Botanical seed oils from plants such as borage and echium have shown modest efficacy in a number of animal and human inflammation models and disease. These botanicals contain 18C-PUFAs (ALA, SDA and GLA) that can be metabolized into 20–22 carbon PUFAs such as EPA, DHA, DGLA and AA. All of these have been shown to impact eicosanoid generation. However, a better understanding of the in vivo biochemistry of potentially bioactive PUFAs found in these botanical seed oils and oil combinations and their capacity to block inflammatory processes including eicosanoid production is needed to enhance the effectiveness of botanical seed oils. The current study utilized various BO/EO combinations to understand these processes.
Supplementation with BO/EO combinations increases plasma levels of n-3 and n-6, 18 carbon and 20–22 carbon PUFAs during the supplementation periods (Figure 2 and Figure 3). Of note, circulating levels of three PUFAs, DGLA, EPA and DPA increased after supplementation. It is likely that DGLA increased as a result of GLA found in both BO and EO. As discussed above, GLA is readily elongated to DGLA in cells and tissues utilizing an enzyme encoded for by a gene known as elongase 5 (ELOVL5). Once formed, DGLA is incorporated into inflammatory cells and tissues and competes with AA for the action of cytosolic phospholipase A2 and cyclooxygenase to form PGE1. Additionally DGLA is converted to a 15-lipoxygenase product, 15-hydroxyeicosatrienoic acid (15 HeTrE) by human mononuclear leukocytes . 15-HETrE has been demonstrated to be a potent blocker of LTB4 formation.
Additionally this botanical oil combination increased circulating levels of EPA and DPA This contribution was like due to EO addition since it contains the precursor PUFAs, ALA and SDA. Providing 0.25 g/d to 1.75 g/d of SDA and 0.57 g/d to 4.02 g/d of ALA from EO led to significant and dose-dependent increases in circulating EPA and DPA; plasma EPA concentrations rose more than 2-fold in the group receiving the highest concentration of SDA (Figure 2). This increase in EPA and DPA is likely a result of SDA and not ALA as in vivo SDA conversion to EPA is 4–5 fold more efficient than ALA. However, some epidemiological studies suggest that ALA-containing oils (from seed oils such as flax; Linum usitatissimum L.)  may provide independent protection from cardiovascular disease.
Numerous studies show the biological impact of EPA and DPA. EPA reduces AA metabolism through several mechanisms including decreasing AA mobilization from membrane phospholipids, competition for cylooxygenase and 5-lipoxygenease and reducing the expression of AA metabolizing enzymes and proinflammatory cytokines. Additionally, EPA can serve as a substrate for prostaglandin formation generating “3-series” prostaglandin products including PGD3, PGE3, PGF3α, PGI3, and TxA3 and “5 series” leukotriene products including LTB5 and LTC5. Reduced asthma symptoms with n-3 PUFA ingestion have been shown to be related to 5-series leukotriene production . With regard to inflammation, DPA is beginning to receive attention. DPA is converted to 11-hydroxy-7,9,13,16,19-DPA and 14-hydroxy7,10,12,16,19-DPA, which inhibit aggregation of platelets and contain 10-fold greater capacity to elicit endothelial cell migration than EPA, a biological process critical to wound healing [22, 23]. There were no changes in plasma levels of DHA, likely reflecting the poor bioconversion of EPA to DHA.
Previous studies have shown that GLA-containing oils such as BO have the potential to increase circulating AA which could enhance inflammation and platelet aggregation through increased thromboxane formation . However, there were no changes in AA levels as a result of BO/EO supplementation. It is possible that the observed increase in EPA resulting from of the botanical combination is a feedback inhibitor of AA production via the ∆5 desaturation step. EPA has been demonstrated to inhibit the in vivo and in vitro desaturation of DGLA to form AA [25, 26]. In any event, the BO/EO combination led to an increase in three 20–22 carbon PUFAs, DGLA, EPA and DPA that have been demonstrated to inhibit AA metabolism and attenuate inflammation without increasing circulating levels of AA.
The final objective of this paper was to determine whether these botanical oil combinations had the capacity to inhibit leukotriene generation from two inflammatory cells, basophils and neutrophils, isolated from subjects with mild asthma who had supplemented their diet with BO/EO. Prior studies of dietary supplementation with GLA have demonstrated a reduction in ex vivo leukotriene generation in whole blood or neutrophils stimulated with calcium ionophore A23187 or with zymosan [13, 26, 27]. Basophils , IgE , and cysteinyl leukotrienes  have been strongly implicated in the pathobiology of asthma. We therefore assessed the effects of BO/EO combinations on the generation of cysteinyl leukotrienes from basophils stimulated through the high affinity IgE receptor, a physiologically relevant stimulus for asthma. Significant inhibition of basophil cysteinyl leukotriene generation was noted within one week of dietary supplementation (Figure 4). Interestingly, the time dependence of this inhibition varied between groups. The least robust inhibition was observed in Group 4, in which subjects received the lowest dose of SDA. Although there is considerable variation in the extent of inhibition of ex vivo leukotriene generation among individuals and the groups were relatively small, the between group variation was statistically significant. Furthermore, a comparable variation in supplementation-induced inhibition of leukotriene generation was observed in response to A23187- stimulation of neutrophils (Figure 6). The data therefore suggest that providing SDA in the diet contributed to the extent of inhibition of leukotriene generation, consistent with data showing that dietary supplementation with EPA leads to inhibition of ex vivo leukotriene generation [31–33].
In a receptor-independent manner, A23187 robustly stimulates human neutrophils to elicit maximal generation of LTB4, the product of leukotriene biosynthesis in neutrophils. Utilization of this stimulus allows an assessment by RP-HPLC of the non-enzymatic degradation products of the proximal intermediates of leukotriene biosynthesis. Dietary supplementation with BO/EO combinations led to a significant inhibition of A23187-stimulated leukotriene generation (Figure 6) that was almost as great as the inhibition of basophil cysteinyl leukotriene generation (Figure 4). We recently reported that dietary supplementation with BO and fish oil led to reduced expression of phosphatidylinositol 3-kinase, a key signaling molecule, in circulating mononuclear cells . It is therefore possible that the inhibition of cysteinyl leukotriene generation that we observed in basophils was due, at least in part, to inhibition of signaling through FcϵRI. However, the inhibition of leukotriene biosynthesis in neutrophils stimulated through A23187, a receptor independent stimulus, argues for a more direct effect of BO and EO on leukotriene biosynthesis.