Objective

2

To establish an ultra-high performance liquid chromatography-tandem triple quadrupole mass spectrometry（UHPLC-QqQ-MS） for determination of the active ingredients in Erdongtang， and to predict the targets and pathways of anti-insulin resistance action of this formula.

Method

2

The analysis was performed on an ACQUITY UPLC BEH C

18

column（2.1 mm×100 mm， 1.7 μm） with the mobile phase of 0.1% formic acid aqueous solution（A）-acetonitrile（B） for gradient elution（0-3 min， 90%-87%A； 3-6 min， 87%-86%A； 6-9 min， 86%-83%A； 9-11 min， 83%-75%A； 11-18 min， 75%-70%A； 18-19 min， 70%-52%A； 19-22 min， 52%A； 22-25 min， 52%-5%A； 25-27 min， 5%-90%A； 27-30 min， 90%A）. The contents of active ingredients in Erdongtang was detected by electrospray ionization（ESI） and multiple reaction monitoring（MRM） mode under positive and negative ion modes. On this basis， network pharmacology was applied to predict the targets and pathways of Erdongtang exerting anti-insulin resistance effect.

Result

2

The 20 active ingredients in Erdongtang showed good linear relationships within a certain mass concentration range， and the precision， stability， repeatability and recovery rate were good. The results of determination showed that the ingredients with high content in 15 batches of samples were baicalein（1 259.39-1 635.78 mg·L

-1

）， baicalin（1 078.37-1 411.52 mg·L

-1

）， the ingredients with medium content were mangiferin（148.59-217.04 mg·L

-1

）， timosaponin BⅡ（245.10-604.89 mg·L

-1

）， quercetin-3-

O

-glucuronide（89.30-423.26 mg·L

-1

）， rutin（46.91-1 553.61 mg·L

-1

）， glycyrrhizic acid（55.97-391.47 mg·L

-1

）， neomangiferin（37.45-127.03 mg·L

-1

）， nuciferine（0.89-63.48 mg·L

-1

）， hyperoside（6.96-136.78 mg·L

-1

）， liquiritin（30.89-122.78 mg·L

-1

）， liquiritigenin（26.64-110.67 mg·L

-1

）， protodioscin（58.57-284.26 mg·L

-1

）， the ingredients with low content were wogonin（7.16-20.74 mg·L

-1

）， pseudoprotodioscin（5.49-22.96 mg·L

-1

）， ginsenoside Rb

1

（7.31-23.87 mg·L

-1

）， ginsenoside Rg

1

（10.78-28.33 mg·L

-1

）， ginsenoside Re（7.78-24.76 mg·L

-1

）， ophiopogonin D（2.08-4.29 mg·L

-1

）， methylophiopogonanone A（0.74-1.67 mg·L

-1

）. The results of network pharmacology indicated that the mechanism of anti-insulin resistance exerted by Erdongtang might be related to the phosphatidylinositol 3-kinase/protein kinase B（PI3K/Akt） signaling pathway.

Conclusion

2

The established UHPLC-QqQ-MS has the advantages of simple sample processing， strong exclusivity and high sensitivity， and can simultaneously determine the contents of the main ingredients from seven herbs in Erdongtang， which can lay the foundation for the development of Erdongtang compound preparations. The results of the network pharmacology can provide a reference for the mechanism study of Erdongtang in the treatment of type 2 diabetes mellitus.