SYNTHESIS, CHARACTERIZATION AND PHARMACOLOGICAL EVALUATION OF SOME NOVEL 2-METHYL-4(3H)-QUINAZOLINONE DERIVATIVES BEARING PYRAZOLINE MOIETY

The heterocyclic molecules containing nitrogen atoms, particularly five and six-membered such as pyrazoles and quinazolines have been recognized as potential scaffolds for therapeutic investigations. Several synthetic compounds containing pyrazoles have shown different biological properties such as anti-inflammatory¹, anticonvulsant², antihypertensive³, antimicrobial⁴, antibacterial⁵, anticancer⁶, antidepressant⁷ and antitubercular⁸ activities. Also, quinazoline nuclei have been extensively used in different compounds to get a broad spectrum of pharmacological activities, comprising analgesic⁹, anti-inflammatory¹⁰, anticonvulsant¹¹, antimicrobial¹², antibacterial¹³, antifungal¹⁴, anticancer¹⁵, antimalarial¹⁶, antioxidant¹⁷, anti-HIV¹⁸, antiviral¹⁹, anti-influenza²⁰, antitubercular²¹, anti-depressant²², antihypertensive²³ as well as anti-histamine²⁴ activities.

On the other hand, various therapeutic activities have been reported for both pyrazole as well as quinazolines moieties. As a part of our continued program on the chemistry of 3H-quinazolin-4-one ring systems, we recently developed a simple and efficient approach to a wide range of such derivatives. These results prompted us to synthesize a series of novel 2-methyl-3H-quinazolin-4-one derivatives containing a pyrazole, pyrazoline or pyrimidinone ring, intending to obtain some novel heterocyclic systems with potentially enhanced biological properties.

2.1. Instrumentation and Synthesis

All starting materials, reagents, and solvents were purchased from commercial suppliers like Merck (Germany) and Sigma Aldrich Chemical Co. Analytical thin-layer chromatography (TLC) was conducted using Merck silica gel 60F254 plates. Infrared (IR) was recorded using a Fourier-transform IR (FT-IR) spectrophotometer. Proton nuclear magnetic resonance (¹H NMR) spectra were recorded by a Bruker 500 MHz spectrophotometer and chemical shifts are expressed as ppm with tetramethylsilane (TMS) as the internal standard. Compounds melting points were determined by the melting point apparatus and are uncorrected.

2.1.1. Synthesis of 3-(4-acetylphenyl)-2-methylquinazolin-4-one (1): Anthranilic acid (1.37g, 0.01M) was dissolved in 30 ml of dry pyridine (0.79 g, 0.01 mol) by stirring slowly at room temperature. The solution was cooled to 0-5°C and a solution of acetyl chloride (1.56 gm, 0.02 M) in dry pyridine (30 ml) was added slowly with constant stirring. After this addition, the reaction mixture was further stirred for half an hour at room temperature and set aside for 1hr. The pasty mass obtained was added with p-aminoacetophenone (1.35g, 0.01 M) in pyridine and refluxed for 6 hr. Excess of solvent was distilled off. The resulting mixture was cooled and poured over crushed ice. The solid was filtered, washed with cold water, and recrystallized from a suitable solvent.

2.1.2. Synthesis of 2-methyl-3-[4-(-3-aryl substituted prop-2-enoyl) phenyl] quinazolin-4-ones (2a-2j): Synthesized 3-(4-acetylphenyl)-2-methylquinazolin-4-one (0.01M) was added to different aromatic aldehydes (0.01M) in 30ml of ethanol and 40% sodium hydroxide solution. The mixture was stirred for 2hr at room temperature and kept in the refrigerator for 24hr. The content was poured on crushed ice and neutralized with 10% HCl, the product was filtered, dried and recrystallized from a suitable solvent.

2.1.3. Synthesis of 2-methyl-3-[4-(4-aryl substituted-4,5-dihydro-1H-pyrazol-3-yl) phenyl] quinazolin-4(3H)-ones (3a-3j): A mixture of compound 2 (0.01 M) and hydrazine hydrate (0.01 M, dissolved in 2 ml glacial acetic acid) was heated at reflux temperature for 6 hr. After completion of the reaction, the mixture was cooled to room temperature and poured into ice-cold water, filtered and recrystallized.

2.1.4. Synthesis of 2-methyl-3-[4-(4-aryl substituted-1-carbaldehyde-2-pyrazolin-3-yl) phenyl] quinazolin-4-ones (4a-4j): To a mixture of compound 2 (0.01 mol) and hydrazine hydrate (0.05M, 1.6ml) add formic acid (40ml) and reflux for 26 hr. On completion of the reaction (TLC monitoring) the resulting solution was poured into ice-cold water and allowed to stand overnight. Precipitate formed was filtered and recrystallized.

Scheme 1: Synthetic pathway for the preparation of title compounds (1, 2a–2j, 3a–3j, 4a–4j)

2.1.5. Physical Characterization

The physical characterization data including yield, melting point, molecular weight, elemental formula and thin layer chromatography (TLC) Rf values for all synthesized pyrazoline derivatives (2a–2j, 3a–3j and 4a–4j) are summarized below.

Table 1: Physical Characterization of the intermediate (2a – 2j)

Table 2: Physical Characterization of the title compounds (3a – 3j and 4a – 4j)

2.2. Spectral Data and Elemental Analysis

The structures of newly synthesized compounds were characterized by FTIR, ¹H NMR, and mass spectrometry.

Table 4: Interpretation of ¹H NMR and MS spectral data of title compounds

Table 5: Data of CHNS elemental analyses of title compounds

3.1. Acute toxicity study: Acute toxicity study was performed for the synthesized compounds to ascertain safe dose by acute oral toxic class method of Organization of Economic Co-operation and Development, as per 423 guidelines. Synthesized compound’s (3a-4j) dose was fixed as 50 mg/kg for pharmacological study. No mortality or behavioral abnormalities were observed at this dose level.

3.2. Anticonvulsant Evaluation

Table 6: Anticonvulsant activity of 2-methyl-4(3H)-quinazolinones (MES method)

FIG: 1 — Graphical representation of Anticonvulsant Activity (MES Method)

Table 8: Anticonvulsant activity of 2-methyl-4(3H)-quinazolinones (PTZ method)

FIG: 2 — Graphical representation of Anticonvulsant Activity (PTZ Method)

3.3. Analgesic Evaluation

Table 9: Writhing and analgesic activity (Aspirin comparison)

FIG: 4 — Graphical representation of Analgesic Activity (Writhing Test)

Table 10: Writhing and analgesic activity (Pentazocine comparison)

FIG: 5 & 6 — Graphical representation of Analgesic Activity (Hot Plate & Tail Flick)

3.4. Anti-inflammatory Evaluation

Table 11 & 12: Percentage Inhibition of Paw Edema (Carrageenan induced)

FIG: 7 & 8 — Graphical representation of Anti-inflammatory Activity Comparison

In conclusion, a series of novel 2-methyl-4(3H)-quinazolinone derivatives bearing pyrazoline moieties (3a–3j and 4a–4j) were successfully synthesized in good yields and structurally characterized by FTIR, ¹H NMR, mass spectrometry and elemental analysis. Pharmacological screening demonstrated that compounds containing electron-withdrawing and methoxy groups (such as 4c, 4h, and 4i) exhibited significant anti-inflammatory and analgesic properties comparable to standard reference drugs without any acute toxicity.

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