Low Plasma Lycopene Concentration Is Associated With Increased Intima-Media Thickness of the Carotid Artery Wall

Low Plasma Lycopene Concentration Is Associated With
Increased Intima-Media Thickness of the
Carotid Artery Wall
Tiina Rissanen, Sari Voutilainen, Kristiina Nyyssönen, Riitta Salonen, Jukka T. Salonen
Abstract—Although a number of epidemiological studies have evaluated the association between ?-carotene and the risk
of cardiovascular diseases, there has been little research on the role of lycopene, an acyclic form of ?-carotene, with
regard to the risk of cardiovascular disease. We investigated the relationship between plasma concentrations of lycopene
and intima-media thickness of the common carotid artery wall (CCA-IMT) in 520 middle-aged men and women (aged
45 to 69 years) in eastern Finland. They were examined from 1994 to 1995 at the baseline of the Antioxidant
Supplementation in Atherosclerosis Prevention (ASAP) study, a randomized trial concerning the effect of vitamin E and
C supplementation on atherosclerotic progression. The subjects were classified into 2 categories according to the median
concentration of plasma lycopene (0.12 ?mol/L in men and 0.15 ?mol/L in women). Mean CCA-IMT of the right and
left common carotid arteries was 1.18 mm in men and 0.95 mm in women with plasma lycopene levels lower than the
median and 0.97 mm in men (P?0.001 for difference) and 0.89 mm in women (P?0.027 for difference) with higher
levels of plasma lycopene. In ANCOVA adjusting for cardiovascular risk factors and intake of nutrients, in men, low
levels of plasma lycopene were associated with a 17.8% increment in CCA-IMT (P?0.003 for difference). In women,
the difference did not remain significant after the adjustments. We conclude that low plasma lycopene concentrations
are associated with early atherosclerosis, manifested as increased CCA-IMT, in middle-aged men living in eastern
Finland. (Arterioscler Thromb Vasc Biol. 2000;20:2677-2681.)
Key Words: lycopene ? atherosclerosis ? carotid arteries ? nutrition ? carotenoids
Low circulating levels of carotenoids have been presumed creased oxidative modification of LDL11 may be one of the
to play a role in atherogenesis. There is some supporting
mechanisms by which lycopene could reduce the risk of
data from studies within populations that have indicated that
coronary heart disease (CHD) and atherosclerotic
high levels of carotenoids in blood1,2 or adipose tissue3,4 are
progression.
associated with decreased risk of cardiovascular disease
Very few previous studies have dealt with the association
(CVD). A few studies have evaluated the association between
between a low concentration of plasma lycopene and early
?-carotene and the risk of CVD, and the results have been
atherosclerosis, as manifested by increased intima-media
inconsistent.4,5 Moreover, there has been little interest in the
thickness of the common carotid artery wall (CCA-IMT). The
role of lycopene, an acyclic form of ?-carotene, with regard
purpose of the present study was to test the hypothesis that
to CVD risk. Like other carotenoids, lycopene is a natural
healthy men and women with decreased levels of serum
pigment synthesized by plants and microorganisms but not by
lycopene have increased thickness of the CCA-IMT.
animals.6 Lycopene gives the familiar red color to tomato
products and is one of the major carotenoids in the diet of
Methods
Europeans and North Americans. Lycopene, which is an
antioxidant carotenoid without provitamin A activity, has
Study Design and Population
been shown to be a more potent antioxidant than ?- or
The Antioxidant Supplementation in Atherosclerosis Prevention
?-carotene.7 In the test tube, lycopene in LDL is used before
(ASAP) study is a randomized, double-masked, placebo-controlled,
?-carotene, lutein, zeaxanthin, or cryptoxanthin in copper-
2?2 factorial trial concerning the effect of vitamin E and C
supplementation in the prevention of atherosclerotic progression in
induced LDL oxidation reaction.8 Men who have high titers
smoking and nonsmoking men and postmenopausal women.12 The
of autoantibodies against oxidatively modified LDL and
study was approved by the Research Ethics Committee of the
those with elevated serum 7?-hydroxycholesterol levels have
University of Kuopio. All study subjects gave their written informed
accelerated progression of carotid atherosclerosis.9,10 A de-
consent.
Received March 27, 2000; revision accepted July 4, 2000.
From the Research Institute of Public Health, University of Kuopio (T.R., S.V., K.N., R.S., J.T.S.), Kuopio, Finland, and Inner Savo Health Centre
(J.T.S.), Finland.
Correspondence to Prof J.T. Salonen, Research Institute of Public Health, University of Kuopio, PO Box 1627, 70211 Kuopio, Finland. E-mail
jukka.salonen@uku.fi
© 2000 American Heart Association, Inc.
Arterioscler Thromb Vasc Biol. is available at http://www.atvbaha.org
2677

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2678
Arterioscler Thromb Vasc Biol.
December 2000
The subjects were regularly smoking (?5 cigarettes/d) or non-
determined with a plasma pool analyzed in 14 separate batches. The
smoking men and postmenopausal women aged 45 to 69 years with
coefficient of variation for lycopene was 10.1%.
a serum cholesterol concentration of ?5.0 mmol/L at a screening
Dietary intake of foods and nutrients was assessed at baseline by
visit. Subjects were not entered into the trial if they had premeno-
4-day instructed food recording. Instructions were given, and com-
pause or regular oral estrogen substitution therapy, regular intake of
pleted food recordings were checked by a nutritionist. The intake of
antioxidants, acetylsalicylic acid or any other drug with antioxidative
nutrients was calculated by use of NUTRICA version 2.5 software.
properties, severe obesity (body mass index ?32 kg/m2), type 1
The data bank of NUTRICA is complied by using mainly Finnish
diabetes, uncontrolled hypertension (diastolic blood pressure
values of nutrient composition of foods. All nutrients were adjusted
?105 mm Hg), any condition limiting mobility (making study visits
for dietary energy intake by use of the residual method. Energy
impossible), severe disease that could shorten life expectancy, or
adjustment is based on the rationale that a larger, more physically
other disease or condition worsening the adherence to the measure-
active person requires a high caloric intake, which is associated with
ments or treatment. The recruitment of the subjects was carried out
a higher absolute intake of all nutrients. Therefore, energy adjust-
in 1993 and the spring of 1994. For the present study, blood for
ment takes into account differences in energy requirements among
lycopene and other chemical measurements was drawn before
individuals. The residuals were standardized by the mean nutrient
antioxidant supplementation. Subjects came to the baseline visits at
intake of a subject consuming 10 MJ/d, the approximate average
the Research Institute of Public Health, University of Kuopio,
total energy intake in the present study population.
between October 1994 and October 1995.
Serum cholesterol was determined from fresh samples with an
enzymatic colorimetric method (Konelab). Serum LDL cholesterol
Ultrasonographic Assessment of CCA-IMT
was measured after precipitation with polyvinyl sulfate (Boehring-
er-Mannheim), and HDL cholesterol was measured from supernatant
Two identical Biosound Phase 2 systems were used (Biosound)
after magnesium chloride dextran sulfate precipitation. Serum tri-
equipped with a 8- to 10-MHz annular array transducer, with a
glycerides were measured colorimetrically (Boehringer-Mannheim).
measurement precision of 0.03 mm. The scannings were videotaped
Plasma total homocysteine (tHcy) concentration was determined
with a PAL S-VHS Panasonic AG 7330E VCR. The ultrasono-
with HPLC.15 Blood pressure was measured manually with the
graphic examinations were carried out by 3 well-trained ultrasound
subject in a sitting position after a rest of 10 minutes; there were 3
technicians. The ultrasonographic scanning of the common carotid
measurements at 3-minute intervals. The body mass index was
arteries (CCAs), the carotid bulbs, and the proximal internal carotid
computed as the ratio of weight to the square of height (in kilograms
artery was performed after a supine rest of 10 minutes, the subject in
per square meter).
the supine position. First, a diagnostic examination of the entire
accessible carotid tree was performed to find the most severe lesions.
Statistical Analysis
Second, the site of the greatest IMT at baseline in the CCA far wall
was located and scanned thoroughly from 3 angles: anterolateral,
Data were analyzed by using either SPSS statistical software for the
lateral, and posterolateral. IMT measurements from videotapes were
IBM RS/6000 workstation or SPSS 9.01 for Windows 98. Mean age,
made at the same site and angle at all examinations of each subject,
ratio of waist-to-hip circumference, systolic blood pressure, diastolic
which was the site with the greatest IMT (in any angle) that was
blood pressure, serum triglycerides, serum HDL cholesterol, LDL
clearly visible at baseline in the far wall of in CCA below the bulb.
cholesterol, total cholesterol, and intake of nutrients are reported as
At this location, IMT was measured in diastole for a length of 10 mm
mean?SD, and cigarette smoking is reported as a percentage.
(or shorter, if not visible) in 1 angle for the far wall. Most often, this
Subjects were classified into 2 categories according to their concen-
was the distal centimeter of the CCA.
trations of plasma lycopene and ?-carotene. We compared the
Computer analysis of ultrasound images to measure IMT was
higher-than-median level with the lower level. The statistical signif-
performed with a reading station equipped with Data Translation DT
icance of differences between these 2 lycopene groups in the main
2861 video frame grabber interfaced to a Panasonic AG 7355 VCR.
characteristics of the subjects was studied by the Student t test.
The association between plasma lycopene and ?-carotene and
Prosound software, developed by Robert Selzer (University of
ultrasonographically assessed CCA-IMT was tested for statistical
Southern California, Los Angeles), with automated boundary detec-
significance also by using ANCOVA, adjusting for age, smoking,
tion was used. IMT was determined as the average difference at, on
serum triglycerides, serum HDL and LDL cholesterol, plasma
average, 100 points between the intima-lumen and media-adventitia
concentration of tHcy, systolic blood pressure, ultrasound observer,
interface.13 The “mean IMT” was computed as the mean of ?100
and intake of 4 nutrients (saturated fatty acids, vitamin C, vitamin E,
IMT measurements in the right CCA and another 100 measurements
and fiber). All tests were 2-tailed, and a value of P?0.05 was
in the left CCA.
considered significant.
For measurement variability, 3 technicians scanned 10 subjects
twice during a week in 1995. The videotapes from all scannings were
read by 1 observer. The repeat correlations for the mean CCA-IMT
Results
were 0.988, 0.995, and 0.998, and pairwise interobserver correlations
The mean?SD concentration of plasma lycopene in the study
were 0.975, 0.983, and 0.995. Computer analysis of ultrasound
population was 0.16?0.11 ?mol/L. Mean plasma levels of
images to measure IMT was performed by use of a PC with a Data
lycopene were higher in women (0.17?0.11 ?mol/L) than in
Translation DT 2861 video frame grabber interfaced to a Panasonic
men (0.14?0.12 ?mol/L, P?0.007 for difference). The
AG 7355 VCR. The Prosound software developed by Robert Selzer
was used.
plasma concentration of lycopene ranged from 0 to 0.62
?mol/L in men and from 0 to 0.64 ?mol/L in women.
Chemical and Dietary Measurements
The main characteristics of the subjects are presented in the
EDTA blood samples were obtained from subjects between 8:00 and
Table. For men, age (P?0.001), plasma concentration of
10:00 AM after an overnight fast. Subjects were instructed to abstain
tHcy (P?0.004), and dietary vitamin C (P?0.010) had a
from alcohol for at least 1 week before the visit. The subjects were
statistically significant difference between the men who had
also instructed to avoid strenuous exercise during the previous 24
plasma levels of lycopene lower than the median (?0.12
hours. Plasma and serum was separated within 60 minutes and stored
at ?80°C until analyzed. Plasma for lycopene determination was
?mol/L) and those who had values higher than the median
extracted with ethanol and hexane; the measurements were con-
(?0.12 ?mol/L). The women with lower levels of plasma
ducted by reversed-phase high-performance liquid chromatography
lycopene (?0.15 ?mol/L) differed significantly with regard
(HPLC) with diode-array UV detection in samples that had been kept
to age (P?0.001), HDL cholesterol (P?0.001), systolic
at ?80°C for 3 to 15 months.14 With this method, the limit of
blood pressure (P?0.016), plasma concentration of tHcy
detection for lycopene was 0.01 to 0.02 ?mol/L. In the statistical
analysis, values below the limits of detection for the assay batch
(P?0.010), and dietary vitamin C intake (P?0.016) from
were marked as 0.00 ?mol/L. The coefficients of variation were
women with higher levels of plasma lycopene. Mean IMT of

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Rissanen et al
Lycopene and Thickness of Carotid Artery Wall
2679
Main Characteristics of Subjects
Plasma Lycopene, ?mol/L
Men (n?256)
Women (n?264)
P for
P for
?0.12
?0.12
Difference
?0.15
?0.15
Difference
n
136
120
144
120
Age, y
61.6?5.0
58.3?6.2
?0.001
61.1?4.6
58.1?5.8
?0.001
Body mass index, kg/m2
26.0?3.0
26.1?2.8
0.901
26.4?3.5
25.8?2.9
0.173
Waist-to-hip ratio
0.95?0.05
0.95?0.05
0.935
0.82?0.05
0.82?0.04
0.900
Systolic blood pressure, mm Hg
135.3?17.7
131.6?18.8
0.103
136.1?20.8
130.2?18.4
0.016
Diastolic blood pressure, mm Hg
79.8?8.7
80.9?9.0
0.304
78.7?9.5
77.8?8.3
0.401
Serum total cholesterol, mmol/L
6.30?0.92
6.36?0.88
0.542
6.44?1.00
6.54?1.09
0.454
Serum HDL cholesterol, mmol/L
1.11?0.27
1.16?0.30
0.165
1.31?0.34
1.48?0.36
?0.001
Serum LDL cholesterol, mmol/L
4.56?0.96
4.56?0.89
0.997
4.53?1.00
4.42?1.12
0.428
Serum triglycerides, mmol/L
1.65?0.76
1.64?0.92
0.897
1.58?0.90
1.52?0.71
0.721
Serum homocysteine, ?mol/L
11.09?2.56
10.10?2.51
0.004
9.74?2.39
9.02?2.08
0.010
Dietary saturated fatty acids, E %
16.6?4.5
15.7?3.7
0.072
15.6?3.6
15.3?3.5
0.416
Dietary fiber, g/d*
21.9?6.7
20.3?7.4
0.065
20.6?4.6
21.1?5.3
0.430
Dietary vitamin C, mg/d*
80.6?52.4
100.2?66.3
0.010
94.6?53.8
113.7?73.9
0.016
Dietary vitamin E, mg/d*
9.5?4.6
9.6?3.6
0.826
9.7?2.4
9.8?2.3
0.691
Smoking, %†
53
41
0.053
49
47
0.671
IMT, mm
1.18?0.39
0.97?0.20
?0.001
0.95?0.25
0.89?0.19
0.027
Values are mean?SD. E% indicates percentage of energy.
*Adjusted for energy intake.
†Proportion of each group of plasma levels of lycopene.
the right and left CCA was 1.18 mm in men and 0.95 mm in
terol, plasma tHcy, and systolic blood pressure), ultrasound
women with low plasma lycopene levels and 0.97 mm in men
observer, and intake of 4 nutrients (proportion of saturated
(P?0.001 for difference) and 0.89 mm in women (P?0.012
fatty acids of total daily energy, vitamin C, vitamin E, and
for difference) with higher plasma levels of lycopene. In men
fiber), low plasma lycopene levels were associated with
in the highest quarter of plasma lycopene level, the CCA-IMT
17.8% increased IMT in men compared with plasma levels of
was 17.5%, and in women, it was 5.6% lower than in men and
lycopene higher than median (P?0.003 for difference). In
women in the lowest quarter of plasma lycopene level
women, the difference did not remain significant after the
(Figure).
adjustments.
In ANCOVA, adjusting for other cardiovascular risk fac-
We conducted analyses for ?-carotene similar to those
tors (age, serum triglycerides, serum HDL and LDL choles-
presented above for lycopene. In ANCOVA, adjusting for
risk factors, there was no statistically significant association
between plasma concentration of ?-carotene and IMT.
We also repeated analysis in smokers and nonsmokers. The
association between plasma levels of lycopene and IMT
seems to be stronger among nonsmokers than among smok-
ers. Adjusted IMT was significantly higher (P?0.006) in
nonsmokers and nonsignificantly higher (P?0.070) in smok-
ers with lower plasma levels of lycopene compared those
with higher plasma lycopene levels. The association between
the plasma level of ?-carotene and the IMT did not differ in
smokers and nonsmokers.
We also divided subjects into 2 categories according to
IMT (?1.00 mm and ?1.00 mm). The mean level of plasma
lycopene was 0.16 ?mol/L in men and 0.17 ?mol/L in
women with lower IMT and 0.12 ?mol/L in men (P?0.003
for difference) and 0.16 in women (P?0.225 for difference)
with higher IMT.
Mean IMT of carotid artery wall in quarters of plasma lycopene
concentration in middle-aged Finnish men and women. Plasma
Discussion
lycopene concentration quarters for men were ?0.06, 0.06 to
Our main finding is that in middle-aged men but not in
0.11, 0.12 to 0.18, and ?0.18 ?mol/L; for women, they were
?0.09, 0.09 to 0.15, 0.16 to 0.22, and ?0.22 ?mol/L.
middle-aged women from eastern Finland, a low concentra-

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2680
Arterioscler Thromb Vasc Biol.
December 2000
tion of plasma lycopene is associated with early atheroscle-
The oxidative modification of LDL particles may play a
rosis, as manifested by increased thickness of the CCA wall.
role in the formation of foam cells, atherosclerotic lesions,
Two previous studies have dealt with the concentration of
and CHD.9,23 Antioxidants can inhibit the oxidative modifi-
blood lycopene and the thickness of the artery wall. In the
cation of LDL, may retard atherosclerotic progression, and,
Atherosclerosis Risk in Communities (ARIC) study,16 there
consequently, prevent clinical complications of atherosclero-
were 231 age-matched, sex-matched, race-matched, and field
sis, such as MI.24,25 Lycopene and other carotenoids have
center–matched case-control pairs selected from the larger
been shown to act as antioxidants.7,11,26,27 Carotenoids found
study population. Cases exceeded the 90th percentile of IMT
in plasma can quench singlet oxygen, a potential initiator of
in the cohort, and the control subjects were below the 75th
lipid peroxidation.26 Lycopene, the open-chain isomer of
percentile of IMT for all arterial segments. The cases had
?-carotene, exhibits the highest physical quenching rate
nonsignificantly lower levels of serum lycopene and ?- and
constant of all carotenoids with singlet oxygen.7
?-carotene. The odds ratio of being above the 90 percentile of
The mean CCA-IMT in the present subjects was somewhat
IMT, related to low serum levels of lycopene, after adjusting
higher than that reported in most other studies. In the ARIC
for risk factors was 0.81 (95% CI 0.60 to 1.08). In the ARIC
study,16 the mean IMT of the carotid artery was 1.2 mm for
study,17 high dietary intake of provitamin A carotenoids was
the cases and 0.6 mm for the controls. In the Perth Carotid
associated with lower prevalence of carotid plaques, but this
Ultrasound Disease Assessment Study (CUDAS),28 the aver-
association was not statistically significant. The dietary lyco-
age mean of carotid artery IMT in men was 0.73 mm. In the
pene intake was not assessed. In the prospective Rotterdam
EVA study,19 the mean IMT was 0.70 in men and 0.65 in
Study18 on serum carotenoids and atherosclerosis, serum
women. In the Cardiovascular Health Study,21 the mean
lycopene was the only carotenoid associated with decreased
CCA-IMT was 1.03 mm in subjects aged ?65 years; thus, the
subjects were ?10 years older than in the present study. In
risk of aortic atherosclerosis (odds ratio 0.55). However, the
the population in eastern Finland, CCA-IMT seems to be
association was nonsignificant. In the Etude sur le Vieillise-
rather high. This is consistent with the high occurrence of
ment Artériel (EVA) Study,19 in the highest quarter of plasma
clinical CHD in eastern Finland. Moreover, the serum LDL
total carotenoids was associated with lower CCA-IMT in
cholesterol in the present study was higher than that in other
women (P?0.014 for a linear trend between quarters) and
studies, and half of the present study subjects smoked
men (P?0.02 for a linear trend between quarters), but after
regularly. The mean plasma concentration of lycopene in the
adjustment this association was nonsignificant. Plasma levels
present study was lower than that in other population-based
of lycopene were not measured.
studies1,16,29; in only 1 earlier study18 was the mean level of
Common carotid plaques and increased IMT have been
plasma lycopene similar to that in the present study. Evi-
shown to predict coronary events.20,21 The association be-
dently, the consumption of tomatoes or tomato products in
tween blood or tissue concentration of lycopene and risk of a
Finland may be lower than that in most other European
CHD has been studied in a cross-sectional study. In a
countries.
multicenter European Community Multicenter Study on An-
It is possible that the plasma level of lycopene could be an
tioxidants, Myocardial Infarction, and Breast Cancer (EU-
indicator for other favorable dietary factors. However, the
RAMIC) study,3 subjects who had suffered a myocardial
effect of lycopene was significant after an adjustment for 3
infarction (MI) had lower concentrations of lycopene in
other dietary factors, the intakes of saturated fatty acids, fiber,
adipose tissue than did the control subjects. In a nested
and vitamin C. Also, smokers had lower plasma levels of
case-control study22 that compared subjects who developed
lycopene than did nonsmokers. This could be due to either
MI with healthy control subjects, the cases had lower serum
dietary differences or a consequence of smoking itself.
concentration of lycopene than did the control subjects, but
However, we adjusted for smoking in our statistical analysis
the difference was not significant. Only in smokers was a low
to shown that the increased CCA-IMT in subjects with low
serum level of lycopene associated with a significantly
plasma levels of lycopene was not simply a consequence of a
increased risk of subsequent MI.
higher proportion of smokers.
In the present study, the association between the plasma
In conclusion, the present study shows that low levels of
level of lycopene and IMT appeared to be stronger among
plasma lycopene are associated with increased CCA-IMT in
nonsmokers than among smokers. We found that an associ-
middle-aged men, but not in women, from eastern Finland.
ation between the plasma level of lycopene and IMT exists in
Because increased IMT has been shown to predict coronary
nonsmokers and is weak in smokers. There are 2 explanations
events,20,21 this finding suggests that the plasma level of
for this difference. First, mean daily intake of carotenoids
lycopene, particularly a biomarker of tomato-rich food intake,
could be lower in smokers than in nonsmokers. An alternative
may play a role in the early stages of atherogenesis. Conse-
explanation is that because smoking is such a strong risk
quently, our results support the previous evidence that plant-
factor itself, smokers do not benefit from lycopene intake as
dominated diet contributes to cardiovascular health.
much as nonsmokers, or they have a greater need of
antioxidants.
Acknowledgments
The effect of lycopene on IMT was different in women
This study was partly funded by the Finnish Academy and the
than in men. There are some possible causes of this differ-
Juho Vainio Foundation. We are grateful to our staff of almost 30
people and Oy Jurilab Ltd for helping with data collection.
ence. Women have a better diet than do men. This is the most
likely explanation for the lack of association with IMT.
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