original papers - Advances in Clinical and Experimental Medicine

ORIGINAL PAPERS
Adv Clin Exp Med 2008, 17, 1, 15–26
ISSN 1230−025X
© Copyright by Silesian Piasts
University of Medicine in Wrocław
BOŻENA REGULSKA−ILOW, RAFAŁ ILOW
The Influence of Quercetin on Fatty−Acid Content
in Selected Organs of Rats on Diets with Fresh
and Oxidized Fat
Wpływ kwercetyny na skład kwasów tłuszczowych
w wybranych narządach szczurów doświadczalnych
na diecie z tłuszczem świeżym i utlenionym
Department of Food Science and Nutrition, Silesian Piasts University of Medicine in Wrocław, Poland
Abstract
Background. The kind and quality of dietary fat may condition the biological activity of quercetin, which is man−
ifested by its influence on the activity of n−3, n−6, and n−9 metabolic pathway enzymes.
Objectives. Evaluation of the effect of quercetin on fatty−acid content in the plasma and selected organs of exper−
imental rats in a condition of oxidative stress due to oxidized dietary fats.
Material and Methods. Using rats fed a diet with an 8% fat content and a 0.05% cholesterol supplement, the influ−
ence of quercetin on the content of fatty acids in the plasma, liver, kidneys, heart, and lung was assessed. Fatty
extracts from the biological material were obtained by means of the Folch method. The composition of fatty acids
(as the percentage of the total level of acids) in the form of methyl esters was determined by gas chromatography
using a 100−m glass capillary column. The source of pro−oxidants in the rats’ diet was either oxidized sunflower
oil or oxidized lard. The four−week experiment involved 80 male Buffalo rats, of which 40 received 0.075%
quercetin as a dietary supplement.
Results. Quercetin affected the fatty−acid content in the investigated organs in the animals fed vegetable fat. The
degree of changes depended on the quality of the fat. Fat statistically significantly changed the content of C16, C18,
C18 = 2, C20 = 2, C22 = 6, and C20 = 4 fatty acids in the liver and plasma and C18 = 1 n−9 in the kidneys. The
content of C18 = 2 was higher by a factor of 3.2 in the liver, 2.8 in the kidneys, plasma, and lungs, and 1.6 in the
heart of rats fed on fresh oil and quercetin. A diet with oxidized oil and quercetin resulted in increases in C18 = 2
of 1.7−fold in the plasma, 1.6−fold in the liver, 1.5−fold in the lungs, 1.4−fold in the kidneys, and 1.2−fold in the heart
compared with the control group. Preference of the n−9 pathway of fatty−acid synthesis was observed.
Conclusions. Quercetin affected the metabolism of fatty acids in the organism when the dietary fat was rich in
polyunsaturated fatty acids. Probably polyunsaturated fatty acids, with they become components of the cell mem−
branes, enable quercetin to pass inside the cells of tissues and to exert the biogenic effect (Adv Clin Exp Med
2008, 17, 1, 15–26).
Key words: fatty acids, oxidized fats, quercetin, rats, liver.
Streszczenie
Wprowadzenie. Rodzaj i jakość tłuszczu w diecie mogą warunkować aktywność biologiczną kwercetyny, przeja−
wiającą się jej wpływem na aktywność enzymów szlaków metabolicznych n−3, n−6, n−9.
Cel pracy. Ocena wpływu kwercetyny na skład kwasów tłuszczowych w osoczu i wybranych narządach szczurów
doświadczalnych w warunkach stresu oksydacyjnego wywołanego utlenionymi tłuszczami pokarmowymi.
Materiał i metody. Podawano szczurom diety z 8% zawartością tłuszczu oraz 0,5% dodatkiem cholesterolu i oce−
niano wpływ kwercetyny na skład kwasów tłuszczowych w osoczu, wątrobie, nerkach, sercu i płucach. Ekstrakty
tłuszczowe z materiału biologicznego otrzymano metodą Folcha. Skład kwasów tłuszczowych (jako% sumy kwa−
sów) w postaci estrów metylowych oznaczono metodą chromatografii gazowej z zastosowaniem kolumny kapilar−
nej szklanej o długości 100 m. Źródłem prooksydantów w diecie szczurów były utlenione: olej słonecznikowy
i smalec. Czterotygodniowe doświadczenie żywieniowe przeprowadzono z udziałem 80 szczurów, samców, rasy
Buffalo, których 40 otrzymywało 0.075% dodatek kwercetyny jako suplement diety.
16
B. REGULSKA−ILOW, R. ILOW
Wyniki. Kwercetyna wpływała na skład kwasów tłuszczowych w wybranych narządach w grupach zwierząt kar−
mionych tłuszczem roślinnym, a wielkość zmian zależała od jego jakości. W istotny statystycznie sposób zmieniał
się skład następujących kwasów tłuszczowych: C16, C18, C18 = 2, C20 = 2, C22 = 6, w wątrobie i osoczu także
C20 = 4, a w nerkach C18 = 1 n−9. Udział C18 = 2 zwiększył się 3,2 raza w wątrobach, 2,8 raza w nerkach, oso−
czu i płucach oraz 1,6 raza w sercach szczurów karmionych olejem świeżym i kwercetyną. Na diecie z olejem utle−
nionym i kwercetyną skład C18 = 2 zwiększył się 1,7 raza w osoczu, 1,6 raza w wątrobach, 1,5 raza w płucach,
1,4 raza w nerkach i 1,2 raza w sercach w stosunku do grupy kontrolnej. Obserwowano preferencję syntezy kwa−
sów szlaku n−9.
Wnioski. Kwercetyna wpływała na metabolizm kwasów tłuszczowych w organizmie wtedy, gdy był spożywany
tłuszcz bogaty w wielonienasycone kwasy tłuszczowe. WKT, stając się składnikiem błon komórkowych, umożli−
wiają kwercetynie wniknięcie do wnętrza komórek tkanek i biogenne działanie (Adv Clin Exp Med 2008, 17, 1,
15–26).
Słowa kluczowe: kwasy tłuszczowe, utlenione tłuszcze, kwercetyna, szczury, wątroba.
Epidemiological studies have shown that
quercetin constitutes about 75% of ingested
bioflavonoids and the source of its intake depends
on the dietary habits of the investigated popula−
tion. It is present in diets rich in vegetables, fruit,
and condiments [1–7]. Quercetin is also a compo−
nent of pharmaceutical products commonly used
in the treatment of colds. Interest in quercetin is
associated with its prevalence and widespread use
as well as its extremely broad spectrum of action.
It also has one of the highest antioxidative poten−
tials among this group of compounds [6]. It is
responsible for inhibition of the oxidation of fatty
acids, which are components of phospholipids in
cell membranes [12]; it also exerts a hepatoprotec−
tive effect [8–11]. Quercetin has been shown to
affect the activities of numerous enzymes [2,
13–18]. Its anti−inflammatory properties are the
consequence of its inhibition of cyclooxygenase
and lipooxygenase activity [14, 18]. Its effect on
the activities of elongase and desaturase, which
participate in fatty−acid metabolism, may result in
a change in the fatty−acid content of tissues.
Quercetin inhibits the activity of hyaluronidase
and thereby shows protective properties on blood
vessels [19]. Its antiatherogenic and cholagogic
action has been confirmed experimentally [20].
Dietary fat affects the composition of fatty
acids in the phospholipids constituting tissue cell
membranes. An adequate content of polyunsatu−
rated fatty acids in membrane lipids conditions
their liquidity and permeability [21–23]. The lack
of studies on the influence of dietary components
that potentially modify the action of quercetin was
the basis for designing the present experiment. It
was assumed that factors that may condition the
effect of quercetin include the kind (vegetable or
animal) and quality (fresh or oxidized) of ingested
fats. The quality of the oxidized fat added to the
food was evaluated by means of its peroxide and
anisidine values and the content of the polar frac−
tion and it corresponded to the quality of deep−fry−
ing oil from fast−food restaurants.
Material and Methods
The Experiment
The study was carried out on eighty male
Buffalo rats with initial and final body weights of
147 ± 28.3 g and 226.0 ± 28.4 g. For the four
weeks of the experiment the rats were kept in
appropriate conditions (room temperature, 12−
hour light−dark rhythm). All procedures for the
animal experiments were approved by official
authorities.
The animals were divided into eight groups,
10 animals per group. Forty rats had a diet that was
8% sunflower oil, with 20 receiving oxidized oil
and 20 fresh oil. Ten of the former and 10 of the
latter had quercetin as a supplement to their diet.
The other 40 rats had a diet that was 8% lard, with
20 receiving oxidized lard and 20 fresh lard. Ten
of the former and 10 of the latter also had the
quercetin supplement. The supplement came to
0.75 g per kilo of diet (Quercetin Dihydrate
reagent, Fluka, cat. No. 83370). The diet was pre−
pared according to the method described in [24].
The rats had unlimited access to the diet and to
water. Their consumption of diet and water was
checked every two days and their body mass mea−
sured once a week.
Preparation of Oxidized Fat
for Use as an Ingredient
of the Diet
Two equal portions of pork lard and two of
refined sunflower oil were made up. One portion
of each was kept fresh, while the other was sub−
jected to thermal oxidation. The oxidation process
was carried out by heating the fat under a quartz
lamp for 65 hours. A 2.5−cm thick layer of fat was
placed in a porcelain dish, 21 x 29 cm. The quartz
lamp was kept 14 cm from the surface of the fat.
The initial temperature of the fat was 50°C and its
17
The Influence of Quercetin on Fatty−Acid Content in Selected Rat Organs
Table 1. Fatty−acid composition, expressed as the percentage of total fatty−acid content, and quality indicators for the fresh
and oxidized fats added to the rats’ fodder
Tabela 1. Skład kwasów tłuszczowych (jako udział procentowy w puli wszystkich kwasów) i wskaźniki jakości tłuszczu
świeżego i utlenionego dodawanego do diety
Analysed indicator
(Badane wskaźniki)
Fresh lard
(Świeży smalec)
Oxidized lard
(Utleniony smalec)
Fresh sunflower oil
(Świeży olej
słonecznikowy)
Oxidized sunflower oil
(Utleniony olej
słonecznikowy)
Peroxide value
(Liczba nadtlenkowa)
meg O2/kg
mg O2/100 g fat
1.5 ± 0.0
0.046
169.3 ± 0.2
5.28
3.4 ± 0.0
0.107
666.7 ± 0.0
20.8
Anisidine value
(Liczba anizydynowa)
0.5 ± 0.2
39.1 ± 0.4
4.5 ± 0.0
68.8 ± 0.8
Polar fraction [%]
(Frakcja polarna)
1.3
16.9
2.1
36.5
Saturated acids
(Kwasy nasycone)
43.3
43.6
8.6
24.5
Monounsaturated acids
(Kwasy jednonienasycone)
45.1
45.5
22.3
30.6
Polyunsaturated acids
(Kwasy wielonienasycone)
10.7
9.8
68.2
42.9
Unidentified acids
(Kwasy niezidentyfikowane)
0.9
1.1
0.9
2.0
Fatty acid type sum
(Suma kwasów tłuszczowych)
final temperature was not higher than 75°C. The
oxidation conditions were established based on the
work of Ziemlański et al. [25]. In the fresh and
oxidized fats, the content of the polar fraction was
determined, and the fatty−acid component and the
content of the primary and secondary products of
oxidation were measured as the peroxide and ani−
sidine values (Table 1).
Extraction of Lipid
Components From the
Investigated Material
According to the Folch
Method [26]
After being weighed and ground, the individ−
ual organs were homogenized with 10 ml of
a chloroform:methanol mixture (2:1). The extract
with sediment was transferred to 100−ml bottles
and shaken mechanically for 5 minutes and then
centrifuged for 10 minutes. The chloroform:
methanol extract was transferred to a separator and
the residue was shaken again with 10 ml of the
extracting mixture, chloroform:methanol (2 : 1)
and centrifuged. Then the chloroform:methanol
extracts collected in the separator were treated
with 15 ml of a 0.9% solution of potassium chlo−
ride, hand shaken, and left for separation of the
layers. The lower, chloroform layer was collected
in Erlenmeyer flasks containing anhydrous sodium
sulfate, dried for 24 hours, and filtered into bottles.
The extracts were stored at +4°C in dark, tightly
closed, 50−ml bottles.
Determination of the Percentage
of Fatty Acids in Extracts From
Plasma and Organs by Gas
Chromatography
The composition of fatty acids in the form of
methyl esters was determined by means of gas
chromatography using the 6890 N with Agilent
ChemStation, version A.8.03 (Agilent Techno−
logies) with a 100−m glass capillary column 0.25
in diameter and a stationary phase of CP−Sil 88.
Identification of the acids was performed by com−
paring their retention times with standards. The
content of the particular fatty acids and their iso−
mers was expressed as a percentage of the total fat.
Stage I
The chloroform extract was collected into an
estrification tube and chloroform was evaporated
with a nitrogen stream in a glycerin bath at 70°C.
The mixture was treated with 1 ml of 2 M KOH in
18
B. REGULSKA−ILOW, R. ILOW
75% methanol. The test tube was sealed with
a stopper and heated for 60 minutes at 70°C. After
cooling, 1 ml of hexane was added. After 5 minutes of shaking, the hexane layer was discarded
and the extraction process was repeated. Then 1 ml
of 2 M aqueous HCl solution was added and heated for 30 minutes in a tightly sealed test tube in
a glycerin bath. After cooling, the contents were
treated with 1 ml of hexane and a saturated solution of NaCl so that the hexane layer was in the
narrowing of the estrification tube. After 5 minutes
of shaking, the hexane solution was collected with
a Pasteur pipette into vials containing anhydrous
sodium sulfate. The extraction was repeated with
another 1 ml of hexane. The combined hexane
solutions were dried for 24 hours by anhydrous
sodium sulfate.
Stage II
The dried hexane solution was transferred to
the estrification tube and evaporated with a nitro−
gen stream in a glycerin bath at 70°C. After the
complete evaporation of hexane, the mixture was
treated with 1 ml of 0.5 M KOH in anhydrous
methanol, sealed with a stopper, and heated for 30
minutes at 70°C. After cooling, 1 ml of a 1.25
M HCl solution in anhydrous methanol was added
and heated again in a tightly sealed test tube. After
cooling, the mixture was treated with 1 ml of hexane and a saturated NaCl solution so that the hexane layer was in the narrowing of the tube. After
5 minutes of shaking, the hexane solution was collected with a Pasteur pipette into vials containing
anhydrous sodium sulfate. The extraction was
repeated with another 1 ml of hexane. The solution
was left to evaporate over anhydrous sodium sulfate. The solution which was above the sodium
sulfate was transferred to a test tube and condensed by evaporation with a nitrogen stream. The
solution, condensed to about 0.02 ml, was injected
into the chromatograph at an amount of 0.001 ml.
The percent content of fatty acids was calcu−
lated as the quotient of the peaks and the distance
from the solvent fronting.
Statistical Assessment
of the Results
The normality of the distribution in the exam−
ined groups was assessed with Shapiro−
Wilk’s W test. By lack of a normal distribution,
logarithmic transformation was applied. The sig−
nificance of the differences between the average
values for the examined groups was assessed using
one−way analysis of variance (p < 0.05). Data were
tested for homogeneity of variances with
Levene’s test. To assess the influence of differ−
ences on the analysis of variance (the average post
hoc comparison), Tukey’s Honest Significant
Difference test (HSD) was applied. The Cochran−
Cox test was used in cases of a normal distribution
of variables but a lack of homogeneity of the vari−
ance. The non−parametric Kruskal−Wallis test was
used for comparisons in cases of a lack of a normal
distribution and a lack of homogeneity of variance
of the variables. All the statistical calculations
were done with STATISTICA 6.0 PL (StatSoft
Inc., USA).
Results
Fat Added to the Diet (Table 1)
As a result of oxidation, the peroxide value
increased 194−fold in sunflower oil and 114−fold in
lard. The anisidine value increased after oxidation
15 times in sunflower oil and 65 times in lard. The
applied experimental conditions of fat oxidation
resulted in a significant decomposition of polyun−
saturated fatty acids in the sunflower oil, their con−
tent in oxidized oil falling by 25%. The composi−
tion of fatty acids in oxidized lard changed only
slightly in relation to the composition in its fresh
counterpart. The content of the polar fraction in
fats increased due to oxidation from 1.3 to 23.9%
in lard and from 2.1 to 42.3% in oil, i.e. 18.4 and
20.1 times, respectively.
The Effect of Quercetin on the
Percent Content of Fatty Acids
in Selected Tissues
in Experimental Rats
(Tables 2 and 3)
The influence of quercetin on the content of
fatty acids in the selected organs was observed in
the groups of animals fed on a diet containing oil,
the range of changes correlating with the quality of
the fat. It involved mainly the n−6, n−3, and n−9
metabolic pathways and statistically significantly
changed the content of the fatty acids in the inves−
tigated organs: C16, C18, C18 = 2, C20 = 2, C22
= 6, and C20 = 4 in plasma and liver and C18 = 1,
n−9, in the kidneys.
Under the effect of quercetin a 3.2−fold
increase in the content of linoleic acid (C18 = 2, n−6)
was observed in the liver, 2.8−fold in the kidneys,
plasma, and lung, and 1.6−fold in the hearts of the
rats fed fresh oil. In the group of animals receiving
oxidized oil and quercetin, the content of C18 = 2
was higher by a factor of 1.7 in the plasma, 1.6 in
20.7 ± 1.0
13.1 ± 1.8
19.3 ± 2.8
3.3 ± 0.4
9.7 ± 1.5
1.0 ± 0.6
19.6 ± 2.7
4.1 ± 1.0
26.2 ± 0.7
6.2 ± 1.0
33.3 ± 2.2
3.8 ± 0.3
8.9 ± 0.8
0.2 ± 0.0
4.9 ± 0.7
0.3 ± 0.1
21.1 ± 0.3
8.3 ± 0.7
29.9 ± 1.5
3.5 ± 0.1
10.4 ± 0.5
1.0 ± 0.0
13.7 ± 0.6
1.5 ± 0.1
Kidneys (Nerki)
C16
C18
C18 = n9
C18 = n7
C18 = 2
C20 = 2
C20 = 4
C22 = 6
Plasma (Osocze)
C16
C18
C18 = n9
C18 = n7
C18 = 2
C20 = 2
C20 = 4
C22 = 6
Control oil
– group 1
(Olej – grupa 1
kontrolna)
Liver (Wątroba)
C16
C18
C18 = n9
C18 = n7
C18 = 2
C20 = 2
C20 = 4
C22 = 6
Fatty acids
(Kwasy tłuszczowe)
20.9 ± 0.2
9.0 ± 0.3
34.5 ± 0.5
4.0 ± 0.3
8.7 ± 0.7
1.7 ± 0.1
7.2 ± 0.3
1.1 ± 0.1
27.9 ± 1.3
5.7 ± 0.9
37.6 ± 2.3
4.7 ± 0.6
3.7 ± 0.6
0.3 ± 0.1
4.1 ± 1.0
0.3 ± 0.1
21.9 ± 1.7
6.7 ± 0.8
29.4 ± 2.0
4.2 ± 0.4
7.6 ± 0.7
1.5 ± 0.4
10.6 ± 2.1
3.3 ± 0.7
Control lard
– group 2
(Smalec – grupa 2
kontrolna)
19.1 ± 0.5
7.2 ± 0.6
21.0 ± 1.4
3.3 ± 0.2
28.8 ± 2.3
0.1 ± 0.1
9.0 ± 0.4
0.6 ± 0.1
23.1 ± 1.1
4.1 ± 0.7
26.1 ± 1.3
2.9 ± 0.2
24.6 ± 1.3
0.1 ± 0.0
4.7 ± 0.8
0.2 ± 0.0
17.9 ± 1.2
6.3 ± 1.4
18.2 ± 2.1
3.4 ± 0.2
31.0 ± 1.6
0.4 ± 0.1
12.0 ± 2.5
1.3 ± 0.2
Oil + quercetin
– group 3
(Olej + kwerce−
tyna – grupa 3)
20.8 ± 0.2
7.7 ± 0.3
33.7 ± 1.0
4.2 ± 0.2
11.4 ± 1.0
0.7 ± 0.0
8.3 ± 0.2
1.2 ± 0.0
26.8 ± 1.1
5.1 ± 0.8
37.6 ± 2.2
4.9 ± 0.5
4.6 ± 0.6
0.2 ± 0.0
4.1 ± 0.8
0.3 ± 0.0
21.4 ± 0.9
8.7 ± 2.2
29.1 ± 2.5
4.5 ± 0.5
8.5 ± 1.5
0.7 ± 0.3
11.3 ± 2.1
3.0 ± 0.6
Lard + quercetin
– group 4
(Smalec + kwer−
cetyna – grupa 4)
21.3 ± 0.3
9.1 ± 0.3
28.7 ± 0.3
3.5 ± 0.0
10.3 ± 0.1
0.9 ± 0.6
14.8 ± 0.3
1.6 ± 0.0
24.7 ± 1.3
4.5 ± 0.5
31.8 ± 1.8
4.0 ± 0.2
9.0 ± 1.2
0.2 ± 0.1
5.4 ± 1.0
0.4 ± 0.1
20.3 ± 1.3
10.9 ± 1.5
18.3 ± 2.2
3.4 ± 0.4
10.5 ± 2.1
1.2 ± 0.6
20.3 ± 3.4
5.0 ± 0.5
Oxidized oil
– group 5
(Olej utleniony
– grupa 5)
21.6 ± 0.3
8.5 ± 0.3
34.8 ± 0.3
3.6 ± 0.3
9.3 ± 0.2
1.7 ± 0.0
7.9 ± 0.2
1.1 ± 0.0
28.8 ± 0.7
5.0 ± 0.8
35.1 ± 1.7
4.2 ± 0.2
4.2 ± 0.6
0.2 ± 0.0
3.6 ± 0.3
0.3 ± 0.0
22.8 ± 1.9
8.6 ± 1.4
29.3 ± 2.7
4.1 ± 0.4
7.3 ± 1.0
1.4 ± 0.6
10.1 ± 2.9
2.8 ± 0.8
Oxidized lard
– group 6
(Smalec utle−
niony – grupa 6)
Tabela 2. Wpływ różnych rodzajów diety na skład kwasów tłuszczowych (%) w wybranych narządach szczurów laboratoryjnych (x ± SD)
Table 2. The influence of the kind of diet on fatty−acid content (%) in selected organs of laboratory rats (X ± SD)
21.6 ± 0.3
7.9 ± 0.4
26.9 ± 0.3
3.4 ± 0.1
17.6 ± 0.4
0.2 ± 0.0
10.8 ± 0.2
1.0 ± 0.1
27.3 ± 1.3
5.3 ± 1.2
29.4 ± 1.8
3.5 ± 0.3
13.0 ± 1.5
0.1 ± 0.0
4.8 ± 1.3
0.3 ± 0.0
20.3 ± 1.2
9.7 ± 1.4
17.2 ± 1.7
3.4 ± 0.27
17.0 ± 1.4
0.3 ± 0.1
18.3 ± 2.5
3.2 ± 0.4
Oxidized oil
+ quercetin
– group 7
(Olej utleniony
+ kwercetyna
– grupa 7)
22.1 ± 0.1
8.8 ± 0.3
33.7 ± 0.6
4.1 ± 0.2
10.2 ± 0.2
1.0 ± 0.0
7.4 ± 0.2
1.0 ± 0.1
28.3 ± 0.7
5.3 ± 0.8
35.2 ± 1.2
4.5 ± 0.3
4.7 ± 0.5
0.2 ± 0.1
4.2 ± 0.8
0.3 ± 0.1
22.5 ± 0.9
6.6 ± 0.9
30.0 ± 1.9
4.1 ± 0.3
8.8 ± 1.1
0.8 ± 0.3
9.7 ± 1.5
2.6 ± 0.3
Oxidized lard
+ quercetin
– group 8
(Smalec utleniony
+ kwercetyna
– grupa 8)
The Influence of Quercetin on Fatty−Acid Content in Selected Rat Organs
19
15.4 ± 2.1
9.3 ± 1.5
13.1 ± 3.3
4.1 ± 0.1
18.0 ± 2.1
0.2 ± 0.1
20.6 ± 4.1
6.1 ± 1.4
29.8 ± 0.5
6.8 ± 0.4
32.1 ± 2.0
4.4 ± 0.2
6.4 ± 0.5
0.2 ± 0.0
3.6 ± 0.7
0.3 ± 0.1
Lungs (Płuca)
C16
C18
C18 = n9
C18 = n7
C18 = 2
C20 = 2
C20 = 4
C22 = 6
Control oil
– group 1
(Olej – grupa 1
kontrolna)
Heart (Serce)
C16
C18
C18 = n9
C18 = n7
C18 = 2
C20 = 2
C20 = 4
C22 = 6
Fatty acids
(Kwasy tłuszczowe)
29.7 ± 1.1
6.1 ± 0.4
36.3 ± 1.6
5.1 ± 0.4
2.8 ± 0.3
0.1 ± 0.0
2.9 ± 0.6
0.3 ± 0.1
13.9 ± 1.5
7.4 ± 0.6
15.3 ± 2.5
4.7 ± 0.2
15.6 ± 1.2
1.0 ± 0.3
21.8 ± 2.6
7.7 ± 1.1
Control lard
– group 2
(Smalec – grupa 2
kontrolna)
27.1 ± 1.2
5.6 ± 0.9
27.9 ± 2.6
3.6 ± 1.0
17.9 ± 1.3
0.3 ± 0.1
3.2 ± 1.1
0.2 ± 0.1
12.7 ± 1.7
7.4 ± 0.6
10.8 ± 2.5
3.5 ± 0.3
28.6 ± 1.0
0.1 ± 0.0
19.7 ± 3.0
5.5 ± 1.4
Oil + quercetin
– group 3
(Olej + kwerce−
tyna – grupa 3)
29.4 ± 0.8
4.9 ± 0.5
36.7 ± 2.2
4.9 ±0.3
3.9 ±0.7
0.1 ± 0.0
3.8 ± 1.1
0.4 ± 0.1
15.3 ± 2.2
8.3 ± 1.1
16.1 ± 4.1
4.5 ± 0.2
14.9 ± 2.3
0.5 ± 0.2
20.2 ± 3.5
6.8 ±1.2
Lard + quercetin
– group 4
(Smalec + kwer−
cetyna – grupa 4)
29.1 ±0.7
5.0 ± 0.7
30.5 ± 1.6
4.8 ± 0.3
6.5 ± 0.9
0.1 ±0.0
3.9 ± 0.5
0.4 ± 0.1
16.0 ± 1.3
8.4 ± 0.6
14.9 ± 2.4
4.1 ± 0.3
18.3 ± 1.6
0.5 ± 0.2
18.4 ± 2.3
6.5 ± 1.2
Oxidized oil
– group 5
(Olej utleniony
– grupa 5)
29.4 ± 1.3
5.0 ± 0.7
32.7 ± 2.9
4.6 ± 0.2
4.7 ± 1.3
0.1 ± 0.0
3.4 ± 1.3
0.3 ± 0.1
17.7 ± 3.8
7.9 ± 2.0
19.5 ± 6.0
3.9 ± 0.4
13.3 ± 3.3
0.6 ± 0.2
16.4 ± 5.4
5.8 ± 2.0
Oxidized lard
– group 6
(Smalec utle−
niony – grupa 6)
Tabela 2. Wpływ różnych rodzajów diety na skład kwasów tłuszczowych (%) w wybranych narządach szczurów laboratoryjnych (x ± SD) – cd.
Table 2. The influence of the kind of diet on fatty−acid content (%) in selected organs of laboratory rats (X ± SD) – continued
29.3 ± 0.6
5.3 ± 0.9
29.5 ± 2.6
4.0 ± 0.3
9.9 ± 1.2
0.2 ± 0.0
3.8 ± 1.0
0.3 ± 0.1
15.8 ± 2.9
7.4 ± 1.1
14.8 ± 4.5
3.4 ± 0.3
22.8 ± 2.7
0.1 ± 0.0
16.9 ± 4.3
5.5 ± 1.4
Oxidized oil
+ quercetin
– group 7
(Olej utleniony
+ kwercetyna
– grupa 7)
29.2 ± 1.1
4.5 ± 0.4
32.1 ± 1.8
4.7 ± 0.2
4.6 ± 0.8
0.1 ± 0.0
3.4 ± 0.9
0.3 ± 0.1
14.8 ± 1.7
7.5 ± 0.7
14.7 ± 2.9
3.8 ± 0.1
16.0 ± 1.9
0.5 ± 0.2
20.6 ± 2.8
7.7 ± 1.0
Oxidized lard
+ quercetin
– group 8
(Smalec utleniony
+ kwercetyna
– grupa 8)
20
B. REGULSKA−ILOW, R. ILOW
21
The Influence of Quercetin on Fatty−Acid Content in Selected Rat Organs
Table 3. Comparison of the effect of a diet with oil on the percent content of fatty acids in selected organ of experimental
rats; significance of differences
Tabela 3. Porównanie wpływu diety z olejem na skład kwasów tłuszczowych w wybranych narządach szczurów doświad−
czalnych, różnice istotne statystycznie
Fatty acids
(Kwasy tłuszczowe)
Group 1 vs. 3
(Grupa 1 vs 3)
Group 3 vs. 7
(Grupa 3 vs 7)
Group 1 vs. 5
(Grupa 1 vs 5)
Group 5 vs. 7
(Grupa 5 vs 7)
Liver (Wątroba)
C16
C18
C18 = 2
C20 = 2
C20 = 4
C22 = 6
0.0003
0.0001
0.0001
0.0000²
0.0001
0.0000¹
0.0024
0.0002
0.0001
ns.²
0.0001
0.0044¹
ns.
0.0291
ns.
ns.²
ns.
ns.¹
ns.
ns.
0.0001
0.0000²
ns.
ns.¹
Kidneys (Nerki)
C16
C18
C18 = 1n9
C18 = 2
C20 = 2
C22 = 6
0.0001
0.0001
0.0001
0.0001
0.0000²
0.0001
0.0001
0.0346
0.0007
0.0001
ns.²
0.0002
0.0313
0.0013
ns.
0.0046
0.0046²
ns.
0.0001
ns.
0.0489
0.0001
0.0000²
0.0077
Plasma (Osocze)
C16
C18
C18 = 1n9
C18 = 2
C20 = 2
C20 = 4
C22 = 6
0.0001
0.0003
0.0001
0.0000
0.0000
0.0001
0.0001
ns.
ns.
0.0001
0.0003
ns.
ns.
0.0291
ns.
ns.
ns.
ns.
ns.
ns.
ns.
ns.
0.0024
ns.
0.0001
0.0346
0.0001
0.0001
Heart (Serce)
C16
C18
C18 = 2
C20 = 2
0.0489
0.0346
0.0001
0.0489
0.0346
ns.
0.0077
ns.
ns.
ns.
ns.
ns.
ns.
ns.
0.0346
0.0001
Lung (Płuca)
C16
C16 = 1
C18
C18 = 1n7
C18 = 2
C20 = 2
C22 = 6
0.0003¹
ns.
0.0014²
ns.¹
0.0001²
0.0000²
0.0440
0.0478
ns.¹
ns.²
ns.¹
0.0000²
0.0015²
ns.¹
ns.¹
0.0178¹
0.0001²
ns.¹
ns.²
ns.²
ns.¹
ns.¹
ns.¹
ns.²
0.0059
0.0001²
0.0155²
ns.¹
ns. – insignificant differences.
ns. – różnica nieistotna statystycznie.
¹ Kruskal-Wallis test.
¹ Test Kruskala-Wallisa.
² Cochran-Cox test.
² Test Cochrana-Coxa.
the livers, 1.5 in the lungs, 1.4 in the kidneys, and
1.2 in the hearts compared with the control group.
Fig. 1 illustrates the changes in the content of C18
= 2 fatty acid in the investigated organs.
The liver and plasma of the rats on a diet with
fresh oil revealed decreased arachidonic acid (C20
= 4, n−6) content in all the fatty acids, by a factor
of 1.5 in the liver and 0.7 in the plasma. A similar
effect of quercetin was observed in animals fed
fodder containing oxidized oil, but it was twofold
lower. The accumulation of linoleic acid did not
significantly affect the content of arachidonic acid
in the lungs, hearts, and kidneys (Fig. 2).
Decreases in the content of all the products of
the n−9 metabolic pathway, i.e. C18, C18 = 1, and
C20 = 2, were observed in the organs of the ani−
mals on diets containing quercetin compared with
controls. The content of stearic acid (C18), the ini−
tial substrate of metabolism, decreased twofold in
the livers, by about 20% in the hearts and lungs,
13% in the plasma, and 25% in the kidneys of the
animals receiving fresh oil with quercetin. The kid−
22
B. REGULSKA−ILOW, R. ILOW
35
Fig. 1. Comparison of
the effect of quercetin on
C18 = 2 acid content in
selected organs of rats on
a diet with oil
30
25
20
Ryc. 1. Porównanie wpły−
wu kwercetyny na skład
kwasu C18 = 2 w wybra−
nych narządach szczurów
na diecie z olejem
15
10
5
0
fresh oil
świeży olej
lungs
płuca
fresh oil + quercetin
świeży olej
+ kwercetyna
kidney
nerki
oxidized oil
utleniony olej
liver
wątroba
oxidized oil + quercetin
utleniony olej
+ kwercetyna
plasma
osocze
heart
serce
25
Fig. 2. Comparison of
the effect of quercetin on
C20 = 4 acid content in
selected organs of rats on
a diet with oil
20
15
10
5
0
fresh oil
świeży olej
lungs
płuca
fresh oil + quercetin
świeży olej
+ kwercetyna
kidneys
nerki
oxidized oil
utleniony olej
liver
wątroba
oxidized oil
+ quercetin
utleniony olej
+ kwercetyna
plasma
osocze
Ryc. 2. Porównanie
wpływu kwercetyny na
skład kwasu C20 = 4
w wybranych narządach
szczurów na diecie
z olejem
heart
serce
1.4
Fig. 3. Comparison of
the effect of quercetin on
C20 = 2 acid content in
selected organs of rats on
a diet with oil
1.2
1
0.8
0.6
0.4
0.2
0
fresh oil
świeży olej
lungs
płuca
fresh oil + quercetin
świeży olej
+ kwercetyna
kidney
nerki
oxidized oil
utleniony olej
liver
wątroba
neys also showed a 22% decreased content of oleic
acid (C18 = 1), which was also 1.2 times lower in
the hearts and lungs compared with controls.
Moreover, quercetin affected the content of an
indirect product of n−9 acid metabolism, i.e.
plasma
osocze
oxidized oil
+ quercetin
utleniony olej
+ kwercetyna
Ryc. 3. Porównanie
wpływu kwercetyny na
skład kwasu C20 = 2
w wybranych narządach
szczurów na diecie
z olejem
heart
serce
eicosadienoic acid (C20 = 2), in the investigated
organs regardless of the quality and kind of fat
added to the animals’ diet. The content of C20 = 2
fatty acid in the liver and kidney of animals receiv−
ing fresh oil was decreased by a factor of 2.5, in
The Influence of Quercetin on Fatty−Acid Content in Selected Rat Organs
the plasma by 10, and in the hearts by about 5
under the effect of flavonoid. On the other hand,
changes in the content of C20 = 2 acid in the lungs,
where a twofold increase was observed, revealed
a reverse trend. In the livers of the animals fed oxi−
dized oil and quercetin, the content of C20 = 2 acid
was four times lower and in the kidneys two times
lower (Fig. 3).
The content of eicosadienoic acid in the livers
of the animals receiving fodder with lard and
quercetin was 2 times lower, in the kidneys 1.5
times lower, in the plasma 2.4 times lower, and in
the hearts 2.0 times lower. The livers of the ani−
mals fed fodder containing oxidized lard and
quercetin showed a 1.75 times lower content of
C20 = 2, in the plasma the levels were 1.7 times
lower, and in the hearts 5 times lower. The effect
of quercetin observed under the experimental con−
ditions on the content of C20 = 2 fatty acid in the
organs of the animals fed fodder containing lard
may result from the high participation of oleic
acid, a substrate of the n−9 fatty−acid metabolic
pathway, in the fat (Fig. 4).
The addition of quercetin resulted in
a decrease in the content of C16 acid in the livers,
kidneys, lungs, and plasma of the rats fed fresh oil.
The kidneys also revealed an increased content of
palmitic acid in the group of animals fed oxidized
oil with the addition of quercetin, which was 11%
higher than in the control group.
The livers and kidneys of the animals receiv−
ing fodder with fresh oil and quercetin revealed
a decreased in the C22 = 6/C18 = 3α index com−
pared with the control group. Its value was 7 times
lower in the liver and 2 times lower in the kidneys.
The acid content ratio proves the activity of the n−
3 metabolic pathway enzymes, i.e. the synthesis of
docosahexaenoic acid from linolenic acid.
Inhibition of the activity of the enzyme ∆−6 desat−
urase by quercetin in the rats’ livers caused
a twofold increase in the content of C18 = 3α acid.
Accumulation of the acid in the liver resulted in
a threefold decrease in the content of C22 = 6 acid,
while in the kidneys the level of docosahexaenoic
acid fell by a factor of 1.5.
The Effect of the Quality
of the Fat Added to the Diet
on the Fatty Acid Levels
in Rat Tissues (Tables 2 and 3)
Statistically significant differences in the level
of fatty acids in the liver between the groups
receiving fresh and oxidized oil were observed in
case of C18 acid and consequently in the total
level of saturated fats. The content of stearic acid
23
decreased by 17% in livers, 27% in the kidneys,
and 26% in the lungs of the rats receiving oxidized
oil compared with the control group.
Some kidneys also revealed other statistically
significant changes in the content of fatty acids
occurring under the effect of modified vegetable
fat. The observations demonstrated decreased lev−
els of C16 and C18 acids and the total content of
saturated fats and an increased content of C20 = 2
acid. The quality of the lard added to the rats’ diet
affected only the content of C20 = 2 acid in the
kidneys. Its level was decreased by about 27% in
the group of animals fed oxidized lard.
Discussion
The content of the polar fraction in the oxi−
dized fats added to the diet of the rats in this exper−
iment disqualified oil as suitable for consumption.
The content of polar compounds exceeded the
acceptable level, which is 25%. Frying fat reaches
this value after 5–6 days of use in the same deep−
fat fryer [28]. The polar fraction has a fluid con−
sistency and under home or restaurant conditions
its removal from fat is impossible. It is absorbed
into or adsorbed onto the surface of the fried prod−
uct and thus it becomes a dietary component. The
polar fraction decreases the digestibility of dietary
fat, but it is the ingestion of the products of lipid
peroxide metabolism that endangers our health.
The content of the primary and secondary products
of fatty−acid oxidation, evaluated by means of the
peroxide and anisidine values, in the oxidized fats
added to the diet increased significantly. The prod−
ucts of fat oxidation destroy the cell structure and
impair its functioning. Peroxides may damage
intestinal mucous membrane, and aldehydes and
other secondary products of oxidation may exert
a toxic effect on animals’ organs [27, 29].
Under the experimental conditions, an effect of
quercetin on the content of fatty acids in the inves−
tigated organs was observed in the group of ani−
mals on diets which included oil, and the range of
changes correlated with the quality of the oil, it
being lower in a diet with oxidized fat. Sunflower
oil contains primarily polyunsaturated fatty acids,
which increase its liquidity and membrane perme−
ability. Under the experimental conditions, oil con−
tained four times more polyunsaturated acids, even
after oxidation, than lard. The change in cell mem−
brane permeability caused in the experiment by the
kind of fat added to the diet probably conditioned
the activity of quercetin or its lack. Lard contains
mainly saturated and monounsaturated fatty acids
which, after being embedded into cell membranes,
increase their rigidity and decrease permeability.
24
B. REGULSKA−ILOW, R. ILOW
Table 4. Comparison of the effect of a diet with lard on
the percent content of fatty acids in selected organ of
experimental rats; significance of differences
Tabela 4. Porównanie wpływu diety ze smalcem na
udział kwasów tłuszczowych w wybranych narządach
szczurów doświadczalnych, różnice istotne statystycznie
Fatty acids
(Kwasy
tłuszczowe)
Group
2 vs. 4
(Grupa
2 vs 4)
Group
4 vs. 8
(Grupa
2 vs 4)
Group
2 vs. 6
(Grupa
2 vs 4)
Group
6 vs. 8
(Grupa
2 vs 4)
Liver
(Wątroba)
C18
C20 = 2
ns.
0.0010²
0.0331
ns²
ns.
ns²
ns.
0.0007²
Kidneys
(Nerki)
C16
C18 = 2
C20 = 2
ns
0.0009
0.0002²
0.0404
ns.
ns.²
ns.
ns.
0.0092²
ns.
ns.
ns.²
Lung
(Płuca)
C18 = 2
C20 = 2
0.0026
ns.²
ns.²
ns.²
0.0050²
0.0009²
ns.²
ns.²
Plasma
(Osocze)
C20 = 2
0.0001
ns.
ns.
0.0001
Heart
(Serce)
C20 = 2
0.0050
ns.
ns.
ns.
ns – insignificant differences.
ns. – różnica nieistotna statystycznie.
¹ Kruskal-Wallis test.
¹ Test Kruskala-Wallisa.
² Cochran-Cox test.
² Test Cochrana-Coxa.
The liver is the main organ of metabolic
changes in the organism; changes in the fatty−acid
content were thus more pronounced in this organ
than in the others. The observed changes in the
content of individual fatty acids from the main
metabolic pathways in the investigated organs
were probably a consequence of the inhibitory
effect of quercetin on the activities of ∆−6 desat−
urase and elongases. Especially the blockade of
C18 = 2 acid metabolism to fatty acids with
a higher number of unsaturated bonds and longer
hydrocarbon chains, e.g. arachidonic acid, was
explicitly visible. The consequence of linoleic acid
accumulation included a 1.5−fold decrease in the
content of arachidonic acid in the liver and plasma.
In the remaining organs the increase in C18 = 2
acid content was not accompanied by an altered
content of C20 = 4 acid. The above changes con−
cerned only animals on a diet containing fresh oil.
The C20 = 4/C18 = 2 index determined in the plas−
ma reflects the metabolism of acids in the liver and
informs about the activity of enzymes participat−
ing in the biosynthesis of arachidonic acid from
linoleic acid [30]. The metabolism of linoleic acid
and arachidonic acid in the hearts, kidneys, and
lungs had a similar course.
Under the experimental conditions, inhibition
of the metabolism of acids belonging to the n−6
and n−3 pathway led to a preference for the n−9
pathway of acid synthesis. This was a conse−
quence of metabolic competition for enzymes par−
ticipating in the metabolism of acids from these
pathways. [31, 32]. Linoleic and α−linolenic acids,
substrates of the metabolism of acids from the n−6
and n−3 family, must be delivered with diet, while
precursors for the synthesis of the n−9 pathway
acids may come from resynthesis in the organism,
as mammals’ cells are able to produce only those
unsaturated fatty acids which have the double−
bond situated near carbon 9, starting from the
methyl group. A high content of linoleic acid,
γ−linolenic, dihomo−γ−linolenic, or arachidonic
acids in the diet leads to the inhibition of oleate
desaturation, which results from a competition for
∆−6 desaturase [31].
Lack of an effect of quercetin in rats fed lard
(except for C20 = 2 metabolism) and its presence
in the groups receiving oil was probably associat−
ed with the permeability of cell membranes, which
correlates with the composition of dietary fat. The
high content of polyunsaturated fats in sunflower
oil increased its liquidity and the permeability of
cell membranes, which enabled quercetin to pene−
trate into the cells and exert a biogenic effect.
Changes in the content of fatty acids in the organs
of the animals on diet with fresh oil and quercetin
were more pronounced than those in the rats
receiving oxidized oil, probably due to the fact that
the fatty−acid polymers which were formed during
oxidation decreased the digestion of fat.
From available literature it can be seen that
quercetin affects the metabolism of fatty acids by
changing the activity of numerous enzymes, e.g.
elongase and desaturase, lipooxygenase, cyclooxy−
genase. Information concerning the direction of
fatty−acid metabolism is ambiguous and it seems to
depend on the experimental conditions [33–35].
Factors which may affect the range and direction of
metabolism include the kind and amount of dietary
fat, the kind and amount of flavonoid, the animal
species, as well as the duration of the experiment.
The above factors caused the changes in fatty−acid
content in the livers of the experimental animals
observed under the effect of bioflavonoid by other
authors to be ambiguous [33, 36, 37]. The content
of C18 and C16 acids in the liver of rats receiving
a diet with fat rich in polyunsaturated fats
decreased, while the content of C18 = 2 and C20 =
The Influence of Quercetin on Fatty−Acid Content in Selected Rat Organs
4 acids increased [33]. Changes in the content of
these acids were similar to those observed in the
present experiment, except for C20 = 4.
In the present experiment a blockade of n−6
and n−3 pathway of acid metabolism under the
effect of quercitin, with a preference for the syn−
thesis of n−9 pathway acids, was observed.
A similar effect of the quality of fat (fresh or
oxidized) on fatty−acid content in the livers of the
animals as observed in the present experiment was
described by Lopez−Varela et al. [36] and
Ammouche et al. [37]. The observed changes in
the fatty−acid content correlated positively with
the content of the polar fraction in the oxidized fat.
The composition of fatty acids in numerous
25
tissues, especially in the liver, reflects their content
in the diet [2]. Such an effect of oxidized oil on
fatty−acid content was not observed in the present
experiment.
In conclusion, quercetin, a natural component
of diet or its supplement, affects fatty−acid metab−
olism in the organism on consumption of fat rich
in polyunsaturated fatty acids. Polyunsaturated
fatty acids, which become components of cell
membranes, enable quercetin to penetrate into tis−
sue cells and exert a biogenic effect. Patients
administered pharmaceutical products containing
quercetin and also consuming products which con−
tain quercitin should therefore be recommended to
ingest oils or soft margarine.
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Address for correspondence:
Bożena Regulska−Ilow
Department of Food Science and Nutrition
Silesian Piasts University of Medicine
Nankiera 1
50−140 Wrocław
Poland
Tel.: +48 71 78 40 209
E−mail: [email protected]
Conflict of interest: None declared
Received: 16.01.2008
Revised: 25.01.2008
Accepted: 7.02.2008