Toluene-buffer biphasic system

Fig. 20.3. Calibration plots for methyl mercury obtained in phosphate buffer (curve a) and biphasic system (phosphate buffer/toluene mixture curve b) using the glucose oxidase biosensor and invertase in solution 0.15 pg/mL, I app = + 0.60 V vs. Ag/AgCl reaction time = 5 min and incubation time-10 min.

Fig. 20.4. Calibration curve for inhibition of invertase (0.05 pg/mL) by methyl mercury after 10-min incubation using biphasic system (phosphate buffer/ toluene mixture) and glucose oxidase biosensor. Eapp — +0.60 V vs. Ag/AgCl and reaction time = 5 min.

The synthetic reactions are also carried out in water-immiscible solvents such as benzene, toluene, chloroform, and tetrahydrofuran in the presence of a buffer. The pH of the buffer has been varied from 5.5 to 7.5 to study the kinetics of the polymerization in the biphasic system. The enzyme and the monomer are solubilized in aqueous and solvent phases, respectively. The polymerization reaction is initiated by the addition of hydrogen peroxide in small increments to the aqueous phase. The polymer is formed at the biphasic interphase and falls out of the reaction mixture as polymerization progresses. Precipitate formed is isolated by centrifugation, washed, and purified to determine the nonlinear optical properties. The schematics of the reaction are given in Fig. 4. Other reaction conditions such as the enzyme, monomer and hydrogen peroxide concentration, and reaction time are given above and were published earlier [11]. 

Example 1. Kinetic Resolution in a Biphasic System [20]. A solution of antibody 84G3 (500 mg, 6.67 pmol active site, 0.0086 mol%) in phosphate buffered saline (PBS pH 7.4, 87.5 mL) at room temperature vas added to a solution of ( )-45 (20 g, 77.4 mmol) in toluene (600 mL) in a Teflon tube. The mixture vas shaken at 250 rpm. under argon at the same temperature and the ee vas monitored by chiral-phase HP LG. When ee > 99% vas reached (65 h) the mixture vas cooled to —20 °G for several hours to enable the aqueous phase to freeze. The organic phase vas decanted. The frozen aqueous phase vas left to thaw and extracted vith toluene (3x3 vol. equiv.) and 1 1 EtOAc-toluene (3x3 vol. equiv.). The combined organic phase vas dried over Na2S04, concentrated in vacuo, and purifled by silica gel column chromatography (1 3 EtOAc-hexane) to afford (S)-45 (10 g, 50%). 

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