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PACKAGING TECHNOLOGY AND SCIENCE Packag. Technol. Sci. 2007; 20: 155–163 Published online 12 September 2006 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/pts.750

Study of Mechanical Properties, Oxygen Permeability and AFM Topography of Zein Films Plasticized by Polyols By Babak Ghanbarzadeh,1* A. R. Oromiehie,2 Mohamad Musavi,3 Pasquale Massimiliano Falcone,4 Zahra Emam D-Jomeh3 and Elhame Razmi Rad3 1

Department of Food Science and Technology, Faculty of Agriculture, University of Tabriz, Abrasan Street,Tabriz, Iran Iran Polymers and Petrochemical Institute, Pazhoohesh Street, PO Box 14965/159,Tehran, Iran 3 Department of Biosystems Engineering, Faculty of Agriculture, University of Tehran, Daneshkadeh Square, PO Box 31587-78659, Karaj, Iran 4 Department of Food Science (DISA), University of Foggia, 71100 Foggia, Italy 2

The use of plastic for packaging has grown extensively in recent years. In this context, biodegradable films can be a source of energy saving and an important issue for environmental protection. Zein protein (prolamin of corn) is one of the best biopolymers for edible film making and polyols are convenient plasticizers for biopolymers. Polyols (sorbitol, glycerol and mannitol) at three levels (0.5, 0.7 and 1 g/g zein) were used as plasticizers and the tensile properties, oxygen permeability (OP) and AFM topography of zein films were studied. Films plasticized by sorbitol had a relatively higher ultimate tensile strength (UTS) than films containing glycerol and mannitol at low levels of plasticizers (0.25, 0.7 g/g zein). There was no significant difference between the strain at break values (SB) of films plasticized by sorbitol and glycerol at low levels of plasticizers, while films plasticized by sorbitol had higher SB than the films containing glycerol and mannitol at a high level of plasticizer (1 g/g zein). Pure zein films had low oxygen permeability (OP), and increasing the plasticizer level to 0.5 g/g zein decreased OP values in films containing sorbitol and glycerol. Films containing sorbitol and mannitol had the lowest and highest OP values, respectively. AFM images were used to evaluate the surface morphology (qualitative parameter) and roughness (quantitative parameter) of zein films. Films plasticized by glycerol had smoother surfaces and a lower roughness parameter (Rq). No relationship between OP values and the roughness of the zein films was observed. Copyright © 2006 John Wiley & Sons, Ltd. Received: 5 April 2006; Revised 14 June 2006; Accepted 16 June 2006 KEY WORDS:

zein edible films; polyols; mechanical properties; atomic force microscopy; roughness; oxygen permeability

* Correspondence to: B. Ghanbarzadeh, Department of Food Science and Technology, Faculty of Agriculture, University of Tabriz, Abrasan Street, Tabriz, Iran. E-mail: [email protected]

Copyright © 2006 John Wiley & Sons, Ltd.

Packaging Technology and Science

B. GHANBARZADEH ET AL.

density polyethylene (HDPE), which are not good oxygen barriers.9 The OP values of zein films at low RH approach those of the best synthetic oxygen barriers, ethylene–vinyl alcohol copolymer (EVOH) and polyvinylidene chloride (PVDC). The casting zein films had higher oxygen permeability than carbon dioxide permeability.8 The RH of the environment affects the oxygen barrier properties of zein and other protein films considerably. Moisture has a plasticizing or swelling effect on hydrophilic polymers, which increases gas permeability.10 With the advent of atomic force microscopy (AFM), new possibilities have been opened up to evaluate the features of and biopolymers. Nanoscale measurements by AFM allow the influence of different factors on the hardness, elasticity and permeability of the film surface to be quantified, which is extremely useful for the design of high-performance edible food packaging.11 Measurements of the topography and roughness can be undertaken with extremely high resolution. This technique has been used to characterize the surface morphology of whey protein films.11,12 The structure, development and orientation of cast and stretched resin films were observed with AFM by Lai and Padua.13 AFM images of cast films showed uniform deposition of components with no salient structural features. The uniform deposition of zein in cast films explained their surface gloss. The main objectives of this work were to study the effects of polyols on the tensile properties, oxygen permeability (OP) and topography of zein films and the correlation between roughness of the surface and oxygen permeability (OP).

INTRODUCTION Using edible films and coatings in the food industry has the following advantages: controlling moisture, oxygen, aroma and oil transferrance in food systems, carrying functional ingredients (e.g. antimicrobial agents to improve safety and stability of foods, antioxidants to prevent lipid oxidation, and flavourings and pigments to improve food sensory properties), protection of food from mechanical damage and reducing synthetic packaging materials.1,2 Several factors, such as additives (e.g. plasticizers and cross linking agents), solvent type, temperature, relative humidity (RH), film production method and lamination, affect the tensile properties of zein films.2,3 Generally, plasticizers decrease the ultimate tensile strength (UTS) and increase the strain at break (SB) of zein films.4 The UTS of zein films was increased by adding cross-linking agents to the alcoholic film forming solutions.5,6 Rakotonirainy and Padua18 studied the effects of lamination and coating with drying oils (tung oil, linseed and soy bean oil) on tensile properties. Laminated films were clearer, tougher and more flexible than non-treated films. Coating process increased UTS and SB. The coating acted as a composite layer which prevented crack propagation and it increased film strength. The ductility of resin zein films increased at high RH,7 e.g. SB increased from 12% to 30% and Young’s modulus decreased from 267 to 150 MPa when the RH increased from 50% to 98%, while the UTS was not significantly affected (p > 0.05) affected. This was attributed to zein plasticization by moisture. The tensile properties of resin zein films containing fatty acids were affected by fatty acid content.4 Increasing the oleic acid level from 0.5 to 1.0 g/g zein decreased UTS and increased SB values of zein films. Films containing 0.5 g oleic acid/g zein had 9 MPa UTS and 12% SB, while films prepared with 0.6 g oleic acid/g zein had 6 MPa UTS and 44% SB. The ability of films and coatings to diminish oxygen transferrance is important for controlling the respiration of fresh fruits and vegetables. In addition, oxygen causes lipid oxidation, which decreases food quality and shortens food shelflife.1,8 In general, at low to intermediate RH, protein films have lower oxygen permeability (OP) than low-density polyethylene (LDPE) and high-


MATERIALS AND METHODS Materials Zein, regular grade, was obtained from Freeman Industries Inc. (Tuckahoe, NY). Other materials were: sorbitol (MW 182), mannitol (MW 182), glycerol (MW 92) and Ca(NO3)2 2H2O, purchased from the Merck Corporation; ethanol (95%) obtained from Razi Corp. and ethanol 80% prepared from it.

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Packag. Technol. Sci. 2007; 20: 155–163 DOI: 10.1002/pts

PROPERTIES OF ZEIN FILMS PLASTICIZED BY POLYOLS


Preparation of zein films Zein dispersions were obtained by dissolving zein to 20%(w/v) in warm (80°C) aqueous ethanol 80%. Polyols were added to the solution at 0.5, 0.7 or 1 g/g zein and stirred in a mixer at 300 r.p.m. for 10 min. Zein–plasticizer dispersions were precipitated by the addition of cold water (5°C) as result of hydrophobic aggregation. Resins were collected as soft solids and kneaded in a mixer for separation of the remaining alcohol and water to obtain cohesive mouldable resins. The resins were rolled and then pressed in a hot press (80°C, 25 MPa) between two metal surfaces to form zein films, which were were dried under ambient conditions for 48 h. Film thickness was measured using a dial gauge micrometer (B. C. Ames Co., Waltham, MA). Films of ~0.2 (±0.03) mm thickness were used in the experiments.

Figure 1. Measurement of oxygen permeability by the equal-pressure method. sure on the two sides is equal but oxygen partial pressure is different. Under the function of oxygen concentration difference, oxygen transits through the film and is diverted into the sensor by nitrogen carrier gas. Testing is complete when the concentration of oxygen in the nitrogen side atmosphere is constant. The oxygen transmission rate (O2 TR) of the film can be calculated according to the oxygen quantity that is accurately measured by the sensor in nitrogen carrier gas. Oxygen permeability (OP) was calculated by dividing the steady-state transmission rate of oxygen gas by the difference in oxygen partial pressure between the sides of the film and multiplying by mean thickness:

Mechanical properties A universal testing machine, Model M/10 (MTS Co.) was used for mechanical properties measurement. The specimens were preconditioned for 48 h at 25°C and 50% RH inside a desiccator containing saturated solutions of calcium nitrate. Films of ~0.2 mm thickness were prepared by cutting into strips 5.0 mm wide. Film strips were placed in the grips of the testing machine, which were set at an initial grip separation of 25 mm. The tensile properties measured included ultimate tensile strength (UTS), strain at break (SB), yield tensile strength (YTS) and Young’s modulus (E). Three replicates were run for each film treatment.

P=

where P = oxygen permeability, Q = quantity of oxygen, X = film thickness, A = area of permeation, t = permeation time and ∆p = the partial pressure difference of the permeant gas across the film. Permeability was expressed in cm3/m/cm2/d/kPa.

Oxygen permeability The measurements of oxygen permeability were carried out based on the equal pressure method (ASTM D3985-81)14 using an Ox-Tran apparatus (Mocon Inc., Minneapolis, MN) at 25°C. The samples were conditioned for 3 days at 50% RH before measurements. In the equal pressure method, the test specimen is held such that it separates two sides of a test chamber. One side is exposed to a nitrogen atmosphere while the other is exposed to an oxygen gas (Figure 1). The pres-


Q X × t.A. ∆p

Atomic force microscopy (AFM) The AFM provided topographic images and roughness of scanned samples. The surface morphology of the films was analysed using dynamic

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SPM acquisition mode and contact SPM acquisition mode (AFM, Nanotec Electronica S.L., Madrid, Spain) with different scan sizes: 1. 2.5 × 2.5 mm scan size and a 74.1 nm vertical range. 2. 5 × 5 mm scan size and a 239.3 nm vertical range. 3. 10 × 10 mm scan size and a 1.4 mm vertical range. 4. 30 × 30 mm scan size and a 2 mm vertical range. Two kinds of cantilevers were used. An NSC12 cantilever (MicroMasch) with a spring constant of 14 N/m, and an AC160TS-2 cantilever (Olympus) with a spring constant of 42 N/m. Three different images were taken of each sample at four scan sizes. The basic principle of AFM is the recording of the deflection of a cantilever over each x,y coordinate of the sample as it is scanned under the probe. The x,y-registered data are then assembled into a 3D surface map. The resulting image from the AFM is a map of forces detected over each point on the sample.11 The most frequently quantitative parameters of roughness, Rq, were calculated using the data from the images with appropriate software. Average roughness (Ra) is the arithmetic mean of the height – deviations from the profile mean value (Z ). Ra is written as: Ra

∑ =

N

Figure 2.Typical load (tensile stress)–strain (elongation) curve of resin zein film. 10 software. General linear model (GLM) procedures were utilized for analysis of variance (ANOVA). The Duncan’s multiple range test was used to determine any significant difference between specific means at p < 0.05.

RESULTS AND DISCUSSION

Zi − Z

i =1

N

Mechanical properties

where:

∑ Z=

N

i =0

A typical tensile stress (load)–strain curve for zein films is shown in Figure 2. Zein films are generally brittle and require the addition of plasticizer to improve their flexibility.4,5,13,15–17 Plasticizers are low molecular mass organic compounds and they are added to soften rigid polymers. The stress–strain curve exhibited greater deviation from linearity as the concentration of plasticizers increased. The effects of different polyolic plasticizers at three levels (0.5, 0.7 and 1 g plasticizer/g zein) on the mechanical properties are illustrated in Figure 3A, B, C. Significant differences (p < 0.05) were found between films containing different polyols for ultimate tensile strenght, strain at break and Young’s modulus. Ultimate tensile strength (UTS) is the maximum tensile stress that a material can sustain before the onset of permanent deformation or failure.

Zi

N and Zi = the height values in profile (histogram; – nm), Z = arithmetic means of heights (nm) and N = number of data points in the profile. The root mean square (RMS) of roughness (Rq) is the root of mean square of height deviations from the mean of heights: N

∑ (Z

i

Rq =

i =1

− Z)

2

N

Statistical analysis Statistical analyses for tensile properties and oxygen permeability were performed using Spss


158


Ultimate Tensile Strenght (MPa)


(A)

25

UTS of polymeric materials decreases by plasticization, but a low level of plasticizer is needed to increase association within polymer chains, and zein films without plasticizer are very brittle. These results were agreement with the study of Lai and Padua,17 who indicated that the addition of palmitic acid in the ratio 0.5 g/g zein or stearic acid:0.25 g/g zein increased the tensile strength of the zein sheets substantially. Films plasticized by sorbitol had a relatively higher tensile strength than films containing glycerol and mannitol at 0.25 g/g zein, but at a higher plasticizer level (1 g/g zein) films containing glycerol had the highest UTS. The low UTS of films plasticized by mannitol could be attributed to low solubility in zein solution and rapid crystallization of mannitol in high concentrations. Films containing mannitol had a chalky white appearance that made those unfavourable. Strain at break (SB) is the maximum elongation (before failure occurred) divided by the original sample length and usually indicated as percentage tensile strain. SB shows the flexibility and extensibility of films and is important for the shaping of films. Increasing the plasticizer levels increased the SB of zein films at all samples. This could be attributed to an increase of chain mobility and lubrication in the film matrix. Films plasticized by sorbitol had a higher SB than films plasticized by glycerol and mannitol. Films plasticized by mannitol had a lower SB than films plasticized by glycerol and sorbitol. Overall, the elongation of zein films was small. Within 1 week, loss of flexibility was observed in all the films. This could be attributed to migration of plasticizers to the surface of new zein films, due to the relatively hydrophobic nature of the zein films matrix.4 Young’s modulus (elastic modulus, E) is the ratio of tensile stress over tensile strain and is determined from the slope of the stress–strain curve at the elastic limit. It is a measure of the force necessary to deform a test specimen and is related to the rigidity of material. Increasing the polyolic plasticizers level from 0.5 to 1 g/g zein decreased E of zein films continuously. Zein films containing mannitol and sorbitol had the highest and lowest E, respectively. As films containing sorbitol showed higher UTS and SB than those containing glycerol and mannitol, it seems that sorbitol is a better plasticizer than glycerol and mannitol, because the film is considered as a good film that

g e

d

20

a

15

a b

10

d

Sorbitol

f

C

Mannitol Glycerol

5

0 0.5

0.7

1

plasticizer level (g/g zein)

Strain at Break (%)

e

(B)

4 3.5

f

d

c

3

a

2.5 2

a

a

a

b

Sorbitol

1.5

Mannitol Glycerol

1 0.5 0 0.5

0.7

1

Plasticizer level (g/g zein)

Young moduluse (Mpa)

1800

a b a (C)

1600

e

e f d

f

g

1400 1200 1000 800

Sorbitol

600

Mannitol

400

Glycerol

200 0 0.5

0.7

1

Plasticizer level (g/g zein)

Figure 3. Effect of plasticizer levels and type on tension properties of zein films: ultimate tensile strength (A), strain at break (B) and Young’s modulus (C). Columns with the same letter are not significantly different (p < 0.05). Increasing the plasticizers levels increased the UTS of zein films in all the specimens, except when mannitol content increased from 0.7 to 1 g/g zein (Figure 3). This could be attributed to the increase of polymer chains association in films matrix by plasticization, up to certain level. In general, the



159




Table 1. Oxygen permeability of zein filmsa (cm3/m/cm2/d/kpa) Plasticizer content (g/g zein)b Film type Pure Zein Zein Zein a b

zein + sorbitol + mannitol + glycerol

0.5

0.7

1

18.6167 ± 0.2714 a 15.9933 ± 0.7283 a 40.8933 ± 1.7616 d 17.3533 ± 0.2312 a

21.8367 ± 1.0013 b 47.3967 ± 2.9593 e 31.5633 ± 1.7652 g

26.2667 ± 1.9083 c 51.4433 ± 2.1548 f 35.4500 ± 0. 9753 h

Values are average of five samples ± standard deviation. Means with the same letter are not significantly different ( p < 0.05).

shows high SB and UTS. A film that shows high UTS and low SB is considered a brittle film, and a film that has high SB and low UTS is considered weak film.

the film matrix. Zein films containing sorbitol and mannitol had lowest and highest OP, respectively. Mannitol had very low compatibility with zein and did not distribute in the zein matrix. Factors affecting the OP of plasticized films are the physical state and the molecular weight of the plasticizer, the chemical interaction between plasticizer and oxygen, and the type of film structure (polymer crystallinity, density, orientation, molecular weight and cross-linking).1 Glycerol has a lower molecular size than sorbitol, so that it has a higher plasticizing effect and as a result films plasticized by glycerol had greater OP value than films containing sorbitol. Sorbitol can create higher numbers of hydrogen bonds in the film matrix. An increase of hydrogen bonds can cause a decrease of OP in films. Also, sorbitol is less hygroscopic than glycerol. In addition, glycerol is liquid and sorbitol is a solid plasticizer, and this factor can affect the plasticization process.

Oxygen permeability The oxygen permeability (OP) values of different resin zein films are presented in Table 1. All the samples had relatively low OP values (15.99– 51.44 cm3/m/cm2/d/kpa). The oxygen and carbon dioxide permeability values of zein films prepared from the casting method were previously reported to be low at 0–50% RH8 but the OP of resin zein films has not been reported. The OP of casting zein film at 0% RH was one to two orders of magnitude less than methyl cellulose and hydroxypropyl cellulose films at the same test temperature and RH.18 However, they were higher than those of wheat gluten and collagen films.19,20 Oxygen molecules apparently can more readily permeate through the zein helical conformation than through the highly cross-linked gluten structure or the highly associated fibrillar nature of collagen. As shown in Table 1, resin zein film without plasticizer had a low OP, and increasing the plasticizer level to 0.5 g/g zein decreased the OP in films containing sorbitol and glycerol, but this difference was not significant (p < 0.5). It was probably due to an increase of association between polymer chains in low levels of plasticizer. Increasing polyols levels from 0.5 to 0.7 and 1 g/g zein increased OP values drastically. This could be attributed to an increase of biopolymer chain mobility and creation of void spaces in


Atomic force microscopy The atomic force microscopy (AFM) technique is a powerful tool for studying surfaces and has been used to provide qualitative and quantitative information about biopolymers at the nanometer scale that are often inaccessible by any other experimental technique.21 AFM can be useful to identify structural differences of films prepared by different methods. The morphology (qualitative parameter) and roughness (quantitative parameter) of the zein films were analysed by AFM. Topography images

160




(Gl)

(GL)

6.0µ m

(S)

(S)

6.0µ m (M)

(M)

6.0µ m

Figure 4. AFM images of zein resin films containing 0.7 g g zein different polyolic plasticizers at 30 × 30 mm scan size (Gl, glycerol; S, sorbitol; M, mannitol).These three images were chosen as representatives from each sample scanned at different scan size.


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bly due to the use of low levels of plasticizers for film making and the increase of chain association with low amounts of plasticizers. Films plasticized by sorbitol had the best UTS and SB and films containing mannitol did not show suitable tensile properties. Adding plasticizers to 0.5 g/g zein decreased the OP values in films containing sorbitol and glycerol (none significantly, p < 0.5). Increasing polyols from 0.5 to 0.7 and 1 g/g zein increased the OP of films considerably. This could be attributed to an increase of biopolymer chain mobility and, as a result, an increase in gas permeability. Films plasticized by sorbitol had the lowest OP. AFM surface analysis showed that films containing glycerol had smoother surfaces than other films. Films containing mannitol and glycerol had the highest and lowest roughness (Rq), respectively. There was no correlation found between the Rq and OP of zein fims. As a result, mechanical properties and oxygen permeability are very important parameters for films used in food packaging. These factors were affected by plasticizer levels and types. It is therefore important to determine the optimum level and type of plasticizers for production of high quality zein films for use in food coating or packaging.

– Table 2. Comparison of Z and Rq values obtained from AFM images of different films Film type Zein Zein Zein Zein Zein Zein Zein Zein Zein Zein Zein Zein

+ + + + + + + + + + + +

sorbitol sorbitol sorbitol sorbitol mannitol mannitol mannitol mannitol glycerol glycerol glycerol glycerol

Scan size (mm)

– Z (nm)

Rq (nm)

× × × × × × × × × × × ×

38.09 95.65 1000.26 1487.40 70.76 410.04 487.94 595.20 37.06 317.51 402.46 592.21

14.47 53.75 187.81 279.34 20.21 93.53 135.89 295.59 13.62 61.34 72.65 210.43

2.5 5 10 30 2.5 5 10 30 2.5 5 10 30

2.5 5 10 30 2.5 5 10 30 2.5 5 10 30

showed that films containing glycerol had a smoother surface than those containing sorbitol and mannitol (Figure 4). Films plasticized by mannitol had the highest incidence of pin-holes and projections. Table 2 summarizes the roughness parameter (Rq) of different zein films in different scan sizes. As shown, in all scan sizes films containing glycerol had the lowest Rq and films plasticized by mannitol had the highest Rq. Herrmann and co-workers11 used AFM for evaluating the surfaces of edible films produced with whey protein concentrate (WPC). The effects of WPC and plasticizer concentration were characterized in terms of water vapour permeability (WVP) and roughness parameters. Their results showed a relationship between water vapour permeability and area roughness. The results of our research did not clearly show a relationship between the oxygen permeability and roughness of different zein films, because zein films containing sorbitol had lower OP values than films plasticized by glycerol, but higher roughness.

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CONCLUSION Increasing the plasticizers increased the UTS of the zein films containing polyolic plasticizers in all specimens, except when mannitol content increased from 0.7 to 1 g/g zein. This was proba-


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