666 FEIGL AND JUNGREIS: SPOT TESTS FOR PHENOLS AND [Vol. 83 Spot Tests for Phenols and Alkylated Anilines Based on the Duff Formylation BY F. FEIGL AXD ES. JUNGREIS TRANSLATED BY R. E. OESPER (Laboratdrio da Produgdo Mzneral, Mznzstdrzo da Agrzcultura, Rzo de Janezro, B$ azzl) (Unzuerszty of Cznciizizatz, Czncznnatz, Ohzo) The formylation of phenols and dialkylanilines, as developed by Duff, has been extended to monoalkylanilines, and is accomplished by brief heating a t 150" to 160' C with a mixture of crystalline oxalic acid and hexamethylene- tetramine. The resulting o-hydroxyaldehydes and p-dialkylaminobenzalde- hydes or 9-monoalkylaminobenzaldehydes can be detected readily by either production of fluorescent aldazines with .hydrazine or formation of orange Schiff's bases with benzidine.These test:: for phenols or alkylated anilines come within the scope of spot-test analysis and have microanalytical sensitivity. DUFF^ has reported that satisfactory yields of o-hydroxyaldehydes and P-dialkylamino- benzaldehydes can be obtained by formylation of phenols and dialkylanilines withDec., 19581 ALKYLATED ANILINES BASED ON THE DUFF FORMYLATION 667 hexamethylenetetramine (hexamine). The procedures involve two partial reactions. In- itially, hexamine reacts in its trialkyl form,, and, when heated, participates in the following condensations- OH I OH I CH,--N=CH, + NH, . , 3 * (1) \=/- In fact, if hexamine is dry-heated with phenols or dialkylanilines, the evolution of ammonia from the reacting mass can be readily demonstrated by means of appropriate indicator-papers.In the second stage, the condensation products (I) are hydrolysed in their isomeric form (11) by dilute mineral acids to give the respective aldehydes according to reaction (3). + R I I)N<$I=N-CH3 + H,O + H+ -+ /-L-/- \ /-\ CHO+NH,-CH, (3) For the condensation reactions, Duff recommends conditions in which hydrolysis of the hexamine is kept a t a minimum or even avoided. This is achieved in reaction (1) by heating phenol and hexamine to 150" C for 15 minutes with a mixture of glycerol and boric acid that has previously been kept at 170" C for 30 minutes to remove water. Reaction (2) was accomplished by heating dialkylanilines and hexamine under reflux in a boiling-water bath for 5 hours with a mixture of glacial acetic acid and 90 per cent.formic acid. I t has been found that the formylations, i.e., reactions (l), ( 2 ) and (3), occur easily and readily when phenols or dialkylanilines are melted with a mixture of hexamine and oxalic acid dihydrate, although the latter, by virtue of its water of crystallisation, is known3 to bring about numerous hydrolyses, including the cleavage of hexamine by the following react ion- (CH,),N4 + 6H,O -+ 6CH,O + 4NH, I t appears, therefore, that reactions (1) and (2), as well as this hydrolysis, occur rapidly. I t should likewise be noted that equilibrium reactions are involved (which lead to extensive transformations of phenols and dialkylanilines) and that the reaction products are removed from the equilibrium mixture by the hydrated oxalic acid; ammonia by formation of am- monium oxalate, and condensation products by hydrolysis, as shown in reaction (3).This assumption is supported by the fact that microgram amounts yield the respective aldehydes after heating for 5 minutes at 150" C with oxalic acid and hexamine. Possibly the oxalic acid melt leads to improved yields of aldehydes, but this was not investigated, as our objective was to ascertain whether or not the Duff formylation can be used to detect phenols and alkylated anilines. Satisfactory results were achieved by fusing with oxalic acid, dissolving the reaction mixture in water and testing for aldehyde. The necessary operations are easily conducted by spot-test techniques. The tests described are additional examples to demonstrate one important point, namely, that suitably modified syntheses and methods of formation can be used as a basis for the detection of organic compounds and functional groups that take part in these changes4668 FEIGL AND JUNGREIS: SPOT TESTS FOR PHENOLS AND [Vol.83 DETECTION OF PHENOLS WITH FREE ortho-POsITIONs To detect o-hydroxyaldehydes formed in the Duff formylation, use is made of the im- mediate condensation of these compounds with hydrazine5 according to the following reaction- OH 013 OH I ,- I /-\ ' :-CH=N-N=CH--// +2H,O 2 Y-N-CHO + 2NH,-NH, -+ \=/ \=/ \=/ The aldazines formed in this reaction are resistant to dilute acids and show a yellow-green (or sometimes orange) fluorescence in ultra-violet light. When phenol and cresols undergo formylation, the resulting o-hydroxyaldehydes vaporise at the reaction temperature.Such aldehydes can be detected by holding a piece of filter- paper moistened with hydrazine sulphate solution over the melt. -4 stain, which exhibits yellow-green fluorescence, can be easily discerned even when no more than a trace of volatile aldehyde is produced. REAGEXTS- methylenetetramine. sodium acetate in 100ml of water. PROCEDCRE- add 1 drop of a solution of the sample in ethanol or diethyl ether. test-tube, and warm gently to remove the solvent. can be used, instead of its solution.) previously heated to 150" C. temperature for 1 to 2 minutes, and then remove the test-tube. hydrazine sulphate reagent solution to the reaction mass. shake the suspension, place it on a filter-paper, and dry for a short time, filter-paper in ultra-violet light. exhibit a blue-green, or sometimes orange, fluorescence.RESULTS- Use of the proposed procedure resulted in the detection of 0.25pg each of phmol, &-naphthol and resorcinol, 1 pg of 9-hydroxydiphenyl, 2 pg of salicylic acid, 5 pg of o-hydroxy- diphenyl and 15 pg of 2 : 7-dihydroxynaphthalene. No aldazine-fluorescence reaction was given by di-P-naphthol, naphthoresorcinol, 2 : 4-dinitroresorcinol or 1 : 8-dihydroxynaphtha- lene-1 : 3 : 6-trisulphonic acid. The fact that certain phenols, contrary to expectation, cannot be detected by the proposed procedure shows that no formylation occurred in the ortho-position, but this still leaves open the question of the possible introduction of a -CHO group meta or pava to the phenolic -OH group.To test this point, di-P-naphthol, naphthoresorcinol, 2 : 4-dinitro- resorcinol and 1 : 8-dihydroxynaphthalene-1 : 3 : 6-trisulphonic acid were heated to 150" C with hexamine. Ammonia was evolved, which indicates a condensation analogous to reaction (1). When phenols that show no aldazine-fluorescence reaction are melted with oxalic acid - hexamine mixture and the reaction mass is dissolved in water, the addition of benzidine produces a brown-yellow precipitate, which indicates the formation of a Schiff's base of the resulting aldehyde. These experiments show that a formylation has occurred, but, as yet, no information is at hand regarding its position and extent. It is remarkable that loss of ammonia can still be detected when 150 to 200 pg of these phenols are heated to 160°C with hexamine.Phenetole and acetylsalicylic acid were tested as representative of phenolic derivatives with free ortho-positions. The former gave no aldazine-fluorescence reaction, but the latter responded strongly (0.5 pg was detected). The response of acetylsalicylic acid is obviously due to the fact that, when heated with oxalic acid, hydrolytic splitting-off of acetic acid Oxalic acid - hexamine mixtw,e-Xx equal weights of crystalline oxalic acid and hexa- Hydrazzne sulphate reagent solution-Dissolve 10 g each of h ydrazine sulphate and The mixture should be freshly prepared before use. Place several centigrams of oxalic acid - hexamine mixture in a micro test-tube, and Nix the contents of the (If desired, a trace of the solid sample Place the test-tube in a glycerol bath that has been Raise the temperature of the bath to 160" C, maintain this When cool, add 1 drop of Add 1 drop of water, if necessary, Examine the If the sample contained phenols, the stains on the filter-paperDec., 19581 ALKYLATED ANILINES BASED ON THE DUFF FORMYLATION 669 occurs with production of salicylic acid.Other phenolic esters may be expected to behave analogously. The formylation test for phenols is impaired by the presence of aromatic amines and their N-alkyl derivatives, since, as shown by the positive aldazine-fluorescence response, o-hydroxyaldehydes may also be formed. A preliminary separation is therefore required. In many instances, this can be successfully accomplished by adding mineral acid or alkali hydroxide solution to the test solution and then extracting with diethyl ether; the aqueous layer will contain the salts of the amines or phenols.DETECTION OF MONO AXD DIALKYLANILINES The 9-dialkylaminobenzaldehydes produced by the Duff formylation can be detected by formation of orange Schiff’s bases with benzidine6 according to the following reaction- As in the test for phenols, there is no need to isolate the aldehyde after the fusion reaction; the test can be conducted on the aqueous solution of the oxalic acid melt. I t is noteworthy that monoalkylanilines (and even aniline itself) can be formylated by heating with oxalic acid - hexamine mixture. It appears that, during formylations, not only are -C,HO groups introduced in para-positions, but reactions also occur by which amines are converted to phenols.This is shown by the fact that addition of hydrazine salts to an aqueous solution of the oxalic acid melt produces the aldazine-fluorescence reaction of o-hydroxylaldehydes. PROCEDURE- The procedure is similar to that described for phenols, except that the cooled reaction mixture is treated with 1 to 2 drops of water, and the solution (or suspension) is placed on filter-paper impregnated with a solution of benzidine in diethyl ether. If the test is positive, an orange stain appears, the intensity of which depends on the amount of aromatic aldehyde formed. RESULTS- Use of the proposed procedure resulted in the detection of 2 pg each of dimethylaniline, diethylaniline and monomethylaniline and 3 pg of monoethylaniline. The test cannot be applied directly in the presence of either aldehydes that form coloured Schiff’s bases with benzidine or phenols that are formylated by hexamine.If such substances are present, the bases must be separated beforehand; this can be readily accomplished by ,dissolving the sample in acid and treating the solution with molybdophosphoric acid. The bases can then be recovered from the precipitate by adding alkali and extracting with diethyl ether. If mono and dialkylanilines are to be detected in the presence of aniline, the aniline should be converted to phenol by warming with nitrous acid (alkali nitrite plus hydrochloric acid). If the solution is then treated with alkali hydroxide solution and shaken with diethyl ether, the ethereal solution can be tested for mono and dialkylanilines by the proposed procedure. 1. 2. 3. 4. 5 . 6. REFERENCES Duff, J . C . , J . Chem. Soc., 1941, 547; 1945, 276. Losekann, G., Chem. Ztg., 1890, 14, 1409. Feigl, F., Jungreis, E., and Stark-Slayer, C., Anal. Chim. Acta, in the press. Feigl, F., “Spot Tests in Organic Analysis,” Fifth Edition, Elsevier Publishing Co., Amsterdam and Kew York, 1956, Chapter 1 . -, 09. cit., p. 185. Chakravarti, S. N., and Roy, M. B., Amzlyst, 1937, 62, 603. Received March 215t, 1958