Comparing the Growth Characteristics of Three Bacteria Involved in Degrading Rubbers

Three bacteria isolated for degradation of rubbers were compared for their growth characteristics derived from the Bradford protein assay and turbidity (optical density, OD) measurement. Both Alcaligenes xylosoxidans T2 and Pseudomonas aeruginosa GP10 were fast-growing bacteria while Nocardia corynebacterioides S3 was a slow grower utilizing rubber as the sole source of carbon and energy, but the extent of degradation was lower by the formers than the latter. A. xylosoxidans T2, P. aeruginosa GP10 and N. corynebacterioides S3 showed a typical sigmoidal growth pattern based on binding of Coomassie Brilliant Blue G250 to bacterial proteins and spectrophotometrical measurement at 600 nm. Both assays showed similar growth characteristics for all three bacteria in this study. Degradation of rubber was more pronounced by N. corynebacterioides S3 than either A. xylosoxidans T2 or P. aeruginosa GP10 during 70 days of incubation. Our results suggest that slow-growing bacteria may play a much greater role in degrading polymeric materials than was previously believed.     

Confirmation of colonization of degrading bacterium strain SC-17 on poly(3-hydroxybutyrate) cast film

To clarify the mechanism of microbial degradation owing to colonization ofPseudomonas sp. strain SC-17 on a poly(3-hydroxybutyrate) (PHB) cast film surface, morphological and spectroscopic analyses of the degraded film were investigated and colonization kinetics on the films is discussed. By spectroscopic analysis of unique hemispherical degradation marks, cells of strain SC-17 adhering to the marks' surface were confirmed. To account for the hemispherical hole formation and their linear enlargement with culture time, a three-dimensional colony growth model toward the interior of the film was developed. The model explained the hemispherical hole formation well. It was concluded that the hemispherical holes resulted from the colonization of strain SC-17 on the film surfaces. It was further determined that the microbial degradation by strain SC-17 is initiated from small pits formed on the PHB film surface.     

Degradation and mineralization of cellulose acetate in simulated thermophilic compost environments

Residual cellulose acetate (CA) films with initial degree of substitution (DS) values of 1.7 and 2.5 (CA DS-1.7 and DS-2.5) were recovered from a simulated thermophilic compost exposure and characterized by gel permeation chromatography (GPC), proton nuclear magnetic resonance (¹H NMR), and scanning electron microscopy (SEM) to determine changes in polymer molecular weight and DS and to study microbial colonization and surface morphology, respectively. During the aerobic degradation of CA DS-1.7 and CA DS-2.5 films exposed for 7 and 18 days, respectively, the number-average molecular weight (M _n) of residual polymer decreased by 30.4% on day 5 and 20.3% on day 16, respectively. Furthermore, a decrease in the degree of substitution from 1.69 to 1.27 (4-day exposure) and from 2.51 to 2.18 (12-day exposure) was observed for the respective CA samples. In contrast, CA films (DS-1.7 and DS-2.5) which were exposed to abiotic control vessels for identical time periods showed no significant changes inM _n and DS. SEM photographs of CA (DS-1.7 and DS-2.5) film surfaces after compost exposures revealed severe erosion and corresponding microbial colonization. Similar exposure times for CA films in abiotic control vessels resulted in only minor changes in surface characteristics by SEM observations. The conversion of CA DS-1.7 and DS-2.5 to CO₂ was monitored by respirometry. In these studies, powdered CA was placed in a predigested compost matrix which was maintained at 53°C and 60% moisture content throughout the incubation period. A lag phase of 10- and 25-day duration for CA DS-1.7 and DS-2.5, respectively, was observed, after which the rate of degradation increased rapidly. Mineralization of exposed CA DS-1.7 and DS-2.5 powders reported as the percentage theoretical CO₂ recovered reached 72.4 and 77.6% in 24 and 60 days, respectively. The results of this study demonstrated that microbial degradation of CA films exposed to aerobic thermophilic laboratory-scale compost reactors not only results in film weight loss but also causes severe film pitting and a corresponding decrease in chainM _n and degree of substitution for the residual material. Furthermore, conversions to greater than 70% of the theoretical recovered CO₂ for CA (DS 1.7 and 2.5) substrates indicate high degrees of CA mineralization.Guest Editor: Dr. Graham Swift, Rohm & Haas.     

Biodegradability of chitin- and chitosan-containing films in soil environment

The biodegradation of polyethylene-chitin (PE-chitin) and polyethylene-chitosan (PE-chitosan) films, containing 10% by weight chitin or chitosan, by pure microbial cultures and in a soil environment was studied. Three soil-inhabited organsims,Serratia marcescens, Pseudomonas aeruginosa, andBeauveria bassiana were able to utilize chitin and chitosan in prepared PE-chitin and PE-chitosan films after eight weeks of incubation at 25°C in a basal medium containing no source of carbon or nitrogen. In a soil environment, the biodegradation of those films was studied and compared with a commercial biodegradable film containing 6% by the weight of corn starch. In soil placed in the lab, 73.4% of the chitosan and 84.7% of the chitin in the films were degraded, while 46.5% of the starch in the commercial film was degraded after six months of incubation. In an open field, 100% of the chitin and 100% of the chitosan in the films were degraded, but only 85% of the starch in the commercial film was degraded after six months of incubation. The weight of controls, (polyethylene films), remained mainly stable during the incubation period. Both PE-chitin and PE-chitosan films degraded at a higher rate than the commercial starch-based film in a soil environment indicating the potential use of chitin-based films for the manufacturing of biodegradable packaging materials.     

Biodegradability of Chemically Modified Starch (RS4)/PVA Blend Films: Part 2

Biodegradable films were successfully prepared by using cornstarch (CS), chemically modified starch (RS4), polyvinyl alcohol (PVA), glycerol (GL), and citric acid (CA). The physical properties and biodegradability of the films using CS, RS4, and additives were investigated. The results of the investigation revealed that the RS4-added film was better than the CS-added film in tensile strength (TS), elongation at break (%E), swelling behavior (SB) and solubility (S). Especially, the RS4/PVA blend film with CA as an additive showed physical properties superior to other films. Furthermore, when the film was dried at low temperature, the properties of the films clearly improved because the hydrogen bonding was activated at low temperature. The biodegradation of films was carried out using the enzymatic, microbiological and soil burial test. The enzyme used in this study was amyloglucosidase (AMG), α-amylase (α-AM) and β-amylase (β-AM). At the enzymatic degradation test, the GL-added films had an approximately 60% degradation, while the CA-added films were degraded about 25%. The low degradation value on CA-added film is attributed to low pH of film added CA that deactivated the enzymatic reaction. The microbiological degradation teat was performed by using Bacillus subtilis and Aspergillus niger.     

Pure-culture and enzymatic assay for starch-polyethylene degradable plastic biodegradation withStreptomyces species

Eleven starch-polyethylene degradable plastic films were prepared from masterbatches from Archer Daniels Midland Inc. (ADM), EcoStar Inc. (SLS), and Fully Compounded Plastic Inc. The biodegradability of initial and 70°C heat-treated materials was determined using a pure-culture assay withStreptomyces badius 252,S. setonii 75Vi2, orS. viridosporus T7A or without bacterial culture (control). Films were treated with 10-foldS. setonii culture concentrates and compared with inactive enzyme controls. Changes in each films mechanical property, molecular weight distribution, and Fourier-transformed infrared spectrum (FT-IR) were determined, and results were evaluated for significant differences by analysis of variance. Cell mass accumulation on each film was quite pronounced. In pure-culture studies, biodegradation was demonstrated for ADM-7 and SLS-2 initial films and for ADM-6 heat-treated films, whereas after 3-week treatment with activeS. setonii culture concentrates (enzyme assay), reductions in mechanical properties and changes in FT-IR spectrum were illustrated by all the films except SLS-2. Thus the absence of biofilm formation on the film surface permitted enzymatic attack of the materials. Furthermore, inhibition of chemical oxidative degradation in the pure-culture assay was demonstrated for ADM-11, SLS-5, and SLS-10 initial materials and for ADM-4, ADM-7, SLS-8, and SLS-10 heat-treated films. These data suggest that biological and chemical degradation were directly affected by the reduction in oxygen tension on the plastic film surface due to cell mass accumulation. This same phenomenon could be the cause for slow degradation rates in nature.Journal Paper No. J-15061 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa. Project Nos. 0178 and 2889.     

Effects of higher-order structure of poly(3-hydroxybutyrate) on its biodegradation. II. Effects of crystal structure on microbial degradation

As one of a series of studies concerning the relationship between the higher-order structure and the biodegradability of a biodegradable plastic, the effects of the crystal structure of the plastic on microbial degradation were investigated. Bacterial poly(d-(–)-3-hydroxybutyrate) (PHB) films which had a wide range of crystallinity were prepared by the melt-quenching method. Results of the microbial degradation indicated that the development of crystallinity evidently depressed the microbial degradability. From scanning electron microscopy (SEM) observations, it is suggested that the microbial degradation proceeded in at least two manners. One was preferential degradation of the amorphous region leaving the crystalline lamellae intact, which was considered to be a homogeneous enzymatic degradation over the surface. The other was nonpreferential spherical degradation on the surface. The SEMs indicate that the spherical holes were the result of colonization by degrading bacteria. The holes varied in size and number with the change of crystal structure. Therefore, it is considered that the crystal structure of PHB also influenced the physiological behavior of the degrading bacteria on the PHB surface.     

Measurement of the biodegradation of starch-based materials by enzymatic methods and composting

The aim of this study was to evaluate the suitability of in vitro enzymatic methods for assaying the biodegradability of new starch-based biopolymers. The materials studied included commercial starch-based materials and thermoplastic starch films prepared by extrusion from glycerol and native potato starch, native barley starch, or crosslinked amylomaize starch. Enzymatic hydrolysis was performed using excessBacillus licheniformis -amylase andAspergillus niger glucoamylase at 37°C and 80°C. The degree of degradation was determined by measuring the dissolved carbohydrates and the weight loss of the samples. Biodegradation was also determined by incubating the samples in a compost environment and measuring the weight loss after composting. The results indicated that the enzymatic method is a rapid means of obtaining preliminary information about the biodegradability of starch-based materials. Other methods are needed to investigate more accurately the extent of biodegradability, especially in the case of complex materials in which starch is blended with other polymers.     

Thermal Properties and Biodegradability Studies of Poly(3-hydroxybutyrate-co-3-hydroxyvalerate)

For investigating the relationship between thermal properties and biodegradability of poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), several films of PHBV containing different polyhydroxyvalerate (HV) fractions were subjected to degradation in different conditions for up to 49 days. Differential scanning calorimetry (DSC), thermogravimetry (TG), specimen weight loss and scanning electron microscopy (SEM) were performed to characterize the thermal properties and enzymatic biodegradability of PHBV. The experimental results suggest that the degradation rates of PHBV films increase with decreasing crystallinity; the degradability of PHBV occurring from the surface is very significant under enzymatic hydrolysis; the crystallinity of PHBV decreased with the increase of HV fraction in PHBV; and no decrease in molecular weight was observed in the partially-degraded polymer.     

The influence of soil macroinvertebrates on primary biodegradation of starch-containing polyethylene films

The primary biodegradability of polyethylene (PE) films containing different percentages of cornstarch (0–50%) and other additives (prooxidant, oxidized polyethylene) was tested using four species of earthworms (Eisenia fetida, Lumbricus terrestris, Aporectodea trapezoides, Aporectodea tuberculata), three species of cockroaches (Periplaneta americana, Blaberus sp.,Blattella germanica), termites (Reticulotermes flavipes), sowbugs (Porcellio laevis), and crickets (Acheta domesticus). These studies were conducted to elucidate the potential role of soil macroinvertebrates in degrading starch/PE biodegradable plastics. The results of the macroinvertebrate bioassays indicate that crickets, cockroaches, and sowbugs consumed starch-containing PE films most readily. In addition, the degree to which the films were attacked and consumed was directly related to the starch content of the film. Films with oxidized polyethylene and those containing prooxidant (vegetable oil and a transition metal catalyst) were also consumed. None of the four species of earthworms tested or the termites showed any activity toward the starch/polyethylene films. These results have important implications for determining the fate of novel plastic formulations which claim to be biodegradable in natural environments. Studies such as these, coupled with studies on microbial degradation, will help provide the type of information needed to assess the environmental fate of biodegradable starch/PE plastics and fill the voids in the scientific database regarding this rapidly developing field.     

Biodegradation of Epoxy and MF Modified Polyurethane Films Derived from a Sustainable Resource

Mesua ferrea L. seed oil (MFLSO) modified polyurethanes blends with epoxy and melamine formaldehyde (MF) resins have been studied for biodegradation with two techniques, namely microbial degradation (broth culture technique) and natural soil burial degradation. In the former technique, rate of increase in bacterial growth in polymer matrix was monitored for 12 days via a visible spectrophotometer at the wavelength of 600 nm using McFarland turbidity as the standard. The soil burial method was performed using three different soils under ambient conditions over a period of 6 months to correlate with natural degradation. Microorganism attack after the soil burial biodegradation of 180 days was realized by the measurement of loss of weight and mechanical properties. Biodegradation of the films was also evidenced by SEM, TGA and FTIR spectroscopic studies. The loss in intensity of the bands at ca. 1735 cm⁻¹ and ca. 1050 cm⁻¹ for ester linkages indicates biodegradation of the blends through degradation of ester group. Both microbial and soil burial studies showed polyurethane/epoxy blends to be more biodegradable than polyurethane/MF blends. Further almost one step degradation in TG analysis suggests degradation for both the blends to occur by breakage of ester links. The biodegradation of the blends were further confirmed by SEM analyses. The study reveals that the modified MFLSO based polyurethane blends deserve the potential to be applicable as “green binders” for polymer composite and surface coating applications.     

Preparation and Biodegradation of Starch/Polycaprolactone Films

Starch granules were modified with trisodium trimetaphosphate (TSTP) and characterized by P³¹-NMR, FTIR and DSC. Seventy-micron films were prepared from modified starch and polycaprolactone blends by solvent casting technique. Three different types of films—PCL (100% polycaprolactone), MOD-ST/PCL (50% modified starch and 50% polycaprolactone blend) and NONMOD-ST/PCL (50% nonmodified starch and 50% polycaprolactone blends)—were prepared, and their thermal, mechanical, and morphologic properties were investigated to show the increased performance of PCL with the addition of starch and also the effect of modification. It was observed that with the addition of starch the Young's modulus of polycaprolactone was increased and became less ductile, whereas tensile strength and elongation at break values decreased. Biodegradation of these films was inspected under different aerobic environments with the presence of Pseudomonas putida, activated sludge, and compost. It was observed that whereas P. putida had almost no effect on degradation during 90 days, with the presence of activated sludge, considerable deformation of films was observed even in the first 7 days of degradation. In a compost environment, degradation was even faster, and all polymer films were broken into pieces within first 7 days of degradation and no film remained after 15 days.     

Effects of Electron-Beam Irradiation and Ultraviolet Light (365 nm) on Polylactic Acid Plastic Films

Strips of Ca-I [polylactic acid (PLA) monolayer plastic films from Cargill Dow Polymers LLC, Minnetonka, MN] cut in the machine and nonmachine directions were irradiated with an electron beam using a CIRCLE III Linear Accelerator (MeV Industries S.A., Jouy-en-Josas, Cedex, France). The effects of 33-kGy irradiation on the physical properties of the Ca-I strips were studied. In addition, the effects of ultraviolet (UV) light (365-nm) illumination on the degradation of three PLA plastic films, Ch-I (PLA monolayer plastic films from Chronopol, Golden, CO), GII (PLA trilayer plastic films from Cargill Dow Polymers LLC), MN), and Ca-I (PLA monolayer plastic films from Cargill Dow Polymers LLC) were analyzed by a modified ASTM D5208-91 method. Results showed that irradiation had decreased the weight-average molecular weight (M _w), stress at break, percentage of elongation, and strain energy of Ca-I by 35.5, 26.7, 32.3, and 44.8%, respectively (P < 0.01). The shelf life of the irradiated Ca-I strips at 5°C and <20 ± 5% RH was about 6 months. The degradation rate of Ch-I, GII, and Ca-I with no UV light treatment at 55°C and 10% RH was 2512, 5618, and 3785 M _w/week, respectively. Under the UV light illumination (365 nm), the degradation rate of Ch-I, GII, and Ca-I, was 2982, 8722, and 7467 M _w/week, respectively. Hence, the degradation rate of GII and Ca-I was increased 55 and 97% by UV light (P < 0.008), respectively. This trend was not observed in Ch-I because its starting M _w (78,000 g/mol) was close to the tensile strength lost range (50,000 to 75,000 g/mol) of PLA films. To our knowledge, this is the first study to demonstrate that UV light does further enhance the degradation of PLA films.     

Preparation and Characterization of Compatible and Degradable Thermoplastic Starch/Polyethylene Film

The degradability of the compatible thermoplastic starch/polyethylene film was investigated by weight loss percent (WLP), Fourier Transform Infrared (FT-IR) Spectroscopy, and Scanning Electron Microscope (SEM). The compatible film was prepared by using the particles of thermoplastic starch/polyethylene blends that were produced by one-step reactive extrusion. The weight of the film after degradation reduced more than 3% for 30 days and 4% for 60 days. The FTIR results revealed that both starch and polyethylene in the film exhibited varying degrees of degradation. SEM photographs of the films after degradation showed that starch particles in the film disintegrated into smaller particles or separated out of the film surface. Degradation studies demonstrated that the compatible thermoplastic starch/polyethylene film had increased degradability at the given degradable environment. The information implies that this film could be utilized as a degradable plastic.     

Preparation and Properties of Propylene Glycol Plasticized Wheat Protein Isolate Novel Green Films

Novel bio-based green films were prepared using wheat protein isolate (WPI) by solution casting method using Propylene Glycol as a plasticizer for packaging applications. The effect of the plasticizer content (10, 15, 20 and 25 wt%) on mechanical properties (tensile strength, young’s modulus and  % of elongation) was investigated. A thermal degradation and phase transition of the prepared WPI was assessed by means of TGA and DSC analysis. The results showed that the tensile strength and young’s modulus decreased and  % of elongation increased with increasing PG content. The ATR-FTIR and SEM were used for structural characterization and morphology of the films, respectively. FTIR studies reveals that the intensity of the bands corresponding to the amide groups increases with increasing PG content tending to increase protein–PG interactions. Further, the glass transition temperature was decreased and the thermal stability of the WPI was found to be increased by plasticization. The overall thermal stability of the films was improved and is attributed to the increase in mobility of the polymer chains.     

Effects of Temperature and Relative Humidity on Polylactic Acid Plastic Degradation

Three high molecular weight (120,000 to 200,000 g mol^–1) polylactic acid (PLA) plastic films from Chronopol (Ch-I) and Cargill Dow Polymers (GII and Ca-I) were analyzed for their degradation under various temperature and relative humidity (RH) conditions. Two sets of plastic films, each containing 11 samples, were randomly hung in a temperature/humidity-controlled chamber by means of plastic-coated paper clips. The tested conditions were 28, 40, and 55°C at 50 and 100% RH, respectively, and 55°C at 10% RH. The three tested PLA films started to lose their tensile properties when their weight-average molecular weight (M _w) was in the range of 50,000 to 75,000 g mol^–1. The average degradation rate of Ch-I, GII, and Ca-I was 28,931, 27,361, and 63,025 M _w/week, respectively. Hence, GII had a faster degradation rate than Ch-I and Ca-I under all tested conditions. The degradation rate of PLA plastics was enhanced by the increase in temperature and relative humidity. This trend was observed in all three PLA plastics (Ca-I, GII, and Ch-I). Of the three tested films, Ch-I was the first to lose its mechanical properties, whereas Ca-I demonstrated the slowest loss, with mechanical properties under all tested conditions.     

A spectrophotometric method for detection of enzymatic degradation of thin polymer films

An assay method has been developed for monitoring the enzymatic degradation of thin films of translucent polymers. The method was based on the observation that when a solution-cast film of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) was exposed to a solution of a depolymerase fromPseudomonas lemoignei, the surface of the film roughened and the film became visibly turbid. This increase in turbidity could be measured spectrophotometrically and was reproducible during the initial stage of degradation. Turbidity correlated very closely with film weight loss early in the degradation but reached a maximum value before extensive degradation had taken place. For a given set of films, this correlation was independent of the concentration of the enzyme used, although it did vary with the mode of enzyme exposure. The turbidity was associated with the exposure of crystalline domains due to the removal of amorphous material from the film surface. The increase in crystallinity at the surface was verified by attenuated total reflectance infrared spectroscopy (ATRIR). In conjunction with SEM, weight loss, and ATRIR, the film turbidity assay provided much semiquantitative insight into the mechanism of the enzymatic degradation reaction. This assay was used to study the enzymatic degradation of films of PHBV solution blended with cellulose acetate esters (CAE). The presence of only 25% of CAE of degree of substitution 2.9 severely hampered the enzymatic degradability of PHBV, a result which is consistent with the environmental degradation of these same samples exposed to activated sludge.     

Hydrolytic and enzymatic degradation of poly(γ-glutamic acid) hydrogels and their application in slow-release systems for proteins

The hydrolytic and enzymatic degradation of newly developed hydrogels, produced by cross-linking purified poly(-glutamic acid) (PGA) with dihaloalkane compounds, was studied and is reported in this paper. Analysis of hydrolysis of the hydrogel as a function of pH indicated that the hydrolysis occurred slowly at neutral pH, but fast in both acidic and alkaline solutions, while the polymer could be hydrolyzed rapidly only in acidic solutions. The ester bonds were more sensitive to hydrolysis than peptide bonds. The biodegradability of the hydrogel and polymer was further confirmed when enzymatic degradation was studied by three enzymes (cathepsin B, pronase E, and trypsin), which were able to cleave both ester and peptide bonds gradually. A slow-release system for porcine somatotropin (pST) formed by using the hydrogel as matrix to entrap the hormone was evaluatedin vitro andin vivo. Results demonstrated that the hydrogel was able to release the hormone for a period of 20–30 days and indicated its potential application in slow-release systems for bioactive materials, especially macromolecules, such as peptides and proteins.     

Process,Characterization and Biodegradability of Aliphatic Aromatic Polyester/Sisal Fiber Composites

The biodegradability, morphology, and mechanical properties of composite materials made of Poly(butylene adipate-co-terephthalate) (PBAT) and sisal fiber (SF) were evaluated. Composites containing acrylic acid-grafted PBAT (PBAT-g-AA/SF) exhibited noticeably superior mechanical properties due to greater compatibility between the two components. The dispersion of SF in the PBAT-g-AA matrix was highly homogeneous as a result of ester formation between the carboxyl groups of PBAT-g-AA and hydroxyl groups in SF and the consequent creation of branched and cross-linked macromolecules. Each composite was subjected to biodegradation tests in Rhizopus oryzae compost. Morphological observations indicated severe disruption of film structure after 60 days of incubation, and both the PBAT and the PBAT-g-AA/SF composite films were eventually completely degraded. Water resistance of PBAT-g-AA/SF was higher than that of PBAT/SF, although weight loss of composites buried in Rhizopus oryzae compost indicated that both were biodegradable, even at high levels of SF substitution. The PBAT-g-AA/SF films were more biodegradable than those made of PBAT, implying a strong connection between these characteristics and biodegradability.     

Oxygen and Water Barrier Properties of Coated Whey Protein and Chitosan Films

Films of whey protein and chitosan acetic acid salt have lower oxygen permeability than, for example, ethylene-co-vinylalcohol under dry conditions, but water and water vapor seriously impair the gas barrier properties. To reduce the oxygen permeability at 90% relative humidity and the water-vapor transmission rate at 100% relative humidity, the films were coated with an alkyd, a beeswax compound, or a nitrocellulose lacquer. Permeability and transmission rate measurements were performed in accordance with standard methods and showed that the beeswax compound and the nitrocellulose were appropriate as water-vapor barriers. Overall migration to water was measured after 10 days exposure time, with the coated surface exposed to the water, showing that the alkyd-coated and the nitrocellulose-coated films were both below the safety limit for food contact. Water absorbency tests, performed by the Cobb method, showed that the films coated with the beeswax compound or with nitrocellulose lacquer exhibit lower absorbency than the alkyd-coated films.