Synthesis and biological evaluation of geniposide derivatives as potent and selective PTPlB inhibitors

Article abstract of DOI:10.1016/j.ejmech.2020.112508

Herein a series of Geniposide derivatives were designed, synthesized and evaluated as protein tyrosine phosphatase 1B (PTPlB) inhibitors. Most of these compounds exhibited potent in vitro PTP1B inhibitory activities, the representative 7a and 17f were found to be the most potent inhibitors against the enzyme with IC₅₀ values of 0.35 and 0.41 μM, respectively. More importantly, they showcased 4 to10-fold selectivity over SHP2 and 3-fold over TCPTP. Further biological activity studies revealed that compounds 7a, 17b and 17f could effectively enhance insulin-stimulated glucose uptake with no significant cytotoxicity. Subsequent molecular docking and structural activity relationship analyses demonstrated that the glucose scaffold, benzylated glycosyl groups, and arylethenesulfonic acid ester significantly impact on the activity and selectivity.

Full text of DOI:10.1016/j.ejmech.2020.112508

Journal Pre-proof
Synthesis and Biological Evaluation of Geniposide derivatives as potent and selective
PTPlB inhibitors
Shuwen Lei, Dongdong Zhang, Yunyue Qi, Sharmin Reza Chowdhury, Ran Sun,
Juntao Wang, Yi Du, Lei Fu, Faqin Jiang
PII:
S0223-5234(20)30480-3
DOI:
https://doi.org/10.1016/j.ejmech.2020.112508
EJMECH 112508
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 28 April 2020
Revised Date: 20 May 2020
Accepted Date: 23 May 2020
Please cite this article as: S. Lei, D. Zhang, Y. Qi, S.R. Chowdhury, R. Sun, J. Wang, Y. Du, L. Fu, F.
Jiang, Synthesis and Biological Evaluation of Geniposide derivatives as potent and selective PTPlB
inhibitors, European Journal of Medicinal Chemistry, https://doi.org/10.1016/j.ejmech.2020.112508.
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Graphical abstract

Synthesis and Biological Evaluation of Geniposide derivatives as potent and
selective PTPlB inhibitors
Shuwen Leia, Dongdong Zhanga, Yunyue Qia, Sharmin Reza Chowdhurya, Ran Suna, Juntao
Wanga, Yi Duc, Lei Fua* and Faqin Jianga*
a. School of Pharmacy, Shanghai Jiao Tong University, No. 800 Dongchuan Rd. Minhang
District, Shanghai, 200240, PR China.
b. Viva Biotech Ltd. (Shanghai), No. 334 Aidisheng Rd., Pudong District, Shanghai, 201203,
PR China.
c. Xinhua hospital affiliated to Shanghai Jiao Tong University，School of Medicine, No. 1665
Kongjiang Rd., Yangpu District, Shanghai, 200092, PR China
Abstract
Herein a series of Geniposide derivatives were designed, synthesized and evaluated as
protein tyrosine phosphatase 1B (PTP l B) inhibitors. Most of these compounds exhibited
potent in vitro PTP1B inhibitory activities, the representative 7a and 17f were found to be the
most potent inhibitors against the enzyme with IC50 value of 0.35 and 0.41 µM, respectively.
More importantly, they showcased 4 to10-fold selectivity over SHP2 and 3-fold over TCPTP.
Further biological activity studies revealed that compounds 17b and 17f could effectively
enhance insulin-stimulated glucose uptake with no significant cytotoxicity. Subsequent
molecular docking and structural activity relationship analyses demonstrated that the glucose
scaffold, benzylated glycosyl groups, and arylethenesulfonic acid ester significantly impact
on the activity and selectivity.
Key Words: Type 2 diabetes mellitus, PTP1B inhibitors, Selectivity, Geniposide, Genipin

1. Introduction
Protein tyrosine phosphatases (PTPs) play important roles in insulin signaling
pathway by dephosphorylating tyrosine residues on insulin receptor (IR) and related
downstream substrate proteins [1-2]. Protein tyrosine phosphatase 1B (PTP1B), the
first discovered PTP family member, is a key negative regulator and effective
therapeutic target for the treatment of type 2 diabetes mellitus (T2DM). PTP1B is a
highly druggable target due to the presence of its conserved and polar active pocket.
However, discovery of drugs that target PTPlB is still a challenging task due to the
presence of the highly homologous proteins such as TCPTP and SHP2. Great effort
has been made, but none of the PTP1B inhibitors have survived through all three
phases of clinical trials, mostly due to their poor selectivity and low bioavailability
[3-4].
Natural products are the abundant resources for drug discovery. Gardenia
jasminoides Ellis, a traditional Chinese medicine, has been prescribed to treat T2DM
for several decades. This natural product contains more than 40 ingredients, among
which geniposide was found to be the most active component [5]. Geniposide (Fig. 1),
a carbohydrate compound, has a good biocompatibility and spatial flexibility [6].
Genipin (Fig. 1), a natural product also derived from the Gardenia jasminoides Ellis,
is an excellent water-soluble cross-linker.[5,7-8] Both geniposide and genipin have
many pharmacological effects, such as anti-anxiety, neuroprotective activity,
anti-Alzheimer’s disease, antithrombosis effects, anti-tumor effects and hypoglycemic
activity [9-13].
Fig. 1. The structure of geniposide and genipin.

Studies showed that introducing hydrophobic groups (Fig. 2, I) or phosphorylated
tyrosine (pTyr) mimics to a molecule (Fig. 2, II) can enhance its binding affinity to
PTP1B and the cell membrane permeability [14,15]. Our previous work also revealed
that arylethenesulfonic acid ester derivatives could act as membrane-permeable
PTP1B inhibitors (Fig. 2, III) [16-19]. In 2009, Zhang et al. demonstrated that
kinsenoside, an active compound coupled with a glycosyl moiety (Fig. 2, IV) has a
good inhibitory activity against PTP1B (IC50=5.14 μM) and high selectivity over other
PTPs [20].
The structure of geniposide is much similar to kinsenoside. Our molecular
docking analysis of geniposide binding to the PTP1B (Fig. 3) indicates that the
glycosyl moiety of geniposide locates in the catalytic active site of PTP1B and forms
hydrogen bonds with Arg220, Ser215, Ala216 and Gln265. This binding
conformation is much like that of the kinsenoside.
Based on the structural and binding conformation similarities between kinsenoside
and geniposide, we hypothesize that geniposide derivatives would offer inhibitory
activity towards PTP1B and the glycosyl group is likely to be the key moiety in our
novel PTP1B inhibitors.
Herein, we report the design, synthesis, and evaluation of geniposide derivatives
as a novel class of potent and selective PTP1B inhibitors. Specifically, to fine-tune the
hydrophobicity of geniposide, we benzylate its glycosyl groups and introduce a
variety of sulfonic esters mimicking pTyr analogs, individually and collectively.

N
N
N
O
O
O
O
O
O
OH
OH
BnO
BnO
(H2C)n
N
N
N
N
O
O
O
O
OMe
OBn
I
II
IC50=6.7±0.5 M (PTP1B)
IC50=2.8±1.0 M (PTP1B)
O
S
OH
O
O
O
H
O
HO
HO
N
N
O
H
Ph
O
OH
S
IV
IC50=5.14 M (PTP1B)
III
IC50=1.9 M (PTP1B)
Fig. 2. Representative PTP1B inhibitors.
Fig. 3. Docking of geniposide to PTP1B (PDB code 1AAX). (a) Binding conformation of
geniposide in PTP1B. (b) Geniposide in the PTP1B active site.
2. Results and discussion
2.1. Chemistry
The synthetic routes of three series of geniposide derivatives are outlined in
Schemes 1-3. Geniposide was hydrolyzed with 2 M LiOH solution overnight to
afford compound 1. The hydroxyl groups of 1 were protected by acetyl group in the
presence of Ac2O and AcONa under reflux for 2 h to give 2. Compound 2 was
condensed with various amines with EDCI overnight to give intermediate 3, which
was hydrolyzed with 2 M LiOH in 2 h to yield compounds 4a-4g.

Compounds 5, 6, 7a-7b were synthesized in three steps starting from geniposide.
The hydroxyl groups of geniposide were protected through BnBr treatment to develop
5. Subsequently, 5 was hydrolyzed with 1 M NaOH solution to give 6. Compound 6
was interacted with various amines to generate compounds 7a-7b.
Compounds 17a-17f were synthesized according to the procedure indicated in
Scheme 3. Esterification of genipin with different alcohols was catalyzed by a drop of
hydrochloric acid to obtain compounds 9a-9c. Compounds 9a-9b were chlorinated by
PhSO2Cl to yield compounds 10a-10b. The halogen of 9a-9b was substituted with
compound 11 to give 12a-12b. Subsequently, compounds 12a-12b was hydrolyzed
with 2 M LiOH to afford the 13a-13b. The coupling of 14 with compounds 13a-13b
was carried out by Wittig-Horner reactions to generate respective compounds 15a-15b.
The final compounds 17a-17f were synthesized by a condensation between compounds
15a-15b and various amines.
O
H
R1
O
H
OH
O
H
O
O
H
OH
b
c
a
O
O
O
O
H
H
AcO
O
O
AcO
H
O
O
H
OAc
OAc
HO
O
O
OAc
OAc
HO
O
O
OH
OH
AcO
AcO
HO
OH
HO
OH
OAc
OAc
OH
OH
3a-3g
2
Geniposide
1
O
H
R1
d
O
H
HO
O
O
OH
OH
HO
OH
4a-4g
Scheme 1. Synthesis of compounds 4a-4g. Reagents and conditions: (a) MeOH, 2 M LiOH·H2O,
50 °C, overnight, 90%; (b) Ac2O, AcONa, reflux, 2 h, 68%; (c) EDCI, HOBT, DIPEA, DMAP,
DMF, r.t., overnight, 19%-27%; (d) THF/H2O, 2 M LiOH·H2O, r.t., 2 h, 86%-97%.

Scheme 2. Synthesis of compounds 5, 6, 7a-7b. Reagents and conditions: (a) BnBr, NaH, TBAI,
DMF, r.t., overnight, 42%; (b) MeOH, 1 M NaOH·H2O, reflux, overnight, 90%; (c) Amine, EDCI,
HOBT, DIPEA, DMAP, DMF, r.t., overnight, 32%-52%.
O
H
O
O
H
O
O
H
O
O
H
O
OH
OHC
OHC
a
b
11
R3OH
+
O
O
O
c
O
H
H
H
HO
OH
H
O
8a R3=CH3
8b R3=Bn
O
Cl
O
R3
HO
OH
O
R3
R3
Genipin
12a-b
9a-c
8c R3=CH2CH3
10a-b
O
H
OH
O
O
H
O O
O
S
O
S
O
P
O
OHC
O
O
d
O
14
O
H
e
O
O
R3
H
O
O
R3
15a-b
13a-b
H
N
R1
O
O
H
O
S
R1-NH2
16
O
O
f
H
O
O
R3
17a-f
Scheme 3. Synthesis of compounds 17a-17f. Reagents and conditions: (a) HCl, reflux, 7 h,
56%-98%; (b) PhSO2Cl, TEA, DCM, DMAP, r.t., 40h, 48%-49%; (c) K2CO3, ACN, reflux, 12 h,
89%-91%; (d) MeOH, 2 M LiOH·H2O, r.t., 30 h, 73%-90%; (e) NaH, THF, r.t., 12 h, 42%-46%; (f)
HATU, TEA, DMF, r.t., 2.5 h, 23%-36%.
2.2. Inhibition of PTP1B
PTP1B inhibitory assay was performed on the synthesized ligands according to
the known method with sodium vanadate as a positive control [21-23]. The results are
shown in Table 1, where we firstly introduced glycosyl ring to the core structure and
verified the impact brought by different amides on the inhibitory activity. Different
anilines were introduced to improve their hydrophobic property (4a, 4b, 4c and 4d).
Compared with parent compound geniposide (IC50>500 μM), all showed obvious
enhancement in activity with IC50 value ranging from 17.69 to 100 µM accordingly
with the increase of their cLogP values. An ethyl group or an F- on the ortho- position
brings more positive effect to the ligand activity than 2-methoxyl group. We further

introduced more bulky aromatic rings on the para- position of the aniline (4e, 4f and
4g). They all generate moderate activity to PTP1B. The best ligand of this series is 4g,
which has an IC50 of 17 μM. However, the improvement is not significant.
Inspired by the former findings about increasing hydrophobicity of PTP1B
ligands, we tried to benzylate all free hydroxyl group of glycosyl moiety in the second
round of modification. As shown in Table 1, the modification brings significant
increase of hydrophobicity with cLog P value= 2.4-4.8, which lead to great
improvement in ligand activity (5, 6, 7a-7g) accordingly with IC50 value=0.35-0.68
μM. We made such an improvement regardless the different substituted amides
attached on the core structure. We then concluded that the benzylation of the glysosyl
group weights more than the modification of the amide building block. The most
potent ligands are 7a and 7b, whose IC50 values are 0.35 μM and 0.40 μM
respectively.
Though the benzylation of the glycosyl ring significantly boost the PTP1B
inhibition activity, the molecular weight increased greatly and became less drug-like .
We herein considered to use a smaller hydrophobic group as a substitution. Not only
it could bring up similar chemical physical property to the ligands as benzylated
glucose, but it has a much smaller size which makes the compounds more drug-like.
Another modification is that a sulfonic esters as pTyr mimics was introduced to the
core structure. We hope the combination of the two modifications would bring about
more significant improvement to the ligand activity. We therefore constructed genipin
derivatives as the third round modification. As shown in Table 1, genipin derivatives
(IC50=0.41-13.15 μM) with methyl or benzyl at R3 position and a sulfonic esters at R2
position exhibited better activity than genipin (IC50>500 μM). On the contrary to the
previous two rounds of modification, the benzamides greatly affect the activity of the
ligands (17a vs 9a, 17f vs 9b). The benzamide modification boost the activity over
100 times. The best ligands of this series are 17c and 17f, whose IC50 values are 1.05
μM and 0.41 μM, respectively. Based on the above findings, SARs of geniposide
derivatives can be concluded as follows, benzylation of the glycosyl groups of

geniposide or introduction of sulfonic esters as pTyr mimics to genipin can increase
the hydrophobicity of the ligand and achieve high active PTP1B inhibitors.
Table 1 PTP1B inhibitory activities of genipin and geniposide derivatives
Compound
Geniposide
R1
R2
H
R3
H
cLogPa
-2.980
IC50(μM)b
-OCH3
>500
Genipin
-OCH3
H
H
-1.016
-1. 976
>500
>100
4a
4b
4c
4d
4e
4f
4g
5
H
H
-1.714
-1.906
-2.214
-0.676
0.499
-0.362
2.497
2.480
3.614
4.803
20.82
18.27
50.76
25.23
21.74
17.69
0.68
H
H
H
H
-OCH3
-OH
-Bn
-Bn
-Bn
-Bn
6
0.38
7a
7b
0.35
0.40
9a
9b
-OCH3
-OCH3
-OCH3
-OCH3
H
H
H
-CH3
-Bn
-0.731
0.348
-0.586
2.232
236.14
40.30
64.74
13.15
9c
-CH2CH3
-Bn
15b
17a
-CH3
3.459
2.67

17b
17c
17d
17e
17f
-CH3
-CH3
-CH3
-Bn
-Bn
-
4.634
5.121
5.445
4.538
6.199
0.43
1.05
0.75
0.99
0.41
0.046
-
-
Sodium vanadate
a Values are calculated by Indraw software;
b Values are means of triplicates, repeated two times;
2.3. Selectivity over TCPTP and SHP2
Those with excellent PTP1B inhibitory activity were further assayed for their
selectivities over the other PTPs, TCPTP and SHP2. The results are shown in Table 2.
As shown in Table 2, most of the representatives of geniposide derivatives (4c & g)
had no inhibitory activity towards TCPTP or SHP2. The geniposide derivative (6)
showed moderate selective inhibitory activity over TCPTP and SHP2. Approximately
4-fold selectivity over TCPTP and 9-fold selectivity over SHP2 were obtained.
Genipin derivatives (17b & f) exhibited 3-fold selectivity over TCPTP and 4-10-fold
selectivity over SHP2.
Table 2 Inhibitory activities of geniposide derivatives on PTP1B, TCPTP and SHP2
Compound
IC50(μM)a
TCPTP
>500
PTP1B
>500
>500
18.27
17.69
0.38
SHP2
>500
>500
>100
>100
3.32
1.24
3.97
NT
Geniposide
Genipin
>500
4c
>100
4g
>100
6
1.32
17b
0.43
0.95
17f
0.41
1.29
Sodium vanadateb
0.046
0.014
a Values are means of triplicates, repeated two times;
b Sodium vanadate as a positive control;
NT means not tested.

2.4. Effects of compounds on cell viability
In order to determine the cytotoxicity of geniposide derivatives, ten compounds
(5, 6, 7a, 17a, 7b, 17b, 17c, 17d, 17e, 17f) were selected to evaluate their effects on
Chinese Hamster Ovarian (CHO) cell viability. The data is given in Table 3.
Table 3 Cytotoxicity of compounds 5, 6, 7a, 7b, 17a, 17b, 17c, 17d, 17e, 17f against CHO cells.
Compound
CHO IC50 (μM)a
5
42
6
>200
7a
>200
7b
>200
17a
17b
17c
17d
17e
17f
>200
>200
>200
>200
62
>200
a Values are means of triplicates, repeated two times;
Overall, compounds 6, 7a, 7b, 17a, 17b, 17c, 17d, 17f exhibited low toxicity
against CHO cell-line. These results indicated that the newly prepared PTP1B
inhibitors have no significant cytotoxicity.
2.5. Effects of compounds 17b and 17f on insulin-stimulated glucose uptake
It has been reported that PTP1B inhibition leads to a remarkable improvement in
insulin sensitivity and glucose metabolism [24]. We evaluated compounds 17b and
17f on 2-NBDG uptake in HepG2 cells to determine the effect. As shown in Fig. 4,
insulin-stimulated glucose uptake in HepG2 cells was increased by Pioglitazone
which was used as the positive control, and the increased percentages were 33.8%,
46.0%, and 63.5% at the concentrations of 5, 10 and 20 μM, respectively. The glucose
uptake in HepG2 cells was also significantly increased by treatment with tested
compounds (17b and 17f). The increased percentages were 38.6%, 68% and 69.6%
for 17b, and 43.8%, 68.3% and 64.4% for 17f at 5, 10 and 20 μM, respectively. The
results were slightly better than the positive control.

Fig. 4. Effect of compound 17b and 17f on insulin-stimulated glucose uptake. Each value was
presented as mean ±SD, n=3; (**) P<0.01 vs the insulin-treated group.
2.6. Molecular docking study

Fig. 5. Docking of compound 4g, 7a and 17f to PTP1B (PDB code 1AAX). (a) Binding
conformation of compound 4g in PTP1B. (b) Compound 4g in the PTP1B active site. (c) Binding
conformation of compound 7a in PTP1B. (d) Compound 7a in the PTP1B active site. (e) Binding
conformation of compound 17f in PTP1B. (f) Compound 17f in the PTP1B active site.
The representative compounds 4g, 7a and 17f were selected for molecular
docking studies (Fig. 5). The glycosyl moiety of compound 4g was found to locate in
the catalytic active site of PTP1B. The hydroxyl group on the glycosyl group formed
multiple hydrogen bonds with Ser215, Asp180, Ser221, Ala216, Phe181 and Arg220.
The diphenyl ether partially extended into the secondary aryl phosphorylation binding
site and established hydrophobic interaction with several residues including Lys35,
Arg46. The core structure of geniposide formed a π-π conjugation with Tyr45. These
multiple interactions determined the activity of 4g. The benzyl groups of 7a bound to
the peripheral hydrophobic surface of PTP1B, formed hydrophobic interactions with
Arg220, Asp180, and Gln261, which enhanced the binding affinity with PTP1B. The
amide carbonyl formed two hydrogen bonds with Arg46. The sulfonic acid ester
formed hydrogen bonds with Try45 and Lys119. In addition, the benzyl rings on the
glycosyl group formed a π-π conjugation with Phe181, which improved the binding to
PTP1B. The sulfonic acid ester of 17f extended to the catalytic active site of PTP1B
and formed hydrogen bonds with Arg23 and Arg253, which enhanced the binding
affinity. The diphenylether side chain formed hydrophobic interactions with Lys115
and Asp28. The benzyl groups of 17f formed a π-π conjugation with Phe181. Then,
the core structure of genipin formed a hydrogen bond with Ser215, which increased
the binding to PTP1B.

3. Conclusions
In this study, a series of geniposide and genipin derivatives were designed and
synthesized as potent and selective PTP1B inhibitors. Most compounds demonstrated
potent PTP1B inhibitory activity and moderate selectivity over TCPTP and SHP2.
SARs study showed that benzylated glycosyl group on geniposide derivatives and
sulfonic ester as pTyr mimics on genipin derivatives are vital for the enhancement of
hydrophobic property and activity. Molecular docking analysis suggested the binding
mode and explained the reasons for better inhibitory activity and selectivity. We
further verified the effects of compounds 17b and 17f on insulin-stimulated glucose
uptake with no significant cytotoxicity. This study demonstrated a good strategy to
discover effective and safe genipin-based PTP1B inhibitors.
4. Experiment section
4.1. Chemistry section
All the starting solvents and reagents were obtained from commercial suppliers,
without further purification. PTP1B, TCPTP, SHP2 proteins were purchased from
Viva Biotech (Shanghai) Ltd. 1H NMR spectra was recorded on an Agilent 400 MHz
NMR. 13C NMR spectra was recorded on an Agilent 100 MHz NMR. Chemical
shifts were expressed in parts per million (ppm). Coupling constants were in units of
Hertz (Hz). Splitting patterns describe apparent multiplicities were designated as s
(singlet), d (doublet), t (triplet), q (quartet), m (multiplet) and br (broad).
High-resolution mass spectra (HRMS) and compound purity data were acquired on an
Agilent 6200 TOF LC/MS system, UV detector (220 and 254 nm). All the reactions
were monitored by Thin-layer chromatography (silica gel, aluminum sheets 60 F254),
which was conducted with Qingdao Huanghai (China). All the crude products were
purified by column chromatography using silica gel (100-200 mesh or 300-400mesh),
purchased from Qingdao Haiyang Chemical Co. Ltd. (China).
4.1.1. Synthesis of geniposidic acid (1)
Geniposide (3 g, 0.00773 mol) was dissolved in methanol (25 mL) and
hydrolyzed with 2 M LiOH solution (8 mL, 0.01546 mol) at 50 °C overnight. Then 1
N hydrochloric acid solution (10 mL) was added by drops until the PH became a

weak acid. The reaction solution was concentrated under reduced pressure to afford a
light brown solid (compound 1) in 90% yield. 1H NMR (400 MHz, CD3OD): δ 7.43 (s,
1H), 5.64 (br. s., 1H), 5.08 (d, J=6.65 Hz, 1H), 5.02 (d, J=5.09 Hz, 1H), 4.95 (d,
J=5.09 Hz, 1H), 4.92 (d, J=5.48 Hz, 1H), 4.70 (t, J=5.48 Hz, 1H), 4.49 (d, J=7.83 Hz,
1H), 4.44 (t, J=5.87 Hz, 1H), 4.08 (br. s., 1H), 3.90-4.02 (m, 3H), 3.34-3.42 (m, 1H),
2.91-3.16 (m, 6H), 2.57-2.70 (m, 2H), 2.45-2.48 (m, 1H).
4.1.2. Synthesis of 7-acetyl-1-(3,4,5,6-tetraacetate)geniposidic acid (2)
To the solution of compound 1 (2.6 g, 0.00695 mol) in acetic anhydride (20 mL),
sodium acetate (2 g, 0.024 mol) was slowly added and heated under reflux for 1 hour.
After the initial reaction was completed, water (50 mL) was added, and the reaction
solvent was extracted with ethyl acetate (40 mL×3). The combined organic was
washed with saturated brine (30 mL×3), dried over anhydrous sodium sulfate, and
concentrated under reduced pressure to generate a crude product. The crude product
was purified by silica gel column purification (petroleum ether: ethyl acetate=2:1) to
give compound 2 as a light yellow solid (3.05 g, 0.00522 mol) with a yield of 68%.
1H NMR (400 MHz, CDCl3): δ 7.53 (s, 1H), 5.87 (br. s., 1H), 5.20-5.27 (m, 1H),
5.09-5.17 (m, 2H), 5.02 (t, J=8.80 Hz, 1H), 4.88 (d, J=8.22 Hz, 1H), 4.70 (d, J=8.22
Hz, 2H), 4.22-4.29 (m, 1H), 4.19 (d, J=2.35 Hz, 1H), 3.70-3.75 (m, 1H), 3.20-3.27 (m,
1H), 2.86-2.95 (m, 2H), 2.23-2.26 (m, 3H), 2.09 (d, J=3.13 Hz, 12H).
4.1.3. General method for the synthesis of compound 3a-3g
To the stirred solution of compound 2 (200 mg, 0.00034 mol) in DMF (3 mL),
EDCI (78 mg, 0.00041 mol), HOBT (55 mg, 0.00041 mol) and DIPEA (0.2 mL,
0.00102 mol) was added, and stirred at room temperature for 2 hours. Then organic
amine (0.00037 mol) and DMAP (63 mg, 0.00051 mol) was added, and continually
stirred at room temperature overnight. After the reaction was completed, the solution
was added to ice water (30 mL) and extracted with ethyl acetate (20 mL×3). The
combined organic solvent was washed with 1 N aqueous citric acid (20 mL×3), 1 N
aqueous dilute hydrochloric acid (20 mL×3) and saturated brine (20 mL×3). The

organic phase was dried over anhydrous sodium sulfate, concentrated under reduced
pressure, and purified by column chromatography on silica gel to afford compound
3a-3g in a yield of 19-27%.
4.1.3.1. 7-acetyl-1-(3,4,5,6-tetraacetate)-4-((phenylcarbamoyl)amide)geniposide 3a
Yield 21%. 1H NMR (400 MHz, CDCl3): δ 7.49 (d, J=7.43 Hz, 2H), 7.32 (d,
J=6.65 Hz, 2H), 7.18 (br. s., 1H), 7.10 (d, J=7.83 Hz, 1H), 5.82 (br. s., 1H), 5.18-5.29
(m, 2H), 5.10 (t, J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz, 1H), 4.87 (d, J=7.83 Hz, 1H),
4.62-4.76 (m, 2H), 4.26 (d, J=11.35 Hz, 1H), 4.15 (d, J=11.35 Hz, 1H), 3.72 (d,
J=9.78 Hz, 1H), 3.31 (br. s., 1H), 3.02 (br. s., 1H), 2.82 (d, J=9.39 Hz, 1H), 2.26 (d,
J=19.17 Hz, 1H), 2.15 (d, J=18.39 Hz, 2H), 1.94-2.03 (m, 14H).
4.1.3.2.
7-acetyl-1-(3,4,5,6-tetraacetate)-4-(((2-fluorophenyl)carbamoyl)amide)geniposide 3b
Yield 21%. 1H NMR (400 MHz, CDCl3): δ 8.28 (t, J=8.22 Hz, 1H), 7.40 (br. s.,
1H), 7.34 (br. s., 1H), 7.00-7.15 (m, 3H), 5.82 (br. s., 1H), 5.18-5.29 (m, 2H), 5.10 (t,
J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz, 1H), 4.87 (d, J=7.83 Hz, 1H), 4.62-4.76 (m, 2H),
4.26 (d, J=11.35 Hz, 1H), 4.15 (d, J=11.35 Hz, 1H), 3.72 (d, J=9.78 Hz, 1H), 3.31 (br.
s., 1H), 3.02 (br. s., 1H), 2.82 (d, J=9.39 Hz, 1H), 2.26 (d, J=19.17 Hz, 1H), 2.15 (d,
J=18.39 Hz, 2H), 1.94-2.03 (m, 14H).
4.1.3.3.
7-acetyl-1-(3,4,5,6-tetraacetate)-4-(((2-ethylphenyl)carbamoyl)amide)geniposide 3c
Yield 20%. 1H NMR (400 MHz, CDCl3): δ 7.84 (d, J=7.43 Hz, 1H), 7.32 (s, 1H),
7.20 (br. s., 2H), 7.06-7.14 (m, 2H), 5.82 (br. s., 1H), 5.18-5.29 (m, 2H), 5.10 (t,
J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz, 1H), 4.87 (d, J=7.83 Hz, 1H), 4.62-4.76 (m, 2H),
4.26 (d, J=11.35 Hz, 1H), 4.15 (d, J=11.35 Hz, 1H), 3.72 (d, J=9.78 Hz, 1H), 3.31 (br.
s., 1H), 3.02 (br. s., 1H), 2.82 (d, J=9.39 Hz, 1H), 2.60 (q, J=7.56 Hz, 2H), 2.26 (d,
J=19.17 Hz, 1H), 2.15 (d, J=18.39 Hz, 2H), 1.94-2.03 (m, 14H), 1.29-1.38 (m, 3H).

4.1.3.4.
7-acetyl-1-(3,4,5,6-tetraacetate)-4-(((2-methoxyphenyl)carbamoyl)amide)geniposide
3d
Yield 27%. 1H NMR (400 MHz, CDCl3): δ 8.34 (d, J=7.43 Hz, 1H), 7.89 (br. s.,
1H), 7.35 (br. s., 1H), 7.01 (d, J=7.04 Hz, 1H), 6.94 (t, J=7.24 Hz, 1H), 6.87 (d,
J=7.43 Hz, 1H), 5.82 (br. s., 1H), 5.18-5.29 (m, 2H), 5.10 (t, J=9.98 Hz, 1H), 5.00 (t,
J=9.00 Hz, 1H), 4.87 (d, J=7.83 Hz, 1H), 4.62-4.76 (m, 2H), 4.26 (d, J=11.35 Hz, 1H),
4.15 (d, J=11.35 Hz, 1H), 3.84-3.92 (s, 3H), 3.72 (d, J=9.78 Hz, 1H), 3.31 (br. s., 1H),
3.02 (br. s., 1H), 2.82 (d, J=9.39 Hz, 1H), 2.26 (d, J=19.17 Hz, 1H), 2.15 (d, J=18.39
Hz, 2H), 1.94-2.03 (m, 14H).
4.1.3.5.
7-acetyl-1-(3,4,5,6-tetraacetate)-4-(((4-phenoxyphenyl)carbamoyl)amide)geniposide
3e
Yield 19%. 1H NMR (400 MHz, CDCl3): δ 7.45 (d, J=8.22 Hz, 2H), 7.30 (t,
J=8.02 Hz, 2H), 7.16 (br. s., 1H), 7.07 (t, J=7.43 Hz, 1H), 6.93-7.01 (m, 3H), 5.82 (br.
s., 1H), 5.18-5.29 (m, 2H), 5.10 (t, J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz, 1H), 4.87 (d,
J=7.83 Hz, 1H), 4.62-4.76 (m, 2H), 4.26 (d, J=11.35 Hz, 1H), 4.15 (d, J=11.35 Hz,
1H), 3.72 (d, J=9.78 Hz, 1H), 3.31 (br. s., 1H), 3.02 (br. s., 1H), 2.82 (d, J=9.39 Hz,
1H), 2.26 (d, J=19.17 Hz, 1H), 2.15 (d, J=18.39 Hz, 2H), 1.94-2.03 (m, 14H).
4.1.3.6.
7-acetyl-1-(3,4,5,6-tetraacetate)-4-(((4-([1,1’-biphenyl]-4-yloxy)phenyl)carbamoyl)a
mide)geniposide 3f
Yield 26%. 1H NMR (400 MHz, CDCl3): δ 7.50-7.57 (m, 4H), 7.48 (d, J=8.61 Hz,
2H), 7.37-7.44 (m, 2H), 7.32 (d, J=7.43 Hz, 1H), 7.19 (br. s., 1H), 7.02 (d, J=7.43 Hz,
4H), 5.82 (br. s., 1H), 5.18-5.29 (m, 2H), 5.10 (t, J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz,
1H), 4.87 (d, J=7.83 Hz, 1H), 4.62-4.76 (m, 2H), 4.26 (d, J=11.35 Hz, 1H), 4.15 (d,
J=11.35 Hz, 1H), 3.72 (d, J=9.78 Hz, 1H), 3.31 (br. s., 1H), 3.02 (br. s., 1H), 2.82 (d,

J=9.39 Hz, 1H), 2.26 (d, J=19.17 Hz, 1H), 2.15 (d, J=18.39 Hz, 2H), 1.94-2.03 (m,
14H).
4.1.3.7.
7-acetyl-1-(3,4,5,6-tetraacetate)-4-(((4-(4-(tert-butyl)phenoxy)phenyl)carbamoyl)ami
de)geniposide 3g
Yield 24%. 1H NMR (400 MHz, CDCl3): δ 7.43 (d, J=8.61 Hz, 2H), 7.31 (d,
J=8.22 Hz, 2H), 7.15 (br. s., 1H), 6.96 (d, J=8.22 Hz, 2H), 6.89 (d, J=7.43 Hz, 2H),
5.82 (br. s., 1H), 5.18-5.29 (m, 2H), 5.10 (t, J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz, 1H),
4.87 (d, J=7.83 Hz, 1H), 4.62-4.76 (m, 2H), 4.26 (d, J=11.35 Hz, 1H), 4.15 (d,
J=11.35 Hz, 1H), 3.72 (d, J=9.78 Hz, 1H), 3.31 (br. s., 1H), 3.02 (br. s., 1H), 2.82 (d,
J=9.39 Hz, 1H), 2.26 (d, J=19.17 Hz, 1H), 2.15 (d, J=18.39 Hz, 2H), 1.94-2.03 (m,
14H), 1.29 (s, 9H).
4.1.4. General method for the synthesis of 4a-4g
Compound 3(a-g) (50 mg, 0.00007 mol) were dissolved in THF/H2O (3 mL/1
mL). Then, 2 N aqueous lithium hydroxide solution (0.2 mL, 0.00042 mol) was added
and the mixture was stirred at room temperature for 2 hours. Then 1 N hydrochloric
acid solution (0.5 mL) was added dropwise until the PH to weak acid. The organic
solvent was concentrated under reduced pressure and purified by column
chromatography on silica gel to get compound 4a-4g in a yield of 86-97%.
4.1.4.1. 4-((phenylcarbamoyl)amide)geniposide 4a
Brown solid, yield 86%.1H NMR (400 MHz, CD3OD): δ 7.54 (d, J=7.43 Hz, 2H),
7.32 (d, J=6.65 Hz, 2H), 7.18 (br. s., 1H), 7.10 (d, J=7.83 Hz, 1H), 5.82 (br. s., 1H),
5.18-5.29 (m, 2H), 5.10 (t, J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz, 1H), 4.87 (d, J=7.83
Hz, 1H), 4.62-4.76 (m, 2H), 4.26 (d, J=11.35 Hz, 1H), 4.15 (d, J=11.35 Hz, 1H), 3.72
(d, J=9.78 Hz, 1H), 3.31 (br. s., 1H), 3.02 (br. s., 1H), 2.82 (d, J=9.39 Hz, 1H), 2.26
(d, J=19.17 Hz, 1H). 13C NMR (100 MHz, CD3OD): δ 167.4, 147.3, 143.3, 138.3,
128.3, 126.7, 123.8, 120.9, 120.8, 115.6, 98.7, 96.3, 96.2, 76.9, 76.4, 74.9, 73.5, 70.1,

67.8, 61.2, 60.0, 46.0, 37.7, 34.8. HRMS (ESI): Calcd. for C22H27NO9 [M+H]+
450.1759, found 450.1771.
4.1.4.2. 4-(((2-fluorophenyl)carbamoyl)amide)geniposide 4b
Light green solid, yield 91%.1H NMR (400 MHz, CD3OD): δ 8.28 (t, J=8.22 Hz,
1H), 7.40 (br. s., 1H), 7.34 (br. s., 1H), 7.00-7.15 (m, 3H), 5.82 (br. s., 1H), 5.18-5.29
(m, 2H), 5.10 (t, J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz, 1H), 4.87 (d, J=7.83 Hz, 1H),
4.62-4.76 (m, 2H), 4.26 (d, J=11.35 Hz, 1H), 4.15 (d, J=11.35 Hz, 1H), 3.72 (d,
J=9.78 Hz, 1H), 3.31 (br. s., 1H), 3.02 (br. s., 1H), 2.82 (d, J=9.39 Hz, 1H), 2.26 (d,
J=19.17 Hz, 1H). 13C NMR (100 MHz, CD3OD): δ 167.4, 147.3, 143.3, 138.3, 128.3,
126.7, 123.8, 120.9, 115.6, 98.7, 96.3, 76.9, 76.4, 74.9, 73.5, 70.1, 67.8, 61.2, 60.0,
46.0, 37.7, 34.8. HRMS (ESI): Calcd. for C22H26FNO9 [M+H]+ 468.1664, found
468.9272.
4.1.4.3. 4-(((2-ethylphenyl)carbamoyl)amide)geniposide 4c
White solid, yield 91%.1H NMR (400 MHz, CD3OD): δ 7,84 (d, J=7.43 Hz, 1H),
7.32 (s, 1H), 7.20 (br. s., 2H), 7.06-7.14 (m, 2H), 5.82 (br. s., 1H), 5.18-5.29 (m, 2H),
5.10 (t, J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz, 1H), 4.87 (d, J=7.83 Hz, 1H), 4.62-4.76
(m, 2H), 4.26 (d, J=11.35 Hz, 1H), 4.15 (d, J=11.35 Hz, 1H), 3.72 (d, J=9.78 Hz, 1H),
3.31 (br. s., 1H), 3.02 (br. s., 1H), 2.82 (d, J=9.39 Hz, 1H), 2.60 (q, J=7.56 Hz, 2H),
2.26 (d, J=19.17 Hz, 1H), 1.29-1.38 (m, 3H). 13C NMR (100 MHz, CD3OD): δ 168.1,
147.6, 143.4, 140.3, 134.9, 128.5, 127.3, 126.7, 126.6, 125.9, 114.9, 98.7, 96.4, 76.9,
76.4, 73.5, 70.1, 61.2, 60.0, 45.9, 37.9, 34.9, 24.2, 13.5. HRMS (ESI): Calcd. for
C24H31NO9 [M+H]+ 478.2072, found 478.2074.
4.1.4.4. 4-(((2-methoxyphenyl)carbamoyl)amide)geniposide 4d
Black solid, yield 87%.1H NMR (400 MHz, CD3OD): δ 8.34 (d, J=7.43 Hz, 1H),
7.89 (br. s., 1H), 7.35 (br. s., 1H), 7.01 (d, J=7.04 Hz, 1H), 6.94 (t, J=7.24 Hz, 1H),
6.87 (d, J=7.43 Hz, 1H), 5.82 (br. s., 1H), 5.18-5.29 (m, 2H), 5.10 (t, J=9.98 Hz, 1H),
5.00 (t, J=9.00 Hz, 1H), 4.87 (d, J=7.83 Hz, 1H), 4.62-4.76 (m, 2H), 4.26 (d, J=11.35

Hz, 1H), 4.15 (d, J=11.35 Hz, 1H), 3.84-3.92 (s, 3H), 3.72 (d, J=9.78 Hz, 1H), 3.31
(br. s., 1H), 3.02 (br. s., 1H), 2.82 (d, J=9.39 Hz, 1H), 2.26 (d, J=19.17 Hz, 1H). 13C
NMR (100 MHz, CD3OD): δ 168.1, 147.6, 143.4, 140.3, 134.9, 128.5, 127.3, 126.7,
126.6, 125.9, 114.9, 98.7, 96.4, 76.9, 76.4, 73.5, 70.1, 61.2, 60.0, 55.8, 45.9, 37.9,
34.9. HRMS (ESI): Calcd. for C23H29NO9 [M+H]+ 480.1864, found 480.1859.
4.1.4.5. 4-(((4-phenoxyphenyl)carbamoyl)amide)geniposide 4e
Green solid, yield 97%.1H NMR (400 MHz, CD3OD): δ 7.45 (d, J=8.22 Hz, 2H),
7.30 (t, J=8.02 Hz, 2H), 7.16 (br. s., 1H), 7.07 (t, J=7.43 Hz, 1H), 6.93-7.01 (m, 3H),
5.82 (br. s., 1H), 5.18-5.29 (m, 2H), 5.10 (t, J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz, 1H),
4.87 (d, J=7.83 Hz, 1H), 4.62-4.76 (m, 2H), 4.26 (d, J=11.35 Hz, 1H), 4.15 (d,
J=11.35 Hz, 1H), 3.72 (d, J=9.78 Hz, 1H), 3.31 (br. s., 1H), 3.02 (br. s., 1H), 2.82 (d,
J=9.39 Hz, 1H), 2.26 (d, J=19.17 Hz, 1H). 13C NMR (100 MHz, CD3OD): δ 167.3,
157.6, 153.4, 147.4, 143.3, 133.9, 129.5, 126.7, 122.8, 122.7, 122.6, 118.8, 118.7,
118.0, 115.4, 98.7, 96.3, 76.9, 76.4, 73.5, 70.1, 61.2, 60.0, 45.9, 37.8, 34.8, 29.4.
HRMS (ESI): Calcd. for C28H31NO10 [M+H]+ 542.2021, found 542.2002.
4.1.4.6. 4-(((4-([1,1’-biphenyl]-4-yloxy)phenyl)carbamoyl)amide)geniposide 4f
Brown solid, yield 91%.1H NMR (400 MHz, CD3OD): δ 7.50-7.57 (m, 4H), 7.48
(d, J=8.61 Hz, 2H), 7.37-7.44 (m, 2H), 7.32 (d, J=7.43 Hz, 1H), 7.19 (br. s., 1H), 7.02
(d, J=7.43 Hz, 4H), 5.82 (br. s., 1H), 5.18-5.29 (m, 2H), 5.10 (t, J=9.98 Hz, 1H), 5.00
(t, J=9.00 Hz, 1H), 4.87 (d, J=7.83 Hz, 1H), 4.62-4.76 (m, 2H), 4.26 (d, J=11.35 Hz,
1H), 4.15 (d, J=11.35 Hz, 1H), 3.72 (d, J=9.78 Hz, 1H), 3.31 (br. s., 1H), 3.02 (br. s.,
1H), 2.82 (d, J=9.39 Hz, 1H), 2.26 (d, J=19.17 Hz, 1H). 13C NMR (100 MHz,
CD3OD): δ 167.3, 157.1, 153.3, 147.5, 143.2, 140.2, 136.0, 134.0, 128.5, 128.0, 126.8,
126.7, 126.3, 122.8, 122.7, 118.9, 118.3, 115.4, 98.7, 96.4, 76.8, 76.3, 73.4, 70.1, 61.1,
60.0, 45.9, 37.8, 34.8, 22.5. HRMS (ESI): Calcd. for C34H35NO10 [M+H]+ 618.2334,
found 618.2299.
4.1.4.7. 4-(((4-(4-(tert-butyl)phenoxy)phenyl)carbamoyl)amide)geniposide 4g

Green solid, yield 97%.1H NMR (400 MHz, CD3OD): δ 7.43 (d, J=8.61 Hz, 2H),
7.31 (d, J=8.22 Hz, 2H), 7.15 (br. s., 1H), 6.96 (d, J=8.22 Hz, 2H), 6.89 (d, J=7.43 Hz,
2H), 5.82 (br. s., 1H), 5.18-5.29 (m, 2H), 5.10 (t, J=9.98 Hz, 1H), 5.00 (t, J=9.00 Hz,
1H), 4.87 (d, J=7.83 Hz, 1H), 4.62-4.76 (m, 2H), 4.26 (d, J=11.35 Hz, 1H), 4.15 (d,
J=11.35 Hz, 1H), 3.72 (d, J=9.78 Hz, 1H), 3.31 (br. s., 1H), 3.02 (br. s., 1H), 2.82 (d,
J=9.39 Hz, 1H), 2.26 (d, J=19.17 Hz, 1H), 1.29 (s, 9H). 13C NMR (100 MHz,
CD3OD): δ 167.3, 155.0, 153.8, 147.6, 145.8, 143.1, 133.5, 126.9, 126.3, 122.8, 122.7,
118.4, 117.8, 115.4, 115.3, 98.7, 96.4, 76.7, 76.2, 73.4, 70.1, 67.8, 66.9, 61.1, 60.0,
45.9, 37.8, 34.8, 33.7, 30.7, 22.4. HRMS (ESI): Calcd. for C32H39NO10 [M+H]+
598.2647, found 598.2619.
4.1.5. Synthesis of 7-benzyl-1-(3,4,5,6-tetrabenzyl)geniposide (compound 5)
To a stirred solution of geniposide (3 g, 0.00773 mol) in DMF (150 mL), in an
ice-water bath, 60% sodium hydride (1.854 g, 0.04638 mol) was slowly added. After
the mixture was stirred for 1 hour, benzyl bromide (5.5 mL, 0.04638 mol) was slowly
added dropwise and stirred at room temperature overnight. After the reaction was
over, the reaction solution was added to ice water (900 mL) and extracted with ethyl
acetate (200 mL×3), then the combined organic solvent was washed with 1 N
hydrochloric acid solution (50 mL×2) and saturated brine (100 mL×3), dried over
anhydrous sodium sulfate and concentrated under reduced pressure to get crude
product. The crude product was purified by silica gel column (petroleum ether: ethyl
acetate=7:1) to give compound 5 as a colorless liquid (2.667 g, 0.00318 mol) with a
yield of 42%. 1H NMR (400 MHz, CDCl3): δ 7.48 (s, 1H), 7.21-7.32 (m, 23H),
7.15-7.20 (m, 2H), 5.83 (br. s., 1H), 5.30 (d, J=5.8 Hz, 1H), 4.86-4.93 (m, 2H),
4.75-4.84 (m, 3H), 4.44-4.64 (m, 7H), 4.19 (br. s., 2H), 4.11 (q, J=7.0 Hz, 1H), 3.65
(s, 3H), 3.40-3.48 (m, 2H), 3.28 (q, J=7.4 Hz, 1H), 2.95 (t, J=6.8 Hz, 1H), 2.87 (dd,
J=7.8, 16.4 Hz, 1H), 2.14-2.27 (m, 2H).
4.1.6. Synthesis of 7-benzyl-1-(3,4,5,6-tetrabenzyl)geniposidic acid (compound 6)

To a stirred solution of compound 5 (2.667 g, 0.00318 mol) in methanol (20 mL),
1 N aqueous sodium hydroxide (6.36 mL, 0.00636 mol) was added and refluxed
overnight. After the reaction was over, the reaction solution was concentrated under
reduced pressure, and 1 N hydrochloric acid solution was added dropwise until the PH
to 1-2. A white solid precipitated, filtered and dried at vacuum to give compound 6
(2.36 g, 0.017 mol) as a pale yellow solid, yield of 90%. 1H NMR (400 MHz, CDCl3):
δ 7.48 (s, 1H), 7.21-7.32 (m, 23H), 7.15-7.20 (m, 2H), 5.83 (br. s., 1H), 5.30 (d, J=5.8
Hz, 1H), 4.86-4.93 (m, 2H), 4.75-4.84 (m, 3H), 4.44-4.64 (m, 7H), 4.19 (br. s., 2H),
4.11 (q, J=7.0 Hz, 1H), 3.40-3.48 (m, 2H), 3.28 (q, J=7.4 Hz, 1H), 2.95 (t, J=6.8 Hz,
1H), 2.87 (dd, J=7.8, 16.4 Hz, 1H), 2.14-2.27 (m, 2H). 13C NMR (100 MHz, CDCl3):
δ174.9, 148.8, 140.3, 138.6, 138.4, 138.2, 138.1, 129.9, 128.5,128.3, 127.9, 127.8,
127.7, 127.5, 127.4, 116.3, 99.3, 96.8, 84.3, 81.4, 77.4, 77.2, 75.5, 74.9, 74.5, 73.2,
72.4, 68.5, 46.3, 39.1, 36.1. HRMS (ESI): Calcd. for C51H52O10 [M+Na]+ 847.3453,
found 847.3455.
4.1.7. General method for the synthesis of 7a-7b
Compound 6 (200 mg, 0.00024 mol) was dissolved in DMF (4 mL), then EDCI
(56 mg, 0.00029 mol), HOBT (39 mg, 0.00029 mol) and DIPEA (0.13 mL, 0.00072
mol) were added and stirred at room temperature for 2 hours. Then organic amine
(0.00026 mol) and DMAP (45 mg, 0.00036 mol) were added and stirred at room
temperature overnight. After the reaction was completed, the solution was added to
ice water (30 mL) and extracted with ethyl acetate (20 mL×3). The combined organic
solvent were washed with 1 N aqueous citric acid (20 mL×3), 1 N aqueous dilute
hydrochloric acid (20 mL×3) and saturated brine (20 mL×3), dried over anhydrous
sodium sulfate, concentrated under reduced pressure and purified by column
chromatography on silica gel to afford compound 7a-7b in a yield of 32-52%.
4.1.7.1. 7-benzyl-1-(3,4,5,6-tetrabenzyl)-4-(4-ethylvinylsulfonate
benzyl)carbamoyl)amide)geniposide 7a

Light yellow solid, yield 32%. 1H NMR (400 MHz, CDCl3): δ 8.07 (d, J=8.6 Hz,
1H), 8.01 (s, 1H), 7.19-7.43 (m, 29H), 5.93 (br. s., 1H), 5.46 (d, J=6.2 Hz, 1H),
4.80-5.00 (m, 5H), 4.72 (d, J=10.9 Hz, 1H), 4.48-4.62 (m, 5H), 4.25 (br. s., 2H),
3.63-3.78 (m, 4H), 3.41-3.56 (m, 4H), 3.09 (t, J=6.8 Hz, 1H), 2.99 (dd, J=8.2, 16.4 Hz,
1H), 2.39 (dd, J=5.4, 17.2 Hz, 1H), 0.83-0.93 (m, 3H). 13C NMR (100 MHz, CDCl3):
δ 163.1, 156.6, 143.5, 139.6, 138.5, 138.2, 138.1, 138.0, 137.9, 129.9, 128.9, 128.6,
128.5, 128.5, 128.4, 128.3, 128.0, 127.9, 127.9, 127.9, 127.8, 127.7, 127.7, 127.6,
124.7, 120.4, 108.4, 107.0, 99.7, 97.0, 84.5, 81.7, 77.5, 75.8, 75.3, 75.1, 75.0, 73.5,
72.4, 68.6, 67.9, 46.4, 38.4, 34.1. HRMS (ESI): Calcd. for C62H65NO12S [M+K]+
1086.38, found 1086.44.
4.1.7.2.
7-benzyl-1-(3,4,5,6-tetrabenzyl)-4-(((4-phenoxyphenyl)carbamoyl)amide)geniposide
7b
White solid, yield 52%. 1H NMR (400 MHz, CDCl3): δ 7.40 (d, J=8.6 Hz, 2H),
7.25-7.33 (m, 19H), 7.24 (s, 6H), 7.20 (d, J=4.7 Hz, 3H), 7.16 (d, J=7.4 Hz, 2H),
7.04-7.10 (m, 2H), 6.96 (d, J=8.6 Hz, 4H), 5.84 (br. s., 1H), 5.30 (d, J=6.2 Hz, 1H),
4.91 (d, J=10.9 Hz, 2H), 4.75-4.85 (m, 3H), 4.63 (d, J=10.5 Hz, 1H), 4.44-4.56 (m,
5H), 4.20 (s, 2H), 3.56-3.73 (m, 5H), 3.37-3.49 (m, 3H), 2.99-3.05 (m, 1H), 2.83 (d,
J=8.6 Hz, 1H), 2.17-2.30 (m, 2H), 1.98 (br. s., 1H). 13C NMR (100 MHz, CDCl3): δ
165.2, 157.6, 153.3, 147.2, 140.0, 138.5, 138.2, 138.1, 138.0, 138.0, 133.4, 129.7,
129.2, 128.4, 128.4, 128.3, 128.2, 128.0, 127.8, 127.7, 127.7, 127.6, 127.6, 123.0,
121.8, 119.7, 118.3, 115.6, 99.3, 95.8, 84.5, 81.8, 77.6, 75.8, 75.2, 75.1, 74.9, 73.4,
72.4, 68.7, 67.9, 46.9, 38.3, 34.2. HRMS (ESI): Calcd. for C63H61NO10 [M+Na]+
1014.4188, found 1014.4153.
4.1.8. General method for the synthesis of 9a-9c
Genipin (10 g, 0.044 mol) was dissolved in corresponding alcohols (8a-8c) (70
mL). The solution was catalyzed by two drops of concentrated hydrochloric acid, and
then heated at reflux for 7 hours. After this reaction was completed, the solution was

concentrated under reduced pressure. A few drops of 1 N sodium hydroxide solution
were added to adjust the pH to neutral, and then it was extracted three times with
ethyl acetate. The combined organic phase was washed three times with saturated
brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure
to get compound 9a-9c in a yield of 56-98%.
4.1.8.1. 1-methoxygenipin 9a
Brown oily liquid, yield 98%.1H NMR (400 MHz, CDCl3): δ 7.41-7.54 (m, 1H),
5.75-5.88 (m, 1H), 4.46 (d, J=8.2 Hz, 1H), 4.12-4.28 (m, 2H), 3.64-3.75 (m, 3H),
3.40-3.54 (m, 3H), 3.13-3.23 (m, 1H), 2.86 (dd, J=8.5, 16.5 Hz, 1H), 2.59 (t, J=7.6 Hz,
1H), 2.06 (dd, J=9.0, 16.4 Hz, 1H).
4.1.8.2. 1-benzyloxygenipin 9b
Yellow oil, yield 56%.1H NMR (400 MHz, CDCl3): δ 7.56 (s, 1H), 7.28-7.42 (m,
5H), 5.82 (br. s., 1H), 5.00 (d, J=11.3 Hz, 1H), 4.60-4.72 (m, 2H), 4.23 (br. s., 2H),
3.68-3.77 (m, 3H), 3.13-3.27 (m, 1H), 2.74-2.94 (m, 1H), 2.69 (t, J=8.0 Hz, 1H), 2.08
(dd, J=9.0, 16.4 Hz, 1H).
4.1.8.3. 1-ethoxygenipin 9c
Brown oil, yield 97%.1H NMR (400 MHz, CDCl3): δ 7.50 (s, 1H), 5.82 (br. s.,
1H), 4.53 (d, J=8.4 Hz, 1H), 4.25 (br. s., 2H), 4.03 (qd, J=7.1, 9.5 Hz, 2H), 3.69-3.74
(m, 3H), 3.14-3.24 (m, 1H), 2.87 (tdd, J=1.3, 8.5, 16.5 Hz, 1H), 2.58 (t, J=8.0 Hz, 1H),
2.01-2.12 (m, 1H), 1.17-1.29 (t, 3H).
4.1.9. General method for the synthesis of 10a-10b
Compound 9a-9b (0.044 mol), triethylamine (8.905 g, 0.088 mol) was dissolved
in dichloromethane (70 mL), and slowly added to benzenesulfonyl chloride (11.656 g,
0.066 mol) in ice water. After catalyzed by the addition of DMAP (200 mg), the
reaction was carried out at room temperature for 40 hours. When the reaction was
completed, the reaction solution was washed with 1 N sodium hydroxide solution (10

mL×3) and extracted with dichloromethane (30 mL×3). The combined organic
solvent was washed with 1 N hydrochloric acid solution and saturated brine (30
mL×3), dried over anhydrous sodium sulfate, concentrated under reduced pressure
and purified by silica gel column chromatography to generate compound 10a-10b in a
yield of 48-49%.
4.1.9.1. 1-methoxy-2-(chloromethyl)genipin 10a
Colorless liquid, yield 49%.1H NMR (400 MHz, CDCl3): δ 7.52 (s, 1H),
5.95-6.01 (m, 1H), 4.51 (d, J=7.8 Hz, 1H), 4.20-4.25 (m, 2H), 3.73 (s, 3H), 3.57 (s,
3H), 3.24 (q, J=8.4 Hz, 1H), 2.91 (dd, J=8.4, 15.8 Hz, 1H), 2.73-2.81 (m, 1H),
2.01-2.18 (m, 1H).
4.1.9.2. 1-benzyloxy-2-(chloromethyl)genipin 10b
Light yellow liquid, yield 48%.1H NMR (400 MHz, CDCl3): δ 7.56 (s, 1H),
7.28-7.42 (m, 5H), 5.82 (br. s., 1H), 5.00 (d, J=11.3Hz, 1H), 4.60-4.72 (m, 2H), 4.23
(br. s., 2H), 3.68-3.77 (m, 3H), 3.13-3.27 (m, 1H), 2.74-2.94 (m, 1H), 2.69 (t, J=8.0
Hz, 1H), 2.08 (dd, J=9.0, 16.4 Hz, 1H).
4.1.10. General method for the synthesis of 12a-12b
Compound 10a-10b (0.022 mol), p-hydroxybenzaldehyde (3.175 g, 0.026 mol)
and anhydrous potassium carbonate (4.561 g, 0.033 mol) was dissolved in acetonitrile
(50 mL) and the mixture was heated under reflux for 12 hours. After the reaction was
completed, it was concentrated under reduced pressure. The residue was washed with
1 N sodium hydroxide solution (10 mL), and extracted with dichloromethane (30
mL×3). The combined organic solvent was washed with 1 N hydrochloric acid
solution, saturated brine and dried over anhydrous sodium sulfate, concentrated under
reduced pressure to afford compound 12a-12b in a yield of 89-91%.
4.1.10.1. 1-methoxy-2-((4-formylphenoxy)methyl)genipin 12a

Brown liquid, yield 89%.1H NMR (400 MHz, CDCl3): δ 9.90 (s, 1H), 7.83-7.86
(m, 2H), 7.50-7.55 (m, 1H), 7.01-7.05 (m, J=8.6 Hz, 2H), 5.96-6.03 (m, 1H),
4.69-4.84 (m, 2H), 4.53 (d, J=7.8 Hz, 1H), 3.74 (s, 3H), 3.56-3.60 (m, 3H), 3.20-3.29
(m, 1H), 2.87-2.99 (m, 1H), 2.70 (t, J=7.6 Hz, 1H), 2.13 (dd, J=8.7, 16.9 Hz, 1H).
4.1.10.2. 1-benzyloxy-2-((4-formylphenoxy)methyl)genipin 12b
White solid, yield 91%.1H NMR (400 MHz, CDCl3): δ 9.79-9.87 (m, 1H),
7.69-7.79 (m, 3H), 7.53 (s, 1H), 7.15-7.25 (m, 4H), 6.89 (d, J=8.6 Hz, 2H), 5.92 (br. s.
1H), 4.97 (d, J=11.3 Hz, 1H), 4.57-4.75 (m, 4H), 3.69-3.73 (m, 3H), 3.10-3.28 (m,
1H), 2.90 (dd, J=8.4, 16.0 Hz, 1H), 2.74 (t, J=8.0 Hz, 1H), 2.08 (dd, J=8.6, 16.8 Hz,
1H).
4.1.11. General method for the synthesis of 13a-13b
To a stirred solution of compound 12a-12b (0.019 mol) in methanol (50 mL), 2 N
aqueous lithium hydroxide solution (47.5 mL, 0.095 mol) was added at room
temperature and maintained for 30 hours. After the reaction was completed, the
reaction solution was concentrated under reduced pressure, and adjusted PH value to
1-2 with 1 N hydrochloric acid solution, and dried to get compound 13a-13b in a yield
of 73-90%.
4.1.11.1. 1-methoxy-2-((4-formylphenoxy)methyl)genipher-5-carboxylic acid 13a
Yellow solid, yield 90%.1H NMR (400 MHz, CDCl3): δ 9.90 (s, 1H), 7.83-7.86
(m, 2H), 7.50-7.55 (m, 1H), 7.01-7.05 (m, J=8.6 Hz, 2H), 5.96-6.03 (m, 1H),
4.69-4.84 (m, 2H), 4.53 (d, J=7.8 Hz, 1H), 3.74 (s, 3H), 3.20-3.29 (m, 1H), 2.87-2.99
(m, 1H), 2.70 (t, J=7.6 Hz, 1H), 2.13 (dd, J=8.7, 16.9 Hz, 1H).
4.1.11.2. 1-benzyloxy-2-((4-formylphenoxy)methyl)genipher-5-carboxylic acid 13b
White solid, yield 73%.1H NMR (400 MHz, CDCl3): δ 9.79-9.87 (m, 1H),
7.69-7.79 (m, 3H), 7.53 (s, 1H), 7.15-7.25 (m, 4H), 6.89 (d, J=8.6 Hz, 2H), 5.92 (br. s.

1H), 4.97 (d, J=11.3 Hz, 1H), 4.57-4.75 (m, 4H), 3.10-3.28 (m, 1H), 2.90 (dd, J=8.4,
16.0 Hz, 1H), 2.74 (t, J=8.0 Hz, 1H), 2.08 (dd, J=8.6, 16.8 Hz, 1H).
4.1.12. General method for the synthesis of 15a-15b
Compound 13a-13b (0.017 mol) and ethyl
(diethoxyphosphoryl)methanesulfonate (6.194 g, 0.024 mol) was dissolved in
tetrahydrofuran (30 mL). 60% sodium hydrogen (2.040 g, 0.051 mol) was added
slowly in portions and stirred at room temperature for 12 hours. After the completion
of this reaction, the mixture was concentrated under reduced pressure, adjusted pH to
neutral with a 1 N hydrochloric acid solution and extracted with ethyl acetate (30
mL×3). The organic phase was washed with saturated brine, dried over anhydrous
sodium sulfate, concentrated under reduced pressure, and purified with silica gel
column to afford compound 15a-15b in a yield of 42-46%.
4.1.12.1. 1-methoxy-2-((4-ethylvinylsulfonate phenoxy)methyl)genipin-5-carboxylic
acid 15a
White solid, yield 42%.1H NMR (400 MHz, CDCl3): δ 7.65 (s, 1H), 7.55 (d,
J=15.6 Hz, 1H), 7.43-7.49 (m, J=8.6 Hz, 2H), 6.94-6.99 (m, J=8.6 Hz, 2H), 6.60 (d,
J=15.6 Hz, 1H), 5.98 (br. s., 1H), 4.65-4.79 (m, 2H), 4.57 (d, J=7.8 Hz, 1H),
4.08-4.26 (m, 2H), 3.59 (s, 3H), 3.19-3.28 (m, 1H), 2.94 (dd, J=8.8, 16.6 Hz, 1H),
2.71 (t, J=8.0 Hz, 1H), 2.16 (dd, J=8.4, 16.6 Hz, 1H), 1.32-1.44 (t, 3H).
4.1.12.2. 1-benzyloxy-2-((4-ethylvinylsulfonate phenoxy)methyl)genipin-5-carboxylic
acid 15b
White solid, yield 46%.1H NMR (400 MHz, CDCl3): δ 7.69-7.79 (m, 3H), 7.55 (d,
J=15.6 Hz, 1H), 7.25 (s, 1H), 7.15-7.25 (m, 4H), 6.89 (d, J=8.6 Hz, 2H), 6.60 (d,
J=15.6 Hz, 1H), 5.92 (br. s., 1H), 4.97 (d, J=11.3 Hz, 1H), 4.57-4.75 (m, 4H),
4.08-4.26 (m, 2H), 3.10-3.28 (m, 1H), 2.90 (dd, J=8.4, 16.0 Hz, 1H), 2.74 (t, J=8.0 Hz,
1H), 2.08 (dd, J=8.6, 16.8 Hz, 1H), 1.32-1.44 (t, 3H).

4.1.12. General method for the synthesis of 17a-d
Compound 15a (0.00014 mol) and organic amine (0.00015 mol) was dissolved in
DMF (3 mL), stirred in ice water bath. Then HATU (64 mg, 0.00017 mol) and
triethylamine (28 mg, 0.00028 mol) were added and stirred at room temperature for
2.5 hours. After this reaction was completed, the solution was added to ice water (30
mL) and 1 N hydrochloric acid dropwise solution to PH =1-2. Yellow solids were
precipitated, filtered, and then purified by silica gel column chromatography to
develop compound 17a-17d in a yield of 27-36%.
4.1.12.1. 1-methoxy-2-((4- ethylvinylsulfonate
phenoxy)methyl)-5-((4-(p-phenyleneoxy)phenyl)amide))genipin 17a
Brown solid, yield 30%.1H NMR (400 MHz, CDCl3): δ 7.55 (d, J=15.4 Hz, 1H),
7.41-7.52 (m, 5H), 7.28-7.36 (m, 3H), 7.06-7.12 (m, 1H), 6.94-7.03 (m, 5H), 6.60 (d,
J=15.4 Hz, 1H), 5.96-6.04 (m, 1H), 4.61-4.81 (m, 2H), 4.57 (d, J=7.8 Hz, 1H),
4.08-4.27 (m, 2H), 3.52-3.64 (m, 3H), 3.34-3.45 (m, 1H), 2.86-2.97 (m, 1H), 2.78 (t,
J=7.9 Hz, 1H), 2.18-2.29 (m, 1H), 1.35-1.44 (t, 3H). 13C NMR (100 MHz, CDCl3): δ
165.2, 161.2, 157.5, 153.4, 147.9, 144.3, 138.6, 133.3, 130.2, 130.0, 129.6, 124.7,
123.0, 121.9, 119.6, 118.3, 115.2, 115.2, 114.6, 102.2, 66.5, 66.4, 57.0, 46.4, 38.7,
35.6, 14.1. HRMS (ESI): Calcd. for C33H33NO8S [M+Na]+ 626.1819, found 626.1815.
4.1.12.2. 1-methoxy-2-((4- ethylvinylsulfonate
phenoxy)methyl)-5-((4-p-diphenylether)phenyl)amide))genipin 17b
Brown liquid, yield 27%.1H NMR (400 MHz, CDCl3): δ 7.55-7.59 (m, 4H),
7.51-7.55 (m, 2H), 7.49 (d, J=7.4 Hz, 2H), 7.41-7.47 (m, 4H), 7.31-7.36 (m, 1H),
7.02-7.09 (m, 4H), 6.97 (d, J=8.8 Hz, 2H), 6.60 (d, J=15.6 Hz, 1H), 5.96-6.04 (m,
1H), 4.66-4.81 (m, 2H), 4.57 (d, J=7.8 Hz, 1H), 4.09-4.27 (m, 2H), 3.60 (s, 3H),
3.35-3.44 (m, 1H), 2.88-2.98 (m, 2H), 2.79 (t, J=7.8 Hz, 1H), 2.20-2.31 (m, 1H),
1.38-1.45 (t, 3H). 13C NMR (100 MHz, CDCl3): δ 165.2, 161.2, 157.1, 153.3, 150.5,
147.9, 144.3, 140.4, 138.6, 136.1, 133.4, 133.1, 130.3, 130.0, 128.8, 128.4, 127.0,
126.9, 124.7, 121.9, 119.8, 118.5, 118.3, 115.3, 114.6, 102.2, 66.6, 66.4, 57.0, 46.4,

38.8, 35.6, 14.9. HRMS (ESI): Calcd. for C39H37NO8S [M+Na]+ 702.2132, found
702.2127.
4.1.12.3. 1-methoxy-2-((4- ethylvinylsulfonate
phenoxy)methyl)-5-((4-(2-chloro-4-fluoro-phenylene ether)benzeneamide)genipin 17c
Brown liquid, yield 36%.1H NMR (400 MHz, CDCl3): δ 7.37-7.58 (m, 7H),
7.17-7.23 (m, 1H), 7.07 (dt, J=1.3, 7.7 Hz, 1H), 6.91-6.98 (m, 4H), 6.59 (d, J=15.4 Hz,
1H), 5.94-6.03 (m, 1H), 4.65-4.80 (m, 2H), 4.55 (d, J=7.8 Hz, 1H), 4.09-4.25 (m, 2H),
3.58 (s, 3H), 3.36-3.42 (m, 1H), 2.90 (dd, J=8.6, 16.2 Hz, 1H), 2.76 (t, J=7.9 Hz, 1H),
2.22 (dd, J=8.3, 16.3 Hz, 1H), 1.36-1.43 (m, 3H). 13C NMR (100 MHz, CDCl3): δ
165.2, 161.2, 153.1, 152.7, 147.9, 144.4, 138.6, 133.5, 130.7, 130.3, 130.1, 127.9,
125.4, 124.7, 124.5, 121.9, 120.2, 118.7, 118.3, 115.3, 114.6, 102.2, 66.6, 66.4, 57.0,
46.4, 38.7, 35.6, 14.8. HRMS (ESI): Calcd. for C33H31BrClNO8S [M+Na]+ 738, found
739.
4.1.12.4. 1-methoxy-2-((4- ethylvinylsulfonate
phenoxy)methyl)-5-((4-(biphenylether)phenyl)amide)genipin 17d
Brown solid, yield 32%.1H NMR (400 MHz, CDCl3): δ 7.76-7.82 (m, 2H), 7.65
(d, J=7.8 Hz, 1H), 7.47-7.53 (m, 3H), 7.40-7.46 (m, 3H), 7.30-7.40 (m, 2H),
7.19-7.25(m, 2H), 7.00-7.05 (m, 2H), 6.93 (d, J=8.6 Hz, 2H), 6.57 (d, J=15.2 Hz, 1H),
5.94-6.02 (m, 1H), 4.62-4.77 (m, 2H), 4.54 (d, J=7.8 Hz, 1H), 4.06-4.22 (m, 2H), 3.56
(s, 3H), 3.37 (q, J=8.2 Hz, 1H), 2.90 (dd, J=8.4, 16.2 Hz, 1H), 2.72-2.80 (m, 1H),
2.17-2.27 (m, 1H), 1.37 (t, J=7.2 Hz, 3H). 13C NMR (100 MHz, CDCl3): δ 165.2,
161.2, 155.4, 153.3, 148.0, 144.3, 138.6, 134.2, 133.6, 130.3, 130.0, 129.9, 127.7,
127.0, 126.5, 124.7, 124.6, 122.0, 120.0, 119.6, 118.3, 115.3, 114.6, 113.3, 102.3,
66.6, 66.4, 57.0, 46.4, 38.8, 35.6, 14.9. HRMS (ESI): Calcd. for C37H35NO8S
[M+Na]+ 676.1976, found 676.1972.
4.1.13. General method for the synthesis of 17e-17f