Simultaneous Determination Of Eight Sexual Hormones In Cosmetics By High Performance Liquid Chromatography With Diode Array And Fluorescence Detectors

Sex hormones(including estrogen, androgen and progesterone) is a steroid hormone, which have physiological functions of controlling accessory sex organs and secondary sexual characteristics. Short-term use of hormone-containing cosmetics increase skin elasticity, whiten skin and remove wrinkles, but long-term use can cause pigmentation, skin atrophy, human metabolic disorders and even cancer. In view of the problem of safety in cosmetics, corresponding standards and regulations have been carried out to regulate the adding of sexual hormones to cosmetics, and recommended testing methods of sex hormones in China《Hygienic Standard for Cosmetics》(2007 edition). Scientists are also actively seeking more rapid and sensitive method to satisfy the increasing sex hormones composition detection.The paper has chosen eight representative hormones(including estriol, estradiol, testosterone, ethinyl estradiol, diethylstilbestrol, megestrol acetate, progesterone and testosterone propionate) as research objects, considering the existing methods and the add probability of sex hormones in samples,systematically studied the pretreatment and the detection method. In the selection of pretreatment methods, solution and cream cosmetics have adopted different pretreatment methods. In the selection of detection methods, high performance liquid chromatography with diode array and fluorescence detectors was selected. A method of simple, rapid and sensitive detection of 8 sexual hormones in cosmetics by high performance liquid chromatography with diode array and fluorescence detectors has been developed, through the optimization of sample pretreatment condition and detection of HPLC.The research contents and results were as follows:1. In this paper, ultrasonic assisted extraction technology was the main method, and have established pretreatment method for solution and cream cosmetics. By investigating different extraction solvent and extraction time on the extraction effect of sex hormones, solution cosmetic samples extracted with methanol and the extract can be directly detected by HPLC, but for cream cosmetics, the extract was extracted with methanol need to be purified by solid phase extraction column before being detected by HPLC, and solid-phase extraction conditions have been optimized.Optimal solution cosmetics pretreatment conditions were as follows: the extraction solvent was methanol, ultrasonic extraction for 20 minutes; Optimal cream cosmetics pretreatment conditions were: sample dispersion with saturated sodium chloride solution,the extraction solvent was methanol, ultrasonic extraction for 20 minutes, and purified through Waters Oasis HLB column, leached with 30% methanol aqueous solution, abandoned to effluent, eluted with methanol and collected the eluent.2. In the study of liquid chromatographic conditions, this thesis examined influence of the chromatographic column, flow ratio and elution procedure, column temperature, flow rate, sample quantity and detection wavelength on the separation and response value degree of sex hormones, and established determination of eight sexual hormones in cosmetics by high performance liquid chromatography with diode array and fluorescence detectors. The optimal chromatographic conditions were: this thesis selected Agilent ZORBAX Eclipse XDB-C18 column for separation, acetonitrile and water as the mobile phase with gradient elution program.DAD detected in 196 nm, 242 nm and 288 nm wavelength at the same time. In order to avoid using a single DAD to be easy to produce the phenomenon of false positive, detected with FLD, the excitation wavelength was 280 nm and the emission wavelength was 310 nm. The retention time of chromatographic peak for qualitative and the external standard method for quantification, and realized the simultaneous determination of eight sex hormones within 20 minutes.The detection limits of this method were from 0.1μg/g to 0.5μg/g, the average recoveries ranged from 90.3% to 102.9%, and the relative standard deviation(RSD)were from 0.4% to 4.8%. The method is simple, rapid,sensitive and accurate. The method can be applied to the determination of eight sexual hormones in commercial cosmetics, and contribute to providing the scientific basis and technical support for the cosmetics hygiene supervision. The establishment of the method is of great significance for the relevant agencies to strengthen supervision of the quality of cosmetics and protect the health of consumers.