LysR Family Regulator LttR Controls Production of Conjugated Linoleic Acid in Lactobacillus plantarum by Directly Activating the cla Operon.

Conjugated linoleic acids (CLAs) have attracted more attention as functional lipids due to their potential physiological activities, including anticancer, anti-inflammatory, anti-cardiovascular disease, and antidiabetes activities. Microbiological synthesis of CLA has become a compelling method due to its high isomer selectivity and convenient separation and purification processes. In Lactobacillus plantarum, the generation of CLA from linoleic acids (LAs) requires the combination of CLA oleate hydratase (CLA-HY), CLA short-chain dehydrogenase (CLA-DH), and CLA acetoacetate decarboxylase (CLA-DC), which are separately encoded by cla-hy, cla-dh, and cla-dc. However, the regulatory mechanisms of CLA synthesis remain unknown. In this study, we found that a LysR family transcriptional regulator, LTTR, directly bound to the promoter region of the cla operon and activated the transcription of cla-dh and cla-dc. The binding motif was also predicted by bioinformatics analysis and verified by electrophoretic mobility shift assays (EMSAs) and DNase I footprinting assays. The lttR overexpression strain showed a 5-fold increase in CLA production. Moreover, we uncovered that the transcription of lttR is activated by LA. These results indicate that LttR senses LA and promotes CLA production by activating the transcription of cla-dh and cla-dc. This study reveals a new regulatory mechanism in CLA biotransformation and provides a new potential metabolic engineering strategy to increase the yield of CLA.IMPORTANCE Our work has identified a novel transcriptional regulator, LTTR, that regulates the production of CLA by activating the transcription of cla-dh and cla-dc, essential genes participating in CLA synthesis in Lactobacillus plantarum This study provides insight into the regulatory mechanism of CLA synthesis and broadens our understanding of the synthesis and regulatory mechanisms of the biosynthesis of CLA.

Effect of Pufa Substrates on Fatty Acid Profile of Bifidobacterium breve Ncimb 702258 and CLA/CLNA Production in Commercial Semi-Skimmed Milk.

Current research on lipids is highlighting their relevant role in metabolic/signaling pathways. Conjugated fatty acids (CFA), namely isomers of linoleic and linolenic acid (i.e. CLA and CLNA, respectively) can positively modulate inflammation processes and energy metabolism, promoting anti-carcinogenic and antioxidant effects, improved lipid profiles and insulin resistance, among others. Bioactive doses have been indicated to be above 1 g/d, yet these cannot be achieved through a moderate intake (i.e. 1-2 servings) of natural sources, and certain CLA-containing products have limited commercial availability. Such handicaps have fueled research interest in finding alternative fortification strategies. In recent years, screening of dairy products for CFA-producing bacteria has attracted much attention and has led to the identification of some promising strains, including Bifidobacterium breve NCIMB 702258. This strain has shown interesting producing capabilities in model systems as well as positive modulation of lipid metabolism activities in animal studies. Accordingly, the aim of this research work was to assay B. breve NCIMB 702258 in semi-skimmed milk to produce a probiotic fermented dairy product enriched in bioactive CLA and CLNA. The effect of substrates (LA, α-LNA and γ-LNA) on growth performance and membrane fatty acids profile was also studied, as these potential modifications have been associated to stress response. When tested in cys-MRS culture medium, LA, α-LNA and γ-LNA impaired the fatty acid synthesis by B. breve since membrane concentrations for stearic and oleic acids decreased. Variations in the C18:1 c11 and lactobacillic acid concentrations, may suggest that these substrates are also affecting the membrane fluidity. Bifidobacterium breve CFA production capacity was first assessed in cys-MRS with LA, α-LNA, γ-LNA or all substrates together at 0.5 mg/mL each. This strain did not produce CFA from γ-LNA, but converted 31.12% of LA and 68.20% of α-LNA into CLA and CLNA, respectively, after incubation for 24 h at 37 °C. In a second phase, B. breve was inoculated in a commercial semi-skimmed milk with LA, α-LNA or both at 0.5 mg/mL each. Bifidobacterium breve revealed a limited capacity to synthesize CLA isomers, but was able to produce 0.062-0.115 mg/mL CLNA after 24 h at 37 °C. However, organoleptic problems were reported which need to be addressed in future studies. These results show that although CFA were produced at too low concentrations to be able to achieve solely the bioactive dose in one daily portion size, fermented dairy products are a suitable vector to deliver B. breve NCIMB 702258.

Microbial Production of Conjugated Linoleic Acid and Conjugated Linolenic Acid Relies on a Multienzymatic System.

Conjugated linoleic acids (CLAs) and conjugated linolenic acids (CLNAs) have gained significant attention due to their anticarcinogenic and lipid/energy metabolism-modulatory effects. However, their concentration in foodstuffs is insufficient for any therapeutic application to be implemented. From a biotechnological standpoint, microbial production of these conjugated fatty acids (CFAs) has been explored as an alternative, and strains of the genera Propionibacterium, Lactobacillus, and Bifidobacterium have shown promising producing capacities. Current screening research works are generally based on direct analytical determination of production capacity (e.g., trial and error), representing an important bottleneck in these studies. This review aims to summarize the available information regarding identified genes and proteins involved in CLA/CLNA production by these groups of bacteria and, consequently, the possible enzymatic reactions behind such metabolic processes. Linoleate isomerase (LAI) was the first enzyme to be described to be involved in the microbiological transformation of linoleic acids (LAs) and linolenic acids (LNAs) into CFA isomers. Thus, the availability of lai gene sequences has allowed the development of genetic screening tools. Nevertheless, several studies have reported that LAIs have significant homology with myosin-cross-reactive antigen (MCRA) proteins, which are involved in the synthesis of hydroxy fatty acids, as shown by hydratase activity. Furthermore, it has been suggested that CLA and/or CLNA production results from a stress response performed by the activation of more than one gene in a multiple-step reaction. Studies on CFA biochemical pathways are essential to understand and characterize the metabolic mechanism behind this process, unraveling all the gene products that may be involved. As some of these bacteria have shown modulation of lipid metabolism in vivo, further research to be focused on this topic may help us to understand the role of the gut microbiota in human health.

Bacterial conjugated linoleic acid production and their applications.

Conjugated linoleic acid (CLA) has been shown to exert various potential physiological properties including anti-carcinogenic, anti-obesity, anti-cardiovascular and anti-diabetic activities, and consequently has been considered as a promising food supplement. Bacterial biosynthesis of CLA is an attractive approach for commercial production due to its high isomer-selectivity and convenient purification process. Many bacterial species have been reported to convert free linoleic acid (LA) to CLA, hitherto only the precise CLA-producing mechanisms in Propionibacterium acnes and Lactobacillus plantarum have been illustrated completely, prompting the development of recombinant technology used in CLA production. The purpose of the article is to review the bacterial CLA producers as well as the recent progress on describing the mechanism of microbial CLA-production. Furthermore, the advances and potential in the heterologous expression of CLA genetic determinants will be presented.

One-pot conjugated linoleic acid production from castor oil by Rhizopus oryzae lipase and resting cells of Lactobacillus plantarum.

Conjugated linoleic acid (CLA) has attracted as novel type of fatty acids having unusual health-promoting properties such as anticarcinogenic and antiobesitic effects. The present work employed castor oil as substrate for one-pot production of CLA using washed cells of Lactobacillus plantarum (L. plantarum) and lipases as catalysts. Among the screened lipases, the lipase Rhizopus oryzae (ROL) greatly assisted resting cells to produce CLA. Mass spectral analysis of the product showed that two major isomers of CLA were produced in the reaction mixture i.e. cis-9, trans-11 56.55% and trans-10, cis-12 43.45%. Optimum factors for CLA synthesis were found as substrate concentration (8 mg/mL), pH (6.5), washed cell concentration (12% w/v), and incubation time of 20 h. Hence, the combination of ROL with L. plantarum offers one pot production of CLA selectively using castor oil as a cost-effective substrate.

Synthesis of the suspected trans-11,cis-13 conjugated linoleic acid isomer in ruminant mammary tissue by FADS3-catalyzed Δ13-desaturation of vaccenic acid.

The octadecadienoic conjugated linoleic acid (CLA) isomer with trans-11 and cis-13 double bonds (trans-11,cis-13 CLA) has been described in ruminant milk. For now, this specific CLA is suspected to derive exclusively from ruminal biohydrogenation of dietary α-linolenic acid. However, in rodents, the fatty acid desaturase 3 (FADS3) gene was recently shown to code for an enzyme able to catalyze the unexpected Δ13-desaturation of vaccenic acid, producing a Δ11,13-CLA with all the structural characteristics of the trans-11,cis-13 isomer, although no commercial standard exists for complete conclusive identification. Because the FADS3 gene has already been reported in bovine animals, we hypothesized in the present study that an alternative direct FADS3-catalyzed Δ13-desaturation of vaccenic acid in mammary tissue may therefore co-exist with α-linolenic acid biohydrogenation to explain the final ruminant milk trans-11,cis-13 CLA presence. Here, we first confirm that the FADS3 gene is present in ruminant mammal genomic sequence databases. Second, we demonstrate that the Δ11,13-CLA found in milk fat and the highly probable trans-11,cis-13 CLA isomer produced by rodent FADS3 possess exactly the same structural characteristics. Then, we show that bovine mammary MAC-T and BME-UV epithelial cells express both FADS3 and stearoyl-CoA desaturase 1 (SCD1) mRNA and are able to synthesize both the suspected trans-11,cis-13 CLA and cis-9,trans-11CLA (rumenic acid) isomers when incubated with vaccenic acid. Finally, the concomitant presence of the suspected trans-11,cis-13 CLA isomer with FADS3 mRNA was shown in goat mammary tissue, whereas both were conversely very low or even absent in goat liver. Therefore, this study provides several lines of evidence that, by analogy with rumenic acid, trans-11,cis-13 CLA may originate both from ruminal biohydrogenation and from direct FADS3-catalyzed Δ13-desaturation of vaccenic acid in mammary tissue.

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.

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.

An overview of conjugated linoleic acid: microbial production and application.

Conjugated linoleic acid (CLA) has attracted considerable attention in health due to its important physiological properties proved in several in vivo experiments. Many bacteria, especially some probiotics, are able to produce CLA from the linoleic acid (LA) present in milk. In this review, CLA production by microorganisms is described. Then factors on the influencing the microbial production and the initial CLA content in milk fat are introduced. After a glimpse on the content of CLA in dairy products and human body, health benefits of CLA including anti-cancer, anti-diabetic, antiathrosclerosis and anti-osteoporosis properties, as well as prevention of body fat increase and function as stimulator of the immunity system are explained.

Genetic engineering of Yarrowia lipolytica for enhanced production of trans-10, cis-12 conjugated linoleic acid.

BACKGROUND: Conjugated linoleic acid (CLA) has been extensively studied for decades because of its health benefits including cancer prevention, anti-atherogenic and anti-obesity effects, and modulation of the immune system. We previously described the production of trans-10, cis-12 CLA in Yarrowia lipolytica by expressing the gene coding for linoleic acid isomerase from Propionibacterium acnes (pai). However the stable strain produced CLA at about 0.08% of dry cell weight (DCW), a level of production which was not high enough for practical applications. The goal of the present study was to enhance production of CLA by genetic engineering of Y. lipolytica strains. RESULTS: We have now co-expressed the delta 12-desaturase gene (FADS12, d12) from Mortierella alpina together with the codon-optimized linoleic acid isomerase (opai) gene in Y. lipolytica, expressed under the control of promoter hp16d modified by fusing 12 copies of UAS1B to the original promoter hp4d. A multi-copy integration plasmid was used to further enhance the expression of both genes. Using glucose as the sole carbon source, the genetically-modified Y. lipolytica produced trans-10, cis-12-CLA at a level of up to 10% of total fatty acids and 0.4% of DCW. Furthermore, when the recombinant yeast was grown with soybean oil, trans-10, cis-12-CLA now accumulated at a level of up to 44% of total fatty acids, which represented 30% of DCW after 38.5 h of cultivation. In addition, trans-10, cis-12-CLA was also detected in the growth medium up to 0.9 g/l. CONCLUSIONS: We have successfully produced trans-10, cis-12-CLA with a titre of 4 g/l of culture (3.1 g/l in cells and 0.9 g/l in culture medium). Our results demonstrate the potential use of Y. lipolytica as a promising microbial cell factory for trans-10, cis-12-CLA production.

Appraisal of conjugated linoleic acid production by probiotic potential of Pediococcus spp. GS4.

Probiotics with ability to produce conjugated linoleic acid (CLA) is considered as an additive health benefit property for its known role in colon cancer mitigation. The conversion involves the biohydrogenation of the unsaturated fatty acid into conjugated form. Probiotic strain Pediococcus spp. GS4 was efficiently able to biohydrogenate linoleic acid (LA) into its conjugated form within 48 h of incubation. Quantum of CLA produced with a concentration of 121 µg/ml and sustained cell viability of 8.94 log cfu/ml maximally. Moreover, antibacterial effect of LA on the strain ability for biohydrogenation was examined at different concentrations and concluded to have a direct relationship between LA and amount of CLA produced. The efficiency of the strain for CLA production at different pH was also estimated and found maximum at pH 6.0 with 149 µg/ml while this ability was reduced at pH 9.0 to 63 µg/ml. Sesame oil, which is rich in the triacylglycerol form of LA, was also found to act as a substrate for CLA production by Pediococcus spp. GS4 with the aid of lipase-catalyzed triacylglycerol hydrolysis and amount of CLA produced was 31 µg/ml at 0.2 % while 150 µg/ml at 1.0 % of lipolysed oil in skim milk medium. Conjugated form was analyzed using UV scanning, RP-HPLC, and GC-MS. This study also focused on the alternative use of lipolysed sesame oil instead of costly LA for biohydrogenation and could be a potential source for the industrial production of CLA.

Production of a conjugated fatty acid by Bifidobacterium breve LMC520 from α-linolenic acid: conjugated linolenic acid (CLnA).

This study was performed to characterize natural CLnA isomer production by Bifidobacterium breve LMC520 of human origin in comparison to conjugated linoleic acid (CLA) production. B. breve LMC520 was found to be highly active in terms of CLnA production, of which the major portion was identified as cis-9,trans-11,cis-15 CLnA isomer by GC-MS and NMR analysis. B. breve LMC520 was incubated for 48 h using MRS medium (containing 0.05% L-cysteine · HCl) under different environmental conditions such as atmosphere, pH, and substrate concentration. The high conversion rate of α-linolenic acid (α-LNA) to CLnA (99%) was retained up to 2 mM α-LNA, and the production was proportionally increased nearly 7-fold with 8 mM by the 6 h of incubation under anaerobic conditions at a wide range of pH values (between 5 and 9). When α-LNA was compared with linoleic acid (LA) as a substrate for isomerization by B. breve LMC520, the conversion of α-LNA was higher than that of LA. These results demonstrated that specific CLnA isomer could be produced through active bacterial conversion at an optimized condition. Because many conjugated octadecatrienoic acids in nature are shown to play many positive roles, the noble isomer found in this study has potential as a functional source.

Conversion of t11t13 CLA into c9t11 CLA in Caco-2 cells and inhibition by sterculic oil.

BACKGROUND: Conjugated linoleic acids (CLA), and principally c9t11 CLA, are suspected to have numerous preventive properties regarding non-infectious pathologies such as inflammatory diseases, atherosclerosis and several types of cancer. C9t11 CLA is produced in the rumen during biohydrogenation of linoleic acid, but can also be synthesized in mammalian tissues from trans-vaccenic acid (C18:1 t11) through the action of delta-9 desaturase (D9D). For several years, it is also known that c9t11 CLA can be synthesized from conjugated linolenic acids (CLnA), i.e. c9t11c13 CLnA and c9t11t13 CLnA. This study aimed at investigating to which extent and by which route c9t11 CLA can be produced from another isomer of CLA, the t11t13 CLA that is structurally very similar to c9t11t13 CLnA, in Caco-2 cells. METHODOLOGY/PRINCIPAL FINDINGS: Caco-2 cells were incubated for 24 h with 20 µmol/l of t11t13 CLA in the absence or presence of sterculic oil used as an inhibitor of D9D. Caco-2 cells were able to convert t11t13 CLA into c9t11 CLA, and c9t11t13 CLnA was formed as an intermediate compound. In the presence of sterculic oil, the production of this intermediate was decreased by 46% and the formation of c9t11 CLA was decreased by 26%. No other metabolite was detected. CONCLUSIONS/SIGNIFICANCE: These results not only highlight the conversion of t11t13 CLA into c9t11 CLA but demonstrate also that this conversion involves first a desaturation step catalysed by D9D to produce c9t11t13 CLnA and then the action of another enzyme reducing the double bond on the Δ13 position.

Rumen microbial response in production of CLA and methane to safflower oil in association with fish oil or/and fumarate.

Supplementation effect of fish oil and/or fumarate on production of conjugated linoleic acid (CLA) and methane by rumen microbes was examined when incubated with safflower oil. One hundred and twenty milligrams of safflower oil (SO), safflower oil with 24 mg fish oil (SOFO), safflower oil with 24 mmol/L fumarate (SOFA), or safflower oil with 24 mg fish oil and 24 mmol/L fumarate (SOFOFA) were added to the 90 mL culture solution. The culture solution was also made without any supplements (control). The SOFA and SOFOFA increased pH and propionate (C3) compared to other treatments from 3 h incubation time. An accumulated amount of total methane (CH(4) ) for 12 h incubation was decreased by all the supplements compared to control. The concentrations of c9,t11CLA for all the incubation times were increased in the treatments of SOFO, SOFA and SOFOFA compared to SO. The highest concentration of c9,t11CLA was observed from SOFOFA among all the treatments at all incubation times. Overall data indicate that supplementation of combined fumarate and/or fish oil when incubated with safflower oil could depress CH(4) generation and increase production of C(3) and CLA under the condition of current in vitro study.

Genetic variation at the goat hormone-sensitive lipase ( LIPE) gene and its association with milk yield and composition.

Hormone-sensitive lipase (LIPE) plays a fundamental role in the regulation of energy balance by releasing free fatty acids from adipose triacylglycerol stores. These fatty acids can be subsequently transferred to other body compartments to be oxidized or employed in other biochemical reactions. This enzymic function is particularly important in lactating animals because the synthesis of milk components involves the mobilization of lipid depots to satisfy the large energy demands of the mammary gland. In the current study, we partially sequenced the goat LIPE gene in several individuals. In doing so, we identified two synonymous polymorphisms at exons 2 (c.327C>A>T, triallelic polymorphism) and 3 (c.558C>T). Moreover, we found a mis-sense polymorphism at exon 6 (c.1162G>T) that involves an alanine to serine substitution at position 388. Analysis with Polyphen and Panther softwares revealed that this amino acid replacement is expected to be neutral. Performance of an association analysis with a variety of milk traits revealed that goat LIPE genotype has highly suggestive effects on milk yield (P=0.0032) as well as on C18:3 n-6g (P=0.0051), trans-10 cis-12 CLA (P=0.007) and C12:0 (P=0.0084) milk contents. These associations are concordant with the preference of LIPE to selectively mobilize medium-chain and unsaturated fatty acids.

Effects of fish oil and starch added to a diet containing sunflower-seed oil on dairy goat performance, milk fatty acid composition and in vivo delta9-desaturation of [13C]vaccenic acid.

The potential benefits on human health have prompted an interest in developing nutritional strategies for specifically increasing rumenic acid (RA) in ruminant milk. The aims of the present study were to (i) compare two dietary treatments with lipid supplements on milk yield and composition, (ii) measure the in vivo delta9-desaturation of vaccenic acid (VA) to RA using 13C-labelled VA and (iii) determine the effect of the dietary treatments on this variable. Treatments were 90 g sunflower-seed oil (SO) per d or 60 g sunflower-seed oil and 30 g fish oil per d plus additional starch (SFO), in a grassland hay-based diet given to eight Alpine goats in a 2 x 2 cross-over design with 21 d experimental periods. Milk yield and composition were similar between treatments. Goats fed SFO had higher milk 6 : 0-16 : 0 concentration, lower milk sigmaC18 concentrations and showed no effect on milk VA and RA, compared with SO. At the end of the experiment, intravenous injection of 1.5 g [13C]VA followed by measurements of milk lipid 13C enrichment showed that in vivo 31.7 and 31.6 % of VA was delta9-desaturated into milk RA in the caprine with the SO and SFO treatments, respectively. The expression of genes encoding for delta9-desaturase (or stearoyl-CoA desaturase; SCD1, SCD5) in mammary tissues and four milk delta9-desaturation ratios were similar between treatments. In conclusion, the present study provides the first estimates of in vivo endogenous synthesis of RA (63-73 % of milk RA) from VA in goats, and shows no difference between the two lipid supplements compared.

Proteomic analysis of endogenous conjugated linoleic acid biosynthesis in lactating rats and mouse mammary gland epithelia cells (HC11).

This study was conducted to investigate the amount of CLA synthesized endogenously by rat mammary tissues in response to TVA (a precursor for cis-9, trans-11 CLA endogenous synthesis) treatment as well as the differences in the protein expression of genes encoding the biosynthesis of CLA in rat mammary tissue and mouse mammary gland epithelia cells (HC11). Treatment with TVA resulted in improved CLA productivity. Furthermore, 2-DE revealed two spots in samples of mammary tissues and one spot in samples of mammary gland epithelia cells (HC11) that were consistently altered in the TVA treatment groups when compared with the control group (non-fatty acid). The mRNA expression patterns of three of the proteins (PDI, PRDX2, LAMR1), as measured by real-time PCR, were similar to the pattern of protein abundance. In addition, the expression of SCD mRNA in the mammary tissue of rats and HC11 cell treated with TVA was higher than in the control group. Our results suggest that the identified proteins may be related to CLA biosynthesis in mammary tissue.

Optimization of a reconstituted skim milk based medium for enhanced CLA production by bifidobacteria.

AIMS: To determine the effect of a range of supplements on the bioconversion of linoleic acid to conjugated linoleic acid (CLA) by Bifidobacterium breve NCIMB 702258 in reconstituted skim milk (RSM). RESULTS: Seven supplements (yeast extract, casein hydrolysate, tryptone, l-cysteine hydrochloride, sodium acetate, sodium butyrate and sodium propionate) were identified as increasing the bioconversion of linoleic acid to c9, t11 CLA. Using these supplements, the percentage bioconversion of linoleic acid (0.35 mg ml(-l)) to the c9, t11 CLA isomer was elevated from 15.5 +/- 1.1% in 20% RSM (w/v) to 48.1 +/- 2.2% in the supplemented RSM. Through additional supplementation of 20 mg m1(-1) inulin and optimization of inoculum and linoleic acid concentration, the percentage bioconversion to c9, t11 CLA was increased to 55.0 + 3.2%. CONCLUSIONS: Through supplementation, the concentration of CLA produced by bifidobacteria in RSM can be increased to levels comparable to those observed in the synthetic medium cys-MRS. SIGNIFICANCE AND IMPACT OF THE STUDY: The impact of 22 supplements on the production of the c9, t11 CLA isomer by the strain B. breve NCIMB 702258 in milk has been determined. The results provide an understanding of the factors, which influence CLA production by bifidobacteria in RSM.

Production of conjugated linoleic acid by probiotic Lactobacillus acidophilus La-5.

AIMS: To study the ability of the probiotic culture Lactobacillus acidophilus La-5 to produce conjugated linoleic acid (CLA), which is a potent anti-carcinogenic agent. METHODS AND RESULTS: The conversion of linoleic acid to CLA was studied both by fermentation in a synthetic medium and by incubation of washed cells. Accumulation of CLA was monitored by gas chromatography analysis of the biomass and supernatants. While the fermentation conditions applied may not be optimal to observe CLA production in growing La-5 cells, the total CLA surpassed 50% of the original content in the washed cells after 48 h under both aerobic and micro-aerobic conditions. The restriction of oxygen did not increase the yield, but favoured the formation of trans, trans isomers. CONCLUSIONS: The capability of L. acidophilus La-5 to produce CLA is not dependant on the presence of milk fat or anaerobic conditions. Regulation of CLA production in this strain needs to be further investigated to exploit the CLA potential in fermented foods. SIGNIFICANCE AND IMPACT OF THE STUDY: Knowledge gained through the conditions on the accumulation of CLA would provide further insight into the fermentation of probiotic dairy products. The capacity of the nongrowing cells to produce CLA is also of great relevance for the emerging nonfermented probiotic foods.

The effect of solids dilution rate and oil source on trans C18:1 and conjugated linoleic acid production by ruminal microbes in continuous culture.

The objective of this study was to evaluate the effect of solids dilution rate (SDR) and oil source [soybean oil (SBO) or linseed oil (LSO)] on the ruminal production of trans C18:1 and conjugated linoleic acid (CLA). A dual-flow continuous culture system consisting of 4 fermenters was used in a 4 x 4 Latin square design with a factorial arrangement of treatments over 4 consecutive periods of 10 d each. Treatment diets (50:50 forage to concentrate) were fed at 120 g/d of dry matter (DM) in 3 equal portions. The concentrate mix contained 1% fish oil and either 2% SBO or 2% LSO on a DM basis. Treatments were as follows: 1) SBO at 6%/h SDR, 2) SBO at 3%/h SDR, 3) LSO at 6%/h SDR, and 4) LSO at 3%/h SDR. The oil source by SDR interaction was not significant for trans C18:1 and CLA fatty acids. The concentrations of trans C18:1 and vaccenic acid were greater in effluents when diets were supplemented with SBO vs. LSO (37.11 vs. 34.09 and 32.71 vs. 29.70 mg/g of DM, respectively) and at high SDR than low SDR (37.60 vs. 33.61 and 32.72 vs. 29.61 mg/g of DM, respectively). The concentration of cis-9, trans-11 CLA in effluents was also greater with SBO than LSO (0.81 vs. 0.40 mg/g of DM) supplementation and at high SDR than low SDR (0.68 vs. 0.54 mg/g of DM). Biohydrogenation of linoleic acid and linolenic acid increased at higher SDR within each oil treatment. Based on these results, SBO supplementation at high SDR enhances ruminal production of vaccenic acid, and therefore could potentially enhance cis-9, trans-11 CLA in milk fat through synthesis by Delta9-desaturase.