Abstract
Docosahexaenoic acid (DHA) enriched in brain can yield many important degradation products after the attack of hydroxyl radicals, which is known to serve as a nutraceutical and neuroprotective effects. Oxidative stress is a commonly observed feature of Alzheimer’s disease (AD). Therefore, uniformly radiolabeled DHA plays an important role in studying the oxidative fate of DHA in vivo and vitro. However, carbon isotope labeled DHA isn’t commercially available now. The heterotrophic microalgae Crypthecodinium cohnii (C. cohnii) has been identified as a prolific producer of DHA. In this study, the growth rate and DHA production in C. cohnii were optimized in a new defined media, and the biosynthesis of U-13C-DHA from U-13C-glucose and U-14C-DHA from U-14C-glucose were analyzed by HPLC–MS/MS. Approximately 40 nmoles of U-13C-DHA with higher isotopic purity of 96.8% was produced in a 300 μL batch, and ~ 0.23 μCi of U-14C-DHA with significant specific activity of 5–6 Ci/mol was produced in a 300 μL batch. It was found that C. cohnii had the optimal growth and DHA accumulation at 25 °C in this defined media (C/N = 10). An efficient protocol for the biosynthesis of U-13C-DHA and U-14C-DHA were set up firstly, which provides the basic support for the analysis of oxidative degradation products of DHA in AD.
Highlights
Docosahexaenoic acid (DHA) as a predominant ω-3 polyunsaturated fatty acid (ω-3 PUFA) is known to play multi-functional roles in brain diseases (Sun et al 2018; Harauma et al 2017)
The total ion current of purified U-13C-DHA was got at m/z = 324–352 in enhanced mass spectrometer (EMS) mode
In this experiment, C. cohnii was cultivated in the same conditions, but carbon isotope labeling defined media were used for the biosynthesis of U-13C-DHA and U-14C-DHA
Summary
Docosahexaenoic acid (DHA) as a predominant ω-3 polyunsaturated fatty acid (ω-3 PUFA) is known to play multi-functional roles in brain diseases (Sun et al 2018; Harauma et al 2017). PUFAs, such as arachidonic acid (ARA) and DHA, are abundant in brain and especially vulnerable to the attack of hydroxyl radical, which can induce the production of many degradation products (Corsinovi et al 2011; Nowak 2013). ARA from membrane phospholipids can be released by phospholipase A2 in cytoplasm, DHA is connected to action of the phospholipase A 2. DHA can make enzymatic conversion by 15-lipoxygenase to form important lipid mediators including the resolvins and neuroprotectins (Strokin et al 2004). DHA can make non-enzymatic conversion by the oxygen free radicals (ROS), which would induce the production of 4-hydoxy-2-hexenal (HHE), an aldehyde derivative of ω-3 PUFA oxidation which
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