INTRODUCTION:

Synthesis of ethyl 2-(4-halobenzyl)-3-oxobutanoate was carried out (figure 1) by using addition of substituted benzyl bromide to a suspension of t-BuOK in THF¹. Two substituted benzyl bromide was used i.e bromo benzyl bromide and chloro benzyl bromide. This reaction is a type of nucleophilic substitution. The three different ethyl 2-(4-substituted benzyl)-3-oxobutanoate compound was synthesize. All the synthesized compounds are listed in Table 1 and their structures are determined by elemental analyses and spectral data (IR, 1 H-NMR, EI-MS) and its pharmacological activity was determine by using prediction of activity spectra for substance (PASS) shown in table 2 and 3^2,3.

PASS Server, this server predicted activity spectrum of a chemical compound as a Pa (probable activity) and Pi (probable inactivity)⁴. If Pa ˃ 0.7, the substance is very likely to exhibit the activity in experiment, but the chance of substance being the analogue of known pharmaceutical agent is high. If 0.5 < Pa <o.7 the substance is likely to exhibit activity in experiment, but the probability is less, and the substance is unlike known pharmaceutical agents. If Pa is < 0.5 the substance is unlikely to exhibit the activity in experiment. However if the presence of this activity is confirmed in the experiment the substance might to be new molecule entity^5,6. The PASS study of ethyl 2-(4-bromobenzyl)-3-oxobutanoate and ethyl 2-(4-chlorobenzyl)-3-oxobutanoate was carried out. It shows various pharmacological activities such as L-glucuronate reductase inhibitor, Gluconate 2-dehydrogenase (acceptor) inhibitor, Centromere associated protein inhibitor, Eye irritation, inactive, GST A substrate, Pectate lyase inhibitor, 5-O-(4-coumaroyl)-D-quinate 3'-monooxygenase inhibitor, Acetylcholine neuromuscular blocking agent, Antihypoxic, Aspulvinone dimethylallyl transferase inhibitor.

MATERIAL AND METHOD:

Material: All starting materials and reagents for the synthesis were purchased from commercial sources. Distilled solvents were used for carrying out reactions as well as column chromatography. All the reactions as well as chromatographic processes were monitored by TLC technique on precoated silica gel plates of size 3cm x 1cm procured from E-Merck., followed by exposure to iodine vapors and/or UV irradiation for detection and/or Anisaldehyde reagent and/or Ninhydrin reagent, then it was charred or heated on a hot plate.

Method: To a suspension of t-BuOK (4.74 g, 42.46 mmol) in THF (70mL) was added ethyl acetoacetate (4.9 mL, 38.42mmol) at 0⁰C and the resulting solution was stirred for 30 minutes followed by addition of substituted benzyl bromide (38.42mmol). The reaction mixture was then refluxed for 12h. The progress of reaction was monitored using TLC. After completion of reaction, the reaction mixture was concentrated on rotary evaporator. The residue was diluted with 30ml water and the product was then extracted with ethyl acetate (3 × 30mL). The combined organic extracts were washed with brine, dried over Na₂SO₄, filtered, and concentrated under vacuum to give oily liquid, which was used directly for next reaction or purified by silica gel column chromatography (compound 3a-c) for spectral analysis (Mobile phase: 1 % EtOAc: Petroleum ether)⁷. Melting points of the synthesized compounds were recorded on “ANALAB, model-ThermoCal” melting point apparatus shown in table 1. The IR spectra were recorded on “Shimadzu IR solution 1.60” Solids were recorded as KBr pallets and liquids as thin film or CHCl3 solution on NaCl discs.1H NMR spectra were recorded on Agilent 400MHz. Chemical shifts were in parts per million (δ ppm) downfield with respect to tetramethylsilane (TMS) as internal standard. Molecular mass was recorded by GC-MS and only mass spectra presented here with mass spectra on Agilent MDS.

Figure 1: Synthesis of ethyl 2-(4-halobenzyl)-3-oxobutanoate

Table 1: Characteristics of ethyl 2-(4-halobenzyl)-3-oxobutanoate

No	Compound	Yield	Boiling Point	Nature	*R_f
3a		40%	ND	Colourless oily liquid	0.51
3b		54%	ND	Golden yellow oil	0.46

3c		50%	ND	Golden yellow oil	0.48

RESULTS:

IR Spectroscopy:

The IR spectra of ethyl 2-(4-halobenzyl)-3-oxobutanoate (3a-c) was recorded and shown in figure 2,3 and 4.

Figure 2: IR spectrum of ethyl 2-benzyl-3-oxobutanoate (3a)

Figure 3: IR spectrum of ethyl 2-(4-bromobenzyl)-3-oxobutanoate (3b)

Figure 4: IR spectrum of ethyl 2-(4-chlorobenzyl)-3-oxobutanoate (3c)

NMR Spectroscopy: ¹H NMR spectrum of ethyl 2-benzyl-3-oxobutanoate (3a) was recorded and shown in figure 5.

Figure 5: ¹H NMR spectrum of ethyl 2-benzyl-3-oxobutanoate (3a)

Mass Spectroscopy:

Mass spectra of ethyl 2-benzyl-3-oxobutanoate (3a) was recorded and shown in figure 6.

Figure 6: Mass spectrum of ethyl 2-benzyl-3-oxobutanoate

Prediction of activity spectra for substance:

Predicted activity (Pa) and Predicted inactivity (Pi) of ethyl 2-(4-bromobenzyl)-3-oxobutanoate and ethyl 2-(4-chlorobenzyl)-3-oxobutanoate was shown in table 2 and 3.

Table 2: predicted activity (Pa) and Predicted inactivity (Pi) of ethyl 2-(4-bromobenzyl)-3-oxobutanoate

Pa	Pi	Biological Activity
0.912	0.002	L-glucuronate reductase inhibitor
0.880	0.005	Gluconate 2-dehydrogenase (acceptor) inhibitor
0.851	0.005	Prolyl aminopeptidase inhibitor
0.800	0.018	Mucomembranous protector
0.810	0.031	Aspulvinone dimethylallyltransferase inhibitor
0.777	0.001	3-Methyl-2-oxobutanoate hydroxymethyltransferase inhibitor
0.777	0.005	Centromere associated protein inhibitor
0.785	0.013	Feruloyl esterase inhibitor
0.771	0.004	Antihypoxic
0.730	0.017	5-O-(4-coumaroyl)-D-quinate 3'-monooxygenase inhibitor
0.706	0.012	2-Hydroxymuconate-semialdehyde hydrolase inhibitor
0.673	0.005	Pectate lyase inhibitor
0.690	0.024	GST A substrate
0.700	0.039	Polyporopepsin inhibitor
0.694	0.048	Antieczematic
0.653	0.009	Acetylcholine neuromuscular blocking agent
0.652	0.015	Linoleate diol synthase inhibitor
0.629	0.003	Rhodotorulapepsin inhibitor
0.649	0.025	Arginine 2-monooxygenase inhibitor
0.634	0.011	Eye irritation, inactive
0.632	0.013	Cutinase inhibitor
0.632	0.016	Platelet aggregation stimulant
0.654	0.o43	Fibrinolytic
0.614	0.008	Alkylglycerone-phosphate synthase inhibitor
0.614	0.013	N-formylmethionyl-peptidase inhibitor
0.617	0.019	1,4-Lactonase inhibitor
0.618	0.024	Cytoprotectant
0.658	0.069	Testosterone 17beta-dehydrogenase (NADP+) inhibitor
0.589	0.004	Antiuremic
0.605	0.023	Preneoplastic conditions treatment
0.599	0.018	All-trans-retinyl-palmitate hydrolase inhibitor
0.625	0.046	Glutamyl endopeptidase II inhibitor
0.654	0.047	Phobic disorders treatment

Table 3: predicted activity (Pa) and Predicted inactivity (Pi) of ethyl 2-(4-chlorobenzyl)-3-oxobutanoate

Pa	Pi	Biological Activity
0.910	0.004	Gluconate 2-dehydrogenase (acceptor) inhibitor
0.887	0.003	5-O-(4-coumaroyl)-D-quinate 3'-monooxygenase inhibitor
0.879	0.009	Phobic disorders treatment
0.855	0.004	2-Hydroxymuconate-semialdehyde hydrolase inhibitor
0.829	0.005	Linoleate diol synthase inhibitor
0.812	0.017	Antiseborrheic
0.791	0.005	Cutinase inhibitor
0.797	0.021	Chlordecone reductase inhibitor
0.777	0.001	3-Methyl-2-oxobutanoate hydroxymethyltransferase inhibitor
0.790	0.021	Mucomembranous protector
0.757	0.008	Fibrinolytic
0.740	0.002	Pediculicide
0.742	0.005	Preneoplastic conditions treatment
0.752	0.016	GST A substrate
0.730	0.012	Arylsulfate sulfotransferase inhibitor
0.735	0.032	Polyporopepsin inhibitor
0.717	0.017	Arginine 2-monooxygenase inhibitor
0.711	0.012	Lipoprotein lipase inhibitor
0.745	0.052	Ubiquinol-cytochrome-c reductase inhibitor
0.703	0.011	Lipid metabolism regulator
0.693	0.008	Antihypoxic
0.684	0.005	H+-exporting ATPase inhibitor
0.683	0.007	Eye irritation, inactive
0.718	0.044	CYP2J substrate
0.682	0.014	Creatininase inhibitor
0.696	0.034	CYP2J2 substrate
0.694	0.048	Antieczematic

DISCUSSION:

Synthesis of ethyl 2-(4-halobenzyl)-3-oxobutanoate (3a-c):

Synthesis of compounds 3a-c was done by nucleophilic substitution of substituted benzyl bromides on ethyl acetoacetate in presence of potassium tertiary butoxide in THF.

The IR spectra for compounds 3a-c showed:

· Weak aromatic C-H stretch peak appeared between 3020-3030 cm^-1

· Medium aliphatic C-H stretch peak appeared between 2930-2985 cm^-1

· A sharp peak for ester C=O stretch appeared between 1735-1740 cm^-1

· A sharp peak for ketone C=O stretch appeared between 1712-1716 cm^-1

· Strong peak for sp² C-O stretch appeared between 1213-1246 cm^-1

· Strong peak for sp³ C-O stretch appeared between 1020-1090 cm^-1

· Strong peak for mono-substituted benzene ring appeared at 700 cm^-1

· Strong peak for di-substituted benzene ring appeared between 811-814 cm^-1

¹H NMR and GC-MS data of ethyl 2-benzyl-3-oxobutanoate (3a)

· Multiplet of five aromatic protons appeared near δ 7.47-6.96 ppm.

· Quartet of two and triplet of three protons of COOCH₂CH₃ appeared near δ 4.1 and δ 1.2 ppm respectively.

· Triplet of one proton of O-CH₃ appeared near δ 3.8 ppm.

· Doublet of two protons of –CH₂next to benzene ring appeared at δ 3.18 ppm.

· Singlet of three protons next to C=O appeared at δ 2.24 ppm.\

· Molecular ion peak appeared at 220.

In PASS study, ethyl 2-(4-bromobenzyl)-3-oxobutanoate and ethyl 2-(4-chlorobenzyl)-3-oxobutanoate shows various pharmacological activities such as L-glucuronate reductase inhibitor, Gluconate 2-dehydrogenase (acceptor) inhibitor, Centromere associated protein inhibitor, Eye irritation, inactive, GST A substrate, Pectate lyase inhibitor, 5-O-(4-coumaroyl)-D-quinate 3'-monooxygenase inhibitor, Acetylcholine neuromuscular blocking agent, Antihypoxic, Aspulvinone dimethylallyl transferase inhibitor.

CONCLUSION:

In the present paper, the synthesis of ethyl 2-(4-halobenzyl)-3-oxobutanoate was carried out and its pharmacological activity determine by using PASS. This compound shows various pharmacological activities so its target point for further synthesize various substituted new compound and determine its Pharmacological sactivity.

CONFLICTS OF INTEREST:

Authors do not have any conflict of interest.

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Received on 13.07.2021 Modified on 04.11.2021

Asian J. Res. Pharm. Sci. 2022; 12(2):102-106.

DOI: 10.52711/2231-5659.2022.00017