Content uploaded by Svetlana Momchilova
Author content
All content in this area was uploaded by Svetlana Momchilova on Aug 08, 2017
Content may be subject to copyright.
65
Bulgarian Chemical Communications, Volume 49, Special issue G (pp.65 –70) 2017
Fatty acids, tocopherols and oxidative stability of hazelnuts during storage
S.M. Momchilova1*, S.P. Taneva1, М.D. Zlatanov2,
G.A. Antova2, M.J. Angelova-Romova2, E. Blagoeva3
1 Institute of Organic Chemistry with Centre of Phytochemistry, Bulgarian Academy of Sciences,
Acad. G. Bonchev Str., bl. 9, 1113 Sofia, Bulgaria
2 Department of Chemical Technology, University of Plovdiv “Paisii Hilendarski”,
24 Tzar Assen Str., 4000 Plovdiv, Bulgaria
3Agricultural Experiment Station, 1 Minjorska Str., 6600 Kardzhali, Bulgaria
Received November 13, 2016; Revised December 6, 2016
Hazelnuts of three cultivars (Ata Baba, Ran Trapezundski and Tonda Gentile) grown in Bulgaria were stored for 1,
3, 6 and 12 months at different conditions: unshelled (in shell) or shelled (kernels), at 4oC (fridge) or at 20oC (in
shadow), and then were analyzed about fatty acids composition, tocopherols and oxidative stability (induction period)
of their oil. The results revealed that fatty acids were not practically changed up to 12 months in spite of the different
storage conditions whereas the tocopherols amount gradually decreased and that trend was slightly stronger at 20oC
than at 4oC as well as for nuts in shell than the kernels. Concerning oxidative stability, the oil from initial hazelnuts had
long induction period (40, 47 and 58 hours for Ata Baba, Ran Trapezundski and Tonda Gentile cultivars, respectively).
This oxidative stability gradually decreased during the storage period with the same trend, namely the hazelnuts stored
at 20oC in shell had slightly shorter induction periods than the corresponding kernels stored at 4oC. All these results
indicate that it is possible to store hazelnuts up to 12 months without considerable and harmful changes in their main
lipid characteristics. In order to keep their quality at most it is preferable to store them as kernels at 4oC.
Key words: hazelnuts, storage, fatty acids, tocopherols, oxidative stability
INTRODUCTION
Nuts have been known and used as food since
ancient times [1] and during the years their health
benefits have been clearly recognized [2, 3].
Among all nuts hazelnuts are preferable because of
their delicious taste, delicate flavor and beneficial
nutritional properties. However, due to high fat
content (above 60%) they tend to deteriorate as a
result of lipid oxidation. Since the hazelnuts are
consumed mainly as kernels the storage conditions
are of crucial importance to keep their quality for
longer periods. In spite of the significance of that
problem, there are not too many publications on the
effects of storage conditions on the hazelnut lipid
characteristics. Up to now the following have been
studied: the storage atmosphere (air or enriched
with oxygen vs. inert gases or vacuum [4‒8]),
temperature (mainly 4-7oC vs. 20-25oC [4, 7‒10], in
single cases ‒25oC [8, 11] or even 55oC [12]), the
presence of nut shell [4, 5, 9, 10, 13]. The storage
periods are usually up to 12 months [4, 5, 7‒12,
14], but also periods of two [13] or four [15] years
have been tested. The investigated hazelnut
characteristics are: textural [6, 11] or sensory [6, 7,
10] attributes, total phenolics [7, 8, 10], acidity
and/or peroxide value [4, 7, 10, 13], hexanal
content [7, 8], conjugated dienes and trienes [4],
α-tocopherols [12, 13], fatty acids [5, 9, 13‒15].
Unfortunately, in these investigations the effects of
only single storage condition on particular lipid
attribute have been examined. Moreover, some
results are contradictory. For that reasons the aim of
our work was to study the effects of the two most
popular and easy to perform even at home storage
conditions, namely the temperature (4oC in fridge
and 20oC in shadow) and the presence of nut shell,
on the fatty acids, tocopherols and oxidative
stability (Induction period in hours) of the oil from
hazelnuts stored up to 12 months. Thus, some more
general recommendations could be given in order to
save the quality of nuts not only as
industrial/market product but also as preferable
dietary stored at home.
EXPERIMENTAL
Samples
Hazelnut cultivars Ata Baba (Corylus pontica C.
Koch), Ran Trapezundski (Corylus maxima Mill.)
and Tonda Gentile (Corylus avellana L.) were
grown in orchards near the town of Kardzhali,
Bulgaria. Fruits of three consecutive crops (2009-
2011) were collected and corresponding portions of
them were stored as kernels (shelled) or in shell,
both types in polyethylene bags, at room
temperature (20°C, in shadow) or in refrigerator
* To whom all correspondence should be sent.
E-mail: svetlana@orgchm.bas.bg
© 2017 Bulgarian Academy of Sciences, Union of Chemists in Bulgaria
S.M. Momchilova et al.: “Fatty acids, tocopherols and oxidative stability of hazelnuts during storage”
66
(4°C) for 1, 3, 6 and 12 months. Initial portions of
hazelnuts were used as respective references (0
months storage). Reagents
All reagents and solvents were of analytical
grade and were used without additional
purification. The reference fatty acid methyl esters
and tocopherols were from Sigma-Aldrich
Chemical Co. (St. Louis, MO, USA).
Extraction of lipids
Hazelnuts (each sample of about 20 g kernels
after removing the shell) were ground and the oil
was extracted with n-hexane in Soxhlet apparatus
for 8 h [16]. The solvent was distilled in rotary
evaporator and stock hexane solution of the oil was
prepared.
Analysis of fatty acid (FA) composition
Fatty acids were determined by gas
chromatography (GC) of their methyl esters
(FAME). For the purpose, about 50 mg oil were
transmethylated with 1 % sulfuric acid in methanol
[17]. Then the FAME were purified by preparative
silica gel thin-layer chromatography (TLC) on 20 x
20 cm glass plates developed with hexane-acetone
(100:6, v/v) mobile phase. A drop of methyl oleate
solution was used as a reference near the edge of
the plate. After development, the zones were
visualized under UV light (366 nm) after spraying
with 0.1 % ethanolic 2`,7`-dichlorofluorescein.
FAME zone (Rf ~ 0.7) was scraped and eluted with
diethyl ether in a small glass column. The solvent
was evaporated under a gentle stream of nitrogen
and the rest was dissolved in hexane to give 1 %
solution of FAME. GC was performed on a Trace
GC Ultra (Thermo Scientific, Bremen, Germany)
gas chromatograph equiped with FID and a
DB-225 60 m x 0.25 mm x 0.25 µm column
(J&W Scientific, USA). The column temperature
was programmed from 150оC to 240°C with
5°C/min and held at this temperature for 20 min.
The injector and detector temperatures were 260oC
and 280оC, respectively. Helium was the carrier gas
at flow rate of 1.0 mL/min. Reference FAME
mixture was used for peak identification according
to the retention times. The analyses were performed
in triplicate. The software package XcaliburTM 2.0,
Revision 2.0 SR 1 (Thermo Scientific, Bremen,
Germany) was used to record and process the data.
Analysis of tocopherols
Tocopherols were analyzed directly by HPLC
on 250 mm x 4 mm Nucleosil Si 50-5 column
(Macherey-Nagel), eluted by hexane-dioxane
(96:4, v/v) and fluorescent detection at 290 nm
excitement and 330 nm emission [18].
Determination of oxidative stability
Oxidative stability (Induction period in hours)
was measured at 100°C (3 g oil sample, air flow 20
L/h) by Rancimat 679 (Metrohm, Switzerland)
equipment. Data processing
Mean values from three consecutive years, each
with three parallel measurements, were compared
by Student`s t-test (Microsoft Excel software) [19].
RESULTS AND DISCUSSION
Fatty acid composition
The main fatty acids in the investigated
hazelnuts are oleic (18:1, from 73.3 % to 80.4 %
depending on the cultivar, Fig. 1), linoleic (18:2,
with values of 7.2 %, 9.8 % and 14.5 % for Tonda
Gentile, Ran Trapezundski and Ata Baba cultivars
respectively, Fig. 2), palmitic (16:0, in the range 6.8
‒ 7.8 %, Fig. 3) and stearic (18:0, between 2.3 %
and 2.8 %, Fig. 4). Other fatty acids as palmitoleic
(16:1), linolenic (18:3), arachidic (20:0) etc. are
below 0.2 %. As could be seen (Figs. 1 ‒ 4), the
cultivars Tonda Gentile and Ata Baba differ almost
twice in their linoleic acid proportion. Nevertheless,
fatty acids composition of the investigated
hazelnuts in general is similar to that of Turkish [5],
Italian, Spanish [12, 20], Croatian, French [21]
cultivars. Unlike the available information about
fresh hazelnuts the findings published about fatty
acids composition of stored nuts are not too many,
not systematic and indeed discrepant. Moreover,
usually the effect of only single storage condition
on fatty acids has been examined. Thus,
Koyuncu et al. [14] have observed in kernels from
three Turkish cultivars slight decrease of 18:2 after
the 6th month storage at 21 oC in vacuum bags,
without effect of the shell presence or absence on
the fatty acids composition [5]. Other authors [9]
have noticed increase of 18:0 (from 2.6 % to 3.1 %)
after the 8th month and decrease of 18:3 (being 0.1
% in the initial oil) after the 12th month in whole
hazelnuts stored at 20 oC but without such changes
in kernels stored at 4 oC. Unfortunately, no
comparative experiments with whole nuts stored at
4oC and kernels at 20oC were implemented.
On the other hand, Ebrahem et al. [13] have
shown that 18:1 and 18:2 have not changed in
kernels and in whole hazelnuts stored at 0 oC up to
12 months. Similarly, Beyhan et al. [15] have
stored whole hazelnuts in warehouse up to 4 years
practically without differences in their fatty acid
composition. In addition to these results, our
S.M. Momchilova et al.: “Fatty acids, tocopherols and oxidative stability of hazelnuts during storage”
67
experiments confirme that fatty acids have not been
changed during hazelnut storage up to 12 months
irrespective of different conditions such as
temperature (4 oC or 20 oC) and the presence or
absence of shell, as well as irrespective of the
differences in the initial fatty acid composition of
the three analyzed cultivars (Figs. 1 ‒ 4).
Fig. 1. Oleic acid (9-18:1) content of the oil from three cultivars hazelnuts stored up to 12 months at different
conditions: at 20oC or 4oC, as kernels (k) or in shell (s).
Fig. 2. Linoleic acid (9,12-18:2) content of the oil from three cultivars hazelnuts stored up to 12 months at different
conditions: at 20oC or 4oC, as kernels (k) or in shell (s).
Fig. 3. Palmitic acid (16:0) content of the oil from three cultivars hazelnuts stored up to 12 months at different
conditions: at 20oC or 4oC, as kernels (k) or in shell (s).
Fig. 4. Stearic acid (18:0) content of the oil from three cultivars hazelnuts stored up to 12 months at different
conditions: at 20oC or 4oC, as kernels (k) or in shell (s).
Tocopherols
Tocopherols are important biologically active
substances with significant role of strong natural
antioxidants. Since their availability prevents the
oils from lipid oxidation higher amount of
tocopherols is valuable and desirable virtue.
Hazelnut oil from different varieties contains
tocopherols in quite wide range: from 115 to
S.M. Momchilova et al.: “Fatty acids, tocopherols and oxidative stability of hazelnuts during storage”
68
600 mg/kg [12, 20 ‒ 24]. Oils from the three
cultivars presented here have quite high initial
amounts: 560 mg/kg in Ran Trapezundski,
520 mg/kg in Ata Baba and 500 mg/kg in Tonda
Gentile (Fig. 5).
Fig. 5. Total tocopherols in the oil from three cultivars hazelnuts stored up to 12 months at different conditions: at 20oC
or 4oC, as kernels (k) or in shell (s).
Fig. 6. Oxidative stability (induction period) of the oil from three cultivars hazelnuts stored up to 12 months at different
conditions: at 20oC or 4oC, as kernels (k) or in shell (s).
After the first storage month these amounts
gradually decrease and that trend is slightly
stronger at 20oC than at 4oC as well as for nuts in
shell than the kernels. Likewise, respective
reducing of the individual α-, β- and -isomers is
observed (not shown here). As can be seen (Fig. 5)
hazelnuts stored at 4oC as kernels contain slightly
more tocopherols than that stored at 20oC in shell,
irrespective of the differences between the three
cultivars. Only two publications were found in the
literature concerning determination of tocopherols
during hazelnuts storage. They revealed that (i) the
remaining amount of α-tocopherol after 9 months
storage of kernels was higher at lower temperature
(three investigated cultivars) [12], and (ii) hazelnuts
stored up to 12 months at 0oC and at 10oC
contained slightly higher amount of α-tocopherol as
kernels than in shell (one investigated cultivar)
[13]. Despite the scarce information, these findings
confirm our results that storage of hazelnuts at 4oC
as kernels is preferable for keeping their quality for
longer period. Oxidative stability
The oxidative stability of oils is important
indicator for their admissible shelf-life. It depends
on the fatty acid composition, the availability of
natural antioxidants (tocopherols, etc.) as well as on
the presence of synergists (e.g. phytosterols). The
oxidative stability of hazelnut oil is among the
highest for vegetable oils: 16 h of refined [24, 25]
or 20 ‒ 40 h of cold-pressed [26] samples (at the
same experimental conditions as presented here).
The fresh oils from investigated here three cultivars
have even better indexes, i.e. 40 h, 48 h and 59 h
for Ata Baba, Ran Trapezundsli and Tonda Gentile,
respectively (Fig. 6).
During storage of nuts, these values have
gradually decreased and some effect of the
temperature and shell can be seen, namely the
decrease of oxidative stability is stronger at 20oC
than at 4oC as well as the kernels have slightly
higher stability than nuts in shell. Some reasonable
interpretation of these results might be the moisture
held by the shell which induces/favors undesirable
processes as lipid oxidation and degradation.
Comparing the three analyzed cultivars, the highest
oxidative stability of Tonda Gentile can be
explained by the lowest percentage of linoleic acid
(Fig. 2) regardless of the lowest amount of
tocopherols (as main antioxidant in that oils,
Fig. 5). Unfortunately, no data in the literature have
been found to compare with these results excepting
only partial characteristics of lipid oxidation such
as acidity, peroxide value [4, 7, 10, 13], antioxidant
capacity (expressed as Trolox equivalent [8]) or
conjugated dienes and trienes [4]. Thus, the authors
have observed gradual increase during storage of
acidity and peroxide values at that in greater extent
at higher temperature (25oC vs. 7oC for kernels and
whole nuts [4]; or 20oC in shell vs. kernels at
4oC [10]). On the other hand, no significant effects
of the storage conditions (temperature and shell) on
the antioxidant capacity (as Trolox equivalent, [8])
S.M. Momchilova et al.: “Fatty acids, tocopherols and oxidative stability of hazelnuts during storage”
69
as well as on the conjugated dienes and trienes [4]
have been observed.
CONCLUSION
Fatty acids have not been changed significantly
during hazelnuts storage up to 12 months
irrespective of different conditions such as
temperature (4oC or 20oC) and the presence or
absence of shell, as well as irrespective of the
differences in the initial fatty acid composition of
the three analyzed cultivars. The fresh oils have
quite high initial amounts of tocopherols which
gradually decrease during storage and that trend is
slightly stronger at 20oC than at 4oC as well as for
nuts in shell than as kernels. The oxidative stability
of the fresh oils decrease similarly during the
hazelnuts storage. Thus, it is possible to store
hazelnuts up to 12 months without considerable and
harmful changes in their main lipid characteristics.
In order to keep their quality at most it is preferable
to store the hazelnuts as kernels at 4oC.
Acknowledgements: Partial financial support from the
Bulgarian Fund for Scientific Research (project DO 02-
239) was gratefully acknowledged.
REFERENCES
1. E. Ros, Nutrients, 2, 652 (2010).
2. M. L. Dreher, C. V. Maher, P. Kearney, Nutr. Rev.,
54, 241 (1996).
3. J. Mukuddem-Petersen, W. Oosthuizen, J. C. Jerling,
J. Nutr., 135, 2082 (2005).
4. M. B. San Martin, T. Fernandez-Garcia, A. Romero,
A. Lopez, J. Food Proc. Preserv., 25, 309 (2001).
5. M. A. Koyuncu, J. Food Qual., 27, 304 (2004).
6. R. Massantini, M. Cortini, M. L. Mordacchini Alfani,
M. Guerrieri, D. Monarca, M. Cecchini, Acta Hort.,
845, 635 (2009).
7. D. De Santis, A. Fardelli, F. Mencarelli, Acta Hort.,
845, 693 (2009).
8. D. Ghirardello, M. Bertolino, S. Belviso, B. Dal
Bello, M. Giordano, L. Rolle, V. Gerbi, M.
Antonucci, N. Spigolon, G. Zeppa, Postharv. Biol.
Technol., 112, 95 (2016).
9. D. Ghirardello, C. Contessa, N. Valentini, G. Zeppa,
L. Rolle, V. Gerbi, R. Botta, Postharv. Biol.
Technol., 81, 37 (2013).
10. D. Ghirardello, G. Zeppa, L. Rolle, V. Gerbi, C.
Contessa, N. Valentini, R. Botta, Acta Hort., 1052,
305 (2014).
11. D. Ghirardello, L. Rolle, G. Zeppa, Acta Hort., 845,
681 (2009).
12. A. S. Pershern, W. M. Breene, E. C. Lulai, J. Food
Proc. Preserv., 19, 9 (1995).
13. K. S. Ebrahem, D. G. Richardson, R. M. Tetley, Acta
Hort., 351, 677 (1994).
14. M. A. Koyuncu, A. Islam, M. Kucuk, Grasas y
Aceites, 56, 263 (2005).
15. O. Beyhan, N. Yilmaz, S. Bulut, M. Aktas, E. Ozsoy,
Int. J. Agric. Biol., 13, 741 (2011).
16. ISO 659:2009, Oilseeds – Determination of oil
content (Reference method), p. 12 (2009).
17. W. W. Christie, Lipid Analysis, The Oily Press,
Bridgwater, England, 2003.
18. ISO 9936:2006. Determination of tocopherol and
tocotrienol contents by HPLC (2006).
19. ISO 6886:2006. Determination of oxidative stability
(2006).
20. P. L. Benitez-Sánchez, M. León-Camacho, R.
Aparicio, Eur. Food Res. Technol., 218, 13 (2003).
21. C. Crews, P. Hough, J. Godward, P. Brereton, M.
Lees, S. Guiet, W. Winkelmann, J. Agric. Food
Chem., 53, 4843 (2005).
22. G. P. Savage, D. L. McNeil, P. C. Dutta, J. Am. Oil
Chem. Soc., 74, 755 (1997).
23. C. Alasalvar, J. S. Amaral, G. Satir, F. Shahidi, Food
Chem., 113, 919 (2009).
24. S. R. P. Madawala, S. P. Kochhar, P. C. Dutta,
Grasas y Aceites, 63, 143 (2012).
25. S. P. Kochhar, C. J. Henry, Int. J. Food Sci. Nutr.,
60, 289 (2009).
26. H. Ciemniewska-Zytkiewicz, K. Ratusz, J. Brys, M.
Reder, P. Koczon, J. Therm. Anal. Calorim., DOI
10.1007/s10973-014-3861-9 (2014).
S.M. Momchilova et al.: “Fatty acids, tocopherols and oxidative stability of hazelnuts during storage”
70
МАСТНИ КИСЕЛИНИ, ТОКОФЕРОЛИ И ОКИСЛИТЕЛНА СТАБИЛНОСТ НА
ЛЕШНИЦИ ПО ВРЕМЕ НА СЪХРАНЕНИЕТО ИМ
С. М. Момчилова1, С. П. Танева1, М. Д. Златанов2, Г. А. Антова2, М. Й. Ангелова-Ромова2,
Е. Благоева3
1 Институт по органична химия с Център по фитохимия, Българска академия на науките,
ул. „Акад. Г. Бончев“, блок 9, 1113 София, България
2 Катедра „Химична технология“, ПУ „П. Хилендарски“, ул. „Цар Асен“ 24, 4000 Пловдив, България
3 Опитна станция по земеделие, ул. „Миньорска“ 1, 6600 Кърджали, България
Постъпила на 13 ноември 2016 г.; приета на 6 декември 2016 г.
(Резюме)
Лешници от три сорта (Ата Баба, Ран Трапезундски и Тонда Джентиле), отглеждани в България,
са съхранявани за 1, 3, 6 и 12 месеца при различни условия: с черупки или като ядки, при 4oC (в
хладилник) или при 20oC (на сянка), след което са анализирани мастно-киселинния състав,
токоферолите и окислителната стабилност (индукционен период в часове) на маслата им. Резултатите
показват, че мастните киселини практически не се променят до 12 месеца независимо от различните
условия на съхранение, докато количеството на токоферолите постепенно намалява, и тази тенденция
е малко по-силно изразена при 20oC, отколкото при 4oC, както и при лешниците с черупки, в
сравнение с ядките. Подобно съответно намаляване се наблюдава и при индивидуалните α-, β- и
-изомери. Относно окислителната стабилност, маслата от изходните лешници имат дълъг
индукционен период (40, 47 и 58 часа, съответно при сортовете Ата Баба, Ран Трапезундски и Тонда
Джентиле). Тази окислителна стабилност постепенно намалява по време на съхранението по същия
начин, както при токоферолите, т.е. лешниците, съхранявани при 20oC с черупки имат малко по-къс
индукционен период. Всички тези резултати показват, че e възможно лешниците да бъдат
съхранявани до 12 месеца, без да настъпят значителни и вредни за здравето промени в основните им
липидни характеристики. За да се запази качеството им в най-голяма степен, е препоръчително те да
бъдат съхранявани като ядки при 4oC.
Ключови думи: лешници, съхранение, мастни киселини, токофероли, оксидантна стабилност