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Polymer Degradation and Stability 94 (2009) 432–437

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Polymer Degradation and Stability

journal homepage: www.elsevier .com/locate/polydegstab

Studies on biodegradability of copolymers of lactic acid, terephthalicacid and ethylene glycol

R.K. Soni*, Shweta Soam, Krishna DuttDepartment of Chemistry, Ch. Charan Singh University, Meerut 250005, Uttar Pradesh, India

a r t i c l e i n f o

Article history:Received 25 July 2008Received in revised form4 November 2008Accepted 6 November 2008Available online 27 November 2008

Keywords:Lactic acidTerephthalic acidCopolymerFungiBiodegradation

* Corresponding author. Tel.: þ91 121 2958009.E-mail address: rksoni_rks@yahoo.com (R.K. Soni)

0141-3910/$ – see front matter � 2008 Elsevier Ltd.doi:10.1016/j.polymdegradstab.2008.11.014

a b s t r a c t

The copolymers were synthesized by the condensation of lactic acid, terephthalic acid and ethyleneglycol. Synthesized copolymers were characterized for various properties such as acid value, hydroxylvalue and number average molecular weight, etc. The copolymers were analyzed by FTIR. Copolymerswere biodegraded by different fungal species such as Aspergillus sp., Mucor sp., Alternaria sp. andRhizopus sp., etc. The extent of biodegradation was examined by weight loss and scanning electronmicroscopy. Biodegradation of copolymer with greater amount of lactic acid was faster than thebiodegradation of copolymer with lesser amount of lactic acid.

� 2008 Elsevier Ltd. All rights reserved.

1. Introduction

Polyesters have recently become materials of considerableinterest because of their potential biodegradability. It was foundthat due to their hydrolysable ester bonds, polyesters are biode-gradable. There are two kinds of polyesters: aliphatic andaromatic. Their biodegradability is completely different. Mulleret al. [1] concluded that pure aromatic polyesters are quiteinsensitive to any hydrolytic degradation it was observed thatdirect microbial or enzymatic attack of pure aromatic polyester arenot significant [2,3]. Researcher has recently claimed that aromaticpolyester could be disintegrated by microbial strains of Tricho-sporum and Arthrobactor in a time scale of weeks. Some growth ofAspergillus niger was found on the surface of aromatic polyesters[4]. On the contrary, aliphatic polyester is considered to besusceptible to microbial attack. Aliphatic polyester degradation isseen as a two step process: the first is depolymerization, or surfaceerosion. The second is enzymatic hydrolysis, which produceswater soluble intermediates that can be assimilated by microbialcells [1].

In order to solve the problem of aromatic polyester non-degradability, aliphatic–aromatic copolyesters were made. The

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idea was that by introducing aliphatic components into aromaticpolyester, its hydrolytic susceptibility should be increased [5].Raw materials consisting of copolyesters of 1,4 butanediol, ter-ephthalic acid and adipic acid (BTA copolyesters are preferentiallyused for commercial biodegradable copolyesters. The greater thefraction of terephthalic acid, the lower rate of biodegradation ofthe copolyesters. In the biodegradation assaying of the copo-lyesters, Thermomonospora fusca DSM 43793 was used. Copo-lyesters have been degraded within days [6]. The process ofaliphatic polyester degradation from depolymerization, aromaticand aliphatic oligomers are intermediates that are, at least,slightly water soluble and can be metabolized by microorganisms.Among these polymers much progress has been made in poly-lactic acid (PLA), polyglycolic acid (PGA) and their copolymer(PLGA). PLA and PGA and their copolymer can be synthesized inwide range of molecular weights by following two processes: 1)direct condensation reaction of lactic acid/or glycolic acid whichleads to low molecular weight polymers [7,8] 2) ring openingpolymerization of cyclic dimers, i.e. lactide and glycolide, in thepresence of metal catalysts to synthesize high molecular weightpolymers [9,10].

In the present work, polycondensation approach was attemptedto produce medium molecular weight copolymers of lactic acid,terephthalic acid and ethylene glycol. Copolymers so obtained werecharacterized for acid value, hydroxyl value and number averagemolecular weight, etc. Biodegradation studies were done by weightloss and scanning electron microscopy.

Fig. 1. Copolymer Y.

Table 1Summarised results of number average molecular weight of the synthesizedcopolymers.

Name Acid number Hydroxyl number Number averagemolecular weight

Y 0.020 0.023 10,000W 0.019 0.026 10,526

R.K. Soni et al. / Polymer Degradation and Stability 94 (2009) 432–437 433

2. Experimental

2.1. Materials

The � DL lactic acid (AR) and ethylene glycol were from Merc,terephthalic acid was from Sigma–Aldrich Chemical Co. whilestannous chloride was from Qualigens. All reagents were used as-received.

2.2. Synthesis of copolymers

Condensation reaction of copolymers was synthesized usinglactic acid, terephthalic acid and ethylene glycol.

Lactic acid (0.50 mol), terephthalic acid (0.25 mol) and ethyleneglycol (0.25 mol) were taken in three-necked round bottom flask. Athermometer was fitted to the neck, a stirrer to the other and DeanStark was fitted to third neck. In first step temperature was kept at100 �C for 7 h. In second step the reaction was carried out at 260 �Cfor 10 h and 0.1% SnCl2 was added as a catalyst with constant stirring.The reaction mixture was poured in a Petri dish, having silica gelcoating. After cooling, copolymers were removed from Petri dish.

Fig. 2. Copolymer W.

Lactic acid (0.10 mol) with terephthalic acid (1.9 mol) andethylene glycol (1.9 mol) was taken in three-necked round bottomflask. The procedure is same as described above.

3. Characterization

Copolymers were synthesized by condensation reaction.Copolymers so obtained were characterized for acid value, hydroxylvalue and number average molecular weight, etc.

3.1. Acid value (ASTM D 1639)

Acid number was determined by dissolving 2.5 g polymericmaterial in ethanol and was titrated against 0.1 N of standardizedKOH (using phenolphthalein as an indicator) until a light pinkcolour of the solution persisted.

The acid number was calculated by the following expression:

Acid number ¼ 56:1VN=m

where V, the volume of KOH; N, normality of the KOH; m, weight ofpolymeric sample taken.

3.2. Hydroxyl value (ASTM D 2840)

Hydroxyl value was determined according to ASTM D 2840method A. Approximately 0.50 g of sample was taken in a 50 mlphthoylating mixture and hydrolyzed by adding 100 ml of chilleddistilled water in another flask. Under vigorous stirring, 20 mlbenzene was added. The resulting solution was titrated against0.5 N standardized KOH (using phenolphthalein as an indicator). Ablank run without sample was also performed. The hydroxyl valuewas calculated with the help of following expression:

Fig. 3. FTIR of copolymer W.

Fig. 4. FTIR of copolymer Y.

+ +C COOH

COOH

COOHHOOC

CH3

CH2OH

CH2OH

CH2 CH2

CH2 CH2

HO

H

CH3

HO

H

C COO

CH3

HO

H

C COO

Polymerization

COO

O

O

O

O

C

C

Fig. 5. Copolymerization of lactic acid, terephthalic acid with ethylene glycol.

Table 2Biodegradation of copolymer of lactic acid with terephthalic acid and ethylene glycol(W) through different fungal species.

Fungi Species After0 days

After15 days

After30 days

After45 days

After60 days

Alternaria alternata 0% 18% 40% 53% 65%Pencillium sp. 0% 5% 43% 55% 59%Rhizopus sp. 0% 10% 45% 53% 55%Mucor sp. 0% 15% 33% 43% 53%Aspergillus niger 0% 10% 33% 43% 68%Aspergillus clavatus 0% 3% 18% 43% 48%Aspergillus versicolor 0% 13% 26% 35% 45%Aspergillus fumigatus 0% 3% 31% 43% 51%

Table 3Biodegradation of copolymer of lactic acid with terephthalic acid and ethylene glycol(Y) through different fungal species.

Fungi Species After0 days

After15 days

After30 days

After45 days

After60 days

Alternaria alternata 0% 11% 25% 38% 70%Pencillium sp. 0% 6% 15% 47% 62%Rhizopus sp. 0% 9% 16% 44% 60%Mucor sp. 0% 8% 14% 42% 65%Aspergillus niger 0% 8% 14% 46% 81%Aspergillus clavatus 0% 14% 29% 41% 64%Aspergillus versicolor 0% 13% 28% 27% 68%Aspergillus fumigatus 0% 10% 18% 46% 55%

R.K. Soni et al. / Polymer Degradation and Stability 94 (2009) 432–437434

Hydroxyl number ¼ 56:1ðV1 � V2ÞN=m

where V1, volume of 0.5 KOH used for titration of the blank;V2volume of 0.5 KOH used for titration of the sample; N, normalityof KOH; m, weight of the polymer sample.

3.3. Number average molecular weight (Mn)

The number average molecular weight was calculated using thefollowing expression:

Number average molecular weightðMnÞ ¼ F100=C

F, functionality of polymer; C, acid value.

3.4. IR spectroscopy

The FTIR spectra of synthesized copolymers were recordedusing Perkin Elmer model spectrum BX Series FTIR. The spectrawere recorded by using KBr Pallets. A total scan 16 scan per sampleat resolution of 4 cm�1 were obtained over mid-IR region of 4000–400 cm�1.

4. Biodegradation

Synthesized copolymers were tested for their biodegradabilityby fungal species. Eight fungal species were selected for this study.These were A. niger, Aspergillus versicolor, Aspergillus clavatus,Aspergillus fumigatus, Alternaria alternata, Mucor sp., Penicillium sp.

Fig. 6. SEM of copolymer Y (before biodegradation).

Fig. 7. SEM of copolymer Y (After 30 days of biodegradation by Aspergillus niger). Fig. 8. SEM of copolymer Y (After 30 days of biodegradation by Aspergillus niger).

R.K. Soni et al. / Polymer Degradation and Stability 94 (2009) 432–437 435

and Rhizopus sp. Identified fungi were procured from MicrobiologyLaboratory, Department of Botany, CCS University campus Meerut.

Potato Dextrose Agar medium was prepared and 15–20 ml ofthis medium was poured in each Petri dish aseptically. Each Petridish was inoculated centrally with a bit of mycelium and sporesfrom the culture of fungus. Petri plates were incubated at 25�1 �Cfor 7 days in BOD incubator. Entire growths of fungus were scrap-ped from petri plates. Entire material was placed in conical flaskhaving 100 ml sterilized water. This suspension was used to inoc-ulate the samples of copolymers. The samples were taken in conicalflasks having prepared suspension. These samples were incubatedin at 25 �C for 15, 30, 45 and 60 days. The samples were removedfrom suspension. The samples were biowashed by water and driedunder vacuum. The biodegraded copolymers were examined byweight loss and scanning electron microscopy.

4.1. Weight loss

Weight loss was calculated by the following expression:

Weight loss ¼ Weight of sample before degradation�Weight of sample after degradationWeight of sample before degradation

� 100

4.2. Scanning election microscopy

SEM studies were done by TEOL JSM – 840 Scanning ElectronMicroscope.

5. Results and discussion

5.1. Synthesis of copolymers

Synthesized copolymers of lactic acid, terephthalic acid andethylene glycol were obtained by condensation reaction. Thecopolymer having greater amount of lactic acid was yellow col-oured solid mass and designated as ‘Y’ (Fig. 1). The copolymer,having less amount of lactic acid was white coloured solid mass anddesignated as ‘W’ (Fig. 2). The acid value, hydroxyl value andnumber average molecular weight was summarized in Table 1.The

synthesized copolymer was medium molecular weight polymer ofthe range of 10,000.

Infrared spectroscopy of the products indicates that esterformation in products, the presence of aromatic hydrocarbon andthe IR spectra of product ‘W’ are illustrated in Fig. 3.

The presence of a sharp band at 1716 cm�1 may be attributed tothe presence of C]O stretching in aryl ester. Two bands at1134 cm�1 and 1281 cm�1 indicate C–O str. in the ester. The broadpeak at 3447 cm�1 may be attributed to O–H str. of alcohol anda weak band at 3064 cm�1 indicates the presence Ar–H str. inaromatic hydrocarbons. The bands at 1507 cm�1 and 1575 cm�1

indicates C]C str. in aromatic hydrocarbon.The IR spectrum of ‘Y’ is illustrated in Fig. 4. The presence of

a band at 1718 cm�1 may be attributed to the presence of C]O str.in aryl ester. Two bands at 1120 cm�1 and 1272 indicated C–O str. inester. Broad band at 3435 cm�1 indicates free hydroxyl group. Aweak band at 1410 cm�1 indicates C–H str. in –CH2– group. On thebasis of IR spectrum, proposed reaction scheme is shown Fig. 5.

5.2. Biodegradation by fungal species

5.2.1. Weight loss after biodegradationThe percentage weight loss of ‘W’ and ‘Y’ is presented in Tables 2

and 3. Table 2 shows highest biodegradation occurs in the case ofA. niger as compared to other fungal species. The A. alternata gavegood results also. The least result was obtained by biodegradationby A. versicolor. In the case of Penicillium sp. and A. clavatus andA. fumigatus the initial rate of loss is lowest from 3% to 5% incopolymer W. The rate of biodegradation of W is faster for Penicilliumsp. after period of 15 days. However the initial loss was slow for thissp. The Alternaria sp. loss after 60 days is quit comparatively highthan other sp. Table 3 shows summarized results of biodegradationof copolymer of lactic acid with terephthalic acid and ethylene glycolhaving excess amount of lactic acid as good results of copolymerbiodegradation. The degradation rate was much faster in the case of

Fig. 11. SEM of copolymer W (After 30 days of biodegradation by Aspergillus niger).

-COO-CH -CH O-C2 2

CH3O

H

O

O

O

-O-C- COO-CH -CH -OOC-C-O-2 2

H

H

H

H

H-O O

cH3

C 3H

Copolymer of lactic acid and terephthalic acid, ethylene glycol

-O-C- -COO-C -C -OOC-C-O-2 2H H

O

O

OO

Asp

O-C-

O

OO

O

HisAsp

ser

+O-H N-HO OO O

His

O- ----H

ser

N N-HO OO O

O O

-

C N

Lipases

Fig. 9. SEM of copolymer W (before biodegradation).

R.K. Soni et al. / Polymer Degradation and Stability 94 (2009) 432–437436

copolymer Y. The biodegradation weight loss was observed 81% inthe case of A. niger after 60 days. Similarly A. alternata shows similarrate with weight loss of 70%. The rate of degradation was foundhigher with copolymer Y. The percentage of terephthalic acid in thecopolymers W and Y depends on the percentage of terephthalicacid. High amount weight loss was found with high amount oflactic acid.

5.2.2. Scanning electron microscopySEM studies were done after biowashing the samples by water

and ethanol (50:50). Biodegradation was appeared clearly on thesurface. SEM micrographs were scanned before and after biodeg-radation. Fig. 6 indicates the SEM micrographs of undegradedcopolymer Y. Fig. 7 shows the SEM of copolymer Y taken after 30days of biodegradation. After 30 days of inoculating the A. niger tothe copolymer Y has biodegraded. Figs. 7 and 8 reveal that sphericalholes probably resulting from localized enzymatic action withcolonization of fungi. Copolymers lost their properties beforesignificant weight loses were observed.

Fig. 9 indicates the SEM micrographs of undegraded copolymerW. After 30 days of inoculating the A. niger to the copolymer Wwas biodegraded (Fig. 10). The SEM of ‘W’ was scanned after 30days of biodegradation by A. niger revealing that spherical holes

Fig. 10. SEM of copolymer W (After 30 days of biodegradation by Aspergillus niger).

probably resulting from localized enzymatic action with coloni-zation of fungi (Figs. 10 and 11) at the magnification of 300 and600. Copolymers lost their properties before significant weightloses were observed.

O

O

O

O

O

O

O

+

His

His

His

H

H

H

H

H

+

+H

O

O H

CH3

CH3

Asp

Asp

Asp

H-O

H-O

C

C

-COO-CH -CH -COO-C-O2 2

-COO-CH -CH -COO-C-O-2 2

ser

ser

ser

O

O

N

N

N

N

N

N

O O

O O

Fig. 12. Mechanism of enzymatic degradation (cleavage of ester bond by lipases).

R.K. Soni et al. / Polymer Degradation and Stability 94 (2009) 432–437 437

6. Purposed mechanism of degradation

Biodegradation of plastics is usually a heterogeneous processbecause of water insolubility and size of polymer molecule;microorganisms are not able to pick up the polymer directly intothe cells where most of the chemical processes take place, but firsthave to secrete extracellular enzyme which depolymerize thepolymers outside the cells. If the molar mass of the polymers issufficiently reduced to generate water soluble intermediates, thesecan be transported into the microorganism and introduced thereinto metabolic pathways [11]. As a final result of these processesmicrobial metabolic end products such as water, carbon dioxide,methane etc. and new biomass are produced. In many cases, thefirst step in the degradation process, the reduction of molecularmass, is the rate-limiting factor of these plastics biodegradation.Most of the enzymes used in the degradation of polyesters aremicrobial enzymes, originated from fungi .The hydrolases family,amongst others consisting of esterases, lipases, proteases,amidases, epoxide hydrolases, nitrolases and glycosidases, isa group of enzymes that can catalyses bond cleavage by reactingwith water [12]. Lipases can be isolated by various fungi. Lipasescatalyze the hydrolysis of water insoluble long chain esters ofsynthesized copolymer as shown in Fig. 12 [13].

7. Conclusions

Copolymers of lactic acid, terephthalic acid and ethylene glycolcan be synthesized by taking different ratio of monomers. Copol-ymers can be biodegraded by different fungal sp. Such as Aspergillussp., Alternaria sp., Penicillium, Rhizopus sp., and Mucor. Synthesizedcopolymer has ester bond. The enzyme lipases catalyze thehydrolysis of water. The biodegradation of ‘Y’ is greater due to thepresence of greater extent of lactic acid.

Acknowledgement

The authors thankfully acknowledge the contribution of Prof.M. U. Charaya, Department of Botany, Chaudhary Charan SinghUniversity, Meerut, U.P, India for providing microbial strains forthis work.

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