Neuroprotective Effects of Geopung-Chunghyuldan Based on Its Salvianolic Acid B Content Using an In Vivo Stroke Model

Abstract

Background: Geopung-Chunghyuldan (GCD) has neuroprotective properties. Salviae miltiorrhizae Radix plays an essential role in GCD's effect. The Salviae miltiorrhizae Radix marker compound is salvianolic acid B; however, its content is not uniform among samples. This study aimed to evaluate the neuroprotective effects of GCD based on salvianolic acid B content.

Methods: The neuroprotective effects of GCD based on the salvianolic acid B content were evaluated by measuring infarct volume 24 h after permanent middle cerebral artery occlusion in an in vivo stroke model. For the experimental group, each GCD was administered immediately before surgery. The control groups were administered distilled water and aspirin (30 mg/kg) in the same way. The salvianolic acid B content in five types of Salviae Miltiorrhizae Radix (two Chinese and three Korean regions) based on different cultivation regions was analyzed by high-performance liquid chromatography.

Results: Three samples met the Korean and Chinese Pharmacopeia standards for salvianolic acid B. However, two samples did not. GCDs with high salvianolic acid B showed marked neuroprotective effects compared to the control groups, whereas GCDs with low salvianolic acid B did not.

Conclusions: The salvianolic acid B content of Salviae miltiorrhizae Radix affects the neuroprotection effect of GCD. Stable, raw Salviae miltiorrhizae Radix is essential for GCD homogenization.

Keywords: Geopung-Chunghyuldan; Salviae miltiorrhizae Radix; neuroprotection; permanent middle cerebral artery occlusion; salvianolic acid B.

Figure 1

Results of high-performance liquid chromatography chromatogram for the salvianolic acid B in five types of Salviae mitiorrhizae Radix. (A) Standardized salvianolic acid B. (B) Salvianolic acid B in C1. (C) Salvianolic acid B in C2. (D) Salvianolic acid B in K1. (E) Salvianolic acid B in K2. (F) Salvianolic acid B in K3. K1, K2, and K3 showed high peak values similar to that of the standardized salvianolic acid B. C1 and C2 showed low peak values. C, Chinese Salviae miltiorrhizae Radix; K, Korean Salviae miltiorrhizae Radix.

Figure 2

Effects of experimental drug A60+B(C1) 30 mg/kg on ischemic brain injury after permanent middle cerebral artery occlusion (pMCAO). Infarct volume was measured 24 h after pMCAO. DW and all drugs were administered just before pMCAO. (A) Individual infarct volume values for each group (n = 13–17 mice in each group). (B Upper) Representative photographs of infarcted brain slices from DW- and drug-treated mice. (B Lower) There was no significant difference in infarct volume among groups. DW, distilled water; ASA, Aspirin^®; A, Ethanol extract of Coptidis Rhizoma, Phellodendri Cortex, Scutellariae Radix, Gardeniae Fructus, and Rhei Rhizoma; B(C1), Ethanol extract of 2018 Chinese Salviae miltiorrhizae Radix and Notoginseng Radix. Data represent means ± standard deviation.

Figure 3

Effects of experimental drug A60+B(C2) 30 mg/kg on ischemic brain injury after permanent middle cerebral artery occlusion (pMCAO). Infarct volume was measured 24 h after pMCAO. DW and all drugs were administered just before pMCAO. (A) Individual infarct volume values for each group (n = 9–17 mice in each group). (B Upper) Representative photographs of infarcted brain slices from DW- and drug-treated mice. (B Lower) No significant difference in infarct volume was observed among groups. DW, distilled water; ASA, Aspirin^®; A, Ethanol extract of Coptidis Rhizoma, Phellodendri Cortex, Scutellariae Radix, Gardeniae Fructus, and Rhei Rhizoma; B(C2), Ethanol extract of 2019 Chinese Salviae miltiorrhizae Radix and Notoginseng Radix. Data represent means ± standard deviation.

Figure 4

Effects of experimental drug A60+B(K1) 30 mg/kg on ischemic brain injury after permanent middle cerebral artery occlusion (pMCAO). Infarct volume was measured 24 h after pMCAO. DW and all drugs were administered just before pMCAO. (A) Individual infarct volume values for each group (n = 15–19 mice for each group). (B Upper) Representative photographs of infarcted brain slices from DW- and drug-treated mice. (B Lower) The group of experimental drug A+BK1 exhibited a significant decrease in infarct volume after pMCAO compared to groups DW and A60. DW, distilled water; ASA, Aspirin^®; A, Ethanol extract of Coptidis Rhizoma, Phellodendri Cortex, Scutellariae Radix, Gardeniae Fructus, and Rhei Rhizoma; B(K1), Ethanol extract of 2019 Yeongyang Korean Salviae miltiorrhizae Radix and Notoginseng Radix. Data represent means ± standard deviation. * p < 0.05 vs. DW; ** p < 0.01 vs. A60 by analysis of variance followed by Tukey’s test.

Figure 5

Effects of experimental drug A60+B(K2) 30 mg/kg on ischemic brain injury after permanent middle cerebral artery occlusion (pMCAO). Infarct volume was measured 24 h after pMCAO. DW and all drugs were administered just before pMCAO. (A) Individual infarct volume values for each group (n = 15–19 mice for each group). (B Upper) Representative photographs of infarcted brain slices from DW- and drug-treated mice. (B Lower) The group of experimental drug A+BK2 exhibited a significant decrease in infarct volume after pMCAO compared to groups DW and A60. DW, distilled water; ASA, Aspirin^®; A, Ethanol extract of Coptidis Rhizoma, Phellodendri Cortex, Scutellariae Radix, Gardeniae Fructus, and Rhei Rhizoma; B(K2), Ethanol extract of 2019 Jangheung Korean Salviae miltiorrhizae Radix and Notoginseng Radix. Data represent means ± standard deviation. * p < 0.05 vs. DW; ** p < 0.01 vs. A60 by analysis of variance followed by Tukey’s test.

Figure 6

Effects of experimental drug A60+B(K3) 30 mg/kg on ischemic brain injury after permanent middle cerebral artery occlusion (pMCAO). Infarct volume was measured 24 h after pMCAO. DW and all drugs were administered just before pMCAO. (A) Individual infarct volume values for each group (n = 15–19 mice for each group). (B Upper) Representative photographs of infarcted brain slices from DW- and drug-treated mice. (B Lower) The group administered the experimental drug A+BK3 exhibited a significant decrease in infarct volume after pMCAO compared to the A60 group. DW, distilled water; ASA, Aspirin^®; A, Ethanol extract of Coptidis Rhizoma, Phellodendri Cortex, Scutellariae Radix, Gardeniae Fructus, and Rhei Rhizoma; B(K3), Ethanol extract of 2020 Korean Salviae miltiorrhizae Radix and Notoginseng Radix. Data represent means ± standard deviation. * p < 0.05 vs. A60 by analysis of variance followed by Tukey’s test.