Fa y Acid Hyd oxy y osyl Es e s o Oli e Oils A e Bioaccessible
Acco ding o Simula ed In Vi o Gas oin es inal Diges ion:
Un a eling he Role o Diges i e Enzymes on Thei S abili y
Ca olina Alemán-Jiménez,
#
Raul Domínguez-Pe les,
#
Juana I. Gallego-Gómez, Agus ín Simonelli-Munoz,
Espé ance Moine, Thie y Du and, Céline C aus e, Fede ico Fe e es, Angel Gil-Izquie do,*
and Sonia Medina*
Ci e This: J. Ag ic. Food Chem. 2021, 69, 14165−14175
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ABSTRACT: Recen ly, new bioac i e compounds we e iden ified in oli e oil, lipophenols, which a e composed o a a y acid (FA)
and a phenolic co e, such as HT (HT-FA). Howe e , hei bioaccessibili y emains unknown. Thus, he p esen s udy unco e s he
impac o he sepa a e phases o gas oin es inal diges ion on he elease and s abili y o HT-FAs om oily ma ices unde in i o
simula ed condi ions. Acco dingly, i was ound ha he bioaccessibili y o HT de i a i es is la gely dependen on he ype o FA ha
es e ifies HT, as well as he ood ma ix. Also, he gene a ion o HT-FAs du ing in es inal diges ion was obse ed, wi h panc ea in
being he enzyme esponsible, o a highe ex en , o he de no o o ma ion o lipophenolic de i a i es. These findings p omp us o
iden i y new applica ions o oily ma ices and hei byp oduc s as po en ial unc ional ing edien s o he p omo ion o heal h, whe e
he possible o ma ion o new lipophenols du ing diges ion should be aken in o conside a ion.
KEYWORDS: oli e oil, hyd oxy y osol, lipophenols, in i o gas oin es inal diges ion, bioaccessibili y, lipases
■INTRODUCTION
Oli e d upes and de i ed p oduc s a e conside ed pa o he
Medi e anean die and co-con ibu e o he heal hy cha ac e -
is ics o his die , especially because o hei con en o mono-
and polyunsa u a ed a y acids (FAs) and (poly)phenols such
as hyd oxy y osol (HT).
1,2
Mo eo e , oleic acid is he mos
abundan FA in oli e oil, and HT is i s p incipal phenolic
compound.
3
Also, he p esence o lipophenol s uc u es,
composed o an FA and a phenolic co e, has been epo ed
in edible oils. These molecules esul om he es e ifica ion o
HT wi h a y acids (e.g.,α-linolenic, linoleic, and oleic acids
(HT-ALA, HT-LA, and HT-OA, co espondingly)).
4,5
To
da e, ex a i gin and i gin oli e oils (EVOO and VOO,
espec i ely) a e he oily ma ices wi h he highes
concen a ion o hese compounds.
5
Lipophenols offe unc ional ad an ages ela i e o hei
molecula cons i uen s (HT and FAs). Specifically, hese
compounds a e cha ac e ized by a highe lipophilici y, cell
memb ane affini y, and imp o ed an ioxidan ac i i ies.
6−8
Howe e , despi e he sugges ion o lipophenol’s biological
powe ,
9,10
he ac ual unc ions emain almos unexplo ed.
Ne e heless, ega ding hei an ioxidan p ope y, i may
p og essi ely be augmen ed along wi h he ex ension o he FA
chain leng h up o a c i ical poin , when addi ional ex ension
could lowe he adical sca enging powe . Because o his
phenomenon, he so-called cu offeffec , sho - o medium-
chain lipophilic es e s o phenolic compounds (C8−C12) a e
mo e effec i e an ioxidan s ela i e o long-chain es e s.
11
In
con as , o he s udies ha e associa ed equal o highe ac i i y
wi h long-chain es e s (C18−C20).
12,13
Aside om hese conside a ions, he bioaccessibili y o
lipophenols emains o be assessed. In his ega d, acco ding o
p e ious s udies, in ense hyd olysis o HT ace a e a e
panc ea in and bile sal diges ion has been epo ed,
14
while
y osol acyl es e s a e hyd olyzed by panc ea ic lipase o
p oduce ee y osol, showing a ce ain sus ained- elease
beha io .
15,16
These esul s sugges ha he ime ame o
e ie ing biological ac i i ies om phenolic compounds could
be ex ended due o lipophenol’s bioac i i y, hus allowing us o
ob ain addi ional heal h benefi s.
15
These au ho s epo ed ha
he s abili y o y osol es e s unde he physicochemical
condi ions associa ed wi h he in i o simula ion o gas o-
in es inal (GI) diges ion was co ela ed bo h wi h he chain
leng h and wi h he numbe o unsa u a ions.
16
None heless,
he effec o he diges i e p ocess on he elease o HT-FAs
and hei s abili y emains unde explo ed. Also, he expe -
imen al app oach implemen ed hus a , based on he analysis
o au hen ic s anda ds ins ead o ood ma ices, is negligible
because i igno es he effec o he ma ix h oughou he
diges ion.
14
To o e come his limi a ion, a simula ed GI
Recei ed: Augus 30, 2021
Re ised: No embe 11, 2021
Accep ed: No embe 12, 2021
Published: No embe 19, 2021
A iclepubs.acs.o g/JAFC
© 2021 Ame ican Chemical Socie y 14165
h ps://doi.o g/10.1021/acs.ja c.1c05373
J. Ag ic. Food Chem. 2021, 69, 14165−14175
Downloaded ia UNIV DE ALMERIA on Decembe 1, 2021 a 19:13:42 (UTC).
See h ps://pubs.acs.o g/sha ingguidelines o op ions on how o legi ima ely sha e published a icles.
diges ion by in i o models could be a aluable, ime-sa ing
ool, as i equi es ewe esou ces o ob aining p elimina y
bu ex emely impo an in o ma ion o he e alua ion o he
s uc u al and chemical changes unde GI condi ions.
17
Acco ding o his backg ound in o ma ion and p e ious
findings, he p esen wo k aims o gain u he insigh in he
esul s o he GI diges ion on HT-ALA, HT-LA, and HT-OA
ound in EVOO and VOO o iden i y he lipophenols eleased
om he ood ma ix ha emain s able du ing GI diges ion
and ha a e a ailable o abso p ion. Also, he e alua ion o he
effec o pepsin, panc ea in, and panc ea ic lipase allowed us o
disco e he influence o he diges i e enzymes on he s abili y
o lipophenols. The majo ou comes e ie ed con ibu ed o
se ing up he bioaccessibili y o HT-FAs in he small in es ine
compa ed wi h unes e ified HT and o unde s anding he
enzyma ic mechanisms esponsible o hem, p o iding he
heo e ical basis o he ans o ma ion o lipophenols du ing
he diges i e p ocess.
■MATERIALS AND METHODS
Chemicals and Reagen s. The au hen ic s anda ds o high-pu i y
HT-FAs (HT-ALA, HT-LA, and HT-OA) we e syn hesized and ully
cha ac e ized by nuclea magne ic esonance (NMR)-based analysis
by he Ins i u des Biomolecules Max Mousse on (IBMM)
(Mon pellie , F ance) acco ding o p e iously published p ocedu es.
4
Ace one and bu yla ed hyd oxyanisole (BHA) we e pu chased om
Sigma-Ald ich (S . Louis, MO, USA), and all LC−MS-g ade sol en s
(deionized wa e , me hanol, and ace oni ile) we e om J.T. Bake
(Phillipsbu g, NJ, USA). A ce ified e e ence o FA s anda ds om
Sigma-Ald ich was used o hei iden ifica ion and quan ifica ion.
Hyd oxy y osol, wi h he ollowing specifica ions: pu i y 99.6%,
mois u e 3.7%, and pH 4.26 (1 M aqueous solu ion), was p o ided
by Sep ox BIOTECH S.L. (Mu cia, Spain). All lipophenols we e
dissol ed in dime hyl sul oxide (DMSO) om Sigma Ald ich (S .
Louis, MO, USA) o ob ain a s ock solu ion (a he mM ange o
concen a ion), and hen successi e dilu ions we e p epa ed in
me hanol/deionized Milli-Q wa e (50:50, / ). All s ock solu ions o
lipophenols we e s o ed a −20 °C in he da k. Po cine pepsin
(P6887), panc ea in om po cine panc eas (P7545, 8 ×USP), and
lipase om po cine panc eas (L3126) we e ob ained om Sigma-
Ald ich Co. (S . Louis, MO, USA). All o he eagen s we e o
analy ical g ade.
Fa y Acid Composi ion Analysis o Edible Oils. Fa y acids o
EVOO, VOO, and flaxseed oil (FO) we e ex ac ed acco ding o he
me hodology employed p e iously
5
and published by he FAO
(FAOLEX No. LEX-FAOC141241, h p://www. ao.o g/ aolex/
esul s/de ails/es/c/%20LEX-FAOC141241/) o he analysis o
ege able oils. B iefly, samples (1 g) we e weighed and d ied a 70
±10 °C o 6 h. The a y acid con en was ob ained by he Soxhle
ex ac ion wi h die hyl e he and hep ane and by es e ifica ion wi h
me hanolic NaOH (1 M). Fa y acids we e de e mined by gas
ch oma og aphy coupled o a flame ioniza ion de ec o (GC-FID)
(6890 GC Agilen Technologies, Wald onn, Ge many). The absolu e
concen a ion (g/100 g w) o a y acids in he ege able oils (n=3)
was calcula ed acco ding o he o mula indi idual a y acid (g/100 g
w) = indi idual a y acid (% o a )/100 × o al a (g/100 g w),
which was used acco ding o FAO/INFOODS guidelines o
con e ing uni s, denomina o s, and exp ession (2012).
18
Vege able Oil Samples and P epa a ion o Analy ical
Ex ac s o Hyd oxy y osol Fa y Acid Es e s. Ex a i gin oli e
oil (Picual mono a ie al; Olimend os S.L. (Mu cia, Spain)), VOO
(Picual mono a ie al; Sal ado Gallego El Jo a S.L. (Albace e,
Spain)), and FO (Labo a o ios Almond S.L. (Mu cia, Spain)) we e
chosen due o hei dissimila concen a ion o HT es e ified wi h
ALA, LA, and OA, wi h FO used as he nega i e con ol sample due
o i s lack o HT-FA es e con en .
5
All oil samples we e kep in da k
glass bo les, closed wi h sc ew caps, and s o ed a 4 °C o a oid oil
oxida ion o he g ea es possible ex en . The HT-FAs we e ex ac ed
om ege able oils acco ding o he me hodology desc ibed
p e iously.
5
In Vi o Simula ed Gas oin es inal Diges ion. To ep oduce
he sepa a e diges ion phases (gas ic, in es inal, and GI), he edible
oils (EVOO, VOO, and FO) we e p ocessed based on he
ha monized in i o diges ion p o ocol desc ibed in he li e a u e,
19,20
using he simula ed gas ic and in es inal fluids (SGF and SIF,
espec i ely) s ock elec oly e solu ion, de eloped acco ding o he
in o ma ion p o ided in Table 1.
A e diges ion, he samples we e cen i uged a 1600g o 5 min a
4°C o sepa a e he bioaccessible ac ion (BF) and he uppe oily
phase o esidual ac ion (RF); he concen a ion o HT and HT-FAs
in each ac ion was analyzed. Blanks (nega i e con ols) wi hou
enzymes we e p ocessed and analyzed in pa allel unde equal
condi ions. All samples we e p o ec ed om ligh o e he en i e
p ocess. Bo h BF and RF ac ions we e ozen immedia ely a −80 °C
and lyophilized. Fo he HT and HT-FA ex ac ion, he lyophilized
samples we e hen dissol ed in 3 mL o an ace one/BHA
(99.995:0.005, /w) solu ion, o exed o 1 min, sonica ed o 30
min a 40 kHz, and cen i uged a 8750g o 5 min a 4 °C acco ding
o he p ocedu e desc ibed.
5
The samples we e d ied using a
SpeedVac concen a o , and he d y ex ac s we e econs i u ed wi h
500 μL o MeOH, sonica ed o 10 min, cen i uged a 8750g o 5
min, and fil e ed h ough a 0.45 μmfil e (Millipo e, Bu ling on, MA,
USA).
UHPLC-ESI-QqQ-MS/MS Analysis. The sepa a ion and iden i-
fica ion o HT and HT-FAs we e pe o med using a UHPLC coupled
wi h a iple quad upole MS/MS (Agilen Technologies, Waldb onn,
Ge many) acco ding o he me hodology epo ed in he li e a u e.
5
The e en ion imes eco ded o unes e ified HT, HT-ALA, HT-LA,
and HT-OA we e 0.6, 3.8, 4.0, and 4.4 min, espec i ely. The analyses
we e pe o med by mul iple eac ion moni o ing (MRM) in he
nega i e mode, and he quan ifica ion and confi ma ion MRM
ansi ions we e as ollows: m/z 153 > 123 a bi a y mass uni s
(amu) and m/z 153 > 95 amu, espec i ely, o HT; m/z 413 > 277
amu and m/z 413 > 260 amu, co espondingly, o HT-ALA; m/z 415
> 279 amu and m/z 415 > 262 amu, espec i ely, o HT-LA; and m/z
417 > 281 amu and m/z 417 > 264 amu, co espondingly, o HT-
OA. Da a acquisi ion and p ocessing we e done using he MassHun e
so wa e e sion B.08.00 (Agilen Technologies, Waldb onn,
Ge many). Bo h HT and es e s o HT we e quan ified using
e e ence s anda ds, and he concen a ions we e exp essed as
nanog ams pe g am o esh weigh (ng/g w).
S a is ical Analyses. All he analy ical ex ac ions o he oily
ma ices conside ed in he p esen wo k, as well as he p oduc s o he
gas ic, in es inal, and GI diges ions, we e analyzed in iplica e (n=
3), and he da a we e exp essed as he mean ±SD. S a is ical analyses
we e pe o med a 5% o he significance le el using he SPSS 27.0
Table 1. P epa a ion o Simula ed Gas ic Fluid (SGF) and Simula ed In es inal Fluid (SIF)
cons i uen
a
(mmol/L)
simula ed fluids KCl KH2PO4NaHCO3NaCl MgCl2(H2O)6(NH4)2CO3
simula ed gas ic fluid (SGF; pH 3) 6.90 0.90 25.00 47.20 0.10 0.50
simula ed in es inal fluid (SIF; pH 8) 6.80 0.80 85.00 38.40 0.13
a
KCl, po assium chlo ide; KH2PO4, po assium phospha e monobasic; NaHCO3, sodium bica bona e; NaCl, sodium chlo ide; MgCl2(H2O)6,
magnesium chlo ide hexahyd a e; and (NH4)2CO3, ammonium ca bona e.
Jou nal o Ag icul u al and Food Chemis y pubs.acs.o g/JAFC A icle
h ps://doi.o g/10.1021/acs.ja c.1c05373
J. Ag ic. Food Chem. 2021, 69, 14165−14175
14166
Figu e 1. Concen a ion (ng/g w) o HT and i s lipophenolic de i a i es in ex a i gin oli e oil (EVOO) and i s bioaccessible and esidual
ac ions (A) and bioaccessibili y esul ing om he indi idual gas ic and in es inal diges ions (B). N.d., no de ec ed. Ba s wi h a diffe en
lowe case le e indica e s a is ically significan diffe ences among ma ices a p< 0.05 acco ding o he analysis o a iance (ANOVA) and Tukey’s
mul iple ange es .
Figu e 2. Concen a ion (ng/g w) o HT and i s lipophenolic de i a i es in i gin oli e oil (VOO) and i s bioaccessible and esidual ac ions (A)
and bioaccessibili y esul ing om he indi idual gas ic and in es inal diges ions (B). N.d., no de ec ed. Ba s wi h a diffe en lowe case le e
among ma ices indica e s a is ically significan diffe ences among ma ices a p< 0.05 acco ding o he analysis o a iance (ANOVA) and Tukey’s
mul iple ange es .
Jou nal o Ag icul u al and Food Chemis y pubs.acs.o g/JAFC A icle
h ps://doi.o g/10.1021/acs.ja c.1c05373
J. Ag ic. Food Chem. 2021, 69, 14165−14175
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so wa e package (LEAD Technologies, Inc., Cha lo e, NC, USA).
Da a we e subjec ed o a one-way analysis o a iance (ANOVA). The
no mal dis ibu ion o he esiduals and he homogenei y o a iance
we e es ed wi h he Kolmogo o −Smi no and Le ene es s,
espec i ely. When s a is ical diffe ences we e iden ified, he a iables
we e compa ed using pos hoc analysis wi h Tukey’s mul iple ange
es .
■RESULTS AND DISCUSSION
Quan i a i e P ofile o Hyd oxy y osol-Fa y Acids in
Ex a Vi gin Oli e Oil and Vi gin Oli e Oil. To unde s and
he bioaccessibili y o HT-FAs, fi s , he quan i a i e p ofile o
unes e ified and es e ified HT in EVOO and VOO edible oils
was se up. As expec ed, he e en ion ime o HT-FAs
displayed a e e se co ela ion wi h he deg ee o unsa u a ion
o i s lipid pa . Thus, HT-ALA, HT-LA, and HT-OA we e
elu ed a 3.8, 4.0, and 4.4 min, espec i ely, which a e
consis en wi h p e ious expe imen al esul s
5
and o he
lipophenolic de i a i es such as y osol es e s.
16
A e his
ask, i was obse ed ha he amoun o unes e ified and
es e ified HT o bo h oli e oils was simila (Figu es 1 and 2)
bu diffe en om ha co esponding o FO, which exhibi ed
an absence o HT FAs (da a no shown).
In his way, he unes e ified HT was p esen as 874.74 and
934.09 ng/g w in EVOO and VOO, espec i ely. These
concen a ions we e in line wi h hose a ailable in he Phenol-
Explo e da abase (h p://phenol-explo e .eu/) on he HT
con en o EVOO (0.01−3.47 mg/100 g w) and VOO
(<0.01−7.43 mg/100 g w). Rega ding he HT-FAs, alues o
2.80, 1.54, and 20.00 ng/g we e eco ded o HT-ALA, HT-
LA, and HT-OA, espec i ely, in EVOO and 4.56, 2.59, and
11.90 ng/g in VOO (Figu es 1 and 2, co espondingly), which
a e in ag eemen wi h p e ious epo s on ma ching ood
ma ices and analy es,
5
al hough impo an ac o s such as
a ie y, geog aphical o igin, ipening s age, and p oduc ion and
ex ac ion p ocesses may play an impo an ole in lipophenol
con en .
21
Fa y Acid Composi ion o he Edible Oils Assessed.
As e e ed o be o e, i is wo h no ing ha he bioaccessibili y
o specific bioac i e compounds may be enhanced by he
con en o FAs in he ood ma ix h ough dispe sing ood
componen s in he diges i e ac o by p omo ing he
sec e ion o panc ea ic juice.
22
In his scena io, as one o he
objec i es o he cu en s udy was o desc ibe he
bioaccessibili y o he o HT-FAs, he quan i a i e p ofile o
a y acids o he edible oils unde conside a ion (EVOO,
VOO, and FO) was c ea ed, gi en i s cen al impo ance o he
o ma ion o he a ge bioac i e compounds, he HT-FAs. As
expec ed and acco ding o p e ious desc ip ions in he
li e a u e, he FA composi ion o he ege able oils conside ed
in he p esen wo k (EVOO, VOO, and FO) in ol ed
sa u a ed, monounsa u a ed, and polyunsa u a ed a y acids
(SFAs, MUFAs, and PUFAs, co espondingly).
23
Specifically,
he oily ma ices s udied con ained a o al o 13 indi idual FAs
(Table 2).
The SFAs de ec ed we e palmi ic (C16:0) and s ea ic
(C18:0) acids, which showed s a is ically significan diffe ences
be ween samples (p< 0.05), wi h EVOO being he plan oil
wi h he highes con en o palmi ic acid (12.07 g/100 g w) in
compa ison wi h VOO and FO (10.97 and 5.77 g/100 g w,
espec i ely). On he o he hand, FO p o ided he g ea es
concen a ion o s ea ic acid (4.40 g/100 g w) ha su passed
by 41.7%, on a e age, he con en s eco ded in EVOO and
VOO, which emained in simila bu lowe le els.
Mo eo e , significan diffe ences we e eco ded ega ding
he con en o palmi oleic and oleic acids (C16:1 n7 and
C18:1 n9, espec i ely) be ween he di e se edible oils
analyzed (p< 0.05), wi h oleic acid unde lined as ha ing he
mos abundan MUFA in all ege able oils conside ed (in he
ange o 19.97−80.10 g/100 g w). The highes concen a ion
o oleic acid was ound in VOO (80.10 g/100 g w) ollowed
by EVOO (3.2% lowe ) and FO (75.1% lowe ) (Table 2).
Finally, when analyzing he con en o PUFAs, linoleic
(C18:2 n-6) and α-linolenic (C18:3 n-3) acids we e ound a
he highes concen a ions in FO (15.27 and 53.53 g/100 g w,
espec i ely), whe eas EVOO and VOO displayed alues in a
simila lowe le el (4.27 and 0.62 g/100 g w, on a e age,
espec i ely) (Table 2).
The composi ion esul s ega ding SFAs, MUFAs, and
PUFAs a e in acco dance wi h in o ma ion p e iously epo ed
in he ame o s udies compa ing he FA composi ion o
se e al ypes o edible oils.
5,24
In his ega d, he FA p ofile,
oge he join ly wi h he p esence o HT in he ma ices
e e ed o in he s udy o HT lipophenolic de i a i es, would
p o ide a e y help ul choice o assessing he effec o GI
diges ion on he lipophenolic p ofile o ege able oils, which is
essen ial o disco e ing he biological in e es o hese
compounds.
Bioaccessibili y o Fa y Acid Es e s o Hyd oxy y -
osol om Ex a Vi gin Oli e Oil and Vi gin Oli e Oil. Fo
his fi s desc ip ion o he GI bioaccessibili y o he HT
lipophenols, an op imized and s anda dized s a ic me hod was
applied o he in i o simula ion o GI diges ion, which allows
mimicking he physicochemical condi ions in he GI ac , in
i o, as closely as possible.
19,20
The lipophenols o HT, HT-
ALA, HT-LA, and HT-OA we e analyzed in he h ee
ege able oils u ilized in he p esen s udy. FO was selec ed
Table 2. Concen a ion o Fa y Acids (g/100 g w) in he
Ex a Vi gin Oli e Oil (EVOO), Vi gin Oli e Oil (VO), and
Flaxseed Oil (FO) Analyzed in he P esen Wo k
edible oils
a y acids
a
EVOO VOO FO LSD (p< 0.05)
C14:0 0.03a
b
0.03a 0.03a <0.01
C16:0 12.07a 10.97b 5.77c 0.17
C16:1 n7 0.93a 0.83a 0.10b 0.09
C17:0 0.10a 0.10a 0.10a <0.01
C17:1 n8 0.10a 0.10a 0.10a <0.01
C18:0 2.50c 2.63b 4.40a 0.06
C18:1 n9 77.53b 80.10a 19.97c 0.17
C18:2 n6 4.43b 4.10c 15.27a 0.09
C18:3 n3 0.63b 0.60b 53.53a 0.09
C20:0 0.37a 0.60a 0.33a 0.09
C20:1 0.30a 0.30a 0.10b <0.01
C22:0 0.10a 0.10a 0.10a <0.01
C24:0 0.47a 0.10b 0.10b 0.06
a
C14:0: my is ic acid; C16:0: palmi ic acid; C16:1 n7: palmi oleic
acid; C17:0: hep adecanoic acid; C17:1 n8: hep adecenoic acid;
C18:0: s ea ic acid; C18:1 n9: oleic acid; C18:2 n6: linoleic acid;
C18:3 n3: alpha-linolenic acid; C20:0: a achidic acid; C20:1:
eicosenoic acid; C22:0: behenic acid; and C24:0: lignoce ic acid.
b
Means (n= 3) wi hin a ow wi h diffe en lowe case le e s a e
significan ly diffe en a p< 0.05 acco ding o he analysis o a iance
(ANOVA) and Tukey’s mul iple ange es .
Jou nal o Ag icul u al and Food Chemis y pubs.acs.o g/JAFC A icle
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as a nega i e con ol as i does no con ain ei he HT o i s
lipophenolic de i a i es acco ding o ou p e ious wo k.
5
These molecules we e also assessed in he diges a es
ob ained a e in i o gas ic, in es inal, and GI diges ion o
he ege able oils men ioned abo e. In his ega d, bo h he
uppe oily o esidual phase (RF) and he lowe micella o
bioaccessible ac ion (BF) esul ing om diges ion we e
collec ed and analyzed o e ie e accu a e in o ma ion on he
heo e ical and po en ial bioa ailable ac ions o hese new
lipophilic phenolic compounds, and also in e con e sions
be ween unes e ified HT and i s es e s de i a i es, which
would condi ion u u e in i o esea ch aimed a add essing he
ac ual bioa ailabili y o HT ( ee and es e ified).
The in i o GI diges ion pe o med on EVOO (Figu e 1A)
and VOO (Figu e 2A) e ealed ha ALA-, LA-, and OA-based
lipophenols o es e ified HT we e p esen in he BF. The
concen a ions o HT-FAs co esponding o EVOO we e 3.68,
4.04, and 239.41 ng/g w, while o he VOO samples, hese
we e 8.22, 1.80, and 92.00 ng/g w o HT-ALA, HT-LA, and
HT-OA, espec i ely, e idencing ha he es e ified o ms o
HT we e bioaccessible. Howe e , he a e o bioaccessibili y
depended on bo h he specific FA-based es e ifica ion and he
composi ional ea u es o he ood ma ix, as al eady epo ed
o ee HT pha macokine ics and bioa ailabili y, in i o, a e
inges ion o diffe en oily ma ices.
25
These esul s a e in good ag eemen wi h p e ious epo s
on he bioaccessibili y o o he es e s o HT, namely,
phospha idyl-hyd oxy y osol (PHT, phospholipid de i a i es
o HT wi h phospha idylcholine).
17
In hei wo k, Ma in e al.
desc ibed ha he po ion o HT wi hin he BF, ega ding he
es e ified o m (PHT), was significan ly highe ela i e o
unes e ified HT.
17
In addi ion, al hough a lowe concen-
a ions han in BF, bo h unes e ified HT and es e ified HT
we e de ec ed in RF (Figu es 1A and 2A). Jus as wi h PHT,
he amphiphilic p ope ies o HT es e ified wi h FAs could
cause i s dispe sion in he aqueous media, which in u n
indi ec ly enabled he dispe sion o he ehicula ed HT.
17
When analyzing he bioaccessibili y a e o HT and i s
lipophenol de i a i es, i was ound ha he unes e ified o m
p esen ed a bioaccessibili y o 70.9 and 15.4% o EVOO and
VOO, espec i ely. These esul s a e in line wi h p e ious
desc ip ions conce ning an impo an loss o HT in he
duodenal compa men (∼50%, on a e age) e ie ed om
alpe ujo (oli e-mill was e) diges ions.
26
These au ho s
desc ibed ha when diges ing only he a ge bioac i e
compounds in analy ical sol en s (wi hou he p esence o
addi ional ood cons i uen s, such as s a ch, casein, and fibe ),
he bioaccessibili y o HT was enhanced by ∼20%. This effec
could be due o he p esence o he e e ed mac omolecules,
which can bind polyphenols and e ain hem wi hin he ood
ma ix. Hence, se e al au ho s ha e emphasized he signifi-
cance and complexi y o phenolic compound in e ac ions
wi hin he ood ma ices associa ed o hei bioaccessibil-
i y.
26,27
In ela ion o he concen a ions o lipophenols in EVOO,
he concen a ion o HT-FAs in BF inc eased 1.3-, 3.0-, and
11.9- old o HT-ALA, HT-LA, and HT-OA, espec i ely
(Figu e 1A). This ac may be due o he in e ac ions be ween
ee HT and FAs, whe e HT could ac as a nucleophilic
compound able o ap ALA, LA, and OA p esen in he oily
ma ix o o m he co esponding es e s ia enzyma ic o
chemical means unde GI condi ions,
28
he e o e exe ing
″nega i e″effec s on ee HT bioaccessibili y bu amelio a ing
he concen a ion o he es e ified o ms o HT in he
diges a e. A simila beha io was obse ed o VOO as a esul
o he GI diges ion, which esul ed in a 1.8- and 7.7- old ise in
concen a ion o HT-ALA and HT-OA, espec i ely, in he
BF when compa ed o hose ound in VOO, wi h he excep ion
o HT-LA, whose bioaccessibili y dec eased up o ∼70%, on
a e age (Figu e 2A). In addi ion, i should also be s essed ha
he mos abundan HT-FA in he RF was HT-OA, which
inc eased 2.0- and 5.6- old in compa ison wi h he amoun s
ound in EVOO and VOO, espec i ely (Figu es 1A and 2and
Table 2). This diffe en ial occu ence o HT-FAs om EVOO
and VOO a e GI diges ion could be owed o he in e ac ions
o hese compounds wi h he ood ma ix componen s such as
amino acids and p o eins, in spi e o hei low con en (0.07−
2.4 mg/kg). The p esence o hese componen s depends on
he ipening s age o oli e cul i a , among o he ac o s,
29,30
and may gene a e a y acid-binding p o ein eac ions
31,32
ha
can hampe he bioaccessibili y o hese compounds.
Con en ionally, he RF has no been conside ed o he
e alua ion o he bioaccessibili y o phenolic compounds a e
he diges ions. Howe e , his ac ion is o high ele ance
because o i s high concen a ion o polyphenols ha a e no
abso bed in he small in es ine, which can be me abolized by
he gu mic obio a,
33
gi ing ise o addi ional bioac i e
de i a i es ha could also con ibu e o he heal hy a ibu es
o oods. Thus, bo h ac ions (BF and RF) esul ing om GI
diges ion may be esponsible o he posi i e heal h effec s
a ibu ed o plan -based oods, including edible oils.
34
In his
ma e , he capaci y o he in es inal mic obio a o modi y he
quan i a i e phy ochemical p ofile o he diges ion p oduc s
and he eby he bioa ailabili y/abso p ion capaci y o bioac i e
compounds p esen in oli e and oli e oils ha ha e no been
abso bed in he small in es ine is ema kable, and as a esul ,
he subs a e o hese me abolic eac ions (nonabso bed
phenolics) should no be dis ega ded. In ac , he gu
mic obio a plays a p ominen ole in he bio ans o ma ion
o bo h es e ified and aglycone o ms o phenolic compounds,
bu he me abolism o lipophenols by he gu mic obio a is s ill
unclea . To da e, he e is only one s udy ha e alua ed he
ecal mic obial me abolism o lipophenols u ilizing an in i o
e men a ion model as well as he in i o abso p ion and
plasma pha macokine ics o y osol es e s in a s, whe e
s anda ds o y osol and y osol es e s we e adminis a ed in
he d inking wa e .
35
The au ho s concluded ha y osol es e
de i a i es ac ed longe in i o han na i e y osol by obse ing
a second abso p ion peak in pha macokine ic p ofiles, possibly
due o mic obio a deg ada ion, wi h he es e ified molecules
exhibi ing an imp o ed bioa ailabili y as compa ed o ha o
ee y osol. The incuba ion o y osol es e s in he
e men a ion solu ion libe a ed ee y osol in a ime-
dependen manne , indica ing he occu ence o hyd olysis.
35
Howe e , his finding should be es ed wi h a s ed wi h
na u al sou ces o hese lipophenols, no wi h s anda ds, and in
he na u al concen a ions in which hey a e ound.
Con ibu ion o he Gas ic and In es inal Diges ion
Phases o he Bioaccessibili y o HT-FAs. To unde s and
he impac o he diges ion p ocess on he bioaccessibili y o
HT-FAs, he gas ic diges a es om EVOO and VOO we e
also analyzed (Figu es 1B and 2), which p o ided aluable
in o ma ion on he elease o he a ge lipophenols on each
diges ion s age. Upon his analysis, i was obse ed ha
unes e ified HT and i s lipophenol de i a i es we e p esen a
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high concen a ions in he RF ob ained om he gas ic
diges ion o bo h EVOO and VOO.
When e alua ing he impac o gas ic diges ion on he
EVOO lipophenols, low concen a ions o HT and HT-FAs
we e obse ed in he BF o all indi idual analy es (no
de ec ed, no de ec ed, and 1.77 and 0.33 ng/g o HT-ALA,
HT-LA, HT-OA, and HT) (Figu e 1B). Simila esul s we e
ob ained ega ding VOO (no de ec ed and 0.13, 0.82, and
0.52 ng/g o HT-ALA, HT-LA, HT-OA, and HT) (Figu e
2B).
The low concen a ions eco ded could be due o he labili y
o lipophenols agains he physicochemical condi ions and he
enzyma ic ac i i y p ope o he gas ic diges ion, which would
be associa ed wi h a low elease o s abili y o he a ge
analy es. This would be consis en wi h p e ious epo s on he
bioaccessibili y o HT in oli e lea es,
36
u he suppo ing he
influence o he ood ma ix, he gas ic enzymes, o acidic
condi ions p e iously desc ibed.
37
None heless, he de elop-
men o in es inal diges ion on he chyme p o ided a high
concen a ion o ee HT in he BF. This could be due o he
hyd olysis o oleu opein and i s aglycone du ing in es inal
diges ion ha depends on he lipase ac i i y, hus leading o
he appea ance o HT, as p e iously s a ed by Rocche i e al.
38
In e es ingly, as a esul o in es inal diges ion, he concen-
a ion o HT-FAs significan ly inc eased in he BF o e he
en i e GI p ocess, en a i ely due o he pu a i e enzyma ic
ac i i y o panc ea ic lipase ha may ca alyze bo h he
hyd olysis o syn hesis o es e s.
39
As a as we know, o da e, mos esea ch on he
bioaccessibili y o lipophenols has been pe o med on solu ions
o au hen ic s anda ds exposed o he GI physicochemical
condi ions. The e o e, despi e he e e ed a ibu es o he
syn hesis o lipophenols as a esul o he diges i e enzyma ic
ac i i y, addi ional wo ks ha e e idenced ha HT de i a i es,
such as hyd oxy y osol ace a e, unde go in ense hyd olysis
du ing in es inal diges ion mainly due o he enzyma ic ac i i y
o panc ea in.
14
Simila ly, i has been epo ed ha y osol acyl
es e s a e hyd olyzed by panc ea ic lipase o p oduce ee
y osol.
15,16
Mo eo e , he s abili y o es e a ol es e s wi h
cap ylic acid du ing diges ion is nega i ely co ela ed wi h he
deg ee o subs i u ion o he FA moie y, gi en ha a e 120
min o incuba ion a 37 °C, ∼54 and ∼11% o monoes e s and
dies e s a e hyd olyzed, espec i ely, while no hyd olysis o he
ies e s has been no iced.
40
Howe e , hese esul s ha e been
associa ed wi h an ″a ificially″ob ained (only wi h chemically
o enzyma ically syn hesized s anda ds) highe bioaccessibili y
ela i e o he effec s ha occu du ing he GI diges ion o
oods because hese models do no allow moni o ing he
ex ac i e capaci y o he p ocess, he in e con e sion be ween
molecules, o he de no o syn hesis o he a ge analy es ( om
he complex pool o molecules p esen in he ood ma ix).
This gap in knowledge has been o e whelmed by a s udy on
he bioaccessibili y o HT wi h alpe ujo (solid byp oduc o
oli e oil ex ac ion). This s udy desc ibed a lowe bioacces-
sibili y o HT when applying GI diges ion on a ege able
ma ix due o he p esence o fibe s and suga s ha could
in e ac wi h HT and he diges i e enzymes, modula ing he
bioaccessibili y a e.
26
Acco dingly, he assessmen o he
bioaccessibili y o HT o HT-FAs needs o include a ood
ma ix o ob ain obus and nonspecula i e conclusions.
The ele ance o he in es inal diges ion phase o he
p esence o HT lipophenols in he p oduc o he GI diges ion
p omp ed us o explo e he effec o he in es inal diges ion
conside ed indi idually on he in ac ood ma ix, a oiding he
unce ainly de imen al effec ha could be occu ing du ing
he gas ic s age (Table 3). Thus, when he in es inal diges ion
was pe o med on bo h EVOO and VOO (wi hou he
p e ious gas ic phase), he concen a ion o HT lipophenols
obse ed in he BF was highe ela i e o ha ob ained
ollowing he physiological wo kflow (Table 3). The con en o
HT-ALA, HT-LA, and HT-OA in BF o he in es inally
diges ed EVOO was ∼64-, ∼8-, and ∼15- old highe ,
espec i ely, in compa ison wi h he BF ob ained a e a
comple e GI diges ion (gas ic plus in es inal diges ions). This
end was simila o VOO, whe e he con en o lipophenols in
he BF inc eased ∼4-, ∼3-, and ∼13- old o HT-ALA, HT-LA,
and HT-OA, espec i ely, ela i e o BF ob ained om he
sequen ial gas ic and in es inal diges ions (Table 3).
On he con a y, he highes concen a ion o unes e ified
HT was ound in he BF om in es inal diges ions de eloped
on gas ic diges a es o EVOO (619.88 ng/g w) in compa ison
wi h he di ec in es inal diges ion o he in ac ood ma ix
(55.0% lowe , p< 0.001) (Table 3), while ega ding VOO, no
s a is ically significan diffe ences we e ound, ein o cing he
Table 3. Concen a ion (ng/g w) o he Bioaccessible and Residual F ac ions (BF and RF, Respec i ely) o HT and I s
Lipophenolic De i a i es as a Resul o he Only In es inal Diges ion on he Diffe en Subs a es (Ex a Vi gin Oli e Oil,
Vi gin Oli e Oil, and Thei Gas ic Diges a es)
concen a ion (ng/g w)
subs a e o he in es inal diges ion
gas ic diges a e oli e oil
oil ma ix analy e
a
oli e oil BF RF BF RF
EVOO
a
HT 874.47 ±41.88a
b
619.88 ±144.42b 64.46 ±12.70d 279.05 ±63.95c 542.00 ±34.15b
HT-ALA 2.80 ±0.95b 3.68 ±2.30b 0.61 ±0.06b 235.96 ±23.77a 0.64 ±0.02b
HT-LA 1.54 ±0.49bc 4.04 ±1.97b N.d. 34.19 ±1.61a N.d.
HT-OA 20.00 ±8.11c 237.64 ±76.94b 12.68 ±2.72c 3599.94 ±107.20a 102.35 ±0.41bc
VOO HT 934.09 ±29.53a 143.76 ±25.08bc 39.58 ±1.27c 207.75 ±131.67b 251.66 ±3.38b
HT-ALA 4.56 ±1.52b 8.22 ±1.79b 5.12 ±1.25b 32.44 ±5.17a 4.59 ±0.54b
HT-LA 2.59 ±0.46b 1.98 ±0.44b N.d. 5.87 ±1.51a N.d.
HT-OA 11.90 ±4.90b 91.18 ±2.97b 63.90 ±4.21b 1205.44 ±160.57a 85.81 ±7.79b
a
HT, hyd oxy y osol; HT-ALA, hyd oxy y osol es e ified wi h α-linolenic acid; HT-LA, hyd oxy y osol es e ified wi h linoleic acid; and HT-OA,
hyd oxy y osol es e ified wi h oleic acid.
b
Da a a e shown as means ±SD (n= 3). Da a wi hin a single ow ollowed by he same lowe case le e
a e no significan ly diffe en a p< 0.001 acco ding o he analysis o a iance (ANOVA) and Tukey’s mul iple ange es . N.d., no de ec ed.
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ele ance o he ood ma ix composi ion o he final
bioaccessibili y esul s. The dis inc beha io in he diges i e
hallma k o HT and i s lipophenol de i a i es could be due o
he spa ing solubili y o HT in oli e oil and i s consequen
loca ion in he aqueous phase (>99%), while, boos ed by he
amphiphilic na u e, HT-FAs a e dis ibu ed be ween bo h he
oily and aqueous phases.
41
The eby, as expec ed, HT-OA was he mos abundan
lipophenol in he ood ma ix and he GI diges a es (Table 3).
This lipophenol should be o med h ough he enzyma ic
es e ifica ion o HT wi h OA, which is he mos abundan FA
in he oily ma ices conside ed in he p esen wo k (EVOO
and VOO) (Table 2). Based on he findings om he
o ma ion o HT-FAs unde in es inal diges i e condi ions, he
use o p o ec o s o HT agains he gas ic condi ions migh be
o in e es o ake ad an age o he bioac i i ies o HT-FAs
om biological o echnological poin s o iew.
Rela i e Influence o he In es inal Enzymes on he
Gene a ion o Hyd oxy y osol Lipophenols. Gi en he
ele ance o he in es inal s age o he bioaccessibili y o HT
and i s lipophenolic de i a i es, he sepa a e and join effec s o
in es inal enzymes (panc ea in and panc ea ic lipase) on hese
compounds we e u he in es iga ed o gain u he unde -
s anding o hei ole on he elease o HT-FAs in o he
in es inal lumen and hei s abili y. Wi h his objec i e, EVOO
was diges ed using SIF con aining indi idual and combined
in es inal enzymes a 37 °C o 120 min. The incuba ion o
EVOOin heSIFwi hou anydiges i eenzymewas
conside ed as a nega i e con ol. This app oach allowed
e ie ing c i ical in o ma ion on hei ela i e influence on he
bioaccessibili y o HT-FAs.
Lipases ( iacylglyce ol acyl hyd olases, EC 3.1.1.3.) a e
bioca alys s wi h a high selec i i y and s abili y ha ca alyze
hyd olysis, es e ifica ion, anses e ifica ion, and alcoholysis
eac ions.
42
To da e, lipases ha e been applied o he
p oduc ion o FA de i a i es by he ag o- ood and
nu aceu ical indus ies because o hei po en ial uses as
fla o ing es e s, a y acid es e s o an ioxidan s, and s uc u ed
lipids wi h high egio- and s e eo-selec i i ies.
43
Up o now, he
biosyn hesis o HT-FAs has been done using comme cial
enzymes on pu e s anda d compounds.
39
These eac ions ha e
been epo ed as being closely dependen on he acyl chain
leng h and he deg ee o unsa u a ion o FAs ha affec he
con e sion yield o he es e ifica ion p ocess due o diffe ences
in hei spa ial configu a ion.
44,45
In he cu en wo k, he gene a ion o HT es e s wi h an
equi alen ca bon alkyl chain (C18) bu wi h diffe en deg ees
o unsa u a ion, C18:1, C18:2, and C18:3 co esponding o
HT es e ified wi h OA, LA, and ALA, espec i ely, was
moni o ed, o he fi s ime, a e he in es inal diges ion o
EVOO wi h indi idual and combined in es inal enzymes
(panc ea in and panc ea ic lipase). The con e sion yield o
HT-FAs was mo e p onounced when he eac ion was
ca alyzed only by panc ea in han when panc ea ic lipase was
Figu e 3. Rep esen a i e UHPLC-ESI-QqQ-MS/MS-ex ac ed ion ch oma og ams co esponding o he bioaccessible lipophenols o
hyd oxy y osol (HT-ALA, hyd oxy y osol-α-linolenic acid; HT-LA, hyd oxy y osol-linoleic acid; and HT-OA, hyd oxy y osol-oleic acid) in
diges a es ob ained a e applying diffe en enzyme combina ions on ex a i gin oli e oil du ing simula ed in i o in es inal diges ion using he
quan ifica ion MRM ansi ions. CPS, cha ges pe second.
Table 4. Concen a ion (ng/g w) o Hyd oxy y osol Es e ified wi h Fa y Acids a e In es inal Diges ion o Ex a Vi gin Oli e
Oil wi h Diffe en Combina ions o Diges i e Enzymes
analy e
a
con ol (no enzyme) panc ea in panc ea ic lipase panc ea in + panc ea ic lipase
HT-ALA 4.58 ±1.44b
b
228.85 ±14.87a 193.94 ±27.88a 173.40 ±4.55
a
HT-LA 0.68 ±0.17c 25.77 ±2.25a 21.33 ±7.50ab 14.54 ±0.42b
HT-OA 31.83 ±2.02c 4853.34 ±69.57a 2235.37 ±244.55b 2876.17 ±315.30b
a
HT-ALA: hyd oxy y osol es e ified wi h α-linolenic acid; HT-LA: hyd oxy y osol es e ified wi h linoleic acid; and HT-OA: hyd oxy y osol
es e ified wi h oleic acid.
b
Mean ±s anda d de ia ion (n= 3) ollowed by he same lowe case le e a e no s a is ically significan a p< 0.05
acco ding o he analysis o a iance (ANOVA) and Tukey’s mul iple ange es .
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p esen (alone o in combina ion wi h panc ea in) (Figu e 3).
HT-OA was he lipophenol wi h he highes concen a ion in
he in es inal diges a e, independen ly o he enzymes o hei
combina ion applied (4853.34, 2235.37, and 2876.17 ng/g
wi h panc ea in, panc ea ic lipase, and combina ions o he wo
enzymes, espec i ely), and showed no s a is ical diffe ences
be ween panc ea ic lipase and he combina ion o bo h
enzymes (p> 0.05) (Table 4). Rega ding HT-LA and HT-
ALA, he final concen a ions achie ed in he in es inal
diges a e, as well as he diffe ences be ween enzyma ic
condi ions, a ied o a lesse ex en ela i e o HT-OA (Figu e
3and Table 4). This may be due o he deg ee o unsa u a ion
o he FAs in ol ed in he es e ifica ion o HT, which may
affec hei con o ma ion and he eby he eac ion ime, as
s a ed by p e ious esea ch.
44
The gene a ion o HT-FAs has al eady been desc ibed in
oily ood ma ices a e o i ying ood ma ices wi h HT,
5
as
well as in oli e oil byp oduc s du ing p ocessing and s o age,
while his was no de ec able in in ac oli es.
46
A hypo hesis
suppo ing hese findings sugges s ha HT es e s could be
o med independen ly as a esul o he es e ase ac i i y om
lipase-posi i e yeas s, which a e p esen in plan -based oods.
Howe e , he molecula mechanisms esponsible o he de
no o o ma iono lipophenols,aswellas heop imal
condi ions needed o he success ul de elopmen o such
eac ions, emain unclea . Likewise, ecen ly, he o ma ion o
y osol es e s, pa icula ly wi h OA, has been desc ibed,
iden i ying c ushing and kneading p ocesses when enzymes a e
ac i a ed and he eac ions igge ed.
47
Wi hin his ame, he
p esen s udy con ibu es o he unde s anding o he
gene a ion o lipophilic hyd oxy y osyl es e s du ing GI
diges ion in oily ma ices, wi h a discussion o some possible
scena ios being necessa y.
The diffe en efficiency ega ding he o ma ion o he
sepa a e lipophenols (HT-ALA, HT-LA, and HT-OA) may be
associa ed wi h he deg ee o unsa u a ion, which in u n
modifies he FA spa ial configu a ion and he lipase selec i i y.
Mo eo e , he concen a ion o he p ecu so s in he ood
ma ix (bo h unes e ified HT and FAs) seems o be c i ical o
he syn hesis o HT-FAs du ing diges ion as well since he
con en o ee OA in EVOO and VOO was, on a e age, ∼99
and ∼95% in compa ison wi h he ALA and LA concen a ion,
espec i ely (Table 2). This is in ag eemen wi h ecen
esea ch, which u ilized nanopa icles wi h immobilized lipase
om The momyces lanuginosus and employed oli e lea aqueous
ex ac o he syn hesis o bioac i e hyd oxy y osyl FA es e s.
The s udy epo ed ha he con e sion yields (%) o OA
(C18:1), LA (C18:2), and my is ic acid (C14:0) we e 55.0,
50.4, and 64.3%, espec i ely, es ablishing ha he numbe o
double bonds had a di ec co ela ion wi h he molecula
s uc u e o he FA, which, as a esul , would affec he ad ance
o he acyla ion eac ions.
45
On he con a y, addi ional s udies
wo king wi h he pu e s anda d o long-chain unsa u a ed FAs
indica ed ha he e was no ela ionship be ween chain leng h
and con e sion, as he highes con e sion was ob ained wi h
eicosapen aenoic acid (EPA) (68.0%) ela i e o OA (46.0%).
The a ia ion in hese findings may be caused by he selec i i y
o lipase o he FAs, wi h he deg ee o unsa u a ion being a
mo e ele an ac o a he han FA chain leng h.
39
In addi ion,
lipase pe o ms he e ogeneous eac ions, and i s ca aly ic
ac i i y is maximum when he enzyme is adso bed a an oil−
wa e in e ace. Gene ally, long-chain es e s a e mainly
dispe sed in he oil d ople , while sho -chain es e s a e
ound mos ly in he wa e d ople .
48
Fo lipophenols, a cu off
effec ha in ol es a oo-sho o a oo-long hyd ophobic chain
does no ensu e a p ope in e acial loca ion, which in u n
does no esul o an op imal lipase ac i i y,
49
and he cu off
effec is simply a na u al consequence o he diffe en ial
solubili y o an ioxidan s in he aqueous, in e acial, and oil
egions o an emulsion.
50
Fu he mo e, o he phenolic acyl es e s, such as y osol-FAs,
ha e shown a sus ained- elease beha io o ee y osol
molecules du ing he ime-consuming diges ion p ocess
16
ha could imp o e he bioac i i y scope o polyphenols
because o ex ended ac ions. In his sense, he cu en wo k
b ings new knowledge and s a egies o he design o slow-
elease o mula ions o HT-FAs ha could enhance he
phenolic loading cycle ime by p olonging he e minal hal -
li e and he eby leng hening hei bioac i i y. This is
pa icula ly impo an , as se e al biological p ope ies ha e
been ecen ly sugges ed o hese compounds, namely,
an idiabe ic, p oli e a i e, and an ioxidan capaci ies.
47,51
In
addi ion, i has been demons a ed ha he es e ifica ion o
phenolic compounds did no comp omise he bioac i i y o he
na i e molecule bu ins ead may enhance i s bioa ailabili y and
expand i s applica ion domains.
52
As concluding ema ks, he p esen s udy p o ides new
e idence on he bioaccessibili y o lipophenolic de i a i es o
HT om di e se oli e oils (EVOO and VOO), showing ha
hey a e bioaccessible a e pe o ming a simula ion o GI
diges ion unde in i o condi ions, depending la gely on he
ype o FA es e i ying HT as well as he ood ma ix. Mo e
impo an ly, he gene a ion o HT-FAs du ing in es inal
diges ion was epo ed in he p esen wo k, as well as he
iden ifica ion o he enzymes esponsible o mos o he
syn hesis iden ified (panc ea in). These findings a e o c i ical
ele ance om a clinical and echnological poin o iew
because o he po en ial biological effec s ha could be
expec ed om de no o syn hesized lipophenols, which could
ha e a c i ical impac on bo h he hal -li e o plan -based oods
and he biological benefi s al eady desc ibed on hose oods
and oods uffs con aining he componen s o lipophenols (e.g.,
HT and FAs). Indeed, he majo ou comes e ie ed sugges
ha he use o oily ma ices and hei byp oduc s as po en ial
unc ional ing edien s, oods, supplemen s, o cosme ics, o
heal h p omo ion and disease isk educ ion, should ake in o
conside a ion he o ma ion o new lipophenols du ing
diges ion and, consequen ly, he ans o ma ion o he final
bioac i e scope and he effec s wi hin he ame o specific
pa hophysiological p ocesses. Ul ima ely, u he assays con-
ce ning he bioa ailabili y o HT-FAs a e s ill needed, wi h a
high impo ance placed on alida ing whe he lipophenol
molecules would mig a e ac oss he epi helial ba ie , hei
me aboliza ion ci cula ing eac ion, he anspo e s o hese
compounds, and he me abolic e en s ha affec hei final
shel -li e in he o ganism and bioac i i y in he sepa a e issues
and cell ypes.
■AUTHOR INFORMATION
Co esponding Au ho s
A

ngel Gil-Izquie do −Depa men o Food Science and
Technology, Resea ch G oup on Quali y, Sa e y and
Bioac i i y o Plan Foods, CEBAS-CSIC, Uni e si y Campus
o Espina do, Mu cia 30100, Spain; o cid.o g/0000-
0001-7646-0386; Phone: +34968396200;
Email: [email p o ec ed]
Jou nal o Ag icul u al and Food Chemis y pubs.acs.o g/JAFC A icle
h ps://doi.o g/10.1021/acs.ja c.1c05373
J. Ag ic. Food Chem. 2021, 69, 14165−14175
14172
Sonia Medina −Depa men o Food Science and Technology,
Resea ch G oup on Quali y, Sa e y and Bioac i i y o Plan
Foods, CEBAS-CSIC, Uni e si y Campus o Espina do,
Mu cia 30100, Spain; o cid.o g/0000-0002-7231-6480;
Phone: +34968396200; Email: [email p o ec ed]
Au ho s
Ca olina Alemán-Jiménez −Depa amen o de En e me ía,
Uni e sidad Ca ólica de Mu cia, Mu cia 30107, Spain
Raul Domínguez-Pe les −Depa men o Food Science and
Technology, Resea ch G oup on Quali y, Sa e y and
Bioac i i y o Plan Foods, CEBAS-CSIC, Uni e si y Campus
o Espina do, Mu cia 30100, Spain; o cid.o g/0000-
0001-6232-712X
Juana I. Gallego-Gómez −Depa amen o de En e me ía,
Uni e sidad Ca ólica de Mu cia, Mu cia 30107, Spain
Agus ín Simonelli-Munoz −Depa amen o de En e me ía,
Fisio e apia y Medicina. Uni e sidad de Alme ía, Alme ía
04120, Spain
Espé ance Moine −Ins i u des Biomolécules Max Mousse on
(IBMM), UMR 5247, CNRS, Uni e si y o Mon pellie ,
Mon pellie 34093, F ance
Thie y Du and −Ins i u des Biomolécules Max Mousse on
(IBMM), UMR 5247, CNRS, Uni e si y o Mon pellie ,
Mon pellie 34093, F ance
Céline C aus e −Ins i u des Biomolécules Max Mousse on
(IBMM), UMR 5247, CNRS, Uni e si y o Mon pellie ,
Mon pellie 34093, F ance; o cid.o g/0000-0002-5714-
8749
Fede ico Fe e es −Depa men o Food Technology and
Nu i ion, Molecula Recogni ion and Encapsula ion (REM)
G oup, Uni e sidad Ca ólica de Mu cia, UCAM, Mu cia
30107, Spain
Comple e con ac in o ma ion is a ailable a :
h ps://pubs.acs.o g/10.1021/acs.ja c.1c05373
Au ho Con ibu ions
#
C.A.-J. and R.D.-P. wo au ho s con ibu ed equally o he
p esen wo k.
Funding
This wo k was pa ially unded by he ″FundacionSe
neca de la
Region de Mu cia″G upo de Excelencia 19900/GERM/15.
S.M. was suppo ed by a Pos doc o al Con ac (Saa ed a-
Faja do 21078/SF/19) om he Agency o Science and
Technology o he Region de Mu cia−″FundacionSe
neca″
(Spain).
No es
The au ho s decla e no compe ing financial in e es .
■ACKNOWLEDGMENTS
The au ho s hank he company Olimend os S.L. (Mu cia,
Spain) o he supply o ex a i gin oli e oils.
■ABBREVIATIONS
BF, bioaccessible ac ion; BHA, bu yla ed hyd oxyanisole;
DMSO, dime hyl sul oxide; EVOO, ex a i gin oli e oil; FAs,
a y acids; FO, flaxseed oil; HT, hyd oxy y osol; HT-ALA,
hyd oxy y osol es e ified wi h α-linolenic acid; HT-FA,
hyd oxy y osol es e ified wi h a y acid; HT-LA, hyd oxy y -
osol es e ified wi h linoleic acid; HT-OA, hyd oxy y osol
es e ified wi h oleic acid; MRM, mul iple eac ion moni o ing;
MUFAs, monounsa u a ed a y acids; NMR, nuclea magne ic
esonance; PUFAs, polyunsa u a ed a y acids; RF, esidual
ac ion; SFAs, sa u a ed a y acids; SGF, simula ed gas ic
fluid; SIF, simula ed in es inal fluid; VOO, i gin oli e oil
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