ChCl: Gly (DESs) promote environmentally benign synthesis of xanthene derivatives and their antitubercular activity

Article abstract of DOI:10.3390/molecules26123667

A ChCl: Gly (DESs) promoted environmentally benign method was developed for the first time using the reaction of aryl aldehydes and dimedone to give excellent yields of xanthene analogues. The major application of this present protocol is the use of green

Full text of DOI:10.3390/molecules26123667

molecules
Article
ChCl: Gly (DESs) Promote Environmentally Benign Synthesis
of Xanthene Derivatives and Their Antitubercular Activity
Mashooq A. Bhat 1,
*
, Ahmed M. Naglah 2,3 , Siddique Akber Ansari 1 , Hanaa M. Al-Tuwajiria 1
and Abdullah Al-Dhfyan 4,5
1
2
3
4
5
Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University,
Riyadh 11451, Saudi Arabia; ansariakber@gmail.com (S.A.A.); mashooqbhat@rediffmail.com (H.M.A.-T.)
Department of Pharmaceutical Chemistry, Drug Exploration and Development Chair (DEDC),
College of Pharmacy, King Saud University, Riyadh 11451, Saudi Arabia; anaglah@ksu.edu.sa
Peptide Chemistry Department, Chemical Industries Research Division, National Research Centre,
Dokki, Cairo 12622, Egypt
Department of Pharmacology and Toxicology, College of Pharmacy, King Saud University,
Riyadh 11451, Saudi Arabia; aaldhfyan@kfshrc.edu.sa
Stem Cell & Tissue Re-Engineering Program, Research Center, King Faisal Specialized Hospital,
MBC-03, P.O. Box 3354, Riyadh 11211, Saudi Arabia
*
Correspondence: mabhat@ksu.edu.sa; Tel.: +966-558164097
Abstract: A ChCl: Gly (DESs) promoted environmentally benign method was developed for the
first time using the reaction of aryl aldehydes and dimedone to give excellent yields of xanthene
analogues. The major application of this present protocol is the use of green solvent, a wide range
of substrate, short reaction times, ease of recovery, the recyclability of the catalyst, high reaction
yield, and ChCl: Gly as an alternative catalyst and solvent. In addition to this, all the synthesized
compounds were evaluated for their in vitro antimycobacterial activity against M. tuberculosis H37Ra
ꢀꢁꢂꢀꢃꢄꢅꢆꢇ
ꢀꢁꢂꢃꢄꢅꢆ
Citation: Bhat, M.A.; Naglah, A.M.;
Akber Ansari, S.; Al-Tuwajiria, H.M.;
Al-Dhfyan, A. ChCl: Gly (DESs)
Promote Environmentally Benign
Synthesis of Xanthene Derivatives
and Their Antitubercular Activity.
Molecules 2021, 26, 3667.
(MTB) and M. bovis BCG strains. The compounds 3d
activity against MTB and M. bovis strains with minimum inhibitory concentration (MIC) values of
2.5 15.10 g/mL and 0.26–14.92 g/mL, respectively. The compounds 3e 3f, and 3j were found to
, 3e, 3f, and 3j showed significant antitubercular
−
µ
µ
,
https://doi.org/10.3390/
be nontoxic against MCF-7, A549, HCT 116, and THP-1 cell lines. All the prepared compounds were
confirmed by 1H NMR and 13C NMR analysis.
molecules26123667
Academic Editors: Richard A. Bunce,
Jean-Marc Sabatier and
Keywords: ChCl: Gly; environmentally benign; xanthenes; antitubercular; antibacterial activity;
cytotoxicity
Margherita Brindisi
Received: 30 December 2020
Accepted: 21 April 2021
Published: 16 June 2021
1. Introduction
In the last few years, deep eutectic solvents (DESs) have emerged as ecofriendly
and environmentally benign reaction protocols because of the exclusive physicochemical
properties, including negligible vapor pressure, nontoxicity, relatively wide liquid range,
Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
published maps and institutional affil-
iations.
biodegradability, and bio renewability [1,2]. DESs are easily prepared by the reaction of
hydrogen bond donor compounds, such as urea, succinic acid, malonic acid, and zinc
chloride with choline chloride as a quaternary ammonium salts. DESs also behave in a
different way than their individual components. The melting point of DESs is less than
their parent components due to charge delocalization occurring through hydrogen bonding
between the hydrogen donor component and halide ion [
promoting medium for the electrophilic activation of aldehydes in the reaction [
chloride-based DESs have been efficiently used for condensation reactions [ ], benzylation
of phenol [ ], synthesis of spirooxoindole [ ], Pictet–Spengler reaction [ ], Clauson–Kaas
reaction [ ], Perkin reaction [10], N-alkylation of amines [11], synthesis of pyrroles and
furans [ ], isolation of residual KOH content from biodiesel [12], deacidification of palm
oil [13], synthesis of highly substituted cyclohexa-1,3-dienamines [14], extraction of pheno-
3
]. The glycerol acts as an efficient
Copyright:
© 2021 by the authors.
4]. Choline
Licensee MDPI, Basel, Switzerland.
This article is an open access article
distributed under the terms and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/).
5
6
7
8
9
8
Molecules 2021, 26, 3667. https://doi.org/10.3390/molecules26123667
https://www.mdpi.com/journal/molecules

Molecules 2021, 26, 3667
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lic compounds from oil mixtures [15], extraction of vanillin [16], and pretreatment of rice
straw for butanol fermentation [17].
Tuberculosis is the ninth leading cause of death worldwide from a single infectious
agent, ranking above human immune deficiency virus (HIV)/acquired immune deficiency
syndrome (AIDS). It is a fatal airborne disease caused by Mycobacterium tuberculosis (Mtb),
which affects the lung and is also responsible for infection in other sites of the body [18].
The global tuberculosis report of 2019 given by the World Health Organization (WHO)
shows that TB is the leading cause of deaths due to antimicrobial resistance and among
people with HIV [19]. More than half a million new TB cases were notified to WHO by
national authorities [20]. Hence, the development of new therapeutic agents against TB is
the urgent need of the time.
According to a literature survey, xanthene derivatives are endowed with various
biological activities, including potent antimicrobial [21], analgesic [22], antimalarial [23],
anti-inflammatory [24], antiviral [25], antioxidant [26], antiproliferative activity [27], an-
ticancer [28], and urease activity [29]; inhibition of trypanothione reductase [23]; bone
morphogenetic protein (BMP-2)-targeted osteogenic agents [30]; selective positive allosteric
modulators of the δ-opioid receptors [31]; and estrogen receptors [32]. The structures of
biologically active xanthene analogues are presented in Figure 1.
Figure 1. Xanthene-incorporated bioactive molecules.
The expansion of a synthetic route for the synthesis of these biologically active com-
pounds using a facile, environmentally benign and nontoxic catalyst is of enormous sig-
nificance from the academic as well as industrial points of view. In the last few years, a
large number of protocols have been developed by researchers, including sulfuric acid or
hydrochloric acid [33], silica sulfuric acid [34], sulfamic acid [35], p-dodecylbenzenesulfonic
acid [36], boric acid [37], p-toluene sulfonic acid (p-TSA) [38], NaHSO4-SiO2 [39], TiO2-
2−
SO4
[40], molecular iodine [41], amberlyst-15 [42], and cyanuric chloride [43]. However,
a large number of the above routes have some drawbacks, such as the use of hazardous
reagents, long reaction times, strong acidic conditions and lower yields of final products.
Therefore, there is an urgent need for innovative and environmentally benign protocols to
synthesize biologically active compounds with ease. Keeping in mind the pharmacological
significance of xanthene derivatives and application of DESs for organic transformation,
and in continuation of our work on the synthesis and bio evaluation of numerous com-
pounds [44], herein, we report for the first time choline chloride: glycerol (ChCl: Gly)-based

Molecules 2021, 26, 3667
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synthesis of xanthene derivatives in excellent yields. The synthesized products were evalu-
ated for their antitubercular, cytotoxicity and antibacterial activities.
2. Materials and Methods
2.1. Preparation of ChCl: Gly
A mixture of ChCl (7.00 g, 50 mmol) and glycerol (11.65 g, 100 mmol) was added to
a round-bottomed flask. The reaction mixture was stirred at 70 ◦C until the formation of
colorless homogeneous liquid. This reaction follows the green chemistry rules [45].
2.2. General Procedure for Synthesis of Xanthene Derivatives
A mixture of benzaldehyde 1a (1 mmol) and dimedone
2 (1 mmol) in ChCl: Gly
(2 mL) was stirred at a temperature of 80 ◦C. The progress of the reaction was monitored
by thin-layer chromatography [n-hexane/ethyl acetate (3:7)] as a mobile phase. After
completion of the reaction, the reaction mass was allowed to cool for 30 min and poured on
ice-cold water. Thus, the solid compound obtained was filtered, dried out, and crystallized
in ethanol to give the pure product. The synthesized compound was confirmed by m.p.,
1H NMR, and 13C NMR.
2.2.1. 3,3,6,6-Tetramethyl-9-phenyl-3,4,5,6,7,9-hexahydro-1H-xanthene-1,8(2H)-dione (3a
)
Compoun◦d 3a was obtained from condensation reaction 1a and
2 as a white solid;
11.93 (s, 1H), 7.29–7.26 (t,
1
m.p.: 205–206 C; Yield: 93%; H NMR (500 MHz, CDCl3)
δ
J = 7.6 Hz, 2H), 7.19–7.09 (m, 3H), 5.56 (d, J = 60.1 Hz, 1H), 2.49–2.30 (m, 9H), 1.24–1.11 (dd,
J = 64.4, 29.0 Hz, 12H); 13C NMR (101 MHz, CDCl3)
δ
195.65 (C=O), 164.07 (C=C-O), 143.16
(Ar-H), 135.05 (Ar-H), 129.49 (Ar-H), 127.51 (Ar-H), 119.84 (Ar-H), 50.45 (CH), 42.23 (CH2),
32.45 (CH2), 31.34 (CH2), 29.56 (CH3) and 29.33 (CH3); High-resolution mass spectroscopy
(HRMS) (ESI-qTOF): Calcd for C23H27O3 [M+H]+, 351.0920: found: 351.0903.
2.2.2. 3,3,6,6-Tetramethyl-9-(m-tolyl)-3,4,5,6,7,9-hexahydro-1H-xanthene-1,8(2H)-dione (3b)
Compoun◦d 3b was obtained from condensation reaction 1a and
2 as a white solid;
7.10–6.99 (m, 3H), 6.86 (d,
1
m.p.: 208–210 C; Yield: 86%; H NMR (400 MHz, CDCl3)
δ
J = 6.8 Hz, 1H), 4.68 (s, 1H), 2.44 (s, 4H), 2.24 (s, 3H), 2.20–2.10 (t, J = 13.8 Hz, 4H), 1.05
(s, 6H), 0.95 (s, 6H); 13C NMR (101 MHz, CDCl3)
δ
196.31 (C=O), 162.23 (C=C-O), 144.01
(Ar-H), 137.21 (Ar-H), 129.38 (Ar-H), 127.84 (Ar-H), 127.13 (Ar-H), 125.21 (Ar-H), 115.67
(Ar-H), 50.76 (CH), 40.84 (CH2), 32.17 (CH2), 31.68 (CH2), 29.24 (CH2), 27.30 (CH3) and
21.48 (CH3); High-resolution mass spectroscopy (HRMS) (ESI-qTOF): Calcd for C24H29O3
[M + H]+, 365.0650: found: 365.0673.
2.2.3. 3,3,6,6-Tetramethyl-9-(p-tolyl)-3,4,5,6,7,9-hexahydro-1H-xanthene-1,8(2H)-dione (3c
Compound 3c was obtained from condensation reaction 1c and as a yellow solid;
m.p.: 222–224 ◦C; Yield: 90%; 1H NMR (400 MHz, CDCl3)
7.15–7.14 (d, J = 7.8 Hz, 2H),
6.99–6.97 (d, J = 7.5 Hz, 2H), 4.68 (s, 1H), 2.43 (s, 4H), 2.20 (s, 3H), 2.17–2.09 (m, 4H),
1.06 (s, 6H), 0.95 (s, 6H); 13C NMR (101 MHz, CDCl3)
196.36 (C=O), 162.17 (C=C-O),
)
2
δ
δ
141.21(Ar-C), 135.61(Ar-C), 128.71 (Ar-C), 128.20 (Ar-C), 115.67 (Ar-C), 50.75 (CH), 40.81
(CH2), 32.13 (CH2), 31.40 (CH2), 29.23 (CH2), 27.31 (CH3) and 21.02 (CH3); High-resolution
mass spectroscopy (HRMS) (ESI-qTOF): Calcd for C24H29O3 [M + H]+, 365.5041: found:
365.5012.
2.2.4. 9-(3-Methoxyphenyl)-3,3,6,6-tetramethyl-3,4,5,6,7,9-hexahydro-1H-xanthene-
1,8(2H)-dione (3d)
Compound 3d was obtained from condensation reaction 1d and
2 as a yellow solid;
m.p.: 162–164 ◦C; Yield: 84%; 1H NMR (400 MHz, CDCl3)
δ
7.05–7.01 (t, J = 8.1 Hz, 1H),
6.80–6.79 (d, J = 6.9 Hz, 2H), 6.57–6.55 (d, J = 7.0 Hz, 1H), 4.66 (s, 1H), 3.66 (s, 3H), 2.40
(s, 4H), 2.17–2.06 (q, J = 16.2 Hz, 4H), 1.01 (s, 6H), 0.91 (s, 6H); 13C NMR (101 MHz,
CDCl3)
δ 196.26 (C=O), 162.35 (C=C-O), 159.26 (Ar-C), 145.68 (Ar-C), 128.77 (Ar-C), 120.72

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(Ar-C), 115.42 (Ar-C), 114.26 (Ar-C), 111.78 (Ar-C), 55.01(OCH3), 50.70 (CH), 40.75 (CH2),
32.09 (CH2), 31.73 (CH2), 29.18 (CH3) and 27.29 (CH3); High-resolution mass spectroscopy
(HRMS) (ESI-qTOF): Calcd for C24H29O4 [M+H]+, 381.1631: found: 381.1645.
2.2.5. 9-(4-Methoxyphenyl)-3,3,6,6-tetramethyl-3,4,5,6,7,9-hexahydro-1H-xanthene-
1,8(2H)-dione (3e)
Compound 3e was obtained from condensation reaction 1e and
m.p.: 248–250 ◦C; Yield: 90%; 1H NMR (400 MHz, CDCl3)
7.17–7.15 (m, 2H), 6.71–6.69
(dd, J = 5.8, 2.6 Hz, 2H), 4.65 (s, 1H), 3.68 (d, J = 4.9 Hz, 3H), 2.42 (s, 4H), 2.20–2.09 (m,
4H), 1.05 (s, 6H), 0.94 (s, 6H); 13C NMR (101 MHz, CDCl3)
196.40 (C=O), 162.05 (C=C-O),
2 as a yellow solid;
δ
δ
157.90 (Ar-C), 136.47 (Ar-C), 129.25 (Ar-C), 115.72 (Ar-C), 113.41 (Ar-C), 55.05 (CH3), 50.74
(CH), 40.81 (CH2), 32.14 (CH2), 30.92 (CH2), 29.23 (CH3) and 27.29 (CH3); High-resolution
mass spectroscopy (HRMS) (ESI-qTOF): Calcd for C24H29O4 [M+H]+, 381.1150: found:
381.1132.
2.2.6. 9-(3,4-Dimethoxyphenyl)-3,3,6,6-tetramethyl-3,4,5,6,7,9-hexahydro-1H-xanthene-
1,8(2H)-dione (3f)
Compoun◦d 3f was obtained from condensation reaction 1f and
2 as a yellow solid;
6.88 (s, 1H), 6.75–6.68 (td,
1
m.p.: 200–202 C; Yield: 89%; H NMR (400 MHz, CDCl3)
δ
J = 8.2, 4.3 Hz, 2H), 4.68 (s, 1H), 3.83 (d, J = 1.8 Hz, 3H), 3.77 (d, J = 1.7 Hz, 3H), 2.44 (s,
4H), 2.24–2.14 (m, 4H), 1.09 (s, 6H), 0.98 (s, 6H); 13C NMR (101 MHz, CDCl3)
δ
196.42
(C=O), 162.08 (C=C-O), 148.44 (Ar-C), 147.45 (Ar-C), 136.98 (Ar-C), 120.09 (Ar-C), 115.73
(Ar-C), 112.31 (Ar-C), 110.85 (Ar-C), 55.85 (CH3), 50.74 (CH), 40.87 (CH2), 32.15 (CH2),
31.20 (CH2), 29.30 (CH3) and 27.14 (CH3); High-resolution mass spectroscopy (HRMS)
(ESI-qTOF): Calcd for C25H31O5 [M+H]+, 411.2860: found: 411.2845.
2.2.7. 3,3,6,6-Tetramethyl-9-(3-nitrophenyl)-3,4,5,6,7,9-hexahydro-1H-xanthene-1,8(2H)-
dione (3g)
Compound 3g was obtained from condensation reaction 1g and
m.p.: 172–174 ◦C; Yield: 82%; 1H NMR (400 MHz, CDCl3)
8.01–7.95 (m, 2H), 7.79–7.77
(d, J = 6.7 Hz, 1H), 7.40–7.38 (d, J = 7.8 Hz, 1H), 4.81 (s, 1H), 2.49 (s, 4H), 2.25–2.12 (q,
J = 16.3 Hz, 4H), 1.09 (s, 6H), 0.97 (s, 6H), 13C NMR (101 MHz, CDCl3)
196.30 (C=O),
2 as a red solid;
δ
δ
162.98 (C=C-O), 148.32 (Ar-C), 146.26 (Ar-C), 135.72 (Ar-C), 128.76 (Ar-C), 112.48 (Ar-
C), 121.64 (Ar-C), 114.53 (Ar-C), 50.62 (CH), 40.81 (CH2), 32.22 (CH2), 32.07 (CH2), 29.17
(CH3)and 27.29 (CH3); High-resolution mass spectroscopy (HRMS) (ESI-qTOF): Calcd for
C23H26NO5 [M+H]+, 396.3451: found: 396.3469.
2.2.8. 9-(3-Iodophenyl)-3,3,6,6-tetramethyl-3,4,5,6,7,9-hexahydro-1H-xanthene-1,8(2H)-
dione (3h)
Compound 3h was obtained from condensation reaction 1h and
solid; m.p.: 276–278 ◦C; Yield: 84%; 1H NMR (400 MHz, CDCl3)
7.53 (s, 1H), 7.42–7.41
(d, J = 7.8 Hz, 1H),7.32–7.24 (d, J = 7.6 Hz, 1H), 6.96–6.92 (t, J = 7.8 Hz, 1H), 4.60 (s, 1H),
2.45 (s, 4H), 2.24–2.14 (m, 4H), 1.08 (s, 6H), 0.99 (s, 6H); 13C NMR (101 MHz, CDCl3)
2 as a pale yellow
δ
δ
196.21 (C=O), 162.48 (C=C-O), 146.36 (Ar-C), 137.07 (Ar-C), 135.48 (Ar-C), 129.76 (Ar-C),
128.20 (Ar-C), 115.05 (Ar-C), 94.07 (Ar-C), 50.67 (CH), 40.84 (CH2), 32.21 (CH2), 31.61 (CH2),
29.67 (CH2), 29.20 (CH3) and 27.32 (CH3); High-resolution mass spectroscopy (HRMS)
(ESI-qTOF): Calcd for C23H26IO3 [M+H]+, 477.3570: found: 477.3526.
2.2.9. 9-(4-Bromophenyl)-3,3,6,6-tetramethyl-3,4,5,6,7,9-hexahydro-1H-xanthene-1,8(2H)-
dione (3i)
Compound 3i was obtained from condensation reaction 1i and
2
as a red solid; m.p.:
7.34–7.32 (d, J = 6.9 Hz, 2H), 7.18–7.16
(d, J = 6.8 Hz, 2H), 4.69 (s, 1H), 2.46 (s, 4H), 2.25–2.14 (q, J = 16.4 Hz, 4H), 1.10 (s, 6H), 0.98
(s, 6H); 13C NMR (101 MHz, CDCl3)
196.26 (C=O), 162.41 (C=C-O), 143.18 (Ar-C), 131.09
◦
238–240 C; Yield: 86%; 1H NMR (400 MHz, CDCl3)
δ
δ

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(Ar-C), 130.13 (Ar-C), 120.18 (Ar-C), 115.14 (Ar-C), 50.65 (CH), 40.81 (CH2), 32.15 (CH2),
31.52 (CH2), 29.21 (CH3) and 27.26 (CH3); High-resolution mass spectroscopy (HRMS)
(ESI-qTOF):Calcd for C23H26BrO3 [M+H]+, 430.2450: found: 430.2400.
2.2.10. 9-(4-Chlorophenyl)-3,3,6,6-tetramethyl-3,4,5,6,7,9-hexahydro-1H-xanthene-
1,8(2H)-dione (3j)
Compound 3j was obtained from condensation reaction 1j and
2
as a yellow solid;
7.24 (d, J = 8.4 Hz, 2H),
7.21–7.14 (m, 2H), 4.71 (s, 1H), 2.47 (s, 4H), 2.19 (q, J = 16.3 Hz, 4H), 1.10 (s, 6H), 0.98 (s,
6H); 13C NMR (101 MHz, CDCl3)
196.28 (C=O), 162.44 (C=C-O), 142.70 (Ar-C), 131.93
◦
1
m.p.: 238–240 C; Yield: 90%; H NMR (400 MHz, CDCl3)
δ
δ
(Ar-C), 129.74 (Ar-C), 128.13 (Ar-C), 115.18 (Ar-C), 50.66 (CH), 40.79 (CH2), 32.14 (CH2),
31.43 (CH2), 29.21 (CH3) and 27.24 (CH3); High-resolution mass spectroscopy (HRMS)
(ESI-qTOF): Calcd for C23H26ClO3 [M+H]+, 385.1165: found: 385.1129.
2.2.11. 9-(4-Hydroxyphenyl)-3,3,6,6-tetramethyl-3,4,5,6,7,9-hexahydro-1H-xanthene-
1,8(2H)-dione (3l)
Compound 3l was obtained from condensation reaction 1l and
248–250 ◦C; Yield: 83%; 1H NMR (400 MHz, CDCl3)
7.31 (s, 1H), 7.07–7.05 (d, J = 7.8 Hz,
2H), 6.55–6.53 (d, J = 7.8 Hz, 2H), 4.66 (s, 1H), 2.46 (s, 4H), 2.26–2.16 (q, J = 16.4 Hz, 4H), 1.08
(s, 6H), 0.99 (s, 6H). 13C NMR (101 MHz, CDCl3)
197.30 (C=O), 162.45 (C=C-O), 154.70
2 as a red solid; m.p.:
δ
δ
(Ar-C), 135.47 (Ar-C), 129.29 (Ar-C), 115.85 (Ar-C), 115.23 (Ar-C), 50.72 (CH2), 40.81 (CH2),
32.23 (CH2), 30.93 (CH2), 29.11 (CH3) and 27.35 (CH3); High-resolution mass spectroscopy
(HRMS) (ESI-qTOF): Calcd for C23H27O4 [M+H]+, 367.0652: found: 367.0673.
2.2.12. 9-Cyclohexyl-3,3,6,6-tetramethyl-3,4,5,6,7,9-hexahydro-1H-xanthene-1,8(2H)-
dione (3m)
Compound 3m was obtained from condensation reaction 1m and
m.p.: 176–178 ◦C; Yield: 82%; 1H NMR (400 MHz, CDCl3)
7.67 (s, 2H), 5.46 (s, 1H), 3.31
(s, 2H), 2.50 (s, 4H), 2.23 (s, 4H), 1.22 (s, 1H), 1.06–0.85 (d, J = 18.0 Hz, 12H); 13C NMR
(101 MHz, CDCl3) 193.30 (C=O), 165.54(C=C-O), 111.77 (Ar-C), 50.43 (CH), 40.71 (CH2),
2 as a white solid;
δ
δ
34.51 (CH2), 33.01 (CH2), 31.25 (CH2), 29.20 (CH2), 27.42 (CH2), 25.93 (CH2), 24.52 (Cy-C),
23.21 (Cy-C), 23.15 (CH3) and 23.07 (CH3); High-resolution mass spectroscopy (HRMS)
(ESI-qTOF): Calcd for C23H33O3 [M+H]+, 357.1081: found: 357.1064.
3. Results and Discussion
3.1. Chemistry
We studied the model reaction using benzaldehyde (1a) (1 mmol), and dimedone
(2) (1 mmol) for the optimization of the reaction solvents, catalysts, temperatures, and
observation of this study is described in Scheme 1 and Table 1.
Scheme 1. Model reaction.
Initially, we observed that the titled product was not perceived when the model
reaction was carried out in the absence of a catalyst (Table 1, Entry 1). In the next step, we
tested the reaction using different choline chloride-derived DESs as a reaction medium
◦
at 70 C, which resulted in the desired product 3a in lower yields, and the results are
given in (Table 1, Entry 2–10). In the presence of ChCl: Gly, this results in the formation of
compound 3a in a good yield of 93% at 80 ◦C for 45 min. It was observed that ChCl: Gly
(1:2) was a pre-eminent catalyst for this conversion. According to these results, the catalyst

Molecules 2021, 26, 3667
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plays an important role in this transformation. We studied several DES catalysts. Among
them, ChCl: Gly was highly active due to its relatively low viscosity and high thermal
stability [46]. The remaining DESs showed lower activity because of direct interactions of
the reactants with DESs, variation in acidity, and the physical properties of DESs. Due to
these main reasons, the smooth interaction between reactants was not possible. The ChCl:
Gly DES exhibited self-association of glycerol and choline chloride through the interaction
of hydrogen bonding, which is the main reason for its high catalytic efficiency.
Table 1. Optimization of the reaction conditions a.
◦
Yield (%) b
Entry
Solvent
Temperature ( C)
Time (min)
1
2
3
-
70
70
70
520
80
80
-
60
58
ChCl:Urea(1:2)
ChCl:Thiourea(1:2)
ChCl:Adipic
Acid (1:1)
ChCl:Oxalic
Acid(1:1)
ChCl:Malonic
Acid(1:1)
ChCl:Succinic
Acid(1:1)
5
4
7
6
70
70
70
70
80
80
80
80
28
63
38
46
9
8
ChCl:ZnCl2 (1:2)
ChCl:FeCl3 (1:2)
ChCl:Gly (1:1)
ChCl:Gly (1:2)
ChCl:Gly (1:3)
ChCl:Gly (1:2)
ChCl:Gly (1:2)
Gly
70
70
70
70
70
80
90
80
80
85
85
80
60
60
45
45
80
520
30
35
76
82
83
93
93
62
25
10
11
12
13
14
15
16
ChCl
a
Reaction conditions: benzaldehyde
Choline Chloride; Gly: Glycerol.
1 (1 mmol), dimedone 2
(1 mmol and DES (2.0 g); b Isolated yields. ChCl:
In the next step, we also screened the effect of temperature on the model reaction.
The model reaction was stirred at a different temperature ranges from 70 to 90 ◦C, with an
increment of 10 ◦C each time. The results exhibited that an increase in the temperature of
the model reaction increased the product of the reaction. All these observations suggest
that 80 ◦C is an optimum temperature in terms of reaction time and yield (Table 1, Entries
11, 13–14).
When the reaction was performed with the help of the glycerol, results in a 62% yield
of the titled product as shown in (Table 1, Entry 15). Whereas when the model reaction was
◦
performed using the ChCl, results decrease in the yield of titles compound 3a at 80 C was
shown in (Table 1, Entry 16). This result proves that the excellent yield of the final product
is due to the ChCl: Gly (1:2) and not to its individual components, glycerol and choline
chloride. This is because of extensive hydrogen bonding between glycerol and quaternary
ammonium salt choline chloride (Scheme 2) [46].
Scheme 2. Hydrogen bonding between glycerol and choline chloride.
Furthermore, we also investigated the catalyst loading on the model reactions. The
model reaction was carried out at different catalyst concentrations of 0.5, 1.0, 1.5, 2.0, and
◦
2.5 g of ChCl: Gly (1:2) at 80 C. Better results were observed when the model reaction was

Molecules 2021, 26, 3667
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carried out using 2.0 g of ChCl: Gly (1:2) to give 93% yields of the titled product (Table 2,
Entry 4).
Table 2. Optimization of deep eutectic solvent a.
◦
% Yield b
Entry
ChCl: Gly (g)
Time (min)
Temp ( C)
1
2
3
4
5
0.5
1
1.5
2
45
45
45
45
45
80
80
80
80
80
28
58
78
93
94
2.5
a
Reaction conditions: benzaldehyde 1 (1 mmol), dimedone 2 (1 mmol) and ChCl: Gly (2.0 g); b Isolated yields.
The recycling and reuse of ChCl: Gly is the main advantage of our planned method.
We studied the reusability of the ChCl: Gly. The reaction was carried out using aldehyde
and dimedone under the optimized reaction parameters.
After the finishing point of the reaction, the reaction mass was poured on ice-cold
water and product was extracted with EtOAc (2
×
20 mL) and an organic layer was washed
with 5% NaHCO3 then dried over anhydrous NaSO4. In this reaction, ChCl: Gly (1:2)
was recovered by simply evaporating water from the reaction mass. The recyclability of
the catalytic efficiency of ChCl: Gly was tested for five consecutive cycles. There was no
significant decrease in the yield observed as shown in Figure 1 (Table 3).
Table 3. Recycling of ChCl: Gly (DES) for the synthesis of compound 3a a.
% Yield b
Entry
Run
Time (min)
1
2
3
4
5
6
Fresh
45
45
45
45
45
45
93
93
91
89
85
82
1
2
3
4
5
a
Reaction conditions: benzaldehyde 1 (1 mmol), dimedone 2 (1 mmol) and ChCl: Gly (2.0 g); b Isolated yields.
ChCl: Gly (1:2) exhibited prominent catalytic activity with extra thermal stability. Zhao
◦
S. H. et al. and Gary A. Baker described the thermal strength of ChCl: Gly at 150 C [47]. We
◦
−1
also confirmed this thermo-gravimetric analysis in a nitrogen atmosphere at 250 C min
(Figure 2). This study sugge◦sts that the ChCl: Gly is stable up to 150 ◦C. In the next step,
ChCl: Gly is heated at 100 C for 300 min, resulting in no weight loss being observed.
According to these results, our synthesized ChCl: Gly is stable below 100 ◦C.
Figure 2. Thermo-gravimetric analysis of ChCl: Gly (DES).

Molecules 2021, 26, 3667
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We studied the stability of ChCl: Gly during recyclability of 3a with the help of IR of
the recovered ChCl: Gly. The Fourier-transform infrared spectroscopy (FTIR) spectrum
of pure ChCl: Gly showed peaks at 1040, 1482, and 2950 cm−1 and a broad peak at
around 3324 cm−1. There was no considerable change in the Fourier-transform infrared
spectroscopy (FTIR) spectra of pure ChCl: Gly and the IR spectra of ChCl: Gly recovered
after the fifth cycle (Figure 3).
Figure 3. FTIR spectra of (
of reaction (c) ChCl: Gly measured after 300 min at 100 C.
a
) ChCl: Gly measured before reaction (
b) ChCl: Gly measured after fifth cycle during recyclability
◦
We undertook a comparative study of the ChCl: Gly with other previously reported
catalysts for the synthesis of xanthene analogues. The comparison results proved that ChCl:
Gly is the better catalyst in terms of its excellent yield and reusability with less reaction
time (Table 4, Entry 10). In conclusion, ChCl: Gly is an efficient and greener approach for
the synthesis of xanthene analogues.
Table 4. Comparative catalytic study of ChCl: Gly with the reported catalysts.
Entry
Catalyst
Amount of Catalyst Time (min) Yield (%)
Solvent/Condition
Ref.
1
2
β-CD (1 mmol)
SBSSA
1 mmol
30 mg
10 h
10 h
96
98
EtOH/60 ◦C
EtOH/reflux
[bmim][BF4]/80 ◦C
CH3CN/Reflux
Solvent free/110 ◦C
[48]
[49]
FeCl3.6H2O (10
mol%)
3
4
5
10 mol%
200 mg
50 mg
6 h
5 h
5 h
92
92
94
[50]
[51]
[52]
Amberlyst-15
Cellulose sulfonic
acid
[bmim][HSO4]
(42 mol%)
6
42 mol%
3 h
85
Solvent free/100 ◦C
[40]
Fe3+ montmorillonite
Zr(DP)2
7
8
9
85 mg
10 mol %
30 mol %
6 h
24 h
5 h
94
98
86
EtOH/reflux
EtOH/80 ◦C
CH3CN/reflux
ChCl:Gly act as a
solvent
[53]
[54]
[55]
ZnO-CH3COCl
10
ChCl:Gly
2.0 g
45 min
93
Present work
1
1
Compound 3a was confirmed by H NMR and 13C NMR analysis. In H NMR
spectra of the compound, 3a shows two distinct singlets at
two methyl groups. Peaks were observed at 2.49 and 2.30 ppm for the -CH2-C=O and
CH2 protons, respectively. These two signals confirmed that a dimedone ring is present
in our synthesized compound. The peak observed at 5.56 ppm is due to the presence
of -CH proton in compound 3a. In the 13C NMR spectrum of compound 3a, the peak at
195.65 ppm shows the presence of a -C=O group. The methine, methylene, and methane
peaks observed at 50.43, 42.23, 32.45, 31.34, 29.56, and 29.33 ppm suggest the formation
of the final compound 3a.
δ 1.21 and 1.24 ppm for the
δ
δ
δ
δ

Molecules 2021, 26, 3667
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In conclusion, the effectiveness and better reaction time for the model reaction was
observed at 80 ◦C by using 2 g of ChCl: Gly as a catalyst. With excellent reaction condi-
tions in hand, the adaptableness of this approach was employing the synthesis xanthene
analogues (3a–l). Various substituents on aryl aldehyde, including methoxy, methyl, nitro,
halogen (-Cl,-Br, -I), and hydroxyl groups, were used. The synthesis of the compounds
(
3b–l) using the optimized reactions conditions and results are shown in Scheme 3. The
result clearly suggests that the condensation reactions using ChCl: Gly catalyst show an
excellent and remarkable performance irrespective of the electron-withdrawing/-donating
groups present on the aryl aldehydes (Table 2), hence this method is facile, efficient, and
general for the synthesis of xanthene analogues. All the synthesized final compounds
(
3a–l) were well characterized by 1H NMR and 13C NMR spectroscopic techniques and are
incorporated in the Supplementary Material.
a
Scheme 3. Synthesis of xanthene derivatives (3a–l) by using ChCl: Gly as a catalyst. Reaction
conditions: aldehydes (1a–m) (1 mmol) and dimedone (2) (1 mmol) in ChCl: Gly (2.0 g) stirred at
◦
45 C; b isolated yields, cmelting points are in good agreements with previously reported compounds
in the literature [50].

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The reaction mechanism cycle for the preparation of xanthene analogues employing
ChCl: Gly is the catalyst. In the first step, benzaldehyde activated by ChCl: Gly results in
the formation of intermediate
the enol product II. In the third step, intermediate
the product III. Further formation of the alkylation product
II and III via removal of the H2O molecule. In the next step, intramolecular cyclization
of occurs to give VI. In the last step, elimination of the H2O molecule using ChCl: Gly
I
. In the next step, ChCl: Gly reacts with dimedone to give
reacts with II to afford the addition of
occurs due to the reaction of
I
V
V
results in the formation of the titled xanthene analogues 3a and regeneration of the catalyst.
The detailed reaction mechanism is presented in Scheme 4.
Scheme 4. Reaction mechanism cycle for the preparation of compounds 3a.
3.2. Biological Activity
3.2.1. Antitubercular Activity Screening
The synthesized xanthene derivatives (3a–m) were screened for their in vitro anti-
tubercular activity against MTB H37Ra American Type Culture Collection 25177 (ATCC
25177) and M. bovis BCG (ATCC 35743) strain in liquid medium [56]. We developed the
well-known (2,3-Bis-(2-Methoxy-4-Nitro-5-Sulfophenyl)-2H-Tetrazolium-5-Carboxanilide)
(XTT) reduction menadione assay (XRMA) of an antitubercular screening protocol (incorpo-
rated in the Supplementary Material) using a standard reference, i.e., rifampicin drug, and
the minimum inhibitory concentration (MIC) and inhibitory concentration (IC50) values
are presented in Table 5.
Among all the synthesized xanthene derivatives compounds, 3d
were found to have prominent antimycobacterial activity against the MTB H37Ra and
M. bovis BCG strain with MIC values of 2.5–15.10 and 0.26–14.92 g/mL, respectively.
However, the rest of the xanthene derivatives 3a 3b 3c 3g 3h 3i 3k, and 3l were inactive
against the MTB H37Ra and M. bovis BCG strain with MIC ≥ 30 µg/mL.
, 3e, 3f, 3j, and 3k
µ
,
,
,
,
,
,

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Table 5. In vitro antimycobacterial activity of xanthene conjugates in (µg/mL).
MTB H37Ra
M. bovis BCG
Compound
a
a
a
a
MIC
IC50
MIC
IC50
3a
3b
3c
3d
3e
3f
3g
3h
3i
>30
>30
>30
15.10
4.20
2.5
>30
>30
>30
4.74
7.8
>30
>30
0.045
>30
>30
>30
3.20
0.74
0.79
>30
>30
>30
0.50
2.3
>30
>30
>30
>30
>30
>30
3.40
0.28
0.32
>30
>30
>30
0.6
14.92
0.26
1.22
>30
>30
>30
2.58
9.4
3j
3k
3l
5.8
>30
>30
>30
>30
0.0017
>30
>30
0.017
3m
Rifampicin
b
0.0015
a
MIC/IC50 in mg/mL. Antitubercular activity of all compounds was investigated using serial dose-dependent
b
dilutions protocol. Rifampicin as a standard drug acts as a positive control.
3.2.2. Structure–Activity Relationship (SAR)
According to the activity data, the xanthene analogues showed excellent antitubercular
activity and the results are presented in Table 5. The biological activity results revealed that
the activity was significantly impacted by various substituents present on the aryl ring.
Firstly, we will elaborate the antitubercular activity of xanthene analogues against the
MTB H37Ra strain. From the xanthene analogues series (3a–m), compound 3a, without any
substitution on the aryl ring, displayed lesser potency against the MTB H37Ra strain with
an MIC value >30
are displayed in Table 5. Compounds 3b (R2 = -methyl) and 3c (R3 = -methyl) showed
lower antitubercular activity against the MTB H37Ra strain with MIC 30 g/mL. When
the -methoxy group was introduced in compound 3d (R2 = -methoxy), it showed excellent
activity against the MTB H37Ra strain with MIC = 15.10 g/mL. When a -methoxy group
was present in the aryl ring at the para position as in compound 3e (R3 = -methoxy), it
showed excellent activity against the MTB H37Ra strain with an MIC value of 4.20 g/mL
µg/mL as compared to the standard drug rifampicin and the results
≥
µ
µ
µ
with reference to the standard drug. When a -methoxy group was introduced at both R2
and R3 in compound 3f, it exhibited prominent antimycobacterial activity against the MTB
H37Ra strain with MIC = 2.25 µg/mL.
When a nitro group was introduced in compound 3g (R2 = -NO2), it exhibited less
activity against the MTB H37Ra strain with MIC
drug. Replacing the nitro group with an iodo group in compound 3h (R2 = -I) resulted in
inactivity against the MTB H37Ra strain with MIC 30 g/mL. Installation of a bromo
group at the para position of the aryl group in 3i (R3 = -Br) resulted in inactivity against
the MTB H37Ra strain with MIC 30 g/mL. Replacing a bromo by a chloro group at the
para position in compound 3j (R3 = -Cl) resulted in the compound being highly potent
against the MTB H37Ra strain with MIC = 4.74 g/mL. When the position of the chloro
group was changed in 3k (R2 = -Cl), it showed moderate antitubercular activity against the
MTB H37Ra strain with MIC = 8.34 g/mL as compared to compound 3j. When a hydroxy
group was present at the para position in compound 3l (R3 = -OH), it was inactive against
the MTB H37Ra strain with MIC 30 g/mL. In compound 3m, when the aryl ring was
replaced by a cyclohexane ring, it was found to be inactive against the MTB H37Ra strain
with MIC 30 g/mL. Hence, among all the synthesized xanthene derivatives 3a–m
compounds 3d 3e 3f 3j, and 3k were found to be highly potent against the MTB H37Ra
strain and the details are disclosed in Table 5.
≥
30 µg/mL with reference to the standard
≥
µ
≥
µ
µ
µ
≥
µ
≥
µ
,
,
,
,
Further, all the compounds were also tested for antitubercular activity against the M.
bovis BCG strain. From the series (3a–m), compound 3a, without any substituent present

Molecules 2021, 26, 3667
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on the aryl ring, showed no activity against the M. bovis BCG strain with MIC
and the results are presented in Table 5. Compounds 3b i (R2 = -Me) and 3c (R3 = -Me) were
found to be inactive against the M. bovis BCG strain with MIC 30 g/mL. When methoxy
was introduced in compound 3d (R2 = -OMe), it presented prominent antitubercular activity
against the M. bovis BCG strain MIC = 14.92 g/mL. Introduction of a methoxy group at the
para position of the aryl ring in compound 3e (R3 = -OMe) and 3f R2 = R3 = -OMe) resulted
in potency against the M. bovis BCG strain with MIC = 0.26 and 1.22 g/mL, respectively.
≥
30 µg/mL
≥
µ
µ
(
µ
When a nitro group was present at the R2 position in compound 3g (R2 = -NO2), it was
found to be inactive against the M. bovis BCG strain with MIC ≥ 30 µg/mL.
Replacing the nitro group by an iodo group in 3h (R2 = -I), the compound was found
to be inactive towards the M. bovis BCG strain with MIC
bromo group at the para position of the aryl group in 3i (R3 = -Br) resulted in the compound
being inactive against the M. bovis BCG strain with MIC 30 g/mL. When the chloro
group was present at the para position in 3j (R3 = -Cl), the compound was highly active
against the M. bovis BCG strain with MIC = 2.58 g/mL. When the position of the chloro
group was changed in 3k (R2 = -Cl), it showed moderate antitubercular activity against
the M. bovis BCG strain with MIC = 6.13 g/mL as compared to compound 3j. Compound
≥
30 µg/mL. Introduction of a
≥
µ
µ
µ
3l (R3 = -OH) did not show antitubercular activity against the M. bovis BCG strain with
MIC ≥ 30 µg/mL. In compound 3m, the aryl group was replaced by a cyclohexane ring
and the antitubercular activity against the M. bovis BCG strain was decreased, with an
MIC value >30 µg/mL. Hence, among all the xanthene derivatives (3a–m), compounds
3d, 3e, 3f, 3k, and 3l were highly potent against the M. bovis BCG strain and the results are
presented in Table 5.
3.3. Cytotoxicity
Among the prepared xanthene analogues, the most active compounds 3d, 3e, 3f,
3j, and 3k were further evaluated for cytotoxicity against human cell lines, including
MCF-7, A549, HCT 116, and THP-1, using the well-developed tetrazolium salt (3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay [57]. The toxicity
results revealed that most of these xanthene analogues were nontoxic and showed specific
activity against the MTB and M. bovis BCG strains, with GI50/GI90 (>100
µg/mL) (Table 6).
Table 6. In vitro cytotoxicity of active xanthene analogues (GI50 in µg/mL).
Compound
MCF-7
A549
HCT 116
THP-1
3d
3e
3f
3j
3k
>100
>100
>100
>100
>100
>100
0.0048
>100
>100
>100
>100
>100
>100
0.0035
>100
>100
>100
>100
>100
>100
0.0260
>100
>100
>100
>100
>100
>100
0.1374
Rifampicin
Paclitaxel
GI50 indicates the concentration that inhibits 50% of the growth of cells.
3.4. Selectivity Index (SI)
SI determines the most active compounds selective towards mycobacteria but remain
nontoxic against host human cancer cell lines. The drug susceptibility study indicates that
when the SI was >10, then these compounds were more selective as antitubercular agents.
Therefore, the most active compounds 3e (SI = 20), 3f (SI = 44), 3j (SI = 18), and 3k (SI = 14)
exhibited >10 SI, revealing that these xanthene analogues act as excellent antitubercular
agents. The detailed study is summarized in Table 7.

Molecules 2021, 26, 3667
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Table 7. Selectivity index against dormant MTB H37Ra and M. bovis BCG.
MCF-7
A549
HCT 116
THP-1
Compound
MTB
BCG
MTB
BCG
MTB
BCG
MTB
BCG
3d
3e
3f
3j
3k
7
20
44
18
14
196
7
330
82
40
12
7
20
42
18
14
196
7
333
82
40
12
7
20
42
18
14
196
7
330
82
40
12
7
20
44
18
14
196
7
330
82
40
12
Rifampicin
222
222
222
222
3.5. Antibacterial Activity
The specificity of the most active xanthene analogues 3d
,
3e, 3f, 3j, and 3k was
estimated for their antibacterial activity against the Gram-negative bacteria Escherichia
coli and Pseudomonas fluorescence and the Gram-positive bacteria Staphylococcus aureus and
Bacillus subtilis. According to the antibacterial activity data, the most potent compounds
were inactive towards the tested bacterial strains. This activity results show that the most
potent compounds were highly specific against the MTB H37Ra and M. bovis BCG strains
and the detailed study is described in Table 8.
Table 8. Antibacterial activity of compounds MIC (µg/mL).
Compound
E. coli
P. fluorescens
S. aureus
B. subtilis
3d
3e
>30
>30
>30
>30
>30
>30
>30
>30
3f
3j
3k
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
>30
Kanamycin
Ampicillin
3.34 ± 0.41
1.01 ± 0.09
>61.92
2.86 ± 0.78
2.78 ± 0.85
4.17 ± 1.04
12.47 ± 1.28
29.53 ± 1.88
3.6. ADME Properties
The success of a drug is determined not only by good efficacy but also by an acceptable
ADME (absorption, distribution, metabolism, and excretion) profile. In this study, we cal-
culated the molecular volume (MV), molecular weight (MW), logarithm of the partition co-
efficient (miLogP), number of hydrogen bond acceptors (n-ON), number of hydrogen bond
donors (n-OHNH), topological polar surface area (TPSA), number of rotatable bonds (n-
ROTB), and Lipinski’s rule of five [58] using the Molinspiration online property calculation
toolkit [59]. Absorption (% ABS) was calculated by: % ABS = 109 − (0.345 × TPSA) [60].
The drug-likeness model score (a collective property of the physicochemical proper-
ties, pharmacokinetics, and pharmacodynamics of a compound represented by a numerical
value) was computed by MolSoft software [61]. A computational study of all the synthe-
sized compounds was performed for the prediction of ADME properties and the values
obtained are presented in Table 9. It was observed that the compounds exhibited a good %
ABS (% absorption), ranging from 69.11 to 85.02%.
Furthermore, the compounds 3j and 3k did not violate Lipinski’s rule of five (miLogP ≤ 5).
A molecule that is likely to be developed as an orally active drug candidate should show
no more than one violation of the following four criteria: miLogP (octanol-water partition
coefficient)
number of hydrogen bond donors
score, the higher the probability that the particular molecule will be active. All the tested
compounds followed the criteria for orally active drugs and therefore, these compounds
may have good potential for eventual development as oral agents.
≤
5, molecular weight
≤
500, number of hydrogen bond acceptors
5 [62]. The larger the value of the drug-likeness model
≤
10, and
≤

Molecules 2021, 26, 3667
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Table 9. Pharmacokinetic parameters important for good oral bioavailability and its drug likeness model score.
TPSA (A2)
-
Com
Rule
% ABS
-
n-ROTB
-
MV
-
MW
<500
miLogP
≤5
n-ON
<10
n-OHNH
<5
Lipinski Violation
Drug Likeness
Model Score
≤1
3a
3b
3c
3d
3e
3f
94.03
94.03
94.03
90.84
90.84
87.66
78.22
94.03
94.03
94.03
87.05
94.03
43.38
43.38
43.38
52.61
52.61
61.84
89.20
43.38
43.38
43.38
63.60
43.38
1
1
2
2
3
2
1
1
1
1
1
1
336.59 350.46
353.15 364.49
353.15 364.49
362.13 380.48
362.13 380.48
5.20
5.62
5.65
5.23
5.25
4.84
5.13
6.26
6.01
5.88
4.72
5.79
3
3
3
4
4
5
6
3
3
3
4
3
0
0
0
0
0
0
0
0
0
0
0
1
1
1
1
2
2
3
1
1
1
1
1
1
−1.31
−0.98
−1.27
−0.89
−1.09
0.41
−0.85
−0.87
−1.16
−0.77
−0.72
−1.32
387.8
410.51
3g
3h
3i
359.92 395.45
360.58 476.35
354.48 429.35
350.12 384.90
344.61 366.46
355.18 356.51
3j
3k
3l
Com: Compound, % ABS: percentage absorption, TPSA: topological polar surface area, n-ROTB: number of rotatable bonds, MV: molecular
volume, MW: molecular weight, milogP: logarithm of partition coefficient of the compound between n-octanol and water, n-ON acceptors:
number of hydrogen bond acceptors, n-OHNH donors: number of hydrogen bonds donors.
4. Conclusions
In conclusion, a facile and environmentally benign approach was developed for the
synthesis of xanthene analogues using a recyclable and inexpensive ChCl: Gly catalytic
solvent-free system. This reaction protocol revealed several advantages, including high
atom efficiency, mild reaction conditions, uniqueness, easy workup procedure, clean
reaction profiles, and eco friendliness. Additionally, all the prepared xanthene analogues
were screened for their in vitro antitubercular activity against the MTB H37Ra and M. bovis
BCG strains. Among all the screened compounds, 3d
highly active compounds, with MIC values in the range of 2.5
M. bovis BCG strain and 0.26–14.92 g/mL against the MTB H37Ra strain, respectively. The
,
3e,
3f
,
3j, and 3k were found to be
−
15.10 g/mL against the
µ
µ
most active compounds were found to be nontoxic against MCF-7, A549, HCT 116, and
THP-1 cancer cell lines by employing the MTT method. Highly potent xanthene derivatives
3d, 3e, 3f, 3j, and 3k were found to have a selectivity index of >10 against MCF-7, A549,
HCT 116, and THP-1, which suggests that they can act as promising antitubercular agents.
All these results confirm that compounds 3d, 3e, 3f, 3j, and 3k can be further modified as
lead drug molecules.
Supplementary Materials: The following are available online. Biological activity protocol and
supplementary data (copies of 1H and 13C NMR spectra of all the synthesized compounds).
Author Contributions: M.A.B. and H.M.A.-T. conceived and designed the experiments; M.A.B. and
S.A.A. performed chemical synthesis and purifications; A.M.N. and A.A.-D. performed biological
assays (tubercular activity against MTB and M. bovis BCG strains); M.A.B. wrote the paper; S.A.A.
helped in the revision of the manuscript. All authors have read and agreed to the published version
of the manuscript.
Funding: The authors would like to extend their sincere appreciation to the Deanship of Scientific Re-
search at King Saud University for funding this research through Research Group no: (RG—1435-006).
Institutional Review Board Statement: Not applicable.
Informed Consent Statement: Not applicable.
Data Availability Statement: Biological activity protocol and supplementary data (copies of 1H and
13C NMR spectra of all the synthesized compounds) associated with this article can be found through
the “Supplemental Content” section of this article’s webpage.
Acknowledgments: The authors would like to extend their sincere appreciation to the Deanship of
Scientific Research at King Saud University for funding this research through Research Group no:
(RG—1435-006).
Conflicts of Interest: The authors declare no conflict of interest.
Sample Availability: Samples of the compounds are available from authors.

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