荷荷巴油提取
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50 mL cultures were collected and centrifuged at 8000g for 5 min. Cell pellets were
then washed once with 10 mL of distilled water and lyophilized at -20 ℃ for 1 days.
Lyophilized cells were then weighed to obtain the dry cell weight (DCW). To
trans-esterify total fatty acids, 1 mL of 3 N HCl in methanol (Sigma) and 0.1 mL
chloroform were added to lyophilized cells. After incubation at 70℃for 3h, the
reaction mixture was cooled to room temperature, and 2 mL of saturated NaCl
solution was added followed by 15 s of vortex. Two milliliters of hexane were added,
and the reaction was agitated by vortex for 15 s. After a brief centrifugation step at
3000g for 1min, the hexane(top) layer containing fatty acid methyl esters (FAME)
was then analyzed on gas chromatography–mass spectrometry (GC–MS) using an
HP 6890 Series GC with an Agilent 5973 Network MSD equipped with a DB5
column (Thermo). The GC program was as follows: The initial temperature of 40 ℃
was maintained for 3 min, then ramped to 250℃ at a rate of 20 ℃/min and held
there for 5 min. The lipid content (%) is calculated as total fatty acid amount (mg) per
dry cell weight (mg)X100.
50mL的培养物中收集，并在8000转速离心5分钟。细胞沉淀物再洗涤一次，用10ml的蒸馏水并冷冻干燥，在-20℃进行1天。然后冷冻干燥细胞，称重，以获得干细胞重量（DCW）。到反式酯化总脂肪酸，1毫升在3N HCl的甲醇溶液（Sigma）和0.1毫升氯仿加入到冷冻干燥的细胞。孵育于70℃3小时后，将反应混合物冷却至室温，并加入2mL的饱和的NaCl溶液，接着加入涡旋15秒。两毫升己烷加入，并将反应物搅拌通过涡旋15秒。在3000g1分钟的简短离心步骤后，将己烷（顶）层含有脂肪酸甲基酯（FAME），然后使用HP6890系列气相色谱仪的Agilent5973网络MSD气相色谱 - 质谱（GC-MS）分析配备了DB5柱（热）。 GC的程序如下：在40℃的初始温度下保持3分钟，然后升温到250℃以20℃/ min的速率和5分钟，保持在那里。脂质含量（％）被计算为每干细胞重量（mg）×100总脂肪酸量（mg）。
