SUPPLEMENTARY MATERIAL
Antiurease Activity of Plants Growing in the Czech Republic
Petra Hřibováa, Elian Khazneha, Milan Žemličkaa, Emil Švajdlenkaa, Mohammed M.
Ghoneimb, c, Khaled M. Elokelyd and Samir A. Rossb,e,*
aDepartment
of Natural Drugs, Faculty of Pharmacy, University of Veterinary and
Pharmaceutical Sciences Brno, Palackého 1-3, 612 42 Brno, Czech Republic
b
National Center for Natural Products Research, The University of Mississippi, University,
MS 38677, USA.
cDepartment
of Pharmacognosy, Faculty of Pharmacy, The University of Al-Azhar, Cairo
11371, Egypt.
dDepartment
of Medicinal Chemistry, School of Pharmacy, The University of Mississippi,
University, MS 38677, USA.
eDepartment
of Pharmacognosy, School of Pharmacy, The University of Mississippi,
University, MS 38677, USA.
*Corresponding author. Tel.: +1-662-915-1031; fax: +1-662-915-7989
E-mail address: sross@olemiss.edu (S.A. Ross)
Abstract
The antiurease activity of the aqueous extracts of 42 plants growing in the Czech Republic
was investigated. A phenol-hypochlorite reaction was used for determination of ammonia
produced by urease. The inhibitory activity of the extracts at concentration of 0.2 mg/ml
varied from 17.8% to 80.0%. Extracts from six Potentilla species showed inhibitory activity
against jack bean urease. They were further investigated for their phenolic constituents and
the major compounds were submitted to molecular docking. The results showed that both jack
bean urease and Helicobacter pylori urease were inhibited by quercetin-3-O-β-Dgalactopyranoside-6´´-gallate (1), myricetin-3-O-β-D-glucuronide (2), tiliroside (3) and Btype procyanidin (4). The antiurease activity of the investigated Potentilla species is probably
due to the presence of complex phenolic constituents such as flavonoid glycosides and
catechin dimers.
Key words: Docking; Phenolic constituents; Phenol-hypochlorite; Potentilla species; Urease.
Table S1:
Plant Name*
Aesculus carnea (leaves)
Allium ursinum (flowers)
Allium ursinum (leaves)
Catalpa bignonioides (leaves)
Chelidonium majus
Chrysanthemum balsamita
Clematis recta
Clinopodium vulgare
Echium vulgare
Eruca sativa
Fagopyrum aesculentum
Hyssopus officinalis
Inula britanica
Lamium album
Lyriodendron spp.
Magnolia spp. (leaves)
Marubium vulgare
Meum athamanticum
Nigella damascena
Nigella integrifolia
Nonea pulla
Origanum vulgare
Potentilla anserina
Potentilla arenaria
Potentilla argentea
Potentilla aurea
Potentilla erecta
Potentilla fruticosa
Prunella grandiflora
Salvia sclarea
Solidago virgaurea
Sophora japonica
Verbena officinalis
Yucca filamentosa
Acetohydroxamic acid
Herbarium No.
PAC-07
PAC-08
PAC-09
PAC-11
PAC-14
PAC-15
PAC-16
PAC-17
PAC-21
PAC-22
PAC-23
PAC-24
PAC-27
PAC-28
PAC-29
PAC-30
PAC-31
PAC-32
PAC-33
PAC-34
PAC-35
PAC-36
PAC-49
PAC-50
PAC-51
PAC-52
PAC-53
PAC-54
PAC-37
PAC-38
PAC-39
PAC-40
PAC-41
PAC-42
Family
Hippocastanaceae
Liliaceae
Liliaceae
Bignoniaceae
Papaveraceae
Asteraceae
Ranunculaceae
Lamiaceae
Boraginaceae
Brassicaceae
Polygonaceae
Lamiaceae
Asteraceae
Lamiaceae
Magnoliaceae
Magnoliaceae
Lamiaceae
Apiaceae
Ranunculaceae
Ranunculaceae
Boraginaceae
Lamiaceae
Rosaceae
Rosaceae
Rosaceae
Rosaceae
Rosaceae
Rosaceae
Lamiaceae
Lamiaceae
Asteraceae
Fabaceae
Verbenaceae
Asparagaceae
-
Urease Inhibition ± SEM† [%]
71.0 ± 3.1
67.2 ± 3.6
79.2 ± 0.3
67.4 ± 1.5
62.3 ± 1.0
80.0 ± 0.6
62.0 ± 2.2
68.3 ± 3.2
70.8 ± 2.9
70.5 ± 2.4
71.9 ± 1.4
74.3 ± 2.5
62.0 ± 2.3
61.1 ± 1.8
66.3 ± 3.6
60.9 ± 5.9
61.2 ± 4.9
74.0 ± 3.1
64.2 ± 2.4
69.6 ± 2.6
71.9 ± 1.7
69.7 ± 1.5
73.7 ± 1.9
44.4 ± 0.3
76.0 ± 0.9
56.9 ± 4.2
72.8 ± 2.6
72.2 ± 5.0
63.4 ± 1.6
78.0 ± 3.0
75.6 ± 3.1
60.7 ± 1.9
67.4 ± 7.6
79.0 ± 1.1
72.6 ± 3.6
*Unless otherwise noted in parenthesis, the aerial part of the flowering plant was tested.
†Standard mean error from the three determinations.
Plant extracts that did not show significant antiurease activity: Borago officinalis (PAC-10), Catalpa
ovata (leaves) (PAC-12), Centaurea segetum (PAC-13), Dalanum segetum (PAC-18), Dentaria
bulbifera (PAC-19), Duchesnea indica (PAC-20), Impatiens balsamita (PAC-25), Indigofera tinctoria
(PAC-26).
Table S2:
tR [min]
Maxima UV [nm]
[M-H]- m/z
HHDP-glucose
0.9
228, 262 sh
481
301, 275
a,b,c,d,e,f
bis-HHDP-glucose
1.0
234, 250 sh
783
765, 722.5
a,b,c,d,e,f
epigallocatechin gallate
1.3
230, 274
457
441
b,c,d,e,f
bis-HHDP-glucose
2.1
234, 250 sh
783
481, 301
a,c,d,f
HHDP-galloyl-glucose
2.4
224, 270 sh
633
301
a,b,c,d,e,f
HHDP-galloyl-glucose
2.7
224, 270 sh
633
481, 301, 257
d,f
proanthocyanidin dimer (Procyanidin B)
3.4
222, 278
577
451, 425, 407, 289
a,b,c,d,f
catechin
3.6
226, 278
289
245, 205
f
proanthocyanidin dimer (Procyanidin B)
4.6
222, 278
577
451, 425, 407, 289
a,b,f
HHDP-galloyl-glucose
4.9
228, 270 sh
633
498, 301
c
bis-HHDP-glucose
5.0
230, 250 sh
783
481, 301
a,f
quercetin-diglucuronide
5.1
256, 350
653
477, 301
b,e
quercetin-dihexoside
5.3
256, 352
625
463, 301
d
quercetin-hexoside-glucuronide
256, 354
639
463, 301
b,d
luteolin-diglucuronide
5.6
6.3
256, 348
637
461, 285
b,e
(iso)rhamnetin-diglucuronide
6.6
254, 352
667
491, 315, 255
b,e
rutin
7.2
266, 344
609
463
d
myricetin-3-O-β-D-glucuronide
quercetin-3-O-β-D-galactopyranoside6''-gallate
9.1
256, 354
493
317, 259
a,b,d
9.6
254, 368
615
463, 301
a,b,c,e,f
quercetin-hexoside
9.9
256, 356
463
301
a,b,c,d,f
quercetin-glucuronide
10.0
256, 352
477
301, 179
c,f
kaempferol-hexoside
11.0
266, 348
447
285
f
(iso)rhamnetin-hexoside
11.0
256, 352
477
315, 165
f
(iso)rhamnetin-pentoside
kaempferol-3-O-coumaroylglucoside
(tiliroside)
11.8
256, 352
447
315
f
14.4
268, 314
593
285
a,c,f
Compound
MS2
Occurence
HPLC-DAD-MS2 data for compounds found in water extracts of Potentilla spp.
a P. anserina, b P. arenaria, c P. argentea, d P. aurea, e P. erecta, f P. fruticosa; sh =
shoulder; HHDP = hexahydroxydiphenyl-glucose.
Figure S1: Compounds 1-4
100
0
2
4.575
4
6
8
10
12
0
14
17.081
14.005
12.510
12.739
14.420
10.984
11.287
11.487
11.679
11.818
11.964
12.180
2
10.508
9.643
1
10.780
600
10.308
4.186
0.932
mAU
10.9.092252
9.145
9.275
9.503
7.966
8.080
8.239
8.407
8.58.89518
8.747
7.080
7.294
7.454
7.623
6.861
6.058
6.369
6.520
2.083
4
4.966
5.139
5.393
3.402
3.640
2.435
2.721
1.453
1.624
1.814
1.085
200
1.295
0.777
Figure S2: Chromatogram of Potentilla fruticosa with the major peak of quercetin-3-O-β-D-galactopyranoside-6''-gallate (1)
DAD1A, Sig=254,4Ref=400,50(C:\USERS\P...\PETRAH_ANTIUREASE\EXPRESSRHUS2013-07-0718-59-32\056-1601.D)
500
400
300
3
16
min
Figure S3. The binding modes of the compounds in the Helicobacter pylori urease binding
pocket. The ligand is shown as sticks with grey carbons. The binding pocket is displayed as
partial charge colored surface. The compounds fit well inside the binding pocket (compound 1
up to the left, compound 2 up to the right, compound 3 down to the left and compound 4
down to the right). The figure was prepared by PyMol.
Figure S4. The 2D interaction profile of compound 1 in the active site of jack bean urease
(left) and Helicobacter pylori (right). The interaction is much more stronger in case of
Helicobacter pylori and mostly as hydrogen bonding, ion–dipole and hydrophobic
interactions.