Optimizing Nigella Oil Extraction Temperature for Sustainable Production

Abstract

Understanding the extraction process of Nigella oil is crucial due to its significant impact on yield, quality, and therapeutic effectiveness. This study explored the effects of various screw press temperatures (40 °C, 50 °C, 60 °C, 70 °C, and 80 °C) on the quantity and quality of Nigella oil to optimize conditions that maximize yield while maintaining its nutritional and therapeutic attributes. Our findings indicate a linear increase in oil yield as screw press temperatures rose from 40 °C to 80 °C. There were no significant differences observed in total phenolic content (TPC), cupric reducing antioxidant capacity (CUPRAC), or the composition and ratio of fatty acids across oils extracted at different temperatures. However, the ferric-reducing antioxidant power (FRAP) was highest in oils extracted at 60 °C, while the thymoquinone (TQ) content peaked between 40 °C and 60 °C. These results underscore the importance of optimizing screw press temperatures to strike a balance between maximizing oil yield and preserving its valuable therapeutic and nutritional properties

1. Introduction

The oil from Nigella sativa seeds, which is also known as black cumin or Kalonji seed oil, plays a crucial role in nutrition by providing essential fatty acids, vitamins, and energy. Additionally, they hold significant value in the cosmetics and pharmaceutical industries due to the presence of phytoconstituents with medicinal properties, specifically thymoquinone (TQ) [1,2,3,4,5].

Several extraction systems are used to extract Nigella oil, including ultrasound-assisted extraction, microwave-assisted extraction, supercritical fluid extraction, mechanical extraction, Soxhlet extraction, and reflux extraction [3]. Over the past few decades, researchers have conducted comparative studies to understand the effect of these different extraction methods on Nigella oil yield and the concentration of its phytoconstituents. Their findings indicate that the oil yield and the concentration of phytoconstituents vary significantly depending on the extraction method used [6,7,8,9,10,11,12,13,14].

Among all the extraction methods, the screw press method, also known as mechanical pressing, has garnered significant attention due to its numerous advantages. These include high-quality product output, lower energy requirements, a more ecological and nontoxic approach, lower investment costs, and a comparatively simple extraction process [3,15,16,17,18]. Such benefits make the screw press method crucial for sustainable oil production by contributing to environmental preservation, resource efficiency, and economic viability.

Unlike solvent extraction, which uses chemicals like hexane, the screw press method does not involve any chemical solvents. This significantly reduces the risk of environmental contamination and ensures that byproducts are free of chemical residues, making them safer for use in animal feed or as fertilizers [19,20]. Utilizing these byproducts contributes to a circular economy, where waste is minimized and resources are continuously reused. Additionally, the low investment cost and easy handling of this method make it economically viable for small-scale producers, promoting sustainable local agriculture and production. Furthermore, the quality of oil is of prime importance and cannot be compromised. The mechanical nature of the screw press method has been reported to retain more natural nutrients and flavors in the oil, which are often lost in chemical extraction processes [20]. This results in higher quality, more nutritious oil. However, various parameters such as characteristics of the raw material (moisture content, oil content), feeding rate, extraction temperature, rotation speed of the screw, diameter of the restriction dye, and pre-treatment can affect the oil yield and the proportion of phytoconstituents in the screw press method. Therefore, it is important to identify the optimal conditions for these parameters [8,15,21,22].

Some researchers have investigated the optimal conditions for screw press extraction of Nigella oil using different parameters. For instance, Sakdasri et al. applied a combination of three different feed rates, temperatures, and moisture contents to determine the ideal extraction condition in terms of oil yield and TQ [18]. Deli et al. also utilized various physical parameters of the screw press oil expeller, such as nozzle size, extraction speed, diameter of the shaft screw, and extraction temperature, to identify the optimal extraction condition in terms of oil yield [8]. However, when optimizing the conditions for Nigella oil extraction, it is crucial to consider additional quality parameters, including TPC, antioxidant capacity, and fatty acid composition, in addition to oil yield and TQ content due to their significant influence in the nutritional, therapeutic, and commercial value of Nigella oil.

The aim of this research was to optimize the oil yield, bioactive compounds, and fatty acids composition of screw-pressed Nigella oil by varying the temperature while keeping other parameters constant. This comparative study of the screw press extraction method intends to support both industries and local farmers in obtaining the highest oil yield with minimal compromise on oil quality.

2. Materials and Methods

2.1. Materials

The seeds of N. sativa, procured from My Natural Beauty (https://www.mynaturalbeauty.com.au/) via an online shopping website (https://www.ebay.com.au/) on 3 June 2022, were used in this study.

Additionally, all reagents sourced from ChemSupply (Gillman, SA, Australia) or Sigma-Aldrich (Melbourne, VIC, Australia) were of analytical grade. Unless stated otherwise, Milli-Q^® water was utilized for dilutions and assays in chemical analysis. The seeds, reagents, and solutions were stored in a dark environment at 4 °C until further analysis.

2.2. Moisture Content

The moisture content of seed powder or oil was determined gravimetrically following the protocol applied in our previous report [23]. Briefly, 3 g of seed powder or oil were spread in an aluminum tray and heated to 103 °C in a Memmert drying oven (Memmert UM400, Schwabach, Germany) until a constant weight was achieved (90–120 min). Moisture content was then calculated using the equation below, and the results were expressed as a % of the total weight of sample:

$$MoistureContent\left(\%\right)={\displaystyle \frac{freshweightofsample\left(\mathrm{g}\right)-moisturefreeweightofsample\left(\mathrm{g}\right)}{freshweightofsample\left(\mathrm{g}\right)}}\times 100\%$$

(1)

2.3. Screw Press Extraction

Nigella seeds (20 g) were extracted using an oil press machine (an automatic oil extractor with pressure and temperature control voltage: 110 V/220 V, power: 600–1500 W, size: 42 × 16 × 32 cm³, Wgwioo brand). The extraction was performed at five different temperatures, 40 °C, 50 °C, 60 °C, 70 °C and 80 °C, while keeping the rotating speed, seed feeding rate, and moisture content constant at 58 rpm, 20 g/min, and 6.57% w/w, respectively.

The materials left in barrel and screw press during the extraction were considered waste. The weight of the waste material remaining around the screw and the barrel was determined by measuring the difference in weight before and after extraction. The accumulated cake in the vessel during extraction was collected and stored in plastic bags. The oil obtained from the pressing process was collected in individual 10 mL centrifuge tubes. These tubes contained a mixture of oil and sludge resulting from the pressing. Subsequently, the tubes were centrifuged at 4000 rpm for 20 min (Heraeus X1 Multifuge, Thermo Fisher Scientific, Melbourne, VIC, Australia). Following centrifugation, the sludge settled at the bottom of the tubes while the clean oil remained above it. The oil was then carefully decanted into fresh 10 mL centrifuge tubes. To ensure that only sludge or sediments remained, a washing step was performed using n-hexane, as recommended in previous literature, to remove any residual oil adhering to the tubes [24]. Two mL of n-hexane was added to each tube, vortexed, and then centrifuged again before the hexane layer was removed with a glass pasture pipette. The tubes containing solid residue were then dried to remove any remaining hexane. Some compounds, such as TQ present in Nigella oil, are highly sensitive to light and heat [25,26,27,28]. According to Ghosheh et al. [25], protecting the oil with aluminum foil and storing it under refrigeration can keep oil extract stable for up to two months. Therefore, immediately after screw press extraction, the oil samples were wrapped in aluminum foil and stored at 4 °C in a refrigerator until further chemical analysis.

2.4. Oil and Solid Components

The solid (sludge, cake, and waste remained in screw press) and oil yield of the extracted seeds were estimated using formulas from previous research [23,29,30,31]. The results were expressed as a % of the dry weight of the seeds.

$$Oilyield\left(\%\right)={\displaystyle \frac{weightofoil\left(\mathrm{g}\right)}{seedweightusedforscrewpressextraction\left(\mathrm{g}\right)}}\times \left(100-moisture\%\right)$$

(2)

$$Solidyield\left(\%\right)={\displaystyle \frac{weightofsolid\left(\mathrm{g}\right)}{seedweightusedforscrewpressextraction\left(\mathrm{g}\right)}}\times \left(100-moisture\%\right)$$

(3)

2.5. Test Sample Preparation

The test sample was prepared following the protocol applied by previous researchers [32,33,34]. Briefly, 1 g of screw press extracted Nigella seed oil was transferred into a 10 mL centrifuge tube. It was then mixed with 1 mL of hexane and 7 mL of extraction solvent (90% methanol/10% water) and vortexed for 10 s. The centrifuge tube containing the sample was mixed using an end-over-end shaker (Ratek RM4) for 60 min at 50 rpm. After this, the sample was centrifuged for 10 min at 3000 rpm. The methanolic extract supernatant was collected with a Pasteur pipette. The extraction was repeated with 7 mL of 90% methanol, but the end-over-end shaker time was reduced to 20 min. The methanolic extract from the second extraction was removed after centrifugation and combined with the initial supernatant and made up to a constant volume of 14 mL and stored at 4 °C in a refrigerator until further analysis.

2.6. Total Phenolic Content (TPC) of Oil

The Folin–Ciocalteu method was used to measure the total phenolic content (TPC) of the samples [35]. For the assay, 400 µL of diluted sample extract was added to a centrifuge tube with 2 mL of 1:10 diluted Folin–Ciocalteu reagent, vortexed, and incubated in darkness at room temperature for 10 min. Then, 2 mL of 7.5% sodium carbonate solution was added, vortexed, and incubated at 40 °C for 30 min. Absorbance was recorded at 760 nm using an Ultraviolet-visible (UV-Vis) spectrophotometer after measuring the blank with Milli-Q^® water. TPC was determined based on a standard solution of gallic acid (20 to 100 mg/L) with excellent linearity (R² = 0.9967), using the equation y = 0.0095x + 0.0075. Results were expressed in mg gallic acid equivalents (GAE) per 100 g of moisture-free sample weight (mg GAE/100 g DW).

2.7. Ferric-Reducing Antioxidant Power (FRAP) Analysis

The ferric-reducing antioxidant power (FRAP) assay method was used in this study following the protocol developed by Benzie and Strain [36]. Briefly, a 300 mM acetate buffer (pH 3.56) was prepared using sodium acetate and glacial acetic acid. Solutions of 10 mM TPTZ (2,4,6-Tri(2-pyridyl)-s-triazine) in 40 mM hydrochloric acid (HCl) and 20 mM ferric chloride were also prepared. The FRAP reagent was made by mixing these solutions in a 10:1:1 ratio and protected with aluminum foil. A 100 µL of diluted test sample and 3 mL of FRAP reagent were mixed properly and equilibrated at 37 °C for 4 min. After incubation, absorbance reading of sample at 593 nm was taken using a spectrophotometer. Trolox, a water-soluble vitamin E analog, was used as the standard. A 1000 mg/L Trolox stock solution in ethanol was diluted to 10–150 mg/L for the standard curve, which had a linearity of R² = 0.9992 and the equation y = 0.0056x + 0.0712. Total antioxidant capacity was expressed as milligrams of Trolox equivalents per 100 g of moisture-free sample weight (mg TE/100 g DW).

2.8. Cupric-Reducing Antioxidant Capacity (CUPRAC) Analysis

The cupric-reducing antioxidant capacity (CUPRAC) method, described by Apak et al., was used to determine the total antioxidant capacity (TAC) of the samples [37]. Briefly, solutions of 10 mM aqueous copper (II) chloride, 1 M aqueous ammonium acetate, and 7.5 mM neocuproine in ethanol were prepared. Thereafter, 1 mL of each reagent and 1 mL of Milli-Q^® water were mixed together in centrifuge tube before the addition of 100 µL of test sample. The tube was then vortexed and incubated at 50 °C for 30 min. After incubation, absorbance readings of sample at 450 nm were recorded using a spectrophotometer. Trolox served as the spectroscopy standard, prepared as a 1000 mg/L stock solution in ethanol and diluted to create a standard curve ranging from 50 to 600 mg/L. The CUPRAC value of the samples was quantified based on the equivalent absorbance of Trolox within this range, with a calibration curve showing high linearity (R² = 0.9988; equation: y = 0.0014x + 0.1686). Results were expressed as milligrams of Trolox equivalents per 100 g of moisture-free sample weight (mg TE/100 g DW).

2.9. Quantification of Thymoquinone (TQ)

TQ in Nigella oil was quantified using high-performance liquid chromatography (HPLC) with an Agilent 1100 system, comprising an autosampler, vacuum degasser, quaternary pump, and multi-wavelength detector module. Conditions followed Mani et al. [38], employing an Agilent Eclipse XDB-C18 column (150 × 4.6 mm², 5 µm particle size) with an isocratic mobile phase of water and methanol (40:60, v/v) at 1 mL/min flow rate. Sample injection volume was 5 µL, analysis was performed at room temperature, and UV detection at 254 nm for TQ. Identification relied on retention times and UV spectra compared to pure standards. Quantitative analysis used external standardization with calibration curves showing high linearity (R² = 0.9997). Results were expressed using the equation y = 17.21x + 12.584, based on peak area measurements of standards prepared in the 10–200 ppm range. The stock and standard solutions of TQ were prepared with methanol [39].

2.10. Fatty Acid Analysis

2.10.1. Deriv Atization (Methyl Ester Preparation)

Methyl ester preparation followed the modified sodium methoxide protocol described by O ‘Fallon et al. [40]. Briefly, 0.25 g of oil was mixed with 2 mL of 0.4 M sodium hydroxide (NaOH) reagent solution (prepared with methanol) and incubated at 55 °C for 1.5 h. After vertexing, 2 mL of saturated sodium bicarbonate (NaHCO₃) reagent solution (prepared with Milli-Q^® water) and 3 mL of hexane were added, vortexed, and centrifuged at 3000 rpm for 10 min. Thereafter, the hexane layer, containing fatty acid methyl esters (FAMEs), was collected, washed with Milli-Q^® water twice, and stored at 4 °C in a GC vial. The prepared FAME was then diluted with hexane (1:9) before further analysis.

2.10.2. Gas Chromatography-Mass Spectrometry (GC-MS) Analysis

Fatty acid methyl esters were analyzed using gas chromatography–mass spectrometry (GC-MS) following a protocol applied in the previous studies [41,42]. The Shimadzu QP2010 Plus system featured a Restek FAMEWAX column (30 m × 0.32 mm ID × 0.25 µm) with an injection volume of 0.5 μL in split mode (split ratio 10) at 250 °C. Helium flowed at 2 mL/min. The oven temperature ramped from 195 °C to 240 °C at 5 °C/min, holding for 1 min. Total runtime was 35 min. Ion source and MS interface were held at 230 °C. FAMEs were identified by scanning from 50 to 500 m/z and confirmed against Restek Food Industry FAME Mix (REST-35077) standards. Quantification of each fatty acid was completed based on the calibration curve equation, and the value was expressed in mg/g of oil.

2.11. Iodine Value

Fatty acids data obtained from GC-MS was used to estimate the iodine value (IV), as suggested in the literature [43,44]. The percentage composition of each unsaturated fatty acid in Nigella oil and its corresponding triglyceride factor were used for the IV calculation, as shown in the equation below:

$$IV=\sum \left(CiFi\right)$$

(4)

where IV is the iodine value, Ci is the percentage of individual fatty acid, and Fi is the corresponding triglyceride factor.

2.12. Statistical Analysis

The experiment was performed with 5 technical replications for each treatment. Values were expressed as mean ± standard deviation (SD). Data were analyzed by one-way ANOVA using IBM SPSS software version 28.000 (190). Wherever the p values were found to be less than 0.05, these were considered statistically significant. Furthermore, Pearson’s linear correlation test was employed to elucidate the associations between variables.

3. Results and Discussion

3.1. Oil and Solid Components

The results of Nigella oil samples obtained using the screw press method under five different temperatures (40, 50, 60, 70, and 80 °C) are presented in

Table 1. Oil and solid composition in screw-pressed Nigella seeds under different levels of temperatures.

Screw Press Temperature	Oil %	Sludge %	Cake %	Waste in Screw and Barrel %	Solid (Sludge, Cake and Waste) %
Extraction at 40 °C	10.9 ± 0.8 a	3.5 ± 0.1 a	55.5 ± 0.6 c	23.5 ± 1.2 d	82.6 ± 0.8 a
Extraction at 50 °C	12.9 ± 1.1 a	3.7 ± 0.5 a	54.3 ± 0.6 bc	22.6 ± 1.2 cd	80.6 ± 1.1 b
Extraction at 60 °C	15.6 ± 1.3 b	3.5 ± 0.3 a	54.0 ± 1.5 abc	20.4 ± 1.6 bc	77.8 ± 1.3 c
Extraction at 70 °C	19.0 ± 1.7 c	3.6 ± 0.3 a	52.5 ± 1.3 ab	18.2 ± 1.0 ab	74.4 ± 1.7 d
Extraction at 80 °C	21.4 ± 0.8 d	3.4 ± 0.2 a	51.8 ± 1.7 a	16.9 ± 1.4 a	72.1 ± 0.8 d

Values ± SD (n = 5) with different letters along the column are statistically different (p < 0.05).

. The results clearly demonstrate a significant effect of temperature on the oil yield. As shown in the table, the oil yield ranged between 10.9 and 21.4%, with the lowest yield obtained at 40 °C and the highest at 80 °C. The oil yield at 40 °C (10.9%) and 50 °C (12.9%) showed no significant difference, after which it gradually increased with higher extraction temperatures up to 80 °C.

Previous studies have also explored the impact of different temperatures in screw press extraction methods on oil yield, confirming significant variability [8,18]. For example, Sakdasri et al. [18] investigated different parameters, including screw press temperatures, and observed Nigella oil yields ranging from 8.8 to 31.6%, while Deli et al. [8] also investigated the effect of screw press temperatures on Nigella oil yield and reported its range between 15.2% to 22.7%.

In the present study, an increase in yield with temperature was observed, corroborated by other studies [45,46]. For example, Rotimi [46] reported an increase in soybean oil yield while extracting oil in screw pressed machine under a temperature between 50 °C to 90 °C; however, it decreased after then.

However, the present results contrast with some other earlier reports. For instance, Deli et al. [8] applied five different temperatures (50, 60, 70, 80, 90, and 100 °C) to extract Nigella oil from a screw press machine (nozzle size of 6 mm, shaft screw diameter of 8 mm, rotational speed at 21 rpm) and found higher oil yields at 50 °C (22.7%) compared to lower yields at 100 °C (15.2%). They observed a decrease in oil yield with increasing the extraction temperature [8]. This irregular relationship between screw press temperature and oil yield may be attributed to various factors such as seed moisture content, feeding rates, applied pressure, screw press speed, restriction size, nozzle size, and shaft screw diameter [18,21,45,47]. To support this, Sakdasri et al. [18] investigated different combinations of feeding rates (36, 45, and 54 g/min), screw press temperatures (40, 55, and 70 °C), and seed moisture contents (6%, 12%, and 18%) to study their effect on Nigella oil yield. They observed both increasing and decreasing patterns of oil yield with increasing temperature in their study. For example, when expelling seed samples with 6% moisture content at different feeding rates and temperatures, they noted an increase in oil yield with temperature, whereas they observed a decrease in oil yield when expelling samples containing 12% moisture [18]. This variability underscores the complexity of optimizing screw press extraction conditions to achieve maximum oil yield from Nigella seeds, highlighting the need for careful consideration of multiple factors influencing the extraction process.

The solid components (sludge, cake, and waste left inside the screw press machine during extraction) were also quantified (Table 1). It is noteworthy from the table that the sludge content in the oil obtained from different extraction temperatures ranged from 3.4 to 3.7%, with no significant differences. The factors such as moisture content and pretreatment methods, pressure, extraction duration, and screw speed might exert a more pronounced effect on sludge content compared to temperature [48].

Furthermore, the % of waste and cake in the present study ranged between 16.9 and 23.5% and 51.8 and 55.5%, respectively. The overall solid component (sludge, cake, and waste left in the screw and barrel) obtained from the extraction ranged between 72.1 and 82.6%. The highest value of solid content was observed in the 40 °C extraction, while the lowest was observed in the 80 °C extraction.

Overall, the present study reveals a direct relationship between screw press extraction temperature and oil yield but an inverse relationship with byproducts (solid components). Therefore, optimizing the temperature for extracting Nigella oil can have significant environmental and economic benefits from a sustainability perspective. For example, many global manufacturing companies produce large volumes of cold-pressed Nigella oil every year, which necessitates a substantial amount of raw materials, resulting in lower oil yields and higher amounts of byproducts. Optimizing the extraction temperature with sustainability in mind could lead to more efficient use of raw materials and a reduction in waste byproducts. This would not only increase the economic efficiency of oil production but also reduce the environmental impact. By minimizing waste and maximizing yield, the overall footprint of Nigella oil production could be significantly lowered. This approach aligns with sustainable development goals, emphasizing resource efficiency, waste reduction, and the use of environmentally friendly production methods.

3.2. TPC and Antioxidant Capacity

The results pertaining to the effect of screw press extraction temperatures in TPC and the antioxidant capacity of Nigella oil are summarized in

Table 2. Chemical characteristics in Nigella oil extracted with screw press machine at different temperatures.

Oil Extraction Temperature	TPC (mg GAE/100 g)	FRAP (mg TE/100 g)	CUPRAC (mg TE/100 g)
Extraction at 40 °C	90.4 ± 7.7 a	414.9 ± 27.2 a	411.4 ± 19.5 a
Extraction at 50 °C	89.4 ± 5.0 a	409.1 ± 17.3 a	380.4 ± 28.4 a
Extraction at 60 °C	89.9 ± 8.9 a	493.9 ± 46.6 b	410.3 ± 19.4 a
Extraction at 70 °C	85.8 ± 8.5 a	422.1 ± 31.2 a	391.8 ± 38.7 a
Extraction at 80 °C	81.1 ± 2.0 a	389.0 ± 6.7 a	417.0 ± 39.6 a

Values ± SD (n = 5) with different letters along the column are statistically different (p < 0.05).

. As shown in the table, TPC in the oils obtained varied from 81.1 to 90.4 mg GAE/100 g. The lowest TPC was observed at 80 °C extraction temperature, while the highest was at 40 °C. Similarly, the CUPRAC values ranged between 380.4 and 417 mg TE/100 g, with the lowest value obtained at 50 °C and the highest at 80 °C. However, TPC and CUPRAC values across all five different levels did not show significant differences. Regarding FRAP values, they ranged between 389 and 493.9 mg TE/100 g. The lowest FRAP value was recorded at 80 °C extraction, whereas the highest was at 60 °C. The FRAP values of oils extracted at 40 °C (414.9 mg TE/100 g), 50 °C (409.1 mg TE/100 g), and 70 °C (422.1 mg TE/100 g) exhibited no significant difference to those obtained at 80 °C.

While previous studies have not specifically explored the effect of screw press temperatures on TPC and antioxidant capacity, numerous investigations worldwide have evaluated these parameters in cold-pressed Nigella oils. The values reported in most of these studies align closely with the findings of the present study [14,49,50,51,52]. However, variability in TPC values has been noted in the literature. For example, Kiralan et al. [11] studied cold-pressed oil in Turkey and reported TPC values as low as 3.6 mg GAE/100 g, whereas Haron et al. [53] reported TPC ranging between 96 and 760 mg GAE/100 g for Yemeni and Malaysian Nigella seed oils. Furthermore, the present study observed that the TPC and antioxidant capacity of Nigella oil are comparatively higher compared to the reported values of many other edible oils, including linseed oil, pumpkin oil, milk thistle oil, rapeseed oil, camelina oil, and sunflower oil [54,55]. These findings highlight the potential of Nigella oil as a rich source of health-promoting compounds. While the present study found no significant difference in the TPC of Nigella oil extracted with the screw press method across five different temperatures, past research on various vegetable oils has reported diverse findings reading the effect of temperature on TPC. For example, Rababah et al. [56] investigated almonds, walnuts, and pistachios in Spain, extracting oils from these nuts using the screw press method at temperatures of 50 °C, 100 °C, 150 °C, and 200 °C. They noted no significant difference in TPC between almond and pistachio oils across the different extraction temperatures but observed a significant increase in TPC in walnut oil as the extraction temperature rose [56]. Similarly, He et al. [57] examined TPC in cold-pressed rapeseed oil (pressed at room temperature) versus hot-pressed (pressed at 120 °C), finding significantly higher TPC in the hot-pressed oil compared to the cold-pressed oil. This irregular trend in TPC across different extraction temperatures may be influenced by factors such as seed moisture content [58], the release of phenolic compounds from bound structures, or chemical alterations of phenolics [59].

3.3. Thymoquinone

The superimposed HPLC chromatograms of TQ obtained from one replicate of an oil sample screw pressed at different temperatures (40–80 °C) have been shown in Figure 1 as an example. Variations in TQ content in response to extraction temperature were observed in the present study, with concentrations ranging between 1027.4 and 1244.7 mg/100 g (

Table 3. TQ content in Nigella oil extracted with screw press machine at different temperatures.

Parameter	Oil Extraction Temperatures
	40 °C	50 °C	60 °C	70 °C	80 °C
TQ (mg/100 g)	1228.2 ± 57.3 c	1244.7 ± 51.6 c	1171.7 ± 64.3 bc	1102.7 ± 59.3 ab	1027.4 ± 86.5 a

Values ± SD (n = 5) with different letters along the row are statistically different (p < 0.05).

). The lowest value of TQ was observed when extraction was conducted at 80 °C, while the higher value was found at 50 °C.

Notably, the total TQ content in oil extracted at 50 °C showed no significant difference to that at 40 °C (1228.2 mg/100 g) and 60 °C (1171.7 mg/100 g). This sensitivity of TQ to temperature variations has been well-reported in the literature [18,60,61]. Ahmad et al. [60] and Pagola et al. [62] reported that TQ has a melting point between 49 and 50 °C and starts decomposing at 65 °C, with complete degradation occurring around 213 °C. Given its medicinal importance, the stability of TQ has been extensively investigated by many researchers [27,28,63]. Pathan et al. and Salmani et al. [28] observed significant degradation of TQ under acidic, basic, oxidation, and UV light conditions. For example, Salmani et al. [28] noted rapid degradation under light exposure, regardless of pH and solvent type. Additionally, Smith and Tess [63] reported that prolonged light exposure leads to the gradual conversion of TQ to dithymoquinone (70–80%), which further undergoes redox reactions to give other degradation products.

Optimizing extraction temperature to achieve maximum concentration of important phytoconstituents, such as TQ, without compromising the oil yield is crucial from a sustainability perspective. Since the present study observed the direct proportion and inverse proportion of oil yield and TQ, respectively, with the extraction temperature, balancing these factors is essential for sustainable production. By optimizing extraction processes to maintain high TQ content while maximizing oil yield, we can ensure efficient resource utilization and minimize waste.

Furthermore, TQ has been widely recognized for its potent antioxidant properties by many researchers [64,65,66]. However, the present findings contrast with these reports. Despite the highest and the lowest values of TQ being observed in oils extracted at 40 °C and 80 °C, respectively, the present study found that the TPC and antioxidant capacity were similar across these oil samples. Notably, the increase in TQ content did not correspond to a significant increase in the TPC or antioxidant capacity.

3.4. Fatty Acid Composition

The fatty acids composition of Nigella oil extracted via screw press at five different temperatures is illustrated in

Table 4. Composition of fatty acids in the Nigella screw-pressed oil at different temperatures.

Fatty Acids	Fatty Acid (mg/g of Oil) in Nigella Oil Extracted at Different Temperatures
Saturated Fatty Acids (SFAs)	40 °C	50 °C	60 °C	70 °C	80 °C
Myristic acid (C14:0)	2.1 ± 0.0 a	2.1 ± 0.1 a	2.0 ± 0.1 a	2.0 ± 0.1 a	2.0 ± 0.1 a
Palmitic acid (C16:0)	26.0 ± 0.8 ab	25.8 ± 0.5 ab	25.3 ± 1.0 a	27.7 ± 1.9 b	25.8 ± 0.7 ab
Margaric acid (C17:0)	0.9 ± 0.1 a	0.9 ± 0.0 a	0.9 ± 0.0 a	0.9 ± 0.1 a	0.9 ± 0.1 a
Stearic acid (C18:0)	7.6 ± 0.2 a	7.5 ± 0.1 a	7.4 ± 0.2 a	7.6 ± 0.3 a	7.5 ± 0.2 a
Arachidic acid (C20:0)	2.7 ± 0.0 a	2.7 ± 0.1 a	2.5 ± 0.1 a	2.6 ± 0.2 a	2.6 ± 0.1 a
Total SFAs	39.3 ± 1.1 ab	39.0 ± 0.4 ab	38.1 ± 1.2 a	40.7 ± 2.1 b	38.8 ± 0.8 ab
Monounsaturated Fatty Acids (MUFAs)
Palmitoleic acid (C16:1(cis-9))	1.5 ± 0.0 a	1.5 ± 0.1 a	1.4 ± 0.1 a	1.4 ± 0.1 a	1.5 ± 0.1 a
Oleic acid (C18:1)	26.2 ± 0.8 a	26.0 ± 0.5 a	25.5 ± 0.9 a	26.6 ± 1.7 a	26.1 ± 0.8 a
11-Eicosenoic acid (C20:1 (cis-11))	1.3 ± 0.1 a	1.3 ± 0.1 a	1.3 ± 0.1 a	1.2 ± 0.1 a	1.2 ± 0.1 a
Total MUFAs	29.0 ± 0.9 a	28.8 ± 0.4 a	28.2 ± 0.9 a	29.2 ± 1.7 a	28.8 ± 0.9 a
Polyunsaturated Fatty Acids (PUFAs)
Linoleic acid (C18:2)	107.7 ± 4.1 a	106.6 ± 2.0 a	107.3 ± 5.1 a	108.5 ± 8.9 a	106.9 ± 3.8 a
Alpha-Linolenic acid (C18:3)	1.4 ± 0.1 ab	1.5 ± 0.0 b	1.3 ± 0.1 a	1.5 ± 0.1 b	1.5 ± 0.1 ab
11,14-Eicosadienoic acid (C20:2)	5.5 ± 0.1 a	5.4 ± 0.3 a	5.5 ± 0.2 a	5.7 ± 0.2 a	5.6 ± 0.2 a
Total PUFAs	114.6 ± 4.1 a	113.5 ± 2.3 a	114.1 ± 5.2 a	115.7 ± 8.9 a	113.9 ± 3.9 a
Total fatty acids	182.9 ± 6.1 a	181.3 ± 3.0 a	180.4 ± 6.0 a	185.6 ± 11.3 a	181.5 ± 5.2 a
Total MUFAs + PUFAs	143.6 ± 5.0 a	142.3 ± 2.6 a	142.3 ± 5.5 a	144.8 ± 10.5 a	142.7 ± 4.5 a
Ratio MUFAs/SFAs	0.7 ± 0.0 a	0.7 ± 0.0 a	0.7 ± 0.0 a	0.7 ± 0.0 a	0.7 ± 0.0 a
Ratio PUFAs/SFAs	2.9 ± 0.0 a	2.9 ± 0.0 a	3.0 ± 0.2 a	2.8 ± 0.2 a	2.9 ± 0.1 a
Ratio MUFAs + PUFAs/SFAs	3.6 ± 0.0 a	3.7 ± 0.0 a	3.7 ± 0.2 a	3.6 ± 0.3 a	3.7 ± 0.1 a

Values ± SD (n = 5) with different letters along the raw are statistically different (p < 0.05).

. The results indicate no significant differences in the fatty acid profiles among the oil samples. While no similar studies on Nigella seeds were found in the literature, a study by Rabadán et al. [22] investigated almond, walnut, and pistachio oils screw press extracted at different temperatures (50, 100, 150, and 200 °C) and similarly found no significant effect of temperature on the fatty acid composition of these nut oils.

A total of 11 fatty acids were detected, collectively contributing between 180.4 and 185.6 mg/g oil in the studied samples. The saturated fatty acids (SFAs) included myristic, palmitic, margaric, stearic, and arachidic acids, while monounsaturated fatty acids (MUFAs) and polyunsaturated fatty acids (PUFAs) each compromised palmitoleic, oleic and 11-Eicosenoic acid, and linoleic, alpha-linolenic and 11,14-Eicosadienoic acid, respectively. The predominant fatty acids were linoleic, oleic, and palmitic acids, each exceeding 25 mg/g of oil in all samples, with linoleic acid being the most abundant at over 100 mg/g of oil. The total content of SFAs, MUFAs, PUFAs, and unsaturated fatty acids (UFAs) in the studied oil samples ranged between 38.1 and 40.7, 28.2 and 29.2, 113.5 and 115.7, and 142.3 and 144.8 mg/g of oil, respectively. Comparable results have been reported in previous studies on Nigella oils worldwide [51,52,67]. For example, Lutterodt et al. [51] investigated six different cold-pressed Nigella oils and found SFAs, MUFAs, and PUFAs within the ranges of 15.64–16.23, 23–24.91, and 59.02–61.36 g/100 g of total fatty acids, respectively, with linoleic, oleic and palmitic acids being major constituents.

In the present study, fatty acids accounted for approximately 18% of the total weight of the oil. Since previous studies on the fatty acid composition of Nigella oil from various sources have reported percentages based on the total observed fatty acids peak area %, understanding the contribution of fatty acids to the total weight of these Nigella oils to compare with present results remains challenging. Some studies have aimed to elucidate the fatty acid contribution in different oils [68,69]. For example, Adjepong et al. analyzed fatty acids in coconut oil, palm oil, vegetable oil, and palm kernel oil in Ghana, observing approximately 15%, 45%, 46%, and 19% fatty acids, respectively, relative to the total weight of these oils [68]. Similarly, Jumbe et al. investigated the fatty acid composition in sunflower oil, korie oil, and red palm oil in Tanzania, reporting approximately 37%, 87%, and 70%, respectively, of fatty acids relative to the total oil weight, which exceeds the values observed in the present study [69]. The Nigella oil is known to contain not only fatty acids but also substantial concentrations of other non-volatile components, such as sterols, tocopherols, saponins, and alkaloids [70,71]. Moreover, a considerable portion of the volatile compounds, with thymoquinone being one of the most prominent in the oil, have also been well documented [72,73,74,75]. Given the complex chemical composition of Nigella oil, it is likely that the remaining 82% is made up of both volatile and other nonvolatile components.

Furthermore, the ratios of MUFAs/SFAs, PUFAs/SFAs, and (MUFAS + PUFAS)/SFAs indicate the nutritional quality of the oil for human consumption. A higher ratio of MUFAs/SFAs, PUFAs/SFAs, and (MUFAs + PUFAs)/SFAs signifies lower SFAs and higher concentrations of UFAs in the seeds. In our study, no significant differences were observed in these ratios, which were approximately 0.7, 2.8–3, and 3.6–3.7, respectively. The fatty acid ratios observed in the present study are comparable with the values reported in the literature, including Saxena et al., who have reported MUFAs/SFAs, PUFAs/SFAs, and (MUFAs + PUFAs)/SFAs in the range between 0.4 and 1.3, 4 and 6.4, and 4.6 and 7.3, respectively, when studying different genotypes of Nigella oil in India [49]. The fatty acid ratios observed in the present study are comparable with other alternative sources of Nigella oil as well, such as oregano oil [76] and sea buckthorn oil [77]. Furthermore, both the higher and the lower values of PUFAs/SFAs than the values observed in the present study have been reported by Kostik et al. while investigating different edible oils [78]. Their report showed PUFAs/SFAs ranging from 0.005 to 10.55, with coconut oil at the low end (0.005) and safflower oil at the high end (10.5). The coconut oil, corn oil, cottonseed oil, palm kernel oil, olive oil, and peanut oil displayed lower PUFAs/SFAs values compared to the results obtained in this study. On the other hand, linseed oil, soybean oil, sunflower seed oil, safflower oil, and canola oil exhibited comparatively higher values [78].

3.5. Iodine Value (IV)

The IV provides information about the degree of unsaturation in oil. As the number of double bonds increases, it directly impacts IV [79,80]. The IV of Nigella oil extracted at different temperatures was observed in the range between 99.2 and 101.8 (

Table 5. IV in Nigella oil extracted at different temperatures.

Parameter	Oil Extraction Temperatures
	40 °C	50 °C	60 °C	70 °C	80 °C
IV	101.2	101.0	101.8	99.2	101.0

). The lowest value was in the oil extracted at 70 °C, and the highest value was in the oil extracted at 60 °C. The previous studies have reported varying IVs ranging from 98 to 128 for Nigella oil from different locations [70,81,82,83].

Given that no significant effect of extraction temperatures on fatty acid composition, TPC, CUPRAC, IV, and stable levels of TQ between 40 °C to 60 °C including the higher FRAP value at 60 °C were observed in the present study, it suggests that maintaining an extraction temperature of 60 °C may be optimal. This temperature choice can enhance resource efficiency, economic benefits, and product quality, thereby contributing to the overall sustainability of the production process.

Additionally, there has been a growing demand for Nigella oil in the international market driven by the needs of various industries such as pharmaceuticals, nutraceuticals, flavoring, cosmetics, and culinary industries [84,85,86]. For instance, according to the Black Seed Oil Market Research: Size, Trends, And Forecast 2033, assessed on 15 July 2024 [87], the global oil market for Nigella was valued at USD 20.6 million in 2023 and is expected to reach USD 42.5 million by 2033, with a compound annual growth rate (CAGR) of 7.7% from 2024 to 2033. Given the substantial quantity of Nigella seeds required to meet this demand, it is crucial to adopt an optimized extraction method that ensures both high oil yield and quality. This method not only enhances the efficiency of the extraction process but also guarantees that the final product meets the highest quality standards. By implementing these optimized extraction methods, producers can strike a balance between high production volumes and the integrity and potency of Nigella oil, thus securing a stable position in the competitive global market. At the same time, the large quantity of produced byproducts can also be utilized for different purposes, such as animal feeds, instead of being wasted [88,89]. This is not possible with solvent extraction methods due to the use of soap stock acids in chemical refining, bleaching earth, metals, and pigments, catalysts used in filters for the hardening process, as well as and mucilage from degumming and deodorizer distillate [90].

3.6. Correlation of Different Variables

Table 6. Correlations among different variables.

	TPC	FRAP	CUPRAC	TQ	C14:0	C16:0	C17:0	C18:0	C20:0	C16:1 (cis-9)	C18:1	C20:1 (cis-11)	C18:2	C18:3 (cis-9,12,15)	C20:2 (cis-11,14)	ΣSFA	ΣMUFA	ΣPUFA	ΣFAs	MUFA + PUFA	MUFA/SFA	PUFA/SFA
FRAP	0.088
CUPRAC	−0.078	0.279
TQ	0.654 **	0.039	−0.427 *
C14:0	−0.284	−0.034	0.165	0.068
C16:0	−0.027	−0.121	0.060	−0.159	−0.071
C17:0	−0.221	0.000	0.175	−0.021	0.895 **	−0.129
C18:0	0.224	−0.062	0.230	0.128	0.014	0.576 **	−0.007
C20:0	−0.238	−0.113	0.250	0.091	0.929 **	−0.058	0.829 **	0.073
C16:1(cis-9)	−0.258	−0.107	0.016	0.063	0.809 **	−0.235	0.719 **	−0.035	0.752 **
C18:1	0.143	0.026	0.219	0.011	−0.254	0.687 **	−0.268	0.896 **	−0.196	−0.305
C20:1 (cis-11)	−0.149	0.156	0.077	0.163	0.379	−0.235	0.339	0.040	0.397 *	0.396	−0.080
C18:2	0.148	0.085	0.152	0.071	−0.403 *	0.382	−0.389	0.673 **	−0.373	−0.373	0.773 **	−0.068
C18:3 (cis-9,12,15)	−0.460 *	−0.497 *	−0.146	−0.288	0.420 *	0.275	0.467 *	0.004	0.399 *	0.267	−0.086	−0.088	−0.317
C20:2 (cis-11,14)	−0.200	−0.091	0.072	−0.170	−0.240	0.498 *	−0.289	0.244	−0.287	−0.411 *	0.451 *	−0.209	0.495 *	0.063
ΣSFA	−0.039	−0.131	0.130	−0.109	0.131	0.970 **	0.062	0.679 **	0.151	−0.060	0.708 **	−0.128	0.369	0.332	0.429 *
ΣMUFA	0.112	0.033	0.232	0.032	−0.155	0.662 **	−0.180	0.917 **	−0.099	−0.190	0.989 **	0.045	0.752 **	−0.072	0.407 *	0.709 **
ΣPUFA	0.126	0.068	0.149	0.056	−0.397 *	0.404 *	−0.385	0.674 **	−0.371	−0.379	0.779 **	−0.078	0.999 **	−0.286	0.532 **	0.390	0.757 **
ΣFAs	0.104	0.028	0.177	0.024	−0.296	0.621 **	−0.306	0.801 **	−0.263	−0.330	0.898 **	−0.080	0.954 **	−0.155	0.560 **	0.624 **	0.883 **	0.960 **
MUFA + PUFA	0.128	0.064	0.169	0.054	−0.369	0.464 *	−0.363	0.740 **	−0.336	−0.360	0.844 **	−0.059	0.992 **	−0.259	0.529 **	0.459 *	0.826 **	0.993 **	0.981 **
MUFA/SFA	0.189	0.225	0.116	0.185	−0.377	−0.446 *	−0.321	0.256	−0.339	−0.161	0.318	0.238	0.462 *	−0.544 **	−0.044	−0.430 *	0.332	0.441 *	0.290	0.438 *
PUFA/SFA	0.141	0.175	0.027	0.138	−0.493 *	−0.391	−0.430 *	0.075	−0.493 *	−0.318	0.162	0.031	0.672 **	−0.553 **	0.191	−0.436 *	0.139	0.657 **	0.424 *	0.591 **	0.755 **
MUFA + PUFA/SFA	0.152	0.187	0.040	0.148	−0.492 *	−0.410 *	−0.428 *	0.102	−0.487 *	−0.306	0.187	0.059	0.663 **	−0.568 **	0.165	−0.447 *	0.169	0.647 **	0.418 *	0.587 **	0.810 **	0.996 **

** Correlation is significant at the 0.01 level; * correlation is significant at the 0.05 level; the following sample size of the variables was used to study the correlation: fatty acids (n = 25), TPC, antioxidant capacity, and TQ (n = 50); three colors scale, red, white and green, was used to represent the combination between −1, 0, and 1, respectively.

presents the results of Pearson’s correlation analysis to explore potential connections among different phytoconstituents, including TPC, antioxidant capacity, TQ, individual fatty acids, and their ratios. Upon examining the table, it is evident that no significant correlations were observed among TPC, FRAP, and CUPRAC. Additionally, TQ did not correlate with the other variables, except for TPC and CUPRAC. In these cases, TQ exhibited a moderate positive correlation with TPC (r = 0.654, p < 0.01) and a weak negative correlation with CUPRAC (r = −0.427, p < 0.01). Furthermore, no significant correlations were found between TPC, CUPRAC, and FRAP with individual fatty acids, except for C18:3, where TPC and FRAP showed a weak negative correlation (r = −0.460, p < 0.05 and r = −0.497, p < 0.01, respectively). Moreover, SFA and PUFA showed a strong positive correlation with MUFA (r = 0.709, p < 0.01 and r = 0.757, p < 0.05, respectively), but there was no significant correlation between SFA and PUFA. Given the limited availability of correlation analyses in studies involving Nigella oil samples, further research is necessary to validate these findings and explore additional relationships among these phytoconstituents.

4. Conclusions

The study investigated the effect of screw press extraction temperatures on the oil yield and key phytochemical characteristics of Nigella oil. The results showed increasing of Nigella oil yield with higher screw press temperatures between 40 and 80 °C, while TQ content decreased with increasing temperatures. The study also revealed that the ratios of fatty acids, TPC, and CUPRAC in the oil samples extracted at five different temperatures were comparable, except for the FRAP value, which was notably higher in the oil extracted at 60 °C. Considering the comprehensive assessment of oil yield, TPC, antioxidant capacity (FRAP and CUPRAC), TQ levels, fatty acid ratios, and IV, it is concluded that extracting Nigella oil at 60 °C yielded superior results with minimal loss of oil yields and contents of valuable bioactive compounds.

Additionally, the study found no strong correlations between TPC, FRAP, CUPRAC, and thymoquinone levels. Similarly, there were no strong correlations of TPC, CUPRAC, and FRAP with individual fatty acids, suggesting that the antioxidant capacity and phenolic content of the oil are influenced by a complex interplay of factors rather than being directly linked to specific fatty acids or thymoquinone content. This highlights the importance of considering multiple parameters when evaluating the quality and health benefits of Nigella oil.