Immobilized Porcine Pancreas Lipase for Polymer Synthesis

Chapter 9

Immobilized Porcine Pancreas Lipase for Polymer Synthesis

Downloaded by UNIV OF PITTSBURGH on May 13, 2016 | http://pubs.acs.org Publication Date: September 19, 2008 | doi: 10.1021/bk-2008-0999.ch009

Feng He Key Laboratory of Biomedical Polymers of the Ministry of Education, College of Chemistry and Molecular Sciences, Wuhan University, Wuhan 430072, People's Republic of China

Porcine Pancreas Lipase (PPL), a common lipase with low cost, could be immobilized on silica particles with good stability and recyclablity. Then the immoblized lipase (IPPL) was employed as the catalyst for polymer synthesis, such as polyesters, polycarbonates, polyphosphates, and their copolymers. Here we present a mini-review of some works in our lab within this area.

Introduction Biodegradable polymers are receiving more and more attentions for their wide application in biomedical uses, such as drug carriers, matrices in tissue engineering, surgical sutures, etc (1). Sn(II) 2-ethylhexanoate, which has been approved for surgical and pharmacological applications by the F D A , is generally employed as the catalyst for the synthesis of biomedical polymers. However, it has been reported that Sn(II) 2-ethylhexanoate cannot be removed by a purification process such as the dissolution/precipitation method, thus the residual Sn may be concentrated within matrix remnants after hydrolytic degradation (2). To avoid the potential harmful effects of metallic residues in biomedical polymer materials, enzymatic polymerization is one of the powerful candidates for polymer synthesis (3). Enzymes, natural kinds of protein without toxicity, have remarkable properties 144

© 2008 American Chemical Society Cheng and Gross; Polymer Biocatalysis and Biomaterials II ACS Symposium Series; American Chemical Society: Washington, DC, 2008.


145 such as high catalytic and high selectivity under mild reaction conditions. Up to now, various kinds of biodegradable polymers have been synthesized by enzymatic polymerization, such as polyesters (4,5), polycarbonates (6,7), polyphosphates (8) and their copolymers. Among them, enzymatic ring-opening polymerization has received much attention as a nçw methodology of biodegradable polymer synthesis for lactones, cyclic carbonates and other cyclic monomers (9-12). However, to our knowledge, most previous studies of enzyme-catalyzed polymerizations have avoided temperatures > 90 oC, which is likely due to thermal deactivation of enzyme catalyst (13-15). It has been found that enzyme immobilization can improve the stability and recyclablity of native enzyme (16). Silica particles, activated by methanesulfonic acid, are effective and economic inorganic carriers for enzyme immobilization (17). Herein, we present a minireview of our works about immobilized porcine pancareas lipase on silica particles (IPPL) for polymer synthesis, such as polycarbonates, polyesters, polyphosphates and their copolymers.

IPPL For Polycarbonates Synthesis Aliphatic polycarbonates are a class of surface erosion biodegradable polymers attracting great interests due to their good biocompatibility, favorable mechanical properties and low toxicity (18J9). The polymerization of aliphatic cyclic carbonates such as trimethylene carbonate (TMC) have been extensively studied (20,21). Ο

Ο

ο

Ο

IPPL

-(-OCH CH CH OC 2

2

2

100 °C, in bulk Figure 1. Ring-opening polymerization of TMC catalyzed by IPPL.

In our study (6), porcine pancreas lipase (PPL) immobilized on silica particles (narrow distributed micron particles) was employed for ring-opening polymerization of T M C . No evidence of decarboxylation occurring during the polymerization. The results showed that silica microparticles improved immobilization efficiency much more. The most preferable polymerization temperature of T M C was 100 °C during 24h polymerization. The M of the resulting polymers was significantly increased compared with that catalyzed by n

Cheng and Gross; Polymer Biocatalysis and Biomaterials II ACS Symposium Series; American Chemical Society: Washington, DC, 2008.

146 P P L while the yield had no marked change. Furthermore, the recovered IPPL could be repeatedly used for many times. It is very interesting that the catalytic activity of recovered IPPL would increase and tend to keep constant after repeated uses. The highest M of P T M C 87400 was obtained at around 0.1 wt% of the seventh recovered IPPL. The excellent recyclablity of IPPL is very helpful to its further industry applications. n


Table 1. T M C polymerization catalyzed by recycled IPPL at 100 °C for 24h. Recycled Time l 2 3 4 5 6 7 Τ a

b

a

b

0

IPPL Conc.(wt%) 1.0 1.0 1.0 1.0 1.0 1.0 1.0 0.1

M

MJM

n

Yield(%) 70 75 81 78 83 81 72 69

n

14700 25400 30800 45200 61700 63000 64000 87400

1.92 1.77 2.11 2.01 1.94 2.53 2.20 2.06

IPPL used for the first time. Recycled IPPL used for the second time. The rest is the same as this method. Carried out by 0.1 wt% recycled IPPL which was the seventh used.

IPPL For Polyesters Synthesis In recent years, the enzymatic synthesis of biodegradable polyesters was focused on the polycondensation method (22,23). Among the very few successful example of enzymatic ring-opening polymerization for polyesters synthesis, Novozyme-435 (immobilized lipase Β from Candida antartica) has been proved an effective catalyst for polycaprolactone (PCL) synthesis in toluene (24). Considering the low cost and high recyclablity of IPPL, we also

Ο I! Ο IPPL

—^OCH CH CH CH CH (^ 2

2

2

2

2

180 °C, in bulk

Figure 2. Ring-opening polymerization of ε-CL catalyzed by IPPL.


147

employed the IPPL for ring-opening polymerization of ε-caprolactone (CL) with/without solvent. However, compared with Novozym-435, IPPL presented very low catalytic activity for P C L synthesis in toluene, and almost no P C L could be obtained. Thus, the IPPL-catalyzed polymerization of C L was carried out without solvent (in bulk). The results showed that M of resulting P C L was significantly increased compared with that catalyzed by native PPL. Higher temperature and longer reaction time both contributed to gain P C L with higher molecular weight, while the yield had almost no change (25). In addition, for evaluating the recyclablity of IPPL for the polymerization of C L , the most severe reaction conditions (180 °C, 240 h) were adopted in the recycling experiments. It was found that the recovered IPPL could be used again with compatible high catalytic activity. The highest M of 21300 of P C L could be obtained at 5.18 wt% of the reused IPPL at 180 °C for 240 h.


n

n

IPPL For Polyphosphates Synthesis Polyphosphates are one of the most promising biodegradable materials for their biocompatibility, low toxicity and biodegradability in biomedical use. They have the similar structure to nucleic acid and teichoic acids which are major components of cell walls, particularly in some bacteria and responsible for a number of biological functions. Polyphosphates and their copolymers have been used for gene therapy, tissue engineering and controlled drug delivery (26-30). In our lab, IPPL was also employed successfully for ring-opening polymerization of cyclic phosphate (ethylene isobutyl phosphate, EIBP) (8). A good coupling yield of IPPL was achieved to 41.88 mg of native lipase/g of silica particles. After incubation in water at 90 °C for l h , the activity of IPPL can retain above 70%. The optimum polymerization temperature was 70 °C. The enzymatic ringopening polymerization was achieved in bulk with M values ranging from 1600 to 5800 g/mol. It was found recovered IPPL worked more actively for the polymerization of EIBP and M was significantly increased. n

n

Ο

OCH CH(CH ) 2

3

2

70 °C, in bulk OCH CH(CH ) 2

Figure 3. Ring-opening polymerization of EIBP catalyzed by IPPL.


3

2

148


IPPL For Copolymers Synthesis Although various kinds of biodegradable polymers have been studied widely and then used in biomedical field, there are also many properties of these materials would be improved to fit various applications. One useful strategy for modifying the properties of biodegradable polymers is copolymerization (57). Another effective method is to introduce pendant functional groups to polymer materials (32). Though few studies on enzymatic copolymerization have been reported, it would be likely to attract more and more attentions in the future, due to its combined advantages of copolymerization and enzymatic polymerization. Herein we introduced some results on the enzymatic ring-opening copoly merization in our lab.

IPPL-catalyzed ring-opening copolymerization of BTMC with DTC IPPL with different size were employed for ring-opening copolymerization of 5-benzyloxy-trimethylene carbonate ( B T M C ) with 5,5-dimethyl-trimethylene carbonate (DTC) in bulk (33).

Table 2. Synthesis of poIy(BTMC-co-DTC) catalyzed by IPPL with different size 3

IPPL IE-1 IE-1 IE-1 IE-1 IE-2 IE-2 IE-2 IE-2 IE-3 IE-3 IE-3 IE-3

b

Conc.(wt%)

Yield%

0.1 0.2 0.5 1.0 0.1 0.2 0.5 1.0 0.1 0.2 0.5 2.0

82 80 82 86 83 77 82 83 94 92 73 39

13600 11400 6500 5900 26400 16500 10000 6900 19600 15300 5500 4700

2.05 2.88 2.36 2.75 1.74 1.84 2.22 2.66 1.80 3.16 2.75 2.28

a

Carried out in bulk at 150 °C for 24h with equal feed molar ratio of BTMC/DTC. Carrier sizes of IPPL: 150-250 μπι (IE-1), 75-150μπι (IE-2) and Ιμηι (IE-3). Determined using GPC. SOURCE: Reproduced with permission from reference 33. Copyright 2003 Elsevier.

b

c


149 Ο

Ο

Α

Λ

ο

ο

0 0 Ο Ο IPPL /II ν ,11 ν ( Ι + I J . u » ^COCH2CCH 0H COCH CHCH dttn

Immobilized Porcine Pancreas Lipase for Polymer Synthesis

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