The aim of this study was to evaluate whether plasma incorporation, apparent retroconversion, or β-oxidation of 13C-DHA differed in the healthy elderly compared to healthy young adults. We observed that within the post-prandial period (+ 4 h) the elderly had at least 4 fold higher 13C-DHA in plasma TG and FFA, and 1.9 fold higher β-oxidation to 13C-CO2. 13C-DHA was also 2.5 fold higher in plasma CE of the elderly after 7d, and its apparent retroconversion was 2.2 fold higher in the elderly 24 h after the oral dose. Hence, our results indicate that there are significant differences in DHA metabolism during healthy aging which compliment, extend and possibly help explain previous reports showing somewhat higher plasma DHA in the elderly [5–9].
The distribution of 13C-DHA we observed in plasma lipid classes of our young study participants agrees with previous published papers [24, 25, 31]. We speculate that the higher early rise in 13C-DHA in plasma TG and FFA in the elderly (Figure 2) may potentially be explained by the elderly having both higher postprandial production of very low density lipoproteins which are rich in TG , as well as a higher post-prandial FFA response . However, one limitation is that postprandial TG-rich lipoproteins were not measured in this study so this explanation remains to be confirmed. The 4-fold higher 13C-DHA in plasma CE in the elderly after 7d persisted out to 28d and may be due to different lipoprotein metabolism in the elderly as suggested by their (i) higher plasma total cholesterol (Table 1), (ii) higher plasma residence time of low density lipoprotein , and/or (iii) lower turnover of low density lipoprotein .
The appearance of 13C in other omega-3 PUFA besides DHA (mostly in 13C-EPA) was about 2 times higher in the elderly 24 h to 7d after tracer intake. This measure of apparent retroconversion of 13C-DHA to 13C-EPA could occur by one cycle of β-oxidation and chain shortening . Subsequent chain lengthening of 13C-EPA could explain the appearance of 13C-DPA. Alternatively, recycling of carbon from PUFA through 13C-acetate into newly synthesized fatty acids is a well known phenomenon , and could potentially become part of ALA because humans can make small amounts of ALA from 16:3 n-3, or omega-3 PUFA longer than ALA via conventional elongation .
The mean total 13C-EPA and 13C-DPA appearing in plasma is well below 10% even at 28d post-dose, so this amount of apparent retroconversion would seem unlikely to fully account for net > 5% rise in plasma phospholipid EPA after DHA supplementation reported elsewhere , unless the process were substantially up-regulated using multigram DHA supplements. Raised plasma EPA after a DHA supplement may also be due to reduced EPA turnover by sparing mechanisms with enhanced dietary DHA.
We show for the first time that 13C-CO2 from β-oxidation of 13C-DHA peaked at 4 h post-intake and was 1.9 fold higher in the elderly (Figure 3). These data confirm a preliminary report  showing that humans β-oxidize 13C-DHA much more slowly than 13C-ALA . The major regulatory mechanism for the control of β-oxidation is the availability of substrate, mostly as fatty acids in postprandial circulating TG and FFA . The higher β-oxidation of 13C-DHA at 4 h post-dose in our elderly participants is in line with their higher 13C-DHA in plasma TG and FFA 4 h post-dose (Figure 2). After 7d, 13C-CO2 was still above baseline in both groups although plasma 13C-DHA concentration had returned close to baseline in TG and FFA. Cumulative β-oxidation of 13C-DHA was not different in the young and the elderly and reached 35 - 38% after 7d, which is about half that seen for the same dose of 13C-ALA over the same time period . Under the conditions of this study, these results give a rough estimate of about 3 wk to turnover DHA to expired CO2, or a DHA biological half-life of about 10 d. However, overall β-oxidation was not higher in the elderly even though the elderly did have 13C-EPA derived from 13C-DHA. There is no implicit reason why the age-related difference in recovery of 13C as 13C-EPA should be directly related to net loss of 13C-DHA to 13CO2; both processes involve a complex interchange of carbon. Furthermore, we do not yet know how the EPA and other omega-3 fatty acids became labelled with 13C; if it was via ‘direct’ retroconversion, i.e. by chain shortening, there is no implicit reason why this form of retroconversion should be coupled to the β-oxidation of DHA; if it was via oxidation to acetyl-CoA, perhaps one would expect a closer correlation between β-oxidation of DHA and EPA labelling. Analytically, we cannot yet distinguish between these two possibilities. Incidentally, the elderly did have higher β-oxidation, but only at the earliest time point (+ 4 h).
The possible relevance of aging-related changes in DHA metabolism to risk of chronic diseases, particularly cognitive decline, remains to be established. Fish intake seems to decrease the risk of cognitive decline  and EPA and DHA in blood are biomarkers of fish intake  but, paradoxically, an overview of the literature shows that lower blood DHA is not seen in Alzheimer's disease or other forms of dementia [2, 39]. Indeed, the aging-associated changes in DHA metabolism we report here suggest that higher plasma DHA in the elderly could actually mask lower DHA availability to the brain. Another factor that was not considered in this small study is the presence of apolipoprotein E ε4 gene polymorphisms which appear to alter plasma EPA and DHA metabolism .
Because of the limited availability of the tracer and its very high cost, we had only six participants per group. However, a tracer is much more sensitive and specific than a fish oil supplement such that low number of participants per group can still generate useful and statistically valid results . Indeed, previous studies using 13C-DHA used only 3 - 4 participants [24, 25, 29, 31].