Table of Contents
Journal of Amino Acids
Volume 2014, Article ID 721291, 14 pages
http://dx.doi.org/10.1155/2014/721291
Research Article

Design and Synthesis of Novel Isoxazole Tethered Quinone-Amino Acid Hybrids

1Department of Medicinal Chemistry, GVK Biosciences Pvt. Ltd., Plot No. 28, IDA, Nacharam, Hyderabad, Andhra Pradesh 500 076, India
2Centre for Chemical Sciences & Technology, Institute of Science and Technology, Jawaharlal Nehru Technological University, Kukatpally, Hyderabad, Andhra Pradesh 500 072, India

Received 26 June 2014; Accepted 14 October 2014; Published 19 November 2014

Academic Editor: Sambasivarao Kotha

Copyright © 2014 P. Ravi Kumar et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

A new series of isoxazole tethered quinone-amino acid hybrids has been designed and synthesized involving 1,3-dipolar cycloaddition reaction followed by an oxidation reaction using cerium ammonium nitrate (CAN). Using this method, for the first time various isoxazole tethered quinone-phenyl alanine and quinone-alanine hybrids were synthesized from simple commercially available 4-bromobenzyl bromide, propargyl bromide, and 2,5-dimethoxybenzaldehyde in good yield.

1. Introduction

Compounds containing the quinone group present an important class of biologically active molecules that are widespread in nature [13]. The discoveries of antibiotic [4, 5] and antitumor [6] properties assigned to several natural quinones have raised interest among scientists for use as pharmaceuticals. While antibiotics display an enormous diversity in chemical structures, quinone antibiotics such as Adriamycin, Mitomycin C, and Streptonigrin deserve special attention [710]. In this context, search of new molecules containing quinone moiety has always fascinated the organic as well as medicinal chemist.

Isoxazole derivatives are an important class of heterocyclic pharmaceuticals and bioactive natural products because of their significant and wide spectrum of biological activities, including potent and selective antagonism of the NMDA receptor and anti-HIV activity. [11, 12]. It shows antihyperglycemic [13], analgesic [14], anti-inflammatory [15], antifungal [16], and antibacterial activity [17]. 3,5-Disubstituted isoxazole derivatives which are biological active include muscimol, dihydromuscimol, micafungin, and cycloserine [18, 19]. Unnatural amino acids, the nonproteinogenic α-amino acids that occur either naturally or chemically synthesized, have been used widely as chiral building block. They have been also used as molecular scaffolds in constructing combinatorial libraries [20]. They represent a powerful tool in drug discovery when incorporated into therapeutic peptidomimetics and peptide analogs [21]. The seminal work on the synthesis of unnatural amino acids has been done by O’Donnell and Maruoka independently, which accelerated the application of this class of amino acid for practical applications [22, 23].

Synthesis of hybrid natural products has gained momentum in recent years [2426]. It is expected that combining features of more than one biologically active natural segment in a single molecule may result in pronounced pharmacological activity while retaining high diversity and biological relevance. There are a few reports describing the preparation of quinone-hybrid with other natural products. For example, quinone-amino acids [27], sugar-oxasteroid-quinone [28], quinone-annonaceous acetogenins [29], and conduritol-carba-sugar [30] hybrids have been described using different synthetic protocol.

In our continuation endeavour to prepare novel hybrid molecules containing variety of natural products [31], we developed interest in the synthesis of novel isoxazole tethered quinone-amino acid hybrid natural products, and herein we report our initial results. Depending on the hybrid pattern, hybrid molecules containing amino acids and either quinone or isoxazole were prepared by various groups using different methods (Figure 1). For example, AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) is a type of glutamatergic ion channels in the central nervous system which can be considered as isoxazole-amino acid hybrid. A series of novel AMPA analogues were prepared in order to evaluate it as drug candidates for neurological disorder [32]. Abenquine D is an amino acid quinone hybrid which is composed of an amino acid linked to an N-acetyl-amino benzoquinone. Abenquines A–D are new bioactive secondary metabolites found in the fermentation broth of Streptomyces sp. stain DB634 which was isolated from the soils of Chilean highland of Atacama desert. The abenquines show inhibitory activity against bacteria, dermatophytic fungi, and phosphodiesterase type 4b [33]. It is noteworthy to mention here that amino acid attached to the quinone is relevant to the enzyme inhibitory activity. Similarly, IRL 3461 is a potent and bifunctional endothelin antagonist. IRL 346a is an isoxazole-amino acid hybrid prepared from 4-methyl-acetophenone in nine steps synthetic protocol [34]. Katritzky et al. have prepared naphthoquinone-amino acid conjugates starting from naphthoquinone and L-amino acids by a Michael type mechanism in aqueous ethanol solution at RT in the presence of triethylamine [35]. Kotha group has used a “building block approach” to synthesize the quinone-amino acid hybrids through ethylene cross-enyne metathesis and Diels-Alder reaction as the key step [27]. But there are no reports of isoxazole tethered quinone amino acid hybrids as per the literature search. To the best of our knowledge, this is the first report on the synthesis of new series of isoxazole tethered quinone-amino acid hybrid natural products.

721291.fig.001
Figure 1: Selected examples of amino acid hybrids.

In view of the importance of these three classes of natural products, we have designed a new class of hybrid structures 1 or 2 (Figure 2) in an effort to combine the activity of amino acid moiety and the quinone unit using isoxazole ring as linker. These hybrids may have significant biological activity and so an efficient strategy to these hybrid molecules would allow us to construct diverse hybrid analogues.

721291.fig.002
Figure 2: Isoxazole tethered quinone amino acid hybrid.

2. Materials and Methods

All reactions were carried out in oven-dried glassware with magnetic stirrers under an argon atmosphere. THF was dried over Na/benzophenone and DCM was dried over CaH2. Commercially available chemicals were purchased from Sigma-Aldrich and Alfa Aesar. EtOAc and pet ether were distilled before use. All melting points were taken in open capillaries and are uncorrected. Analytical thin-layer chromatography (TLC) was performed on commercially available Merck TLC Silica gel 60 F254. Silica gel column chromatography was performed on silica gel 60 (spherical 100–200 μm). FTIR spectra were recorded on Perkin-Elmer FT/IR-4000 spectrophotometer and only the characteristic peaks are reported. Mass spectra were scanned on a Shimadzu LCMS 2010 spectrometer. 1H NMR spectra were recorded on Varian-400 (400 MHz) spectrometer. Chemical shifts of 1H NMR spectra were reported relative to tetramethylsilane. 13C NMR spectra were recorded on Varian-400 (100 MHz) spectrometer. Chemical shifts of 13C NMR spectra were reported to be relative to CDCl3  (77.0). Splitting patterns were reported as s, singlet; d, doublet; t, triplet; q, quartet; m, multiplet; dd, double doublet; and br, broad.

2.1. Experimental Procedure for the Preparation of Methyl 2-((tert-Butoxycarbonyl)amino)-3-(4-((trimethylsilyl)ethynyl)phenyl)propanoate (4a)

To a solution of compound 3a (1.0 g, 2.80 mmol) in triethylamine (10 mL), PdCl2(PPh3)2 (0.098 g, 0.14 mmol), CuI (0.013 g, 0.07 mmol), and trimethylsilylacetylene (0.411 g, 4.20 mmol) were added under argon atmosphere and heated at 80°C in a sealed tube for 12 h. The progress of the reaction was monitored by TLC analysis (20% ethyl acetate/pet ether). After completion of the reaction, the reaction mixture was filtered. The filtrate was evaporated to give the crude product which was charged on silica gel column. The column was eluted with 20% ethyl acetate/pet ether to give the compound 4a (0.800 g, 76% yield) as light yellow liquid.

IR (KBr, cm−1): 3375, 2961, 2158, 1716, 1505, 1250, 1168, 865, 843. 1H NMR (400 MHz, CDCl3): δ = 7.42–7.36 (m, 2H), 7.06 (d, J = 7.8 Hz, 2H), 4.94 (d, J = 8.1 Hz, 1H), 4.57 (d, J = 7.4 Hz, 1H), 3.69 (s, 3H), 3.09 (td, J = 14.2, 6.1 Hz, 2H), 1.42 (s, 9H), 0.2 (s, 9H). MS (EI): m/z 375 (M + 1, 100).

2.2. Experimental Procedure for the Preparation of Methyl 2-Pivalamido-3-(4-((trimethylsilyl)ethynyl)phenyl)propanoate (4b)

To a solution of compound 3b (3.5 g, 10.26 mmol) in triethylamine (20 mL), PdCl2(PPh3)2 (0.359 g, 0.51 mmol), CuI (0.048 g, 0.25 mmol), and trimethylsilylacetylene (1.20 g, 12.31 mmol) were added under argon atmosphere and heated at 90°C in a sealed tube for 12 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). After completion of the reaction, the reaction mixture was filtered. The filtrate was evaporated to give the crude reaction mixture which was charged on silica gel column. The column was eluted with 20% ethyl acetate/pet ether to give the compound 4b (1.8 g, 48% yield) as off-white solid.

m.p. 143–145°C. IR (KBr, cm−1): 3328, 2958, 2158, 1751, 1638, 1205, 841. 1H NMR (300 MHz, DMSO): δ = 7.40 (dd, J = 1.7, 7.8 Hz, 2H), 7.20 (dd, J = 10.3, 8.1 Hz, 2H), 4.53–4.38 (m, 1H), 3.61 (d, J = 1.7 Hz, 3H), 3.18–2.89 (m, 2H), 1.00 (d, J = 1.6 Hz, 9H), 0.2 (s, 9H). MS (EI): m/z 360 (M + 1, 100).

2.3. Experimental Procedure for the Preparation of Methyl 2-((tert-Butoxycarbonyl)amino)-3-(4-ethynylphenyl)propanoate (5a)

To a solution of compound 4a (0.800 g, 2.13 mmol) in THF (10 mL), 1 M TBAF in THF (4.26 mL, 4.26 mmol) was added at −70°C and stirred for 2 h. The progress of the reaction was monitored by TLC analysis (20% ethyl acetate/pet ether). After the reaction was complete, the reaction mixture was quenched with water (10 mL) and extracted with ethyl acetate thrice. The organic layers were combined and washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 5a (0.650 g, 95% yield) as light yellow solid.

m.p. 94–97°C. IR (KBr, cm−1): 3355, 2974, 2103, 1739, 1682, 1519, 1170, 826. 1H NMR (400 MHz, CDCl3): δ = 7.42 (d, J = 7.8 Hz, 2H), 7.09 (d, J = 7.8 Hz, 2H), 4.96 (s, 1H), 4.58 (d, J = 7.8 Hz, 1H), 3.71 (d, J = 1.0 Hz, 3H), 3.21–2.94 (m, 3H), 1.42 (s, 9H). MS (EI): m/z 303 (M + 1, 100).

2.4. Experimental Procedure for the Preparation of Methyl 3-(4-Ethynylphenyl)-2-pivalamidopropanoate (5b)

To a solution of compound 4b (1.0 g, 3.84 mmol) in THF (20 mL), 1 M TBAF in THF (3.8 mL, 7.66 mmol) was added at −70°C and stirred for 2 h. The progress of the reaction was monitored by TLC analysis (20% ethyl acetate/pet ether). After the reaction was complete, the reaction mixture was quenched with water (20 mL) and extracted with ethyl acetate thrice. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 5b (0.650 g, 82% yield) as off-white solid.

m.p. 65–68°C. IR (KBr, cm−1): 3326, 2957, 1750, 1737, 1637, 1522, 1202, 1116. 1H NMR (400 MHz, DMSO): δ = 7.39–7.33 (m, 2H), 7.27–7.20 (m, 2H), 4.46 (m, 1H), 4.12 (s, 1H), 3.61 (s, 3H), 3.18–2.90 (m, 2H), 1.00 (d, J = 2.2 Hz, 9H). MS (EI): m/z 288 (M + 1, 100).

2.5. Experimental Procedure for the Preparation of 2,5-Dimethoxybenzaldehyde Oxime (7a)

To a solution of compound 6a (1 g, 6.02 mmol) in MeOH (10 mL), NaOAc (0.98 g, 12.04 mmol) and NH2OH·HCl (0.62 g, 9.03 mmol) were added under nitrogen atmosphere. Then the reaction mixture was stirred at RT for 2 h. The progress of the reaction was monitored by TLC analysis (20% ethyl acetate/pet ether). After completion of the reaction, the solvent was evaporated, quenched with water (20 mL), and extracted with ethyl acetate thrice. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 7a (1.0 g, 91% yield) as off-white solid.

m.p. 105–107°C. IR (KBr, cm−1): 3245, 2838, 1577, 1503, 1277, 1232, 1038, 970. 1H NMR (400 MHz, CDCl3): δ = 8.5 (s, 1H), 7.6–8.0 (br s, 1H), 7.3 (m, 1H), 6.9 (m, 1H), 6.8 (m, 1H), 3.8 (s, 3H), 3.7 (s, 3H). MS (EI): m/z 181 (M+, 100).

2.6. Experimental Procedure for the Preparation of 2,5-Dimethoxy-4-methylbenzaldehyde Oxime (7b)

To a solution of compound 6b (3.0 g, 16.6 mmol) in MeOH (30 mL), NaOAc (2.73 g, 33.3 mmol) and NH2OH·HCl (1.73 g, 24.9 mmol) were added under nitrogen atmosphere. Then the reaction mixture was stirred at RT for 2 h. The progress of the reaction was monitored by TLC analysis (20% ethyl acetate/pet ether). After completion of the reaction, the solvent was evaporated, quenched with water (20 mL), and extracted with ethyl acetate thrice. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 7b (3.05 g, 93% yield) as off-white solid.

m.p. 125–129°C. IR (KBr, cm−1): 3187, 2995, 1613, 1405, 1212, 1045. 1H NMR (400 MHz, CDCl3): δ = 11.10 (s, 1H), 8.20 (s, 1H), 7.15 (s, 1H), 6.9 (s, 1H), 3.75 (s, 6H), 2.2 (s, 3H). MS (EI): m/z 195 (M+, 100).

2.7. Experimental Procedure for the Preparation of Methyl 2-((tert-Butoxycarbonyl)amino)-3-(4-(3-(2,5-dimethoxyphenyl)isoxazol-5-yl)phenyl)propanoate (8a)

To a solution of compound 7a (0.150 g, 0.83 mmol) in dichloromethane (8 mL), compound 5a (0.27 g, 0.91 mmol), triethylamine (0.12 g, 1.24 mmol), and NaOCl (9–12% in H2O, 8 mL) were added at 0°C under nitrogen atmosphere. Then the reaction mixture was stirred at RT for 12 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). After completion of the reaction, the solvent was evaporated, quenched with water (20 mL), and extracted with dichloromethane. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 8a (0.28 g, 71% yield) as light yellow liquid.

IR (KBr, cm−1): 3304, 2970, 2939, 1712, 1627, 1500, 1276, 1222, 1170, 1045. 1H NMR (300 MHz, CDCl3): δ = 7.78 (d, J = 8.0 Hz, 2H), 7.59–7.48 (m, 1H), 7.24 (s, 2H), 7.04 (s, 1H), 6.98 (t, J = 1.8 Hz, 2H), 5.00 (s, 1H), 4.62 (s, 1H), 3.97–3.63 (m, 9H), 3.22 (d, J = 3.1 Hz, 2H), 1.42 (s, 9H). MS (EI): m/z 482 (M + 1, 100).

2.8. Experimental Procedure for the Preparation of Methyl 3-(4-(3-(2,5-Dimethoxyphenyl)isoxazol-5-yl)phenyl)-2-pivalamidopropanoate (8b)

To a solution of compound 7a (0.2 g, 1.11 mmol) in dichloromethane (10 mL), compound 5b (0.35 g, 1.22 mmol), triethylamine (0.16 g, 1.66 mmol), and NaOCl (9–12% in water, 10 mL) were added at 0°C under nitrogen atmosphere. Then the reaction mixture was stirred at RT for 12 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). After completion of the reaction, the solvent was evaporated, quenched with water (20 mL), and extracted with dichloromethane. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent gives the crude reaction mixture which was charged on silica gel column. The column was eluted with 25% ethyl acetate/pet ether to give the compound 8b (0.3 g, 58% yield) as light yellow liquid.

IR (KBr, cm−1): 3437, 2926, 1623, 1275, 1260, 764, 750. 1H NMR (300 MHz, CDCl3): δ = 7.82–7.73 (m, 2H), 7.55–7.47 (m, 1H), 7.24–7.16 (m, 2H), 7.05 (s, 1H), 6.98 (t, J = 1.9 Hz, 2H), 6.12 (d, J = 7.5 Hz, 1H), 4.90 (dt, J = 7.5, 5.8 Hz, 1H), 3.89 (s, 3H), 3.84 (s, 3H), 3.81–3.74 (m, 3H), 3.33–3.07 (m, 2H), 1.17 (s, 9H). 13C NMR (100 MHz, CDCl3): 177.9, 172.0, 169.0, 160.4, 153.6, 151.6, 138.0, 129.8, 126.6, 125.8, 118.4, 117.3, 114.9, 113.5, 113.0, 112.4, 100.9, 56.2, 55.8, 52.7, 52.4, 42.7, 38.7, 38.6, 37.7, 29.6. HRMS (ESI): Calcd for C26H31N2O6 [M + H]+: 467.2182; Found: 467.2455.

2.9. Experimental Procedure for the Preparation of Methyl 2-((tert-Butoxycarbonyl)amino)-3-(4-(3-(2,5-dimethoxy-4-methylphenyl)isoxazol-5-yl)phenyl)propanoate (8c)

To a solution of compound 7b (0.2 g, 1.02 mmol) in dichloromethane (10 mL), compound 5a (0.34 g, 1.12 mmol), triethylamine (0.15 g, 1.53 mmol), and NaOCl (9–12% in H2O, 10 mL) were added at 0°C under nitrogen atmosphere. Then the reaction mixture was stirred at RT for 12 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). After completion of the reaction, water (20 mL) was added and extracted with dichloromethane thrice. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. The solvent was evaporated to give the crude reaction mixture which was charged on silica gel column. Elution of the column with 20% ethyl acetate/pet ether gave the compound 8c (0.41 g, 80% yield) as off-white solid.

m.p 147–150°C. IR (KBr, cm−1): 3373, 3149, 2932, 1742, 1702, 1520, 1216, 1044. 1H NMR (300 MHz, CDCl3): δ = 7.78 (d, J = 7.8 Hz, 2H), 7.45 (s, 1H), 7.26 (d, J = 1.4 Hz, 2H), 7.06 (s, 1H), 6.85 (s, 1H), 5.03 (d, J = 8.4 Hz, 1H), 4.63 (q, J = 6.7 Hz, 1H), 3.87 (d, J = 4.6 Hz, 6H), 3.73 (s, 3H), 3.17 (td, J = 16.2, 15.0, 5.8 Hz, 2H), 2.28 (s, 3H), 1.42 (s, 9H). 13C NMR (100 MHz, CDCl3): 172.0, 168.9, 160.5, 155.0, 151.9, 151.1, 138.1, 130.0, 129.8, 126.6, 125.9, 115.3, 115.0, 110.3, 100.9, 80.0, 56.3, 55.9, 54.2, 52.3, 38.3, 28.2, 16.6. MS (EI) m/z 496 (M + 1, 100). HRMS (ESI): Calcd for C27H33N2O7 [M + H]+: 497.1882; Found: 497.2288.

2.10. Experimental Procedure for the Preparation of Methyl 3-(4-(3-(2,5-Dimethoxy-4-methylphenyl)isoxazol-5-yl)phenyl)-2-pivalamidopropanoate (8d)

To a solution of compound 7b (0.15 g, 0.76 mmol) in dichloromethane (10 mL), compound 5b (0.24 g, 0.84 mmol), triethylamine (0.116 g, 1.15 mmol), and NaOCl (9–12% in water, 10 mL) were added at 0°C under nitrogen atmosphere. Then the reaction mixture was stirred at RT for 12 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). After completion of the reaction, water (10 mL) was added and the reaction mixture was extracted with dichloromethane thrice. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. The solvent was evaporated to give the crude reaction which was charged on silica gel column. Elution of the column with 25% ethyl acetate/pet ether gave the compound 8d (0.31 g, 83% yield) as light yellow liquid.

IR (KBr, cm−1): 3444, 2929, 1637, 1473, 1275, 1261, 1215, 764. 1H NMR (300 MHz, DMSO): δ = 7.81 (dd, J = 13.0, 8.0 Hz, 3H), 7.41 (d, J = 8.1 Hz, 2H), 7.32 (d, J = 3.3 Hz, 2H), 7.07 (s, 1H), 4.51 (s, 1H), 3.85 (s, 3H), 3.80 (s, 3H), 3.64 (s, 3H), 3.24–2.96 (m, 2H), 2.20 (d, J = 19.4 Hz, 3H), 1.02 (s, 9H). MS (EI): m/z 480 (M + 1, 100).

2.11. Experimental Procedure for the Preparation of Methyl 2-((tert-Butoxycarbonyl)amino)-3-(4-(3-(3,6-dioxocyclohexa-1,4-dien-1-yl)isoxazol-5-yl)phenyl)propanoate (9a)

To a solution of compound 8a (0.15 g, 0.31 mmol) in acetonitrile (6 mL) and H2O (1 mL), CAN (0.511 g, 0.93 mmol) was added and the reaction mixture was stirred at RT for 1 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). After completion of the reaction, water (10 mL) was added and extracted with ethyl acetate thrice. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 9a (0.12 g, 85% yield) as yellow solid.

m.p. 125–127°C. IR (KBr, cm−1): 3355, 2979, 1743, 1720, 1654, 1522, 1288, 1251, 1167. 1H NMR (300 MHz, CDCl3): δ = 7.77 (d, J = 7.9 Hz, 2H), 7.48 (d, J = 1.9 Hz, 1H), 7.26 (d, J = 0.9 Hz, 2H), 7.12 (d, J = 0.9 Hz, 1H), 6.96–6.86 (m, 2H), 5.02 (s, 1H), 4.63 (s, 1H), 3.74 (d, J = 1.0 Hz, 3H), 3.29–3.02 (m, 2H), 1.42 (s, 9H). 13C NMR (100 MHz, CDCl3): 186.8, 185.2, 171.9, 170.7, 156.5, 154.9, 139.0, 136.8, 136.6, 134.4, 133.3, 130.0, 126.0, 125.7, 100.7, 80.1, 54.2, 52.3, 38.3, 28.2. HRMS (ESI): Calcd for C24H24N2O7 [M + H]+: 453.1662; Found: 453.1640.

2.12. Experimental Procedure for the Preparation of Methyl 3-(4-(3-(3,6-Dioxocyclohexa-1,4-dien-1-yl)isoxazol-5-yl)phenyl)-2-pivalamidopropanoate (9b)

To a solution of compound 8b (0.2 g, 0.42 mmol) in acetonitrile (8 mL) and H2O (2 mL), CAN (0.705 g, 1.28 mmol) was added and the reaction mixture was stirred at RT for 1 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). After completion of the reaction, water (20 mL) was added and extracted with ethyl acetate thrice. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 9b (0.136 g, 72% yield) as yellow solid.

m.p. 115–117°C. IR (KBr, cm−1): 3321, 2958, 1749, 1656, 1640, 1531, 1286, 1199, 1107. 1H NMR (300 MHz, CDCl3) δ = 7.76 (d, J = 7.9 Hz, 2H), 7.47 (d, J = 2.2 Hz, 1H), 7.23 (d, J = 7.9 Hz, 2H), 7.12 (s, 1H), 6.90 (d, J = 2.7 Hz, 2H), 6.13 (d, J = 7.7 Hz, 1H), 4.90 (q, J = 6.1 Hz, 1H), 3.77 (s, 3H), 3.37–3.04 (m, 2H), 1.17 (s, 9H). 13C NMR (100 MHz, CDCl3): 186.8, 185.1, 177.9, 172.0, 170.6, 156.5, 138.9, 136.8, 136.6, 134.4, 133.3, 130.0, 125.9, 125.7, 100.7, 52.7, 52.4, 38.6, 37.7, 27.3. HRMS (ESI): Calcd for C24H25N2O6 [M + H]+: 437.1713; Found: 437.1879.

2.13. Experimental Procedure for the Preparation of Methyl 2-((tert-Butoxycarbonyl)amino)-3-(4-(3-(4-methyl-3,6-dioxocyclohexa-1,4-dien-1-yl)isoxazol-5-yl)phenyl)propanoate (9c)

To a solution of compound 8c (0.23 g, 0.46 mmol) in acetonitrile (10 mL) and H2O (2 mL), CAN (0.76 g, 1.39 mmol) was added and the reaction mixture was stirred at RT for 1 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). After completion of the reaction, water (20 mL) was added and extracted with ethyl acetate thrice. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 9c (0.205 g, 95% yield) as yellow solid.

m.p. 153–155°C. IR (KBr, cm−1): 3364, 2979, 1732, 1693, 1660, 1524, 1252, 1170, 1020. 1H NMR (300 MHz, CDCl3): δ = 7.83–7.69 (m, 2H), 7.45 (s, 1H), 7.34–7.20 (m, 2H), 7.11 (s, 1H), 6.74 (q, J = 1.5 Hz, 1H), 5.04 (d, J = 8.1 Hz, 1H), 4.63 (q, J = 6.7 Hz, 1H), 3.74 (s, 3H), 3.29–3.00 (m, 2H), 2.13 (d, J = 1.6 Hz, 3H), 1.42 (s, 9H). 13C NMR (100 MHz, CDCl3): 187.4, 185.4, 171.9, 170.5, 156.5, 154.9, 146.2, 138.9, 134.3, 133.5, 133.4, 130.0, 126.0, 125.7, 100.8, 80.1, 54.2, 52.3, 38.3, 28.2, 15.5. HRMS (ESI): Calcd for C25H27N2O7 [M + H]+: 467.1818; Found: 467.1611.

2.14. Experimental Procedure for the Preparation of Methyl 3-(4-(3-(4-Methyl-3,6-dioxocyclohexa-1,4-dien-1-yl)isoxazol-5-yl)phenyl)-2-pivalamidopropanoate (9d)

To a solution of compound 8d (0.3 g, 0.62 mmol) in acetonitrile (12 mL) and H2O (3 mL), CAN (1.02 g, 1.86 mmol) was added and the reaction mixture was stirred at RT for 1 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). After completion of the reaction, water (20 mL) was added and extracted with ethyl acetate thrice. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 9d (0.25 g, 89% yield) as yellow solid.

m.p. 130–132°C. IR (KBr, cm−1): 3379, 2959, 2924, 1734, 1657, 1237, 1020, 807. 1H NMR (300 MHz, CDCl3): δ = 7.82–7.68 (m, 2H), 7.45 (d, J = 1.3 Hz, 1H), 7.22 (d, J = 7.9 Hz, 2H), 7.12 (d, J = 1.0 Hz, 1H), 6.74 (q, J = 1.4 Hz, 1H), 6.12 (d, J = 7.4 Hz, 1H), 4.90 (q, J = 6.2 Hz, 1H), 3.76 (d, J = 0.9 Hz, 3H), 3.34–3.05 (m, 2H), 2.18–2.07 (m, 3H), 1.17 (d, J = 0.9 Hz, 9H). 13C NMR (100 MHz, CDCl3): 187.3, 185.4, 177.9, 172.0, 170.4, 156.5, 146.2, 138.8, 134.2, 133.5, 133.4, 130.0, 125.9, 125.8, 100.9, 52.7, 52.4, 38.6, 37.7, 27.3, 15.5. HRMS (ESI): Calcd for C25H27N2O7 [M + H]+: 467.1818; Found: 467.1611.

2.15. Experimental Procedure for the Preparation of (3-(2,5-Dimethoxyphenyl)isoxazol-5-yl)methanol (15a)

To a solution of compound 7a (2 g, 11.11 mmol) in ethyl acetate (20 mL), compound 14 (2.01 g, 16.66 mmol), N-chlorosuccinamide (2.21 g, 16.66 mmol), and NaHCO3 (1.86 g, 22.22 mmol) were added and the reaction mixture was refluxed for 16 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). Then, water (20 mL) was added and the reaction mixture was extracted with ethyl acetate. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent gave the crude product which was purified by column chromatography to give the compound 15a (2.1 g, 80% yield) as white solid.

m.p. 69–73°C. IR (KBr, cm−1): 3330, 2943, 1709, 1510, 1295, 1225, 1036. 1H NMR (400 MHz, CDCl3): δ = 7.45 (d, J = 2.9 Hz, 1H), 7.01–6.90 (m, 2H), 6.79 (s, 1H), 4.82 (d, J = 6.3 Hz, 2H), 3.83 (d, J = 12.0 Hz, 6H), 2.19 (t, J = 6.5 Hz, 1H). MS (EI): m/z 235 (M + 1, 100).

2.16. Experimental Procedure for the Preparation of (3-(2,5-Dimethoxy-4-methylphenyl)isoxazol-5-yl)methanol (15b)

To a solution of compound 7b (1.0 g, 5.12 mmol) in ethyl acetate (20 mL), compound 14 (0.93 g, 7.69 mmol), N-chlorosuccinamide (1.02 g, 7.69 mmol), and NaHCO3 (0.861 g, 10.2 mmol) were added and refluxed for 16 hr. The progress of the reaction was monitored by TLC analysis (30% Ethyl acetate/pet ether). Then, water (20 mL) was added and the reaction mixture was extracted with ethyl acetate. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent gave the crude product which was purified by column chromatography to give the compound 15b (1.0 g, 90% yield) as off-white solid.

m.p. 58–62°C. IR (KBr, cm−1): 3426, 2940, 2129, 1715, 1216, 1038. 1H NMR (300 MHz, DMSO): δ = 11.05 (s, 1H), 7.27 (s, 1H), 7.05 (s, 1H), 6.74 (s, 1H), 4.60 (d, J = 0.8 Hz, 2H), 3.80 (d, J = 10.2 Hz, 6H), 2.21 (s, 3H). MS (EI): m/z 250 (M + 1, 100).

2.17. Experimental Procedure for the Preparation of 5-(Bromomethyl)-3-(2,5-dimethoxyphenyl)isoxazole (16a)

To a solution of compound 15a (1.5 g, 6.38 mmol) in dichloromethane (15 mL), phosphorous tribromide (2.59 g, 9.57 mmol) was added at 0°C under nitrogen atmosphere. Then the reaction mixture was stirred at RT for 16 hr. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). Then, water (10 mL) was added and the reaction mixture was extracted with dichloromethane. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. The solvent was evaporated and the crude reaction mixture was purified by column chromatography to give the compound 16a (1.2 g, 63% yield) as white solid.

m.p. 71–75°C. IR (KBr, cm−1): 3432, 2925, 1852, 1603, 1465, 1270, 1021. 1H NMR (400 MHz, CDCl3): δ = 7.47 (d, J = 3.0 Hz, 1H), 7.02–6.91 (m, 2H), 6.87 (s, 1H), 4.52 (s, 2H), 3.84 (d, J = 15.1 Hz, 6H). MS (EI): m/z 297 (M + 1, 100).

2.18. Experimental Procedure for the Preparation of 5-(Bromomethyl)-3-(2,5-dimethoxy-4-methylphenyl)isoxazole (16b)

To a solution of compound 15b (1.0 g, 4.0 mmol) in dichloromethane (20 mL), phosphorous tribromide (1.62 g, 6.0 mmol) was added at 0°C under nitrogen atmosphere. Then the reaction mixture was stirred at RT for 16 hr. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). Then, water (20 mL) was added and the reaction mixture was extracted with dichloromethane. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. The solvent was evaporated and the crude reaction mixture was purified by column chromatography to give the compound 16b (1.0 g, 80% yield) as brown solid.

m.p. 65–68°C. IR (KBr, cm−1): 3445, 2936, 1716, 1471, 1285, 1218, 1042. 1H NMR (400 MHz, CDCl3): δ = 7.39 (s, 1H), 6.82 (d, J = 6.4 Hz, 2H), 4.82 (dd, J = 6.5 Hz, 2H), 3.85 (s, 6H), 2.27 (s, 3H). MS (EI): m/z 311 (M + 1, 100).

2.19. Experimental Procedure for the Preparation of Methyl 3-(3-(2,5-Dimethoxyphenyl)isoxazol-5-yl)-2-((diphenylmethylene)amino)propanoate (17a)

To a solution of compound 10 (1.12 g, 4.44 mmol) in acetonitrile (20 mL), K2CO3 (2.78 g, 20.2 mmol) was added under nitrogen atmosphere and stirred at RT for 1 hr. Then compound 16a (1.2 g, 4.04 mmol) was added and the reaction mixture was refluxed for 16 hr. The progress of the reaction was monitored by TLC analysis (20% ethyl acetate/pet ether). Then, reaction mixture was filtered and filtrate was evaporated. The crude reaction mixture was purified by column chromatography to give the compound 17a (1.5 g, 74% yield) as light yellow liquid.

IR (KBr, cm−1): 3292, 2952, 1740, 1510, 1276, 1227, 1043. 1H NMR (300 MHz, CDCl3): δ = 7.63 (d, J = 6.9 Hz, 2H), 7.44–7.28 (m, 7H), 7.04–6.83 (m, 4H), 6.52 (s, 1H), 4.47 (dd, J = 7.5, 5.8 Hz, 1H), 3.78 (d, J = 8.5 Hz, 6H), 3.59 (s, 3H), 3.53–3.41 (m, 2H). MS (EI): m/z 470 (M + 1, 100).

2.20. Experimental Procedure for the Preparation of Methyl 3-(3-(2,5-Dimethoxy-4-methylphenyl)isoxazol-5-yl)-2-((diphenylmethylene)amino)propanoate (17b)

To a solution of compound 10 (0.9 g, 3.55 mmol) in acetonitrile (20 mL), K2CO3 (2.45 g, 17.7 mmol) was added under nitrogen atmosphere and stirred at RT for 1 hr. Then compound 16b (1.21 g, 3.91 mmol) was added and the reaction mixture was refluxed for 16 hr. The progress of the reaction was monitored by TLC analysis (20% ethyl acetate/pet ether). Then, reaction mixture was filtered and filtrate was evaporated. The crude reaction mixture was purified by column chromatography to give the compound 17b (1.1 g, 69% yield) as off-white solid.

m.p. 142–146°C. IR (KBr, cm−1): 3447, 2949, 1736, 1284, 1213, 1041. 1H NMR (300 MHz, CDCl3): δ = 7.63 (d, J = 8.2 Hz, 2H), 7.49–7.24 (m, 7H), 7.04–6.92 (m, 2H), 6.75 (s, 1H), 6.53 (s, 1H), 4.61–4.36 (m, 1H), 3.79 (dd, J = 16.5 Hz, 6H), 3.59 (d, J = 0.9 Hz, 3H), 3.54–3.35 (m, 2H), 2.24 (s, 3H). MS (EI): m/z 484 (M + 1, 100).

2.21. Experimental Procedure for the Preparation of Methyl 2-Amino-3-(3-(2,5-dimethoxyphenyl)isoxazol-5-yl)propanoate (18a)

To a solution of compound 17a (1.5 g, 3.19 mmol) in diethyl ether (20 mL), 1 M HCl (20 mL) was added at 0°C. Then the reaction mixture was stirred at RT for 16 hr. The progress of the reaction was monitored by TLC analysis (10% methanol/chloroform). The layers were separated and the aqueous layer was basified with aqueous ammonia until PH 10 and extracted with ethyl acetate. The combined organic layers were washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent gave the compound 18a (0.850 g, 87% yield) as pale yellow liquid.

IR (KBr, cm−1): 3383, 2953, 1738, 1602, 1471, 1227, 1042. 1H NMR (300 MHz, DMSO): δ = 7.27 (d, J = 3.0 Hz, 1H), 7.17–6.97 (m, 2H), 6.67 (s, 1H), 3.78 (d, J = 17.1 Hz, 7H), 3.64 (s, 3H), 3.07 (qd, J = 15.1, 6.6 Hz, 2H), 1.99 (s, 2H). MS (EI): m/z 306 (M + 1, 100).

2.22. Experimental Procedure for the Preparation of Methyl 2-Amino-3-(3-(2,5-dimethoxy-4-methylphenyl)isoxazol-5-yl)propanoate (18b)

To a solution of compound 17b (0.5 g, 1.03 mmol) in diethyl ether (10 mL), 1 M HCl (10 mL) was added at 0°C. Then the reaction mixture was stirred at RT for 16 hr. The progress of the reaction was monitored by TLC analysis (10% methanol/chloroform). The layers were separated and the aqueous layer was basified with aqueous ammonia until PH 10 and extracted with ethyl acetate. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent gave the compound 18b (0.27 g, 82% yield) as off-white solid.

m.p. 218–221°C. IR (KBr, cm−1): 3468, 2838, 1741, 1472, 1250, 1220, 1041. 1H NMR (300 MHz, DMSO): δ = 8.58 (s, 2H), 7.28 (s, 1H), 7.06 (s, 1H), 6.82 (s, 1H), 4.53 (t, J = 6.1 Hz, 1H), 3.93–3.65 (m, 9H), 3.41 (d, J = 6.2 Hz, 2H), 2.22 (s, 3H). MS (EI): m/z 320 (M + 1, 100).

2.23. Experimental Procedure for the Preparation of Methyl 2-((tert-Butoxycarbonyl)amino)-3-(3-(2,5-dimethoxyphenyl)isoxazol-5-yl)propanoate (19a)

To a solution of compound 18a (0.300 g, 0.98 mmol) in dichloromethane (15 mL), triethylamine (0.19 g, 1.98 mmol) was added. Then (Boc)2O (0.23 g, 1.07 mmol) was added and the reaction mixture was stirred at RT for 16 hr. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). Then, water (10 mL) was added and the reaction mixture was extracted with dichloromethane. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent gave the crude compound which was purified by column chromatography to give the compound 19a (0.380 g, 95% yield) as light brown solid.

m.p. 110–113°C. IR (KBr, cm−1): 3372, 2948, 1742, 1690, 1524, 1274, 1220, 1025. 1H NMR (300 MHz, CDCl3): δ = 7.44 (d, J = 2.8 Hz, 1H), 7.01–6.88 (m, 2H), 6.62 (s, 1H), 5.24 (s, 1H), 4.70 (s, 1H), 3.90–3.72 (m, 9H), 3.37 (d, J = 6.0 Hz, 2H), 1.44 (s, 9H). 13C NMR (100 MHz, CDCl3): 171.2, 167.4, 159.9, 155.0, 153.6, 151.5, 118.2, 117.3, 113.4, 113.0, 104.5, 80.2, 56.1, 55.8, 52.6, 52.1, 29.7, 28.2. MS (EI): m/z 406 (M + 1, 100). HRMS (ESI) Calcd for C20H26N2O7 [M + H]+: 407.1818; Found: 407.1743.

2.24. Experimental Procedure for the Preparation of Methyl 3-(3-(2,5-Dimethoxyphenyl)isoxazol-5-yl)-2-pivalamidopropanoate (19b)

To a solution of compound 18a (0.300 g, 0.98 mmol) in DCM (15 mL), dimethylaminopyridine (0.012 g, 0.098 mmol) was added. Then pivaloyl chloride (0.356 g, 2.94 mmol) was added and the reaction mixture was stirred at RT for 16 hr. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). Then, water (10 mL) was added and the reaction mixture was extracted with dichloromethane. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent gave the crude compound which was purified by column chromatography to give the compound 19b (0.300 g, 78% yield) as light yellow liquid.

IR (KBr, cm−1): 3366, 2959, 1745, 1651, 1511, 1267, 1227, 1043. 1H NMR (300 MHz, CDCl3): δ = 7.44 (d, J = 2.8 Hz, 1H), 7.03–6.85 (m, 2H), 6.59 (d, J = 2.3 Hz, 1H), 6.38 (d, J = 7.0 Hz, 1H), 5.00–4.80 (m, 1H), 3.95–3.70 (m, 9H), 3.56–3.28 (m, 2H), 1.22 (d, J = 2.3 Hz, 9H). 13C NMR (100 MHz, CDCl3): 178.2, 171.1, 167.3, 159.9, 153.6, 151.5, 118.1, 117.4, 113.3, 112.9, 104.7, 56.0, 55.8, 52.8, 50.8, 38.6, 28.9, 27.3, 27.0. MS (EI): m/z 390 (M + 1, 100).

2.25. Experimental Procedure for the Preparation of Methyl 2-((tert-Butoxycarbonyl)amino)-3-(3-(2,5-dimethoxy-4-methylphenyl)isoxazol-5-yl)propanoate (19c)

To a solution of compound 18b (0.2 g, 0.62 mmol) in dichloromethane (10 mL), triethyl amine (0.12 g, 1.25 mmol) was added. Then (Boc)2O (0.15 g, 0.68 mmol) was added and the reaction mixture was stirred at RT for 16 hr. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). Then, water (10 mL) was added and the reaction mixture was extracted with dichloromethane. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent gave the crude compound which was purified by column chromatography to give the compound 19c (0.25 g, 95% yield) as off-white solid.

m.p. 103–107°C. IR (KBr, cm−1): 3344, 2928, 2846, 1733, 1677, 1526, 1219, 1048. 1H NMR (300 MHz, DMSO): δ = 7.47 (d, J = 8.3 Hz, 1H), 7.26 (s, 1H), 7.04 (s, 1H), 6.68 (d, J = 4.3 Hz, 1H), 4.39 (t, J = 9.2 Hz, 1H), 3.91–3.73 (m, 6H), 3.65 (d, J = 10.3 Hz, 3H), 3.28–3.04 (m, 2H), 2.21 (s, 3H), 1.35 (s, 9H). 13C NMR (100 MHz, DMSO-d6): 171.9, 168.7, 159.2, 155.2, 151.2, 150.7, 129.3, 115.3, 114.6, 109.6, 109.5, 103.6, 78.5, 56.1, 55.5, 52.1, 51.9, 28.0, 26.8, 16.2. MS (EI): m/z 420 (M + 1, 100). HRMS (ESI) Calcd for C21H29N2O7 [M + H]+: 421.1975; Found: 421.1988.

2.26. Experimental Procedure for the Preparation of Methyl 3-(3-(2,5-Dimethoxy-4-methylphenyl)isoxazol-5-yl)-2-pivalamidopropanoate (19d)

To a solution of compound 18b (0.3 g, 0.93 mmol) in dichloromethane (10 mL), dimethylaminopyridine (0.011 g, 0.093 mmol) was added. Then pivaloyl chloride (0.34 g, 2.81 mmol) was added and the reaction mixture was stirred at RT for 16 hr. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). Then, water (10 mL) was added and the reaction mixture was extracted with dichloromethane. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent gave the crude compound which was purified by column chromatography to give the compound 19d (0.31 g, 82% yield) as off-white solid.

m.p. 116–120°C. IR (KBr, cm−1): 3323, 2963, 1741, 1531, 1431, 1217, 1041. 1H NMR (300 MHz, DMSO): δ = 7.98 (d, J = 8.0 Hz, 1H), 7.25 (s, 1H), 7.03 (s, 1H), 6.65 (s, 1H), 4.61 (td, J = 8.2, 6.6 Hz, 1H), 3.78 (d, J = 5.9 Hz, 6H), 3.66 (s, 3H), 3.38–3.24 (m, 2H), 2.21 (s, 3H), 1.07 (s, 9H). 13C NMR (100 MHz, DMSO-d6): 177.5, 171.2, 169.0, 159.2, 151.2, 150.7, 129.2, 115.3, 114.7, 109.6, 103.6, 56.0, 55.5, 52.1, 50.4, 37.8, 27.5, 27.0, 16.2. MS (EI): m/z 404 (M + 1, 100). HRMS (ESI): Calcd for C21H29N2O6 [M + H]+: 405.2026; Found: 405.1703.

2.27. Experimental Procedure for the Preparation of Methyl 2-((tert-Butoxycarbonyl)amino)-3-(3-(3,6-dioxocyclohexa-1,4-dien-1-yl)isoxazol-5-yl)propanoate (20a)

To a solution of compound 19a (0.250 g, 0.61 mmol) in acetonitrile (10 mL) and H2O (2 mL), CAN (1.01 g, 1.84 mmol) was added and the reaction mixture was stirred at RT for 1 h. The progress of the reaction was monitored by TLC analysis (20% ethyl acetate/Pet ether). Then, water (10 mL) was added and extracted with ethyl acetate. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 20a (0.150 g, 64% yield) as yellow solid.

m.p. 110–112°C. IR (KBr, cm−1): 3359, 2979, 2955, 1749, 1688, 1524, 1283, 1163. 1H NMR (300 MHz, CDCl3): δ = 7.44–7.36 (m, 1H), 6.87 (d, J = 1.5 Hz, 2H), 6.69 (s, 1H), 5.22 (s, 1H), 4.68 (s, 1H), 3.80 (s, 3H), 3.53–3.25 (m, 2H), 1.44 (s, 9H). 13C NMR (100 MHz, CDCl3): 186.7, 185.0, 170.9, 169.3, 156.1, 154.9, 136.7, 136.5, 134.4, 133.3, 104.4, 80.4, 52.8, 52.0, 29.8, 28.2. MS (EI): m/z 376 (M + 1, 100). HRMS (ESI): Calcd for C18H20N2O7 [M + H]+: 377.1349; Found: 377.0531.

2.28. Experimental Procedure for the Preparation of Methyl 3-(3-(3,6-Dioxocyclohexa-1,4-dien-1-yl)isoxazol-5-yl)-2-pivalamidopropanoate (20b)

To a solution of compound 19b (0.2 g, 0.51 mmol) in acetonitrile (10 mL) and H2O (2 mL), CAN (0.560 g, 1.025 mmol) was added and the reaction mixture was stirred at RT for 1 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). Then, water (10 mL) was added and extracted with ethyl acetate. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 20b (0.160 g, 86% yield) as yellow solid.

m.p. 107–109°C. IR (KBr, cm−1): 3312, 2961, 2872, 1752, 1662, 1639, 1534, 1287, 1206, 1093. 1H NMR (400 MHz, CDCl3): δ = 7.39 (s, 1H), 6.87 (s, 2H), 6.65 (s, 1H), 6.36 (d, J = 7.1 Hz, 1H), 4.90 (q, J = 5.7 Hz, 1H), 3.82 (s, 3H), 3.53 (dd, J = 15.4, 5.4 Hz, 1H), 3.37 (dd, J = 15.3, 5.3 Hz, 1H), 1.22 (s, 9H). 13C NMR (100 MHz, CDCl3): 186.7, 184.9, 178.2, 170.9, 169.2, 156.0, 136.7, 136.5, 134.3, 133.3, 104.5, 52.9, 50.8, 38.7, 29.0, 27.3. MS (EI): m/z 360 (M + 1, 100). HRMS (ESI): Calcd for C18H21N2O6 [M + H]+: 361.1400; Found: 361.1388.

2.29. Experimental Procedure for the Preparation of Methyl 2-((tert-Butoxycarbonyl)amino)-3-(3-(4-methyl-3,6-dioxocyclohexa-1,4-dien-1-yl)isoxazol-5-yl)propanoate (20c)

To a solution of compound 19c (0.2 g, 0.47 mmol) in acetonitrile (10 mL) and H2O (2 mL), CAN (0.52 g, 0.95 mmol) was added and the reaction mixture was stirred at RT for 1 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). Then, water (10 mL) was added and extracted with ethyl acetate. The combined organic layer was washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 20c (0.185 g, 86% yield) as yellow solid.

m.p. 89–93°C. IR (KBr, cm−1): 3379, 2977, 1742, 1695, 1654, 1524, 1239, 1170, 1040. 1H NMR (300 MHz, CDCl3): δ = 7.36 (s, 1H), 6.74–6.62 (m, 2H), 5.23 (d, J = 6.3 Hz, 1H), 4.68 (s, 1H), 3.80 (s, 3H), 3.39 (m, 2H), 2.25–1.99 (m, 3H), 1.44 (s, 9H). 13C NMR (100 MHz, CDCl3): 187.3, 185.2, 170.9, 169.1, 156.1, 154.9, 146.2, 134.2, 133.5, 133.4, 109.9, 104.5, 52.8, 52.0, 29.8, 28.2, 15.5. MS (EI): m/z 390 (M + 1, 100). HRMS (ESI): Calcd for C19H23N2O7 [M + H]+: 391.1505; Found: 391.1601.

2.30. Experimental Procedure for the Preparation of Methyl 3-(3-(4-Methyl-3,6-dioxocyclohexa-1,4-dien-1-yl)isoxazol-5-yl)-2-pivalamidopropanoate (20d)

To a solution of compound 19d (0.2 g, 0.49 mmol) in acetonitrile (10 mL) and H2O (2 mL), CAN (0.54 g, 0.99 mmol) was added and the reaction mixture was stirred at RT for 1 h. The progress of the reaction was monitored by TLC analysis (30% ethyl acetate/pet ether). After completion of the reaction, water (10 mL) was added and extracted with ethyl acetate thrice. The organic layers were combined, washed with water, brined, and dried over anhydrous Na2SO4. Evaporation of the solvent in high vacuum gave the compound 20d (0.175 g, 94% yield) as yellow solid.

m.p.118–120°C. IR (KBr, cm−1): 3351, 2958, 2872, 1735, 1657, 1524, 1230, 1042. 1H NMR (300 MHz, CDCl3): δ =7.36 (s, 1H), 6.70 (q, J = 1.6 Hz, 1H), 6.68 (s, 1H), 6.35 (d, J = 7.3 Hz, 1H), 4.89 (dt, J = 7.2, 5.2 Hz, 1H), 3.82 (s, 3H), 3.59–3.44 (m, 1H), 3.36 (dd, J = 15.2, 5.2 Hz, 1H), 2.11 (d, J = 1.7 Hz, 3H), 1.21 (s, 9H). 13C NMR (100 MHz, CDCl3): 187.3, 185.2, 178.2, 171.0, 169.0, 156.1, 146.2, 134.1, 133.5, 133.4, 104.6, 52.9, 50.8, 38.7, 29.0, 27.3, 15.5. MS (EI): m/z 374 (M + 1, 100). HRMS (ESI): Calcd for C19H23N2O6 [M + H]+: 375.1556; Found: 375.1468.

3. Results and Discussion

Our proposed strategy was based on a simple and lucid cycloaddition reaction [4245] of alkyne 5 with oxime 7 to prepare isoxazole-amino acid hybrids 8 (Scheme 1). The alkyne 5 was prepared in five steps using the protocol developed by O’Donnell et al. starting from N-(diphenylmethylene) glycine methyl ester [46, 47] using appropriate benzyl or propargyl halides. Subsequent oxidation of the suitable placed methoxy group in the aromatic ring of compound 8 will provide isoxazole tethered quinone-amino acid hybrids 9.

721291.sch.001
Scheme 1: Synthesis of isoxazole tethered quinone phenylalanine hybrids.

To start with, the compound 3a or 3b was prepared according to literature procedure [48] starting from 4-bromobenzyl bromide. Then the compound 3a or 3b reacted with TMS-acetylene in the presence of CuI/Et3N/PdCl2(PPh3)2 in reflux condition to give compound 4a or 4b (Scheme 1). Then TMS group was deprotected using TBAF to give the key acetylenic amino acid ready for cycloaddition reaction. 2,5-Dimethoxy benzaldehyde 6a or 6b reacted with hydroxylamine hydrochloride (NH2OH·HCl) to produce the oxime derivative 7a or 7b. The compound 7a or 7b was subjected to the key 1,3-dipolar cycloaddition reaction with actetylenic amino acid 5a or 5b in the presence of NaOCl/Et3N in DCM as solvent. The isoxazoles 8a–d were smoothly formed using this Huisgen’s one pot protocol. We did not observe the formation of any other isomer and nitrile oxide dimerization product in this reaction. Any undesired by-products resulting from aromatic halogenations reaction were not observed in our case.

Then the compound 8a was oxidized with CAN to give the desired isoxazole tethered quinone-amino acid 9a in very good yield. The compound 9a was characterized by 1H-NMR, 13C-NMR, and HRMS. For example, the characteristic isoxazole proton at 7.1δ and quinone proton at 6.8δ in 1H-NMR confirms the oxidation of compound 8a to generate compound 9a. The two carbonyl peaks at 186.8 and 185.2δ in 13C-NMR spectrum validate the benzoquinone moiety. Using similar sequence (Scheme 1) the target compounds 9b–d (Table 1) were prepared and characterized by spectral data. The isoxazole tethered quinone-amino acids show equilibrium between hydroquinone (9ah) and benzoquinone (9a) in liquid chromatography mass spectrometry (LC-MS) analysis condition as shown in Figure 3. It may be possible that benzoquinone forms a reduced species during the ionization process in LC-MS condition [49].

tab1
Table 1: The yields and purity of the novel isoxazole-phenyl alanine and isoxazole tethered quinone-phenyl alanine hybrids.
721291.fig.003
Figure 3: LC-MS analysis of compound 9a (showing the integrated percentage).

Encouraged by this result, we turn our attention to the preparation of propargyl amino acid as starting material. Various acetylene building blocks containing an amino acid moiety were prepared from Schiff-base N-(diphenylmethylene)glycine ester 10 using the literature procedure [50]. Thus, alkylation of 10 with propargyl bromide 11 in the presence of K2CO3/CH3CN in reflux condition gave the propargylated derivative 12. Then the compound 12 was subjected to 1,3-dipolar cycloaddition reaction with the oxime 7a in the presence of NCS/NaHCO3 in ethyl acetate as solvent to give very low yield of the compound 17a (Scheme 2). We tried couple of conditions to improve the yield of compound 17a but without any success. Maybe either steric factor or the instability of compound 12 is the main reason behind the low yield of the cycloaddition product. Then we turn our attention to the compound 13 which was prepared from compound 12 by hydrolysis reaction followed by protection of the amino group. We tried several conditions (Table 2) to improve the yield of the compound 19b without any success.

tab2
Table 2: The yields of the compound 19b using various 1,3-cycloaddition reaction conditions.
721291.sch.002
Scheme 2: Synthesis of 3,5-disubstituted isoxazole derivative.

Alternatively we plan to introduce the amino acid at the end of reaction sequence to prepare the isoxazole tethered quinone-amino acid hybrid (Scheme 3). Thus, the oxime derivative 7a or 7b was subjected to cycloaddition reaction with propargyl alcohol 14 using NCS/NaHCO3 condition in ethyl acetate as solvent. Under this reaction condition, the 3-aryl-isoxazole derivative 15a or 15b was prepared in good yield. Then the hydroxyl methyl group in the compound 15a or 15b was converted to bromo methyl group using PBr3/DCM condition to give the compound 16a or 16b.

721291.sch.003
Scheme 3: Synthesis of isoxazole tethered glycine quinone hybrids.

Gratifyingly, the key step alkylation reaction of 10 with compound 16a in the presence of K2CO3/CH3CN in reflux condition gave the 3,5-substituted isoxazole derivative 17a or 17b. The compound 17a was hydrolyzed in the presence of 1 N HCl/diethyl ether and the resulting amino ester 18a was protected with either Boc2O or pivaloyl chloride to obtain Boc derivative 19a or pivaloyl derivative 19b,  respectively. Then the compound 19a was oxidized with CAN to give the desired target isoxazole tethered quinone amino acid 20a in very good yield. The compound 20a was characterized by 1H-NMR, 13C-NMR, and HRMS spectral data. In a similar sequence the target compounds 20b–d were prepared and characterized by spectral data. The purity of the final isoxazole tethered quinone amino acids (Table 3) was obtained by LC-MS analysis which showed equilibrium between quinone and hydroquinone as observed earlier (Figure 3).

tab3
Table 3: The yields and purity of the novel isoxazole-glycine and isoxazole tethered quinone-glycine hybrids.

It is noteworthy to mention here that previously inaccessible quinone amino acids (19a–d and 20a–d) containing isoxazole moiety were synthesized in very good yield. As indicated in Table 3, the proting group (Boc or pivaloyl) has no effect on the yield of cycloaddition as well as oxidation reaction to give the isoxazole tethered quinone amino acids.

In conclusion, we have developed an efficient and simple method to synthesize isoxazole tethered quinone-amino acid hybrids using 1,3-dipolar cycloaddition and oxidation reactions as key steps in good yields. We believe that this methodology will find a widespread application for the synthesis of 2-aryl-benzoquinone and its derivatives. Further application of this methodology for the synthesis of isoxazole tethered quinone-peptide hybrid as well as preparation of tetrazole tethered quinone-amino acid hybrid is undergoing in our group.

Conflict of Interests

The authors declare that there is no conflict of interests regarding the publication of this paper.

Acknowledgments

The authors are grateful to GVK Biosciences Pvt. Ltd., for the financial support and encouragement. Help from the analytical department for the analytical data is appreciated. The authors thank Dr. Sudhir Kumar Singh for his invaluable support and motivation.

References

  1. P. M. Dewick, Medicinal Natural Products, Chichester, UK, Wiley, 2nd edition, 2002.
  2. Y. Kimura, “New anticancer agents: In vitro and in vivo evaluation of the antitumor and antimetastatic actions of various compounds isolated from medicinal plants,” In Vivo, vol. 19, no. 1, pp. 37–60, 2005. View at Google Scholar · View at Scopus
  3. B. Nowicka and J. Kruk, “Occurrence, biosynthesis and function of isoprenoid quinones,” Biochimica et Biophysica Acta, vol. 1797, no. 9, pp. 1587–1605, 2010. View at Publisher · View at Google Scholar · View at Scopus
  4. Z. S. Saify, N. Mushtaq, F. Noor, S. Takween, and M. Arif, “Role of quinone moiety as antitumour agtents: a review,” Pakistan Journal of Pharmaceutical Sciences, vol. 12, no. 2, pp. 21–31, 1999. View at Google Scholar
  5. D. R. A. Mans, J. Retel, J. M. S. van Maanen et al., “Role of the semi-quinone free radical of the anti-tumour agent etoposide (VP-16-213) in the inactivation of single- and double-stranded ΦX174 DNA,” British Journal of Cancer, vol. 62, no. 1, pp. 54–60, 1990. View at Publisher · View at Google Scholar · View at Scopus
  6. K. Ishiguro, K. Takahashi, K. Yazawa, S. Sakiyama, and T. Arai, “Binding of saframycin A, a heterocyclic quinone anti-tumor antibiotic to DNA as revealed by the use of the antibiotic labeled with [14C]tyrosine or [14C]cyanide,” The Journal of Biological Chemistry, vol. 256, no. 5, pp. 2162–2167, 1981. View at Google Scholar · View at Scopus
  7. D. L. Boger, M. Yasuda, L. A. Mitscher, S. D. Drake, P. A. Kitos, and S. C. Thompson, “Streptonigrin and lavendamycin partial structures. Probes for the minimum, potent pharmacophore of streptonigrin, lavendamycin, and synthetic quinoline-5,8-diones,” Journal of Medicinal Chemistry, vol. 30, no. 10, pp. 1918–1928, 1987. View at Publisher · View at Google Scholar · View at Scopus
  8. W. J. Pigram, W. Fuller, and L. D. Hamilton, “Stereochemistry of intercalation: interaction of daunomycin with DNA,” Nature: New Biology, vol. 235, no. 7, pp. 17–19, 1972. View at Google Scholar · View at Scopus
  9. V. N. Iyer and W. Szybalski, “Mitomycins and porfiromycin: chemical mechanism of activation and cross-linking of DNA,” Science, vol. 145, no. 3627, pp. 55–58, 1964. View at Publisher · View at Google Scholar · View at Scopus
  10. R. Cone, S. K. Hasan, J. W. Lown, and A. R. Morgan, “The mechanism of the degradation of DNA by streptonigrin,” Canadian Journal of Biochemistry, vol. 54, no. 3, pp. 219–223, 1976. View at Publisher · View at Google Scholar · View at Scopus
  11. P. Conti, M. de Amici, G. Grazioso et al., “Synthesis, binding affinity at glutamic acid receptors, neuroprotective effects, and molecular modeling investigation of novel dihydroisoxazole amino acids,” Journal of Medicinal Chemistry, vol. 48, no. 20, pp. 6315–6325, 2005. View at Publisher · View at Google Scholar · View at Scopus
  12. S. Srivastava, L. K. Bajpai, S. Batra et al., “In search of new chemical entities with spermicidal and anti-HIV activities,” Bioorganic and Medicinal Chemistry, vol. 7, no. 11, pp. 2607–2613, 1999. View at Publisher · View at Google Scholar · View at Scopus
  13. A. Kumar, R. A. Maurya, S. Sharma et al., “Design and synthesis of 3,5-diarylisoxazole derivatives as novel class of anti-hyperglycemic and lipid lowering agents,” Bioorganic and Medicinal Chemistry, vol. 17, no. 14, pp. 5285–5292, 2009. View at Publisher · View at Google Scholar · View at Scopus
  14. T. Karabasanagouda, A. V. Adhikari, and M. Girisha, “Synthesis of some new pyrazolines and isoxazoles carrying 4-methylthiophenyl moiety as potential analgesic and antiinflammatory agents,” Indian Journal of Chemistry—Section B Organic and Medicinal Chemistry, vol. 48, no. 3, pp. 430–437, 2009. View at Google Scholar · View at Scopus
  15. B. R. Dravyakar, D. P. Kawade, P. B. Khedekar, and K. P. Bhusari, “Design and syntheses of some new diphenylaminoisoxazolines as potent anti-inflammatory agent,” Indian Journal of Chemistry B: Organic and Medicinal Chemistry, vol. 47, no. 10, pp. 1559–1567, 2008. View at Google Scholar · View at Scopus
  16. S. B. Jadhav, R. A. Shastri, K. V. Gaikwad, and S. V. Gaikwad, “Synthesis and antimicrobial studies of some novel pyrazoline and isoxazoline derivatives,” E-Journal of Chemistry, vol. 6, supplement 1, pp. S183–S188, 2009. View at Publisher · View at Google Scholar · View at Scopus
  17. J. J. Talley, D. L. Brown, J. S. Carter et al., “4-[5-Methyl-3-phenylisoxazol-4-yl]-benzenesulfonamide, Valdecoxib: a potent and selective inhibitor of COX-2,” Journal of Medicinal Chemistry, vol. 43, no. 5, pp. 775–777, 2000. View at Publisher · View at Google Scholar · View at Scopus
  18. T. M. V. D. Pinho e Melo, “Recent advances on the synthesis and reactivity of isoxazoles,” Current Organic Chemistry, vol. 9, no. 10, pp. 925–958, 2005. View at Publisher · View at Google Scholar · View at Scopus
  19. A. R. Katritzky, M. A. C. Button, and S. N. Denisenko, “Efficient synthesis of 3,5-functionalized isoxazoles and isoxazolines via 1,3-dipolar cycloaddition reactions of 1-propargyl- and 1-allylbenzotriazoles,” Journal of Heterocyclic Chemistry, vol. 37, no. 6, pp. 1505–1510, 2000. View at Publisher · View at Google Scholar · View at Scopus
  20. S. Kotha, “The building block approach to unusual α-amino acid derivatives and peptides,” Accounts of Chemical Research, vol. 36, no. 5, pp. 342–351, 2003. View at Publisher · View at Google Scholar · View at Scopus
  21. D. A. Dougherty, “Unnatural amino acids as probes of protein structure and function,” Current Opinion in Chemical Biology, vol. 4, no. 6, pp. 645–652, 2000. View at Publisher · View at Google Scholar · View at Scopus
  22. M. J. O'Donnell, “The preparation of optically active α-amino acids from the benzophenone imines of glycine derivatives,” Aldrichimica Acta, vol. 34, no. 1, pp. 3–15, 2001. View at Google Scholar · View at Scopus
  23. T. Hashimoto and K. Maruoka, “Recent development and application of chiral phase-transfer catalysts,” Chemical Reviews, vol. 107, no. 12, pp. 5656–5682, 2007. View at Publisher · View at Google Scholar · View at Scopus
  24. L. F. Tietze, H. P. Bell, and S. Chandrasekhar, “Natural product hybrids as new leads for drug discovery,” Angewandte Chemie, vol. 42, no. 34, pp. 3996–4028, 2003. View at Publisher · View at Google Scholar · View at Scopus
  25. G. Mehta and V. Singh, “Hybrid systems through natural product leads: an approach towards new molecular entities,” Chemical Society Reviews, vol. 31, no. 6, pp. 324–334, 2002. View at Publisher · View at Google Scholar · View at Scopus
  26. M. Decker, “Hybrid molecules incorporating natural products: applications in cancer therapy, neurodegenerative disorders and beyond,” Current Medicinal Chemistry, vol. 18, no. 10, pp. 1464–1475, 2011. View at Publisher · View at Google Scholar · View at Scopus
  27. S. Kotha, K. Mandal, S. Banerjee, and S. M. Mobin, “Synthesis of novel quinone-amino acid hybrids via cross-enyne metathesis and Diels-Alder reaction as key steps,” European Journal of Organic Chemistry, no. 8, pp. 1244–1255, 2007. View at Publisher · View at Google Scholar · View at Scopus
  28. K. P. Kaliappan and V. Ravikumar, “Design and synthesis of novel sugar-oxasteroid-quinone hybrids,” Organic and Biomolecular Chemistry, vol. 3, no. 5, pp. 848–851, 2005. View at Publisher · View at Google Scholar · View at Scopus
  29. S. Hoppen, U. Emde, T. Friedrich, L. Grubert, and U. Koert, “Natural product hybrids: design, synthesis and biological evaluation of quinone-annonaceous acetogenins,” Angewandte Chemie International Edition in English, vol. 39, no. 12, pp. 2099–2102, 2000. View at Publisher · View at Google Scholar
  30. G. Mehta and S. S. Ramesh, “Polycyclitols Novel conduritol and carbasugar hybrids as new glycosidase inhibitors,” Canadian Journal of Chemistry, vol. 83, no. 6-7, pp. 581–594, 2005. View at Publisher · View at Google Scholar
  31. P. R. Kumar, M. Behera, K. Raghavulu, A. J. Shree, and S. Yennam, “Synthesis of novel isoxazole-benzoquinone hybrids via 1,3-dipolar cycloaddition reaction as key step,” Tetrahedron Letters, vol. 53, no. 32, pp. 4108–4113, 2012. View at Publisher · View at Google Scholar · View at Scopus
  32. D. J. Burkhart, B. Twamley, and N. R. Natale, “A new direct synthesis of ACPA and novel AMPA analogues,” Tetrahedron Letters, vol. 42, no. 48, pp. 8415–8418, 2001. View at Publisher · View at Google Scholar · View at Scopus
  33. D. Schulz, P. Beese, B. Ohlendorf et al., “Abenquines A-D: aminoquinone derivatives produced by Streptomyces sp. strain DB634,” Journal of Antibiotics, vol. 64, no. 12, pp. 763–768, 2011. View at Publisher · View at Google Scholar · View at Scopus
  34. J. Sakaki, T. Murata, Y. Yuumoto et al., “Discovery of IRL 3461: a novel and potent endothelin antagonist with balanced ETA/ETB affinity,” Bioorganic and Medicinal Chemistry Letters, vol. 8, no. 16, pp. 2241–2246, 1998. View at Publisher · View at Google Scholar · View at Scopus
  35. A. R. Katritzky, L. Huang, and R. Sakhuja, “Efficient syntheses of naphthoquinone-dipeptides,” Synthesis, no. 12, Article ID M06709SS, pp. 2011–2016, 2010. View at Publisher · View at Google Scholar · View at Scopus
  36. O. Moriya, H. Takenaka, M. Iyoda, Y. Urata, and T. Endo, “Generation of nitrite oxides via O-tributylstannyl aldoximes, application to the synthesis of isoxazolines and isoxazoles,” Journal of the Chemical Society Perkin Transactions I, no. 4, pp. 413–417, 1994. View at Google Scholar
  37. F. Himo, T. Lovell, R. Hilgraf et al., “Copper(I)-catalyzed synthesis of azoles. DFT study predicts unprecedented reactivity and intermediates,” Journal of the American Chemical Society, vol. 127, no. 1, pp. 210–216, 2005. View at Publisher · View at Google Scholar · View at Scopus
  38. S. Minakata, S. Okumura, T. Nagamachi, and Y. Takeda, “Generation of nitrile oxides from oximes using t -BuOI and their cycloaddition,” Organic Letters, vol. 13, no. 11, pp. 2966–2969, 2011. View at Publisher · View at Google Scholar · View at Scopus
  39. R. D. Jadhav, H. D. Mistry, H. Motiwala et al., “A facile one-pot synthesis of 3,5-disubstituted isoxazole derivatives using hydroxy (tosyloxy) iodobenzene,” Journal of Heterocyclic Chemistry, vol. 50, no. 4, pp. 774–780, 2013. View at Publisher · View at Google Scholar · View at Scopus
  40. A. Dondoni, P. P. Giovannini, and A. Massi, “Assembling heterocycle-tethered C-glycosyl and α-amino acid residues via 1,3-dipolar cycloaddition reactions,” Organic Letters, vol. 6, no. 17, pp. 2929–2932, 2004. View at Publisher · View at Google Scholar · View at Scopus
  41. S. Bhosale, S. Kurhade, U. V. Prasad, V. P. Palle, and D. Bhuniya, “Efficient synthesis of isoxazoles and isoxazolines from aldoximes using Magtrieve CrO2,” Tetrahedron Letters, vol. 50, no. 27, pp. 3948–3951, 2009. View at Publisher · View at Google Scholar · View at Scopus
  42. R. Huisgen, “1,3-Dipolar cycloaddition. Past and future,” Angewandte Chemie International Edition, vol. 2, no. 10, pp. 565–568, 1963. View at Google Scholar
  43. S. Kanemasa and O. Tsuge, “Recent advances in synthetic applications of nitrile oxide cycloaddition (1981–1989),” Heterocycles, vol. 30, no. 1, pp. 719–736, 1990. View at Publisher · View at Google Scholar · View at Scopus
  44. L. M. Stanley and M. P. Sibi, “Enantioselective copper-catalyzed 1,3-dipolar cycloadditions,” Chemical Reviews, vol. 108, no. 8, pp. 2887–2902, 2008. View at Publisher · View at Google Scholar · View at Scopus
  45. R. Sustmann, “Rolf Huisgen's contribution to organic chemistry, emphasizing 1,3-dipolar cycloadditions,” Heterocycles, vol. 40, no. 1, pp. 1–18, 1995. View at Publisher · View at Google Scholar · View at Scopus
  46. M. J. O'Donnell, K. Wojciechowski, L. Ghosez, M. Navarro, F. Sainte, and J. P. Antoine, “Alkylation of protected a-amino acid derivatives in the presence of potassium carbonate,” Synthesis, no. 4, pp. 313–315, 1984. View at Google Scholar
  47. M. J. O'Donnell, W. D. Bennett, and S. Wu, “The stereoselective synthesis of α-amino acids by phase-transfer catalysis,” Journal of the American Chemical Society, vol. 111, no. 6, pp. 2353–2355, 1989. View at Publisher · View at Google Scholar · View at Scopus
  48. M. J. O'Donnell, F. Delgado, and R. S. Pottorf, “Enantioselective solid-phase synthesis of α-amino acid derivatives,” Tetrahedron, vol. 55, no. 20, pp. 6347–6362, 1999. View at Publisher · View at Google Scholar · View at Scopus
  49. B. M. Hoey, J. Butler, and A. J. Swallow, “Reductive activation of mitomycin C,” Biochemistry, vol. 27, no. 7, pp. 2608–2614, 1988. View at Publisher · View at Google Scholar · View at Scopus
  50. P. Nun, V. Pérez, M. Calmès, J. Martinez, and F. Lamaty, “Preparation of chiral amino esters by asymmetric phase-transfer catalyzed alkylations of schiff bases in a ball mill,” Chemistry—A European Journal, vol. 18, no. 12, pp. 3773–3779, 2012. View at Publisher · View at Google Scholar · View at Scopus