WO2010089765A1

WO2010089765A1 - Enzymatic polymerization process for the production of polylactide polymers

Info

Publication number: WO2010089765A1
Application number: PCT/IN2010/000010
Authority: WO
Inventors: Venkata Ramakhrishna Sonti; Ambadas Gore Mahesh
Original assignee: Praj Industries Limited
Priority date: 2009-01-07
Filing date: 2010-01-07
Publication date: 2010-08-12

Abstract

The present invention provides a process for producing polylactide polymer comprising subjecting the lactide to enzymatic ring opening polymerization condition in the presence of immobilized laccase enzyme by reacting lactide and immobilized laccase enzyme, followed by evacuating the reaction mixture and further polymerizing the lactide, dissolving the polymerization product in chloroform, separating the enzyme catalyst, concentrating the polylactide polymer, and purifying the polylactide polymer.

Description

ENZYMATIC POLYMERIZATION PROCESS FOR THE PRODUCTION OF

POLYLACTIDE POLYMERS

CROSS- REFERENCE TO RELATED APPLICATION

This application claims priority from provisional patent application filed on January 07, 2009 at the Indian patent office having patent application no. 1413/MUM/2008.

FIELD OF THE INVENTION

The present invention relates to a process for production of polylactide polymer. In particular the present invention provides an enzymatic ring opening polymerization (ROP) of lactide to synthesize polylactide [PLA] polymer using an immobilized laccase enzyme catalyst.

BACKGROUND OF THE INVENTION

Typically various methods have been developed for the polylactide polymer to replace the non biodegradable polymers. Advanced technology in petrochemical based polymers has brought many benefits to mankind. However, the extensive use of petrochemical polymer based products, particularly as disposable items, has considerably disturbed and damaged the ecosystem because petrochemical polymer based waste products are non-biodegradable.

In recent years, with the critical situation of the worsening of the global environment with global warming and the like, the construction of systems with sustainable use of materials has been accelerated from the viewpoint of using effectively the limited carbon resources and conserving limited energy resources. In the case of polymer products, after use the products are reused as they are (examples of this case include the conversion of PET bottles to fibrous material), or recycled or discarded. For recycling processes, material recycling processes, chemical recycling processes, thermal recycling processes, and the like are used. However, material recycling processes involve deterioration in quality, such as a drop in molecular weight, chemical recycling processes consume much energy and thermal recycling process generate a large amount of carbon dioxide gas. Thus, each of these processes involves problems.

The concern for the adverse environmental impact of such non-biodegradable plastic wastes is growing worldwide. The methods for disposal of plastic wastes are limited. Incineration of plastic waste generates toxic air pollution and landfill sites for disposal of such plastic waste products are limited. Besides, the petroleum resources are becoming limited. Apart from the viewpoint of reuse, attention has been paid to the so-called biodegradable polymers, which are degraded by bacteria and the like in the ground, as polymers imposing only small loads on environment.

Various such biodegradable polymers have been suggested. For example, for biodegradable polymers biodegradable polyesters are known. Typical examples of biodegradable polyesters produced by chemical synthesis include polycaprolactone (PCL), polylactide (PLA), polyhydroxybutyric acid (PHB), and aliphatic polyesters made from diol and succinic acid, such as polyethylene succinate, polybutylene succinate (PBS) and polybutylene succinate/adipate copolymer (PBS/A).

Among these, as well as polylactide and polycaprolactone, polybutylene succinate (PBS) has been investigated in an attempt to make it practicable as a typical chemically-synthesized biodegradable plastic, since PBS can be obtained from 1,4- butanediol and succinic acid by a petrochemical industrial process. Among the above- mentioned examples, polylactide (PLA) is a polymer yielded by polycondensation of lactic acid, which is obtained by fermenting corn starch, or the like, which is a renewable resource. It can be said that this polymer is a low environment load polymer, which does not cause a direct increase in the total amount of carbon dioxide gas even if the polymer is finally biodegraded or burned up. Lactic acid or its dimmer (lactide) thereof, which is the raw material of the polymer, has already been produced with a high efficiency by research and development over many years.

Polylactide (PLA) offers a substitute for petroleum based plastics, as PLA is a biodegradable thermoplastic which is derived from renewable resources such as corn starch or sugarcanes by a combination of fermentation and chemical polymerization. PLA polymers have found wide range of applications in medical and pharmaceutical industries such as surgical sutures, implant devices, controlled drug delivery, food packaging industry and artificial skin because of their biocompatibility and biodegradability. It is also considered as the most promising biodegradable plastic because of its high strength, thermoplasticity, fabricability, biodegradability and availability from renewable resources.

Polylactide has strength equivalent to that of polyethylene or polystyrene, has a higher transparency than other biodegradable plastics, and is superior in weather resistance, heat resistance, workability and the like. PLA has already been put to practical use such as in covering materials for agriculture, fibers, earth-retaining netting, weed- preventing bags and the like. Accordingly, PLA is a biodegradable plastic that has currently been developed furthest towards practical use.

There are two major approaches used to produce PLA from the lactic acid either by direct condensation polymerization of lactic acid or ring-opening polymerization through the lactide intermediate. The first approach is least attractive because of low molecular weight of resulting polymers. In the second approach, transition metal ion catalyst is used for the ring opening of lactide intermediate which leads to the contamination of polymer. Further, high temperature polymerization technique employed in this approach of using metal ion catalyst, leads to racemization of lactic acid. Hence the polymers produced using the second approach cannot be employed for the biomedical applications. There, is therefore a need for enzymatic polymerization technique of the present invention which enables to overcome the above-mentioned limitations.

SUMMARY OF INVENTION

BRIEF DESCRIPTION OF THE DRAWING

Figure 1 illustrates the entire process flow chart for the enzymatic synthesis of Polylactide from L-lactide using immobilized laccase enzyme. The process of the present invention is illustrated with a flow chart which is intended to illustrate the steps of the process and is not intended to be taken restrictively to imply any limitation on the scope of the present invention in the accompanying figure.

Figure 2 is a graph showing ¹H NMR spectrum of a produced polylactide polymer in Example 2 and 3 of the invention.

DETAILED DESCRIPTION OF INVENTION

The following is a detailed description and explanation of the preferred embodiments of the process of the invention with some examples thereof. In order to better appreciate the invention, it is described with reference to the process flow chart in figure 1 which illustrates the preferred embodiment of the invention.

As indicated in figure 1, a process for synthesis (10) of Polylactide from lactide using immobilized enzyme catalyst laccase wherein the L-lactide and immobilized enzyme catalyst laccase is mixed together (12) in the polymerization test tube. The reaction mixture is evacuated (14) at about 50⁰C to about 80⁰C for about 1 hour to 2 hours and the test tubes kept under vacuum for about 1 hr at about 60⁰C in order to remove the traces of moisture and the test tube with reaction mixture is sealed under vacuum. The reaction mixture is subjected to polymerization reaction (16) of lactide by an immobilized enzyme catalyst laccase at about 80⁰C to about 130⁰C. After the polymerization reaction, the reaction mixture is dissolved into chloroform (18) followed by filtration in order to remove (20) the immobilized enzyme catalyst laccase. The filtrate chloroform solution subjected to concentration (22) to obtain the polymerization product; Polylactide. The crude Polylactide polymer is precipitated into methanol (24) the purification of Polylactide is facilitated by dissolving into chloroform and reprecipitating into methanol as discussed in step (22) and (24). The crude Polylactide is further separated by filtration and dried under vacuum (26). The dried Polylactide (28) is stored under vacuum at room temperature.

For obtaining the L-lactide, the lactide synthesis is carried out as per the prior art method wherein lactide is prepared by using lactic acid by chemical synthesis method. The polymerization of lactide conducted by a simple operation using immobilized enzyme catalyst laccase. Accordingly, the invention makes it possible to construct a complete- cycle type polymer material using system which is of environmentally acceptable and is capable of reusing carbon resources completely.

In one of the embodiment of the present invention uses immobilized laccase enzyme for a ring opening polymerization of lactide for the synthesis of Polylactide. Laccases (EC 1.10.3.2) are copper-containing oxidase enzymes act on phenols and similar molecules, performing one-electron oxidations.

The polymer of Polylactide is synthesized using immobilized enzyme catalyst laccases. The enzymatic synthesis process is initiated by evacuation of the lactide with an addition of about 0.5 % to about 6.5 % immobilized laccase enzyme in an oil bath for about 1 hour to about 2 hours. The reaction mixture is sealed under vacuum and temperature of the reaction mixture is gradually increased to about 80 C to about 130 C. Reaction is carried out for about 24 hours to about 120 hours for facilitating the polymerization process. The polymer thus produced is reprocessed by dissolution in chloroform reprecipitation with methanol and further drying to obtain a white production with about 17 % to about 75 % purity.

Following examples illustrate the preferred embodiments of the present invention and are not limiting of the specification and claims in any way.

Example 1 : Lactide Synthesis.

The lactide synthesis was carried out as per the prior art method. L- Lactic acid (88%) is purchased from Merck chemicals, India. Dibutyl tin oxide is purchased from Aldrich Chemicals, India. The three neck flasks kept in an oil bath for heating and mixing of 246 gm of L-lactic acid is done using magnetic stirrer. Temperature was progressively increased to about 140⁰C under nitrogen flushing for the removal of water as distillate. Reaction mass then cooled to about 90° C and about 1.8 gm of dibutyl tin oxide catalyst was then added to the reaction mixture. Temperature was then progressively increased to about 160° C and vacuum was increased to about 60 mm Hg over a period of about 2 hrs for the removal of remaining water. After removal of water reaction mixture was cooled to about 90° C. Water condenser was replaced to hot water condenser and receiver was cooled with ice and temperature was progressively increased to about 260 C and vacuum to about 4 mm Hg over a period of about 5 hrs for distillation. 120 gm crude lactide was obtained with a yield of about 90% based on lactic acid. Crude dilactice is purified by crystallization from ethyl acetate using crude lactide and ethyl acetate ratio 5:3 by dissolving lactide into 100 ml Round Bottom flast and solution was allowed to cool at 5° C for 2-4 hrs crystallization of lactide to occur. The second recrysatlization with ethyl acetate resulted in purified lactide. Lactide filtered to separate and stored under vacuum.

Example 2: Polymerization of L- Lactide using immobilized laccase enzyme.

Immobilized laccase [Novoprime Base 268], obtained from Novozyme, India. In a 50 ml polymerization test tube about 6 gm of purified L-lactide was added with the predetermined amount of immobilized laccase enzyme as shown in Table 1. The test tubes kept under vacuum for about 1 hr at about 60⁰C to remove any traces of moisture and test tube was vacuum sealed. Polymerization of lactide carried out at predetermined temperature and time as shown in Table 1. After the reaction, the reaction mixture was dissolved into chloroform and enzyme is separated by filtration. PLA thus synthesized was reprocessed by dissolution in chloroform followed by reprecipitation with methanol and drying under vacuum. The results are summarized as shown in Table 1. These polymers further characterized using GPC (Gel Permeation Chromatography), TGA (Thermogravimetric analysis) and DSC (Differential Scanning calorimetry) and results are summarized in Table 1 and 2.

Table 1: L-lactide polymerization using laccase

a = weight % amount of enzyme in weight % based on L-lactide b = Yield calculated based on presence of L-lactide Mw = Weight average molecular weight D = Mw/Mn i.e. Polydispersity

Example: 3 Purification and characterization of Polylactide synthesized by immobilized laccase enzyme.

The PLA was purified by filtration and further dried under vacuum. Characterization of dried PLA was done using GPC (Gel Permeation Chromatography), TGA (Thermogravimetric analysis), and DSC (Differential Scanning calorimetry) as shown in table 1 and table 2.

As shown in figure 2, H NMR spectra were recorded on Bruker superconducting FT- NMR AC 300 operating at 300 MHz using CDCl₃ as a solvent. As shown in table 2. Differential scanning calorimetry (DSC) was performed using TA [Waters India] DSC equipped with a Thermal Analysis Data Station (TADS) at a heating rate of 10°C/min in nitrogen atmosphere. Temperature range used was about 0 C to about 200⁰C. Duplicate determinations were carried out for each sample. Thermogravimetric analysis [TGA] analyses were recorded on TA [Water India] TGA Q 500, Instrument. Temperature range used was about O⁰C to about 400⁰C and heating rate 10°C/min in nitrogen atmosphere. The molecular weight analyses were recorded (as shown in Table 1.) using Agilent Gel Permeation Chromatography [GPC] system. THF solvent was used with the flow rate lml/min. Ultrasyragel columns were used with polystyrene standards. Melting point and moisture content was determined using Mettler India instrument. Duplicate determinations were carried out for each sample.

Table 2: Characterization of PLA synthesized by laccase

Tg: Glass transition temp. Tc: Crystallization Temp., Tm: Melting Temp.

The ring opening polymerization of lactide and the polymerization process of the invention using immobilized laccase may be performed by a simple operation by use of one pot, is mild in reaction conditions, and consumes low energy. In the case that the ring opening polymerization is conducted by chemical degradation or thermal degradation, both ends of a generated low molecular weight compound is irregular and such compound cannot be re-polymerized into a polymer. Accordingly, the ring opening polymerization using immobilized laccase enzyme makes it possible to construct a complete-cycle type polymer material using system which is environmentally acceptable and is capable of reusing carbon resources completely. Thus, the industrial utility of value of this invention is extremely high.

Claims

WE CLAIM

1. A process for producing polylactide polymer comprising subjecting a lactide to enzymatic ring opening polymerization condition in the presence of a laccase enzyme by reacting lactide and laccase enzyme, followed by evacuating the reaction mixture, polymerizing the lactide, dissolving the polymerization product in chloroform, separating the enzyme catalyst, concentrating the polylactide polymer, and purifying the polylactide polymer

2. A process as claimed in claim 1, wherein the lactide is L-lactide.

3. A process as claimed in claim 1, wherein the laccase enzyme is in the immobilized form.

4. A_. process as claimed in claim 1, wherein evacuating the reaction mixture at about 50⁰C to about 80⁰C for about 1 hour to 2 hours.

5. A process as claimed in claim 1, wherein polymerizing the lactide in* presence of laccase enzyme at about 80 C to about 130⁰C for about 24 hours to 120 hours

6. A process as claimed in claim 1, wherein separating the laccase enzyme from the reaction mixture by filtration.

7. A process as claimed in claim 1, wherein concentrating the polylactide polymer in presence of methanol.

8. A process as claimed in claim 1, wherein purifying the polylactide polymer by filtration and dried under vacuum.