Efficient synthesis of 13C-labelled erythromycin biosynthetic intermediate. 1: S-2-acetylaminoethyl (2R,3R,4R,5R)-3,5-diacetoxy-2,4-dimethyl- 4-([13C]methoxy)-heptanethioate

Article abstract of DOI:10.1002/jlcr.1507

An efficient ¹³C-labelling synthesis of the putative erythromycin biosynthetic intermediate, S-2-acetylaminoethyl (2R,3R,4R,5R)-3,5-diacetoxy-2,4-dimethyl-4-([¹³C]methoxy) heptanethioate, which would be useful for the investigation of the chain elongation mechanism in erythromycin biosynthesis, was achieved by utilizing iodo[¹³C]methane and (2S,3R,4R,5R)-4-hydroxy-3,5-O-isopropylidene-2, 4-dimethylheptanol, obtained in our previous studies on erythromycin A synthesis. Copyright

Full text of DOI:10.1002/jlcr.1507

Research Article
Received 3 October 2007,
Revised 22 January 2008,
Accepted 2 February 2008
Published online 27 March 2008 in Wiley Interscience
(www.interscience.wiley.com) DOI: 10.1002/jlcr.1507
13
Efficient synthesis of C-labelled
erythromycin biosynthetic intermediate.
1: S-2-acetylaminoethyl (2R,3R,4R,5R)-3,5-
diacetoxy-2,4-dimethyl-4-([13C]methoxy)-
heptanethioate
Ã
Katsumi Iida,a Masahiro Kajiwara,a Tomoko Inoue-Tanihata,b
Mineo Fukui{,b Tadashi Nakatay,b and Takeshi Oishizb
An efficient 13C-labelling synthesis of the putative erythromycin biosynthetic intermediate, S-2-acetylaminoethyl
(2R,3R,4R,5R)-3,5-diacetoxy-2,4-dimethyl-4-([13C]methoxy)heptanethioate, which would be useful for the investigation of
the chain elongation mechanism in erythromycin biosynthesis, was achieved by utilizing iodo[13C]methane and
(2S,3R,4R,5R)-4-hydroxy-3,5-O-isopropylidene-2,4-dimethylheptanol, obtained in our previous studies on erythromycin A
synthesis.
Keywords: 13C-labelling synthesis; erythromycin biosynthetic intermediate; iodo[13C]methane
Introduction
Results and discussion
Methods previously reported for the synthesis of erythromycin13
are unsuitable for efficient 13C-labelling, and we required a more
efficient synthetic strategy. Here, we focused on the introduc-
tion of a 13C-label into the optically active synthetic inter-
mediates obtained in our previous work.14,15
Initially, we looked at the 13C-labelled putative erythromycin
biosynthetic intermediate (3), which is the equivalent of the C-
9–C-21 segment of erythromycin A (1) shown inside the dotted
square in Scheme 1, though the incorporation of 3 in S.
erythraea might lead to 12-([13C]methoxy)erythromycin A (2). As
shown in Scheme 1, 3 should be obtainable from the primary
alcohol 5 via oxidation of the hydroxyl group, thioesterification
with 2-acetylaminoethanethiol, and deprotection of the
Erythromycin A, a 14-membered macrolide antibiotic produced
by Saccharopolyspora erythraea, is widely used in clinical
medicine. Corcoran et al. investigated the origin of the carbon
atoms of 6-deoxyerythronolide B, the first biosynthetic macro-
lide intermediate of erythromycin A, by means of feeding
experiments with 14C-labelled compounds in S. erythraea.1,2
Subsequently, Cane et al. evaluated the chain elongation
mechanism leading to 6-deoxyerythronolide B by means of
feeding experiments with 2H-, 13C-, and/or 18O-labelled
compounds, especially 2H- and/or 13C-labelled S-2-acetylami-
noethyl (2S,3R)-3-hydroxy-2-methylpentanethioate.,3–7 Groups
led by Leadlay and Katz employed a genetic approach to
examine the biosynthetic pathways to erythromycin.8–11 Based
on the domain organization of the 6-deoxyerythronolide B
synthase proteins encoded in the ery A region of the S. erythraea
genome, they predicted the biosynthetic pathways to 6-
deoxyerythronolide B.
{Deceased.
yPresent address: Department of Chemistry, Faculty of Science, Tokyo University
of Science, 1-3 Kagurazaka, Shinjuku-ku, Tokyo 162-8601, Japan.
We were interested in the chain elongation mechanism in
erythromycin biosynthesis, and aimed at approaching this
issue by using 13C-labelled erythromycin biosynthetic inter-
mediates. The intact incorporation of a 13C-labelled triketide
zRetired.
aDepartment of Medicinal Chemistry, Meiji Pharmaceutical University, 2-522-1
Noshio, Kiyose-shi, Tokyo 204-8588, Japan
into tylosine,
a 16-membered macrolide antibiotic, was
reported by Hutchinson and Coworkers,12 but the incorpora- bRIKEN (The Institute of Physical and Chemical Research), 2-1 Hirosawa, Wako-
shi, Saitama 351-0198, Japan
tion of other labelled putative polyketide intermediates into
erythromycin has not been described. Here, we describe the
efficient 13C-labelling synthesis of a putative erythromycin
biosynthetic intermediate.
*Correspondence to: Katsumi Iida, Department of Medicinal Chemistry, Meiji
Pharmaceutical University, 2-522-1 Noshio, Kiyose-shi, Tokyo 204-8588, Japan.
E-mail: iida@my-pharm.ac.jp
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K. Iida et al.
Scheme 1. Strategy for syntheses of 13C-labelled putative erythromycin intermediates 3 or 4 from 13CH3I and 6.
Scheme 2
acetonide. The primary alcohol 5 should be obtainable from diol 99% yield. Oxidation of 5 (unlabelled) with ruthenium (IV)
6 synthesized in our previous work14 via 13C-methylation of the oxide in the presence of sodium metaperiodate (NaIO4) in a
secondary
hydroxyl
group
with
iodo[13C]methane two-phase solution of CCl4, CH3CN, and 0.1 M Na phosphate
after regioselective protection of the primary hydroxyl group buffer at room temperature for 2 h gave acid 9 in a 68%
with tert-butyldiphenylsilyl chloride (TBDPSCl), followed yield. The thioesterification of 9 with 2-acetylaminoethanethiol
by desilylation. As shown in Scheme 2, we attempted to in DMF in the presence of diphenylphosphoryl azide and
synthesize
3
(unlabelled) using
6
and iodomethane. Et3N under an argon atmosphere at room temperature for 16 h
Regioselective silylation of the primary hydroxyl group of 6 gave thioester 10 in a 95% yield.6,16 However, deprotection of
with TBDPSCl in the presence of imidazole under an the acetonide of 10 did not afford diol 3 (unlabelled), but
argon atmosphere at room temperature for 1 h quantitatively d-lactone 11 was obtained in a 67% yield. The chair form of 11
afforded the secondary alcohol 7. Methylation of the secondary shown in Scheme
2 might be the lowest-energy chair
hydroxyl group of 7 with KH and iodomethane under an conformation, as C-2–Me, C-3–OH, and C-5–Et of 11 can take
argon atmosphere at 01C for 30 min and then at room equatorial positions. Therefore, 11 might be generated from
temperature for 30 min gave ether 8 (unlabelled) in an 10 through the attack of the 5-hydroxyl oxygen on the carbonyl
82% yield. The desilylation of 8 (unlabelled) with n-tetrabuty- carbon with the elimination of 2-acetylaminoethanethioxyl
lammonium fluoride (n-Bu4NF) in THF at room temperature after desilylation. Consequently, the synthesis of 3 appeared
for 96 h afforded the primary alcohol 5 (unlabelled) in a to be difficult.
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J. Label Compd. Radiopharm 2008, 51 213–217

K. Iida et al.
Scheme 3
As shown in Scheme 1, we next examined the synthesis of the
13C-labelled putative erythromycin biosynthetic intermediate 4,
in which the 3,5-dihydroxyl groups of 3 are protected with
acetyl groups to exclude formation of d-lactone 11. As shown in
Scheme 3, ether 8 was obtained via 13C-methylation of 7 with
iodo[13C]methane as before (Scheme 2). Treatment of 8 with
pyridinium p-toluenesulfonate (PPTS) in MeOH at room tem-
perature for 27 h gave diol 12. This reaction was repeated twice
more with recovered 8, and 12 was obtained in an overall yield
of 88%. Diacetylation of the hydroxyl groups of 12 in pyridine
and acetic anhydride in the presence of 4-dimethylaminopyr-
idine (DMAP) at room temperature for 23 h afforded ester 13 in
a 94% yield. The desilylation of 13 with n-Bu4NF in the presence
of acetic acid, which was added to decrease the loss of the
acetyl group from C-3 or C-5, in THF at room temperature for
89 h gave the primary alcohol 14 in a 78% yield. Oxidation of 14
with ruthenium (IV) oxide in the presence of NaIO4 in a two-
phase solution of CCl4, CH3CN, and 0.1 M Na phosphate buffer at
room temperature for 1 h provided acid 15 in an 81% yield. The
thioesterification of 15 with 2-acetylaminoethanethiol in DMF in
the presence of diphenylphosphoryl azide and Et3N under an
argon atmosphere at room temperature for 14 h gave the
desired thioester 4 in a 75% yield.
mixture was stirred for 1 h at room temperature. The reaction
was quenched with water (5 ml) at 01C, and the whole was
extracted with Et2O. The combined extract was washed with
water and brine, dried over anhydrous MgSO4, and evaporated
to give 7 (10.38 g, quant.); 1H NMR (CDCl3) d: 0.86 (d, 3H,
J = 6.7 Hz, 2-CH3), 1.00 (t, 3H, J = 7.0 Hz, 7-H3), 1.04 (s, 9H, Si-
C(CH3)3), 1.20 (s, 3H, 4-CH3), 1.29 (s, 3H, isopropylidene-CH3),
1.36 (s, 3H, isopropylidene-CH3), 1.38 (m, 2H, 6-H2), 2.16 (m, 1H,
2-H), 3.22 (s, 1H, 4-OH), 3.45 (dd, 1H, J = 2.8, 6.7 Hz, 5-H), 3.54 (d,
1H, J = 6.3 Hz, 3-H), 3.59 (dd, 1H, J = 4.9, 9.8 Hz, 1-H), 3.67 (dd, 1H,
J = 7.9, 9.8 Hz, 1-H), 7.65–7.68 (m, 10H, phenyl-H10).
(2S,3R,4R,5R)-1-tert-Butyldiphenylsilyloxy-3,5-O-isopropyli-
dene-2,4-dimethyl-4-([13C]methoxy)heptanol (8)
To a suspension of KH (50% in oil, 574 mg, 7.2 mmol; previously
washed with dry hexane) in dry THF (10 ml) was added
dropwise a solution of 7 (3.04 g, 6.5 mmol) in dry THF (6 ml) at
01C under an argon atmosphere. The suspension was stirred
for 30 min, then iodo[13C]methane (0.45 ml, 7.2 mmol) at 01C
was added dropwise under an argon atmosphere. The
whole was stirred for 30 min at 01C, and then for 30 min at
room temperature. The reaction was quenched with sat. NH4Cl
aq., and the mixture was extracted with Et2O. The combined
extract was washed with brine, dried over anhydrous MgSO4,
and evaporated. Chromatography of the crude product on silica
gel, eluting with Et2O:hexane (1:50–5:95) and then AcOEt/
Experimental
Materials and instruments
1
hexane (5:95–1:9–1:4), gave 8 (1.90 g, 82%); H NMR (CDCl3) d:
Iodo[13C]methane (99 atom% 13C) was purchased from Cam-
bridge Isotope Laboratories. (2S,3R,4R,5R)-4-Hydroxy-3,5-O-iso-
propylidene-2,4-dimethylheptanol (6) (495% e.e.) synthesized
in our previous work14 was used. All other chemicals were of
analytical grade and commercially available. 1H NMR and 13C
NMR spectra were recorded on a JEOL GX-400, GX-500, or GSX-
400 (1H: 400 or 500 MHz and 13C: 100 MHz) spectrometer. IR
spectra were recorded on a Jasco VALORA-III FT-IR spectrometer.
MS spectra were obtained on a JEOL JMS-DX302 spectrometer.
0.93 (d, 3H, J = 7.3 Hz, 2-CH3), 0.98 (t, 3H, J = 7.3 Hz, 7-H3), 1.05 (s,
9H, SiC(CH3)3), 1.08 (s, 3H, 4-CH3), 1.28 (s, 3H, isopropylidene-
CH3), 1.35 (s, 3H, isopropylidene-CH3), 1.38 (m, 1H, 6-H), 1.54 (m,
1H, 6-H), 2.01 (m, 1H, 2-H), 3.26 (d, 3H, J = 141 Hz, 4-O13CH3), 3.49
(dd, 1H, J = 4.9, 9.8 Hz, 1-H), 3.56 (dd, 1H, J = 7.9, 9.8 Hz, 1-H), 3.57
(dd, 1H, J = 2.4, 11.0 Hz, 5-H), 3.76 (d, 1H, J = 3.7 Hz, 3-H),
7.65–7.68 (m, 10H, phenyl-H10), data of
8 (unlabelled)
1
synthesized by using iodomethane; H NMR (CDCl3) d: 0.93 (d,
3H, J = 7.3 Hz, 2-CH3), 0.98 (t, 3H, J = 7.3 Hz, 7-H3), 1.05 (s, 9H,
SiC(CH3)3), 1.07 (s, 3H, 4-CH3), 1.27 (s, 3H, isopropylidene-CH3),
1.36 (s, 3H, isopropylidene-CH3), 1.38 (m, 1H, 6-H), 1.54
(m, 1H, 6-H), 2.01 (m, 1H, 2-H), 3.26 (s, 3H, 4-OCH3), 3.49 (dd,
1H, J = 4.9, 9.8 Hz, 1-H), 3.56 (dd, 1H, J = 7.9, 9.8 Hz, 1-H), 3.57 (dd,
1H, J = 2.2, 10.1 Hz, 5-H), 3.60 (d, 1H, J = 3.7 Hz, 3-H), 7.65–7.68
(m, 10H, phenyl-H10).
(2S,3R,4R,5R)-1-tert-Butyldiphenylsilyloxy-4-hydroxy-3,5-O-
isopropylidene-2,4-dimethylheptanol (7)
To a solution of 6 (4.97 g, 21.0 mmol) and imidazole (3.57 g,
52.4 mmol) in dry DMF (20 ml) was added dropwise TBDPSCl
(6.0 ml, 23.7 mmol) at 01C under an argon atmosphere. The
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K. Iida et al.
was added anhydrous MgSO4, and the whole was stirred for
30 min at room temperature. The suspension was filtered and
the filtrate was evaporated. Preparative TLC (0.5 mm ꢁ 20 cm2)
of the crude product on silica gel and development twice with
CHCl3/MeOH (20:1) gave 11 (19 mg, 67%); 1H NMR (CDCl3) d:
1.09 (t, 3H, J = 7.3 Hz, 7-H3), 1.18 (s, 3H, 4-CH3), 1.42 (d, 3H,
J = 7.0 Hz, 2-CH3), 1.61 (m, 1H, 6-H), 1.75 (m, 1H, 6-H), 2.26 (m, 1H,
3-OH), 2.47 (dq, 1H, J = 3.1, 7.0 Hz, 2-H), 3.37 (s, 3H, 4-OCH3), 3.72
(d, 1H, J = 10.1 Hz, 3-H), 3.97 (dd, 1H, J = 2.1, 10.4 Hz, 5-H).
(2S,3R,4R,5R)-3,5-O-Isopropylidene-2,4-dimethyl-4-methox-
yheptanol (5) (unlabelled)
To a solution of 8 (unlabelled) (891 mg, 2.5 mmol) in THF (3 ml)
was added n-Bu4NF ꢀ 3H2O (945 mg, 3.6 mmol) at room tem-
perature, and the mixture was stirred for 96 h. The reaction was
quenched with sat. NH4Cl aq., and the mixture was extracted
with Et2O. The combined extract was washed with brine, dried
over anhydrous MgSO4, and evaporated. Chromatography of
the crude product on silica gel with AcOEt/hexane (1:4–1:2–1:1)
gave 5 (unlabelled) (608 mg, 99%); 1H NMR (CDCl3) d: 0.99 (t, 3H,
J = 7.3 Hz, 7-H3), 1.03 (d, 3H, J = 6.8 Hz, 2-CH3), 1.15 (s, 3H, 4-CH3),
1.29 (s, 3H, isopropylidene-CH3), 1.37 (s, 3H, isopropylidene-CH3),
1.38 (m, 1H, 6-H), 1.54 (m, 1H, 6-H), 2.03 (m, 1H, 2-H), 3.08 (m, 1H,
1-OH), 3.38 (d, 1H, J = 4.9 Hz, 3-H), 3.39 (s, 3H, 4-OCH3), 3.58 (t,
2H, J = 4.9 Hz, 1-H2), 3.72 (dd, 1H, J = 2.0, 10.5 Hz, 5-H).
(2S,3R,4R,5R)-1-tert-Butyldiphenylsilyloxy-3,5-dihydroxy-
2,4-dimethyl-4-([13C]methoxy)heptanol (12)
To a solution of 8 (1.02 g, 2.1 mmol) in MeOH (5 ml) was added
PPTS (400 mg, 1.6 mmol) at room temperature, and the mixture
was stirred for 27 h. The reaction mixture was diluted with Et2O,
washed with sat. NaHCO3 aq. and brine, dried over anhydrous
MgSO4, and evaporated. Chromatography of the crude product
on silica gel with AcOEt/hexane (5:95–1:9–1:3) gave 12 and
recovered 8, and the recovered 8 was subjected to the same
reaction twice more (overall 823 mg, overall 88%); 1H NMR
(CDCl3) d: 0.02 (d, 3H, J = 7.0 Hz, 2-CH3), 0.11 (s, 9H, SiC(CH3)3),
0.14 (t, 3H, J = 7.3 Hz, 7-H3), 0.53 (m, 2H, 6-H2), 0.61 (s, 3H, 4-CH3),
1.00 (m, 1H, 2-H), 2.01 (d, 1H, J = 6.1 Hz, 5-OH), 2.31 (d, 1H,
J = 6.4 Hz, 3-OH), 2.34 (d, 3H, J = 141 Hz, 4-O13CH3), 2.61 (dd, 1H,
J = 4.9, 9.8 Hz, 1-H), 2.68 (dd, 1H, J = 7.3, 9.8 Hz, 1-H), 2.68 (m, 1H,
5-H), 3.17 (m, 1H, 3-H), 7.65–7.68 (m, 10H, phenyl-H10).
(2R,3R,4R,5R)-3,5-O-Isopropylidene-2,4-dimethyl-4-methox-
yheptanoic acid (9)
To a two-phase solution of 5 (unlabelled) (598 mg, 2.4 mmol) in
CCl4 (5 ml), CH3CN (5 ml) and 0.1 M Na phosphate buffer (7.5 ml)
were added NaIO4 (1.56 g, 7.3 mmol) and ruthenium (IV) oxide
(55%, 19 mg, 63.4 mmol) at room temperature, and the whole
was stirred vigorously for 2 h. The reaction mixture was diluted
with sat. NaCl aq. and extracted with Et2O. The combined extract
was dried over anhydrous MgSO4 and evaporated. Chromato-
graphy of the crude product on silica gel with AcOEt/hexane
1
(1:3–1:1–2:1–4:1) gave 9 (430 mg, 68%); H NMR (CDCl3) d: 0.98
(2S,3R,4R,5R)-3,5-Diacetoxy-1-tert-butyldiphenylsilyloxy-
2,4-dimethyl-4-([13C]methoxy)heptanol (13)
(t, 3H, J = 6.8 Hz, 7-H3), 1.16 (s, 3H, 4-CH3), 1.29 (d, 3H, J = 7.3 Hz,
2-CH3), 1.33 (s, 3H, isopropylidene-CH3), 1.39 (s, 3H, isopropy-
lidene-CH3), 1.39 (m, 1H, 6-H), 1.53 (m, 1H, 6-H), 2.80 (quintet,
1H, J = 7.3 Hz, 2-H), 3.31 (s, 3H, 4-OCH3), 3.60 (dd, 1H, J = 2.9,
8.8 Hz, 5-H), 3.76 (d, 1H, J = 7.3 Hz, 3-H).
Pyridine (2 ml, 24.7 mmol) and acetic anhydride (2 ml,
21.2 mmol) were added to 12 (401 mg, 0.9 mmol) at room
temperature, and the mixture was stirred for 3 h. To this solution
was added DMAP (30 mg, 0.25 mmol) at room temperature, and
the whole was stirred for 23 h. The reaction mixture was
evaporated. Chromatography of the residue on silica gel with
AcOEt/hexane (5:95–1:9) gave 13 (446 mg, 94%); 1H NMR (CDCl3)
d: 0.89 (d, 3H, J = 7.0 Hz, 2-CH3), 0.90 (t, 3H, J = 7.6 Hz, 7-H3), 1.05
(s, 9H, SiC(CH3)3), 1.15 (s, 3H, 4-CH3), 1.56 (m, 1H, 6-H), 1.72 (m,
1H, 6-H), 2.02 (s, 3H, COCH3), 2.05 (s, 3H, COCH3), 2.27 (m, 1H, 2-
H), 3.36 (d, 3H, J = 142 Hz, 4-O13CH3), 3.40 (dd, 1H, J = 6.4, 9.8 Hz,
1-H), 3.48 (dd, 1H, J = 8.5, 9.8 Hz, 1-H), 5.17 (d, 1H, J = 2.1 Hz, 3-H),
5.23 (dd, 1H, J = 2.1, 10.4 Hz, 5-H), 7.65–7.68 (m, 10H, phenyl-H10).
S-2-Acetylaminoethyl (2R,3R,4R,5R)-3,5-O-Isopropylidene-
2,4-dimethyl-4-methoxyheptanethioate (10)
To a solution of 9 (171mg, 0.65 mmol) and 2-acetylaminoetha-
nethiol (386mg, 3.2mmol) in DMF (0.4 ml) was added diphenyl-
phosphoryl azide (0.28 ml, 1.3mmol) at room temperature under
an argon atmosphere. To this solution was added dropwise Et3N
(0.36 ml, 2.6 mmol) at 01C, and the whole was stirred for 16 h at
room temperature. The reaction was quenched with sat. NH4Cl
aq. and extracted with Et2O. The combined extract was washed
with 10% NaOH aq. and brine, dried over anhydrous MgSO4, and
evaporated. Chromatography of the crude product on silica gel
with AcOEt/hexane (2:1–4:1)–CHCl3/MeOH (20:1) gave 10
(224 mg, 95%); 1H NMR (CDCl3) d: 0.98 (t, 3H, J = 7.1 Hz, 7-H3),
1.06 (s, 3H, 4-CH3), 1.26 (d, 3H, J = 7.1Hz, 2-CH3), 1.31 (s, 3H,
isopropylidene-CH3), 1.36 (s, 3H, isopropylidene-CH3), 1.38 (m,
1H, 6-H), 1.49 (m, 1H, 6-H), 1.95 (s, 3H, NHCOCH3), 3.02 (dt, 2H,
J = 2.9, 6.6Hz, SCH2), 3.06 (dq, 1H, J = 1.2, 8.1Hz, 2-H), 3.32 (s, 3H,
4-OCH3), 3.45 (m, 2H, SCH2CH2), 3.65 (dd, 1H, J = 2.0, 8.8 Hz, 5-H),
3.80 (d, 1H, J = 8.3 Hz, 3-H), 5.93 (brs, 1H, NH).
(2S,3R,4R,5R)-3,5-Diacetoxy-2,4-dimethyl-4-([13C]methoxy)-
heptanol (14)
To a solution of 13 (104 mg, 0.20 mmol) in THF (0.5 ml) were
added dropwise acetic acid (30 ml, 0.52 mmol) and n-Bu4NF
(1.0 M in THF, 0.4 ml, 0.4 mmol) at room temperature, and the
mixture was stirred for 89 h. The reaction was quenched with
sat. NH4Cl aq., and the whole was extracted with Et2O. The
combined extract was washed with sat. NaHCO3 aq. and brine,
dried over anhydrous MgSO4, and evaporated. Chromatography
of the crude product on silica gel with AcOEt/hexane (1:4–1:2)
gave 14 (54 mg, 78%); 1H NMR (CDCl3) d: 0.91 (t, 3H, J = 7.3 Hz, 7-
H3), 0.93 (d, 3H, J = 7.0 Hz, 2-CH3), 1.20 (s, 3H, 4-CH3), 1.58 (m, 1H,
6-H), 1.72 (m, 1H, 6-H), 2.03 (s, 3H, COCH3), 2.13 (s, 3H, COCH3),
2.21 (m, 1H, 2-H), 2.94 (dd, 1H, J = 3.1, 5.5 Hz, 1-OH), 3.18 (ddd,
1H, J = 3.1, 5.8, 9.5 Hz, 1-H), 3.38 (d, 3H, J = 142 Hz, 4-O13CH3),
3.44 (ddd, 1H, J = 5.5, 8.6, 9.5 Hz, 1-H), 5.01 (d, 1H, J = 2.1 Hz, 3-H),
5.26 (dd, 1H, J = 1.8, 10.4 Hz, 5-H).
(2R,3R,4S,5R)-2,4-Dimethyl-3-hydroxy-4-methoxyheptan-5-
olide (11)
A solution of 10 (50 mg, 0.14 mmol) in 48% HF/CH3CN (5:95,
1 ml) was stirred for 1 h at room temperature, then added
dropwise to a suspension of NaHCO3 in CH2Cl2, and the whole
was stirred for 30 min at room temperature. To this suspension
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J. Label Compd. Radiopharm 2008, 51 213–217

K. Iida et al.
(2R,3R,4R,5R)-3,5-Diacetoxy-2,4-dimethyl-4-([13C]methoxy)-
heptanoic acid (15)
Conclusion
Reaction of the optically active erythromycin A synthetic
intermediate 6 obtained in our previous work with iodo[13C]-
methane did not afford the desired 3, as d-lactone 11 was
generated during deprotection of the 3,5-dihydroxyl groups in
the final step. Therefore, the 3,5-dihydroxyl groups of 3 were
protected by diacetylation, and 4 was successfully obtained for
biosynthetic studies on erythromycin.
To a two-phase solution of 14 (711 mg, 2.1 mmol) in CCl4 (5 ml),
CH3CN (5 ml), and 0.1 M Na phosphate buffer (7.5 ml) were
added NaIO4 (1.33 g, 6.2 mmol) and ruthenium (IV) oxide (55%,
8.0 mg, 26.7 mmol) at room temperature, and the whole was
stirred vigorously for 1 h. The reaction mixture was diluted with
sat. NaCl aq. and extracted with Et2O. The combined extract was
dried over anhydrous MgSO4 and evaporated. Chromatography
of the crude product on silica gel with AcOEt/hexane
1
(1:2–2:1–4:1) gave 15 (615 mg, 81%); H NMR (CDCl3) d: 0.90 (t,
3H, J = 7.3 Hz, 7-H3), 1.14 (d, 3H, J = 7.3 Hz, 2-CH3), 1.18 (s, 3H, 4-
CH3), 1.52 (m, 1H, 6-H), 1.74 (m, 1H, 6-H), 2.05 (s, 3H, COCH3),
2.08 (s, 3H, COCH3), 3.06 (quintet, 1H, J = 7.3 Hz, 2-H), 3.28 (d, 3H,
J = 142 Hz, 4-O13CH3), 5.23 (dd, 1H, J = 2.7, 10.4 Hz, 5-H), 5.34 (d,
1H, J = 7.3 Hz, 3-H).
References
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To a solution of 15 (278 mg, 0.77 mmol) and 2-acetylaminoetha-
nethiol (826mg, 6.9mmol) in DMF (0.4 ml) was added diphenyl-
phosphoryl azide (0.5 ml, 2.3 mmol) at room temperature under
an argon atmosphere. To this solution was added dropwise Et3N
(0.65 ml, 4.7 mmol) at 01C, and the whole was stirred for 14 h at
room temperature. The reaction was quenched with sat. NH4Cl
aq. and the mixture was extracted with Et2O. The combined
extract was washed with 10% NaOH aq. and brine, dried over
anhydrous MgSO4, and evaporated. Chromatography of the
crude product on silica gel with AcOEt/hexane (2:1–4:1) gave 4
(238 mg, 75%); 1H NMR (CDCl3) d: 0.91 (t, 3H, J = 7.6Hz, 7-H3), 1.19
(d, 3H, J = 7.3Hz, 2-CH3), 1.21 (s, 3H, 4-CH3),1.52 (m, 1H, 6-H), 1.74
(m, 1H, 6-H), 1.96 (s, 3H, NHCOCH3), 2.05 (s, 3H, COCH3), 2.09 (s,
3H, COCH3), 3.05 (m, 2H, SCH2), 3.17 (m, 1H, 2-H), 3.30 (d, 3H,
J = 142 Hz, 4-O13CH3), 3.47 (m, 2H, SCH2CH2), 5.23 (dd, 1H, J = 2.1,
10.4 Hz, 5-H), 5.34 (d, 1H, J = 5.5Hz, 3-H), 6.14 (brs, 1H, NH); 13C
NMR (CDCl3) d: 51.78 (4-O13CH3); IR (CHCl3) cmꢂ1: 3450, 3401,
3008, 2940, 1733, 1668, 1526, 1460, 1373, 1248, 1085, 1056, 965;
FAB-MS (glycerol) m/z: 407 (MH1).
J. Label Compd. Radiopharm 2008, 51 213–217
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