- Open Access
The effect of ultrasound pretreatment on some selected physicochemical properties of black cumin (Nigella Sativa)
© The Author(s) 2018
- Received: 28 March 2018
- Accepted: 12 July 2018
- Published: 10 August 2018
In the present study, the effects of ultrasound pretreatment parameters including irradiation time and power on the quantity of the extracted phenolic compounds quantity as well as on some selected physicochemical properties of the extracted oils including oil extraction efficiency, acidity and peroxide values, color, and refractive index of the extracted oil of black cumin seeds with the use of cold press have been studied.
For each parameter, three different levels (30, 60, and 90 W) for the ultrasound power and (30, 45, and 60 min) and for the ultrasound irradiation time were studied. Each experiment was performed in three replications.
The achieved results revealed that, with enhancements in the applied ultrasound power, the oil extraction efficiency, acidity value, total phenolic content, peroxide value, and color parameters increased significantly (P < 0.01). Enhancements in ultrasound irradiation time have not significantly increased the oil extraction efficiency, acidity value, total phenolic content, and peroxide value as well as the oil refractive index (P < 0.05). As the highest oil extraction efficiency (39.93%) was obtained from the seeds when the applied ultrasound power and time were 90 W and 60 min respectively, and the lowest acidity value of oil was achieved once the applied power and time of ultrasound were 30 W and 30 min respectively. The application of ultrasound as pretreatment has not shown any significant effects on the refractive index of the extracted oils (P > 0.05).
In summary, it could be mentioned that the application of ultrasound pretreatment in the oil extraction might improve the oil extraction efficiency, the extracted oil’s quality, and the extracted phenolic compounds content.
- Ultrasound pretreatment
- Black cumin
- Total phenols
- Physicochemical properties
The oils and fats not only from their potential health benefits point of view, but also from commercial point of views have attracted considerable attention of researches. These compounds might provide considerable quantities of required energy of body cells, required essential fatty acids, as well as fat-soluble vitamins .
Recently, with a growth and increase of general knowledge in this area, the demand of people for application of oils increased due to different reasons including their potential health benefits and providing desirable taste and high energy quantities. Regarding the fact that Iran needs to import oilseeds from other countries in producing vegetable oil or in other words rather than 90% of the consumed oil in Iran is provided from foreign countries, therefore the application of available oil sources in Iran for achieving self-healing and enhancement of the cultivated area under common oilseeds and exploration as well as cultivating novel oil sources are steps toward providing the required oil sources in countries suffering from lack of appropriate edible oil sources. Oils and fats prepared from various plant species and animal sources may have different metabolic, physical, and chemical properties regarding their species . Nigella sativa (black cumin) seeds, a dicotyledon of Ranunculaceae family, have been used since thousands of years ago as a spice and food preservative [3–5]. Thymoquinone, the main ingredient of the essential oil of N. sativa seeds, was also capable to reveal beneficial effects on acute gastric ulcer . In addition, thymoquinone and its reduced product named thymohydroquinone have been reported to have antibacterial activities and might be associated with some further antibiotic activities . The moisture, protein, and carbohydrate as well as oil contents of oilseeds are generally found in the range of 0.52–7.43%, 20–27%, 23.5–33.2%, and 25–32% respectively . Black cumin seeds contain tocopherol contents including α, β, and γ types. Among them, sterols and beta-cytosterols are the most important and predominant compounds in the extracted oil of black cumin seeds followed by Stigma sterols demonstrating 18.6 and 69% quantities respectively. Different approaches might be used to extract oil from oil seeds such as solvent extraction including Soxhlet and mechanical methods . Oil extraction with the use of mechanical approach is simple, safer, and less expensive compared to oil extraction with solvent and it has more advantages .
Recently, green extraction technologies including microwave, ultrasound, pulse electric field, instant controlled pressure drop, supercritical fluid processing in the frontiers of food processing, food chemistry, and food microbiology are not considered as novel approaches and have been used for rather than 30 years by academia and in different industries under extreme or non-classical conditions (currently a dynamically developing area in applied research and industry). Alternatives to conventional processing, preservation, and extraction procedures may increase the production efficiency and contribute to environmental preservation by reducing the use of water and solvents, elimination of wastewater, fossil energy, and generation of hazardous substances. Within those constraints, green food processing has to be introduced on the basis of green chemistry and green engineering: green food processing is based on the discovery and design of different technical processes reducing energy and water consumption, allows recycling of by-products through bio-refinery, and ensures a safe and high-quality product . It is almost a decade that novel extraction approaches have attracted considerable attention. In the abovementioned approaches, it has been tried to extract the oil in a most effective way to reduce the consumed solvent, time, and temperature and increase oil extraction efficiency and to reduce process expenses . For instance, with the use of a combined method in extraction of functional compounds of pomegranate fruit, the tough and wooden parts of the fruit must be digested and decomposed with the assistance of further methods.
Application of novel approaches including microwave and pulsed electric field in most of cases might increase the oil extraction rate and efficiency . Ultrasound is a novel technology used frequently in nutrition science [13, 14]. Ultrasound is described as waves irradiated in the waves in the range of 18–20 kHz. When ultrasound wave is applied in liquid or semi-liquid products, acoustic pressure as well as hydrostatic pressure are created.
In an unlimited level, acoustic pressure normally is considered as a sinusoidal wave and depends on the frequency, time, and maximum wave length . Oil extraction with ultrasound assistance is going to be considered as a replacement of traditional approaches in oil extraction industry due to having high extraction efficiency, less energy, and solvent consumption; moreover, it is considered as an improved approach in plant-based products particularly in extraction of compounds with lower molecular weight . The positive role of ultrasound waves in extraction might be relevant to its effects on destruction of cells’ membrane and as a result its impacts on extraction environment . Ultrasound application includes processes that could enhance the rate of extraction, quality of the extracted oil, safety of the process, and reduce time of extraction .
Ultrasound advantages are associated to cavitation process. Small produced bubbles in liquid phase might grow when the product is exposed to ultrasound waves and might fluctuate due to pressure alterations before the bubbles’ elimination .
The effective parameters on cavitations are frequency, liquid viscosity, solvent vapor pressure, external pressure, temperature, and available gas. Ultrasound frequency is one of the most effective factors in the size of the created bubbles. The effective parameters on cavitations are frequency, liquid viscosity, solvent vapor pressure, external pressure, temperature, and available gas. Ultrasound frequency is one of the most effective factors in the size of the created bubbles. The creation of cavitation needs high surface tension between gas and liquids; therefore, in liquids with low surface tension, less cavitation is created. Moreover, the trapped gas bubbles may facilitate the original cores of cavitation . The triggering of materials from substrates may reach the cavitation bubble near the surface and then through compression cycles, bubble collapse happens. Slow flow of liquid with high rate is sucked to the surface of the particles and therefore in high pressure and temperature destructs the plant cellular septum and triggers the products .
An increasing number of companies already used the different ultrasonic technology, either by adapting their conventional or innovative systems of extraction or by alterations in their installation. In industrial scale, the majority of extracted compounds are directly used, as in liquor/or wine as well as beer production or can be used as food and cosmetic additives, in the case of essential oil and bioactive molecules. Euphytos and Giotto companies are Italian companies specialized in natural extracts from herbs, fruits, and vegetables with the use of ultrasound technology to improve the flavor and quality of the obtained extracts . Zhang et al. , Lin et al. , and Goula  reported that the oil extraction efficiency with the use of ultrasound pretreatment might show enhancements. In the present study, the properties of the extracted oil of black cumin seeds with the use of ultrasound pretreatment have been studied and reported.
The black cumin seeds used in the current research were provided from the animal feed stores of Gonbad-e-Kavoos at spring 2017, then they were transferred to Azad University of Gonbad-e-Kavoos for oil extraction process. The chemical materials used in the present research including sodium hydroxide, phenolphthalein, sodium carbonate, glycolic acid, folic acid, Folin–Ciocalteu reagent (FCR), sodium thiosulfate, potassium iodide, and ethyl alcohol were provided from Merck Company (Germany) and all of the chemicals and reagents were provided from analytical grade.
Sample preparation and the pretreatments applied conditions before the oil extraction process
In the present research, the black cumin seeds (containing 41.8% oil content) were prepared; after moving to the lab, they were sieved and kept in plastic bags which were resistant to air and moisture penetration until the experimental day. Then, 500 g of sample for each treatment was shifted in a 1.5-l container that has been located inside the ultrasonic bath. Afterward, they were affected by the different conditions of ultrasound pretreatments (30, 60, and 90 W) with the constant frequency equal to 25 kHz, and ultrasound irradiation time (30, 45, and 60 min). After the pretreatments were completed, the oilseeds were extracted with the use of a screw press with 33 rpm, and the selected experiments were performed.
The oil extraction efficiency
Acidity value determination
where N indicates the normality of sodium hydroxide (NaOH), V presents the volume of the consumed NaOH, W represents the weight of the sample (g), and A expresses the fatty acids content based on oleic acid in 100 g−1 sample.
Determination of the refractive index
The refractive index value of the extracted oil was detected with the use of a refractometer device in the temperature equals to 25 °C according to the released approach by (AOCS Cc 7-25).
Color index measurement
Determination of the peroxide index
where S indicates the volume of the consumed sodium thiosulfate solution in milliliters, M expresses sodium thiosulfate molarity, W presents weight of the extracted oil in grams, and PO is the oil peroxide value in mEq of oxygen per kg−1 of the extracted oil.
Total phenolic compounds content
The total phenolic compounds have been determined through colorimetric method with the use of Folin-Ciocalteu’s phenol reagent. In this regard, 0.5 mL of sample was diluted up to 5 mL with distilled water. Then, 0.5 mL of Folin-Ciocalteu’s reagent was added to the diluted sample. After 3 min, 0.5 mL of 10% sodium carbonate was added to the abovementioned solution and was stand in room temperature at darkness for an hour. Finally, the samples’ absorption was recorded at room temperature with the use of a UV spectrophotometer at wavelength of 760 nm. Gallic acid was used to draw a standard curve (0 to 1000 μg in mL) and the total phenolic compounds content was calculated and the achieved data were reported as mg Gallic acid equivalent per each kg of sample.
The data analysis was performed using a completely random design in the form of 3 × 3 factorial experiments with three levels of irradiation power and three levels of irradiation time, each one in three different replications. The SAS software was used to analyze the information and Duncan’s multiple range test was used to compare the data mean.
The oil extraction efficiency
The influence of ultrasound power on some selected properties of samples
Ultrasound power (W)
Oil extraction efficiency (%)
Acidity (oleic acid%)
Peroxide (meq oxygen on kg oil)
Total phenolic content (ppm)
The influence of ultrasound irradiation time on some properties of samples
Ultrasound time (min)
Oil extraction efficiency (%)
Acidity (oleic acid%)
Peroxide (meq oxygen on kg oil)
Total phenolic content (ppm)
The ultrasound effect depends on the alterations in the environment, in other words, when cavitation bubbles implode in a solid surface, a distortion in the pressure zone occurs and increases up to 400 km h−1. This high rate implosion leads to the removal and rupture of cell membranes or increases their porosity, facilitating the mass transfer from the cells’ interior [26, 27]. Cavitation speed, sequential contraction, and expansions depend on the ultrasound frequency and cause deformations and create micro-channels ; as a result, it reduces the thickness of the boundary layer . The controlling mechanism in high-power ultrasound extraction is also attributed to the mechanical alterations, cavitations, and thermal effects.
The interaction of ultrasound power and time on some properties of the selected samples
Ultrasound power (W)
Ultrasound time (min)
Oil extraction efficiency (%)
Acidity (based on oleic acid%)
Peroxide (meq oxygen per kg of the extracted oil)
Total phenolic content (ppm)
27.29 ± 0.25i
9.68 ± 0.31i
109.8 ± 0.53i
8.00 ± 0.08i
71.2 ± 1.31i
34.15 ± 0.11e
10.26 ± 0.09g
110.3 ± 1.17h
8.50 ± 0.13h
79.4 ± 2.08h
36.78 ± 0.17d
11.57 ± 0.27c
117.2 ± 1.11f
9.50 ± 0.06g
89.0 ± 0.89g
29.04 ± 0.01h
10.12 ± 0.16h
112.4 ± 1.24g
10.50 ± 0.21f
92.4 ± 0.33f
33.49 ± 0.05f
10.70 ± 0. 17f
119.4 ± 1.15e
11.00 ± 0.04e
102.9 ± 1.46e
37.22 ± 0.10c
11.83 ± 0.01b
124.6 ± 1.01d
11.50 ± 0.11d
109.7 ± 1.71d
31.05 ± 0.13g
10.83 ± 0.01e
131.2 ± 0.13c
11.70 ± 0.09c
115.9 ± 0.19c
37.40 ± 0.51a
11.09 ± 0.09d
132.3 ± 0.09b
13.50 ± 0.15b
117.0 ± 0.31b
39.93 ± 0.19a
12.86 ± 0.17a
136.5 ± 0.47a
15.00 ± 0.33a
121.1 ± 0.42a
Different microscopic observations (SEM, environmental scanning electron microscopy (e-SEM), cyto-histochemistry) have been used for spacial and temporal localization of metabolites in rosemary leaves, that is one of the most studied and important plant species due to containing antioxidant metabolites used frequently in different food industries, during conventional and ultrasound extraction. The study permits to highlight that ultrasound-impacted rosemary leaves not by a single or different mechanisms in function of ultrasound power, as described by previous studies, but by a chain detexturation mechanism in a special order: local erosion, shear forces, sonoporation, fragmentation, capillary effect, and detexturation. These detexturation impacts followed a special order during ultrasound treatment leading at the end to the total detexturation of rosemary leaves .
The effect of operational parameters on acidity value of the extracted oil
The achieved results demonstrated that with enhancements in ultrasound power and irradiation time, the acidity value of the extracted oil may increase (Tables 1 and 2). Lipolytic enzymes under the thin grain shell are not able to hydrolyze fats in normal cells but in damaged cells these enzymes may start their activities and hydrolyze the oils and fats (substrate) and as a result may enhance the acidity of the extracted oils.
As shown in Fig. 1, application of ultrasound may lead to alterations in black cumin seed structure [10, 33]. The achieved data in Table 3 also presented as well that minimum acidity value of oil is obtained when the ultrasound power and time were in their minimum values. Yamaguchi et al.  found out that once the polymerization grade leads to enhancements in superoxide scavenging, the photonic capability may also increase. Obviously, with enhancements in process temperature and ultrasound pretreatment time, the condition gets appropriate to eliminate antioxidant compounds and generate free radicals . Acidity value might be used as a purity examination of the extracted oil and reveals that if oils had already started decomposition reactions.
The refractive index of the extracted oil
The refractive index (RI) is often used as a criterion for purity determination of the extracted oils. The RI of saturated fatty acids reveals a linear enhancement with escalating chain length. When RI is detected at a temperature above 40 °C, the RI of unsaturated FA increases with a degree of unsaturation. In both cases, the RI value was detected equal to 1.470, which is within the range reported in the literature for vegetable oils: olive oil (1.468–1.471), rapeseeds oil (1.465–1.469), sunflower seeds oil (1.467–1.469), and pumpkin seeds oil (1.466–1.474) as well as black cumin seeds oil (1.469–1.473) . Tables 1, 2, and 3 presented as well that the use of different pretreatments have not affected significantly on refractive index (P > 0.05). Bakhshabadi et al.  studied the effects of pulsed electric fields and microwave pretreatments on some selected physicochemical properties of black cumin seeds oil and reported that these treatments have not affected significantly on refractive index of this oil.
The effects of ultrasound power and time on the color parameters of the extracted oil
Color is one of most important parameter demonstrating food products quality; this parameter has an important role in attracting customers. Further properties including smell, taste, and texture are parameters that are judged by customers after shopping of the products . The achieved results of the current study demonstrated that with an enhancement in ultrasound power, color parameter of the extracted oil increased; the similar results were achieved when ultrasound irradiation time boosted (Tables 1 and 2). The cause of color enhancement of the extracted oils might be justified by the exhausting of pigments such as chlorophylls in the extracted oil and decomposition of phospholipids through ultrasound pretreatment processing. Jiménez et al.  reported that application of ultrasound might increase tocopherols, chlorophylls, and carotenoids contents in the extracted oils of olive oil.
The influence of operative parameters on peroxide value of the extracted oil
The achieved results presented that with enhancements in ultrasound power from 30 to 90 w, peroxide value of samples increased almost 12.14% and with enhancements in irradiation time from 30 to 60 min, peroxide value might increase almost 54.55% (Tables 1 and 2). The cause of enhancements in peroxide value might be attributed to oxidation reactions of fatty acids; this phenomenon may increase with boosting in the power and time of ultrasound pretreatment. Chemat et al.  found out that ultrasound waves might release free radicals as well as further volatile compounds and as a result increases peroxide value of the extracted oils.
Peroxides and hydroperoxides and the primary oxidation products decompose to form aldehydes, ketones, acids, alcohols, hydrocarbons, and other compounds. These substances are formed through thermal oxidation of oils and fats. The products of oil degradation as defined previously by International Union of Pure and Applied Chemistry  are polar materials. The polar fractions are composed from free fatty acids, monoglycerides, oxidized triglycerides, and oligomeric triglycerides.
Peroxide value is applied to detect the quantity of peroxide values in the extracted oils; these substances are important intermediate products of oxidative reactions since they decompose via transition metal irradiation and due to enhanced temperatures create free radicals . These results were in solid agreement of the achieved results by .
Table 1 indicates that with enhancements in ultrasound power from 30 to 90 W, the quantity of phenolic compounds increased from 93.21 to 106.6 ppm. With enhancements in ultrasound treatment time, the contact between solvent and solid materials increases leading to decomposition of cells’ membrane and as a result mass transfer increases; it means enhancements in diffusion of phenolic compounds to solvent. On the other hand, ultrasound may lead to initiation in polymerization reaction in short time and as a result might increase the extracted phenolic contents [43, 44]. Other studies [22, 45] achieved similar outcomes. Corrales et al. , in a study, extracted the available anthocyanins in grape waste with the use of ultrasounds, high hydrostatic pressure, and pulsed electric field and studied the extraction efficiency. The obtained outcomes demonstrated that 1 h after the extraction process, total phenolic compounds of the treated samples with the use of novel pretreatment approaches was 5% higher compared to control sample. Furthermore with the use of novel extraction approaches, antioxidant activity of the extracted infusions increased.
On the other hand,  demonstrated that the application of protracted ultrasound process in the extraction of plant-based phenolic compounds may lead to enhancements in extraction efficiency.
The degradation may result in a modification of macromolecular structures and a decrease of molecular weight leading to an improvement of the solubilization of polymers. However, the ultrasound effects have to be assessed on more complex structures since it is not obvious that these simplified models described for ultrasound impacts on single polymers would be valid for plant materials, which are composed of a large network of various polymers.
Table 3 demonstrates that the maximum amount of phenolic compounds is achieved once the power and time of ultrasound pretreatments were in the maximum degree.
The obtained results showed that with enhancements in ultrasound power, oil extraction efficiency, acidity and peroxide values, color parameter, and total phenolic contents increased. An increase in ultrasound irradiation time has shown the similar results. The application of ultrasound pretreatment has not presented significant effects on refractive index of the extracted oils. Regarding the achieved outcomes of the present research, it could be mentioned that the application of ultrasound pretreatments to treat the black cumin seeds before the extraction process with cold press approach may improve the quantitative and qualitative properties of the extracted oil. On the other hand, regarding the abovementioned advantages of ultrasound pretreatment and its applications and its impacts in the enhancements of extraction efficiency and phenolic compounds, it may be demonstrated that this approach may be used in medicinal and invaluable plant applications in industrial scales.
The authors would like to appreciate the Department of Chemistry, Gonbad Kavoos Branch, Islamic Azad University, Gonbad Kavoos, Iran, for the facilities provided during the present study.
Availability of data and materials
The data may be released for the reviewers and editor not for public due to the policy of the authors and University.
MM supervised the present research work and gave the required tips and scientific instructions. VF and HB did the experiments and obtained the data. All the achieved data were analyzed by all the authors. The article was prepared by VF and HB. All authors read and approved the final manuscript.
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