To simultaneously determine the contents of p-coumaric acid, chlorogenic acid, 5-caffeoylquinic acid, 4-caffeoylquinic acid, caffeic acid and ferulic acid in Imperatae Rhizoma concentrated granules, an ultra-high performance liquid chromatography (UPLC) with two internal references method (TIRM) was established and validated. Chromatographic separation was achieved on a ZORBAX RRHD Eclipse Plus C18 column (2.1 mm×100 mm, 1.8 μm) using 1.7 mmol·L-1 oxalic acid in water and methanol as mobile phase. The flow rate was 0.4 mL·min-1 and the column temperature was set as 35℃. The relative correction factors (RCFs) of caffeic acid and ferulic acid using p-coumaric acid as internal reference were calculated and the RCFs of 4-caffeoylquinic acid and 5-caffeoylquinic acid were calculated using chlorogenic acid as the internal reference. The TIRM was fully validated for linearity, accuracy, repeatability, stability and recovery so that it could be compared with the external standard method (ESM). The RCFs of 5-caffeoylquinic acid, 4-caffeoylquinic acid, caffeic acid, and ferulic acid were 1.069, 1.022, 1.368, and 1.493, respectively. The TIRM and ESM were used to determine the contents of six ingredients in Imperatae Rhizoma concentrated granules from different manufacturers and the variation between results was within acceptable limits. In conclusion, the newly established TIRM allowed simultaneous determination of six ingredients (p-coumaric acid, chlorogenic acid, 5-caffeoylquinic acid, 4-caffeoylquinic acid, caffeic acid, ferulic acid) in Imperatae Rhizoma concentrated granules, providing support for the quality control of this traditional Chinese medicine.
| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |