ARSENOTUNGSTIC ACID AS AN ALKALOID AL REAGENT BY EDWIN ROBERT LITTMAN THESIS FOR THE D E G R E E O F KACHE L O R O F SCI E N G E IN CHEMISTRY COLLEGE OK LIBERAL ARTS AND SCIENCES UNIVERSITY OF ILLINOIS ] « :•_> \> "v I 0 ?^u Wl "5 ro UNIVERSITY OF ILLINOIS P c iu Mav 24 192 P - L I92 THIS IS TO CERTIFY THAT THE THESIS PREPARED UNDER MY SUPERVISION BY E_l_'J^r__PpJbejrt__L.i t_t n lanri ENTITLED Arsenotunget i c Acid as an Alkaloidal Reagent. IS APPROVED BY ME AS FULFILLING THIS PART OF THE REQUIREMENTS FOR THE DEGREE OF Pack el or of Science in Cherrist-rv. Instructor in Charge HEAD OF DEPARTMENT OF K.'i ri'i no n j us.. . ■ m ACICN 0 V/LEDGrBL/ENT I wish to take this opportunity of expressing my sincere thanhs to Dr. George Denton Beal, who, by his good will and invaluable assistance has made this inves tigat ion «. success* Digitized by the Internet Archive in 2015 https://archive.org/details/arsenotungsticacOOIitt HISTORICAL OUTLINE Many complex metallic acids yield precipitates with organic bases. The best known example is the precipitation of an ammonium salt of phosphomolybdi c acid, (ME^JgPO^.lSMoO^, when ammonium molybdate is added to a solution of phophoric acid, as the first step in the determination of phosphorus from practically all sources. The use of phosphornolybdic acid has since that time been extended to the precipitation of more complex bases. Ph<£jjho tungstic acid has been used to a much greater extent in this way than has phophomolybdic acid. The especial value of phopho tungstic acid lies in its use in the separation of the protein and non-protein forms of nitrogen, as a substitute for the copper hydroxide reagent of Stutzer. 1 p Bertrand has prepared a series of silico- tungstic acids of which one form of the silicc-duodeci tungstic acid, 4K£0, SiQg, ' 7 : lEWOg, 2ZRgQ, has been used successfully by the Chapin method for the determination of nicotine. Further studies of silico- tungstic acid as an alkaloids! reagent have been made by Beal and Peterson and by Bueh and Beal^ • These reagents have shown so much promise that it has been decided to study the behavior of other complex metallic acids, including the arseno- tun stic acids, with particular reference to their availability as precipitating reagents for alkaloids. « , _ o_ Chemical literature yields little on the subject of arseno tungstic acids* Among the references to the preparation of various forms of the acid so far mention has only been made twice to the reactions of arseno tungstic acids with organic compounds. These are to the work of S.R. Benedic t* and Morris Macleod on the reaction between arseno tungstic acid and uric acid. Arseno tungstic acid has been suggested as a reagent for the determination of uric acid, based upon the reducing action of the uric acid on the tungsten. It was found that uric acid reduced the arseno tungstic acid to "tungsten blue". From this has been developed a colorometric method for uric acid. The reaction, however, does not depend upon the formation af a complex molecule containing the compounds. In neither of the two articles above was any attempt made to isolate the acid or any of its salts. The authors state that the composition of the acid was unknown. In 1888 an article a was published by Gibb on the preparation of arseno tungstic acids according to the method of Fr emery. Gibbs, in his own work obtained a salt of the following composition: lo'.VOg .As^O^. 6AggO 11H 0. The method of Gibbs is as follows:- To an aqueous solution of arsenic acid add freshly precipitated barium tungstate, followed by sulphuric acid to decompose the barium tungstate. Filter the mixture and evaporate on the water bath until crystals just form. Remove the dish and allow to cool. The precipitate is a mixture of two acids alpha and beta. _ „ , . -3- Dissolve this precipitate in boiling water. Add Potassium chloride to form the potassium salts. A precipitate should separate immediately. Allow Hie solution to cool, when yellow prisms should deposit. As soon as this occurs decant the liquid. The two products so obtained must be recrystallized until pure. The first precipitate is the alpha and the second the -beta salt. The formulae are: alpha ASgO ; . .9 W0 r/ • 14HgO beta As 0 _ . 18 W0 f . .XH 0 2 5 3 2 Since 1888 no satisfactory method for the preparation of an acid of definite composition has been published. It is true that quite a number of acids have been prepared, but there is doubt as to their composition, yields and methods of preparation. With but one exception the preparation of the acids involved such proceedures as fractional crystallization to separate the Af various compounds formed. Zehrman^ in 1900 published a paper on the preparation of an arseno tungstate of definite composition. His method is as follows:- To a saturated solution of sodium- tungstate add "syrupy" arsenic acid until the solution is distinctly acid, then one half as much again, finally add solid ammonium chloride to precipitate the ammonium arseno tungs tat . The product must be purified by repeated salting out. It has the formula 3(UH 4 ) 2 0. ASgO~. 181/0,,. 14H 2 0. This method was used as a starting point. - 4 - THEQRETIOAL CONSIDERATION. There is no definite theory to account for the combination of arsenic acid and a tungstate to form a complex. In view of this lack the condition of the experiments must be closely 4 followed in order to duplicate results. The method of Hehrman was modified in an attempt to get better yields of the pure salt. The modification changed only the method of precipitation. Instead ox saturating the solution with ammonium chloride and immediately filtering the product, the salt was added until a precipitate just formed and then allowed to stand. At the end of twenty four hours a very crystalline precipitate will have been deposited. This precipitate is a pure salt, as purification by salting out v/ith ammonium chloride twice gave no changes in the analysis. When an attempt was made to analyse the salt for arsenic many difficulties had to be overcome before concordant results could be obtained. The arsenic v/as precipitated as magnesium ammonium arsenate, but even after three precipitations the ignited residue showed the presence of tungstic oxide (WQ_). An electrolysis of a solution of the ammonium arseno tungstate caused violent "popping" but no liberation of arsine. This "popping" and the fact that the salt changed color but did not decrepitate at 100° 0. led to a further investigation as to its physical properties. The salt was placed in a dilatometer and the uulL heateu. slowly xo 11^° heigth of the column , * -5- (Mijol) was read for each degree rise in temp era ture. The curve (.Figure I.) shows that there is a transition point at 97° - 98° C. At study of the changes in crystal structure further advances the probability of meta-stability. A sample of the dry salt when placed on a heated microscope (Figure 3) stage changes in color from yellow to green as soon as the temp- erature reaches the transition point, but the change is crystal structure is not so rapid. If, however, a drop of water is added to the sample on the stage the crustal change occours as soon as the water has evaporated. Figure two shows the original salt (a) and after treatment wi th water and heat (b). The failure of the arsenic precipitation can be laid to the meta stable compound being present to some extent if we assume that the meta stable compound will not give up its arsenic as readily as the stable material. It was then decided to decompose the complex by reducing the arsenic and distilling the arsenous chloride (AsOl^). The most effective reducing agent found was titanous chloride (TiClg). Hydrogen sulfide reduces the compound and precipitates arsenous sulphide (As^S^) only under pressure and at high temp- erature. Sulfur dioxide has no effect. Zinc and hydrochloric acid reduces the arseno tungstate but wi th no evolution of arsine, (AsH^). Electrical reduction partly deposits metallic arsenic but does not liberate arsine. The accompanying sketch shov/s the type of apparatus used. The distillate should not come in contact with any rubber because of the presence of sulfur therein. F, s .3 jQea cQSh 5 Yea m C/ear drawn fletf AsCIs Distillation FtasR Ha Ccls B ottom of F\a, sM Q round Joint m imm - 6 - The results of the arsenic analysis, by precipitation, are probably low, as five precipitations were made to free it from tungsten. As previously stated there are only two references to the reactions of arseno tungstic acid with organic compounds and both of these refer to uric acid only. The present inves tigation v/as undertaken in the hope that the arsenic- tungsten combination would behave in the same manner as the silico- tungsten, phospho tongs ten, and phospho- molybdenum compounds. In order to test the arseno- tungstate, qualitative analyses were made upon quinine, quinidine, chincholine, and cinchonidine. Other organic nitrogen compounds were also used &nd the results recorded in the following table. Compoun d Formula Reaction Note: + reaction Quinine Quinidine + Chinchonine +■ Cinchonidine ■h £ Styryl 3 Benzoyl amino 4 tuinazolone /y H ♦ 1 ! 9 -CH* CK O "-NH-CGO c — 0 t . ■ -7- . ^ H Acetanilide Q N -CQGH^ p-Brom H N - COGH- A Acetanilide 0 Br p-Nitro ethyl COGK s * - C 2 H 5 Acetanilide 0 ho 2 Ethyl Acetanilide N - COCH„ 0' C 2 H 5 3 — Phenacetin (V° °2 E 5 V '~ HHOOOHg — Isobutyl Alcohol GH S- ° ^ CK-CE OH GH * 2 o — Secondary Butyl alcohol „OH CH 1 X CH 0 CH - GH B 2 3 — Pyafcidine o | 4 - Quinoline oo 4 - H p G 4 ° OH, O' m 'GE CH a 2 2 4- Piperidine . -8- Phenyl amino Propionic acid ' m 2. A- OH CH^ - ky - co h 2 — Trimethyl Amine (C W ~h p-Nitro ethyl /C mC 2 H 5 Ana line 0 »o s — Anisole y\ -QCH* o ■ — Phenyl amino Thiazole C S C - NHp H- C N + 2 Styryl 3 Phene tedyl 4 Quinazolone /V * C - CH=CH - H - o - OCgHg Anthraniiic Acid or - co 2 h — Uric Acid H - N - C =• 0 » / 0 = C C - NHx 1 H C=0 H - N - C - HH' — -9- 2 Styryl O N V N C -CH*CK - <3 N - UK - COCA* 4 ^uiiiazoxone "C 0 2 Styryl 3 Amino 4 Qninazolone GHrGK -< > 1!H 2 -h - 10 - After determining the extend of the possibilities in the qualitative precipitation of the alkaloids, the next step was the determination of the "factor" to be used in quantitative work. The "factor" is explained by the equation: Wt. WO^ x "factor" Wt. alkaloid. The general method for the determination of the "factor" follows: Dissolve the alkaloid in dilute sulphuric acid with a slight excess. Add an excess of the arseno tungstate sol- ution (l c.c. .01 gm. ) and heat to boiling, filter hot and wash thoroughly with hot water. Ignite the filter and precipitate in a weighed crucible at a low red heat and weigh as tungstic oxide (WOg). The "factors" for the four above mentioned aldaloids are:- Quinine .22565 Qu ini dine .30638 Cinchonine — -- .24268 Cinchonidine ----- — .26341 In each case the quantitativeness of the precipitation was tested by the standard methods of alkaloid extraction after decomp- osition of the precipitate by sodium or potassium hydroxide. The following table shows the results. Allcalc id Weight taken Weight recovered Quinine .01985 .02005 (Mean) Quinidine .02505 .02496 Cinchonine .02497 .02491 Cinchonidine .02772 .02764 - 11 - I&x-ait IIvlHLT TAL Preparation of the Ammonium arseno tungstate To 200 c.c. of saturated, solution of crystallized sodium tungstate (Na 2 *70 4 .2Hg0) ( approximately 85 grams^ add 80 c.c of f freshly prepared arsenic acid. (h^AsQ^) with constant stirring. The temperature of the solution will rise appreciably. Cool the mixture to 25° C. and add 40 gm. of ammonium chloride (NR^Cl) with stirring. Continue to stir rapidly for 15 minutes and allow the mixture i?o stand 24 hours. Pilter the mixture with suction and centrifuge the crystals until dry. The compound prepared in this way needs no further purification. Several trials were made using commercial arsenic acid ( 8 O/ 0 ) but a discolored product always resulted. The arsenic acid used was prepared as follows*.- To 500 c.c of concentrated nitric acid (C.P.) add arsenous oxide (AsgOgJin 10 gm. portions with stirring until no further reaction takes place. Pilter through an asbeston mat and use as soon as possible. The above reaction proceeds as follows :- 2H 0 As A 2HN0 2H AsO NO NO 2 2 0 O 0 4 2 ANALYSIS of the Salt Ignite a sample ( .2 - .5 gm. ) at the lowest possible red The residue is ifYO^. heat to content weight - 12 - Tun^sten Sample .1449 .1765 Crucible WO,, 5 10.0905 9.5952 ’ 1 along 9.9675 9.4461 WO* .125 .1491 84.8$ 84.5$ Pisco a sample ( .5 - 1.0 gm. ) in a distilling flask (See sketch) and add 100 c.c of hydrochloric acid (20 $) . Shake until the material is dissolved then add 50 c.c of titanous chloride solution (15$). Distil the solution in a current of cydrogen chloride. Neutralize the distillate with solid potash (KOH) using phenolphthalein as indicator. Just acidify with hydrochloric acid and again neutralize with sodium bicarbonate (NaliGO-) and add 5 gm. in excess. Titrate this solution with standard iodine using starch as an indicator. Arsenic Sample .5488 .1252 C.c. I soln( .1668) 1.15 .47 As 0 ($) 5.52$ 5.50$ The ammonia was liberated with sodium hydroxide and distill- ed in a Kjeldahl apparatus. Ammonia Sample 2.4058 C.c. acid( .1055) 25.70 C.c.alkali( .0971) .00 2.6769 57.0 7.62 5 • QO/b (hh 4 ) 2 o 2.9355a -13- From the above analysts the following ratios were obtained :- (1) AsgC^WOg 1:24.18 (2) (EE 4 ) 2 0:As 2 0 5 5:78:1 (3) H o 0:As 0_ 32.2:1 2 2 5 These ratios show the following formula for the ammonium arseno tungstate: 4(UE 4 ) 2 0.As 2 0 & .24W0^.52E 2 0. The specific gravity of the salt was determined by displacement of ligroin and was 5.343 referred to water. Determination of the "factors" Dissolve the alkaloid Salt in water (or just enough 10 $ sulfuric acid if the free alkaloid is used) and add 5 c.c. of 10; a sulfuric acid. To this solution add an excels of arseno- tungstate solution and heat to boiling. Filter hot and wash the precipitate thorougly Tilth boiling water. When the excess water has drained off place the wet filter in a weighed crucible and ignite very sl o wly . never allowing the paper to more than glow. Vi/hen the paper has been thoroughly charred ignite at a faint red heat to constant weight (WOg). The following results were obtained. Data on Factors quinine fuinine Hydrochloride .01S85 Dish and Residua — - — - — — 25*6411 Dish alone -- — --------------- 25.6250 Residue .0161 Factor .2256 .01985 22.4086 22.3920 .0166 I , , « . * -14- Qu ini dine Quinidine Sulphate Dish and Residue Dish alone Residue Factor .3064 .025035 10.1455 10.0565 .0890 .025035 9.5619 9.4925 .0894 C inchonine Chinchonine Hydrochloride .02497 Dish and Residue 9.5755 Dish alone 9.4828 Residue .0927 Factor .24268 .02497 9.5316 9.4390 .0926 Cinch o ni&ine Cinchonidine Hydrochloride .02772 Dish and Residue 9.5329 Dish alone 9.4394 Residue *0935 Factor .2634 .02772 9.6789 9.5852 .0937 Qualitative Tests on Organic Compounds The compounds were dissolved in a solvent common to the reagent and the compound and acidified with dilute sulfuric acid - 15- Fur the r St udies on Ars eno- tungs ti c Acid The ars eno tungstate when treated in aquous solution with ammonium oarhonate precipitates a white, crystalline product. This material was not analysed for arsenic but showed 83.7 % W0 3 . If the mixture of arsenic acid and sodium tungstate is sat- urated with ammonia (MHg) a white amorphous compound precipitates. It c 021 tains ab 09 . t 66 . 5>& WOg. The tungsten in the £41'70 3 compound is not precipitated with acids until the arsenic is removed. This indicates a stable complex. When electrolyzed so as to theoretically release the arsenic as arsine a solution of the ars eno tungstate turns blue but gives no arsine. When the same solution is treated in the same way but with the cathode in a porus cup metallic arsenic is partially precipitated. Sulphur dioxide will not reduce tie arsenic or tunsten. Hydrogen sulphide causes reduction only at 100° 0. , and under pressure; (a pressure flash was used). Iron salts cause a blue-green coloration in an acid solution of the sal t -16- SUMMARY of WORK. A rapid method for the preparation of an ammonium arseno tungstate has been devised. The salt has a definite composition, 4( .As^O^. 24 WO .32H fl>. The ammonium arseno tungstate precipitates certain definite alkaloids. Other organic nitrogen compounds are precipitated by the reagent, but only when apparently, the basicity reaches a definite point. A method has been devised whereby alkaloids may be quantitatively determined through the use of ammonium ar s eno tungs ta t e . Some properties of the ammonium arseno tungstate have been investigated and the results recorded. . • • * - • • ) ' . . ' . . <1 I • . . -17- B I BL I QGRAPHY * J.G •Drummond- Observations on the Phospho tungstates of Certain Bases and Amino Acids. Bioehem. J. IB, 5 - 24 (1918) 1. Stutser Untersuchungen ueber die quantitative Bestim- mung des Proteinstichstoff es und die Trennung der Pro teins toff e von anderen in Pflarijjen vorkommenden Sticks toffverbindungen. J. f. laudwirtjischoft 1880, 28, 103; 1881, 29, 473; 1886, 34, 151 2. Bertraud — Comptes Rendus 128 - 742 - 1899 Bull. Soc. Ghim. 2,434 (3) (1900) 3. Chapin — The determination of Nicotine in Nicotine Solutions and Tabacco Extracts. Bull. No. 133— U.S.Dep’t Ag. Bur. of Animal Husbandry. 4. Beal and Peterson - Silico Tungstic Acid as an Alkaloidal Reagent. Thesis - U. of Illinois 1920 5. Buck and Beal - Extraction of Alkaloids by Means of Immiscible Solvents. Thesis. U. of 111. 1921 6. S.R. Benedict - Uric Acid in Blood J. Biol. Ghem. LI - 1. 7. J.L. Morris and A.G.Macleod - Golorometric Deterimation of Uric acid in Blood. J. Biol. Chem. 1-1 8. Gibbs - Annalen 245, 1888, 50 9. Kehrman - Zt. Anorg. Ghem. 22, 1900, 290 • » .. •• . « -19- GMRRAL REh'ERWGlSS Graelin -ICraut Berichte Zt. Zryst* ii ii Rosooe and Schorlemer Ab egg 0. Dommer Ho ffinan- Lexicon Chem. Abst. 3/2- 594, 595, 602, 695 IV, 1834, 296 21, 1893, 313 22, 1893, 307 V. 2, p. 1061 3/3 V.2, p.616 V.15, Ho. 1, p.145