Skip to main content
NCBI home page
ACS AuthorChoice logoLink to ACS AuthorChoice
. 2025 Nov 6;41(45):30188–30195. doi: 10.1021/acs.langmuir.5c03546

Influence of Interfacial Interactions on the Formation of Zein–Ethyl Cellulose Composite Films

Jeannine Bonilla 1,*, Yuyang Zhang 1, Loong-Tak Lim 1,*
PMCID: PMC12633697  PMID: 41198052

Abstract

In this study, submicrometer zein particle suspensions (ZPS) were prepared by antisolvent precipitation from aqueous ethanol (Eth, 90% w/w) or isopropanol (Iso, 90% w/w) solutions. The resulting ZPS were incorporated into an ethyl cellulose solution (EC, 4% w/w), followed by casting to obtain continuous composite films. The zein particle size distribution was evaluated by laser diffraction, confocal laser scanning microscopy, and atomic force microscopy analyses. The ZPS prepared from 90% aqueous ethanol had zein particles with a smaller average particle size (Dv50 = 0.56 μm) compared to those prepared from 90% aqueous isopropanol (Dv50 = 3.74 μm). Transparent and coherent zein–EC composite films exhibited different scanning electron microscopy features depending on the types of solvent (ethanol = Eth or isopropanol = Iso) and surfactants (Span 85 or Tween 80) used. Atomic force microscopy analysis confirmed the affinity between zein particles and the EC phase when prepared in pure ethanol or in ethanol- or isopropanol-based formulations containing Tween 80. The Hansen Solubility Parameter concept was applied to elucidate how the compatibility of various components affected the microstructures and material properties of the zein–EC films. Overall, results from this study have contributed to increased understanding of the effects of solvents and surfactants on the formation of submicrometer zein particles and zein–EC composite films.

graphic file with name la5c03546_0010.jpg

graphic file with name la5c03546_0008.jpg

1. Introduction

Zein, a major storage protein in corn, is biocompatible, biodegradable, amphiphilic, and has self-assembling properties. − As a prolamin, zein is soluble in aqueous ethanol (60% to 90% v/v ethanol) and isopropanol (65% to 90% v/v isopropanol) solutions. , Due to its relatively hydrophobic nature, zein can be converted into spherical colloidal nanoparticles from aqueous alcohol solutions through antisolvent precipitation. − ,− This method, also known as liquid–liquid dispersion or phase separation, involves the addition of a nonsolvent to a polymer solution (or vice versa) to induce supersaturation, causing the polymer to precipitate and form particles with diameters ranging from microns to nanometers. , The size distribution and morphology of the zein particles, prepared using the antisolvent method, are affected by the alcohol type, pH, initial concentrations of alcohol and zein, as well as the presence of additives (e.g., surfactants, plasticizers).

Zein protein can form films that exhibit moisture barrier properties but lack flexibility and toughness. These shortcomings can be overcome by combining zein with other polymers, such as ethyl cellulose (EC). EC is a food-grade polymer with desirable properties (i.e., biocompatible, biodegradable, optically transparent, and gas/moisture barrier) that are ideal for food, pharmaceutical, and biomedical applications. −

Zein and EC, respectively, are well-known as film and coating formers; however, a composite system based on zein nanoparticle suspensions embedded in EC film matrices remains unexplored. − ,,− In this study, a novel approach for the preparation of narrow zein particle size distributions was achieved by the antisolvent method. The resulting ZPS was subsequently incorporated into an alcoholic EC solution to form transparent, coherent composite films through controlled solvent evaporation. Interfacial interactions were systematically studied, including electrostatic repulsion, interfacial tension, and hydrogen bonding, which collectively govern the formation of submicrometer zein particles and their assembly into zein–EC composite films. By evaluating solvent and surfactant effects, the stabilization mechanisms of zein suspensions during antisolvent precipitation and zein–EC composite film formation were investigated.

The specific objectives of this study are to (1) develop stable submicrometer zein particles by the antisolvent precipitation method; (2) study the interfacial interaction driving the formation of zein–EC composite films at 50 °C (minimum film-forming temperature); (3) examine the microstructure of the composite films; and (4) apply Hansen Solubility Parameters (HSP) to elucidate how components’ solubility/compatibility affect the composite film formation.

2. Materials and Methods

2.1. Materials

Zein prolamine from corn (grade Z3625), EC (22 cP, 48% ethoxy content), Span 85 (MW = 957.52 g/mol, HLB = 1.2), and sodium carbonate (Na2CO3, pH 11.2) were purchased from Sigma-Aldrich Canada (Oakville, ON, Canada). Ethanol (HPLC grade), isopropanol (molecular biology grade), sodium hydroxide (NaOH), hydrochloric acid (HCl), and Tween 80 (MW = 1310 g/mol, HLB = 15) were supplied by Thermo Fisher Scientific (Mississauga, ON, Canada).

2.2. Preparation of Submicrometer Zein Particles

Zein (4% w/w) solutions were prepared using an antisolvent process. The zein powder was dispersed in either aqueous ethanol or aqueous isopropanol solvents and stirred with a magnetic stirrer at 450 rpm for 2 h at 22 °C to form homogeneous zein solutions. Preliminary experiments using 70% and 80% (w/w) aqueous alcohol solutions led to rapid precipitation and the formation of heterogeneous zein aggregates. In contrast, 90% (w/w) ethanol or isopropanol resulted in stable suspensions with zein particles of a narrower size distribution. Therefore, aqueous ethanol and isopropanol at the 90% (w/w) level were selected for subsequent experiments. Separately, the deionized water was adjusted to pH 10 using Na2CO3 with a pH meter (Accumet Excel XL20, Fisher Scientific, Ottawa, ON, Canada). Then, the zein solution was added slowly, at a rate of approximately 3 mL/min, to deionized water (zein:water 1:3 w/w, pH 10) under constant stirring at 450 rpm at 22 °C to induce zein particle precipitation. The resulting zein suspension, henceforth termed ZPS, remained stable for 3 days, during which phase separation was not observed.

2.3. Preparation of Zein–EC Composite Films

EC (4% w/w) solution was prepared by dissolving the polymer in either pure ethanol (Eth) or pure isopropanol (Iso). To prepare the composite-forming suspension, 4 mL of ZPS (4% w/w), prepared in Section , was added to 100 mL of EC solution (4% w/w) and stirred for 2 h at 22 °C. Then, either Span 85 (S, hydrophobic nature) or Tween 80 (T, hydrophilic nature) was added at 20% w/w (based on total polymer content) and stirred for 1 h at 22 °C, resulting in a final surfactant concentration of 0.83% (w/w) in the final film-forming suspension. The resulting suspensions were cast on glass plates (33 cm × 20 cm) and dried in an environmental chamber (model MLR-350H, Sanyo Electric Co., Ltd., Osaka, Japan) at 50 °C and 65% relative humidity to form the final composite film samples.

2.4. Characterization of the Zein Particle Suspension

2.4.1. ζ-Potential and Particle Size Distribution

ZPS produced in either aqueous ethanol (90% w/w) or aqueous isopropanol (90% w/w) was analyzed at 25 °C using particle electrophoresis measurements (Zetasizer Nano ZS, Malvern Instruments Ltd., Malvern, UK). Particle size analysis was carried out using a laser diffractometer (Mastersizer 3000; Malvern Instruments, Malvern, UK) coupled to a Hydro Medium Volume automated dispersion unit. Aqueous ethanol or isopropanol solvent (90%, w/w; refractive index = 1.36 ± 0.01) was used as the dispersant depending on the ZPS sample. The ZPS prepared in Section was slowly added to the automated dispersion unit (Hydro MV, Malvern Panalytical, Malvern, UK) and stirred at 2000 rpm until an obscuration of 10% was obtained. Mie theory was applied to invert the scattering pattern into a particle size distribution, considering a refractive index of 1.49 ± 0.01 for zein and 0 as the absorbance value. Results from laser diffraction were expressed in terms of volume distribution (Dv50, i.e., volume average particle size), which is equivalent to the particle diameter. Measurements were repeated at least three times.

2.4.2. Confocal Laser Scanning Microscope

To visualize the submicrometer zein particles, 30 μL of rhodamine B fluorescent dye was added to 10 mL of ZPS prepared in aqueous ethanol or isopropanol (90%, w/w). A drop of either suspension sample was placed on a glass slide and examined using a confocal laser scanning microscope (Zeiss LSM 980, Carl Zeiss Canada Ltd., Toronto, ON, Canada). The microscope was equipped with an Airyscan 2, containing 32 concentrically arranged detection channels, with a maximum resolution of 120/120 nm. The images were obtained and processed using image analysis software (ZEISS ZEN Microscopy, version 3.1, Carl Zeiss Microscopy Deutschland GmbH, Oberkochen, Deutschland).

2.4.3. Atomic Force Microscopy Analysis

Zein particles were examined by using an atomic force microscope (AFM+, Anasys Instruments Inc., Santa Barbara, CA, USA) in tapping mode with a cantilever probe (part no. ACTA-10, L = 125 μm, W = 30 μm, frequency = 200–400 kHz, spring constant = 13–77 N/m; AppNano, Mountain View, CA, USA). A 10 μL ZPS sample, prepared in aqueous ethanol or isopropanol (90%, w/w), was deposited onto a mica surface (AFM mica disc, 10 mm, quality grade V1, Ted Pella Inc., Redding, CA, USA) and allowed to dry completely prior to analysis. The analysis of the resulting particle height image (15 μm × 15 μm) was performed with an image analysis software (Gwyddion, version 2.63, www.gwyddion.net), where the statistical function height distribution was applied. This function shows the peak count (ρ) as a function of the peak height (z). In addition, the average height distribution was analyzed.

For the composite film samples, surface topography and phase images (10 × 10 μm) were analyzed in tapping mode (AFM+, Anasys Instruments Inc., Santa Barbara, CA, USA). Mean roughness (R a), i.e., the value of height irregularities, and height particle distribution, along with their corresponding average values, for all composite film samples were analyzed using image analysis software (Gwyddion, version 2.63, www.gwyddion.net).

2.4.4. Hansen Solubility Parameters (HSPs)

Compatibility between the composite film components (i.e., zein, EC, ethanol, isopropanol, aqueous ethanol, aqueous isopropanol, Tween 80, and/or Span 85) was evaluated using the HSPs. This compatibility was analyzed based on R A (MPa1/2), which represents the straight-line distance between two components (e.g., polymer–solvent or polymer–surfactant) in the HSP space. Thus, the R A value indicates the difference in the energy contributions from dispersion forces (δd), dipolar intermolecular forces (δp), and hydrogen bonds (δh) between each component, as described in eq :

RA={4(δd1−δd2)2+(δp1−δp2)2+(δh1−δh2)2}1/2 1

where subscripts 1 and 2 indicate components 1 and 2, respectively. A small R A indicates that the two components are more compatible and vice versa. The compatibility between two components can be further expressed using the Relative Energy Density (RED) number, as defined in eq :

RED=RAR0 2

where R A represents the distance between two components in the HSP space, while R 0 denotes the radius of interaction in the same HSP space, defining the region within which solvents are likely to dissolve the polymer. Solvents with R A ≤ R 0 are regarded as “good solvents”, able to dissolve or effectively interact with the polymer. As a guideline, RED ≤1 implies high compatibility between the two components, while RED ≥1 indicates low compatibility. The HSPs (i.e., δd, δp, and δh) and R 0 values of all substances were obtained from the library of the HSPiP software package (HSPiP, fifth edition, version 5.0.02).

2.5. Characterization of the Zein–EC Composite Films

2.5.1. Thickness and Visual Appearance

Film thickness was determined with a digital micrometer (Testing Machines Inc., Islandia, NY, USA) to the nearest 1 μm. Six measurements were taken for each film, obtaining an average thickness of 28 ± 1 μm for all films. Photographs of the composite film samples were taken with a digital camera (Canon SX 400is, Canon Solutions America, Melville, IL, USA) to examine their visual appearance. Film color was measured by a spectrophotometer (CR-400 Meter, Konica Minolta, Ramsey, NJ, USA) at room temperature. The L* value (lightness) was measured for different films. The instrument was calibrated using a white reflector standard plate with reference values (L = 95.13, a = 0.00, and b = 1.47).

2.5.2. Scanning Electron Microscopy (SEM)

Images of the cross sections of all composite films were acquired by using a scanning electron microscope (Quanta FEG 250, FEI Company, Hillsboro, OR, USA). Composite film samples were cryo-fractured in liquid nitrogen, coated with a thin conductive layer of gold–palladium using a sputter coater (Desk V TSC, Denton Vacuum, Moorestown, NJ, USA), and then mounted on a metal stub using double-sided carbon tape. The samples were analyzed by using an accelerating voltage of 10 kV.

2.6. Data Analysis

Particle size and height distributions were analyzed using the SAS University Edition software package (SAS Institute Inc., Cary, NC, USA), with the PROC GLIMMIX procedure and ANOVA, to determine significant differences between treatments at the 95% confidence level. Mean values were compared using Tukey’s honest significant difference test.

3. Results and Discussion

3.1. Characterization of the Zein Particle Suspension

3.1.1. Visual Appearance, pH, and ζ-Potential

The zein solutions prepared in aqueous ethanol or isopropanol (90%, w/w) were yellow and translucent. After antisolvent precipitation in water at pH 10, the resulting suspensions were milky and opaque in appearance. Zein has an isoelectric point of 6.2 and tends to precipitate or aggregate in pH-neutral media. In our preliminary studies, ZPS produced from aqueous ethanol or isopropanol solutions at 90% w/w, with initial pH values of 6.2 and 5.9, respectively, were adjusted to various pH levels using HCl (pH 2 and 4), NaOH (pH 8), and Na2CO3 (pH 10). The resulting suspensions adjusted to pH 2, 4, and 8 were unstable and prone to aggregation during the antisolvent process. However, the adjustment to pH 10 substantially increased the stability of the dispersion and favored the formation of submicrometer zein particles. ζ-potential measurements of ZPS solutions prepared in 90% (w/w) aqueous ethanol or aqueous isopropanol were −42.5 and −35.4 mV, respectively, confirming the formation of stable colloidal systems. These findings are consistent with the strong hydrophobic attraction and weak electrostatic repulsion near the isoelectric point of zein (pH = 6.2), which lead to particle aggregation.

3.1.2. Particle Size Distribution

During the antisolvent precipitation process, rapid diffusion of deionized water into zein solution (ratio 3:1) induced zein supersaturation, leading to particle formation. Zein particles prepared from aqueous ethanol had a smaller average particle size (Dv50 of 0.56 ± 0.40 μm) compared with those prepared from aqueous isopropanol (Dv50 of 3.74 ± 0.03 μm) (Figure ). These results suggested that the higher dipolar (δp = 8.8 MPa1/2) and hydrogen bonding (δh = 19.4 MPa1/2) parameters of ethanol, compared to those of isopropanol (δp = 6.1 MPa1/2, δh = 16.4 MPa1/2), , might have facilitated stronger interactions with polar amino acid residues in zein (e.g., serine, threonine, asparagine, and glutamine). These interactions likely promoted a more effective dispersion of zein in ethanol, thereby favoring the formation of smaller particles. Similar effects of solvent polarity on zein particle size were reported by Chen et al. in a study, where 90% (v/v) aqueous ethanol solution produced smaller zein particles (14.85 nm) compared to 90% (v/v) aqueous isopropanol solution (≤100 nm). Kim and Xu also reported that the hydrodynamic radius of zein decreased from ∼100 nm to <10 nm when prepared in 90% (v/v) aqueous ethanol compared to 70% (v/v) and 80% (v/v) solutions.

1.

1

Open in a new tab

Particle size distribution of zein particle suspension (ZPS) prepared in (1) aqueous ethanol (90%, w/w) or (2) aqueous isopropanol (90%, w/w) after antisolvent precipitation method.

3.1.3. Confocal Laser Scanning Microscope

To confirm the presence of submicrometer zein particles in the ZPS samples, hydrophilic Rhodamine B dye, with an emission wavelength of 610 nm, was dispersed within the ZPS for preferential staining of the aqueous phase. A red fluorescence of dispersed submicrometer zein particles against a dark background was observed after excitation with CLSM. As shown in Figure , red fluorescing spherical geometries with average diameters of 0.68 ± 0.20 μm and 1.72 ± 0.20 μm were observed for ZPS prepared in aqueous ethanol or isopropanol, respectively. This observation confirmed the presence of submicrometer particles in ZPS, which is in accordance with the findings described in Section .

2.

2

Open in a new tab

Confocal laser scanning microscopy images of the zein particles stained by rhodamine B. The dye was dispersed within the zein particle suspension (ZPS) previously prepared using zein (4% w/w) in (1) aqueous ethanol (90%, w/w) or (2) aqueous isopropanol (90%, w/w).

3.1.4. Atomic Force Microscopy Analysis

The zein particle height distribution and average height are summarized in Figure . As shown, zein particles prepared in aqueous ethanol showed a narrower and lower particle height distribution (from 0.00 to 0.42 μm, average height = 0.08 ± 0.01 μm), as compared with a broader particle height distribution of those prepared in aqueous isopropanol (from 0.00 to 1.27 μm, average height = 0.26 ± 1.0 μm). These results can be correlated with the zein particle size distribution presented in Section , which shows that the aqueous ethanol solvent favored the formation of smaller zein particles compared to the aqueous isopropanol solvent. The number of zein aggregates decreased significantly as the ethanol–water solvent ratio increased from 70 to 90% (w/w). Due to its amphiphilicity, zein likely formed micelle-like structures at lower ethanol concentrations but not at elevated ethanol concentrations.

3.

3

Open in a new tab

Surface topography of zein particles prepared in the ZPS sample with (A) aqueous ethanol (90%, w/w) or (B) aqueous isopropanol (90%, w/w). The corresponding particle height distribution plot shows peak count (ρ) as a function of peak height (z) and average height for each preparation.

3.1.5. Hansen Solubility Parameters (HSPs)

The affinity of zein for aqueous ethanol or isopropanol (90%, w/w) solutions is summarized in Table . As shown, zein exhibited comparable solubility (R A = 13.5) in both aqueous solutions, with marginal differences in RED values around 1 (1.13 and 1.14 for aqueous ethanol and aqueous isopropanol, respectively), indicating limited compatibility between zein and either solvent. These results suggest that the R A and RED values alone do not fully account for the observed differences in particle formation. Other important factors include molecular interaction (e.g., protein–protein interactions), solvent dynamics (e.g., conformational changes in protein structure in response to the solvent environment and ratios used), and/or experimental conditions (e.g., ionic interactions potentially arising from increased pH from 6.2 to pH 10). Similar RED values have been reported by other researchers for zein solutions prepared in aqueous ethanol (70% w/w, RED = 1.017) and aqueous isopropanol (70% w/w, RED = 1.043), indicating that zein had marginally higher compatibility with aqueous ethanol solvent compared to aqueous isopropanol.

1. HSPsR A and RED Values for Zein Nanoparticles in Different Aqueous Solvents and for Zein–EC Composite Films Using Different Solvents and Surfactants.
  δd (MPa1/2) δp (MPa1/2) δh (MPa1/2) R A (MPa1/2) RED
Zein (target) 22.4 9.8 19.4    
Ethanol:water (90:10, w/w)       13.5 1.13
Isopropanol:water (90:10, w/w)       13.5 1.14
Tween 80 (T) 16.8 6.5 9.5 15.21 1.29
Span 85 (S) 16.1 3.8 3.7 21.01 1.77
Ethyl cellulose (EC, target) 22.7 0.5 16.5    
Zein 22.4 9.8 19.4 9.76 0.49
Pure ethanol (Eth) 15.8 8.8 19.4 16.36 0.82
Pure isopropanol (Iso) 15.8 6.1 16.4 14.89 0.75
Tween 80 (T) 16.8 6.5 9.5 14.97 0.75
Span 85 (S) 16.1 3.8 3.7 18.68 0.94
Zein-EC, Eth-S, or Eth-T       15.90 0.84
Zein-EC, Iso-S, or Iso-T       14.80 0.78
Open in a new tab
a

R A = Distance of polymer–solvent or polymer–surfactant in Hansen space.

b

RED = Relative energy difference.

EC polymer had a high affinity toward zein polymer (RED = 0.49). Moreover, EC showed higher compatibility with isopropanol as a solvent (R A = 14.89; RED = 0.75) compared to ethanol (R A = 16.36; RED = 0.82). This trend was also observed in zein–EC composite film formulations, regardless of the surfactant used, where it exhibited higher affinity for isopropanol (R A = 14.80; RED = 0.78) compared to ethanol (R A = 15.90; RED = 0.84).

In terms of the surfactant effect, Tween 80 showed higher compatibility with pure EC (RED = 0.75) and pure zein (RED = 1.29) as compared with Span 85 (RED = 0.94 and 1.77 for EC and zein, respectively). These results suggest that Tween 80, with a higher molecular weight (Tween 80 = 1310 g/mol; Span 85 = 957.52 g/mol) and a higher hydrophilic–lipophilic balance index (Tween 80 = 15.0; Span 85 = 1.2), might have lowered the interfacial tension in the composite film suspensions, thereby promoting the formation of more homogeneous films compared to composite films with Span 85.

3.2. Characterization of the Zein–EC Composite Films

3.2.1. Visual Appearance

The appearance of zein–EC composite films dried at 50 °C and 65% RH is shown in Figure . The film samples were coherent and could be peeled off easily from the glass cast plates. All films were optically transparent, except for Eth-S films, which were relatively opaque.

4.

4

Open in a new tab

Visual appearance of film samples against a black background, and L* value of zein–EC composite films prepared by casting method at 50 °C and 65% RH, using different solvents and surfactants: (A) Eth; (B) Eth-T; (C) Eth-S; (D) Iso; (E) Iso-T; and (F) Iso-S (Eth = pure ethanol; Iso = pure isopropanol; T = Tween 80; S = Span 85).

Without surfactants, composite films prepared from ethanol were more homogeneous in appearance than those prepared from isopropanol (Figure A,D, respectively), possibly due to the slower evaporation rate of isopropanol, which promoted the formation of zein aggregates. The incorporation of Tween 80 into films prepared from both alcohols greatly facilitated the zein particle dispersion and film-formation processes (Figure B,E, respectively), resulting in a more homogeneous zein particle dispersion than films prepared with Span 85. Moreover, Figure C shows that Span 85-containing films prepared from ethanol solvent had a heterogeneous morphology with dispersed particles and inclusions of hydrophobic Span 85 on the film surface, resulting in reduced visual transparency as compared to Span 85-containing films prepared in isopropanol (Figure F).

3.2.2. Scanning Electron Microscopy (SEM)

SEM micrographs of cross sections of the composite films are shown in Figure . Films prepared from ethanol (Figure A) displayed small zein particles (≤1 μm) throughout the entire cross section. At a drying temperature of 50 °C, the rapid evaporation of ethanol might have resulted in the fusion of zein particles during drying, thereby decreasing light scattering and forming a transparent and homogeneous composite film. By contrast, film prepared from isopropanol (Figure D) showed a heterogeneous cross section, possibly due to the slower evaporation rate of isopropanol, which promoted the formation of zein aggregates at 50 °C. On the other hand, ethanol composite films containing Tween 80 (Figure B) exhibited a compact, homogeneous structure with submicrometer zein particles. In contrast, those containing Span 85 (Figure C) displayed more dispersed particles and surface-localized inclusions of the hydrophobic surfactant, as previously noted in Section . Moreover, the presence of zein aggregates in films prepared from isopropanol containing Tween 80 (Figure E) or Span 85 (Figure F) was evident, suggesting a heterogeneous structure throughout both composite films.

5.

5

Open in a new tab

SEM micrographs of zein–EC composite films prepared with different solvents and surfactants: (A) Eth; (B) Eth-T; (C) Eth-S; (D) Iso; (E) Iso-T; and (F) Iso-S (Eth = pure ethanol; Iso = pure isopropanol; T = Tween 80; S = Span 85).

3.2.3. Atomic Force Microscopy (AFM, Tapping Mode)

Phase and height images of the zein–EC composite film surface topographies are shown in Figures and , respectively. Films prepared from ethanol without surfactant (Figure A) exhibited lower surface roughness (R a = 4.23 nm) compared to those prepared from isopropanol (R a = 19.5 nm, Figure D), supporting the observations discussed in Sections and . Moreover, the phase image of the Span 85-containing film prepared in ethanol (Figure C) showed the highest R a value (Eth-S = 26.3 nm) among all film formulations, revealing their surface heterogeneity, characterized by more pronounced bright areas compared to the other composite films. The bright areas can be attributed to zein–surfactant aggregates that exhibit viscoelastic properties of a higher phase shift due to increased energy dissipation during the AFM probe–sample interaction. On the other hand, the dark areas correspond to the EC-rich domains with stiffer and more elastic properties, characterized by lower energy dissipation during probe–surface interaction.

6.

6

Open in a new tab

Phase images of zein–EC composite films prepared with different solvents and surfactants: (A) Eth; (B) Eth-T; (C) Eth-S; (D) Iso; (E) Iso-T; and (F) Iso-S (Eth = pure ethanol; Iso = pure isopropanol; T = Tween 80; S = Span 85). Mean roughness value = R a: mean roughness.

7.

7

Open in a new tab

Surface topography of zein–EC composite films prepared with different solvents and surfactants: (A) Eth; (B) Eth-T; (C) Eth-S; (D) Iso; (E) Iso-T; and (F) Iso-S (Eth = pure ethanol; Iso = pure isopropanol; T = Tween 80; S = Span 85). The corresponding height distribution plot shows peak count (ρ) as a function of peak height (z) and the average height for each preparation.

The height profile analysis showed that the films prepared from isopropanol Tween 80-containing films (Figure E) had a narrow height distribution with an average height of 11.52 ± 0.03 nm, indicating a relatively smoother surface compared to other composite film formulations. Conversely, films prepared from ethanol Span 85-containing solutions had a considerably higher average height value (152.10 ± 0.67 nm, Figure C) compared to the other composite films (Figure B,D,F), indicating a more heterogeneous film structure. This microstructural heterogeneity is consistent with the limited dispersion of zein particles within the EC matrix and the increased surface roughness observed in Figures C and C, respectively. The heterogeneous zein aggregates scattered light, leading to increased film opacity, as noted in Section .

4. Conclusions

Using the antisolvent precipitation method, zein particles prepared with aqueous ethanol (90%, w/w) were smaller than those prepared with aqueous isopropanol (90%, w/w), as revealed by laser diffractometry, confocal laser scanning microscopy, and AFM analyses. From Hansen Solubility Parameter (HSP) analysis, the higher dipolar and hydrogen bonding interactions of ethanol with zein promoted the formation of smaller particles than those prepared in isopropanol. The microstructures of zein:EC composite films prepared from these zein particles were influenced by the type of solvent and surfactant used. The composite films were visually transparent, except for Eth-S, where dispersed particles and inclusions of hydrophobic Span 85 were observed on the surface, indicating lower compatibility between zein–EC polymers, ethanol, and Span 85. SEM analysis revealed homogeneous structures in pure Eth or Eth-T composite films. AFM-tapping mode confirmed that films prepared with either ethanol or isopropanol containing Tween 80 (Eth-T and Iso-T) presented a narrower and lower height distribution compared to the other films. These results aligned with HSP predictions. Overall, this study provided an insight into how solvent and surfactant affect zein particle formation and the morphology of EC composite films containing the zein particles. In order to use the zein-EC composite films for structural and barrier applications, further research is needed to characterize their mechanical, thermal, and barrier properties. Additionally, the zein particles embedded in the EC matrices may serve as carrier for bioactive compounds. For this purpose, future work should evaluate the encapsulation efficiency and controlled release profiles under conditions relevant to the intended applications.

Acknowledgments

This research was made possible through support and funding from the Natural Sciences and Engineering Research Council of Canada (NSERCALLRP 560774-20). The authors acknowledge the support of the University of Guelph Molecular and Cellular Imaging Facility for the use of the confocal laser microscope. Y.Z.’s stipend and tuition are cosupported by the China Scholarship Council and the University of Guelph.

The authors declare no competing financial interest.

References

  1. Zhang Z., Li X., Sang S., McClements D. J., Chen L., Long J., Jiao A., Jin Z., Qiu C.. Preparation, properties and interaction of curcumin loaded zein/HP-β-CD nanoparticles based on electrostatic interactions by antisolvent co-precipitation. Food Chem. 2023;403:134344. doi: 10.1016/j.foodchem.2022.134344. [DOI] [PubMed] [Google Scholar]
  2. Inchaurraga L., Martínez-López A. L., Abdulkarim M., Gumbleton M., Quincoces G., Peñuelas I., Martin-Arbella N., Irache J. M.. Modulation of the fate of zein nanoparticles by their coating with a Gantrez® AN-thiamine polymer conjugate. Int. J. Pharm. 2019;1:100006. doi: 10.1016/j.ijpx.2019.100006. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Pascoli M., De Lima R., Fraceto L. F.. Zein nanoparticles and strategies to improve colloidal stability: A mini-review. Front. Chem. 2018;6:6. doi: 10.3389/fchem.2018.00006. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Dai L., Sun C., Li R., Mao L., Liu F., Gao Y.. Structural characterization, formation mechanism and stability of curcumin in zein-lecithin composite nanoparticles fabricated by antisolvent co-precipitation. Food Chem. 2017;237:1163–1171. doi: 10.1016/j.foodchem.2017.05.134. [DOI] [PubMed] [Google Scholar]
  5. Spasojević L., Katona J., Bučko S., Savić S. M., Petrović L., Budinčić J. M., Tasić N., Aidarova S., Sharipova A.. Edible water barrier films prepared from aqueous dispersions of zein nanoparticles. LWT. 2019;109:350–358. doi: 10.1016/j.lwt.2019.04.038. [DOI] [Google Scholar]
  6. Chen Y., Ye R., Liu J.. Understanding of dispersion and aggregation of suspensions of zein nanoparticles in aqueous alcohol solutions after thermal treatment. Ind. Crops Prod. 2013;50:764–770. doi: 10.1016/j.indcrop.2013.08.023. [DOI] [Google Scholar]
  7. Yan X., Li M., Xu X., Liu X., Liu F.. Zein-based nano-delivery systems for encapsulation and protection of hydrophobic bioactives: A review. Front. Nutr. 2022;9:999373. doi: 10.3389/fnut.2022.999373. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Souza E. M. C., Ferreira M. R. A., Soares L. A. L.. Pickering emulsions stabilized by zein particles and their complexes and possibilities of use in the food industry: A review. Food Hydrocolloids. 2022;131:107781. doi: 10.1016/j.foodhyd.2022.107781. [DOI] [Google Scholar]
  9. Liu C., Xu B., McClements D. J., Xu X., Cui S., Gao L., Zhou L., Xiong L., Sun Q., Dai L.. Properties of curcumin-loaded zein-tea saponin nanoparticles prepared by antisolvent co-precipitation and precipitation. Food Chem. 2022;391:133224. doi: 10.1016/j.foodchem.2022.133224. [DOI] [PubMed] [Google Scholar]
  10. Sun C., Dai L., He X., Liu F., Yuan F., Gao Y.. Effect of heat treatment on physical, structural, thermal and morphological characteristics of zein in ethanol-water solution. Food Hydrocolloids. 2016;58:11–19. doi: 10.1016/j.foodhyd.2016.02.014. [DOI] [Google Scholar]
  11. Tortorella S., Maturi M., Buratti V. V., Vozzolo G., Locatelli E., Sambri L., Franchini M. C.. Zein as a versatile biopolymer: Different shapes for different biomedical applications. RSC Adv. 2021;11:39004–39026. doi: 10.1039/D1RA07424E. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Patel A., Hu Y., Tiwari J. K., Velikov K. P.. Synthesis and characterisation of zein–curcumin colloidal particles. Soft Matter. 2010;6:6192–6199. doi: 10.1039/c0sm00800a. [DOI] [Google Scholar]
  13. Patnode K., Rasulev B., Voronov A.. Synergistic behavior of plant proteins and biobased latexes in Bioplastic food packaging materials: Experimental and machine learning study. Appl. Mater. Interfaces. 2022;14:8384–8393. doi: 10.1021/acsami.1c21650. [DOI] [PubMed] [Google Scholar]
  14. Jaski A. C., Schmitz F., Horta R. P., Cadorin L., Da Silva B. J. G., Andreaus J., Paes M. C. D., Riegel-Vidotti I. C., Zimmermann L. M.. Zein - a plant-based material of growing importance: New perspectives for innovative uses. Ind. Crops Prod. 2022;186:115250. doi: 10.1016/j.indcrop.2022.115250. [DOI] [Google Scholar]
  15. Horvat G., Žvab K., Knez Ž., Novak Z.. Simple, one-pot method for preparing transparent ethyl cellulose films with good mechanical properties. Polymers. 2022;14:2399. doi: 10.3390/polym14122399. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Parashar B., Thakur A., Thakur V., Prashar D.. Ethyl cellulose and zein: A mini review. Int. J. Inst. Pharm. Life Sci. 2012;2(3):274–279. [Google Scholar]
  17. Guo X., Ren C., Zhang Y., Cui H., Shi C.. Stability of zein-based films and their mechanism of change during storage at different temperatures and relative humidity. J. Food Process. Preserv. 2020;44(9):e14671. doi: 10.1111/jfpp.14671. [DOI] [Google Scholar]
  18. Liang J., Xia Q., Wang S., Li J., Huang Q., Ludescher R. D.. Influence of glycerol on the molecular mobility, oxygen permeability and microstructure of amorphous zein films. Food Hydrocolloids. 2015;44:94–100. doi: 10.1016/j.foodhyd.2014.09.002. [DOI] [Google Scholar]
  19. Chen Y., Ye R., Liu J.. Effects of different concentrations of ethanol and isopropanol on physicochemical properties of zein-based films. Ind. Crops Prod. 2014;53:140–147. doi: 10.1016/j.indcrop.2013.12.034. [DOI] [Google Scholar]
  20. Goudar N., Vanjeri V. N., Hiremani V. D., Gasti T., D’souza O. J., Chougale R. B., Masti S. P.. Evaluation of spectroscopic, microscopic and hydrophobic properties of cetrimide and polyethylene glycol-400 plasticized ethyl cellulose films. Chem. Data Collect. 2021;36:100796. doi: 10.1016/j.cdc.2021.100796. [DOI] [Google Scholar]
  21. Su X., Yang Z., Tan K. B., Chen J., Huang J., Li Q.. Preparation and characterization of ethyl cellulose film modified with capsaicin. Carbohydr. Polym. 2020;241:116259. doi: 10.1016/j.carbpol.2020.116259. [DOI] [PubMed] [Google Scholar]
  22. Mahnaj T., Ahmed S. U., Plakogiannis F. M.. Characterization of ethyl cellulose polymer. Pharm. Dev. Technol. 2013;18(5):982–989. doi: 10.3109/10837450.2011.604781. [DOI] [PubMed] [Google Scholar]
  23. Hansen, C. M. Hansen Solubility Parameters: A user’s handbook; Second ed.; CRC Press, 2007, p. 544. [Google Scholar]
  24. Sato T., Hamada Y., Sumikawa M., Araki S., Yamamoto H.. Solubility of oxygen in organic solvents and calculation of the Hansen Solubility Parameters of Oxygen. Ind. Eng. Chem. Res. 2014;53:19331–19337. doi: 10.1021/ie502386t. [DOI] [Google Scholar]
  25. Rutkevicius M., Allred S., Velev O. D., Velikov K. P.. Stabilization of oil continuous emulsions with colloidal particles from water-insoluble plant proteins. Food Hydrocolloids. 2018;82:89–95. doi: 10.1016/j.foodhyd.2018.04.004. [DOI] [Google Scholar]
  26. Kim S., Xu J.. Aggregate formation of zein and its structural inversion in aqueous ethanol. J. Cereal Sci. 2008;47:1–5. doi: 10.1016/j.jcs.2007.08.004. [DOI] [Google Scholar]
  27. Giteru S. G., Ali M. A., Oey I.. Review: Recent progress in understanding fundamental interactions and applications of zein. Food Hydrocolloids. 2021;120:106948. doi: 10.1016/j.foodhyd.2021.106948. [DOI] [Google Scholar]
  28. Moomand K., Lim L.-T.. Effects of solvent and n-3 rich fish oil on physicochemical properties of electrospun zein fibres. Food Hydrocolloids. 2015;46:191–200. doi: 10.1016/j.foodhyd.2014.12.014. [DOI] [Google Scholar]

Articles from Langmuir are provided here courtesy of American Chemical Society

RESOURCES