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1 From the Unité Maladies Métaboliques et Micronutriments, INRA, Clermont-Ferrand/Theix, Saint-Genès-Champanelle, France (VT, NC, PG, VA-B, and PB); the Station de Technologie des Produits Végétaux, INRA Domaine St Paul, Avignon, France (CC-V and M-JA); and the Laboratoire de Nutrition Humaine, CRNH d'Auvergne, Clermont-Ferrand, France (CB).
2 The lutein and lycopene pills were donated by Rod Ausich (Kemin Industries Inc, Des Moines, IA) and Zohar Nir (Lycored Ltd, Beer-Sheva, Israel), respectively. 3 Address reprint requests to P Borel, INRA, UMMM, Centre INRA de Clermont-Ferrand/Theix, 63122 Saint-Genès-Champanelle, France. E-mail: patrick.borel{at}clermont.inra.fr.
ABSTRACT
Background: The results of epidemiologic studies have consistently shown associations between dietary intake or plasma carotenoid status and incidence of cancers and cardiovascular and eye diseases.
Objective: The aim was to assess whether vegetable-borne carotenoids (lycopene, lutein, and ß-carotene) compete for intestinal absorption and whether this affects the plasma status of carotenoids in the medium term (ie, after 3 wk).
Design: During 3-wk periods separated by 3-wk washout periods, 20 women were supplemented with either 96 g tomato purée/d (14.98 mg lycopene + 1.50 mg ß-carotene), 92 g cooked chopped spinach/d (11.93 mg lutein + 7.96 mg ß-carotene), 96 g tomato purée/d + 92 g chopped spinach/d, 96 g tomato purée/d + 2 lutein pills (12 mg lutein), or 92 g chopped spinach/d + 1 lycopene pill (15 mg lycopene). Plasma carotenoids were measured before and after each supplementation period. The subjects also participated in postprandial experiments in which they ingested meals containing double amounts of the supplements described above. Carotenoids were measured in chylomicrons to assess the interaction of carotenoids on absorption.
Results: Adding a second carotenoid to a meal that provided a first carotenoid diminished the chylomicron response to the first carotenoid. However, cosupplementation with a second carotenoid of a diet supplemented with a first carotenoid did not diminish the medium-term plasma response to the first carotenoid.
Conclusion: Consumption of carotenoids from different vegetable sources does not diminish plasma carotenoid concentrations in the medium term, despite the finding in postprandial testing of competitive inhibitory interactions among different carotenoids.
Key Words: Carotenoids • bioavailability • spinach • tomatoes • antioxidants • women • ß-carotene • lycopene • lutein • France
INTRODUCTION
The results of epidemiologic studies conducted over the past 30 y have consistently shown associations between dietary intake of fruit and vegetables and a reduced risk of several diseases. Among the components of fruit and vegetables, carotenoids—whose antioxidant properties have been clearly shown (1)—are likely to play an important role (2–10). In recent intervention trials, however, supranutritional doses of synthetic ß-carotene had unexpected and worrying results: smokers supplemented during 6 y with 25–50 mg ß-carotene/d had a higher incidence of lung cancer than did placebo-treated control subjects (11, 12). This finding contradicted the results of epidemiologic studies that consistently reported a protective effect against lung cancer of foods rich in ß-carotene (4). Such a contradiction illustrates the need for a better understanding of the biology of carotenoids in humans. Special attention should be given to the fate of intakes close to usual dietary habits, ie, with nutritional amounts of vegetable-borne carotenoids.
A key factor to investigate is the bioavailability of carotenoids, ie, the fraction of the ingested dose, possibly metabolized, that will reach the target site or sites. Intestinal absorption is a major determinant of bioavailability. Carotenoids are hydrophobic molecules, whose absorption pathway closely follows that of lipids (13). Numerous factors can interfere at different levels of this pathway, including competition between various lipid-soluble compounds or matrix effects (14–16). Interactions between carotenoids during intestinal absorption have been investigated in several studies (17). The available evidence in humans concerns only the interactions between purified or synthetic carotenoids: ß-carotene with lutein (18, 19), lycopene (20), or canthaxanthin (21, 22). Although the picture is not always consistent, it appears that providing various purified or synthetic carotenoids in the same meal results in modifications in the postprandial response of the carotenoids. Mechanisms potentially involved include competition for incorporation into lipid droplets (23) and then into mixed micelles, resulting in competition for intestinal absorption.
Carotenoids might also interact for incorporation and transport in lipoproteins. Another set of studies addressed the effect of supplementation with a single, purified carotenoid on the plasma concentrations of other carotenoids. Some studies showed no effect of supplementation with purified or synthetic ß-carotene on the concentrations of several plasma carotenoids (24–27). In contrast, other studies showed that purified or synthetic ß-carotene can diminish concentrations of lutein (28–30) and lycopene (31), and others showed that ß-carotene supplementation can even increase serum lycopene (32) and -carotene (29, 32).
In this study carried out in healthy volunteers, we investigated the interaction between vegetable-borne lycopene and lutein, 2 carotenoids with different hydrophobicities whose interactions have not been investigated so far. The interaction was studied by 2 means: with use of a postprandial study, which mainly examined the intestinal absorption step of the bioavailability concept, and in a supplementation study during which other physiologic and regulatory mechanisms may intervene.
SUBJECTS AND METHODS
Study population
Twenty young, female, nonsmoking volunteers aged 21–39 y were recruited for this study. None of the subjects took any oral medication, apart from oral contraceptives, or supplements of any kind both during the month before the study started and during the study period. The study was approved by the regional committee on human experimentation of the university hospital in Clermont-Ferrand (France), and we obtained written consent from each volunteer. The subjects were apparently healthy, according to clinical examination and disease history. Their lean and fat masses were measured by bioelectrical impedance analysis with a BIA 101A instrument (RJL Systems, Mt Clemens, MI). Fasting plasma lipid concentrations were measured to assess the normality of the subjects' lipid metabolism and their glycemia. The subjects' usual diet was monitored with use of a 5-d food recall. This dietary recall was analyzed for nutrient composition by using diet analyzer software (GENI; Micro 6, Nancy, France). The database of the software was completed for carotenoids with use of a carotenoid food-composition database (33).
Because of the potentially very long duration of the study, the subjects were randomly divided into 2 groups. Before the study began, we checked that there were no significant differences in all the variables measured between the 2 groups. Also, because the variability of response to carotenoids is high (34), it was important to verify that the carotenoid response did not differ significantly between the 2 groups (ie, that low and high responders were equally distributed). This point was checked by providing the same tomato purée meal to the 2 groups in the postprandial study and the same tomato purée diet to the 2 groups in the medium-term supplementation study.
Postprandial experiments
Postprandial experiments were performed on the first day of each supplementation period. After the subjects had fasted overnight for 12 h, an antecubital vein was catheterized with an intravenous cannula equipped with disposable obturators (Becton Dickinson, Meylan, France). A baseline fasting blood sample was collected and the subjects ingested a test meal within 20 min. The composition of the meal was as follows: 60 g wheat semolina (cooked and hydrated with 120 mL water), 40 g peanut oil, 2 pieces of bread (45 g), 1 cooked egg white (35 g), 1 serving of nonfat yogurt (125 g), and the carotenoid-rich source. Group 1 received 192 g tomato purée, 192 g tomato purée + 184 g chopped spinach, or 192 g tomato purée + 4 pills containing 6 mg lutein each as the carotenoid-rich source. Group 2 received 192 g tomato purée, 184 g chopped spinach, or 184 g chopped spinach + 2 pills containing 15 mg lycopene each. Blood samples were collected 2, 3, 4, and 6 h after the start of the meal. Samples were stored at -80°C until analyzed.
Medium-term (3-wk) supplementation experiments
During 3-wk study periods separated by 3-wk washout periods, the subjects ingested products rich in carotenoids in addition to their usual diet during lunch or dinner. The design was as follows: subjects in group 1 received 96 g pasteurized tomato purée/d (INRA, Unité expérimentale d’nologie de Pech rouge, Gruissan, France) for 3 wk. The composition of the tomato purée is shown in Table 1; 96 g provided 14.98 mg lycopene and 0.42 mg lutein. Then, after a 3-wk washout period (during which the subjects were asked to avoid consuming lycopene-rich products), group 1 received 96 g tomato purée/d + 92 g cooked chopped spinach/d (Stoc Supermarket, Clermont-Ferrand, France) for 3 wk. The composition of the chopped spinach is shown in Table 1; 92 g provided 11.93 mg lutein and no lycopene. Finally, after another 3-wk washout period, group 1 received 96 g tomato purée/d + 2 pills/d containing 6 mg lutein each and 0.3 mg zeaxanthin each. The carotenoid content of the pills was purified from marigold flowers (Kemin Industries Inc, Des Moines, IA).
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TABLE 1 . Composition of the tomato purée (96 g) and chopped spinach (92 g)1
Subjects in group 2 first received 96 g tomato purée/d. Then, after a 3-wk washout period (during which the subjects were asked to avoid consuming lycopene-rich products), group 2 received 92 g chopped spinach/d for 3 wk. Finally, after another 3-wk washout period, group 2 received 92 g chopped spinach/d + 1 pill/d containing 15 mg lycopene as 6% oleoresin extracted from tomatoes (Lycored Ltd, Beer-Sheva, Israel).
Note that a second dietary recall was performed during the first supplementation period (ie, during the supplementation with tomato purée). This showed that the energy and macronutrient intakes of the subjects did not change significantly during this period compared with their initial dietary intake. Note also that 92 g chopped spinach/d provided only 0.069 MJ/d when the subjects ate 7 MJ/d.
Plasma carotenoids, triacylglycerols, and cholesterol were measured in blood samples collected after the subjects had fasted for 12 h overnight the day before the start of each supplementation period and after a similar fasting period the day after each supplementation period had ended. Compliance was checked by regularly phoning the subjects, by reviewing a notebook in which the volunteers recorded when they ate the supplements, and by measuring plasma carotenoid concentrations after supplementation.
Chylomicron preparation
Blood was collected in EDTA-treated evacuated tubes and plasma was prepared immediately by centrifugation (910 x g, 4°C, 10 min). Chylomicrons were then isolated from 5 mL plasma layered under 5 mL NaCl (9 g/L) by ultracentrifugation (130000 x g, 28 min, 10°C). Chylomicrons were stored at -80°C under nitrogen until analyzed.
Analytic determinations
Plasma and chylomicron carotenoids were extracted twice with ethanol and hexane. Echinenone (Hoffmann-La Roche, Basel, Switzerland) was used as an internal standard. Carotenoids (lutein, lycopene, ß-carotene, -carotene, ß-cryptoxanthin, and zeaxanthin) were quantified by reversed-phase HPLC on a Kontron apparatus (Zurich, Switzerland) with detection at 450 nm (Kontron Detector 430). Carotenoids were separated by using 2 columns set in series (a Nucleosil C18, 150 x 4.6 mm, 3 µm; followed by a Vydac C18, 250 x 4.6 mm) purchased from Interchim (Montluçon, France). The mobile phase was a mixture of acetonitrile: methanol:dichloromethane:water (70:15:10:5, by vol). Quantification was conducted by using the Kontron MT 2 software.
Triacylglycerols and cholesterol were assayed in plasma and chylomicrons by using enzymatic colorimetric methods with commercial kits (Biomerieux, Craponne, France). The concentrations were measured spectrophotometrically (Hitachi U 2001; Hitachi Instruments, Inc, San Jose, CA) at 505 and 500 nm for triacylglycerols and cholesterol, respectively.
Statistical analysis
Results are expressed as means ± SEMs. The areas under the curves (AUCs) of the postprandial chylomicron responses were calculated by the trapezoidal rule. AUCs obtained in the postprandial experiment were compared by using a factorial one-way analysis of variance (ANOVA). When a significant (P < 0.05) difference was detected, means were compared by using the post hoc Tukey-Kramer test. For the medium-term supplementation experiment, ANOVAs for repeated measurements were used to determine whether there were significant variations in plasma carotenoids or lipids during supplementation. When a significant difference was detected, means were compared by using the post hoc Tukey-Kramer test. The statistical comparisons were performed with STATVIEW software (version 5.0; SAS Institute Inc, Cary, NC).
RESULTS
Characteristics of the study population
There were no significant differences in subject characteristics between the 2 groups at baseline (Table 2). Additionally, there were no significant differences in fasting plasma carotenoid concentrations between the 2 groups at baseline (Table 3).
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TABLE 2 . Subject characteristics at the beginning of the study1
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TABLE 3 . Fasting plasma carotenoid concentrations at the beginning of the study1
The chylomicron lycopene response (AUC) to the tomato meal was not significantly different between the 2 groups (data not shown), and the medium-term plasma lycopene, ß-carotene, and lutein responses to the tomato purée supplementation were not significantly different between the 2 groups. These results strongly suggest that the 2 groups did not respond differently to the main carotenoids studied (lycopene, ß-carotene, and lutein). Nevertheless, it cannot be totally discounted that the groups might have responded differently to other carotenoids.
Effect of different carotenoid-rich sources on postprandial chylomicron triacylglycerol responses
There was no significant difference between the overall chylomicron triacylglycerol responses (AUCs) obtained after the different meals. Also, there was no significant difference in the time at which the maximal concentration of chylomicron triacylglycerols appeared in the plasma (all the peaks were at 2 h) (data not shown).
Effect of lutein sources on postprandial chylomicron lycopene responses to tomato purée
The mean response curves of lycopene in the chylomicron fraction after meals providing tomato purée, tomato purée + chopped spinach, or tomato purée + lutein pills are shown in Figure 1A. Ingestion of tomato purée alone produced a rapid increase in chylomicron lycopene, with a peak at 2 h (59 ± 11 nmol/L). When a source of lutein was absorbed together with the lycopene source, the chylomicron lycopene response was diminished, as shown by the lower AUCs for the tomato + spinach meal and the tomato + lutein meal than for the tomato meal (191 ± 27 and 167 ± 17 compared with 273 ± 32 nmol·h/L, respectively; Figure 2A).
FIGURE 1. . Mean (±SEM) postprandial changes in lycopene (A), lutein (B), and ß-carotene (C and D) in the chylomicron fraction after the ingestion of a meal that provided either tomato purée (), tomato purée + chopped spinach (), tomato purée + lutein pills (•), chopped spinach (), or chopped spinach + one lycopene pill (). n = 10. Note that in C, ß-carotene was provided mainly by tomato purée whereas in D it was provided mainly by chopped spinach. For better clarity, the y axes of the different panels differ slightly.
FIGURE 2. . Mean (±SEM) areas under the curve (AUCs) of the chylomicron carotenoid responses obtained after the ingestion of a meal that provided either tomato purée (Tom), tomato purée + chopped spinach (Tom + Spi), tomato purée + lutein pills (Tom + Lut), chopped spinach (Spi), or chopped spinach + one lycopene pill (Spi + Lyc). Note that in C, ß-carotene was provided mainly by tomato purée whereas in D it was provided mainly by chopped spinach. For better clarity, the y axis of panel D is slightly different from the others. n = 10. Bars in the same panel with different letters are significantly different, P < 0.05 (factorial ANOVA followed by post hoc Tukey-Kramer test).
Effect of lycopene sources on postprandial chylomicron lutein response to chopped spinach
The chylomicron lutein responses obtained after the consumption of chopped spinach alone or of chopped spinach + lycopene-rich sources are shown in Figure 1B. Chylomicron lutein concentrations increased to a maximum peak at 4 h (39 ± 9 nmol/L) after the consumption of chopped spinach; the increase was less after the consumption of chopped spinach + tomato purée or + lycopene. Consequently, the overall chylomicron lutein responses were significantly lower when tomato purée or lycopene pills were added to the chopped spinach meal: 107 ± 13 and 108 ± 15 compared with 187 ± 26 nmol·h/L for the spinach + tomato meal, the spinach + lycopene meal, and the spinach meal, respectively (Figure 2B).
Effect of lycopene-rich or lutein-rich sources on postprandial chylomicron ß -carotene responses to ß -carotene–containing meals
The appearance of ß-carotene in the chylomicron fraction after the consumption of meals that provided ß-carotene (spinach and tomatoes contain appreciable amounts of ß-carotene) is shown in Figure 1, C and D. After the consumption of tomato purée (which provided 3 mg ß-carotene), the chylomicron ß-carotene concentration increased and peaked at 2 h (50 ± 6 nmol/L) and then declined slowly (Figure 1C). When chopped spinach or lutein pills were added to the tomato purée, the chylomicron ß-carotene response was diminished (Figure 1C), as shown by the significantly lower AUCs for the tomato + spinach meal and the tomato + lutein meal than for the tomato meal (151 ± 15 and 107 ± 6 compared with 245 ± 17 nmol·h/L, respectively; Figure 2C). Surprisingly, the chylomicron ß-carotene response to the chopped spinach meal that provided 16 mg ß-carotene (Figure 2D) was lower than its response to the tomato purée meal that provided 3 mg ß-carotene (Figure 2C). The addition of tomato purée or lycopene pills to the chopped spinach slightly but nonsignificantly diminished the chylomicron ß-carotene response (Figure 2D).
Effect of medium-term supplementation with carotenoid-rich sources on plasma carotenoid concentrations
The changes in plasma carotenoid concentrations after 3 wk of supplementation with the different sources of carotenoids are shown in Figures 3 and 4. Supplementation with the carotenoid-rich sources used in this study significantly affected the concentrations of all carotenoids measured except those of -carotene and ß-cryptoxanthin. Consumption of tomato purée for 3 wk increased plasma ß-carotene and lutein concentrations (Figure 3). Surprisingly, the increase in plasma lycopene, although detectable, was not significant. Chopped spinach supplementation induced a dramatic increase in plasma lutein (2.6-fold) and an increase in plasma ß-carotene (1.6-fold), but did not significantly affect plasma lycopene. Tomato purée + chopped spinach supplementation led to an increase in the 3 major carotenoids: 3.2-, 1.8-, and 1.4-fold for lutein, lycopene, and ß-carotene, respectively. Supplementation with tomato purée + lutein pills led to increases in lycopene and ß-carotene similar to those observed when tomato purée was ingested with chopped spinach, but the increase in plasma lutein was higher. The consumption of chopped spinach + lycopene pills produced results similar to those observed after spinach supplementation.
FIGURE 3. . Mean (±SEM) concentrations of the major plasma carotenoids during the 3-wk supplementation study in groups 1 (A) and 2 (B). , ß-carotene; , lutein; , lycopene; 3 wk Tom, after the consumption of 96 g tomato purée/d for 3 wk; 3 wk WO: after 3 wk of the subjects' usual diets (washout); 3 wk Tom + Spi, after the consumption of 96 g tomato purée/d + 92 g chopped spinach/d for 3 wk; 3 wk Tom + Lut, after the consumption of 96 g tomato purée/d + 2 lutein pills/d for 3 wk; 3 wk Spi, after the consumption of 92 g chopped spinach/d for 3 wk; 3 wk Spi + Lyc, after the consumption of 92 g chopped spinach/d + 1 lycopene pill/d for 3 wk. n = 10. There was a significant effect of diet on all plasma carotenoid concentrations, P < 0.05 (ANOVA for paired values). A filled symbol indicates that the mean value measured after supplementation was significantly different from the corresponding mean value measured before supplementation, P < 0.05 (post-hoc Tukey-Kramer test).
FIGURE 4. . Mean (±SEM) concentrations of other plasma carotenoids during the 3-wk supplementation study in groups 1 (A) and 2 (B). , -carotene; , zeaxanthin; , ß-cryptoxanthin; 3 wk Tom, after the consumption of 96 g tomato purée/d for 3 wk; 3 wk WO: after 3 wk of the subjects' usual diets (washout); 3 wk Tom + Spi, after the consumption of 96 g tomato purée/d + 92 g chopped spinach/d for 3 wk; 3 wk Tom + Lut, after the consumption of 96 g tomato purée/d + 2 lutein pills/d for 3 wk; 3 wk Spi, after the consumption of 92 g chopped spinach/d for 3 wk; 3 wk Spi + Lyc, after the consumption of 92 g chopped spinach/d + 1 lycopene pill/d for 3 wk. n = 10. There was a significant effect of diet on plasma zeaxanthin concentrations, P < 0.05 (ANOVA for paired values).
Concerning other carotenoids (Figure 4), only zeaxanthin was significantly affected by supplementation with carotenoid sources. Zeaxanthin increased markedly (although nonsignificantly compared with the values measured before supplementation) after supplementation with spinach or lutein pills (with or without tomato purée).
Effect of medium-term supplementation with carotenoid-rich sources on plasma lipids
Supplementation with the different sources of carotenoids did not significantly affect the concentrations of plasma triacylglycerols and cholesterol (Table 4).
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TABLE 4 . Change in plasma triacylglycerol and cholesterol during the 3-wk supplementation period1
DISCUSSION
The aim of this study was to assess whether there are interactions between vegetable-borne carotenoids regarding their bioavailability and whether this affects the medium-term plasma status of carotenoids. The data obtained also made it possible to draw important conclusions about the effects of carotenoid-rich sources on plasma lipids.
It is well established that consuming a diet rich in fruit and vegetables helps to reduce the risk of cardiovascular diseases. It has also been suggested that chylomicrons are involved in the etiology of these diseases. Thus, because fruit and vegetables are particularly rich in carotenoids, it was interesting to assess whether carotenoid-rich vegetables can affect the chylomicron triacylglycerol response to a fatty meal. The results were unambiguous: adding vegetable sources of carotenoids or purified carotenoids had no significant effect on the chylomicron triacylglycerol response to meals that provided 40 g fat. This result agrees with the results of others (36). Thus, the beneficial effect of fruit and vegetables on cardiovascular diseases is probably not the consequence of an effect of carotenoids on the time course of postprandial chylomicrons. The medium-term supplementation study performed in this protocol also enabled us to assess the effects of supplementation with carotenoids and vegetable purée on fasting plasma lipids. The finding of no significant changes in fasting plasma triacylglycerols or cholesterol after the consumption of carotenoid-rich diets (Table 4) confirms and extends to other carotenoids what was shown by others for lycopene (37).
The postprandial experiment was done to assess whether there are interactions between vegetable-borne carotenoids regarding their bioavailability (absorption or incorporation into chylomicrons). Adding chopped spinach to tomato purée diminished the chylomicron lycopene response (AUC) to tomatoes (Figures 1A and 2A). Note that this effect was not the consequence of a lower secretion of chylomicron triacylglycerols. The effect of adding purified lutein was similar to the effect observed with spinach. This strongly suggests that the factor in spinach that reduced the chylomicron lycopene response was lutein and that there was no additional effect of other spinach components (eg, fiber) on lycopene bioavailability. Adding lycopene, either in its natural vegetable matrix or as a purified supplement, to a spinach meal significantly decreased the chylomicron lutein response to spinach (Figures 1B and 2B). This also suggests that lycopene competes with lutein for incorporation into the chylomicron fraction and that the tomato matrix has no additional effect on lutein bioavailability.
The data obtained also made it possible to assess whether there was competition between lycopene or lutein and ß-carotene. The chylomicron ß-carotene response was lower when purified lutein was added to a meal that provided ß-carotene (the tomato meal in this case; Figures 1C and 2C). The chylomicron ß-carotene response was also lower, although nonsignificantly so, when purified lycopene was added to a meal that provided ß-carotene (the spinach meal in that case; Figures 1D and 2D). On the whole, these results suggest that lutein and possibly lycopene can diminish ß-carotene bioavailability.
The first conclusion of this paper is that lycopene, lutein, and ß-carotene, provided in their natural vegetable matrices, interact for incorporation into chylomicrons. This result agrees with previous results obtained with synthetic (22) and purified (19) carotenoids. The most likely mechanism for this interaction is competition between carotenoids for incorporation into the mixed micelles during digestion; note, however, that Garrett et al (38) did not find such evidence with use of Caco-2 cells. Thus, another explanation could be that there is competition between carotenoids for uptake and metabolism in the enterocyte or incorporation into the chylomicrons. Additional experiments are needed to identify the mechanism or mechanisms involved.
Having shown that carotenoids compete for incorporation into the chylomicron fraction, the question arises as to the consequence of this phenomenon on the plasma status of carotenoids. This is the question we aimed to answer in the medium-term supplementation experiment. The results obtained surprisingly showed that adding lutein-rich supplements to tomato purée enhanced the plasma lycopene response to tomato purée. More precisely, the plasma lycopene concentration increased by 16% after consumption of the tomato purée for 3 wk compared with the value measured before supplementation, whereas it increased by 85% and 92% after consumption of the tomato purée + spinach supplement or the lutein pill for 3 wk, respectively (Figure 3). This result agrees with the result of Johnson et al (20), who showed that the serum lycopene response was higher when lycopene was ingested with ß-carotene than when ingested alone. However, it apparently does not agree with the result of the postprandial experiment performed in the present study, which clearly showed that lutein diminished the chylomicron lycopene response to tomato purée. It also does not agree with the results of Castenmiller et al (15) and van het Hof et al (39), who showed that consumption of purified carotenoids decreases serum lycopene. To explain this apparent discrepancy, we suggest that other mechanisms were involved that overlapped the negative effect of lutein on lycopene bioavailability. The most probable hypothesis is that lutein induced an antioxidant-sparing effect, resulting in an enhanced status of lycopene in plasma. Indeed, the presence of cis isomers of lycopene and of numerous lycopene degradation products in human plasma (40) suggests that this carotenoid is particularly sensitive to oxidation.
Having shown that lutein can improve plasma lycopene status in the medium term, the question arises as to the effect of lycopene on lutein status. Our results showed that adding lycopene-rich sources to chopped spinach had no adverse effect on the 3-wk plasma lutein response. More precisely, plasma lutein increased 2.6-fold after the spinach diet, 2.3-fold after the spinach + lycopene diet, and 3.2-fold after the spinach + tomato diet (Figure 3). Thus, as previously discussed, there is a discrepancy between the results of the postprandial study, which showed that lycopene reduces the bioavailability of lutein, and the results of the medium-term supplementation experiment, which showed that there was no marked effect of cosupplementation with lycopene on the plasma lutein response to a lutein-rich diet.
Supplementation with tomato purée (which provided 14.98 mg lycopene and 1.50 mg ß-carotene) or purified lycopene did not significantly affect the plasma concentrations of the other carotenoids studied (zeaxanthin, -carotene, and ß-cryptoxanthin). Supplementation with spinach (which provided 11.93 mg lutein, 7.96 mg ß-carotene, and 0.3 mg zeaxanthin) or lutein pills (which provided 12 mg lutein and 0.3 mg zeaxanthin) increased plasma zeaxanthin by 80% and had no significant effect on plasma -carotene and ß-cryptoxanthin. This increase in zeaxanthin after supplementation with spinach or lutein pills was probably due to the zeaxanthin present in these lutein-rich sources. Taken together, these results suggest that lycopene, lutein, and ß-carotene do not markedly interfere with the bioavailability of zeaxanthin, -carotene, and ß-cryptoxanthin and that they do not affect the concentrations of these carotenoids in the plasma.
These results confirm and extend to other carotenoids the observations made in previous studies that ß-carotene supplementation does not affect the plasma concentration of other carotenoids (24, 26, 37). Note that although there was no apparent negative effect of bioavailability interactions on plasma carotenoid concentrations in the medium term, it cannot be totally excluded that systematic small interactions affect the long-term (eg, after months or years) storage and handling of carotenoids.
In conclusion, we clearly showed that there is competition between vegetable-borne lutein, lycopene, and ß-carotene regarding their appearance in the chylomicron fraction. This strongly suggests that these carotenoids compete for intestinal absorption, for incorporation into the chylomicrons, or both. However, the results of the medium-term supplementation experiment showed that this interaction has no adverse effect on the medium-term plasma status of carotenoids, suggesting that other mechanisms likely overlap the negative effect of carotenoid interaction on bioavailability. Future research should not depend solely on postprandial testing to examine these interactions because such testing can be misleading. Our findings strengthen the recommendation to eat a diet rich in a variety of fruit and vegetables rather than to attempt to eat a single carotenoid. Multiple carotenoids, at least over the short term, do not interfere with each other in terms of bioavailability and may provide some additive or synergistic preventive benefits.
ACKNOWLEDGMENTS
We thank Marion Brandolini for the dietary recall analysis; Lilianne Morin, Paulette Rousset, Sylvie Bureau, and Maryse Reich for technical assistance; and Eliane Albuisson for statistical advice.
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