# Analysis of Yield of Eleutherosides B and E inAcanthopanax divaricatus and A. koreanum Grown with Varying Cultivation Methods

Authors: Jeong Min Lee; Myoung-Hee Lee; Suk-Bok Pae; Ki-Won Oh; Chan-Sik Jung; In-Youl Baek; Sanghyun Lee
Journal: The Scientific World Journal
 (2014)

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## Abstract

An analysis of the yield of eleutherosides B and E inAcanthopanax divaricatus and A. koreanum was performed using high performance liquid chromatography to evaluate production by different cultivation methods. In A. divaricatus and A. koreanum, the total content of eleutherosides B and E was 2.466–7.360 mg/g varying by plant section, 3.886–11.506 mg/g by pinching site, 3.655–10.083 mg/g by planting time, and 3.652–10.108 mg/g by fertilizer ratio. Thus the total content of eleutherosides B and E in A. divaricatus and A. koreanum differed depending on cultivation methods. These results present useful information for high eleutheroside content applications in A. divaricatus and A. koreanum. This information can affect selection of plant section and cultivation methods for nutraceutical, pharmaceutical, and cosmeceutical material development.


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## Body

## 1. Introduction

Acanthopanax species are commonly known as Siberian ginseng, touch-me-not, devil’s shrub, prickly, and wild pepper [1]. A diverse group of chemical compounds isolated fromAcanthopanax species was named “eleutherosides.” The group of eleutherosides consists of eleutherosides A (daucosterol), B (syringin), B1 (isofraxidin-7-O-glucoside), B2, B3, B4 (sesamin), C (methyl-α-D-galactoside), D, E (syringaresinol-di-O-β-D-glucoside), E1 (syringaresinol-O-β-D-glucoside), E2 (episyringaresinol 4′′-O-β-D-glucoside), F, G, I, K, L, and M (hederasaponin B) [2–5]. Among the eleutherosides, eleutheroside E fromA. senticosus has the most noticeable stimulant and antistress effects [6].There have been many studies on the activities of eleutherosides B and E. Eleutherosides B and E have been reported to have protective effects against amyloidβ(25–35)-induced neuritic atrophy in cultured rat cortical neurons [7], neuroprotective effects against transient global cerebral ischemia in rats [8], protective effects in dopaminergic neurons in Parkinson’s disease mice [9], and antioxidant properties [10]. There have been many studies determining the effects of eleutherosides B and E. The quantitative analysis of eleutherosides B and E fromAcanthopanax species has been reported from various parts of members ofAcanthopanax species [11, 12], includingA. sessiliflorus fruits and fermented wine made from them [13] and in the roots ofA. senticosus [14].Acanthopanax species are cultivated and grow wild in various areas in Korea, and the variety of their pharmacological effects has attracted consumers’ interests. However, there are many difficulties in producing high qualityAcanthopanax species, depending upon the location and method of culture, which affect different pharmacological ingredients [15].This study, therefore, analyzed eleutherosides B and E inA. divaricatus andA. koreanum, depending on plant section and cultivation method, using high performance liquid chromatography (HPLC) with the goal of optimizing the content of eleutherosides B and E by cultivation method, to suggest efficient cultivation methods forA. divaricatus andA. koreanum.


## 2. Materials and Methods

### 2.1. Plant Materials

A. divaricatus (Voucher number LEE11-1) andA. koreanum (Voucher number LEE11-2) were collected at Yeongcheon Agricultural Technology & Extension Center, Yeongcheon, Korea. Plant section (Ps) was designated as Ps-1, -2, -3, -4, or -5 (Figure 1). Different cultivation method criteria consisted of pinching site (30 and 60 cm), planting time (March 30, April 15, and April 30), and fertilizer ratio (N-P-K, 10.5-8.5-8.5: 50 kg/10a; 2N-P-K, 21-8.5-8.5: 50 kg/10a; N-2P-K, 10.5-17-8.5: 50 kg/10a; N-P-2K, 10.5-8.5-17: 50 kg/10a; 2N-2P-2K, 21-17-17: 50 kg/10a).Sample collection fractions by plant section (a) and by upper and lower parts of plants (b).
(a)
(b)


### 2.2. Cultivation Methods

Tilling was carried out on March 20, during which fully fermented compost (1,500 kg/10a) was added to the soil, andA. divaricatus andA. koreanum seedlings were planted on March 30. The compound fertilizer (2N-2P-2K, 21-17-17) fertilized the soil once in the June 10. The harvest time of all samples was February 13 of the following year. The cultivation conditions (2007) were 13.1°C average temperature and 1,142 mm average precipitation. The soil conditions were mature soil, pH 7.0, 3.4% soil organic content, 435 ppm available phosphate, and 0.36, 3.0, and 1.2 cmol+/kg of K, Ca, and Mg, respectively [16]. All samples were cultivated under the same above conditions.


### 2.3. Apparatus and Chemicals

Evaporation was conducted using an Eyela rotary evaporator system (Tokyo, Japan) under refluxin vacuo. HPLC chromatograms were recorded with an Agilent 1100 series HPLC (Waldbronn, Germany) equipped with Agilent 1100 series G1311A Quat pump and Agilent 1100 series G1315B detector. A Discovery C18 (4.6 × 250 mm, 5 μm) column was purchased from Sigma-Aldrich Co. (PA, USA). Water and acetonitrile used in this research were of HPLC grade, and all other reagents were of analytical grade.


### 2.4. Preparation of Eleutherosides B (1) and E (2)

The air-dried powdered stems ofA. senticosus were extracted with H2O. The extract was suspended in H2O and then partitioned successively with equal volumes of chloroform, ethyl acetate, andn-butanol. For qualitative and quantitative analysis, a combined eleutherosides B (CHCl3 : MeOH = 95 : 5) and E (CHCl3 : MeOH = 90 : 10) isolate was obtained by repeated column chromatography from the n-butanol fraction of stem ofA. senticosus [10, 12].Eleutheroside B (1): FAB-MS:m/z 373 [M+H]+; 1H-NMR (500 MHz, DMSO-d6): δ 6.73 (2H, s, H-2,6), 6.46 (1H, d,J= 15.9 Hz, H-7), 6.33 (1H, dt,J = 15.9, 5.1 Hz, H-8), 4.84 (1H, d,J = 7.5 Hz, glucosyl H-1), 4.11 (1H, dd,J = 5.1, 1.4 Hz, H-9a), 4.09 (1H, dd,J = 5.1, 1.4 Hz, H-9b), 3.77 (6H, s, 2 × OMe); 13C-NMR (125 MHz, DMSO-d6): δ 152.7 (C-3,5), 133.0 (C-4), 131.0 (C-7), 129.0 (C-8), 128.1 (C-1), 104.5 (C-2,6), 103.1 (Glc C-1), 77.4 (Glc C-5), 76.5 (Glc C-3), 74.9 (Glc C-2), 71.0 (Glc C-4), 62.0 (C-9), 60.5 (Glc C-6), 56.3 (OMe).Eleutheroside E (2): FAB-MS:m/z 743 [M+H]+; 1H-NMR (500 MHz, DMSO-d6): δ 6.67 (4H, s, H-2′,6′), 4.88 (2H, d,J = 7.3 Hz, glucosyl H-1), 4.67 (2H, d,J = 3.6 Hz, H-2), 4.28 (2H, dd,J = 8.5, 6.6 Hz, H-4eq), 4.20 (2H, dd,J = 8.5, 3.0 Hz, H-4ax), 3.76 (12H, s, 4 × OMe), 3.19 (2H, m, H-1); 13C-NMR (125 MHz, DMSO-d6): δ 153.2 (C-3′,5′), 138.1 (C-4′), 134.1 (C-1′), 104.6 (C-2′,6′), 103.3 (Glc C-1), 85.7 (C-2), 77.5 (Glc C-5), 76.7 (Glc C-3), 74.5 (Glc C-2), 72.1 (C-4), 70.2 (Glc C-4), 61.2 (Glc C-6), 57.0 (OMe), 54.2 (C-1).


### 2.5. Sample Preparation

To analyze the eleutherosides B (1) and E (2) content inA. divaricatus andA. koreanum, 5 g of each ofA. divaricatus andA. koreanum was extracted with 50% MeOH (3 × 100 mL) by reflux and evaporatedin vacuo. The residue was dissolved in 1 mL of MeOH and filtered with a 0.45 μm filter. The resulting solution was used for HPLC analysis.


### 2.6. HPLC Conditions

HPLC separation of eleutherosides B (1) and E (2) for qualitative and quantitative analysis was performed using a reverse phase system. A Discovery C18 (4.6 × 250 mm, 5 μm) column was used with a mobile phase that consisted of water and acetonitrile. A gradient solvent system of water and acetonitrile (90 : 10 to 70 : 30 for 20 min) was used for the elution program. UV detection was conducted at 350 nm. The injection volume was 10 μL and the flow rate was 1 mL/min. The temperature was maintained at 25°C. All injections were performed in triplicate.


### 2.7. Calibration Curve

A stock solution (1 mg/mL) of each eleutherosides B (1) and E (2) isolate was prepared in MeOH, and then the solution content was successively reduced to 50% to create different concentrations. The analyte contents were determined from the corresponding calibration curves. The calibration functions of the eleutherosides B (1) and E (2) isolate were calculated using the peak area (Y), concentration (X, μg/10 μL), and mean values (n=3) ± standard deviation (SD).


### 2.8. Statistical Analysis

Data for each sample was expressed as mean ± SD. ANOVA using the SAS Enterprise Guide software was calculated and the significance between the means of each group was determined using Duncan’s multiple test.


## 2.1. Plant Materials

A. divaricatus (Voucher number LEE11-1) andA. koreanum (Voucher number LEE11-2) were collected at Yeongcheon Agricultural Technology & Extension Center, Yeongcheon, Korea. Plant section (Ps) was designated as Ps-1, -2, -3, -4, or -5 (Figure 1). Different cultivation method criteria consisted of pinching site (30 and 60 cm), planting time (March 30, April 15, and April 30), and fertilizer ratio (N-P-K, 10.5-8.5-8.5: 50 kg/10a; 2N-P-K, 21-8.5-8.5: 50 kg/10a; N-2P-K, 10.5-17-8.5: 50 kg/10a; N-P-2K, 10.5-8.5-17: 50 kg/10a; 2N-2P-2K, 21-17-17: 50 kg/10a).Sample collection fractions by plant section (a) and by upper and lower parts of plants (b).
(a)
(b)


## 2.2. Cultivation Methods

Tilling was carried out on March 20, during which fully fermented compost (1,500 kg/10a) was added to the soil, andA. divaricatus andA. koreanum seedlings were planted on March 30. The compound fertilizer (2N-2P-2K, 21-17-17) fertilized the soil once in the June 10. The harvest time of all samples was February 13 of the following year. The cultivation conditions (2007) were 13.1°C average temperature and 1,142 mm average precipitation. The soil conditions were mature soil, pH 7.0, 3.4% soil organic content, 435 ppm available phosphate, and 0.36, 3.0, and 1.2 cmol+/kg of K, Ca, and Mg, respectively [16]. All samples were cultivated under the same above conditions.


## 2.3. Apparatus and Chemicals

Evaporation was conducted using an Eyela rotary evaporator system (Tokyo, Japan) under refluxin vacuo. HPLC chromatograms were recorded with an Agilent 1100 series HPLC (Waldbronn, Germany) equipped with Agilent 1100 series G1311A Quat pump and Agilent 1100 series G1315B detector. A Discovery C18 (4.6 × 250 mm, 5 μm) column was purchased from Sigma-Aldrich Co. (PA, USA). Water and acetonitrile used in this research were of HPLC grade, and all other reagents were of analytical grade.


## 2.4. Preparation of Eleutherosides B (1) and E (2)

The air-dried powdered stems ofA. senticosus were extracted with H2O. The extract was suspended in H2O and then partitioned successively with equal volumes of chloroform, ethyl acetate, andn-butanol. For qualitative and quantitative analysis, a combined eleutherosides B (CHCl3 : MeOH = 95 : 5) and E (CHCl3 : MeOH = 90 : 10) isolate was obtained by repeated column chromatography from the n-butanol fraction of stem ofA. senticosus [10, 12].Eleutheroside B (1): FAB-MS:m/z 373 [M+H]+; 1H-NMR (500 MHz, DMSO-d6): δ 6.73 (2H, s, H-2,6), 6.46 (1H, d,J= 15.9 Hz, H-7), 6.33 (1H, dt,J = 15.9, 5.1 Hz, H-8), 4.84 (1H, d,J = 7.5 Hz, glucosyl H-1), 4.11 (1H, dd,J = 5.1, 1.4 Hz, H-9a), 4.09 (1H, dd,J = 5.1, 1.4 Hz, H-9b), 3.77 (6H, s, 2 × OMe); 13C-NMR (125 MHz, DMSO-d6): δ 152.7 (C-3,5), 133.0 (C-4), 131.0 (C-7), 129.0 (C-8), 128.1 (C-1), 104.5 (C-2,6), 103.1 (Glc C-1), 77.4 (Glc C-5), 76.5 (Glc C-3), 74.9 (Glc C-2), 71.0 (Glc C-4), 62.0 (C-9), 60.5 (Glc C-6), 56.3 (OMe).Eleutheroside E (2): FAB-MS:m/z 743 [M+H]+; 1H-NMR (500 MHz, DMSO-d6): δ 6.67 (4H, s, H-2′,6′), 4.88 (2H, d,J = 7.3 Hz, glucosyl H-1), 4.67 (2H, d,J = 3.6 Hz, H-2), 4.28 (2H, dd,J = 8.5, 6.6 Hz, H-4eq), 4.20 (2H, dd,J = 8.5, 3.0 Hz, H-4ax), 3.76 (12H, s, 4 × OMe), 3.19 (2H, m, H-1); 13C-NMR (125 MHz, DMSO-d6): δ 153.2 (C-3′,5′), 138.1 (C-4′), 134.1 (C-1′), 104.6 (C-2′,6′), 103.3 (Glc C-1), 85.7 (C-2), 77.5 (Glc C-5), 76.7 (Glc C-3), 74.5 (Glc C-2), 72.1 (C-4), 70.2 (Glc C-4), 61.2 (Glc C-6), 57.0 (OMe), 54.2 (C-1).


## 2.5. Sample Preparation

To analyze the eleutherosides B (1) and E (2) content inA. divaricatus andA. koreanum, 5 g of each ofA. divaricatus andA. koreanum was extracted with 50% MeOH (3 × 100 mL) by reflux and evaporatedin vacuo. The residue was dissolved in 1 mL of MeOH and filtered with a 0.45 μm filter. The resulting solution was used for HPLC analysis.


## 2.6. HPLC Conditions

HPLC separation of eleutherosides B (1) and E (2) for qualitative and quantitative analysis was performed using a reverse phase system. A Discovery C18 (4.6 × 250 mm, 5 μm) column was used with a mobile phase that consisted of water and acetonitrile. A gradient solvent system of water and acetonitrile (90 : 10 to 70 : 30 for 20 min) was used for the elution program. UV detection was conducted at 350 nm. The injection volume was 10 μL and the flow rate was 1 mL/min. The temperature was maintained at 25°C. All injections were performed in triplicate.


## 2.7. Calibration Curve

A stock solution (1 mg/mL) of each eleutherosides B (1) and E (2) isolate was prepared in MeOH, and then the solution content was successively reduced to 50% to create different concentrations. The analyte contents were determined from the corresponding calibration curves. The calibration functions of the eleutherosides B (1) and E (2) isolate were calculated using the peak area (Y), concentration (X, μg/10 μL), and mean values (n=3) ± standard deviation (SD).


## 2.8. Statistical Analysis

Data for each sample was expressed as mean ± SD. ANOVA using the SAS Enterprise Guide software was calculated and the significance between the means of each group was determined using Duncan’s multiple test.


## 3. Results and Discussion

Content analysis of eleutherosides B (1) and E (2) inA. divaricatus andA. koreanum by plant section and cultivation method was conducted by HPLC. Eleutherosides B (1) and E (2) (Figure 2) have previously been isolated fromA. senticosus,A. divaricatus, andA. koreanum [12, 17, 18].Figure 2
Chemical structures of eleutherosides B (1) and E (2).HPLC was used to separate eleutherosides B (1) and E (2) for qualitative and quantitative analysis using a reverse phase system. HPLC conditions were selected to analyze eleutherosides B (1) and E (2) with good linearity (eleutheroside B, r2=0.9999; eleutheroside E, r2=0.9997). The contents of eleutherosides B (1) and E (2) inA. divaricatus andA. koreanum were determined, examining variations in plant section, pinching site, planting time, and fertilizer ratio, as described in Section 2. The HPLC chromatograms of eleutherosides B (1) and E (2) standards and ofA. divaricatus andA. koreanum 50% MeOH extracts are shown in Figure 3.HPLC chromatograms of standards of eleutherosides B (1) and E (2) (a) and of the MeOH extracts fromA. divaricatus grown with fertilizer ratio N-2P-K (b) and fromA. koreanum grown with fertilizer ratio 2N-2P-2K (c).
(a)
(b)
(c)The total eleutherosides B (1) and E (2) content in different plant sections (Ps-1 to -5) ranged from 2.466 to 5.841 mg/g inA. divaricatus and from 4.339 to 7.360 mg/g inA. koreanum (Table 1). The total eleutherosides B (1) and E (2) content in the upper part (Ps-1) was greater than that in the lower part (Ps-5) ofA. divaricatus, butthe result inA. koreanum was the opposite. The total eleutherosides B (1) and E (2) content was the highest in Ps-1 ofA. divaricatus and Ps-5 ofA. koreanum.Table 1
The contents of eleutherosides B (1) and E (2) in A. divaricatusandA. koreanumdivided by plant section.
Sample
Plant section*
Content (mg/g)
1
2
Total
A. divaricatus
Ps-1
—
5.841 ± 0.054a
5.841 ± 0.054
Ps-2
—
3.443 ± 0.147c
3.443 ± 0.147
Ps-3
—
3.922 ± 0.019b
3.922 ± 0.019
Ps-4
0.034 ± 0.013
3.078 ± 0.046d
3.112 ± 0.059
Ps-5
—
2.466 ± 0.060e
2.466 ± 0.060
A. koreanum
Ps-1
3.229 ± 0.101b
3.722 ± 0.008a
6.951 ± 0.109
Ps-2
2.482 ± 0.021e
1.857 ± 0.010d
4.339 ± 0.031
Ps-3
2.601 ± 0.020d
1.822 ± 0.004e
4.423 ± 0.024
Ps-4
3.080 ± 0.036c
2.365 ± 0.004c
5.445 ± 0.040
Ps-5
3.801 ± 0.013a
3.559 ± 0.020b
7.360 ± 0.033
*Plant sections are as shown in Figure 1(a).
Data are presented as the mean ± SD (n = 3) in mg/g of the dried samples.
Means followed by the different superscript letters (a–e) are significantly different by Duncan's multiple range tests.The total eleutherosides B (1) and E (2) content was found to be similar in the upper and lower parts ofA. divaricatus andA. koreanum when different cultivation methods were employed. Pinching (30 and 60 cm) yielded total eleutherosides B (1) and E (2) contents of 6.301–6.827 and 7.858–11.506 mg/g in the upper and lower parts ofA. divaricatus, respectively, while inA. koreanum, the total eleutherosides B (1) and E (2) contents were 4.439–7.847 and 3.886–4.161 mg/g in the upper and lower parts, respectively (Table 2). When planting time was varied (March 30, April 15, and April 30), the total eleutherosides B (1) and E (2) contents were found to be 4.290–6.526 and 3.769–10.083 mg/g in the upper and lower parts ofA. divaricatus, respectively, while the total eleutherosides B (1) and E (2) contents were found to be 4.546–5.693 and 3.655–6.529 mg/g in the upper and lower parts ofA. koreanum, respectively (Table 3). When fertilizer ratio was varied (N-P-K, 2N-P-K, N-2P-K, N-P-2K, and 2N-2P-2K), the total eleutherosides B (1) and E (2) contents were found to range from 4.417 to 6.905 and from 3.652 to 7.227 mg/g in the upper and lower parts ofA. divaricatus, respectively, and from 4.591 to 10.108 and from 3.834 to 9.079 mg/g in the upper and lower parts ofA. koreanum, respectively (Table 4).Table 2
The contents of eleutherosides B (1) and E (2) in A. divaricatusandA. koreanumcultivated by different pinching sites.
Sample
Pinching*
Content (mg/g)
1
2
Total
A. divaricatus
Upper part
30 cm
Trace
6.827 ± 0.007
6.827 ± 0.007
60 cm
Trace
6.301 ± 0.007
6.301 ± 0.007
Lower part
30 cm
Trace
11.506 ± 0.012
11.506 ± 0.012
60 cm
0.011 ± 0.008
7.847 ± 0.007
7.858 ± 0.015
A. koreanum
Upper part
30 cm
4.735 ± 0.521
3.112 ± 0.003
7.847 ± 0.524
60 cm
2.188 ± 0.025
1.698 ± 0.008
3.886 ± 0.033
Lower part
30 cm
3.099 ± 0.024
1.340 ± 0.003
4.439 ± 0.027
60 cm
2.497 ± 0.008
1.664 ± 0.007
4.161 ± 0.015
*Upper and lower parts of the plant are as shown in Figure 1(b).
Data are presented as the mean ± SD (n = 3) in mg/g of the dried samples.Table 3
The contents of eleutherosides B (1) and E (2) in A. divaricatusandA. koreanumplanted at varying times.
Sample
Planting time
Content (mg/g)
1
2
Total
A. divaricatus
Upper part
March 30
Trace
6.491 ± 0.012
6.491 ± 0.012
April 15
0.005 ± 0.002
4.285 ± 0.006
4.290 ± 0.008
April 30
0.016 ± 0.001
6.510 ± 0.015
6.526 ± 0.016
Lower part
March 30
—
3.769 ± 0.004
3.769 ± 0.004
April 15
0.002 ± 0.001
3.985 ± 0.007
3.987 ± 0.008
April 30
Trace
10.083 ± 0.012
10.083 ± 0.012
A. koreanum
Upper part
March 30
3.378 ± 0.012
2.315 ± 0.014
5.693 ± 0.026
April 15
2.436 ± 0.015
2.110 ± 0.016
4.546 ± 0.031
April 30
2.834 ± 0.007
2.349 ± 0.007
5.183 ± 0.014
Lower part
March 30
3.590 ± 0.033
2.939 ± 0.013
6.529 ± 0.046
April 15
1.887 ± 0.035
1.768 ± 0.012
3.655 ± 0.047
April 30
3.225 ± 0.028
2.915 ± 0.024
6.140 ± 0.052
Plant divisions and data presentation are as shown in Table2.Table 4
The contents of eleutherosides B (1) and E (2) in A. divaricatusandA. koreanumcultivated with varying fertilizer ratio.
Sample
Fertilizer ratio
Content (mg/g)
1
2
Total
A. divaricatus
Upper part
N-P-K
0.006 ± 0.003
5.623 ± 0.001
5.629 ± 0.004
2N-P-K
0.008 ± 0.001
5.280 ± 0.006
5.288 ± 0.007
N-2P-K
Trace
6.905 ± 0.021
6.905 ± 0.021
N-P-2K
0.018 ± 0.007
4.399 ± 0.022
4.417 ± 0.029
2N-2P-2K
Trace
5.711 ± 0.007
5.711 ± 0.007
Lower part
N-P-K
Trace
6.865 ± 0.007
6.865 ± 0.007
2N-P-K
0.012 ± 0.006
3.640 ± 0.008
3.652 ± 0.014
N-2P-K
0.013 ± 0.004
7.214 ± 0.022
7.227 ± 0.026
N-P-2K
Trace
7.011 ± 0.008
7.011 ± 0.008
2N-2P-2K
Trace
6.976 ± 0.006
6.976 ± 0.006
A. koreanum
Upper part
N-P-K
4.798 ± 0.030
3.101 ± 0.030
7.899 ± 0.060
2N-P-K
3.203 ± 0.022
2.902 ± 0.030
6.105 ± 0.052
N-2P-K
2.137 ± 0.024
2.454 ± 0.008
4.591 ± 0.032
N-P-2K
3.473 ± 0.017
2.451 ± 0.012
5.924 ± 0.029
2N-2P-2K
6.314 ± 0.007
3.794 ± 0.022
10.108 ± 0.029
Lower part
N-P-K
2.083 ± 0.020
2.848 ± 0.027
4.931 ± 0.047
2N-P-K
1.615 ± 0.039
2.219 ± 0.006
3.834 ± 0.045
N-2P-K
2.707 ± 0.044
2.494 ± 0.005
5.201 ± 0.049
N-P-2K
1.855 ± 0.018
2.088 ± 0.015
3.943 ± 0.033
2N-2P-2K
4.918 ± 0.018
4.161 ± 0.008
9.079 ± 0.026
Plant divisions and data presentation are as shown in Table2.A previous paper reported that eleutherosides B (1) and E (2) contents were 0 and 1.804 mg/g in the stem ofA. divaricatus, 0 and 1.016 mg/g in the root ofA. divaricatus, 0.621 and 0.885 mg/g in the stem ofA. koreanum, and 0.478 and 0.538 mg/g in the root ofA. koreanum, respectively [11]. Our results showed a similar trend.The best conditions to increase the eleutherosides B (1) and E (2) content inA. divaricatus were determined to be with 30 cm pinching, April 30 planting time, and a N-2P-K fertilizer ratio. In comparison, forA. koreanum, the eleutherosides B (1) and E (2) content was obtained with 30 cm pinching, March 30 planting time, and a 2N-2P-2 K fertilizer ratio. Moreover, the total eleutherosides B (1) and E (2) contents in the upper part of the plant were greater than in the lower part ofA. divaricatus by pinching and fertilizer ratio and in the lower part of the plant they were greater than in the upper part ofA. divaricatus by planting time. However, the result of theA. koreanum was the opposite. Therefore, the cultivation methods ofAcanthopanax species should be established on an individual basis for each species considered. In conclusion, these results can be applied to optimize eleutherosides B (1) and E (2) production for harvesting fromA. divaricatus andA. koreanum in nutraceutical, pharmaceutical, and cosmeceutical development.



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