Acquisition uricase with high uric acid decomposing activity from bacteria as raw materials to product drugs for prevention and treatment of increased blood uric acid. 

Theoretical results:

• 50 strains of uricase producing bacteria were isolated from soil. A strain CN3 have the ability to produce high levels of extracellular enzyme in a short time had been chosen. This strain was identified as Pseudomonas putida.
• Have found the suitable conditions for P. putida CN3 product high extracellular uricase: 0.5% glucose, 0.3% yeast extract, 0.5% uric acid, initial pH 8, the culture temperature 30°C, the culture time 48h, shaking speed 250 rpm.
• The strain CN3 was incubated in 5L fermenter at 30°C, stirring speed 250 rpm, the aeration blower 1.0 vvm, automatically controlled pH 8.0 ± 0.1. After 30h of fermentation, the activity of total uricase reached 1.21 U/ml with the specific activity 1.33 U/mg.
• The crude extracellular uricase from P. putida CN3 was purified by filtration and concentrated enzyme by ultrafiltration membrane 30 kDa and 8 kDa on the Tangential Flow Filtration System Millipore to collect enzyme with activity ~ 5 U/mg with purity levels increased 3.74 times. Further purified by fractional precipitation with ethanol concentrations of 70-80%, add 0.2% CaCl₂ to collect uricase with activity 7.15 U/mg for the purified level increased 5.38 times compare the crude enzyme.
• Identified characteristics of the extracellular uricase biosynthesis from P. putida CN3: Molecular weight ~ 25 kDa, optimum pH and temperature of this enzyme is 8.5 and 35°C, pI 4.4, Km 3.31 mM. This enzyme was stable in the pH 7-9 at 4°C for 24 hours and was denatured at temperatures > 45°C, but it still kept 50% of initially activity at 55°C. Enzyme sustainable at storage temperature -20°C and 4°C. Metal ions, Ca²⁺ and Cu²⁺ also improved enzyme activity to 112 and 120%, while the metal ions, Co²⁺, Mn²⁺, Mg²⁺, Fe²⁺, Zn²⁺, and Al³⁺ influence inhibitory activity of uricase. 50 mM borate buffer at pH 8.5 was the best for P. putida CN3 produced extracellular uricase.
• Produced 10g of lyophilized uricase.
• Developed a procedure to produce extracellular uricase from strains of P. putida CN3.

Applied results: The results are significant for the future application.

Pseudomonas putida was first found to have extracellular uricase. Researching on uricase from bacteria to produce drug for prevention and treatment of increased blood uric acid is a new problem in Vietnam.

In the former, the ratio of high blood uric acid in Vietnamese estimated about 1%-2%, but now that figure was much higher. To apply results into the life, need to research continuously:

• More purified enzyme products to obtain higher purity level
• Research toxicity levels of uricase from the strain of P. putida CN3 on animals
• The half-life of uricase
• The ability allergy of enzyme in animals
• Ability to adapt to the environment and the enzymes in the human stomach
• Study the stability of the product