![[Feedback]](/icons/banner/feedbackACT.gif){kind=icon}
![[For Subscribers]](/icons/banner/subscriberACT.gif){kind=icon}
![[Archive]](/icons/banner/archiveACT.gif){kind=icon}
![[Search]](/icons/banner/searchACT.gif){kind=icon}
![[Contents]](/icons/banner/tocACT.gif){kind=icon}
|
|
|
|
|
||
Department of Clinical Nutrition, University of Kuopio, 70211 Kuopio, Finland
2To whom correspondence should be addressed.
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
KEY WORDS: • plant stanol ester • plant sterol • sitostanol • cholesterol • humans
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
). In addition, other forms of plant sterols have been used
up to 53 g (Pollak and Kritchevsky 1981
). In some
previous studies, sitostanol has been found to lower serum cholesterol
concentrations more effectively than sitosterol (Becker et al. 1993
, Heinemann et al. 1986
). Several studies
have shown that about 2.0–3.0 g of sitostanol from full-fat
sitostanol ester margarines or mayonnaises significantly reduces serum
total and LDL cholesterol concentrations without affecting serum HDL
cholesterol or triglyceride concentrations (Gylling et al. 1995
, and 1997
, Gylling and Miettinen 1994
, Miettinen et al. 1995
, Niinikoski et al. 1997
, Vanhanen et al. 1994
,
Weststrate and Meijer 1998
). Furthermore, it has been
reported that plant sterols reduce cholesterol concentrations within
2–3 wk of initiation of treatment (Jones et al. 1997
).
Previous studies on plant sterols from different origins suggest at
least 0.8–1 g/d should be consumed to have a significant effect on
serum lipids (Hendriks et al. 1999
, Miettinen and Vanhanen 1994
, Vanhanen et al. 1994
). In the
study of Miettinen et al. (1995)
, the cholesterol-lowering effect
of 2.6 g of sitostanol was slightly, but significantly greater
than that of 1.8 g of sitostanol. But it is still unclear, what
the dose is beyond which no apparent additional benefit can be
obtained. Previously no controlled primary dose-response study with
several different doses of >98.5% esterified plant stanols has been
done.
Plant stanols and sterols may interfere with the absorption of
fat-soluble vitamins and carotenoids while lowering serum
cholesterol concentrations (Gylling et al. 1996
,
Hallikainen and Uusitupa 1999
, Hendriks et al. 1999
, Weststrate and Meijer 1998
). Therefore, it
is important to investigate whether stanol ester dose, having
significant cholesterol-lowering effect, has an effect on serum
carotenoids and fat-soluble vitamin concentrations as well.
Finally, the aim of this dose-response study was to determine the dose-response curve for serum total and LDL cholesterol during different doses of plant stanols (0–3.2 g) added as plant stanol esters into rapeseed oil-based margarines. In addition, we investigated the effects of different doses of stanol ester on serum plant sterol concentrations to obtain information on absorption and bioavailability of plant sterols and stanols.
|
METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
Altogether 26 subjects (men and women) were recruited to the study from
former studies carried out at the Department of Clinical Nutrition,
University of Kuopio and from the local society of the Finnish Heart
Association. The main inclusion criteria were as follows: serum total
cholesterol 5.0–8.5 mmol/L and total triglycerides below 3.5 mmol/L
after the pretrial period, ages 25–65 y, normal liver, kidney and
thyroid function, willingness to participate, no lipid-lowering
medication, no unstable coronary heart disease, no alcohol abuse (>45
g of ethanol/d) and no irregular eating habits. Four subjects dropped
out during the study: one during the pretrial period due to personal
reasons, one during the first dose period due to prolonged constipation
for which medication possibly affecting serum lipids (a plantago ovata
product, Visiblin®) was prescribed, one during the second-dose
period due to prolonged infection (bronchitis, stomatitis) and one
during the fourth-dose period due to prostatitis. One subject had a
hormone-releasing intrauterine device, one subject used hormone
substitution medication, four subjects used postmenopausal hormone
replacement therapy, and one subject used a calcium channel blocker and
one subject renin-angiotensin system affecting medication for the
treatment of hypertension, but he stopped the medication in the middle
of the third-dose period. One of the subjects was a smoker. The
subjects were requested to maintain their medication, weight, alcohol
consumption, smoking habits and physical activity constant during the
entire study. Baseline characteristics of 22 subjects are shown in
Table 1
.
|
Study design.
The study was carried out from January to June 1998 at the Department of Clinical Nutrition, University of Kuopio with a randomized single-blind, repeated measures design. After a 1-wk pretrial period, all subjects consumed five different doses of plant stanol added as plant stanol ester into the rapeseed oil-based margarine. Each dose was taken for 4 wk in the same order. The order of dose periods was randomly determined and was as follows: 2.4, 3.2, 1.6, 0 (control) and 0.8 g.
Routine laboratory measurements were taken to ensure normal health status at the first and at the last visit of the study. In addition, previous and present diseases, current medication, alcohol and tobacco consumption, physical activity, use of vitamins or other nutrient supplements were interviewed by a structured questionnaire at the first visit of the study. Alcohol and tobacco consumption and physical activity were reviewed also at the last visit to the study unit. Furthermore, possible changes in diseases, medication, use of vitamin or nutrient supplements were recorded during the study. Blood samples were taken from fasting subjects at the beginning of the pretrial period (-1 wk), at the beginning of the first dose period (0 wk) and at the middle and the end of each period (2, 4, 6, 8, 10, 12, 14, 16, 18 and 20 wk). A study period of 4-wk long was used to achieve the steady state with the present dose. Body weight was recorded at each visit. The possible adverse effects and symptoms were interviewed using a structured questionnaire at the end of each dose period.
Diet.
The composition of low erucic acid rapeseed oil-based margarines
(Raisio Group Plc., Raisio, Finland) is presented in Table 2
. The total amount of fat in the test margarines ranged between 70 and
81% and the amount of absorbable fat 68 and 70%. Added plant stanol
ester-containing spreads were prepared using commercially available
plant sterols by recrystallization, hydrogenation to form plant stanols
and esterification to produce fatty acid esters of the obtained plant
stanols. The daily dose of the test margarine was 25 g taken in
two to three portions with meals. The daily amounts of total stanols
based on the actual amount of stanols in the test spreads were 0 g
containing no added stanols, 0.81 g (planned 0.8 g)
consisting of 0.62 g sitostanol and 0.19 g campestanol,
1.56 g (planned 1.6 g) consisting of 1.19 g sitostanol
and 0.37 g campestanol, 2.29 g (planned 2.4 g)
consisting of 1.74 g sitostanol and 0.55 g campestanol and
3.03 g (planned 3.2 g) consisting of 2.30 g sitostanol
and 0.73 g campestanol. During the pretrial period, the spread did
not contain added stanols. Vitamin A (4.45 µg retinol equivalents/g)
and vitamin D (0.064 µg/g) were added to each spread.
|
Subjects followed a standardized background diet throughout the study.
The composition of the background diet resembled the habitual diet of
the subjects and was the following: 34% of energy
(E%)3
from fat including <12 E% saturated, 14 E% monounsaturated and 8 E%
polyunsaturated fat, and 23.8 mg/MJ dietary cholesterol. The subjects
received oral and written instructions on the diet by food groups at
their own energy level. The diet plan included precise amounts and
quality of fat and cheese (low-fat), and only the precise quality of
liquid milk (fat-free or low-fat) and meat products (low-fat). The diet
plan was made for eight energy levels (6.7–12.6 MJ/d). Energy
requirement of the subjects was estimated according to
Harris-Benedict formula to which the energy needs as a result of
physical activity were added (Alpers et al. 1986
). The
feasibility of the background and test diets was improved by providing
test margarines, rapeseed oil, salad dressing and low-fat cheese
for the subjects free of charge.
If the subject’s habitual diet did not meet the goal for the
composition, the diet was modified during the pretrial period.
Adherence to the background diet was monitored by 3-d food records kept
before the end of the pretrial period and by 4-d food records kept
before the end of each dose period. One of the recording days was a
weekend day or the person’s day off from work. The subjects recorded
their food consumption after consulting a booklet containing
photographs of food portions (Haapa et al. 1985
). At
study visits, the amounts and qualities of foods in the records were
checked by the nutritionist for completion, filling in data that were
lacking.
The nutrient intake was calculated using the Micro-Nutrica® dietary
analysis program (version 2.0; Finnish Social Insurance Institute,
Turku, Finland). The food composition database is based on analyses of
the Finnish food and international food composition tables
(Rastas et al. 1993
). In addition, the database was
updated for the purposes of the present study.
Laboratory measurements.
All measurements were done and venous blood samples were obtained after
a 12-h overnight fast by using standardized methods. Body weight was
measured with a digital scale. Since the phase of menstrual cycle may
have an effect on serum cholesterol concentration (Cullinane et al. 1995
) in premenopausal women, the study measurements at
each dose period were performed at d 5–10 of the cycle.
Lipoproteins were separated by ultracentrifugation for 18 h at a
density of 1.006 kg/L to remove VLDL fraction. HDL in the infranatant
was separated from LDL by precipitation of LDL with dextran sulfate and
magnesium chloride (Penttilä et al. 1981
). LDL
cholesterol was calculated as the difference between the mass of
cholesterol in the infranatant and HDL, and VLDL cholesterol was
calculated as the difference between the whole serum and the
infranatant. Enzymatic photometric methods were used for the
determination of cholesterol and triglycerides from whole serum and
separated lipoproteins using commercial kits (Monotest® Cholesterol
and Triglyceride GPO-PAP; Boehringer Mannheim GmbH Diagnostica,
Mannheim, Germany) and a Kone Specific Clinical Analyser (Kone, Espoo,
Finland).
Serum samples for 
- and ß-carotene, lycopene and fat-soluble
vitamins, and apolipoprotein A-I (apo A-I) and B and plant sterols
were stored at -70°°C until analyzed at the
end of the study. Analyses of apo were based on the measurement of
immunoprecipitation enhanced by polyethylene glycol at 340 nm. A Kone
Specific Clinical Analyzer and apo A-I and apo B reagents from Kone
Corporation were used.
Serum - and ß-carotene, lycopene and fat-soluble vitamins were
analyzed by the HPLC system (Perkin-Elmer, Norwalk, CT) equipped with a
C18 column (Waters, Milford, MA) (Driskell et al. 1983
,
Kaplan et al. 1987
, Parviainen 1983
).
Serum plant sterols were measured by gas-liquid chromatograph (HP
5890 Series II, Hewlett Packard, Delaware, Little Falls, Wilmington,
DE) from nonsaponifiable serum material equipped with 50-m long Ultra 1
capillary column (methyl-polysiloxane) [Hewlett-Packard, Little
Falls, DE] for plant sterols and equipped with a 50-m long
Ultra 2 capillary column (phenyl-methyl-siloxane) (Hewlett-Packard) for
sitostanol and campestanol (Miettinen 1988
,
Miettinen and Koivisto 1983
). Serum plant sterols were
determined twice from same samples, and the mean values of two
determinations were used in the statistical analysis.
Plasma glucose was analyzed by enzymatic photometric method using reagent Granutest 100 (Merck, Darmstadt, Germany) with a Kone Specific Clinical Analyser (Kone).
Apo E genotypes were analyzed with the restriction fragment length
polymorphism-polymerase chain reaction method described by
Tsukamoto et al. (1993)
with a slight modification.
Statistical analyses.
All statistical analyses were performed with SPSS for windows 6.0.1 statistics program (SPSS, Chicago, IL). The results are given as means ± SD in text and tables, and as means ± SEM in figure.
The main comparison was made among the mean values at the end of each dose period. In the Results and Discussion sections only these end measurements and their percentage changes are presented. The percentage changes were calculated comparing the end measurements of each dose period to the end measurement of the control period. To eliminate the effects of changes in lipoprotein concentrations, serum carotenoid, tocopherol and plant sterol values are given besides crude concentrations also in terms of mmol/mol of cholesterol, which express ratios to total cholesterol.
Normal distribution of variables was checked with Shapiro Wilks test
before the further analyses (Norusis 1993
). If a
variable was not normally distributed, statistical analysis was made
after logarithmic transformation. Repeated measures ANOVA was used to
compare the overall changes in continuous variables among different
dose periods. Two-tailed comparisons with paired t
test were used in the further analyses. For variables (intake of
alcohol, fiber and vitamin A, and serum -carotene, ß-carotene,
lycopene and campestanol) which were not normally distributed not even
after logarithmic transformation Friedman Two-tailed ANOVA test and
Wilcoxon’s matched-pairs signed rank test or Mann-Whitney test
was used. To control the overall level, Bonferroni adjustment was
used. Wilcoxon’s matched-pairs signed rank test was used to
compare alcohol consumption, smoking habits and physical activity,
which were reviewed by the questionnaires at the beginning and at the
end of the study.
Power of the study was 0.80 based on assumption to be able to detect a
0.4–0.5 mmol/L difference in serum total cholesterol response between
the different doses with the present number of subjects and probability
for type I error = 0.05.
|
RESULTS |
|---|
|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
Baseline characteristics of the subjects are presented in Table 1
.
Blood hemoglobin and thrombocytes, and serum thyroid stimulating
hormone, 
-glutamyl and alanine amino transferase and
creatinine were all within the normal ranges at the beginning and the
end of study. Body mass index did not change significantly during the
study (Table 1)
. Physical activity, alcohol consumption and smoking
habits remained stable according to questionnaires. During the study,
five subjects had gastrointestinal symptoms (stomach pain/discomfort,
flatulence or constipation), and two subjects had skin symptoms
(eczema, itching or dry skin). The symptoms occurred occasionally and
they were not related to the dose of stanol ester.
Feasibility of the diet.
The mean daily consumption of margarine was between 25.2 and 25.5 g during the different dose periods. Thus the actual mean daily intake of stanol was 0.82 ± 0.0 g (0.63 ± 0.0 g sitostanol and 0.19 ± 0.0 g campestanol), 1.59 ± 0.02 g (1.22 ± 0.02 g sitostanol and 0.37 ± 0.0 g campestanol), 2.33 ± 0.05 g (1.77 ± 0.04 g sitostanol and 0.56 ± 0.01 g campestanol) and 3.05 ± 0.09 g (2.32 ± 0.07 g sitostanol and 0.74 ± 0.02 g campestanol) in the 0.8, 1.6, 2.4 and 3.2 g dose periods, respectively.
The actual composition of the diet during the different dose periods is
presented in Table 3
. There were no significant differences in the intake of fat,
monounsaturated and polyunsaturated fatty acids, cholesterol,
carbohydrates nor in the intake of fat-soluble vitamins and
ß-carotene among the different dose periods. However, the intake of
saturated fatty acids was significantly lower (1.8 E%) during the
2.4-g dose period than during the control period, but there were no
significant differences in the intake of saturated fatty acids between
any other two dose periods. Furthermore, the intake of alcohol was
significantly lower (difference 1.6–1.7 E%) during the 3.2, 2.4 and
1.6 g dose periods than during the control period. The intake of
fiber was significantly lower (difference 0.5–0.6 g/MJ) during the
control and 0.8-g dose periods than during the 2.4- and 1.6-g dose
periods.
|
Serum lipids and lipoproteins.
The concentrations of serum lipids, lipoproteins and apo at the end of
each dose period are shown in Table 4
. Figure 1(A
,B
,C)
presents the percentage differences in serum total cholesterol,
LDL cholesterol and apo B compared to the control dose, respectively.
|
|
. There were no
significant differences in the mean concentration of serum total
cholesterol between the 3.2- and the 2.4-g dose periods, between the
2.4- and the 1.6-g dose periods, or between the 0.8-g dose and the
control periods. The mean concentration of the serum total cholesterol
tended to be lower (P = 0.054) at the end of the 3.2-
than at the end of the 1.6-g dose period.
There were no significant differences in the mean concentration of the
serum LDL cholesterol among the 3.2-, 2.4- and 1.6-g dose periods, or
between the dose periods of the 1.6 g and the 0.8 g, or the
0.8 g and the control (Table 4)
. The mean concentration of the
serum LDL cholesterol was significantly lower at the end of 3.2- and
2.4-g dose periods than at the end of the 0.8-g dose and control
periods. Furthermore, serum LDL cholesterol concentration was
significantly lower at the end of the 1.6-g dose than at the end of the
control period. The percentage changes in serum total and LDL
cholesterol concentration calculated in reference to control were
parallel to the changes in mean concentration (Fig. 1)
.
The mean concentration of serum VLDL cholesterol did not differ
significantly among the 3.2-, 2.4-, 1.6- and 0.8-g dose periods (Table 4)
. Furthermore, there was no significant difference in the mean
concentration of serum VLDL cholesterol between the 0.8-g dose and the
control periods. The mean concentration of the serum VLDL cholesterol
was significantly lower at the end of the 3.2-, 2.4- and 1.6-g dose
periods than at the end of the control period.
There were no significant changes in serum HDL cholesterol and total
triglyceride concentrations during the entire study (Table 4)
.
Serum apo B decreased significantly at the dose of 0.8 g (8.7%,
P < 0.001) compared to the control (Fig. 1)
. There
were no significant differences in the mean concentration or percentage
decrease of serum apo B in pairwise comparisons after Bonferroni
correction among the 3.2-, 2.4-, 1.6- and 0.8-g dose periods (Table 4
,
Fig. 1
).
Serum apo A-I concentration did not change significantly during the
study (Table 4)
. Furthermore, there were no significant differences in
serum apo A-I/apo B ratio among the 3.2-, 2.4-, 1.6- and 0.8-g dose
periods. However, apo A-I/apo B ratio was significantly higher at the
3.2-, 1.6- and 0.8-g dose periods than at the control period and in
addition, the apo A-I/apo B ratio tended to be higher (P
= 0.068) at the 2.4-g dose than at the control period.
In a secondary analysis, there were no significant differences in the percentage changes of LDL cholesterol concentration between apo E 3:3 (n = 14) and 3:4 (n = 8) groups (-2.3 vs. -0.6%, -6.2 vs. -4.5%, -9.4 vs. -10.1% and -11.8 vs. -8.1%, apo E 3:3 vs. 3:4, at the 0.8-, 1.6-, 2.4- and 3.2-g dose periods vs. reference to control, respectively).
Plant sterols.
The higher the dose, the lower the serum plant sterol concentration was
(Table 5
). The mean values of serum campesterol were 4.75 ± 3.01 µmol/L
(24.5 ± 11.6%) to 8.67 ± 4.76 µmol/L (44.7 ± 10.4%) lower at the end of the test dose periods compared to the
control period. Serum campesterol concentration was significantly lower
at the end of the 3.2-, 2.4- and 1.6-g dose periods than at the end of
the 0.8-g dose period. Furthermore, serum campesterol concentration was
lower at the end of the 3.2-g dose period than at the end of the 2.4-g
dose period. The changes in campesterol/total cholesterol ratio were
parallel with the changes in absolute serum campesterol concentration
(Table 5)
.
|
.
Serum sitosterol concentrations were 2.21 ± 1.41 µmol/L (25.2
± 10.7%) to 3.83 ± 2.02 µmol/L (43.9 ± 8.7%)
lower at the end of the test dose periods compared to the end of the
control period.
Serum plant stanol concentrations rose with increasing the
dose. However, their concentrations remained very low in serum
throughout the entire study. The serum concentrations of campestanol
were 0.19 ± 0.11 µmol/L to 0.27 ± 0.16 µmol/L greater
at the end of the test dose periods than at the end of the control
period. In addition, serum campestanol concentration was significantly
greater at the end of the 3.2-g dose period than at the of 0.8-g dose
period. The campestanol/total cholesterol ratio was significantly
greater (difference 0.04 ± 0.03–0.06 ± 0.03 mmol/mol of
cholesterol) at the end of all dose periods than at the end of the
control period (Table 5)
. In addition, the ratio was significantly
greater at the end the 3.2-g dose period than at the end of the other
test dose periods.
The serum sitostanol concentration was significantly greater
(difference 0.19 ± 0.21–0.36 ± 0.25 µmol/L) at the end
of all dose periods than at the end of the control period, but among
the test dose periods there were no significant differences in serum
sitostanol concentration (Table 5)
. Changes in the sitostanol/total
cholesterol ratio (increase from 0.05 ± 0.04 to 0.08 ± 0.05
mmol/mol of cholesterol) were parallel to the absolute changes in
sitostanol concentration, except that the ratio was significantly
greater also at the end of the 3.2-g than at the end of the 0.8-g dose
period.
Carotenoids and fat-soluble vitamins.
There were no significant changes in serum retinol, -carotene,
ß-carotene or + ß-carotene concentrations nor their ratios to
the serum total cholesterol concentrations during the different dose
periods (Table 6
).
|
. There were no significant
differences in the serum lycopene concentrations in the men throughout
the study. However, in women, serum lycopene concentrations were
significantly greater at the end of the control and 0.8-g dose periods
than at the end of the 2.4-g dose period (Table 6)
. In women, the
lycopene/total cholesterol ratio was significantly greater at the end
of the 0.8-g dose period than at the end of 2.4-g dose and control
periods (Table 6)
.
Serum -tocopherol concentration was significantly lower at the end
of all experimental dose periods than at the end of the control period.
In addition, serum -tocopherol concentration was significantly lower
at the end of the 3.2-g dose period than at the end of the 0.8-g dose
period (Table 6)
. Serum -tocopherol concentration was significantly
lower only at the end of the 3.2-g and the 2.4-g dose periods than at
the end of the control period (Table 6)
. Furthermore, the changes in
serum +-tocopherol concentration were parallel to the changes in
the serum -tocopherol concentration during the trial (Table 6)
.
However, after relating the serum -, - and +-tocopherol to
the serum total cholesterol concentration, there were no significant
differences among the different periods.
Serum 25-hydroxycholecalciferol concentration was significantly lower
at the end of the control than at the end of the 0.8-g dose period
(Table 6)
. There were no significant differences in serum
25-hydroxycholecalsiferol concentration between any other dose periods
after the Bonferroni correction.
|
DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
Why does the cholesterol-lowering effect of stanol ester seem then
to level off with higher doses? After hydrolysis, the cholesterol
absorption is dependent on micelle formation, and the amount and type
of bile acids influence this micelle formation and consequently
cholesterol absorption. It is believed that the
cholesterol-lowering effect of plant sterols and stanols is based
on their competition with cholesterol incorporation into mixed micelles
(Ikeda and Sugano 1998
). It can be assumed that if there
occurs an excessive amount of plant stanols in the small intestine to
that of cholesterol, no additional benefit can be obtained with
increasing doses of plant stanol esters. In adults, 1000–1500 mg of
cholesterol, biliary and dietary origin, enters the lumen of small
intestine daily. Therefore, the full saturation effect would be reached
with doses of around 2.0–3.0 g of plant stanols, as also suggested by
the present results.
The dose of 0.8 g stanol did not significantly affect serum total
and LDL cholesterol concentrations, but resulted in 8.7% decrease in
apo B concentration in comparison to the control dose, suggesting a
reduction of apo B containing particles even with a low dose of stanol
ester. The results of the present study are parallel to the results of
previous studies, in which sitostanol has reduced total and LDL
cholesterol concentrations, when the ingestion of sitostanol has been
from 1.5 g (Becker et al. 1993
, Heinemann et al. 1986
) up to 3.4 g/d (Gylling and Miettinen 1994
, Gylling et al. 1995
and 1997
,
Hallikainen and Uusitupa, 1999
, Miettinen et al. 1995
, Niinikoski et al. 1997
, Vanhanen et al. 1993
and 1994
, Weststrate and Meijer 1998
).
Our results are also in agreement with earlier studies (Hendriks et al. 1999
, Miettinen and Vanhanen 1994
,
Vanhanen et al. 1994
) in which it has been shown that at
least 0.8–1 g/d of plant sterol should be consumed before clinically
remarkable cholesterol-lowering effects can be observed. Stanol
ester inhibits the cholesterol absorption so that less dietary and
endogenous cholesterol enters via portal circulation the liver.
Depletion of intracellular cholesterol in the liver could result in the
upregulation of LDL receptor activity and consequently cause an
enhanced clearance of apo B containing particles. It has been
hypothesized by Gylling and Miettinen (Gylling and Miettinen 1994
, Miettinen and Gylling 1999
) that decreased
VLDL and IDL cholesterol concentrations caused by removal of these
cholesterol-rich particles also results in decrease of their
conversion to LDL. This phenomenon might explain the small discrepancy
in reduction of LDL cholesterol and apo B concentrations with the dose
of 0.8 g, because one might expect a greater reduction in LDL
cholesterol based on the change in apo B concentration which is the
major apo in LDL particle. The other possibility is that LDL particles
in circulation after ingestion of 0.8 g dose of stanol are more
cholesterol-rich, but the results on VLDL levels in the present
study support the first-described explanation, the enhanced
clearance of apo B containing particles.
Serum campesterol concentration is shown to correlate positively with
intestinal cholesterol absorption (Miettinen et al. 1990
, Tilvis and Miettinen 1986
). In the present
study, serum campesterol, sitosterol and avenasterol concentrations
were significantly lower already at the end of the 0.8-g dose period
compared with the control period, reflecting that plant stanols inhibit
effectively intestinal cholesterol absorption even with the small dose
of stanol ester. Both in former human and animal absorption studies,
sitostanol has been found to be minimally absorbed and campestanol to
some extent (Hassan and Rampone 1979
, Heinemann et al. 1993
, Lütjohann et al. 1993
,
Xu et al. 1999
). In this study, higher serum
concentrations of sitostanol and campestanol at the end of the test
dose periods as compared to the control period indicate that small
amounts of sitostanol and campestanol are absorbed from the intestine.
However, it should be noticed that the absorbed amounts are really
neglible compared to the given dose; at the stanol dose of 0.8 and
3.2 g the increase of serum sitostanol in reference to control was
only about 0.19 µmol/L (80.7 µg/L) and 0.36 µmol/L (149.2
µg/L), respectively. The respective increase in serum campestanol was
0.19 µmol/L (75.2 µg/L) and 0.27 µmol/L (108.3 µg/L). These
results are in-line with the results of Gylling et al. (1999)
.
Besides the negligible absorption of plant stanols, the low-serum
concentrations could also result from the fast and effective clearance
of absorbed stanols.
In the present study, all subjects consumed each test margarine in the
same randomly determined order, and each subject served as his/her own
control. The benefit of the present study design is that it eliminates
the between-individual variation. The order of the dose periods was
randomized to control for systematic bias due to the order of periods.
According to the chosen design, the main comparisons were made between
the mean values at the end of each period. Dose period of a 4-wk
duration can be considered sufficient to eliminate the carry-over
effect of the previous dose period to the next one, and in addition, to
bring out the effects of a given dose on serum cholesterol
concentrations. In earlier studies it has been shown that plant sterols
reduce cholesterol concentrations within 2–3 wk of initiation of
treatment (Jones et al. 1997
). That is also in agreement
with our previous study (Hallikainen and Uusitupa 1999
).
On the other hand, the serum cholesterol concentration returned to an
initial value within 2–3 wk, upon cessation of the ingestion of plant
sterols (Farquhar et al. 1956
, Heinemann et al. 1986
).
The differences in the nutrient intake among the different dose periods were occasional and minor. Thus the differences in lipid responses among the dose periods can be ascribed to the differences in the amount of active compound rather than differences in background diet. Furthermore, body mass index did not change significantly during the study.
Serum VLDL cholesterol concentration was significantly lower at the
3.2-, 2.4- and 1.6-g dose periods compared with the control period. The
significant decrease could be due to the effect of plant stanols, but
it could more likely be ascribed to slightly, but significantly, higher
alcohol consumption, which might have increased VLDL cholesterol
concentration at the end of the control period (Steinberg et al. 1991
). Temporary increased alcohol consumption was probably due
to the fact that the eve of May Day and May Day were at the end of the
control period. In Finland, alcohol consumption belongs to the
celebration of May Day.
It has been assumed that sitostanol ester could reduce serum total and
LDL cholesterol concentration more effectively in subjects with the apo
E allele 4 than those with allele 2 or 3 (Miettinen and Vanhanen 1994
, Vanhanen et al. 1993
), but our results do
not support this assumption. In the present study there were no
significant differences in percentage reduction in LDL cholesterol
between subjects with apo genotype 3:3 and 3:4 during the different
dose periods. However, when interpreting this result, it should be kept
in mind that the sizes of two apo E genotype groups were rather small.
During the study there were no significant changes in the
concentrations of serum retinol, - and ß-carotene, and in the
concentration of serum tocopherols related to the serum total
cholesterol concentrations. Serum lycopene concentration did not change
significantly in men during the study, whereas in women there were
significant differences among the different dose periods even after
standardization for serum total cholesterol concentration. However, the
differences were not related to the dose of stanol ester. Women had
lower concentrations of serum lycopene than men, which could be due
partly to their older age (Vogel et al. 1997
) (mean age
52 vs. 42 y, women vs. men). Also the changes in serum
25-hydroxycholecalsiferol were not related to the dose of stanol ester.
Based on previous plant sterol studies (Gylling et al. 1996
, Hallikainen and Uusitupa 1999
,
Hendriks et al. 1999
, Weststrate and Meijer 1998
), it seems that plant sterol would have some effect on
serum carotenoid concentrations. However, the results are variable:
Before lipid standardization serum carotenoid concentrations might have
decreased significantly, but after lipid standardization these changes
have not usually been great or significant (Gylling et al. 1996
, Hallikainen and Uusitupa 1993,
Hendriks et al. 1999
, Weststrate and Meijer 1998
). These differences in the results can not be due
to different carotenoid contents in test margarines, because both test
and control margarines have been similarly vitaminized in those studies
(Gylling et al. 1996
, Hallikainen and Uusitupa 1999
, Hendriks et al. 1999
, Weststrate and Meijer, 1998
). Therefore, the differences in the results
might be a consequence of variability in composition of background
diets. The diets in the studies of Gylling et al. (1996)
, Hendriks et al. (1999)
and Weststrate and Meijer (1998)
were not standardized like
in our earlier (Hallikainen and Uusitupa 1999
) or
present study. In our studies the instructions for intake of vegetables
were given to the subjects. According to our findings, the
effects of plant sterol on serum carotenoid concentrations were minor
and clinically unimportant. However, additional studies will be needed
to discover long-term effects of plant sterol on carotenoid
concentrations.
In conclusion, significant reduction of serum total and LDL cholesterol concentrations is reached with the dose of 1.6 g stanol, and increasing the dose of stanol from 2.4 g to 3.2 g does not provide clinically significant additional benefits. Interestingly, the 0.8-g dose of stanol resulted in 8.7% reduction in apo B concentration. Serum plant stanol concentrations rose slightly with the dose; however, their concentrations remained extremely low in serum.
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
3 Abbreviations used: apo , apolipoprotein; E%, energy percentage.
Manuscript received June 29, 1999. Initial review completed September 21, 1999. Revision accepted November 29, 1999.
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMETHODSRESULTSDISCUSSIONREFERENCES |
|---|
1. Alpers D. H., Clouse R. E., Stenson W. F. Protein and Calorie Requirements. Manual of Nutritional Therapeutics 1986:131-159 Little, Brown and Company Toronto.
2. Becker M., Staab D., von Bergmann K. Treatment of severe familial hypercholesterolemia in childhood with sitosterol and sitostanol. J. Pediatr. 1993;122:292-296[Medline]
3. Cullinane E. M., Yurgalevitch S. M., Saritelli A. L., Herbert P. N., Thompson P. D. Variations in plasma volume affect total and low-density lipoprotein cholesterol concentrations during the menstrual cycle. Metabolism 1995;44:965-971[Medline]
4.
Driskell W. J., Bashor M. M., Neese J. W. Beta-carotene determined in serum by liquid chromatography with an internal standard. Clin. Chem. 1983;29:1042-1044
5. Farquhar J. W., Smith R. E., Dempsey M. E. The effect of beta sitosterol on the serum lipids of young men with arteriosclerotic heart disease. Circulation 1956;XIV:77-82
6. Gylling H., Miettinen T. A. Serum cholesterol and cholesterol and lipoprotein metabolism in hypercholesterolaemic NIDDM patients before and during sitostanol ester-margarine treatment. Diabetologia 1994;37:773-780[Medline]
7.
Gylling H., Puska P., Vartiainen E., Miettinen T. A. Serum retinol, -tocopherol, carotenes and lipid peroxide production during serum cholesterol lowering by sitostanol ester margarine in a mildly hypercholesterolemic population. Circulation 1996;94(Suppl. 1):578(abs.)
8.
Gylling H., Puska P., Vartiainen E., Miettinen T. A. Serum sterols during stanol ester feeding in a mildly hypercholesterolemic in a mildly hypercholesterolemic population. J. Lipid Res. 1999;40:593-600
9.
Gylling H., Radhakrishnan R., Miettinen T. A. Reduction of serum cholesterol in postmenopausal women with previous myocardial infarction and cholesterol malabsorption induced by dietary sitostanol ester margarine. Women and dietary sitostanol. Circulation 1997;96:4226-4231
10. Gylling H., Siimes M., Miettinen T. A. Sitostanol ester margarine in dietary treatment of children with familial hypercholesterolemia. J. Lipid Res. 1995;36:1807-1812[Abstract]
11. Haapa E., Toponen T., Pietinen P., Räsänen L. Annoskuvakirja 1985 Sosial Insurance Institution Helsinki, Finland (in Finnish) Portion size booklet
12.
Hallikainen M. A., Uusitupa M. I. J. Effects of two low-fat stanol ester-containing margarines on serum cholesterol concentrations as part of a low-fat diet in hypercholesterolemic subjects. Am. J. Clin. Nutr. 1999;69:403-410
13. Hassan A. S., Rampone A. J. Intestinal absorption and lymphatic transport of cholesterol and ß-sitostanol in rats. J. Lipid Res. 1979;20:646-653[Abstract]
14. Heinemann T., Axtmann G., von Bergmann K. Comparison of intestinal absorption of cholesterol with different plant sterols in man. Eur. J. Clin. Invest. 1993;23:827-831[Medline]
15. Heinemann T., Leiss O., von Bergmann K. Effect of low-dose sitostanol on serum cholesterol in patients with hypercholesterolemia. Atherosclerosis 1986;61:219-223[Medline]
16. Hendriks H. F. J., Weststrate J. A., van Vlient T., Meijer G. W. Spreads enriched with three different levels of vegetable oil sterols and the degree of cholesterol lowering in normocholesterolaemic and mildly hypercholesterolaemic subjects. Eur. J. Clin. Nutr. 1999;53:319-327[Medline]
17. Ikeda I., Sugano M. Inhibition of cholesterol absorption by plant sterols for mass intervention. Curr. Opin. Lipidol. 1998;9:527-531[Medline]
18. Jones P. J. H., MacDougall D. E., Ntanios F., Vanstone C. A. Dietary phytosterols as cholesterol-lowering agents in humans. Can. J. Physiol. Pharmacol. 1997;75:217-227[Medline]
19. Kaplan L. A., Miller J. A., Stein E. A. Simultaneous measurement of serum retinol, tocopherols, carotenes, and carotenoids by high performance liquid chromatography. J. Clin. Lab. Anal. 1987;1:147-152
20. Lütjohann D., Meese C. O., Crouse J. R., von Bergmann K. Evaluation of deuterated cholesterol and deuterated sitostanol for measurement of cholesterol absorption in human. J. Lipid Res. 1993;34:1039-1046[Abstract]
21. Miettinen T. A. Cholesterol metabolism during ketoconazole treatment in man. J. Lipid Res. 1988;29:43-51[Abstract]
22. Miettinen T. A., Gylling H. Regulation of cholesterol metabolism by dietary plant sterols. Curr. Opin. Lipidol. 1999;10:9-14[Medline]
23. Miettinen T. A., Koivisto P. Non-cholesterol sterols and bile acid production in hypercholesterolaemic patients with ileal bypass. Paumgartner G. Stiehl A. Gerok W. eds. Bile Acid and Cholesterol in Health and Disease 1983:183-187 MTP Press Lancaster.
24.
Miettinen T. A., Puska P., Gylling H., Vanhanen H., Vartiainen E. Reduction of serum cholesterol with sitostanol-ester margarine in a mildly hypercholesterolemic population. N. Engl. J. Med. 1995;333:1308-1312
25.
Miettinen T. A., Tilvis R. S., Kesäniemi Y. A. Serum plant sterols and cholesterol precursors reflect cholesterol absorption and synthesis in volunteers of a randomly selected male population. Am. J. Epidemiol. 1990;131:20-31
26. Miettinen T. A., Vanhanen H. T. Dietary sitostanol related to absorption, synthesis and serum level of cholesterol in different apolipoprotein E phenotypes. Atherosclerosis 1994;105:217-226[Medline]
27. Niinikoski H., Viikari J., Palmu T. Cholesterol-lowering effect and sensory properties of sitostanol ester margarine in normocholesterolemic adults. Scand. J. Nutr. 1997;41:9-12
28. Norusis M. J. SPSS for Windows Base System User’s Guide. Release 6 0 1993 SPSS Chicago, IL.
29. Parviainen M. T. The fat-soluble vitamins A, D and E - their metabolism, binding-proteins and determination from human serum. Acta Universitatis Tamperensis 1983;159:49-54
30. Penttilä I. M., Voutilainen E., Laitinen O., Juutilainen P. Comparison of different analytical and precipitation methods for the direct estimation of high-density lipoprotein cholesterol. Scand. J. Clin. Lab. Invest. 1981;41:353-360[Medline]
31. Pollak O. J., Kritchevsky D. Phytosterols-sitosterol as cholesterol depressants. Sitosterol. Monographs on Atherosclerosis 1981;Vol. 10:60-118 Karger Basel, Switzerland.
32. Rastas M., Seppänen R., Knuts L.-R., Karvetti R.-L., Varo P. Nutrient Composition of Foods 1993 Social Insurance Institution Helsinki, Finland.
33. Steinberg D., Pearson T. A., Kuller L. H. Alcohol and atherosclerosis. Ann. Intern. Med. 1991;114:967-976
34.
Tilvis R. S., Miettinen T. A Serum plant sterols and their relation to cholesterol absorption. Am. J. Clin. Nutr. 1986;43:92-97
35. Tsukamoto K., Watanabe T., Matsushima T., Kinoshita M., Kato H., Hashimoto Y., Kurokawa K., Teramoto T. Determination by PCR-RFLP of apo E genotype in a Japanese population. J. Lab. Clin. Med. 1993;121:598-602[Medline]
36. Vanhanen H. T., Blomqvist S., Ehnholm C., Hyvönen M., Jauhiainen M., Torstila I., Miettinen T. A. Serum cholesterol, cholesterol precursors, and plant sterols in hypercholesterolemic subjects with different apo E phenotypes during dietary sitostanol ester treatment. J. Lipid Res. 1993;34:1535-1544[Abstract]
37. Vanhanen H. T., Kajander J., Lehtovirta H., Miettinen T. A. Serum levels, absorption efficiency, faecal elimination and synthesis of cholesterol during increasing doses of dietary sitostanol esters in hypercholesterolaemic subjects. Clin. Sci. 1994;87:61-67[Medline]
38.
Vogel S., Contois J. H., Tucker K. L., Wilson P. W., Schaefer E. J., Lammi-Keefe C. J. Plasma retinol and plasma and lipoprotein tocopherol and carotenoid concentrations in healthy elderly participants of the Framingham Heart Study. Am. J. Clin. Nutr. 1997;66:950-958
39. Weststrate J. A., Meijer G. W. Plant sterol-enriched margarines and reduction of plasma total- and LDL-cholesterol concentrations in normocholesterolaemic and mildly hypercholesterolaemic subjects. Eur. J. Clin. Nutr. 1998;52:334-343[Medline]
40.
Xu G., Salen G., Tint G. S., Batta A. K., Shefer S. Campestanol (24-methyl-5-cholestan-3ß-ol) absorption and distribution in New Zealand white rabbits: effects of dietary sitostanol. Metabolism 1999;48:363-368[Medline]
This article has been cited by other articles:
|
|
 |
|
  O. J. G. Schiepers, R. H. M. de Groot, M. P. J. van Boxtel, J. Jolles, A. de Jong, D. Lutjohann, J. Plat, and R. P. Mensink Consuming Functional Foods Enriched with Plant Sterol or Stanol Esters for 85 Weeks Does Not Affect Neurocognitive Functioning or Mood in Statin-Treated Hypercholesterolemic Individuals J. Nutr., July 1, 2009; 139(7): 1368 - 1373. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 V. S. Retelny, A. Neuendorf, and J. L. Roth Nutrition Protocols for the Prevention of Cardiovascular Disease Nutr Clin Pract, October 1, 2008; 23(5): 468 - 476. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I. Rudkowska Functional foods for cardiovascular disease in women Menopause Int, June 1, 2008; 14(2): 63 - 69. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Naumann, J. Plat, A. D.M. Kester, and R. P. Mensink The Baseline Serum Lipoprotein Profile Is Related to Plant Stanol Induced Changes in Serum Lipoprotein Cholesterol and Triacylglycerol Concentrations J. Am. Coll. Nutr., February 1, 2008; 27(1): 117 - 126. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Theuwissen and R. P. Mensink Simultaneous Intake of {beta}-Glucan and Plant Stanol Esters Affects Lipid Metabolism in Slightly Hypercholesterolemic Subjects J. Nutr., March 1, 2007; 137(3): 583 - 588. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. K Nies, A. A Cymbala, S. L Kasten, D. G Lamprecht, and K. L Olson Complementary and Alternative Therapies for the Management of Dyslipidemia Ann. Pharmacother., November 1, 2006; 40(11): 1984 - 1992. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. S. AbuMweis, C. A. Vanstone, N. Ebine, A. Kassis, L. M. Ausman, P. J. H. Jones, and A. H. Lichtenstein Intake of a Single Morning Dose of Standard and Novel Plant Sterol Preparations for 4 Weeks Does Not Dramatically Affect Plasma Lipid Concentrations in Humans J. Nutr., April 1, 2006; 136(4): 1012 - 1016. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I. A Castro, L. P Barroso, and P. Sinnecker Functional foods for coronary heart disease risk reduction: a meta-analysis using a multivariate approach Am. J. Clinical Nutrition, July 1, 2005; 82(1): 32 - 40. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. W. Lau, M. Journoud, and P. J. Jones Plant sterols are efficacious in lowering plasma LDL and non-HDL cholesterol in hypercholesterolemic type 2 diabetic and nondiabetic persons Am. J. Clinical Nutrition, June 1, 2005; 81(6): 1351 - 1358. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. C. Hayes, A. Pronczuk, and D. Perlman Nonesterified Phytosterols Dissolved and Recrystallized in Oil Reduce Plasma Cholesterol in Gerbils and Humans J. Nutr., June 1, 2004; 134(6): 1395 - 1399. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. H. Ju, L. M. Clausen, K. F. Allred, A. L. Almada, and W. G. Helferich {beta}-Sitosterol, {beta}-Sitosterol Glucoside, and a Mixture of {beta}-Sitosterol and {beta}-Sitosterol Glucoside Modulate the Growth of Estrogen-Responsive Breast Cancer Cells In Vitro and in Ovariectomized Athymic Mice J. Nutr., May 1, 2004; 134(5): 1145 - 1151. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. J. H. Jones, C. A. Vanstone, M. Raeini-Sarjaz, and M.-P. St-Onge Phytosterols in low- and nonfat beverages as part of a controlled diet fail to lower plasma lipid levels J. Lipid Res., September 1, 2003; 44(9): 1713 - 1719. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. B. Katan, S. M. Grundy, P. Jones, M. Law, T. Miettinen, R. Paoletti, and Stresa Workshop Participants Efficacy and Safety of Plant Stanols and Sterols in the Management of Blood Cholesterol Levels Mayo Clin. Proc., August 1, 2003; 78(8): 965 - 978. [Abstract] [PDF]
|
|
|
|
 |
|
 M. Evans, A. Roberts, and A. Rees Pharmacological management of hyperlipidaemia The British Journal of Diabetes & Vascular Disease, May 1, 2003; 3(3): 204 - 210. [Abstract] [PDF]
|
|
|
|
|
|
L. F Masson, G. McNeill, and A. Avenell Genetic variation and the lipid response to dietary intervention: a systematic review Am. J. Clinical Nutrition, May 1, 2003; 77(5): 1098 - 1111. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. S. Ewart, L. K. Cole, J. Kralovec, H. Layton, J. M. Curtis, J. L. C. Wright, and M. G. Murphy Fish Oil Containing Phytosterol Esters Alters Blood Lipid Profiles and Left Ventricle Generation of Thromboxane A2 in Adult Guinea Pigs J. Nutr., June 1, 2002; 132(6): 1149 - 1152. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. H. Lichtenstein and R. J. Deckelbaum Stanol/Sterol Ester-Containing Foods and Blood Cholesterol Levels : A Statement for Healthcare Professionals From the Nutrition Committee of the Council on Nutrition, Physical Activity, and Metabolism of the American Heart Association Circulation, February 27, 2001; 103(8): 1177 - 1179. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
