Design, synthesis and structure–activity relationships of novel N11-, C12- and C13-substituted 15-membered homo-aza-clarithromycin derivatives against various resistant bacteria

Article abstract of DOI:10.1016/j.bioorg.2021.104992

Bacterial infections are still the main significant problem of public health in the world, and their elimination will greatly rely on the discovery of antibacterial drugs. In the processes of our searching for novel macrolide derivatives with excellent activity against sensitive and resistant bacteria, three series of novel N11-, C12- and C13-substituted 15-membered homo-aza-clarithromycin derivatives were designed and synthesized as Series A, B and C by creatively opening the lactone ring of clarithromycin (CAM), introducing various 4-substituted phenyl-1H-1,2,3-triazole side chains at the N11, C12 or C13 position of CAM and macrolactonization. The results from their in vitro antibacterial activity demonstrated that compounds 20c, 20d and 20f displayed not only the most potent activity against S. aureus ATCC25923 with the MIC values of 0.5, 0.5 and 0.5 μg/mL, but also greatly improved activity against B. subtilis ATCC9372 with the MIC values of less than or equal to 0.25, 0.25 and 0.25 μg/mL, respectively. In particular, compound 11g exhibited the strongest antibacterial effectiveness against all the tested resistant bacterial strains and had well balanced activity with the MIC values of 4–8 μg/mL. Further study on minimum bactericidal concentration and kinetics confirmed that compound 11g possessed a bacteriostatic effect on bacterial proliferation. Moreover, the results of molecular docking revealed an potential additional binding force between compound 11g and U790 in addition to the normal binding force of macrolide skeleton, which may explain why this compound performed the most potent activity against resistant bacteria. The results of cytotoxic assay indicated that compounds 20c, 20d and 20f were non-toxic to human breast cancer MCF-7 cells at its effective antibacterial concentration.

Full text of DOI:10.1016/j.bioorg.2021.104992

Bioorganic Chemistry 113 (2021) 104992
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Design, synthesis and structure–activity relationships of novel N11-, C12-
and C13-substituted 15-membered homo-aza-clarithromycin derivatives
against various resistant bacteria
Yinhui Qin, Di Song, Yuetai Teng, Xingbang Liu, Panpan Zhang, Nan Zhang, Na Zhang,
Weijin Chen, Shutao Ma*
Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Sciences, Cheeloo College of Medicine,
Shandong University, 44 West Wenhua Road, Jinan 250012, PR China
A R T I C L E I N F O
A B S T R A C T
Keywords:
Bacterial infections are still the main significant problem of public health in the world, and their elimination will
greatly rely on the discovery of antibacterial drugs. In the processes of our searching for novel macrolide de-
rivatives with excellent activity against sensitive and resistant bacteria, three series of novel N11-, C12- and C13-
substituted 15-membered homo-aza-clarithromycin derivatives were designed and synthesized as Series A, B and
C by creatively opening the lactone ring of clarithromycin (CAM), introducing various 4-substituted phenyl-1H-
1,2,3-triazole side chains at the N11, C12 or C13 position of CAM and macrolactonization. The results from their
in vitro antibacterial activity demonstrated that compounds 20c, 20d and 20f displayed not only the most potent
activity against S. aureus ATCC25923 with the MIC values of 0.5, 0.5 and 0.5 µg/mL, but also greatly improved
activity against B. subtilis ATCC9372 with the MIC values of less than or equal to 0.25, 0.25 and 0.25 µg/mL,
respectively. In particular, compound 11g exhibited the strongest antibacterial effectiveness against all the tested
resistant bacterial strains and had well balanced activity with the MIC values of 4–8 µg/mL. Further study on
minimum bactericidal concentration and kinetics confirmed that compound 11g possessed a bacteriostatic effect
on bacterial proliferation. Moreover, the results of molecular docking revealed an potential additional binding
force between compound 11g and U790 in addition to the normal binding force of macrolide skeleton, which
may explain why this compound performed the most potent activity against resistant bacteria. The results of
cytotoxic assay indicated that compounds 20c, 20d and 20f were non-toxic to human breast cancer MCF-7 cells
at its effective antibacterial concentration.
Homo-aza-clarithromycin derivatives
Synthesis
Antibacterial activity
Structure-activity relationships
1. Introduction
importantly, macrolide antibiotics have always been a hot antibacterial
field for scientists.
Development of potent antibacterial agents is a permanently new
and unremitting investigation in the therapeutic field. Still, researchers
want to find the best antibacterial agent. Recently, some novel hetero-
cyclic compounds such as pyranopyrazoles, tri-substituted pyrazole,
dihydropyridines and bis(indolyl)methanes derivatives were designed
and synthesized and evaluated for their biological activities. The results
showed that representative compounds exhibited not only significantly
potent antibacterial activity against Bacillus subtilis ATCC6633, Escher-
ichia coli ATCC8739 and Proteus vulgaris ATCC29213, but also excellent
antifungal activity against Aspergillus niger MTCC1881 [1–5]. More
Macrolide antibiotics produced by Streptomyces, are a kind of weak
alkaline compounds which contain a fourteen or sixteen-membered
lactone ring. They have similar antibacterial spectrum and activity
without cross-resistance with other commonly used antibiotics. Thus,
macrolide antibiotics have become one of the most widely used oral
antibacterial drugs in the field of anti-infective therapy [6]. Erythro-
mycin (Fig. 1) is the first macrolide antibiotic, which was approved by
the US Food and Drug Administration for the treatment of respiratory
tract infections caused by Gram-positive bacteria in the 1950s [7].
Erythromycin has excellent antibacterial activity and low toxicity
* Corresponding author at: Department of Medicinal Chemistry, School of Pharmaceutical Sciences, Shandong University, 44, West Culture Road, Jinan 250012, PR
China.
E-mail address: mashutao@sdu.edu.cn (S. Ma).
https://doi.org/10.1016/j.bioorg.2021.104992
Received 13 February 2021; Received in revised form 12 April 2021; Accepted 11 May 2021
Available online 18 May 2021
0045-2068/© 2021 Elsevier Inc. All rights reserved.

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
clinically, but it is hindered by its narrow antibacterial spectrum and
poor stability in gastric acid [8]. Thus, second generation macrolide
antibiotics have been developed to overcome those shortcomings, which
are semisynthetic derivatives of erythromycin, such as clarithromycin
(CAM), azithromycin (AZM) and roxithromycin (Fig. 1). Their drug
properties are more greatly enhanced than those of erythromycin,
including higher acid stability, stronger antibacterial activity, wider
antibacterial spectrum, better pharmacokinetic properties and fewer
gastrointestinal side effects [9–12]. Consequently, these second gener-
ation macrolide antibiotics have replaced erythromycin as first-line
treatment for respiratory tract infections, and especially, have played
an important role in the treatment of mycoplasma and chlamydia
pneumonia and soft tissue infections. However, with the widespread use
of second generation macrolide antibiotics in clinical practice, some
pathogens have gradually developed resistance to them, which has
brought great difficulties to the treatment of the above infectious dis-
eases. Thus, it is imperative to strengthen the exploration of new gen-
eration of macrolide antibiotics [13]. Telithromycin, cethromycin and
solithromycin (Fig. 1) are the representative drugs of third generation
macrolide antibiotics, which exhibit excellent antibacterial activity
against methicillin-resistant Staphylococcus aureus and multi-drug
resistant Streptococcus pneumoniae [14–16]. In particular, their anti-
bacterial activity against erythromycin-resistant bacteria is much
stronger than that of the first two generations of macrolide antibiotics
[17,18], which is attributed to their aralkyl side chains in structure,
enhancing the binding force of the third generation macrolide antibi-
otics for ribosomes of resistant bacteria [19–23]. With their increasing
clinical application, third generation macrolide antibiotics still face the
serious problems of hepatotoxicity and bacterial resistance. Therefore, it
is necessary to strive to discover newer chemical entities of macrolide
antibiotics with potent activity against resistant bacteria.
and have made positive progress. For example, the methods for mac-
rolactone skeleton reconstruction have been established by Tomoaki
Miura [28] and Tomohiro Sugimoto [29].
In order to find new macrolide derivatives with novel macrolactone
skeletons, 15-membered 11a-homo-aza-clarithromycin derivatives were
designed and synthesized using the above synthetic methods in our
previous study. Among which, some derivatives exhibited excellent
antibacterial activity against sensitive bacteria [30]. For example,
38bpre, 38lpre and 38vpre displayed the strongest activity against
S. aureus ATCC25923 and Bacillus subtilis ATCC9372 with the minimum
inhibitory concentration (MIC) values 0.25 µg/mL, which were better
than the others, and also better than (or equal to) CAM. Moreover, the
antibacterial activity of 38dpre and 38fpre against most sensitive bac-
teria was comparable to that of CAM. For instance, they exhibited
excellent antibacterial activity against B. subtilis ATCC9372 with an MIC
value of 0.25 µg/mL. As for the activity against the four resistant bac-
teria strains of S. aureus ATCC31007, S. aureus ATCC43300,
S. pneumoniae AB11 and S. pyogenes 2, 9epre (Fig. 2), which is C13
substituted 15-membered 11a-homo-aza-clarithromycin derivative with
4-butylphenyl-1H-1,2,3-triazole linked to the C13 position of the 15-
membered lactone by a methylene, displayed 16–32 fold improvement
over CAM. 23epre showed greatly improved activity against S. aureus
ATCC31007, S. pneumoniae AB11 and S. pyogenes 2, which is C12
substituted 15-membered 11a-homo-aza-clarithromycin, possessing the
same side chain at the C12 position of the 15-membered lactone to 9epre
.
However, the replacement of n-butyl group at benzene ring by other
groups resulted in poor antibacterial activity.
The computational analyses of 9epre and CAM with the crystal
structure of the Escherichia coli ribosome (PDB code: 4V7S [31]) by
In recent years, many new macrolide derivatives have been obtained
by the structural modification of macrolides to improve antibacterial
activity against various bacterial strains [24]. For instance, novel
carbonate-triazole derivatives of AZM and N-alkylammonium bromides
of AZM and CAM were designed, synthesized by Anna Janas et al. and
their biological studies revealed potential antibacterial activity of qua-
ternary N-alkylammonium bromides of CAM as compared to AZM [25].
It is reported that macrolide derivatives provided by structural modifi-
cations at the C13 position of erythromycin are very active against
resistant bacteria [26,27]. However, these modifications are blocked by
the lack of chemical reaction activity at the C13 position. Nevertheless,
researchers have been trying their best to overcome these difficulties
Fig. 2. Structure of compound 9epre and its essential group against resis-
tant bacteria.
Fig. 1. Structures of representative macrolides.
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Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
autodock indicated that the 4-n-butylphenyl-1H-1,2,3-triazole side
C12 position, and macrolactonization (Fig. 5). In fact, in addition to the
different introduction position and extension direction of 4-alkylphenyl-
1H-1,2,3-triazole side chains, the design ideas of Series B and C are
similar to that of Series A, which are to explore the possible secondary
force produced by the introduced side chain and nucleotide binding sites
of 23S rRNA.
chain can extend to the bacterial ribosomal nucleotide residue U790,
may generate π-π stacking, electrostatic or hydrophobic interaction with
U2609, A752 and U790 (Fig. 3a) [19,32]. There is a strong hydrogen
bond interaction of the 4′’–OH of 9epre and G2505. In addition, the
conformation of 15-membered macrolactone skeleton has a certain de-
gree of change compared with that of CAM (Fig. 3b). In particular, the 4-
n-butylphenyl-1H-1,2,3-triazole side chain at the C13 position may
produce additional hydrophobic interaction with U790 in the binding
site of the ribosome, leading to dramatically increased activity against
resistant bacteria strains.
2. Chemistry
2.1. Synthesis of N11-substituted 15-membered homo-aza-clarithromycin
derivatives (Series A)
The above findings encourage us continue to explore the effects of
introducing 4-n-butylphenyl-1H-1,2,3-triazole side chains into other
positions of 15-membered macrolactone skeleton, such as N11 position,
on activity against resistant bacteria. Thus, N11-substituted 15-
membered homoclarithromycin derivatives (Series A) were designed
and synthesized by introduction of 4-alkylphenyl-1H-1,2,3-triazole side
chains into the N11 position of 15-membered macrolactone skeleton
(Fig. 4). The design ideas of Series A are as follows: (1) Changing the
length (extension or shortening) and types (non-oxygen-containing or
oxygen-containing alkyl group, and branched or straight alkyl group,
etc.) of the alkyl group on the benzene ring, which may approach U790
The synthetic route of N11-substituted 15-membered homo-aza-
clarithromycin derivatives 11a-11 m (Series A) from CAM 1 is shown
in Scheme 1. CAM 1 was reduced in the presence of NaBH4 to yield 9(S)-
dihydroclarithromycin 2 [34], which was then subjected to selective
protection of 2 with TESCl to give TES ether 3 [35]. After conversion of 3
into the corresponding acyclic aldehyde 4 through oxidation reaction,
which was treated with ethanolamine 5 in the aid of NaBH(OAc)3 to
obtain the secondary amine 6. Alkylation of 6 produced the corre-
sponding tertiary amines 8a-8m by reductive amination of the second-
ary amine with various substituted aldehydes 7a-7m [36].
Saponification of the ester groups of 8a-8m was carried out in the
presence of the base catalyst LiOH to form carboxylic acids 9a-9m in
good yields. Macrocyclization [37] of 9a-9m afforded TES-protected
N11-substituted 15-membered homo-aza-clarithromycin derivatives
10a-10m, which were followed by deprotection of the silane groups
from 10a-10m to provide the target compounds 11a-11m (Series A).
Various substituted aldehydes 7a-7m were prepared as key in-
termediates for the synthesis of Series A (Scheme 2). Conversion of 3-
bromopropanol 12 into azide product 13 through substitution reaction
with NaN3 [38], which was followed by treatment with various
substituted phenylacetylenes 14 to give the corresponding alcohols 15
[39]. Finally, 15 was oxidized with Dess-Martin periodinane to afford
the desired aldehydes 7a-7m in excellent yields [40].
of 23S rRNA in bacterial ribosome and produce stronger π-π stacking,
electrostatic or hydrophobic interaction with it; (2) Altering position of
introduced 4-alkyllphenyl-1H-1,2,3-triazole side chain is to generate
stronger
π-π stacking, electrostatic or hydrophobic interaction with
U790; (3) Besides, the 2′-OH of 15-membered macrolactone skeleton
may still bind to A2058 and A2059 of 23S rRNA of bacterial ribosome
well, producing activity against sensitive bacteria [33].
In our previous study [30], only 9epre and 9fpre with 4-alkyllphenyl-
1H-1,2,3-triazole side chains at the C13 position, and 23epre and 23fpre
with 4-alkyllphenyl-1H-1,2,3-triazole side chains at the C12 position
were explored. However, it is still unclear how the length (extension or
shortening) and types (non-oxygen-containing or oxygen-containing
alkyl groups, and branched or straight alkyl group, etc.) of the alkyl
group on the benzene ring affect activity against sensitive and resistant
and bacteria. In addition, the effect of the configuration (R or S) of C13
and C12 on antibacterial activity was further investigated. In order to
clarify the above problems, C13-substituted 15-membered homo-aza-
clarithromycin derivatives (Series B) and C12-substituted 15-membered
homo-aza-clarithromycin derivatives (Series C) were designed and
synthesized through opening of CAM lactone ring, introduction of 4-
alkylphenyl-1H-1,2,3-triazole side chains into the C13 position and the
2.2. Synthesis of C13-substituted 15-membered homo-aza-clarithromycin
derivatives (Series B)
The 15-membered C13-substituted homo-aza-clarithromycin de-
rivatives 20a-20l (Scheme 3) were synthesized from 4 by using the
similar cyclization process as previously described [30].
The synthetic route of intermediates 16a-16l for the synthesis of
Fig. 3. (a) Model of compound 9epre docked into E. coli ribosome (PDB code: 4V7S [31]). In the model, the predicted hydrogen bonds between compound 9epre and
the nucleotide residues are indicated by dotted lines; (b) The docking model of compound 9epre (green) and CAM (yellow) in E. coli ribosome. Both are located in
approximately the same spatial area in E. coli ribosome. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of
this article.)
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Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
Fig. 4. Brief design strategy of Series A.
Fig. 5. Brief design of Series B and C.
Series B is shown in Scheme 4. Conversion of R-epichlorohydrin 21 into
the corresponding azide 22 through azidation [41] was followed by
treatment with corresponding substituted phenylacetylenes 23 with the
aid of CuSO4 and L-ascorbic acid sodium to afford chloroethanol 24 [39].
Epoxidation of 24 was carried out in the presence of NaOH to form the
epoxide 25 [42], and subsequently, 25 was stereoselectively ring-
opened to provide various substituted aminoalcohols 16a, 16c, 16e,
16g, 16i and 16k [35]. Their enantiomers 16b, 16d, 16f, 16h, 16j and
16l were also prepared from S-epichlorohydrin 26 by applying of the
same synthetic method described above.
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Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
Scheme 1. Regents and conditions: (a) NaBH4,
CH3OH-THF, rt, 24 h, 52%; (b) TESCl, imidazole,
DMF, 0 ◦C to rt, 18 h, 68%; (c) Pb(OAc)4, CHCl3,
0
◦C, 0.5 h; (d) 5, NaBH(OAc)3, CHCl3, rt, 4 h,
72% in two steps from 3; (e) 7a-7m, NaBH
(OAc)3, CHCl3, rt, 12 h; (f) LiOH, THF-C2H5OH-
H2O, rt, 6 h, 18–70% in two steps from 6; (g) i) 2,
4, 6-trichlorobenzoyl chloride, Et3N, THF, rt, 4 h;
ii) DMAP, CH3CN, reflux, 0.5 h, 42–82%; (h)
Hydrogen fluoride-pyridine (65% in hydrofluoric
acid), THF, rt, 18 h, 32–80%.
2.3. Synthesis of C12-substituted 15-membered homo-aza-clarithromycin
derivatives (Series C)
starting material.
3. Results and discussion
The 15-membered C12 substituted homo-aza-clarithromycin de-
rivatives 34a-34l (Scheme 5) were synthesized from 4 by using the
similar cyclization process as previously described [30].
3.1. In vitro antibacterial activity of N11-, C12- and C13-substituted
homo-aza-clarithromycin derivatives
The synthetic route of intermediates 30a-30l for the synthesis of
Series C is outlined in Scheme 6. The Boc-L-serine methyl ester 35 was
derivatived into azide product 37 by methanesulfylation with MsCl, and
then nucleophilic substitution with NaN3 [43]. Cyclization of 37 with
substituted phenylacetylenes 23 produced triazole product 38 [39],
which was followed by reduction with NaBH4 to give alcohol 39 [44].
Finally, deprotection of 39 under the conditions of concentrated hy-
drochloric acid formed the various substituted aminoalcohols 30a, 30c,
30e, 30g, 30i and 30k. Similarly, their enantiomers 30b, 30d, 30f, 30h,
30j and 30l were prepared from Boc-D-serine methyl ester 40 as a
The in vitro antibacterial activity against various sensitive and
resistant bacterial strains were reported as minimum inhibitory con-
centration (MIC) that were determined by the broth microdilution
method [45]. The tested sensitive bacterial strains included three
different penicillin-susceptible strains (E. coli ATCC25922, P. aeruginosa
ATCC27853 and B. subtilis ATCC9372) and one erythromycin-
susceptible strain (S. aureus ATCC25923). In addition, a variety of
resistant bacterial strains were included in the tests in order to identify
the potential N11-, C12- and C13-substituted 15-membered homo-aza-
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Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
Scheme 2. Regents and conditions: (a) NaN3, H2O, 80 ◦C, 24 h, 88%; (b) 14, CuSO4, L-ascorbic acid sodium salt, MeOH-H2O, 40 ◦C, 24 h, 51–98%; (c) Dess-Martin
periodinane, DCM, rt, 1 h, 62–91%.
clarithromycin derivatives that could overcome bacterial resistance.
Three different erythromycin-resistant strains (S. pneumoniae B1,
S. pneumoniae AB11 and S. pyogenes 1), one penicillin-resistant strain
(S. aureus ATCC31007) and one methicillin-resistant strain (S. aureus
ATCC43300) were also tested for the antibacterial activity of com-
pounds. CAM and AZM were used as the reference compounds in vitro
antibacterial activity assays.
11l) or 2-pyridine-1H-1,2,3-triazole (11m) at the N11 position. We also
observed that their lipid/water partition coefficients (ClogP) values did
not affect the anti-sensitive bacteria activity of Series A.
On the other hand, the SARs of Series A against resistant bacteria are
also summarized as follows. The substituent on the 4-phenyl-1H-1,2,3-
triazole and its length is critical to antibacterial activity as shown by
11a-11g vs 11h-11m. Introduction of an alkyl chain into the 4-phenyl-
1H-1,2,3-triazole at the N11 position (11a-11g) leads to dramatically
increased activity against resistant strains. For example, the ClogP
values of 11a-11g generally increased with the extension of alkyl chain
on the 4-phenyl-1H-1,2,3-triazole, while their activity against
S. pneumoniae AB11 was gradually enhanced (Fig. S1, see Supplemen-
tary Data). Obviously, 11g possessed the most significant activity
against resistant bacteria with a balanced spectrum. Those findings
clearly indicate that the alkyl chains with appropriate length and flex-
3.1.1. In vitro antibacterial activity of Series A (11a-11m)
The antibacterial activity of Series A is shown in Table 1. In general,
the compounds of Series A were inactive against E. coli ATCC25922 and
P. aeruginosa ATCC27853. In contrast, the compounds of Series A were
observed to exhibit excellent antibacterial activity against S. aureus
ATCC25923 (MIC = 1–8 µg/mL) and B. subtilis ATCC9372 (MIC = 1–4
µg/mL). Among them, 11b showed the most potent activity against
S. aureus ATCC25923 (MIC = 1 µg/mL) and B. subtilis ATCC9372 (MIC =
1 µg/mL). As for activity against the five resistant bacterial strains
(S. aureus ATCC31007, S. aureus ATCC43300, S. pneumoniae B1,
S. pneumoniae AB11 and S. pyogenes 1), 11f and 11g were found to
display a substantial improvement (a 16–64 fold increase) over CAM.
Moreover, 11c, 11d, 11e, 11j, 11k and 11l were also more active than
CAM against the above five resistant bacterial strains. In particular, 11g
exerted the strongest antibacterial effectiveness against the above five
resistant bacterial strains with balanced activity (MIC = 4 ~ 8 µg/mL).
Based on the MIC data mentioned above in our study, and the
structure–activity relationships (SARs) of Series A against sensitive
bacterial strains are summarized as follows. Introduction of 4-
substituted phenyl-1H-1,2,3-triazole or 2-pyridine-1H-1,2,3-triazole
side chain at the N11 position results in a very similar profile in activity
against S. aureus ATCC25923 and B. subtilis ATCC9372. 11a-11c and
11h-11m were slightly less active than CAM against S. aureus
ATCC25923 and B. subtilis ATCC9372, but the antibacterial activity of
these two kinds of compounds is similar. Actually, introduction of the 4-
substituted phenyl-1H-1,2,3-triazole side chain at the N11 position may
not affect the binding ability of 2′-OH to A2058 and A2059 in domain V
of 23S rRNA of bacterial ribosome [33]. In contrast, loss of activity
against E. coli ATCC25922 and P. aeruginosa ATCC27853 was observed
for introduction of various 4-substituted phenyl-1H-1,2,3-triazole (11a-
ibility may easy generate the π-π stacking, electrostatic or hydrophobic
interaction with U790 of 23S rRNA [19,32]. We infer that the
enhancement of the antibacterial activity achieved by introducing the
alkyl group is mainly due to production of a secondary interaction with
the binding sites of 23S rRNA. Besides, 11h-11j had similar trend in
antibacterial activity to 11a-11g. Introduction of halogen group into the
4-phenyl-1H-1,2,3-triazole (11k and 11l) results in very potent activity
against resistant bacteria (MIC ranges from 16 to 128 µg/mL). It may be
because that introduction of halogen group can improve the lip-
osolubility of compounds, which may explain why 11k and 11l have the
desired profile of antibacterial activity.
3.1.2. In vitro antibacterial activity of Series B (20a-20l)
The antibacterial activity of Series B is shown in Table 2. Most
compounds in series B were very effective against S. aureus ATCC25923
(MIC = 0.5–2 µg/mL) and B. subtilis ATCC9372 (MIC = 0.25–1 µg/mL).
Among them, 20c, 20d and 20f exhibited not only the most potent ac-
tivity against S. aureus ATCC25923 (MIC = 0.5, 0.5 and 0.5 µg/mL), but
also the strongest activity against B. subtilis ATCC9372 (MIC ≤ 0.25,
0.25 and 0.25 µg/mL). Interestingly, all compounds of Series B possessed
improved activity against the five resistant bacterial strains of S. aureus
ATCC31007, S. aureus ATCC43300, S. pneumoniae B1, S. pneumoniae
AB11 and S. pyogenes 1 compared with CAM and AZM. In particular, 20g
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Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
Scheme 3. Regents and conditions: (a) 16a-16l, NaBH(OAc)3, CHCl3, rt, 4 h; (b) HCHO, NaBH(OAc)3, CHCl3, rt, 4 h; (c) LiOH, THF-C2H5OH-H2O, rt, 6 h, 27–88% in
four steps from 3; (d) i) 2, 4, 6-trichlorobenzoyl chloride, Et3N, THF, rt, 4 h; ii) DMAP, CH3CN, reflux, 0.5 h, 41–89%; (e) Hydrogen fluoride-pyridine (65% in
hydrofluoric acid), THF, rt, 18 h, 24–63%.
and 20h showed the most greatly improved activity (4–8 µg/mL) against
the above five resistant bacterial strains, which was 16–32-fold stronger
than those of CAM and AZM, while 20c, 20e, 20f and 20l exhibited
moderately enhanced activity as compared to CAM. However, all com-
pounds of Series B completely abolished the activity against E. coli
ATCC25922 and P. aeruginosa ATCC27853.
better activity than S- isomer against S. aureus ATCC25923 and
B. subtilis ATCC9372 (The order of activity: 20f > 20e, 20h > 20g, 20j
> 20i and 20l > 20k). Thus, the R-configuration of C13 has favorable
effects on antibacterial activity.
On the other hand, introduction of an alkyl group into the 4-phenyl-
1H-1,2,3-triazole at the C13 position greatly increased activity against
resistant strains (20a, 20c, 20e and 20g). Moreover, we also observed
that the longer the alkyl group was, the higher the ClogP value was, and
the stronger its antibacterial activity was. For instance, 20g with n-amyl
group had the most potent activity against S. pneumoniae AB11 (The
order of activity: 20g > 20e > 20c > 20a) (Fig. S2, see Supplementary
Data). The above findings may indicate that the length and flexibility of
the alkyl group has a dramatic effect on the binding force of compounds
(20a, 20c, 20e and 20g) for new binding site U790 [19,32]. Similarly,
the same trend was also observed for 20b, 20d, 20f and 20h (Fig. S3, see
Supplementary Data). Besides, the activity of the compounds with R-
configuration at the C13 position was essentially equivalent to that of
The SARs of Series B against sensitive bacterial strains have been
systematically summarized as follows. Introduction of the 4-substituted
phenyl-1H-1,2,3-triazole side chain at the C13 position can significantly
improve antibacterial activity against S. aureus ATCC25923 and
B. subtilis ATCC9372, which was illustrated by the examples of 20a-20h
with the MIC values of 1–8
μg/mL. In a remarkable contrast, this
modification is not necessary for activity against E. coli ATCC25922 and
P. aeruginosa ATCC27853, which was confirmed by the fact that the MIC
values of 20a-20l are more than 32 μg/mL. The more important concern
is that C13 is chirality, which results in the compounds of Series B with
R- and S-configurations. R-isomer, in general, exhibited 8- to 64-fold
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Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
Scheme 4. Regents and conditions: (a) NaN3, AcOH-H2O, 30 ◦C, 5 h, 86–90%; (b) 23, CuSO4, L-ascorbic acid sodium salt, CH3OH-H2O, 35–40 ◦C, 24 h, 62–97%; (c)
NaOH, CH3CN-H2O, rt, 2 h; (d) 25% NH3⋅H2O, CH3CN, rt, 3 h, 41–76% in two steps from 24 or 28.
the compounds with S-configuration against resistant bacteria (The
order of activity: 20a = 20b, 20c ≈ 20d, 20e ≈ 20f, 20g ≈ 20h, 20i =
20j and 20k ≈ 20l), which reveals that the chirality of C13 has no in-
fluence on activity against resistant bacteria.
ATCC9372 (The order of activity: 34e > 34f, 34i > 34j and 34k > 34l).
On the other hand, comparison of the activity of 34a, 34c, 34e and
34g against resistant bacteria with the reference CAM reveals that the
order of activity of the above compounds against S. pneumoniae AB11 is
the 34g > 34e ≈ 34c > 34a, which displays that the length of the
introduced alkyl group is beneficial to the activity against S. pneumoniae
AB11 (Fig. S4, see Supplementary Data). That may be because the
suitable length and flexibility of the alkyl group are good for tightly
interacting with the new binding site U790 [19,32]. Besides, 34b, 34d,
34f and 34h possess the similar trend in antibacterial activity to 34a,
34c, 34e and 34g (Fig. S5, see Supplementary Data). The relationships
between ClogP values and the activity of 34a-34h against resistant
bacteria revealed that the higher values of ClogP (>6) of compound led
to its better antibacterial activity. It is worth noting that isomers 34a and
34b exhibited same activity against resistant bacteria, which indicates
that the configuration of C12 position is does not affect the antibacterial
activity. Similarly, the conclusion also applies to 34c and 34d, 34e and
34f, 34g and 34h. However, the antibacterial activity of compounds
with alkoxy group is not different from that of the above compounds. For
example, 34i with alkoxy group displayed a 2–4-fold improvement in
antibacterial activity as compared to its epimer 34j against resistant
bacteria, and 34k with alkoxy group exhibited increased antibacterial
activity as compared to its epimer 34l as well.
3.1.3. In vitro antibacterial activity of Series C (34a-34l)
The antibacterial activity of Series C is shown in Table 3. In this
series, 34h, 34i and 34k were found to possess potent activity against
B. subtilis ATCC9372 with the MIC values of 1, 1 and 0.5 µg/mL,
respectively, which were fundamentally equivalent to that CAM. They
also showed activity against S. aureus ATCC25923 with the MIC values
of 4, 2 and 1 µg/mL, respectively, but were slightly less active than CAM.
Especially, 34k exhibited the most potent and balanced activity against
both S. aureus ATCC25923 and B. subtilis ATCC9372. 34g and 34h had
excellent activity against the five resistant bacterial strains of S. aureus
ATCC31007, S. aureus ATCC43300, S. pneumoniae B1, S. pneumoniae
AB11 and S. pyogenes 1, which were 16- to 64-fold more active than
those of CAM. In addition, 34c, 34d, 34e and 34k had a moderate ac-
tivity against the above resistant bacterial strains, which were generally
4- to 16-fold better than those of CAM. Among them, 34g exhibited the
most potent and balanced activity against with the MIC values of 4–8 µg/
mL. However, the compounds of Series C (34a-34l) exhibited poor ac-
tivity against E. coli ATCC25922 and P. aeruginosa ATCC27853.
According to the data of antibacterial activity, the SARs of Series C
against sensitive strains were summarized in detail. Introduction of a 4-
substituted phenyl-1H-1,2,3-triazole side chain at the C12 position fails
to improve activity against E. coli ATCC25922 and P. aeruginosa
ATCC27853. In contrast, moderate improvements in activity against
S. aureus ATCC25923 and B. subtilis ATCC9372 were observed for
introduction of the above side chain. Similar to Series B, introduction of
a 4-substituted phenyl-1H-1,2,3-triazole side chain at the C12 position
leads to the chirality of C12, thereby producing the compounds with R-
and S-configurations. As for their antibacterial activity, the compounds
with S-configuration was in general 8- to 64-fold better than the com-
pounds with R-configuration against S. aureus ATCC25923 and B. subtilis
3.1.4. Overall SARs of Series A, B and C
Overall, in the antibacterial activity against susceptible bacteria, the
activity of these three series is B > A > C. This indicates that antibac-
terial activity depends on the position of the 4-substituted phenyl-1H-
1,2,3-triazole side chain attached to the macrolide skeleton. For
example, introduction of the 4-substituted phenyl-1H-1,2,3-triazole side
chain at the C13 position had the greatest effect on the antibacterial
activity, followed by that at the N11 position and the worst at the C12
position. The above findings may show that introduction of the appro-
priate side chains at the C13 position has little effect on the binding of 2′-
OH to A2058 and A2059 in domain V of 23S rRNA [33]. On the other
8

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
Scheme 5. Regents and conditions: (a) 30a-30l, NaBH(OAc)3, CHCl3, rt, 4 h; (b) HCHO, NaBH(OAc)3, CHCl3, rt, 4 h; (c) LiOH, THF-C2H5OH-H2O, rt, 6 h, 15–77% in
four steps from 3; (d) i) 2, 4, 6-trichlorobenzoyl chloride, Et3N, THF, rt, 4 h; ii) DMAP, CH3CN, reflux, 0.5 h, 22–81%; (e) Hydrogen fluoride-pyridine (65% in
hydrofluoric acid), THF, rt, 18 h, 24–83%.
hand, Series A, B and C displayed similar activity against resistant
bacteria, which reveals that the introduction position of the 4-
substituted phenyl-1H-1,2,3-triazole side chain is not related to the
antibacterial activity. It is worth noting that introduction of 4-n-amyl-
phenyl-1H-1,2,3-triazole side chain could lead to the strongest activity
against resistant bacteria with a balanced spectrum (11g, 20g, 20h, 34g
and 34h). This may be due to n-amyl group with the most suitable length
and flexibility for interaction with U790 [19,32]. Besides, it also might
be explained by the possibility that higher lipophilic compounds were
easily transported to the binding sites of microorganism. Therefore, we
can draw the important conclusions as follows: (1) When introduction of
an alkyl group into phenyl-1H-1,2,3-triazole side chain at the N11, C12
or C13 position, the longer the alkyl chain is (5 carbon is the best), the
stronger the activity against resistant bacteria is; (2) The compounds
containing alkyl groups are more active against resistant bacteria than
the compounds containing alkoxy groups; (3) The introduction of 4-
substituted phenyl-1H-1,2,3-triazole side chain at the N11, C12 and
C13 position of 15-membered skeleton can effectively enhance the ac-
tivity against resistant bacteria.
3.2. Molecular docking
In order to investigate the potential binding mode of compounds,
11g and 20c were selected as representative compounds for molecular
docking because they were the most potent compounds against drug-
resistant and susceptible bacteria. Here, the computational analyses of
11g, 20c and CAM were explored utilizing a reported crystal structure of
E. coli ribosome bound to antibiotics (PDB code: 4V7S [31]) and auto-
dock. The molecular docking of 11g and CAM is shown in Fig. 6. We
observed that the macrolide skeleton of 11g and CAM are basically in
the same space (Fig. 6a), while the 4-n-amylphenyl-1H-1,2,3-triazole
side chain of 11g extends to the vicinity of U790 (Fig. 6b). This indicates
9

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
Scheme 6. Regents and conditions: (a) MsCl, DCM, Et3N, 0 ◦C, 0.5 h; (b) NaN3, DMF, 50 ◦C, 0.5 h; (c) 23, CuSO4, L-ascorbic acid sodium salt, CH3OH-H2O, 35–40 ◦C,
48 h, 36–62%; (d) NaBH4, CH3OH, 0 ◦C, 2 h; (e) aq HCl, EtOAc, rt, 4 h, 40–85% in two steps from 38 or 43.
that the side chain may bind to U790 by
π-
π
stacking, electrostatic or
3.3. Bactericidal or bacteriostatic effect
hydrophobic interaction. We also observed that the aralkyl side chains
of 11g and telithromycin had the same spatial extension direction,
which implied that theypossessed the same binding sites (Fig. 6c). In the
same way, the model of 20c and CAM docked into E. coli ribosome (PDB
code: 4V7S [31]) was presented in Fig. 7. The results indicate that the
stereo conformation of 15-membered homo-aza-clarithromycin skeleton
of 20c and the macrocyclic lactone of CAM are located in approximately
the same spatial area in E. coli ribosome (Fig. 7a). In addition, the strong
hydrogen bond interaction of the 2′-OH of 20c with A2058 and A2059
was also observed (Fig. 7b). This may be helpful to explain why 20c has
excellent activity against susceptible bacteria.
The minimal bactericidal concentration (MBC) values of some
promising compounds against susceptible bacteria (S. aureus
ATCC25923) and resistant bacteria (S. pneumoniae AB11) was deter-
mined to confirm whether those promising compounds were bacteri-
cidal or bacteriostatic (Table 4). The results showed that 20c, 20d and
20f possessed the MBC/MIC values of 64–128, which suggests that they
are bacteriostatic agents with regard to S. aureus ATCC25923. In
contrast, the MBC/MIC values of 16 were observed for 11f, 11g, 20g,
20h and 34g, which indicates that they exhibit bacteriostatic effect
against S. pneumoniae AB11. In short, the above compounds exhibit
potent activity against S. aureus ATCC25923 and S. pneumoniae AB11 by
bacteriostatic mechanism.
10

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
Table 1
In vitro antibacterial activity and ClogP of Series A (11a-11m).
Minimum inhibitory concentration / MIC (µg/mL)
Compound
S. aureus
E. coli
P. aeruginosa
B. subtilis
S. aureus
S. aureus
S. pneumoniae
S. pneumoniae
S. pyogenes
ClogPj
ATCC25923a
ATCC25922b
ATCC27853c
ATCC9372d
ATCC31007e
ATCC43300f
B1g
AB11h
1i
11a
11b
11c
11d
11e
11f
2
>128
>128
>128
>128
>128
>128
>128
>128
>128
>128
>128
>128
>128
64
128
128
128
128
128
128
128
128
128
128
128
128
128
16
1
256
64
256
64
64
32
16
8
128
128
32
64
32
16
16
8
3.79
4.28
4.94
5.47
5.61
6.00
6.53
3.90
4.43
4.96
3.97
4.54
2.67
2.64
2.37
1
1
2
1
32
32
4
2
16
16
32
8
2
16
8
16
8
16
8
2
8
8
8
8
11g
11h
11i
8
4
4
4
8
4
8
2
1
256
128
32
128
64
64
32
32
32
16
256
>256
>256
256
128
32
64
32
16
32
16
64
256
256
2
2
11j
2
2
32
11k
11l
2
1
64
64
128
32
2
1
32
32
11m
AZM
CAM
2
1
>256
>256
>256
>256
>256
>256
128
256
128
≤0.25
≤0.25
≤0.25
≤0.25
128
32
a
S. aureus ATCC25923: Staphylococcus aureus ATCC25923, erythromycin-susceptible strain;
E. coli ATCC25922: Escherichia coli ATCC25922, penicillin-susceptible strain;
b
c
d
e
f
P. aeruginosa ATCC27853: Pseudomonas aeruginosa ATCC27853, penicillin-susceptible strain, not characterized;
B. subtilis ATCC9372: Bacillus subtilis ATCC9372, penicillin-susceptible strain;
S. aureus ATCC31007: Staphylococcus aureus ATCC31007, penicillin-resistant strain;
S. aureus ATCC43300: Staphylococcus aureus ATCC43300, methicillin-resistant strain;
g
h
i
S. pneumoniae B1: Streptococcus pneumoniae B1, erythromycin-resistant strain expressing the ermB gene;
S. pneumoniae AB11: Streptococcus pneumoniae AB11, erythromycin-resistant strain expressing the ermB and mefA genes;
S. pyogenes 1: Streptococcus pyogenes 1, erythromycin-resistant strain isolated clinically;
j
ClogP: The theoretically calculated values of ClogP for all the target compounds were calculated using SYBYL-X 2.1.1.
Table 2
In vitro antibacterial activity and ClogP of Series B (20a-20 l).
Minimum inhibitory concentration / MIC (µg/mL)
Compound
S. aureus
E. coli
P. aeruginosa
B. subtilis
S. aureus
S. aureus
S. pneumoniae
S. pneumoniae
S. pyogenes
ClogPj
ATCC25923a
ATCC25922b
ATCC27853c
ATCC9372d
ATCC31007e
ATCC43300f
B1g
AB11h
1i
20a
20b
20c
20d
20e
20f
2
64
128
64
1
64
64
64
32
64
8
64
64
32
64
16
32
8
64
64
32
64
16
32
4
64
64
32
64
16
16
8
3.97
3.97
4.50
4.50
5.03
5.03
6.09
6.09
3.99
3.99
4.52
4.52
2.64
2.37
1
32
1
64
0.5
0.5
8
128
32
128
64
≤0.25
≤0.25
2
32
128
16
>128
32
128
64
0.5
8
≤0.25
8
32
32
8
20g
20h
20i
>128
>128
>128
64
128
>128
128
64
8
1
1
8
8
8
8
8
32
2
64
64
64
64
32
32
>256
>256
64
64
64
64
>256
>256
64
64
32
32
256
128
64
64
32
32
256
256
20j
1
64
20k
20l
16
1
>128
64
128
64
4
64
0.5
32
AZM
CAM
≤0.25
≤0.25
64
16
≤0.25
≤0.25
>256
>256
128
32
a
S. aureus ATCC25923: Staphylococcus aureus ATCC25923, erythromycin-susceptible strain;
E. coli ATCC25922: Escherichia coli ATCC25922, penicillin-susceptible strain;
b
c
d
e
f
P. aeruginosa ATCC27853: Pseudomonas aeruginosa ATCC27853, penicillin-susceptible strain, not characterized;
B. subtilis ATCC9372: Bacillus subtilis ATCC9372, penicillin-susceptible strain;
S. aureus ATCC31007: Staphylococcus aureus ATCC31007, penicillin-resistant strain;
S. aureus ATCC43300: Staphylococcus aureus ATCC43300, methicillin-resistant strain;
g
h
i
S. pneumoniae B1: Streptococcus pneumoniae B1, erythromycin-resistant strain expressing the ermB gene;
S. pneumoniae AB11: Streptococcus pneumoniae AB11, erythromycin-resistant strain expressing the ermB and mefA genes;
S. pyogenes 1: Streptococcus pyogenes 1, erythromycin-resistant strain isolated clinically;
j
ClogP: The theoretically calculated values of ClogP for all the target compounds were calculated using SYBYL-X 2.1.1.
3.4. Study on bactericidal kinetics
S. pneumoniae AB11 CFU/mL (0–12 h), respectively. In contrary, this
compound showed excellent bacteriostatic activity as it did not produce
a significant log10 reduction in S. pneumoniae AB11 CFU/mL (12–24 h).
Thus, 11g have a bacteriostatic effect on bacterial proliferation, which is
dependent on its concentration and timing.
Based on the above mentioned MBC/MIC data, 11g was found to
have a certain degree of bactericidal activity. Next, we investigated how
rapidly 11g were able to reduce a high amount of S. pneumoniae AB11
using a time-bactericidal assay (Fig. 8). The results displayed that at the
concentrations of 1, 2 and 4 MIC, this compound exhibited bactericidal
activity as it produced a 2.7 log10, 3.14 log10 and 3.31 log10 reduction in
11

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
Table 3
In vitro antibacterial activity and ClogP of Series C (34a-34 l).
Minimum inhibitory concentration / MIC (µg/mL)
Compound
S. aureus
E. coli
P. aeruginosa
B. subtilis
S. aureus
S. aureus
S. pneumoniae
S. pneumoniae
S. pyogenes
ClogPj
ATCC25923a
ATCC25922b
ATCC27853c
ATCC9372d
ATCC31007e
ATCC43300f
B1g
AB11h
1i
34a
34b
34c
34d
34e
34f
16
16
16
32
8
>128
>128
>128
>128
>128
>128
>128
>128
>128
>128
128
128
128
128
128
128
128
128
128
128
128
64
8
128
128
32
128
128
32
128
128
32
64
64
64
16
16
32
16
8
3.97
3.97
4.50
4.50
5.03
5.03
6.09
6.09
3.99
3.99
4.51
4.51
2.64
2.37
16
128
32
8
8
32
32
32
32
4
16
32
32
32
64
8
32
128
8
64
64
32
34g
34h
34i
4
4
8
8
4
1
8
8
8
8
8
2
1
64
64
64
64
64
128
32
32
256
256
34j
64
1
32
256
32
256
32
256
32
128
32
34k
34l
0.5
16
64
≤0.25
≤0.25
>128
64
128
16
128
>256
>256
128
>256
>256
128
>256
>256
128
256
128
AZM
CAM
≤0.25
≤0.25
128
32
a
S. aureus ATCC25923: Staphylococcus aureus ATCC25923, erythromycin-susceptible strain;
E. coli ATCC25922: Escherichia coli ATCC25922, penicillin-susceptible strain;
b
c
d
e
f
P. aeruginosa ATCC27853: Pseudomonas aeruginosa ATCC27853, penicillin-susceptible strain, not characterized;
B. subtilis ATCC9372: Bacillus subtilis ATCC9372, penicillin-susceptible strain;
S. aureus ATCC31007: Staphylococcus aureus ATCC31007, penicillin-resistant strain;
S. aureus ATCC43300: Staphylococcus aureus ATCC43300, methicillin-resistant strain;
g
h
i
S. pneumoniae B1: Streptococcus pneumoniae B1, erythromycin-resistant strain expressing the ermB gene;
S. pneumoniae AB11: Streptococcus pneumoniae AB11, erythromycin-resistant strain expressing the ermB and mefA genes;
S. pyogenes 1: Streptococcus pyogenes 1, erythromycin-resistant strain isolated clinically;
j
ClogP: The theoretically calculated values of ClogP for all the target compounds were calculated using SYBYL-X 2.1.1.
3.5. Cytotoxic assay
5. Experimental section
In the three series, 20c, 20d and 20f were selected for cytotoxic assay
due to their potent activity against S. aureus ATCC25923 and B. subtilis
ATCC9372 with the MIC values in the low concentration range
(0.25–0.5 µg/mL), which is equivalent to that of CAM. The 3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method
was used to evaluate the cytotoxicity of those compounds to MCF-7
breast cancer cells (Fig. 9). The result represented that 20c and 20f
were non-toxic to MCF-7 breast cancer cells at the concentration of 8 µg/
mL, because their cell survival rate was still more than 50% at this
concentration, and 20d was non-toxic to MCF-7 breast cancer cells at the
concentration of 16 µg/mL. Therefore, 20c, 20d and 20f exhibit no
cytotoxicity at its effective antibacterial concentration.
5.1. Synthetic procedures
The structures were identified by spectral analysis. 1H NMR and 13
C
NMR spectra were all given on Bruker Avance DRX600 spectrometer
(600 MHz for 1H NMR, 100 or 150 MHz for 13C NMR) and measured in
CDCl3 or d6-DMSO. Mass spectra and high resolution mass spectra were
recorded on the API 4000 instrument and Orbitrap analyzer, respec-
tively. The melting points of compounds were determined on an un-
corrected RY-1 melting point apparatus. All reagents and solvents were
used directly without prior separation and purification, except where
noted. The chemical synthesis reaction was monitored by thin layer
chromatography. Isolation and purification of intermediates and target
compounds were carried out on the column chromatography of silica
gel.
4. Conclusion
Novel N11-, C12- and C13-substituted 15-membered homo-aza-
clarithromycin derivatives against various sensitive and resistant bac-
teria were designed, synthesized and evaluated for their antibacterial
activity. The C13-substituted derivatives 20c, 20d and 20f possessed
both the most potent activity against S. aureus ATCC25923 with the MIC
values of 0.5, 0.5 and 0.5 µg/mL, and B. subtilis ATCC9372 with the MIC
values of less than or equal to 0.25, 0.25 and 0.25 µg/mL, respectively.
In particular, the N11-substituted derivative 11g exhibited the strongest
activity against all the tested resistant bacterial strains with the MIC
values of 4–8 µg/mL, and exerted a bacteriostatic effect on bacterial
proliferation. This may be because it produced an additional binding
force with U790 in addition to the normal binding force of macrolide
skeleton. Moreover, 20c, 20d and 20f were non-toxic to human breast
cancer MCF-7 cells at its effective antibacterial concentration. Taken all
together, 20c, 20d, 20f and 11g become visible as forward-looking and
promising lead compounds for further investigation.
5.1.1. Synthesis of N11-substituted 15-membered homo-aza-clarithromycin
derivatives (Series A)
5.1.1.1. 3-Azidopropan-1-ol (13). 3-Bromo-1-propanol 12 (1 g, 7.25
mmol) in H2O (10 mL) was reacted with NaN3 (0.94 g, 14.5 mmol) at
80 ◦C for 24 h. DCM (20 mL) was added, the organic phase was sepa-
rated and aqueous layer was thoroughly extracted with DCM (20 mL ×
2). The combined organic layers were dried over Na2SO4, filtered and
evaporated in vacuo to yield 3-azidopropan-1-ol 13 (0.64 g, 88%) as a
colorless oil.
5.1.1.1. 3-(4-Phenyl-1H-1,2,3-triazol-1-yl)propan-1-ol (15). 13 (0.5 g,
4.95 mmol) in 75% methanol solution (20 mL) was reacted with phe-
nylacetylene 14 (0.5 g, 4.95 mmol) in the presence of CuSO4 (49 mg)
and L-ascorbic acid sodium salt (147 mg) at 40 ◦C for 24 h. The reaction
mixture was evaporated in vacuo and the residue was separated chro-
matographically (DCM/CH3OH = 30:1) to provide the alcohol 15 (0.8 g,
80%) as a white solid. Mp: 85–88 ◦C, TLC Rf = 0.47 (DCM/CH3OH =
20:1); 1H NMR (600 MHz, CDCl3, δ ppm): 7.84–7.81 (m, 3H), 7.43 (t, J
= 7.7 Hz, 2H), 7.34 (t, J = 7.4 Hz, 1H), 4.58 (t, J = 6.7 Hz, 2H), 3.70 (t, J
= 5.8 Hz, 2H), 2.21–2.17 (m, 2H), 1.72 (s, 1H); MS (ESI) m/z calcd for
C
11H14N3O [M + H]+: 204.1, found: 204.2.
12

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
Fig. 6. (a) Model of compound 11 g (wathet) and CAM (yellow) docked into E. coli ribosome (PDB code: 4V7S [31]). Both are located in approximately the same
spatial area in E. coli ribosome; (b) The docking model of compound 11g (wathet) in E. coli ribosome; (c) Model of compound 11g (wathet) and telithromycin
(reddish brown) docked into E. coli ribosome. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of
this article.)
Fig. 7. (a) Model of compound 20c (navy blue) and CAM (yellow) docked into E. coli ribosome; (b) The docking model of compound 20c (navy blue) in E. coli
ribosome. In the model, the predicted hydrogen bonds between compound 20c and the nucleotide residues are indicated by dotted lines. (For interpretation of the
references to colour in this figure legend, the reader is referred to the web version of this article.)
13

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
NH3⋅H2O = 30:1:0.1 ~ 20:1:0.1 ~ 10:1:0.1) to form the 9(S)–OH-2
(10.5 g, 52%) as a white solid. Mp: 208–210 ◦C, TLC Rf = 0.45 (DCM/
CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR (600 MHz, CDCl3, δ ppm): 5.72
(d, J = 9.8 Hz, 1H), 5.30 (s, 1H), 5.21 (dd, J = 11.2, 2.2 Hz, 1H), 4.96
(dd, J = 20.3, 4.7 Hz, 1H), 4.50 (d, J = 7.1 Hz, 1H), 4.34 (d, J = 1.6 Hz,
1H), 4.05–4.03 (m, 1H), 3.83–3.72 (m, 3H), 3.53–3.45 (m, 3H), 3.37 (s,
3H), 3.33 (s, 3H), 3.29–3.21 (m, 2H), 3.04–2.95 (m, 3H), 2.39–2.33 (m,
7H), 2.19–2.13 (m, 2H), 2.02–1.84 (m, 4H), 1.53–1.44 (m, 4H), 1.38 (s,
3H), 1.32–1.22 (m, 15H), 1.13–1.08 (m, 9H), 0.84 (t, J = 7.4 Hz, 3H);
MS (ESI) m/z calcd for C38H72NO13 [M + H]+: 750.5, found: 750.8.
Table 4
Comparison of MIC and MBC values for some promising compounds against
S. aureus ATCC25923 and S. pneumoniae AB11.
Compound
MBC (µg/mL)
MIC (µg/mL)
MBC/MIC
S. aureus ATCC25923a
20c
20d
20f
64
0.5
128
64
32
0.5
64
0.5
128
≥512
CAM
128
≤0.25
S. pneumoniae AB11b
11f
128
64
8
16
16
16
16
16
>16
1
11g
20g
20h
34g
34h
CAM
4
5.1.1.4. 2′, 4′’, 9(S)-Triethylsilane-6-O-methylerythromycin A (3). To a
solution of 2 (8 g, 10.7 mmol) and imidazole (7.2 g, 105.8 mmol) in
anhydrous DMF (100 mL) was added dropwise TESCl (5.5 g, 36.5 mmol)
at 0 ◦C. The reaction mixture was stirred at room temperature for 18 h
under nitrogen atmosphere. The reaction mixture was quenched by
addition of H2O (100 mL), then EtOAc (100 mL) and n-hexane (100 mL)
were added to this mixture. The organic layer was separated and washed
with saturated NH4Cl solution (100 mL). The organic layer was dried
over anhydrous Na2SO4, filtered and evaporated in vacuo, and the res-
64
4
128
128
>128
128
8
8
8
128
a
S. aureus ATCC25923: Staphylococcus aureus ATCC25923, erythromycin-
susceptible strain;
b
S. pneumoniae AB11: Streptococcus pneumoniae AB11, erythromycin-resistant
strain expressing the ermB and mefA gene.
idue
was
separated
chromatographically
(PE/acetone
=
5.1.1.2. 3-(4-Phenyl-1H-1,2,3-triazol-1-yl)propanal (7a). To a solution
of 15 (0.5 g, 2.46 mmol) in anhydrous DCM (20 mL) was added Dess-
Martin periodinane (1.15 g, 2.71 mmol) at room temperature. After
stirring the reaction mixture for 1 h, the reaction mixture was quenched
with saturated NaHCO3 solution (20 mL). The organic phase was sepa-
rated and aqueous layer was extracted with DCM (20 mL × 2). The
combined organic layers were dried over anhydrous Na2SO4, filtered,
and evaporated in vacuo. The residue was separated chromatographi-
cally (DCM/CH3OH = 30:1) to obtain the aldehyde 7a (0.45 g, 91%) as a
light yellow oil.
5.1.1.3. 9-Dihydro-6-O-methylerythromycin A (2). To a solution of CAM
1 (20 g, 26.8 mmol) dissolved in anhydrous THF (150 mL) and CH3OH
(300 mL) was added NaBH4 (6 g, 158.6 mmol) in three batches within 1
h at 0 ◦C. The reaction mixture was stirred at room temperature for 24 h
under nitrogen atmosphere. The reaction mixture was concentrated in
vacuo to dryness, and then added DCM (200 mL) and saturated NH4Cl
solution (200 mL). The organic phase was separated and aqueous layer
was extracted with DCM (100 mL × 2). The combined organic layers
were dried over anhydrous Na2SO4, filtered and evaporated in vacuo,
and the residue was separated chromatographically (DCM/CH3OH/
Fig. 9. The toxicity assay of compounds 20c, 20d and 20f to MCF-7 breast
cancer cells, and CAM served as a control.
Fig. 8. Bactericidal kinetics of compound 11g against S. pneumoniae AB11.
14

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
80:1–50:1–30:1–10:1) to give the TES ether 3 (7.9 g, 68%) as a white
foam. Mp: 86–89 ◦C, TLC Rf = 0.76 (PE/acetone = 3:1); 1H NMR (600
MHz, CDCl3, δ ppm): 5.08 (dd, J = 11.2, 2.3 Hz, 1H), 4.86 (d, J = 4.6 Hz,
1H), 4.48 (s, 1H), 4.32 (d, J = 7.0 Hz, 1H), 4.26–4.24 (m, 1H), 3.88 (d, J
= 9.7 Hz, 1H), 3.80 (s, 1H), 3.64 (d, J = 9.0 Hz, 1H), 3.51–3.48 (m, 1H),
3.36–3.32 (m, 4H), 3.29 (s, 3H), 3.25–3.21 (m, 2H), 3.12 (dd, J = 9.7,
7.0 Hz, 1H), 2.88–2.86 (m, 1H), 2.54–2.50 (m, 1H), 2.38 (d, J = 14.8 Hz,
1H), 2.19 (s, 6H), 2.10–2.09 (m, 1H), 1.97–1.95 (m, 1H), 1.85–1.82 (m,
1H), 1.68 (d, J = 14.3 Hz, 1H), 1.64–1.61 (m, 1H), 1.52 (d, J = 5.0 Hz,
1H), 1.49–1.46 (m, 4H), 1.29–0.96 (m, 44H), 0.96–0.89 (m, 10H), 0.82
(t, J = 7.4 Hz, 3H), 0.71–0.54 (m, 18H); MS (ESI) m/z calcd for
mL). The organic phase was separated and aqueous layer was extracted
with DCM (20 mL × 2). The combined organic layers were dried over
anhydrous Na2SO4, filtered, and evaporated in vacuo. The residue was
separated chromatographically (PE/acetone = 15:1 ~ 10:1) to obtain
the cyclic intermediate 10a (250 mg, 63%) as a colorless oil. TLC Rf =
0.54 (PE/acetone = 3:1); 1H NMR (600 MHz, CDCl3, δ ppm): 7.87–7.74
(m, 3H), 7.44–7.41 (m, 2H), 7.33 (t, J = 7.4 Hz, 1H), 5.38–5.32 (m, 1H),
4.78–4.69 (m, 1H), 4.54–4.45 (m, 3H), 4.32–4.13 (m, 2H), 4.12–4.01
(m, 1H), 3.70–3.63 (m, 1H), 3.30–3.09 (m, 9H), 2.85–2.74 (m, 2H),
2.74–2.56 (m, 3H), 2.56–2.38 (m, 4H), 2.23–2.01 (m, 9H), 1.87–1.79
(m, 1H), 1.62–1.61 (m, 5H), 1.32 (s, 3H), 1.22–1.15 (m, 17H), 0.99–0.87
(m, 33H), 0.80–0.57 (m, 18H); MS (ESI) m/z calcd for C64H120N5O11Si3
[M + H]+: 1218.8, [M + 2H]2+/2: 609.9, found: 1219.1, 610.4.
C
56H115NO13Si3 [M + 2H]2+/2: 546.9, found: 547.4.
5.1.1.5. Ring-opening intermediate with secondary amine (6). To a solu-
tion of 3 (2 g, 1.83 mmol) in anhydrous CHCl3 (20 mL) was added Pb
(OAc)4 (0.97 g, 2.19 mmol) at 0 ◦C. The reaction mixture was stirred at
0 ◦C for 0.5 h under nitrogen atmosphere. The aldehyde 4 was obtained
in a high yield and was directly used in the next step without
purification.
5.1.1.8. 11-N-{[3-(4-Phenyl-1H-1,2,3-triazol-1-yl)-1-yl]propyl}-9(S)-
hydroxyclarithromycin (11a). Hydrogen fluoride–pyridine (56 mg, 0.37
mmol, 65%) was added to a solution of 10a (150 mg, 0.12 mmol) in THF
(5 mL) at room temperature, and the resulting mixture was stirred at
room temperature for 18 h. The reaction mixture was quenched with
saturated NaHCO3 solution (20 mL) and diluted with DCM (20 mL). The
organic phase was separated and aqueous layer was extracted with DCM
(20 mL × 2). The combined organic layers were dried over anhydrous
Na2SO4, filtered, and evaporated in vacuo. The residue was separated
chromatographically (DCM/CH3OH/NH3⋅H2O = 10:1:0.1) to give
compound 11a (50 mg, 46%) as a white solid. Mp: 84–86 ◦C, TLC Rf =
0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR (600 MHz, CDCl3, δ
ppm): 7.90 (s, 1H), 7.84 (d, J = 7.4 Hz, 2H), 7.44–7.41 (m, 2H), 7.33 (t,
J = 7.4 Hz, 1H), 4.88 (d, J = 4.6 Hz, 1H), 4.48–4.44 (m, 3H), 4.38 (s,
1H), 4.10 (s, 1H), 4.06–4.01 (m, 1H), 3.92 (d, J = 11.7 Hz, 1H), 3.75 (d,
J = 7.5 Hz, 1H), 3.57–3.48 (m, 2H), 3.34–3.19 (m, 8H), 3.02 (t, J = 8.6
Hz, 2H), 2.87–2.84 (m, 1H), 2.72–2.51 (m, 5H), 2.47–2.15 (m, 14H),
1.94–1.91 (m, 1H), 1.71–1.66 (m, 2H), 1.46–1.42 (m, 1H), 1.34 (s, 3H),
1.30–1.25 (m, 10H), 1.19–1.18 (m, 3H), 1.09–1.07 (m, 3H), 0.89–0.87
(m, 3H), 0.79 (d, J = 6.1 Hz, 3H); 13C NMR (150 MHz, CDCl3, δ ppm):
176.74, 147.70, 130.71, 128.82, 128.79, 128.06, 125.75, 125.72,
120.14, 103.33, 96.28, 80.51, 78.10, 72.73, 72.71, 71.01, 68.85, 68.20,
65.52, 65.36, 52.48, 50.33, 49.49, 48.18, 45.21, 40.34, 39.39, 35.06,
34.11, 31.83, 29.31, 28.04, 27.21, 25.74, 23.49, 22.61, 21.60, 21.40,
18.31, 14.08, 13.99, 10.38; HRMS (ESI) m/z calcd for C46H78N5O11 [M
+ H]+: 876.5698, [M + 2H]2+/2: 438.7888, found: 876.5691,
438.8012.
Then, ethanolamine 5 (224 mg, 3.67 mmol) and NaBH(OAc)3 (1.16
g, 5.47 mmol) were added to a solution of 4 at 0 ◦C. The reaction mixture
was stirred at room temperature for 4 h and then quenched with satu-
rated NaHCO3 solution (20 mL). The organic phase was separated and
aqueous layer was extracted with DCM (20 mL × 2). The combined
organic layers were dried over anhydrous Na2SO4, filtered, and evapo-
rated in vacuo. The residue was separated chromatographically (DCM/
CH3OH/NH3⋅H2O = 10:1:0.1) to obtain the crude secondary amine 6
(1.5 g, 72% in two steps from intermediate 3) as a colorless oil.
5.1.1.6. Ring-opening intermediate with carboxylic acid (9a). NaBH
(OAc)3 (369 mg, 1.74 mmol) and 7a (233 mg, 1.16 mmol) were added to
a solution of 6 (660 mg, 0.58 mmol) in anhydrous CHCl3 (20 mL) at 0 ◦C,
and the reaction mixture was stirred at room temperature for 12 h. The
reaction mixture was quenched with saturated NaHCO3 solution (20
mL). The organic phase was separated and aqueous layer was extracted
with DCM (20 mL × 2). The combined organic layers were dried over
anhydrous Na2SO4, filtered, and evaporated in vacuo. The residue was
separated chromatographically (DCM/CH3OH/NH3⋅H2O = 30:1:0.1) to
obtain the crude ester 8a as a colorless oil.
LiOH (69 mg, 2.9 mmol) was added to a solution of 8a in the mixed
solvents of THF (15 mL), EtOH (5 mL) and H2O (5 mL) and the reaction
mixture was stirred at room temperature for 6 h. The reaction mixture
was evaporated in vacuo and the residue was separated chromato-
graphically (DCM/CH3OH/NH3⋅H2O = 15:1:0.1 ~ 10:1:0.1) to afford
the carboxylic acid 9a (0.4 g, 55% in two steps from intermediate 6) as a
colorless oil. TLC Rf = 0.51 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.91–7.79 (m, 3H), 7.44–7.39 (m, 2H),
7.33 (dd, J = 14.7, 7.4 Hz, 1H), 5.41–5.34 (m, 1H), 4.80 (dd, J = 32.1,
4.6 Hz, 1H), 4.66–4.55 (m, 1H), 4.53–4.38 (m, 2H), 4.31–4.22 (m, 1H),
3.97 (s, 1H), 3.74 (d, J = 6.1 Hz, 1H), 3.65–3.58 (m, 1H), 3.30–3.18 (m,
10H), 3.03 (s, 1H), 2.81–2.71 (m, 2H), 2.68–2.59 (m, 3H), 2.53–2.42 (m,
3H), 2.36–2.15 (m, 10H), 2.04–2.00 (m, 2H), 1.84–1.75 (m, 1H),
1.69–1.61 (m, 2H), 1.49–1.45 (m, 1H), 1.25–1.23 (m, 17H), 1.17 (d, J =
4.6 Hz, 3H), 1.08 (d, J = 7.4 Hz, 3H), 0.99–0.88 (m, 30H), 0.69–0.58 (m,
18H); MS (ESI) m/z calcd for C64H122N5O12Si3 [M + H]+: 1236.8, [M +
2H]2+/2: 618.9, found: 1237.2, 619.5.
According to the synthetic procedure of compound 11a, the other
compounds (11b-11 ) in Series A were also prepared.
5.1.1.9. 11-N-{[3-(4-p-Methylphenyl-1H-1,2,3-triazol-1-yl)-1-yl]propyl}-
9(S)-hydroxyclarithromycin (11b). White solid, yield 55%, mp:
90–92 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.84 (s, 1H), 7.72 (d, J = 8.0 Hz, 2H), 7.23 (d,
J = 8.0 Hz, 2H), 4.88 (d, J = 4.8 Hz, 1H), 4.56–4.36 (m, 5H), 4.08 (s,
1H), 4.03–4.01 (m, 1H), 3.91 (d, J = 12.0 Hz, 1H), 3.75 (d, J = 7.2 Hz,
1H), 3.54 (s, 2H), 3.34–3.16 (m, 8H), 3.03 (t, J = 8.7 Hz, 1H), 2.95–2.91
(m, 1H), 2.86–2.84 (m, 1H), 2.73–2.52 (m, 5H), 2.46–2.05 (m, 17H),
1.91 (s, 1H), 1.58 (d, J = 5.0 Hz, 2H), 1.56–1.55 (m, 1H), 1.33 (s, 3H),
1.28–1.25 (m, 10H), 1.18 (d, J = 7.3 Hz, 3H), 1.07 (d, J = 7.2 Hz, 3H),
0.87 (d, J = 6.9 Hz, 3H), 0.79 (d, J = 6.7 Hz, 3H); 13C NMR (150 MHz,
CDCl3, δ ppm): 176.69, 147.78, 137.91, 129.48, 127.85, 125.65, 119.76,
103.14, 96.24, 80.44, 79.48, 78.23, 78.06, 72.76, 71.01, 68.68, 65.59,
65.34, 53.22, 52.47, 50.33, 49.49, 48.17, 45.14, 36.62, 35.03, 34.03,
29.70, 29.32, 27.99, 27.22, 25.55, 22.69, 21.60, 21.32, 21.29, 18.32,
14.05, 10.42; HRMS (ESI) m/z calcd for C47H80N5O11 [M + H]+:
890.5854, [M + 2H]2+/2: 445.7967, found: 890.5858, 445.8036.
5.1.1.7. 15-Membered 11a-homo-aza-clarithromycin intermediate (10a).
To a suspension of 9a (400 mg, 0.32 mmol) and Et3N (324 mg, 3.2
mmol) in THF (5 mL) was added 2,4,6-trichlorobenzoyl chloride (234
mg, 0.96 mmol) at room temperature. The reaction solution was stirred
at room temperature for 4 h under nitrogen atmosphere. DMAP (985 mg,
8 mmol) and CH3CN (30 mL) were added to the reaction solution, and
refluxed for 0.5 h. The reaction mixture was concentrated in vacuo to
dryness, and then added DCM (20 mL) and saturated NH4Cl solution (20
5.1.1.10. 11-N-{[3-(4-p-Ethylphenyl-1H-1,2,3-triazol-1-yl)-1-yl]propyl}-
9(S)-hydroxyclarithromycin (11c). White solid, yield 43%, mp:
96–98 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
15

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
(600 MHz, CDCl3, δ ppm): 7.84 (s, 1H), 7.74 (d, J = 8.1 Hz, 2H), 7.25 (s,
2H), 4.88 (d, J = 4.5 Hz, 1H), 4.46–4.42 (m, 4H), 4.08 (s, 1H), 4.03–4.00
(m, 1H), 3.91 (d, J = 12.0 Hz, 1H), 3.75 (d, J = 7.1 Hz, 1H), 3.55 (s, 2H),
3.31–3.21 (m, 8H), 3.06–3.02 (m, 2H), 2.86–2.84 (m, 1H), 2.70–2.60
(m, 7H), 2.43–2.10 (m, 14H), 1.92 (s, 1H), 1.67–1.66 (m, 2H), 1.56–1.55
(m, 1H), 1.33 (s, 3H), 1.28–1.25 (m, 13H), 1.18 (d, J = 7.3 Hz, 3H), 1.07
5.1.1.14. 11-N-{[3-(4-p-n-Amylphenyl-1H-1,2,3-triazol-1-yl)-1-yl]pro-
pyl}-9(S)-hydroxyclarithromycin (11g). White solid, yield 32%, mp:
97–99 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.85 (s, 1H), 7.74 (d, J = 8.1 Hz, 2H), 7.23 (d,
J = 8.1 Hz, 2H), 4.88 (d, J = 4.5 Hz, 1H), 4.46–4.44 (m, 4H), 4.09–4.01
(m, 2H), 3.92 (d, J = 11.9 Hz, 1H), 3.75 (d, J = 7.4 Hz, 1H), 3.53 (s, 2H),
3.39–3.14 (m, 8H), 3.03–3.01 (m, 1H), 2.91–2.84 (m, 2H), 2.72–2.02
(m, 21H), 1.93–1.91 (m, 1H), 1.65–1.63 (m, 4H), 1.55 (d, J = 4.8 Hz,
1H), 1.38–1.32 (m, 7H), 1.29–1.25 (m, 10H), 1.18 (d, J = 7.3 Hz, 3H),
(d, J = 7.1 Hz, 3H), 0.87 (d, J = 6.7 Hz, 3H), 0.78 (d, J = 6.2 Hz, 3H); 13
C
NMR (150 MHz, CDCl3, δ ppm): 175.62, 147.56, 138.14, 129.48,
127.45, 125.49, 120.27, 102.09, 96.30, 80.13, 80.04, 78.49, 77.70,
72.80, 70.93, 68.20, 68.01, 67.88, 65.89, 65.16, 62.59, 61.45, 50.48,
49.29, 44.49, 35.03, 29.61, 29.23, 21.45, 21.21, 20.88, 18.77, 18.21,
14.69, 10.51; MS (ESI) m/z calcd for C48H82N5O11 [M + H]+: 904.6, [M
+ 2H]2+/2: 452.8, found: 904.8, 453.2.
1.08 (d, J = 6.5 Hz, 3H), 0.91–0.87 (m, 6H), 0.79 (d, J = 6.3 Hz, 3H); 13
C
NMR (100 MHz, CDCl3, δ ppm): δ 176.43, 147.62, 142.77, 128.63,
127.91, 125.47, 119.54, 103.04, 96.09, 80.29, 79.24, 78.08, 77.90,
72.57, 70.85, 68.55, 65.38, 65.20, 61.64, 60.36, 53.12, 52.35, 50.12,
49.28, 47.97, 44.99, 40.86, 40.21, 36.48, 35.51, 34.89, 33.88, 31.26,
30.85, 29.48, 27.83, 22.32, 21.38, 21.14, 18.12, 13.80, 10.20; MS (ESI)
m/z calcd for C51H88N5O11 [M + H]+: 946.6, [M + 2H]2+/2: 473.8,
found: 947.0, 474.0.
5.1.1.11. 11-N-{[3-(4-p-n-Propylphenyl-1H-1,2,3-triazol-1-yl)-1-yl]pro-
pyl}-9(S)-hydroxyclarithromycin (11d). White solid, yield 41%, mp:
78–80 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.83 (s, 1H), 7.73 (d, J = 8.1 Hz, 2H), 7.23 (d,
J = 8.1 Hz, 2H), 4.88 (d, J = 4.4 Hz, 1H), 4.50–4.39 (m, 5H), 4.07 (s,
1H), 4.03–4.00 (m, 1H), 3.91 (d, J = 12.1 Hz, 1H), 3.75 (d, J = 7.0 Hz,
1H), 3.60–3.53 (m, 2H), 3.34–3.28 (m, 4H), 3.22–3.20 (m, 4H), 3.03 (s,
2H), 2.86–2.84 (m, 1H), 2.70–2.60 (m, 7H), 2.45–2.11 (m, 14H),
1.92–1.90 (m, 1H), 1.68–1.65 (m, 4H), 1.56 (d, J = 4.9 Hz, 1H), 1.32 (s,
3H), 1.27–1.26 (m, 10H), 1.18 (d, J = 7.3 Hz, 3H), 1.06 (d, J = 7.3 Hz,
3H), 0.95 (t, J = 7.3 Hz, 3H), 0.86 (d, J = 7.1 Hz, 3H), 0.78 (d, J = 6.4
Hz, 3H); MS (ESI) m/z calcd for C49H84N5O11 [M + H]+: 918.6, [M +
2H]2+/2: 459.8, found: 918.9, 460.2.
5.1.1.15. 11-N-{[3-(4-p-Methoxyphenyl-1H-1,2,3-triazol-1-yl)-1-yl]pro-
pyl}-9(S)-hydroxyclarithromycin (11h). White solid, yield 80%, mp:
85–87 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.80 (s, 1H), 7.75 (d, J = 8.7 Hz, 2H), 6.96 (d,
J = 8.7 Hz, 2H), 4.88 (d, J = 4.4 Hz, 1H), 4.47–4.41 (m, 4H), 4.08 (s,
1H), 4.03–3.99 (m, 1H), 3.91 (d, J = 12.2 Hz, 1H), 3.85 (s, 3H), 3.75 (d,
J = 7.2 Hz, 1H), 3.59–3.50 (m, 2H), 3.39–3.16 (m, 8H), 3.03 (s, 1H),
2.94–2.92 (m, 1H), 2.86–2.84 (m, 1H), 2.71–2.53 (m, 5H), 2.48–1.97
(m, 14H), 1.94–1.90 (m, 1H), 1.68–1.66 (m, 2H), 1.55 (d, J = 4.8 Hz,
1H), 1.32 (s, 3H), 1.29–1.25 (m, 10H), 1.18 (d, J = 7.3 Hz, 3H), 1.06 (d,
J = 7.1 Hz, 3H), 0.86 (d, J = 7.1 Hz, 3H), 0.78 (d, J = 6.4 Hz, 3H); 13
C
5.1.1.12. 11-N-{[3-(4-p-Tert-butylphenyl-1H-1,2,3-triazol-1-yl)-1-yl]pro-
pyl}-9(S)-hydroxyclarithromycin (11e). White solid, yield 49%, mp:
110–112 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.84 (s, 1H), 7.76 (d, J = 8.4 Hz, 2H),
7.45 (d, J = 8.4 Hz, 2H), 4.88 (d, J = 4.5 Hz, 1H), 4.54–4.36 (m, 5H),
4.08–4.00 (m, 2H), 3.91 (d, J = 12.0 Hz, 1H), 3.75 (d, J = 7.1 Hz, 1H),
3.57–3.51 (m, 2H), 3.34–3.19 (m, 8H), 3.08–2.98 (m, 2H), 2.86–2.84
(m, 1H), 2.70–2.53 (m, 5H), 2.45–2.04 (m, 14H), 1.94–1.90 (m, 1H),
1.58–1.55 (m, 3H), 1.35–1.33 (m, 12H), 1.29–1.25 (m, 10H), 1.18 (d, J
= 7.3 Hz, 3H), 1.07 (d, J = 6.8 Hz, 3H), 0.87 (d, J = 6.8 Hz, 3H), 0.79 (d,
J = 6.2 Hz, 3H); 13C NMR (150 MHz, CDCl3, δ ppm): 176.54, 151.11,
147.63, 127.78, 125.64, 125.62, 125.41, 125.38, 119.71, 119.65,
103.02, 96.17, 80.36, 79.53, 78.18, 77.97, 72.71, 70.94, 68.55, 65.55,
65.29, 53.18, 52.43, 50.25, 49.39, 48.10, 45.05, 36.58, 34.97, 34.57,
33.94, 31.21, 29.61, 27.91, 27.13, 21.51, 21.20, 18.34, 18.24, 14.02,
13.97, 11.84, 10.35; MS (ESI) m/z calcd for C50H86N5O11 [M + H]+:
932.6, [M + 2H]2+/2: 466.8, found: 932.8, 467.2.
NMR (150 MHz, CDCl3, δ ppm): δ 176.55, 159.49, 147.48, 126.95,
123.36, 119.22, 114.14, 102.99, 96.15, 80.34, 79.51, 78.41, 78.16,
77.96, 72.70, 70.92, 68.53, 65.53, 65.28, 55.23, 53.14, 52.40, 50.23,
49.39, 48.07, 45.04, 40.36, 36.58, 34.96, 33.90, 29.59, 29.22, 27.90,
27.12, 22.58, 21.49, 21.19, 18.23, 13.97, 10.33; MS (ESI) m/z calcd for
C
47H80N5O12 [M + H]+: 906.6, [M + 2H]2+/2: 453.8, found: 906.8,
454.2.
5.1.1.16. 11-N-{[3-(4-p-Ethoxyphenyl-1H-1,2,3-triazol-1-yl)-1-yl]pro-
pyl}-9(S)-hydroxyclarithromycin (11i). White solid, yield 58%, mp:
101–103 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.81 (s, 1H), 7.75 (d, J = 8.8 Hz, 2H),
6.94 (d, J = 8.8 Hz, 2H), 4.88 (d, J = 4.6 Hz, 1H), 4.45–4.43 (m, 3H),
4.36 (t, J = 10.2 Hz, 1H), 4.11–4.03 (m, 4H), 3.92 (d, J = 12.0 Hz, 1H),
3.75 (d, J = 7.6 Hz, 1H), 3.55–3.49 (m, 2H), 3.35–3.20 (m, 8H), 3.02 (t,
J = 7.6 Hz, 1H), 2.92–2.84 (m, 2H), 2.69–2.47 (m, 5H), 2.47–2.10 (m,
14H), 1.92–1.91 (m, 1H), 1.70–1.66 (m, 2H), 1.54 (d, J = 4.9 Hz, 1H),
1.43 (t, J = 7.0 Hz, 3H), 1.34 (s, 3H), 1.30–1.24 (m, 10H), 1.18 (d, J =
7.3 Hz, 3H), 1.09 (d, J = 7.3 Hz, 3H), 0.87 (d, J = 7.1 Hz, 3H), 0.79 (d, J
= 6.6 Hz, 3H); 13C NMR (100 MHz, CDCl3, δ ppm): 176.43, 158.73,
147.41, 126.81, 123.11, 119.05, 114.58, 103.01, 96.06, 80.27, 79.22,
78.33, 78.05, 77.87, 72.56, 70.83, 68.52, 65.36, 65.17, 63.30, 61.61,
60.36, 53.07, 52.32, 50.10, 49.26, 47.93, 44.97, 40.85, 40.20, 36.46,
34.87, 33.84, 31.28, 29.49, 27.79, 21.37, 21.12, 18.10, 14.60, 13.79,
10.17; MS (ESI) m/z calcd for C48H82N5O12 [M + H]+: 920.6, [M +
2H]2+/2: 460.8, found: 920.8, 461.2.
5.1.1.13. 11-N-{[3-(4-p-n-Butylphenyl-1H-1,2,3-triazol-1-yl)-1-yl]pro-
pyl}-9(S)-hydroxyclarithromycin (11f). White solid, yield 36%, mp:
94–96 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.85 (s, 1H), 7.74 (d, J = 8.0 Hz, 2H), 7.23 (d,
J = 8.0 Hz, 2H), 4.88 (d, J = 4.6 Hz, 1H), 4.46–4.44 (m, 3H), 4.37 (t, J =
10.3 Hz, 1H), 4.10 (s, 1H), 4.04 (dd, J = 9.0, 6.4 Hz, 1H), 3.92 (d, J =
12.1 Hz, 1H), 3.75 (d, J = 7.2 Hz, 1H), 3.52 (s, 2H), 3.35–3.20 (m, 8H),
3.02–3.01 (m, 1H), 2.90–2.83 (m, 2H), 2.65–2.62 (m, 2H), 2.58–2.06
(m, 19H), 1.92–1.91 (m, 1H), 1.70–1.68 (m, 2H), 1.63–1.61 (m, 2H),
1.54 (d, J = 4.8 Hz, 1H), 1.40–1.36 (m, 2H), 1.34 (s, 3H), 1.30–1.25 (m,
10H), 1.18 (d, J = 7.3 Hz, 3H), 1.09 (d, J = 7.1 Hz, 3H), 0.94 (t, J = 7.4
Hz, 3H), 0.88 (d, J = 6.9 Hz, 3H), 0.79 (d, J = 6.3 Hz, 3H); 13C NMR
(100 MHz, CDCl3, δ ppm): δ 176.42, 147.62, 142.74, 128.63, 127.91,
125.47, 119.53, 103.01, 96.08, 80.28, 79.27, 78.35, 78.07, 77.89,
72.57, 70.84, 68.52, 65.39, 65.20, 61.64, 60.39, 53.11, 52.34, 50.12,
49.27, 47.97, 44.98, 40.84, 40.22, 36.48, 35.22, 34.89, 33.87, 33.33,
27.82, 22.11, 21.38, 21.13, 18.12, 13.80, 13.72, 10.20; MS (ESI) m/z
calcd for C50H86N5O11 [M + H]+: 932.6, [M + 2H]2+/2: 466.8, found:
932.8, 467.1.
5.1.1.17. 11-N-{[3-(4-p-Propoxyphenyl-1H-1,2,3-triazol-1-yl)-1-yl]pro-
pyl}-9(S)-hydroxyclarithromycin (11j). White solid, yield 61%, mp:
87–89 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.80 (s, 1H), 7.74 (d, J = 8.8 Hz, 2H), 6.95 (d,
J = 8.8 Hz, 2H), 4.88 (d, J = 4.7 Hz, 1H), 4.45–4.43 (m, 3H), 4.39–4.36
(m, 1H), 4.10–4.02 (m, 2H), 3.96 (t, J = 6.6 Hz, 2H), 3.93–3.91 (m, 1H),
3.75 (d, J = 7.5 Hz, 1H), 3.52 (s, 2H), 3.33–3.20 (m, 8H), 3.02 (t, J = 7.9
Hz, 1H), 2.93–2.89 (m, 1H), 2.87–2.84 (m, 1H), 2.70–2.51 (m, 5H),
2.49–2.09 (m, 14H), 1.93–1.91 (m, 1H), 1.85–1.81 (m, 2H), 1.67–1.64
16

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
(m, 2H), 1.55 (d, J = 4.8 Hz, 1H), 1.34 (s, 3H), 1.30–1.24 (m, 10H), 1.18
(d, J = 7.3 Hz, 3H), 1.09 (d, J = 7.2 Hz, 3H), 1.05 (t, J = 7.4 Hz, 3H),
0.87 (d, J = 7.1 Hz, 3H), 0.79 (d, J = 6.7 Hz, 3H); 13C NMR (100 MHz,
CDCl3, δ ppm): 176.42, 158.94, 147.42, 126.79, 123.06, 119.04, 114.61,
103.04, 96.06, 80.27, 79.19, 78.34, 78.05, 77.88, 72.55, 70.84, 69.39,
68.53, 65.35, 65.17, 60.33, 53.08, 52.34, 50.08, 49.25, 47.92, 44.98,
40.86, 40.17, 36.48, 34.88, 33.85, 29.40, 27.79, 22.37, 21.36, 21.13,
18.10, 13.76, 10.27, 10.16; MS (ESI) m/z calcd for C49H84N5O12 [M +
H]+: 934.6, [M + 2H]2+/2: 467.8, found: 934.9, 468.1.
column chromatography (PE/EtOAc = 6:1) yielded azide 22 (1.27 g,
86%) as a light yellow oil.
5.1.2.2. (R)-1-Chloro-3-[4-(p-tolyl)-1H-1,2,3-triazol-1-yl]propan-2-ol
(24). 22 (0.35 g, 2.58 mmol) in 75% methanol solution (20 mL) was
reacted with 4-ethynyltoluene 23 (0.25 g, 2.15 mmol) in the presence of
CuSO4 (22 mg) and L-ascorbic acid sodium salt (66 mg) at 40 ◦C for 24 h.
The reaction mixture was evaporated in vacuo and the residue was
separated chromatographically (DCM/CH3OH = 30:1) to form the
◦
alcohol 24 (0.51 g, 94%) as a white solid. Mp: 118–120 C, TLC Rf =
5.1.1.18. 11-N-{[3-(4-p-Fluorophenyl-1H-1,2,3-triazol-1-yl)-1-yl]pro-
pyl}-9(S)-hydroxyclarithromycin (11k). White solid, yield 35%, mp:
85–87 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.86–7.78 (m, 3H), 7.14–7.10 (m, 2H), 4.88
(d, J = 4.7 Hz, 1H), 4.49–4.46 (m, 4H), 4.05 (s, 1H), 4.00–3.97 (m, 1H),
3.91 (d, J = 12.2 Hz, 1H), 3.74 (d, J = 2.7 Hz, 1H), 3.60–3.55 (m, 2H),
3.36–3.10 (m, 8H), 3.04 (s, 1H), 2.99–2.94 (m, 1H), 2.91–2.71 (m, 6H),
2.62–1.98 (m, 14H), 1.91 (s, 1H), 1.65–1.63 (m, 2H), 1.57 (d, J = 4.8 Hz,
1H), 1.31 (s, 3H), 1.29–1.26 (m, 10H), 1.18 (d, J = 7.3 Hz, 3H), 1.03 (d,
J = 7.3 Hz, 3H), 0.86 (d, J = 8.4 Hz, 3H), 0.82–0.75 (m, 3H); MS (ESI)
m/z calcd for C46H77FN5O11 [M + H]+: 894.6, [M + 2H]2+/2: 447.8,
found: 894.8, 448.2.
0.32 (DCM/CH3OH = 30:1); 1H NMR (600 MHz, CDCl3, δ ppm): 7.83 (s,
1H), 7.64 (d, J = 8.1 Hz, 2H), 7.20 (d, J = 7.9 Hz, 2H), 4.67 (dd, J =
14.0, 3.4 Hz, 1H), 4.50 (dd, J = 14.0, 7.0 Hz, 1H), 4.42–4.38 (m, 1H),
3.61–3.60 (m, 2H), 2.38 (s, 3H); MS (ESI) m/z calcd for C12H15ClN3O
[M + H]+: 252.1, found: 252.2.
5.1.2.3. (S)-1-Amino-[3-(4-(p-tolyl)-1H-1,2,3-triazol-1-yl]propan-2-ol
(16a). NaOH solution (5 mL, 10 M) was added to a solution of alcohol
24 (0.5 g, 1.99 mmol) in CH3CN (2 mL) and stirred at room temperature
for 1 h. The reaction mixture was diluted with H2O (10 mL) and
extracted with DCM (20 mL × 3). The combined organic layers were
dried over anhydrous Na2SO4, filtered and evaporated in vacuo to yield
the epoxide 25 as a white solid. This intermediate 25 can be used
directly in the next step without purification.
5.1.1.19. 11-N-{[3-(4-p-Chlorophenyl-1H-1,2,3-triazol-1-yl)-1-yl]pro-
pyl}-9(S)-hydroxyclarithromycin (11l). White solid, yield 46%, mp:
84–86 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.90 (s, 1H), 7.77 (d, J = 8.5 Hz, 2H), 7.40 (d,
J = 8.5 Hz, 2H), 4.88 (d, J = 4.7 Hz, 1H), 4.52–4.41 (m, 4H), 4.05 (s,
1H), 3.99 (dd, J = 9.1, 6.3 Hz, 1H), 3.92 (d, J = 12.3 Hz, 1H), 3.75 (d, J
= 6.6 Hz, 1H), 3.62–3.55 (m, 2H), 3.33–3.20 (m, 8H), 3.05–3.00 (m,
2H), 2.99–2.95 (m, 1H), 2.87–2.73 (m, 5H), 2.36–2.00 (m, 14H), 1.91 (s,
1H), 1.68 (s, 2H), 1.56 (s, 1H), 1.31 (s, 3H), 1.27–1.25 (m, 10H), 1.18 (d,
J = 7.3 Hz, 3H), 1.02 (d, J = 7.4 Hz, 3H), 0.87 (d, J = 7.3 Hz, 3H),
0.81–0.75 (m, 3H); MS (ESI) m/z calcd for C46H77ClN5O11 [M + H]+:
910.5, [M + 2H]2+/2: 455.8, found: 910.8, 456.2.
25% NH3⋅H2O solution (20 mL) was added to a solution of the
epoxide 25 in CH3CN (2 mL) and stirred at room temperature for 3 h.
The reaction mixture was diluted with H2O (10 mL) and extracted with
DCM (30 mL × 3). The combined organic layers were dried over
anhydrous Na2SO4, filtered and evaporated in vacuo. Purification by
column chromatography (DCM/CH3OH/NH3⋅H2O = 10:1:0.1) afforded
substituted ethanolamine 16a (0.27 g, 58% in two steps from interme-
diate 24) as a white solid. Mp: 135–137 ◦C, TLC Rf = 0.11 (DCM/
1
CH3OH/NH3⋅H2O = 10:1:0.1); H NMR (600 MHz, DMSO‑d6, δ ppm):
8.42 (s, 1H), 7.74 (d, J = 8.1 Hz, 2H), 7.25 (d, J = 8.1 Hz, 2H), 5.12 (s,
1H), 4.49 (dd, J = 13.8, 4.0 Hz, 1H), 4.29–4.25 (m, 1H), 3.80–3.76 (m,
1H), 2.57–2.53 (m, 2H), 2.33 (s, 3H), 1.63 (s, 2H); MS (ESI) m/z calcd for
C
12H17N4O [M + H]+: 233.1, found: 233.3.
5.1.1.20. 11-N-{[3-(4-o-Pyridyl-1H-1,2,3-triazol-1-yl)-1-yl]propyl}-9(S)-
hydroxyclarithromycin (11m). White solid, yield 39%, mp: 96–98 ◦C,
TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR (600 MHz,
CDCl3, δ ppm): 8.58 (d, J = 4.7 Hz, 1H), 8.22 (s, 1H), 8.16 (d, J = 7.9 Hz,
1H), 7.78 (t, J = 7.3 Hz, 1H), 7.24 (d, J = 4.9 Hz, 1H), 4.88 (d, J = 4.7
Hz, 1H), 4.54–4.45 (m, 4H), 4.38–4.35 (m, 1H), 4.08–4.01 (m, 2H), 3.92
(d, J = 12.1 Hz, 1H), 3.75 (d, J = 7.2 Hz, 1H), 3.58–3.48 (m, 2H),
3.34–3.20 (m, 8H), 3.02 (d, J = 8.8 Hz, 1H), 2.89–2.84 (m, 2H),
2.63–2.52 (m, 5H), 2.39–2.05 (m, 14H), 1.91 (s, 1H), 1.71 (dd, J = 13.9,
7.2 Hz, 2H), 1.57–1.55 (m, 1H), 1.33 (s, 3H), 1.28–1.24 (m, 10H), 1.19
(d, J = 7.3 Hz, 3H), 1.07 (d, J = 7.4 Hz, 3H), 0.86 (d, J = 7.0 Hz, 3H),
0.80 (d, J = 6.7 Hz, 3H); 13C NMR (100 MHz, CDCl3, δ ppm): 176.42,
150.06, 149.11, 148.06, 136.67, 122.57, 122.12, 119.99, 102.76, 95.96,
80.17, 79.37, 78.23, 78.04, 77.82, 72.57, 70.78, 68.24, 65.38, 65.03,
60.17, 52.33, 50.07, 49.23, 48.22, 44.85, 40.21, 36.49, 34.81, 33.79,
30.20, 29.43, 27.73, 21.33, 21.01, 18.09, 13.82, 10.10; MS (ESI) m/z
calcd for C45H77N6O11 [M + H]+: 877.6, [M + 2H]2+/2: 439.3, found:
877.8, 439.7.
5.1.2.4. Ring-opening intermediate with carboxylic acid (18a). To a so-
lution of 3 (469 mg, 0.43 mmol) in anhydrous CHCl3 (20 mL) was added
Pb(OAc)4 (228 mg, 0.52 mmol) at 0 ◦C. The reaction mixture was stirred
◦
at 0 C for 0.5 h under nitrogen atmosphere. The aldehyde 4 was ob-
tained in a high yield and was directly used in the next step without
purification.
To a solution of 4 was added 16a (200 mg, 0.86 mmol) and NaBH
(OAc)3 (273 mg, 1.29 mmol) at 0 ◦C . The reaction mixture was stirred at
room temperature for 4 h under nitrogen atmosphere. Then, NaBH
(OAc)3 (273 mg, 1.29 mmol) and 37% aqueous formaldehyde solution
(209 mg, 2.58 mmol) were added and stirred at room temperature for
another 4 h under nitrogen atmosphere. The reaction mixture was
quenched with saturated NaHCO3 solution (20 mL). The organic phase
was separated and aqueous layer was extracted with DCM (20 mL × 2).
The combined organic layers were dried over anhydrous Na2SO4,
filtered, and evaporated in vacuo. The residue was separated chro-
matographically (DCM/CH3OH/NH3⋅H2O = 30:1:0.1) to yield the crude
ester 17a as a colorless oil.
5.1.2. Synthesis of C13-substituted 15-membered homoclarithromycin
derivatives (Series B)
LiOH (51 mg, 2.15 mmol) was added to a solution of 17a in the
mixed solvents of THF (15 mL), EtOH (5 mL) and H2O (5 mL) and the
reaction mixture was stirred at room temperature for 6 h. The reaction
mixture was evaporated in vacuo and the residue was separated chro-
matographically (DCM/CH3OH/NH3⋅H2O = 15:1:0.1 ~ 10:1:0.1) to
give the carboxylic acid 18a (0.37 g, 70% in four steps from interme-
diate 3) as a white solid. Mp: 89–92 ◦C, TLC Rf = 0.42 (DCM/CH3OH/
NH3⋅H2O = 15:1:0.1); 1H NMR (600 MHz, CDCl3, δ ppm): 7.99 (s, 1H),
7.72 (d, J = 8.1 Hz, 2H), 7.23 (d, J = 8.1 Hz, 2H), 4.79 (d, J = 4.6 Hz,
5.1.2.1. (R)-1-Azido-3-chloropropan-2-ol (22). To a solution of R-
epichlorohydrin 21 (1 g, 10.9 mmol) in AcOH (10 mL) and H2O (10 mL)
was added NaN3 (3.7 g, 56.9 mmol) at room temperature. The reaction
mixture was stirred at 30 ◦C for 5 h, and then EtOAc (20 mL) was added.
The organic phase was separated and aqueous layer was extracted with
EtOAc (20 mL × 3). The combined organic phases were dried over
anhydrous Na2SO4, filtered, and evaporated in vacuo. Purification using
17

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
1H), 4.55 (dd, J = 13.8, 4.0 Hz, 1H), 4.48–4.41 (m, 3H), 4.26 (dd, J =
9.0, 6.4 Hz, 1H), 3.93–3.92 (m, 1H), 3.73 (d, J = 6.0 Hz, 1H), 3.63–3.59
(m, 1H), 3.30 (s, 3H), 3.25–3.16 (m, 7H), 3.07 (d, J = 10.0 Hz, 1H), 2.79
(s, 1H), 2.68–2.64 (m, 1H), 2.58 (s, 3H), 2.39–2.25 (m, 8H), 2.19 (s, 6H),
1.79–1.74 (m, 1H), 1.62–1.55 (m, 2H), 1.45 (dd, J = 15.0, 4.9 Hz, 1H),
1.28 (s, 3H), 1.27–1.25 (m, 1H), 1.22 (d, J = 6.3 Hz, 3H), 1.18–1.14 (m,
10H), 1.11 (d, J = 6.8 Hz, 3H), 1.08 (d, J = 7.4 Hz, 3H), 0.98–0.91 (m,
30H), 0.67–0.57 (m, 18H); MS (ESI) m/z calcd for C64H123N5O12Si3 [M
+ 2H]2+/2: 618.9, found: 619.5.
5.1.2.7. 13(R)-[1-(4-p-Methylphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (20b). White solid, yield 63%, mp:
96–99 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.78–7.70 (m, 3H), 7.23 (d, J = 7.9 Hz, 2H),
5.21 (s, 1H), 4.88–4.78 (m, 1H), 4.67–4.55 (m, 2H), 4.45 (d, J = 7.0 Hz,
1H), 4.11–3.95 (m, 2H), 3.88–3.83 (m, 1H), 3.66–3.51 (m, 2H),
3.34–3.14 (m, 8H), 3.04–2.97 (m, 2H), 2.90–2.86 (m, 1H), 2.82–2.54
(m, 5H), 2.54–2.18 (m, 12H), 2.15–2.02 (m, 2H), 1.94–1.91 (m, 1H),
1.80 (s, 1H), 1.67–1.61 (m, 2H), 1.53 (dd, J = 14.4, 3.8 Hz, 1H),
1.39–1.20 (m, 16H), 1.13 (d, J = 6.8 Hz, 3H), 1.02 (d, J = 6.8 Hz, 3H),
0.97 (d, J = 7.1 Hz, 3H); 13C NMR (150 MHz, CDCl3, δ ppm): 180.13,
147.95, 138.13, 129.54, 127.61, 125.72, 125.66, 120.07, 96.48, 80.43,
77.73, 72.68, 70.98, 68.41, 66.35, 65.81, 65.40, 65.35, 51.47, 51.37,
50.02, 49.43, 49.39, 40.52, 35.21, 34.83, 30.44, 21.62, 21.45, 21.29,
21.23, 21.14, 20.97, 18.68, 18.27, 10.27; HRMS (ESI) m/z calcd for
5.1.2.5. 15-Membered
11a-homo-aza-clarithromycin
intermediate
(19a). 18a (370 mg, 0.30 mmol), Et3N (303 mg, 3 mmol) and 2,4,6-tri-
chlorobenzoyl chloride (219 mg, 0.9 mmol) were added in THF (5 mL) at
room temperature. The reaction solution was stirred at room tempera-
ture for 4 h under nitrogen atmosphere. DMAP (922 mg, 7.5 mmol) and
CH3CN (30 mL) were added to the reaction solution, and refluxed for
0.5 h. The reaction mixture was concentrated in vacuo to dryness, and
then DCM (20 mL) and saturated NH4Cl solution (20 mL) were added.
The organic phase was separated and aqueous layer was extracted with
DCM (20 mL × 2). The combined organic layers were dried over
anhydrous Na2SO4, filtered, and evaporated in vacuo. The residue was
separated chromatographically (PE/acetone = 15:1 ~ 10:1) to afford
the cyclic intermediate 19a (150 mg, 41%) as a white solid. Mp:
85–88 ◦C, TLC Rf = 0.53 (PE/acetone = 3:1); 1H NMR (600 MHz, CDCl3,
δ ppm): 7.72–7.70 (m, 3H), 7.23 (d, J = 8.0 Hz, 2H), 5.25 (s, 1H),
4.81–4.76 (m, 1H), 4.72–4.69 (m, 1H), 4.57–4.52 (m, 1H), 4.39 (d, J =
6.9 Hz, 1H), 4.27–4.17 (m, 2H), 3.65–3.62 (m, 1H), 3.57–3.53 (m, 1H),
3.31 (s, 1H), 3.27 (s, 3H), 3.21–3.16 (m, 5H), 2.79–2.73 (m, 2H),
2.52–2.42 (m, 3H), 2.38–2.37 (m, 4H), 2.32–2.27 (m, 3H), 2.20 (s, 6H),
2.13–2.10 (m, 1H), 1.96–1.90 (m, 2H), 1.84–1.81 (m, 1H), 1.63–1.61
(m, 2H), 1.37 (d, J = 6.7 Hz, 1H), 1.31 (s, 3H), 1.24–1.22 (m, 4H),
1.16–1.14 (m, 13H), 0.99–0.95 (m, 33H), 0.65–0.59 (m, 18H); MS (ESI)
m/z calcd for C64H121N5O11Si3 [M + 2H]2+/2: 609.9, found: 610.5.
C
46H78N5O11 [M + H]+: 876.5698, [M + 2H]2+/2: 438.7888, found:
876.5655, 438.7967.
5.1.2.8. 13(S)-[1-(4-p-Ethylphenyl-1H-1,2,3-triazol-1-yl)-1-methylene]-9
(S)-hydroxyclarithromycin (20c). White solid, yield 36%, mp:
98–100 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.78–7.73 (m, 3H), 7.27 (s, 2H), 5.54 (s, 1H),
4.79 (d, J = 4.4 Hz, 1H), 4.55–4.54 (m, 1H), 4.46 (d, J = 7.1 Hz, 1H),
4.26 (s, 1H), 4.03–3.89 (m, 2H), 3.72 (d, J = 6.5 Hz, 1H), 3.53–3.46 (m,
2H), 3.32 (s, 1H), 3.26 (s, 3H), 3.21 (s, 3H), 3.11–3.07 (m, 1H),
3.02–2.97 (m, 2H), 2.92–2.89 (m, 1H), 2.82–2.66 (m, 7H), 2.48–2.15
(m, 9H), 2.13–1.97 (m, 3H), 1.88–1.85 (m, 1H), 1.82–1.73 (m, 2H),
1.46–1.45(m, 1H), 1.29–1.25 (m, 16H), 1.19 (d, J = 7.5 Hz, 3H), 0.98 (d,
J = 5.5 Hz, 3H), 0.84 (d, J = 4.2 Hz, 3H), 0.72 (d, J = 6.1 Hz, 3H); 13
C
NMR (150 MHz, CDCl3, δ ppm): 175.97, 148.03, 144.52, 128.29,
127.63, 125.67, 125.64, 102.37, 96.17, 79.88, 77.85, 74.18, 72.72,
70.81, 70.76, 67.91, 65.59, 65.16, 51.52, 50.12, 49.34, 44.93, 43.98,
37.22, 34.81, 32.37, 28.57, 21.52, 21.41, 21.37, 21.09, 20.96, 20.85,
19.67, 18.54, 18.19, 17.89, 15.93, 15.41, 14.75, 12.20, 10.72; MS (ESI)
m/z calcd for C47H80N5O11 [M + H]+: 890.6, [M + 2H]2+/2: 445.8,
found: 890.8, 446.2.
5.1.2.6. 13(S)-[1-(4-p-Methylphenyl-1H-1,2,3-triazol-1-yl)-1-methylene]-
9(S)-hydroxyclarithromycin (20a). Hydrogen fluoride–pyridine (56 mg,
0.37 mmol, 65%) was added to 19a (150 mg, 0.12 mmol) in THF (5 mL)
at room temperature, and the resulting mixture was stirred at room
temperature for 18 h. The reaction mixture was quenched with saturated
NaHCO3 solution (20 mL) and diluted with DCM (20 mL). The organic
phase was separated and aqueous layer was extracted with DCM (20 mL
× 2). The combined organic layers were dried over anhydrous Na2SO4,
filtered, and evaporated in vacuo. The residue was separated chro-
matographically (DCM/CH3OH/NH3⋅H2O = 10:1:0.1) to obtain com-
pound 20a (55 mg, 51%) as a white solid. Mp: 98–100 ◦C, TLC Rf = 0.40
(DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR (600 MHz, CDCl3, δ
ppm): 7.76–7.70 (m, 3H), 7.24 (d, J = 7.9 Hz, 2H), 5.54–5.52 (m, 1H),
4.81 (d, J = 4.1 Hz, 1H), 4.56 (dd, J = 14.6, 4.5 Hz, 1H), 4.49 (dd, J =
14.5, 5.1 Hz, 1H), 4.43 (d, J = 6.9 Hz, 1H), 4.28 (s, 1H), 4.02–3.97 (m,
1H), 3.72 (d, J = 7.1 Hz, 1H), 3.55–3.49 (m, 2H), 3.26 (s, 3H), 3.22–3.04
(m, 5H), 3.02–2.93 (m, 2H), 2.93–2.89 (m, 1H), 2.79–2.58 (m, 5H),
2.58–2.24 (m, 12H), 2.21–2.11 (m, 3H), 1.88–1.85 (m, 1H), 1.81–1.71
(m, 2H), 1.45 (dd, J = 15.1, 4.7 Hz, 1H), 1.29 (s, 3H), 1.28 (d, J = 6.3 Hz,
3H), 1.26–1.18 (m, 10H), 1.01 (d, J = 6.7 Hz, 3H), 0.83 (d, J = 6.9 Hz,
3H), 0.72 (d, J = 6.5 Hz, 3H); 13C NMR (150 MHz, CDCl3, δ ppm):
176.14, 148.11, 138.22, 129.58, 127.50, 125.66, 125.63, 120.07,
102.83, 96.19, 80.03, 78.73, 77.99, 72.76, 70.98, 68.34, 65.59, 65.23,
63.05, 59.01, 51.65, 50.17, 49.45, 45.15, 44.18, 34.89, 32.54, 30.73,
21.52, 21.30, 21.10, 21.04, 18.27, 18.21, 14.78, 10.80; HRMS (ESI) m/z
calcd for C46H78N5O11 [M + H]+: 876.5698, [M + 2H]2+/2: 438.7880,
found: 876.5699, 438.7960.
5.1.2.9. 13(R)-[1-(4-p-Ethylphenyl-1H-1,2,3-triazol-1-yl)-1-methylene]-9
(S)-hydroxyclarithromycin (20d). White solid, yield 55%, mp:
100–103 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.74 (dd, J = 13.1, 6.3 Hz, 3H), 7.25 (s,
2H), 5.20 (s, 1H), 4.82 (s, 1H), 4.66–4.58 (m, 2H), 4.44 (d, J = 7.1 Hz,
1H), 4.04–4.01 (m, 1H), 3.83 (s, 1H), 3.61–3.49 (m, 2H), 3.33 (s, 1H),
3.24 (s, 3H), 3.19 (s, 3H), 3.14 (s, 1H), 3.00–2.96 (m, 2H), 2.90–2.87 (m,
1H), 2.81–2.58 (m, 7H), 2.55–2.33 (m, 9H), 2.26–2.19 (m, 2H),
2.08–2.07 (m, 1H), 1.94–1.91 (m, 1H), 1.84–1.76 (m, 2H), 1.54–1.51
(m, 1H), 1.30–1.19 (m, 19H), 1.14 (d, J = 6.9 Hz, 3H), 1.02 (d, J = 7.0
Hz, 3H), 0.97 (d, J = 6.8 Hz, 3H); 13C NMR (100 MHz, CDCl3, δ ppm):
175.59, 147.62, 144.20, 128.06, 127.63, 125.49, 119.93, 96.25, 80.15,
77.52, 72.43, 70.68, 68.02, 65.97, 65.16, 51.16, 49.78, 49.15, 45.02,
40.23, 40.07, 38.19, 36.88, 34.69, 32.61, 30.14, 29.56, 28.39, 21.30,
20.96, 20.87, 20.79, 18.39, 18.01, 15.23, 9.96; MS (ESI) m/z calcd for
C
47H80N5O11 [M + H]+: 890.6, [M + 2H]2+/2: 445.8, found: 890.8,
446.1.
5.1.2.10. 13(S)-[1-(4-p-n-Propylphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (20e). White solid, yield 50%, mp:
98–101 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.73 (d, J = 8.0 Hz, 3H), 7.24 (d, J = 8.0 Hz,
2H), 5.53 (d, J = 9.6 Hz, 1H), 4.83 (d, J = 4.2 Hz, 1H), 4.55–4.54 (m,
1H), 4.49 (dd, J = 14.5, 5.2 Hz, 1H), 4.42 (d, J = 7.1 Hz, 1H), 4.29 (s,
1H), 4.04–3.99 (m, 1H), 3.72 (d, J = 7.4 Hz, 1H), 3.51–3.50 (m, 2H),
3.38–3.20 (m, 8H), 3.04–2.94 (m, 2H), 2.63–2.60 (m, 2H), 2.58–2.43
(m, 5H), 2.38–2.14 (m, 9H), 2.09–2.01 (m, 2H), 1.95–1.75 (m, 4H),
1.69–1.63 (m, 2H), 1.45 (dd, J = 15.2, 4.7 Hz, 1H), 1.30 (s, 3H), 1.28 (d,
According to the synthetic procedure of compound 20a, the other
compounds (20b-20 l) in Series B were also prepared.
18

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
J = 6.2 Hz, 3H), 1.25–1.18 (m, 10H), 1.03 (d, J = 7.0 Hz, 3H), 0.96 (t, J
J = 14.5, 4.5 Hz, 1H), 4.48 (dd, J = 14.5, 5.1 Hz, 1H), 4.43 (d, J = 6.0
Hz, 1H), 4.29 (s, 1H), 4.09–4.05 (m, 2H), 4.02–4.00 (m, 1H), 3.72 (d, J
= 7.2 Hz, 1H), 3.51 (d, J = 8.0 Hz, 2H), 3.35–3.11 (m, 8H), 3.01–2.90
(m, 3H), 2.72–2.68 (m, 2H), 2.58–2.07 (m, 14H), 2.02 (d, J = 9.4 Hz,
1H), 1.88–1.85 (m, 1H), 1.80–1.71 (m, 2H), 1.47–1.46 (m, 1H), 1.44 (t,
J = 7.0 Hz, 3H), 1.30–1.18 (m, 16H), 1.02–1.01 (m, 3H), 0.83 (d, J = 7.0
Hz, 3H), 0.72 (d, J = 6.6 Hz, 3H); 13C NMR (100 MHz, CDCl3, δ ppm):
175.92, 158.92, 147.72, 126.80, 122.73, 119.38, 114.68, 102.84, 96.01,
79.91, 79.54, 78.62, 77.84, 72.53, 70.81, 68.23, 67.68, 65.34, 65.04,
63.34, 62.81, 58.88, 51.44, 49.93, 49.21, 45.01, 43.96, 41.09, 40.31,
36.67, 34.74, 32.33, 29.46, 22.00, 21.29, 20.94, 18.06, 14.58, 14.50,
10.48; MS (ESI) m/z calcd for C47H81N5O12 [M + 2H]2+/2: 453.8, found:
454.1.
= 7.3 Hz, 3H), 0.84 (d, J = 7.0 Hz, 3H), 0.72 (d, J = 6.5 Hz, 3H); 13
C
NMR (100 MHz, CDCl3, δ ppm): 175.91, 147.88, 142.76, 128.75,
127.55, 125.40, 119.89, 102.79, 96.00, 79.87, 79.51, 78.61, 77.81,
72.51, 70.75, 68.18, 67.61, 65.31, 65.02, 62.78, 58.83, 51.43, 49.91,
49.19, 44.98, 43.93, 41.06, 40.30, 37.57, 36.65, 34.72, 32.30, 30.74,
24.21, 21.99, 21.28, 20.92, 18.03, 14.47, 13.53, 10.46; MS (ESI) m/z
calcd for C48H82N5O11 [M + H]+: 904.6, [M + 2H]2+/2: 452.8, found:
904.8, 453.1.
5.1.2.11. 13(R)-[1-(4-p-n-Propylphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (20f). White solid, yield 52%, mp:
90–92 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.75–7.72 (m, 3H), 7.23 (d, J = 8.1 Hz, 2H),
5.16 (s, 1H), 4.81 (s, 1H), 4.72–4.59 (m, 3H), 4.42 (d, J = 7.2 Hz, 1H),
4.06–4.01 (m, 1H), 3.81 (s, 1H), 3.56–3.46 (m, 2H), 3.32–3.18 (m, 8H),
3.00–2.90 (m, 3H), 2.62–2.60 (m, 2H), 2.57–2.25 (m, 14H), 2.21–2.11
(m, 3H), 1.94–1.91 (m, 1H), 1.83 (s, 2H), 1.68–1.64 (m, 2H), 1.52–1.49
(m, 1H), 1.31–1.22 (m, 16H), 1.15 (d, J = 6.9 Hz, 3H), 1.05 (d, J = 10.1
Hz, 3H), 0.98–0.94 (m, 6H); MS (ESI) m/z calcd for C48H82N5O11 [M +
H]+: 904.6, [M + 2H]2+/2: 452.8, found: 904.8, 453.2.
5.1.2.15. 13(R)-[1-(4-p-Ethoxyphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (20j). White solid, yield 24%, mp:
95–98 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.78–7.64 (m, 3H), 6.94 (d, J = 8.8 Hz, 2H),
5.20 (s, 1H), 4.84 (s, 1H), 4.66–4.56 (m, 2H), 4.45 (d, J = 7.1 Hz, 1H),
4.09–4.02 (m, 3H), 3.84 (s, 1H), 3.59–3.47 (m, 2H), 3.34–3.13 (m, 8H),
3.02–2.96 (m, 2H), 2.90–2.86 (m, 1H), 2.81–2.21 (m, 14H), 2.20–2.00
(m, 3H), 1.96–1.92 (m, 1H), 1.85–1.72 (m, 2H), 1.53 (d, J = 4.2 Hz, 1H),
1.43 (t, J = 7.0 Hz, 3H), 1.38–1.17 (m, 16H), 1.14 (d, J = 6.5 Hz, 3H),
1.01–1.00 (m, 3H), 0.96–0.95 (m, 3H); MS (ESI) m/z calcd for
5.1.2.12. 13(S)-[1-(4-p-n-Amylphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (20g). White solid, yield 27%, mp:
94–96 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.73 (d, J = 8.0 Hz, 3H), 7.24 (d, J = 8.2 Hz,
2H), 5.53–5.51 (m, 1H), 4.84–4.83 (m, 1H), 4.56 (dd, J = 14.4, 4.5 Hz,
1H), 4.49 (dd, J = 14.5, 5.2 Hz, 1H), 4.41 (d, J = 6.9 Hz, 1H), 4.30 (s,
1H), 4.03–4.01 (m, 1H), 3.72 (d, J = 7.5 Hz, 1H), 3.50 (d, J = 7.3 Hz,
2H), 3.37–3.16 (m, 8H), 2.99–2.92 (m, 2H), 2.70 (d, J = 12.7 Hz, 1H),
2.64–2.62 (m, 3H), 2.56–2.18 (m, 14H), 2.16–2.08 (m, 2H), 2.01 (d, J =
10.8 Hz, 1H), 1.88–1.85 (m, 1H), 1.75–1.74 (m, 2H), 1.65–1.61 (m, 2H),
1.45 (dd, J = 15.1, 4.6 Hz, 1H), 1.35–1.20 (m, 20H), 1.03 (d, J = 6.3 Hz,
3H), 0.90 (t, J = 7.0 Hz, 3H), 0.84 (d, J = 7.0 Hz, 3H), 0.72 (d, J = 6.6
Hz, 3H); 13C NMR (100 MHz, CDCl3, δ ppm): 175.94, 147.89, 143.03,
128.69, 127.53, 125.42, 119.86, 102.95, 96.01, 79.91, 79.39, 78.63,
77.85, 72.49, 70.82, 68.29, 65.29, 64.98, 62.77, 58.86, 51.44, 49.91,
49.20, 45.03, 43.95, 40.22, 35.48, 34.74, 32.31, 31.23, 30.82, 30.33,
22.28, 21.98, 21.29, 20.98, 18.04, 14.46, 13.77, 10.45; MS (ESI) m/z
calcd for C50H86N5O11 [M + H]+: 932.6, [M + 2H]2+/2: 466.8, found:
932.8, 467.3.
C
47H81N5O12 [M + 2H]2+/2: 453.8, found: 454.1.
5.1.2.16. 13(S)-[1-(4-p-Propoxyphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (20k). White solid, yield 39%, mp:
104–106 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.73 (d, J = 8.7 Hz, 2H), 7.68 (s, 1H),
6.96 (d, J = 8.7 Hz, 2H), 5.53 (d, J = 10.0 Hz, 1H), 4.79–4.78 (m, 1H),
4.57–4.45 (m, 3H), 4.26 (s, 1H), 3.96 (t, J = 6.6 Hz, 3H), 3.72 (d, J = 6.6
Hz, 1H), 3.53–3.52 (m, 2H), 3.27–3.21 (m, 8H), 3.01–2.99 (m, 2H),
2.92–2.89 (m, 1H), 2.78 (s, 3H), 2.74–2.72 (m, 1H), 2.55–2.21 (m, 10H),
2.17–2.05 (m, 3H), 1.85–1.76 (m, 5H), 1.47–1.46 (m, 1H), 1.28–1.23
(m, 16H), 1.19 (d, J = 7.5 Hz, 3H), 1.05 (t, J = 7.4 Hz, 3H), 0.83 (d, J =
6.9 Hz, 3H), 0.72 (d, J = 6.6 Hz, 3H); 13C NMR (100 MHz, CDCl3, δ
ppm): 175.90, 159.12, 147.73, 126.78, 122.63, 119.39, 114.69, 102.64,
96.00, 79.85, 79.70, 78.59, 77.80, 72.55, 70.72, 69.41, 68.07, 67.64,
65.37, 65.05, 62.84, 58.83, 51.41, 49.94, 49.21, 44.95, 43.92, 41.04,
40.37, 36.70, 34.71, 32.30, 31.24, 22.35, 22.03, 21.29, 20.87, 18.05,
14.54, 10.50, 10.27; MS (ESI) m/z calcd for C48H83N5O12 [M + 2H]2+/2:
460.8, found: 460.9.
5.1.2.13. 13(R)-[1-(4-p-n-Amylphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (20h). White solid, yield 54%, mp:
102–104 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.73 (dd, J = 13.8, 7.2 Hz, 3H), 7.23 (d,
J = 8.1 Hz, 2H), 5.18 (s, 1H), 4.82 (s, 1H), 4.73–4.58 (m, 3H), 4.43 (d, J
= 6.7 Hz, 1H), 4.04–4.02 (m, 1H), 3.82 (s, 1H), 3.59–3.49 (m, 2H),
3.33–3.20 (m, 8H), 2.98–2.86 (m, 3H), 2.64–2.61 (m, 2H), 2.54–2.16
(m, 14H), 2.16–1.94 (m, 3H), 1.94–1.92 (m, 1H), 1.88–1.81 (m, 2H),
1.64–1.62 (m, 2H), 1.54 (s, 1H), 1.35–1.20 (m, 20H), 1.15 (d, J = 6.9 Hz,
3H), 1.03 (d, J = 6.0 Hz, 3H), 0.97 (d, J = 6.7 Hz, 3H), 0.90 (t, J = 6.9
Hz, 3H); 13C NMR (100 MHz, CDCl3, δ ppm): 175.52, 147.72, 142.93,
128.64, 127.65, 125.43, 119.84, 103.11, 96.30, 80.17, 79.11, 77.79,
77.63, 72.42, 71.54, 70.84, 68.32, 65.84, 65.18, 51.18, 49.88, 49.19,
45.10, 40.22, 38.14, 36.89, 35.47, 34.83, 32.65, 31.22, 30.83, 30.21,
22.28, 21.32, 21.10, 20.92, 18.39, 13.77, 9.96; MS (ESI) m/z calcd for
5.1.2.17. 13(R)-[1-(4-p-Propoxyphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (20l). White solid, yield 39%, mp:
88–90 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.74–7.63 (m, 3H), 6.95 (dd, J = 8.9, 2.5 Hz,
2H), 5.15 (s, 1H), 4.81 (s, 1H), 4.68–4.57 (m, 2H), 4.42 (d, J = 7.2 Hz,
1H), 4.06–4.00 (m, 1H), 3.95 (t, J = 6.6 Hz, 2H), 3.83–3.76 (m, 1H),
3.60–3.43 (m, 2H), 3.35–3.18 (m, 8H), 2.99–2.90 (m, 3H), 2.62–2.16
(m, 16H), 2.11–2.10 (m, 1H), 1.94–1.91 (m, 1H), 1.88–1.81 (m, 4H),
1.52–1.49 (m, 1H), 1.32–1.21 (m, 16H), 1.16–1.12 (m, 3H), 1.05 (t, J =
7.4 Hz, 6H), 0.99–0.97 (m, 3H); MS (ESI) m/z calcd for C48H83N5O12 [M
+ 2H]2+/2: 460.8, found: 461.2.
5.1.3. Synthesis of C12-substituted 15-membered homoclarithromycin
derivatives (Series C)
C
50H86N5O11 [M + H]+: 932.6, [M + 2H]2+/2: 466.8, found: 932.8,
467.2.
5.1.3.1. (S)-Methyl 3-azido-2-[(tert-butoxycarbonyl)amino]propanoate
(37). MsC l (10 mL 1 M solution in DCM 10 mmol) was added dropwise
to a solution of N-Boc-L-serine methyl ester 35 (2 g 9.1 mmol) and Et3N
5.1.2.14. 13(S)-[1-(4-p-Ethoxyphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (20i). White solid, yield 54%, mp:
104–106 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.74 (d, J = 8.7 Hz, 2H), 7.68 (s, 1H),
6.95 (d, J = 8.7 Hz, 2H), 5.52 (d, J = 9.2 Hz, 1H), 4.82 (s, 1H), 4.55 (dd,
◦
(1.1 g 10.9 mmol) in dry DCM (50 mL) at 0 C under nitrogen atmo-
sphere. The reaction mixture was stirred at 0 ◦C for 0.5 h and then H2O
(50 mL) was added. The organic phase was separated and aqueous layer
19

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
was extracted with DCM (50 mL × 2). The combined organic layers were
dried over anhydrous Na2SO4 filtered and evaporated in vacuo to form
the methanesulfonate 36 as a colorless oil. NaN3 (1.4 g 21.5 mmol) was
added to a solution of methanesulfonate 36 in DMF (60 mL) at room
temperature. The reaction mixture was stirred at 50 ◦C for 0.5 h and then
H2O (60 mL) and EtOAc (60 mL) was added. The organic phase was
separated and aqueous layer was extracted with EtOAc (60 mL × 2). The
combined organic layers were dried over anhydrous Na2SO4 filtered and
evaporated in vacuo to provide the azide 37 (8.74 g) as a colorless oil.
The crude intermediate 37 was used directly for the next step without
purification.
chromatographically (DCM/CH3OH/NH3⋅H2O = 30:1:0.1) to yield the
crude ester 31a as a colorless oil.
LiOH (44 mg, 1.83 mmol) was added to a solution of the above 31a in
the mixed solvents of THF (15 mL), EtOH (5 mL) and H2O (5 mL) and the
reaction mixture was stirred at room temperature for 6 h. The reaction
mixture was evaporated in vacuo to dryness and the residue was sepa-
rated chromatographically (DCM/CH3OH/NH3⋅H2O = 15:1:0.1 ~
10:1:0.1) to give the carboxylic acid 32a (70 mg, 15% in four steps from
intermediate 3) as a white solid. Mp: 83–86 ◦C, TLC Rf = 0.42 (DCM/
CH3OH/NH3⋅H2O = 15:1:0.1); 1H NMR (600 MHz, CDCl3, δ ppm): 7.80
(s, 1H), 7.70 (d, J = 8.1 Hz, 2H), 7.23 (d, J = 8.1 Hz, 2H), 4.75 (s, 1H),
4.68–4.60 (m, 1H), 4.55–4.49 (m, 1H), 4.44–4.35 (m, 1H), 4.29–4.26
(m, 1H), 4.04–3.89 (m, 2H), 3.84–3.76 (m, 1H), 3.60–3.56 (m, 1H), 3.37
(s, 1H), 3.31 (s, 3H), 3.24–3.10 (m, 5H), 2.87–2.58 (m, 5H), 2.44 (s, 3H),
2.38 (s, 3H), 2.33–2.11 (m, 8H), 2.07–1.99 (m, 1H), 1.93–1.86 (m, 1H),
1.77–1.72 (m, 1H), 1.71–1.52 (m, 2H), 1.49 (dd, J = 14.9, 4.9 Hz, 1H),
1.26 (s, 3H), 1.24–1.20 (m, 4H), 1.18–1.05 (m, 13H), 0.98–0.86 (m,
33H), 0.69–0.55 (m, 18H); MS (ESI) m/z calcd for C64H123N5O12Si3 [M
+ 2H]2+/2: 618.9, found: 619.5.
5.1.3.2. (S)-Methyl
2-[(tert-butoxycarbonyl)amino]-3-[4-(p-tolyl)-1H-
1,2,3-triazol-1-yl]propanoate (38). 37 (8.74 g) in 75% methanol solu-
tion (20 mL) was reacted with 4-ethynyltoluene 23 (0.48 g, 4.14 mmol)
in the presence of CuSO4 (82 mg) and L-ascorbic acid sodium salt (246
◦
mg) at 35 ~ 40 C for 48 h. The reaction mixture was evaporated in
vacuo and the residue was separated chromatographically (PE/EtOAc =
4:1 ~ 1:1) to form the 1,2,3-triazole 38 (0.92 g, 62%) as a white solid.
1
◦
Mp: 146–149 C, TLC Rf = 0.58 (DCM/MeOH = 30:1); H NMR (600
MHz, CDCl3, δ ppm): 7.70 (d, J = 8.3 Hz, 3H), 7.24 (d, J = 8.1 Hz, 2H),
5.41 (d, J = 6.4 Hz, 1H), 4.91 (dd, J = 13.9, 3.9 Hz, 1H), 4.83 (dd, J =
14.0, 4.4 Hz, 1H), 4.76–4.75 (m, 1H), 3.80 (s, 3H), 2.38 (s, 3H), 1.44 (s,
9H); MS (ESI) m/z calcd for C18H25N4O4 [M + H]+: 361.2, found: 361.4.
5.1.3.5. 15-Membered 11a-homo-aza-clarithromycin intermediate (33a).
To a suspension of 32a (70 mg, 0.06 mmol) and Et3N (58 mg, 0.57
mmol) in THF (5 mL) was added 2,4,6-trichlorobenzoyl chloride (42 mg,
0.17 mmol) at room temperature. The reaction solution was stirred at
room temperature for 4 h under nitrogen atmosphere, and was added to
a refluxed solution of DMAP (175 mg, 1.42 mmol) in CH3CN (30 mL) for
0.5 h. The reaction mixture was concentrated in vacuo to dryness, and
then DCM (20 mL) and saturated NH4Cl solution (20 mL) were added.
The organic phase was separated and aqueous layer was extracted with
DCM (20 mL × 2). The combined organic layers were dried over
anhydrous Na2SO4, filtered, and evaporated in vacuo. The residue was
separated chromatographically (PE/acetone = 15:1 ~ 10:1) to afford
the cyclic intermediate 33a (50 mg, 72%) as a white solid. Mp: 81–84 ◦C,
TLC Rf = 0.53 (PE/acetone = 3:1); 1H NMR (600 MHz, CDCl3, δ ppm):
7.77–7.67 (m, 3H), 7.23 (d, J = 7.8 Hz, 2H), 4.71 (s, 1H), 4.47–4.38 (m,
3H), 4.30–4.25 (m, 1H), 4.21–4.14 (m, 1H), 4.10–4.04 (m, 1H), 3.67 (s,
1H), 3.57–3.54 (m, 1H), 3.52–3.49 (m, 1H), 3.28 (s, 3H), 3.21–3.17 (m,
5H), 2.71–2.60 (m, 2H), 2.53–2.24 (m, 11H), 2.24–2.19 (m, 6H),
2.06–2.01 (m, 1H), 1.78–1.72 (s, 2H), 1.61–1.55 (m, 2H), 1.44–1.40 (m,
1H), 1.26 (s, 3H), 1.24–1.22 (m, 1H), 1.22–1.20 (m, 3H), 1.18–1.12 (m,
10H), 1.09–1.08 (m, 3H), 1.00–0.92 (m, 33H), 0.68–0.55 (m, 18H); MS
(ESI) m/z calcd for C64H121N5O11Si3 [M + 2H]2+/2: 609.9, found:
610.4.
5.1.3.3. (S)-2-Amino-3-[4-(p-tolyl)-1H-1,2,3-triazol-1-yl]propan-1-ol
(30a). NaBH4 (284 mg, 7.5 mmol) was added to a solution of 1,2,3-tri-
azole 38 (0.9 g, 2.5 mmol) in MeOH (20 mL) at 0 ◦C. The reaction so-
lution was stirred at 0 ◦C for 2 h under nitrogen atmosphere. The
reaction mixture was concentrated in vacuo, and then H2O (20 mL) and
DCM (20 mL) were added. The organic phase was separated and aqueous
layer was extracted with DCM (20 mL × 2). The combined organic layers
were dried over anhydrous Na2SO4, filtered and evaporated in vacuo to
obtain the primary alcohol 39 as a white solid.
The concentrated HCl (4 mL, 48 mmol) was added to a solution of
primary alcohol 39 in EtOAc (10 mL) at room temperature. The reaction
mixture was stirred at room temperature for 4 h, and then MeOH (4 mL)
and NaHCO3 (5 g, 59.5 mmol) was added. The resulting mixture was
stirred at room temperature for 0.5 h and was evaporated in vacuo to
dryness. The residue was separated chromatographically (DCM/
CH3OH/NH3⋅H2O = 10:1:0.1) to form the substituted ethanolamine 30a
(0.35 g, 60% in two steps from intermediate 38) as a white solid. Mp:
135–138 ◦C, TLC Rf = 0.36 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, DMSO‑d6, δ ppm): 8.46 (s, 1H), 7.73 (d, J = 8.1 Hz, 2H),
7.25 (d, J = 7.9 Hz, 2H), 4.44 (dd, J = 13.6, 4.9 Hz, 1H), 4.18 (dd, J =
13.6, 7.8 Hz, 1H), 3.32–3.29 (m, 2H), 3.13–3.09 (m, 1H), 2.33 (s, 3H),
1.90 (s, 1H); MS (ESI) m/z calcd for C12H17N4O [M + H]+: 233.1, found:
233.3.
5.1.3.6. 12(S)-[1-(4-p-Methylphenyl-1H-1,2,3-triazol-1-yl)-1-methylene]-
9(S)-hydroxyclarithromycin (34a). To a solution of 33a (50 mg, 0.04
mmol) in THF (5 mL) was added hydrogen fluoride–pyridine (19 mg,
0.12 mmol, 65%) at room temperature, and the resulting mixture was
stirred at room temperature for 18 h. The reaction mixture was
quenched with saturated NaHCO3 solution (20 mL) and diluted with
DCM (20 mL). The organic phase was separated and aqueous layer was
extracted with DCM (20 mL × 2). The combined organic layers were
dried over anhydrous Na2SO4, filtered, and evaporated in vacuo. The
residue was separated chromatographically (DCM/CH3OH/NH3⋅H2O =
10:1:0.1) to obtain compound 34a (15 mg, 42%) as a white solid. Mp:
91–94 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.85 (s, 1H), 7.71 (d, J = 8.0 Hz, 2H), 7.24 (d,
J = 8.0 Hz, 2H), 4.75–4.74 (m, 1H), 4.57 (dd, J = 14.2, 6.6 Hz, 1H), 4.52
(d, J = 7.2 Hz, 1H), 4.41 (dd, J = 14.0, 7.4 Hz, 1H), 4.28 (d, J = 11.9 Hz,
1H), 4.17 (s, 1H), 4.05–3.99 (m, 2H), 3.84 (d, J = 4.3 Hz, 1H), 3.49–3.48
(m, 2H), 3.27 (s, 3H), 3.18–3.15 (m, 4H), 3.03 (d, J = 8.8 Hz, 1H),
2.95–2.88 (m, 2H), 2.46–2.38 (m, 11H), 2.32–2.21 (m, 6H), 2.06–1.95
(m, 5H), 1.78–1.76 (m, 1H), 1.58 (dd, J = 14.8, 4.9 Hz, 1H), 1.26–1.25
(m, 13H), 1.17 (d, J = 1.9 Hz, 3H), 1.02 (d, J = 7.3 Hz, 3H), 0.91 (d, J =
6.1 Hz, 3H), 0.76 (d, J = 6.3 Hz, 3H); 13C NMR (150 MHz, CDCl3, δ
5.1.3.4. Ring-opening intermediate with carboxylic acid (32a). To a so-
lution of 3 (400 mg, 0.37 mmol) in anhydrous CHCl3 (20 mL) was added
Pb(OAc)4 (195 mg, 0.44 mmol) at 0 ◦C. The reaction mixture was stirred
◦
at 0 C for 0.5 h under nitrogen atmosphere. The aldehyde 4 was ob-
tained in a high yield and was directly used in the next step without
purification.
To a solution of 4 were added 30a (170 mg, 0.73 mmol) and NaBH
(OAc)3 (233 mg, 1.1 mmol) at 0 ◦C. The reaction mixture was stirred at
room temperature for 4 h under nitrogen atmosphere. Then, NaBH
(OAc)3 (233 mg, 1.1 mmol) and 37% aqueous formaldehyde solution
(179 mg, 2.2 mmol) were added and stirred at room temperature for
another 4 h under nitrogen atmosphere. The reaction mixture was
quenched with saturated NaHCO3 solution (20 mL). The organic phase
was separated and aqueous layer was extracted with DCM (20 mL × 2).
The combined organic layers were dried over anhydrous Na2SO4,
filtered, and evaporated in vacuo. The residue was separated
20

Y. Qin et al.
Bioorganic Chemistry 113 (2021) 104992
ppm): 175.60, 148.13, 138.23, 129.59, 127.51, 125.63, 120.10, 80.60,
77.46, 72.90, 71.66, 70.59, 68.30, 68.10, 68.05, 65.62, 63.64, 49.43,
46.08, 43.87, 35.94, 34.68, 31.91, 30.31, 29.78, 29.70, 29.61, 29.52,
29.32, 29.24, 27.22, 25.54, 22.69, 21.69, 21.30, 20.90, 20.79, 18.87,
18.42, 14.12; HRMS (ESI) m/z calcd for C46H78N5O11 [M + H]+:
876.5698, [M + 2H]2+/2: 438.7888, found: 876.5682, 438.7885.
According to the synthetic procedure of compound 34a, the other
compounds (34b-34l) in Series C were also prepared.
(600 MHz, CDCl3, δ ppm): 7.86 (s, 1H), 7.73 (d, J = 8.1 Hz, 2H), 7.24 (d,
J = 8.1 Hz, 2H), 4.74 (s, 1H), 4.58 (dd, J = 14.0, 6.5 Hz, 1H), 4.52 (d, J
= 7.2 Hz, 1H), 4.41 (dd, J = 14.1, 7.3 Hz, 2H), 4.28 (d, J = 10.4 Hz, 1H),
4.20 (s, 1H), 4.03–3.94 (m, 3H), 3.85 (d, J = 3.8 Hz, 1H), 3.51–3.48 (m,
2H), 3.28–3.24 (m, 4H), 3.15 (s, 3H), 3.04–3.03 (m, 1H), 2.98–2.93 (m,
2H), 2.63–2.60 (m, 2H), 2.45–2.17 (m, 14H), 2.05–1.99 (m, 3H),
1.96–1.95 (m, 1H), 1.79–1.76 (m, 2H), 1.67–1.65 (m, 2H), 1.58–1.57
(m, 1H), 1.28 (s, 3H), 1.25–1.24 (m, 10H), 1.16 (d, J = 7.3 Hz, 3H), 1.01
(d, J = 7.2 Hz, 3H), 0.96–0.95 (m, 3H), 0.87 (d, J = 7.2 Hz, 3H),
0.76–0.75 (m, 3H); MS (ESI) m/z calcd for C48H83N5O11 [M + 2H]2+/2:
452.8, found: 453.2.
5.1.3.7. 12(R)-[1-(4-p-Methylphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (34b). White solid, yield 83%, mp:
99–102 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.78 (s, 1H), 7.67 (d, J = 8.0 Hz, 2H), 7.23 (d,
J = 8.0 Hz, 2H), 4.85 (d, J = 3.8 Hz, 1H), 4.69 (s, 1H), 4.52 (d, J = 7.0
Hz, 1H), 4.40 (s, 2H), 4.06 (d, J = 7.3 Hz, 1H), 4.01–3.96 (m, 1H), 3.75
(d, J = 7.3 Hz, 2H), 3.61–3.53 (m, 2H), 3.30–3.25 (m, 7H), 3.06–3.03
(m, 3H), 2.90–2.86 (m, 2H), 2.86–2.82 (m, 3H), 2.57 (s, 1H), 2.48 (s,
3H), 2.38 (s, 3H), 2.34–2.16 (m, 5H), 1.92–1.84 (m, 2H), 1.66–1.51 (m,
7H), 1.30–1.26 (m, 13H), 1.19 (d, J = 7.2 Hz, 3H), 1.12 (d, J = 6.9 Hz,
3H), 0.83–0.81 (m, 6H); 13C NMR (150 MHz, CDCl3, δ ppm): 175.73,
147.67, 138.24, 129.58, 127.56, 125.60, 120.38, 102.19, 96.41, 80.23,
80.15, 78.59, 77.80, 72.90, 71.04, 68.30, 68.11, 67.98, 65.99, 65.26,
62.69, 61.56, 50.58, 49.39, 44.59, 35.13, 29.71, 29.33, 21.56, 21.31,
20.99, 18.87, 18.32, 14.80, 10.62; HRMS (ESI) m/z calcd for
5.1.3.11. 12(R)-[1-(4-p-n-Propylphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (34f). White solid, yield 77%, mp:
106–108 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.78 (s, 1H), 7.70 (d, J = 8.0 Hz, 2H),
7.23 (d, J = 8.0 Hz, 2H), 4.86 (d, J = 4.3 Hz, 1H), 4.60 (s, 1H), 4.50 (d, J
= 7.0 Hz, 1H), 4.41–4.36 (m, 2H), 4.07–3.99 (m, 2H), 3.75 (d, J = 7.3
Hz, 1H), 3.47–3.39 (m, 2H), 3.30–3.25 (m, 7H), 3.05–3.02 (m, 1H),
2.92–2.86 (m, 2H), 2.63–2.60 (m, 2H), 2.48–2.21 (m, 11H), 2.08–1.91
(m, 3H), 1.73–1.56 (m, 9H), 1.32 (s, 3H), 1.30–1.25 (m, 10H), 1.18 (d, J
= 6.7 Hz, 3H), 1.10 (d, J = 6.9 Hz, 3H), 0.95 (d, J = 7.4 Hz, 3H),
0.87–0.81 (m, 6H); 13C NMR (100 MHz, CDCl3, δ ppm): 175.66, 147.56,
142.70, 128.72, 127.65, 125.39, 120.05, 102.34, 95.92, 80.09, 79.75,
78.58, 77.69, 72.63, 70.81, 68.04, 65.61, 65.07, 62.38, 61.58, 50.31,
49.16, 44.43, 40.45, 40.30, 37.57, 36.18, 34.88, 31.89, 30.81, 29.44,
24.19, 21.31, 20.94, 18.09, 14.29, 13.51, 10.15; MS (ESI) m/z calcd for
C
46H78N5O11 [M + H]+: 876.5698, [M + 2H]2+/2: 438.7888, found:
876.5658, 438.7889.
C
48H83N5O11 [M + 2H]2+/2: 452.8, found: 453.1.
5.1.3.8. 12(S)-[1-(4-p-Ethylphenyl-1H-1,2,3-triazol-1-yl)-1-methylene]-9
(S)-hydroxyclarithromycin (34c). White solid, yield 24%, mp: 92–94 ◦C,
TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR (600 MHz,
CDCl3, δ ppm): 7.83 (s, 1H), 7.74–7.73 (m, 3H), 7.25 (s, 1H), 4.77 (d, J
= 4.1 Hz, 1H), 4.53 (dd, J = 13.6, 6.7 Hz, 1H), 4.48 (d, J = 7.1 Hz, 1H),
4.39 (dd, J = 13.9, 7.4 Hz, 2H), 4.22 (d, J = 11.7 Hz, 1H), 4.15 (s, 1H),
4.05–3.96 (m, 3H), 3.80 (d, J = 5.1 Hz, 1H), 3.49–3.48 (m, 2H),
3.29–3.25 (m, 4H), 3.16 (s, 3H), 3.00–2.99 (m, 1H), 2.97–2.89 (m, 2H),
2.70–2.66 (m, 2H), 2.48–2.27 (m, 14H), 2.02–1.99 (m, 3H), 1.81–1.71
(m, 3H), 1.58–1.53 (m, 1H), 1.30 (s, 3H), 1.27–1.23 (m, 13H), 1.17 (d, J
= 7.3 Hz, 3H), 1.05 (d, J = 6.9 Hz, 3H), 0.92 (d, J = 6.6 Hz, 3H),
0.81–0.80 (m, 3H); 13C NMR (150 MHz, CDCl3, δ ppm): 175.44, 147.96,
144.34, 128.25, 127.84, 125.63, 119.94, 103.01, 95.70, 95.51, 80.61,
78.64, 78.46, 77.92, 77.81, 72.72, 70.79, 68.64, 65.77, 65.41, 61.84,
50.15, 49.34, 49.23, 47.73, 44.05, 40.33, 34.92, 34.61, 34.16, 28.58,
21.54, 21.51, 21.15, 21.11, 20.76, 18.13, 18.07, 15.40, 13.02, 10.96;
MS (ESI) m/z calcd for C47H81N5O11 [M + 2H]2+/2: 445.8, found: 446.1.
5.1.3.12. 12(S)-[1-(4-p-n-Amylphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (34g). White solid, yield 30%, mp:
96–98 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.83 (s, 1H), 7.73 (d, J = 8.1 Hz, 2H), 7.24 (d,
J = 8.1 Hz, 2H), 4.77–4.76 (m, 1H), 4.54 (dd, J = 12.1, 7.0 Hz, 1H), 4.48
(d, J = 7.0 Hz, 1H), 4.39 (dd, J = 14.0, 7.4 Hz, 1H), 4.23 (d, J = 10.8 Hz,
1H), 4.16 (s, 1H), 4.08–3.94 (m, 3H), 3.81 (s, 1H), 3.50–3.45 (m, 2H),
3.28 (s, 3H), 3.21–3.16 (m, 4H), 3.02–3.00 (m, 1H), 2.96–2.89 (m, 2H),
2.64–2.61 (m, 2H), 2.48–2.21 (m, 12H), 2.06–1.98 (m, 3H), 1.81–1.75
(m, 2H), 1.68–1.61 (m, 6H), 1.34–1.32 (m, 4H), 1.29 (s, 3H), 1.27–1.21
(m, 10H), 1.17 (d, J = 7.3 Hz, 3H), 1.05 (d, J = 6.2 Hz, 3H), 0.93–0.88
(m, 6H), 0.83–0.80 (m, 3H); MS (ESI) m/z calcd for C50H87N5O11 [M +
2H]2+/2: 466.8, found: 467.2.
5.1.3.13. 12(R)-[1-(4-p-n-Amylphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (34h). White solid, yield 74%, mp:
102–104 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.78 (s, 1H), 7.71 (d, J = 8.1 Hz, 2H),
7.23 (d, J = 8.1 Hz, 2H), 4.87 (d, J = 4.4 Hz, 1H), 4.60–4.54 (m, 1H),
4.49 (d, J = 7.1 Hz, 1H), 4.45–4.41 (m, 1H), 4.36–4.32 (m, 1H),
4.06–4.01 (m, 2H), 3.81 (d, J = 7.7 Hz, 1H), 3.75 (d, J = 7.4 Hz, 2H),
3.58–3.49 (m, 2H), 3.31–3.25 (m, 7H), 3.04 (s, 1H), 2.90–2.88 (m, 2H),
2.64–2.61 (m, 2H), 2.48–2.25 (m, 12H), 2.07 (s, 2H), 1.96–1.80 (m, 4H),
1.68–1.55 (m, 5H), 1.33–1.24 (m, 17H), 1.18 (d, J = 7.2 Hz, 3H), 1.10
(d, J = 7.1 Hz, 3H), 0.91–0.84 (m, 9H); 13C NMR (100 MHz, CDCl3, δ
ppm): 175.73, 147.52, 142.90, 128.61, 127.55, 125.35, 120.01, 102.37,
95.82, 80.06, 79.71, 78.57, 77.68, 72.57, 70.77, 67.94, 65.50, 64.83,
62.29, 61.60, 50.27, 49.11, 44.40, 40.52, 40.02, 36.11, 35.41, 34.81,
31.16, 30.75, 30.53, 22.22, 21.25, 20.92, 18.07, 14.23, 13.72, 10.03;
MS (ESI) m/z calcd for C50H87N5O11 [M + 2H]2+/2: 466.8, found: 467.1.
5.1.3.9. 12(R)-[1-(4-p-Ethylphenyl-1H-1,2,3-triazol-1-yl)-1-methylene]-9
(S)-hydroxyclarithromycin (34d). White solid, yield 52%, mp: 92–95 ◦C,
TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR (600 MHz,
CDCl3, δ ppm): 7.79 (s, 1H), 7.72–7.70 (m, 3H), 7.25 (s, 1H), 4.87 (d, J
= 4.2 Hz, 1H), 4.61 (s, 1H), 4.50 (d, J = 7.0 Hz, 1H), 4.45–4.36 (m, 3H),
4.07–4.00 (m, 3H), 3.75 (d, J = 7.5 Hz, 1H), 3.44–3.38 (m, 2H),
3.30–3.25 (m, 7H), 3.05–3.02 (m, 1H), 2.91–2.86 (m, 2H), 2.70–2.66
(m, 2H), 2.48–2.16 (m, 14H), 2.06–1.91 (m, 3H), 1.87–1.77 (m, 3H),
1.56 (d, J = 4.7 Hz, 1H), 1.32 (s, 3H), 1.27–1.23 (m, 13H), 1.19 (d, J =
6.9 Hz, 3H), 1.10 (d, J = 6.9 Hz, 3H), 0.87–0.83 (m, 6H); 13C NMR (150
MHz, CDCl3, δ ppm): 175.79, 147.68, 144.43, 128.26, 127.77, 125.62,
120.18, 102.45, 96.08, 80.22, 79.88, 78.70, 77.81, 72.76, 70.96, 68.19,
68.01, 65.77, 65.20, 62.53, 61.68, 50.47, 49.30, 44.55, 40.55, 36.29,
35.01, 32.04, 29.59, 28.57, 21.46, 21.07, 18.76, 18.23, 15.39, 14.46,
10.32; MS (ESI) m/z calcd for C47H81N5O11 [M + 2H]2+/2: 445.8, found:
446.1.
5.1.3.14. 12(S)-[1-(4-p-Ethoxyphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (34i). White solid, yield 35%, mp:
106–108 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.78 (s, 1H), 7.73 (d, J = 8.8 Hz, 2H),
6.95 (d, J = 8.8 Hz, 2H), 4.76–4.75 (m, 1H), 4.57–4.53 (m, 1H), 4.50 (d,
5.1.3.10. 12(S)-[1-(4-p-n-Propylphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (34e). White solid, yield 34%, mp:
84–87 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
21

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Bioorganic Chemistry 113 (2021) 104992
J = 7.2 Hz, 1H), 4.41–4.37 (m, 1H), 4.26 (d, J = 11.3 Hz, 1H), 4.17 (s,
1H), 4.07 (d, J = 7.0 Hz, 2H), 4.03–3.94 (m, 4H), 3.83 (d, J = 4.8 Hz,
1H), 3.49–3.47 (m, 2H), 3.27–3.25 (m, 4H), 3.15 (s, 3H), 3.03–3.01 (m,
1H), 2.95–2.89 (m, 2H), 2.49–2.21 (m, 14H), 2.01–1.94 (m, 3H),
1.80–1.75 (m, 3H), 1.58 (d, J = 5.0 Hz, 1H), 1.43 (t, J = 7.0 Hz, 3H),
1.27–1.24 (m, 13H), 1.17 (d, J = 7.3 Hz, 3H), 1.03 (d, J = 7.3 Hz, 3H),
0.91 (d, J = 6.2 Hz, 3H), 0.78 (d, J = 6.1 Hz, 3H); MS (ESI) m/z calcd for
and incubated for 24 h at 37 ◦C. The turbidity of each well was observed,
and the MIC was determined as the concentration at which no visible
bacterial proliferation was seen. The solution (10 µL), from wells where
no visible bacterial proliferation was seen (in the MIC 96-well plates),
onto culture medium plates. The plates were subcultured for 24 h at
37 ◦C. The MBC was determined as the lowest concentration at which no
bacterial colonies were seen.
C
47H81N5O12 [M + 2H]2+/2: 453.8, found: 454.1.
5.3. Docking methods
5.1.3.15. 12(R)-[1-(4-p-Ethoxyphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (34j). White solid, yield 63%, mp:
102–104 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H
NMR (600 MHz, CDCl3, δ ppm): 7.74–7.69 (m, 3H), 6.94 (d, J = 7.0 Hz,
2H), 4.86 (d, J = 4.2 Hz, 1H), 4.61 (s, 1H), 4.50 (d, J = 7.1 Hz, 1H), 4.38
(d, J = 28.9 Hz, 2H), 4.09–3.99 (m, 4H), 3.79–3.75 (m, 2H), 3.50–3.39
(m, 2H), 3.33–3.17 (m, 7H), 3.04–3.03 (m, 1H), 2.89–2.82 (m, 2H),
2.82–2.58 (m, 5H), 2.48 (s, 3H), 2.39–2.18 (m, 5H), 2.04–1.89 (m, 2H),
1.86–1.67 (m, 5H), 1.56 (d, J = 4.7 Hz, 1H), 1.44 (t, J = 7.0 Hz, 3H),
1.32–1.24 (m, 13H), 1.18 (d, J = 7.1 Hz, 3H), 1.10 (d, J = 6.9 Hz, 3H),
0.86–0.82 (m, 6H); 13C NMR (100 MHz, CDCl3, δ ppm): 175.65, 158.83,
147.34, 126.75, 122.73, 119.60, 114.62, 102.14, 95.94, 80.00, 79.86,
78.47, 77.64, 72.64, 70.71, 67.82, 65.62, 65.05, 63.30, 62.35, 61.51,
50.28, 49.13, 44.40, 36.19, 34.86, 32.01, 31.43, 21.28, 21.00, 20.84,
18.07, 14.54, 14.34, 10.18; MS (ESI) m/z calcd for C47H81N5O12 [M +
2H]2+/2: 453.8, found: 453.7.
The crystal structure of the E. coli ribosome bound to the peptidyl
transferase center was obtained from the Protein Data Bank (code:
4V7S). Compounds 9epre, 11g and 20c were docked by Ledock, and
CAM was used as the control compound. The alkyl group of compounds
11g were docked into binding site, which may bind to bacterial ribo-
some by the π-π stacking, electrostatic and hydrophobic interaction. The
binding conformation was given by using the conformation search
function of simulated annealing-genetic algorithm.
5.4. Time-bactericidal assay
S. pneumoniae AB11 cells in log-phase were diluted to approximate
106 CFU/mL and exposed to a series of concentrations of compound 11g
(0.5 × MIC, 1 × MIC, 2 × MIC and 4 × MIC, MIC = 4 µg/mL) in CAMH
broth. The solution were obtained from each treatment after 0, 3, 6, 9,
12 and 24 h of incubation at 37 ◦C. The solution was serially diluted in
CAMH broth by using 10-fold dilution method. Therefore, 10 concen-
trations of bacterial solution were obtained and were plated onto TSA
plates. This plates were incubated for 24 h at 37 ◦C prior to viable CFU/
mL was provided.
5.1.3.16. 12(S)-[1-(4-p-Propoxyphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (34k). White solid, yield 47%, mp:
88–91 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.78–7.72 (m, 3H), 6.96 (d, J = 8.8 Hz, 2H),
4.75 (s, 1H), 4.59–4.47 (m, 3H), 4.39 (dd, J = 14.1, 7.5 Hz, 1H),
4.31–4.27 (m, 1H), 4.23–4.16 (m, 1H), 4.03–4.00 (m, 1H), 3.96 (t, J =
6.6 Hz, 2H), 3.84 (d, J = 4.4 Hz, 1H), 3.49–3.47 (m, 2H), 3.26–3.14 (m,
7H), 3.03 (s, 1H), 2.97–2.91 (m, 2H), 2.85–2.65 (m, 6H), 2.49–2.14 (m,
11H), 2.03–1.92 (m, 3H), 1.85–1.77 (m, 3H), 1.28–1.24 (m, 13H), 1.17
(d, J = 7.3 Hz, 3H), 1.05 (t, J = 7.4 Hz, 3H), 1.01 (d, J = 7.0 Hz, 3H),
0.91 (d, J = 5.9 Hz, 3H), 0.77–0.76 (m, 3H); MS (ESI) m/z calcd for
5.5. Cytotoxicity assay
A 28 h continuous MTT method was used to evaluate the toxicity of
compounds 20c, 20d and 20f to MCF-7 breast cancer cells. After the
MCF-7 breast cancer cells (approximately 104 cells/well) were cultured
in 96-well plates for 24 h, the drug solution of 20c, 20d and 20f was
added. After the above mixtures were cultured for 24 h, the solution of
MTT was added, and then continued to be cultured for 4 h at 37 ◦C. The
liquid in the mixture was carefully absorbed and discarded, and then
DMSO was added. The absorbance of each well was obtained at 570 nm
on an enzyme-labeled instrument.
C
48H83N5O12 [M + 2H]2+/2: 460.8, found: 461.1.
5.1.3.17. 12(R)-[1-(4-p-Propoxyphenyl-1H-1,2,3-triazol-1-yl)-1-methy-
lene]-9(S)-hydroxyclarithromycin (34l). White solid, yield 57%, mp:
84–86 ◦C, TLC Rf = 0.40 (DCM/CH3OH/NH3⋅H2O = 10:1:0.1); 1H NMR
(600 MHz, CDCl3, δ ppm): 7.73–7.68 (m, 3H), 6.94 (d, J = 8.8 Hz, 2H),
4.85 (d, J = 3.8 Hz, 1H), 4.68 (s, 1H), 4.52 (d, J = 7.0 Hz, 1H), 4.39 (s,
2H), 4.06–4.05 (m, 1H), 4.01–3.94 (m, 4H), 3.74 (s, 1H), 3.62–3.55 (m,
2H), 3.30–3.28 (m, 4H), 3.24 (s, 3H), 3.12–3.01 (m, 4H), 2.94–2.83 (m,
6H), 2.60–2.13 (m, 10H), 1.92 (s, 1H), 1.85–1.81 (m, 2H), 1.58 (dd, J =
15.0, 4.7 Hz, 3H), 1.30–1.26 (m, 13H), 1.19 (d, J = 8.1 Hz, 3H), 1.11 (d,
J = 6.3 Hz, 3H), 1.05 (t, J = 7.4 Hz, 3H), 0.83–0.81 (m, 6H); MS (ESI) m/
z calcd for C48H83N5O12 [M + 2H]2+/2: 460.8, found: 461.2.
Declaration of Competing Interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Acknowledgements
This work was supported financially by National Natural Science
Foundation of China (81973179 and 81673284) and Key Project of
Major Project of New and Old Kinetic Energy Conversion in 2020.
5.2. Bactericidal or bacteriostatic assay
The MIC values of Series A, B and C against sensitive bacterial strains
(S. aureus ATCC25923, E. coli ATCC25922, P. aeruginosa ATCC27853
and B. subtilis ATCC9372) and resistant bacterial strains (S. aureus
ATCC31007, S. aureus ATCC43300, S. pneumonia B1, S. pneumoniae
AB11 and S. pyogenes 1) were determined using CAM and AZM as
reference compounds. The MIC values of compounds were determined
by the broth microdilution method. The prepared compounds and the
two reference compounds were added to in cation adjusted Mueller-
Hinton (CAMH) in a 96-well plate and well mixed. Then, the concen-
tration of each compound in a 96-well plate was range from 0.25 to 128
µg/mL or from 0.5 to 256 µg/mL by using 2-fold dilution method.
Bacteria are inoculated into the above prepared solution of each well
Appendix A. Supplementary material
Supplementary data to this article can be found online at https://doi.
org/10.1016/j.bioorg.2021.104992.
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