New chiral derivatives of xanthones: Synthesis and investigation of enantioselectivity as inhibitors of growth of human tumor cell lines

Article abstract of DOI:10.1016/j.bmc.2013.12.042

A highly efficient and practical methodology for synthesis of new chiral derivatives of xanthones (CDXs) in enantiomerically pure form has been developed According to this approach, thirty CDXs (3-32) were synthesized by coupling a carboxyxanthone (1) and a carboxymethoxyxanthone (2) with both enantiomers of commercially available chiral building blocks, namely six amino alcohols, one amine and one amino ester The activation of the carboxylic acid group of the xanthonic scaffold was carried out with the coupling reagent O-(benzotriazol-1-yl)-N-N-N′-N′-tetramethyluronium tetrafluoroborate (TBTU), in the presence of a catalytic amount of TEA in anhydrous THF The coupling reactions with the chiral blocks were performed at room temperature with short reactions times, excellent yields (ranging from 94% to 99%), and very high enantiomeric excess The synthesized CDXs were evaluated for their effect on the in vitro growth of three human tumor cell lines, namely A375-C5 (melanoma), MCF-7 (breast adenocarcinoma), and NCI-H460 (non-small cell lung cancer) The most active compound was CDX 15 being active in all human tumor cell lines with values of GI₅₀ of 32.15 ± 2.03 μM for A375-C5, 22.55 ± 1.99 μM for MCF-7, and 14.05 ± 1.82 μM for NCI-H460 Nevertheless, some CDXs showed cell-type selectivity Furthermore, the growth inhibitory effects, in some cases, demonstrated to be depending on the stereochemistry of the CDXs An interesting example was observed with the enantiomers 3 and 4, which demonstrated high enantioselectivity for MCF-7 and NCI-H460 cell lines It can be inferred that the effects on the growth of the human tumor cell lines can be ascribed not only to the nature and positions of substituents on the xanthonic scaffold but also to the stereochemistry of the CDXs Some considerations regarding structure-activity relationship within this class of compounds will be highlighted

Full text of DOI:10.1016/j.bmc.2013.12.042

Bioorganic & Medicinal Chemistry 22 (2014) 1049–1062
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
New chiral derivatives of xanthones: Synthesis and investigation
of enantioselectivity as inhibitors of growth of human tumor cell
lines
Carla Fernandes a,b,c, , Kamonporn Masawang a,d, , Maria Elizabeth Tiritan a,c,e, Emília Sousa a,b,c
,
Virgínia de Lima f, Carlos Afonso a,b,c, Hassan Bousbaa a,c,e, Wanwisa Sudprasert d, Madalena Pedro a,e,
,
⇑
Madalena M. Pinto a,b,c,
⇑
a Centro de Química Medicinal da Universidade do Porto (CEQUIMED-UP), Rua de Jorge Viterbo Ferreira 228, 4050-313 Porto, Portugal
b Laboratório de Química Orgânica e Farmacêutica, Departamento de Ciências Químicas, Faculdade de Farmácia, Universidade do Porto, Rua de Jorge Viterbo Ferreira 228,
4050-313 Porto, Portugal
c Centro Interdisciplinar de Investigação Marinha e Ambiental (CIIMAR/CIMAR), Universidade do Porto, Rua dos Bragas 289, 4050-123 Porto, Portugal
d Faculty of Science, Kasetsart University, 10900 Bangkok, Thailand
e Cooperativa de Ensino Superior, Politécnico e Universitário (CESPU), Centro de Investigação em Ciências da Saúde, Instituto Superior de Ciências da Saúde-Norte (CICS-ISCS-N),
Rua Central de Gandra 1317, 4585-116 Gandra PRD, Portugal
f Universidade Federal do Rio de Janeiro, Instituto de Química, LADETEC—LAB RES, Rio de Janeiro, RJ, Brazil
a r t i c l e i n f o
a b s t r a c t
Article history:
A highly efficient and practical methodology for synthesis of new chiral derivatives of xanthones (CDXs)
in enantiomerically pure form has been developed. According to this approach, thirty CDXs (3–32) were
synthesized by coupling a carboxyxanthone (1) and a carboxymethoxyxanthone (2) with both enantio-
mers of commercially available chiral building blocks, namely six amino alcohols, one amine and one
amino ester. The activation of the carboxylic acid group of the xanthonic scaffold was carried out with
the coupling reagent O-(benzotriazol-1-yl)-N-N-N0-N0-tetramethyluronium tetrafluoroborate (TBTU), in
the presence of a catalytic amount of TEA in anhydrous THF. The coupling reactions with the chiral blocks
were performed at room temperature with short reactions times, excellent yields (ranging from 94% to
99%), and very high enantiomeric excess. The synthesized CDXs were evaluated for their effect on the
in vitro growth of three human tumor cell lines, namely A375-C5 (melanoma), MCF-7 (breast adenocar-
cinoma), and NCI-H460 (non-small cell lung cancer). The most active compound was CDX 15 being active
Received 11 September 2013
Revised 10 December 2013
Accepted 17 December 2013
Available online 31 December 2013
Keywords:
Chiral derivatives of xanthones
Enantiomerically pure
TBTU
Antitumor
Enantioselectivity
in all human tumor cell lines with values of GI50 of 32.15 2.03
lM for A375-C5, 22.55 1.99 lM for
MCF-7, and 14.05 1.82 M for NCI-H460. Nevertheless, some CDXs showed cell-type selectivity. Fur-
l
thermore, the growth inhibitory effects, in some cases, demonstrated to be depending on the stereochem-
istry of the CDXs. An interesting example was observed with the enantiomers 3 and 4, which
demonstrated high enantioselectivity for MCF-7 and NCI-H460 cell lines. It can be inferred that the effects
on the growth of the human tumor cell lines can be ascribed not only to the nature and positions of sub-
stituents on the xanthonic scaffold but also to the stereochemistry of the CDXs. Some considerations
regarding structure–activity relationship within this class of compounds will be highlighted.
Ó 2013 Elsevier Ltd. All rights reserved.
1. Introduction
requirements gave a great contribution to the considerable in-
creased interest in this field. The gains in potency, efficacy, safety
The number of enantiomerically pure drugs in the pharmaceu-
tical market has been increasing.1 The different biological/pharma-
cological activities of enantiomers, together with the advances in
synthetic and analytical methodologies as well as the regulatory
and selectivity obtained by treatment with single enantiomers
rather than racemates are undeniable.2 The enantiomerically pure
drugs can be obtained either by preparative resolution of a
cracemate or by enantioselective synthesis of the desired enantio-
mer. Regarding the early steps of drug development, the HPLC
chromatographic resolution of a racemate using chiral stationary
phases (CSPs) is the preferential approach since it can provide both
enantiomers with high enantiomeric excess (ee).3 Concerning
enantioselective synthesis, it can be useful when a large amount
⇑
Corresponding authors. Tel.: +351 24157190 (M.P.); tel.: +351 222078692;
fax: +351 226093390 (M.M.P.).
E-mail addresses: madalena.pedro@iscsn.cespu.pt (M. Pedro), madalena@
ff.up.pt, madalenakijjoa@gmail.com (M.M. Pinto).
These authors equally contributed to this work.
0968-0896/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmc.2013.12.042

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C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
enantiomers using commercially available chiral reagent in enanti-
omerically pure form as building blocks can be an interesting ap-
proach to obtain both enantiomers in high enantiomeric purity.
Xanthone (9H-xanthon-9-one) derivatives comprise an impor-
tant class of oxygenated heterocycles associated to a large spec-
trum of biological and pharmacological activities.4–9 For the last
several years, searching of new xanthone derivatives with poten-
tial biological/pharmacological properties has remained in the area
of interest of our group.5–18 Despite the massive research regarding
xanthone derivatives, there are few examples in the literature con-
cerning synthetic chiral derivatives of xanthones (CDXs).17,19–24
Among them there are, for example, chiral xanthonolignoids with
an inhibitory effect on the in vitro growth of different human tu-
mor cell lines.17 Moreover, the referred biological activity demon-
strated to be dependent on the stereochemistry.25
In this paper we report the synthesis of CDXs in enantiomeri-
cally pure form by coupling two carboxyxanthone derivatives,
6-methoxy-9-oxo-9H-xanthene-2-carboxylic acid (1) and 2-((9-
oxo-9H-xanthen-3-yl)oxy)acetic acid (2) (Fig. 1), with both
enantiomers of eight commercially available chiral building
blocks, using O-(benzotriazol-1-yl)-N-N-N0-N0-tetramethyluronium
Figure 1. Structures of carboxyxanthone 1 and carboxymethoxyxanthone 2 used as
chemical substrates.
of only one enantiomer is required.3 However, if both enantiomers
are required two independent syntheses are necessary, which is
usually a difficult task when specific and expensive reagents, such
as asymmetric catalysts, enzymes, chiral auxiliaries, are needed.
Besides, the enantiomeric purity of the final product usually is
not high. On the other hand, the strategy to obtain pure
Figure 2. Structures of CDXs 3–32 (the used numbering concerns the NMR assignments).

C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
1051
tetrafluoroborate (TBTU) as the coupling reagent. This methodol-
ogy required mild reaction conditions, was straightforwardly and
very high yields were achieved with any degree of racemisation.
Thirty CDXs were obtained (Fig. 2) with six (15–18, 31 and 32)
being here described for the first time. The synthesized CDXs, in
enantiomerically pure form, were evaluated for their effect on
the in vitro growth of three human tumor cell lines namely
A375-C5 (melanoma), MCF-7 (breast adenocarcinoma), and
NCI-H460 (non-small cell lung cancer). The structural aspects, con-
cerning the nature and positions of substituents on the xanthonic
scaffold as well as the stereochemistry of the library of CDXs, are
discussed. Some considerations regarding structure–activity
relationship are also described.
ether synthesis, based on the nucleophilic substitution of 3-hydro-
xy-9H-xanthen-9-one (37) with methyl bromoacetate in alkaline
medium (Scheme 1). The methyl ester 38 was then hydrolyzed
providing the carboxymethoxyxanthone building block 2. To our
best knowledge, the synthesis of the carboxymethoxyxanthone 2
through the alkaline cyclization of the benzophenone intermediate
(35) using MW irradiation is described here for the first time.
2.1.2. Synthesis of chiral derivatives of xanthones 3–32
In the course of our studies aiming to enlarge our library of new
biologically active CDXs, we were interested in establishing an effi-
cient, practical, and mild methodology for coupling carboxyxanth-
one derivatives with enantiomerically pure chiral building blocks
through an amide bond. Amines, amino alcohols and amino esters
as building blocks were chosen to demonstrate the efficiency and
applicability of our methodology.
2. Results and discussion
2.1. Chemistry
Activation of a carboxyl group for the formation of amide bonds
can be achieved either by conversion into more reactive functional
groups such as acyl halide, anhydride, acyl azide or by in situ acti-
vation using coupling reagents. In the case of the carboxylic acid
groups on the xanthones 1–2, using tionyl chloride to obtain the
respective acyl halides, an extensive degradation of the chemical
substrates was observed. In the past few years, the design and syn-
thesis of innovative coupling reagents has been an area of intense
investigation and several became commercially available.31–34
Among of them, the coupling reagent dicyclo-hexylcarboiimide
(DCC) was used in the scope of this study and furnished the desir-
able products; however, the formation of the secondary product
dicyclohexylurea led to very low yields (data not shown). Nowa-
days, the most popular coupling reagents are probably onium
(phosphonium and uronium) salts.35 One of them, TBTU demon-
strated to be very efficient for the synthesis of different classes of
compounds such as peptides,36 amides,37,38 phenylhydrazides,37
esters,39,40 1,3,4, oxadiazoles,41 acid azides,42 and thiol esters.43
This coupling reagent was also employed by our group to success-
fully synthesize of thirty CDXs (3–32) by coupling reactions of two
different carboxyxanthone derivatives (1 and 2) with a variety of
2.1.1. Synthesis of carboxyxanthone derivatives 1 and 2
Carboxyxanthone derivatives are not only interesting bioactive
compounds26,27 but they are also very important as basis to molec-
ular modifications tending to obtain new chiral derivatives.19 The
carboxyxanthone 1 was obtained in our group via Ullmann reac-
tion, with the formation of a diaryl ether intermediate, as described
elsewhere.28 The carboxymethoxyxanthone 2 was synthesized via
a benzophenone intermediate by a multi-step pathway as illus-
trated on Scheme 1. The syntheses of the benzophenone 3529 as
well as the 3-methoxy-9H-xanthen-9-one (36)29,30 and 3-hydro-
xy-9H-xanthen-9-one (37) intermediates have already been
described.30 However, in order to decrease the reaction time and
to improve the yield, the alkaline cyclization of the benzophenone
35 was carried out under microwave (MW) irradiation instead of
conventional heating. With MW methodology, the reaction time
was reduced from 36 h to 30 min. Furthermore, the yield of the
synthesis of 3-methoxy-9H-xanthen-9-one (36) increased from
83% to 92%. The methyl 2-((9-oxo-9H-xanthen-3-yl)oxy)acetate
(38) (described here for the first time) was obtained by Williamson
Scheme 1. Total synthesis of carboxymethoxyxanthone 2.

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C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
Table 1
Synthesis of CDXs 3–18 by coupling reactions of carboxyxanthone 1 with chiral compounds 39–54
O
O
O
TBTU, TEA,
COOH
THF anhydrous
R
R2
+
rt
R1
H3CO
O
H3CO
O
1
H2N
39-54
3-18
Compound
R
R1
Compound
R2
Time (h)
0.5
Yield (%)
98
ee (%)
>99
39a
CH3
3a
N
H
40b
41a
CH3
4b
5a
N
H
0.5
2
97
96
>98
>99
OH
CH(CH3)2
CH2OH
CH2OH
N
H
42b
43a
CH(CH3)2
6b
7a
2
2
98
98
>99
>99
OH
OH
OH
N
H
CH2CH(CH3)2
CH2OH
CH2OH
CH2OH
CH2OH
N
H
44b
45a
46b
CH2CH(CH3)2
8b
2
3
3
99
96
95
>99
>99
>99
N
H
9a
OH
N
H
10b
OH
N
H
OH
47a
48b
CH3
CH3
CH2OH
CH2OH
11a
12b
0.5
0.5
97
96
>99
>99
N
H
OH
OH
OH
N
H
49a
50b
H
H
CH(CH3)OH
13a
14b
0.5
0.5
96
97
>99
>99
N
H
N
H
CH(CH3)OH
CHOH
51c
52d
15c
16d
5
5
94
95
>97
>98
OH
N
H
CHOH
OH
N
H
O
53a
54b
CH3
CH3
COOC(CH3)3
COOC(CH3)3
17a
18b
4
4
96
97
>99
>99
N
H
O
O
O
N
H
a
b
c
(R)-Enantiomer.
(S)-Enantiomer.
(R,S)-Enantiomer.
(S,R)-Enantiomer.
d
commercially available chiral blocks, namely (R)-(+)-
zylamine (39), (S)-(ꢀ)-
(41), (S)-(+)-valinol (42), (R)-(ꢀ)-leucinol (43), (S)-(+)-leucinol
(44), (R)-(ꢀ)- -phenylglycinol (45), (S)-(+)- -phenylglycinol (46),
a
-dimethylben-
(R)-(ꢀ)-2-amino-1-propanol (47), (S)-(+)-2-amino-1-propanol (48),
(R)-(ꢀ)-1-amino-2-propanol (49), (S)-(+)-1-amino-2-propanol (50),
(1R,2S)-(ꢀ)-2-amino-1,2-diphenylethanol (51), (1S,2R)-(+)-2-
amino-1,2-diphenylethanol (52), (R)-alanine tert-butyl ester (53),
a
-dimethylbenzylamine (40), (R)-(ꢀ)-valinol
a
a

C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
1053
(S)-alanine tert-butyl ester (54) (Tables 1 and 2). Our selection in-
cluded both enantiomers of enantiomerically pure building blocks
with no tendency towards racemization or enantiomeric intercon-
version, and having a primary amine as reactive group for amide
formation. The coupling reactions were performed in anhydrous
THF using equivalent amounts of the carboxyxanthone derivative
substrate, the corresponding commercial chiral block and TBTU
at room temperature with 2 equiv of TEA.
desired amides in excellent yields (ranging from 94% to 99%).
Actually, TBTU can also be used at room temperature for the syn-
thesis of esters from carboxylic acids and either aliphatic alcohols
or phenols in the presence of organic bases.39,40 Interestingly,
according to these yields, the coupling reactions with chiral amino
alcohols (41–52) were highly selective for the more nucleophilic
group of the building blocks, that is, the amine instead of the
alcohol group.
As shown in Tables 1 and 2, the syntheses worked equally well
with both carboxyxanthone derivatives (1 and 2) affording the
Regarding the reaction times, some differences were observed.
Indeed, the coupling reactions with the amines or amino alcohols
Table 2
Synthesis of CDXs 19–32 by coupling reactions of carboxymethoxyxanthone 2 with chiral compounds 39–52
O
O
O
TBTU, TEA,
THF anhydrous
R
+
H2N
R2
rt
R1
O
O
OCH2COOH
O
19-32
39-52
2
Compound
R
R1
Compound
R2
Time (h)
0.5
Yield (%)
97
ee (%)
>99
H
N
39a
CH3
19a
H
N
40b
41a
42b
CH3
20b
21a
22b
0.5
2
96
95
97
>98
>99
>99
H
N
CH(CH3)2
CH(CH3)2
CH2OH
CH2OH
OH
OH
OH
H
N
2
H
N
43a
44b
45a
46b
CH2CH(CH3)2
CH2CH(CH3)2
CH2OH
CH2OH
CH2OH
CH2OH
23a
24b
25a
26b
2
2
3
3
98
96
94
95
>99
>99
>99
>99
H
N
OH
H
N
OH
H
N
OH
H
N
47a
48b
CH3
CH3
CH2OH
CH2OH
27a
28b
0.5
0.5
97
96
>99
>99
OH
H
N
OH
H
N
49a
50b
H
H
CH(CH3)OH
CH(CH3)OH
29a
30b
0.5
0.5
98
97
>99
>99
OH
OH
H
N
H
N
51c
52d
31c
32d
5
5
95
94
>98
>99
CHOH
CHOH
OH
H
N
OH
a
b
c
(R)-Enantiomer.
(S)-Enantiomer.
(R,S)-Enantiomer.
(S,R)-Enantiomer.
d

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C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
bearing alkyl groups were faster and completed after 0.5–2 h,
while the reactions involving amino esters or amino alcohols
bearing aryl groups were only completed after 3–5 h. However,
concerning the amino alcohols bearing alkyl groups, it was found
that increasing the size of the alkyl group, reaction times also aug-
mented. For example, the coupling reactions for amino alcohols
with a methyl group (47–50) were completed after 0.5 h, while
the reactions for amino alcohols with an isopropyl group (41–42)
needed 2 h to be completed. Furthermore, the coupling reactions
with the amino alcohols possessing two aryl groups in the chiral
moiety (51–52) were slower (5 h) compared with the reactions
involving amino alcohols with one aryl group (3 h) (45–46). It
can be inferred, that the steric hindrance created by the bulky alkyl
groups and aryl groups of the amino alcohols may be responsible
for the observed results.
2.2. Biological evaluation
2.2.1. Effect on the growth of human tumor cell lines
The library of thirty CDXs, in enantiomerically pure form, were
evaluated on the in vitro growth of three human tumor cell lines:
A375-C5 (melanoma), MCF-7 (breast adenocarcinoma), and
NCI-H460 (non-small cell lung cancer). This drug-screening proce-
dure, adopted from the National Cancer Institute (NCI, USA), uses
the protein binding dye, SRB, to assess cell growth.46 For each com-
pound,
a dose–response curve was established. The results,
expressed as the concentration that was able to cause 50% cell
growth inhibition (GI50), are summarized in Table 3.
The overall results obtained demonstrate that some CDXs exhib-
itedinterestinggrowthinhibitoryeffectsonthetestedhumantumor
cell lines. Actually, the most active CDX in all human tumor cell lines
The purification procedures were easy, involving simple wash-
ing of the crude reaction products with 1 M HCl solution and
saturated solution of NaHCO3 to remove the by-product
1-hydroxybenzotriazole, as well as the excess of reagents, followed
by recrystallization from ethyl acetate/n-hexane or EtOH or chloro-
form/n-hexane to afford the CDXs.
was compound 15 presenting values of GI50 = 32.15 2.03
GI50 = 22.55 1.99 M and GI50 = 14.05 1.82 M for A375-C5,
MCF-7, and NCI-H460, respectively.
lM,
l
l
Moreover, the evaluation of the growth inhibitory effect of the
series of CDXs synthesized by coupling the carboxyxanthone 1
with different chiral amino alcohols ( CDXs 5–16) allowed taking
some considerations regarding structure–activity relationship.
Accordingly, it was found that the CDXs bearing aryl groups in
the chiral moiety (9–10, 15–16) were the most active compounds
in all human tumor cell lines tested. However, a slight decrease
of activity was observed for the CDXs possessing only one aryl
group (9 and 10) when compared with the CDXs possessing two
aryl groups (15 and 16). Regarding the CDXs bearing alkyl groups
in the chiral moiety (5–8, 11–14) it was observed that decreasing
the size of the alkyl group the growth inhibitor effect also de-
creased or was abolished. For example, the CDXs with an isopropyl
group in the chiral moiety, namely CDXs 7 and 8 presented values
HPLC using different types of CSPs was used to determine the
enantiomeric purity of the synthesized CDXs (Tables 1 and 2; see
Section
4 for details). The HPLC methods development are
described elsewhere.44,45
Thus, the synthetic methodology used herein provides to be
very efficient, broad-scope applicability, inexpensive and opera-
tionally simplest for obtaining both enantiomers with high ee.
Additionally, the synthesis procedure can be easily scaled-up for
both enantiomers in order to obtain available quantities for biolog-
ical/pharmacological assays.
Table 3
Growth inhibitory activity of thirty CDXs on the growth of three human tumor cell lines
Compound
GI50 (lM)
A375-C5 (melanoma)
MCF-7 (breast adenocarcinoma)
NCI-H460 (non-small cell lung cancer)
3
4
5
6
>150
>150
102.10 1.24
>150
>150
85.88 5.30
42.62 1.77
114.06 3.80
>150
91.91 6.27
125.34 9.81
>150
7
8
9
112.94 8.38
69.72 0.03
57.72 2.90
45.95 5.84
>150
>150
>150
>150
32.15 2.03
51.69 5.77
>150
>150
>150
112.96 7.10
63.33 3.51
54.25 3.06
43.22 6.51
>150
>150
>150
127.63 19.93
22.55 1.99
36.54 2.95
>150
100.96 11.35
62.61 3.86
53.88 3.89
43.32 4.97
>150
>150
>150
>150
14.05 1.82
24.88 1.37
>150
>150
>150
>150
>150
>150
92.17 9.42
63.73 6.77
67.75 2.38
38.75 4.92
>150
133.63 9.81
>150
>150
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
>150
>150
>150
>150
>150
134.17 9.77
114.55 16.65
105.05 13.45
79.37 4.26
66.24 2.59
46.27 6.14
134.74 7.79
>150
>150
>150
>150
>150
>150
94.88 9.17
65.78 7.76
71.37 3.95
46.69 2.03
>150
>150
>150
>150
>150
>150
>150
>150
⁄Results are presented as GI50 after a continuous exposure of 48 h and represent means SEM from at least three independent experiments performed in duplicate.
Doxorubicin was used as positive control, GI50: A375-C5 = 136.20 12.13 nM; MCF-7 = 59.77 3.81 nM; NCI-H460 = 34.07 1.95 nM.

C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
1055
of GI50 = 112.94 8.38
A375-C5, while the CDXs with a methyl group (11–14) were inac-
tive (GI50 >150 M). Thus, it can be inferred that the presence of
aryl or bulky alkyl groups in the chiral moiety may be important
for the activity.
l
M
and GI50 = 62.72 0.03
l
M
for
series FTIR (software: WinFirst v.2.10) spectrophotometer in KBr
microplates (cmꢀ1). 1H and 13C NMR spectra were taken in CDCl3
or DMSO-d6 at room temperature on Bruker Avance 300 instru-
ment (300.13 or 500.13 MHz for 1H and 75.47 or 125.77 MHz for
13C). Chemical shifts are expressed in d (ppm) values relative to
tetramethylsilane (TMS) as an internal reference. Coupling con-
stants are reported in hertz (Hz). 13C NMR assignments were made
by 2D HSQC and HMBC experiments (long-range C, H coupling
constants were optimized to 7 and 1 Hz). MS spectra were re-
corded as EI (electronic impact) mode on a VG Autospec Q spec-
trometer (m/z) and HRMS mass spectra were measured on a
Bruker Daltonics micrOTOF Mass Spectrometer, recorded as ESI
(electrospray) mode in Centro de Apoio Científico e Tecnolóxico á
Investigation (C.A.C.T.I.), University of Vigo, Spain. TLC was per-
formed using Merck silica gel 60 (GF254) plates, with appropriate
mobile phases and detection at 254 and/or 365 nm. Column
chromatography was carried out using Merck silica gel 60
(0.040–0.063 mm). Optical rotation measurements were carried
out on a Polartronic Universal polarimeter. Liquid chromatography
was performed using a HPLC system consisted of two Shimadzu LC
10-ADvp pumps, a FCV-10AL solvent selector valve, an automatic
l
Furthermore, comparing the results obtained with the CDXs
15–16 and the CDXs 31–32, possessing the same chiral moiety while
the xanthonic chemical substrates is the carboxyxanthone 1 and
carboxymethoxyxanthone 2, respectively, it can be seen that: the
first pair of compounds exhibited the highest growth inhibitory
effects on the three human tumor cell lines, while the second pair
was inactive. Accordingly, it is interesting to point out that the nat-
ure of the substituents and their positions on the xanthonic scaffold
also have an important role in growth inhibitory effect.
Regarding the influence of the enantiomeric configuration on the
activity, enantioselectivity between the enantiomeric pairs of CDXs
7–8 and 15–16, on the three human tumor cell lines, were observed.
For example, the CDX 7 presented a significant (p < 0.01) weaker
activity for all cell lines tested (GI50 = 112.94 8.38
GI50 = 112.96 7.10 M and GI50 = 100.96 11.35 M for A375-C5,
MCF-7 and NCI-H460 respectively), while its enantiomer ( CDX 8)
was more active presenting values of GI50 = 69.72 0.03 M,
GI50 = 63.33 3.51 M and GI50 = 62.61 3.86 M for A375-C5,
MCF-7 and NCI-H460, respectively.
lM,
l
l
l
injector SIL10-Advp, a SPD-10AV UV/VIS detector with a
l
l
SCL-10Avp interface or a HPLC system consisted of two Shimadzu
LC 10-AD pumps, an automatic injector SIL10-AF, a SPD-10A
UV/VIS detector with a CBM-10A interface or a System 880-PU
Intelligent HPLC Pump (Jasco) equipped with a Rheodyne 7125
Another interesting example of enantioselectivity was observed
between the enantiomers 3 and 4 in MCF-7 and NCI-H460 human
tumor cell lines. In fact, considering NCI-H460 cell line, CDX 3 pre-
injector fitted with a 20 lL loop, a 875-UV Intelligent UV/VIS
sented a slight inhibitory activity (GI50 = 85.88 5.30
CDX 4 was able to inhibit the growth of this cell line with a
GI50 = 42.62 1.77 M; in MCF-7 cell line, CDX 3 was considered
not active (GI50 >150 M), while CDX 4 presented a weaker activity
(GI50 = 91.91 6.27 M) (p < 0.01). Considering these results as
well as the incapacity of both enantiomers to inhibit the growth
of the human tumor cell line A375-C5, it can be emphasized that
cell-type selectivity also occurred.
l
M), while
Detector, and a Chromatography Station for Windows, version
1.7 DLL. Chirobiotic T column (15 ꢁ 0.46 cm ID size column) was
commercially available from Sigma-Aldrich. Polysaccharide
columns were prepared as described elsewhere47,48 and consisted
of amylose tris-3,5-dimethylphenylcarbamate and cellulose tris-
3,5-dimethylphenylcarbamate coated onto APS-Nucleosil (500 Å,
l
l
l
7
l
m, 20% w/w), and packed into a stainless-steel 15 ꢁ 0.46 cm
ID size column. (S,S)-Whelk-O1 column (15 ꢁ 0.46 cm ID size
column) was commercially available from Regis Technologies,
Inc. Both enantiomers of the optically pure reagents, valinol,
3. Conclusions
leucinol, 2-amino-1-propanol, 1-amino-2-propanol,
cinol, 2-amino-1,2-diphenylethanol, alanine tert-butyl ester and
-dimethylbenzylamine, were obtained from Fluka or Sigma–
a-phenylgly-
Coupling reactions of carboxyxanthone derivatives with a vari-
ety of chiral building blocks, such as amines, amino alcohols, and
amino esters demonstrated to be very efficient under very mild
conditions using TBTU. Six new and twenty four previously de-
scribed chiral derivatives of xanthones were synthesized with
excellent yields, very high enantiomeric excess and requiring short
reaction times. The present methodology has also the advantages
of operational simplicity, use of ready available, inexpensive and
low toxic materials, and simple reaction work-up. The MW-as-
sisted organic synthesis was successfully applied in the alkaline
cyclization of a benzophenone intermediate, with an increased
yield and a remarkable shorter reaction time, when compared with
the conventional heating conditions.
a
Aldrich. Other reagents and solvents were commercially available
materials at pro analysis or HPLC grade, from Sigma–Aldrich, Merck
or Fluka, and used without further purification. MW reactions were
performed using an Ethos MicroSYNTH 1600 Microwave Labstation
from Milestone (ThermoUnicam, Portugal).
4.2. Synthesis of the carboxyxanthone 6-methoxy-9-oxo-9H-
xanthene-2-carboxylic acid (1)
Compound 1 was obtained via Ullmann reaction as described
elsewhere.28
Some CDXs exhibited interesting dose-dependent growth inhib-
itory effects on the evaluated human tumor cell lines. Furthermore,
it can be inferred that the effects on the growth of the human tu-
mor cell lines can be ascribed not only to the nature and positions
of substituents on the xanthonic scaffold, but also are associated
with the stereochemistry of the CDXs concerning enantioselectivi-
ty in some cases.
4.3. Synthesis of the carboxymethoxyxanthone 2-((9-oxo-9H-
xanthen-3-yl)oxy)acetic acid (2) and intermediates 35–38
The synthetic route used to synthesize compound 2 is outlined
in Scheme 1, which involved a multi-step pathway.
4.3.1. (2-Hydroxy-4-methoxyphenyl)(2-methoxyphenyl)metha
none (35)
4. Experimental
4.1. General
Compound 35 was synthesized (9.42 g, 62%) and characterized
according to the described procedure.29
4.3.2. 3-Methoxy-9H-xanthen-9-one (36)
A mixture of compound 35 (10.00 g, 38.76 mmol) and NaOH
(18.80 g, 0.47 mol) was placed into a 270 mL Teflon reactor,
Melting points are uncorrected and were obtained in a Köfler
microscope. IR spectra were recorded on an ATIMattson Genesis

1056
C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
provided with
a
fiber-optic probe sensor, and MeOH/water
4.4. General procedure for the synthesis of chiral derivatives of
xanthones 3–32
(150 mL, 2:1 v:v) were added. The mixture was submitted to
30 min of microwave (MW) irradiation at 300 W of potency. After
cooling, the solid was filtered and washed with water (100 mL).
The crude product was recrystallized from MeOH to afford the
compound 36 as a white solid (5.69 g, 65%). Compound 36 was
characterized according to the described procedure.30
6-Methoxy-9-oxo-9H-xanthene-2-carboxylic acid (1) or 2-((9-
oxo-9H-xanthen-3-yl)oxy)acetic acid (2) (100 mg, 0.37 mmol)
was dissolved in anhydrous THF (20 mL) and TEA (103 lL,
0.74 mmol) was added. Following, TBTU (120 mg, 0.37 mmol)
and an appropriate chiral reagent (0.37 mmol) were added. The
mixture was stirred at room temperature for 0.5 to 5 h. After com-
pletion of the reaction, the solvent was evaporated under reduced
pressure and the crude product was dissolved in 50 mL of dichlo-
romethane (3–10, 13–14, 17–30), ethyl acetate (11–12) or chloro-
form (15–16, 31–32). This solution was washed with 1 M HCl
solution (2 ꢁ 25 mL), saturated solution of NaHCO3 (2 ꢁ 30 mL),
and water (3 ꢁ 50 mL). The organic layer was dried with anhy-
drous sodium sulfate, filtered and the solvent was evaporated un-
der reduced pressure. The product was recrystallized from ethyl
acetate/n-hexane (3–4, 7–14, 19–20, 23–30), EtOH (5–6, 21–22),
chloroform/n-hexane (15–16, 31–32), or MeOH/water (17–18) to
afford the chiral derivative of xanthone. Compounds 3–32 were
identified by their spectroscopic and analytical data.
4.3.3. 3-Hydroxy-9H-xanthen-9-one (37)
Compound 37 was synthesized (7.13 g, 76%)49 and character-
ized30 according to the described procedure.
4.3.4. Methyl 2-((9-oxo-9H-xanthen-3-yl)oxy)acetate (38)
Compound 37 (10.00 g, 47.17 mmol) was dissolved in anhy-
drous acetone (400 mL) and K2CO3 (7.65 g, 55.43 mmol) and
BrCH2COOCH3 (5 mL, 54.38 mmol) were added. The mixture
was kept under reflux and magnetic stirring for 24 h. Then, the
mixture was concentrated under reduced pressure and n-hexane
was added. The solid was separated by filtration and purified by
column chromatography (silica gel, dichloromethane/ethyl ace-
tate in gradient) to afford compound 38 as
a white solid
~
(9.11 g, 68%); mp: 156–159 °C; IR (KBr):
m
1737, 1617, 1463,
1433, 1378, 1291, 1244, 821, 760 cmꢀ1
;
1H NMR (300.13 MHz,
4.4.1. (R)-6-Methoxy-9-oxo-N-(1-(p-tolyl)ethyl)-9H-xanthene-2-
[D6]DMSO): d = 8.19 (1H, dd, J = 7.4 and 0.9 Hz, H-8), 8.13 (1H,
d, J = 9.0 Hz, H-1), 7.87 (1H, ddd, J = 8.0, 7.8 and 1.2 Hz, H-6),
7.65 (1H, d, J = 8.1 Hz, H-5), 7.50 (1H, ddd, J = 7.5, 7.3 and
0.9 Hz, H-7), 7.21 (1H, d, J = 2.4 Hz, H-4), 7.12 (1H, dd, J = 8.8
and 2.4 Hz, H-2), 5.06 (2H, s, OCH2), 3.75 ppm (3H, s, OCH3);
13C NMR (75.47 MHz, [D6]DMSO): d = 175.0 (C-9), 168.6 (C@O, es-
ter), 163.2 (C-3), 157.3 (C-4a), 155.6 (C-10a), 135.2 (C-6), 127.7
(C-1), 125.9 (C-8), 124.4 (C-7), 121.2 (C-8a), 118.0 (C-5), 115.5
(C-9a), 113.8 (C-2), 101.6 (C-4), 65.0 (OCH2), 52.0 ppm (OCH3);
MS (ESI) m/z (%): 286 [M+H]++1 (20), 285 [M+H]+ (100), 201
(15), 175 (12), 158 (73), 130 (18); HRMS (ESI) m/z [M+H]+ calcd
for C16H12O5: 285.07575, found: 285.07629.
carboxamide (3)
Compound 3 was obtained as a white soꢃlid (140 mg, 98%); mp:
221–223 °C (ethyl acetate/n-hexane); ½a D25
ꢂ
ꢀ125.3 (c = 10.70 ꢁ
C
10ꢀ3 g/mL in dichloromethane); IR (KBr):
3336, 1660, 1611,
~
m
1531, 1477, 1440, 1269, 836 cmꢀ1 1H NMR (300.13 MHz, CDCl3):
;
d = 8.56 (1H, d, J = 2.3 Hz, H-1), 8.32 (1H, dd, J = 8.7 and 2.3 Hz,
H-3), 8.25 (1H, d, J = 8.9 Hz, H-8), 7.53 (1H, d, J = 8.7 Hz, H-4),
7.31 (2H, d, J = 8.0 Hz, H-200 and H-600), 7.18 (2H, d, J = 8.0 Hz,
H-300 and H-500), 6.99 (1H, dd, J = 8.9 and 2.3 Hz, H-7), 6.92 (1H, d,
J = 2.3 Hz, H-5), 6.59 (1H, d, J = 7.4 Hz, NH), 5.32 (1H, m, H-10),
3.95 (3H, s, Ar-OCH3), 2.34 (3H, s, Ar-CH3), 1.63 ppm (3H, d,
J = 6.9 Hz, H-20); 13C NMR (75.47 MHz, CDCl3): d = 175.9 (C-9),
165.5 (C@O, amide), 164.8 (C-6), 158.0 (C-10a), 157.9 (C-4a),
139.9 (C-100), 137.2 (C-400), 134.3 (C-3), 130.1 (C-2), 129.4 (C-300
and C-500), 128.3 (C-8), 126.2 (C-200 and C-600), 124.0 (C-1), 121.0
(C-9a), 118.5 (C-4), 115.5 (C-8a), 113.8 (C-7), 100.3 (C-5), 55.9
(Ar-OCH3), 49.3 (C-10), 21.7 (Ar-CH3), 21.1 ppm (C-20); MS (ESI)
m/z (%): 389 [M+H]++1 (29), 388 [M+H]+ (100), 354 (23), 326
(38), 270 (9), 219 (17), 197 (6); HRMS (ESI) m/z [M+H]+ calcd for
C24H21NO4: 388.15433, found: 388.15343; ee >99% (HPLC;
Column: amylose tris-3,5-dimethylphenylcarbamate coated onto
4.3.5. 2-((9-Oxo-9H-xanthen-3-yl)oxy)acetic acid (2)
Compound 38 (9.40 g, 33.07 mmol) was dissolved in dichlo-
romethane/MeOH (800 mL, 1:1 v/v) and 5 M NaOH solution
(64 mL) was added. The mixture was kept at room temperature
and magnetic stirring for 5 h. After this period, the dichloro-
methane and MeOH were evaporated under reduced pressure
and the suspension was washed with dichloromethane
(2 ꢁ 100 mL). The aqueous phase was acidified with 5 M HCl
solution resulting in the formation of a precipitate that was col-
lected by filtration under reduced pressure and dissolved in sat-
urated NaHCO3 solution. This solution was washed again with
dichloromethane (2 ꢁ 100 mL) and the aqueous phase was acid-
ified with 5 M HCl solution. The precipitate formed was col-
lected by filtration under reduced pressure, washed with
water, and recrystallized from MeOH to provide compound 2
as a white solid (6.96 g, 78%); mp: 211–213 °C (MeOH); IR
~
APS-Nucleosil (500 Å, 7 lm, 20% w/w), Mobile phase: EtOH/ACN
(50:50 v:v), 0.5 mL/min, kmax 254 nm).
4.4.2. (S)-6-Methoxy-9-oxo-N-(1-(p-tolyl)ethyl)-9H-xanthene-2-
carboxamide (4)
Compound 4 was synthesized (139 mg, 97%) and characterized
according to the described procedure.28
(KBr):
m
2858, 1712, 1609, 1461, 1431, 1374, 1231, 852,
4.4.3. (R)-N-(1-Hydroxy-3-methylbutan-2-yl)-6-methoxy-9-oxo-
9H-xanthene-2-carboxamide (5)
751 cmꢀ1
;
1H NMR (300.13 MHz, [D6]DMSO): d = 13.32 (1H, s,
COOH), 8.19 (1H, dd, J = 7.4 and 0.9 Hz, H-8), 8.13 (1H, d,
J = 9.0 Hz, H-1), 7.87 (1H, ddd, J = 8.2, 7.8 and 1.2 Hz, H-6),
7.65 (1H, d, J = 8.4 Hz, H-5), 7.49 (1H, ddd, J = 7.5, 7.3 and
0.9 Hz, H-7), 7.16 (1H, d, J = 2.4 Hz, H-4), 7.11 (1H, dd, J = 8.8
and 2.4 Hz, H-2), 4.94 ppm (2H, s, OCH2); 13C NMR (75.47 MHz,
[D6]DMSO): d = 175.0 (C-9), 169.5 (COOH), 163.4 (C-3), 157.3
(C-4a), 155.6 (C-10a), 135.2 (C-6), 127.7 (C-1), 125.9 (C-8),
124.4 (C-7), 121.2 (C-8a), 118.0 (C-5), 115.3 (C-9a), 113.8
(C-2), 101.5 (C-4), 65.0 ppm (OCH2); HRMS (ESI) m/z [M+H]+
calcd for C15H10O5: 271.06010, found: 271.06006.
Compound 5 was obtained as a white solid (126 mg, 96%); mp:
ꢃC
180–182 °C (EtOH); ½a D25
ꢂ
+14.8 (c = 10.08 ꢁ 10ꢀ3 g/mL in dichlo-
3370, 3241, 1655, 1619, 1539, 1474,
1H NMR (300.13 MHz, [D6]DMSO):
~
romethane); IR (KBr):
m
1444, 1270, 828 cmꢀ1
;
d = 8.73 (1H, d, J = 2.3 Hz, H-1), 8.38 (1H, d, J = 8.7 Hz, NH), 8.31
(1H, dd, J = 8.7 and 2.3 Hz, H-3), 8.15 (1H, d, J = 8.9 Hz, H-8), 7.71
(1H, d, J = 8.7 Hz, H-4), 7.21 (1H, d, J = 2.3 Hz, H-5), 7.10 (1H, dd,
J = 8.9 and 2.3 Hz, H-7), 4.65 (1H, t, J = 5.6 Hz, OH), 3.95 (3H, s,
Ar-OCH3), 3.85 (1H, m, H-10), 3.56 (2H, m, CH2OH), 1.97 (1H, m,
H-20), 0.93 (3H, d, J = 7.0 Hz, H-30a)⁄, 0.91 ppm (3H, d, J = 7.0 Hz,

C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
1057
H-30b)⁄; 13C NMR (75.47 MHz, [D6]DMSO): d = 175.0 (C-9), 165.2
(C@O, amide), 165.1 (C-6), 157.6 (C-10a), 157.1 (C-4a), 134.1
(C-3), 130.9 (C-2), 127.8 (C-8), 125.4 (C-1), 120.7 (C-9a), 118.0
(C-4), 115.0 (C-8a), 114.0 (C-7), 100.8 (C-5), 61.3 (CH2OH), 57.0
(Ar-OCH3), 56.3 (C-10), 28.7 (C-20), 19.7 (C-30a)⁄, 18.8 ppm
(C-30b)⁄; MS (ESI) m/z (%): 357 [M+H]++1 (28), 356 [M+H]+ (100),
326 (27), 255 (70), 233 (36), 197 (3), 155 (10); HRMS (ESI) m/z
[M+H]+ calcd for C20H21NO5: 356.14925, found: 356.14967; ee
>99% (HPLC; Column: Chirobiotic T, Mobile phase: n-hexane/EtOH
(80:20 v:v), 0.5 mL/min, kmax 254 nm).
56.2 ppm (Ar-OCH3); MS (ESI) m/z (%): 391 [M+H]++1 (28), 390
[M+H]+ (100), 354 (3), 255 (12), 197 (5), 179 (8); HRMS (ESI) m/z
[M+H]+ calcd for C23H19NO5: 390.13360, found: 390.13209; ee
>99% (HPLC; Column: amylose tris-3,5-dimethylphenylcarbamate
coated onto APS-Nucleosil (500 Å, 7 lm, 20% w/w), Mobile phase:
EtOH/ACN (50:50 v:v), 0.5 mL/min, kmax 254 nm).
4.4.8. (S)-N-(2-Hydroxy-1-phenylethyl)-6-methoxy-9-oxo-9H-
xanthene-2-carboxamide (10)
Compound 10 was obtained as a white solid (138 mg, 96%); mp:
ꢃC
⁄Asterisks denote assignments that may be interchanged.
94–96 °C (ethyl acetate/n-hexane); ½a D25
ꢂ
ꢀ76.0 (c = 2.70 ꢁ 10ꢀ3 g/mL
~
m
in dichloromethane); IR (KBr):
3395, 3284, 1633, 1618, 1537, 1475,
4.4.4. (S)-N-(1-Hydroxy-3-methylbutan-2-yl)-6-methoxy-9-oxo-
9H-xanthene-2-carboxamide (6)
1440, 1266, 830 cmꢀ1 1H NMR (300.13 MHz, [D6]DMSO): d = 9.11
;
(1H, d, J = 8.0 Hz, NH), 8.81 (1H, d, J = 2.2 Hz, H-1), 8.32 (1H, dd,
J = 8.8 and 2.2 Hz, H-3), 8.15 (1H, d, J = 8.9 Hz, H-8), 7.72 (1H, d,
J = 8.8 Hz, H-4), 7.42 (2H, d, J = 7.1 Hz, H-200 and H-600), 7.33 (2H, t,
J = 7.1 Hz, H-300 and H-500), 7.25 (1H, d, J = 7.1 Hz, H-400), 7.21 (1H, d,
J = 2.3 Hz, H-5), 7.10 (1H, dd, J = 8.9 and 2.3 Hz, H-7), 5.11 (1H, m,
H-10), 5.01 (1H, t, J = 5.8 Hz, OH), 3.95 (3H, s, Ar-OCH3), 3.70 ppm
(2H, m, CH2OH); 13C NMR (75.47 MHz, [D6]DMSO): d = 174.9 (C-9),
165.2 (C@O, amide), 164.8 (C-6), 157.5 (C-10a), 157.2 (C-4a), 141.3
(C-100), 134.1 (C-3), 130.4 (C-2), 128.1 (C-300 and C-500), 127.7 (C-8),
127.0 (C-200 and C-600), 127.0 (C-400), 125.4 (C-1), 120.7 (C-9a), 118.1
(C-4), 115.0 (C-8a), 114.0 (C-7), 100.8 (C-5), 64.4 (CH2OH), 56.3
(C-10), 56.2 ppm (Ar-OCH3); MS (ESI) m/z (%): 391 [M+H]++1 (26),
390 [M+H]+ (100), 348 (18), 255 (57), 197 (14), 191 (49), 179 (22);
HRMS (ESI) m/z [M+H]+ calcd for C23H19NO5: 390.13360, found:
390.13331; ee >99% (HPLC; Column: amylose tris-3,5-dimethylphe-
Compound 6 was synthesized (129 mg, 98%) and characterized
according to the described procedure.28
4.4.5. (R)-N-(1-Hydroxy-4-methylpentan-2-yl)-6-methoxy-9-
oxo-9H-xanthene-2-carboxamide (7)
Compound 7 was obtained as a white solid (134 mg, 98%); mp:
ꢃC
156–158 °C (ethyl acetate/n-hexane);
½
a 2D5
ꢂ
+25.0 (c = 11.40
3360, 3283, 1651,
ꢁ 10ꢀ3 g/mL in dichloromethane); IR (KBr):
m
~
1617, 1546, 1468, 1443, 1261, 801 cmꢀ1 1H NMR (300.13 MHz,
;
[D6]DMSO): d = 8.71 (1H, d, J = 2.1 Hz, H-1), 8.40 (1H, d, J = 8.6 Hz,
NH), 8.30 (1H, dd, J = 8.7 and 2.2 Hz, H-3), 8.14 (1H, d, J = 8.9 Hz,
H-8), 7.70 (1H, d, J = 8.7 Hz, H-4), 7.21 (1H, d, J = 2.2 Hz, H-5),
7.10 (1H, dd, J = 8.9 and 2.2 Hz, H-7), 4.76 (1H, t, J = 5.7 Hz, OH),
4.12 (1H, m, H-10), 3.95 (3H, s, Ar-OCH3), 3.43 (2H, m, CH2OH),
1.63 (⁄2H, m, H-20), 1.45 (1H, m, H-30), 0.91 (3H, d, J = 6.0 Hz,
H-40a) , 0.90 ppm (3H, d, J = 6.0 Hz, H-40b)⁄; 13C NMR (75.47 MHz,
[D6]DMSO): d = 174.9 (C-9), 165.2 (C@O, amide), 164.7 (C-6),
157.6 (C-10a), 157.1 (C-4a), 134.0 (C-3), 130.7 (C-2), 127.8 (C-8),
125.3 (C-1), 120.7 (C-9a), 118.1 (C-4), 115.0 (C-8a), 114.0 (C-7),
100.8 (C-5), 63.9 (CH2OH), 56.3 (Ar-OCH3), 49.8 (C-10), 39.0 (C-20),
24.5 (C-30), 23.5 (C-40a)⁄, 21.9 ppm (C-40b)⁄; MS (ESI) m/z (%): 371
[M+H]++1 (28), 370 [M+H]+ (100), 328 (8), 306 (8), 219 (4), 197
(4), 171 (18); HRMS (ESI) m/z [M+H]+ calcd for C21H23NO5:
370.16490, found: 370.16552; ee >99% (HPLC; Column: amylose
tris-3,5-dimethylphenylcarbamate coated onto APS-Nucleosil
nylcarbamate coated onto APS-Nucleosil (500 Å, 7 lm, 20% w/w),
Mobile phase: EtOH/ACN (50:50 v:v), 0.5 mL/min, kmax 254 nm).
4.4.9. (R)-N-(1-Hydroxypropan-2-yl)-6-methoxy-9-oxo-9H-
xanthene-2-carboxamide (11)
Compound 11 was obtained as a white solid (117 mg, 97%); mp:
ꢃC
233–235 °C (ethyl acetate/n-hexane);
½
a 2D5
ꢂ
ꢀ83.3 (c = 0.72
ꢁ 10ꢀ6 g/mL in acetone); IR (KBr):
m
3380, 3312, 1657, 1617,
~
1551, 1479, 1442, 1265, 821 cmꢀ1
;
1H NMR (300.13 MHz,
[D6]DMSO): d = 8.71 (1H, d, J = 2.1 Hz, H-1), 8.49 (1H, d, J = 8.1 Hz,
NH), 8.30 (1H, dd, J = 8.8 and 2.1 Hz, H-3), 8.14 (1H, d, J = 8.9 Hz,
H-8), 7.71 (1H, d, J = 8.8 Hz, H-4), 7.21 (1H, d, J = 2.2 Hz, H-5),
7.10 (1H, dd, J = 8.9 and 2.2 Hz, H-7), 4.79 (1H, t, J = 5.7 Hz, OH),
4.03 (2H, m, CH2OH), 3.95 (3H, s, Ar-OCH3), 3.49 (1H, m, H-10),
1.16 ppm (3H, d, J = 6.7 Hz, H-20); 13C NMR (75.47 MHz,
[D6]DMSO): d = 174.9 (C-9), 165.2 (C@O, amide), 164.4 (C-6),
157.5 (C-10a), 157.1 (C-4a), 134.0 (C-3), 130.6 (C-2), 127.7 (C-8),
125.3 (C-1), 120.6 (C-9a), 118.0 (C-4), 114.9 (C-8a), 114.0 (C-7),
100.8 (C-5), 64.3 (C-10), 56.3 (Ar-OCH3), 47.5 (CH2OH), 17.0 ppm
(C-20); MS (ESI) m/z (%): 329 [M+H]++1 (18), 328 [M+H]+ (100),
282 (17), 218 (13); HRMS (ESI) m/z [M+H]+ calcd for C18H17NO5:
328.11795, found: 328.11798; ee >99% (HPLC; Column: amylose
tris-3,5-dimethylphenylcarbamate coated onto APS-Nucleosil
(500 Å, 7 lm, 20% w/w), Mobile phase: EtOH/ACN (50:50 v:v),
0.5 mL/min, kmax 254 nm).
⁄Asterisks denote assignments that may be interchanged.
4.4.6. (S)-N-(1-Hydroxy-4-methylpentan-2-yl)-6-methoxy-9-
oxo-9H-xanthene-2-carboxamide (8)
Compound 8 was synthesized (135 mg, 99%) and characterized
according to the described procedure.28
4.4.7. (R)-N-(2-Hydroxy-1-phenylethyl)-6-methoxy-9-oxo-9H-
xanthene-2-carboxamide (9)
Compound 9 was obtained as a white solid (137 mg, 95%); mp:
ꢃC
94–96 °C (ethyl acetate/n-hexane); ½a D25
ꢂ
+75.6 (c = 2.70 ꢁ 10ꢀ3 g/mL
(500 Å, 7 lm, 20% w/w), Mobile phase: EtOH/ACN (50:50 v:v),
0.5 mL/min, kmax 254 nm).
~
m
in dichloromethane); IR (KBr):
3395, 3255, 1633, 1618, 1544,
1473, 1430, 1258, 816 cmꢀ1 1H NMR (300.13 MHz, [D6]DMSO):
;
d = 9.12 (1H, d, J = 8.0 Hz, NH), 8.80 (1H, d, J = 2.2 Hz, H-1), 8.32
(1H, dd, J = 8.8 and 2.2 Hz, H-3), 8.15 (1H, d, J = 8.9 Hz, H-8), 7.72
(1H, d, J = 8.8 Hz, H-4), 7.42 (2H, d, J = 7.2 Hz, H-200 and H-600),
7.33 (2H, t, J = 7.2 Hz, H-300 and H-500), 7.25 (1H, d, J = 7.2 Hz,
H-400), 7.21 (1H, d, J = 2.3 Hz, H-5), 7.09 (1H, dd, J = 8.9 and
2.3 Hz, H-7), 5.11 (1H, m, H-10), 5.03 (1H, t, J = 5.8 Hz, OH), 3.95
(3H, s, Ar-OCH3), 3.70 ppm (2H, m, CH2OH); 13C NMR
(75.47 MHz, [D6]DMSO): d = 174.9 (C-9), 165.3 (C@O, amide),
164.8 (C-6), 157.6 (C-10a), 157.2 (C-4a), 141.3 (C-100), 134.1 (C-3),
130.4 (C-2), 128.2 (C-300 and C-500), 127.8 (C-8), 127.1 (C-200 and
C-600), 127.0 (C-400), 125.5 (C-1), 120.7 (C-9a), 118.2 (C-4), 115.0
(C-8a), 114.0 (C-7), 100.8 (C-5), 64.5 (CH2OH), 56.3 (C-10),
4.4.10. (S)-N-(1-Hydroxypropan-2-yl)-6-methoxy-9-oxo-9H-
xanthene-2-carboxamide (12)
Compound 12 was obtained as a white solid (116 mg, 96%); mp:
ꢃC
233–235 °C (ethyl acetate/n-hexane);
½
a 2D5
ꢂ
+83.3 (c = 0.72
ꢁ 10ꢀ6 g/mL in acetone); IR (KBr):
m
3387, 3312, 1652, 1620,
~
1543, 1486, 1443, 1261, 819 cmꢀ1
;
1H NMR (300.13 MHz,
[D6]DMSO): d = 8.72 (1H, d, J = 2.2 Hz, H-1), 8.49 (1H, d, J = 8.0 Hz,
NH), 8.30 (1H, dd, J = 8.8 and 2.2 Hz, H-3), 8.14 (1H, d, J = 8.9 Hz,
H-8), 7.70 (1H, d, J = 8.7 Hz, H-4), 7.21 (1H, d, J = 2.2 Hz, H-5),
7.10 (1H, dd, J = 8.9 and 2.2 Hz, H-7), 4.77 (1H, t, J = 5.7 Hz, OH),
4.03 (2H, m, CH2OH), 3.95 (3H, s, Ar-OCH3), 3.49 (1H, m, H-10),
1.16 ppm (3H, d, J = 6.7 Hz, H-20); 13C NMR (75.47 MHz,

1058
C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
[D6]DMSO): d = 174.9 (C-9), 165.2 (C@O, amide), 164.4 (C-6), 157.5
(C-10a), 157.1 (C-4a), 134.0 (C-3), 130.6 (C-2), 127.7 (C-8), 125.3
(C-1), 120.6 (C-9a), 118.0 (C-4), 114.9 (C-8a), 114.0 (C-7), 100.8
(C-5), 64.3 (C-10), 56.3 (Ar-OCH3), 47.5 (CH2OH), 17.0 ppm (C-20);
MS (ESI) m/z (%): 329 [M+H]++1 (25), 328 [M+H]+ (100), 255 (79),
233 (45), 197 (4), 155 (12); HRMS (ESI) m/z [M+H]+ calcd for
H-1), 8.13 (1H, d, J = 8.8 Hz, H-8), 8.07 (1H, dd, J = 8.8 and 2.0 Hz,
H-3), 7.64 (1H, d, J = 8.8 Hz, H-4), 7.48 (2H, d, J = 7.0 Hz, H-200 and
H-600), 7.46 (2H, d, J = 7.0 Hz, H-2000 and H-6000), 7.31 (2H, t,
J = 7.0 Hz, H-300 and H-500), 7.28 (2H, t, J = 7.0 Hz, H-3000 and H-5000),
7.24 (1H, d, J = 7.0 Hz, H-400), 7.19 (1H, d, J = 7.0 Hz, H-4000), 7.19
(1H, d, J = 2.3 Hz, H-5), 7.09 (1H, dd, J = 8.8 and 2.3 Hz, H-7), 5.48
(1H, d, J = 4.9 Hz, OH), 5.17 (1H, dd, J = 8.9 and 8.7 Hz, H-10), 4.97
(1H, dd, J = 8.7 and 4.9 Hz CHOH) 3.94 ppm (3H, s, Ar-OCH3); 13C
NMR (75.47 MHz, [D6]DMSO): d = 174.7 (C-9), 165.2 (C@O, amide),
163.9 (C-6), 157.5 (C-10a), 157.1 (C-4a), 143.6 (C-100), 141.3 (C-1000),
133.8 (C-3), 130.4 (C-2), 128.4 (C-300 and C-500), 128.4 (C-3000 and
C-5000),127.6 (C-8), 127.6 (C-200 and C-600), 127.1 (C-2000 and C-6000),
126.9 (C-400), 126.7 (C-4000),125.2 (C-1), 120.6 (C-9a), 118.1 (C-4),
114.9 (C-8a), 114.0 (C-7), 100.8 (C-5), 74.5 (CHOH), 59.2 (C-10),
56.2 ppm (Ar-OCH3); MS (ESI) m/z (%): 467 [M+H]++1 (31), 466
[M+H]+ (100), 448 (49), 391 (7), 282 (40), 256 (11); HRMS (ESI)
m/z [M+H]+ calcd for C29H23NO5: 466.16490, found: 466.16486;
ee >97% (HPLC; Column: (S,S)-Whelk-O1, Mobile phase: ACN/
MeOH (50:50 v:v), 1.0 mL/min, kmax 254 nm).
C
18H17NO5: 328.11795, found: 328.11914; ee >99% (HPLC;
Column: amylose tris-3,5-dimethylphenylcarbamate coated onto
APS-Nucleosil (500 Å, 7 m, 20% w/w), Mobile phase: EtOH/ACN
l
(50:50 v:v), 0.5 mL/min, kmax 254 nm).
4.4.11. (R)-N-(2-Hydroxypropyl)-6-methoxy-9-oxo-9H-
xanthene-2-carboxamide (13)
Compound 13 was obtained as a white solid (116 mg, 96%); mp:
ꢃC
208–210 °C (ethyl acetate/n-hexane);
10ꢀ6 g/mL in dichloromethane); IR (KBr):
1612, 1535, 1473, 1446, 1278, 838 cmꢀ1
½
a 2D5
ꢂ
ꢀ81.0 (c = 0.74 ꢁ
~
m
3440, 3370, 1657,
;
1H NMR (300.13 MHz,
CDCl3): d = 8.62 (1H, d, J = 2.2 Hz, H-1), 8.30 (1H, dd, J = 8.8 and
2.2 Hz, H-3), 8.26 (1H, d, J = 8.9 Hz, H-8), 7.54 (1H, d, J = 8.8 Hz,
H-4), 6.99 (1H, dd, J = 8.9 and 2.4 Hz, H-7), 6.92 (1H, d, J = 2.4 Hz,
H-5), 6.87 (1H, s, NH), 4.09 (1H, m, H-20), 3.96 (3H, s, Ar-OCH3),
3.72 (1H, m, H-10)⁄, 3.37 (1H, m, H-10)⁄, 2.58 (1H, s, OH),
1.31 ppm (3H, d, J = 6.3 Hz, CH3); 13C NMR (125.77 MHz, CDCl3):
d = 175.8 (C-9), 167.0 (C@O, amide), 165.5 (C-6), 157.9 (C-10a),
157.0 (C-4a), 134.2 (C-3), 129.8 (C-2), 128.4 (C-8), 124.4 (C-1),
121.1 (C-9a), 118.6 (C-4), 115.5 (C-8a), 113.8 (C-7), 100.4 (C-5),
67.6 (C-20), 55.9 (Ar-OCH3), 47.6 (C-10), 21.2 ppm (CH3); MS (ESI)
m/z (%): 329 [M+H]++1 (19), 328 [M+H]+ (100), 282 (9), 218 (75);
HRMS (ESI) m/z [M+H]+ calcd for C18H17NO5: 328.11795, found:
328.11808; ee >99% (HPLC; Column: amylose tris-3,5-dimethylphe-
4.4.14. N-((1S,2R)-2-Hydroxy-1,2-diphenylethyl)-6-methoxy-9-
oxo-9H-xanthene-2-carboxamide (16)
Compound 16 was obtained as a white solid (163 mg, 95%); mp:
ꢃC
243–244 °C (chloroform/n-hexane);
ꢁ 10ꢀ3 g/mL in chloroform); IR (KBr):
1526, 1478, 1441, 1260, 834 cmꢀ1
½
a 2D5
ꢂ
+175.7 (c = 0.33
~
m
3444, 3340, 1636, 1618,
;
1H NMR (300.13 MHz,
[D6]DMSO): d = 9.00 (1H, d, J = 8.9 Hz, NH), 8.53 (1H, d, J = 2.1 Hz,
H-1), 8.13 (1H, d, J = 8.8 Hz, H-8), 8.07 (1H, dd, J = 8.8 and 2.1 Hz,
H-3), 7.64 (1H, d, J = 8.8 Hz, H-4), 7.48 (2H, d, J = 7.0 Hz, H-200 and
H-600), 7.46 (2H, d, J = 7.0 Hz, H-2000 and H-6000), 7.31 (2H, t,
J = 7.0 Hz, H-300 and H-500), 7.28 (2H, t, J = 7.0 Hz, H-3000 and H-5000),
7.24 (1H, d, J = 7.0 Hz, H-400), 7.19 (1H, d, J = 7.0 Hz, H-4000), 7.19
(1H, d, J = 2.3 Hz, H-5), 7.09 (1H, dd, J = 8.8 and 2.3 Hz, H-7), 5.48
(1H, d, J = 4.9 Hz, OH), 5.17 (1H, dd, J = 8.9 and 8.7 Hz, H-10), 4.97
(1H, dd, J = 8.7 and 4.9 Hz CHOH) 3.94 ppm (3H, s, Ar-OCH3); 13C
NMR (75.47 MHz, [D6]DMSO): d = 174.7 (C-9), 165.2 (C@O, amide),
163.9 (C-6), 157.5 (C-10a), 157.1 (C-4a), 143.6 (C-100), 141.3 (C-1000),
133.8 (C-3), 130.4 (C-2), 128.4 (C-300 and C-500), 128.4 (C-3000 and
C-5000),127.6 (C-8), 127.6 (C-200 and C-600), 127.1 (C-2000 and C-6000),
126.9 (C-400), 126.7 (C-4000),125.2 (C-1), 120.6 (C-9a), 118.1 (C-4),
114.9 (C-8a), 114.0 (C-7), 100.8 (C-5), 74.5 (CHOH), 59.2 (C-10),
56.2 ppm (Ar-OCH3); MS (ESI) m/z (%): 467 [M+H]++1 (30), 466
[M+H]+ (100), 448 (40), 391 (4), 282 (27), 256 (7); HRMS (ESI)
m/z [M+H]+ calcd for C29H23NO5: 466.16490, found: 466.16495;
ee >98% (HPLC; Column: (S,S)-Whelk-O1, Mobile phase:
ACN/MeOH (50:50 v:v), 1.0 mL/min, kmax 254 nm).
nylcarbamate coated onto APS-Nucleosil (500 Å, 7 lm, 20% w/w),
Mobile phase: n-hexane/2-PrOH (50:50 v:v), 0.5 mL/min, kmax
254 nm).
⁄Asterisks denote assignments that may be interchanged.
4.4.12. (S)-N-(2-Hydroxypropyl)-6-methoxy-9-oxo-9H-
xanthene-2-carboxamide (14)
Compound 14 was obtained as a white solid (117 mg, 97%); mp:
ꢃC
208–210 °C (ethyl acetate/n-hexane);
½
a 2D5
ꢂ
+81.2 (c = 0.74
ꢁ 10ꢀ6 g/mL in dichloromethane); IR (KBr):
m
3440, 3371, 1658,
~
1613, 1534, 1474, 1446, 1277, 837 cmꢀ1 1H NMR (300.13 MHz,
;
CDCl3): d = 8.62 (1H, d, J = 2.2 Hz, H-1), 8.29 (1H, dd, J = 8.8 and
2.2 Hz, H-3), 8.25 (1H, d, J = 8.9 Hz, H-8), 7.53 (1H, d, J = 8.8 Hz,
H-4), 6.99 (1H, dd, J = 8.9 and 2.3 Hz, H-7), 6.91 (1H, d, J = 2.3 Hz,
H-5), 6.87 (1H, s, NH), 4.09 (1H, m, H-20), 3.96 (3H, s, Ar-OCH3),
3.72 (1H, m, H-10)⁄, 3.37 (1H, m, H-10)⁄, 2.58 (1H, s, OH),
1.30 ppm (3H, d, J = 6.3 Hz, CH3); 13C NMR (75.47 MHz, CDCl3):
d = 174.9 (C-9), 165.3 (C@O, amide), 165.1 (C-6), 157.6 (C-10a),
157.2 (C-4a), 133.9 (C-3), 130.5 (C-2), 127.8 (C-8), 125.4 (C-1),
120.7 (C-9a), 118.2 (C-4), 115.0 (C-8a), 114.1 (C-7), 100.8 (C-5),
67.6 (C-20), 55.9 (Ar-OCH3), 47.6 (C-10), 21.1 ppm (CH3); MS (ESI)
m/z (%): 329 [M+H]++1 (19), 328 [M+H]+ (74), 255 (100), 233
(53), 197 (6), 155 (13); HRMS (ESI) m/z [M+H]+ calcd for
4.4.15. (R)-tert-Butyl 2-(6-methoxy-9-oxo-9H-xanthene-2-
carboxamido)propanoate (17)
Compound 17 was obtained as a white solid (141 mg, 96%); mp:
ꢃC
160–162 °C (MeOH/water); ½a D25
ꢂ
ꢀ29.8 (c = 0.67 ꢁ 10ꢀ3 g/mL in
~
dichloromethane); IR (KBr):
m
3389, 1735, 1645, 1622, 1525,
1480, 1458, 826 cmꢀ1 1H NMR (300.13 MHz, CDCl3): d = 8.67
;
C
18H17NO5: 328.11795, found: 328.11732; ee >99% (HPLC; Col-
umn: amylose tris-3,5-dimethylphenylcarbamate coated onto
APS-Nucleosil (500 Å, 7 m, 20% w/w), Mobile phase: n-hexane/
(1H, d, J = 2.3 Hz, H-1), 8.28 (1H, dd, J = 8.8 and 2.3 Hz, H-3), 8.27
(1H, d, J = 8.8 Hz, H-8), 7.54 (1H, d, J = 8.8 Hz, H-4), 6.99 (1H, dd,
J = 8.8 and 2.4 Hz, H-7), 6.92 (1H, d, J = 2.4 Hz, H-5), 6.91 (1H, d,
J = 7.2 Hz, NH), 4.70 (1H, quint, J = 7.2 Hz, H-10), 3.96 (3H, s,
Ar-OCH3), 1.53 (3H, d, J = 7.2 Hz, H-20), 1.51 ppm (9H, s,
(C(CH3)3)); 13C NMR (125.77 MHz, CDCl3): d = 175.7 (C-9), 172.2
(C@O, ester), 165.4 (C@O, amide), 165.2 (C-6), 158.0 (C-10a),
157.9 (C-4a), 133.9 (C-3), 129.8 (C-2), 128.4 (C-8), 124.8 (C-1),
121.3 (C-9a), 118.5 (C-4), 115.6 (C-8a), 113.7 (C-7), 100.4 (C-5),
82.3 (C-10), 55.9 (Ar-OCH3), 49.2 (C(CH3)3), 29.7 (C-20), 28.0 ppm
((CH3)3); MS (ESI) m/z (%): 399 [M+H]++1 (27), 398 [M+H]+ (100),
342 (12), 191 (10), 148 (26), 139 (30); HRMS (ESI) m/z [M+H]+ calcd
for C22H23NO6: 398.15981, found: 398.15952; ee >99% (HPLC;
l
2-PrOH (50:50 v:v), 0.5 mL/min, kmax 254 nm).
⁄Asterisks denote assignments that may be interchanged.
4.4.13. N-((1R,2S)-2-Hydroxy-1,2-diphenylethyl)-6-methoxy-9-
oxo-9H-xanthene-2-carboxamide (15)
Compound 15 was obtained as a white soꢃlid (162 mg, 94%); mp:
C
244–246 °C (chloroform/n-hexane);
ꢁ 10ꢀ3 g/mL in chloroform); IR (KBr):
1528, 1478, 1443, 1266, 834 cmꢀ1
½
a 2D5
m
ꢂ
ꢀ181.8 (c = 0.33
~
3434, 3341, 1633, 1618,
;
1H NMR (300.13 MHz,
[D6]DMSO): d = 9.00 (1H, d, J = 8.9 Hz, NH), 8.53 (1H, d, J = 2.0 Hz,

C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
1059
Column: amylose tris-3,5-dimethylphenylcarbamate coated onto
APS-Nucleosil (500 Å, 7 m, 20% w/w), Mobile phase: MeOH,
0.5 mL/min, kmax 254 nm).
163.4 (C-3), 157.3 (C-4a), 155.6 (C-10a), 141.2 (C-100), 135.7 (C-6),
135.2 (C-400), 128.7 (C-300 and C-500), 127.6 (C-1), 126.0 (C-8), 125.9
(C-200 and C-600), 124.4 (C-7), 121.2 (C-8a), 117.9 (C-5), 115.3 (C-9a),
114.2 (C-2), 101.5 (C-4), 67.3 (OCH2), 47.5 (C-10), 22.1 (Ar-CH3),
20.6 ppm (C-20); MS (ESI) m/z (%): 389 [M+H]++1 (29), 388 [M+H]+
(100), 349 (5), 270 (8), 219 (7), 149 (5); HRMS (ESI) m/z [M+H]+ calcd
for C24H21NO4: 388.15490, found: 388.15560; ee >98% (HPLC;
Column: amylose tris-3,5-dimethylphenylcarbamate coated onto
l
4.4.16. (S)-tert-Butyl 2-(6-methoxy-9-oxo-9H-xanthene-2-
carboxamido)propanoate (18)
Compound 18 was obtained as a white solid (142 mg, 97%); mp:
ꢃC
160–162 °C (MeOH/water); ½a D25
ꢂ
+29.9 (c = 0.67 ꢁ 10ꢀ3 g/mL in
~
dichloromethane); IR (KBr):
m
3389, 1735, 1645, 1622, 1525,
APS-Nucleosil (500 Å,
7 lm, 20% w/w), Mobile phase: ACN,
1480, 1458, 826 cmꢀ1 1H NMR (300.13 MHz, CDCl3): d = 8.67
;
0.5 mL/min, kmax 254 nm).
(1H, d, J = 2.3 Hz, H-1), 8.28 (1H, dd, J = 8.8 and 2.3 Hz, H-3), 8.27
(1H, d, J = 8.8 Hz, H-8), 7.54 (1H, d, J = 8.8 Hz, H-4), 6.99 (1H, dd,
J = 8.8 and 2.4 Hz, H-7), 6.92 (1H, d, J = 2.4 Hz, H-5), 6.91 (1H, d,
J = 7.0 Hz, NH), 4.70 (1H, quint, J = 7.0 Hz, H-10), 3.96 (3H, s, Ar-
OCH3), 1.53 (3H, d, J = 7.0 Hz, H-20), 1.51 ppm (9H, s, (C(CH3)3);
13C NMR (125.77 MHz, CDCl3): d = 175.7 (C-9), 172.2 (C@O, ester),
165.4 (C@O, amide), 165.2 (C-6), 158.0 (C-10a), 157.9 (C-4a),
133.9 (C-3), 129.8 (C-2), 128.4 (C-8), 124.8 (C-1), 121.3 (C-9a),
118.5 (C-4), 115.6 (C-8a), 113.7 (C-7), 100.4 (C-5), 82.3 (C-10),
55.9 (Ar-OCH3), 49.2 (C(CH3)3), 29.7 (C-20), 28.0 ppm ((CH3)3); MS
(ESI) m/z (%): 399 [M+H]++1 (25), 398 [M+H]+ (100), 342 (11),
191 (20), 148 (84), 139 (6); HRMS (ESI) m/z [M+H]+ calcd for
C22H23NO6: 398.15981, found: 398.15884; ee >99% (HPLC; Col-
umn: amylose tris-3,5-dimethylphenylcarbamate coated onto
4.4.19. (R)-N-(1-Hydroxy-3-methylbutan-2-yl)-2-((9-oxo-9H-
xanthen-3-yl)oxy)acetamide (21)
Compound 21 was obtained as a white solid (125 mg, 95%); mp:
ꢃC
184–186 °C (EtOH); ½a D25
ꢂ
ꢀ8.3 (c = 1.20 ꢁ 10ꢀ3 g/mL in dichloro-
~
m
methane); IR (KBr):
3412, 3276, 1645, 1614, 1559, 1461, 1438,
1242, 755; 1H NMR (300.13 MHz, CDCl3): d = 8.37 (1H, dd, J = 7.6
and 1.6 Hz, H-8), 8.35 (1H, d, J = 8.8 Hz, H-1), 7.77 (1H, ddd,
J = 8.0, 7.5 and 1.6 Hz, H-6), 7.52 (1H, dd, J = 8.0 and 0.6 Hz, H-5),
7.44 (1H, ddd, J = 7.6, 7.5 and 0.6 Hz, H-7), 7.06 (1H, dd, J = 8.8
and 2.4 Hz, H-2), 6.99 (1H, d, J = 2.4 Hz, H-4), 6.73 (1H, d,
J = 8.1 Hz, NH), 4.71 (2H, s, OCH2), 3.90 (1H, m, H-10), 3.78 (2H,
m, CH2OH), 1.97 (1H, m, H-20), 1.02 (3H, d, J = 6.7 Hz, H-30a)⁄,
0.97 ppm (3H, d, J = 6.7 Hz, H-30b)⁄; 13C NMR (75.47 MHz, CDCl3):
d = 176.2 (C-9), 167.7 (C@O, amide), 162.1 (C-3), 157.8 (C-4a),
156.2 (C-10a), 134.6 (C-6), 128.9 (C-1), 126.7 (C-8), 124.2 (C-7),
121.9 (C-8a), 117.8 (C-5), 116.9 (C-9a), 113.1 (C-2), 101.5 (C-4⁄),
67.5 (OCH2), 63.7 (CH2OH), 56.9 (C-10), 28.9 (C-20), 19.5 (C-30a) ,
18.7 ppm (C-30b)⁄; MS (ESI) m/z (%): 357 [M+H]++1 (19), 356
[M+H]+ (100), 325 (5), 282 (20), 256 (5); HRMS (ESI) m/z [M+H]+
calcd for C20H21NO5: 356.14925, found: 356.14918; ee >99%
(HPLC; Column: amylose tris-3,5-dimethylphenylcarbamate
APS-Nucleosil (500 Å, 7
lm, 20% w/w), Mobile phase: MeOH,
0.5 mL/min, kmax 254 nm).
4.4.17. (R)-2-((9-Oxo-9H-xanthen-3-yl)oxy)-N-(1-(p-
tolyl)ethyl)acetamide (19)
Compound 19 was obtained as a white solid (138 mg, 97%); mp:
ꢃC
172–174 °C (ethyl acetate/n-hexane);
10ꢀ3 g/mL in dichloromethane); IR (KBr):
1549, 1458, 1436, 840 cmꢀ1
½
a 2D5
ꢂ
ꢀ15.6 (c = 5.75 ꢁ
~
m
3286, 1647, 1611,
;
1H NMR (300.13 MHz, [D6]DMSO):
coated onto APS-Nucleosil (500 Å, 7 lm, 20% w/w), Mobile phase:
d = 8.63 (1H, d, J = 8.6 Hz, NH), 8.18 (1H, dd, J = 8.0 and 1.7 Hz,
H-8), 8.12 (1H, d, J = 8.8 Hz, H-1), 7.86 (1H, ddd, J = 8.5, 7.1 and
1.7 Hz, H-6), 7.65 (1H, dd, J = 8.5 and 1.0 Hz, H-5), 7.48 (1H, ddd,
J = 8.0, 7.1 and 1.0 Hz, H-7), 7.20 (2H, d, J = 8.0 Hz, H-200 and H-600),
7.12 (1H, dd, J = 8.8 and 2.3 Hz, H-2), 7.08 (1H, d, J = 2.3 Hz, H-4),
7.07 (2H, d, J = 8.0 Hz, H-300 and H-500), 4.98 (1H, m, H-10), 4.78 (2H,
s, OCH2), 2.22 (3H, s, Ar-CH3), 1.40 ppm (3H, d, J = 7.0 Hz, H-20); 13C
NMR (75.47 MHz, [D6]DMSO): d = 175.0 (C-9), 165.9 (C@O, amide),
163.5 (C-3), 157.6 (C-4a), 155.6 (C-10a), 141.2 (C-100), 135.7 (C-6),
135.2 (C-400), 128.8 (C-300 and C-500), 127.6 (C-1), 126.0 (C-8), 125.9
(C-200 and C-600), 124.4 (C-7), 121.2 (C-8a), 118.0 (C-5), 115.3 (C-9a),
114.2 (C-2), 101.5 (C-4), 67.3 (OCH2), 47.5 (C-10), 22.2 (Ar-CH3),
20.6 ppm (C-20); MS (ESI) m/z (%): 389 [M+H]++1 (29), 388 [M+H]+
(100), 349 (3), 270 (8), 149 (13); HRMS (ESI) m/z [M+H]+ calcd for
MeOH, 0.5 mL/min, kmax 254 nm).
⁄Asterisks denote assignments that may be interchanged.
4.4.20. (S)-N-(1-Hydroxy-3-methylbutan-2-yl)-2-((9-oxo-9H-
xanthen-3-yl)oxy)acetamide (22)
Compound 22 was obtained as a white solid (127 mg, 97%); mp:
ꢃC
184–186 °C (EtOH); ½a D25
ꢂ
+ 8.5 (c = 1.20 ꢁ 10ꢀ3 g/mL in dichloro-
~
m
methane); IR (KBr):
3407, 3276, 1644, 1613, 1554, 1460, 1438,
1240, 755 cmꢀ1 1H NMR (300.13 MHz, CDCl3): d = 8.37 (1H, dd,
;
J = 7.6 and 1.6 Hz, H-8), 8.35 (1H, d, J = 8.8 Hz, H-1), 7.77 (1H,
ddd, J = 8.0, 7.5 and 1.6 Hz, H-6), 7.52 (1H, dd, J = 8.0 and 0.6 Hz,
H-5), 7.44 (1H, ddd, J = 7.6, 7.5 and 0.6 Hz, H-7), 7.06 (1H, dd,
J = 8.8 and 2.4 Hz, H-2), 6.99 (1H, d, J = 2.4 Hz, H-4), 6.73 (1H, d,
J = 8.1 Hz, NH), 4.71 (2H, s, OCH2), 3.90 (1H, m, H-10), 3.78 (2H,
m, CH2OH), 1.98 (1H, m, H-20), 1.02 (3H, d, J = 6.7 Hz, H-30a)⁄,
0.98 ppm (3H, d, J = 6.7 Hz, H-30b)⁄; 13C NMR (75.47 MHz, CDCl3):
d = 176.2 (C-9), 167.7 (C@O, amide), 162.1 (C-3), 157.8 (C-4a),
156.2 (C-10a), 134.6 (C-6), 128.9 (C-1), 126.7 (C-8), 124.2 (C-7),
121.9 (C-8a), 117.8 (C-5), 116.9 (C-9a), 113.1 (C-2), 101.5 (C-4⁄),
67.5 (OCH2), 63.7 (CH2OH), 56.9 (C-10), 28.9 (C-20), 19.5 (C-30a) ,
18.7 ppm (C-30b)⁄; MS (ESI) m/z (%): 357 [M+H]++1 (21), 356
[M+H]+ (100), 325 (5), 282 (20), 256 (6); HRMS (ESI) m/z [M+H]+
calcd for C20H21NO5: 356.14925, found: 356.14923; ee >99%
(HPLC; Column: amylose tris-3,5-dimethylphenylcarbamate
C24H21NO4: 388.15490, found: 388.15550; ee >99% (HPLC; Column:
amylosetris-3,5-dimethylphenylcarbamatecoated ontoAPS-Nucle-
osil (500 Å, 7
254 nm).
lm, 20% w/w), Mobile phase: ACN, 0.5 mL/min, kmax
4.4.18. (S)-2-((9-Oxo-9H-xanthen-3-yl)oxy)-N-(1-(p-
tolyl)ethyl)acetamide (20)
Compound 20 was obtained as a white solid (137 mg, 96%); mp:
ꢃC
172–174 °C (ethyl acetate/n-hexane);
10ꢀ3 g/mL in dichloromethane); IR (KBr):
1549, 1457, 1433, 834 cmꢀ1
½
a 2D5
ꢂ
+14.3 (c = 5.75 ꢁ
~
m
3286, 1647, 1610,
;
1H NMR (300.13 MHz, [D6]DMSO):
coated onto APS-Nucleosil (500 Å, 7 lm, 20% w/w), Mobile phase:
d = 8.63 (1H, d, J = 8.6 Hz, NH), 8.18 (1H, dd, J = 8.0 and 1.7 Hz,
H-8), 8.12 (1H, d, J = 8.8 Hz, H-1), 7.86 (1H, ddd, J = 8.5, 7.1 and
1.7 Hz, H-6), 7.65 (1H, dd, J = 8.5 and 1.0 Hz, H-5), 7.48 (1H, ddd,
J = 8.0, 7.1 and 1.0 Hz, H-7), 7.21 (2H, d, J = 8.1 Hz, H-200 and H-600),
7.12 (1H, dd, J = 8.8 and 2.3 Hz, H-2), 7.08 (1H, d, J = 2.3 Hz, H-4),
7.07 (2H, d, J = 8.1 Hz, H-300 and H-500), 4.98 (1H, m, H-10), 4.78 (2H,
s, OCH2), 2.22 (3H, s, Ar-CH3), 1.40 ppm (3H, d, J = 7.0 Hz, H-20); 13C
NMR (75.47 MHz, [D6]DMSO): d = 174.9 (C-9), 165.9 (C@O, amide),
MeOH, 0.5 mL/min, kmax 254 nm).
⁄Asterisks denote assignments that may be interchanged.
4.4.21. (R)-N-(1-Hydroxy-4-methylpentan-2-yl)-2-((9-oxo-9H-
xanthen-3-yl)oxy)acetamide (23)
Compound 23 was obtained as a white solid (134 mg, 98%); mp:
ꢃC
154–156 °C (ethyl acetate/n-hexane); ½a D25
ꢂ
+18.0 (c = 10.00 ꢁ
10ꢀ3 g/mL in dichloromethane); IR (KBr):
3406, 3350–3312,
~
m

1060
C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
1652, 1613, 1539, 1461, 1436, 1252, 756 cmꢀ1
;
1H NMR
101.6 (C-4), 67.3 (OCH2), 64.5 (CH2OH), 55.0 ppm (C-10); MS (ESI)
.
(300.13 MHz, [D6]DMSO): d = 8.20 (1H, dd, J = 8.0 and 1.6 Hz,
H-8), 8.13 (1H, d, J = 9.4 Hz, H-1), 7.87 (1H, ddd, J = 8.2, 7.6 and
1.6 Hz, H-6), 7.64 (1H, dd, J = 8.2 and 0.9 Hz, H-5), 7.49 (1H, ddd,
J = 8.0, 7.6 and 0.9 Hz, H-7), 7.13 (1H, dd, J = 9.4 and 2.4 Hz, H-2),
7.09 (1H, d, J = 2.4 Hz, H-4), 4.72 (2H, s, OCH2), 3.90 (1H, m,
H-10), 3.75 (2H, m, CH2OH), 1.54 (1H, m, H-30), 1.32 (2H, m, H-20),
0.84 (3H, d, J = 6.6 Hz, H-40a)⁄, 0.81 ppm (3H, d, J = 6.6 Hz,
H-40b)⁄; 13C NMR (75.47 MHz, [D6]DMSO): d = 174.9 (C-9), 166.4
(C@O, amide), 163.4 (C-3), 157.3 (C-4a), 155.6 (C-10a), 135.2
(C-6), 127.6 (C-1), 125.9 (C-8), 124.4 (C-7), 121.2 (C-8a), 117.9
(C-5), 115.3 (C-9a), 114.2 (C-2), 101.5 (C-4), 67.3 (OCH2), 63.7
(CH2OH), 48.7 (⁄C-10), 39.0 (C-20), 24.2 (C-30), 23.4 (C-40a)⁄,
21.7 ppm (C-40b) ; MS (ESI) m/z (%): 371 [M+H]++1 (23), 370
[M+H]+ (100), 328 (30), 282 (23), 256 (6); HRMS (ESI) m/z [M+H]+
calcd C21H23NO5: 370.16490, found: 370.16485; ee >99% (HPLC;
Column: amylose tris-3,5-dimethylphenylcarbamate coated onto
m/z (%): 391 [M+H] +1 (23), 390 [M+H]+ (100), 320 (5), 267 (16);
HRMS (ESI) m/z [M+H]+ calcd for C23H19NO5: 390.13360, found:
390.13370; ee >99% (HPLC; Column: cellulose tris-3,5-dimethyl-
phenylcarbamate coated onto APS-Nucleosil (500 Å, 7 lm, 20% w/
w), Mobile phase: n-hexane/EtOH (70:30 v:v), 0.5 mL/min, kmax
254 nm).
⁄Asterisks denote assignments that may be interchanged.
4.4.24. (S)-N-(2-Hydroxy-1-phenylethyl)-2-((9-oxo-9H-xanthen-
3-yl)oxy)acetamide (26)
Compound 26 was obtained as a white solid (135 mg, 94%); mp:
ꢃC
181–182 °C (ethyl acetate/n-hexane);
½
a 2D5
ꢂ
ꢀ16.0 (c = 2.90
ꢁ 10ꢀ3 g/mL in dichloromethane); IR (KBr):
m
3369, 3316, 1667,
~
1612, 1549, 1461, 1434, 1257, 842 cmꢀ1 1H NMR (300.13 MHz,
;
[D6]DMSO): d = 8.60 (1H, d, J = 8.4 Hz, NH), 8.18 (1H, dd, J = 7.7
and 1.7 Hz, H-8), 8.12 (1H, d, J = 9.5 Hz, H-1), 7.87 (1H, ddd,
J = 8.0, 7.3 and 1.7 Hz, H-6), 7.65 (1H, dd, J = 8.0 and 0.9 Hz, H-5),
7.48 (1H, ddd, J = 7.7, 7.3 and 0.9 Hz, H-7), 7.33 (2H, d, J = 6.7 Hz,
H-200 and H-600), 7.30(2H, t, J = 6.7 Hz, H-300 and H-500), 7.22 (1H, d,
J = 6.7 Hz, H-400), 7.14 (1H, dd, J = 9.5 and 2.3 Hz, H-2), 7.13 (1H,
d, J = 2.3 Hz, H-4), 4.99 (1H, t, J = 5.6 Hz, OH), 4.92 (1H, m, H-10),
4.84 (1H, d, J = 14.6 Hz, OCH2)⁄, 4.77 (1H, d, J = 14.6 Hz, OCH2)⁄,
3.63 ppm (2H, m, CH2OH); 13C NMR (75.47 MHz, [D6]DMSO):
d = 174.9 (C-9), 166.6 (C@O, amide), 163.5 (C-3), 157.3 (C-4a),
155.6 (C-10a), 140.8 (C-100), 135.2 (C-6), 128.1 (C-300 and C-500),
127.6 (C-1), 127.0 (C-200 and C-600), 126.9 (C-400), 126.0 (C-8),
124.4 (C-7), 121.2 (C-8a), 118.0 (C-5), 115.3 (C-9a), 114.2 (C-2),
101.5 (C-4), 67.3 (OCH2), 64.5 (CH2OH), 55.0 ppm (C-10); MS (ESI)
m/z (%): 391 [M+H]++1 (30), 390 [M+H]+ (100), 354 (21), 255
(15), 191 (23), 179 (7); HRMS (ESI) m/z [M+H]+ calcd for
APS-Nucleosil (500 Å,
7 lm, 20% w/w), Mobile phase: ACN,
0.5 mL/min, kmax 254 nm).
⁄Asterisks denote assignments that may be interchanged.
4.4.22. (S)-N-(1-Hydroxy-4-methylpentan-2-yl)-2-((9-oxo-9H-
xanthen-3-yl)oxy)acetamide (24)
Compound 24 was obtained as a white solid (131 mg, 96%); mp:
ꢃC
154–156 °C (ethyl acetate/n-hexane); ½a D25
ꢂ
ꢀ18.4 (c = 10.00 ꢁ
10ꢀ3g/mL in dichloromethane); IR (KBr):
m
3410, 3350–3312,
~
1654, 1615, 1541, 1462, 1437, 1253, 758 cmꢀ1
;
1H NMR
(500.13 MHz, [D6]DMSO): d = 8.19 (1H, dd, J = 8.0 and 1.6 Hz,
H-8), 8.13 (1H, d, J = 9.4 Hz, H-1), 7.87 (1H, ddd, J = 8.2, 7.6 and
1.6 Hz, H-6), 7.64 (1H, dd, J = 8.2 and 0.9 Hz, H-5), 7.49 (1H, ddd,
J = 8.0, 7.6 and 0.9 Hz, H-7), 7.13 (1H, dd, J = 9.4 and 2.4 Hz, H-2),
7.09 (1H, d, J = 2.4 Hz, H-4), 4.71 (2H, s, OCH2), 3.90 (1H, m,
H-10), 3.78 (2H, m, CH2OH), 1.55 (1H, m, H-30), 1.32 (2H, m, H-20),
0.84 (3H, d, J = 6.6 Hz, H-40a)⁄, 0.81 ppm (3H, d, J = 6.6 Hz,
H-40b)⁄; 13C NMR (75.47 MHz, [D6]DMSO): d = 174.9 (C-9), 166.4
(C@O, amide), 163.4 (C-3), 157.3 (C-4a), 155.6 (C-10a), 135.2
(C-6), 127.6 (C-1), 125.9 (C-8), 124.4 (C-7), 121.2 (C-8a), 117.9
(C-5), 115.3 (C-9a), 114.2 (C-2), 101.5 (C-4), 67.3 (OCH2), 63.7
(CH2OH), 48.7 (⁄C-10), 39.0 (C-20), 24.2 (C-30), 23.4 (C-40a)⁄,
21.7 ppm (C-40b) ; MS (ESI) m/z (%): 371 [M+H]++1 (23), 370
[M+H]+ (100), 356 (6), 282 (34), 256 (9), 236 (5); HRMS (ESI) m/z
[M+H]+ calcd for C21H23NO5: 370.16490, found: 370.16487; ee
>99% (HPLC; Column: amylose tris-3,5-dimethylphenylcarbamate
C
23H19NO5: 390.13360, found: 390.13252; ee >99% (HPLC; Col-
umn: cellulose tris-3,5-dimethylphenylcarbamate coated onto
APS-Nucleosil (500 Å, 7 m, 20% w/w), Mobile phase: n-hexane/
l
EtOH (70:30 v:v), 0.5 mL/min, kmax 254 nm).
⁄Asterisks denote assignments that may be interchanged.
4.4.25. (R)-N-(1-Hydroxypropan-2-yl)-2-((9-oxo-9H-xanthen-3-
yl)oxy)acetamide (27)
Compound 27 was obtained as a white solid (117 mg, 97%); mp:
ꢃC
183–185 °C (ethyl acetate/n-hexane);
½
a 2D5
m
ꢂ
+6.2 (c = 3.33
ꢁ 10ꢀ3 g/mL in dichloromethane); IR (KBr):
3383, 3313, 1650,
~
1616, 1552, 1460, 1434, 1250, 753 cmꢀ1 1H NMR (300.13 MHz,
;
coated onto APS-Nucleosil (500 Å, 7
l
m, 20% w/w), Mobile phase:
CDCl3): d = 8.37 (1H, dd, J = 7.6 and 1.6 Hz, H-8), 8.35 (1H, d,
J = 8.8 Hz, H-1), 7.77 (1H, ddd, J = 8.3, 7.5 and 1.6 Hz, H-6), 7.52
(1H, dd, J = 8.3 and 0.8 Hz, H-5), 7.44 (1H, ddd, J = 7.6, 7.5 and
0.8 Hz, H-7), 7.06 (1H, dd, J = 8.8 and 2.4 Hz, H-2), 6.98 (1H, d,
J = 2.4 Hz, H-4), 6.73 (1H, d, J = 7.1 Hz, NH), 4.67 (2H, s, OCH2),
4.26 (1H, m, H-10), 3.73 (2H, m, CH2OH), 1.31 ppm (3H, d,
J = 6.8 Hz, H-20); 13C NMR (75.47 MHz, CDCl3): d = 176.2 (C-9),
167.3 (C@O, amide), 162.0 (C-3), 157.8 (C-4a), 156.2 (C-10a),
134.6 (C-6), 128.9 (C-1), 126.7 (C-8), 124.2 (C-7), 121.9 (C-8a),
117.8 (C-5), 116.9 (C-9a), 113.2 (C-2), 101.5 (C-4), 67.4 (OCH2),
66.6 (C-10), 47.5 (CH2OH), 17.0 ppm (C-20); MS (ESI) m/z (%): 329
[M+H]++1 (19), 328 [M+H]+ (100), 282 (10), 256 (2); HRMS (ESI)
m/z [M+H]+ calcd for C18H17NO5: 328.11795, found: 328.11795;
ee >99% (HPLC; Column: amylose tris-3,5-dimethylphenylcarba-
ACN, 0.5 mL/min, kmax 254 nm).
⁄Asterisks denote assignments that may be interchanged.
4.4.23. (R)-N-(2-Hydroxy-1-phenylethyl)-2-((9-oxo-9H-xanthen-
3-yl)oxy)acetamide (25)
Compound 25 was obtained as a white solid (137 mg, 95%); mp:
ꢃC
181–182 °C (ethyl acetate/n-hexane);
½
a 2D5
ꢂ
+16.2 (c = 2.90
ꢁ 10ꢀ3 g/mL in dichloromethane); IR (KBr):
3368, 3308, 1659,
~
m
1606, 1541, 1451, 1428, 1252, 834 cmꢀ1 1H NMR (300.13 MHz,
;
[D6]DMSO): d = 8.61 (1H, d, J = 8.3 Hz, NH), 8.19 (1H, dd, J = 7.7
and 1.7 Hz, H-8), 8.13 (1H, d, J = 9.4 Hz, H-1), 7.87 (1H, ddd,
J = 8.0, 7.3 and 1.7 Hz, H-6), 7.65 (1H, dd, J = 8.0 and 0.9 Hz, H-5),
7.49 (1H, ddd, J = 7.7, 7.3 and 0.9 Hz, H-7), 7.33 (2H, d, J = 6.7 Hz,
H-200 and H-600), 7.30(2H, t, J = 6.7 Hz, H-300 and H-500), 7.22 (1H, d,
J = 6.7 Hz, H-400), 7.14 (1H, dd, J = 9.4 and 2.3 Hz, H-2), 7.13 (1H,
d, J = 2.3 Hz, H-4), 5.02 (1H, t, J = 5.6 Hz, OH), 4.93 (1H, m, H-10),
4.83 (1H, d, J = 14.7 Hz, OCH2)⁄, 4.78 (1H, d, J = 14.7 Hz, OCH2)⁄,
3.63 ppm (2H, m, CH2OH); 13C NMR (75.47 MHz, [D6]DMSO):
d = 175.0 (C-9), 166.7 (C@O, amide), 163.5 (C-3), 157.4 (C-4a),
155.7 (C-10a), 140.8 (C-100), 135.3 (C-6), 128.1 (C-300 and C-500),
127.7 (C-1), 127.0 (C-200 and C-600), 126.9 (C-400), 126.0 (C-8),
124.5 (C-7), 121.2 (C-8a), 118.0 (C-5), 115.4 (C-9a), 114.2 (C-2),
mate coated onto APS-Nucleosil (500 Å, 7 lm, 20% w/w), Mobile
phase: ACN, 0.5 mL/min, kmax 254 nm).
4.4.26. (S)-N-(1-Hydroxypropan-2-yl)-2-((9-oxo-9H-xanthen-3-
yl)oxy)acetamide (28)
Compound 28 was obtained as a white solidꢃ(116 mg, 96%); mp:
C
182–184 °C (ethyl acetate/n-hexane);
½
a 2D5
m
ꢂ
ꢀ6.1 (c = 3.33
ꢁ 10ꢀ3 g/mL in dichloromethane); IR (KBr):
3405, 3313, 1651,
~
1613, 1541, 1461, 1436, 1252, 755 cmꢀ1 1H NMR (300.13 MHz,
;

C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
1061
~
CDCl3): d = 8.38 (1H, dd, J = 7.6 and 1.6 Hz, H-8), 8.35 (1H, d,
J = 8.8 Hz, H-1), 7.77 (1H, ddd, J = 8.3, 7.5 and 1.6 Hz, H-6), 7.52
(1H, dd, J = 8.3 and 0.8 Hz, H-5), 7.44 (1H, ddd, J = 7.6, 7.5 and
0.8 Hz, H-7), 7.06 (1H, dd, J = 8.8 and 2.4 Hz, H-2), 6.98 (1H, d,
J = 2.4 Hz, H-4), 6.73 (1H, d, J = 7.0 Hz, NH), 4.67 (2H, s, OCH2),
4.26 (1H, m, H-10), 3.73 (2H, m, CH2OH), 1.31 ppm (3H, d,
J = 6.8 Hz, H-20); 13C NMR (75.47 MHz, CDCl3): d = 176.2 (C-9),
167.3 (C@O, amide), 162.0 (C-3), 157.8 (C-4a), 156.2 (C-10a),
134.6 (C-6), 128.9 (C-1), 126.7 (C-8), 124.2 (C-7), 121.9 (C-8a),
117.8 (C-5), 116.9 (C-9a), 113.2 (C-2), 101.5 (C-4), 67.4 (OCH2),
66.6 (C-10), 47.5 (CH2OH), 17.0 ppm (C-20); MS (ESI) m/z (%): 329
[M+H]++1 (20), 328 [M+H]+ (100), 282 (14), 256 (5); HRMS (ESI)
m/z [M+H]+ calcd for C18H17NO5: 328.11795, found: 328.11791;
ee >99% (HPLC; Column: amylose tris-3,5-dimethylphenylcarba-
g/mL in chloroform); IR (KBr):
m
3412–3190, 3290, 1666, 1617,
;
1546, 1461, 1441, 1167, 759 cmꢀ1
1H NMR (300.13 MHz, [D6-
]DMSO): d = 8.53 (1H, d, J = 8.9 Hz, NH), 8.19 (1H, dd, J = 7.9 and
1.5 Hz, H-8), 8.08 (1H, d, J = 8.9 Hz, H-1), 7.88 (1H, ddd, J = 8.5, 7.1
and 1.5 Hz, H-6), 7.65 (1H, dd, J = 8.5 and 0.8 Hz, H-5), 7.49 (1H,
ddd, J = 7.9, 7.1 and 0.8 Hz, H-7), 7.19 (10H, m, H-Ar), 6.98 (1H, dd,
J = 8.9 and 2.2 Hz, H-2), 6.92 (1H, d, J = 2.2 Hz, H-4), 5.57 (1H d,
J = 4.9 Hz, OH), 5.01 (1H, dd, J = 8.9 and 6.9 Hz, H-10), 4.87 (1H, dd,
J = 6.9 and 4.9 Hz CHOH), 4.66 (1H, d, J = 15.1 Hz, OCH2)⁄, 4.61 ppm
(1H, d, J = 15.1 Hz, OCH2)⁄; 13C NMR (75.47 MHz, [D6]DMSO):
d = 175.0 (C-9), 165.8 (C@O, amide), 163.3 (C-3), 157.3 (C-4a),
155.6 (C-10a), 142.8 (C-100), 140.1 (C-1000), 135.2 (C-6), 128.3 (C-300
and C-500), 128.3 (C-3000 and C-5000), 127.6 (C-1), 127.5 (C-200 and C-
600), 127.0 (C-2000 and C-6000), 126.8 (C-400), 126.7 (C-4000), 126.0 (C-8),
124.4 (C-7), 121.2 (C-8a), 118.0 (C-5), 115.3 (C-9a), 114.0 (C-2),
101.4 (C-4), 74.4 (CHOH), 67.1 (OCH2), 58.3 ppm (C-10); MS (ESI)
m/z (%): 467 [M+H]++1 (27), 466 [M+H]+ (83), 448 (100), 304 (22),
282 (31), 256 (8); HRMS (ESI) m/z [M+H]+ calcd for C29H23NO5:
466.16490, found: 466.16502; ee >98% (HPLC; Column: (S,S)-
Whelk-O1, Mobile phase: ACN/MeOH (50:50 v:v), 1.0 mL/min, kmax
254 nm).
mate coated onto APS-Nucleosil (500 Å, 7 lm, 20% w/w), Mobile
phase: ACN, 0.5 mL/min, kmax 254 nm).
4.4.27. (R)-N-(2-Hydroxypropyl)-2-((9-oxo-9H-xanthen-3-
yl)oxy)acetamide (29)
Compound 29 was obtained as a white solid (119 mg, 98%); mp:
ꢃ
161–163 °C (ethyl acetate/n-hexane);
½
a 2D5 C
ꢂ
ꢀ12.8 (c = 3.10
ꢁ 10ꢀ3 g/mL in dichloromethane); IR (KBr):
m
3440, 3285, 1652,
⁄Asterisks denote assignments that may be interchanged.
~
1611, 1564, 1457, 1319, 1252, 753 cmꢀ1 1H NMR (300.13 MHz,
;
CDCl3): d = 8.38 (1H, dd, J = 7.6 and 1.6 Hz, H-8), 8.35 (1H, d,
J = 8.8 Hz, H-1), 7.77 (1H, ddd, J = 8.4, 7.5 and 1.6 Hz, H-6), 7.53 (1H,
dd, J = 8.4 and 0.6 Hz, H-5), 7.44 (1H, ddd, J = 7.6, 7.5 and 0.6 Hz, H-
7), 7.06 (1H, dd, J = 8.8 and 2.4 Hz, H-2), 6.98 (1H, d, J = 2⁄.4 Hz, H-4),
4.70 (2H, s, OCH2), 4.04 (1H, m, H-20), 3.64 (1H, m, H-10) , 3.27 (1H,
m, H-10)⁄, 1.28 ppm (3H, d, J = 6.3 Hz, CH3); 13C NMR (75.47 MHz,
CDCl3): d = 176.2 (C-9), 167.8 (C@O, amide), 162.1 (C-3), 157.8 (C-
4a), 156.2 (C-10a), 134.6 (C-6), 128.9 (C-1), 126.7 (C-8), 124.2 (C-7),
121.9 (C-8a), 117.8 (C-5), 116.9 (C-9a), 113.2 (C-2), 101.5 (C-4), 67.4
(OCH2), 67.3 (C-20), 46.4 (C-10), 21.1 ppm (CH3); MS (ESI) m/z (%):
329 [M+H]++1 (20), 328 [M+H]+ (100), 282 (14), 256 (4); HRMS (ESI)
m/z [M+H]+ calcd for C18H17NO5: 328.11795, found: 328.11794; ee
>99% (HPLC; Column: amylose tris-3,5-dimethylphenylcarbamate
4.4.30. N-((1S,2R)-2-Hydroxy-1,2-diphenylethyl)-2-((9-oxo-9H-
xanthen-3-yl)oxy)acetamide (32)
Compound 32 was obtained as a white solid (162 mg, 94%); mp:
ꢃ
216–219 °C (chloroform/n-hexane); ½a D25 C
ꢂ
+66.6 (c = 0.33 ꢁ 10ꢀ3 g/
3412–3190, 3290, 1665, 1616, 1545,
1H NMR (300.13 MHz, [D6]DMSO):
~
mL in chloroform); IR (KBr):
m
1460, 1439, 1165, 759 cmꢀ1
;
d = 8.53 (1H, d, J = 8.9 Hz, NH), 8.19 (1H, dd, J = 7.9 and 1.6 Hz, H-8),
8.08 (1H, d, J = 8.9 Hz, H-1), 7.88 (1H, ddd, J = 8.5, 7.1 and 1.6 Hz,
H-6), 7.65 (1H, dd, J = 8.5 and 0.9 Hz, H-5), 7.49 (1H, ddd, J = 7.9, 7.1
and 0.9 Hz, H-7), 7.19 (10H, m, H-Ar), 6.98 (1H, dd, J = 8.9 and
2.2 Hz, H-2), 6.92 (1H, d, J = 2.2 Hz, H-4), 5.57 (1H d, J = 4.9 Hz, OH),
5.01 (1H, dd, J = 8.9 and 6.9 Hz, H-10), 4.87 (1H, dd, J = 6.9 and 4.9 Hz
CHOH), 4.66 (1H, d, J = 15.1 Hz, OCH2)⁄, 4.61 ppm (1H, d, J = 15.1 Hz,
OCH2)⁄; 13C NMR (75.47 MHz, [D6]DMSO): d = 175.0 (C-9), 165.8
(C@O, amide), 163.3 (C-3), 157.3 (C-4a), 155.6 (C-10a), 142.8 (C-100),
140.1 (C-1000), 135.2 (C-6), 128.3 (C-300 and C-500), 128.3 (C-3000 and
C-5000), 127.6 (C-1), 127.5 (C-200 and C-600), 127.0 (C-2000 and C-6000),
126.8 (C-400), 126.7 (C-4000), 126.0 (C-8), 124.4 (C-7), 121.2 (C-8a),
118.0 (C-5), 115.3 (C-9a), 114.0 (C-2), 101.4 (C-4), 74.4 (CHOH), 67.1
(OCH2), 58.3 ppm (C-10); MS (ESI) m/z (%): 467 [M+H]++1 (18), 466
[M+H]+ (56), 448 (100), 304 (53), 282 (41), 256 (13); HRMS (ESI) m/z
[M+H]+ calcd for C29H23NO5: 466.16490, found: 466.16495; ee >99%
(HPLC; Column: (S,S)-Whelk-O1, Mobile phase: ACN/MeOH (50:50
v:v), 1.0 mL/min, kmax 254 nm).
coated onto APS-Nucleosil (500 Å, 7 lm, 20% w/w), Mobile phase:
ACN, 0.5 mL/min, kmax 254 nm).
4.4.28. (S)-N-(2-Hydroxypropyl)-2-((9-oxo-9H-xanthen-3-
yl)oxy)acetamide (30)
Compound 30 was obtained as a white solid (117 mg, 97%); mp:
ꢃ
161–163 °C (ethyl acetate/n-hexane);
½
a 2D5 C
ꢂ
+12.9 (c = 3.10
3438, 3285, 1658,
ꢁ 10ꢀ3 g/mL in dichloromethane); IR (KBr):
m
~
1613, 1560, 1458, 1320, 1252, 754 cmꢀ1 1H NMR (300.13 MHz,
;
CDCl3): d = 8.38 (1H, dd, J = 7.6 and 1.6 Hz, H-8), 8.35 (1H, d,
J = 8.8 Hz, H-1), 7.77 (1H, ddd, J = 8.4, 7.5 and 1.6 Hz, H-6), 7.53 (1H,
dd, J = 8.4 and 0.6 Hz, H-5), 7.44 (1H, ddd, J = 7.6, 7.5 and 0.6 Hz,
H-7), 7.06 (1H, dd, J = 8.8 and 2.4 Hz, H-2), 6.98 (1H, d, J = 2.4 Hz,
H-4), 4.70 (2H, s, OCH2), 4.04 (1H, m, H-20), 3.64 (1H, m, H-10)⁄, 3.27
(1H, m, H-10)⁄, 1.28 ppm (3H, d, J = 6.3 Hz, CH3); 13C NMR
(75.47 MHz, CDCl3): d = 176.2 (C-9), 167.8 (C@O, amide), 162.1
(C-3), 157.8 (C-4a), 156.2 (C-10a), 134.6 (C-6), 128.9 (C-1), 126.7
(C-8), 124.2 (C-7), 121.9 (C-8a), 117.8 (C-5), 116.9 (C-9a), 113.2
(C-2), 101.5 (C-4), 67.4 (OCH2), 67.3 (C-20), 46.4 (C-10), 21.1 ppm
(CH3); MS (ESI) m/z (%): 329 [M+H]++1 (19), 328 [M+H]+ (100), 282
(17), 256 (5); HRMS (ESI) m/z [M+H]+ calcd for C18H17NO5:
328.11795, found: 328.11794; ee >99% (HPLC; Column: amylose
tris-3,5-dimethylphenylcarbamate coated onto APS-Nucleosil
⁄Asterisks denote assignments that may be interchanged.
4.5. Biological evaluation
4.5.1. Cell cultures
The three human tumor cell lines, A375-C5 (melanoma), MCF-7
(breast adenocarcinoma), and NCI-H460 (non-small cell lung can-
cer), were grown as monolayer and routinely maintained in cell
culture medium RPMI-1640 (with Glutamax, Lonza) supplemented
with 5% heat-inactivated fetal bovine serum (FBS), and incubated
in a humidified incubator at 37 °C with 5% CO2 (Hera Cell, Herae-
us). Cell number and viability were routinely determined with Try-
pan Blue (Sigma) exclusion assay. All experiments were performed
with cells in exponential growth with viabilities over 90% and re-
peated at least three times.
(500 Å, 7
254 nm).
lm, 20% w/w), Mobile phase: ACN, 0.5 mL/min, kmax
4.4.29. N-((1R,2S)-2-Hydroxy-1,2-diphenylethyl)-2-((9-oxo-9H-
xanthen-3-yl)oxy)acetamide (31)
4.5.2. Cell growth assay inhibitory assay
Compound 31 was obtained as a white solid (163 mg, 95%); mp:
The effects of compounds on the growth of human tumor cell
lines were evaluated according to the procedure adopted by the
ꢃC
215–218 °C (chloroform/n-hexane); ½a D25
ꢂ
ꢀ66.7 (c = 0.33 ꢁ 10ꢀ3

1062
C. Fernandes et al. / Bioorg. Med. Chem. 22 (2014) 1049–1062
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ited 50% of the net cell growth, was calculated as described.46
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This work was funded by FCT – Fundação para a Ciência e a
Tecnologia under the project CEQUIMED-PEst-OE/SAU/UI4040/
2011; CESPU 10-GCQF-CICS-09 project; FCT-GRICES/CAPES 00770
29/05/08 project; University of Porto/Santander Totta (Investi-
gação Científica na Pré-Graduação) projects; and also funded by
FEDER funds through the COMPETE program under the project
FCOMP-01-0124-FEDER-011057.
The authors also gratefully acknowledge the BRC, ScAWAKE and
Bioscience Program funds from Faculty of Science and a grant from
Department of Zoology and the graduate school scholarship of
Kasetsart University, Thailand to KM; and also thank to Sara Cravo
for technical support in microwave methods.
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