A low intake of antioxidant nutrients is associated with poor semen quality in patients attending fertility clinics.

Abstract

To compare specific nutrient intake between normospermic and oligoasthenoteratospermic patients attending infertility clinics in two Mediterranean provinces of Spain.
Case-control study.
Private fertility clinics in southeastern Spain.
Thirty men with poor semen quality (case subjects) and 31 normospermic control subjects of couples attending our fertility clinics.
We recorded dietary habits and nutrient consumption using a food frequency questionnaire adapted to meet specific study objectives.
We calculated nutrient intakes by multiplying the frequency of use for each food by the nutrient composition of the portion size specified on the food frequency questionnaire and by addition across all foods to obtain a total nutrient intake for each individual. Semen quality was assessed by measuring volume, concentration, motility, and morphology. Hormones levels were also analyzed in case and control subjects.
In the logistic regression, control subjects had a significantly higher intake of carbohydrates, fiber, folate, vitamin C, and lycopene and lower intakes of proteins and total fat.
A low intake of antioxidant nutrients was associated with a poor semen quality in this case-control study of Spanish men attending infertility clinics.

Full text

Page 1

A low intake of antioxidant nutrients is associated with
poor semen quality in patients attending fertility
clinics
Jaime Mendiola, Ph.D.,a,bAlberto M. Torres-Cantero, M.D., Dr.P.H.,b,cJes? us Vioque, M.D., Ph.D.,c,d
Jos? e M. Moreno-Grau, Ph.D.,eJorge Ten, Ph.D.,aManuela Roca, M.D., Ph.D.,b
Stella Moreno-Grau, Ph.D.,eand Rafael Bernabeu, M.D., Ph.D.a,f
aDepartment of Reproductive Biology and Medicine, Instituto Bernabeu, Alicante;bDepartment of Preventive Medicine and
Public Health, School of Medicine, University of Murcia, Espinardo;cEl Centro de Investigaci? on Biom? edica en Red de
Epidemiolog? ıa y Salud P? ublica (CIBERESP), Elche;dDepartment of Public Health, Miguel Hernandez University, Elche;
eDepartment of Environmental and Chemical Engineering, Technical University of Cartagena, Cartagena; andfReproductive
Medicine Chair, Miguel Hernandez University–Instituto Bernabeu, Alicante, Spain
Objective: To compare specific nutrient intake between normospermic and oligoasthenoteratospermic patients at-
tending infertility clinics in two Mediterranean provinces of Spain.
Design: Case-control study.
Setting: Private fertility clinics in southeastern Spain.
Patient(s): Thirty men with poor semen quality (case subjects) and 31 normospermic control subjects of couples
attending our fertility clinics.
Intervention(s): We recorded dietary habits and nutrient consumption using a food frequency questionnaire adap-
ted to meet specific study objectives.
Main Outcome Measure(s): We calculated nutrient intakes by multiplying the frequency of use for each food by
thenutrientcompositionoftheportionsizespecifiedonthefoodfrequencyquestionnaireandbyadditionacrossall
foods to obtain a total nutrient intake for each individual. Semen quality was assessed by measuring volume, con-
centration, motility, and morphology. Hormones levels were also analyzed in case and control subjects.
Result(s): In the logistic regression, control subjects had a significantly higher intake of carbohydrates, fiber,
folate, vitamin C, and lycopene and lower intakes of proteins and total fat.
Conclusion(s): A low intake of antioxidant nutrients was associated with a poor semen quality in this case-control
study of Spanish men attending infertility clinics. (Fertil Steril?2009;-:-–-. ?2009 by American Society for
Reproductive Medicine.)
Key Words: Food frequency, male infertility, antioxidants, vitamins
Several studies have suggested that human semen quality and
fertility have been declining during the last decades (1–6).
Deterioration in seminal samples has been related to environ-
mental and occupational pollutants, changes in lifestyles,
exposure to toxics, and dietary habits (7–9).
Concerning dietary factors, a lower intake of some anti-
oxidant nutrients, such as vitamins A, C, and E, carnitines,
folate, zinc, and selenium, has been associated with male
infertility (10–14). In a cross-sectional study of 97 healthy
male volunteers, a higher nutrient intake of vitamins C, E
and b-carotene was associated with a higher sperm count
number and motility (15). Lewis et al. (16) found that
men with the lowest intake of dietary antioxidants had
less sperm motility; the consumption of <5 servings/day
of fruits and vegetables as well as the vitamin C intake
were significantly lower among infertile men than among
control subjects.
In a prior analysis comparing intakes of food items in the
same study population, we found that control subjects had
a higher intake of skim milk, shellfish, tomatoes, and lettuce,
and case subjects consumed more yogurt, meat products, and
potatoes. In the logistic regression model, case subjects had
lower intake of lettuce, tomatoes, and fruits (apricots and
peaches) and significantly higher intake of dairy and meat
products (17). However, in that study we could not attribute
those differences to specific nutrients.
The aim of the present study was to compare specific
nutrient intake between normospermic and oligoasthenotera-
tospermic patients attending infertility clinics in two Medi-
terranean provinces of Spain.
Received August 26, 2008; revised October 6, 2008; accepted October
27, 2008.
J.M. has nothing to disclose. A.T.-C. has nothing to disclose. J.V. has
nothing to disclose. J.M.-G. has nothing to disclose. J.T. has nothing
to disclose. M.R. has nothing to disclose. S.M.-G. has nothing to dis-
close. R.B. has nothing to disclose.
Supported inpartbyTheSenecaFoundation,RegionalAgencyofScience
and Technology, Department of Education and Culture of Murcia
(Ref.00694/PI/04),andtheReproductiveMedicineChairofMiguelHer-
nandez University–Instituto Bernabeu and El Centro de Investigaci? on
Biom? edica en Red de Epidemiolog? ıa y Salud P? ublica (CIBERESP).
Reprint requests:Dr.Jaime Mendiola. Division ofPreventive Medicineand
Public Health, University of Murcia, 30100, Espinardo (Murcia), Spain
(FAX: þ34968363947; E-mail: mendiola.j@gmail.com).
0015-0282/09/$36.00
doi:10.1016/j.fertnstert.2008.10.075
Fertility and Sterility?Vol. -, No. -, - 2009
1
Copyright ª2009 American Society for Reproductive Medicine, Published by Elsevier Inc.
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MATERIALS AND METHODS
Design and patients
Themenofcouplesattendingthethreeinfertilitycentersofthe
Instituto Bernabeu in Murcia and Alicante (southeastern
Spain)between2005and2007wereclassifiedintotwogroups
on the basis of seminal quality following World Health Orga-
nization(WHO)criteria(18):1)casesubjects(n¼30;17from
Alicanteand13fromMurcia)composedofmenwithsevereor
moderateoligozoospermia(<5millionspermatazoa(spz)/mL
or 5–20 million spz/mL, respectfully) and severe teratozoo-
spermia[<6%normalformsstrictcriteria(19)];and2)control
subjects (n ¼ 31, 18 from Alicante and 13 from Murcia) com-
posedofnormospermicpatients(R20millionspz/mL,R50%
motile sperm, and R14% normal forms strict criteria).
Another ten patients and twelve controls that were invited to
participaterefusedtobeincludedinthestudy;therefore,there
were no significant differences between the refusal rates in
caseandcontrolsubjects.Subjectsprovidedatleasttwosemen
samples. Abstinence time ranged from 3 to 5 days without
significant differences between case and control subjects.
Following WHO recommendations, the interval between the
two collections was between 7 days and 3 weeks (18).
Semen analyses were performed by the same technician,
which was blind for other clinical data. Semen parameters
evaluated included: ejaculate volume, sperm concentration,
percentage of motile sperm (grades a and b according to
WHO criteria), and percentage of normal forms following
Kruger’s strict criteria (19). To assess motility, a 5-mL aliquot
was added to a Makler chamber (Sefi-Medical Instruments,
Haifa,Israel)andspermwasvisualizedbyphase-contrastmi-
croscope (Olympus CX21; Olympus, Tokyo, Japan) at ?200
magnification. A 20-mL semen aliquot smear was dried at
room temperature and stained with Papanicolau stain (20).
Methods used in this study have been previously described
(17). This study was approved by the Institutional Review
Board of our clinics, and patients were included in the study
after giving informed written consent.
Data collection
Study participants were interviewed by the same trained
fieldworker, who was blinded regarding case-control patient
status, using structured questionnaires to collect information
for sociodemographic characteristics, tobacco use, and die-
tary assessment. At the clinical examination, the fieldworker
followed the same standard protocol in all centers to obtain
anthropometric measurements. Height was measured to the
nearest 0.1 cm with subjects standing without shoes and
with their backs to the stadiometer. Body weight was mea-
sured in light clothes to the nearest 0.1 kg on a digital scale
which was placed on a firm flat surface. The body mass index
(BMI) was calculated from weight and height measurements
as kg/m2. We also collected information on occupation his-
tory using a questionnaire adapted from the one developed
by the U.S. Agency for Toxic Substances and Disease Regis-
try in cooperation with the National Institute for Occupa-
tional Safety and Health (21). Past exposure to chemicals
was assessed from the occupational history as reported by
case and control subjects. It was considered that the person
had a previous occupational toxic exposure if he answered
that he had been exposed to any of the chemicals that were
listed. Detailed information of this study methodology may
be found elsewhere (22).
Dietary assessment
A semiquantitative food frequency questionnaire (FFQ) of
93 food items was used to assess usual dietary intake. The
FFQ was a modified version of the Harvard questionnaire
(23) that was previously validated and adapted for adult
Spanish populations (24, 25). When comparing the nutrient
intakesfromtheFFQwiththosefromfour1-weekdietaryre-
cords, the average correlation coefficients for 1-year validity
and reproducibility of nutrient intakes were 0.47 and 0.40,
respectively, a similar range to other established diet ques-
tionnaires (26). For a similar version of this FFQ adapted
for an elderly population in the same area of Spain, we
observed satisfactory correlation coefficients between
plasma concentration and the nutrient intake of carotenoids
and vitamin C: 0.20 and 0.36, respectively (27).
Participants in the present study were asked how often, on
average, they had consumed each type item during the previ-
ous year. Standard units or serving sizes were specified for
each food item in the FFQ. The questionnaire had nine pos-
sible responses, ranging from ‘‘never or less than once per
month’’ to ‘‘six or more times per day.’’ The response to
each food item was converted to average daily intake for
each individual participant. The average daily intakes for
each fruit and vegetable were summed to compute the total
fruit and vegetable intake. Nutrient values were primarily ob-
tained from food composition tables from the United States
Department of Agriculture and other published sources
(28). We calculated nutrient intakes by multiplying the fre-
quency of use for each food by the nutrient composition of
the portion size specified on the FFQ and by addition across
all foods to obtain a total nutrient intake for each individual.
None of the patients had dietary or vitamin supplements dur-
ing the total duration of study. Alcohol consumption was as-
sessed within an FFQ including specific items on wine, beer,
and liquor and specifying standard portions for all of them.
We calculated ethanol intake by multiplying the frequency
of consumption of each beverage by the alcohol content of
the specified portion size and summing across beverages.
Analysis
Statistical analysis was performed with the statistical pack-
age SPSS 15.0 (SPSS, Chicago, IL). Results are presented
as percentage and mean with standard deviation (SD). De-
scriptive means are presented using untransformed data.
We used unpaired Student t tests for means comparison of
continuous variables for case and control subjects. c2tests
were used for categoric variables. The level of statistical
significance was set at .05, and all tests were two-tailed.
Statistical tests for nutrient intakes were made with calorie-
adjusted values by calculating the residuals from a linear
2
Mendiola et al.
Antioxidant nutrients and semen quality
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regression with the log e of the nutrient modeled as the
dependent variable and the log e of total energy intake as
the independent variable (26).
Adjusted odds ratios (ORs) and 95% confidence intervals
(CIs) were calculated using unconditional logistic regression.
Toexploretheassociationbetweensemenquality(normosper-
mia yes/no) and nutrients intake we used the lowest tertile of
every nutrient intake adjusted for total energy as the reference
category. Tertiles of nutrient intakes were estimated based on
thewholedistributionofcaseandcontrolsubjects.Thefollow-
ing potential confounders were included in the models: age
(years), total energy intake (kcal), tobacco smoking (no/yes),
and previous occupational toxic exposures (no/yes). Tests for
trend inthe ORs acrossexposure stratawerecalculatedfor nu-
trient intakes by using logistic models that included categoric
terms as continuous variables in a model with all of the poten-
tial confounders. For trend tests, we used the likelihood ratio
test statistic with one degree of freedom.
RESULTS
Table1showsthedistributionofcasesandcontrolsaccording
to personal characteristics, hormone levels, and semen
parameters.Thedistributionofage,BMI,andhormonelevels
were similar between case and control subjects. Control
subjects had higher absolute intakes of carbohydrates, fiber,
folate, vitamin C, and lycopene (P<.05).
Table 2 shows risk estimates according to nutrient intakes
by tertiles using logistic regression models. Control subjects
presented significantly higher intakes of carbohydrates, fiber,
folate, vitamin C, and lycopene (P<.05) and lower intakes of
proteins and total fat (P<.05). For these nutrients, statisti-
cally significant trends were observed (P<.05).
Significant differences also existed between case and
control subjects in occupational exposure. A multiple
logistic regression model was used incorporating the nutri-
ents that had significantly different intakes between case
TABLE 1
Personal characteristics, semen quality parameters, and nutrient intakes among case and control
subjects.
VariablesCase subjects (n [ 30)Control subjects (n [ 31)
Age (yrs)
Body mass index (kg/m2)
Current smoker
Current alcohol drinking
Hormonal levels
FSH (mUI/mL)
LH (mUI/mL)
T (ng/mL)
Semen characteristicsa
Volume (mL)
Concentration (million/mL)
Sperm motility (grades a þ b, %)
Normal morphologyb(%)
Nutrient intakes
Protein (g)
Carbohydrates (g)
Total fat (g)
Fiber (g)
Folate (mg)
Vitamin C (mg)
Vitamin E (mg)
Lycopene (mg)
b-Carotene (mg)
Selenium (mg)
Kilocalorie intake/day
34.2 ? 3.7
23.2 ? 1.0
8 (26.7%)
14 (46.7)
32.8 ? 3.9
23.5 ? 1.1
11 (35.5%)
18 (58.1%)
6.4 ? 2.1
4.2 ? 1.2
5.3 ? 1.6
6.5 ? 1.4
4.1 ? 1.4
5.4 ? 1.3
3.8 ? 1.2
3.5 ? 2.1
27.4 ? 18.6
3.7 ? 1.5
3.5 ? 1.4
39.5 ? 14.6
52.2 ? 12.3
22.3 ? 4.5
91.8 ? 33.9
132.2 ? 46.6
94.1 ? 23.3
11.8 ? 4.4
217.6 ? 76.6
45.5 ? 20.9
9.8 ? 2.1
2,890.5 ? 1,529.9
3,647.3 ? 2,311.1
108.6 ? 41.0
1,767.7 ? 484.1
94.4 ? 33.3
156.7 ? 44.1c
99.0 ? 27.9
14.2 ? 3.9c
269.5 ? 74.8c
57.9 ? 21.8c
10.9 ? 2.3
4,238.5 ? 2,044.9c
4,351.4 ? 2,008.8
115.4 ? 39.0
1,921.5 ? 530.7
aMean of the two samples.
bNormal morphology defined according to the strict criteria by Kruger et al. (19).
cControl subjects presented significantly higher intakes of the following nutrients: carbohydrates, fiber, folate, vitamin C,
and lycopene (P<.05).
Mendiola. Antioxidant nutrients and semen quality. Fertil Steril 2009.
Fertility and Sterility?
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and control subjects in the univariate analyses and occupa-
tional exposure. In the logistic regression, control subjects
had a significantly higher intake of carbohydrates, fiber, fo-
late, vitamin C, and lycopene (P<.05) and lower intakes of
proteins and total fat, and had been exposed less frequently
to occupational toxics than case subjects (P<.05).
DISCUSSION
Ourstudysuggeststhatpoorsemenqualitymaybeassociated
with a lower intake of carbohydrates, fiber, folate, vitamin C,
and lycopene and a higher intake of protein and total fat.
Overall, a low intake of antioxidant nutrients might seem to
have a negative effect on semen quality.
TABLE 2
Adjusted odds ratios (ORs) for poor/low semen quality according to usual nutrient intake.
Nutrient
intake
in tertiles
Case
subjects
Control
subjects
ORa
(95% CI)
P
value
ORb
(95% CI)
P
value
ORc
(95% CI)
P
value
Test for
trend, P
Protein
Low
Medium
High
Carbohydrates
Low
Medium
High
Total Fat
Low
Medium
High
Fiber
Low
Medium
High
Folate
Low
Medium
High
Vitamin C
Low
Medium
High
Vitamin E
Low
Medium
High
Lycopene
Low
Medium
High
b-Carotene
Low
Medium
High
Selenium
Low
Medium
High
8
6
12
14
5
1
0.66 (0.17–2.54)
4.92 (1.23–19.62)
1
0.83 (0.20–3.45)
5.25 (1.29–21.52)
1
.55
.02
.80
.02
1.27 (0.25–6.41)
8.27 (1.58–43.34)
.77
.01 16.011
14
8
8
61
0.32 (0.08–1.22)
0.26 (0.07–0.99)
1
0.36 (0.09–1.44)
0.21 (0.05–0.85)
1
12
13
.10
.05
.15
.03
0.32 (0.06–1.65)
0.09 (0.02–0.59)
.17
.01.001
7
9
13
11
7
1
1.67 (0.44–6.28)
3.46 (0.93–12.85)
1
2.61 (0.61–11.22)
5.04 (1.21–20.95)
1
.45
.06
.20
.03 11.81 (1.87–74.65) <.01
4.91 (0.86–28.17).07
14.009
13
11
6
7
9
1
0.68 (0.18–2.63)
0.19 (0.05–0.74)
1
0.84 (0.20–3.56)
0.14 (0.03–0.64)
1
.58
.02
.82
.01
0.87 (0.17–4.43)
0.13 (0.02–0.68)
.87
.0115.018
13
10
7
71
0.53 (0.14–1.97)
0.23 (0.06–0.88)
1
0.60 (0.15–2.43)
0.16 (0.04–0.73)
1
10
14
.34
.03
.48
.02
0.44 (0.09–2.18)
0.13 (0.02–0.71)
.31
.02.024
14
10
6
61
0.37 (0.09–1.47)
0.14 (0.03–0.57) <.01
1
0.35 (0.08–1.50)
0.11 (0.02–0.52)
1
10
15
.16.160.27 (0.05–1.40)
0.09 (0.02–0.51)
.12
<.01
<.01.008
13
8
9
51
0.20 (0.05–0.80)
0.32 (0.08–1.26)
1
0.16 (0.04–0.71)
0.36 (0.09–1.55)
1
14
12
.02
.10
.02
.17
0.22 (0.04–1.11)
0.52 (0.11–2.49)
.07
.41
12
11
7
8
9
1
0.77 (0.20–3.05)
0.29 (0.08–1.09)
1
0.68 (0.16–2.81)
0.25 (0.06–1.00)
1
.72
.07
.59
.05
0.66 (0.13–3.37)
0.16 (0.03–0.82)
.62
.0314 .034
13
6
11
71
0.22 (0.05–0.92)
0.44 (0.12–1.61)
1
0.26 (0.06–1.12)
0.41 (0.11–1.60)
1
13
11
.04
.21
.07
.20
0.20 (0.04–1.10)
0.53 (0.12–2.34)
.06
.40
11
6
13
91
0.30 (0.07–1.18)
1.48 (0.40–5.52)
1
0.27 (0.07–1.17)
1.52 (0.40–5.90)
1
14
8
.08
.56
.08
.54
0.45 (0.10–2.11)
2.04 (0.45–9.21)
.31
.36
aOR adjusted for age and total energy intake.
bOR adjusted for age, total energy intake, BMI, and smoking.
cOR adjusted for age, total energy intake, BMI, smoking, and previous occupational toxic exposures.
Mendiola. Antioxidant nutrients and semen quality. Fertil Steril 2009.
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A higher antioxidant dietary intake has been associated
with higher sperm numbers and motility in healthy nonsmok-
ing men (15). A recent review of intervention studies about
the effect of antioxidants on semen parameters concluded
that vitamins A, C, E, carnitine, and glutathione improve
semen quality in male factor infertility (14). Oral supplemen-
tation with vitamin C or E and folic acid plus zinc sulfate has
also been shown to improve sperm counts, motility, and
morphology in infertile patients (11, 29, 30).
In the present study, we have found a positive association
between semen quality and vitamin C intake. The association
between vitamin E and b-carotene was not significant once
we adjusted by previous occupational toxic exposures. We
did not find an association between selenium intake and
semen quality,which isinagreementwith recent intervention
studies in which selenium supplements did not improve
semen quality (31, 32). We also found a positive association
between folate intake and semen quality, which hasbeen sup-
ported in intervention studies that have shown that supple-
mentation with folic acid significantly increased sperm
quality in subfertile males (11, 33).
Moreover, in a recent article, Young et al. (34) found in
a healthy population of nonsmoking men that men with
high folate intake (>75th percentile) had lower frequencies
of sperm with X, 21, sex nullisomy, and a lower aggregate
measure of sperm aneuploidy (P%.04) compared with men
with lower intakes. Finally, we found that lycopene intake
was positively associated with semen quality. Lycopene is
a powerful carotenoid antioxidant, and therefore, the effect
we found is consistent with the hypothesis discussed ear-
lier. We have found only an experimental study in which
the semen quality of 30 men with idiopathic nonobstruc-
tive oligo/astheno/teratozoospermia improved after receiv-
ing 2,000 mg lycopene (35). However that study was not
randomized.
As in other case-control studies, the present study may
present limitations common to observational studies. Be-
sides,somerealassociationscouldhaveremained undetected
owingtosamplesizelimitations.However,thestrengthofthe
associations, the existence of a dose response, the control for
other potential confounders such as age, tobacco smoking,
andoccupationalfactors,andtheconsistencywithotherstud-
ies in humans and other animals support that the study results
are real, and that nutrient intake, particularly low intake of
antioxidant nutrients, is likely to be causally related to poor
semen quality (15, 16, 36). Our findings might be compatible
with the hypothesis that oxidative stress associated with in-
creased reactive oxygen species generation and reduced anti-
oxidant capacity is negatively correlated with sperm
concentration, motility, and morphology in infertile men
(37–40).
In conclusion, the present case-control study suggests that
the risk of poor semen quality is associated with a low intake
of some antioxidant nutrients in a Mediterranean population
of Spanish men.
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