Effects of Fatty Acids and Catechins on Sebum Production in a Human Model
Topical antiandrogenic activity of several fatty acid and catechin was first evaluated in the hamster flank organ assay or the rat assay. To further confirm the effectiveness of antiandrogenic compounds and suitability for human use, tests were performed on a human male subject. The ideal compounds for human treatment are those that are topically and locally active but do not show systemic antiandrogenic activity, especially in the cases involving young males. In the following example, two classes of compounds were tested by measuring sebum secretion from the forehead of an adult male treated topically.
A. METHODS
1. Determination of Forehead Sebum Production
A 63-year old Asian male volunteer was used to test and analyze sebum production from the forehead region. The forehead was washed thoroughly by soap twice and cleaned by 70% isopropyl alcohol twice. Sebum production was measured 30 to 60 minutes later by a sebum meter (Courage/Khazaka Electronic GmbH, Germany). The sebum meter tape probe (7 mm.times.8 mm) covered 56 mm.sup.2 area in each measurement. Ten measurements were made within the 4 cm square area (16 cm.sup.2) located at the middle of the left or right side forehead between the eyebrow and the hair line.
The sebum meter detected the difference in the transparency of the tape before and after the tape was placed on the forehead for 30 seconds and expressed the difference in an arbitrary number (S-value) between 0 to 300 (or higher). S-values of sebum accumulated on the foreheads of men are usually 200 to 300. Skin surface on hands usually showed a very low number (5 to 20). The S-value for forehead immediately after washing was less than 5. For men, the S-value gradually increased to about 50 within 30 minutes after washing and reached 100 to 200 in 45 minutes to 55 minutes.
To determine the rate of sebum production, the left and the right forehead areas were measured alternatively and each time at the comparable areas on the two sides. Ten measurements on each side (i.e., 20 measurements for two sides) could take about 15-20 minutes and the sebum-values ranged between 30 to 200. The S-values were different considerably at different areas of the forehead and could be influenced by environmental, including weather, diet, and physiological conditions. However, the ratio of the total S-value (the sum of 10 measurements) for the left and the total S-value for the right forehead was constant. For the Asian male tested in this experiment, the L/R ratios measured over a six month's period was within 1.15 to 1.38 if the S-values were determined 30 to 50 minutes after the forehead was washed thoroughly. Therefore, compounds applied to the left forehead that reduced the L/R ratio to lower than 1.1 were considered as topically active agents for suppression of sebum production.
B. RESULTS
1. .gamma.-LA Inhibition of Human Forehead Sebum
In the experiment shown in FIG. 24, 0.2 ml of borage oil (containing 18% of .gamma.LA) in a gel capsule was applied to the left forehead twice daily for 23 days. During this period, L/R ratio reduced from 1.28.+-.0.03 down to 1.05.+-.0.01. After the borage oil treatment was stopped, the L/R ratio returned to 1.20.+-.0.14. The effect of borage oil on the sebum production on the left forehead was relatively small, possibly due to the fact that most .gamma.LA in the borage oil was in the form of triglyceride that did not inhibit 5.alpha.-reductase (see Table 2). Free acid released from the glyceride by nonenzymic or enzymic action was probably responsible for the effect.
After the borage oil application was stopped and the L/R ratio recovered to 1.33, 20 mg of pure .gamma.-LA was applied to the left forehead twice each day for 6 days. The L/R ratio decreased to 0.22 during this period. After the .gamma.-LA application was stopped, the L/R ratio recovered slowly to 1.20.+-.0.14 over the period of 16 days. The finding clearly showed that .gamma.-LA was superior than borage oil in quickly suppressing sebum production from forehead of a human male subject.
2. Catechin Inhibition of Human Forehead Sebum Production
Twenty mg of (-)epigallocatechin gallate (EGCG) in 0.2 ml 70% ethanol was applied to the left forehead twice a day for 6 days (FIG. 25). The L/R ratio decrease from 1.20.+-.0.02 to 0.71.+-.0.04 during this period. After the EGCG treatment was stopped, the L/R ratio gradually recovered to 1.19.+-.0.02 within 16 days. Subsequent treatment of the left forehead with 20 mg (-)epicatechin in 0.2 ml 70% ethanol twice a day for 6 days reduced the L/R ratio to 1.02.+-.0.1. After the application was stopped, the L/R ratio gradually increased to the normal value of 1.21.+-.0.01 in 10 days. Clearly EGCG was more effective in reducing the sebum production from forehead than (-)epicatechin.
That contrasts with roughly ~30% or so inhibitions of T-stimulated flank-organ growth by spironolactone in other typical hamster studies.