Quinic acid derivatives from Pimpinella brachycarpa exert anti-neuroinflammatory activity in lipopolysaccharide-induced microglia

Article abstract of DOI:10.1016/j.bmcl.2013.01.115

Five new quinic acid derivatives (1-5), together with 10 known quinic acid derivatives (6-15), were isolated from the MeOH extract of Pimpinella brachycarpa (Umbelliferae). Their structures were established on the basis of spectroscopic analyses including extensive 2D NMR studies (COSY, HMQC and HMBC). Isolated compounds 1-15 were evaluated for their inhibitory activities on nitric oxide (NO) production in an activated murine microglial cell line. Compounds 2, 3, 8 and 11 significantly inhibited NO production without high cell toxicity in lipopolysaccharide (LPS)-activated BV-2 cells, a microglia cell line (IC₅₀ = 4.66, 12.52, 9.04 and 12.11 μM, respectively).

Full text of DOI:10.1016/j.bmcl.2013.01.115

Bioorganic & Medicinal Chemistry Letters 23 (2013) 2140–2144
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Quinic acid derivatives from Pimpinella brachycarpa exert
anti-neuroinflammatory activity in lipopolysaccharide-induced
microglia
Seung Young Lee a, Eunjung Moon b, Sun Yeou Kim b,c, Kang Ro Lee a,
⇑
a Natural Products Laboratory, School of Pharmacy, Sungkyunkwan University, 300 Chonchon-dong, Jangan-ku, Suwon, Gyeonggi-do 440-746, Republic of Korea
b Gachon Institute of Pharmaceutical Science, Gachon University, #191 Hambakmoe-ro, Yeonsu-gu, Incheon 406-799, Republic of Korea
c College of Pharmacy, Gachon University, #191 Hambakmoe-ro, Yeonsu-gu, Incheon 406-799, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
Five new quinic acid derivatives (1–5), together with 10 known quinic acid derivatives (6–15), were isolated
from the MeOH extract of Pimpinella brachycarpa (Umbelliferae). Their structures were established on the
basis of spectroscopic analyses including extensive 2D NMR studies (COSY, HMQC and HMBC). Isolated
compounds 1–15 were evaluated for their inhibitory activities on nitric oxide (NO) production in an
activated murine microglial cell line. Compounds 2, 3, 8 and 11 significantly inhibited NO production
Received 7 November 2012
Revised 21 January 2013
Accepted 24 January 2013
Available online 8 February 2013
without high cell toxicity in lipopolysaccharide (LPS)-activated BV-2 cells,
a microglia cell line
Keywords:
Pimpinella brachycarpa
Umbelliferae
(IC50 = 4.66, 12.52, 9.04 and 12.11
lM, respectively).
Ó 2013 Elsevier Ltd. All rights reserved.
Quinic acid derivative
Anti-inflammatory effect
Microglia, which is a type of neuroglia, is similar to macro-
phages in the CNS. Activated microglia secrete several inflamma-
tory mediators such as NO and TNF-alpha. Overproduction of
toxic mediators by microglia can induce pathogenesis of several
neurodegenerative diseases such as ischemia, Alzheimers’s disease,
Parkinsons’s disease and Multiple sclerosis.1
H
OR2
6
O
5
4
OH
H
H
2
7
HO
1
3
Based on the assays in search for anti-inflammatory activities
from different plant sources, the MeOH extract from the aerial
parts of Pimpinella brachycarpa revealed significant anti-neuroin-
flammatory activities in BV-2 cells.
OR1
OH
R1
R2
Pimpinella brachycarpa (Umbelliferae) is widely distributed in
Europe, Africa, and Asia.2 In Korea, P. brachycarpa, called as cham-
na-mul, is well known as a vegetable and a popular edible herb.3 This
species has also been used in Korean folk medicine for the treatment
of gastrointestinal disturbances, bronchial asthma, insomnia, and
persistent cough.4 In previous phytochemical studies, terpenes,4
flavonoids,5 and essential oil components6 were isolated from this
source. The MeOH extract of P. brachycarpa has exhibited antibacte-
rial, antioxidative, anti-proliferative,3 antifungal,7 and antithrom-
botic activities8 in a previous study. Also, the EtOH extract of
P. brachycarpa showed positive effects on ethanol-induced chronic
hepatotoxicity in rat liver and on lipid metabolism in rats fed with
a high cholesterol diet.4,9 Quinic acid derivatives are a large family
O
OCH3
1
2
3
4
5
trans-caffeoyl
trans-caffeoyl
A
trans-coumaroyl
cis-coumaroyl
trans-coumaroyl
A
trans-caffeoyl
cis-coumaroyl
cis-coumaroyl
OH
OH
A:
Figure 1. Chemical structures of compounds 1–5.
of esters formed between quinic acid and one or more of several phe-
nylpropanoic acids such as caffeic acid, ferulic acid, coumaric acid,
sinapic acid and cinnamic acid.10 Quinic acid derivatives have vari-
ous beneficial effects including antioxidant, anti-inflammatory,
anti-HIV, anti-hepatitis B virus, hypoglycaemic, and hepatoprotec-
tive activities and inhibition of mutagenesis and carcinogenesis.11–16
The aim of the present study was to identify the potential
components related to the anti-inflammatory activity of the MeOH
extract from P. brachycarpa on BV-2 microglial cells. Column
⇑
Corresponding author. Tel.: +82 31 290 7710; fax: +82 31 290 7730.
E-mail address: krlee@skku.ac.kr (K.R. Lee).
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.01.115

S. Y. Lee et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2140–2144
2141
Table 2
chromatographic separation of the MeOH extract from the aerial
13C NMR data of compounds 1–5 (in CD3OD, 125 MHz, d in ppm)
parts of P. brachycarpa furnished five new quinic acid derivatives
(1–5) (Fig. 1), together with 10 known quinic acid derivatives (6–
15). The structures of these new compounds were elucidated on
the basis of 1D and 2D NMR spectroscopic data analyses (1H, 13C
NMR, COSY, HMQC and HMBC), as well as chemical means. Also,
we evaluated the inhibitory effect of the isolated compounds (1–
15) from P. brachycarpa on NO production in LPS-activated BV-2
cells, a microglial cell line.
Position
1
2
3
4
5
1
2
3
4
5
6
79.8
34.4
68.2
71.6
70.4
35.7
80.1
34.5
68.3
71.6
70.7
35.8
79.7
34.2
68.2
71.5
70.4
35.2
79.8
34.2
68.3
71.7
69.8
35.8
79.8
34.0
68.2
71.5
69.9
35.9
173.7
126.2
132.3
114.4
158.7
114.4
132.3
143.9
115.4
166.1
7
173.7
126.0
114.0
145.9
148.4
115.3
121.8
146.2
114.2
166.8
173.8
126.7
113.9
145.6
148.4
115.3
121.9
146.2
114.2
166.8
173.6
132.1
113.4
145.4
145.1
115.1
118.3
79.8
173.7
125.8
129.8
115.3
159.8
115.3
129.8
145.6
114.0
166.6
10
Compound (1) was isolated as a colorless gum. The molecular
formula C25H25O11 was determined by the positive ion HRFABMS
m/z 501.1400 [M+H]+ (calcd for 501.1397). The 1H NMR spectrum
of 1 (Table 1) showed the presence of three oxygenated methine
proton signals at dH 5.38 (1H, m, H-5), 4.29 (1H, m, H-3) and
3.78 (1H, m, H-4) and two methylene proton signals at dH 2.57,
2.07 (2H, m, H2-6) and 2.44, 2.43 (2H, m, H2-2). In the 13C NMR
spectrum (Table 2), 7 carbon signals appeared, including oxygen-
ated three methine carbons at dC 71.6 (C-4), 70.4 (C-5) and 68.2
(C-3), two methylene carbons at dC 35.7 (C-6) and 34.4 (C-2), one
quaternary carbon at dC 79.8 (C-1) and one carbonyl carbon at dC
173.7 (C-7). These spectral data implied that 1 could be a quinic
acid derivative.17,18 In addition, 1H (Table 1) and 13C (Table 2)
NMR spectra of 1 showed trans-caffeoyl group signals19 [dH 7.57
(1H, d, J = 16.0 Hz, H-70), 7.05 (1H, d, J = 2.0 Hz, H-20), 6.96 (1H,
dd, J = 8.5, 2.0 Hz, H-60), 6.78 (1H, d, J = 8.5 Hz, H-50) and 6.29
(1H, d, J = 16.0 Hz, H-80); dC 166.8 (C-90), 148.4 (C-40), 146.2
(C-70), 145.9 (C-30), 126.0 (C-10), 121.8 (C-60), 115.3 (C-50), 114.2
(C-80) and 114.0 (C-20)] and trans-p-coumaroyl group signals19
[dH 7.65 (d, J = 16.0 Hz, H-700), 7.46 (d, J = 8.5 Hz, H-200, 600), 6.80
(d, J = 8.5 Hz, H-300, 500), and 6.33 (d, J = 16.0 Hz, H-800); dC 167.4
(C-900), 160.1 (C-400), 145.8 (C-700), 130.0 (C-200, 600), 126.6 (C-100),
and 115.6 (C-300, 500)]. The position of the trans-p-coumaroyl group
was confirmed by the HMBC experiment, in which a correlation
was observed between the H-5 (dH 5.38) of the quinic acid moiety
and the C-900 (dC 167.4) of the trans-p-coumaroyl unit as shown in
20
30
40
50
60
70
80
43.4
170.4
55.5
90
70-OMe
100
126.6
130.0
115.6
160.1
115.6
130.0
145.8
114.1
167.4
131.9
113.4
145.4
145.1
115.0
118.4
79.9
43.3
171.0
55.5
126.7
113.9
145.6
148.4
115.3
121.8
146.1
114.1
167.3
126.0
132.3
114.4
158.7
114.4
132.3
143.8
115.2
166.2
126.1
132.3
114.3
158.6
114.3
132.3
143.8
115.3
165.7
200
300
400
500
600
700
800
900
700-OMe
of 6 which was isolated from Arnica chamissonis.20 The major dif-
ference was the acyl moiety at C-5, indicating the presence of the
trans-p-coumaroyl unit in 1 instead of the trans-caffeoyl unit in
6. Alkaline methanolysis (3% KOH in MeOH) of
1 afforded
trans-p-coumaric acid methyl ester,21 trans-caffeic acid methyl es-
ter,22 and quinic acid,17,18 which were confirmed by comparison of
their 1H NMR and optical rotation values with literature values,
respectively. Thus, the structure of 1 was determined to be 1-O-
trans-caffeoyl-5-O-trans-p-coumaroylquinic acid.
Figure 2. The downfield shift of the signals at H-5 (
DdH +1.2) and at
C-5 ( dC +2.5), compared to the corresponding signals of quinic
D
acid18 which was isolated from Centella asiatica, suggested that
the trans-p-coumaroyl unit should be placed at C-5 of 1.18 The
trans-caffeoyl unit was placed at C-1, since the carbon chemical
shift of the oxygenated quaternary carbon (C-1) was downfield
shifted to dC 79.8, compared to that of quinic acid containing a free
hydroxyl group at C-1 (dC 74.5 in 7; 73.5 in 8; 74.8 in 9).18 The
above 1H and 13C NMR spectral data of 1 were very similar to those
Compound (2) was isolated as a colorless gum. The molecular
formula C26H28NaO13 was inferred from the positive ion HRESIMS
m/z 571.1431 [M+Na]+ (calcd for 571.1428). The 1H (Table 1) and
13C (Table 2) NMR data were similar to those of 1, except for the
presence of 7,8-dihydro-7-methoxycaffeoyl signals23 [dH 6.74
(1H, d, J = 2.0 Hz, H-200), 6.73 (1H, d, J = 8.5 Hz, H-500), 6.66 (1H,
dd, J = 8.5, 2.0 Hz, H-600), 4.48 (1H, m, H-700), 3.16 (3H, s, 700-OMe)
Table 1
1H NMR data of compounds 1–5 (in CD3OD, 500 MHz, d in ppm, J in Hz)
Position
1
2
3
4
5
2
3
2.43 m, 2.44 m
4.29 m
2.35 m, 2.48 m
4.23 m
2.38 m, 2.40 m
4.26 m
2.33 m, 2.45 m
4.23 m
2.35 m, 2.48 m
4.23 m
4
3.78 m
3.68 m
3.75 m
3.70 m
3.69 m
5
5.38 m
5.29 m
5.33 m
5.37 m
5.31 m
6
2.07 m, 2.57 m
7.05 d (2.0)
1.98 m, 2.48 m
7.05 d (2.0)
2.04 m, 2.50 m
6.74 d (2.0)
1.98 m, 2.54 m
7.46 d (8.5)
6.80 d (8.5)
6.80 d (8.5)
7.46 d (8.5)
7.65 d (16.0)
6.36 d (16.0)
1.98 m, 2.52 m
7.65 d (8.5)
6.74 d (8.5)
6.74 d (8.5)
7.65 d (8.5)
6.86 d (13.0)
5.82 d (13.0)
20
30
50
6.78 d(8.5)
6.76 d (8.0)
6.73 d (8.5)
6.66 dd (8.5, 2.0)
4.51 m
2.57 m, 2.74 m
3.18 s
60
6.96 dd (8.5, 2.0)
7.57 d (16.0)
6.29 d (16.0)
6.95 dd (8.0, 2.0)
7.57 d (16.0)
6.28 d (16.0)
70
80
70-OMe
200
7.46 d (8.5)
6.80 d (8.5)
6.80 d (8.5)
7.46 d (8.5)
7.65 d (16.0)
6.33 d (16.0)
6.74 d (2.0)
7.05 d (2.0)
7.65 d (8.5)
6.75 d (8.5)
6.75 d (8.5)
7.65 d (8.5)
6.86 d (13.0)
5.79 d (13.0)
7.64 d (8.5)
6.73 d (8.5)
6.73 d (8.5)
7.64 d (8.5)
6.86 d (13.0)
5.78 d (13.0)
300
500
6.73 d (8.5)
6.66 dd (8.5, 2.0)
4.48 m
2.59 m, 2.74 m
3.16 s
6.76 d (8.5)
600
6.96 dd (8.5, 2.0)
7.57 d (16.0)
6.25 d (16.0)
700
800
700-OMe

2142
S. Y. Lee et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2140–2144
OH
OH
OH
OH
OH
OH
7
O
OH
O
OH
O
O
OH
O
O
1
6
9'
2
5 O
O
O
O
OCH3
O
4
O
O
9''
3
HO
HO
O
HO
O
OH
OH
OH
OCH3
OH
OH
OH
OH
OH
1
2
3
HO
OH
O
OH
O
OH
O
O
O
O
OH
O
O
OH
HO
O
HO
O
OH
OH
4
5
Figure 2. Key HMBC (?) correlations of 1–5.
and 2.74, 2.59 (2H, m, H-800); dC 171.0 (C-900), 145.4 (C-300), 145.1
(C-400), 131.9 (C-100), 118.4 (C-600), 115.0 (C-500), 113.4 (C-200), 79.9
(C-700), 55.5 (700-OMe) and 43.3 (C-800)] in 2 instead of the trans-p-
coumaroyl group in 1. The position of the 7,8-dihydro-7-meth-
oxycaffeoyl group was confirmed by the HMBC experiment, in
which a correlation was observed between the H-5 (dH 5.29) of
the quinic acid moiety and the C-900 (dC 171.0) of the 7,8-dihy-
dro-7-methoxycaffeoyl unit. In addition, this HMBC correlation be-
tween the methoxy signals at dH 3.16 and C-700 (dC 79.9) implied
that the methoxy group was located at C-700 (Fig. 2). Alkaline hydro-
lysis of 2 afforded trans-caffeic acid methyl ester,22 quinic acid17,18
and 7,8-dihydro-7-methoxycaffeic acid methyl ester (2a). The 1H,
13C NMR, optical rotation value and HRFABMS data of 2a were sim-
ilar to those of pisnoninol I isolated from Pisonia unbellifera,24 ex-
cept for the presence of an additional methoxy signal (dH 3.15; dC
55.9). 2a had the same planar structure as taraxafolin isolated from
Taraxacum formosanum,23 except for the optical rotation value. The
optical rotation of 7,8-dihydro-7-methoxycaffeic acid methyl ester
acid methyl ester.23,25 Therefore, the structure of 3 was determined
to be 1-O-7,8-dihydro-7a-methoxycaffeoyl-5-O-trans-caffeoylqui-
nic acid.
Compound (4) was obtained as a colorless gum, and its molec-
ular formula 25H25O10 was inferred from the positive ion
C
HRFABMS m/z 485.1446 [M+H]+ (calcd for 485.1448). The 1H and
13C NMR data were very similar to those of 15, except for the pres-
ence of additional trans-p-coumaroyl group signals19 [dH 7.65 (1H,
d, J = 16.0 Hz, H-70), 7.46 (2H, d, J = 8.5 Hz, H-20,60), 6.80 (2H, d,
J = 8.5 Hz, H-30,50) and 6.36 (1H, d, J = 16.0 Hz, H-80); dC 166.6 (C-
90), 159.8 (C-40), 145.6 (C-70), 129.8 (C-20,60), 125.8 (C-10), 115.3
(C-30,50) and 114.0 (C-80)]. The location of another trans-p-couma-
royl group should be placed at C-1 (dC 79.8) in 4 due to the
downfield shift of the signal at C-1 (DdC +3.0), compared to the cor-
responding signal of 15. Alkaline methanolysis (3% KOH in MeOH)
of 4 yielded trans-p-coumaric acid methyl ester, cis-p-coumaric
acid methyl ester21 and quinic acid.17,18 Therefore, the structure
of 4 was determined to be 1-O-trans-p-coumaroyl-5-O-cis-p-cou-
maroylquinic acid.
([a] +30.5) (Ref. 24 [a] +37.8) was almost of the same value but of
opposite sign to that of taraxafolin C ([
that 7,8-dihydro-7-methoxycaffeic acid methyl ester could be a
a
] ꢀ32.0), which suggested
Compound (5) was obtained as a colorless gum, and its molec-
ular formula C25H25O10 was inferred from the positive ion
stereoisomer of taraxafolin. The above data indicated that the
HRFABMS m/z 485.1450 [M+H]+ (calcd for 485.1448). The 1H and
13C NMR data were very similar to those of 4 except for the pres-
ence of cis-p-coumaroyl group signals26 [dH 7.65 (2H, d, J = 8.5 Hz,
H-20,60), 6.86 (1H, d, J = 13.0 Hz, H-70), 6.74 (2H, d, J = 8.5 Hz, H-
30,50) and 5.82 (1H, d, J = 13.0 Hz, H-80); dC 166.1 (C-90), 158.7
(C-40), 143.9 (C-70), 132.3 (C-20,60), 126.2 (C-10), 115.4 (C-80) and
114.4 (C-30,50)] in 5, instead of the trans-p-coumaroyl group in 4.
The coupling constant values (13.0 Hz) at H-70 (dH 6.86) and H-80
(dH 5.82) in the p-coumaroyl unit proton signals indicated the cis
form.26 Alkaline hydrolysis (3% KOH in MeOH) of compound 5 gave
quinic acid,17,18 and cis-p-coumaric acid methyl ester.21 Thus, the
structure of 5 was determined to be 1,5-di-O-cis-p-coumaroylqui-
nic acid.
methoxyl group at C-7 has a
-orientation.25 Therefore, the structure
of 2 was determined to be 1-O-trans-caffeoyl-5-O-7,8-dihydro-7a-
methoxycaffeoylquinic acid.
Compound (3) was obtained as a colorless gum, and its molec-
ular formula C26H28NaO13 was inferred from the positive ion HRE-
SIMS m/z 571.1432 [M+Na]+ (calcd for 571.1428). The 1H (Table 1)
and 13C (Table 2) NMR data were very similar to those of 2. The dif-
ference was the position of two acyl groups. The location of the
trans-caffeoyl unit was determined by HMBC correlation between
H-5 (dC 5.33) of the quinic acid moiety and C-900 (dC 167.3) of the
trans-caffeoyl unit. In addition, a comparison of the carbon signal
(dC 79.8, C-1) of 1 with the corresponding signal (dC 79.7) of 3 re-
vealed that another acyl group, 7,8-dihydro-7
unit, should be placed at C-1 in 3. Alkaline hydrolysis of 3 with
a
-methoxycaffeoyl
The structures of the known compounds (6–15) were identified
as 1,5-O-trans-dicaffeoylquinic acid (6),20 3,4-O-trans-caffeoylqui-
nic acid methyl ester (7),27 3,5-O-trans-dicaffeoylquinic acid
methyl ester (8),15 4,5-O-trans-dicaffeoylquinic acid methyl ester
3% KOH in MeOH at room temperature afforded trans-caffeic acid
methyl ester,22 quinic acid17,18 and 7,8-dihydro-7
a-methoxycaffeic

S. Y. Lee et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2140–2144
2143
Table 3
those factors, NO is a primary released factor in activated microglia.
The IC50 values of 2, 3, 8 and 11 were 4.66, 12.52, 9.04, and
Inhibitory effect on NO production of compounds 1–15 in LPS-activated BV-2 cells
a
Sample
IC50
(lM)
Cell viabilityb (%)
12.11
trol), showed a low IC50 value of 16.32
2, 3, 8 and 11 were more active than the
l
M, respectively.
L
-NMMA, a NOS inhibitor (a positive con-
M (Table 3). Compounds
-NMMA. In particular,
l
1
2
3
4
5
6
7
8
9
10
11
12
13
14
23.53
4.66
94.3 1.9
93.2 1.0
90.9 2.3
98.1 2.1
96.0 3.7
93.7 3.8
96.8 2.9
91.1 6.8
92.2 3.8
92.6 1.2
95.3 3.0
90.6 0.9
108.1 7.5
108.8 0.9
108.7 5.5
103.2 4.3
L
12.52
59.82
52.79
22.63
20.15
9.04
24.58
16.57
12.11
17.04
44.99
81.97
86.41
16.32
compounds 2 and 8 exhibited very strong anti-neuroinflammatory
properties. Moreover, both compounds 2 and 8 did not affect cell
viability at a concentration of 20 lM. To confirm effects of these
compounds on NO production in different cell line, NO levels in
the medium of murine macrophage RAW264.7 activated by LPS
were also measured. The IC50 values of 2 and 8 were 174.89 and
42.87
lM. And both compounds had no influence on cell viability
at a concentration of 20
l
M in RAW264.7 cells (data not shown).
These data confirmed that the inhibitory effects of compounds 2
and 8 on NO production were more potent in BV2 than RAW264.7.
In microglia, NO production is primarily regulated by the induc-
ible nitric oxide synthase (iNOS).32 Based on western blot analysis
and iNOS activity assay, we determined whether the NO inhibitory
effects of compounds 2 and 8 are related to the regulation of the
protein expression and/or enzymatic activity of iNOS. As shown in
15
NMMAc
a
IC50 value of each compound was defined as the concentration (
50% inhibition of NO production in LPS-activated BV-2 cells.
lM) that caused
b
Cell viability after treatment with 20 lM of each extract was expressed as a
percentage (%) of the LPS only treatment group. The results are averages of three
Figure 3A, pretreatment with 10 lM of compound 2 led to a de-
crease in iNOS protein level. However, compound 8 did not showed
independent experiments, and the data are expressed as mean SD.
c
NMMA as a positive control.
an inhibitory effect on LPS-induced iNOS expression in concentra-
tion range of 1–10
and 8 reduced iNOS activity in LPS-pretreated BV-2. One and
10 M of compound 2, and 1 and 5 M of compound 8 inhibited
iNOS activity, significantly. To investigate the precise mechanisms
of compounds 2 and 8 on NO regulation, NO radical scavenging as-
say was also performed. As shown inFigure 3C, 5 and 10 lM of com-
pound 2 reduced the accumulation of nitrite upon decomposition of
NO donor, sodium nitroprusside (SNP). Moreover, the effect of com-
lM. As shown in Figure 3B, both compounds 2
(9),27 4-O-trans-caffeoylquinic acid methyl ester (10),28 5-O-trans-
caffeoylquinic acid methyl ester (11),28 5-O-cis-caffeoylquinic acid
methyl ester (12),29 4-O-trans-coumaroylquinic acid methyl ester
(13),30 5-O-trans-coumaroylquinic acid methyl ester (14),31 and
5-O-cis-coumaroylquinic acid methyl ester (15)26 by comparison
of their spectroscopic data with those reported in the literature.
Lipopolysaccharide (LPS) is an important activator of microglia.
Overactivated microglia cells secrete excessive inflammatory and
cytotoxic mediators which are deleterious to neuronal cells. Among
l
l
pound 8 is similar to that of compound 2 in 1–10
lM. These results
indicate that quinic acid derivatives isolated from P. brachycarpa
Figure 3. Effects of compounds 2 and 8 on NO regulation. (A) Effects of compounds 2 and 8 on LPS-induced iNOS expression in BV-2 cells. (B) Effects of compounds 2 and 8 on
iNOS activity in BV-2 cells. (C) Effects of compounds 2 and 8 on NO scavenging. Sodium nitroprusside (SNP) is a NO generator. #p <0.05 indicates statistically significant
difference between the control and LPS alone-treated groups. ⁄p <0.05 indicate statistically significant differences compared to treatment with LPS alone.

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S. Y. Lee et al. / Bioorg. Med. Chem. Lett. 23 (2013) 2140–2144
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exert attenuate LPS-induced pro-inflammatory responses in
microglial BV2 cells through down regulation of NO. NO regulation
of compounds 2 and 8 might be attributed to the inhibition of iNOS
expression, enzymatic activity, and NO scavenging property in all.
In Korea, P. brachycarpa, called as cham-na-mul, is well known as
a vegetable and a popular edible herb. In the course of our contin-
uing search for biologically active constituents from Korean medic-
inal plants, we investigated the MeOH extract from the aerial parts
of P. brachycarpa and isolated five new quinic acid derivatives (1–5),
together with 10 known quinic acid derivatives (6–15). The chem-
ical structures of the new compounds (1–5) were determined by
the mean of 1H, 13C NMR, COSY, HMQC, HMBC and HRMS. Com-
pounds 2 and 8 isolated from P. brachycarpa may have neuroprotec-
tive properties by inhibiting inflammatory activation of microglia.
This study supports that P. brachycarpa, a well known vegetable,
may be an invaluable functional food source for the regulation of
neurodegenerative diseases related with neuroinflammation.
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Acknowledgments
This research was supported by the Basic Science Research
Program through the National Research Foundation of Korea
(NRF), funded by the Ministry of Education, Science andTechnology
(2011-0028285). We thank Drs. E. J. Bang, S. G. Kim, and J. J. Seo at
the Korea Basic Science Institute for their assistance with the NMR
spectroscopic and mass spectrometric measurements.
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