Chapter 3

Biopolymer Composite Materials

with Antimicrobial Effects Applied
to the Food Industry

Kelvia Álvarez, Vera A. Alvarez and Tomy J. Gutiérrez

Abstract Over the past decade, there has been a marked increase in consumer
demand for healthy foods that are convenient, safe, with a longer useful life, and
packaged using eco-materials. This has prompted both scientists and the food
industry to investigate new strategies for food processing, handling, and packaging.
In particular, films and coatings made from biodegradable and edible composite
materials with antimicrobial effects, either due to intrinsic characteristics or through
the incorporation of traditional or natural antimicrobial composites have been
developed. These materials have proved to be a novel alternative to extend the
shelf-life of foods, while maintaining their physical, chemical, and sensory prop-
erties and, most importantly, ensuring food safety. They also serve as a barrier
against moisture loss from foods and the entry of oxygen. In addition, they can
convey different bioactive compounds, some of which have an antimicrobial effect
on important pathogenic microorganisms, thus ensuring food safety. All of this with
an advantageous cost–benefit ratio. In this chapter, we review various
biopolymer-based antimicrobial composites incorporated in films and coatings and
their effects on different food matrices. Finally, factors that should be considered
when developing composite materials with antimicrobial effects, as well as toxi-
cological aspects and the regulatory status of these materials are discussed.

K. Álvarez
T.J. Gutiérrez

Faculty of Sciences, Institute of Food Science and Technology (ICTA), Central University of
Venezuela (UCV), PO Box 47097, Caracas 1041-A, Venezuela
V.A. Alvarez (&)
T.J. Gutiérrez (&)
Thermoplastic Composite Materials (CoMP) Group, Faculty of Engineering, Institute of
Research in Materials Science and Technology (INTEMA), National University of Mar del
Plata (UNMdP), National Council of Scientific and Technical Research (CONICET), Colón
10850, Mar del Plata 7600, Buenos Aires, Argentina
e-mail: alvarezvera@fi.mdp.edu.ar; [email protected]
T.J. Gutiérrez
e-mail: tomy.gutierrez@fi.mdp.edu.ar; [email protected]

© Springer International Publishing AG 2018 57

V.K. Thakur and M.K. Thakur (eds.), Functional Biopolymers,
Springer Series on Polymer and Composite Materials,
https://doi.org/10.1007/978-3-319-66417-0_3
58 K. Álvarez et al.

Keywords Antimicrobials Composite materials Edible coatings




Edible films Natural fillers Packaging Thermoplastic materials

1 Introduction

Lifestyle and diet changes driven by the patterns of modern society have led to a
greater acquisition by consumers of ready-to-eat products. These foods should
provide adequate nutritional values for the proper functioning of the body and be
microbiologically safe. The packaging should also be practical and, when dis-
carded, friendly to the environment (Cagri et al. 2004). Such points have proved to
be a challenge for the food and polymer industries, both technologically (devel-
opment of new strategies for processing, handling, and packaging) and logistically
(management, marketing, and appropriate delivery systems). This is largely because
ready-to-eat foods require greater manipulation during processing, and are therefore
more susceptible to attack by deteriorative and pathogenic microorganisms. This
could lead to potential public health problems if they are not adequately processed,
handled, and distributed (Beuchat 1996; Díaz-Cinco et al. 2005). Proper treatment,
packaging, and storage are thus required to prevent contamination of these products
and increase their useful life (Cagri et al. 2004).
The food industry has traditionally implemented several strategies to ensure food
safety, namely cooling, dehydration, acidification, packaging in a modified atmo-
sphere, fermentation, and/or the use of antimicrobials. This with the aim to control
the growth of microorganisms in foods such as sausages, sliced cheeses,
ready-to-cook meat products, and fresh-cut fruit and vegetables (Davidson et al.
2013). New preservation techniques are, however, being studied and successfully
applied to these products. One of these is the use of edible coatings and films based
on biodegradable polymers with added antimicrobial substances (Wang et al. 2016).
Of course, it must be said that a wide variety of packaging materials of inedible
origins, such as polypropylene and polyethylene films, are already available to the
food industry. However, although these materials do protect food from environ-
mental pollution, they are non-biodegradable and thus not environmentally friendly
(Michalska-Pożoga et al. 2016; Thakur et al. 2014c). Because of this, packaging
materials made with edible films and coatings derived from proteins, lipids, and
polysaccharides are being studied and applied to food products. These biodegrad-
able and environmentally friendly materials have been shown to act as barriers
against water vapor, gases, and volatile compounds. They also serve as carriers of
antimicrobial substances, antioxidants, flavorings, and vitamins, which can increase
the quality, safety, functionality, and useful life of foodstuffs (Gennadios et al.
1997; Cagri et al. 2004; Lin and Zhao 2007; Rojas-Graü et al. 2009).
Two fundamental procedures have been used to control the behavior of
microorganisms in food through the application of films: 1—the incorporation of
antimicrobial agents into the polymeric matrix and 2—the generation of bioactive
polymeric matrices on the surface of foods, which hinder the growth of
3 Biopolymer Composite Materials with Antimicrobial Effects … 59

microorganisms (Silvera-Almitrán et al. 2012). As regards the first of these,

numerous traditional and natural antimicrobials have been incorporated into films,
including benzoates, propionates, sorbates, parabens, acidifying agents (acetic acid,
lactic acid, etc.), curing salts (sodium chloride and sodium nitrite), natural agents
(essential oils, lysozyme, liquid smoke), and bacteriocins (Cagri et al. 2004;
Raybaudi-Massilia and Mosqueda-Melgar 2012). For example, amino groups on
the C2 of films prepared from chitosan act on the biological capabilities of
microorganisms. This affects their metabolic processes, thus restricting their growth
(Silvera-Almitrán et al. 2012). In this chapter, we focus on edible films and coatings
derived from proteins, lipids, and polysaccharides, either with antimicrobial sub-
stances added or with their own intrinsic antimicrobial activity, applied to
ready-to-eat foods.

2 Antimicrobial Agents Incorporated into Edible Films

and Coatings

Food antimicrobials are chemical compounds or substances that may retard the
growth of microbials or cause their death when they are incorporated into a food
matrix (Davidson and Zivanovic 2003). The main targets for these antimicrobials
are toxin-producing pathogenic microorganisms, microorganisms that cause
infections, and deteriorative microorganisms whose metabolic end products pro-
duce undesirable odors and flavors, problems with texture and the discoloration of
the product (Davidson et al. 2013).
The grouping of antimicrobials into specific types is not easy, since some of
those are considered as traditional and/or synthetics can also be found naturally in
foods. Nevertheless, they are currently classified by some authors as traditional and
naturals (naturally occurring compounds), while taking into account certain con-
siderations (Raybaudi-Massilia et al. 2009; Davidson et al. 2013) as follows.
Antimicrobials are called traditional when they: (1) have been used for many
years, (2) have been tested in many countries as food antimicrobials, and (3) have
been produced by chemical synthesis. This classification does not imply that a
traditional or synthetic preservative is any less effective from a microbiological
point of view than one of natural origin, or vice versa.
Nonetheless, natural antimicrobial agents derived from plants, animals, micro-
bial, and inorganic nanoparticles are preferred nowadays by consumers, due to their
active promotion as “non-harmful” additives to the organism, and thus “healthier”
than nonnatural or synthetic chemical substances. Table 1 shows some traditional
and natural antimicrobial agents, which have been added to food packaging
materials in order to obtain functional biopolymers with antimicrobial activity.
In addition, the incorporation of antimicrobial agents such as organic acids,
bacteriocins, enzymes, plant extracts, and polysaccharides into edible films and
coatings, has proven to be a novel alternative to retard or inhibit the growth of
Table 1 Some traditional and natural antimicrobial agents for the development of functional biopolymers
60

Antimicrobial agent Biopolymeric material References

Film Coating
Traditional Malic acid Alginate, milk Gadang et al. (2008), Raybaudi-Massilia et al. (2008a, b)
whey protein
Acetic acid Chitosan Bégin and Van Calsteren (1999)
Formic acid Chitosan Bégin and Van Calsteren (1999)
Lactic acid Chitosan Bégin and Van Calsteren (1999), Ye et al. (2008)
Lauric acid Soy protein Dawson et al. (2007)
Propionic acid Zein Cellulose Janes et al. (2002), Valencia-Chamorro et al. (2009)
Potassium sorbate Chitosan Cellulose Pranoto et al. (2005a, b), Ye et al. (2008), Valencia-Chamorro
et al. (2009)
Sodium benzoate Chitosan Cellulose Ye et al. (2008), Valencia-Chamorro et al. (2009)
Natural or Chitosan Chitosan, Chitosan Bégin and Van Calsteren (1999), Pen and Jiang (2003),
new methylcellulose Devlieghere et al. (2004), Dong et al. (2004), González-Aguilar
et al. (2005, 2009), Pranoto et al. (2005a, b), Sebti et al. (2005),
Chien et al. (2007), Sangsuwan et al. (2008), Ye et al. (2008),
Dutta et al. (2009), Vargas et al. (2009)
Enzymes Chitosan, alginate, Chitosan Cha et al. (2002), Seol et al. (2009).
(lysozyme and carrageenan
ovotransferin)
Essential oils Chitosan, alginate, soy Alginate, apple Pranoto et al. (2005a, b), Oussalah et al. (2006a, b), Rojas-Graü
protein puree et al. (2007), Raybaudi-Massilia et al. (2008a, b)
Bacteriocins Chitosan, alginate, Milk whey Cha et al. (2002), Janes et al. (2002), Franklin et al. (2004),
(nisin, enterocins carrageenan, soy protein, protein Pranoto et al. (2005a, b), Dawson et al. (2007), Millette et al.
and pediocins) cellulose, zein (2007), Gadang et al. (2008), Marcos et al. (2008), Santiago-Silva
et al. (2009)
Grapeseed and Alginate, carrageenan, Milk whey Cha et al. (2002), Theivendran et al. (2006), Gadang et al. (2008),
grapefruit extracts gelatin protein, soy Hong et al. (2009)
K. Álvarez et al.

protein
(continued)
Table 1 (continued)
Antimicrobial agent Biopolymeric material References
Film Coating
Green tea extract Gelatin Soy protein Theivendran et al. (2006), Hong et al. (2009)
Species powder Caseinate, milk whey Outtara et al. (2002)
protein
Vanillin Alginate, Rojas-Graü et al. (2007), Sangsuwan et al. (2008)
cellulose, apple
puree, chitosan
3 Biopolymer Composite Materials with Antimicrobial Effects …
61
Table 2 Effect of functional biopolymers on foods
62

Matrix type Antimicrobial Food Effect References

added applied
Alginate Nisin Ground beef Reductions of more than 3.0 log CFU/g of the population Cutter and
of Brochothrix thermosphacta Siragusa
(1997)
Alginate Nisin Chicken Reductions of more than 4.0 log CFU/g of the population Natrajan and
of Salmonella enterica ser. Typhimurium Sheldon (2000)
Alginate Nisin Meat fillets Reductions of more than 2.5 log CFU/g of the population Millette et al.
of Staphylococcus aureus (2007)
Alginate Lysozyme/Nisin Smoked Reductions in populations of L. monocytogenes and Datta et al.
salmon Salmonella enterica serovar Anatum in a range between (2008)
2.2 and 2.8 log CFU/g
Alginate Enterotoxin A and Baked Bacteriostatic effect on the population of L. Marcos et al.
B sliced ham monocytogenes (2008)
Alginate Essential oil of Meat fillets Reduction of more than 1.0 log CFU/g of the population Oussalah et al.
oregano, cinnamon of S. enterica serovar Typhimurium applying alginate (2006a, b)
and savory films with essential oils of oregano or cinnamon.
Reduction of more than 2.0 log CFU/g of the population
of E. coli O157:H7 applying alginate films with oregano
essential oil
Alginate Essential oil of Baked ham Reductions of more than 2.0 log CFU/g of populations of Oussalah et al.
oregano, cinnamon, and bologna Salmonella enterica serovar Typhimurium and L. (2007)
and savory sliced monocytogenes in bologna and cooked ham slices
applying alginate films with cinnamon essential oil
Carrageenan Ovotransferin Fresh Reductions of 2.0 log CFU/g for the total population of Seol et al.
chicken microorganisms and 3 log CFU/g for E. coli when (2009)
carrageenan films were applied with ovotransferin and
EDTA
(continued)
K. Álvarez et al.
Table 2 (continued)
Matrix type Antimicrobial Food Effect References
added applied
Chitosan Cinnamaldehyde Cooked The growth of Enterobacteriaceae and Serratia Ouattara et al.
and acetic, ham, liquefaciens was delayed or completely inhibited, while (2000)
propionic, and bologna, the lactic acid bacteria were unaffected
lauric acids and pastrami
Chitosan Lysozyme Mozzarella Reduction of 1.0 log CFU/g of populations of L. Duan et al.
cheese monocytogenes, E. coli, and Pseudomonas fluorescens (2007)
Chitosan Natamycin Cheese Reductions of 1.1 log CFU/g of yeast and mold Fajardo et al.
populations (2010)
Chitosan Potassium sorbate, Smoked Inactivation of the population of L. monocytogenes Jiang et al.
sodium lactate, and salmon (2011)
diacetate
Chitosan Green tea extract Pork Complete inhibition of microbial growth Siripatrawan
sausages and Noipha
(2012)
CMC Potassium sorbate Fresh Inhibition of mold growth Sayanjali et al.
pistachio (2011)
Cellulose Pediocin Baked Films with pediocin (50%) caused a reduction of 2.0 log Santiago-Silva
sliced ham CFU/g of the population of L. innocua while films with et al. (2009)
3 Biopolymer Composite Materials with Antimicrobial Effects …

pediocin to 25 or 50% only caused a reduction of 0.5 log

CFU/g of the population of Salmonella sp.
Cellulose Nisin “Frankfurt”- Reductions of about 2.0 log CFU/g in the population of Nguyen et al.
type L. monocytogenes and 3.3 log CFU/g in the population of (2008)
sausages total aerobics
Methylcellulose/hydroxypropyl Nisin Sausages Reductions of more than 2.0 log CFU/g of the population Franklin et al.
methylcellulose of L. monocytogenes (2004)
(continued)
63
Table 2 (continued)
64

Matrix type Antimicrobial Food Effect References

added applied
Apple-based films Cinnamaldehyde Meat and Antimicrobial activity of the films turned out to be Ravishankar
and carvacrol chicken dependent on the concentration, temperature, and et al. (2009)
antimicrobial, being the greatest reductions in the
populations of Salmonella enterica and E. coli O157:H7
(4.3–6.8 log CFU/g) at a concentration 3% of either
antimicrobial (cinnamaldehyde or carvacrol) at 23 °C.
While at 4 °C the reductions achieved in these
populations were lower and carvacrol was more effective
than the cinnamaldehyde
Apple-based films Cinnamaldehyde Ham Films containing carvacrol were more effective than those Ravishankar
and carvacrol containing cinnamaldehyde to reduce the population of L. et al. (2009)
monocytogenes inoculated. Besides, greater reductions
were found at 23 °C than at 4 °C
Apple-based films Cinnamaldehyde Chicken Films containing cinnamaldehyde were more effective Mild et al.
and carvacrol than those containing carvacrol to reduce the population (2011)
of Campylobacter jejuni. Besides, greater reductions of
that population were observed at 23 °C compared to 4 °C
K. Álvarez et al.
3 Biopolymer Composite Materials with Antimicrobial Effects … 65

Fig. 1 Mechanisms of action of functional biopolymers from the incorporation of antimicrobial

agents

bacteria, yeasts, and molds in a wide range of foods. The resulting improvement in
food quality and safety has meant that the application of these agents is becoming
ever more common (Table 2). Figure 1 summarizes the possible mechanisms of
action of antimicrobial agents used in the development of functional biopolymers.

2.1 Traditional Antimicrobial Agents

Organic acids are the most traditional and commonly used preservation agents in
the food industry. Some of these, for example, citric, malic, lauric, propionic,
tartaric, benzoic, sorbic and lactic acid, have been incorporated into edible films and
coatings, either indirectly or in salt form. Their spectrum of antimicrobial action is
very wide, and results from a direct decrease in the pH of the substrate or growth
medium due to an increase in the proton concentration. This alters the permeability
of the cell membrane due to interactions between the membrane proteins and
phospholipids, thus affecting its functioning (Davidson et al. 2013). Furthermore,
the non-dissociated forms of weak organic acids can penetrate the lipidic bilayer of
the cell membrane of microorganisms more easily. Once inside, the acid molecule is
forcibly dissociated into anions and protons due to the near-neutral pH in the cell
cytoplasm. This leads to cell inactivation by damage to cell signaling, active
transport mechanisms, and genetic material (Stratford and Eklund 2003).
66 K. Álvarez et al.

2.2 Natural Antimicrobial Agents

Depending on their origin, natural antimicrobials may be classified as vegetable,

animal, microbial, or inorganic.

2.2.1 Agents of Vegetable Origin

Vegetable antimicrobials are those compounds naturally present in different plant

parts, such as stems, bark, roots, flowers, and fruits, which have an inhibitory or
bactericidal effect on microorganisms. Among the compounds in this group that
have been applied to biopolymer-based edible films and coatings are essential oils,
spices, and other extracts.

Essential Oils

Essential oils are aromatic oily liquids derived from plant parts (flowers, seeds,
leaves, twigs, grass, wood, fruits, and roots). They consist of a complex mixture of
compounds such as terpenes, alcohols, ketones, phenols, acid, aldehydes, and esters
(Burt 2004; Ayala-Zavala et al. 2005) that can be obtained by fermentation,
extraction, or distillation. Essential oils and their components have been used since
ancient times as flavorings in food. However, some of them, for example, those
extracted from oregano, cinnamon, clove, lemongrass, palmarosa, thyme, savory,
pepper and garlic, and some of their active compounds, such as carvacrol, cin-
namaldehydes, eugenol, citral, geraniol, and thymol, have recently been incorpo-
rated as antimicrobial agents into biopolymer-based edible films and coatings
(Oussalah et al. 2004, 2006a; Pranoto et al. 2005a; Raybaudi-Massiilia et al. 2008a,
b; Rojas-Graü et al. 2009). There is no one mechanism of action of essential oils on
microorganisms, as they consist of multiple chemical compounds that act on dif-
ferent targets on and within microbial cells (Burt 2004). Nychas et al. (2003), Burt
(2004) and Oussalah et al. (2006b) indicate that essential oils can cause degradation
of the cell wall, damage to the cytoplasmic membrane and membrane proteins, the
output of cell content, coagulation of the cytoplasm, and a decrease in the proton
motive force. Nychas et al. (2003) suggest that the mode of action of essential oils is
dependent on their concentration, such that low concentrations inhibit enzymes
associated with energy production, while high concentrations can cause the pre-
cipitation of proteins.

Vanillin

Vanillin (4-hydroxy-3-methoxybenzaldehyde) is a phenolic aldehyde present in

vanilla beans. It has been used in edible coatings both as a flavoring and an
3 Biopolymer Composite Materials with Antimicrobial Effects … 67

antimicrobial agent (Rojas-Graü et al. 2007). Vanillin is effective against

Gram-positive and Gram-negative microorganisms, yeasts, and molds (Walton
et al. 2003), but is most active against this last and non-lactic Gram-positive bac-
teria (Davidson et al. 2013). According to scientific studies, the inhibitory and
bactericidal activity of vanillin lies mainly in its ability to negatively affect the
integrity of the microorganism cytoplasmic membrane. This results in the loss of
ions, thus unbalancing pH homeostasis and inhibiting respiratory activity
(Fitzgerald et al. 2004).

Green Tea, Grapefruit, and Grape Seed Extracts

Green tea (GTE), grape seed (GS), and grapefruit seed (GFS) extracts have also
been added to edible coatings in order to obtain composite biopolymers, due to the
fact that they contain high concentrations of phenolic compounds. Polyphenols,
particularly flavonoids such as catechins, are the most abundant compounds in GTE
(Almajano et al. 2008) and many of its antimicrobial and antioxidant properties
have been attributed to catechin fractions such as epicatechin (EC), epigallocatechin
(EGC), epigallocatechin gallate (EGCg), epicatechin gallate (ECg), catechin gallate
(Cg), and gallocatechin gallate (GCg) (Frei and Higdon 2003). Taylor et al. (2005)
note that studies conducted in the past 20 years have shown that polyphenolic
catechins in green tea, particularly EGCg and ECg, can inhibit microbial growth in
a wide range of Gram-positive and Gram-negative microorganisms. Shimamura
et al. (2007) showed that the antimicrobial activity of these two catechins can be
attributed mainly to the galloyl fraction (g), which is absent in EC and EGC. These
authors also reported that EGCg can be directly linked to the precipitation of the
peptidoglycan layer in microorganisms, thus interfering with their biosynthesis.
Gram-positive bacteria are more sensitive to this action than Gram-negative bac-
teria, due to the presence of an outer membrane in the latter that serves as a barrier
to antibacterials. The physiological function of the outer membrane of
Gram-negative bacteria, and the low affinity between the polysaccharides contained
within it and EGCg limit the linking of EGCg to the peptidoglycan layer, thus
protecting them from antimicrobial activity (Shimamura et al. 2007).
GS and GFS are also rich sources of monomeric phenolic compounds such as
the catechins EC and ECg, procyanidin dimer, trimer, and tetramer, which can act
as antiviral and anti-mutagenic agents (Saito et al. 1998). GS and GFS are effective
mainly against Gram-positive bacteria, with gallic acid the main active component
(Jayaprakasha et al. 2003). The mechanisms of antimicrobial action proposed for
GS and GFS, are that the fractions containing phenolic components and pro-
cyanidins may act on cell membrane proteins, producing the loss of K+, glutamic
acid, and intracellular RNA, among others, as well as altering the fatty acid com-
position of the membrane (Rozes and Peres 1998; Heggers et al. 2002).
68 K. Álvarez et al.

2.2.2 Agents of Animal Origin

Among the antimicrobials incorporated into edible films and coatings originating
from animals are enzymes such as lysozyme and ovotransferin, as well as the
polysaccharide, chitosan.

Enzymes

Lysozyme (peptidoglycan N-acetylmuramoyl hydrolase EC3.2.1.17) is an enzyme

found in mammalian milk, tears and other secretions, bird eggs, insects, and fish.
This enzyme catalyzes the hydrolysis of the glycosidic a-1, 4 bond between N-
acetylmuramic acid and N-acetylglucosamine found in the peptidoglycan layer of
bacterial cell walls (Davidson et al. 2013). Lysozyme is more effective against
Gram-positive bacteria than Gram-negative bacteria, probably because in the for-
mer the peptidoglycan layer of the cell wall is more exposed. It is currently clas-
sified as a traditional antimicrobial and has been approved by several countries for
its use in foods (Davidson et al. 2013).
Another enzyme, ovotransferrin (commonly known as “conalbumin”) is the
second most important protein in egg white. It is a glycoprotein belonging to the
transferrin family, linked to iron, which transports and sequesters iron ions (Fe+3)
(Ko et al. 2008). Ovotransferrin inhibits several Gram-positive and Gram-negative
bacteria and yeasts including: Escherichia coli, Pseudomonas spp., Streptococcus
mutans, Staphylococcus aureus, Bacillus cereus, Salmonella enteritidis, and
Candida, although it is most effective in Gram-positve bacteria (Ko et al. 2008;
Davidson et al. 2013). The mechanism of ovotransferrin antimicrobial action is not
yet fully understood, but it was thought to have the ability to bind to Fe+3, since this
is the principal mechanism of action of ovotransferrin. More recent studies have,
however, shown that its antimicrobial action is through a direct interaction with the
bacterial surface. This produces an increase in the permeability of the outer
membrane of Gram-negative bacteria, which can subsequently cause damage to the
biological functions of the cytoplasmic membrane (Ibrahim et al. 2000).

Chitosan

Another type of antimicrobial of animal origin used in biopolymeric coatings and

films is chitosan, a linear binary heteropolysaccharide composed of N-acet-
ylglucosamine and glucosamine associated with the b-(1, 4) link (Thakur and
Thakur 2014; Thakur and Voicu 2016). Chitosan is obtained commercially by the
deacetylation of chitin, an abundant constituent of fungi and the exoskeletons of
crustaceans and insects (Pranoto et al. 2005b; Sebti et al. 2005). This compound
could potentially be used in a wide range of applications due to its biodegradability,
biocompatibility, antimicrobial activity, nontoxicity, and versatile physical and
chemical properties (Dutta et al. 2009). The mechanisms of the antimicrobial action
3 Biopolymer Composite Materials with Antimicrobial Effects … 69

of chitin, chitosan, and their derivatives are still largely unknown. However, studies
done by Liu et al. (2004) showed that chitosan increases the permeability of both
the outer and inner membranes of bacterial cells to break point, with the subsequent
leakage of cytoplasmic content. The authors attributed this damage or bactericidal
effect to the electrostatic interaction between the positively charged amino groups
(NH3 þ ) of the chitosan and the negatively charged phosphoryl groups (PO3  ) of
the phospholipidic components of the cell membrane.

2.2.3 Agents of Microbial Origin

Among the antimicrobial compounds of microbial origin incorporated into

biopolymer composites (Thakur et al. 2014a, b) are bacteriocins such as nisin,
pediocin, and enterocin.
Bacteriocins are peptides or proteins produced by different microorganisms that
have an inhibitory or bactericidal effect on other microorganisms. Bacteriocins
produced by lactic acid bacteria are separated into two main groups: lactacins and
lactocins (produced by Lactobacillus spp.), and lysine (produced by Lactococcus
lactis) and pediocins (produced by Pedicoccus acidilactici) (Marugg 1991;
Davidson and Zivanovic 2003). Another group of commonly studied bacteriocins
are enterocins, which are produced by species in the genus Enterococcus. Of these
bacteriocins, the most studied and applied has been nisin, a small heat stable 34
amino acid peptide. Nisin shows bactericidal activity against Gram-positive bacteria
and their spores, but is ineffective against Gram-negative bacteria (Helander and
Mattila-Sandholm 2000). The low antimicrobial effectiveness of nisin against
Gram-negative bacteria is due to the fact that they have a protective outer mem-
brane covering both the cytoplasmic membrane and the peptidoglycan layer which
is also thinner. The inner surface of this outer membrane contains phosphoglyc-
erides, and the outer surface lipopolysaccharide molecules (LPS). These together
create a hydrophilic surface (Nikaido 1996) that acts as a barrier against the pen-
etration of hydrophobic and macromolecular substances. Nisin is a hydrophobic
macromolecule and is thus unable to traverse the outer membranes of
Gram-negative bacteria, which means it cannot reach its site of action (Helander
and Mattila-Sandholm 2000).
On the other hand, the mechanism of action of lysine involves its interaction
with precursors of peptidoglycan and the formation of transient pores in the
cytoplasmic membrane of the target organism (Abee et al. 1994). This leads to a
decrease in proton motive force and a loss of cell ions, amino acids, and ATP
(Crandall and Montville 1998). Lysine can also interfere in the biosynthesis of cell
walls, however, some researchers have suggested that this may simply be a con-
sequence of energy loss and the depolarization of the membrane resulting from pore
formation and the onset of autolysis (Thomas et al. 2000).
Pediocins and enterocins have not been well characterized, and their mode of
action on microorganisms is still not at all clear. Their antimicrobial nature has been
70 K. Álvarez et al.

demonstrated, however, with strong activity reported against pathogenic microor-

ganisms (Sobrino-López and Martín-Belloso 2008; Santiago-Silva et al. 2009).
Bhunia et al. (1991) indicated that pediocin initially causes the loss of viability of
sensitive cells when it binds to nonspecific receptors (probably lipoteichoic acids).
Once these nonspecific sites are saturated, the pediocin molecules bind to specific
receptors, leading to changes in membrane integrity. This causes the cells to lose
potassium ions and other small molecules, as well as lowering their ability to
multiply. In some strains, the cell membrane may also lose its structural integrity
and undergo lysis. In resistant Gram-positive bacteria, the specific receptors are
absent or are not available for bonding, and in Gram-negative bacteria both non-
specific and pediocin-specific receptors are absent. As regards enterocins, Krämer
and Brandis (1975) indicated that they may inhibit DNA, RNA, and protein syn-
thesis, as well as preventing the accumulation of isoleucine, and causing the exit of
already accumulated isoleucine. These effects on biochemical processes may
indicate an obstruction of the energy supply and an alteration in membrane
permeability.

2.2.4 Agents of Inorganic Origin

Inorganic antimicrobial agents, such as zinc oxide, titanium oxide, and silver, have
been frequently incorporated as nanoparticles (NPs) in biopolymer-based matrices
with the purpose of inhibiting the microbial growth of different pathogenic and
deteriorative microorganisms (Table 3). In addition, NPs have helped to improve
the physicochemical and mechanical properties of these matrices, which in recent
years has generated the development of functional biopolymers that can be used as
food packaging materials. Nevertheless, the migration of these NPs toward food is a
key point in determining their antimicrobial efficacy. In addition, NPs have been
subject of discussion due to their potentially counterproductive effects on the health
of consumers (Llorens et al. 2012). Because of this, the food and polymer industries
have been cautious over the use of these types of antimicrobial agents.

3 Factors to Consider in the Choice of Antimicrobials

for Obtaining Functional Biopolymers

Antimicrobial effectiveness depends on several factors related to the food itself,

storage conditions, handling, and the type of microorganism to be controlled. The
preservation of food is best achieved when the following aspects are known and
taken into account: microorganisms to be inhibited, type of antimicrobial and its
concentration, temperature and time of storage, food pH, buffering capacity, and the
presence of other agents that may affect its useful life (Davidson et al. 2013).
3 Biopolymer Composite Materials with Antimicrobial Effects … 71

Table 3 Functional biopolymers made from inorganic nanocomposites with antimicrobial effects
Inorganic Microorganisms References
nanocomposites inhibited
Zinc oxide Escherichia coli Becheri et al. (2008), Liu et al. (2009), Rahman et al.
O157:H7 (2013), El Saeed et al. (2015), Shankar et al. (2015)
Titanium oxide Diaporthe Wist et al. (2002), Chawengkijwanich and Hayata
actinidiae, (2008), Lan et al. (2013), Ammendolia et al. (2014),
Escherichia coli, Othman et al. (2014), Bogdan et al. (2015)
Listeria
monocytogenes,
Penicillium
expansum,
Pseudomonas
aeruginosa,
Salmonella
choleraesuis,
Stayphylococcus
aureus, and
Vibrio
parahaemolyticus
Silver Bacillus Sondi and Salopek-Sondi (2004), Sharma et al. (2009),
megaterium, Valodkar et al. (2010), Yoksan and Chirachanchai
Bacillus subtilis, (2010), Božanić et al. (2011), Dallas et al. (2011),
E. coli, Mohanty et al. (2012), Raji et al. (2012), Wang et al.
Janthinobacterium (2013), Kahrilas et al. (2014), Salunke et al. (2014),
lividum, Taheri et al. (2014), Franci et al. (2015), Ibrahim
Klebsiella (2015), Latif et al. (2015)
pneumonia,
M. smegmatis,
P. aeruginosa,
P. aeruginosa,
S. aureus,
S. epidermidis,
S. flexneri, and
S. typhi

According to Gould (1989), the factors affecting antimicrobial activity may be

classified into microbial, intrinsic, extrinsic, and processing. Microbial factors
influence inherent resistance (vegetative cells vs. spores, differences between
strains), start number and growth rate, interaction with other microorganisms
(competition, antagonism), cell composition (Gram reaction), cell status (lesions),
and the ability to form films. Intrinsic factors are those associated with the food
itself such as nutrients, pH, buffering capacity, oxide-reduction potential, and water
activity. Extrinsic factors include storage time, temperature, atmosphere and relative
humidity. Finally, processing factors affecting antimicrobial activity are food
composition, the replacement of native microflora, changes in microbial popula-
tions, and changes in the microstructure of foods.
72 K. Álvarez et al.

4 Determination of Antimicrobial Activity

The principal method used to evaluate the antimicrobial activity of films derived
from composite biopolymers is the disk diffusion method. This method, one could
say, is an adaptation of the method developed by Davidson and Parish (1989) used
to determine the antimicrobial activity of chemical agents including antibiotics. The
disk diffusion method consists in cutting approximately 1 cm2 discs from the films
to be evaluated and placing them on the surface of solidified agar (previously
inoculated with specific microorganisms) contained in Petri dishes. The petri dishes
with the disks and inoculated agar are then incubated at the optimum growth
temperature for microorganisms for a set time, after which the areas or zones of
inhibition caused by the action exerted by the antimicrobial agents are measured.
Padgett et al. (1998), Cagri et al. (2001), Cha et al. (2002), Rojas-Graü et al. (2006),
Seydim and Sarikus (2006) and Rosales-Oballos et al. (2012) all used this method
and found that it was useful to evaluate antimicrobial activity at different concen-
trations of antimicrobials, since this enabled researchers to pinpoint the optimum
concentration for each one, based on the measurement of the zone of inhibition of
microorganism growth around the discs.
As a general rule, the antimicrobial activity of coatings and films based on
functional polymers can be determined by two methods: dilution and the disk
diffusion method.

4.1 Dilution Method

This method consists in placing different concentrations of the antimicrobial in a

liquid or solid medium, which is then inoculated with the microorganism under
study. For example, Rojas-Graü et al. (2007) evaluated the antimicrobial activity of
an edible film-forming solution prepared from alginate and apple puree against
E. coli O157:H7. The solution also contained oregano, cinnamon, and lemon grass
essential oils and/or their active components: carvacrol, cinnamaldehyde, and citral.
Film-forming solutions containing one or more of the essential oils or their active
components were diluted with saline solution at pH 3.3, and placed in microplate
wells which were then were inoculated with E. coli O157:H7. The concentration of
each compound capable of reducing the population of the microorganism to 50%,
denominated bactericidal activity 50 (BA50), was then determined.

4.2 Disk Diffusion Method

The disk diffusion method consists in placing discs of the films containing different
concentrations of the antimicrobial substance to be evaluated on agar plates
3 Biopolymer Composite Materials with Antimicrobial Effects … 73

Fig. 2 Representation of the disk diffusion method from functional biopolymers

previously inoculated with the microorganism to be studied (Fig. 2). Padgett et al.
(1998), Cagri et al. (2001), Cha et al. (2002), Seydim and Sarikus (2006) and
Rojas-Graü et al. (2006) all used this method and found that it was useful to
evaluate antimicrobial activity at different concentrations of antimicrobials, since
this enabled researchers to pinpoint the optimum concentration for each one, based
on the detection and measurement of the zone of inhibition of microorganism
growth around the discs.

5 Edible Films and Coatings with Antimicrobial Effects

Applied to Foodstuffs

The main purpose of food packaging is to preserve the quality and safety of foods,
from the time of manufacture to the moment of consumption. Thus, packaging
materials must act as a barrier to the passage of food contaminating agents. The
best-known packaging materials that meet this objective are polyethylene or
copolymer-based materials, which have been used by the food industry for over
60 years (Cutter 2006). Nonetheless, one of the limitations of these materials is that
they are not biodegradable, which means that they represent a source of
74 K. Álvarez et al.

environmental pollution. This has motivated scientists to develop packaging

materials made from biodegradable substances that may also be edible, thus
reducing environmental pollution while ensuring that the packaging meets the
principal objective of food protection.
A novel alternative to traditional packaging materials has been the use of edible
films, which can provide barrier properties and limit the passage of oxygen, thus
preventing the contamination and deterioration of foodstuffs. The growth of
microorganisms can be inhibited by simply suppressing oxygen entry, although
films may also be prepared from matrices with antimicrobial properties such as
chitosan. In addition, these functional biopolymers can serve as carriers of
antimicrobial substances of either natural or synthetic origin (Rojas-Graü et al.
2009). One of the advantages of incorporating antimicrobial agents into
biopolymer-based films is the reduction in the rate of diffusion of these compounds
from foods, thus guaranteeing their action over a longer period of time on the
surface of the food, where the greatest contamination usually occurs (Cagri et al.
2004).

5.1 Meat Products

The contamination of meat products usually starts from their external surface due to
the microbial load coming from improper handling, processing machinery, and
packaging. Antimicrobial films can prevent the contamination of meat and meat
products during refrigerated storage, but can also be used to inhibit microbial
growth on the surface of processed fresh products, consequently extending their
useful life (Cagri et al. 2004).
According to the literature, antimicrobial films applied to meat products have
been developed mainly from polysaccharides (alginate, cellulose, methylcellulose,
and hydroxypropylmethylcellulose) and proteins (zein, caseinate, whey, soy, and
gelatin) (Thakur et al. 2016; Thakur and Kessler 2014a, b; Voicu et al. 2016;
Corobea et al 2016).
Oussalah et al. (2004) evaluated the antimicrobial effects of edible films prepared
from milk protein containing oregano essential oil (1%) and/or pepper essential oil
(1%) against E. coli O157:H7 and Pseudomonas spp. inoculated on beef steaks
stored for 7 days at 4 °C. These authors found that the films containing oregano
essential oil were the most effective, with a reduction in microbial populations of
about 1 log cycle compared to control samples and films containing pepper essential
oil. Oussalah et al (2006a) also determined the effect of alginate-based films con-
taining 1% oregano, cinnamon, or savory (Satureja spp.) essential oils on popu-
lations of Salmonella typhimurium or E. coli O157:H7 inoculated into beef filets
stored for 5 days at 4 °C. The authors reported that the films containing oregano or
cinnamon essential oils were most effective against S. typhimurium (causing a
decrease in the population of about 1 log cycle), while films made with oregano
essential oil were the most effective against E. coli O157:H7 (population decreased
3 Biopolymer Composite Materials with Antimicrobial Effects … 75

by 1.97 log cycles). Millette et al. (2007) evaluated the antimicrobial effect of an
alginate-based film incorporating two different amounts of nisin (500 and 1000 IU)
to control the growth of S. aureus inoculated onto beef steaks stored for 14 days at
4 °C. After 7 days, reductions in microorganism populations of 0.91 and
1.86 CFU/cm2 on the beef steaks covered with the films containing 500 or
1000 IU/mL of nisin, respectively, were observed. Studies done by Zinoviadou
et al. (2010) on fresh beef covered with films made from whey protein isolate
containing either sodium lactate (NaL) or e-polylysine (e-PL), showed a reduction
in the growth rate of total native flora and a total inhibition in the growth of lactic
acid bacteria when a concentration of 0.75% of e-PL was added to the films. In
addition, the growth of the total flora and Pseudomonas spp. was significantly
inhibited with the addition of 2% NaL. Ouattara et al. (2002) achieved a slight
reduction (by 0.5 log cycles) in the total counts of microorganisms naturally present
in ground beef during the first 4 days of storage at 4 °C, after covering with a film
prepared from a mixture of proteins (calcium caseinate and soy protein isolate) and
3% powdered thyme, sage, and rosemary. Nevertheless, populations of Brochothrix
thermosphacta, Staphylococcus spp., lactic acid bacteria, coliforms, enterobacteri-
aceae, and Pseudomonas were not significantly affected by the use of this antimi-
crobial film.
On the other hand, Seol et al. (2009) were able to reduce populations of total
microorganisms and E. coli by 1.8 and 2.7 CFU/g, respectively, in chicken breast
covered by a zein-based film containing EDTA (5 mM) and ovotransferrin (25 mg),
stored for 7 days at 5 °C. Similarly, Janes et al. (2002) found that populations of
Listeria monocytogenes decreased by 1.69 or 2.03 log cycles in ready-to-eat
chicken stored for 8 days at 4 °C, by incorporating either nisin (1000 UI/g), or
nisin (1000 UI/g)—calcium propionate (1%), respectively, into a zein-based film
used to cover the chicken. In addition, the antimicrobial effect of nisin alone in the
edible film was intensified (by up to 2.59 log reductions) when the storage tem-
perature was increased to 8 °C.
In other studies, Hong et al. (2009) covered pork loin with a film based on a
gelatin/red algae agarose mixture containing grapefruit seed extract (0.08%) or
green tea extract (2.8%). The antimicrobial action of the film inhibited the growth of
E. coli O157:H7 and reduced populations of L. monocytogenes by about 1 log cycle
after 7 days of storage at 4 °C. Similarly, Kang et al. (2007) evaluated the effect of
an edible coating prepared from pectin containing green tea powder (0.5%) on the
quality of pork burgers packaged in air or under vacuum and stored for 14 days at
10 °C. The authors reported that the initial population of total aerobic microor-
ganisms (104 CFU/mg) declined to undetectable levels after 7 days under vacuum
conditions. However, under normal atmosphere (air) it rose to 105 CFU/mg, and
after 7 days at 10 °C reached 108 CFU/mg.
Dawson et al. (2007) evaluated the antimicrobial effectiveness of soy
protein-based films containing lauric acid (8%) and nisin (400 IU/g), against L.
monocytogenes inoculated into turkey mortadella. The L. moncytogenes population
was successfully reduced by 1 log cycle after 21 days at 4 °C. An even greater
antimicrobial effect was observed by Santiago-Silva et al. (2009) who showed that
76 K. Álvarez et al.

pediocin (25% or 50%) incorporated into an emulsion for cellulose-based films lead
to a reduction in the populations of the pathogens Listeria and Salmonella inocu-
lated into ham slices. These antimicrobial films were, however, more effective
against Listeria than Salmonella: 50% pediocin reduced Listeria populations by 2
log cycles but Salmonella populations by only 0.5 log cycles compared to the
control. Similarly, Gadang et al. (2008) found that populations of L. monoytogenes
and E. coli O157:H7 inoculated into Frankfurt-type sausages stored at 4 °C for
28 days, decreased by 2 log cycles and populations of S. typhimurium by 1 log
cycle when covered with edible films prepared from whey protein containing a
combination of nisin (6,000 UI/g), malic acid (1%), and grape seed extract (0.5%),
plus EDTA (1.6 mg/mL) for S. typhimurium and E. coli O157:H7. Similar results
were reported by Theivendran et al. (2006) who evaluated the bactericidal effect of
nisin (10,000 IU/g) combined with grape seed extract (1%) or green tea extract
(1%) against L. monocytogenes inoculated into Frankfurt-type sausages submerged
in film-forming solutions prepared from soy protein. These authors observed a
decrease the population of L. monocytogenes by more than 2 log cycles after
28 days of storage at 4 or 10 °C. Similarly, Franklin et al. (2004) achieved a
reduction in populations of L. monocytogenes in hot dog-type sausage stored for
28 days at 4 °C by more than 2 log cycles using 2,500 IU/mL of nisin, and the
same result at a lower concentration with films made from
methylcellulose/hydroxypropylmethylcellulose. Finally, Marcos et al. (2008)
achieved a bacteriostatic effect against L. monocytogenes inoculated into cooked
ham during the first 8 days at 1 °C, after incorporating enterocins A and B into
alginate-based films.

5.2 Fishery Products

Films applied to fishery products have been developed mainly using gelatin, either
alone or in combination with chitosan. Gelatin has been widely studied for its
ability to form films and its capacity to protect food from dehydration, light and
oxygen when used as an external cover. It is also one of the first materials proposed
as a carrier of bioactive compounds (Gómez-Guillén et al. 2009). The most
abundant sources of gelatin are the skin and bones of pigs and cattle. However, in
recent years gelatin extracted from fish has become an important alternative.
López-Caballero et al. (2005) evaluated the preservative effect of a coating made
from a mixture of chitosan and fish (Lepidohombus boscii) skin gelatin on cod
burgers. These researchers found that the coating protected the burgers from
microbial spoilage when they were refrigerated and stored for 8 or 11 days,
reporting microorganism counts about 2 log cycles lower than those of control
samples. The coatings were especially effective against Gram-negative bacteria
such as Enterobacteriaceae and Pseudomonas spp. In addition, Gómez-Estaca et al.
(2007) showed that gelatin/chitosan-based edible films containing rosemary and
3 Biopolymer Composite Materials with Antimicrobial Effects … 77

oregano extract lead to a decrease in the total counts of aerobic and sulfide-reducing
bacteria from 2 to 3 log cycles on cold-smoked sardines after 16 days at 5 °C.
In the same way, Datta et al. (2008) reported that coatings derived from calcium
alginate with incorporated oyster lysozyme or hen egg lysozyme applied to smoked
ham was effective in reducing levels of inoculated Salmonella var. Anatum (about
2.2 log cycles) and L. monocytogenes (about 2.7 log cycles).

5.3 Cheese

Fresh cheeses may suffer deterioration through the action of Gram-negative psy-
chrotrophic bacteria (Pseudomonas, Flavobacterium, and Alcaligenes), coliforms,
yeasts, and molds that reach them as postprocessing contaminats. The spoilage of
ripened cheeses, however, is generally due to the surface growth of molds and
yeasts, particularly if they are exposed to oxygen. Furthermore, the use of a slow
growing starter culture can allow the development of pathogenic microorganisms
such as Staphylococcus, Salmonella, Listeria, and enteropathogenic E. coli, which
may either come from raw milk or postprocessing contamination.
A novel alternative to avoid this deterioration, and thus both extend the useful
life of cheeses and ensure the safety of these products, is the use of antimicrobial
films. Cao-Hoang et al. (2010) demonstrated that the application of sodium
caseinate-based films, with the addition of sorbitol as a plasticizer and nisin
(1,000 IU/cm2) as an antimicrobial, reduced a population of Listeria innocua
inoculated onto the surface of ripened cheese by 1.1 log cycles after a storage
period of 7 days at 4 °C.

5.4 Fruits and Vegetables

Edible coatings with antimicrobial agents have gained importance as a new tech-
nology to reduce the deleterious effects produced during the storage of processed
and whole fruits and fresh-cut vegetables. These effects are more severe in the
latter, as the cut damages the cell structure of the tissues. This causes the cell
constituents to leak out, resulting in changes to the color and texture, and shortening
useful microbiological life. In these situations, edible coatings and films can
function as carriers of different preservation agents such as antioxidants, antimi-
crobials, and textural and nutritional preservatives, as well as vitamins and minerals
that counteract the effects of pathogenic and deteriorative microorganisms.
Valencia-Chamorro et al. (2009) studied the antifungal effect of hydroxypropyl
methylcellulose-based coatings containing different mixtures of potassium sorbate
(2%), sodium benzoate (2.5%) and sodium propionate (0.5%) on the postharvest
conservation of whole “Valencia” oranges. The authors observed that the coatings
significantly reduced the effects caused by strains of Penicillium digitatum and
78 K. Álvarez et al.

Penicillium italicum inoculated onto the surface of oranges stored at 20 °C. In

addition they showed that edible coatings with potassium sorbate combined with
sodium propionate were the most effective after a storage period of 60 days at 5 °C.
Park et al. (2005) evaluated the antifungal effects of coatings prepared from chi-
tosan (2%) and hydroxypropyl methylcellulose (1%) with or without added
potassium sorbate (0.3%) on fresh strawberries inoculated with strains of
Cladosporium sp. or Rhizopus sp. and stored for 23 days at 5 °C. The authors
reported that the incorporation of potassium sorbate had no significant additive
effect against the inoculated molds compared to the chitosan coatings alone.
However, all the coatings studied decreased total aerobic and coliform counts.
Caillet et al. (2006) found that carrots inoculated with L. innocua (103 CFU/g)
and covered with an antimicrobial coating prepared from a combination of calcium
caseinate, whey protein isolate, carboxymethylcellulose, and pectin containing
trans-cinnamaldehyde (0.025%) showed a slight reduction (0.5 log cycles) in the
population of the microorganism after 21 days at 4 °C.
Rojas-Graü et al. (2007) evaluated the antimicrobial effect of lemongrass (1 and
1.5%), oregano (0.1 and 0.5%), and vanillin (0.3 and 0.6%) essential oils incor-
porated into film-forming solutions made from alginate and apple puree on native
flora (psychrophilic aerobic, molds, and yeasts) and L. innocua inoculated into
fresh-cut apples. They found that all the antimicrobials studied significantly
inhibited the native flora during 21 days of storage at 4 °C. The lemongrass and
oregano essential oils exerted a greater antimicrobial effect than the vanillin on L.
innocua, however, producing a reduction in its population of up to 4 log cycles.
Likewise, Raybaudi-Massilia et al. (2008a) covered fresh melon pieces inoculated
with S. enteritidis with alginate-based edible coatings containing malic acid, either
alone or combined with cinnamon leaf, palamarosa, and lemongrass essential oils at
0.3 and 0.7%, or their active components (eugenol, geraniol, and citral) at 0.5%.
The coatings significantly reduced the populations of S. enteritidis by about 3–5 log
cycles and inhibited the growth of native flora after storage for more than 21 days at
5 °C. Raybaudi-Massilia et al. (2008b) obtained similar results for fresh apple
pieces inoculated with E. coli O157:H7, this time coated with alginate-based films
containing malic acid, and/or cinnamon bark, clove and lemongrass essential oils at
0.3 and 0.7%, or their active components (cinnamaldehyde, eugenol, and citral) at
0.5%. In this case, the authors achieved population reductions of more than 4 log
cycles after 30 days of refrigerated storage. The growth of the native flora was also
inhibited for over more than 30 days of storage at 5 °C.
Chien et al. (2007) evaluated the effects of edible coatings derived from chitosan
(0.5, 1 and 2%) on the growth of native flora for fresh-cut mango stored for 7 days
at 6 °C. Populations, expressed as the total count of microorganisms, were inhibited
in samples coated with different concentrations of chitosan (5.53, 5.41,
5.30 CFU/g) compared to the control (6.41 CFU/g). Nevertheless, a greater effec-
tiveness of chitosan-based coatings was observed by González-Aguilar et al.
(2009), who found that populations of mesophilic aerobics on fresh-cut papayas,
covered with low molecular weight (2%) and medium molecular weight (1 and 2%)
chitosan coatings, decreased from 2.8 CFU/g to undetectable levels after 14 days
3 Biopolymer Composite Materials with Antimicrobial Effects … 79

storage at 5 °C. These authors also reported that the growth of molds and yeasts
was inhibited on coated papayas compared to the controls. These differences in the
effectiveness of chitosan-based coatings may be due mainly to the type of fruit
studied and the initial microbial load.
Finally, Sangsuwan et al. (2008) evaluated the effectiveness of
chitosan/methylcellulose coatings. These authors observed that populations of
inoculated E. coli in coated fresh-cut melon pieces (5.18 UFC/piece) significantly
decreased during storage to undetectable levels, while populations of
Saccharomyces cerevisiae only slightly decreased in coated fresh-cut melons, but
significantly decreased in coated fresh-cut pineapples, compared to the controls.
These authors also found that the addition of vanillin to the
chitosan/methylcellulose coatings increased their effectiveness against both the
inoculated microorganisms.

6 Impacts on Sensory Attributes

Edible films and coatings are generally consumed while still in place. It is thus
important that the incorporation of compounds such as antimicrobials, antioxidants,
and nutraceuticals does not affect consumer acceptance (Rojas-Graü et al. 2009).
However, some authors have indicated that adding antimicrobial agents to edible
coatings could affect the sensory attributes of food, especially when plant essential
oils are used (Burt 2004). For example, Rojas-Graü et al. (2007) observed that the
sensory attributes of fresh-cut apples were negatively affected when covered with a
coating prepared from alginate-apple puree containing 0.1% (v/v) of oregano
essential oil. Similarly, Raybaudi-Massilia et al. (2008a, b) found that fresh-cut
apple and melon pieces wrapped in coatings derived from alginate and containing
cinnamon, lemongrass, palmarosa, and nail essential oils at 0.7% affected sensory
attributes such as texture, color, and flavor. In order to reduce these effects, it is thus
recommended that essencial oils be used only at very low concentrations and
possibly in combination with alternative preservatives. Nevertheless,
Raybaudi-Massilia et al. (2008a) reported that the addition of 0.3% (v/v) of pal-
marosa essential oil to alginate-based films coating fresh-cut melon pieces actually
turned out to be promising, since this combination was well accepted by the pan-
elists, despite being perceived by them.
In another study, Eswaranandam et al. (2006) found that antimicrobial coatings
derived from soy protein containing the organic acids malic and lactic acid at 2.6%
(w/v) did not affect the sensory properties of fresh-cut melon.
Rojas-Graü et al. (2007) evaluated the sensory quality of fresh-cut apples cov-
ered with an alginate-apple puree film containing vanillin at 0.3% (w/w), finding
that the taste of the coated product was acceptable to the panelists. In contrast,
Walton et al. (2003) noted that a limitation of vanillin, despite being a GRAS
substance, is the strong flavor it imparts when used at the minimum inhibitory
80 K. Álvarez et al.

concentrations (>0.2%) required in some foods. These variations in the results

reported by researchers are probably influenced by the type of food coated.
The effects of other antimicrobial composites such as chitosan, used as a base for
the formulation of antimicrobial coatings, has also been evaluated as regards their
sensory attributes. For example, the sensory qualities of fresh strawberries covered
with chitosan (0.6%) were not significantly affected after 7 days storage at 2 °C
(Han et al. 2005). Similarly, Chien et al. (2007) reported that the sensory attributes
of fresh-cut mango coated with 0.5, 1, and 2% chitosan did not vary significantly
after 3 days of storage at 6 °C, but were significantly altered after 7 days for the
mangos coated with 1 and 2% chitosan.

7 Toxicological Aspects of Composite Biopolymers

with Antimicrobial Effects

Most scientific studies focusing on the development of functional biopolymers

made from natural fillers with antimicrobial effects have been undertaken, funda-
mentally, for their application in the food industry. Nonetheless, there is still a great
vacuum in relation to the toxicological characteristics of these materials. The
application of antimicrobial agents derived from inorganic nanocomposites in food
has been particularly questioned, since they can penetrate the intestinal wall, with
smaller particles tending to spread more quickly through the mucus layer than
larger particles. The rate of diffusion also depends on the charge of the particle;
anionic particles have been shown to traverse the epithelial surface more easily,
while cationic particles remain trapped in the mucus (Szentkuti 1997). In addition,
ligand binding or coating with surfactants can affect the transcellular absorption of
particles in the gastrointestinal tract (Russell-Jones et al. 1999; Hoet et al. 2004).
Furthermore, if encapsulated particles or NPs are protected or sterically hindered,
this can facilitate or inhibit their entry into the bloodstream, resulting in an either
homogeneous or heterogeneous distribution in the human body (Gabor et al. 2004).
Once nano-encapsulated or NP antimicrobials pass through the gastrointestinal
epithelium and into the bloodstream, they may interact with various components of
the blood, such as plasmatic proteins, coagulation factors, platelets, and red and
white blood cells (Nemmar et al. 2002). These interactions can have a substantial
effect on the distribution and excretion of NPs (Dobrovolskaia 2007). For example,
the hydrophobic surfaces of nanospheres are highly susceptible to opsonization and
depuration by the reticulo-endothelial system, resulting in the kidnapping of par-
ticles within organs such as the liver and spleen (Letchford and Burt 2007).
Borm and Kreyling (2004), Oberdorster et al. (2005a), Nel et al. (2006), Powers
et al. (2006), Hagens et al. (2007) and Oberdorster et al. (2007a, b) have observed
that the functionalities of NPs, e.g., particle size, size distribution, potential
agglomeration and surface charge, can change in different biological matrices, and
that this depends on compounds that are present in the matrix and thermodynamic
3 Biopolymer Composite Materials with Antimicrobial Effects … 81

conditions. Interactions between the matrix and the NP’s can also change as a result
of dilution. In practice this means that the appearance of a NP can be expected to
change following sample processing (e.g., freeze-thaw cycles, heating, dilution). It
has, therefore, so far not been possible to establish a single parameter that describes
the possible toxicity of an antimicrobial compound, in particular depending on the
amount employed (SCENIHR 2007). Several authors have indicated that toxicity
tests will have to be undertaken on a case-by-case basis using different
dose-describing parameters (Oberdorster et al. 2005b; Thomas and Sayre 2005;
Powers et al. 2006). In order to give an idea of the magnitude of the studies that
need to be carried out to determine the toxicity of an antimicrobial packaging
material, some toxicological aspects are discussed in Table 4.

Table 4 Toxicological studies in functional biopolymers made from inorganic nanocomposites

Toxicology Effect of inorganic nanocomposites References
study
In vitro NPs may trigger the release of Nel et al. (2006), Donaldson et al.
toxicity reactive species and thereby cause (2007), Donaldson and Seaton
oxidative stress and subsequent (2007), Oberdorster et al. (2007b)
inflammation through interaction
with the reticulo-endothelial system
Toxicokinetics There are experimental data so far Jani et al. (1990), Ballou et al.
suggesting that the characteristics of (2004), Florence (2005), Roszek
NPs (e.g., size, surface charge, et al. (2005), des Rieux et al.
functionalized groups) can influence (2006), Singh et al. (2006), Hagens
the absorption, distribution, et al. (2007)
metabolism, and excretion (ADME)
of NPs present in food. However,
not much is known about the
relationship between these
physicochemical characteristics and
the behavior of NPs in the body
Neurotoxicity Several studies have indicated that Hillyer and Albrecht (2001), Borm
NPs can pass the blood–brain et al. (2006), Silva (2007)
barrier following systemic
availability of PNs. Nevertheless, it
is not clear if this is a generic effect
of all NPs or only a subgroup. This
highlights the need of kinetic
studies, and increased attention
toxicologists to neurotoxicity in
their search for possible effects on
target tissues. Therefore, this fact is
highly relevant for its consideration
(continued)
82 K. Álvarez et al.

Table 4 (continued)
Toxicology Effect of inorganic nanocomposites References
study
Acute toxicity In the past decade, the toxicological Jia et al. (2005), Zhang et al.
results made orally have noted that (2005), Chen et al. (2006), Wang
depending on the particle size (e.g., et al. (2006, 2007, 2008)
copper, selenium, zinc, and zinc
oxide and titanium dioxide NPs),
the matrix in which this content and
the chemical composition of the
NPs, the acute toxicity may occur at
higher doses
Long term Nowadays there is no data on the Wang et al. (2006, 2007, 2008)
toxicity toxicity after chronic or acute low
dose oral exposure. Although
inorganic nanocomposites can have
effects on the immune and
inflammatory and cardiovascular
system, including may provoke
oxidative stress and/or activation of
pro-inflammatory cytokines in the
liver, lungs, brain, and heart. There
could also be effects on the
cardiovascular system, which in
turn may have pro-thrombotic
effects and adverse effects on the
cardiac function (acute myocardial
infarction and adverse effects on
heart rate). Furthermore, they
possibly may produce genotoxicity,
carcinogenesis, and teratogenicity,
but no data on these latter points
that are still available
Reprotoxicity Another aspect to consider and that Fujimoto et al. (2005), Tran et al.
cannot be excluded is the transfer of (2005)
NPs through the placenta, including
excretion via breast milk, which
could lead to embryotoxicity as a
result of exposure to NPs. Data
addressing the distribution of NPs to
the reproductive cells is currently
unavailable. Besides, there are no
clear data showing the distribution
of PNs in the fetus. This leads to the
recommendation that reprotoxicity
needs to be carefully considered
when there is evidence that NPs
pass into the placenta
(continued)
3 Biopolymer Composite Materials with Antimicrobial Effects … 83

Table 4 (continued)
Toxicology Effect of inorganic nanocomposites References
study
Allergenicity Even for conventional chemicals Govers et al. (1994), Borm and
(or little is known on the induction of Kreyling (2004), Gabor et al.
sensitization) food allergy and the type of (2004), des Rieux et al. (2006),
exposure required inducing such Hong et al. (2006), Kabanov
responses. In the case of NPs, this (2006), Dobrovolskaia (2007)
becomes extra prominent for two
reasons. First of all, it is the possible
adjuvant activity of NPs that
introduces additional uncertainty.
And second, because of the actively
charged surfaces of NPs it can
absorb biomolecules as they pass
through the GI tract. This might
result in changed exposure of the
cellular lining of the intestine. In
addition, the surface properties, e.g.,
coatings are important determinants
for the active uptake of
encapsulates, but might also be a
reason for concern. For example,
lectins used for coatings are highly
immunogenic, can be cytotoxic or
induce inflammatory responses and
gastrointestinal irritation

8 Regulatory Status

Edible films and coatings can be classified, according to European and American
regulations, as food products, food ingredients, and substances in contact with food,
or food packaging materials (ED 1995, 1998; USFDA 2006a, b). Nevertheless,
because films and coatings constitute an integral part of the edible portion of food
they should be subject to the same regulations as those governing food ingredients
(Guilbert and Gontard 1995). The main government agencies responsible for
dealing with issues related to food additives are the U.S. Food and Drug
Administration (USFDA), the European Committee for Standardization (CEN),
Codex Alimentarius, the Food and Agriculture Organization (FAO) of the United
Nations and the World Health Organization (WHO) (Raju and Bawa 2006).
The USFDA states that any compound to be included in a formulation should be
“Generally Recognized as Safe” (GRAS) or regulated as a food additive and used
within specific limits (USFDA 2006a, b, 2009, 2010). In Europe, the ingredients
that may be incorporated into film-forming solutions are recognized primarily as
food additives and are named in the list of additives for general purposes, although
pectins, beeswax, polysorbates, fatty acids, lecithin, and arabic and karaya gums,
are mentioned separately as coating applications (ED 1995). In any case, the use of
84 K. Álvarez et al.

these film-forming substances is permitted as long as the principle of quantum satis

is respected. Recently, the CEN introduced specific purity criteria for food additives
(ED 2008).
Due to the fact that edible films and coatings may have ingredients with a
functional effect, the inclusion of these compounds should be mentioned on the
label. In Europe, the use of food additives should always be labeled on the pack-
aging according to their category, for example, antioxidants, preservatives, color-
ings, among others. In the corresponding regulations of most countries, chemical
substances added as antimicrobials are considered as food additives if their main
purpose is the extension of the useful life of the food (Table 5). However, each
country has its own regulations and lists of approved additives (ED 1995, 1998,
2008; USFDA 2006a, b, 2009).
Another important point within the regulatory status is the presence of allergens.
Many edible films and coatings are made from ingredients that could cause allergic
reactions.
Among the substances identified as allergens, those derived from milk, soy-
beans, marine foods (fish, shellfish, crustaceans, etc.), peanuts, walnuts, and wheat
are the most important. Several edible films and coatings are composed of milk
proteins (whey, casein), wheat protein (gluten), soy protein, and peanut protein. The
presence of a known allergen incorporated into an edible film and/or food coating
should thus be very clearly labeled (Franssen and Knochta 2003).

Table 5 Regulatory status of the main antimicrobials incorporated into edible films and coatings
to obtain functional biopolymers
Antimicrobial agent Regulatory Amount permitted References
status
Chitosan GRAS, GRN GMP USFDA
73, GRN 170 (2010)
Essential oils of plants and GRAS, GMP USFDA
their active compounds 21 CFR (2010)
(carvacrol, cinnamon, 182.10,
cinnamaldehyde, citral, 21 CFR
cinnamic acid, citric acid, 182.20,
clove, eugenol, garlic, 21 CFR
geraniol, lemon, lime, 184.1317,
lemongrass, mandarin, 21 CFR
oregano, palmorosa) 182.60,
21 CFR
184.1257
Grape seed extract CAS-RN GMP USFDA
085594-37-2, (2009)
21 CFR
182.20
(continued)
3 Biopolymer Composite Materials with Antimicrobial Effects … 85

Table 5 (continued)
Antimicrobial agent Regulatory Amount permitted References
status
Grapefruit seed extract CAS-RN GMP USFDA
090045-43-5 (2009)
Green tea extract GRN 259 GMP, up to 540 mg/per USFDA
portion in juice (2006a, b)
Lysozyme GRAS, GMP, Losso et al.
GRN 64,  5.5 mg/kg in (2000),
E-1105 Frankfurt-type sausages, USFDA
 4.4 mg/kg in chicken (2006a, b,
and beef products 2010)
Nisin GRAS, GMP, USFDA
GRN 65,  6.9 mg/kg in (2010),
21 CFR Frankfurt-type sausages, FSANZ
184.1538, 5.5–12.5 mg/kg in chicken (2007)
E-234, and beef products
A565,
Codex
standard A-8
Vanillin GRAS, GMP USFDA
21 CFR (2010)
182.60
Organic acids (acetic, GRAS, GMP Doores
lactic, malic, propionic, 21 CFR (1993)
benzoic, and their salts) 184.1005,
21 CFR
184.1021,
21 CFR
184.1221,
21 CFR
184.1069,
21 CFR
182.3640
Inorganic nanocomposites EU In Europe, the legislation EU
(zinc oxide, titanium oxide No. 10/2011 currently applies an overall No. 10/2011,
silver) migration limit of 10 mg Silvestre et al.
constituent per dm2 surface (2011)
area to all substances that
can migrate from food
contact materials up to
foodstuffs. For a liter cubic
packaging containing 1 kg
of food, this equates to a
(continued)
86 K. Álvarez et al.

Table 5 (continued)
Antimicrobial agent Regulatory Amount permitted References
status
migration of 60 mg of
substance per kg of food.
However, with the
exception of a few
materials (specifically
listed in Annex 1 of the
legislation) nanomaterial
risk assessment has to be
performed on a
case-by-case basis
GRAS generally recognized as safe, GRN GRAS news according to USFDA, A Application
according to FSANZ, 21 CFR: title 21 code of regulations of the United States, E code number of
food additives in the European Union, CAS-RN chemical abstract service-registration number,
GMP according to good manufacturing practices

Hen egg proteins such as ovotransferrin (Gal d 3) and lysozyme (Gal d 4), which
reside in the egg white fraction and are used as antimicrobials in edible films, have
traditionally been involved in the development of food allergies. However, clinical
reactions to ovotransferrin and lysozyme have rarely been reported. Nonetheless, a
hypersensitive reaction mediated by immunoglobulin E (IgE) may occur in people
allergic to these enzymes (Pérez-Calderón et al. 2007). Thus, to make the use of
lysozyme and ovotransferin safe, their presence should be clearly identified on the
product label (USDA 2010).

9 Conclusions and Future Perspectives

It has been amply demonstrated that films and coatings with antimicrobial prop-
erties can be developed as novel alternative packaging materials that ensure the
safety of different food types, including meat, fish, fruits, vegetables, and dairy
products such as cheese, as well as extending their useful life. Nonetheless, for
these materials to be successfully applied the following aspects should be taken into
account: food composition, identification of the active component for forming the
film or coating, the antimicrobial to use, the type and number of microorganisms to
control, and the intrinsic, extrinsic, and processing factors that may affect the
product characteristics. In addition, antimicrobial agents should be incorporated in
moderate amounts in order to prevent their negative impact on the sensory char-
acteristics of foods, thus ensuring consumer acceptance.
In the future we expect that materials currently underutilized or discarded by the
food industry, such as vegetable waste, plants, and crustacean shells, among others,
will be used to obtain base materials such as cellulose and chitosan for the for-
mulation of films and coatings, thus producing biodegradable materials that aid
environmental conservation.
3 Biopolymer Composite Materials with Antimicrobial Effects … 87

In recent years there have been a number of research studies on naturally

occurring compounds such as plant extracts that show functional and antimicrobial
properties when tested in vitro against pathogenic microorganisms of importance in
foods, such as L. monocytogenes, E. coli O157:H7, Salmonella spp., S. aureus
among others. Nevertheless, their direct application in foods or as components of
films and coatings has not been fully investigated. For this reason, we expect to see
the continuing emergence of new proposals for the formulation of packaging
materials including natural antimicrobial agents.

Acknowledgements The authors would like to thank National Council of Scientific and
Technical Research (CONICET) (Postdoctoral fellowship internal PDTS-Resolution 2417) and
National University of Mar del Plata (UNMdP) for the financial support.

References

Abee T, Rombouts FM, Hugenholtz J, Guihard G, Letellier L (1994) Mode of action of nisin Z
against Listeria monocytogenes Scott A grown at high and low temperatures. Appl Environ
Microb 60(6):1962–1968
Almajano MP, Carbo R, Jiménez JAL, Gordon MH (2008) Antioxidant and antimicrobial
activities of tea infusions. Food Chem 108(1):55–63
Ammendolia MG, Iosi F, De Berardis B, Guccione G, Superti F, Conte MP, Longhi C (2014)
Listeria monocytogenes behaviour in presence of non-UV-irradiated titanium dioxide
nanoparticles. PLoS ONE 9(1):e84986
Ayala-Zavala JF, Villegas-Ochoa MA, Cuamea-Navarro F, González-Aguilar GA (2005)
Compuestos volátiles de origen natural. Nueva alternativa para la conservación. En: Nuevas
tecnologıas de conservación de productos vegetales frescos cortados. González-Aguilar GA,
Gargea AA, Cuamea-Navarro F (ed). Sonora, Mexico: CIAD AC, pp 314–338
Ballou B, Lagerholm BC, Ernst LA, Bruchez MP, Waggoner AS (2004) Noninvasive imaging of
quantum dots in mice. Bioconj Chem 15(1):79–86
Becheri A, Dürr M, Nostro PL, Baglioni P (2008) Synthesis and characterization of zinc oxide
nanoparticles: application to textiles as UV-absorbers. J Nanopart Res 10(4):679–689
Bégin A, Van Calsteren M-R (1999) Antimicrobial films produced from chitosan. Int J Biol
Macromol 26(1):63–67
Beuchat LR (1996) Pathogenic microorganisms associated with fresh produce. J Food Prot 59
(2):204–216
Bhunia AK, Johnson MC, Ray B, Kalchayanand N (1991) Mode of action of pediocin AcH from
Pediococcus acidilactici H on sensitive bacterial strains. J Appl Bacteriol 70(1):25–33
Bogdan J, Jackowska-Tracz A, Zarzyńska J, Pławińska-Czarnak J (2015) Chances and limitations
of nanosized titanium dioxide practical application in view of its physicochemical properties.
Nanoscale Res Lett 10(1):1–10
Borm PJA, Kreyling W (2004) Toxicological hazards of inhaled nanoparticles—potential
implications for drug delivery. J Nanosci Nanotechnol 4(5):521–531
Borm P, Klaessig FC, Landry TD, Moudgil B, Pauluhn J, Thomas K, Trottier R, Wood S (2006)
Research strategies for safety evaluation of nanomaterials, part V: role of dissolution in
biological fate and effects of nanoscale particles. Toxicol Sci 90(1):23–32
Božanić DK, Djoković V, Dimitrijević-Branković S, Krsmanović R, McPherson M, Nair PS,
Georges MK, Radhakrishnan T (2011) Inhibition of microbial growth by silver–starch
nanocomposite thin films. J Biomat Sci Polym E 22(17):2343–2355
88 K. Álvarez et al.

Burt S (2004) Essential oils: their antibacterial properties and potential applications in foods—a
review. Int J Food Microbiol 94(3):223–253
Cagri A, Ustunol Z, Ryser ET (2001) Antimicrobial, mechanical, and moisture barrier properties
of low pH whey protein-based edible films containing p-aminobenzoic or sorbic acids. J Food
Sci 66(6):865–870
Cagri A, Ustunol Z, Ryser ET (2004) Antimicrobial edible films and coatings. J Food Prot 67
(4):833–848
Caillet S, Millette M, Salmieri S, Lacroix M (2006) Combined effects of antimicrobial coating,
modified atmosphere packaging, and gamma irradiation on Listeria innocua present in
ready-to-use carrots (Daucus carota). J Food Prot 69(1):80–85
Cao-Hoang L, Grégoire L, Chaine A, Waché Y (2010) Importance and efficiency of in-depth
antimicrobial activity for the control of listeria development with nisin-incorporated sodium
caseinate films. Food Control 21(9):1227–1233
Cha DS, Choi JH, Chinnan MS, Park HJ (2002) Antimicrobial films based on Na-alginate and
j-carrageenan. LWT Food Sci Technol 35(8):715–719
Chawengkijwanich C, Hayata Y (2008) Development of TiO2 powder-coated food packaging film
and its ability to inactivate Escherichia coli in vitro and in actual tests. Int J Food Microbiol
123(3):288–292
Chen Z, Meng H, Xing G, Chen C, Zhao Y, Jia G, Wang T, Yuan H, Ye C, Zhao F, Chai Z,
Zhu C, Fang X, Ma B, Wan L (2006) Acute toxicological effects of copper nanoparticles
in vivo. Toxicol Lett 163(2):109–120
Chien PJ, Sheu F, Yang FH (2007) Effects of edible chitosan coating on quality and shelf life of
sliced mango fruit. J Food Eng 78(1):225–229
Commission Regulation (EU) No. 10/2011 of 14 of January 2011 on plastic materials and articles
intended to come in contact with food. Off J Eur Un
Corobea MC, Muhulet O, Miculescu F, Antoniac IV, Vuluga Z, Florea D et al (2016) Novel
nanocomposite membranes from cellulose acetate and clay-silica nanowires. Polym Adv
Technol 27(12):1586–1595
Crandall AD, Montville TJ (1998) Nisin resistance in Listeria monocytogenes ATCC 700302 is a
complex phenotype. Appl Environ Microb 64(1):231–237
Cutter CN (2006) Opportunities for bio-based packaging technologies to improve the quality and
safety of fresh and further processed muscle foods. Meat Sci 74(1):131–142
Cutter CN, Siragusa GR (1997) Growth of Brochothrix thermosphacta in ground beef following
treatments with nisin in calcium alginate gels. Food Microbiol 14(5):425–430
Dallas P, Sharma VK, Zboril R (2011) Silver polymeric nanocomposites as advanced
antimicrobial agents: classification, synthetic paths, applications, and perspectives. Adv
Colloid Interface 166(1):119–135
Datta S, Janes ME, Xue QG, Losso J, La Peyre JF (2008) Control of Listeria monocytogenes and
Salmonella anatum on the surface of smoked salmon coated with calcium alginate coating
containing oyster lysozyme and nisin. J Food Sci 73(2):M67–M71
Davidson PM, Parish ME (1989) Methods for testing the efficacy of food antimicrobials. Food
Technol Chicago 43:148–155
Davidson PM, Zivanovic S (2003) The use of natural antimicrobials. In: Zeuthen P,
BoghSorensen L (eds) Food preservation techniques. CRC Press LLC, Boca Ratón, pp 5–30
Davidson PM, Taylor TM, Schmidt SE (2013) Chemical preservatives and natural antimicrobial
compounds. In: Doyle MP, Beuchat LR (eds) Food microbiology. Fundamentals and frontiers,
4a edn. ASM Press, Washington, pp 765–801
Dawson PL, Carl GD, Acton JC, Han IY (2007) Efecto de películas a base de soya impregnadas
con ácido láurico y nisina sobre el crecimiento de Listeria monocytogenes en mortadella de
pavo. Mundo lácteo y cárnico mayo/junio 13–21
des Rieux A, Fievez V, Garinot M, Schneider YJ, Preat V (2006) Nanoparticles as potential oral
delivery systems of proteins and vaccines: a mechanistic approach. J Control Release 116(1):
1–27
3 Biopolymer Composite Materials with Antimicrobial Effects … 89

Devlieghere F, Vermeulen A, Debevere J (2004) Chitosan: antimicrobial activity, interactions with

food components and applicability as a coating on fruit and vegetables. Food Microbiol 21
(6):703–714
Díaz-Cinco ME, Acedo-Félix E, García-Gálaz A (2005) Principales microorganismos patógenos y
de deterioro. In: González-Aguilar GA, Gardea AA, Cuamea-Navarro F (eds) Nuevas
tecnologías de conservación de productos vegetales fresocs cortados. CIAD, México,
pp 216–240
Dobrovolskaia M (2007) Immunological properties of engineered nanomaterials. Nat Nanotechnol
2(8):469–478
Donaldson K, Seaton A (2007) The Janus faces of nanoparticles. J Nanosci Nanotechnol 7
(12):4607–4611
Donaldson K, Faus S, Borm PJA, Stone V (2007) Approaches to the toxicological testing of
particles. In: Donaldson K, Borm PJA (eds) Particle toxicology. CRC Press, Taylor and Francis
Group, London, pp 299–316
Dong H, Cheng L, Tan J, Zheng K, Jiang Y (2004) Effects of chitosan coating on quality and shelf
life of peeled litchi fruit. J Food Eng 64(3):355–358
Doores S (1993) Organic acids. In: Davidson PM, Branen AL (eds) Antimicrobials in food. Marcel
Dekker, Nueva York, pp 95–136
Duan J, Park S, Daeschel M, Zhao Y (2007) Antimicrobial chitosan-lysozyme (CL) films and
coatings for enhancing microbial safety of mozzarella cheese. J Food Sci 72(9):M355–M362
Dutta PK, Tripathi S, Mehrotra GK, Dutta J (2009) Perspectives for chitosan based antimicrobial
films in food applications. Food Chem 114(4):1173–1182
El Saeed AM, El-Fattah MA, Azzam AM (2015) Synthesis of ZnO nanoparticles and studying its
influence on the antimicrobial, anticorrosion and mechanical behavior of polyurethane
composite for surface coating. Dyes Pigments 121:282–289
Eswaranandam S, Hettiarachchy NS, Meullenet JF (2006) Effect of malic and lactic acid
incorporated soy protein coatings on the sensory attributes of whole apple and fresh-cut
cantaloupe. J Food Sci 71(3):S307–S313
European Parlament and Council Directive N_98/72/EC (ED) (1998) Food additive other than
colors and sweeteners. http://ec.europa.eu/Food/fs/sfp/addit_Flavor/flav11_en.pdf
European Parlament and Council Directive N_2008/84/EC (ED) (2008) Laying down specific
purety criteria on food additives other than colours and sweeteners. http://eur-lex.europa.eu/
lexUniServ/Lex-UriServ.do?url1/4OJ:L2008:253:0001;0175:EN:PDF
European Parlament and Council Directive No. 95/2/EC (ED) (1995) Food additive other than
colors and sweeteners. http://ec.europa.eu/Food/fs/sfp/addit_Flavor/flav11_en.pdf
Fajardo P, Martins JT, Fuciños C, Pastrana L, Teixeira JA, Vicente AA (2010) Evaluation of a
chitosan-based edible film as carrier of natamycin to improve the storability of Saloio cheese.
J Food Eng 101(4):349–356
Fitzgerald DJ, Stratford M, Gasson MJ, Ueckert J, Bos A, Narbad A (2004) Mode of antimicrobial
action of vanillin against Escherichia coli, Lactobacillus plantarum and Listeria innocua.
J Appl Microbiol 97(1):104–113
Florence AT (2005) Nanoparticle uptake by the oral route: fulfilling its potential? Drug Discov
Today 2(1):75–81
Food Standards Australian New Zeland (FSANZ) (2007) Application A565. Use of nisin in
processed meat product. http://www.foodstandards.gov.au
Franci G, Falanga A, Galdiero S, Palomba L, Rai M, Morelli G, Galdiero M (2015) Silver
nanoparticles as potential antibacterial agents. Molecules 20(5):8856–8874
Franklin NB, Cooksey KD, Getty KJ (2004) Inhibition of Listeria monocytogenes on the surface
of individually packaged hot dogs with a packaging film coating containing nisin. J Food Prot
67(3):480–485
Franssen LR, Knochta JM (2003) Edible coatings containing natural antimicrobials for processed
foods. In: Roller S (ed) Natural antimicrobials for minimal processing of food. CRC Press,
Boca Ratón, pp 250–262
90 K. Álvarez et al.

Frei B, Higdon JV (2003) Antioxidant activity of tea polyphenols in vivo: evidence from animal
studies. J Nutr 133(10):3275S–3284S
Fujimoto A, Tsukue N, Watanabe M, Sugawara I, Yanagisawa R, Takano H, Yoshida S, Takeda K
(2005) Diesel exhaust affects immunological action in the placentas of mice. Environ Toxicol
20(4):431–440
Gabor F, Bogner E, Weissenboeck A, Wirth M (2004) The lectin-cell interaction and its
implications to intestinal lectin-mediated drug delivery. Adv Drug Deliver Rev 56(4):459–480
Gadang VP, Hettiarachchy NS, Johnson MG, Owens C (2008) Evaluation of antibacterial activity
of whey protein isolate coating incorporated with nisin, grape seed extract, malic acid, and
EDTA on a turkey frankfurter system. J Food Sci 73(8):M389–M394
Gennadios A, Hanna MA, Kurth LB (1997) Application of edible coatings on meats, poultry and
seafoods: a review. LWT Food Sci Technol 30(4):337–350
Gómez-Estaca J, Montero P, Giménez B, Gómez-Guillén MC (2007) Effect of functional edible
films and high pressure processing on microbial and oxidative spoilage in cold-smoked sardine
(Sardina pilchardus). Food Chem 105(2):511–520
Gómez-Guillén MC, Pérez-Mateos M, Gómez-Estaca J, López-Caballero E, Giménez B,
Montero P (2009) Fish gelatin: a renewable material for developing active biodegradable
films. Trends Food Sci Tech 20(1):3–16
González-Aguilar GA, Monroy-García IN, Goycoolea-Valencia F, Díaz-Cinco ME, Ayala-Zavala
JF (2005) Cubiertas comestibles de quitosano. Una alternative para prevenir el deterioro
microbiano y conserver la calidad de papayas frescas cortadas. In: González-Aguilar G,
Cuamea-Navarro F (eds) Nuevas tecnologías de conservación y envasado de frutas y hortalizas.
Hermosillo, México, CIAD, pp 121–133
González-Aguilar GA, Valenzuela-Soto E, Lizardi-Mendoza J, Goycoolea F, Martínez-Téllez MA,
Villegas-Ochoa MA, Monroy-García IN, Ayala-Zavala JF (2009) Effect of chitosan coating in
preventing deterioration and preserving the quality of fresh-cut papaya ‘Maradol’. J Sci Food
Agric 89(1):15–23
Gould GW (1989) Introduction. In: Gould GW (ed) Mechanisms of action of food preservation
procedures. Elsevier Applied Science, Londres, pp 1–42
Govers M, Termont D, Vanaken GA, Vandermeer R (1994) Characterization of the adsorption of
conjugated and unconjugated bile-acids to insoluble, amorphous calcium-phosphate. J Lipid
Res 35(5):741–748
Guilbert S, Gontard N (1995) Edible and biodegradable food packing. In: Ackermann P,
Jágerstand M, Ohlsson T (eds) Food and packing material—chemical interactions. The Royal
Society of Chemistry, London, pp 159–168
Hagens WI, Oomen AG, de Jong WH, Cassee FR, Sips AJ (2007) What do we (need to) know
about the kinetic properties of nanoparticles in the body? Regul Toxicol Pharm 49(3):217–229
Han C, Lederer C, McDaniel M, Zhao Y (2005) Sensory evaluation of fresh strawberries (Fragaria
ananassa) coated with chitosan-based edible coatings. J Food Sci 70(3):S172–S178
Heggers JP, Cottingham J, Gusman J, Reagor L, McCoy L, Carino E, Cox R, Zhao JG (2002) The
effectiveness of processed grapefruit-seed extract as an antibacterial agent: II. Mechanism of
action and in vitro toxicity. J Altern Complement Med 8(3):333–340
Helander IM, Mattila-Sandholm T (2000) Permeability barrier of the Gram-negative bacterial outer
membrane with special reference to nisin. Int J Food Microbiol 60(2):153–161
Hillyer JF, Albrecht RM (2001) Gastrointestinal persorption and tissue distribution of differently
sized colloidal gold nanoparticles. J Pharm Sci 90(12):1927–1936
Hoet P, Bruske-Hohlfeld I, Salata O (2004) Nanoparticles—known and unknown health risks.
J Nanobiotechnol 2(1):12
Hong S, Leroueil PR, Janus EK, Peters JL, Kober MM, Islam MT, Orr BG, Baker JR, Banaszak
Holl MM (2006) Interaction of polycationic polymers with supported lipid bilayers and cells:
nanoscale hole formation and enhanced membrane permeability. Bioconj Chem 17(3):728–734
Hong YH, Lim GO, Song KB (2009) Physical properties of Gelidium corneum–gelatin blend films
containing grapefruit seed extract or green tea extract and its application in the packaging of
pork loins. J Food Sci 74(1):C6–C10
3 Biopolymer Composite Materials with Antimicrobial Effects … 91

Ibrahim HM (2015) Green synthesis and characterization of silver nanoparticles using banana peel
extract and their antimicrobial activity against representative microorganisms. J Radiat Res
Appl Sci 8(3):265–275
Ibrahim HR, Sugimoto Y, Aoki T (2000) Ovotransferrin antimicrobial peptide (OTAP-92) kills
bacteria through a membrane damage mechanism. BBA Gen Subj 1523(2):196–205
Janes ME, Kooshesh S, Johnson MG (2002) Control of Listeria monocytogenes on the surface of
refrigerated, ready-to-eat chicken coated with edible zein film coatings containing nisin and/or
calcium propionate. J Food Sci 67(7):2754–2757
Jani P, Halbert GW, Langridge J, Florence AT (1990) Nanoparticle uptake by the rat
gastrointestinal mucosa: quantitation and particle size dependency. J Pharm Pharmacol 42
(12):821–826
Jayaprakasha GK, Selvi T, Sakariah KK (2003) Antibacterial and antioxidant activities of grape
(Vitis vinifera) seed extracts. Food Res Int 36(2):117–122
Jia X, Li N, Chen J (2005) A subchronic toxicity study of elemental Nano-Se in Sprague-Dawley
rats. Life Sci 76(17):1989–2003
Jiang Z, Neetoo H, Chen H (2011) Control of Listeria monocytogenes on cold-smoked salmon
using chitosan-based antimicrobial coatings and films. J Food Sci 76(1):M22–M26
Kabanov AV (2006) Polymer genomics: an insight into pharmacology and toxicology of
nanomedicine. Adv Drug Deliver Rev 58(15):1597–1621
Kahrilas GA, Haggren W, Read RL, Wally LM, Fredrick SJ, Hiskey M, Prieto AL, Owens JE
(2014) Investigation of antibacterial activity by silver nanoparticles prepared by
microwave-assisted green syntheses with soluble starch, dextrose, and arabinose. ACS
Sustain Chem Eng 2(4):590–598
Kang HJ, Jo C, Kwon JH, Kim JH, Chung HJ, Byun MW (2007) Effect of a pectin-based edible
coating containing green tea powder on the quality of irradiated pork patty. Food Control 18
(5):430–435
Ko KY, Mendonca AF, Ahn DU (2008) Effect of ethylenediaminetetraacetate and lysozyme on the
antimicrobial activity of ovotransferrin against Listeria monocytogenes. Poult Sci 87(8):1649–
1658
Krämer J, Brandis H (1975) Mode of action of two Streptococcus faecium bacteriocins.
Antimicrob Agents Chemother 7(2):117–120
Lan Y, Lu Y, Ren Z (2013) Mini review on photocatalysis of titanium dioxide nanoparticles and
their solar applications. Nano Energy 2(5):1031–1045
Latif U, Al-Rubeaan K, Saeb AT (2015) A review on antimicrobial chitosan-silver nanocom-
posites: a roadmap toward pathogen targeted synthesis. Int J Polym Mater 64(9):448–458
Letchford K, Burt H (2007) A review of the formation and classification of amphiphilic block
copolymer nanoparticulate structures: micelles, nanospheres, nanocapsules and polymersomes.
Eur J Pharm Biopharm 65(3):259–269
Lin D, Zhao Y (2007) Innovations in the development and application of edible coatings for fresh
and minimally processed fruits and vegetables. Compr Rev Food Sci F 6(3):60–75
Liu H, Du Y, Wang X, Sun L (2004) Chitosan kills bacteria through cell membrane damage. Int J
Food Microbiol 95(2):147–155
Liu Y, He L, Mustapha A, Li H, Hu ZQ, Lin M (2009) Antibacterial activities of zinc oxide
nanoparticles against Escherichia coli O157:H7. J Appl Microbiol 107(4):1193–1201
Llorens A, Lloret E, Picouet PA, Trbojevich R, Fernandez A (2012) Metallic-based micro and
nanocomposites in food contact materials and active food packaging. Trends Food Sci Technol
24(1):19–29
López-Caballero ME, Gómez-Guillén MC, Pérez-Mateos M, Montero P (2005) A chitosan-gelatin
blend as a coating for fish patties. Food Hydrocolloid 19(2):303–311
Losso JN, Nakai S, Charter EA (2000) Lysozyme. In: Naidu AS (ed) Natural food antimicrobial
systems. CRC Press, Boca Ratón, pp 185–210
Marcos B, Aymerich T, Monfort JM, Garriga M (2008) High-pressure processing and
antimicrobial biodegradable packaging to control Listeria monocytogenes during storage of
cooked ham. Food Microbiol 25(1):177–182
92 K. Álvarez et al.

Marugg JD (1991) Bacteriocins, their role in developing natural products. Food Biotechnol 5
(3):305–312
Michalska-Pożoga I, Tomkowski R, Rydzkowski T, Thakur VK (2016) Towards the usage of
image analysis technique to measure particles size and composition in wood-polymer
composites. Ind Crops Prod 92:149–156
Mild RM, Joens LA, Friedman M, Olsen CW, McHugh TH, Law B, Ravishankar S (2011)
Antimicrobial edible apple films inactivate antibiotic resistant and susceptible Campylobacter
jejuni strains on chicken breast. J Food Sci 76(3):M163–M168
Millette MCLT, Le Tien C, Smoragiewicz W, Lacroix M (2007) Inhibition of Staphylococcus
aureus on beef by nisin-containing modified alginate films and beads. Food Control 18(7):
878–884
Mohanty S, Mishra S, Jena P, Jacob B, Sarkar B, Sonawane A (2012) An investigation on the
antibacterial, cytotoxic, and antibiofilm efficacy of starch-stabilized silver nanoparticles.
Nanomed Nanotechnol 8(6):916–924
Natrajan N, Sheldon B (2000) Inhibition of Salmonella on poultry skin using protein and
polysaccharide-based films containing a nisin formulation. J Food Prot 63(9):1268–1272
Nel A, Xia T, Madler L, Li N (2006) Toxic potential of materials at the nanolevel. Science 311
(5761):622–627
Nemmar A, Hoet PHM, Vanquickenborne B, Dinsdale D, Thomeer M, Hoylaerts MF,
Vanbilloen H, Mortelmans L, Nemery B (2002) Passage of inhaled particles into the blood
circulation in humans. Circulation 105(4):411–414
Nguyen VT, Gidley MJ, Dykes GA (2008) Potential of a nisin-containing bacterial cellulose film
to inhibit Listeria monocytogenes on processed meats. Food Microbiol 25(3):471–478
Nikaido H (1996) Other mebrane. In: Neidhardt FC (ed) Escherichia coli and Salmonella cellular
and molecular biology. D.C. ASM Press, Washington, pp 29–47
Nychas GJE, Skandamis PN, Tassou CC (2003) Antimicrobials from herbs and spices. In: Roller S
(ed) Natural antimicrobials for the minimal processing of foods. CRC Press, Washington,
pp 177–199
Oberdorster G, Maynard A, Donaldson K, Castranova V, Fitzpatrick J, Ausman K, Carter J,
Karn B, Kreyling W, Lai D (2005a) Principles for characterizing the potential human health
effects from exposure to nanomaterials: elements of a screening strategy. Part Fibre Toxicol 2:8
Oberdorster G, Oberdorster E, Oberdorster J (2005b) Nanotoxicology: an emerging discipline
evolving from studies of ultrafine particles. Environ Health Perspect 113(7):823–839
Oberdorster G, Oberdorster E, Oberdorster J (2007a) Concepts of nanoparticle dose metric and
response metric. Environ Health Perspect 115(6):A290
Oberdorster G, Stone V, Donaldson K (2007b) Toxicology of nanoparticles: a historical
perspective. Nanotoxicology 1(1):2–25
Othman SH, Abd Salam NR, Zainal N, Kadir Basha R, Talib RA (2014) Antimicrobial activity of
TiO2 nanoparticle-coated film for potential food packaging applications. Int J Photoenergy,
pp 1–6
Ouattara B, Simard RE, Piette G, Bejín A, Holler RA (2000) Inhibition of surface spoilage bacteria
in processed meats by application of antimicrobial films prepared with chitosan. Int J Food
Microbiol 62(1):139–148
Ouattara B, Giroux M, Yefsah R, Smoragiewicz W, Saucier L, Borsa J, Lacroix M (2002)
Microbiological and biochemical characteristics of ground beef as affected by gamma
irradiation, food additives and edible coating film. Radiat Phys Chem 63(3):299–304
Oussalah M, Caillet S, Salmiéri S, Saucier L, Lacroix M (2004) Antimicrobial and antioxidant
effects of milk protein-based film containing essential oils for the preservation of whole beef
muscle. J Agric Food Chem 52(18):5598–5605
Oussalah M, Caillet S, Saucier L, Lacroix M (2006a) Antimicrobial effects of alginate-based film
containing essential oils for the preservation of whole beef muscle. J Food Prot 69(10):2364–
2369
3 Biopolymer Composite Materials with Antimicrobial Effects … 93

Oussalah M, Caillet S, Lacroix M (2006b) Mechanism of action of spanish oregano, chinese

cinnamon, and savory essential oils against cell membranes and walls of Escherichia coli
O157:H7 and Listeria monocytogenes. J Food Prot 69(5):1046–1055
Oussalah M, Caillet S, Salmieri S, Saucier L, Lacroix M (2007) Antimicrobial effects of
alginate-based film containing essential oils o Listeria monocytogenes and Salmonella
thyphimurium present in bologna and ham. J Food Prot 70(4):901–908
Padgett T, Han IY, Dawson PL (1998) Incorporation of food-grade antimicrobial compounds into
biodegradable packaging films. J Food Prot 61(10):1330–1335
Park SI, Stan SD, Daeschel MA, Zhao Y (2005) Antifungal coatings on fresh strawberries
(Fragaria  ananassa) to control mold growth during cold storage. J Food Sci 70(4):M202–
M207
Pen LT, Jiang YM (2003) Effects of chitosan coating on shelf life and quality of fresh-cut Chinese
water chestnut. LWT Food Sci Technol 36(3):359–364
Pérez-Calderón R, Gonzalo-Garijo MA, Lamilla-Yerga A, Mangas-Santos R, Moreno-Gaston I
(2007) Recurrent angioedema due to lysozyme allergy. J Investig Allergol Clin 17(4):264–266
Powers KW, Brown SC, Krishna VB, Wasdo SC, Moudgil BM, Roberts SM (2006) Research
strategies for safety evaluation of nanomaterials. Part VI. Characterization of nanoscale
particles for toxicological evaluation. Toxicol Sci 90(2):296–303
Pranoto Y, Salokhe VM, Rakshit SK (2005a) Physical and antibacte rial properties of
alginate-based edible film incorporated with garlic oil. Food Res Int 38(3):267–272
Pranoto Y, Rakshit SK, Salokhe VM (2005b) Enhancing antimicrobial activity of chitosan films
by incorporating garlic oil, potassium sorbate and nisin. LWT Food Sci Technol 38(8):859–
865
Rahman MAA, Mahmud S, Alias AK, Nor AFM (2013) Effect of nanorod zinc oxide on electrical
and optical properties of starch-based polymer nanocomposites. J Phys Sci 24(1):17–28
Raji V, Chakraborty M, Parikh PA (2012) Synthesis of starch-stabilized silver nanoparticles and
their antimicrobial activity. Part Sci Technol 30(6):565–577
Raju PS, Bawa AS (2006) Food additives in fruit processing. In: Hui YH (ed) Handbook of fruits
and fruit processing. Blackwell Publishing, Ames, pp 145–170
Ravishankar S, Zhu L, Olsen CW, McHugh TH, Friedman M (2009) Edible apple film wraps
containing plant antimicrobials inactivate foodborne pathogens on meat and poultry products.
J Food Sci 74(8):M440–M445
Raybaudi-Massilia RM, Mosqueda-Melgar J (2012) Polysaccharides as carriers and protectors of
additives and bioactive compounds in foods. In: Karunaratn DN (ed) The complex world of
polysaccharides. Editorial InTech, Rijeka, pp 429–454
Raybaudi-Massilia RM, Mosqueda-Melgar J, Martín-Belloso O (2008a) Edible alginate-based
coating as carrier of antimicrobials to improve shelf-life and safety of fresh-cut melon. Int J
Food Microbiol 121(3):313–327
Raybaudi-Massilia RM, Rojas-Graü MA, Mosqueda-Melgar J, Martin-Belloso O (2008b)
Comparative study on essential oils incorporated into an alginate-based edible coating to
assure the safety and quality of fresh-cut fuji apples. J Food Prot 71(6):1150–1161
Raybaudi-Massilia RM, Mosqueda-Melgar J, Soliva-Fortuny R, Martín-Belloso O (2009) Control
of pathogenic and spoilage microorganisms in fresh-cut fruits and fruit juices by traditional and
alternative natural antimicrobials. Compr Rev Food Sci F 8(3):157–180
Rojas-Graü MA, Avena-Bustillos RJ, Friedman M, Henika PR, Martín-Belloso O, McHugh TH
(2006) Mechanical, barrier, and antimicrobial properties of apple puree edible films containing
plant essential oils. J Agric Food Chem 54(24):9262–9267
Rojas-Graü MA, Raybaudi-Massilia RM, Soliva-Fortuny RC, Avena-Bustillos RJ, McHugh TH,
Martín-Belloso O (2007) Apple puree-alginate edible coating as carrier of antimicrobial agents
to prolong shelf-life of fresh-cut apples. Postharvest Biol Technol 45(2):254–264
Rojas-Graü MA, Soliva-Fortuny R, Martín-Belloso O (2009) Edible coatings to incorporate active
ingredients to fresh-cut fruits: a review. Trends Food Sci Technol 20(10):438–447
Rosales-Oballos Y, Raybaudi-Massilia R, Mosqueda-Melgar J, Tapia de Daza MS, Tomé-
Boschian E (2012) Propiedades mecánicas, de barrera y antimicrobianas de películas de
94 K. Álvarez et al.

quitosano y películas de alginato de sodio con aceites esenciales y nisina. Revista de la

Facultad de Farmacia y Bioquímica Universidad de los Andes. Venezuela 54(1):12–16
Roszek B, de Jong W, Geertsma R (2005) Nanotechnology in medical applications: state-of-the-art
in materials and devices. RIVM report. 265001001/2005
Rozes N, Peres C (1998) Effects of phenolic compounds on the growth and the fatty acid
composition of Lactobacillus plantarum. Appl Microbiol Biotechnol 49(1):108–111
Russell-Jones GJ, Veitch H, Arthur L (1999) Lectin-mediated transport of nanoparticles across
Caco-2 and OK cells. Int J Pharm 190(2):165–174
Saito M, Hosoyama H, Ariga T, Kataoka S, Yamaji N (1998) Antiulcer activity of grape seed
extract and procyanidins. J Agric Food Chem 46(4):1460–1464
Salunke GR, Ghosh S, Kumar RS, Khade S, Vashisth P, Kale T, Chopade S, Pruthi V, Kundu G,
Bellare JR, Chopade BA (2014) Rapid efficient synthesis and characterization of silver, gold,
and bimetallic nanoparticles from the medicinal plant Plumbago zeylanica and their application
in biofilm control. Int J Nanomed 9:2635
Sangsuwan J, Rattanapanone N, Rachtanapun P (2008) Effect of chitosan/methyl cellulose films
on microbial and quality characteristics of fresh-cut cantaloupe and pineapple. Postharvest Biol
Technol 49(3):403–410
Santiago-Silva P, Soares NF, Nóbrega JE, Júnior MA, Barbosa KB, Volp ACP, Zerdas ERMA,
Würlitzer NJ (2009) Antimicrobial efficiency of film incorporated with pediocin (ALTA®
2351) on preservation of sliced ham. Food Control 20(1):85–89
Sayanjali S, Ghanbarzadeh B, Ghiassifar S (2011) Evaluation of antimicrobial and physical
properties of edible film based on carboxymethyl cellulose containing potassium sorbate on
some mycotoxigenic Aspergillus species in fresh pistachios. LWT Food Sci Technol 44
(4):1133–1138
Scientific Committee on Emerging and Newly Identified Health Risk (SCENIHR) (2007) Opinion
on: the appropriateness of the risk assessment methodology in accordance with the technical
guidance documents for new and existing substances for assessing the risks of nanomaterials
European Commission Health and Consumer Protection Directorate-General. Directorate C—
Public Health and Risk Assesment C7—Risk Assessment
Sebti I, Martial-Gros A, Carnet-Pantiez A, Grelier S, Coma V (2005) Chitosan polymer as
bioactive coating and film against Aspergillus niger contamination. J Food Sci 70(2):M100–
M104
Seol KH, Lim DG, Jang A, Jo C, Lee M (2009) Antimicrobial effect of j-carrageenan-based edible
film containing ovotransferrin in fresh chicken breast stored at 5 °C. Meat Sci 83(3):479–483
Seydim AC, Sarikus G (2006) Antimicrobial activity of whey protein based edible films
incorporated with oregano, rosemary and garlic essential oils. Food Res Int 39(5):639–644
Shankar S, Teng X, Li G, Rhim JW (2015) Preparation, characterization, and antimicrobial activity
of gelatin/ZnO nanocomposite films. Food Hydrocolloid 45:264–271
Sharma VK, Yngard RA, Lin Y (2009) Silver nanoparticles: green synthesis and their
antimicrobial activities. Adv Colloid Interface 145(1):83–96
Shimamura T, Zhao WH, Hu ZQ (2007) Mechanism of action and potential for use of tea catechin
as an antiinfective agent. Anti Infective Agents Med Chem 6(1):57–62 (Formerly Current
Medicinal Chemistry-Anti-Infective Agents)
Silva GA (2007) Nanotechnology approaches for drug and small molecule delivery across the
blood-brain barrier. Surg Neurol 67(2):113–116
Silvera-Almitrán C, Escobar-Giannini D, Repiso-Ibánez L, Márquez-Romero R (2012)
Aplicaciones de películas y cubiertas comestibles y métodos combinados para mejorar sus
propiedades. In: Olivas-Orozco GI, González-Aguilar GA, Martín-Belloso O, Soliva-Fortuny
R (eds) Películas y recubrimientos comestibles. Propiedades y aplicaciones en alimentos. Clave
Editorial, México, pp 497–518
Silvestre C, Duraccio D, Cimmino S (2011) Food packaging based on polymer nanomaterials.
Prog Polym Sci 36(12):1766–1782
3 Biopolymer Composite Materials with Antimicrobial Effects … 95

Singh R, Pantarotto D, Lacerda L, Pastorin G, Klumpp C, Prato M, Bianco A, Kostarelos K (2006)

Tissue biodistribution and blood clearance rates of intravenously administered carbon nanotube
radiotracers. Proc Natl Acad Sci USA 103(9):3357–3362
Siripatrawan U, Noipha S (2012) Active film from chitosan incorporating green tea extract for
shelf life extension of pork sausages. Food Hydrocolloid 27(1):102–108
Sobrino-López A, Martín-Belloso O (2008) Use of nisin and other bacteriocins for preservation of
dairy products. Int Dairy J 18(4):329–343
Sondi I, Salopek-Sondi B (2004) Silver nanoparticles as antimicrobial agent: a case study on
E. coli as a model for Gram-negative bacteria. J Colloid Interface Sci 275(1):177–182
Stratford M, Eklund T (2003) Organic acids and esters. In: Russell NJ, Gould GW (eds) Food
preservatives. Kluwer Academic/Plenum Publishers, Londres, pp 48–84
Szentkuti L (1997) Light microscopical observations on luminally administered dyes, dextrans,
nanospheres and microspheres in the pre-epithelial mucus gel layer of the rat distal colon.
J Control Release 46(3):233–242
Taheri S, Baier G, Majewski P, Barton M, Förch R, Landfester K, Vasilev K (2014) Synthesis and
antibacterial properties of a hybrid of silver–potato starch nanocapsules by
miniemulsion/polyaddition polymerization. J Mater Chem 2(13):1838–1845
Taylor PW, Hamilton-Miller JM, Stapleton PD (2005) Antimicrobial properties of green tea
catechins. Food Sci Technol Bull 2:71
Thakur VK, Kessler MR (2014a) Free radical induced graft copolymerization of ethyl acrylate
onto SOY for multifunctional materials. Mater Today Commun 1(1–2):34–41
Thakur VK, Kessler MR (2014b) Synthesis and characterization of AN-g-SOY for sustainable
polymer composites. ACS Sustain Chem Eng 2(10):2454–2460
Thakur VK, Thakur MK (2014) Recent advances in graft copolymerization and applications of
chitosan: a review. ACS Sustain Chem Eng 2(12):2637–2652
Thakur VK, Voicu SI (2016) Recent advances in cellulose and chitosan based membranes for
water purification: a concise review. Carbohydr Polym 146:148–165
Thakur VK, Thunga M, Madbouly SA, Kessler MR (2014a) PMMA-g-SOY as a sustainable novel
dielectric material. RSC Adv 4(35):18240–18249
Thakur VK, Grewell D, Thunga M, Kessler MR (2014b) Novel composites from eco-friendly soy
flour/SBS triblock copolymer. Macromol Mater Eng 299(8):953–958
Thakur VK, Vennerberg D, Kessler MR (2014c) Green aqueous surface modification of
polypropylene for novel polymer nanocomposites. ACS Appl Mater Interfaces 6(12):9349–
9356
Thakur MK, Thakur VK, Gupta RK, Pappu A (2016) Synthesis and applications of biodegradable
soy based graft copolymers: a review. ACS Sustain Chem Eng 4(1):1–17
Theivendran S, Hettiarachchy NS, Johnson MG (2006) Inhibition of Listeria monocytogenes by
nisin combined with grape seed extract or green tea extract in soy protein film coated on turkey
frankfurters. J Food Sci 71(2):M39–M44
Thomas K, Sayre P (2005) Research strategies for safety evaluation of nanomaterials, part I:
evaluating the human health implications of exposure to nanoscale materials. Toxicol Sci 87
(2):316–321
Thomas LV, Clarkson MR, Delves-Broughton J (2000) Nisin. In: Naidu AS (ed) Natural food
antimicrobial systems. CRC Press, Boca Ratón, pp 463–524
Tran CL, Danaldson K, Stones V, Fernandez T, Ford A, Christofi N, Ayres JG, Steiner M,
Hurley JF, Aitken RJ, Seaton A (2005) A scoping study to identify hazard data needs for
addressing risks presented by nanoparticles and nanotubes. Institute of Occupational Medicine
(IOM) research report, Edinburgh, United Kingdom
US Food and Drug Administration (USFDA) (2006a) Food additives permitted for direct addition
to food for human consumption 21CFR172, subpart C. Coatings, films and related substances
US Food and Drug Administration (USFDA) (2006b) GRAS substances (SCOGS) database.
http://www.fda.gov/Food/FoodIngredientsPackaging/GenerallyRecognizedasSafeGRAS/
GRASSubstancesSCOGSDatabase/default.htm
96 K. Álvarez et al.

US Food and Drug Administration (USFDA) (2009) Everything added to food in the United
States. http://www.fda.gov/Food/foodingredientspackaging/ucm115326.htm
US Food and Drug Administration (USFDA) (2010) Listing of food additive status Part I and
Part II. http://www.fda.gov/Food/FoodIngre.dientsPackaging/FoodAdditives/FoodAdditive
Listings/ucm091048.htm
Valencia-Chamorro SA, Pérez-Gago MB, del Río MÁ, Palou L (2009) Effect of antifungal
hydroxypropyl methylcellulose (HPMC)-lipid edible composite coatings on postharvest decay
development and quality attributes of cold-stored ‘Valencia’ oranges. Postharvest Biol Technol
54(2):72–79
Valodkar M, Sharma P, Kanchan DK, Thakore S (2010) Conducting and antimicrobial properties
of silver nanowire-waxy starch nanocomposites. Int J Green Nanotechnol Phys Chem 2(1):
P10–P19
Vargas M, Chiralt A, Albors A, González-Martínez C (2009) Effect of chitosan-based edible
coatings applied by vacuum impregnation on quality preservation of fresh-cut carrot.
Postharvest Biol Technol 51(2):263–271
Voicu SI, Condruz RM, Mitran V, Cimpean A, Miculescu F, Andronescu C, Thakur VK (2016)
Sericin covalent immobilization onto cellulose acetate membrane for biomedical applications.
ACS Sustain Chem Eng 4(3):1765–1774
Walton NJ, Mayer MJ, Narbad A (2003) Vanillin. Phytochemistry 63(5):505–515
Wang B, Feng WY, Wang TC, Jia G, Wang M, Shi JW, Zhang F, Zhao YL, Chai ZF (2006) Acute
toxicity of nano- and micro-scale zinc powder in healthy adult mice. Toxicol Lett 161(2):
115–123
Wang J, Zhou G, Chen C, Yu H, Wang T, Ma Y, Jia G, Gao Y, Li B, Sun J (2007) Acute toxicity
and biodistribution of different sized titanium dioxide particles in mice after oral administra-
tion. Toxicol Lett 168(2):176–185
Wang B, Feng WY, Wang M, Wang TC, Gu YQ, Zhu MT, Ouyang H, Shi JW, Zhang F, Zhao YL
(2008) Acute toxicological impact of nano- and submicroscaled zinc oxide powder on healthy
adult mice. J Nanopart Res 10(2):263–276
Wang J, Dong Z, Huang J, Li J, Liu K, Jin J, Ma J (2013) Synthesis of Ag nanoparticles decorated
multiwalled carbon nanotubes using dialdehyde starch as complexant and reductant for
antibacterial purposes. RSC Adv 3(3):918–922
Wang B, Mireles K, Rock M, Li Y, Thakur VK, Gao D, Kessler MR (2016) Synthesis and
preparation of bio-based ROMP thermosets from functionalized renewable isosorbide
derivative. Macromol Chem Phys 217(7):871–879
Wist J, Sanabria J, Dierolf C, Torres W, Pulgarin C (2002) Evaluation of photocatalytic
disinfection of crude water for drinking-water production. J Photoch Photobiol A 147(3):
241–246
Ye M, Neetoo H, Chen H (2008) Effectiveness of chitosan-coated plastic films incorporating
antimicrobials in inhibition of Listeria monocytogenes on cold-smoked salmon. Int J Food
Microbiol 127(3):235–240
Yoksan R, Chirachanchai S (2010) Silver nanoparticle-loaded chitosan-starch based films:
fabrication and evaluation of tensile, barrier and antimicrobial properties. Mater Sci Eng C 30
(6):891–897
Zhang J, Wang H, Yan X, Zhang L (2005) Comparison of short-term toxicity between Nano-Se
and selenite in mice. Life Sci 76(10):1099–1109
Zinoviadou KG, Koutsoumanis KP, Biliaderis CG (2010) Physical and thermo-mechanical
properties of whey protein isolate films containing antimicrobials, and their effect against
spoilage flora of fresh beef. Food Hydrocolloid 24(1):49–59