The results of this study showed that 6 weeks of supplemental fish oil significantly increased lean mass, and significantly reduced fat mass in healthy adults. This is in agreement with Couet et al. , who observed a significant 0.88 kg reduction in fat mass, and a non-significant 0.20 kg increase in lean mass following 3 weeks of an increased consumption of fish oil. In their study, they added fish oil to the diet, but kept total fat and energy constant between the treatments. In the present study, the fish oil was added on top of an ad libitum diet, with instructions given to the subjects to maintain their normal dietary patterns throughout the study. Similarly, Hill et al  found a significant reduction in fat mass following 12 weeks of supplementation with fish oil in overweight subjects. They also observed an increase in lean mass in the fish oil group, however, like the data reported by Couet et al. , it did not reach significance. Thorsdottir et al.  recently found that supplementation with fish oil, or inclusion of fish in an energy-restricted diet resulted in significantly greater weight loss in young men. Additionally, they found that young men taking the fish oil supplements had a significantly greater reduction in waist circumference compared to the control group, or the group that increased their dietary intake of fish.
Unlike the Couet et al. study , we did not observe an increase in RMR, or a decrease in RER following fish oil treatment. The failure to find an increase in RMR following fish oil treatment is hard to explain given the significant increase in lean mass observed in the present study. Several studies have shown that lean mass is the largest determinant of RMR [28–30], and decreasing lean mass decreases RMR , while increasing lean mass increases RMR . Therefore, it would be expected that the increase in lean mass would correspond to an increased RMR following fish oil treatment. In the Couet et al. study , metabolic data were measured for 45 min following a 90 min rest period. This is a longer time period than the 40 min used in the present study. However, it is doubtful that this methodological difference between the studies contributed to the differing effects observed for RMR and RER values since recent studies have shown that very short rest periods (as little as 5 min) produce reproducible results that correlate extremely well with RMR measures made over much longer time periods [33, 34]. It is also unlikely that the use of a subset (n = 24) of the total subject population can explain the failure to observe any metabolic changes since analysis of the 24 subjects found that they responded similar to the entire group in regards to body composition changes. It remains unclear why the increased lean mass observed following fish oil treatment did not correspond to an increase in RMR.
Intuitively it would make sense that if fat mass was reduced, but resting metabolic rate did not change following fish oil treatment, then the amount of calories coming from the oxidation of fatty acids should be increased. However, this was not the case in the present study. Although there was an absolute reduction in the RER following fish oil treatment (which would indicate an increased oxidation of fatty acids), the difference was not statistically significant. While it is possible that a type II error was committed and the reduction in RER was a real effect, it is also possible that the fish oil treatment increased fat oxidation at other times during the day such as during exercise , or during the post-prandial period .
A potential shortcoming of the present study was not using dietary records to monitor the subjects' intake during the study. Although there are several potential problems with the use of dietary records (for a review of inaccuracies with self-recorded diet records see ), they would have provided us with some insight into the dietary habits of the subjects during the study. It therefore remains a possibility that the fish oil supplements resulted in the subjects changing their normal dietary habits. Although increasing dietary fat does not generally cause a decrease in voluntary fat intake , it has been shown that fish oil may reduce appetite , which could have led to the subjects consuming less total calories during the study. While a reduction in volitional food intake would explain the observed reduction in fat mass following fish oil treatment, it does not explain the increase in lean mass we observed.
Although other studies have observed a significant [3, 5], or insignificant [21, 22], increase in lean mass following fish oil treatment, to date no study has determined the mechanism by which dietary fish oil causes an increased accretion of lean mass. One possibility lies in the well-documented ability of dietary omega 3 fatty acids to reduce inflammatory cytokines , since inflammatory cytokines have the ability to increase protein degradation mainly by activating the ATP-ubiquitin-dependent pathway [41–45]. It is possible then, that dietary fish oil is simply decreasing the breakdown of protein tissue caused by inflammatory cytokines, and this results in an increased accretion of protein over time.
An alternative possibility is that fish oil supplementation was able to increase lean mass by reducing cortisol levels since it is well established that cortisol increases protein catabolism [46–49]. The significant negative correlation (r = -0.504, p = 0.02) observed in the fish oil group between the change in lean mass and the change in salivary cortisol concentrations would support this hypothesis. Although other studies have observed a decrease in cortisol levels following fish oil consumption , the exact mechanism(s) responsible are currently unknown. However, it is possible that the reduction of IL-6 as a result of fish oil consumption  is causing a reduction in cortisol production since it has been shown that IL-6 induces increases in cortisol levels [51, 52]. It is unclear whether it is the well-documented ability of fish oil to reduce inflammatory cytokines, the reduction in cortisol, or a combination of both, that resulted in the increased lean mass observed in the present study following fish oil treatment. More work is needed to determine the mechanism(s) responsible for the accretion of lean mass following fish oil consumption.
The role of cortisol in obesity is poorly understood. Excessive cortisol levels, such as those observed in patients with Cushing's disease, results in substantial fat mass gains - especially in the abdominal region [17, 19]. However, there is disagreement between studies about the relationship between values of cortisol that are within a normal physiological range, and obesity . Nevertheless, several studies have shown an association with higher levels of cortisol and fat mass [53–58]. In the present study, there was a significant correlation between the change in salivary cortisol and the change in fat mass following fish oil treatment (r = 0.661, p = 0.001). Recent work by Purnell et al.  has shown that a reduction in fat mass as a result of dieting does not lower cortisol production, which would suggest that the relationship observed in the present study between salivary cortisol and fat mass was not simply a result of the reduction in fat mass. However, further work is needed to determine exactly how the reduction in cortisol levels may have influenced fat loss observed in the FO group.
In conclusion, 6 weeks of supplemental fish oil significantly increased lean mass, and significantly reduced fat mass in healthy adults. Given the short duration of this study, it is unclear how these changes would impact long-term body composition changes and more research is needed to determine the impact of chronic fish oil supplementation on long-term body composition. The reduction in salivary cortisol following fish oil treatment was significantly correlated with the increased fat free mass and the decreased fat mass observed. To the best of our knowledge, this is the first time that this association has been described in the literature. Since higher salivary cortisol levels are associated with higher mortality rates , the reduction in salivary cortisol levels observed in the present study following fish oil supplementation likely has significant implications beyond positive changes in body composition.