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.

中文翻译：

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隐甲藻中均匀碳同位素标记的二十二碳六烯酸的生物合成。

 抽象的

大脑中富含的二十二碳六烯酸 (DHA) 在羟基自由基的攻击后可以产生许多重要的降解产物，已知其具有营养保健和神经保护作用。氧化应激是阿尔茨海默病 (AD) 的一个常见特征。因此，统一放射性标记的DHA在体内外研究DHA氧化命运方面发挥着重要作用。然而，碳同位素标记的 DHA 目前尚未上市。异养微藻Crypthecodinium cohnii ( C. cohnii ) 已被确定为 DHA 的高产者。在本研究中， C. cohnii的生长速率和 DHA 产量在新的成分确定培养基中进行了优化，并从 U- ¹³ C-葡萄糖生物合成 U- ¹³ C-DHA 和从 U- ¹⁴ C-DHA 生物合成 U- ¹⁴ C-DHA通过 HPLC-MS/MS 分析 C-葡萄糖。在 300 μL 批次中生产了约 40 nmole 的 U- ¹³ C-DHA，其同位素纯度高达 96.8%，在 300 μL 批次中生产了约 0.23 μCi 的 U- ¹⁴ C-DHA，其显着比活性为 5-6 Ci/mol。 300 µL 批次。结果发现， C. cohnii在该限定培养基中于 25 °C 时具有最佳生长和 DHA 积累 (C/N = 10)。首次建立了U- ¹³ C-DHA和U- ¹⁴ C-DHA生物合成的高效方案，为AD中DHA氧化降解产物的分析提供了基础支持。