Synthesis and Anti-Hypercholesterolemic Evaluations of Calcium Salts of 4-Substituted Quinoline-Based Mevalonic Acid

Article abstract of DOI:10.1111/j.1747-0285.2011.01198.x

A series of calcium (3R,5S,6E)-7-[4,6,7,8-substituted-quinoline-3-yl]-3,5-dihydroxy-hept-6-enoates was synthesized from the lactones, and the anti-hypercholesterolemic evaluation in quails was presented in this report. It was found that most of the compou

Full text of DOI:10.1111/j.1747-0285.2011.01198.x

ª 2011 John Wiley & Sons A/S
doi: 10.1111/j.1747-0285.2011.01198.x
Chem Biol Drug Des 2011; 78: 730–733
Research Letter
Synthesis and Anti-Hypercholesterolemic
Evaluations of Calcium Salts of 4-Substituted
Quinoline-Based Mevalonic Acid
alent to that of pitavastatin, atorvastatin and fluvastatin. Among
them, compounds 1a, 1b, 1c, 1d, 1e, and 1h showed more potent
than rosuvastatin. Generally, it was known (6) that the ring-opened
dihydroxyheptenoate form was more active than the corresponding
lactone form, especially in vivo, so that most of the statins in the
market are calcium salts, such as pitavastatin calcium, atorvastatin
calcium, and rosuvastatin calcium. Herein, in this paper, we like to
report the preparation of the calcium salts from these ten com-
pounds and anti-hypercholesterolemic studies in quails.
Qun Hao, Zhengyan Cai, Jing Pan, Yongjia
Li, Yuye Xia, Yang Min, Yuchen Sheng and
Weicheng Zhou*
State Key Lab of New Drug & Pharmaceutical Process, Shanghai
Key Lab of Anti-Infectives, Shanghai Institute of Pharmaceutical
Industry, State Institute of Pharmaceutical Industry, Shanghai
200437, China
*Corresponding author: Weicheng Zhou, profzhouwc@yahoo.com.cn
A
series of calcium (3R,5S,6E)-7-[4,6,7,8-substi-
tuted-quinoline-3-yl]-3,5-dihydroxy-hept-6-enoates
was synthesized from the lactones, and the anti-
hypercholesterolemic evaluation in quails was pre-
sented in this report. It was found that most of
the compounds significantly decreased levels of
total cholesterol and low-density lipoprotein. 2e
showed better hypolipidemic effect than atorvast-
atin, and 2d and 2j exhibited comparable efficacy
to atorvastatin. These three compounds were
selected as the hypocholesterolemic candidates
for further evaluation.
Materials and Methods
Analysis
Melting points were determined on an Electrothermal Melting
Point Apparatus and were uncorrected. 1H NMR spectra were
recorded at 400 MHz on a Varian Inova-400 spectrometer in
DMSO-d6 with chemical shifts (d) given in ppm relative to tetra-
methylsilane (TMS) as an internal standard. Mass spectra were
recorded by an Q-TOF Mass Spectrometer using electrospray ioni-
zation (ESI). HPLC was performed in Agilent1100 with Ultimate
XB-C18 column (Welch Materials, Inc., Ellicott City, MD, USA;
250 · 4.6 mm, 5 lm) as the stationary phase and tetrahydrofuran
(THF) ⁄ H2O = 2:8 (pH 4.5 adjusted by NH4OAc and HOAc) as the
mobile phase, detected at 270 nm.
Key words: anti-hypercholesterolemic evaluation, calcium salts,
3-hydroxy-3-methyl glutaryl-coenzyme A, mevalonic acid, quinoline
Received 25 January 2010, revised 22 June 2011 and accepted for pub-
lication 1 July 2011
Synthesis
Hypercholesterolemia is a well-known risk factor for atherosclerotic
diseases, especially for coronary heart disease (1). Currently, the
most common method of treating hypercholesteromia is the use of
3-hydroxy-3-methyl glutaryl-coenzyme A (HMG CoA) reductase inhib-
itors, also known as statins, such as lovastatin, simvastatin, pra-
vastatin, fluvastatin, atorvastatin, rosuvastatin, and pitavastatin.
Their mechanism of action is based on the reversible and competi-
tive inhibition of HMG CoA reductase, which is the rate-limiting
enzyme of cholesterol biosynthesis in the liver and non-hepatic tis-
sues, ultimately resulting in up-regulation of the low-density lipo-
protein (LDL) receptor and the clearing of LDL particles from the
blood stream (2).
General methods
Scheme 1 illustrated the synthesis of calcium (3R,5S,6E)-7-[4,6,7,8-
substituted-quinoline-3-yl]-3,5-dihydroxy-hept-6-enoates 2. The prep-
aration started from the reported lactones 1 (3,5). At first, we tried
to convert lactone 1 in THF to the corresponding carboxylate
sodium salt at 0 ꢀC by saponification with NaOH aqueous solution,
then HCl solution was added to adjust pH 7–8, the solution was
concentrated under reduced pressure to evaporate THF at 40 ꢀC,
and CaCl2 aq solution was added to generate calcium salts 2. In
the process, especially during the evaporation of THF, it was found
that the product was contaminated with the Z-isomer (3). Some-
times this main impurity contributed 3–5% in the product deter-
mined by HPLC. Therefore, another method was tried. After the
hydrolysis of compound 1 with aqueous NaOH, the pH of the solu-
tion was adjusted to 2 to give its carboxylic acid 4, which was
crystallized with ethanol to control the quality. Then, NaOH aq solu-
As parts of our program of research on novel hypocholesterolemic
agents, a series of 4-phenoxy or phenylthio-quinoline mevalonolac-
tones 1 was synthesized in our laboratory (3–5). In the in vitro test
for the evaluation of inhibition on the rat HMG CoA reductase, ten
compounds 1a–j (Figure 1) showed the activity better than or equiv-
730

Synthesis and Anti-Hypercholesterolemic Evaluations of Calcium Salts
R
1a: X=S, R=4-CH(CH3)2, R1=F, R2=F, R3=F
1b: X=S, R=3-OCH3, R1=F, R2=SC6H4-3-OCH3, R3=H
1c: X=S, R=4-CH(CH3)2, R1=SC6H4-4-CH(CH3)2, R2=SC6H4-4-CH(CH3)2, R3=SC6H4-4-CH(CH3)2
1d: X=S, R=H, R1=F, R2=SC6H5, R3=H
OH
O
X
N
1e: X=S, R=4-F, R1=F, R2=F, R3=F
R1
R2
1f: X=S, R=3-OCH3, R1=F, R2=Cl, R3=H
O
1g: X=S, R=4-CH(CH3)2, R1=F, R2=SC6H4-4-CH(CH3)2, R3=H
1h: X=O, R=4F, R1=F, R2=Cl, R3=H
X=S or O
1i: X=O, R=H, R1=OC6H5, R2=OC6H5, R3=OC6H5
1j: X=O, R=H, R1=F, R2=OC6H5, R3=F
R3
1
Figure 1: Structure of 10 active 3-hydroxy-3-methyl glutaryl-coenzyme A reductase inhibitors.
R
R
R
OH
O
X
N
X
N
OH OH
X
N
a
R1
R2
R1
R2
COOCa1/2
R1
R2
O
COOCa1/2
OH OH
R3
R3
R3
3
2
1
b
c
R
X
N
OH OH
R1
R2
COOH
R3
4
Scheme 1: Synthesis of calcium (3R,5S,6E)-7-[4,6,7,8-substituted-quinoline-3-yl]-3,5-dihydroxy-hept-6-enoates 2. Reagents and conditions:
(a) i. NaOH aq, EtOH:THF = 1:1, 0 ꢀC, ii. CaCl2 aq, H2O; (b) NaOH aq, EtOH:THF = 1:1, 0 ꢀC, HCl aq; (c) NaOH aq, CaCl2 aq.
tion was added to the suspension of compound 4 in 66% ethanol
to get the clear solution of its sodium salt at 0 ꢀC, and CaCl2 solu-
tion was added to generate the calcium salt 2. Through this
approach, the final product had the purity of more than 99% by
HPLC.
General procedure for preparation of
compounds 2a–2j
One molar aqueous sodium hydroxide solution (4.50 mL) was added
dropwise to a suspension of compound 4e (2.00 g, 4.30 mmol) in
ethanol (20 mL) and water (10 mL). The mixture was stirred at 0 ꢀC
to get a clear solution. Then, 0.47 g of calcium chloride (4.30 mmol)
in 1 mL of water was added. After stirring for 1 h, the precipitate
was filtrated, washed with water and ethanol, and dried at 50 ꢀC
to afford the title compound 2e (1.60 g, 76.9%), mp: 174 ꢀC (dec),
General procedure for preparation of
compounds 4a–4j
26
One molar aqueous sodium hydroxide solution (16.7 mL) was added
dropwise to a solution of compound 1e (5.00 g, 11.1 mmol) in eth-
anol (50 mL). After the mixture was stirred for 1 h at 0 ꢀC, 1 M HCl
(25 mL) was added to the solution and then ethyl acetate was
added. The organic layer was separated, washed with brine, dried
over Na2SO4, and concentrated to afford a yellow solid. Recrystalli-
zation from 50 mL of ethanol provided 4.1 g of compound 4e
½aꢀD = +10.8 (c = 1, THF).
All other carboxylic acid 4 were converted to the corresponding cal-
cium salts 2 by the similar procedure, and the physical and spectra
data of 2a–j were shown in Tables 1 and 2.
26
1
(80.0%) as crystals. mp: 168–170 ꢀC, ½aꢀD = +33.0 (c = 1, THF), H
NMR d ppm in DMSO-d6: 11.09 (br.s, 1H), 9.30 (s, 1H), 8.12–8.07
(m, 1H), 7.28 (d, 1H, J = 15.6 Hz), 7.22–7.08 (m, 4H), 6.76 (dd, 1H,
J = 5.2, 15.6 Hz), 4.38 (m, 1H), 4.01–4.00 (m, 1H), 2.31–2.25 (m,
2H), 1.70–1.57 (m, 2H).
Biology
Pharmacological evaluation in hyperlipemia
quails
According to the literature (7), 230 male quails were housed in a
room kept at 23 € 2 ꢀC, 50–70% relative humidity, and illuminated
from 7:00 to 19:00 h. They were fed with standard chow for
7 days, then divided into 23 groups randomly including normal,
All other lactones 1 were converted to the corresponding carboxylic
acid 4 by the similar procedure.
Chem Biol Drug Des 2011; 78: 730–733
731

Hao et al.
Table 1: Physical data of the target compounds 2a–ja
R
X
N
OH OH
2+
Ca
R1
COO
R2
2
R3
20
c
No.
Formula
Mp (ꢀC)b
½aꢀD
MS ESI+(M + 1)
2a
2b
2c
2d
2e
2f
C50H46F6N2O8S2Ca
C60H54F2N2O12S4Ca
C52H56NO4S4Nae
C56H46F2N2O8S4Ca
C44H32F8N2O8S2Ca
C46H40F2N2Cl2O10S2Ca
C68H70F2N2O8S4Ca
C44H34Cl2F4N2O10Ca
C80H64N2O16Ca
118
152
118
136
174
174
206
190
140
146
+15.3
+3.0
1021.29
1201.33
888.34e
1081.26
973.10
993.15
1249.46
937.28
+18.2
)13.3
+10.8
+16.8
+7.0
2g
2h
2i
+20.2d
+13.9
+17.7d
1349.43
1053.44
2j
C56H44F4N2O12Ca
aThe definition of X, R, R1, R2, and R3 is the same as Figure 1.
bAll compounds melted with decomposition.
cThe optical values were measured in THF, c = 1.
dThe optical values were measured in THF ⁄ H2O = 2:1, c = 1.
eThe compound 2c was prepared as sodium salt, but the MS gave the molecular weight of the acid.
ESI, electrospray ionization.
Table 2: 1H NMR data of the target compounds 2a–j
No.
1H NMR (d; ppm) 400 MHz in DMSO-d6
2a
9.31 (s, 1H), 8.12–8.07 (m, 1H), 7.34–7.29 (m,1H), 7.16–7.05 (m, 4H), 6.77 (dd, 1H, J = 5.2, 16.4 Hz), 4.41–4.37 (m, 1H), 4.06–4.02
(m, 1H), 2.43–2.26 (m, 2H), 1.68–1.63 (m, 2H)
2b
2c
2d
2e
2f
9.19 (m, 1H), 7.99 (d, J = 11.5 Hz), 7.61 (d, 1H, J = 7.2 Hz), 7.46–7.41 (m,1H), 7.28–7.06 (m, 5H), 6.78–6.67 (m,3H), 6.58–6.56
(m, 1H), 4.37–4.36 (m, 1H), 4.02 (m, 1H), 3.67 (s, 3H), 3.78 (s, 3H), 2.42–2.25 (m, 2H), 1.67–1.60 (m, 2H)
9.12 (s, 1H), 7.83 (s, 1H), 7.31–7.23 (m, 5H), 7.19–6.98 (m, 8H), 6.87–6.85 (m, 2H), 6.67–6.60 (m, 3H), 4.35–4.30 (m, 1H), 3.92–3.90
(m, 1H), 2.98–2.91 (m, 1H), 2.83–2.69 (m, 3H), 2.07–1.82 (m, 2H), 1.56–1.44 (m, 2H), 1.26–1.11 (m, 24H)
9.17 (s, 1H), 7.99 (d, 1H, J = 11.5 Hz), 7.59–7.51 (m, 5H), 7.28–7.08 (m, 6H), 6.71 (dd, 1H, J = 6.8, 15.0 Hz), 4.38–4.34
(m, 1H), 4.02–3.95 (m, 1H), 2.37–2.20 (m, 2H), 1.66–1.56 (m, 2H)
9.24 (s, 1H), 8.06–8.02 (m, 1H), 7.26–7.07 (m, 5H), 6.73 (dd, 1H, J = 5.2, 16.4 Hz), 4.41–4.36 (m, 1H), 3.95–3.91 (m, 1H), 2.22–2.03
(m, 2H), 1.65–1.52 (m, 2H)
9.23 (s, 1H), 8.03 (d, 1H, J = 10 Hz), 8.21 (d, 1H, J = 7.6 Hz), 7.22–6.51 (m, 5H),7.24 (d, 1H, J = 16 Hz), 4.40–4.38 (m, 1H),
4.03–3.95 (m, 1H), 3.64 (s, 3H), 2.24–2.05 (m, 2H), 1.64–1.53 (m, 2H)
2g
9.07 (s,1H), 7.93 (d, 1H, J = 11.2 Hz), 7.49–7.47 (m, 2H), 7.42–7.36 (m, 3H), 7.09–6.96(m, 4H), 6.67 (dd, 1H, J = 4.8, 16 Hz), 4.37–4.36
(m, 1H), 3.96–3.95 (m, 1H), 2.96–2.92 (m, 1H), 2.77–2.74 (m, 1H), 2.24–2.06 (m, 2H), 1.66–1.55 (m, 2H), 1.22 (d, 6H, J = 6.8 Hz),
1.09 (d, 6H, J = 6.8 Hz)
2h
9.25 (s, 1H), 8.30 (d, 1H, J = 6.8 Hz), 7.63 (d, 1H, J = 10 Hz), 7.17–7.13 (m, 2H), 6.93–6.75 (m, 2H), 6.72–6.65 (m, 2H), 4.31–4.27
(m, 1H), 3.97–3.91 (m, 1H), 2.36–2.19 (m, 2H), 1.63–1.52 (m, 2H)
2i
2j
9.09 (s, 1H), 7.31–7.21 (m, 7H), 7.13–6.62 (m, 14H), 4.28–4.24 (m, 1H), 3.97–3.91 (m,1H), 2.37–2.20 (m, 2H), 1.64–1.49 (m, 2H)
9.29 (s, 1H), 7.51 (dd, 1H, J = 1.6, 11.2 Hz), 7.40–7.33 (m, 4H), 7.16–7.05 (m, 4H), 7.05–6.93 (m, 2H), 6.75–6.72 (m, 2H), 4.29
(d, 1H, J = 4.8 Hz), 3.97–3.93 (m, 1H), 2.50–2.20 (m, 2H), 2.50–2.20 (m, 2H), 1.62–1.55 (m, 2H)
negative control, atorvastatin (Atr, 20 mg ⁄ kg), 2a (5, 20 mg ⁄ kg),
2b (5, 20 mg ⁄ kg), 2c (5, 20 mg ⁄ kg), 2d (5, 20 mg ⁄ kg), 2e (5,
20 mg ⁄ kg), 2f (5, 20 mg ⁄ kg), 2j (5, 20 mg ⁄ kg), 2h (5, 20 mg ⁄ kg),
2i (5, 20 mg ⁄ kg), and 2j (5, 20 mg ⁄ kg). The normal group was
fed with standard chow, and the negative control was fed with
hypercholesterol chow including 10% lard and 0.5% cholesterol.
The other groups were fed with hypercholesterol chow (10% lard,
0.5% cholesterol) and corresponding drugs suspended in 0.5%
sodium carboxymethylcellulose (CMC-Na) once daily for 28 days.
After the final administration, they were fasted overnight, and
blood was withdrawn from jugular vein. The plasma was sepa-
rated and measured for triglyceride (TG), total cholesterol (TC),
LDL, and high-density lipoprotein level (HDL) using Hitachi auto-
matic biochem analyzer-7080. The ratio of HDL ⁄ LDL was intro-
duced to evaluate the ability of the tested compound for selective
reduction in LDL. The results are shown in Table 3.
732
Chem Biol Drug Des 2011; 78: 730–733

Synthesis and Anti-Hypercholesterolemic Evaluations of Calcium Salts
Table 3: Hypolipidemic effect of compounds 2a–j in Quails
No.
Dosage (mg ⁄ kg)
TC (mM)
TG (mM)
HDL-C (mM)
LDL-C (mM)
H ⁄ L
Normal
Negative
Atorvastatin
2a
–
4.67 € 1.13**
12.42 € 4.05
6.51 € 1.61**
8.65 € 2.48*
8.66 € 0.99*
9.66 € 3.69
10.55 € 5.65
8.72 € 3.57
8.05 € 0.78**
11.16 € 3.26
7.54 € 1.96**
9.11 € 3.89
5.57 € 2.41**
11.63 € 4.20
8.53 € 1.81**
14.13 € 4.29
8.88 € 3.27
11.64 € 3.67
7.61 € 1.62**
9.75 € 5.53
0.81 € 0.37
0.57 € 0.14
0.76 € 0.39
0.70 € 0.21
4.47 € 7.06
1.95 € 3.99
0.58 € 0.28
0.59 € 0.13
0.51 € 0.25
0.53 € 0.23
1.46 € 1.48
1.50 € 1.02
0.45 € 0.27
0.56 € 0.16
0.42 € 0.18
0.56 € 0.17
0.35 € 0.07
0.74 € 0.35
0.96 € 0.78
1.24 € 1.11
1.26 € 0.60
0.54 € 0.19
0.97 € 0.94
2.97 € 0.59**
4.73 € 1.03
4.39 € 0.86
4.89 € 1.16
4.74 € 1.57
5.35 € 1.97
5.35 € 1.56
5.84 € 1.69
5.31 € 0.68
6.61 € 0.80**
4.93 € 0.99
4.75 € 0.78
3.73 € 1.53
6.32 € 1.38
5.06 € 0.91
6.93 € 1.28**
5.46 € 1.57
6.63 € 1.32**
4.66 € 1.20
4.98 € 1.58
4.98 € 0.95
6.21 € 1.85
3.77 € 1.03
1.69 € 1.05**
8.25 € 3.53
2.51 € 1.14**
4.61 € 2.06*
3.66 € 1.45**
5.07 € 3.02
2.18 € 0.89**
0.71 € 0.42
2.19 € 1.46*
1.21 € 0.45*
1.37 € 0.25**
1.29 € 0.68
0.5% CMC-Na
20
5
20
5
20
5
20
5
20
5
20
5
20
5
20
5
20
5
20
5
20
2b
2c
2d
2e
2f
6.40 € 4.96
1.06 € 0.37
3.99 € 2.95*
3.63 € 0.81**
6.33 € 3.04
2.73 € 1.62**
3.46 € 2.09**
1.94 € 1.30**
6.51 € 3.53*
4.30 € 1.65*
9.32 € 4.02
3.95 € 2.42*
6.40 € 3.27
3.62 € 1.53**
5.16 € 4.61
1.91 € 0.76*
1.54 € 0.42**
1.23 € 0.46*
2.19 € 0.90**
1.58 € 0.42**
2.21 € 0.65**
1.24 € 0.60
1.33 € 0.53*
0.92 € 0.45
2g
2h
2i
1.79 € 0.72**
1.15 € 0.31*
1.44 € 0.59*
1.56 € 0.79*
1.54 € 0.63**
1.43 € 0.77*
2.11 € 0.92**
9.18 € 3.72
12.4 € 7.70
6.25 € 2.52**
4.08 € 2.75*
6.75 € 5.97
2.52 € 2.16**
2j
*p < 0.05; **p < 0.01 significantly different from the negative control.
TG, triglyceride; TC, total cholesterol; LDL, low-density lipoprotein; HDL high-density lipoprotein level.
Results
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Conclusion
In conclusion, we have described the synthesis and anti-hypercho-
lesterolemic evaluation of a series of calcium salts of 4-substituted
quinoline-based mevalonic acid. In anti-hypercholesterolemic studies
in quails, it was found that most of the compounds significantly
decrease levels of TC and LDL. 2e showed better hypolipidemic
effect than atorvastatin, and 2d and 2j exhibited comparable effi-
cacy to atorvastatin. These three compounds were selected as the
hypocholesterolemic candidates for further evaluation.
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Acknowledgments
The work was supported by 'New Drug Innovation (Grant No.
2009ZX09301-007)' and ''973 project (Grant No 2010CB735601)''
from Ministry of Science and Technology of China.
Chem Biol Drug Des 2011; 78: 730–733
733