Int J Curr Pharm Res, Vol 13, Issue 1, 81-86Original Article



1Department Pharmaceutical Chemistry C. U. Shah College of Pharmacy, S. N. D. T. Women’s University, Sir Vithaldas Vidyavihar, Juhu Road, Santacruz (West), Mumbai 400049, Maharashtra, India, 2Biocare Research India Pvt. Ltd.,1st Floor, Kanth Complex, Above New Limda Travels, Near Mahavir Jain Vidyalaya, Paldi Cross Road, Ahmedabad, 3Department Pharmaceutical Chemistry, Ideal College of Pharmacy and Research, Kalyan, Maharashtra, India 421306

Received: 15 Sep 2020, Revised and Accepted: 17 Nov 2020


Objective: Synthesis of N-1 Substituted Indolylchalcone Hybrids and evaluation of anti-angiogenic activity using Chorioallantoic Membrane (CAM) Assay.

Methods: Claisen-schmidt reaction is used for the synthesis of 30 Indolylchalcone hybrids, it involves condensation of N-1 substituted indole-3-carboxaldehyde and N1 substituted 2-acetyl-benzimidazole. The phase transfer catalyst, a green catalyst such as anhydrous potassium carbonate (K2CO3) and PEG-400 are used in the alkylation and arylation. All synthesized indolylchalcone hybrids were evaluated for their antiangiogenic activity by in vivo-chorioallantoic membrane (CAM) assay method.

Results: The synthesized indolylchalcone compounds are evaluated. The morphometric study was carried out as described by Melkonian et al. (2002). The Compounds with code C-2, I-1, I-2 are showing the more potent effect on the dose-dependent assay of CAM. The compounds with code C-1, C-3, E-1 to E-3, M-1 and M-5 shows the significant activity, however, though the compounds with code B-1, B-2, CL-1 and A-5 were showing antiangiogenic effect at 0.1 µM, but does not show any significant activity on dose-dependent assay of CAM.

Conclusion: The synthesized Indolylchalcones as shown in the graph possess very good dose-dependent anti-angiogenic activities. The potency of anti-angiogenetic activity shows that methyl>Ethyl>Cl-benzyl>Benzyl>Isobutyl. 2-acetyl benzimidazole analogs have possible future scope to develop as potent angiogenesis inhibitors.

Keywords: Angiogenesis, Indole chalcone, CAM assay


Angiogenesis or neovascularization is a combination of two Greek words, “angio” and “genesis” means vase and birth, respectively. Development of these blood capillaries, vessels plays a critical role in cell division, proliferation and movement [1]. More than 30 y ago, the scientist Folkman proposed that the role of these blood vessel to supply food, oxygen and nutrious to malignant tumor because of that growth was dependent on the development of tumor-associated blood vessels, a process called angiogenesis [2]. Targeting angiogenesis has proven to be an effective strategy in several malignant tumour not only colorectal, gastric and lung tumour but also breast, cervical and ovary tumour [3]. Anti-vascular approaches to cancer therapy may be primarily divided into antiangiogenesis inhibitors and vascular disrupting agents (VDAs) [4, 5].

The well-known marketed anti-angiogenic inhibitors drug, bevacizumab (Avastin) [6], and sorafenib (Nexavar, BAY 43-9006) and sunitinib malate (Sutent, SU11248) [7] have been approved by the United States Food and Drug Administration for the treatment of cancer patients. However, these marketed drugs has serious side effects, such as hypertension, bleeding and gastrointestinal perforation; they have been associated with currently available anti-VEGF agents, limiting their chronic use [8]. Hence, there is an urgent need to find a chalcone hybrid molecule that can be more potent towards cancerous cell and less toxic to normal cell but also may overcome the problem of multidrug-resistant strains.

It is well know that all type of heterocyclic ring comprises a very core active moiety or the pharmacophore [9-12]. Derivatives of nitrogen-containing fused heterocyclic compounds such as benzimidazole and indole core molecule have found number of application as an anticancer agent [13]. Literature survey reveals that there is interest in the indolylchalcone systems for use in cancer therapy and such indolylchalcone have the potential to improve the selectivity of chemotherapeutic agents and hence reduce unwanted side effects and multidrug resistance MDR syndrome [14, 15]. The well-known antiangiogenic drug sunitinib also has indole core, taking this in an account, chemistry of 3-substitution on indole moity shows that these has various biological activities and synthesis of such derivatives which has a skeleton that contains an indole cores and are also in chemistry of benzimidazole 2-substitution are valid for biological activities. Considering this in our study of rational research, we fused these indole and benzimidazole moieties with chalcone like structure (fig. 1). Thus three scaffolds moieties in our design are based on the diversity of these moieties in nature as well as in various marketed drugs.

Now a day’s 2-acetylbenzimidazole molecule and indole-3-carboxaldehyde are associated with a wide range of biological activities such as anticancer, anti-inflammatory, analgesic and anthelmintic etc. The N-substituted fused heterocyclic compounds are usually biologically active and may be used as potential therapeutic alternatives to the antitumor drug.


Synthesis of indolylchalcone

In this research study, a series of 30 Indolylchalcone hybrids were synthesized by Claisen-schmidt condensation of N-1 substituted indole-3-carboxaldehyde and N1 substituted 2-acetyl-benzimidazole. The reaction involved in the synthesis of indolylchalcone hybrids as shown in (Scheme 1). The N-alkylation/arylation, mono-methylation on indole-3-carboxaldehyde were achieved by using alkylating and arylating agents in the presence of phase transfer catalyst, a green catalyst such as anhydrous potassium carbonate (K2CO3) and PEG-400. The N1 substitution reaction on indole-3-carboxaldehyde and 2-aceyl-benzimidazole as shown in (scheme 1A and scheme 1B). The physicochemical properties and analytical data of synthesized indolylchalcone compounds are mentioned in table 1 and 2, respectively.

Scheme 1: Synthesis of indole chalcone

Scheme 1A: Synthesis of N-substituted indole-3-carbaldehyde derivatives

Scheme 1B: Synthesis of N-derivatives of 2-acetyl benzimidazole

Anti-angiogenic activity

Chicken egg chorioallantoic membrane assay (CAM)

The fertile eggs of 6th day was procured from IPDP poultry Maqarba, Ahmedabad and Angiogenesis inhibitory activities of 30 indolylchalcone hybrids were evaluated using quantitative CAM assay [16-18], Out of which 15 Indolylchalcone hybrids shows most inhibitory effect on in vivo assay models of angiogenesis as shown in (table 3). The morphometric study was carried out as described by Melkonian et al. (2002). Further, these indolylchalcone hybrids also shows the dose-dependent percentage inhibition of antiangiogenic activity of tested compounds calculated as per given formula (fig. 1)

% inhibition = [(vessel number of untreated CAM-vessel number of CAM treated with test compounds)/vessel number of untreated CAM] x100


The Compounds with code C-2, I-1, I-2 are showing more potent effect on dose dependent assay of CAM when compared to standard. Compounds with code C-2 and I-2 consist of N-1 substitution on ethyl group on both the indole and benzimidazole. Compound with code I-1 is unsubstituted indole, which consists of N-1 substitution of iso-butyl moity on benzimidazole. The compounds with code C-1, C-3, E-1 to E-3, M-1 and M-5 shows the significant activity, however, though the compounds with code B-1, B-2, CL-1 and A-5 were showing antiangiogenic effect at 0.1 µM, but does not show any significant activity on dose-dependent assay of CAM.

Table 1: Physicochemical properties of synthesized indolylchalcone

S. No. Name of the compound R-1 R-2 Molecular weight Molecular formula % yield of chalcone reaction Melting point RF
C-1 H H 287.31532 C18H13N3O 77 265 °C 0.6123
C-2 Ethyl H 315.36848 C20H17N3O 65 213 °C 0.1711
C-3 Benzyl H 377.43786 C25H19N3O 68 249 °C 0.45234
C-4 2-Cl benzyl H 411.8892 C25H18ClN3O 85 223 °C 0.2105
B-1 H Benzyl 377.4376 C25H19N3O 70 139 °C 0.2630
B-2 Ethyl Benzyl 405.49102 C27H23N3O 69 155 °C 0.756
B-3 Benzyl Benzyl 467.5604 C32H25N3O 72 183 °C 0.842
B-4 2-Cl benzyl Benzyl 502.00546 C32H24ClN3O 75 171 °C 0.754
E-1 H Ethyl 315.36898 C20H17N3O 55 205 °C 0.641
E-2 Ethyl Ethyl 343.42164 C22H21N3O 60 177 °C 0.6728
E-3 Benzyl Ethyl 405.49102 C27H23N3O 65 216 °C 0.5553
E-4 2-Cl benzyl Ethyl 439.93608 C27H22ClN3O 77 149 °C 0.453
CL-1 H 2-Cl benzyl 411.8892 C25H18ClN3O 79 219 °C 0.2156
CL-2 Ethyl 2-Cl benzyl 439.9360 C27H22ClN3O 81 181 °C 0.7800
CL-3 Benzyl 2-Cl benzyl 502.00546 C32H24ClN3O 84 232 °C 0.8600
CL-4 2-Cl benzyl 2-Cl benzyl 536.45052 C32H23Cl2N3O 68 226 °C 0.8039
A-1 Allyl H 327.37918 C21H19N3O 65 122 °C 0.1428
A-2 Allyl Ethyl 355.43234 C23H23N3O 68 101 °C 0.3564
A-3 Allyl Benzyl 417.50172 C28H25N3O 71 132 °C 0.5870
A-4 Allyl 2-Cl benzyl 451.94678 C28H24ClN3O 60 139 °C 0.5612
A-5 Allyl Iso-Butyl 399.5279 C25H27N3O 70 95 °C 0.4012
M-1 Methyl H 301.3419 C19H15N3O 71 142 °C 0.1451
M-2 Methyl Ethyl 329.39506 C21H19N3O 65 224 °C 0.4166
M-3 Methyl Benzyl 391.46444 C26H21N3O 66 231 °C 0.4833
M-4 Methyl 2-Cl benzyl 425.9095 C26H20ClN3O 80 220 °C 0.6353
M-5 Methyl Iso-Butyl 357.4422 C23H23N3O 79 141 °C 0.5833
I-1 H Iso-butyl 343.42164 C22H21N3O 65 131 °C 0.6857
I-2 Ethyl Iso-butyl 371.4748 C24H25N3O 70 116 °C 0.5421
I-3 Benzyl Iso-butyl 433.54418 C29H27N3O 73 129 °C 0.5483
I-4 2-Cl benzyl Iso-butyl 467.94678 C29H26ClN3O 68 125 °C 0.6140

Table 2: Analytical data of synthesized indolylchalcone

S. No. Name of the compound Wavenumber (cm-1) Chemical shift (δ ppm)
C-1 3424, 3226(NH str), 1639(C=O),1562(CH=CH Str), 7.26-7.30 (m, 4H, Ar-CH),7.5 (d, H, C-O-CH), 7.7(br, 2H, NH), 8.05 (t,3H, Ar) 8.26 (d, 1H CH=CH), 8.29(d, 2H, Ar)ppm.
C-2 3224(NH Str) 3055, 2934(CH2CH3) 2978 (-CH3-Str)1646(C=O) 1523(C=H Str) 1.55 (s,3H, CH3) 4.24(m, 2H,CH2) 7.75 (d, H, C-O-CH)7.29-7.33 (m, 4H, Ar-CH), 7.96(d, 1H, CH=CH), 8.13 (t, 3H, Ar-CH), 8.42(d,2H, Ar-CH), 10.71(br, 1H, NH) ppm


(C=H Str),

5.50(s, 2H, CH2), 7.51(d, H, C-O-CH), 7.25-7.36 (m, 8H, Ar-CH),7.81(d, 1H, CH=CH), 8.06 (t, 3H, Ar-CH), 8.23(d,2H, Ar-CH), 13.24(br, 1H, NH)ppm.
C-4 3248(NH-Str), 3120,3058(CH str, Ar),1658 (C=O),1525(C=H Str),737 (C-Cl) 5.57(s, 2H, CH2), 7.85(d, 1H, CH=CH)7.51(d, 1H, C-O-CH), 7.19-7.33 (m, 8H, Ar-CH), 8.10 (m, 4H, Ar-CH) 13.1(br, 1H, NH)ppm.
B-1 3208(NH str),3068(CH str, Ar), 1650(C=O), 1530(CH=CH Str), 1477(C=N strAr) 3.46(s, 2H, CH2), 6.03 (d, 1H, C-O-CH),7.77, (d, 1H, CH=CH) 7.19 to 7.77 (m, 8H, Ar-CH), 8.16(t,3H,Ar), 8.31(d,2H,Ar-CH), 12.87 (br, 1H, NH)ppm.
B-2 3059,3032,3007(CH str, Ar) 2980(CH2CH3), 1650(C=O),1525(CH=CH Str). 1.50(t,3H, CH3) 4.18(q, 2H,CH2), 6.06(s,2H, Ar-CH2) 7.21(d, 1H, C-O-CH), 7.60(d, 1H, CH=CH), 7.23-7.36 (m, 8H, Ar-CH) 7.85(t, 3H, Ar-CH), 7.96(d,2H, Ar-CH) ppm.
B-3 3061,3031, (CH str, Ar) 2927(-CH2), 1651(C=O) 1525(CH=CH Str). 5.47(s, 2H, CH2-Ar), 5.71 (s,2H, CH2-Ar), 6.32 (d, H, C-O-CH), 7.21(d, 1H, CH=CH), 6.39to 7.50 (m, 13H, Ar-CH), 8.04 (m,5H, Ar-CH) ppm.
B-4 3061,3031(CHAr-str,),2927(CH2Str), 1651(C=O), 1525(CH=CH str), 623(C-Cl) 5.45(s, 2H, CH2-Ar), 6.00 (s,2H, CH2-Ar), 6.75 (d, 1H, C-O-CH), 7.21(d, 1H, CH=CH), 7.24-7.63 (m, 14H, Ar-CH), 7.90-8.21(m,4H, Ar-CH) ppm.
E-1 3229(NH str), 2961(CH2CH3), 1638(C=O),1433(C=C strAr), 1.38 (s, 3H, CH3) 4.70(q, 2H,CH2), 7.40 (d, H, C-O-CH), 7.26-7.41 (m, 4H, Ar-CH),7.50 (d, 1H, CH=CH),7.89(d, 2H, Ar-CH), 8.12 (t, 3H, Ar-CH), 11.82(br, 1H, NH)ppm.
E-2 3053(CH str, Ar), 2934(CH2CH3), 2978(CH2CH3),1650(C=O), 1524 (CH=CH Str). 1.45(t, 3H, CH3), 1.52(t, 3H, CH3), 4.18(q, 2H, CH2), 4.76(q, 2H, CH2), 7.65(d, 1H, C-O-CH), 7.96 (d, H, CH=CH), 7.25-7.45 (m, 5H, Ar-CH), 8.21(t, 3H, Ar-CH) ppm.
E-3 3090, 3059, (CH str, Ar), 2980(CH2CH3), 1736(CH-C-O), 1651(C=O), 1569, 1582 (CH=CH). 1.49(s, 3H, CH3), 4.74 (q, 4H,-CH2 CH-3), 5.29(d, 2H,CH2-Ar), 7.62(d, H, C-O-CH), 715(d, 2H,Ar-CH) 7.25-7.40 (m, 8H, Ar-CH), 7.93(d, 1H, CH=CH), 8.20 (t, 3H, Ar-CH) ppm.
E-4 3441, 3057(CH str, Ar),2971(CH2CH3),2932,1654 (C=O),1084,623 (C-Cl), 592 (C=N) 1.51(t, 3H, CH3), 4.76 (q,2H,-CH2 CH3), 5.44(d, 2H,CH2-Ar),7.60
(d, H, C-O-CH),7.23-7.38 (m, 8H, Ar-CH), 7.92(d, 1H, CH=CH),
8.1-8.35(m, 4H,Ar)ppm.
CL-1 3173(N-H str),3059, 2973, 2922,1775(CH-str), 1651(C=O), 1570(C=C), 737 (C-Cl) 6.05(s, 2H, CH2-Ar), 6.40 (d, 1H, C-O-CH), 7.08 (d, 1H, CH=CH) 7.25-7.37 (m,8H, Ar) 7.94 (m, 4H, Ar-CH), 11.91(br, 1H, NH).
CL-2 3093,3059(CH str, Ar), 1647(C=O),1525(C-H Str),5 733.5(C-Cl).

1.47 (t,3H, CH3) 4.23(m, 2H, CH2) 6.06(s, 2H, Ar-CH2),6.42(d,1H, C-O-CH

7.13(d, 1H, CH=CH),), 7.23-7.54 (m, 8H, Ar-CH), 8.07(m, 4H, Ar-CH)

CL-3 3054(CH str, Ar), 3029, 1656(C=O), 1569(CH=CH, Str), 739 (C-Cl) 5.51(s, 2H, CH2-Ar), 5.54 (s,2H, CH2-Ar) 6.75 (d, 1H, C-O-CH),) 7.41(d, 1H, CH=CH), 7.23-7.34(m, 12H,Ar-CH), 8.11 (m, 5H, Ar-CH).
CL-4 3058(CH str, Ar), 1643(C=O),1522(C-H Str), 738(C-Cl) 5.58(s, 2H, CH2-Ar), 6.08 (s,2H, CH2-Ar), 6.39(d, 1H, C-O-CH), 6.89(d, 1H, CH=CH), 7.13-7.28 (m, 13H, Ar-CH), 7.96(m, 4H, Ar-CH)
A-1 3254(N-H str), 3063,2931(CH str, Ar), 1649(C=O), 1525(C=C Str). 3.63(d, 2H, CH2), 4.91 (d, 2H, N-CH2), 5.24(m, 1H, CH=CH2), 6.89(d, 1H, C-O-CH), 7.13-8.19 (m, 8H, Ar-CH) 7.77 (d,1H,CH=CH), 11.91(br, 1H, NH)ppm.
A-2 3294,3092,3052,2980, (CH str, Ar) 1650(C=O), 1527(C=C Str), 1447(N-CH2), 1.49 (s,3H, CH3) 5⋅28(d, 2H, NCH2), 5.98 (m, 1H, CH=CH2),, 5⋅14 (s, 2H, CH2), 7.63 (d, H, C-O-CH)7.94(d, 1H, CH=CH), 7.22-7.99(m, 8H, Ar-CH), 8.13(q, 2H, CH2) ppm.
A-3 3091, 3026, 3054, 2985(CH str, Ar), 1644(C=O), 1524(CH=CH Str), 4.7(s,2H,Ar-CH2), 5.18 (m, 1H, = CH2), 5⋅26 (d, 1H, =CH2) 5⋅84(d, 2H, NCH2), 7.77(d, H, C-O-CH), 8.02(d, 1H, CH=CH), 7.23-8.30(m, 13H, Ar-CH), 8.12(s, 2H, CH2) ppm.
A-4 3107,3058 (CH str, Ar), 1768(C=CH-C), 1650(C=O), 1527(CH=CH), 741(C-Cl). 4.7(s,2H,CH2), 5.95 (m, 1H, CH= CH2), 5⋅14(d, 2H, NCH2), 5⋅18(d, 1H, =CH2), 6.49 (d, H, C-O-CH), 7.00-7.69(m, 12H, Ar-CH) 7.98(d, 1H, CH=CH) ppm.
A-5 3399, 3053(CH str, Ar), 2958, 2871,(CH3CH2str) 1687,1651(C=O), 1525(CH=CH Str), 0.92(s, 6H, CH3-CH3), 2.30(m,1H,CH=CH2), 4.53(s, 2H,CH2), 4.55(s,2H,CH2), 5.95 (m, 1H, CH= CH2), 5⋅14(d, 2H, NCH2), 7.61 (d, H, C-O-CH),7.25-7.46(m, 6H, Ar-CH), 7.98(d, 1H, CH=CH), 8.12-8.25(t, 2H, Ar)ppm.
M-1 3390(N-H str), 3213, 3062(CH str, Ar), 2937, 1647 (C=O), 1566, 1521 (CH=CH Str). 3.65 (s, 3H, CH3), 7.34-7.53 (m, 4H, Ar-CH), 7.74 (d, H, C-O-CH) 8.02(d, 2H, Ar), 8.10(t, 3H, Ar), 8.25(d, 1H,CH=CH) 13.18 (br, 1H, NH)ppm.
M-2 3430,3215, 3047(CH str, Ar),3101, 1648(C=O), 1528(CH=CH str), 1462(N-CH2), 592 (C=CH) 1.44(t,3H,CH3), 3.53(s,3H, CH3),4.77(d,2H,CH2), 7.53-7.55(m, 4H, Ar-CH), 7.66(d, 1H, C-O-CH), 7.88 (d, H, CH=CH),8.01-8.11 (m, 4H, Ar-CH) ppm.
M-3 3093, 3055(CH str, Ar),2931(R-CH3,str), 1647(C=O), 1527(CH=CH Str). 3.89(s,3H,CH3), 7.31-7.48 (m, 8H, Ar-CH), 7.59(d, 1H, C-O-CH), 7.93 (d, H, CH=CH), 7.95-8.01 (m, 3H, Ar-CH), 8.11(d, 2H, Ar-CH)ppm.
M-4 3402, 3202, 3286, 3096(CH str, Ar), 3055, 2987(R-CH3), 1646(C=O),1527(CH=CH Str), 1441(N-CH2), 1064, 633(C-Cl) 3.89(s, 3H, CH3), 6.08(s, 2H, CH2), 6.38 (d, H, C-O-CH),7.24 (d, 1H, CH=CH), 7.35-7.49 (m, 8H, Ar-CH), 7.94-8.13(m, 4H, Ar-CH) ppm.
M-5 3104, 3053(CH str, Ar), 2934, 1643(C=O), 1524(CH-CH Str). ; 0.90(s, 6H, CH3-CH3), 1.6 (s,3H, CH3), 2.21(m, 1H, CH=CH2), 4.57 (q, 2H,CH2), 7.68(d, 1H, C-O-CH), 7.87 (d, H, CH=CH), 7.31-7.36 (m, 4H, Ar-CH), 8.02-8.11(m, 3H, Ar-CH) ppm.
I-1 3176(NH-Str), 2955, 1898(CH str, Ar), 1634(C=O), 1519(CH=CH Str). 1.35(s,6H,CH3-CH3), 2.27(m,1H,CH=CH2), 4.57(d, 2H,CH2), 7.61 (d, H, C-O-CH), 7.87(d, 1H, CH=CH) 7.18-7.65 (m, 5H, Ar-CH), 7.96-8.16 (m, 4H, Ar-CH),11.86(br, 1H, NH) ppm.
I-2 3368, 3107, 3053(CH str, Ar),1642(C=O),1522(CH=CH str) 0.92(s, 6H, CH3-CH3), 1.50 (t,3H, CH3-CH2), 2.36(m, 1H,CH=CH2), 4.20 (q, 2H,CH2),4.56(d, 2H, CH2), 7.46 (d, H, C-O-CH), 7.25-7.40 (m, 5H, Ar-CH), 7.95 (d, 1H, CH=CH), 8.11-8.21(m, 3H,Ar-CH) ppm.
I-3 3083,3056, 3029 (Ar-CH str), 1646(C=O), 1526(CH=CH Str). 0.65(d, 6H, CH3-CH3), 2.02(m, 1H, CH=CH2), 4.39(d, 2H, CH2), 5.3(s, 2H, CH2-Ar),7.09-7.19(m, 9H, Ar-CH), 7.27(d, 1H, C-O-CH), 7.72(d, 1H, CH=CH), 7.77(d, 1H, Ar-CH), 7.97-8.22(t, 3H, Ar-CH)ppm.
I-4 3294,3090(CH=CHAr),1650(C=O), 1525(CH=CH str), 625(C-Cl) 0.73(d, 6H, CH3-CH3), 2.05(m, 1H,CH=CH2), 4.27(d, 1H, CH2), 5.47(s, 2H, CH2),6.77(d, 1H, C-O-CH), 7.10-7.40(m, 8H, Ar-CH), 7.77(d, 1H, CH=CH), 7.80-8.21(m, 4H,Ar-CH) ppm.

Table 3: The % inhibitory effect of antiangiogenic activity N-1 substituted indolylchalconehybrids and in a CAM assay in a concentration of 0.1 µM/egg

Code of compound

No% avascular CAM/Total


Std. error of deviation Code of compound

No % avascular CAM/Total


Std. error of deviation Code of compound

No% avascular CAM/Total


Std. error of deviation
B-1 14/20(70%) 1.826 CL-3 2/20 (10%) 0.5774 A-1 6/20(30%) 1.390
B-2 13/20(65%) 1.390 CL-4 7/20(35%) 1.390 A-2 5/20(25%) 0.5774
B-3 8/20(40%) 1.390 E-1 18/20(90%) 0.5774 A-3 4/20(20%) 1.390
B-4 7/20(35%) 1.390 E-2 19/20(95%) 0.2582 A-4 3/20 (15%) 13.336
C-1 18/20(90%) 0.5774 E-3 15/20(75%) 1.155 A-5 14/20(70%) 0.8165
C-2 17/20(85%) 1.317 E-4 6/20(30%) 1.390 M-1 15/20(75%) 1.155
C-3 16/20(80%) 0.9309 I-1 16/20(80%) 0.9309 M-2 8/20(40%) 6.731
C-4 15/20(75%) 1.155 I-2 18/20(90%) 0.8165 M-3 7/20(35%) 0.8165
CL-1 17/20(85%) 1.317 I-3 7/20(35%) 0.8165 M-4 6/20(30%) 1.390
CL-2 5/20(25%) 1.317 I-4 6/20(30%) 1.390 M-5 16/20(80%) 0.7303

*SD (n=6)

(a) C-2 at dose 2.5 µM (b) C-2at dose 5 µM (c) C-2at dose10 µM
(d) I-2at dose 2.5 µM (e) I-2at dose 5 µM (f) I-2at dose10 µM
(g)-Ve Control (h) DMSO (i)+Ve Control

Fig. 1A: Effect of dose-dependent assay of synthesized indolylchalcone hybrids of benzimidazole on in vivo CAM angiogenesis: CAM were treated with different series of dose such as 2.5 µM, 5 µM and 10 µM treatment groups indolyl chalcone hybrids at day 8th, the percentage of inhibition of blood vesel formation, compared to untreated conrol, was determined. (a to i) Result suggests that indolyl chalcone hybrids compound code C-2 and I-2 inhibited angiogenesis expressed as (⁄p =<0.001 vs DMSO)

Fig. 1A: Effect of dose-dependent assay of synthesized indolylchalcone hybrids of benzimidazole on in vivo CAM angiogenesis


We found during our CAM Assay Study that there was noticeable decrease in number of blood vessels as compared with the negative control group, as shown in fig. 1A. Furthermore, One-way ANOVA was used to compare the means of groups. Results showed that the different series of dose such as 2.5µM, 5µM and 10µM treatment groups exhibited varying response based on their %inhibition on blood vessel count (fig. 1B). These newly synthesized indolylchalcone hybrids were statistically significant at P = 0.05. Considering this we can draw an inference that amongst 30 synthesized compounds 11 compounds of indolylchalcone hybrids has potent anti-angiogenic properties.


We have developed an inexpensive, simple, and eco-friendly synthesis of N-alkyl derivatives of two fused heterocyclic compounds of indole-3-carboxaldehyde and 2-acetylbenzimidazole. The 11 compounds of indolylchalcones as shown in graph possess very good dose-dependent anti-angiogenic activities. The N-1 substitution is valid on benzimidazole such as ethyl, benzyl and also 2-Cl substituted group on phenyl shows significant potent anticancer activity. The potency of anti-angiogenetic activity shows that methyl>Ethyl>Cl-benzyl>Benzyl>Isobutyl.

The present study suggests that N-1 substituted Indolylchalcone hybrids of 2-acetyl benzimidazole might be promising analogs of angiogenesis inhibitors to manage the uncontrolled neovascularization occurring during malignant tumor development.


Authors take this opportunity to thank UGC-BSR for funding this project. Authors also thankful to the Director of SAIF of Punjab University, Chandigarh for recording IR–spectra and NMR-spectra of synthesized compounds.


All the authors have contributed equally to this study.


Declared none


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