13C-NMR (75 MHz, CDCl3) 175.66, 153.88, 132.74, 129.56, 115.21, 69.60, 52.56, 35.93, 29.95 ppm; HRMS (ESI) calculated for C11H14O4 [M + Na]+ 233.0892, found 233.0823. Intermediate S4, obtained from last step of reaction without further purification, was dissolved in acetonitrile (2 mL) at 0 C. B and leupeptin. Both nostosin A and B contain three subunits, 2-hydroxy-4-(4-hydroxyphenyl)butanoic acid (Hhpba), = ?2.4 (1.1, MeOH)) with natural product ([]= ?2.9 (0.08, H2O)), while nostosin B 1a shows the opposite sign ([]= 1.6 (1.0, MeOH)). From these analytical data, we believe that nostosin 1b was the real structure of the natural product, the slight differences on NMR spectra should be arisen from the different experimental conditions for data acquisition, i.e., the concentration of sample and pH of the solution. To further identify the stereochemistry of nostosin B, the synthetic samples (1a and 1b) were co-injected with the authentic sample (natural product sample) using high-performance liquid chromatography (Physique 3). Both reverse phase column and chiral column gave the same results, indicating that synthetic nostosin B 1b has identical retention time with natural nostosin B, unambiguously defining the stereochemistry of Hhpba as = ?28.7 (1.0, CH2Cl2); 1H-NMR (500 MHz, CDCl3) 7.72C7.70 (m, 2H), 7.53C7.51 (m, 2H), 7.37C7.32 (m, 2H), 7.25C7.22 (m, 2H), 6.45 (br, 1H), 6.13 (br, 2H), 5.63 (br, 1H), 4.41C4.37 (dd, = 7.5, 10.5 Hz, 1H), 4.26C4.22 (m, 1H), 4.14C4.10 (m, 1H), 4.03C3.90 (m, 1H), 3.52 (s, 2H), 3.18 (br, 1H), 3.10 (br, 1H), 2.89 (s, 2H), 2.54 (s, 3H), 2.48 (s, 3H), 2.04 (s, 3H), 1.88C1.78 (m, 2H), 1.57C1.44 (m, 4H), 1.41 (s, 6H), 1.16C1.12 (m, 1H), 0.90C0.88 (m, 6H), 0.85 (s, 9H), 0.01 (s, 6H). 13C-NMR (125 MHz, CDCl3) 171.79, 158.74, 156.68, 156.22, 143.94, 143.69, 141.38, 138.45, 132.40, 127.82, 127.21, 125.11, 124.58, 120.07, 117.46, 88.34, 67.26, 65.02, 60.03, 47.22, 43.35, 41.22, 37.50, 29.15, 28.65, 25.94, 25.75, 25.33, 25.10, 19.30, 18.34, 17.95, 15.54, 12.50, 11.46, ?5.42, ?5.44 ppm; HRMS (ESI) calculated for C17H20O3 [M + Na]+ 884.4530, found 884.4434. Compound 6 (55 mg, 0.064 mmol) was dissolved in CH3CN (2 mL) and cooled to 0 C, after diethylamine (0.07 mL, 0.64 mmol) was added, the reaction mixture was brought to room heat and monitored by TLC. Upon the consumption of all starting materials, the reaction mixture was concentrated in vacuo. The residue was dissolved in DCM (2 mL) and concentrated in vacuo, these procedures were repeated twice. The residue was dried under high vacuum for 1 h to give the crude amine 7, which was used directly without further purification. 4.3. Synthesis of the Hhpba Fragment 4.3.1. Synthesis of = 1.5 (1.1, CHCl3); 1H-NMR (500 MHz, CDCl3) 7.48C7.35 (m, 5H), 7.16 (d, = 8.0 Hz, 2H), 6.95 (d, = 8.0 Hz, 2H), 5.07 (s, 2H), 4.19C4.08 (m, 1H), 4.04 (dd, = 7.9, 5.8 Hz, 1H), 3.56 (t, = 7.5 Hz, 1H), 2.79C2.60 (m, 2H), 2.04C1.73 (m, 2H), 1.49 (s, 3H), 1.41 (s, 3H). 13C-NMR (125 MHz, CDCl3) 157.16, 137.21, 133.92, 129.34, 128.61, 127.95, 127.51, 114.84, 108.74, 75.41, 70.05, 69.40, 35.59, 31.19, 27.08, 25.83 ppm; HRMS (ESI) calculated for C20H24O3 [M + Na]+ 335.1725, found 335.1744. Compound 16 (0.50 g, 1.60 mmol) was dissolved in methanol (10 mL) and cooled to 0 C, after PTSA (30 mg, 0.16 mmol) was added, the reaction mixture was stirred at room temperature for 16 h. The reaction solution was concentrated in vacuo, the residue was dissolved in ethyl acetate (50 mL) and washed with saturated aqueous answer of sodium bicarbonate (50 mL) and brine (50 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated in vacuo. The residue was purified by flash chromatography to afford the desired diol compound S1 (0.43 g, 99%) as clear oil. []= ?14.7 (1.0, MeOH); 1H-NMR (500 MHz, CDCl3) Rabbit Polyclonal to PEG3 7.47C7.29 (m, 5H), 7.12 (d, = 8.5 Hz, 2H), 6.91 (d, = 8.6 Hz, 2H), 5.04 (s, 2H), 3.78C3.57 (m, 2H), 3.46 (dd, = 11.1, 7.6 Hz, 1H), 2.80C2.55 (m, 2H), 2.28 N6-(4-Hydroxybenzyl)adenosine (s, 1H), 2.08 (s, 1H), 1.72 (m, 2H). 13C NMR (125 MHz, CDCl3) 157.16, 137.20, 134.01, 129.33, 129.31, 128.57, 128.55, 127.89, 127.47, 127.44, 114.92, 114.88, 77.27, 77.01, 76.76, N6-(4-Hydroxybenzyl)adenosine 71.52,.13C-NMR (125 MHz, CDCl3) 173.28, 157.22, 137.59, 137.27, 133.44, 129.48, 128.57, 128.45, 128.15, 128.13, 127.90, 127.46, 114.91, 72.44, 70.12, 70.09, 51.84, 34.75, 30.52 ppm; HRMS (ESI) calculated for C25H26O4 [M + H]+ 391.1831, found 391.1889. Methyl ester S3 (60 mg, 0.154 mmol) was dissolved in ethyl acetate (2 mL), after Pd/C (6 mg, 10% Pd on charcoal) was added under a protective flow of nitrogen, the reaction vessel was sealed and purged by hydrogen. a separate windows Physique 1 Natural peptide aldehyde and structure of nostosins A and B and leupeptin. Both nostosin A and B contain three subunits, 2-hydroxy-4-(4-hydroxyphenyl)butanoic acid (Hhpba), = ?2.4 (1.1, MeOH)) with natural product ([]= ?2.9 (0.08, H2O)), while nostosin B 1a shows the opposite sign ([]= 1.6 (1.0, MeOH)). From these analytical data, we believe that nostosin 1b was the real structure of the natural product, the slight differences on NMR spectra should be arisen from the different experimental conditions for data acquisition, i.e., the concentration of sample and pH of the solution. To further identify the stereochemistry of nostosin B, the synthetic samples (1a and 1b) were co-injected with the authentic sample (natural product sample) using high-performance liquid chromatography (Physique 3). Both reverse phase column and chiral column gave the same results, indicating that synthetic nostosin B 1b has identical retention time with natural nostosin B, unambiguously defining the stereochemistry of Hhpba as = ?28.7 (1.0, CH2Cl2); 1H-NMR (500 MHz, CDCl3) 7.72C7.70 (m, 2H), 7.53C7.51 (m, 2H), 7.37C7.32 (m, 2H), 7.25C7.22 (m, 2H), 6.45 (br, 1H), 6.13 (br, 2H), 5.63 (br, 1H), 4.41C4.37 (dd, = 7.5, 10.5 Hz, 1H), 4.26C4.22 (m, 1H), 4.14C4.10 (m, 1H), 4.03C3.90 (m, 1H), 3.52 (s, 2H), 3.18 (br, 1H), 3.10 (br, 1H), 2.89 (s, 2H), 2.54 (s, 3H), 2.48 (s, 3H), 2.04 (s, 3H), 1.88C1.78 (m, 2H), 1.57C1.44 (m, 4H), 1.41 (s, 6H), 1.16C1.12 (m, 1H), 0.90C0.88 (m, 6H), 0.85 (s, 9H), 0.01 (s, 6H). 13C-NMR (125 MHz, CDCl3) 171.79, 158.74, 156.68, 156.22, 143.94, 143.69, 141.38, 138.45, 132.40, 127.82, 127.21, 125.11, 124.58, 120.07, 117.46, 88.34, 67.26, 65.02, 60.03, 47.22, 43.35, 41.22, 37.50, 29.15, 28.65, 25.94, 25.75, 25.33, 25.10, 19.30, 18.34, 17.95, 15.54, 12.50, 11.46, ?5.42, ?5.44 ppm; HRMS (ESI) calculated for C17H20O3 [M + Na]+ 884.4530, found 884.4434. Compound 6 (55 mg, 0.064 mmol) was dissolved in CH3CN (2 mL) and cooled N6-(4-Hydroxybenzyl)adenosine to 0 C, after diethylamine (0.07 mL, 0.64 mmol) was added, the reaction mixture was brought to room temperature and monitored by TLC. Upon the consumption of all starting materials, the reaction mixture was concentrated in vacuo. The residue was dissolved in DCM (2 mL) and concentrated in vacuo, these procedures were repeated twice. The residue was dried under high vacuum for 1 h to give the crude amine 7, which was used directly without further purification. 4.3. Synthesis of the Hhpba Fragment 4.3.1. Synthesis of = 1.5 (1.1, CHCl3); 1H-NMR (500 MHz, CDCl3) 7.48C7.35 (m, 5H), 7.16 (d, = 8.0 Hz, 2H), 6.95 (d, = 8.0 Hz, 2H), 5.07 (s, 2H), 4.19C4.08 (m, 1H), 4.04 (dd, = 7.9, 5.8 Hz, 1H), 3.56 (t, = 7.5 Hz, 1H), 2.79C2.60 (m, 2H), 2.04C1.73 (m, 2H), 1.49 (s, 3H), 1.41 (s, 3H). 13C-NMR (125 MHz, CDCl3) 157.16, 137.21, 133.92, 129.34, 128.61, 127.95, 127.51, 114.84, 108.74, 75.41, 70.05, 69.40, 35.59, 31.19, 27.08, 25.83 ppm; HRMS (ESI) calculated for C20H24O3 [M + Na]+ 335.1725, found 335.1744. Compound 16 (0.50 g, 1.60 mmol) was dissolved in methanol (10 mL) and cooled to 0 C, after PTSA (30 mg, 0.16 mmol) was added, the reaction mixture was stirred at room temperature for 16 h. The reaction solution was concentrated in vacuo, the residue was dissolved in ethyl acetate (50 mL) and washed with saturated aqueous solution of sodium bicarbonate (50 mL) and brine (50 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated in vacuo. The residue was purified by flash chromatography to afford the desired diol compound S1 (0.43 g, 99%) as clear oil. []= ?14.7 (1.0, MeOH); 1H-NMR (500 MHz, CDCl3) 7.47C7.29 (m, 5H), 7.12 (d, = 8.5 Hz, 2H), 6.91 (d, = 8.6 Hz, 2H), 5.04 (s, 2H), 3.78C3.57 (m, 2H), 3.46 (dd, = 11.1, 7.6 Hz, 1H), 2.80C2.55 (m, 2H), 2.28 (s, 1H), 2.08 (s, 1H), 1.72 (m, 2H). 13C NMR (125 MHz, CDCl3) 157.16, 137.20, 134.01, 129.33, 129.31, 128.57, 128.55, 127.89, 127.47, 127.44, 114.92, 114.88, 77.27, 77.01, 76.76, 71.52, 70.12, 66.82, 34.87, 30.90 ppm; HRMS (ESI) calculated for C17H20O3 [M + Na]+ 295.1412, found 295.1456. Diol compound S1 (136 mg, 0.50 mmol) was dissolved in DCM (5 mL) and cooled to 0 C, after triethyl amine (0.35 mL, 2.5 mmol) and triphenylmethyl.