PROTEINS OF EPIDERMIS, RELATION TO AGING SKIN 331 CHART III--PROCEDURE FOR THE EXTRACTIO•q OF THE PROTEINS FROM EPIDERMIS WITH A 2 M UREA SO•.UTION Beef snout epidermis (53 grams) I I. Extracted with 400 ml. 2 M urea 7 days at 0øC. I I--Extract (dialyzed)-] I I pH 5.5 P(A) pH 4.5 P(B) (50 mg.) (52 mg.) Residue beef snout epidermis I II. Extracted with 400 mi. 2 M urea 7 days at 0øC. I [ I Residue beef snout epidermis •-Extract (dialyzed)n pH 5.5 P(A) pH 4.5 P(B) III. Extracted with 400 mi. 2 M urea 14 days at 0øC. vExtract (dialyzed) Residue beef snout epider- I 1 mis (discarded) pH 5.5 P(A) pH 4.5 P(B) (19 mg.) (22 mg.) Proteins IIA and IIB lost. Proteins lB and IIIB purified as previously described (24). For the fibrous proteins (IA and IIIA) the last centrifugation was done in a Spinco Ultra- centrifuge at 130,050 X g for 5 hr. P -- protein. A Perkin-Elmer, Model 28 electrophoresis apparatus was employed using standard techniques (27, 28). Electrophoresis was carried out in phos- phate buffer, pH 7.0 (r/2, 0.20). ... ?•½.. ,.-[•..,..., .... ,.,., : -C• ? - ..•.. ß • -• •..:.• ,: '• ,• •%•.,,., ,: • .• •:•. •'.•.• ...... •.. •....? :- -.•.• Figure 1.•Beef snout epidermis, X 80. Note the various strata this tissue.

332 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS RESULTS AND DISCUSSION Beef snout epidermis because of its availability and thickness is a good source of material for studies on the structural proteins of epidermis. This tissue has all the strata of epidermis well defined and it is covered with a thick coat of keratin (Fig. 1). At high magnification the tonofibrils be- tween adjacent squamous cells are well delineated (Fig. 2). Beef snout skin left in 6 M urea solution at 0 ø for one hour showed tonofibrils in many cells, particularly in the outer Malpighian stratum, whereas this tissue left ........ ..,.. %,, ':.a, ...... •.', '•,,•, '• ?:':.,.:i-$, ".'%.: ....... 4. ß %'• '•1 ' .. "?. '.'"' •-•= ½ 7 ...xz, ..? . . ......... 4. :.4. ':..:½•r:' •.:o•k•½•,.: •t•. ' - [igure 2.•Beef snout epidermis, Malpighian layer, X 450. •ote the tono•bdls between adjacent cells. --•. -, .:•.• :• '-• •- • •. •v:'? .... 7•...• ....... ..• : '% • ..... . 4 •'• "• ..... .:•_ . .... • %... . .... .. ,, .•. .•... • .... • •. .... -• •--• '-L.- .2 ?' • • :•ø-:.. :."' -.-s-•- '•i:, ........ ' "_ . .., . . Figure &--Beef snout epidermis, X 450 which had been in a 6 M urea solution for St/,. hours at 0øC. Note empty spaces between some of the cells and pycnotic nuclei.