User:Julian C. Leos/methoduanl
Formation of Nanoreactor
Employ the microfluidic method for the production of liposomes as nanoreactor because it is a simple and low cost alternative.
This method involves dissolving a lipid in ethanol solution, which is rapidly injected counter-flow to an enzyme solution with deionized water to produce spontaneously vesicles, SUV primarily, encapsulating enzymes. Subsequently, we performed an ultrafiltration to removed the residual ethanol.
Considerations to decrease the size of the liposomes:
• Decrease the size of the syringe
• Increased hydrodynamic pressure
• Decreased lipid concentration alcohol solution
• Decrease in the phase transition temperature in the lipid bilayer
• No cholesterol (decreased membrane rigidity)
• Changing the flow rate
Recommendations for the formation of liposomes
The higher the concentration of lipids greater proportion of the aqueous phase may be entrapped.
The average life of a liposome increases with increasing lipid dose.
The encapsulation rate increases to make slower hydration.
The addition of charged phospholipids increases the distance between the lipid membranes.
Caracterizacion del nanoreactor
Dynamic light scattering is therefore the most used method to estimate the average hydrodynamic radius of particles in suspension and polydispersity index and covers a range from a few nanometers to microns.
Because the last reaction that occurs in the liposome is the decomposition of hydrogen peroxide, the enzyme activity of the nanoreactor will be evaluated by UV spectrophotometry monitoring the products delivered by the catalase.
The system consist in having the nanoreactors in contact with the uric acid, then the serie of reactions catalyzed by the enzymes to the final products, which are the allantoin, water and oxygen.
In this way, the enzyme activity will be evaluated in the industrial application with the corresponding substrate and products