Abstract:To investigate the changes in the mass fraction of flavonoids in the germination process of black wheat, a high-performance liquid chromatography (HPLC) method was established for simultaneous determination of 10 flavonoids in germinated black wheat. An Agilent 1260 ZORBAX SB-C18 column (4.6 mm×250 mm×5 μm) was used as the stationary phase, with a gradient elution of acetonitrile and 0.8% phosphoric acid aqueous solution as the mobile phase. The flow rate was set at 0.8 mL/min, with a detection wavelength of 225 nm and the column temperature maintained at 25 ℃. The results showed that the mass concentration of 10 flavonoids, including puerarin, epicatechin, rutin, luteolin, quercetin, apigenin, naringenin, kaempferol, isorhamnetin, and chalcone, exhibited a good linear relationship with peak area (R2≥0.999 1, n=6) within the mass concentration range of 20~100 μg/mL. Precision, repeatability, and stability tests of these compounds showed that the relative standard deviation (RSD) of the peak area was less than 2.00%, and the spiked recovery rate ranged from 98.72% to 102.03%, demonstrating the accuracy and reliability of the established method. Additionally, the extraction method for 10 flavonoids in germinated black wheat was optimized, with the best efficiency achieved using a hydrochloric acid-ethanol solution with a 70% ethanol volume fraction(a solid-to-liquid ratio of 1 g∶25 mL), and refluxing at 85 ℃ for 90 min. The mass fractions of 10 flavonoid were also determined in black wheat germinated for 1, 3, 5, and 7 d. This research lays a foundation for studying functional bioactive ingredients in germinated black wheat and offers a reference for the quality control and evaluation of its products.
