Effect of Nanoparticles on Lipid Peroxidation in Plants

*Shahla Hashemi*

## **Abstract**

The size of the nanoparticles is between 1 and 100 nm. Nanoparticles are widely used in consumer and medical products, as well as in agricultural and industrial applications. The excessive use nanoparticles increases its release into the environment. Plants are an important part of the environment that is affected by nanoparticles. Studies have examined the effect of nanoparticles on plants. The results showed that high concentrations of nanoparticles showed a negative effect. Reactive oxygen species generation is a toxicological mechanism of nanoparticles in plants. When the production of radicals is greater than its removal, oxidative stress occurs. The key indicator of oxidative stress is lipid peroxidation. The unsaturated fatty acids in the cell membrane are a major target for radicals. Radical absorbs hydrogen from unsaturated fatty acids to form water. Therefore, the fatty acid has a non-coupled electron, which is then able to capture oxygen and form a peroxyl radical. Lipid peroxyl radical can lead to a chain of radical production. Enzymatic and nonenzymatic systems exist for the removal of radicals in plants. Enzymatic systems include catalase, guaiacol peroxidase, ascorbate peroxidase, superoxide dismutase, glutathione reductase, and dehydroascorbate reductase. Nonenzymatic systems include ascorbate and carotenoids, glutathione, tocopherol, and phenolic compounds.

**Keywords:** nanoparticles, reactive oxygen species, malondialdehyde, catalase, ascorbate, glutathione

## **1. Introduction**

The nanoparticles have a size of less than 100 nm in at least one dimension. Due to the specific properties of nanoparticles, in particular the high-surfaceto-volume ratio, they have been used for several applications. For example, nanoparticles are used in the fields of biosensors and electronics, cosmetic industries, wastewater treatment, biomedicines, cancer therapy, and targeted drug delivery [1, 2]. The excessive use of nanoparticles results in the release of these materials into the environment. The environment includes plants, the main producers of the food chain, which are affected by nanoparticles. Nanoparticles are absorbed by plants and transmitted to various parts of the plants and affect them. Several factors such as physicochemical properties of nanoparticles, plant species, and exposure conditions contribute to the absorption and transfer of nanoparticles. Size, magnetic properties, surface charge, composition, crystalline state, and surface functionalization are some of the physical properties of nanoparticles that are important in their absorption

into the plant. Nanoparticles are introduced into the plant by various methods, for example, through penetration into the coating of seeds, during absorption of nutrient by the root, and entering the cuticle and stomata of the leaf. After absorbing nanoparticles, these materials can accumulate or move through the vascular system to the shoot. The first cell-level barrier to move nanoparticles is the cell wall. The size of the pores in the cell wall is 5–20 nm. Therefore, nanoparticles of less than 20 nm in size can easily pass through the pores. But nanoparticles with sizes larger than 20 nm through routes such as ion channels, endocytosis, and aquaporins and creation of new pores pass the cell wall of the barrier. The next barrier is the plasma membrane. The role of the plasma membrane is controlling the passage of materials in and out of the cell. Protein and lipids are two main parts of the plasma membrane structure. Plasma membrane lipids play an important role in determining cellular structures, regulating fluid membrane and signal transduction. Lipids are not only present in the plasma membrane but also in all parts of the plant. Plants have a diverse range of lipids including fatty acids, sterol lipids, glycolipids, sphingolipids, phospholipids, and waxes. In this chapter, we discussed the effects of nanoparticulate toxicity on the lipids of plants and plant defense mechanisms against this toxicity.

## **2. Interaction of nanoparticles with plants**

There are reports that nanoparticles can lead to stress through release of reactive oxygen species (ROS) in plants. The lack of balance between the production and removal of ROS leads to the production of oxidative stresses with oxidative damage to DNA, proteins, and fats. There are two unpaired electrons in separate orbitals in the outer shell of oxygen. This oxygen structure makes it a candidate for the production of ROS*.* ROS are free radical species and non-free radical oxygen*.* Radicals can have neutral, negative, or positive charge. Free radical is an atom or group of atoms that have one or more unpaired electrons. Free radical oxygen species contains the hydroxyl radicals (• OH) and free radicals superoxide anion (• O2 <sup>−</sup>). Non-free radical species containing hydrogen peroxide (H2O2) are various forms of activated oxygen resulted from oxidative biological reactions or exogenous factors (**Figure 1**). Radicals are naturally produced as intermediate biochemical reactions, but excess production of these radicals damages the plant and should be eliminated by the antioxidant system.

**59**

**Figure 3.**

**Figure 2.**

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

thione reductase (GR).

**3. Alpha-tocopherol**

molecules of single oxygen [3].

*Chemical structure of alpha-tocopherol [4].*

ascorbic acid or other antioxidants [4] (**Figure 3**).

*The role of antioxidant tocopherol. Polyunsaturated fatty acids (PUFA).*

The antioxidant system in the plant contains an enzymatic and nonenzymatic system. The nonenzymatic antioxidant system contains alpha-tocopherol, flavonoids, ascorbate, glutathione and phenolic compounds, and carotenoids, while the enzymatic antioxidant system includes catalase (CAT), ascorbate peroxidase (APX), superoxide dismutase (SOD), peroxidase (POX), and gluta-

Alpha-tocopherol is a hydrophobic antioxidant that is produced by all plants (**Figure 2**). This compound is present primarily in the cell membrane and plays a key role in the collection of proxy lipid radicals from lipid peroxidation. One of the most prominent properties of tocopherol is their ability to turn off single oxygen, and it is estimated that a tocopherol molecule alone can neutralize about 120

Alpha-tocopherol also acts as an end point for peroxidation reactions of unsaturated fats, which is converted to radical tocopheroxyl by reaction with lipid peroxyl radicals. Radical tocopheroxyl can be converted to tocopherol by reaction with

**Figure 1.** *Oxygen and some reactive oxygen species.*

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

The antioxidant system in the plant contains an enzymatic and nonenzymatic system. The nonenzymatic antioxidant system contains alpha-tocopherol, flavonoids, ascorbate, glutathione and phenolic compounds, and carotenoids, while the enzymatic antioxidant system includes catalase (CAT), ascorbate peroxidase (APX), superoxide dismutase (SOD), peroxidase (POX), and glutathione reductase (GR).

## **3. Alpha-tocopherol**

*Advances in Lipid Metabolism*

against this toxicity.

radicals superoxide anion (•

**2. Interaction of nanoparticles with plants**

Free radical oxygen species contains the hydroxyl radicals (•

O2

into the plant. Nanoparticles are introduced into the plant by various methods, for example, through penetration into the coating of seeds, during absorption of nutrient by the root, and entering the cuticle and stomata of the leaf. After absorbing nanoparticles, these materials can accumulate or move through the vascular system to the shoot. The first cell-level barrier to move nanoparticles is the cell wall. The size of the pores in the cell wall is 5–20 nm. Therefore, nanoparticles of less than 20 nm in size can easily pass through the pores. But nanoparticles with sizes larger than 20 nm through routes such as ion channels, endocytosis, and aquaporins and creation of new pores pass the cell wall of the barrier. The next barrier is the plasma membrane. The role of the plasma membrane is controlling the passage of materials in and out of the cell. Protein and lipids are two main parts of the plasma membrane structure. Plasma membrane lipids play an important role in determining cellular structures, regulating fluid membrane and signal transduction. Lipids are not only present in the plasma membrane but also in all parts of the plant. Plants have a diverse range of lipids including fatty acids, sterol lipids, glycolipids, sphingolipids, phospholipids, and waxes. In this chapter, we discussed the effects of nanoparticulate toxicity on the lipids of plants and plant defense mechanisms

There are reports that nanoparticles can lead to stress through release of reactive oxygen species (ROS) in plants. The lack of balance between the production and removal of ROS leads to the production of oxidative stresses with oxidative damage to DNA, proteins, and fats. There are two unpaired electrons in separate orbitals in the outer shell of oxygen. This oxygen structure makes it a candidate for the production of ROS*.* ROS are free radical species and non-free radical oxygen*.* Radicals can have neutral, negative, or positive charge. Free radical is an atom or group of atoms that have one or more unpaired electrons.

peroxide (H2O2) are various forms of activated oxygen resulted from oxidative biological reactions or exogenous factors (**Figure 1**). Radicals are naturally produced as intermediate biochemical reactions, but excess production of these radicals damages the plant and should be eliminated by the antioxidant system.

OH) and free

<sup>−</sup>). Non-free radical species containing hydrogen

**58**

**Figure 1.**

*Oxygen and some reactive oxygen species.*

Alpha-tocopherol is a hydrophobic antioxidant that is produced by all plants (**Figure 2**). This compound is present primarily in the cell membrane and plays a key role in the collection of proxy lipid radicals from lipid peroxidation. One of the most prominent properties of tocopherol is their ability to turn off single oxygen, and it is estimated that a tocopherol molecule alone can neutralize about 120 molecules of single oxygen [3].

Alpha-tocopherol also acts as an end point for peroxidation reactions of unsaturated fats, which is converted to radical tocopheroxyl by reaction with lipid peroxyl radicals. Radical tocopheroxyl can be converted to tocopherol by reaction with ascorbic acid or other antioxidants [4] (**Figure 3**).

**Figure 2.** *Chemical structure of alpha-tocopherol [4].*

**Figure 3.** *The role of antioxidant tocopherol. Polyunsaturated fatty acids (PUFA).*

## **4. Ascorbic acid**

Ascorbic acid is one of the most powerful antioxidants that has been found in a variety of plant cells, organelles, and apoplastic space. In physiological conditions, ascorbic acid is often reduced. The ability of ascorbate to give the electron in a wide range of enzymatic and nonenzymatic reactions has transformed this substance into active oxygen species detoxification compound. Ascorbic acid plays a role in collecting superoxide, hydroxyl radicals, and singlet oxygen or converting hydrogen peroxide through the reaction of ascorbate peroxidase into water [5]. The conversion of hydrogen peroxide to water in ascorbate-glutathione cycle leads to the conversion of ascorbate to monodehydroascorbate. These compounds have a short life-span and convert into ascorbate by interfering with the enzymes of monodehydroascorbate reductase and NADPH (**Figure 4**).

#### **Figure 4.**

*Ascorbate-glutathione cycle. ASC, ascorbate; MDHA, monodehydroascorbate; DHA, dehydroascorbate; GSH, glutathione; GSSG, glutathione disulfide; APX, ascorbate peroxidase; MDHAR, monodehydroascorbate reductase; DHAR, dehydroascorbate reductase; GR, glutathione reductase [6].*

## **5. Glutathione**

Glutathione, γ-glutamyl-cysteinyl-glycine, is a tripeptide that has been found in all parts of the cell, such as cytosol, endoplasmic reticulum**,** chloroplast, vacuoles, and mitochondria [7]. Glutathione is one of the main sources of thiol in most plant cells. Due to the reactivity of the glutathione thiol group, this substance has been widely recognized for a wide range of biochemical reactions. The central cysteine in the glutathione molecule has created a high potential for reduction in this molecule. Reduced glutathione can remove the hydrogen peroxide [8]. The main role of glutathione in antioxidant defense is due to its ability to produce reduced ascorbic through the ascorbate-glutathione cycle. Some researchers have reported that glutathione protects the cell from oxidative stress by reacting thiol groups with singlet oxygen and radical hydroxyl [5, 9, 10].

## **6. Phenolic compounds**

Phenolic compounds of a group of secondary metabolites include flavonoids and tannins, which are found in plant tissues abundantly. Most plants synthesize

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*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

pounds and convert them into radical cation [12]:

compounds releases electrons more easily.

the role of antioxidant activity [14, 15].

tion of singlet oxygen (**Figure 5**)

There are two types of carotenoids in plant tissues:

1.Carotenoids that only contain hydrocarbons (carotene)

compounds (POH).

deactivation [12].

in H2O2-scavenging [13].

(xanthophyll).

(**Figure 5**)

**7. Carotenoids**

phenolic compounds in natural conditions, but their synthesis and accumulation are induced by stresses of nanoparticles [11]. Phenolic compounds play a role in H2O2-scavenging, quenching of singlet oxygen*,* and reducing or inhibiting lipid oxidation [14, 15]. There are two main mechanisms for the protective role of phenolic

In the first mechanism, the hydrogen atoms of phenolic compounds are elimi-

R˙ + POH → RH + PO.

In evaluating the phenolic compounds' action in this mechanism, the bond dissociation energy of the O–H bonds is an important parameter, because the weakening of the OH bond increases the activity of phenolic compounds for radical

In the second mechanism, free radicals can take electrons from phenolic com-

R˙ + POH → R<sup>−</sup> + POH˙

According to the second mechanism, the lower ionization potential of phenolic

Therefore, the activity of phenolic compounds is easily estimated by calculating ionization potential and the bond dissociation energy of the O–H bonds [12]. Bendary et al. suggested that the phenolic compounds perform scavenging of H2O2 from the first mechanism. The number of hydroxyl groups and the aromatic ring substitution pattern are all important associated factors. The ortho and para position substitution with another hydroxyl group is another important factor that plays

Carotenoids are tetraterpenes that exist in the photosynthetic and nonphotosynthetic tissues of the plants and synthesize from isoprenoid biosynthesis pathway*.* Carotenoids act as auxiliary pigment in chloroplasts, but their main role is

2.Carotenoids which in addition to the hydrocarbon chain have oxygen atoms

These compounds carry the antioxidant role through the following routes:

2.The reaction with excite chlorophyll and gaining energy to prevent the forma-

1.Eliminating singlet oxygen and wasting energy in the form of heat

3.Waste high energy of exciting through the xanthophyll cycle [14, 15]

+

nated by free radical (R˙), and the phenolic compounds become radical:

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

*Advances in Lipid Metabolism*

droascorbate reductase and NADPH (**Figure 4**).

singlet oxygen and radical hydroxyl [5, 9, 10].

**6. Phenolic compounds**

Ascorbic acid is one of the most powerful antioxidants that has been found in a variety of plant cells, organelles, and apoplastic space. In physiological conditions, ascorbic acid is often reduced. The ability of ascorbate to give the electron in a wide range of enzymatic and nonenzymatic reactions has transformed this substance into active oxygen species detoxification compound. Ascorbic acid plays a role in collecting superoxide, hydroxyl radicals, and singlet oxygen or converting hydrogen peroxide through the reaction of ascorbate peroxidase into water [5]. The conversion of hydrogen peroxide to water in ascorbate-glutathione cycle leads to the conversion of ascorbate to monodehydroascorbate. These compounds have a short life-span and convert into ascorbate by interfering with the enzymes of monodehy-

Glutathione, γ-glutamyl-cysteinyl-glycine, is a tripeptide that has been found in all parts of the cell, such as cytosol, endoplasmic reticulum**,** chloroplast, vacuoles, and mitochondria [7]. Glutathione is one of the main sources of thiol in most plant cells. Due to the reactivity of the glutathione thiol group, this substance has been widely recognized for a wide range of biochemical reactions. The central cysteine in the glutathione molecule has created a high potential for reduction in this molecule. Reduced glutathione can remove the hydrogen peroxide [8]. The main role of glutathione in antioxidant defense is due to its ability to produce reduced ascorbic through the ascorbate-glutathione cycle. Some researchers have reported that glutathione protects the cell from oxidative stress by reacting thiol groups with

*Ascorbate-glutathione cycle. ASC, ascorbate; MDHA, monodehydroascorbate; DHA, dehydroascorbate; GSH, glutathione; GSSG, glutathione disulfide; APX, ascorbate peroxidase; MDHAR, monodehydroascorbate* 

*reductase; DHAR, dehydroascorbate reductase; GR, glutathione reductase [6].*

Phenolic compounds of a group of secondary metabolites include flavonoids and tannins, which are found in plant tissues abundantly. Most plants synthesize

**4. Ascorbic acid**

**60**

**5. Glutathione**

**Figure 4.**

phenolic compounds in natural conditions, but their synthesis and accumulation are induced by stresses of nanoparticles [11]. Phenolic compounds play a role in H2O2-scavenging, quenching of singlet oxygen*,* and reducing or inhibiting lipid oxidation [14, 15]. There are two main mechanisms for the protective role of phenolic compounds (POH).

In the first mechanism, the hydrogen atoms of phenolic compounds are eliminated by free radical (R˙), and the phenolic compounds become radical:

$$\text{R}^{\cdot}\text{ + POH}\rightarrow\text{RH}\text{ + POV}$$

In evaluating the phenolic compounds' action in this mechanism, the bond dissociation energy of the O–H bonds is an important parameter, because the weakening of the OH bond increases the activity of phenolic compounds for radical deactivation [12].

In the second mechanism, free radicals can take electrons from phenolic compounds and convert them into radical cation [12]:

$$\mathbb{R}^+\text{ + }\mathrm{POH}\rightarrow\mathbb{R}^-\text{ + }\mathrm{POH}^{\cdots\*}$$

According to the second mechanism, the lower ionization potential of phenolic compounds releases electrons more easily.

Therefore, the activity of phenolic compounds is easily estimated by calculating ionization potential and the bond dissociation energy of the O–H bonds [12]. Bendary et al. suggested that the phenolic compounds perform scavenging of H2O2 from the first mechanism. The number of hydroxyl groups and the aromatic ring substitution pattern are all important associated factors. The ortho and para position substitution with another hydroxyl group is another important factor that plays in H2O2-scavenging [13].

## **7. Carotenoids**

Carotenoids are tetraterpenes that exist in the photosynthetic and nonphotosynthetic tissues of the plants and synthesize from isoprenoid biosynthesis pathway*.* Carotenoids act as auxiliary pigment in chloroplasts, but their main role is the role of antioxidant activity [14, 15].

There are two types of carotenoids in plant tissues:


These compounds carry the antioxidant role through the following routes:


**Figure 5.**

*Schematic representation of singlet oxygen formation upon excitation of chlorophyll (Chl) and the role of carotenoids (car) in protection against photooxidative damage. The symbol \* indicates excited states.*

## **8. The xanthophyll cycle**

In green tissues, zeaxanthin epoxidase can be zeaxanthin converted to violaxanthin via the intermediate antheraxanthin. This is a reversible reaction; violaxanthin de-epoxidase converts violaxanthin to zeaxanthin by antheraxanthin. The relative concentration of zeaxanthin/violaxanthin is controlled by the xanthophyll cycle in plant photosynthetic tissues, which is a collection of light and dark control reactions.

Under high light conditions, violaxanthin de-epoxidase activated and converted violaxanthin to zeaxanthin. In dark conditions zeaxanthin is converted into violaxanthin [16] (**Figure 6**).

**63**

oxidation.

**12. Guaiacolperoxidase**

vacuole, and cell wall [21].

**13. Superoxide dismutase (SOD)**

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

Catalase has a tetramer enzyme that breaks H2O2 into water and oxygen. Catalase has a lower affinity for H2O2, so it can remove H2O2 at high concentrations [17]. Hydrogen peroxide is very toxic to plant cells, especially in the chloroplast. Hydrogen peroxide at very low concentrations prevents the activity of the enzymes of the calvin cycle, especially the enzymes with *sulfhydryl* group such as glyceraldehyde 3-phosphate dehydrogenase and fructose 1,6

Peroxides are a group of antioxidant enzymes that cause the decomposition of hydrogen peroxide with the oxidation of a substance. Peroxidases are located in the cytosol, vacuole, chloroplast, and extracellular space and are classified based on

Ascorbate peroxidase is an antioxidant enzyme that participates in the ascorbate-glutathione cycle, and its activity has been reported in the chloroplast, cytosol, peroxisome, and apoplast. This enzyme uses ascorbate as a reducing agent and decomposes hydrogen peroxide into water and oxygen [20]. The high concentration of ascorbate peroxidase to hydrogen peroxide shows that the ascorbate-glutathione cycle plays a vital role in controlling the level of radicals in cellular organs. In ascorbate-glutathione cycle with ascorbate peroxidase enzyme activity, ascorbate is oxidized to monodehydroascorbate, and ascorbate production is required to continue the cycle. In this cycle, the enzymes of monodehydroascorbate reductase (MADAR), dehydroascorbate reductase (DHAR), and glutathione reductase (GR) are active and reduce ascorbate using water and glutathione. Using NADPH, monodehydroascorbate reductase converts monodehydroascorbate to ascorbate. While dehydroascorbate reductase catalyzes dehydroascorbate to ascorbate using glutathione (GSH)

Guaiacol peroxidase oxidize guaiacol. This enzyme is also present in the cytosol,

Superoxide dismutase is one of the enzymes that is located in all intracellular organs and apoplast and is very important in the defense against active oxygen species. This enzyme converts the radical superoxide to H2O2, which H2O2 should be detoxified during the next stages of antioxidant defense. In the presence of the superoxide dismutase enzyme, this reaction occurs 10,000 times faster [22].

**9. Catalase**

bisphosphatase [18, 19].

their combined composition.

**11. Ascorbate peroxidase**

**10. Peroxidases**

**Figure 6.** *The xanthophyll cycle.*

## **9. Catalase**

*Advances in Lipid Metabolism*

**8. The xanthophyll cycle**

anthin [16] (**Figure 6**).

reactions.

**Figure 5.**

In green tissues, zeaxanthin epoxidase can be zeaxanthin converted to violaxanthin via the intermediate antheraxanthin. This is a reversible reaction; violaxanthin de-epoxidase converts violaxanthin to zeaxanthin by antheraxanthin. The relative concentration of zeaxanthin/violaxanthin is controlled by the xanthophyll cycle in plant photosynthetic tissues, which is a collection of light and dark control

*Schematic representation of singlet oxygen formation upon excitation of chlorophyll (Chl) and the role of carotenoids (car) in protection against photooxidative damage. The symbol \* indicates excited states.*

Under high light conditions, violaxanthin de-epoxidase activated and converted violaxanthin to zeaxanthin. In dark conditions zeaxanthin is converted into violax-

**62**

**Figure 6.**

*The xanthophyll cycle.*

Catalase has a tetramer enzyme that breaks H2O2 into water and oxygen. Catalase has a lower affinity for H2O2, so it can remove H2O2 at high concentrations [17]. Hydrogen peroxide is very toxic to plant cells, especially in the chloroplast. Hydrogen peroxide at very low concentrations prevents the activity of the enzymes of the calvin cycle, especially the enzymes with *sulfhydryl* group such as glyceraldehyde 3-phosphate dehydrogenase and fructose 1,6 bisphosphatase [18, 19].

## **10. Peroxidases**

Peroxides are a group of antioxidant enzymes that cause the decomposition of hydrogen peroxide with the oxidation of a substance. Peroxidases are located in the cytosol, vacuole, chloroplast, and extracellular space and are classified based on their combined composition.

## **11. Ascorbate peroxidase**

Ascorbate peroxidase is an antioxidant enzyme that participates in the ascorbate-glutathione cycle, and its activity has been reported in the chloroplast, cytosol, peroxisome, and apoplast. This enzyme uses ascorbate as a reducing agent and decomposes hydrogen peroxide into water and oxygen [20]. The high concentration of ascorbate peroxidase to hydrogen peroxide shows that the ascorbate-glutathione cycle plays a vital role in controlling the level of radicals in cellular organs. In ascorbate-glutathione cycle with ascorbate peroxidase enzyme activity, ascorbate is oxidized to monodehydroascorbate, and ascorbate production is required to continue the cycle. In this cycle, the enzymes of monodehydroascorbate reductase (MADAR), dehydroascorbate reductase (DHAR), and glutathione reductase (GR) are active and reduce ascorbate using water and glutathione. Using NADPH, monodehydroascorbate reductase converts monodehydroascorbate to ascorbate. While dehydroascorbate reductase catalyzes dehydroascorbate to ascorbate using glutathione (GSH) oxidation.

## **12. Guaiacolperoxidase**

Guaiacol peroxidase oxidize guaiacol. This enzyme is also present in the cytosol, vacuole, and cell wall [21].

## **13. Superoxide dismutase (SOD)**

Superoxide dismutase is one of the enzymes that is located in all intracellular organs and apoplast and is very important in the defense against active oxygen species. This enzyme converts the radical superoxide to H2O2, which H2O2 should be detoxified during the next stages of antioxidant defense. In the presence of the superoxide dismutase enzyme, this reaction occurs 10,000 times faster [22].

**Figure 7.** *Formation and elimination of reactive oxygen species.*

This enzyme is divided into three groups based on its cofactor:


There are genome types of SOD in the nucleus that are synthesized in the cytoplasm and then transmitted to different organs [23]. In the chloroplast, the superoxide dismutase enzyme is in two forms attached to the thylakoid membrane and is free in the stroma. The forms attached to the thylakoid membrane and the free enzyme, at the site of production and inward stroma, convert radical superoxide into H2O2 [24].

In summary, the activity of enzymes was shown in **Figure 7**.

## **14. Lipids and oxidation**

Lipids are major constituents of prokaryotic and eukaryotic membranes. Besides serving as structural components of the plasma membrane and intracellular membranes, they provide diverse biological functions in energy and carbon storage, signal transduction, and stress responses. Plants contain a diverse set of lipids including fatty acids, phospholipids, glycolipids, sterol lipids, sphingolipids, and waxes. Polyunsaturated fatty acids (PUFAs) are lipid components commonly and easily oxidized by unbalanced ROS (mainly hydroxyl radical due to its indiscriminative reactive character).

## **15. Effect of reactive oxygen species on lipids**

Cell membrane is one of the primary goals of many environmental stresses. Therefore, maintaining the integrity and stability of the membrane under stress is one of the signs of stress tolerance [25]. Polyunsaturated fatty acids are one of the most important membrane lipid compounds that are very sensitive to peroxidation. The main reason for the harmful effects of ROS is their ability to start the chain reaction of oxidation of unsaturated fatty acids, which leads to lipid peroxidation and membrane degradation.

The reaction is divided into three major steps: initiation, propagation, and termination.

#### **Initiation**

A radical fatty acid is produced at the initiation stage. Oxygen reactive species (ROS), such as OH, combines with hydrogen atom of unsaturated fatty acid to produce water and radical fatty acids (**Figure 8**).

**65**

**Propagation**

**Figure 9.**

**Figure 8.**

**Termination**

and different radical fatty acid (**Figure 9**).

*The propagation phase of peroxidation of unsaturated fatty acids [26].*

**16. Formation malondialdehyde (MDA)**

when two radicals react together. So the radical reaction stops.

The stability of the fatty acid radical is low, so that it reacts with molecular oxygen, resulting in the formation of peroxyl-fatty acid radical. This radical also has low stability, which reacts with another free fatty acid and produces a lipid peroxide

It always produces another radical, while a radical reacts with a non-radical. This process is called the "chain reaction mechanism." A non-radical species is produced

As previously mentioned, the polyunsaturated fatty acid (PUFA) acyl chain is attacked by free radicals and creates a radical fat that reacts easily with an oxygen molecule and forms a lipid peroxyl radical. The lipid peroxyl radical can attack neighboring PUFAs, propagating a chain reaction. A linolenic acid (18:3) peroxyl radical can also react internally, forming a cyclic peroxyl radical, which spontaneously reacts with a second oxygen molecule and is subsequently reduced to phytoprostane G1 (PPG1). Phytoprostane G1 (PPG1) either spontaneously decays, forming MDA and

other alkanes and alkenes, or forms other phytoprostanes [27] (**Figure 10**).

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

*The initiation phase of peroxidation of unsaturated fatty acids [26].*

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

**Figure 8.** *The initiation phase of peroxidation of unsaturated fatty acids [26].*

#### **Figure 9.**

*Advances in Lipid Metabolism*

**Figure 7.**

This enzyme is divided into three groups based on its cofactor:

In summary, the activity of enzymes was shown in **Figure 7**.

(mainly hydroxyl radical due to its indiscriminative reactive character).

**15. Effect of reactive oxygen species on lipids**

produce water and radical fatty acids (**Figure 8**).

There are genome types of SOD in the nucleus that are synthesized in the cytoplasm and then transmitted to different organs [23]. In the chloroplast, the superoxide dismutase enzyme is in two forms attached to the thylakoid membrane and is free in the stroma. The forms attached to the thylakoid membrane and the free enzyme, at the site of production and inward stroma, convert radical superoxide into H2O2 [24].

Lipids are major constituents of prokaryotic and eukaryotic membranes. Besides serving as structural components of the plasma membrane and intracellular membranes, they provide diverse biological functions in energy and carbon storage, signal transduction, and stress responses. Plants contain a diverse set of lipids including fatty acids, phospholipids, glycolipids, sterol lipids, sphingolipids, and waxes. Polyunsaturated fatty acids (PUFAs) are lipid components commonly and easily oxidized by unbalanced ROS

Cell membrane is one of the primary goals of many environmental stresses. Therefore, maintaining the integrity and stability of the membrane under stress is one of the signs of stress tolerance [25]. Polyunsaturated fatty acids are one of the most important membrane lipid compounds that are very sensitive to peroxidation. The main reason for the harmful effects of ROS is their ability to start the chain reaction of oxidation of unsaturated fatty acids, which leads to lipid peroxidation

The reaction is divided into three major steps: initiation, propagation, and

A radical fatty acid is produced at the initiation stage. Oxygen reactive species (ROS), such as OH, combines with hydrogen atom of unsaturated fatty acid to

1.Cu/Zn SOD in the chloroplasts and cytosol

2.Fe/SOD in the chloroplasts of some plants

3.Mn/SOD in the mitochondrial matrix

*Formation and elimination of reactive oxygen species.*

**14. Lipids and oxidation**

and membrane degradation.

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termination. **Initiation** *The propagation phase of peroxidation of unsaturated fatty acids [26].*

#### **Propagation**

The stability of the fatty acid radical is low, so that it reacts with molecular oxygen, resulting in the formation of peroxyl-fatty acid radical. This radical also has low stability, which reacts with another free fatty acid and produces a lipid peroxide and different radical fatty acid (**Figure 9**).

#### **Termination**

It always produces another radical, while a radical reacts with a non-radical. This process is called the "chain reaction mechanism." A non-radical species is produced when two radicals react together. So the radical reaction stops.

### **16. Formation malondialdehyde (MDA)**

As previously mentioned, the polyunsaturated fatty acid (PUFA) acyl chain is attacked by free radicals and creates a radical fat that reacts easily with an oxygen molecule and forms a lipid peroxyl radical. The lipid peroxyl radical can attack neighboring PUFAs, propagating a chain reaction. A linolenic acid (18:3) peroxyl radical can also react internally, forming a cyclic peroxyl radical, which spontaneously reacts with a second oxygen molecule and is subsequently reduced to phytoprostane G1 (PPG1). Phytoprostane G1 (PPG1) either spontaneously decays, forming MDA and other alkanes and alkenes, or forms other phytoprostanes [27] (**Figure 10**).

## **17. Malondialdehyde (MDA) assay**

Lipid peroxidation was measured according to Heath and Packer (1968) by measuring the concentration of MDA. According to this method, 0.2 g of tissue was homogenized in 2 ml 0.1% (w:v) trichloroacetic acid (TCA) solution. The homogenate was centrifuged at 10,000 g (rcf) at 4°C for 10 min and 2 ml of supernatant transfer to new tube and then added 1 ml 20% TCA containing 0.5% (w:v) thiobarbituric acid (TBA). The reaction mixture was incubated in boiling water for 30 min at 95°C followed by placing the tubes on an ice bath to stop the reaction. The homogenate was centrifuged at 10,000 g for 15 min, and the absorbance was read at 532 nm [28]. The unspecific turbidity was corrected by A600 subtracting from A530. The amount of MDA–TBA complex (red pigment) was expressed as μmol/g FW and calculated by the extinction coefficient 155 mM − 1 cm − 1 using the formula (**Figure 11**).

*MDA* = (*A*<sup>532</sup> <sup>−</sup> *<sup>A</sup>*600) \_\_\_\_\_\_\_\_\_\_\_\_ 155 × *Amount of reaction mixture* \_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_ *tissue*

**Figure 11.** *The formation of MDA–TBA complex [29].*

### **18. Radical scavenging assays**

DPPH• is a stable free radical with purple color and a strong absorption band in the range of 515–520 nm. DPPH takes an electron or a hydrogen atom from an antioxidant accumulation molecule to become a stable DPPH molecule, in the presence of antioxidant compounds. The reduced form of DPPH is pale yellow. By studying spectrophotometric color changes, it may determine the antioxidant activity.

**67**

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

of DPPH•

antioxidant.

antioxidant.

**19. ABTS+**

[33, 34].

nanotechnology.

from the following formula:

 **assay**

to measure the antioxidant capacity.

control is measured at 415 nm [32].

of ion pumps dependent on H+

**21. Benefits of nanotechnology**

An antioxidant compound with a larger free radical scavenging capacity reduces DPPH further. Therefore, there is less purple color in the sample. The DPPH assay was performed according to the method developed by Blois (1958) slightly modified by

methanol was stirred. Then a standard or sample (50 mL) was added with 2.95 mL

DPPH scavenging effects(%) = \_

A0: The absorbance is at 515 nm of the radical (DPPH) in the absence of

A1: The absorbance is at 515 nm of the radical (DPPH) in the presence of

Another important evaluation for antioxidant activity is the ABTS+

ABTS•+ is intensely colored (dark green). Reduction of color ABTS•+ radical is used

By using spectrophotometer, a decrease in absorbance by test compound and

The lipoxygenase enzyme is one of the oxidative enzymes. This enzyme catalyzes the addition of molecular oxygen to unsaturated fatty acids, produces unsaturated fatty hydroperoxides, and accelerates lipid peroxidation. Free radicals produced by lipoxygenase cause irregularities in the selective membrane permeability. This irregularity leads to an increase in ion leakage, a decrease in the activity

Nanomaterials can offer many applications in mechanical industries especially in coating, lubricants, and adhesive applications. The magnetic nanoparticles such as Fe3O4 are employed in the biomedical and clinical fields. TiO2 nanoparticles find an application in cosmetics, pigments, sunscreen products, solar cells, and photocatalysis [35]. However, human beings must take caution in using nanoparticles and

assay, by peroxyl radicals or other oxidants, ABTS+

**20. The role of enzymes in peroxidation of lipids**

Decrease of absorbance was read at 517 nm. DPPH scavenging effect is obtained

*A*0 − *A*1 *A*0

ATPase, and changes in the cell membrane potential

× 100

in 80% (v/v)

test. In this

is oxidized to its radical cation.

Brand-Williams et al. [30, 31]. For 40 min, a solution of 1 mM DPPH•

solution and placed for 30 min in the dark.

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

An antioxidant compound with a larger free radical scavenging capacity reduces DPPH further. Therefore, there is less purple color in the sample. The DPPH assay was performed according to the method developed by Blois (1958) slightly modified by Brand-Williams et al. [30, 31]. For 40 min, a solution of 1 mM DPPH• in 80% (v/v) methanol was stirred. Then a standard or sample (50 mL) was added with 2.95 mL of DPPH• solution and placed for 30 min in the dark.

Decrease of absorbance was read at 517 nm. DPPH scavenging effect is obtained from the following formula: DPPH scavenging effects(%) = \_

*A*0 − *A*1 *A*0 × 100

A0: The absorbance is at 515 nm of the radical (DPPH) in the absence of antioxidant.

A1: The absorbance is at 515 nm of the radical (DPPH) in the presence of antioxidant.

#### **19. ABTS+ assay**

*Advances in Lipid Metabolism*

**17. Malondialdehyde (MDA) assay**

**18. Radical scavenging assays**

*The formation of MDA–TBA complex [29].*

Lipid peroxidation was measured according to Heath and Packer (1968) by measuring the concentration of MDA. According to this method, 0.2 g of tissue was homogenized in 2 ml 0.1% (w:v) trichloroacetic acid (TCA) solution. The homogenate was centrifuged at 10,000 g (rcf) at 4°C for 10 min and 2 ml of supernatant transfer to new tube and then added 1 ml 20% TCA containing 0.5% (w:v) thiobarbituric acid (TBA). The reaction mixture was incubated in boiling water for 30 min at 95°C followed by placing the tubes on an ice bath to stop the reaction. The homogenate was centrifuged at 10,000 g for 15 min, and the absorbance was read at 532 nm [28]. The unspecific turbidity was corrected by A600 subtracting from A530. The amount of MDA–TBA complex (red pigment) was expressed as μmol/g FW and calculated by

the extinction coefficient 155 mM − 1 cm − 1 using the formula (**Figure 11**).

\_\_\_\_\_\_\_\_\_\_\_\_ 155 ×

*Amount of reaction mixture* \_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_\_ *tissue*

is a stable free radical with purple color and a strong absorption band in

the range of 515–520 nm. DPPH takes an electron or a hydrogen atom from an antioxidant accumulation molecule to become a stable DPPH molecule, in the presence of antioxidant compounds. The reduced form of DPPH is pale yellow. By studying spectrophotometric color changes, it may determine the antioxidant activity.

*MDA* = (*A*<sup>532</sup> <sup>−</sup> *<sup>A</sup>*600)

**66**

DPPH•

**Figure 11.**

**Figure 10.**

*Lipid peroxidation [27].*

Another important evaluation for antioxidant activity is the ABTS<sup>+</sup> test. In this assay, by peroxyl radicals or other oxidants, ABTS+ is oxidized to its radical cation. ABTS•+ is intensely colored (dark green). Reduction of color ABTS•+ radical is used to measure the antioxidant capacity.

By using spectrophotometer, a decrease in absorbance by test compound and control is measured at 415 nm [32].

## **20. The role of enzymes in peroxidation of lipids**

The lipoxygenase enzyme is one of the oxidative enzymes. This enzyme catalyzes the addition of molecular oxygen to unsaturated fatty acids, produces unsaturated fatty hydroperoxides, and accelerates lipid peroxidation. Free radicals produced by lipoxygenase cause irregularities in the selective membrane permeability. This irregularity leads to an increase in ion leakage, a decrease in the activity of ion pumps dependent on H+ ATPase, and changes in the cell membrane potential [33, 34].

### **21. Benefits of nanotechnology**

Nanomaterials can offer many applications in mechanical industries especially in coating, lubricants, and adhesive applications. The magnetic nanoparticles such as Fe3O4 are employed in the biomedical and clinical fields. TiO2 nanoparticles find an application in cosmetics, pigments, sunscreen products, solar cells, and photocatalysis [35]. However, human beings must take caution in using nanoparticles and nanotechnology.

*Advances in Lipid Metabolism*

## **Author details**

Shahla Hashemi1,2

1 Biology Department, Faculty of Science, Shahid Bahonar University of Kerman, Iran

2 Young Researcher's Society, Shahid Bahonar University of Kerman, Kerman, Iran

\*Address all correspondence to: shahlahashemi15@yahoo.com

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

**69**

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

> radicals, and singlet oxygen. Journal of Agricultural and Food Chemistry.

2000;**48**(11):5677-5684

2018;**69**(1):97-109

[11] Chung I-M, Rajakumar G, Thiruvengadam M. Effect of silver nanoparticles on phenolic compounds production and biological activities in hairy root cultures of *Cucumis anguria*. Acta Biologica Hungarica.

[12] Bendary E, Francis R, Ali H, Sarwat M, El Hady S. Antioxidant and structure–activity relationships (Sars) of some phenolic and anilines compounds. Annals of Agricultural

Science. 2013;**58**(2):173-181

Plant. 2018;**11**(1):58-74

[13] Ma X, Li H, Dong J, Qian W. Determination of hydrogen peroxide scavenging activity of phenolic acids by employing gold nanoshells precursor composites as nanoprobes. Food Chemistry. 2011;**126**(2):698-704

[14] Sun T, Yuan H, Cao H, Yazdani M, Tadmor Y, Li L. Carotenoid metabolism in plants: The role of plastids. Molecular

[15] Nisar N, Li L, Lu S, Khin NC, Pogson BJ. Carotenoid metabolism in plants. Molecular Plant. 2015;**8**(1):68-82

[16] Demmig-Adams B, Adams WW. Antioxidants in photosynthesis and human nutrition. Science. 2002;**298**(5601):2149-2153

[17] Wang W-B, Kim Y-H, Lee H-S, Kim K-Y, Deng X-P, Kwak S-S. Analysis of antioxidant enzyme activity during germination of alfalfa under salt and drought stresses. Plant Physiology and Biochemistry. 2009;**47**(7):570-577

[18] Pan Y, Wu LJ, Yu ZL. Effect of salt and drought stress on antioxidant enzymes activities and SOD isoenzymes of liquorice (Glycyrrhiza uralensis

[1] Peralta-Videa JR, Zhao L, Lopez-Moreno ML, de la Rosa G, Hong J, Gardea-Torresdey JL. Nanomaterials and the environment: A review for the biennium 2008-2010. Journal of Hazardous Materials. 2011;**186**(1):1-15

[2] Nel A, Xia T, Mädler L, Li N. Toxic potential of materials at the nanolevel. Science. 2006;**311**(5761):622-627

[3] Yu S-W, Tang K-X. Map kinase cascades responding to environmental stress in plants. Acta Botanica Sinica.

[4] Hashim S, Aisha AF, Majid AMSA, Ismail Z. A Validated Rp-Hplc Method for Quantification of Alpha-Tocopherol in *Elaeis guineensis* Leaf Extracts. 2013

[5] Sharma P, Dubey RS. Drought induces oxidative stress and enhances the activities of antioxidant enzymes in growing rice seedlings. Plant Growth Regulation. 2005;**46**(3):209-221

[6] Locato V, Cimini S, De Gara L. Strategies to increase vitamin C in plants: From plant defense perspective to food biofortification. Frontiers in

Plant Science. 2013;**4**:152

Trends in Plant Science. 2002;**7**(9):405-410

1988;**27**(4):969-978

[7] Mittler R. Oxidative stress, antioxidants and stress tolerance.

[8] Larson RA. The antioxidants of higher plants. Phytochemistry.

[9] Asada K. The water-water cycle in chloroplasts: Scavenging of active oxygens and dissipation of excess photons. Annual Review of Plant Biology. 1999;**50**(1):601-639

[10] Wang SY, Jiao H. Scavenging capacity of berry crops on superoxide radicals, hydrogen peroxide, hydroxyl

2004;**46**(2):127-136

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*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

## **References**

*Advances in Lipid Metabolism*

**68**

Iran

**Author details**

Shahla Hashemi1,2

1 Biology Department, Faculty of Science, Shahid Bahonar University of Kerman,

2 Young Researcher's Society, Shahid Bahonar University of Kerman, Kerman, Iran

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

\*Address all correspondence to: shahlahashemi15@yahoo.com

provided the original work is properly cited.

[1] Peralta-Videa JR, Zhao L, Lopez-Moreno ML, de la Rosa G, Hong J, Gardea-Torresdey JL. Nanomaterials and the environment: A review for the biennium 2008-2010. Journal of Hazardous Materials. 2011;**186**(1):1-15

[2] Nel A, Xia T, Mädler L, Li N. Toxic potential of materials at the nanolevel. Science. 2006;**311**(5761):622-627

[3] Yu S-W, Tang K-X. Map kinase cascades responding to environmental stress in plants. Acta Botanica Sinica. 2004;**46**(2):127-136

[4] Hashim S, Aisha AF, Majid AMSA, Ismail Z. A Validated Rp-Hplc Method for Quantification of Alpha-Tocopherol in *Elaeis guineensis* Leaf Extracts. 2013

[5] Sharma P, Dubey RS. Drought induces oxidative stress and enhances the activities of antioxidant enzymes in growing rice seedlings. Plant Growth Regulation. 2005;**46**(3):209-221

[6] Locato V, Cimini S, De Gara L. Strategies to increase vitamin C in plants: From plant defense perspective to food biofortification. Frontiers in Plant Science. 2013;**4**:152

[7] Mittler R. Oxidative stress, antioxidants and stress tolerance. Trends in Plant Science. 2002;**7**(9):405-410

[8] Larson RA. The antioxidants of higher plants. Phytochemistry. 1988;**27**(4):969-978

[9] Asada K. The water-water cycle in chloroplasts: Scavenging of active oxygens and dissipation of excess photons. Annual Review of Plant Biology. 1999;**50**(1):601-639

[10] Wang SY, Jiao H. Scavenging capacity of berry crops on superoxide radicals, hydrogen peroxide, hydroxyl radicals, and singlet oxygen. Journal of Agricultural and Food Chemistry. 2000;**48**(11):5677-5684

[11] Chung I-M, Rajakumar G, Thiruvengadam M. Effect of silver nanoparticles on phenolic compounds production and biological activities in hairy root cultures of *Cucumis anguria*. Acta Biologica Hungarica. 2018;**69**(1):97-109

[12] Bendary E, Francis R, Ali H, Sarwat M, El Hady S. Antioxidant and structure–activity relationships (Sars) of some phenolic and anilines compounds. Annals of Agricultural Science. 2013;**58**(2):173-181

[13] Ma X, Li H, Dong J, Qian W. Determination of hydrogen peroxide scavenging activity of phenolic acids by employing gold nanoshells precursor composites as nanoprobes. Food Chemistry. 2011;**126**(2):698-704

[14] Sun T, Yuan H, Cao H, Yazdani M, Tadmor Y, Li L. Carotenoid metabolism in plants: The role of plastids. Molecular Plant. 2018;**11**(1):58-74

[15] Nisar N, Li L, Lu S, Khin NC, Pogson BJ. Carotenoid metabolism in plants. Molecular Plant. 2015;**8**(1):68-82

[16] Demmig-Adams B, Adams WW. Antioxidants in photosynthesis and human nutrition. Science. 2002;**298**(5601):2149-2153

[17] Wang W-B, Kim Y-H, Lee H-S, Kim K-Y, Deng X-P, Kwak S-S. Analysis of antioxidant enzyme activity during germination of alfalfa under salt and drought stresses. Plant Physiology and Biochemistry. 2009;**47**(7):570-577

[18] Pan Y, Wu LJ, Yu ZL. Effect of salt and drought stress on antioxidant enzymes activities and SOD isoenzymes of liquorice (Glycyrrhiza uralensis

fisch). Plant Growth Regulation. 2006;**49**(2-3):157-165

[19] Takeda T, Yokota A, Shigeoka S. Resistance of photosynthesis to hydrogen peroxide in algae. Plant and Cell Physiology. 1995;**36**(6):1089-1095

[20] Sgherri CLM, Maffei M, Navari-Izzo F. Antioxidative enzymes in wheat subjected to increasing water deficit and rewatering. Journal of Plant Physiology. 2000;**157**(3):273-279

[21] Jaleel CA, Riadh K, Gopi R, Manivannan P, Ines J, Al-Juburi HJ, et al. Antioxidant defense responses: Physiological plasticity in higher plants under abiotic constraints. Acta Physiologiae Plantarum. 2009;**31**(3):427-436

[22] Blokhina O, Virolainen E, Fagerstedt KV. Antioxidants, oxidative damage and oxygen deprivation stress: A review. Annals of Botany. 2003;**91**(2):179-194

[23] Weisiger RA, Fridovich I. Mitochondrial superoxide dismutase site of synthesis and intramitochondrial localization. Journal of Biological Chemistry. 1973;**248**(13):4793-4796

[24] Jithesh M, Prashanth S, Sivaprakash K, Parida AK. Antioxidative response mechanisms in halophytes: Their role in stress defence. Journal of Genetics. 2006;**85**(3):237

[25] Sudhakar C, Lakshmi A, Giridarakumar S. Changes in the antioxidant enzyme efficacy in two high yielding genotypes of mulberry (*Morus alba* L.) under Nacl salinity. Plant Science. 2001;**161**(3):613-619

[26] Sachdeva M, Karan M, Singh T, Dhingra S. Oxidants and antioxidants in complementary and alternative medicine: A review. Spatula DD. 2014;**4**(1):1-16

[27] Sattler SE, Mène-Saffrané L, Farmer EE, Krischke M, Mueller MJ, DellaPenna D. Nonenzymatic lipid peroxidation reprograms gene expression and activates defense markers in arabidopsis tocopheroldeficient mutants. The Plant Cell. 2006;**18**(12):3706-3720

[28] Heath RL, Packer L. Photoperoxidation in isolated chloroplasts: I. kinetics and stoichiometry of fatty acid peroxidation. Archives of Biochemistry and Biophysics. 1968;**125**(1):189-198

[29] Shafel T, Lee SH, Jun S. Food preservation technology at subzero temperatures: A review. Journal of Biosystems Engineering. 2015;**40**(3):261-270

[30] Blois MS. Antioxidant determinations by the use of a stable free radical. Nature. 1958;**181**(4617):1199

[31] Brand-Williams W, Cuvelier M-E, Berset C. Use of a free radical method to evaluate antioxidant activity. LWT- Food Science and Technology. 1995;**28**(1):25-30

[32] Re R, Pellegrini N, Proteggente A, Pannala A, Yang M, Rice-Evans C. Antioxidant activity applying an improved Abts radical cation decolorization assay. Free Radical Biology and Medicine. 1999;**26**(9-10):1231-1237

[33] Kubi J. Exogenous spermidine alters in different way membrane permeability and lipid peroxidation in water stressed barley leaves. Acta Physiologiae Plantarum. 2006;**28**(1):27-33

[34] Juan M, Rivero RM, Romero L, Ruiz JM. Evaluation of some nutritional and biochemical indicators in selecting salt-resistant tomato cultivars. Environmental and Experimental Botany. 2005;**54**(3):193-201

**71**

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

[35] Hashemi S, Asrar Z, Pourseyedi S, Nadernejad N. Investigation of Zno nanoparticles on Proline, anthocyanin

contents and photosynthetic pigments and lipid peroxidation in the soybean. IET Nanobiotechnology.

2018;**13**(1):66-70

*Effect of Nanoparticles on Lipid Peroxidation in Plants DOI: http://dx.doi.org/10.5772/intechopen.88202*

[35] Hashemi S, Asrar Z, Pourseyedi S, Nadernejad N. Investigation of Zno nanoparticles on Proline, anthocyanin contents and photosynthetic pigments and lipid peroxidation in the soybean. IET Nanobiotechnology. 2018;**13**(1):66-70

*Advances in Lipid Metabolism*

2006;**49**(2-3):157-165

2000;**157**(3):273-279

2009;**31**(3):427-436

2003;**91**(2):179-194

fisch). Plant Growth Regulation.

[27] Sattler SE, Mène-Saffrané L, Farmer EE, Krischke M, Mueller MJ, DellaPenna D. Nonenzymatic lipid peroxidation reprograms gene expression and activates defense markers in arabidopsis tocopheroldeficient mutants. The Plant Cell.

2006;**18**(12):3706-3720

[28] Heath RL, Packer L. Photoperoxidation in isolated chloroplasts: I. kinetics and stoichiometry of fatty acid

2015;**40**(3):261-270

1958;**181**(4617):1199

1995;**28**(1):25-30

[30] Blois MS. Antioxidant determinations by the use of a stable free radical. Nature.

[31] Brand-Williams W, Cuvelier M-E, Berset C. Use of a free radical method to evaluate antioxidant activity. LWT- Food Science and Technology.

[32] Re R, Pellegrini N, Proteggente A, Pannala A, Yang M, Rice-Evans C. Antioxidant activity applying an improved Abts radical cation decolorization assay. Free Radical Biology and Medicine. 1999;**26**(9-10):1231-1237

[33] Kubi J. Exogenous spermidine alters in different way membrane permeability and lipid peroxidation in water stressed

barley leaves. Acta Physiologiae Plantarum. 2006;**28**(1):27-33

salt-resistant tomato cultivars. Environmental and Experimental Botany. 2005;**54**(3):193-201

[34] Juan M, Rivero RM, Romero L, Ruiz JM. Evaluation of some nutritional and biochemical indicators in selecting

peroxidation. Archives of Biochemistry and Biophysics. 1968;**125**(1):189-198

[29] Shafel T, Lee SH, Jun S. Food preservation technology at subzero temperatures: A review. Journal of Biosystems Engineering.

[19] Takeda T, Yokota A, Shigeoka S. Resistance of photosynthesis to hydrogen peroxide in algae. Plant and Cell Physiology. 1995;**36**(6):1089-1095

[20] Sgherri CLM, Maffei M, Navari-Izzo F. Antioxidative enzymes in wheat subjected to increasing water deficit and rewatering. Journal of Plant Physiology.

[21] Jaleel CA, Riadh K, Gopi R, Manivannan P, Ines J, Al-Juburi HJ, et al. Antioxidant defense responses: Physiological plasticity in higher plants under abiotic constraints. Acta Physiologiae Plantarum.

[22] Blokhina O, Virolainen E,

[23] Weisiger RA, Fridovich I. Mitochondrial superoxide dismutase site of synthesis and intramitochondrial localization. Journal of Biological Chemistry.

1973;**248**(13):4793-4796

[25] Sudhakar C, Lakshmi A, Giridarakumar S. Changes in the antioxidant enzyme efficacy in two high yielding genotypes of mulberry (*Morus alba* L.) under Nacl salinity. Plant Science. 2001;**161**(3):613-619

[26] Sachdeva M, Karan M, Singh T, Dhingra S. Oxidants and antioxidants in complementary and alternative medicine: A review. Spatula DD.

2006;**85**(3):237

2014;**4**(1):1-16

Fagerstedt KV. Antioxidants, oxidative damage and oxygen deprivation stress: A review. Annals of Botany.

[24] Jithesh M, Prashanth S, Sivaprakash K, Parida AK. Antioxidative response mechanisms in halophytes: Their role in stress defence. Journal of Genetics.

**70**

**73**

**Chapter 5**

**Abstract**

**1. Introduction**

**1.1 Fatty acids (FAs)**

Fatty Acid Compositions in

Fish fermentation differs from one region to another in the world. Different types of fish, different fermentation conditions, and different fermentation processes are used, thus resulting in different fermented fish products. The most investigated fermented fish products in regard to the fatty acid contents are Kejeik from Sudan, Fseekh from Egypt, Hatahata-zushi from Japan, and Tareeh and Mehiawah from the Middle East. The results of those studies were not consistent regarding the effect of the fermentation process on the contents of saturated fatty acids (SFAs), monounsaturated fatty acids (MUFAs), and polyunsaturated fatty acids (PUFAs). Some of those studies reported an increase in the level of SFAs and a decrease in the PUFAs contents, while other studies reported the opposite. The fermentation process itself was attributed to different microorganisms such as lactic acid bacteria (LAB), halophilic bacteria, the bacterial flora of *Micrococcus* and *Bacillus* species, and a new bacillus strain named *Bacillus mojavensis-ASK*. Autolytic enzymes from

the fish were also reported to be responsible for the fermentation process.

Lipids are considered as one major group of the naturally occurring organic molecules, along with carbohydrates, proteins, and nucleic acids [1]. Lipids are characterized according to their solubility (physical property) rather than their structure, in which they are insoluble in water, but soluble in nonpolar organic solvents, such as chloroform and benzene [1, 2]. All lipids are composed of carbon, hydrogen, and oxygen atoms; however, some lipids contain phosphorus, sulfur, nitrogen, or other elements [3]. Lipids are divided into three classes, (a) triacylglycerol's (TGs), which are used as long-term energy stores such as fats and oils; (b) phospholipids (PLs), which function primarily in cell membranes; and (c) steroids, like cholesterol which is a component of animal cell membranes and a precursor in the synthesis of various steroid hormones [1, 3]. Lipids play a structural role in the cell membranes in combination with proteins to give the membranes their semipermeable property [1]. In addition, lipids give the membranes

Fatty acids are the building blocks for the majority of lipids especially TGs and PLs [3, 4]. FAs are composed of a long hydrocarbon chain (nonpolar) that is

**Keywords:** fatty acids, fish, fermentation, PUFAs

their shape and protect them from the external environment [3].

Fermented Fish Products

*Afnan Freije and Aysha Mohamed Alkaabi*

## **Chapter 5**

## Fatty Acid Compositions in Fermented Fish Products

*Afnan Freije and Aysha Mohamed Alkaabi*

## **Abstract**

Fish fermentation differs from one region to another in the world. Different types of fish, different fermentation conditions, and different fermentation processes are used, thus resulting in different fermented fish products. The most investigated fermented fish products in regard to the fatty acid contents are Kejeik from Sudan, Fseekh from Egypt, Hatahata-zushi from Japan, and Tareeh and Mehiawah from the Middle East. The results of those studies were not consistent regarding the effect of the fermentation process on the contents of saturated fatty acids (SFAs), monounsaturated fatty acids (MUFAs), and polyunsaturated fatty acids (PUFAs). Some of those studies reported an increase in the level of SFAs and a decrease in the PUFAs contents, while other studies reported the opposite. The fermentation process itself was attributed to different microorganisms such as lactic acid bacteria (LAB), halophilic bacteria, the bacterial flora of *Micrococcus* and *Bacillus* species, and a new bacillus strain named *Bacillus mojavensis-ASK*. Autolytic enzymes from the fish were also reported to be responsible for the fermentation process.

**Keywords:** fatty acids, fish, fermentation, PUFAs

## **1. Introduction**

Lipids are considered as one major group of the naturally occurring organic molecules, along with carbohydrates, proteins, and nucleic acids [1]. Lipids are characterized according to their solubility (physical property) rather than their structure, in which they are insoluble in water, but soluble in nonpolar organic solvents, such as chloroform and benzene [1, 2]. All lipids are composed of carbon, hydrogen, and oxygen atoms; however, some lipids contain phosphorus, sulfur, nitrogen, or other elements [3]. Lipids are divided into three classes, (a) triacylglycerol's (TGs), which are used as long-term energy stores such as fats and oils; (b) phospholipids (PLs), which function primarily in cell membranes; and (c) steroids, like cholesterol which is a component of animal cell membranes and a precursor in the synthesis of various steroid hormones [1, 3]. Lipids play a structural role in the cell membranes in combination with proteins to give the membranes their semipermeable property [1]. In addition, lipids give the membranes their shape and protect them from the external environment [3].

#### **1.1 Fatty acids (FAs)**

Fatty acids are the building blocks for the majority of lipids especially TGs and PLs [3, 4]. FAs are composed of a long hydrocarbon chain (nonpolar) that is conjugated to a carboxyl group (polar) which is an acidic functional group [5]. FAs hydrocarbon chains range in length from 2 to 80 but commonly from 12 up to 24. Chain length from 2 to 6 are called short-chain, from 6 to 10 are called mediumchain, and 12 up to 24 are called long-chain [6]. FAs containing 16 and 18 carbons are the most prevalent. The majority of FAs have an even number of carbon atoms because they are synthesized by combining the C2 units of acetyl CoA [7, 8]. FAs are usually synthesized by the enzyme fatty acid synthase that is responsible to convert acetyl CoA into fatty acid [5]. The hydrocarbon chains of FAs are usually unbranched and can be divided into saturated and unsaturated [9]. Saturated fatty acids (SFAs) have no double bond in their hydrocarbon chain, while unsaturated fatty acids (UFAs) have one or more double bonds in their chain. These double bonds cause the formation of bents or "kinks" in the fatty acid chains making them liquid at room temperature [3].

UFAs are divided into monounsaturated fatty acids (MUFAs) which have one double bond in their chain and polyunsaturated fatty acids (PUFAs) which have two or more double bonds [10]. The double bonds locations follow a unique pattern; the MUFAs usually have the double bond between carbons 9 and 10 (Δ<sup>9</sup> ) where C1 is the carboxyl carbon. The second double bond in PUFAs is mostly between carbons 12 and 13 (Δ12). PUFAs do not normally have conjugated double bonds (─CH〓CH─CH〓CH─), instead their double bonds are usually separated by at least one methylene group (─CH〓CH─CH2─CH〓CH─) [7–10]. The stereochemistry of the double bond in the naturally occurring UFAs is usually *cis*, i.e., the two hydrogens on the carbon atoms of the double bond are on the same side of the molecule [9, 10]. *Trans* FAs are formed during the hydrogenation process of UFAs to produce SFAs [2]. In the *trans*isomers, the two hydrogens on the carbon atoms of the double bond are on the opposite sides of the molecule [10].

### **1.2 Nomenclature of fatty acids**

Fatty acids can be named using (a) systematic naming addressed in detail by the International Union of Pure and Applied Chemists and the International Union of Biochemistry and Molecular Biology (IUPAC-IUBMB) and (b) common naming [4]. The systematic naming of FAs ends with the suffix "oic acid" on to the name of the parent hydrocarbon. However, the ionized form of the fatty acid at physiological pH ends with the suffix "ate" rather than "oic acid." FAs are named according to the total number of carbon atoms, in addition to the number and position of double bonds, if present. The carbon atoms in FAs are numbered from the carboxylic acid residue (C1), and the position of the double bond is described by the symbol delta (Δ) followed by the number of the first carbon involved in the bond. For example, the full systematic name for palmitoleic acid (common name) is *cis*-Δ<sup>9</sup> hexadecenoic acid, and it is written as 16:1(Δ<sup>9</sup> ) in which it consists of 16 carbons in its hydrocarbon chain with a double bond positioned between carbons 9 and 10 [10].

The common names for FAs reflect their prominent food sources such as palmitic acid (C16:0) found in palm oil and *arachidonic* acid (*cis*5, *cis*8, *cis*11, *cis*14–20:4; from *arachis*, meaning legume or peanut) found in peanut butter [11]. The common names are much simpler than the systematic names; however, they do not give any information about the structure of the fatty acids.

Omega (ω, n) system is an alternative system of naming fatty acids. In this system, carbon atoms are numbered relative to the methyl end of the molecule. Omega system can be distinguished from the IUPAC naming system in the bases of the following: (a) omega nomenclature is only applied to unsaturated fatty acids, (b) omega system does not identify whether the double bond have *cis* or *trans*

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*Fatty Acid Compositions in Fermented Fish Products DOI: http://dx.doi.org/10.5772/intechopen.90110*

to produce long-chain polyunsaturated fatty acids [14].

ω-6 fatty acid in omega system [11].

*1.3.1 Omega-3 and omega-6 fatty acids*

members of the family (**Figure 1**) [12, 15].

been around 10:1 to 20:1 [17].

(steatohepatitis)] [18].

seeds such as flaxseeds, walnuts, soybeans [12].

*1.3.2 The role of omega-3 and omega-6 fatty acids*

biologically active molecules that regulate body functions [12].

**1.3 Essential fatty acids (EFAs)**

configuration, and (c) omega system does not identify the location of other double bonds in the molecule. For example, linoleic acid C18:2n-6(Δ9,12) can be written as

Fatty acids that the body needs but cannot synthesize in sufficient amounts to meet physiological needs due to the absence of the required enzymes are called essential fatty acids. The body cannot synthesize two polyunsaturated fatty acids: linoleic acid (C18:2n6, LA) and alpha-linolenic acid (C18:3n-3, ALA); therefore, they must be supplied by the diet [12]. Animal cells are unable to introduce double bonds in the n-3 and n-6 positions because they are deficient in certain required desaturase enzymes. However, these cells have the ability to introduce double bonds into all other positions in fatty acid hydrocarbon chain [13]. Dietary EFAs are used

Although omega-6 FAs were the first to be described as an essential fatty acids in the 1920s, omega-3 FAs take more attention than omega-6 [13]. Linolenic acid (ω-3) is the parent of the omega-3 FA family in which it can be used to produce other

Alpha-linolenic acid is the precursor for linolenic acid that is used to synthesize two active biological components: eicosapentaenoic acid (C20:5n-3, EPA) and docosahexaenoic acid (C22:6n-3, DHA) [12, 14, 16]. ALA is found in plant oils, nuts, and

Aquatic ecosystems are the principal sources of DHA and EPA in the biosphere provided by the fish and fish oils in human diet [12, 16]. On the other hand, linoleic acid can be used to produce other members of the omega-6 family like arachidonic acid (C20:4n-6, AA) that acts as a starting material for a number of eicosanoids, i.e.,

Omega-6 FAs are found in seeds, nuts, and vegetable oils of corn, sesame, and sunflower [12]. Before the industrialization, the ratio of omega-6 to omega-3 (n-6/n-3) was around 1:1 to 2:1 due the abundant consumption of vegetables and seafood high in omega-3 fatty acids. However, there is a gradual change in this ratio with the industrialization, mainly due to the consumption of refined oils and seeds with a high content of omega-6 fatty acids in which the (n-6/n-3) average ratio has

This imbalanced n-6/n-3 ratio can even be worsened by the overnutrition habit associated with the Western diet around the world. Overnutrition, associated with an increased amount of FAs made available for oxidation in the liver, favors the pro-inflammatory state due to the depletion of n-3 long-chain PUFA (n-3 LCPUFA) such as EPA and DHA, elevated n-6/n-3 ratio, hyperinsulinemia, and insulin resistance (IR). Such changes may result in the development of nonalcoholic fatty liver disease (NAFLD), steatosis [hepatocyte triacylglycerol accumulation, and cirrhosis

The consumption of omega-3, omega-6 PUFAs, and their derivatives has various beneficial effects, ranging from fetal development to cancer prevention [19]. PUFAs have a preventive effect against arterial hypertension, asthma, inflammatory

diseases, human breast cancer, and disorders of the immune system [20].

configuration, and (c) omega system does not identify the location of other double bonds in the molecule. For example, linoleic acid C18:2n-6(Δ9,12) can be written as ω-6 fatty acid in omega system [11].

## **1.3 Essential fatty acids (EFAs)**

*Advances in Lipid Metabolism*

liquid at room temperature [3].

**1.2 Nomenclature of fatty acids**

(common name) is *cis*-Δ<sup>9</sup>

between carbons 9 and 10 [10].

conjugated to a carboxyl group (polar) which is an acidic functional group [5]. FAs hydrocarbon chains range in length from 2 to 80 but commonly from 12 up to 24. Chain length from 2 to 6 are called short-chain, from 6 to 10 are called mediumchain, and 12 up to 24 are called long-chain [6]. FAs containing 16 and 18 carbons are the most prevalent. The majority of FAs have an even number of carbon atoms because they are synthesized by combining the C2 units of acetyl CoA [7, 8]. FAs are usually synthesized by the enzyme fatty acid synthase that is responsible to convert acetyl CoA into fatty acid [5]. The hydrocarbon chains of FAs are usually unbranched and can be divided into saturated and unsaturated [9]. Saturated fatty acids (SFAs) have no double bond in their hydrocarbon chain, while unsaturated fatty acids (UFAs) have one or more double bonds in their chain. These double bonds cause the formation of bents or "kinks" in the fatty acid chains making them

UFAs are divided into monounsaturated fatty acids (MUFAs) which have one double bond in their chain and polyunsaturated fatty acids (PUFAs) which have two or more double bonds [10]. The double bonds locations follow a unique pattern;

C1 is the carboxyl carbon. The second double bond in PUFAs is mostly between carbons 12 and 13 (Δ12). PUFAs do not normally have conjugated double bonds (─CH〓CH─CH〓CH─), instead their double bonds are usually separated by at least one methylene group (─CH〓CH─CH2─CH〓CH─) [7–10]. The stereochemistry of the double bond in the naturally occurring UFAs is usually *cis*, i.e., the two hydrogens on the carbon atoms of the double bond are on the same side of the molecule [9, 10]. *Trans* FAs are formed during the hydrogenation process of UFAs to produce SFAs [2]. In the *trans*isomers, the two hydrogens on the carbon atoms of the double

Fatty acids can be named using (a) systematic naming addressed in detail by the International Union of Pure and Applied Chemists and the International Union of Biochemistry and Molecular Biology (IUPAC-IUBMB) and (b) common naming [4]. The systematic naming of FAs ends with the suffix "oic acid" on to the name of the parent hydrocarbon. However, the ionized form of the fatty acid at physiological pH ends with the suffix "ate" rather than "oic acid." FAs are named according to the total number of carbon atoms, in addition to the number and position of double bonds, if present. The carbon atoms in FAs are numbered from the carboxylic acid residue (C1), and the position of the double bond is described by the symbol delta (Δ) followed by the number of the first carbon involved in the bond. For example, the full systematic name for palmitoleic acid

it consists of 16 carbons in its hydrocarbon chain with a double bond positioned

The common names for FAs reflect their prominent food sources such as palmitic acid (C16:0) found in palm oil and *arachidonic* acid (*cis*5, *cis*8, *cis*11, *cis*14–20:4; from *arachis*, meaning legume or peanut) found in peanut butter [11]. The common names are much simpler than the systematic names; however, they do

Omega (ω, n) system is an alternative system of naming fatty acids. In this system, carbon atoms are numbered relative to the methyl end of the molecule. Omega system can be distinguished from the IUPAC naming system in the bases of the following: (a) omega nomenclature is only applied to unsaturated fatty acids, (b) omega system does not identify whether the double bond have *cis* or *trans*

not give any information about the structure of the fatty acids.

hexadecenoic acid, and it is written as 16:1(Δ<sup>9</sup>

) where

) in which

the MUFAs usually have the double bond between carbons 9 and 10 (Δ<sup>9</sup>

bond are on the opposite sides of the molecule [10].

**74**

Fatty acids that the body needs but cannot synthesize in sufficient amounts to meet physiological needs due to the absence of the required enzymes are called essential fatty acids. The body cannot synthesize two polyunsaturated fatty acids: linoleic acid (C18:2n6, LA) and alpha-linolenic acid (C18:3n-3, ALA); therefore, they must be supplied by the diet [12]. Animal cells are unable to introduce double bonds in the n-3 and n-6 positions because they are deficient in certain required desaturase enzymes. However, these cells have the ability to introduce double bonds into all other positions in fatty acid hydrocarbon chain [13]. Dietary EFAs are used to produce long-chain polyunsaturated fatty acids [14].

## *1.3.1 Omega-3 and omega-6 fatty acids*

Although omega-6 FAs were the first to be described as an essential fatty acids in the 1920s, omega-3 FAs take more attention than omega-6 [13]. Linolenic acid (ω-3) is the parent of the omega-3 FA family in which it can be used to produce other members of the family (**Figure 1**) [12, 15].

Alpha-linolenic acid is the precursor for linolenic acid that is used to synthesize two active biological components: eicosapentaenoic acid (C20:5n-3, EPA) and docosahexaenoic acid (C22:6n-3, DHA) [12, 14, 16]. ALA is found in plant oils, nuts, and seeds such as flaxseeds, walnuts, soybeans [12].

Aquatic ecosystems are the principal sources of DHA and EPA in the biosphere provided by the fish and fish oils in human diet [12, 16]. On the other hand, linoleic acid can be used to produce other members of the omega-6 family like arachidonic acid (C20:4n-6, AA) that acts as a starting material for a number of eicosanoids, i.e., biologically active molecules that regulate body functions [12].

Omega-6 FAs are found in seeds, nuts, and vegetable oils of corn, sesame, and sunflower [12]. Before the industrialization, the ratio of omega-6 to omega-3 (n-6/n-3) was around 1:1 to 2:1 due the abundant consumption of vegetables and seafood high in omega-3 fatty acids. However, there is a gradual change in this ratio with the industrialization, mainly due to the consumption of refined oils and seeds with a high content of omega-6 fatty acids in which the (n-6/n-3) average ratio has been around 10:1 to 20:1 [17].

This imbalanced n-6/n-3 ratio can even be worsened by the overnutrition habit associated with the Western diet around the world. Overnutrition, associated with an increased amount of FAs made available for oxidation in the liver, favors the pro-inflammatory state due to the depletion of n-3 long-chain PUFA (n-3 LCPUFA) such as EPA and DHA, elevated n-6/n-3 ratio, hyperinsulinemia, and insulin resistance (IR). Such changes may result in the development of nonalcoholic fatty liver disease (NAFLD), steatosis [hepatocyte triacylglycerol accumulation, and cirrhosis (steatohepatitis)] [18].

## *1.3.2 The role of omega-3 and omega-6 fatty acids*

The consumption of omega-3, omega-6 PUFAs, and their derivatives has various beneficial effects, ranging from fetal development to cancer prevention [19]. PUFAs have a preventive effect against arterial hypertension, asthma, inflammatory diseases, human breast cancer, and disorders of the immune system [20].

#### **Figure 1.**

*Omega-6 and omega-3 PUFAs and their respective sources and metabolic derivatives [15].*

n-3 FAs protect against several types of cardiovascular diseases such as myocardial infarction, atherosclerosis, arrhythmia, hypertension, and human coronary artery disease [19, 21]. Moreover, consumption of n-3 FAs interferes with prostaglandin metabolism, which can reduce the platelet aggregation and adhesion to blood vessels, which reduce the blood pressure [22]. Omega-3 PUFAs can reduce the incidence of high cholesterol level in the blood, psoriasis, cancer, and arthritis [23].

The n-3 LCPUFA DHA plays a major role in the development of the nervous system during the life of fetus and neonates [24]. The study of Barrera et al. (2018) has shown that a low dietary intake of n-3 LCPUFA in pregnant Chilean women has resulted in a significant decline in their erythrocytes and breast milk DHA levels. Therefore, the improvement of the quality of FAs intake specifically DHA was recommended during pregnancy and lactation periods in order to supply adequate amount of DHA to embryos and neonates [25]. In addition, n-3 supplements intake

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*Fatty Acid Compositions in Fermented Fish Products DOI: http://dx.doi.org/10.5772/intechopen.90110*

of these compounds in the diet [15].

**1.4 Metabolism of long-chain PUFAs**

**1.5 Sources of omega-3 PUFAs**

*1.3.3 EPA and DHA*

was suggested to improve the low level of n-3 LCPUFA associated with NAFLD patients in order to lower their n-6/n-3 ratio and thus reduce the inflammatory

Omega-6 PUFAs are converted to other important compounds through various enzymatic reactions to the key intermediate arachidonic acid, which is subsequently converted to eicosanoids that act somewhat like hormones such as prostaglandins, thromboxanes, and leukotrienes [15]. Eicosanoids play an important role in muscle relaxation and contraction, blood clot formation, blood vessel contraction and dilution, blood lipid regulation, and immune response to injury and infections [12]. Omega-3 PUFAs are considered as more potent anti-inflammatory agents than n-6 PUFAs, but the effects of n-6 PUFAs are more dominant due to the abundance

Alpha-linolenic acid can be used in the body as a precursor to produce EPA; however, EPA can be directly obtained through the consumption of fish oils. Moreover, EPA can be converted to DHA and also to eicosanoids that are essential for inflammatory signaling, such as prostaglandins, thromboxanes, and leukotrienes. DHA can also be converted to eicosanoids to produce anti-inflammatory mediators such as protectins and resolvins [15]. In 2004, The International Society for the Study of Fatty Acids and Lipids (ISSFAL) has recommended the minimum intake of 0.5 g/day

ALA and LA obtained from diet can be converted into longer chains of PUFAs

by the help of two important enzymes, desaturase and elongase, that work to increase the degree of unsaturation. Elongase works by adding carbon atoms into the chain, while desaturase works to introduce double bonds by removing hydrogen atoms [27]. The n-3 and n-6 FA families compete especially at the rate-limiting Δ6 desaturase enzyme that both pathways start with. Usually desaturase enzymes display highest affinity to ALA (n-3 family) more than LA (n-6 family) [28]. The n-3 pathway starts with ALA (C18:3n-3) and ends with DHA (C22:6n-3), while the n-6 pathway starts with LA (C18:2n-6) and ends usually with AA (C20:4n-6) [27, 29]. The major difference between n-3 and n-6 pathways is that n-6 pathway usually does not proceed beyond AA; however, the n-3 pathway involves more complex steps. In the n-3 pathway, there are two elongation steps after the formation of EPA (C20:5n-3), leading to the formation of tetracosapentaenoic acid (C24:5n-3), followed by the reduction by Δ6 desaturase enzyme to produce tetracosahexaenoic acid (Nisinic acid) (C24:6n-3). The final step in n-3 pathway yields DHA (22:6n-3) by the retroconversion of (24:6n-3) to (22:6n-3) which involves peroxisomal ß-oxidation step (**Figure 2**) [27, 30]. In the mammalian cells, the FAs from (n-3) and (n-6) families cannot be interconverted because of lack of Δ12 or Δ15 desaturase

status as a treatment for the nutritional hepatic steatosis in adults [24].

of EPA and DHA for the prevention of cardiovascular disease [26].

enzymes, while the interconversion step takes place in plants [28].

Fish is a major source of animal protein diet in many countries that is easily digestible than red meat because fish flesh contains long muscle fibers [18]. Consumption of fish have many benefits for human health due to its high content of essential n-3 PUFAs, namely, EPA and DHA [14, 31, 32].The FAs content of fish varies according to diet, i.e., availability of planktons [33], environmental conditions

#### *Fatty Acid Compositions in Fermented Fish Products DOI: http://dx.doi.org/10.5772/intechopen.90110*

was suggested to improve the low level of n-3 LCPUFA associated with NAFLD patients in order to lower their n-6/n-3 ratio and thus reduce the inflammatory status as a treatment for the nutritional hepatic steatosis in adults [24].

Omega-6 PUFAs are converted to other important compounds through various enzymatic reactions to the key intermediate arachidonic acid, which is subsequently converted to eicosanoids that act somewhat like hormones such as prostaglandins, thromboxanes, and leukotrienes [15]. Eicosanoids play an important role in muscle relaxation and contraction, blood clot formation, blood vessel contraction and dilution, blood lipid regulation, and immune response to injury and infections [12].

Omega-3 PUFAs are considered as more potent anti-inflammatory agents than n-6 PUFAs, but the effects of n-6 PUFAs are more dominant due to the abundance of these compounds in the diet [15].

## *1.3.3 EPA and DHA*

*Advances in Lipid Metabolism*

**76**

**Figure 1.**

n-3 FAs protect against several types of cardiovascular diseases such as myocardial infarction, atherosclerosis, arrhythmia, hypertension, and human coronary artery disease [19, 21]. Moreover, consumption of n-3 FAs interferes with prostaglandin metabolism, which can reduce the platelet aggregation and adhesion to blood vessels, which reduce the blood pressure [22]. Omega-3 PUFAs can reduce the incidence of high cholesterol level in the blood, psoriasis, cancer, and arthritis [23]. The n-3 LCPUFA DHA plays a major role in the development of the nervous system during the life of fetus and neonates [24]. The study of Barrera et al. (2018) has shown that a low dietary intake of n-3 LCPUFA in pregnant Chilean women has resulted in a significant decline in their erythrocytes and breast milk DHA levels. Therefore, the improvement of the quality of FAs intake specifically DHA was recommended during pregnancy and lactation periods in order to supply adequate amount of DHA to embryos and neonates [25]. In addition, n-3 supplements intake

*Omega-6 and omega-3 PUFAs and their respective sources and metabolic derivatives [15].*

Alpha-linolenic acid can be used in the body as a precursor to produce EPA; however, EPA can be directly obtained through the consumption of fish oils. Moreover, EPA can be converted to DHA and also to eicosanoids that are essential for inflammatory signaling, such as prostaglandins, thromboxanes, and leukotrienes. DHA can also be converted to eicosanoids to produce anti-inflammatory mediators such as protectins and resolvins [15]. In 2004, The International Society for the Study of Fatty Acids and Lipids (ISSFAL) has recommended the minimum intake of 0.5 g/day of EPA and DHA for the prevention of cardiovascular disease [26].

## **1.4 Metabolism of long-chain PUFAs**

ALA and LA obtained from diet can be converted into longer chains of PUFAs by the help of two important enzymes, desaturase and elongase, that work to increase the degree of unsaturation. Elongase works by adding carbon atoms into the chain, while desaturase works to introduce double bonds by removing hydrogen atoms [27]. The n-3 and n-6 FA families compete especially at the rate-limiting Δ6 desaturase enzyme that both pathways start with. Usually desaturase enzymes display highest affinity to ALA (n-3 family) more than LA (n-6 family) [28]. The n-3 pathway starts with ALA (C18:3n-3) and ends with DHA (C22:6n-3), while the n-6 pathway starts with LA (C18:2n-6) and ends usually with AA (C20:4n-6) [27, 29].

The major difference between n-3 and n-6 pathways is that n-6 pathway usually does not proceed beyond AA; however, the n-3 pathway involves more complex steps. In the n-3 pathway, there are two elongation steps after the formation of EPA (C20:5n-3), leading to the formation of tetracosapentaenoic acid (C24:5n-3), followed by the reduction by Δ6 desaturase enzyme to produce tetracosahexaenoic acid (Nisinic acid) (C24:6n-3). The final step in n-3 pathway yields DHA (22:6n-3) by the retroconversion of (24:6n-3) to (22:6n-3) which involves peroxisomal ß-oxidation step (**Figure 2**) [27, 30]. In the mammalian cells, the FAs from (n-3) and (n-6) families cannot be interconverted because of lack of Δ12 or Δ15 desaturase enzymes, while the interconversion step takes place in plants [28].

## **1.5 Sources of omega-3 PUFAs**

Fish is a major source of animal protein diet in many countries that is easily digestible than red meat because fish flesh contains long muscle fibers [18]. Consumption of fish have many benefits for human health due to its high content of essential n-3 PUFAs, namely, EPA and DHA [14, 31, 32].The FAs content of fish varies according to diet, i.e., availability of planktons [33], environmental conditions

**Figure 2.**

*The elongation-desaturation pathway for the metabolism of n-6 and n-3 polyunsaturated fatty acids [30].*

[15], and seasonal variation [34]. A regular consumption of fatty fish prevents cardiovascular disease and neural disorders [35].

PUFAs are important for brain and retina development since studies with animals have shown that deficiencies in n-3 FAs decrease the concentration of DHA in the brain and retina tissues [16]. Fish and terrestrial animals are not able to synthesize n-3 or n-6 PUFAs. The primary producers for PUFAs are marine photosynthetic organisms including phytoplankton, macroalgae, and seaweeds [36].

Only some microalgae species are effective producers for EPA and DHA; therefore, aquatic ecosystems are the principal sources of these two essential PUFAs in the biosphere, in which humans obtain these FAs through the consumption of fish and other marine products (**Figure 2**) [30, 37]. Fish need PUFAs to tolerate low

**79**

*Fatty Acid Compositions in Fermented Fish Products DOI: http://dx.doi.org/10.5772/intechopen.90110*

diet rich in marine fish like Eskimos and Japanese [40].

microorganisms, normally in the presence of salt [48–50].

spoilage organisms [52].

**2.1 Fish fermentation all over the world**

*2.1.1 Asian fermented fish products*

*2.1.2 European fermented fish products*

**2. Fish fermentation in relation to fatty acids**

water temperature; thus, low amounts are expected in wormer water like tropical waters [38]. Fish oil become very popular and plays an important role in the prevention of cardiovascular disease and some type of cancers like breast, colon, and prostate [39]. It was observed that there are lower incidence of cardiovascular diseases, hypertension, and autoimmune disorders in populations that consume

Fresh fish is prone to spoilage which is caused by both microbiological and chemical reactions [41]. Lipid deterioration limits the shelf life of oily fish during storage [42]. Lipid hydrolysis induced by lipases and phospholipases produce free FAs that are susceptible to further oxidation to produce low molecular weight compounds responsible for the rancid of fish products [43]. Fish quality may decline during processing and storage mainly due to the oxidation of the PUFAs which is related to the production of unpleasant flavors and odors [44, 45].EPA and DHA are especially susceptible to oxidation during heating or other culinary treatments [46]. A long time ago, people used to preserve food either by sun drying or salting methods [47]. Salting of fish is an old-age technology that is still in use nowadays even in developing countries due to its simplicity and low cost [47, 48]. Salting fatty fish involves a certain degree of fermentation which is brought by autolytic enzymes from the fish and microorganisms in the presence of high concentrations of salt [47]. Fish fermentation is the transformation of organic substances into simpler compounds like peptides and amino acids by the action of endogenous enzymes or

Lactic acid bacteria (LAB) are used to ferment fish along with other food materials like dairy, meat, vegetables, and beverage products [51] resulting in shelf life extension and the addition of new aromas and consistencies [32]. Fermentation by lactic acid bacteria preserves food by the production of lactic acid and other organic acids, which help to reduce pH of the food and inhibit the growth of pathogenic and

Many types of fish sauce and paste are famous in Japan, Southeast Asia, and India. Traditional fermented Japanese seafood is Hatahata-zushi which is processed with boiled rice. Hatahata-zushi is prepared by soaking the sandfish (*Arctoscopus japonicas*) in water, salt, and rice vinegar, which is then fermented with boiled rice and some vegetables [53]. Bagoong is a fish paste produced in the Philippines through the neutral fermentation process of whole fish or shrimp in the presence of 20–25% salt, while Patis is a Philippine yellowish fish sauce prepared from sardines, anchovies, and shrimps [33, 50]. "Suanyu" is a Chinese low-salt fermented whole fish snack which is prepared from fresh fish mixed with cooked carbohydrates, salt, and spices [54]. Lona ilish is a salt fermented fish from India which has strong aroma mixed with some sweet, fruity, and acidic flavors with some saltiness [47].

Scandinavia is the main producer of fermented fish products in Europe. Surstromming is produced in Sweden and rakefish in Norway. These fermented fish *Fatty Acid Compositions in Fermented Fish Products DOI: http://dx.doi.org/10.5772/intechopen.90110*

*Advances in Lipid Metabolism*

**78**

**Figure 2.**

[15], and seasonal variation [34]. A regular consumption of fatty fish prevents

*The elongation-desaturation pathway for the metabolism of n-6 and n-3 polyunsaturated fatty acids [30].*

organisms including phytoplankton, macroalgae, and seaweeds [36].

PUFAs are important for brain and retina development since studies with animals have shown that deficiencies in n-3 FAs decrease the concentration of DHA in the brain and retina tissues [16]. Fish and terrestrial animals are not able to synthesize n-3 or n-6 PUFAs. The primary producers for PUFAs are marine photosynthetic

Only some microalgae species are effective producers for EPA and DHA; therefore, aquatic ecosystems are the principal sources of these two essential PUFAs in the biosphere, in which humans obtain these FAs through the consumption of fish and other marine products (**Figure 2**) [30, 37]. Fish need PUFAs to tolerate low

cardiovascular disease and neural disorders [35].

water temperature; thus, low amounts are expected in wormer water like tropical waters [38]. Fish oil become very popular and plays an important role in the prevention of cardiovascular disease and some type of cancers like breast, colon, and prostate [39]. It was observed that there are lower incidence of cardiovascular diseases, hypertension, and autoimmune disorders in populations that consume diet rich in marine fish like Eskimos and Japanese [40].

## **2. Fish fermentation in relation to fatty acids**

Fresh fish is prone to spoilage which is caused by both microbiological and chemical reactions [41]. Lipid deterioration limits the shelf life of oily fish during storage [42]. Lipid hydrolysis induced by lipases and phospholipases produce free FAs that are susceptible to further oxidation to produce low molecular weight compounds responsible for the rancid of fish products [43]. Fish quality may decline during processing and storage mainly due to the oxidation of the PUFAs which is related to the production of unpleasant flavors and odors [44, 45].EPA and DHA are especially susceptible to oxidation during heating or other culinary treatments [46].

A long time ago, people used to preserve food either by sun drying or salting methods [47]. Salting of fish is an old-age technology that is still in use nowadays even in developing countries due to its simplicity and low cost [47, 48]. Salting fatty fish involves a certain degree of fermentation which is brought by autolytic enzymes from the fish and microorganisms in the presence of high concentrations of salt [47]. Fish fermentation is the transformation of organic substances into simpler compounds like peptides and amino acids by the action of endogenous enzymes or microorganisms, normally in the presence of salt [48–50].

Lactic acid bacteria (LAB) are used to ferment fish along with other food materials like dairy, meat, vegetables, and beverage products [51] resulting in shelf life extension and the addition of new aromas and consistencies [32]. Fermentation by lactic acid bacteria preserves food by the production of lactic acid and other organic acids, which help to reduce pH of the food and inhibit the growth of pathogenic and spoilage organisms [52].

#### **2.1 Fish fermentation all over the world**

#### *2.1.1 Asian fermented fish products*

Many types of fish sauce and paste are famous in Japan, Southeast Asia, and India. Traditional fermented Japanese seafood is Hatahata-zushi which is processed with boiled rice. Hatahata-zushi is prepared by soaking the sandfish (*Arctoscopus japonicas*) in water, salt, and rice vinegar, which is then fermented with boiled rice and some vegetables [53]. Bagoong is a fish paste produced in the Philippines through the neutral fermentation process of whole fish or shrimp in the presence of 20–25% salt, while Patis is a Philippine yellowish fish sauce prepared from sardines, anchovies, and shrimps [33, 50]. "Suanyu" is a Chinese low-salt fermented whole fish snack which is prepared from fresh fish mixed with cooked carbohydrates, salt, and spices [54]. Lona ilish is a salt fermented fish from India which has strong aroma mixed with some sweet, fruity, and acidic flavors with some saltiness [47].

#### *2.1.2 European fermented fish products*

Scandinavia is the main producer of fermented fish products in Europe. Surstromming is produced in Sweden and rakefish in Norway. These fermented fish

#### *Advances in Lipid Metabolism*

are made by immersing whole herring and trout in brine (salty water) for 1–2 days, eviscerating, and retaining the roe or milt in barrels with fresh brine. The final fermented product is packed in cans and usually consumed on special occasions [49]. "Tidbits" is another Scandinavian product that is canned with vinegar, sugar, and spices after maturation. In France, the fish *Engraulis encrasicholus*is is salted to prepare anchovies, while in southern France, a fish sauce called "Pissala" is prepared from small fish of the *Engraulis* sp., *Aphya* sp., and *Gobius* sp. [49].

### *2.1.3 Middle East, African, and Asian fermented fish products*

Many types of fermented fish are prepared in the Middle East. "Fseekh" is a famous fermented fish prepared in Egypt and Sudan from different types of fish [49]. Kejeik is a fermented dried fish produce that is common in Sudan and central Africa which is powdered and thickened with okra after boiling [48]. The FA content of Bouri fish varies with fish size; in which it was much greater in small size than large size fish because of the higher activity of lipolytic enzymes in small fish as stated by the authors [48]. However, the fresh and fermented product Fseekh of both fish sizes had the same FA profile. The high salt content was found to have no effect on those enzymes responsible for the liberation of free fatty acids from the lipids [48]. The ratio of UFAs/SFAs decreased as the amount of UFAs decreased significantly after the salting and fermenting process. Moreover, all SFAs except palmitic acid (C16:0) increased significantly. The major SFA was C16:0, and the major UFAs were palmitoleic acid (C16:1n-7) and oleic acid (C18:1n-9). Appreciable amounts of PUFAs such as C18:2n-6, C18:3n-3, stearidonic acid (C18:4n-3, SDA), C20:5n-3, docosapentaenoic acid (C22:5n-3, DPA), and C22:6n-3 were also present. The presence of the odd-chain pentadecylic acid (C15:0) and heptadecanoic acid (C17:0) FAs is considered a unique characteristic of the Bouri fish oil [48].

The analysis of fatty acid compositions was done for different fermented seafoods all over the world. For example, the analysis of Hatahata-zushi which is a Japanese fermented fish product of sandfish has shown no change in the fatty acid compositions throughout the fermentation process; however, the content of SFAs and MUFAs increased, while the content of PUFAs decreased markedly during the process of fermentation. The highest concentrations of the FAs in "Hatahata-zushi" were C18:1n-9, C16:0, C22:6n-3, C20:5n-3, and C16:1n-7, respectively [53].

It has been noticed that bacterial enzymes play an important role in fish fermentation in which the aroma in fermented fish products is claimed to be derived from the activity of halophilic bacteria [47]. The enzymes that participate in the production of PUFAs are thought to be either fish tissue enzymes or microbial enzymes [47]. The traditional fermented fish product Lona ilish of Northeast India has shown that salting plays an important role in the fermentation and preservation of food. Salt preserves the fermented products through the reduction of water activity (aw) of the system, thus rendering a condition less suitable (low moisture) for the microbial growth. Generally, food pathogenic bacteria are inhibited when the water activity is 0.92 or less which is equivalent to NaCl concentration of 13% (w/v). It was reported that halophilic bacteria were responsible for the fermentation process of fish product Lona ilish since these bacteria can tolerate high salt conditions. The bacterial flora of *Micrococcus* and *Bacillus* species were reported to be involved in the processing of'Lona ilish [47]. Other microorganisms are found in the spontaneously fermented Chinese fish product Suanyu such as lactic acid bacteria, *Staphylococcus*, and yeast [54].

The two main products of fish fermentation in the Arabian Gulf region are fish paste and sauce. It is difficult to classify these products due to the lack of standardization throughout the world. Generally, fish paste is a thick product

**81**

*Fatty Acid Compositions in Fermented Fish Products DOI: http://dx.doi.org/10.5772/intechopen.90110*

Tareeh [49, 55].

**3. Conclusion**

the mystery behind such contradictions.

**Acknowledgements**

with whole fermented fish, while fish sauce is a thinner product with additives such as spices and cereals. The color of fish paste or sauce is usually brown resembling soy sauce. Fish paste is more nutritious than fish sauce [49]. Tareeh is a salt fermented fish paste in the Arabian Gulf countries prepared from a high-fat fish *Sardinella* spp. [33, 49]. The famous fish sauce in the Arabian Gulf Countries is called Mehiawah that is prepared by the addition of spices to the fermented fish

The fatty acid contents of the fish product Tareeh and Mehiawah of white sardinella (Oom) were recently investigated [56, 57]. The study of Freije et al. (2018) was conducted in order to determine the effect of fermentation under high salting conditions (20%) for 8 weeks on weekly basis on the fatty acid compositions of white sardinella (Sardinella albella) [57]. The fatty acid compositions of Tareeh have shown a great deal of variation as the fermentation process proceeded. The concentrations of SFAs and MUFAs significantly declined in Tareeh samples compared to raw fish, whereas the concentrations of UFAs and PUFAs were significantly increased starting from week 4 of fermentation. The amount of n-6 FAs as well as n-3 FAs increased during the fermentation process, and the enzymatic activities of the elongases and desaturases were found to have higher affinity to n-3 FAs than n-6 FAs. The ratio of n-6/n-3 was decreased after 8 weeks of fermentation, while the proportions of EPA + DHA were increased. This unique fermentation process might have a great application in the food industry [56]. It was also concluded that the elongation and desaturation process was carried out by single new bacterial

strain from the *Bacillus* species named *Bacillus mojavensis-ASK* [58, 59].

The studies that investigated fatty acid compositions in fermented fish products all over the world are limited. Each of those studies has given different results without any common consistency among them. Some of those studies have reported a decrease in UFAs and an increase in SFAs, while others reported increased SFAs and MUFAs but decreased PUFAs during fermentations. On the contrary, the most recent studies have reported declined levels of SFAs and MUFAs and a rapid increase in PUFAs. Such contradicted results can be attributed to the difference in the type of fermented fish, fermentation conditions, and the addition of different additives. Therefore, this process requires thorough investigations in order to reveal

The authors are grateful to the Department of Biology, College of Science, University of Bahrain, where the study was conducted and financially supported. *Fatty Acid Compositions in Fermented Fish Products DOI: http://dx.doi.org/10.5772/intechopen.90110*

*Advances in Lipid Metabolism*

are made by immersing whole herring and trout in brine (salty water) for 1–2 days, eviscerating, and retaining the roe or milt in barrels with fresh brine. The final fermented product is packed in cans and usually consumed on special occasions [49]. "Tidbits" is another Scandinavian product that is canned with vinegar, sugar, and spices after maturation. In France, the fish *Engraulis encrasicholus*is is salted to prepare anchovies, while in southern France, a fish sauce called "Pissala" is prepared from small fish of the *Engraulis* sp., *Aphya* sp., and *Gobius* sp. [49].

Many types of fermented fish are prepared in the Middle East. "Fseekh" is a famous fermented fish prepared in Egypt and Sudan from different types of fish [49]. Kejeik is a fermented dried fish produce that is common in Sudan and central Africa which is powdered and thickened with okra after boiling [48]. The FA content of Bouri fish varies with fish size; in which it was much greater in small size than large size fish because of the higher activity of lipolytic enzymes in small fish as stated by the authors [48]. However, the fresh and fermented product Fseekh of both fish sizes had the same FA profile. The high salt content was found to have no effect on those enzymes responsible for the liberation of free fatty acids from the lipids [48]. The ratio of UFAs/SFAs decreased as the amount of UFAs decreased significantly after the salting and fermenting process. Moreover, all SFAs except palmitic acid (C16:0) increased significantly. The major SFA was C16:0, and the major UFAs were palmitoleic acid (C16:1n-7) and oleic acid (C18:1n-9). Appreciable amounts of PUFAs such as C18:2n-6, C18:3n-3, stearidonic acid (C18:4n-3, SDA), C20:5n-3, docosapentaenoic acid (C22:5n-3, DPA), and C22:6n-3 were also present. The presence of the odd-chain pentadecylic acid (C15:0) and heptadecanoic acid (C17:0) FAs is considered a unique characteristic of the Bouri fish oil [48]. The analysis of fatty acid compositions was done for different fermented seafoods all over the world. For example, the analysis of Hatahata-zushi which is a Japanese fermented fish product of sandfish has shown no change in the fatty acid compositions throughout the fermentation process; however, the content of SFAs and MUFAs increased, while the content of PUFAs decreased markedly during the process of fermentation. The highest concentrations of the FAs in "Hatahata-zushi"

were C18:1n-9, C16:0, C22:6n-3, C20:5n-3, and C16:1n-7, respectively [53].

It has been noticed that bacterial enzymes play an important role in fish fermentation in which the aroma in fermented fish products is claimed to be derived from the activity of halophilic bacteria [47]. The enzymes that participate in the production of PUFAs are thought to be either fish tissue enzymes or microbial enzymes [47]. The traditional fermented fish product Lona ilish of Northeast India has shown that salting plays an important role in the fermentation and preservation of food. Salt preserves the fermented products through the reduction of water activity (aw) of the system, thus rendering a condition less suitable (low moisture) for the microbial growth. Generally, food pathogenic bacteria are inhibited when the water activity is 0.92 or less which is equivalent to NaCl concentration of 13% (w/v). It was reported that halophilic bacteria were responsible for the fermentation process of fish product Lona ilish since these bacteria can tolerate high salt conditions. The bacterial flora of *Micrococcus* and *Bacillus* species were reported to be involved in the processing of'Lona ilish [47]. Other microorganisms are found in the spontaneously fermented Chinese fish product Suanyu such as lactic acid bacteria, *Staphylococcus*,

The two main products of fish fermentation in the Arabian Gulf region are fish paste and sauce. It is difficult to classify these products due to the lack of standardization throughout the world. Generally, fish paste is a thick product

*2.1.3 Middle East, African, and Asian fermented fish products*

**80**

and yeast [54].

with whole fermented fish, while fish sauce is a thinner product with additives such as spices and cereals. The color of fish paste or sauce is usually brown resembling soy sauce. Fish paste is more nutritious than fish sauce [49]. Tareeh is a salt fermented fish paste in the Arabian Gulf countries prepared from a high-fat fish *Sardinella* spp. [33, 49]. The famous fish sauce in the Arabian Gulf Countries is called Mehiawah that is prepared by the addition of spices to the fermented fish Tareeh [49, 55].

The fatty acid contents of the fish product Tareeh and Mehiawah of white sardinella (Oom) were recently investigated [56, 57]. The study of Freije et al. (2018) was conducted in order to determine the effect of fermentation under high salting conditions (20%) for 8 weeks on weekly basis on the fatty acid compositions of white sardinella (Sardinella albella) [57]. The fatty acid compositions of Tareeh have shown a great deal of variation as the fermentation process proceeded. The concentrations of SFAs and MUFAs significantly declined in Tareeh samples compared to raw fish, whereas the concentrations of UFAs and PUFAs were significantly increased starting from week 4 of fermentation. The amount of n-6 FAs as well as n-3 FAs increased during the fermentation process, and the enzymatic activities of the elongases and desaturases were found to have higher affinity to n-3 FAs than n-6 FAs. The ratio of n-6/n-3 was decreased after 8 weeks of fermentation, while the proportions of EPA + DHA were increased. This unique fermentation process might have a great application in the food industry [56]. It was also concluded that the elongation and desaturation process was carried out by single new bacterial strain from the *Bacillus* species named *Bacillus mojavensis-ASK* [58, 59].

## **3. Conclusion**

The studies that investigated fatty acid compositions in fermented fish products all over the world are limited. Each of those studies has given different results without any common consistency among them. Some of those studies have reported a decrease in UFAs and an increase in SFAs, while others reported increased SFAs and MUFAs but decreased PUFAs during fermentations. On the contrary, the most recent studies have reported declined levels of SFAs and MUFAs and a rapid increase in PUFAs. Such contradicted results can be attributed to the difference in the type of fermented fish, fermentation conditions, and the addition of different additives. Therefore, this process requires thorough investigations in order to reveal the mystery behind such contradictions.

### **Acknowledgements**

The authors are grateful to the Department of Biology, College of Science, University of Bahrain, where the study was conducted and financially supported. *Advances in Lipid Metabolism*

## **Author details**

Afnan Freije1 \* and Aysha Mohamed Alkaabi2

1 Department of Biology, College of Science, University of Bahrain, Kingdom of Bahrain

2 University of Bahrain, Sakhir, Kingdom of Bahrain

\*Address all correspondence to: afreije@uob.edu.bh

© 2020 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

**83**

*Fatty Acid Compositions in Fermented Fish Products DOI: http://dx.doi.org/10.5772/intechopen.90110*

[1] Karp G. Cell and Molecular Biology. 5th ed. Vol. 42. Hoboken: John Wiley & [11] McGuire M, Beerman KA. Lipids in Table of Food Comparison for

[12] Sizer F, Whitney E. The Lipids: Fats, Oils, Phospholipids, and Sterols In Nutrition Concepts and Controversies. 12th ed. USA: Wadsworth, Cengage

[13] Shireman R. Essential Fatty Acids in Encyclopedia of Food Sciences and Nutrition, 2nd ed. London: Academic

Learning; 2011. pp. 150-188

Press; 2003. pp. 2169-2176

[14] Gladyshev MI, Sushchik NN, Anishchenko OV, Gubanenko GA, Demirchieva SM, Kalachova GS. Effect of boiling and frying on the content of essential polyunsaturated fatty acids in muscle tissues of four fish species. Food

Chemistry. 2007;**101**:1694-1700

[16] Rebah FB, Abdelmouleh A,

of lipid content and fatty acid

[17] Bulla MK, Simionato JI, Matsushita M, CorÓ FAG,

Schwid SR. Polyunsaturated fatty acids and their potential therapeutic role in multiple sclerosis. Nature Clinical Practice. Neurology. 2008;**5**:82-92

Kammou W, Yezza A. Seasonal variation

composition of *Sardinella aurita* from the Tunisian coast. Journal of the Marine Biological Association of the United Kingdom. 2010;**90**(3):569-573

Shimokomaki M, Visentainer JV, et al. Proximate composition and fatty acid profile of raw and roasted salt-dried sardines (*Sardinella Brasiliensis*). Food and Nutrition Sciences. 2011;**2**:440-443

[18] Valenzuela R, Videla LA. The importance of the long-chain

polyunsaturated fatty acid n-6/n-3 ratio in development of non-alcoholic fatty

[15] Mehta LR, Dworkin RH,

Nutritional Sciences from Fundamentals to Food. 3rd ed. Belmont: Wadsworth, Cengage Learning; 2013. pp. 217-268

Fundamentals of Organic Chemistry. 6th ed. Thomson Brooks/Cole: Belmont;

[3] Mader SS. Biology. 12th ed. New York: McGraw-Hill companies; 2013. pp. 42-46

[4] Fahy E, Subramaniam S, Brown HA, Glass CK, Merrill AH Jr, Murphy RC, et al. A comprehensive classification system for lipids. The Journal of Lipid

[5] Wakil SJ. Mechanism of fatty acid synthesis. Journal of Lipid Research.

[6] Leonard AE, Pereira SL, Sprecher H, Huang YS. Elongation of long-chain fatty acids. Progress in Lipid Research.

Membranes, and Cellular Transport In Concepts in Biochemistry. 1st ed. Pacific Grove: Brooks/Cole Publishing Company A division of International Thomson Publishing Inc.; 1999. pp. 238-275

Sons, Inc; 2008. pp. 47-49

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Research. 2005;**46**:839-862

1961;**2**(1):1-24

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pp. 208-240

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[9] Campbell MK, Farrell SO. Lipids and proteins are associated in biological membranes. In: White A, Olafsson J, Bowen L, Weber L, Van Camp S, editors. Biochemistry. 7th ed. China: Brooks/Cole, Cengage Learning; 2012. pp. 193-225

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*Fatty Acid Compositions in Fermented Fish Products DOI: http://dx.doi.org/10.5772/intechopen.90110*

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*Advances in Lipid Metabolism*

**82**

**Author details**

Kingdom of Bahrain

\* and Aysha Mohamed Alkaabi2

2 University of Bahrain, Sakhir, Kingdom of Bahrain

\*Address all correspondence to: afreije@uob.edu.bh

provided the original work is properly cited.

1 Department of Biology, College of Science, University of Bahrain,

© 2020 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

Afnan Freije1

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Function. 2011;**2**:644-648

2011;**124**:24-28

2003;**92**(4):308-316

1979;**2**(8140):433-435

2017;**124**:1-10

[19] Louly AWOA, Gaydou EM, ElKebir MVO. Muscle lipids and fatty acids profiles of three edible fish from the Mauritanian coast: *Epinephelus aeneus*, *Cephalopholis taeniops* and *Serranus scriba*. Food Chemistry.

[20] Njinkoué JM, Barnathan G, Miralles J, Gaydou EM, Samb A. Lipids and fatty acids in muscle, liver and skin of three edible fish from the Senegalese coast: *Sardinella maderensis*, *Sardinella aurita*, and *Cephalopholis taeniops*. Comparative Biochemisty and Physiliogy Part B. 2002;**131**:395-402

[21] Hirafuji M, Machida T, Hamaue N, Minami M. Cardiovascular protective effects of n-3 polyunsaturated fatty acids with special emphasis on docosahexaenoic acid. Journal of Pharmacological Sciences.

[22] Dyerberg J, Bang HO. Haemostatic function and platelet polyunsaturated

[23] Ward OP. Microbial production of long-chain polyunsaturated fatty acids. Inform. 1995;**6**:683-688

fatty acids in Eskimos. Lancet.

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Docosahexaenoic acid (DHA), a fundamental fatty acid for the brain: Newdietry sources. Prostaglandins, Leukotrienes, and Essential Fatty Acids.

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## *Edited by Rodrigo Valenzuela Baez*

This edited volume is a collection of reviewed and relevant research chapters concerning developments within the field of lipid metabolism. It includes scholarly contributions from experts in the field that cover such topics as roles of lipids in cancer, analytical tools for lipid assessment in biological assays, plant lipid metabolism, the effect of nanoparticles on lipid peroxidation in plants, and fatty acid compositions in fermented fish products.

This book provides a thorough overview of the latest research efforts by international authors on lipid metabolism, and opens new possible research paths for further novel developments.

Published in London, UK © 2020 IntechOpen © vjanez / iStock

Advances in Lipid Metabolism

Advances in

Lipid Metabolism

*Edited by Rodrigo Valenzuela Baez*