Synthesis of coumarin derivatives containing pyrazole and indenone rings as potent antioxidant and antihyperglycemic agents

R. Kenchappa; Yadav D. Bodke; A. Chandrashekar; M. Aruna Sindhe; S.K. Peethambar

doi:10.1016/j.arabjc.2014.05.029

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Original article

10 (

2_suppl

); S3895-S3906

doi:

10.1016/j.arabjc.2014.05.029

Synthesis of coumarin derivatives containing pyrazole and indenone rings as potent antioxidant and antihyperglycemic agents

R. Kenchappa^{^a}, Yadav D. Bodke^{^a,⁎}, A. Chandrashekar^{^a,^b}, M. Aruna Sindhe^{^a}, S.K. Peethambar^{^c}

a Department of P.G. Studies and Research in Industrial Chemistry, School of Chemical Science, Jnana Sahyadri, Kuvempu University, Shankaraghatta-577451, Shivamogga, Karnataka, India

b Biocon Ltd., 20th KM, Hosur Road, Electronic City, Bangalore 560 100, Karnataka, India

c Department of P.G. Studies and Research in Biochemistry, Kuvempu University, Shankaraghatta-577451, Shivamogga, Karnataka, India

⁎Corresponding author. Tel.: +91 8282256228; fax: +91 8282256262. ydbodke@gmail.com (Yadav D. Bodke)

Received: 2013-8-8, Accepted: 2014-5-31, Published: 2014-05-01

Disclaimer:
This article was originally published by Elsevier and was migrated to Scientific Scholar after the change of Publisher.

Peer review under responsibility of King Saud University.

Abstract

The title compounds (5a–l) were synthesized by Claisen–Schmidt condensation of 3-(6-substituted-2-oxo-2H-chromen-3-yl)-1-(4-substituted)-1H-pyrazole-4-carbaldehyde (4a–l) derivatives with 5,6-dimethoxy-2,3-dihydro-1H-inden-1-one at reflux temperature. The synthesized compounds were screened for antioxidant, in vitro and in vivo antihyperglycemic activities.

Abstract

The present work describes the synthesis of 6-substituted-3-(1-(4-substituted)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene)methyl)-1H-pyrazol-3-yl)-2H-chromen-2-one derivatives (5a–l) by Claisen–Schmidt condensation of 3-(6-substituted-2-oxo-2H-chromen-3-yl)-1-(4-substituted)-1H-pyrazole-4-carbaldehyde derivatives (4a–l) with 5,6-dimethoxy-2,3-dihydro-1H-inden-1-one at reflux temperature for 8 h. The synthesized compounds were screened for in vitro antioxidant activity. Compounds 5e and 5g showed promising DPPH radical scavenging activity with IC₅₀ 54.14–56.19 μg/mL, ferrous ion chelating ability with IC₅₀ 53.75–56.89 μg/mL and reductive capability with IC₅₀ 58.01–62.57 μg/mL. The selected compounds were also tested for in vivo antihyperglycemic activity against Streptozotocin–nicotinamide induced Adult Wistar rats. Compounds 5e and 5g showed significant decrease in glucose concentration (115 and 138 mg/dL) with the dose of 100 mg/kg.

Keywords

Coumarin

Pyrazole

Indenone

Antioxidant

Antihyperglycemic

Streptozotocin–nicotinamide

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1 Introduction

Human bodies are protected from oxidative stress by natural enzymatic and non-enzymatic antioxidant defensive system, whose capacity is affected by age, diet, and health status of the individual (Chun et al., 2003). Therefore, only endogenous antioxidant defenses are not absolutely efficient. Dietary antioxidants are required to diminish the cumulative effects of oxidative damage due to excess ROS (reactive oxygen species) that remains in our system (Lim and Murtijaya, 2007).These free radicals may oxidize nucleic acids, proteins, or lipids which occur through a chain reaction and may form potentially toxic end products (Guo et al., 2005; Chunhuan et al., 2010). Therefore, oxidation can cause not only deterioration of foodstuffs but also harm living organisms.

When free radicals overcome our body’s ability to neutralize them, they attack the body itself. As a result free radicals disturb the action of cellular DNA, which directs key cellular activities. Cells with damaged DNA stagnate and are prone to developing cancer and growths. This kind of damage also accelerates the aging process and diabetes (Hyun et al., 2006) directly causing wrinkles and age spots and severely taxing the immune system. They may also responsible for other diseases such as cardiovascular diseases (Singh and Jialal, 2006), neural disorders (Sas et al., 2007), Alzheimer’s disease (Smith et al., 2000), alcohol-induced liver disease (Arteel et al., 2006). Therefore, the searches for new antioxidants have received much attention.

Diabetes mellitus is a chronic and multifactorial disease characterized by a high level of blood glucose and impaired insulin action (Neogi et al., 2003; Saltiel et al., 2001). It generally results in decrease in insulin secretion that translates into impaired carbohydrate (glucose) use, which is the characteristic feature of diabetes mellitus. This, results in hyperglycemia in the blood, which could potentially cause serious debilitating conditions including cardiovascular morbidity, nerve damage, and mortality (Gegory et al., 2001). Non-insulin dependent diabetes mellitus (NIDDM) accounts for approximately 80–90% of all diabetes cases (King and Rewers, 1993). Although significant progress has been achieved in the treatment of type-2 diabetes mellitus (T2DM) over last decade, the biochemical and molecular mechanisms underlying insulin resistance have not been clearly indicated (Kus et al., 2011; Sangle et al., 2012). Therefore, it is essential to control blood glucose levels during the early stages of the disease.

Coumarins, a group of benzopyrone derivatives (Saija et al., 1995), have been recognized as one of the largest and most widespread class of plant constituents occurring throughout the plant kingdom (Es-Safi et al., 2007; Anju et al., 2016). Moreover coumarin containing other heterocyclic compounds have been found to exhibit various pharmacological activities including spasmolytic, antioxidant, anticoagulant, antiviral (Stancho et al., 2008; Jovanovic et al., 1994), antimicrobial (Chimenti et al., 2010; Vijaya Laxmi et al., 2013), analgesic (Khan et al., 2005; Ghate et al., 2005), antitubercular (Karali et al., 2002), antimalarial (Gajanan et al., 2010), antidiabetic (Gireesh et al., 2016) and antitumor (Xin-Hua et al., 2012) activities.

Pyrazoline derivatives have exhibited various biological activities such as antioxidant, antimicrobial (Ahmad et al., 2010; Padmaja et al., 2011; Mamta et al., 2015 ), analgesic, anti-inflammatory (Isloor et al., 2000), anti-hyperglycemic and sedative-hypnotic activities (Braulio et al., 1999; Rizk et al., 2011).

Prompted by the above results and in continuation of our research work on biologically active compounds (Kenchappa et al., 2013,2017) we synthesized coumarin derivatives containing pyrazole and indenone moiety and screened them for antioxidant and antidiabetic activities.

2 Materials and methods

2.1

2.1 Chemistry

5,6-Dimethoxyindenone with 98% purity was purchased from Sigma Aldrich Company. Melting points were recorded on an electro thermal melting point apparatus and are uncorrected. ¹H and ¹³CNMR spectra were recorded on a Bruker 400 MHz spectrometer and chemical shifts are shown in δ values (ppm) with tetramethylsilane (TMS) as internal standard. LC–MS was obtained using an Agilent LC/MS instrument (Electronic Ionization method). The FT-IR spectra were taken in KBr pellets (100 mg) using a Shimadzu FT-IR spectrophotometer. Column chromatography was performed using silica gel (230–400 mesh), silica gel GF254 plates from Merck were used for TLC and spots were identified either by UV or dipping the plates in potassium permanganate solution.

2.1.1

2.1.1 General procedure for the synthesis of 3-(6-substituted-2-oxo-2H-chromen-3-yl)-1-(4-substituted)-1H-pyrazole-4-carbaldehyde derivatives (4a-l)

To the cold solution of DMF (1.0 mL, 0.014 mol), POCl₃ (1.3 mL 0.014 mol) was added drop wise for half an hour by maintaining the temperature at 0–5 °C. To this solution, 3-[(1E)-1-(2-phenylhydrazinylidene) ethyl]-2H-chromen-2-one (0.0035 mol) (3a–l) was added and the reaction mixture was stirred. After complete addition, the stirring was continued for 30 min, at 0–5 °C and then the temperature was raised to 80 °C for about 6 h. Completion of reaction was confirmed by TLC, reaction mixture was poured into crushed ice and neutralized by 10% NaOH solution. The solid precipitated out was filtered, dried and recrystallized from ethanol.

The formation of compounds 4a (m.p.; 183–186 °C) and 4b (m.p.;−185 to 187 °C) was confirmed by comparing its melting points with the literature value (Vijaya Laxmi et al., 2013).

2.1.2

2.1.2 1-(4-Chlorophenyl)-3-(2-oxo-2H-chromen-3-yl)-1H-pyrazole-4-carbaldehyde (4c)

Yield 70%; m.p. 189–192 °C; Mol. Formula: C₁₉H₁₁ClN₂O₃; IR (KBr, cm⁻¹): 1714 (C⚌O), 1675 (CHO), 1625 (C⚌N), 1590 (C⚌C); ¹H NMR (DMSO-d₆, 400 MHz) δ: 7.30–8.10 (m, 9H), 9.43 (s, 1H), 10.22 (s, 1H).MS: m/z = 370 [M⁺], 372 [M+2].

2.1.3

2.1.3 3-(6-Bromo-2-oxo-2H-chromen-3-yl)-1-(4-chlorophenyl)-1H-pyrazole-4-carbaldehyde (4d)

Yield 75%; m.p. 189–192 °C; Mol. Formula: C₁₉H₁₀BrClN₂O₃; IR (KBr, cm⁻¹): 1700 (C⚌O), 1671 (CHO), 1621 (C⚌N), 1592 (C⚌C); ¹H NMR (DMSO-d₆, 400 MHz) δ:7.31–8.15 (m, 8H), 9.45 (s, 1H), 10.23 (s, 1H).MS: m/z = 430 [M+], 432 [M+2], 434 [M+4].

2.1.4

2.1.4 1-(4-Methylphenyl)-3-(2-oxo-2H-chromen-3-yl)-1H-pyrazole-4-carbaldehyde (4e)

Yield 72%; m.p. 194–197 °C; Mol. Formula: C₂₀H₁₄N₂O₃; IR (KBr, cm⁻¹): 1710 (C⚌O), 1689 (CHO), 1625 (C⚌N), 1588 (C⚌C); ¹HNMR (DMSO-d₆, 400 MHz) δ: 3.20 (s, 3H, CH₃), 7.20–7.93 (m, 9H), 9.40 (s, 1H), 9.97 (s, 1H).MS: m/z = 330 [M⁺].

2.1.5

2.1.5 3-(6-Bromo-2-oxo-2H-chromen-3-yl)-1-(4-methylphenyl)-1H-pyrazole-4-carbaldehyde (4f)

Yield 68%; m.p.199–202 °C; Mol. Formula: C₂₀H₁₃BrN₂O₃; IR (KBr, cm⁻¹): 1715 (C⚌O), 1691 (CHO), 1621 (C⚌N), 1585 (C⚌C); ¹HNMR (DMSO-d₆, 400 MHz) δ: 3.25(s, 3H, CH₃), 7.22–8.10 (m, 8H), 9.43 (s, 1H), 10.10 (s, 1H). MS: m/z = 408 [M⁺], 410 [M+2].

2.1.6

2.1.6 1-(4-Methoxyphenyl)-3-(2-oxo-2H-chromen-3-yl)-1H-pyrazole-4-carbaldehyde (4g)

Yield 77%; m.p. 177–180 °C; Mol. Formula: C₂₀H₁₄N₂O₄; IR (KBr, cm⁻¹): 1718 (C⚌O), 1685(CHO), 1618 (C⚌N), 1595 (C⚌C); ¹HNMR (DMSO-d₆, 400 MHz) δ: 3.88 (s, 3H, OCH₃), 7.13–8.06 (m, 9H), 9.42 (s, 1H), 9.94 (s, 1H).MS: m/z = 346.33 [M⁺].

2.1.7

2.1.7 3-(6-Bromo-2-oxo-2H-chromen-3-yl)-1-(4-methoxyphenyl)-1H-pyrazole-4-carbaldehyde (4h)

Yield 78%; m.p. 181–183 °C; Mol. Formula: C₂₀H₁₃BrN₂O₄; IR (KBr, cm⁻¹): 1712 (C⚌O), 1687 (CHO), 1620 (C⚌N),1591 (C⚌C); ¹H NMR (DMSO-d₆, 400 MHz) δ: 3.35(s, 3H, OCH₃), 7.30–8.13 (m, 8H), 9.43 (s, 1H), 10.21 (s, 1H). MS: m/z = 424 [M⁺], 426 [M+2].

2.1.8

2.1.8 1-(4-Nitrophenyl)-3-(2-oxo-2H-chromen-3-yl)-1H-pyrazole-4-carbaldehyde (4i)

Yield 75%; m.p. 189–192 °C; Mol. Formula: C₁₉H₁₁N₃O₅; IR (KBr, cm⁻¹): 1721 (C⚌O), 1681 (CHO), 1620 (C⚌N), 1599 (C⚌C); ¹H NMR (DMSO-d₆, 400 MHz) δ: 6.90–8.0 (m, 9H), 9.43 (s, 1H), 10.01 (s, 1H).MS: m/z = 361 [M⁺].

2.1.9

2.1.9 3-(6-Bromo-2-oxo-2H-chromen-3-yl)-1-(4-nitrophenyl)-1H-pyrazole-4-Carbaldehyde (4j)

Yield 76%; m.p.187–189 °C; Mol. Formula: C₁₉H₁₀BrN₃O₅; IR (KBr, cm⁻¹): 1719 (C⚌O), 1684 (CHO), 1630 (C⚌N), 1625, 1520 (N—O), 1585 (C⚌C); ¹HNMR (DMSO-d₆, 400 MHz) δ: 6.95–8.05 (m, 8H), 9.48 (s, 1H), 10.15 (s, 1H).MS: m/z = 439 [M⁺], 441 [M+2].

2.1.10

2.1.10 1-(4-Fluorophenyl)-3-(2-oxo-2H-chromen-3-yl)-1H-pyrazole-4-carbaldehyde (4k)

Yield 72%; m.p. 161–164 °C; Mol. Formula: C₁₉H₁₁FN₂O₃; IR (KBr, cm⁻¹): 1709 (C⚌O), 1680 (CHO), 1611 (C⚌N), 1603 (C⚌C); ¹H NMR (DMSO-d₆, 400 MHz) δ: 7.13–8.06 (m, 9H), 9.45 (s, 1H), 9.98 (s, 1H).MS: m/z = 334 [M⁺].

2.1.11

2.1.11 3-(6-Bromo-2-oxo-2H-chromen-3-yl)-1-(4-fluorophenyl)-1H-pyrazole-4-carbaldehyde (4l)

Yield 68%; m.p. 166–168 °C; Mol. Formula: C₁₉H₁₀BrFN₂O₃; IR (KBr, cm⁻¹): 1705 (C⚌O), 1682 (CHO), 1618 (C⚌N), 1589 (C⚌C); ¹HNMR (DMSO-d₆, 400 MHz) δ: 7.10–8.0 (m, 8H), 9.42 (s, 1H), 10.12 (s, 1H).MS: m/z = 412 [M⁺], 414 [M+2].

2.1.12

2.1.12 Synthesis of 3-(4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-4-substituted-phenyl-1H-pyrazol-3-yl)-6-substituted-2H-chromen-2-onederivatives (5a–l)

The mixture of 5,6-dimethoxy-2,3-dihydro-1H-inden-1-one (0.5 g 0.002 mol) and 3-(6-substituted-2-oxo-2H-chromen-3-yl)-1-(4-substituted)-1H-pyrazole-4-carbaldehyde (4a–l) (0.002 mol) was stirred in sodium methoxide solution under reflux condition for 8 h. and completion of the reaction was confirmed by TLC. The reaction mixture was poured into cold water and neutralized with dilute HCl. The solid separated was filtered, dried and recrystallized with ethanol to furnish the title compounds.

2.1.13

2.1.13 3-(4-((Z)-(5,6-Dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-phenyl-1H-pyrazol-3-yl)-2H-chromen-2-one (5a)

Yield 73%; m.p. 224–227 °C; Mol. Formula: C₃₀H₂₂N₂O₅; IR (KBr, cm⁻¹): 3063 (Ar—H), 1705 (coumarin C⚌O), 1678 (indenone C⚌O), 1619 (C⚌N), 1596 (C⚌C); ¹HNMR (DMSO-d₆, 400 MHz) δ: 3.28 (3H, s, OCH₃), 3.41 (3H, s, OCH₃), 4.75 (2H, s, CH₂), 7.39–8.89 (12 H, m, Ar—H), 8.90 (1H, s, CH-methine), 9.80 (1H, s, C⚌C—H), ¹³CNMR (CDCl₃) δ: 48.9 (CH₂), 57.8 (2-OCH₃), 102.2, 103.1, 108.0, 110.5 (2C), 112.1, 114.3, 118.4, 119.5, 122.3, 123.2, 126.5, 129.1, 130.1 (2C), 131.2, 132.8, 138.5, 139.8, 142.9, 148.4, 150.1, 152.4, 159.6, 161.5, 168.2 (coumarin C⚌O), 171.5 (indenone C⚌O).MS: m/z 490.50 [M⁺].

2.1.14

2.1.14 6-Bromo-3-(4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-phenyl-1H-pyrazol-3-yl)-2H-chromen-2-one (5b)

Yield 71%; m.p. 221–223 °C; Mol. Formula: C₃₀H₂₁BrN₂O₅; IR (KBr, cm⁻¹): 3060 (Ar—H), 1707 (coumarin C⚌O), 1670 (indenone C⚌O), 1610 (C⚌N), 1590 (C⚌C); 684 (C—Br); ¹H NMR (DMSO-d₆, 400 MHz) δ: 3.00 (3H, s, OCH₃), 3.09 (3H, s,OCH₃), 4.33 (2H, s, CH₂), 7.09–8.53 (11 H, m, Ar—H), 8.56 (1H, s, CH methine), 9.68 (1H, s, C⚌C—H),¹³CNMR (CDCl₃) δ: 49.0 (CH₂), 57.9 (2-OCH₃), 103.2, 104.1, 109.0, 111.6 (2C), 113.2, 116.3, 119.3, 121.5, 123.5, 125.5, 128.6, 131.1, 132.1 (2C), 133.1, 135.8, 139.5, 141.8, 143.6, 150.4, 153.1, 155.4, 161.6, 165.5, 169.2 (coumarin C⚌O), 175.5 (indenone C⚌O).MS: m/z 568 [M⁺], 570 [M+2].

2.1.15

2.1.15 3-(1-(4-Chlorophenyl)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene)methyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5c)

Yield 81%; m.p. 233–225 °C; Mol. Formula: C₃₀H₂₁ClN₂O₅; IR (KBr, cm⁻¹): 3052 (Ar—H), 1703 (coumarin C⚌O), 1675 (indenone C⚌O), 1615 (C⚌N), 1604 (C⚌C); 680 (C—Cl); ¹H NMR (DMSO-d₆, 400 MHz) δ: 3.20 (3H, s, OCH₃), 3.39 (3H, s,OCH₃), 4.65 (2H, s, CH₂), 7.31–8.82 (11 H, m, Ar—H), 8.82 (1H, s, CH methine), 9.69 (1H, s, C⚌C—H),¹³CNMR (CDCl₃) δ: 45.0 (CH₂), 57.9 (2-OCH₃), 103.6, 104.7, 109.7, 112.0 (2C), 113.7, 117.1, 119.5, 122.0, 123.7, 126.0, 128.9, 131.5, 133.1 (2C), 134.1, 136.2, 139.5, 142.0, 144.2, 151.4, 153.1, 156.4, 161.6, 166.5, 169.2 (coumarin C⚌O), 176.5 (indenone C⚌O).MS: m/z 524[M⁺], 526 [M+2].

2.1.16

2.1.16 6-Bromo-3-(1-(4-chlorophenyl)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5d)

Yield 82%; m.p. 241–243 °C; Mol. Formula: C₃₀H₂₀BrClN₂O₅; IR (KBr, cm⁻¹): 3060 (Ar—H), 1713 (coumarin C⚌O), 1678 (indenone C⚌O), 1610 (C⚌N), 1605 (C⚌C); 687 (C—Br), 680 (C—Cl); ¹H NMR (DMSO-d₆, 400 MHz) δ: 3.24 (3H, s, OCH₃), 3.37 (3H, s,OCH₃), 4.71 (2H, s, CH₂), 7.35–8.85 (10H, m, Ar—H), 8.86 (1H, s, CH methine), 9.74 (1H, s, C⚌C—H), ¹³CNMR (CDCl₃) δ: 45.0 (CH₂), 57.9 (2-OCH₃), 102.6, 104.7, 109.7, 112.0 (2C), 114.7, 116.1, 120.5, 122.0, 123.7, 125.0, 127.9, 130.0, 132.5 (2C), 134.1, 136.7, 139.5, 142.0, 144.2, 150.4, 153.1, 156.4, 162.2, 165.1, 169.0 (coumarin C⚌O), 175.5 (indenone C⚌O).MS: m/z 602[M⁺], 604 [M+2], 606 [M+4].

2.1.17

2.1.17 3-(4-((Z)-(5,6-Dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-p-tolyl-1H-pyrazol-3-yl)-2H-chromen-2-one (5e)

Yield 72%; m.p. 242–245 °C; Mol. Formula: C₃₁H₂₄N₂O₅; IR (KBr, cm⁻¹): 3060 (Ar—H), 1703 (coumarin C⚌O), 1667 (indenone C⚌O), 1615 (C⚌N), 1605 (C⚌C); ¹H NMR (DMSO-d₆, 400 MHz) δ:2.52 (3H, s, CH₃)3.22 (3H, s, OCH₃), 3.35 (3H, s,OCH₃), 4.72 (2H, s, CH₂), 7.36–8.86 (11H, m, Ar—H), 8.88 (1H, s, CH methine), 9.73 (1H, s, C⚌C—H), ¹³CNMR (CDCl₃) δ: 25.6 (CH₃), 43.0 (CH₂), 57.1 (2-OCH₃), 103.2, 105.2, 108.6, 112.0 (2C), 114.7, 116.1, 119.5, 122.0, 123.5, 125.0, 127.9, 130.0, 132.2 (2C), 134.1, 136.5, 138.2, 142.3, 144.7, 151.1, 154.1, 157.1, 161.4, 165.1, 168.0 (coumarin C⚌O), 174.2 (indenone C⚌O). MS: m/z 504 [M⁺].

2.1.18

2.1.18 6-Bromo-3-(4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-p-tolyl-1H-pyrazol-3-yl)-2H-chromen-2-one (5f)

Yield 68%; m.p. 251–253 °C; Mol. Formula: C₃₁H₂₃BrN₂O₅; IR (KBr, cm⁻¹): 3068 (Ar—H), 1710 (coumarin C⚌O), 1669 (indenone C⚌O), 1617 (C⚌N), 1600 (C⚌C); 687 (C—Br); ¹H NMR (DMSO-d₆, 400 MHz) δ: 2.50 (3H, s, CH₃)3.25 (3H, s, OCH₃), 3.38 (3H, s,OCH₃), 4.72 (2H, s, CH₂), 7.35–8.85 (10H, m, Ar—H), 8.87 (1H, s, CH methine), 9.77 (1H, s, C⚌C—H),¹³CNMR (CDCl₃) δ: 24.3 (CH₃), 44.0 (CH₂), 57.6 (2-OCH₃), 102.2, 105.2, 107.6, 112.0 (2C), 113.7, 116.1, 119.5, 121.0, 123.5, 125.0, 127.0, 130.0, 132.2 (2C), 134.1, 136.5, 138.2, 142.3, 144.7, 152.1, 154.1, 157.1, 162.4, 165.1, 168.9 (coumarin C⚌O), 175.3 (indenone C⚌O). MS: m/z 582 [M⁺], 584 [M+2].

2.1.19

2.1.19 3-(4-((Z)-(5,6-Dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-(4-methoxy phenyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5g)

Yield 74%; m.p. 228–231 °C; Mol. Formula: C₃₁H₂₄N₂O₆; IR (KBr, cm⁻¹): 2989 (Ar—H), 1712 (coumarin C⚌O), 1687 (indenone C⚌O), 1624 (C⚌N), 1603 (C⚌C); ¹H NMR (DMSO-d₆, 400 MHz) δ: 2.78 (3H, s, OCH₃)3.30 (3H, s, OCH₃), 3.40 (3H, s,OCH₃), 4.34 (2H, s, CH₂), 7.40–8.90(11H, m, Ar—H), 8.90 (1H, s, CH methine), 9.82 (1H, s, C⚌C—H), ¹³CNMR (CDCl₃) δ: 25.3 (OCH₃), 45.0 (CH₂), 58.6 (2-OCH₃), 103.2, 105.2, 107.8, 112.0 (2C), 114.7, 116.1, 119.0, 121.0, 122.9, 125.0, 127.0, 129.5, 132.2 (2C), 134.1, 136.3, 139.2, 142.3, 146.7, 152.1, 155.1, 157.1, 161.4, 166.1, 168.9 (coumarin C⚌O), 174.3 (indenone C⚌O).MS: m/z 520 [M⁺].

2.1.20

2.1.20 6-Bromo-3-(4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene)methyl)-1-(4-methoxy phenyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5h)

Yield 76%; m.p. 234–237 °C; Mol. Formula: C₃₁H₂₃BrN₂O₆; IR (KBr, cm⁻¹): 2995 (Ar—H), 1709 (coumarin C⚌O), 1681 (indenone C⚌O), 1621 (C⚌N), 1600 (C⚌C), 687 (C—Br); ¹H NMR (DMSO-d₆, 400 MHz) δ: 2.76 (3H, s, CH₃) 3.29 (3H, s, OCH₃), 3.42 (3H, s,OCH₃), 4.76 (2H, s, CH₂), 7.42–8.91 (10H, m, Ar—H), 8.92 (1H, s, CH methine), 9.84 (1H, s, C⚌C—H), ¹³CNMR(CDCl₃) δ: 25.5 (OCH₃), 46.0 (CH₂), 57.2 (2-OCH₃), 102.3, 105.2, 106.8, 111.0 (2C), 114.7, 116.1, 118.0, 121.0, 123.5, 125.0, 127.0, 129.3, 132.0 (2C), 134.2, 136.9, 139.2, 142.5, 145.6, 151.1, 155.2, 158.4, 161.4, 165.4, 169.0 (coumarin C⚌O), 175.3 (indenone C⚌O). MS: m/z 598 [M⁺], 600 [M+2].

2.1.21

2.1.21 3-(4-((Z)-(5,6-Dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-(4-nitro phenyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5i)

Yield 81%; m.p. 267–270 °C; Mol. Formula: C₃₀H₂₁N₃O₇; IR (KBr, cm⁻¹): 2995 (Ar—H), 1709 (coumarin C⚌O), 1676 (indenone C⚌O), 1626 (C⚌N), 1598 (C⚌C), 1553 (NO₂); ¹H NMR (DMSO-d₆, 400 MHz) δ: 3.28 (3H, s, OCH₃), 3.40 (3H, s,OCH₃), 4.75 (2H, s, CH₂), 7.44–8.92 (11H, m, Ar—H), 8.93 (1H, s, CH methine), 9.83 (1H, s, C⚌C—H), ¹³CNMR (CDCl₃) δ: 46.0 (CH₂), 58.1 (2-OCH₃), 103.4, 105.2, 107.8, 111.0 (2C), 113.7, 116.1, 118.0, 121.3, 123.5, 125.3, 127.0, 129.3, 131.4 (2C), 134.2, 137.4, 139.2, 142.5, 145.3, 152.3, 155.2, 157.3, 160.4, 164.4, 168.0 (coumarin C⚌O), 176.3 (indenone C⚌O). MS: m/z 535 [M⁺].

2.1.22

2.1.22 6-Bromo-3-(4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-(4-nitro phenyl)-1H-pyrazol-3-yl)-2H-chromen-2-one(5j)

Yield 82%; m.p. 278–28 °C; Mol. Formula: C₃₀H₂₀BrN₃O₇; IR (KBr, cm⁻¹): 3016 (Ar—H), 1703 (coumarin C⚌O), 1686 (indenone C⚌O), 1620 (C⚌N), 1596 (C⚌C), 1552 (NO₂); ¹H NMR (DMSO-d₆, 400 MHz) δ:2.97 (3H, s, OCH₃), 3.06 (3H, s,OCH₃), 4.30 (2H, s, CH₂), 7.06–8.49(10H, m, Ar—H), 8.50 (1H, s, CH methine), 9.72 (1H, s, C⚌C—H),¹³CNMR (CDCl₃) δ: 45.0 (CH₂), 57.1 (2-OCH₃), 102.4, 105.2, 108.2, 111.0 (2C), 113.7, 115.5, 118.0, 121.4, 123.5, 125.3, 128.0, 131.5, 133.4 (2C), 136.2, 138.4, 141.2, 144.5, 148.3, 152.3, 155.2, 158.3, 162.4, 165.4, 169.0 (coumarin C⚌O), 175.3 (indenone C⚌O).MS: m/z 613[M⁺], 615 [M+2].

2.1.23

2.1.23 3-(1-(4-Fluorophenyl)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene)methyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5k)

Yield 77%; m.p. >300 °C; Mol. Formula: C₃₀H₂₁FN₂O₅; IR (KBr, cm⁻¹): 3016 (Ar—H), 1705 (coumarin C⚌O), 1684 (indenone C⚌O), 1620 (C⚌N), 1596 (C⚌C), 1552 (NO₂); ¹H NMR (DMSO-d₆, 400 MHz) δ: 3.26 (3H, s, OCH₃), 3.42 (3H, s,OCH₃), 4.70 (2H, s, CH₂), 7.33–8.83 (11 H, m, Ar—H), 8.85 (1H, s, CH methine), 9.75 (1H, s, C⚌C—H),¹³CNMR (CDCl₃) δ: 43.0 (CH₂), 56.2 (2-OCH₃), 103.4, 106.2, 109.3, 111.0 (2C), 114.7, 116.5, 118.0, 120.4, 122.5, 125.3, 127.0, 130.5, 133.4 (2C), 136.2, 138.4, 143.4, 145.2, 148.3, 153.3, 155.2, 159.3, 162.4, 166.4, 169.0 (coumarin C⚌O), 176.3 (indenone C⚌O). MS: m/z 508 [M⁺].

2.1.24

2.1.24 6-Bromo-3-(1-(4-fluorophenyl)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5l)

Yield 70%; m.p. 293–296 °C; Mol. Formula: C₃₀H₂₀BrFN₂O₅; IR (KBr, cm⁻¹): 3012 (Ar—H), 1712 (coumarin C⚌O), 1682 (indenone C⚌O), 1621 (C⚌N), 1593 (C⚌C); ¹H NMR (DMSO d₆, 400 MHz) δ: 3.23 (3H, s, OCH₃), 3.40 (3H, s,OCH₃), 4.72 (2H, s, CH₂), 7.38–8.87 (10H, m, Ar—H), 8.89 (1H, s, CH methine), 9.79 (1H, s, C⚌C—H), ¹³CNMR (CDCl₃) δ: 44.0 (CH₂), 57.4 (2-OCH₃), 104.4, 107.2, 109.1, 112.0 (2C), 114.2, 116.4, 119.0, 121.7, 122.5, 125.3, 127.0, 131.5, 133.4 (2C), 136.2, 138.4, 142.4, 144.2, 148.3, 152.5, 155.4, 152.3,163.4, 166.4, 170.0 (coumarin C⚌O), 175.0 (indenone C⚌O). MS: m/z 586 [M⁺], 588 [M+2].

2.2

2.2 Pharmacology

2.2.1

2.2.1 Antioxidant activity

2.2.1.1

2.2.1.1 Free radical scavenging activity by DPPH method

Free radical scavenging capacities of synthesized compounds were determined according to the reported procedure (Braca et al., 2001). The newly synthesized compounds at different concentrations (25–100 μg/mL) were added to each test tube and volume was made up to 4 mL using methanol. To this, 3 mL of 0.004% DPPH in methanol was added and the mixtures were incubated at room temperature under dark condition for 30 min. The absorbance was recorded at 517 nm using a UV–Visible spectrophotometer (Shimadzu UV-1800, Japan). Butylated hydroxyl toluene (BHT), dissolved in distilled water was used as a reference. Control sample was prepared using the same volume without any compound and BHT, 95% methanol served as blank. Test was performed in triplicate and the results were averaged. Radical scavenging activity was calculated using the formula: $% of radical scavenging activity = [(A_{control} - A_{test}) / A_{control}] \times 100$ where A_control is the absorbance of the control sample (DPPH solution without test sample) and A_test is the absorbance of the test sample (DPPH solution + test compound).

2.2.2

2.2.2 Chelating effect on ferrous ions

The chelating effect was determined according to the literature method (Nevcihan et al., 2010). The test solution (2 mL) of different concentrations (25–100 μg/mL) in methanol was added to a solution of 2 mM FeCl₂ (0.05 mL) and the reaction was initiated by adding 5 mM ferrozine (0.2 mL) and the total volume was adjusted to 5 mL with methanol. Then, the mixture was shaken vigorously and left at room temperature for 10 min. Absorbance of the solution was measured spectrophotometrically at 562 nm. EDTA was used as a standard. The inhibition percentage of ferrozine-Fe⁺² complex formations was calculated using the formula: $Metal chelating effect (%) = [(A_{control} - A_{sample}) / A_{control}] \times 100$ where A_control is the absorbance of control (control contains FeCl₂ ferrozine complex) and A_sample is the absorbance of test compounds. Test was performed in triplicate, and the results were averaged.

2.2.3

2.2.3 Total reductive capability

The reductive capability of the compounds was determined according to the literature method (Oyaizu, 1986). The solution of test compound (1 mL) at different concentrations (25–100 μg/mL) in methanol was mixed with phosphate buffer (2.5 mL, 0.2 mol/L pH 6.5) and potassium ferricyanide (2.5 mL, 1%) and the mixture was incubated at 50 °C for 20 min. At the end of the incubation, trichloroacetic acid (2.5 mL, 10%) was added to the mixture, and centrifuged at 3000 rpm for 10 min. The upper layer solution was collected and mixed with distilled water (2.5 mL) and 0.1% ferric chloride (0.5 mL, 0.1%) solution. The absorbance was measured at 700 nm against a blank. Test was performed in triplicate, and the results were averaged.

2.2.4

2.2.4 In-vivo anti-hyperglycemic activity

2.2.4.1

2.2.4.1 Animal housing and maintenance

Adult Wistar rats weighing between 200 and 300 g were used in this study. Animals were taken care as per the Regulations of Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA), Government of India. The ‘Form B’ for carrying out animal experimentation was reviewed and approved by the Institutional Animal Ethics Committee (IAEC). Animals were maintained in a controlled environment with 22–25 °C temperature, 50–60% humidity, a light/dark cycle of 12 h each and 15–20 air changes per hour. Animals were fed, ad libitum, with certified Irradiated Laboratory Rodent Diet (Nutri lab brand, Tetragon Chemie Pvt. Ltd, Bangalore) except during the fasting & study period.

2.2.4.2

2.2.4.2 Oral glucose tolerance test in normal rats

Overnight fasted normal rats were divided into twenty groups of six rats each. They were orally administered with vehicle, six synthesized compounds 5a, 5c and 5e–5h (10, 50 and 100 mg/kg) and metformin (60 mg/kg), respectively. Glucose (2 g/kg) was fed 30 min after the administration of compounds (Shirwaikar and Rajendran, 2006). Blood was withdrawn through the tail vein at 0, 30, 60 and 120 min of glucose administration to observe changes in the blood glucose level.

2.2.4.3

2.2.4.3 Hypoglycemic activity assay

Test substance and reference standard (metformin) suspension were prepared with vehicle containing 0.5% (w/v) of carboxymethyl cellulose and 0.1% Tween 80. Streptozotocin (STZ) was dissolved in citrate buffer (pH 4.5) and nicotinamide in normal saline solution (0.9% NaCl solution). Diabetes was induced in overnight fasted rats by a single intraperitoneal (i.p.) injection of 60 mg/kg of streptozotocin and 120 mg/kg of nicotinamide in Wistar rats (of both sexes) (Shirwaikar and Rajendran2006). Hyperglycemia was confirmed by the elevated non fasting glucose levels in blood, determined 48 h after diabetes induction using a Glucometer (Accu-check Advantage, Roche diagnostics Mannheim, Germany) and strips. Animals with blood glucose concentrations more than 250 mg/dL were used for the study. Each study group involved 6 diabetic rats. Cage cards indicating the study number, animal number & treatment group details were affixed to the corresponding cages. Standard drug and test compounds were given orally at 10, 50 and 100 mg/kg. The change in the body weight of rats after induction diabetes was noted. The blood sample was collected from caudal vein of the tested rats. Reduction in blood glucose produced by the compound was recorded on 1st, 2nd, 3rd, 4th, 7th and 14th day. At the end of experiment, animals were euthanized by CO₂ asphyxiation and organ weights were recorded.

Statistical analysis was performed by One-way ANOVA followed by Dunnett’s multiple comparison tests. P < 0.05 was considered as a significant change.

3 Result and discussion

3.1

3.1 Chemistry

Reaction sequences employed for the synthesis of title compounds is shown in Scheme 1. The key intermediates, 6-substituted-3-{(1E)-1-[2-(4-substituted) hydrazinylidene] ethyl}-2H-chromen-2-one (3a–l) were synthesized by stirring the reaction mixture of 6-substituted 3-acetyl coumarin (1a–b) with p-substituted phenyl hydrazine hydrochloride (2a–f) in dry ethanol using sodium acetate at room temperature (Goudarshivannanavar et al., 2009). Vilsmeier formylation of these compounds (3a–l) at reflux temperature for 6 h gave the expected formyl pyrazole derivatives (4a–l) in good yield. Further, Claisen–Schmidt condensation of 3-(6-substituted-2-oxo-2H-chromen-3-yl)-1-(4-substituted)-1H-pyrazole-4-carbaldehyde derivatives (4a–l) with 5,6-dimethoxy-2,3-dihydro-1H-inden-1-one at room temperature for 8 h led to the formation of corresponding 6-substituted-3-(1-(4-substituted)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene)methyl)-1H-pyrazol-3-yl)-2H-chromen-2-onederivatives (5a–l).

Synthesis of 6-substituted-3-(1-(4-substituted)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5a–l) derivatives.

All pyrazole coumarin indenone analogs provided satisfactory spectral data. In IR spectra, bands in the region 1671–1691 cm⁻¹ were attributed to the –CHO group of (4a–l) series and bands in the region 1625, 1520 cm⁻¹ were attributed to symmetric and asymmetric frequency of the NO₂ group of the compound 4j. Bands at 1703–1713 cm⁻¹ obtained from the lactone ring of coumarin C⚌O, 1667–1687 cm⁻¹ stretching frequencies correspond to the C⚌O groups of indenone. In ¹HNMR spectra, the singlet between δ 9.94–10.23ppm corresponds to the presence of the –CHO group of the compounds 4(a–l). The absence of aldehyde proton signal at δ 9.94–10.21 ppm and the presence of a signal in the range δ 9.68–9.91 ppm (C⚌C—H) support the formation of compounds 5(a–l) and the singlet in the range of 8.50–9.2 ppm due to the methine proton. In ¹³C-NMR, signal at δ 171.5–176.3 assigned to carbonyl carbon in indenone, signal at δ 168.2–169.2 confirmed lactone carbonyl of 5(a–l) and the signal between 43-49 ppm due to CH₂ carbon in the indenone ring. Signal that appeared between 56.2-58.6 corresponds to the OCH₃ carbon in indenone ring. Other signals are in good agreement with the target compounds.

3.2

3.2 Pharmacology

3.2.1

3.2.1 Free radical scavenging activity by DPPH method

The newly synthesized compounds (5a–l) were screened for their radical scavenging activity by the DPPH method. The activity results of the newly synthesized compounds are represented in Fig. 1.

DPPH free radical scavenging assay (%) of test compounds (5a–l).

We found that, most of the compounds showed considerable free radical scavenging activity. Compounds 5e–h have shown the strongest free radical scavenging activity among the tested compounds with IC₅₀ ranging between 54.14 and 59.76 μg/mL. The compound with no substitution on phenyl ring 5a with IC₅₀ 64.75 μg/mL and the compounds 5c and 5k substituted with halogens showed good scavenging effect with IC₅₀ 70.14 and 88.29 μg/mL. The other tested compounds displayed moderate to good scavenging activity. The IC₅₀ value of the reference standard BHT was recorded as 46.95 μg/mL.

3.2.2

3.2.2 Chelating effects on ferrous ions

The iron chelating studies measure the ability of antioxidants to compete with ferrozine in chelating ferrous ion (Elmastaş et al., 2006). It was reported that the compounds containing two or more functional groups like OH, SH, COOH, C⚌O, NR₂ and oxygen in a favorable structure–function configuration can show the activity of metal chelation (Yuan et al., 2005). The ferrous ion-chelating activity of the newly synthesized compounds is represented in Fig. 2. From the activity data it was observed that, the compounds substituted with electron donating groups on the indenone ring and phenyl ring 5e–5h exhibited excellent activity with IC₅₀ value 53.75–62.30 μg/mL. Compounds 5a and 5k displayed satisfactory activity with IC₅₀ 65.95 and 7.76 μg/mL. Compound 5c showed moderate activity with IC₅₀ 93.94 μg/mL. The IC₅₀ value of the reference standard EDTA was recorded as 44.11 μg/mL.

Ferrous ion chelating assay (%) of test compounds (5a–l).

3.2.3

3.2.3 Total reductive capability

Fe(III) reduction is often used as an indicator of electron donating activity, which is an important mechanism of phenolic antioxidant action. In the reducing power assay, the presence of antioxidant in the synthesized compounds would result in the reduction of Fe⁺³ to Fe⁺² complex by donating an electron. The amount of Fe⁺² complexes was then monitored by measuring the formation of Perl’s Prussian blue at 700 nm. Increasing absorbance at 700 nm indicates an increase in reducing ability (Nabavi et al., 2008). The results are represented in Fig. 3. It was found that the reducing power of all the synthesized compounds increased with the increase in their concentrations. The best reducing power was presented by the compound 5g containing two methoxy groups on indenone and a methoxy group on phenyl ring with IC₅₀ 58.01 μg/mL. Compounds substituted with electron donating groups 5e, 5f and 5h exhibited good activity with IC₅₀ ranging between 50 62.57 and 69.17 μg/mL. The other tested compounds showed less activity compared to standard. The IC₅₀ value of the reference standard Butylated hydroxyl anisole BHA was recorded as 50.00 μg/mL.

Total reductive capability assay (%) of test compounds (5a–l).

IC₅₀ values of DPPH radical scavenging ferrous ion chelating activity and total reductive capability of test compounds are given in Table 1.

Table 1 Half maximum inhibition concentration (IC₅₀) values for DPPH free radical scavenging, ferrous ion chelating and total reductive capability activity of test compounds.

Compounds	DPPH (IC₅₀ μg/mL)	Fe⁺² ion Chelating activity (IC₅₀ μg/mL)	Total reductive capability (IC₅₀ μg/mL)
5a	64.75 ± 0.11	65.95 ± 0.11	73.09 ± 0.11
5b	107.7 ± 0.32	105.01 ± 0.31	108.17 ± 0.31
5c	70.14 ± 0.10	93.94 ± 0.29	84.21 ± 0.29
5d	185.24 ± 0.15	189.29 ± 0.21	217.20 ± 0.21
5e	56.19 ± 0.18	53.75 ± 0.15	62.57 ± 0.15
5f	59.76 ± 0.21	59.45 ± 0.20	69.17 ± 0.20
5g	54.14 ± 0.13	56.89 ± 0.20	58.01 ± 0.20
5h	57.73 ± 0.13	62.30 ± 0.17	65.52 ± 0.17
5i	92.15 ± 0.17	117.36 ± 0.13	157.96 ± 0.13
5j	101.41 ± 0.13	153.87 ± 0.13	289.02 ± 0.13
5k	88.29 ± 0.21	70.76 ± 0.13	92.59 ± 0.13
5l	141.56 ± 0.13	131.02 ± 0.13	133.90 ± 0.13
Std^a,b,c	46.95 ± 0.17	44.11 ± 0.15	50.00 ± 0.15

Each value is expressed as mean ± SD of three replicates.

a Std-BHT is used as standard for DPPH radical scavenging activity.

b Std-EDTA is used as a standard for Fe⁺² ion chelating activity.

c Std-BHA is used as a standard for reductive capability.

3.2.4

3.2.4 Oral glucose tolerance test in normal rats

Oral glucose tolerance tests of synthesized compounds demonstrated the highest blood glucose level at 30 min following glucose administration, with decreasing blood glucose levels thereafter (Figs. 4–6 ). Compared with the groups, i.e. normal control, standard and with the tested compounds, compounds 5g and 5e showed a significant reduction in the blood glucose level. Additionally at 120 min following glucose administration, the 100 mg/kg 5g and 5e groups had significantly (P ⩽ 0.005) a greater drop in blood glucose than the 10 and 50 mg/kg groups. The other tested groups exhibited moderate to good decrease in the blood glucose level.

Effect of synthesized compounds on oral glucose tolerance at the concentration of 10 mg/kg of body weight.

Effect of synthesized compounds on oral glucose tolerance at the concentration of 50 mg/kg of body weight.

Effect of synthesized compounds on oral glucose tolerance at the concentration of 100 mg/kg of body weight.

3.2.5

3.2.5 In-vivo anti-hyperglycemic activity

The antidiabetic potential of the newly synthesized compounds was evaluated in STZ and nicotinamide induced T2DM in Wistar rats (Watcho et al. 2012). Blood glucose levels of compounds 5a, 5c and 5e–5h in diabetes in STZ/nicotinamide model were checked for 14 days.

Compounds 5g and 5e having an electron donating group on the phenyl ring showed prominent decrease in glucose concentration (115 and 138 mg/dL) with the dose of 100 mg/kg. It was interesting to note that the compound 5h substituted with electron withdrawing (Br) as well as electron donating (OCH₃) groups showed moderate to good activity. No significant decrease in the plasma glucose level was observed in compound 5f. Compounds 5a and 5c showed less activity by increase in the plasma glucose level (278 mg/dL and 387 mg/dL) with the dose of 100 mg/kg. The lowering of plasma glucose in diabetic rats after treatment with compounds 5a, 5c and 5e–h at concentrations 10, 50 and 100 mg/kg is represented in Figs 7–9 , respectively.

Significant lowering of plasma glucose in diabetic rats treated with compounds at the concentration of 10 mg/kg of body weight.

Significant lowering of plasma glucose in diabetic rats treated with compounds at the concentration of 50 mg/kg of body weight.

Significant lowering of plasma glucose in diabetic rats treated with compounds at the concentration of 100 mg/kg of body weight.

Treatment with the test substance for 14 days showed an increase in body weight (Table 2) compared to diabetic control; no significant increase in liver weight at 100 mg/kg; and no change in kidney weight were observed with treatment.

Table 2 Effect of synthesized compounds on body weight in STZ–nicotinamide induced diabetic rats.

Sl no.	Loss or gain of body weight of experimental rats
	10 mg/kg bw		50 mg/kg bw		100 mg/kg bw
	0th day	14th day	0th day	14th day	0th day	14th day
Normal control	195.01 ± 4.08	224.09 ± 9.94	ND	ND	ND	ND
Diabetic control	190.56 ± 9.45	168.13 ± 15.83	ND	ND	ND	ND
Standard	193.45 ± 5.03^a	197.07 ± 3.57 ^a	198.37 ± 4.03^a	203.67 ± 5.57^a	199.00 6.03^a,b	218.07 ± 3.57 ^a
5a	191.51 ± 4.12	192.08 ± 2.34	190.33 ± 2.14	193.25 ± 3.47	195.00 ± 3.13^a	193.05 ± 2.43^a
5c	188.51 ± 2.18	190.08 ± 2.04^a	190.33 ± 2.11^a,b	189.25 ± 4.17	194.00 ± 3.21	192.12 ± 2.43
5e	189.23 ± 4.23^a,b,c	192.01 ± 3.57	194.31 ± 2.03^a	200.25 ± 4.27	196.00 ± 2.03	210.17 ± 5.18
5f	188.41 ± 3.13	191.05 ± 2.34	192.23 ± 3.12^a,b,c	196.25 ± 2.17	195.00 ± 3.13^a	197.04 ± 2.43
5g	190.35 ± 3.03^a	194.04 ± 2.67 ^a	196.37 ± 2.21	201.67 ± 3.24^a	198.00 ± 3.43	216.02 ± 2.34^a,b
5h	192.27 ± 3.13	193.08 ± 4.54	195.33 ± 4.02	199.25 ± 5.57	200.00 ± 2.03^a	206.07 ± 4.57

Values are mean ± SEM of 6 animals in each group.

a P < 0.05 compared with the normal.

b P < 0.05 compared with diabetic control.

c P < 0.05 compared with the glibenclamide treated group. ND-Not determined.

4 Conclusion

A new series of coumarin derivatives containing pyrazole and indenone moiety have been synthesized and the synthesized compounds were well characterized by IR, ¹HNMR, ¹³CNMR and LC–MS spectroscopic methods. Our adopted method for the synthesis of target compounds was effective in terms of time and yield. The reaction conditions are simple, and they are not sensitive to oxygen and water, which make it easy to operate at room temperature. The presence of pyrazole ring endows these species with important pharmacological and therapeutic interest. From the structure–activity relationship studies it revealed that, the presence of electron withdrawing group/halogen substituted compounds showed good antioxidant property. In case of antihyperglycemic activity, compounds 5g and 5e showed prominent antidiabetic activity. Further research needs to be done to fully understand how the synthesized compounds are working in the pancreas and by what mechanism it is able to lower glucose levels in Streptozotocin–nicotinamide induced diabetic mice.

Acknowledgements

The authors are thankful to the Chairman, Department of Industrial Chemistry, Kuvempu University, Shankaraghatta for providing the laboratory facilities. One of the authors (Kenchappa R.) is thankful to the UGC for the award of Rajiv Gandhi National Fellowship (RGNF).

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Goudarshivannanavar B.C., Jayadevappa H., Mahadevan K.M., . Indian J. Chem.. 2009;48B:1419-1423.
Guo J.N., Weng X.C., Wu H., Li Q.H., Bi K.S., . Food Chem.. 2005;91:287-292.
Hyun D.H., Hernandez J.O., Mattson M.P., de Cabo R., . Aging Res. Rev.. 2006;5:209-220.
Isloor A.M., Kalluraya B., Rao M., . J. Saudi Chem. Soc.. 2000;4(3):265-270.
Jovanovic S.V., Steenken S., Tosic M., Marjanovic B., Simic M.G., . J. Am. Chem. Soc.. 1994;116:4846-4851.
Karali N., Kocabalkanli A., Gursoy A., Ates O., . Farmaco. 2002;57:589-593.
Kenchappa R., Bodke Y.D., Peethambar S.K., SandeepTelkar, Venkatesh K.B., . Med. Chem. Res.. 2013;22:4787-4797.
Kenchappa R., Bodke Y.D., Chandrashekar A., Telkar Sandeep, Manjunatha K.S., Aruna Sindhe M., . Arabian J. Chem.. 2017;10:S1336-S1344.
Khan A.I., Kulkarni V.M., Gopal M., Shahabuddin S.M., Chung-Ming S., . Bioorg. Med. Chem. Lett.. 2005;15:3587.
King H., Rewers M., . Diabetes Care. 1993;16:157-177.
Kus V., Flachs P., Kuda O., Bardova K., . PLoS One. 2011;6(11):e27126.
[CrossRef]
Lim Y.Y., Murtijaya J., . Food Sci. Technol.. 2007;40:1664-1669.
Mamta R., Mohamad Y., Salman A.K., Sahota P.P., Pandove G., . Arabian J. Chem.. 2015;8:S174-S180.
Nabavi S.M., Ebrahimzadeh M.A., Nabavi S.F., Hamidinia A., Bekhradnia A.R., . Pharmacol. Online. 2008;2:560-567.
Neogi P., Lakner F.J., Medicherla S., Cheng J., Dey D., Gowri M., Nag B., Sharma S.D., Pickford L.B., Gross C., . Bioorg. Med. Chem.. 2003;11:4059-4067.
Nevcihan G., Cengiz S., Bektas T., Halil S.M., . Food Sci. Biotech.. 2010;19(3):691-696.
Oyaizu M., . Jpn. J. Nutr.. 1986;44:307-315.
Padmaja A., Rajasekhar C., Muralikrishna A., Padmavathi V., . Eur. J. Med. Chem.. 2011;46:5034-5038.
Rizk H.F., El-Badawi M.A., Ibrahim S.A., El-Borai M.A., . Arabian J. Chem.. 2011;4:37-44.
Saija A., Scalese M., Lanza M., Marzullo D., Bonina F., Castelli F., . Free Radical Biol. Med.. 1995;19:481-486.
Saltiel A.R., Kahn C.R., . Nature. 2001;414:799-806.
Sangle G.V., Lauffer L.M., Grieco A., . Endocrinology. 2012;153:564.
Sas K., Robotka H., Toldi J., Vecsei L., . J. Neurol. Sci.. 2007;257:221-239.
Shirwaikar A., Rajendran K., . J. Ethnopharmacol.. 2006;107(2):285-290.
Singh U., Jialal I., . Pathophysiology. 2006;13:129-142.
Smith M.A., Rottkamp C.A., Nunomura A., Raina A.K., Perry G., . Biochim. Biophys. Acta. 2000;1502:139-144.
Stancho S., Georgi M., Frank J., Ilia M., . Eur. J. Med. Chem.. 2008;43:694-706.
Vijaya Laxmi S., Suresh K.B., Rajitha B., . Med. Chem. Res.. 2013;22:768-774.
Watcho P., Achountsa J.H.G., Mbiakop C.U., Wankeu N.M., Nguelefack T.B., Kamanyi A., . BMC Complem. Altern. Med.. 2012;12:264.
Xin-Hua L., Jun L., Jing B.S., Bao-An S., Xing-Bao Qi., . Eur. J. Med. Chem.. 2012;51:294-299.
Yuan Y.V., Bone D.E., Carrington M.F., . Food Chem.. 2005;91:485-494.

Introduction

Materials and methods

Chemistry
Pharmacology

Result and discussion

Chemistry
Pharmacology

Conclusion

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[13] Ghate M., Kusanur A.R., Kulkarni V.M., . Eur. J. Med. Chem.. 2005;40:882.

[14] Gireesh T., Ravindra R.K., Pramod P.K., Sheetal B.M., . Arabian J. Chem.. 2016;9:S306-S312.

[15] Goudarshivannanavar B.C., Jayadevappa H., Mahadevan K.M., . Indian J. Chem.. 2009;48B:1419-1423.

[16] Guo J.N., Weng X.C., Wu H., Li Q.H., Bi K.S., . Food Chem.. 2005;91:287-292.

[17] Hyun D.H., Hernandez J.O., Mattson M.P., de Cabo R., . Aging Res. Rev.. 2006;5:209-220.

[18] Isloor A.M., Kalluraya B., Rao M., . J. Saudi Chem. Soc.. 2000;4(3):265-270.

[19] Jovanovic S.V., Steenken S., Tosic M., Marjanovic B., Simic M.G., . J. Am. Chem. Soc.. 1994;116:4846-4851.

[20] Karali N., Kocabalkanli A., Gursoy A., Ates O., . Farmaco. 2002;57:589-593.

[21] Kenchappa R., Bodke Y.D., Peethambar S.K., SandeepTelkar, Venkatesh K.B., . Med. Chem. Res.. 2013;22:4787-4797.

[22] Kenchappa R., Bodke Y.D., Chandrashekar A., Telkar Sandeep, Manjunatha K.S., Aruna Sindhe M., . Arabian J. Chem.. 2017;10:S1336-S1344.

[23] Khan A.I., Kulkarni V.M., Gopal M., Shahabuddin S.M., Chung-Ming S., . Bioorg. Med. Chem. Lett.. 2005;15:3587.

[24] King H., Rewers M., . Diabetes Care. 1993;16:157-177.

[25] Kus V., Flachs P., Kuda O., Bardova K., . PLoS One. 2011;6(11):e27126.
[CrossRef]

[26] Lim Y.Y., Murtijaya J., . Food Sci. Technol.. 2007;40:1664-1669.

[27] Mamta R., Mohamad Y., Salman A.K., Sahota P.P., Pandove G., . Arabian J. Chem.. 2015;8:S174-S180.

[28] Nabavi S.M., Ebrahimzadeh M.A., Nabavi S.F., Hamidinia A., Bekhradnia A.R., . Pharmacol. Online. 2008;2:560-567.

[29] Neogi P., Lakner F.J., Medicherla S., Cheng J., Dey D., Gowri M., Nag B., Sharma S.D., Pickford L.B., Gross C., . Bioorg. Med. Chem.. 2003;11:4059-4067.

[30] Nevcihan G., Cengiz S., Bektas T., Halil S.M., . Food Sci. Biotech.. 2010;19(3):691-696.

[31] Oyaizu M., . Jpn. J. Nutr.. 1986;44:307-315.

[32] Padmaja A., Rajasekhar C., Muralikrishna A., Padmavathi V., . Eur. J. Med. Chem.. 2011;46:5034-5038.

[33] Rizk H.F., El-Badawi M.A., Ibrahim S.A., El-Borai M.A., . Arabian J. Chem.. 2011;4:37-44.

[34] Saija A., Scalese M., Lanza M., Marzullo D., Bonina F., Castelli F., . Free Radical Biol. Med.. 1995;19:481-486.

[35] Saltiel A.R., Kahn C.R., . Nature. 2001;414:799-806.

[36] Sangle G.V., Lauffer L.M., Grieco A., . Endocrinology. 2012;153:564.

[37] Sas K., Robotka H., Toldi J., Vecsei L., . J. Neurol. Sci.. 2007;257:221-239.

[38] Shirwaikar A., Rajendran K., . J. Ethnopharmacol.. 2006;107(2):285-290.

[39] Singh U., Jialal I., . Pathophysiology. 2006;13:129-142.

[40] Smith M.A., Rottkamp C.A., Nunomura A., Raina A.K., Perry G., . Biochim. Biophys. Acta. 2000;1502:139-144.

[41] Stancho S., Georgi M., Frank J., Ilia M., . Eur. J. Med. Chem.. 2008;43:694-706.

[42] Vijaya Laxmi S., Suresh K.B., Rajitha B., . Med. Chem. Res.. 2013;22:768-774.

[43] Watcho P., Achountsa J.H.G., Mbiakop C.U., Wankeu N.M., Nguelefack T.B., Kamanyi A., . BMC Complem. Altern. Med.. 2012;12:264.

[44] Xin-Hua L., Jun L., Jing B.S., Bao-An S., Xing-Bao Qi., . Eur. J. Med. Chem.. 2012;51:294-299.

[45] Yuan Y.V., Bone D.E., Carrington M.F., . Food Chem.. 2005;91:485-494.

Synthesis of coumarin derivatives containing pyrazole and indenone rings as potent antioxidant and antihyperglycemic agents

Abstract

Abstract

Keywords

Coumarin

Pyrazole

Indenone

Antioxidant

Antihyperglycemic

Streptozotocin–nicotinamide

1 Introduction

2 Materials and methods

2.1 Chemistry

2.1.1 General procedure for the synthesis of 3-(6-substituted-2-oxo-2H-chromen-3-yl)-1-(4-substituted)-1H-pyrazole-4-carbaldehyde derivatives (4a-l)

2.1.2 1-(4-Chlorophenyl)-3-(2-oxo-2H-chromen-3-yl)-1H-pyrazole-4-carbaldehyde (4c)

2.1.3 3-(6-Bromo-2-oxo-2H-chromen-3-yl)-1-(4-chlorophenyl)-1H-pyrazole-4-carbaldehyde (4d)

2.1.4 1-(4-Methylphenyl)-3-(2-oxo-2H-chromen-3-yl)-1H-pyrazole-4-carbaldehyde (4e)

2.1.5 3-(6-Bromo-2-oxo-2H-chromen-3-yl)-1-(4-methylphenyl)-1H-pyrazole-4-carbaldehyde (4f)

2.1.6 1-(4-Methoxyphenyl)-3-(2-oxo-2H-chromen-3-yl)-1H-pyrazole-4-carbaldehyde (4g)

2.1.7 3-(6-Bromo-2-oxo-2H-chromen-3-yl)-1-(4-methoxyphenyl)-1H-pyrazole-4-carbaldehyde (4h)

2.1.8 1-(4-Nitrophenyl)-3-(2-oxo-2H-chromen-3-yl)-1H-pyrazole-4-carbaldehyde (4i)

2.1.9 3-(6-Bromo-2-oxo-2H-chromen-3-yl)-1-(4-nitrophenyl)-1H-pyrazole-4-Carbaldehyde (4j)

2.1.10 1-(4-Fluorophenyl)-3-(2-oxo-2H-chromen-3-yl)-1H-pyrazole-4-carbaldehyde (4k)

2.1.11 3-(6-Bromo-2-oxo-2H-chromen-3-yl)-1-(4-fluorophenyl)-1H-pyrazole-4-carbaldehyde (4l)

2.1.12 Synthesis of 3-(4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-4-substituted-phenyl-1H-pyrazol-3-yl)-6-substituted-2H-chromen-2-onederivatives (5a–l)

2.1.13 3-(4-((Z)-(5,6-Dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-phenyl-1H-pyrazol-3-yl)-2H-chromen-2-one (5a)

2.1.14 6-Bromo-3-(4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-phenyl-1H-pyrazol-3-yl)-2H-chromen-2-one (5b)

2.1.15 3-(1-(4-Chlorophenyl)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene)methyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5c)

2.1.16 6-Bromo-3-(1-(4-chlorophenyl)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5d)

2.1.17 3-(4-((Z)-(5,6-Dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-p-tolyl-1H-pyrazol-3-yl)-2H-chromen-2-one (5e)

2.1.18 6-Bromo-3-(4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-p-tolyl-1H-pyrazol-3-yl)-2H-chromen-2-one (5f)

2.1.19 3-(4-((Z)-(5,6-Dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-(4-methoxy phenyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5g)

2.1.20 6-Bromo-3-(4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene)methyl)-1-(4-methoxy phenyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5h)

2.1.21 3-(4-((Z)-(5,6-Dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-(4-nitro phenyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5i)

2.1.22 6-Bromo-3-(4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1-(4-nitro phenyl)-1H-pyrazol-3-yl)-2H-chromen-2-one(5j)

2.1.23 3-(1-(4-Fluorophenyl)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene)methyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5k)

2.1.24 6-Bromo-3-(1-(4-fluorophenyl)-4-((Z)-(5,6-dimethoxy-1-oxo-1H-inden-2(3H)-ylidene) methyl)-1H-pyrazol-3-yl)-2H-chromen-2-one (5l)

2.2 Pharmacology

2.2.1 Antioxidant activity

2.2.1.1 Free radical scavenging activity by DPPH method

2.2.2 Chelating effect on ferrous ions

2.2.3 Total reductive capability

2.2.4 In-vivo anti-hyperglycemic activity

2.2.4.1 Animal housing and maintenance

2.2.4.2 Oral glucose tolerance test in normal rats

2.2.4.3 Hypoglycemic activity assay

3 Result and discussion

3.1 Chemistry

3.2 Pharmacology

3.2.1 Free radical scavenging activity by DPPH method

3.2.2 Chelating effects on ferrous ions

3.2.3 Total reductive capability

3.2.4 Oral glucose tolerance test in normal rats

3.2.5 In-vivo anti-hyperglycemic activity

4 Conclusion

Acknowledgements

References

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