead salt melting a t 116-II~', a lithium salt decomposing a t 195' and gradually liquefying t o 2 1 0 - 2 1 5 ' , a t which temperature it formed a brown liquid. From t h e solubility of t h e acid and its salts t h e percentage composition, t h e decomposing point of the lithium salt, t h e melting point of t h e lead salt, and t h e non-lowering of t h e melting point when mixed. with lignoceric acid from soil, i t may be concluded t h a t the acid derived from the rotten oak wood b y dry distillation is lignoceric acid identical with t h e lignoceric acid obtained b y Hell4 from beech wood t a r and from "carnaubon" from ox kidneys b y Rosenheim and MacLean.' The lignoceric acid, or a closely related substance, exists in t h e wood, for i t has been obtained b y extraction with alcohol. To obtain this acid t h e finely ground mood was extracted with hot alcohol and the part which separated on cooling was saponified with alcoholic soda. P a r t of t h e sodium salt was insoluble in hot alcohol. T h e filtrate from t h e hot alcoholic soda was freed f r o m alcohol, with replacement by water. T h e hot water mixture was filtered hot. The filtrate treated with sulfuric acid gave a crystalline acid which was purified b y crystallization from alcohol and ether. The free acid gave a lithium salt insoluble in methyl alcohol. The acid freed from lithium melted at 80-82' and was found t o be identical with t h e acid obtained from wood b y dry distillation. The second acid obtained in the dry distillation of wood, the acid melting a t 8 j ' with a lithium salt soluble in hot methyl alcohol, is believed t o be t h e inactive cerebronic acid described b y Levene and West.4 It is a white, easily pulverizable, non-hygroscopic substance soluble in ether and warm .alcohol. I 2
Ann. chim. p h y s . , ( 7 1 I (1896), 145. Hell and Hermann, Bev.. 13 (ISSO), 1713.
SOILFERTILITY INVES?IGATIONS DEPAR?MEK'C
OF A G R I C U L T U R E
WASHIKGTOB,
D. C.
METHOD OF EXTRACTION AS AFFECTING THE DETERMINATION OF PHOSPHORIC ACID IN SOILS By
HALEA N D a '. L. HARTLEP Received July 29, 1916
HaRRISON
While determining t h e phosphoric acid in soils during some work done in cooperation with t h e Missiouri State Fruit Experiment Station, occasion was also taken t o compare 0. L. Brauer's2 method of extracting t h e phosphoric acid 2 hrs. with 2 N HNOs with t h e official method of the Association of Official Agricultural Chemists3 of extracting I O hrs. with HC1 (SP. gr. I . 1 1 j). T h e soils were taken b y hIr. 4. D. Kilham of t h e
a Meyer, Brod, and Soyka, Mn?ialrh., 34 (1913), 1113.
1 2
Biochem. J . , 9 (1915), 103.
II
On cooling the alcohol it separates o u t in round masses, often in thick lamellae, which show a fine radiating structure. I t is free from nitrogen and burns with t h e smell of burning fat. I t s sodium salt is insoluble in water and not r e r y soluble in hot alcohol, especially in hot alkaline alcohol. On rubbing, i t becomes electrified. T h e general properties of t h e acid, its solubility, t h e solubility of t h e sodium and t h e lithium salts suggest t h e inactive form of cerebronic acid described b y Lerene and West. A further indication t h a t cerebronic acid exists in rotten wood is t h a t from a sodium alcoholate extract of t h e rotten wood material resembling cerebronic acid has been obtained. T h e alkaline alcohol solution was filtered hot and cooled in a n ice box. The precipitate which formed was slightly soluble in boiling water. The part insoluble in hot water was recrystallized successively from 95, 80 and 60 per cent alcohol. The nearly colorless material made a fine suspension in hot j o per cent alcohol and the turbid mixture passed through filter paper. ~On concentrating t h e solution, white matter settled out in mamillary masses, which on. drying melted a t 106-108O, t h e melting point of active cerebronic acid. This material was soluble in warm glacial acetic acid, from which i t settled out as a jelly on cooling. T h e p a r t most soluble in cold glacial acetic acid melted a t 90-94'. Cerebronic acid, as described b y Levene and Jacobs,' has three forms: ( I ) t h e optically active form melting a t 106-108'; ( 2 ) t h e inactive form melting a t 82-85'; and (3) a mixture of these melting about 92' All in all, it is believed t h a t cerebronic acid exists in rotten wood, though absolute proof was not obtained. A pleasing hypothesis is t h a t ccrebrosides exist on the living tree and t h a t from t h e cerebrosides there forms in t h e rottening of t h e wood, under t h e action of molds, bacteria, and t h e processes of oxidation, cerebronic acid and lignoceric acid. Of the presence of cerebrosides in wood no tests have been made; of t h e presence of cerebronic acid in rotten wood there are indications; and of t h e presence of lignoceric acid, a close relative of these, there is a satisfactory proof.
4 L O G . Lit. 6
Vol. 8 . S o .
8
J . Bid. Chem., 12 (1912), 381. THISJOURNAL, 6 (1914), 1004. Bureau of Chemistry, Bull. 101, 14.
