Compounds (91), (93) (98), (99), (102), (105) (108) and (112) were synthesised in order to be investigated as potential substrates for myrosinase.
4.2.1 Synthesis of Phenyl-1-thio-p-D-olucopyranoside (PTG) (91)^03
HO
S
OH
(91)
1,2,3,4,6-Penta-0-aoetyl“P-D-glucopyranose (6.00 g, 15.4 mmol) was dissolved in dry dichloromethane (70 ml) and stirred under nitrogen. To the resulting solution was added thiophenol (2.04 g, 18.5 mmol) then boron trifluoride-diethyl ether complex (10.92 g, 76.9 mmol). The reaction mixture was stirred at room temperature under nitrogen for 6 hours until the reaction was complete (t.I.c., hexane/ethyl acetate (2:1)). The solution was then washed with saturated sodium carbonate solution (3 x 100 ml) and water (100 ml), dried (Na2S0 4) and concentrated under reduced pressure to give a white solid (6.81 g). This solid was dissolved in dry methanol (150 ml) under nitrogen and to the resulting solution was added 1.0 M sodium methoxide in dry methanol (2.0 ml) under nitrogen. The reaction mixture was stirred at room temperature under nitrogen for 5 days. The solution was then neutralised with glacial acetic acid and concentrated under reduced pressure to give a white solid (3.52 g). This product was
purified by silica chromatography using ethyl acetate/methanol/water (16:2:1) as the eluent to give pure phenyl-1-thio-p-D-glucopyranoside (91) as a white solid (3.04 g, 73%), m.p. 123-6 °G (lit.,1^6 128-130 °C); [a]o -52.5 ° (c 1.0 in HgO) (lit.,176 -71.6 ° (c 1.0 in H2O)); Vmax (nujol)/cm-i 3350
(OH) and 730 (aromatic); ôh (200 MHz; 2H2O) 3.22-3.53 (4H, m., H- 4,5,5a 6b), 3.64 (1H, d.d., J i,2 5.0 Hz, Jz,3 12.4 Hz, H-2), 3.83 (1H, d.d., J2.3
12.4 Hz, J3,4 1.9 Hz, H-3), 4.84 (1H, d., J i,2 5.0 Hz, H-1), 7.35 (3H, m., H- 3',4',5'), 7.51 (2H, m., H-2',6'); ôc (50.3 MHz; 2H2O) 63.60 (0-6), 72.16 (0- 4), 74.53 (0-2), 80.04 (0-5), 82.71 (0-3), 90.11 (0-1), 130.92 (0-4'), 132.16
(C-3',5*), 134.43 (C-2',6') and 134.83 (C-T); m/z (Cl) 290 {[M + NH4]+,
100%) and 180 (12, [M - PhSH + NH4M .
4.2.2 Synthesis of 2.3.4.6-Tetra-0-acetvi-1-bromo-a-D-alucopvranose (a-D-Acetobromoalucose1 (68)204 AcO""^ »— u " Br (68) AcO AcO
To a solution of 1,2,3,4,6-penta-O-acetyl-p-D-glucopyranose (0.50 g, 1.28 mmol) in dry dichloromethane (10 ml) was added acetic anhydride (1.0 ml)
followed by 30% w/v hydrogen bromide in acetic acid (1.0 ml, 2.70 mmol) under nitrogen. The reaction mixture was stirred at room temperature under nitrogen for 48 hours until the reaction was complete (t.I.c., ethyl acetate/light petroleum (3:2)). Toluene (40 ml) was then added and the resulting solution was concentrated under reduced pressure. A further
portion of toluene (20 ml) was added and the solution was again
concentrated under reduced pressure to remove excess acetic acid. The residue was dissolved in dichloromethane (50 ml) and the solution washed with 10% sodium hydrogen carbonate solution (2 x 50 ml) and water (50 ml). The organic layer was separated, dried (MgS0 4) and
concentrated under reduced pressure to give pure 2,3,4,6-tetra-O-acetyl-l- bromo-a-D-glucopyranose (68) as a pale gold syrup (0.45 g, 85.5%) which overnight turned to a white solid, m.p. 86-7 °C (lit.,190 88-9 °C); [a]o +195.9 ° (c 2.42 in CHCI3) (lit.,205 +197.84 ° (c 2.42 in CHCI3)); Vmax (nujol)/cm-l 1730 (CO); ÔH (200 MHz; G2HCI3) 1.91-2.21 (12H, 4s., 4 x CH3), 4.11 (1H, m., H-5), 4.30 (2H, m., H-O^,6^), 4.82 (1H, d.d., J i,2 3.1 Hz, Jz,3 9.6 Hz, H- 2), 5.16 (1H, t., J3,4 9.6 Hz, ^4,5 9.6 Hz, H-4), 5.55 (1H, t., ^2,3 9.6 Hz, ^3,4 9.6 Hz, H-3), 6.61 (1H, d., J i,2 3.1 Hz, H-1); ôc (50.3 MHz; C2HCI3) 21.01 & 21.11 (4 X QH3), 61.39 (0-6), 67.60 (C-4), 70.59 (C-2), 71.00 (C-5), 72.57 (C-3), 86.56 (C-1), 169.91, 170.23, 170.29 & 170.94 (4 x CO); m/z (Cl) 428 & 430 ([M+NH4]T 18%), 331 (8, [M - Br]+) and 213 (29, [M - Br - 20Ac]+).
4.2.3 Synthesis of 2.3.4.6-Tetra-O-acetyl-p-D-alucopyranosvl Isothio- uronium Bromide (9 4 ^77
AcO
NHg (94)'
2,3,4,6-Tetra-0-acetyl-1-bromo-a-D-glucopyranose (68) (6.75 g, 16.4
mmol) was dissolved in dry acetone (30 ml) and to the resulting solution was added thiourea (1.25 g, 16.4 mmol). Once dissolved, the reaction mixture was refluxed for 15 minutes then cooled to room temperature. A white precipitate formed, which was filtered off and washed with acetone to give a white solid (4.33 g). This product was recrystaliised from acetone to give pure 2,3,4,6-tetra-O-acetyi-p-D-giucopyranosyi Isothiouronium bromide (94) as a white solid (3.69 g, 46.1%), m.p. 196-7 °C (d.) (lit.,206 205 °C); (Found: 0, 37.25; H, 4.59; N, 5.72. Calc, for C i5H23BrN2 0 9S: 0, 36.97; H, 4.76; N, 5.75%); [a]o -20.2 ° (c 1.0 in MeOH) (iit.,207 .1 7 . 3 » (c 1.0 in MeOH)); Vmax (nujol)/cm-i 3310-3160 (NH), 1750 (CO) and 1655 (NH);
5h (200 MHz; 2H2O) 1.90-2.09 (12H, 4s., 4 x CH3), 4.12-4.25 (1H, m., H-5), 4.25-4.42 (2H, m., H-6a,6b), 5.12-5.50 (4H. m., H-1,2,3,4); Sc (50.3 MHz;
2H2O) 14.89, 18.08, 18.11 & 18.21 (4 x ÇH3), 55.52 (C-T), 59.92 (C-6), 65.65 (C-4), 67.24 (C-2), 71.43 (C-5), 73.93 (C-3), 79.18 (C-1), 170.46, 170.65, 170.99 & 171.65 (4 x CO); m/z (El) 331 ([W- CH4BrN2S]+, 11%),
169 (35, [CsHgO# and 127 (18, [CeH703]+).
4.2.4 Svnthesis of 2.3.4.6-Tetra-0-acetvl-1-thio-p-D-alucopvranose (95)177
AcO-^
AcO—
OAc
(95)
Potassium metabisulfite (1.68 g, 7.57 mmol) was dissolved in water (20 ml) and the resulting solution was heated to 75 °C. Dichloromethane (25 ml) was then added as the lower phase and 2,3,4,6-tetra-O-acetyl-p-D-gluco- pyranosyl isothiouronium bromide (94) (3.67 g, 7.57 mmol) was added
with stirring. The two-phase system was refluxed for 15 minutes with stirring and then allowed to cool to room temperature. The organic layer was separated and washed with water (3 x 25 ml), dried (MgS0 4) and
concentrated under reduced pressure to give a white solid (1.42 g). This
product was recrystaliised from methanol to give pure 2,3,4,6-tetra-O- acetyl-1 -thio-p-D-glucopyranose (95) as a white solid (1.26 g, 46%), m.p.
72-5 °C (lit.,177 75 oQ). (Found: C, 46.02; H, 5.41. Calc, for C14H20O9S: 0, 46.15; H, 5.53%); [a]o -9.8 ° (c 1.5 in EtOH) (lit.,208 -8.3 ° (c 1.5 in EtOH));
Vm ax (nujol)/cm-i 3460 (SH) and 1735 (00); ôh (200 MHz; C^HCIs) 1.85-
2.35 (12H, 4s., 4 x CH3), 3.69 (1H, m., H-5), 4.13 (1H, d.d., Js,6a 2.5 Hz,
Jea.eb 12.3 Hz, H-6®), 4.26 (1H, d.d., Js.eb 4.8 Hz, Jea^Qb 12.3 Hz, H-6b), 4.54 (1H, t., J i,2 9.6 Hz, H-1), 4.98 (1H, t., J i,2 9.6 Hz, J2.3 9.6 Hz, H-2),
5.10 (1H, t., 1/3,4 9.6 Hz, J4,5 9.6 Hz, H-4), 5.20 (1H, t., Ü2,3 9.6 Hz, u/3,4 9.6 Hz, H-3); ôc (50.3 MHz; C2HCI3) 21.03 (2 x ÇH3), 21.20 (2 x ÇH3), 62.43 (0-6), 68.53 (0-4), 73.97 (0-2), 76.75 (0-5), 77.48 (0-3), 79.14 (0-1), 169.82 (ÇO), 170.08 (ÇO), 170.56 (CO) and 171.11 (ÇO); m/z (01) 382 {[M + NH4]+, 100%), 331 (7, [M - SH]+) and 322 (22, [M - OAc]+).
4.2.5 Svnthesis of p-Nitrophenyl-1-thio-p-D-glucopyranoside (PNPTG^ m
HO HO
OH
(93)
2,3,4,6-Tetra-0-acetyl-1-thio-p-D-glucopyranose (95) (0.95 g, 2.6 mmol) was dissolved in dry dimethylformamide (5 ml) under nitrogen and to the resulting solution was added p-fluoronitrobenzene (0.37 g, 2.6 mmol) and triethylamine (0.29 g, 2.9 mmol) under nitrogen. The reaction mixture was stirred at room temperature under nitrogen for 24 hours. The mixture was then concentrated under reduced pressure to give a yellow solid. This solid was dissolved in dichloromethane (50 ml) and washed with water (3