Major carotenoids in diet, serum and tissues of humans include the hydrocarbon carotenoids -carotene (BC) and lycopene (Krinsky 1993). Both BC and lycopene have antioxidant properties, whereas only BC has pro-vitamin A activity. There are numerous reports of large interindividual variability in the serum concentration of BC, and the serum response to a single oral dose of BC is also highly variable (Brown et al. 1989, Dimitrov et al. 1988, Johnson and Russell 1992, Mathews-Roth and Gulbrandsen 1974). Determinants of serum concentration of BC include dietary factors such as fat and fiber, efficiency of absorption, rate of tissue uptake, and rate of BC metabolism (Bowen et al. 1993). Little information is available on the determinants of serum concentrations of lycopene. A correlation between 7-d food diary lycopene intake and plasma lycopene has been reported (Forman et al. 1993). However, Campbell et al. (1994) reported that there does not seem to be a correlation between serum lycopene and total fruit and vegetable intake. It is not known whether there is large variability in the serum response to an oral dose of lycopene, similar to that for BC. Moreover, it is not known whether, within individuals, the serum response to an oral dose of BC is similar to that of lycopene. Furthermore, it is not known if large doses of BC or lycopene affect the absorption and clearance of the other, as has been shown for BC and canthaxanthin (White et al. 1994) and BC and lutein (Kostic et al. 1995).

This study investigates the serum response in adult human subjects after oral doses of BC and lycopene or combined doses.

SUBJECTS AND METHODS

Table 1. Characteristics of 10 men studied [View Table]

Subjects (n = 10) were housed at the Human Nutrition Research Center for three separate 2-d periods. Two weeks before the study, 10 mL of blood was drawn from fasting subjects to determine the basal concentrations of carotenoids, cholesterol and triglycerides. At that time, subjects were instructed by the metabolic dietician to consume a low carotene diet while at home to lower blood carotene concentrations. Food diaries were kept for 3 d out of each of the 2-wk periods before the study to confirm low carotene consumption. On admission to the Nutrition Center (d 0), subjects consumed at 0800, 1200 and 1800 h a 3138 kJ meal of a liquid diet formulated to contain protein (22 g), carbohydrate (116 g) and a standardized amount of fat (21 g, 25% of total energy) and no carotenoids or retinoids. The liquid diet also contained vitamin E (4.3 mg -tocopherol equivalents), and the percentages of energy from saturated, monosaturated and polyunsaturated fatty acids were 13, 7 and 2%, respectively. The liquid diet contained no fiber.

The study involved the oral ingestion of individual and combined 0.11-mmol (60-mg) doses of all-trans BC (100%, Henkel Corp., LaGrange, IL) and all-trans lycopene (97%, 1% 13-cis lycopene, Makhteshim-Agan of North America, Inc., New York, NY). Doses of BC and lycopene crystals, dispersed in corn oil (1 and 5%, respectively), were provided as gelatin capsules. On d 1 of the study, each subject consumed either BC or lycopene or both with the liquid diet after the first blood sample (10 mL) was taken. The dose type was randomized. Blood (10 mL) was drawn at 1, 3, 5, 7, 9 and 12 h after the carotene dose. The subject consumed the two other 3138 kJ meals at 4 and 10 h after dosing. Subsequently, 10 mL of blood was collected after a 12-h overnight fast. Blood was collected in a Vacutainer with no additives. Serum samples were prepared (1000 × g for 15 min at 4°C) and stored at 70°C for subsequent analysis of carotenoids. After a 2-wk washout period, during which subjects consumed a low carotene diet, study d 0-2 were repeated using another carotenoid dose type. After another 2-wk washout period, during which subjects consumed a low carotenoid diet, study d 0-2 were repeated using the remaining carotenoid dose type.

RESULTS

Serum concentrations of lycopene after ingestion of lycopene or combined doses of BC and lycopene are presented in Figure 2. After the lycopene dose, the mean serum lycopene concentrations significantly increased from baseline at 5 h (P < 0.008). Thereafter, the mean concentration returned to baseline. Serum BC concentrations did not change after the single dose of lycopene (data not shown). Ingestion of a combined dose of BC and lycopene resulted in a gradual increase in the mean concentration of serum lycopene from baseline, which was significant at 24 h (P < 0.05).

The 24-h AUC for serum BC was not different when BC was ingested alone or with lycopene (Table 2). However, the AUC for lycopene was significantly greater when lycopene was ingested with BC (P < 0.05).

Table 2. Serum -carotene and lycopene responses in men given an oral dose of -carotene, lycopene or a combined dose1 [View Table]

The individual serum BC response to the BC dose was not related to the serum lycopene response to the lycopene dose. That is, the 24-h peak concentration and AUC for the BC dose were not correlated to peak concentration and AUC for the lycopene dose (r = 0.285 and 0.018, respectively). In all three dose tests, the intraindividual responses varied.

The serum response (peak concentration, 24-h AUC) was not related to age, pre-study serum BC or lycopene concentrations, or baseline (0 h) BC or lycopene concentrations (data not shown). Baseline concentrations of BC or lycopene were not different among the three tests (Table 2).

Nine of the 10 subjects had a peak BC serum concentration at 12 h or later and one subject had a peak BC serum concentration at 5 h after the BC dose. Eight subjects had a peak BC serum concentration at 12 h or later and two subjects had a peak BC serum concentration at 12 h or later after the combined dose. The time of peak concentration for serum lycopene was earlier for most subjects after the single dose of lycopene. Seven subjects had a peak concentration during the first 12 h after the dose, and three subjects had a peak at 12 h or later. After the combined dose, three subjects has a peak serum concentration of lycopene during the first 12 h and seven subjects had a peak serum concentration at 12 h or later.

DISCUSSION

The determinants of the serum response of BC and lycopene are not known, but one might anticipate that they would be similar. -Carotene and lycopene are major carotenoids in the Western diet and in human serum and tissues (Krinsky 1993). For both BC and lycopene, the serum response to the orally ingested carotenoid is improved when the carotenoid is heated with small amounts of oil (Erdman et al. 1993, Stahl and Sies 1992). Also, the efficiency of the response to these dietary carotenoids is greater at low than at higher doses (Erdman et al. 1993, Stahl and Sies 1992). Both carotenoids are nonpolar and transported exclusively by lipoproteins (Clevidence and Bieri 1993). Given these similarities in their serum response and transport, it is possible that, for an individual, the serum response to one carotenoid would be similar to that of the other.

In the present study, the response to a single oral dose of BC varied among the subjects. The meal given with the BC dose was designed to be high in fat to facilitate carotenoid absorption, and the BC dose was large (0.86 mg/kg for a 70-kg man) with the intent to produce measurable changes in the serum BC concentration. Even so, the range of the increase in BC concentration varied from 17 to 285 nmol/L for the 10 subjects. Our laboratory, as well as others, has reported irregularities in the serum response to a single oral dose of BC (Brown et al. 1989, Dimitrov et al. 1988, Johnson and Russell 1992, Mathews-Roth and Gulbrandsen 1974). A poor response to the BC dose suggests that in some individuals, the serum BC concentration may be maintained over time and is the result of long-term dietary intakes, being little affected by a single large dose. Variation in the serum response to the BC dose could be due to individual differences at several points of BC metabolism, including intestinal absorption, release into and clearance from the circulation, tissue uptake, and release from body stores.

Similar to the response to BC, the response to the single oral dose of lycopene varied among the subjects, with changes in concentrations over the 24-h period ranging from 0 to 257 nmol/L. However, the serum response to the lycopene dose was not related to the serum response to the BC dose. Our results suggest that each carotenoid, when administered alone, has different absorption kinetics. However, the serum response also reflects release into and clearance from the circulation, tissue uptake, and release from body stores, and these may differ between the two carotenoids. In fact, differences in the organ distribution of lycopene and BC have been reported (Kaplan et al. 1990, Stahl et al. 1992).

During the first 24 h of absorption, the peak concentration times for each carotenoid when ingested alone provide further evidence for differing absorption kinetics for BC and lycopene. The time at which serum BC concentrations peaked after the BC dose was significantly later than the peak concentration time for serum lycopene after the lycopene dose. This could mean that, after enterocyte uptake, the release of BC is slower relative to lycopene. This may be due to the presence of enzymes that can bind to BC and possibly delay release from the enterocyte. Previous work from this laboratory and others provides evidence for delayed release of BC from the intestine (Henderson et al. 1989, Johnson and Russell 1992). The earlier peak concentration time for lycopene may be due to a more rapid uptake and release from the enterocytes rather than a faster clearance from the circulation.

In this study, we used the 24-h AUC of the serum concentration vs. time to estimate the early absorption events of BC and lycopene from the gastrointestinal tract. We chose to study the serum carotenoid response for up to 24 h after the administration of the oral dose based on our previous work that showed serum BC concentrations decreased after this time point (Johnson and Russell 1992), although other studies have reported peak serum concentrations to be as late as 48 h after a single oral dose (Brown et al. 1989, Kostic et al 1995, Stahl and Sies 1992). On the basis of the 24-h AUC, lycopene had little effect on the absorption of BC, whereas the combined dose of BC and lycopene enhanced the absorption of lycopene. This observation is due either to an improvement in the serum response of lycopene in the presence of BC or to a very short absorptive period for lycopene. As seen in Figure 1, BC, either in the presence of absence of lycopene, shows a maximum absorption at 24 h, the last time point measured in this study. This time course is similar to those in other studies in the literature (Brown et al. 1989, Johnson and Russell 1992). However, the time course for the lycopene absorption was unusual, in comparison to that reported by Stahl and Sies (1992). Using lycopene present in tomato juice boiled with 1% corn oil for 1 h, these authors found maximum absorption at 24-48 h, whereas we found the maximum at 5 h and a return to baseline by 12-24 h. It is possible that the tomato juice matrix may affect lycopene absorption. We found that only in the presence of BC did the lycopene absorption curve appear similar to that reported for other carotenoids. Why then did administration of lycopene alone give such an unusual absorption pattern? There is a suggestion that lycopene absorption differs from that of the other carotenoids. Campbell et al. (1994) reported that the plasma carotenoid concentrations serve as excellent markers of fruit and vegetable intake, with the exception of plasma lycopene. Although this may be due to absorption, transport or storage differences, our observations would indicate that lycopene administered by itself is not well absorbed, with a very early peak at 5 h. This increase at 5 h was only about half of the maximum increase of serum lycopene achieved after the combined dose. There is the possibility, however, that in this study the peak response occurred later that 24 h.

The results presented in Figure 2 are in sharp contrast to the observation of Prince et al. (1991), who suggested that high dose BC supplementation (300 mg/d for 21 d added to a self-selected diet) resulted in a decrease in serum lycopene concentrations, which they attributed to a possible competition in absorption. The differing results may be due to the differences in dose size and duration of supplementation.

An alternate explanation for the finding of an improved lycopene absorption when administered with BC is that some components in the BC suspension enhanced the solubilization of crystalline lycopene and thereby provided a better serum response. This effect would be a physical-chemical one, however, dependent upon the nature of the preparations of administered carotenoids but not of biological events in the gastrointestinal tract. Other investigators have reported what seems to be the facilitation of absorption of one carotenoid by another. For example, when chicks are supplemented with a mixture of 9-cis-BC and all-trans-BC, they show a marked increase in the absorption of all-trans-BC, in comparison to receiving only all-trans-BC (Ben-Amotz et al. 1989, Mokady et al. 1990). However, the carotenoid preparations were likely well solubilized given that the carotenoid capsules, containing vegetable oil, were dispersed in the high fat liquid meal, and identical procedures were used with both the single and the combined doses.

Our study provides evidence that ingestion of a combined dose of BC and lycopene has minimal effect on the early absorption events of BC but improves those of lycopene. Even when we doubled the mass of carotenoid ingested by adding lycopene, we observed no difference in the 24-h peak absorption of BC. Because carotenoid absorption normally decreases with increased dosage (Brown et al. 1989, Stahl and Sies 1992), our findings suggest that there may be independent pathways for BC and lycopene absorption. Furthermore, the evidence suggests that the presence of BC mobilized the lycopene absorption pathway, which allowed the lycopene to be absorbed to the same extent as the BC when these two carotenoids were administered together. These results should be contrasted to those of Kostic et al. (1995), who reported that BC reduced lutein absorption, whereas lutein had a mixed effect on BC absorption. These differences may be attributable to the fact that both BC and lycopene are hydrocarbons, as opposed to lutein, a hydoxylated carotenoid that may have a different absorption mechanism.