Objective:

2

To develop an interference-free and rapid method to elucidate Guanxin Ⅱ (GXⅡ)'s representative vasodilator absorbed bioactive compounds (ABCs) among enormous phytochemicals.

Methods:

2

The contents of ferulic acid

tanshinol

and hydroxysafflor yellow A (FTA) in GXⅡ/rat serum after the oral administration of GXⅡ (30 g/kg) were detected using ultra-performance liquid chromatography-mass spectrometry. Totally 18 rats were randomly assigned to the control group (0.9% normal saline)

GXⅡ (30 g/kg) and FTA (5

28 and 77 mg/kg) by random number table method. Diastolic coronary flow velocity-time integral (VTI)

i.e.

coronary flow or coronary flow-mediated dilation (CFMD)

and endothelium-intact vascular tension of isolated aortic rings were measured. After 12 h of exposure to blank medium or 0.5 mmol/L H

2

O

2

endothelial cells (ECs) were treated with post-dose GXⅡ of supernatant from deproteinized serum (PGSDS

300 μL PGSDS per 1 mL of culture medium) or FTA (237

1539

and 1510 mg/mL) for 10 min as control

H

2

O

2

PGSDS and FTA groups. Nitric oxide (NO)

vascular endothelial growth factor (VEGF)

endothelin-1 (ET-1)

superoxide dismutase (SOD)

malondialdehyde (MDA) and phosphorylated phosphoinositide 3 kinase (p-PI3K)

phosphorylated protein kinase B (p-AKT)

phosphorylated endothelial nitric oxide synthase (p-eNOS) were analyzed. PGSDS was developed as a GXⅡ proxy of

ex vivo

herbal crude extracts.

Results:

2

PGSDS effectively eliminates false responses caused by crude GXⅡ preparations. When doses equaled the contents in GXⅡ/its post-dose serum

FTA accounted for 98.17% of GXⅡ-added CFMD and 92.99% of PGSDS-reduced vascular tension. In ECs

FTA/PGSDS was found to have significant antioxidant (lower MDA and higher SOD

P

<

0.01) and endothelial function-protective (lower VEGF

ET-1

P

<

0.01) effects. The increases in aortic relaxation

endothelial NO levels and phosphorylated PI3K/Akt/eNOS protein induced by FTA/PGSDS were markedly abolished by NG-nitro-L-arginine methyl ester (L-NA

eNOS inhibitor) and wortmannin (PI3K/AKT inhibitor)

respectively

indicating an endothelium-dependent vasodilation via the PI3K/AKT-eNOS pathway (

P

<

0.01).

Conclusion:

2

This study provides a strategy for rapidly and precisely elucidating GXⅡ's representative

in

/

ex vivo

cardioprotective absorbed bioactive compounds (ABCs)-FTA

suggesting its potential in advancing precision ethnomedicine.