New straightforward synthesis and characterization of a unique 1β- methylcarbapenem antibiotic biapenem bearing a σ-symmetric bicyclotriazoliumthio group as the pendant moiety

Article abstract of DOI:10.1021/jo980462j

Biapenem 1, (1R,5S,6S,)-2-[(6,7-dihydro-5H-pyrazolo[1,2- α][1,2,4]triazolium-6-yl)thio]-6-yl)thio]-6-[(R)-1-hydroxyethyl]-1- methylcarbapen-2-em-3-carboxylate, is a new non-natural 1β-methylcarbapenem antibiotic which exhibits a wide range of antibacterial activity, remarkable chemical stability, and extensive stability against human renal dehydropeptidase-I. Mercaptobicyclotriazolium chloride 2 useful for the pendant moiety of 1 was successfully synthesized starting from hydrazine hydrate 3 along an economically available synthetic route. The thiol 2 was efficiently exploited for an expeditious synthesis of biapenem 1. Characterization (crystal structure, nonbonded S- - -O interaction, conformational analysis, and CH- - -O hydrogen bonds) of 1 was investigated by its X-ray crystallographic, ¹H NMR, and deuteration experiment analyses.

Full text of DOI:10.1021/jo980462j

J . Org. Chem. 1998, 63, 8145-8149
8145
New Str a igh tfor w a r d Syn th esis a n d Ch a r a cter iza tion of a Un iqu e
1â-Meth ylca r ba p en em An tibiotic Bia p en em Bea r in g a σ-Sym m etr ic
Bicyclotr ia zoliu m th io Gr ou p a s th e P en d a n t Moiety
Toshio Kumagai,*,1a Satoshi Tamai,1a Takao Abe,1a Hiroshi Matsunaga,1a
Kazuhiko Hayashi,1a Ikuo Kishi,1a Motoo Shiro,1b and Yoshimitsu Nagao*,1c
The Chemical and Formulation Research Laboratories, Lederle (J apan), Ltd., Kashiwa-cho, Shiki,
Saitama 353-8511, J apan, Rigaku Corporation, 3-9-12 Matsubara-cho, Akishima-shi,
Tokyo 196-8666, J apan, and Faculty of Pharmaceutical Sciences, The University of Tokushima,
Sho-machi, Tokushima 770-8505, J apan
Received March 11, 1998
Biapenem 1, (1R,5S,6S,)-2-[(6,7-dihydro-5H-pyrazolo[1,2-a][1,2,4]triazolium-6-yl)thio]-6-[(R)-1-hy-
droxyethyl]-1-methylcarbapen-2-em-3-carboxylate, is a new non-natural 1â-methylcarbapenem
antibiotic which exhibits a wide range of antibacterial activity, remarkable chemical stability, and
extensive stability against human renal dehydropeptidase-I. Mercaptobicyclotriazolium chloride
2 useful for the pendant moiety of 1 was successfully synthesized starting from hydrazine hydrate
3 along an economically available synthetic route. The thiol 2 was efficiently exploited for an
expeditious synthesis of biapenem 1. Characterization (crystal structure, nonbonded S- - -O
interaction, conformational analysis, and CH- - -O hydrogen bonds) of 1 was investigated by its
1
X-ray crystallographic, H NMR, and deuteration experiment analyses.
In tr od u ction
of strong antibacterial activity, remarkable chemical
stability, and extensive stability against human renal
DHP-I.4c,d In the earlier synthesis of 1, we adopted a
roundabout synthetic route by which the desired bicy-
clotriazolium moiety was constructed after introduction
of 4-mercaptopyrazolidine bis-N-p-nitrobenzyloxycarbo-
nyl derivative6 onto the 1â-methylcarbapenem skeleton
14.3b We now describe an alternative straightforward
synthesis of biapenem 17 and its characterization based
Since the development of a remarkable non-natural 1â-
methylcarbapenem antibiotic by a Merck Sharp & Dohme
research group,2 there have been several reports on new
characteristic 1â-methylcarbapenems, such as Mero-
penem,3a biapenem,3b lenapenem,3c and others.3b,d,4a,b
These 1â-methylcarbapenems are promising as new-
generation non-natural â-lactam antibiotics because of
their excellent biological and chemical behavior.3,4 We
have developed a unique biapenem 13b bearing a σ-sym-
metric (6,7-dihydro-5H-pyrazolo[1,2-a][1,2,4]triazolium-
6-yl)thio (“bicyclotriazolium”thio) group at C2 as the
pendant moiety that is strikingly different from that of
other 1â-methylcarbapenems.3a,c It is generally known
that the C2-pendant moiety of the carbapenem antibiotics
plays an important role in their antibacterial activity,
chemical stability, and stability against renal dehydro-
peptidase-I (DHP-I).5 Biapenem 1 exhibits a wide range
1
on X-ray crystallographic and H NMR analyses and a
deuteration experiment.
(1) (a) Lederle (J apan). (b) Rigaku Corporation. (c) The University
of Tokushima.
(2) Shih, D. H.; Baker, F.; Cama, L.; Christensen, B. G. Heterocycles
1984, 21, 29.
Resu lts a n d Discu ssion
In designing the pendant molecule, a new heterocyclic
compound, mercaptobicyclotriazolium chloride 2, was
chosen on the basis of the following consideration.
Namely, we wished to establish a reaction cascade (vide
infla) for construction of this particular heterocycle by
treatment of a pyrazolidine derivative with ethyl formim-
idate hydrochloride. Thus, we attempted to define an
economically viable synthesis of 2 starting from hydra-
(3) (a) Sunagawa, M.; Matsumura, H.; Inoue, Y.; Fukasawa, M.;
Kato, M. J . Antibiotics 1990, 43, 519. (b) Nagao, Y.; Nagase, Y.;
Kumagai, T.; Matsunaga, H.; Abe, T.; Shimada, O.; Hayashi, T.; Inoue,
Y. J . Org. Chem. 1992, 57, 4243. (c) Ohtake, N.; Okamoto, O.; Mitomo,
R.; Kato, Y.; Yamamoto, K.; Haga, Y.; Fukatsu, Y.; Nakagawa, S. J .
Antibiot. 1997, 50, 598. (d) Wildonger, K. J .; Ratcliffe, R. W. J . Antibiot.
1993, 46, 1866.
(4) (a) Guthikonda, R. N.; Cama, L. D.; Quesada, M.; Woods, M. F.;
Salzmann, T. N.; Christensen, B. G. J . Med. Chem. 1987, 30, 871. (b)
Kim, C. U.; Luh, B. Y.; Misco, P. F.; Hichicock, M. J . J . Med. Chem.
1989, 32, 601. (c) Ubukata, K.; Hikida, M.; Yoshida, M.; Nishiki, K.;
Furukawa, Y.; Tashiro, K.; Konno, M.; Mitsuhashi, S. Antimicrob.
Agents Chemother. 1990, 994. (d) Hikida, M.; Kawashima, K.; Nishiki,
K.; Furukawa, Y.; Nishizawa, K.; Saito, I.; Kuwao, S. Antimicrob.
Agents Chemother. 1992, 481.
(5) Andrus, A.; Baker, F.; Bouffard, F. A.; Cama, L. D.; Christensen,
B. G.; Guthikonda, R. N.; Heck, J . V.; J ohnston, D. B. R.; Leanza, W.
J .; Ratcliffe, R. W.; Salzmann, T. N.; Schmitt, S. M.; Shih, D. H.; Shah,
N. V.; Wildonger, K. J .; Wilkening, R. R. In Recent Advances in the
Chemistry of â-Lactam Antibiotics; Roberts, S. M., Brown, A. G., Eds.;
The Royal Society of Chemistry; London, 1984; pp 86-99.
(6) Kumagai, T.; Tamai, S.; Abe, T.; Nagase, Y.; Inoue, Y.; Nagao,
Y. Heterocycles 1994, 37, 1521.
(7) (a) Kumagai, T.; Matsunaga, H.; Machida, R.; Nagase, Y.; Hikida,
M.; Nagao Y. J apan Kokai Tokkyo Koho (J apanese Patent), 1989, J P
S64-25779. (b) Abe, T.; Tamai, S.; Nagase, Y. J apan Kokai Tokkyo
Koho (J apanese Patent), 1992, J P H4-230267. (c) Abe, T.; Tamai, S.;
Nagase, Y. J apan Kokai Tokkyo Koho (J apanese Patent), 1992, J P H4-
230286. A Merck research group3d also achieved an expeditious
synthesis of biapenem 1 utilizing our cyclization procedure3b,7 for the
bicyclotriazolium derivative.
10.1021/jo980462j CCC: $15.00 © 1998 American Chemical Society
Published on Web 10/16/1998

8146 J . Org. Chem., Vol. 63, No. 23, 1998
Kumagai et al.
Sch em e 1a
F igu r e 1. Computer-generated drawing of 1 derived from the
X-ray coordinates.
bicyclotriazolium derivative 13, which has been previ-
ously reported by our group.3b,7 Namely, the compound
12 was allowed to react with ethyl formimidate hydro-
chloride3b,7 in the presence of KHCO3 in water at 0 °C,
and the crude product was purified on a Dowex 50W-X4
column with 50% MeOH and 6 N HCl-MeOH (1:1) to
furnish a crystalline compound bicyclotriazolium disul-
fide dichloride 13 in 75% yield. An aqueous solution of
13 was treated with a solution of Bu3P in THF at 0 °C,
and the reaction mixture was repeatedly washed with
AcOEt. After evaporation of the resultant water layer
in vacuo, the crude product was crystallized in isopropyl
alcohol to give mercaptobicyclotriazolium chloride 2 as
the hydrate compound (C5H8N3SCl‚H2O) in 70% yield.
Subsequently, the mercaptobicyclotriazolium moiety
was introduced to the 1â-methylcarbapenem skeleton in
a Michael type reaction manner, as shown in Scheme 2.3b
The compound 14, obtained by our earlier synthetic
method,3b was treated with 2 in the presence of diiso-
propylethylamine in a solution of MeCN-acetone-DMF
(1:1:0.1) at 0 °C to afford thioether 15 as colorless needles
in 91% yield. Finally, deprotection of the p-nitrobenzyl
group (PNB) of 15 was efficiently performed by our
method8 employing Zn dust as follows. Treatment of 15
with excess Zn dust in 0.35 M phosphate buffer solution
(pH 5.6) at room temperature followed by the usual
workup3b,8 of the reaction mixture gave the desired
biapenem3b in 80% yield.
Sch em e 2a
zine hydrate 3 and an expeditious synthesis of biapenem
1, as shown in Schemes 1 and 2. Treatment of 3 with
HCO2Et and then acetone gave crystalline compound 4
in 96% yield. Allylation of 4 with allyl bromide employ-
ing K2CO3 in AcOEt at 80 °C followed by deprotection of
ketimine 5 with excess HCO2H at 80 °C afforded bis-
formylated allylhydrazine 6 in 68% yield from 4. Bro-
mination of 6 with Br2 in the presence of catalytic LiBr
in CH2Cl2-MeOH at 0 °C followed by cyclization of the
resultant dibromide 7 with K2CO3 in warm AcOEt
furnished cyclic diformyl product 8 as colorless prisms
in 70% yield from 6. Substitution reaction of 8 with
potassium thioacetate in warm AcOEt gave crystalline
acetylthiolate 9 (95% yield) which was submitted to
methanolysis in MeOH containing KOH followed by
acidification with a small excess of HCO2H to obtain thiol
10 in 98% yield. Air oxidation of 10 with catalytic FeCl3
in MeOH followed by deformylation of the resultant
disulfide 11 with concd HCl in MeOH gave pyrazolidine
disulfide dihydrochloride 12 as colorless prisms in 81%
yield from 10. Then the pyrazolidine derivative 12 was
subjected to the reaction cascade toward the desired
Successful recrystallization of 1 from aqueous EtOH
furnished excellent crystals [colorless needles; mp 210-
218 °C (decomp); [R]20 -33.7° (c 0.5, water)] that were
D
then submitted to X-ray analysis. The crystallographic
structure of 1 is depicted in Figure 1. The conformation
of C6-hydroxyethyl group of 1 is different from that of
imipenem9a and meropenem.9b An intramolecular non-
bonded O12- - -S16 interaction (“close contact”) is recog-
nized in the crystalline structure of 1.10 This O12- - -S16
close contact must be common even in other â-lactam
antibiotics.9a,b,11,12 To learn the conformation of C2 and
C6 side chain groups of 1 in D2O, a nuclear Overhauser
effect (NOE) experiment [1H NMR analysis (400 MHz)]
(8) Kumagai, T.; Abe, T.; Fujimoto, Y.; Hayashi, T.; Inoue, Y.; Nagao,
Y. Heterocycles 1993, 36, 1729.
(9) (a) Ratcliffe, R. W.; Wildonger, K. J .; Michele, L. D.; Douglas, A,
W.; Hajdu, R.; Goegelman, R. T.; Springer, J . P.; Hishfield, J . J . Org.
Chem. 1989, 54, 653. (b) Yanagi, K.; Takeuchi, Y.; Sunagawa, M. Acta
Crystallogr. 1992, C48, 1737. (c) Takeuchi, Y.; Inoue, T.; Sunagawa,
M. J . Antibiot. 1993, 46, 827.

Unique 1â-Methylcarbapenem Antibiotic Biapenem
J . Org. Chem., Vol. 63, No. 23, 1998 8147
Ta ble 1. Deu ter a tion Exp er im en ts of th e
6,7-Dih yd r o-5H-p yr a zolo[1,2-a ][1,2,4]tr ia zoliu m Moiety
F igu r e 2. Selected NOE enhancement for 1.
conditions
percent
temp, deuteration
compound base (mmol) solvent
time
°C
(%)a
1
1
1
Et3N (0.005) D2O
2-5 min rt
100
0
50
none
D2O
D2O
24 h
12 h
rt
50
none
13
13
13
13
15
15
15
Et3N (0.1)
none
none
CD3OD 2-5 min rt
100
50
100
95
67
100
100
D2O
D2O
D2O
24 h
144 h
12 h
rt
rt
50
none
Et3N (0.005) CD3OD 2-5 min rt
Et3N (0.005) CD3OD 20 min rt
Et3N (0.1) CD3OD 2-5 min rt
a
1
Each percent deuteration was determined by H NMR analy-
sis.
3. The angles (<A- - -H-D: 123.6-149.5°) are also
sufficiently large for the hydrogen bonding described
above.
F igu r e 3. Possible intermolecular hydrogen bonds in the
crystal structure of 1.
In order to get an insight into the acidity of both the
C20 and C22 hydrogen atoms of the bicyclotriazolium
moiety, deuteration experiments (200 MHz 1H NMR
analysis) on compounds 1, 13, and 15 were carried out.
The results of these experiments are listed in Table 1.
When these compounds were exposed to CD3OD or D2O
containing 1 mol equiv or 5 mol % of Et3N at room
temperature for 2-5 or 20 min, 100% deuteration of both
C20 and C22 hydrogens was recognized. Surprisingly,
the similar deuteration proceeded even without Et3N at
room temperature or 50 °C. The easy deuteration of the
C20 and C22 hydrogens can be explained by the contri-
bution of ylide 16 to the mechanism of the deuteration
reaction.
was performed. Selected NOE enhancements are il-
lustrated in Figure 2. Interestingly, an NOE enhance-
ment between H1 and H8 was confirmed on the spectrum
of meropenem by Sumitomo Pharm. Co. research group9c
but was not recognized on that of biapenem 1. On the
other hand, an NOE enhancement between H6 and H8
was observed on the spectrum of 1 but was not on that
of meropenem.9c Thus, the predominant conformation of
C6-hydroxyethyl and C2-bicyclotriazoliumthio groups in
the molecule 1 in D2O may be similar to that in the
crystalline structure (Figure 1). Particularly, the con-
formational stability of the bicyclotriazoliumthio group
at C2 may be caused by the intramolecular nonbonded
S- - -O interaction due to n (O12) σ* (S16-C17) orbital
overlap.10
Interestingly, four kinds of possible intermolecular
hydrogen bonds (O13- - -H-O15, O12- - -H-C20, O14- - -
H-C22, and O13- - -H-C22) were suggested in the
crystal structure of 1. The interatomic distances (A- - -
H: ca. 2.03-2.28 Å) between the specific acceptor atom
(A: O12, O13, or O14) and hydrogen atom (H) of the
specific donor (D: O15, C20, or C22) are unusually
shorter than the sum (2.72 Å) of van der Waals radii
between oxygen and hydrogen atoms, as shown in Figure
In conclusion, we have established useful, practical,
and large-scale synthetic procedures for the preparation
of mercaptobicyclotriazolium chloride 2 and biapenem 1
and clarified some interesting physical and chemical
characteristics of 1.
Exp er im en ta l Section
(10) The nonbonded O12- - -S16 atoms distance (2.976 Å) is signifi-
cantly shorter than the sum (3.35 Å) of the van der Waals radii of
oxygen and sulfur atoms. In addition, a linear relationship between
the O12 atom and the S16-C17 σ bond and a plane conformation of
the O12-C11-C3-C2-S16 moiety can support the O12- - -S16 close
contact. See, Kucsman, A.; Kapovits, I. Organic Sulfur Chemistry:
Theoretical and Experimental Advances; Bernardi, F., Csizmadia, I.
G., Mangini, A., Eds.; Elsevier Scientific Publishing Co.: Amsterdam,
1985; pp 191-245.
Gen er a l Meth od s. All reactions were monitored by thin-
layer chromatography. Melting points are uncorrected. The
column chromatographic separations were performed on Wako
gel C-200 (particle size 74-149 µm). Optical rotations and
IR and H NMR (270 MHz) spectra were recorded at Medical
Research Laboratories, Lederle (Japan), Ltd. Elemental analy-
ses and H NMR (200 and 400 MHz) and all mass (MS, HRMS,
FAB, and SIMS) spectra were recorded at Faculty of Phar-
maceutical Sciences, The University of Tokushima.
1-F or m yl-2-isop r op ylid en eh yd r a zin e (4). To a solution
of hydrazine hydrate (377 g, 7.54 mol) in EtOH (760 mL) was
added dropwise ethyl formate (726 mL, 9 mol) at -5 °C over
1
1
(11) Nagao, Y.; Abe, T.; Shimizu, H.; Kumagai, T.; Inoue, Y. J . Chem.
Soc., Chem. Commun. 1989, 821.
(12) The intramolecular nonbonded S- - -O interactions recognized
in several drugs have been reported by Y. Nagao et al. Nagao, Y.;
Hirata, T.; Goto, S.; Sano, S.; Kakei, A.; Iizuka, K.; Shiro, M. J . Am.
Chem. Soc. 1998, 120, 3104.

8148 J . Org. Chem., Vol. 63, No. 23, 1998
Kumagai et al.
1 h. The mixture was stirred at -5 °C for 30 min and at room
temperature for 14 h. The reaction mixture was poured into
acetone (1011 mL, 15 mol) over 30 min and then stirred at
room temperature for 30 min. The resultant solution was
evaporated to dryness in vacuo to give 4 (721 g, 96% yield) as
colorless needles: mp 68-69 °C (EtOH); IR (KBr) νmax 3200,
at 0 °C, and then the mixture was stirred at 0 °C for 10 min
followed by addition of formic acid (12 mL, 0.26 mol). After
filtration of the mixture, the filtrate was evaporated in vacuo.
The residue was chromatographed on a silica gel column with
EtOAc to give 10 (32.3 g, 98%) as a colorless oil: IR (CHCl3)
ν
max 1665 cm-1; 1H NMR (CDCl3, 270 MHz) δ 1.85 (d, 1H, J )
1
1690, 1640 cm-1; H NMR (CDCl3, 270 MHz) δ 1.92 (s, 3H),
5.6 Hz), 2.82-2.91 (m, 1H), 3.19-3.29 (m, 2H), 3.69-3.79 (m,
1H), 3.98 (dd, 1H, J ) 7.9 and 12.2 Hz), 8.44 (s, 1H); HRMS
calcd for C4H8N2OS MW 132.0357, found m/z 132.0345 (M+).
Bis(1-for m ylp yr a zolid in -4-yl) Disu lfid e (11). To a solu-
tion of thiol 10 (33 g, 0.25 mol) in MeOH (330 mL) was added
a solution of FeCl3 (405 mg, 2.5 mmol) in MeOH (40 mL) at 0
°C, and air was bubbled into the solution at room temperature
for 2 h. After evaporation of the reaction mixture in vacuo,
the residue was chromatographed on a silica gel column with
CH2Cl2-EtOH (9:1) to give 11 (31 g, 95%) as a colorless oil:
IR (CHCl3) νmax 1665 cm-1; 1H NMR (CDCl3, 270 MHz) δ 3.10-
4.00 (m, 10H), 8.42 (s, 2H),; HRMS calcd for C8H14N4O2S2 MW
262.0558, found m/z 262.0531 (M+).
2.00 (s, 3H), 8.65 (d, 1H, J ) 9.9 Hz), 9.75 (d, 1H, J ) 9.9 Hz);
HRMS calcd for C4H8N2O MW 100.0637, found m/z 100.0637
(M+).
1-Allyl-1-for m yl-2-isop r op ylid en eh yd r a zin e (5). To a
solution of 4 (254 g, 2.54 mol) in EtOAc (762 mL) were added
allyl bromide (330 mL, 3.81 mol) and powdered K2CO3 (877 g,
6.35 mol). The mixture was stirred at 80 °C for 5 h. After
cooling to room temperature, the reaction mixture was filtered,
and the filtrate was concentrated in vacuo. The oily residue
was purified by vacuum distillation to give 5 (266.7 g, 75%
yield) as a colorless oil. Compound 5 was found to be a mixture
of conformational isomers (3:1) due to the N-CHO bond
rotation: bp 60 °C/4 mmHg; IR (CHCl3) νmax 1660 cm-1
;
1H
Bis(4-p yr a zolid in yl) Disu lfid e Dih yd r och lor id e (12). A
solution of disulfide 11 (31 g, 0.119 mol) in concd HCl (40 mL,
0.48 mol) and MeOH (400 mL) was stirred at room tempera-
ture for 6 h to give a precipitate. The desired precipitate was
filtered off, and the filtrate was evaporated to dryness in vacuo.
The solid residue was suspended with a small amount of
MeOH, and then the precipitate was filtered off. The combined
precipitate was dried in vacuo to give 12 (28 g, 85%) as
colorless prisms: mp 194 °C (aq MeOH); IR (KBr) νmax 3200,
NMR (CDCl3, 270 MHz) δ 1.74 (s, 0.75H), 1.86 (s, 2.25H), 1.99
(s, 0.75H), 2.13 (s, 2.25H), 4.09 (d, 1.5H, J ) 6.3 Hz), 4.23 (d,
0.5H, J ) 6.3 Hz), 5.17-5.37 (m, 2H), 5.66-5.91 (m, 1H), 7.93
(s, 0.25H), 7.97 (s, 0.75H); HRMS calcd for C7H12N2O MW
140.0950, found m/z 140.0974 (M+).
1-Allyl-1,2-d ifor m ylh yd r a zin e(6). A solution of ketimine
5 (210 g, 1.5 mol) in formic acid (420 mL) was stirred at 80 °C
for 15 h followed by evaporation of the solvent in vacuo. The
residue was chromatographed on a silica gel column with
EtOAc to give 6 (173 g, 90%) as a colorless oil: IR (CHCl3)
1
3000-2500 cm-1; H NMR (D2O, 270 MHz) δ 3.39 (dd, 4H, J
) 3.8 and 13.0 Hz), 3.59 (dd, 4H, J ) 6.8 and 13.0 Hz), 3.94-
4.02 (m, 2H); FAB MS m/z 207 [(M - 2Cl - H)+]; FAB HRMS
calcd for C6H17N4S2Cl2 MW - 2Cl - H 207.0738, found m/z
207.0716 [(M - 2Cl - H)+].
ν
max 3400, 1720, 1685 cm-1; 1H NMR (CDCl3, 270 MHz) δ 4.13-
4.18 (m, 2H), 5.23-5.38 (m, 2H), 5.66-5.90 (m, 1H), 8.01-
8.32 (m, 3H); HRMS calcd for C5H8N2O2 MW 128.0586, found
m/z 128.0598 (M+).
Bis(6,7-d ih yd r o-5H-p yr a zolo[1,2-a ][1,2,4]tr ia zoliu m -6-
yl) Disu lfid e Dich lor id e (13). To a solution of 12 (28 g, 0.1
mol) and KHCO3 (20 g, 0.2 mol) in H2O (700 mL) was added
ethyl formimidate hydrochloride (109 g, 1 mol) at 0 °C, and
the mixture was stirred at 0 °C for 10 min. After adjusting to
pH 2 with 6 N HCl, the acidic reaction mixture was evaporated
to dryness in vacuo. The solid residue was suspended with
MeOH (420 mL) and then the undesired precipitate was
filtered off. The filtrate was evaporated in vacuo to give an
oily residue. The residue was chromatographed on a DOWEX-
X4 (H+ type) resin column with MeOH-H2O (1:1) and then 6
N HCl-MeOH (1:1) to give 13 (26.5 g, 75%) as colorless
prisms: mp 183 °C (decomp) (MeOH); 1H NMR (D2O, 270
MHz) δ 4.70-4.85 (m, 6H), 4.85-5.00 (m, 4H), 8.90 (s, 4H);
FAB MS m/z 317 [(M - Cl)+]; FAB HRMS calcd for C10H14N6S2-
Cl2 MW - Cl 317.0410, found m/z 317.0433 [(M - Cl)+].
6,7-Dih yd r o-6-m e r ca p t o-5H -p yr a zolo[1,2-a ][1,2,4]-
tr ia zoliu m Ch lor id e (2). To a solution of 13 (17.7 g, 0.05
mol) in THF (90 mL) and H2O (90 mL) was added n-Bu3P (20.2
g, 0.1 mol) at 0 °C, and then the mixture was stirred at 0 °C
for 1 h. After removal of THF in vacuo, the resultant aqueous
solution was washed with EtOAc followed by concentration
in vacuo. The residue was chromatographed on a Diaion SP-
207 resin column with H2O to give 2 (12.4 g, 70%) as colorless
prisms: mp 127-128 °C (decomp) (n-PrOH); IR (KBr) νmax
1-(2,3-Dibr om op r op yl)-1,2-d ifor m ylh yd r a zin e (7). To
a solution of 6 (154 g, 1.2 mol) in CH2Cl2 (750 mL) were added
a solution of LiBr‚H2O (124 g, 1.2 mol) in MeOH (250 mL) and
a solution of Br2 (199 g, 1.26 mol) in CH2Cl2 (250 mL) at 0 °C,
and the mixture was stirred at 0 °C for 10 min. To the reaction
mixture were successively added a suspension of NaHCO3 (396
g) in H2O (250 mL) and saturated aqueous Na2SO (250 mL).
After separation of the CH2Cl2 layer, the aqueous layer was
extracted with EtOAc (3 × 500 mL). The CH2Cl2 and EtOAc
extracts were combined and dried over MgSO4 followed by
evaporation of the solvent in vacuo. The residue was chro-
matographed on a silica gel column with EtOAc-hexane, (3:
1) to give 7 (326 g, 95%) as a colorless oil: IR (CHCl3) νmax
1715, 1690 cm-1; 1H NMR (CDCl3, 270 MHz) δ 3.60-4.50 (m,
5H), 8.10-8.30 (m, 2H), 8.77 (s, 1H); HRMS calcd for C5H8N2O2-
Br2 MW 285.8954, found m/z 285.8969 (M+).
4-Br om o-1,2-d ifor m ylp yr a zolid in e (8). To a solution of
dibromide 7 (214 g, 0.75 mol) in dry EtOAc (1070 mL) was
added powdered anhydrous K2CO3 (207 g, 1.5 mol), and then
the mixture was stirred at 40 °C for 6 h. After filtration of
the mixture, the filtrate was evaporated in vacuo. The residue
was chromatographed on a silica gel column with EtOAc-
hexane (3:1) to give 8 (114 g, 74%) as colorless prisms: mp
83-84 °C (EtOAc); IR (CHCl3) νmax 1700 cm-1; 1H NMR (CDCl3,
270 MHz) δ 3.80-4.40 (m, 4H), 4.60-4.80 (m, 1H), 8.50 (s,
2H); Anal. Calcd for C5H7N2O2Br: C, 29.01; H, 3.41; N, 13.53.
Found: C, 28.95; H, 3.43; N, 13.75.
2400 cm-1 1H NMR (D2O, 270 MHz) δ 4.20-4.35 (m, 2H),
;
4.70-4.85 (m, 3H), 8.67 (s, 2H); FAB MS m/z 142 [(M - Cl)+];
FAB HRMS calcd for C5H8N3SCl MW - Cl 142.0439, found
m/z 142.0486 [(M - Cl)+]; Anal. Calcd for C5H8N3SCl‚H2O:
C, 30.69; H, 5.15; N, 21.48. Found: C, 30.51; H, 4.76; N, 21.58.
p-Nitr oben zyl (1R,5S,6S)-2-[(6,7-d ih yd r o-5H-p yr a zolo-
[1,2-a ][1,2,4]tr ia zoliu m -6-yl)th io]-6-[(R)-1-h yd r oxyeth yl]-
1-m eth ylca r ba p en -2-em -3-ca r boxyla te Ch lor id e (15). To
a suspension of p-nitrobenzyl (1R,5R,6S)-2-(diphenylphospho-
ryloxy)-6-[(R)-1-hydroxyethyl]-1-methylcarbapen-2-em-3-car-
boxylate 14 (5.95 g, 10 mmol) and thiol 2 (2.3 g, 13 mmol) in
a mixture of MeCN (18 mL), acetone (18 mL), and DMF (1.8
mL) was added diisopropylethylamine (2.1 g, 16.3 mmol) at 0
°C, and the mixture was stirred at 0 °C for 2 h to afford a
precipitate. The desired precipitate was filtered off and
washed with CH2Cl2 followed by dryness in vacuo to give 15
(4.74 g, 91%) as colorless needles: mp 163-168 °C (decomp)
4-(Acetylth io)-1,2-d ifor m ylp yr a zolid in e (9). A mixture
of bromide 8 (89 g, 0.43 mol) and potassium thioacetate (72 g,
0.63 mol) in EtOAc (450 mL) was stirred at 40 °C for 6 h. After
filtration of the reaction mixture, the filtrate was evaporated
in vacuo. The residue was chromatographed on a silica gel
column with EtOAc-hexane (3:1) to give 9 (83 g, 95%) as
colorless prisms: mp 47-48 °C (EtOH); IR (CHCl3) νmax 1700,
1
1695 cm-1; H NMR (CDCl3, 270 MHz) δ 2.37 (s, 3H), 3.50-
3.60 (m, 2H), 4.00-4.20 (m, 3H), 8.42 (s, 2H); Anal. Calcd for
C7H10N2O3S: C, 41.58; H, 4.98; N, 13.85. Found: C, 41.51;
H, 5.03; N, 14.10.
1-F or m yl-4-m er ca p top yr a zolid in e (10). To a solution of
acetylthiolate 9 (50.5 g, 0.25 mol) in MeOH (180 mL) was
added a solution of KOH (14 g, 0.25 mol) in MeOH (130 mL)

Unique 1â-Methylcarbapenem Antibiotic Biapenem
J . Org. Chem., Vol. 63, No. 23, 1998 8149
(DMF-CH2Cl2); [R]25D +55.4° (c 0.5, H2O); IR (KBr) νmax 1760,
1695 cm-1; 1H NMR (CD3OD, 270 MHz) δ 1.40 (d, 3H, J ) 7.3
Hz), 1.43 (d, 3H, J ) 6.3 Hz), 3.53 (dd, 1H, J ) 3.0 and 6.3
Hz), 3.70 (dq, 1H, J ) 7.3 and 9.6 Hz), 4.26 (quint, 1H, J )
6.3 Hz), 4.48 (dd, 1H, J ) 3.0 and 9.6 Hz), 4.81 (d, 1H, J )
12.2 Hz), 4.82 (d, 1H, J ) 12.2 Hz), 5.18-5.31(m, 3H), 5.46 (d,
2H, J ) 13.9 Hz), 7.78 (d, 2H, J ) 8.9 Hz), 8.30 (d, 2H, J )
8.9 Hz), 9.16 (s, 1H), 9.18 (s, 1H); FAB MS m/z 486 [(M - Cl)+];
FAB HRMS calcd for C22H24N5O6SCl MW - Cl 486.1447, found
m/z 486.1425 [(M - Cl)+]. Anal. Calcd for C22H24N5O6SCl‚3/
2H2O: C, 48.13; H, 4.96; N, 12.76. Found: C, 48.44; H, 5.25;
N, 12.92.
Gen er a l P r oced u r e for Deu ter a tion Exp er im en ts of
th e Com p ou n d s 1, 13, a n d 15. To a solution of each
compound (0.1 mmol) in CD3OD (1 mL) or D2O (1 mL) was
added Et3N (0.005 or 0.1 mmol). The mixture was allowed to
stand for a time indicated in Table 1 at room temperature and
then the D-content of C20 and C22 hydrogens in each sample
was determined by the 200 MHz 1H NMR analysis at room
temperature. The deuteration experiments and the D-content
analysis for 1 and 13 in the absence of Et3N were similarly
carried out at room temperature or 50 °C, as described above.
All experimental results are recorded in Table 1.
(1R,5S,6S)-2-[(6,7-Dih yd r o-5H -p yr a zolo[1,2-a ][1,2,4]-
t r ia zoliu m -6-yl)t h io]-6-[(R)-1-h yd r oxyet h yl]-1-m et h yl-
ca r ba p en -2-em -3-ca r boxyla te (1). To a solution of thioether
15 (1.042 g, 2 mmol) in 0.35 M phosphate buffer solution (pH
5.6) (35 mL) was added zinc dust (8.3 g), and then the mixture
was stirred at room temperature for 1 h. The insoluble
materials were filtered off through a Celite pad, and the filtrate
was adjusted to pH 5.5 with 0.1 N HCl followed by concentra-
tion in vacuo. The oily residue was chromatographed on a
Diaion SP-207 resin column with isopropyl alcohol-H2O (5:
95) to give 1 (0.56 g, 80%) as colorless needles: mp 210-218
°C (decomp) (H2O-EtOH); [R]20D -33.7° (c 0.5, H2O); IR (KBr)
νmax 1750, 1600 cm-1; 1H NMR (D2O, 270 MHz) δ 1.26 (d, 3H,
J ) 7.3 Hz), 1.30 (d, 3H, J ) 6.6 Hz), 3.41 (dq, 1H, J ) 7.3
and 9.6 Hz), 3.53 (dd, 1H, J ) 3.0 and 5.9 Hz), 4.27 (dq, 1H,
J ) 5.9 and 6.6 Hz), 4.31 (dd, 1H, J ) 3.0 and 9.6 Hz), 4.71-
4.80 (m, 2H), 4.98 (m, 1H), 5.04-5.15 (m, 2H), 9.02 (s, 1H),
9.04 (s, 1H); SIMS m/z 351 [(M + 1)+]. Anal. Calcd for
Ack n ow led gm en t. This work was partly supported
by the Grant-in-Aid for Scientific Research on Priority
Area No. 8245101 from the Ministry of Education,
Science, Sport, and Culture, of J apanese Government.
Su p p or tin g In for m a tion Ava ila ble: 1H NMR spectra for
new compounds which were not characterized by combustion
analysis and X-ray characterization data for biapenem 1,
including tables of experimental details, atomic coordinates,
anisotropic displacement parameters, bond lengths, bond
angles, torsion angles, nonbonded contacts out to 3.60 Å,
intermolecular hydrogen bonds, ORTEP drawing, and crystal
structure (36 pages). This material is contained in libraries
on microfiche, immediately follows this article in the microfilm
version of the journal, and can be ordered from the ACS; see
any current masthead page for ordering information.
C
15H18N4O4S: C, 51.42; H, 5.18; N, 15.99. Found: C, 51.09;
H, 5.18; N, 15.96.
J O980462J