QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Abstract Full Text (PDF) Purchase Article View Shopping Cart Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Edwards, A. J. Articles by Parker, R. S. Search for Related Content PubMed PubMed Citation Articles by Edwards, A. J. Articles by Parker, R. S.

- and ß-Carotene from a Commercial Carrot Puree Are More Bioavailable to Humans than from Boiled-Mashed Carrots, as Determined Using an Extrinsic Stable Isotope Reference Method

Division of Nutritional Sciences and Department of Food Science, Cornell University, Ithaca, NY 14853 and ^* Gerber Products Company, Fremont, MI 49413

²To whom correspondence should be addressed. E-mail: rsp3{at}cornell.edu.

	ABSTRACT

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
The extent to which processing affects the carotene or vitamin A value of foods is poorly understood. An extrinsic reference method was used to estimate the mass of carotenes and vitamin A derived from various preparations made from the same lot of carrots. Using a repeated-measures design, nine healthy adult subjects consumed test meals of either carrot puree (commercial baby food) or boiled-mashed carrots on separate days; six of the subjects also consumed a test meal of raw-grated carrot. Test meals supplied 34.7 µmol (18.6 mg) carrot ß-carotene (ßC), plus 6 µmol deuterium-labeled retinyl acetate (d₄-RA) in oil solution. Baseline-adjusted carotene and retinyl ester (R-ester) area-under-curve (AUC) responses in the triacylglycerol-rich lipoprotein (TRL) fraction (0–8.5 h) were determined using HPLC and gas chromatography-mass spectrometry. The masses of absorbed ßC, -carotene (C) and R-ester were estimated by comparing their AUC values with that of deuterium-labeled retinyl ester (d₄-R-ester), assuming the latter represented 80% of the d₄-RA reference dose. Absorption of ßC and C was approximately twofold greater from carrot puree than from boiled-mashed carrots, whereas the retinol yield was only marginally (P = 0.11) influenced by treatment. Carotene and R-ester absorption from raw-grated carrot was intermediate to, and did not differ significantly from the cooked preparations. The vitamin A yield (puree, 0.53 mg; boiled-mashed, 0.44 mg) of cooked carrot containing 18.6 mg ßC was substantially less than that predicted by current convention and limited primarily by intestinal carotene uptake. Processing can therefore significantly improve bioavailability of carrot carotenes, and in some cases influence the carotene value more than the intrinsic vitamin A value.

KEY WORDS: • provitamin A carotenoids • ß-carotene • retinol equivalency • stable isotope • bioavailability • humans

	INTRODUCTION

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
Provitamin A carotenoids, predominantly ß-carotene (ßC)³ and -carotene (C), are important sources of vitamin A, particularly in countries where vitamin A deficiency is prevalent and sources of preformed vitamin A are consumed infrequently (1 ). Quantitative information regarding the mass of vitamin A actually derived from provitamin A–containing foods is required to formulate recommendations for prevention of vitamin A deficiency through consumption of carotene-rich foods. Currently, there is no "gold standard" against which to compare the metabolic vitamin A equivalency of different plant foods or the influence of food processing on the amount of vitamin A or carotenoids assimilated from those foods. We recently described a novel modification of the triacylglycerol-rich lipoprotein (TRL) response model in conjunction with use of an extrinsic reference standard of deuterium-labeled retinyl acetate (d₄-RA) to estimate absolute bioavailability of provitamin A carotenes from foods (2 ,3 ).

The retinol activity equivalency (RAE) ratios for ßC and for C from foods were set recently at 12:1 and 24:1, respectively (4 ), a factor of two higher than the previous ratios used to determine retinol equivalency (5 ). These ratios reflect the relatively poor absorption efficiency of provitamin A carotenoids from foods as reported in recent studies (2 ,6 –8 ). Carotenes have been reported to be more bioavailable in purified form than from foods (6 ,9 –14 ). Key meal-based factors in the bioavailability of carotenoids include the presence of fat (15 –18 ) and fiber (19 –22 ) and the nature and surface area of the food matrix itself (23 –25 ), which may be altered by processing. Carotenoid release from the food matrix is required before the carotenoids can be taken up with mixed lipid micelles in the intestine, as reviewed by Parker (26 ). Heat and/or homogenization have been shown to improve bioavailability of ßC from green leafy vegetables (25 ), carrots (23 ,24 ,27 ) and tomatoes (28 ), as assessed by change in serum ßC. However, there are limited data on the mass of metabolic retinol (ROH) actually derived from those foods under meal conditions.

Measurement of the ßC and retinyl ester (R-ester) response in the TRL ( < 1.006 kg/L) fraction of plasma has been applied previously to assess carotenoid bioavailability from water-dispersible ßC beadlets (29 ), tangerine concentrate (30 ), tomatoes and tomato paste (31 ), and ßC supplements (32 ). The TRL approach has the advantage of focusing analysis on the chylomicron (CM)-rich plasma fraction, which contains newly absorbed lipids (including ßC and R-ester) from a recent test meal. However, as it has been applied to date, the TRL approach has been limited due to a lack of control for interindividual variation in CM clearance kinetics or for variation in R-ester recovery in the TRL fraction during ultracentrifugation.

The approach developed in our laboratory (2 ,3 ) involves coadministration of a reference dose of d₄-RA with a ßC-containing test meal. This method permits estimation of the mass of unlabeled vitamin A or intact ßC derived from a test meal containing provitamin A carotenoids (such as ßC) as the sole source of vitamin A. The d₄-RA serves as an "internal standard," controlling for variation in CM clearance kinetics in vivo and for R-ester recovery during sample processing. The d₄-RA also constitutes a reference against which to compare and estimate the mass of unlabeled R-ester derived from cleavage of provitamin A carotenes absorbed from the test meal.

Cooked, homogenized carrots have previously been shown to produce a twofold greater increase in serum ßC than raw carrots, an effect attributed to a reduced particle size of the food (24 ). In the present study, we applied the TRL/d₄-RA approach to compare masses of vitamin A and ßC derived from carrots prepared three ways, each representing a typical form in which carrots are consumed. Test meals contained a commercially processed carrot puree, boiled-mashed carrots or raw-chopped carrots. Carrot meals were all prepared from a single carrot source, but varied in both heat and mechanical treatment conditions.

	SUBJECTS AND METHODS

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
Subjects.

All procedures involving human subjects were approved by the Cornell Committee on Human Subjects Research. Nine apparently healthy, nonsmoking human subjects (5 women and 4 men, ages 23–42 y) were recruited from the Cornell community. Subjects were screened by health history interview and clinical evaluation, and exhibited no evidence of chronic gastrointestinal disease, liver disease or hyperlipidemias. Subjects reported no use of vitamin A or ß-carotene supplements within the past 2 mo. The average body mass index of the nine subjects was 23.3 ± 3.6 kg/m².

Treatments.

A single-dose, pharmacokinetic approach was utilized, in which subjects were fed a morning test meal and blood samples were collected at baseline and at five time points after the meal up to 8.5 h. There were three treatments: 1) carrot puree, 2) boiled-mashed carrot and 3) raw-chopped carrot. All nine subjects were given the first two (cooked carrot) treatments; a subset of 6 subjects was also given the raw carrot treatment. An incomplete randomized block design was used, with a minimum 2-wk washout period between treatments. Subjects were instructed to avoid carotenoid-containing and vitamin A-rich foods for 24 h before each trial to clear the upper gastrointestinal tract of these substances. A list of permitted foods and a list of foods to be avoided were provided to subjects, who completed a food diary for this period. All subjects fasted between their evening meal (consumed before 2000h) and reporting to the laboratory the following morning for the trial period. Subjects were fed test meals individually on separate trial dates. All food consumed during the test day was consumed on site in the presence of at least one investigator to verify complete consumption of the test meal and the absence of outside sources of carotenes or vitamin A.

All test meals contained 6.0 µmol (2 mg) d₄-RA dissolved in safflower oil as described previously (3 ). The d₄-RA solution was prepared in bulk for all trials, and stored under N₂ at 4°C until use. Deuterated (10, 19, 19, 19-²H) retinyl acetate (Cambridge Isotope Laboratories, Andover, MA) was of >98% chemical purity and was determined by gas chromatography-mass spectrometry (GC-MS) to contain 99.5% d₄-RA. The safflower oil source for all trials was high oleate safflower oil, containing 74% monounsaturated and 17% polyunsaturated fat (Pure Pressed Spectrum Naturals Petaluma, CA). The carrot puree was a commercial baby food product (Gerber 2nd Foods Carrots, Fremont, MI), served at room temperature without further treatment. Unit processes involved in the production of the carrot puree included washing and peeling/slicing, pureeing and cooking, filling and capping, and retort processing and cooling. Both the boiled-mashed and the raw-chopped carrots were prepared from the same lot of carrots (controlled source) used to manufacture the commercial puree, intercepted at the processing plant before peeling. Raw, crowned carrots were stored at 2°C, 90% relative humidity under controlled atmosphere refrigeration until meal preparation or analysis. To prepare the boiled-mashed carrots, simulating home preparation of baby food, 5–6 carrots were randomly sampled, washed with water and peeled. The carrots were cut into 5-cm lengths, quartered and further cut lengthwise into pieces between 0.5 and 1.0 cm thick. Approximately one third of the carrot pieces was gently boiled for 40 min in 1 L water, then immediately collected and blotted. The cooked carrot pieces were allowed to cool for 5 min and then passed through a commercial home-use food grinder (Happy Baby Food Grinder, Omron Healthcare, Vernon Hills, IL). The grinder bore a sieve with pores 3 mm in diameter. The resulting mashed carrot preparation was flushed with N₂ and stored overnight in the refrigerator before consumption by subjects or analysis for carotenoid content. To prepare the raw-chopped carrots, carrots were peeled and cut as described above, and then blended in a household food processor (Cuisinart DLC-10E, Cuisinart, Greenwich, CT) for 20 s (two 10-s pulses) at medium speed.

Each test meal consisted of the carrot preparation plus a blended drink containing the d₄-RA and 5 g safflower oil. The amount of carrot fed in each trial was adjusted to provide an equivalent all-trans ßC dose of 18.6 mg, thus controlling for varying water contents of the preparations or for carotene loss during preparation (e.g., during boiling). For the carrot puree, 226 g Gerber 2nd Foods Carrots was fed, providing 18.6 mg all-trans ßC (34.7 µmol), 7.5 mg C (13.9 µmol), 2.0 mg cis₂ ßC (3.7 µmol) and 1.2 mg cis₁ ßC (2.2 µmol). For the boiled-mashed carrot, 175 g carrot was fed, providing 18.6 mg all-trans ßC (34.7 µmol), 7.2 mg C (13.4 µmol) and 1.1 mg cis₂ ßC (2.1 µmol). For the raw carrot, 164 g carrot was fed, providing 18.6 mg all-trans ßC (34.7 µmol) and 6.5 mg C (12.2 µmol). With all test meals, the blended drink was prepared by blending 1 g of the 6 µmol/g d₄-RA stock solution with 70 mL diluted pear juice (filtered through cheese cloth and diluted 1:1 with water), 30.0 g banana and 4 g additional safflower oil. This drink was consumed with the carrot preparation; then the cup and blender were thoroughly rinsed with an additional 100 mL pear juice/water (1:1), which was consumed by the subject to ensure complete ingestion of the components of the test meal.

Subjects were provided with the test meal at 0830 h. Subjects were given 100 mL water 30 min after the test meal and were permitted free access to water after 60 min. Coffee was permitted (without milk) after 1200 h. A standardized, carotenoid-free, low vitamin A lunch was provided 3 h after the test meal. The lunch contained 6 g fat and consisted of one and one-half plain bagel, 23 g cream cheese, 230 g apple juice and choice of fruit (green apple or banana), all purchased locally. In preliminary trials, we found that delay of the second fat dose resulted in a delay in the return to baseline of both carotenoids and unlabeled retinyl ester (d₀-R-ester). Therefore we chose to administer fat at both 0 and 3.25 h to permit release of carotenoids from the intestine within the time frame of the study. A snack consisting of two low fat cookies, providing 1 g fat and 420 kJ, was fed at 1500 h. No other food was permitted during this postdose time period.

Determination of test meal carotenoid content.

Carotenoid contents of the carrot puree, boiled-mashed and raw-chopped carrot treatments were determined by the method of Khachik and Beecher (33 ), with slight modifications. Food samples (6–7 g) with added magnesium carbonate (10% weight of food) were extracted in a Waring blender (Dynamics Corp. of America, New Hartford, CT) with 150 mL tetrahydrofuran (THF) containing 5 mg ethyl ß-apo-8'-carotenoate (Fluka, Milwaukee, WI) as internal standard and 0.01% BHT. The suspension was filtered through no. 1 Whatman filter paper (Whatman International, Maidstone, UK), and the residue reextracted with 100 mL additional THF. The THF extract was concentrated to 80–100 mL on a rotary evaporator at 30°C and then stirred with 150 mL petroleum ether (PE) and 100 mL saturated NaCl solution in a separatory funnel. The hydrophobic, carotenoid-dense PE layer was collected and the water layer removed. The water layer was washed with 2–3 additional aliquots of PE (50–100 mL) until colorless. The PE extracts were combined, dried over magnesium sulfate (200% original weight of food) and filtered through no. 2V Whatman folded filter paper. The PE extract was then sampled, dried and redissolved in mobile phase for analysis of carotenoids by reverse-phase HPLC as described below.

Sample collection and preparation.

Blood samples were drawn by venipuncture into EDTA-containing vacutainers by a trained phlebotomist. Six time points were sampled in each trial: 0 (fasting, baseline), 2.0, 3.5, 4.5, 5.5 and 8.5 h. The test meal, containing the 6 µmol (2 mg) d₄-RA, was administered after collection of the baseline sample. Plasma was separated by low speed centrifugation (500 x g, 5 min) and stored at 2–4°C until ultracentrifugation, which was performed later the same day. The < 1.006 kg/L plasma fraction was obtained by standard ultracentrifugation techniques, as described previously (3 ). Briefly, 4 mL plasma was overlaid with 2 mL 0.195 mol/L NaCl solution ( = 1.006 kg/L) containing 0.27 mmol/L EDTA-Na₂ and 0.001 mol/L NaOH. Tubes were prepared in triplicate for each time point from 4 mL plasma and centrifuged at 180,000 x g_ave for 14 h at 4°C, using a Beckman 50.3 Ti fixed angle rotor (Beckman Instruments, Palo Alto, CA). The < 1.006 kg/L plasma fractions were collected using a tube slicer, flushed with N₂ and stored at -80°C until extraction and analysis.

Total lipids contained in the collected < 1.006 kg/L fraction were extracted and saponified as previously described (34 ) with modifications. Protein was precipitated with 1 volume of ethanol containing 2.5 µg of -tocopherol and 0.1 µg echinenone (as internal standards) and 0.2 g/L BHT; the sample was extracted twice with 3 volumes of hexane. The dried extracts were redissolved in 1.6 mL of 1% ethanolic potassium hydroxide and saponified at 60°C under N₂ for 30 min to convert R-ester to ROH and to remove triacylglycerol. Thus, in the present study, the amount of ROH derived from test meals was assessed from saponification of R-ester in the TRL. Cooled samples were extracted twice with 2 mL hexane after addition of 1.2 mL deionized water. The hexane extract was washed with 2 mL of 100 g/L NaCl followed by a second rinse of 2 mL deionized water. The hexane layer was collected, dried under N₂, and redissolved in 48 µL dimethylformamide plus 250 µL mobile phase (described below) for HPLC analysis.

Analysis of carotenoids and vitamin A in the TRL fraction.

Concentrations of all-trans-ßC, C, retinyl ester-derived ROH [containing both unlabeled (d₀)- and d₄-retinol], -tocopherol, -tocopherol and echinenone in the TRL extract were determined by HPLC using a modification of Thurnham et al. (35 ). A 15-cm Waters Spherisorb ODS-2 column was used with a mobile phase consisting of methanol/acetonitrile/chloroform (47:47:6, v/v/v) and 0.05% triethylamine. Equipment used for HPLC included a Hewlett-Packard Series 1050 Isocratic pump, Waters 996 Photodiode Array Detector and 717 plus Autosampler, Eppendorf CH-30 Column Heater, and Millenium Software Version 2.10 (Waters, Milford, MA). Retention times (RT) of ROH, -tocopherol, echinenone, C and all-trans ßC were 2.8, 5.6, 8.8, 18.1 and 19.6 min, respectively, at a flow rate of 1 mL/min. Analytes were identified by RT relative to authentic standards and UV-visible (UV-vis) spectra, and quantified by reference to standard curves. Six point standard curves were generated for each compound by plotting concentration vs. response and the linear fit of the curve was determined. Equal concentrations of internal standard were used for both standards and samples. "Response" was taken as the height of ROH or carotene divided by the height of the corresponding internal standard. -Tocopherol was used as the internal standard for ROH, and echinenone was used as the internal standard for C, all-trans ßC and cis isomers of ßC. Recoveries of ROH and -tocopherol were determined to be similar through extraction and saponification procedures. Neither internal standard was detected in control TRL fractions. The internal standard used to quantify both - and ß-carotene in the carrot preparations was ethyl ß-apo-8'-carotenoate (RT = 6.1 min). Two cis isomer peaks were designated cis₁ (RT = 20.4 min; max at 448, 348, and 277 nm) and cis₂ (RT = 21.1 min; max at 448, 343 and 281 nm). Because the HPLC method was not optimized for resolution of cis isomers, it is likely that these two peaks represented a mixture of isomers.

The retinyl ester-derived ROH fraction was collected from HPLC, dried under N₂, and silylated with 50 µL of N,O-bis(trimethylsilyl)trifluoroacetimide (TMS) and 100 µL pyridine (Pierce, Rockford, IL) at room temperature in the dark for 1 h. The molar ratio of d₀-retinol-TMS/d₄-retinol-TMS was determined using GC-MS (Hewlett Packard HP 6890 Series Mass Selective Detector & Gas Chromatograph) fitted with a 30-m DB-1 column (i.d., 0.25 mm; film thickness, 0.25 µm, J & W Scientific, Folsom, CA). The GC inlet temperature was 300°C, head pressure 238.5 kPa and split ratio 2.0:1 (split flow 4.8 mL/min). GC oven conditions were as follows: initial temperature 220°C (2-min hold), temperature increased 15°C/min to 260°C (with no hold) and temperature increased 25°C/min to 280°C (hold 10 min). Linear velocity of the helium carrier gas was 68 cm/min. The total run time was 15.57 min. Output was monitored at m/z 358.4 for d₀-retinol-TMS (RT = 5.22 min) and at m/z 362.4 for d₄-retinol-TMS (RT = 5.21 min).

The proportion of total plasma retinyl esters (d₀- and d₄-R-ester) recovered in the upper 1.5 cm of the ultracentrifuge tube (by tube slicing) after centrifugation was determined in a subset of plasma samples collected at various times, i.e., baseline (n = 3), time of maximal TRL-R-ester concentration (n = 9) and time of final blood draw (n = 3). R-esters in the plasma infranatant (lower 4.5 cm) were separated from ROH by normal phase minicolumn chromatography. Briefly, as described earlier (3 ), lipids were extracted from the infranatant, the solvent removed and the residue redissolved in 4 mL hexane/methyl tert-butyl ether (95:5, v/v). This extract was then applied to an Accubond 6-mL solid phase extraction silica cartridge (J & W Scientific), and R-ester eluted with 4 mL additional hexane/methyl tert-butyl ether (95:5, v/v). Retinol, which was retained completely on the column under these conditions, was subsequently eluted with 8 mL of hexane/isopropanol (90:10, v/v). Samples were analyzed by analytical HPLC as described above.

Calculations.

Concentration of total ROH (or retinyl ester-derived retinol) in the TRL, containing both d₀- and d₄-retinol, was determined by HPLC. The molar ratio d₀-retinol-TMS/d₄-retinol-TMS, determined by GC/MS, was multiplied by the concentration of total ROH (from HPLC) to determine the concentrations of labeled and unlabeled retinol in the TRL at each time point. Plots of concentration vs. time were generated, using baseline-corrected values for concentrations of d₀-R-ester, ßC and C. Area-under-curve (AUC) values were then determined by surface area calculations using the experimentally determined coordinates of the curve. To estimate the amount (mass) of ROH or carotene assimilated from the test meal (absolute bioavailability), an absorption efficiency of 80% of the 6 µmol d₄-RA reference dose was assumed, based on published reports of preformed vitamin A absorption obtained from a variety of experimental approaches (36 –39 ). The mass of absorbed carotenes or carotene-derived d₀-retinol was calculated by dividing the AUC of d₀-R-ester or carotenoid by that of d₄-R-ester, and multiplying this quotient by 4.8 µmol.

Retinol equivalencies (RE) of ßC in the carrot meals were calculated by the following formulas, which account for the ratio of ßC:C in carrot (2.5:1 puree, 2.6:1 boiled mashed, 2.9:1 raw), and assuming a ROH biopotency of C as one half that of ßC. Because the ratios of ßC:C for the two cooked carrot preparations were very similar, calculations used a single value (2.6:1). For cooked carrot, RE was calculated as follows: (18.6 mg ßC from dose)/[(mass d₀-ROH_TRL) - (0.5 x 1/3.6 · mass d₀-ROH_TRL)], where the ratio of 1/3.6 represents C/C + ßC. For raw carrot, RE was calculated as follows: (18.6 mg ßC from dose)/[(mass d₀-ROH_TRL) - (0.5 x 1/3.9 · mass d₀-ROH_TRL)]. The latter parenthetical expression represents the predicted contribution of raw carrot C to the mass of absorbed ROH. We also estimated the amount of total carotenes absorbed, either as intact ßC or C or as d₀-R-ester (derived from intestinal cleavage of carrot carotenes). To do this, we initially ignored the contribution of C to TRL R-ester. Assuming a 1:2 stoichiometry of conversion of ßC to ROH, the value of total carotenes absorbed was calculated as follows (mol): [(ßC_TRL + C_TRL + 0.5 · R-ester_TRL · 100)/(ßC_food + C_food)]. To estimate the potential contribution of C to R-ester, the ratio of ßC to C in the carrot and a 1:1 equivalency of C to R-ester were factored into the equation. For example, the percentage of total carrot carotenes assimilated from cooked carrot was recalculated as follows: [(ßC_TRL + C_TRL + 0.5 x 2.6/3.6 · R-ester_TRL + 1 x 1/3.6 · R-ester_TRL · 100)/(ßC_food + C_food)].

The apparent conversion efficiency and ßC:ROH equivalency ratio were also calculated. To make these calculations, the estimated amount of C-derived ROH was subtracted from the total amount of ROH absorbed (as R-ester) to yield a value for ßC-derived ROH. C-Derived ROH was calculated by assuming that the ROH yield from C is one half that of ßC and accounting for the ratio of ßC to C in the carrot meal. Thus ßC-derived ROH is equal to [(mmol d₀-R-ester_TRL) - 0.5 x 1/3.6 · mmol d₀-R-ester_TRL)]. Apparent conversion efficiency, or percentage of ßC taken up that was metabolized to ROH, was calculated as follows (mmol): [(ßC-derived ROH_TRL)/(ßC_TRL as intact ßC + ßC-derived ROH_TRL)] · 100. TRL concentrations for each analyte (d₀-R-ester, d₄-R-ester, ßC, C) at 8.5 and at 2 h were used to determine an 8.5 h/2 h ratio within each subject for each treatment day.

Statistics.

Paired t tests were used to compare the masses of ßC, C, total carotenes (ßC plus C) and d₀-R-ester resulting from the two cooked carrot treatments. The mass of total carotenes absorbed either as intact carotene (ßC + C) or as d₀-R-ester, the ratio of mass of d₀-ROH absorbed to mass of ßC absorbed, the percentage of total carotenes absorbed as a function of carotene dose and d₄-R-ester AUC values were also analyzed by paired t test. All analyses were performed using SPSS for Windows (version 8.0; SPSS, Chicago). A significance level of P < 0.05 and 8 df were applied for each analysis. Using the subset of data from 6 subjects who were given all three carrot treatments, a repeated-measures ANOVA was performed. Data examined were mass and percentage of carotenes absorbed, d₀-ROH absorbed, d₄-R-ester AUC and 8.5 h/2 h ratios for the three treatments. For these analyses, a significance level of P < 0.05 and 5 df were applied.

	RESULTS

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
Carotene content of test meals.

Analysis of the two cooked carrot preparations yielded mean all-trans-ßC contents (on a wet-weight basis) of 8.25 ± 0.09 and 10.65 ± 0.12 mg/100 g for carrot puree and boiled-mashed carrot, respectively. For the raw carrot, mean all-trans-ßC content (on a wet-weight basis) was 11.45 ± 1.86 mg/100 g. The ratios of ßC:C were similar for the cooked carrot preparations (puree, 2.5:1, boiled-mashed, 2.6:1) and slightly higher for the raw carrot (2.9:1). Two cis isomers of ßC were detected by their UV-vis spectra, i.e., cis₁ at 20.4 min (detected in carrot puree) and cis₂ at 21.1 min (detected in TRL fraction, carrot puree and mashed carrot). One isomer of C (RT = 18.8 min) was detected in the carrot puree, but was not detected in TRL fractions. No cis isomers were detected in the raw-chopped carrot.

Response profiles for d₄-R-ester, d₀-R-ester, ßC and C for the cooked carrot preparations.

Baseline-adjusted, postprandial concentrations of d₄-R-ester, d₀-R-ester, ßC and C in the < 1.006 kg/L (TRL) plasma fraction for a typical subject fed the two cooked carrot treatments are shown in Figure 1 . d₄-R-ester AUC did not differ significantly between the two carrot treatments. In 12 of 18 trials, d₄-R-ester and d₀-R-ester peaked simultaneously; in the other six trials, d₄-R-ester peak response was detected slightly earlier than d₀-R-ester. In 17 of 18 trials, concentrations of all four analytes peaked within 3.5–5.5 h postdose and returned to near baseline within 8.5 h. In 15 of 18 trials, the ßC peak time coincided with the C peak time.

View larger version (28K): [in this window] [in a new window]   Figure 1. Baseline-corrected, postprandial concentrations of labeled (d4) and unlabeled (d0) retinyl esters (R-ester), ß-carotene (ßC) and -carotene (C) in the triacylglycerol-rich lipoprotein fraction ( < 1.006 kg/L) of a female volunteer fed carrot puree (A) and boiled-mashed carrot (B). Both carrot test meals contained 2 mg d4-retinyl acetate, 18.6 mg ßC, 7.4 mg C and 5 g fat and were consumed in separate, single-dose trials conducted 2–6 wk apart. Values are means ± SD of triplicate determinations at each time point.

Estimated masses of d₀-retinol and carotenes derived from test meals contained carrot puree or boiled-mashed carrots.

The estimated masses of ßC and of C absorbed intact, calculated assuming 80% absorption efficiency of the d₄-RA reference, were significantly greater for the carrot puree treatment (0.44 ± 0.15 mg ßC, 0.26 ± 0.07 mg C) than for the mashed carrot treatment (0.16 mg ± 0.11 mg ßC, 0.09 ± 0.04 mg C) (Fig. 2 ). The amount of ROH derived per 18.6 mg ßC dose tended to be greater for carrot puree (0.53 ± 0.2 mg ROH) than for boiled-mashed carrot (0.44 ± 0.17 mg ROH) (P = 0.11). In 6 of 9 subjects, the mass of absorbed R-ester was greater from the puree than from the mashed treatment, whereas for 2 subjects, mass of absorbed R-ester was greater with the boiled-mashed treatment, and for 1 subject there was virtually no difference between treatments.

View larger version (18K): [in this window] [in a new window]   Figure 2. Estimated masses of unlabeled retinol (d0-ROH), ß-carotene (ßC) and -carotene (C) absorbed from test meals of commercial carrot puree or boiled-mashed carrots, calculated by comparison of the area-under-curve (AUC) values of ßC, C, and unlabeled retinyl ester (d0-R-ester) with the corresponding AUC value of the deuterium-labeled retinyl ester (d4-R-ester) reference (4.8 µmol). AUC values were obtained by plotting concentration (nmol/L) vs. time curves for 4 analytes in each trial, and determining the surface area of each curve. Values are mean ± SD, n = 9 for the two treatments. Nine healthy human subjects consumed both treatments on separate trial days, in random order.

The sum of combined masses of ßC and C was significantly greater for the carrot puree than for the carrot mash (Fig. 3 ). Similarly, the sum of combined masses of ßC, C and retinyl esters also differed (P < 0.01) between the two treatments (Fig. 3) .

View larger version (15K): [in this window] [in a new window]   Figure 3. Estimated masses of total carotenes (sum of intact carotenes), and total carotenes plus retinyl-ester derived retinol (d0-ROH) absorbed from test meals of commercial carrot puree or boiled-mashed carrots, calculated by comparison of the (area-under-curve) AUC values of ß-carotene (ßC), -carotene (C) and unlabeled retinyl ester (d0-R-ester) with the corresponding AUC value of the deuterium-labeled retinyl ester (d4-R-ester) reference (4.8 µmol) obtained from each trial. AUC values were obtained by plotting concentration (nmol/L) vs. time curves for carotenes and retinol (d4 and d0) for each trial and determining the surface area of each curve. Values are mean ± SD, n = 9 for the two treatments. Nine healthy human subjects consumed both treatments on separate trial days, in randomized order.

View larger version (33K): [in this window] [in a new window]   Figure 4. Mole percentage of individual carotenes and of total carotenes [including carotenes converted to retinyl ester (R-ester)] absorbed from test meals of commercial carrot puree or boiled-mashed carrots, relative to total carotenes consumed. Values for the percentage of total carotenes absorbed were calculated as the sum of intact carotenes plus carotene-derived unlabeled retinyl ester (d0-R-ester), expressed relative to total carotenes [-carotene (C) plus ß-carotene (ßC)] consumed. The relative contributions of and ß-carotene to d0-R-ester was estimated by accounting for the ratio of ßC:C in the carrot (2.6:1 cooked, 2.9:1 raw) and by assuming a stoichiometry of conversion to retinol of 1:2 for ßC and 1:1 for C. Values are mean ± SD, n = 9 for the two treatments. Nine healthy human subjects consumed both treatments on separate trial days, in randomized order.

View larger version (38K): [in this window] [in a new window]   Figure 5. Estimated masses of ß-carotene (ßC) absorbed intact by each subject consuming 18.6 mg ßC as commercial carrot puree (n = 9), boiled-mashed carrots (n = 9) or raw-grated carrot (n = 6). Each subject is represented by a unique column pattern. The mass of absorbed ßC was calculated by comparing triacylglycerol-rich lipoprotein (TRL) ßC area-under-curve (AUC) values against the corresponding AUC for the deuterium-labeled retinyl acetate (d4-RA) reference (4.8 µmol) for each subject and treatment.

For the six subjects receiving all three carrot treatments, the concentrations of ßC and R-ester at 8.5 h were expressed relative to those at 2 h as evidence for a treatment effect on the time course of absorption. The 8.5 h/2 h ratios for ßC were significantly greater for the raw carrot treatment than for the puree treatment (Tukey P = 0.03, Fig. 6A ), suggesting a delay in absorption of ß-carotene from the raw-chopped carrot meal. A similar trend (P = 0.16) was observed for d₀-R-ester (Fig. 6 B).

View larger version (21K): [in this window] [in a new window]   Figure 6. Ratios of the concentrations (nmol/L) of ß-carotene (ßC) (Panel A) or unlabeled retinyl ester (d0-R-ester) (Panel B) in the triacylglycerol-rich lipoprotein (TRL) fraction at 2 h relative to that at 8.5 h, for six subjects receiving all three carrot test meals (commercial carrot puree, boiled-mashed carrot and raw-grated carrot). Each subject is represented by a different symbol.

For the raw carrot treatment (n = 6), the calculated masses of d₀-ROH, ßC and C secreted into the TRL fraction were 0.43 ± 0.27, 0.38 ± 0.28 and 0.19 ± 0.14 mg, respectively. The percentages of ßC and C consumed that were absorbed intact were 1.7 ± 1.3 and 2.4 ± 1.8%, respectively. The molar ratio of ßC:C in the TRL fraction was 2.2 ± 0.6, lower than that of the carrot in the test meal, as was observed with the two cooked carrot treatments. None of these values, based on data from 6 subjects, were significantly different from the corresponding values for either of the two cooked carrot treatments.

Apparent efficiency of conversion of ßC to ROH.

The proportion of ß-carotene taken up by the intestine that was metabolized to ROH was estimated as described above. The apparent ßC conversion efficiency was 44 ± 11% (range: 24–58%) for the carrot puree treatment and 59 ± 12% (range: 36–72%) for boiled-mashed carrots. For the two cooked carrot treatments, this difference in apparent conversion efficiency of ßC to vitamin A was significant. For the raw-chopped carrot treatment, conversion efficiency was 63 ± 10% and was not different from either of the cooked carrot treatments (n = 6), although the power to detect this difference was reduced with use of fewer subjects (n = 6) than in the cooked carrot treatments (n = 9).

	DISCUSSION

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
The TRL/d₄-RA reference approach, which employs coadministration of oil-solubilized, deuterium-labeled preformed vitamin A ester as a reference dose, was developed to permit estimation of the mass of unlabeled vitamin A (d₀-R-ester) derived from single meals containing provitamin A carotenoids. This approach was used to compare the absolute (mass of ROH and carotenes absorbed) and relative bioavailabilities of ßC and C from a commercial carrot baby food puree, boiled-mashed carrot and raw-grated carrot, all prepared from a single source of carrots. Postprandial TRL concentrations of d₀-R-esters and individual carotenes were normalized to the AUC for d₄-RA, thus controlling for intra- and interindividual variations in CM clearance rates. Esterified ROH, which does not readily exchange between lipoproteins and is not resecreted from liver, has been used for >20 y as a marker for these intestinally derived particles (40 –45 ). With this approach, the labeled vitamin A reference dose will ideally mix with the vitamin A and carotenes from the test commodity into a commonly absorbed pool, and will be absorbed with similar efficiency across a variety of meal compositions.

In this study, similarities in absorption patterns for d₀-R-ester, d₄-R-ester, ßC and C in the TRL fraction were evident. In particular, both unlabeled and labeled vitamin A peaked simultaneously in most trials, supporting the assumption that vitamin A metabolically derived from the test meal carotenes and that from the reference dose comixed into a common pool during CM secretion and clearance. Comparison of d₄-R-ester AUC revealed no significant differences between the two cooked carrot treatments or between the cooked and raw treatments. We previously observed similar d₄-RA responses under differing test meal fat and fiber content conditions (3 ). Previous studies have also shown that retinyl acetate is generally well absorbed and that efficiency of absorption of retinyl ester administered in oil solution is similar in healthy individuals (36 ,37 ,46 ).

Using this approach, we observed that the mass of absorbed ßC and C was approximately twofold greater from the commercial carrot puree than from boiled-mashed carrots. This finding is consistent with Rock et al. (24 ) who reported that the increase in serum ßC from chronic consumption of cooked and homogenized carrots was three times greater than the increase in serum ßC from raw carrots. The greater carotene bioavailability of the commercial carrot puree relative to the boiled-mashed carrots may have resulted from reduced particle size (increased surface area), greater heat exposure or a combination of both factors (26 ). Results of several studies support the concept that disruption of the food matrix by heat (47 ), homogenization (27 ,48 ) or both factors (24 ) improves the bioavailability of ßC from carrots, as reflected by serum ßC concentration change. Recently, van het Hof et al. (28 ) reported an interactive effect of heat and homogenization on ßC uptake from canned tomatoes. It has been suggested that mild heat treatment may improve ßC bioavailability from plant foods by weakening carotenoid-protein complexes (49 ).

Heat treatment of carrots promotes solubilization of cell wall pectins and subsequent softening of the tissue (50 ), increases the ratio of soluble to insoluble fiber (51 ) and affects the microstructure of cell walls (52 ,53 ). Soluble fiber has been proposed to alter gastric conditions to inhibit (54 ) or enhance (55 ) lipid absorption. However, although ßC and C from isolated carrot chloroplasts were more bioavailable to ferrets than from unfractionated carrot juice, heating of the chromoplasts or juice had no effect on their bioavailability in that model (23 ). Because soluble fiber (pectin) can interfere with carotenoid absorption (20 ,21 ), predicting the net effect of heat-induced fiber changes on carotenoid absorption is difficult.

Heating may also promote solubilization of carotene complexes, particularly from finely divided foods with a large surface area such as the carrot baby food puree studied here. This product received the majority of its heat exposure after sealing in its glass container, and any solubilized carotenes would have been retained and consumed with the test meal. In contrast, some carotene was clearly lost during preparation of the boiled-mashed carrots, as indicated by coloration of the cooking water. Although the test meals were constructed to deliver identical amounts of ßC, it is possible that a particularly bioavailable subfraction of carotene was lost during boiling. Heating also results in isomerization of carotenoids. The presence of 9-cis ßC increased total uptake of carotenes into micelles in vitro (56 ). The extent to which the elevated cis-ßC content of the commercial carrot puree contributed to the greater carotene bioavailability of this preparation is not known.

Although the commercial puree and boiled-mashed carrot preparations exhibited markedly different carotene bioavailabilities, the mass of assimilated vitamin A was only marginally greater for the puree than for boiled-mashed carrots, and the treatment difference was not significant with nine subjects (P = 0.11). The difference in apparent ßC-to-ROH intestinal conversion efficiencies (44 and 59% in the puree and boiled-mashed treatments, respectively) implies a degree of regulation in the production of ROH by the intestine in healthy individuals and deserves additional study. We earlier reported evidence for saturation of intestinal conversion at 50-mg ßC doses (57 ). Similar estimates of conversion efficiency in well-nourished subjects have been reported by others (29 ,38 ,39 ).

We anticipated that the carotene and vitamin A values of the raw-grated carrot preparation would be significantly lower than those of the two cooked preparations, but this was not the case. This finding, however, agrees with a previous study (58 ) in which no difference in serum ßC change was noted between groups of subjects fed either carrot juice or raw grated carrots. We speculate that longer gastrointestinal residence time may have compensated for the lack of heat treatment and relatively large particle size of this preparation because it has been suggested that the particle size of ingested foods may affect gastric emptying rate (59 ,60 ) and subsequent glucose response (61 ,62 ). Prolonged transit time would be expected to result in a shift in the TRL carotene absorption profile to later times, which we in fact observed. The raw-grated carrot trials were accompanied by significantly greater 8.5 h/2.5 h TRL ßC concentration ratios relative to either of the cooked carrot trials. The fact that the d₄-R-ester profile was unaffected by treatment supports the notion that the delay in carotene and d₀-R-ester absorption was related to the carrot preparation per se.

The ßC:C ratio in the TRL fraction (mean of 1.7) was consistently and considerably lower than the ratio present in the ingested cooked carrot preparations (mean of 2.6). This could have resulted from either preferential conversion of ßC (vs. C) to ROH or from less efficient intestinal uptake of ßC than of C. Whether differential molecular localization or arrangement in carrot tissue could result in more efficient release of C is not known. Other studies have observed greater increase in plasma -carotene than ß-carotene in subjects fed carrots (13 ) or carrot juice (63 ).

Subjects consumed 26 mg provitamin A carotenes, consisting of 18.6 mg ßC and 7.4 mg C, with each test meal. The estimated ROH yields were nearly 23–28% lower than the 1.9 mg ROH predicted by the RAE of 12:1 for ßC and 24:1 for C recently set by the Micronutrient Panel (4 ). This general finding was consistent with that of an earlier study in our laboratory, using the same approach but with smaller doses of raw carrots and spinach (3 ). Although the carrot values obtained here are within the range of those calculated by comparing change in serum ROH levels in malnourished children fed provitamin A vegetables with those fed retinol-rich foods (8 ), certain issues should be noted. First, vitamin A values obtained with the TRL/d₄-RA model reflect only intestinal production of vitamin A from carotene, and may underestimate that from the total body if substantial postabsorptive conversion of carotene to vitamin A does in fact occur. Second, our carotene doses were relatively high, and the dose-response relationship of ROH production from food-borne carotenes is unknown. The efficiency of ß-carotene absorption decreases at greater intakes (36 ,64 ). Reduced apparent absorption efficiency of ßC has been reported to occur with an increase in dose from 12 to 30 mg, based on a decrease in peak plasma ßC response (12 ). Thus it is possible that the RAE ratio of smaller vegetable portions is lower (greater ROH yield relative to carotene consumed) than that from the relatively large portions used in this study. Additionally, RAE ratios obtained in well-nourished subjects may not be equivalent to those typical of vitamin A–deficient populations. Test meal fat content was unlikely to have limited carotene bioavailability in the current study, as suggested by recent data concerning the influence of reduced meal fat content on carotenoid bioavailability (3 ,65 ).

This report describes the application of an extrinsic stable isotope reference method to compare the mass of carotenoids and vitamin A derived from test meals containing carrots processed in different ways. The TRL/d₄-RA model, as illustrated here, can be a useful quantitative tool in the investigation of the determinants of bioavailability of provitamin A carotenoids from specific foods consumed under virtually any meal condition. Markedly greater absorption of ßC and C was obtained from a commercially processed carrot puree, a product subjected to substantial heat treatment and particle size reduction, relative to a boiled-mashed preparation derived from the same lot of carrots. At the same time, the mass of vitamin A derived from the carrot puree was only marginally, and nonsignificantly greater than that of the boiled-mashed carrot preparation, suggesting regulation in the intestinal conversion of carotene to vitamin A.

Although the type of processing of the two heated carrot preparations clearly affected their carotene bioavailability, the subjects also assimilated large amounts of carotenes and vitamin A from raw-grated carrots, possibly due to slower gastrointestinal transit and prolonged particle digestion. The amount of ROH metabolically derived from the cooked carrot preparations was approximately one third that predicted by current RAE ratios. Because the estimated efficiency of intestinal conversion of ßC to ROH was 44–59%, low intestinal uptake of the relatively large dose of carrot carotenes, rather than poor efficiency of conversion of ßC to ROH, appeared responsible for the lower than predicted vitamin A values obtained under these conditions.

	FOOTNOTES

 
¹ Supported by Gerber Products Company, Fremont, MI 49413 and by the Division of Nutritional Sciences and Department of Food Science (assistantships to A.J.E.), Cornell University, Ithaca, NY.

³ Abbreviations used: C, -carotene; AUC, area-under-curve; ßC, ß-carotene; CM, chylomicron; d₀-R-ester, unlabeled retinyl ester; d₄-RA, deuterium-labeled retinyl acetate; d₄-R-ester, deuterium-labeled retinyl ester; PE, petroleum ether; RAE, retinol activity equivalency; R-ester, retinyl ester; ROH, retinol; RT, retention time; THF, tetrahydrofuran; TMS, N,O-bis(trimethylsilyl)trifluoroacetamide; TRL, triacylglycerol-rich lipoprotein; UV-vis, UV-visible.

Manuscript received 21 August 2001. Initial review completed 20 September 2001. Revision accepted 30 October 2001.

	LITERATURE CITED

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 

1. Underwood, B. A. (1984) Vitamin A in animal and human nutrition. Sporn, M. B. Roberts, A. B. Goodman, D. S. eds. Retinoids 1:281-392 Academic Press Orlando, FL. .

2. Parker, R. S., Swanson, J. E., You, C. S., Edwards, A. J. & Huang, T. (1999) Bioavailability of carotenoids in human subjects. Proc. Nutr. Soc. 58:155-162.[Medline]

3. Edwards, A. J., You, C. S., Swanson, J. E. & Parker, R. S. (2001) A novel extrinsic reference method for assessing the vitamin A value of plant foods. Am. J. Clin. Nutr. 74:348-355.[Abstract/Free Full Text]

4. Panel on Micronutrients. Subcommittees on Upper Reference Levels of Nutrients and of Interpretation and Use of Dietary Reference Intakes, Standing Committee on the Scientific Evaluation of Dietary Reference Intakes, Food and Nutrition Board, Institute of Medicine (2001) Dietary Reference Intakes for Vitamin A, Vitamin K, Arsenic, Boron, Chromium, Copper, Iodine, Iron, Manganese, Molybdenum, Nickel, Silicon, Vanadium, and Zinc 2001 National Academy Press Washington, DC. .

5. National Research Council (1989) Recommended Dietary Allowances 1989 National Academy Press Washington, DC. .

6. de Pee, S., West, C. E., Muhilal, , Karyadi, D. & Hautvast, J. G. (1995) Lack of improvement in vitamin A status with increased consumption of dark-green leafy vegetables. Lancet 346:75-81.[Medline]

7. de Pee, S. & West, C. E. (1996) Dietary carotenoids and their role in combating vitamin A deficiency: a review of the literature. Eur. J. Clin. Nutr. 50(suppl. 3):S38-S53.

8. de Pee, S., West, C. E., Permaesih, D., Martuti, S., Muhilal, & Hautvast, J. G. (1998) Orange fruit is more effective than are dark-green, leafy vegetables in increasing serum concentrations of retinol and ß-carotene in schoolchildren in Indonesia. Am. J. Clin. Nutr. 68:1058-1067.[Abstract]

9. Roels, O. A., Trout, M. & Dujacquier, R. (1958) Carotene balances in boys in Ruanda where vitamin A deficiency is prevalent. J. Nutr. 65:115-127.

10. Rao, C. N. & Rao, B. S. (1970) Absorption of dietary carotenes in human subjects. Am. J. Clin. Nutr. 23:105-109.[Abstract]

11. Jensen, C. D., Pattison, T. S., Spiller, G. A., Whittam, J. H. & Scala, J. (1985) Repletion and depletion of serum and ß carotene in humans with carrots and an algae-derived supplement. Acta Vitaminol. Enzymol. 7:189-198.[Medline]

12. Brown, E. D., Micozzi, M. S., Craft, N. E., Bieri, J. G., Beecher, G., Edwards, B. K., Rose, A., Taylor, P. R. & Smith, J. C., Jr (1989) Plasma carotenoids in normal men after a single ingestion of vegetables or purified ß-carotene. Am. J. Clin. Nutr. 49:1258-1265.[Abstract/Free Full Text]

13. Micozzi, M. S., Brown, E. D., Edwards, B. K., Bieri, J. G., Taylor, P. R., Khachik, F., Beecher, G. R. & Smith, J. C., Jr (1992) Plasma carotenoid response to chronic intake of selected foods and ß-carotene supplements in men. Am. J. Clin. Nutr. 55:1120-1125.[Abstract/Free Full Text]

14. van den Berg, H. & van Vliet, T. (1998) Effect of simultaneous, single oral doses of ß-carotene with lutein or lycopene on the ß-carotene and retinyl ester responses in the triacylglycerol-rich lipoprotein fraction of men. Am. J. Clin. Nutr. 68:82-89.[Abstract]

15. Jayaran, P., Reddy, V. & Mohanram, M. (1980) Effect of dietary fat on absorption of ß-carotene from green leafy vegetables in children. Indian J. Med. Res. 71:53-56.[Medline]

16. Dimitrov, N. V., Meyer, C., Ullrey, D. E., Chenoweth, W., Michelakis, A., Malone, W., Boone, C. & Fink, G. (1988) Bioavailability of ß-carotene in humans. Am. J. Clin. Nutr. 48:298-304.[Abstract/Free Full Text]

17. Jalal, F., Nesheim, M. C., Agus, Z., Sanjur, D. & Habicht, J. P. (1998) Serum retinol concentrations in children are affected by food sources of ß-carotene, fat intake, and anthelmintic drug treatment. Am. J. Clin. Nutr. 68:623-629.[Abstract]

18. Takyi, E.E.K. (1999) Children’s consumption of dark green, leafy vegetables with added fat enhances serum retinol. J. Nutr. 129:1549-1554.[Abstract/Free Full Text]

19. Erdman, J. W., Jr, Fahey, G. C., Jr & White, C. B. (1986) Effects of purified dietary fiber sources on ß-carotene utilization by the chick. J. Nutr. 116:2415-2423.

20. Rock, C. L. & Swendseid, M. E. (1992) Plasma ß-carotene response in humans after meals supplemented with dietary pectin. Am. J. Clin. Nutr. 55:96-99.[Abstract/Free Full Text]

21. Riedl, J., Linseisen, J., Hoffmann, J. & Wolfram, G. (1999) Some dietary fibers reduce the absorption of carotenoids in women. J. Nutr. 129:2170-2176.[Abstract/Free Full Text]

22. Deming, D. M., Boileau, A. C., Lee, C. M. & Erdman, J. W., Jr (2000) Amount of dietary fat and type of soluble fiber independently modulate postabsorptive conversion of ß-carotene to vitamin A in Mongolian gerbils. J. Nutr. 130:2789-2796.[Abstract/Free Full Text]

23. Zhou, J. R., Gugger, E. T. & Erdman, J. W., Jr (1996) The crystalline form of carotenes and the food matrix in carrot root decrease the relative bioavailability of beta- and alpha-carotene in the ferret model. J. Am. Coll. Nutr. 15:84-91.[Abstract]

24. Rock, C. L., Lovalvo, J. L., Emenhiser, C., Ruffin, M. T., Flatt, S. W. & Schwartz, S. J. (1998) Bioavailability of ß-carotene is lower in raw than in processed carrots and spinach in women. J. Nutr. 128:913-916.[Abstract/Free Full Text]

25. Castenmiller, J. J., West, C. E., Linssen, J. P., van het Hof, K. H. & Voragen, A. G. (1999) The food matrix of spinach is a limiting factor in determining the bioavailability of ß-carotene and to a lesser extent of lutein in humans. J. Nutr. 129:349-355.[Abstract/Free Full Text]

26. Parker, R. S. (1996) Absorption, metabolism, and transport of carotenoids. FASEB J 10:542-551.[Abstract]

27. van Zeben, W. & Hendriks, T. F. (1948) The absorption of carotene from cooked carrots. Z. Vitam. Forsch. 19:265-266.

28. van het Hof, K. H., de Boer, B. C., Tijburg, L. B., Lucius, B. R., Zijp, I., West, C. E., Hautvast, J. G. & Weststrate, J. A. (2000) Carotenoid bioavailability in humans from tomatoes processed in different ways determined from the carotenoid response in the triglyceride-rich lipoprotein fraction of plasma after a single consumption and in plasma after four days of consumption. J. Nutr. 130:1189-1196.[Abstract/Free Full Text]

29. van Vliet, T., Schreurs, W. H. & van den Berg, H. (1995) Intestinal ß-carotene absorption and cleavage in men: response of ß-carotene and retinyl esters in the triglyceride-rich lipoprotein fraction after a single oral dose of ß-carotene. Am. J. Clin. Nutr. 62:110-116.[Abstract/Free Full Text]

30. Wingerath, T., Stahl, W. & Sies, H. (1995) ß-Cryptoxanthin selectively increases in human chylomicrons upon ingestion of tangerine concentrate rich in ß-cryptoxanthin esters. Arch. Biochem. Biophys. 324:385-390.[Medline]

31. Gartner, C., Stahl, W. & Sies, H. (1997) Lycopene is more bioavailable from tomato paste than from fresh tomatoes. Am. J. Clin. Nutr. 66:116-122.[Abstract/Free Full Text]

32. Johnson, E. J. & Russell, R. M. (1992) Distribution of orally administered ß-carotene among lipoproteins in healthy men. Am. J. Clin. Nutr. 56:128-135.[Abstract/Free Full Text]

33. Khachik, F. & Beecher, G. R. (1987) Application of a C-⁴⁵-ß-carotene as an internal standard for the quantification of carotenoids in yellow/orange vegetables by liquid chromatography. J. Agric. Food Chem. 35:732-738.

34. You, C. S., Parker, R. S., Goodman, K. J., Swanson, J. E. & Corso, T. N. (1996) Evidence of cis-trans isomerization of 9-cis-ß-carotene during absorption in humans. Am. J. Clin. Nutr. 64:177-183.[Abstract]

35. Thurnham, D. I., Smith, E. & Flora, P. S. (1988) Concurrent liquid-chromatographic assay of retinol, -tocopherol, ß-carotene, -carotene, lycopene, and ß-cryptoxanthin in plasma, with tocopherol acetate as internal standard. Clin. Chem. 34:377-381.[Abstract/Free Full Text]

36. Olson, J. A. (1987) Recommended dietary intakes (RDI) of vitamin A in humans. Am. J. Clin. Nutr. 45:704-716.[Abstract/Free Full Text]

37. Sivakumar, B. & Reddy, V. (1972) Absorption of labelled vitamin A in children during infection. Br. J. Nutr. 27:299-304.[Medline]

38. Goodman, D. S., Blomstrand, R., Werner, B., Huang, H. S. & Shiratori, T. (1966) The intestinal absorption and metabolism of vitamin A and ß-carotene in man. J. Clin. Investig. 45:1615-1623.

39. Blomstrand, R. & Werner, B. (1967) Studies on the intestinal absorption of radioactive ß-carotene and vitamin A in man. Conversion of ß-carotene into vitamin A. Scand. J. Clin. Lab. Investig. 19:339-345.[Medline]

40. Hazzard, W. R. & Bierman, E. L. (1976) Delayed clearance of chylomicron remnants following vitamin-A-containing oral fat loads in broad-beta disease (type III hyperlipoproteinemia). Metabolism 25:777-801.[Medline]

41. Ross, A. C. & Zilversmit, D. B. (1977) Use of esterified retinol to trace the degradation of chylomicrons in cholesterol-fed rabbits. Adv. Exp. Med. Biol. 82:142-145.[Medline]

42. Berr, F., Eckel, R. & Kern, F., Jr (1985) Plasma decay of chylomicron remnants is not affected by heparin-stimulated plasma lipolytic activity in normal fasting man. J. Lipid Res. 26:852-859.[Abstract]

43. Berr, F. (1992) Characterization of chylomicron remnant clearance by retinyl palmitate label in normal humans. J. Lipid Res. 33:915-930.[Abstract]

44. Hussain, M. M., Mahley, R. W., Boyles, J. K., Fainaru, M., Brecht, W. J. & Lindquist, P. A. (1989) Chylomicron-chylomicron remnant clearance by liver and bone marrow in rabbits. Factors that modify tissue-specific uptake. J. Biol. Chem. 264:9571-9582.

45. Goodman, D. S. & Blaner, W. S. (1984) Biosynthesis, absorption and hepatic metabolism of retinol. Sporn, M. B. Roberts, A. B. Goodman, D. S. eds. Retinoids 2:2-39 Academic Press Orlando, FL. .

46. Borel, P., Pasquier, B., Armand, M., Tyssandier, V., Grolier, P., Alexandre-Gouabau, M. C., Andre, M., Senft, M., Peyrot, J., Jaussan, V., Lairon, D. & Azais-Braesco, V. (2001) Processing of vitamin A and E in the human gastrointestinal tract. Am. J. Physiol. 280:G95-G103.[Abstract/Free Full Text]

47. Poor, C. L., Bierer, T. L., Merchen, N. R., Fahey, G. C., Jr & Erdman, J. W., Jr (1993) The accumulation of - and ß-carotene in serum and tissues of preruminant calves fed raw and steamed carrot slurries. J. Nutr. 123:1296-1304.

48. Hussein, L. & El-Tohamy, M. (1990) Vitamin A potency of carrot and spinach carotenes in human metabolic studies. Int. J. Vitam. Nutr. Res. 60:229-235.[Medline]

49. Dietz, J. M., Sri, K. S. & Erdman, J. W., Jr (1988) Reversed phase HPLC analysis of - and ß-carotene from selected raw and cooked vegetables. Plant Foods Hum. Nutr. 38:333-341.[Medline]

50. Greve, L. C., McArdle, R. N., Gohicke, J. R. & Labavitch, J. M. (1994) Impact of heating on carrot firmness: changes in cell wall components. J. Agric. Food Chem. 42:2900-2906.

51. Penner, M. H. & Kim, S. (1991) Nonstarch polysaccharide fractions of raw, processed, and cooked carrots. J. Food Sci. 56:1593-1596.

52. Préstamo, G., Fuster, C. & Risueño, M. C. (1998) Effects of blanching and freezing on the structure of carrots cells and their implication for food processing. J. Sci. Food Agric. 77:223-229.

53. Kidmose, U. & Martens, H. J. (1999) Changes in texture, microstructure and nutritional quality of carrot slices during blanching and freezing. J. Sci. Food Agric. 79:1747-1753.

54. Pasquier, B., Armand, M., Castelain, C., Guillon, G., Borel, P., Lafont, H. & Lairon, D. (1996) Emulsification and lipolysis of triacylglycerols are altered by viscous soluble dietary fibres in acidic gastric medium in vitro. Biochem. J. 314:269-275.

55. Fillery-Travis, A. J., Gee, J. M., Waldron, K. W., Robins, M. M. & Johnson, I. T. (1997) Soluble nonstarch polysaccharides derived from complex food matrices do not increase average lipid droplet size during gastric lipid emulsification in rats. J. Nutr. 127:2246-2252.[Abstract/Free Full Text]

56. Levin, G. & Mokady, S. (1995) Incorporation of all-trans- or 9-cis-ß-carotene into mixed micelles in vitro. Lipids 30:177-179.[Medline]

57. Parker, R. S., Brenna, J. T., Swanson, J. E., Goodman, K. J. & Marmor, B. (1997) Assessing metabolism of ß-[¹³C]carotene using high-precision isotope ratio mass spectrometry. Methods Enzymol 282:130-140.[Medline]

58. Törrönen, R., Lehmusaho, M., Häkkinen, S., Hänninen, O. & Mykkänen, H. (1996) Serum ß-carotene response to supplementation with raw carrots, carrot juice or purified ß-carotene in healthy non-smoking women. Nutr. Res. 16:565-575.

59. Meyer, J. H., Ohashi, H., Jehn, D. & Thomson, J. B. (1981) Size of liver particles emptied from the human stomach. Gastroenterology 80:1489-1496.[Medline]

60. Holt, S., Reid, J., Taylor, T. V., Tothill, P. & Heading, R. C. (1982) Gastric emptying of solids in man. Gut 23:292-296.[Abstract/Free Full Text]

61. Mourot, J., Thouvenot, P., Couet, C., Antoine, J. M., Krobicka, A. & Debry, G. (1988) Relationship between the rate of gastric emptying and glucose and insulin responses to starchy foods in young healthy adults. Am. J. Clin. Nutr. 48:1035-1040.[Abstract/Free Full Text]

62. Torsdottir, I., Alpsten, M. & Andersson, H. (1986) Effect of different starchy foods in composite meals on gastric emptying rate and glucose metabolism. II. Comparisons between potatoes. rice and white beans in diabetic subjects. Hum. Nutr. Clin. Nutr 40:397-400.[Medline]

63. Kim, H., Simpson, K.L. & Gerber, L.E. (1988) Serum carotenoids and retinol of human subjects consuming carrot juice. Nutr. Res. 8:1119-1127.

64. Bauernfeind, J. C. (1972) Carotenoid vitamin A precursors and analogs in foods and feeds. J. Agric. Food Chem. 20:456-473.

65. Roodenburg, A. J., Leenen, R., van het Hof, K. H., Weststrate, J. A. & Tijburg, L. B. (2000) Amount of fat in the diet affects bioavailability of lutein esters but not of -carotene, ß-carotene, and vitamin E in humans. Am. J. Clin. Nutr 71:1187-1193.[Abstract/Free Full Text]

This article has been cited by other articles:

				M. J Brown, M. G Ferruzzi, M. L Nguyen, D. A Cooper, A. L Eldridge, S. J Schwartz, and W. S White Carotenoid bioavailability is higher from salads ingested with full-fat than with fat-reduced salad dressings as measured with electrochemical detection Am. J. Clinical Nutrition, August 1, 2004; 80(2): 396 - 403. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Purchase Article View Shopping Cart Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Edwards, A. J. Articles by Parker, R. S. Search for Related Content PubMed PubMed Citation Articles by Edwards, A. J. Articles by Parker, R. S.