Hindawi Publishing Corporationdownloads.hindawi.com/journals/omcl/2019/5465463.f… · Web viewThe results of univariate analysis of variance Groups Counter Sum Average Variance Emodin

Supplementary Material

Phytochemical and antioxidant studies on a rare Rheum cordatum

Losinsk. species from Kazakhstan

Gulsim Zhumashova, Wirginia Kukula-Koch,* Wojciech Koch, Tomasz Baj, Galiya Sayakova, Alma Shukirbekova, Kazimierz Głowniak and Zuriyadda Sakipova1

Figure S1. Compositional variation of the root extracts in relation to the collection date

Figure S2. The differences in the composition of ethanolic extracts in various organs of Rheum cordatum (TIC chromatograms recorded in the negative ionisation mode)

Figure S3. The influence of extractant on the composition of the Rheum cordatum root extract (1- 98% ethanol, 2- 50% ethanol, 3-dichloromethane, 4-water, 5-chloroform extracts)

Figure S4. The MS/MS spectra of the selected major constituents of the extractsAcetyl aloe-emodin

2x10

00.5

11.5

22.5

33.5

44.5

55.5

66.5

77.5

88.5

9-ESI Product Ion (rt: 28.032 min) Frag=170.0V [email protected] (311.0608[z=1] -> **) 2R0518_20_neg.d

311.

0579

269.

0477

224.

0539

248.

0819

169.

0687

298.

0645

197.

3009

255.

0881

114.

6740

129.

0348

238.

6512

158.

0525

230.

5508

361.

7164

149.

0927

324.

2470

216.

2753

338.

3396

Counts vs. Mass-to-Charge (m/z)120 140 160 180 200 220 240 260 280 300 320 340 360

Aloeemodin3x10

0

0.5

1

1.5

2

2.5


269.

0456

225.

0517

341.

0795

180.

0617

125.

0250

485.

2378

829.

1705

927.

7012

392.

1866

1052

.135

3

433.

1833

762.

6471

Counts vs. Mass-to-Charge (m/z)100 200 300 400 500 600 700 800 900 1000 1100

Apigenin galloyl glucose3x10

00.20.40.60.8

11.21.41.61.8

22.22.42.62.8

33.2

-ESI Product Ion (rt: 24.641 min) Frag=170.0V [email protected] (583.1185[z=1] -> **) 2R0518_20_neg.d

583.

1060

169.

0111

269.

0388

435.

0940

341.

0273

311.

9866

125.

0168

417.

0839

225.

0238

651.

2875

151.

0375

187.

0416

Counts vs. Mass-to-Charge (m/z)100 150 200 250 300 350 400 450 500 550 600 650

Apigenin glucoside3x10

0

0.5

1

1.5

2

2.5

3

3.5

4

4.5

5-ESI Product Ion (rt: 18.507 min) Frag=170.0V [email protected] (431.0978[z=1] -> **) 2RCE_20_neg.d

311.

0582

431.

1011

341.

0669

283.

0588

323.

0557

413.

1027

191.

0417

353.

0742

458.

3453

161.

0170

440.

1874

269.

0418

383.

0870

Counts vs. Mass-to-Charge (m/z)80 100 120 140 160 180 200 220 240 260 280 300 320 340 360 380 400 420 440 460 480

Catechin Chrysophanol

3x10

00.25

0.5

0.751

1.251.5

1.75

22.25

2.52.75

3

3.253.5

-ESI Product Ion (rt: 13.485 min) Frag=160.0V [email protected] (289.0698[z=1] -> **) 2RCE_20_neg.d

289.

0687

245.

0792

179.

0300

125.

0250

151.

0319

109.

0339

203.

0698

230.

0588

165.

0197

425.

9288


3x10

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

1.8

2


253.

0508

225.

0555

139.

1122

145.

0375

182.

0392

245.

3283

207.

0473

196.

0511

276.

7628

129.

5038

263.

2006

221.

6105

310.

8081

Counts vs. Mass-to-Charge (m/z)120 130 140 150 160 170 180 190 200 210 220 230 240 250 260 270 280 290 300 310

Emodin4x10

0

0.20.40.60.8

11.2

1.41.61.8

2


269.

0518

225.

0598


Emodin acetyl hexose3x10

00.10.20.30.40.50.60.70.80.9

11.11.21.31.41.51.61.7


473.

1163

269.

0467

577.

4017

254.

0474

311.

0512

282.

0512

186.

0601

413.

0831

431.

1480

456.

1063

349.

9640

495.

4271

201.

0663

397.

2875

508.

5674

128.

6230

542.

1300

146.

8610

160.

0873

328.

5281

559.

4084

375.

2971


Emodin galloylglucose3x10

00.20.40.60.8

11.21.41.61.8

22.22.42.62.8

33.2


583.

1060

169.

0111

269.

0388

435.

0940

341.

0273

311.

9866

125.

0168

417.

0839

225.

0238

651.

2875

151.

0375

187.

0416

Counts vs. Mass-to-Charge (m/z)150 200 250 300 350 400 450 500 550 600 650 700

Emodin glucoside3x10

0

0.5

1

1.5

2

2.5

3

3.5

4

4.5

5

5.5

-ESI Product Ion (rt: 23.986 min) Frag=170.0V [email protected] (431.1022[z=1] -> **) 2RST_20_neg.d

431.

1008

269.

0461

311.

0530

413.

1082

293.

0537

224.

0439


Emodin malonyl hexose3x10

0

0.5

1

1.5

2

2.5

3

3.5

4

4.5

5

5.5

6


473.

0726

517.

0615

269.

0252

431.

0502


Epicatechin3x10

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

1.8

2

2.2


289.

0740

245.

0795

205.

0528

181.

0293

123.

6484

139.

0424

165.

0176

221.

1489

109.

0301

150.

7214

355.

2125

229.

0402

301.

9061

101.

2901

Counts vs. Mass-to-Charge (m/z)80 100 120 140 160 180 200 220 240 260 280 300 320 340

Epicatechin gallate Epigallocatechin gallate

3x10

00.5

11.5

22.5

33.5

44.5


451.

1127

407.

1273

305.

0673

1089

.745

7

167.

0493

125.

0251

242.

0424


3x10

0

1

2

3

4

5

6


169.

0140

457.

0782

305.

0666

125.

0250

261.

0771

331.

0559

Counts vs. Mass-to-Charge (m/z)150 200 250 300 350 400 450 500 550 600 650 700 750

Gallic acid3x10

00.250.5

0.751

1.251.5

1.752

2.252.5


125.

0238

169.

0113

227.

9971


Gallic acid hexoside3x10

00.25

0.50.75

11.25

1.51.75

22.25

2.52.75

33.25

3.53.75

44.25


169.

0125

331.

0639

125.

0267


Kaempferol3x10

00.250.5

0.751

1.251.5

1.752

2.252.5


285.

0353

151.

0021

213.

0501

257.

0364

107.

0137

240.

0416

135.

0108

195.

0415

173.

1125

516.

6141

Counts vs. Mass-to-Charge (m/z)100 150 200 250 300 350 400 450 500

Quercetin3x10

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

1.8

2

2.2


151.

0022

178.

9969

301.

0299

107.

0116

273.

0358

121.

0313

215.

0286

227.

0297

240.

0295

190.

0167

257.

0383

143.

0721

Counts vs. Mass-to-Charge (m/z)120 140 160 180 200 220 240 260 280 300 320 340 360 380 400 420 440 460 480

Tetrahydroxystilbene-O-(acetyl)-hexose3x10

00.20.40.60.8

11.21.41.61.8

22.22.42.62.8

33.23.4


285.

0738

447.

1281

242.

0530

227.

0326

270.

0489

144.

9902

402.

6246

349.

0662

Counts vs. Mass-to-Charge (m/z)125 150 175 200 225 250 275 300 325 350 375 400 425 450 475 500 525 550 575

Table S1: Content of the main constituents determined in the particular plant parts, depending on the solvents used.

Leaves Seeds stems Radix 2017 Radix04.2018

Radix 05.2018

Etha

nol e

xtra

ct

Emodin 0.0614±0.0009 0.0694±0.0017 0.6581±0.0056 0.5174±0.0043 0.3626±0.003 0.8023±0.0152

Aloeemodin

0.0037±0.00006

0.0029±0.00002 0.0135±0.00008 0.0889±0.0021 0.0049±0.0007 0.0089±0.0001

ECG 0.8552±0.0264 0.7824±0.0147 0.0509±0.0011 2.93±0.0921 2.5052±0.0276 2.38±0.0316

EGCG 0.0151±0.0002 0.24±0.0064 0.0001±0.000003 0.4394±0.0034 0.2467±0.0035 0.1265±0.005EC 0.0304±0.0009 0.0736±

0.00270.0116±0.00008 0.0274±0.0012 0.0114±0.0002 0.1100±0.0042

Rutoside 2.108±0.0244 1.614±0.0361 0.1504±0.0026 0.0327±0.0002 0.0155±0.0003 0.0160±0.0003

Kaempferol glucoside

0.0410±0.0004 0.0032±0.0001 0.0043±0.00006 0.0433±0.001 0.0428±0.0006 0.0225±0.0001

DPPH <LOQ 89.80±1.97 <LOQ 92.05±0.39 92.3±0.69 68.13±4.18TPC <LOQ 460±24.70 <LOQ 530.2±37.65 430.2±28.67 720.3±34.22

Etha

nol:w

ater

Emodin 0.5420±0.0114 0.1416±0.0012 0.4482±0.0194 1.29±0.0068 0.9650±0.027 0.7956±0.022Aloeemodin

0.0025±0.00002

<LOQ 0.0293±0.0003 0.1273±0.0007 0.0539±0.0036 0.017±0.0003

ECG 2.1589±0.0131 2.0829±0.0031 1.6876±0.0132 3.084±0.0319 3.3669±0.0262 5.004±0.1265

EGCG 0.2350±0.0017 0.6432±0.0195 0.0363±0.0003 0.2212±0.0028 0.3596±0.0026 0.24±0.0015EC 0.0464±0.0005 0.0416±0.0004 0.0873±0.005 0.0213±0.0000

40.0222±0.0005 0.081±0.0033

Rutoside 0.0757±0.0009 2.0235±0.0106 0.8525±0.009 0.0232±0.00003

0.4144±0.0064 0.022±0.0005


0.1564±0.0051 0.0466±0.0001 0.1408±0.0021 0.0549±0.0017 0.0159±0.0007 0.021±0.0003

DPPH 89.96±0.4559 89.06±1.54 92.74±0.9577 88.48±0.68 89.69±1.63 83.7±2.08TPC 719.8±26.61 669.8±19.86 540.2±30.74 480.3±30.07 462±23.56 620.7±39.93

Dich

loro

met

hene


0.0002±0.00002

<LOQ <LOQ <LOQ <LOQ <LOQ

ECG 0.0032±0.0002 0.0022±0.00002 0.0403±0.0021 0.0019±0.00004

0.0045±0.00009 0.0014±0.00015

EGCG <LOQ 0.0003±0.00002 0.0008±0.00005 0.0002±0.000006

0.0003±0.00002 0.00009±0.00001

EC 0.0007±0.00002

0.0002±0.000002

0.0006±0.00004 0.0004±0.00003

0.0003±0.00002 0.0003±0.00002

Rutoside 0.0106±0.0002 0.0014±0.00005 0.0059±0.0001 0.0003±0.00003

0.0018±0.00001 <LOQ


<LOQ 0.0005±0.00001 0.0019±0.00006 <LOQ <LOQ <LOQ

DPPH <LOQ <LOQ <LOQ <LOQ <LOQ <LOQTPC <LOQ <LOQ <LOQ <LOQ <LOQ <LOQEmodin 0.0197±0.0001 0.1898±0.001 0.06±0.0008 0.8431±0.0308 0.5222±0.0337 0.3369±0.011

Aloeemodin

<LOQ 0.0029±0.000009

0.0027±0.00005 0.0831±0.001 0.0257±0.0028 0.011±0.00008

ECG 1.424±0.0111 1.7212±0.0035

0.6364±0.0134 2.5042±0.0131 2.76±0.1116 2.29±0.0196

EGCG 0.1107±0.0005 0.7922±0.0097 0.0144±0.00004 0.1564±0.0004 0.2335±0.0014 0.1224±0.0028EC 0.0087±0.0000

40.0557±0.0021 0.0341±0.0002 0.0186±0.0000

90.0178±0.0003 0.0421±0.0009

Water

Rutoside 4.8664±0.0604 1.9050±0.0102 0.5992±0.0171 0.0211±0.0003 0.0157±0.0006 0.015±0.0004Kaempferol glucoside

0.1299±0.0016 0.0181±0.00005 0.0567±0.002 0.014±0.00006 0.011±0.0013 0.0049±0.00007

DPPH 45.9±1.38 88.23±1.26 21.83±8.42 82.79±2.07 91.59±1.11 90.6±1.00TPC 279.8±37.35 470.3±18.75 90.33±9.52 320±31.55 498±19.71 751.8±34.78

Chlo

rofo

rm


<LOQ <LOQ <LOQ <LOQ <LOQ <LOQ

ECG 0.0024±0.00002

0.0019±0.00002 0.0035±0.00005 0.0034±0.00005

0.0011±0.00005 0.8647±0.0545

EGCG 0.0001±0.000005

0.0002±0.000001

0.0002±0.00001 0.0006±0.00004

0.0002±0.00002 0.0193±0.00005

EC 0.0001±0.00002

0.0008±0.00003 0.0006±0.0001 <LOQ 0.0003±0.00003 0.0459±0.0015

Rutoside 0.0007±0.0001 0.0005±0.00001 0.0005±0.00006 0.0004±0.00001

0.0039±0.0002 0.0023±0.00005


0.0041±0.00003

<LOQ <LOQ 0.0042±0.00008

0.0102±0.0007 0.0076±0.0003

DPPH <LOQ <LOQ <LOQ <LOQ 21.71±7.36 <LOQTPC <LOQ <LOQ <LOQ <LOQ <LOQ <LOQ

Table S2. The correlations between the single metabolites determined in the extracts

I% Emodin Aloeemodin ECG EGCG EC RutosideKaempferol glucoside

I% 1

Emodin-

0.0173 1

Aloeemodin 0.3448 0.259271 1

ECG 0.8852 -0.05801 0.256912 1

EGCG 0.7165 -0.12694 -0.11936 0.695647 1

EC 0.8673 -0.03761 0.589719 0.721823 0.48493 1

Rutoside 0.3956 -0.35367 -0.05216 0.538240.38586

4 0.256191 1

Kaempferol glucoside 0.5809 0.048616 0.424163 0.759295

0.094772 0.489354

0.415671 1

Table S3. The results of univariate analysis of variance

Groups Counter Sum Average Variance

Emodin 15 3.962698 0.26418 0.10624

Aloeemodin 15 0.0578 0.003853 6.15E-05

ECG 15 11.45336 0.763558 0.703273

EGCG 15 2.088774 0.139252 0.062664

EC 15 0.392295 0.026153 0.000861

Rutoside 15 14.21484 0.947656 1.864206

Kaempferol glucoside 15 0.603319 0.040221 0.003165

ANALYSIS OF VARIANCE

Source of variance SS df MS F p value Test F

between groups13.3592

7 6 2.226544 5.687277 4.18242E-05 2.192518

inside groups38.3665

7 98 0.391496

Sum51.7258

4 104

Figure S5. Emodin variation graph depending on the solvent used

Figure S6. Emodin variation plot depending on the part of the plant used.

Table S4. The list of secondary metabolites present in other species of Rheum or Polygonum gender, which presence was not confirmed in the herein studied extracts by HR-MS studies.

Name NamePhyscion GentisinRhein Aloeemodin acetyl-

hexosideChrysophanol hexoside HydroxyemodinDimer of rhein and emodin and acetyl-hexose

Rhapontigen hexoside

Rhein Resveratrol glucosideRhein hexoside Emodin malonyl-

glucosideChrysophanol acetyl-hexoside

Dimer of emodin and chrysophanol

Documents

Hindawi Publishing Corporationdownloads.hindawi.com/journals/omcl/2019/5465463.f… · Web viewThe results of univariate analysis of variance Groups Counter Sum Average Variance Emodin