the nutritional effects of olestra

The Nutritional Effects of Olestra

Olestra Dose Response on Fat-Soluble and Water-Soluble Nutrientsin the Pig1,2,3

Dale A. Cooper, Delia A. Berry, Victoria A. Spendel, Dennis King, Anthony L. Kiorpes* andJohn C. Peters

The Procter & Gamble Company, Winton Hill Technical Center, Cincinnati, OH 45224 and *Hazleton-Wisconsin, Inc., Madison, WI 53704

ABSTRACT Groups of weanling pigs were fed a purified diet containing graded concentrations of olestra rangingfrom 1.1 to 7.7% (wt/wt) and the NRC*s requirements for micronutrients for 12 wk. Each group consisted of 12pigs, with the exception of the control group, which had 20, with equal numbers of females and castrated males.The purpose of the study was to determine the dose-response effects of olestra on fat-soluble vitamins andselected water-soluble micronutrients. At wk 0, 4, 8 and 12, hematology, clinical chemistry and blood concentra-tions of vitamins A, E, K and B12, and 25-hydroxyvitamin D, 1,25-dihydroxyvitamin D, folate, calcium, iron, zincand adipose concentration of vitamin E were measured. Cumulative weight gain and feed efficiency were deter-mined weekly. Prothrombin time was measured weekly for the control group and the groups fed 5.5 or 7.7% olestra,and monthly for other groups. Liver concentrations of vitamins A, E, and B12 and iron and bone concentrations ofcalcium, phosphorus, zinc and ash were measured for 12 pigs killed at wk 0 and for all animals at wk 12. By wk12, the pigs were eating from 20 to 155 g/d of olestra. Olestra did not affect the pigs’ growth or feed efficiency,indicating that the digestion and absorption of macronutrients were unaffected. Olestra reduced tissue concentra-tions of vitamin A, vitamin E and 25-hydroxyergocalciferol in a dose-responsive manner but did not affect prothrom-bin time. Olestra had no effect on the status of folate, vitamin B12, zinc or iron. Statistically reduced liver concentra-tions of vitamin B12 and iron in groups fed 5.5 or 7.7% olestra and a significant trend in bone ash content witholestra intake were possibly due to the poor vitamin A and/or vitamin E status of the pigs. J. Nutr. 127: 1573S–1588S, 1997.

KEY WORDS: • pigs • olestra • fat-soluble vitamins • minerals • folate • vitamin B12

Olestra is the common name for the mixture of hexa-, nal (GI)4 tract competes with the intestinal micelles forhepta- and octaesters of sucrose formed from long-chain the lipophilic nutrients. Lipophilic molecules that parti-fatty acids derived from edible oils. Olestra (Olean, Pro- tion into the olestra are not incorporated into the mixedcter & Gamble, Cincinnati, OH) has taste and cooking micelles and transported to the intestinal surface (Jandacekcharacteristics similar to those of traditional fats and oils 1982). Consistent with this mechanism, studies in normal(Bernhardt 1988, Kester 1993), but does not contribute healthy human subjects showed that olestra reduced serumany energy to the diet because it is not hydrolyzed by gastric concentrations of carotenoids, a-tocopherol, and 25-hy-lipases and therefore is not absorbed (Mattson and Vol- droxyergocalciferol [25(OH)D2 ], the metabolite of dietarypenhein 1972, Miller et al. 1995). Because of these unique vitamin D, but did not affect vitamin K status (Jones etproperties, olestra can serve as a zero-calorie replacement al. 1991a and 1991b, Koonsvitsky et al. 1997). Decreasesfor conventional fats and oils. in serum cholesterol concentration (Crouse and Grundy

Because olestra is lipophilic and is not absorbed, it can 1979, Fallat et al. 1976, Glueck et al. 1979) and increasesinterfere with the absorption of lipophilic nutrients. This in fecal cholesterol excretion (Jandacek et al. 1980 andinterference occurs because the olestra in the gastrointesti- 1990) have been seen in humans consuming olestra. Stud-

ies in the rat showed that olestra increased fecal cholesterolexcretion (Mattson et al. 1976) and reduced liver vitamin

1 Published as a supplement to The Journal of Nutrition. Guest editors for A stores (Mattson et al. 1979). Studies in the domestic pigthis supplement were John W. Suttie, University of Wisconsin, Department of showed that olestra reduced serum and liver concentrationsBiochemistry and Nutritional Sciences, 420 Henry Mall, Madison, WI and A. C.Ross, Pennsylvania State University, 126 S. Henderson Bldg., University Park,PA 16802.

2 Presented in part at Experimental Biology 94, March 1994, Anaheim, CA[Cooper, D., Berry, D., Jones, M., Spendel, V., Peters, J., King, D., Aldridge, D. & 4 Abbreviations used: BW, body weight; GI, gastrointestinal; MCH, mean cor-

puscular hemoglobin; MCHC, mean corpuscular hemoglobin concentration;Kiorpes, A. (1994) An assessment of the nutritional effects of olestra in the domes-tic pig. FASEB J. 8: A191 (abs. 1103)]. MCV; mean corpuscular volume, 1,25(OH)2D, 1,25-dihydroxyvitamin D; 25(OH)D,

25(OH)D2 / 25(OH)D3; 25(OH)D2, 25-hydroxyergocalciferol; 25(OH)D3, 25-hydro-3 Address correspondence to Suzette J. Middleton, Ph.D., The Procter &Gamble Company, Winton Hill Technical Center, 6071 Center Hill Road, Cincin- xycholecalciferol; PHT, parathyroid hormone; PT, prothrombin time; TIBC, total

iron-binding capacity.nati, OH 45224.

0022-3166/97 $3.00 q 1997 American Society for Nutritional Sciences.

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being fed experimental diets. During this period, they were changedof retinol and a-tocopherol, and serum concentration ofto a purified basal diet that provided levels of micronutrients repre-25(OH)D2 (Cooper et al. 1997c, Daher et al. 1997).senting the NRC’s requirements for 5–10 kg pigs (NRC 1988) andThe partitioning mechanism would lead one to expectcontained 14% fat (30% of digestible energy).that the effect of olestra on fat-soluble nutrients could be

During the acclimation period, the animals were housed three tooffset by introducing additional amounts of the affected five per pen and were observed daily for abnormalities indicative ofnutrients to olestra or olestra-containing foods. Studies in ill health. They underwent a complete physical examination. Hema-the pig (Cooper et al. 1997a and 1997b) and in humans tology and clinical chemistry measurements were made, and body(Koonsvitsky et al. 1997, Schlagheck et al. 1997a) have weights were taken. During the treatment period, the pigs weredemonstrated this outcome. housed in individual pens in a sunlight-free barn with controlled

temperature (above 187C), ambient humidity, and a 12-h ligh:darkThe purpose of this study was to determine the dosecycle. The pens were cleaned once or twice daily to reduce theresponse of olestra on selected nutrients with a broad rangepotential for coprophagy.of lipophilicity, in an animal model that closely resembles

Treatment groups and diets. Twelve pigs (six of each sex) werehumans with respect to the handling of these nutrients.selected randomly at the end of the acclimation period and killedThe domestic weanling pig was chosen as the animalto provide base-line data on nutrient status (base-line group). Themodel. The pig is an appropriate model to use for such remaining 92 pigs were randomized, balanced by body weight and

a study because of the similarity of the GI anatomy and assigned to one of seven treatment groups: 20 to the control groupphysiology and requirements for fat-soluble vitamins of and 10 each to the other six. Each group contained equal numberspigs and humans; there is also an extensive database on of females and castrated males.the pig’s nutrient requirements and metabolism (Miller The groups were fed a purified diet (ICN Biomedicals, Cleveland,

OH) providing the NRC requirements of micronutrients for 5- toand Ullrey 1987). In addition, the pig’s vitamin stores and10-kg swine (National Research Council 1988) and containing 0nutritional indices are responsive to dietary changes. The(control), 1.1, 2.2, 3.3, 4.4, 5.5 or 7.7% (wt/wt) olestra for 12 wk.weanling pig has a rapid growth rate and therefore high

The compositions of the diets are shown in Table A in the Appen-nutrient demands. Because of this, reductions in nutrientdix. The purified basal diet fed to the control group furnished 30%availability are easily detected in the growing pig. Previous of energy from fat and contained 140 g fat/kg of diet. This diet is thestudies have shown that the domestic pig is an appropriate same as that fed in other pig studies and has been shown to produce

model in which to evaluate the nutritional effects of olestra growth equivalent to that produced by standard swine diet (Cooper(Cooper et al. 1997c, Daher et al. 1997). et al. 1997c, Daher et al. 1997). Vitamins and minerals were added

On the basis of the partitioning mechanism (Jandacek to the diet as a premix, as described in Cooper et al. (1997c).The dietary concentrations of nutrients and olestra and the homo-1982), the nutrients with the greatest potential to be af-

geneity of the diets were confirmed by analysis. Because of a formula-fected by olestra are the fat-soluble vitamins. Thereforetion error, the diets provided only 5% of the requirement for vitaminthe effect of olestra on the status of vitamins A, D, EB12. This did not affect the integrity of the study, for reasons discussedand K was investigated. The partitioning mechanism wouldlater.lead one to predict that olestra would not affect the absorp-

The stability of olestra in the diets was confirmed as described intion of water-soluble nutrients. Nevertheless, the status of Cooper et al. (1997c). Stability of dietary nutrients was not deter-selected water-soluble micronutrients was monitored in mined. Previous analyses showed no instability of nutrients in thethis study to provide assurance that olestra does not affect same purified diet over a 4-wk period (Cooper et al. 1997c). Thethese nutrients. Folate and vitamin B12 were chosen as diets fed in this study were prepared in sequential batches so that theexamples of water-soluble nutrients that are digested and pigs never received diets that were older than 4 wk.

The olestra was prepared by the method of Rizzi and Taylor (1978)absorbed by complex multistep processes; hypothetically,and consisted of 80% octaesters and 20% heptaesters. The relativesuch processes provide the opportunity for olestra to inter-composition of the fatty acid chains was 20% palmitic, 5% stearic,fere with cleavage and binding reactions. In addition, these38% oleic, 28% linoleic, 7% behenic and 2% others, the same asnutrients are eaten in microgram amounts, whereas olestrathat used in other pig studies (Cooper et al. 1997a–c, Daher et al.is eaten in gram amounts, thus increasing the possibility 1997). Before being added to the diet, the olestra was used to frythat olestra might interfere with their absorption. potato chips under conditions representative of commercial frying

Calcium, iron and zinc were chosen as examples of wa- processes. This was done to ensure that the olestra fed the pigs waster-soluble micronutrients that many people consume at thermally stressed at least to the same degree as the olestra eaten bylevels that do not exceed and often do not meet recom- humans in savory snacks.

The lowest dietary concentration of olestra, 1.1%, was chosen tomended dietary intakes. Therefore any decrease in the bio-provide the pigs, at the start of the study, with a daily olestra intakeavailability of these micronutrients would be of potentialof about twice the estimated mean chronic human intake of olestranutritional concern.from savory snacks, 3.1 g/d (Webb et al. 1997). On the basis of thenormal growth of these crossbred pigs (Martin and Crenshaw 1989),the pigs in this group were expected to be consuming about eightMATERIALS AND METHODStimes that daily intake by the end of the study. The highest concentra-tion, 7.7%, was judged to be the maximum amount that could be fedThe study was conducted in accordance with the Food and Drugwithout introducing nutritional deficiencies resulting from dilutionAdministration Good Laboratory Practice Regulations for Nonclini-of the diet (Borzelleca 1992). This concentration would provide thecal Laboratory Studies (Hazleton Wisconsin, Madison, WI). All pro-pigs with a daily intake at the start of the study that would be aboutcedures involving animals complied with the Guide for Care and18 times the average chronic human intake, or about 8 times theUse of Agricultural Animals in Agricultural Research and Teaching90th-percentile chronic intake of olestra from savory snacks, 6.9 g/d(Consortium 1988).(Webb et al. 1997). The inclusion of extreme dietary concentrationsAnimals and husbandry. One hundred and four crossbred (one-of olestra increased the opportunity to see effects on water-solublehalf Duroc, one-quarter Landrace, one-quarter Large White) domesticmicronutrients, if such effects existed.pigs were used in the study (University of Wisconsin-Madison Swine

All pigs were exposed to 2 min/d of UV light (FS-40, T12-UVB-Unit, Madison, WI). One half were castrated males and one halfU, National Biological, Twinsbury, OH) to mimic the effect of sun-were females. The pigs were weaned at about 3 wk of age and fed alight on vitamin D synthesis. The 2-min daily exposure period wasstandard corn-soy–based swine starter diet formulated by the Univer-chosen to produce a 50–80% contribution of vitamin D3 to totalsity of Wisconsin-Madison. The pigs were received by the testing

laboratory at 4–5 wk of age and were acclimated for 10 d before vitamin D status (Cooper et al. 1997c). A 50% contribution of vita-


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TABLE 1

Biological response measured at scheduled intervals in pigs fed up to 7.7% olestra for 12 wk

Study week

Measurement 0 1 2 3 4 5 6 7 8 9 10 11 12

Body weight X X X X X X X X X X X X XClinical chemistry1 X X X XHematology1 X X X XSerum

Retinol X X X X25(OH)D2 X X X X1,25(OH)2D2 X X X Xa-Tocopherol X X X XTotal iron X X X XZinc X X X XCalcium X X X X

PlasmaPT3 X X X X X X X X X X X X XFolate X X X X

Adipose tissuea-Tocopherol X X X X

Liver tissueRetinol X Xa-Tocopherol X XCobalamin X XIron X XZinc X X

BoneAsh X XZinc X XCalcium X XPhosphorus X X

1A full battery, as presented in Cooper et al. (1997c).25-Hydroxyergocalciferol [25(OH)D2] and 25-hydroxycholecalciferol [25(OH)D3], separately; total 25(OH)D Å 25(OH)D2 / 25(OH)D3, 1,25(OH)2D

Å 1,25 dihydroxyvitamin D.3PT Å prothrombin time. Measured weekly for the control group and the groups fed 5.5 and 7.7% olestra; measured at wk 0, 4, 8 and 12 for

all other groups.

min D3 to total vitamin D status models the worst-case dependence D [1,25(OH)2D]. In addition, a complete battery of clinical chemistryand hematology parameters was measured, including total iron con-on dietary vitamin D in humans, paralleling the situation for elderly

people living in northern latitudes in winter (Delvin et al. 1979). centration, total iron-binding capacity (TIBC), total zinc concentra-tion and total calcium concentration. Folate concentration and pro-Feeding regimen. The pigs were fed sufficient diet daily to provide

95% of the recommended digestible energy for swine (NRC 1988). thrombin time (PT) were also measured.The liver was removed from each pig killed at wk 0 (base-lineDigestible energy content of the diet was calculated by summing the

digestible energy contributed, per kilogram of diet, by each dietary group) or wk 12 through an incision in the cranial abdomen and wasperfused with PBS. The entire left lateral lobe was frozen in dry iceingredient. The daily feed allotment for each pig was calculated on

the basis of the pig’s body weight at the beginning of each week and and homogenized. Portions of the powder were analyzed for all-trans-retinol, a-tocopherol, vitamin B12 (cyanocobalamin), iron and zinc.the projected weight gain over the week. The projected weight gain

was determined from growth curves for these crossbred pigs given Biopsies of adipose tissue were taken from the interscapular regionat wk 0, 4, 8 and 12 and analyzed for a-tocopherol. The fifth lumbarfree access to a standard corn-soy–based swine diet (Martin and

Crenshaw 1989). Feed was provided to the pigs in three equal-weight vertebra was collected from each pig killed at wk 0 or 12, groundinto a fine powder and analyzed for total ash content and zinc, calciumportions at 0730, 1200 and 1630 h for 45-min intervals. Any feed

not eaten at any specific session was collected and added to the and phosphorus concentrations.Analytical methods. All analytical methods were validated forsubsequent session. Any feed not eaten at the end of the day and any

spillage were collected and weighed to determine daily feed intake. use with biological samples from swine before the study samples wereanalyzed.Observations, necropsy, and tissue sampling. The observation,

measurement, and specimen collection schedule is shown in Table The concentrations of vitamin A (total retinol and retinyl esters)in liver and of vitamin E (a-tocopherol) in liver and adipose tissues1. The pigs were observed daily for clinical signs, including those of

nutritional deficiency, morbidity and mortality. Body weights (BW) were measured by HPLC following the method of Kayden et al.(1983). Samples were saponified with ethanolic KOH, and the vita-were determined weekly. Feed consumption, corrected for uneaten

feed and spillage, was measured daily. Cumulative weight gain, cumu- mins were extracted with hexane. Quantitation was by HPLC usinga silica column (Zorbax, 5-mm, Dupont, Wilmington, DE) and UVlative weight gain/wk-0 weight and cumulative weight gain/wk-0 body

surface area were calculated weekly. Body surface area was estimated detection. Retinol was detected at 325 nm; a-tocopherol was detectedby fluorescence excitation at 292 nm and emission at 325 nm. USPas (wk-0 body weight)0.75 (Kleiber 1975). Digestible feed efficiency

(body weight gain/digestible energy intake) was calculated weekly. standards of all-trans-retinol and a-tocopherol were used to generatestandard curves.Blood was collected at wk 0, 4, 8 and 12. The samples were taken

from the cranial vena cava before the morning feeding. Serum or Concentrations of vitamins A and E in serum were measured byHPLC, following the method of Driskell et al. (1982). Serum samplesplasma was stored at 0207C until analyzed. Serum was analyzed for

all-trans-retinol (vitamin A), a-tocopherol (vitamin E), 25(OH)D2, were deproteinized with ethanol and the vitamins extracted withhexane. Aliquots of extracts were reconstituted in hexane and in-25-hydroxycholecalciferol [25(OH)D3], and 1,25-dihydroxyvitamin


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jected onto an HPLC system equipped with a silica gel column (Zor- TABLE 2bax, 5-mm, DuPont) and a UV detector. Quantitation was by regres-sion analysis using standard curves generated with USP standards of Daily olestra consumption during wk 1, 6 and 12 by pigs fedall-trans-retinol and a-tocopherol.

up to 7.7% olestra for 12 wk1Serum concentrations of 25(OH)D2 and 25(OH)D3 were mea-

sured by HPLC, following the method of Kao and Heser (1984). TheGroup2 wk 1 wk 6 wk 12serum samples were acidified with concentrated HCl, and the 25-

hydroxy metabolites were extracted on prepacked octadecylsilano sil-% g/dica cartridges (C-18 Bond Elute, Analytichem International, Harbor

City, CA). The extracts were purified further by reextracting on1.1 6.0 { 0.8 12.9 { 1.0 20.4 { 0.8aminopropyl cartridges (NH2 Bond Elute, Analytichem Interna-2.2 10.3 { 1.3 22.5 { 1.9 34.9 { 1.1tional). The two 25-hydroxy metabolites were quantified simultane- 3.3 17.4 { 2.2 38.6 { 2.7 61.8 { 3.0ously with a silica column (Zorbax, 5-mm, Dupont) and UV detection 4.4 24.4 { 3.2 52.4 { 4.3 82.1 { 6.6

(Kratos Analytical, Ramsey, NJ). Concentrations of the two metabo- 5.5 30.9 { 3.7 68.8 { 5.3 109 { 4lites were calculated from a single calibration curve established with 7.7 47.6 { 5.3 102 { 7 155 { 11a 25(OH)D3 standard (Duphar, Amsterdam, The Netherlands). Re-covery was determined for each serum sample by adding a 25-hydroxy- 1Values are means { SD; (n Å 12).(25[27]-methyl-3H)cholecalciferol standard (Amersham, Arlington 2Percentage of olestra in diet.Heights, IL) to the sample before extraction.

The serum concentration of 1,25(OH)2D was measured by a radio- analyzed by repeated-measures ANOVA, using time, diet and genderreceptor method using a thymus receptor specific for both 1,25-dihy- as classification variables (Steel and Torrie 1960). To further investi-droxyergocalciferol [1,25(OH)2D] and 1,25-dihydroxycholecalciferol gate for potential differences, two-way ANOVA was conducted at[1,25(OH)3D] (Reinhardt et al. 1984). Briefly, serum samples were each time point, using diet and gender as class variables. Data col-spiked with [3H-26,27]-1,25(OH)2D (INCSTAR, Stillwater, MN), lected at single time points were analyzed by two-way ANOVA, usingextracted with acetonitrile, and purified further with activated C16OH diet and gender as classification variables. When the dose-by-gendercartridges (INCSTAR), following the method of Hollis (1986). interaction was significant, data from males and females were analyzedBound and free hormones were separated by incubation with charcoal. separately; otherwise the data were combined and intergroup compari-After incubation, the supernatants containing the bound hormone sons were made on the combined data. All comparisons were madewere decanted into scintillation tubes and the radioactivity content using a variability estimate from the two-way ANOVA, which in-determined with a scintillation counter (Model 1500, Packard Instru- creased the probability of detecting significant intergroup differences.ments, Downers Grove, IL). When significant differences were indicated by the two-wayPlasma prothrombin time was measured as part of the clinical ANOVA, the protected least significant difference (LSD) multiple-chemistry battery. comparison procedure was used to assess significant pairwise groupPlasma concentration of folate was measured by RIA (Matte and mean differences (Carmer and Swanson 1973, Welsch 1977).Girard 1989, O’Connor et al. 1989). Alkaline denaturation was used When a measured parameter fell below the detection limit of theto free the folic acid from carrier proteins in plasma. 125I-labeled folic analytical method, one half of the detection limit rather than zeroacid (Diagnostics Products, Los Angeles, CA) in buffered dithiothrei- was used as the value for that particular measurement (Helsel 1990).tol was added, and the samples were incubated with b-lactoglobulin All analyses were conducted at the two-tailed 0.05 significanceimmobilized on cellulose particles (Diagnostics Products). After incu- level, using PC SAS Version 6.04 or SAS Version 6.06 softwarebation, the samples were centrifuged and the radioactivity content (SAS Institute, Cary, NC).of the pellets was determined with a gamma counter (Packard Model5780). Quantitation was accomplished by the use of a standard curve.

RESULTSThe liver concentration of cyanocobalamin was measured by amicrobiologic assay (Baker and Frank 1968, Baker et al. 1986). Ali- Olestra consumption. The average daily amounts of olestraquots of liver homogenates were diluted 1:10 with 0.25% aconitic eaten by the pigs during wk 1 ranged from 6.0 g/d, for thoseacid/1.25% sodium cyanide buffer and autoclaved. After centrifuging in the 1.1% olestra group, to 47.6 g/d, for those in the 7.7%to remove debris, the supernatant was diluted again with the aconitic

olestra group (Table 2). Daily intake increased more thanacid/sodium cyanide buffer. Aliquots were added to Erlenmeyer flasksthreefold as the pigs grew over the course of the study. By wkcontaining the basal growth medium and were sterilized by autoclav-12, the pigs fed 1.1% olestra in the diet were eating 20.4 g/ding for 20 min. After being cooled to room temperature in a sterile

room, each flask was incubated with Ochromonas malhamensis for 5 d olestra, and the pigs fed 7.7% were eating 155 g/d.under light at 207C. Growth was measured with a densitometer and Growth of the pigs. No significant differences between thequantified by comparison with a standard growth curve. Recovery sexes were found with respect to the pattern of growth; there-was determined by spiking liver homogenates with a USP sample of fore the growth data for males and females were analyzed incyanocobalamin. The vitamin B12 assay was conducted by Vitamin combination. All of the groups grew at essentially the sameDiagnostics (Cliffwood Beach, NJ). rate. Initial group mean ({ SD) body weights ranged from 11.7To assay liver for iron and zinc, samples of homogenized liver { 1.7 to 12.4 { 1.6 kg. At wk 12, group mean ({ SD) bodywere digested with concentrated nitric acid and hydrogen peroxide.

weights ranged from 71.4{ 7.9 to 75.1{ 6.6 kg. No significantThe resulting solution was filtered and analyzed simultaneously fordifferences in cumulative digestible energy consumption, cu-iron and zinc by atomic emission spectroscopy (ARL 3560, Appliedmulative weight gain or feed efficiency were found amongResearch Laboratories, Sunland, CA), using an inductively coupled

plasma as the excitation source (Dahlquist and Knoll 1978). the groups at any time point. These parameters measured orTo assay for calcium, phosphorus and zinc in bone, fat was ex- calculated at the end of the study are shown in Table 3.

tracted from the ground vertebrae with pentane. The defatted sample General health. No unscheduled deaths occurred, and nowas weighed into a crucible, charred on a hot plate and ashed in a visible indications of clinical nutritional deficiency were foundmuffle furnace. The crucible was cooled and reweighed to determine among the pigs during the study. There were no antemortemash content. The result was expressed as a percentage of fat-free findings to indicate an adverse olestra effect (data not shown).dry weight. The resulting ash then was treated with concentrated

Variations in fecal consistency were noted among the groups.hydrochloric acid, dried, redissolved in hydrochloric acid, filteredGenerally, more observations of pasty feces and fewer observa-and assayed for calcium, phosphorus and zinc by inductively coupledtions of pelleted feces were associated with the groups fedplasma-atomic emission spectroscopy (Dahlquist and Knoll 1978).greater amounts of olestra. These effects result from the pres-Statistical analysis. Data collected as a function of time (growth

parameters, PT, blood and adipose nutrient concentrations) were ence of olestra in the feces.


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TABLE 5TABLE 3

Liver, serum, and adipose vitamin E concentrations for pigsCumulative energy consumption, cumulative weight gain anddigestible feed efficiency for pigs fed up to fed up to 7.7% olestra for 12 wk1

7.7% olestra for 12 wk1Group Liver2 Serum Adipose

Digestible% nmol/g liver mmol/L nmol/g tissueEnergy Weight feed

Group consumption gained efficiency20 18.28 { 3.10 4.22 { 1.13 23.7 { 4.41.1 7.56 { 1.29* 2.30 { 0.41* 12.3 { 2.2*

% MJ kg kg/MJ 2.2 5.63 { 1.04* 1.79 { 0.29* 9.7 { 1.6*3.3 4.67 { 0.76* 1.28 { 0.39* 8.7 { 1.5*0 2059 { 155 62.0 { 6.0 0.030 { 0.0024.4 4.08 { 0.91* 1.04 { 0.33* 7.6 { 1.9*1.1 2071 { 130 62.8 { 6.2 0.030 { 0.0025.5 3.70 { 0.94* 0.84 { 0.23* 6.4 { 0.9*2.2 2071 { 109 60.7 { 5.6 0.029 { 0.0027.7 3.53 { 1.14* 0.89 { 0.36* 6.3 { 1.3*3.3 2013 { 130 60.6 { 5.4 0.030 { 0.002

4.4 2021 { 180 59.4 { 6.5 0.029 { 0.0011Values are means { SD at end of study; 0% group (n Å 20); all5.5 2071 { 126 63.1 { 4.2 0.031 { 0.002

other groups (n Å 12).7.7 2059 { 117 60.1 { 5.4 0.029 { 0.0022Base-line group Å 6.69 { 1.91 nmol/g liver.* Significantly different than control (P õ 0.05).1Values are means { SD; 0% group (n Å 20); all other groups

(n Å 12).2Digestible feed efficiency Å cumulative weight gain/cumulative en- for the pigs fed 1.1 or 7.7% olestra was about 63 and 33% ofergy consumption.

control, respectively. By wk 12, the serum vitamin A concen-tration increased slightly in all olestra-fed animals, both in

Scattered significant differences among the groups were absolute terms and in relation to control.noted in a number of hematology and clinical chemistry pa- The liver concentration of vitamin E (a-tocopherol) de-rameters (data not shown). The differences were not dose creased with increasing dietary concentration of olestra (Tablerelated and were not consistent over time or between males 5). The mean concentration for the pigs fed 1.1% olestra wasand females. It was concluded that the differences represented about 41% of the control value; for the pigs fed 7.7% olestra,normal biological variability. it was about 20% of the control value. The pigs in the control

Fat-soluble vitamins. No significant differences between group increased their liver vitamin E stores by a factor ofmales and females were found in blood and tissue concentra- almost three during the study in relation to the value in thetions of fat-soluble vitamins; therefore the data for males and animals killed at base line (18.3 vs. 6.69 nmol/g). The pigsfemales were combined and analyzed. Separate analysis of the fed 1.1% olestra increased their liver vitamin E stores by aboutdata for each sex produced the same results. 13% over the course of the study.

Liver concentration of vitamin A (retinol) decreased in Serum vitamin E concentration decreased in a dose-respon-a dose-responsive manner with increasing concentrations of sive manner with increasing dietary concentration of olestra,dietary olestra (Table 4). The liver vitamin A concentration paralleling the change in the liver concentration of the vita-for the pigs fed 1.1 or 7.7% olestra was about 36 and 7% of min (Table 5). The dose response was essentially completecontrol, respectively. Liver vitamin A concentration for the within 4 wk (Table C in the Appendix). The largest effectpigs in the control group, 63.3 nmol/g, was about 29% greater was measured at wk 8, when serum vitamin E concentrationthan the value measured for the group killed at base line, 49.2 for the pigs fed 1.1 or 7.7% olestra was about 43 and 18% ofnmol/g. control, respectively. By wk 12, serum vitamin E concentration

Serum concentration of vitamin A also decreased in a dose- was increasing in all olestra-fed groups, both in absolute valuesresponsive manner with increased dietary olestra concentra- and in relation to control.tion (Table 4). The dose response was complete within 8 wk Serum vitamin E concentration normalized with respect to(Table B in the Appendix). At wk 8, the serum concentration serum lipids showed the same response to olestra as did the

nonnormalized concentration (data not shown).The concentration of vitamin E in adipose tissue decreased

TABLE 4 in a dose-responsive manner with increasing dietary concen-tration of olestra (Table 5). Unlike serum vitamin E, adiposeLiver and serum vitamin A concentrations for pigs fed up tovitamin E continued to decrease between wk 8 and 12 for the7.7% olestra for 12 wk1olestra-fed groups (Table D in the Appendix). At wk 12, theadipose vitamin E concentration for the pigs fed 1.1 or 7.7%Group Liver2 Serumolestra was about 52 and 27% of control, respectively.

At wk 12, serum 25(OH)D2 decreased in a dose-responsive% nmol/g liver mmol/Lmanner with increasing olestra dietary concentrations up to

0 63.3 { 11.9 0.64 { 0.19 3.3% but tended to increase with further increases in olestra1.1 23.0 { 6.7* 0.54 { 0.15dietary concentrations (Table 6). The dose response was pres-2.2 18.5 { 6.8* 0.48 { 0.08*ent at olestra dietary concentrations õ5.5% at wk 4 and3.3 8.4 { 4.8* 0.35 { 0.11*

4.4 6.5 { 4.7* 0.28 { 0.19* õ4.4% at wk 8 (Table E in the Appendix).5.5 4.9 { 3.1* 0.23 { 0.07* At wk 12, serum 25(OH)D3 concentration for the group7.7 4.4 { 3.6* 0.27 { 0.12* fed 1.1% olestra was significantly greater than the control

value (Table 6). The group fed 7.7% olestra had a mean serum1Values are means { SD at end of study; 0% group (n Å 20); all25(OH)D3 concentration which was significantly less than theother groups (n Å 12).control value (Table 6). Similar trends were observed at wk2Base-line group Å 49.2 { 6.8 nmol/g liver.

* Significantly different than control (P õ 0.05). 4 and 8 (Table F in the Appendix).


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TABLE 6

Serum 25-hydroxyergocalciferol [25(OH)D2], serum 25-hydroxycholecalciferol [25(OH)D3], total 25-hydroxyvitamin D [25(OH)D] and1,25-dihydroxyvitamin D [1,25(OH)2D] concentrations for pigs fed up to 7.7% olestra for 12 wk1

Group 25(OH)D2 25(OH)D3 25(OH)D 1,25(OH)2D

% nmol/L nmol/L nmol/L pmol/L

0 9.2 { 2.6 61.4 { 16.1 69.5 { 16.0 95 { 141.1 8.2 { 3.1 79.1 { 18.0* 86.0 { 17.1* 100 { 282.2 6.0 { 1.4* 67.9 { 21.9 72.8 { 20.9 100 { 193.3 5.6 { 1.4* 67.3 { 24.8 71.7 { 23.7 121 { 28*4.4 6.0 { 2.3* 64.8 { 13.1 69.6 { 14.4 104 { 215.5 6.1 { 2.1* 56.0 { 14.0 61.1 { 14.1 102 { 177.7 8.0 { 2.7 35.4 { 9.2* 42.8 { 10.3* 133 { 43*

1Values are means { SD at end of study; 0% group (n Å 20); all other groups (n Å 11–12).* Significantly different than control (P õ 0.05).

Serum concentrations of total 25(OH)D reflected fL for males and females, respectively, in the control group(complete MCV data are not shown). The liver concentrationchanges in the serum concentration of 25(OH)D3 , which

contributed from 87 to 89% of the 25(OH)D concentration of vitamin B12 measured at wk 12 in the control group, 64.2{ 13.3 nmol/g liver, was not significantly different than the(Table 6 and Table G in the Appendix). No olestra dose

response was found on the serum concentration of concentration measured in the group killed at base line, 80.4{ 16.2 nmol/g liver (Table 9).1,25(OH)2D (Table 6). A time-dependent decrease in serum

1,25(OH)2D was noted in all groups, including the control Liver iron concentration for the groups fed 5.5 or 7.7%olestra was significantly less than the control value, resultinggroup (Table H in the Appendix).

No effect of olestra was noted on PT (Table 7). PT measure- in a significant trend test (Table 10). No olestra dose responsewas found on serum TIBC or total iron (Table 10 and Table Iments were made weekly for the control, 5.5% olestra and

7.7% olestra groups. No significant differences were found in the Appendix). No olestra effects were noted on hematologyparameters such as RBC, mean corpuscular hemoglobinamong the PT values at any of these time points (data not

shown). (MCH) and mean corpuscular hemoglobin concentration(MCHC) (data not shown).Water-soluble micronutrients. There was no significant

difference between males and females in the response of blood There were no significant differences in liver, bone or serumconcentrations of zinc between the olestra-fed and controlor tissue concentrations of any of the water-soluble nutrients

to the diet. Therefore the data from the two sexes were com- groups and no significant trends in the data (Table 11 andTable J in the Appendix). A significant downward trend inbined and analyzed for intergroup differences. Separate analysis

of the data for each sex produced the same results. bone ash was found with increasing dietary olestra concentra-tion (Table 12). The value for the group fed 4.4% olestra wasNo olestra dose response on plasma folate concentration

was found (Table 8). At wk 4 and 8, plasma folate concentra- significantly less than the control value because one pig hada bone ash value of 55.3%. The mean for the rest of the animalstions for some olestra groups were significantly different than

control: some were less, some greater. in the group was 60.5 { 1.2%, not significantly different thanthe control value of 61.1 { 1.0%. No effect of olestra on boneLiver vitamin B12 concentration for the 7.7% olestra group

was significantly less than the control value and thus produced concentrations of calcium or phosphorus occurred (Table 12).Olestra had no significant effects on serum concentrationa significant trend (Table 9). However, mean corpuscular vol-

ume (MCV), which normally is elevated when vitamin B12 is of calcium or inorganic phosphorus (Table 13 and Table Kin the Appendix). Alkaline phosphatase was also unaffecteddeficient (Herbert 1988), was 54 { 2.5 fL for both males and

females in the 7.7% olestra group, and 52 { 3.5 and 55 { 2.0 by olestra intake (data not shown).

TABLE 7

Plasma prothrombin time for pigs fed up to 7.7% olestra for 12 wk1

Prothrombin time

Group wk 0 wk 4 wk 8 wk 12

% s

0 10.69 { 0.48 11.47 { 0.58 11.42 { 0.53 11.27 { 0.611.1 10.60 { 0.41 11.46 { 0.56 11.31 { 0.49 10.88 { 0.962.2 10.84 { 0.45 11.41 { 0.47 11.28 { 0.71 11.43 { 0.533.3 10.62 { 0.50 11.84 { 0.63 11.27 { 0.75 11.13 { 0.844.4 10.53 { 0.68 11.42 { 0.89 11.58 { 0.39 11.54 { 0.735.5 10.57 { 0.44 11.72 { 0.68 11.63 { 0.49 11.42 { 0.397.7 10.79 { 0.49 11.72 { 0.67 11.42 { 0.45 11.35 { 0.58

1Values are means { SD; 0% group (n Å 20); all other groups (n Å 11–12).


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TABLE 8

Plasma folate concentration for pigs fed up to 7.7% olestra for 12 wk1

Folate

Group wk 0 wk 4 wk 8 wk 12

% nmol/L

0 78.0 { 26.3 74.1 { 16.1 74.3 { 16.5 56.9 { 14.71.1 82.5 { 24.5 77.0 { 12.9 76.8 { 12.7 61.2 { 12.72.2 75.2 { 17.0 84.7 { 16.8* 88.1 { 18.8* 68.0 { 15.23.3 69.6 { 15.4 74.8 { 10.9 58.7 { 11.1* 60.5 { 7.94.4 79.5 { 12.7 69.1 { 13.6 53.9 { 14.0* 54.4 { 9.35.5 78.0 { 23.6 76.6 { 12.0 61.0 { 14.5* 58.7 { 13.47.7 75.9 { 22.0 88.1 { 17.2* 74.8 { 14.5 63.7 { 13.1

1Values are means { SD; 0% group (n Å 20); all other groups (n Å 11–12).* Significantly different than control (P õ 0.05).

DISCUSSION growth was the same for all groups and was essentially thesame as that observed when pigs are fed a standard corn-soy–

To ensure that any effect that olestra might have on fat- based swine diet (Martin and Crenshaw 1989).soluble vitamins and selected water-soluble micronutrients The absorption or utilization of macronutrients was unaf-would be observed and quantified, this study was conducted fected by olestra as evidenced by growth and the lack of anyusing dietary conditions which were exaggerated relative to effect on the amount of digestible energy consumed. Thesehow people will eat olestra. As a result, the effects measured findings are in agreement with those from other long-termare exaggerated relative to those likely to occur when people animal feeding studies conducted as part of the safety evalua-eat olestra snacks in free-living dietary patterns. Exaggerating tion of olestra (Lafranconi et al. 1994, Miller, K. et al. 1991,factors designed into the study included the frequency of oles- Wood et al.1991).tra consumption, the amount of olestra consumed and the In agreement with findings from other studies, there wasmanner (dietary context) in which it was fed. The pigs ate no effect of olestra on absorption of water-soluble nutrientsolestra at every feeding (42 times in 14 d), whereas snacks are despite the exaggerated conditions of the study (Cooper et al.eaten only 5 times in 14 d by the average consumer, only four 1997a, Schlagheck et al. 1997a and 1997b). The lack of anof those occurring with meals (Webb et al. 1996). By the last effect of olestra on the absorption of water-soluble nutrientsweek of the study, the pigs fed the lowest dietary concentration is consistent with the hypothesis that the only way olestraof olestra (1.1%) were eating about three times the estimated interferes with nutrient absorption is via the partitioning90th-percentile chronic human intake from savory snacks, 6.9 mechanism, i.e., lipophilic nutrients partition into the olestrag/d. The olestra was mixed into the diet before feeding, which and become unavailable to the intestinal micelles (Jandacekincreases the effects on the absorption of fat-soluble vitamins 1982).by two- to fivefold relative to the situation in which the olestra Olestra affected the absorption of vitamins A, D2 and E asis eaten in a snack food such as potato chips (Daher et al. expected on the basis of the partitioning mechanism and as1997). observed by others (Glueck et al. 1982, Jones et al. 1991b,

Olestra was well tolerated by the pigs as evidenced by the Koonsvitsky et al. 1997, Mattson et al. 1979, Mellies et al.lack of any health-related antemortem observations or changesin clinical chemistry or hematology measures. The rate of

TABLE 10TABLE 9

Liver iron, serum total iron-binding capacity (TIBC) and serumtotal iron concentration for pigs fed up toLiver vitamin B12 concentration for pigs fed up to

7.7% for 12 wk1 7.7% olestra for 12 wk1

Group Liver iron2,3 Serum TIBC Serum total ironGroup Vitamin B122,3

% nmol/kg % mmol/kg mmol/L mmol/L

0 64.2 { 13.3 0 2.47 { 0.86 113.4 { 10.5 23.7 { 10.91.1 2.27 { 0.68 115.0 { 7.9 26.0 { 9.21.1 67.9 { 21.4

2.2 59.8 { 15.5 2.2 2.61 { 0.56 116.3 { 4.7 26.9 { 8.03.3 2.49 { 0.48 114.4 { 10.8 26.8 { 3.23.3 59.8 { 11.1

4.4 56.1 { 14.0 4.4 2.38 { 0.48 116.2 { 7.4 28.6 { 6.65.5 1.99 { 0.59* 108.2 { 9.6 25.9 { 7.95.5 64.2 { 21.4

7.7 47.2 { 6.6* 7.7 1.90 { 0.43* 116.3 { 8.4 24.5 { 5.4

1Values are means { SD at end of study; 0% group (n Å 20); all1Values are means { SD from end of study; 0% group (n Å 20); allother groups (n Å 12). other groups (n Å 11–12).

2Base-line group Å 1.25 { 0.56 mmol/kg liver.2Base-line group Å 80.4 { 16.2 nmol/kg.3Downward trend (P õ 0.05); r2 Å 0.08. 3Downward trend (P õ 0.05); r2 Å 0.08.

* Significantly different than control (P õ 0.05).* Significantly different than control (P õ 0.05).


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TABLE 13TABLE 11

Serum calcium and inorganic phosphorus concentrations forLiver, bone and serum zinc concentrations for pigs fed up to7.7% olestra for 12 wk1 pigs fed up to 7.7% olestra for 12 wk1

Calcium Inorganic phosphorusGroup Liver2 Bone3 SerumGroup wk 12 wk 12

% mmol/kg wet wt mmol/kg fat-free bone mmol/L% mmol/L

0 2.39 { 0.58 2.48 { 0.23 21.4 { 7.81.1 2.35 { 0.72 2.48 { 0.31 20.8 { 4.7 0 2.56 { 0.13 2.23 { 0.232.2 2.25 { 0.60 2.58 { 0.09 20.7 { 4.4 1.1 2.57 { 0.09 2.33 { 0.193.3 2.14 { 0.41 2.42 { 0.12 20.5 { 4.5 2.2 2.60 { 0.10 2.31 { 0.224.4 2.00 { 0.43 2.35 { 0.20 21.8 { 6.0 3.3 2.59 { 0.09 2.26 { 0.125.5 2.26 { 0.89 2.42 { 0.21 21.1 { 7.1 4.4 2.58 { 0.12 2.42 { 0.307.7 2.48 { 0.63 2.49 { 0.31 20.6 { 4.4 5.5 2.53 { 0.09 2.24 { 0.24

7.7 2.56 { 0.10 2.28 { 0.201Values are means { SD at end of study; 0% group liver and serum

(n Å 20); 0% group bone (n Å 19); all other groups (n Å 11–12). 1Values are means { SD at end of study; 0% group (n Å 20); all2Base-line group Å 1.27 { 0.24 mmol/kg wet wt. other groups (n Å 11–12).3Base-line group Å 2.49 mmol/kg fat-free bone.

1983 and 1985, Schlagheck et al. 1997b). Olestra did not fed 1.1% olestra, the dietary concentration most closely relatedaffect vitamin K status in agreement with results from other to human intake, had vitamin A liver stores of about 33,580studies (Cooper et al. 1997b, Jones et al. 1991a, Koonsvitsky nmol. Although the concentration of vitamin A in the liveret al. 1997, Schlagheck et al. 1997b). was about 53% less for this group than for the group killed at

The dose response of olestra on vitamin A was established base line, there was almost a threefold net gain in total vitaminwith dietary sources of the vitamin, i.e., retinyl palmitate and A stores.b-carotene, similar to the sources in the U.S. diet (Olson Serum vitamin A concentration decreased slightly for the1988). The effect on the liver concentration of vitamin A can control group during the study, from 0.86 to 0.64 mmol/L.be interpreted as an effect on the absorption of the vitamin Serum vitamin A concentration has been found to drop inbecause olestra has been shown not to affect the irreversible pigs after weaning (Miller, E. et al. 1991). The dose responseloss of vitamin A from normal metabolism (Food Additive of olestra on serum vitamin A observed here was not surprisingPetition 7A 3997). In addition, the expansion of the liver in view of the liver concentration of the vitamin. Serum vita-pool size was the same in all groups, another factor that could min A reflects differences in intake of the vitamin when liveraffect the liver concentration of the vitamin. vitamin A is less thanÇ70 nmol/g liver, as it was in this study

The expansion in the size of the liver pool during growth (Hentges et al. 1952, Olson 1984, Sauberlich et al. 1974).of the pigs in this study had a substantial effect on the liver In humans, functional impairment, including impaired darkconcentration of vitamin A. However, the expansion did not adaptation, night blindness and ocular lesions, can occur whenaffect the dose response because it occurred to essentially the serum vitamin A falls below Ç0.35 mmol/L (10 mg/dL). Forsame degree in all groups. Calculations shown that the pigs example, corneal xerophthalmia occurs commonly in Indiankilled at base line had about 11,760 nmol (12,000 g BW 1 2 and Indonesia children with serum vitamin A concentrationsg liver/100 g BW 1 49 nmol vitamin A/g liver) of vitamin A õ 0.35 mmol/L (Gibson 1990). However, in pigs, visible signsliver stores, assuming that liver represents about 2% of body of vitamin A deficiency do not appear unless serum vitaminweight (Filer et al. 1966). At the end of the study, the pigs A concentrations are õ0.18 mmol/L. Hentges et al. (1952)

observed incoordination of movement, paresis of hind legs,tilted heads and night blindness in pigs only after plasma vita-TABLE 12min A concentrations fell below 0.18 mmol/L. In this study,the lowest average serum vitamin A concentration measuredBone ash content, bone calcium and phosphorusamong the pigs fed olestra was 0.23 { 0.07 mmol/L (Table 4).concentration in bone for pigs fed up toNone of the pigs showed clinical signs of vitamin A deficiency7.7% olestra for 12 wk1and there was no decline in growth or feed intake among theolestra-fed groups.Group Ash2,3 Calcium2 Phosphorus2

Liver, serum, and adipose concentrations of vitamin E re-% % fat-free dry wt sponded similarly to olestra intake, consistent with a reduction

in absorption efficiency. Each measure was affected essentially0 61.1 { 1.0 22.3 { 0.7 10.5 { 0.3 in the same way, although adipose concentration of the vita-1.1 61.7 { 1.1 22.3 { 0.6 10.5 { 0.3 min responded more slowly than liver or serum concentrations.2.2 61.5 { 0.8 22.4 { 0.3 10.6 { 0.2 Because olestra affects only the absorption of vitamin E, all3.3 60.6 { 1.1 22.5 { 0.6 10.6 { 0.3

pools of the vitamin would be expected to respond similarly4.4 60.0 { 2.0* 21.9 { 0.8 10.4 { 0.4in growing animals. Bieri (1972) found that plasma, liver and5.5 60.4 { 1.4 22.0 { 0.9 10.5 { 0.4

7.7 60.2 { 2.3 22.0 { 0.7 10.5 { 0.4 other tissue concentrations of vitamin E declined rapidly inweanling rats fed vitamin E–deficient diets; most of the change

1Values are means { SD; 0% group (n Å 19); all other groups (n Å occurred within 2 wk. Machlin et al. (1979) found that11–12); values are from end of study. changes in availability of dietary vitamin E produced parallel2Base-line group Å 50.9 { 1.5, 18.2 { 0.7 and 9.0 { 0.3% fat-free

changes in plasma and tissue concentrations within a fewdry wt for bone ash, calcium and phosphorus, respectively.weeks in young guinea pigs.3P õ 0.05 for trend; r2 Å 0.1.

* Significantly different than control (P õ 0.05). The concentration of vitamin E in adipose tissue changed


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more slowly than serum and liver concentrations in response test noted in this study) was probably not an effect of olestraon vitamin B12 absorption. Vitamin B12 absorption was notto olestra because of a slower expansion of this pool, in relation

to lean tissues, in pigs in this weight range (Shields et al. affected when adult human subjects ate up to 32 g of olestrain a meal along with vitamin B12 (Schlagheck et al. 1997a1983). Machlin et al. (1979) found that the adipose pool of

vitamin E decreased more slowly than other tissue pools in and 1997b) or when pigs were fed up to 7.7% olestra withgraded levels of vitamins A, D, and E for 12 wk, or up to 5.5%young guinea pigs fed a vitamin E–free diet.

At olestra dietary levels below 3.3–5.5%, depending when olestra with graded levels of vitamins A and E for 26 wk(Cooper et al. 1997a and 1997b). In addition, MCV, whichthe measurement was made (wk 4, 8 or 12), the responses of

serum 25(OH)D2 and 25(OH)D3 to olestra were as expected, is normally elevated when vitamin B12 is deficient (Herbert1988), was unaffected in this study, further supporting thei.e., serum 25(OH)D2 declined in a dose-responsive manner

and serum 25(OH)D3 was unaffected. The shape of the conclusion that vitamin B12 absorption was not directly af-fected by olestra. MCV was 54 { 2.5 fL for both males and25(OH)D2 response was similar to the responses observed for

vitamins A and E, indicating that olestra interacts with vita- females in the 7.7% olestra group, in which the liver vitaminB12 concentration was the lowest, compared with control val-min D in the same way as it interacts with the other fat-

soluble vitamins. ues of 52 { 3.5 fL (males) and 55 { 2.0 fL (females).The low liver vitamin B12 concentration for the groups ofThe increase in serum 25(OH)D2 and the decrease in serum

25(OH)D3 observed with long-term consumption of high di- pigs fed 7.7% olestra was possibly a result of the poor vitaminE status of those pigs. The mean liver vitamin E concentrationetary levels of olestra were unexpected. Such effects were not

seen in pigs fed up to 5.5% olestra for 26 wk without exposure for that group, 3.53 nmol/g, was only slightly greater than thevalue of 3.0 nmol/g associated with signs of vitamin E defi-to UV light (Cooper et al. 1997b) or in human subjects con-

suming up to 32 g/d olestra (Schlagheck et al. 1997b). A ciency in pigs (Jensen et al. 1988). It is known that a-tocoph-erol is needed for the conversion of methylcobalamin to adeno-plausible explanation for the reciprocal changes in serum con-

centrations of the two metabolites is competition between sylcobalamin (Turley and Brewster 1993), the major form ofvitamin B12 stored in the liver (Herbert and Das 1994). Thevitamin D3 and vitamin D2 for liver 25-a-hydroxylase. In pigs,

vitamin D3 is the preferred substrate for 25-a-hydroxylase microbiologic assay used to determine the concentration ofvitamin B12 in the liver in this study measures the metaboli-(Horst et al. 1982); therefore an increase in serum 25(OH)D2

might occur if the amount of vitamin D3 available for hydroxyl- cally active forms of the vitamin, including adenosylcobalamin(Baker and Frank 1986, Baker et al. 1986). Pappu et al. (1978)ation decreased.

The declines in serum 25(OH)D3 and serum 25(OH)D showed that the synthesis of adenosylcobalamin is inhibitedin the rat when the liver concentration of vitamin E is low.noted with long-term consumption of 5.5 and 7.7% olestra

may have resulted from an interference with resorption of the The fact that no effects on liver vitamin B12 were observed inthe pig studies in which extra amounts of vitamin E wereenterohaptic-circulating forms of vitamin D3. In these pigs,

almost 90% of the serum 25(OH)D concentration came from added to the diet supports the conclusion that the effects seenin this study were probably related to the vitamin E status of25(OH)D3. Vitamin D3 and 25(OH)D3 are secreted in the

bile, and some fraction is reabsorbed from the intestine (Ar- the pigs and not to a direct effect of olestra on vitamin B12

absorption.naud et al. 1975, Avioli et al. 1967, Nagubandi et al. 1980).A similar change in 25(OH)D3 has been reported for humans The lack of any effect on the status of either vitamin B12

or folate, both water-soluble nutrients absorbed by complexeating large amounts of dietary fiber (Batchelor and Compston1983), and reductions in vitamin D status have been observed multistep processes (Herbert and Colman 1988), provides as-

surance that olestra does not interfere with nutrient availabil-in clinical malabsorption syndromes in which the resorptionof biliary-derived substances is hindered (Batchelor et al. 1982, ity by means other than the partitioning of fat-soluble nutri-

ents into the olestra.Compston et al. 1982).The decline in serum 1,25(OH)2D concentration noted in Although the liver concentrations of iron for the pigs fed

5.5 or 7.7% olestra were significantly less than the controlall groups was a result of aging (Horst and Littledike 1982,Lachenmaier-Currle and Harmeyer 1988). Serum 1,25(OH)2D value, no effects on serum TIBC or serum total iron concentra-

tion or on the hematology parameters RBC, MCH and MCHCis tightly regulated by calcium, phosphorus and parathyroidhormone (PTH) and is a reliable indicator of changes in di- were observed for these groups, indicating that the effects on

the liver concentration were probably not direct effects ofetary vitamin D intake only under deficiency conditions(Schrijver 1991). olestra on iron absorption. Other studies in pigs (Cooper et

al. 1997a and 1997b) and humans (Schlagheck et al. 1997aThe lack of an effect of olestra on PT agrees with resultsfrom other pig (Cooper et al. 1997a and 1997b) and human and 1997b) showed no effect of olestra on iron absorption.

Also, there was no effect of olestra on any of the measures ofstudies (Jones et al. 1991a, Koonsvitsky et al. 1997, Schlaghecket al. 1997a and 1997b). Any effect of olestra on the absorption zinc status of the pigs. If the effect on liver iron concentration

for the 5.5 and 7.7% olestra groups resulted from a direct effectof phylloquinone, and measurements of serum phylloquinoneconcentrations in humans indicate that there is such an effect of olestra on iron absorption, a similar effect might have been

expected on zinc absorption.(Schlagheck et al. 1997a and 1997b), is not sufficient to affectvitamin K functional status. The low liver concentration of iron for the pigs fed the

highest levels of olestra may have resulted from the poor vita-Although vitamin B12 was fed at only 5% of the NRCrequirement, the liver concentrations were in the range known min A status of those pigs. Vitamin A is required for normal

hematopoiesis, and improvements in iron status with vitaminto be responsive to changes in intake of the vitamin (Catronet al. 1952). The amount of vitamin B12 fed was sufficient to A treatment have been observed among children with mar-

ginal iron status (Bloem et al. 1989, Mejia and Chew 1988).allow the pigs in the control group to increase their total poolof vitamin B12 by a factor of almost 5, calculated as discussed The significant decreasing trend in bone ash with increasing

dietary levels of olestra was most likely a secondary effect of theabove for vitamin A.The significantly lower liver vitamin B12 concentration for low vitamin A status of the animals. In vitamin A deficiency,

deposition of calcium in bone is decreased; this decrease canthe pigs fed 7.7% olestra (and the resulting significant trend


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1582S SUPPLEMENT

Developing a Guide for the Care and Use of Agriculture Animals in Agriculturelead to a decrease in bone ash (Navia and Harris 1980). OtherResearch, Champaign, IL.indices of calcium status such as bone and serum calcium and Cooper, D. A., Berry, D. A., Jones, M. B., Kiorpes, A. L. & Peters, J. C. (1997a)

phosphorus concentrations were unaffected, and no effect of Olestra’s effect on the status of vitamins A, D and E in the pig can be offsetby increasing the dietary levels of these vitamins. J. Nutr. 127: 1589S–1608S.olestra has been found in other pig studies. Importantly, when

Cooper, D. A., Berry, D. A., Spendel, V. A., Jones, M. B., Kiorpes, A. L. & Peters,pigs were fed up to 7.7% olestra and graded levels of vitamins J. C. (1997b) Nutritional status of pigs fed olestra with and without in-A, D and E, no effect of olestra on bone ash content or other creased dietary levels of vitamins A and E in long-term studies. J. Nutr. 127:

1609S–1635S.measures of calcium status was seen (Cooper et al. 1997a).Cooper, D. A., Berry, D. A., Spendel, V. A., Kiorpes, A. L. & Peters, J. C. (1997c)Similarly, when pigs were fed up to 5.5% olestra for 26 wk

The domestic pig as a model for evaluating olestra’s nutritional effects. J.and PTH concentration was measured, in addition to the mea- Nutr. 127: 1555S–1565S.

Crouse, J. R. & Grundy, S. M. (1979) Effects of sucrose polyester on choles-sures used in this study, there was no effect of olestra on anyterol metabolism in man. Metabolism 28: 994–1000.of the calcium status measures (Cooper et al. 1997b). PTH,

Daher, G. C., Cooper, D. A. & Peters, J. C. (1997) Physical or temporal separa-in addition to 1,25(OH)2D, is a primary factor in calcium tion of olestra and vitamins A, E and D intake decreases the effect of olestraon the status of the vitamins in the pig. J. Nutr. 127: 1566S–1572S.homeostasis (Avioli 1988), and changes in serum calcium con-

Dahlquist, R. L. & Knoll, J. W. (1978) Inductively coupled plasma-atomic emis-centration lead to reciprocal changes in serum PTH concentra-sion spectrometry: analysis of biological materials and soils for major, trace,tion (Silver 1992). and ultra-trace elements. Appl. Spectrom. 32: 1–29.

Findings from this study generally agreed with those from Delvin, E. E., Glorieux, F. H., Dussault, M., Bourbonnais, R. & Watters, G. (1979)Simultaneous measurement of serum 25-hydroxycholecalciferol and 2-hydro-other pig and human studies and confirmed expectations basedxyergocalciferol. Med. Biol. 57: 165–170.on the partitioning mechanism. Because of the similarities Driskell, W., Neese, J. W., Bryant, C. C. & Bashor, M. (1982) Measurement of

between the GI tracts of weanling pigs and young children vitamin A and vitamin E in human serum by high-performance liquid chroma-tography. J. Chromatogr. 231: 439–444.(Leary and Lecce 1976), these findings are especially important

Fallat, R. W., Glueck, C. J., Lutmer, R. & Mattson, F. H. (1976) Short term studyin understanding potential olestra effects in children. Becauseof sucrose polyester, a nonabsorbable fat-like material as a dietary agent for

of the dietary conditions used in the study, it is unlikely that lowering plasma cholesterol. Am. J. Clin. Nutr. 29: 1204–1215.Filer, L. J., Jr., Churella, H., Knauff, R. & Vaughan, O. W. (1966) Effects ofeffects not seen in the pig will be seen in humans. Further,

dietary calcium, phosphorus, and strontium on growth, organ weights, andthe effects of olestra on the absorption of fat-soluble vitaminsbone composition of miniature swine. In: Swine in Biomedical Research (Bus-will be considerably less in people eating olestra in real-life tad, L. K. & McClellan, R. O., eds.), pp. 151–162. Pacific Northwest Labora-

conditions than those measured in pigs in this study. tory, Seattle, WA.Food Additive Petition 7A 3997, Appendix EC-30, April 1, 1987.Fraser, D. R. (1984) Vitamin D. In: Nutrition Reviews’ Present Knowledge in

Nutrition, pp. 209–225. Nutrition Foundation, Washington, DC.ACKNOWLEDGMENTSGibson, R. S. (1990) Assessment of the status of vitamins A, D, and E. In:

Principles of Nutritional Assessments, pp. 377–412. Oxford University Press,The authors would like to acknowledge D. H. Tallmadge for ana-New York, NY.lytical support and L. J. Bishop, S. J. Middleton and K. D. Lawson

Glueck, C. J., Hastings, M. M., Allen, C., Hogg, E., Baehler, L., Gartside, P. S.,for assistance in preparing the manuscript.Phillips, D., Jones, M., Hollenbach, E. J., Braun, B. & Anastasia, J. V. (1982)Sucrose polyester and covert caloric dilution. Am. J. Clin. Nutr. 35: 1352–1359.LITERATURE CITED Glueck, C. J., Mattson, F. H. & Jandacek, R. J. (1979) The lowering of plasmacholesterol by sucrose polyester in subjects consuming diets with 800, 300Arnaud, S. B., Goldsmith, R. S., Lambert, P. W. & Go, V.L.W. (1975) 25-Hy-or less than 50 mg of cholesterol per day. Am. J. Clin. Nutr. 32: 1636–1644.droxyvitamin D3: evidence of an enterohepatic circulation in man. Proc. Soc.

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1585SOLESTRA DOSE RESPONSE: APPENDIX


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1587SOLESTRA DOSE RESPONSE: APPENDIX


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the nutritional effects of olestra

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