Bacterial Production of Conjugated Linoleic and Linolenic Acid in Foods: A Technological Challenge

Conjugated linoleic acid (CLA) and conjugated linolenic acid (CLNA) isomers are present in foods derived from ruminants as a result of the respective linoleic acid (LA) and α-linolenic acid (LNA) metabolism by ruminal microorganisms and in animals’ tissues. CLA and CLNA have isomer-specific, health-promoting properties, including anticarcinogenic, antiatherogenic, anti-inflammatory, and antidiabetic activity, as well as the ability to reduce body fat. Besides ruminal microorganisms, such as Butyrivibrio fibrisolvens, many food-grade bacteria, such as bifidobacteria, lactic acid bacteria (LAB), and propionibacteria, are able to convert LA and LNA to CLA and CLNA, respectively. Linoleate isomerase activity, responsible for this conversion, is strain-dependent and probably related to the ability of the producer strain to tolerate the toxic effects of LA and LNA. Since natural concentrations of CLA and CLNA in ruminal food products are relatively low to exert their health benefits, food-grade bacteria with linoleate isomerase activity could be used as starter or adjunct cultures to develop functional fermented dairy and meat products with increased levels of CLA and CLNA or included in fermented products as probiotic cultures. However, results obtained so far are below expectations due to technological bottlenecks. More research is needed to assess if bacterial production kinetics can be increased and can match food processing requirements.     

Conjugated linoleic and linolenic acid production kinetics by bifidobacteria differ among strains

There is great interest in conjugated linoleic acid (CLA) and conjugated linolenic acid (CLNA) isomers because of their supposed health-promoting properties. Therefore, the differences in production kinetics of CLA and CLNA isomers from linoleic acid (LA) and α-linolenic acid (α-LNA), respectively, by bifidobacteria were investigated. Laboratory fermentations, supplemented with LA or α-LNA in the fermentation medium, were performed with Bifidobacterium bifidum LMG 10645, Bifidobacterium breve LMG 11040, B. breve LMG 11084, B. breve LMG 11613, B. breve LMG 13194, and Bifidobacterium pseudolongum subsp. pseudolongum LMG 11595. Conversion of LA and α-LNA to CLA and CLNA isomers, respectively, started immediately upon addition of the substrate fatty acids. During the active growth phase, the c9, t11-CLA isomer and the putative c9, t11, c15-CLNA isomer were formed. Further fermentation resulted in a reduction in the concentration of c9, t11-CLA and c9, t11, c15-CLNA and the subsequent production of the t9, t11-CLA isomer and the putative t9, t11, c15-CLNA isomer, respectively. Modelling of the growth and metabolite data indicated differences in production kinetics among the strains. Some strains displayed a high specific conversion of LA and α-LNA despite poor growth, whereas other strains grew well but displayed lower conversion. Production of specific CLA and CLNA isomers by bifidobacteria holds potential for the production of functional foods and could contribute to their probiotic properties.     

共轭亚油酸生理功能及其合成方法

共轭亚油酸（CLA）为亚油酸（LA）的异构体,具有抗癌、降脂、调节免疫、抗粥样硬化等重要的生理功能,目前共轭亚油酸（CLA）的生产大都是利用碱催化亚油酸异构而成,但其异构体繁多。利用微生物转化生产的共轭亚油酸（CLA）,异构体较少,与天然食品中的CLA异构体组成相似,具有很好的商业前景。     

Systematic evaluation on the effectiveness of conjugated linoleic acid in human health

The term CLA (conjugated linoleic acid) corresponds to a mixture of positional and geometric isomers of linoleic acid. Two of these isomers (9c, 11t and 10t, 12c) have biological activity. The milk and dairy products are the most abundant source of conjugated linoleic acid, which refers to a group of positional and geometric isomers of CLA (CLA 18:2 cis-9, cis-12). The following research aims to approach aspects regarding the CLA, as well as its relationship with diseases. Conjugated linoleic acids have been studied for their beneficial effects in the prevention and treatment of many diseases, including obesity, cancer, diabetes, and cardiovascular diseases. Scientific information put together the physiological properties of CLA, which serves as inputs to claim their potential as functional ingredients to be used in the prevention and control of several chronic metabolic disorders.     

Concentration‐dependent antioxidant activities of conjugated linoleic acid and α‐tocopherol in corn oil

BACKGROUND: Antioxidants prevent rancidity (lipid peroxidation) and natural antioxidants, e.g., α‐tocopherol, likely provide additional value to oil‐based food products because of their health benefits. Conjugated linoleic acid (CLA) has potential health benefits and may exhibit antioxidant properties. The main aim of this study was to compare the antioxidant efficacy of α‐tocopherol, trans‐10, cis‐12‐CLA and cis‐9, trans‐11‐CLA (in graded concentrations) added to antioxidant‐stripped corn oil. RESULTS: As compared to α‐tocopherol, both CLA isomers displayed significant inhibition of corn oil lipid peroxidation induced by copper. Inhibition of thiobarbituric acid reactive substances (TBARS) were CLA concentration dependent for both isomers but with significant inhibition occurring at 0.1 and 1 ppm of CLA isomers or α‐tocopherol, respectively (P < 0.05). Graded concentrations of α‐tocopherol, and for both CLA isomers and time, had significant effects on TBARS formation (P < 0.0001). There were significant effects in interactions between graded concentrations and time for both CLA isomers (P < 0.0001) but not for α‐tocopherol (P > 0.05). CONCLUSION: CLA compounds could serve as useful food antioxidants and provide additional value because of their potential bioactivity in disease prevention. Copyright © 2007 Society of Chemical Industry     

Conjugated linoleic acid conversion by six Lactobacillus plantarum strains cultured in MRS broth supplemented with sunflower oil and soymilk

Six strains of Lactobacillus plantarum, isolated from traditional dairy products of minority nationalities, were evaluated for their ability to produce conjugated linoleic acid (CLA) from free linoleic acid in vitro. All the 6 strains were found to be capable of converting linoleic acid to CLA when using sunflower oil as substrate or during soymilk fermentation. The inhibitory effect of linoleic acid on the growth of the L. plantarum was also discussed. The production of CLA was increased with adding high concentration of substrate in sunflower oil and IMAU60042 produced the highest CLA both in sunflower oil and soymilk. The CLA was composted by 2 isomers: cis9, trans11-CLA and tran10, cis12-CLA, and cis9, tran s11-CLA covered the most part of the total CLA formed except for L. plantarum P8. The production of CLA was decreased during the storage of fermented soymilk. The CLA contents decreased significantly in the first week, also more quickly in 2 wk. Especially, tran10, cis12-CLA decreased more rapidly than cis9, tran11-CLA. No dramatic change was observed among other 8 fatty acids in soymilk. The proportion of unsaturated fatty acids varied after fermentation with different L. plantrum strains, but all decreased the during storage. The research on the ability of converting CLA of L. plantrum strains could be basis for the future research and development of fermented soymilk products. PRACTICAL APPLICATION: Desirable probiotic traits, such as acid and bile tolerance, aggregation activity, and antibacterial activity, have been proved for the 6 Lactobacillus plantarum strains. The 6 L. plantarum strains might be used in the fermentation of soymilk to produce multifunctional probiotic soymilk products, especially the rich CLA contents.     

生物转化共轭亚油酸乳杆菌的筛选

以选定的乳杆菌为对象,系统性地探究其生物转化共轭亚油酸(CLA)的规律,并筛选得到高产菌株。首先用酶标仪快速检测法,对5个种属76株乳杆菌生物转化CLA的能力进行了分析,然后利用GC-MS进一步分析高产菌株生物转化CLA产物异构体组成。结果显示:植物乳杆菌大部分可生物转化CLA,但菌株间转化能力存在较大差异;副干酪乳杆菌、干酪乳杆菌和卷曲乳杆菌普遍具备生物转化CLA的能力,但转化能力不高;唾液乳杆菌普遍不具备生物转化CLA的能力。筛选得到3株高产菌株,其中植物乳杆菌CCFM241 CLA的转化能力最高,CLA转化率为16.35%,转化产物存在3种CLA异构体,以t9,t11-CLA为主。     

Conjugated linoleic acids in milk fat: Variation and physiological effects

Much attention has been directed toward conjugated linoleic acid (CLA) since the discovery of its anticarcinogenic properties two decades ago. Many other biological activities have been reported over the past few years confirming that individual CLA isomers present in milk fat have a high health promoting potential. Its possible use in functional dairy products explains the increasing interest of the food industry in CLA research. Recent advances in the analytical methodology offer new possibilities to study the individual effects of the various isomers in biological systems. The aim of this review is to summarize the current knowledge in CLA research including the formation of CLA in cows, analysis of CLA isomers, factors influencing the CLA content in milk, processing of CLA-enriched milk and dairy products, as well as aspects concerning nutrition and health.     

Advances in Research on cis-9, trans-11 Conjugated Linoleic Acid: A Major Functional Conjugated Linoleic Acid Isomer

Conjugated linoleic acid (CLA) consists of a group of positional and geometric conjugated isomers of linoleic acid. Since the identification of CLA as a factor that can inhibit mutagenesis and carcinogenesis, thousands of studies have been conducted in the last several decades. Among the many isomers discovered, cis-9, trans-11 CLA is the most intensively studied because of its multiple, isomer-specific effects in humans and animals. This paper provides an overview of the available data on cis-9, trans-11 CLA, including its isomer-specific effects, biosynthesis, in vivo/in vitro research models, quantification, and the factors influencing its content in ruminant products.     

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.     

Effects of conjugated linoleic acid isomers on lipid metabolism and gluconeogenesis in monolayer cultures of bovine hepatocytes

The objective was to determine the effects of linoleic acid and different isomers of conjugated linoleic acid (CLA) at different concentrations on hepatic lipid and glucose metabolism in the bovine. Monolayer cultures of hepatocytes obtained from 7- to 10-d-old Holstein bull calves were exposed to treatments from 16 to 64 h after plating. The treatments included 1.0 mM palmitic acid plus either 0.1 or 1.0 mM of cis-9, cis-12 linoleic acid, cis-9, trans-11 CLA, or trans-10, cis-12 CLA. Metabolism of palmitic acid to cellular triacylglycerol (TAG) was decreased when media contained cis-9, trans-11 compared with trans-10, cis-12 CLA. Total cellular TAG content was increased for the CLA isomers compared to cis-9, cis-12 linoleic acid. Both CLA isomers increased palmitic acid incorporation into phospholipids, cholesterol, and media triacylglycerol compared with cis-9, cis-12 linoleic acid at a concentration of 1.0 mM. Increasing the concentration of treatment fatty acids from 0.1 to 1.0 mM decreased oxidation of palmitic acid to acid-soluble products, but no effects of fatty acids were observed. There was no treatment effect on rates of gluconeogenesis from propionic acid. Overall, CLA isomers elicited changes in palmitic acid metabolism to cellular and media triacylglycerol, and cellular phospholipids and cholesterol, but had little or no effect on other measured pathways of lipid metabolism or gluconeogenesis in bovine hepatocytes.     

Factors affecting conjugated linoleic acid content in milk and meat

Conjugated linoleic acid (CLA) has been recently studied mainly because of its potential in protecting against cancer, atherogenesis, and diabetes. Conjugated linoleic acid (CLA) is a collective term for a series of conjugated dienoic positional and geometrical isomers of linoleic acid, which are found in relative abundance in milk and tissue fat of ruminants compared with other foods. The cis-9, trans-11 isomer is the principle dietary form of CLA found in ruminant products and is produced by partial ruminal biohydrogenation of linoleic acid or by endogenous synthesis in the tissues themselves. The CLA content in milk and meat is affected by several factors, such as animal's breed, age, diet, and management factors related to feed supplements affecting the diet. Conjugated linoleic acid in milk or meat has been shown to be a stable compound under normal cooking and storage conditions. Total CLA content in milk or dairy products ranges from 0.34 to 1.07% of total fat. Total CLA content in raw or processed beef ranges from 0.12 to 0.68% of total fat. It is currently estimated that the average adult consumes only one third to one half of the amount of CLA that has been shown to reduce cancer in animal studies. For this reason, increasing the CLA contents of milk and meat has the potential to raise the nutritive and therapeutic values of dairy products and meat.     

新型食品营养强化剂-共轭亚油酸

共轭亚油酸 (Conjgatedlinoleicacid ,CLA)是一类含有共轭双键的亚油酸的异构体 ,由于具有特殊的脂肪链结构使得其具有多种生理功能。c9t11和t10cl2异构体是共轭亚油酸的主要活性异构体。本文对共轭亚油酸的生理功能、代谢、安全性等进行综述。     

Effect of ripening on total conjugated linoleic acid and its isomers in buffalo Cheddar cheese

Cheese is a biologically and biochemically dynamic product in which a series of sequential changes take place throughout its manufacture and subsequent ripening. These changes improve nutritional value, body and texture and enrich the flavour. Conjugated linoleic acids (CLA), a group of naturally occurring geometrical and positional isomers of linoleic acid, are present in high concentration in dairy products and fat from ruminant animals. Six isomers of CLA have been separated and identified by gas liquid chromatography (GLC) analysis. Buffalo milk had higher 9c,11t-18:2 isomer (86.14%). Total CLA concentration of CLA isomers was increased significantly ( P <  0.05) on conversion of milk into cheese and subsequent storage. The concentration of the 9c,11t-18:2 isomer was decreased significantly ( P <  0.05), whereas that of trans , trans -isomers (12,14-; 11,13-; 10,12-; 9,11-) increased significantly ( P <  0.05).     

Linoleic acid Isomerase Activity in Enzyme Extracts from Lactobacillus Acidophilus and Propionibacterium Freudenreichii ssp. Shermanii

Enzyme extracts from Lactobacillus acidophilus (CCRC14079) and Propionibacterium freudenreichii ssp. shermanii (CCRC11076) were reacted with linoleic acid at 50 °C for 10 min at pH 5, 6, 7, and 8, and the levels of conjugated linoleic acid (CLA) formation were determined by high performance liquid chromatography. CLA formations were observed in all the reactions catalyzed by the retentates using ultrafiltration membranes with 100 kDa nominal molecular weight cutoff, indicating the presence of the activity of linoleic acid isomerase with nominal molecular weight higher than 100 kDa in the retentates from two cultures. More CLA was formed at pH 5 of L. acidophilus treatment and t10c12, c11t13, and c9t11 CLA were 3 major CLA isomers produced. Results demonstrate a potential for CLA production through linoleic acid isomerase.     

Quantitative and qualitative determination of CLA produced by Bifidobacterium and lactic acid bacteria by combining spectrophotometric and Ag-HPLC techniques

Bifidobacterium and lactic acid bacteria (LAB), especially from the genera Lactobacillus and Lactococcus, are commonly used in the production of fermented dairy products due to their potential probiotic characteristics. Moreover, some strains of these microorganisms also have the ability to produce conjugated linoleic acid (CLA) from linoleic acid (LA), which has attracted much attention as a novel type of beneficial functional fermented milk. In the present work 22 probiotic bacteria were tested for the production of CLA, using a UV screening method and HPLC techniques. Five microorganisms, two strains of the genera Bifidobacterium, two Lactobacillus and one Lactococcus were selected for their ability to produce CLA after incubation in skim milk with free LA as a substrate. It was possible to quantify the production of CLA (in the range of 40–50 μg CLA/ml) and identify the CLA isomers produced as C18:2 cis 9, trans 11 (60–65%), C18:2 trans 10, cis 12 (30–32%), C18:2 trans 9, trans 11 and C18:2 trans 10, trans 12 (2–5%).     

影响乳制品中共轭亚油酸(CLA)含量的因素

概述了共轭亚油（CLA）的形成方式、影响乳制品中CLA含量的因素及如何提高乳中CLA含量的方法。动物的种类、饲料的成分、喂养方式、季节等对原料乳产生的影响．以及加工方法、贮存条件、发酵菌种等对乳制品中CLA的含量的影响。     

气相色谱-质谱联用法分析苏尼特羔羊肉中共轭亚油酸含量

共轭亚油酸是亚油酸的一组几何和位置异构体，具有多种生理功能。作为瘤胃微生物的产物，反刍动物肉制品是共轭亚油酸异构体较丰富的天然来源，本研究利用气相色谱-质谱联用法分离测定了5月龄苏尼特羔羊肌肉和腹部脂肪中共轭亚油酸总含量，研究结果表明，背最长肌间脂肪中CLA相对百分含量平均为0．83％，腹部脂肪中CLA相对百分含量平均为0．99％。     

Optimisation of fermentation variables for conjugated linoleic acid bioconversion by Lactobacillus fermentum DDHI27 in modified skim milk

Effects of fermentation variables on conjugated linoleic acids (CLAs) biosynthesis of Lactobacillus fermentum DDHI27 in modified skim milk were evaluated. A total of 0.5 mg/mL was found to be the optimum linoleic acid concentration for maximum CLA production and this level was used in utilised for all successive experiments. The highest production of CLA isomers was detected at pH 6.0 when compared to other pH levels. Maximum biosynthesis of CLA isomers occurred during stationary growth (48 h) phase at all pH values tested. Validation of CLA isomers produced in selected samples was made by GC‐MS. These results indicate that these parameters strongly influence the CLA production capabilities of the strain.     

乳酸菌合成共轭亚油酸及其应用研究进展

共轭亚油酸(conjugated linoleic acid,CLA)是一种具有多种生理活性的功能性脂肪酸,在食品、保健品中具有广阔的应用前景。然而,天然来源的CLA含量很低,不能满足人类需求。乳酸菌可以合成CLA,且广泛用于食品加工领域,利用乳酸菌开发富含CLA的功能食品是当今的研究热点。为推动CLA功能食品的研究开发,该文对合成共轭亚油酸乳酸菌的研究进展、合成共轭亚油酸乳酸菌的益生性研究以及将产CLA乳酸菌应用到食品和饲料中的研究现状进行综述,并对利用乳酸菌合成CLA进而应用于功能食品进行展望。