Stereoselective oligomerizations catalyzed by lipases in organic solvents

Tetrahedron Printed in

Letters,Vol.28,No.l5,pp Great Britain

1607-1610,1987

0040-4039/87 $3.00 + .oO Pergamon Journals Ltd.

STEREDSSLSCTIVS OLIGOMERIzATIoNs CATALYSED BY LIPASSS IN ORGANICSOLVENTS

Alexey L. Mrgolin,

Jean-lhres Crenne, and Alexander II. Klibanav*

Department of Applied Biological Cambridge, Massachusetts 02139

Sciences,

Massachusetts

Institute

of Technology,

Lipases have been found to act as stereoselective catalysts FE=: oo vcondensation reactions between racemic diesters and achiral diols ior-vice versa) in organic solvents. Optically psntamers have been formed as a result.

Optically

active

polymers possess

have found applications adsorbent6

as catalysts

for separation

synthetic

optically

polymerization

active

solvents active

can catalyze

as the reaction

ification

products9 .

enzyme-catalyzed

different

organic

solvents”.

One of such reactions

The initial

commarcially

available

lipases12

all

subsequent,

scaled-up

active

via

polymers and

initiators2’3.

In

with organic

is stereoselective

(or vice

versa)

stereoselective

(A) and 1,6-hexanediol

were tested

as catalysts

c1~2~2~(0)~rCH2~2MBrC(O)~2~2C1

+

were catalytically afforded

experiments.

of 2 in 50 mG of toluene,

all

leading and diol

transesterto optically

are used

polycondensations

in

model system examined was comprised of bis(2-

(_+)-2,5-dibromoadipate1’

’ , and nearly

Thus far,

to achiral

to us that if a racemic diester

chloroethyl)

reaction’

coupling

when water is replaced

and an alcohol

may bs able to catalyze

and Cbromobacterium sp.”

crystals6.

and

chiral

asymmetric polymerizations.

processes’

medium’.

It occurred

then lipase

and unique propsrties’-3

and liquid

wnomars or their

between a racemic ester

instead,

and

for asynrnetric syntheses4,

of racemic Mmoomsrsusing optically

work, we have explored Lipases

trimsrs

polymers have been prepared by chemical means, e.g.,

of optically

asynnnetric polymerization this

a number of valuable

and reagents

of racemic mixtures5,

active

active

in

active.

the highest We dissolved

(2) in toluene.

HOta J6m -> Lipases

activities

from Aspergillus

niger

and were employed in the

3.7 amoles of racer&

and then 11 g of kspergillus

Nine

of the polycondensation

-

lipase15

i and 13 rmnoles was added.

The

suspension’ ’ was shaken at 45’C and 250 rpm for 7 days when the degree of conversion the ester

reached approximately

SO%, as judged by a gas chromatographic 1607

analysis

of

of both

1608

the 1 remaining and the Z-chloroethanol tion

and the solvent

subjected

and 2-chloroethanol

to preparative

the fractions molecular

obtained

weights

formed.

evaporated under vacuum.

HPLC (hexane/ethyl

acetate

greater

than 450 daltons

and smaller

quantities

IR revealed

the presence

the absence of carboxyl

products (C

4,

respectively,

same system, +1.3”

hydrolysis

of the enzymatically

where A is the diol

were found to be optically

active

of these products

and ‘H NMRstudies

of esters)

and the ClCH,CH,

moiety and B is the acid moiety. and +4.3”,

respectively

in the

with [crli5 of

(c 10, TIiF).““g

Asymmetric enzymatic oligomerization bis(2,2,2-trichloroethyl)

Both

respectively

(14 g15) was used as a catalyst

140 mg of the trizter and 80 mg of the pentamer were formed”

and +1.5”,

by

indicated

formed trimer and pentamer is A-B-A

with [crli5 of +4.5’

When Chromobacterium lipase

lliF).l’t’g

and

with apparent

80 mg of a pentamer,

Analysis

bonds and free OH groups,

groups (thus excluding

Hence, the composition

Fractions

140 mg of a trimer,

of higher polymers were obtained. of ester

The remainder was

as the mobile phase),

were pooled and rechromatographed using

gel permeation HPLC. As a result,

and A-B-A-R-A,

gradient

were analyzed by gel permeation HPLC”.

preparative

moiety.

The enzyme was then removed by filtra-

was also observed when another racemic ester,

(_+)-3-methyladipate,

was used where the asyzznetric

carbon is

Cl,CM,OC(OKH,CHH(U$ DiH,CH2C(ONH2CCl, + HO(CH,),OH -> farther

from the reaction

sites

turned out to be the most active

than in 1.

Porcine pancreatic

catalyst

in this

case,

lipase

(344 mg/mL15)

and under the same conditions

described

above,

260 mg of the A-R-A trimer and 35 mg of the A-R-A-R-A pentamer”

obtained;

[a]:’

of their

mixture was +0.3”

In both examples described, lipase-catalyzed

production

ogy, we then reversed achiral

diester,

(c 30, HeOH).”

a racemic diester

of ester

the situation

bis(2-chloroethyl)

oligomers.

toluene

and in isopropyl

was

ether,

adipate”.

(no 2-chloroethyl [alE5

of -7.9’

catalysts in organic

after

were involved

in

of our methodol-

2,4-pentanediol, lipase,

+ HCCH(CH~ )cII,CH(~

)oH ->

and an

both in

only with the (2R,4R)-(-)

but 15

When 10 g of the enzyme

10 zzzoles each of racemic 2,4-pentanediol

14 days of shaking 350 mg of a mixture of a trimer and pentamer”

moiety by NHR; both ester (c 2, CI$Cl,)

for the production solvents.

diol,

in that reaction.

containing

diol

the generality

Porcine pancreatic

was found to be reactive

isomer of the diol

suspended1 6 in 25 z& of toluene

and the diester,

and an achiral TO test

and employed a chiral

C1cH,cH,oC(0)~Cy~H,cH,C~O~ocH,cH,Cl

not with the (2S,4S)-(+)

as

were

was formed. of optically

bonds and free OH groups present Hence, lipases

active

ester

by IR) with

can act as stereoselective

oligomers

of different

StnXtUreS

1609

R.C. ; Kaiser,

1.

Schulz,

2.

Selegny,

F.;

3.

E. Adv. Polymer Sci.

1965, 4, 239-315.

Pino, P.;

Ciardelli,

Zandomeneghi, M. Ann. Rev. Phys. Chem. 1970, 21, 561-608. E., Ed. Cptically

(Holland),

1979.

Ciardelli,

F.

active

In Encyclopedia

H.F. ; Bikales,

Polymers; D. Reidel

Publishing

Co.:

of Polymer Science and Engineering,

N.M.; Overberger,

C.G.; Menges, G.; Eds.; Wiley:

Dordrecht

2nd ed.;

Mark,

New York, 1986, in

press. 4.

Luisi,

P.L.

Aglietto,

In Reference

M. Pure Appl.

2; pp. 357-401.

5.

Manecke, G.; Lamer, W. In Reference

6.

Plate,

N.A.;

Freidzon,

Goodby, J.W. Science

Ciardelli,

Ya.S.;

1986, 231, 350-355.

Zaks, A.;

Klibanov,

Klibanov,

A.H. CHENTECH 1986, 16, 354-359.

9.

(a) Kirchner,

10.

polycondensation

in aqueous-organic

Lett. acid 11.

All

Yoshimoto,

T.;

diesters

Biol.

by lipase

J.;

1985, 107,

Buono, G.; Triantaphylides,

between achiral

dicarboxylic reported

Chem. 1984, 48, 2805-2808).

C.

Y.;

from the corresponding procedure:

and primary Iwai,

M.;

Ajima -et al. (Ajima, Inada, Y. Biotechnol.

of achiral

in benzene but no convincing

to the general

acids

(Okumura, S.;

Also,

Tamaura, Y.; Saito,

K.;

claimed a polycondensation

were synthesized

EtOH according

A.M. J. Am. Chem. Sot.

mixtures was recently

Takahashi,

1985, 7, 303-306) catalyzed

USA 1985, 82, 3192-3196.

1986, 27, 29-32.

Tominaga, Y. Agric. A.;

M.P.; Klibanov,

(b) Langrand, G.; Baratti,

Lipase-catalyzed glycols

A.M. Proc. Natl. Acad. Sci.

G.; Scollar, Lett.

C.;

2; pp. 403-410.

8.

7072-7076.

E.; Carlini,

Shibaev, V.P. Pure Appl. Chem. 1985, 57, 1715-1726.

7.

Tetrahedron

F.; Chiellini,

Chem. 1980, 52, 1857-1864.

lo-hydroxydecanoic

evidence

was presented.

acyl chlorides

and chlorinated

Sonntag, N.O.V. Chem. Rev. 1953, 52,

237-416. 12.

Lipases

were obtained

as follows:

porcine

pancreatic,

wheat germ from Sigma Chemical Co.; Aspergillus

&

niveus N, Rhizopus orysae FAP, and lipoprotein

13.

Amano International

&rqme Co.;

Specific

activities

of lipases

units/mg

solid,

All

were 11, 500, 8.6,

(X-fold)

10, 10, 45, 750, 2190, and 120

were tested

both “straight

from an aqueous solution increase

for lipoprotein

mixtures were shaken at 45°C; periodically,

Only for the Chrorwbacterium lipase lyophilized

from Pseudomonas sp. from

in toluene were 100 mg/mL, except

The reaction

lipases

lyophilized

lipase

Chrcawbacterium sp. CV from Finnsugar Biochemicals.

withdrawn and assayed by gas chromatography9 14.

and

respectively.

Lipase concentrations mg/ant).

Candida cylindracea,

K, Mucor sp. HAP, Rhizopus

in the specific

frcm pi 7.0 prior

aliguots

were

and “pH-adjusted”’

, i.e., ’

from the bottle”

for the enzymatic activity.

such treatment resulted activity,

(5

.

of the pH optimal

to use.

lipase

and therefore

in a significant this

enxyme was always

1610

15.

The seemingly high amount6 of lipases leading,

as the cozrzercial

reaction

was observed

pancreatic

lipase

reagent diethyl Desnuelle, ability

employed were very crude.

was irreversibly

p-nitrophenyl

inactivated

with the active

phosphate (Maylie,

to catalyze

All

17.

An Ultrastyragel

lipases

are insoluble

used for all hereafter

transesterification in the organic

used.

Waters Associates),

gel permeation BPLC experiments.

enzymatically

with polyethylene

cmrcial

starting

material,

2,5-dibromoadipic

and meso forms (Buchman, E.R.; 1942, 64, 2696-2700)

19.

Although specific 3-methyladipic for the free

optical acids

acids

Reims, A.O.; not leading

rotations

significant)

crolefin Chielini,

difficulties

E’S

encountered

of 2,5-dibrormadipic

4, p. 2117, respectively) for the enzymatically

that while we are not aware of studies

monomers with optically afford

polymerizations E.

of

in

and

active

low optical (Pino,

In Coordination

e.g.,

P.; Gschwald, A.;

Polymerization;

New

rather

produced oligomers.

It is

on asymmetric polycondensations

initiators,

purities,

Press:

suggests

polymerization less

processes

This work was financially Michel Therisod (Received

in

for helpful

Ciardelli,

F.;

Carlini,

C.;

Chien, J.C.W, Ed.; Academic Press:

supported by W.R. Grace & Co. discussions.

USA 13 October

1986)

of

than 10% in the case of

New York, 1975; pp. 25-72). 20.

(2)

N.J. J. Am. Chem.

of Organic Compounds; Oxford University

modest (although

that type usually

of a mixture

Schlatter,

are not known, comparison of our data with much higher values (Dictionary

2, p. 908 and vol.

of racemic

because the

consisted

T.;

of the

(see text).

to racemization.

for esters

York, 1965; vol. worth noting

acid,

Skei,

and because of serious

of the oligoesters

hydrolysis

excess of these esters,

and

of the

The composition

and pentamers was always confirmed by IR and NWRanalyses

We did not determine the enantiomeric

Sot.

glycols.

weights of

for the trimers

is presumably due to the approximation

formed oligoesters

(here and

polyethylene

The molecular

the enzymatic products were within 20% from those calculated

18.

packed in TBF, was

The column was calibrated

with molecular weights from 200 to 8,000 daltons. This difference

its

solvents.

‘IHF was employed as the mobile phase) using 8 different

pentamers. trimers

M.; Sarda, L.;

the enzyme lost

in organic

solvents

column (1000 A pore size,

center-specific

M.P.; Charles,

reactions

No appreciable

Furthermore, when porcine

in the absence of the enzyme.

P. Biochim Biophys. Acta 1969, 178, 196-198),

16.

glycols

used in these experiments are somewhat mis-

preparations

We are grateful

to Dr.