Laccase-catalyzed synthesis of catechol thioethers by reaction of catechols with thiols using air as an oxidant Heba T. Abdel-Mohsen, Jürgen Conrad and Uwe Beifuss* Bioorganische Chemie, Institut für Chemie, Universität Hohenheim Garbenstr. 30, D-70599 tuttgart, Germany Fax: (+49) 711 459 22951; Tel: (+49) 711 459 22171 E-mail: ubeifuss@uni-hohenheim.de Electronic upplementary Information Table of contents 1. General remarks 2 2. General procedure for the laccase-catalyzed domino reaction between catechols and thiols 2 3. ynthesis and analytical data of catechol thioethers 3 4. Computational studies of compounds 3a and 3b 30 5. Calculation of the E-factor, atom economy, TO and TOF of the laccase-catalyzed domino reaction of catechol (1a) and 2-mercaptobenzoxazole (2a) 33. References 35 1
1. General remarks All chemicals were purchased from commercial suppliers. Laccase from Agaricus bisporus ( U / mg) was purchased from AA pezialenzyme. olvents used in extraction and purification were distilled prior to use. The ph of the buffer was adjusted using a ph 330/ET-1 ph-meter. Analytical thin layer chromatography (TLC) was performed on precoated silica gel 0 F 245 aluminium plates (Merck) with visualization under UV light. Flash chromatography was carried out on silica gel M 0, 0.04-0.053 mm (Macherey & agel). Melting points were determined on a Büchi melting point apparatus B-545 with open capillary tubes and are uncorrected. UV/VI spectra were recorded with a Varian Cary 50. IR spectra were measured on a Perkin-Elmer pectrum One (FT-IR-spectrometer). 1 H and 13 C MR spectra were recorded at 300 (75) MHz on a Varian Unity Inova using DMO-d. The chemical shifts were referenced to the solvent signals at δ H/C 2.49 / 39.50 ppm (DMO-d ) relative to TM as internal standards. ghqc, ghmbc and gcoy spectra were recorded on a Varian Unity Inova spectrometer (300 MHz). Coupling constants J [Hz] were directly taken from the spectra and are not averaged. Low resolution electron impact mass spectra (EI- LRM) and exact electron impact mass spectra (HRM) were recorded at 70 ev on a Finnigan MAT 95 instrument. Low resolution electron spray ionisation mass spectra (EI- LRM) and exact electron spray ionisation mass spectra (HRM) were recorded on a Bruker Daltonics (micro TOFQ) instrument. The intensities are reported as percentages relative to the base peak (I = 100%). 2. General procedure for the laccase-catalyzed domino reaction between catechols and thiols A 100 ml round bottomed flask with a magnetic stir bar was charged with a solution or suspension of the catechol 1 (0.3 mmol) and the thiol 2 (0.50 mmol) in methanol (3 ml). Phosphate buffer (0.2 M, ph, 27 ml) and laccase from A. bisporus (10 mg, U/ mg) were added and the mixture was stirred under air at rt for the time given. The reaction mixture was acidified with 2M HCl to ph ~ 4 and saturated with solid acl. The precipitated product was filtered with suction using a Buchner funnel. The filter cake was washed with aq acl (15%, 25 ml) and water. The crude products obtained after drying exhibit a purity of 90-95% (MR). Analytically pure products were obtained by column filtration (CH 2 Cl 2 /EtOAc 5:1 to 1:1) of the crude products. 2
3. ynthesis and analytical data of catechol thioethers 3.1. ynthesis and analytical data of 4-(benzo[d]oxazol-2 -ylthio)benzene-1,2-diol (3a) According to the general procedure, catechol (1a) (9 mg, 0.3 mmol), 2- mercaptobenzoxazole (2a) (7 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 1 h. Workup gave 4- (benzo[d]oxazol-2 -ylthio)benzene-1,2-diol (3a) as a pale brown solid (121 mg, 93%), mp 155 157 C (lit. 1 12 12 C); R f = 0.54 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 28 (log ε, 4.20), 280 (4.1), 24 (3.17) and 202 (4.2); ṽ max (atr)/cm -1 350 310 (OH), 1517, 1493, 1452, 1217 and 1137; δ H (300 MHz; DMO-d ).85 (1H, d, 3 J 5-H,-H 8.1 Hz, -H), 7.00 (1H, dd, 3 J 5-H,-H 8.4 Hz, 4 J 3-H,5-H 2.1 Hz, 5-H), 7.07 (1H, d, 4 J 3-H,5-H 2.4 Hz, 3-H), 7.28 7.31 (2H, m, 5 -H and -H), 7.57 7.2 (2H, m, 4 -H and 7 -H), 9.44 (1H, s, 1-OH or 2-OH), 9.59 (1H, s, 1-OH or 2-OH); δ C (75 MHz; DMO-d ) 110.2 (C-7 ), 113.8 (C-4), 11.58 (C-), 118.51 (C-4 ), 122.05 (C-3), 124.42 (C-5 or C- ), 124.3 (C-5 or C- ), 12.83 (C-5), 141.40 (C-3a ), 14.20 (C-2), 147.99 (C-1), 151.23 (C-7a ), 13.74 (C-2 ); m/z (EI, 70 ev) 259 (M +, 100%), 22 (M + H, 9), 151 (C 7 H 5 O +, 20), 119 (10) and 91 (12); HRM (EI, M + ) found: 259.0281; calcd for C 13 H 9 O 3 : 259.0303. 2.4842 2.4900 2.4959 3.351.8332.805.9852.9927 7.012 7.0200 7.024 7.098 7.2590 7.279 7.2812 7.2941 7.3071 7.3111 7.3291 7.572 7.5815 7.590 7.5981 7.053 7.147 7.200 9.4409 9.5941 2 3 4 5 1 3a 2 O 7a 3a 4 5 7 1.107 0.983 0.993 2.528 2.005 0.987 1.000 3
38.47 38.9432 39.2215 39.3753 39.5000 39.31 39.7784 39.9081 39.918 39.941 40.057 40.3350 110.201 113.850 11.578 118.5087 122.0503 124.4258 124.38 12.8353 141.4025 14.2003 147.9938 151.2257 13.7381 Fig. 1 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 3a in DMO-d 3.2. ynthesis and analytical data of 4-(benzo[d]thiazol-2 -ylthio)benzene-1,2-diol (3b) According to the general procedure, catechol (1a) (9 mg, 0.3 mmol), 2- mercaptobenzothiazole (2b) (84 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave 4- (benzo[d]thiazol-2 -ylthio)benzene-1,2-diol (3b) as a pale yellow solid (120 mg, 87%); mp 198 200 C; R f = 0.53 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 284 (log ε, 4.0) and 20 (4.20); ṽ max (atr)/cm -1 342 (OH), 3054 (C-H), 1593, 1415, 1270 and 1030; δ H (300 MHz; DMO-d ).90 (1H, d, 3 J 5-H,-H 8.1 Hz, -H), 7.07 (1H, dd, 3 J 5-H,-H 8.4 Hz, 4 J 3-H,5-H 2.1 Hz, 5- H), 7.13 (1H, d, 4 J 3-H,5-H 2.1 Hz, 3-H), 7.29 (1H, ddd, 3 J 5 -H, -H 7.2 or 7.9 Hz, 3 J -H,7 -H 7.2 or 7.9 Hz, 4 J 4 -H, -H 1.1 Hz, -H), 7.42 (1H, ddd, 3 J 4 -H,5 -H 7.3 or 8.2 Hz, 3 J 5 -H, -H 7.3 or 8.2 Hz, 4 J 5 -H,7 -H 1.3 Hz, 5 -H), 7.79 (1H, ddd, 3 J 4 -H,5 -H 8.1 Hz, 4 J 4 -H, -H 1.2 or 0. Hz, 5 J 4 -H,7 -H 1.2 or 0. Hz, 4 -H), 7.89 (1H, ddd, 3 J -H,7 -H 8.0 Hz, 4 J 5 -H,7 -H 1.3 or 0.7 Hz, 5 J 4 -H,7 -H 1.3 or 0.7 Hz, 7 -H) and 9.3 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) 11. (C-4), 117.01 (C-), 121.12 (C-4 ), 121.4 (C-7 ), 122.45 (C-3), 124.13 (C- ), 12.27 (C-5 ), 127. (C-5), 134.75 (C-7a ), 14.1 (C-2), 148. (C-1), 153.71 (C-3a ) and 171.99 (C-2 ); m/z (EI, 70 4
ev) 275 (M +, 100%), 242 (M + H, 5), 17 (C 7 H 5 2 +, 8) and 115 (15); HRM (EI, M + ) found: 275.0051; calcd for C 13 H 9 2 O 2 : 275.0075. 2.4843 2.4902 2.491 3.3354 3.3398.8909.9179 7.0514 7.0589 7.0785 7.0859 7.1019 7.1092 7.247 7.281 7.2918 7.3154 7.3188 7.3879 7.3919 7.4154 7.4392 7.4431 7.778 7.8033 7.8735 7.8983 9.22 9.294 2 1 3 4 5 3b 2 3a 4 7a 5 7 0.993 1.810 2.410 1.93 1.572 38.45 38.9430 39.2215 39.5000 39.333 39.7783 39.9245 40.058 40.3351 11.04 117.0128 121.1251 121.35 122.4473 124.1282 12.28 127.575 134.7458 14.133 148.59 153.7138 171.9883 Fig. 2 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 3b in DMO-d 5
3.3. ynthesis and analytical data of 4-(benzo[d]oxazol-2 -ylthio)-3-methylbenzene-1,2- diol (3c) and 5-(benzo[d]oxazol-2 -ylthio)-3-methylbenzene-1,2-diol (5c) According to the general procedure, 3-methylcatechol (1b) (78 mg, 0.3 mmol), 2- mercaptobenzoxazole (2a) (7 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 24 h. Workup gave a mixture of 4-(benzo[d]oxazol-2 -ylthio)-3-methylbenzene-1,2-diol (3c) and 5- (benzo[d]oxazol-2 -ylthio)-3-methylbenzene-1,2-diol (5c) as a brown solid (123 mg, 90%), mp 181 183 C; R f = 0.4 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 28 (log ε, 4.25), 279 (4.23), 247 (4.24) and 20 (4.); ṽ max (atr)/cm -1 3471 (OH), 1592, 1500, 1452, 1235 and 1138; δ H (300 MHz; DMO-d ) of 3c 2.21 (3H, s, CH 3 ),.74 (1H, d, 3 J 5-H,-H 8.2 Hz, -H), 7.05 (1H, d, 3 J 5-H,-H 8.3 Hz, 5-H), 7.24 7.31 (2H, m, 5 -H and -H), 7.55 7.58 (2H, m, 4 - H and 7 -H), 8.8 (1H, br, 1-OH or 2-OH) and 9.85 (1H, br, 1-OH or 2-OH); δ H (300 MHz; DMO-d ) of 5c 2.13 (3H, s, CH 3 ),.94 (1H, d, 4 J 4-H,-H 3.0 Hz, 4-H),.9 (1H, d, 4 J 4-H,-H 3.0 Hz, -H), 7.24 7.31 (2H, m, 5 -H and -H), 7.55 7.58 (2H, m, 4 -H and 7 -H), 8.80 (1H, br, 1-OH or 2-OH) and 9.71 (1H, br, 1-OH or 2-OH); δ C (75 MHz; DMO-d ) of 3c 14.0 (CH 3 ), 110.20 (C-7 ), 113.29 (C-), 114.35 (C-4), 118.40 (C-4 ), 124.25 (C-5 or C- ), 124.57 (C-5 or C- ), 127.70 (C-5), 129.39 (C-3), 141.47 (C-3a ), 144.43 (C-2), 147.80 (C- 1), 151.23 (C-7a ) and 13.71 (C-2 ); δ C (75 MHz; DMO-d ) of 5c 15.79 (CH 3 ), 110.2 (C- 7 ), 112.80 (C-5), 118.40 (C-4 ), 119.45 (C-), 124.41 (C-5 or C- ), 124.2 (C-5 or C- ), 125.92 (C-3), 128.20 (C-4), 141.47 (C-3a ), 145.54 (C-1), 145.99 (C-2), 151.23 (C-7a ) and 13.82 (C-2 ); m/z (EI, 70 ev) 273 (M +, 100%), 240 (M + H, 7), 1 (18) and 73 (13); HRM (EI, M + ) found: 273.0445; calcd for C 14 H 11 O 3 : 273.040.
2.1291 2.2141 2.4842 2.4900 2.4958 3.3381.7327.704.9381.952.9597 7.0389 7.0 7.240 7.2587 7.24 7.279 7.2895 7.3271 7.3093 7.3044 7.551 7.553 7.552 7.5754 7.5822 7.5871 7.5919 7.005 7.02 7.19 7.221 8.829 8.87 8.901 9.9418 2 3 4 1 5 3c 2 3a 4 O 5 7a 7 + 2 3 4 1 5 2 O 5c 3a 4 5 7a 7 0.482 0.998 0.387 0.341 0.35 1.170 1.022 0.509 0.479 14.0395 15.792 38.70 38.9435 39.2213 39.4211 39.5000 39.7125 39.7787 40.0004 40.0572 40.335 110.201 110.218 112.8015 113.2892 114.345 118.3952 118.5012 119.4449 124.2483 124.4048 124.550 124.184 125.9239 127.949 128.2009 129.3927 141.4110 141.41 144.4308 145.543 145.9880 147.8042 151.2185 151.2320 13.704 Fig. 3 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 3c and 5c in DMO-d 7
3.4. ynthesis and analytical data of 4-(benzo[d]thiazol-2 -ylthio)-3-methylbenzene-1,2- diol (3d) and 5-(benzo[d]thiazol-2 -ylthio)-3-methylbenzene-1,2-diol (5d) According to the general procedure, 3-methylcatechol (1b) (78 mg, 0.3 mmol), 2- mercaptobenzothiazole (2b) (84 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave a mixture of 4-(benzo[d]thiazol-2 -ylthio)-3-methylbenzene-1,2-diol (3d) and 5- (benzo[d]thiazol-2 -ylthio)-3-methylbenzene-1,2-diol (5d) as a pale brown solid (138 mg, 95%); mp 18 188 C; R f = 0.74 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 289 (log ε, 4.20), 282 (4.21) and 211 (4.59); ṽ max (atr)/cm -1 320 (OH), 1453, 141, 1415, 1270 and 1010; δ H (300 MHz; DMO-d ) of 3d 2.2 (3H, s, CH 3 ),.81 (1H, d, 3 J 5-H,-H 8.4 Hz, -H), 7.11 (1H, d, 3 J 5-H,-H 8.3 Hz, 5-H), 7.27 7.31 (1H, m, -H), 7.41 (1H, t like ov, 3 J ~ 8.1 Hz, 5 -H), 7.78 (1H, d like ov, 3 J ~ 8.1 Hz, 4 -H), 7.87 (1H, t like ov, 3 J ~ 8.1 Hz, 7 -H) and 8.78 (2H, br, 1-OH and 2-OH); δ H (300 MHz; DMO-d ) of 5d 2.15 (3H, s, CH 3 ), 7.02 (1H, d, 4 J 4-H,-H 1.8 Hz, -H), 7.04 (1H, d, 4 J 4-H,-H 1.8 Hz, 4-H), 7.27 7.31 (1H, m, -H), 7.41 (1H, t like ov, 3 J ~ 8.1 Hz, 5 -H), 7.78 (1H, d like ov, 3 J ~ 8.1 Hz, 4 -H), 7.87 (1H, t like ov, 3 J ~ 8.1 Hz, 7 -H) and 9.02 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) of 3d 13.94 (CH 3 ), 113.80 (C-), 117.45 (C-4), 121.07 (C-4 ), 121.11 (C-7 ), 123.98 (C- ), 12.24 (C-5 ), 128.34 (C-5), 129.48 (C-3), 134.79 (C-7a ), 144.74 (C-2), 148.30 (C-1), 153.88 (C-3a ) and 171.94 (C-2 ); δ C (75 MHz; DMO-d ) of 5d 15.83 (CH 3 ), 115.83 (C-5), 119.79 (C-), 121.07 (C-4 ), 121.11 (C-7 ), 124.10 (C- ), 12.4 (C-3), 12.21 (C-5 ), 128.92 (C-4), 134.7 (C-7a ), 145.90 (C-1), 14.3 (C-2), 153.73 (C-3a ) and 172.14 (C-2 ); m/z (EI, 70 ev) 289 (M +, 100%), 25 (M + H, 8), 210 (1) and 17 (C 7 H 5 + 2, 18); HRM (EI, M + ) found: 289.024; calcd for C 14 H 11 O 2 2 : 289.0231. 8
2.1519 2.207 2.4843 2.4900 2.4958 3.350.7973.8250 7.004 7.0954 7.1231 7.2458 7.2491 7.2730 7.2842 7.294 7.2997 7.3079 7.3113 7.374 7.380 7.4071 7.427 7.4318 7.79 7.798 7.8400 7.85 7.8933 2 3 4 1 5 3d 2 3a 4 5 7a 7 + 2 3 4 1 5 2 5d 3a 4 5 7a 7 1.724 2.22 0.905 1.000 0.888 1.588 1.95 1.337 1.17 13.9321 15.8201 38.54 38.9430 39.221 39.3527 39.5000 39.54 39.7784 39.913 39.9337 39.9558 40.3355 40.0570 113.7971 115.821 117.4539 119.787 121.034 121.1052 121.042 121.440 123.9838 124.097 12.20 12.243 12.4588 128.344 128.9220 129.4709 134.705 134.7858 144.7387 145.8970 14.327 148.2949 153.72 153.8773 171.9379 172.1431 Fig. 4 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 3d and 5d in DMO-d 9
3.5. ynthesis and analytical data of 4-(benzo[d]oxazol-2 -ylthio)-3-methoxybenzene-1,2- diol (3e) and 5-(benzo[d]oxazol-2 -ylthio)-3-methoxybenzene-1,2-diol (5e) According to the general procedure, 3-methoxycatechol (1c) (88 mg, 0.3 mmol), 2- mercaptobenzoxazole (2a) (7 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave a mixture of 4-(benzo[d]oxazol-2 -ylthio)-3-methoxybenzene-1,2-diol (3e) and 5- (benzo[d]oxazol-2 -ylthio)-3-methoxybenzene-1,2-diol (5e) as a pale brown solid (140 mg, 9%), mp 150 152 C; R f = 0.43 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 28 (log ε, 4.18), 279 (4.19), 249 (4.19) and 208 (4.59); ṽ max (atr)/cm -1 3300 (OH), 2994 (C-H), 1499, 1450, 1200, 1131 and 1095; δ H (300 MHz; DMO-d ) of 3e 3.8 (3H, s, OCH 3 ),.7 (1H, d, 3 J 5-H,- H 8.5 Hz, -H),.9 (1H, d, 3 J 5-H,-H 8.4 Hz, 5-H), 7.23 7.31 (2H, m, 5 -H and -H), 7.5 7.3 (2H, m, 4 -H and 7 -H) and 9.22 (2H, br, 1-OH and 2-OH); δ H (300 MHz; DMO-d ) of 5e 3.74 (3H, s, OCH 3 ),.79 (1H, d, 4 J 4-H,-H 3.0 Hz, -H),.82 (1H, d, 4 J 4-H,-H 3.0 Hz, 4-H), 7.23 7.31 (2H, m, 5 -H and -H), 7.5 7.3 (2H, m, 4 -H and 7 -H) and 9.22 (2H, br, 1- OH and 2-OH); δ C (75 MHz; DMO-d ) of 3e 0.1 (OCH 3 ), 108.22 (C-4), 110.11 (C-7 ), 111.71 (C-), 118.41 (C-4 ), 124.2 (C-5 or C- ), 124.58 (C-5 or C- ), 12.21 (C-5), 139.41 (C-2), 141.50 (C-3a ), 149.37 (C-1), 150.05 (C-3), 151.1 (C-7a ) and 13.3 (C-2 ); δ C (75 MHz; DMO-d ) of 5e 5.05 (OCH 3 ), 110.20 (C-7 ), 110.29 (C-4), 113.11 (C-5), 11.11 (C-), 118.54 (C-4 ), 124.44 (C-5 or C- ), 124.4 (C-5 or C- ), 13.78 (C-2), 141.44 (C-3a ), 14.38 (C-1), 148.7 (C-3), 151.24 (C-7a ) and 13.59 (C-2 ); m/z (EI, 70 ev) 289 (M +, 17%), 287 (M + 2, 100), 274 (M + CH 3, 15), 25 (M + H, 20), 255 (50) and 240 (0); HRM (EI, M + ) found: 289.0413; calcd for C 14 H 11 O 4 : 289.0409. 10
2.4900 3.2882 3.3449 3.347 3.3528 3.355 3.304 3.337 3.355 3.37 3.3743 3.378 3.4011 3.4351 3.4557 3.457 3.835 3.7342 3.731.579.81.7941.8234.9789 7.009 7.292 7.2871 7.2994 7.3122 7.5892 7.040 7.173 7.324 O 2 3 4 1 5 3e 2 3a 4 O 5 7a 7 + O 2 3 4 1 5 2 5e 4 5 3a O 7a 7 0.382 1.753 0.151 0.999 0.135 1.345 1.204 1.58 38.48 38.710 38.824 38.9432 39.077 39.099 39.221 39.3479 39.5000 39.214 39.7783 40.3350 40.211 40.2091 40.059 39.9375 5.1329 0.134 108.2155 110.4912 110.2930 110.195 111.7084 113.10 11.1051 118.4054 118.5417 124.253 124.445 124.5218 124.581 124.434 12.210 13.7799 139.404 141.4375 141.4995 14.3832 148.725 149.34 150.0518 151.1581 151.2358 13.5902 13.325 Fig. 5 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 3e and 5e in DMO-d 11
3.. ynthesis and analytical data of 4-(benzo[d]thiazol-2 -ylthio)-3-methoxybenzene-1,2- diol (3f) and 5-(benzo[d]thiazol-2 -ylthio)-3-methoxybenzene-1,2-diol (5f) According to the general procedure, 3-methoxycatechol (1c) (88 mg, 0.3 mmol), 2- mercaptobenzothiazole (2b) (84 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave a mixture of 4-(benzo[d]thiazol-2 -ylthio)-3-methoxybenzene-1,2-diol (3f) and 5- (benzo[d]thiazol-2 -ylthio)-3-methoxybenzene-1,2-diol (5f) as a pale brown solid (145 mg, 95%); mp 108 110 C; R f = 0.55 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 279 (log ε, 4.19) and 217 (4.55); ṽ max (atr)/cm -1 3400 (OH), 3058 (C-H), 1599, 1504, 1420, 119 and 1084; δ H (300 MHz; DMO-d ) of 3f 3.72 (3H, s, OCH 3 ),.72 (1H, d, 3 J 5-H,-H 8.3 Hz, -H), 7.07 (1H, d, 3 J 5-H,-H 8.3 Hz, 5-H), 7.31 (1H, t like ov, 3 J ~ 8.0 Hz, -H), 7.42 (1H, t like ov, 3 J ~ 8.0 Hz, 5 -H), 7.78 (1H, d like ov, 3 J ~ 7.8 Hz, 4 -H), 7.90 (1H, d like ov, 3 J ~ 7.8 Hz, 7 -H) and 9.27 (2H, br, 1-OH and 2-OH); δ H (300 MHz; DMO-d ) of 5f 3.78 (3H, s, OCH 3 ),.84 (1H, d, 4 J 4-H,-H 2.0 Hz, -H),.89 (1H, d, 4 J 4-H,-H 2.0 Hz, 4-H), 7.31 (1H, t like ov, 3 J ~ 8.0 Hz, - H), 7.42 (1H, t like ov, 3 J ~ 8.0 Hz, 5 -H), 7.78 (1H, d like ov, 3 J ~ 7.8 Hz, 4 -H), 7.90 (1H, d like ov, 3 J ~ 7.8 Hz, 7 -H) and 9.27 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) of 3f 5.18 (OCH 3 ), 111.04 (C-4), 112.15 (C-), 121.0 (C-4 ), 121.5 (C-7 ), 124.03 (C- ), 12.20 (C-5 ), 127.32 (C-5), 134.71 (C-7a ), 139.3 (C-2), 149.72 (C-1), 150.8 (C-3), 153.72 (C-3a ) and 19.92 (C-2 ); δ C (75 MHz; DMO-d ) of 5f 5.92 (OCH 3 ), 111.01 (C-4), 115.98 (C-5), 11.54 (C-), 121.13 (C-4 ), 121.5 (C-7 ), 124.14 (C- ), 12.2 (C-5 ), 134.77 (C-7a ), 137.44 (C-2), 14.74 (C-1), 149.13 (C-3), 153.70 (C-3a ) and 171.88 (C-2 ); m/z (EI, 70 ev) 307 (M + + 2, 45%), 305 (M +, 100), 290 (M + CH 3, 28) and 274 (M + OCH 3, 24); HRM (EI, M + ) found: 305.0122; calcd for C 14 H 11 O 3 2 : 305.0180; m/z (EI) 328 (M + + a, 100%) and 30 (M + + 1, 30); HRM (EI, M + + a) found: 328.007; calcd for C 14 H 11 O 3 2 a: 328.0073. 12
2.4844 2.4900 2.4957 3.727 3.7805.7099.7381.8411.8479.8870.8938 7.0343 7.024 7.284 7.2715 7.279 7.2950 7.318 7.4170 7.3935 7.389 7.352 7.3219 7.434 7.4407 7.4445 7.452 7.752 7.79 7.7817 7.7908 7.8082 7.8493 7.875 7.914 7.9412 O 2 3 4 1 5 3f 2 3a 4 5 7a 7 + O 2 3 4 1 5f 5 2 4 5 3a 7a 7 0.34 2.020 0.10 0.999 0.112 1.071 1.038 0.855 0.919 38.50 38.7741 38.7913 38.9432 39.0353 39.085 39.2217 39.3451 39.5000 39.310 39.479 40.4148 40.3355 40.2577 40.2344 40.058 39.9171 39.7785 5.9435 5.9287 5.1837 112.1530 111.0429 111.0122 11.5444 115.9835 121.748 121.510 121.5790 121.555 121.4394 121.1275 121.0571 121.8180 121.8388 124.0257 124.1392 124.2543 124.3424 12.1973 12.2572 134.7139 134.775 137.434 14.731 149.1333 150.773 153.4832 153.92 153.7228 171.8828 Fig. 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 3f and 5f in DMO-d 13
3.7. ynthesis and analytical data of 4-(benzo[d]oxazol-2 -ylthio)-3-fluorobenzene-1,2- diol (3g) and 5-(benzo[d]oxazol-2 -ylthio)-3-fluorobenzene-1,2-diol (5g) According to the general procedure, 3-fluorocatechol (1d) (81 mg, 0.3 mmol), 2- mercaptobenzoxazole (2a) (7 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave a mixture of 4-(benzo[d]oxazol-2 -ylthio)-3-fluorobenzene-1,2-diol (3g) and 5- (benzo[d]oxazol-2 -ylthio)-3-fluorobenzene-1,2-diol (5g) as a pale yellow solid (125 mg, 90%); mp 183 185 C; R f = 0.3 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 285 (log ε, 4.18), 278 (4.18) and 247 (4.21); ṽ max (atr)/cm -1 3470 (OH), 304 (C-H), 1500, 1452 and 1138; δ H (300 MHz; DMO-d ) of 3g.73 (1H, dd, 3 J 5-H,-H 8. Hz, 5 J -H,F 1.4 Hz, -H), 7.0 (1H, dd, 3 J 5-H,-H 8. Hz, 4 J 5-H,F 7. Hz, 5-H), 7.27 7.34 (2H, m, 5 -H and -H), 7.58 7.4 (2H, m, 4 -H and 7 -H) and 9.78 (2H, br, 1-OH and 2-OH); δ H (300 MHz; DMO-d ) of 5g 7.00 (1H, dd, 4 J 4-H,-H 1. or 2.1 Hz, 5 J -H,F 1. or 2.1 Hz, -H), 7.08 (1H, dd, 4 J 4-H,-H 2.2 Hz, 3 J 4-H,F 10.1 Hz, 4-H), 7.27 7.34 (2H, m, 5 -H and -H), 7.58 7.4 (2H, m, 4 -H and 7 -H) and 9.78 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) of 3g 101.77 (d, 2 J C-4,F 1.9 Hz, C-4), 110.34 (C-7 ), 111.88 (d, 4 J C-,F 2.2 Hz, C-), 118.5 (C-4 ), 124.54 (C-5 or C- ), 124.7 (C-5 or C- ), 12.28 (br, C-5), 134.45 (d, 2 J C-2,F 14.73 Hz, C-2), 141.32 (C-3a ), 151.24 (d, 3 J C-1,F.1 Hz, C-1), 151.31 (C-7a ), 152.30 (d, 1 J C-3,F 240.4 Hz, C-3) and 12.50 (C-2 ); δ C (75 MHz; DMO-d ) of 5g 113.45 (br, C-5), 113. (d, 2 J C-4,F 10. Hz, C-4), 110.34 (C-7 ), 118.08 (d, 4 J C-,F 2.2 Hz, C-), 118.4 (C-4 ), 124.54 (C-5 or C- ) 124.7 (C-5 or C- ), 135.97 (d, 2 J C-2,F 13.7 Hz, C-2), 141.32 (C-3a ), 148.18 (d, 3 J C-1,F.5 Hz, C-1), 151.31 (C-7a ), 151.4 (d, 1 J C-3,F 240.5 Hz, C-3) and 13.01 (C-2 ); m/z (EI, 70 ev) 277 (M +, 85%), 257 (M + HF, 100), 22 (1), 171 (15), 151 (C 7 H 5 O +, 19) and 119 (18); HRM (EI, M + ) found: 277.0194; calcd for C 13 H 8 FO 3 : 277.0209. 14
2.4850 2.4900 3.327.7117.7403.9417.94 7.0225 7.050 7.074 7.2819 7.2859 7.2958 7.3055 7.3093 7.3119 7.5720 7.575 7.5884 7.5947 7.037 7.092 7.211 7.252 F 2 3 4 1 5 3g 2 3a 4 O 5 7a 7 + F 2 3 4 1 5g 5 2 3a O 4 5 7a 7 1.000 0.347 1.1 2.19 2.11 38.54 38.9424 39.0511 39.0710 39.0892 39.0981 39.2212 39.345 39.3777 39.5000 39.445 40.333 40.1583 40.05 39.9558 39.7782 101.8870 101.23 110.3394 111.8941 111.845 113.7308 113.5903 113.4507 118.095 118.082 118.527 118.3 124.532 124.122 124.48 124.758 12.2818 134.3490 134.5455 135.8817 13.048 141.3224 148.1402 148.222 150.0409 150.977 151.2027 151.2840 151.3052 153.9025 12.5023 13.0139 Fig. 7 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 3g and 5g in DMO-d 15
3.8. ynthesis and analytical data of 4-(benzo[d]thiazol-2 -ylthio)-3-fluorobenzene-1,2- diol (3h) and 4-(benzo[d]thiazol-2 -ylthio)-3-fluorobenzene-1,2-diol (5h) According to the general procedure, 3-fluorocatechol (1d) (81 mg, 0.3 mmol), 2- mercaptobenzothiazole (2b) (84 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave a mixture of 4-(benzo[d]thiazol-2 -ylthio)-3-fluorobenzene-1,2-diol (3h) and 5- (benzo[d]thiazol-2 -ylthio)-3-fluorobenzene-1,2-diol (5h) as a pale yellow solid (127 mg, 8%); mp 177 179 C; R f = 0.3 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 277 (log ε, 4.19) and 208 (4.53); ṽ max (atr)/cm -1 323 (OH), 1578, 1414, 127 and 1011; δ H (300 MHz; DMOd ) of 3h.78 (1H, dd, 3 J 5-H,-H 8.4 Hz, 5 J -H,F 1.4 Hz, -H), 7.11 (1H, dd, 3 J 5-H,-H 8. Hz, 4 J 5- H,F 7.7 Hz, 5-H), 7.31 (1H, t like ov, 3 J ~ 7.7 Hz, -H), 7.43 (1H, ddd, 3 J 4 -H,5 -H 7.3 or 8.4 Hz, 3 J 5 -H, -H 7.3 or 8.4 Hz, 4 J 5 -H,7 -H 1.4 Hz, 5 -H), 7.81 (1H, d like ov, 3 J 4 -H,5 -H 8.1 Hz, 4 -H), 7.90 (1H, ddd, 3 J -H,7 -H 7.8 Hz, 4 J 5 -H,7 -H 1.3 or 0.7 Hz, 5 J 4 -H,7 -H 1.3 or 0.7 Hz, 7 -H) and 9.78 (2H, br, 1-OH and 2-OH); δ H (300 MHz; DMO-d ) of 5h 7.00 (1H, dd, 4 J 4-H,-H 1. or 2.2 Hz, 5 J -H,F 1. or 2.2 Hz, -H), 7.15 (1H, dd, 3 J 4-H,F 10.0 Hz, 4 J 4-H,-H 2.1 Hz, 4-H), 7.31 (1H, t like ov, 3 J ~ 7.7 Hz, -H), 7.43 (1H, ddd, 3 J 4 -H,5 -H 7.3 or 8.4 Hz, 3 J 5 -H, -H 7.3 or 8.4 Hz, 4 J 5 - H,7 -H 1.4 Hz, 5 -H), 7.81 (1H, d like ov, 3 J 4 -H,5 -H 8.1 Hz, 4 -H), 7.90 (1H, ddd, 3 J -H,7 -H 7.8 Hz, 4 J 5 -H,7 -H 0.7 or 1.3 Hz, 5 J 4 -H,7 -H 0.7 or 1.3 Hz, 7 -H) and 9.78 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) of 3h 104. (d, 2 J C-4,F 1.7 Hz, C-4), 112.31 (d, 4 J C-,F 2.5 Hz, C-), 121.27 (C-4 ), 121.73 (C-7 ), 124.33 (C- ), 12.38 (C-5 ), 12.99 (br, C-5), 134.5 (d, 2 J C-2,F 14.9 Hz, C-2), 134.77 (C-7a ), 151.7 (d, 3 J C-1,F.3 Hz, C-1), 152.32 (d, 1 J C-3,F 240.4 Hz, C- 3), 153.5 (C-3a ) and 170.10 (C-2 ); δ C (75 MHz; DMO-d ) of 5h 114.28 (d, 2 J C-4,F 20.1 Hz, C-4), 11.1 (d, 3 J C-5,F 10.3 Hz, C-5), 118.9 (d, 4 J C-,F 1. Hz, C-), 121.27 (C-4 ), 121.73 (C-7 ), 124.33 (C- ), 12.38 (C-5 ), 134.77 (C-7a ), 13.8 (d, 2 J C-2,F 13.8 Hz, C-2), 148.59 (d, 3 J C-1,F.7 Hz, C-1), 151.18 (d, 1 J C-3,F 240.4 Hz, C-3), 153.59 (C-3a ) and 170.2 (C-2 ); m/z (EI, 70 ev) 293 (M +, 100%), 273 (M + HF, 79), 20 (M + H, 4), 244 (8) and 17 (C 7 H 5 + 2, 10); m/z (EI) 31 (M + + a, 100%) and 294 (M + + 1, 30); HRM (EI, M + + a) found: 315.9893; calcd for C 13 H 8 FO 2 2 a: 315.9873. 1
2.4842 2.4900 2.4958 3.3440 3.3492 3.3515 3.3543.759.7704.7944.7990.9925.9984 7.0040 7.0828 7.1100 7.1233 7.1308 7.138 7.3385 7.3124 7.2880 7.4035 7.4074 7.4311 7.4337 7.4548 7.458 7.795 7.822 7.8891 7.8980 7.9003 7.9152 7.9244 F 2 3 4 1 5 3h 2 3a 4 5 7a 7 + F 2 3 4 1 5 2 5h 4 5 3a 7a 7 1.000 0.40 0.52 0.4 1.707 1.700 1.379 1.504 38.4 38.9432 39.2217 39.5000 39.7785 40.0570 40.3351 104.541 104.789 112.2925 112.325 114.1495 114.4179 11.0989 11.231 118.793 118.7009 121.24 121.730 124.3307 12.3807 12.9859 134.5498 134.70 13.5892 13.7729 148.5458 148.347 150.2289 150.7130 151.7210 151.8045 153.4339 153.5850 153.548 153.7024 153.9183 170.1039 170.227 Fig. 8 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 3h and 5h in DMO-d 17
3.9. ynthesis and analytical data of 4-(benzo[d]oxazol-2 -ylthio)-5-methylbenzene-1,2- diol (a) According to the general procedure, 4-methylcatechol (1h) (78 mg, 0.3 mmol), 2- mercaptobenzoxazole (2a) (7 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave 4- (benzo[d]oxazol-2 -ylthio)-5-methylbenzene-1,2-diol (a) as a pale yellow solid (117 mg, 85%); mp 198 200 C (lit. 1 198 200 C); R f = 0.53 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 287 (log ε, 4.27), 280 (4.23), 24 (4.28) and 20 (4.71); ṽ max (atr)/cm -1 3200 (OH), 3141, 149 1453, 1235 and 1132; δ H (300 MHz; DMO-d ) 2.22 (3H, s, CH 3 ),.80 (1H, s, -H), 7.04 (1H, s, 3-H), 7.25 7.32 (2H, m, 5 -H and -H), 7.5 7.1 (2H, m, 4 -H and 7 -H), 9.20 (1H, s, 1-OH or 2-OH), 9.48 (1H, s, 1-OH or 2-OH); δ C (75 MHz; DMO-d ) 19. (CH 3 ), 110.23 (C-7 ), 112.7 (C-4), 118.07 (C-), 118.44 (C-4 ), 123.1 (C-3), 124.31 (C-5 or C- ) 124.59 (C-5 or C- ), 133.58 (C-5), 141.45 (C-3a ), 143.99 (C-2), 148.17 (C-1), 151.25 (C- 7a ) and 13.51 (C-2 ); m/z (EI, 70 ev) 273 (M +, 100%), 240 (M + H, 40), 151 (C 7 H 5 O +, 42) and 124 (C 7 H 8 O + 2, 32); HRM (EI, M + ) found: 273.0455; calcd for C 14 H 11 O 3 : 273.040. 2.2155 2.4841 2.4898 2.495 3.3495.790 7.041 7.2473 7.255 7.2705 7.2829 7.2952 7.3001 7.5594 7.54 7.5743 7.5811 7.5895 7.598 7.05 7.11 9.194 9.4830 2 1 3 4 5 a 2 3a O 7a 7 4 5 3.250 1.1 1.104 2.371 1.929 1.050 1.000 18
19.13 38.54 38.9433 39.2217 39.5000 39.778 40.0571 40.3353 110.2297 112.735 118.0700 118.431 123.1572 124.301 124.5909 133.577 141.4492 143.980 148.1717 151.2525 13.5157 Fig. 9 1 H (300 MHz) and 13 C (75 MHz) MR spectra of a in DMO-d 3.10. ynthesis and analytical data of 4-(benzo[d]thiazol-2 -ylthio)-5-methylbenzene-1,2- diol (b) According to the general procedure, 4-methylcatechol (1h) (78 mg, 0.3 mmol), 2- mercaptobenzothiazole (2b) (84 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave 4- (benzo[d]thiazol-2 -ylthio)-5-methylbenzene-1,2-diol (b) as a white solid (131 mg, 90%); mp 187 189 C; R f = 0.50 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 299 (log ε, 4.15), 290 (4.21), 282 (3.19) and 210 (4.1); ṽ max (atr)/cm -1 3248 (OH), 3054 (C-H), 1493, 141, 1278 and 1029; δ H (300 MHz; DMO-d ) 2.25 (3H, s, CH 3 ),.85 (1H, s, -H), 7.08 (1H, s, 3-H), 7.2 (1H, ddd, 3 J 5 -H, -H 7.3 or 8.1 Hz, 3 J -H,7 -H 7.3 or 8.1 Hz, 4 J 4 -H, -H 1.2 Hz, -H), 7.42 (1H, ddd, 3 J 4 -H,5 -H 7.3 or 8.4 Hz, 3 J 5 -H, -H 7.3 or 8.4 Hz, 4 J 5 -H,7 -H 1.3 Hz, 5 -H), 7.78 (1H, ddd, 3 J 4 -H,5 -H 8.1 Hz, 4 J 4 -H, -H 1.1 or 0. Hz, 5 J 4 -H,7 -H 1.1 or 0. Hz, 4 -H), 7.87 (1H, ddd, 3 J -H,7 -H 7.9 Hz, 4 J 5 -H,7 -H 1.3 or 0.7 Hz, 5 J 4 -H,7 -H 1.3 or 0.7 Hz, 7 -H) and 9.55 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) 19.52 (CH 3 ), 115.87 (C-4), 118.39 (C-), 121.08 (C-4 ), 121.4 (C-7 ), 123.47 (C-3), 124.04 (C- ) 12.24 (C-5 ), 133.99 (C-5), 134.79 (C-7a ), 144.52 (C-2), 148.88 (C-1), 153.91 (C-3a ) and 171.75 (C-2 ); m/z (EI, 70 ev) 289 (M +, 19
57%), 25 (M + H, 100), 210 (34), 17 (C 7 H 5 2 +, 34) and 123 (34); HRM (EI, M + ) found: 289.0237; calcd for C 14 H 11 O 2 2 : 289.0231. 2.2518 2.4842 2.4900 2.4958 3.4913.8523 7.081 7.2531 7.254 7.2804 7.3038 7.3071 7.3815 7.3853 7.4091 7.4328 7.435 7.795 7.793 7.8540 7.8801 2 3 4 2 3a 4 1 5 b 7a 7 5 3.383 1.077 1.000 1.19 1.219 0.998 1.100 19.5231 38.49 38.9430 39.221 39.5000 39.7785 40.059 40.3359 115.8707 118.3938 121.0804 121.443 123.48 124.0355 12.2432 133.9873 134.7883 144.5218 148.8838 153.9145 171.742 Fig. 10 1 H (300 MHz) and 13 C (75 MHz) MR spectra of b in DMO-d 20
3.11. ynthesis and analytical data of 4-(benzo[d]oxazol-2 -ylthio)-5-ethylbenzene-1,2- diol (c) According to the general procedure, 4-ethylcatechol (1i) (87 mg, 0.3 mmol), 2- mercaptobenzoxazole (2a) (7 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave 4- (benzo[d]oxazol-2 -ylthio)-5-ethylbenzene-1,2-diol (c) as a pale yellow solid (120 mg, 83%); mp 141 143 C; R f = 0.50 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 287 (log ε, 4.19), 248 (4.13) and 20 (4.55); ṽ max (atr)/cm -1 3233 (OH), 1502, 1453 and 1133; δ H (300 MHz; DMO-d ) 1.04 (3H, t, 3 J 1 -H,2 -H 7.5 Hz, 2 -H), 2.1 (2H, q, 3 J 1 -H,2 -H 7.5 Hz, 1 -H),.80 (1H, s, -H), 7.04 (1H, s, 3-H), 7.24 7.30 (2H, m, 5 -H and -H), 7.5 7.1 (2H, m, 4 -H and 7 -H), 9.44 (1H, s, 1-OH or 2-OH) and 9.59 (1H, s, 1-OH or 2-OH); δ C (75 MHz; DMOd ) 15.44 (C-2 ), 2.27 (C-1 ), 110.23 (C-7 ), 112.10 (C-4), 11.5 (C-), 118.47 (C-4 ), 123.47 (C-3), 124.34 (C-5 or C- ) 124.1 (C-5 or C- ), 139.50 (C-5), 141.47 (C-3a ), 144.09 (C-2), 148.43 (C-1), 151.24 (C-7a ) and 13.84 (C-2 ); m/z (EI, 70 ev) 287 (M +, 100%), 254 (M + H, 30), 180 (58), 151 (C 7 H 5 O +, 80) and 123 (44); HRM (EI, M + ) found: 287.018; calcd for C 15 H 13 O 3 : 287.01. 1.011 1.0238 1.037 1.01 2.4840 2.4900 2.4959 2.5731 2.5981 2.232 2.482 3.3835.798 7.038 7.2434 7.215 7.28 7.2790 7.2912 7.295 7.5590 7.540 7.5744 7.5827 7.5881 7.003 7.052 2 3 4 2 3a 4 1 5 1 2 c O 7a 7 5 2.879 2.032 1.000 0.883 2.02 1.552 1.530 21
15.4383 2.278 38.2 38.9431 39.2217 39.3394 39.340 39.5000 39.300 39.7785 40.0572 40.3355 110.2313 112.0935 11.539 118.43 123.4742 124.3425 124.107 139.4549 141.40 144.080 148.4337 151.2438 13.8443 Fig. 11 1 H (300 MHz) and 13 C (75 MHz) MR spectra of c in DMO-d 3.12. ynthesis and analytical data of 4-(benzo[d]thiazol-2 -ylthio)-5-ethylbenzene-1,2- diol (d) According to the general procedure, 4-ethylcatechol (1i) (87 mg, 0.3 mmol), 2- mercaptobenzothiazole (2b) (84 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave 4- (benzo[d]thiazol-2 -ylthio)-5-ethylbenzene-1,2-diol (d) as a pale yellow solid (12 mg, 83%); mp 152 154 C; R f = 0.54 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 290 (log ε, 4.24), 282 (4.24) and 211 (4.3); ṽ max (atr)/cm -1 3275 (OH), 3054 (C-H), 1498, 141, 12 and 1029; δ H (300 MHz; DMO-d ) 1.07 (3H, t, 3 J 1 -H,2 -H 7.2 Hz, 2 -H), 2.3 (2H, q, 3 J 1 -H,2 -H 7.2 Hz, 1 -H),.85 (1H, s, -H), 7.07 (1H, s, 3-H), 7.28 (1H, ddd, 3 J 5 -H, -H 7.2 or 8.0 Hz, 3 J -H,7 -H 7.2 or 8.0 Hz, 4 J 4 -H, -H 1.1 Hz, -H), 7.41 (1H, ddd, 3 J 4 -H,5 -H 7.2 or 8.0 Hz, 3 J 5 -H, -H 7.2 or 8.0 Hz, 4 J 5 -H,7 -H 1.2 Hz, 5 -H), 7.78 (1H, ddd, 3 J 4 -H,5 -H 8.1 Hz, 4 J 4 -H, -H 1.2 or 0.5 Hz, 5 J 4 - H,7 -H 1.2 or 0.5 Hz, 4 -H), 7.87 (1H, ddd, 3 J -H,7 -H 8.0 Hz, 4 J 5 -H,7 -H 1.3 or 0. Hz, 5 J 4 -H,7 -H 1.3 or 0. Hz, 7 -H) and 9.51 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) 15. (C- 2 ), 2.23 (C-1 ), 115.18 (C-4), 11.8 (C-), 121.0 (C-4 ), 121.3 (C-7 ), 123.7 (C-3), 124.02 (C- ), 12.23 (C-5 ), 134.7 (C-7a ), 139.84 (C-5), 144.55 (C-2), 149.05 (C-1), 22
153.83 (C-3a ) and 172.15 (C-2 ); m/z (EI, 70 ev) 303 (M +, 0%), 270 (M + H, 100), 255 (25), 17 (C 7 H 5 2 +, 42) and 123 (3); HRM (EI, M + ) found: 303.0381; calcd for C 15 H 13 O 2 2 : 303.0388. 1.0459 1.0709 1.0959 2.4845 2.4900 2.495 2.5910 2.159 2.410 2.0.8532 7.0725 7.2554 7.2795 7.3033 7.300 7.3814 7.3849 7.4089 7.432 7.431 7.79 7.795 7.85 7.8829 2 3 4 2 3a 4 1 5 1 7a 7 2 5 d 3.252 2.414 1.05 0.952 1.1 1.147 0.951 1.000 2.059 15.51 2.228 38.44 38.9430 39.221 39.5000 39.7785 40.057 40.3347 115.178 11.8572 121.021 121.287 123.2 124.0228 12.2337 134.700 139.8379 144.5452 149.0471 153.8258 172.140 Fig. 12 1 H (300 MHz) and 13 C (75 MHz) MR spectra of d in DMO-d 23
3.13. ynthesis and analytical data of 3-(benzo[d]oxazol-2 -ylthio)-4-nitrobenzene-1,2- diol (7a) According to the general procedure, 4-nitrocatechol (1j) (78 mg, 0.50 mmol), 2- mercaptobenzoxazole (2a) (7 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Column filtration (EtOAc/MeOH 1:2) gave 3-(benzo[d]oxazol-2 -ylthio)-4-nitrobenzene-1,2-diol (7a) as a yellow solid (130 mg, 85%); mp 223 225 C; R f = 0.22 (EtOAc/MeOH = 5:1); λ max (MeC)/nm 314 (log ε, 4.18) and 202 (4.1); ṽ max (atr)/cm -1 3327 (OH), 1587, 1303 and 1207; δ H (300 MHz; DMO-d ).84 (1H, t like, 3 J 5 -H, -H 7.1 Hz, 5 -H),.93 (2H, d like ov, 3 J ~ 7.5 Hz, 4 -H and 7 -H), 7.00 (1H, d, 3 J 5-H,-H 9.2 Hz, -H), 7.03 (1H, t like ov, -H), 8.0 (1H, d, 3 J 5-H,-H 9.3 Hz, 5-H) and 9.35 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) 115.11 (C-), 11.75 (C-7 ), 119.84 (C-5 ), 121.2 (C-4 ), 122.28 (C-5), 123.17 (C-3), 12.11 (C- ), 133.04 (C-4), 134.70 (C-3a ), 138.17 (C-2), 148.1 (C-7a ), 148.39 (C-1) and 159.83 (C-2 ); m/z (EI, 70 ev) 304 (M +, 100%), 258 (M + O 2, 5), 19 (78), 135 (38) and 80 (15); HRM (EI, M + ) found: 304.0154; calcd for C 13 H 8 2 O 5 : 304.0154. 2.4900 3.3297.8432.873.9148.9395.9998 7.0308 8.0424 8.0732 9.344 OH 1 2 3 5 4 O O 2 7a 2 3a 7a 7 4 5 1.175 2.037 2.247 0.998 1.107 24
38.47 38.9431 39.091 39.221 39.5000 39.344 39.7785 40.059 40.3352 115.112 11.7473 119.8418 121.202 122.2820 123.1704 12.1101 133.0349 134.7008 138.154 148.1578 148.3893 159.828 Fig. 13 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 7a in DMO-d 3.14. ynthesis and analytical data of 4-(4,5 -dihydrothiazol-2 -ylthio)benzene-1,2-diol (8a) According to the general procedure, catechol (1a) (9 mg, 0.3 mmol), 2-mercaptothiazoline (2c) (0 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 20 h. Workup gave 4-(4,5 dihydrothiazol-2 -ylthio)benzene-1,2-diol (8a) as a pale yellow solid (95 mg, 83%); mp 15 17 C (lit. 2 > 250 C); R f = 0.27 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 28 (log ε, 3.23) and 20 (4.14); ṽ max (atr)/cm -1 331 (OH), 154, 150, 1401, 1251 and 1029; δ H (300 MHz; DMO-d ) 3.2 (2H, t, 3 J 4 -H,5 -H 8.2 Hz, 5 -H), 4.1 (2H, t, 3 J 4 -H,5 -H 8.2 Hz, 4 -H),.7 (1H, d, 3 J 5-H,-H 7.2 Hz, -H),.87 (1H, dd, 3 J 5-H,-H 8.0 Hz, 4 J 3-H,5-H 2.1 Hz, 5-H),.90 (1H, d, 4 J 3- H,5-H 2.1 Hz, 3-H) and 9.44 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) 34.00 (C-5 ), 5.4 (C-4 ), 11.08 (C-), 117.07 (C-4), 122.1 (C-3) 127.37 (C-5), 145.1 (C-2), 147.87 (C-1) and 17.12 (C-2 ); m/z (EI, 70 ev) 227 (M +, 74%), 22 (M + 1, 100), 17 (M + C 2 H 4, 47) and 141 (1); HRM (EI, M + ) found: 227.0090; calcd for C 9 H 9 O 2 2 : 227.0075. 25
2.4842 2.4900 2.4957 3.235 3.228 3.2898 4.1302 4.1572 4.1844.7458.7730.8571.844.8843.891.9419.9491 3 2 4 5 1 5 8a 2 4 2.55 2.559 1.084 0.971 1.000 2.144 33.9939 38.58 38.942 39.2220 39.5000 39.778 40.058 40.332 5.404 11.0790 117.075 122.02 127.397 145.092 147.8708 17.1193 Fig. 14 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 8a in DMO-d 2
3.15. ynthesis and analytical data of 4-(4,5 -dihydrothiazol-2 -ylthio)-5- methylbenzene-1,2-diol (8b) According to the general procedure, 4-methylcatechol (1h) (78 mg, 0.3 mmol), 2- mercaptothiazoline (2c) (0 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave 4- (4,5 -dihydrothiazol-2 -ylthio)-5-methylbenzene-1,2-diol (8b) as a white solid (90 mg, 74%); mp 192 194 C (lit. 2 179 180 C); R f = 0.25 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 291 (log ε, 3.8) and 209 (4.59); ṽ max (atr)/cm -1 3270 (OH), 152, 142, 1401, 1288 and 1002; δ H (300 MHz; DMO-d ) 2.21 (3H, s, CH 3 ), 3.2 (2H, t, 3 J 4 -H,5 -H 8.1 Hz, 5 -H), 4.1 (2H, t, 3 J 4 - H,5 -H 8.2 Hz, 4 -H),.71 (1H, s, -H),.90 (1H, s, 3-H) and 9.24 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) 19.88 (CH 3 ), 33.95 (C-5 ), 5.51 (C-4 ), 11.37 (C-4), 117.3 (C-), 123.49 (C-3) 133.84 (C-5), 143.45 (C-2), 148.01 (C-1) and 1.82 (C-2 ); m/z (EI, 70 ev) 241 (M +, 100%), 22 (M + CH 3, 3), 208 (M + H, 22), 194 (50) and 154 (32); HRM (EI, M + ) found: 241.0204; calcd for C 10 H 11 O 2 2 : 241.0231. 2.2091 2.4842 2.4900 2.4958 3.2301 3.2573 3.2843 3.333 3.330 4.1284 4.1554 4.182.7047.9203 9.2341 9.2389 3 2 4 1 5 5 8b 2 4 2.910 2.517 2.058 1.070 1.000 1.31 27
19.879 33.9489 38.38 38.9433 39.0487 39.000 39.0810 39.0959 39.221 39.5000 39.324 39.558 39.7785 39.9212 39.929 39.909 40.057 40.157 40.333 5.5075 11.391 117.27 123.4931 133.8348 143.445 148.0050 1.8174 Fig. 15 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 8b in DMO-d 3.1. ynthesis and analytical data of 4-(4,5 -dihydrothiazol-2 -ylthio)-5-ethylbenzene- 1,2-diol (8c) According to the general procedure, 4-ethylcatechol (1i) (87 mg, 0.3 mmol), 2- mercaptothiazoline (2c) (0 mg, 0.50 mmol), methanol (3 ml), phosphate buffer (0.2 M, ph.0, 27 ml) and laccase (0 U, 10 mg, A. bisporus) were reacted for 17 h. Workup gave 4- (4,5 -dihydrothiazol-2 -ylthio)-5-ethylbenzene-1,2-diol (8c) as a pale yellow solid (105 mg, 82%); mp 19 171 C; R f = 0.24 (CH 2 Cl 2 /EtOAc = 5:1); λ max (MeC)/nm 290 (log ε, 3.1) and 210 (4.53); ṽ max (atr)/cm -1 3252 (OH), 157, 1435, 1290 and 1003; δ H (300 MHz; DMOd ) 1.08 (3H, t, 3 J 1 -H,2 -H 7.5 Hz, 2 -H), 2.0 (2H, q, 3 J 1 -H,2 -H 7.5 Hz, 1 -H), 3.25 (2H, t, 3 J 4 -H,5 -H 8.4 Hz, 5 -H), 4.15 (2H, t, 3 J 4 -H,5 -H 8.4 Hz, 4 -H),.71 (1H, s, -H),.91 (1H, s, 3- H) and 9.24 (2H, br, 1-OH and 2-OH); δ C (75 MHz; DMO-d ) 15.48 (C-2 ), 2.31 (C-1 ), 34.04 (C-5 ), 5.51 (C-4 ), 115.7 (C-4), 11.11 (C-), 123.85 (C-3), 139.3 (C-5), 143.49 (C-2), 148.24 (C-1) and 17.29 (C-2 ); m/z (EI, 70 ev) 255 (M +, 50%), 222 (M + H, 17), 208 (44) and 123 (100); HRM (EI, M + ) found: 255.0417; calcd for C 11 H 13 O 2 2 : 255.0388. 28
1.0500 1.0751 1.1000 2.4843 2.4900 2.495 2.5491 2.5740 2.5991 2.241 3.157 3.2212 3.2483 3.2752 3.3497 4.1197 4.14 4.1737.7074.9139 3 2 4 2 1 5 1 5 4 2 8c 3.098 1.981 2.737 2.013 1.0 1.000 1.501 15.4792 2.3097 33.9840 33.9940 34.0084 34.014 34.0352 38.57 38.9420 39.1485 39.2213 39.297 39.12 39.594 39.5000 39.4128 39.3809 39.3437 39.25 39.593 39.781 39.7782 39.8501 39.988 40.0049 40.0571 40.1128 40.330 5.5078 115.74 11.1088 123.843 139.32 143.4901 148.2382 17.2848 Fig. 1 1 H (300 MHz) and 13 C (75 MHz) MR spectra of 8c in DMO-d 29
4. Computational studies of compounds 3a and 3b Calculations reported in this paper were performed within Density Functional Theory, using the Gaussian 03 package. 3 13 C MR chemical shifts of selected compounds 3a and 3b were calculated as follows: the structures were optimized with the MM2 force field implemented in Chem3D Pro. 4 In the second step, the optimized structures were subsequently reoptimized at the AM1 level followed by the RHF/3-21G level and finally by B3LYP/-31G(d) level of theory within the Gaussian 03 package. In the final step, the 13 C MR chemical shielding of the reoptimized geometries were computed once at the mpw1pw91/-311+g(2d,p)//mpw1pw91/-31g(d) level of theory in the gas phase. 3 The references TM and benzene for the MTD approach according to arotti and Pellegrinet 5 were computed in the same manner as for 3a and 3b. For comparison with the experimental 13 C MR chemical shifts the computationally derived 13 C MR chemical shifts were calculated as follows: δ a = σ ref gas pahse σ a gas phase + δ ref where σ ref and σ a are the calculated MR isotropic magnetic shielding tensors of the reference compound and carbon a of the compound of interest: σ TM = 185.81 ppm and σ benzene = 54.41 ppm at the mpw1pw91/-311+g(2d,p)// mpw1pw91/-31g(d) level gas phase; δ ref represents the chemical shift of the reference compound in deuterated DMO: δ TM = 0 ppm; δ benzene = 128.27 ppm. An HP Compaq with a 2.39 GHz processor and 2 GB RAM was used for the calculations. 30
Fig. 17 3D structure of 3a 4.1. Cartesian of 3a ymbol X Y Z C 3.1477780-0.4840430 0.829520 C 3.7115840-0.7013480-0.4388140 C 3.1897990-0.03940-1.541980 C 2.1125270 0.8291420-1.3922370 C 1.554830 1.0388110-0.1340220 C 2.0830170 0.384980 0.9855140 O 3.7429190-1.181840 1.8430090 O 4.7555080-1.5451120-0.5803500 0.2325240 2.2118320 0.072940 C -1.1829770 1.181050 0.0722950 O -1.0498810-0.1314110-0.28520 C -2.3317350-0.12240-0.1923470 C -3.15590 0.449740 0.185200 31
-2.3753490 1.5881030 0.3514700 C -2.7811200-1.894340-0.4331550 C -4.1528990-2.0909980-0.2788750 C -5.0047980-1.0434210 0.098410 C -4.5253010 0.240870 0.3371720 H 3.37800-0.2170330-2.5129140 H 1.7044430 1.342380-2.2549740 H 1.50410 0.5595050 1.987240 H 3.297850-0.995950 2.78990 H 4.97380-1.8999870 0.29570 H -2.1112100-2.91140-0.725430 H -4.59500-3.07710-0.453580 H -.00090-1.2420720 0.202490 H -5.1817780 1.0520870 0.293810 Fig. 18 3D structure of 3b 4.2. Cartesian of 3b 32
ymbol X Y Z C -3.3044020 0.98710 0.7179180 C -3.9989020 0.038130-0.499440 C -3.5753440-0.3043180-1.4598500 C -2.40530-1.1104230-1.218480 C -1.7847920-1.0143530-0.008570 C -2.212120-0.1108720 0.9703280 O -3.8094270 1.23250 1.590070 O -5.070840 1.391990-0.7303230-0.422420-2.109720 0.320890 C 0.919470-1.0407230 0.1412380 2.590-1.7853290 0.2125980 C 3.2525780-0.194130 0.0079470 C 2.2090290 0.7492550-0.1030000 0.9284430 0.23010-0.0180700 C 4.5901870 0.1859100-0.0532420 C 4.8779310 1.53320-0.2293890 C 3.8523580 2.480910-0.3422380 C 2.5205900 2.1001980-0.2809250 H -4.1201130-0.333400-2.395030 H -2.12910-1.8117070-1.9731540 H -1.807220-0.0393470 1.9138900 H -3.242750 1.84280 2.38100 H -5.19100 1.955140 0.0485330 H 5.383190-0.5454140 0.032180 H 5.9132820 1.8537990-0.2811200 H 4.104820 3.527900-0.4807770 H 1.713040 2.8218210-0.387010 5. Calculation of the E-factor, atom economy, TO and TOF of the laccase-catalyzed domino reaction between catechol (1a) and 2-mercaptobenzoxazole (2a) Yield of 3a = 93% 5.1. Calculation of the E-factor 33
OH + H + laccase + MeOH + a 2 HPO 4 + ah 2 PO 4 + HCl O O OH 1a 2a 3a 9 mg 7 mg 10 mg 237.54 mg 125 mg 542.3 mg 14.4 mg 121 mg Total amount of the reactants (taking into account a loss of 10% of the solvent used) = 9 mg + 7 mg + 10 mg + 237.54 mg + 125 mg + 542.3 mg + 14.4 mg = 1074.24 mg. Amount of the final product = 121 mg. Amount of waste = 1074.24-121 = 953.24 mg E-factor = Amount of waste [kg]/amount of product [kg] = 953.24/121 = 7.88 kg kg -1. 5.2. Calculation of the atom economy 7 The atom economy of the reaction was calculated according to the following equation: Molecular weight of the desired product % Atom economy = 100. Molecular weight of all reactants OH + + H + 1/2 O 2 O OH O 1a 2a 3a 110 g/mol 151 g/mol 1 g/mol 259 g/mol H 2 O Atom economy = 100 259/277 = 94% 5.3. Calculation of TO Molecular weight of laccase from A. bisporus = 9 000 g/mol. 8 pecific activity of the laccase = U/mg. 0 U Laccase corresponds to 10 mg, ie 1.0417 10-7 mol = 1.0417 10-4 mmol. TO = Amount of the substrate consumed [mmol] / Amount of catalyst [mmol]. TO = 0.47 [mmol] / 1.0417 10-4 [mmol] = 4512. 5.4. Calculation of TOF TO TOF =. Time TOF = 4512 / 1 h = 282 h -1. 34
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