100 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS are negative. In the Thymol Blue test, a neutral, dilute aqueous solution of the surfactant is treated with an acidic solution of Thymol Blue. If an anionic surfactant is present, the yellow color of the indicator solution changes to reddish-purple. The second type of test for ionic surface active agents, the Dye Transfer test, is based upon the reaction of an ionic surfactant with a water-soluble, so]vent-insoluble dyestuff of oppositely charged type to produce a water- insoluble, solvent-soluble surfactant-dyest•ff complex. R-Na* + DN*C1- -,- DN*R- + Na'C1 - (II1) Anionic Water-Soluble, Water-Insoluble, Surfactant Solvent-Insoluble Solvent-Soluble Cationic Dyestuff Surfactant-Dyestuff Complex RN*C1- + D-Na + --,- RN*D- + Na'•C1- (IV) Cationic Water-Soluble, Water-Insoluble, Surfactant Solvent-Insoluble Solvent-Soluble Anionic Dyestuff Surfactant-Dyestuff Complex Weatherburn (5) has designed an ingenious test of this type which is capable of detecting both anionics and cationics, based upon the observa- tion that the surfactant-dyestuff complex formed by the reaction of an anionic surfactant with a cationic dyestuff, is less stable than the complex formed by the reaction of an anionic surfactant with a cationic surfactant. Thus, RN*C1- q- DN*R- -• RN+R - q-- DN*C1- (V) Cationic Water-Insoluble, Water-Insoluble, Water-Soluble, Surfactant Solvent-Soluble Anionic-Cationic Cationic Surfactant- Complex Dyestuff Dyestuff Complex (Colorless) (Colored) In his test, a few milliliters of a solution of the water-soluble, chloroform- insoluble, cationic dyestuff, Methylene Blue, are added to a few milliliters of immiscible chloroform, and the mixture treated dropwise with a solution of a known anionic surfactant until both the water and the chloroform layers are equally blue in color. That is, the blue color of the water layer due to the presence in it of the water-soluble Methylene Blue equals the blue color of the chloroform layer due to the presence in it of the surfactant-dyestuff complex. This blue-layered mixture is then treated with a few milliliters of a dilute aqueous solution of the unknown surfactant, shaken well, and the layers allowed to separate. If the chloroform (lower) layer is now bluet than the aqueous layer, the unknown is an anionic surfactant (equation III, above) if the water layer is bluet than the chloroform layer, the un- known is a cationic surfactant (equation V) if both layers are more or less the same color, it is nonionic. Of course, the layers will also be the same color if no surfactant is present at all.

IDENTIFICATION OF SURFACE ACTIVE AGENTS 101 Tests for the detection of nonionic surfactants are not as well developed as those fi)r ionic surface active agents. Of course, the presence of this type of surfactant may be deduced from a positive result obtained in the dye solubilization test of Hoyt, mentioned before, followed by negative results in the tests for ionic surfactants. However, this negative type of evidence is always inconclusive and, moreover, would not detect a nonionic admixed with an ionic surfactant. As a result, a goodly number of tests have been proposed for the detection of nonionic surface active agents. Unfortunately, none of these tests are general enough to detect all types of nonionic surfactants, and almost all are subject to interference by both cationic and anionic surfactants. Because of their limited utility they will, with one exception, not be covered in this paper. The one test for detecting nonionic surfactants which is relatively free from interferences is the so-called Heat test or Cloud test (6). This test is not a general one for nonionics, but will only detect nonionic ethylene oxide derivatives. It is based on the well-known fact that nonionic ethylene oxide derivatives become less soluble in water as the temperature is in- creased. The reason for this reverse solubility is believed to be the break- ing of the hydrogen bonds between the water and surfactant molecules as the temperature increases. Since these hydrogen bonds are probably the main factor in causing the surfactant to be soluble in water, as they are broken, the solubility of the surfactant in water decreases. The test, it- self, is rather simple. A dilute aqueous solution of the unknown surfactant is heated gently in a beaker and observed for the appearance of any cloud- ing. If clouding appears, the solution is cooled to observe if it clears upon cooling. If it does, the presence of a nonionic ethylene oxide derivative is indicated. If no cloudiness appears, even at the boiling point, the solution is cooled, and sufl:icient salt is added to make a 10 per cent salt solution, and the test repeated. Materials which are not clearly soluble in water at room temperature may be tested in aqueous alcohol. ISOLATION If a surt'ace active agent has been detected in a composition, and it is desired to obtain further information concerning its nature, it is almost always desirable, if not imperative, to isolate the surfactant before pro- ceeding any further with its analysis. The type of composition in which surfactants appear are now so numerous that it is almost impossible to get reliable qualitative or quantitative test results unless all the other substances present in the composition are known and the analytical meth- ods used are specially selected or modified to avoid interference from the other materials present in the composition. In the usual case where only a limited amount of information concerning the nonsurfactant components is available, isolation of the surfactant is therefore almost always a necessary