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Article
Changes in fatty acid profile and phospholipid molecular species composition of human erythrocytes membranes after hybrid palm and extra virgin olive oil supplementation D. Pacetti, R. Gagliardi, M. Balzano, N. G. Frega, M. L. Ojeda, M. Borrero, A. Ruiz, and P. Lucci J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.6b01903 • Publication Date (Web): 17 Jun 2016 Downloaded from http://pubs.acs.org on June 19, 2016
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Journal of Agricultural and Food Chemistry is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.
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Journal of Agricultural and Food Chemistry is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.
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Journal of Agricultural and Food Chemistry
200 180 160 140 120 100
80 60 40 20 0 C16:0
C18:0
C18:1
C18:2
Tocols mg per 100 g of oil
%
EVOO
HPO
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Total Total phenolic content ppm phenolic content
Journal of Agricultural and Food Chemistry
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Positive ions; base peak
Relative Abundance
100 80
[M+H]+=m/z 760.4
PC (16:0/18:1)
60 40 20
0 0.0
[M+H]+=m/z 718.1
PE (16:0/18:1) Sph (16:0)
[M+H]+=m/z 703.3 0.5
1.0
1.5
2.0
2.5
3.0
3.5 4.0 4.5 Time (min)
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5.0
5.5
6.0
6.5
7.0
7.5
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Journal of Agricultural and Food Chemistry
Positive ions; base peak
Relative Abundance
100 80
[M+H]+=m/z 760.4
PC (16:0/18:1)
60 40 20 0 0.0
[M+H]+=m/z 718.1
PE (16:0/18:1) Sph (16:0)
[M+H]+=m/z 703.3 0.5
1.0
1.5
2.0
2.5
3.0
3.5 4.0 4.5 Time (min)
5.0
5.5
6.0
6.5
7.0
7.5
Hybrid palm oil
Changes in RBC membrane: Phospholipid molecular species composition
Supplementation 25 mL/day - 3 months
Omega-3 Index
Extra virgin olive oil
Red blood cell membrane ACS Paragon Plus Environment
Lipophilic Index
Journal of Agricultural and Food Chemistry
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Changes in fatty acid profile and phospholipid molecular species composition of
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human erythrocytes membranes after hybrid palm and extra virgin olive oil
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supplementation.
4 5 6
D. Pacettia, R. Gagliardia, M. Balzanoa, N. G. Fregaa, M. L. Ojedab, M. Borrerob, A. Ruizc, P.
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Luccid*
8 9 10 11 12 13 14
a
15
Italy
Department of Agricultural, Food, and Environmental Sciences, Marche Polytechnic University, Via Brecce Bianche,
60131, Ancona, Italy b
Department of Nutrition and Biochemistry, Faculty of Sciences, Pontificia Universidad Javeriana, KR 7 # 40-62, Bogota
D.C., Colombia c
Department of Internal Medicine, School of Medicine, Pontificia Universidad Javeriana, KR 7 # 40-62, Bogota D.C.,
Colombia d
Department of Agri-food, Environmental and Animal Sciences, University of Udine, Via Sondrio 2/a. 33100 Udine,
16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37
*
Corresponding author: Department of Agri-food, Environmental and Animal Sciences, University of Udine, via Sondrio 2/a, 33100 Udine, Italy; Tel. +3904325558170– Email:
[email protected];
[email protected] 1 ACS Paragon Plus Environment
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ABSTRACT
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The present work aimed to evaluate and compare, for the first time, the effects of extra virgin olive
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oil (EVOO) and hybrid palm oil (HPO) supplementation on fatty acid profile and phospholipid (PL)
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molecular species composition of human erythrocyte membranes.
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Results supported the effectiveness of both HPO and EVOO supplementation (3-month, 25mL/day)
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in decreasing the lipophilic index of erythrocytes with no significant differences between HPO and
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EVOO groups at month 3. On the other hand, the novel and rapid UPLC/MS/MS method used for PL
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analysis reveals an increase in phosphatidylcholine and phosphatidylethanolamine species esterified
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with polyunsaturated fatty acids.
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The present work demonstrated the ability of both EVOO and HPO to increase the degree of
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unsaturation of erythrocyte membrane lipids with an improvement in the membrane fluidity that could
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be associated with a lower risk to develop cardiovascular diseases.
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Keywords
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Dietary supplementation, erythrocytes phospholipids, extra virgin olive oil, functional oils, hybrid palm oil.
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INTRODUCTION
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A certain role in the maintenance of the functionality of the cells is played by the properties of lipids,
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mainly phospholipids (PL), and associated proteins, which form domains responsible for specialized
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functions. In particular, the membranes lipid composition represents a relevant factor for erythrocytes
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properties and biological functions. In fact, the membrane is the only structural component of the
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non-nucleated red cell, and it is responsible for all of its antigenic, transport and mechanical
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characteristics.1 There is abundant literature on erythrocytes lipids, and their modulation by disease.
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The erythrocytes represent a convenient model for studying mechanisms of oxidative damage, as they
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are available in large quantities, do not contain nuclei and mitochondria and are incapable of protein
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synthesis.
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As a general rule, the total fatty acid (FA) profile of erythrocytes is considered a valuable marker for
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human pathologies associated with a situation of oxidative stress (i.e. cardiovascular disease, cancer,
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inflammatory processes).2,3 For example, the sum of the relative percentages of eicosapentaenoic
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(C20:5 3,EPA) and docosahexaenoic (C22:6 3, DHA) acids among the erythrocytes FA constitute
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the Omega-3 Index, an innovative risk marker for the cardiovascular disease (CVD).4 A low value of
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Omega-3 Index is correlated with a higher risk of cardiovascular events, and it is influenced by several
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factors such as diet, age, diabetes, body mass index, gender and other.5 Erythrocytes FA composition
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is also one of the principal factors modulating the erythrocytes membrane fluidity, and the oxidative
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stress connected to the CVD can modify the erythrocyte FA profile by means of lipid peroxidation,
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thus lowering the membrane fluidity and playing a relevant role in the CVD pathogenesis.6,7 One
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innovative possible way to estimate the erythrocyte membrane fluidity is the lipophilic index, that is
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defined as a weighted mean of the erythrocyte FA melting points. In fact, membrane fluidity is
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strongly determined by the van der Waals forces between FA, that are also strongly linked to their
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melting point.8-10
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Besides the total FA profile, the modification in erythrocytes PL profile and concentration has also
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been associated with the physiological activity of the cell. Recent investigation suggests the use 3 ACS Paragon Plus Environment
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erythrocytes as biomarkers for pathological processes associated with oxidative stress. The alteration
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of the characteristic FA profile of each PL fraction has been related to changes in erythrocytes shapes
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and sizes which can also directly affect their oxygen transport behaviour.11 Furthermore, high levels
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of monounsaturated fatty acids (MUFA), especially of oleic acid, were revealed in erythrocytes
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phosphatidylethanolamine (PE) of patients suffering from adenocarcinoma.12 Diabetes and diabetic
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retinopathy were also associated with a reduction of erythrocyte polyunsaturated fatty acids (PUFAs)
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in PE whereas the increase of the amounts of phosphatidylcholine (PC) species was found in
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erythrocytes of diabetic patients without diabetic retinopathy.13 Jimenez et al.14 found a direct
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relationship among the sphingomyelin (Sph) of erythrocytes membranes and the plasma total
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cholesterol and LDL-cholesterol in patients with significant coronary arterial disease, indicating a
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possible role of Sph on the formation of atherosclerotic plaques.
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Taking into account the growing evidence suggesting the marked influence of dietary lipids on cell
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membrane lipid composition,15 the present paper was aimed to estimate the ability of two selected
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edible oils [Extra Virgin Olive Oil (EVOO) and Hybrid Palm Oil (HPO)] consumption to modulate
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the lipid composition of erythrocytes membrane. For the first time, the effects of HPO on erythrocytes
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fatty acid composition and the influence of EVOO and HPO supplementation on phospholipid
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molecular species composition were investigated. The findings could help to get more knowledge on
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the health benefits of dietary oil and their potential as a functional oil for disease prevention,
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especially CVD.
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In detail, the erythrocytes FA composition of voluntary patients submitted to HPO or EVOO
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supplementations (3-month, 25mL/day) were investigated, with particular interest in the
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determination of innovative risk markers for CVD such as lipophilic and Omega-3 indexes.
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Additionally, the molecular species composition of the most abundant phospholipid classes from
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erythrocytes, such as PC, PE and Sph, were determined by a novel and rapid HILIC-UPLC-ESI(+)-
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MS/MS method.
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The selection of the dietary oils was performed taking account that the regular assumption of EVOO
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improves the human health status, preventing cardiovascular diseases,16 cancer, age related processes
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and other chronical diseases17 and can also modulate the composition of the erythrocyte cell
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membrane.18 At the same time, the HPO, a crude oil obtained from the fruits of the interspecific
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hybrid (Elaeis oleifera x Elaeis guineensis) palm, represents an extremely valuable source of
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antioxidant compounds (carotenes, tocopherols and tocotrienols) that might contribute to lower the
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risk of certain diseases.19-23
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In our latest investigations, we found that HPO consumption had a favorable effect on plasma lipids
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pattern related to CVD risk factors and improved the plasma antioxidant status and that these effects
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were not statistically different from those of EVOO. In detail, a supplementation of 25 mL of HPO
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for a period of three months significantly decreases the low-density lipoprotein cholesterol (LDL-C)
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level and the total cholesterol content in human plasma.24 Furthermore, it significantly increases
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(p
