|Title of Invention||
A PROCESS FOR THE PREPARATION OF POLYMER USEFUL FOR THE CONVERSION OF ESTERS AND AMIDES TO CORRESPONDING ALCOHOLS AND AMINES
|Abstract||The present invention provides a process for the preparation of a new polymeric composition which exhibits catalytic activity similar to hydrolytic enzymes especially -chymotrypsin. The process of the present invention involves reacting vinyl monomers comprising functional groups present in the active site of a-chymotrypsin. The functional groups are brought together by complexation with a metal ion and an appropriate print molecule. The complex is then immobilized by polymerization in the presence of a cross-linker and a U.V. sensitive monomer. The resulting composition exhibits hydrolytic activity similar to -chymotrypsin "and is useful for the conversion of esters and amides to. the corresponding alcohols and amines. The composition offers additional advantages such as enhanced catalytic activity due to improved accessibility in the high surface area .polymeric catalyst, stability at elevated temperatures, ease of recovery from the reaction mixture, ability to withstand pH variations and ability to switch on-off the catalytic activity by photoregulation.|
|Full Text||The present invention relates to a process for the preparation of polymer useful for the conversion of esters and amides to corresponding alcohols and'amines.
Catalytic activity of hydrolytic enzymes in the hydrolysis of esters and amides is characterized by high reaction rates and high selectivity/ However, they undergo irreversible deactivation at higher temperatures, due to pH variations, shear etc. Since the enzymes are expensive, it is desirable to use them for repeated applications. But this is rendered difficult as the enzymes often lose their - catalytic activity during . their recovery from the reaction media. These disadvantages limit their industrial applications. One of the means to get around this problem, is to immobilize the enzymes on suitable polymeric supports- These approaches however cannot completely overcome the limitations of the enzymes. Also the efficacy is limited by the extent of 'immobilization that could be achieved.
Prior- art in this area describes polymers which in solution act as catalysts for the hydrolysis of esters and amides (R.Breslow. , Science/ 218, 53.2, 1982, J.Nh Lehn. ,. Angew. Chem. Int. Ed.- Engl . 27," 90, 1988, V.T. D'Souza, K. Hanabusa, T.O'Leavy, R.C. Gad-wood and MvL. Bender., Biochem. .Biophys. Res. Commun.
129, '727, 1988, H.J. Van den Berg and G. Challa in Syntheses and Separations using Functional Polymers.
Ed. B.C. Sherrington and P. Hodge, John Wiley & Sons, Chichester, 227, 1988). Unfortunately these polymers have very low activity and are difficult to recover from the reaction system.
Highly crosslinked polymers have also been used as catalysts for hydrolysis (A. Leonhardt and K. Mosbach., React. Polym. 6, 285 1987, B. Sellergren and K.J. Shea., Tetrahedron : Asymm 5, 1403, 1994, D.K. Robinson and K. Mosbach., J. Chem. Soc. "Chem. Commun. 969, 1989). These catalysts do not exhibit -chymo-trypsin like catalytic activity nor do they contain all the functional groups involved in the active site of -chymotrypsin.
It is a principal object of the present invention to provide a new polymer composition which is useful for the conversion of esters and ami'des to the corresponding alcohols and amines and which overcomes the disadvantages inherent in the polymers hitherto used in the prior art.
Another object of the present invention is to provide a process for the preparation of a polymer composition for the above mentioned purposes which eliminates the difficulties associated with low reactivity, and problems associated with the recovery of the polymer from.the reaction media and offer high
catalytic activity . and" selectivity for the hydrolysis reactions.
Yet another object of the present invention is to provide a process for the preparation of a polymer composition which -offers additional advantages such as enhanced activity due to improved accessibility, stability at higher temperatures, ability to withstand pH variations and enable switching on-off the activity of the polymer in response to a particular stimulus especially light.
As a result of our sustained research, we have been, able to ascertain that the activity of a polymer composition can be signi'ficantly enhanced when the functional groups present in the polymers are brought - in close proximity'by complexation in the presence of metal ion and a print molecule and are immobilized in . place by polymerization in .the -presence of a crosslinker on the internal surface of macroporous polymers. ' -
Accordingly the present invention provides a process for the preparation of polymer useful for the conversion of esters and amides to corresponding alcohols and amines which comprises
(a) forming a complex between a transition metal ion, a vinyl monomer with a hydroxyl group in the pendant chain,. a vinyl -monomer with a carboxyl group in the pendant chain, and a vinyl monomer with an imidazole group in the pendant chain and a print molecule "having a size and structure similar to the ester or amide substrate to be hydrolyzed and which can complex with the metal" ion in an
organic solvent, (b) immobilizing the complex on an inert support by
polymerizing the vinyl monomer either thermally at a
temperature of 55 to 80 C or in presence of UV light at a
- o .
temperature in the range^of 4 to 40 C with conventional
crosslinking monomer to obtain polymer.
In our copending application No. 109.5/Del/95 we have described and claimed a process for the conversion of esters and amides into their corresponding alcohols, and amines employing the polymer composition prepared by the process of the present invention.
According to a feature of the invention, the polymeric composition may be. prepared in the form of discs, dense microspheres or in the form of a thin film deposited on the internal surfaces of highly porous spherical - beads.
The functional monomers used in the process have
hydroxyl, carboxyl and imidazole groups linked to the
vinyl group through- the pendent link. The monomer .
bearing hydroxyl group may be selected from 2-
hydroxyethyl methacrylate, 2-hydroxypropyl methacrylate,
p-styrene phenol,; -2-hydroxyethyl methacrylamide, N-
methacryloyl serine, Methacryloyl 6-aminocaproyl
serine, p-vinyl benzoyl.-.. 6-aminocaproyl serine and the
like. The monomer' bearing imidazole group may be select
ed from N-methacryloyl L-histidine, N-acrylpyl. L-histi-.-
dine, 4(5)-vinyl imidazole, N-methacryloyl
histamine, methacryloyl 6-aminocaproyl
histidine, p-vinyl benzoyl 6-aminocaproyl histidine. The monomer bearing carboxyl group may be selected from acrylic acid, methacrylic acid, N-methacrloyl L-glutamic acid, N-methacryloyl aspartic acid, methacryl-oyl 6-aminocaproyl aspartic acid, p-vinyl benzoyl 6-aminocaproyl aspartic'acid.
The metal ion used for complex formation may be
+ + +.+ ++ + +
chosen from Ni , Co , Zn , Cu and similar transition metal ions. The individual composition of the monomers and the print molecule' in the complex can be varied. The solvent used for the formation of the complex may be chosen from methanol, ethanol, propa-nol, butanol.
The polymerization temperature used may be in the
o range of 4-40 C for U.V. and gamma irradiation and
55 C-75 C for thermal polymerization. The polymerization can be initiated thermally or by ultra violet light in the presence of a photoinitiator or by gamma irradiation. The inert atmosphere can be maintained using inert gases such as nitrogen or argon. The polymerization of the complex is carried out on the inner surface of a porous bead.
The inert polymer support used for the immobilization of the complex may be selected from poly (glycidyl methacrylate-ethyleneglycol dimethacrylate), poly (methylmethacrylate - ethyleneglycol dimethacrylate) , poly (styrene-divinylbenzene), poly (trimethylol propane trimethacrylate - glycidyl methacrylate), poly
(trimethylol propane trimethacrylate- acrylonitrile), poly (trimethylol propane triacrylate-glycidyl metha-crylate), poly (trimethylol propane triacrylate-acrylonitrile) macroporous particles.
The initiator used for polymerization may be chosen from azobisisobutyronitrile, t-butyl hydrop-eroxide, benzoyl peroxide and the like. The period of polymerization employed may range from 3-24 hours.
.The metal ion can be extracted by dilute acid, ethylene diamine tetra acetic acid solution or by treating the polymers with methanol solution containing 2,2' bipyridyl or 1,10 phenanthroline.
The surface area of the macroporous supports can be
2 varied from 50-800 m ,/g. The spheres can be in the
size range of 35-150 u.
The composition is brought in contact with the ester or amide to be hydrolyzed, in a mixed solvent system, at ambient temperature, in the presence or absence of U.V. light so as to bring about hydrolysis of the amide or alcohol till the reaction is complete.
.The hydrolysis of the amide and ester can be carried out in mixed solvent systems comprising ethanol-buffer, acetonitrile-buffer, dioxane-buffer. The composition of the organic solvent in the mixture can be in the range 10% to 50% and a preferable range is from 20% to 40%. The catalyst to substrate ratio in the reaction can be varied from 1 : 0.05 to 1 : 50.
The pH of the reaction mixture can be in the range pH 6-11 and a preferable range is from pH . 7.5-8.0. The
hydrolysis of amides or esters can be effected in the
o temperature range 20 G-80oC and a preferable range is
from 25 O C-40O C. The hydrolysis of the ester and amides
can be controlled by U.V. light and pH.
The invention will now be described in greater details in the following examples which are provided to illustrate the invention and should not therefore be construed to limit the scope of the present invention.
0.286 gm (0.0022 M) 2-hydroxyl ethyl methacry-late, 0.189 gm (0.0022 M) methacrylic acid, 0.5 gm (0.0022 M) N-methacryloyl L-histidine and 0.932 gm (0.0022 M) N-isobutyryl 6-amino caproyl L-phenyl alanyl 2-amino pyridine were taken-in a 25 ml beaker.
5 ml methanol and 0.524 gm (0.0022 M) CoCl .6H O were
added andv the mixture was stirred for one hour at room temperature.. Methanol was then evaporated under
reduced pressure to give a deep blue coloured
2.5 gm of this complex was taken in a glass test tube. To it 2 gm 2-hydroxyethyl methacrylate, 0.5 ml ethylene glycol dimethacrylate and 0.8 ml t-butyl hydroperoxide were added. The test tube was purged with nitrogen for 10 minutes and then immersed in a
water bath maintained at 65oC. The polymerization was carried out for 16 hours. The polymer was isolated in the form of a cylindrical rod by breaking the test tube, The rod was then cut into discs of 0.09-0.11 cm thickness on a lathe. The yield obtained was 4 gm.
These discs were- dipped into a 1% ~2,2' bipyri-
dyl solution in methanol for 12 hours to remove
+ + Co and the print molecule. The solution of 2,2'
bipyridyl was changed after every 4 hrs. Further the discs were washed with dilute HC1 and dried in a vacuum oven at room, temperature for 48 hours.
The " substrate N-methacryloyl 6-aminocaproyl L-phenylalanyl p-nitrophenol was sorbed into the discs
from an acetone solution. It was then polymerized by
60 exposing the discs to gamma irradiation from a Co
source of 0.25 Mrad/hr for 6 hours. The unreacted monomer was extracted in acetone. The concentration of the catalytic groups based on the moles of imidazole was 2 x 10-5 M/gm of polymer.
0.286 gm (0.0022 M) 2-hydroxyethyl methacrylate, 0.189 gm (0.0022 M) methacrylic acid, 0.5 gm (0.0022 M) N-methacryloyl L-histidine, 0.932 gm (0.0022 M) isobu-tyryl 6-amirio caproyl L-phenylalanine 2-amino pyridine
were mixed well in 5 ml methanol "in a' 25 ml beaker.
0.524 gm (0.0022 M) CoCl .6H O was added and the
solution was stirred for 1 hour at room temperature to give a blue coloured complex. Methanol was evaporated under reduced pressure. 2 gm 2-hydrox-yethyl methacrylate, 0.3 ml ethylene glycol dimethacry-late and 0.030 gm azobisisobutyronitrile were further added to the mixture and air was expelled from the mixture using nitrogen. In a three necked round bottom flask equipped with a stirrer, 47 ml . 35 % NaCl' solution was added. To this, 3 ml IN NaOH and 2
gm MgCl 2.6H2 O were added to give a uniform suspen-
sion of Mg(OH)2. A stream of nitrogen was passed through the solution for 15 minutes. The monomer
mixture, .was added dropwise to the suspension and
o stirred at 1000 rpm at . 75 C for 3 hours. The monomer
mixture was dispersed into fine droplets which were then polymerized into rigid spheres. The spheres were isolated by filtration. The yield obtained.was 3.8 gm. The microspheres were subjected to repeated extraction with 1 % methanolic- solution of 2,2' bipyridyl and dilute HC1, followed by drying in a vacuum oven at room temperature for 48 hrs. The concentration of the catalytic groups based on the number of moles of imidazole was found to be 2.5 x 10-5 M/gm.
0.286 gm (0.0022 M) 2-hydroxyethyl methacrylate', 0.189 gm (6.0022 M) methacrylic acid, 0.5 gm (0.0022 M) N-methacryloyl L-histidine and 0.932 gm (0.0022 M) isobutyryl 6-aminocaproyl-L-phenylalanine-2-amino pyridine were mixed together in 5 ml methanol. 0.524
gm (0.0022 M) CoCl . 6H O was added and stirred
well for one hour to give a blue complex. Methanol was evaporated under reduced pressure. Further 0.3 gm 2-methacryloyl hydroxy ethyl p-phenyl azobenzo-ate, 1.7'gm 2-hydroxyethyl methacrylate, 0.3 ml ethyle-neglycol dimethacrylate and 0.030 gm azobisisobutyro-nitrile were added and the mixture was purged with nitrogen for 10 minutes.
In a three necked round bottom flask equipped with a stirrer, 47 ml 35 % NaCl solution was added.
To this, 3 ml IN NaOH and 2 gm MgCl 2.6H2 O were added
to give a uniform suspension of Mg(OH) . This mixture
was purged with nitrogen for 15 minutes. The
monomer mixture was added dropwise to the Mg(OH)
2 o suspension.. The temperature was maintained at 75 C
and the mixture was stirred at 1000 rpm for 3 hrs. Spheres were isolated by filtration. The yield obtained was: 3.5 gm.
The spheres were subjected to repeated extraction with 1% 2,2' bipyridyl solution in methanol and then with dilute HC1 for ,12 hrs followed by drying in vacuum oven at room temperature for 48 hrs. The catalytic group concentration was: found to be 2.5 x 10-5 M / gm.
3.6 gm of glycidyl methacrylate, 8.4 gm ethyleneg-lycol dimethacrylate and 0.120 gm azobisisobutyroni-trile were mixed with 16 gm cyclohexanol and suspended into a solution of 88 ml 1% polyvinylpyrrolidone
5 (M.W. 3.6 x 10 ) in, a 100 ml round bottomed flask.
Air was expelled from the reaction vessel by a
stream of nitrogen. Stirring rate was adjusted to
o 1000 rpm and the polymerization was performed at 70 C
o for 2 hrs. and at 80 C for 6 hrs. The mixture was
a'llowed to cool for 2 hrs. The spheres were isolated
by filtration and washed with water and alcohol
o repeatedly and dried in vacuum oven at 40 C for 48
hrs. Spheres in the size.range 37-45 u were chosen.
0.286 gm (0.0022 M) 2-hydroxyethyl methacrylate, 0.189 gm (0.0022 M) methacrylic acid, 0.5 gm (0.0022 M) N-methacryloyl L-histidine, 0.932 gm (0.0022 M) isobu-tyryl 6-aminocaproyl-L-phenylalanine-2-aminopyridine, 0.1 ml ethylene glycol dimethacrylate, 0.524 gm
(0.0022M) CoCl2 .6H2 O and 0.1 gm azobisisobutyronitrile
were mixed together in 5 ml methanol. 1 gm of macroporous spheres was added to this and stirred for 24 hours and filtered. , The monomer mixture that was
adsorbed on the internal surface of the spheres was
o then polymerized thermally at 75 C for 24 hrs.
The spheres were subjected to repeated extraction with methanolic solution of 2,2' bipyridyl and dilute HCl followed by drying in vacuum oven at room temperature for 48 .hrs. The catalytic group concentration in the surface- imprinted polymer was found to be 2 . 1 x 10-4 M / gm.
The complex .comprising 2-hydroxyethyl methacrylate (0.0022 M) , methacrylic acid (0.0022 M) , N-methacryloyl L-histidine (0.0022 M) , N-isobutyryl 6-aminocaproyl L-
phenylalahyl 2-aminopyridine (0.0022 M), and CoCl 2.6H2 0
(0.0022 M), with 0.5 ml ethylene glycol dimethacrylate and 0.125 gm azobisisobutyronitrile was adsorbed on the surface of 1 gm poly (glycidyl methacrylate-ethylene
glycol dimethacrylate).microspheres. The polymerization
o was initiated by. U.V. light at 4 C for 6 .hours. The
spheres were subjected to repeated extraction with 1 % 2,2' bipyridyl solution in methanol and with dilute HCl followed by drying in a vacuum oven at room temperature for '48 hours. The active site concentration based on the imidazole was 1.8 x 10-6 M / gm.
We claim :
1. A process for the ,preparation of polymer -useful for the conversion of esters and amides to corresponding alcohols and amines which comprises : .
(a) forming a complex between a transition metal ion, a
vinyl monomer with a hydroxyl group in the pendant chain,
a vinyl monomer with a carboxyl group in the pendant
chain, and a vinyl monomer with an imidazole group in the
pendant chain and:a. print molecule having a size and
structure . similar to the ester or amide substrate to be
hydrolyzed and which can complex with the metal ion in an
(b) immobilizing .the complex on an inert support by
polymerizing the vinyl .monomer either .thermally at a
temperature of 55 to 80 C or in presence of UV light at a
temperature . in the range of 4 to 40 C with conventional
cros-sl inking monomer to obtain polymer.
2 . A process as claimed in claim 1 wherein at least, one of the vinyl monomer contain the hydroxyl,. carboxyl, or imidazole group as a terminal group in the pendant chain.
3. A process as claimed, in claims 1-2 .wherein the . hy droxyl group bearing vinyl monomer is selected from .the group consisting - of 2-hydroxyl ethyl methacrylate, 2-
hydroxyl propyl methacrylate, p-styrene phenol, N-methacryolyl serine, 2 hydroxyl.ethyl methecrylane N-methacryolyl,serine, methecryloyl 6-aminocaproyl serine, and p-vinyl benzoyl 6-aminocaproyl serine.
4. A process as claimed in claims 1-3 wherein the
imidazole group bearing vinyl monomer is selected from
the group consisting of N-methacryloyl L-histidene, N-
acryloyl L-histidine, 4(5) vinyl imidazole, N-methacryl
oyl histamine, methacryloyl 6-aminocaproyl histidine and
p-vinyl benzoyl .6-ami.ne caproyl hist idine.
5. A process as claimed in claims - 1-4 wherein the
carboxyl group bearing vinyl monomer is selected from the
group consisting of acrylic acid, methacrylic acid, N-
methacryloyl . L-glutamic " acid, N-methacryloyl aspartic
acid, methacryolyl 6-aminocaproyl aspartic acid and p-
vinyl benzoyl 6-aminocaproyl aspartic acid,
6. A process as claimed in claims 1-5. wherein the metal
ion is selected from-the group consisting of Ni ., Co ,
Zn , Cu , Mg++ , Mn++ &' Fe++
7. A process as claimed in claims 1-6 wherein the com
plex is formed in a solvent selected from the group
consisting of ethanol,. ..methanol, propanol, butanol,
aetetonitrile, ethyl acetate, dioxine and dichlorometh-
8. A process as claimed.in claims 1-7 wherein the
cros.slinking monomer is selected from the group consist
ing of ethylene glycol dimethacrylate, triethylene gly-
col, dimethacrylate, divinyl benzene, trimethyloyl pro
pane trimethacrylate, N,N-bisacrylamide and trimethylol
9. A process, for the preparation of polymer useful for
the conversion of esters and amides to corresponding
alcohols and amines substantially as herein described
with reference to the examples.
|Indian Patent Application Number||1094/DEL/1995|
|PG Journal Number||12/2013|
|Date of Filing||14-Jan-1995|
|Name of Patentee||COUNCIL OF SCIENTEFIC AND INDUSTRIAL RESEARCH|
|Applicant Address||RAFI MARG, NEW DLEHI-110001, INDIA.|
|PCT International Classification Number||C08L 33/08|
|PCT International Application Number||N/A|
|PCT International Filing date|