Screening of 56 Herbal formulas covered by the National Health Insurance Service on Dementia-related Factors

Article information

J Korean Med. 2018;39(3):1-16
Publication date (electronic) : 2018 September 30
doi : https://doi.org/10.13048/jkm.18020
1Clinical Medicine Division, Korea Institute of Oriental Medicine, Daejeon 34054, Republic of Korea
2College of Pharmacy, Chungnam National University
Correspondence to:정수진(Soo-Jin Jeong) 대전 유성구 유성대로 1672 한국한의학연구원 임상의학부 Tel: +82-42-868-9651, Fax: +82-41-521-7007, E-mail: sjijeong@kiom.re.kr
Received 2018 July 12; Revised 2018 September 7; Accepted 2018 September 7.

Abstract

Objectives

The purpose of this study was to investigate the effects of 56 herbal formulae covered by the National Health Insurance Corporation (NHIC) on dementia-related biomarkers and neuronal cell changes.

Methods

The 56 herbal formulae were extracted with 70% ethanol at 100°C for 2 h. The antioxidant properties was measured by radical scavenging assay using ABTS+ radical. The acetylcholinesterase (AChE) activity was tested by Ellman’s assay and amyloid-β (Aβ) aggregation was determined using fluorescence method. To estimate the inhibitory effects of herbal formulae on neuronal cell death and inflammation using HT22 hippocampal cells and BV-2 microglia, respectively.

Results

Among the 56 herbal formulae, Dangguiyukhwangtang, Banhasasimtang, Samhwangsasimtang, Cheongwiesan, Hwangryunhaedoktang, Banhabaekchulchunmatang, Jaeumganghwatang, Cheongseoikgitang, and Hoechunyanggyuksan has a significant inhibitory effects on acetylcholinesterase (AChE) activity. Doinseunggitang and Samhwangsasimtang exerted the effect on the inhibition of amyloid-β (Aβ) aggregation. Additionally, 10 herbal formulae affected AChE and Aβ aggregation revealed antioxidant activity as well as neuroprotective and anti-neuroinflammation effects in neuronal cell lines.

Conclusions

10 herbal formulae that have been shown to be effective against the major dementia markers have been shown to have antioxidant activity, neuronal cell protection and inhibition of brain inflammation. Further investigation of these herbal formulae will need to be validated in dementia animal models.

Inhibition (%)=[1-{(S-S)/(C-C)}]×100
aggregation inhibition (%)=[1-{(S-S)/(C-C)}]×100
Fig. 1

Inhibitory effects of ethanol extract of herbal formulas on acetylcholinesterase (AChE) activity. (A) Dangguiyukhwangtang (DYT), (B) Banhasasimtang (BST), (C) Samhwangsasimtang (SST), (D) Cheongwiesan (CS), (E) Hwangryunhaedoktang (HHT), (F) Banhabaekchulchunmatang (BBCT), (G) Jaeumganghwatang (JGT), (H) Cheongseoikgitang (CIT), and (I) Hoechunyanggyuksan (HYS).

Fig. 2

Inhibitory effects of ethanol extracts of herbal formulas on amyloid β (Aβ) aggregation. (A) Doinseunggitang (DST) and (B) Samhwangsasimtang (SST).

Fig. 3

ABTS radical scavenging activity of ethanol extracts of dementia effective herbal formulas. (A) Dangguiy-ukhwangtang (DYT), (B) Doinseunggitang (DST), (C) Banhasasimtang (BST), (D) Samhwangsasimtang (SST), (E) Cheongwiesan (CS), (F) Hwangryunhaedoktang (HHT), (G) Banhabaekchulchunmatang (BBCT), (H) Jaeumganghwatang (JGT), (I) Cheongseoikgitang (CIT), and (J) Hoechunyanggyuksan (HYS).

Fig. 4

Neuroprotective effects of ethanol extracts of dementia effective herbal formulas. HT22 cells were exposed to H2O2 in the absence or presence of herbal formula extracts for 6 h. Cell viability was determined using CCK assay kit. The results are expressed as mean ± SEM of three independent experiments. ###P< 0.01 vs vehicle control cells; *, P<0.05 **, P<0.01 ***, P<0.001 vs H2O2-treated cells. (A) Dangguiyukhwangtang (DYT), (B) Doinseunggitang (DST), (C) Banhasasimtang (BST), (D) Samhwangsasimtang (SST), (E) Cheongwiesan (CS), (F) Hwangryunhaedoktang (HHT), (G) Banhabaekchulchunmatang (BBCT), (H) Jaeumganghwatang (JGT), (I) Cheongseoikgitang (CIT), and (J) Hoechunyanggyuksan (HYS).

Fig. 5

Anti-neuroinflammatory effects of ethanol extracts of dementia effective herbal formulas. BV-2 cells were pretreated with herbal formula extracts for 2 h and stimulated with LPS for additional 22 h. Culture supernatants were collected and subjected to NO assay using Greiss reagents. Data represent mean ± SEM from three independent experiments. ###, P<0.001 vs untreated control; *, P<0.05 **, P<0.01 ***, P<0.001 vs LPS-treated group. (A) Dangguiyukhwangtang (DYT), (B) Doinseunggitang (DST), (C) Banhasasimtang (BST), (D) Samhwangsasimtang (SST), (E) Cheongwiesan (CS), (F) Hwangryunhaedoktang (HHT), (G) Banhabaekchulchunmatang (BBCT), (H) Jaeumganghwatang (JGT), (I) Cheongseoikgitang (CIT), and (J) Hoechunyanggyuksan (HYS).

Inhibitory Activity of the 56 Herbal Formulas in the National Health Insurance Coverage on AChE Activity and Amyloid-β Aggregation in vitro (at 100 μg/mL)

Cytotoxicity of Dementia Effective Herbal Formulas Against HT22 cells.

Cytotoxicity of Dementia Effective Herbal Formulas Against BV-2 cells.

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Article information Continued

Fig. 1

Inhibitory effects of ethanol extract of herbal formulas on acetylcholinesterase (AChE) activity. (A) Dangguiyukhwangtang (DYT), (B) Banhasasimtang (BST), (C) Samhwangsasimtang (SST), (D) Cheongwiesan (CS), (E) Hwangryunhaedoktang (HHT), (F) Banhabaekchulchunmatang (BBCT), (G) Jaeumganghwatang (JGT), (H) Cheongseoikgitang (CIT), and (I) Hoechunyanggyuksan (HYS).

Fig. 2

Inhibitory effects of ethanol extracts of herbal formulas on amyloid β (Aβ) aggregation. (A) Doinseunggitang (DST) and (B) Samhwangsasimtang (SST).

Fig. 3

ABTS radical scavenging activity of ethanol extracts of dementia effective herbal formulas. (A) Dangguiy-ukhwangtang (DYT), (B) Doinseunggitang (DST), (C) Banhasasimtang (BST), (D) Samhwangsasimtang (SST), (E) Cheongwiesan (CS), (F) Hwangryunhaedoktang (HHT), (G) Banhabaekchulchunmatang (BBCT), (H) Jaeumganghwatang (JGT), (I) Cheongseoikgitang (CIT), and (J) Hoechunyanggyuksan (HYS).

Fig. 4

Neuroprotective effects of ethanol extracts of dementia effective herbal formulas. HT22 cells were exposed to H2O2 in the absence or presence of herbal formula extracts for 6 h. Cell viability was determined using CCK assay kit. The results are expressed as mean ± SEM of three independent experiments. ###P< 0.01 vs vehicle control cells; *, P<0.05 **, P<0.01 ***, P<0.001 vs H2O2-treated cells. (A) Dangguiyukhwangtang (DYT), (B) Doinseunggitang (DST), (C) Banhasasimtang (BST), (D) Samhwangsasimtang (SST), (E) Cheongwiesan (CS), (F) Hwangryunhaedoktang (HHT), (G) Banhabaekchulchunmatang (BBCT), (H) Jaeumganghwatang (JGT), (I) Cheongseoikgitang (CIT), and (J) Hoechunyanggyuksan (HYS).

Fig. 5

Anti-neuroinflammatory effects of ethanol extracts of dementia effective herbal formulas. BV-2 cells were pretreated with herbal formula extracts for 2 h and stimulated with LPS for additional 22 h. Culture supernatants were collected and subjected to NO assay using Greiss reagents. Data represent mean ± SEM from three independent experiments. ###, P<0.001 vs untreated control; *, P<0.05 **, P<0.01 ***, P<0.001 vs LPS-treated group. (A) Dangguiyukhwangtang (DYT), (B) Doinseunggitang (DST), (C) Banhasasimtang (BST), (D) Samhwangsasimtang (SST), (E) Cheongwiesan (CS), (F) Hwangryunhaedoktang (HHT), (G) Banhabaekchulchunmatang (BBCT), (H) Jaeumganghwatang (JGT), (I) Cheongseoikgitang (CIT), and (J) Hoechunyanggyuksan (HYS).

Table 1

Inhibitory Activity of the 56 Herbal Formulas in the National Health Insurance Coverage on AChE Activity and Amyloid-β Aggregation in vitro (at 100 μg/mL)

Herbal formulas Original AChE activity inhibition (%) Amyloid-β aggregation inhibition (%)
1 가미소요산(加味逍遙散) 동의신경정신과학 2.47±0.82 −17.88±6.31
2 당귀육황탕(當歸六黃湯) 한방산부인과학 92.32±0.39 15.38±2.37
3 대시호탕(大柴胡湯) 상한론 3.11±2.26 26.76±17.64
4 대황목단피탕(大黃牧丹皮湯) 동의보감 22.54±2.30 47.47±5.79
5 도인승기탕(桃仁承氣湯) 방약합편 26.42±1.61 74.06±3.44
6 반하사심탕(半夏瀉心湯) 동의보감 83.47±1.00 51.63±1.92
7 백출탕(白朮湯) 동의보감 −18.49±0.78 −17.74±17.80
8 보중익기탕(補中益氣湯) 동원십서 −16.20±1.76 −1.32±1.92
9 보허탕(補虛湯) 의학입문 −16.85±1.65 −5.85±1.10
10 삼황사심탕(三黃瀉心湯) 동의보감 93.30±0.23 76.00±5.57
11 소시호탕(小柴胡湯) 상한론 −8.03±5.33 46.15±2.61
12 시호계지탕(柴胡桂枝湯) 동의보감 −12.85±2.28 36.46±5.62
13 오림산(五淋散) 방약합편 −0.45±1.87 24.56±4.39
14 이중탕(理中湯) 방약합편 −12.23±1.04 23.29±1.60
15 이진탕(二陳湯) 방약합편 −9.22±1.19 −26.37±35.65
16 익위승양탕(益胃升陽湯) 방약합편 −9.85±1.33 −6.76±10.14
17 인진호탕(茵蔯蒿湯) 득효력 30.30±0.32 36.98±2.58
18 조위승기탕(調胃承氣湯) 동의보감 34.33±3.15 64.80±10.10
19 청위산(淸胃散) 방약합편 89.12±0.10 −0.29±16.49
20 팔물탕(八物湯) 방약합편 −2.23±3.80 7.83±15.77
21 황금작약탕(黃金芍藥湯) 방약합편 9.07±1.42 41.89±12.74
22 황련해독탕(黃連解毒湯) 방약합편 96.71±0.31 38.57±5.20
23 갈근탕(葛根湯) 상한론 −4.48±3.32 4.64±0.33
24 갈근해기탕(葛根解肌湯) 방약합편 −13.33±9.08 17.90±0.95
25 구미강활탕(九味羌活湯) 방약합편 −25.32±2.82 16.38±2.43
26 궁소산(芎蘇散) 방약합편 −16.28±2.15 29.62±0.24
27 궁하탕(芎夏湯) 방약합편 −6.82±6.34 21.54±0.54
28 내소산(內消散) 방약합편 −19.34±4.48 4.31±0.06
29 당귀연교음(當歸連翹飮) 동의보감 −18.15±3.04 5.78±4.29
30 대청룡탕(大靑龍湯) 상한론 −24.89±3.82 −3.15±3.40
31 대화중음(大和中飮) 방약합편 −5.67±1.53 7.94±3.70
32 반하백출천마탕(半夏白朮天麻湯) 방약합편 67.71±0.11 −3.94±1.56
33 반하후박탕(半夏厚朴湯) 동의보감 −2.30±1.29 42.42±2.31
34 복령보심탕(茯苓補心湯) 방약합편 −7.89±3.29 21.25±6.75
35 불환금정기산(不換金正氣散) 동의보감 −11.45±2.33 29.31±6.20
36 삼소음(蔘蘇飮) 동의보감 −16.97±2.26 11.76±0.83
37 삼출건비탕(蔘朮健脾湯) 동의보감 −15.97±2.12 0.90±1.80
38 삼호작약탕(蔘胡芍藥湯) 방약합편 −19.57±1.09 25.62±0.48
39 생맥산(生脈散) 천금익방 −27.77±1.66 2.99±3.06
40 소청룡탕(小靑龍湯) 상한론 −13.75±0.41 23.94±2.20
41 승양보위탕(升陽補胃湯) 동의보감 2.22±2.23 22.45±5.68
42 시경반하탕(柴梗半夏湯) 방약합편 −5.00±1.82 47.56±1.95
43 시호소간탕(柴胡疏肝湯) 동양의학대사전 0.23±2.56 28.36±2.78
44 시호청간탕(柴胡淸肝湯) 의종금감 3.07±0.39 44.85±30.86
45 안태음(安胎飮) 동의보감 1.25±1.12 33.49±4.08
46 연교패독산(連翹敗毒散) 동의보감 −7.39±0.81 24.20±6.41
47 오적산(五積散) 방약합편 −1.93±0.26 27.59±4.80
48 인삼패독산(人蔘敗毒散) 동의보감 −3.07±0.43 34.16±1.94
49 자음강화탕(滋陰降火湯) 만병회춘 75.72±0.55 37.48±2.64
50 청상견통탕(淸上蠲痛湯) 제중신편 −2.39±0.71 51.58±3.35
51 청서익기탕(淸署益氣湯) 방약합편 82.60±0.30 25.64±6.89
52 평위산(平胃散) 동의보감 −6.42±0.61 31.08±5.91
53 행소탕(산)(杏蘇湯(散)) 동의보감 −7.73±0.81 25.66±18.14
54 향사평위산(香砂平胃散) 방약합편 1.19±0.65 33.58±7.45
55 형개연교탕(荊芥連翹湯) 방약합편 0.51±0.43 38.42±2.03
56 회춘양격산(回春涼隔散) 방약합편 91.87±0.20 49.94±1.20

Berberine 60.63±0.6

Morin 68.3±6.2

Table 2

Cytotoxicity of Dementia Effective Herbal Formulas Against HT22 cells.

μg/mL 0 12.5 25 50 100

Herbal formula
당귀육황탕 100.00±2.47 100.08±7.36 100.25±11.7 100.40±7.12 100.83±5.36
도인승기탕 100.00±8.63 100.27±2.30 100.68±9.44 100.44±8.58 84.89±6.54
반하사심탕 100.00±3.23 101.82±4.37 100.29±8.48 80.75±8.39 67.01±0.46
삼황사심탕 100.00±7.55 100.92±1.81 100.58±9.79 100.83±9.50 84.83±2.96
청위산 100.00±8.52 102.53±5.83 100.07±3.31 100.40±3.01 100.95±3.43
황련해독탕 100.00±8.89 100.89±3.55 100.48±7.84 106.01±5.02 71.12±0.13
반하백출천마탕 100.00±7.80 100.98±3.18 100.48±7.84 106.01±5.02 71.12±0.13
자음강화탕 100.00±6.31 109.26±7.06 102.77±5.95 94.10±4.37 81.54±1.69
청서익기탕 100.00±2.14 100.74±1.47 100.69±1.25 100.30±4.14 86.70±1.59
회춘양격탕 100.00±3.67 100.51±4.82 100.86±2.18 109.49±4.92 95.66±4.72

Table 3

Cytotoxicity of Dementia Effective Herbal Formulas Against BV-2 cells.

μg/mL 0 12.5 25 50 100

Herbal formula
당귀육황탕 100.00±5.27 102.59±4.92 93.39±4.07 90.08±2.43 83.00±2.50
도인승기탕 100.00±1.38 102.56±2.58 102.78±3.27 95.89±4.55 84.96±5.36
반하사심탕 100.00±3.50 95.99±1.11 98.88±1.79 96.17±1.39 85.98±1.30
삼황사심탕 100.00±8.54 80.96±2.24 73.97±2.81 70.32±2.37 66.39±1.03
청위산 100.00±1.53 97.00±2.48 92.33±2.54 90.15±1.98 83.14±1.33
황련해독탕 100.00±2.95 113.31±5.74 99.46±4.13 91.64±4.15 77.36±2.19
반하백출천마탕 100.00±1.92 99.83±1.66 96.63±1.79 91.97±3.05 91.00±0.90
자음강화탕 100.00±1.19 95.42±1.89 91.31±2.68 90.57±0.88 90.44±1.00
청서익기탕 100.00±1.15 98.55±2.81 94.08±2.15 92.57±1.53 91.27±1.50
회춘양격탕 100.00±2.65 98.31±2.08 95.85±1.40 93.65±0.19 93.44±1.84