.]. Soc. Cosmet. Chem., 36, 223-230 (May/June 1985) Determination of N-nitrosodiethanolamine (NDELA) in cosmetic ingredients MITCHELL D. ERICKSON, DUANE B. LAKINGS, ARBOR D. DRINKWINE, and JAMES L. SPIGARELLI, Midwest Research Institute, 425 Volker Boulevard, Kansas City, MO 64110. Received December 1 O, 1984. Presented in part at the Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy, Atlantic City, N J, March 10-14, 1980. Synopsis Methods for the extraction and cleanup of several cosmetic ingredient matrices are presented. Diethanoi- amine and triethanolamine matrices were diluted with ethyl acetate and dried with sodium sulfate. Amphoteric and quaternium matrices were slurried with silica gel and methanol and then evaporated to dryness. The dried sample was then transferred to a silica gel column and eluted with acetone/methylene chloride (60/40). Morpholine matrices were diluted with methanol/methylene chloride (2/98). All samples were analyzed by high performance liquid chromatography-thermal energy analyzer. Recovery corrections for losses during extraction and cleanup were based on the recovery of •4C-N-nitrosodiethanolamine mea- sured by scintillation counting. The methods were validated, showing an accuracy of 99% and a precision of + 22% over a range of about 50-1,000 ng/g. Detection limits are estimated to be below 50 ng/g for all matrices studied. INTRODUCTION N-Nitrosamines have long been recognized as a group of hazardous compounds about 80% of those tested have been found to be carcinogenic to test animals (1-4). Their carcinogenicity, mutagenicity, health effects, environmental presence, and occurrence in commercial products have been reviewed (1-11). In 1977, it was reported that N- nitrosodiethanolamine (NDELA) had been found in cutting fluids (12), tobacco (13), cosmetics (14), lotions (14), and shampoos (14). Subsequently, NDELA was shown to be a carcinogen in the rat (15). Since these reports, scientists in industry and govern- ment have been working to develop and improve upon analytical methods for detecting NDELA (16). NDELA can be formed in cosmetics or their ingredients as a reaction product of alkanolamines such as triethanolamine, alkanolamides, and other amino compounds used in cosmetic formulations with a nitrosating agent, possibly 2-bromo- 2-nitro- 1,3-propanediol ( 17, 18). Since prevention of human exposure to NDELA is of paramount interest, determination of NDELA in cosmetics and their ingredients is necessary for adequate consumer pro- tection. Although the procedure used by Fan et al. (14) in the original work was apparently adequate for certain matrices, it has been reported (18) to be unreliable, especially with aqueous matrices. 223

224 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS As part of an overall program to develop methods for detecting NDELA in cosmetic matrices (16,19), the work reported here was designed to develop and validate methods for the extraction and cleanup of NDELA in several specific cosmetic ingredient matrices. EXPERIMENTAL APPARATUS The isocratic high performance liquid chromatography (HPLC) system consisted of a single Waters Associates (Milford, Mass.) Model 6000A pump, Waters Model U6K injector, Carle Instruments (Anaheim, Calif.) Model 2025 four-way micro-switching valve, Waters Model 440 UV detector operated at 254 nm, and a Thermo Electron Corporation (Waltham, Mass.) Thermal Energy Analyzer (TEA) Model 502. The UV and TEA detectors were used in series, the UV used first. The TEA was operated at a furnace temperature of 550øC, argon and oxygen pressures were 0.5 torr, photomul- tiplier voltage was 1475 V DC, and the cold trap was dry ice/isopropanol. The columns were Waters p•Bondapak CN (300 x 4 mm ID) a Laboratory Data Control (Riviera Beach, Fla.) 5 p•m Spherisorb CN (250 x 4.6 mm ID) with a Whatman (Clifton, N.Y.) Co:Pell octadecyl silane guard column and a Laboratory Data Control 5 •m Spherisorb Silica (250 x 4.6 mm ID) column. Unless otherwise specified, flow rates were 2 ml/min isooctane/methylene chloride/methanol (75/19/6 volumetric ratios are used throughout this paper). The micro-switching valve was placed in the system between the UV detector and the TEA detector to permit the analyst to switch the column eluent from the TEA detector to waste when desired. Radioactive decay rates were obtained using a Packard Tri-Carb liquid scintillation counter (Packard Instrument Company, Downers Grove, Ill.). REAGENTS All solvents were HPLC "distilled in glass" grade (Burdick and Jackson, Muskegon, Mich.) and filtered prior to use in the HPLC system. The •4C-NDELA was obtained from Dohm Products, Inc. (North Hollywood, Calif.) Cosmetic ingredient matrices were supplied by the sponsor (see Acknowledgements). RECOVERY CORRECTION USING •4C-NDELA Duplicate weighed aliquots of each sample were spiked with 200 p•l of 130 ng/ml 14C- NDELA (10 mCi/mmol) in 4% methanol/methylene chloride. Following all extraction and cleanup steps, a quantitative aliquot (generally 1.0 ml of 5) was transferred to a scintillation vial containing 15 ml Aquasol universal cocktail and counted for 10 min (or adequate time to get sufficiently precise counting statistics) using a liquid scintil- lation counter. Appropriate blanks and controls (aliquots of the original •4C-NDELA spiking solution) were also counted with the samples for background correction and calculation of percent recovery. PROCEDURE HPLC-TEA a,alysis. All extracts were analyzed by isocratic HPLC-TEA using isooc- tane/methylene chloride/methanol (75/19/6 v/v/v) on a 5 p• Spherisorb CN column.