Synthesis, characterization, computational and biological study of novel azabenzo[a]phenothiazine and azabenzo[b]phenoxazine heterocycles as potential antibiotic agent

Article abstract of DOI:10.1007/s00044-017-2131-3

Two angular phenothiazines and one angular phenoxazine were successfully synthesized via anhydrous base condensation reaction of 2,6-diamino-4-chloropyrimidine-5-thiol, with 7-chloro-5,8-quinolinequinone,2-aminothiophenol and 2-aminophenol, respectively. Condensation reaction between 2-6-diamino-4-chloropyrimidine-5-thiol and 7-chloro-5,8-quinolinequinone in the presence of anhydrous sodium carbonate yielded 10-amino-8-chloro-1,9,11-triaza-5H-benze[a]phenothiazine-5-one, 1-aza-5H-benzo[a]phenothiazine-5-one and 1-aza-5H-benzo[a]phenoxazine-5-one were produced with anhydrous basic condensation between 7-chloro-5,8-quinolinequinone and 2-aminothiophenol and 2-aminophenol respectively. These angular azaphenothiazin-5-ones and angular azaphenoxazine-5-one were converted to their derivatives via palladium(o)/piperazine ligand utilizing Mizoroki–Heck cross coupling tandem reaction to obtain six derivatized compounds. The synthesized compounds are intensely coloured and their structural elucidation were established by combined spectroscopic and elemental analytical data. In silico and in vitro screening methods were used to investigate the antibacterial potencies of the compounds. All the compounds, except one, interacted with Type I SPase, an unconventional validated antibiotic enzyme targeted in combating antibacterial resistant, at low micromolar range. They also showed activity against the tested bacteria: Bacillus cereus, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa. In fact, B. cereus exhibited more susceptibility towards four of the compounds than the standard drug—ciprofloxacin. The predicted binding modes of four compounds with outstanding activities were finally studied to identify vital ligand–protein interactions, which can serve as template during activity optimization process.

Full text of DOI:10.1007/s00044-017-2131-3

MEDICINAL
CHEMISTRY
RESEARCH
Medicinal Chemistry Research
https://doi.org/10.1007/s00044-017-2131-3
ORIGINAL RESEARCH
Synthesis, characterization, computational and biological study of
novel azabenzo[a]phenothiazine and azabenzo[b]phenoxazine
heterocycles as potential antibiotic agent
Fidelia N. Ibeanu1 Efeturi A. Onoabedje2 Akachukwu Ibezim3,4 Uchechukwu C. Okoro2
●
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Received: 2 October 2017 / Accepted: 29 December 2017
© Springer Science+Business Media, LLC, part of Springer Nature 2018
Abstract
Two angular phenothiazines and one angular phenoxazine were successfully synthesized via anhydrous base condensation
reaction of 2,6-diamino-4-chloropyrimidine-5-thiol, with 7-chloro-5,8-quinolinequinone,2-aminothiophenol and 2-amino-
phenol, respectively. Condensation reaction between 2-6-diamino-4-chloropyrimidine-5-thiol and 7-chloro-5,8-quinoline-
quinone in the presence of anhydrous sodium carbonate yielded 10-amino-8-chloro-1,9,11-triaza-5H-benze[a]phenothiazine-
5-one, 1-aza-5H-benzo[a]phenothiazine-5-one and 1-aza-5H-benzo[a]phenoxazine-5-one were produced with anhydrous
basic condensation between 7-chloro-5,8-quinolinequinone and 2-aminothiophenol and 2-aminophenol respectively. These
angular azaphenothiazin-5-ones and angular azaphenoxazine-5-one were converted to their derivatives via palladium(o)/
piperazine ligand utilizing Mizoroki–Heck cross coupling tandem reaction to obtain six derivatized compounds. The
synthesized compounds are intensely coloured and their structural elucidation were established by combined spectroscopic
and elemental analytical data. In silico and in vitro screening methods were used to investigate the antibacterial potencies of
the compounds. All the compounds, except one, interacted with Type I SPase, an unconventional validated antibiotic
enzyme targeted in combating antibacterial resistant, at low micromolar range. They also showed activity against the tested
bacteria: Bacillus cereus, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa. In fact, B. cereus exhibited
more susceptibility towards four of the compounds than the standard drug—ciprofloxacin. The predicted binding modes of
four compounds with outstanding activities were finally studied to identify vital ligand–protein interactions, which can serve
as template during activity optimization process.
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Keywords Phenoxazine Phenothiazine Benzophenothiazine Benzophenoxazine Antimicrobial Antibiotic.
Introduction
Previous reports have highlighted the numerous usefulness
of phenothiazine 1 and phenoxazine 2 and their derivatives
most especially as drugs (Pluta et al. 2011; Okafor 1986;
Onoabedje et al. 2017) (Scheme 1).
* Efeturi A. Onoabedje
efeturi.onoabedje@unn.edu.ng
* Akachukwu Ibezim
H
N
akachukwu.ibezim@unn.edu.ng
1
Natural Science Unit, School of General Studies, University of
Nigeria, Nsukka, Nigeria
X
2
Department of Pure and Industrial Chemistry, University of
Nigeria, Nsukka, Nigeria
X = 1 ( S ) ; 2 ( O )
3
Department of Pharmaceutical and Medicinal Chemistry,
University of Nigeria, Nsukka, Nigeria
The phenothiazine and phenoxazine nuclei are the pha-
macophores in various antimicrobial (Onoabedje et al.
2016; Wainwright et al. 2012) antimalarial (Ge et al. 2010;
Wesolowska et al. 2006), sedative, tranquilizer (Okafor
4
Skaggs School of Pharmacy and Pharmaceutical Sciences, Center
for Discovery and Innovation in Parasitic Diseases, University of
California, San Diego, 9500 Gilman Drive, PSB 4262, La Jolla,
CA 92093, USA

Medicinal Chemistry Research
Cl
Scheme 1 Synthesis of 2,4-
diamino-6-chloropyrimidine- 5-
thiol
Cl
N
Cl
KSCN, Br2
SCN
NH2
N
SH
20 % KOH
heat, H3O+
N
N
gl. AcOH
-5 - 0 oC
H2N
N
2
NH2
H2N
H2N
N
3
NH2
NO2
OH
i. Conc HNO3/H2O
ii. KOH
NO
OH
Scheme 2 Synthesis of 7-
chloroquinoline-5,8-dione
NaNO2, Conc HCl
0 - 5 oC, H2O
. HCl
iii. AcOH
N
N
N
OH
4
i. KOH/H2O
ii. 5% excess NaOCl
iii. AcOH
O
NH2
NO2
KOH, H2O
K2Cr2O7
N
Cl
6 M H2SO4, H2O,
N
Cl
Na2S2O4, N2 g
Cl
N
O
5
OH
OH
1977), antituberculotic (Okafor et al. 1981; Sridhar et al.
2015), antiproliferative, immunosupressive, antiviral (Weso-
lowska et al. 2006) and anticancer (Pluta et al. 2010; Thim-
maiah et al. 1992) agents. While phenothiazine does not have
their core in the world of natural compounds, phenoxazine is
well known components of actinomycin D, an antibiotic
produced by Streptomyces antibioticus (Pluta et al. 2010;
Jaszczyn et al. 2012). Natural products containing phenox-
azine moiety exhibit anticancer, antibacterial, antiviral and
antifugal properties (Barness et al. 2015). Besides the
numerous biological importance, many phenothiazine and
phenoxazine derivatives are fluoresecent, they form stable
radical cations which are used as chromophores for photo-
induced electron transfer experiments and organic light
emitting diodes (Turdean et al. 2009; Sailer et al. 2006; Zhu
et al. 2005; Okamoto et al. 2005). The versatile utility of
phenothiazine and phenoxazine have motivated their intensive
and extensive structural modifications in order to obtain better
drugs and scientific materials. We have recently reported the
synthesis of new aryl, styryl, alkynyl, thiophenyl and furanyl
derivatives of phenoxazine and phenothiazines which pos-
sesses broad and potent antimicrobial activity (Onoabedje
et al. 2016) (Scheme 2). Therefore, as an extension of the
previous work, we are reporting a convenient protocol for the
transformation of angular chlorophenothiazines and angular
phenoxazines and their antimicrobial properties.
purification. All compounds were synthesized in the senior
research chemistry laboratory of Department of Pure and
Industrial chemistry, University of Nigeria, Nsukka. Melt-
ing points were determined with a Fisher-John apparatus
and were uncorrected. Most of the reactions were carried
out in inert atmosphere (Nitogen atmosphere). UV-visible
and IR spectra were recorded on UV2500PC series using
matched 1 cm quartz cells and on a Shimadzu FTIR-8400s
Fourier Transform Infrared (KBr pellets) respectively in
National Research Institute Chemical Technology, Zaria.
Nuclear Magnetic resonance (1HNMR and 13C NMR)
spectra were determined using a Bruker AV-400 and a
JEOL-JNM LA-400 spectrometer from Strathclyde Institute
of Pharmacy and Biomedical Science, University of
Strathclyde, Glasglow, Scotland. Chemical shifts are
recorded on the δ-scale(neat) and coupling constant (J)
reported in hertz. Multiplicity indicated using the following
abbreviations; d for doublet, dd for doublet of doublet, t for
triplet and m for multiplet. Elemental analysis was carried
out at the Central Science Laboratory, University of Cairo,
Egypt on a CE440 Elemental Analyzer.
Molecular simulation
X-ray crystal structure of bacteria Type I signal peptidase
(Type I SPase) in complex with its substrate (PDB code
4WVI) was retrieved from protein databank (Berman et al.
2000). Molecular operating environment (MOE) software
was used to treat the protein as described in our earlier work
(Ntie-Kang et al. 2014). The graphical user interface and
QuSAR module implemented in MOE package were
respectively used to generate the three dimensional struc-
tures and calculate the following molecular descriptors
(after energy minimizing the molecules to a 0.001 kcal/mol
Experimental section
General information
Starting materials and reagents were purchased from Sigma-
Aldrich chemical company and were used without further

Medicinal Chemistry Research
energy gradient using Merck Molecular (MMFF94) force-
field (Halgren 1996): molecular weight (MW), lipophilicity
(log P), hydrogen bond acceptor/donor (HBA/HBD) and
number of rotatable bond (NRB) of the newly synthesized
phenothiazines and phenoxazines.
AutoDock 4.2.0 was used to carry out docking compu-
tation (Morris et al. 1998). A grid box size of 40 40 40 A3
points (spacing between the grid points of 0.375 Å) was
employed which centered on the mass center (10.631,
29.727, −3.746) of the crystallographic macromolecule
encompassing all the active site atoms. The dock protocol
used was validated by ensuring that it gives a docked pose
of the substrate which differs from that of the X-ray crys-
tallography by less than 2.0 Å.
H), 9.32–9.19 (1 H, m, Ar-H), 8.66–8.56 (2 H, m, Ar-H),
7.94–7.34 (4 H, m, Ar-H). 13C NMR (DMSO6, 100 MHz):
180.7 (C=O), 149.4 (C), 148.8 (C), 147.9 (C), 144.6 (C),
142.4 (C), 141.1 (C), 140.3 (C), 132.1 (CH), 129.3 (CH),
128.5 (CH), 126.4 (CH), 125.8 (CH), 120.0 (CH), 117.0
(CH). (Anal.calcd. for C15H8N2OS: C, 68.18; H, 3.03.
Found: C, 68.30; H, 2.98).
1-Aza-5H-benzo[a]phenoxazin-5-one 13
A reaction mixture containing 2–aminophenol (0. 47 g, 5
mmol), anhydrous sodium carbonate (1.06 g, 10 mmol) and
benzene (40 ml) mixed with DMF (5 ml) in a round bottom
flask was refluxed at 70–75 °C for 45 min while stirring.
7–Chloro–5,8–quinolinequinone (0.97 g, 5 mmol) was
added and stirring continued at the same temperature for 6
h. At the end of the reaction, solvent evaporated in vacuum,
and the crude product recrystallized from acetone. Yield =
1.15 g (87%), mp 210–212 °C (dec.). UV-visible (DMSO)
λmax 366 nm (4.00), 450 (3.22). IR(νmax,cm−1): 3109 (Ar-
10-Amino-8-chloro-1,9,11-triaza-5H-benzo[a]
phenothiazine-5-one 6
To a suspension of 2,6-diamino-4-chloropyrimidine-5-thiol
(0.88 g, 5 mmol) in benzene (40 ml) mixed with dimethyl-
formamide (DMF), (5 ml) was added Na2CO3 (1.06 g,10
mmol).The mixture was heated to boiling temperature
before adding 7-chloro-5,8-quinolinequinone (0.97 g, 5
mM) and entire mixture refluxed for 6 h while stirring in a
round bottom flask. At the end of the reflux period the
mixture was filtered and the filtrate exposed in a dish for the
solvent to evaporate. The yellow-brown solid product was
recrystallized from acetone, after treatment with activated
charcoal, filtered and air-dried. Yield = 0.90 g (57%), mp
198–200 °C. UV-visible (DMSO) λmax 366.5 (4.00), 440
(3.76), 753(2.05). IR (νmax, cm−1): 3380 (–NH2), 1657
(C=O), 1589,1566 (C=C), 1494, 1274,1172, 925, 881,
806, 746, 677, 630. 1H NMR (DMSO-d6, 400 MHz,): δ 9.42
9.19 (1 H, m, Ar-H); 8.63–8.44 (2 H, m, Ar-H), 7.94–7.80
(1 H, m, Ar-H) 4.87(2 H, s, br, NH2). 13C NMR (DMSO-d6,
100): 181.5 (C=O), 167.7 (C), 162.9 (C), 148.5(C), 146.7
(C), 136.1 (CH), 135.4 (CH), 132.1 (CH), 130.7 (CH),
128.4 (CH), 126.4 (CH), 124.5 (CH), 115.97 (CH). (Anal.
Calcd. for C13H6ClN5OS: C, 64.56; H, 2.81. Found: C,
64.60; H, 2.77).
1
H); 1656 (C=O), 1575, 1276, 1200, 910, 876, 780 710. H
NMR (DMSO-d6, 400 MHz,): δ 9.42–9.21 (1 H, m, Ar-H),
8.64–8.20 (2H, m, Ar-H), 7.94–7.76 (1 H, m, Ar-H),
7.10–7.00 (1 H, m, Ar-H), 6.76–6.42 (3H, m, Ar-H). 13C
NMR (DMSO6, 100 MHz): 175.1 (C=O), 152.4 (C), 150.3
(C), 149.5 (C), 145.8 (C), 143.6 (C), 142.0 (C), 132.1 (CH),
131.7 (CH), 129.0 (CH), 126.3 (CH), 124.5 (CH), 117.1
(CH), 116.7 (CH), 115.4 (CH). (Anal.calcd. for C15H8N2O2:
C, 72.58; H, 3.32. Found C,73.15; H, 3.32).
The general procedure for the Mizoroki–Heck cross-
coupling reactions
In a round bottom flask equipped with a stirrer, piperazine
ligand (0.005 g, 0.01 mmol), dppb(PdCl2) catalyst (0.006 g,
0.01 mmol) and methanol (10 ml) were stirred at room
temperature under nitrogen atmosphere. After stirring for 5
min an intimate mixture of 10-amino-8-chloro-1,9,11-
triaza-5-benzo[a]phenothiazin-5-one (0.947 g, 3 mmol), aryl
halide (1 mmol) and K2CO3 (0.193 g, 1.4 mmol) were
added. The reaction mixture was heated to 60 °C for 4 h.
After the reflux period, the solvent was removed by vacuum
distillation. Crushed ice (5 g) was added to the resultant
residual mixture which precipitated out the solid product.
The product was further recrystallized from aqueous etha-
nol. The general procedure was employed in synthesing
compounds 7, 8, 11–15.
1-Aza-5H-benzo[a]phenothiazin-5-one 10
Compound 10 was prepared by coupling of 2-
aminothiophenol (0.63 g 5 mmol) with 7-chloro-5,8-quino-
linequinone (0.97 g, 5 Mm) following similar procedure for
synthesis of compound 6. The crude product was recrys-
tallized from acetone after treatment with activated charcoal
to give brown coloured solid. Yield = 1.20 g (91%), mp
130–132 °C. UV-visible (DMSO) λmax 366.5 (4.00). 410
(2.44). IR(νmax, cm−1): 1648 (C=O), 1609 (C=C), 1568,
1479, 1392, 1276, 1178, 1147, 1018, 926, 876, 750, 678.
1H NMR (DMSO-d6, 400 MHz,): δ 9.48 - 9.40 (1 H, m, Ar-
10-Amino-8-chloro-6(4-nitrophenyl)1,9,11-triaza-5H-
benzo[a]phenothiazin-5-one 7
The coupling of 10-amino-8-chloro-1,9,11-triaza-5H-benzo
[a]phenothiazin-5-one
(0.947 g,
3 mmol),
and

Medicinal Chemistry Research
4-iodonitrobenzene (0.248 g, 1 mmol) afforded compound 7
as brown solid, % yield (1.10 g, 84%). Mp > 250 °C (dec).
UV-visible (DMSO) λmax 367(4.00), 500 (3.42), 508 (3.42),
750(1.07), 779(1.072). IR (νmax,cm−1): 3350 (–NH2,), 1645
(C = O), 1591, 1404, 1334, 1255, 1168, 1004, 848, 744,
682. 1H NMR (DMSO-d6, 400 MHz,): δ 9.45–9.42 (1 H, dd,
J = 8.93, 18.84 Hz), 8.19–8.04 (1 H, d, J = 8.50 Hz),
7.97–7.55 (3 H, m), 6.73–6.58 (2 H, m), 3.50 (2 H, s,
–NH2). 13C NMR (DMSO6, 100 MHz): 178.22 (C=O),
156.3 (C), 155.10 (C), 153.0 (C), 150.9 (C), 142.9 (C),
143.91 (C), 142.45 (C), 139.3 (C), 135.9 (C), 134.1 (C),
132.2 (C), 130.9 (C), 126.4 (CH), 125.0 (CH), 122.7 (CH),
119.3 (CH), 119.7 (CH), 112.6 (CH). (Anal. calcd. for
C19H9N5SO3Cl: C, 52.12; H, 2.30; N, 19.19; S, 7.32.
Found: C, 51.72; H, 2.01; N,19.10; S, 7.02).
(CH), 127.8 (CH), 124.6 (CH), 120.2 (CH), 119.2 (CH),
118.7 (CH), 115.9 (CH). Anal. calcd. for C21H11N3SO3:
C,65.45; H,2.85. Found: C, 65.05; H, 2.45.
6-(4-Carboxyphenyl)-1-aza-5H-benzo[a]
phenothiazin-5-one 12
Reaction of 1-aza-5H-benzo[a] phenothiazin-5-one 10
(0.792 g, 3 mmol), 4-iodobenzoic acid (0.248 g, 1 mmol)
afforded as greenish yellow solid after recrystallized from
aqueous ethanol. Yield = 0.94 g (78%), mp > 250 °C (dec).
UV-visible (DMSO) λmax 367 (5.00), 433 (2.70). IR (νmax
,
cm−1): 1710 (COOH), 1634 (C=O), 1560, 1473, 1394,
1271, 1147, 1010, 922, 879, 746, 680, 623. 1H NMR
(DMSO-d6, 400 MHz,):
δ 11.35 (1 H, s, –COOH),
9.46–9.18 (2 H, m, Ar-H), 8.70–8.57 (2 H, m, Ar-H),
7.12–6.99 (3 H, m, Ar-H), 6.74–6.41 (4 H, m, Ar-H). 13C
NMR (DMSO6, 100 MHz): 188.5 (COOH), 171.9 (C = O),
150.8(C), 148.0 (C), 146.1 (C), 145.4 (C), 136.4 (C), 136.0
(2 C), 132.0 (2 C), 131.9 (CH), 131.7 (CH), 128.3 (CH),
125.8 (CH), 124.1 (CH), 123.6 (CH), 120.3 (CH), 118.4
(CH), 117.0 (CH), 115.9 (CH), 112.7 (CH). (Anal. Calcd.
for C22H12N2SO3: C, 68.74; H, 3.15. Found: C, 68.78; H,
3.10).
10-Amino-8-chloro-6(4-carboxyphenyl)-1,9,11-
triaza-5H-benzo[a] phenothiazin-5-one 8
The reaction of 10-amino-8-chloro-1,9,11-triaza-5-benzo[a]
phenothiazin-5-one,
6
(0.947 g,
3 mmol)
with
4–iodobenzoic acid (0.248 g, 1 mmol) afforded compound 8
as yellowish brown powdered solid after recrystallized from
aqueous ethanol. Yield = 1.1 g (85%), mp > 250 °C (dec).
UV-visible (DMSO) λmax 343 (4.92); 378 (4.00); 556
(1.09), 661(1.08); 744 (1.18); 794 (1.22). IR (νmax, cm−1):
3430 (–NH2), 1740 (COOH), 1660 (C=O), 1570, 1404,
6–(4–Nitrophenyl)–1-aza-5H–benzo[a]–phenoxazin-
5-one 14
1
1334, 1260, 1178, 1053, 923, 881, 767, 678. H NMR
(DMSO-d6, 400 MHz,): δ 12.1 (1 H, s, br, -COOH),
9.45–9.43 (1 H, m, Ar-H), 9.33–9.20 (1 H, m, Ar-H),
9.07–8.57 (5 H, m, Ar-H), 3.50 (2 H, br, -NH2). 13C NMR
(DMSO6, 100 MHz): 190.4 (COOH), 177.5 (C=O), 154.5
(C), 152.1 (C), 150.6 (C), 147.7 (C), 145.5 (C), 141.3 (C),
140.9 (C), 136.6 (C), 132.0 (C), 130.8 (C), 128.32 (CH),
125.4 (CH), 123.7 (CH), 120.9 (CH), 119.2 (CH), 115.8
(CH), 112.1 (CH). (Anal.calcd. for C20H10N4SO3Cl: C,
54.99; H, 2.54. Found: C, 55.39; H, 2.94).
Compound 14 was obtained by reaction of 1-aza-5H-benzo
[a]phenoxazin-5-one
13
(0.744 g,
3 mmol)
and
4–iodonitrobenzene (0.248 g, 1 mmol) to afford greenish
solid after recrystallization from aqueous ethanol. Yield =
0.92 g (70%), mp 216–218 °C (dec). UV-visible (DMSO)
λmax 366(4.00), 540 (3.15). IR (νmax, cm−1): 1644 (C=O),
1593, 1504, 1400, 1330, 1259, 1165, 1010, 846, 746, 682,
1
& 624. H NMR (DMSO-d6, 400 MHz,): δ 8.12 8.06 (4 H,
m, Ar-H), 7.99–7.93 (4 H, m, Ar-H), 7.48–7.47 (1 H, m, Ar-
H), 6.71–6.59 (2 H, m, Ar-H). 13C NMR (DMSO6, 100
MHz): 177.0 (C=O), 150.5(C), 145.3 (C), 144.1 (C), 142.3
(C), 140.5 (C), 139.9 (C), 136.7 (C), 135.4 (C), 132.9 (C),
133.7 (CH), 130.1 (CH), 129.6 (CH), 127.0 (CH), 126.9
(CH), 124.3 (CH), 122.5 (CH), 120.8 (CH), 119.7(CH),
113.1 (CH). (Anal. calcd. for C21H11N3O4: C, 68.29;
H,3.00. Found: C,67.89; H, 2.60).
6-(4-Nitrophenyl)-1-aza-5H-benzo[a]phenothiazin-5-
one, 11
Compound 11 was obtained by reaction of 1-aza-5H-benzo
[a]phenothiazin-5-one 10, (0.792 g, 3 mmol) with 4-
iodonitrobenzene (0.248 g, 1 mmol) as a orange solid.
Yield = 1.06 g (86%), mp > 250–252 °C. (dec). UV-visible
(DMSO) λmax 290 (4.00); 366 (3.00); 511(1.38). IR (νmax
,
6-(4-Carboxyphenyl)-1-aza-5H-benzo[a]phenoxazin-
5-one 15
cm−1): 3430 (–NH2), 1626 (C=O), 1589, 1479, 1313, 1267,
1
1192, 1097, 977, 844, 750, 669, 621. H NMR (DMSO-d6,
400 MHz,): δ 8.13–7.93 (4 H, m, Ar-H), 7.43–7.40 (4 H, m,
Ar-H), 7.35–7.12 (3 H, m, Ar-H). 13C NMR (DMSO6, 100
MHz): 181.0 (C=O), 156.3 (C), 151.4 (C), 148.0 (C), 146.9
(C), 145.2 (C), 143.9 (C), 140.5 (C), 137.6 (C), 136.1 (C),
132.7 (C), 123.8 (CH), 127.0 (CH), 126.1 (CH), 129.4
The reaction of 1-aza-5H-benzo[a]phenoxazin-5-one 13
(0.744 g, 3 mmol) with 4–iodo–benzoic acid (0.248 g, 1
mmol) gave compound 15 as dark brown solid. Yield =
0.95 g (82%), mp 240–242 °C (dec). UV-visible (λmax
,
DMSO-d6): 332(1.48), 339(1.49), 442(2.50). IR (νmax, cm

Medicinal Chemistry Research
−1): 1755 (COOH), 1667 (C = O), 1589, 1554, 1494, 1392,
1271, 1174, 1143, 1006, 923, 877, 829, 752, 682, 621. H
examined for inhibition zones and the observed zones were
measured and recorded in millimeters.
1
NMR (DMSO-d6, 400 MHz,): δ 12.41 (1 H, s, COOH),
9.50–9.21 (3 H, m, Ar-H), 8.70–8.32 (4 H, m Ar-H),
7.77–7.49 (4 H, m, Ar-H). 13C NMR (DMSO6, 100 MHz):
191(COOH), 168.7 (C=O), 154.6 (C), 152.2 (C), 150.4 (C),
149.5 (C), 148.3 (C), 145.5 (C), 144.5 (C), 140.6 (C), 138.2
(CH), 137.7(CH), 136.3(CH), 134.5 (CH), 132.9(CH),
131.9(CH), 130.4(CH), 128.8(CH), 126.6(CH), 125.1(CH),
120.7(CH), 115.3(CH). Anal. calcd. for C22H12N2O4:
C,71.74;H,3.28; N,7.60. Found: C, 71.34; H, 3.20; N, 7.21.
Determination of minimum inhibitory concentration
(MIC)
The minimum inhibitory concentrations (MIC) were deter-
mined using the agar dilution method. A set of six capped
small test tubes were used for each synthesized sample
against each organism. Nutrient agar solution was prepared
according to the level of turbidity of the solution in the test
tubes. The test tube containing the solution of lowest con-
centration of the sample that produced a clear solution was
taken and recorded as the MIC of the synthesized sample.
The screening effect of the synthesized compounds was
compared with the standard drugs, ciprofloxacin and
ketoconazole.
Antimicrobial screening test of the synthesized
angular phenothiazines and phenoxazines
Antimicrobial analyses were carried out using the agar-well
diffusion method Barry and Theornsberry 1985; Bauer et al.
1966). Fresh and pure clinical isolates of Bacillus subtillis,
staphylococcus aureus, Enterococcus faecalis, Pseudo-
mones aeruginosa, Escherichia coli, Candida albicans and
Aspergilus niger from Faculty of Pharmaceustical Sciences,
Univeristy of Nigeria, Nsukka were used for the tests. Stock
solutions of the respectives synthesized compounds were
prepared by initially dissolving 0.04 g in 2 ml of dimethyl
sulphoxide, DMSO, to obtain stock solutions of con-
centration 20 mg/ml each. From the stock solution, con-
centration of 10, 5, 2.5, 1.25, 0.125 and 0.3125 mg/ml were
prepared by serial dilution. Inoculation of the prepared agar
plates with the organism was done using a wire loop to
transfer a strand of the organism into the plate followed by
cross-streaking with the same wire loop to achieve uniform
spread on the plate. The bores (8 mm in diameter) were
aseptically made into the plates using sterilized cork borer.
A synthesized compound of known concentration was
introduced into the well using a sterilized syringe. The
plates were incubated at 37 °C for bacteria and 25 °C for
fungi for 24 h. At the expiration of the time the plates were
Results and discussion
Synthesis and characterization of the compounds
The syntheses of the novel derivatives of angular azaphe-
nothiazines and angular azaphenoxazines started with
synthesis of key intermediates. 4-Choro-2,6-diaminopyr-
imidine-5-thiol 3 with melting point 201–203 °C (Lit mp
202–203 °C) (Okafor 1975), was synthesized by three-step
thionation of 4-chloro-2,6-diaminopyrimidine 2 (Scheme 1)
(Okafor 1973).
The second intermediate, 7-chloroquinoline-5,8-dione 5,
with melting point 172–174 °C (Lit. mp 173.5–174.5 °C),
was prepared by multistep conversion of 8-
hydroxylnapthoquinoline 4 (Petrow and Sturgeon 1954)
(Scheme 2).
The cross-coupling of equimolar amounts of 4-chloro-2,
6-diaminopyrimidine-5-thiol 3 with 7-chloroquinoline-5,
Cl
O
O
N
Scheme 3 Synthesis of 10-
amino-8-chloro-1,9,11-triaza-5H
benzo[a]phenothiazine-5-one 6
Na2CO3
SH
N
H2N
N
N
+
N
C6H6/DMF
70 - 75oC
H2N
N
NH2
Cl
N
O
S
, 6h
Cl
3
6
5
N
Scheme 4 The synthesis of 4-
nitro-phenyl and 4-
carboxylphenyl derivatives of
compound 6
N
N
H N
N
N
S
I
COOH
I
NO2
2
H2N
N
N
S
H2N
N
N
N
N
O
N
, 4h
O
, 4h
dppb/PdCl2
dppb/PdCl2
O
S
Cl
Cl
Cl
6
8
7
COOH
NO2

Medicinal Chemistry Research
O
O
N
Scheme 5 Synthesis of 1-aza-
5H-benzo[a]phenothiazin-5-one
10
Na2CO3
SH
N
+
C6H6/DMF
reflux
NH2
Cl
N
O
S
9
10
5
N
Scheme 6 The synthesis of 4-
nitro-phenyl and 4-
carboxylphenyl derivatives of
compound 10
N
N
S
N
N
I
COOH
I
NO2
N
S
O
O
S
PdCl2/dppb
PdCl2/dppb
O
12
11
10
COOH
N
NO2
N
Scheme 7 The synthesis of 4-
nitro-phenyl and 4-
carboxylphenyl derivatives of
compound 13
N
N
N
I
COOH
I
NO2
N
O
O
O
O
PdCl2/dppb
O
PdCl2/dppb
O
14
15
13
COOH
NO2
8-dione 5 under anhydrous base catalyzed reaction afforded
a brownish solid compound, identified as 10-amino-8-
chloroquinoline-5,8-dione supplied a yellow powder solid
identified as 7-oxa-1,12-diazabenzo[a]anthracene-5-one 10,
melting point at 210–212 °C (Scheme 5).
chloro-1,9,11-triaza-5H-benzo[a]phenothiazine-5-one
6
melted at 130–132 °C (Scheme 3). The IR showed char-
acteristic absorptions at 3380, 1653 cm−1 which were
assigned to –NH2 and C=O, respectively. The multiplet
signals at δ 9.48–7.34 were assigned to aromatic protons.
The palladium(0)/piperazine catalyzed cross-coupling of
10-amino-8-chloro-7-thia-1,9,11,12-tetrabenzo[a]anthra-
cen-5-one 6 with 1-iodo-4-nitrobenzoic acid and 4-
iodobenzoic acid afforded new 10-amino-8-chloro-6-(4-
nitrophenyl)-1,9,11-triaza-5H-benzo[a]phenothiazin-5-one
The structure of compound 10 is in agreement with spectral
and elemental analytical data. Besides the aromatic protons
that integrated for eight protons as multiplet at δ 9.48–7.34 the
IR displayed absorption for C=O at 1648 cm−1.
The Mizoroki-Heck cross coupling of compound 10 with
1-iodo-4-nitrobenzoic acid and 4-iodobenzoic acid afforded
brownish coloured solids identified as compounds 6-(4-
nitrophenyl)-1-aza-5H-benzo[a]phenothiazin-5-one 11 and
6-(4-carboxyphenyl)-1-aza-5H-benzo[a]phenothiazin-5-one
12, respectively in excellent yields after recrystallization
from ethanol (Scheme 6).
7
and 10-amino-8-chloro-6-(4-carboxyphenyl)-1,9,11-
triaza-5H-benzo[a]phenothiazin-5-one 8, respectively
(Scheme 4) in high yields. In this reaction, a 1:1 mixture of
1,4-bis(diphenylphosphino)butane-palladium(II)chloride
precatalyst and a synthesized 1,4-bis(2-hydroxy-3,5-di-
tertbutylbenzyl)piperazine ligand (Mohanty et al. 2008)
were used in combination with K2CO3 under refluxing
methanol provided the optimal yields of the products. The
UV-visible spectra of compounds 7 and 8 exhibited bath-
ochromic shifts due to extended conjugation. The char-
acteristic absortions for –NH2 and C=O functional groups
were found at 3350 and 1645, respectively for compound 7
The spectral data are in agreement with the assign
structures and molecular formula of the synthesized com-
pounds. Compounds 11 and 12 showed red shifts in the
1
UV-visible spectra due to extended conjugations. The H
NMR –COOH absorption appeared offset in compound 12.
The IR bands at 1710, 1634 for compound 12 were assigned
to –COOH and C=O, respectively.
Similar procedure used in synthesis of compounds 6 and
10 was employed to prepare compound 13. The coupling of
equimolar amount of aminothiophenol with 7-chlor-
oquinoline-5,8-dione supplied 7-oxa-1,12-diazabenzo[a]
athracen-5-one 13, which was converted into 6-(4-nitrope-
nyl)-1-aza-5H-benzo[a]phenoxazin-5-one 14 and 6-(4-car-
boxyl)-1-aza-5H-benzo[a]phenoxazin-5-one 15 via Heck
reactions. Like compounds 6 and 10, compound 13 IR band
for carbonyl (Scheme 7) 1656 cm−1 (C = O) is lower than
1
and at 3430 and 1660 for compound 8. The H NMR sig-
nals for the carboxyl proton in compound 8 appeared offset
as a weak peak at 2.10 ppm. Other spectral and analytical
data were in agreement with the synthesized compounds.
In a similar procedure to the above, the reaction of
equimolar mixture of 2-aminothiophenol and 7-

Medicinal Chemistry Research
that expected for 7-chloroquinoline-5,8-dione (1690 cm−1
)
high number of HBA and this is reflected in the lipophilicity
values. This account for the reason compounds (10, 11, 13
and 14) with zero HBD value had relatively good lipophi-
licity (at the range of 3.029–4.735). Overall, drug-likeness
check revealed all the compounds will be well exposed in
systemic circulation when administered orally.
probably due to resonant effects in which the ionic reso-
nance form contribute appreciably to the ground state
(Agrawal and Mital 1975). The same lowering of carbonyl
IR frequencies were observes in all the synthesized deri-
vatives. The molecular structures of the synthesized com-
pounds were established by NMR, IR, UV and elemental
analysis data.
Antibacteria testing
How drug-like are the compounds
Docking calculations
Several cases are widely reported in literature where com-
pounds with interesting pharmacological activities failed to
make it to the shelves of market due to poor/bad pharma-
cokinetic properties (Ibezim et al. 2015). This has resulted
to investigation of these properties early in drug develop-
ment process. One of the most investigated of these prop-
erties is oral bioavailability profile. This is of particular
importance because most drugs are formulated in oral
administrable forms (Ibezim et al. 2017). Oral bioavail-
ability, also known as drug-likeness, of molecules is
determined computationally through calculation of certain
molecular descriptors and comparing the results to a set of
criteria outlined by Lipinski. Lipinski proposed rules, now
generally referred to as 'Lipinski’s rule of five' (ro5) which
qualifies a compound to be drug-like if it possesses the
following properties; MW less than five, lipophilicity (log
P) less than five, number of HBA/HBD groups less than ten/
five and NRB less than five (Lipinski et al. 1997).
All the compounds have physical-chemical parameters
which fall within the acceptable ranges for a drug-like
molecule according Lipinski ro5 (Table 1). However, the
MW of compound 7 and 8 seems quite high (436.839 and
435.851 Da respectively) and could be of concern if more
functional groups or moieties will be added during chemical
structural modification process for activity optimization.
Also, it was observed that the low HBD property of the
compounds was greatly compensated by their relatively
Rising cases of drug resistant bacteria (DRB)is now an
obvious global public health threat which has got the
attention of World Health Organization and Infectious
Disease Society of America (Worthington and Melander
2013). In the United States alone, a review performed by
Klein and Coworkers revealed that, more people died from
methicillin-resistant Staphylococcus aureus infections than
those who died of HIV/AIDS, emphysema, Parkinson’s
disease and homicide combined (Klein et al. 2007). Scien-
tists in both academia and industry seem to agree that one of
the effective ways of designing new molecules to combat
DRB is through tackling unconventional targets (Smitha
Rao et al. 2014 one of which is Type I signal peptidase
(Type I SPase).Type I SPase is an essential validated anti-
bacterial drug target which plays key roles in bacterial
secretory pathway through catalyzing the cleavage of
amino-terminal signal peptide from the translocated pre-
protein and to release exported proteins from the membrane
to reach their correct cellular or extra-cellular destination
(Tuteja 2005).
The best docked conformations of the test compounds
representing the lowest theoretical binding free energy of
the ligand–target complexes showed that all the compounds
interacted favourably with Type I SPase (Table 2). All
inhibited the activity of the enzyme at micromolar con-
centration except 6 which did at millimolar range.
Throughout, it appears attachment of benzoic acid and
nitrobenzene to either parent scaffolds afforded increased
interaction and therefore stronger binding with Type I
SPase. Although none of the new compounds bound to
Type 1 SPase stronger than its cocrystallized substrate
(maltose; −9.30 kcal/mol), their physicochemical proper-
ties, low inhibitory concentration and good ligand efficiency
present them as promising/potential antibacterial agents.
Therefore, it was necessary to confirm their predicted
potencies experimentally through whole bacteria assay.
Table 1 Physicochemical properties for evaluating drug-likeness
according to ro5
Compounds
NRB
MW
HBA
HBD
logP (o/w)
6
0
2
2
0
2
2
0
2
2
315.744
436.839
435.851
298.753
419.848
418.860
282.686
403.781
402.793
6
9
8
3
6
5
4
7
6
1
1
2
0
0
1
0
0
1
1.158
2.864
2.606
3.029
4.735
4.477
2.393
4.099
3.841
7
8
10
11
12
13
14
15
Experimental screening of the compounds against
selected bacteria
In vitro antibacterial screening results of the compounds are
shown in Table 3. It was observed that the compounds

Medicinal Chemistry Research
demonstrated varying activity against the studied organ-
isms. B. cereus was most susceptible to the compounds
being inhibited at a concentration as low as 0.25 µg/mL by
compound 6. Only compound 10 showed activity against E.
coli at a measurable MIC level (1.38 µg/mL). In compar-
ison, the compounds were more active against the tested
Gram positive bacteria (B. cereus and S. aureus) than the
tested Gram negative bacteria (P.aeruginosaand E. coli).
Many research findings attribute the relatively poor anti-
biotic effect of compound candidates against Gram negative
bacteria to the presence of lipopolysaccharide (LPS) layer
enveloping their cell wall. However, except for compound 6
which might get stocked in the LPS layer due to its very low
lipophilicity (log P = 1.158), the lipophilicity of the rest of
the compounds apparently guarantees easy passage of this
layer to reach intercellular targets. The discrepancies
between docking predictions and experimental assay is
more obvious in the performance of compound 12 where
docking scores it high whereas it performed very poorly
experimentally. The reason for differing results between
these two techniques is due to the fact that the in vitro
screening was not carried out directly on the enzyme used in
docking. Notwithstanding, the in silico result suggests that
in a situation where antibiotic resistant occurs, these com-
pounds would possibly thrive because they have different
target enzyme (Type I SPase) other than the ones known to
contribute/participate in bacteria resistant.
In the overall, compounds 7, 10, 11 and 15 emerged as
the best potent against the studied bacteria because of
inhibition of both Gram positive and negative bacteria at
low MIC some even lower than the used approved antibiotic
drug (ciprofloxacin).
Binding mode prediction of 7, 10, 11 and 15
Understanding the intermolecular interaction existing
between a putative drug molecule and a protein target goes
a long way to facilitate activity optimization of that mole-
cule. In that light, analysis of the binding conformation of
selected better performing compounds were studied as
shown in Fig. 1a–d.
Interactions of the compounds with Type I SPase were
grossly characterized with π-π contacts with aromatic rings
of Tyr161, Tyr216, Trp68 and Trp236 residues and
hydrogen bond relationships with Arg72 and Arg350 NH
groups. Compound 7 gained stronger binding with the
protein by forming extra lipophilic and hydrogen bonding
with the Trp346 aromatic ring and Asp20 carboxylate
moiety through its nitrophenyl aromatic ring and amino
group respectively. It appears the pose of compound 10
enabled it to make more hydrogen contact with Lys21 and
Ala69 NH through its chlorine atom. Both compounds 7
Table 2 Dock scores of the newly synthesized compoundstoward
Type I SPase binding site
Compounds Inhibition
constant
Binding free
energy
Ligand
efficiency
6
3.23
−3.40
−4.51
−4.67
−5.75
−7.03
−7.78
−6.08
−6.54
−6.57
0.16
0.15
0.16
0.29
0.24
0.27
0.3
7
492.55
378.48
60.99
7.08
8
10
11
12
13
14
15
1.97
35.13
16.15
15.24
0.23
0.23
Inhibition constant (Ki) values are in µM except for compound 6
which is in mM concentration
Table 3 Results of inhibition
zone diameter (IZD) in
Compounds
bacteria
millimetre (mm) of synthesized
compounds (20 mg/mL)
Gram positive
B.c.
Gram negative
S.a.
P.a
E.c.
6
22 0.55 (0.25)
14 0.11 (0.63)
10 0.44 (1.00)
27 0.55 (0.44)
16 0.55 (0.50)
7
19 1.20 (0.80)
12 1.50 (1.00)
–
4
6
7
18 0.44 (0.40)
3
8
16 0.07 (0.89)
-
10
20 0.07 (0.42)
–
–
14 0.35 (1.38)
11
23 1.44 (0.40)
–
12
–
7
–
13
–
17 0.22 (0.63)
12 0.55 (0.91)
22 0.40 (0.45)
25 0.22 (0.50)
4
5
14
15 0.25 (0.96)
28 0.10 (0.40)
31 0.08 (0.55)
5
4
15
5
4
Ciprofloxacin
25 0.10 (0.44)
22 0.22 (0.63)
Minimum inhibitory concentration (MIC) in µg/mL in bracket. B. c (Bacillus cereus); S. a (Staphylococcus
aureus); E. c (Escherichia coli); P. s (Pseudomonas aeruginosa); Conc. = μg/mL of DMSO

Medicinal Chemistry Research
Fig. 1 Dock pose of compounds
7 (a), 10 (b), 11 (c) and 15 (d)
toward Type I SPase binding
site. The atoms are in their
standard colours for both protein
residues and ligands, i.e.,
oxygen atom in red, chlorine
atom in green, hydrogen atom in
white, sulphur atom in yellow
etc. Polar contacts are shown in
dash lines
and 11 made intercourse with same residues with the
exception of that initiated by the amino group in 7. The best
dock pose of compound 15 gave a pose which made con-
tact, although with the same protein residues as the rest, but
using different part of the compound. Unlike the hetero
nitrogen atom of either phenothiazine or phenoxazine that
participated in hydrogen bonding, there was no observable
contribution made by sulphur and oxygen atoms in the
interaction of the compounds with the test protein. More-
over, it appears that the four candidates hydrogen bond with
lysine (Lys21 for 7, 10 and 11 and Lys48 for 15), an
essential residue for signal peptidase 1 activity (Dalbey
2013). Also, the compounds (7, 10, 11 and 15) made con-
tact with Type I SPase through the same residues as did the
maltose which suggests they might be acting as activators
rather than inhibitors. However, their activity in vitro indi-
cated that they are inhibitors (Ting et al. 2016).
one and 1-aza-5H-benzo[a]phenonoxazin-5-one and their
derivatives were particularly susceptible to Bacillus subtilis
and Staphylloccccus aureus. Further research work is in
progress to optimize the reaction conditions and develop a
better catalytic system to explore related cross-coupling
protocols so as to diversify target molecular motifs. In
addition, results of computational and biological screening
respectively showed that the new compounds interacted
favourably with an unconventional validated antibiotic
enzyme targeted to treat drug resistant bacteria—Type I
SPase and inhibited the activities of both Gram positive and
Gram negative bacteria with some having lower MICs than
the used approved drug—ciprofloxacin.
Compliance with ethical standards
Conflict of interest The authors declare that they have no conflict of
interest.
Conclusion
References
We have reported convenient protocol for functionalisation
of heterocyclic phenothiazine as well as phenoxazine in
moderate to high yields to afford highly coloured deriva-
tives at low temperatures and short time via Mizoroki–Heck
reaction system. The reaction conditions tolerated both
carbonyl functional groups and unprotected –NH2 moieties
in substrates. The 10-amino-8-chloro-1,9,11-triaza-5H-
benzo[a]phenothiazine, 1-aza-5H-benzo[a]phenothiazin-5-
Agrawal NL, Mital RL (1975) Synthesis of benzo[a][1, 4]benzothia-
zino[3,2-c]phenothiazines and 5H-benzo[a]phenothiazin-5-ones.
J Chem Eng Data 20(2):201
Barness EC, Bezerra-Gomez P, Nett M, Dandamycin HC, Chan-
drananimycin E (2015) benzoxazines from streptomycesgriseus. J
Antibiot 68:463
Barry AL, Thornsberry C (1985) Susceptibilty test, diffusion test
procedure. J Chem Path 19:492–500

Medicinal Chemistry Research
Bauer AW, Kirby WMM, Sherris JC, Track M (1966) Antibiotic
susceptibility testing standardized single disk method. Am J Clin
Path 45(4):493–496
Berman HM, Westbrook J, Feng Z, Gilliland G, Bhat TN, Weissig H,
Shindyalov IN, Bourne PE (2000) The protein data bank. Nucleic
Acids Res 28(1):10592235.
Dalbey RE (2013) Signal peptidase 1. In: Rawlings ND, Salvesen G
(eds) Handbook of proteolytic enzymes, vol 2, 3rd edn. Academic
Press, Washington, DC, pp 3493–3497
Ge JF, Arai C, Yang M, Md. AB, Lu J, Ismail NSM, Wittlin S, Kaiser
M, Brun R, Charman SA, Nguyen T, Morizzi J (2010) Discovery
of novel benzo[a]phenoxazine SSJ-183 as a drug candidate for
malaria. ACS Medicinal. Chem Lett 1:360–364
Okamoto T, Kozaki M, Doe M, Uchida M, Wang G, Okada K (2005)
1,4-benzoxazino[2,3-b]phenoxazine and its sulphur analogues:
synthesis, properties, and application to organic light emitting
diodes. Chem Mater 17(22):5504–5511
Onoabedje EA, Ibezim A, Okafor SN, Onoabedje US, Okoro UC
(2016) Oxazin-5-ones as a novel class of penicillin binding pro-
tein inhibitors: design, synthesis and structure activity relation-
ship PLoS ONE 11(10):e0163467
Onoabedje EA, Uchechukwu CO, Knight DW (2017) Rapid access to
new angular phenothiazine and phenoxazine dyes. J Heterocycl
Chem 54(1):206–214
Onoabedje EA, Okoro UC, Sarkar A, Knight DW (2016) Synthesis
and structure of new alkynyl derivatives of phenothiazine and
phenoxazine. J Sulfur Chem 34(3):269–281
Halgren TA (1996) Merck molecular forcefield. I. Basis, form, scope,
parameterization, and performance of MMFF94. J Comput
17:490–641
Petrow V, Sturgeon B (1954) Some quinoline-5,8-quinones. J Chem
Soc: 570–574
Ibezim A, Debnath B, Ntie-Kang F, Mbah CJ (2017) Binding of anti-
trypanosoma natural products from Africa flora against selected
drug targets: a docking study. Med Chem Res 26:562–579.
Ibezim A, Onyia K, Ntie-Kang F, Nwodo NJ (2015) Drug-like prop-
erties of potential anti-cancer compounds from Cameroonian
flora: a virtual study. J Appl Pharm Sci 5(06):133–137
Jaszczyszyn A, Gasiorowski K, Swiatek P, Malinka W, Cieslik-
Boczula K, Petrus J, Czarnik-Matusewicz B (2012) Chemical
structure of phenothiazines and their biological activity. Phar-
macol Report 64(1):16
Klein E, Smith DL, Laxminarayan R (2007) Hospitalizations and
death caused by methicillin-resistant Staphylococcus aureus,
United States, 1999-2005. Emerg Infect Dis 13(12):1840–1846
Lipinski CA, Lombardo F, Dominy BW, Feeney PJ (1997) Experi-
mental and computational approaches to estimate solubility and
permeability in drug discovery and development settings. Adv
Drug Deliv Rev 23:3–25
Pluta K, Jelen M, Morak-Mlodawska B, Zimecki M, Artym J, Kocieba
M (2010) Anticancer activity of newly synthessedazaphenothia-
zines from NCI’s anticancer screening bank. Pharmacol Rep
62:319–332
Pluta K, Morak-Miodawska B, Jelen M (2011) Recent progress in the
biological activities of synthesized phenothiazines. Eur J Med
Chem 46:3179–3189
Sailer M, Nonnenmacher M, Oeser T, Muller JJ (2006) Synthesis and
electronic properties of 3-acceptor-substituted and 3,7-bisaccep-
tor-substituted phenothiazines. Eur J Org Chem 423
Smitha Rao CV, Evelien De W, Anastassios E, Jozef A (2014)
Antibiotic targeting of the bacterial secretory pathway. Biochim
Biophys Acta 1843:1762–1783
Sridhar BT, Girish K, Channu BC, Thimmaiah KN, Kumara MN
(2015) Antibacterial activity of phenoxazine derivatives. J Chem
Pharm Res 7(4):1074–1075
Thimmaiah KN, Horton JK, Seshadri R, Isreal M, Houghton JA,
Harwood FC, Houghton PJ (1992) Synthesis and chemical
characterization of N-substituted phenoxazines directed towards
reversing vinca alkaloid in multidrug-resistant cancer cells. J Med
Chem 35:3358–3363
Moharty S, Suresh P, Balakrishna MS, Mague JT (2008) An inex-
pensive and highly stable ligand, 1,4-bis(2-hydroxy-3,5-di-tert-
butylbenzyl)piperizine for Mizoroki-Heck cross-coupling reac-
tions. Tetrahedron 64:240–247
Morris GM, Goodsell DS, Halliday RS, Huey R, Hart WE, Belew RK
et al. (1998) Automated docking using a lamarckian genetic
algorithm and an empirical binding free energy function. J Comp
Chem 19:1639–1662.
Ting YT, Harris PWR, Batot G, Brimble MA, Baker EN, Young PG
(2016) Peptide binding to a bacterial signal peptidase visualized
by peptide tethering and carrier-driven crystallization. Lucrj
3:10–19
Ntie-Kang F, Nwodo NJ, Ibezim A, Simoben CV, Karaman B, Ngwa
VF et al. (2014) Molecular modeling of potential anticancer
Turdean R, Bogdan E, Terec A, Petran A, Vlase L, Turcu I, Grosu I
(2009) Synthesis and structure of new 3,7,10-substituted-phe-
nothiazine derivatives. Cent Eur J Chem 7(1):111
agents from African medicinal plants.
J Chem Model
54:2433–2450.
Tuteja R (2005) Type I signal peptidase: a review. Arch Biochem
Biophys 441:107–111
Wainwright M, Amaral L, Kristiansen JE (2012) The evolution of
antimycobacterialagants from non-antibiotics Open J pharmacol
2:2–11
Wesolowska O, Molnar J, Westman G, Samuelsson K, Kawase M,
Ocsovszki I, Motohashi N, Michalak K (2006) Benzo[a]pheno-
zaxine: a new group of potent P-glycoprotein inhibitors. vivo
20:109
Worthington RJ, Melander C (2013) Combination approach to combat
multi-drug resistant bacteria. Trend Biotechnol 31(3):177–184
Zhu Y, Kulkarini AP, Jenekhe SA (2005) Phenoxazine based emission
donor-acceptor materials for efficient organic light emitting
diodes. Chem Mater 17(21):5225–5227
Okafor CO (1973) Studies in the heterocyclic studies VII: The use of
Kaufmann’s reaction as a route to o-aminomercaptopyridine. J
Org Chem 38:26–80
Okafor CO (1975) Studies in the heterocyclic series X: 1,3,9-triaza-
phenothiazine ring system. A new phenothiazine ring. J Org
Chem 40(19):2753–2755
Okafor CO (1977) Studies in the heterocyclic series XIII. New CNS-
depressants derived from 1,9-Diazaphenoxazine and two isomeric
triazaphenothiazine. Eur J Med Chem 12(3):249
Okafor CO (1986) The chemistry and applications of angular phe-
nothiazines derivatives. Dye Pigment 7:249–287
Okafor CO, Uche OI, Akpanisi ES (1981) A novel tetraaza-analog of
phenothiazine. J Heterocycl Chem 18:1589