Physicochemical and Functional Properties of Ethiopian (Roba Variety) Peanut (Arachishypogaea L.) for Industrial Use

Special Article - Antioxidants in Foods

Austin J Nutri Food Sci. 2018; 6(3): 1107.

Physicochemical and Functional Properties of Ethiopian (Roba Variety) Peanut (Arachishypogaea L.) for Industrial Use

Gebremedhin G1,2*, Shimelis A², Tarekegn B³ and Cernak M4

¹Department of Food Process Engineering and Postharvest Technology, Ambo University, Ethiopia

²School of Chemical and Bioengineering, Addis Ababa Institute of Technology, Ethiopia

³Addis Ababa Science and Technology University, Ethiopia

4Department of Physical Electronics, Masaryk University, Czech Republic

*Corresponding author: Gebremedhin Gebremariam, Department of Food Process Engineering and Postharvest Technology, Ambo University, Ethiopia

Received: May 28, 2018; Accepted: June 27, 2018; Published: July 04, 2018


Peanut (Arachishypogaea L.) is considered as a highly nutritious and as a functional food has been growing. Target tests and experiment were carried out to ascertain the physiochemical and functional property of peanut. Rova variety, Ethiopian peanut was analyzed for its oil content, fatty acid profiles, total phenolic content, along with antioxidant scavenging activity, peroxide value, unsaturated fatty acid, saturated fatty acid, acid value, refractive index, saponification and un saponification matter, microstructure, hardness, moisture, vitamin E (alpha), contact angle, iodine value, density, dynamic viscosity, flash, fire points, color (L*, a* and b*) and sensory evaluation. The mean value for the peanut analyzed were, oil: 53.73%, O/L ratio: 1.335, saturated fatty acid: 77.41%, unsaturated fatty acid: 22.05%, peroxide value: 1.56, iodine value: 89.23, Saponification value: 182.37, Refractive index: 1.45, density: 0.91, dynamic viscosity: 55.72mPa.s, flashpoint: 230.0, fire point: 245, total polyphenols: 200.23, and hardness: 109.90N. This indicates the peanut with high oil content, high antioxidant capacity, with the desired composition of fatty acids and vitamin E were identified which would be useful for the industrial purpose to develop nutritional superior peanut products.

Keywords: Peanut; Fatty acid profile; Functional properties; Microstructure; Physical properties


The peanut (Arachis Hypogea L.) is an annual legume grown in the tropics and temperate regions around the globe [1] and widely consumed throughout the world and is one among the five widely grown oil crops in Ethiopia [2]. It is widely used as an economic food enhancement to counter malnutrition owing to its high nutritional value [3]. It is a globally important oilseed valued as a source of high-quality cooking oil, crude protein, crude fat, crude fiber, water, ash, total sugar, amino acids, fatty acids, vitamins, minerals, phytosterol, resveratrol, squalene, and other anti-nutritional factors [4] and appreciated worldwide as an affordable, flavorful, serving as a primary ingredient for peanut butter, confections, and nutritional bars, among other finished products.

The most attractive and impressive oilseed crop contains so many bio-constituents for human betterment such as protein, oil (linoleic acid and oleic acid, also a good source of Omega-6 fatty acids and Omega-3 fatty acids) and vitamin E are the most important, and they act on reactive oxygen species, as an anti-oxidant as a curative for early ageing of human being [5]. To use for the different purpose the local variety of peanut (Roba), the physicochemical and functional characteristics of the oil and the peanut seed should be quantified. The behavior of the peanut seed and oil helps to know dietary use, quality and shelf life determination. The peanut seed (Rova variety), no compositional data in terms of being dietary, physical, and function properties have been reported, and no studies have been conducted on its potential as a new variety. Therefore, the present study analyzed the physicochemical characteristics and functional properties of the peanut seed based on recent techniques and methods available, finally, the generated information use for processors, the exporters, breeders, as well as by the researchers engaged in improving the quality of Ethiopia peanut and in other world.

Materials and Methods

Chemicals and samples

n-Hexane, methanol, 95% ethanol, potassium hydroxide, sodium thiosulphate, potassium iodide, chloroform, glacial acetic acid, 2,2-Diphenyl-1-picryhydrzyl radical (DPPH), Folin-Ciocalteu, sodium carbonate, starch, BHT, ascorbic acid, petroleum ether, ethanol, acetone, Gallic acid standard, and vitamin E standard (a). A domestic, commercial peanut (Roba variety) was obtained from the Were Research Center, Oromia, Ethiopia.

Extraction methods

The extraction was performed in duplicate, with solvent, n-hexane (99% purity). An automated Soxhlet set (The Soxhlet extractor SXT- 06, Shaanxi, China) was used to extract peanut oil. To achieve this, 5g of sample was packed in a cartridge placed inside a 250ml extractor device. The sample was extracted for 8h at the solvent’s boiling point temperature. Then extra solvent form sample oil was removed by rotary vacuum evaporator. The extracted was stored in brown bottle at refrigerator for further analysis.

The total oil was calculated using the following;

Analysis of peanut oil

The Acid value, peroxide value, iodine number, saponification value and un saponification matter oil were determined according to standard methods (AOAC, 2010). Density and dynamic viscosity of peanut oil were measured at 20°C based on method [6]. The refractive index of seed oil was measured at 20°C using an Abbemat 550 refractometer and droplets of oil were added to the measuring prism using a disposable pipette. The Flash and Fire was measured using in frary gun thermometer, open cup, torch nozzle and heater based on ASTM method.

Fatty acid determination

The lipid fraction of peanut seed oil samples was extracted and fatty acids methyl esters were prepared according to [7] and the fatty acid profile was determined by gas chromatography with mass spectrophotometer (GC-MS).

Fatty acid methylation

In a 50mL round bottom flask fitted with reflux condenser, the Soxhet extracted oil (1gm) was placed and dissolved in 2% methanolic potassium hydroxide (10mL) prepared by mixing KOH with methanol. The mixture was heated on a water bath at 50°C for 1h. The reaction mixture was allowed to cool down to room temperature and saturated NaCl (3mL) was added to the reaction mixture and the solution was swirled gently several times. N-Hexane (20mL) was added into the solution and then the mixture was transferred to a separator funnel. The organic layer (upper layer) was separated, dried over anhydrous sodium sulfate and filtered through a man No.1 filter paper. The solvent was removed by rotary evaporation. The transesterified sample was prepared at 10ppm concentration in triplicate for GC-MS analysis.

GC–MS instrument

Oils extracted from peanut was be analyzed by gas chromatography (Agilent technologies 7820A GC system coupled with Agilent technologies 5977E MSD, USA). The chromatographic capillary column (HP-5) 30m long and 0.25mm in internal diameter was used. Ultra-high purity (99.999%) helium gas, as the carrier gas, was used at constant flow mode. An Agilent 7820A auto sampler was used to inject 1μL of the sample with a split less injection mode into the inlet heated to 275°C. Oven temperature was programmed with the initial column temperature of 60°C held for 2 min, and then, the temperature was increased at a rate of 10°C/min until the column temperature was reached 200°C, and then heated at the rate of 3°C/min till the temperature reached 240°C. No mass spectra were collected during the first 4 min of the solvent delay. The transfer line and the ion source temperature will 280°C and 230°C, respectively. The detector voltage wills 1600 V, and the electron energy will 70 eV. Mass spectra were collected from 40–600 m/z. The parameters, such as the quality, and probability values of peaks identified was made through a library search using NIST 2014. 1μL of oils peanut were injected into gas chromatography coupled with mass spectrometry (GC-MS) and each component of essential oils were identified by comparing its mass spectrum with reference data from the equipment database (NIST 2014 Mass Spectral Library).

Measurements of physical properties of peanut seed

Moisture content of whole peanut was dried in a forced air oven at 130°C for 6 hours based on the method of Young, Whitaker [8]. The weight differences before and after oven drying was used to calculate moisture content (MC; % dry weight).

Color measurement of peanut seeds

Surface color of peanut was measured using colorimeter, model of CM-700d/600d (Konica Minolta, INC, Japan) and recorded in L* (lightness/darkness), a* (redness/greenness), and b* (yellowness/ blueness) color values. The polycarbonate measuring dish was filled peanut samples to determine the average value of 10 replications. The illuminant was D65 and the standard observer was 10°. The colorimeter was calibrated using black and white.

Hardness of peanut seeds

The hardness of peanut sample was analyzed using Texture Analyzer model (TA plus machine, AMENTEK Lloyd Instruments Ltd, Forum House, UK) installed with the Nexygen Plus software. The compression was applied on a peanut sample placed on the plate using a cylindrical probe. The mean value of the maximum peak of the first compression (N) in the force-time curves were considered to evaluate hardness of peanut seeds. Five measurements were performed at each plasma experiments.

Contact angle measurement

The contact angle measurement was performed according to the method described by [9]. The surface wet ability of the peanut samples was evaluated at room temperature using DSA 30 (Kruss, Germany) software controlled system for quick measuring of static and dynamic contact angles sessile drop technique. A drop of ultra- pure water with a volume of 0.5μl was placed on the flat horizontal sample surface with a micro syringe and was immediately (within 1 s) and automatically photographed with a CCD camera, 61 fps (780 × 580px) or 311fps (780 × 60px). The software controlled x, y, z-axes of the sample with a resolution of 0.1°and contact angle ranges from1 to 180°. The contact-angle was computationally determined from the captured images and reported the average of at least five measurements placed on different peanut samples and at different positions on the peanut.

Microstructure of peanut seeds

Acording the [9], the peanut samples were placed on SEM stubs and were coated with a thin layer of platinum (Pt) before sputter (Q150R). After sputtering, the surface morphology of peanuts was examined by Scanning electron microscope (MIRA3 made by Tuscan, Brno, and is fully PC controlled SEM equipped with a Schottky Field Emission electron).

Analysis of Functional Properties of Peanut Seeds

Total phenol and antioxidant activities

Sample preparation for extraction: Peanut seeds were milled (High Speed Universal Disintegrator (FW100) Grinder, China) with a speed of rotating knife (2400rpm) and passed through a mesh size 16 sieve to obtain identically sized particles then, was retained in a sealed bag in a refrigerator (1-2°C) until use. Milled peanut seed particle size is important to facilitate analyses of mass transfer during the extraction of oil and antioxidant. Peanut seeds were defatted first with n-hexane (10% w/v) using a soxhlet extraction unit for 8h. The defatted samples were then air-dried and extracted with methanol (100mL) using an incubator shaker (Thermo Shaker Incubator, Model, THZ-103B, China). All suspensions were then filtered through a Whatman No.1 filter paper and the residues re extracted twice, each time with additional 100ml of the same solvent. The filtrates were combined and the solvent evaporated under reduced pressure using a rotary evaporator (Eyela, Model N-1000) at 40°C. The methanolic extracts were used for the determination of total polyphenol and antioxidant activity.

Extraction of peanut for analysis of the antioxidant and polyphenols: Samples were extracted based on the procedures used by Bishi, Lokesh [10]. Briefly, five gram of dried groundnut powder was extracted by stirring with 50ml of methanol at 25°C at 150rmp for 24hrs using temperature shaker incubator (ZHWY-103B) and then filtered through what man No. 4 paper. The residue was then extracted two wise with addition of 50ml methanol as the above procedure. The combined methanol extracts were evaporated at 40°C to dryness using rotary evaporator (Stuart R3300). The crude extracts were weighed to calculate the yield and re-dissolved in methanol at the concentration of 30mg/ml and stored in a refrigerator (-4°C), until used for further work.

Measurement of antioxidant activities and total polyphenol

Total polyphones contents (TPC) determination: A modified Folin-Ciocalteu procedure as described by [11] was used for the determination of total polyphenol contents. Samples (0.1mL) were mixed with 1.0mL of the Folin- Ciocalteu reagent (previously diluted with distilled water 1:10 v/v), and the reaction was terminated using 1mL of 7.5% sodium carbonate. The mixture was vortexed for 15sec for color development. After 30 min incubation at room temperature (28±1°C), the absorbance was measured at 765nm using a UV-VIS spectrophotometer (Perkin Elmer Lamda 950 UV/Vis/NIR). The standard curve was prepared Figures 1 & 2 using gallic acid standard solutions of known concentrations, a linear calibration graph was constructed with gallic acid concentrations of 20, 50, 100, 150, 200, and 250μg/ml and the results were expressed as mg gallic acid equivalent/100g sample.

C= total polyphenol content(mg/gm); c- concentration of gallic acid (mg/ml); V- volume of extract in assay (ml) M- mass of pure plant methanolic extract(gm).

Free Radical Scavenging Assay (DPPH): The effect of methanol extracts on DPPH radical was estimated according to Win, Abdul- Hamid [12]. A 0.004% freshly prepared solution of DPPH radical solution in methanol was prepared and then 4ml of this solution was mixed with methanol extract of sample. Finally, the samples were incubated for 30 min in the dark at room temperature. Scavenging capacity was read by spectrophotometric (Perkin Elmer Lamda 950 UV/Vis/NIR) by monitoring the decrease in absorbance at 517nm. This absorption maximum was first verified by scanning freshly prepared DPPH from 200-800 nm using the scan mode of the spectrophotometer. Free Radical Scavenging activity DPPH in percent (%) was then calculated below and the scavenging activity as shown in Figure 3.

where Ao is the absorbance of the control and A1 is the absorbance of the sample.

Vitamin E analysis (a): Vitamin E(a) concentration of the sample was measured according to [13]. For the separation of alpha vitamin E, the sample was saponified in the following way; 5.0g of sample was mixed with 10ml of 50% KOH in 40ml of ethanol at 95°C for 45 min and was added 2 drops of pyrogallol crystal (to prevent oxidation and serves as and antioxidant). Mix and agitate with 10min intervals. This solution was subsequently transferred to a separating funnel along with 40ml distilled water and 20mL ethanol successively in order to get rid of the aqueous phase. 40ml deionized water was added to the sample and transferred to the digested sample into the separating funnel. Next 20ml ethanol and 70ml petroleum ether was added to each sample and shake the sample following by mechanical shaker for 5minutes. After shaking wait for few minutes and separate the pure solution and transferred into another separating funnel. Then, 70ml hexane was added to the remaining digested sample and shakes again using the mechanical shaker for 5 minute wait to allow separating the pure solution from the separating funnel and transferred to the other separating funnel. 30ml hexane and 30ml petroleum ether was added to the sample to the remaining digested and shake as the previous principle and separated as previously procedure. Next, 100ml distilled water was added to the pure collected sample on the separating funnel for washing and separate the bottom solution and repeat 3 times. Checked using phenolphthalein indicator until colorless was occurring. The sample was filtered using what man No.540, 125mm only the bottom portion to remove if water remains. The filtered sample was transferred and collected to the round flask and evaporated using rotary vacuum evaporator (773mbar and 40°C), dried the remaining sample using nitrogen, reconstituted using the addition of 10ml methanol, sonicated with ultrasonic for two minutes and finally, transferred to brown vial glass for further analysis.

Quantification of Vitamin E (alpha): Vitamin E content was measured by reversed phase HPLC and DAD -detection at 292nm wavelength. 10μL of sample was injected into a reversed-phase C18 column (SBC18, 250mm ×3, 5μm i.d.). 98% methanol and 2% distilled water was used as a mobile phase during analysis with a flow rate of 1ml/min. The column was purged with methanol for 10 min after each sample analysis to remove the traces of impurities and residues. The vitamin E was identified by comparison with the standard. Standards of a vitamin E were diluted with methanol. The stock solutions of vitamin E for the procedure were 10, 20, 30, 40, 60 and 80 PPm respectively. An equal volume of vitamin E was mixed, diluted with methanol, and placed in an auto sampler vial. The mixture was vortexes before use. Its concentration was determined by the absorbance maximums of the solutions using according to Beer’s Law. Calculations of the unknown were done by comparison of peak areas and the calculated concentrations of the standard solutions as indicated in Figure 1.