Enhanced incorporation of dietary DHA into lymph phospholipids by altering its molecular carrier

Abstract

Several previous studies indicated that for optimal uptake by the brain, docosahexaenoic acid (DHA) should be present as phospholipid in the plasma. However most of dietary DHA is absorbed as triacylglycerol (TAG) because it is released as free fatty acid during digestion of either TAG-DHA (fish oil) or sn-2-DHA phospholipid (krill oil), and subsequently incorporated into TAG of chylomicrons. We tested the hypothesis that the absorption of DHA as phospholipid can be increased if it is present in the sn-1 position of dietary phospholipid or in lysophosphatidylcholine (LPC), because it would escape the hydrolysis by pancreatic phospholipase A2. We infused micelle containing the DHA either as LPC or as free acid, into the duodenum of lymph cannulated rats, and analyzed the chylomicrons and HDL of the lymph for the DHA-containing lipids. The results show that while the total amount of DHA absorbed was comparable from the two types of micelle, the percentage of DHA recovered in lymph phospholipids was 5 times greater with LPC-DHA, compared to free DHA. Furthermore, the amount of DHA recovered in lymph HDL was increased by 2-fold when LPC-DHA micelle was infused. These results could potentially lead to a novel strategy to increase brain DHA levels through the diet.

Keywords: Chylomicrons/HDL; Fish oil/DHA; Lymph; Lysophosphatidylcholine; Micelles; Phospholipids/absorption.

Fig. 1. Total amount of DHA secreted into lymph

Total amount of DHA secreted in 6 h in individual experiments is shown. Although there were individual variations among the animals in each group, the mean values were not significantly different between the two groups. The dotted line is the mean value and the solid lines are SEM. N= 8 for both groups. The total amount of DHA infused into the duodenum was 24 µmoles (7.88 mg) in both groups.

Fig. 1. Total amount of DHA secreted into lymph

Total amount of DHA secreted in 6 h in individual experiments is shown. Although there were individual variations among the animals in each group, the mean values were not significantly different between the two groups. The dotted line is the mean value and the solid lines are SEM. N= 8 for both groups. The total amount of DHA infused into the duodenum was 24 µmoles (7.88 mg) in both groups.

Fig. 2. Percent of total DHA recovered in HDL fraction of the lymph

HDL and CM from each lymph fraction were separated by centrifugation at a density of 1.063 g/ml, and their fatty acid composition was determined by GC/MS, as described in Section 2.4. The values shown are percentages of total lymph DHA which was recovered in HDL, and are mean ± SEM of 9 experiments for LPC-DHA and 7 experiments for free DHA. The remaining DHA was CM fraction. ** p< 0.01, free DHA vs LPC-DHA (unpaired t test)

Fig. 3. Percent of DHA recovered in PC

The PC and TAG from the lipids of total lymph, HDL, and CM were separated on aminopropyl columns, and the fatty acid composition was determined by GC/MS. The percent of DHA recovered in PC was calculated from this data. The PC fraction contained a small amount of DHA as PE (<10%), since these two lipids were not separated in this procedure. All the remaining DHA was in TAG. The results shown are mean ± SEM. (n= 10 total lymph for LPC-DHA, n=8 for total lymph for free DHA, n=5 each HDL and CM for both micelle). * P<0.05; ** p<0.005 free DHA vs LPC-DHA (unpaired t test)

Fig. 4. Molecular species of TAG containing DHA in HDL

The molecular species composition of DHA-containing TAG species was determined by MRM in LC/MS as described in Section 2.5 (and supplementary data, Table 1). The nomenclature of the species is based on the fatty acid composition, but the position of the fatty acids on the glycerol backbone has not been determined. The values shown are mean ± SEM of 9 experiments for LPC-DHA and 7 experiments for free DHA. * p <05, free DHA vs LPC-DHA (unpaired t test)

Fig. 5. Molecular species of TAG containing DHA in lymph CM

The molecular species composition was determined by MRM in LC/MS as described in Section 2.5 (and supplementary data, Table 1). The values shown are mean ± SEM of 9 experiments for LPC-DHA and 7 experiments for free DHA. * p <0.05, free DHA vs LPC-DHA (unpaired t test)

Fig. 6. Molecular species of DHA-containing PC in lymph HDL

The molecular species composition of PCs that contained DHA was determined by MRM, as described in Section 2.5 (and supplementary data, Table 1). The position occupied by DHA was not determined. The values shown are mean ± SEM of 9 experiments for LPC-DHA, and 7 experiments for free DHA * p < 0.05, ** p < 0.005, Free DHA vs LPC-DHA (unpaired t test)

Fig. 7. Molecular species of DHA-containing PC in CM

The molecular species composition of PCs containing DHA was determined by MRM, as described in Section 2.5 (and supplementary data, Table 1). The position occupied by DHA was not determined. The values shown are mean ± SEM of 9 experiments for LPC-DHA, and 7 experiments for free DHA * p< 0.05, Free DHA vs LPC-DHA (unpaired t test)

Hypothesis 1

Absorption of TAG-DHA and sn-2 DHA PC

Hypothesis 2

Absorption of sn-1 DHA PC