CHAPTER TWO

2.0 LITERATURE REVIEW

2.1 VEGETABLE OILS

Vegetable oils are obtained from plant seeds such as seeds such as sunflower, peanut, sesame, soy beans cotton seeds (Nzikou et al., 2010). They belong to a class of organic compounds called lipids. Vegetable oils contain a major component known as triglyceride which makes up 98% of its content and also the minor components (2%), comprising of free fatty acids, mono- and diglycerides, phosphatides, glycolipids, pigments, sterols, tocopherols, flavonoids, tannins, and trace metals (O’brien, 2018). Vegetable oils are widely used in food preparation, cosmetics and pharmaceutical industries, margarine, soap and detergent, paint and varnishes (Othman and Ngassapa, 2010).

Besides vegetable oils, other plant oils known as essential oils are obtained from plant materials such as flowers, leaves, bark, wood, roots or peels (Bansal, 2016).

2.2 FATTY ACIDS IN VEGETABLE OILS

Fatty acids are a class of aliphatic monocarboxylic acids composed of a carboxyl group and a hydrocarbon chain. Fatty acids are distinguished from one another by the nature of their hydrocarbon chain which can vary in length from 4 to 24 carbon atoms which can be saturated or unsaturated (one double bond, two or more double bonds). The most common fatty acids in vegetable oils are those containing 18 carbon atoms which includes stearic acid, oleic acid, linoleic, and linolenic acids (Ratnayake and Galli, 2009).

Figure 1: Structures of some C18 common fatty acids in vegetable oils (Ratnayake and Galli, 2009)

2.2.1 Nomenclature of Fatty Acids

The chemical nomenclature requires that carbon atoms be counted from the carboxyl end of the fatty acid (Davidson and Cantrill, 1985). However, for biological activity, carbon atoms are numbered from the terminal methyl group to the first carbon of the ethylenic bond. Such a classification is designated by the symbol ϖ-x, ϖx, or n-x, nx, where x denotes the position of the double bond closest to the terminal methyl group. Fatty acid abbreviations are made according to the number of carbon atoms in the molecule and the number of cis ethylenic double bonds. The general assumption is that all multiple double bonds are methylene interrupted (IUPAC-IUB commission on Biochemical nomenclature, 1978).

For example, linoleic acid with two double bonds, where one is located on the sixth carbon atom counted from the methyl group, will be abbreviated as C18:2n-6 (Mead and Fulco, 1976).

2.2.2 Formation of a Triglyceride in Vegetable Oils

A triglyceride (triacylglycerol) is an ester formed by combination of three fatty acid molecules and a glycerol alcohol. The reaction occurs when three hydroxyl (OH^–) groups of a single glycerol molecule react with the carboxyl group (COOH^–) of three fatty acids to form an ester bond. The fatty acids may be the same or different in structure.

Figure 2: Formation of triglyceride molecule (Boudreaux, 2020)

2.3 CLASSIFICATION OF FATTY ACIDS

Fatty acids may be classified according to the length of the hydrocarbon carbon chain as short, medium or long chained. Besides the hydrocarbon chain length, fatty acids may also be classified as unsaturated (Containing carbon-carbon double bond) and saturated (containing carbon-carbon single bond) (Turicka et al., 2011).

2.3.1 Saturated Fatty Acids

Saturated fatty acids contain only carbon-carbon single bonds in the hydrocarbon chain and naturally occurring saturated fatty acids usually have even number of carbon atoms (table 2.1) shows some major saturated fatty acids in vegetable oils.

Table 2.1:Major saturated fatty acids in vegetable oils (Gunstone, 1996)

(Source: Gunstone, 1996)

2.3.2 Unsaturated Fatty Acids

Unsaturated fatty acids contain one or more carbon-carbon double bond in the hydrocarbon chain and the double bonds may be located in the cis or trans configuration. Unsaturated fatty acids with only one double bonds are referred to as monounsaturated fatty acids (MUFA), for example, oleic acid (18:1n9). Whereas those with two or more double bonds are called polyunsaturated fatty acids (PUFA), for example, linoleic acid (18:2n6) and linolenic acid (18:3n3). Table 2.2 summarizes some major unsaturated fatty acids.

Table 2.2: Major unsaturated fatty acids in vegetable oils (Gunstone, 1996)

(Source: Gunstone, 1996)

2.3.3 Essential Fatty Acids (EFAs)

These are polyunsaturated fatty acids that are not biosynthesized by the human body, and therefore must be obtained from the diet. They are called essential because they are required for important biological processes like synthesis of prostaglandins and hormones (Glick and Fischer, 2013). Essential fatty acids have been reported to suppress production of pro-inflammatory compounds, and hence very important in control of diseases. Vegetable oils such as sunflower and sesame are some of the major and cheap sources of essential fatty acids, for example, alpha-linolenic acid (an omega-3 fatty acid) and linoleic (an omega-6 fatty acid).

2.3.3.1 Omega-3 Essential Fatty Acids

Vegetable oils are the main sources of omega-3 fatty acids especially alpha-linolenic (ALA). This fatty acid possesses health benefits such as anti-inflammatory, antiarrhythmic and antithrombotic properties (Weylandt et al., 2012). Fish and fish oil are rich sources of other omega-3 fatty acids specifically eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA).

2.3.3.2 Omega-6 Essential Fatty Acids

Vegetable oils, seeds, and nuts are dietary sources of omega-6 fatty acids, for example, linoleic acid. Al-Khudairy et al. (2015) reported that omega-6 fatty acids are important in the body for reducing risk of heart disease, lowering total cholesterol levels, lowering “bad” (LDL) cholesterol levels, raising “good” HDL cholesterol levels, and reducing cancer risks.

The essential fatty acids in vegetable oils are nutritionally important, but most susceptible to oxidative degradation during storage, which may cause deterioration of oils leading to loss of nutritional value, alteration of sensory properties such as aroma, flavour, and colour of vegetable oils.

2.3.3.3 Previous studies on fatty acid composition

Various studies have reported fatty acid composition of different vegetable oils.

Musalima et al., (2019) reported fatty acid composition of peanut oil from Uganda contained 39.71-55.89% oleic acid, 20.21-35.59% linoleic and 11.91-17.16% palmitic acids.

Achola et al., (2017) reported a range of 14.61 to 18.61% palmitic acid, 26.79 to 33.44% linoleic acid 21.9-34.46, 2.19 to 3.46% stearic acid in peanut oils from Uganda.

Flagella et al., 2002 reported that standard sunflower oil contains approximately 15% saturated, 85% unsaturated fatty acid consisting of (14-43%) oleic acid and 44-75% linoleic acid.

In recent years, high quality sunflower oils have been produced with a range of composition via development of mid-oleic type (43.1 to 71.8%) and high oleic type (75 to 90.7%) sunflower varieties that has high oleic acid content than standard sunflower type (flagella et al., 2002).

Hwang, (2005) reported that Fatty acid composition in Sesame seeds consist of abundant unsaturated fatty acid, like oleic acid (35.9-42.3%) and linoleic acid 41.5 – 47.9% from 80% of total fatty acids. Yoshida et al., (2000) reported fatty acid composition of seasame oil as 44% oleic, 34% linoleic and 10% palmitic and 7% stearic acids. High contents of linoleic acids and linolenic acids are another merit of sim sim oils as food source (Park, 2010)

Sesame seeds also contains less than 20% saturated fatty acids mainly palmitic (7.9 to 12% and stearic acid 4.8 to 6.1%. worldwide, fatty acid composition in sesame oil is variable among the different varieties of sesame seeds such as black brown.

Rehman et al., (2007) reported that Fatty acid composition of sesame seeds depends on different factors such as climatic situations soil conditions and ripeness of plants.

2.4 IMPACT OF FATTY ACID COMPOSITION ON HUMAN NUTRITION AND HEALTH

The ratio of unsaturated to saturated fatty acids in edible oils and fats is very important for human nutrition. While high levels of saturated fatty acids are desirable to increase oil stability, on the other hand, nutritionally they become undesirable, because high levels of saturated fatty acids are frequently considered to have an influence in increasing the concentration of low density lipoproteins (LDL), affecting the ratio of LDL to HDL (high density lipoproteins), a risk marker for Cardio Vascular Disease (CVD) (Barbour et al., 2015). Oleic acid, a monounsaturated fatty acid (MUFA) is however, thought to reverse the above effects. Oleic acid (C18:1n9) has also been associated with several human health benefits, including; decreased risk of cardio vascular disease (CVD) by reducing the levels of serum low-density lipoproteins (LDL) cholesterol; and maintaining the levels of high-density lipoproteins (HDL), without causing significant weight gain (Barbour et al., 2015).

Kris-Etherton (1999) reported that MUFAs decrease plasma triglyceride levels in comparison with carbohydrates. In addition, the MUFAs help in hindering the development of adrenoleukodystrophy (ALD) (Rizzo et al.,1986) and reversing inhibitory effects of insulin production (Vassiliou et al., 2009). MUFAs may also decrease platelet aggregation and increase ûbrinolysis, thereby protecting against thrombogenesis (Kris-Etherton, 1999). It also has anti-inflammatory properties that activate different pathways of immune competent cells (Carrillo et al., 2012).

Polyunsaturated fatty acids (PUFAs) such as linoleic (C18:2n6) are recognized for their susceptibility to oxidative rancidity because when heated at high temperatures makes it dangerous for human consumption (Isleib et al.,2006), and this instability leads to formation of trans-fatty acid, which has detrimental effect on human health as it causes cardiovascular disease (CVD) (Wang et al., 2015). In addition, linoleic acid is a metabolic precursor to arachidonic acid and eicosanoids, which has been associated with an increased risk of inflammation, cancers, CVD, and neurological disorders (Whelan, 2008).

2.4.1 Nutritional Index of Oils

The relationship between saturated and polyunsaturated FA content is expressed as nutritional index (P/S index). This value is an important parameter for determination of nutritional value of vegetable oils (Kostik et al, 2008). Vegetable oils with P/S index greater than 1 are considered to have a high nutritional value. Lawton et al. (2000) indicated that higher value of P/S index means a smaller deposition of lipids in the body. Other studies elsewhere have reported the P/S index value for some vegetable oils such as sunflower, peanut, soybean, and safflower as 6.76, 1.04, 4.26, and 10.55, respectively (Zambiazi et al., 2007; Daniewski, 2003).

2.5 BIOSYNTHESIS OF FATTY ACIDS

Fatty acid biosynthesis is a complex process due to compartmentalization of the biosynthesis pathway in different organelles in plant cells, and the extensive lipid movement from one organelle to another (Buchanan, 2000). An overview of various steps in the fatty acid biosynthesis pathway in plants is illustrated by Buchanan (2000) in (Figure3).

Figure 3: Fatty acid biosynthesis in plants (Buchanan, 2000)

Steps involved in fatty acid biosynthesis in plants; –

Step 1: Fatty acid biosynthesis begins with an ATP-dependent carboxylation process in which Acetyl-CoA carboxylase (also known as ACCase) catalyses the formation of malonyl-CoA (also known as activated Acetyl-CoA). Biotin carboxylase activates CO₂ by attaching it to nitrogen in the biotin ring of the biotin carboxyl carrier protein (BCCP). The flexible biotin arm of BCCP carries the activated CO₂ from the biotin carboxylase active site to the carboxyl transferase site and this enzyme transfers activated CO₂ from biotin to Acetyl-CoA, producing malonyl-CoA. ACCase is highly regulated enzyme which is active in light and inhibited in dark conditions. Also, being a part of the first step of lipid synthesis, this enzyme can be inhibited by a class of herbicides called ACCase inhibitors. The herbicide compounds affect meristem of the grasses and kill them by ceasing production of cell membrane.

Step 2: Malonyl-CoA transacylase (MT) exchanges the CoA for the ACP (Acyl Carrier Protein) which is an essential protein co-factor in fatty acid biosynthesis.

Step 3: The 3-ketoacyl-ACP synthase (KAS) enzyme has three isoforms – (KAS) I, II and III. KAS III is utilized during the first condensation reaction, which includes a carbon-carbon bond formation between C1 of an acetate primer and C2 of the malonyl-ACP. KAS I is active with C4-C14 acyl-ACPs and during production of C6:0 to C16:0. KAS II accepts only longer carbon chains and elongation of 16:0-ACP to C18:0-ACP requires KAS II.

Step 4: The 3-ketoacyl-ACP reductase (KR) enzyme reduces the keto group from 3-ketoacyl-ACP to form 3-hydroxyacyl-ACP.

Step 5: The 3-hydroxyacyl-ACP dehydratase catalyzes the removal of water from 3-hydroxyacyl-ACP to form the 2,3-trans-enoyl-ACP and in the next step, enoyl-ACP reductase converts 2,3-trans-enoyl-ACP to its corresponding saturated acyl-ACP.

Step 6: Enoyl-ACP reductase (ER) reduces the double bond and the repetitive condensation of 2 carbon units produces 16:0=ACP and 18:0-ACP. Each cycle of fatty acid synthesis adds two carbons to the acyl chain and the reaction stops at 16:0 or 18:0 when thioesterase terminates the biosynthesis cycle. 18:1-ACP is produced using 16.18:0-ACP using a stearoyl-ACP desaturase as a catalyst. 16:1-Δ9 is desaturated by using a soluble acyl-ACP Δ9 desaturase enzyme. Fatty acid elongation happens in the endoplasmic reticulum.

Several desaturase enzymes catalyze the formation of cis-double bonds which causes kinks in the fatty acid chain (Figure 4) and generate a diverse variety of unsaturated fatty acids in plant membranes and storage reserves.

Figure4: Introduction of double bonds in plant lipids (Buchanan, 2000)

2.6 FACTORS AFFECTING THE FATTY ACID COMPOSITION IN VEGETABLE OILS

Fatty acid composition determines the physical and chemical characteristics of vegetable oils, and the fatty acid composition is not always constant but influenced by factors such as genetic, environmental, and oil extraction methods.

2.6.1 Genetic Factors

Fatty acid biosynthesis in plant cells is controlled by a group of genes identified as sad1, sad2, fad2a, fad2b, fad3a, and fad3b, which are collectively known as desaturase (Thambugala et al., 2013). The genes act by encoding enzymes that perform fatty acid synthesis in the plant cells.  Variation in fatty acid composition in the different vegetable oils could be due to the differential expression of the genes during seed development and maturation (Baud and Graham, 2006).

In peanut, sesame and sunflower seeds, it has been observed that increase in the oleic acid content leads to the reduction in the levels of palmitic, stearic, and linoleic acids, which suggests that there is close linkage of genes controlling these fatty acids (Barkley et al., 2013). Variations in fatty acid compositions among different crop varieties of the same species is due to the cumulative effect of several minor genes that modify the expression of fad3a and fad3b (Vrintel et al., 2005). Baud and Lepiniec (2009) reported that the genes expression programme related to the fatty acid synthesis are activated during the maturation phase of the seed, and most genes encoding fatty acid synthesis display a bell shaped pattern of expression during seed development. The fad2 genes are thought to be the rate limiting genes of the fatty acid biosynthesis pathway, and are highly influenced by environment (Fofana et al., 2006), and (Esteban et al., 2004) also reported that there is a significant gene environment interaction which determines the fatty acid composition differences.

2.6.2 Environmental Factors

Environmental growth conditions of plants such as temperature, soil, moisture, altitude, and light affects the expression of the alleles and relative activity of the genes responsible for fatty acid biosynthesis in plant cells. Therefore, vegetable oils obtained from plants seeds of the same species but from different localities may have different fatty acid compositions (Alizadeh, 2010).

2.6.2.1 Temperature

Varying temperatures during growth and maturation seasons of plants affects the level of gene expression which cause alteration in quantities of fatty acids of vegetable oil (Fofana et al., 2008). Byfield and Upchurch (2007) also confirmed that at high temperature during growth and maturation seasons of plant seeds, linoleic acid content reduces while oleic acid content increases because fatty acid desaturase enzymes, encoded by three genes converts linoleic acid to oleic acid, and hence altering the fatty acid composition in some seeds such as sunflower, soybeans, peanut, and sesame.

2.6.2.2 Soil

Soil contains nutrients such as nitrogen, potassium, and phosphorous which are used in the biosynthesis of fatty acids, and the concentration of these nutrients in the soils may influence the quality and composition of fatty acids in plant seeds (Stepien et al., 2017).

Kaptan et al. (2017) reported that the application of NPK fertilizers to the soils significantly increased the concentration of lignoceric and arachidic acids, whereas myristic and palmitic acids were decreased. On the other hand, fertilization had no significant change on unsaturated fatty acid content observed, except eicosenoic acid.

2.6.2.3 Water

Igbadun et al. (2006) showed that the crop yield response is very much dependent on the amount of water applied at different crop development stages than the overall seasonal water applied. Drought stress in the flowering stage of a plant leads to reduced availability of carbohydrates (glucosinolate) for fatty acid biosynthesis, and may cause reduction in the concentration of saturated fatty acids in the plant seeds (Geogel et al., 2007). Drought stress also leads to oxidation of polyunsaturated fatty acids in the plant seeds thus altering the fatty acid composition in plant seeds (Singh and Sinha, 2005).

2.7 FACTORS AFFECTING NUTRITIONAL QUALITY OF VEGETABLE OILS

There is still no standard defining the quality characteristics of vegetable oils for use as food and industrial applications, but there are a number of factors such as fatty acid compositions, oil extraction and processing methods, seed quality and storage systems that are used to evaluate the quality of a good vegetable oil (Okparanta et al., 2018; Abulude et al., 2007).

2.7.1 Fatty Acid Composition and Oil Oxidation

The quality of vegetable oils largely depends on the quantity of oleic and linoleic acids, which constitutes more than 80% of total fatty acids. Good quality vegetable oil contains high proportion of fatty acids like oleic, palmitic, and stearic acids which are resistant to autoxidation. The linoleic and linoleic acids are nutritionally important, but are susceptible to oxidation because their acyl residues are chemically reactive which adversely impacts oil stability and quality. Oxidative degradation of oil leads to loss of nutritional value, alteration of sensory properties like flavour, aroma, and colour. The degree of autoxidation and the potential for deterioration are important parameters of the vegetable oils. It has been reported that factors such as temperature, light, processing procedures, and oxygen concentration are the main causes of oxidation in vegetable oils (Kamal, 2006; Merrill et al., 2008).

2.7.2 Extraction and Processing Methods of Vegetable Oils

The traditional methods are crude, largely unscientific, inefficient, and yielding poor quality oil that contain substantial amount of contaminants.

The solvent extraction method is commonly applied to oilseeds with low oil content (< 20%), like soybean. This method has some disadvantages such as poor product quality caused by high processing temperatures, and a relatively high number of processing steps (Dawidowicz et al., 2008; Takadas and Doker, 2017).

Mechanical press methods are often used to extract vegetable oil from oilseeds having oil content higher than 20% (Sinha et al., 2015). There are two types of mechanical press method namely, cold-press and hot-press methods.

Cold-press or scarification method is carried out at low temperature below 50⁰C. Cold-pressed method are safer than hot-pressed method because high temperature is avoided in the cold press method. In cold-pressed oils, the purity and natural properties of oils are preserved (Azadmard- Damirchi et al., 2011; Bhatol, 2013).

The hot-press method is carried out at elevated temperature and pressure. This method results in decreased oxidative stability, degradation of valuable oil components and reduced oil shelf-life of the vegetable oil.

2.7.3 Refining and Storage

Inadequate handling and storage facilities for oilseeds and vegetable oils during refining and transportation may lead to contamination of oil products by soil, insects, rodents or microorganisms, and may cause loss or gain of moisture, odor or flavour (Ngassapa and Othman, 2001). The longer the seeds are stored, the higher the contamination and deterioration, and the higher the free fatty acid contents of the oil.

Oil refining processes may lead to contamination of vegetable oils with heavy metals which accelerates oxidation reaction.  Oil refining removes antioxidants which leads to acceleration of oxidation reaction that reduces shelf-life. Addition of synthetic antioxidants such as BHT, BHA and BTHQ may also lower oil quality. To retain oil quality, care must be taken while storing vegetable oils for a period of time to prevent their deformation as they easily undergo oxidative deterioration, hence shortening their shelf life.

Bukola et al. (2015) reported that oil stored in a refrigerator has a greater nutritional quality than that of a cupboard and shelf upon long storage because the peroxide and acid values obtained from such oils are lower than the other means of storage.

2.7.4 Pro-oxidants

Pro-oxidants in oil have a detrimental effect on oil stability. Metals act as pro-oxidants by electron transfer whereby they liberate radicals from fatty acids or hydroperoxides as in the following reactions (Gordon, 1990):

M^(n+l)+ + RH              Mⁿ⁺ + H⁺ R·

ROOH + Mn+                      RO· + OH- + M^(n+l)+

ROOH + M^(n+l)+              ROO· + H+ + Mⁿ⁺

Two of the more active metals to induce oxidation are copper and iron of which copper is the most pro-oxidative (Garrido, Frias, Diaz and Hardisson, 1994).

2.8 VEGETABLE OIL EXTRACTION METHODS

Most vegetable oils are extracted from plant seeds, and the oil yield will depend on seed variety, soil, and environmental conditions around the oil-bearing plant, as well as on pretreatment procedure, and on the particular extraction method (s) used (Mariana et al., 2013). Selection of suitable extraction method(s) is a key factor, and may depend on whether small or large scale oil extraction is intended. Vegetable oil producers widely use methods like traditional, solvent extraction, and mechanical expression to extract vegetable oils from plant seeds. The percentage yield and percentage oil recovery are evaluated using the expressions;

% Oil yield (1),

% Oil recovery =  (2),

Where Moil = mass of oil (kg),

Mseed= mass of oilseed (kg),

X = oil content of oil seed.

2.8.1 Traditional Method

Oluwole et al. (2012) reported that the traditional method of vegetable oil extraction involves, collection of seed pods, shelling of the pods/winnowing, roasting/drying of seeds to reduce moisture, grinding the roasted seeds by mortar and pestle or in between two stones to form a paste. The paste is mixed with water, and boiled to obtain the oil by floating and skimming. The oil is then dried by further heating.

The traditional method is tedious, time consuming, energy sapping, largely unscientific, inefficient, and yielding poor quality oil (Olaniyan and Yusuf, 2012). This method is mainly practiced among the rural communities to obtain vegetable oils on a small scale from peanut and sesame seeds, and also sheanut.

2.8.2 Solvent Extraction Method

In soxhlet extraction, normally a solid material containing the desired compound is placed inside a thimble made from thick filter paper, which is loaded into the main chamber of the Soxhlet extractor. The soxhlet extractor is placed onto a flask containing the extraction solvent-hexane and is then equipped with a condenser, and the solvent is heated to reflux. The solvent vapour travels up a distillation arm and floods into the chamber housing the thimble of solid. The condenser ensures that any solvent vapour cools, and drips back down into the chamber housing the solid material.

The chamber containing the solid material slowly fills with warm solvent. Some of the desired compound will then dissolve in the warm solvent. When the Soxhlet chamber is almost full, the chamber is automatically emptied by a siphon side arm, with the solvent running back down to the distillation flask. This cycle may be allowed to repeat many times, over hours or days.

During each cycle, a portion of the non-volatile compound dissolves in the solvent. After many cycles, the desired compound is concentrated in the distillation flask. The advantage of this system is that instead of many portions of warm solvent being passed through the sample, just one batch of solvent is recycled.

After extraction, the solvent is removed, typically by means of a rotary evaporator, yielding the extracted compound. The non-soluble portion of the extracted solid remains in the thimble and is usually discarded.

The solvent extraction method is commonly applied to plant seeds with low oil content (<20%). This method is considered as one of the most efficient methods in vegetable oil extraction, with less residual oil left in the cake or meal (Tayde et al., 2011). The choice of solvent is based mainly on the maximum leaching characteristics of the desired solute substrate (Dutta et al., 2015). Solvents commonly used are hexane, diethyl ether, petroleum ether, and ethanol. Other considerations are high solvent-solute ratio, relative volatility of solvent to oil, oil viscosity and polarity, and cost (Muzenda et al., 2012; Takadas and Doker, 2017). Hexane a hydrocarbon, with chemical formula C₆H_14, boiling and melting points 68.7°C and -95.3°C respectively, has become the solvent of choice for solvent extraction (liquid-liquid extraction) because of its high stability, low evaporation loss, low corrosiveness, little greasy residue, and better odor and flavour of the extracted products (Muzenda et al., 2012). Solvent extraction using hexane has several drawbacks, including high capital equipment cost and operational expenditures, the perpetual hazard of fire and/or explosion, as well as the residual solvent associated with both oil and meal.

The primary prerequisite for solvent extraction of oils is the rupturing of the seed or feed material to render the cell wall more porous, and complete rupturing of the cell wall is necessary for rapid extraction. Soxhlet based solvent extraction process is the primary means of extracting vegetable oil from oleaginous materials. The Soxhlet process is also widely used in laboratory scale oil extraction (Abdelaziz et al., 2014), but large scale operation of this process would require a commercial solvent extractor (Ogunniyi, 2006). The major advantage of the Soxhlet process is solvent recycling (over and over) during extraction. However, disadvantages of the Soxhlet method include high solvent requirement, time, and energy consumption (Takadas and Doker, 2017), as well as sample being diluted in large volumes of solvent (Rassem et al., 2016).

2.8.3 Mechanical Expression Method

Mechanical expression involves the application of pressure using hydraulic or