Bioavailability of long-chain omega-3 polyunsaturated fatty acid enriched luncheon meats.

Abstract

Objective: To determine the acute and chronic effects of low doses of long chain (LC) n-3 polyunsaturated fatty acids (PUFA) on plasma LC n-3 PUFA levels.

Design: In the acute study, six healthy omnivores, avoiding fish meals on the day prior to the study, provided a fasting blood sample initially and post prandially at four hours. In the chronic study, 12 healthy subjects provided a fasting blood sample at baseline and three weeks after daily consumption of the test food.

Main outcome measures: Plasma non-esterified fatty acid and phospholipid LC n-3 PUFA composition.

Statistical analysis: Differences in plasma non-esterified fatty acids and phospholipid LC n-3 PUFA. A pre- and post-consumption of the test food were assessed using paired t-tests (SPSS).

Results: The acute study demonstrated that a low dose of LC n-3 PUFA (25% eicosapentaenoic acid and 75% docosahexaenoic acid) significantly increased eicosapentaenoic acid levels in plasma of human subjects postprandially from 0.30% to 0.42% of total non-esterified fatty acids, a per cent change of 39% (P < 0.05). The chronic study demonstrated a significant increase in total n-3 phospholipids from zero weeks (5.48% of total fatty acids) to three weeks (7.92% of total fatty acids), representing a per cent increase of 44% (P = 0.006). The n-6 to n-3 ratio of LC PUFA phospholipids demonstrated a significant reduction from 5.1 at zero weeks to 4.07 at three weeks, representing a reduction of 20% (P = 0.006).

Conclusions: These findings demonstrate the bioavailability of LC n-3 PUFA consumed as a low-fat omega-3-enriched luncheon meat.

Key words: docosahexaenoic acid, docosapenaetnoic acid, eicosapentaenoic acid, omega-3 fatty acids 

(Nutr Diet 2005; 62: 130-137)

Introduction

There is considerable interest in the role of n-3 polyunsaturated fatty acids (PUFA) in human health and in growth and development. (1-3) A number of key studies conducted over the past decade have shown the potential benefits of foods containing n-3 fatty acids. (2,4-9) The consumption of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) results in a decrease in the amount of arachidonic acid (AA) in the membrane phospholipids of cells involved in inflammation and immunity. In addition, long-chain (LC) n-3 PUFA appear to inhibit the release of AA. This results in a decreased capacity of immune cells to synthesise pro-inflammatory eicosanoids from AA. (10) Furthermore, n-3 fatty acids increase bleeding time by decreasing platelet aggregation, blood viscosity and fibrinogen, thus decreasing the tendency of thrombus formation. (9,11) Consequently, increasing the consumption of n-3 PUFA reduces the risk for heart disease and the severity of symptoms of rheumatoid arthritis and other inflammatory diseases. (9,11)

In a study that compared a seven-day weighed food record with plasma fatty acid concentrations, Sinclair et al. found that the Australian diet contained low levels of LC n-3 PUFA; this diet has been estimated to be between 100 and 190 mg/day for adults, which is much lower than the dietary intake of 650 mg/day recommended by the International Society for the Study of Fatty Acids and Lipids (ISSFAL). (2,12) Ollis et al. found that the major food sources of n-6 PUFA in the Australian diet were polyunsaturated margarines, some nuts/seeds, bread, snacks, desserts and takeaways, whereas canola oil and margarine were rich sources of the n-3 PUFA, [alpha]-linolenic acid (ALA). (13) The recommendation made by ISSFAL indicates 'The increase of ALA, together with EPA and DHA (all n-3 PUFA), and reduction of vegetable oils with high n-6 PUFA, linoleic acid (LA) content, are necessary to achieve a healthier diet in Western countries'. (2) However, some common n-3 PUFA food sources may not be suitable for individuals of differing cultural, religious or ethnic backgrounds. Thus, it may be beneficial to incorporate LC n-3 PUFA into some frequently used foods, for example small goods, bread, milk and eggs. There are a number of these LC n-3 PUFA-enriched functional foods available in Australia, for example Tip Top Up Omega-3 white bread (Tip-Top, Chatswood, NSW, Australia), omega-3-enriched Body Eggs (Chanteclair, Blacktown, NSW, Australia), Heart Plus omega-3-enriched milk (Browns Dairy, Balcatta, WA, Australia), Meadow Lea Hi-Omega omega-3-enriched margarine (Goodman Fielder, Ryde, NSW, Australia).

Studies have been published examining the ability of n-3-enriched foods to influence plasma lipids in healthy humans, for example, omega-3 PUFA-enriched margarine, (6) omega-3 PUFA-enriched eggs, (14) omega-3 PUFA-enriched bread (15) and omega-3 PUFA-enriched milk. (7) These studies have used foods containing 300-910 mg LC n-3 PUFA per serving. The present study uses a lower level of omega-3 PUFA enrichment, which could provide a more practical approach by avoiding extensive changes to habitual diets. (15) This is the first study to provide bioavailability information on the effect of low-dose LC n-3 PUFA in processed foods.

The hypothesis of the present study is that the consumption of a low-dose LC n-3 PUFA in 190 g of enriched small goods will increase the LC n-3 PUFA status of human subjects. The aim of the present study was to observe the effects of a single serve of luncheon meat enriched with ~190 mg of LC n-3 PUFA postprandially (the acute study) and the effect of a daily serve of luncheon meat enriched with ~190 mg of LC n-3 PUFA for three weeks (the chronic study) on plasma LC n-3 PUFA status in human subjects.

Methods

The dietary intervention study consisted of two parts, a four-hour acute study and a three-week chronic study. The present study aimed to determine the short-term and long-term effects of consumption of LC n-3 PUFA-enriched luncheon meats on the plasma fatty acid and plasma phospholipid status of human subjects.

Subjects

In total, 18 healthy free-living omnivores (an omnivore was defined as someone who eats meat at least five times a week), 15 females and 3 males were recruited among students and staff at the University of Sydney. In total, 5 females and 1 male participated in the acute study, and 10 females and 2 males were in the chronic study. Participant numbers were determined by checking the literature. The exclusion criteria were hyperlipidaemia, hypertension, atherosclerosis, non-alcoholic steatohepatitis, hepatitis, acute and chronic renal failure, type 1 and type 2 diabetes mellitus, high alcohol consumption (more than 10 standard drinks per week) and nicotine use. Subjects were excluded if they had a history of dieting. This was ascertained by questions on restrained eating, following a diet that restricted food choice and fasting. Subjects who had taken dietary fatty acid supplements and medications for heart disease, lipid lowering and diabetes were also excluded. All of the volunteers successfully completed the study.

All subjects gave their written consent before participating in the present study. The study protocol was approved by the Human Research Ethics Committee, the University of Sydney.

Study design and diet

In the acute study, six subjects were advised to follow their habitual diet for one week prior to the study day. One day before the study, foods rich in n-3 and n-6 PUFA (e.g. seafood, nuts, peanut butter, polyunsaturated margarines, soy and linseed bread) were excluded. On the study day, the subjects recorded all the foods consumed on the previous day. They were given the LC n-3 PUFA-enriched luncheon meat serving as a sandwich with monounsaturated margarine, tea, coffee, or juice. Subjects consumed this meal following the initial fasting blood collection. They were also requested to avoid strenuous physical exercise and to limit their food intake over the next four hours to water, zero calorie drinks and/or an apple. They remained in the testing facility for four hours.

In the chronic study, 12 subjects were required to consume the LC n-3 PUFA-enriched luncheon meat for three weeks. During this three-week period, foods rich in n-3 and n-6 PUFA (e.g. seafood, nuts, peanut butter, soy and linseed bread, oils and margarine) were excluded. A cool bag was used for each subject to collect their quantity of LC n-3 PUFA-enriched luncheon meats to take home. The subjects were seen at least once a week while they collected their supply of luncheon meats. This contact was designed to maintain their interest in the study, to encourage the avoidance of prohibited foods and to ensure their compliance and cooperation. A 24-hour food recall was recorded for each the subjects at baseline, one week and three weeks when their blood samples were collected.

A food exclusion list was given to all the subjects prior to the study. They were also requested to avoid strenuous physical exercise on the day before the study.

Blood collection

In the present study, 5 mL venous blood was drawn from each subject into an ethylenediaminetetraacetic-acid coated blood tube between 7:00 a.m. and 9:30 a.m. on the study day after an overnight fast. In the acute study, additional blood samples were collected at two and four hours after the ingestion of a sandwich with the LC n-3 PUFA-enriched luncheon meats. In the chronic study, additional blood samples were collected at baseline, one week and three weeks after a 12-hour fast. The blood samples were placed on ice and centrifuged at 3000 r.p.m., 4[degrees]C for 10 min. Plasma was stored at -80[degrees]C within an hour until further processing. Blood was drawn by DHV who has venesection accreditation from the Pathology Foundation, Suite 3, Fourth Floor, 118 Queen Street, Melbourne, Vic. 3000, Australia.

Anthropometry assessment

In the acute study, the subjects' height, weight and blood pressure were measured during the pre- and post-study times. In the chronic study, the measurements were recorded at baseline, one week and three weeks. They were weighed in light clothing without shoes, and their blood pressures were measured using a sphygmomanometer. Body mass index was calculated in kg/[m.sup.2].

Percentage of body fat was quantified via measurement of bioelectrical impedance with a Seac SFB3 four-terminal portable analyser (supplied by Impedimed, Mansfield, Qld, Australia). (16,17) Bioelectrical impedance analysis is based on the principle that the lean body mass is highly conductive with its water and electrolyte content whereas fat is a poor conductor of the applied current. (18,19) Therefore, the resistance of the body to an electrical current is inversely related to the lean body mass. (18) The subjects' age, gender, height and weight were taken into account during the measurement. The fat mass was calculated by subtracting the lean body mass from the total body weight. The percentage of body fat was calculated by dividing the fat mass by the total body weight. The validity and reproducibility of bioelectrical impedance analysis were confirmed previously. (20)

Dietary assessment

Food frequency questionnaire

A semi-quantitative food frequency questionnaire (FFQ) was designed to be self-instructive and has been validated for use in the ranking and the quantification of dietary fat intakes. (8,21) FoodWorks v3.1, Fatty Acid database (Xyris Software (Australia), Highgate Hill, Qld, Australia) was used to analyse the nutrient intake from the FFQ and the 24-hour recall. Published food composition tables, (22) 'The Omega Plan' (23) and 'The Omega-3 Life Program: your food guide for health and healing' (24) were provided for subjects to read to encourage their understanding of the dietary restrictions used in the present study.

The 24-hour recall

In the acute study, the consumption of the LC n-3 PUFA-enriched luncheon meat was observed by the investigator. The 24-hour recall was used as a means of checking the compliance of the subjects as well as correlation of the FFQ values. Subjects were required to recall all the foods consumed on the previous day. If the subject could not avoid foods that were supposed to be excluded, they were requested to record the name and amount of those foods in the 24-hour recall. Subjects in the chronic study provided a 24-hour recall at zero, one and three weeks as a dietary compliance check and an FFQ correlation check.

Test food

An Australian manufacturer, Hans Continental Small-goods, Colmslie, Qld, Australia has been involved in increasing the level of LC n-3 PUFA in a range of low and reduced fat products. The product was designed to meet the requirements of the National Heart Foundation of Australia recommendations on dietary fats. (25) The n-3-enriched luncheon meats were provided by the manufacturer. Microencapsulation technology was used to enrich the meats with tuna oil, which is a traditional component of the human diet. Microencapsulation involves the incorporation of liquid oil in microcapsules that release their contents under specific conditions. (26) This technology has increased the variety of foods into which fish oil can be incorporated and thus increases the shelf-life of these enriched foods by reducing the potential for oxidation of the fatty acids. (26) A total of 190 g of luncheon meat was given to the subjects in the form of vienna franks, sliced chicken, sliced strassburg and strassburg roll. This 190-g amount provided 200 mg of LC n-3 PUFA that contained approximately 75% of DHA and 25% of EPA. The meats were stored in a cold room in the Human Nutrition Unit and packed individually as required. Helga's Traditional white bread (Quality Bakers Australia, Macquarie Park, NSW, Australia) and Monosun monounsaturated sunflower margarine (Flora, Unilever Australasia, Epping, NSW, Australia) were served to subjects during the acute study.

Laboratory fatty acid analysis

All organic solvents, glacial acetic acid and sodium chloride were obtained from Asia Pacific Specialty Chemicals (Seven Hills, NSW, Australia). Thin layer chromatography lipid standards, butylated hydroxy toluene, boron trifluoride: methanol reagent and dichlorofluorescein were obtained from Sigma-Aldrich (Castle Hill, NSW, Australia).

Plasma lipids were extracted using a method modified from Folch et al. (27) In total, 1 mL of plasma was added to 20 mL of 2 : 1 (v/v) AR grade chloroform : methanol mixture containing 0.005% (w/v) of butylated hydroxy toluene (as an antioxidant) in glass screw lid tubes and vortexed for 1 min. Afterwards, 4 mL of 0.9% (w/v) aqueous sodium chloride solution was added and vortexed for a further 15 seconds. The tubes were placed in ice for 45 minutes to allow the layers to separate. The lower chloroform layer was removed and evaporated under a dry nitrogen gas stream.

The lipid extract was reconstituted in 100 [micro]L of hexane for separation by thin layer chromatography. Sample extracts and lipid standards (for identification of sample lipid fractions) were spotted onto separate 2-cm tracks on 250 [micro]m silica gel 60 plates (Merck, Kilsyth, Vic., Australia). The plate was then developed for approximately 45 min in a sealed glass tank containing a 85 : 15 : 1 (v/v/v) hexane : diethyl ether : glacial acetic acid mobile phase containing 0.005% (w/v) of butylated hydroxy toluene, then sprayed with a 1% (w/v) 2',7'-dichlorofluorescein in methanol solution. Lipid fractions were visualised under ultraviolet light and identified by comparison with lipid fraction standards. The non-esterified fatty acids (NEFA) and phospholipid fractions were scraped off the plate into 10 mL glass screw lid tubes.

Derivitisation of NEFA and phospholipid fractions into fatty acid methyl esters was performed by adding 1 mL each of 14% (w/v) hexane and boron trifluoride : methanol reagent to the silica gel scrapings, and the tubes sealed firmly with Teflon tape, then heated at 100[degrees]C for 1 h. After cooling, 2 mL of distilled water and 1 mL of hexane was added and the tubes vortexed for 1 min. The top organic layer was removed and the solvent evaporated under dry nitrogen gas stream. The methylated lipid fractions were then reconstituted in 100 [micro]L of toluene for gas chromatography analysis of fatty acid methyl esters, which was performed by the Child Nutrition Research Centre (Flinders Medical Centre, Bedford Park, SA, Australia).

Statistical analysis

Paired t-test was used to analyse the results. An alpha level of 0.05 was set to determine statistical significance, and all analyses were conducted using SPSS 11 (SPSS, Chicago, IL, USA, SPSS for Windows, version 11, 2001).

Results

Study population

Subject characteristics are given in Table 1. Mean age was 28 and 33 years, respectively, in the acute and the chronic study. Mean body mass index was 21 and 24 kg/[m.sup.2], respectively. Mean percentage of body fat was 21.9% and 24.7%, respectively. Subjects' weight remained stable throughout both study interventions.

Habitual dietary intake

The data for habitual intake of food components from the FFQ are given in Table 2. In the acute study, mean daily energy intake was 9995[+ or -]2110 kJ. Overall, the contribution to daily dietary energy intake was 21% for protein, 26.5% for fat, 48.5% for carbohydrate and 1.5% for alcohol. Mean fat intake was 71.5[+ or -]14.5 g/day. Of this, saturated fat was 9.3% of energy intake; monounsaturated fat, 11.7% and polyunsaturated fat, 5.5%. Mean LC n-3 PUFA intake was 390 mg with 6.08 g of n-6 PUFA. The n-6 : n-3 ratio was 15.6 in the acute study population. Main sources of LC n-3 PUFA are from fish and shellfish; main sources of n-6 PUFA are from nuts, soy products, salad dressing, takeaway foods and snacks. In the chronic study, mean daily energy intake was 9356 [+ or -] 686 kJ. The contribution to daily dietary energy intake was 20% for protein, 31% for fat, 44% for carbohydrate and 2.8% for alcohol.

Mean fat intake was 78.4 [+ or -] 13.6 g/day. Of this, saturated fat was 12.4% of energy intake; monounsaturated fat, 13.4% and polyunsaturated fat, 5.2%. Mean LC n-3 PUFA intake was 580 mg with 5.05 g of n-6 PUFA. The n-6 : n-3 ratio was 8.7 in the chronic study population.

Dietary intervention

The test food provided for the study participants in the acute and the chronic groups was 64 g of vienna franks, 28 g of sliced chicken, 32 g of sliced strassburg and 66 g of strassburg roll. This quantity of luncheon meat (190 g) provided 200 mg of LC n-3 PUFA. The LC n-3 PUFA content was 75% of DHA and 25% of EPA (Table 3). Participants in the acute study were observed to consume 190 g of luncheon meat. Participants in the chronic study completed a 24-hour food recall on each visit, and their compliance was assessed by comparison with LC n-3 PUFA results of their plasma fatty acid and phospholipid analyses. All participants appeared to be compliant.

Plasma NEFA in acute study

Long-chain n-3 PUFA are EPA, docosapentaenoic acid (DPA) and DHA. Changes in these fatty acids in plasma NEFA are presented in Figure 1. EPA, DPA and DHA levels increased from time zero to four hours after consumption of the test meal. Plasma NEFA showed an immediate and significant increase in EPA following the test meat from time zero to 4 hours after intake of the n-3-enriched meats (P = 0.043). DPA and DHA increased from time zero; however, both increases were insignificant (Figure 1). Meanwhile, the percentage change in LC PUFA, EPA, DPA and DHA increased by 39%, 37% and 53%, respectively, at four hours after the intake of LC n-3 PUFA-enriched meats.

[FIGURE 1 OMITTED]

Plasma NEFA and phospholipid status in chronic study

The changes of EPA, DPA and DHA in plasma phospholipids are presented in Figure 2. There were significant increases in EPA (P = 0.04), total n-3 PUFA (P = 0.006) and n-6 : n-3 ratio (P = 0.006) at three weeks. Meanwhile, the percentage change in LC PUFA, EPA, DPA and DHA increased by 25%, 47% and 55%, respectively, at three weeks from time zero. Total LC n-3 PUFA increased by 44% and the n-6 : n-3 ratio decreased by 20% after three-week consumption of LC n-3-enriched meats. The changes of EPA, DPA and DHA in plasma NEFA in chronic study are presented in Figure 3. DHA increased by 163%, which was not significant; however, EPA decreased significantly by 22% from zero to three weeks (P = 0.043).

Discussion

Non-esterified fatty acids in plasma are used as a biomarker to determine the effect of a single serve of LC n-3 PUFA-enriched luncheon meat on postprandial plasma status. EPA NEFA increased significantly from 0.30 [+ or -] 0.02% of fatty acids to 0.42 [+ or -] 0.05% of fatty acids. NEFA in the chronic study are used to assess compliance to the prescribed diet as NEFA reflects the fatty acids consumed in the daily diet. Plasma phospholipids are used to assess the longer term intake of fatty acids. The daily serve of LC n-3 PUFA-enriched luncheon meat over three weeks resulted in a significant increase in EPA phospholipids from 0.93 [+ or -] 0.16 to 1.16 [+ or -] 0.16% of fatty acids. Total n-3 PUFA increased significantly from 5.48 [+ or -] 0.80 to 7.92 [+ or -] 0.53% of fatty acids, and the ratio of n-6 : n-3 decreased significantly from 5.10 [+ or -] 0.41 to 4.07[+ or -]0.24. These results demonstrate that supplementation of a low level of LC n-3 PUFA in luncheon meat can provide an important dietary source of LC n-3 PUFA.

[FIGURE 2 OMITTED]

[FIGURE 3 OMITTED]

The richest food sources of LC n-3 PUFA are found in marine foods, such as fish and shellfish. Small amounts of LC n-3 PUFA can be found in lean meat, egg, milk and dairy products. (2,28) Humans cannot obtain significant amounts of DHA from the conversion of ALA as the conversion rate is only 10-15%. This low rate of conversion takes place providing n-6 LA in the diet is low. LA competes directly with ALA for the enzyme systems involved in desaturation and elongation. (29) As a functional food, the enriched luncheon meat offers an alternative to the use of fish oil capsules or fish as a source of LC n-3 PUFA in the diet. The results of this bioavailability study demonstrate that the LC n-3 PUFA-enriched luncheon meat did improve plasma LC n-3 PUFA status in both four-hour postprandial period and three-week period. The results are also consistent with other studies using a variety of LC n-3 PUFA-enriched food, for example the bread enrichment study by Yep et al. (15) In the egg enrichment study by Farrell, the results showed significant increases in EPA, DHA and total n-3 PUFA, as well as a significant decrease of the n-6 : n-3 ratio. (14) This was achieved after 16 and 22 weeks consumption of a 60-g n-3-enriched egg (containing minimum 300 mg n-3 PUFA, of which 50% was EPA) daily compared with the control group. The high level of incorporation of EPA and DHA into the plasma fatty acids in the present study indicates that the LC n-3 PUFA (Table 3) in microencapsulated oil were well absorbed and bio-available.

The total n-3 and n-6 intakes determined by FFQ in the present study population was 390 mg and 6.08 g in the acute study and 580 mg and 5.05 g in the chronic study, respectively. The reported dietary intake of n-3 PUFA was higher than the average intake recorded for the National Nutrition Survey. (13) Additionally, the n-6 : n-3 ratio of the study groups was 15.6 for the acute study and 8.7 for the chronic study, which is higher than the 8.1 Australian population mean. (13) The mean habitual diet of the acute and the chronic groups was similar, except for the n-6 PUFA content. This may reflect the use of peanut oil in the food preparation methods of the predominantly Asian group in the acute study. The high ratio of n-6 : n-3 in the diets of the acute group would not influence results as only post-prandial measures were reported. The western diet is rich in n-6 PUFA, especially LA, the most abundant PUFA (30) compared with small amounts of n-3 PUFA (100-190 mg) normally available in the diet, suggesting that the study group were not fully representative of the typical western population. (31) There is also a possibility that the FFQ may underestimate usual energy intake, therefore, absolute estimates of nutrient intake may be underestimated. (18) Furthermore, the small sample size (n = 4 Asians and 2 Caucasians for the acute study; n = 2 Asians and 10 Caucasians for the chronic study) may be another limitation of the present pilot study. A control group was not used in the present study, this may limit the value of these findings. The key to efficient LC n-3 PUFA incorporation in cellular membrane phospholipids is to limit n-6 PUFA dietary intake. AA competitively inhibits EPA incorporation in cellular membrane phospholipids. (1,2,23)

The significant increase of total LC n-3 PUFA levels in plasma NEFA in the acute study can be explained by the LC n-3 PUFA content of the luncheon meats and the restriction of n-6 PUFA in the background diet. According to the nutrition information provided by the manufacturer, the n-3 content was estimated as 205 mg per serving of franks, 210 mg per serving of strassburg and 500 mg per serving of chicken. The n-6 : n-3 ratio in NEFA in acute study was increased from 3.97 to 4.67 after four hours. In the chronic study, the n-6 : n-3 ratio in NEFA decreased from 5.01 to 4.07 after three weeks consumption of n-3-enriched meats. Sinclair et al. reported that diets rich in kangaroo meat or lean beef also induced significantly higher plasma AA levels than did vegetarian diets. (12,28) When too much AA is present in the diet, the enzyme systems preferentially elongate AA instead of n-3 PUFA with the result that the n-6 derived precursors of eicosanoids are utilised for eicosanoid production. This results in the increase of class 2 thromboxanes, leukotrienes and prostaglandins. The elevation of plasma phospholipid levels of LC n-3 PUFA and their incorporation in cell membrane phospholipids will result in an increase in class 3 thromboxanes, leukotrienes and prostaglandins, which are significantly less potent and less active than the class 2 metabolites. (32)

Several biological markers can be used as a measure of LC n-3 PUFA status in humans, for example, plasma, platelets, erythrocytes in blood, adipose tissue and cheek cells. (33) All have been reported to correlate reasonably well with dietary intervention trials. (4,7,8,34,35) In the present study, plasma was subjected to analysis as plasma NEFA can be acutely influenced by a single meal. (33) For a longer-term intake, phospholipids are a better indicator to monitor dietary changes in subjects following a special diet, as phospholipids are the active site for LC n-3 PUFA. (33)

Plasma non-esterified EPA content was significantly increased after four hours in the acute study (Figure 1), whereas EPA (in plasma phospholipids) was significantly increased after three weeks in the chronic study (Figure 2). This finding supports prior work that demonstrated that plasma non-esterified EPA reflects short-term n-3 PUFA intake, (36) whereas phospholipids EPA is a biomarker of longer-term LC n-3 PUFA intake. (37)

In a previous study healthy subjects consumed a test meal where fish oil provided the sole fat source of 0.5 g/kg body weight for 12 hours. (38) Both EPA and DHA in plasma NEFA increased significantly by the fourth hour of the test period over baseline, and remained at the peak levels for four to five hours. In the present study, DHA in NEFA and phospholipids tended to be higher than EPA, notwithstanding the fact that most sources of dietary n-3 PUFA contain more EPA than DHA. This difference was also found in the previous study. (38) This is attributed to a tendency of DHA to accumulate in the NEFA fractions. (38) A possible explanation for this is that the enzymatic capacity for the elongation and desaturation of EPA to DHA is low, but with chronically low intakes of EPA it is of sufficient capacity to allow maximal conversion of EPA to DHA. (38)

The quantity of EPA and DHA consumed is known to be important in determining the physiological effects of these fatty acids. The luncheon meats contain 75% of DHA and 25% of EPA, which explains the obvious increase of DHA in both the acute and chronic studies. Some previous studies on the n-3 PUFA-enriched margarine, (6) eggs (14) and milk (7) have used foods containing 300-910 mg LC n-3 PUFA per serving. They all demonstrated a marked increase of n-3 PUFA in plasma. The present study offered a lower dose of 200 mg of LC n-3 PUFA to the luncheon meat, with similar results. This indicates that the enrichment of LC n-3 PUFA can be as low as 200 mg, which increased human plasma PUFA levels, although the use of a control group would have strengthened these results. However, it still does not meet the recommendation of 650 mg per day made by ISSFAL. (2)

In summary, the consumption of 200 mg LC n-3-enriched luncheon meats (75% DHA and 25% EPA) increased plasma LC n-3 PUFA levels. Our results illustrate that it is feasible to use manufactured enriched foods as a means of increasing LC n-3 PUFA intake for a healthy population. The present study further confirms that functional foods could be used as a good source of LC n-3 PUFA when individuals do not like, do not eat or cannot obtain fish.

Acknowledgements

The present study was sponsored by Hans Continental Smallgoods (Colmslie, Qld, Australia). We are grateful to Wendy Morgan of Innovations and Solutions, North Sydney, NSW, Australia, for her introduction to Hans Continental Smallgoods. Our thanks also go to Anne-Marie Moxham, brand manager of Hans Continental Smallgoods for the coordination of this consultancy.

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Human Nutrition Unit, School of Molecular and Microbial Biosciences, The University of Sydney, Camperdown

D.H. Volker, APD, PhD

X. Weng, BMed, MNutrDiet, APD

P. Quaggiotto, BSc

Correspondence: D. Volker, formerly, Human Nutrition Unit, School of Molecular and Microbial Biosciences, G08, The University of Sydney, Camperdown, NSW 2006, Australia. Email: [e-mail] D.H. Volker was responsible for the study design and preparation and amendments to the paper. X. Weng was responsible for subject recruitment and supervision of study participants. P. Quaggiotto was responsible for sample processing and data analysis.

Table 1. Physical characteristics and study sample (mean [+ or -] SEM)

                  Acute                    Chronic

Sample size (n)     6                       12
Age (years)        28.0 [+ or -] 0.7        33.0 [+ or -] 3.0
BMI                21.0 [+ or -] 1.0        24.0 [+ or -] 1.0
  (kg/[m.sup.2])
% Body fat         21.9 [+ or -] 1.4        24.7 [+ or -] 1.1
Height (cm)       165.6 [+ or -] 4.0       170.1 [+ or -] 2.3
Weight at time     58.2 [+ or -] 5.6        69.8 [+ or -] 4.6
  zero (kg)
BP at time zero   116/68 [+ or -] 2.4/3.3  114/70 [+ or -] 3.6/2.9
  (mmHg)
Weight at four     59.0 [+ or -] 5.5       --
  hours (kg)
BP at four hours  117/67 [+ or -] 2.8/2.5  --
  (mmHg)
Weight at three   --                        69.7 [+ or -] 4.5
  weeks (kg)
BP at three       --                       112/71 [+ or -] 2.6/2.5
  weeks (mmHg)

The male: female ratio was 2 : 4. The subjects in the acute study were
shorter and lighter than the chronic group, reflecting the predominance
of Asian subjects. The chronic group was predominantly non-Asian with a
gender ratio of 2 males and 10 females. (10) The blood pressure (BP)
decrease in the chronic group at the completion of the study may reflect
the vasodilatory effects of LC n-3 polyunsaturated fatty acids.
BMI = body mass index.

Table 2. Energy and selected nutrient intakes of the study sample (mean
[+ or -] SEM)

                         Acute                 Chronic

Energy (kJ)              9995.0 [+ or -] 2110  9356.0 [+ or -] 686
Protein (%)                21.0 [+ or -] 2.2     20.0 [+ or -] 0.9
Total fat (%)              26.5 [+ or -] 3.0     31.0 [+ or -] 1.6
Carbohydrate (%)           48.5 [+ or -] 2.7     44.0 [+ or -] 2.3
Alcohol (%)                 1.5 [+ or -] 1.2      2.8 [+ or -] 0.7
Saturated fat (%)           9.3 [+ or -] 1.3     12.4 [+ or -] 0.9
Monounsaturated fat (%)    11.7 [+ or -] 1.6     13.4 [+ or -] 0.8
Polyunsaturated fat (%)     5.5 [+ or -] 1.0      5.2 [+ or -] 0.4
n-3 PUFA (g)                0.4 [+ or -] 0.2      0.5 [+ or -] 0.1
n-6 PUFA (g)                6.1 [+ or -] 3.3      5.1 [+ or -] 0.9
LC PUFA n-3 (g)             0.1 [+ or -] 1.6      0.1 [+ or -] 0.03
Ratio of n-6 : n-3         15.6                   8.7

The subjects for the present study were recruited from the staff and
student population of the School of Molecular and Microbial Biosciences
at the University of Sydney. The mean habitual diet of the acute and the
chronic groups were not significantly different (P < 0.05).
LC PUFA = long-chain polyunsaturated fatty acids; PUFA = polyunsaturated
fatty acids.

Table 3. Nutritional composition of the four types of luncheon meat
consumed by the study sample

Average quantity                     Strassburg  Strassburg  Chicken
(per 100 g)           Vienna franks  (sliced)    (roll)      (sliced)

Energy (kJ)           427.0          543.0       543.0       535.0
Protein (g)            13.2           15.4        15.4        21.7
Fat (total) (g)         2.9            2.9         2.9         2.0
Saturated fat (g)       1.4            1.3         1.3         0.7
Trans fat (g)          <0.1           <0.1        <0.1        <0.1
MUF (g)                 1.1            1.3         1.3         0.7
PUF (g)                 0.3            0.3         0.3         0.5
DHA (mg)               72.0           68.0        68.0       135.0
EPA (mg)               24.0           23.0        23.0        45.0
Carbohydrate (total)    7.7            8.4         8.4         5.3
  (g)
Sugars (g)              1.5            3.0         3.0         1.3
Sodium (mg)           740.0          740.0       740.0       740.0
Gluten (g)              0.0            0.0         0.0         0.0

The luncheon meats consumed in the present bioavailability study were
designed as functional foods as a low-fat source of LC n-3 PUFA. The
four types of luncheon meats chosen for enrichment with
microencapsulated tuna oil (Clover Corporation, Driphorm 25, North
Sydney, NSW, Australia) were the best selling products in the luncheon
meat range of Hans Continental Smallgoods.
DHA = docosahexaenoic acid; EPA = eicosapentaenoic acid; MUFA =
monounsaturated fat; PUFA = polyunsaturated fat.