TABLE 1

Effect of various solubilizing agents on the solubility of rifalazil and production of 32-hydroxy-rifalazil catalyzed by MLM

Reaction conditions: 100 mM potassium phosphate buffer, pH 7.4, containing 20 μl/ml MLM, 100 mg/l rifalazil, 0.4 U/ml glucose-6-phosphate dehydrogenase, 1.3 mM NADP, 3.3 mM glucose-6-phosphate, and 1.3 mM MgCl2. Total reaction volume was 1 ml. Reaction temperature 37°C, shaking rate 150 rpm, reaction time 16 h. Under these experimental conditions and using an MLM batch specific to this experiment, the volumetric productivity of the additive-free system was 0.6 mg product/l reaction volume. Volumetric productivity is determined as the amount of product generated in the unit reaction volume over a specified period of time.

Solubilizing Agent
Tested Concentration Range
Solubility of Rifalazil
Estimated Relative Volumetric Productivity for 32-Hydroxy-rifalazila
mg/l
None 10 1.0
Tween 80 0.01-1% w/v >1000b 0.6c
Alkamuls EL-620 0.01-1% w/v >1000b 0.6c
Polyvinylpyrrolidone, molecular mass 10 kDa 0.2-5% w/v >100b 0.8c
Polyvinylpyrrolidone, molecular mass 40 kDa 0.2-5% w/v >100b 1.1c
Bovine serum albumin 1-20 mg/ml >100b 0.6c
Glycerol
1-20% v/v
10-50
0.3d

  • ↵ a Estimated from the HPLC peak area.

  • ↵ b Estimated at the lowest tested concentration of the solubilizing agent.

  • ↵ c Estimated at the lowest tested concentration of the solubilizing agent; increasing concentrations of the agent resulted in a significant decrease of the estimated yield.

  • ↵ d At 20% glycerol.