HETEROCYCLES

Paper | Regular issue | Vol. 85, No. 11, 2012, pp. 2693-2712
Received, 6th August, 2012, Accepted, 31st August, 2012, Published online, 12th September, 2012.
DOI: 10.3987/COM-12-12565

■ Design, Synthesis and Biological Activity Evaluation of 2,5-Diphenyl-1,3,4-oxadiazole Derivatives as Novel Inhibitors of Fructose-1,6-bisphosphatase

Hai-Bing He, Li-Xin Gao, Yue-Yang Zhou, Ting Liu, Jie Tang, Xue-Ping Gong, Wen-Wei Qiu, Jing-Ya Li,* Jia Li,* and Fan Yang*

Shanghai Engineering Research Center of Molecular Theraputics and New Drug Development, East China Normal University, North Zhongshan Road, Shanghai 200062, China

Abstract

Fructose-1,6-bisphosphatase (FBPase), an important gluconeogenic enzyme, catalyzes the hydrolysis of fructose 1,6-bisphosphate to fructose 6-phosphate. The effort to discover new FBPase inhibitors was carried out by high-throughput screening (HTS) of a library of 56,000 lead-like compounds, and a 2,5-diphenyl-1,3,4-oxadiazole (3a, IC50 = 15.45 μM) which bearing no phosphate group was identified as a potential FBPase inhibitor for the first time. Structure-activity-relationship (SAR) research of a series of analogues obtained by modifying the substituent groups and replacing the 1,3,4-oxadiazole with several other heterocycles disclosed the key structure and substituent groups related to the binding with FBPase.

INTRODUCTION
Type-2 diabetes mellitus (T2DM) is associated with an increased rate of hepatic glucose production that contributes to fasting hyperglycemia. Specifically, increased endogenous glucose production can be accounted for by increased rates of hepatic gluconeogenesis (GNG).1 The rate of gluconeogenesis is governed by the activity of three regulatory and irreversible enzymes, phosphoenolpyruvate carboxykinase (PEPCK), fructose-1,6-bisphosphatase (FBPase), and glucose-6-phosphatase (G6Pase).2 As a key enzyme of gluconeogenic pathways, FBPase catalyzes the hydrolysis of fructose 1,6-bisphosphate (F1,6-P2) to fructose-6-phosphate (F6-P) and inorganic phosphate,3 which is a rate-controlling reaction of hepatic and renal gluconeogenesis.4 Unlike both PEPCK and G6Pase, FBPase is not only a rate-controlling enzyme within the GNG pathway but also functions only within the GNG pathway.5 Therefore FBPase is an attractive target in the development of new anti-diabetic pharmaceuticals.

FBPase is generally considered to be regulated in vivo via the allosteric interactions of AMP.6 Therefore the AMP-site was recognized as a specific target of FBPase inhibitor research. Several clusters of small molecules were reported as AMP-site allosteric inhibitors of FBPase. Among them, two AMP mimics 1 and 2 were highlighted.7-9 However, there are still challenges for the AMP mimics due to the abundance of AMP-binding enzymes controlling other key biosynthetic pathways resulting in issues with specificity. Other difficulties that need to be overcome include the hydrophilic nature of AMP sites and their reliance on the negatively charged phosphate group of AMP for binding affinity.10 Therefore, in the past decade, a lot of effort was made to discover new FBPase inhibitors structurally distinct from the AMP analogues without phosphate groups.11-15

In order to search for new hits with FBPase inhibitory activity, we carried out a high-throughput screening (HTS) campaign of a library of 56,000 lead-like small molecular compounds. As a result, 3a was recognized as a lead compound with a moderate FBPase inhibitory activity (IC50 = 15.45 µM). The scaffold of 3a contains three typical elements: the 1,3,4-oxadiazole ring, amide bond and other three substituted phenyl groups. Because the amide bond usually plays key roles in binding to the target sites of enzymes and usually are responsible for good pharmacokinetic and toxic properties. So at the first stage of SAR, the amide bond was reserved. Additionally, phenyl B was reserved without any modification due to the complication caused by multi-substitution. The rest areas of the structure with high modifiability, phenyl A, B and the 1,3,4-oxadiazole ring were investigated systematically.

RESULTS AND DISCUSSION
1. Chemistry. All the final products (3, 8, 9 and 24) were prepared via the key nitro-containing intermediates 6 and 11. The synthesis of 1,3,4-oxadiazole 6 was followed the classical procedures: dehydrated cyclization of dihydrazide 5 with POCl3 in acetonitrile (Scheme 1).16 Condensation of compounds 7 with the corresponding carboxylic acid using EDC as the condensation reagent gave 3, 8 or 9. The dihydroxy compound 10 was prepared by demethylation of 3i with boron tribromide.17

The reaction of 6i with Lawesson’s reagent afforded 1,3,4-thiadiazole 11a.18 11b was produced from 6i with benzylamine at a high temperature (Scheme 2).19

The synthesis of heterocyclic rings that containing two heteroatoms (11c~11g) was shown in Scheme 3. Bromination of 4'-methylacetophenone afforded the key initial material 12.20 Compound 12 reacted with thioamide 14 to produce 2,4-diphenylthiazole 11c in one step.21 2,5-diphenylthiazole 11g and 2,5-diphenyloxazole 11f were obtained by the reaction of 18 with Lawesson’s reagent or phosphorus oxychloride, respectively.22,23 Compound 18 was synthesized from 12 via the azide substitution, hydrogenation and acylation successively.24 Cyclization of 15 in acetic acid with the presence of ammonium acetate yielded a mixture of 11d (18%) and 11e (27%).25

The analogue 3,5-diphenyl-1,2,4-oxadiazole (11h and 11i) was synthesized by a common procedure as shown in Scheme 4. A nucleophilic addition of hydroxylamine to the cyano afforded 20 in a high yield. Condensation of 20 with benzoyl chloride in 2-picoline produced the precipitate 22. The dehydrative cyclization of 22 was easily accomplished in refluxing toluene to give 11h and 11i.26,27

In order to avoid the possible hydrogenation of the heterocyclic rings, the reduction of NO2 group (6 and 11) to NH2 group (7 and 23) was carried out in the presence of zinc and ammonium chloride.28 Similar to the preparation of 3, 24 was obtained via the condensation of compounds 23 with (3,4-dimethoxyphenyl)acetic acid using EDC as the condensation reagent. Debenzylation of 24b in hydrogen with Pd/C as the catalyst gave 25 (Scheme 5).

2. In vitro biological evaluation. All the compounds were evaluated in the enzyme inhibition assay against FBPase by a method reported by Doris Rittmann.29 Initially, in order to enhance the activity against FBPase, we investigated the substituent groups on phenyl A and B. Due to the conjugated system constituted by three aromatic rings, the inductive effect of the substituent groups on ring A had significant effect on activity (Table 1). Removing of the methyl group at C-4 (3b) resulted in the elimination of the activity. However, the C-3 position of ring A showed some tolerance of electron-withdrawing groups. 3-Br substituted 3c showed an enhanced threefold (IC50 = 4.92 µM) activity than the lead compound 3a (IC50 = 15.45 µM). Methoxycarbonyl substituted 3f also kept a similar activity to 3a. However, the introduction of more electron-withdrawing group cyano (3d) and trifluoromethyl (3e) led to the elimination of the activity. Moreover, we still investigated effects of the substituent groups at C-4 and C-2 position. Similar to 3b, the electron-withdrawing 4-Cl (3g) or 4-F (3h) substituted compound didn’t show significant inhibitory activity to the enzyme. In contrast, by introducing electron-donating methyl at C-4 position, we obtained 3i (IC50 = 4.51 µM) with an improved threefold activity than 3a. A bigger and more hydrophobic isopropyl (3j, IC50 = 7.24 µM) at C-4 showed a decreased activity. In addition, the combination of the optimized group at C-3 and C-4 position (3k, 3l) didn’t display an encouraging result. Modification of C-2 on ring A gave two compounds (3m, 3n), which showed no significant inhibition activity against FBPase. When it comes to the substituents on phenyl B, the comparison of 3i with 8 and 9 showed the importance of 3’,4’-dimethoxy unit for the inhibitory activity. Remarkably, when the 3’,4’-dimethoxy group was replaced by 3’,4’-dihydroxy group (10), the activity reduced by half (IC50 = 9.21 µM). The result revealed that a hydrogen-bonding acceptor on ring B is essential for the binding with FBPase instead of a hydrogen-bonding donor. The result convinced the importance of 4-methyl group on phenyl A ring and 3,4-dimethoxy substituent on phenyl B ring, which were reserved in the following SAR research.

On the reservation of 4-methyl at phenyl A and 3,4-dimethoxy at phenyl B, we tried to replace the 1,3,4-oxadiazole unit with typical bioisosteres: a class of five-membered heterocycles (24a~24i, 25). The IC50 value of these compounds in Table 2 demonstrated that none of them could provide a similar affinity with the enzyme as 1,3,4-oxadiazole (Table 1). Whether the oxygen atom was replaced with sulfur (24a) or nitrogen (24b, 25) the activity diminished dramatically. Transposition of the nitrogen atom on oxadiazole ring also caused the lost of activity (24h, 24i). Furthermore, all the heterocyclic rings that containing two heteroatoms (24c~24g) didn’t show activities against FBPase. The results demonstrated the key role of 1,3,4-oxadiazole moiety for the inhibition against the enzyme.

CONCLUSION
In this paper we reported a series of compounds with a novel scaffold as inhibitors against fructose 1,6-bisphosphatase. The substituent groups on phenyl A and phenyl B and a class of heterocyclic bioisosteres were synthesized and evaluated systematically. Some of these compounds showed moderate to good inhibitory activity against FBPase. The two most potent compounds 3i (IC50 = 4.51 µM) and 3c (IC50 = 4.92 µM) showed improved threefold FBPase inhibitory activity than the lead compound 3a. The SAR research demonstrated the characteristics of the substituents at different positions essential to the activities and the key role of the 1,3,4-oxadiazole ring. These novel 2,5-diphenyl-1,3,4-oxadiazole derivatives could be promising lead compounds for the development of a new class of FBPase inhibitors. Further SAR research and evaluation of new derivatives are still in progress.

EXPERIMENTAL
General. 1H (400 and 500 MHz) and 13C (100 and 125 MHz) NMR spectra were recorded on a JEOL-400 or Bruker AM-500 Fourier transform spectrometer. The chemical shifts were reported (δ in ppm) using the δ = 7.26, 2.5 signals of CDCl3, DMSO-d6 (1H NMR), and using the δ = 77.23, 39.51 signals of CDCl3, DMSO-d6 (13C NMR) as internal standards. High-resolution mass data were obtained on a Micromass Tof II spectrometer.
General procedure A, synthesis of dihydrazide 5. 3-Nitrobenzohydrazide (4, 10 mmol), substituted benzoic acid (10 mmol), EDC (13 mmol) and HOBt (13 mmol) were suspended in anhydrous DCM (25 mL). To this mixture, 3-picoline (14 mmol) was added. After stirring at room temperature for 12 h, the reaction mixture was filtered. The filter cake was washed with 5% HCl and water successively, dried in oven and used without any purification.
General procedure B, synthesis of 6. Dihydrazide 5 (10 mmol) was suspended in anhydrous MeCN (20 mL). To this mixture POCl3 (2 mL) was added. After stirring at reflux for 8 h, the reaction mixture was evaporated in vacuo. The resultant residue was then purified by column chromatography (DCM / hexane = 1:1).
General procedure C, synthesis of 7 or 23. Nitro compounds 6 or 11 (10 mmol), zinc dust (50 mmol), ammonium chloride (100 mmol) were suspended in a mixed solution of MeOH (15 mL), THF (15 mL) and water (15 mL). The mixture was heated to reflux for 4 h. After filtration, the filtration was concentrated in vacuo and purified by column chromatography (DCM / EtOAc= 5:1).
General procedure D, synthesis of 3 or 24. Amino compounds 7 or 23 (2 mmol), substituted phenylacetic acid (2 mmol), EDC (2.5 mmol) and HOBt (2.5 mmol) were dissolved in anhydrous DCM (10 mL). To this solution 3-picoline (3 mmol) was added. After stirring at room temperature for 20 h, the reaction mixture was diluted with DCM (10 mL), and then washed with 5% HCl, water, 5% NaOH and saturated aqueous NaHCO3 successively. After dried over anhydrous Na2SO4, the organic phase was evaporated in vacuo and then purified by column chromatography (DCM / EtOAc / MeOH = 25: 5: 1).
N-(3-(5-(3-Chlorophenyl)-1,3,4-oxadiazol-2-yl)phenyl)-2-(3,4-dimethoxyphenyl)acetamide (3a). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, DMSO-d6): δ 3.63 (s, 2H), 3.75 (s, 3H), 3.78 (s, 3H), 6.89-6.94 (m, 2H), 6.99 (s, 1H), 7.57 (t, J = 7.8 Hz, 1H), 7.68 (t, J = 7.8 Hz, 1H), 7.74 (d, J = 7.5 Hz, 1H), 7.84~7.88 (m, 2H), 8.07 (d, J = 7.5 Hz, 1H), 8.12 (s, 1H), 8.46 (s, 1H), 10.44 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.8, 164.3, 163.9, 148.6, 147.7, 140.1, 134.1, 131.8, 131.5, 130.0, 128.0, 126.1, 125.3, 123.5, 122.4, 121.5, 121.1, 116.8, 113.2, 113.1, 111.9, 55.5, 55.4, 42.9; HRMS (ESI): Calcd for C24H21ClN3O4 [M+H]+, 450.1215; Found, 450.1218.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-phenyl-1,3,4-oxadiazol-2-yl)phenyl)acetamide (3b). Synthesized according to the General procedure D. White solid; mp 152-154 oC; 1H NMR (500 MHz, DMSO-d6): δ 3.62 (s, 2H), 3.73 (s, 3H), 3.76 (s, 3H), 6.87~6.92 (m, 2H), 6.98 (s, 1H), 7.55 (t, J = 8.0 Hz, 1H), 7.63-7.64 (m, 3H), 7.79 (d, J = 7.6 Hz, 1H), 7.83 (d, J = 7.6 Hz, 1H), 8.09 (d, J = 6.2 Hz, 2H), 8.45 (s, 1H), 10.38 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.8, 164.0, 163.9, 148.6, 147.7, 140.1, 132.0, 129.9, 129.4 (2C), 128.0, 126.6 (2C), 123.7, 123.3, 122.3, 121.3, 121.1, 116.7, 113.1, 111.9, 55.5, 55.4, 42.9; HRMS (ESI): Calcd for C24H22N3O4 [M+H]+, 416.1610; Found, 416.1642.
N-(3-(5-(3-Bromophenyl)-1,3,4-oxadiazol-2-yl)phenyl)-2-(3,4-dimethoxyphenyl)acetamide (3c). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, CDCl3): δ 3.73 (s, 2H), 3.86 (s, 3H), 3.87 (s, 3H), 6.85-6.89 (m, 3H), 7.36 (t, J = 7.9 Hz, 1H), 7.42 (t, J = 8.0 Hz, 1H), 7.64 (d, J = 8.0 Hz, 1H), 7.78 (d, J = 8.2 Hz, 1H), 7.81 (d, J = 7.8 Hz, 1H), 7.90 (s, 1H), 8.01 (d, J = 7.9 Hz, 1H), 8.18 (s, 1H), 8.21 (s, 1H); 13C NMR (125 MHz, CDCl3): δ 170.0, 164.5, 163.3, 149.4, 148.5, 138.7, 134.7, 130.6, 129.8, 129.6, 129.0, 126.5, 125.4, 124.0, 123.3, 123.1, 122.7, 121.7, 117.9, 112.5, 111.6, 55.9 (2C), 44.2; HRMS (ESI): Calcd for C24H21BrN3O4 [M+H]+, 494.0715; Found, 494.0718.
N-(3-(5-(3-Cyanophenyl)-1,3,4-oxadiazol-2-yl)phenyl)-2-(3,4-dimethoxyphenyl)acetamide (3d). Synthesized according to the General procedure D. White solid; mp 186-187 oC; 1H NMR (500 MHz, DMSO-d6): δ 3.61 (s, 2H), 3.73 (s, 3H), 3.76 (s, 3H), 6.87-6.92 (m, 2H), 6.97 (s, 1H), 7.56 (t, J = 8.0 Hz, 1H), 7.82-7.85 (m, 3H), 8.10 (d, J = 7.5 Hz, 1H), 8.39 (d, J = 8.0 Hz, 1H), 8.45 (s, 1H), 8.52 (s, 1H), 10.41 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.8, 164.4, 162.6, 148.6, 147.7, 140.1, 135.3, 131.0, 130.7, 130.2, 130.0, 128.0, 124.6, 123.5, 122.5, 121.6, 121.1, 117.8, 116.9, 113.1, 112.7, 111.9, 55.5, (2C), 42.9; HRMS (ESI): Calcd for C25H21N4O4 [M+H]+, 441.1557; Found, 441.1532.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-(3-(trifluoromethyl)phenyl)-1,3,4-oxadiazol-2-yl)phenyl)acetamide(3e). Synthesized according to the General procedure D. White solid; mp 177-179 oC; 1H NMR (500 MHz, CDCl3): δ 3.74 (s, 2H), 3.88 (s, 3H), 3.89 (s, 3H), 6.87-6.89 (m, 3H), 7.45 (t, J = 8.0 Hz, 1H), 7.66 (t, J = 7.8 Hz, 1H), 7.74 (s, 1H), 7.80 (d, J = 7.8 Hz, 2H), 7.84 (d, J = 7.7 Hz, 1H), 8.18 (s, 1H), 8.29 (d, J = 7.7 Hz, 1H), 8.5 (s, 1H); 13C NMR (125 MHz, CDCl3): δ 169.9, 164.7, 163.5, 149.4, 148.6, 138.7, 131.7 (q), 130.0, 129.9, 129.8, 128.3, 128.2, 126.4, 124.5, 124.0, 123.7 (q), 123.3, 122.7, 121.7, 118.0, 112.5, 111.7, 55.9 (2C), 44.3; HRMS (ESI): Calcd for C25H21F3N3O4 [M+H]+, 484.1479; Found, 484.1449.
Methyl 3-(5-(3-(2-(3,4-dimethoxyphenyl)acetamido)phenyl)-1,3,4-oxadiazol-2-yl)benzoate (3f). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, DMSO-d6): δ 3.61 (s, 2H), 3.73 (s, 3H), 3.76 (s, 3H), 3.87 (s, 3H), 6.87-6.92 (m, 2H), 6.98 (s, 1H), 7.56 (t, J = 8.0 Hz, 1H), 7.77 (t, J = 7.8 Hz, 1H), 7.82 (d, J = 7.6 Hz, 1H), 7.87 (d, J = 8.4 Hz, 1H), 8.19 (d, J = 7.6 Hz, 1H), 8.33 (d, J = 7.8 Hz, 1H), 8.41 (s, 1H), 8.59 (s, 1H), 10.43 (s, 1H); HRMS (ESI): Calcd for C26H23N3NaO6 [M+Na]+, 496.1485; Found, 496.1541.
N-(3-(5-(4-Chlorophenyl)-1,3,4-oxadiazol-2-yl)phenyl)-2-(3,4-dimethoxyphenyl)acetamide (3g). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, DMSO-d6): δ 3.62 (s, 2H), 3.75 (s, 3H), 3.78 (s, 3H), 6.88-6.92 (m, 2H), 6.99 (s, 1H), 7.57 (t, J = 7.9 Hz, 1H), 7.73 (d, J = 8.4 Hz, 2H), 7.81-7.85 (m, 2H), 8.12 (d, J = 8.4 Hz, 2H), 8.48 (s, 1H), 10.44 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.8, 164.1, 163.2, 148.5, 147.7, 140.1, 136.7, 130.0, 129.6 (2c), 128.4 (2c), 128.0, 123.6, 122.3, 122.2, 121.4, 121.1, 116.7, 113.1, 111.9, 55.5 (2c), 42.9; HRMS (ESI): Calcd for C24H20ClN3O4 [M+H]+, 450.1215; Found, 450.1257.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-(4-fluorophenyl)-1,3,4-oxadiazol-2-yl)phenyl)acetamide (3h). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, CDCl3): δ 3.73 (s, 2H), 3.87 (s, 3H), 3.89 (s, 3H), 6.85-6.88 (m, 3H), 7.19 (t, J = 8.5 Hz, 2H), 7.43 (t, J = 8.0 Hz, 1H), 7.67 (s, 1H), 7.74 (d, J = 8.1 Hz, 1H), 7.82 (d, J = 7.7 Hz, 1H), 8.08-8.11 (m, 2H), 8.17 (s, 1H); 13C NMR (125 MHz, CDCl3): δ 169.8, 165.1, 164.0, 163.2, 163.1, 148.5, 147.7, 140.1, 129.9, 129.3, 128.0, 123.6, 122.3, 121.3, 121.1, 120.0, 116.8, 116.7, 116.6, 113.1, 111.8, 55.5, 55.4, 42.9; HRMS (ESI): Calcd for C24H21FN3O4 [M+H]+, 434.1516; Found, 434.1537.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-p-tolyl-1,3,4-oxadiazol-2-yl)phenyl)acetamide (3i). Synthesized according to the General procedure D. White solid; mp 169-170 oC; 1H NMR (500 MHz, DMSO-d6): δ 2.41 (s, 3H), 3.62 (s, 2H), 3.74 (s, 3H), 3.77 (s, 3H), 6.88~6.93 (m, 2H), 6.93 (s, 1H), 7.44 (d, J = 7.6 Hz, 2H), 7.56 (t, J = 8.0 Hz, 1H), 7.79 (d, J = 7.5 Hz, 1H), 7.83 (d, J = 7.8 Hz, 1H), 7.98 (d, J = 7.5 Hz, 2H), 8.44 (s, 1H), 10.43 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.8, 164.1, 163.7, 148.6, 147.6, 142.2, 140.1, 130.0 (2C), 128.0, 126.6 (2C), 123.8, 122.2, 121.3, 121.1 (2C), 120.6, 116.7, 113.1, 111.9, 55.5, 55.5, 42.9, 21.1; HRMS (ESI): Calcd for C25H24N3O4 [M+H]+, 430.1761; Found, 430.1756.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-(4-isopropylphenyl)-1,3,4-oxadiazol-2-yl)phenyl)acetamide (3j). Synthesized according to the General procedure D. White solid; mp 138-140 oC; 1H NMR (500 MHz, CDCl3): δ 1.28 (d, J = 6.9 Hz, 6H), 2.93-3.02 (m, 1H), 3.72 (s, 2H), 3.87 (s, 3H), 3.88 (s, 3H), 6.81-6.89 (m, 3H), 7.35 (d, J = 8.3 Hz, 2H), 7.43 (t, J = 7.3 Hz, 1H), 7.81 (d, J = 8.0 Hz, 2H), 7.89 (s, 1H), 8.01 (d, J = 8.3 Hz, 1H), 8.16 (s, 1H); 13C NMR (125 MHz, CDCl3): δ 170.0, 164.8, 164.0, 153.2, 149.3, 148.5, 138.6, 129.7, 127.1 (2C), 127.0 (2C), 126.5, 124.3, 122.5, 121.6, 121.5, 121.0, 117.8, 112.5, 111.2, 55.8, 55.7, 44.2, 34.2, 23.6 (2C); HRMS (ESI): Calcd for C27H28N3O4 [M+H]+, 458.2080; Found, 458.2082.
N-(3-(5-(3,5-Dichlorophenyl)-1,3,4-oxadiazol-2-yl)phenyl)-2-(3,4-dimethoxyphenyl)acetamide (3k). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, DMSO-d6): δ 3.59 (s, 2H), 3.72 (s, 3H), 3.75 (s, 3H), 6.86 (d, J = 8.1 Hz, 1H), 6.89 (d, J = 8.2 Hz, 1H), 6.96 (s, 1H), 7.55 (t, J = 8.0 Hz, 1H), 7.84~7.86 (m, 2H), 7.92 (s, 1H), 8.08 (s, 1H), 8.09 (s, 1H), 8.42 (s, 1H), 10.38 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.8, 164.5, 161.9, 148.5, 147.7, 140.1, 135.2 (2C), 131.3, 130.0, 128.0, 126.5, 125.1 (2C), 123.4, 122.6, 121.7, 121.1, 116.8, 113.1, 111.8, 55.5, 55.4, 42.9. HRMS (ESI): Calcd for C24H20Cl2N3O4 [M+H]+, 484.0831; Found, 484.0871.
N-(3-(5-(3-Chloro-4-methylphenyl)-1,3,4-oxadiazol-2-yl)phenyl)-2-(3,4-dimethoxyphenyl)acetamide (3l). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, CDCl3): δ 2.41 (s, 3H), 3.72 (s, 2H), 3.84 (s, 3H), 3.85 (s, 3H), 6.83-6.88 (m, 3H), 7.32 (d, J = 7.5 Hz, 1H), 7.40 (t, J = 8.0 Hz, 1H), 7.77-7.79 (m, 2H), 7.83 (dd, J1 = 1.5 Hz, J2 = 7.9 Hz, 1H), 8.01 (d, J = 1.4 Hz, 1H), 8.07 (s, 1H), 8.18 (s, 1H); 13C NMR (125 MHz, CDCl3): δ 170.0, 164.2, 163.6, 149.3, 148.5, 140.3, 138.7, 135.1, 131.5, 129.7, 127.2, 126.6, 124.9, 124.1, 123.2, 122.6 (2C), 121.6, 117.9, 112.5, 111.6, 55.8 (2C), 44.1, 20.2; HRMS (ESI): Calcd for C25H23ClN3O4 [M+H]+, 464.1377; Found, 464.1377.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-(2-methoxyphenyl)-1,3,4-oxadiazol-2-yl)phenyl)acetamide (3m). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, DMSO-d6): δ 3.59 (s, 2H), 3.71 (s, 3H), 3.74 (s, 3H), 3.92 (s, 3H), 6.86 (d, J = 8.1 Hz, 1H), 6.89 (d, J = 8.1 Hz, 1H), 6.96 (s, 1H), 7.13 (t, J = 7.5 Hz, 1H), 7.26 (d, J = 8.5 Hz, 1H), 7.53 (t, J = 7.9 Hz, 1H), 7.60 (t, J = 7.5 Hz, 1H), 7.72 (d, J = 7.8 Hz, 1H), 7.81 (d, J = 7.9 Hz, 1H), 7.91 (d, J = 6.8 Hz, 1H), 8.41 (s, 1H), 10.39 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.8, 163.8, 162.8, 157.5, 148.6, 147.7, 140.1, 133.6, 130.2, 130.0, 128.0, 123.8, 122.1, 121.2, 121.1, 120.8, 116.7, 113.1, 112.7, 112.2, 111.9, 56.1, 55.5 (2C), 42.9; HRMS (ESI): Calcd for C25H24N3O5 [M+H]+, 446.1716; Found, 446.1731.
N-(3-(5-(2-Chlorophenyl)-1,3,4-oxadiazol-2-yl)phenyl)-2-(3,4-dimethoxyphenyl)acetamide (3n). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, DMSO-d6): δ 3.63 (s, 2H), 3.75 (s, 3H), 3.78 (s, 3H), 6.89 (d, J = 8.2 Hz, 1H), 6.92 (d, J = 8.1 Hz, 1H), 6.99 (s, 1H), 7.56~7.63 (m, 2H), 7.69 (t, J = 7.5 Hz, 1H), 7.75~7.80 (m, 2H), 7.88 (d, J = 8.1 Hz, 1H), 8.12 (d, J = 7.7 Hz, 1H), 8.46 (s, 1H), 10.45 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.9, 164.3, 162.2, 148.5, 147.7, 140.2, 133.3, 131.8, 131.3, 131.2, 130.1, 128.0, 127.9, 123.5, 122.5, 122.4, 121.4, 121.1, 116.8, 113.1, 111.8, 55.5 (2C), 42.9; HRMS (ESI): Calcd for C24H21ClN3O4 [M+H]+, 450.1215; Found, 450.1212.
3-Nitrobenzohydrazide (4). Methyl 3-nitrobenzoate (18.1 g, 0.1 mol) was dissolved in dioxane (50 mL). To this solution, 85% hydrazine hydrate (23.5 g, 0.4 mol) was added. After stirring at reflux for 6 h, the reaction mixture was concentrated in vacuo to give 4 as a yellow solid without any purification.
N'-(4-Methylbenzoyl)-3-nitrobenzohydrazide (5i), synthesized according to the General procedure A. White solid; Yield: 67%. 1H NMR (400 MHz, DMSO-d6): δ 2.37 (s, 3H), 7.32 (d, J = 7.2 Hz, 2H), 7.84-7.86 (m, 3H), 8.36 (d, J = 7.6 Hz, 1H), 8.44 (d, J = 8.0 Hz, 1H), 8.75 (s, 1H), 10.56 (s, 1H), 10.88 (s, 1H); Calcd for C15H14N3O4 [M+H]+, 300.0979; Found, 300.0976.
2-(3-Nitrophenyl)-5-p-tolyl-1,3,4-oxadiazole (6i), synthesized according to the General procedure B. White solid; Yield: 86%. 1H NMR (400 MHz, CDCl3): δ 2.46 (s, 3H), 7.36 (d, J = 6.8 Hz, 2H), 7.76 (s, 1H), 8.04 (d, J = 7.3 Hz, 2H), 8.40 (d, J = 7.3 Hz, 1H), 8.50 (d, J = 7.1 Hz, 1H), 8.93 (s, 1H); 13C NMR (100 MHz, CDCl3): δ 165.5, 162.4, 148.7, 142.9, 132.4, 130.4, 129.9 (2C), 127.1 (2C), 125.9, 125.7, 121.6, 120.6, 21.7; HRMS (ESI): Calcd for C15H12N3O3 [M+H]+, 282.0873; Found, 282.0869.
3-(5-p-Tolyl-1,3,4-oxadiazol-2-yl)aniline (7i). Synthesized according to the General procedure C. White solid; Yield: 83%. 1H NMR (400 MHz, CDCl3): δ 2.36 (s, 3H), 3.82 (s, 2H), 6.77 (s, 1H), 7.19-7.42 (m, 5H), 7.93 (d, J = 7.6 Hz, 2H); HRMS (ESI): Calcd for C15H13N3NaO [M+Na]+, 274.0956; Found, 274.0999.
2-(4-Chlorophenyl)-N-(3-(5-p-tolyl-1,3,4-oxadiazol-2-yl)phenyl)acetamide (8). Synthesized according to the General procedure D. White solid; mp 211-213 oC; 1H NMR (500 MHz, DMSO-d6): δ 2.40 (s, 3H), 3.69 (s, 2H), 7.35~7.38 (m, 4H), 7.43 (d, J = 8.0 Hz, 2H), 7.54 (t, J = 8.0 Hz, 1H), 7.77~7.81 (m, 2H), 7.97 (d, J = 8.1 Hz, 2H), 8.41 (s, 1H), 10.47 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.2, 164.1, 163.7, 142.2, 140.0, 134.7, 131.4, 131.1 (2C), 130.0 (2C), 129.5, 128.2 (2C), 126.5 (2C), 123.8, 122.2, 121.4, 120.6, 116.7, 42.4, 21.1; HRMS (ESI): Calcd for C23H19ClN3O2 [M+H]+, 404.1166; Found, 404.1277.
2-p-Tolyl-N-(3-(5-p-tolyl-1,3,4-oxadiazol-2-yl)phenyl)acetamide (9). Synthesized according to the General procedure D. White solid; mp 195-197 oC; 1H NMR (500 MHz, DMSO-d6): δ 2.27 (s, 3H), 2.47 (s, 3H), 3.62 (s, 2H), 7.13 (d, J = 7.7 Hz, 2H), 7.23 (d, J = 7.7 Hz, 2H), 7.53 (t, J = 7.9 Hz, 1H), 7.78 (d, J = 7.7 Hz, 2H), 7.81 (d, J = 7.7 Hz, 2H), 7.97 (d, J = 7.9 Hz, 2H), 8.42 (s, 1H), 10.42 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 170.2, 164.5, 164.2, 142.7, 141.5, 140.5, 136.1, 133.1, 130.4 (2C), 129.4 (2C), 129.3 (2C), 127.0 (2C), 124.2, 122.6, 121.7, 121.0, 117.1, 43.4, 21.6, 21.1; HRMS (ESI): Calcd for C24H22N3O2 [M+H]+, 384.1712; Found, 384.1751.
2-(3,4-Dihydroxyphenyl)-N-(3-(5-p-tolyl-1,3,4-oxadiazol-2-yl)phenyl)acetamide (10). 3a (0.1 g, 0.23 mmol) was dissolved in DCM (10 mL) and cooled to -20 oC. To the solution was added the solution of BBr3 (1 mmol) in DCM (1 mL). After stirring at -20 oC for 8 h, the solution was washed by water and dry over anhydrous Na2SO4. The resultant solution was then evaporated in vacuo to give 10. White solid; Yield: 0.09 g (92%). 1H NMR (500 MHz, DMSO-d6): δ 3.61 (s, 2H), 3.73 (s, 3H), 3.76 (s, 3H), 6.88~6.92 (m, 2H), 6.97 (s, 1H), 7.05 (t, J = 7.5 Hz, 1H), 7.11 (d, J = 8.2 Hz, 1H), 7.48 (t, J = 7.9 Hz, 1H), 7.56 (t, J = 7.9 Hz, 1H), 7.77 (d, J = 7.6 Hz, 1H), 7.83 (d, J = 8.1 Hz, 1H), 7.88 (d, J = 7.1 Hz, 1H), 8.45 (s, 1H), 10.34 (s, 1H), 10.46 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.8, 163.5, 163.3, 156.3, 148.6, 147.7, 140.1, 133.4, 129.9, 128.7, 128.0, 123.6, 122.3, 121.1 (2C), 119.7, 117.1, 116.7, 113.2, 111.9, 109.5, 55.5 (2C), 42.9; HRMS (ESI): Calcd for C24H21N3O5 [M+Na]+, 454.1379; Found, 454.1399.
2-(3-Nitrophenyl)-5-p-tolyl-1,3,4-thiadiazole (11a). 6i (1.62 g, 5.42 mmol) and Lawesson’s reagent (2.42 g, 6 mmol) were suspended in anhydrous THF (20 mL). The mixture was heated to reflux for 6 h. The reaction mixture was cooled to rt and then filtrated. The cake was collected, washed with EtOH and ether and dried in oven to give 11a. Yellow solid; Yield: 0.88g (55%). 1H NMR (400 MHz, CDCl3): δ 2.43 (s, 3H), 7.31 (d, J = 8.3 Hz, 2H), 7.70 (t, J = 8.1 Hz, 1H), 7.90 (d, J = 8.3 Hz, 1H), 8.34 (d, J = 8.1 Hz, 1H), 8.38 (d, J = 8.1 Hz, 2H), 8.79 (s, 1H); HRMS (ESI): Calcd for C15H12N3O2S [M+H]+; 298.0650; Found, 298.0677.
4-Benzyl-3-(3-nitrophenyl)-5-p-tolyl-4H-1,2,4-triazole (11b). To a mixture of BnNH2 (5 mL), TsOH (0.25 g) and mesitylene (25 mL), 6i (1.7 g, 6.0 mmol) was added. The mixture was heated to reflux for 13 h. After cooled to rt, the mixture was filtrated. The filter cake was collected and dissolved in DCM (30 mL). After removing the insoluble solid by filtration, the filtrate was concentrated and purified by column chromatography (DCM / EtOAc = 10:1) to give 11b. White solid; Yield: 1.03 g (46%). 1H NMR (500 MHz, CDCl3): δ 2.38 (s, 3H), 5.33 (s, 2H), 6.90 (d, J = 4.9 Hz, 2H), 7.24~7.30 (m, 5H), 7.52 (d, J = 6.4 Hz, 2H), 7.57 (s, 1H), 7.95 (s, 1H), 8.24 (d, J = 7.3 Hz, 1H), 8.38 (s, 1H); HRMS (ESI): Calcd for C22H19N4O2 [M+H]+, 371.1508; Found, 371.1535.
2-(3-Nitrophenyl)-4-p-tolylthiazole (11c). 14 (0.2 g, 1.1 mmol) and 12 (0.22 g, 1.1 mmol) were dissolved in EtOH (10 mL). The solution was then heated to reflux for 3 h. After cooling to rt, white precipitate emerged immediately. The precipitate was filtered and dried in oven to give 11c. White solid; Yield: 0.2 g (61%). HRMS (ESI): Calcd for C16H13N2O2S [M+H]+, 296.0698; Found, 296.0677.
2-(3-Nitrophenyl)-4-p-tolyloxazole (11d) and 2-(3-nitrophenyl)-4-p-tolyl-1H-imidazole (11e). The solution of 15 and ammonium acetate in acetic acid was refluxed for 6 h and then poured into ice water before a mass of light yellow precipitate appeared. The solid was collected by filtration and purified by column chromatography (hexane / EtOAc = 10 / 1) to give 11d (White solid; Yield: 0.1 g, 18%) and 11e (White solid; Yield: 0.15 g, 26.8%). 11d: HRMS (ESI): Calcd for C16H13N2O3 [M+H]+, 281.0926; Found 281.0885. 11e: HRMS (ESI): Calcd for C16H14N3O2 [M+H]+, 280.1086; Found: 280.1092.
2-(3-Nitrophenyl)-5-p-tolyloxazole (11f). 18 (0.3 g, 1 mmol) was refluxed in POCl3 (10 mL) for 3 h. After evaporating in vacuo the resultant residue was diluted with DCM (10 mL) and then poured into ice water. The DCM phase was separated and washed with saturated aqueous NaHCO3. After drying over anhydrous Na2SO4, the solution was evaporated in vacuo to give 11f. White solid; Yield: 0.28 g (99%). Calcd for C16H13N2O3 [M+H]+, 281.0926; Found: 281.0913.
2-(3-Nitrophenyl)-5-p-tolylthiazole (11g). The mixture of 18 (0.3 g, 1.0 mmol) and Lawesson’s reagent (0.8 g, 2.0 mmol) in dry THF (15 mL) was refluxed for 3 h. After being evaporated in vacuo the resultant residue was purified by column chromatography (hexane / EtOAc = 10/1) to give 11g. White solid; Yield: 0.2 g (66%). 1H NMR (400 MHz, CDCl3) δ 2.40 (s, 3H), 7.25 (d, J = 7.8 Hz, 2H), 7.51 (d, J = 6.3 Hz, 2H), 7.63-7.66 (m, 1H), 8.00 (s, 1H), 8.24-8.30 (m, 2H), 8.79 (s, 1H).
5-(3-Nitrophenyl)-3-p-tolyl-1,2,4-oxadiazole (11h). To a solution of 20h (0.8 g, 5.3 mmol) in 2-picoline (6 mL) was added 3-nitrobenzoyl chloride (6.4 mmol) at 0~5 oC. After stirring at rt overnight, a mass of precipitate emerged. The solid was collected by filtration and then refluxed in toluene for 3 h. The reaction mixture was then concentrated in vacuo and was washed with hexane. The resultant solid was dried in oven to give 11h. White solid; Yield: 0.65 g (43%). 1H NMR (500 MHz, CDCl3) δ 2.40 (s, 3H), 7.26-7.29 (m, 2H), 7.75 (t, J = 7.7Hz, 1H), 7.99 (d, J = 7.7 Hz, 2H), 8.41(d, J = 7.8 Hz, 1H), 8.48 (d, J = 7.8 Hz, 1H), 8.99 (s, 1H); 13C NMR (125 MHz, CDCl3) δ 173.2, 169.1, 148.4, 141.8, 133.4, 130.3, 129.5 (2C), 127.3 (2C), 126.8, 125.7, 123.3, 123.0, 122.9; HRMS (ESI): Calcd for C15H12N3O3 [M+H]+, 282.0879; Found, 282.0877.
3-(3-Nitrophenyl)-5-p-tolyl-1,2,4-oxadiazole (11i). White solid; 11i was synthesized from 3-nitrobenzonitrile according to the synthesis method of 11h.
2-Bromo-1-p-tolylethanone (12). To the solution of 4-methylacetophenone (2.68 g, 20 mmol) in MeOH was added two drops of hydrobromic acid. A solution of bromine (1 mL, 20 mmol) in MeOH (10 mL) was added dropwise. After stirring at rt for 3 h, the reaction mixture was concentrated to afford 9. Yellow solid; Yield: 3.7 g (92%). 1H NMR (CDCl3, 400 MHz): δ 7.83 (d, J = 8.0 Hz, 2H), 7.23 (d, J = 8.0 Hz, 2H), 4.37 (s, 2H), 2.37 (s, 3H).
3-Nitrobenzamide (13). To 30% ammonium hydroxide (60 mL) was added a solution of 3-nitrobenzoyl chloride (2.5 g, 15 mmol) in toluene (20 mL). White precipitate emerged immediately. The resultant precipitate was filtered and dried in oven to give 10. White solid; Yield: 1.5g (60%).
3-Nitrobenzothioamide (14). 13 (0.4 g, 2.4 mmol) and P2S5 (0.54 g, 2.4 mmol) were dissolved in THF (20 mL). The solution was then heated to reflux for 3 h. After cooling to rt the reaction mixture was concentrated in vacuum and then purified by column chromatography (hexane / DCM = 1 / 1) to give 14. Yellow solid; Yield: 0.2 g (46%). HRMS (ESI): Calcd for C7H7N2O2S [M+H]+, 183.0228; Found, 183.0200.
2-Oxo-2-p-tolylethyl 3-nitrobenzoate (15). 3-Nitrobenzoic acid (2.17 g, 13 mmol), 12 (2.5 g, 12.5 mmol) and DBU (1.9 g, 12.5 mmol) were dissolved in DMF. After heating at 60 oC for 3 h, the reaction mixture was poured into water (100 mL), extracted by Et2O, and dried over anhydrous Na2SO4. Purification by column chromatography (hexane / EtOAc = 10 / 1) afforded 15. White solid; Yield: 2.5 g (67%). 1H NMR (CDCl3, 400 MHz): δ 8.91 (s, 1H), 8.40 (d, J = 8.0 Hz, 2H), 7.96 (s, 1H), 7.85 (d, J = 8.0 Hz, 2H), 7.63 (s, J = 8.0 Hz, 1H), 7.25 (s, J = 8.0 Hz, 1H), 5.56 (s, 2H), 2.38 (s, 3H). HRMS (ESI): Calcd for C16H14NO5 [M+H]+, 300.0872; Found, 300.0833.
2-Azido-1-p-tolylethanone (16). To a solution of 12 (1 g, 5 mmol) in DMF (20 mL) was added NaN3 (1 g, 15 mmol). After stirring at rt for 4 h, water (100 mL) was added to reaction mixture, and the mixture was extracted by EtOAc, dried over anhydrous Na2SO4 and evaporated in vacuo to give a light yellow liquid 16. Yield: 0.85 g (97 %).
2-Amino-1-p-tolylethanone (17). 10% Pd/C (0.08 g, moisture: 50%) was suspended in a solution of 16 (0.85 g, 4.9 mmol) in a mixture of concentrated hydrogen chloride (2 mL) and MeOH (20 mL). After stirring at rt in hydrogen atmosphere overnight, the reaction mixture was filtered. The resultant filtration was evaporated in vacuo and recrystallized from EtOAc (20 mL) to give 17. White solid; Yield: 0.6 g (82%).
3-Nitro-N-(2-oxo-2-p-tolylethyl)benzamide (18). To a mixture of 17 (4 mmol) and triethylamine (0.9 mL, 6 mmol) in DCM (20 mL) was added dropwise the solution of 3-nitrobenzoyl chloride (4 mmol) in DCM (20 mL) at 0-5 oC. After stirring at 0-5 oC overnight, the reaction mixture was washed with 10% aqueous Na2CO3, 1N HCl and brine successfully and then dried over anhydrous Na2SO4. The resultant solution was evaporated in vacuo and purified by column chromatography (hexane / EtOAc = 5 / 1) to give 18. White solid; Yield: 0.8 g (67%). 1H NMR (400 MHz, CDCl3): δ 2.45 (s, 3H), 4.96 (d, J = 4.4 Hz, 2H), 7.34 (d, J = 8.0 Hz, 1H), 7.41 (brs, 1H), 7.69 (t, J = 8.0 Hz, 1H), 7.94 (d, J = 8.0 Hz, 2H), 8.22 (d, J = 8.4 Hz, 1H), 8.39 (d, J = 8.4 Hz ,1H), 8.74 (s, 1H).
N'-Hydroxy-4-methylbenzimidamide (20h). 4-Methylbenzonitrile (3.6 g, 30.7 mmol), K2CO3 (4.4 g, 31.9 mmol) and hydroxylamine hydrochloride (3 g, 43.1 mmol) were suspended in EtOH (50 mL). The mixture was then heated to reflux for 8 h. After cooling to rt, the mixture was filtered and then evaporated in vacuo and purified by column chromatography (DCM / EtOAc = 5/1) to give 20h. White solid; Yield: 3.69 g (80%). 1H NMR (500 MHz, CDCl3): δ 2.29 (s, 3H), 5.78 (s, 2H), 7.16 (d, J = 8.0 Hz, 2H), 7.57(d, J = 8.0 Hz, 2H), 9.61 (brs, 1H); 13C NMR (125 MHz, DMSO-d6): 150.9, 138.3, 130.6, 129.4, 129.2, 128.7, 125.4, 20.9. HRMS (ESI): Calcd for C8H11N2O [M+H]+, 151.0866; Found, 151.0813.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-p-tolyl-1,3,4-thiadiazol-2-yl)phenyl)acetamide (24a). Synthesized according to the General procedure D. White solid; 1H NMR (400 MHz, CDCl3): δ 2.43(s, 3H), 3.73 (s, 2H), 3.90 (s, 3H), 3.92 (s, 3H), 6.85 (s, 1H), 6.89~6.91 (m, 2H), 7.29~7.32 (m, 3H), 7.42 (t, J = 8.0 Hz, 1H), 7.69 (d, J = 8.0 Hz, 1H), 7.76 (d, J = 8.0 Hz, 1H), 7.88 (d, J = 8.0 Hz, 2H), 7.99 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.8, 167.8, 167.3, 148.6, 147.7 141.6, 140.2, 131.7, 130.0 (2C), 129.9, 128.1, 127.6 (2C), 126.8, 122.4, 121.7, 121.1, 117.6, 113.1, 111.9, 55.5 (2C), 42.9, 21.0; HRMS (ESI): Calcd for C25H24N3O3S [M+H]+, 446.1533; Found, 446.1532.
N-(3-(4-Benzyl-5-p-tolyl-4H-1,2,4-triazol-3-yl)phenyl)-2-(3,4-dimethoxyphenyl)acetamide (24b). Synthesized according to the General procedure D. White solid; mp 82-84 oC; 1H NMR (500 MHz, DMSO-d6): δ 2.34 (s, 3H), 3.57 (s, 2H), 3.72 (s, 3H), 3.74 (s, 3H), 5.37 (s, 2H), 6.72 (d, J = 6.5 Hz, 2H), 6.83-6.95 (m, 3H), 7.16-7.18 (m, 3H), 7.24 (d, J = 7.5 Hz, 1H), 7.29 (d, J = 7.5 Hz, 2H), 7.39 (t, J = 7.5 Hz, 1H), 7.52 (d, J = 8.0 Hz, 2H), 7.65 (d, J = 7.5 Hz, 2H), 8.01 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.7, 155.3, 154.9, 148.6, 147.7, 139.8, 139.7, 136.1, 129.5 (2C), 129.4, 128.8 (2C), 128.6 (2C), 128.2, 127.8, 127.6, 125.7 (2C), 124.5, 122.9, 121.2, 120.6, 119. 6, 113.1, 111.9, 55.6, 55.5, 47.7, 42.9, 20.9; HRMS (ESI): Calcd for C32H31N4O3 [M+H]+, 519.2396; Found, 519.2413.
2-(3,4-Dimethoxyphenyl)-N-(3-(4-p-tolylthiazol-2-yl)phenyl)acetamide (24c). Synthesized according to the General procedure D. White solid; 1H NMR (CDCl3, 400 MHz): δ 7.99 (s, 1H), 7.84 (d, J = 8.0 Hz, 2H), 7.70 -7.66 (m , 3H), 7.39 -7.36 (m, 2H), 7.21 (d, J = 8.0 Hz, 2H), 6.90 (m, 2H), 6.84 (s, 1H), 3.91 (s, 3H), 3.90 (s, 3H), 3.72 (s, 2H), 2.39 (s, 3H). 13C NMR (100 MHz, CDCl3): δ 169.7, 167.1, 156.4, 149.5, 148.7, 138.4, 138.1, 134.4, 131.7, 129.6, 129.4 (2C), 126.7, 126.4 (2C), 122.6, 121.8, 121.4, 117.5, 112.6, 112.2, 111.8, 55.9 (2C), 44.4, 21.3. HRMS (ESI): Calcd for C26H25N2O3S [M+H]+, 445.1586; Found: 445.1555.
2-(3,4-Dimethoxyphenyl)-N-(3-(4-p-tolyloxazol-2-yl)phenyl)acetamide (24d). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, CDCl3) δ 2.38 (s, 3H), 3.73 (s, 2H), 3.89 (s, 3H), 3.90 (s, 3H), 6.84 (s, 1H), 6.92 (m, J = 7.5 Hz, 2H), 7.42 (m, J = 8.0 Hz, 1H), 7.38-7.42 (m, 2H), 7.68 (d, J = 7.9 Hz, 2H), 7.79-7.83 (m, 3H), 7.91 (s, 1H), 7.93 (s, 1H). 13C NMR (125 MHz, CDCl3): δ 169.5, 161.2, 149.6, 148.7, 142.1, 138.1, 138.0, 133.1, 129.5, 129.4, 128.1, 126.5, 125.5, 122.3, 121.8, 121.7, 117.4, 112.5, 111.8, 55.9 (2C), 44.4, 20.8. HRMS (ESI): Calcd for C26H25N2O4 [M+H]+, 429.1814; Found: 429.1817.
2-(3,4-Dimethoxyphenyl)-N-(3-(4-p-tolyl-1H-imidazol-2-yl)phenyl)acetamide (24e). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, CDCl3): δ 2.30 (s, 3H), 3.51 (s, 2H), 3.75 (s, 3H), 3.79 (s, 3H), 6.66-6.79 (m, 3H), 7.05-7.11 (m, 4H), 7.36 (d, J = 7.8 Hz, 1H), 7.47-7.50 (m, 3H), 7.87 (s, 1H ), 8.19 (s, 2H). 13C NMR (125 MHz, CDCl3): δ 170.4, 161.2, 149.2, 148.4, 146.6, 137.9, 136.7 (2C), 130.9, 129.4, 129.3(2C), 126.6 (2C), 124.7, 121.7, 121.6, 120.1, 117.1, 112.4, 111.5, 55.8 (2C) , 43.9, 21.2. HRMS (ESI): Calcd for C26H26N3O3 [M+H]+, 428.1974; Found: 428.1979.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-p-tolyloxazol-2-yl)phenyl)acetamide (24f). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, CDCl3) δ 2.38 (s, 3H), 3.72 (s, 2H), 3.89 (s, 3H), 3.90 (s, 3H), 6.85 (s, 1H), 6.89 (m, 2H), 7.22 (d, J = 7.9 Hz, 2H), 7.35 (s, 1H), 7.38-7.42 (m, 2H), 7.58 (d, J = 8.0 Hz, 2H), 7.76 (d, J = 7.9 Hz, 1H), 7.81 (d, J = 7.7 Hz, 1H), 7.96 (s, 1H). 13C NMR (125 MHz, CDCl3): δ 169.6, 160.2, 151.7, 149.5, 148.6, 138.6, 138.2, 129.6 (2C), 128.0 (2C), 126.4 (2C), 125.0, 124.2, 122.6, 122.1, 121.8, 121.6, 117.1, 112.4, 111.7, 55.9 (2C), 44.4, 21.4. HRMS (ESI): Calcd for C26H25N2O4 [M+H]+, 429.1736; Found: 429.1716.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-p-tolylthiazol-2-yl)phenyl)acetamide (24g). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, CDCl3): δ 2.38 (s, 3H), 3.71 (s, 2H), 3.89 (s, 3H), 3.90 (s, 3H), 6.84 (s, 1H), 6.89 (m, 2H), 7.21 (d, J = 7.8 Hz, 2H), 7.34 (d, J = 7.9 Hz, 1H), 7.38 (d, J = 7.9 Hz, 1H), 7.47 (d, J = 8.0 Hz, 2H), 7.63 (d, J = 7.6 Hz, 1H), 7.73 (d, J = 8.2 Hz, 1H), 7.87 (s, 1H ), 7.94 (s, 1H). HRMS (ESI): Calcd for C26H25N2O3S [M+H]+, 445.1586; Found: 445.1576.
2-(3,4-Dimethoxyphenyl)-N-(3-(3-p-tolyl-1,2,4-oxadiazol-5-yl)phenyl)acetamide (24h). Synthesized according to the General procedure D. White solid; 1H NMR (500 MHz, DMSO-d6): δ 2.40 (s, 3H), 3.62 (s, 2H), 3.74 (s, 3H), 3.77 (s, 3H), 6.88-6.92 (m, 3H), 6.99 (s, 1H), 7.40 (d, J = 6.5 Hz, 2H), 7.59 (t, J = 7.6 Hz, 1H), 7.84 (d, J = 6.5 Hz, 1H), 7.89 (d, J = 7.7 Hz, 1H), 7.98 (d, J = 6.5 Hz, 2H), 8.55 (s, 1H), 10.45 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 175.2, 170.0, 168.2, 148.6, 147.7, 141.6, 140.2, 130.1, 129.8 (2C), 128.0 (2C), 127.0 (2C), 123.8, 123.4, 122.4, 121.2, 117.9, 113.1, 111.8, 55.5 (2C), 43.0, 21.1; HRMS (ESI): Calcd for C25H24N3O4 [M+H]+, 430.1767; Found, 430.1761.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-p-tolyl-1,2,4-oxadiazol-3-yl)phenyl)acetamide (24i). Synthesized according to the General procedure D. White solid; mp 178-180 oC; 1H NMR (500 MHz, DMSO-d6): δ 2.44 (s, 3H), 3.60 (s, 2H), 3.73 (s, 3H), 3.76 (s, 3H), 6.87-6.92 (m, 2H), 6.97 (s, 1H), 7.47 (d, J = 8.0 Hz, 2H), 7.52 (t, J = 7.5 Hz, 1H), 7.76 (d, J = 7.5 Hz, 1H), 7.83 (d, J = 8.0 Hz, 1H), 8.07 (d, J = 8.0 Hz, 2H), 8.42 (s, 1H), 10.36 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 175.4, 169.7, 168.1, 148.6, 147.7, 143.7, 140.0, 130.0 (2C), 129.6, 128.1, 127.8 (2C), 126.6, 121.8, 121.7, 121.1, 120.6, 117.5, 113.1, 111.9, 55.5, 55.4, 42.9, 21.1; HRMS (ESI): Calcd for C25H24N3O4 [M+H]+, 430.1767; Found, 430.1761.
2-(3,4-Dimethoxyphenyl)-N-(3-(5-p-tolyl-4H-1,2,4-triazol-3-yl)phenyl)acetamide (25). The triazole 22b (0.12 g, 0.28 mmol) and 10% Pd/C (moisture: 50%, 20 mg) were suspended in MeOH (10 mL), the mixture was hydrogenated at 2.5 MPa for 24 h. The catalyst was removed by filtration and the filtrate was evaporated under reduced pressure to give 23. mp 208-211 oC; Yield: 98 mg (99%). 1H NMR (500 MHz, DMSO-d6): δ 2.36 (s, 3H), 3.65 (s, 2H), 3.76 (s, 3H), 3.79 (s, 3H), 6.85-6.92 (m, 4H), 6.99 (s, 1H), 7.33 (d, J = 8.0 Hz, 2H), 7.42 (t, J = 7.9 Hz, 1H), 7.74 (t, J = 7.8 Hz, 1H), 7.97 (d, J = 8.0 Hz, 2H), 8.36 (s, 1H), 10.37 (s, 1H); 13C NMR (125 MHz, DMSO-d6): δ 169.5, 148.5, 147.6, 139.7, 139.3, 129.4 (2C), 129.1, 128.9, 128.3, 125.9 (2C), 121.1, 121.0, 120.7, 119.9, 116.8, 113.1, 113.0, 111.9, 111.8, 55.5 (2C), 42.9, 20.9; HRMS (ESI): Calcd for C25H25N4O3 [M+H]+, 429.1927; Found, 429.1923.

Biological activity against fructose-1,6-diphosphatase (FBPase). FBPase recombinant protein was expressed and purified according to previous report.29 FBPase activity was measured in a coupled spectrophotometric assay containing 5 mM MgCl2, 66.7 mM KCl, 66.7 mM MOPS pH 7.5, 0.25 mM NADP+, yeast glucose-6-phosphate-dehydrogenase (0.4 U/mL), yeast phosphoglucoisomerase (0.7 U/mL) and 325 nmol/L FBPase. Fructose 6-phosphate formed by the reaction of FBPase was converted to glucose 6-phosphate and subsequently to 6-phosphogluconate by coupling to phosphoglucoisomerase and glucose-6-dehydrogenase and the concomitant formation of NADPH was detected at 340 nm.

ACKNOWLEDGEMENTS
This research was financially supported by National Nature Science Foundation of China (81125023), Shanghai Science and Technology Council (No 11DZ2292200 and 10142200800), National Science and Technology Major Projects for Major New Drugs Innovation and Development (2012ZX09304011 and 2012ZX09301-001-004), Chinese Academy of Sciences (KSCS2-EW-R-15). We also thank the Laboratory of Organic Functional Molecules, Sino-French Institute of ECNU for support.

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