B. Spingler, S. Mundwiler, P. Ruiz-Sánchez, D. R. van Staveren, R. Alberto
FULL PAPER
mentary crystallographic data for this paper. They can be obtained
free of charge from The Cambridge Crystallographic Data Centre
via www.ccdc.cam.ac.uk/data_request/cif.
(125.8 MHz, CD3OD, 293 K): δ = 181.72, 180.27, 177.77, 177.64,
177.50, 175.86, 175.69, 175.47, 174.72, 174.36, 173.84, 173.05,
167.25, 167.05, 160.65, 149.63, 143.58, 138.90, 138.43, 135.60,
134.00, 131.59, 125.10, 124.06, 118.02, 112.62, 108.82, 105.24,
95.76, 88.11, 86.56, 83.84 (d, JC,P = 5.6 Hz), 76.49, 75.65, 73.77 (d,
JC,P = 6.1 Hz), 70.85, 62.84, 61.75, 60.86, 60.51, 57.79, 57.02, 55.73,
55.15, 53.10, 52.62, 46.95, 44.02, 43.19, 40.25, 38.59, 36.37, 35.45,
33.78, 33.63, 33.13, 32.52, 32.45, 29.79, 28.21, 27.78, 27.57, 21.09,
20.60, 20.51, 20.33 (d, JC,P = 3.8 Hz), 20.09, 17.65, 17.24, 16.53,
16.25, 14.77 ppm. 31P NMR (300 MHz, CD3OD, 300 K): δ = 1.373
ppm. ESI-MS: m/z = 1612 [M + Na]+, 1590 [M + H]+, 806 [M +
H + Na]2+, 795.1 [M + 2 H]2+. UV/Vis (methanol): λ (ε,
Lmol–1 cm–1) = 279.0 (13400), 361.1 (23300), 549.1 (7200) nm. Free
acid Rt = 10.6 min (A1/B). Full 1H and 13C NMR spectra are
shown in the Supporting Information.
Synthesis of Vitamin B12 b- and d-Acids 1 and 2: Vitamin B12
(1.88 g, 1.39 mmol) was hydrolysed in HCl (0.1 , 190 mL) as de-
scribed in literature.[8] The purification was modified in the follow-
ing way. After desalting the crude reaction mixture by phenol ex-
tractions, three fractions were isolated on a Dowex column. The
first fraction contained exclusively 2, the second one a mixture of
1 and 2, and the third one contained 2 and the e-acid. The second
fraction (mixture of 1 and 2) was separated by preparative HPLC
(column: Waters XTerra Prep RP8, 5 µm, 30ϫ100 mm; buffer A1;
gradient: 0.5%min–1 starting from 100% A1). The mixture of 2
and the e-acid was separated on the same system but using the
buffer A2. Compound 1 was isolated in a yield of 280.6 mg
(14.9%), compound 2 in a yield of 131.5 mg (7.0%), and the e-acid
was obtained in a yield of 94.26 mg (5.0%). Assignment of the
isomers was carried out by comparison with the HPLC retention
order given in literature.
Synthesis of 6: Complex [NEt4]2[ReBr3(CO)3] (75 mg, 0.09 mmol)
was dissolved in water (10 mL), and a solution of fully deprotected
3 (36 mg, 0.13 mmol) in methanol was added. The solution was
heated to 70–80 °C for 2 h. The proceeding of the reaction was
monitored by HPLC: the peak with Rt = 6.2 min ([Re(OH2)3-
(CO)3]+) disappeared, and a new peak (product 6) with Rt
=
Synthesis of 4: A solution of freshly prepared 3 (361 µmol) in water
(1 mL) was added to 1 (65.0 mg, 48.1 µmol). EDC (46.1 mg,
240 µmol) was added, and the pH was adjusted to 5.5 with NaOH
(0.1 ). After the mixture was stirred at room temperature for 15 h,
HPLC analysis exhibited about 50% of product 4. More EDC
(46.1 mg, 240 µmol) was added, but even extended stirring at room
temperature did not lead to a significant amount of formation of
more 4. The solvent was removed in vacuo, and the residue was
purified by preparative HPLC (buffer A1, gradient: 0.5%min–1
starting from 100% A1). The main fraction was collected, the sol-
vent was removed in vacuo, and the product was desalted to give
4 in a yield of 25.8 mg (16.2 µmol, 33.3%). Rt = 13.1 min (gradient
A1/B). 1H NMR (500 MHz, CD3OD, 300 K): δ = 8.43 (d, J =
4.9 Hz, 1 H, L12), 7.77 (t, J = 7.7 Hz, 1 H, L10), 7.58 (d, J =
7.8 Hz, 1 H, L9), 7.28 (t, J = 6.9 Hz, 1 H, L11), 7.25 (s, 1 H, B7),
7.12 (s, 1 H, B2), 6.56 (s, 1 H, B4), 6.27 (d, J = 3.0 Hz, 1 H, R1),
6.04 (s, 1 H, C10) ppm. 13C NMR (125.8 MHz, CD3OD, 293 K):
δ = 181.65, 180.20, 177.67, 176.73, 175.65, 175.58, 175.43, 174.72,
174.48, 174.34, 173.19, 167.22, 166.98, 160.49, 160.44, 149.67,
143.55, 138.86, 138.34, 135.72, 133.88, 131.56, 125.07, 124.04,
15.1 min appeared. After 2 h, the conversion was complete. When
the reaction mixture was cooled, a white powder precipitated from
solution. This precipitate was filtered, washed with small amounts
of cold water and dried in vacuo. Yield: 32 mg (72%); the rest re-
mained in the filtrate. It is important to apply temperatures above
1
70 °C in order to ensure ester hydrolysis during complexation. H
NMR (300 MHz, CD3OD, 300 K): δ = 8.83 (d, J = 5.7 Hz, 1 H),
8.11 (t, J = 7.8 Hz, 1 H), 7.74 (d, J = 8.1 Hz, 1 H), 7.56 (t, 1 H, J
= 6.9 Hz), 4.78 (d, 1 H, J = 15.6 Hz), 4.55 (d, 1 H, J = 15.9 Hz),
3.90 (d, 1 H, J = 17.1 Hz), 3.70 (m, 2 H), 3.53 (d, J = 16.8 Hz, 1
H), 3.04 (t, J = 7.5 Hz, 2 H), 2.18 (m, 2 H) ppm. ESI-MS: m/z =
493.80 [M + H]+. C12H18F3N3O7Re (6·TFA): calcd. C 25.76, H
3.24, N 7.51; found C 25.23, H 3.21, N 7.33.
Synthesis of 8: Two synthetic procedures are possible. The direct
reaction of 4 with [Re(OH2)3(CO)3]+ in water gives compound 8 in
good yield. Alternatively, the complex 6 can be preformed by the
reaction of fully deprotected 3 and [Re(OH2)3(CO)3]+ followed by
coupling to 1 as described in the following part. Compound 1
(26.7 mg, 19.8 µmol), complex
6 (29.2 mg, 60 µmol), EDC
117.92, 112.65, 108.77, 105.21, 95.76, 88.05, 86.50, 83.74 (d, JC,P
=
(11.5 mg, 60 µmol) and N-hydroxysuccinimide (6.9 mg, 60 µmol)
were dissolved in a mixture of water (5 mL) and DMSO (0.5 mL),
and the pH was adjusted to 5.5. After 5 h at room temperature,
HPLC analysis exhibited about 33% of product 8. More EDC and
N-hydroxysuccinimide were added and the stirring was continued
for 3 d with addition of EDC and N-hydroxysuccinimide in 24-h
intervals. The water was removed in vacuo, and the product was
precipitated by adding diethyl ether. The oily suspension was cen-
trifuged and decanted. Washing with diethyl ether was repeated
twice (5 mL each time) until a fine precipitate formed. The crude
product was dried at high vacuum, purified by preparative HPLC
(A1, gradient 1%min–1 starting from 100% A1) and desalted to
5.6 Hz), 76.40, 75.55, 73.69 (d, JC,P = 6.1 Hz), 71.66, 70.80, 70.56,
62.74, 61.79, 60.85, 60.45, 57.75, 56.97, 55.91, 55.67, 55.13, 53.31,
52.64, 50–49 (1 C below solvent), 46.91, 43.99, 43.16, 40.22, 38.88,
37.06, 35.43, 33.58, 33.09, 33.06, 32.42, 29.74, 28.01, 27.96, 27.52,
21.07, 20.61, 20.43, 20.26 (d, JC,P = 3.8 Hz), 20.02, 17.67, 17.16,
16.51, 16.21, 14.71 ppm. ESI-MS: m/z = 806.5 [M + H + Na]2+
,
795.6 [M + 2 H]2+. UV/Vis (methanol): λ (ε, Lmol–1 cm–1) = 278.0
1
(8500), 361.1 (26500), 549.1 (8000) nm. A full H NMR spectrum
is shown in the Supporting Information. The ester can be hydro-
lysed to the acid. Because of self-catalysed cleavage processes (see
above) this compound has to be labelled immediately. Labelling
can be performed directly with 4 without the need for prior de-
esterification. Rt = 10.8 min.
give 8 in a yield of 9.1 mg (23%). Rt = 14.2 min (gradient A1/B).
1
IR (KBr): ν = 2927, 2134, 2024, 1913, 1897, 1662 cm–1. H NMR
˜
Synthesis of 5: The procedure is similar to the preparation of 4.
Compound 2 (9.3 mg, 6.9 µmol) was treated with 3 (7 µmol) and
EDC (6.6 mg, 34 µmol) as described for the synthesis of 4. After
HPLC purification, the product 5 was isolated in a yield of 3.6 mg
(33%). Rt = 15.5 min (gradient A1/B). 1H NMR (500 MHz,
CD3OD, 300 K): δ = 8.41 (d, J = 4.9 Hz, 1 H, L12), 7.79 (t, J =
7.7 Hz, 1 H, L10), 7.56 (d, J = 7.8 Hz, 1 H, L9), 7.30 (t, J = 5.6 Hz,
(500 MHz, CD3OD, 300 K): δ = 8.1 (d, J = 4.5 Hz, 1 H, L12), 8.10
(t, J = 8 Hz, 1 H, L10), 7.73 (m, 1 H, L9), 7.54 (m, 1 H, L11), 7.26
(s, 1 H, B7), 7.14 (s, 1 H, B2), 6.59 (m, 1 H, B4), 6.27 (s, 1 H, R1),
6.05 (s, 1 H, C10) ppm. 13C NMR (125.8 MHz, CD3OD, 300 K):
δ = 198.39, 198.24, 197.46, 183.33, 181.75, 180.28, 177.72, 177.68,
176.77, 175.74, 175.69, 175.46 (2 C), 174.90, 174.76, 174.36, 167.27
(2 C), 160.90, 153.72, 143.63, 141.85, 138.39, 135.77, 133.97,
1 H, L11), 7.25 (s, 1 H, B7), 7.12 (s, 1 H, B2), 6.56 (s, 1 H, B4), 131.63, 127.20, 125.19, 117.99, 112.65, 105.25, 102.57, 95.74, 88.13,
6.27 (d, J = 3.0 Hz, 1 H, R1), 6.05 (s, 1 H, C10) ppm. 13C NMR
86.59, 83.83 (d, JC,P = 4.5 Hz), 76.52, 75.74, 73.72 (d, JC,P
=
2646
www.eurjic.org
© 2007 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Inorg. Chem. 2007, 2641–2647