Relationship between dietary phospholipid classes and neutral lipid absorption in newly-weaned turbot, shape Scophthalmus maximus

A 28-day feeding trial was conducted for comparing the effect of different dietary phospholipid (PL) classes on the growth of post-larval turbot and on the incorporation of dietary neutral lipid fatty acids into their body lipids. Prior to the experiment the turbot were weaned for one week on a PL-free diet. The nine experimental diets were isolipidic and contained an equal amount of highly unsaturated fatty acids in the form of ethyl esters. They differed by their PL content (0, 1 or 2%) and by the PL class composition of the added soybean PL fractions.Compared to the PL-free diet, diets enriched with phosphatidylcholine (PC) resulted in a better growth, a higher triglyceride content (% body dry matter) and increased levels of docosahexaenoic acid (% total fatty acids) in each of the examined body lipid classes (neutral lipid, phosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol). The effects of the other soybean PL fractions were less explicit than those noted for soybean PC.The results support the idea that dietary PC plays a role in the intestinal absorption of neutral lipid fatty acids. This might, at least partially, explain the superiority of PC for enhancing growth. Abbreviations: DHA – docosahexaenoic acid (22:6n-3); EPA – eicosapentaenoic acid (20:5n-3); HUFA – highly unsaturated fatty acid; PA – phosphatidic acid; PC – phosphatidylcholine; PE – phosphatidylethanolamine; PI – phosphatidylinositol; PL – phospholipid; PS – phosphatidylserine; PUFA – polyunsaturated fatty acid.     

梨磷脂酰胆碱转移蛋白基因的克隆及其表达分析

据转录组测序结果,结合RACE(cDNA末端快速扩增技术)和Genome walking(染色体步移技术)获得了梨磷脂酰胆碱转移蛋白基因的完整读码框序列,其开放阅读框长为1 323 bp,编码的440个氨基酸序列与葡萄磷脂酰胆碱转移蛋白质的同源相似性达73%,其具有START domain(启动域)、ZnF_TAZ(TAZ锌指结构)、DCD(双氰胺)保守结构域,将梨磷脂酰胆碱转移蛋白基因命名为PePCTP1。PePCTP1基因与GFP融合,构建植物表达载体,洋葱表皮亚细胞定位观察结果显示,PePCTP1基因编码的蛋白质分布在细胞膜上。用激素生长素(IAA)、脱落酸(ABA)、水杨酸(SA)、赤霉素(GA)、细胞分裂素(6-BA)及NaCl诱导处理,发现与对照(CK)相比,SA和GA处理后PePCTP1基因表达量显著上升;NaCl和IAA处理后PePCTP1基因的表达量随时间逐渐上升;而ABA和6-BA处理不能增加目的基因的表达量。据此推测PePCTP1可能参与梨的逆境胁迫抗性反应。     

Role of molecular species catabolism in the temperature-induced restructuring of phosphatidylcholines in liver microsomes of thermally-acclimated rainbow trout (Oncorhynchus mykiss)

Most previous studies of the temperature-induced restructuring of phospholipid molecular species composition have examined steps in the biosynthesis of phospholipids to explain the accumulation of unsaturated fatty acids in membranes of cold-acclimated poikilotherms. In contrast, the present study explores the role of phospholipases in this restructuring process by determining the rates of degradation of specific molecular species of phosphatidylcholine, using enzymes (microsomes) freshly isolated from the liver of rainbow trout. (Oncorhynchus mykiss) acclimated to either 5° or 20°C. The substrate preparation employed to assay phospholipase activity possessed a range of molecular species, all radiolabeled with 1-¹⁴C-palmitic acid at thesn-1 position, similar to that present in native trout liver microsomes. After defined periods of incubation (120 and 240 min at 5°C; 60 and 120 min at 20°C), phospholipids were extracted from the reaction mixture and the distribution of radioactivity among the molecular species of phosphatidylcholine was determined by HPLC/liquid scintillation counting. In general, molecular species catabolism was not significantly influenced by either assay or acclimation temperature. Only in 20°C-acclimated fish did a reduction in assay temperature (from to 20 to 5°C) result in significantly increased proportions of radioactivity being recovered in one polyunsaturated fatty acid-containing species (16:0/22:6-PC). It is concluded: 1) that phospholipase specificity, assayed under conditions approximating thosein situ, is not significantly influenced by temperature; and 2), that the increased proportions of unsaturated fatty acid-containing molecular species of phosphatidylcholine observed at low temperatures must reflect the specificity of biosynthetic rather than degradative processes.     

响应面法提取南极磷虾磷脂及其分子种组成的测定

南极磷虾(Euphausia superba)生物储量巨大,含有丰富的磷脂酰胆碱(phosphatidylcholine,PC),而PC是生物生长的必需磷脂。现今南极磷虾的研究主要针对磷虾油和蛋白,尚未见有PC分子组成的系统研究报道。该研究使用超声辅助Bligh Dyer脂质提取法提取南极磷虾中磷脂,并采用Box-Benhnken设计原理优化提取工艺,以正离子模式下胆碱碎片(m/z 184)为特异性碎片,利用鸟枪脂质组学法鉴定南极磷虾组织中的PC分子组成。结果表明,南极磷虾磷脂的最佳提取条件为超声时间21.0 min、超声温度30℃、溶剂用量3.1 mL,此条件下磷脂实际提取率的平均值为(7.59±0.03)%。利用鸟枪脂质组学法鉴定出南极磷虾中共含有36种PC分子,其中EPA/DHA脂肪酸链含量占47.75%,质量分数为269.82μg·g~(-1)。研究表明南极磷虾组织中含有丰富的EPA和DHA,而磷脂状态存在的ω-3脂肪酸具有较高的利用效率,这为开发南极磷虾类营养保健产品提供数据支持和理论基础。     

乙醇提取冷冻结晶去杂精制磷脂酰胆碱

该文研究了在低温条件下，用乙醇作为溶剂，通过冷冻结晶去杂，从粗制大豆磷脂中精制磷脂酰胆碱的方法。通过单因素试验确定了影响磷脂酰胆碱纯度的相关因素。用响应面试验对精制条件进行了优化，结果表明冷冻时间、冷冻温度、液料比都对磷脂酰胆碱的纯度产生一定的影响。其中冷冻时间，冷冻温度的影响最为显著。经响应面分析优化得到了最佳工艺条件为:温度为-20℃、冷冻时间为23.56 h、液料比为8.73∶1(V/m)。在此条件下得到磷脂酰胆碱的预测纯度为70.73%。经过验证实验和高效液相法检测，实测在此条件下：磷脂酰胆碱的纯度为70.1%,得率为23.0%。中试结果为：磷脂酰胆碱的纯度为68.8%,得率为22.7%。表明此方法简单可行，适宜于工业化生产。     

Purification and characterization of phospholipase A2 from the pyloric caeca of red sea bream, Pagrus major

A phospholipase A₂ was purified 55,000-fold in a yield of 10% from the lipid-free extract of powder of the pyloric caeca of red sea bream to near homogeneity by sequential column chromatography on S-sepharose fast flow, butyl-cellulofine, Asahipak ES-502C cation-exchange HPLC, TSK gel G3000SW gel-filtration HPLC, and Asahipak ODP-50 reversed-phase HPLC. The final preparation showed a single band with the apparent molecular mass of 14 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and an estimated specific activity was 717 µmol min^-1 mg^-1 protein. The purified enzyme had a pH optimum in the range of pH 8.0–9.0 and required the presence of both 8 mM of Ca²⁺ and from 2 to 10 mM of sodium deoxycholate for its maximal activity, using 2 mM of phosphatidylcholine as a substrate. The purified enzyme preferentially hydrolyzed the 2-acyl ester bonds of both phosphatidylglycerol and phosphatidylcholine in the presence of sodium deoxycholate, followed in order by phosphatidylethanolamine and phosphatidyl-serine. In contrast to porcine pancreatic PLA₂, pyloric caeca PLA₂ hydrolyzed mixed-micellar phosphatidylcholine substrate effectively, regardless of the kinds of bile salts used. These results indicate that Ca²⁺-dependent low molecular mass PLA₂, so called secretory PLA₂, occurs in the pyloric caeca of red sea beam.     

Dietary phosphatidylcholine requirements in larval and postlarval Penaeus japonicus Bate

Three nutritional experiments were carried out to investigate the effect of dietary phosphatidylcholine (PC) on the growth (length, weight and metamorphosis) and survival (rearing and stress) of larval and postlarval Penaeus japonicus Bate. Purified soybean phosphatidylcholine (SPC, 95% purity), hen egg-PC (EPC, 94% purity), and de-oiled soybean lecithin (DSL, PC 23% purity) were used as sources of dietary PC. The results indicated that optimal metamorphosis in larval P. japonicus was obtained at levels of 15–30 g kg^?1 of dietary SPC. Feeding trials with postlarval P. japonicus showed that low levels of dietary SPC (15 g kg^?1) were more beneficial than higher levels of SPC (30 g kg^?1) or DSL (65 g kg^?1), although early postlarvae (<3 mg dry weight) presented a higher requirement for PC than later stages. Shrimp fed the 150 g kg^?1 PC-supplemented diets exhibited significantly better resistance to salinity stress than those fed a PC-deficient diet. A high level of dietary soybean lecithin (65 g kg^?1 DSL), providing 15 g kg^?1 of dietary PC, was no more effective for young postlarvae of P. japonicus than 15 g kg^?1 of purified PC alone, which suggested that the other phospholipids (mainly phosphatidylethanolamine and phosphatidylinositol) in the soybean lecithin are not required for postlarval shrimp, at least when there is already an adequate source of PC.     

Effect of two choline sources on the performance of postlarval Marsupenaeus japonicus bate

The present study was conducted to clarify the effect and the interaction between two choline sources [choline chloride (CC) and soybean phosphatidylcholine (SPC)] on the performances of postlarval shrimp, Marsupenaeus japonicus. A 30‐day feeding experiment was conducted as a 2 × 2 factorial design. The postlarval shrimp were fed with four κ‐carrageenan micro‐bound diets that contained two levels of CC (0.6 and 1.2 g kg⁻¹) and two levels of SPC (20 and 40 g kg⁻¹). A significant (P < 0.05) interaction was also detected between both choline sources in terms of the weight gain, apparent feed efficiency ratio, apparent protein efficiency ratio and total lipid, free choline and phosphatidylcholine contents of the shrimp whole body except for survival, moisture, and protein contents of the whole body. At the lower dietary SPC (20 g kg⁻¹) level, increasing of CC level from 0.6 to 1.2 g kg⁻¹ significantly enhanced the above parameters, whereas such positive effect of dietary CC was diminished at the higher dietary SPC (40 g kg⁻¹) level. Standing on the different viewpoint, the effect of supplemental SPC was also affected with the dietary levels of CC.     

高压脉冲电场辅助溶剂提取蛋黄卵磷脂效果及品质保证

为提高对蛋黄卵磷脂(PC)的提取率,该文利用高压脉冲电场(PEF)辅助有机溶剂来进行提取试验。试验选定了影响PC提取率的3个主要因素:PEF场强、脉冲数和提取溶剂含水率,并采用Box-Behnken试验设计对提取工艺进行优化;并且,通过中红外光谱扫描分析了经PEF处理前后的PC分子结构变化。结果表明:PC提取的最佳工艺条件为场强39kV/cm,脉冲数31,提取溶剂含水率9%,其PC提取率比无PEF处理的传统有机溶剂提取法提高了10.2%;PEF对PC分子结构并没有显著影响,证实了PEF对食品营养成分破坏少,是一种温和的处理方式。该研究可为PEF技术应用于蛋黄卵磷脂的提取提供参考。