The Preliminary Research On Extraction Process Of ε-poly-lysine From The Fermentation Broth

ε-poly-Lysine(ε-PL) is a homo-polymer of L-lysine generated by micro-organisms, withthe molecular weight generally ranging from2500to4500Da，mainly used as biological foodpreservation. Japan is always dominated in the ε-PL research, in recent years, it also hascaused the domestic scholars’ widespread interests because of its higher application value andbroad market prospect. At present, domestic research on ε-PL focuses on improvement offermentation production by screening and mutagenesis of ε-PL producing strains andoptimizing fermentation strategy. However, with fermentation production increasing,extraction of ε-PL has become the important research content. The extraction process restrictsthe quality of the products and their industrial production cost, therefore, the study ofextraction technology has a real economic significance.In this frame, around establishing a primary extraction processs and on the basis ofpreliminary study, we mainly optimized three key unit operations: the removal of protein, theadsorption of chroma and ultrafiltration concentration. The structure and antibacterial activityof the purified sample were identified. The main results obtained were as follows:(1) With ε-PL loss rate and protein removal rate as the indexes, compared the effect ofprotein removal by flocculation and the heat treatment, as a result, when the fermentationbroth pH was adjusted to6.0and heated to80℃with maintenance of60min, the proteinremoval rate reached82%-85%, ε-PL loss rate was less than5%. With ε-PL loss rate andchroma removal rate as the indexes, we optimized the decolorization conditions: a addition of1%active charcoal LT-720, fermentation broth pH4.0, time2h, temperature85℃, thechroma removal rate was about75-80%, ε-PL yield reached91.3±2.0%. With ε-PL loss rateas the index, compared the effect on concentration and purification of ε-PL by differentmembranes with molecular weight cutoff of10kDa,5kDa,1kDa, as a result,1kDaregenerated cellulose membrane was chosen with ε-PL yield96.1±1.0%.(2) Based on optimization of the unit operations, combined with ion exchange andorganic solvent precipitation technology, we established the primary extraction process ofε-PL, the total yield was51.2±0.5%, protein removal rate was up to98.8±0.5%. Afterultrafiltration, the chlorine content of the samples varied from31.8±1.1%to28.8±1.4%, ashvaried from29.4±1.8%to16.7±3.8%; After organic solvent precipitation, the chlorinecontent dropped to19.1±0.5%, ash fell to3.2±0.8%.(3) The finally purified sample was hygroscopic yellow powder, protein content was2.5±0.5%, the purity was88.6±1.4%. By amino acid analysis of the sample hydrolysate,L-lysine was identified as the polymer monomer of ε-PL. Through MALDI-TOF massspectrometry analysis, the number average molecular weight was3642.5, the weight averagemolecular mass was3695.1, polymerization degree was28.4. The molecular weightdistribution was in general accord with the other three products. Antibacterial experimentproved that the ε-PL sample exhibited a good antibacterial activity to yeast, staphylococcusaureus, gas bacill1us, escherichia coli, proteus.