Studies On Hyaluronic Acid By Streptococci Zooepidemicus

Hyaluronic acid ( HA ) is a high molecular weight linear polysaccharide composed of repeating disaccharide units of D-glucuronic acid (GlclIA) and N-acetyl-D-glucosamine (GlcNAc) linked by 3-1-3 and P-1-4 glycoside bonds. Because of its special physiological action, extraordinary rheological property and moister-holding function, hyaluronic acid is extensively applied to areas including medical research, clinical therapy, and cosmetic industry. Due to shortage of material source, high cost, and low output, more attention has turned from tradition method of producing hyaluronic acid by extraction of animal tissues to microbial route. HA has high affinity to combine with water. The water solution of HA has a property of high viscosity. HA can be added to cosmetics to keep the moisture of skin. HA can also be used in ophthalmology, arthritis heating and other medical areas. HA exists in connective tissues of mammals and the capsule of group A and group C of Streptococci. The aim of this thesis is to study the production of HA by fermentation of Streptococci zooepidemicus.The objective of this dissertation was to develop a fermentation bioprocess by a strain of Streptococci zooepidemicus. The research content included, (l)the breeding of HA producing strains from protoplast of Streptococcus zooepidemicus^ NTG induced mutagenesis; (2) studies on fermentation condition of H-35; (3) isolation and purification of HA in fermentation broth;(4) studies on the properties of HA. Main results are as follows: 1. Hydrolysis of cell wall with lysozyme, the highest regeneration rateof protoplast was obtained. The optimum concentrations of lysozyme.. temperature and time were 0. 6mg/mL> 30C and 20 minutes respectively. The yield of H-35 was 250% higher than that of the original strain.2. The optimum culture conditions of producing HA : initial concentration of glucose 4%, yeast extract 2%, NaCl 0.2%, KH2P04 0.2%, MgS04 7H 20 0.06%, 20mL medium in 250mL flask, the temperature 35C, initial pH7.0 , fermentation period 18hours. Finally the yield of hyaluronic acid was 1.43g/L.3. Successful purification of the crude HA was achieved by anion exchange resin DEAE-cellulose column chromatography using 0. 6mol/LNaCl solution as an eluant.