Conjugated Linoleic Acid Production Research Articles

Effect of Minor Oil Constituents on Soy Oil Conjugated Linoleic Acid Production

Conjugated linoleic acid (CLA) is produced by photoisomerization of soy oil linoleic acid. Yields increase with the degree of oil refining, but the effect of specific minor oil components is not known. Therefore, the objectives were to determine the effect of each soy oil minor component on CLA yields and oxidative stability after processing, to determine the effect of soy oil minor constituent interactions on CLA yields and oxidative stability, and to determine how soy oil Magnesol adsorption pretreatment affects CLA yields. Soy oils with varying levels of peroxides, tocopherols, phospholipids, free fatty acids (FFA), and lutein were each UV irradiated, and the CLA content and oxidative stability were determined. A peroxide value of above 0.8 greatly decreased CLA yields, as did phospholipids above 500 ppm. Tocopherols enhanced CLA production at low levels and reduced yields at high concentrations, while lutein and FFA had little effect. High CLA yields corresponded with a lower oil oxidative stability. The interactions between the minor components showed similar trends as seen in the single component study. These findings were supported by observations that Magnesol adsorption removed large quantities of phospholipids and peroxides from soy oil and greatly increased CLA yields but reduced the oxidative stability. Minor components, particularly peroxides and phospholipids, need to be removed from the oil to optimize CLA yields.

Microbial production of conjugated fatty acids

AbstractConjugated 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 of production 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 microorganisms. Lactic acid bacteria produced CLA from linoleic acid. The CLA, which was obtained as the free fatty acid form, comprised a mixture of cis‐9,trans‐11‐octadecadienoic acid (18:2) and trans‐9,trans‐11‐18:2. Furthermore, lactic acid bacteria transformed ricinoleic acid [12‐hydroxy‐cis‐9‐octadecenoic acid (18:1)] to CLA (a mixture of cis‐9,trans‐11‐18:2 and trans‐9,trans‐11‐18:2). Castor oil, 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. Filamentous fungi transformed trans‐vaccenic acid (trans‐11‐18:1) to cis‐9,trans‐11‐18:2 by Δ9 desaturation. This CLA was obtained as a triacylglycerol. In addition, lactic acid bacteria produced conjugated trienoic fatty acids from α‐ and γ‐linolenic acid. The trienoic fatty acids produced from α‐linolenic acid were cis‐9,trans‐11,cis‐15‐octadecatrienoic acid (18:3) and trans‐9,trans‐11,cis‐15‐18:3. Those produced from γ‐linolenic acid were cis‐6,cis‐9,trans‐11‐18:3 and cis‐6,trans‐9,trans‐11‐18:3.

Influence of sunflower oil on conjugated linoleic acid production byLactobacillus acidophilus andLactobacillus casei

This study was conducted to test the conjugated linoleic acid (CLA) producing potential ofLactobacillus acidophilus NCDC-14 andLactobacillus casei NCDC-19, where sunflower oil (a rich source of linoleic acid) was used as a substrate for CLA production in skim milk at varying concentrations ranging from 0.25–1.0%.Lactobacillus casei produced higher amount of CLA i.e. 11.0 mg/g of fat at a concentration of 1.0% sunflower oil, whereasL. acidophilus produced only 2.73 mg/g of fat. However, at the lower concentration of sunflower oil (0.25%), there was no considerable difference (i.e. 6.92 and 6.73 mg/g of fat forL. casei andL. acidophilus, respectively) in the CLA producing ability of both the cultures, indicating that the increased amount of oil in the medium led to the inhibition of either the growth ofL. acidophilus or the CLA conversion. Hence,L. casei may extend a possibility to be used in the preparation of different fermented milk products for enhanced CLA content.

Characterization of Conjugated Linoleic Acid Production by Bifidobacterium breve LMC 520

This study was performed to characterize the CLA production ability of a bacterial strain, Bifidobacterium breve LMC 520, which can actively convert linoleic acid (LA) to cis-9,trans-11 conjugated linoleic acid (CLA), a major isomer derived from microbial enzymatic conversion. The culture conditions were optimized to improve CLA production under the aerobic conditions. B. breve LMC 520 was tested with different amounts of LA in varied culture conditions, such as air, additives, and pH. A maximal level of CLA production (up to 90% of substrate) was obtained after 24 h of incubation in culture medium containing 1 mM LA at pH 5.5 and under anaerobic conditions. There was no decline in the CLA level with prolonged incubation until 48 h. When the effect of pre-incubation with LA on CLA production was tested, there was no significant difference between the CLA-producing activity of pre-incubated and untreated bacteria at the third passage but there was a significant reduction in CLA production by the pre-incubated cells after the fourth passage. These results demonstrate that the CLA-producing activity of B. breve LMC 520 could be maximized by numerous environmental factors. The data also indicate its potential for increasing CLA accumulation in dairy products when B. breve LMC 520 is used as a functional starter culture.

Conjugated Linoleic Acid in Rumen Liquor: Their Relation with Other Carbon 18 Fatty Acids and Rumen pH

This study was focused on the quantitative analysis of Conjugated Linoleic Acids (CLAs) in rumen liquor and assessment of the relationship between CLAs and other carbon 18 fatty acids and rumen pH of cattle slaughtered for meat consumption. The fatty acids were extracted using modified Folch method and determined by gas chromatography. The mean (n = 22) level of CLAs in the rumen liquor was 14.97 ±7.46 mg/100 mL. Cis-9, trans-11(c9, t11) was the dominant isomer of CLA followed by trans-10,cis-12 (t10, c12). The level of CLAs was positively correlated with the concentration of octadecatrienoic (18:3), cis-9, cis-12 octadecadienoic (18:2), trans-11octadecenoic (18:1) and octadecenoic (18:0) acids. Ruminal pH was positively correlated with the amount of c9, t11 and total CLA and negatively correlated with t10, c12 CLA isomer. All these correlations and relationships indicated that CLAs production in the rumen is dependent on availability of carbon 18 fatty acids and pH of the rumen liquor.

Linolenic Acid in Association with Malate or Fumarate Increased CLA Production and Reduced Methane Generation by Rumen Microbes

An in vitro study was conducted to investigate the effect of propionate precursor (malate or fumarate) on fermentation characteristics, production of conjugated linoleic acid (CLA) and methane (CH₄) by rumen microbes when incubated with linoleic acid (C 18:2 ) as a CLA precursor. Total CH₄ production for 12h incubation, however, was greatly reduced (P＜0.0001) by all the supplements compared to that of control, and its production from M-LA or F-LA treatment was smaller than that from LA treatment. Supplementation of M-LA or F-LA also increased concentrations of cis9, trans11-CLA (P＜0.039 - P＜0.001) for all incubation times and trans10,cis 12-CLA at 1h (P＜0.013), 3h(P＜0.036) and 12h (P＜0.025) incubation times compared to LA supplementation.

Production of conjugated linoleic acid by probioticLactobacillus acidophilusLa-5

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. 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. 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. 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.

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

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). 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%. 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. 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.

Mechanism of conjugated linoleic acid and vaccenic acid formation in human faecal suspensions and pure cultures of intestinal bacteria

Faecal bacteria from four human donors and six species of human intestinal bacteria known to metabolize linoleic acid (LA) were incubated with LA in deuterium oxide-enriched medium to investigate the mechanisms of conjugated linoleic acid (CLA) and vaccenic acid (VA) formation. The main CLA products in faecal suspensions, rumenic acid (cis-9,trans-11-CLA; RA) and trans-9,trans-11-CLA, were labelled at C-13, as were other 9,11 geometric isomers. Traces of trans-10,cis-12-CLA formed were labelled to a much lower extent. In pure culture, Bifidobacterium breve NCFB 2258 formed labelled RA and trans-9,trans-11-CLA, while Butyrivibrio fibrisolvens 16.4, Roseburia hominis A2-183T, Roseburia inulinivorans A2-192T and Ruminococcus obeum-like strain A2-162 converted LA to VA, labelled in a manner indicating that VA was formed via C-13-labelled RA. Propionibacterium freudenreichii subsp. shermanii DSM 4902T, a possible probiotic, formed mainly RA with smaller amounts of trans-10,cis-12-CLA and trans-9,trans-11-CLA, labelled the same as in the mixed microbiota. Ricinoleic acid (12-OH-cis-9-18 : 1) did not form CLA in the mixed microbiota, in contrast to CLA formation described for Lactobacillus plantarum. These results were similar to those reported for the mixed microbiota of the rumen. Thus, although the bacterial genera and species responsible for biohydrogenation in the rumen and the human intestine differ, and a second route of RA formation via a 10-OH-18 : 1 is present in the intestine, the overall labelling patterns of different CLA isomers formation are common to both gut ecosystems. A hydrogen-abstraction enzymic mechanism is proposed that may explain the role of a 10-OH-18 : 1 intermediate in 9,11-CLA formation in pure and mixed cultures.

Kinetics of microbial hydrogenation of free linoleic acid to conjugated linoleic acids

To investigate the ability of selected probiotic bacterial strains to produce conjugated linoleic acid (CLA) and also to estimate the biohydrogenation kinetics of Lactobacillus acidophilus on the production of CLA from free linoleic acid (LA). Six probiotic bacteria, Lact. paracasei, Lact. rhamnosus GG, Lact. acidophilus ADH, and Bifidobacterium longum B6, Lact. brevis, and Lact. casei, were used to examine their ability to convert LA to CLA. LA tolerance was evaluated by addition of different LA concentrations in MRS broth. Lact. acidophilus showed the major tolerant to LA and the greatest CLA-producing ability (36-48 microg ml(-1) of CLA). The rate-controlling steps were k(2) and k(1) for the addition of 1 and 3 mg ml(-1) of LA, respectively. The percentage of CLA conversion was higher in MRS broth supplemented with 1 mg ml(-1) (65%) than 3 mg ml(-1) (26%). The results provide useful information and new approach for understanding the biohydrogenation mechanisms of CLA production. This study would help elucidate the pathway from LA to stearic acid (SA), known as biohydrogenation. In addition, the use of selected probiotic bacteria might lead to a significant improvement in food safety.

Utilization of Monolinolein as a Substrate for Conjugated Linoleic Acid Production by Bifidobacterium breve LMC 520 of Human Neonatal Origin

This study was designed to isolate bifidobacteria from human intestines that efficiently converts monolinolein, a monoglyceride form of linoleic acid, into conjugated linoleic acid (CLA), as well as to optimize culture conditions for improving CLA production during milk fermentation. Among 150 screened neonatal bifidobacteria, Bifidobacterium breve LMC 520 showed the highest CLA-producing ability and was tested with different types of fat substrates at various concentrations to determine the optimal conditions for CLA production. Monolinolein was tested as a substrate for CLA production. The incubation time optimized for CLA production was 24 h, and CLA production was proportionally increased with monolinolein concentration. The incubation of LMC 520 with commercial starter strains caused minimal reduction in CLA production. Our results demonstrate that the CLA-producing ability of B. breve LMC 520 could offer beneficial effects when utilized as a starter culture for the development of functional dairy products.

Conjugated linoleic acid producing potential of lactobacilli isolated from the rumen of cattle

Lactobacilli isolated from the rumen of cattle were subjected to morphological and biochemical characterizations followed by PCR-based identification. Among isolates, Lactobacillus brevis was found to be the most prevalent species in the rumen. For in vitro conjugated linoleic acid (CLA) production, the two isolates of L. brevis and one each of Lactobacillus viridescens and Lactobacillus lactis were selected. The sunflower oil (i.e., 0.25, 0.5 and 1.0%; a rich source of linoleic acid) was added to skim milk as a substrate for CLA production by isolates at 37 degrees C/12 h. L. brevis 02 was found to be the most potential CLA producer (10.53 mg CLA/g fat) at 0.25% concentration of sunflower oil followed by L. brevis 01 (8.27 mg CLA/g fat). However, at higher level of sunflower oil (i.e., 1.0%), L. lactis was the highest CLA producer (9.22 mg/g fat) when compared to L. brevis and L. viridescens. The results indicated that L. brevis and/or CLA production was inhibited with increasing concentration of sunflower oil in skim milk. In contrast, L. lactis and L. viridescens could tolerate the increasing concentrations of sunflower oil and produced higher CLA. Overall, L. brevis extends a possibility to be used as a direct-fed microbial for ruminants to increase the CLA content in milk, however, in vivo trials are needed for validation of results obtained.

Bioconversion of linoleic acid to conjugated linoleic acid by Lactobacillus reuteri under different growth conditions

AbstractBACKGROUND: Lactobacillus reuteri was grown in De Man/Rogosa/Sharpe (MRS) broth (initial pH 6.5) supplemented with free linoleic acid (LA) at different concentrations (5, 10, 20 and 30 mg mL−1) and incubated aerobically at different temperatures (4, 10, 16, 22 and 30 °C) in order to test its ability to accomplish the bioconversion of LA to conjugated linoleic acid (CLA). Temperatures and LA concentrations producing the highest conversion of LA to CLA in the initial trials were tested further using micro‐anaerobic conditions and a lower initial pH (5.5).RESULTS: Data showed that production of CLA exhibited variations with regard to the fermentation conditions used. The highest production of CLA (0.108 mg mL−1) was measured in a broth containing 20 mg mL−1 free LA that was incubated aerobically at 10 °C for 30 h. When the initial pH of the reaction medium was reduced from 6.5 to 5.5, CLA production decreased. Micro‐aerobic conditions reduced the ability of Lb. reuteri to produce CLA, since production of CLA under aerobic conditions was at least 1.4 times greater.CONCLUSION: Production of CLA by Lb. reuteri at low temperatures and relatively high substrate concentrations provides novel opportunities for the development of functional foods with the benefits of enrichment in CLA and probiotic bacteria. Copyright © 2008 Society of Chemical Industry

Synthesis of Conjugated Linoleic Acid by Human-Derived Bifidobacterium breve LMC 017: Utilization as a Functional Starter Culture for Milk Fermentation

This study was performed to discover bifidobacteria isolated from human intestines that optimally convert linoleic acid (LA) to conjugated linoleic acid (CLA) and to optimize the culture conditions of milk fermentation. One hundred and fifty neonatal bifidobacteria were screened for CLA-producing ability, and Bifidobacterium breve LMC 017 was selected as it showed about 90% conversion of free LA in MRS broth. The selected strain showed resistance at 0.5% LA in microaerophillic conditions. When monolinolein (LA 90%) was used as a substrate for CLA production, the conversion rate was lower compared to free LA, but the growth rate was unaffected during the milk fermentation. There was no significant difference in CLA production between aerobic and anaerobic conditions, and little decline in CLA was shown after the maximal CLA level had been reached. CLA production increased by 80% with 24 h of incubation in milk containing additional skim milk (5%), where the proteins may have facilitated the production of CLA by enhancing the interaction of substrate with the bacteria. CLA production did not decline after 9 h of fermentation and an additional 12 weeks of storage with other commercial starters. This demonstrates the possibility of using this strain as a costarter in the production of CLA-enriched yogurt.

Dietary manipulations to increase the concentration of conjugated linoleic acid in milk

Ten Holstein cows between 8 and 12 weeks in lactation were used to investigate the effect of feeding full-fat soybean, full-fat sunflower, and a Ca-soap source (Profat) on the conjugated linoleic acid (CLA) content of milk. Cows were fed the experimental fat sources in the dosage of 500 g crude fat daily. The results indicated that milk CLA content increased in relation to the linoleic acid concentration of experimental fat supplements, namely full-fat sunflower increased the most and Profat increased the least the CLA concentration in milk. The strength of the correlation was r=0.62 between the linoleic acid concentration in feed and the CLA content in milk. The strength of correlation increased to r=0.69 when both linoleic acid and linolenic acid concentration of feed were used in the calculation. Considering milk production and the daily production of CLA in milk, the following equation described the relationship between the linoleic acid content of fat supplements and CLA concentration in milk: x=167....

Australian plants with potential to inhibit bacteria and processes involved in ruminal biohydrogenation of fatty acids

Conjugated linoleic acids (CLA) are health-promoting fatty acids found in foods derived from ruminant products that are formed in the rumen during bacterial biohydrogenation of linoleic acid (LA). Although selective antimicrobials might increase CLA production by manipulation of ruminal microflora, feeding of antibiotic growth promoters to livestock is declining due to fears of development of antibiotic resistance in human pathogens. This has initiated a search for alternatives, including plants containing bioactive compounds, which can have similar positive effects on ruminal microflora, but without negative impact on animal and human health. In this study we investigated effects of natural compounds in Australian plants on the bacteria and processes involved in ruminal biohydrogenation. Ethanolic extracts and essential oils were obtained from 91 Australian plants collected during 2004/2005 in Western Australia. Minimal Inhibitory Concentrations for the main bacterial species involved in rumen biohydrogenation were established using an agar dilution method, and effects of selected candidates on LA biohydrogenation was examined using an in vitro system with a mixed rumen bacterial population. A wide range of the plant extracts had selective inhibitory effect towards Clostridium proteoclasticum (which forms stearate from LA) without affecting Butyrivibrio fibrisolvens (which forms CLA and vaccenic acid, but not stearate). However, only a few plants, including Acacia iteaphylla and Kennedia eximia, inhibited LA metabolism or stearate formation in the mixed bacterial community in vitro. Further experiments are needed to investigate effects of selected candidates on CLA production in vivo.

Influence of Sodium Glycocholate on Production of Conjugated Linoleic Acid by Cells of Lactobacillus reuteri ATCC 55739

The possible influence of a bile salt on production of conjugated linoleic acid (CLA) by Lactobacillus reuteri ATCC 55739 was evaluated. Cells of the lactobacilli grown in MRS broth with and without linoleic acid (LA, 0.2%) were harvested and washed. The washed cells were added to buffer containing 0.2% LA and incubated 18 h at 37 degrees C. The cells, which had been grown without LA, transformed LA into CLA (mainly c9t11-C18:2) better than did those cells grown with it. When sodium glycocholate (0.3%) was added to the washed cell suspensions, about the same level of CLA was formed as in its absence regardless of whether or not the cells had been grown in broth supplemented with free LA. Thus, glycocholate that occurs in humans did not influence production of CLA by resting cells of the lactobacilli.

Conjugated linoleic acid conversion by dairy bacteria cultured in MRS broth and buffalo milk

To evaluate strains of Lactobacilli, Bifidobacteria and Streptococci for their ability to produce conjugated linoleic acid (CLA) from free linoleic acid (LA). Eight dairy bacteria tolerant to LA were grown in MRS broth containing LA (200 microg ml(-1)) and CLA was assessed. Seven bacteria were able to form CLA after 24 h of incubation, varying percentage conversion between 17% and 36%. Lactobacillus casei, Lactobacillus rhamnosus, Bifidobacterium bifidum and Streptococcus thermophilus showed the highest LA conversion and were inoculated into buffalo milk supplemented with different concentration of LA. The production of CLA at 200 microg ml(-1) of LA was two- or threefold in milk than MRS broth. All evaluated strains were able to produce CLA from high LA levels (1000 microg ml(-1)). The most tolerant strain to LA was Lact. casei. Lacttobacillus rhamnosus produced the maximum level of CLA at high LA concentrations (800 microg ml(-1)). The selected bacteria may be considered as adjunct cultures to be included on dairy fermented products manufacture. Low concentration of LA must be added to the medium to enhance CLA formation. The production of CLA by strains using milks from regional farms as medium offer a possible mechanism to enhance this beneficial compound in dairy products and those the possibility to develop functional foods.

Partial genetic characterization of Stearoyl Coa-Desaturase’s structural region in Bubalus bubalis

Conjugated Linoleic Acids (CLAs) comprise a family of positional and geometric isomers of linoleic acid. The main form of CLA, cis-9, trans-11-C18:2 show positive effects in cancer prevention and treatment. The major dietary sources of these fatty acids are derived from ruminant animals, in particular dairy products. In these animals, the endogenous synthesis mainly occurs in mammary gland by the action of enzyme Stearoyl CoA Desaturase (SCD). Different levels of expression and activity of SCD in mammary gland can explain partially the variation of CLA levels in fat milk. Considering a great fat concentration in bubaline milk and the benefit of a high and positive correlation between fat milk and CLA production, this study was carried on with the intention of sequencing and characterizing part of the gene that codifies SCD in buffaloes. Genomic DNA was extracted from blood samples of lactating bubaline which begins to the breed Murrah. After the (acho que nao precisa desse the) extractions, PCR (Polymerase Chain Reaction) reactions were made by using primers Z ▸◂ D1 and E1 ó ▸◂ F1. The fragments obtained in PCR were cloned into “T” vectors and transformed in competent cells DH10B line. After this, three samples of each fragment were sequenced from 5′ and 3′ extremities using a BigDye kit in an automatic sequencer. Sequences were edited in a consensus of each fragment and were submitted to BLAST-n / NCBI for similarity comparisions among other species. The sequence obtained with Z ▸◂ D1 primers shows 938 bp enclosing exons 1 and 2 and intron 1. The primers E1 ▸◂ F1 show 70 bp corresponding to exon 3 of bubaline SCD gene. Similarities were obtained between 85% and 97% among bubaline sequences and sequences of SCD gene described in human, mouse, rat, swine, bovine, caprine and ovine species. This study has permitted the identification and partial characterization of SCD codifing region in Bubalus bubalis specie.

Purification and gene sequencing of conjugated linoleic acid reductase from a gastrointestinal bacterium, Butyrivibrio fibrisolvens

To characterize the cause for the lack of conjugated linoleic acid (CLA) reductase (CLA-R) activity in the Butyrivibrio fibrisolvens MDT-5 strain that rapidly isomerizes linoleic acid (LA) to CLA without hydrogenation, the CLA-R was purified and its gene (cla-r) sequence was determined. CLA-R was purified to near homogeneity as a 53-kDa monomeric protein from the high CLA-R activity-expressing strain MDT-10. The purified CLA-R recognized conjugated double bonds. Unsaturated fatty acids containing 18 carbons markedly increased the CLA-R expression at the transcriptional level. Complete sequencing of the cla-r gene revealed that the CLA-R is a novel protein. Sequence analysis of the cla-r gene from the MDT-5 strain revealed that the MDT-5 CLA-R protein sequence differed from that of the MDT-10 at four consecutive amino acids. Northern and Western blotting analyses confirmed that the cla-r mRNA and protein are expressed normally in MDT-5. Strain MDT-5 expresses the CLA-R protein that lacks enzyme activity because of mutation, which explains why MDT-5 exclusively produces CLA from LA. The cla-r gene was sequenced for the first time. Exogenous fatty acids affected the cla-r transcription. These results will provide additional knowledge on biohydrogenation, and may also augment the CLA production in the gastrointestinal tract.