Bianchi Sara E, Teixeira Helder F, Kaiser Samuel, Ortega George G, Schneider Paulo Henrique, Bassani Valquiria L
Programa de Pós-Graduação em Ciências Farmacêuticas, Faculdade de Farmácia, Universidade Federal do Rio Grande do Sul, Av. Ipiranga 2752, CEP 90610-000, Porto Alegre, RS, Brazil.
Programa de Pós-Graduação em Química, Instituto de Química-Universidade Federal do Rio Grande do Sul, Av. Bento Gonçalves 9500, CEP 91501-970, Porto Alegre, RS, Brazil.
J Chromatogr B Analyt Technol Biomed Life Sci. 2016 May 1;1020:43-52. doi: 10.1016/j.jchromb.2016.03.012. Epub 2016 Mar 15.
Coumestrol is present in several species of the Fabaceae family widely distributed in plants. The estrogenic and antioxidant activities of this molecule show its potential as skin anti-aging agent. These characteristics reveal the interest in developing analytical methodology for permeation studies, as well as to know the stability of coumestrol identifying the major degradation products. Thus, the present study was designed, first, to develop and validate a versatile liquid chromatography (HPLC) method to quantify coumestrol in a hydrogel formulation in different porcine skin layers (stratum corneum, epidermis, and dermis) in permeation tests. In the stability-indicating test coumestrol samples were exposed to stress conditions: temperature, UVC light, oxidative, acid and alkaline media. The degradation products, as well as the constituents extracted from the hydrogel, adhesive tape or skin were not eluted in the retention time of the coumestrol. Hence, the HPLC method showed to be versatile, specific, accurate, precise and robust showing excellent performance for quantifying coumestrol in complex matrices involving skin permeation studies. Coumestrol recovery from porcine ear skin was found to be in the range of 97.07-107.28 μg/mL; the intra-day precision (repeatability) and intermediate precision (inter-day precision), respectively lower than 4.71% and 2.09%. The analysis using ultra-performance liquid chromatography coupled to a quadrupole time-of-flight high definition mass spectrometry detector (UPLC-QTOF/HDMS) suggest the MS fragmentation patterns and the chemical structure of the main degradation products. These results represent new and relevant findings for the development of coumestrol pharmaceutical and cosmetic products.
香豆雌酚存在于豆科植物的多个物种中，广泛分布于植物界。该分子的雌激素活性和抗氧化活性表明其具有作为皮肤抗老化剂的潜力。这些特性显示出开发用于渗透研究的分析方法以及了解香豆雌酚稳定性并鉴定主要降解产物的重要性。因此，本研究首先旨在开发并验证一种通用的液相色谱法（HPLC），用于在渗透试验中对不同猪皮肤层（角质层、表皮和真皮）的水凝胶制剂中的香豆雌酚进行定量。在稳定性指示试验中，香豆雌酚样品被置于应激条件下：温度、紫外线C光、氧化、酸性和碱性介质。降解产物以及从水凝胶、胶带或皮肤中提取的成分在香豆雌酚的保留时间内未被洗脱。因此，HPLC方法显示出通用性、特异性、准确性、精密度和稳健性，在涉及皮肤渗透研究的复杂基质中对香豆雌酚进行定量时表现出色。从猪耳皮肤中回收的香豆雌酚浓度范围为97.07 - 107.28μg/mL；日内精密度（重复性）和中间精密度（日间精密度）分别低于4.71%和2.09%。使用超高效液相色谱与四极杆飞行时间高分辨质谱检测器联用（UPLC-QTOF/HDMS）进行的分析表明了主要降解产物的质谱裂解模式和化学结构。这些结果为香豆雌酚药物和化妆品的开发提供了新的相关发现。
