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To establish methods of thin-layer chromatography(TLC) qualitative identification and quantitative analysis of multi-components by single-marker (QAMS) according to the characterization results of the main components of Iris tectorum Maxim..
Firstly，the main components of Iris tectorum were characterized by ultra performance liquid chromatography tandem time-of-flight mass spectrometry(UPLC-Q TOF MS/MS) technology，using an ACQUITY UPLC® BEH C18 column (100 mm×2.1 mm，1.7 μm) with gradient elution using 0.1% formic acid-water (A)-acetonitrile (B) as mobile phase，and the data were collected in positive ion mode. In addition，the qualitative identification method was established by the TLC. Finally，quantitative analysis was performed on an Agilent 5 TC-C18 column (250 mm×4.6 mm，5 μm). The mobile phase was 0.05% phosphoric acid water(A)-acetonitrile(B) with gradient elution. The QAMS method was established by using tectorigenin as an internal standard to establish its relative correction factor with tectoridin and irigenin，and the contents of three isoflavones were calculated by the correction factor. At the same time，the accuracy and feasibility of the QAMS method was verified by the external standard method(ESM).
Five isoflavones were characterized (tectoridin，iristectorin B，tectorigenin，irigenin and iristectorigenin A) by UPLC-Q TOF MS/MS. A TLC identification method for four components，tectoridin，iristectorin B，tectorigenin and irigenin was established. In this study，the linearity of the three components of tectoridin，tectorigenin and irigenin were good in a certain concentration range (r>0.999)，with the average recoveries of 97.7%-100.8% and the RSDs of 1.1%-2.9%，and the relative correction factors of tectoridin and irigenin were 1.114 5 and 0.827 0，respectively. And the repeatability of the correction factors was good under different experimental conditions (RSD<5%). There was no significant difference between the contents of the three analytes in ten different batches of Iris tectorum determined by QAMS and ESM. The contents of tectorigenin，tectoridin and irigenin in 10 batches of Iris tectorum from different habitats determined by QAMS were not significantly different from those determined by ESM.
Based on the characterized components of Iris tectorum medicinal materials，the TLC and QAMS methods with high-content and stable ingredients are established. The methods are simple and stable，which can be used for qualitative and quantitative analysis and quality evaluation of Iris tectorum.
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| 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 |
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