Meat and Meat Processing

FOOD SCIENCE AND TECHNOLOGY
MEAT AND MEAT PROCESSING
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MEAT AND MEAT PROCESSING
DERRICK B. MCCARTHY
EDITOR
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CONTENTS
Preface vii
Chapter 1 Incorporation of Agro-Industrial By-Products in the
Diets of Animals: Improvement of Meat Quality
Characteristics with Minimal Cost 1
Panagiotis E. Simitzis
Chapter 2 Lipid and Protein Oxidation in Meat 43
Ana P. A. A. Salim, Fernanda M. Viana and
Carlos A. Conte-Junior
Chapter 3 Dry-Cured Meats: Quality, Safety and
Nutritional Aspects 87
Paulina Kęska and Joanna Stadnik
Chapter 4 Analytical Techniques for Trace-Element
Determination in Meat Samples 111
Jefferson S. de Gois, Eduardo S. Chaves and
Aderval S. Luna
Chapter 5 The Use of Electrical Stimulation in
Meat Production 133
Paolo Polidori and Silvia Vincenzetti
Chapter 6 Quality and Nutritional Characteristics of
Donkey Meat 155
Paolo Polidori and Silvia Vincenzetti
vi Contents
Chapter 7 Nitrites/Nitrates in Processed Meat:

Risks and Benefits 179
Małgorzata Karwowska and Anna Kononiuk
Index 195
PREFACE
Meat products occupy quite an exceptional position in the preferences of

the consumers and the interest of food industry, since they provide human
organism with high quality proteins, vitamins and minerals. At the same time,
a constant challenge is faced by the health sector around the world to uncover
the causes associated with the etiology of several diseases. Much attention has
therefore been paid to develop meat products with physiological functions that
promote human health, which is discussed in Chapter One. In Chapter Two,
the authors describe how understanding and monitoring the oxidation
processes is essential for the development of technological strategies to
improve the meat industry. Chapter Three presents the latest achievements in
the production of dry-cured meat products as well as updated scientific and
technological information on the safety, quality and nutritional properties of
this group of meat products. Chapter Four presents an overview of the main
sample preparation methods and analytical techniques applied for trace-
element determination in meat samples. Chapter Five describes what is known
about the effects of the use of electrical stimulation of carcasses of meat
animals, including the effects on meat tenderness and meat sensorial
characteristics. Chapter Six covers the quality of donkey carcass and donkey
meat quality parameters, showing its chemical and sensorial characteristics
(when possible in different muscles) and evaluating the effects of the age of
slaughtering. Finally, in Chapter Seven, the authors present both the benefits
and risks of nitrites and nitrates, as well as the possibilities of using acid whey
as an alternative to nitrites/nitrates in meat products.
Chapter 1 - Meat products occupy quite an exceptional position in the
preferences of the consumers and the interest of food industry, since they
provide human organism with high quality proteins, vitamins and minerals. At
viii Derrick B. McCarthy
the same time, a constant challenge is faced by the health sector around the
world to uncover the causes associated with the etiology of several diseases.
Much attention has therefore been paid to develop meat products with
physiological functions that promote human health.
The use of several by-products of the agro-industry in the diets of farm
animals could combine the improvement of meat quality characteristics and
the preservation of human health with the prevention of problems that are
related with their disposal into the environment. Fortification of meat with
bioactive compounds that are contained in the above by-products appears to
improve the quality of the derived products and protect consumers against
oxidation and the negative implications of free radicals. Additionally, cost of
production is reduced and profit of farmers is increased. However, our
knowledge regarding their activities in animal organism is still rather limited
and a further clarification is necessary in order to establish the regular use of
these by-products in animal meat-producing systems.
Chapter 2 - Oxidation reactions are the main cause of quality loss and
reduction of acceptance on meat matrix. Oxidative deterioration of meat
occurs mainly on lipids and proteins and impairs many attributes, leading to
discoloration, texture modifications and developing off-flavors. Therefore, the
market value is reduced, causing relevant economic losses. Lipid oxidation is a
chain reaction consisted of initiation, propagation, and termination, which can
be performed by enzymatic and non-enzymatic processes. Moreover, protein
oxidation can be defined as a covalent modification, induced by direct
oxidation of the amino acid side chains or conjugation with secondary
compounds that are derived mainly from the oxidation of polyunsaturated fatty
acids. The balance of these reactions depends on the action of prooxidants and
antioxidants agents. Nonetheless, although lipid and protein oxidation appear
to be related processes, the order of occurrence of these events is not well
understood. In order to reduce or prevent oxidative reactions, many strategies
have been applied during pre and post-harvest periods. Furthermore, both
oxidation processes are influenced by intrinsic and extrinsic factors, such as
animal breed and species, muscle types and composition, as well as animal
management, storage temperature and oxygen exposure. Analytical
determination of oxidation on meat matrix is performed mainly by the
measurement of secondary compounds and chemical groups generated during
oxidation processes. Spectrophotometric and chromatographic methods are the
most commonly applied tools to perform these analyses. Understanding and
monitoring the oxidation processes is essential for the development of
technological strategies aiming the quality improvement of meat industry.
Preface ix
Chapter 3 - Dry-cured meat products, such as dry-cured hams, loin or

sausages, are some of the most representative traditional foods that are
produced and consumed in different places throughout the world. In
Mediterranean countries, such as Italy, Spain, Portugal or France, there is a
firmly established tradition of eating dry-cured meat products, and these
products are well known not only on the local market but also on a global
scale. These meat products have a unique taste, smell and texture that result
from the use of specific formulations and production technology. Also, the
different microflora of lactic acid bacteria that are typical for that part of the
world can differentiate the organoleptic qualities of dry-cured meat. Despite
the various production processes of dry-cured meats, curing, fermentation and
ageing constitute typical stages of each. Meat is a valuable nutritious food
which, if left untreated, will spoil within a few days. However, certain
preservative techniques may extend its stability for several days, weeks or
months. Curing is a meat preservation method which uses the preservative
effect of salt (in a high concentration) and, to a lesser extent, sodium nitrite,
mainly in the form of curing salt. Then a period of curing, fermentation and
ageing is required for the full development of the typical sensory and
rheological properties of cured meat products. Today, many people have a
growing awareness of the relation between diet and health aspects. Dry-cured
meat products seem to have a negative image here, mainly because of their
high content of sodium chloride and nitrogen compounds as well as the
possibility of microbial poisoning as a result of non-thermal preservation
methods. Along with the growing concerns regarding consumer health, much
attention is currently being paid to new developments in the production of dry-
cured products which consist of the introduction of modifications in
production technology while maintaining the characteristic flavour, aroma and
consistency of dry-cured meats. Alternative methods for the production of
meat products as compared to curing with nitrite (III) also include sodium
curing with the use of natural sources of nitrate (V) as vegetable-based
ingredients (such as celery juices) with the simultaneous addition of
denitrifying bacteria culture (mainly Staphylococcus) or acid whey. The use of
vacuum conditions and low temperatures during meat ageing is another
solution implemented to extend the shelf life of dry-cured meat products. The
negative image of meat and meat products may be changed by reports of
biologically active peptides isolated from dry-cured meat products,
particularly those with antioxidant (preventing the negative effects of fat
oxidation), antimicrobial (improving the microbiological quality of products)
and antihypertensive (lowering the blood pressure as a counterweight to the
x Derrick B. McCarthy
presence of sodium chloride) properties. This chapter presents the latest

achievements in the production of dry-cured meat products as well as updated
scientific and technological information on the safety, quality and nutritional
properties of this group of meat products.
Chapter 4 - Obtaining information about trace-element content in meat
samples is a critical approach to assess meat quality and its potential impact on
the human health. Different techniques might be used to monitor the trace
element concentration in meat samples, such as inductively coupled plasma
mass spectrometry, ion chromatography, capillary electrophoresis, atomic
absorption spectrometry, among others. Most techniques require at least one
sample pretreatment step to solubilize the analytes from the sample in the
aqueous medium before analysis. Procedures that are available for sample
pretreatment of meat samples include dissolution in alkaline media,
microwave assisted digestion using concentrated acids, combustion
techniques, extraction procedures using acidic media, and direct solid sample
analysis. Therefore, this entry presents an overview of the main sample
preparation methods and analytical techniques applied for trace-element
determination in meat samples.
Chapter 5 - The present chapter describes what is known about the effects
of the use of electrical stimulation of carcasses of meat animals, including the
effects on meat tenderness and meat sensorial characteristics. Electrical
stimulation as a process involves passing an electric current through the
carcass of freshly slaughtered animals. Electrical stimulation has been
extensively used since the 1950s to hasten the onset of rigor mortis and to
modify steps of the glycolytic pathway. Many studies conducted in the USA,
in New Zealand, Australia and Europe have involved a variety of electrical
stimulation methods on different types of meat animals. Data reported in many
studies suggest that electrical stimulation, through hastening rigor changes,
can significantly reduce in the carcasses of meat animals the phenomenon of
cold shortening, one of the major cause of meat toughness. Although it is well
established that electrical stimulation increases the rate of post mortem
glycolysis, other biochemical and biophysical effects have been implicated
with the use of this technology, including the possibility that electrical
stimulation also results in physical disruption of muscle structure. Electrical
stimulation can be considered as a part of the total meat production chain from
slaughter to final sale, and has particular advantages for hot boning, where the
shortening and toughening conditions that would occur for non stimulated
muscles during chilling are avoided.
Preface xi
Chapter 6 - Meat has exerted a crucial role in human evolution and is an

important component of a healthy and well balanced diet due to its nutritional
richness. The aim of the present chapter is to shed light on the nutritional
composition of donkey meat and the implications for human health. Donkeys
are not perceived as multi-use animals. Cattle, buffaloes and camels are
usually kept for their milk and their meat as well as for work. In many areas
donkeys are not sold for their meat. One of many exceptions is Lesotho where
donkeys are culled for meat when they are considered too old to work, and for
this reason donkeys are relatively expensive in this Country. In the rest of the
world, the lower cost of donkeys makes them more affordable to small
farmers. On the other hand, donkey meat can be considered a good alternative
in red meat consumption, being a dietary meat. Donkey meat is in fact
characterized by low fat, low cholesterol content, a favourable fatty acid
profile and is rich in iron. Today consumers are health conscious and demand
high quality food products; they require leaner meat, with less fat (the minimal
fat level required to maintain juiciness and flavour) and a consistent quality.
Ultimately, the success of any food product is determined by the consumer’s
acceptance. Meat quality and acceptability is determined by its physico-
chemical characteristics, although consumer preferences for meat are difficult
to define. In this context, this chapter will describe the quality of donkey
carcass and donkey meat quality parameters, showing its chemical and
sensorial characteristics (when possible in different muscles) and evaluating
the effects of the age of slaughtering.
Chapter 7 - High intake of processed meat has been associated with
increased risk of many diseases. Some additives used in processed meat
production, especially nitrites/nitrates, are of concern. Nitrite has been in the
spotlight for decades because of its involvement in the formation of nitroso-
compounds, such as carcinogenic N-nitrosoamines. Particularly red meat and
meat products, which are a good source of heme iron, have been related to
increased risk of cancer. For this reason there has been pressure on the
consumer side to eliminate the use of nitrites/nitrates in meat product
formulations. On the other hand, the elimination of nitrites/nitrates from the
production of meat products is problematic as their use contributes to color
development, flavor, antioxidant properties and microbiological stability.
Nitrites and/or nitrates are used to improve meat product safety as they prevent
the growth of most pathogenic and spoilage organisms, including Clostridium
botulinum and Staphylococcus aureus. Moreover, nitrite retards oxidative
rancidity during storage of the meat product. During the curing process, nitrite
is converted to nitric oxide (NO) via reduction reactions by curing adjuncts
xii Derrick B. McCarthy
(sodium ascorbate, sometimes bacteria), which may act as a bacteriocidal

agent by blocking the thiol group that comprises the active center of nonheme
iron-sulfur proteins. However, added nitrite is usually not completely degraded
to NO during the curing process, and excess amounts of residual nitrite have
been associated with the formation of carcinogenic nitrosamines.
In the authors’ previous studies, acid whey was investigated for its
potential use as a substitute for nitrites/nitrates in meat products. The obtained
results showed that acid whey had a positive effect on the physicochemical
qualities of non-nitrite meat products. In this context, the aim of this paper is
to present both the benefits and risks of nitrites/nitrates as well as the
possibilities of using acid whey as an alternative to nitrites/nitrates in meat
products.
In: Meat and Meat Processing ISBN: 978-1-53612-210-7
Editor: Derrick B. McCarthy © 2017 Nova Science Publishers, Inc.
Chapter 1
INCORPORATION OF AGRO-INDUSTRIAL
BY-PRODUCTS IN THE DIETS OF ANIMALS:
IMPROVEMENT OF MEAT QUALITY
CHARACTERISTICS WITH MINIMAL COST
Panagiotis E. Simitzis*, PhD

Department of Animal Breeding and Husbandry,
Faculty of Animal Science and Aquaculture,
Agricultural University of Athens, Greece
ABSTRACT
Meat products occupy quite an exceptional position in the
preferences of the consumers and the interest of food industry, since they
provide human organism with high quality proteins, vitamins and
minerals. At the same time, a constant challenge is faced by the health
sector around the world to uncover the causes associated with the
etiology of several diseases. Much attention has therefore been paid to
develop meat products with physiological functions that promote human
health.
The use of several by-products of the agro-industry in the diets of
farm animals could combine the improvement of meat quality
characteristics and the preservation of human health with the prevention
of problems that are related with their disposal into the environment.
*
Corresponding Author: pansimitzis@aua.gr.
2 Panagiotis E. Simitzis
Fortification of meat with bioactive compounds that are contained in the

above by-products appears to improve the quality of the derived products
and protect consumers against oxidation and the negative implications of
free radicals. Additionally, cost of production is reduced and profit of
farmers is increased. However, our knowledge regarding their activities
in animal organism is still rather limited and a further clarification is
necessary in order to establish the regular use of these by-products in
animal meat-producing systems.
Keywords: agro-industrial by-products, meat, oxidation, antioxidants
1. LIPID OXIDATION AND ANTIOXIDANTS IN BIOLOGICAL

AND FOOD SYSTEMS
1.1. Oxidation Procedures
Meat products are essential components in the diets of developed

countries, since they are important sources for protein, fat, essential amino
acids, minerals, vitamins and other nutrients. Meat consumption is mainly
affected by the product characteristics (species, sensory and nutritional
properties, safety, price etc.) but also consumer and environment-related
factors closely linked to health, social and economic status. However, lipids
and proteins of meat are easily susceptible to oxidative damage due to the
rapid depletion of endogenous antioxidants after slaughter. Oxidation of lipids
and free radicals’ production are natural processes occurring in meat
processing systems and result in undesirable off-flavors (rancid) and
discoloration (fading, browning or degradation) that make meat unpalatable
and could cause its rejection. In addition, there is also nutritive losses due to
degradation of essential fatty acids (phenylalanine, tryptophan etc.) and
vitamins. Oxidative stability of meat is therefore the main concern for all the
stakeholders involved in the animal production chain, including the primary
producers, processors, distributors and retailers (Embuscado, 2015; Falowo et
al., 2014; Simitzis and Deligeorgis, 2011).
Lipids of meat could be categorized as triacylglycerides, phospholipids
and sterols. These components are chemically unstable and therefore
susceptible to oxidation especially during post-mortem handling and storage.
Meat mincing, cooking and other processing prior to refrigerated storage
disrupt muscle cell membranes accelerating lipid oxidation (Falowo et al.,
Incorporation of Agro-Industrial By-Products in the Diets … 3
2014). Oxidation procedure further destroys the membrane structure, disturbs

transport processes and causes losses in the function of the cell organelles.
Polyunsaturated fatty acids (PUFAs) are responsible for the maintenance of
physiologically important cell membrane properties including fluidity and
permeability. The peroxy radicals react with PUFAs and form hydroperoxides
(ROOH), which later decompose to produce the volatile non-radical aromatic
compounds (aldehydes, alkanes and conjugated dienes etc.) that adversely
affect lipids, pigments, proteins, carbohydrates vitamins and the overall quality
of meat products by causing loss of nutritive value and limiting shelf-life
(Pisoschi and Pop, 2015). Many factors seem to affect lipid oxidation in
animal tissues, such as species, sex, age, anatomical location, diet,
environmental – storage temperature, light, exposure to air – oxygen access
and phospholipid composition and content. On the other hand, various
biochemical components involving trace minerals, enzymes and vitamins
protect the cellular structure and function against oxidative damage (Gordon,
2001).
1.2. Antioxidants and Antioxidant Systems
Living organisms have developed specific mechanisms that are known as

“the endogenous antioxidant system” and are located in organelles, subcellular
compartments or the extracellular space with the intention to prevent oxidative
injury in the cell. These antioxidants can act at different steps of the oxidative
radical process (initiation, propagation and chain termination) (Yin et al.,
2011). The first level of the antioxidant defense system consists of the
antioxidant enzymes (glutathione peroxidase, catalase and superoxide
dismutase) and the metal-binding proteins (transferrin, ferritin, lactoferrin,
ceruloplasmin) that remove precursors of free radicals, inactivate catalysts or
limit the radicals initiators due to their bond with metals such as iron and
copper as metal chelators to stabilize them in an inactive or insoluble form.
The first level is usually not sufficient to completely prevent free radical
formation, since some peroxyl radicals escape and several chain breaking
antioxidants (vitamin E, vitamin A, carotenoids, ascorbic acid etc.) inhibit
peroxidation by donating electrons to break and terminate the oxidation cycle,
by scavenging peroxyl radical intermediates and by keeping the chain length
of the propagation reaction as small as possible (second level of cellular
antioxidant defense). Even the second level is sometimes not able to prevent
lipid peroxidation and the integrity of the biological molecules is damaged. In
this case, the third level of antioxidant defense that includes lipolytic,
proteolytic and other enzymes is activated and eliminates or repairs the
damaged molecules. The cooperation between the three levels of antioxidant
defense in the cell is vital for maximum protection from the deleterious effects
of free radicals (Falowo et al., 2014; Haliwell and Gutteridge, 1996; Pisoschi
and Pop, 2015).
Cells can usually tolerate mild oxidative stress by additional synthesis of
antioxidants in order to restore the critical balance between antioxidants and
free radicals. Stress conditions of environmental (increased temperature,
humidity etc.) or nutritional (high levels of PUFAs, toxicants, deficiencies of
vitamins or elements etc.) origin could increase production of free radicals that
cause damage and disrupt integrity of cell membranes. When free radicals are
produced in excess and the antioxidant defense system cannot tolerate them,
damage to the cellular components and harmful autoimmune responses are
observed (Barbieri and Sestili, 2012). Natural antioxidants (vitamins,
carotenoids, phenolics etc.) and optimal levels of several elements (Mn, Cu,
Zn, Se etc.) in animal diet contribute in the maintenance of efficient
antioxidant levels in animal tissues and prevent the negative effects of free
radicals (Pisoschi and Pop, 2015).
1.3. Antioxidants, Meat Products and Nutrition-Related Diseases
A great interest in applying antioxidants from natural sources to increase

the shelf life of meat products is nowadays considerably enhanced due to
consumer preference for natural occurring ingredients and concerns about the
possible toxic effects of synthetic antioxidants (butylated hydroxyanisole
(BHA), butylated hydroxytoluene (BHT), tert-butylhydroquinone (TBHQ),
gallates etc). Natural antioxidants are mainly phenolic compounds and the
most important groups are the tocopherols, flavonoids and phenolic acids.
Food antioxidants are readily assimilated by the organism and render
antioxidant activity mainly as a result or their role as reducing agents,
hydrogen donors, and singlet oxygen quenchers. Some antioxidants also have
the ability to chelate metal ions which act as catalysts in oxidation reactions.
At the same time, lipoxygenase and cyclooxygenase action is inhibited,
enzymes that are responsible for the development of oxidative rancidity in
foods (Embuscado, 2015).
Meat products are nowadays intended not only to cover the nutritional
needs but also contribute in fortifying organism against nutrition-related
diseases. The involvement of the reactive oxygenated/nitrogenated species,

such as superoxide anion radical, hydroxyl, alkoxyl and lipid peroxyl radicals,
nitric oxide and peroxynitrite in several diseases has been extensively
demonstrated in the literature (Pamplona and Costantini, 2011). The unbalance
between these oxidation products and the components of the antioxidant
defense system may trigger oxidative damage in the cell, described as over-
expression of oncogene genes, generation of mutagen compounds, promotion
of atherogenic activity, senile plaque occurrence or inflammation. The
deterioration of cell structure and function can further lead to cardiovascular
diseases (coronary heart disease, atherosclerosis), diabetes, neurodegeneration,
kidney diseases, osteoporosis and several forms of cancer (breast, prostate,
pancreas, esophagus, stomach, colon etc.) (Pisoschi and Pop, 2015; Roleira et
al., 2015; Shen et al., 2012).
Dietary antioxidants appear as a first rated alternative that could combat
the multiple health risks associated with oxidative stress and contribute in the
maintenance of functional integrity of cell organelles (Falowo et al., 2014).
The consumption of meat rich in antioxidants can reinforce the activity of the
endogenous antioxidant system against degenerative diseases linked to
oxidative stress and reactive oxygenated species-related tissue damage.
Antioxidants contained in animal products are often lost during processing,
handling or storage due to their rapid depletion after slaughter, necessitating
their further exogenous supplementation. In general, antioxidants can
effectively minimize rancidity and retard lipid peroxidation without any
damage to sensory or nutritional properties of meat products, resulting in
maintenance of quality and enhancement of shelf life. The traditional practice
of adding antioxidants during processing can still play a very important role
since the added compounds have the potential for enhancing the activity of the
inherent antioxidant system by inhibiting degradation of product, delaying
onset of rancid flavors and stabilizing the color (Gordon et al., 2001; Karre et
al., 2013; Shah et al., 2014).
Furthermore, the dietary supplementation with antioxidants appears as a
simple and convenient strategy to uniformly introduce a natural antioxidant
into phospholipid membranes, where it may effectively inhibit the oxidative
reactions at their localized sites and appears as a more effective way of
retarding lipid oxidation of meat products compared to post mortem addition
(Govaris et al., 2004). However, the main problem with the dietary
supplementation of antioxidants is usually their low bio-availability. Their
concentration in the diet could be very high, but their levels in blood are low
and their concentration in muscle tissues usually is negligible (Surai, 2014).
Each year, medical costs for major chronic diseases related with the action
of free radicals, such as cardiovascular disease, diabetes, cancer, osteoporosis
and obesity are worldwide increased. Many of these disorders are directly
linked to the human diet and could be prevented by adopting a healthier food
supply as a preventive health care strategy. Supplementation of animal diets
with antioxidants could serve in this direction, since they diminish free radical
induced tissue damage by preventing the formation of radicals, by scavenging
them, or by promoting their decomposition in the derived animal products that
are further consumed by humans (Decker and Park, 2010).
APPLICATION OF AGRO-INDUSTRIAL BY-PRODUCTS

IN ANIMAL PRODUCTION AND EFFECTS ON MEAT
QUALITY CHARACTERISTICS
2.1. Introduction
A constant increase of livestock production cost is observed due to the

high prices of the main dietary ingredients, i.e., cereal grains and soybean
products. Low input feeding strategies based on the agro-industrial by-
products are therefore necessary (Vasta and Luciano, 2011). Apart from their
nutritional value, these by-products contain several secondary compounds that
fortify animal products and boost human health. Due to their low cost, they are
considered as a cheap source of functional additives – nutraceuticals and their
inclusion in animal diets minimizes the environmental effects induced by their
disposal (due to their high organic load) and enables the sustainability of high-
added value ingredients inside food chain. However, the variation in their
composition among different regions and the limited knowledge concerning
aspects such as their application (mode, dose etc.), functional pathways,
bioactivity, bioavailability and interactions with other ingredients in animal
gastrointestinal tract pose limitations on their regular use (Galanakis, 2012;
Tufarelli et al., 2013).
Alternative feeding strategies also gain ground among the stakeholders
due to the constant increased demand for clean, natural or eco-green animal
products that are safe and healthy and obtained from sustainable and drugs-
free farming systems (Font i Furnols et al., 2011). Agro-industrial by-products
could be used either as raw material or after processing, such as drying for the
reduction of moisture content or other advanced techniques for the collection
of specific compounds well known as nutraceuticals. These compounds

usually possess antibacterial, coccidiostatic, anthelmintic, antiviral, anti-
inflammatory and antioxidant properties that could improve health status of
animals and the quality of the derived meat products (Zhang et al., 2010).
2.2. Apple Pomace
Apple (Malus domestica) processing generates huge quantities of solid

residues (~ 20-30%) that consist of a mixture of skin, pulp and seeds and are
generally known as ‘apple pomace.’ These residues are highly biodegradable
and their disposal represents a serious environmental problem (Dhillon et al.,
2013). Alternatively, they could serve as an important and inexpensive source
of food and feed ingredients, although vary in nutrient density depending on
the morphology of the original stock and the extraction technique used. As
presented in Table 1, apple pomace fortified meat products appear to possess
improved quality properties (Jung et al., 2015), since they contain suitable
enzyme activities for meat protein stabilization (Lantto et al., 2006). Apple by-
products can be profitably utilized to develop fibre enriched meat products,
such as sausages (Yadav et al., 2016). Moreover, apple polyphenols appear to
successfully retard lipid oxidation of chinese-style sausages (Yu et al., 2015),
of both pork and beef sliced cooked cured hams (SCCH) during chilled storage
and inhibit discoloration in pork SCCH (Sun et al., 2010). At the same time,
they prevent linoleic acid and cholesterol oxidation in sausages (Osada et al.,
2000) and have an inhibitory effect on the formation of heterocyclic amines in
pan fried beef patties (Sabally et al., 2016).
Apple pomace could also be used as a component of animal diets, since it
contains a high content of dietary fiber, polyunsaturated fatty acids and
desirable bioactive substances such as vitamins, anthocyanins and phenolic
acids. As a result, tissues of animals are fortified with these natural compounds
and the quality of the derived products is improved (Pieszka et al., 2015). As it
has been demonstrated, dietary addition of a fermented apple supplement (2%)
seems to improve growth, feed intake, and carcass weight in finishing pigs. At
the same time, meat moisture and crude protein contents are decreased, pH24
and water holding capacity values are increased and marbling of fresh meat
but also tenderness, juiciness, flavor and overall acceptability of cooked meat
are improved (Lee et al., 2009). Improvement of growth parameters and meat
antioxidant capacity were also observed after the inclusion of 1% fermented
apple pomace in the diets of broilers (Kang et al., 2009).
Table 1. Effects of agro-industrial fruit by-products addition on the quality characteristics of meat products
By-product Level Meat product Effect Reference

Apple pomace (PM) 3.5% PM Pork meat Protein stabilization. Lantto et al., 2006
or polyphenol (PL) 5-15% PM Mutton nuggets Reduction of pH, hardness - texture, flavor and Huda et al., 2014
overall acceptability scores.
1-5% PM Goshtaba (mutton) Reduction of pH values and thiobarbituric acid Rather et al., 2015
reactive substances.
3-9% PM Chicken sausages Reduction of moisture and protein content. No effect Yadav et al., 2016
on sensory attributes.
0.1% PL Pork sausages Inhibition of linoleic acid and cholesterol oxidation. Osada et al., 2000
3-10% PL Cured beef and pork Reduction of lipid oxidation values. No effect on Sun et al., 2010
hams protein oxidation.
5% PL Chinese-style Exhibition of phospholipid protective capacity and Yu et al., 2015
sausages improvement of oxidative stability.
0.1-0.3% PL Pan fried beef patties Minimization of the formation of genotoxic Sabally et al., 2016
heterocyclic aromatic amines.
Citrus fibre (CF) or 0.5-2%CF Bologna sausages Reduction of residual nitrite levels and lipid Fernandez-Gines
citrus fibre washing oxidation values. Increase of redness and hardness. et al., 2003
water (CFWW) or 2.5-10% CF Cooked & dry-cured Reduction of residual nitrite levels and pH values. Fernandez-Lopez
lemon albedo (LA) sausages Increase of lightness. et al., 2004
0.5-2% CF Dry cured sausages Reduction of residual nitrite levels and thiobarbituric Fernandez-Lopez
acid values. et al., 2007
1-2% CF Dry fermented Reduction of residual nitrite levels and increase of Fernandez-Lopez et
sausages micrococcus growth. al., 2008
1% CF Mortadella Reduction of residual nitrite levels and lipid Viuda-Martos et al.,
oxidation values. Increase of hardness and lightness 2010a
& reduction of yellowness.
1% CF Non fermented dry No effect on pH values and color properties. Sayas-Barberá et al.,
cured sausages Reduction of residual nitrite levels and increase 2012
of lactic acid bacteria growth.
5-10% CFWW Bologna sausages Reduction of residual nitrite levels and lipid Viuda-Martos et al.,
oxidation values. 2009b
5-10% CFWW Bologna sausages Reduction of moisture content, residual nitrite Viuda-Martos et al.,
levels and lipid oxidation values. No effect on 2010c
color, pH and textural properties.
2.5-10% LA Non fermented dry Reduction of residual nitrite levels and Aleson-Carbonell
cured sausages thiobarbituric acid values. et al., 2004
2.5-7.5% LA Beef burgers Reduction of thiobarbituric acid values, hardness Aleson-Carbonell
and yellowness. et al., 2005
2.5-5% LA Bologna sausages Reduction of residual nitrite levels. Fernandez-Gines
et al., 2004
Grape pomace (GP) or seed 0.3-0.6% GP Chicken patties Improvement of oxidative stability. Sayago-Ayerdi
extract (GSE) or seed flour et al., 2009b
(GSF) or dietary fiber 4-16% GSE Minced turkey meat Improvement of oxidative stability. Mielnik et al., 2006
(GDF) 1% GSE Turkey frankfurters Control of the growth and recontamination of L. Theivendran et al.,
monocytogenes, E. coli and S. typhimurium. 2006; Sivarooban
et al., 2007
0.3% GSE Beef patties Inhibition of microbial spoilage. Improvement of Banon et al., 2007
redness and lipid oxidation values. No effect on
sensory attributes.
Table 1. (Continued)

0.1% GSE Pork patties Reduction of thiobarbituric acid values. Increase Carpenter et al., 2007
of redness. No effect on mesophilic plate counts,
pH values and sensory attributes.
0.1% GSE Chicken thigh meat Improvement of oxidative stability, mitigation of Brannan, 2008; 2009
the prooxidative effects of NaCl. Reduction of
lightness and yellowness & increase of redness.
0.02% GSE Pork patties Improvement of oxidative stability. No effect on Sasse et al., 2008
redness and yellowness.
5% GSE Processed chicken Improvement of the oxidative stability. Shirahigue et al.,
meat 2010
0.06% GSE Pork burgers Improvement of the oxidative stability, color Garrido et al., 2011
attributes and overall acceptability.
0.1-0.5% GSE Beef sausages Improvement of the oxidative stability, odor and Kulkarni et al., 2011
color scores.
1-10% GSE Beef Patties Inhibition of Enterobacteriaceae, coliform Sagdic et al., 2011
bacteria, lipolytic bacteria, yeasts and molds
GSE (60 mg of Chicken meat Reduction of color parameters and lipid Selani et al., 2011
TP/kg) oxidation values
2-8% GSE Fried beef patties Improvement of oxidative stability. No effect on Gibis and Weiss,
sensory attributes. 2012
0.1% GSE Restructured Reduction of thiobarbituric acid values, total Reddy et al., 2013
mutton slices psychrophilic and coliform counts. Improvement
of color, flavor, juiciness and overall palatability.
0.2-1% GSE Dry cured sausage Improvement of oxidative stability. Increase of Lorenzo et al., 2013
(chorizo) redness & reduction of hardness and total volatile
compounds.
Level Meat product Effect Reference
1% GSE Pork patties Improvement of oxidative stability and control of Lorenzo et al., 2014
color deterioration. Inhibition of lactic acid
bacteria, Pseudomonas and psychotropic aerobic
bacteria.
0.3% GSE Chicken nuggets Reduction of total plate, psychrophilic, yeast and Kaur et al., 2015
mold counts and thiobarbituric acid reactive
substances levels. Improvement of sensory
attributes.
0.5-2% GDF Chicken Reduction of lightness, yellowness and lipid Sayago-Ayerdi et al.,
hamburgers oxidation values. Increase of redness. 2009a
0.5-5% GSF Beef frankfurters Reduction of color parameters values and lipid Özvural and Vural,
oxidation values. 2011
Pomegranate powder (PP) PP (50-200 mg Chicken patties Reduction of thiobarbituric acid reactive Naveena et al.,
or peel extract (PE) gallic acid substances levels and lightness values. No 2008a; 2008b
equivalents/kg) differences on sensory attributes.
2% PP Goat meat Reduction of thiobarbituric acid reactive Devatkal and
substances levels. Naveena, 2010
0.5% PP Goat meat patties Reduction of thiobarbituric acid reactive Devatkal et al., 2010
substances and lightness values. No differences
on sensory attributes.
2% PP Chicken patties Reduction of thiobarbituric acid reactive Devatkal et al., 2011
substances levels.
4% PP Goat meat chunks Improvement of the textural properties Narsaiah et al., 2011
(tenderization).
5% PP Chicken meat balls Reduction of standard plates counts, Chandralekha et al.,
thiobarbituric acid reactive substances, cooking 2012
loss and pH values. Improvement of sensory
attributes.

PP (200 mg Pork meat Reduction of standard plate counts, Qin et al., 2013
gallic acid thiobarbituric acid reactive substances, pH and
equivalents/kg) lightness values. Improvement of overall
acceptability.
1-3% PP Beef sausages Improvement of oxidative stability, microbial El-Nashi et al., 2015
counts, cooking quality and sensory attributes.
0.1% PE Chicken meat Inhibition of Staphylococcus aureus, Bacillus Kanatt et al., 2010
cereus and Pseudomonas sp. growth.
Improvement of oxidative stability.
24.7 mg/g PE Paté Inhibition of Listeria monocytogenes growth. Hayrapetyan et al.,
2012
1% PE Goat meat and Reduction of thiobarbituric acid reactive Devatkal et al., 2014
nuggets substances levels.
0.5% PE Beef and chicken Inhibition of total heterocyclic aromatic amine Keşkekoğlu and
meatballs formation. Üren, 2014
0.5-1.5% PE Mutton ribs Reduction of microbial, yeast and mold counts Dua et al., 2016
and thiobarbituric acid reactive substances
levels. Improvement of sensory attributes.
3% PE Chicken nuggets Reduction of total plate, psychrophilic, yeast Kaur et al., 2015
and mold counts and thiobarbituric acid
reactive substances levels. Improvement of
sensory attributes.
0.5-1% PE Beef meatballs Reduction of lipid and protein oxidation values Turgut et al., 2016;
and improvement of sensory scores. 2017
Apple pomace is a rich source of dietary fiber and pectin, but also
bioactive molecules, such as phenolic compounds (gallic acid, chlorogenic
acid, catechins, quercetin and phloridzin) (Grigoras et al., 2013). As apple
skins are a rich source of quercetin glycosides, several in vivo studies have
been conducted with quercetin to determine its effects on the quality
characteristics of the derived meat products. Quercetin dietary
supplementation in broilers (0.5 or 1 g/kg, from 0 to 42 days) could prolong
meat shelf life by reducing lipid oxidation values (Goliomytis
et al., 2015). At the same time, quercetin incorporation in the diets of broilers
(0.1, 0.2 or 0.3 g/kg) appeared to improve oxidative stability, color, texture
and sensory characteristics of nuggets (Sohaib et al., 2015a) and breast
(Sohaib et al., 2015b) meat. Furthermore, inclusion of quercetin at the level of
200 mg/kg in the broilers’ diet could influence meat fatty acid composition of
the pectoralis major muscle in broilers, by reducing the relative proportion of
monounsaturated fatty acids (MUFA), n-6 polyunsaturated fatty acids (PUFA)
and n-6: n-3 fatty acid ratio (Oskoueian et al., 2013).
2.3. Citrus Pulp
Citrus fruits have a small edible portion and large amounts of waste
material. Dried citrus pulp is the main by-product from the citrus-processing
industry and produced after extraction of the juice from citrus fruits and drying
of the residues. It is a mixture of peel, inside portions and culled fruits of the
citrus family (60-65% peels, 30-35% segment pulp and 0-10% seeds) and
represents a rich source of energy, fiber and calcium. The presence of
functional dietary fiber and antioxidants in citrus by-products allow their
application in food processing to obtain healthy products. As shown in Table
1, citrus by-products (lemon albedo and orange fiber powder) have already
been added to meat products (cooked and dry-cured sausages, cooked turkey
meat, mortadella etc.) with positive effects on their quality characteristics,
such as pH, color parameters and lipid oxidation indices (Fernandez-Lopez et
al., 2004; Contini et al., 2014; Viuda-Martos et al., 2010a). Moreover, citrus
cy-products could serve as a potential ingredient to reduce the nitrite level in
meat products (Viuda-Martos et al., 2009a).
Dried citrus pulp is a cheap feed especially during summer when grass is
limited in countries around the Mediterranean and moderates dependence of
livestock (ruminants) on grains that can be consumed by humans. At the same
time, its utilization in animal diets has a positive effect on preventing problems
related with its disposal into the environment and the necessity for costly
waste management programs. As it has been demonstrated, replacing cereals
with dried citrus pulp in concentrated-based diets (24-35%) could improve
meat oxidative (Inserra et al., 2014) and protein (Gravador et al., 2014)
stability and increase intramuscular polyunsaturated fatty acids (PUFAs)
concentration (Lanza et al., 2015) in sheep. In contrast, other researchers
found no effect of dietary citrus pulp, at different levels (30-45%) (Caparra et
al., 2007; Scerra et al., 2001), and orange pulp (10%) (Lanza et al., 2001)
supplementation on feed intake, growth performance and carcass
characteristics in lambs.
Apart from ruminants, dried citrus pulp could also be used in diets for
monogastric animals. In broilers, dietary supplementation with citrus sinensis
peel extract (1.5 or 3.0%) did not influence final weight, hot carcass weight
and carcass yield, although thigh and breast weight were negatively affected at
the level of 3% (Ebrahimi et al., 2013). In an experiment of the same authors,
addition of the same extract at the level of 1.25 g/kg resulted in a significantly
lower abdominal fat content in broilers (Ebrahimi et al., 2014). Furthermore,
incorporation of citrus pulp in the diet at the level of 10% had a negative effect
on body weight, although polyunsaturated fatty acids content in chicken meat
increased (Murao et al., 2008). TBARS values of breast and thigh meat were
also reduced after the addition of Citrus junos by-products fermented with
multistrain probiotics at the level of 5-20 g/kg in the diets of broilers (Ahmed
et al., 2014). Oxidative stability of chicken leg and duck breast meat was also
improved after the dietary supplementation with a dry extract from orange
(Citrus aurantium) peel (Marzoni et al., 2014). In ostrich, citrus pulp inclusion
(20%) in the diet reduced meat cooking loss, increased polyunsaturated fatty
acids content and ω6/ω3 ratio (Lanza et al., 2004).
The inclusion of ensiled citrus pulp at the level of 5-10% in diets for
growing pigs did not affect back-fat thickness, color parameters and the
proportion and composition of saturated and unsaturated fatty acids in pork
meat (Cerisuelo et al., 2010). However, an increase in the levels of the
polyunsaturated fatty acids in the Longissimus lumborum muscle of Nero
Siciliano pigs was observed after their feeding with citrus pulp (1 kg/animal)
and barleycorn (Chiofalo et al., 2007). Replacing 15% of the dietary dry
matter with dried citrus pulp did not have a significant effect on carcass traits,
meat quality or sensory characteristics in pigs (Crosswhite et al., 2013). On the
other hand, Watanabe et al. (2010) found a negative linear effect of dietary
supplementation with citrus pulp (10-30%) on final body weight, carcass yield
and pork meat color characteristics.
Dietary fibers from citrus fruits are highly soluble and have an additional
advantage over fibers from other sources due to the presence of associated
bioactive compounds (i.e., flavonoids). Bioflavonoids (hesperidin, naringin
etc) are mostly found in the pulp, albedo, membranes and the pith of citrus and
usually contain one or more aromatic hydroxyl groups, which actively
scavenge free radicals and are responsible for their intense antioxidant and
anti-inflammatory properties (Bampidis and Robinson, 2006; Marin et al.,
2007). They compose a class of secondary plant phenolics and belong to the
non-volatile compounds of the citrus essential oil, with a concentration
between 1 and 15% (in citrus essential oil). Flavonoids are introduced in the
animal metabolism as glycosides and polymers that are further absorbed in the
small intestine and entered the circulatory system in the form of glucuronide,
sulfate and methylated metabolites which are perceived as xenobiotics by the
body and are rapidly removed from the bloodstream. Although metabolism of
these compounds remains an elusive and controversial issue over the years,
enteric absorption occurs sufficiently to reduce plasma indices of oxidant
status (Walle, 2004; Crozier et al., 2010). In general, participation of
flavonoids and their metabolites in human diet contributes in the prevention of
cardiovascular diseases (coronary heart disease, atherosclerosis) and some
forms of cancer (breast, prostate, pancreas, esophagus, stomach, colon, etc)
(Fisher and Phillips, 2008). They also appear to affect the activity of different
enzymes (kinases, phosphodiesterases, cyclooxygenases, lipoxygenases and
phospholipases) that play an important role in the inflammation occurrence at
a cellular level (Lee and Kim, 2010).
Several studies have been published during the previous decade that
describe the in vivo effects of flavonoids and particularly hesperidin on meat
properties. Although, meat quality characteristics, such as pH, color, water
holding capacity and shear force values were not significantly influenced,
measurement of lipid oxidation values showed that hesperidin inclusion in the
diet positively affected meat antioxidant properties during storage in lambs
(1.5-3 g/kg) (Simitzis et al., 2013) and broilers (1.5-3 g/kg) (Simitzis et al.,
2011) but not in rabbits (1-2 g/kg) (Simitzis et al., 2014). Naringin
incorporation in the diet at the levels of 0.75-1.50 g/kg also improved
oxidative stability in broiler meat (Goliomytis et al., 2015). Dietary hesperidin
supplementation (5, 10, 15 or 20 mg/kg) positively improved the fatty acid and
lipid metabolite profile (Kamboh and Zhu et al., 2013a) and the water holding
capacity and lipid oxidative stability (Kamboh and Zhu, 2013b) of broiler
breast meat in a dose-dependent fashion. Thus, hesperidin could be a feasible
alternative of antioxidant plants/herbs and synthetic feed additives for the

production of healthier animals and meat especially in poultry.
2.4. Grape Pomace or Marc
Grape (Vitis vinifera) pomace or marc is derived as a result of

destemming, crushing and pressing of grape. It consists mainly of peels, stems,
and seeds and represents about 20% of the weight of the grape processed into
wine (Llobera and Canellas, 2007). It contains a wide range of polyphenols
(i.e., anthocyanins in purple and flavan-3-ols in white varieties) that are
considered to possess several biological properties, such as antioxidant, anti-
inflammatory, anti-cancer, antimicrobial, antiviral, cardioprotective,
neuroprotective and hepatoprotective (Egert and Rimbach, 2011; Georgiev et
al., 2014; McCullough et al., 2012). Due to these activities, the application of
grape by-products is an increasing trend in the food industry as alternative to
chemical or synthetic antioxidants to inhibit lipid oxidation during refrigerated
and frozen storage and thus extend raw or processed meat products’ shelf life
(Brannan, 2008; Garrido et al., 2011; Mielnik et al., 2006; Sayago-Ayerdi et
al., 2009a; Selani et al., 2011; Shirahigue et al., 2010). At the same time, grape
seed extract appears to inhibit microbial spoilage of meat products caused by
Listeria monocytogenes, Salmonella typhimurium, Escherichia coli and
Aeromonas hydrophila (Ahn et al., 2007; Gadang et al., 2008; Reddy et al.,
2013; Sivarooban et al., 2007; Theivendran et al., 2006) (Table 1).
Recent research has stressed the importance of by-products from wine
processing as plant materials that could be used in animal production (Brenes
et al., 2016). Dietary supplementation with grape pomace at the levels of 15,
30 or 60 g/kg in broilers increased the antioxidant capacity of breast muscle,
although growth performance and weights of abdominal fat, liver, pancreas
and spleen were not affected (Brenes et al., 2008). Goñi
et al. (2007) reached to the same conclusions after the addition of grape
pomace at the level of 5, 15 and 30 g/kg in the diet of broilers. Moreover,
inclusion of grape pomace at the levels of 5 and 10% in broilers’ diet reduced
the susceptibility of meat to lipid oxidation and increased polyunsaturated fatty
acids content, without negative effects on body weight, feed intake and feed
efficiency (Chamorro et al., 2015). On the other hand, Francesch and Cartana
(2015) found an increased percentage of unsaturated fatty acids in the meat of
broilers fed with a grape seed supplemented diet (5%). Dietary
supplementation with resveratrol - that is found in the skin of grapes - at the
level of 200, 400 or 600 mg/kg appears to improve growth performance and
reduce oxidative stress in heat-stressed black-boned chickens by increasing
serum growth hormone concentrations and modulating the expression of heat
shock genes in organs of the immune system (Liu et al., 2014).
In pigs, dietary inclusion of grape pomace (30 g/kg) fermented by
Saccharomyces boulardii increased marbling score, values of redness (a*) and
yellowness (b*) and the anti-oxidative ability (lower TBARS) of pork meat
(Yan and Kim, 2011). At the same time, addition of flavonoids extracted from
grapes (2g/kg) in the diet of Iberian pig significantly decreased the lipid
oxidation values of the derived meat (Gonzalez and Tejeda, 2007). On the
other hand, O’Grady et al. (2008) found that dietary supplementation of pig
diets with grape seed extract (100 or 300 or 700 mg/kg) did not affect lipid
oxidation rates in longissimus dorsi muscle. Bertol et al., (2017) did not also
find a significant effect of grape pomace inclusion in the pigs’ diets at the
levels of 3-10% on pork oxidative stability. However, addition of resveratrol
in the finishing diets of pigs (0.3 or 0.6 g/kg) appeared as an effective way of
ameliorating pork meat quality, possibly due to the improved muscle fiber
characteristics and antioxidative capacity (increased glutathione peroxidase
activity and decreased malonaldehyde content) (Zhang et al., 2015).
In lambs, utilization of grape pomace up to 10% in diet did not have
negative effects on growth performance (Bahrami et al., 2010). Moreover,
addition of grape seed extract (25 g/kg of dry matter) in lamb diet protected
the derived meat against lipid oxidation (Jeronimo et al., 2012), although meat
fatty acid profile was not modified (Jeronimo et al., 2010). Dietary
supplementation with a red wine extract rich in polyphenols (900 ppm) also
improved meat oxidative stability in lambs (Rivas-Cañedo et al., 2013).
Finally, supplementation (2.5%) with the grape seed extracts 2,4-heptadien-1-
al, 2-ethylphenol, 2-nonenal, 3-hydroxy-2-butanone appeared to cause a
reduced lipid autoxidation rate in lamb meat (Vasta et al., 2010).
2.5. Olive Cake
Olive (Olea europaea) cultivation plays an important economic and social

role in the Mediterranean region, where are located the top 3 world producers
countries; Spain, Italy and Greece. The use of olive oil in the Mediterranean
diet has been associated with a reduced incidence of cardiovascular disease
and cancer due to its phenolic compounds (Kris-Etherton et al., 2002). Olive
oil has already been added to meat products (sausages, salami, mortadella,
beef patties etc.) with positive effects on their quality characteristics

(Muguerza et al., 2001; Reddy et al., 2015; Severini et al., 2003; Lopez-Lopez
et al., 2010).
However, olive oil extraction generates substantial amounts of by-
products (pulp, olive kernels, skin and water) that are potential pollutants;
from each kg of olives, approximately 800 g of olive cake are obtained
(Camposeo et al., 2013; Chouchene et al., 2010). A variety of substances with
proven antioxidant and radical scavenging activity, such as hydroxytyrosol
(3,4-DHPEA), tyrosol (p-HPEA) and their secoiridoid derivatives (dialdheydic
form of decarboxymethyl elenolic acid, 3,4-DHPEA-EDA or p-HPEA-EDA)
as well as verbascoside, is contained in olive processing residues or olive cake
(Amro et al., 2002). As a result, these residues could be applied post mortem
in meat products for improving their oxidative and microbial stability
(Hawashin et al., 2016; Muiño et al., 2017) (Table 2).
Utilization of the derived cake as a part of the livestock diet might
therefore alleviate the environmental pollution caused and minimize the costs
related to waste management and animal feeding, since animals become less
dependent on conventional feeds such as cereal grains. It can be used as fresh,
ensiled, dried or as a component of concentrated pellets and multi-nutrient
feed blocks in animal and especially ruminant diets. Unfortunately, its
composition is not stable and varies according to cultivation conditions,
method of oil extraction and preservation characteristics (Molina-Alcaide and
Yanez-Ruiz, 2008).
The addition of olive cake to the concentrate at the level of 15% had no
significant negative effect on daily gain, carcass weight and dressing
percentage of lambs, although muscle content of fat and proteins decreased
(Mioc et al., 2007). Moreover, carcass characteristics and major cuts were not
affected by inclusion of olive pulp silage (up to 30%) in lamb diet (Taheri et
al., 2013). Lamb carcass weight and quality were also not influenced after the
application of a dry olive cake-based ration (320 g/kg), but reduced palmitic
and increased oleic and stearic acid contents in samples of subcutaneous fat
were found (Vera et al., 2009). On the other hand, the level of polyunsaturated
fatty acids (Mele et al., 2014) and the production of volatile organic
compounds (Gravador et al., 2015) in lamb meat were not influenced
after feeding with a concentrate containing 35% of olive cake.
Table 2. Effects of agro-industrial olive and tomato by-products addition on the quality characteristics
of meat products

Olive cake 2-6% OCP Beef patties Reduction of pH, thiobarbituric acid reactive Hawashin et al., 2016
powder (OCP) substances levels and total plate counts. Improvement
or waste extract of sensory attributes.
(OWE) OWE (100- Lamb meat patties Improvement of oxidative stability and control of color Muíño et al., 2017
400 mg deterioration.
GAE/kg)
Tomato powder 2% PD Beef frankfurters Reduction of pH, nitrite and oxidation level. Eyiler and Oztan,
(PD) or pomace Improvement of consumer acceptability. 2011
(PM) or paste 1.2-1.5% PD Low fat pork Increase of yellowness, redness and water holding Kim et al., 2011
(PS) sausages capacity. Reduction of pH, thiobarbituric acid reactive
substances, cohesiveness and springiness values.
Improvement of the consumer acceptability.
0.4-0.8% PD Pork and beef Reduction of pH. Increase of yellowness and redness. Modzelewska-
meatloaf No effect on cooking loss and oxidative stability. Kapitula, 2012
0.5-1% PD Pork patties Reduction of pH and thiobarbituric acid reactive Kim et al., 2013
substances. Increase of yellowness and redness.
Improvement of sensory attributes.
1.5-3% PD Pork luncheon roll Reduction of pH and hardness. Increase of yellowness, Hayes et al., 2013
redness, thiobarbituric acid reactive substances and
cohesiveness.
2% PD Chicken nuggets Reduction of total plate, psychrophilic, yeast and mold Kaur et al., 2015
counts and thiobarbituric acid reactive substances
levels. Improvement of sensory attributes.

0.6-1.2% PM Fermented sausages Differences in color properties. Increase of hardness Calvo et al., 2008
(salchichón) and reduction of cohesiveness as the levels of PM
increased. No effect on sensory attributes.
1.5-6% PM Beef hamburgers Increase of redness, yellowness and hardness. Garcia et al., 2009
0.30% PM Ηigh-pressure (600- Improvement of oxidative stability by decreasing the Alves et al., 2012
800 MPa) processed formation of secondary oxidation products.
minced chicken meat
1-7% PM Beef ham and Differences in color properties. Improvement of the Savadkoohi et al.,
frankfurters consumer acceptability. 2014
3-9% PM Chicken sausages Reduction of moisture content. No effect on sensory Yadav et al., 2016
attributes.
5-15% PS Beef patties Reduction of pH and thiobarbituric acid reactive Candogan, 2002
substances. Increase of yellowness and redness. No
difference in desirability scores.
12% PS Beef frankfurters Reduction of pH and nitrite levels & increase of Deda et al., 2007
thiobarbituric acid values, lightness and yellowness.
2-10% PS Mortadella Reduction of total protein. Increase of yellowness, Domenech-Asensi
softness and oxidative stability. No effect on sensory et al., 2013
attributes.
10-20% PS Pork frankfurters Increase of yellowness, redness and antioxidant Valenzuela-
properties. Melendres et al.,
2014
Moreover, no effect of olive cake on lamb carcass traits, meat chemical

composition, quality characteristics (color, cooking loss and tenderness) and
sensory attributes (tenderness, fat degree and flavor intensity) was
demonstrated (Foti et al., 2003; Hamdi et al., 2016). Joven et al. (2014)
reached to the same conclusions after the partial replacing of barley by
increasing levels of olive cake in the diet of finishing pigs; no significant
effects on carcass and meat characteristics were observed. In broilers, dietary
addition of olive cake up to 10% did not adversely affect carcass traits and
inner organs weight (Al-Harthi, 2016).
According to recent studies, the inclusion of olive cake in animal diets
could positively influence the quality and antioxidant capacity of the derived
products, such as beef (Branciari et al., 2015), lamb meat (Luciano et al.,
2013), rabbit meat (Dal Bosco et al., 2012) and pork (Doyle et al., 2006). The
above results clearly demonstrate that the inclusion of olive cake into the
livestock diets could be proposed as an advantageous strategy especially in the
Mediterranean areas allowing exploitation of an important agro-industrial by-
product to reduce production costs for livestock feeding while enhancing the
quality of meat products.
2.6. Pomegranate By-Products
Pomegranate (Punica granatum L.) is a fruit originates from Iran and

Northern India that its global production has greatly increased in recent years
due to its properties against cardiovascular disease, diabetes and different
types of cancer (Viuda-Martos et al., 2010b). Pomegranate also possesses
great antioxidant activity that has been attributed to its high polyphenolic
content (punicalagins, punicalins, gallagic acid and ellagic acid) (Cam et al.,
2014). Pomegranate seed pulp is a by-product of the pomegranate juice
industry that contains powerful antioxidants, anti-inflammatory, anti-coccidial,
anthelmintic compounds, vitamin E, sterols, phenols and natural estrogens
(Dkhil, 2013). There is a wide range of possible applications of pomegranate
peel, rind and seed powder or extract in various meat products and several
studies have been already implemented to establish their regular use due to
their antioxidant (Devatkal and Naveena, 2010; Devatkal et al., 2010; 2014;
Qin et al., 2013; Turgut et al., 2016; Vaithiyanathan et al., 2011) and
antimicrobial (Hayrapetyan et al., 2012; Kanatt et al., 2010) properties (Table
1).
Addition of dried pomegranate seed pulp (50, 100 or 150 g/kg in dry
matter basis) linearly increased fat content and decreased shear force, drip
loss, total aerobic bacterial count and lipid oxidation of longissimus lumborum
muscle in kids. At the same time, a linear increase of linoleic (C18:2 n-6),
alpha-linolenic (C18:3 n-3), n-6 and n-3 polyunsaturated fatty acids and a
decrease in the ratio of n-6/n-3 in both muscle and adipose tissues, and a linear
increase in vaccenic acid, conjugated linoleic acid and punicic acid
concentration in subcutaneous and intramuscular fat were observed (Emami et
al., 2015). In lambs, dietary pomegranate silage supplementation improved
quality characteristics and antioxidant potential of meat, as indicated by the
increase in essential fatty acids, linoleic, α-linolenic and trans-10, cis-12 CLA
acids in intramuscular fat and the increase in total phenolic content and
antioxidant activity (Kotsampasi et al., 2014).
In broilers, supplementation of diets with up to 2% pomegranate by-
products improved meat fatty acid profile (reduction of saturated and increase
of mono-unsaturated and n- 3 fatty acids) and reduced lipid oxidation values of
broiler meat (Ahmed et al., 2015). Szymczyk and Szczurek (2016) reached to
the same conclusions after the addition of dietary pomegranate seed oil at the
level 0.5-1.5% in the diets of broilers; the content of PUFAs increased and that
of MUFAs decreased as a result of a substantial deposition of conjugated
linoleic acid isomers into breast lipids. Inclusion of pomegranate peel extract
at the levels of 200-300 mg/kg also improved the antioxidant capacity and
quality indices of broilers breast meat (Saleh et al., 2017).
2.7. Tomato Pomace
Tomato (Solanum lycopersicum) is one of the most important vegetable

crops in the world, with the amount of the related industry wastes to be
estimated at up to 50 thousand tons per year. This by-product of the tomato
canning industry is called “tomato pomace” and represents around 4% of
tomatoes original weight. It contains valuable nutritional compounds, such as
fibers (59.0%), proteins (19.3%), sugars (25.7%), pectins (7.6%), fat (5.9%),
minerals (3.9%) and antioxidants (e.g., lycopene) (values in dry weight basis)
(Del Valle et al., 2006) and it can be therefore used for the improvement of
color, sensory and textural attributes of processed meat products, such as ham,
sausages, hamburgers etc. (Savadkoohi et al., 2014; Viuda-Martos et al., 2014)
(Table 2).
Tomato pomace can also be fed to livestock, fresh, dried or ensiled with or
without additives as an alternative cheap source of energy and other nutrients.
In an experiment conducted in pigs, dietary supplementation with tomato by-
products (3 or 5%) seemed to slightly affect pork meat characteristics
(tenderness) (Chung et al., 2014). Tomato silage can be added up to 30% of
dry matter basis in fermentable liquid diets for growing-finishing pigs, since it
improves growth performance without affecting carcass characteristics
(Aguilera-Soto et al., 2014). In fattening rabbits, the dietary addition of tomato
pomace at the level of 6% resulted in increased polyunsaturated fatty acids’
levels and yellowness (b*) and chroma values (Peiretti et al., 2013). Increase
of total polyunsaturated fatty acids and of unsaturated/saturated fatty acid ratio
was also observed in the meat of quails fed with a dried tomato pulp
supplemented diet at the level of 10%. However, effects on the oxidative
stability of meat were not exerted at this concentration but at 5% (Botsoglou et
al., 2004). Sahin et al. (2008) reached to the same conclusions;
malondialdehyde levels in serum, liver and muscle of quails decreased after
dietary tomato powder supplementation at the levels of 2.5-5%.
Tomato by-products contain a great variety of bioactive components,
principally lycopene, which have been demonstrated to possess antioxidant,
hypolipidemic, and anticarcinogenic activities (Sanchez-Zapata et al., 2014).
Lycopene antioxidant properties include a considerable reactive oxygen
species (ROS) scavenging activity that allows lycopene to prevent lipid
peroxidation and DNA damage. At the same time, lycopene induces enzymes
of the cellular antioxidant defense systems by activating the antioxidant
response element transcription system (Kelkel et al., 2011). Several
epidemiological studies associate lycopene intake with a decreased risk of
cardiovascular diseases and several forms of cancer (Story et al., 2010).
Meat from lambs fed with lycopene supplemented diets (0.05, 0.1 or 0.2
g/kg) tended to be darker and redder as indicated by lower lightness and hue
angle values at 24 h. Muscle fat content and lipid oxidation values decreased
in the lycopene-supplemented groups, while polyunsaturated fatty acids
increased compared to the control group (Jiang et al., 2015a). Inclusion of
lycopene in lambs’ diet (0.2 g/kg) also appeared to improve the antioxidant
status and optimize the plasma lipid profile, by reducing plasma
malondialdehyde level, total cholesterol, total triglycerides and low-density
lipoprotein cholesterol and increasing plasma antioxidant vitamin E level, total
antioxidant capacity and activities of catalase, glutathione peroxidase and
superoxide dismutase (Jiang et al., 2015b).
Dietary supplementation with lycopene (0.75 g/kg) positively influenced

growth performance parameters and meat quality characteristics in broilers
(Englmaierova et al., 2011). Lycopene incorporation in broilers’ diets (12
mg/kg) also resulted in a reduction in the concentration of cholesterol in thigh
muscle (Rozbicka-Wieczorek et al., 2014). However, Pozzo et al., (2013)
found no effect of lycopene inclusion (0.5 g/kg) on broilers’ growth, slaughter
performance or antioxidant enzymes in the breast meat, thigh meat, liver and
kidney.
CONCLUSION
Meat industry is one of the fastest growing animal production subsectors
and the demand for high-quality meat products is rapidly increasing
worldwide. However, a constant increase of livestock production cost is
observed due to the prices of cereal grains and soybean products. Low input
feeding strategies are therefore necessary, based on alternative feeding
resources, such as the agro-industrial by-products. At the same time, inclusion
of these by-products in animal diets minimizes the environmental effects
induced by their disposal due to their high organic load and enables the
sustainability of high-added value ingredients - nutraceuticals inside food
chain.
In response to recent demand for healthier products, meat industry is
seeking natural solutions to improve quality and boost sales. Meat products
through their modified composition and/or processing conditions should
prevent or limit the presence of certain potentially harmful compounds, and/or
increase the possibility of achieving certain desired traits with the subsequent
added benefits to health. Despite the fact that the understanding of
nutraceuticals mode of action is a prerequisite for their regular application in
animal production, our knowledge regarding their activities in animal
organism is still rather limited and there is a strong need for information
regarding their absorption, distribution, metabolism and excretion. The
understanding of these mechanisms would be beneficial for commercial
animal production, giving us the opportunity to curb the challenges of quality
losses and to preserve the functionality of meat by adjusting the optimal agro-
industrial by-products’ supplementation for different animal species and
establishing their regular use.
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Chapter 2
LIPID AND PROTEIN OXIDATION IN MEAT
Ana P. A. A. Salim1,2, Fernanda M. Viana1,2 and

Carlos A. Conte-Junior1,2,3,*
1
Universidade Federal Fluminense, Department of Food Technology,
Niterói, RJ, Brazil
2
Universidade Federal do Rio de Janeiro, Chemistry Institute,
Food Science Program, Rio de Janeiro, RJ, Brazil
3
Fundação Oswaldo Cruz, National Institute of Health Quality Control,
Rio de Janeiro, RJ, Brazil
ABSTRACT
Oxidation reactions are the main cause of quality loss and reduction
of acceptance on meat matrix. Oxidative deterioration of meat occurs
mainly on lipids and proteins and impairs many attributes, leading to
discoloration, texture modifications and developing off-flavors.
Therefore, the market value is reduced, causing relevant economic losses.
Lipid oxidation is a chain reaction consisted of initiation, propagation,
and termination, which can be performed by enzymatic and non-
enzymatic processes. Moreover, protein oxidation can be defined as a
covalent modification, induced by direct oxidation of the amino acid side
chains or conjugation with secondary compounds that are derived mainly
from the oxidation of polyunsaturated fatty acids. The balance of these
reactions depends on the action of prooxidants and antioxidants agents.
Nonetheless, although lipid and protein oxidation appear to be related
processes, the order of occurrence of these events is not well understood.
In order to reduce or prevent oxidative reactions, many strategies have
44 Ana P. A. A. Salim, Fernanda M. Viana, and Carlos A. Conte-Junior
been applied during pre and post-harvest periods. Furthermore, both

oxidation processes are influenced by intrinsic and extrinsic factors, such
as animal breed and species, muscle types and composition, as well as
animal management, storage temperature and oxygen exposure.
Analytical determination of oxidation on meat matrix is performed
mainly by the measurement of secondary compounds and chemical
groups generated during oxidation processes. Spectrophotometric and
chromatographic methods are the most commonly applied tools to
perform these analyses. Understanding and monitoring the oxidation
processes is essential for the development of technological strategies
aiming the quality improvement of meat industry.
Keywords: meat processing, oxidative stability, lipid oxidation, protein

oxidation
1. INTRODUCTION
Oxidation processes are constituted of complexes chain-reactions and their
adverse effects lead to quality loss in meat. Furthermore, the decrease on
consumers acceptance causes reduction of market values, negatively impacting
the economy [1, 2]. There are several traits associated to the determination of
meat quality, nonetheless the most critical to define purchase decisions are
related to sensorial attributes such as color, texture and flavor. Lipids and
proteins are important meat components and are directly associated with the
maintenance of the mentioned parameters [3–5]. The oxidative stability of
these compounds is highly influenced by their quantitative composition and
chemical structure, which varies according to many intrinsic and extrinsic
factors such as the animal species, breed, sex, age and feeding type [6–9].
Lipids are distributed in the intra and extracellular spaces of meat as a
variety of classes including diacylglyerols, monoacylglycerols, triacylglycerols
and phospholipids. These classes are basically constituted by the esterification
of fatty acids with glycerol, and differ on the number of ester bonds and on the
presence of other compounds (e.g., phosphoric acid and organic bases) [10,
11]. Fatty acids can be classified as saturated (SFA), monounsaturated
(MUFA) or polyunsaturated (PUFA) and are essential to the formation of most
lipids, being responsible for their physicochemical and technological
characteristics [12]. In this context, MUFA and PUFA are more susceptible to
oxidation process due to the presence of double bonds with lower dissociation
energies [13, 14].
Lipid and Protein Oxidation in Meat 45
Proteins are the major organic components on meat and can be

divided into three groups: sarcoplasmic, myofibrillar and stromal. They are
synthesized from amino acids sequences linked by peptide bonds. The
proportion and arrangement of these amino acids varies according to the
proteins functionality, also the exposure to oxidation relies on their position
inside the chain [12, 15]. Additionally, the presence of some amino acids (e.g.,
phenylalanine, tryptophan, histidine, cysteine, tyrosine, proline, methionine,
arginine and lysine) increases the susceptibly to oxidative reactions due to
their facility of convert to carbonyl compounds [16].
Because of the great vulnerability of meat to oxidative processes, the
employment of combined analytical methodologies is necessary to monitor
biochemical and sensorial modifications during the whole production chain [7,
17]. Moreover, the understanding of the.g.e general mechanisms and
consequences of these reactions are important for the development of control
strategies in order to favor the maintenance of meat quality, leading to the
enhancement of this industrial sector.
2. LIPID OXIDATION
Lipid oxidation (LOX) occurs in the presence of oxygen and can be
performed by enzymatic reactions, which are induced mainly by the action of
lipoxygenases that are not predominant in meat matrix; or non-enzymatic
reactions that are triggered by the presence of initiators/catalytic agents (e.g.,
high temperature, light and metals). The most common oxidative mechanisms
are autoxidation and photooxidation [12, 18]. Autoxidation is defined as a free
radical chain process, initiated by the reaction of fatty acid radicals or reactive
oxygen species (ROS) with the triplet oxygen (common form in the
atmosphere) and requires high activation energy [19, 20]. Otherwise, the
photooxidation involves the light exposure of photosensitizers, which transfer
energy to lipids or oxygen species [21, 22].
2.1. General Mechanisms
In general, the LOX reaction comprises three stages: initiation,

propagation and termination. Initiation starts with the abstraction of a
hydrogen atom from a methylene group of the fatty acids chain, stimulated by
catalytic agents, originating alkyl free radicals and leaving unpaired electrons
at the carbons of the chain [8, 12]. These carbon radicals, when originated
from PUFA, usually stabilize by a double-bond rearrangement, resulting in
conjugated dienes [12]. At the propagation phase, triplet oxygen or ROS
continue to react, now with the alkyl free radical to form a covalent bond,
resulting in lipid peroxyl radicals [23, 24]. Then, these radicals continue the
process by reacting with other unsaturated fatty acids present on the meat, to
form lipid hydroperoxides, which are the main primary products of LOX [25,
26]. These compounds are very unstable and rapidly decompose to a range of
secondary oxidation products such as aldehydes, ketones, alcohols and other
volatile substances [27]. The propagation stage may be repeated several times,
with the accumulated radicals reacting among each other, originating non-
radical compounds [28, 29]. Once there are no hydrogen source or radical
species left, the chain is interrupted at the termination phase [28, 30].
2.1.1. Primary and Secondary Products of Lipid Oxidation

LOX development is followed by the production of compounds that are
classified as primary or secondary products and can be used as oxidation
indicators. The primary compounds include conjugated dienes and
hydroperoxides, which are the most common [20, 31]. These compounds are
not responsible for the sensorial changes on meat, however they can act as
substrate for several enzymatic reactions and generally undergo further
oxidation, originating the LOX secondary products [12, 27].
There is a wide variety of secondary products and their type depends on
many factors like the initial reaction conditions and the matrix fatty acids
composition. Specific examples of these comprise malondialdehyde, propanal
and hexanal, some of which are responsible for the characteristic rancid off-
flavor [32]. Moreover, the secondary products of LOX can react with other
compounds of the meat matrix and stimulate their oxidation. One of the most
important interactions is with the proteins, where the reaction with amino acids
containing reactive side chains leads to the formation of carbonyl groups [1,
33]. Therefore, POX and LOX processes appear to be related, even though the
order of occurrence of these events was not established yet.
2.2. Methods to Determine Lipid Oxidation
There are many methods available to determine the oxidation levels on

meat and most of them are based on the different compounds that are formed
during the LOX process (Table 1). The choice of the analytical method should
be based on the objective of the research and on the sensorial characteristics of
the matrix, since each phase of LOX presents characteristic biomarkers [19].
Furthermore, the combination of different techniques must be supported in
order to obtain an overview regarding the meat oxidative status.
2.2.1. Determination of Primary Products of LOX
2.2.1.1. Hydroperoxides
The hydroperoxides are the main primary products and they can be used
as biomarkers for the initial stages of LOX, in which the sensorial
modifications at the meat are not observed yet. However, their determination
can induce an underestimation of the oxidative degradation of the evaluated
sample, once these compounds are unstable and rapidly decomposed on
secondary products [18, 32].
The majority of the analyses utilized to quantify the hydroperoxides are
supported on their capacity of oxidize various reagents like iodide or ferrous
iron. These methods require the prior extraction of the lipids and, in some
cases, the addition of other substances (e.g., xylenol orange) that will form
complexes with the oxidized reagents, so they can be measured by
spectrophotometric methods [19].
Recently, chromatographic techniques such as high-performance liquid
chromatography (HPLC) and gas chromatography (GC) are also being
employed to hydroperoxides determination. Despite the greater sensibility of
these methods, they present higher complexity and usually require more time
and investment. Then, the previously described techniques are preferred due to
their simplicity [27].
2.2.1.2. Conjugated Dienes

The conjugated dienes are relatively stable compounds formed at the
PUFA chains during the early phases of LOX. These products are absorbed in
the UV range and can be measured by spectrophotometry assays, nonetheless a
disadvantage of method is the possibility of suffer interference from
components on the meat matrix (e.g., heme proteins), which absorb at the
same region. Therefore, the use of this technique is limited by the potential
overestimation of the results [34, 3].
Table 1. Analytical methods of determination of

lipid oxidation in meat
Matrix Lipid oxidation Analytical method References

biomarker
Beef meat Malondialdehyde Spectrophotometry (TBARS) [41–43]
Hydroperoxides; Spectrophotometry; [44]
Conjugated dienes; Spectrophotometry;
Malondialdehyde Spectrophotometry (TBARS)
Volatile compounds GC-MS [45]
Poultry Hydroperoxides; Spectrophotometry; [46]
meat Malondialdehyde Spectrophotometry (TBARS)
Malondialdehyde Spectrophotometry (TBARS) [47–49]
Poultry Malondialdehyde; Spectrophotometry (TBARS), [50, 51]

meat Volatile compounds GC-MS
Malondialdehyde Spectrophotometry (TBARS) [52–54]
Fish meat Hydroperoxides; Spectrophotometry; GC-MS [55, 56]
Volatile compounds
Pork meat Hydroperoxides; Spectrophotometry; [57]
Conjugated dienes; Spectrophotometry; [58]
Lamb meat Malondialdehyde Spectrophotometry (TBARS) [59, 60]
Hydroperoxides; Spectrophotometry; [61]
Conjugated dienes; Spectrophotometry;
2.2.2. Determination of Secondary Products of LOX
2.2.2.1. Malondialdehyde
Malondialdehyde (MDA) is known as the most important secondary
oxidation product due to its ability of interact with amino groups of proteins
[35]. The thiobarbituric acid reactive substances (TBARS) spectrometric assay
is the main method employed for the quantification of this compound in meat
[19, 36]. Although the variations of TBARS technique are abundant, the
general procedure consists of the sample homogenization with an acid
(commonly trichloroacetic acid), followed by the addition of thiobarbituric
acid (TBA), which reacts with MDA, resulting in the formation of a pink-
colored complex that offers a maximum absorbance peak at 532 nm [19, 36,
37].
Some protocols submit the TBA-MDA mixture to high temperatures,

however this might induce the fatty acids oxidation, leading to results
misrepresentation. In addition, TBARS assay may also be impaired by the fact
that TBA is not selective to MDA and reacts with other components (e.g.,
carbohydrates and amino acids) [38, 39]. Regardless of the availability of GC
and HPLC techniques to evaluate MDA concentrations on meat, the same
drawbacks described in the hydroperoxides chromatographic determination are
observed [19].
2.2.2.2. Volatile Compounds: Propanal and Hexanal

The volatile compounds are primarily responsible for the flavor
modifications caused by LOX at the meat matrix. Among these compounds,
propanal and hexanal are the most studied and they originate from the
oxidative degradation of n-3 and n-6 PUFA families, respectively [19, 34].
Many sophisticated analytical methods such as gas chromatography–mass
spectrometry (GC-MS) and high-performance size exclusion chromatography
(HPSEC) are involved at the quantification of these products, however the
wide application of these techniques is impaired by the requirement of
laborious preparation steps [34, 40].
3. PROTEIN OXIDATION
Proteins are important targets for oxidation reactions due their abundance
in meat matrix. The mechanisms of protein oxidation (POX) involve different
pathways, in which the nature of the oxidation products is dependent of the
protein targets and how the reactions are initiated [62]. POX can occur on
amino acid side chains or on the peptide backbone and results in loss of
sulfhydryl groups, formation of intra- and intermolecular cross-links and
production of carbonyl derivatives [21]. Furthermore, the generation of
carbonyl compounds is the most important modification of POX in meat.
3.1. General Mechanisms
In general, POX is initiated by a reactive oxygen species (ROS), such as

the superoxide, hydroperoxyl, peroxyl, and hydroxyl radicals, which are
usually produced from external or internal muscle factors. ROS promotes an
abstraction of a hydrogen atom from the protein leading to a generation of a
carbon-centered radical. From a carbon-centered, can be generated an alkyl

peroxy radical and an alkyl peroxide, through the reaction with iron or another
hydrogen atom. In addition, POX reactions can lead to generation of alkoxy
radical and hydroxyl derivative, by reaction with free peroxyl radical or
through the reaction with iron [1]. Moreover, POX can induce the generation
of intermolecular cross-links, by the reaction between two carbon-centered
radicals in absence of oxygen [21].
3.1.1. Generation of Protein Carbonyl Derivatives

Carbonylation is the most common modification promoted by POX
reactions. The carbonyls are yielded through an irreversible and non-
enzymatic process of oxidation of the amino acids (lysine, arginine, proline,
and threonine), glycation with presence of reducing sugars, fragmentation of
peptide backbone and reaction with secondary compounds from lipid
peroxidation (e.g., 4-hydroxy-2-nonenal and malondialdehyde) [63, 64]. The
determination of carbonyl content is widely used to analyze the extension of
oxidative reactions in meat.
3.2. Methods to Determine Protein Oxidation
As in LOX, the extent of POX can be determined by a variety of methods

(Table 2). The choice of the analytical method employed is highly dependent
of the compound to be analyzed, which varies according to the type of the
matrix and the purpose of research. In addition, despite the existence of high
accurate analytical methodologies for the determination of POX, the use of
spectrophotometry is the most consolidated, due the efficiency and
convenience.
3.2.1. Determination of Carbonyl Content

The DNPH method is a quantification technique that allows the
determination of the total amount of carbonyls in a meat matrix. In this
method, the protein carbonyl compounds react with DNPH to generate 2,4-
dinitrophenyl hydrazones, which is read at absorbance of 370 nm. The
carbonyl content is determinate spectrophotometrically and the results are
expressed as nmols per mg of protein [1, 63, 65].
Table 2. Analytical methods of determination of protein oxidation in meat
Matrix Protein oxidation Analytical method References

biomarker
Beef meat Carbonyl Spectrophotometry [41, 72–74]
(DNPH)
α-aminoadipic HPLC-FLD [75, 76]
semialdehyde
α-aminoadipic and γ- HPLC-FLD [77]
glutamic semialdehydes
Poultry meat Carbonyl Spectrophotometry [2, 78, 79]
(DNPH)
Fish meat Carbonyl Spectrophotometry [80–82]
(DNPH)
α -aminoadipic and γ- LC-ESI-MS [83]
glutamic semialdehydes
Pork meat Carbonyl Spectrophotometry [84]
(DNPH)
Carbonyl Spectrophotometry [85]
(DNPH)
Lamb meat Carbonyl; Spectrophotometry
α -aminoadipic and (DNPH); [86]
γ-glutamic HPLC-FLD
semialdehydes
Carbonyl Spectrophotometry [87]
(DNPH)
In meat matrix, the protein carbonyl has been successfully applied as a

marker of POX however, the DNPH method not reflects with accuracy the
extent of POX. Despite the simplicity and convenience of the method some
oxidative modifications in proteins do not lead to generation of carbonyl
compounds and consequently are not detected by DNPH [64, 65].
Additionally, carbonyls can be generated by different pathways, which are not
related to oxidation of amino acid residues, such as the addition of lipid
oxidation products, which could lead to an overestimation of carbonyl content
[66, 67].
For the determination of carbonyls is also used the high-performance
liquid chromatography (HPLC) for the separation of hydrazones, after the
reaction with DNPH. The separation by HPLC demonstrates high accuracy
and selectivity, however the technique requires many steps, which makes the
process expensive and late. In addition, the derivatization process requires a
strong sample acidification, which can promote undesirable changes in meat

sample (e.g., amine decomposition) [68].
3.2.2. Determination of α-Aminoadipic Semialdehydes and

γ-Glutamic Semialdehydes
The α-aminoadipic semialdehydes (AAS) and γ-glutamic semialdehydes
(GGS) are carbonyl products, used as biomarkers of protein oxidation. AAS is
obtained from deamination of lysine, while GGS is formed through the
oxidation of arginine and proline residues [66]. Several techniques can be
performed to determine AAS and GGS in meat matrix, such as gas
chromatography, high-performance liquid chromatography and LC–ESI–MS,
which are presenting highly correlation with the carbonyl contents in meat
samples [69–71].
4. PROOXIDANTS
The beginning of oxidative deterioration in meat lipids and proteins is
influenced by direct or indirect interactions of oxygen species with exogenous
(e.g., high temperature and light) and endogenous (e.g., metals and enzymes)
initiators. These factors act as prooxidants in meat matrix, promoting the
generation of fatty acids/amino acids radicals and ROS [7].
Triplet and singlet are the major forms of oxygen responsible for
oxidation processes in meat. The triplet oxygen is the most abundant ground
atmospheric form. Due to their electronic configuration, the direct reaction
with fatty acids and amino acids is unlikely. Therefore, the formation of free
radicals is required to overcome the electrochemical obstacles and start
oxidation through free radical chain reaction pathway. Otherwise, the singlet
oxygen is an excited state and can react directly with meat components,
without the intermediate production of carbon-based free radicals [20, 22].
The reduction of oxygen during other compounds oxidation leads to
the development of ROS, which comprise radical (e.g., superoxide anion,
hydroxyl, peroxy, alkoxy and hydroperoxy) and non-radical (e.g., hydrogen
peroxide and singlet oxygen) oxygen derivatives. These species can behave as
precursors or initiators on oxidation reactions. One of the most important
precursor at lipid and proteins oxidative processes is the hydrogen peroxide,
which generates hydroxyl radicals, which exhibit high reaction rates. ROS
chemical and quantitative variations will depend on the meat composition and
on the environmental conditions [88, 89].
4.1. High Temperature
High temperature decreases the activation energy of oxidation reactions

and can induce ROS production by different mechanisms. Heating conditions
favor the generation of hydroxyl and alkyl radicals by the decomposition of
hydroperoxides and the break of covalent bonds present on lipids and proteins.
Additionally, the disruption of cells caused by this factor releases oxygen from
meat components such as oxymyoglobin, increasing several ROS production
[90, 91].
4.2. Light
Light exposure can initiate two types of photooxidation reactions at the

presence of photosensitizers. At type I photooxidation, the photosensitizers
(e.g., riboflavin and myoglobin) are electronically excited by light and can
initiate oxidation reactions by the direct interaction with meat compounds,
leading to the formation of several ROS. Type II photooxidation is
characterized by the transfer of energy from the excited photosensitizer to the
triplet oxygen, activating the singlet oxygen. This ROS reacts faster than
triplet oxygen, producing allylic hydroperoxides through the transfer of double
bonds [24, 34].
4.3. Metals
The presence of transition metals such as iron and copper (as a minor
component of muscle) in meat favors the oxidative processes and ROS
production especially through Fenton and Haber-Weiss reactions. The first one
involves the hydroxyl radical generation through the reaction of the hydrogen
peroxide with iron or cupper ions. At the second reaction type, these metal
ions catalyze the reaction between the hydrogen peroxide and the superoxide
anion, also producing hydroxyl radicals. In addition, transition metal ions are
also involved on the formation of alkoxy radicals due to the fast reaction with
hydroperoxides [27, 92].
The major source of iron in meat matrix is the myoglobin (Mb). This
heme protein can catalyze oxidation reactions by the release of iron from the
heme molecule or by the generation of hypervalent species such as
ferrylmyoglobin [93].
4.4. Enzymes
The enzymatic catalysis of oxidative processes can be promoted mainly by

lipoxygenases, cyclooxygenases and peroxidases. These endogenous factors
are involved at the initiation step of oxidation reactions through free radical
mechanisms, leading to the development of specific hydroperoxides.
Moreover, the enzymatic formation of singlet oxygen and hydroxyl radicals
may influences theses reactions. However, the participation of the catalytic
enzymes at the oxidation of lipids and proteins on meat is limited, once they
are easily inactivated by external factors such as heat, during post-harvest
processing and storage [90, 94].
5. ANTIOXIDANTS
Antioxidants are compounds that can hinder or inhibit oxidative processes,
protecting biological systems against the potentially damaging effects of ROS
and several free radicals. After harvest, during the conversion of muscle to
meat, most of the antioxidant substances naturally present at the matrix are
depleted, leading to an imbalance of prooxidative and antioxidative factors and
consequently to the development of oxidation reactions [95]. Therefore, in
order to increase the concentration of antioxidants at the matrix, some dietary
and technological strategies are being successfully employed in meat
production [96, 97].
The antioxidants can be classified, based on their mechanism of action, as
primary, synergistic, oxygen scavengers and chelators. Primary antioxidants
promote the inactivation of the free radicals by donating a hydrogen atom or
accepting an electron from these radicals. The synergistic ones represent
substances with low antioxidant activity if applied alone, but their action is
improved when combined with the primary antioxidants. The oxygen
scavengers remove the oxygen present at the matrix, which becomes
unavailable to initiate or propagate oxidation reactions. The chelators prevent
metal-catalyzed reactions mainly by the formation of insoluble metal
complexes [98, 99].
Additionally, the antioxidants may also be grouped as synthetic or natural,
according to their origin. Synthetic antioxidants are commonly used as
preservatives on meat industry due to their relatively low cost and high
efficacy. Despite being less effective, the application of natural antioxidants is
considered by consumers a safer and healthier procedure to decrease oxidation
reactions in meat. Therefore, although recent studies have shown that the
toxicity of some synthetic antioxidants is low, the current tendency is their
replacement with natural alternatives [7, 99].
5.1. Synthetic Antioxidants
The main synthetic antioxidants used by meat industry are butylated

hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and propyl gallate
(PG). BHA and BHT are monophenolic compounds and they act as oxygen
scavengers and present great synergistic effects when combined with other
synthetic antioxidants such as PG. Regarding these compounds safety, recent
studies affirm that BHA and BHT carcinogenic effects in humans are relevant
only at high doses. However, BHT may cause hemorrhages due to its ability of
reduce vitamin K, which is an important cofactor in blood coagulation. The
acceptable daily intake (ADI) recommended by the joint FAO/WHO Expert
Committee on Food Additives (JECFA) and by the European Food Safety
Authority (EFSA) are, respectively, 0-0.50 mg/kg bw and 0-0.25 mg/kg bw for
BHT; 0–0.3 mg/kg bw and 0-1.0 mg/kg bw for BHA [100–102].
The PG is prepared by esterification of gallic acid with propyl alcohol and
acts as a very efficient peroxyl radical scavenger. During the application of
this substance, the concomitant use of metal chelators is necessary, because
when in contact with iron and copper, the PG forms complexes that can
modify the sensorial characteristics of the matrix. According to EFSA, the
recommended ADI for this antioxidant is 0-0.5 mg/kg bw [103, 104].
5.2. Natural Antioxidants
Herbs, spices and fruits are the major sources of natural antioxidants
compounds such as phenolic substances and vitamins. Some examples of these
plants that are widely applied at meat industry include berries of several
families, rosemary (Rosmarinus officinalis L.) and oregano (Origanum vulgare
L.) The phenolic compounds (e.g., phenolic acids and flavonoids) are
secondary products of plants metabolism and their antioxidant action depends
on the arrangement of hydroxyl groups in the molecules. They can act as
reducing agents, oxygen scavengers and metal chelators. Additionally, some
phenolic compounds may also inhibit prooxidant enzymes and enhance the
activity of antioxidant ones [28, 97].
Among the vitamins present in plants, vitamin E and vitamin C constitute

the most relevant by their high antioxidant capacity. Vitamin E is a lipophilic
antioxidant and act neutralizing free radicals through the donation of a
hydrogen atom from their hydroxyl group, forming non-radical species that
can be reduced to tocopherol dimers or to quinones. Vitamin C is a hydrophilic
antioxidant and eliminates free radicals, forming an ascorbyl radical, which is
stable and causes less oxidative damage. Due to their different solubility
properties, the combined application of vitamin E and vitamin C can present
synergistic effects [102, 105].
6. INFLUENCE OF PRE-HARVEST FACTORS ON

OXIDATION PROCESSES
Oxidation reactions is the main cause of loss of meat quality and are
directly affected by intrinsic or animal factors (e.g., genetic, breed, sex, age
and muscle type) as well as the extrinsic (e.g., diet) factors. Among these, by
means of the dietary strategy is possible to control the muscle oxidation
susceptibility through the manipulation of muscle fatty acid content and
antioxidant intake.
6.1. Intrinsic Factors
6.1.1. Genetic and Breed

Genetic variation includes differences among species, between breeds in
addition to animal-to-animal variation. Breed is mainly influence by the major
genes, as the stress sensitivity gene in pigs and double muscle genes in cattle,
which have impact on meat color, texture, and oxidative susceptibility [106,
107].
Genetic variation includes differences among species, between breeds in
addition to animal-to-animal variation. Breed is mainly influence by the major
genes, as the stress sensitivity gene in pigs and double muscle genes in cattle,
which have impact on meat color, texture, and oxidative susceptibility [107].
Regarding the RN gene, their effects are related to a high content of muscle
glycogen and extended pH decline, which increase drip loss and reduced the
meat yield. In cattle, the double muscle gene is referred as a mutation on the
coding region of myostatin gene, in which increase the number muscle fibers,
as well as increase the fiber sizes [108]. For beef quality traits, these genetic
modifications reduce the fat content and, overall the levels of total SFA and
MUFA in the muscle [108, 109].
6.1.2. Animal Age and Sex

Animal age and sex affect the oxidative stability of meat, by means of
changing the total lipid content and composition of fatty acids during growth
[110]. Age promotes the increase of live weight and carcass weight as well as
the increase the intramuscular fat. In addition, there also an increase of the
total lipids, phospholipid, neutral lipids and oleic acid, however, the content
PUFA reduces [111]. In addition, the muscle fiber composition is also
affected. Animal age, increase the fiber-cross sectional area, which promotes a
decrease of tenderness. Moreover, the muscle fibers present a more oxidative
pattern, with the increase of oxidative fibers and myoglobin content, which
corresponds to color changes in the matrix, such as the increase of redness
[112]. Furthermore, protein oxidation is also increased by age, in which the
antioxidant capacity of muscle reduce and consequently the ability to eliminate
oxidized proteins. Once the accumulation of oxidized proteins occurs, further
oxidative reactions could be initiated [113, 114].
Animal sex mainly affects the fat deposition. Males present thinner
subcutaneous fat than females, which is related to sexual hormones
metabolism. The castrate male carcasses contains more than females and
uncastrated male pigs, which have more lean meat carcass [110, 115].
6.1.3. Muscle Type

The muscle type influences directly the susceptibility to oxidative
reactions. The muscle fibers composition will determine the whole metabolism
of muscle and can be divided in oxidative (red) and glycolytic (white) fibers.
In general, muscles composed mainly by oxidative fibers (e.g., Psoas major)
are more susceptible to oxidative reactions due the greater content of lipids
and myoglobin, whereas glycolytic muscles (e.g., Longissimus dorsi) are more
affected by protein denaturation occasioned by fast decline of pH [41, 83,
116].
In addition, differences in fatty acid composition and heme iron content
also influence the susceptibility of oxidative reactions among species. Beef
contains higher amount of SFA and heme iron than pork and poultry, which
make it the most affected by lipid oxidation. However, regarding the PUFA
poultry presents greater contents than pork and beef [117].
6.2. Extrinsic Factors
6.2.1. Diet
Animal diet affects directly the oxidative stability of meat. The influence
of diet is related to the fatty acid composition and the concentration of
antioxidants intake, which varies according to animal species, supplementation
employed and feed system [118].
Pasture-feed and grain-feed are the mainly feeding systems employed and
present differences on antioxidant content intake (e.g., vitamin E and
carotenoids). Pasture-feed increased concentrations of α-linolenic acid (18:3 n-
3) in meat and conjugated linoleic acid (CLA), whereas grain-diet is rich in
linoleic acid (18:2 n-6). Ruminants, through the consumption of forages, have
a low-fat diet and rich of PUFA. However, in the rumen occurs the microbial
biohydrogenation of PUFA, which leads to the mainly SFA absorption in the
intestine and further deposited in muscle and tissues. Monogastrics (e.g., pigs)
absorbed the dietary fatty acids intact in the intestine and are readily
incorporated to the muscle and fat. Thus, the fatty acid composition of pork
meat can be modified through the addition of adequate lipid source in the feed
[119, 120].
Pasture-feed cattle present lower content of total lipid than the grain-feed.
However, pasture-feed contains more suitable fatty acid composition, with
high levels of myristic (C14:0), palmitic (C16:0) and stearic acid (C18:0). For
beef, the linoleic acid (C18:2), alpha-linolenic acid (C18:3) are the most
important PUFA, and the influence of diet is widely expressed, once pasture-
feed beef present high levels of MUFA, PUFA and n-3 fatty acids when
comparing to grain-feed resulting in resulting in positive ratio (n-6:n-3) [118,
121].
In pigs, the dietary management exerts a more relevant effect on the fatty
composition. The fat supplementation increases the deposition and improves
the modification of fatty acids into muscle and tissue. PUFA are originated
exclusively from the diet, in which the addition of linseed oils in feed increase
in the levels of alpha-linolenic acid (C18:3) as well as the content of the n-3
fatty acid EPA (eicosapentaenoic; C22:5) and DHA (docosahexaenoic;
C22:6). Additionally, feed supplementation with soy, peanut and corn oils
increase the levels of linoleic acid (C18:2), an n-6 fatty acid. Pigs with a high
PUFA diet present less SFA in their muscle and tissues, and more content of
linoleic acid (C18:2) and alpha-linolenic acid (C18:3). In contrast with cattle,
the presence of conjugated linoleic acid (CLA) in pork meat only occurs via
supplementation, which is employed aiming the increase the presence of this
compound in meat. For pork meat the omega ratio (n-6:n-3) vary according to
dietary management, in which feeding grass or with oil supplementation
increase the n-3 PUFA resulting in a positive ratio (n-6:n-3) [110, 115, 122].
Despite the interest of meat industry in increase ratios of PUFA in meat,
enhancing their nutritive values, the meat oxidative stability is affected by
PUFA imbalance, which turns the muscle more susceptible for oxidation
reactions, and thus, promoting discoloration and development of off-flavor.
Vitamin E (a-tocopherol) supplementation is important pre-harvest strategy
used to avoid lipid oxidation. Vitamin E acts inhibiting the peroxidation of
PUFA, induced by free radicals. Dietary levels above 200 mg/kg, protects
meat against lipid oxidation improving oxidative stability [121, 123].
6.2.2. Pre-Slaughter Management

Oxidative stress occurs when a stimulus (e.g., lighting, handling or animal
loading) exceeds the duration or intensity, altering the animal homeostasis.
Pre-harvest management and transportation are the critical stages before
animal slaughter, in which the oxidative stress can occurs, compromising the
animal welfare and meat quality [124, 125]. During pre-slaughter period, the
stress sources can be occasioned by long-term stress (e.g., handling, mixing
animals, animal loading and transport) or short-term stress (e.g., illumination
and conduction to stunner). Both are related with undesirable effects on meat
such as DFD (dark firm and dry) and PSE (pale, soft and exudative) meat.
Long-term stress occasioned DFD meat, through the glycogen depletion and
maintenance of high pH during post mortem period, whereas, PSE meat is
occasioned by short-term stress, and related to fast pH drop combined to high
carcass temperature, leading to a muscle proteolysis [8, 126].
7. INFLUENCE OF POST-HARVEST FACTORS ON

OXIDATION PROCESSES
There are many post-harvest strategies employed by meat industry, aiming
to ensure the meat safety, prolong shelf life and the maintenance of
physicochemical characteristics, which involve since of the temperature
management until the choice of package and application of preservation
methods. Despite the strategies benefits their use can compromise the
oxidative stability of meat. Understanding the mechanism of action of these
methodologies is important for the correct application in each matrix, avoiding

the compromise of meat quality.
7.1. Freezing
Freezing is highly employed for meat matrix, once allows the

conservation for long periods with a few losses of the quality parameters.
Freezing temperatures delay physicochemical reactions, through the reduction
of unfrozen water available, and by the difficult of pro-oxidants to achieve the
susceptible molecules and start the oxidation process [117]. However, even
under freezing temperatures the oxidation reactions still occurs. The formation
of ice crystals promote cellular disruption at the meat matrix, increasing its
susceptibility to oxidative reactions, triggered by natural meat compounds
(e.g., myoglobin and iron). The range of damage promoted by oxidative
reactions is dependent of freeze temperature, which determines the amount of
unfrozen water remains available to the development of biochemical reactions.
The unfrozen water important in terms of oxidation since creates a high
oxidative environment due the concentration of solutes and pro-oxidants [63].
Temperatures equal or less than - 40°C are recommended to freeze meat, in
which about 90% of water can be frozen, retarding the majority of biochemical
reactions. However, in higher temperatures (-12ºC, -18ºC) great part of water
remains unfrozen inside the muscle fibers, allowing LOX and POX reactions
[127]. Among species, fish and poultry meat are the most affected by
oxidation, due their fatty acid profile with high content of PUFA, which
affects important quality traits, such as reduction of water hold capacity
(WHC), discoloration and increase of tenderness [46, 77].
7.2. Cooking
The majority of meat matrixes are subject to cooking before the

consumption. Thermal treatments influence oxidative reactions, once the
temperatures achieved during process increase the free radical production,
besides promoting the denaturation and inactivation of antioxidant enzymes,
allowing the action of pro-oxidants [20, 76]. The extent of oxidative reactions
is determined by the cooking method, temperature and time. High cooking
temperatures and time (e.g., roasting) increase the extent of lipid and protein
oxidation. In addition, some cooking methods (e.g., microwave) promote the
generation of oxidation products, even at low temperature time, decreasing the

content of PUFA in meat. Moreover, when meat is cooked with oils and fats
(e.g., frying), can occur oxidation reactions in both, in addition to the
modification of meat fatty acid composition and antioxidant content, affecting
oxidative stability of meat [128–130]. Regarding the sensory traits, color is the
first parameter affected by cooking due to the denaturation of myoglobin,
providing to the meat a brown coloration. Moreover, WHC and cooking yields
are also affected by cooking. The heat transference promotes the oxidation and
thermal denaturation of proteins, leading to structural changes and affecting
the meat quality parameters [87].
7.3. Irradiation
Gama irradiation is a preservation method widely employed in meat,

which is related to the increase of LOX and POX. Resulting from high-energy
incidence on lipids, the production of free radicals occurs, which are
responsible to accelerate the autoxidation of PUFA. Generally, irradiated meat
(beef, pork, chicken and fish) presents an increase in SFA and a decrease of
PUFA contents. Regarding the irradiation dosage, a maximum of 3 kGy is
established for use in fish and poultry, whereas dosage up to 5.0 kGy can be
used in pork, and a dosage until 7.0 kGy is allowed for beef. Although meat
irradiation with dosage above 10 kGy did not demonstrate deleterious effect
for consumer health, the use of this dose range can promote several
biochemical changes in meat, compromising the sensorial traits [21, 131, 132].
The first parameter affected by irradiation is the color, in which the oxidation
of myoglobin leads to greater meat discoloration [131]. In addition, tenderness
is also affected, due to the change on protein structure, favoring the formation
of aggregates and cross-links, which increase hardness and affects negatively
the meat tenderness [133].
7.4. High hydrostatic pressure
High hydrostatic pressure (HHP) is as emergent technology utilized by

meat industry, aiming the assurance of meat safety and prolonging the shelf
life without the use of additives. However, despite the benefits of HHP, this
technology is related with the increase of oxidative reactions in meat. The
application of HHP favors LOX and POX, since it induces changes on protein
structure, promoting denaturation and formation of aggregates. In addition,

HHP promotes membrane damage and releases iron, which is responsible to
trigger oxidative reactions [134]. Pressure levels between 300-600 MPa are
related to the increase of oxidative reactions in beef, pork, chicken and fish,
whereas pressure levels below 300 MPa demonstrate little generation of
secondary products from LOX and POX [135, 136]. Regarding meat quality
traits, tenderness is the texture parameter mainly affected by HHP, due to
changes on myofibrillar structure. Moreover, HHP promote undesirable color
changes (browning), mainly on read meat, due to the denaturation of
myoglobin, negatively affecting meat appearance [137].
7.5. Comminution
Deboning, mincing, grounding and slicing are practices routinely

employed by meat industry and have impact on LOX and POX. These
practices increase the surface exposed to oxygen and mix the oxidant catalysts
such as iron, turning the meat more susceptible to the attack of ROS [21]. In
order to hinder the effects of comminution on meat, the addition of antioxidant
directly onto meat or by package are being employed [2, 28]. Synthetic or
natural antioxidants can be applied as liquid extract, in ground meat, aiming to
delay the oxidative reactions. Spices and herbs are widely utilized as powder
or liquid extract, demonstrating antioxidant activity. In addition, the use of
grape seed extract is also being used in beef, pork, poultry and fish.
Furthermore, extracts of tea catechin have being shown potential to inhibit
LOX and POX in beef, pork, poultry and fish. Antioxidants can also be
incorporated into packaging (e.g., active packaging; AP), in which they
interact with meat, delaying the oxidative reactions and extending shelf life
[28, 95, 97].
7.6. Packaging
Among the preservation methods, the most used for processing of fresh
meat is modified atmosphere packaging (MAP), which includes vacuum (VP)
and package with gas mixtures. The exposure to oxygen can be favored or
reduced according to the type of package utilized, which directly influences
the susceptibility of meat to oxidative reactions [138]. The VP is mainly
employed for meat cuts. The oxygen impermeability hinders the oxidative
reactions during storage, however promotes surface color changes, from red to
purplish-red. In MAP oxygen (O2), nitrogen (N2) and carbon dioxide (CO2) are
employed, and the mixtures will vary according to the meat matrix. The use of
O2 promotes the maintenance of the bright-cherry red color of fresh meat, due
the oxygenation of myoglobin. In most of retail markets, high oxygen content
(70–80%) in MAP is used for fresh meat. However, the application of high O2
concentrations favors the oxidative reactions, with formation of inter-
molecular cross-links, decreasing tenderness, and increasing drip loss,
affecting the juiciness [139, 140]. N2 is an inert gas, used in MAP to displace
oxygen and to act as filler, avoiding the package collapse. Due to its low
solubility in water and fat, the use of this gas does not promote meat color
changes. CO2 promote the increase of meat shelf life due the capacity of delay
bacterial growth. The absorption of CO2 is dependent of meat moisture and fat
content and, when used in high concentrations, CO2 reduces the meat pH
affecting the WHC and meat color, with increase of drip loss and meat
discoloration [141]. Regarding gas mixtures, for red meat (e.g., beef and pork)
is utilized the concentrations of 20–30% CO2 and 70– 80% O2, aiming the
color maintenance and bacteriological inhibition. For poultry and fish the
oxygen reduction is recommended, using gas mixtures of 40% CO2 30% N2
30% O2, in order to avoid the oxidation of PUFA [142].
8. QUALITY PARAMETERS AFFECTED BY

OXIDATIVE PROCESSES
At the point of sale, meat quality is related with intrinsic attributes that are
important for meat acceptance and consumption. These parameters include the
nutritional value, color, texture and water holding capacity, which are widely
affected by oxidative reactions. Thus, the comprehension of muscle
biochemistry and the mechanisms behind the oxidative stability of meat, such
as the factors that decrease meat quality is important in order to delay the
oxidative process and maintain the quality parameters.
8.1. Color
Color is the first quality parameter observed at the moment of purchase

and is determined for myoglobin. Consumers desire a cherry-red color and use
this parameter as an indicator of freshness and wholesomeness. The changes

occasioned by oxidation promote meat discoloration, leading to product
rejection and economic losses [123, 143]. In fresh meat, Mb can exist in four
different forms, according their redox state: deoxymyoglobin (purplish-red
color; DMb), oxymyoglobin (bright-cherry red color; OMb),
carboxymyoglobin (bright-cherry red color; CMb) and metmyoglobin (brown
color; MMb). POX and LOX promote the Mb oxidation, in which DMb and
OMb are oxidized to MMb, leading to meat discoloration. In addition to Mb
redox states, the Mb content is associated to greater rates of meat
discoloration. Beef present more Mb content than pork and chicken, being the
more susceptible to discoloration. In fish, differences in myoglobin content
will occur according to the color of fish meat (light or dark muscle). However,
in tuna, occurs a higher Mb oxidation rates in comparison to other species,
regardless of Mb content. Tuna Mb presents, on their primary structure,
oxidizable residues (e.g., cysteine residue), which promotes the high rates of
Mb oxidation and meat discoloration, in comparison to beef, pork and chicken
[4, 144].
8.2. Texture
Meat texture is determinate by muscle composition (e.g., muscle fiber

type, connective tissue and collagen content) and is influenced by oxidative
reactions. Among texture parameters, tenderness and hardness are the main
affected by POX and LOX, being widely important for consumer acceptance.
The increase of hardness is undesirable, once promotes meat toughening,
negatively affecting the texture traits. Under oxidative conditions, texture is
affected by inactivation of proteolytic enzymes, responsible for meat
tenderization, along with the generation of protein cross-links. In this context,
fish and chicken meat are the most affected by the changes in tenderness,
caused by oxidative reactions, followed by pork and beef [145–148].
8.3. Water Holding Capacity (WHC)
The capacity to retain moisture is an important quality parameter of meat.

Oxidative reactions induce protein structural changes, altering their
functionality, as the water-holding capacity (WHC), promoting drip loss and
decreasing the meat juiciness. The drip results in economic losses for meat
industry due to reduction of cut yield, weight loss and consumer rejection
[149]. The reduction of WHC is related to the formation of protein cross-links,
which increases the spaces between muscle fibers and consequent allows the
water release. In addition, changes in inter and intramolecular interactions
result in protein denaturation with loss of conformation, contributing to
increase of protein hydrophobicity and reduction of WHC in meat. Among
species, fish and pork are more susceptible to oxidative impact in juiciness, in
comparison with chicken and beef, which are related, besides to oxidative
reactions, to genetic factors (e.g., halothane gene) and muscle composition [1,
140, 148].
CONCLUSION
In this chapter, become evident that the oxidative stability of meat is
directly related to their physicochemical and technological characteristics,
which vary according to the type of matrix, and is related to the intrinsic and
extrinsic factors as well as the processing steps in which the meat is subjected.
Moreover, despite the existence of many methodologies for the determination
of oxidation products, the choice of method is widely dependent on the meat
matrix type, the proposal of the study and also on the target analytes.
Furthermore, the comprehension of meat biochemical and the technological
factors that affects their oxidative stability is widely important to establish
adequate control strategies specifically for each matrix and for their processing
stage.
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BIOGRAPHICAL SKETCHES
Ana Paula Amaral de Alcântara Salim
Affiliation: Universidade Federal Fluminense and Universidade Federal

do Rio de Janeiro
Education: Degree of Medicine Veterinarian (D.V.M.); Master of

Science (M.Sc.); Philosophic Doctor Candidate (Ph.D.Candidate)
Address: Rua Vital Brazil Filho, n. 64. Santa Rosa, Niterói, Rio de
Janeiro, Brazil. CEP: 24.230-340
Professional Appointments: Researcher of Food Technology of Faculty

of Veterinarian of Universidade Federal Fluminense and Ph.D. Candidate of
Food Science of Chemistry Institute of Universidade Federal do Rio de Janeiro
Publications Last 3 Years:
Viana, F. M.; Canto, A. C. V. C. S.; Costa-Lima, B. R. C.; Salim, A. P. A.

A.; Conte-Junior, C. A. Color stability and lipid oxidation of broiler
breast meat from animals raised on organic versus non-organic
production systems. Poultry Science, v. pew, 1 - 7, 2016.
Canto, Anna C. V. C. S.; Costa-Lima, Bruno R. C.; Suman, S. P.;
Monteiro, M. L. G.; Viana, F. M.; Salim, Ana Paula A. A.; Nair, M.
N.; Silva, T. J. P.; Conte-Junior, C. A. Color attributes and oxidative
stability of longissimus lumborum and psoas major muscles from
Nellore bulls. Meat Science, v.121, p.19 - 26, 2016.
Canto, Anna C. V. C. S.; Costa-Lima, B. R. C.; Suman, S. P.; Monteiro, M.
L. G.; Marsico, E. T.; Conte-Junior, C. A.; Franco, R. M.; Salim, Ana
Paula A. A.; Torrezan, R.; Silva, T. J. P. Fatty acid profile and
bacteriological quality of caiman meat subjected to high hydrostatic
pressure. Lebensmittel-Wissenschaft + Technologie/Food Science +
Technology, v.63, p.872 - 877, 2015.
Simoes, J. S.; Mársico, E. T.; Lázaro, C. A.; Ferreira, M. S.; Franco, R. M.;
Pereira, Ana Paula A. A. S.; Conte-Junior, C. A. Microbiological,
physical and chemical characteristics of freshwater prawns
(Macrobrachium rosenbergii) in modified-atmosphere packaging.

International Journal of Food Science & Technology, v.50, p.128 -
135, 2015.
Fernanda Medeiros Viana
Name: Fernanda Medeiros Viana

do Rio de Janeiro
Education: Degree of Medicine Veterinarian (D.V.M.); Master of

Science (M.Sc.); Philosophic Doctor Candidate (Ph.D. Candidate)
Professional Appointments: Researcher of Food Technology of Faculty

of Veterinary Medicine of Universidade Federal Fluminense and Ph.D.
Candidate of Food Science of Chemistry Institute of Universidade Federal do
Rio de Janeiro
Viana, F. M.; Canto, A. C. V. C. S.; Costa-Lima, B. R. C.; Salim, A. P. A.

A.; Conte-Junior, C. A. Color stability and lipid oxidation of broiler
breast meat from animals raised on organic versus non-organic
production systems. Poultry Science, v. pew, 1 - 7, 2016.
Canto, Anna C. V. C. S.; Costa-Lima, Bruno R. C.; Suman, S. P.;
Monteiro, M. L. G.; Viana, F. M.; Salim, Ana Paula A. A.; Nair, M.
N.; Silva, T. J. P.; Conte-Junior, C. A. Color attributes and oxidative
stability of longissimus lumborum and psoas major muscles from
Nellore bulls. Meat Science, v.121, p.19 - 26, 2016.
Carlos Adam Conte-Junior

do Rio de Janeiro
Education: Degree of Medicine Veterinarian (D.V.M.) of Universidade
Federal Fluminense; Master of Science (M.Sc.) in Food Science in the
Universidade Federal do Rio de Janeiro; Philosophy Doctor (Ph.D.) in
Medicine Veterinary in the Universidade Federal Fluminense and Nutrition
and Food Science by the Universidad Complutense de Madrid (Madrid,
Spain).
Research and Professional Experience:
2014-2015 Post-doctorate of Gemone Center, University of California,

Davis.
2014-2015 Visiting professor of Department of Food Science and
Technology, University of California, Davis
2008-2011 Researcher, Department of Biochemistry, Rio de Janeiro
Federal University, Brazil.
2008 Visiting Scientist (7 months) in Karolinska Institutet,
working with Characterization/identification of novel fatty
acids secreted by probiotic as inducers of FIAF expression in
HT-29 cells. Stockholm, Sweden.
2007 Visiting Scientist (5 months) in Medish Centrum at the Vrije
Universiteit Amsterdam working with analysis of folic acid
produced by probiotic bacteria. Amsterdam, Netherlands.
2007 Visiting Scientist (2 months) in Dipartimento di Scienze e
Tecnologie Veterinarie per la Sicurezza Alimentar at the
Universita Degli Studi di Milano working with
chromatographic techniques in food analysis. Milan, Italy.
Professional Appointments:
Professor of Department of Food Technology, Universidade Federal

Fluminense (UFF)
Professor of Food Science Program, Universidade Federal do Rio de

Janeiro (UFRJ)
Scientist of Research Foundation of the State of Rio de Janeiro (FAPERJ)
Researcher of National Council of Technological and Scientific
Development (CNPq)
Honors:
2014 Scientist Award, Rio de Janeiro Research Supporting Foundation.

2013 Researcher Award of National Council of Technological and
Scientific Development.
2012 Young Scientist Award, Rio de Janeiro Research Supporting
Foundation.
2011 First Place Award, Oral Presentation, 21th Seminary Vasconcelos
Torres, awarded by the National Council for Scientific and
Technological Development.
2011 Outstanding Paper Presentation Award, 34th Annual Meeting of
Brazilian Chemical Society, awarded by the Brazilian Chemical
Society.
2010 Outstanding Paper Presentation Award, 7th Brazilian Congress of
Food Microbiology, awarded by the Brazilian Society of
Microbiology.
Rodrigues, B. L.; Alvares, T. S.; Sampaio, G. S. L.; C, C. C.; Araujo, J. V. A.;

Franco, R. M.; Mano, S. B.; Conte-Junior, C. A. Influence of vacuum and
modified atmosphere packaging in combination with UV-C radiation on
the shelf life of rainbow trout (Oncorhynchus mykiss) fillets. Food
Control, 60, 596-605, 2016.
Guimaraes, C. F.; Marsico, E. T.; Lazaro, C. A.; Assis, M. T. Q. M.;
Guimaraes, A. M. P.; Hofmeister, A. W.; Mano, S. B.; Conte-Junior, C.
A.. Effect of the anatomical point of hanging and dripping time on water
retention of chicken carcasses. The Journal of Applied Poultry Research,
v. 25, p. 80-84, 2016.
Guedes-Oliveira, J. M.; Salgado, R. L.; Costa-Lima, B. R. C.; Guedes-
Oliveira, J.; Conte-Junior, C. A. Washed cashew apple fiber (Anacardium
occidentale L.) as fat replacer in chicken patties. Lebensmittel-
Wissenschaft + Technologie/Food Science + Technology, 71, 268-273,

2016.
Bottino, F. O.; Rodrigues, B. L.; Ribeiro, J. D. N.; Lazaro, C. A.; Conte Junior,
C. A. Influence of UV-C Radiation on Shelf Life of Vacuum Package
Tambacu (Colossoma macropomum -×- Piaractus mesopotamicus) Fillets.
Journal of Food Processing and Preservation, 1-11, 2016.
Canto, A. C. V. C. S; Costa-Lima, B. R. C; Suman, S. P.; Monteiro, M. L. G.;
Viana, F. M; Salim, A. P. A. A.; Nair, M. N.; Silva, T. J. P.; Conte-Junior,
C. A. Color attributes and oxidative stability of longissimus lumborum and
psoas major muscles from Nellore bulls. Meat Science, 121, 19-26, 2016.
Guimarães, C. F. M.; Mársico, E. T.; Monteiro, M. L. G.; Lemos, M.; Mano,
S. B.; Conte Junior, C. A. The chemical quality of frozen Vietnamese
fillets. Food Science & Nutrition, v. 4, p. 398-408, 2016.
Viana, F. M.; Canto, A. C. V. C. S.; Costa-Lima, B. R. C.; Salim, A. P. A. A.;
Conte-Junior, C. A. Color stability and lipid oxidation of broiler breast
meat from animals raised on organic versus non-organic production
systems. Poultry Science (Print), v. 95, p. pew331, 2016.
Carneiro, C. S.; Mársico, E. T.; Ribeiro, R. O. R.; Conte-Júnior, C. A.; Mano,
S. B.; Augusto, C. J. C.; Oliveira De Jesus, E. F. Low-Field Nuclear
Magnetic Resonance (LF NMR 1H) to assess the mobility of water during
storage of salted fish (Sardinella brasiliensis). Journal of Food
Engineering, 169, 321-325, 2016.
Lanzarin, M.; Ritter, D. O.; Novaes, S. F.; Monteiro, M. L. G.; Filho, E. S.
Almeida; Mársico, E. T.; Franco, R. M.; Conte, C. A.; Freitas, M. Q.
Quality Index Method (QIM) for ice stored gutted Amazonian Pintado
(Pseudoplatystoma fasciatum × Leiarius marmoratus) and estimation of
shelf life. Lebensmittel-Wissenschaft + Technologie/ Food Science +
Technology, v. 65, p. 363-370, 2016.
Gomes, V. S.; Mano, S. B.; Freitas, M. Q.; Conte Junior, C. A.; Santos, E. B.
Meat characteristics of cattle fed diets containing whole cottonseed.
Arquivo Brasileiro de Medicina Veterinária e Zootecnia, v. 68, p. 1069-
1076, 2016.
Ritter, D. O.; Lanzarin, M.; Novaes, S. F.; Monteiro, M. L. G.; Almeida Filho,
E. S.; Mársico, E. T.; Franco, R. M.; Conte-Junior, C.A.; Freitas, M. Q.
Quality Index Method (QIM) for gutted ice-stored hybrid tambatinga
(Colossoma macropomum×Piaractus brachypomum) and study of shelf
life. Lebensmittel-Wissenschaft + Technologie/Food Science +

Technology, v. 67, p. 55-61, 2016.
Afonso, A. M.; Almeida, P. C.; Bravo, S. A. C.; Araujo, J. V. A.; Marsico, E.
T.; Conte Junior, C. A.; Freitas, M. Q.; Mano, S. B. Metodologia de abate
de girinos de rã-touro para obtenção de filés de cauda e subprodutos não
comestíveis. Revista Brasileira de Ciência Veterinária (Impresso), v. 23,
p. 104-108, 2016.
Bessa, D. P.; Teixeira, C. E.; Franco, R. M.; Freitas, M. Q.; Monteiro, M. L.
G.; Conte Junior, C. A.; Gaze, L. V.; Siva, F. A.; Mársico, E. T.
Functional sausage made from mechanically separated tilapia meat. Italian
Journal of Food Sciences, v. 28, p. 426-439, 2016.
Tigre Maia, L. R. F.; Mano, S. B.; Mársico, E. T.; Conte Júnior, C. A.;
Guimarães, C. F. M.; Brum, D.V. A new sample collection method for the
assessment of the percentage of water absorbed in frozen and quick-frozen
poultry cuts (chicken breasts with skin). Analytical Methods (Print), v. 8,
p. 7080-7086, 2016.
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M. L. G.; Conte-Junior, C. A.; Freitas, M. Q.; Cruz, A. G.; Santos, E. B.;
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restructured caiman steaks containing microbial transglutaminase and salt
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R. S.; Conte-Junior, C. A. Flours and Instant Soup from Tilapia Wastes as
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Marsico, E. T.; Conte, C. A.; Mano, S. B. Microbiological quality and
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Santana, A. S.; Conte-Junior, C. A.; Franco, R. M.; Silva, T. J. P. High
hydrostatic pressure minimizes formation of biogenic amines in
refrigerated caiman meat. Meat Science, v. 96, p. 481, 2014.
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Costa-Lima, B. R. C.; Mano, S. B.; Franco, R. M. Effects of ultraviolet
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during refrigerated storage. Poultry Science (Print), v. 93, p. 2304-2313,
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Santos, E. B.; Silva, T. J. P. Instrumental and sensory texture of low-
sodium restructured cooked Caiman steaks. Meat Science, v. 96, p. 453,
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2014.
Chapter 3
DRY-CURED MEATS:
QUALITY, SAFETY
AND NUTRITIONAL ASPECTS
Paulina Kęska* and Joanna Stadnik

Department of Meat Technology and Food Quality,
Faculty of Food Science and Biotechnology,
University of Life Sciences in Lublin, Lublin, Poland
ABSTRACT
Dry-cured meat products, such as dry-cured hams, loin or sausages,
are some of the most representative traditional foods that are produced
and consumed in different places throughout the world. In Mediterranean
countries, such as Italy, Spain, Portugal or France, there is a firmly
established tradition of eating dry-cured meat products, and these
products are well known not only on the local market but also on a global
scale. These meat products have a unique taste, smell and texture that
result from the use of specific formulations and production technology.
Also, the different microflora of lactic acid bacteria that are typical for
that part of the world can differentiate the organoleptic qualities of dry-
cured meat. Despite the various production processes of dry-cured meats,
curing, fermentation and ageing constitute typical stages of each. Meat is
a valuable nutritious food which, if left untreated, will spoil within a few
days. However, certain preservative techniques may extend its stability
*
Corresponding author: email: paulina.keska@up.lublin.pl,
88 Paulina Kęska and Joanna Stadnik
for several days, weeks or months. Curing is a meat preservation method

which uses the preservative effect of salt (in a high concentration) and, to
a lesser extent, sodium nitrite, mainly in the form of curing salt. Then a
period of curing, fermentation and ageing is required for the full
development of the typical sensory and rheological properties of cured
meat products. Today, many people have a growing awareness of the
relation between diet and health aspects. Dry-cured meat products seem
to have a negative image here, mainly because of their high content of
sodium chloride and nitrogen compounds as well as the possibility of
microbial poisoning as a result of non-thermal preservation methods.
Along with the growing concerns regarding consumer health, much
attention is currently being paid to new developments in the production of
dry-cured products which consist of the introduction of modifications in
production technology while maintaining the characteristic flavour,
aroma and consistency of dry-cured meats. Alternative methods for the
production of meat products as compared to curing with nitrite (III) also
include sodium curing with the use of natural sources of nitrate (V) as
vegetable-based ingredients (such as celery juices) with the simultaneous
addition of denitrifying bacteria culture (mainly Staphylococcus) or acid
whey. The use of vacuum conditions and low temperatures during meat
ageing is another solution implemented to extend the shelf life of dry-
cured meat products. The negative image of meat and meat products may
be changed by reports of biologically active peptides isolated from dry-
cured meat products, particularly those with antioxidant (preventing the
negative effects of fat oxidation), antimicrobial (improving the
microbiological quality of products) and antihypertensive (lowering the
blood pressure as a counterweight to the presence of sodium chloride)
properties. This chapter presents the latest achievements in the production
of dry-cured meat products as well as updated scientific and technological
information on the safety, quality and nutritional properties of this group
of meat products.
Keywords: meat, curing, fermentation, bioactive compounds
INTRODUCTION
Dry-cured meat products are known throughout the world as traditional
food products. This tradition follows from the characteristic recipe and
production style that is typical for different countries and areas, with every
region of the world having perfected its recipes for the production of raw
ripening products as related to the differences between various types of dry-
cured meat products. These differences primarily pertain to the organoleptic
Dry-Cured Meats 89
and physicochemical properties resulting from the livestock used for slaughter
(breed of animals, feeding, their weight and age) and the raw material
(depending on the country of origin the type of meat product is traditionally
manufactured by commonly using whole leg pork or beef (mainly in Spain and
Italy), individual pieces of whole muscle (ham or loin) or a combination of
various components (dry fermented sausage), e.g., m. longissimus dorsi, rectus
femoris, vastus lateralis, medialis and intermedius. The properties of dry-
cured meat products may also differ depending on the differences in the
technological practice used by different manufacturers. Among the many
traditional recipes, the production technology of dry-cured meat products is
mainly based on stages of salting, fermentation and ageing, during which the
raw material undergoes a series of biochemical and physicochemical reactions
that affect the sensory properties and microbiological safety. All of the
technological procedures used during the production of dry-cured products
(such as adding nitrite salt and, optionally, starter cultures) and the conversion
of chemical and physicochemical properties (low pH resulting from the
increase in lactic acid bacteria (LAB), low redox potential and water activity)
are implemented for preservation, colour development and flavour
enhancement. The production of a safe foodstuff with a longer shelf life thanks
to innovative technologies in food production reduces the risk of diseases
associated with the consumption of foods based on meat. However,
consumers’ new lifestyle choices and the growing demand for organic foods
with reduced levels of chemical preservatives have led to the emergence of a
new trend of eating traditional food or food that is based on traditional
processing methods such as fermentation. Although today meat may be
preserved via freezing, refrigeration and heat treatment, traditional fermented
sausages and dry-cured hams, loin or neck are still produced in large
quantities. For this reason, meat and dry-cured meat products are characterised
by high potential for innovation, which makes it possible to meet increasingly
diversified consumer expectations in relation to this group of food products.
Contemporary knowledge in the field of meat technology points to a number
of possible directions in the modification of the composition and nutritional
value of meat products. The production of food with unique health-promoting
qualities that is directed specifically at certain customer groups may be a
chance of development for producers of meat and meat products who are
looking for new groups of consumers for their products.
1. MAIN STEPS IN THE PRODUCTION

OF DRY-CURED MEAT PRODUCTS
Salting/Curing
Salting (this step is also called curing in the literature, when salt is used as
an addition or a combination of salt, nitrates, nitrites and/or sugar are added) is
used as a method of preserving meat by reducing the moisture level. Salting is
accompanied by the osmotic dehydration process – moisture levels fall as the
salt is diffused into the muscle. This allows for better penetration of the salt
inside the tissue and allows water loss. Before the salting step, some elements
such as skin and fat, which are barriers to salt diffusion, are removed to
facilitate the penetration of ingredients (Toldrá, 2008). The salting steps in
dry-cured meat products, e.g., ham, neck or loin, may be performed via two
main techniques (Petrova et al., 2015). In the first technique, which uses an
undetermined supply of salt, a piece of meat is completely covered with salt
(Coutron-Gambotti et al., 1999). In this case the raw meat is placed in
containers with holes at the bottom to permit the drainage of excess moisture.
Sometimes, such as in the case of Spanish dry-cured ham, the relevant stage of
salting is preceded by a pre-salting stage using a mixture of curing ingredients
(salt, nitrite and nitrate) for some period of time, e.g., 30 min. After this the
salting stage takes place during which meat is completely covered with solid
salt and placed in a cold room (e.g., 1-3°C and 90-95% relative humidity for
12 days for Spanish ham) (Escudero et al., 2013). The second method of
salting that is used in the production of dry-cured products is an accurate
supply of salt, which is characteristic of, e.g., Italian dry-cured ham (Virgili et
al., 2007). In this case, a predetermined amount of salt is rubbed onto the
surface of the meat tissue. The salting step allows for low salt addition in order
to achieve a moderate salt content in the final outcome. In this case, the salting
time is determined by the fragment size of the muscle. The next step is the step
of resting, in which the products are often rinsed to remove excess salt and
placed on racks to rest (Coutron-Gambotti et al., 1999). In the literature, this
step is sometimes called post-salting, which is intended to make the salt and
nitrates/nitrites diffuse throughout the entire structure of the muscle tissue. The
length of the resting stage is limited by many factors, including the size of the
piece of raw material, the area ratio of lean mass, pH, the intramuscular fat
content (diffusion barrier salt) and the temperature in the room. The addition
of salt (as sodium chloride (NaCl)) or curing salt affects the chemical and
Dry-Cured Meats 91
biochemical reactions, such as proteolysis, lipolysis and oxidation, which

contribute to texture development and typical taste (due to the release of the
precursors of flavour and odour of the macromolecular constituents of the
meat or salt which directly result from the addition of NaCl) of the meat
products. The addition of NaCl has an impact on the technological (e.g.,
increasing the water-holding capacity and improving the properties of the
gelling and emulsifying properties of the proteins of meat) and microbiological
factors. This is an important feature of NaCl in terms of controlling bacterial
flora which limits the growth of microorganisms that are responsible for
spoiling meat products, particularly in the early stages of production. In
addition to sodium chloride, nitrogen compounds such as nitrate and nitrite are
applied at this stage of production. These curing salts are added as preservative
agents due to their antimicrobial activity as the result of the presence of nitrite,
which is primarily related to the inhibition of growth of Clostridium botulinum
and its toxins (Sebranek & Bacus, 2007; Sebranek et al., 2012). Furthermore,
nitrite plays several other roles in this technology, such as having an effect on
the colour and formation of flavour in cured meat and its contribution to the
oxidative stability of lipids (Marco et al., 2008; Hospital et al., 2015; Sidira
et al., 2016).
Smoking
Smoking is not a general, technological part of preparing the dry-cured

product of a piece of meat, such as ham, loin or neck. It is an optional stage of
production and is more commonly used in the production of dry fermented
sausage. It is carried out for the product’s characteristic taste but, more
importantly, it protects the surface development of mould and yeast due to the
bacteriostatic and bactericidal compounds in smoke (Toldrá, 2008; Asefa
et al., 2009; Petrova et al., 2015).
Fermentation
There is no explicitly designated beginning and ending step of the

fermentation process. It is assumed that from the moment when lactic acid
bacteria (autochthonous or industrial) begin to grow and multiply in the
framework of natural processes, the process of converting carbohydrates into
lactic acid, and thus fermentation, begins and will last until the depletion of
nutrient sources for the microflora. In most industries, fermentation processes

are used with commercial and industrial micro-organisms as a starter culture
comprising LAB predominantly of Lactobacillus and Pediococcus, whose
fermentation properties provide a number of desirable changes in the product
and increase process efficiency, thus leading to consistent quality of the
products. The main advantage of using LAB in meat fermentation is their
ability to decompose carbohydrates to lactic acid which thus lowers the pH or
acidification of meat. In addition, reduced meat pH limits the growth and
development of unwanted microflora, thus increasing the microbiological
purity of dry-cured meat products. The effect of limiting pH refers, among
other things, to spoilage bacteria which are responsible, on the one hand, for
poor microbiological quality of the final products and, on the other hand, for
excessive proteolysis which leads to defects in the texture of the products
(Skwarek & Dolatowski 2013). Furthermore, under conditions of low pH there
is a gelling of meat protein, which enhances the texture of the meat in dry-
cured products. Additionally, the fermentation is amplified by the organoleptic
characteristics of the final product. Generally, the presence of lactic acid gives
a slightly sour taste to meat products. During the fermentation of meat,
compounded processes shape the sensory impressions. Thus, the secretion of
secondary metabolites by the fermentation of LAB, such as diacetyl, improves
the sensory qualities of dry-cured meat. The role of LAB also lies in the
reduction of nitrate V to nitrite III, which is further decomposed to nitrogen
oxide that interacts with myoglobin and creates nitrosomyoglobin. It is this
compound that is responsible for the characteristic red colour of fermented
meat products. An essential role is also played by enzymes derived from LAB
which reduce the amount of peroxides and thus the rancidity of fats or stabilise
the colour of the products (Aquilanti et al., 2007; Prpich et al., 2015; Sidiraet
al., 2016). Also, probiotic products are becoming the fastest growing segment
of the food industry. Although start-fermentation microorganisms, such as
lactic acid bacteria, are commonly used in meat products, commercial use of
probiotic microorganisms in fermented meat products is not yet widespread.
The results of various studies indicate that probiotic strains, single or as a
mixture of strains, such as Lactobacillus paracasei, Lactobacillus rhamnosus
LOCK900 (also called Lactobacillus casei LOCK 0900), Lactobacillus
acidophilus Bauer and Bifidobacterium bifidumBB12, can be used in meat
processing (Kołożyn-Krajewska & Dolatowski 2012; Wójciak et al., 2012;
Okoń & Dolatowski 2014; Liberaet al., 2015; Neffe-Skocińska et al., 2015).
The use of probiotic LAB is a problem for food technologists, mainly due to
the high level of native microflora which effectively competes with other
Dry-Cured Meats 93
microorganisms during processing, thus an assortment of meat products such

as loin, ham, neck or sausages require the development of separate production
procedures.
Drying and Ageing
Stage ageing/drying has two main objectives. The first is to dry the raw
material to ca. 32% weight loss. This effect can be achieved by implementing
appropriate technological parameters. Air velocity, temperature and relative
humidity are critical factors that influence the rate of water diffusion from the
interior of the product and determine the rate of evaporation from the surface,
e.g., in Wójciak et al.,(2012) this effect was reached after 21 days of
production of organic, probiotic fermented sausage (the other parameters were
18ºC and 70%-85% relative humidity). Loss of water (as steam) that occurs
during drying reduces microbial activity due to the loss of water available for
the maintenance of osmotic pressure. In addition, the mass and volume
decrease, the texture hardens and the development of aromatic compounds
takes place (Zukal & Incze, 2010). The hard, dry surface is susceptible to
spoilage microorganisms, and the interior will have the tendency to dry
incompletely. These external factors must be synchronised in order to prevent
excessive dehydration of the meat surface and inadequate water diffusion from
the interior to the surface of the tissue. The steam will leave the surface of the
product only when the water activity in the muscle is higher than the air
humidity around the product. Since water vapour leaves the surface of the
product, humidity increases. The result is that the relative water vapour
content in the environment is higher, thus the rate of diffusion of water from
the product is limited. To continue the process of moisture loss, the ambient
air has to become drier and warmer. This is achieved by environmental control
systems (Zukal & Incze, 2010). Water loss of the raw material is also
important due to protective properties against microorganism. Ensuring
sufficient water availability for the microbial metabolism can determine the
pace of the microbes’ growth. Also the presence of solutes, such as salt or
sugar, to increase their concentration in the drying process makes it difficult
for the bacteria to maintain osmotic pressure. The parameter describing the
availability of water in the product is water activity (aw), defined as the ratio of
steam to medium pure water at the same temperature. Water activity and
microbial growth are closely linked. The water activity of fresh meat is
approx. 0.99, so it quickly breaks down. Microbial growth is inhibited by
water activity of less than 0.75; therefore, manipulation of the activity of water
can affect the growth of microorganisms. Another objective of ageing is to
ensure an adequate time period for the enzymatic and biochemical reactions,
among which proteolysis directly affects the products’ texture by breaking
down the structural proteins of the muscles, primarily myofibrillar proteins.
During the proteolytic breakdown, proteins are released along with peptides
and free amino acids, which form and/or are the precursors of flavour
compounds. In turn, lipolysis is the first step of changing fat into aromatic
components. During ageing, triacylglycerols are hydrolysed, which results in
the formation of free fatty acids. They are then more easily degradable in the
subsequent oxidative metabolism. Oxidation is the next stage of transforming
lipid components responsible for the flavour and aroma of products. Lipid
oxidation is catalysed by factors such as light, elevated temperature and the
presence of salts in the production process (Damodaran et al., 2008). The first
products of oxidative metabolism are peroxides, which can form secondary
oxidation products such as aldehydes, ketones, hydrocarbons, alcohols, esters
or lactones. These are important elements forming the distinct aroma of dry-
cured meat products (Toldra & Flores, 1998; Gandemer, 2002). Thus, the
aging time is dependent on the desired sensory profile.
2. TECHNOLOGICAL CHALLENGES AND NEW SOLUTIONS

Given the changing needs of consumers, a sector of the meat industry
includes new processing technologies and the use of custom component
ingredients that make food more natural and organic. Consumers demand
healthier meat products that have low content levels of chemical additives and
possess bioactive, health-promoting ingredients such as unsaturated fatty acids
or biologically active peptides and amino acids. Overcoming the difficulties in
the production of organic meat products and the elimination of chemical
preservatives are new courses of action for producers (Weiss et al., 2010).
Modification of the Bioactive Compounds Level in Meat

and Meat Products
Measures aimed at modifying the composition and nutritional value of

meat products are aimed primarily at eliminating or reducing the content of
undesirable constituents and enriching meat and meat products in biologically
Dry-Cured Meats 95
active substances. This effect can be achieved via actions taken at the level of
livestock for slaughter as well as via changes in the meat product formulations
(Jiménez-Colmenero, 2007; Olmedilla-Alonso et al., 2013). The results of
research conducted in this area are an indication of the possibility of granting
meat products functional properties, e.g., by modifying the composition of
fatty acids, decreasing the sodium ions or adding functional ingredients, such
as herbs and spices, Lactobacilli or other bioactive components (Higgs, 2000;
Hoffmann et al., 2010; Olmedilla-Alonso et al., 2013; Stadnik & Kęska, 2015;
Bolger et al., 2017). The type of meat, way of breeding and raising livestock
are the main actions necessary to obtain functional products, including dry-
curing products (Fernández-Ginés et al., 2005; Olmedilla-Alonso et al., 2013).
Beef and pork are the most popular sources of raw material for the production
of raw ripening products. Researchers have also pointed to the meat of other
species as nutritionally valuable sources for use in the meat industry. Paleari et
al.,(2003) and Soriano et al.,(2006) proposed horsemeat and the meat of wild
boar, deer and goat for the production of raw ripening products. They
emphasised that the test model products had a reduced fat content which was
characterised by significant values of unsaturated fatty acids. Particularly one
factor in several of the species is the amount of PUFAs, some of which may
play an important role as precursors of anti-thrombotic factors (Simopoulos
1989; Gebauer et al., 2006). High nutritional value was also confirmed by the
high content of free amino acids, in particular amino acids that had an
undisputed nutritional value. Besides meat, the type of farming is essential in
achieving the quality of raw ripening products. There is no doubt that
ecological meat has better nutritional properties than meat from conventional
production. Ecological meat thus has a much better composition of animal
fats; the sensory quality of organic meat as evaluated by consumers is better,
mainly due to the content of intramuscular fat which conditions better flavour
and tenderness of the product. A problem appearing in numerous publications
is greatly accelerated organic lipid peroxidation in meat which results from the
modified fatty acid composition (Wójciak, 2012).
One of the main directions to improve the nutritional and functional
quality is to reduce the fat content in meat. The chemical composition of meat
can be shaped intravitally through the selection, crossbreeding and line feeding
method (modifying the composition of the feed). The proper selection of
breeds for crossing reduces the fat content of the external carcass and the level
of unwanted intramuscular fat while maintaining the optimal intramuscular fat
content. The removal of visible fat during the cutting and punching of essential
parts further reduces the meat’s overall fat content (Jiménez-Colmenero, 2007;
Olmedilla-Alonso et al., 2013). In improving the nutritional value of meat, in

addition to the amount of fat in the diet an important role is also played by its
composition, i.e., the relative proportions of saturated, monounsaturated and
polyunsaturated fatty acids (PUFAs) (Connor, 2000; Fernández-Ginés et al.,
2005). In studying the role of dietary factors in preventing NCDs, the attention
of researchers was primarily focused on the possibility of increasing the share
of dietary polyunsaturated n-3 fatty acids and thus reducing the non-preferred
ratio of n-6 to n-3 (Marciniak-Łukasiak, 2011; Olmedilla-Alonso et al., 2013).
Measures aimed at enriching animal fat in polyunsaturated n-3 fatty acids may
be taken at the stage of cultivation, e.g., by modifying the composition of the
compound feed. For this purpose, feed concentrates rich in linoleic acid (n-6)
are replaced by mixtures containing components which are a source of
polyunsaturated n-3 fatty acids (linseed oil, flax seed, seaweed, fish oil).
Modification of the fatty acid profile via the feed takes place primarily in
monogastric animals, as PUFAs are substantially biohydrogenated to saturated
fatty acids in ruminants (Zymon & Strzetelski, 2010). Increasing the share of
polyunsaturated n-3 fatty acids in the diet of the animals requires the use of
appropriate protection against oxidation processes via the addition of
antioxidant vitamins, e.g., vitamin E, β-carotene or vitamin C. In addition to
loss of oxidation stability, enhancement of PUFA n-3 may also adversely
affect the raw material’s sensory and technological suitability (excessive
softness, mush fat) and negatively affect its stability (Decker & Park, 2010;
Piotrowska et al., 2012).
Technological possibilities limiting the fat and cholesterol content in dry
cured products are focused primarily on cured fermented sausage at the
milling and mixing meat stuffing step. This strategy is one of the most
important approaches to the development of potential meat-based functional
foods. It is impossible to completely eliminate fat from raw maturing sausages
as fat is a carrier of flavour and a precursor of volatile aromatic compounds.
Moreover, the production of fermented sausages with a very low fat content is
problematic due to excessive weight loss during drying of the product, the
sausage’s unacceptable appearance due to its wrinkled surface (insufficient
filling in the sausage casing) and the carbonising step when smoking is used.
Liaros et al.,(2009) proposed the use of vacuum packaging during ripening as
an effective strategy to produce low-fat fermented sausages without adversely
affecting their appearance. Reducing the level of fat in the production of dry
fermented sausage is achieved by mixing lean meat and fat, and limiting the
fat content of the finished product by up to 35% without a reduction in product
acceptability to the consumer is possible. Another possible solution for
Dry-Cured Meats 97
reducing the fat content in dry sausages is the use of suitable fat replacers,
such as plant oils. Ansorena and Astiasarán (2004) concluded that the use of
linseed oil as a partial replacement in the production of pork fat-dry fermented
sausages decreased the n-6 to n-3 ratio (from 14.1 to 2.1), thus resulting in an
increase of the α- linolenic acid increment. To date, fat content has been
replaced by soy oil (Muguerza et al., 2013a), olive oil (Muguerza et al., 2003b;
Bolumar et al., 2015), konjac gel (Ruiz-Capillas et al., 2012), fibres
(Fernández-López et al., 2008; Salazar et al., 2009) and inulin (Mendoza et al.,
2001; Menegas et al., 2013). It is technically easier to incorporate liquid oils in
fresh and cooked meat products than in dry-cured meat products (Bolumar et
al., 2015). However, other strategies to achieve stable incorporation of liquid
oils into dry-cured meat products were also examined, e.g., Turkish fermented
sausages (Sucuk) were prepared by replacing 15, 30 and 50% beef fat with
hazelnut oil included as pre-emulsified with simplesse®100 (whey protein
powder). The results indicate that the replacement of fat beef with pre-
emulsified hazelnut oil by up to 50% in the Sucuk formulation significantly
influenced the nutritional quality without adversely affecting the quality of
ripening. Despite the increase in MUFA and PUFA fractions, no problems
connected with oxidation were detected (Yildiz-Turp & Serdaroglu, 2008).
Virgin olive oil was also included as a pre-emulsified fat with the soy protein
isolate in the production of Turkish sausage. It was found that replacing beef
fat with olive oil had a positive effect on the sensory quality and on the
reduction of cholesterol in Turkish soudjouk (sucuk) sausages (Kayaardi &
Gök, 2004). Another approach to the use of olive oil for the production of dry-
cured fermented sausages was presented by Bolumar et al.,(2015). However,
the incorporation of olive oil is an inconvenient technology and gives an
unacceptable product as the olive oil is not retained within the sausage matrix.
It is also possible to create tri-dimensional networks or gel structures by using
the denaturation ability of HPP treatments. Bolumar et al., (2015) obtained
HPP-meat by treating pork lean meat trimmings with HPP for 5 min at 600
MPa at room temperature in the model products. The cooked meat can be
further incorporated into the final product.
Sodium Chloride Control
Currently, the trend towards a high quality diet with a low sodium content
has promoted the study of meat products manufactured with less sodium
chloride. Previous studies on the reduction of sodium chloride were aimed
primarily at limiting the addition of NaCl to meat products to a minimal level

of acceptable sensory quality and allowing a product with microbiological
criteria (Coutron-Gambotti et al., 1999; Corral et al., 2014). Another approach
has been to replace a portion of the NaCl with other chlorides (KCl, MgCl2,
CaCl2) (Wu et al., 2014; Lorenzo et al., 2015; dos Santos et al., 2015).
Establishing proper partial NaCl (%) replacement with other chlorides is very
important for the effective control of the rate of proteolysis and lipolysis,
thereby controlling the final taste of the product, particularly since potassium
contributes to a bitter taste. Curing salts with up to 40% KCl in dry cured
bacon (Wu et al., 2014) or 50% KCl in Italian salami (Zanardi et al., 2010)
were used without a significant negative impact on their sensory properties.
Replacement of the NaCl part with other non-chloride salts (lactate or
phosphate) in raw ripening products was also the subject of investigation
(Gelabert et al., 2003; Guàrdia et al., 2008). The results indicate that even with
the replacement of NaCl it is possible to achieve an acceptable meat product
without affecting the taste, texture, colour and microbial stability. An
alternative to chemical compounds are methods of improving salt diffusion via
high pressure treatment (HPP) or ultrasound technology in meat (McDonnell
et al., 2014; Ojha et al., 2016). Ferrini et al.,(2012) stated that NaCl reduction
was possible by salt replacement with KCl in combination with HPP.
Nitrate Control
Curing that involves nitrates and nitrites shapes the physicochemical

properties (desired colour, flavour, aroma, etc.), oxidative stability and, most
importantly, inhibits the growth of pathogenic microorganisms, in particular
Clostridium botulinum and Listeria monocytogenes. Regardless of the benefits
arising from the use of sodium nitrate and sodium nitrite, the ability of high
reactivity with second-amines leading to the formation of carcinogenic
nitrosamines has limited their use (Sebranek & Bacus, 2007). Currently, an
increasing number of products are appearing on the market which are referred
to as “non-nitrite, i.e., where there is no declared addition of nitrites in the
production process” (Sebranek et al., 2012). The curing process of such dry-
cured products is to reduce nitrates (V) contained in other additives, e.g., fruit
or vegetables. Magrinyà et al.,(2009) and Magrinyà et al.,(2016) used natural
sources of nitrates (V) in the form of celery concentrate with the simultaneous
addition of cultures of denitrifying bacteria (Staphylococcus carnosus which
possesses nitrate reductase activity) as alternative methods to cure dry-cured
Dry-Cured Meats 99
sausage with nitrate (III) sodium. Another option is the use of acid whey for
the production of maturing raw meat products. Acid whey is a by-product in
cheese-making. It comprises a plurality of metabolites derived from lactic acid
bacteria strains, often probiotic ones possessing bacteriostatic and bactericidal
properties (e.g., acid, lactic acid, and bacteriocins), which support the effect of
meat preservation and ensure sensory acceptability. Due to its “natural”
antioxidant properties, acid whey can be used to stabilise oxidative processes
via its ability to inactivate pro-oxidative heme proteins (ferrylmyoglobin) and
to chelate prooxidant transition metals to β-lactoglobulin and lactoferrin,
respectively (Colbert & Decker, 1991). The possible mechanisms of
antioxidant whey also include free radical scavenging by amino acids such as
cysteine and tyrosine (Tong et al., 2000). To determine the functionality of
substitutes, nitrite in organic sausages, sea salt and AW in conjunction with
mustard were tested (Karwowska et al., 2014a; Karwowska et al., 2014b;
Wójciak et al., 2014, Wójciak et al., 2015). The results indicated that these
components have a positive impact on the physicochemical features of non-
nitrate sausages. Wójciak et al., (2015) also studied the influence of acid whey
and probiotic strains (Lactobacillus casei LOCK 0900, Lactobacillus casei
LOCK 0908 and Lactobacillus paracasei LOCK0919) as a replacement for the
microbial stability and sensory quality of organic fermented sausage. The
microbiological quality of ecological fermented products with probiotic strains
and acid whey was superior as compared to the controls with the addition of
curatives. Thus, acid whey can be used effectively to improve microbiological
quality without adversely affecting organic sausage quality. Stadnik and
Stasiak(2016) studied the effect of acid whey on the physicochemical
properties of non-nitrite organic dry-cured pork loin. The combination of acid
whey and sea salt resulted in an effective lowering of the pH of organic
fermented loin without nitrite. Sea salt in combination with acid whey
effectively reduced the browning reaction involved in the formation of dark
colour in the product. Variants with acid whey were characterised by a similar
PUFA content in comparison with the cured loin, thus the presence of PUFAs
in trials with AW could have a protective effect against oxidation at a level
comparable to nitrite.
Addition of Natural Plant Extract

with Antioxidant/Antimicrobial Properties
The inhibition of undesirable oxidation processes, such as oxidation of

lipids, which occur in dry-cured meat products is small due to the protective
effect of bioactive compounds derived from plants. Some products’ small
compounds act as free radical scavengers, chelating metal ions and quenchers
of singlet oxygen, thereby they delay the oxidative degradation of lipid and
improve the quality and nutritional value of food (Li et al., 2013; Lorenzo et
al., 2013; Pateiro et al., 2015; Jiang, et al., 2016; Wójciak & Dolatowski,
2016). In recent years, many researchers have evaluated antioxidant properties
from different plants and vegetables. Among the many compounds, the
flavonoids and polyphenols that are present in green tea were tested due to
their potent antioxidant and nutraceutical activity. Neffe-Skocińska et
al.,(2015) used a green tea extract and the Lb. rhamnosus strain LOCK900 as
a probiotic strain to improve selected physicochemical properties and sensory
quality in dry-cured pork loin. The results indicated that the application of
green tea extract had no negative effect on the growth and survival of LAB
and on the overall sensory quality. Moreover, the addition of Lb.
rhamnosusLOCK900 and green tea extract inhibited oxidation lipids. Also, the
effects of plant polyphenols (tea polyphenol, grape seed extract and gingerol)
and α-tocopherol on the quality and microbial safety of dry-cured bacons were
studied (Wang et al., 2015). The authors indicated the role of plant extract in
lipid inhibition, reducing aerobic plate counts, Enterobacteriaceae,
Micrococcaceae, yeast and moulds as well as inhibiting the formation of
biogenic amine (putrescine, cadaverine, tyramine and spermine). Also, green
tea polyphenol was most effective in reducing lipid oxidation and lower
residual nitrite and N-nitrosamine levels in dry-cured sausage (Li et al., 2013).
The effect of grape seed and chestnut extract as natural antioxidants (on
physicochemical, lipid oxidation, microbial and sensory characteristics) of
dry-fermented sausage (Lorenzo et al., 2013) or chorizo (Spanish dry-cured
sausage) (Pateiro et al., 2015) was also investigated. The effect of adding
grape seed extract (and Bifidobacterium animalis ssp. lactisBB-12 as a
probiotic strain) on the inhibition of hydrolysis of fats during two-month
storage of dry-cured neck was also confirmed (Libera & Stasiak, 2015). In
turn, Wójciak & Dolatowski(2016) evaluated the addition of acid whey with
plant extracts, i.e., mustard, rosemary leaves and juniper berries, which are a
rich source of natural antioxidants (polyphenols, terpenoids and vitamins), on
the quality and shelf-life of organic pork roast during vacuum storage. The
Dry-Cured Meats 101
authors showed that the joint action of plant extracts in combination with acid
whey reduced oxidation of lipids in a manner similar or identical to that of
using nitrate. Their results also indicated effective antibacterial ingredients that
were used in production. The results of the microbiological assays of samples
with natural antioxidants of plant origin did not differ from the results of the
control samples (containing nitrates) (Wójciak & Dolatowski, 2016). These
results indicate that plant extracts are effective antioxidants and can be used to
produce raw ripening products in order to improve quality and ensure safe
products.
The growth of certain microorganisms, such as mould and yeast, on the
surface of dry-cured meat products affects their organoleptic properties.
Enzymes involved in proteolysis and degradation of amino acids, lipolysis and
oxidation processes release taste and smell precursors. Some forms of
microorganisms also retard rancidity and colour stability through catalase
activity and via hindering the penetration of oxygen and light (Bruna et al.,
2001). However, uncontrolled growth of microorganisms can lead to a loss of
quality characteristics and to product contamination with pathogenic
microflora. As an example, mould, in suitable environmental conditions, can
produce green, grey or yellow discolouration in dry-cured meat products and
toxic and allergenic metabolites can penetrate the meat. As an alternative to
chemical preservatives such as potassium sorbate, there has been growing
interest in natural sources against pathogenic flora, such as essential oils. The
superficial antifungal activity of oregano essential oil in Spanish fermented
dry-cured sausages (“salchichón”) was determined (Martín-Sánchez et al.,
2011). The surface application of oregano essential oils reduced mould
contamination on the surface without significantly affecting the drying
process. Moreover, the addition of oregano essential oils led to a higher
amount of unsaturated fatty acids. Additionally, the essential oils of oregano
had no effect on the sensory properties, which resulted in even better texture
by increasing hardness. El Adab et al.,(2016) tested the effects of adding
essential oils from oregano and thyme on the microbiological, biochemical and
sensory characteristics of Tunisian dry-fermented poultry meat sausage and
confirmed these oils’ antimicrobial activity. Their presence in the product
contributed to a reduction in the number of Enterobacteriaceae, total coliform
counts and Staphylococcus aureus counts, thus raising the hygienic quality of
dry fermented sausages.
The antifungal activity of polyphenols extracted from olive mill
wastewater was examined as an alternative to chemical molecules in
improving food safety (Chaves-López et al., 2015). This treatment of reducing
or eliminating the undesired growth of fungi on the surface of dry fermented

sausages was effective and can be used as a potential alternative to synthetic
antifungal compounds to preserve the product from both oxidation and
undesired fungi without changing its sensory characteristics (Chaves-López et
al., 2015).
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Chapter 4
ANALYTICAL TECHNIQUES FOR TRACE-

ELEMENT DETERMINATION
IN MEAT SAMPLES
Jefferson S. de Gois1,, Eduardo S. Chaves2

and Aderval S. Luna1
1
Department of Analytical Chemistry, Rio de Janeiro State University,
Rio de Janeiro, Brazil
2
Department of Chemical Engineering, Federal University of Technology -
Paraná, Paraná, Brazil
ABSTRACT
Obtaining information about trace-element content in meat samples
is a critical approach to assess meat quality and its potential impact on the
human health. Different techniques might be used to monitor the trace
element concentration in meat samples, such as inductively coupled
plasma mass spectrometry, ion chromatography, capillary
electrophoresis, atomic absorption spectrometry, among others. Most
techniques require at least one sample pretreatment step to solubilize the
analytes from the sample in the aqueous medium before analysis.
Procedures that are available for sample pretreatment of meat samples
include dissolution in alkaline media, microwave assisted digestion using
concentrated acids, combustion techniques, extraction procedures using

Corresponding author: Jefferson S. de Gois. Email:jefferson.gois@uerj.br.
112 Jefferson S. de Gois, Eduardo S. Chaves and Aderval S. Luna
acidic media, and direct solid sample analysis. Therefore, this entry
presents an overview of the main sample preparation methods and
analytical techniques applied for trace-element determination in meat
samples.
Keywords: meat analysis, trace-element in meat, meat quality control
INTRODUCTION
Meat is one of the most consumed foods in the word, the concentration of
trace-elements in meat samples may provide information on its nutritional
content and also about the presence of potentially toxic elements such as Hg
and Pb (Tsuji et al., 2016).
Numerous analytical techniques have been applied to access the elemental
composition of such matrices; in general, these techniques provide different
limits of detection (LOD) and quantification (LOQ) among them, and require
the analyte in a liquid solution before analysis.
Sample preparation can be carried out to release the analyte from the meat
samples to a liquid solution; the proper selection of the sample preparation
procedure will depend on the sample’s matrix, analyte and analyte
concentration (in this case a preconcentration step might be required).
Direct solid sample analysis is also an option for trace-element
determination in meat samples, with the advantage of not requiring the analyte
in an aqueous solution and therefore, time-consuming sample preparation steps
may be avoided.
Thus, the aim of this chapter is to provide a general overview of the
available methods and techniques for trace-element determination addressing
the current discussion from the sample pretreatment to the instrumental
determination.
TRACE-ELEMENTS IN MEAT
The definition of meat may differ over the world due to characteristics that
are taken into account for controlling its composition and the composition of
meat products. According to the definition provided by Codex Alimentarius,
meat is defined as “all the edible parts of any animal slaughtered in an
abattoir,” and meat products are defined as “the products intended for human
Analytical Techniques for Trace-Element Determination … 113
consumption containing meat from mammals, poultry, and game meat”

(FAO/WHO, 1992).
Meat is mainly composed of water, proteins, amino acids, minerals, fats,
vitamins, and small amounts of carbohydrates (FAO, 2017a). Thus, meat and
meat products are important sources of proteins, vitamins, minerals, and some
micronutrients for the human nutrition (FAO, 2017b).
Worldwide meat consumption can be directly related to the culture of the
population and production costs (OECD-FAO, 2017). The average meat
consumption from the year of 2006 to 2016 was reported by the Organisation
for Economic Co-operation and Development (OECD). The data, presented in
Table 1, shows that, in general, the major consumer of meat per capita are the
US, Brazil, countries of European Union followed by Russia and South Africa.
Besides the nutritional benefits related to the consumption of meat and
meat products to human health regarding macro and micronutrients, there is a
growing concern about some potentially toxic trace elements that may be
present in meat. Most of those elements and species present the ability of
bioaccumulation. These potential toxic elements are normally released into
aquatic and terrestrial ecosystems as a result of anthropogenic actions (Alturiqi
et al., 2012).
Cadmium and Pb, for example, are considered toxic elements that have no
biological function; their presence in foods is a result of environmental
contamination by anthropogenic activities and food processing. According to
the World Health Organization, Pb is frequently detected in meat, mainly in
organ meats and wild game (FAO/WHO, 2011).
The maximum level for contaminant elements in foodstuff has been
established by the Commission of the European Communities through the
Commission Regulation (EC) n 1881/2006 of December 19th, 2006. It
establishes that the maximum tolerated concentration of Pb in bovine, sheep,
pig, and poultry meat is 0.10 mg kg-1. Additionally, muscle fish, some
crustaceans, and bivalve mollusks have limits for Pb concentration of 0.30,
0.50 and 1.50 mg kg-1, respectively (European Commission, 2006).
The maximum concentration established for Cd is lower (0.050 mg kg-1)
when compared to Pb, for some crustaceans and bivalve mollusks the
maximum levels of Cd are respectively 0.50 and 1.0 mg kg-1 (European
Commission, 2006). These examples demonstrate that those elements have its
own maximum concentration of tolerance depending on the food source, and it
is mainly considered due to its toxicological impacts and bioaccumulation
potential. Apart from health effects, in some cases, the maximum levels of
trace-elements in foods may be established based on the available
methodologies for their detection and quantification. In addition, permitted

maximum concentrations of these contaminants may vary according to the
regulation adopted by the countries.
Therefore, the existence of reliable high-throughput methods to measure
and monitor the concentration of trace-elements in meat products is of great
importance in food industries. It allows the application of analytical techniques
to meat quality control preventing crossover contamination and assuring food
safety.
The determination of essential and potentially toxic elements in meat
samples and meat products has been carried out by several analytical
techniques. Some examples may be pointed out as the determination of Cd,
Hg, Pb, and Sn in canned meat from Egypt, which was conducted to compare
with established maximum levels and identifying the contamination sources.
According to the authors, significant concentrations of Cd and Pb were found
and linked to contamination from production processes (Khalafalla et al.,
2016). High concentration of Cd and Pb were also found in frozen and canned
pork meat products commercialized at Chennai city in India (Santhu et al.,
2008).
Table 1. Meat consumption in some regions around the world

from 2006-2016
Period 2006-2016 Average consumption in kg per capita

Beef Pork Poultry Sheep
World 6.591 12.220 12.751 1.685
EU 10.892 32.531 21.075 2.057
US 27.100 22.059 46.031 0.385
Brazil 25.173 10.831 37.454 0.400
India 1.071 0.232 1.542 0.483
Thailand 2.066 10.474 9,095 0.035
China 3.447 29.490 11.008 2.697
Russia 12.877 18.073 23.290 1.152
South Africa 11.107 3.763 28.786 3.125
Sub-Saharan Africa 3.308 1.114 2.230 2.129
*
Data collected from The Organisation for Economic Co-operation and Development
(OECD-FAO, 2017).
The concentration of Fe, Mg, Mn, P, and Zn was determined in meat

samples using inductively coupled plasma optical emission spectrometry (ICP-
OES) and alkaline solubilization as sample preparation procedure. The
proposed method was successfully applied for elemental determination in

several meat samples, and the concentrations of all elements were detected at
mg g-1. According to the obtained results, the authors pointed out that meat
processing may be associated to the obtained results for Fe, Mg, and P (Nunes
et al., 2013).
SAMPLING AND SAMPLE STORAGE

Although the discussion in this chapter is addressed to meat sample
analysis, some general concepts of sampling and sample storage will be
discussed in this section given its importance for trace element determination,
since, contamination, sample deterioration, and analyte loss may jeopardize
the accuracy of the analysis.
The sampling step of meat samples is a critical point for trace element
determination and must be carefully carried out to avoid contamination or
analyte loss. Thus, proper sample apparatus must be employed, e.g., Fe
devices should be avoided for the determination of this element, the apparatus
used for sampling should also be inert, so the characteristics of the sample are
not modified (European Commission, 2007).
For sample storage, as a general rule, the procedure must keep the original
features of the analyte. Although in the case of total analyte concentration its
chemical form may not be essential, in the event of chemical speciation, the
sample must be stored in such way that the chemical species of interest do not
suffer mutation or degradation (Salomons, 1988).
One example is the European Commission Regulation that established
methods for sampling, sample storage, and analysis to monitor the
concentration of Pb, Cd, Hg, and Sn in foodstuff, including meat samples. This
regulation highlights that care shall be taken to avoid contamination. Thus
sample must be packed in a clean and inert container, and all apparatus should
be washed with pure water and acid in the case of metal determination
(European Commission, 2007).
Given the different characteristics of analytes and matrices, each case
should be carefully analyzed; e.g., Hg determination may suffer from analyte
loss when the samples are stored for prolonged period of time (de Gois and
Borges, 2014), chemical speciation of Cr VI e Cr III in meat samples may be
challenging due to the transformation of Cr VI to Cr III (Marques et al., 2011),
As speciation is susceptible to degradation of the organic As species within
time between sampling and analysis (Amayo et al., 2014), among others.
Therefore, the local regulation and literature should be consulted to

perform sampling and sample storage adequately, when no regulation is
available, tests shall be carried out to assure no contamination, analyte loss or
even analyte deterioration.
SAMPLE PREPARATION
Sample preparation is one of the most important steps for trace-element
determination in meat samples, during which the analyte is released in a
suitable solution for analysis by the selected analytical technique.
Sample drying, grinding, homogenization and particle size selection are
usually the first steps in a sample preparation procedure; these steps are crucial
to assure homogeneous distribution of the analyte in the sample before sample
dissolution procedures, digestion, and extraction, among others. Even though
these sample pretreatment steps seem straightforward, they are prone to issues
such as analyte loss and contamination (European Commission, 2007).
Volatile species such as Hg are susceptible to be lost as a result of sample
heating while drying and grinding, in these cases, systems that avoid heating,
e.g., freeze-drying, lyophilization, and cryogenic mill are preferred as opposed
to conventional drying and grinding procedures. Feldmann (2003) addressed
this theme in a useful book chapter inferring the problems and possible
solutions for a series of cases.
Particle size selection may be performed by simple sieving, however,
contamination must be avoided by proper material selection of the sieve, e.g.,
plastic materials may be a good choice for metallic elements, whereas for
other elements such as the halogens the use of metallic sieve shall not be a
problem (European Commission, 2007).
After the procedures of drying, grinding, and homogenization, most
analytical techniques require the analyte solubilization in an aqueous solution
prior analysis, in this step, the analyte is separated (or partially separated) from
the sample matrix and retained in a liquid solution for further analysis.
Most popular sample preparation procedures applied to determine the total
trace-elements content are the acid digestion using inorganic acids
concomitantly with microwave energy or temperature, extraction procedures
using diluted acids, pyrohydrolysis, solubilization in alkaline media, and
combustion techniques. Direct solid sample analysis has also proven to be an
efficient strategy for trace element determination by reducing the need for
additional sample pretreatment steps, thus minimizing reagents and equipment.
The proper choice of the sample pretreatment technique must take into
account the sample matrix, the analyte, and its concentration in the sample.
Wet digestion in close vessels using a microwave oven or a thermic heating is
trustworthy for the determination of non-volatiles elements in meat samples
(Millour et al., 2011), however volatile species may be lost due heating and
sometimes the formation of volatile species like HX (X = halogen) in the case
of halogens may occur (de Gois et al., 2015).
In wet digestion in close vessels using a microwave oven or a thermic
heating, normally an amount of homogenized sample (from about 0.1 to 2.0
mg) is weighed into a proper flask, an amount of digesting solution (e.g.,
HNO3, HNO3 + H2O2, H2SO4, HF, HCl, Aqua-Regia, among others) is added,
and the flask is then submitted to heating (Ojeda and Rojas, 2014).
The combination of reagents and the temperatures depend on the sample
matrix and the analyte characteristics. This procedure promotes degradation of
the sample matrix and solubilization of the inorganic analyte in a liquid
solution, although care must be taken in order not to produce non-soluble
species of the analyte (Ojeda and Rojas, 2014).
Volatile species may require different sample pretreatment strategies like
pyrohydrolysis and combustion techniques. These have been successfully
applied for the determination of volatile species in various sample matrices
including meat samples (Barnes et al., 2014; Flores et al., 2007).
The basic principle of pyrohydrolysis is the volatilization of the analyte
that is separated from the sample matrix by heating; the volatilized analyte is
then carried to a trap solution for posterior determination. Hence, this
procedure is only suitable for the analytes that are volatile at the temperature
applied. In pyrohydrolysis, the sample is placed in a sample holder inserted
into a quartz tube, followed by heating of the system. The heating promotes
the volatilization of volatile species, which are carried by the carrier gas and
water vapor to a condenser and subsequently to a trap solution; the fractions
are then collected and analyzed (Langenauer and Krähenbühl, 1993).
Combustion procedures, on the other hand, may be applied for both,
volatile and non-volatile elements, requiring the sample to be prone to
combustion. Thus, organic matrices such as meat samples are good candidates
for these sample treatment techniques (Flores et al., 2007). Some important
combustion techniques are the oxygen flask, combustion bomb and
microwave-induced combustion (MIC).
Combustion bomb presents the advantage of simplicity and ability to
decompose samples in only a few minutes (normally less than 30 min
including cooling step). The sample (at least 0.5 mg) is combusted inside of a
stainless steel vessel pressurized with oxygen. The products generated by the
sample combustion are absorbed into a suitable solution at the bottom of the
bomb (Souza et al., 2002). Due to the high temperature generated from the
combustion, it is necessary to cool down the system before removing the
resulting solution. Usually, water or an ice bath may be employed for this
purpose.
Another combustion system that can be applied is the oxygen flask, which
is a simple combustion system composed of a glass vessel containing oxygen
at atmospheric pressure and an absorbing solution inserted at the bottom of the
vessel (Souza et al., 2002). The sample (usually less than 100 mg) is wrapped
with paper and positioned in a platinum holder for subsequent combustion; the
system is pressurized with oxygen up to atmospheric pressure and closed for
posterior combustion. Sample ignition is started by an electrical current
applied to a Pt wire or by a focused infrared lamp; another option is the use of
a piece of paper which is usually ignited before the introduction of the Pt
holder into the vessel (Mesko et al., 2016).
A more recent developed system for sample preparation, is the MIC,
which has been applied for the determination of several analytes in a wide
range of samples including meat (Barin and Flores, 2014). In this procedure,
the ignition of the sample is started by microwave irradiation.
Similarly to other combustion procedures, in MIC the gasses resulting
from the combustion are absorbed in a solution, although in MIC the
absorbing solution is subject to reflux (Barin and Flores, 2014). After the
system is cooled down the solution can be removed, diluted and analyzed
(Flores et al., 2007).
Extraction procedures or even partial degradation of samples are very
attractive options for trace element determination in meat samples, with the
advantage of simplicity and low cost when compared to wet digestion or even
combustion techniques.
Sample pretreatment promoting solubilization of the sample in alkaline
media or analyte extraction using diluted acids assisted by ultrasound energy
has gained increasing interest and provided results that were comparable to
those obtained with normal wet digestion and combustion techniques
(Nóbrega et al., 2006).
One example of the use of alkaline media for meat sample preparation is
the use of tetramethylammonium hydroxide and heating for trace element
determination. This procedure promotes partial decomposition and partial (or
complete) solubilization of the sample, allowing its analysis after the selection
of the proper analytical technique. The procedure is also simple and fast
providing high sample throughput and reduced risk of contamination

(Matusiewicz and Golik, 2004).
Analyte extraction may replace sample decomposition in several
applications. Ultrasound energy and diluted acids are typically employed to
extract the analytes from the sample, resulting in simple methods with high
sample throughput. In this procedure, one amount of the homogenized sample
in a powder form is mixed with an extraction solution (e.g., HNO3, HNO3 +
HF, Aqua Regia) and submitted to shaking, ultrasound energy, microwave
energy, among others (Jagtap and Maher, 2016).
A problem related to sample preparation procedures may be observed
when the analyte concentration in the sample is low. Sample dilution can
jeopardize the analyte determination by some analytical techniques. In these
cases, the LODs can be improved after enrichment by preconcentration
procedures such as dispersive liquid-liquid microextraction (Shrivas and
Jaiswal, 2013), solid phase extraction (Pourjavid et al., 2014), among others.
Another very attractive way to analyze meat samples is using solid and
slurry sample analysis, which decreases the risk of contamination, increases
the detection capability, and reduce the need for special apparatus for sample
pretreatment. Moreover, these procedures allow micro-homogeneity studies
(de Andrade et al., 2016).
Few techniques may require some sample pretreatment to produce pellets
or another form that is suitable for sample introduction into the instrument to
perform direct solid sample analysis, although in some techniques the sample
may be introduced as a powder or in situ (Pereira et al., 2014).
A general problem regarding direct solid sample analysis is that it requires
more capabilities do deal with interferences from the analytical technique, due
to the presence of relatively large quantities of the matrix (de Andrade et al.,
2016).
Analytical calibration that may be applied to the solid samples providing
accurate results may be challenging in direct solid sample analysis. Normally
the behavior of the analyte in the samples should be as similar as possible to
the behavior of the analyte in the standards (Kurfürst, 1998).
In solid sample analysis, the chemical form of the analyte is sometimes
strongly different from the chemical form of the analyte in the solid samples,
compromising the use of a calibration against aqueous standards. In these
cases, calibration may be performed using solid samples, such as certified
reference materials (CRM) (Kurfürst, 1998).
ANALYTICAL TECHNIQUES
FOR TRACE-ELEMENT DETERMINATION
The choice of the analytical technique for trace-element determination

should take into account the analyte characteristics, analyte concentration,
presence of interferences, sample throughput and costs.
Some analytical techniques may be applied for trace-elements
determination in meat samples, among those, electrochemical and
spectrometric methods stand out in publications and applications. Therefore,
potentiometry, voltammetry, atomic absorption spectrometry (AAS), ICP-
OES, inductively coupled plasma mass spectrometry (ICP-MS), atomic
fluorescence spectrometry (AFS), X-ray fluorescence spectroscopy (XRF),
Neutron activation analysis (NAA) and separation techniques (ion
chromatography and capillary electrophoresis) will be briefly described in this
section.
Potentiometric and voltammetric techniques are well-established
electrochemical techniques that allow measuring free and bonded ions, and in
some cases, also the determination of the oxidation state of some elements
(Brown and Milton, 2005).
Generally, potentiometric measurements are performed via ion selective
electrode (ISE) monitoring variations in the equilibrium voltage under zero
current conditions, where, the device equilibrium potential can be described as
a Nernstian function of the analyte activity in the sample solution (Brown and
Milton, 2005; Tajik et al., 2013).
Ion selective electrodes have been applied for trace-elements
determination in different samples, including biologic fluids and meat samples
after proper sample pretreatment (Brown and Milton, 2005; Tajik et al., 2013),
reaching LODs in a mass fraction in the order of 10-11. The main problem
associated with the use of ISE in complex matrices is the presence of
interferences when the sample pretreatment is not able to efficiently separate
the analyte from the matrix components (Brown and Milton, 2005).
The determination of Zn, Cd, and Pb was directly performed using
potentiometric stripping analysis with a continuous flow cell. As reported by
Monos and Palmero (2006), the results obtained for these elements were in
agreement with those obtained using ICP-MS. The direct determination of Cd,
Pb, and Cu has also been reported, the LODs achieved by the proposed
method were 0.30, 1.7 and 3.8 µg L-1, respectively (Suturović et al., 2014).
Analyte preconcentration may also be achieved using voltammetry

techniques by accumulating the analyte on electrode surface prior analysis, for
those methods the achieved LODs can reach up to 10-12, in a mass fraction
(Brown and Milton, 2005).
Some applications of anodic stripping voltammetry include the
determination of Se(IV) (Ochab et al., 2017) traces of Pb(II) and Cd(II)
(Zhang et al., 2016), Tl(I) (Węgiel et al., 2016) and Ag(I) (El-Mai et al., 2016)
achieving LODs at ng L-1 level.
Spectrometric techniques are powerful tools that can provide high
sensitivity, accurate and precise results for the determination of trace-elements
in a wide range of samples. Among the spectrometric techniques, AAS, ICP-
OES, ICP-MS, and AFS are the most widely applied techniques for trace-
element determination (Korn et al., 2007).
The basic principle of an AAS analysis is the absorption of radiation from
a primary source by gaseous atoms in the ground state (Becker, 2005).
Different atomizers may be used in order obtain gaseous atoms of the analyte
in the ground state; flame and electrothermal atomizers are the most common.
Thermospray, cold vapor generation (in the case of Hg) and atomization by
electrothermic or flame heating in a quartz tube are also used.
In flame atomic absorption spectrometry (FAAS) the liquid sample is
typically introduced in liquid form, and the atomization occurs in a flame
normally composed of the mixture of acetylene/air or acetylene/nitrous oxide.
This technique provides enough sensitivity for the determination of several
elements in the range of mg L-1.
The use of a flame composed by acetylene/air is suited for the
determination of most elements, although for elements for which refractory
oxides may be formed, such as Ca, Al, Ti, W and Mo, the mixture
acetylene/nitrous oxide may be preferred or acetylene/air with the use of a
releasing agent, such as La (Welz and Sperling, 1999).
Comparing with FAAS analysis, the determination of trace-elements using
electrothermal atomization atomic absorption spectrometry (ET-AAS) is more
sensitive and provides improved LODs, which can also be enhanced by
combination with preconcentration procedures (Becker, 2005; Welz and
Sperling, 1999).
In electrothermal atomization atomic spectrometry, using a graphite
furnace (GF-AAS), the sample may be introduced in liquid form, slurry or
even in the solid form using appropriate apparatus, one of the main benefits of
this technique is that the LODs are in general 10 to 100 times better than those
with FAAS (Becker, 2005). The improvement in LOD associated to GF-AAS
is mainly due to the high efficiency of atomization and the high residence time
of the atoms into the graphite tube.
Chemical vapor generation (CVG) is applied to determine the hydride
forming elements (e.g., Hg, As, Se, Sb, and Sn), the hydride form is then
atomized by heating in e.g., a quartz cell using electrothermal heating or
flame. In cold vapor (CV), which is applied for Hg determination, the analyte
is reduced to gaseous Hg0 which can be measured directly using AAS (Welz
and Sperling, 1999).
The CVG and CV techniques may also be used in concomitance with a
graphite furnace, tungsten coil or thermospray atomizer. Moreover,
determination of volatiles species generated by CVG can also be determined
by AFS, FAAS, and ICP-MS (Gil et al., 2005; Becker, 2005). The success of
CVG is due to characteristics such as high analyte transport efficiency to the
atomizer and, in some cases, potential for speciation analysis (Welz and
Sperling, 1999).
High-resolution continuum source atomic absorption spectrometry (HR-
CS AAS) is one improvement of the AAS techniques. This technique uses a
high-intensity continuum source (xenon short-arc lamp), which enables the
determination of most elements using only one radiation source, besides
allowing the possibility of simultaneous determination. Correction for the
continuous and structured background is also an important improvement of the
AAS techniques when using HR-CS AAS (Welz et al., 2005).
Elements that show absorption lines at UV-Vacuum region may be
determined relatively free of interferences via high-resolution molecular
absorption spectrometry (MAS HR-AAS) by producing diatomic molecules in
the atomizer and measuring in one or more of their absorption bands, for
example, F and Cl may be determined via molecular absorption of strontium
monofluoride and strontium monocloride, respectively (Butcher, 2013;
Limburg and Einax, 2013; Ozbek and Akman, 2012; Flórez and Resano, 2013,
Pereira et al., 2014).
Inductively coupled plasma optical emission spectrometry is an analytical
technique for multi-element determination in a wide range of samples
(Brenner et al., 1999). In this technique, an inductively coupled plasma (ICP)
ionization source is employed to produce atomized and ionized excited-state
species that emit radiation at specific wavelengths. The main advantages of the
ICP-OES are the potential for multi-element determination, adequate LOD for
most applications, wide linear range and high sample throughput (Dean,
2003).
In ICP-OES the sample is usually introduced in the liquid form using a

nebulizer, although ETV, CVG, CV and laser ablation may also be employed
(Shizhong et al., 2005; Giné, 1999; Dubois et al., 2005; Bressy et al., 2013).
Inductively coupled plasma mass spectrometry is another multi-element
technique that relies on the use of an ICP, in this case as an ion source. In this
technique, the ions generated in the ICP are carried to a mass analyzer that
separates them according to their mass charge ratio (m/z), the ions are then
directed to a detection system. One remarkable characteristic of the ICP-MS is
the combination of high energy of the ICP with a mass spectrometer obtaining
an isotopic and elemental analyzer at the same instrument and achieving LODs
down to ng L-1 (Becker, 2007; Houk et al., 1992). The determination of sixteen
elements in different food samples after sample microwave assisted digestion
using concentrated acids was performed by ICP-MS achieving LODs in ng g-1
(Nardi, 2009). Concentrations of non-essential and essential trace-elements
were determined in meat samples from cattle reared in different systems by
ICP-MS. The samples were also prepared by microwave assisted acid
digestion and the LODs obtained were in the range of µg g-1 (Blanco-Penedo
et al., 2010).
One of the critical issues for ICP-OES and ICP-MS analysis is the
presence of spectral and non-spectral interferences. Spectral interferences in
ICP-MS are especially problematic for elements at low mass, generally lower
than 80 u.m.a. Different approaches have been proposed in order to correct
spectral interferences in ICP-MS such as the use of special equipment with
collision and reaction cells, triple quadrupole ICP-MS, the use of non-
traditional sample introduction systems, high-resolution instruments, or even
classical corrections using mathematic equations. Blank subtraction and
separation techniques have also been used in order to remove potential
interferences.
The correction of non-spectral interferences, on the other hand, may be
challenging, the most applied techniques are the use of matrix-matching
standards, standard addition, internal standardization and separation
techniques (Vanhaecke, 2002; Frei and Gerdes, 2009; Castro et al., 2008;
Tanner et al., 2002; De Muynck et al., 2009).
Atomic fluorescence spectrometry is an atomic emission technique that
has also been extensively applied for trace-element determination (Greenfield,
1995). This technique combined with hydride generation (HG AFS) allowed
the determination of As, Sb, Se, Te, and Bi with LODs from 0.10 to 0.45 μg g-
1 (Feng and Fu, 1998). Cadmium determination by vapor generation using
AFS was performed in seafood achieving a LOD of 0.012 μg L-1 (Yang et al.,
2015). Moreover, Pb was determined by laser-excited atomic fluorescence

using graphite furnace atomization system and a LOD of 10 ng L-1 was
achieved (Wagner, 1996).
X-ray fluorescence (XRF) is routinely applied to determine metals and
non-metals in a broad range of samples, including meat and meat products.
The XRF sensitivity depends on the energy of radiation, the geometry and the
detector used (Brown and Milton, 2005).
The use of total reflection XRF (TXRF) can improve the LOD down to 2
pg, and with synchrotron radiation source and high photon fluxes, LOD as low
as 0.03 pg can be achieved. Additionally, heavy charged particles can induce
X-ray emission (PIXE, particle-induced X-ray emission), allowing
determinations with low LOD and precision greater than 5% (Brown and
Milton, 2005). The TXRF technique was successfully applied for the
determination of trace-elements such as Cr, Mn, Fe, Ni, Cu, Zn, As, and Pb in
freshwater rotifers and ciliates (Woelfl et al., 2016), and for trace and ultra-
trace analysis in liquid samples (Marguí et al., 2014).
In activation analysis techniques, the sample composition is determined
based on the activation of the stable isotopes followed by radiation
measurements. Neutron activation analysis is the most common activation
analysis technique, in which neutrons are used to irradiate and activate the
analytes (Ali et al., 2017; Brown and Milton, 2005).
During analysis, radionuclides with specific decay are produced by a
nuclear reaction between neutron and the analyte isotope; the natural
radionuclide emission is then monitored by an appropriate detector. In NAA,
mass fractions lower than 10-15 can be detected. Analysis of solids and liquids
samples can also be performed by NAA (Ali et al., 2017; Brown and Milton,
2005).
Separation techniques, especially combined with ICP-MS/OES, AAS, and
AFS, are frequently used for metal and metalloid speciation, although
macromolecules speciation and separation of ions of different charges are
easily performed by capillary electrophoresis (CE) while gas chromatography
(CG) coupled to ICP-MS allows the determinations of volatile species with
high sensitivity (Kroukamp et al., 2016).
Speciation of trace Hg in seafood was performed by HPLC after a
preconcentration step, the LODs obtained for Hg(II), methylmercury (MeHg),
ethylmercury (EtHg), and phenylmercury (PhHg) were in the range of ng g-1
(Dong et al., 2004). Metal ions, and the species Cu(II), Zn(II), Pb(II), and
Mn(II), were determined by capillary electrophoresis, reaching LODs of 10-8
mol (Isoo et al., 2003).
Table 2 presents some selected applications of trace-element

determination, pointing out sample preparation and analytical techniques. The
combination of these techniques is responsible for the successful results
achieved by the authors.
In some cases, the LODs were improved by the use of direct solid sample
analysis coupled to a sensitive analytical technique, as in the event of the
determinations by ETV-ICP-MS, INAA, and XRF. In other cases, as the LOD
was not a critical factor, but rather the analyte species, the sample preparation
was performed by sequential extraction procedures and the analytes detected
by HPLC-ICP-MS.
CONCLUSION
The trace-element determination in meat samples is an important task that
should be performed carefully. Contamination and analyte loss are common
issues related to trace element determination; hence attention shall be taken
during sampling and sample storage.
Most analytical techniques require the sample to be prepared prior to
analysis, and there is currently different sample preparation techniques
available. The choice of the sample preparation technique must take into
account relevant factors such as sample matrix, the analyte and its
concentration in the sample, whereas, in some cases, preconcentration steps
might be required in order to improve the LODs of the analytical methods.
Furthermore, direct solid sample analysis may be performed providing a
more straightforward approach, although only some analytical techniques
allow for its use. The choice of the analytical technique that may be applied
for trace-element determination shall consider the analyte characteristics,
analyte concentration, spectral and non-spectral interferences, sample
throughput and cost. Among the analytical techniques currently available for
trace-element determination in meat samples, electrochemical and
spectrometric methods stand out, although separation techniques may also be
applied concomitant to these techniques providing a powerful tool for
chemical speciation.
Table 2. Selected applications for trace-element determination in meat samples
Sample Analyte Sample preparation Analytical LOQs Reference

technique
Molluscs, crustaceans, Li, Al, V, Mn, Co, Ni, Closed-vessel ICP-MS From 0.001 µg g-1 (Millour et al.,
and fishes Cu, Zn, Ga, Ge, As, digestion to 0.100 µg g-1 2011)
Sr, Mo, Ag, Cd, Sn,
Sb, Te, Ba, Hg, and Pb
Bovine muscle, bovine Cl Direct solid sample ETV-ICP- 5 µg g-1 (de Gois et al.,
liver, pig kidney, muscle analysis MS 2015)
tissue, and beef liver
Bovine and fish tissues Ca, Cu, K, Mg, Na, P, Oxygen bomb ICP-OES Not informed (Souza et al.,
S, and Zn combustion 2002)
Chicken, duck, pork, Cd, Hg, Pb, As, Cr, Closed-vessel ICP-MS Not informed (Wu et al.,
freshwater fish, marine Cu, Fe, Zn, Mn, Mo, digestion 2016)
fish, and shrimp Ni, Co, Se, and Ti
Fish As, Co, Fe, Hg, and Zn Direct solid sample INAA From 0.003 to 1 (Avigliano et
analysis μg g-1 al., 2016)
Pig, bovine, lamb, Zn Ultrasound-assisted FAAS 1.8 μg g-1 (Yebra-
rabbit, veal, and turkey extraction Biurrun et al.,
2005)
Rats Pb Alkaline GF-AAS From 51 to 71 μg (de Sousa et
solubilization and g-1 al., 2013)
closed-vessel
microwave digestion
Sample Analyte Sample preparation Analytical LOQs Reference
technique
Fish As, Cd, Co, Cr, Cu, Microwave-induced ICP-MS From 0.001 to 0.016 (Macie et al.,
Fe, Mn, Mo, Ni, Se, combustión μg g-1 2014)
and Zn
Fish Ca Alkaline solubilization GF-AAS 0.13 μg g-1 (Gomes et al.,
2016)
Muscle Na and K Open-vessel digestion IC Not informed (Haber and
Atalla, 1986)
Fish Arsenolipids Sequential extraction HPLC-ICP- Not informed (Amayo et al.,
MS 2014)
Coral Mg, P, S, and Sr Direct solid sample XRF Not informed (Nguyen et al.,
analysis 2014)
Lobster, bovine Mn, Rb, and Ni Direct solid sample HR-CS GF From 0.002 to 0.1 μg de Andrade et
muscle, and oyster analysis AAS g-1 al., 2016
ACKNOWLEDGMENTS
The authors are thankful to Conselho Nacional de Desenvolvimento
Científico e Tecnológico (CNPq), Coordenação de Aperfeiçoamento de
Pessoal de Nível Superior (CAPES), Fundação Araucária, Fundação de
Amparo a Pesquisa no Rio de Janeiro (FAPERJ) and Rio de Janeiro State
University – Prociência.
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Chapter 5
THE USE OF ELECTRICAL STIMULATION IN

MEAT PRODUCTION
Paolo Polidori1,* and Silvia Vincenzetti2

1
School of Pharmacy, University of Camerino, Camerino (MC), Italy
2
School of Biosciences and Veterinary Sciences, University of Camerino,
Matelica (MC), Italy
ABSTRACT
The present chapter describes what is known about the effects of the
use of electrical stimulation of carcasses of meat animals, including the
effects on meat tenderness and meat sensorial characteristics. Electrical
stimulation as a process involves passing an electric current through the
carcass of freshly slaughtered animals. Electrical stimulation has been
extensively used since the 1950s to hasten the onset of rigor mortis and to
modify steps of the glycolytic pathway. Many studies conducted in the
USA, in New Zealand, Australia and Europe have involved a variety of
electrical stimulation methods on different types of meat animals. Data
reported in many studies suggest that electrical stimulation, through
hastening rigor changes, can significantly reduce in the carcasses of meat
animals the phenomenon of cold shortening, one of the major cause of
meat toughness. Although it is well established that electrical stimulation
increases the rate of post mortem glycolysis, other biochemical and
biophysical effects have been implicated with the use of this technology,
*
Corresponding author: Email: paolo.polidori@unicam.it.
134 Paolo Polidori and Silvia Vincenzetti
including the possibility that electrical stimulation also results in physical

disruption of muscle structure. Electrical stimulation can be considered as
a part of the total meat production chain from slaughter to final sale, and
has particular advantages for hot boning, where the shortening and
toughening conditions that would occur for non stimulated muscles
during chilling are avoided.
Keywords: electrical stimulation, meat production, meat tenderness.
INTRODUCTION
Tenderness is generally judged as the most important quality parameter of
fresh meat. A number of procedures have been developed for increased meat
tenderness: suspension via the pelvic bone, mechanical restraint of muscles,
conditioning, cooler ageing, high temperature conditioning, delayed chilling,
blade or needle tenderization, use of tropical plant or fungal enzymes, etc.
(Polidori et al., 1996). All of these procedures cause changes in meat
tenderness via effects on the contracting machinery (muscle fibres), the
collecting-harnessing-reinforcing structures (connective tissues), or both.
Factors affecting muscle tenderness have been extensively studied over the
past 50 years. Initially, the connective tissue component of meat received the
greatest attention; since 1960, the state of muscle contraction following rigor
mortis has been the most intensively studied (Ouali, 1990; Thompson, 2002).
The discovery that muscle shortening is one of the major causes of meat
toughness has led to the realization that post mortem treatments will usually
outweigh live-animal factors such as breed, age, stress and preslaughter state
in determining palatability (Locker & Hagyard, 1963; Marsh & Leet, 1966;
Penney & Dransfield, 1979; Jeacocke, 1984). Since cold shortening and,
indeed, thaw shortening occur only in pre rigor muscle (Marsh & Thompson,
1958), it is evident that these can be prevented by ensuring that rigor mortis is
achieved before meat is either chilled or frozen. Rigor development in meat
producing animals can take up to 36 h, so the aim of the meat scientists has
been to hasten rigor mortis process, defined when adenosine triphosphate
(ATP) production ceases. Early muscle studies indicated that rigor
development can be brought forward by ante mortem stress, by maintaining a
high carcass temperature or by electrical stimulation of carcasses (Chrystall &
Devine, 1985). The first approach is in conflict with the actual legislation
The Use of Electrical Stimulation in Meat Production 135
concerning animal welfare; the second one is expensive, and for this reason is
not widely applied. Electrical Stimulation (ES) of carcasses soon after
slaughter emerges as the most ingenious and practical approach in inducing
early rigor development. It does this by causing muscles to undergo work via
anaerobic glycolysis, resulting in an initial pH fall (pH) followed by a change
in the rate of pH fall (dpH/dt); the combined effect is that the muscles enter
rigor mortis before the muscle temperature falls to values producing cold
shortening and toughening (Devine et al., 2014).
Rigor Mortis
The merit of using ES in meat technology is intimately associated with the

development of rigor mortis in muscle, with the cold shortening phenomena,
with thaw rigor, and with meat ageing. Brief attention is therefore given to
these topics in preparation for the more extensive discussion on ES which
follows.
After an animal dies, its muscles live on in the pre rigor state. They are
reversibly extensible and can be excited to contract. If starved of oxygen, they
enter rigor mortis some minutes or hours post mortem and become
nonexcitable and rigid. It is obvious that, in the carcasses of meat animals,
various muscles will have different rates of fall of temperature post mortem,
according to their proximity to the exterior and their insulation. As a result, the
rates of post mortem glycolysis will tend to be higher in muscles which are
slow to cool, and vice versa (Lawrie, 1991). From extensive studies of rigor
development, it is evident that the biochemical changes are likely to be the
same for all vertebrate species, with the disappearance of ATP and creatine
phosphate (CP) and, in glycolytic activity, the appearance of considerable
quantities of lactate ions, up to 100 mol/g muscle (Chrystall & Hagyard,
1976; Lee et al., 2000; Hopkins et al., 2014). The ultimate pH of muscle is
inversely related to the accumulation of these ions, being relatively high
(approximately pH 7.0) in excessively exhausted animals and low
(approximately pH 5.5) in well fed and rested animals (Hopkins et al., 2011).
Cold Shortening
The tenderness of meat removed from the carcass in a pre rigor condition
is highly dependent ion the extent of the cold shortening which occurs after
excision (Marsh & Leet, 1966). The relationship between shortening and
tenderness is complex; Locker & Hagyard (1963) examined the shortening
which accompanies or precedes rigor onset at low temperature (2°C). Their
investigations revealed an interesting cold shortening phenomenon in which
the exposure of excised fresh bovine muscles to temperature near the freezing
point causes very appreciable shortening. Muscle from beef animals shortens
by 50% or more prior to and during the development of rigor mortis if held at
2°C (Chrystall & Devine, 1985). The magnitude of this so-called cold
shortening increases with the drop in temperature toward 0°C and it is reduced
by increasing post mortem delay before chilling.
Red muscles are relatively susceptible to cold shortening, while so-called
white muscles are minimally affected by the conditions which cause cold
shortening (Devine et al., 1984). A few muscles such as the muscles of the
neck, are severed during carcass dressing and are likely to shorten. Some other
muscles, such as the Longissimus dorsi, remain attached to the skeleton, but
since most of the constituent fibres insert into flexible epimysium, the muscles
are able to shorten. In fact, even muscles that are fixed at both ends are
capable of cold shortening over a part of their length if they are subjected to a
differential chilling rate along their length. Shortening produced in pre rigor
muscle is the greatest possible cause of toughening in cooked meat, animal age
can be merely a secondary cause (Devine et al., 1999). If rigor muscle is aged
for a number of days before cooking, it usually becomes appreciably more
tender. The decrease in tenderness which is associated with the onset of rigor
mortis is gradually reversed as the time of post rigor conditioning increases
(Lawrie, 1991).
Electrical Stimulation
The association between meat and electricity dates back to some of the
earliest muscle physiology experiments: from Galvani’s time the use of
electricity to study muscle function increased (Chrystall & Devine, 1992). The
earliest reported use of electricity for meat improvement is its purposed by
Benjamin Franklin in 1749 to electrocute turkeys with the result that they were
uncommonly tender (Devine et al., 2014). During the 1950s it was shown that
ES could improve meat tenderness of beef, but no commercial application of
the process occurred. Stimulation of horse muscle has been used to facilitate a
trial of microbial growth on pre rigor and rigor muscles from the same
animals (Ingram & Ingram, 1955).
Several hypotheses have been suggested that would account for the meat
tenderizing action of early post mortem ES. The originators of the process
(Harsham & Deatherage, 1951) proposed that activity of acid proteases was
increased by the rapid acidification induced by the treatment. The subsequent
discovery of cold shortening led, some years later, to an alternative
explanation, according to which ES accelerates rigor onset and so prevents
shortening and the toughening associated with it (Hwang et al., 2003).
The incorporation of ES into the slaughtering process was first used in
New Zealand (Carse, 1973) and then Australia to avoid toughness resulting
from cold shortening. ES has become of increasing interest to meat processors
because it requires little changes in normal abattoir practice and the removal of
meat from the carcass pre rigor (hot-boning) could become a practical
possibility. ES involves passing an electric current through the bodies or
carcasses of freshly harvested animals. This electric current causes the muscles
to contract, increasing the rate of glycolysis and results in an immediate
reduction in muscle pH (pH) that ranges from 0.6 pH units at 35°C to 0.018
units at 15°C, suggesting that ES of warm carcasses should take place soon
after slaughter to maximize efficacy (Devine et al., 2014).
There are several physical methods by which ES could be applied, many
different possible electrical specifications, ad in reality many different
perceptions of the response. Most commercially used ES systems employ the
conveying rail as ground, and a live electrode contact some other part of the
body, carcass or side. In the most basic systems the live electric contact is a
clip manually applied to the head end of the animal’s body that is suspended
by one or both hind legs (See Figure 1).
More sophistication is required as voltages increases and as application of
the electrode becomes automated. Safety has been of utmost importance
during experimentation and implementation of ES in New Zealand, Australia,
the USA and Europe to the point that in some instances safety concerns have
effectively prevented commercial adoption of the process.
Figure 1. Electrical Stimulation of sheep carcass.
ELECTRICAL STIMULATION PARAMETERS

Harsham and Deatherage (1951) used voltages ranging from less than 50
to greater than 3000 V peak, the latter giving better current distribution
throughout the beef cercasses. Many researchers have shown how both low
and high voltage systems can benefit meat eating quality for both beef and
sheep meat (Polidori et al., 1999; Shaw et al., 2005; Hopkins et al., 2006). A
new generation of ES parameters was developed in Australia for both
sheep/lambs and beef, based on pre-dressing medium-voltage stimulation, to
avoid the danger associated with high-voltage (Toohey et al., 2008). This
study used an optimised setting of 800 milliamperes (mA), varying voltage
with a peak of 300 V, a pulse width of 0.5 milliseconds (ms) and a unipolar
waveform. The stimulation treatment was applied for approximately 60 s with
a frequency of 15 Hz. The results obtained in that study confirmed that ES
effectively increased the rate of pH decline when the current is administered to
wool on carcasses with no negative impact on any meat quality trait., with an
improvement in tenderness determined 24 h after slaughtering.
Factors Affecting pH
The rate of post mortem pH and temperature decline can significantly

impact on the resulting tenderness of meat (van de Ven et al., 2014). For
optimal eating quality sheep meat should reach pH 6 when the carcass
temperature is between 18 and 35°C (Thompson et al., 2005). ES acts by
accelerating the onset of rigor (pH 6.0) resulting in a higher temperature at
which a carcass enters rigor (Hwang et al., 2003). The temperature at the point
a carcass reaches pH 6.0 and enters rigor can be used to predict meat quality
(Thompson et al., 2005). If the carcass temperature falls too fast before the
onset of rigor then cold shortening may result, which can have adverse effects
on meat tenderness (Tornberg, 1996).
The Asutralian Sheep Meat Eating Quality (SMEQ) program identified
various temperature ranges for optimal eating quality depending on the market
for the product (Toohey et al., 2008). It was concluded that the target
temperature range to achieve pH 6.0 should be 18-25°C for short aged meat
destined for the domestic market.
When muscles of freshly slaughtered animals are electrically stimulated,
they contract. There is a concomitant increase in biochemical reactions in the
muscle cells leading to an accumulation of lactate resulting in an immediate
drop in the muscle pH (pH). The magnitude of pH is governed by muscle
fibre type, initial glycogen stores within the muscle, the electrical parameters
(current, frequency, pulse shape, stimulation duration), muscle temperature,
and the time after death at which ES is applied (Devine et al., 2014).
The increase in glycolytic rate can be explained by ES causing a reduction
in the energy of activation, specifically the amount of energy necessary to start
the reaction in excess of that already possessed by the molecules. If the energy
of activation is high, the rate is low and vice versa. In addition, pH is
strongly affected by temperature, being faster at higher temperatures, so that
an increase has a greater effect on pH of stimulated muscles than that of
unstimulated muscle. These changes are possibly a consequence of irreversible
changes to ATPase activity that dictate the rate of ATP hydrolysis and
therefore pH decline (Devine et al., 2014).
Frequency of ES
The higher is the pH, longer is the time for muscles of stimulated
carcasses to reach pH 6.0; the frequency of the applied voltage is an important
determining factor. At frequencies between 10 and 20 pulses/sec pH values
are, respectively, 40 and 75% greater than tat 50 and 100 pulses/sec (Polidori
et al., 1996). The frequency optimum of around 9-16 pulses/sec seems to hold
for most muscles of sheep and beef carcasses. When considering such complex
interactions, a clearcut assembly of the influences of stimulation parameters is
not to be expected. Thus, with short stimulation periods, high frequencies (50
to 100 pulses/sec) give higher pH values than those produced by lower
frequencies. If longer stimulation periods can be used (120 sec), then 9-16
pulses/sec give the highest pH. The lower frequencies produce a slightly lower
peak tetanic tension than the higher frequencies but maintain their peak
tension for a considerable longer time.
An advantage of lower stimulation frequencies is the lowered energy
input. For example, at 14.28 pulses/sec, the energy input is only one-seventh
of that at 100 pulses/sec; this significantly reduces heating at the electrode
contacts and in the musculature (Chrystall & Devine, 1978). In an experiment
based on the application of 60 Hz frequency ES on beef sides, Takahashi et al.
(1984) found that this frequency produced very extensive fracturing with
breaks appearing on average every 6 mm of fibre length. This treatment
resulted in a very significant tenderizing relative to that observed in the
unstimulated control sides.
TENDERIZATION MECHANISMS OF
ELECTRICAL STIMULATION
Meat tenderness has been considered by many authors as the prime
determinant of consumer satisfaction with meat purchase. Although the
ultimate measure of tenderness or toughness lies with the consumer, objective
assessments can be made with a wide variety of mechanical devices, because
these devices can reliably indicate differences attributable to animal and
processing factors (Destefanis et al., 2008). Meat tenderness-toughness
depends on both the myofibrillar strength and the connective tissue strength.
ES seems mainly to modify the myofibrillar strength, although a study
suggests that ES could also have an impact on the connective tissue

component (Mills et al., 1989).
The major improvement in tenderness of electrically stimulated meat was
originally due to prevention of cold shortening, as above mentioned. However,
ES also appears to improve tenderness above that which can be accounted for
inhibition of cold shortening. In fact, ES generally improves tenderness even
though no differences in sarcomere length may be evident between stimulated
and unstimulated muscles (Uytterhaegen et al., 1992; Toohey et al., 2008;
Polidori et al., 2016). Evidence suggests that ES may also benefit tenderness
by causing the rapid release of lysosomal enzymes and/or by physical
disruption of the electrically stimulated muscle fibres. The linkage between
improved meat tenderness and physical disruption is plausible, as ES treatment
has improved tenderness under circumstances where no cold shortening was
evident. However, it is unclear whether it is the physical disruption per se that
has caused the effect or whether the physical disruption facilitates ageing in
other ways, such as enhancing proteolysis (Devine et al., 2014).
Lysosomal enzymes are known to have the ability to degrade the
myofibrillar proteins under the high temperature and low pH conditions
prevailing in post mortem muscle (Sorinmade et al., 1982). Release of the
lysosomal enzymes plays an important role in meat tenderization following
application of ES. The disruption of the lysosomal membranes with the release
of the lysosomal enzymes appears to be responsible, at least in part, for the
increased tenderness of electrically stimulated meat.
Figure 2. Contacts for ES: rectal probe (left) and clip for nostrils (right).
Sorinmade et al. (1982) presented ultrastructural evidence that ES causes

contracture bands with superstretching of the myofibrils, resulting in the
absence or presence of poorly defined A-bands, I-bands and Z-bands. This
further substantiates the fact that physical disruption is another mechanisms
whereby tenderization occurs as a consequence of ES.
ES accelerates activation of calpains and subsequent proteolysis of
myofibrilar/cytoskeletal proteins, which results in improvement of meat
tenderness (Lee et al., 2000) since the rate constants for the post mortem
decline in calpain and for tenderization were similar, suggesting the loss of
calpain activities is inversely related to tenderization. There are several
possible explanations why ES might increase the activity of specific enzymes
such as the calpains. It may be due to some intrinsic effect associated with the
rapid pH decline, with a low pH at high temperatures, that affects the
processes governing the activation and inactivation of the calcium dependent
proteases, or it could be due to a flow-on effect associated with a significant
increase in free calcium, which leads to activation of the calpains, especially
-calpain (Devine et al., 2014).
Effects of ES on Different Meat Animals
ES causes an increase in the rate of post mortem glycolysis and prevents

excessive cold shortening during rigor in common livestock species. Hwang et
al. (2003) described the three classical area by which ES is proposed to elicit
changes in post mortem muscle:
1. Prevention of cold-induced shortening by ensuring rigor mortis

occurs under optimal conditions;
2. Physical disruption of the muscle fibres;
3. Acceleration of proteolysis
Acceleration of proteolysis could be classified as a secondary effect

mediated through the time-temperature-pH interactions, affecting factors such
as enzyme stability and activity. ES of carcasses after slaughter is a process
that can have a significant effect on meat tenderness, and for this reason it has
been used for beef, pigs, deer, goats, sheep, cattle, buffalo, poultry, alpacas
and donkeys (see Table 1).
The first study in which ES was used to hasten the rigor mortis process
was conducted in 1951 by Harsham and Deatherage. These workers stimulated
beef carcasses by attaching multiple electrodes on the surfaces and stimulating

at 1700-3500 V and 50 pulses/sec. Their pH/time curves closely resemble
those obtained in more recent works: They showed that ES helped to tenderise
meat and suggested that this was due to release of catheptic enzymes during
ES. No more was heard of the notion of ES of large meat animals until it was
resurrected in New Zealand (Carse, 1973) for lambs and later for beef
(Bendall, 1980). The review published by Hwang et al. (2003) summarize the
biochemical and physical effects of ES on beef and sheep meat; the
conclusions stated that ES can result in no or a detrimental effect on meat
tenderness. Provided pre-slaughter animal status and chilling regime are taken
into account when the total energy input from ES is decided, meat tenderness
will be improved. However, ES does not improve inherently tender meat
beyond baseline toughness.
High voltage ES (700 V, 1400 V peak, pulses 1 sec/on – 1 sec/off, 60 Hz)
on buffalo carcasses resulted in a significantly more rapid pH fall in muscle
Longissimus dorsi thoracis when compared to non stimulated control (Soares
& Arêas, 1995). ES produced buffalo meat with better texture characteristics,
and the final product from stimulated carcasses had superior quality compared
to control group. The results obtained in this study indicated that that ES can
be applied in buffalo slaughter, and results in a shorter period for the onset of
rigor mortis.
ES has been used on broiler carcasses to accelerate the production of
boneless meat by reducing or eliminating the need for the costly ageing
process (Sams, 1999). ES systems using high amperages of 350 to 500 mA per
bird induce such forceful contractions that the muscle not only exercises to
accelerate ATP depletion, but tears itself. The physical disruption tenderize the
meat as the rigor mortis acceleration from the exercising prevents toughening.
The combination of these two mechanisms has generally made high amperage
ES more effective at reducing the need for the ageing period and effective
without combining it with other procedure. High amperage ES results in
sufficient reduction in the toughening to produce meat deboned immediately
after chilling (1.5 to 2 h post mortem) that would be considered slightly to
moderate tender to consumers (Sams, 1999). Using the same amperage and
pulsing that is effective on broilers, ES was not effective in reducing the need
for ageing in turkeys (Owens & Sams, 1997) or ducks (Owens et al., 1997.
Differences in muscle fibre type and metabolism (Walker et al., 1996) may
account for the different response to ES among species.
A study conducted on 96 goats of similar age, weight, quality grade and
yield grade were conducted by McKeith et al. (1979). Results obtained in this
study indicated that ES improved tenderness of goat muscles and that ES can
be performed at any of several stages during the slaughter-dressing sequence,
Advantages for a particular site and for split vs unsplit carcasses were not
apparent, thus meat packers can install ES equipment at any point in the
slaughter-dressing sequence where space is available and safety of workmen is
not compromised.
The first study in which ES was applied to Danish Landrace pigs was
carried out by Hallund & Bendall (1965); the acceleration of pH fall after ES
lasted during the whole course of the pH-time curves. Gigiel et al. (1984)
demonstrated that ES produced more tender pork after conventional chilling of
Large White x Landrace pigs, while Westervelt & Stouffer (1978) did not find
improvement in tenderness of the Longissimus muscle obtained by electrically
stimulated Yorkshire hogs. The Authors suggested that the unexplained
mechanisms of response of porcine muscle Longissimus to ES may be similar
to the cold-shortening characteristics of pork muscle which are only a tenth of
those observed in the bovine and ovine species. A study conducted by Wiley et
al. (1989) revealed that loin chops obtained from electrically stimulated hog
carcasses were less tender than those from unstimulated carcasses. The
updated knowledge about use of ES on pigs said that with optimum technical
parameters ES is effective on pork, especially for some breeds; very stress-
susceptible breeds such as Pietrain do not give good results (Devine et al.,
2014). ES cannot produce a PSE-like condition, because PSE in pork arises
from severe myosin denaturation pre rigor and in worst cases a lot of drip is
produced because myosin is a major muscle protein. Tenderization occurs via
cytoskeletal denaturation post rigor mortis of smaller amounts (<10%)
cytoskeletal proteins, therefore no extremes of drip thus no PSE.
A total of 14 female red deer were included in a study on the effects of
low voltage carcass electrical stimulation on meat tenderness, colour stability
and water-holding capacity (Wiklund et al., 2001). Carcasses were randomly
allocated to either electrical stimulation treatment (90-95 V unipolar pulses,
7.5 ms duration, 15 Hz for a duration of 55 s) or no electrical stimulation.
Compared to controls, ES increased the rate of muscle pH decline and
produced lower shear forces at 1 day, 1 week and 3 weeks post-mortem, but
these differences disappeared by 6 and 12 weeks post-mortem. This study
showed no detrimental effects of using ES meat colour stability or drip loss.
TABLE 1. STUDIES WHICH HAVE MEASURED THE EFFECTS OF ES IN MEAT ANIMALS
Species Voltage applied Authors

Lambs 250 Carse, 1973
Lambs 3600 Chrystall & Hagyard, 1976
Pigs 110 Westervelt & Stouffer, 1978
Pigs 550 Wiley et al., 1989
Goats 100 McKeith et al., 1979
Beef 32 Taylor & Marshall, 1980
Beef 300 Smulders et al., 1986
Buffalo 700 Soares & Arêas, 1995
Turkey hens 570 Owens & Sams, 1997
Broiler 450 Owens & Sams, 1998
Deer 95 Wiklund et al., 2001
Alpaca 300 Smith et al., 2016
Donkeys 28 Polidori et al., 2016
In a study conducted on 50 huacaya alpacas (Smith et al., 2016), the use of

a medium voltage (300 V) ES significantly reduced on muscle Longissimus
thoracis et lumborum purge values by 3% and shear force at 5 and 10 days
post mortem in both Longissimus thoracis et lumborum and in
Semimembranosus muscles; consumers rated ES samples higher on
tenderness, juiciness, flavour and overall rating. This study showed that ES of
alpaca carcasses is recommended as it significantly improved meat quality
attributes and reduced the incidence of lean alpaca carcasses experiencing cold
induced shortening.
The effects of early post mortem low voltage electrical stimulation (28 V,
60 Hz) on biochemical changes and on final tenderness in Longissimus
thoracis et lumborum muscle from donkey carcasses was studied (Polidori et
al., 2016). ES significantly accelerated the glycolytic process during the first
hours post mortem, with a faster pH fall at 1, 3 and 6 h post mortem, and a
reduction in ATP content at 3 and 6 h post mortem. Tenderness was
significantly improved (P < 0.05) at 4 and 7 days post mortem by the
application of ES.
Effects of ES on Meat Colour
ES is applied with the aim of ultimately improving meat tenderness, but

there have been reported some minor effects such as ES-mediated meat colour
changes. Meat colour is an important quality trait to monitor given consumer

decisions at the pint of purchase are influenced by meat colour more than any
other quality factor and consumers use the degree of discoloration as an
indication of freshness (Mancini & Hunt, 2005). It seems unlikely that
electricity on its own affects meat colour. The effect possibly arises because
the process depletes the metabolites of surviving oxidative pathways in the
muscle, or because the fast pH fall causes the muscle proteins from treated
carcasses to approach their isoelectric point much sooner, thereby “opening
up” the structure and easing oxygenation of myoglobin (Polidori et al., 1996).
The latter explanation seems at least partially responsible because, although
the cut surface from electrically stimulated sides develops a brighter red than
those from control sides when exposed to the air at 12-20 h post mortem, by
this time both electrically stimulated and control muscles would have attained
their ultimate pH, the structure in both would be “open” and residual oxygen
utilization of the same order (Lawrie, 1991).
In a study conducted by Toohey et al. (2008) in which was used a medium
voltage (300 V, 15 Hz) on 40 stimulated lambs, compared to 40 control lambs,
there was no significant difference between lightness L*, redness a* or
yellowness b* values in the two groups of animals. Similar results were found
in a study conduced by Channon et al. (2005) where it was determined that use
of ES had no effect on the meat colour of lambs.
A study in which the same voltage (300 V) has been used to stimulate 25
alpaca carcasses (Smith et al., 2016) found a significant difference (P<0.05) in
lightness L*, with meat samples from stimulated carcasses being lighter than
those from unstimulated carcasses. This increase in lightness was thought to be
primarily due to an effect of the increased glycolysis and pH decline occurring
due to ES. This trend in lightness agrees with other studies in which was
investigated the effects of low and high voltage ES on lamb carcasses, with the
result of an increase of L* values from the control of 28.0 to the value of 29.6
for low voltage and up to 31.4 for high voltage (Shaw et al., 2005).
Finally, a study performed on donkey carcasses electrically stimulated
using low voltage (28 V, 60 Hz) found a significant improvement (P<0.05) in
lightness L* determined in meat from stimulated carcasses (35.4) compared to
control group (33.2), while the difference in redness (a*) and yellowness (b*)
values was not statistically significant (Polidori et al., 2016). This increase in
lightness has been mainly attributed to the acceleration of the glycolytic
process due to electrical stimulation, with the faster pH decline and ATP
depletion occurred in stimulated donkey muscle.
CONCLUSION
ES is considered a valid method for accelerating the post mortem
biochemical changes in muscle. Thus, full rigor is established in stimulated
carcasses and the final pH is reached in their muscles within about 4 h from
stimulation, compared to 15-20 h in unstimulated carcasses. Industrial
application of this process to tenderize meat as well as facilitate the earlier
processing and utilization of meat through normal distribution channels has
made ES one of the most talked-about innovations in the meat industry.
Advantages for the packer, retailer, purveyor and consumer exist due to the
multiple benefits obtained when ES is used as an integral step in the process of
converting live animals to meat and meat products. ES has become one of the
most useful potential tools in meat technology, particularly in view of the
increasing tendency in commercial practice to cool carcasses as soon as
possible after slaughter.
The challenge for further development of ES systems is optimisation of
the activation of the enzyme systems, possibly by chilling regimes to ensure
rigor mortis close to 15°C, within the constraints of food safety concerns and
bearing in mind the different fibre composition of muscles. In this sense a
more targeted approach such as the use of local specific stimulation of certain
muscle groups, susceptible to rapid chilling effects, may be more beneficial for
improving tenderness.
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BIOGRAPHICAL SKETCH
Paolo Polidori
Affiliation: School of Pharmacy, University of Camerino, Italy

Education: Master Degree in Agriculture, University of Milan, Italy;
PhD in Animal Sciences, University of Milan, Italy.
Address: via Circonvallazione 93/95, 62024, Matelica (MC), Italy.
1. Protein profile characterization of donkey milk.

2. Nutritional characteristics of donkey milk.

3. Functional and nutraceutical foods.
 From November 1993 to October 2000: Researcher in Animal

Husbandry in the Faculty of Veterinary Medicine, University of
Camerino, Italy.
 From November 2000 to December 2005: Associate Professor in
Animal Husbandry in the Faculty of Veterinary Medicine, University
of Camerino, Italy.
 Since January 2006: Full Professor in Animal Nutrition in the School
of Pharmacy, University of Camerino, Italy
Honors:
 From September 1995 to March 1996: Visiting scientist at the Muscle

Biology Lab, University of Wisconsin, Madison, USA (six months
period, Italian National Research Council grant).
 From August 1998 to June 1999: Visiting scientist at the Department
of Food Science and Technology, University of Tennessee, Knoxville,
USA (11 months period, North Atlantic Treaty Organization grant).
 From January 2008 to December 2010: Member of the Editorial
Board of the journal Small Ruminant Research, Elsevier Publisher,
The Netherlands.
 Since March 2009: Member of Expert database of EFSA (European
Food Safety Agency), Parma, Italy.
 Since 2011: Member of the Editorial Board of the Open Journal of
Animal Science, SCIRP Publisher, USA.
 Since 2011: Member of the Editorial Board of the Journal of Food
Research, Canadian Centre of Science and Education, Canada.
 Since 2013: Member of the Editorial Board of the International
Journal of Child Health and Nutrition, LifeScience Global, Canada.
Peer Reviewed Publications Last 3 Years:
1. Use of Donkey Milk in Children with Cow’s Milk Protein Allergy.

Polidori P., Vincenzetti S.
Foods, 2013, 2, 151-159, doi: 10.3390/foods2020151
2. Meat Quality in Donkey Foals.

Italian Journal of Food Science, 2013, 25, 390-393
3. Effects of Thermal Treatments on Donkey Milk Nutritional
Characteristics.
Recent Patents on Food, Nutrition & Agriculture, 2013, 5, 182-187.
4. Carcass Characteristics, Meat Quality and Nutritional value of
Horsemeat: A review.
Lorenzo J.M., Sarriés M.V., Tateo A., Polidori P., Franco D., Lanza
M.
Meat Science, 2014, 96, 1478-1488.
5. Profile of Nucleosides and Nucleotides in Donkey’s Milk.
Vincenzetti S., Pucciarelli S., Nucci C., Polzonetti V., Cammertoni
N., Polidori P.
Nucleosides, Nucleotides and Nucleic Acids, 2014, 33, 656-667.
6. A comparison of the carcass and meat quality of Martina Franca
donkey foals aged 8 or 12 months.
Polidori P., Pucciarelli S. Ariani A., Polzonetti V., Vincenzetti S.
Meat Science, 2015, 106, 6-10.
7. Use of Donkey Milk in Cases of Cow’s Milk Protein Allergies.
Polidori P., Ariani A., Vincenzetti S.
International Journal of Child Health and Nutrition, 2015, 4, 174-
179.
8. The effects of low voltage electrical stimulation on donkey meat.
Polidori P., Ariani A., Micozzi D., Vincenzetti S.
Meat Science, 2016, 119, 160-164.
9. Comparative proteomic analysis of two clam species: Chamelea
gallina and Tapes philippinarum.
Vincenzetti S., Felici A., Ciarrocchi G., Pucciarelli S., Ricciutelli M.,
Ariani A., Polzonetti V., Polidori P.
Food Chemistry, 2017, 219, 223-229.
Book Chapters
 Effect of Dephosphorylation on Donkey Milk Caseins.

Vincenzetti S., Vita A., Carpi F.M., Micozzi D., Polidori P.
In: Trends in Veterinary Sciences, 2013, Eds. C. Boiti, A. Ferlazzo, A.

Gaiti, A. Pugliese, Springer-Verlag, Berlin, Germany, ISBN: 978-3-
642-36487-7, pp. 21-25.
 Oleic Acid in Milk of Different Mammalian Species.
In: Oleic Acid. Dietary Sources, Functions and Health Benefits, 2013,
Ed. Luciano P. Silva, Nova Science Publishers, Inc., New York, USA,
ISBN: 978-1-62618-332-2, pp. 127-140.
 Caseins Characteristics in Equid and Human Milk.
Vincenzetti S., Ariani A., Polidori P.
In: Caseins – Properties, Functions and Health Implications. 2016, Ed.
Laurence Mendoza, Nova Science Publishers, Inc., New York, USA,
ISBN: 978-1-63485-337-8, pp. 47-75.
 Farm Management and Feeding Strategies for Donkey Milk
Production.
In: Agricultural Research Updates, Volume 14. 2016, Eds. P.
Gorawala & S. Mandhatri, Nova Science Publishers, Inc., New York,
USA, ISBN: 978-1-53610-344-1, pp. 93-117.
Chapter 6
QUALITY AND NUTRITIONAL

CHARACTERISTICS OF DONKEY MEAT
Paolo Polidori1,* and Silvia Vincenzetti2

1
School of Pharmacy, University of Camerino, Italy
2
School of Biosciences and Veterinary Sciences,
University of Camerino, Italy
ABSTRACT
Meat has exerted a crucial role in human evolution and is an
important component of a healthy and well balanced diet due to its
nutritional richness. The aim of the present chapter is to shed light on the
nutritional composition of donkey meat and the implications for human
health. Donkeys are not perceived as multi-use animals. Cattle, buffaloes
and camels are usually kept for their milk and their meat as well as for
work. In many areas donkeys are not sold for their meat. One of many
exceptions is Lesotho where donkeys are culled for meat when they are
considered too old to work, and for this reason donkeys are relatively
expensive in this Country. In the rest of the world, the lower cost of
donkeys makes them more affordable to small farmers. On the other
hand, donkey meat can be considered a good alternative in red meat
consumption, being a dietary meat. Donkey meat is in fact characterized
by low fat, low cholesterol content, a favourable fatty acid profile and is
* Corresponding Author address, Email: paolo.polidori@unicam.it.

rich in iron. Today consumers are health conscious and demand high
quality food products; they require leaner meat, with less fat (the minimal
fat level required to maintain juiciness and flavour) and a consistent
quality. Ultimately, the success of any food product is determined by the
consumer’s acceptance. Meat quality and acceptability is determined by
its physico-chemical characteristics, although consumer preferences for
meat are difficult to define. In this context, this chapter will describe the
quality of donkey carcass and donkey meat quality parameters, showing
its chemical and sensorial characteristics (when possible in different
muscles) and evaluating the effects of the age of slaughtering.
Keywords: donkey meat, donkey carcass, meat quality, meat nutritional value
INTRODUCTION
Donkeys are said to have originated in north-east Africa and then spread
to other parts of the world. The world donkey population is about 44 million;
half is found in Asia, just over one quarter in Africa and the rest mainly in
Latin America. There are pictures of donkeys in the tombs of the Egyptian
pharaohs and 82 biblical references to donkeys. The ancient Romans used
donkeys for pack transport and agriculture.
Donkey (Equus asinus, Perissodactyla) is a domestic animal belonging to
the equine family, which includes horses, zebras and mules, and its progenitor
was the small gray donkey of northern Africa (Equus africanus) domesticated
around 4000 BC on the shores of the Mediterranean Sea. Donkey has been
domesticated for thousands of years (Aganga et al., 2003), and has contributed
to the development of various civilizations. Despite the increase in
mechanization throughout the world, donkeys have still an important role to
play in transport of people and goods in arid and semi-arid areas where roads
are poor or non-existent. Donkeys are therefore easy to manage and not too
demanding in terms of feeding. They can almost survive on poor quality feeds
and thrive under adverse climatic conditions, and they can also tolerate a
considerable heat and dehydration (Aganga et al., 1994).
Quality and Nutritional Characteristics of Donkey Meat 157
Figure 1. Amiata breed (Italy).
Donkeys are not perceived as multi-use animals. Cattle, buffaloes and

camels are usually kept for their milk and their meat as well as for work. In
many areas donkeys are not sold for their meat. One of many exceptions is
Lesotho where donkeys are culled for meat when they are considered too old
to work, and for this reason donkeys are relatively expensive in this Country
(Fernando & Starkey, 2000). In the rest of the world, the lower cost of
donkeys makes them more affordable to small farmers. On the other hand,
horse meat is considered a good alternative in red meat consumption, being
considered as a dietary meat, and this has led to an increase of their
consumption in recent years (Lorenzo et al., 2014). Horse meat is in fact
characterized by low fat, low cholesterol content, a favourable fatty acid
profile and is rich in iron (Franco et al. 2011). Today consumers are health
conscious and demand high quality food products; they require leaner meat,
with less fat (the minimal fat level required to maintain juiciness and flavour)
and a consistent quality. Ultimately, the success of any food product is
determined by the consumer’s acceptance. Meat quality and acceptability is
determined by its physico-chemical characteristics, although consumer
preferences for meat are difficult to define. Horse meat has a good reputation
for consumers (Belaunzaran et al., 2015); in this context, the aim of this
chapter is to assess the quality of donkey meat, showing its chemical and
sensorial characteristics (when possible in different muscles) and evaluating
the effects of the age of slaughtering.
DONKEY CARCASS
The composition of meat cannot be described simply in terms of the
different components and their percentages, since meat production involves
also the evaluation of the entire carcass, along with the muscles, fatty tissues,
bones, tendons, edible organs and glands. This obviously gives a wide range of
components and thus of composition and nutritive value. As firstly
demonstrated by Aganga et al. (2003), donkeys have a carcass yield in the
range between 54.5 to 59.5%, according to the different age, with the lowest
value obtained for the oldest animals, slaughtered at the age of 7 years, while
the highest value had been obtained with animals slaughtered at the age of 5
years. The dressing out percentage depends in fact upon the stage of maturity,
degree of finish, breed and the intestinal contents (offals). Donkeys as non-
ruminants have a slightly lower offal contents. Since they lack a rumen,
therefore, most of the digestion takes place in the caecum and the small
intestines. Basing on the data, as animals get older, their carcass yield declines.
This is due to the fact that fat, particularly subcutaneous fat is the least tissue
to mature, thus older animals tend to be fatter and the percentages of muscle
and bone decreasing progressively.
A study (Polidori et al., 2008) performed using 15 entire donkey males of
the Martina Franca breed, slaughtered at 15 months of age and a mean fasted
final body weight of 18137 kg, determined warm (98.7 kg) and cold (96.7
kg) carcass weights, shown in Table 1, together with warm (54.5%) and cold
(53.3%) dressing percentage. Obviously, comparisons with the dressing
percentages of horses can be made, but most of the horses used for meat
production belong to specialized breeds or to genetic types very different from
donkeys: in these studies carcass weight and dressing percentage were usually
higher compared than those obtained in donkeys.
Table 1. Donkey carcass characteristics (means±S.E.)
(n = 15)
Warm carcass weight (Kg) 98.71.22
Cold carcass weight (kg) 96.71.13
Warm dressing % 54.50.83
Cold dressing % 53.30.92
Source: Adapted from Polidori et al. (2008).
MEAT QUALITY
Donkeys can constitute an important source of meat in arid and semi-arid
areas, but in the past the potential of donkey breeding for meat production has
received little attention. In most of the countries where the donkeys are still
used as work animals or for milk production, they are slaughtered at an
advanced age, at the end of their useful working life. This age factor probably
accounts for the general opinion that donkey meat is unacceptably tough, and
in fact is mostly destined to be transformed into salami or other salted meat-
based products (Marino et al., 2015). However, not all male donkeys reared on
farms can be used for breeding, but meat from young males is an easy way to
obtain a cheap meat, and to increase the income of local farmers. Compared to
other livestock, donkey has an exceptional ability to survive under adverse
climatic conditions such as high temperatures, low rainfall and scarcity of
feed. Therefore, it offers an ideal animal for meat production in arid and semi-
arid regions of the world. Donkey is a good source of meat in areas where the
climate adversely affects other animal’s production efficiency. The aim of the
following sections is to describe the different quality parameters of donkey
meat, such as meat tenderness, chemical composition, mineral content and
fatty acid profile.
Figure 2. Catalan breed (Spain).
Meat Tenderness
Tenderness is probably the most important factor considered by the

consumer in assessing the eating quality of meat; without adequate tenderness,
consumers are unlikely to appreciate the more subtle characteristics of flavour
and juiciness. Two structural components determine the tenderness of meat.
The collagen of connective tissue has firstly been recognized as an important
influence on quality, and till the 1970’s was considered as the sole cause of
tenderness variation. The contractile apparatus, by contrast, was established as
a potential contributor to toughness only in the early 1960’s. This does not
mean that connective tissue has been relegated to play merely a minor role as a
toughening agent: in some circumstances toughness is due almost entirely to
collagen, whereas in others it is caused almost exclusively by the contractile
machinery (Marsh, 1977). There is a list of the factors known to influence
meat tenderness, they may be roughly divided into three groups:
 Factors which are determined before the birth of the animal (species,
breed, sex, etc.).
 Factors modified by management during life (age, animal feeding,
etc.)
 Factors affected by treatment after slaughter (ageing, cooking method,
etc.)
After an animal has been slaughtered, meat tenderness can be affected

only by the factors belonging to the third group, and depends mainly on the
variations in myofibrillar proteins, connective tissue proteins, water content
and state, as well as the interactions between the components during cooking
(Fabiansson & Reuterswärd, 1985).
In a study performed in order to evaluate donkey meat tenderness
(Polidori et al., 2011), 40 entire donkey males of the Martina Franca breed
were used. They were reared in an extensive pasture system in the same farm
in the south of Italy and receiving the same diet. Twenty animals (Group 1)
were slaughtered at 12 months of age and a mean fasted final body weight of
14827 kg. The other 20 animals (Group 2) were slaughtered at 18 months of
age and a mean fasted final body weight of 20245 kg. After 24 h storage in
the cold room (2°C), Longissimus thoracis muscle (LT) samples (weighting
approximately 400 g) were collected from the left side of each carcass,
between the 12th and the 13th rib. Samples were stored for two and for seven
days post slaughter before evaluating the shear force values. Two days after
slaughtering no significant differences were determined (Table 2) between
meat samples collected from donkeys slaughtered at 12 months of age
(6.25±0.53 kg/cm2) and from donkeys belonging to Group 2 (6.53±0.41
kg/cm2). At seven days post slaughter, the shear force values determined for
LT muscles obtained from donkeys belonging to Group 1 (5.15±0.31 kg/cm2)
were significantly lower (P<0.01) compared to the values registered in the
same group at two days. The shear force obtained seven days post slaughter in
Group 2 showed higher values (5.88±0.23 kg/cm2) compared with Group 1 but
significantly lower (P<0.05) with the results obtained in the same animals at
two days.
Table 2. Tenderness evaluation (kg/cm2) of donkey Longissimus thoracis

after two and seven days post mortem (means±S.E.)
Two days post mortem Mean S.E.

Group 1 (n = 20) 6.25a 0.53
Group 2 (n = 20) 6.53a 0.41
Seven days post mortem
Group 1 (n = 20) 5.15c 0.31
Group 2 (n = 20) 5.88b 0.23
Group 1: animals slaughtered at 12 months of age (n = 20)
Group 2: animals slaughtered at 18 months of age (n = 20)
Different letters in the column indicate significant difference (b: P<0.05; c: P<0.01)
Source: adapted from Polidori et al. (2011).
The results of this study demonstrated that, after an adequate ageing

period, donkey meat tenderness can be considered very similar to the same
quality parameter determined on horse meat, according also to the results
obtained in Sanfratellano and Halflinger foals by Lanza et al. (2009), and
confirming the hypothesis that an ageing period of seven days is considered
necessary to improve meat tenderness for most of the red meats (Thompson,
2002).
Intra-muscular collagen content and maturation are considered as primary
factors contributing to meat tenderness (Maiorano et al., 2001). Literature
reports differences in collagen properties among various muscles of the same
animals due to different muscle metabolism, structure, physiological function,
localisation and growth rate (Lawrie, 1985). A study conducted by Polidori &
Vincenzetti (2013) determined collagen concentration in muscles
Semimembranosus (SM) and Semitendinosus (ST) taken from 10 entire
donkey foals slaughtered at 10 months of age and with a mean final body
weight of 126 kg. Collagen content was significantly (P<0.01) higher in ST
muscle (44.2 g/mg) compared to SM muscle (32.1 g/mg). Studies
conducted on male and female horse foals showed higher values in total
collagen amount compared to donkey foals (Sarriés & Beriain, 2005). The
results obtained by Tateo et al. (2008) confirmed the significantly higher
statistical difference between total collagen content determined in ST muscle
compared to SM muscle taken from Italian Heavy Draft Horses.
Electrical stimulation has been extensively used since the 1950s to hasten
rigor mortis and to modify steps of the glycolytic pathway (Hwang et al.,
2003). It is generally accepted that the accelerated post mortem glycolysis

induced by electrical stimulation can prevent the development of toughness
caused by cold shortening (Fabiansson & Laser Reuterswärd, 1985; Polidori et
al., 1996; Devine et al., 2014). In order to determine whether a low voltage (28
V peak) early post mortem electrical stimulation, designed to accelerate
glycolysis and proteolysis, could significantly improve the tenderness of
donkey meat obtained from 16 entire male donkeys crossbred foals (Martina
Franca x Ragusana), slaughtered at 8 months of age and at a mean final body
weight of 10622 kg, a study (Polidori et al., 2016) demonstrated that use of
low voltage electrical stimulation in donkey carcasses can improve some meat
quality traits, reducing shear force values in electrically stimulated donkey
meat.
Meat Chemical Composition
Donkey meat has been historically obtained from animals that were
slaughtered at the end of their working lives; for this reason the meat usually
had not good sensorial and nutritional characteristics, as perceived by a lot of
consumers. Meat production from male foals can be an interesting source of
proteins and a cheap alternative to other red meats. A study had been
performed to evaluate the effects of different slaughter age on chemical
composition of donkey meat obtained from Martina Franca breed entire males
(Polidori et al., 2011). Twenty animals (Group 1) were slaughtered at 12
months of age and a mean fasted final body weight of 14827 kg. The other 20
animals (Group 2) were slaughtered at 18 months of age and a mean fasted
final body weight of 20245 kg. After 24 h storage in the cold room (2°C),
Longissimus thoracis muscle (LT) samples were collected in order to evaluate
chemical composition and glycogen content. Donkeys slaughtered at 12
months of age showed a significant (P<0.05) lower fat content, specifically
2.41 g/100 g compared with 3.71 g/100 g obtained in older animals (Table 3).
Cholesterol content was similar in both the groups of animals, 67.4 mg/100 g
in Group 1 and 68.7 mg/100 g in Group 2, confirming that cholesterol level in
meat is not strictly related to fat content (Lawrie 1985). Protein content
increased significantly (P<0.05) in donkeys slaughtered at 18 months of age,
showing a value of 22.3 g/100 g compared with 21.4 g/100 g obtained in
Group 1. Ash and glycogen content did not show significant differences
between the two groups. Moisture content was significantly lower (P<0.05) in
samples of LT collected from animals belonging to Group 2 (72.5 g/100g)

compared to Group 1 (74.8 g/100 g), due to the increase of protein and fat
content in older animals. Glycogen and cholesterol content were very close to
the values obtained in many studies conducted on horse meat (Badiani et al.,
1997; Lorenzo et al., 2014).
An evaluation of mineral levels in donkey meat samples was carried out
using 15 entire donkey males of the Martina Franca breed, slaughtered at 15
months of age and a mean fasted final body weight of 18137 kg (Polidori et
al., 2008). Potassium (343 mg/100 g) and phosphorus (212 mg/100 g) were the
two main represented minerals, followed by sodium (52 mg/100 g) and
magnesium (24 mg/100 g). Iron content (3.80 mg/100 g) was higher than zinc
content (3.67 mg/100 g): the levels of these micro elements, essential in the
human diet, were similar with those determined in beef meat (Lawrie, 1985).
The donkey meat had a calcium content of 8.6 mg/100 g. The mineral content
of donkey meat varied widely between animals (Table 4).
Table 3. Chemical characteristics (mean ± S.E.) of the Longissimus

thoracis muscle taken in 12 old months (Group 1) and in 18 old months
(Group 2) entire donkey males
Group MeanS.E.
Moisture (%) 1 74.8a3.31
2 72.5b2.21
Fat (%) 1 2.41a0.71
2 3.71b0.43
Protein (%) 1 21.4a2.95
2 22.3b3.01
Ash (%) 1 1.04a0.77
2 1.10b0.89
Glycogen (%) 1 0.36a0.03
2 0.42a0.07
Cholesterol (mg/100 g) 1 67.4a0.81
2 68.7a0.93
Different letters in the column indicate significant difference (b: P<0.05).
Table 4. Mean values (±S.E.) of minerals (mg/100 g) of Longissimus

Thoracis muscle of donkeys carcasses
Mineral Mean±S.E.
Calcium 8.65±2.13
Magnesium 24.8±6.71
Potassium 343.7±65.9
Phosphorus 212.9±56.7
Sodium 52.5±13.3
Zinc 3.67±0.78
Iron 3.80±1.01
Fatty Acid Composition
There is growing consensus that the dietary habits adopted by Western

societies over the past century have contributed to an increased risk of
coronary heart disease, hypertension, diabetes and cancer (Coates & Ayerza,
2004). Clinical data strongly support a relationship between the incidence of
coronary heart disease and consumption of cholesterol and saturated fatty
acids. Consumption of fish would provide a more balanced fatty acid diet, but
for many people increasing their intake of fish is not readily accepted because
of taste preferences. A study performed (Polidori et al., 2009) using 12 entire
donkey males of the Martina Franca breed, slaughtered at 14 months of age
with a mean fasted final body weight of 16913 kg, demonstrated that fatty
acid composition of the intramuscular fat determined on muscle Longissimus
Thoracis et Lumborum and on muscle Biceps Femoris did not show significant
differences between the two muscles examined, with the exception of myristic
acid (C14:0), that was significantly greater in the Biceps Femoris (Table 5).
An interesting content of polyunsaturated fatty acids (PUFA) was found in
both muscles, The two most abundant fatty acids in both muscles were oleic
acid (18:1cis9) and palmitic acid (C16:0); saturated fatty acid (SFA) content
was very similar in both muscles (Table 5).
According to the results shown in Table 5, donkey meat is characterised
by a high content of unsaturated fatty acids, particularly PUFA, some of which
can play important healthy roles, as described before in this chapter.
Meat Colour
In a study performed in order to determine rheological characteristics

(Polidori et al., 2009), meat samples were taken four days post mortem from
muscles Longissimus thoracis et lumborum (LTL) and Biceps femoris (BF)
from 12 Martina Franca donkey males, slaughtered at 14 months of age and a
mean final body weight of 169 kg. Colorimetric parameters were measured to
determine L* (lightness), a* (redness), b* (yellowness) and chroma. Results
showed a significantly (P<0.05) higher lightness (L*) and redness (a*) in the
LTL compared to the BF, respectively 35.86 versus 31.34 and 10.43 versus
9.23 (Table 6), indicating a darker colour of the LTL compared with the hind
leg muscle BF. Colour is considered the most important sensory attribute
affecting consumer purchasing decisions of red meat, because red colour is
associated with freshness. The L* values obtained indicated that donkey meat
colour can be considered very similar to other red meats, such as beef (Boakye
& Mittal, 1996), horse meat (Tateo et al., 2008) and lamb (Luciano et al.,
2009). Redness (a*) and yellowness (b*) indexes can be important in
evaluating meat quality when it is possible to determine changes over time,
because their changes describe meat colour deterioration from red to brown,
reflecting myoglobin concentration and its redox state in muscle (Mancini &
Hunt, 2005). In this study a* and b* values were determined only four days
after slaughter; in the future it will be important to follow changes during
storage. Chroma (Table 6) was significantly (P<0.05) different between the
LTL and BF, with values of respectively 10.46 and 9.26. Chroma describes the
intensity of a fundamental colour with respect to the amount of white light in
the background (Boakye & Mittal, 1996); the differences between the two
muscles demonstrated for all indexes evaluated a darker colour in the LTL
compared with the BF.
An experiment performed with the aim of determining the effect of two
different slaughter age (8 vs 12 months) on 16 entire donkey male foals of the
Martina Franca breed (Polidori et al., 2015) determined that the colorimetric
characteristics of donkey meat showed not significant differences in lightness
(L*) and redness (a*) in the LTL muscle collected from foals slaughtered at 8
and 12 months of age, respectively 33.57 versus 32.34 for L* and 12.24 versus
11.49 for a* (Table 7). Similarly, the age of the animals did not affect the
colorimetric characteristics, according to previous results obtained in horse
meat (Sarriés & Beriain, 2006), confirming therefore that feeding systems play
an important role in differentiating between meat samples on the basis of
colour. Meat colour is also influenced by the myoglobin content, and the
myoglobin content within a species varies with age (Lawrie, 1985). A possible
explanation of the results obtained in that study could be that haemoglobin
contents in LTL muscle did not differ between foals slaughtered at 8 and 12
months of age.
Table 5. Fatty acid composition (% total fatty acids) determined in

donkey Longissimus Thoracis and Biceps Femoris muscles (means±S.E.)
Fatty acid LT (n = 12) BF (n = 12)

a
C14:0 3.88±0.53 4.51±0.74b
C16:0 29.77±2.98 29.44±2.01
C16:1 3.16±0.64 3.78±0.58
C18:0 7.43±0.87 6.83±1.01
C18:1 29.65±3.23 29.54±2.88
C18:2 18.75±2.86 19.43±3.01
C18:3 4.32±0.89 3.89±0.76
C20:1 0.95±0.11 0.93±0.09
C20:4 2.09±0.37 1.65±0.29
SFA 41.08±2.02 40.78±1.91
MUFA 33.76±1.68 34.25±1.55
PUFA 25.16±1.21 24.97±1.01
SFA: saturated fatty acids
MUFA: monounsaturated fatty acids
PUFA: polyunsaturated fatty acids
Different letters in the same row indicate a significant difference (b: P<0.05)
Table 6. Colour parameters for the LTL and BF donkey muscles

(means±S.E.)
Parameter LTL BF
a
L* 35.86±1.49 31.34±1.63b
a
a* 10.43±0.38 9.23±0.43b
b* - 0.78±0.11 - 0.67±0.21
a
Chroma 10.46±0.66 9.26±0.58b
Different letters in the same row indicate a significant difference (b: P<0.05)
Table 7. Colour parameters (means±S.E.) of the LTL muscle of donkeys

slaughtered at different ages
Slaughter age (months)

8 12
(n = 8) (n = 8)
L* 33.57±2.94 32.34±2.36
a* 12.24±0.48 11.49±0.83
b* 8.76±0.22 7.87±0.13
Source: Adapted from Polidori et al. (2015).
Amino Acid Composition
Amino acid composition has been determined in donkey meat (Polidori et

al., 2009). The content of the different amino acids is shown in Table 8; both
the LTL and BF had higher essential amino acid percentages, respectively
52.88% in LTL and 51.26% in BF, compared with the total amino acid
contents. Arginine was included between the essential amino acids, as done by
Hoffman et al. (2005), because arginine is considered a conditionally essential
amino acid. The essential amino acids at the highest concentration in donkey
meat were lysine (1.77 g/100 g in LTL, 1.63 g/100 g in BF) and leucine (1.51
g/100 g in LTL, 1.60 g/100 g in BF), as shown in Table 8. No statistically
significant differences in amino acid contents were found between the
muscles. Values were very similar than those given by Badiani et al. (1997) in
a review on horse meat quality characteristics, while Lorenzo & Pateiro (2013)
found histidine as the most abundant amino acid in meat obtained by 12 foals
of Galician Mountain breed slaughtered at 15 months of age.
Evaluating the results shown in table 8, we can affirm that the nutritional
qualities of donkey meat are also confirmed by the high content of essential
amino acids, a factor that is very important in determining food quality. The
ratio between essential/non-essential amino acids was larger than 50% also in
another study (Polidori et al., 2015) performed on donkey foals slaughtered at
8 months of age (52.8%) and at 12 months of age (51.3%) confirming the high
biological values of donkey meat proteins.
A study conducted by Paleari et al. (2003) compared the amino acids
content determined in deer, boar, horse, beef and goat meat; beef meat had a
significant lower content (47.4%), while both deer and goat meat showed in
that study higher proportions of essential amino acids, respectively 58.8% and
54.8%. The results obtained for donkey meat can be considered close to horse
(54.5%) and boar meat (50.0%).
Table 8. Amino acid composition (g/100 g muscle) of LTL and BF muscles

(means±S.E.) of donkey
LTL BF
Essential
Arginine 1.44±0.18 1.38±0.21
Histidine 0.86±0.08 0.93±0.09
Isoleucine 1.05±0.11 0.99±0.14
Leucine 1.51±0.65 1.60±0.51
Lysine 1.77±0.34 1.63±0.48
Methionine 0.74±0.13 0.65±0.18
Phenylalanine 0.83±0.17 0.76±0.13
Threonine 0.88±0.14 0.91±0.13
Tryptophan 0.24±0.08 0.19±0.12
Valine 1.01±0.34 1.09±0.45
Non essential
Alanine 1.22±0.23 1.09±0.11
Aspartic acid 1.79±0.45 1.92±0.59
Cystine 0.22±0.05 0.18±0.05
Glutamine 3.09±1.25 3.26±1.58
Glycine 0.97±0.36 0.84±0.65
Proline 0.95±0.58 1.00±0.42
Serine 0.75±0.23 0.64±0.20
Tyrosine 0.59±0.17 0.70±0.29
Total 19.91 19.76

Essential AA (%) 10.33 (52.88%) 10.13 (51.26%)
DONKEY MEAT PRODUCTS

Due to the continuous evolution of the economic and social countries,
food demands have been transformed and modified. Cured, fermented and
dried meat products from different species (Paleari et al., 2003) have recently
appeared on the market and are being sold alongside traditional beef and pork
products. The production of typical processed meat products could be a tool to
increase the value of donkey meat. Traditional salting, fermenting and drying
technologies have been used since ancient times to produce dry cured meat
products available throughout the year and consumed in many countries
(Marino et al., 2015).
Salami is a very popular fermented meat product, its quality depends on
the variations in raw meat, formulation and manufacturing processes. Among
cured meat products, bresaola is a product originated in different areas of north
Italy since the 15th century as a way of preserving both beef and horse meat
(known as “slinzega”). Products similar to bresaola are consumed in Brazil
(known as “charqui,” “manta de carne de sl”), in Spain (known as “cecinas” or
“cecina de leon,” Canton of Grisons, Switzerland (known as “Viandes de
Grison” or “Bindenfleisch”), in France, Doubs region (known as “Bresi”) and
in Peru (known as “charqui” and produced with both lama and alpaca meat).
A study performed with the aim to evaluate the nutritional properties of
bresaola and salami from donkey meat compared with respective conventional
products (Marino et al., 2015) demonstrated that donkey bresaola and salami
showed higher content of protein and lower content of fat compared with beef
bresaola and pork salami. Significant differences in unsaturation level of fatty
acids were found in this study, particularly, donkey meat products showed
lower saturated fatty acids and higher polyunsaturated fatty acid content.
Furthermore, donkey meat products, especially bresaola, showed the highest
content of essential amino acids. Both donkey meat products resulted to be
more tender than conventional products, in addition donkey bresaola showed
also higher consumer acceptability. Basically, this study demonstrated the
possibility of processing donkey meat into products comparable to traditional
ones with a high nutritional value.
Figure 3. Donkey butchery in China.
CONCLUSION
Donkey, as a domestic animal, could be bred not only for leisure activities
or for working as a draught animal, but also for donkey meat production, being
this kind of meat a quite popular food in China (see Figure 3) or in south
America. Donkey meat production under sustainable extensive systems should
be encouraged in order to maintain endangered local donkey breeds, to obtain
a healthy product and finally to conserve natural resources (mountain areas),
as grazing could provide greater diversity of habitats benefiting fauna and
flora that bring environmental and social advantages to rural areas. Moreover,
donkey meat production could be managed as short productions chains
improving the local autochthonous breeds reared according to low input
production systems by applying feeding strategies to improve quality
standards either in fresh meat or meat products. Production of donkey meat
shows a very important potential to be considered in the economical
development of many countries; there is a big challenge for the farmers to use
donkeys as meat animals, not only for the aspects related to the breeding
development for these animals, but especially in the rational productive aspect.
This approach can be very useful economically for many donkey farmers all
over the world.
The nutritional characteristics of donkey meat show interesting aspects in

comparison to the usual red meat; when related to the human health
parameters, this kind of meat can be favourably accepted by the consumers. In
fact, donkey meat presents low levels of lipids and cholesterol, and shows a
beneficial relationship between the different fatty acids.
The studies performed till now determined that donkey meat is good
quality meat; data obtained indicates that the meat is very high in crude
protein, characterized by low fat content and high in important minerals such
as Potassium, Phosphorus, Iron and Zinc.
Donkey, especially males, can be used as cheap meat animals, and donkey
meat can easily have a market also in the western countries, considering the
high quality level shown by this kind of red meat.
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Lanza, M., Landi, C., Scerra, M., Galofaro, V., & Pennisi, P. (2009). Meat
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BIOGRAPHICAL SKETCH
Paolo Polidori
Affiliation: School of Pharmacy, University of Camerino, Italy

Education: Master Degree in Agriculture, University of Milan, Italy;

PhD in Animal Sciences, University of Milan, Italy.
Address: via Circonvallazione 93/95, 62024, Matelica (MC), Italy.
1. Nutritional properties of alternative red meats.

2. Nutritional characteristics of donkey milk.
3. Functional and nutraceutical foods.
 From November 1993 to October 2000: Researcher in Animal

Husbandry in the Faculty of Veterinary Medicine, University of
Camerino, Italy.
 From November 2000 to December 2005: Associate Professor in
Animal Husbandry in the Faculty of Veterinary Medicine, University
of Camerino, Italy.
 Since January 2006: Full Professor in Animal Nutrition in the School
of Pharmacy, University of Camerino, Italy
Honors:
 From September 1995 to March 1996: Visiting scientist at the Muscle

Biology Lab, University of Wisconsin, Madison, USA (six months
period, Italian National Research Council grant).
 From August 1998 to June 1999: Visiting scientist at the Department
of Food Science and Technology, University of Tennessee, Knoxville,
USA (11 months period, North Atlantic Treaty Organization grant).
 From January 2008 to December 2010: Member of the Editorial
Board of the journal Small Ruminant Research, Elsevier Publisher,
The Netherlands.
 Since March 2009: Member of Expert database of EFSA (European
Food Safety Agency), Parma, Italy.
 Since 2011: Member of the Editorial Board of the Open Journal of
Animal Science, SCIRP Publisher, USA.
 Since 2011: Member of the Editorial Board of the Journal of Food

Research, Canadian Centre of Science and Education, Canada.
 Since 2013: Member of the Editorial Board of the International
Journal of Child Health and Nutrition, LifeScience Global, Canada.
Peer Reviewed Publications Last 4 Years:
1. Use of Donkey Milk in Children with Cow’s Milk Protein Allergy.

Foods, 2013, 2, 151-159, doi: 10.3390/foods2020151
2. Meat Quality in Donkey Foals.

Italian Journal of Food Science, 2013, 25, 390-393
3. Effects of Thermal Treatments on Donkey Milk Nutritional

Characteristics.
Recent Patents on Food, Nutrition & Agriculture, 2013, 5, 182-187.
4. Carcass Characteristics, Meat Quality and Nutritional value of

Horsemeat: A review.
Lorenzo J.M., Sarriés M.V., Tateo A., Polidori P., Franco D., Lanza M.
Meat Science, 2014, 96, 1478-1488.
5. Profile of Nucleosides and Nucleotides in Donkey’s Milk.

Vincenzetti S., Pucciarelli S., Nucci C., Polzonetti V., Cammertoni N.,
Polidori P.
Nucleosides, Nucleotides and Nucleic Acids, 2014, 33, 656-667.
6. A comparison of the carcass and meat quality of Martina Franca

donkey foals aged 8 or 12 months.
Polidori P., Pucciarelli S. Ariani A., Polzonetti V., Vincenzetti S.
Meat Science, 2015, 106, 6-10.
7. Use of Donkey Milk in Cases of Cow’s Milk Protein Allergies.

Polidori P., Ariani A., Vincenzetti S.
International Journal of Child Health and Nutrition, 2015, 4, 174-179.
8. The effects of low voltage electrical stimulation on donkey meat.

Polidori P., Ariani A., Micozzi D., Vincenzetti S.
Meat Science, 2016, 119, 160-164.
9. Comparative proteomic analysis of two clam species: Chamelea

gallina and Tapes philippinarum.
Vincenzetti S., Felici A., Ciarrocchi G., Pucciarelli S., Ricciutelli M.,
Ariani A., Polzonetti V., Polidori P.
Food Chemistry, 2017, 219, 223-229.
Book Chapters
 Effect of Dephosphorylation on Donkey Milk Caseins.

Vincenzetti S., Vita A., Carpi F.M., Micozzi D., Polidori P.
In: Trends in Veterinary Sciences, 2013, Eds. C. Boiti, A. Ferlazzo, A.
Gaiti, A. Pugliese, Springer-Verlag, Berlin, Germany, ISBN: 978-3-642-
36487-7, pp. 21-25.
 Oleic Acid in Milk of Different Mammalian Species.

In: Oleic Acid. Dietary Sources, Functions and Health Benefits, 2013, Ed.
Luciano P. Silva, Nova Science Publishers, Inc., New York, USA, ISBN: 978-
1-62618-332-2, pp. 127-140.
 Caseins Characteristics in Equid and Human Milk.

Vincenzetti S., Ariani A., Polidori P.
In: Caseins – Properties, Functions and Health Implications. 2016, Ed.
Laurence Mendoza, Nova Science Publishers, Inc., New York, USA, ISBN:
978-1-63485-337-8, pp. 47-75.
 Farm Management and Feeding Strategies for Donkey Milk

Production.
In: Agricultural Research Updates, Volume 14. 2016, Eds. P. Gorawala &
S. Mandhatri, Nova Science Publishers, Inc., New York, USA, ISBN: 978-1-
53610-344-1, pp. 93-117.
Chapter 7
NITRITES/NITRATES IN PROCESSED MEAT:

RISKS AND BENEFITS
Małgorzata Karwowska* and Anna Kononiuk

Department of Meat Technology and Food Quality, University of Life
Sciences in Lublin, Lublin, Poland
ABSTRACT
High intake of processed meat has been associated with increased
risk of many diseases. Some additives used in processed meat production,
especially nitrites/nitrates, are of concern. Nitrite has been in the spotlight
for decades because of its involvement in the formation of nitroso-
compounds, such as carcinogenic N-nitrosoamines. Particularly red meat
and meat products, which are a good source of heme iron, have been
related to increased risk of cancer. For this reason there has been pressure
on the consumer side to eliminate the use of nitrites/nitrates in meat
product formulations. On the other hand, the elimination of
nitrites/nitrates from the production of meat products is problematic as
their use contributes to color development, flavor, antioxidant properties
and microbiological stability. Nitrites and/or nitrates are used to improve
meat product safety as they prevent the growth of most pathogenic and
spoilage organisms, including Clostridium botulinum and Staphylococcus
aureus. Moreover, nitrite retards oxidative rancidity during storage of the
meat product. During the curing process, nitrite is converted to nitric
*
Corresponding author: Małgorzata Karwowska. Address: ul. Skromna 8, 20-704 Lublin, Poland.
Email: malgorzata.karwowska@up.lublin.pl.
180 Małgorzata Karwowska and Anna Kononiuk
oxide (NO) via reduction reactions by curing adjuncts (sodium ascorbate,

sometimes bacteria), which may act as a bacteriocidal agent by blocking
the thiol group that comprises the active center of nonheme iron-sulfur
proteins. However, added nitrite is usually not completely degraded to
NO during the curing process, and excess amounts of residual nitrite have
been associated with the formation of carcinogenic nitrosamines.
In our previous studies, acid whey was investigated for its potential
use as a substitute for nitrites/nitrates in meat products. The obtained
results showed that acid whey had a positive effect on the
physicochemical qualities of non-nitrite meat products. In this context,
the aim of this paper is to present both the benefits and risks of
nitrites/nitrates as well as the possibilities of using acid whey as an
alternative to nitrites/nitrates in meat products.
Keywords: nitrites/nitrates, processed meat
INTRODUCTION
The health effects of the dietary consumption of meat and meat products
have been attributed to meat’s constituents, including high biological value
protein, iron, and vitamin B12 as well as other B complex vitamins, zinc,
selenium and phosphorus (Pereira and Vincente, 2013). As reported by
Williams (2007), red meat provides ca. 25% of the recommended dietary
intake of riboflavin, niacin, vitamin B6 and pantothenic acid per 100 g and
two-thirds of the daily requirement of vitamin B12 per 100 g. According to
Watanabe (2007), animal foods are the major dietary sources of vitamin B12.
Vitamin B12 deficiency is associated with megaloblastic anemia and a high
level of blood homocysteine, which is a cardiovascular disease risk factor, and
with depressive symptoms and neurological impairment (Green and Miller,
2005; Agarwal, 2011). Despite the health benefits of meat in the diet, high
intake of processed meat has been associated with increased risk of many
diseases. Several studies have associated processed meat and red red intake
with increased all-cause mortality (Larsson and Orsini, 2014), increased risk of
stroke (Chen et al., 2013) and type 2 diabetes (Aune et al., 2009). The
epidemiological studies reviewed by Abid et al. (2014) provide data to support
the role of red and processed meat in colorectal cancer and some evidence for
other cancer sites, including the liver, kidneys and, the prostate. Heme iron has
been indicated as one of the meat compounds associated with the presence of
Nitrites/Nitrates in Processed Meat 181
cancer. Very high iron intake is associated with increased risk of colorectal
cancer (Balder et al., 2006). Moreover, some additives used in processed meat
production, especially nitrites/nitrates, are of concern. Other authors have
presented conflicting results and conclusions (Alexander et al., 2011;
Alexander et al., 2015).
The International Agency for Research on Cancer (IARC) classifies
nitrates and nitrites as “probably carcinogenic to humans” (Group 2A) under
certain conditions (i.e., ingested nitrate or nitrite under conditions that result in
endogenous nitrosation) which could lead to the formation of known
carcinogens such as N-nitroso compounds (IARC, 2010). In 2015, the IARC
declared processed meat (meat that has been transformed through such
processes as salting, curing, fermentation, smoking) to be a Group 1
carcinogen based on data related to colorectal and stomach cancer. Moreover,
red meat (mammalian muscle meat including beef, veal, pork, lamb, mutton,
horse and goat) consumption was classified as probably carcinogenic to
humans (Bouvard et al., 2015). In this context, recent research has focused on
finding alternatives for nitrates/nitrites in meat processing. Natural curing
without the addition of nitrites/nitrates is possible thanks to the specific
properties certain starter cultures possess as well as with the use of natural
juices, dried fruit and vegetable concentrates. Nitrates are widely found in
plants and soil as they are a part of the nitrogen cycle in nature (O’Donnell,
2009). High levels of nitrates have been found in celery, thus it is used in the
“natural curing” of meat (Sebranek and Bacus, 2007), in which a natural
source of nitrates should be used in combination with nitrate- reducing
bacterial cultures, e.g., Staphylococcus carnosus (Szymański and Kołożyn-
Krajewska, 2014).
In our previous studies, liquid acid whey obtained from cottage cheese
production was investigated for its potential use as a substitute for
nitrites/nitrates in meat products. The obtained results indicated that acid whey
had a positive effect on the quality of nitrite-free meat products (Karwowska et
al., 2014; Karwowska et al., 2015; Karwowska and Dolatowski, 2017).
In reference to the above, the goal of this paper was to summarize the
recent literature on the benefits and risks of using nitrites/nitrates and on the
possibilities of using acid whey as an alternative to nitrites/nitrates in meat
products.
SODIUM NITRITE/NITRATE IN PROCESSED MEAT

Nitrites (sodium nitrite - E249, potassium nitrite - E250) and nitrates
(sodium nitrate - E251, potassium nitrate - E252) are approved food additives
in the European Union. Commission Regulation (EU) No 1129/2011
determines the maximum amount of nitrites and nitrates that may be added
during meat processing as meat preservatives. The amount of nitrite permitted
for use as a food additive in cured meat is currently 150 mg kg-1, with the
exception of sterilized meat products for which the limit was set at 100 mg kg-
1. The addition of nitrate (V) is allowed only in uncooked meat, up to 150 mg
kg-1. Other detailed requirements for higher maximum levels for nitrates have
been specified for some traditional meat products.
Curing is a conservation technique that is widely used in meat processing
to prolong the shelf-life of meat products (Bázan-Lugo et al., 2012; Marco et
al., 2006). Curing meat means adding either nitrite or nitrate, or both, together
with salt to the meat. Nitrate becomes an active curing agent only after
reduction to nitrite. The reduction of nitrates to nitrites is carried out mainly by
bacteria possessing nitrate reductase activity (staphylococci and micrococci),
which are naturally present in meat or added during processing (Hammes,
2012). In acid conditions of meat, generally pH 5.6-5.8, nitrites are converted
to a variety of compounds such as nitrous acid (HNO2), nitric oxide (NO) and
nitrates (Honikel, 2004). The use of additives in cured meat batters such as
ascorbic acid or ascorbate helps to decrease the number of nitrites in meat
products. As reported by Honikel (2008), the action of ascorbate is based on
reaction with oxygen, by forming dehydroascorbate, and reduces the amount
of nitrites that could be oxidized to nitrates. The formation of nitric oxide from
nitrites is a prerequisite step for reactions in the curing process. The decrease
in the amount of nitric oxide is due to its reactions with myoglobin and other
substrates in meat, including amino acids such as cysteine (Honikel, 2008).
Circa 10-20% of originally added nitrites, referred to as residual nitrite, is
typically present in meat products after production. This amount of residual
nitrite slowly declines during the storage period of cured meat products
(Sindelar and Milkowski, 2012).
The beneficial effects of nitrites/nitrates in cured meat products is related
to their functionality. The addition of nitrate/nitrite during curing processes
shows the positive effect of color enhancement in meat products. Via a series
of reactions, nitric oxide is bound to the iron ion located in the center of the
porphyrin ring system of myoglobin and forms nitrosomyoglobin. Upon
heating, this pigment forms the more stable red pigment nitroso-chromogen, of
cured meat. The denaturated form of nitrosomyoglobin can be created either
via the application of heat or in the low-acid environment of raw fermented
meat products. Sebranek and Bacus (2007) reports that the addition of ascorbic
acid can increase the conversion of nitrite to nitric oxide and thus speed up the
reaction of nitric oxide with myoglobin.
The role of nitrite/nitrate in the development of flavor typical of cured
meat is not fully understood. As reported by Jira (2004), several compounds
are formed when nitrite is bound to proteins and lipids. Since nitrites are
bound to sulfur- containing amino acids of meat proteins, SH-residues with a
specific aroma and flavor are formed and contribute to the flavor of cured
meat products. However, isolation of cured meat’s flavor compounds is
difficult (Sindelar and Milkowski, 2012).
The antimicrobial role of nitrites/nitrates in cured meat products has been
well documented. They contribute to meat product safety as they inhibit the
growth of Clostridium botulinum and thereby the formation of a neurotoxic
compound that is known as botulinum toxin (Xi et al., 2011). The
antimicrobial activity of nitrites/nitrates in cured meat products is likely
attributed to reactions associated with the generation of nitric oxide or nitrous
acid from nitrites (Møller and Skibsted, 2002). The activity is additionally
reinforced by other antimicrobial hurdles including heat treatment, a low- acid
environment, salt and other ingredients that are added during the curing
process, water activity and redox potential (Tompkin, 2005).
Nitrite acts against lipid oxidation primarily due to oxygen deletion. The
nitric oxide molecule can be oxidized to form NO2 causing oxygen
sequestering (Honikel, 2008). The oxidation of meat lipids is inhibited in such
conditions. Moreover, nitrite acts against lipid oxidation through the binding
of heme and by preventing the release of catalytic iron. Consequently, free
iron ions are not available for the initiation of the lipid oxidation process
(Andrée et al., 2010). Furthermore, nitrites are also able to bind heme and non-
heme iron and stabilize polyunsaturated fatty acids forming nitro-nitroso
derivatives (Weiss et al., 2010). The cell membranes are thus protected against
lipid peroxidation. Sebranek and Bacus (2007) indicated that nitrite levels
down to 50 ppm can have an antioxidant effect reducing TBA values by up to
68% for pork, chicken and beef.
HEALTH RISK AND BENEFITS WITH THE USE

OF NITRATE/NITRITE
Exposure to nitrate/nitrite may come from internal nitrate production as

well as from external sources, mainly from food (80%) (vegetables, fruits and
meat products) and water (14%) (WHO, 2007). According to Larsson et al.
(2011), approximately 98% of nitrate intake from food is attributed to
vegetables and fruits, while only 2% comes from cured meat products. In
addition to dietary intake, nitrates are formed endogenously. In mammals, the
main source of nitrates formed endogenously is the L-arginine-NO synthesis
pathway which is active in numerous cell types. Nitric oxide is produced from
L-arginine and molecular oxygen by nitric oxide synthase (Lundberg et al.,
2009). In contrast to nitrates, nitrites are mainly formed from the endogenous
bioconversion of dietary nitrates to nitrites in saliva (Govari and Pexara,
2015).
The intake of a certain amount of nitrates comprises a part of the nitrogen
cycle in the human body. The physiological disposition of nitrate, nitrite and
nitric oxide (NO) from dietary and endogenous sources includes: 1) the action
of bacterial nitrate reductases on the tongue and mammalian enzymes that
have nitrate reductase activity in tissues; 2) bacterial nitrate reductases; and 3)
mammalian enzymes with nitrite reductase activity (Hord et al. 2009).
Nitrate can be reduced to nitrite and nitric oxide when needed
physiologically or as a part of pathological processes depending on local
conditions, e.g., inflammation and tissue oxygenation. In vivo conversion of
nitrates to nitrites significantly enhances the nitrates’ toxic potency. The main
effect of nitrite related to toxicity is methemoglobin in infants, usually as a
consequence of excessive nitrite exposure (WHO, 2007). A case of acute
human intoxication by nitrites has been described by Matteucci et al. (2008).
Intoxication by nitrites in a woman and her son took place after they had
consumed turkey meat that was contaminated with a very high level of nitrite
(ranging from 6,000 to 10,000 mg kg-1 (Matteucci et al., 2008).
The long-term toxicity of nitrites/nitrates in food is associated with
increased risk of cancer. The relationship between nitrite consumption and
gastrointestinal cancers is associated with findings that ingested nitrites may
react with secondary amines or N-alkylamides to generate carcinogenic N-
nitroso compounds (Mensinga et al., 2003). This group of nitrosamines
includes volatile and non- volatile nitrosamines; the majority of the first group
are known to be carcinogenic. According to the classification of carcinogenic

compounds, N-nitrosodimethylamine (NDMA), and N-nitrosodiethylamine
(NDEA) belong to the group of probable carcinogens, whereas N-
nitrosodibutylamine (NDBA), N-nitrosopiperidine (NPIP) and N-
nitrosopyrrolidine (NPYR) belong to the group of possible carcinogens
(Yurchenko and Mölder, 2005). Karovičová and Kohajdová (2005) claimed
that putrescine (PUT) from ornithine, cadaverine (CAD) from lysine, and
spermine (SPM) and spermidine (SPD) from putrescine can be the precursors
of carcinogenic N-nitrosamines. Wei et al. (2009) suggested that NPIP can be
formed from cadaverine. As pointed by Andrée et al. (2010) nitrosoamines
formation is favored under specific conditions including presence of secondary
of amines, low pH (less than 6.55) and thermal processing at high temperature
(usually >130C) or long storage period at room temperature.
When reviewing the toxicological effects of nitrates and nitrites, a
committee of FAO / WHO experts determined that the acceptable daily intake
(ADI) for nitrate (V) should not exceed 3.7 mg kg-1, and in the case of nitrates
(III) it should not exceed 0.07 mg kg-1 (WHO, 2011). According to recent
studies, nitric oxide (NO) affects a number of key physiological processes in
the human body. Dietary consumption of nitrates has been shown to have
numerous health benefits, mainly related to improved cardiovascular function
(Bedale et al., 2016). Additionally, nitrites are associated with lower blood
pressure (Lundberg et al., 2006).
According to Lundberg et al. (2008) nitrate from food may be paramount
in the future of cardiovascular medicine by allowing generation of NO to
localized areas experiencing oxygen deficiency and blood deficiency
(ischemia). Studies have demonstrated the beneficial effects of short-term
dietary nitrite/nitrate supplementation on insulin resistant (Ohtake et al., 2015).
Their results suggest that nitrite from dietary sources provides an alternative
source of nitric oxide and subsequently improves the insulin-mediated
signaling, the metabolic and histological features in diabetic mice. Porcelli et
al. (2015) carried out the research whose purpose was to evaluate the effects of
short-term nitrate supplementation on endurance performance in subjects with
different levels of aerobic fitness. The results of this study suggest that the
individual aerobic fitness level affects the ergogenic benefits induced by
dietary nitrate supplementation.
AN ALTERNATIVE TO NITRITES/NITRATES
IN MEAT PRODUCTS
Due to the potential health effects of nitrite, there has been strong pressure
to either decrease or eliminate the use of nitrite in the curing process in order
to reduce the risk of nitrosamine formation and thereby any potential health
risks (Sindelar and Milkowski, 2012).
In the process of natural curing without the addition of nitrites/nitrates,
natural sources of nitrate (natural juices, dried fruit and vegetable
concentrates) and a starter culture with nitrate reductase activity to
subsequently produce nitrite, e.g., Staphylococcus carnosus, are used. Nitrates
are widely found in plants as they are a part of the nitrogen cycle in nature
(O’Donnell, 2009). In plant material, the highest level of nitrate is generally
found in leafy vegetables. High levels of nitrate have been found in celery,
lettuce and beets (Sebranek and Bacus, 2007). The most common ingredient
used to manufacture “uncured’ meat products has been unrefined sea salt
derived directly from the evaporation of sea water without the addition of free-
flow additives and retaining natural trace minerals (Heinerman & Anderson,
2001). Sebranek and Bacus (2007) reviewed the production of naturally cured
products in which ingredients that are high in nitrate (sea salt, celery
juice/powder) and starter culture with nitrate reductase activity are used to
subsequently produce nitrite and cured meat products without directly adding
sodium nitrite. As reported by Sebranek and Bacus (2007), Mediterranean Sea
salt contains 1.1 ppm of nitrate and 1.2 ppm of nitrite. Sindelar et al. (2007)
applied vegetable juice powder and starter culture as a nitrite replacer in
cooked frankfurter-style sausage. They concluded that meat products without
nitrate/nitrite addition can be manufactured with vegetable juice powder and a
starter culture containing Staphylococcus carnosus, with quality and sensory
attributes similar to traditionally cured products.
In our previous studies, acid whey was investigated for its potential use as
a substitute for nitrite/nitrate in meat products (Table 1). The obtained results
showed that acid whey had a positive effect on the physicochemical qualities
of non-nitrite meat products. The results obtained by Wójciak et al. (2014,
2015) showed that sausage samples with acid whey had overall sensory quality
that was similar to controls with curing salt. It is worth noting that the color of
the sausage was assessed as high in the sample in which a curing mixture was
used as in the nitrite-free sample with acid whey addition.
Table 1. Use of acid whey in the production of nitrite/nitrate-free

meat products
Description of texts analyzed Reference

The effect of acid whey and freeze-dried cranberries (used (Karwowska and
separately and in combination) on physicochemical characteristics, Dolatowski,
lipid oxidation and fatty acid composition of nitrite-free fermented 2017)
sausage made from deer meat and pork fat
The influence of natural preservatives, rosemary, juniper berries (Wójciak and
and mustard seed extract in combination with acid whey in the Dolatowski,
production of organic fermented sausage as an alternative to 2016)
nitrite/nitrate
The effect of acid whey with a plant extract (mustard, rosemary, (Wójciak and
juniper berry) combination on the shelf-life of organic roast pork Dolatowski,
2016)
The effect of marination in acid whey on selected pathogenic (Wójciak and
bacteria, N-nitrosamines, biogenic amines, amino acids, TBARS Solska, 2016)
value changes in fermented beef during ripening
The effect of acid whey on the physicochemical properties of non- (Stadnik and
nitrite organic dry-cured pork loin during chilling storage Stasiak, 2016)
The effect of acid whey and probiotic strains (Lactobacillus casei (Wójciak et al.,
LOCK 0900, L. casei LOCK 0908 and L. paracasei LOCK 0919) 2015)
on microbiological stability and sensory acceptance of organic
fermented sausage
The effect of acid whey on instrumental color value (CIE L*, a*, (Wójciak and
b*), total heme pigment and heme iron concentration in uncured Dolatowski,
fermented sausage 2015)
The influence of adding acid whey and mustard seed on oxidation (Karwowska et
stability (conjugated dienes and secondary products of lipid al., 2015)
oxidation, changes in fatty acid composition and antioxidant
capacity) of nitrite-free organic fermented sausage during 90 days
of vacuum storage
The effect of acid whey and two unusual starter cultures, i.e., three (Wójciak et al.,
probiotic strains (Lactobacillus casei LOCK 0900, Lactobacillus 2015)
casei LOCK 0908 and Lactobacillus paracasei LOCK 0919), on
model fermented sausage-type products by focusing on oxidative
stability via measuring instrumental color (L*, a*, b* values),
conjugated dienes and TBARS
The influence of adding acid whey with a combination of native (Karwowska et
and autoclaved mustard seed on the stability of organic model al., 2014)
sausages during 30 days of vacuum storage (by measuring the
primary and secondary products of lipid oxidation, changes in the
fatty acid composition, oxidation-reduction potential and
antioxidant capacity)
The elimination of nitrates is crucial in organic meat products as an

important principle of organic production is that producers use a minimum of
chemical substances during food production. In order to reproduce the
properties of nitrite/nitrate in organic sausage, sea salt and acid whey were
tested in combination with mustard seed (Karwowska et al., 2014, Karwowska
et al., 2015). The obtained results showed that these ingredients when used
together had a positive effect on the physicochemical qualities of non-nitrite
sausages. Karwowska et al. (2014) concluded that the use of acid whey and
mustard seed may offer meat processors the opportunity to develop organic
nitrite- free meat products with improved oxidation stability during storage.
Acid whey has been suggested to be a ‘natural’ antioxidant thanks to its ability
to inactivate pro-oxidative heme proteins and to chelate pro-oxidant transition
metals by β-lactoglobulin and lactoferrin, respectively (Colbert and Decker,
1991).
Since the basic problem in the production of nitrite-free meat products is
the formation of chemical and microbiological toxicants, studies have been
conducted to determine selected pathogenic bacteria, biogenic amines and N-
nitrosamine contents in fermented beef subjected to extended aging (Wójciak
and Solska, 2016). The highest concentration of total biogenic amine was
found in fermented beef marinated with acid whey (ca. 5 times higher as
compared to a cured sample), whereas no N-nitrosamines were detected in any
of the aged beef samples.
CONCLUSION
Nitrites/nitrates are important additives in meat processing due to their
functionality and beneficial effect on the quality of meat, and particularly on
the safety of meat products. Dietary nitrite at low doses has been proved as
beneficial to human health; unfortunately, despite the proven positive effects
on human health, a major concern of nitrates/nitrites in cured meat products is
related to their potential to form N-nitroso compounds, yet the numerous
functions of nitrates make it difficult to replace them with one component.
However, developing technologies, the selection of appropriate process
parameters and combinations of natural components possessing both
antioxidant and antimicrobial activity can lead to an elimination or significant
reduction in the use of nitrates in meat processing.
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INDEX
amines, 7, 8, 12, 31, 33, 38, 52, 69, 83, 84,

A 85, 86, 98, 100, 108, 184, 187, 188, 192,
193
abstraction, 45, 49
amino, viii, 2, 43, 45, 46, 48, 49, 50, 51, 52,
access, 3, 112
94, 95, 99, 101, 108, 113, 168, 170, 173,
acid, vii, ix, xi, xii, 8, 9, 10, 11, 12, 13, 16,
182, 183, 187, 193
17, 18, 19, 20, 21, 22, 23, 25, 29, 30, 31,
amino acid, viii, 2, 43, 45, 46, 49, 50, 51,
32, 33, 34, 35, 37, 38, 40, 44, 45, 48, 55,
52, 94, 95, 99, 101, 108, 113, 168, 170,
56, 57, 58, 60, 61, 68, 74, 75, 76, 78, 84,
173, 182, 183, 187, 193
88, 91, 95, 96, 97, 99, 100, 102, 103,
Amino acid composition, 168, 169
104, 108, 109, 115, 116, 123, 137,
amino groups, 48
155,157, 159, 165, 167, 168, 169, 170,
analytical technique, vii, x, 112, 114, 116,
172, 173, 180, 181, 182, 183, 186, 187,
119, 120, 122, 125
188, 190, 191, 192, 193
animal welfare, 59, 135
acid whey, vii, ix, xii, 88, 99, 100, 102, 104,
anti-cancer, 16
108, 109, 180, 181, 186, 187, 188, 190,
antioxidants, viii, ix, xi, 2, 3, 4, 5, 6, 7, 13,
191, 192, 193
14, 15, 16, 17, 18, 20, 21, 22, 23, 24, 27,
acidic, x, 112
29, 30, 31, 33, 34, 35, 36, 37, 38, 39, 40,
activation energy, 45, 53
43, 54, 55, 56, 57, 58, 60, 62, 66, 67, 68,
additives, xi, 6, 23, 40, 61, 94, 98, 179, 181,
69, 70, 71, 72, 73, 74, 75, 88, 96, 99,
182, 186, 188
100, 103, 104, 105, 107, 108, 179, 183,
adenosine triphosphate, 134
187, 188
adipose, 22, 29, 66, 74, 103
apple pomace, 7, 13, 32, 33, 34, 37, 42
adipose tissue, 22, 29, 66, 74, 103
arginine, 45, 50, 52, 168, 184
adverse effects, 44, 139
aromatic compounds, 3, 93, 96
agro-industrial by-products, 2, 6, 24, 25
ascorbic acid, 3, 72, 105, 182, 183
aldehydes, 3, 46, 68, 71, 94
assessment, 69, 71, 83, 107
alkaline media, x, 111, 116, 118
atherosclerosis, 5, 15
alpha-tocopherol, 40
atmosphere, 35, 45, 62, 68, 69, 72, 77, 79,
alternative energy, 27
81, 83, 147
ambient air, 93
atmospheric pressure, 118
196 Index
atomic emission spectrometry, 130 calibration, 119

ATP, 134, 135, 139, 143, 145, 146 cancer, xi, 5, 6, 15, 18, 21, 23, 34, 165, 179,
180, 181, 184, 189
capillary, x, 111, 120, 124, 125
B carbohydrates, 3, 49, 91, 113
carbon, 46, 50, 52, 63, 85
bacteria, ix, xii, 9, 10, 80, 87, 89, 91, 93, 99,
carbon dioxide, 63, 85
149, 180, 182, 187, 188
carbon-centered radicals, 50
bacteriocins, 99
carbonyl groups, 46
bacteriostatic, 91, 99
carcinogen, 181
beef, 7, 8, 10, 18, 19, 21, 25, 26, 27, 29, 31,
cardiovascular disease, 5, 6, 15, 18, 21, 23,
32, 33, 34, 35, 38, 40, 41, 57, 58, 61, 62,
34, 35, 180, 190
63, 64, 65, 66, 68, 71, 72, 73, 75, 76, 77,
cardiovascular diseases, 5, 15, 23
89, 97, 104, 109, 126, 136, 138, 140,
cardiovascular function, 185
142, 143, 148, 149, 150, 151, 164, 166,
carotenoids, 3, 4, 58
168, 170, 172, 173, 181, 183, 187, 188,
cattle, 56, 58, 75, 82, 123, 142
193
cell membranes, 2, 4, 183
beneficial effect, 182, 185, 188
cell organelles, 3, 5
benefits, vii, xii, 24, 59, 61, 67, 98, 103,
chemical, vii, viii, xi, 16, 21, 30, 32, 35, 36,
104, 113, 122, 147, 180, 181, 185, 190
39, 41, 44, 52, 72, 78, 82, 83, 84, 85, 89,
benzo(a)pyrene, 129
90, 94, 95, 98, 101, 103, 105, 109, 115,
bioactive compounds, viii, 2, 15, 32, 88, 100
119, 125, 156, 157, 159, 163, 172, 174,
bioavailability, 6, 192
188, 191, 193
biochemistry, 63, 67, 68
chemical characteristics, xi, 78, 83, 156,
bioconversion, 184
157, 174
biological systems, 54
chicken, 10, 12, 14, 20, 25, 26, 27, 28, 29,
biomarkers, 47, 52, 189
30, 33, 35, 36, 38, 39, 40, 41, 42, 61, 62,
blood, ix, 5, 55, 88, 180, 185
64, 65, 69, 72, 81, 83, 85, 86, 106, 183
blood pressure, ix, 88, 185
cholesterol, xi, 7, 8, 24, 35, 36, 70, 76, 96,
body weight, 14, 15, 17, 158, 161, 162, 163,
155, 157, 163, 164, 165, 172
164, 165, 166
chromatographic technique, 47, 80
bone, 39, 134, 158
chromatography, x, 47, 49, 52, 68, 111, 120,
Botswana, 172
124, 193
Brazil, 43, 78, 79, 80, 83, 111, 113, 114,
chronic diseases, 6
170
citrus pulp, 13, 14, 15, 27, 28, 30, 31, 32,
breeding, 95, 159, 171
34, 36, 42, 70
bresaola, 170, 173
cognitive impairment, 189
buffalo, 142, 143, 150
cold shortening, x, 133, 134, 135, 136, 137,
by-products, viii, 1, 2, 6, 7, 8, 14, 16, 17, 18,
139, 141, 142, 149, 163
19, 22, 23, 24, 25, 26, 28, 30, 36
collagen, 64, 149, 160, 162, 173
color, xi, 5, 9, 10, 11, 13, 14, 15, 16, 19, 20,
C 21, 23, 25, 28, 34, 36, 44, 56, 57, 61, 62,
63, 65, 68, 69, 75, 76, 77, 173, 179, 182,
caecum, 158 186, 187, 192, 193
calcium, 13, 142, 164 colorectal cancer, 180, 189
Index 197
colour stability, 101, 144, 151, 173 creatine, 135

combined effect, 33, 72, 135 CRM, 120
combustion, x, 112, 116, 117, 118, 126 cultivation, 18, 96
commercial, 25, 92, 136, 137, 147 cultivation conditions, 18
compensation, 189 culture, ix, 39, 88, 92, 105, 106, 107, 113,
complex interactions, 74, 140 186
complexity, 47 curing, ix, xi, 87, 88, 90, 95, 98, 106, 107,
composition, viii, xi, 3, 6, 13, 15, 18, 21, 24, 179, 181, 182, 183, 186, 190
25, 27, 30, 31, 32, 33, 35, 36, 37, 38, 40, curing process, xi, 98, 107, 179, 182, 183,
41, 44, 46, 52, 57, 58, 61, 64, 65, 66, 72, 186
74, 75, 89, 94, 95, 102, 103, 108, 109, cysteine, 45, 64, 99, 182
112, 124, 129, 147, 155, 158, 159, 163, cytochrome, 74
165, 167, 168, 169, 172, 173, 174, 187,
190, 191, 192
compounds, viii, xi, 2, 5, 6, 7, 10, 15, 22, D
23, 24, 32, 34, 38, 40, 41, 43, 44, 45, 46,
decomposition, 6, 52, 53, 119
47, 48, 49, 50, 51, 52, 53, 54, 55, 60, 68,
deficiency, 180, 185, 189, 190
73, 88, 91, 94, 98, 100, 102, 106, 107,
degradation, 2, 5, 27, 41, 47, 49, 100, 101,
179, 180, 182, 183, 185, 191
115, 116, 117, 118, 149
comprehension, 63, 65
dehydration, 90, 93, 156
conditionally essential, 168
denaturation, 57, 60, 62, 65, 97, 144
conditioning, 134, 136
denitrifying, ix, 88, 99
conjugated dienes, 3, 46, 47, 187
deposition, 22, 57, 58, 74
conjugation, viii, 43
depressive symptoms, 180
connective tissue, 64, 134, 140, 160, 161
derivatives, 18, 49, 52, 183
conservation, 60, 182
detection, 112, 114, 119, 123
constituents, 91, 94, 180
detection system, 123
consumers, vii, viii, xi, 1, 2, 44, 54, 83, 89,
developed countries, 2
94, 95, 143, 145, 146, 156, 157, 160,
diabetes, 5, 6, 21, 165
163, 172
diet, ix, xi, 3, 4, 5, 6, 13, 14, 15, 16, 17, 18,
consumption, xi, 2, 5, 41, 58, 60, 63, 89,
21, 23, 24, 25, 26, 27, 30, 33, 34, 35, 37,
102, 113, 114, 155, 157, 165, 172, 180,
40, 56, 58, 65, 68, 70, 71, 72, 75, 88, 96,
181, 184, 185, 189, 191
97, 155, 161, 164, 165, 180, 191, 192
contamination, 101, 114, 115, 116, 119
dietary fat, 58
control group, 24, 143, 146
dietary fiber, 9, 13, 14, 30, 38
controversial, 15
dietary habits, 165
controversies, 192
dietary intake, 180, 184
cooking, 2, 11, 12, 15, 19, 21, 33, 60, 71,
dietary supplementation, 5, 13, 14, 15, 17,
76, 136, 161
23, 28, 29, 31, 38, 39, 40
copper, 3, 53, 55, 73
diffusion, 90, 93, 98, 106
coronary heart disease, 5, 15, 165
digestibility, 26, 31, 33, 42, 72
cost, viii, xi, 2, 6, 24, 28, 54, 118, 125, 155,
digestion, x, 111, 116, 117, 118, 123, 126,
157
127, 158
country of origin, 30, 89
diseases, vii, viii, xi, 1, 5, 89, 179, 180
covalent bond, 46, 53
disposition, 184
198 Index
dissociation, 44 equipment, 117, 123, 144

distribution, 25, 116, 138 ESI, 51, 52, 71
diversity, 102, 171 esophagus, 5, 15
DNA, 23 essential fatty acids, 2, 22
DNA damage, 23 Europe, x, 133, 137
Donkey, v, xi, 152, 153, 154, 155, 156, 158, European Commission, 113, 115, 116, 129
159, 163, 169, 171, 172, 176, 177 European Union, 113, 182
Donkey Carcass, vii, xi, 145, 146, 156, 158, evaporation, 93, 186
163 evidence, 142, 180
donkey meat, vii, xi, 150, 153, 155, 156, evolution, xi, 107, 155, 169
158, 159, 161, 162, 163, 164, 165, 166, exercise performance, 192
168, 169, 170, 171, 172, 174, 177 exposure, viii, 3, 44, 45, 53, 62, 136, 184,
double bonds, 44, 53 191
drinking water, 192 extraction, x, 7, 13, 18, 31, 47, 112, 116,
dry matter, 15, 17, 22, 23, 26 118, 119, 125, 126, 127
Dry-cured meat, ix, 87, 88, 108 extracts, 18, 25, 26, 27, 28, 31, 32, 35, 38,
drying, 6, 13, 33, 93, 96, 101, 116, 170 39, 62, 70, 71, 100, 109, 193
E F
ecology, 102 families, 49, 55

economic losses, viii, 43, 64 farmers, viii, xi, 2, 155, 157, 159, 171
elderly population, 189 fat, ix, xi, 2, 14, 15, 17, 19, 21, 22, 23, 26,
electric current, x, 133, 137 29, 32, 33, 35, 37, 40, 41, 57, 58, 63, 74,
electric field, 69 75, 81, 88, 90, 94, 95, 96, 102, 103, 104,
Electrical Stimulation, vii, x, 133, 134, 144, 105, 106, 107, 109, 155, 157, 158, 163,
145, 146, 147, 148, 149, 150, 151, 153, 165, 170, 172, 187
162, 163, 172, 173, 174, 177 fatty acid composition, 13, 31, 32, 33, 37,
electricity, 136, 146 38, 40, 57, 58, 61, 74, 75, 95, 108, 165,
electrode surface, 121 187, 190
electrodes, 120, 143 fatty acids, 3, 15, 17, 22, 23, 44, 45, 46, 49,
electrophoresis, x, 111, 120, 124, 125 52, 57, 58, 66, 80, 94, 95, 96, 101, 102,
emission, 114, 122, 123, 124 108, 165, 167, 170, 172
energy, 13, 23, 45, 53, 61, 116, 118, 119, feed additives, 16, 67, 104
123, 124, 139, 140 fermentation, ix, 87, 88, 89, 91, 102, 106,
environment, viii, 1, 2, 14, 60, 93, 183 181
environmental conditions, 52, 101 fermentation and ageing, ix, 87, 89
environmental contamination, 113 fiber(s), 13, 15, 17, 23, 30, 39, 42, 56, 57,
environmental control, 93 60, 64, 65, 81, 103, 149
environmental effects, 6, 24 fish, 38, 60, 61, 62, 63, 64, 65, 71, 72, 76,
enzymes, 3, 4, 7, 15, 23, 24, 27,52, 54, 55, 82, 84, 96, 113, 126, 165
60, 92, 134, 141, 142, 143, 147, 184 fish oil, 38, 96
epidemiologic, 189 fitness, 185, 191
epidemiology, 189 flame, 121, 122
equilibrium, 120 flavonoids, 4, 15, 16, 17, 28, 29, 42, 55, 100
Index 199
flavor, xi, 8, 10, 13, 21, 44, 46, 49, 59, 107, grass, 14, 59, 75
108, 179, 183 grazing, 32, 171
flora, 91, 101, 171 growth, xi, 8, 9, 12, 13, 14, 17, 23, 24, 25,
fluorescence, 120, 123, 124, 130 26, 27, 28, 29, 30, 31, 32, 34, 39, 40, 42,
foals, 153, 162, 163, 166, 168, 173, 174, 57, 63, 91, 92, 93, 98, 100, 101, 102,
176 137, 149, 162, 179, 183
folic acid, 80, 190 growth hormone, 17
food, vii, ix, xi, 1, 6, 7, 14, 16, 24, 30, 31, growth rate, 162
40, 66, 67, 71, 73, 74, 75, 76, 80, 87, 88, guidelines, 192
92, 94, 100, 101, 103, 104, 113, 114,
123, 147, 156, 157, 168, 169, 171, 182,
184, 185, 188, 192 H
food additive, 73, 74, 182
halogens, 116, 117
food chain, 6, 24
hardness, 8, 9, 10, 19, 20, 61, 64, 101
food industry, vii, 1, 16, 92
harvesting, 27
food production, 89, 188
health, vii, viii, ix, xi, 1, 2, 5, 6, 7, 24, 61,
food products, xi, 88, 156, 157
67, 73, 74, 88, 89, 94, 102, 104, 113,
food safety, 76, 101, 114, 147
156, 157, 180, 185, 186, 188, 190
force, 16, 22, 145, 148, 161, 163
health effects, 73, 74, 113, 180, 186
formation, xi, 6, 7, 8, 12, 20, 31, 33, 38, 44,
health-promoting, 89, 94
46, 48, 49, 52, 53, 54, 60, 61, 62, 63, 65,
heart disease, 165
73, 76, 86, 91, 94, 98, 100, 105, 117,
heat shock protein, 35
179, 181, 182, 183, 185, 186, 188
heat transfer, 61
France, ix, 30, 87, 170
heme, xi, 47, 53, 57, 77, 99, 179, 183, 187,
free radicals, viii, 2, 3, 4, 6, 15, 45, 52, 54,
188
56, 59, 61
histidine, 45, 168
freezing, 60, 69, 76, 85, 89, 136
homeostasis, 59
freshwater, 78, 83, 84, 124, 126
homocysteine, 180
fruits, 13, 15, 55, 184
homogeneity, 119
functional food, 27, 28, 96, 103, 104, 107
horses, 156, 158, 174
functional foods, 28, 96, 103, 104, 107
human, vii, viii, x, xi, 1, 6, 15, 41, 67, 70,
105, 111, 113, 155, 164, 172, 184, 185,
G 188, 191
human body, 184, 185
gastrointestinal tract, 6 human health, vii, viii, x, xi, 1, 6, 41, 67, 70,
gel, 34, 97, 107 111, 113, 155, 172, 188
genes, 5, 17, 56, 74 humidity, 4, 26, 90, 93
genetic factors, 65, 74 hydrogen, 4, 45, 49, 52, 53, 54, 56
global scale, ix, 87 hydrolysis, 100, 139
glutathione, 3, 17, 24 hydroperoxides, 3, 46, 47, 49, 53, 54
glycerol, 44 hydrophobicity, 65
Glycogen, 56, 59, 139, 163, 164 hydroxide, 118
glycolysis, x, 133, 135, 137, 142, 146, 147, hydroxyl, 5, 15, 49, 52, 53, 54, 55, 56
163 hydroxyl groups, 15, 55
graphite, 121, 122, 124 hypertension, 165
200 Index
hypothesis, 162 Lactobacillus, 92, 99, 108, 187

lactoferrin, 3, 99, 188
L-arginine, 184
I laser ablation, 123
Latin America, 156
incidence, 18, 61, 145, 165
lead, 5, 44, 50, 51, 101, 129, 181, 188
income, 159
legislation, 134
incubation time, 192
light, xi, 3, 45, 52, 53, 64, 69, 86, 94, 101,
India, 21, 37, 114
155, 166
industry, vii, viii, 1, 13, 22, 24, 39, 44, 54,
linoleic acid, 7, 8, 22, 58, 69, 96
55, 59, 61, 62, 65, 75, 92, 94, 95, 114,
lipases, 108
147
lipid oxidation, 2, 5, 7, 8, 9, 10, 11, 13, 14,
infants, 184
16, 17, 22, 23, 25, 27, 28, 31, 32, 36, 37,
inflammation, 5, 15, 184
38, 39, 44, 48, 51, 57, 59, 65, 66, 67, 68,
ingredients, ix, 4, 6, 7, 24, 41, 70, 88, 90,
69, 70, 72, 73, 76, 78, 79, 82, 100, 104,
94, 95, 101, 183, 186, 188
105, 106, 183, 187, 190, 191, 193
inhibition, 63, 67, 91, 100, 141
lipid peroxidation, 3, 5, 23, 33, 38, 42, 50,
inhibitor, 106
67, 95, 183
initiation, viii, 3, 43, 45, 54, 183
lipids, viii, 2, 3, 22, 35, 43, 44, 45, 47, 52,
insulin, 185, 191
53, 54, 57, 61, 67, 70, 73, 91, 100, 172,
insulin resistance, 191
183
integrity, 3, 4, 5
lipolysis, 91, 94, 98, 101, 103, 105
interference, 47
liquid chromatography, 47, 51, 52, 71
intoxication, 184
liquids, 124
intramuscular fat, 22, 32, 35, 57, 90, 95,
Listeria monocytogenes, 12, 16, 32, 40, 98,
165, 173
193
ionization, 122, 193
liver, 17, 23, 24, 72, 126, 180
ionizing radiation, 76
livestock, 6, 14, 18, 21, 23, 24, 89, 95, 142,
ions, 53, 95, 120, 123, 124, 135
159, 173
iron, xi, 3, 47, 50, 53, 55, 57, 60, 62, 73,
local conditions, 184
156, 157, 179, 180, 182, 183, 187
low temperatures, ix, 88
irradiation, 61, 65, 69, 76, 85, 86, 118
low-density lipoprotein, 24
issues, 107, 116, 123, 125, 192
lycopene, 23, 24, 27, 29, 31, 32, 33, 37, 38
Italy, ix, 18, 80, 87, 89, 133, 151, 152, 155,
lysine, 45, 50, 52, 68, 168, 185
157, 161, 170, 174, 175
M
K
machinery, 134, 160
ketones, 46, 94
macromolecules, 124
kidney, 5, 24, 126, 180
magnitude, 136, 139
kinetics, 149
Maillard reaction, 71
majority, 47, 60, 184
L mammals, 113, 184
management, viii, 44, 58, 59, 161
lactic acid, ix, 9, 10, 87, 89, 91, 99 manipulation, 56, 94
Index 201
manufacturing, 170 metabolism, 15, 25, 34, 42, 55, 57, 93, 143,
marine fish, 126 149, 162
MAS, 122 metabolites, 15, 33, 92, 99, 101, 146
mass, x, 49, 90, 93, 111, 120, 121, 123, 124, metal ions, 4, 53, 100
193 metals, 3, 45, 52, 124
mass spectrometry, x, 49, 111, 120, 123, methemoglobinemia, 191
193 microorganisms, 91, 92, 93, 98, 101
materials, 17, 116, 120 mixing, 59, 96
matrix, viii, 43, 45, 46, 47, 49, 50, 51, 52, modifications, viii, ix, 43, 45, 47, 49, 51, 57,
53, 54, 55, 57, 60, 63, 65, 97, 112, 116, 66, 88
117, 119, 120, 123, 125 moisture, 6, 8, 9, 13, 20, 63, 64, 66, 90, 93
matrixes, 60 moisture content, 6, 9, 20
measurement, viii, 16, 44, 68 mold, 10, 11, 12, 19, 106
meat analysis, 112 molecular oxygen, 184
meat colour, 144, 145, 146, 149, 166, 173 molecular weight, 108
meat consumption, xi, 113, 155, 157, 189 molecules, 3, 13, 55, 60, 101, 122, 139
meat nutritional value, 156 monounsaturated fatty acids, 13, 167
meat processing, 2, 44, 92, 115, 181, 182, mortality, 35, 180, 191
188 muscle contraction, 134
meat production, x, xi, 54, 134, 158, 159, muscles, vii, x, xi, 38, 57, 66, 68, 71, 78, 79,
171, 179, 181 82, 94, 103, 134, 135, 136, 137, 139,
meat products, vii, viii, ix, xi, xii, 1, 3, 4, 5, 140, 141, 144, 145, 146, 147, 148, 149,
7, 8, 13, 14, 16, 18, 19, 21, 22, 23, 24, 150, 151, 156, 158, 161, 162, 165, 166,
28, 29, 30, 33, 36, 37, 39, 41, 42, 66, 67, 167, 168, 169, 173
68, 72, 73, 76, 87, 88, 90, 92, 94, 97, 99, mutation, 56, 74, 115
100, 101, 102, 103, 104, 105, 107, 108, myoglobin, 53, 57, 60, 61, 62, 63, 73, 92,
112, 113, 114, 124, 129, 147, 170, 171, 146, 166, 173, 182
179, 180, 181, 182, 183, 184, 186, 187, myosin, 73, 144
188, 189, 190, 192
meat quality, viii, x, xi, 1, 15, 16, 17, 24, 27,
28, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, N
40, 42, 44, 45, 56, 59, 60, 61, 62, 63, 66,
NaCl, 10, 90, 98, 103, 104, 105, 109
70, 74, 75, 111, 112, 114, 138, 139, 145,
National Research Council, 152, 175
148, 149, 150, 153, 156, 163, 166, 168,
natural compound, 13
173, 174, 176
natural resources, 171
meat quality control, 112, 114
negative effects, ix, 4, 17, 88
meat tenderness, vii, x, 61, 133, 134, 136,
Netherlands, 68, 80, 152, 173, 175, 189
139, 141, 142, 143, 144, 145, 149, 150,
New Zealand, x, 133, 137, 143, 148
151, 159, 160, 161, 162, 173, 174
nitrates, vii, xi, xii, 90, 98, 101, 179, 180,
medicine, 185, 190
181, 182, 183, 184, 185, 186, 188, 190,
Mediterranean, ix, 14, 18, 21, 87, 156, 186
192
Mediterranean countries, ix, 87
nitric oxide, xi, 5, 180, 182, 183, 184, 185,
membranes, 5, 15, 141
191
meta-analysis, 189, 191
nitric oxide synthase, 184
202 Index
nitrite, ix, xi, xii, 8, 9, 14, 19, 20, 28, 32, 37, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79,
41, 88, 89, 90, 92, 98, 100, 104, 105, 82, 88, 91, 94, 96, 97, 99, 100, 101, 102,
107, 108, 179, 180, 181, 182, 183, 184, 104, 105, 106, 120, 183, 187, 188, 190,
185, 186, 187, 188, 189, 190, 191, 192 191, 193
nitrites/nitrates, vii, xi, xii, 179, 180, 181, oxidation products, 5, 20, 46, 49, 61, 65, 76,
182, 183, 184, 186 94
nitrogen, ix, 63, 88, 91, 92, 181, 184, 186 oxidation rate, 17, 64
nitrogen compounds, ix, 88, 91 oxidative damage, 2, 3, 5, 56
nitrosamines, xii, 98, 105, 180, 184, 187, oxidative reaction, viii, 43, 45, 50, 57, 60,
188 61, 62, 63, 64, 65
nitroso compounds, 181, 184, 188 oxidative stability, 8, 9, 10, 12, 13, 16, 17,
N-nitroso compounds, 181, 184, 188 18, 19, 20, 23, 25, 26, 28, 29, 31, 38, 40,
non-metals, 124 44, 57, 58, 59, 61, 63, 65, 66, 68, 76, 78,
Nuclear Magnetic Resonance, 82 79, 82, 91, 98, 187
nutraceutical, 100, 152, 175 oxidative stress, 4, 5, 17, 37, 38, 59
nutrient, 2, 7, 18, 23, 27, 31, 42, , 75 92 oxygen, viii, 3, 4, 26, 27, 44, 45, 46, 50, 52,
nutrition, 4, 26, 66, 73, 74, 105, 113, 191 53, 54, 55, 62, 66, 67, 68, 77, 100, 101,
nutritional, xi, 2, 4, 5, 6, 23, 29, 37, 41, 63, 117, 118, 135, 146, 182, 183, 185
71, 83, 89, 94, 95, 97, 102, 104, 106,
108, 112, 113, 155, 163, 168, 170, 172,
173, 190, 191 P
Pakistan, 172
O pancreas, 5, 15, 17
pantothenic acid, 180
OECD, 113, 114, 131 pasture, 36, 58, 71, 161
oil, 15, 18, 22, 31, 37, 40, 41, 42, 59, 70, 71, pathway, x, 6, 49, 51, 52, 133, 146, 162,
96, 97, 101, 102, 106, 109 184, 191
oleic acid, 57, 72, 165 pectoralis major, 13
olive cake, 18, 21, 25, 26, 31, 32, 33, 35, 36, peptides, ix, 45, 49, 50, 88, 94, 103, 108
40, 41, 70 permeability, 3, 105
olive oil, 18, 26, 35, 36, 39, 97, 102, 104, peroxidation, 3, 41, 59, 67
106 peroxide, 50, 52, 53
omega-3, 26, 38, 105 peroxynitrite, 5
opportunities, 172 pH, 8, 9, 10, 11, 12, 14, 16, 19, 20, 56, 57,
organic food, 89 59, 63, 89, 90, 92, 99, 135, 137, 138,
organism, vii, viii, 1, 2, 4, 24 139, 140, 141, 142, 143, 144, 145, 146,
organs, 17, 21, 158 147, 148, 151, 182, 185, 192
ornithine, 185 phenolic compounds, 4, 13, 18, 28, 55, 73
osmotic pressure, 93 phenylalanine, 2, 45
osteoporosis, 5, 6 phosphate, 98, 135
oxidation, vii, viii, ix, 2, 3, 4, 5, 7, 8, 9, 10, phospholipids, 2, 44, 76
11, 12, 13, 14, 16, 17, 19, 20, 22, 23, 25, phosphorus, 164, 180
27, 28, 31, 32, 35, 36, 37, 38, 39, 40, 41, photooxidation, 45, 53
43, 44, 45, 46, 48, 49, 50, 51, 52, 53, 54, photosensitizers, 45, 53
56, 57, 59, 60, 61, 63, 64, 65, 66, 67, 68,
Index 203
physicochemical characteristics, 59, 76, protein oxidation, viii, 8, 12, 40, 41, 43, 44,
108, 187, 192 49, 51, 52, 57, 60, 65, 69, 70, 71, 72, 75,
physicochemical properties, 26, 89, 98, 100, 76, 77
105, 106, 187 protein structure, 61, 62
physiology, 136, 191 proteins, vii, viii, xii, 1, 2, 3, 21, 23, 43, 44,
pigs, 13, 15, 17, 21, 23, 25, 26, 27, 28, 33, 45, 46, 47, 48, 51, 52, 53, 54, 57, 61, 66,
34, 42, 56, 57, 58, 76, 142, 144, 148 70, 71, 73, 91, 94, 99, 113, 141, 142,
plants, 16, 41, 55, 56, 100, 181, 186 144, 146, 161, 163, 168, 180, 183, 188
PM, 8, 19, 20 proteolysis, 59, 91, 92, 94, 98, 101, 103,
Poland, 87, 179 105, 106, 141, 142, 149, 163
pollutants, 18 proteolytic enzyme, 64
pollution, 18 pulp, 7, 13, 14, 15, 18, 21, 22, 23, 26, 27,
polycyclic aromatic hydrocarbon, 193 28, 29, 30, 31, 32, 34, 36, 39, 40, 42, 70
polymers, 15 pure water, 93, 115
polyphenols, 7, 16, 18, 72, 100, 101, 105 purity, 92
polyunsaturated fat, viii, 13, 14, 15, 17, 21,
22, 23, 24, 35, 43, 70, 71, 96, 165, 167,
170, 183 Q
polyunsaturated fatty acids, viii, 13, 14, 15,
quality, vii, viii, ix, xi, 1, 2, 3, 5, 7, 8, 12,
17, 21, 22, 23, 24, 35, 43, 70, 71, 96,
13, 14, 18, 19, 21, 22, 24, 26, 27, 28, 29,
165, 167, 183
30, 33, 36, 38, 39, 40, 41, 42, 43, 44, 57,
pomegranate by-products, 22
60, 63, 64, 66, 67, 68, 69, 71, 72, 73, 74,
population, 113, 156, 190
75, 76, 77, 78, 82, 84, 85, 86, 88, 92, 95,
potassium, 98, 101, 104, 109, 182
97, 99, 100, 101, 102, 103, 104, 105,106,
poultry, 16, 33, 57, 60, 61, 62, 63, 74, 75,
107, 108, 109, 134, 138, 139, 143, 145,
83, 84, 101, 103, 113, 142
146, 147, 148, 149, 150, 151, 156, 157,
preparation, vii, x, 49, 112, 115, 116, 118,
159, 160, 162, 168, 170, 171, 172, 173,
119, 125, 126, 127, 135
174, 181, 186, 188, 189, 192, 193
preservation, viii, ix, 1, 18, 59, 61, 62, 67,
quality and nutritional, vii, x, 88, 100
88, 89, 99
quality control, 112, 114
preservative, ix, 87, 91
quality improvement, viii, 44
prevention, viii, 1, 15, 34, 141, 189
quality standards, 171
primary antioxidants, 54
quantification, 48, 49, 50, 112, 114
primary products, 46, 47
quartz, 117, 121, 122
probiotic, 39, 80, 92, 93, 99, 100, 105, 106,
quercetin, 13, 37, 40
107, 108, 109, 187, 193
probiotics, 14, 25
processed meat, xi, 16, 23, 67, 70, 104, 170, R
179, 180, 181, 189, 191
producers, 2, 18, 89, 94, 188 radiation, 69, 81, 121, 122, 124
production costs, 21, 113 radical formation, 3
production technology, ix, 87, 89 radical mechanism, 54
proline, 45, 50, 52 radicals, 3, 4, 5, 6, 32, 45, 46, 49, 52, 53, 54,
propagation, viii, 3, 43, 45 56
protection, 4, 25, 39, 96 rancid, 2, 5, 46
204 Index
reaction rate, 52 seed, 9, 16, 17, 22, 26, 27, 28, 29, 30, 31,
reactions, viii, xi, 4, 5, 43, 44, 45, 46, 49, 32, 33, 34, 35, 36, 37, 40, 41, 62, 96,
50, 52, 53, 54, 55, 56, 57, 59, 60, 62, 63, 100, 104, 105, 109, 187, 188, 191
64, 66, 72, 89, 91, 94, 139, 180, 182, 183 selenium, 38, 180
reactive oxygen, 5, 23, 45, 49, 72 sensitivity, 56, 121, 124
rectus femoris, 89 sex, 3, 44, 56, 57, 161, 173
red wine, 18, 38 shape, 92, 139
reducing sugars, 50 shear, 16, 22, 144, 145, 148, 161, 163
reflectance spectra, 77 sheep, 14, 74, 113, 138, 139, 140, 142, 143,
regions of the world, 159 149, 150, 173
rejection, 2, 64, 65 shelf life, ix, 4, 5, 13, 16, 26, 33, 35, 41, 59,
requirement, 49, 180, 182 61, 62, 63, 66, 69, 81, 82, 83, 85, 88, 89
researchers, 14, 96, 100, 138 showing, vii, xi, 156, 158, 163
residues, 7, 13, 18, 39, 51, 52, 64, 183 side chain, viii, 43, 46, 49
resolution, 122, 123 skeletal muscle, 26
resources, 24, 75 skin, 7, 17, 18, 83, 90
response, 23, 24, 35, 137, 143, 144 small intestine, 15, 158
resveratrol, 17, 42 smoking, 96, 181, 190
retail, 63, 147 sodium, ix, xii, 37, 83, 85, 86, 88, 90, 95,
riboflavin, 53, 180 97, 98, 103, 104, 106, 109, 164, 180,
rigor mortis, x, 133, 134, 135, 136, 142, 182, 186, 191
143, 144, 147, 149, 162 solid phase, 119
risk, vii, xi, xii, 23, 89, 119, 165, 179, 180, solid waste, 27
181, 184, 186, 189 solubility, 56, 63
ROOH, 3 solution, ix, 41, 88, 96, 112, 116, 117, 118,
room temperature, 86, 97, 185 119, 120
rural areas, 171 South Africa, 113, 114
Russia, 113, 114 Spain, ix, 18, 30, 80, 87, 89, 160, 170
speciation, 115, 122, 124, 125
species, viii, 2, 3, 5, 23, 25, 26, 44, 45, 46,
S 49, 52, 53, 56, 57, 58, 60, 64, 65, 72, 77,
84, 95, 107, 113, 115, 116, 117, 122,
safety, vii, x, xi, 2, 55, 59, 61, 76, 88, 89,
124, 125, 135, 142, 143, 144, 153, 161,
100, 101, 114, 137, 144, 147, 179, 183,
167, 170, 174, 177
188, 189, 192
specifications, 137
Salami, 170
spectrophotometric method, 47
salting, 89, 90, 106, 170, 181
spectrophotometry, 47, 50
salts, 91, 94, 98, 107
spectroscopy, 71, 120
Sample preparation, 112, 116, 126, 127
stability, ix, xi, 2, 8, 9, 10, 12, 13, 14, 16,
sample storage, 115, 116, 125
17, 18, 19, 20, 23, 25, 26, 27, 28, 29, 31,
sampling, 115, 116, 125, 129
34, 37, 38, 40, 44, 57, 58, 59, 61, 63, 65,
saturated fat, 23, 96, 165, 167, 170
66, 67, 68, 69, 70, 75, 76, 78, 79, 82, 87,
saturated fatty acids, 96, 165, 167, 170
91, 96, 98, 142, 144, 179, 187, 188
scarcity, 159
staphylococci, 182
scavengers, 54, 55, 100
science, 189, 190
Index 205
state, 52, 64, 77, 83, 120, 121, 122, 134, textural character, 106
135, 161, 166, 189 texture, viii, ix, 8, 13, 43, 44, 56, 62, 63, 64,
stimulation, vii, x, 133, 134, 138, 139, 140, 69, 71, 76, 86, 87, 91, 92, 93, 98, 101,
144, 145, 146, 147, 148, 149, 150, 151, 143, 174
153, 162, 172, 173, 174, 177 therapeutics, 191
storage, viii, xi, 2, 5, 7, 16, 26, 27, 28, 29, threonine, 50
30, 31, 33, 35, 39, 40, 41, 44, 54, 63, 65, tissue, 5, 6, 58, 74, 90, 93, 126, 134, 141,
68, 69, 70, 71, 72, 75, 76, 77, 82, 86, 158, 160, 184
100, 106, 108, 115, 116, 125, 161, 163, tocopherols, 4
166, 179, 182, 185, 187, 188, 189, 193 tomato pomace, 23, 28, 37, 38, 42
stress, 5, 35, 56, 59, 75, 134, 144 total cholesterol, 24
structural changes, 61, 64, 77 total energy, 143
structural protein, 94 toxic effect, 4
structure, x, 3, 5, 34, 44, 62, 64, 66, 90, 134, toxicity, 55, 184
146, 162 trace-element in meat, 112
style, 7, 8, 42, 88, 186, 192 traits, 15, 21, 24, 32, 40, 44, 57, 60, 61, 62,
substitution, 39, 104, 109 64, 72, 163
substrates, 28, 46, 182 transition metal, 53, 99, 188
subtraction, 123 transition metal ions, 53
sulfur, xii, 180, 183 transport, 3, 59, 122, 156
Sun, 7, 8, 40, 66, 76 transport processes, 3
supplementation, 5, 14, 16, 17, 18, 22, 23, transportation, 59, 172
24, 25, 32, 33, 34, 35, 36, 37, 38, 39, 42, treatment, 26, 76, 89, 98, 101, 117, 137,
58, 59, 70, 75, 185, 191, 192 138, 140, 141, 144, 161, 183
susceptibility, 17, 31, 56, 57, 60, 62 triacylglycerides, 2
sustainability, 6, 24, 83 triglycerides, 24
Switzerland, 170, 192 trimmings, 97
synergistic effect, 55, 56 tryptophan, 2, 45
synthesis, 4, 184 Turkey, 9, 37, 145
type 2 diabetes, 180, 189
tyramine, 100
T tyrosine, 45, 99, 169
target, 65, 139

TBA, 48, 49, 183 U
techniques, vii, ix, x, 6, 47, 49, 52, 67, 87,
90, 111, 112, 114, 116, 117, 118, 119, ultrasound, 98, 118, 119
120, 121, 122, 123, 124, 125 underlying mechanisms, 75
technology(ies), ix, x, 31, 61, 68, 75, 77, 88, United Kingdom, 30
89, 91, 94, 97, 98, 133, 135, 147, 150, United States (USA), x, 39, 107, 133, 137,
170, 188 148, 152, 154, 175, 177
temperature, viii, 3, 4, 44, 45, 52, 53, 59, 60,
69, 72, 90, 93, 116, 117, 118, 134, 135,
136, 139, 141, 142, 148, 151, 185
tendons, 158
terrestrial ecosystems, 113
206 Index
V W
vacuum, ix, 28, 62, 69, 76, 77, 81, 85, 88, waste, 13, 14, 18, 19, 36, 41, 84, 85
96, 100, 105, 151, 187 waste management, 14, 18
vapor, 121, 122, 124 waste water, 41, 101
variations, 48, 52, 120, 161, 170 water, 8, 13, 16, 18, 19, 35, 41, 60, 63, 64,
varieties, 16, 38 71, 72, 77, 81, 82, 83, 89, 90, 93, 103,
vastus lateralis, 89 106, 113, 117, 118, 144, 151, 161, 183,
vegetable oil, 32 184, 186, 192
vegetables, 98, 100, 184, 186, 191 water diffusion, 93
vitamin A, 3 water quality, 192
vitamin B1, 180 water vapor, 117
vitamin B12, 180 wavelengths, 122
vitamin B6, 180 weight loss, 65, 93, 96
vitamin C, 56, 96 Wisconsin, 152, 175
vitamin E, 3, 22, 24, 29, 31, 38, 56, 58, 70, World Health Organization (WHO), 55,
96 113, 129, 184, 185, 192
vitamin K, 55 worldwide, 6, 24
vitamins, vii, 1, 2, 3, 4, 13, 55, 56, 96, 100,
113, 180
volatile organic compounds, 21 Y
volatilization, 117
yeast, 11, 12, 19, 25, 91, 100, 101, 103
vulnerability, 45
yield, 14, 15, 56, 65, 143, 158

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FOOD SCIENCE AND TECHNOLOGY

MEAT AND MEAT PROCESSING

Additional books in this series can be found on Nova’s website

Additional e-books in this series can be found on Nova’s website

MEAT AND MEAT PROCESSING

NOTICE TO THE READER

Library of Congress Cataloging-in-Publication Data

ISBN:  H%RRN

Published by Nova Science Publishers, Inc. † New York

Chapter 7 Nitrites/Nitrates in Processed Meat:

Meat products occupy quite an exceptional position in the preferences of

Chapter 3 - Dry-cured meat products, such as dry-cured hams, loin or

presence of sodium chloride) properties. This chapter presents the latest

Chapter 6 - Meat has exerted a crucial role in human evolution and is an

(sodium ascorbate, sometimes bacteria), which may act as a bacteriocidal

Panagiotis E. Simitzis*, PhD

Fortification of meat with bioactive compounds that are contained in the

Keywords: agro-industrial by-products, meat, oxidation, antioxidants

1. LIPID OXIDATION AND ANTIOXIDANTS IN BIOLOGICAL

1.1. Oxidation Procedures

Meat products are essential components in the diets of developed

2014). Oxidation procedure further destroys the membrane structure, disturbs

1.2. Antioxidants and Antioxidant Systems

Living organisms have developed specific mechanisms that are known as

1.3. Antioxidants, Meat Products and Nutrition-Related Diseases

A great interest in applying antioxidants from natural sources to increase

diseases. The involvement of the reactive oxygenated/nitrogenated species,

APPLICATION OF AGRO-INDUSTRIAL BY-PRODUCTS

A constant increase of livestock production cost is observed due to the

of specific compounds well known as nutraceuticals. These compounds

2.2. Apple Pomace

Apple (Malus domestica) processing generates huge quantities of solid

By-product Level Meat product Effect Reference

By-product Level Meat product Effect Reference

By-product Level Meat product Effect Reference

2.3. Citrus Pulp

alternative of antioxidant plants/herbs and synthetic feed additives for the

2.4. Grape Pomace or Marc

Grape (Vitis vinifera) pomace or marc is derived as a result of

2.5. Olive Cake

Olive (Olea europaea) cultivation plays an important economic and social

beef patties etc.) with positive effects on their quality characteristics

By-product Level Meat product Effect Reference

By-product Level Meat product Effect Reference

Moreover, no effect of olive cake on lamb carcass traits, meat chemical

2.6. Pomegranate By-Products

Pomegranate (Punica granatum L.) is a fruit originates from Iran and

2.7. Tomato Pomace

Tomato (Solanum lycopersicum) is one of the most important vegetable

Dietary supplementation with lycopene (0.75 g/kg) positively influenced

Proceedings of Western Section of American Society of Animal Science

Fernandez-Lopez, J., Fernandez-Gines, J. M., Aleson-Carbonell, Sendra, E.,

Hawashin, M. D., Al-Juhaimi, F., Ahmed, I. A. M., Ghafoor, K., Babiker, E.

Kris-Etherton, P.M., Hecker, K.D., Bonanome, A., Coval, S.M., Binkoski,

Lorenzo, J. M., González-Rodríguez, R. M., Sánchez, M., Amado, I. R.,

Mourao, J. L., Pinheiro, V. M., Prates, J. A. M., Bessa, R. J. B., Ferreira, L. M.

Pamplona, R., Costantini, D., 2011. Molecular and structural antioxidant

Rozbicka-Wieczorek, A. J., Wiesyk E., Brzoska, F., Sliwinski, B., Kowalczyk,

quality of meat nuggets from broilers by dietary quercetin and alpha-

LIPID AND PROTEIN OXIDATION IN MEAT

Ana P. A. A. Salim1,2, Fernanda M. Viana1,2 and

been applied during pre and post-harvest periods. Furthermore, both

Keywords: meat processing, oxidative stability, lipid oxidation, protein

Proteins are the major organic components on meat and can be

ISBN: H%RRN

[90] Morello, MJ; Shahidi, F; Ho, C. Free Radicals in Foods : chemistry,