Academic Edi o : Gian anco Picone
Recei ed: 31 Ma ch 2025
Re ised: 25 Ap il 2025
Accep ed: 27 Ap il 2025
Published: 30 Ap il 2025
Ci a ion: Goicoechea-Oses, E. 1H
NMR S udy o he Lipid Composi ion,
Oxida i e and Hyd oly ic S a us o he
Co e ing Oils o Canned Sa dines
A e Long-Te m S o age. Foods 2025,
14, 1589. h ps://doi.o g/10.3390/
oods14091589
Copy igh : © 2025 by he au ho .
Licensee MDPI, Basel, Swi ze land.
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A icle
1H NMR S udy o he Lipid Composi ion, Oxida i e and
Hyd oly ic S a us o he Co e ing Oils o Canned Sa dines A e
Long-Te m S o age
Enca nacion Goicoechea-Oses
Food Technology, Facul y o Pha macy, Lasca ay Resea ch Cen e , Uni e si y o he Basque Coun y (UPV/EHU),
01006 Vi o ia-Gas eiz, Spain; [email p o ec ed]; Tel.: +34-945013083
Abs ac : The co e ing oils o wen y- wo comme cially canned sa dines we e s udied
by P o on Nuclea Magne ic Resonance spec oscopy (
1
H NMR) eshly pu chased and
also a e s o age a oom empe a u e o i een yea s. The illing oils s udied we e
oli e oils (one ex a- i gin oli e oil), sun lowe oils, soybean oils, and ege able oils
(unspeci ied o igin). The aim was o ob ain quali a i e and quan i a i e in o ma ion on lipid
composi ion, oxida i e and hyd oly ic s a us, and on he changes occu ing du ing s o age.
Jus a e pu chase, in all he samples, he mig a ion o ish omega-3 polyunsa u a ed
eicosapen aenoic (EPA, C20:5
ω
3) and docosahexaenoic (DHA, C22:6
ω
3) acyl g oups was
epo ed; he occu ence o oxida i e o hyd oly ic eac ions was no obse ed. A e
s o age, he main change in he spec a was he p esence o signals due o hyd oly ic
compounds (mainly 1,3-diglyce ides, oge he wi h 1,2-diglyce ides, 1-monoglyce ides,
and lowe p opo ions o 2-monoglyce ides). In ele en samples e y low concen a ions o
sa u a ed aldehydes (alkanals) we e de ec ed, which is conside ed a low oxida i e s a us.
I is sugges ed ha he abo e-men ioned pa ial glyce ides and alkanals mig a ed om
sa dine muscle o he oils. The con en in omega-3 lipids in he oils a e s o age indica ed
he occu ence o lipid in e change be ween he sa dine muscle and he packing oil in
bo h di ec ions.
Keywo ds: co e ing liquid; ege able oil; canned sa dines; inned ish; p o on nuclea
magne ic esonance spec oscopy (1H NMR)
1. In oduc ion
Fish is known o p o ide high con en s o impo an mac o- and mic onu ien s o he
human die , such as omega-3 polyunsa u a ed lipids (
ω
3), especially eicosapen aenoic (EPA,
C20:5
ω
3) and docosahexaenoic (DHA, C22:6
ω
3) acyl g oups, essen ial aminoacids and
p o eins, liposoluble i amins, and mic oelemen s, among o he s. In ecen yea s, special
a en ion has been paid o he po en ial o o age ish, as his a o dable and abundan
sea ood ca ego y is conside ed as he mos nu i ious ish species wi h he lowes ca bon
oo p in [
1
]. A species o o age ish widely consumed in some Eu opean coun ies a e
sa dines, no only esh, bu also as canned p oduc s. Canning is applied o p olong he
shel li e o his pe ishable ood p oduc . In his p ocess, b ie ly, sa dines a e p ecooked,
packed wi h a illing medium, sealed in a he me ic con aine , and subjec ed o he mal
ea men (s e iliza ion a 110–130
◦
C o 25–120 min) o achie e comme cial s e ili y [
2
].
As a esul o his se e e hea ea men , ish enzymes and mic oo ganisms a e inac i a ed.
A e wa ds, he ma u a ion p ocess s a s and con inues du ing s o age un il he cans
a e consumed.
Foods 2025,14, 1589 h ps://doi.o g/10.3390/ oods14091589
Foods 2025,14, 1589 2 o 14
In canned sa dines, he illing medium is gene ally a ege able oil, bu o he kinds
o co e ing medium a e also possible, like oma o sauce o ma inade [
3
]. The quali y
and na u e o he oils employed a e o g ea impo ance as hey can a ec no only he
nu i ional and senso y alue o sa dines bu also hei p ice and shel li e. I mus be
no ed ha al hough he e a e se e al s udies on he nu i ional and quali y con ol o
canned ish p oduc s, he e a e no so many on ha o hei co e ing oils [
4
–
7
]. Some
s udies ocused on he assessmen o he genuineness o he oil used as liquid medium,
bu his is a di icul ask due o he lipid in e change ha occu s be ween he co e ing oil
and he ish [
8
]. Mo eo e , jus h ee s udies ha e ocused on he p esence o oxida i e
and hyd oly ic compounds in he co e ing oils o canned ish [
9
–
11
], which is a subjec
o g ea in e es due o i s po en ial implica ions o consume s’ heal h. In his ega d
i mus be no ed ha in ecen yea s, special a en ion has been paid o he addi ion o
po en ial na u al an ioxidan s o he packing medium o canned sa dines [
12
] and o he
ish species [
13
,
14
] o enhance lipid s abili y. Rega ding he me hodology employed o
s udy lipid main componen s, in mos o hese p e ious wo ks, de i a iza ion o he a y
acyl chains o he co esponding me hyl es e s (FAMEs) ollowed by gas ch oma og aphic
(GC) me hods was used. This is qui e labo ious, and in ol es a complex se ies o chemical
manipula ions ha , oge he wi h he high empe a u es o GC injec o and o en, may gi e
ise o he a i ac ual oxida ion o he compounds subjec s o s udy [15].
In ecen decades, in e es has eme ged in he use o spec oscopic echniques, such as
P o on Nuclea Magne ic Resonance spec oscopy (
1
H NMR), o s udy ood lipid samples
as a whole, as his echnique can p o ide quali a i e and quan i a i e in o ma ion on lipid
componen s and on he occu ence o deg ada i e eac ions, like oxida ion and hyd olysis,
in a single un o a ew minu es and wi hou any p e ious sample modi ica ion [16–19].
In his con ex , he aim o his wo k is o s udy, by
1
H NMR, he quali y and sa e y
o he co e ing oils o comme cial canned sa dines, no only eshly pu chased bu also
a e being s o ed o i een yea s a oom empe a u e. Special a en ion will be paid o
he po en ial changes in he composi ion in main acyl g oups o he packing oils, and also
o he p esence o oxida i e o hyd oly ic compounds. This is he i s ime ha his kind o
s udy has been ca ied ou o such a long- e m s o age pe iod.
2. Ma e ials and Me hods
2.1. Samples
Twen y- wo comme cially canned sa dines (Sa dina pilcha dus, also called Eu opean
sa dine) o di e en b ands we e acqui ed in local supe ma ke s ( ou cans o each sam-
ple) [
3
]. These wen y- wo samples we e selec ed because hey ep esen ed all he di e en
canned sa dines ha we e ma ke ed a ha ime (2010) in local supe ma ke s in Vi o ia-
Gas eiz (Spain). Acco ding o he labelling, ou kinds o oils we e employed as co e ing
mediums: oli e oil (OO) in se en samples (OO5 sample con ained ex a- i gin oli e oil,
bu he o he s we e made o e ined oli e oil blended wi h i gin oli e oil); sun lowe oil
(SFO) in six samples; soybean oil (SYO) in ou samples; and ege able oil o unspeci ied
o igin (VO) in i e samples. I was only possible o ind one sample o sa dines canned
in ex a- i gin oli e oil; p obably due o i s high p ice, his highe quali y oil is used in
mo e expensi e canned ish, such as una [
11
]. I mus be no ed ha in mos o he samples,
he co e ing medium was he oil, bu in h ee o hem, in addi ion o oil, hey also con-
ained many o he ing edien s like inega o ege ables; o poin ou his ac , he name o
hese samples was ma ked wi h an as e isk: SFO6*, SYO3*, and SYO4*. The in o ma ion
p o ided by he p oduce s in he labelling is shown in Supplemen a y Table S1. Jus a e
acqui ing he cans in 2010, wo cans o each sample we e opened and he co e ing oil
Foods 2025,14, 1589 3 o 14
was sepa a ed by decan a ion, ollowed by il a ion h ough anhyd ous sodium sulpha e
(Pan eac, Ba celona, Spain). Samples we e ozen a −20 ◦C un il hei s udy by 1H NMR.
2.2. S o age
Remaining canned sa dines we e kep closed in a cupboa d a oom empe a u e in
he da k o i een yea s. A e wa ds, wo cans o each sample we e opened, and hei
co e ing oil was s udied as desc ibed abo e. The samples we e named simila ly bu wi h
he p e ix ‘ST_’.
2.3. 1H Nuclea Magne ic Resonance Spec a Acquisi ion and De i ed Da a
The
1
H NMR spec a o he samples we e acqui ed in duplica e using a B uke A ance
400 spec ome e ope a ing a 400 MHz (B uke Scien i ic Ins umen s, Bille ica, MA, USA).
Fo sample p epa a ion, 200
µ
L o oil was mixed in a 5 mm diame e
1
H NMR ube wi h
400
µ
L o deu e a ed chlo o o m (CDCl
3
, 99.8%), con aining a small p opo ion o non-
deu e a ed chlo o o m and 0.03% o e ame hylsilane (TMS). This was used as e e ence
compound o calib a e he chemical shi a 0.0 ppm. NMR deu e a ed sol en was acqui ed
om Eu iso op (Sain -Aubin, F ance). The acquisi ion pa ame e s we e as ollows: spec al
wid h 6250 Hz, elaxa ion delay 3 s, numbe o scans 64, acquisi ion ime 2.621 s, and
pulse wid h 90
◦
, wi h a o al acquisi ion ime o 6 min and 20 s. The assignmen o he
1
H NMR signals (based on hei chemical shi s and mul iplici ies) was pe o med using
s anda ds and/o he li e a u e, as p e iously desc ibed, and is p o ided in Table 1[
16
–
21
].
The elaxa ion delay and acquisi ion ime used o ob ain he spec a ensu ed comple e
elaxa ion o he sample p o ons, allowing he signal a ea o be p opo ional o he numbe
o p o ons gene a ing hem. This enabled he use o
1
H NMR signals o es ima e he
mola pe cen age o he se e al kinds o acyl g oups, glyce ides, and oxida ion p oduc s
p esen in he samples, as desc ibed in he Supplemen a y Ma e ial. The
1
H NMR spec a
illus a ing he wo igu es we e plo ed a a ixed absolu e in ensi y alue o ensu e alidi y
o compa a i e pu poses and p ocessed using he MNo a p og am (Mes elab Resea ch,
San iago de Compos ela, Spain).
Table 1. Chemical shi assignmen s and mul iplici ies o he
1
H NMR signals in CDCl
3
o he main
p o ons o glyce ides and a y acids, and o ce ain oxida ion compounds p esen in he co e ing oils
o canned sa dines be o e and a e i een-yea s o age, in ag eemen wi h p e ious s udies [
16
–
21
].
The signal le e s ag ee wi h hose gi en in Figu es 1 and 2.
Signal Chemical
Shi (ppm) Mul i-Plici y Func ional G oup
Type o P o ons Compound
Main Acyl G oups (AG) and Fa y Acids (FA)
A0.88 -CH3sa u a ed, monounsa u a ed ω-9
and/o ω-7 AG and FA
0.89 -CH3unsa u a ed ω-6 AG and FA
B0.97 -CH3unsa u a ed ω-3 AG and FA
C1.19–1.42 m * -(CH2)n-AG and FA
D1
1.61 m -OCO-CH2-CH2-AG in TG, excep o DHA and EPA AG
1.62 m -OCO-CH2-CH2-AG in 1,2-DG, excep o DHA and EPA AG
1.63 m -OCO-CH2-CH2-,
COOH-CH2-CH2-
AG in 1,3-DG,1-MG and FA, excep o DHA
and EPA AG/FA
1.64 m -OCO-CH2-CH2-AG in 2-MG, excep o DHA and EPA AG
D2 1.69 m -OCO-CH2-CH2- EPA AG in TG
1.72 m COOH-CH2-CH2- EPA FA
E1.92–2.15 m ** -CH2-CH=CH- AG and FA, excep o -CH2- o DHA AG/FA in
β-posi ion in ela ion o ca bonyl g oup
Foods 2025,14, 1589 4 o 14
Table 1. Con .
Signal Chemical
Shi (ppm) Mul i-Plici y Func ional G oup
Type o P o ons Compound
Main Acyl G oups (AG) and Fa y Acids (FA)
F1
2.26–2.36 d -OCO-CH2-AG in TG, excep o DHA AG
2.33 m -OCO-CH2-AG in 1,2-DG, excep o DHA AG
2.35 -OCO-CH2-
COOH-CH2-
AG in 1,3-DG,1-MG and FA, excep o DHA
AG/FA
2.38 -OCO-CH2-AG in 2-MG, excep o DHA AG
F2 2.37–2.41 m -OCO-CH2-CH2- DHA AG in TG
2.39–2.44 m COOH-CH2-CH2- DHA FA
G2.77 =HC-CH2-CH= diunsa u a ed ω-6 AG and FA
H2.77–2.90 m =HC-CH2-CH= polyunsa u a ed ω-3 (and ω-6) AG and FA
I3.65 ddd ROCH2-CHOH-CH2OH glyce yl g oup in 1-MG
J3.73 m *** ROCH2-CH(OR′)-CH2OH glyce yl g oup in 1,2-DG
K3.84 m *** HOCH2-CH(OR)-CH2OH glyce yl g oup in 2-MG
L3.94 m ROCH2-CHOH-CH2OH glyce yl g oup in 1-MG
M4.05–4.21 m ROCH2-CHOH-CH2OR′glyce yl g oup in 1,3-DG
N4.18 ddd ROCH2-CHOH-CH2OH glyce yl g oup in 1-MG
O4.22 dd,dd ROCH2-CH(OR′)-CH2OR′′ glyce yl g oup in TG
P4.28 ddd ROCH2-CH(OR′)-CH2OH glyce yl g oup in 1,2-DG
Q4.93 m HOCH2-CH(OR)-CH2OH glyce yl g oup in 2-MG
R5.08 m ROCH2-CH(OR′)-CH2OH glyce yl g oup in 1,2-DG
S5.27 m ROCH2-CH(OR′)-CH2OR′glyce yl g oup in TG
T5.28–5.7 m -CH=CH-AG and FA
Seconda y oxida ion compounds
Aldehydes
a9.75 -CHO alkanals
Abb e ia ions: , iple ; m, mul iple ; d, double ; s, single ; DHA, docosahexaenoa e (C22:6
Ñ
3); EPA, eicosapen-
aenoa e (C20:5
ω
3); 1,3-DG, 1,3-diglyce ide; 1-MG, 1-monoglyce ide; TG, iglyce ide; 1,2-DG, 1,2-diglyce ide;
2-MG, 2-monoglyce ide. * o e lapping o mul iple s o me hylenic p o ons in he di e en acyl g oups ei he in
be a-posi ion, o u he , in ela ion o double bonds, o in gamma-posi ion, o u he , in ela ion o he ca bonyl
g oup; ** o e lapping o mul iple s o he alpha-me hylenic p o ons in ela ion o a single double bond o he
di e en unsa u a ed acyl g oups; *** his signal shows di e en mul iplici y i he spec um is acqui ed om he
pu e compound o aking pa in he mix u e.
2.4. S a is ical Analysis
Da a p o ided in Tables 2–4 a e a e age alues o he se e al de e mina ions o each
sample, oge he wi h he co esponding s anda d de ia ion, calcula ed using Mic oso
Excel 2016.
3. Resul s and Discussion
3.1. 1H NMR S udy o he Co e ing Oils o Canned Sa dines F eshly Pu chased
I is well known ha ege able oils, like ish lipids, a e composed mainly o iglyc-
e ides (TG), which can ha e sa u a ed, mono-, di-, and polyunsa u a ed acyl g oups (AG) in
di e en p opo ions depending on hei na u e. The p o ons o hese acyl g oups gene a e
di e en main signals om 0 o 5.5 ppm in he
1
H NMR spec a. Figu e 1shows he spec a
o ou illing oils o di e en na u e; hese a e soybean (SYO2), sun lowe (SFO2), ege able
oil o unspeci ied o igin (VO4), and oli e oil (OO5, ex a- i gin oli e oil). In he uppe side
o his igu e, some signals ha e been p ope ly enla ged o compa a i e pu poses. The
assignmen o he signals is gi en in Table 1, in ag eemen wi h p e ious s udies [16–21].
Signal A, be ween 0.84 and 0.94 ppm, is due o he o e lapping o he iple s o
me hylic p o ons o he sa u a ed (Sa ), monounsa u a ed
ω
9 and/o
ω
7 (MU) acyl g oups,
and ha o he diunsa u a ed
ω
6 (DU
ω
6) acyl g oups. Thus, signal A can p o ide six clea ly
dis inguishable peaks, h ee o hem belonging o he me hylic p o ons o sa u a ed and
Foods 2025,14, 1589 5 o 14
monounsa u a ed acyl g oups ( iple cen ed a 0.879 ppm) and he o he h ee belonging o
diunsa u a ed acyl g oups ( iple cen ed a 0.889 ppm). As can be obse ed in he spec a
o SYO2, SFO2, and VO4, he iple a 0.889 ppm is highe han he iple a 0.879 ppm,
e idencing a highe p opo ion o diunsa u a ed
ω
6 acyl g oups (mainly linoleic, C18:2
ω
6)
han o sa u a ed and monounsa u a ed acyl g oups. The opposi e is obse ed in signal
A o OO5 spec um, in ag eemen wi h he high con en o monounsa u a ed oleic acyl
g oups (C18:1ω9) in oli e oils.
Foods 2025, 14, x FOR PEER REVIEW 5 o 14
Figu e 1. 1H NMR spec a be ween 0 and 5.5 ppm o co e ing oils o di e en na u e: soybean oil
(SYO2), sun lowe oil (SFO2), ege able oil (VO4), and ex a- i gin oli e oil (OO5). Some spec al
egions we e p ope ly enla ged in he uppe pa o he igu e o compa a i e pu poses. Signal
le e s ag ee wi h hose in Table 1.
Figu e 2. Some 1H NMR spec al egions o wo samples be o e (SYO1, OO1) and a e being sub-
mi ed o s o age a oom empe a u e o i een yea s (ST_SYO1, ST_OO1): (a) Regions whe e p o-
ons in he glyce yl backbone o mono-, di-, and iglyce ides a e obse ed, (b) Regions whe e he
Figu e 1.
1
H NMR spec a be ween 0 and 5.5 ppm o co e ing oils o di e en na u e: soybean oil
(SYO2), sun lowe oil (SFO2), ege able oil (VO4), and ex a- i gin oli e oil (OO5). Some spec al egions
we e p ope ly enla ged in he uppe pa o he igu e o compa a i e pu poses. Signal le e s ag ee
wi h hose in Table 1.
Signal B, be ween 0.94 and 1.00 ppm, is due o he iple o me hylic p o ons o
ω
3
polyunsa u a ed acyl g oups, which is cen e ed a 0.972 ppm and can co espond ei he
o he linolenic acyl g oups o he o iginal ege able oil (C18:3
ω
3), o , i mig a ion has
occu ed, also o he
ω
3 sa dine lipids; his is mainly EPA and DHA [
22
]. As can be
obse ed, signal B shows a highe in ensi y in he spec a o SYO2 and OO5 han in hose
o SFO2 and VO4. This is in ag eemen wi h he high con en in linolenic acyl g oups
in soybean oils (4.5–11.0%), and wi h hei almos comple e absence in sun lowe oils
(<0.3%) [
23
]. Rega ding OO5 spec a, he in ensi y o signal B is much highe han ha o a
ypical oli e oil, which con ains low p opo ions o linolenic (<1.5%); his ac sugges s ha
in his co e ing oil, Ñ3 ish lipids a e also p esen .
Signal C, be ween 1.19 and 1.42 ppm, is due o me hylenic p o ons ei he in posi ion
be a, o u he in ela ion o double bonds, o in posi ion gamma, o u he in ela ion o
he ca bonyl g oup in he di e en acyl g oups. I has wo main peaks, one a 1.257 ppm
co esponding o he me hylenic p o ons o sa u a ed acyl g oups and he o he one nea
1.300 ppm associa ed wi h he o e lapping o me hylenic p o ons o all he unsa u a ed
acyl g oups [
17
]. In addi ion, i has a shoulde nea 1.280 ppm ela ed o monounsa u a ed
acyl g oups. This was obse ed in he OO5 spec um, bu i is no enla ged in Figu e 1.
Foods 2025,14, 1589 6 o 14
Signal D1, be ween 1.54 and 1.67 ppm, is due o me hylenic p o ons in he be a posi ion
in ela ion o he ca bonyl g oup, excep hose o EPA and DHA acyl g oups (see Figu e 1).
As is known, his signal does no show signi ican di e ences in mul iplici y o in chemical
shi s be ween he
1
H NMR spec a o he di e en acyl g oups [
17
]. Signal D2, be ween
1.67 and 1.74 ppm, is due o me hylenic p o ons in he be a posi ion in ela ion o he
ca bonyl g oup o EPA acyl g oups. Thus, al hough i is absen in he
1
H NMR spec a o
ege able oils [
17
], i can be obse ed in he enla ged spec a o he co e ing oils shown in
Figu e 1, especially in OO5 and SYO2, e idencing he mig a ion o EPA acyl g oups om
he sa dine muscle o he packing oil.
Signal E, be ween 1.92 and 2.15 ppm, is due o he o e lapping o he a ious signals
o allylic p o ons, ha is, o alpha-me hylenic p o ons in ela ion o a single double bond
in he di e en acyl g oups, excep hose o DHA acyl g oup, which a e also in be a-
posi ion in ela ion o he ca bonyl g oup. As sa u a ed acyl g oups do no ha e double
bonds, hey make no con ibu ion o his signal. Rega ding his signal, he oli e oil
sample OO5 is clea ly dis inguishable om he o he co e ing oils, because he peaks a
2.002 and 2.020 ppm
co esponding o monounsa u a ed acyl g oups a e much highe han
hose a
2.036, 2.056, and 2.074 ppm
due o diunsa u a ed
ω
6 acyl g oups (mainly linoleic).
I mus be no ed ha in he spec a o OO5 and SYO2, a peak a 2.093 ppm can be obse ed
due o he allylic p o ons o
ω
3 acyl g oups. This is in ag eemen wi h wha was obse ed
in signal B.
Signal F1, be ween 2.26 and 2.36 ppm, is due o me hylenic p o ons in he alpha-
posi ion in ela ion o he ca bonyl g oup, excep hose o DHA acyl g oup (see Figu e 1).
This signal does no show signi ican di e ences in mul iplici y o in chemical shi s
be ween he
1
H NMR spec a o he di e en acyl g oups suppo ed on iglyce ides; o
his eason, i is no use ul o disc imina e co e ing oils. Signal F2 be ween 2.37 and 2.41 is
due o me hylenic p o ons in alpha and be a-posi ions in ela ion o he ca bonyl g oup o
DHA acyl g oups, being p esen in he spec a o ish lipids bu no in hose o ege able
oils [
16
,
20
]. As shown in he enla ged spec a o Figu e 1, signal F2 is p esen in he ou
co e ing oils, mainly in OO5 and SYO2; his is in ag eemen wi h he abo e commen ed on
signal D2 due o EPA acyl g oups.
Signal Gis a iple o he bis-allylic p o ons o diunsa u a ed
ω
6 (linoleic) acyl g oups,
ha is o hei me hylenic p o ons in
α
-posi ion in ela ion o wo double bonds. This signal
is pa ially o e lapped wi h Signal H, also due o he bis-allylic p o ons bu o he es o
polyunsa u a ed acyl g oups, which a e mainly
ω
3. As can be obse ed in he spec a o
Figu e 1, he in ensi y o signal G is much highe in SYO2, SFO2, and VO4 han in OO5, in
ag eemen wi h ha commen ed on abo e ega ding signals A and E. On he o he hand,
he in ensi y o signal H is highe in OO5 and SYO2 han in SFO2 and VO4, which is in
ag eemen wi h ha epo ed conce ning signals B, D2, E (peak a 2.093 ppm), and F2.
Fu he mo e, signals due o he p o ons o he glyce ol backbone o TG can also be
obse ed. Signal Oa 4.10–4.34 ppm is due o he p o ons bonded o ca bon a oms 1 and 3
o he glyce yl g oup, and signal Sa 5.23–5.30 ppm due o hose o ca bon a om 2. The
la e signal o e laps sligh ly wi h signal T, a 5.28–5.47 ppm, due o ole inic p o ons o
all he unsa u a ed acyl g oups. These signals O, S, and T do no show di e ences in
mul iplici y o chemical shi s be ween he di e en acyl g oups and hus do no p o ide
use ul in o ma ion o disc imina ing be ween di e en oil samples.
No only quali a i e bu also quan i a i e in o ma ion can be ob ained om he s udy
o hese
1
H NMR signals. Table 2p o ides he mola pe cen ages o he di e en kinds o
acyl g oups p esen in he illing oils. In gene al, oli e oils showed he highes p opo ion
o monounsa u a ed acyl g oups (MU, mainly oleic) and he lowes o diunsa u a ed
ω
6 acyl g oups (DU
ω
6, mainly linoleic). In he seed oils, much highe p opo ions o
Foods 2025,14, 1589 7 o 14
DU
ω
6 we e obse ed. I is ema kable ha in all he samples, he p esence o DHA
(0.8–8.3%) and o EPA (1.0–9.0%) was epo ed in a iable p opo ions. These esul s
a e in ag eemen wi h p e ious s udies ca ied ou by ch oma og aphic echniques ha
epo ed he in e change o lipids be ween canned sa dine muscle and he oli e oil used
as packing medium [
8
,
24
], which sugges ed ha he e is a di usion g adien ha causes
ha he acyl g oup p opo ions o sa dine lipids and he co e ing oil end o be simila . In
o he ish species, like una canned in soybean oil, simila esul s ha e also been epo ed
by ch oma og aphic echniques [
5
]. As can be obse ed in Table 2, soybean oil samples
con ained he highes p opo ions o o al
Ñ
3 lipids, due o he ac ha in addi ion o
DHA and EPA mig a ed om sa dine muscle, his kind o oil na u ally con ains highe
p opo ions o linolenic acyl g oups (4.5–11.0%) han oli e (<1.5%) o sun lowe (<0.3%)
oils [
23
]. Rega ding sa u a ed acyl g oups, a iable p opo ions we e de ec ed in all he
samples. The highes p opo ion o sa u a ed acyl g oups was obse ed in a soybean oil
sample, SYO4*. As his sample also con ained he highes p opo ions o DHA and EPA,
his ac could be a ibu ed o he mig a ion o sa dine sa u a ed lipids o he illing oil,
in ag eemen wi h p e ious s udies on sa dines canned in oli e oil [
24
]. I mus be no ed
ha his sample SYO4* is one o hose in which he oil is no he main ing edien o he
illing medium, and hus ish lipids a e mo e concen a ed han in o he samples (see Table
S1). The ac ha hese a e comme cial samples p epa ed om aw ma e ials o unknown
ini ial composi ion, which ha e di e en inal weigh s and bes -be o e da es (see Table S1)
and which ha e unde gone di e en manu ac u ing p ocesses, makes i di icul o d aw
u he conclusions abou he a iabili y o he p opo ions o acyl g oups in he co e ing
oils epo ed in Table 2.
Table 2. Mola pe cen ages o he main acyl g oups o he co e ing oils o canned sa dines eshly
pu chased in local supe ma ke s, es ima ed by 1H NMR.
Sample To al ω3 DHA EPA DUÑ6 MU To al U Sa
OO1 6.8 ±0.0 2.8 ±0.0 3.8 ±0.0 7.6 ±0.1 68.9 ±0.9 83.2 ±1.0 16.8 ±1.0
OO2 5.7 ±0.1 2.3 ±0.1 3.3 ±0.2 6.5 ±0.2 73.4 ±0.1 85.5 ±0.1 14.5 ±0.2
OO3 8.0 ±0.1 3.7 ±0.1 3.7 ±0.1 8.3 ±0.2 65.0 ±0.6 81.3 ±0.3 18.7 ±0.3
OO4 4.0 ±0.0 1.8 ±0.0 1.7 ±0.2 5.9 ±0.2 75.4 ±0.2 85.3 ±0.1 14.7 ±0.1
OO5 10.6 ±0.1 4.0 ±0.1 5.9 ±0.2 6.4 ±0.1 65.4 ±0.4 82.4 ±0.3 17.6 ±0.3
OO6 7.9 ±0.1 2.1 ±0.0 5.7 ±0.1 7.9 ±0.0 69.9 ±0.5 85.6 ±0.6 14.4 ±0.6
OO7 2.7 ±0.4 0.9 ±0.0 1.8 ±0.2 6.7 ±0.1 78.2 ±1.0 87.6 ±0.5 12.4 ±0.5
SFO1 2.5 ±0.1 0.8 ±0.1 1.4 ±0.1 55.1 ±0.8 30.4 ±0.8 88.0 ±0.0 12.0 ±0.0
SFO2 4.8 ±0.1 0.9 ±0.0 3.8 ±0.2 54.4 ±0.5 27.6 ±1.4 86.7 ±1.0 13.3 ±1.0
SFO3 1.9 ±0.1 0.8 ±0.0 1.0 ±0.1 64.0 ±0.9 23.0 ±1.4 88.9 ±0.4 11.1 ±0.5
SFO4 6.2 ±0.1 2.1 ±0.0 3.8 ±0.0 45.9 ±3.6 34.1 ±3.7 86.2 ±0.0 13.8 ±0.0
SFO5 1.7 ±0.1 0.6 ±0.1 1.1 ±0.2 66.4 ±1.1 23.1 ±0.2 91.2 ±1.0 8.8 ±1.0
SFO6* 13.3 ±0.2 5.8 ±0.2 6.5 ±0.3 37.7 ±1.3 31.5 ±1.8 82.4 ±0.4 17.6 ±0.4
SYO1 15.2 ±0.0 4.9 ±0.1 6.0 ±0.2 40.8 ±2.6 24.0 ±3.1 80.0 ±0.5 20.0 ±0.5
SYO2 12.9 ±0.3 3.0 ±0.1 5.6 ±0.3 45.7 ±0.5 23.7 ±0.3 82.3 ±0.0 17.8 ±0.0
SYO3* 14.1 ±0.1 3.4 ±0.0 5.9 ±0.1 41.9 ±0.4 24.6 ±0.4 80.6 ±0.1 19.4 ±0.1
SYO4* 21.5 ±0.2 8.3 ±0.1 9.0 ±0.1 28.3 ±0.5 25.9 ±0.1 75.7 ±0.6 24.3 ±0.7
VO1 9.0 ±0.3 1.7 ±0.1 2.5 ±0.1 51.5 ±1.8 23.3 ±2.5 83.8 ±0.3 16.2 ±0.4
VO2 2.1 ±0.0 0.8 ±0.0 1.2 ±0.1 61.8 ±1.8 25.0 ±1.4 88.8 ±0.4 11.2 ±0.4
VO3 9.2 ±0.0 4.4 ±0.0 4.3 ±0.0 32.2 ±1.7 42.3 ±0.3 83.8 ±2.0 16.3 ±2.0
VO4 4.8 ±0.2 1.3 ±0.3 3.2 ±0.7 48.8 ±2.3 32.8 ±2.7 86.4 ±0.2 13.6 ±0.3
VO5 5.1 ±1.0 2.3 ±0.1 2.6 ±1.0 55.3 ±4.1 27.5 ±4.6 87.9 ±0.4 12.1 ±0.4
OO: oli e oil; SFO: sun lowe oil; SYO: soybean oil; VO: ege able oil;
ω
-3: omega-3; DHA: docosahexaenoic; EPA:
eicosapen aenoic; DU
Ñ
6: diunsa u a ed omega-6; MU: monounsa u a ed, Sa : sa u a ed; U: o al unsa u a ed.
As e isked samples did no con ain oil as he main ing edien o he illing medium.
Foods 2025,14, 1589 8 o 14
In sho ,
1
H NMR p o ided a g ea deal o in o ma ion on he lipid composi ion o he
co e ing oils. I allowed us o de ec he mig a ion o sa dine
ω
3 polyunsa u a ed lipids,
e idenced by he p esence in he spec a o signal D2 ela ed o EPA and signal F2 ela ed
o DHA acyl g oups, which a e absen in he ypical spec a o ege able oils. Mo eo e ,
signals A and E made i possible o dis inguish wi h he naked eye he
1
H NMR spec a
o oli e oils om he o he seed oil samples, due o he high con en o monounsa u a ed
oleic acyl g oups in he o me . Howe e , he spec a o sun lowe , soybean, and ege able
oils showed many simila i ies, p obably because mos o he oils labelled as ‘ ege able’ a e,
indeed, sun lowe oil, soybean oil, o mix u es he eo , among o he s. This ac , oge he
wi h he mig a ion o ish lipids, makes he assessmen o he genuineness o he oil used
as liquid medium in canned sa dines di icul . Simila esul s we e epo ed by means
o a enua ed o al e lec ion Fou ie ans o m in a ed spec oscopy (ATR-FTIR) and
chemome ics on he au hen ica ion o packing oils om comme cial canned una, which
di e en ia ed oli e oil om seed oils (sun lowe and ege able oils) [
25
]. In his ega d, i
mus be no ed ha i a huge numbe o
1
H NMR spec a o co e ing oils o di e en na u e
we e s udied oge he wi h he powe o mul i a ia e analyses, his echnique could be a
e y good inge p in ing and disc imina ing me hod [26].
As o he oxida i e s a us o he co e ing oils, no p o on signals ela ed o p ima y
o o seconda y oxida ion p oduc s we e obse ed in hei
1
H NMR spec a [
18
], which
e idenced ha no ele an lipid oxida ion eac ions had occu ed in he co e ing oils.
These esul s a e in ag eemen wi h hose ob ained in he s udy o he oxida i e s a us o
co e ing oli e oils o canned una by Pe oxide Value [
9
]. On he con a y, a highe oxida i e
deg ada ion le el was epo ed in seed oils (sun lowe , soybean, co n, ege able) han
in oli e oils used as illing mediums in canned sa dines, una, ancho ies, and macke el,
acco ding o hei pe cen age o pola compounds [10,11].
Conce ning he hyd oly ic s a us o he co e ing oils, in he spec a o all he samples,
incipien signal Ja 3.73 ppm was obse ed; his is due o he me hylenic p o ons in ca bon
a om 3 o he glyce yl backbone o 1,2-diglyce ides (1,2-DG). This small p opo ion o
1,2-DG is ypically p esen in ege able oils and is conside ed negligible. In a p e ious
s udy, a highe hyd oly ic le el (F ee Fa y Acids pe cen age, FFA%) was epo ed o oli e
oils han o seed oils used as illing mediums in di e en canned ish, bu his ac was
a ibu ed o he e ining p ocess o seed oils ha emo e hese compounds. Rega dless,
in all he samples, FFA% was below he legal limi s es ablished o he di e en ypes o
oils [10,11].
3.2. 1H NMR S udy o he Co e ing Oils o Canned Sa dines A e Fi een-Yea S o age
When he
1
H NMR spec a o he co e ing oils submi ed o i een-yea s o age we e
s udied, he main change obse ed was he appea ance o new signals be ween 3.6 and
5.1 ppm due o p o ons in he glyce yl backbone o mono- and diglyce ides. Figu e 2a
shows hese enla ged egions o he spec a o one soybean and one oli e co e ing oils
be o e (SYO1, OO1) and a e s o age (ST_SYO1, ST_OO1) o compa a i e pu poses. In
addi ion o signal O due o p o ons in he glyce yl backbone o TG, which is ypical in
he spec um o edible oils, new signals can be obse ed due o p o ons in he glyce yl
backbone o pa ial glyce ides (see Table 1): signal M o 1,3-diglyce ides (1,3-DG), signals J,
Pand R ela ed o 1,2-diglyce ides (1,2-DG), signals I,Land N o 1-monoglyce ides (1-MG),
and signals Kand Q o 2-monoglyce ides (2-MG).
Table 3shows he a e age mola pe cen ages o he TG, 1,3-DG, 1,2-DG, 1-MG, 2-MG,
and glyce ol (Gol) de ec ed in he co e ing oils a e s o age. I mus be no ed ha in
edible oils, TG accoun s o app oxima ely 97–98% o o al glyce ides p esen . As can be
obse ed in Table 3, in s o ed co e ing oils, TG anged om 78.7% in ST_OO5 o 37.9% in
Foods 2025,14, 1589 9 o 14
ST_OO1. The spec um o he la e sample is shown in Figu e 2a and a g ea dec ease in
he in ensi y o signal O can be obse ed a e s o age. The pa ial glyce ide ha showed
he highes p opo ions in he co e ing oils we e 1,3-DG, anging om 24.0% in ST_OO1 o
12.0% in ST_OO5. They we e gene a ed due o he b eakdown o he es e bond in ca bon
2 o he glyce yl backbone o TG, also eleasing one a y acid (FA). Mo eo e , 1,2-DG
we e also de ec ed in he samples, bu in lowe p opo ions ( anging om 9.9% in ST_OO1
o 5.0% in ST_OO2 and ST_OO5). They we e o med due o he b eakdown o he es e
bond in ca bon 3 o he glyce yl backbone o TG. Rega ding monoglyce ides, 1-MG we e
p esen in all samples, wi h sligh ly lowe p opo ions han hose o 1,2-DG ( anging om
15.6% in ST_OO1 o 3.0% in ST_OO5). On he o he hand, 2-MG we e also de ec ed in
all samples, bu in much lowe p opo ions (<1.0%), which we e lowe han hose o Gol.
The highe p opo ions o 1-MG han 2-MG can be explained by he abo e-men ioned
p e e en ial o ma ion o 1,3-DG, because as hyd olysis ad ances, 1,3-DG gi e ise o 1-
MG and one FA. These esul s a e in ag eemen wi h p e ious s udies ca ied ou by
13
C
NMR on lipid hyd olysis occu ing in aw and hea - ea ed una muscle du ing indus ial
canning [
27
,
28
]. Du ing ozen s o age o aw muscle, a p e e en ial o ma ion o 1,2-DG
was epo ed due o he ac i i y o lipoly ic enzymes. Howe e , a e he mal p ocessing
(cooking and s e iliza ion in cans), a di e en lypolisis mechanism was e idenced, epo ing
a p e e en ial o ma ion o 1,3-DG. These esul s we e a ibu ed o a physical b eakdown
o he es e bond in ca bon a om 2 o TG due o a hea e ec , because lipase enzymes we e
no ac i e due o he high empe a u es applied du ing canning [27].
Foods 2025, 14, x FOR PEER REVIEW 5 o 14
Figu e 1. 1H NMR spec a be ween 0 and 5.5 ppm o co e ing oils o di e en na u e: soybean oil
(SYO2), sun lowe oil (SFO2), ege able oil (VO4), and ex a- i gin oli e oil (OO5). Some spec al
egions we e p ope ly enla ged in he uppe pa o he igu e o compa a i e pu poses. Signal
le e s ag ee wi h hose in Table 1.
Figu e 2. Some 1H NMR spec al egions o wo samples be o e (SYO1, OO1) and a e being sub-
mi ed o s o age a oom empe a u e o i een yea s (ST_SYO1, ST_OO1): (a) Regions whe e p o-
ons in he glyce yl backbone o mono-, di-, and iglyce ides a e obse ed, (b) Regions whe e he
Figu e 2. Some
1
H NMR spec al egions o wo samples be o e (SYO1, OO1) and a e being
submi ed o s o age a oom empe a u e o i een yea s (ST_SYO1, ST_OO1): (a) Regions whe e
p o ons in he glyce yl backbone o mono-, di-, and iglyce ides a e obse ed, (b) Regions whe e he
main signals ela ed o polyunsa u a ed omega-3 lipids a e obse ed. Signal le e s ag ee wi h hose
in Table 1.
This inc ease in pa ial glyce ides obse ed in all he co e ing oils a e s o age is in
ag eemen wi h o he s udies on canned una s o ed o up o 6 yea s, which epo ed ha
an inc ease in he s o age ime p oduced highe FFA% in ish muscle and in he co e ing
oil (oli e oil) [
9
]. In ha s udy, cans ha con ained only oli e oil (blank oils wi hou una)
