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Department of Biochemistry, Biophysics & Molecular Biology, Iowa State University, Ames, IA 50011
4To whom correspondence should be addressed. E-mail: abarua{at}iastate.edu.
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ABSTRACT |
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 TOP ABSTRACT MATERIALS AND METHODSRESULTSRetinoids and carotenoids in...DISCUSSIONLITERATURE CITED |
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KEY WORDS: • ß-carotene • vitamin A • absorption • bioconversion • rats
Although vitamin A has been known as an essential micronutrient for almost 90 years (1
,2
), vitamin A deficiency (VAD)5 is still a major public health problem in developing countries (3
). Vitamin A can be obtained easily from animal sources. However, a large section of the human population depend on dietary carotenoids as the primary source of the vitamin. It is not clear whether the occurrence of VAD is due to the lack of consumption of a carotenoid-rich diet or to the many other factors that are involved in the bioavailability and bioconversion of the ingested carotenoids (4
–8
). Until recently, a dietary conversion factor of 6 µg of ß-carotene or 12 µg of other provitamin A carotenoids was regarded as equal to 1 µg retinol (ROL) (retinol equivalent) (9
). However, some studies had shown that the bioavailability of carotenoids was not as efficient as it was previously thought (4
,5
). Therefore, the Dietary Reference Intake Committee recently recommended that 12 µg of ß-carotene (BC) or 24 µg of other provitamin A carotenoids be considered equal to 1 µg ROL (retinol activity equivalent) (10
). It is not clear whether vitamin A status is another factor that plays a role in the bioavailability and bioconversion of BC to vitamin A.
The purpose of the present study was to gain further information concerning the bioavailability and bioconversion to vitamin A of a single oral dose of 3H-labeled-BC dissolved in corn oil or dispersed in aqueous Tween 80 in rats of different vitamin A status.
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MATERIALS AND METHODS |
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TOPABSTRACTMATERIALS AND METHODSRESULTSRetinoids and carotenoids in...DISCUSSIONLITERATURE CITED |
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Chemicals and solvents.
Hexane, methanol, dichloromethane, ethyl acetate, acetone and anhydrous diethyl ether (Fisher Scientific, Fair Lawn, NJ) were used. Whenever available, HPLC-grade solvents were used for HPLC.
Synthetic crystalline all-trans BC was a gift from Hoffmann-La Roche, Basel, Switzerland. 11,12-3H-ROL [250 µCi (9.25 MBq) in 125 µL ethanol containing 1 g/L d-
-tocopherol] was purchased from New England Nuclear, Boston, MA. 11,12-3H-ß-carotene (TBC) was chemically synthesized by a modification of a published procedure (11
) starting from 11,12-3H-ROL. The purity of the compound was tested by HPLC.
Measurement of radioactivity.
HPLC fractions were collected every minute in scintillation vials (20 mL) placed on a fraction collector (ISCO model Retriever IV, Lincoln, NE). Solutions or HPLC fractions were mixed well with Scintiverse BD (Fisher Scientific, Fair Lawn, NJ)(10 mL), and radioactivity was counted in a scintillation counter (Packard model 1600 TR).
Standard retinoids and ß-carotene.
The concentrations of standard ROL, retinoic acid (RA), retinyl acetate (RAc) and retinyl palmitate were determined from known E (1%, 1cm) (12
). The concentrations of TBC and newly formed retinoids in extracts were determined from the amount of radioactivity present in individual peaks obtained during HPLC analysis. The identities of individual compounds were determined not only from retention times but also by examining the absorption spectra with the photodiode array detector used during HPLC. Retinyl esters (RE) present in serum and tissues separated into several ester peaks, with retinyl palmitate the most predominant ester. Because all of the RE standards were not available, the concentration of other RE in serum and tissues was determined with respect to retinyl palmitate. Therefore, the concentration of RE reported in this study is the sum of all retinyl esters.
Preparation of the oral dose of 11,12-3H-ß-carotene.
The oral dose of TBC was prepared as described in previous studies (13
,14
). Briefly, a solution of TBC (2000 µg) in hexane was divided into two parts and transferred into separate mortars. The solvent in each mortar was evaporated under a gentle stream of argon.
Carotene in oil dose.
Corn oil (0.6 mL) was added to the residue in one mortar, and the mixture was ground until a clear solution was obtained. The solution was transferred to a glass vial, and the mortar was washed with 0.1-mL aliquots of corn oil (x 4) to transfer any residual carotene. The pooled solution was centrifuged at 500 x g for 1 min, and the supernatant was removed carefully. The concentration of TBC was determined as described below. This was the TBC in oil dose.
Carotene in aqueous Tween dose.
Tween 80 (500 µL) was added to the residue in the second mortar and mixed well with the pestle to dissolve the residue. Water (500 µL) was added and mixed well. The solution was transferred to a glass tube. The mortar was rinsed with water. The pooled solution was centrifuged at 500 x g for 1 min, and the supernatant was removed carefully. The concentration of TBC was determined as described below. This was the TBC in Tween 80/water dose.
Measurement of concentration and dosage of TBC.
Aliquots of both solutions were used to determine the exact concentration of TBC spectrophotometrically (11
). The radioactivity in each solution was measured. Both solutions were diluted appropriately to obtain a final concentration of 1.52 mmol/L of TBC with a specific activity of 35,777 dpm/µg. An aliquot of 100 µL of this corn oil or Tween 80/water solution containing 0.152 µmol (81.6 µg) (2.92 x 106 dpm) of TBC was administered to each rat.
Rats.
Weanling male Sprague-Dawley rats (n = 18) were purchased through Iowa State University Laboratory Animal Resources (LAR) and housed at a facility in the department maintained by LAR. All experiments were in accord with National Institutes of Health Guidelines for the Use of Animals and were approved by the University Committee on the Use of Animals in Research. The rats were kept in individual cages and fed a vitamin A–deficient diet (Diet No. 904646) (13
) supplied by ICN, Cleveland, OH, for 4 wk.
For daily supplementation with vitamin A, crystalline RAc was ground with corn oil in a mortar to obtain a clear solution (stock solution). An aliquot of the stock solution was dissolved in hexane, and the concentration of RAc was determined spectrophotometrically. The stock solution was diluted with varying amounts of corn oil to obtain 0.75, 2.24 and 6.72 mmol/L RAc.
The design of the feeding experiment is shown in Fig. 1
. The weanling rats (n = 18) were divided into 4 groups. Group 1 was fed 100 µL of corn oil only containing no RAc (-A). The second, third and fourth group received 100 µL of the corn oil solution containing 0.75 mmol/L (+A), or 2.24 mmol/L (++A), or 6.72 mmol/L (+++A) RAc so as to supply 7, 21 and 63 µg RAc/(rat · d), respectively.
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Serum.
The method for the extraction of retinoids and carotenoids in serum was described recently (13
–15
). However, the volume of serum used was 1 mL for each extraction. The volumes of extracting solvents were doubled accordingly.
Liver, small intestinal wall and mucosa, small intestinal contents, large intestine and stomach.
Because administered BC would constantly move from the stomach to the small intestine and then to the large intestine, there might be gradation in the distribution of the dose in different parts of these organs. To avoid such differences, except for liver, the entire organ and/or its contents were ground and mixed well. A portion of the ground tissue or contents was used for extraction and analysis of retinoids and carotenoids. Instead of extracting the whole liver, portions (
100 mg) of liver tissues were collected from different regions of the liver and ground; 1 g of the ground liver was used. The contents of the entire small intestine were removed by gently squeezing from the upper side of the small intestine downward with a pair of tweezers. The majority of rats did not excrete any feces during the 4-h study period. Feces excreted by one or two rats were mixed with the large intestine and its contents for extraction of carotenoids to make sure that any radioactivity and/or carotenoid excreted in the fecal matter was retained. The method of extraction of retinoids and carotenoids was the same as previously described (13
,15
). The pooled extract was evaporated completely and the residue was dissolved in 2-propanol/dichloromethane (2:1)(100–500 µL). Aliquots were analyzed by HPLC as described below.
Reversed-phase gradient HPLC.
Simultaneous analysis of carotenoids and retinoids was carried out by reversed-phase gradient HPLC procedures (13
,15
). A HPLC system (Waters, Milford, MA), equipped with Millennium software and a model 996 photodiode array detector was used along with a Rainin C18 Microsorb-MV 3-µm (3.6 x 100 mm) column.
Statistical analysis.
Comparisons of differences in radioactivity in serum, liver, other tissues, urine and feces of rats in the different groups were done with Student’s t tests (16
). Further analysis was performed using GraphPad Prism version 3.00 for Windows, GraphPad Software (San Diego, CA, www.graphpad.com). Data were evaluated by two-way ANOVA. Comparisons were performed across -A and +++A groups, and between groups of different vitamin A status. Using the same computer software, the unpaired t test was used to compare rats given a specific dose in oil or the aqueous dispersion. Newly formed retinoids in serum and liver were also compared. Differences with P <0.05 were considered significant.
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RESULTS |
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TOPABSTRACTMATERIALS AND METHODSRESULTSRetinoids and carotenoids in...DISCUSSIONLITERATURE CITED |
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HPLC analysis of the sera of rats fed the vitamin A–deficient diet receiving corn oil or RAc supplements for 4 wk showed varying levels of ROL. The concentration of ROL in serum of rats administered corn oil only (-A) was 0.63 ± 0.16 µmol/L. Concentrations in those receiving 7, 21 and 63 µg/d RAc were 0.83 ± 0.21, 1.84 ± 0.18 and 2.18 ± 0.29 µmol/L, respectively. No RE were detected.
3H-Retinoids in serum of rats after a dose of 11,12-3H-ß-carotene.
A total of 0.89–1.76% of the administered dose of radioactivity was recovered in the sera of rats 4 h after administration of TBC in oil or the Tween 80/water dispersion (Table 1
). HPLC analysis of the extracts showed 3H-ROL as the major radioactive peak in the sera of rats, but varying amounts of 3H-RE and 3H-RA were also present. The HPLC profile of the radioactive retinoids in the serum of an -A rat 4 h after the dose is shown in Figure 2A
. No TBC was detected in the serum of any rat. The concentration of newly formed ROL 4 h after the dose was significantly higher in the sera of -A rats than in +++A rats. On the other hand, the concentration of newly formed RE was significantly higher in +++A rats than in -A rats (Fig. 3
).
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Retinoids and carotenoids in organs and tissues |
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TOPABSTRACTMATERIALS AND METHODSRESULTSRetinoids and carotenoids in...DISCUSSIONLITERATURE CITED |
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Varying amounts of radioactivity (11.4–25.7%) were present in the stomach 4 h after the oral dose of TBC (Table 1)
. HPLC analysis of the stomach extracts showed that TBC (89%) was the predominant carotenoid (Fig. 4A
). The HPLC profile of the radioactive compounds in the stomach of an -A rat 4 h after the dose is shown in Figure 4
A. Two minor carotenoids were also present. One of these carotenoids has been tentatively identified as the 5,6-epoxide of TBC (9%). The other carotenoid (2%) resembled ß-cryptoxanthin in its spectrum and HPLC behavior (13
). No retinoids were detected in the stomach.
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The mucosa of rats administered TBC contained 0.87–3.2% of the administered radioactivity 4 h after the dose (Table 1)
. HPLC analysis of the extracts showed that 3H-RE predominated over 3H-ROL and TBC 4 h after the dose. 3H-RA was detected in most mucosa samples. In one sample, 3H-retinal (
max 370 nm) was identified as a major radioactive peak (41%) followed by 3H-ROL (max 325 nm) (25%), 3H-RE (23%)(max 326 nm) and TBC (10%)(max 485, 457, 428 nm). No form of apo-ß-carotenoids (aldehyde, alcohol, acid or ester) was detected in any mucosa sample. The HPLC profile of radioactive compounds present in the mucosa of an -A rat is shown in Figure 4
B.
Contents of small intestine.
An average of 8.27% of radioactivity was obtained from the contents of the small intestine (Table 1)
. HPLC analysis of the extracts showed TBC as the major radioactive compound (Fig. 4
C). Traces of other 3H-compounds were detected in the contents, but it was very likely that they were derived from degradation of TBC or from traces of mucosa remaining in the intestinal contents as a contaminant.
Large intestine and its contents.
At 4 h after the administration of TBC, almost 54% of the administered radioactivity was present in the large intestine and its contents (Table 1)
. HPLC analysis showed that > 90% of the radioactivity in the extracts was due to TBC (Fig. 4
D). Small amounts of 3H-carotenoids (10%), similar to the carotenoids seen in the stomach, were present. No retinoids were detected. The amount of radioactivity in the large intestine of rats fed the TBC dose in the aqueous dispersion was significantly higher than in those receiving the dose in oil (Table 1)
.
Liver.
The accumulation of radioactivity in the liver 4 h after administering TBC was 0.93–2.4% of the administered dose (Table 1)
. 3H-retinoids, but no TBC, were detected in the liver samples. The HPLC profile of the radioactive compounds in an extract of the liver of an -A rat 4 h after the dose is shown in Figure 2
B. At 4 h after the dose, the percentages of 3H-RE and 3H-ROL varied with the vitamin A status of the rats. The percentages of 3H-RE were 43, 50, 55, and 66 and those of 3H-ROL were 50, 43, 37 and 29 in -A, +A, ++A and +++A rats, respectively. Although more free ROL than RE was present in -A rats, more RE than free ROL was present in the livers of +++A rats. At 24 h after the dose, the concentration of free 3H-ROL had dropped to 9%, whereas the concentration of 3H-RE had increased to 91%.
Other tissues.
TBC was not detected in other tissues, viz., lung, kidney and heart. However, small amounts of radioactive retinoids were detected in these tissues. The percentages of the radioactivity recovered from these tissues were: kidney, 0.6%; lung, 0.01%; and heart, 0.05%. In kidneys, 3H-ROL (50–78%) predominated over 3H-RE (11–43%) 4 h after the dose. The HPLC profile of radioactive retinoids in the kidney of an -A rat 4 h after the dose is shown in Figure 2
C. A trace of 3H-RA was also present. In the heart, a small amount of radioactivity due to 3H-ROL was detected. The lungs contained predominantly 3H-RE (79%), followed by 3H-ROL (20%) and 3H-RA (Fig. 2
D).
Therefore, 85% of the administered dose of TBC was recovered intact and/or as retinoids (Table 1)
. The fate of the remaining 15% of the radioactivity is unknown.
Urine and feces.
Analysis of urine collected for 24 h after the dose showed that 0.57 ± 0.12 and 0.68 ± 0.27% of the administered dose was excreted in the urine by rats that received the oil and the aqueous dispersion, respectively. The excretions of radioactivity in the feces collected for 24 h after the dose were 49.23 ± 18.2 and 50.8 ± 5.56% from oil and the aqueous dispersion, respectively. Excretions of radioactivity were not affected by the vehicle used to administer the dose.
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DISCUSSION |
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TOPABSTRACTMATERIALS AND METHODSRESULTSRetinoids and carotenoids in...DISCUSSIONLITERATURE CITED |
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). A more reliable method would be to use a label such as 2H (17
) or 13C (18
) or a radioactive label such as 3H or 14C (19
–21
) in the BC molecule to obtain accurate information about absorbed BC and its metabolites. In this study, we used TBC to trace the distribution of BC and its metabolites. We used rats (22
) because it was possible to analyze all organs as well as plasma to study the distribution of the label. Rats were killed 4 h after the dose to examine the nature of the circulating metabolites in blood. Past studies have shown that newly formed retinoids disappear from circulation during transport for storage in liver and target tissues. Moreover, in a recent study in rats in which unlabeled ß-carotene was used, precise information about the distribution of the dose and its metabolites was known when the rats were killed within 4 h after the dose (23
). Excretion was minimal, and > 80% of the dose had moved from the stomach to the small intestine where absorption and conversion would occur (1
). Therefore, in this study we killed the majority of rats 4 h after the dose.
A few rats were killed 24 h after the dose to determine the rate of excretion of radioactivity. We found that a single oral dose of TBC resulted in absorption and conversion of BC to vitamin A as judged from appearance of radioactive retinoids in the serums, liver and other tissues of rats. The mean circulating radioactivity in serum of +++A rats fell from 1.25% at 4 h to 0.34% after 24 h (Table 1)
. During passage through the stomach, a small amount of the dose was converted, possibly by chemical means, to polar oxygenated compounds that were identified in a previous study in this laboratory (13
). The small intestinal mucosa contained predominantly 3H-retinoids and less TBC, and the contents contained predominantly TBC. The large intestine with its contents had intact TBC as the predominant compound along with small amounts of polar carotenoids that resembled the stomach oxycarotenoids. No retinoids were detected in the large intestine. In one of the classic studies on the intestinal conversion of carotene in rats, the highest concentrations of vitamin A were found in the middle sections and no vitamin A was found in the large intestine (24
,25
). During the present study, a separate experiment was carried out to study the conversion, if any, of TBC to retinoids as follows: a rat was anaesthetized, and a small incision was made to reach the large intestine. TBC dissolved in a few drops of dimethyl sulfoxide was injected directly into the mid-section of the large intestine and the rat was returned to its cage. The rat was killed 4 h after the injection. No radioactivity was detected in serum or liver. The result of the present study is in agreement with the earlier findings (1
,24
,25
) that once carotene enters the large intestine, it is neither absorbed nor converted to vitamin A.
The radioactivity in serum, liver, kidney and lung was associated with retinoids only and accounted for < 5% of the dose of TBC. If the absorption and conversion of the remaining TBC in the stomach and small intestine continued at the same rate as that at which the dose moved from the stomach into the small intestine, an additional 1.2% of the radioactivity would appear as retinoids in serum, liver and other tissues. Thus, it can be expected that at the end of digestion, a maximum of 5–6% of radioactivity would appear as retinoids from a single oral dose of BC. The present findings reveal that the absorption of BC was inefficient, and that only a small amount of BC was converted to vitamin A (19
).
The rats used in the present study were of different vitamin A status. The absorption and conversion of ß-carotene to vitamin A did not differ among the groups of different vitamin A status 4 or 24 h after the dose (Table 1)
. Similarly, the absorption or conversion generally did not differ between rats administered the dose in oil or in the aqueous dispersion. However, significantly more of the dose was found in the large intestine and its contents 4 h after the dose in oil when the groups were combined, indicating that it was retained longer than in the rats administered the dose in the aqueous dispersion. It should be emphasized that the number of rats used per group was too small to draw any definite conclusions. A future study will use a larger number of rats and repeated daily doses of BC for several days to determine whether vitamin A status and the vehicle used affect absorption. The present results pertain to rats and may not be applicable to other species, including humans.
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FOOTNOTES |
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2 Supported by U.S. Department of Agriculture-NRICGP 00–37200-4290 and National Institutes of Health DK 39733.
3 Present address: Department of Chemistry, Gauhati University, Guwahati 781014, India.
5 Abbreviations used: BC, ß-carotene; RAc, retinyl acetate; RA, retinoic acid; RE, retinyl ester; ROL, retinol; TBC, 11,12-3H-ß-carotene; VAD, vitamin A deficiency.
Manuscript received 22 August 2002. Initial review completed 13 September 2002. Revision accepted 18 October 2002.
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LITERATURE CITED |
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TOPABSTRACTMATERIALS AND METHODSRESULTSRetinoids and carotenoids in...DISCUSSIONLITERATURE CITED |
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1. Moore, T. (1957) Vitamin A 1957 Elsevier London, UK.
2. Olson, J. A. (1999) Carotenoids. Shils, M. E. Olson, J. A. Shike, M. Ross, A. C. eds. Modern Nutrition in Health and Disease 9th ed. 1999:525-541 Williams & Wilkins Philadelphia, PA. .
3. Underwood, B. (1994) Vitamin A in human nutrition: public health considerations. Sporn, M. B Roberts, A. B. Goodman, D. S eds. The Retinoids 2nd ed. 1994:211-227 Raven Press New York NY. .
4. Castenmiller, J.J.M. & West, C. E. (1998) Bioavailability and bioconversion of carotenoids. Annu. Rev. Nutr. 18:19-38.[Medline]
5. dee Pee, S. & West, C. E. (1996) Dietary carotenoids and their role in combating vitamin A-deficiency: a review of literature. Eur. J. Clin. Nutr. 50:S38-S63.
6. Yeum, K. & Russell, R. M. (2002) Carotenoid bioavailability and bioconversion. Annu. Rev. Nutr. 22:483-504.[Medline]
7. Erdman, J. W., Jr, Bierer, T. L. & Gugger, E. T. (1993) Absorption and transport of carotenoids. Ann. N.Y. Acad. Sci. 691:76-85.[Medline]
8. Parker, R. S. (1997) Bioavailability of carotenoids. Eur. J. Clin. Nutr. 51:S86-S90.
9. FAO/WHO Joint Expert Consultation (1988) Requirements of vitamin A, iron, folate and vitamin B12. FAO Food and Nutrition Series No. 23 1988 FAO Rome, Italy.
10. Food and Nutrition Board Standing Committee of the Scientific Evaluation of Dietary Reference Intakes (2001) Dietary Reference Intakes for vitamin A, vitamin K, arsenic, boron, chromium, copper, iodine, iron, manganese, molybdenum, nickel, silicon, vanadium and zinc 2001 Institute of Medicine, National Academy of Sciences Washington, DC.
11. Bergen, H. R. & Olson, J. A. (1989) Synthesis of deuterated ß-carotene. J. Label. Compd. Radiopharm. 27:783-786.
12. Barua, A. B, Olson, J. A, Furr, H. C. & van Breeman, R. B. (2000) Vitamin A and carotenoids. De Leenheer, A. P. Lambert, W. E. Bocxlaer, J. F. eds. Chromatographic Analysis of Vitamins 3rd ed. 2000:1-74 Marcel Dekker Basel, Switzerland .
13. Barua, A. B. & Olson, J. A. (2000) ß-Carotene is converted primarily to retinoids in rats in vivo. J. Nutr. 130:1996-2001.
14. Barua, A. B. & Olson, J. A. (2001) Xanthophyll epoxides, unlike ß-carotene monoepoxides, are not detectibly absorbed by humans. J. Nutr. 131:3212-3215.
15. Barua, A. B. & Olson, J. A. (1998) Reversed-phase gradient high-performance liquid chromatographic procedure for simultaneous analysis of very polar to nonpolar retinoids, carotenoids and tocopherols in animal and plant samples. J. Chromatogr. A. 707:69-79.[Medline]
16. Snedecor, G. W. & Cochran, W. G. (1989) Statistical methods 8th ed. 1989:83-106 University Press Ames, IA.
17. Tang, G., Grusak, M. A., Qin, J., Dolnikowski, G. G. & Russell, R. M. (1999) Vitamin A activity of spinach determined by isotope reference method 1999 Report of XIX International Vitamin A Consultative Group Meeting Washington, DC.
18. Parker, R. S., Swanson, J. E., Marmor, B., Goodman, K., Spielman, A. B., Brenna, J. T., Viereck, S. M. & Canfield, W. K. (1993) Study of ß-carotene metabolism in humans using 13C-ß-carotene and high precision isotope ratio mass spectrometry. Ann. N.Y. Acad. Sci. 691:86-95.[Medline]
19. Chen, Y., Oace, S. M. & Wolf, G. (1999) Studies on the effect of dose size on the absorption of ß-carotene by the rat in vivo. Int. J. Vitam. Nutr. Res. 69:8-15.[Medline]
20. Krinsky, N. I., Mathews-Roth, M. M., Welankiwar, S., Seghal, P. K., Lausen, N. C. & Russell, M. (1990) The metabolism of [14C]-ß-carotene and the presence of other carotenoids in rats and monkeys. J. Nutr. 120:81-87.
21. Ducker, S. R., Lin, Y., Buchholz, B. A., Schneider, P. D., Lame, M. W., Segall, H. J., Vogel, J. S. & Clifford, A. J. (2000) Long-term kinetic study of ß-carotene, using accelerator mass spectrometry in an adult volunteer. J Lipid Res. 41:1790-1800.
22. Lee, C. M, Boileau, A. C., Boileau, T. W., Williams, A. W., Swanson, K. S., Heintz, K. A. & Erdman, J. W., Jr (1999) Review of animal models in carotenoid research. J. Nutr. 129:2271-2277.
23. Barua, A. B. (2002) Absorption and conversion of a single oral dose of carotene in corn oil to vitamin A in Sprague-Dawley rats with low reserve of vitamin A. Int. J. Vitam. Nutr. Res. (in press).
24. Thompson, S. Y., Ganguly, J. & Kon, S. K. (1949) The conversion of ß-carotene to vitamin A in the intestine. Br. J. Nutr. 3:50-78.
25. Thompson, S. Y., Braude, R., Coates, M. E., Cowie, A. T., Ganguly, J. & Kon, S. K. (1950) The conversion of ß-carotene to vitamin A in the intestine. Br. J. Nutr. 4:398-420.
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