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© 2007 American Society for Nutrition J. Nutr. 137:945-952, April 2007

Nutrient Physiology, Metabolism, and Nutrient-Nutrient Interactions

Decreasing Linoleic Acid with Constant {alpha}-Linolenic Acid in Dietary Fats Increases (n-3) Eicosapentaenoic Acid in Plasma Phospholipids in Healthy Men1

Y. Angela Liou, D. Janette King, Deborah Zibrik and Sheila M. Innis*

The Nutrition Research Program, Child and Family Research Institute, University of British Columbia, Vancouver, Canada V52 4H40

* To whom correspondence should be addressed. E-mail: sinnis{at}interchange.ubc.ca.


   ABSTRACT
TOP
 ABSTRACT
Introduction
 Subjects and Methods
 Results
 Discussion
 LITERATURE CITED
 
High linoleic acid (LA) intakes have been suggested to reduce {alpha}-linolenic acid [ALA, 18:3(n-3)] metabolism to eicosapentaenoic acid [EPA, 20:5(n-3)] and docosahexaenoic acid [DHA, 22:6(n-3)], and favor high arachidonic acid [ARA, 20:4(n-6)]. We used a randomized cross-over study with men (n = 22) to compare the effect of replacing vegetable oils high in LA with oils low in LA in foods, while maintaining constant ALA, for 4 wk each, on plasma (n-3) fatty acids. Nonvegetable sources of fat, except fish and seafoods, were unrestricted. We determined plasma phospholipid fatty acids at wk 0, 2, 4, 6, and 8, and triglycerides, cholesterol, serum CRP, and IL-6, and platelet aggregation at wk 0, 4, and 8. LA and ALA intakes were 3.8 ± 0.12% and 1.0 ± 0.05%, and 10.5 ± 0.53% and 1.1 ± 0.06% energy with LA:ALA ratios of 4:0 and 10:1 during the low and high LA diets, respectively. The plasma phospholipid LA was higher and EPA was lower during the high than during the low LA diet period (P < 0.001), but DHA declined over the 8-wk period (r = –0.425, P < 0.001). The plasma phospholipid ARA:EPA ratios were (mean ± SEM) 20.7 ± 1.52 and 12.9 ± 1.01 after 4 wk consuming the high or low LA diets, respectively (P < 0.001); LA was inversely associated with EPA (r = –0.729, P < 0.001) but positively associated with ARA:EPA (r = 0.432, P < 0.001). LA intake did not influence ALA, ARA, DPA, DHA, or total, LDL or HDL cholesterol, CRP or IL-6, or platelet aggregation. In conclusion, high LA intakes decrease plasma phospholipid EPA and increase the ARA:EPA ratio, but do not favor higher ARA.


   Introduction
TOP
 ABSTRACT
 Introduction
Subjects and Methods
 Results
 Discussion
 LITERATURE CITED
 
Clinical and epidemiological studies provide evidence that, when compared with dietary saturated fatty acids, the consumption of linoleic acid [LA,2 18:2(n-6)] results in a lower plasma total and LDL cholesterol (1,2). In addition to its effects in modifying plasma lipid risk factors for cardiovascular disease (CVD), LA is a dietary essential fatty acid and is converted through desaturation and elongation to arachidonic acid [ARA, 20:4(n-6)], which is essential in growth, cell signaling, and as an eicosanoid precursor (3). Dietary intakes of LA have increased over the last century from ~3% of energy in the early 1900s and now contributes ~85–90% of total polyunsaturated fatty acids and 5–7% of dietary energy in the U.S., Canada, and many westernized nations due largely to an increased consumption of LA-rich vegetable oils (4–7). The dietary intake of LA required to prevent biochemical and clinical signs of (n-6) fatty acid deficiency, however, is only ~2% total energy (8), and in rodents maximum tissue levels of ARA are achieved at intakes of ~2.8% dietary energy from LA (9).

Recent studies emphasized the favorable effects of (n-3) fatty acids in reducing the risk of CVD, possibly involving multiple mechanisms including eicosanoid metabolism, inflammatory mediators, platelet aggregability, hemostatic and myocardial function, and plasma lipids (10–18). The (n-3) fatty acids are present in the diet as {alpha}-linolenic acid [ALA; 18:3(n-3)], eicosapentaenoic acid [EPA; 20:5(n-3)] and docosahexaenoic acid [DHA; 22:6(n-3)]. ALA is found in some vegetable oils such as soybean, canola, and flax seed oil, but many oils such as corn, safflower, sunflower, olive, and hydrogenated fats contain <1.0% fatty acids as ALA. EPA and DHA are present in the diet in animal tissue lipids, with the richest dietary source being fatty fish. Some studies suggested that diets high in ALA, as well as diets high in EPA and DHA are associated with lower levels of proinflammatory and higher levels of anti-inflammatory cytokines, although other studies found no effects of dietary (n-3) fatty acids on inflammatory mediators (19–22). In contrast, although eicosanoids derived from ARA are generally considered proinflammatory (10), ARA-derived metabolites may also have important anti-inflammatory actions (23), and, in epidemiological studies, higher plasma levels of both ARA and (n-3) fatty acids were associated with the lowest level of inflammatory markers (19,22).

The first enzymatic step in the desaturation of LA and ALA involves desaturation by the microsomal {Delta}6 desaturase (3,8). High intakes of LA have been suggested to decrease the desaturation of ALA to EPA and DHA and favor higher ARA through competition for the {Delta}6 desaturase (21,24). Supplementation with EPA and DHA from fish or fish oils, on the other hand, results in reduced tissue and plasma ARA, and increased blood lipid EPA and DHA (25–27). Our objective was to determine whether replacing vegetable oils and fats in bakery and snack foods, margarines, and cooking and salad oils with oils low in LA, compared with oils high in LA, increases (n-3) fatty acids and lowers (n-6) fatty acids in plasma lipids of adult men. To address this objective, we prepared foods using high or low LA oils to provide ~55% of total dietary fat intakes, and a total dietary intake of 3.8 or 10.5% energy from LA, with a constant intake of 1% energy ALA. To achieve a lower LA, we used oils with oleic acid [OA,18:1(n-9)], thus keeping saturated fatty acids constant. Fish and seafood were avoided during the study to minimize confounding effects of a dietary intake of preformed EPA and DHA.


   Subjects and Methods
TOP
 ABSTRACT
 Introduction
 Subjects and Methods
Results
 Discussion
 LITERATURE CITED
 
    Subjects. Eligible subjects were nonvegetarian, nonsmoking men, 20–45 y of age, BMI 18.5–29.9, and not known to have hypertension, hyperlipidemia, glucose intolerance, diabetes, or any other disease likely to affect lipid metabolism, or to consume any fatty acid, lipid or antioxidant supplements, or medications likely to interfere with this study. Twenty-four subjects were enrolled, 2 of whom withdrew in the 1st 28 d of the study. The study was approved by the Clinical Research Ethics Board of the University of British Columbia, and the Research Review Committee of the Children's and Women's Heath Center of British Columbia. Informed written consent was obtained from all subjects.

    Study design. This was a 10-wk study involving a 2-wk prestudy phase, in which fish and seafood were avoided, followed by a randomized cross-over, in which the intake of LA from vegetable oils was altered without modifying the intake of fats from meat, poultry, or dairy foods. The subjects consumed foods prepared with oils high LA or low in LA, with constant ALA, for 4 wk for each diet (high LA–low LA and low LA–high LA). Our objective was to maintain a constant dietary intake of ~1% energy from ALA with either high or low LA, to provide a total dietary LA:ALA ratio of ~4:1 or 10:1 by modifying only the intake of LA from vegetable oils and fats. Changes in LA intake were achieved through the use of oils with high or low in LA and OA but with similar saturated fatty acids and ALA. Meat, poultry, eggs, dairy products, fruits and vegetables, cereals and grains and all foods containing no fat were unrestricted throughout the study. The subjects were requested to consume no salad or cooking oils, mayonnaise, salad dressings, sauces, bakery foods, desert, or snack foods other than those provided during the 8-wk cross-over portion of the study. Information sessions were given at the beginning of the study and then prior to each 4-wk period to assist subjects in adhering to the protocol and avoiding foods containing vegetable fats. Each subject attended the Nutrition Research Program each week, at which time they were given foods for the following week, and dietary records from the previous week were collected and reviewed.

    Food development and preparation. Because of the potential for oxidative and thermolytic changes in high ALA oils during frying (28), we prepared baked foods, mayonnaises, and salad dressings containing low or high LA, with similar ALA such that 3 servings/d would result in a total dietary intake of ~1% of dietary energy as ALA. We derived this by extrapolation of the probable intake of ALA from all food sources other than vegetable oils, fish, and seafood. An oil blend containing 90 parts high oleic safflower oil or 90 parts high linoleic sunflower oil with 10 parts flax seed oil (Flax Canada 2015) was prepared and contained (g/100 g fatty acids) 13.7% LA and 6.5% ALA, or 56.5% LA and 6.9% ALA, with LA:ALA ratios of 2.1:1 and 8.2:1, respectively, for the high and low LA oil blends (Table 1). OA was 69.5 and 24.2% fatty acids in the high and low LA blends, respectively. Then, recipes for cookies, savory and sweet breads, cakes, deserts, and snacks were developed using the oil blends, and the stability of LA and ALA during preparation and storage was verified by GLC and GLC-MS/MS (29). Foods were prepared weekly in our nutrition research kitchen, wrapped in preweighed individual portions, and frozen. Subjects selected foods from weekly production lists. Mayonnaises and salad dressings were prepared using the high and low LA oil blends (Table 1) in commercial facilities through collaboration with English Bay Baking, Delta, British Columbia. The subjects were given high OA safflower oil, olive oil, and Becel with olive oil margarine during the low LA diet and sunflower oil and Fleischmann's margarine during the high LA diet period (Unilever Canada) for home use.


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  		TABLE 1 Fatty acid composition of oil blends and margarines

 
    Dietary analysis. Dietary intakes were estimated using three 24-h food records, kept at least 4 d apart, during the two 4-wk study periods. Data from the food records was entered into Food Processor (version 8.2, ESHA Research) and analyzed for total fat and fatty acid intakes using the Canadian Nutrient file and recipes used in this study (30).

    Blood samples and analyses. Venous blood was collected into 7 mL tubes containing EDTA or sodium citrate as the anticoagulant, or clotting tubes (BD Vacutainer Systems, Becton Dickinson) after an overnight fast at the beginning of the study, and again every 2 wk throughout the study. Plasma and serum were separated by centrifugation at 2000 x g for 15 min at 4°C, then made into aliquots and frozen at –70°C for later analyses.

Plasma lipids were extracted and phospholipids and triacylglceyrols separated using thin-layer chromatography. The fatty acids were converted to their respective methyl esters prior to separation and quantified using GLC (29). Plasma total and HDL cholesterol and triacylglycerols were analyzed using enzymatic reagents (Diagnostic Chemicals) (31). LDL cholesterol was calculated using the Friedewald equation (32). Serum C-reactive protein (CRP) and IL-6 were determined using high sensitivity, commercial immunoassays (American Laboratory Products, and QuantiKine, R and D Systems, respectively). The interassay and intra-assay variability for CRP were 11.7 and 2.7% and for IL-6 were 12.0 and 5.7%, respectively. Platelet aggregation was assayed with a Dade Platelet Function Analyzer (PFA 100, Dade Behring) with ADP as the agonist, according to the manufacturer's instructions.

    Statistical analysis. Values are expressed as means ± SEM. The potential effects of the order in which the diets were consumed were tested before considering the effects of the high compared with low LA diet by ANOVA (33). A significant effect for order (the order in which the low and high LA diets were consumed) was not found for any of the fatty acids except ARA in the 2nd 4-wk period. Therefore, the results of ARA for the 1st 4-wk period only were used to test for differences between the high and low LA diets, with a priori contrasts to determine differences in the fatty acids of interest at the end of each 4-wk period. Paired t tests were used to determine differences within the diet groups at the end of the two 4-wk diet periods. Tukey's test was used to determine differences due to time within a diet group, excluding the baseline (wk 0) value, which was not part of the controlled 8-wk diet period. Linear regression was used to assess the potential relation between the plasma phospholipid LA and EPA, the ARA:EPA ratio, and to assess the change in the plasma phospholipid DHA during the study. All statistical analyses were performed using SPSS software (version 13.0 for Windows). Significance was determined at P < 0.05.


   Results
TOP
 ABSTRACT
 Introduction
 Subjects and Methods
 Results
Discussion
 LITERATURE CITED
 
    Characteristics of study subjects. The mean age, BMI, fasting plasma lipids, glucose or phospholipid fatty acids (Table 2), or triglycerides (data not shown) did not differ among the men randomized to 2 study groups. A blood sample was not collected from 1 subject who was randomized to the high LA–low LA group on wk 8, the last day of the study.


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  		TABLE 2 Characteristics of study subjects1

 
    Dietary fatty acid intakes. During the study period, the mean fat intake from all sources was 32.5% total energy (Table 3). The foods provided contributed ~55% of the total fat intake. By modifying only the vegetable fat portion of the diet, we achieved an intake of 3.8 ± 0.12 and 10.5 ± 0.53% total energy from LA, with an LA:ALA ratio of 4:1 and 10:1 during the high and low LA diet periods, respectively. In addition, there was no overlap in the range of LA intake among the men in the low LA (2.9–5.2% total energy) and high LA (7.0–15.8% energy) diet periods, showing that the dietary intake of LA was modified for all subjects in our study. Foods prepared with flax oil contributed ~70% of the total intake of ALA during the study and resulted in total intake of ~1% energy from ALA throughout the study. Because we exchanged oils high in LA oil with oils low in LA and high in OA, the intake of OA also differed between the high and low LA diet periods, although the intake of saturated fatty acids was not different.


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  		TABLE 3 Dietary fat and fatty acid intakes during the high and low linoleic diet periods1

 
    Plasma lipid fatty acids. The fasting plasma phospholipid levels of total saturated fatty acids, monounsaturated fatty acids, (n-6) and (n-3) fatty acids, and the ARA:EPA ratio after consuming the high and low LA diets for 4 wk were measured and recorded (Table 4). Because an order effect was found for ARA, a diet x time interaction was found for ALA, and a time effect was found for DHA. Time effect results showed the levels of the major (n-6) and (n-3) fatty acids for the 2 groups of men (high LA–low LA and low LA–high LA), rather than the means for each of the high and low dietary LA periods (Fig. 1). The plasma phospholipid total saturated fatty acids did not differ after 4 wk of consuming the high and low LA diets, but the total monounsaturated fatty acids, (n-3) fatty acids, and ARA:EPA ratio were higher, and the total (n-6) fatty acids was lower in the men after 4 wk of consuming the low LA compared with the high LA diet (Table 4). The plasma phospholipid LA and EPA after 4 wk of consuming the high (n = 22) or low LA diet (n = 21) were 29.4 ± 0.78 and 23.7 ± 0.67 (P < 0.001) and 0.58 ± 0.05 and 0.93 ± 0.07 g/100 total fatty acids (P < 0.0001), respectively. Notably, the plasma phospholipid level of LA increased and EPA decreased during the 1st 2 wk of consuming the high LA diet, and LA decreased and EPA increased between 0 and 2 wk of consuming the low LA diet, with no further changes in these fatty acids with a continuation of the diet periods to wk 4. Neither the plasma phospholipid level of ARA nor docosapentaenoic acid [DPA, 22:5(n-6)] changed as a result of the change attributable to the change in dietary LA (Fig. 1A). We found a diet x time interaction for the plasma phospholipid ALA (Fig. 1B). Those men who consumed the low LA diet in the first 4 wk had a significant increase in their plasma phospholipid ALA level at wk 2, then a decrease in ALA at wk 4, whereas those men who consumed the low LA diet in the second 4 wk period had a higher plasma phospholipid ALA after 4 wk consuming the high LA diet (Fig. 1B). Neither the low nor high LA diet affected plasma phospholipid (n-3) DPA; the plasma phospholipid (n-3) DPA was 1.18 ± 0.09 and 0.94 ± 0.05 g/100 g fatty acids at wk 4 of the study among those who consumed the high and low LA diets, respectively, and 1.07 ± 0.05 and 1.02 ± 1.04 g/100 g after 4 wk on the cross-over to the low and high LA diet, respectively (P > 0.05). In contrast to EPA, the plasma phospholipid level of DHA did not differ between the low and high LA diet periods, but did decrease in both groups of men throughout the study. Linear regression analysis showed that the plasma phospholipid DHA decreased (r = –0.423, P < 0.0001) at a rate of 0.104 g/(100 g plasma phospholipid fatty acids · wk).


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  		TABLE 4 Fasting plasma phospholipid fatty acids in adult men after consuming low LA then high LA diets, or vice versa, each for 28 d1

 

Figure 1
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  		Figure 1  Plasma phospholipid major (n-6) (A) and (n-3) (B) fatty acids in men consuming low and high LA diets in a random cross-over design. Values are means ± SEM, n = 10–12. Significant effect of 2diet, 3diet x time, and 4time, P < 0.05 (by 2-way ANOVA); a,bdifference within a group due to time (Tukey's test); {dagger},§ difference between the low and high LA diet, P < 0.05.

 
The plasma phospholipid level of LA was inversely associated with EPA at each time point (data not shown), and using the results for all time points and subjects (r = –0.729, P < 0.0001, CI –0.073 to –0.051, n = 109) (Fig. 2). We also found an association between the plasma phospholipid level of LA and the ARA:EPA ratio (r = 0.432, P < 0.0001, CI 0.487 to 1.130, n = 109) (Fig. 2).


Figure 2
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  		Figure 2  Scatterplots of the associations between the plasma phospholipid LA and EPA (left) and LA and ARA:EPA ratio (right) in men who consumed low and high LA diets in a random cross-over design. Fatty acids are expressed as g/100 g total plasma phospholipid fatty acids. The 5th–95th CI are shown.

 
Plasma triacylglycerol fatty acids had higher LA and lower EPA after 4 wk of consuming the high compared with the low LA diet, with 17.4 ± 2.11 and 22.5 ± 2.08 g/100 g LA and 0.16 ± 0.02 and 0.13 ± 0.02 g/100 EPA at wk 4 and 8, respectively, in the men randomized to the low LA–high LA group (n = 12), and 24.0 ± 2.23 and 16.0 ± 1.21 g/100 g LA and 0.17 ± 0.03 and 0.29 ± 0.10 g/100 EPA at wk 4 and 8, respectively, in the men randomized to the high LA–low LA group, (n = 10). Also, as found in the plasma phospholipids, the plasma triacylglycerol level of DHA decreased by ~20% over the 8-wk study period, with no effect of dietary LA (data not shown).

    Plasma lipids, inflammatory markers, and platelet aggregation. After 4 wk of consuming the low (n = 21) and the high LA (n = 22) diets, there was no difference in plasma total cholesterol (4.05 ± 0.18 and 3.94 ± 0.18 mmol/L), HDL cholesterol (1.32 ± 0.65 and 1.31+0.067 mmol/L), LDL cholesterol (2.29 ± 0.16 and 2.33 ± 0.16 mmol/L), LDL:HDL cholesterol ratio (1.84 ± 0.17 and 1.89 ± 0.19), or triacylglycerols (0.96 ± 0.17 and 0.65 ± 0.08 mmol/L). Inspection of the results for the plasma triacylglycerols, however, suggested that men with a fasting plasma triacylglycerol >0.90 mmol/L after 4 wk of consuming the low LA diet had a lower triacylglycerol after 4 wk consuming the high LA diet, 1.62 ± 0.28 and 0.88 ± 0.16 mmol/L, respectively, n = 9, with no effect of diet order. The fasting plasma triacylglycerols in the other men were 0.47 ± 0.04 and 0.48 ± 0.053 mmol/L after 4 wk on the low and high LA diets, respectively, n = 12. The serum CRP was 0.56 ± 0.15 and 0.60 ± 0.21 ng/L (n = 19) and IL-6 was 0.96 ± 0.33 and 0.93 ± 0.30 ng/L (n = 21) after 4 wk on the high LA diet (n = 22) or low LA diet (n = 21), respectively, P > 0.05. Results for CRP were below the limit of detection, or were >3 SD above the mean for n = 2 subjects. Platelet aggregation in response to ADP did not differ after 4 wk of consuming the low (81 ± 1.63) compared with high (85 ± 2.51) LA diets.


   Discussion
TOP
 ABSTRACT
 Introduction
 Subjects and Methods
 Results
 Discussion
LITERATURE CITED
 
This study demonstrates that changing the fats and oils in the diets of adult men to achieve an intake of 3.8% energy from LA while maintaining an intake of 1% energy from ALA results in higher EPA and higher EPA:ARA ratio in plasma phospholipids than a LA intake of 10.5% energy. The study was designed to address the effects of lowering the dietary (n-6):(n-3) fatty acid ratio by reducing the intake of LA and increasing OA from vegetable oils while maintaining a constant intake of 1% energy from ALA. To reduce the possibility of low ALA intakes, we prepared an oil blend containing 10% flax seed oil with 90% (v:v) of a high OA or high LA oil such that 2–3 servings representing 5 g fat or 30 g baked food servings would, in addition to the intake of ALA from dairy foods, eggs, and meats, achieve a mean intake of ~1% dietary energy from ALA. An intake of 1% energy from ALA is about double the current mean ALA intake in North America (4). Using this approach, and without restricting the intake of meat, poultry, eggs, or dairy foods, we provided ~55% of the total fat intake with the result that LA intakes were 2.9–5.2 and 7.8–17.2% total energy among the men during the low and high LA diet periods, respectively. The results provide a clear demonstration that lowering dietary LA intake increases EPA in plasma phospholipids. Notably, our results also show that although an intake of 10.5% energy from LA with a LA:ALA ratio of 10:1 resulted in lower plasma phospholipid levels of EPA, it did not favor higher levels of ARA compared with an intake of 3.8% energy from LA with a LA:ALA ratio of 4:1.

Previous reports suggest that high dietary intakes of LA lead to reduced desaturation of ALA due to competition between LA and ALA for {Delta}6 desaturase, thus favoring increased tissue ARA and reduced levels of the longer chain (n-3) fatty acids (21,24). The current ratio of (n-6):(n-3) fatty acids in many western diets is in excess of 10:1, and a ratio of 4:1 or lower has been suggested as optimal for human health (21,24). Our results, which show a higher plasma phospholipid EPA when the dietary LA:ALA ratio was 4:1 rather than 10:1, are consistent with the hypothesis that a high dietary LA inhibits the desaturation of ALA. An effect of high dietary LA in reducing plasma phospholipid EPA has been reported previously (34). In addition, we found a strong inverse association between the plasma phospholipid LA and EPA that predicted that for every 10% increase in LA, EPA decreased by 0.64 g/100 g fatty acids. The association between LA and EPA showed no evidence of a plateau at lower levels of LA intake, which suggests that additional lowering of LA would result in further increases in the plasma phospholipid EPA.

Consistent with the results of our study, the results of several other studies also show no increase in plasma or blood cell lipid ARA, regardless of changes in LA intake (22,26,34,35). The consistently high levels of LA and ARA in plasma phospholipids, with LA:ALA ratios >60:1, even when the dietary LA intake was reduced to 3.8% energy in an LA:ALA ratio of 4:1 (Fig. 1), suggests possible differences in the metabolic handling of LA and ALA. Early in vitro studies, reported that {Delta}6 desaturase shows strong substrate preference in the order of ALA > LA > OA at saturating substrate concentrations, and that ALA and LA are mutually competitive (36). However, the intracellular substrate concentrations of LA and ALA are below enzyme saturation, and considerably lower for ALA than LA (37). In addition, ALA appears to be rapidly ß-oxidized, perhaps due to activation by acyl CoA synthase in the mitochondria (38,39). It is possible that preferential channeling of LA toward acylation and desaturation rather than ß-oxidation may explain the results of these and other studies to show high LA and ARA in plasma lipids, regardless of a low intake of LA (22,26,34,35) (Fig. 1). As in studies of rodents (9), our results in humans suggest that the conversion of LA to ARA and incorporation of ARA into tissue lipids is already maximal at intakes of ~3.8% energy LA. Competition among (n-6) and (n-3) fatty acids for acylation could also be important. In this regard, several studies showed that the incorporation of dietary EPA and DHA into tissue and blood lipids is lower during high LA intakes (26,39,40). Similarly, studies using a stable isotope tracer also suggest that (n-6) fatty acids may compete with (n-3) fatty acids at the level of incorporation in to acylated lipids, rather than as inhibitors of ALA conversion (41).

Several additional important points arise from the present study. Although the plasma phospholipid EPA levels decreased or increased within 2 wk of consuming the high or low LA diets, respectively, DHA declined throughout the study. The decrease in the plasma phospholipid DHA from 3.9 at wk 0 to 3.0 g/100 g fatty acids at wk 8 of the study represented a decrease (P < 0.0001) in DHA at a rate of 0.104 g/100 g plasma phospholipid fatty acids per week. Studies in vegans report 1.4% DHA in plasma phospholipid fatty acids (42), which is considerably lower the level than attained during our 8-wk study. On the other hand, 1% dietary energy from ALA in an LA:ALA ratio of 4:1 clearly does not maintain a plasma phospholipid level of DHA as high as that achieved by a mixed diet that includes fish. Stable isotopic tracer studies show that the conversion ALA to DHA, particularly at the step of EPA to DHA is very low in humans (43–45). However, whereas the decrease in plasma phospholipid levels of DHA in our study is reasonably explained by a low intake of preformed DHA and low rates of incorporating DHA derived from ALA into plasma lipids, the plasma phospholipid EPA levels increased when the dietary LA was decreased (Fig. 1A, B). Studies showing that supplementation with ALA or stearadonic acid [18:4(n-3)] increases blood lipid EPA with little or no increase DHA in humans (46–50), also show that (n-3) fatty acid metabolism is limited beyond EPA. However, in our study, high dietary LA did not reduce DHA in plasma phospholipids, which is consistent with the results of stable isotope tracer studies that found no evidence that high LA interferes with DHA synthesis in animals (41). However, we note that steady-state levels of DHA were not achieved during our study.

DPA [22:5(n-6)] is the product of elongation, {Delta}6 desaturation, and chain shortening of ARA and is increased in animals fed (n-3) fatty acid–deficient diet (3,8). An increase in DPA did not occur with the decrease in plasma phospholipid DHA in the men in our study. Possibly, 1% energy from ALA meets the requirements for (n-3) fatty acids in humans, or limited elongation of carbon chain 22 (n-6) and (n-3) fatty acids and {Delta}6 desaturation of 24:4(n-6) and 24:5(n-3) may both be low in humans (51). Changes in serum CRP, IL-6, or platelet aggregation were not attributable to changes in dietary LA or the LA:ALA ratio, although the subjects in our study were not at risk for disease, and values for these measures were within normal ranges.

In summary, our study indicates that a diet with 1% energy from ALA and a LA:ALA ratio of 4:1 results in higher plasma phospholipid EPA and a 40% lower ARA:EPA ratio than a diet containing a LA:ALA ratio of 10:1, with no change in the plasma phospholipid the level of ARA. Interestingly, whereas the plasma phospholipid ARA:EPA ratio in the men at the start of our study was 12.5 ± 0.08, the ratios after 4 wk of consuming the high LA or low LA diet were 20.7 ± 1.52 and 12.9 ± 1.01, respectively. Several recent studies suggested that diets high in ALA may be important in lowering the risk of CVD (52–54), and recent studies suggested that ARA-derived eicosanoids also have important anti-inflammatory actions (19,22). A notable difference between diets high in ALA with a low LA:ALA ratio and diets high in EPA and DHA is that, whereas dietary EPA and DHA reduce blood lipid ARA, no such effect occurred in our study, which involved relatively simple changes in LA intake. Future studies need to address the relevance of modifying ARA:EPA ratio with and without changing ARA with respect to disease prevention and in individuals with pre-existing disease.


   ACKNOWLEDGMENTS
 
We acknowledge with thanks to R. Esdaile, J. Pamar, and L. Yeung for their assistance in food preparation and in the conduct of this study. R. W. Friesen assisted with preparation of the mayonnaise and salad dressings and with blood samples, and A. Lee provided assistance with the analysis of foods. We gratefully acknowledge the support of our biostatistical expert, R. A. Milner in the data analysis.


   FOOTNOTES
 
1 This study was supported by grants from the Canadian Foundation for Dietetic Research and Flax Canada 2015. Y.A.L. was supported by a student stipend from the Molly Towell Perinatal Research Foundation and S.M.I. was supported by a distinguished scholar award from the Michael Smith Foundation for Health Research. Back

2 Abbreviations used: ALA, {alpha}-linolenic acid; ARA, arachidonic acid; CVD, cardiovascular disease; DHA, docosahexaenoic acid; DPA, docosapentaenoic acid; EPA, eicosapentaenoic acid; LA, linoleic acid; OA, oleic acid. Back

Manuscript received 23 June 2006. Initial review completed 9 August 2006. Revision accepted 24 January 2007.


   LITERATURE CITED
TOP
 ABSTRACT
 Introduction
 Subjects and Methods
 Results
 Discussion
 LITERATURE CITED
 

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