费氏丙酸杆菌谢氏亚种转化芝麻油生成共轭亚油酸的研究

对费氏丙酸杆菌谢氏亚种CGMCC1.2231菌体细胞转化芝麻油生成共轭亚油酸进行了研究。首先通过单因素实验,研究了诱导产酶阶段亚油酸的添加量,以及转化反应温度、缓冲体系pH值、乳化剂种类对共轭亚油酸生成量的影响,进而通过正交实验得到最佳转化条件。在诱导产酶培养阶段加入0.15%(v/v)的亚油酸,转化温度30℃,反应体系pH值为7.4,使用牛血清白蛋白乳化底物的条件下,共轭亚油酸的最高生成量可达45.682mg/mL。     

一株植物乳杆菌转化生成共轭亚油酸的特性研究

报道了一株植物乳杆菌(L.plantarumLT2-6)发酵转化亚油酸(LA)生成共轭亚油酸(CLA)的特性研究。微氧环境有利于CLA的生成;温度为37℃、初始pH为7.0、底物LA浓度为0.075%时,菌体生长及CLA生成量较高。时间曲线结果表明,接种后8h,CLA开始生成;发酵24h时,CLA生成量达到最高(0.29g/L),LA转化率为387%。生成的CLA产物主要为cis9,trans11/trans9,cis11-CLA。     

产共轭亚油酸鼠李糖乳杆菌的筛选与鉴定

从东北传统酸菜汁中筛选出具有产共轭亚油酸(CLA)能力的乳酸菌菌株15株,采用紫外分光度计法测定发酵液中共轭亚油酸产量,其中菌株A53.2的产量最高,在MRS培养基中添加0.4mg/mL亚油酸(LA),37℃发酵24h,CLA产量达14.91μg/mL。通过对菌体的形态特征和生化特性分析,并结合16SrDNA分子生物学以及系统发育分析鉴定菌株A53.2为鼠李糖乳杆菌。     

响应面法优化植物乳杆菌lp15-2-1产共轭亚油酸发酵条件

对植物乳杆菌(Lactobacillus plantarum)lp15-2-1 转化亚油酸生成共轭亚油酸(CLA)发酵工艺进行研究。通过单因素试验确定最佳接种量、培养温度、初始pH 值、培养时间、亚油酸添加时间、亚油酸质量浓度。采用响应面Box-Benhnken 试验设计,建立初始pH 值、培养温度和亚油酸质量浓度的二次多项式回归方程模型,所得植物乳杆菌发酵产共轭亚油酸的最佳参数为：温度30℃、亚油酸质量浓度0.21mg/mL、初始pH6.3,此条件下,CLA得率为44.77μg/mL,CLA 理论得率为45.326μg/mL,转化率为21.32%,与优化前转化率7.78% 相比,有了很大提高。     

一株嗜酸乳杆菌合成共轭亚油酸的条件优化

以嗜酸乳杆菌(Lactobacillus acidophilus)1.1854为出发菌株,经紫外诱变和诱导处理,获得1株共轭亚油酸(CLA)高产菌株,通过五因素二次正交旋转组合设计对该菌株合成共轭亚油酸的条件进行了优化。其优化条件为:培养温度37℃、pH值4.5～5.5、LA浓度0.1%、接种量5%～7%、培养时间24h,此时CLA产率达31.2%。     

产共轭亚油酸干酪乳杆菌的诱变选育

共轭亚油酸(CLA)是世界公认的具有抗癌、减肥和抗动脉硬化作用的脂肪酸.以共轭亚油酸的生成菌株干酪乳杆菌为出发菌株,经紫外诱变处理,筛选出一株高产共轭亚油酸的菌株,并优化其培养条件,在亚油酸添加量0.05%,乳糖添加量2%,接种量4%,初始pH6.5,37℃发酵36 h时,共轭亚油酸生物合成量可达394.1μg/mL.     

保加利亚乳杆菌诱导产亚油酸异构酶条件研究

亚油酸异构酶可由保加利亚乳杆菌经诱导产生,可以将亚油酸(LA)转化为共轭亚油酸(CLA)。本实验对诱导保加利亚乳杆菌产亚油酸异构酶的条件进行研究,利用紫外和气质联用仪(GC-MS)检测所生成的CLA。结果表明：在培养基中添加1.5‰(V/V) LA 时所产酶的共轭亚油酸转化率最高;温度为36℃,培养36h 为较适的培养条件;单独添加0.1%(m/V)的乳糖或0.1%(m/V)的氯化钠有利于诱导产酶;在培养基中直接添加LA 的效果优于培养3至12h 后再进行添加。诱导所产酶可将LA 转化为CLA,且含有9c,11t-CLA 异构体。     

一株嗜酸乳杆菌产共轭亚油酸条件的优化

共轭亚油酸(conjugated linoleic acid,CLA)是具有抗癌、减肥等许多生理功能的天然脂肪酸.生物法合成共轭亚油酸无毒且异构体单一,但目前产量较低.为提高CLA产量,以嗜酸乳杆菌(Lactobacillus acidophilus)为出发菌株,优化其培养条件.结果表明:发酵液初始pH为6.5,红花油添加量为0.15%,接种量5%,发酵温度为37°C,发酵时间为60h时最有利于共轭亚油酸的合成,此时CLA的合成最为94.7μg/mL.     

植物乳杆菌发酵脱脂牛乳生成共轭亚油酸过程的研究

为提高发酵牛乳共轭亚油酸(CLA)含量,以植物乳杆菌(Lactobacillus plantarum LT2-6)发酵添加亚油酸的脱脂牛乳,并对其生成CLA的过程进行了研究。结果表明,接种后6h CLA开始生成,24h CLA生成量达到最高,再延长发酵时间后CLA生成量缓慢下降。接种3～12h,牛乳滴定酸度快速增加,12h后滴定酸度增加缓慢。发酵18h时CLA的生成速率最大;24h时,牛乳pH降到4.0,CLA生成基本停止。牛乳发酵过程中,pH下降是影响植物乳杆菌LT2-6转化LA生成CLA的主要因素。脱脂牛乳中加入LA,经植物乳杆菌LT2-6发酵、脂肪酸萃取和甲酯化,并用气相色谱检测,结果显示:生成的CLA主要为cis9、trans11/trans9和cis11-CLA。     

乳酸菌UV_3-9包埋固定化技术发酵产共轭亚油酸

采用包埋法将乳酸菌UV3-9固定化处理后,对该固定化乳酸菌小球进行了发酵生产共轭亚油酸(CLA)的研究。结果显示,固定化小球的最佳直径为2～3 mm,固定化小球在氯化钙和硼酸混合液中需浸泡过夜;固定后的小球发酵产CLA最佳条件为:发酵温度37℃,初始pH 6.5,发酵时间28 h,底物LA添加量为0.3%,CLA最大产量达到92.061μg/mL。该固定化乳酸菌小球已完成了7批次的共轭亚油酸发酵,其产量均在80.476μg/mL以上。而且固定化UV3-9的CLA发酵产量是游离UV3-9的1.28倍。     

Effects of Linoleic Acid on Conjugated Linoleic Acid Production by Planktonic Rumen Bacteria from Grain-fed Cows

Ruminal conjugated linoleic acid (CLA) production from linoleic acid (LA) was characterized in vitro. Rumen bacteria from grain‐fed cows were more active in BH than those from hay‐fed cows. Particleassociated bacteria produced more hydrogenated products leaving less CLA than the planktonic bacteria (P < 0.05). CLA production by planktonic bacteria did not always correlate to LA given; longer incubations generally decreased CLA concentration and increased c9, t11/t10, c12 ratio, especially at higher LA concentrations. The preincubated cells to LA produced more CLA than the unexposed ones and the increase was more evident with c9, t11 CLA (P < 0.05). This study provides insight into how cattle diet and LA feedings affect ruminal CLA production.     

Production of free conjugated linoleic acid by Lactobacillus acidophilus and Lactobacillus casei of human intestinal origin

A gas chromatographic procedure was used for analysis of conjugated linoleic acid (CLA) isomers cis-9, trans-11-octadecadienoic; trans-10, cis-12 octadecadienoic; and trans-9, trans-11-octadecadienoic (c9t11, t10c12, t9t11) produced by lactobacilli. Four different cultures, two strains each of Lactobacillus acidophilus and Lactobacillus casei were tested for their ability to produce CLA from free linoleic acid in MRS broth supplemented with linoleic acid. Different concentrations of linoleic acid (0, 0.05, 0.1, 0.2 and 0.5 mg/ml) were added to MRS broth, inoculated with the lactobacilli, and incubated at 37 degrees C. Viable counts and amounts of individual isomers of CLA (c9t11, t10c12, t9t11) were measured at 0, 24, 48, and 72 h. All the cultures were able to produce free CLA in media supplemented with linoleic acid. Maximum production of CLA (80.14 to 131.63 microg/ml) was observed at 24 h of incubation in broth containing 0.02% of free linoleic acid. No significant (P > 0.05) increases in total CLA levels were observed after 24 h of incubation. The ability of the cultures to produce CLA in skim milk supplemented with 0.02% free linoleic acid also was studied. In this medium, the total amounts of free CLA after 24 h of incubation ranged from 54.31 to 116.53 microg/ml. The use of lactic acid bacteria able to form free CLA in cultured dairy products may have potential health or nutritional benefits. Free CLA in the products likely would be more readily available for absorption from the digestive tract than if it were incorporated into the cells of the starter culture.     

生物转化法生产共轭亚油酸及其研究进展

共轭亚油酸（CLA）是一种具有多种重要生理功能的天然脂肪酸，具有抗癌、抗动脉粥样硬化、抗糖尿病、调节免疫等多种功能。论述了利用生物转化法合成共轭亚油酸的生产菌株、培养条件和培养模式以及利用基因工程等菌株改良手段以提高生产效率的国内外研究进展；并简述了生物法工业化生产共轭亚油酸的前景。     

产共轭亚油酸乳酸菌的筛选及生产条件的研究

从分离到的乳酸菌中筛选到1株共轭亚油酸高产菌株,乳杆菌L1(Lactobacillus sp.),在MRS培养基上,乳杆菌L1产生共轭亚油酸的最适亚油酸的浓度是0.1%(v/v),最适培养时间是42h,参与共轭亚油酸形成的酶是胞内酶,并且可能是由多种酶共同作用的结果.建立了共轭亚油酸的紫外光谱扫描和高效液相色谱检测方法.     

产共轭亚油酸植物乳杆菌的筛选、鉴定与诱变

从传统泡菜中筛选得到1 株乳酸菌lp15 能够合成共轭亚油酸(CLA),经16S rRNA 全序列分析法和API 系统鉴定法鉴定为植物乳杆菌(Lactobacillus plantarum)。利用人工诱变方法,以lp15 为出发菌株,采用紫外线、硫酸二乙酯(DES)依次诱变处理,经进一步液体发酵复筛获得多株突变菌株,CLA合成能力较出发菌株提高了29.3%～52.2%。其中lp15-2-1 突变菌株为CLA 生成能力最高菌株,MRS 培养液中添加0.2mg/mL LA 培养48h,CLA 产量达30.13μg/mL。经气相色谱(GC)检测分析,产物中cis9, trans11- 共轭亚油酸(c9, t11-CLA)占76.5%,trans10, cis12-共轭亚油酸(t10,c12-CLA)占23.5%。     

Production of conjugated fatty acids by lactic acid bacteria

Conjugated fatty acids have attracted much attention as a novel type of biologically beneficial functional lipid. Some isomers of conjugated linoleic acid (CLA) reduce carcinogenesis, atherosclerosis, and body fat. Considering the use of CLA for medicinal and nutraceutical purposes, a safe isomer-selective process is required. The introduction of biological reactions for CLA production could be an answer. We screened microbial reactions useful for CLA production, and found several unique reactions in lactic acid bacteria. Lactic acid bacteria produced CLA from linoleic acid. The produced CLA comprised a mixture of cis-9,trans-11-octadecadienoic acid (18:2) and trans-9,trans-11-18:2. Lactobacillus plantarum AKU 1009a was selected as a potential CLA producer. Using washed cells of L. plantarum AKU 1009a as a catalyst, CLA production from linoleic acid reached 40 mg/ml under the optimized conditions. The CLA-producing reaction was found to consist of two successive reactions, i.e., hydration of linoleic acid to 10-hydroxy-12-octadecenoic acid and dehydrating isomerization of the hydroxy fatty acid to CLA. On the basis of these results, the transformation of hydroxy fatty acids by lactic acid bacteria was investigated. Lactic acid bacteria transformed ricinoleic acid (12-hydroxy-cis-9-octadecenoic acid) to CLA (a mixture of cis-9,trans-11-18:2 and trans-9,trans-11-18:2). Castor oil, which is rich in the triacylglycerol form of ricinoleic acid, was also found to act as a substrate for CLA production by lactic acid bacteria with the aid of lipase-catalyzed triacylglycerol hydrolysis. L. plantarum AKU 1009a produced conjugated trienoic fatty acids from alpha- and gamma-linolenic acid. The trienoic fatty acids produced from alpha-linolenic acid were identified as cis-9,trans-11,cis-15-octadecatrienoic acid (18:3) and trans-9,trans-11,cis-15-18:3. Those produced from gamma-linolenic were cis-6,cis-9,trans-11-18:3 and cis-6,trans-9,trans-11-18:3. The conjugated trienoic fatty acids produced from alpha- and gamma-linolenic acid were further saturated by L. plantarum AKU 1009a to trans-10,cis-15-18:2 and cis-6,trans-10-18:2, respectively.     

瘤胃中干酪乳杆菌的亚油酸异构酶基因的克隆与表达

以从黄牛瘤胃中分离到的一株具有将亚油酸转化为c9,t11-共轭亚油酸(conjugated linoleic acid,CLA)的干酪乳杆菌(Lactobacillus casei)Fx为出发菌株,提取其基因组DNA,聚合酶链式反应(polymerase chain reaction,PCR)扩增得到1 700 bp大小的亚油酸异构酶(linoleic acid isomerase,LAI)基因片段,将该基因片段纯化后进行TA克隆,得到重组质粒p UCm-T-LAI,将重组质粒p UCm-T-LAI和表达质粒p ET-Dsb A同时进行双酶切,连接得到重组表达载体p ET-Dsb A-LAI,经PCR鉴定和酶切后,将重组表达载体转化到大肠杆菌BL21中,得到具有LAI活性的重组菌株,能将亚油酸转化为c9,t11-CLA,表明从Lactobacillus casei Fx成功克隆LAI,该研究将有助于深入了解不同瘤胃细菌特异性合成不同CLA异构体的LAI基因差异。     

生物合成共轭亚油酸菌种的筛选与鉴定

从传统泡菜和生牛乳中筛选出一株乳酸菌ZS2058能生物合成共轭亚油酸,经API系统鉴定为植物乳杆菌(Lactobacillusplantarum).该菌株在MRS培养基中将质量分数11.6%的亚油酸(1.024mg/mL)转化为共轭亚油酸,经气相色谱分析证实c9,t11 18∶2占75.9%,t10,c12 18∶2占24.1%.