Synthesis of Novel N,N-Disubstituted Ethyl P-[2-(1 H-Indol-3-YL)Acetyl]Phosphoramidates and Their Biological Activity

Article abstract of DOI:10.1080/10426507.2015.1054930

A series of novel N,N-disubstituted ethyl P-[2-(1H-indol-3-yl)acetyl]phosphoramid-ates were synthesized. All title compounds were characterized by IR, NMR (¹H, ¹³C, ³¹P) spectra, mass spectra, and C, H, N analysis. The anticancer activity of the title compounds was evaluated in two human cell lines MCF-7 (breast cancer) and HeLa (Cervical cancer) cell lines by the use of the MTT assay, and the IC₅₀ values were determined. The derivatives with substituted piperazines exhibited significant cytotoxicity against the tested cell lines at 5 μM and displayed low IC₅₀ values in the region of 5.8-8.8 μM for MCF-7 cell lines and 14.8-16.4 μM for HeLa cell lines when compared with the standard doxorubicin (5.4 and 14.2 μM).

Full text of DOI:10.1080/10426507.2015.1054930

Phosphorus, Sulfur, and Silicon and the Related Elements
ISSN: 1042-6507 (Print) 1563-5325 (Online) Journal homepage: http://www.tandfonline.com/loi/gpss20
Synthesis of novel n,n-disubstituted ethyl p-[2-
(1h-indol-3-yl)acetyl]phosphoramidates and their
biological activity
K. Venkata Ramana, G. Madhava, K. Ravendra Babu, D. Subbarao, K. Hema
Kumar & C. Naga Raju
To cite this article: K. Venkata Ramana, G. Madhava, K. Ravendra Babu, D. Subbarao, K. Hema
Kumar & C. Naga Raju (2015): Synthesis of novel n,n-disubstituted ethyl p-[2-(1h-indol-3-
yl)acetyl]phosphoramidates and their biological activity, Phosphorus, Sulfur, and Silicon and
the Related Elements, DOI: 10.1080/10426507.2015.1054930
To link to this article: http://dx.doi.org/10.1080/10426507.2015.1054930
Accepted online: 30 Sep 2015.
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Running Head: Synthesis of N,N-disubstituted P-[2-(1H-indol-3-yl)acetyl]phosphoramidates.
SYNTHESIS OF NOVEL N,N-DISUBSTITUTED ETHYL P-[2-(1H-INDOL-3-
YL)ACETYL]PHOSPHORAMIDATES AND THEIR BIOLOGICAL ACTIVITY
K. Venkata Ramanaa, G. Madhavaa, K. Ravendra Babua, D. Subbaraob, K. Hema Kumarc
and C. Naga Rajua*
aDepartment of Chemistry, Sri Venkateswara University, Tirupati -517 502, A.P., India.
bResearch & Development Center, API Division, Micro Labs Ltd., Jigani-Bommasandra Link
Road, Bangalore-560100, Karnataka, India.
cAptus BioSciences Private Limited, Mahabub Nagar -509002, A. P., India.
* Corresponding author E-mail address: rajuchamarthi10@gmail.com
Phone number: +91-9703193375. Fax: 0877-2289555.
Abstract
A series of novel N,N-disubstituted ethyl P-[2-(1H-indol-3-yl)acetyl]phosphoramidates were
13
31
synthesized. All title compounds were characterized by IR, NMR (1H, C, P) spectra, mass
spectra and C, H, N analysis. The anticancer activity of the title compounds was evaluated in two
human cell lines MCF-7 (breast cancer) and HeLa (Cervical cancer) cell lines by use of the MTT
assay, and the IC50 values were determined. The derivatives with substituted piperazines
exhibited significant cytotoxicity against the tested cell lines at 5 µM and displayed low IC50
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values in the region of 5.8-8.8 µM for MCF-7 cell lines and 14.8-16.4 µM for HeLa cell lines
when compared with the standard doxorubicin (5.4 and 14.2 µM).
Keywords: Indole-3-acetic acid (IAA); ketophosponates; cytotoxicity; MCF-7 cell lines; HeLa
cell lines; MTT assay.
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INTRODUCTION
Cancer is an undoubtedly serious and life threatening disease with a complex pathogenesis.
A number of natural products like taxol or vinca alkaloids have been used for the treatment of
cancer that includes DNA-alkylating agents and antimitotic agents. However, the complex
synthesis, difficult formulation, lack of oral availability makes these drugs suboptimum for
clinical treatment of cancer.1 A new era has entered into molecularly targeted therapeutics, which
is highly selective and not associated with serious toxicities of conventional cytotoxic drugs.2
Hence, the considerable interest to search for potential anticancer drugs has led to the discovery
of small synthetic molecules with anti-carcinogenic activity and limited harmful side effects.
Indole-based derivatives are commonly used heterocycles and have been identified in
various biological products as potential antibacterial, anticancer, antiviral agents and protein
3-5
inhibitors
and play a crucial role in medicinal chemistry and agrochemistry fields. Many
natural and synthetic bioactive indolyl heterocycles like 5-(3-indolyl)-oxazoles (1a),
indolylthiazoles (1b) are known for their cytotoxic activities and Meridianin (2) (Fig. 1) has
shown good anticancer activity against MCF-7 breast cancer cell lines.6
Phosphoramidates play an important role in a wide variety of structurally diverse natural
and biologically active compounds, from glycolipids to nucleic acids.7 In nucleotide and
oligonucleotide chemistry, phosphoramidites and phosphoramidates occupy a well established
position. Nucleoside phosphoramidates 8 are commonly used as basic building blocks in modern
chemical synthesis of oligonucleotides, modified oligonucleotides bearing either 31-51-P-N
bonds.9 Phosphoramidates are often used as prodrugs to increase the water solubility and hence
increase the bio-availability of the drug.10 A series of nucleoside 51-phosphoramidates and
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nucleoside 51-thiophosphoramidates were designed to act as membrane-soluble prodrugs of
bioactive free nucleotides and showed selective anti-HIV activity in MT-4 cells.11 Based on the
above facts and extensive biological nature of the indole nucleus and phosphoramidates and in
continuation of our research on the design of biologically active new phosphorus molecules,12 we
are interested in the design of some new hybrid molecules that comprise two pharmacophores in
a single molecule with an intention to enhance the biological activity. We synthesized thus a
series of novel N,N′-disubstituted ethyl P-[2-(1H-indol-3-yl)acetyl]phosphoramidates with
moderate to high yields from indoleacetic acid and diethyl substituted cyclic/phenyl/heterocyclic
phosphoramidites (7a-m). The newly synthesized molecules were tested for their anticancer
activity in two human cell lines MCF-7 (breast cancer) and HeLa (Cervical cancer) cell lines
using the MTT assay.
RESULTS AND DISCUSSION
Chemistry
A series of 13 novel N,N′-disubstituted acylphosphoramidate derivatives containing an
indole nucleus were synthesized. The synthetic route is shown in Scheme 1. The target
molecules, N,N′-disubstituted ethyl P-[2-(1H-indol-3-yl)acetyl]benzyl, -aryl, cyclic, or
heterocyclic phosphonamidates (8a-m) were synthesized by the Michaelis-Arbuzov reaction.
The title compounds were prepared in three-steps. In the first-step, indole-3-acetic acid (3) was
reacted with thionyl chloride in anhydrous THF at 0-10 °C followed by 40 °C to convert the acid
into the acid chloride (4).13 Diethyl substituted cyclic, phenyl, or heterocyclic phosphoramidites
(7a-m) were prepared by the reaction of various cyclic, phenyl, and heterocyclic amines (5a-m)
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with diethyl chlorophosphite (6) in the presence of TEA at 20-40 °C under N2 atmosphere.
Finally, the intermediates (7a-m) were reacted (Michaelis-Arbuzov reaction) with indole-3-
acetyl chloride (4) at 40-50 °C to afford the title compounds (8a-m). The keto phosphoramidates
(8a-m) were obtained in moderate to high yields. The reason might be that these molecules
enhance the nucleophilicity of the trivalent phosphorus atom which led to enhanced reactivity in
the Michaelis-Arbuzov reaction. The chemical structures of the title compounds were established
by elemental analysis, IR, NMR (1H, 13 C, 31P) spectroscopy and mass spectral data.
Spectroscopic Data
The appearance of bands at 1212-1234 cm1 of the IR spectra confirmed the formation of a
P=O function during the Michaelis-Arbuzov reaction. The bands at 3330-3390 cm1 (NH) and
1610-1660 cm1 (C=O), confirmed the presence of these functional groups in the title
1
compounds.14 In the H NMR spectra, the chemical shifts of one ethoxy group linked to
phosphorus were observed at 1.30-1.60 ppm for 3H (t), 3.50-4.80 ppm for 2H (q), 6.85-7.60 ppm
for aromatic protons, 9.90-10.80 ppm for NH (s). The 13C NMR signals at 189.0-181.2 ppm for
C=O confirmed the formation of the title compounds. In the 31P NMR spectra, the appearance of
signals at δ 21.4 to 26.8 confirms the nature of pentavalent phosphorus.15 In addition, the
13
expected C NMR chemical shift values, the molecular ion peaks and fragmented daughter ion
peaks in their mass spectra and the composition correlation of C, H, N elements in their
elemental analysis provided further evidence for the structure of title compounds (8a-m).
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Anticancer activity
The
newly
synthesized
N,N′-disubstituted
ethyl
P-[2-(1H-indol-3-yl)acetyl]
benzyl/aryl/cyclic/hetrocyclic phosphonamidates (8a-m) were evaluated for their anticancer
activity against two human cell lines MCF-7 (breast cancer) and HeLa (Cervical cancer) cell
lines by the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay24 using
doxorubicin as a positive control. The results are presented in Tables S 1 and S 2 (Supplemental
Materials). The IC50 value represents the drug concentration required to inhibit the cell growth
by 50%. Most of the title compounds exhibited good anticancer activity against breast and
cervical cancer cell lines at 5 µM concentration and reasonably low IC50 values (Table 2).
Among the title compounds, 8i, 8j, 8k and 8l exhibited good inhibitory activity against the two
human cell lines (MCF-7 and HeLa) when compared with doxorubicin. The compound 8i
exhibited better inhibitory activity compared with the remaining title compounds and nearly
equal activity at 5 µM concentration when compared (Table S 1) with the positive control. The
title compounds 8i, 8j, 8k and 8l showed an inhibitory effect against MCF-7 with IC50 values
5.8±0.82, 6.2±2.20, 6.6±1.28 and 8.8±2.14 comparable with doxorubicin (IC50 = 5.4±1.26). The
compounds 8i, 8j, 8k and 8l showed 50% inhibitory activities against HeLa cell lines with IC50
values 14.8±1.42, 15.2±1.68, 15.6±2.12 and 16.4±3.20, compared with doxorubicin (14.2±1.52).
The compound 8i has nearly the same IC50 value (14.8) when compared with doxorubicin (14.2).
Activity results proved that the aromatic, heterocyclic, or acyclic amines substituents at the
indole ring of the ketophosphoramidates play a crucial role in imparting the anticancer activity.
Hence, 8i might be considered as a lead molecule among the title compounds.
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CONCLUSIONS
In summary, a synthesis of a series of new hybrid molecules, N,N′-disubstituted ethyl P-[2-
(1H-indol-3-yl)acetyl]phosphoramidates derivatives (8a-m) was accomplished for biological
interest that encompasses two pharmacophoric units such as the indolyl and the phosphoramidite
motif with incorporation of biologically active molecules/functional groups in single compound.
Cytotoxicity was evaluated for the title compounds (8a-m) against two human cell lines, MCF-7
(breast cancer) and HeLa (Cervical cancer) cell lines using MTT assay and IC50 values were also
determined. The structure-activity studies demonstrated that molecules such as
fluorophenylpiperazinyl, pyrimidyl, nitrophenylpiperazinyl and chlorophenylpiperazinyl linked
to basic indolylphosphoramidates were mainly influenced the anticancer activity. The majority of
the title compounds exhibited good anticancer activity, where as the compounds 8i, 8j, 8k and 8l
displayed promising activity. The results endowed with a foundation for future design and
development of anticancer agents and excellent contribution for the phosphorus chemistry.
EXPERIMENTAL
All chemicals and reagents used for the synthesis were purchased from Sigma-Aldrich. All
solvents used for spectroscopic and other physical studies were reagent grade and were further
purified by literature methods.16 Melting points (m. p.) were obtained with a digital Guna
melting apparatus and are uncorrected. IR spectra (max/cm1) were recorded on a Perkin Elmer
1
283 unit as KBr discs. H, 13C, 31P NMR spectra were recorded on a Bruker 400 NMR
1
spectrometer operating at 400 MHz for H, 100 MHz for 13C and 161.9 MHz for 31P NMR using
1
13
DMSO-d6 as solvent and TMS as internal standard for H and C, and 85% H3PO4 as external
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31
standard for P. Chemical shifts (δ) and coupling constants (J) are reported in ppm and Hz
respectively. Standard abbreviations indicating multiplicity are used as follows: ‘s’ for singlet,
‘d’ for doublet, ‘dd’ for doublet of doublets, ‘t’ for triplet, ‘q’ for quartet, ‘m’ for multiplet and
‘br’ for broad. LC-MS spectra were recorded on a Jeol SX 102 DA/600 Mass spectrometer.
Elemental analyses were performed on a Thermo Finnigan Instrument at University of
Hyderabad, Hyderabad, India. Analytical thin-layer chromatography (TLC) was carried out on
precoated plates and spots were visualized with UV light.
General procedure for the synthesis of the title compounds (8a-m)
Indole-3-acetic acid (IAA) (3) (1 mmol, 0.18 g) was taken in a 50 mL round-bottomed
flask containing 10 mL of THF. SOCl2 (1.2 mmol, 0.1 mL) was added drop-wise with stirring at
0-5 °C. The reaction mixture was stirred for 2 h at 10-40 °C to afford 2-(1H-indol-3-yl)acetyl
chloride (4) and the completion of the reaction was checked by TLC. Diethyl
phosphorochloridite (6) (1.2 mmol, 0.12 mL) was dissolved in 5 mL of THF and Et3N (1.2
mmol, 0.17 mL) was added and the contents were flushed with N2. Thiophen-2-ylmethanamine
(5a) (1 mmol, 0.12 g) in 5 mL of THF was added drop-wise to 6 under a N2 atmosphere at 20-40
°C. The completion of the reaction was monitored by TLC. The precipitated Et3N·HCl was
filtered to get diethyl thiophen-2-ylmethylphosphoramidite (7a). Finally, to this filtrate (7a) a
solution of 2-(1H-indol-3-yl)acetyl chloride (4) was added at 20 °C and the reaction mixture was
stirred at 40-50 °C for 3 h under a N2 atmosphere. After completion of the reaction as indicated
by TLC, the reaction mixture was concentrated under reduced pressure to afford crude ethyl P-
[2-(1H-indol-3-yl)ethyl]-N-(thiophen-2-ylmethyl)phosphoramidate (8a). The crude product was
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purified by column chromatography using 4:1 ethyl acetate: hexane as eluent. The same
successful procedure was employed for the preparation of the remaining title compounds 8b-m.
Representative 1H, 13C and 31P NMR spectra for compound 8a are presented in the
Supplemental Materials (Figures S 1 – S 3)
Ethyl P-[2-(1H-indol-3-yl)acetyl]-N-(thiophen-2-ylmethyl)phosphoramidate (8a).
Yield: 80%; Brown solid, mp. 192-194 °C, IR: 3380 (NHAromatic), 1624 (C=O), 1218 (P=O), 914
1
(P-O-Caliphatic), 872 (P-NHaliphatic); H NMR: δ 10.24 (1H, NH), 7.54 (d, 1H, J = 5.2 Hz, ArH),
7.45 (d, 1H, J = 6.6 Hz, ArH), 7.37-7.16 (m, 3H, ArH), 7.05-6.85 (m, 3H, Ar'H), 4.56 (qt, 2H, J
= 6.4 Hz, P-OCH2CH3), 3.83 (d, 2H, J = 5.4 Hz, NHCH2), 3.54 (s, 2H, CH2), 2.26 (t, 1H, J = 6.2
13
Hz, NHCH2), 1.53 (t, 3H, J = 4.2 Hz, P-OCH2CH3); C NMR: δ 186.0 (C-11), 136.2 (C-8),
128.2 (C-4'), 127.2 (C-3'), 126.6 (C-9), 126.2 (C-5'), 125.6 (C-2'), 123.2 (C-2), 121.5 (C-6),
119.4 (C-5), 118.8 (C-4), 112.7 (C-7), 108.2 (C-3), 64.2 (C-12), 38.4 (C-14), 17.4 (C-10), 16.1
(C-13); 31P NMR: δ 22.6; LC-MS m/z (%): 360 (100) [M+]; Anal. Calcd. for C17H19N2O3PS: C,
56.66; H, 5.87; N, 15.55. Found: C, 56.61, H, 5.84; N, 15.52.
Ethyl P-[2-(1H-indol-3-yl)acetyl]-N-(pyridin-3-ylmethyl)phosphoramidate (8b)
Yield: 79%; Black solid, mp. 201-203 ºC, IR (υmax, KBr, cm-1): 3360 (NHAromatic), 1632
1
(C=O), 1222 (P=O), 916 (P-O-Caliphatic), 872 (P-NHaliphatic); H NMR (400 MHz, DMSO-d6): δ
10.34 (s, 1H, NH), 7.51 (d, 1H, J = 4.4 Hz, ArH), 7.38 (d, 1H, J = 4.6 Hz, ArH), 7.48-7.25 (m,
3H, ArH), 7.02-6.88 (4H, m, Ar'H), 4.61 (qt, 2H, J = 4.2 Hz, P-OCH2CH3), 3.78 (d, 2H, J = 5.2
Hz, NHCH2), 3.62 (s, 2H, ArCH2), 2.24 ( t, 1H, J = 5.4 Hz, NHCH2), 1.60 (3H, t, J = 6.4 Hz, P-
OCH2CH3); 13C NMR (100 MHz, DMSO-d6): 188.4 (C-11), 146.7 (C-6'), 136.5 (C-5'), 136.4 (C-
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8), 128.6 (C-4'), 127.5 (C-3'), 127.2 (C-9), 126.2 (C-2'), 123.4 (C-2), 122.1 (C-6), 119.7 (C-5),
31
118.6 (C-4), 113.2 (C-7), 109.4 (C-3), 64.8 (C-12), 39.2 (C-14), 17.6 (C-10), 16.7 (C-13); P
NMR (161.9 MHz, DMSO-d6): δ 21.4; LC-MS m/z (%): 357 (100) [M+.]; Anal. Calcd. for
C18H20N3O3P: C, 60.50; H, 5.64; N, 11.76. Found: C, 60.48, H, 5.62; N, 11.73.
Ethyl N-[2-(1H-imidazol-4-yl)ethyl-P-[(2-(1H-indol-3-yl)acetyl]phosphoramidate (8c)
Yield: 78%; Light brown solid, mp. 195-197 ºC, IR (υmax, KBr, cm-1): 3358 (NHaromatic),
1
1652 (C=O), 1224 (P=O), 918 (P-O-Caliphatic), 881 (P-NHaliphatic); H NMR (400 MHz, DMSO-
d6): δ 12.24 (s, 1H, NHimidazole), 10.37 (s, 1H, NH), 7.56 (d, 1H, J = 6.1 Hz, ArH), 7.52 (d, 1H, J
= 5.6 Hz, Ar'H), 7.42 (d, 1H, J = 4.4 Hz, Ar'H), 7.36 (d, 1H, J = 5.2 Hz, Ar-H), 7.30-7.12 (m,
3H, ArH), 4.54 (qt, 2H, J = 6.4 Hz, P-OCH2CH3), 3.72 (s, 2H, ArCH2), 3.62-3.51 (m, 2H,
NHCH2CH2), 3.01 (t, 2H, J =5.8 Hz, NHCH2CH2), 2.54 ( t, 1H, J = 4.0 Hz, NHCH2), 1.57 (t,
13
3H, J = 5.2 Hz, P-OCH2CH3); C NMR (100 MHz, DMSO-d6): δ 189.0 (C-11), 136.4 (C-8),
136.2 (C-2'), 132.5 (C-4'), 127.2 (C-9), 123.4 (C-2), 122.1 (C-6), 119.7 (C-5), 118.6 (C-4), 116.2
(C-5'), 113.2 (C-7), 109.4 (C-3), 64.8 (C-12), 42.5 (C-14), 32.6 (C-15), 17.6 (C-10), 16.7 (C-13);
31P NMR (161.9 MHz, DMSO-d6): δ 25.2; LC-MS m/z (%): 360 (100) [M+.]; Anal. Calcd. for
C17H21N4O3P: C, 56.66; H, 5.87; N, 15.55. Found: C, 56.62, H, 5.85; N, 15.52.
Ethyl P-[2-(1H-indol-3-yl)acetyl]-N-(4-fluorobenzyl)phosphoramidate (8d)
Yield: 82%; Reddish brown solid, mp. 166-168 ºC, IR (υmax, KBr, cm-1): 3338
1
(NHaromatic), 1654 (C=O), 1228 (P=O), 914 (P-O-Caliphatic), 882 (P-NHaliphatic); H-NMR (400
MHz, DMSO-d6): δ 10.72 (s, 1H, NH), 7.52 (d, 2H, J = 5.8 Hz, Ar'H), 7.48 (d, 1H, J = 5.8 Hz,
ArH), 7.42 (d, 2H, J = 5.4 Hz, Ar'H), 7.38 (d, 1H, J = 6.2 Hz, ArH), 7.28-6.84 (m, 3H, ArH),
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4.52 (qt, 2H, J = 4.4 Hz, P-OCH2CH3), 3.64 (s, 2H, ArCH2), 3.56 (d, 2H, J = 4.4 Hz, NHCH2),
13
2.62 ( t, 1H, J = 4.6 Hz, NHCH2), 1.59 (3H, t, J = 3.6 Hz, P-OCH2CH3); C NMR (100 MHz,
DMSO-d6): δ 185.0 (C-11), 160.2 (C-1'), 137.2 (C-8), 136.2 (C-4'), 127.5 (C-5'), 127.3 (C-9),
126.2 (C-3'), 123.6 (C-2), 122.3 (C-6), 119.6 (C-5), 118.9 (C-4), 116.4 (C-6'), 115.6 (C-2'), 114.5
31
(C-7), 109.6 (C-3), 64.4 (C-12), 42.5 (C-14), 42.4 (C-15), 17.2 (C-10), 16.6 (C-13); P NMR
(161.9 MHz, DMSO-d6): δ 25.6; LC-MS m/z (%): 374 (100) [M+.]; Anal. Calcd. for C19H20
FN2O3P: C, 60.96; H, 5.39; N, 7.48. Found: C, 60.92, H, 5.37; N, 7.45.
Ethyl P-[2-(1H-indol-3-yl)acetyl]-N-(3-chlorobenzyl)phosphoramidate (8e)
Yield: 84%; Black solid, mp. 151-153 ºC, IR (υmax, KBr, cm-1): 3332 (NHaromatic), 1610
1
(C=O), 1212 (P=O), 910 (P-O-Caliphatic), 862 (P-NHaliphatic); H NMR (400 MHz, DMSO-d6) δ
10.72 (s, 1H, NH), 7.48 (d, 1H, J = 5.4 Hz, ArH), 7.36 (d, 1H, J = 4.4 Hz, ArH), 7.34 (d, 2H, J =
5.2 Hz, Ar'H), 7.32-7.15 (m, 3H, ArH), 7.12 (d, 2H, J = 5.4 Hz, Ar'H), 4.48 (qt, 2H, J = 6.4 Hz,
P-OCH2CH3), 3.68 (s, 2H, ArCH2), 3.48 (d, 2H, J = 5.6 Hz, NHCH2), 2.49 ( t, 1H, J = 5.8 Hz,
13
NHCH2), 1.47 (3H, t, J = 5.6 Hz, P-OCH2CH3); C NMR (100 MHz, DMSO-d6): δ 187.0 (C-
11), 143.5 (C-5'), 137.7 (C-8), 134.2 (C-1'), 130.4 (C-6'), 127.8 (C-9), 126.6 (C-2'), 126.2 (C-3'),
124.4 (C-4'), 123.2 (C-2), 122.5 (C-6), 120.2 (C-5), 118.4 (C-4), 114.6 (C-7), 109.3 (C-3), 65.2
(C-12), 43.6 (C-14), 42.8 (C-15), 17.5 (C-10), 16.5 (C-13); 31P NMR (161.9 MHz, DMSO-d6): δ
23.0; LC-MS m/z (%): 390 (100) [M+.], 392 (33) [M+2]; Anal. Calcd. for C19H20ClN2O3P: C,
58.39; H, 5.16; N, 7.17. Found: C, 58.37, H, 5.13; N, 7.14.
Ethyl P-[2-(1H-indol-3-yl)acetyl-N-(3,4-dihydroxyphenethyl)phosphoramidate (8f)
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Yield: 77%; Brown solid, mp. 195-197 ºC, IR (υmax, KBr, cm-1): 3347 (NHaromatic), 1628
1
(C=O), 1216 (P=O), 919 (P-O-Caliphatic), 874 (P-NHaliphatic); H NMR (400 MHz, DMSO-d6): δ
10.62 (s, 1H, NH), 9.43 (s, 1H, Ar'OH), 9.32 (s, 1H, Ar'OH), 7.55 (d, 1H, J = 6.4 Hz, ArH), 7.37
(d, 1H, J = 6.2 Hz, ArH), 7.35-7.14 (m, 3H, ArH), 6.94 (d, 1H, J = 5.6Hz, Ar'H), 6.87 (d, 1H, J =
6.4 Hz, Ar'H), 6.82 (s, 1H, Ar'H), 4.51 (qt, 2H, J = 5.2 Hz, P-OCH2CH3), 3.46 (s, 2H, ArCH2),
2.72 (s, 2H, HNCH2), 2.68 (t, 2H, J =4.8 Hz, NCH2CH2), 2.45 (t, 1H, J = 5.8 Hz, NHCH2), 1.37
(3H, t, J = 5.4 Hz, P-OCH2CH3); 13C NMR (100 MHz, DMSO-d6): δ 182.4 (C-11), 145.2 (C-3'),
143.5 (C-4'), 137.7 (C-8), 135.4 (C-1'), 126.5 (C-9), 124.2 (C-6), 122.5 (C-2), 122.3 (C-6'); 121.6
(C-5), 118.6 (C-4), 117.4 (C-7), 116.4 (C-2'), 116.2 (C-5'), 107.4 (C-3), 64.5 (C-12), 47.4 (C-14),
31
39.2 (C-15), 17.8 (C-10), 17.7 (C-13); P NMR (161.9 MHz, DMSO-d6): δ 22.8; LC-MS m/z
(%): 402 (100) [M+.]; Anal. Calcd. for C20H23N2O5P: C, 59.70; H, 5.76; N, 6.96. Found: C,
59.67, H, 5.74; N, 6.95.
Ethyl 2-(1H-indol-3-yl)acetyl(4-methylpiperazin-1-yl)phosphinate (8g)
Yield: 83%; Black solid, mp. 146-148 ºC, IR (υmax, KBr, cm-1): 3330 (NHaromatic), 1642
(C=O), 1220 (P=O), 914 (P-O-Caliphatic), 856 (P-Naliphatic); 1H NMR (400 MHz, DMSO-d6): δ 9.90
(s, 1H, NH), 7.45 (d, 1H, J = 6.2 Hz, ArH), 7.42 (d, 1H, J = 5.6 Hz, ArH), 7.32-6.88 (m, 3H,
ArH), 3.68 (s, 2H, ArCH2), 3.50 (qt, 2H, J = 5.2 Hz, P-OCH2CH3), 2.48 (s, 2H, NCH2), 2.38 (t,
2H, J = 4.4 Hz, NCH2CH2), 2.35 (t, 2H, J = 4.2 Hz, NCH2CH2), 2.31 (s, 2H, NCH2), 1.82 (s, 3H,
NCH3), 1.30 (3H, t, J = 4.8 Hz, P-OCH2CH3); 13C NMR (100 MHz, DMSO-d6): δ 182.0 (C-11),
138.1 (C-8), 125.2 (C-9), 123.7 (C-2), 122.2 (C-6), 120.5 (C-5), 118.7 (C-4), 115.2 (C-7), 108.5
(C-3), 65.3 (C-12), 56.6 (C-3'), 56.3 (C-5'), 47.2 (C-2'), 45.7 (C-6'), 21.4 (C-4'), 17.9 (C-10), 16.1
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31
(C-13); P NMR (161.9 MHz, DMSO-d6): δ 23.5; LC-MS m/z (%): 349 (100) [M+.]; Anal.
Calcd. for C17H24N3O3P: C, 58.44; H, 6.92; N, 12.03. Found: C, 58.40, H, 6.89; N, 12.01.
Ethyl 2-(1H-indol-3-yl)acetyl(thiomorpholino)phosphinate (8h)
Yield: 85%; Brown solid, mp. 166-168 ºC, IR (υmax, KBr, cm-1): 3342 (NHaromatic), 1639
1
(C=O), 1216 (P=O), 918 (P-O-Caliphatic), 851 (P-Naliphatic); H NMR (400 MHz, DMSO-d6): δ
10.41 (s, 1H, NH), 7.45 (d, 1H, J = 5.7 Hz, ArH), 7.42 (d, 1H, J = 5.2 Hz, ArH), 7.36-7.12 (m,
3H, ArH), 3.68 (qt, 2H, J = 5.4 Hz, P-OCH2CH3), 3.36 (s, 2H, ArCH2), 2.42 (s, 2H, NCH2), 2.37
( s, 2H, NCH2), 2.34 (t, 2H, J = 4.0 Hz, NCH2CH2), 2.28 (t, 2H, J = 3.8 Hz, NCH2CH2), 1.50
(3H, t, J = 4.6 Hz, P-OCH2CH3); 13C NMR (100 MHz, DMSO-d6) δ: 183.0 (C-11), 138.4 (C-8),
125.6 (C-9), 122.8 (C-2), 122.6 (C-6), 121.2 (C-5), 118.6 (C-4), 115.5 (C-7), 108.9 (C-3), 65.6
31
(C-12), 46.5 (C-6'), 46.2 (C-2'), 29.6 (C-3'), 29.3 (C-5'), 17.5 (C-10), 16.4 (C-13); P NMR
(161.9 MHz, DMSO-d6): δ 24.2; m/z (%): 352 (100) [M+.]; Anal. Calcd. for C16H21N2O3PS: C,
54.53; H, 6.01; N, 7.95. Found: C, 54.51, H, 5.97; N, 7.92.
Ethyl 2-(1H-indol-3-yl)acetyl[4-(4-fluorophenyl)piperazin-1-yl]phosphinate (8i)
Yield: 86%; Dark brown solid, mp. 175-177 ºC, IR (υmax, KBr, cm-1): 3390 (NHaromatic),
1635 (C=O), 1232 (P=O), 912 (P-O-Caliphatic), 847 (P-Naliphatic); 1H NMR (400 MHz, DMSO-d6):
δ 10.80 (s, 1H, NH), 7.38 (d, 1H, J = 5.8 Hz, ArH), 7.32 (d, 1H, J = 5.4 Hz, ArH), 7.30-7.18 (m,
3H, ArH), 7.12 (d, 2H, J = 6.4 Hz, Ar'H), 6.78 (d, 2H, J = 6.2 Hz, Ar'H), 4.32 (qt, 2H, J = 5.2
Hz, P-OCH2CH3), 3.18 (s, 2H, ArCH2), 2.32 (s, 2H, NCH2), 2.27 ( s, 2H, NCH2), 2.18 (t, 2H, J =
4.4 Hz,NCH2CH2), 2.14 (t, 2H, J = 4.6 Hz, NCH2CH2), 1.44 (3H, t, J = 5.4 Hz, P-OCH2CH3);
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13C NMR (100 MHz, DMSO-d6): δ 181.4 (C-11), 156.5 (C-10'), 145.6 (C-7'), 138.1 (C-8), 125.7
(C-9), 123.4 (C-2), 122.2 (C-6), 121.6 (C-5), 118.4 (C-4), 117.4 (C-9'), 116.4 (C-11'), 116.2 (C-
8'), 115.7 (C-7), 115.4 (C-12'), 108.2 (C-3), 65.4 (C-12), 46.8 (C-6'), 46.7 (C-2'), 29.5 (C-5'),
31
28.6 (C-3'), 17.2 (C-10), 16.2 (C-13); P NMR (161.9 MHz, DMSO-d6): δ 26.8; LC-MS m/z
(%): 429 (100) [M+.]; Anal. Calcd. for C22H25 FN3O3P: C, 61.53; H, 5.87; N, 9.79. Found: C,
61.48, H, 5.83; N, 9.74.
Ethyl 2-(1H-indol-3-yl)acetyl[4-(pyrimidin-2-yl)piperazin-1-yl]phosphinate (8j)
Yield: 87%; Reddish brown solid, mp. 144-146 ºC, IR (υmax, KBr, cm-1): 3342
(NHaromatic), 1630 (C=O), 1230 (P=O), 920 (P-O-Caliphatic), 836 (P-Naliphatic); 1H NMR (400 MHz,
DMSO-d6): δ 10.28 (s, 1H, NH), 7.56 (d, 2H, J = 5.8 Hz, Ar'H), 7.34 (d, 1H, J = 6.2 Hz, ArH),
7.25 (d, 1H, J = 5.8 Hz, ArH), 7.32-7.18 (m, 3H, ArH), 7.12 (t, 1H, J = 4.4 Hz, Ar'H), 4.38 (qt,
2H, J = 3.6 Hz, P-OCH2CH3), 3.26 (s, 2H, ArCH2), 2.32 (s, 2H, NCH2), 2.28 ( s, 2H, NCH2),
2.16 (t, 2H, J = 4.4 Hz, NCH2CH2), 2.12 (t, 2H, J = 4.6 Hz, NCH2CH2), 1.48 (3H, t, J = 4.2 Hz,
13
P-OCH2CH3); C NMR (100 MHz, DMSO-d6): δ 181.2 (C-11), 161.6 (C-7'), 156.8 (C-11'),
156.2 (C-9'), 138.5 (C-8), 125.4 (C-9), 125.2 (C-2), 122.6 (C-6), 121.5 (C-5), 117.5 (C-4), 116.5
(C-10'), 115.8 (C-7), 108.4 (C-3), 64.6 (C-12), 49.8 (C-5'), 49.4 (C-3'), 47.4 (C-2'), 47.0 (C-6'),
17.6 (C-10), 16.5 (C-13); 31P NMR (161.9 MHz, DMSO-d6): δ 22.8; LC-MS m/z (%): 413 (100)
[M+.]; Anal. Calcd. for C20H24 N5O3P: C, 58.11; H, 5.85; N, 16.94. Found: C, 58.09, H, 5.82; N,
16.92.
Ethyl 2-(1H-indol-3-yl)acetyl[4-(4-nitrophenyl)piperazin-1-yl]phosphinate (8k)
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Yield: 74%; Brown solid, mp. 174-178 ºC, IR (υmax, KBr, cm-1): 3336 (NHaromatic), 1660
1
(C=O), 1234 (P=O), 924 (P-O-Caliphatic), 845 (P-Naliphatic); H NMR (400 MHz, DMSO-d6): δ
10.54 (s, 1H, NH), 7.54 (d, 2H, J = 6.4 Hz, Ar'H), 7.14 (d, 2H, J = 6.6 Hz, Ar'H), 7.42 (d, 1H, J
= 6.4 Hz, ArH), 7.32 (d, 1H, J = 6.2 Hz, ArH), 7.28-7.11 (m, 3H, ArH), 4.80 (qt, 2H, J = 4.0 Hz,
P-OCH2CH3), 3.14 (s, 2H, ArCH2), 2.28 (s, 2H, NCH2), 2.24 ( s, 2H, NCH2), 2.14 (t, 2H, J = 4.2
13
Hz, NCH2CH2), 2.12 (t, 2H, J = 4.4 Hz, NCH2CH2), 1.56 (3H, t, J = 3.6 Hz, P-OCH2CH3), C
NMR (100 MHz, DMSO-d6): δ 182.7 (C-11), 155.2 (C-7'), 138.4 (C-8), 136.2 (C-10'), 126.2 (C-
11'), 125.3 (C-9), 124.5 (C-9'), 123.5 (C-2), 122.5 (C-6), 121.4 (C-5), 118.7 (C-4), 116.0 (C-7),
112.4 (C-8'), 112.2 (C-12'), 107.6 (C-3), 64.2 (C-12), 46.9 (C-2'), 46.5 (C-6'), 29.7 (C-5'), 28.4
31
(C-3'), 17.6 (C-10), 16.8 (C-13); P NMR (161.9 MHz, DMSO-d6): δ 24.6; LC-MS m/z (%):
456 (100) [M+.]; Anal. Calcd. for C22H25 N4O5P: C, 57.89; H, 5.52; N, 12.27. Found: C, 57.84,
H, 5.49; N, 12.22.
Ethyl 2-(1H-indol-3-yl)acetyl[4-(4-chlorophenyl)piperazin-1-yl]phosphinate (8l)
Yield: 75%; Dark red solid, mp. 184-186 ºC. IR (υmax, KBr, cm-1) 3346 (N-Haromatic),
1
1642 (C=O), 1226 (P=O), 921 (P-O-Caliphatic), 864 (P-N aliphatic), cm-1; H NMR (400 MHz,
DMSO-d6): δ 10.62 (s, 1H, NH), 7.52 (d, 1H, J = 6.4 Hz, ArH), 7.44 (d, 1H, J = 6.2 Hz, ArH),
7.36 (d, 2H, J = 6.8 Hz, Ar'H), 7.34-7.16 (m, 3H, ArH), 6.74 (d, 2H, J = 7.2 Hz, Ar'H), 4.54 (qt,
2H, J = 3.8 Hz, P-OCH2CH3), 3.12 (s, 2H, ArCH2), 2.34 (s, 2H, NCH2), 2.26 (s, 2H, NCH2),
2.19 (t, 2H, J = 4.0 Hz, NCH2CH2), 2.17 (t, 2H, J = 4.2 Hz, NCH2CH2), 1.58 (3H, t, J = 4.0 Hz,
P-OCH2CH3); 13C NMR (100 MHz, DMSO-d6): δ 185.7 (C-11), 148.2 (C-7'), 138.7 (C-8), 129.5
(C-9'), 129.4 (C-10'), 128.2 (C-11'), 125.6 (C-9), 123.6 (C-2), 122.8 (C-6), 121.5 (C-5), 118.9
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(C-4), 116.4 (C-7), 115.6 (C-12'), 106.4 (C-8') 107.3 (C-3), 64.6 (C-12), 47.2 (C-2'), 46.8 (C-6'),
30.2 (C-5'), 29.5 (C-3'), 17.3 (C-13), 17.2 (C-10); 31P NMR (161.9 MHz, DMSO-d6): δ 23.8; LC-
MS m/z (%): 445 (100) [M+.], 447 (33) [M+2]; Anal. Calcd. for C22H25 ClN3O3P: C, 59.26; H,
5.65; N, 9.42. Found: C, 59.20, H, 5.62; N, 9.38.
Ethyl 2-(1H-indol-3-yl)acetyl[4-(pyridin-2-yl)piperazin-1-yl]phosphinate (8m)
Yield: 76%; Brown, mp. 174-178 ºC, IR (υmax, KBr, cm-1): 3352 (N-H Aromatic), 1648
1
(C=O), 1226 (P=O), 926 (P-O-Caliphatic), 847 (P-Naliphatic); H NMR (400 MHz, DMSO-d6): δ
10.54 (s, 1H, NH), 8.06 (t, 1H, J = 5.4 Hz, Ar'H), 7.58 (t, 1H, J = 5.8 Hz, Ar'H), 7.52 (d, 1H, J =
6.2 Hz, Ar'H), 7.38 (d, 1H, J = 6.2 Hz, ArH), 7.36 (d, 1H, J = 5.8 Hz, ArH), 7.35-7.15 (m, 3H,
ArH), 6.74 (d, 1H, J = 6.6 Hz, Ar'H), 4.61 (qt, 2H, J = 4.2 Hz, P-OCH2CH3), 3.25 (s, 2H,
ArCH2), 2.37 (s, 2H, NCH2), 2.28 ( s, 2H, NCH2), 2.21 (t, 2H, J = 4.2 Hz, NCH2CH2), 2.19 (t,
2H, J = 4.4 Hz, NCH2CH2), 1.64 (3H, t, J = 3.8 Hz, P-OCH2CH3); 13C NMR (100 MHz, DMSO-
d6): δ 188.2 (C-11), 158.4 (C-7'), 148.4 (C-11'), 139.4 (C-9'), 138.4 (C-8), 125.8 (C-9), 123.2 (C-
6), 122.8 (C-2), 121.8 (C-5), 119.2 (C-4), 117.6 (C-10'), 116.7 (C-7), 107.6 (C-3), 106.4 (C-8'),
64.8 (C-12), 47.4 (C-2'), 46.9 (C-6'), 30.4 (C-5'), 29.8 (C-3'), 17.5 (C-13), 17.4 (C-10); 31P NMR
(161.9 MHz, DMSO-d6): δ 25.2; LC-MS m/z (%): 412 (100) [M+.]; Anal. Calcd. for
C21H25N4O3P: C, 59.06; H, 6.00; N, 14.50. Found: C, 59.01, H, 5.97; N, 14.45.
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ACKNOWLEDGEMENTS
The author K. Venkata Ramana expresses his grateful thanks to DST-PURSE Programme,
S. V. University, Tirupati, for giving financial assistance through DST-PURSE- JRF Fellowship.
One of the authors, G. Madhava is thankful to University Grants Commission (UGC), New
Delhi, India for providing financial assistance by awarding a Senior Research Fellowship (SRF).
The authors acknowledge with gratitude the help of Aptus Bio-Sciences Private Limited,
Mahabubnagar-509002, A.P, India and Hyderabad Central University, India for providing
anticancer activity and spectral data respectively.
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REFERENCES
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[3]
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[4]
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Hanna-Elias, A.; Manallack, D. T.; Berque-Bestel, I.; Irving, H. R.; Coupar, I. M.;
Iskander, M. N. Eur. J. Med. Chem. 2009, 44, 2952-2959.
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Schmidt, J. M. J. Nat. Prod. 2002, 65, 1793-1797.
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601.
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N
R
N
O
EtO
C
NH2
Y
P
N
N
Y = O and S
O
N
H
N
H
Indolyloxazoles (1a)
Indolylthiazoles (1b)
N
Titile compounds 8(a-m)
Meridianin (2)
H
Figure 1. Two biologically active anticancer agents having indole nucleus
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EtO
EtO
P Cl
R-H
+
5(a-m)
6
E
T
t
3
H
N
F
,
,
2
N
0
2
-
4
0
Et3N.HCl
o
O
C
EtO
C
P
R
Cl
OH
EtO
THF, N2
40-50 oC
SOCl2/ THF
C
C
O
P R
+
O
10-40 oC
O
EtO
N
H
N
H
N
H
8(a-m)
7 (a-m)
3
4
Compound
Compound
R
R
Compound
R
H
N
N
N
N
N
F
NH
8i
8j
8a
8b
8e
S
Cl
N
N
H
N
N
H
OH
OH
N
NH
8f
8k
8l
N
N
NO2
Cl
N
8c
N
N
8g
H
N
N
N
Me
H
N
N
N
S
8h
8m
N
N
8d
F
Scheme 1. Synthesis of ethyl P-2-(1H-indol-3-yl)acetyl-N-substituted phosphoramidates (8a-m).
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