Abstract
This study examined the oxidation stability and volatile compounds in samples of black cumin seed oil at 60 and 100°C. Oxidative changes of black cumin oils followed by periodic determination of its conjugated dienes and trienes during accelerated conditions (60 and 100°C). Black cumin oil showed high resistance to oxidation during thermal oxidation (60 and 100°C). In addition to oxidative stability, volatile compounds in the oil were determined by headspace solid phase microextraction with gas chromatography-mass spectrometry analysis technique. The major componenent of the oil was p-cymene (44.77%) followed by thymoquinone (28.62%). Some volatile compounds were lost rapidly during thermal oxidation at the end of storage. Levels of thymoquinone, 4-terpineol, α-longipinene, carvacrol, and isolongifolene reduced slowly and remained more stable during storage at two temperatures (60 and 100°C).
INTRODUCTION
Nigella sativa L. is an annual plant of the family Ranunculacea. It is native to the Mediterranean region, through west Asia to northern India. The plant is known as known as “black cumin” and as “çörek otu” in Turkey. Black cumin seeds have been used as a spice from ancient times and were mainly used for medicinal purposes, to treat a variety of health conditions.[Citation1,Citation2] In addition to medicinal uses, the seeds are commercially important for their uses in food production, as a condiment in some meat dishes, and as a means of enriching the flavor of bakery products and cheese.
The major compounds of Nigella seeds are essential oil (volatile oil) and fixed oil (non-volatile oil, as also called fatty oil). Nigella essential oil possess anti-inflammatory, antimicrobial, anticancer,[Citation3,Citation4] and antioxidant activity.[Citation5,Citation6] The volatile oil content of seeds averages 0.5 to 1.4%.[Citation2] Harzallah et al.[Citation4] reported that the main constituents of Nigella essential oil from Tunisia were p-cymene (49.48%), α-thujene (18.93%), α-pinene (5.44%), β-pinene (4.31%), and γ-terpinene (3.69%).
Black cumin seed oil is generally obtained by the cold press method. The oil of seeds is used for consumption purposes. These oils have stronger radical scavenger activity than some other vegetable oils.[Citation7] Nigella seed oil has better oxidation stability than coriander and niger seed oils at 60°C.[Citation8] Some spices have fixed and volatile oil. Some volatile compounds migrate to oil during cold pressing of these seeds. The oxidation stability of oil has been determined by some researchers but there is no study within the literature about alteration of volatile compounds in black cumin oil during accelerated oxidation. This oxidation conditions (60 and 100°C) give rapid information on the oxidative stability of oils. These storage conditions may be correlated with cooking applications. Volatile compounds are important part of black cumin oil, so alteration of volatile compounds in black cumin oil could be determined in these storage conditions.
In the present study, black cumin seed oil was subjected to accelerated oxidation conditions (60 and 100°C) and analyzed by HS-GC/MS. The main objective was to determine volatile compounds under these conditions.
EXPERIMENTAL PROCEDURES
Material
Pressed black cumin oil was purchased from an essential oil factory (NU-KA) and refrigerated at approximately +4°C until use for storage and analysis.
Methods
Accelerated oxidation condition
Samples of oil weighing 12 g (±0.01 g) were placed in 20 mL clear glass bottles. The oxidation reaction was accelerated in a forced-draft air oven set at 60 ± 2°C for 0, 2, 4, 10, and 18 days. A second series of tests was applied at 100°C under the same conditions mentioned above. The samples were removed from the oven after 0, 1, 2, 3, 5, 7, and 9 h. The conjugated dienes (CDs) and conjugated trienes (CTs) are used to evaluate oxidation products, these values were determined by measuring the specific extinctions at 232 and 270 nm. Samples were diluted with n-hexane in a cell of 1 cm path length and K232 and K270 were calculated following the standard method (Ch 5–91 methods of in American Oil Chemist’s Society).[Citation9] In addition to oxidation test, samples were analyzed for volatile compounds with two replications.
Volatile Compounds Analyses
Approximately 1.5 g of the sample was inserted into a 20 mL headspace screw top vial and allowed to equilibrate for 15 min at 35°C. The headspace of the samples was extracted for 45 min at 35°C using a CTC Combi PAL auto sampler with 75 μm carboxen/polydimethylsiloxane (CAR/PDMS) solid phase micro extraction (SPME) fiber. The volatile compounds were desorbed by directly inserting the fiber for 10 min into the injection port of the GC maintained at 250°C.
Analyses of volatile compounds were performed using an Agilent model 7890 series gas chromatograph in combination with a CTC Combi PAL autosampler and an Agilent 5975 N mass selective detector. The compounds were separated in a DB-624 (J&W Scientific, 30 m, 0.25 mm i.d., 1.4 μm film thickness), working with the following temperature program: 40°C, hold for 5 min; 3°C/min up to 110°C; 4°C/min up to 150°C; 10°C/min up to 210°C, hold for 12 min. The temperatures for the injection port, ion source, quadrupole and interface were set at 250, 230, 150, and 240°C, respectively. Mass spectra were obtained in the electron impact at 70 eV in full scan and a scan range from m/z 41–400. Identification of compounds detected by comparing mass spectra and Kovats Index (KI) with the authentic standards and published data, as well as by comparing their mass spectra with the MS library of Nist05 and Wiley7.0. KI were calculated using the series of n-hydrocarbons (C7-C20).
Statistics
Statistical analyses were conducted using SPSS (Statistical Program for Social Sciences, SPSS Corporation, Chicago, IL, USA) version 16.0 for Windows. Experimental data were evaluated using analysis of variance (ANOVA) and significant differences between the means of the groups from volatile compounds during storage (p < 0.05) were determined by Duncan’s multiple range test, using SPSS (version 16.0 for Windows).
RESULTS AND DISCUSSION
Incubation at 60°C
CDs and CTs were determined using the spesific absorptivity at 232 and 270 nm in the UV region.[Citation10] shows CDs (K232) and CTs (K270) of samples during 18 days storage. Up to 10 days, the CDs showed no differences between samples. After 10 days, the CD value of the oil reached 27.08 from an initial value of 8.06. The variation of CTs was stabile during storage at 60°C. CT is a measure of secondary oxidation products such as unsaturated ketones and aldehydes and generally increased during the middle or late phase of oxidation.[Citation11]
Table 1 CD and CT of black cumin oil stored at 60°C
Black cumin seed oil contained the lowest level of oxidation products up to 10 days according the CD data of the oils. The results were similar to those obtained by Ramadan and Mörsel,[Citation7] who found that black cumin oil exhibited strong oxidation stability according CD and CT data during 21 days storage at 60°C.[Citation8] Pressed black cumin oil showed good antioxidant activity. The pressed oils from black cumin contain fixed oil and volatile compounds, which showed antiradical and antioxidant activity.[Citation12] The results of GC-MS analysis of the volatile compounds in black cumin oil () indicate that the oil was characterized mainly by terpenes; thymoquinone and p-cymene are major compounds in the volatile fraction of black cumin oil, with relative concentrations of 44.77 and 28.62%, respectively ().
Table 2 Volatile compounds quantified as AU × 10−4 in the headspace of black cumin oils during storage at 60°C
Volatile compounds in oils changed with oxidation at 60°C. Most of terpenes decreased during oxidation at 60°C. Most of them disappeared quickly with time. The levels of 1,8-cineole, camphor, and thymol could not be detected after two days of storage. The α-pinene, β-myrcene, and β-thujone levels could not be detected at the fourth day, whereas β–pinene and α-phellandrene was lost in moderate time (10th day). Sabinene, α-terpinene, limonene, γ-terpinene, α-terpinolene, p-cymen-8-ol, cis- and trans-dihydrocarvone content decreased at the end of storage (18th day) at 60°C. In addition to these compounds, p-cymene decreased dramatically during storage.
Some compounds were resistant to oxidation and decreased slowly with oxidation of oil. Among the volatile compounds of oils, thymoquinone, 4-terpineol, bornyl acetate, α-longipinene, carvacrol, and isolongifolene remained stable. Some volatile products from lipid oxidation occurred with increasing oxidation. Hexanal, 2-pentyl furan, cis-2-heptenal increased. Pentenal, 2-hexenal, 2,4-heptadienal, trans-2-octenal, nonanal and 2,4-nonadienal occurred after oxidation progress and increased up to 217.01, 136.18, 375.51, 842.59, 114.24, and 74.46 AU×10−4, respectively. After 18 days storage, remarkable differences were observed in the CDs of oxidized oils; these compounds are related to oxidation.
Incubation at 100°C
shows the CD and CT developments during the storage of black cumin seed oils at 100°C for 9 h. There was no significant increase in these values during storage (p > 0.05). Volatile compounds varied during storage at 100°C (). The volatile components 2-pentyl furan, β-thujone, camphor, p-cymen-8-ol, and thymol disappeared after 2 h of storage. In addition to these compounds, β-myrcene, and 1,8-cineole content disappeared at the ninth hour of storage at 100°C.
Table 3 CD and CT of black cumin oil stored at 100°C
Table 4 Volatile compounds quantified as AU × 10−4 in the headspace of black cumin oils during storage at 100°C
The major volatile compounds p-cymene and thymoquinone decreased with storage duration and showed a sudden reduction between 7 to 9 h, from 23392.44 to 15066 AU×10−4. In addition to major, from the seventh hour onwards, sabinene, β-pinene, α-phellandrene, γ-terpinene, α-terpinolene reduced by approximately half. Headspace sampling technique showed that the α-pinene and α-thujone contents of the oil decreased significantly (p ≤ 0.05) after 5 h of storage.
The levels of 4-terpineol, α-longipinene, carvacrol, isolongifolene, cis- dihydrocarvone, and trans-dihydrocarvone decreased gradually during storage and the tolerances of these compounds towards to oxidation were comparable and even better than that of other compounds in black cumin oil.
Hexanal, cis-2-heptenal, and nonanal content of oils increased with storage time. Hexanal is formed oxidation of linolenic acid (C18:2) and its content increased with storage in potato chips fried sunflower oil,[Citation13] and some vegetable oils.[Citation14] However, pentanal formation increased up to 5 h storage and on the seventh hour of storage, the level of pentanal decreased during storage and at the ninth hour this compound disappeared.
Singh et al.[Citation15] studied the antioxidant effect of black cumin essential oil and its acetone extract on rapeseed oil. The essential oil and extract were able to reduce the formation of hydroperoxides and showed antioxidant activity in model systems. Burits and Bucar[Citation4] found that thymoquinone, carvacrol, trans-anethole, and 4-terpineol from black cumin oil had strong antiradical activity. In addition to essential oil and extracts, some cold pressed black cumin oils showed strong resistance to oxidation.[Citation15] In this study, black cumin oil showed stronger resistance to oxidation during storage in two accelerated models. In this respect, the results are in agreement with those reported.[Citation15] The antioxidant activity of black cumin oil may be caused by phenolic compounds and individual phenolic compounds, as indicated by the results.[Citation5,Citation15] However, there was no information about volatile compounds in oil during oxidation at accelerated storage conditions. Thymoquinone, 4-terpineol, α-longipinene, carvacrol, and isolongifolene remained more stable than other compounds in the oil under the two storage conditions. Burits and Bucar[Citation5] emphasize the antiradical activity of thymoquinone, carvacrol and 4-terpineol from black cumin oil and Lutterodt et al.[Citation12] suggested that thymoqinone may contribute to the oxidation stability of black cumin fixed oil.
CONCLUSION
In conclusion, cold pressed nigella oil showed good oxidation stability at 60 and 100°C. Major volatile compounds in Nigella oil such as thymoquinone showed stability during these accelerated conditions. Terpenes and other some volatile compounds dramatically decreased with oxidation increased. Some volatile oxidation compounds such as pentanal, cis-2-heptenal occurred at the end of storage temperatures (60 and 100°C).
REFERENCES
- Al-Saleh, I.A.; Billedo, G.; El-Doush II. Levels of selenium, DL-α-tocopherol, DL-γ-tocopherol, all-trans-retinol, thymoquinone, and thymol in different brands of Nigella sativa seeds. Journal of Food Composition and Analysis 2006, 19, 167–175.
- Malhotra, S.K. Nigella. Handbook of Herbs and Spices Edited by K.V. Peter. CRC Press: Washington, DC., 2004.
- Bourgou, S.; Pichette, A.; Marzouk, B.; Legault, J. Bioactivities of black cumin essential oil and its main terpenes from Tunisia. South African Journal of Botany 2010, 76, 210–216.
- Harzallah, H.J.; Kouidhi, B.; Flamini, G.; Bakhrouf, A.; Mahjoub, T. Chemical composition, antimicrobial potential against cariogenic bacteria and cytotoxic activity of Tunisian Nigella sativa essential oil and thymoquinone. Food Chemistry 2011, 129, 1469–1474.
- Burits, M.; Bucar, F. Antioxidant activity of Nigella sativa essential oil. Phytotherapy Research 2000, 14, 323–328.
- Erkan, N.; Ayranci, G.; Ayranci, E. Antioxidant activities of rosemary (Rosmarinus officinalis L.) extract, blackseed (Nigella sativa L.) essential oil, carnosic acid, rosmarinic acid, and sesamol. Food Chemistry 2008, 110, 76–82.
- Ramadan, M.F.; Mörsel, J.T. Antiradical performance of some common and nontraditional vegetable oils. Inform 2004, 15, 553–555.
- Ramadan, M.F.; Mörsel, J.T. Oxidative stability of black cumin (Nigella sativa L.), coriander (Coriandrum sativum L.), and niger (Guizotia abyssinica Cass.) crude seed oils upon stripping. European Journal of Lipid Science and Technology 2004, 106, 35–43.
- American Oil Chemists’ Society. Determination of specific extinction of oils and fats, ultraviolet absorption. In: Official methods and recommended practices of the American Oil Chemists Society. Official method, 2005, Ch 5–91.
- Wanasundara, U.N.; Shahidi, F. Canola extract as an alternative natural antioxidant for canola oil. Journal of the American Oil Chemists’ Society 1994, 71, 817–822.
- Shahidi, F.; Zhong, Y. Lipidoxidation: Measurement methods. In: Bailey’s Industrial Oil and Fat Products; Shahidi, F.; Ed.; John Wiley & Sons: Hobiken, NJ, USA, 2005; Vol. 1 pp. 357–386.
- Lutterodt, H.; Luther, M.; Slavin, M.; Yin, J.J.; Parry, J.; Gao, J.M.; Yu, L.L. Fatty acid profile, thymoquinone content, oxidative stability, and antioxidant properties of cold-pressed black cumin seed oils. LWT-Food Science and Technology 2010, 43, 1409–1413.
- Lee, J.H.; Pangloli, P. Volatile compounds and storage stability of potato chips fried in mid-oleic sunflower oil. International Journal of Food Properties 2013, 16 (3), 563–573.
- Jeleń, H.H.; Obuchowska, M.; Zawirska-Wojtasiak, R.; Wasowicz, E. Headspace solid-phase microextraction use for the characterization of volatile compounds in vegetable oils of different sensory quality. Journal of Agricultural and Food Chemistry 2000, 48, 2360–2367.
- Singh, G.; Marimuthu, P.; de Heluani, C.S.; Catalan, C. Chemical constituents and antimicrobial and antioxidant potentials of essential oil and acetone extract of Nigella sativa seeds. Journal of the Science of Food & Agriculture 2005, 85, 2297–2306.
- Olivares, A.; Navarro, J.L.; Flores, M. Effect of fat content on aroma generation during processing of dry fermented sausages. Meat Science 2011, 87, 264–273.
- Kaban, G. Volatile compounds of traditional Turkish dry fermented sausage (Sucuk). International Journal of Food Properties 2010, 13 (3), 525–534.
- Marco, A.; Navarro, J.L.; Flores,M. Quantitation of selected odor-active constituents in dry fermented sausages prepared with different curing salts. Journal of Agricultural and Food Chemistry 2007, 55, 3058–3065.