Olestra is a mixture of hexa-, hepta- and octaesters of sucrose formed from long-chain fatty acids derived from edible oils. Olestra (Olean, Procter & Gamble, Cincinnati, OH) has taste and cooking characteristics similar to those of traditional dietary fats and oils (Bernhardt 1988, Kester 1993) but is not absorbed (Mattson and Volpenhein 1972, Miller et al. 1995). Consequently, olestra contributes no energy to the diet and can serve as a zero-calorie replacement for conventional fats and oils. Olestra is approved for use in preparing savory snacks such as potato chips and crackers.

Because it is lipophilic and is not absorbed, olestra can interfere with the absorption of fat-soluble micronutrients such as the fat-soluble vitamins. This interference occurs because some portion of the fat-soluble vitamins in the gastrointestinal (GI)⁴ tract partitions into the olestra and therefore becomes unavailable to the micelle-mediated absorptive process (Jandacek 1982). The extent to which olestra reduces the absorption of fat-soluble vitamins has been established in studies in the pig and in normal healthy human subjects (Cooper et al. 1997a and 1997b, Schlagheck et al. 1997b).

Animal and human studies have shown that olestra's effects on the absorption of fat-soluble vitamins can be offset by adding extra amounts of the vitamins to the diet. Mattson et al. (1979) measured the liver vitamin A content of rats fed 15% olestra in a vitamin A-free casein-based diet supplemented with two concentrations of retinyl palmitate. After 3 d of consuming the diet, the rats were killed and the vitamin A content of the liver was measured. The total amounts of retinyl palmitate fed over the 3-d period were 0, 1230 and 2420 IU [0, 677 and 1331 RE (µg retinol equivalents), respectively]. The mean liver content of vitamin A for the three groups was <150 IU (<50 RE), 599 IU (180 RE) and 1143 IU (343 RE), respectively.

In another study, mice were fed 0, 2.5, 5 or 10% olestra for 2 y. The diets were supplemented with 2500 IU (1375 RE)/kg of vitamin A (retinyl palmitate), 750 IU (18.8 µg)/kg of vitamin D and 160-640 IU [107-429 mg -tocopherol equivalents (-TE)/kg of vitamin E] (Lafranconi et al. 1994). Liver concentrations of vitamins A and E were measured periodically throughout the study; the serum concentration of total 25-hydroxyvitamin D [25(OH)D] was measured after 9 mo. The liver concentrations of vitamins A and E for the olestra-fed groups were comparable to control values. The serum 25(OH)D concentrations for the groups fed 2.5 or 5% olestra were comparable to control values; the concentrations for the group fed 10% olestra were 70 and 85% of control for males and females, respectively.

Free-living human subjects were given 18 g/d olestra with and without added d--tocopheryl acetate for 16 wk (Koonsvitsky et al. 1997). Serum -tocopherol concentration was reduced by 6%, relative to control, for the olestra group. The addition of 1.1 mg d--tocopheryl acetate/g olestra restored serum -tocopherol concentration by about one third.

This study was undertaken to establish the relationship between the dietary concentration of olestra and the amount of vitamins A, D and E required to restore tissue concentrations of the vitamins to control concentrations, i.e., the restoration levels. It was of particular interest to determine whether the restoration level is a linear function of the dietary concentration of olestra. Another purpose of the study was to confirm that olestra does not affect vitamin K function, the status of water-soluble micronutrients or the absorption or utilization of macronutrients.

The study was conducted in the domestic pig. Previous studies have shown that the domestic pig is an appropriate model in which to evaluate the nutritional effects of olestra (Cooper et al. 1997c, Daher et al. 1997). The pig's GI anatomy and physiology are very similar to those of humans (Graham and Aman 1987, Leary and Lecce 1976), and its vitamin stores and nutritional indices respond to dietary changes (Miller and Ullrey 1987). The dose-responsive effects of olestra on tissue concentrations of vitamins A and E and 25-hydroxyvitamin D₂ have been established in the pig (Cooper et al. 1997c).

MATERIALS AND METHODS

Table 1. Treatment groups and target levels of vitamins A, D and E in diets fed to pigs for 12 wk1 [View Table]

The medium concentration (MV) of vitamins A, D and E added to the diets was chosen on the basis of data from a 4-wk study (Cooper et al. 1997c). The high (HV) and low (LV) vitamin concentrations were chosen to provide tissue concentrations of the vitamins that evenly bracketed the concentration provided by the medium vitamin concentration, and to provide overlap in the range of vitamins added at each olestra concentration. The total dietary concentrations of vitamins A, D, and E are shown in Table 1, for each dietary concentration of olestra.

The extra vitamin A added to the basal diet was retinyl palmitate, the extra vitamin E was added as d--tocopheryl acetate, and the extra vitamin D was added as ergocalciferol.

The olestra used in the study had the same composition as that used in other pig studies (Cooper et al. 1997a-c, Daher et al. 1997). It was heated before being added to the diet, as described in Cooper et al. (1997b). The dietary concentrations tested, 1.1, 4.4 and 7.7%, were the low, medium and high dietary concentrations tested in a dose-response study in the pig (Cooper et al. 1997b). The lowest concentration provided a daily intake at the start of the study (5-6 g/d) that is about twice the estimated mean human chronic intake of olestra from savory snacks, 3.1 g/d (Webb et al. 1997). The highest dietary concentration, 7.7%, was the maximum concentration judged reasonable to test without potentially introducing nutritional deficiencies in the animals as a result of dilution of the diet with the nondigestible olestra (Borzelleca 1992).

The stability of olestra in the diets was confirmed as described in Cooper et al. (1997c). The concentrations of other dietary nutrients and the homogeneity of the diets were confirmed by analysis. Because of a formulation error, the diets provided only about 6% of the NRC requirement for vitamin B₁₂. The integrity of the study was unaffected by this lower-than-target concentration of the vitamin as discussed in a later section.

The concentrations of olestra and vitamins A, D and E in the diets were measured by the methods described by Cooper et al. (1997c). These measured concentrations were used to calculate the extra amounts of vitamins A, D and E required to restore tissue concentrations of these vitamins to control concentrations for pigs fed olestra-containing diets.

To determine the vitamin restoration levels, expressions were developed that related changes in tissue concentrations of the vitamin, relative to those produced by the basal diet, to dietary concentrations of the vitamin, over and above the concentration in the basal diet, and to the dietary concentration of olestra. These expressions were developed by a multiple-regression procedure. Multiple regression, in contrast to linear regression, was used because it allowed the simultaneous investigation of both independent variables (dietary concentrations of vitamins and olestra).

Before the regression was performed, differences were calculated between tissue concentrations of the vitamins measured in individual pigs fed olestra-containing diets and the mean values in the control group (the pigs fed the basal diet). This calculation yielded the net tissue response to the supplemental vitamin added over and above the concentrations in the basal diet and to the olestra in the diet. Similarly, the concentrations of vitamins in the olestra-containing diets were converted to amounts over and above the amounts in the basal diet. Because the basal diet contained no olestra, the dietary concentrations of olestra were over and above basal amounts.

A second-order zero-intercept model was used for the multiple regression. A zero-intercept model is indicated because additional vitamin is not needed when the diet contains no olestra. The model is described by the following general equation: where C is the difference between individual and mean control tissue vitamin concentration, OA is the percentage of olestra in diet, V is the amount of vitamin in the olestra-containing diet over and above the amount in the basal (control) diet, B_n are constants and error is the variability not explained by the model.

The Shapiro-Wilk test indicated that the errors were normally distributed for all responses (Shapiro and Wilk 1965). The model assumed that there were no interactions among the vitamins.

RESULTS

Table 2. Average daily olestra consumption during wk 1, 6 and 12 by pigs fed olestra and graded levels of vitamins A, D, and E for 12 wk1 [View Table]

At the end of the study, there were no significant differences among the groups in cumulative digestible energy consumption, in cumulative weight gain or feed efficiency (Table 3). No significant differences occurred among the groups for these parameters at intermediate time points or for growth expressed relative to wk-0 body weight or wk-0 body surface area (data not shown). Growth of all groups was essentially the same as that observed in the 12-wk dose-response study (Cooper et al. 1997b) and similar to that observed for pigs fed a standard swine diet (Martin and Crenshaw 1989).

Table 3. Cumulative energy consumption, cumulative weight gain and digestible feed efficiency for pigs fed olestra and graded levels of vitamins A, D and E for 12 wk1 [View Table]

Scattered significant differences among the groups were noted in some hematology and clinical chemistry parameters (data not shown). The differences, however, were not consistent over time, within groups fed the same concentrations of olestra, or between sexes, and thus were concluded to represent normal biological variability.

One animal in the 1.1% MV group died during wk 12 of the study. Necropsy observations suggested that the cause of death was blood loss from a gastric ulcer. It is unlikely that the ulcer was related to olestra consumption for several reasons. The animal was in the group fed the lowest dietary olestra concentration in the study. No deaths were observed in a 12-wk dose-response study in which pigs were fed up to 7.7% olestra (Cooper et al. 1997b) or in a 26-wk study in which pigs were fed up to 5.5% olestra (Cooper et al. 1997a). Gastric ulcers are common in swine and are due to various etiologic factors (Reese et al. 1966). Thus it is likely that the ulcer resulted from one or more of the processes that normally cause ulcers in swine.

Vitamin A. The liver concentration of vitamin A (total retinol) increased with increasing dietary concentrations of the vitamin in all olestra-fed groups (Fig. 1, Table C in the Appendix). For each dietary olestra concentration, the range of liver vitamin A concentrations produced by adding vitamin A to the diet encompassed the concentration measured in the control animals.

Multiple regression of the liver vitamin A produced the following relationship between liver vitamin A concentration and dietary concentrations of vitamin A and olestra: where _LA is the difference between the vitamin A liver concentration for a pig fed olestra with added vitamin A and the mean vitamin A liver concentration for the control group, OA is the dietary concentration of olestra (wt/wt%), and V_A is the dietary concentration of vitamin A (IU/kg diet), i.e., the restoration level. The response surface produced by this equation showed that the relationship was essentially linear for the range of dietary olestra concentrations tested. To derive an expression describing the relationship between the restoration level of vitamin A (V_A) and the dietary concentration of olestra (OA), this equation was solved for the three dietary concentrations of olestra tested, 1.1, 4.4 and 7.7%, by setting _LA equal to 0 and inserting the dietary concentration of olestra. Then a linear regression was performed on the three resulting values of V_A , using a zero-intercept model. Figure 2 shows the linear relationship between V_A , expressed as RE/kg diet and the dietary concentration of olestra, along with the specific restoration levels obtained from solution of the above equation for 1.1, 4.4 and 7.7% olestra. The equation for the relationship is as follows: where RL_A is the amount of vitamin A, expressed as RE/kg of diet, required to restore liver vitamin A concentration to the control concentration and OA is the dietary concentration of olestra, expressed as (wt/wt)% in the diet.

As vitamin A was added to the diet, the serum concentration of vitamin A increased in a dose-responsive manner (Table D in the Appendix). Serum vitamin A concentration is proportional to liver stores of the vitamin only when those stores are less than ~50 nmol/g liver (Hentges et al. 1952). This relationship was not true for many of the groups (Table C in the Appendix); therefore the serum data were not used to calculate a restoration level of vitamin A.

Vitamin E. The liver concentration of vitamin E increased in a dose-responsive manner for all olestra-fed groups as the dietary concentration of the vitamin was increased (Fig. 3 and Table E in the appendix). For each dietary olestra concentration, the amounts of vitamin E added to the diet produced liver vitamin E concentrations that encompassed the mean value measured in the control animals.

The relationship between vitamin E liver concentration and the dietary concentrations of vitamin E and olestra, derived by multiple regression of the liver vitamin E data as described above, is expressed by the following equation; where _LE is the difference between the liver concentration of vitamin E for a pig fed olestra with added vitamin E and the mean vitamin E liver concentration for the control group, OA is the dietary concentration of olestra (wt/wt%), and V_E is the dietary concentration of vitamin E (-TE/kg diet), i.e., the restoration level. As with vitamin A, the response surface produced by the above equation showed that the relationship is essentially linear over the range of olestra intakes tested.

The procedure described above produced the relationship shown in Figure 4. The equation describing the relationship is as follows: where RL_LE is expressed in -TE/kg diet and OA is expressed as a percentage in the diet (wt/wt).

The vitamin E concentration in the serum increased in a dose-responsive manner with increased concentrations of added vitamin E at all time points (Table F in the Appendix). At wk 12, the serum vitamin E concentration produced by the added vitamin E encompassed the control values for the 4.4 and 7.7% olestra groups (Fig. 5). The serum vitamin E concentrations for the 1.1% olestra groups fell slightly below the control value.

The relationship between serum vitamin E concentration and the dietary concentrations of vitamin E and olestra, obtained by multiple regression of the wk-12 serum concentration data, is expressed by the following equation:

Here _SE is the difference between the serum vitamin E concentration in a pig fed olestra for 12 wk and the mean serum vitamin E concentration for the control group. OA and V_E are as defined above. Like the relationship derived from the liver vitamin E data, this relationship is essentially linear over the range of dietary concentrations of olestra tested.

By solving this equation for the three concentrations of olestra tested and carrying out a linear regression on the three values obtained, the relationship between the amounts of additional vitamin E needed to restore serum vitamin E concentration (RL_SE) and the dietary concentration of olestra (OA) shown in Figure 6 was found. The equation of this relationship is as follows: where the restoration level, RL_SE , is expressed as -TE/kg diet and OA is expressed as a percentage in the diet (wt/wt). This equation agrees closely with that derived from the liver vitamin E concentration.

The concentration of vitamin E in adipose tissue increased in a dose-responsive manner as vitamin E was added to the diet (Table G in the Appendix). At wk 12, the resulting concentrations for the groups fed 1.1 or 7.7% olestra encompassed the control value; the concentration for the group fed 4.4% olestra with the high concentration of vitamin E was essentially the same as the control value. At wk 8, only the concentrations in the group fed 7.7% olestra encompassed the control value. The adipose concentration data were not used to calculate a restoration level of vitamin E.

Vitamin D. Adding ergocalciferol to the olestra-containing diets produced increases in serum 25-hydroxyergocalciferol [25(OH)D₂] concentration (Fig. 7 and Table H in the appendix). At wk 12, for a given concentration of added ergocalciferol, serum 25(OH)D₂ concentration also increased with increasing dietary concentrations of olestra. Similar but less clearly defined trends were present at wk 8 and 4. 

The serum 25(OH)D₂ concentration data measured for the groups fed 4.4 and 7.7% olestra were greater than control values for all concentrations of added ergocalciferol. This finding suggests that the serum 25(OH)D₂ results were confounded because of a competitive interaction between vitamin D₂ and high concentrations of vitamin D₃ resulting from the UV exposure. Such an interaction was noted in the 12-wk dose response study (Cooper et al. 1997b). In that study, serum 25(OH)D₂ concentration increased and serum 25-hydroxycholecalciferol [25(OH)D₃] concentration decreased when the dietary concentration of olestra was increased above 3.3%. Similar changes occurred in this study. Because of this interaction, a relationship describing the dependence of the restoration level on the dietary concentration of olestra was not developed for vitamin D.

At wk 12 (Table 4) and wk 8 (Table I in the Appendix), serum 25(OH)D₃ concentration decreased with increases in dietary olestra concentration. Because 25(OH)D₃ contributed >80% to the total concentration of 25-hydroxyvitamin D in the serum, the decreases in 25(OH)D₃ resulted in decreases in 25(OH)D in most groups with increasing dietary concentration of olestra (Table 4 and Table J in the Appendix).

Table 4. Serum 25-hydroxycholecalciferol [25(OH)D3], serum total 25-hydroxyvitamin D [25(OH)D] and serum 1,25 dihydroxyvitamin D [1,25(OH)2D] concentrations for pigs fed olestra and graded levels of vitamin A, D and E for 12 wk1 [View Table]

The serum concentrations of 1,25-dihydroxyvitamin D [1,25(OH)₂D] for the olestra-fed groups were not significantly different from the control value (Table 4 and Table K in the Appendix). Serum 1,25(OH)₂D concentration decreased with time in all groups.

Vitamin K. Olestra had no biologically meaningful effects on prothrombin time (PT); mean PT values measured at wk 0, 4, 8 and 12 are shown in Table 5. PT was also measured at wk 1, 2, 3, 5, 6, 7, 9, 10 and 11 for the control and 7.7% olestra groups (data not shown). At wk 10 and 11, the PT values in the three 7.7% olestra groups were prolonged relative to the control value by <0.5 s, but the differences were significant.

Table 5. Plasma prothrombin time (PT) for pigs fed olestra and graded levels of vitamins A, D and E for 12 wk1 [View Table]

Olestra did not affect the status of any of the water-soluble micronutrients, although significant differences between control and olestra-fed groups were observed in three instances. There was no dose-response effect on plasma folate concentration, although values for the 1.1 and 7.7% MV groups were significantly less than the control value at wk 8 and 12 (Table L in the Appendix). No significant differences were found between control and olestra-fed groups for the liver concentrations of vitamin B₁₂ and iron (Table M in the Appendix). Also, olestra had no significant effects on serum total iron-binding capacity (TIBC) or total iron concentration at any time point (Tables N and O in the Appendix).

Table 6. Plasma folate concentration for pigs fed olestra and graded levels of vitamins A, D and E for 12 wk1 [View Table]

Table 7. Liver vitamin B12 concentration for pigs fed olestra and graded levels of vitamins A, D and E for 12 wk1 [View Table]

No significant differences were found between control and olestra-fed groups in the liver and bone concentration of zinc (Table P in the Appendix). Relative to control, serum zinc concentrations were significantly elevated for the 1.1% LV and 1.1% HV groups at wk 8 but not wk 4 or 12 (Table Q in the Appendix).

Ash, calcium, and phosphorus concentration in bone (Table R in the Appendix) and calcium concentration in serum (Table S in the Appendix) were not significantly different than control values.

The serum concentration of inorganic phosphorus measured at wk 12 for the 4.4% LV group was significantly greater than control (Table T in the Appendix). No significant differences were found at other time points or for other dietary olestra concentrations.

Table 8. Liver iron concentration, serum total iron-binding capacity (TIBC) and serum total iron concentration for pigs fed olestra and graded levels of vitamins A, D and E for 12 wk1 [View Table]

Table 9. Liver, bone and serum zinc concentrations for pigs fed olestra and graded levels of vitamins A, D and E for 12 wk1 [View Table]

Table 10. Amounts of ash, calcium and phosphorus in bone of pigs fed olestra and graded levels of vitamins A, D and E for 12 wk1 [View Table]

Table 11. Serum calcium and inorganic phosphorus concentrations for pigs fed olestra and graded levels of vitamins A, D and E for 12 wk1 [View Table]

DISCUSSION

All pigs grew at a rate similar to that observed by others for these crossbred pigs fed standard swine diet (Martin and Crenshaw 1989). No differences were seen among the groups in digestible feed efficiency, an indication that olestra does not affect the absorption and utilization of macronutrients. This finding is consistent with observations in other pig studies (Cooper et al. 1997a and 1997b).

This study showed that the effects of olestra on the availability of the fat-soluble vitamins A, D and E can be offset over a range of dietary concentrations of olestra by providing additional amounts of the vitamins in the diet. Further, the amounts of the vitamins required are essentially linear functions of the amount of olestra in the diet. This means that tissue concentrations of these vitamins can be maintained at control concentrations when olestra-containing foods are consumed.

The addition of vitamin A to the diet produced dose-responsive increases in the liver concentration of the vitamin. The resulting concentrations encompassed the control values, allowing the relationship between the restoration level and the dietary concentration of olestra to be determined. The amount of vitamin A required to maintain liver vitamin A concentration at control concentration was essentially linear. That is, a constant amount of added vitamin A per unit amount of olestra in the diet was needed to offset the olestra effect over the dietary concentrations of olestra tested. These dietary concentrations encompass and exceed expected human intakes of olestra.

Liver and serum concentrations of vitamin E both increased in a dose-responsive manner when increasing amounts of vitamin E were added to the olestra-containing diets. The relationships between the restoration level and the dietary concentration of olestra determined from the liver data and from the serum data were the same. Like vitamin A, the amount of vitamin E required to offset the olestra effect was constant per unit amount of olestra in the diet, over the dietary concentrations of olestra tested.

Parallel and equivalent decreases in liver and serum concentrations of vitamin E in response to increasing dietary concentrations of olestra have been seen in other pig studies (Cooper et al. 1997a and 1997b). This behavior is consistent with the fact that liver stores of vitamin E are mobilized rapidly when vitamin E intake is reduced; they become depleted in parallel with the circulating pool (Bieri 1972, Bunnell et al. 1975, Horwitt et al. 1956). In this study, liver and serum concentrations of vitamin E responded in a parallel manner to addition of vitamin E to the olestra-containing diets. These observations indicate that serum vitamin E concentration is a reliable measure of vitamin E status and can be reliably used to monitor vitamin E status in humans.

Studies have shown that the concentration of vitamin E in adipose tissue of animals fed vitamin E-free diets declines less rapidly than other tissue concentrations (Bieri 1972, Machlin et al. 1979). Because of the large size and the slower response of the adipose vitamin E pool to changes in vitamin E intake, it might be hypothesized that adipose stores of the vitamin would buffer changes in other tissue stores. The changes in liver, serum and adipose concentrations of vitamin E observed in this and other studies in the pig do not support that hypothesis (Cooper et al. 1997a and 1997b).

The response of serum 25(OH)D₂ concentration to changes in dietary ergocalciferol concentration was fundamentally the same as the responses of vitamins A and E. The nature of the response of serum 25(OH)D₂ concentration to olestra intake was essentially the same as the responses noted for vitamins A and E . This was also found in humans (Schlagheck et al. 1997b) and in pigs not exposed to ultraviolet light (Cooper et al. 1997a). The serum concentrations of 25(OH)D₂ for the groups fed 1.1% olestra and graded concentrations of ergocalciferol in this study showed that serum concentration of 25(OH)D₂ could be restored. Although it was not possible to develop a relationship between the serum 25(OH)D₂ concentration and the dietary concentrations of ergocalciferol and olestra, as was done for vitamins A and E, the similarities in the responses of tissue concentrations of all of the vitamins to dietary concentrations of the vitamins and olestra suggest that a similar relationship exists between vitamin D restoration concentration and dietary olestra concentration.

The apparent interaction between vitamins D₃ and D₂, which increased serum 25(OH)D₂ concentrations, also occurred in the dose-response study in which the pigs were exposed to ultraviolet light daily throughout the study (Cooper et al. 1997b). No such interaction was observed in humans (Schlagheck et al. 1997a and 1997b) or in pigs fed up to 5.5% olestra for 26 wk without being exposed to ultraviolet light (Cooper et al. 1997a). A restoration level for vitamin D, 0.07 µg ergocalciferol/g olestra, was determined from the human data (Schlagheck et al. 1997a).

Possible reasons for the behavior of the serum concentrations of 25(OH)D₂ and 25(OH)D₃ at the higher dietary concentrations of olestra are as follows: 1 ) the competitive interaction between vitamins D₂ and D₃ for liver 25--hydroxylase and 2 ) the interference with the reabsorption of vitamin D₃ and 25(OH)D₃ secreted in the bile. Vitamin D₃ is the preferred substrate for 25--hydroxylase in pigs (Horst et al. 1982); therefore a decrease in the availability of vitamin D₃ would be expected to lead to an increase in serum 25(OH)D₂ . Both vitamin D₃ and 25(OH)D₃ are secreted in the bile and some fraction of each is reabsorbed (Arnuad et al. 1975, Avioli et al. 1967).

A decline in the half-life of serum 25(OH)D₃ concentration has been seen with long-term consumption of large amounts of dietary fiber (Batchelor and Compston 1983). It is postulated that this decline results from an interference with the enterohepatic circulation of the metabolite, which occurs when the metabolite binds to the fiber. Reductions in vitamin D status have also been seen in patients with malabsorption syndromes that involve interference with the reabsorption of biliary-derived substances (Batchelor et al. 1982, Compston et al. 1982).

Significantly lower serum 25(OH)D concentrations for the pigs fed 4.4 and 7.7% olestra reflected decreases in the contribution of 25(OH)D₃ to total 25(OH)D, which was more than 80%. These changes do not indicate an effect of olestra on overall vitamin D status. No effect of olestra on overall vitamin D status was observed in free-living human subjects consuming 18 g/d olestra for 16 wk (Koonsvitsky et al. 1997). In that study, the contribution to total vitamin D status coming from the diet was ~10-15%, a contribution typical of individuals living in northern latitudes in winter (Jones 1978, Jones et al. 1991). Olestra also did not affect overall vitamin D status in subjects consuming 20 g/d olestra along with a 20-µg daily capsule of ergocalciferol for 6 wk (Jones et al. 1991) or in subjects consuming up to 32 g/d olestra for 8 wk (Schlagheck et al. 1997b).

The decrease in serum 1,25(OH)₂D concentration that was seen in all groups in this study is consistent with an age effect observed in pigs by others (Horst and Littledike 1982). The same effect has been seen in other pig studies (Cooper et al. 1997a and 1997b).

Results of this study, which showed that olestra did not affect vitamin K status, as measured by PT, or the absorption of water-soluble nutrients, are in agreement with findings from other studies in the pig (Cooper et al. 1997a and 1997b) and studies in humans (Koonsvitsky et al. 1997, Schlagheck et al. 1997a and 1997b).

The few statistical effects noted in the measures of status of the water-soluble nutrients were not dose responsive or consistent over time, indicating that they resulted from normal biological variability. Significant differences observed in the 12-wk dose-response study in liver vitamin B₁₂ concentration and bone ash content were not seen in this study, supporting the interpretation that those effects were probably related to the poor vitamin A and E status of the pigs. In this study, vitamin A and E status was maintained by the addition of extra amounts of those vitamins to the diet.

This study illustrates that the effect of olestra on the absorption of fat-soluble vitamins can be effectively offset by adding extra amounts of the vitamins to the diet. Importantly, the amounts of vitamins required are predictable and are constant per unit mass of olestra in the diet. This was demonstrated for vitamins A and E and is also true for vitamin D, although that relationship was not specifically demonstrated in this study. This conclusion can be drawn for vitamin D because the responses of the circulating concentration of 25-hydroxyvitamin D to olestra and to added vitamin D are the same as the responses noted for the other vitamins, an indication that the interactions that occur between olestra and fat-soluble vitamins are fundamentally the same for all of the vitamins.

Further evidence that olestra does not affect vitamin K functional status, the status of water-soluble nutrients or the absorption and digestion of macronutrients is provided by this study.

ACKNOWLEDGMENTS