3 .1~5~19
-- 2
This is a division of copending a~plication
serial no. 357,816, filed August 7, 1980
~ACE~ROUND O~ ~ INVEI~IO~
Field of the invention
This invention relate~ to new semi-synthetiG amino-
glycosidic antibiotic~9 particularly 6"-deoxydibekacin,
4",6"-dideoxydi~ekacin, 1~ 4-amino-2-hydroxybut~ryl)-
6"-deoxydibekacin and l-N (I.-4-amino-2-hydroxybutyryl)-
4"96"-dideo~ydibekacin which are each the new compound
u~eful as antibacterial agents~ Th~s in~ention also
relates to processes for -the production of these new
compounds, Thi~ invention further relates to an antiW
bacterial composition comprising one of the~e new compou~d~
: as the active ingredient.
Descript~on of the prior art
Dibekacin, namely ~ dideo~ykanamycin ~ wa~ semi-
synthetically produced from kanamycin ~ by the presentinventors (see Japanese patent publication No. 7595/75;
Japanese patent ~o. 794,612; U.S. patent ~o. 3,75~,973~.
Dîbekacin has been used extensively in therapeutic treat-
ment o~ various bacterial in*ections as a chemotherapeutic
agent which ls active against the kanamycin-sen~itire
bacteria and also against various kanam~cin-resi~tant
bac-~eria~ We, the present inventors, produced semi-
syntheticallg l-N~ 4-amino-~-hydro~ybu-tyryl)-dibe~acin
which i8 a chemotherapeutic agent e~fective again~t
dibekacin-resistan~ bacteria (see JapanQ~e patent publi-
cation No. ~3~629/77; U.S~ patent Nou 4,107,424). Recently,
we also produced semi-synthetically 6"-deoxyamikacin
.` ~k
15~7~19
(i,e~ l-N~ 4-amino-2-hydroxybutyryl)-6"-deoæykanamycin A)
and 4",6"-dideoæyamikacin (i.e. 1-N-(L~4-amino 2~hydro~y-
b~tyry~)-4",6" dideoæykanamycin A) (Japane~e patent appli-
cation No. 54733/79) which each have a low oto-toxicit~
and are confirmed to show an antibacterial activity as
hlgh a~ that o~ ~mikacin9 namely 1~ 4-amino-2-hydroæy-
butyryl)-kanamycin A~
~U~MARY 0~ THE INVFNTION
In our ~urther re~earches9 we have now sucGeeded at
first time in synthetioally producing the new 6'~-deoxy or
4",6i'-dideo~y derivative~ o~ dibekacin and 1~ 4-amino-
- 2-hydroxybutyrgl~-dibekacin, respectively. Furthermore,
ue have ~ound that these new 6"~deoæy derivatives and
4"36"~dideoxy derivatives now synthetized e~hibit not only
a low oto-to~icity but al~o ~how an antibacterial activity
as high a~ that of dibekacin or 1 N~ 4-amino-2-hydro~y-
butyryl)-dibekacin~
DETAILBD DESCRIPTION OF THE I~VENTIO~
According to a flr~t aspect of this i~vention,
there-~ore~ there i~ provided a~ a new compo~nd a deoæy
derivative of dibakacin repre~ented by the general
~ormula (I~
~ 1~5~
- 4
CH3
R ~ ~ \ ~ ~ ~L~
~2~ ~ 0 2 (I)
0 ~
H2N ~ ~g2
~herein R i~ a hydro~yl group or a hydrogen atom, a~d an
a~id addition ~alt thereo~. The new compoùnd of the above
formula (I) where R is a h~dro~yl group i~ 6" deoxydibekacin,
and the new compound of the formula (I3 where R is a hydrogen
atom is 4",6"-dideoxydibekacin.
According to a ~econd a~pect of this invention9
there is provided as a new compound a deoxy derivative of
l-N~ 4-amino~2-hydroxybutyryl)-dibekacin repre~ented by
the general formula (II)
~75~9
5 --
H2~ ou ~ OH 7
O ~ ~ ~ (II)
HN ~ ~ NH2
~ CO
CHOE
l~2a~2~H2
wherein R is a hydro~yl group or a hydrogen atom, and an
acid addition salt thereo~. The new compound of ~he above
formula (II) where R is a hydroxyl group is 1~ 4-2mino-
2-hydro:~:ybutyr~ 6"-deoxydibekacin, a~d the new compou~d
o~ the ~ormula (II) where R is a hydrogen atom i~ l-N-
(I~4-~mino-2-hydro~ybutyrylj-4" 9 6"-dideoxydibekacin.
The physico-chemical and biological propertis~ o~
the above-mentioned four new compounds according to the
first and second a~pect~ o~ this in~ention ar~ a~ f`ollows:-
(l) 6"~-Deoxydibekacin ~esqui-carbona-te i~ a ~ub-
~tance in the form of a colorle3~ powder decomposing gradually
near 131~C and ~howing a speci~ic optical rotation ~a]26_ ~lOl~
(c 0.44, w~ter). It~ elemental analy~i~ i3 coincident with
the theoretical values o~ Cl8~37N507 ~/ 2 3
H 7.63%, N 13.25~). In ma~ spectrometr~, it gave a value
7!58~9
6 --
of m/e 435 (M+). This substance give~ a single spot
(positi~e to ninhydrin) at Rf 0.38 in a thin layer chromato-
graphy on silica gel developed with a mixture of butanol-
ethanol-chloroform water (4:5:2:5 by volume) as the develop
ment solvent~
( 2 ) 4 ", 6 "-Dide oxydibe~acin sesqui-carbonate is a
substance in the form of a colorless powder decomposing
gradually ne~r 129~ and showing a specific optical rotation
~]D3 = + 126~ (c 0.5, water). It~ elemental analysis i~
coincident with the theoretical values o~ ~18H~7N~06-3/2 H2C03
(~ 45~69%, H 7.87%, ~ 13.66%). In mass spectrometeryD it
gave a value o~ m/e 420 (M ~ his substance gives a
single spot (positive to ninhydrin) at Rf Ob45 in the
above-mentioned ~ilica gel thin layer chromatography.
(~) 1 N~ 4-amino-2rhydro~ybutyryl)-6"-deoxy-
dibekaci~ sesqui-carbonate is a substance in the form of a
colorless powder decomposing at 132-1~gC and showing a
specific optical rotation ~a]23 = + 73o (c 0.3, water).
Its elemental analysis is coincident with the theoretical
22H42~69 3/2H~c03 (c 44. 97~o, H 7. ~3~ 13 395~)
Thi~ sub~tance gi~es a single spot (positive to ninhydrin)
at Rf` 0.10 in the above-mentioned silica gel thi~ la~er
chromatograph~
(4) l~N-(Lr4 amino-2-hy~ro~yhu~ryl)-4",6"-dideox.vdibekacin
se~qui-carbonate is a sub~tance in the ~orm o~ a colorless
powder ha~ing a decomposition point at 142~147C ~nd showing
a speci~ic optical rotation [a~24 - ~ 84 (c 0.5, water).
~ 175819
-- 7 --
It~ elemental analy~is i~ coincident with the theoretical
22~4~N68 3/2 H2C03 (C 46-15~ H 7.42%, ~ 13.74~).
~his substance give~ a ~ingle spot at Rf 0.21 in the above-
mentioned silica gel thin layer chromatographyO
Th~ minim~un inhibitory concentrationæ (mcg/m~) of
6"-deoxydibekacin (abbre~iated as 6"-DDBB)~ 4",6"-dideo~y-
dibekacin (abbreviated a~ 4'1~6"-DDRB), 1~ 4-amino 2-
hydroxybutyry~)-6"-deo~ydibekacin (abbreviated as AHB-6"
DDEB) and 1 N-~I,4-amino-2-hydro~ybutyryl) 4",61'-dideo~y-
; 10 dibekacin (abbreviated as AHB-4'~,6'1-DDKB) of this invention
against various microorganism~ were determined according to
~erial dilution method on a nutrient agar medium at ~7~C~
the estimation being made after 18 hours incubation. ~or
compari~on purpose, the minimum inhibitory concentrations
~ dibekacin (abbreviated as DKB) were also determined in
the same manner a~ ~tated above.
The antibacterial spectra of these new and known
~u~stances are shown in Table 1 below.
. ,
~ ~ ~S~L9
__ .
=a) o ~D
. _ ~ C~l ~ Lr~~ P~ O ~ O ~ L~ a
_ ~ o r~ JLS~ ~ ~ L(~ In 1
_ ~I r I r~ O
I ~ ~ ~ O O O ~ ~ O ~ ~ O O
~Q ~ V
~4Q
__ ~ . ~ ~
~ _ . .. _
~~ O ~
1-1Ir~ N 1~ 15~ L~ tr~ O O o ~\ ~ 0~ .D CO
~ t-i r~ J ~ ~ t~ U~ L~
~_ _ ~ ~ O O O ~ ~ C~ ~i ~ O O
~3 ~
~~ ~ o ~
e~ iq ~ 0 0 0 ~ D W
~::1~ o - ~
Z. _1 O O ~ ~ O ~ o ~ o o
_ ~ 1~ O O O O ~ O ~~I r-l Or-l
~ U~ ~ O
_ . r~
f~ CO O ~ ~D
~4 ~ c~ o In o~
~ . .. ~ t~ C~J C~J C~J 1~ Ll~ ~n In
r l Q O O~ ~ o
. a ~ N Lf ~ O O O tD L~ O' r~ r-l 0 ~1
r~ _ ~r~r C~ r~
n~ ~-
,a:~ o ~D ~:7 ~ - - ------- _ _ ,, ,__ _
E~r~ o o o
~4 . ~c~l ~ ~m In 11
'. ~ V O ~ O
. ~ ~ O O O O O ~ O ~1 ~i ~ O ~
,~ o~ ~ ~ Lr~ 0~
_ ~ _ . , _ ,,, . ~_
, ~ O
O
r~ .
,~ 0 V 00
D ~ ~ O E~ 0
O E; ~~d ~ cn I O ,~
c~
O H ~ J CUC~
U~ ~D ~ C) V P~ rl ~ Il~--1 r-¦~1
.,1t1 - =rl ~ ; V P El --i
~ ~ O ~0 ~
~5 U) P H ~rl ~1 ~ ~ ~U a)
. ~ tl~ ~ c) ~ 1 .~ E~ rl
h rQ ~ r-l ~ Cd r~
O ~ ~ t~ r lrl :~ ~ O
C) C) ~ rl El c )
O O
' E~ C) ~ q C) -~ f-~ ~r
r-l ' ., r-l O ~ ~q ~ Ul ~ -1~ 0
~ ,0 C) ~r~
,S~ , o
~ h ~rl r~l r-l ~I r-l R ,o a~
a~ v c) ~J t ~rl ~1 ~ O ~
-1~ V V ~ V
.? ~ n~ ~S
rt? . ~ n r~ r~
9 ~ 8 1 9
r-i ~ o ~ O ~ 9
V
-
O ~ 0~ 0 ,~
~i ~ Q O 1~ 0 0 0 0 0 0 S
- - - - - - - - - - - - - - - - - -
tn. ~ ~ O m u~ ~i o ~ o ,i o
O C~l ~ O O O
~- ~- ~- ~ ~ O Lt~ ~L ~ O ~ c, o o
o ~ c~l ~) o o
- - ~ L - - - - -
c~o o ~ ~0 ~ ~-~
cC ~ ¢ ~ ~C) H
æ ,~ v
~I r~ ~3 r- r,l r-J r-l r-l r~ 1 C~
o h ~1
r-l ;~ h
O C,~
,1 ., ~ a~ , I r~
h _ _ _ _ h _ _ _ rl 1~l ~1
~, , ~
-- ~ . .... _ _ .. ~ ._ . . . ~ ._ , _
~lo- 117~81~
O ~D O ~ ~ O ~ ~ O ~ ~ ~D
~ ~ O ~J O r~ L~ N 1::
~, .. . . _ . .. . . ._ . . . ~
~, ~ ~ o o ,~ ~ 8 ~ o ~ ~
. '
O ~D O ~ ~ C~ ~ ~ O O ~ C~J O ~ O
~ J l ~ ~ ~ .-,
- - ~~~ ~~ r - - - ------ -- - - - . . _ _ __ _ _- _ - _ __ _ . .
Lr~ ~
~ o ~ o ~ ~ <:~ o g 8 ~t ~D N ~ ~
r~ ,~, r-~
. . _ _.. _ ~1~ -- --- j~_ , _ _ ,
~D O ~ O ~D r-l O V g g O S~
~,t ~t r I ~ r-
~,,,, ___ , , _
,. 1.
N
r p~
t~ rl cn r-l
~lt ~ ~r ~t
F r
rt ~ rt C~ r-t r-l
rt ~5 J a) rt S-l
O ~ 1 0
Ulr~J ~3 d a~ h ~ ~ rl rl Ctl ~
~13~tt rr I 1> F I CIS ~ Id F~ t ~ i
~1~ c~ rl rl rl ~ (1~ El
,r~ ~ ~ r~ r~ O
a~ o o a) Q) a)c~ r-l ~1 r
t~O~ E 1~ ~ t ~ h 5-~ h ~ p
rl ~ r-- O O
~ t~ ~ h
. . _ _
!
7 5 8 1 9
. . . , ~
LL~ ~ ~
.. ~,
O ~ C~J O L~ O ~ O
o ,~ ~ ~ ~ ~ ~ o
~ ,~
~ U~
~ ~ ~ o C) o ~ o
U~ ~ ~ o ~ o ~1 o
~, ~
, . . . ~
o o o ~ ~ o ~ o
o o o ~ o ~ ~U o
~. ~1 ~,1 ~ ~ ,~
. _ . ... ,
L-~
~ ~U
C~ ~ O~ O O ~ O
O ~ U~ O O O ~
.. __ .
U~ ~
O O ~O O O ~D O
O U~ O O O ~
5~
. ~
~ O
LS~ ~ ~ ~ o
rll ~1 0
Fq N P I FCI M
C) . ,1
R
o~
~q
r~ = - = - - - r
Q) ' Q)
- 12 _ 1~758~9
~ rom the above table, it is ~een that the ne~
compound~ of the general formulae (I) and (II) according
to thi~ invention inhibit the growth o~ many kinds o~
bacterial strains. The new compounds of this in~en-tion
e~hibit a low acute toxicity to animal~ and men. It ha~
bee~ e~timatea that the intra~enous ID50 of 6"-deo~y-
dibekacin and 4?',6"-dideo~ydibekacin in mice were more
than 60 mg~kg, and also that the intravenous ID50 o~
l-N-(I-4-amino-2 hydroxybutyryl)-6"-deoxydibekacin and
l-N (I-4-amino-2~hydro~ybutyryl)-4"p6"-dideo~ydibekacin
in mice were more th~n 80 mg/kg.
As compared to dibekacin (DEB~ 9 6"-deoxydibekacin
and 4"96"-dideoxydibekacin of this ~nvention are 3ame or
more active against ~anamycin-sensitive and re~istant
bacteria. 1-N-(I~4-Amino-2-hydroxybu~yryl)-6"-deoxydibekacin
and l-N (I~4-amino-2-hydroxybutyryl)-4",6"-dideoxydibekacin
of thi~ inventlon show a similar activity to that o~ l-N-
~I-4-amino-~ hydroxybutyryl)-dibekacin. ~iside3, 6"-deoxy-
dibekacin~ 4",6"-dideoxydibekacin, l-N-(~-4-amino-2--hydro~y-
butyryl)-6"-deoxydibekacin and 1-N~ 4-amino-2-hydrox~-
: butyryl)-4",~"-diaeoxydibekacin o~ this invention ha~e a
low oto-toxioity~
T'he new compounds o~ thi~ invention, that is, 6"-
deoxydibekacin, 4",6"-dideo~ydibekacing 1-N-(I-4-amino-2-
hydroxybutyryl)-6"-deoxydibekacin and 1-N~ 4-amino-2-
hydroxybutyryl)-4",6"-dideoxydibekacin are usually obtained
in the ~orm o~ its free base or a hydrate or a carbonate
1~7~9
~ 13
thereof. ~he new compound~ o~ this in~ention each may
readily be converted into a form of a pharmaceutically
acceptable acid addition salt thereof, such as the
hydrochloride, hydrobromide, ~ulYate t pho6phate, nitrate,
acetate~ maleate, citrate~ a~corbate~ methanesulfonate
and the like by reaeting with the appropriatey pharma-
ceutically acceptable inorganic or organic acid in an
aqueous medium.
~he new cempound~ of the formula (I) or (II)
according to thi~ invention and its pharmaceutically
acceptable acid addition ~alt may be admi~istered orally,
intraperitoneally, intravenou~ly/ subcutaneously or
in~ramuscularly using any pharmaceutlcal ~orm known to
the art ~or such administration and in a similar manner
to the known kanamycin~. For insta~ce, the new compounds
of thi~ invention may be admini~tered orally using any
pharmaceutical form kno~n to the art Yor oral administra-
tion. Example~ of the pharmaceutical forms for oral
administration are powdersg capsules, tablets9 syrup and
the like. A suitable dose of the new compounds o~ this
invention for e~ective treatment of bacterial in~ections
is in a ran~e of 0.1 to l g. per person a day when it i~
given orally. It i~ preferred that said do~e should be
orally administered in three to four aliquot~ per day~
The new compounds of this invention may also be admin-
istered by intramuscular injec-tion at a dosage of 50 to
500 mg per person two to four times per day. Moreover,
~ 17~19
-- 14 --
the new compounds of -this invention may b~ formulated
into an ointment for external application which con-tains
the active compound at a concentration o~ 005-5~ by weight
in mixture with a known ointment base such as polyethylene
glycol r ~urthermore, the new compound~ of this invention
are each use~ul for sterilization o~ surgical instruments
and sanitary material3.
According to a thlrd aspect of this invention,
thereforep there i~ provlded an antibacterial compo~ition
comprising as the active ingredient 6"-deoxydibekacin,
4";6"-dideoxydibekacin, l-N~ 4-amino-2-hydro~ybut~ryl)-
6"-deo~ydibekacin or 1 ~ 4 amino-2-hydro~ybutyryl)-4",6"-
dideoxydibekac~n or a pharmaceutically acceptable acid
addition salt thereof, in an antibacterially e~ecti~e
amount to i~hibit the growth of bacteria, in combination
with a carrier for the active ingredient compoundO
Next, the production of the new compounds o~ the
~ormula (I) or (II) according to thi~ invention is de~-
cribed.
6"-Deoxydibekacin and 4"~6"-dideoxydibekacin may
be produced 3emi-synthetically starting from dibekacin~
~ 4-Amlno-2-hydroxybutyryl)-6"-deo~ydibekacin and
l-N~ 4-amino-2-hydro~ybutyryl)-4",6"-dideoxydibekacin
may be produced ~emi-synthetioally 3tarting from 1-~(I-4-
amino-2-hydro~ybutyryl)-dibekacin. While, l-N-(L~4-amino-
2-hydro~ybutyryl)-6"~deoxydibekacin and 1-N~ 4-amino-
2-hydroxybutyryl)-4",6'~-dideoxydibekacin may also be
1 ~ 75~ ~ 9
-- 15 --
produced semi-synthetically using 6"-deo~ydjbekacin and
- 4",6"-dideo~ydibekacin, re~pectively, as the ~tarti.ng
material.
According to a fourth a~pect of thi~ i~vention9
there ie provided a proces~ for the pxoduction of 6"-
deo~ydibekacin or 4",6"-dideoæydibekacin of the ge~eral
formula (I)
c~3
R ~ H2N
E2N ~ OH
O ~ (I)
H2N NH2
wherein ~ i~ a h~dro~l group or a hydrogen atom (R i~ a
hydroxyl group in re~pect of 6"-deoxgdibekacin while R is
a hydrogen atom in re~pect of 4"~6"-dideoxydibekacin)~
which comprise~:-
(a) protecting with a known amino-protecting group
the ~ive amino group~ o~ dibekacin of the formula (III)
.
I 375~9
-- 16 _
HO ~
~lo ~ 2
H ~H2
in a known manner to produce an amino-protected dibekacin
derivative of the ~ormula ~III')
Ht)~\ A
HO ~ \ B~
A ~ ~ ~ B
O ~
\ A (IIIl)
A > ~ ~<
wherein A is a hydrogen atom and B i9 a mono-~alent amino~
protecting group, or A and B taken together ~orm a di-valent
amino~protec-ting grOUp3
(b) protecting with a know~ di-valent hydro~yl-
protecting group simultaneou~ly the 4"- and 6"-hydro~yl
~7581g
- 17 -
group~ of the amino-protected dibekacin derivative (III')
in a kno~n manner to produce a protected derivati~e o~ the
formula (IIIIl)
6"
> N N < (III")
wherein A and ~ have the ~ame meanings as defi~ed above
and a group o~ the ~ormula X \ ~ i8 a di-valen~
y/
hydroxyl~protecting group where X and Y are hydrogen atoms
both (i.e. methylene group), alkyl groups both (alkylidene
group, particularly an isopropylidene group~, each a hydrogen
atom and an alkyl group9particularly an alkyl group of 1~6
oarbon atom~ (alkylidene group), an aryl group, particularly
a phenyl group (benzylideno group~ or an alkoxy group
(alko~y-methylene group), or alternativel~ the group of
the ~ormula X~ ~ i~ a cgcloalkylidene group, particularly
a cyclohex~lidene group~
(c) protecting wi.th a known mono-valent hydroxyl-
protecting group either the 2"-hydro~yl group alone or
-31 1758~9
-- 18 --
bo-th the 5- and 2" hydro~yl group~ of ~aid protected
derivative (III") in a known manner to produce an amino
protected and hydroxyl-protected dibekacin compound of
the formula (IV)
X
< A
>N < B
wherein A~ ~ and the group o~ the ~ormula ~ ~ / ha~e
~/ ~
the ~ame meaning~ as defined above, D i~ a mono valent
hydro~yl prot~cting group in the form o~ an acyl group,
particularly an alkanoyl ~uch a~ acetyl or an ~royl ~uch
a~ ben~oyl, and E i~ a hydrogen atom or a mono-valent
hydroxyl-protecting group in the form of an acyl group
which may be the ~ame a~ or di~ferent ~rom the group D,
(d) removing the group of protecting ~imult~neou~].y
the 4"- and 6"-hydro~yl group~ o~ said protected dibekacin
compound (IV) therefrom in a known manner to produce a
partially protected dibekaoln compound of the ~ormula (V)
I ~758~9
-- 19 --
HO- \
A\ ~ Z<A
A>~ ~ (V)
wherein A, B, D and E have the same meanings as de~:ined
ab OV2 ~
(e) ~ulfonylating with a sulfonylating agent the
6"-hydro~yl group alone or s~multaneously both the ~11,
and 6"-hydroxyl groups of said partially protected di-
bekaoin compound (V) in a known manner to produce a mono-
sulfonylated or di ~ulfonylated derlvative of the
~ormula ~V')
GSO~ \
~OD ~
N < B
8 ~ ~
~ 20 -
wherein A, ~, D and E have the same meanings as defined
above, G is a lower alkyl group of 1~4 carbon atoms9
particularly methgl or ethyl 9 an aryl group such a~ phenyl
group or p-methylphenyl group, or an aralkyl group,
p~rticularly a phenyl-lower alkyl group ~uch a3 benzyl
group, and G' is a hydroxyl group or i~ the same a~ the
group GS03- ~hown in the formula (V')~
(~) reacting the 6"-mono-~ulfonylated or 4"~6"Wdi-
3ulfonylated derivative (V') with an aIkali metal iodide
or bromide in a known manner to replace -the 6"-sulfonyloxy
group or the 4" and 6"-sul~onyloxy groups by iodo or
bromo group, respectively, and thereby to produce the
corresponding 6"-mono-iodo or bromo derivative or the
corre~ponding 4"~6"-di-iodo or bromo derivative,
(g) reducing said 6"-mono-iodo or bromo derivative
or oaid 4",6'l~di-iodo or bromo derivative with hydrogen in
the pre~ence o~ a hydrogenoly~is cataly~t to effect the
dehalogenation and thereby to produce a corresponding 6"-
mono-deoxy derivative (which i~ correspo~ding to such a
compound o~ the ~ormula (V') but where the group GS03-
ha~ been con~erted into a hydrogen atom and G' i~ remaining
aa the hydroxyl group) or the corre~ponding 4"?6l'-dideoxy
derivative (which i~ correspondlng to ~uch a compound of
the formula (V') but where the group GS03- and the group G'
oach have been converted into a hydrogen atom) represented
by the formula (V")
~758~9
- 21
6"
C~
4" ~ O >N
\N ~ ~ ~ f o~<
/ O~ OE
O/~ 0
A~ N ~ < (~"~
wherein A9 ~ D and E are as de~ined above, and R is a
hydroxyl group or a hydrogen a-tom, and
- ~h) remo~ing the remaining ami~o-pr~tecting groups
and the remaining hydroxyl-protecting groups from the
6"-mono-deoxy derivative or 4",6"-di-deoxy deriva~ive (V")
in a known manner to produce 6"-deo~ydibekacin or 4",6"-
dideoxydi~ekacin (I~
The process o~ the abo~e-mentioned ~ourth aspect
of thi~ invention may~ if nece~sary~ include a further
~tep o~ converting 6"-deoxydibekacin or 4"~6"-dideoxy-
dibekacin into a pharmaceutically acceptable acid additio~
3alt thereof.by reacting with a pharmaceutioally acceptable
inorganio or organic acid in a known ma~nerO
The procedures for carrying out the proces3 ~or
the production o~ 6"-deo~ydibekacin or 4",6" dideoxydibekacin
are now describea.
In the ~irst step (a) o~ the present proces~, the
-3 17581g
~ 22 -
five amino group3 o~ the ~tarting dibekacin (III) are
protected with a known amino-protecting group in a known
manner.
~he amino-protecting group available for the pro-
tection of the amino groups of dibekacin may be any known
amino-protecting group which is ordinarily u~ed in the
conventional synthesis o~ peptideæ~ However, the amino-
protecting group employed must be of the nature that it is
removable readily by ~uch a procedure and under ~uch re-
action condition~ which will substantially not break theglyco~ide li~kage o~ dibekacin when the removal o~ the
amino~protecting ~roups is effected ~rom the amino~
protected 6"-deoxydibekacin or 4"~6"~dideo~ydibekacin
as produced in the process of thi~ i~vention. Suitable
e~amples of the mono-valent amino-protecting group which
are available ~a~ the group 3) for the above purpose
include an alko2ycarbonyl group o~ 2~6 carbon atom~ such
a3 tert-butoxycarbonyl and tert-amylo~ycarbonyl; a cyclo-
alk~loxycarbonyl group o~ 3~7 carbon atoms such as cyclo-
he~ylo2ycarbonyl; an aralkyloxycarbonyl group ~uch a3benzylo~ycarbonyl and p-methoxybenzyloxycarbonyl; a~d a
oubotituted al~anoyl group o~ 2~5 carbon atom~ ~uch as
trifluoroacetyl and o-nitrophenoxyacetyl. Pre~erred
e~ample~ o~ the di-valent amino-protecting group available
(as the groupo A and B taken togother) ~or the above-
mentioned purpo~ inelude phthaloyl g~oup and a group of
Schi~ base type such as salicylidene. The introduction
I ~ 75~ ~ 9
~ 23 -
of the amino-protecting group into dibekacin may be
achie~ed by reacting the latter with an appropriate
reagent ~or the introductio~ o~ the amino-protecting
group which is in the ~orm of an acid halide, acid a~ide,
active ester or acid anhydride, in the same manner as
described e.g. in the speci~ications o~ U.S. Patent Nos~
~,929~762 and 3,939~14~ a~ well as ~.E. Patent No. 1,426,90~.
In the ~econd step (b) o~ the present process, the
4"- and 6"-hydroxyl groups of dibek~cin are protected
qimultaneou~ly as a preliminary step for blocking the 2"-
hydroxyl group occa~ionally together with the 5~hydroxyl
group o~ dibekacin.
A~ the di valent hydroxyl-protecting group for
protecting simultaneously both the 4"- and 6"-hydrogyl
groups of dibekacin, there may be employed such a prot~ctive
group which is frequently used ~or blocking the hydro~yl
group~ of 1,3-glycol conventionalLy in the chemistry o~
sugar~. Suitable e~amples o~ the di-valent hydroxy:L-
pro-tecting group ( X ~ ~ ) include an alkylidene group
such a~ methylene, ethylidene and isopropylidene; ~1
aralkylidene group ~uch a~ benzylidene (when the alkylidene
or aralkylide~e group is used ~or this purpose, the 4"-
and 6"~hydroxyl g~oup~ are blocked by being con~erted into
the ~orm o~ an acetal); a cycloalkylidene group such as
cyclohexylidene and tetrahydro-4-pyranylidene (when the
cycloalkylidene group is used for this purpose, the 4"-
~ ~l 7~8 1 9
- 24 -
and 6"-hydroxyl groups are blocked by being con~erted into
the form o~ a ketal); a lower aIkoxy-alkylidene group~
such a3 alkoxy~methylene, particularly methoxy-methylene
(when the alko~y-a~kylidene group is employed for thi~
purpose, the 4-l_ and 6"-hydroxyl groups are blocked by
being con~erted into the form of a cyclic ortho ester)
and the like. In order to introduce the above di-valen~
hydro~yl~protecting group into the 4"-- and 6"-hydroxyl
groups of the N-protected dibekacin derivative ~III'),
the latter is reacted with an appropriate aldehyde or
ketone in a known manner in the presence of an acid
catalyst by exchange reaction with an acetal or a ketal
or a~ orthoformate~ ~he N-protected dibekacin deri~ative
~III') may prefera~ly be reacted with formaldehyde or
2,2-dimethoxypropane for the alkylidenation; with
benæaldeh~de for the aralkylidenation; with l,l-dimethoxy-
cyclohexane for the cycloalkylidenation; or with trimethyl
orthoformate for the alko~y-alkylidenation at a temperatura
o~ e.g. 10~80C in the presence of an acid catalyst such
a~ p-tolene~ul~onic acid or sulfuric acid in a known manner
as de~cribed in U.S. Patent ~99299762, '~he above protection
of the 4"- and 6" hydroxyl gIoups of the N-protected di-
dekacin derivati~e (III') gi~e~ the 4",6't-0-protected
dibekacin derivative (III") containing -the two hydroxyl
groups which remain in the free ~tate at the 5- ~nd 2"-
positions .
In the third step (c) of the present proces~ the
1~75~19
-- 25 ~
2'~-hydroxyl grOUp9 occasionally together with the 5-hydroxyl
group of dibekacin, is protected in a known manner. As the
available hydroxyl-protective group~ (D,E) for blocking
both the 5- and 2"-hydro~yl groups or the 2"~hydro~yl
group alone, there may be mentioned a mono-valent hydro~yl-
protecting group which is an acyl group, particularly a
lower alkanoyl group such a3 acetyl; or an aroyl group
such a~ benzoyl. ~or the introduction of the mono-valent
hydroxyl-protecing group (Dg~), the 4"~6"-O~protected
dibekacin derivati~e (III") is acy1ated by reacting with
an acid anhydride, acid halide or active ester o~ the
acid which is containing the acyl group to be introduced
into both the 5- and 2"-hydroxyl group~ or into the 2"- i
hydroxyl group of the dibe~acin compound. The acylation
may readily be achieved in a known manner. Thi~ acglation
u~ually gives a larger proportion of the 2"-mono-0-acyl
derivative and a minor proportion of the 5,2"~di 0-acyl
derivative owing to that the 5-hydroxyl group is generally
les~ reactive than the 2"-hydroxyl group. The 5,2"-di-0-
acyl derivative a~ well a~ the 2"-mono-0-acyl derivative
o~ the ~ormula (IV) may equally be utilized in the ~ext
step (d) o~ the pres~nt proces~.
In -this way, the third step tc) of the pre~ent
process affords the amino-protected and hyaroxyl-protected
dibekacin compound (IV), and thi~ compound is then ~ub-
~ected to the selective removal of the di-valent hydroxyl-
protecting group ~ ~ from the 4"- and 6"-hydro~yl
, C ~
1~7~8~9
-- 26 --
groups there~rom. The reaction for selec-ti~ely removing
the di-valent hydroxyl-protec-ting group may readily proceed
by subjecting the dibekacin compound (IV) to hydrolysis
under wea~ly acidic conditions in the presence of a weak
acid ~uch a~ aqueoue acetic acid or diluted hyarochloric
acid, giving the partially protected dibekacin compound (V)
~hich is an important intermediate useful in this irl~entio~.
This acid hydroly~is may usuall~ be effected at a t~mperature
of 20C to 100C.
The partially protected dibekarin compound (V) ~o
obtained is containing the liberated 4"- and 6"-hydroxyl
group~. In order to achieve the production of 6"-deogy-
dibekacin or 4",6"-dideogydibekacin according to this
in~ention9 it is necessary to e~ect the removal o~ the
6"-hydro~yl group (~or the 6"-deoxygenation) or the removal
of both the 4"- and 6"~hydroxyl groups from the partially
protected dibekacin compound (V) (for the 4",6"-dideox~-
genation). For thiq mono-deoxygenation or di-deoxygenation9
the following step~ are followed in the process o~ the fourth
a~pect of thi~ in~ention.
Thu~, in the sul~onylation step (e) o~ the proce~s
o~ the fourth aspect invention, the 6"-hydroxyl group or
the 6"- and 4"~hydroxyl group~ oY the partially protected
dibe~acin compound (V) i8 or are alkylsulfonylatedf aryl-
sul~onylatod or aralkylsul~onylated by reacting with asul~onylating agent of tha formula
GS0~-X
1 ~75819
- 27
wherein G is ~he lower alkyl group, the aryl group or the
aralkyl group as de~ined hereinbe~ore and X is chlorine or
bromine atom, in the reaction medium consi~ti~g of eg. dry
pyridine. The sulfonylating agent GS02-X ma~ be an alkyl-
~ulfonylating agent ~uch a~ methanesuleonyl chloride; an
- arylsulfonylating agent suoh as p-toluene~ulfonyl chloride
or an aralkylsulfonylating agent such as benzyl ulfonyl
chloride or bromide. This sulfonylating step (e) gives
the mono-sul~onylated product of which the 6~'-hydroxyl
group alone ha~ been sulfonylated9 and/or the di-s~lfonylated
product of which the 4"- and 6~'-hydroxyl group8 both have
been sulfonylated. When the compound (V) is reacted with
p-toluene~ul~onyl chloride ~in a substantially equimolar
proportion) at ambient temperature~ the 6"-mono-0-tosylated
product (corresponding to the compo~nd of the formula (V~)
where the group GS03- is tosyloxy group p-CE~C6H4S03- and
the group G' is the hydro~yl group) is produced as the main
product in par~icular. On the other hand, ~hen the compound
(V) is reaoted with methanesulfonyl chloride in a ~ub-
~tantlally 2 molar proportions or more at ambient tem-
perature, the 4",6"-di-0-mesylated product (corre~ponding
to the compound o~ the ~ormula (V') where the group GSO~-
and the group G' both are the mesyloxy group CH3S03~
produced in a favorable yield. ~hese 6"-mono~O tosylated
product and 4",6"-di-0--me~ylated product both are the
intermediate useful in this invention. G~nerally, the
~ulfonylation o~ the 4"- and 6'l-hydroxyl groups in the
1~7~9
- 28 -
above step ~8) may be effected at a temperature of 10 -to
100C and most pre-ferably at ambient temperature or at a
temperat~rs o~ 10 to 60C and for a reaction time of
~0 mi~ute~ to 1 day.
The reaction mix-ture from the above sulfonylation
step (e) i3 then admixed with a small volume of water to
decompose the residual amount of the unreacted sulfonylation
agent, ~ollowed by concentration to drynes~ under reduced
pressure to give a crude residue compri~ing the sul~onylatio~
produc-t~ This residue is dissolved in a volume of chloro-
~orm, and the resulting solution is washed with an aqueou~
potassium hydrogen sulfate~ with aqueous saturated ~odium
hydrogen carbonate and with water. The chloroform phase
is then dried with anhydrou~ sodium sulfate and then con
centrated to dryness under reduced pressure to give a crude
powder compri~ing the sul~onylation produc-ts. When thi~
crude powder is subjected to a column chromatography on
~ilica gel developed with a mi~ture of ~hloroform-ethanol,
the 6"-mono-0-sulfonylated product and the 4",6"-di-0-
sul~onylated product may be obtained separately.
In the next step (f) o~ the present process, the
6"-mono-0-sulfonylated product or the 4",6"-di-0-~ulfonylated
product of the formula (V') i~ reacted with an alkali metal
~odide such a~ sodium iodide or an alkali metal bromide
~uch a~ ~odium bromide in an iner-t organic solvent such as
dry dimethylformamide, dimethyl~ulfoxide, acetone 9 dio~ane
and the like. The reaction may properly be e~fected at a
~ 1~581
-- 29 --
reaction temperature o~ 50 to 150C and for a reaction
time o~ 10 minutes to 1 day~ By thi~ reac-tion9 the 6"-
mono-0-sulfonylated product is iodlnated or brominated to
give the 6"-mono-iodo or bromo derivative or the 4",6"-
di-iodo or bromo derivative.
In the further step (g) of the present process,
the 6"-mono~halo derivative or the 4",6"~di-halo derivative
obtained as above is reduced with hydrogen in a known
marmer in the presence o~ a hydrogenolysi~ catalyst such
a~ Raney nickelg palladium, platinum and the like and in
solution in an inert organic solvent such a~ dioxane,
methanol and the lik~ to e~fect the de-iodination or de-
bromination. Thi~ de-iodinatio~ or de-bromination by
hydrogenolysi~ may properly be effected at a temperature
o~ from ambient temperature to 100C and for a reaction
time of ~0 minutes to 24 hours under atmospheric pressure
or even under an ele~ated pressure of 10 kg/cm2. This
dehalogenation step (g) af-~ords the 6"-mono-deoxy derivative
or the 4",6"-dideo~y derivative o~ the formula (V"). In
thi~ way, the 6"-deoxygenation or the 4",6"~dideo~ygenation
has been achieved.
In the last step (h) of the present process, the 6"
deoxy derivative or the 4",6"-di deox~ derivati~e (V")
obtained in the above step ~g) is sub~ected to the treat-
merlt ~or removal o~ the residual protective groups accordingto the corlventio~al deprotecting technique. ~he acyl group
which ls present a~ the mono-valent hydroxyl-protecting
~75~9
30 --
group (D~E) may readily be removed by alkaline hydroly~i3
at ambient temperature, for example~ by treating with 12~o
ammonia-methanol. ~en the amino-protecting group (B) is
an aralkyloxycarbonyl group, thi~ type of the amina-
protecting group is removed concurrently in the catalytic
reduction step (g) o~ the present process, When the amino-
protecting group (B) is o~ the kind other than the aralkyloxy-
carbonyl group, the removal of such amino-protec~ing group
may ea~ily be conducted in a kno~n manner, for example, by
hydrolysis with a weak acid ~uch as a~ueous acetic acid~
In thi~ way, the desired product, that is, 6"-deo~ydibekaci~
(when R is hydro~yl grOllp in the ~ormula (I)~ or 4''v6''-
dideo~ydibekacin (~Jhen R i~ a hydrogen atom in the -~ormula
(I)).
Puri~ication o~ 6" deo~ydibekacin or 4",6"-dideoxy~
dibekacin so obtained may pre~erably be conducted by column
chromatography on an cation-e~change resin containing
~arboxylic function~. To this end, it is recommendable
to make the chromatographic purification by adsorption on
Amberlite IRC 50 or CG-50 (NH4 form or a mi~ture o~ NH4
form and H ~orm) (a product of Rohm & Haas Co,, U.S.~)
-~ollowed by elution with a dilute aqueous a~monia.
The new compound of the ~ormula (II) according to
the aforesaid second aspect of thi~ invention includes l-N--
(I-4~amino-2-hydroxybutyryl)-6" deoxydibekacin and l-N-
(I~4-amino-2-hydroxybutyryl)-4",6"-dideoxydîbekacin.
These particular new compounds may be produced by acylation
* trade mark
. ' I .1 7S~lg
- 31 -
of the l-amino group of 6"-deoxydibekacin or 4"96"-dideoxy-
dibekacin with I~4-amino-2-hydroxylbutyryl group.
Aocording to the fifth a~pect of thi~ invention~
therefore, there i~ provided a proces~ for the production
of 1~ 4-amino-2-hydroxybutyryl)-6"--deoxydibekacin or
l-N~ 4-amino-2-hydroxybutyryl)-4",6"-dideoxydibekacin
of the ~ormula (II)
E2 ~ ~2/ =
0~
H~ ~H2
CO (II)
CHOH
CH2CH2~I2
wherein R i~ a hydroxyl group or a hydrogen atom, which
comprise~
(a3 acylating the l-amino group of 6"-deoxydibekacin
or 4",6"-dideoxydibekacin or a partially amino-protected
derlvative thereof repre~ented by the formula (I~)
- 1 ~7~8
-- 32 --
CH A
R ~ , / ~ \ >N
O
E2 N~B
wherein R is a hydroxyl group or a hydrogen atom9 and A is
a hydrogen atom and at least one B i~ a mono-valent amino-
protecting group but the other ~(s) lS or are each a hydroge~
atom~ or at least one pair o~ A and ~ taken together form
a di-valent amino~protecting group 'DUt the other A and B
are each a hydroge~ atom, and the amino-protecting groups
repre3ented by A and B may be equal to each other or di-
~ferent ~rom each other, by reaction with an amino-protected
derivative of ~-4-amino-2~hydroxybutyric acid or a ~unctional
equi~al~nt thereto, to produce a l~N-acylated product o~
6"-deo~ydibekacin or 4't,6"~dideoxydibekacin repre~ented by
the ~ormula (X)
1~7~19
CH
E2~ f------ Z 2
o~ OH o
~
CO (~)
CHOH
CH2CE2NH~ ~ I
wherein Al is a hydrogen ato~ and ~' is a hydrogen atom
or a mono-valent amino-protecting group, or A' and ~'
taken together form a di-valent amino-protecting group~
or to produce an amino protected l-N-acylated product o~
6" deoxydibekacin or 4",6"-dideoxydibekacin repre~erlted
by the formula ~X')
1 1 7~ 9
- 34 -
CH A
A \ ~ ~ ~ A
B / . OH OH
0, ~ ~ .
~ _ N <
CO
aXOH (X ' )
l~2CH2~ <
:B '
~ .
: wherein R, A, ~9 A~ and ~' are as defined in the above
~ormula ~IX) or (~) 9 and
(b) removing the rema-ining amino~protecting group(~)
~rom the l-N-acylated product of the formula (X) or ~X') i~
a kno~n manner, to produca the compound of the ~ormula (II).
The process oY the fifth aspect of thi~ inventio~
may include a ~urther step of converting the compound (II)
into a pharmaceutically acceptable acid addition salt
thereof by reacting with a pharmaceutically acceptable
inorganic or organic acid in a known manner, if desirad.
~ he procedures ~or oarrylng out the process o~ the
~i~th a~pect o~ this in~ention are now described in more
detailO
In aarrying out the present process, it is possible
~ 17~9
-- 35 --
to employ as the star-ting material 6"-deoxydibekacin or
4",6"-dideoxydibekacin ~I) of which amino group~ are not
protected at all, in the ~orm of the free acid or in the
form of an acid addition salt with an appropriate acid ~uch
a~ hydrochloric acid or sulfuric acid. However, it i~
preferable to employ a~ the ~tarting material ~uch a
partially amino~protected derivative o~ 6"-deoxydibekacin
or 4",6"-dideo~ydibekacin according to the ~ormula ~IX)
in which all or ~ome o~ the amino groups other than the
l-amino group ha~e been protected with known amino-
protecting grOUp(6) and which may be prepared by intro-
duction o~ a known amino-protecting group into the compound
o~ the -~ormula (I) by means OI a known amino-protecting
technique previously adop~ed in the synthesis o~ some
; 15 known deoxy derivatives of kan~mycin B. ~or the pre-
paration of the partially amino-protected 6"-deoxydibekacin
or 4",6"-dideo~ydibekacin derivative of the formula (IX)9
it is ~ea~ible to utilize the amino-protecting technique~
which were employed, for in~tance, in the preparation o~
the 6'-N-benzyloxycarbonyl derivative o~ kanamycin ~ a~
de~cribed in the ~peclfication of U.S. patent ~o. 3 9 781,268
or U.S. patent No~ 3,929,762; the preparation o~ 2',6'-di-
N-tert-buto~ycarbonyl kanamycin B or 6'-N benzylo~ycarbonyl
kanamycin B, or the mono-~ or di-~-tert-butoxycarbo~lyl
and e~en tri-N-tert-butoxycarbonyl derirati~o of 6'-N-
ben~ylo~ycarbony]-kanamycin ~, ei.ther i~olated or in mixture
thereo~, a~ described in the speci~ication o~ U.E. patent
~ ~ 75~ g
-- 36 --
No 1,426,908 or ~.S. patent No. ~93g~l43; or the pre-
paration of 2',3,3"~6'-tetra-N-formyl derivati~e of
kanamycin B as described in the specification of Belgian
patent ~o. 817J546.
In general, suitable examples of the amino-protecting
group which may be used for the protection of some amino
group~ of the compound o~ the general formula (I) as the
initial material or may be present in the partiall~ amino-
protected 6"-deo~ydibekacin or 4"96"-dideo~ydibekacin
derivative o~ the ~ormula ~IX) may be an ordinary amino-
protecting group, including an alko~ycarbonyl group ~uch
as tert-butoxycarbonyl and tert-amylo~ycarbonyl; a cyclo-
alkylo~ycarbon~l group such a~ cyclohexyloxycarbonyl; an
aralkyloxycarbonyl group such as benzyloxycarbonyl; an
acyl group ~uch a~ trifluoroacetyl and o~nitrophenoxyacetyl;
a phosphinothioyl group ~uch a~ diphenylpho~phinothioyl and
dimethylphosphinothioyl; a phosphinyl group such as di-
phenylphosphinyl, and the like. Preferred e~amples of the
di-valent amino-protecting group include phthaloyl group
and a group of Schiff base -type such as ~alicylidene.
~he introduction o~ the amino-protecting group o~ the~e
kinds ~ay be conducted by reacting the compound o~ the
~ormula (I) with an appropriate known reagent for intro-
duction o~ the amino-protecting group which may be in the
~orm of an acid halide, acid a~ide, active ester or acid
anhydride and the like, in the manner known in the con-
ventional synthesis of peptides. By chosing the quantity
i ~l 75~19
- 37
of the reagent ~or introduction of the amino-protecting
group employed in a proportion o~ 0.5 to 6 mol. per mol.
of the compound of the formula ~I), it is po~ible to
prepare a mi~ture o~ di~fere~t, partially amino-protected
derivative~ (IX) at any ratio, due to the di~`ference in
the reacti~ity of the respective amino group8 of the
compouna (I).
In the process of -the fifth aspect of thi~ in~ention,
it i~ practical to employ as the starting material such an
: 10 amino-protected 6"-deoxydibekacin or 4",6"-dideo~ydibekacin
derivati~e in which all or some o~ the amino groups other
than the l-amino group have or ha~ been blocked, ~or example,
a 3,2',6l~3"-tetra-~-protected deri~ative t a 3,2i,6'-tri
N-protected derivati~e9 a 2',6',3"-tri-N-pro-tected deri~ative,
a 2',6' di-N-protected derivative and a 6'-mono-~-protected
derivative. Be3ides, a mixture o~ two or more of these
partially ~-protected deri~ative~ may, without being puri~ied,
be used ~or the l-N-acylation step of the present proces3.
In order to ensure that the desired product o`f the
ganeral ~ormula (II) can be produced in a high yield in
accordance with the process o~ the ~i~th aspect in~ention,
it needs only that ju~t the l-amino group of 6"-deo~y-
dibe~acin or 4",6" dideoxydibe~acin is selectively aoylated
with L-4-~a~ino 2-hydroxgbutgric acld. Accordingl~, it will
be evident that mo~t pre~era~ly, a 3,2'96',3"-tetra-N-
protected derivative o~ the compound (I), that is, the
ami~o-protected derivative o~ the compound (I) in whioh
~758~9
-- 38 --
all the amino groups other than the l-amino group have
been blocked with the protective group~ i~ emp].oyed as
the starting material to be l-N~acylated in the pre~ent
proces~.
~o prepare the ~,2',6',3"-tetra-N-protected
derivative of the ~ormula (IX) from the compound of the
~ormula (I), the following procedure may be used, for
instance. ~hus, there can be applied a known method o~
U.~. patent ~o. 4,136,254 of ~agabhu~han et al by which
a 3~2~,5'-tri-~ acylated protected derivative o~ kanamycin
is ~repared by reacting kanamycin ~ with a di-valent
transition metal cation9 ~or example~ cation o~ copper
~II), nic~el ~II), cobalt (II) etc. for the formation of
a metal complex of kanar~cin ~, reacting thi~ kan~mycin ~-
me~al comple~ with an acylation agent known a~ the amino
protecting group-introducing reagent ~or the protec-tion
of all the amino group~ other than the l-amino and 3"-
amino group~ o~ the kanamycin ~ moiety ol the kanamycin
~-metal complex (said l- and 3"-amino group~ having been
blocked by complexing with the di-valent metal cation in
the kanamycin B-metal complex), and then remo~ing the
di-valent metal ca-tion from ~aid complex, egO, by treat-
ment with hydrogen ~ul~ide or by treatment with aqueou~
ammonia~ Or, -there oan be applied a method o~ our co-
pending Japanese patent application No. 138402/78
(corresponding to our co-pending U.S. pa-tent application
No 090,591; oo-pending U.E. patent application No.7938894;
,;
1~7~81g
- ~9 -
~elgian patent ~oO 879,925) by which a ~,2',6'-tri N
acylated protected derivative o~ kanamycin ~ is prepared
in a ~imilar way to the aforeæaid known method of
Nagabhushan et al except that ~inc cation i~ employed
in stead of -the di-valent tran~ition metal cation. In
thi~ way9 a 3,2',6'~tri-N-protected derivati~e of the
formula (I2) can be prepared from the compound of the
formula (I~ in a high yield. ~he 3"-amino group of thi~
3,2',6'-tri-N-protected deri~ative (IX) so prepared ca~
further be protected by the ~elective acylation according
to a ~elective 3"-~-acylation method of our co-pending
Japan~se patent application NoO 7~064/79 (~ee claim 15 o~
said Belgian patent No. 879,923) for the production of an
amino-protected derivative o2 an aminoglycosidic anti-
biotic of which all the amino group9 other than the l-amino
group have been protected selectively, ~o that a 312',6',3"-
tetra-N-protected derivative o~ the compound (I) can be
prepared in a high yield. In accordance with the ~elective
~"-N-acylation method of the co-pending Japane~e patent
application No. 73064/79 (as described in the claim 15 of
the ~elgian patent No. 879,923), the above-mentioned 3,2',6'-
tri-~-protected derivative o~ the compound (I) i~ reacted
with a ~ormic acid alkyl e~ter, a di-halo or tri-halo-
alkanoic acid alkyl e~ter, formyl~midazole or an N-alkanoyl-
imidazole a~ the acylation agent, whereby the 3"-amino
group can be acylated ~electively with the acyl re~idue
of the acylation agent employed in a high yield, without
, .
8 1 g
- ~o -
involving the acylation of the l-amino group of said
3,2',6'-tri-N-protected deriv~tive. The ~ 9 2',6'93"-
tetra ~-acylated derivative~ for e~ample, 3,2',6'-tri-
N~benzylo~ycarbonyl~ N-tri-fluoroacetyl deri~ative, o~
6"-deoxydibekacin or 4",6'1-dideoxydibekacin which may be
obtained by applying the above-mentioned method~ of the
.S. patent ~oO 4 91~6, 254 and of the ~elgian patent
~o. 879,923 is a most preferred material to be 1-~-
acylated selectively with the ~-4-amino-2-hydro~ybutyric
acid in the l-N-acylation step of the present process.
In the proces~ of this fifth a~pect in~entio~5
the l-amino group o~ the compound of the formula (I) or
the l-amino group of the partially amino-protected
deri~ative~ (IX) thereo~, either isolated or in mi~tura
~ two or more of them, i~ acylated with ~-4-amino-2-
hydro~ybutyric acid of which the amino group i3 not
protected or has been protected. This l-N-acylation may
be conducted according to any of the co~ventional methods
for the ~ynthe~is of peptides, for instance~ according
to the known dicyclohaxylcarbodiimide method, the known
mixed acid anhydride method, the known a~ide method or
the active e~ter method and the like, u~ing I-4-amino-2-
hydroxybutyric acid a~ such or in the f`orm of its reaotiva
derivative ~as the functional equivalent thereo~). Ae the
amino-protecting group ~or protection o~ the amino group
of X~-amino-2-hydro~ybutyri.c acid ma~ be employed such
an amino-protecting group which i~ the ~ame as or di~erent
3~75~1g
41 -
~rom the one pre~ent in the compound (Ig~. Particularly~
a preferred amino-protecting group for thi~ p~rpose i~
tert-butoxycarbonyl group which is easily removable by
treatment with aqueou~ trifluoroacetic acid or acetic
acid or with diluted aqueou~ hydrochloric acid. Benzyloxy-
carbonyl group ~hich i~ r~ovable by a conventional hydro~
genolysis in the pre~ence of a catalyst such as palladium
or platinum o~ide i~ a convenient ~-protecting group.
The 1 N-acylation in the present pro~ess may pre
~erably be carried out in an aqueous organic solvent
according to the active e~ter method u~ing I-4-amino-2-
hydroxybutyric acid in the form of it~ active e~ter.
For example, ~--hydro~ysuccinimide e~er of ~-4 benzylo~y-
carbonylamino-2-hydro~ybutyric acid may preferably be u~ed
a~ the active e~t~r which may be prepared by a conventional
method of preparing the active e~ter. Thi~ active e~ter
pre~erably may be u~ed in a proportion of from 005 to 3
molar equivalents and pre~erably o~ from l to 1.5 molar
equivalents per mol of the 6"-deoxydibekacin or 4"~6"-
dideo2ydibekacin compound to be l-N-acylated. The aqueou~
organic ~olvent u~ed in the reaction medium may be a water-
mi~cible organic ~olvent ~uch a~ dioxane, lt2-dimetho~y-
ethane, dimethyl~ormamlde, tetrahydrofuran, and the like.
The l-N~acylation may be e~ected at ambient temperature
or, lf de~ired, at an elevated temperature of 20~90C and
for a reaction time o~ ~everal hour~ and preferably o~
5~6 hour~.
1~5
-- 42
When the l-N-acyla-tion in the present proces~ i3
conducted using as the starting material such a partially
amino pro~ected derivative in which ~ome9 but not allg of
the amino groups other than the l-amino group has or have
bee~ protec~ed, for e~ample, the 6'-N-protected derivative
of ~"-deoxydibekacin or 4"96"-dideo~ydibekacin, the acylation
products a~ ~ormed may partially be purified by a column
chromatography, for example, on silica gel so that the ~n-
reacted ~-ta~ting material is removed, giving a mixture o~
the de~ired l-N-mono-acylated product with the otherwise
N-acylated products, as the case be in the synthesi~ of
4-amino-2-hydro~ybutyryl)-3',4'-dideoxykanamycin
~ as described in the specification of U.3. patant No~4~107,4240
: These miged acylation products may, without being purified
and/or i~olated, be sub~ected immediatelg to the sub~e-
~uent de-protecting ~tep of the present process, followed
by the purification and isolation so that the desired l-N-
mono-acylated produc-t is obtained.
In the second step of the proce~s of this fifth
a~peot invention, the l-N-acylatioll product (including
the mixed acglation product~) a~ obtained in the 1-~-
acylation step of the present proces~ is sub~ected to the
remo~al of the ami~o-protecting groups, if thesa are still
remai~ing i~ the l-N-acylation product. The removal of
the protectlng groups i8 effected by a conventional de-
protecting technique Thus, the amino-protecting group
of the alkoxycarbonyl type i8 removed by weak acid
S 8 1 9
-- 43 --
hydroly~i~ using an aqueous solution of trifluorcacetic
acid or acetic acid and the like or a diluted aqueou~
solution of a~ inorganic acid such as hydrochloric acid
The aral~ylo2ycarbonyl gro~ such as benzyloxycarbonyl
may be removed by an ordinary catalytic reduction (hydro-
genolysis). When phthaloyl group is present as the ~mino~
protecting group~ it can be removed by heating in a solutio~
o~ hydra~ine hydrate in a lvwer alkanol.
The deprotected acylation product as obtained from
the ~econd, de-protec~ing step of the present process may
contain comprise the desired l-N-acylation product o~ the
formula ~II) together with some i~omers thereofO The
de~ired 1~ 4-amino-2-hydro~ybutyryl) derivative (II)
may be isolated and purified chromatographically using a
cation exchanger conta~ning carboxylic ~unctions, such as
Amberlite CG-50 (a product o~ Rohm & Haas Co., U.S.A.) or
CM-Sephadex C-25 (a product of Pharmacia Co. 9 Sweden) and
as~aying the antlbacterial activity of the fraction~ of
the eluate by means o~ a proper kanamycin-sensitive strain
and kanamycin-resistant strain o~ bacteria.
Besides, the new compound o~ the ~ormula (II)
according to the second aspect of this invention may also
be prepared starting from the known compound, 1-N-(I-4-
amino-2-hydroxybutyryl)-3',4'-dideo~ykanamycin B~ that is,
1~ 4 amino-2-hgdro2ybutyryl)-dibekacln. According to
the sixth aspect o~ this invention, therefore, there i~
provided a process ~or the production of l-N-(I-4-amino-
* trade mark
3 17~9
-- 44 --
2-hydro~ybutyryl) 6"-deoxydibekacin or -4",6"-dideoxy-
dibe~acin of the formula (II)
CH3
X ~ f7 ;;~Z~I2
~
H~ E2
CO
I (II)
CHOE
CH2~E2~H2
wherein R i3 a hydro~yl. group or a hydrogen atom, which
comprise~
(a) protecting with a known amino-protecting group
the fi~e amino group~ of l-N~ 4-amino-2-hydroxybutyryl)-
dibekacin of the ~ormula (VI)
3 ~7~9
- 45
HO ~
2 OH O~ O
O ~
HN ~ NH2 (YI)
CO
CHOH
CH2 CHz~I2
ln a known manner to produce an amino-protected derivative
of the formula (VI')
HO ~ A
A ~ ~
B OH ~ A
7 N < (YI ?
ao
CHOE
CH2CH2<~
3 1 7~8~ 9
- 46 ;
wherein A i~ a hydrogen atom and ~ i~ a mono-valent ~miho-
protecting group9 or A and ~ taken together ~orm a di~valent
amino-protecting group9
(b) protecting with a known di-valent hydroxyl-
: 5 protecting group simultaneously the 4"- and 6"-hydroxyl
group~ of the amino-protected derivative ~VI') in a know~
. manner to produce a protected derivative of the ~ormula
(VI")
~`C'O ~ A> ~ 7 < A
~>~ OH
~ ~ VI")
HN ~ ~ ~
I .
CO
C~OH
i A
CH2CH2N~ E3
wherein A and ~ ha~e the ~ame meaning~ a~ defined above
and the group of the ~ormula X ~ ~ i9 a di-valent
y~ ~
hydrox~l-protocting group where X an~ Y are each a hydrogen
atom, an a~kyl groupt an aryl group, or an alkoxy group,
or the group of the formula X~ / is a cycloalkylidene
group, Y / ~
~ ,1 7~8~'g
~ 47 -
(c) protecting with a known mono-valent hydroxyl~
protecting group either the two, 2"- and 2"'-hydro~yl groups
or the three, 5-, 2"- and 2"Lhydroxyl groups of said pro-
tected derivative tVI") in a known manner to produce an
amino-protected and hydro~yl-protected derivative o~ the
~ormula (VII)
A ~ ~ < B
N<
ao
2"' CHOD . (VII)
I A
CH2CH2N<
wherein A, ~ and the group of the formula ~C ~ have
the ~ame meanings a~ defined above, D is a mono-valent
hydroxyl protecting group in the form of an acyl group,
and E i~ a hydrogen atom or a mono-valent hydroxyl-
protecting group in the ~orm of an acyl group~
(d) removing the group ~ ~C~ ) of protecting
the 4"~ and 6"-hydro~yl groups of said amino-protected
and hydro~yl-protected derivative (VXI) therefrom in a
known manner to produce a partially pro-tected compound
~7
-- 48 --
o~ the formula (VIII)
~O ~ A
~ O\ / N ~ ~ < A
A ~ ~ O
B OD O~ o
0 ~
H~_ <~
CO
CHOD (VIII)
- CE2CH~<
~herein A, ~ D and ~ have the same meanings a~ defined
abo~e,
(e) sul~onylating with a sulfonylation agent the
6"-hydroxyl group alone or ~imultaneou~ly both the 411_
a~d 6"~-hydroxyl group~ o-~ ~aid partially protected com-
pound (VIII) in a known manner to produce a mono-
~ulfonylated or di-~ulfonylated derivative o~ the
~ormula (VIIX')
1 ~75819
49 --
GSO3 ~
OD~ _ Y
<
CO
I (VIII')
GHOD
CH2CH2N<
: . .
wherein A, B, D and E have the same meaningæ a~ defined
abo~e, G is a lower alkyl group o~ 1-4 carbon atoms, an
aryl group such a~ phenyl or p--methylphenyl, or an aralkyl
5 group such as benzyl~ and G' is a hydro~yl group or is the
~ame as the group GSU3- shown in the formula (VIII'~,
(~) reacting the 6'l-mono-sulfonylated or 4",6"-di-
sul~onylated derivative (VIII') with an alkali metal iodide
or bromide in a known manner to replace the 6"-~ul~onylox~
10 group or the 4"- and 6"-sulfonyloxy groups by io~o or bromo
group, respeo~iv01y, and thereby to produce the corresponding
6"-mono-iodo or brvmo derivative or the corresponding 4",6"-
di-iodo or bromo derivative,
(g) reduci~ ~aid 6"-mono-iodo or bromo derivative
or said 4",6"-di-iodo or bromo der.ivative with hydrogen
J .~ 7581g
- 50 ~
in the presence of a hydrogenolysis cataly~t to effect the
dehaloge~ation and thereby to produce the corresponding
6"-mono-deoxy derivative (which is corresponding to ~uch
a compound of the formula (~III') but where the group GS03-
has been converted into a hydrogen atom and G' i~ remaininga~ the hydro~yl group~ or the corresponding 4"76"-did00~y
deri~ative (which i~ corresponding to such a compound of
the formula (VIII') but ~here the group GSO~- and the
Group G' each ha~e been convertea into a hydrogen atom~
represented by the formula (VIII")
c~3
R \B ~ ~ ~ < A
A ~ ~ ~ ~ B
OD 0~ 0
o~,~_
. ~ \ < A
bo B
CHOD (VIII")
I A
CH2CH2N<
wherei~ A, ~7 D and E are as defined above and R i~ a
hydro~gl group or a hydrogen atom, and
(h) removing the remaining amino-protecting group~
and the remaining hydroxyl-protecting group~ from the 6"-
mono-deoxy derivative or 4t',6"-di-deoxy derivative (VIïI")
I ~7~19
-- 51 --
in a known manner to produce l~ 4-amino-2 hydroxybutyryl)-
6"-deo~y or -4~',6~-dideoxydibekacin o~ the ~ormula (Il).
The proce~ of the above-mentioned sixth aspect o~
thi~ inven-tion mayp if necessary9 include a furthsr ~tep of
converting the compound o~ the formula (II) into a pharma-
ceutically acceptable acid addition salt thereof by reacting
with a pharmaceutically acceptable inorganic or or~anic acid
in a known manner
~he step~ (a~ to (h) of the proces~ of the ~i~th
aspect invention may be carried out in the same manner as
hereinbefore describea in respect o~ the ~tep~ (a) to (h)
of the proce~ o~ ~he aforesaid fourth aspect in~ention,
re~pectively. Accordingly, the description~ of the procedure~
for carrying out the respective steps of the proce~ of the
~i~th aspect invention are omitted here.
~ his invention is now illustrated with reference to
the following Examples to which this in~ention is not limited~
E~amples 1 and 2 are illustrative of the first and fourth
aspects of this in~ention, E~amples 4 and 5 are illu~trati~e
of the ~econd and fi~th aspects of this invention~ ~nd
Ex~mpla 3 i~ illustrati~a o~ the si~th aspect of thi~
invention.
~e~
(a) Synthe~is o~ 2"-0-benzoyl-1J3,2',6',3"-penta-
N--tert-butoxycarbonyl~dibekacin
Dibekacin ~3.0 g, 6.65 m mol) wa~ di~solved in 10 m~
of water, and the resulting so.Lutio~ was admixed with
1 17S819
4.63 ~Q (33.24 m mol) of triethylamine and 30 mQ of methanolO To
the admixture was dropwise added a solution of 11.5 g (50 m mol)
of tert-butyl S-4,6-dimethylpyrimid-2-ylthiocarbona-te in 20 mQ
of methanol, followed by stirring at 60C for 5 hours -to effect the
reaction of introducing the tert-butoxycarbonyl group as the N-
protecting group into the starting dibekacin. The reaction
solution was concentrated under reduced pressure to about a half
volume and then poured into 650 mQ of water, followed by standing
overnight in a refrigerator. The precipitate formed was removed
by iltration and washed with 300 mQ of water and with 150 mQ of
ethyl ether to give 4.73 g (yield 75%) of a faintly brown powder
of l,3,2',6',3"-penta-N-tert-hutoxycarbonyl-dibekacin.
This powder (1.0 g, 1.05 m mol) was dissolved in 21 mQ
of anhydrous dimethylformamide, and the resultant solution was
further admixed with 432 mg (3 m mol) of l,l-dimethoxycyclo-
hexane and 42 mg (0.24 m mol~ of p-toluene-sulfonic acid, followed
; by stirring at ambient temperature for 16 hours to effect the
reaction of introducing the 4",6"-0-cyclohexylident group. The
reaction solution was admixed with 0.1 mQ of triethylamine and
then concentrated to dryness under reduced pressure. The residue
was taken up in 100 mR of chloroform and the solution obtained was
; washed with the equal volume of water and dried on anhydrous
sodium sulfate, followed by concentration under reduced pressure,
affording 1.10 g (yield 100%) of a faintly yellow pow~er of
1,3,2',6',3"-penta-N tert-
- 52 -
3 ~75819
-- 53 ~
butoxycarbonyl~4" 9 6"-0-cyclohe~ylidene~dibekacinO
~ hi~ powder ~800 mg~ 00775 m mol) wa~ dis~ol~ed in
16 m~ of dry pyridine, and the solution was admixed with
260 mg (1.86 m mol) of benzoyl chlori.de, ~ollowed by
stirring at 60C overnight to effect the reaction o~
introducing the 2"-0-benzoy~ group. The reaction 901ution
wa~ admi~ed with 1 mæ of methanol to decompose the e~cesJ
of the benæoyl chlorida unreacted. ~he reaction solution
wa~ then concentrated to a æmaller volume under reduced
pre~ure to give an oilO ~hi~ oil wa~ taken up in 50 m~
o~ chloroform, and the resulting solution was washed with
the ~ame volume o~ 5~o aqueous potassium hydrogen sul~ate~
then with the same volume of ~aturated aqueou~ ~odium
hydrogen carbonate, and finall~ with the ~ame volume of
water~ The ~olution in chloro~orm wa~ separated, dehydratea
on anhydrous sodium ~ul~ate and co~centratea under reduced
pre~sur~0 ~he residue was washed with small volumes o~
chloroform and n-hexane, giving 826 mg (yield 94~) o~ a
~aintly ~ellew powder of 2"rO-benzoyl-1,3j2',6',3"-penta-
N~tert-buto~ycarbo~yl-4",6"-0-cyclohegylidene-dibeka~in.
This powder (815 mg~ 0.718 m mol) was dis~olved in
20 m~ o~ a mi~ture of acetio acid7 methanol and water
(2:1:1 by volume), and the resulting solution was stirred
at 50C ~or 1 hour and at 40-C ~or ~ hours to e~ect the
hydrol~tic removal o~ the 4~,6~-0-cyclohe~lidene group4
~he reaction solu-tion wa~ concentrated under reduced
pre~re and the re~idue was taken up in 50 m~ o~
~ ~7$8~9
- 54
chloro~orm. The ~ol.ution was ~ashed with the same volume
of aqueous saturated sodium hydrogen carbonate and with
the same volume of water, followed by dehydratio~ of the
chloro~orm pha~e on anhydrou~ oodium sulfate and con~
centration of the solution in chloroform to dryness under
reduced pressure to glve 758 mg (yield 100%~ o~ a falntly
yellow powdsr o~ 2"-0-benzoyl-1,3,2' 9 6',3"-penta-~ tert-
buto~ycarbonyl-d~bekacin
(b~ Synthe~is of 4"96i'-dideo~ydibekaci~
The 2'~ 0-benzoyl-1~3~2',6' 9 3"~penta-~-tert buto~y-
carbonyl-dibekacin (850 mg, 0.805 m mol) obtained in the
above procedure of E~ample l(a) was dissol~ed in 20 m~ of
anhydrou~ pyridine, followed by addition o~ 483 mg (4~24
m mol) of mesyl chloride thereto and reaction of the ra-
sulting admixture at 40-50 a overnight to ef~ect the
mesylationO ~he reaction mixture was admixed with a small
volume of water for decomposition o~ the unreacted mesyl
chloride, followed by concentration to dryness under
reduced pre~sure. ~he residue was taken up in 100 m~ of
chloroform and the solution obtained wa~ washed ~7ith the
same volume o-f 5% aqueou3 potassium hydrogen sulfate, with
the aame volume o-f aqueous saturated sodium hydrogen carbo-
~ate and with th.e same volume o~ water, and the chloro~o~m
phase was d~hydrated on anhydrous sodium sulfata and con-
centrated to dryness under reduced pressure to a~-ford a
lightly brown powder o~ 2"-0-benzoyl-1,3,2'~6',3"-penta-~-
tert-butoxycarbonyl-4",6"-di-0-mesyl-dibekacin. ~his
I ~ 7581 9
- 55 -
crude powdar was purified by a colum~ chromatography on
silica gel (100 g9 Wako Gel C~100, a product of Wako
Junyaku Co., Japan~ developed with a mixture of chloro-
~orm and ethanol (100:1 by volume) to gi~e 859 ~g (yield
88%) of a puri~ied colorles~ powder~
This colorless powder (350 mg, 0.289 m mol) was
dissol~ed in 7 m~ o~ anhydrous dimeth~lformamide9 and the
resulting solution was admi~ed with a large exces~ ~3.5 g)
o~ sodium iodide~ followed by ~tirring at 95 C for 5 hours
under argon atmosphere to effect the 4"96"-di-iodinat~onO
The reaction ~olution was admixed with 50 mQ of water, and
the precipitate formed was removed by filtration a~d washed
with water~ The precipitate collected was dissolved in
50 m~ of chloroform~ and the solution wa~ washed with the
same volume of 20% aqueou~ sodium thiosulfate and then
with the same volume of water. ~he chloro~orm pha~e was
dehydrated on anhydrous sodium sul~ate and concentrated to
dryness under reduced pressure, giving a colorless powder
(~59 mg). This powder was purified by a chromatograph~
on thin layer of ~ilica gel (Silica Gel ~ 500t K~ plate,
a produrt o~ Camag AG., Swis~) developed with a mixture o~
chloroform and ethanol (20:1 by volums), when the product
pre~ent in the region o~ the silica gel thin layer cor~e-
sponding to R~ 0.6 was colleoted. A colorless powder
(210 mg, yield 57%) of 2"-0-benzoyl-1~3,2',6',3"-penta-
N-tert-butoxyoarbonyl-4",6"-di-iodo-dibekacin was afforded.
Thi~ colorless powder (210 mg) was dissolved in
* trade mark
1 ~7~9
-- 56 --
5 m~ of dioxane~ and the solution was admixed with 50 mg
of Raney nickel catalyst (R-200~ a product o~ ~ikko Rika
CoO, Japan) and was subjected to the catalytic hydrogenoly~
~i~ with h~drogen at a hydrogen pressure of 3,6 Kg/cm2 in a
Parr apparatus overnight at ambient temperature to effect
the de-iodination. The reaction mi~ture was filtered to
remove the catalyst there~rom, and the filtrate was con-
centrated to dryness under reduced pressure. ~he residue
was dissolved in 5 mQ o~ 12% ammonia~methanol and the
solution was allowed to stand o~ernight to e~ect the
debenzo~lation~ The reaction solution was then con-
cen-trated under reduced pressure, and the solid residue
wa~ taken up in 20 m~ of chloroform~ followed by washing
with water. The solution in chloroform was then dried on
anhydrou~ sodium sulfate and concentrated to dryness under
reduced pressure, ~ielding a colorless powder. This powder
was dissolved in 2 m~ of a ~olution of 90~ tri~luoroacetic
acid in water and the~ allowed to stand at ambient tem-
perature for 45 minutes to ef~ect the removal of the tert~
butoxgcarbonyl groups. The reaction ~olution was again
co~centrated under reduced pressure and the re~idue wa~
wa~hed with a ~olume of e~hyl ether to give a faintly
yellow powder (comprising the tri~luoroacetate o~ the
desired 4",6'l-dideoxydibekacin). ~his powder was taken
up in a volume o~ water and passed through a column of
17 mQ of Amberlite CG-50 (NH~+) ~or adsorption of the
de~ired product. The Amberlite column wa~ washed with
~ ~ 7
-- 57 --
water and then eluted with 0 4M aqueous ammonia. The
biologically active eluate was collected and concentrated
to dryness under reduced pressure to afford 36 mg of a
colorles~ powder o~ the desired 4",6"-dideo~ydibekacin in
the ~orm of its sesqui-carbonate. Yield 52~. Thîs powder
decomposed slowly at about 129C and gave a specific optical
rotation ~]2~ = + 126 (c 0,5~ water).
~2a~
Synthesis of 6"-deoxydibekacin
The 2"-0-ben~oyl-1,3,2',6'~3"-penta-~-tert-buto2y-
car~o~yl-dibekacin ~500 mg, 0.473 m mol) obtained ~n the
procedure o~ ~xample l(a) wa~ dis~olved in 10 m~ of an-
hydrous pyridine, and the resulting solution was aam~xed
with 130 mg (1.14 m ~ol) of mesyl chloride. The admixture
was ~tirred at 30C overnight to ef~ect the mesylat~on.
The reaction solution was admixed wlth a small volume o~
water, followed by standing for 30 minutes at ~mbient
temperature to decompose the unreacted mesyl chloride.
The reaction solution was concentrated to dryness ~der
reduced pressure, and the residue was di~solved in 50 mQ
o~ chloro~orm. ~he solution in chloro~orm was washed with
the ~ame volume of 5% aqueous potassium hydrogen sulfate,
with the ~ame volume of aqueous saturated sodium hydrogen
carbonate and then with the ~ame volume of water. The
chloro~orm pha~e was dried on anhydrous ~odium sul~ate
and concentrated to dryne~s under reduced pre~sure to
give a lightly brown powder. This powder was purified
~758~9
-- 58 --
by a column chromatography on silica gel (Wako Gel C-200,
a product o~ Wako Junyaku CoO, Japa~) developed with a
mi~ture o~ chloro~orm and ethanol (50:1 by volume) to give
114 mg (yield 20~o) of a colorless powder of 2"~0-benzogl~
1,3,2',G',~"-penta-N-tert-butoxycarbonyl-4",6"-di-0-mesyl~
dibekacin and 250 mg (yield 47~) of a colorless powder of
2"-O-benzoyl-1,3,2'~6'j3'l-penta-N-tert-buto~ycarbonyl-6"-
0-mesyl-dibekacin.
The colorless powder (39 mg, 0.0344 m mol) of the
latter, that is, the 6"-0-mesylated product was dis~olved
in 1 m~ o~ anhydrou~ dimethylformamide, and the resultant
solution was admixed with 390 mg (2,6 m mol) of sodium
iodide, ~ollowed by agitating the admixture at 90C for
2 hours to effect the 6"-iodinationO The reaction solution
was admixed with 30 m~ of chloroform, and the ~olution
obtained wa~ washed with three 30 m~-portions of aqueous
~aturated sodium chloride~ with 30 m~ o~ aqueous 20~ sodium
thiosul~ate and then with 30 m~ of water. ~he chloroform
pha~e was dried on anhydrou~ sodium sulfate and concentrated
to dryness under reduced pressure, giving 38 mg of a color-
less powder of 2"-0-ben~oyl-1,3p2',6',3"-penta-N-tert-
butoxycarbonyl-6"-iodo-dibekacin.
~hi~ powder wa~ dissol~ed in 1.5 m~ of dioxane and
the solution was admixed with 20 mg of Raney nickel catalyst
(R-200, a product of ~ikko Rika Co., Japan) and then sub-
jected to a catalytic hydrogenolysis at a hydrogen pressure
of 3.6 ~g/cm2 *or 24 hours in a Paar apparatus to effect
3 175
-- 59 -
the de~iodinationO The reaction mi~tu~e was filtered to
remove the catalyst there~rom, and the filtrate was con-
centrated to dryness under reduced pressure~ The residue
was taken up in 2 m~ of a solution of 12~ ammonia in
methanol. The resulting solution was allowed to ~tand
at ambient temperature overnight to effect the de~benzoyla-
tion. The reaction solution was concentrated to dryness
under reduced pressure to a~ford a colorless powder. ~his
powder wa~ dissolved in 1 mQ o~ aqueou~ 90~ tri*luoroacetic
acid, followed by standing at ambient temperature ~or 45
mi~utes to e~fect the removal o~ the tert-butogycarbonyl
groups. The reaction solution was concentrated to dryness
under reduced pressurei and the residue was wa~hed ~ith
a volume o~ ethyl ether to give a faintly yellow powder
(comprising the trifluoroacetate of the desired 6"~deoxy-
dibekacin). This powder was dissolved in water ancl the
aqueous solution was passed through a column of 5 m~ of
Amberlite CG~50 (~E4~ ~or adsorption of the desired
product. The Amberlite column was washed with water and
then eluted with 004M aqueou~ ammonia. The active eluate
wa~ collected and concentrated to dryness under reduced
pre~ure to give 12 ~g of a colorle~ powder of 6"-deo~y-
dibekacin in the form of it~ ~esqui-carbona-te. The ~ield
was amounting to 75% as calcula-ted from the intermediate
6"-0-mesylated product mentioned hereinbe~ore. This 6"-
deogydibekacin sesqui~carbonate decomposed slowly at about
1~1C and gave a specific optical rotation [a]D6 = ~ 101-
~75819
_ 60 --
(c 0044, water).
(a) Synthesis of 2"92"'-di-0-benzoyl-3,2',6'73",4"9-
penta-N-tert-buto~ycarbonyl-l NA (I-4-amino 2
hydro~ybutyryl)-dibekacin
l-N~ 4-Amlno--2-hydroxybutyryl)~dibekacin (679 mg~,
1.23 m mol) wa~ dissolved in 14 m~ of aqueous 50% dio~ane9
and to the re3ulting solution was added 1 m~ ~7.2 m mol) of
triethylamine and then dropwise a ~olution of 2.96 g (12.3
m mol) of tert-butyl S-4,6-dimethylp~rimid-2-ylthlocarbonate
in 7 m~ of dioxane ~he admi~ture obtained was ~tirred at
ambient temperature for 40 hours to e~fect the ~-tert-
butoxycarbonylation, followed by concentration of the
reaction solution to dryness under reduced pressure. The
re~idue was washed with 100 m~ portions of n-hexane and
water, a~fording 1.0 g ~yield 82~) of a faintly yellow
powder of 3,2' 9 6' 9 ~1; 4"'-penta-N-tert-buto2ycarbo~yl-l-N-
(I-4-amino-2-hydro~ybutyryl)-dibekacin.
This powder (1.0 gO~ 1 01 m mol) was ta~en up in
20 mb of anhydrous dimethylformamide, and the ~olution
obtained wa~ admixed with 4 mg of p-toluene~ulfonic ac~d
a~ld 0.5 m~ (4.1 m moL) of 2,2-dimetho~ypropanel fol:Lowed
by ~tirrin~ the admlxture at ambient temperature for 17
hours to effect the 4",6"-0~ opropylidenation. The re~
action solution, after addition of one drop of triethyl-
amine thereto, was concentrated to dryness under reduced
pre~sure, and the re~idue wa~ dissolved in 150 m~ of
1~7~19
61 --
ehloroformO The solution in chloroform was then washed with
two lO0 m~ portion~ of aqueous satura-ted sodium hydrogen
carbonate and then with lO0 m~ o~ aqueous saturated ~odium
chloride~ and the chloroform phase was subsequently dried
on anhydrous sodium sulfate and concentrated to drynes~ to
give l.l g of a lightly yellow crude powder of 3,2' 7 6',3",4"1-
penta-N-t ert-butoxycarbon~l-4",6"-0-isopropylidene-l-N~ 4-
amino-2-hydroxybu-t~ryl)-d ibekac in.
This powder was dissolved in 30 m~ of chloroform and
purifiea chromatographically in a column of silica gel
(100 g., Wako Gel C~200, a product of Wako Junyaku COr ~
Japan) de~eloped with a mi~ture of chloro~orm and ethanol
(20:1 by ~olume). The fractions of the eluate which con~
talned the desired product were concentrated to dryne3s
under reduced pressure to afford 842 mg of a purified
colorless powder of the desired product (yield 77~).
This powder (840 mg, 0.78 m mol) was dissolved in
10 m~ of anhydrous pyridine to which was then addecL 0O4 m~
(3.4 m mol) of benzoyl chloride. The resulting solution
wa~ ~tirred at ambient temperature for l8 hour~ to`e~ect
the 2",2"'-di-0 benzoylation. The reaction solution wa~
admixed with one drop o~ water to decompose the un~eacted,
exces~ive benzoyl chloride, and the reaction solution was
-then concentrated to dryness under reduced pressure. The
residue wa~ taken up in lO0 m~ of chloroform and the
~olution wa~ washed with lO0 m~ of 0.25N hydrochloric acid,
with three 50 m~ portion~ o~ aqueou~ saturated sodium
1J175~19
62
hydrogen carbonate and then with 100 m~ of aqueou~ ~aturated
~odium chloride. The ~olution in chloroform wa~ dried on
anhydrous sodium sul~ate and concentrated to drynes~ under
reduced pres~ure to a~ford 1.017 g (yield 100%) of a faintly
yellow powder of 2",2"'-di-0-benzoyl 3,2'~6'~3'`,4"'-penta-
N-tert-buto~ycarbonyl-4"96"-0-i~opropylidene-l-N-(I-4-amino-
2-hydroxybutyryl)-dibekacin.
Thi~ powder (975 mg9 0.76 m mol) was dissolved in
20 m~ of a mixture of acetic acid, methanol and water
(2:1:1 by volume) and the resulting solution was agitated
at ambient temperature for 15 hours to ef~ect the de-
i~opropylidenation, and the reaotion solution was con-
centrated to dryne~s under reduced pressure. The re~idue
wa~ dis~ol~ed in 100 m~ of chloro~orm and the solution
wa~ wa~hed with two 100 m~ portion~ of aqueous saturated
sodium hydrogen carbonate and then with 100 m~ o~ aqusou~
saturated sodium chloride. ~he chloro~orm phase (the
solution) wa~ dehydrated with anhydrous sodium sul~ate
and then concentrated to dryness under reduced pressure
to give 873 mg (yield 93%) of a faintly yellow powder o~
2l',2"'-di-0-benzoyl-3,2',6',3",4"'-penta-N-tert-butoxy-
carbonyl-1-~-(1-4-amino-2-hydroxybutyryl)-dibekacin~
(b) Synthe31~ o~ 1-N~ 4-amino-2-hydro~ybutyryl)-
4",6'1-dideoxydibekac in and 1-N-(L-4-amino-2-
hydroxybutyryl)-6"-deoxydibekacin
2",2"'-Di-0-benzoyl-3,2',6',3",4"' -penta-N-tert-
butoxycarbonyl-l-N-(L-4-amino-2-hydroxybutyryl)-dibekacin
~ ~75819
6~ -
(250 mg, 0~2 m mol) obtained in the .E~ample 3(a) was
dissolved in 5 m~ of anhydrous pyridine, and the resulting
solution, after addition of 0.06 m~ 3 m mol) of mesyl
chloride thereto, was agitated at ambient temperature for
16 hour~ to effect the mesylation. To the reaction
solution was added one drop of water to decompose the
unreacted~ excessive mesyl chloride. The reaction solution
wa~ then concentrated to dryness under reduced pressure and
the residue was dissolved in 25 m~ of chloro~orm. The
solution obtained was washed with 25 m~ of 0~2N hyclrochloric
acid, with three 25 mQ portions of aqueous saturated sodium
hydrogen carbonate and then with 25 m~ of aqueou~ ~aturated
qodium chlorîde, and subsequently the solution in chloro- i
form was dried on anhydrous sodium sulfate, ~ollowed by
concentration to dryness under reduced pres~qure, giving
271 mg (yield 96%) of a lightly brown powder comprising
2"~2"'-di-0-ben~oyl-392',6',~",4"'-penta-N-tert-buto~y-
carbonyl-4",6"-di-0-mesyl-1-N-(I~4-amino-2-hydroxybutyryl)
dibekacin (which gave a single spot at Rf 0.25 in a silica
gel thin layer chromatography developed with chloroform-
methanol (30:1)) and a minor proportion o~ 2",2"'-di-0-
benzoyl-3,2'~6',3",4"'~penta-N-tert-buto~ycarbonyl-6"~0-
mesyl-l-N~ 4 amino-2-hydroxybutyryl)-dibekacin (which
gave a single spot at Rf 0.18 :Ln the same silica gel thin
layer chromatography ~ust mentioned above).
Thi~ powder (2~0 mg, 0.155 m mol) was dissolved in
4.4 m~ of dry dimethyl~ormamide7 and the ~olution was
3 ~1 75~1 ~
admixed with a large excess (2.3 g) of sodium iodide, followed
by agitation at 90 C for 6 hours to effect the substitutive
iodination. The reaction solution was admixed with 50 mQ of
water and the precipitate as formed was removed by filtration.
The precipitate collected was dissolved in 20 mQ of chloroform
and the solution was washed with two 20 mQ portions of aqueuos
20g6 sodium thio~ulfate and then with 20 mQ o~ aqueous saturated
sodium chloride. The solution in chloroform was dried on anhydrous
sodium sul~ate and concentrated to dryness under reduced pressure
to give 224 mg (yield 99~6) of a colorless powder comprising a main
proportion of 2",2"'-di-O~benzoyl-3,2',6',3",4"'-penta-N-tert-
butoxyearbonyl-4",6"-di-iodo-l-N-(L-4-amino-2-hydroxybutyryl) -
dibekacin (which gave a single spot at Rf 0.40 in a silica gel
thin layer ehromatography developed with chloroform-ethanol (30:1)
and a minor proportion of the corresponding 6"-mono-iodo product,
namely 2",2"l-di-0-benzoyl-3,2',6',3",4"'-penta-N-tert butoxy-
carbonyl-6"-mono-iodo-1-N-(L-4-amino-2-hydroxybutyryl)-dibekacin,
(which gave a single spot at Rf 0.31 in the same silica gel thin
layer chromatoyraphy just mentioned above).
This powder (220 mg, 0.15 m mol) was dissolved in 5 mQ
of dioxanel an~ the solution was subjected to a catalytic hydro-
genolysis under a hydrogen pressure of 3.6 Kg/cm for 5 hours in a
Paar apparatus in the presence of a Raney nickel catalyst (R~200,
a produet of Nikko Rika Co., Japan) to eEfect the de-iodination.
The reaction solution was filtered to remove the catalyst. The
filtrate was concentrated to dr~ness (169 mg) under recluced pressure
-- 64 --
~ I758
-- 65 --
and the re~idue was taken up in-to 10 m~ of 12~ ammonia
methanol. The resulting solution was allowed to ~tand at
ambient temperature overnight to effect the de-benzoylation~
The reaction ~olu-tion was concentrated to dryness (1~1 mg)
under reduced pressure, and the residue was di~solved in
2 m~ of aqueous 90% trifluoroacetic acid, followed by being
allowed to stand at ambient temperature for 45 minute~ to
e~fect the removal of the ter~-bu-to~ycarbonyl groups.
The reaction solution wa~ concentrated to dryness under
reduced pressure and the residue was washed with ethyl
ether, a-~fording a faintly yellow powder (comprising the
trifluoroacetate~ of l-N-(1~4-amino-2-hydroxybu-tyry1)-4"~6"-
dideoxy- and 6"-deoxydibekacin~). This powder was di~solved
in 2 m~ o~ water and adjusted to pH 7.8 by addition of lN
aqueous ammonia, and the resulting solution was passed
through a column of 20 m~ of Amberlite CG-50 (NH4~) for
ad~orption o~ the desired products. The ~mberlite col~unn
wa~ washed with water (68 m~), with 0.2N aqueous ammonia
(70 m~) and then with 0~5N aqueous ammonia (88 m~)) followed
by elution with 0.6N aqueous ammonia (80 mQ) and then with
0~7N aqueou~ ammonia (50 m~ he eluate ~rom -the elution
with the 0.6N aqueous ~mmonia were combined together and
concentrated to dryne~s undar reduced pressure to give
7.2 mg (yield 8~o) Of a colorles~ powder o~ 1-N-(L-4 amino-
2-hydroxybutyryl)-6"-deoxydibekacin (as the sesqui~oarbonate).
The fractions from the elutlon with the 0.7~ aqueous ammonia
were combined together and concentrated to drynes~ under
1J17S~9
- 66 -
reduced pre~s~e to give 18.8 mg (yield 21~o) of a colorles~
powder of l~ 4-amino-2-hydro~ybu-tyryl)~4",6"-dideoxy-
dibekacin (as the sesqui-carbonate). This product der
composed a-t 142~147C and ga~e a ~pecific optical rotation
~D4 = + 84 (c 0.57 water)~
E~ample_4
Synthesis of 1-~ 4-amino-2--hydroxybutyryl)-6"-
deoxydibekaein
6"-Deo~ydibekacin (as the sesqui-carbonate) (26.4 mg~
0.05 m mol) obtained in the Example 2 was dissolved in 0~5 mQ
of anhydrou~ dimethylsulfoxide~ and the resulting solution
was admixed with 54.7 mg (0.24 m mol) of zine aeetate
(Zn(C~3C02)2 2H20) and stlrred at ambient t~mperature for
20 hours to effect the formation o~ the comple~ of 6"
deoxydibekacin with æinc cation. The ~olution eontaining
said compleg was admixed with 41~2 mg (0~165 m mol) of
benzyloxycarbonyl-~-hydroxysuccinimide ester, and the
admixture wa~ agitated for 20 hours at ambient temperature
to effect the benzylo~ycarbonylation of the 3-, 2'- and
6'-amino groups of 6"-deo~ydibekaein which were not com-
ple~ing with the zinc cation~ The reaction solution wa~
admixed with 30 mQ of water and a small volume of eon
eentrated aqueou~ ammonia 90 that the reaction solution
wa~ adju~ted to pH 11, whereby the breakdown of the zine-
complexing with the 3~2'J6'-tri-N-benzyloxyearbonylated
6"-deoxydibekaein was effeeted, The reaction solution ~o
treated whieh eontained the preeipitate as formed was
~7~819
-- 67 --
filtered, and the precipitate collected (which comprised
the 392',6'~tri-N-ben~yloxycarbonyl~6"-deo~ydibekacin)
was dissolved in 5 m~ of dimethylsulfoxide~ To the re-
sultant solution was added a ~olution of 0.02 m~ (0.14 m mol~
of ethyl trifluoroacetate in 1 m~ of dimethylsul-fo~ide,
followed by agitation at ambient temperature for 3 hours
to effect the selective 3"-N-trifluoroacetyla-tion and to
give the reaction 301ution containing 3~2',6'-tri-N-benzyl-
oxycarbo~yl 3"-N-trifluoroacetyl-6"-deo~ydibekacin. Thi~
reaction solution was admixed with 0~01 mQ (0.07 m mol) of
triethylamine and with 26.3 mg (0.075 m mol) of N~h~droxy-
succinimide ester o~ ~-4-benzyloxycarbonylamino-2-hydroxy-
butyric acid, and the admi~ture obtalned wa~ stirred for
15 hours at ambient temperatuL~e to e~fect the acylation of
the l-amino group of the ~"-deo~ydi~ekacin compound. The
reaction mi~ture was admixed with 2 m~ of aqueous saturated
sodium chloride and 2 m~ of ethyl acetate, and the ethyl
acetate phase together with the precipitate formed wa3
concentrated to dryne~ to give 44.2 mg of a powder. This
20 powder wa~ admixed wîth 2 mQ of a mixture of tetrahydrofuran
and lN aqueou~ ammonia (1:1 by ~olume), and the resultant
admixture was stirred at ambient -temperature for 20 hour~
to effect the removal of the trifluoroacetyl g~oup. The
reaction solution obtained was concentrated to a ~mall
volume, followed by addition of 2 ~ of lN aqueous ammonia
there-to~ The precipitate formed was removed by filtration
and washed with water. This precipitate was dissolved in
1~758~9
_ 68 --
20 mQ of a mixture of aoetic acid, methanol and water
(2:1:1 by volume~ and the resulting solution9 after
addition of 50 mg of 5% palladium-on-carbon thereto~ wa~
subjected to catalytic hydrogenolysis by passage of a
hydrogen stream through said solution for 6 hours at
ambient temperature. The reaction mixture was filtered
to remove the catalyqt therefrom, and the filtrate was
concentrated to dryness under reduced pressure. The
residue was taken up into 0.3 m~ of water and the aqueous
solution was adjusted to pH 7.8 by addition o~ lN aqueous
ammonia. This solution wa~ charged into a column o~ 2 mQ
of Amberlite CG-50 (~E4+ ~orm) ~or adsorption o~ the
desired product. The col~mn was then washed with 5 mQ
of water and with 5 m~ of O.lN aqueous ammonia, followed
by elution with 5 m~ of 0.5N aqueous ammonia and then with
10 m~ o~ 0.8~ aqueous ammonia. The fractions from the
elution with the 0~8N aqueous ammonia were combined together
and concentrated to dryness under reduced pressure to give
10.0 mg of a colorle~s powder o~ the desired 1-N~ 4-
amino~2-hyaroxybutyryl)-6"-deo~ydibekacin ~sesqui-carbonate).
Yield 32% Thi~ product decomposed at 132~139C ancL gave a
0pecific optioal rotation [~D3 ~ ~ 73 (c 0.3, water).
. ~ ~
Synthe~is o~ l-N-(L-4-amino-2-hydroxybutyryl)-4",6"-
dideoxydibekacin
The process of the above ~xample 4 was repeated
using 25.6 mg (0~05 m mol) of 4",6'l-dideoxydibekac:in sesqui-
1~75~19
_ ~9 _
carbonate instead of the 6"-deo~ydibekacin. A colorless
powder o~ l-N~(L-4-amino-2 hydroxybutyryl)-4",6~'-dideoxy-
dibekacin (sesqui-carbonate) was ob-tained. ~ield 9.0 mg
~290 .
