84 JOURNAL OF THE SOCIETY O17 COSMETIC CHEMISTS '• comparing the pen. etrating abilities of et•cient types of mercurial oint- ments. This method is so sensitive that if as little as 9 micrograms enter the body of a 250-gm. rat, 25 to 50 per Cent thereof can be detected in the kidneys in 24 hours. The procedure which we have used for assaying mercurial oint- ments may be summed up as fol- lows: A clipped area on the back is inuncted for 3 minutes with a mer- cury ointment, covered with rubber sheeting or celluloid, and contact of the ointment with the skin main- tained for 24:'hours. At the end of this time, the animal is sacrificed by exsanguination under ether an- esthetic, and the kidneys removed for analysis of mercury (5). All results are expressed in micrograms of mercury per gram of wet tissue. Owing to the natural variability in test animals, 6 to 8 are used for an assay in order to be able to recognize significant differences be- tween inunction procedures or oint- ments. Before discussing the findings of chief interest, namely, the effects of various types of ointment bases and mercury compounds on the cutaneous penetration of mercury, a number of factors relating espe- cially to the test a. nimals and the methods of application must be con- sidered briefly. L Effect of Covering the 2rnunction Site According to Rothman (6), cover- ing the skin results in interferences. with the normal escape of moisture.

STUDIES ON THE PERMEABILITY OF THE SKIN TO MERCURY. 85 Moisture thus accumulated may produce softening and maceration of the stratum corneum, a condition which may be favorable to the re- tention of substances in close con- tact with the skin, and possibly also to penetration. While in our assays the inuncted areas of skin were al- ways kept covered to avoid con- tamination errors, it seemed desir- able to test whether this technique affected skin penetration. The re- sults are 'shown in Table 2. It is clear that covering the inuncted area increases the absorption of mercury from every one of the six different ointments tested. ß TABLE 2--EFFECT oF COVERING THE IN- UNCTED SKIN AREA ON THE CUTANEOUS PENETRATION OF MERCURY IN THE RABBIT Storage of mercury in the kidney is used as a measure of penetration Micrograms Mercury per Gram Wet Kidney Calomel Covered Uncovered Ointment Skin* Skin* 1 26 7 2 45 13 3 17 4 20 9 5 6 4 6 22 5 -- Av. 23 8 * Average of two animals on each treat- ment. 2. Location of the site of Inunction Generally speaking the abdominal skin in the rat and rabbit is con- sidered thinner than the back skin. A comparison of the penetration of mercury at these two sites was, therefore, made and is shown in Table 3. It is to be noted that the site of inunction does not signifi- cantly affect absorption of mer- cury. ' 3. Size of the'Inunction Site In spite of the best precautions, seepage of the ointments sometimes occurred so that in effect a larger area of skin came into contact with the ointment than originally planned. In Table 4 are shown the results of deliberately halving the inunction areas which were rou- tinely 8 per cent of the total body surface. The amount of mercury stored in the kidney is not halved but reduced by roughly one-third. From these data it can be inferred that small changes in exposure area cannot seriously influence the out- come of the results. 4'. Effect of IFeight, Sex, and Litter The studies were generally con- ducted on groups of rats homogene- ous with respect to age, weight, and sex, but heterogeneous with respect to litter. As examples of three pos- sible variants, litter, sex, and weight were studied. It may be presumed that these variants could operate to produce differences between skin as characterized by such factors as texture, thickness, subcutaneous fat, number of hair follicles, etc., or' differences between organs, such as the a13ility of the kidney to excrete or store.mercury. .It was found, however, that the cutaneous pene- tration of mercury, as judged by the level in the 'kidneys, is not sig- nificantly affected by the above fac- tors.