Enzyme catalyzed method of forming organosilicon esters

The present invention relates to the formation of organosilicon esters using an enzymatic route.

Ester and polyester compounds may be synthesized with an acid or base catalyst at high temperatures for long periods. Although these reaction conditions favor the equilibrium of polycondensation, they also promote uncontrolled side reactions, redistribution of monomers sequences, cross linking, and broad molecular weight distributions. Additionally, the typical acid and base catalysts are not regioselective and may catalyze esterification at all reactive groups on a polyfunctional monomer. The acid and base catalysts may cause the decomposition of potentially useful functional groups, such as epoxy groups, and bonds, such as siloxane bonds. Thus, the ability to control the material structure is essentially lost.

The patent <patcit id="pcit0001" dnum="US6359097B"><text>US6359097</text></patcit> discloses functionalized polyorganosiloxanes and a process for the preparation thereof by chemical synthesis starting from a polyhydroorganosiloxane. Among the compounds disclosed are also functionalized polyorganosiloxanes having a cyclic structure.

The patent <patcit id="pcit0002" dnum="US6288129B"><text>US6288129</text></patcit> discloses a process for preparing acrylic esters and/or methacrylic esters of hydroxy-functional siloxanes and/or polyoxyalkylene-modified siloxanes and their use. The process comprises the enzymatic esterification of hydroxy-functional siloxanes and/or polyoxyalkylene-modified siloxanes.

Traditional methods of synthesizing amide compounds may not be selective. For example, the methods may not be regioselective and/or enantioselective. Therefore, methods of synthesizing amides may not provide the ability to control the material structure.

Thus, the need remains in the relevant art for improved methods of forming structurally defined organosilicon esters and amides, and the need remains in the relevant art for more structurally defined organosilicon esters and amides.

In accordance with the present invention, a method of forming an organosilicon ester is provided as described by the claims.

The method comprises contacting a hydrolase enzyme selected from an esterase, lipase, protease and combinations thereof with a cyclic organosilicon reactant and an organic reactant, wherein: <ul id="ul0001" list-style="none" compact="compact"><li>said cyclic organosilicon reactant comprises a cyclic organosilicon with siloxane bonds having the formula <chemistry id="chem0001" num="0001"><img file="EP1546159B1_D0001.tif" /></chemistry> wherein: <ul id="ul0002" list-style="none" compact="compact"><li>each R is independently selected from alkyl, haloalkyl, unsaturated alkyl, aryl, hydroxy, alkoxy, hydrogen, -(OSiR₂)_x-OSiR₃, or R';</li><li>at least one of R=R';</li><li>x is 0 or greater than 0;</li><li>y is equal to or greater than 3; and</li><li>R' is: <chemistry id="chem0002" num="0002"><img file="EP1546159B1_D0002.tif" /></chemistry> wherein: <ul id="ul0003" list-style="none" compact="compact"><li>A is a substituted or unsubstituted hydrocarbon substituent, wherein said hydrocarbon may be substituted such that said hydrocarbon comprises a halogen-, ether-, alkoxy-, phenyl-, or unsaturated-functional hydrocarbon and combinations thereof;</li><li>R" is independently carboxylic acid, ester, amide, or alcohol; and</li><li>m is 0 or greater than 0;</li></ul></li><li>said organic reactant is selected from the group consisting of hexanol, 2- hydroxylethylmethacrylate, hexanediol, polyethylene glycol, adipic acid, octanoic acid, phenyl acetic acid methyl ester, and dimethyl adipate; and said enzyme catalyzes the formation of an ester bond between carboxylic acid, ester, or amide functional groups of said organosilicon reactant or said organic reactant and alcohol functional groups of said organosilicon reactant or said organic reactant to form said organosilicon ester; wherein the method is carried out under reaction conditions that do not cleave the siloxane bonds of the cyclic organosilicon reactant.</li></ul></li></ul><ul id="ul0004" list-style="none" compact="compact"><li><figref idref="f0001">Fig. 1</figref> is a ¹³C DEPT NMR spectrum of the esterification of Example 1.</li><li><figref idref="f0002">Fig. 2</figref> is a GC MS of the esterification of Example 1.</li><li><figref idref="f0003">Fig. 3</figref> is a ¹H NMR spectrum of the esterification of Example 2.</li><li><figref idref="f0004">Fig. 4</figref> is an ESI MS of the esterification of Example 2.</li><li><figref idref="f0005">Fig. 5</figref> is a ¹H NMR spectrum of the esterification of Example 3.</li><li><figref idref="f0006">Fig. 6</figref> is an ESI MS of the esterification of Example 3.</li><li><figref idref="f0007">Fig. 7</figref> is a ¹H NMR spectrum of the esterification of Example 4.</li><li><figref idref="f0008">Fig. 8</figref> is ¹H NMR spectrum of the esterification of Example 6.</li><li><figref idref="f0009 f0010">Figs. 9-10</figref> are ¹³C DEPT NMR spectrum of the esterification of Example 8.</li></ul>

The present invention relates to the reaction of a cyclic organosilicon reactant and an organic reactant in the presence of a hydrolase enzyme to form organosilicon esters. The reaction involves the formation of an ester bond between at least one carboxylic acid, ester, or amide functional group and at least one alcohol functional group, as shown in the reaction sequences below: <chemistry id="chem0003" num="0003"><img file="EP1546159B1_D0003.tif" /></chemistry><chemistry id="chem0004" num="0004"><img file="EP1546159B1_D0004.tif" /></chemistry><chemistry id="chem0005" num="0005"><img file="EP1546159B1_D0005.tif" /></chemistry>

In the above reactions, X and Y represent a portion of a cyclic organosilicon or an organic compound other than the carboxylic acid, ester, alcohol, or amide functional groups. It will be understood that the ester or amide functional groups are not limited to the methylated ester or amide as shown in the above reactions. Rather, the ester or amide functional group may have any desired group substituted for the methyl group shown.

The cyclic organosilicon reactant is a cyclic organosilicon species. The cyclic organosilicon reactant has at least three siloxane bonds. The cyclic organosilicon reactant has at least one carboxylic acid, ester, alcohol, or amide functional group, and the cyclic organosilicon reactant may be monofunctional or polyfunctional. The carboxylic acid, ester, alcohol, or amide functional groups may be pendant, terminal, or be in any other suitable location. However, the carboxylic acid, ester, alcohol, or amide functional groups may not be directly bonded to the silicon. Suitable cyclic organosilicon reactants having at least three siloxane bonds correspond to Formula (II), as shown below: <chemistry id="chem0006" num="0006"><img file="EP1546159B1_D0006.tif" /></chemistry> wherein: <ul id="ul0005" list-style="none" compact="compact"><li>each R is</li><li>independently selected from alkyl, haloalkyl, unsaturated alkyl, aryl, hydroxy, alkoxy, hydrogen, -(OSiR₂)_x-OSiR₃, or R';</li><li>at least one R=R';</li><li>x is 0 or greater than 0;</li><li>y is equal to or greater than 3; and</li><li>R' is: <chemistry id="chem0007" num="0007"><img file="EP1546159B1_D0007.tif" /></chemistry> wherein: <ul id="ul0006" list-style="none" compact="compact"><li>A is a substituted or unsubstituted hydrocarbon substituent, wherein said hydrocarbon may be substituted such that said hydrocarbon comprises a halogen-, ether-, alkoxy-, phenyl-, or unsaturated-functional hydrocarbon and combinations thereof;</li><li>R" is independently carboxylic acid, ester, amide, or alcohol; and</li><li>m is 0 or greater than 0.</li></ul></li></ul>

In accordance with an embodiment of the present invention, x may be between 0 to about 250. The organosilicon reactant may be selected such that y is between about 3 to about 6. The organosilicon reactant may be selected such that A is a C₃ to C₂₀ hydrocarbon, and m may be between 0 and about 250.

It will be understood by those skilled in the art that the portion of the organosilicon structure shown within the brackets of Formula (II) do not have to be <u>identical</u> repeating units. Rather, the R groups may be independently chosen for each of the repeating units. It will be understood that alkyl, haloalkyl, unsaturated alkyl, alkoxy, and hydrocarbon may be substituents having one carbon or more than one carbon. It will be further understood that when A is a substituted hydrocarbon the substitutions may be in the middle of the carbon chain or pendant on the carbon chain. However, the substitution will not generally be terminal on the end of the carbon chain next to R". The organosilicon reactant of Formula (II), is an acceptable substrate that allows the hydrolase enzyme to selectively catalyze the esterification reaction at the reactive sites of the organosilicon reactant. The reactive sites of the organosilicon reactant comprise the carboxylic acid, ester, amide, or alcohol functional groups.

Organic reactants useful in the present invention include hexanol, 2-hydroxyethylmethacrylate, hexanediol, polyethylene glycol, adipic acid, octanoic acid, phenyl acetic acid methyl ester, and dimethyl adipate.

Generally, the at least one functional group of the organic reactant comprises an alcohol functional group when at least one of R" of Formula II comprises carboxylic acid, ester, or amide. The at least one functional group of the organic reactant comprises a carboxylic acid functional group or ester functional group, when at least one of R" of Formula II comprise alcohol. It will be understood that the organic reactant may have more than one functional group, and it will be further understood that the functional groups may be the same functional group or different functional groups.

In accordance with an embodiment of the present invention, the organic reactant may be in a liquid form, and a liquid-to-liquid reaction may occur between the organic reactant and the organosilicon reactant. Thus, the reaction may occur in bulk. In accordance with another embodiment of the present invention, the organic reactant or organosilicon reactant may be a solid and the reaction may be between a liquid and a solid. When the organic reactant or organosilicon reactant is in the form of a solid, the reactions of the present invention may occur on the surface of the solid. Thus, the surface of the solids may be modified.

The organosilicon reactant and the organic reactant are contacted with a hydrolase enzyme in order to catalyze the formation of the ester bond. It will be understood that the organosilicon and organic reactant may be contacted with the hydrolase enzyme sequentially. The hydrolase enzyme is generally an esterase, lipase, or protease and combinations thereof, and the hydrolase enzyme is more generally lipase or protease. The hydrolase enzyme may be derived from a bacterial, fungal, or mammalian source, or the hydrolase enzyme may be derived from any other suitable source. For example, the hydrolase enzyme may be Novozyme 435® (N435) lipase available from Novozyme (Bagsvaerd, Denmark), <i>Candida cylindricia</i> lipase type VII (CCL) available from Sigma (St. Louis, MO), porcine pancreatic lipase (PPL) available from Sigma (St. Louis, MO), and protease enzymes such as subtilisin and papain available from Sigma (St. Louis, MO). The enzyme is generally present as a heterogeneous suspension, and the enzyme may be lyophilized or immobilized.

The hydrolase enzyme catalyzes the formation of an ester bond between a carboxylic acid, ester, or amide functional group and an alcohol functional group in a one-step reaction. Thus, the enzyme catalyzes the formation of an ester bond between carboxylic acid, ester, or amide functional groups of the organosilicon reactant or the organic reactant and alcohol functional groups of the organic reactant or the organosilicon reactant to form the organosilicon ester.

The organosilicon ester may be a structurally defined compound of the formula: <chemistry id="chem0008" num="0008"><img file="EP1546159B1_D0008.tif" /></chemistry> wherein: <ul id="ul0007" list-style="none" compact="compact"><li>each R¹ is independently selected from alkyl, haloalkyl, unsaturated alkyl, aryl, hydroxy, alkoxy, hydrogen, -(OSiR¹₂)_x-OSiR¹₃, or R"';</li><li>at least one of R¹=R''';</li><li>y is equal to or greater than 3; and</li><li>R''' is: <chemistry id="chem0009" num="0009"><img file="EP1546159B1_D0009.tif" /></chemistry> or <chemistry id="chem0010" num="0010"><img file="EP1546159B1_D0010.tif" /></chemistry> wherein: <ul id="ul0008" list-style="none" compact="compact"><li>A is a substituted or unsubstituted hydrocarbon substituent, wherein said hydrocarbon may be substituted such that said hydrocarbon comprises a halogen-, ether-, alkoxy-, phenyl-, or unsaturated-functional hydrocarbon and combinations thereof;</li><li>R² is an organic compound; and</li><li>m is 0 or greater than 0.</li></ul></li></ul>

It will be apparent to those having skill in the art that more than one functional group may be present on an organosilicon reactant and/or on an organic reactant. Therefore, organosilicon ester monomers, macromers, and polymers may be formed using the enzymatic method of the present invention. For example, the polymers may be of the -AABB- variety wherein AA represents an organosilicon reactant having two functional groups (AA) and BB represents an organic reactant having two functional groups (BB) that react to form ester bonds. Alternatively, the polymers may be of the terpolymer type or any other suitable type of polymer. Additionally, the polymers may be crosslinked in any suitable manner.

The hydrolase enzyme selectively catalyzes the formation of the ester bonds. For example, the hydrolase enzyme may be regioselective or enantioselective, or combinations of both. Thus, structurally defined organosilicon esters may be formed in accordance with the present invention.

When an interfacial enzyme is used, the organosilicon reactants appear to enhance the rate of esterification catalyzed by the enzyme in comparison to the rate of esterification catalyzed by the enzyme when organic materials are used in place of organosilicon materials. Lipase is an example of an interfacial enzyme. Thus, the use of organosilicon reactants may be advantageous in forming organic materials containing ester bonds.

The reactions are generally carried out under mild reaction conditions. The temperature of the reactions is generally between about 20°C and 100°C, and the reaction is more generally carried out at a temperature of between about 40°C to about 70°C. No acid or base catalysts are required. The reactions may be performed under solventless (neat) conditions, or the reactions may be performed utilizing a solvent. Suitable solvents include, but are not limited to, hexane, toluene, xylene, and other hydrophobic alkyl or aromatic organic solvents.

The ability to synthesize organosilicon esters under mild reaction conditions is advantageous because the mild reaction conditions do not cleave the siloxane bonds of the siloxane reactants. Therefore, novel organosilicon esters may be synthesized. Additionally, the various functional groups of the organosilicon and organic reactants may be chosen to introduce a desired functionality into the resulting organosilicon ester. For example, a hydrophobic organosilicon and a hydrophilic organic may be chosen such that the resulting organosilicon ester is an amphiphilic molecule having desired surfactant properties. Alternatively, phenol functional groups may be chosen and then selectively coupled after the formation of the organosilicon ester to synthesize an electrically conductive organosilicon ester. Fibers, films, coatings, gels and other materials may be synthesized using this enzymatic route having desired properties such as, for example, adhesiveness and the ability to self assemble.

In order that the invention may be more readily understood, reference is made to the following examples.

Example 1

Novozyme 435 (N435), <i>Candida cylindricia</i> (CCL), and porcine pancreatic (PPL) lipase were used to catalyze the esterification of trimethylsilylpropionic acid with hexanol as shown in the following diagram: <chemistry id="chem0011" num="0011"><img file="EP1546159B1_D0011.tif" /></chemistry> The reactions were conducted with constant stirring (i.e. magnetic stir bar) in a two-neck round-bottom flask attached to a Dean-Stark trap and a water-cooled condenser in a heated oil bath. The reactions were performed in refluxing hexane (i.e. 70°C pot, 90°C bath temperature). The organic reactant was added to the organosilicon-solvent solution at 70°C. After homogenization, the dried enzyme was added to the reaction mixture and the reactions were conducted for 24 hours. The reactions were formulated with organosilicon:organic and solvent:monomer mole ratios equal to 1:1 and 10:1, respectively. The enzyme:monomer weight ratio was 1:10. The lipases formed heterogeneous suspensions in the stirred organic mixtures. In the presence of the Dean-Stark trap, the formation of a hexane-water azeotrope further promoted the esterification reactions.

After the reaction, chloroform (∼10 mL) was added to the mixture to remove the contents from the flask and filter out the enzyme. Subsequently, the filtrate was evaporated on a rotary evaporator and dried in a vacuum oven overnight at ∼45°C in order to isolate the product. The reactions were characterized by ¹H and ¹³C nuclear magnetic resonance spectroscopy (NMR), gas chromatography-mass spectrometry (GC-MS), gas chromatography - flame ionization detection (GC-FID), and electrospray mass spectrometry (ESI MS). <figref idref="f0001">Fig. 1</figref> illustrates the ¹³C NMR of the esterification. <figref idref="f0002">Fig. 2</figref> illustrates the GC-MS of the esterification. Based on the analyses, lipase was observed to catalyze the esterification reaction. Comparatively, N435 was determined to be more active than CCL or PPL. The materials were observed to contain a mixture of products and reactants.

Example 2

N435, CCL, and PPL were used to catalyze the esterification of 1,3-bis(3-carboxypropyl)tetramethyldisiloxane (i.e., diacid disiloxane) with hexanol as shown in the following diagram: <chemistry id="chem0012" num="0012"><img file="EP1546159B1_D0012.tif" /></chemistry>

The reactions were conducted with constant stirring (i.e. magnetic stir bar) in a two-neck round-bottom flask attached to a Dean-Stark trap and a water-cooled condenser in a heated oil bath. The reactions were performed in refluxing hexane. The organic reactant was added to the organosilicon-solvent solution at 70°C. After homogenization, dried lipase enzyme was added to the reaction mixture and the reactions were conducted for 24 hours. The reactions were formulated with organosilicon:organic and solvent:monomer mole ratios equal to 1:1 and 10:1, respectively. The enzyme:monomer weight ratio was 1:10. In the presence of the Dean-Stark trap, the formation of a hexane-water azeotrope further promoted the esterification reactions.

After the reaction, chloroform (∼10 mL) was added to the mixture to remove the contents from the flask and filter out the enzyme. Subsequently, the filtrate was evaporated on a rotary evaporator and dried in a vacuum oven overnight at ∼45°C in order to isolate the product. The reactions were characterized by ¹H and ¹³C NMR, GC-MS, GC-FID, and ESI MS. <figref idref="f0003">Fig. 3</figref> illustrates the ¹H NMR of the esterification. <figref idref="f0004">Fig. 4</figref> illustrates the ESI MS of the esterification. Based on the analyses, lipase was observed to catalyze the esterification reaction. Comparatively, N435 was determined to be more active than CCL or PPL. The materials were observed to contain a mixture of products and reactants.

Example 3

N435 was used to react the diacid disiloxane of Example 2 with 2-hydroxyethylmethacrylate (HEMA) in refluxing hexane or neat at 70°C for 1 hour as shown in the following diagram: <chemistry id="chem0013" num="0013"><img file="EP1546159B1_D0013.tif" /></chemistry>

The reactions were conducted with constant stirring (i.e. magnetic stir bar) in a two-neck round-bottom flask attached to a Dean-Stark trap and a water-cooled condenser in a heated oil bath. The reactions were formulated with organosilicon:organic and solvent:monomer mole ratios equal to 1:2 and 10:1, respectively. The enzyme:monomer weight ratio was 1:10. In addition to the Dean-Stark trap, molecular sieves (i.e. sieve:enzyme weight ratio = 2:1) were used to adsorb water in order to further promote the esterification reaction.

After the reaction, chloroform (∼10 mL) was added to the mixture to remove the contents from the flask and filter out the enzyme. Subsequently, the filtrate was evaporated on a rotary evaporator and dried in a vacuum oven overnight at ∼45°C in order to isolate the product.

In hexane and neat media, 82% and 91 % of the HEMA was consumed within 1 hour, respectively. In the presence of molecular sieves, aliquots of the reaction were analyzed by ¹H NMR every 5 minutes. Based on the spectral data, HEMA was nearly consumed within the first 5 minutes. After 1 hour, the neat reaction was filtered and dried to isolate a crude oily product. The product was characterized by ¹H and ¹³C NMR, ESI MS, Fourier transform infrared spectroscopy (FTIR), and gel permeation chromatography-refractive index detection (GPC-RI). <figref idref="f0005">Fig. 5</figref> illustrates the ¹H NMR of the esterification. <figref idref="f0006">Fig. 6</figref> illustrates the ESI MS of the esterification. Based on the molecular characterization, the material was observed to contain a mixture of mono- and diester products band reactants.

Example 4

An organosilicon polyester was synthesized by reacting a diacid disiloxane (AA) and hexanediol (BB) with N435 in refluxing hexane at 70°C for 6 days as shown in the diagram below: <chemistry id="chem0014" num="0014"><img file="EP1546159B1_D0014.tif" /></chemistry>

The reaction was conducted with constant stirring (i.e. magnetic stir bar) in a two-neck round-bottom flask attached to a Dean-Stark trap and a water-cooled condenser in a heated oil bath. The reaction was formulated with organosilicon:organic and solvent:monomer mole ratios equal to 1:1 and 10:1, respectively. The enzyme:monomer weight ratio was 1:10. The use of the Dean-Stark trap further promoted the esterification reaction.

The progress of the reaction was monitored by ¹H NMR. After 6 days, the mixture was filtered, evaporated, and dried to isolate an oily product. The material was characterized by ¹H, ¹³C, and ²⁹Si NMR, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), FTIR, GPC-RI, thermal gravimetric analysis (TGA), and differential scanning calorimetry (DSC). <figref idref="f0007">Fig. 7</figref> illustrates the ¹H NMR of the esterification.

The material was observed to contain, primarily, linear as well as cyclic -[AABB]_x- organosilicon polyesters. No residual diacid disiloxane was observed. Based on the GPC-RI results, the Mn and Mw values were 5140 and 15710, respectively, with a 3.1 polydisperity. Based on the thermal analysis, the material experienced a critical mass loss at 381°C vs. 164°C (i.e. diacid disiloxane reactant). The Tg was measured to be -80°C. In comparison to the diacid disiloxane (Tg = -76°C), similar amounts of energy were required to achieve molecular motion. During the DSC heating cycles, crystalline phases (Tm) were observed at -47°C &amp; -38°C (0.7 J/g), and 92°C (0.9 J/g). A cold crystallization (Tcc = -15°C, 50.4 J/g) and two crystalline phases (Tm = 30°C, 45 J/g; 46°C, 8.6 J/g) were detected in the diacid disiloxane.

Example 5

An organosilicon polyester was synthesized by reacting a diacid-endblocked polydimethylsiloxane (diacid PDMS, AA) and polyethylene glycol (BB) with N435 in refluxing hexane at 70°C for 6 days as shown in the following diagram: <chemistry id="chem0015" num="0015"><img file="EP1546159B1_D0015.tif" /></chemistry>

The reaction was conducted with constant stirring (i.e. magnetic stir bar) in a two-neck round-bottom flask attached to a Dean-Stark trap and a water-cooled condenser in a heated oil bath. The reaction was formulated with organosilicon:organic and solvent:monomer mole ratios equal to 1:1 and 10:1, respectively. The enzyme:monomer weight ratio was 1:10. The use of the Dean-Stark trap further promoted the esterification reaction.

The progress of the diacid PDMS esterification was monitored by ¹H NMR. Based on the spectral data, N435 was observed to catalyze the formation of ester bonds (i.e., -[AABB]_x-) with large polymeric substrates over 4 days.

Example 6

N435 was used to catalyze the polyesterification of a carbinol-endblocked polydimethylsiloxane (diol PDMS, BB) and adipic acid (AA) in refluxing hexane at 70°C for 6 days as shown in the following diagram: <chemistry id="chem0016" num="0016"><img file="EP1546159B1_D0016.tif" /></chemistry>

The reaction was conducted with constant stirring (i.e. magnetic stir bar) in a two-neck round-bottom flask attached to a Dean-Stark trap and a water-cooled condenser in a heated oil bath. The reaction was formulated with organosilicon:organic and solvent:monomer mole ratios equal to 1:1 and 10:1, respectively. The enzyme:monomer weight ratio was 1:10. The use of a Dean-Stark trap further promoted the esterification reaction.

The progress of the diol PDMS esterification was monitored by ¹H NMR. After 4 days of reaction, chloroform (∼10 mL) was added to the mixture to remove the contents from the flask and filter out the enzyme. Subsequently, the filtrate was evaporated on a rotary evaporator and dried in a vacuum oven overnight at ∼45°C. in order to isolate the product. The material was characterized by ¹H NMR and FTIR. <figref idref="f0008">Fig. 8</figref> illustrates the ¹H NMR of the esterification. Based on the spectral data, N435 was observed to catalyze the formation of ester bonds (i.e.,-[BBAA]_x-) with a carbinol-functional PDMS substrate.

Example 7

N435 was used to catalyze the polyesterification of the diol PDMS (BB) and an acid-endcapped polyethylene glycol (AA) in refluxing hexane at 70°C for 6 days as shown below: <chemistry id="chem0017" num="0017"><img file="EP1546159B1_D0017.tif" /></chemistry>

The reaction was conducted with constant stirring (i.e. magnetic stir bar) in a two-neck round-bottom flask attached to a Dean-Stark trap and a water-cooled condenser in a heated oil bath. The reaction was formulated with organosilicon:organic and solvent:monomer mole ratios equal to 1:1 and 10:1, respectively. The enzyme:monomer weight ratio was 1:10. The use of the Dean-Stark trap further promoted the esterification reaction.

After 4 days of reaction, the mixture was filtered, evaporated, and dried to isolate the product. The material was characterized by ¹H NMR, ESI MS, MALDI-TOF MS, and GPC-RI. N435 was observed to catalyze the formation of ester bonds with large polymeric substrates. The organosilicon polyester material was also observed to contain residual reactants. Based on the GPC-RI results, the Mn and Mw values were 5500 and 19000, respectively, with a 5.4 polydisperity.

Example 8

N435 lipase-catalyzed organosilicon polyester terpolymers were synthesized by reacting either the diacid disiloxane or diacid PDMS (x) with hexanediol (y) and adipic acid (z) in refluxing hexane or neat at 70°C for 4 days. The diagram below shows lipase-catalyzed diacid disiloxane (A) and diacid PDMS (B) polyester terpolymers. <chemistry id="chem0018" num="0018"><img file="EP1546159B1_D0018.tif" /></chemistry><chemistry id="chem0019" num="0019"><img file="EP1546159B1_D0019.tif" /></chemistry>

The reactions were conducted with constant stirring (i.e. magnetic stir bar) in a two-neck round-bottom flask attached to a Dean-Stark trap and a water-cooled condenser in a heated oil bath. The stoichiometry of the reactions were formulated with diacid:diol and solvent:monomer mole ratios equal to 1:1 (i.e. 1:2:1 and 0.5:2:1.5 x:y:z mole ratios) and 10:1, respectively. The enzyme:monomer weight ratio was 1:10. The use of a Dean-Stark trap further promoted the esterification reaction.

The progress of the reactions were monitored daily by ¹H NMR. After 4 days, the mixtures were filtered, evaporated, and dried to isolate crude oily products. The materials were characterized by ¹H, ¹³C, and ²⁹Si NMR, FTIR, GPC-RI, GPC-MALS, TGA, and DSC. <figref idref="f0009 f0010">Figs. 9-10</figref> illustrate the ¹³C NMR analyses of the esterification. Based on the material characterization, the diacid molecules were consumed during the lipase-catalyzed esterification reactions. The Mn, Mw, and polydispersity values of the disiloxane- and PDMS-functional terpolymers were calculated to be 3500-3700, 14000-15000, and 3.8-6.8 at x:y:z equal to 1:2:1, and 3200-4900, 20000-22000, and 4.0-6.8 at x:y:z equal to 0.5:2:1.5.

Based on the thermal analysis of the 0.5x:2y:1.5z terpolymers, the diacid disiloxane- (A) and diacid PDMS- (B) polyester terpolymers experienced critical mass losses at 353°C vs. 226°C (diacid disiloxane reactant) and 393°C vs. 283°C (diacid PDMS reactant), respectively. The temperature was observed to increase with the increase in organosilicon concentration. The Tg values were measured to be -76°C (A) and -117°C (B). In comparison to the diacid disiloxane (Tg =-76°C) and diacid PDMS (Tg = -119°C) reactants, similar amounts of energy were required to achieve molecular motion. During the DSC heating cycles, cold crystallization and crystalline phases were observed at ∼45°C (Tcc, 2.7 J/g), -1°C (Tm, 17.8 J/g), and 35°C (Tm, 2.8 J/g) in terpolymer A. A cold crystallization (Tcc = -15°C, 50.4 J/g) and two crystalline phases (Tm = 30°C, 45 J/g; 46°C, 8.6 J/g) were detected in the diacid disiloxane. Cold crystallization (Tcc =-101°C, 0.3 J/g) and multiple crystalline phases (Tm = -46°C &amp; -28°C, 14.6 J/g; 42°C &amp; 53°C, 0.7 J/g; 91 °C, 0.9 J/g) were observed in terpolymer B. A crystalline phase (Tm =-60°C, 16.8 J/g) was detected in the diacid PDMS. Given the range of energy values, the size of the regions of order or the degree of molecular interactions varied throughout the materials.