PYRIDOXINE-3,4-DIACYLATES IN COSMETICS 351 pyridoxine tripalmitate. He concluded from a series of experiments that pyridoxine tripalmitate is very effective in keeping the human skin healthy and beautiful. The large-scale preparation of pyridoxine tripalmitate is not easy, nor is this compound satisfactory for cosmetic use because of its insufficient fat-solubility. A new commercial method for the synthesis of heat- and light-stable and fat-soluble derivatives of pyridoxine, pyridoxine-3,4-diacylates, has been developed in this laboratory. Patents for this method of prepara- tion are now pending in the United States of America, Great Britain, France, Switzerland, West Germany, and Japan. The chemical and physical properties, biological activity, toxicity and effects on the skin of these diacylates have been examined. It ap- pears that the higher fatty acid diesters are heat- and light-stable, fat- soluble and hydrolyzed into free pyridoxine in vivo and show the biologi- cal activity of vitamin B6. The present report deals mainly with the dibutyrate, dioctanoate, dilaurate, and dipalmitate of pyridoxine. EXPERIMENTAL AND DISCUSSION Materials All the diesters used were white crystalline powders, except the di-iso-octanoate. Some of the properties of these diesters are tabulated in Tables I, II, and III. Heat Stability Two grams of pyridoxine-3,4-dipalmitate was dissolved in 10 g. of olive oil and heated for six hours at 150-160øC. At the end of this heating period, the oil was brown. The oil was then diluted with 100 ml. of petroleum ether and cooled in the refrigerator for two days. The precipitate was removed by filtration and washed with petroleum ether. The crystalline precipitate was pure white and weighed 1.6 g. The crystals showed no depression in melting point when mixed with au- thentic pyridoxine-3,4-dipalmitate. The dilaurate and dioctanoate were tested in a similar manner, and the recovered unchanged esters weighed 1.4 and 1.1 g. respectively. The dibutyrate, however, could not be recovered from the heated brown oil, presumably because most of the diester was destroyed by heat. Light Stability One gram of each of the pyridoxine derivatives was dissolved in 100 g. of 50% aqueous ethyl alcohol. Samples of these solutions were

352 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS TABLE V LDs0 of Pyridoxine-3,4-dipalmitate and Related Compounds (Mice) LDs0, g./kg. Compound Administration 24 Hr. 7 Days Pyridoxin e-3,4-dipahnitate Pyridoxine hydrochloride Oral 7.1 5.1 Subcutaneous 4.4 1.6 Oral 3.8 3.8 Subcutaneous 1.7 1.4 Palmitic acid Oral 5.0 4.2 Subcutaneous . . . 4.2 sealed into test tubes and placed into sunlight. The transmittance of the irradiated solutions was read at 435 mu with a spectrophotometer. The data shown in Table IV indicate that the esters are not severely discolored during long exposure to sunlight and are more stable than pyridoxine hydrochloride. Toxicity * Acute Toxicity The acute toxicity of pyridoxine-3,4-dipalmitate was studied in mice and is recorded as LD•0 in Table V. l?yridoxine hydrochloride and palmitic acid were also tested as control materials. The ester and palmitic acid were dissolved in ethanol and then mixed with propylene glycol. Pyridoxine hydrochloride was studied in an aqueous solution. The mixture and the solution of samples were administered orally and subcutaneously to mice. From the results shown in Table V, it can be concluded that pyr- idoxine-3,4-dipalmitate is less toxic than pyridoxine hydrochloride. Chronic Toxicity Six groups each of five male and five female rats weighing from 85 to 135 g. were used in this test and fed for six months as follows' Group A' Group B' Group C' Standard Diet Standard Diet + 40 mg./kg./day of pyridoxine-HC1 Standard Diet + ,50 mg./kg./day of palmitic acid * This work was conducted at the Osaka City Institute of Hygiene (6).