Molecules 2018, 23, 2117
12 of 17
4.67 (ddt, J = 143.1, 12.4, 7.1 Hz, 1 H), 4.17 (tt, J = 6.6, 2.8 Hz, 2 H), 1.70–1.56 (m, 2 H + 1.5 H), 1.39 (dq,
J = 14.6, 7.3 Hz, 2 H), 1.28 (dt, J = 7.1, 4.5 Hz, 1.5 H), 0.94 (t, J = 7.4 Hz, 3 H); 13C-NMR (101 MHz, CDCl3,
292 K): δ
[ppm] = 172.8 (13C, dd, J = 61.2, 1.2 Hz), 160.4 (s), 65.7 (s), 47.06 (13C, dd, J = 61.3, 34.8 Hz),
30.6 (s), 19.1 (s), 18.8 (13C, dd, J = 34.9, 1.3 Hz), 13.8 (s); ESI-MS: calcd: [(M + H)+] m/z 177.2, found:
[(M + H)+] m/z 177.2.The obtained data of the unlabeled substances matched those reported in [43].
5-Butoxy-4-[13C1]methyl[4,5-13C2]oxazole (10): A solution of butyl-N-formyl-[13C3]alaninate (1.24 g,
7.06 mmol, 1.00 eq.) in dichloromethane was added to a homogenous mixture of celite (2 g), calcium
oxide (2 g) and P2O5 (2.5 g, 17.6 mmol, 2.5 eq.) in dichloromethane (100 mL) under inert atmosphere.
The mixture was stirred at room temperature for 30 min followed by heating to reflux for 48 h. After the
first 24 h, additional 2.5 eq. P2O5 w◦ere added at rt and the heating was continued. A sat. Aqueous
solution of NaHCO3 was added at 0 C, the mixture was filtered and the aqueous phase extracted with
dichloromethane (3
×
50 mL). The combined organic phases were washed with brine (1
×
25 mL),
dried over Na2SO4 and the solvent removed under reduced pressure. The crude product was purified
via column chromatography (pentane/diethyl ether = 3/1) to obtain the product (0.68 g, 4.33 mmol,
62%) as yellow oil. 1H-NMR (400 MHz, CDCl3, 292 K):
δ
[ppm] = 7.37 (dd, J = 6.9, 4.4 Hz, 1 H),
4.08 (td, J = 6.6, 2.9 Hz, 2 H), 2.04 (ddd, J = 128, 7.2, 4.6 Hz, 3 H), 1.76–1.64 (m, 2 H), 1.52–1.41 (m, 2 H),
0.96 (t, J = 7.4 Hz, 3 H); 13C-NMR (101 MHz, CDCl3, 292 K): [ppm] = 154.6 (13C, dd, J = 97.5, 7.2 Hz),
δ
142.4–142.0 (m), 112.0 (13C, dd, J = 97.5, 56.4 Hz), 74.4–74.2 (m), 31.5 (d, J = 2.4 Hz), 18.9 (s), 13.8 (s),
10.1 (13C, dd, J = 56.4, 7.2 Hz); ESI-MS calcd: [(M + H)+] m/z 159.1,found: [(M + H)+] m/z 159.1;
TLC: Rf = 0.34 (pentane/diethyl ether = 3/1 [KMnO4]).The obtained data of the unlabeled substances
matched those reported in [42].
2-[13C1]Methyl-3-hydroxy-4,5-epoxydimethyl[2,3-13C2]pyridine (11): 5-Butoxy-4-[13C1]methyl[4,5-13C2]oxazole
(0.68 g, 4.33 mmol, 1.00 eq.), 2,5-dihydrofuran (9.63 mL, 130 mmol, 30.0 eq.) and trichloroacetic acid
◦
(0.15 g, 0.95 mmol) were heated to 210 C for 5 h in a pressure vial and thereafter allowed to cool
to room temperature overnight. The reaction mixture was concentrated under reduced pressure
and the crude product purified via column chromatography (diethyl ether/methanol = 10/0.5) to
1
obtain the product (0.40 mg, 2.63 mmol, 61%) as brownish solid. H-NMR (400 MHz, MeOD, 292 K):
δ
[ppm] = 7.78 (d, J = 9.0 Hz, 1 H), 5.06 (d, J = 11.0 Hz, 4 H), 2.42 (ddd, J = 127.8, 6.5, 2.8 Hz, 3 H);
13C-NMR (101 MHz, MeOD, 292 K): [ppm] = 149.57 (13C, d, J = 66.6 Hz), 145.99 (13C, dd, J = 66.8,
50.6 Hz), 137.16 (s), 137.08 (s), 130.4–130.1 (m), 72.85 (d, J = 2.4 Hz), 72.33 (m), 17.64 (13C, dd, J = 50.6,
δ
4.4 Hz); ESI-MS calcd: [(M + H)+] m/z 155.1, [(M
−
H)−] m/z 153.0,found: [(M + H)+] m/z 155.1,
[(M − H)−] m/z 153.0; TLC: Rf = 0.27 (diethyl ether/methanol = 10/0.5 [KMnO4]).
[13C3]Pyridoxine (PN, 1): A solution of 2-[13C1]methyl-3-hydroxy-4,5-epoxydimethyl[2,3-13C2]pyridine
(0.40 mg, 2.63 mmol, 1.00 eq.) in 6 mL 48% HBraq. was heated to reflux for one hour. After cooling
to rt, the solvent was evaporated under reduced pressure. Water (25 mL) and freshly prepared AgCl
(8 g) were added and the reaction heated to reflux for one hour. After filtration over a Celite-Pad
and washing with water, the solvent was removed under reduced pressure and the crude product
purified via column chromatography (dichloromethane/methanol = 10/1) to obtain an off-white
powder (0.45 g, 2.15 mmol, 82%). 1H-NMR (400 MHz, D2O, 292 K):
1 H), 5.00 (d, J = 3.9 Hz, 2 H), 4.80 (s, 2 H), 2.64 (ddd, J = 131.4, 6.8, 3.1 Hz, 3 H); 13C-NMR: (101 MHz,
δ [ppm] = 8.16 (d, J = 7.0 Hz,
D2O, 292 K):
127.4, 55.8 (d, J = 3.5 Hz), 54.5 (d), 12.1 (13C, dd, J = 46.5, 1.6 Hz); ESI-MS calcd: [(M
173.1, [(M HCl
H)−] m/z 171.1,found: [(M HCl + H)+] m/z 173.0, [(M – HCl
TLC: Rf = 0.28 (dichloromethane/methanol = 10/1 [KMnO4]).
δ
[ppm] = 150.5 (13C, dd, J = 72.7, 1.5 Hz), 140.5 (13C, dd, J = 72.9, 46.4 Hz), 138.5, 134.5,
−
HCl + H)+] m/z
H)−] m/z 171.5;
−
−
−
−
[13C3]N-(Pyridoxylidene)methylamine (13): A solution of potassium permanganate (0.18 g, 1.16 mmol,
0.60 eq.) in distilled water (20 mL) was added portion wise to a solution of [13C3]pyridoxine (0.40 g,
1.93 mmol, 1.00 eq.) in distilled water (10 mL) over the course of 1 h followed by heating to 70 ◦C
for 1 h. Afterwards, the reaction mixture was reduced to 1/3 of its volume under reduced pressure.