A scalable synthesis of a hydroxamic acid LpxC inhibitor

Article abstract of DOI:10.1021/op300163n

A short and scalable synthesis of chiral hydroxamic acid 1, a LpxC inhibitor, is described. This work discloses a novel diastereoselective addition of 2-nitropropane-generated lithium salt to tert-butanesulfinimine. Moreover, the impurity profiles of reac

Full text of DOI:10.1021/op300163n

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Communication
A scalable synthesis of a hydroxamic acid LpxC inhibitor
ZhongBo Fei, Weiyong Kong, Huaimin Wang, Jianbiao Peng,
Feng Sun, Yueyan Yin, Joginder Bajwa, and xinglong jiang
Org. Process Res. Dev., Just Accepted Manuscript • DOI: 10.1021/op300163n • Publication Date (Web): 24 Jul 2012
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A scalable synthesis of a hydroxamic acid LpxC
inhibitor
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ZhongBo Fei*†, Weiyong Kong†, Huaimin Wang†, Jianbiao Peng†,
Feng Sun†, Yueyan Yin†, Joginder Bajwa‡, Xinglong Jiang‡
†Chemical and Analytical Development, Suzhou Novartis Pharma Technology Co. Ltd,
Changshu, Jiangsu, China 215537
‡ Chemical and Analytical Development, Novartis Pharmaceuticals Corporation, East Hanover,
NJ 07936, USA
KEYWORDS
Diastereoselective addition, reduction, impurities and byproducts
ABSTRACT
A short and scalable synthesis of chiral hydroxamic acid 1, a LpxC inhibitor, is described. This
work discloses a novel diastereoselective addition of 2ꢀnitropropane generated lithium salt to
tertꢀbutanesulfinimine. Moreover, impurity profile of reactions giving oily products and practical
means of purifying these products are discussed in detail.
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INTRODUCTION
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The increasing number of multidrugꢀresistant bacterial pathogens calls for urgent needs for
developing new molecules treating bacterial infectious diseases with novel mechanism.1 The
enzyme LpxC (UDPꢀ3ꢀOꢀ[(R)ꢀ3ꢀhydroxymyristoyl]ꢀGlcNAc deacetylase) catalyzes the
biosynthesis of lipopolysaccharides contained in gramꢀnegative bacterial membranes and has
been identified as pharmaceutical target.2, 3 Hydroxamic acids are an important class of LpxC
inhibitors.4 Here we would like to report a scalable synthesis of a new LpxC inhibitor,
hydroxamic acid derivative 1.
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The original synthesis from medicinal chemistry is shown in Scheme 1. Racemic βꢀnitroꢀ
valine 4 was prepared in a twoꢀstep sequence and coupled with benzoic acid derivative 7 to
afford 8. Following a reduction to 9 and protection to 10, the desired enantiomer 11 was yielded
by chiral preparative separation. Removal of the protecting group and installation of
hydroxylamine afforded desired 1.
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Scheme 1. Original synthesis
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Me
NO2
Me
NO2
H
Me
Me
Me
NO2
KOH
H2O
SOCl2
MeOH
OH
+
O
NH3
+
OH
OMe
H2N
*
Me
H2N
O
*
O
O
2
3
4
HATU,
10
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12
13
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O
DIPEA
DCM
O
Br
O
OH
OMe
Me
KOH
MeOH/THF/H2O
OMe
O
O
NaH, DMF
Me
HO
Me
7
6
5
Me
NH2
Me
NHBoc
Me
NO2
Me
(Boc)2O
NaHCO3
Me
O
Me
O
O
OMe
OMe
Zn, HOAc
EtOH/H2O
N
OMe
*
N
*
N
H
*
H
THF, DCM
Me
O
O
H
O
O
O
O
Me
Me
Me
9
10
8
Me
Me
NH2
O
Me
O
NHBoc
OMe
O
Chiral preparative
HPLC separation
H
N
1, TFA, DCM,rt
N
OH
H
N
H
*
2, MeOH, NH2OH
then HPLC
O
O
O
Me
Me
1
11
This synthesis involving a lateꢀstage chiral separation is not economical for multikilogram
manufacture. An asymmetric version would be more efficient for this purpose. We were
successful in achieving an asymmetric synthesis of 1 and developing the new synthesis to a
practical process for scale up.
The asymmetric synthesis is outlined in Scheme 2. Condensation of ethyl glyoxylate 2 with
(S)ꢀ(ꢀ)ꢀtertꢀbutanesulfinamide 12, followed by diastereoselective addition with 2ꢀnitropropane
lithium salt and cleavage of chiral auxiliary produced the desired enantiomer 15. Coupling amine
15 with acid 7 gave 16, which was reduced and then treated with hydroxylamine to generate 1.
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Scheme 2. Asymmetric synthesis
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O
Me
NO2
S+
Me
H
O
Me
NO2
Me
LiHMDS, THF
NH2
O
H
S+
OEt
S+
OEt
OEt
O
N
N
12
10
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12
13
14
15
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H
O
O
O
MgSO4, toluene
14
2
13
de
96%
O
Me
OH
Me
NO2
Me
NO2
O
Me
O
OEt
N
H
AcCl
7
Me
OEt
H2N
O
O
EtOH
DIPEA,
HATU, CH3CN
O
Me
15
~20% yield over three
ee
16
94% yield
steps >99%
Me
Me
Me
NH2
OEt
Me
NH2
O
O
H
N
Zn
NH2OH
Me
50% aq.
N
N
OH
H
H
O
O
H2O, HOAc, EtOH
O
O
Me
17
75% yield
1
60% isolated yield
ee
>99%
RESULTS AND DISCUSSION
The synthesis began with the known condensation of ethyl glyoxylate 2 with (S)ꢀ(ꢀ)ꢀtertꢀ
butanesulfinamide 125 (Scheme 3). The reported condensation conditions utilize activated
molecular sieves in CH2Cl2 at room temperature, and the reaction takes up to 42 hrs. We found
that these conditions could be replaced with MgSO4 in toluene at elevated temperature (56 °C),
which resulted in 20 h reaction time. After concentration to a ca 85% solution in toluene, the
crude Nꢀsulfinylimino ester 13 was used for the next step without further purification.
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Scheme 3. Synthesis of amine 15
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O
S+
Me
NO2
Me
H
NH2
O
H
S+
OEt
OEt
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12
O
N
O
MgSO4. toluene
0.4 equiv. LiHMDS, THF
o
O
13
2
C
-20
Me
NO2
Me
NO2
Me
Me
OEt
Me
Me
H2N
AcCl
O
O
S+
OEt
S+
+
OEt
EtOH
N
H
N
H
O
O
O
15
~20% yield over three
ee
14
96% de
18
steps >99%
Davis et al. have reported high diastereoselective additions of Grignard reagents to
sulfinylimino ester 135 to access αꢀamino acids. Inspired by this work, we decided to explore the
addition of 2ꢀnitropropane generated anion to sulfinylimino ester 13 to access the desired
synthetic intermediate 14. Even though it could be a straightforward way to access chiral vicinal
diamines in general, such type of additions has not been reported to the best of our knowledge.
To our delight, we achieved such addition with high diastereoselectivity (98:2) by using 0.4
equivalents of LiHMDS and 1.2 equivalents of 2ꢀnitropropane at ꢀ20 °C in THF. The full
conversion of the reaction by using substoichiometric amount of base supports the assumption
that it might be an interesting case of baseꢀcatalyzed addition reaction,6 namely the newly formed
adduct in the event served as additional base to deprotonate the rest of 2ꢀnitropropane for
reaction completion. The stereochemistry of 14 was assigned by comparison with authentic
sample on chiral HPLC, and is consistent with the prediction based on the openꢀtransition state
model depicted by Davis.7 The reaction also proceeded with formation of elimination byproduct
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18 (at 8% by HPLC area) and other unidentified impurities, which was estimated to be ca 40%
yielding and was purified after removal of chiral auxiliary. Our preliminary studies indicated that
either higher amount of LiHMDS or higher reaction temperature led to more elimination to 18,
while lower amount of base or lower reaction temperature resulted in incomplete reaction. Using
potassium tertꢀbutoxide as base also triggered the addition but gave lower selectivity. Using
triethylamine as base led to decomposition of 13.
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Crude 14 was treated with hydrochloric acid in ethanol (prepared by addition of AcCl to
ethanol) to provide 15. Free amine 15 itself is oil and cannot be purified by extractive workup.
We found that concentration of reaction solution followed by addition of TBME leading to direct
precipitation of 15 hydrochloric salt with some ammonium chloride was an efficient way to
remove all organic impurities. Further removal of ammonium chloride by washing product in
IPAc solution with aqueous alkaline solution resulted in 15 in 20% yield over three steps, 97%
chemical purity and 99% ee by HPLC. Even though the overall isolation yield is modest (due to
the addition step), the synthesis is operationally simple to access enantiomeric pure βꢀnitroꢀvaline
15, and hence was adopted for pilotꢀplant manufacture.
Synthesis of acid 7 was achieved in a straightforward manner as shown Scheme 4. Following
the propargylation of phenolic ester 5 with 1ꢀbromoꢀbutꢀ2ꢀyne in acetone, the reaction mixture
containing intermediate 6 was filtered to remove inorganic salts and further treated with aqueous
KOH solution for hydrolysis to 7. Upon acidification of the reaction solution with aqueous HCl,
the benzoic acid derivative 7 was precipitated out of the reaction and isolated in 91% overall
yield and 98% purity by HPLC.
Scheme 4. Synthesis of acid 7
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O
O
O
Br
OH
OMe
KOH
OMe Me
O
O
HO
K2CO3, acetone
acetone, H2O Me
91% overall yield
Me
7
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Amine 15 was then coupled with acid 7 using HATU in the presence of DIPEA to afford 16 and
HOAt, tetramethylurea byproducts (Scheme 5). Compound 16 is oil and its easy purification only
relies on extractive workup. We found HOAt and tetramethylurea was removable by aqueous
washing while the intermediate 19 was not, so it was critical to use slightly excess of amine 15 to
ensure its full consumption. All byproducts and impurities including HOAt, tetramethylurea,
DIPEA and excess of amine 15 were removed by diluting the reaction mixture with IPAc and
sequentially washing with aqueous hydrochloric acid solution (to remove DIPEA and excess
amine 15), aqueous sodium carbonate solution (to remove HOAt), and water. The product 16
was eventually isolated in 94% yield and over 95% purity by HPLC.
Scheme 5. Coupling of amine 15 and acid 7
Me
NO2
Me
NO2
O
Me
Me
O
DIPEA, HATU
OH
+
OEt
OEt
H2N
CH3CN
94%
N
H
O
O
O
Me
O
7
15
16
Me
+
N
N
O
O
N
N
N
O
N
Me
Me
N
N
N
N
O
Me Me
HO
Me
(HOAt)
19
Chemoꢀselective reduction of nitro group over acetylenic function of 16 was achieved with
Zinc dust (<10 ꢁm) in the presence of acetic acid. This is a known exothermic reaction. Our
initial attempt by slow addition of acidic acid to the reaction mixture did not lead to control of
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reaction heat. So that an inverse addition of 16 solution in ethanol to the reaction mixture of Zinc
and acetic acid in ethanol was explored and proved to be efficient at controlling reaction
temperature below 5 °C. It is noteworthy that uncontrolled reaction temperature resulted in
further reduction of acetylenic function of product 17 8. The reduction was always accompanied
by the formation of acid 20.
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Scheme 6. Reduction of 16
Me
Me
NH2
Me
NO2
Me
NH2
Me
Me
O
O
O
Zn
OEt
OH
+
OEt
N
N
N
H
H
O
H
O
O
O
H2O, HOAc, EtOH
75%
O
O
Me
Me
Me
17
20
16
Even though we cannot completely rule out the pathway of hydrolysis of 17 to form 20, it is
more likely formed via reductive cleavage of the cyclized intermediate 22 which was identified
in the reaction mixture by LCꢀMS (Scheme 7). We observed that 16 was quickly reduced to 21
while its further reduction to 17 was slow, which opened up the competing pathway of
cyclization to 22. Therefore higher equivalents of Zinc would increase the reaction rate of
reducing 21 to 17 and disfavor its cyclization to 22 leading to 20. Indeed, we observed that ten
equivalents of Zinc led to the 17/20 ratio of 94/6, while five equivalents led to a ratio of 77/23
under otherwise identical conditions.
Scheme 7. Reduction pathways
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Me
NH2
Me
NO2
Me
N
H
Me
Me
Me
O
Zn
OH
Zn
O
O
OEt
OEt
OEt
N
N
H
N
H
H
O
O
O
O
O
O
7
8
16
Me
Me
Me
17
21
9
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Me
NH2
Me
O
Me
H
Me
Zn
OH
N
O
N
O
H
O
N
O
H
O
Me
O
20
Me
22
The acid impurity 20, acetic acid and Zinc acetate were in any case removed by washing the
crude product in IPAc solution with mixed aqueous alkali and EDTA tetra sodium salt solution.
Subsequent acidꢀbase wash eventually resulted in product 17 in 75% yield and 97% purity by
HPLC. This treatment lowered Zinc content to less than 10 ppm.
Treatment of compound 17 with aqueous hydroxylamine in methanol gave a clean but slow
reaction. It took 60 hrs for completion. Introducing two equivalents of LiOH to the reaction
dramatically increased the reaction rate to 3 hrs for completion. The crude product was purified
by a sequence of salt formation and releasing to the free form to give 1 in 84% isolated yield,
>98% purity and >99% ee by HPLC.
Scheme 8. Completion of the synthesis
Me
Me
NH2
Me
NH2
O
Me
H
N
O
50% aq.
NH2OH
N
OH
OEt
N
H
O
H
O
O
LiOH, methanol
84%
O
Me
Me
1
17
CONCLUSION
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In summary, a short and scalable synthesis for producing hydroxamic acid derivative 1 is
described. Diastereoselective addition of 2ꢀnitropropane derived lithium salt to a tertꢀ
butanesulfinimine is the key reaction in this approach and this in principle could serve as a
general method for preparing chiral vicinal diamines which are quite prevalent in many
molecules of biologically interest. Further detailed studies of this reaction and extension of the
reaction scope are under way in our laboratory.
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EXPERIMENTAL SECTION
NMR spectra were recorded on a Bruker 400 MHz spectrometer. HPLC analysis was
performed using Agilent 1200, and the results were described as area %. LCꢀMS analysis was
performed using Agilent 1200 and 6130 Quadrupole system.
(E)-(S)-2-Methyl-propane-2-sulfinylimino]-acetic acid ethyl ester (13). Under nitrogen
atmosphere (S)ꢀ(ꢀ)ꢀtertꢀbutanesulfinamide 12 (1.45 kg, 11.96 mol) was added to a solution of
ethyl glyoxylate 2 in toluene (2.40 L, 51% wt in toluene, 12.06 mol) in a 20 L flask fitted with a
mechanically stirrer and a condenser. After addition, the mixture was heated to 50 °C and a clear
solution was obtained. MgSO4 (2.88 kg, 23.93 mol) was added to the above solution and the
temperature was increased to 56 °C (internal temperature). The reaction mixture was stirred at 56
°C and monitored by HPLC. After 20 h almost all sulfinamide was consumed (< 1% left). The
reaction mixture was cooled to room temperature, filtered and washed with toluene. The
collected filtrate was concentrated to yield yellow oil (2.67 kg, 85% wt in toluene, measured by
HꢀNMR). The crude Nꢀsulfinylimino ester 13 (2.27 kg, 92%) was used as such for the next step.
1H NMR (CDCl3, 400 MHz): δ 8.01 (s, 1 H), 4.38 (q, 2 H, J = 7.2 Hz), 1.39 (t, 3 H, J = 7.2 Hz),
1.28 (s, 9H).
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(S)-3-Methyl-2-((S)-2-methyl-propane-2-sulfinylamino)-3-nitro-butyric acid ethyl ester
(14). 2ꢀNitropropane (485.5 mL, 5.40 mol) was slowly added to a mechanically stirred and cold
LiHMDS solution (ꢀ20 °C) (1.82 L, 1M in THF, 1.82 mol). During the addition, internal
temperature was kept below ꢀ15 °C. Stirring was continued for 30 min after the addition, and a
solution of Nꢀsulfinylimino ester 13 (1.10 kg, 85% wt in toluene, 4.54 mol) in THF (4.5 L) was
slowly added to the above solution in 80 min and the internal temperature was maintained at
about ꢀ20 °C. After the addition, stirring was continued for 60 min, and saturated aqueous
NH4Cl solution (3 L) was added to the cold reaction mixture followed by the addition of water (1
L). 10 min later, EtOAc (1.5 L) was added to the above mixture and the layers were separated.
The organic layer was washed with 10% brine (2 Lx1). The collected organic layer was
concentrated to yield the crude product containing 14/diastereomer of 14/18 (~98/2/7, 1.10 kg in
total), which was used as such in the next step.
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(S)-2-Amino-3-methyl-3-nitro-butyric acid ethyl ester (15). Acetyl chloride (1.2 L, 16.82
mol) was slowly added to a mechanically stirred and cooled (0 °C) 10 L flask containing
anhydrous ethanol ( 4.2 L). During the addition, the internal temperature was kept below 25 °C.
After the addition, the mixture was stirred for 30 min at 25 °C.
The crude sulfonamide 14 (1.40 kg) in EtOAc (1.4 L) was poured into the prepared HCl
solution. Stirring was continued for 60 min and the solvent was concentrated. MTBE (4.9 L)
was added to the residue and the solid was filtered after vigorous stirring. The filter cake was
washed with MTBE and dried under vacuum to yield the hydrochloride salt of 15 as a white solid
(370 g, >99. 0 % ee, ~20% yield over three steps) containing NH4Cl impurity. 1H NMR (CDCl3,
400 MHz): δ 4.89 (s, 1 H), 4.30 (m, 2H), 1.95 (s, 3H), 1.88 (s, 3H), 1.29 (t, 3 H, J = 7.2 Hz). 13C
NMR (DMSO, 100 MHz): δ 166.0, 87.5, 62.7, 57.5, 24.4, 22.4, 13.7. ESIꢀMS: m/z = 191 [M +
1]+ .
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4-But-2-ynyloxy-benzoic acid (7). To a reactor was sequentially charged 5 (227.2 g, 1.49
mol), K2CO3 (247.1 g, 1.79 mol), acetone (1.15 L) and 1ꢀbromoꢀbutꢀ2ꢀyne (198.6 g, 1.49 mol) at
RT under nitrogen atmosphere. The reaction mixture was heated to reflux for 6 h. After
completion the reaction mixture was cooled to 25 °C. The inorganic salt in the reaction mixture
was filtered and washed with acetone (300 mL).
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To the above filtrate was added a solution of KOH (133.7 g, 2.38 mol) in water (1.4 L). The
mixture was stirred at 40 °C. After completion (1 h), 1.5 L of aqueous HCl (6 N) was slowly
added while the internal temperature was kept below 25 °C. The white solid formed was filtered
and washed with water (to pH=7) and acetone, dried under vacuum to give 7 (258 g, 91% over 2
1
steps). H NMR (DMSO, 400 MHz): δ 7.90 (d, 2 H, J = 8.8 Hz), 7.05 (d, 2 H, J = 8.8 Hz), 4.83
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(d, 2 H, J = 2.0 Hz), 1.89 (s, 3 H). C NMR (DMSO, 100 MHz): δ 166.9, 161.0, 131.2, 123.4,
114.6, 84.0, 74.3, 56.1, 3.1. ESIꢀMS: m/z = 189 [M ꢀ 1]+ .
(S)-2-(4-But-2-ynyloxy-benzoylamino)-3-methyl-3-nitro-butyric acid ethyl ester (16). A
suspension of 15 hydrochloric salt (100 g) in IPAc (2.5 L) was washed with NaOH solution (1
Lx1, 1 M), followed by water (1 Lx1). Then IPAc layer was concentrated to give 15 as light
yellow oil (61.3 g, yield 73%). The oil was dissolved in acetonitrile (760 mL), and 7 (50.8 g,
0.268 mmol), HATU (101.7 g, 0.268 mol) were added then DIPEA (69.1 g, 0.535 mol) at ~ 22
°C. After completion at about 19 h (intermediate 19 was completely consumed), IPAc was added.
The solution was sequentially washed with aqueous HCl (1 M, 0.8 Lx2), water (1.3 L), aqueous
Na2CO3 solution (11% wt, 370 mL) and water (1.3 Lx2). The organic phase was concentrated to
give 16 as an oil (91.3 g, >99% ee, yield 94%). 1H NMR (DMSO, 400 MHz): δ 8.79 (d, 1 H, J =
9.2 Hz), 7.88 (d, 2 H, J = 8.8 Hz), 7.06 (d, 2 H, J = 8.8 Hz), 5.59 (d, 1 H, J = 9.2 Hz), 4.83 (d, 2
H, J = 2.4 Hz), 4.12 (m, 2 H), 1.82 (t, 3 H, J = 2.4 Hz), 1.66 (s, 3 H), 1.56 (s, 3 H), 1.15 (t, 3 H,
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J = 7.2 Hz). 13C NMR (DMSO, 100 MHz): δ 168.3, 166.7, 160.1, 129.7, 125.9, 114.3, 88.7, 83.9,
74.3, 61.4, 56.9, 56.0, 25.0, 21.1, 13.8, 3.1. ESIꢀMS: m/z = 363 [M + 1]+ .
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(S)-2-(4-But-2-ynyloxy-benzoylamino)-3-methyl-3-amino-butyric acid ethyl ester (17). A
stirred mixture of Zn dust (<10 ꢁm) (<10 (17.9 g, 276 mmol) in EtOH (50 mL) and H2O (3.3
mL) was cooled to ꢀ10 °C in 250 mL flask under nitrogen atmosphere, followed by addition of
HOAc (31.4 mL, 552 mmol) in one portion. The resulting mixture was cooled to ꢀ10 °C, and a
solution of 16 (9.3 g, 25.7 mmol) in EtOH (50 mL) was slowly added while internal temperature
was maintained below 5 °C (the addition rate was controlled so as to maintain an internal
temperature below 5 °C). After addition, the mixture was cooled to ꢀ10 °C (internal temp.) and
stirred at this temperature until 16 was fully consumed. The reaction was then warmed to 0 °C
and stirred for 10 h until full conversion of intermediate 21. The reaction mixture was filtered
and concentrated to remove EtOH. To the concentrated mixture was added IPAc (203 mL). The
resulting mixture was cooled to ~22 °C, stirred for 10 min and filtered. The filtrate was cooled to
0 °C with stirring and combined aq. EDTA tetra sodium salt and NaOH solution (0.25 M for
EDTA tetra sodium salt and 4 M for NaOH, 148 mL) was added at such a rate that internal
temperature was controlled below 22 °C (the pH value was checked to be 10~11, if not more
solution will be added to reach this pH value). The separated IPAc layer was washed with
combined EDTA tetra sodium salt and NaOH solution (0.25 M for EDTA tetra sodium salt and 1
M for NaOH, 102 mL x 2), followed by water (62 mLx1). The IPAc layer was concentrated to 20
mL, and extracted with 1M HCl aqueous solution (30.5 mLx1). To the aqueous phase was added
IPAc (204 mL). The mixture was stirred and cooled to 0 °C a. To the mixture was added 1 M
NaOH solution (39.6 mL) at such a rate that internal temperature was maintained below room
temperature. Two phases was separated, the organic phase was washed with water (62 mL) and
concentrated (internal temp. 40 °C, 50 mbar) to give 17 as an oil (6.4 g, yield: 75 %, HPLC
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purity by area normalization: 97.4 %). 1H NMR (DMSO, 400 MHz): δ 8.09 (d, 1 H, J = 8.0 Hz),
7.84 (d, 2 H, J = 8.8 Hz), 7.03 (d, 2 H, J = 8.8 Hz), 4.85 (d, 2 H, J = 2.0 Hz), 4.32 (d, 1 H, J = 7.6
Hz), 4.12 (m, 2 H), 1.82 (t, 3 H, J = 2.0 Hz), 1.19 (t, 3 H, J = 7.2 Hz), 1.13 (s, 3H), 1.10 (s, 3 H).
13C NMR (DMSO, 100 MHz): δ 171.0, 166.3, 159.8, 129.3, 126.6, 114.3, 83.8, 74.3, 62.0, 60.1,
56.0, 51.4, 28.2, 27.9, 14.1, 3.1. ESIꢀMS: m/z = 333 [M + 1]+ .
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N-((s)-2-Amino-1-hydroxycarbamoyl-2-methyl-propyl)-4-but-2-ynyloxy-benzamide (1). A
3ꢀnecked 250ꢀmL round bottom flask was charged with 17 (10.0 g, 30.12 mmol), and methanol
(100 mL). The solution was cooled to 0ꢀ5 °C and 50% aqueous hydroxylamine (22.13 mL, 361.4
mmol) was added followed by LiOH.H2O (2.5 g, 60.24 mmol). The reaction mixture was stirred
at 0ꢀ5 °C. HPLC of an aliquot of the reaction mixture after 3 h at 0ꢀ5 °C indicated completion of
the reaction. Water (100 mL) was added to the reaction mixture at 0ꢀ5 °C. The pH of the mixture
was adjusted to ~8.5 with 6 N HCl. The mixture was concentrated under vacuum. To the residue
were added water (50 mL) and 6 N HCl until pH of the solution was ~6. The solution was
transferred to a 1ꢀL, 3ꢀnecked round bottom flask. Aqueous NaOH (4 N) was slowly added till
pH of the mixture was 8.5. The product precipitated out as oil. The mixture was heated at 45 °C
for 0.5 h. The mixture was slowly cooled to ~22 °C and stirred at ~22 °C for 1 h. The offꢀwhite
solid was filtered and dried under vacuum to give 1 (8.05 g, yield 84%, HPLC purity by area
normalization 98.7%). 1H NMR (DMSO, 400 MHz): δ 7.82 (d, 2 H, J = 8.4 Hz), 7.01 (d, 2 H, J
= 8.4 Hz), 4.80 (d, 2 H, J = 2.4 Hz), 4.26 (d, 1 H, J = 8 Hz), 1.82 (t, 3 H, J = 2.4 Hz), 1.08 (s, 3
H), 0.99 (s, 3 H); 13C NMR (DMSO, 100 MHz): δ 167.2, 165.7, 159.7, 129.2, 126.8, 114.2, 83.8,
74.4, 58.0, 55.9, 51.8, 28.5, 27.2, 3.1. ESIꢀMS: m/z = 320 [M + 1]+ .
AUTHOR INFORMATION
Corresponding Author
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*zhongbo.fei@novartis.com
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ACKNOWLEDGMENT
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We thank Dr. Tangqing Li and Mr. Yunwei Zhu for valuable discussions, Ms. Na Li, Ms.
Lijuan He and Mr. Su Xu for analytic supports, and Dr. Li Zeng and Dr. Anjun Hu for safety
tests. We also thank Dr. Kapa Prasad and Dr. Fabrice Gallou for help with manuscript
preparation.
ABBREVIATIONS
LiHMDS, Lithium bis(trimethylsilyl)amide; THF, Tetrahydrofuran; TBME, tertꢀbutyl methyl
ether;
IPAc,
isopropyl
acetate;
HATU,
Oꢀ(7ꢀAzabenzotriazolꢀ1ꢀyl)ꢀN,N,N′,N′ꢀ
tetramethyluronium hexafluorophosphate; DIPEA, N,NꢀDiisopropylethylamine; HOAt, 1ꢀ
Hydroxyꢀ7ꢀazabenzotriazole; EDTA, Ethylenediaminetetraacetic acid.
REFERENCES
(1) Li, X.; Uchiyama, T.; Raetz, C. R. H.; Hindsgaul, O. Org. Lett. 2003, 5, 539.
(2) Jackman, J. E.; Fierke, C. A.; Tumey, L. N.; Pirrung, M.; Uchiyama, T.; Tahir, S. H.;
Hindsgaul, O.; Raetz, C. R. H. J. Biol. Chem. 2000, 275, 11002.
(3) Sutherland, P. J.; Tobin, A. E.; Rutherford, C.; Price, N. P. J. Biol. Chem. 1998, 273, 4459.
(4) Onishi, H. R.; Pelak, B. A.; Gerchens, L.S.; Silver, L. L.; Kahan, F. M.; Chen, M.; Patchett,
A. A.; Galloway, S. M.; Hyland, S. A.; Anderson, M. S.; Raetz, C. R. H. Science 1996, 274, 980.
(5) Davis, F. A.; McCoull, W. J. Org. Chem. 1999, 64, 3396.
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(6) Ernest, G. J. Am. Chem. Soc. 1985, 107, 4710.
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(7) The stereochemistry was confirmed by comparing with authentic sample after transforming
to 9.
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(8) The reduction of acetylene to alkene in 8 by Zinc was confirmed by LCꢀMS.
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