ARTICLE

pubs.acs.org/JAFC

Chemical Characterization of Sacha Inchi (Plukenetia volubilis L.) Oil

Chiara Fanali,*,† Laura Dugo,† Francesco Cacciola,‡,§ Marco Beccaria,§ Simone Grasso,† Marina Dach
a,†
Paola Dugo,†,§ and Luigi Mondello†,§
†
University Campus Bio-Medico, Via A lvaro del Portillo 21, 00128 Rome, Italy
‡
Chromaleont Srl, A Spin-off of the University of Messina, via Industriale 143, 98124 Messina, Italy
§
Dipartimento Farmaco-chimico, University of Messina, viale Annunziata, 98168 Messina, Italy

ABSTRACT: A chemical characterization of the major components, namely, triacylglycerols (TAGs), polyphenols, and
tocopherols in a Sacha inchi oil derived from cold pressing of the seed, is hereby reported. To tackle such a task, high-
performance liquid chromatography in combination with photodiode array (PDA), fluorescence (RF), and mass spectrometry
(MS) detection was employed. The latter was interfaced with atmospheric pressure chemical ionization and with electrospray
ionization for the analysis of TAGs and polyphenols, respectively, whereas RF detection was tested for the determination of
tocopherol content. Furthermore, fatty acid methyl esters (FAMEs) were evaluated by gas chromatography
flame ionization
detector. A 93% amount of total fatty acids was represented by unsaturated FAMEs with the greatest percentage represented by
linoleic (L) and linolenic (Ln) accounting for approximately 50 and 36%, respectively. The main TAGs (>10%) were represented by
LLnL, LnLnLn, and LnLLn; the latter was present in the oil sample at the highest percentage (22.2%). Among tocopherols, γ-
tocopherol was detected to be the most abundant component (over 50%). The polyphenolic composition was also investigated, and a
total of 15 compounds were positively identified, through the complementary analytical information coming from PDA and MS data.
To the best of our knowledge, this is the first report providing a thorough chemical characterization of a Plukenetia volubilis L. oil.
KEYWORDS: Sacha inchi, triacylglycerols, tocopherols, polyphenols, mass spectrometry

’ INTRODUCTION in the prevention of chronic diseases. The optimal ratio between

Sacha inchi (Plukentia volubilis L.), also known as the Inca
peanut, is a perennial, oleaginous plant of the Euphorbiaceae family,
native to the rain forest of the Andean region of South America.
The plant was known by the natives of the area for thousands of
years as witnessed by several representations reported on vessels
found in Incan tombs.1 The Sacha inchi plant produces star-shaped
green fruits, which yield edible dark brown seeds, slightly enlarged
in the center and squashed toward the edges. The seeds are rich in
oil (35
60%) and proteins (27%) and contain heat-labile sub-
stances with a bitter taste.2 Chancas Indians and other tribal groups
of the region extract oil from the seeds, which is used for the
preparation of various meals. Roasted seeds and cooked leaves are
also an important component of their diets.
To our knowledge, only a few papers have reported the com-
position of Sacha inchi oil and seeds.1,3
7 According to these
works, such an oil is rich in unsaturated fatty acids, about 93% of
the total. In particular, high levels of essential fatty acids were
found, namely, C18:3 ω3 (α-Ln, cis,cis,cis-9,12,15-octadecatrie-
noic acid; α-linolenic) and C18:2 ω6 (L, cis,cis-9,12-octadeca-
dienoic acid; α-linoleic) fatty acids, accounting for approximately
47 and 37%, respectively. Essential fatty acids are intermediate in
the biosynthesis of important compounds in the human body,
such as prostaglandin E1 and its derivates.8,9 Several studies
reported that ω-6 and especially ω-3 unsaturated fatty acids have
beneficial effects on human health by preventing several diseases
like cancer, coronary heart disease, and hypertension; further-
more, a hypocholesterolemic effect was observed when used as
food supplements.10,11 Growing evidence shows how a proper
balance in the diet between ω3 and ω 6 fatty acids plays a key role
linoleic acid and α-linolenic acid in the diet is, in most cases, re-
ported in the literature with values ranging between 4:1 to 5:1,
without exceeding 10:1.12 These values are very often shifted to
the advantage of linoleic acid in the common western diet, re-
aching values as high as 20:1. Such a disproportion is considered
one of the main risk factors leading to the development of chronic
diseases such as obesity, cancer, and cardiovascular disease. ω6
linoleic acid is a precursor of arachidonic acid, which leads to
the production of essential compound such as prostaglandins and
leukotriens, essential for the immune function and platelet aggrega-
tion. However, an excess of arachidonic acid formation could lead
to an abnormal increase of pro-inflammatory mediators, translat-
ing in the development of an inflammatory process. On the other
hand, leukotriens, originating from ω3 fatty acids, show a much
smaller proinflammatory effect. Linoleic and α-linolenic acids
compete for the same enzymes elongase and desaturase for the
syntesis of fatty acids belonging, respectively, to ω6 and ω3 series.
If ω6 consumption in the diet is excessive, a significant inhibition
of the synthesis of EPA and DHA can occur. As a consequence,
great attention has been so far devoted to food supplements con-
taining essential fatty acids and to the proportion between ω3
and ω6 fatty acids.13 The potential use of Sacha inchi products as
ingredients for food supplements is not only related to the fatty
acid profile but also to the amino acid composition of seeds,
Received: August 8, 2011
Accepted: November 4, 2011
Revised: October 24, 2011
Published: November 04, 2011

r 2011 American Chemical Society 13043 dx.doi.org/10.1021/jf203184y | J. Agric. Food Chem. 2011, 59, 13043–13049
Journal of Agricultural and Food Chemistry
showing relatively high levels of cysteine, tyrosine, threonine, and
tryptophan.3 An albumin protein, representing 31% of the total
seed proteins, was also isolated and characterized. It is a glycopro-
tein containing all of the essential amino acids in adequate amounts.14
Moreover, the oil is rich in tocopherols and β-carotene.3,5 For its
composition, especially for the essential fatty acids content, Sacha
inchi oil could be considered in the elaboration of food supplements.
However, very recently, Bueso et al. reported the first case of oc-
cupational allergy induced by P. volubilis seed. This allergy was pro-
bably due to a particular protein allergen not yet identified.15
The aim of this work was to characterize the composition of a
Sacha inchi oil sample by analyzing the main constituents oc-
curring in plant oils: triacylglycerols (TAGs), fatty acid methyl
esters (FAMEs), tocopherols, and polyphenols. TAGs were
analyzed by nonaqueous reversed-phase high-performance liquid
chromatography (NARP HPLC). Detection was performed by
atmospheric pressure chemical ionization (APCI), which is the
most frequently used ionization technique for TAG analysis.16
18
FAMEs were derivatized to methyl esters and analyzed by gas
chromatography
flame ionization detector (GC-FID).19 Toco-
pherols were separated by normal-phase liquid chromatography
(NPLC) coupled to a fluorescence detector (RF). Quantitative
analysis of total phenolic compounds was performed by the
Folin
Ciocalteu method,20 while a polyphenolic fingerprint of
the major polyphenols was attained by HPLC-photodiode array
(PDA)/electrospray ionization (ESI)-MS analysis.
’ MATERIALS AND METHODS
Materials and Reagents. All solvents were of HPLC grade and
were used as received. Methanol (MeOH) and formic acid were pur-
chased from Farmitalia-Carlo Erba (Milan, Italy). Isopropanol (IPA),
acetonitrile (ACN), n-hexane (Hex), acetone, α-tocopherol, γ-toco-
pherol, and δ-tocopherol were provided by Sigma-Aldrich (Milan, Italy).
Samples. Sacha inchi oil sample, guaranteed as nonadulterated, was
obtained by a Peruvian producer.
Fatty Acid Analysis by GC-FID. FAMEs were converted in their
methyl esters by adding 1 mL of sodium methylate in methanol (5% w/v)
to 100 μL of oil and incubating the mixture at 100 °C for 15 min. After the
mixture was cooled, 1 mL of boron trifluoride
methanol (20% BF3)
reagent (Merck, Milan, Italy) was added, and the solution was heated at
100 °C for 15 min. The solution was first cooled and afterward mixed
with 1 mL of n-hexane and 1 mL of water prior to being centrifuged for
2 min at 1254g. The upper layer containing the FAMEs was subsequently
transferred to a 2 mL vial and stored at +4 °C prior to analysis.21
GC-FID analyses were carried out on a GC2010 (Shimadzu, Milan,
Italy) equipped with an AOC-20i autoinjector, a split
splitless injector
(280 °C), and a FID detector. FAMEs separation was performed on a
Supelcowax column (Sigma-Aldrich/Supelco, Bellefonte, PA; 30 m 
0.25 mm i.d.  0.25 μm df).
GC operational parameters were as follows: initial pressure of carrier
gas (He at constant linear velocity, 30 cm/s), 99.5 kPa; temperature pro-
gram 50
280 at 3 °C/min; injection volume, 1 μL; and split ratio, 1:100.
FID parameters were as follows: H2 flow rate, 50 mL/min; air flow rate,
400 mL/min; makeup gas (N2) flow rate, 50 mL/min; sampling fre-
quency, 80 ms; and filter time constant, 200 ms. Data acquisition was
performed using the GCsolution software (version 2.3).
TAG Analysis by HPLC-APCI-MS. HPLC-MS analyses of TAGs
were carried out on a Shimadzu HPLC system (Kyoto, Japan) equipped
with a CBM-20 A controller, two LC-20AB pumps, a DGU-20A5 de-
gasser, a SIL-20AC autosampler, and a LC-MS 2020 mass spectrometer
equipped with an APCI interface (Shimadzu, Kyoto, Japan). As a chroma-
tographic column, an Ascentis Express C18 column (150 mm  4.6 mm,
13044
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2.7 μm d.p.; Supelco) was used. The mobile phase flow rate was 1 mL/min;
ACN (A) and IPA (B) were used, respectively, as the mobile phase in the
following linear gradient mode: 0 min, 0% B; 50 min, 70% B; 55 min, 70%
B; and 56 min, 0% B. The injection volume was 2 μL, and analyses were run
at room temperature.
Applied were the following APCI-MS parameters: APCI mode pos-
itive; mass spectral range, 250
1200 m/z; interval, 0.5 s; scan speed,
2143 amu/s; nebulizing gas (N2) flow, 4.0 mL/min; APCI temperature,
400 °C; heat block, 230 °C; desolvation line (DL) temperature, 250 °C;
DL voltage,
34 V; probe voltage, +4.5 kV; Qarray voltage, 1.0 V; RF
voltage, 90 V; and detection gain, 1.05 kV. Data acquisition was per-
formed using the LabSolution software (version 5.10.153).
Tocopherol Analysis by NPLC-RF. The analyses of tocopherols
were carried out by using a Shimadzu HPLC system (Kyoto, Japan)
equipped with a LC 10 AD Vp high pressure isocratic pump, an SCL-
10A Vp controller, and an RF-10 AXL fluorescence detector (program-
med for excitation at 290 nm and emission at 330 nm). Data acquisition
was performed using the LCsolution software (ver. 1.12).
All of the analyses were performed at ambient temperature (25 °C)
using a microsilica column (Supelco Ascentis SI 250 mm  1.0 mm,
5 μm particle size). The mobile phase was a mixture of Hex:IPA (99:1),
the flow rate was 0.050 mL/min, while the injection volume was 2 μL.
The sample was prepared by dissolving 0.01 g of oil in 1 mL of n-hexane. The
method was validated following the EURACHEM guideline for each com-
ponent, namely, α-tocopherol, γ-tocopherol, and δ-tocopherol.22
Linearity was tested at five different concentrations for each analyte,
performing three replicates per level. Regression lines were built using
the square method. The linearity and the goodness of the curves used
were confirmed using Mandel's23 fitting tests. The significance of the
intercept was established running a t test (significant level, 5%).
Accuracy, in terms of trueness and precision, was assessed at two
different levels, repeating the analysis five times. The Shapiro
Wilk test, to
check the normality of the distribution, and the Dixon and Grubbs tests, to
verify the presence of outliers, were performed before calculating precision
in terms of coefficient of variation (CV %) and limit of repeatability (r).
The limit of detection (LOD) and limit of quantification (LOQ) were
calculated performing 10 times the analysis of a blank sample and applying
the following formulas:
LOD : yd ¼ μb þ 2t  σb
LOQ : yq ¼ μb þ 10  σb
where yd and yq is the signal at the LOD and LOQ, respectively, μb is the
average signal of the blank sample, σb is the blank standard deviation, and
t is the constant of the t Student distribution depending on the confidence
level (95%) and degrees of freedom. LOD and LOQ values were, finally,
obtained by plotting yd and yq in the calibration line.
Total Phenolic Compounds Quantitative Analysis. The total
soluble oil phenolics analysis was performed by using Folin
Ciocalteu
reagent, according to the procedure of Montedoro et al.20 Briefly, the
methanolic extract of Sacha inchi oil was obtained as follows: 10 g of oil
were dissolved in a solution of Tween 20 (2% v/w), added to 10 mL of a
methanol/water (80:20 v/v) solution, and mixed with an Ultra-Turrax
T18 homogenizer (model B003UTSVCW, IKA WERKE GmbH & Co.
KG, Staufen im Breisgau, Germany) at 24000 rpm. The mixture was
centrifuged at 5000g for 10 min, and the methanolic phase was re-
covered. The extraction procedure was repeated two times; the two
methanolic fractions were pooled and stored at
20 °C to eliminate
residual oil droplets. One milliliter of Sacha inchi oil extract was mixed
with 70 mL of water and 5 mL of Folin
Ciocalteu reagent. Five minutes
later, 15 mL of sodium carbonate solution (25% w/v) was added; the
mixture was vortexed and diluted with water to a final volume of 100 mL.
The mixture was then incubated for 2 h at room temperature after which
dx.doi.org/10.1021/jf203184y |J. Agric. Food Chem. 2011, 59, 13043–13049
Journal of Agricultural and Food Chemistry ARTICLE

the absorbance at 765 nm was measured; results were expressed as gallic
acid equivalents (GAE) in milligrams per 100 g of oil (mg GA/100 g
oil). The concentration of polyphenols in sample was derived from a
standard curve of absorbance of gallic acid concentrations ranging from
50 to 500 μg/mL.
Polyphenol Analysis by HPLC-PDA/ESI-MS. Polyphenols were
extracted from Sacha inchi oil by using a slightly modified procedure already
reported in the literature.20 Briefly, 50 g of oil was extracted three times with
10 mL each of a methanol/water (80:20, v/v) mixture. The three fractions
were pooled, and the solution was centrifuged at 5000g for 10 min. After-
ward, the surnatant was recovered, and the solvent was completely removed
in vacuo by using a Speed Vac Plus concentrator (model SC110A, Savant
Instrument, Holbrook, NY). The obtained residue was suspended in 5 mL of
acetonitrile and washed with 10 mL of hexane to remove lipid components.
The hexane extract was discarded, and the ACN solution was concentrated
under vacuum. The dried residue, containing the polyphenols, was sus-
pended in 200 μL of a mixture methanol/water (50:50 v/v).
HPLC-PDA/ESI-MS analyses of polyphenols were performed on a
Prominence LC system (Shimadzu, Milan, Italy) equipped with a CBM
20A controller, two LC-20AD pumps, a CTO-20AC column oven, a
DGU-20A3 degasser, a SIL-20AC autosampler, a SPD-20AD, and
LCMS-2020 mass spectrometer equipped with an ESI interface. Data
acquisition was performed by the LabSolution software (Shimadzu,
Version 5.10.346).
Polyphenols were separated on a Supelco Ascentis Express C18 col-
umn (150 mm  4.6 mm, 2.7 μm particle size). A gradient employing
water (pH = 3; solvent A) and acetonitrile (solvent B) was employed
under the following conditions: 0
60 min, 0
100% B; 50
55 min, 100%
B. The column was reconditioned with the initial eluent composition in
5 min. The mobile phase flow rate was 1 mL/min (split to 0.2 mL/min prior
to MS detection), the column temperature was set at 25 °C, while the
injection volume was 2 μL. The wavelength range was 210
400 nm, and
the chromatograms were extracted at 280 nm. The sampling frequency
was 6.25 Hz, while the time constant was 0.16 s. ESI-MS acquisition was
performed in negative mode under the following conditions: mass
spectral range, 100
700 m/z; interval, 0.5 s; scan speed, 1250 amu/s;
nebulizing gas (N2) flow, 1.5 L/min; heat block, 300 °C; DL tempera-
ture, 300 °C; DL voltage,
34 V; probe voltage, 4.5 kV; Qarray voltage,
1.0 V; RF voltage, 60 V; and detection gain, 1.0 kV.
’ RESULTS AND DISCUSSION
FAMEs Composition in a Sacha Inchi Oil Sample by GC-
FID. Evaluation of Sacha inchi fatty acids profile was performed
by GC-FID analysis of FAMEs. Peak identification was obtained
by means of coinjection of pure standard FAMEs; the determi-
nation of the distribution of single FAMEs was expressed as a
mass fraction of the total FAME content (%), calculated from
Ax =ATOT  100
where Ax refers to the peak area of the FAME considered and
ATOT refers to the total peak area of the FAMEs contained in the
sample. The names of the identified FAMEs along with the
relative percentage are reported in Table 1.
These results show that approximately 93% of total fatty acids
is represented by unsaturated FAMEs, the greatest percentage is
for α-Ln (ω3) and L (ω6) acids, approximately 50 and 36%, re-
spectively (Table 1). Our results are in good agreement with
those reported by other groups.1,3
5 Because of the high content
Table 2. Fatty Acid Composition of Different Edible
Vegetable Oils

Table 1. FAMEs Identified in Sacha Inchi Oil Sample by GC- fatty acid %
FID Analysis Together with Their Peak Area Ratio Percentage
ref vegetable oil 8:0 10:0 12:0 14:0 16:0 18:0 18:1 18:2 18:3 20:1
FAMEs name symbol CN:DBa peak area ratio %
24 palm oil 5 36 2 50 8
palmitic P 16:0 4.3 24 sunflower oil 7 5 19 68
stearic S 18:0 3.0 24 soybean oil 11 4 23 54 8
vaccenic V Δ11
18:1 0.6 24 linseed oil 6 2 19 24 47
oleic O Δ9
18:1c 9.0 24 canola oil 4 2 60 21 10 1
linoleic L Δ9,12
18:2 36.2 24 coconut oil 7 7 48 18 9 3 6 2
linolenic Ln Δ9,12,15
18:3 46.8 25 peanut oil 9 3 52 32
a
CN, carbon number; DB, double bond. Sacha inchi oil 4 3 9 36 47

Figure 1. HPLC-APCI-MS total ion current chromatogram of TAGs in a Sacha inchi oil sample.

13045 dx.doi.org/10.1021/jf203184y |J. Agric. Food Chem. 2011, 59, 13043–13049

Journal of Agricultural and Food Chemistry ARTICLE

Table 3. TAGs Identified in the Sacha Inchi Oil Sample by composition of some common vegetable edible oils as compared
Means of HPLC-APCI-MSa to that of Sacha inchi oil.24,25 Among plant oils reported in
Table 2, only linseed oil contains a percentage of linolenic acid
TAG MW DB PN [M + H]+ DG+ DG+ DG+ area %
comparable to that of Sacha inchi oil, together with low percen-
1 LnLnLn 9 36 874 LnLn 12.3 tages of saturated fatty acids such as palmitic and stearic. Among
873 596 edible plant oils, linseed oil, because of its predominant composi-
2 LnLLn 8 38 876 LnL LnLn 22.2 tion in ω3 fatty acids, has been widely employed as a food sup-
875 598 596
plement showing a particularly favorable nutritional profile.26 It is
also worth noting that the balance between ω3, ω6, and ω9 fatty
3 LLnL 7 40 878 LnL LL 18.2
acids is an important nutritional property of plant oils. For this
877 598 600
reason, plant oils that are rich in polyunsaturated fatty acids could
4 LnOLn 7 40 878 OLn LnLn 7.3 be of great interest for a balanced diet and represent a good alter-
877 600 596 native to fish-based foods and supplements that so far represent
5 PLnLn 6 40 852 LnLn PLn 3.2 the main food sources of ω3 fatty acids.
851 596 574 Triacilglycerol (TAG) Composition in a Sacha Inchi Oil
6 LLL 6 42 880 LL 5.3 Sample by HPLC-APCI-MS. The fatty acid composition can be
879 600 used to evaluate the stability and nutritional quality of fats and
7 OLLn 6 42 880 OLn LLn OL 9.3 oils. However, to understand their physical and functional pro-
879 600 598 602 perties, the determination of the types and amounts of TAG
8 PLLn 5 42 854 PLn LP 4.4
species present in a lipidic sample is also essential. Figure 1 shows
the total ion current (TIC) chromatogram of a Sacha inchi oil
853 574 576
sample. As reported in Table 3, 21 different TAGs were detected
9 SLnLn 6 42 880 LnLn SLn 2.1
and identified; for each TAG, the relative abundance was calc-
879 596 602 ulated as the ratio of the TIC area of the peak with respect to the
10 LOL 5 44 882 LL LO 3.0 sum of TIC areas of all identified TAGs. The latter were identified
881 600 602 according to their HPLC-APCI-MS mass spectra considering m/z
11 OLnO 5 44 882 OLn OO 1.8 values of pseudomolecular (M + H)+ and “diacylglycerol” fragment
881 600 604 (M + H
RCOOH)+ ions. As can be seen from Table 3, some
12 LLP 4 44 856 LP LL 6.9 coelutions were observed (12, LLP + SLLn + POLn; 14, SLL +
855 576 600 OLP + SOLn). These results may be explained considering that
SLLn 5 44 882 SL LLn SLn TAG retention times in NARP-HPLC increase with increasing
881 604 598 602
partition number (PN); the latter is defined as the total carbon
number (CN) in all acyl chains minus two times the number of
POLn 4 44 856 OLn PLn PO
double bonds (DBs), viz. PN = CN
2DB. As a consequence,
855 600 574 578
TAGs with the same PN are, usually, very difficult to resolve.
13 OLO 4 46 884 OO OL 0.8 In addition, the retention behavior of TAGs with the same CN is
883 604 602 strongly influenced by the FAs composition of the individual
14 SLL 4 46 884 LL SL 2.7 TAG, mainly by the unsaturation degree and acyl chain length.16
883 600 604 In agreement with the GC-FID analysis, identified TAGs con-
OLP 3 46 858 PL PO LO tained five different FAs (P, L, Ln, O, and S). The predominant
857 576 578 602 components (>50%) were detected at m/z 873, 875, and 877 and
SOLn 4 46 884 OLn SLn SO were identified as dilinolenoyl-linoleoylglycerol (LnLLn), dili-
883 600 602 606 noleoyl-linolenoylglycerol (LLnL), and trilinolenin (LnLnLn).
15 OOO 3 48 886 OO 0.1
Most of the TAGs in Sacha inchi oil (>80%) contained at least
one residue of linolenic acid. The high percentage of linolenic
885 604
acid in TAGs is very important because unsaturated fatty acids
16 SLO 3 48 886 LO SL SO 0.4
deriving primarily from vegetable oils reduce the levels of total
885 602 604 606 and low density lipoprotein cholesterol, holding promise against
SLP 2 48 860 SL LP SP the risk of cardiovascular diseases.27
859 604 576 580 Moreover, the position of fatty acid in the glycerol skeleton is
a
PNs, DBs, pseudomolecular ([M + H]+), and diglyceride ions (DG+ important from a nutritional and metabolic point of view. The
[M + H
RCOOH]+) corresponding to each identified TAG along stereospecifical position of fatty acids plays a key role in tryglicerol
with the relative concentration expressed as percentage area (area %) are absorption and digestion because FAs at the sn-1 and sn-3 po-
reported. sitions are digested first by lipases, yielding sn-2 monoacylglycerols
and free FAs.28 To our knowledge, this work reports for the first time
of polyunsaturated essential fatty acids, especially ω3 and ω6, a thorough characterization of the TAG fraction in Sacha inchi oil.
Sacha inchi oil is increasingly advertised as a nutritional supple- Tochopherol Composition in a Sacha Inchi Oil Sample by
ment. In fact, it provides more useful nutraceuticals than other NPLC-RF. Tochopherols were analyzed by NPLC coupled with a
common seed oils that contain only remarkable quantities of ω6 fluorimetric detector. Peak identification was obtained by com-
fatty acids. Specifically, edible plant oils contain different per- paring the retention time of the sample with a standard solution
centages of the five common, nutritionally important free fatty (Table 4). The method was validated in terms of linearity, LOD,
acids (FAs) (P, S, O, L, and α-Ln). Table 2 shows the fatty acid LOQ , and accuracy (precision and trueness). Mandel's fitting
13046 dx.doi.org/10.1021/jf203184y |J. Agric. Food Chem. 2011, 59, 13043–13049
Journal of Agricultural and Food Chemistry
Figure 2. HPLC-PDA chromatogram (extracted at 280 nm) of the polyphenolic profile of a Sacha inchi oil sample.
Table 4. Linearity, LOD, LOQ , Accuracy, and Concentration Values for α-, γ-, and δ-Tocopherols Contained in the Sacha Inchi
Oil Sample Tested
μg/kg
linear range tocopherol
compds (mg/kg) content (g/kg) LOD LOQ
α-tocopherol 0.026
13 0.004 0.047 0.053
γ-tocopherol 0.048
24 1.257 0.098 0.104
δ-tocopherol 0.032
16 0.869 0.101 0.107
test was widely satisfactory (Fcalc < Ftab), and R2 was 0.996 for α-,
γ-, and δ-tocopherol, respectively. LOD and LOQ were as
follows: α-tocopherol, 4.7 and 5.3 μg/kg; γ-tocopherol, 9.8 and
10.4 μg/kg; and δ-tocopherol, 10.1 and 10.7 μg/kg, respectively,
equivalent to 0.047 and 0.053 μg/kg of oil for α-tocopherol,
0.098 and 0.104 μg/kg of oil for γ-tocopherol, and 0.101 and
0.107 μg/kg of oil, respectively (Table 4). No outliers were
highlighted by Dixon and Grubbs tests. At both concentrations
assessed, intraday precision results were satisfactory, less than
10% at the lower level and less than 5% at the higher level, for all
considered compounds. Trueness results were lower than 15 and
0.35% in terms of relative error, at the lower and higher levels,
respectively. γ-Tocopherol accounted for more than 50% of the
entire tocopherol content.
α-Tocopherol is considered the most representative antiox-
idant in olive oil, while γ- and δ-tocopherols are found principally
in seed oils like soybean and sunflower, and in particular, a large
amount of γ-tocopherol is present in soybean oil. It has been
reported that the antioxidant activity decreases in the order γ > δ >
β > α tocopherol. As already stated, the greater amounts of γ-
and δ-tocopherols with respect to α could be attributed to their
greater antioxidant capability. This makes the oil more stable to
oxidation.4 From a nutritional point of view, intestinal absorption
of γ-tocopherol is similar to that of α-tocopherol, and it could
play a specific role in preventive side effects of some radicals like
peroxynitrite and NOx.29
Polyphenolic Composition in a Sacha Inchi Oil Sample by
RPLC-ESI-MS. The total phenolic content of Sacha inchi oil was
determined by the Folin
Ciocalteu technique.22 The concen-
tration of phenolics was 6.20 mg/100 g of oil expressed as GAEs.
Thanks to its antioxidant properties, the polyphenolic content
13047
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min level (n = 5) max level (n = 5)
precision trueness precision trueness
(CV %) (relative error %) (CV %) (relative error %)
9.42
14.47 4.67 0.30
7.03
7.17 4.19 0.28
6.67
5.07 4.95 0.31
ensures the oxidative stability of polyunsaturated fatty acids
imparting the characteristic flavor to the oil. Moreover, the effect
of many of them on some common diseases such as hypertension,
atherosclerosis, prevention of certain cancers, and modification of
immune and inflammatory responses is notorious.30 Among edible
oils, olive oil has been extensively characterized, and several studies
have been published on the quantitative analysis of total phenolics
and on their qualitative characterization in extra virgin olive oil
(EVOO).31
33 On the other hand, no data on the polyphenolic
fraction of Sacha inchi have been reported so far. A qualitative char-
acterization of Sacha inchi oil phenolics was performed by RPLC-
PDA/ESI-MS (Figure 2). The partially porous C18 column
allowed good baseline separation for most compounds to be
attained within 50 min of analysis time under gradient condi-
tions. The advantages of shell-packed stationary phases have
previously demonstrated by a number of studies for high-
efficient separations of complex samples.34
38 Polyphenols
were identified through the complementary information com-
ing from PDA and MS spectra along with data already reported
in the literature (Table 5).31
33 Twenty-one phenolic com-
pounds were detected; among them, a number of 15, belonging
to phenyl alcohol, flavonoid, seicoridoid, and lignan classes,
were positively identified.
Concluding, in this study, a characterization of the composi-
tion of a Sacha inchi oil sample through the analysis of the main
constituents, namely, TAGs, FAs, tochopherols, and polyphenols,
has been carried out for the first time. Apart from the FAs profile,
the elucidation of TAG and polyphenol content has been reported
for the first time. LnLLn was detected as the major compound,
whereas among tocopherols, γ-tocopherol turned out to be
the most abundant. Finally, the polyphenolic composition was
dx.doi.org/10.1021/jf203184y |J. Agric. Food Chem. 2011, 59, 13043–13049
Journal of Agricultural and Food Chemistry ARTICLE

Table 5. Polyphenols Identified in a Sacha Inchi Oil Sample by HPLC-PDA/ESI-MS

molecular [M
H]
λmax identification
peak tR (min) formula (fragment ions) UV/vis (nm) compd compd class [ref]

1 10.3 C8H10O3 153 271 hydroxytyrosol phenyl alcohol UV, MS [31, 32]
2 19.4 C8H10O2 137 270 tyrosol phenyl alcohol UV, MS [31, 32]
3 20.0 C14H16O9 327 (311) 278, 325 bergenin isocoumarin UV, MS
4 26.9 C26H25O4 401 (369) 268, 350 alpinumisoflavone flavonoid UV, MS [31]
5 27.5 C21H24O10 435 278, 350 phloretin-glucoside flavonoid UV, MS
6 28.3 C18H21O6 333 270, 309 methyl decarboxymethyl oleuropein aglycon secoiridoid UV, MS [32]
7 29.5 C22H21O9 357 272 pinoresinol lignan UV, MS [31, 32]
8 31.6 C18H21O6 315 369 isorhamnetin-glucoside flavonoid UV, MS
9 31.8 C10H15O3 183 270, 309 oleuropeic acid secoiridoid UV, MS [31]
10 33.1 C9H15O4 187 260 azaleic acid organic acid UV, MS [31]
11 34.6 C15H9O6 285 272, 346 luteolin flavonoid UV, MS [32]
12 36.5 596 (586, 550) 278 unknown
13 37.4 541 278 unknown
14 39.2 479 (431) 278 unknown
15 39.5 479 (431) 275 unknown
16 39.9 479 (431) 278 unknown
17 40.3 C22H26O8 417 275 syringaresinol lignan UV, MS [31, 32]
18 40.6 334 275
19 41.9 C20H22O7 373 272 hydroxy-pinoresinol lignan UV, MS [33]
20 44.6 C15H10O5 269 268, 339 apigenin flavonoid UV, MS [32]
21 45.2 C19H21O8 377 270, 309 oleuropein aglycon secoiridoid UV, MS [31 32]

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The present study demonstrates that Sacha inchi oil is a good
source of linoleic and linolenic acids with considerable amounts
of tocopherols. In addition, the flavonoid composition in the
extracted oil suggests its potential use as dietary source of natural
antioxidants.
’ AUTHOR INFORMATION
Corresponding Author
*Tel: +39-06-225419123. Fax: +39-06-225411972. E-mail: c.fanali@
unicampus.it.
’ ACKNOWLEDGMENT
We gratefully acknowledge Shimadzu and Sigma-Aldrich/
Supelco Corporations for their continuous support.
’ ABBREVIATIONS USED
P, palmitic; L, linoleic; Ln, linolenic; O, oleic; S, stearic; V, vacce-
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liquid chromatography; GC-FID, gas chromatography
flame
ionization detector; LOD, limit of detection; LOQ, limit of quan-
tification; PN, partition number; CN, total carbon number; DBs,
double bonds; GAE, gallic acid equivalents
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