Bioconversion Of Linoleic Acid Into The Conjugated Linoleic Acid By Lactobacillus Plantarum

Conjugated linoleic acid (CLA) has attracted much attention as a novel type of biologically beneficial functional lipid. For example, CLA reduces carcinogenesis, atherosclerosis, and body fat. Presently, CLA is produced through chemical isomerization of linoleic acid (LA), but by-productions of unexpected isomers are produced, which is not beneficial for the use of CLA in food and medicine. However, CLA isomers are much simpler through biology isomerization of LA than through chemical. And it will bring a bright promise for application of CLA, which is isomerized by lactic acid bacteria.The process of CLA production by Lactobacillus plantarum was studied. The research included bacteria isolation, screening, identification and induced mutation, also fermentation dynamics, CLA production from washed cells, LA isomerase purification and its properties were studied, in the end the CLA concentration in milk was increased by fermentation. Results as the follows:1) Lactic acid bacteria L29, with a high ability to produce CLA, was isolated from pickle. L29 was identified as a strain ofLactobacillus plantarum (confidence level 96.1%) .2) Lplantarum L29 was induced by ultraviolet radiation and DES, then a mutation strain LT2-6 was screened out, by which CLA production could increase 18.6 %.3) Fermentation dynamics curve was obtained by the study of the Lplantarum LT2-6 fermentation in batches in 5 L fermentor. A mathematics model was established based on the cells growth of L. plantarum LT2-6, production CLA accumulation and glucose metabolism.4) Washed cells of Lplantarum LT2-6 as the catalyst, the optimal reaction condition for CLA production was: temperature 30℃, pH 7.0, LA concentration 1.5% (w/v) , cells concentration 20%(w/v). After reacted for 64 h, CLA concentration was up to the maximal (7.86 g/L), and 52.4 % of LA was conversed to CLA. Then fatty acid was extracted, methylated and determined by gas chromatography analysis, the majority of resultants was cis-,trans-9,ll-CLA.5) Molecular weight of the LA isomerase of Lplantarum LT2-6 was about 48.8 KDa. The optimal reaction condition for the LA isomerase was pH 6.0, 30℃. This enzyme was non-resistant to heat relatively, if temperature was up to 50℃, stabilization ability of the enzyme declined soon. Kinds of metal ion had different effects on the enzyme activity, lowering sequence was Mg2+ > Ca2+ > Mn2+ > Fe2+ > Zn2+ > Na+. Km and V,,,, dynamics parameters of LA isomerase, were respectively 12.46 mol/L, 1.933x10-1moVh.mg.6) In the process of fermentation of skim milk by Lplantarum LT2-6, 37℃ was suitable for fermentation. The growth of cells was accelerated significantly supplementing with glucose in milk, so production of CLA was increased. Also small molecular nitrogen source materials supplemented in milk had the same effects on CLA production. However, when LA concentration supplemented in milk was more than 0.075 %, the growth of cells and CLA production were restrained. Production of CLA was active during the end of log phase and stationary phase. Fermentation for 24 h, CLA production was themaximum. Results showed that the majority of CLA in resultants was cis-,trans-9,ll-CLA. 0.075 % of LA supplemented in milk, and 50% of LA was bio-conversed into CLA in 37℃, fermentation for 24 h. After fermentation by Lplantarum LT2-6, CLA production was higher 5-6 times with skim milk supplemented LA than with market yoghourt without LA.