Synthesis of novel phthalazine derivatives as pharmacological activities

Article abstract of DOI:10.1002/jhet.3735

Phthalazine derivatives attached to amino acid derivatives were synthesized with high yields. The reaction of phthalazine derivatives with different phthalyl and tosylamino acids such as glycine, alanine, phenylalanine, valine, serine, and threonine in the presence of N,N-dicyclo hexylcarbodiimide (DCC) as a dehydrating agent reagent yielded high yields of the afforded compounds. Phthalylamino acids derivatives were obtained by deprotection of phthalazine derivatives, with the latter heating with hydrazine hydrate. The chemical structures of all phthalazine derivatives were affirmed by elemental analysis and spectral data (IR, MS, ¹HNMR, and ¹³C NMR). Screening out and estimation of the synthesized derivatives for their cytotoxic and antioxidant activity were done, and most of them showed powerful activity in comparison with standard drugs.

Full text of DOI:10.1002/jhet.3735

Received: 21 April 2019
DOI: 10.1002/jhet.3735
Revised: 15 August 2019
Accepted: 25 August 2019
A R T I C L E
Synthesis of novel phthalazine derivatives as
pharmacological activities
Ashraf Farouq Wasfy | Aly A. Aly | Mohamed S. Behalo | Noura S. Mohamed
Chemistry Department, Faculty of
Abstract
Science, Benha University, P.O. Box
13518, Benha, Egypt
Phthalazine derivatives attached to amino acid derivatives were synthesized with
high yields. The reaction of phthalazine derivatives with different phthalyl and
tosylamino acids such as glycine, alanine, phenylalanine, valine, serine, and
threonine in the presence of N,N-dicyclo hexylcarbodiimide (DCC) as a
dehydrating agent reagent yielded high yields of the afforded compounds.
Phthalylamino acids derivatives were obtained by deprotection of phthalazine
derivatives, with the latter heating with hydrazine hydrate. The chemical struc-
tures of all phthalazine derivatives were affirmed by elemental analysis and spec-
Correspondence
Noura Sobhi Mohamed, Chemistry
Department, Faculty of Science, Benha
University, P.O. Box 13518, Benha, Egypt.
Email: nourasobhi@yahoo.com
Funding information
Benha University, Faculty of Science,
Chemistry Department
tral data (IR, MS, HNMR, and 13C NMR). Screening out and estimation of the
1
synthesized derivatives for their cytotoxic and antioxidant activity were done,
and most of them showed powerful activity in comparison with standard drugs.
According to these observations and in continuation of our
work on synthesis heterocycles of pharmacological
1 | INTRODUCTION
Among the different nitrogen containing heterocycles,
phthalazines play an important role in medicinal chemistry
and subsequently have emerged as a pharmacophore. The
phthalazine structural motifs have attracted a great deal of
interest because of their ready accessibility, diverse chemical
reactivity, and wide gamut of biological activities like
interest,[36–38] there was an interest in synthesizing new
molecules involving phthalazine and amino acid moieties
in a single molecular framework that possibly have anti-
tumor, antioxidant, and DNA agents.
antibacterial,[1]
antifungal,[2]
antitumor,[3–5]
anti-
2 | RESULTS AND DISCUSSION
inflammatory,[6–8] and anticonvulsant[9–11] activities. In
organic synthesis, N-protected amino acids are essential key
precursors and have been used in many areas, for example,
peptide synthesis,[12,13] medicinal chemistry,[14–16] as chiral
sources,[17–20] and polymer materials.[21,22] And in the
recent years, they got interest because of their magnificent
pharmacological and therapeutic properties,[23–25] and
phthalazines linked amino acid derivatives as a new class of
The starting molecule 2-[4-(4-oxophthalazin-1-yl)phe-
nyl]-1H-isoindole-1,3-(2H)-dione (2) synthesis was
achieved with high yield via fusion of phthalic anhydride
with 4-(4-aminophenyl)-phthalazin-1-(2H)-one (1) (pre-
pared by reacting γ-keto acids with hydrazine hydrate in
ethanol)[39–41] (Scheme 1). Phthalazinone 2 structure was
1
improved on the basis of its spectral data. H NMR spec-
promising
antimicrobial,[26–28]
antitumor,[19–23]
trum exhibited two signals (exchangeable) at δ 7.99 and
13.20 ppm corresponding to NH and OH protons that cer-
tify existence of lactam-lactim forms of phthalazinone 2.
Also, IR spectrum showed absorption bands at ύ 3208 to
3085 cm−1, 1746 to 1700 cm−1, and 1685 cm−1 due to
NH$OH, 3CO, respectively (Scheme 1).
anihypertensive,[24,26] antidiabetic,[27,28]anti-inflammatory,[29,30]
and vesarelaxant activities.[31] The integration of amino
acid residues in various oxygen-,nitrogen-, and sulfur-con-
taining heterocycles sometimes enhances the biological
profile manifold over that of its parent nucleus.[32–35]
J Heterocyclic Chem. 2019;1–14.
wileyonlinelibrary.com/journal/jhet
© 2019 Wiley Periodicals, Inc.
1

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WASFY ET AL.
SCHEME 1 Synthesis of phthalazinone derivative 2
Synthesis of 2-(4-(4-aminophthalazin-1-yl)phenyl)iso-
indoline-1,3-dione (4) and 2-(4-(4-mercaptophthalazin-
1-yl)phenyl)isoindoline-1,3-dione (5) required as starting
material was accomplished with a good yield via reaction
of phthalazinone derivative 2 with phosphorus pen-
tachloride and phosphorus oxychloride to yield
chlorophthalazine 3, which in turn reacts with ammonium
acetate and thiourea to give aminophthalazine 4 and
mercaptophthalazine 5, respectively. Aminophthalazine
as dehydrating agent to furnish the corresponding
phthalazine derivatives 7 to 9a-f (Scheme 3).
Deprotection of the amino group in 1-N-
(arylaminoacycloxy)-2-[4-(4-oxo-phthalazin-1-yl)phenyl]
isoindole-1,3-dione (7a-f), 1-N-(arylaminoacylamino)-2-[4
-(4-oxo-phthalazin-1-yl)phenyl]
isoindole-1,3-dione
(8a-f), and 1-N-(arylaminoacylthio)-2-[4-(4-oxo-3,4-dihy-
drophthalazin-1-yl) phenyl]isoindole-1,3-dione (9a-f) was
accomplished in good yield (70% to 75%) through their
refluxing with ethanolic solution of hydrazine hydrate for
2 hours to afford unprotected aminoacyl derivatives (10-
12)a-f, respectively (Scheme 3). Appearance of absorption
band of amino group in compounds (10-12)a-f and disap-
pearance of imidic group absorption confirm the
deprotection process.
1
derivative 4 was confirmed by spectral studies. H NMR
spectrum showed signal at δ 8.13 (s, 2H, NH2, exchange-
able with D2O), 8.06 to 7.33 ppm (m, 12H, aromatic pro-
tons). Also, IR spectrum displayed absorption bands at ύ
3469 to 3230 cm−1 and 1780 and 1735 cm−1 due to NH2
and CO imidic, respectively. And the structure of
mercaptophthalazine improved by IR spectrum (cm−1)
showed bands at 2590 (SH), 1788 to 1730 (CO imidic),
Similarly, phthalazines 2, 4, and 5 reacted with tosyl
derivatives of the same amino acids to furnish
phthalazine derivatives (14-16)a-f (Scheme 4).
1
1640 (C N), and 1330 (C S); also, H NMR spectrum
showed signal at δ 11.50 (s,1H,SH), 8.20 to 7.70 ppm (m,
12H, aromatic protons) (Scheme 2).
3 | MATERIALS AND METHODS
Phthalazines 2, 4, and 5 were used as reactive key pre-
cursors to originate a group of amino acids derivatives by
treating them with several N-protected amino acids
followed by estimating their antitumor, antioxidant, and
DNA activity.
Consequently, phthalazines (2, 4, and 5) reacted with
phthalyl derivatives of amino acids such as glycine, DL-ala-
nine, DL-phenylalanine, L-valine, L-serine, and L-threonine
We got all chemical reagents from Sigma-Aldrich. Sol-
vents were purchased from El-Nasr Chemicals in analyti-
cal grade. Also, we did thin-layer chromatography (TLC)
on all products by using silica gel polyester sheets
(Kieselgel 60 F254, 0.20 mm, Merck). All melting points
are in degree centigrade (uncorrected). We recorded IR
spectra (KBr) at the Microanalytical Center (Faculty of
SCHEME 2 Synthesis of phthalazines derivative (4,5)

WASFY ET AL.
3
(0.01 mol), and the solution was heated for 6 hours.[39–41]
After cooling, we filtered off the precipitate and rec-
rystallized dimethylformamide/water to produce the
afforded compound 1. Mp 250ꢀC-252ꢀC; yield 50%; IR
(cm−1) ν: 3295-3064 (NH2, NH$OH), 1685 (CO); Ms: m/
1
z 237 (Mꢁ+); H NMR (DMSO-d6) δ: 5.31 (s, 2H, NH2
exchangeable), 7.99 (s,1H, NH$OH exchangeable),
6.50-8.19 (m, 8H, Ar─H); Anal. Calcd. for C14H11N3O
(Mol.wt.237): C, 70.87; H, 4.67; N, 17.71; Found: C,
70.85; H, 4.69; N, 17.70%.
4.1.2 | 2-[4-(4-Oxo-3,4-dihydrophthalazin-
1-yl)-phenyl]-isoindole-1,3-dione(2)
Fusion of a mixture of compound 1 (0.01 mol) with
phthalic anhydride (0.01 mol) at 250ꢀC for 2 hours; after
cooling, we added water to the residual product, and the
solid that was obtained was filtered off and crystallized
from DMF/H2O to give yellow crystals.[17,18] Mp 305ꢀC-
307ꢀC; yield 93%; IR (cm−1): 3400-3200 (NH$OH),
SCHEME 3 Reactions of phthalazines with phthaloyl amino
acids
1
1780-1730 (2CO), 1600 (C N); Ms: m/z 367 (Mꢁ+); H
NMR (DMSO-d6) δ: 8.17 (s, 1H, NH exchangeable),
7.69-8.19 (m, 12H, Ar─H); 13C NMR, 167.10, 165.02,
162.80, 159.40, 135.04, 134.71, 132.00, 131.10, 130.40,
129.45, 128.50, 127.51, 124.50, 123.70; Anal. Calcd. for
C22H13O3N3 (Mol.wt.367): C, 71.93; H, 3.57; N, 11.44;
Found: C, 71.83; H, 3.60; N 11.40%.
4.1.3 | Synthesis of 2-(4-(4-chlorophthala-
zin-1-yl)phenyl)isoindoline-1,3-dione (3)
SCHEME 4 Reaction of phthalazines with tosyl amino acids
A solution of phthalazinone 2 (0.01 mol) in phosphorus
oxychloride (3 mL) and phosphorus pentachloride
(0.01 mol) was heated for 4 hours in water bath. After
cooling, pour the solution accurately into ice. The prod-
uct was filtered, washed well with water, and crystallized
from ethanol to give 3. Yield, 80%; mp 130ꢀC-132ꢀC. IR
spectrum (KBr, ν, cm−1): 1780-1730 (CO imidic), 1662
Science; Mansoura University) on a Mattson 5000 FTIR
Spectrophotometer. The H NMR spectra were detected
1
on a Varian Spectrophotometer at 300 MHz, using tetra-
methylsilane (TMS) as an internal reference and DMSO-
d6 as solvent at the Microanalytical Center (Faculty of
Science, Ain Shams University).
Elemental analyses were performed on a CHN ana-
lyzer, and all compounds were within 0.4 of the theoret-
ical values. Pharmacological activities were carried in
Pharmacognosy Department, Faculty of Pharmacy, Man-
soura University, Mansoura, Egypt.
1
(C N), 835 (C─Cl); H NMR (CDCl3, δ, ppm): 8.31-7.03
(m, 12H, Ar─H); MS: m/z 385 (Mꢁ+), 386 (M+); Anal.
Calcd. for C22H12ClN3O2 (385.81): C, 68.49; H, 3.14; N,
10.89; Found: C, 68.40; H, 3.10; N, 10.82%.
Synthesis of 2-(4-(4-aminophthalazin-1-yl)phenyl)
isoindoline-1,3-dione (4)
4 | EXPERIMENTAL PROTOCOLS
4.1 | Chemistry
Fusion of an equimolar amount of chlorophthalazine 3 with
ammonium acetate (0.01 mol) in oil bath for 2 hours. After
cooling, add water to the formed solid, filter, dry, and crys-
tallize from ethanol. Yield, 80%; mp 230ꢀC-232ꢀC. IR spec-
trum (cm−1): 3469-3230 (NH2), 3056 (CH─ aromatic),
1780-1735 (CO imidic), 1704 (C N); 1H NMR (DMSO-d6, δ,
ppm): 8.36 (s, 2H, NH2, exchangeable), 8.30-7.88 (m, 12H,
4.1.1 | Synthesis of 4-(4-aminophenyl)
phthalazin-1-(2H)-one (1)
Ethanolic solution of hydrazine hydrate (0.01 mol) was
added to a solution of 2-(4-aminobenzoyl)-benzoic acid

4
WASFY ET AL.
Ar─H); 13C NMR, 167.10, 166.02, 165.00, 152.32, 143.01,
133.01, 132.11, 131.8, 129.40, 128.50, 127.20, 126.20, 123.70,
119.30, 116.40; Anal. Calcd. for C22H14N4O2 (366.38): C,
72.12; H, 3.85; N, 15.29; Found: C, 72.01; H, 3.75; N, 15.20%.
for C32H18N4O6 (Mol.wt.554): C, 69.31; H, 3.27; N, 10.10;
Found: C, 69.21; H, 3.20; N, 10.01%.
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-
1-yl 2-(1,3-dioxoisoindolin-2-yl)propanoate (7b)
Yield, 60%; mp 252ꢀC-254ꢀC; IR (cm−1): ʋmax: 1780-1734
(imidic CO), 1704 (C O of ester), 1591 (C N); Ms: m/z
568 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.42-7.97 (m, 16H,
Ar─H), 4.30 (q, 1H, J = 7.3, CH), 1.71 (d, 3H, J = 7.5,
CH3); 13C NMR, 172.73, 167.32, 165.33, 152.38, 143.34,
133.25, 132.69, 131.40, 129.40, 128.55, 127.90, 126.38,
123.42, 116.50, 56.50, 16.50; Anal. Calcd. for C33H20N4O6
(Mol.wt.568): C, 69.72; H, 3.55; N, 9.85; Found: C,
69.62; H, 3.50; N, 9.81%.
Synthesis of 2-(4-(4-mercaptophthalazin-1-yl)
phenyl)isoindoline-1,3-dione (5)
A solution of chlorophthalazine 3 (0.01 mol) in ethanol
(20 mL) and thiourea (0.01 mol) containing sodium
ethoxide was heated under reflux. After refluxing for
6 hours, pour the reaction mixture into ice cold water
and then add acetic acid to acidify the product. The
formed product was collected by filtration then dried and
crystallized from ethanol to produce yellow crystals.
Yield, 60%; mp 200ꢀC-201ꢀC. IR spectrum (cm−1): 3460
(NH), 2590 (SH), 1788-1730 (CO imidic), 1640 (C N),
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-1-yl
2-(1,3-dioxoisoindolin-2-yl)-3-phenylpropanoate (7c)
Yield, 60%; mp 200ꢀC-202ꢀC; IR (cm−1): ʋmax: 1778-1730
(imidic CO), 1715 (C O of ester), 1607 (C N); Ms: m/z
630 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.43-7.97 (m, 20H,
Ar─H), 4.47(t, 1H, J = 7.2, CH), 3.33 (d, 2H, J = 6.8,
CH2); Anal. Calcd. for C39H24N4O6 (Mol.wt.644): C,
72.66; H, 3.75; N, 8.69; Found: C, 72.66; H,
3.65; N, 8.60%.
1
1330 (C S); H NMR (DMSO-d6, δ, ppm): 11.50 (s, 1H,
NH$SH, exchangeable), 8.20-7.70 (m, 12H, Ar─H); 13C
NMR, 172.73, 167.32, 165.33, 152.38, 143.34, 133.25,
132.69, 131.40, 129.40, 128.55, 127.90, 126.38, 123.42,
116.50, 56.50, 16.5; MS: m/z 383 (M+); Anal. Calcd. for
C22H13N3O2S (383.43): C, 68.92; H, 3.42; N, 10.96;
Found: C, 68.80; H, 3.32; N, 10.85%.
4.1.4 | General procedure for reaction of
(2, 4, 5) with phthalylamino acids:
Formation of (7-9)a-f
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-1-yl
2-(1,3-dioxoisoindolin-2-yl)-3-methylbutanoate (7d)
Yield, 65%; mp 210ꢀC-212ꢀC; IR (cm−1): ʋmax: 1780-1727
(imidic CO), 1710 (C O of ester), 1591 (C N); Ms: m/z
596 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.27-8.27 (m, 16H,
A well-stirred solution of phthalazine derivative 2, amino
phthalazine 4, and mercaptophthalazine 5 (0.01 mol) in
dry tetrahydrofurane (30 mL) and N-phthalayl amino
acids, namely, glycine, DL-alanine, L-phenylalanine, DL-
valine, L-serine, and threonine (0.01 mol), and
dicyclohexylcarbodiimide (0.01 mol) were added at 0ꢀC.
for 24 hours at 0ꢀC, the reaction mixture was stirred, and
stirring was continued at room temperature for another
24 hours. The residual N,N-dicyclohexylurea was filtrated
from the solution. The filterate was evaporated in vacuo,
and the formed product dissolved in ethylacetate (25 mL),
and the solution was filtrated again from the residual
N,N-dicyclohexylurea. The products were attained by
evaporation of filtrate in vacuo and recrystallized from
proper solvent to give compounds (7-9)a-f.
Ar─H), 4.32 (m, 1H, CH), 1.09 (d, 6H, J = 6.8, 2CH3); 13
C
NMR, 168.10, 167.19, 166.00, 152.40, 150.04, 133.11,
132.00, 129.45, 128.50, 127.51, 126.70, 123.70, 66.4, 27.8,
18.9; Anal. Calcd. for C35H24N4O6 (Mol.wt.596): C,
70.46; H, 4.05; N, 9.39; Found: C, 70.36; H, 4.00; N, 9.28%.
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-
1-yl 2-(1,3-dioxoisoindolin-2-yl)-
3-hydroxypropanoate (7e)
Yield, 70%; mp 230ꢀC-232ꢀC; IR (cm−1): ʋmax: 3468 (OH),
1780-1733 (imidic CO), 1700 (C O of ester), 1607 (C N);
Ms: m/z 584 (Mˑ+); 1H NMR (DMSO-d6) δ: 5.55 (s, 1H, OH,
exchangeable with D2O), 7.43-7.95 (m, 16H, Ar─H), 4.43 (q,
1H, J = 7.1, CH), 4.10 (d, 2H, J = 7.5, CH); 13C NMR,
197.60, 193.01, 165.33, 167.10, 165.00, 152.05, 132.01, 129.40,
128.00, 127.70, 126.60, 133.10, 123.70, 72.90, 59.30; Anal.
Calcd. for C33H20N4O7 (Mol.wt.584): C, 67.81; H, 3.45; N,
9.58; Found: C, 67.71; H, 3.40; N, 9.50%.
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-
1-yl 2-(1,3-dioxoisoindolin-2-yl)acetate (7a)
Yield, 70%; mp 265ꢀC-267ꢀC; IR (cm−1): ʋmax: 1781-1733
(imidic CO), 1710 (C O ester), 1591 (C N); Ms: m/z
554 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.43-8.57 (m, 16H,
Ar─H), 4.40 (s, 2H, CH2); 13C NMR, 171.20, 168.10,
167.02, 166.00, 165.12, 152.00, 133.51, 132.01, 129.40,
128.50, 127.50, 126.70,123.70, 119.71, 43.11; Anal. Calcd.
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-1-yl
2-(1,3-dioxoisoindolin-2-yl)-3-hydroxybutanoate (7f)
Yield, 65%; mp 253ꢀC-255ꢀC; IR (cm−1): ʋmax: 3377 (OH),
1775-1735 (imidic CO), 1701 (C O of ester), 1608 (C N);

WASFY ET AL.
5
1
1
Ms: m/z 598 (Mˑ+); H NMR (DMSO-d6) δ: 5.59 (s, 1H,
OH), 7.42-7.97 (m, 16H, Ar─H), 4.95 (q, 1H, J = 6.8, CH),
4.30 (d, 1H, J = 7.0, CH), 1.22 (d, 3H, J = 6.8, CH3); Anal.
Calcd. for C34H22N4O7 (Mol.wt.598): C, 68.22; H, 3.70; N,
9.36; Found: C, 68.15; H, 3.60; N, 9.30%.
(C N); Ms: m/z 595 (Mˑ+); H NMR (DMSO-d6) δ: 9.23
(s,1H, NH, exchangeable), 7.40-7.87 (m, 16H, Ar─H), 4.50
(d, 1H, J = 7.8, CH), 2.87 (m, 1H, CH), 1.22 (d, 6H,
J = 6.8, 7.3, 2CH3); Anal. Calcd. for C35H25N5O5 (Mol.
wt.595): C, 70.58; H, 4.23; N, 11.76; Found: C, 70.50; H,
4.13; N, 11.64%.
2-(1,3-Dioxoisoindolin-2-yl)-N-(4-(4-(1,3-
dioxoisoindolin-2-yl)phenyl)phthalazin-1-yl)
acetamide (8a)
2-(1,3-Dioxoisoindolin-2-yl)-N-(4-(4-(1,3-
dioxoisoindolin-2-yl)phenyl)phthalazin-1-yl)-3-
hydroxypropanamide (8e)
Yield, 60%; mp 250ꢀC-252ꢀC; IR (cm−1): ʋmax: 3350-3250
(NH), 1780-1735 (imidic CO), 1701 (C O of amide), 1608
Yield, 70%; mp 201ꢀC-203ꢀC; IR (cm−1): ʋmax: 3420-3260
(NH, OH), 1780-1735 (imidic CO), 1701 (C O of amide),
1
(C N); Ms: m/z 553 (Mˑ+); H NMR (DMSO-d6) δ: 9.11
1
(s,1H, NH, exchangeable), 7.60-7.97 (m, 16H, Ar─H),
3.83 (s, 2H, CH2); 13C NMR, 168.50, 167.12, 166.00,
165.10, 152.00, 143.51, 133.1, 132.00, 131.81, 129.41,
128.50, 127.20, 126.90, 123.70, 119.71, 116.40, 47.17; Anal.
Calcd. for C32H19N5O5 (Mol.wt.553): C, 69.44; H, 3.46; N,
12.65; Found: C, 69.34; H, 3.40; N, 12.55%.
1608 (C N); Ms: m/z 583 (Mˑ+); H NMR (DMSO-d6) δ:
10.55 (s,1H, NH, exchangeable), 7.40-7.87 (m, 16H,
Ar─H), 4.95 (s, 1H, OH), 4.32 (t, 1H, J = 6.5, CH), 3.43
(d, 2H, J = 8.2, CH2); Anal. Calcd. for C33H21N5O6 (Mol.
wt.583): C, 67.92; H, 3.63; N, 12.00; Found: C, 67.82; H,
3.53; N, 11.95%.
2-(1,3-Dioxoisoindolin-2-yl)-N-(4-(4-(1,3-
dioxoisoindolin-2-yl)phenyl)phthalazin-1-yl)
propanamide (8b)
2-(1,3-Dioxoisoindolin-2-yl)-N-(4-(4-(1,3-
dioxoisoindolin-2-yl)phenyl)phthalazin-1-yl)-
3-hydroxybutanamide (8f)
Yield, 65%; mp 230ꢀC-232ꢀC; IR (cm−1): ʋmax: 3450-3200
(NH), 1780-1735 (imidic CO), 1701 (C O of amide), 1608
Yield, 70%; mp 201ꢀC-203ꢀC; IR (cm−1): ʋmax: 3420-3260
(NH, OH), 1780-1735 (imidic CO), 1701 (C O of amide),
1
1
(C N); Ms: m/z 567 (Mˑ+); H NMR (DMSO-d6) δ: 10.22
1608 (C N); Ms: m/z 597 (Mˑ+); H NMR (DMSO-d6) δ:
(s,1H, NH, exchangeable), 7.60-7.97 (m, 16H, Ar─H), 3.53
(q, 1H, J = 7.5, CH), 1.81(d, 3H, J = 7, CH3); 13C NMR,
172.73, 167.17, 166.20, 143.69, 132.20, 131.80, 129.40, 128.65,
127.70, 126.92, 123.71, 119.30, 116.40, 56.80, 16.50; Anal.
Calcd. for C33H21N5O5 (Mol.wt.567): C, 69.84; H, 3.73; N,
12.34; Found: C, 69.74; H, 3.63; N, 12.23%.
8.80 (s,1H, NH), 7.40-7.87 (m, 16H, Ar─H), 5.50 (s, 1H,
OH), 4.52 (d, 1H, J = 8.5, CH), 2.23 (q, 1H, J = 7.3, CH),
1.20 (d, 3H, J = 6.8, CH3); Anal. Calcd. for C34H23N5O6
(Mol.wt.597): C, 68.34; H, 3.88; N, 11.72; Found: C,
68.25; H, 3.80; N, 11.62%.
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 2-(1,3-dioxoisoindolin-2-yl)
ethanethioate (9a)
2-(1,3-Dioxoisoindolin-2-yl)-N-(4-(4-(1,3-
dioxoisoindolin-2-yl)phenyl)phthalazin-1-yl)-
3-phenylpropanamide (8c)
Yield, 60%; mp 230ꢀC-232ꢀC; IR (cm−1): ʋmax: 1760-1730
(imidic CO), 1720 (C O of ester), 1660 (C N); Ms: m/
Yield, 70%; mp 210ꢀC-212ꢀC; IR (cm−1): ʋmax: 3380-3220
(NH), 1780-1735 (imidic CO), 1701 (C O of amide), 1608
z
570 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.40-7.87
(C N); Ms: m/z 630 (Mˑ+); H NMR (DMSO-d6) δ: 8.23
(m, 16H, Ar─H), 4.52 (s, 2H, CH2); 13C NMR, 196.53,
193.02, 167.01, 165.02, 152.11,133.14, 132.05, 129.40,
128.50, 127.70, 126.60, 123.75, 57.40; Anal. Calcd. for
C32H19N4O5S (Mol.wt.570): C, 67.36; H, 3.18; N, 9.82;
Found: C, 67.30; H, 3.10; N, 9.73%.
1
(s,1H, NH, exchangeable), 7.40-7.87 (m, 16H, Ar─H),
4,40 (t, 1H, J = 6.5, CH), 3.43 (d, 2H, J = 8.1, CH2); 13C
NMR, 171.12, 170.00, 168.20, 167.11, 166.02, 165.45,
152.78, 150.44, 135.91, 133.43, 132.54, 131.85, 129.40,
128.50, 127.51, 126.72, 119.71, 62.84, 34.70; Anal. Calcd.
for C39H25N5O5 (Mol.wt.643): C, 72.78; H, 3.92; N, 10.88;
Found: C, 72.70; H, 3.81; N, 10.80%.
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 2-(1,3-dioxoisoindolin-2-yl)
propanethioate (9b)
2-(1,3-Dioxoisoindolin-2-yl)-N-(4-(4-(1,3-
dioxoisoindolin-2-yl)phenyl)phthalazin-1-yl)-
3-methylbutanamide (8d)
Yield, 65%; mp 210ꢀC-212ꢀC; IR (cm−1): ʋmax: 1770-1735
(imidic CO), 1710 (C O of ester), 1600 (C N); Ms: m/z
584 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.40-8.12 (m, 16H,
Ar─H), 4.47 (q, 1H, J = 6.3, CH), 1.18 (d, 3H, J = 6.8,
CH3); Anal. Calcd. for C33H20N4O5S (Mol.wt.584): C,
Yield, 75%; mp 240ꢀC-242ꢀC; IR (cm−1): ʋmax: 3400-3200
(NH), 1780-1735 (imidic CO), 1701 (C O of amide), 1608

6
WASFY ET AL.
67.80; H, 3.45; N, 9.58; Found: C, 67.71; H,
3.36; N, 9.48%.
4.1.5 | General procedure for reaction of
(7, 8, 9)a-f with hydrazine hydrate:
Formation of compounds (10-12)a-f
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 2-(1,3-dioxoisoindolin-2-yl)-
3-phenylpropanethioate (9c)
To a solution of compounds (7, 8, 9)a-f (0.01 mol) in abso-
lute ethanol (20 mL), hydrazine hydrate (0.01 mol) was
added and refluxed for 2 hours, then left for 24 hours at
room temperature. Evaporation of the solvent in vacuo
resulted in a solid material, which in return was added to
water (10 mL) and the solution acidified with acetic acid till
(pH = 6), then heated for 1 hour on steam bath; the suspen-
sion was diluted with water (15 mL) and filtrated off. The
filtrate was concentrated and cooled to produce the solid,
which crystallized from proper solvent to obtain (10-12)a-f.
Yield, 68%; mp 240ꢀC-245ꢀC; IR (cm−1): ʋmax: 1780-1730
(imidic CO), 1715 (C O of ester), 1610 (C N); Ms: m/z
646 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.45-8.22 (m, 16H,
Ar─H), 3.40 (t, 1H, J = 7.2,CH), 2.89(d, 2H, J = 7.49, CH2);
13C NMR, 197.62, 167.10, 152.08, 136.64, 133.61, 132.70,
129.40, 128.60, 127.74, 126.62, 125.90, 123.01, 72.04, 35.30;
Anal. Calcd. for C39H24N4O5S (Mol.wt.660): C, 70.90; H,
3.66; N, 8.48; Found: C, 70.80; H, 3.60; N, 8.44%.
4-(4-Aminophenyl)phthalazin-1-yl glycinate (10a)
Yield, 75%; mp 240ꢀC-242ꢀC; IR (cm−1): ʋmax: 3360-3210
(2NH2), 1730 (C O of ester), 1610 (C N). The vanishing
of bands corresponding to CO of cyclic imide at 1780,
1730, was highly informative and established the accom-
plishment of deprotection process; Ms: m/z 294 (Mˑ+); 1H
NMR (DMSO-d6) δ: 8.50 (s, 2H, NH2), 7.43-7.97 (m, 8H,
Ar─H), 4.53 (s, 2H, CH2); 13C NMR, 171.00, 168.11,
150.00, 145.60, 133.51, 131.81, 128.30, 127.40, 126.60,
123.00, 119.10, 115.92, 39.90; Anal. Calcd. for
C16H14N4O2 (Mol.wt.294); C, 65.30; H, 4.79; N, 19.04;
Found: C, 65.22; H, 4.70; N, 18.00%.
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 2-(1,3-dioxoisoindolin-2-yl)-
3-methylbutanethioate (9d)
Yield, 70%; mp 240ꢀC-245ꢀC; IR (cm−1): ʋmax: 1780-1730
(imidic CO), 1715 (C O of ester), 1610 (C N); Ms: m/z
612 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.45-8.22 (m, 16H,
Ar─H), 3.40 (s, 1H, CH), 2.55(m, 1H, CH), 1.19 (d, 6H,
J = 8.45, 2CH3); 13C NMR, 197.62, 167.90, 152.00, 133.11,
132.21, 129.40, 127.71, 126.62, 123.70, 76.80, 29.00, 19.45;
Anal. Calcd. for C35H24N4O5S (Mol.wt.612): C, 68.62; H,
3.95; N, 9.15; Found: C, 68.52; H, 3.85; N, 9.08%.
4-(4-Aminophenyl)phthalazin-1-yl alaninate (10b)
Yield, 69%; mp 235ꢀC-238ꢀC; IR (cm−1): ʋmax: 3310-3100
(2NH2), 1730 (C O of ester), 1600 (C N); Ms: m/z
308 (Mˑ+); 1H NMR (DMSO-d6) δ: 8.76 (s, 2H, NH2),
7.50-7.93 (m, 8H, Ar─H), 5.52 (s, 2H, NH2), 3.53 (q, 1H,
J = 7.2, CH), 1.33 (d, 3H, J = 6.8, CH3); 13C NMR, 171.70,
150.31, 145.61, 143.00, 132.80, 131.80, 128.30, 127.60,
126.70, 123.02, 119.70, 116.40, 115.11, 49.51, 16.80; Anal.
Calcd. for C17H16N4O2 (Mol.wt.308); C, 66.22; H, 5.23; N,
18.17; Found: C, 66.20; H, 5.20; N, 18.00%.
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 2-(1,3-dioxoisoindolin-2-yl)-
3-hydroxypropanethio-ate (9e)
Yield, 60%; mp 230ꢀC-235ꢀC; IR (cm−1): ʋmax: 1780-1730
(imidic CO), 1715 (C O of ester), 1610 (C N); Ms: m/z 600
1
(Mˑ+); H NMR (DMSO-d6) δ: 7.45-8.22 (m, 16H, Ar─H),
4.94 (s, 1H, OH), 4.40 (t, 1H, J = 7.5, CH), 3.92 (d, 2H,
J = 7.1, CH2); Anal. Calcd. for C33H20N4O6S (Mol.
wt.600): C, 65.99; H, 3.36; N, 9.33; Found: C, 65.89; H,
3.30; N, 9.23%.
4-(4-Aminophenyl)phthalazin-1-yl 2-amino-
2-phenyla-cetate (10c)
Yield, 75%; mp 239ꢀC-241ꢀC; IR (cm−1): ʋmax: 3450-3200
(2NH2), 1730 (C O of ester), 1600 (C N); Ms: m/z
500 (Mˑ+); 1H NMR (DMSO-d6) δ: 8.40 (s, 2H, NH2),
7.50-7.80 (m, 12H, Ar─H), 4.53 (t, 1H, J = 6.5, CH), 2.46
(d, 2H, J = 7.5, CH2); Anal. Calcd. for C22H18N4O2 (Mol.
wt.370); C, 71.34; H, 4.90; N, 15.13; Found: C, 71.30; H,
4.80; N, 15.10%.
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 2-(1,3-dioxoisoindolin-2-yl)-
3-hydroxybutanethioate (9f)
Yield, 60%; mp 250ꢀC-252ꢀC; IR (cm−1): ʋmax: 1780-1730
(imidic CO), 1715 (C O of ester), 1610 (C N); Ms: m/z
614 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.45-8.22 (m, 16H,
Ar─H), 5.56 (s, 1H, OH), 4.40 (d, 1H, J = 6.9, CH),
2.75(m, 1H, CH), 1.18 (d, 3H, J = 7.5, CH3); 13C NMR,
197.62, 167.10, 152.00, 133.11, 132.21, 129.40, 128.60,
127.71, 126.61, 123.70, 77.50, 65.50, 19.50; Anal. Calcd.
for C34H22N4O6S (Mol.wt.614): C, 66.44; H, 3.61; N, 9.12;
Found: C, 66.32; H, 3.52; N, 9.01%.
4-(4-Aminophenyl)phthalazin-1-yl valinate (10d)
Yield, 70%; mp 230ꢀC-232ꢀC; IR (cm−1): ʋmax: 3380-3150
(2NH2), 1730 (C O of ester), 1600 (C N); Ms: m/z
466 (Mˑ+); 1H NMR (DMSO-d6) δ: 8.11 (s, 2H, NH2),

WASFY ET AL.
7
7.43-7.80 (m, 12H, Ar─H), 4.53 (d, 1H, J = 8.32, CH),
2.15 (m, 1H, CH), 1.46 (d, 6H, J = 7.5, CH3); 13C NMR,
171.00, 168.30, 167.10, 166.01, 165.01, 152.01,133.50,
132.00, 129.40, 128.50, 127.70, 126.70, 119.70, 59.00,
30.40, 18.90; Anal. Calcd. for C19H20N4O2 (Mol.
wt.336); C, 67.84; H, 5.99; N, 16.66; Found: C, 67.80; H,
5.90; N, 16.60%.
2-Amino-N-(4-(4-aminophenyl)phthalazin-1-yl)-3-
phenyl propanamide (11c)
Yield, 70%; mp 240ꢀC-242ꢀC; IR (cm−1): ʋmax: 3350-3100
1
(NH, 2NH2), 1700 (C O amide), 1620 (C N); H NMR
(DMSO-d6) δ: 10.61 (s, 1H, NH, exchangeable), 8.86 (s,
2H, NH2, exchangeable), 7.14-7.86 (m, 17H, Ar─H), 4.53
(t, 1H, J = 6.8, CH), 3.16 (d, 2H, J = 6.8, CH2); Ms: m/z
500 (Mˑ+); Anal. Calcd. for C22H19N5O (Mol.wt.369); C,
71.53; H, 5.18; N, 18.96; Found: C, 71.50; H,
5.10; N, 18.90%.
4-(4-Aminophenyl)phthalazin-1-yl serinate (10e)
Yield, 70%; mp 229ꢀC-231ꢀC; IR (cm−1): ʋmax: 3400-3050
(2NH2), 1730 (C O of ester), 1600 (C N); 1H NMR
(DMSO-d6) δ: 8.52 (s, 2H, NH2), 7.39-7.95 (m, 12H,
Ar─H), 5.45 (s, 1H, OH), 4.53 (t, 1H, J = 7.5, CH), 3.47
(d, 2H, J = 6.8, CH2); Ms: m/z 324 (Mˑ+); Anal. Calcd. for
C17H16N4O3 (Mol.wt.324); C, 62.95; H, 4.97; N, 17.27;
Found: C, 62.90; H, 4.90; N, 17.20%.
2-Amino-N-(4-(4-aminophenyl)phthalazin-1-yl)-3-
methylbutanamide (11d)
Yield, 78%; mp 250ꢀC-254ꢀC; IR (cm−1): ʋmax: 3400-3200
1
(NH, 2NH2), 1690 (C O amide), 1620 (C N); H NMR
(DMSO-d6) δ: 10.21 (s, 1H, NH), 8.33 (s, 2H, NH2),
7.33-7.80 (m, 12H, Ar─H), 4.43 (d, 1H, J = 7.5, CH), 2.30
(m, 1H, CH), 1.26 (d, 6H, J = 6.5, CH3); Ms: m/z
335 (Mˑ+); Anal. Calcd. for C19H21N5O (Mol.wt.335); C,
68.04; H, 6.31; N, 20.88; Found: C, 68.00; H,
6.21; N, 20.80%.
4-(4-Aminophenyl)phthalazin-1-yl 2-amino-
3-hydroxy-butanoate (10f)
Yield, 78%; mp 201ꢀC-203ꢀC; IR (cm−1): ʋmax: 3300-3180
(2NH2), 1730 (C O of ester), 1600 (C N); 1H NMR
(DMSO-d6) δ: 8.96 (s,2H,NH2), 7.32-7.93 (m, 12H, Ar─H),
5.37 (s, 1H, OH), 4.28 (d, 1H, J = 7, CH), 3.36 (q, 1H, J = 7,
CH), 1.16 (d, 3H, J = 7.5, CH2); 13C NMR, 171.00, 168.30,
167.10, 150.01,133.50, 132.00, 129.40, 128.50, 127.70,
123.70, 119.70, 67.40, 60.50, 19.42; Ms: m/z 338 (Mˑ+);
Anal. Calcd. for C18H18N4O3 (Mol.wt.338.14); C, 63.89; H,
5.36; N, 16.56; Found: C, 63.80; H, 5.30; N, 16.50%.
2-Amino-N-(4-(4-aminophenyl)phthalazin-1-yl)-3-
hydroxypropanamide (11e)
Yield, 80%; mp 210ꢀC-212ꢀC; IR (cm−1): ʋmax: 3350 (NH,
2NH2), 3100 (C─H aromatic), 2940 (C─H aliphatic), 1698
1
(C O amide), 1620 (C N); Ms: m/z 323 (Mˑ+); H NMR
(DMSO-d6) δ: 10.43(s, 1H, NH, exchangeable), 8.50 (s,
2H, NH2, exchangeable), 7.43-7.97 (m, 12H, Ar─H), 4.53
(t, 1H, J = 6.5, CH), 3.16 (d, 2H, J = 6.8, CH2); Anal.
Calcd. for C17H17N5O2 (Mol.wt.323); C, 63.15; H, 5.30; N,
21.66; Found: C, 63.10; H, 5.20; N, 21.60%.
2-Amino-N-(4-(4-aminophenyl)phthalazin-1-yl)
acetamide (11a)
Yield, 78%; mp 201ꢀC-203ꢀC; IR (cm−1): ʋmax: 3400-3250
1
(NH, 2NH2), 1680 (C O amide), 1600 (C N); H NMR
2-Amino-N-(4-(4-aminophenyl)phthalazin-1-yl)-3-
hydroxybutanamide (11f)
(DMSO-d6) δ: 8.60 (s, 1H, NH, exchangeable), 7.50 (s, 2H,
NH2, exchangeable), 7.60-7.95 (m, 12H, Ar─H), 4.41 (s,
2H, CH2); Ms: m/z 468 (Mˑ+); Anal. Calcd. for
C16H15N5O (Mol.wt.293); C, 65.52; H, 5.15; N, 23.88;
Found: C, 65.41; H, 5.10; N, 23.80%.
Yield, 70%; mp 210ꢀC-212ꢀC; IR (cm−1): ʋmax: 3350 (NH,
2NH2), 3100 (C─H aromatic), 2940 (C─H aliphatic), 1680
1
(C O of amide), 1620 (C N); Ms: m/z 467 (Mˑ+); H
NMR (DMSO-d6) δ: 9.43 (s, 1H, NH), 8.87 (s, 2H, NH2),
7.43-7.80 (m, 12H, Ar─H), 4.53 (d, 1H, J = 7.5, CH), 3.36
(m, 1H, CH), 1.20 (d, 3H, J = 6.8, CH3); Anal. Calcd. for
C18H19N5O2 (Mol.wt.337); C, 64.08; H, 5.68; N, 20.76;
Found: C, 64.00; H, 5.60; N, 20.70%.
2-Amino-N-(4-(4-aminophenyl)phthalazin-1-yl)
propanamide (11b)
Yield, 80%; mp 198ꢀC-200ꢀC; IR (cm−1): ʋmax: 3360-3010
1
(NH, 2NH2), 1660 (C O amide), 1620 (C N); H NMR
(DMSO-d6) δ: 9.50 (s, 1H, NH, exchangeable), 7.42 (s, 2H,
NH2, exchangeable), 7.43-8.11 (m, 12H, Ar─H), 4.47 (q,
1H, J = 7.3, CH), 1.25 (d, 3H, J = 7, CH3); 13C NMR,
172.70, 167.10, 166.10, 143.01, 132.21, 131.80, 129.40,
128.60, 127.70, 123.70, 119.70, 116.40, 49.50, 16.80; Ms:
m/z 437 (Mˑ+); Anal. Calcd. for C17H17N5O (Mol.
wt.307); C, 66.43; H, 5.58; N, 22.79; Found: C, 66.40; H,
5.50; N, 22.70%.
S-(4-(4-Aminophenyl)phthalazin-1-yl)
2-aminoethanethioate (12a)
Yield, 60%; mp 301ꢀC-303ꢀC; IR (cm−1): ʋmax: 3350-3020
(2NH2), 1710 (C O), 1620 (C N); Ms: m/z 440 (Mˑ+); 1H
NMR (DMSO-d6) δ: 8.50 (s, 2H, NH2), 7.43-7.97 (m, 12H,
Ar─H), 4.53 (s, 2H, CH2); Anal. Calcd. for C16H14N4OS
(Mol.wt.310); C, 61.92; H, 4.55; N, 18.05; Found: C,
61.87; H, 4.50; N, 18.00%.

8
WASFY ET AL.
S-(4-(4-Aminophenyl)phthalazin-1-yl)
2-aminopropanethioate (12b)
4.1.6 | General procedure for reaction of
(2, 4, 5) with N-tosyl-amino acids:
Formation of (14-16)a-f
Yield, 65%; mp 312ꢀC-314ꢀC; IR (cm−1): ʋmax: 3400-3150
(2NH2), 1700 (C O), 1620 (C N); Ms: m/z 454 (Mˑ+); 1H
NMR (DMSO-d6) δ: 8.50 (s, 2H, NH2, exchangeable),
7.40-8.15 (m, 12H, Ar─H), 3.97 (q, 1H, J = 7.1, CH), 1.09
(d, 3H, J = 6.8, CH3); Anal. Calcd. for C17H16N4OS (Mol.
wt.324); C, 62.94; H, 4.97; N, 17.27; Found: C, 62.90; H,
4.90; N, 17.17%.
Similarly, phthalazine-1(1H)-one 2, aminophthalazine 4,
and mercaptophthalazine 5 reacted with N-tosylamino
acids 13a-f in presence of DCCI under the same reaction
conditions to give to afforded compounds (14-16)a-f.
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-
1-yl tosylglycinate (14a)
S-(4-(4-Aminophenyl)phthalazin-1-yl) 2-amino-
3-methylbutanethioate (12c)
Yield, 75%; mp 230ꢀC-232ꢀC; IR (cm−1): ʋmax: 3430 (NH),
1781-1700 (3CO), 1593 (C N), 1380 (SO2); Ms: m/z 578 (Mˑ
Yield, 69%; mp 210ꢀC-212ꢀC; IR (cm−1): ʋmax: 3380-3100
(2NH2), 3100 (C─H aromatic), 1690 (C O), 1620 (C N);
1H NMR (DMSO-d6) δ: 8.11 (s, 2H, NH2, exchangeable),
7.10-7.78 (m, 17H, Ar─H), 4.53 (t, 1H, J = 7.1, CH), 3.16
(d, 2H, J = 6.5, CH2); Ms: m/z 352 (Mˑ+); Anal. Calcd.
for C19H20N4OS (Mol.wt.352); C, 64.75; H, 5.72; N, 15.90;
Found: C, 64.70; H, 5.62; N, 15.80%.
1
+); H NMR (DMSO-d6) δ: 7.81 (s, 1H, NH), 7.32-7.95 (m,
16H, Ar─H), 4.34 (s, 2H, CH2), 2.35 (s, 3H, CH3); 13C NMR,
171.00, 168.30, 167.10, 150.01,136.30, 133.50, 132.00, 129.40,
128.50, 127.70, 126.62, 123.70, 119.70, 43.80; Anal. Calcd. for
C31H22N4O6S (Mol.wt.578); C, 64.35; H, 3.83; N, 9.68;
Found: C, 64.25; H, 3.80; N, 9.62%.
S-(4-(4-Aminophenyl)phthalazin-1-yl) 2-amino-
3-me-thylbutanethioate (12d)
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-
1-yl tosylalaninate (14b)
Yield, 45%; mp 330ꢀC-332ꢀC; IR (cm−1): ʋmax: 3300-3280
(2NH2), 3100 (C─H aromatic), 2940 (C─H aliphatic),
Yield, 70%; mp 235ꢀC-237ꢀC; IR (cm−1): ʋmax: 3368 (NH),
1781-1701 (3CO), 1607 (C N), 1381 (SO2); Ms: m/z
1
1705 (C O), 1620 (C N); Ms: m/z 352 (Mˑ+); H NMR
1
592 (Mˑ+); H NMR (DMSO-d6) δ: 10.90 (s, 1H, NH),
(DMSO-d6) δ: 7.50 (s, 2H, NH2), 7.43-7.97 (m, 16H,
Ar─H), 4.67 (d, 1H, J = 6.2, CH), 2.46 (m, 1H, CH), 1.01
(d, 6H, J = 7.5, 2CH3); Anal. Calcd. for C19H20N4OS
(Mol.wt.352); C, 64.75; H, 5.72; N, 15.90; Found: C,
64.70; H, 5.62; N, 15.80%.
7.03-7.76 (m, 16H, Ar─H), 4.57 (s, 1H, CH), 1.31 (q, 3H,
J = 7.2, CH3), 2.38 (s, 3H, CH3─Ar); Anal. Calcd. for
C32H24N4O6S (Mol.wt.592); C, 64.86; H, 4.08; N, 9.45;
Found: C, 64.80; H, 4.01; N, 9.40%.
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-
1-yl tosylphenylalaninate (14c)
S-(4-(4-Aminophenyl)phthalazin-1-yl) 2-amino-
3-hydro-xypropanethioate (12e)
Yield, 65%; mp 240ꢀC-242ꢀC; IR (cm−1): ʋmax: 3300 (NH),
1781-1735 (3CO), 1589 (C N), 1382 (SO2); Ms: m/z 668 (Mˑ
Yield, 50%; mp 338ꢀC-340ꢀC; IR (cm−1): ʋmax: 3390-3250
(2NH2), 1730 (C O of ester), 1620 (C N); Ms: m/z
340 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.20 (s, 2H, NH2),
7.55-7.87 (m, 12H, Ar─H), 4.42 (t, 1H, J = 7, CH), 2.46 (d,
2H, J = 6.8, CH2), 5.50 (s, 1H, OH); 13C NMR, 197.60,
167.30, 152.01,133.10, 132.20, 129.40, 128.50, 127.70,
126.40, 123.70, 71.10, 64.70; Anal. Calcd. for C17H16N4O2S
(Mol.wt.340); C, 59.98; H, 4.74; N, 16.46; Found: C,
59.90; H, 4.70; N, 16.40%.
1
+); H NMR (DMSO-d6) δ: 11.44 (s,1H, NH), 7.02-8.28 (m,
16H, Ar─H), 4.89 (t, 1H, J = 8.2, CH), 2.68 (d, 2H, J = 7.5,
CH2), 2.28 (s, 3H, CH3); 13C NMR, 171.00, 168.30, 167.10,
150.01,136.30, 141.50, 137.50, 133.50, 132.00, 129.40, 128.50,
127.70, 126.62, 123.70, 119.70, 58.30, 35.82, 21.30; Anal.
Calcd. for C38H28N4O6S (Mol.wt.668); C, 68.25; H, 4.22; N,
8.38; Found: C, 68.20; H, 4.21; N, 8.30%.
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-
1-yl-tosylvalinate (14d)
S-(4-(4-Aminophenyl)phthalazin-1-yl) 2-amino-
3-hydro-xybutanethioate (12f)
Yield, 70%; mp 215ꢀC-217ꢀC; IR (cm−1): ʋmax: 3328
(NH),1785-1700 (3CO), 1575 (C N), 1329 (SO2); 1H NMR
(DMSO-d6) δ: 10.10 (s, 2H, NH2), 7.18-7.30 (m, 16H,
Ar─H), 5.12 (d, 1H, J = 6.3, CH), 3.13-3.46 (m, 1H, CH),
2.45 (s, 3H, CH3─Ar), 1.01 (d, 6H, J = 7.2, 2CH3); Anal.
Calcd. for C34H28N4O6S (Mol.wt.620); Found: C,
65.79; H, 4.55; N, 9.03; O, 15.47; S, 5.17%.
Yield, 70%; mp 338ꢀC-340ꢀC; IR (cm−1): ʋmax: 3300-3260
(2NH2), 1730 (C O), 1620 (C N); Ms: m/z 484 (Mˑ+); 1H
NMR (DMSO-d6) δ: 7.44 (s, 2H, NH2), 7.43-7.97 (m, 12H,
Ar─H), 4.03 (d, 1H, J = 8, CH), 2.11 (m, 1H, CH), 5.22 (s,
1H, OH); Anal. Calcd. for C18H18N4O2S (Mol.wt.354); C,
61.00; H, 5.12; N, 15.81; Found: C, 60.90; H,
5.02; N, 15.71%.

WASFY ET AL.
9
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-
1-yl-tosylserinate (14e)
N-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl)-2-((4-methylphenyl)
sulfonamido)-3-phenylpropa-namide (15c)
Yield, 75%; mp 220ꢀC-222ꢀC; IR (cm−1): ʋmax: 3330 (NH,
OH), 3050 (C─H aromatic), 2950 (C─H aliphatic),
Yield, 70%; mp 200ꢀC-202ꢀC; IR (cm−1): ʋmax: 3380 (NH),
1780-1700 (3CO), 1600 (C N); Ms: m/z 653 (Mˑ+); H
1
1
1780-1700 (3C O), 1600 (C N), 1330 (SO2); H NMR
(DMSO-d6) δ: 8.99 (s,1H, NH), 7.43-7.96 (m, 16H, Ar─H),
5.01(s,1H, OH), 2.34 (d, 2H, J = 6.4, CH2), 4.22 (t, 1H,
J = 6.8, CH), 2.80 (s, 3H, CH3); Anal. Calcd. for
C32H24N4O7S (Mol.wt.608); C, 63.15; H, 3.97; N, 9.21;
Found: C, 63.10; H, 3.91; N, 9.11%.
NMR (DMSO-d6) δ: 11.92 (s, 1H, NH, exchangeable with
D2O), 7.15-8.32 (m, 16H, Ar─H), 4.22 (t,1H, J = 7.5, CH),
2.70 (d, 2H, J = 6.3, CH2), 3.12 (s, 3H, CH3); Anal. Calcd.
for C38H29N5O5S (Mol.wt.667); C, 68.35; H, 4.38; N,
10.49; Found: C, 68.30; H, 4.30; N, 10.39%.
4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)phthalazin-
1-yl 3-hydroxy-2-((4-methylphenyl) sulfonamido)
butanoate (14f)
N-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl)-3-methyl-2-((4-methylphenyl)
sulfonamido)butana-mide (15d)
Yield, 70%; mp 226ꢀC-228ꢀC; IR (cm−1): ʋmax: 3400-3100
(NH, OH), 3080 (C─H aromatic), 2970 (C─H aliphatic),
1780-1700 (3CO), 1600 (C N), 1330 (SO2); Ms: m/z
622 (Mˑ+); 1H NMR (DMSO-d6) δ: 8.42 (s,1H,NH),
7.13-7.90 (m, 16H, Ar─H), 4.90 (s, 1H, OH), 4.34 (d, 1H,
J = 7.5, CH), 2.34 (m,1H, CH), 1.20 (d, 3H, J = 6.8, CH3),
3.55 (s, 3H, CH3─Ar); 13C NMR, 171.00, 168.30, 167.10,
150.01,136.30, 133.50, 132.00, 129.40, 128.50, 127.70,
126.62, 123.70, 119.70, 43.80; Anal. Calcd. for
C32H24N4O7S (Mol.wt.622); C, 63.66; H, 4.21; N, 9.00;
Found: C, 63.60; H, 4.11; N, 9.12%.
Yield, 55%; mp 220ꢀC-222ꢀC; IR (cm−1): ʋmax: 3400 (NH,
CONH), 1780-1700 (3CO), 1600 (C N); Ms: m/z 619 (Mˑ
+); H NMR (DMSO-d6) δ: 10.23 (s, 1H, NH, exchange-
1
able with D2O), 7.14-7.86 (m, 16H, Ar─H), 5.22 (d,1H,
J = 6.4, CH), 3.55-3.65 (m, 1H, CH), 2.75 (d, 6H, J = 7.8,
2CH3), 1.01 (s, 3H, CH3─Ar); Anal. Calcd. for
C34H29N5O5S (Mol.wt.619); C, 65.90; H, 4.72; N, 11.30;
Found: C, 65.81; H, 4.61; N, 11.23%.
N-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl)-3-hydroxy-2-((4-methylphenyl)
sulfonamido)propanamide (15e)
N-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl)-2-((4-methylphenyl)
Yield, 65%; mp 225ꢀC-227ꢀC; IR (cm−1): ʋmax: 3298-3280
(NH, OH), 1780-1710 (3CO), 1600 (C N); Ms: m/z
607 (Mˑ+); 1H NMR (DMSO-d6) δ: 10.55 (s, 1H, NH,
exchangeable with D2O), 7.40-7.86 (m, 16H, Ar─H),
5.50 (s, 1H, OH), 4.52 (d, 1H, J = 6.8, CH), 3.22 (d, 2H,
J = 7.5, CH2), 2.75 (s, 3H, CH3─Ar); 13C NMR, 172.00,
168.30, 167.10, 150.01,136.30, 133.50, 132.00, 129.40,
128.50, 127.70, 126.62, 123.70, 119.70, 60.50, 59.81,
21.43; Anal. Calcd. for C32H25N5O6S (Mol.wt.607); C,
63.25; H, 4.15; N, 11.53; Found: C, 63.12; H,
4.10; N, 11.42%.
sulfonamido)acetamide (15a)
Yield, 60%; mp 203ꢀC-205ꢀC; IR (cm−1): ʋmax: 3400 (NH),
1780-1700 (3CO), 1710 (C O), 1600 (C N); Ms: m/z
577 (Mˑ+); 1H NMR (DMSO-d6) δ: 10.70 (s, 1H, NH,
exchangeable with D2O), 7.13-7.90 (m, 16H, Ar─H), 3.45
(s, 2H, CH2), 2.12 (s, 3H, CH3); 13C NMR, 171.00, 168.30,
167.10, 150.01,136.30, 133.50, 132.00, 129.40, 128.50,
127.70, 126.62, 123.70, 119.70, 43.80, 21.36; Anal. Calcd.
for C31H23N5O5S (Mol.wt.577); C, 64.46; H, 4.01; N,
12.12; Found: C, 64.40; H, 4.00; N, 12.02%.
N-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl)-2-((4-methylphenyl)
N-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl)-3-hydroxy-2-((4-methylphenyl)
sulfonamido)butanamide (15f)
sulfonamido)propanamide (15b)
Yield, 68%; mp 190ꢀC-192ꢀC; IR (cm−1): ʋmax: 3320 (NH),
1780-1710 (3CO), 1710, 1600 (C N); Ms: m/z 591 (Mˑ+);
1H NMR (DMSO-d6) δ: 8.34 (s, 1H, NH, exchangeable
with D2O), 7.03-7.26 (m, 16H, Ar─H), 4.22 (q, 1H,
J = 6.7, CH), 2.35 (s, 3H, CH3─Ar), 1.29 (d, H, J = 7.1,
CH3); 13C NMR, 171.00, 168.30, 167.10, 150.01, 136.30,
133.50, 132.00, 129.40, 128.50, 127.70, 126.62, 123.70,
119.70, 53.40, 21.30, 17.00; Anal. Calcd. for C32H25N5O5S
(Mol.wt.591); C, 64.96; H, 4.26; N, 11.84; Found: C,
64.90; H, 4.22; N, 11.72%.
Yield, 70%; mp 242ꢀC-244ꢀC; IR (cm−1): ʋmax: 3300-3180
(NH, OH), 1780-1730 (3CO), 1600 (C N); Ms: m/z
621 (Mˑ+); 1H NMR (DMSO-d6) δ: 9.12 (s, 1H, NH,
exchangeable with D2O), 7.20-7.56 (m, 16H, Ar─H), 5.20
(s, 1H, OH), 4.52 (d, 1H, J = 7, CH), 3.22 (q, 1H, J = 7.2,
CH), 2.58 (s, 3H, CH3─Ar), 1.32 (d, 3H, J = 7, CH3); Anal.
Calcd. for C33H27N5O6S (Mol.wt.621); C, 63.76; H,
4.38; N, 11.27; Found: C, 63.71; H, 4.30; N, 11.15%.

10
WASFY ET AL.
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 2-((4-methylphenyl)
sulfonamido)ethanethioate (16a)
phthalazin-1-yl) 3-hydroxy-2-((4-methylphenyl)
sulfonamido)propa-nethioate (16e)
Yield, 60%; mp 233ꢀC-235ꢀC; IR (cm−1): ʋmax: 3295 (NH),
1780-1700 (3CO), 1600 (C N), 1350 (SO2); Ms: m/z
594 (Mˑ+); 1H NMR (DMSO-d6) δ: 8.51 (s, 1H, NH,
exchangeable with D2O), 7.40-7.91 (m, 16H, Ar─H), 4.32
(s, 2H, CH2), 2.12 (s, 3H, CH3); 13C NMR, 196.50, 168.30,
167.10, 150.01,136.30, 133.50, 132.00, 129.40, 128.50,
127.70, 126.62, 123.70, 119.70, 54.00, 21.33; Anal. Calcd.
for C31H22N4O5S2 (Mol.wt.594); C, 62.61; H, 3.73; N, 9.42;
Found: C, 62.51; H, 3.62; N, 9.31%.
Yield, 65%; mp 240ꢀC-242ꢀC; IR (cm−1): ʋmax: 3390 (NH),
3080 (C─H aromatic), 2970 (C─H aliphatic), 1780-1690
(3CO), 1600 (C N), 1350 (SO2); Ms: m/z 624 (Mˑ+); H
1
NMR (DMSO-d6) δ: 8.75 (s, 1H, NH, exchangeable with
D2O), 7.20-7.99 (m, 16H, Ar─H), 5.53(s, 1H, OH), 4.12 (t,
1H, J = 7.5, CH), 2.85 (d, 2H, J = 7, CH2), 2.40 (s, 3H, CH3);
Anal. Calcd. for C32H24N4O6S2 (Mol.wt.624); C, 61.53; H,
3.87; N, 8.97; Found: C, 61.41; H, 3.80; N, 8.91%.
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 3-hydroxy-2-((4-methyl phenyl)
sulfonamido)butane-thioate (16f)
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 2-((4-methylphenyl)
Yield, 68%; mp 210ꢀC-212ꢀC; IR (cm−1): ʋmax: 3320 (NH),
1780-1680 (3CO), 1600 (C N), 1350 (SO2); Ms: m/z
638 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.85 (s, 1H, NH,
exchangeable with D2O), 7.52-8.21 (m, 16H, Ar─H),
5.03(s, 1H, OH), 4.42 (d, 1H, J = 6.8, CH), 2.11 (q, 1H,
J = 6.8, CH), 1.12 (d, 3H, J = 7.4, CH3), 2.58 (s, 3H,
CH3─Ar); 13C NMR, 197.76, 168.30, 167.10,
150.01,136.30, 133.50, 132.00, 129.40, 128.50, 127.70,
126.62, 123.70, 119.70, 74.11, 67.30, 19.51; Anal. Calcd.
for C33H26N4O6S2 (Mol.wt.638); C, 62.06; H, 4.10; N, 8.77;
Found: C, 62.00; H, 4.00; N, 8.70%.
sulfonamido)propanethioate (16b)
Yield, 50%; mp 243ꢀC-245ꢀC; IR (cm−1): ʋmax: 3390 (NH),
3080 (C─H aromatic), 1780-1730 (3CO), 1600 (C N), 1350
1
(SO2); Ms: m/z 608 (Mˑ+); H NMR (DMSO-d6) δ:7.85 (s,
1H, NH, exchangeable with D2O), 7.43-8.30 (m, 16H,
Ar─H), 4.25 (q, 1H, CH), 2.36 (s, 3H, CH3─Ar), 1.36 (d, 3H,
J = 7, CH3); Anal. Calcd. for C32H24N4O5S2 (Mol.wt.608); C,
63.14; H, 3.97; N, 9.20; Found: C, 63.10; H, 3.90; N, 9.10%.
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 2-((4-methylphenyl)
sulfonamido)-3-phenylpropane-thioate (16c)
Yield, 55%; mp 240ꢀC-242ꢀC; IR (cm−1): ʋmax: 3750
(NH), 1780-1705 (3CO), 1600 (C N), 1350 (SO2); Ms:
5 | PHARMACOLOGICAL
5.1 | Cytotoxicity and anticancer
evaluation
1
m/z 670 (Mˑ+); H NMR (DMSO-d6) δ: 8.22 (s,1H,NH,
exchangeable with D2O), 7.04-7.66 (m, 16H, Ar─H),
4.29 (t, 1H, J = 7.5, CH), 2.88 (d, 2H, J = 6.9, CH2),
2.35 (s, 3H, CH3); Anal. Calcd. for C38H28N4O5S2 (Mol.
wt.684); C, 66.65; H, 4.12; N, 8.18; Found: C, 66.60; H,
4.08; N, 8.10%.
Different human cancer cell lines, namely, breast cancer
(MCF-7), hepatic cancer (HePG-2), human prostate can-
cer (PC3), and colon cancer (HCT-116) of some novel
synthesized compounds were measured via standard
MTT assay in vitro.[42–44] This colorimetric assay is rely-
ing on the changing of the yellow tetrazolium bromide to
a purple formazan derivative by mitochondrial succinate
dehydrogenase enzyme. The synthesized compounds
showed different degrees of inhibitory activity toward the
tested human tumor cell lines compared with the stan-
dard doxorubicin. Compounds 8c, 10a, 10d, 10f, 10e,
11d, 11e, 11b, and 12e revealed the highest cytotoxic
activity among the studied series toward HePG-2,MCF-7,
PC3, and HCT-116, with IC50 ranges between 4 and
10 mg/mL. The highest cytotoxic activity against PC3 was
shown by compounds 7f, 10a, 10d, 10f, 11d, 11e, 11b,
and 12e with IC50 values 9.72, 5.43, 8.33, 6.22, 6.51, 5.55,
7.22, and 9.05 mg/mL, respectively. Powerful inhibitory
activity was also established by compounds 10a, 11d, 11e,
S-(4-(4-(1,3-Dioxoisoindolin-2-yl)phenyl)
phthalazin-1-yl) 3-methyl-2-((4-methylphenyl)
sulfonamido)butane-thioate (16d)
Yield, 70%; mp 205ꢀC-207ꢀC; IR (cm−1): ʋmax: 3360 (NH),
1780-1700 (3CO), 1600 (C N), 1350 (SO2); Ms: m/z
636 (Mˑ+); 1H NMR (DMSO-d6) δ: 7.31 (s, 1H, NH,
exchangeable with D2O), 7.23-7.46 (m, 16H, Ar─H), 4.50
(d, 1H, J = 7, CH), 3.35-3.13 (m, 1H, CH), 2.49 (s, 3H,
CH3), 1.30 (d, 6H, J = 6.8, 2CH3); 13C NMR, 197.60,
168.30, 167.10, 150.01,136.30, 133.50, 132.00, 129.40,
128.50, 127.70, 126.62, 123.70, 119.70, 69.40, 61.10, 21.33;
Anal. Calcd. for C34H28N4O5S2 (Mol.wt.636); C, 64.14; H,
4.43; N, 8.80; Found: C, 64.10; H, 4.33; N, 8.71%.

WASFY ET AL.
11
and 11b toward MCF-7. At the other side, other com-
pounds showed cytotoxic activity that ranges from mod-
erate to weak toward the human cancer cell lines. It is
concluded from the given data in Table 1 that the exis-
tence of amino acid moieties attached to phthalazines
improves their cytotoxic effect in comparison with
starting phthalazines 2, 4, and 5. Also, combination of
phthaloyl phenyl alanine and phthaloyl L-threonine in
derivatives 8c and 7f upgraded their activity toward
HepG-2 and HCT-116. (Table 1).
In studying the structure-activity relationship (SAR)
between the activity and synthesized analogs, com-
pounds having electron donating groups (OH and
CH3) presents on the compounds (8c, 10(a, d, f, e), 11
(d, e, f) and 12e) were found to be the most potent
anticancer activity. The biological activities also depend
TABLE 1 Cytotoxic activity of some compounds against human tumor cells
In Vitro Cytotoxicity IC50 (μM)a
Compounds
DOXb
7a
HePG-2
MCF-7
PC3
8.87 0.6
HCT-116
4.50 0.2
14.46 1.3
32.37 2.4
12.48 1.2
7.30 0.5
58.53 3.6
9.16 0.8
52.87 3.4
20.68 1.9
30.71 2.1
20.37 2.8
15.44 1.8
7.90 1.3
6.90 0.3
4.85 0.4
2.95 0.2
9.95 0.2
5.27 3.8
7.11 1.7
12.11 1.5
11.24 1.6
10.11 3.7
37.10 2.6
82.48 4.7
91.47 5.1
40.71 3.1
10.16 1.0
29.50 2.2
18.24 1.6
50.60 4.1
30.58 1.9
20.51 2.6
4.17 0.2
26.28 2.1
42.91 3.4
17.62 1.7
15.83 1.4
70.52 4.3
11.50 1.2
46.66 3.6
30.51 2.4
50.16 2.5
32.81 3.2
30.41 2.0
10.14 2.1
11.03 1.1
12.50 1.5
9.64 0.8
6.64 0.8
9.15 2.5
9.50 2.1
15.50 3.01
13.51 3.0
12.50 1.5
42.03 3.2
>100
5.23 0.3
13.19 1.4
34.37 2.6
10.25 1.0
9.13 0.8
89.98 4.9
7.50 0.6
43.14 2.9
16.63 1.5
60.40 4.7
25.37 2.5
25.55 1.9
9.10 1.4
5.91 0.4
5.10 0.7
4.03 0.3
5.03 0.3
7.51 1.0
10.56 1.9
7.56 1.9
22.56 2.9
7.56 2.0
28.38 2.3
>100
28.43 2.3
48.83 3.5
19.40 1.8
9.72 1.0
79.28 4.2
13.89 1.3
51.71 3.5
38.97 2.7
40.83 2.9
50.83 3.5
22.05 2.5
5.43 1.9
8.33 0.9
10.15 0.7
6.22 0.5
7.22 0.5
6.51 1.9
5.55 2.7
10.05 2.05
16.05 2.5
9.05 2.8
44.98 3.1
>100
7b
7d
7f
8b
8c
8e
8f
9a
9b
9e
10a
10d
10e
10f
11b
11d
11e
12a
12b
12e
14c
14d
14e
15a
15b
15d
15e
16a
16d
16e
57.87 3.9
66.78 4.0
18.71 1.9
32.65 2.6
40.51 3.0
60.87 3.0
30.45 1.6
30.61 3.3
70.98 3.8
62.93 3.7
15.48 1.5
23.12 1.9
32.05 2.5
42.82 4.0
27.02 2.2
35.15 2.8
48.78 3.5
81.40 4.7
12.28 1.3
18.22 1.7
22.56 1.9
71.31 3.7
19.42 2.6
20.46 1.5
aIC50 (μM): 1-10 (very strong); 11-20 (strong); 21-50 (moderate); 51-100 (weak) and above 100 (non-cytotoxic).
bDOX, doxorubicin.

12
WASFY ET AL.
on the nature of amino acids presents on the heterocy-
clic moieties. On the basis of these previous results,
amino acids were playing a crucial role in increasing
biological activities. Some of the amino acids played a
key factor for increasing the anticancer activity. Amino
acids containing long-chain aliphatic hydrocarbons
showed more potent anticancer agents compared with
other conjugated amino acids such as glycine and ala-
nine derivatives. This fact may be explained on the
basis of aromaticity and hydrophobicity of phenylalanine,
valine, serine, and threonine possessing good anticancer
properties. The other two amino acid derivatives (glycine
and alanine) were also moderately active, which may be
due to their simple side chain functionalities and steric
hindrance of aliphatic side chains. While the number of
electron-donating groups of amino acids attached to
phthalazine derivatives (8c, 10(a, d, f, e), 11 (d, e, f) and
12e) increases, the anticancer activity also increases com-
pared with the minimum number of electron-donating
groups present on the amino acids attached to
TABLE 2 Results of ABTS assay
TABLE 3 Results of bleomycin-dependent DNA damage assay
on some selected compounds
Compound No.
Absorbance, mean
0.510
0.056
0.378
0.415
0.357
0.327
0.450
0.332
0.428
0.396
0.314
0.471
0.462
0.042
0.302
0.115
0.288
0.105
0.345
0.452
0.276
0.476
0.309
0.406
0.489
0.441
0.434
0.349
0.361
0.385
0.476
0.129
0.411
% Inhibition
0
Compounds
Absorbance of Samples
0.072
Control
L-absorbance
7a
89.0
25.9
18.6
30.0
35.9
11.8
34.9
16.1
22.3
38.4
7.6
Ascorbic acid
7a
0.089
7b
7b
0.106
7d
7d
0.086
7f
7f
0.074
8b
8b
0.127
8c
8c
0.078
8e
8e
0.123
8f
8f
0.097
9a
9a
0.145
9b
9b
0.139
9e
9.4
9e
0.154
10a
10d
10e
10f
91.8
40.8
77.5
43.5
97.4
32.4
11.4
45.9
6.7
10a
10d
10f
10e
11b
11d
11e
12a
12b
12e
14c
14d
14e
15a
15b
15d
15e
16a
16d
16e
0.058
0.069
0.067
0.064
11b
11d
11e
12a
12b
12e
14c
14d
14e
15a
15b
15d
15e
16a
16d
16e
0.074
0.066
0.072
0.063
0.053
39.4
20.4
4.1
0.061
0.111
0.153
13.5
14.9
31.6
29.2
24.5
6.6
0.132
0.089
0.081
0.104
0.093
0192
74.7
19.4
0.119
0.105

WASFY ET AL.
13
phthalazine rings. The presence of strong electron-donat-
ing groups (OH and CH3) on the aromatic ring increases
the hydrophobicity of the molecules and is responsible
for the enhanced anticancer activity. This fact suggests
the importance of the electron-withdrawing groups for
anticancer activity of this series.
It was detected that a lot of the synthesized derivatives
displayed potent cytotoxic activity from their IC50 values.
In studying the SAR, hydrophobic and aromatic amino
acids phenylalanine, valine, serine, and threonine, con-
taining electron-donating groups, were found to most
favor anticancer activity. Our results presented here
could be used as a preliminary stage for the development
of powerful phthalazine conjugated amino acids as anti-
cancer therapies.
5.2 | Antioxidant screening (ABTS)
ABTS, (2,20-azino-bis-3-ethylbenzothiazoline-6-sulfonic
acid diammonium salt), radical cation decolonization test
is a spectrophotometric method greatly used in evaluating
the antioxidant activity of different substances.[45] It can
be applied for both lipophilic and hydrophilic com-
pounds. Oxidation of ABTS gives a radical cation
(ABTS˙+) in presence of potassium persulfate. Because of
their capacity to reduce the preformed radical, the absor-
bance is bleached by antioxidants. The preliminary quali-
tative antioxidant screening results (scavenger activity) of
all the tested compounds are recorded (Table 2). This in
comparison with standard ascorbic acid; compounds 10a,
10e, and 11b showed a potent activity, while compounds
10d and 12a displayed moderate activity. The rest of the
compounds showed weak to mild activity.
ACKNOWLEDGMENTS
This work was supported by the financial assistance from
Benha University, Faculty of Science, Chemistry Depart-
ment, Egypt.
ORCID
Noura S. Mohamed https://orcid.org/0000-0003-2059-
2763
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How to cite this article: Wasfy AF, Aly AA,
Behalo MS, Mohamed NS. Synthesis of novel
phthalazine derivatives as pharmacological
activities. J Heterocyclic Chem. 2019;1–14. https://
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