论文标题:植物乳杆菌洗涤细胞转化生成共轭亚油酸的研究
The Production of Conjugated Linoleic Acid by Washed Cells of Lactobacillus Plantarum
论文作者 柴秋儿
论文导师 孙君社,论文学位 硕士,论文专业 食品科学与工程
论文单位 中国农业大学,点击次数 102,论文页数 62页File Size3912k
2005-03-01论文网 http://www.lw23.com/lunwen_926414402/ 植物乳杆菌;洗涤细胞;共轭亚油酸;亚油酸;豆油
Lactobacillus plantarum; washed cells; conjugated linoleic acid; linoleic acid; soybean oil
植物乳杆菌Lactobacillus plantarum LT2-6为实验室筛选所得,具有转化亚油酸(LinoleicAcid, LA)生成共轭亚油酸(Conjugated Linoleic Acid, CLA)的催化能力。乳杆菌LT2-6在适宜条件下培养24 h后离心,所得菌体用生理盐水或缓沖液洗涤离心收集得LT2-6的洗涤细胞,利用该乳杆菌的洗涤细胞即完整细胞能有效转化生成CLA。 本论文主要研究了LT2-6洗涤细胞的催化性能及其转化生成CLA的途径,并借助气相色谱对最终产物进行了脂肪酸成分检测。 研究结果表明:在以游离亚油酸为底物时,LT2-6的洗涤细胞在优化的反应条件下(0.1M磷酸缓冲液,pH 7.0,30℃,120 r/min)具有较强的催化能力,10%(w/v)的洗涤细胞在48 h内从6mg/mL亚油酸中转化生成4.628mg/mL CLA,转化率达77.1%;20%(w/v)的洗涤细胞在64 h内从15mg/mL亚油酸中转化生成7.861mg/mL CLA,转化率达52.4%;气相色谱结果显示产物CLA主要为cis-,trans-9, 11-CLA。在胰脂酶协助下,LT2-6的洗涤细胞(15%,w/v)在优化条件下(0.1M柠檬酸缓冲液,pH 6.0,30℃,120 r/min,72 h)能将豆油(15mg/mL)中以甘油酯形式存在的亚油酸有效转化为CLA,基于豆油中的亚油酸含量其转化率可达54%。 论文还研究了固定化L.plantarum LT2-6细胞的催化性能。研究发现,采用海藻酸钠包埋法所得的固定化细胞能保留较好的催化能力。在优化条件下即0.6g细胞/g海藻酸钠载体、50mM的Tris-HCl缓冲液(pH 8.0)和37℃,固定化细胞在24 h内能从lmg/mL亚油酸转化生成0.271mg/mL CLA,转化率为27.1%。对固定化细胞循环使用的研究表明,pH 7.5和25℃是固定化细胞循环反应生成CLA的有效pH值和有效温度。有机体系中细胞转化反应的初探实验显示细胞的催化能力与体系中的水分含量有关。
Lactobacillus plantarum LT2-6 was screened as a bacterium having high catalysis to isomerize Linoleic Acid (LA) into Conjugated Linoleic Acid (CLA). After cultivated in MRS medium for 24 h, LT2-6 was harvested by centrifugation, washed with 0.85% NaCl_2 or the right buffer, centrifugated again, and then used as washed cells in the following reaction. The whole cells (or the washed cellls) of LT2-6 were selected as a potent CLA producer from LA in the reaction.In this paper, the catalysis ability of the cells and the conditions for CLA production from LA and soybean oil were investigated using washed cells of LT2-6, and fatty acid analysis of the final product was also carried out by gas chromatography.The study clarified that washed cells of LT2-6 showed effective bioconversion with the free acid form of LA under optimal reaction conditions (0.1M sodium phosphate buffer , pH7.0, 30℃, 120r/min), 4.628 mg/mL CLA was produced from 6 mg/mL LA in 48 h by the catalyst of 10% (w/v) washed cells with a 77.1% yield as to LA; 7.861 mg/mL CLA was produced from 15 mg/mL LA in 64 h by the catalyst of 20% (w/v) cells with a 52.4% yield as to LA; the majority of resultants were cis-,trans-9,ll-CLA. Then the research of cooperation effect of porcine pancreatic lipase and washed cells on CLA production from soybean oil was carried out. The result showed that with the aid of porcine pancreatic lipase, under the optimal conditions (0.1M sodium citrate buffer, pH6.0, 30℃, 120r/min), with 15 mg/mL soybean oil as the substrate and 15% (wet cell, w/v) washed cells as the catalyst, the production of CLA reached a maximum (4.39 mg/mL) at 72 h with a 54% yield as to linoleic acid contained in the soybean oil.Research of different cell immobilizing technologies and bioconversion activities of the immobilized cells was measured in this paper. The experiment results showed that the immobilized cells entrapped by embedding medium of sodium alginate gel under the optimum embedding condition kept the satisfying catalysis and efficient bioconversion; the optimized conditions were 0.6g of cells/(g of carrier), 50 mM Tris-HCl buffer system (pH 8.0) and 37℃, respectively. Under the optimal conditions, the immobilized cells produced 0.271 mg/mL CLA from 1 mg/mL LA in 24 h with a yield of 27.1%. The CLA-producing ability of reused cells was investigated over five reused reactions and was maximal at pH 7.5 and 25℃. The pilot study of CLA production in organic media showed that the biocatalysis of washed cell was related to the water content in this special system.
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