Detection of Small Amounts of Phenolphthalein In the Presence of Emodin and of Chrysophanic Acid EUGEiX1.A H. bIAECHLING. College of Physicians and Surgeons. Columbia University, New York, S . 1..
HEN plienoIplitIialeiii is the main active principle in medicinal preparations, its detection and determination are as a rule readily accomplished (1, I S ) . Khen, however, i t is accompanied by certain laxative plant products, such as cascara, aloes, rhubarb, senna, or frangula, which contain polyhydroxyanthraquinones, its detection may offer considerable difficulties due to the similarity of their chemical behavior (3, 7 ) . These anthraquinone derivatives-namely, frangula-emodin (also present in cascara sagrada and in rhubarb), aloeemodin (also present in senna), and particularly chrysophanic acid (chrysophanol, @-show in alkaline solution a variety of rather stable colors (18, 19), from cherry-red to purplish red, sometimes indistinguishable froin that of phenolphthalein, the presence of which may thus be obscured. On acidification such solutions turn yellow or yellox-ish bron 11 ; phenolphthalein is colorless under these condition.. The similarity of color reactions, including those given with concentrated sulfuric acid (11, 16), and the difficulty in separating the anthraquinone derivatives from phenolphthalein by means of organic solvents (19),made desirable an analytical method which would be characteristic for phenolphthalein and negative for the polyhydroxyanthraquinones in question, and thus permit with certainty the identification of phenolphthalein in some medicinal preparations (4). It was found that the purplish red solution of phenolphthalein in potassium hydroxide when treated n i t h an excess of hydrogen peroxide undergoes a gradual decoloration, resulting from oxidative cleavage. When the colorless alkaline solution is cooled, acidified n i t h dilute sulfuric acid, and extracted with ether, phthalic acid can be isolated from the ethereal extract and identified by its melting point and its ability to yield fluorescein when heated with resorcinol. When the oxidation by hydrogen peroxide in akaline solution was carried out upon certain plant products such as rhubarb powder, cascara sagrada, and polvdered aloes, to which known amounts of phenolphthalein had been added, oxidation products were obtained which also gave fluorescent solutions on heating x i t h resorcinol under suitable conditions. Since some organic acids like oxalic, malic, citric, and succinic respond in this way, all materials to be analyzed must uniformly first be treated with an excess of sodium bicarbonate golution, and the phenolphthalein extracted with ether. This procedure not only achieves the separation of phenolphthalein and the polyhydroxyanthraquinones from the organic acids and other mater-soluble substances, including some anthraglucosides possibly present, but may a t the same time reveal the absence of emodin and of chrysophanic acid in winples from which the ethereal extracts prove to be coloilees. Oxidation experiments carried out upon authentic samples of rhubarb, senna, aloes, and cascara sagrada unmixed n ith phenolphthalein gave uniformly negative rewlts in the test for phthalic acid. Experiments with commercial chrysarobin powder were also negative (5, 6, 17). The absence of phthalic acid in these oxidatioii products can well be explained by the following considerations: The anthraquinone derivatives-namely, frangula-emodin, 1,8,6trihydroxy-3-methylanthraquinone, aloe-emodin, 1,s-dihydroxy-3-oxymethylanthraquinone, and chrysophanic acid, 1,stiihydroxy-3-methylanthraquinone-occurring in the abovementioned plant product- aye related to chiysazin (1,s386
dihydroxyanthraquinone, 10, l a ) , which on oxidation does not yield phthalic acid (8). Chrysaain (Istizin) has also laxative propert,ies.
Procedure A suspension of 0.5 t o 1 gram of the powdered sample in 20 cc. of a freshly prepared solution of sodium bicarbonate is shaken in a separatory funnel three times lvith ether, using 25 cc. for the first and 15 cc. for each subsequent extraction. The combined ethereal extracts are Jyaahed and evaporated to dryness. The residue, containing phenolphthalein and the anthraquinone derivatives originally present, is dissolved in 20 cc. of 10 per cent potassium hydroxide and the red to purple solution is oxidized by successive additions of 30 per cent hydrogen peroxide, preferably in a beaker or a porcelain casserole, heating gently over a low flame. An Erlenmeyer flask should not be used for this process, because of foaming and a tendency of the alkaline tolution to creep. The solution is allowed t o ,settle in the cold. A purplish sediment of alkali salts of anthraquinone derivatives (3, 1,5), if present, is removed, and the alkaline solution is cooled with ice and acidified with an excess of dilute sulfuric acid. Effervescence and a change of color to colorles place. The solution is extracted with three succe ether, using 20, 15, and 15 cc., respectively. The combined ethereal extracts are washed and evaporated first in a casserole t o a small volume at room temperature and finally to dryness in a Pyrex tube surrounded xith vater at about 60" C. The residue in the test tube is heated with an excess of resorcinol in a metal bath at 180" to 200" C. for 20 minutes, or at boiling temperature for 5 minutes over a small flame. No condensing agent is used (9, 14. The resulting melt is dissolved in about 5 cc. of potassium hydroxide and diluted with water. According to the quantity of phenolphthalein present in the original sample, a more or less pronounced green fluorescence will be observed.
The presence of 5 to 10 gamma of phenolphthalein in the original mixture can be detected by this procedure. Cascara sagrada tablet's to which succinic acid and oxalic acid had .been added gare negative fluoiescence bests.
Literature Cited .Lssoc. Official dgr. Chem., Official and Tentatii-e Methods o f rinalysis, 4th ed., p. 569, 1935. Bailey, E. bf., J. IND. EXG.CHEY., 6, 320 (1914). Beal, G. H., and Okey, R., J . Am. Chem. Soc., 39, 716 (191ij. Belote, G. H., and Whitney. H.. :Irch. Dermafol. Syphilol., 36, 279 (1937). Eder, R.. Arch. Pharm., 253, 1 (1915. Hesse, O., Ann., 388, 65 (1912). Hubbard, W.S.,J. IND. ESG. CHEM., 9, 518 (191T). Liebermann, C., Ann., 183, 201 (1876). hfeyer, Hans, "hnalyse und Konstitutionsermittelung organischer \-erbindungcn," 5th ed., p. 414, Berlin, Julius Springer, 1931. Xaylor, C. A, Kith Gardner. J . , J . Ana. Chem. Soc.. 53, 41011 (1931j. Oesterle, 0.A., Arch. Pharm., 237, 82 (1899). Ibid., 250, 301 (1912); 253, 335 (1915). Palkin, S., J. IXD.ENG.C H m z . , 12, 766 (1920). Paul, L., Z . angew. Chem., 24, 1867 (1911). Peel, E,, Arch. Pharm., 257, 254 (1919). Tschirch, d.,and Heuberger, K., Ibid.. 240, 808 (1902). Tutin, F., and Clewer, H. IT. B., J . Chem. Soc., 99, 946 (1911); 101, 290 (1912). Warren, L. E., d m . J . Pharrn., 86, 414 (1914). Westman, L. E., and Rowat, R. M.,J . Am. Phurm. .4ssoc., 7 , 759 (1918). RECEIVED 3Iay 27, 1036.
