Qin Si, Lv Chenghao, Wang Qingshan, Zheng Zhibing, Sun Xi, Tang Minyi, Deng Fangming
Key Laboratory for Food Science and Biotechnology of Hunan Province, College of Food Science and Technology, Hunan Agricultural University, Changsha 410128, China.
Hunan Co-Innovation Center for Utilization of Botanical Functional Ingredients, Hunan Agricultural University, Changsha 410128, China.
Anim Nutr. 2018 Sep;4(3):281-287. doi: 10.1016/j.aninu.2018.05.005. Epub 2018 Jun 20.
Limonin, the main bioactive phytochemical constituent of limonoids with multi-functions, is enriched in citrus fruits and often found at a high concentration in citrus seeds. The present study was attempted to introduce a new and efficient extraction method to isolate limonoids from pummelo seeds, and to evaluate the antioxidant property of the main constituent limonin in HepG2 cells. Three key single factors were identified for the extraction of limonoids from pummelo seeds using the Box-Behnken experiment design of response surface methodology (RSM), and the optimized extraction parameters were treatment with 89.68 mL of anhydrous acetone for 4.62 h at 78.94 °C, while the yield of limonoids was 11.52 mg/g. The structure of isolated main constituent of the limonoids was further identified as limonin by Fourier transform infrared (FT-IR) spectrometer and nuclear magnetic resonance (NMR) spectrum. Moreover, the molecular data in HepG2 cells revealed that limonin exerted its anti-oxidant property mainly by the activation of nuclear factor (erythroid-2)-like 2 (Nrf2)/kelch-like ECH-associated protein 1 (Keap1)- antioxidant response element (ARE) pathway in the form of transcriptional regulation of mRNA and posttranscriptional regulation of Nrf2/Keap1 system. These results demonstrate that pummelo seeds are an ideal source of limonoids, and limonin is proved to exert its anti-oxidant property by the activation of Nrf2/Keap1 pathway.
柠檬苦素是具有多种功能的柠檬苦素类化合物的主要生物活性植物化学成分，富含于柑橘类水果中，且在柑橘种子中含量通常较高。本研究试图引入一种新的高效提取方法，从柚子种子中分离柠檬苦素类化合物，并评估主要成分柠檬苦素在肝癌细胞系HepG2中的抗氧化特性。采用响应面法（RSM）的Box-Behnken实验设计，确定了从柚子种子中提取柠檬苦素类化合物的三个关键单因素，优化后的提取参数为：在78.94℃下用89.68 mL无水丙酮处理4.62 h，此时柠檬苦素类化合物的得率为11.52 mg/g。通过傅里叶变换红外光谱仪（FT-IR）和核磁共振光谱（NMR）进一步鉴定了分离出的柠檬苦素类化合物主要成分的结构为柠檬苦素。此外，HepG2细胞中的分子数据显示，柠檬苦素主要通过激活核因子（红系衍生2）样2（Nrf2）/kelch样ECH相关蛋白1（Keap1）-抗氧化反应元件（ARE）途径，以mRNA转录调控和Nrf2/Keap1系统的转录后调控的形式发挥其抗氧化特性。这些结果表明，柚子种子是柠檬苦素类化合物的理想来源，并且证明柠檬苦素通过激活Nrf2/Keap1途径发挥其抗氧化特性。
