Aqueous, buffers phases

First, the effects of aerobic and anaerobic culture conditions on toxaphene degradation were studied with washed P. putida cells grown on camphor and incubated with no readily usable carbon source. The radioactivities remaining in water after extraction with n-hexane were used as an indicator of metabolic activity. This was further extracted with ethyl acetate after acidification to divide this "total polar metabolites" fraction into aqueous buffer phase and ethyl acetate phase, i.e., the total polar metabolites reported refer to summation of the aqueous buffer and ethyl acetate soluble phases (Table 4). All radioactivities have been corrected by zero time controls and autoclaved 8 hr controls are included in each experiment. 

Of the total recovered radioactivities after AgNO, treatment (water soluble plus AgCl) of the aqueous buffer phase takem from C1-toxaphene cultures (Table 5), 30.3% was found as water soluble metabolites and 69.7% as free chloride ion under aerobic conditions. Under anaerobic conditions, 17.0% was found as water soluble metabolites and 83.0% as free chloride ion. 

Table 5. AgNO, complexation of aqueous buffer phase from washed P. putida cell incubations with...

Fractions Aerobic % C1 Radioactivities of aqueous buffer phases Anaerobic... 

If 20 ml of aqueous buffer phase were extracted with 20 ml of chloroform there would be 1 part of the base in the aqueous phase to 50 parts in the chloroform layer. Since 60 ml of chloroform are used in the extraction there will be 1 part of the base remaining in the aqueous phase and 150 parts in the chloroform layer. 

As noted above, enzymatic hydrocyanations are preferably performed at pH < 5, to suppress the non-enzymatic bacl ound reaction whereas the pH optimum of the common nitrilases is 7. A compromise pH is obviously required and we accordingly assessed the effects of the pH on the MeHnL-mediated hydrocyanation of benzaldehyde (2a, see Figure 16.3) in a biphasic aqueous-diisopropyl ether (DIPE) medium. We found that enantioselectivity was maintained at pH 5.5, which we adopted as a compromise pH for the bienzymatic reactions, provided that the aqueous buffer phase accounted for <10% of the reaction volume. PfNLase was the obvious choice for the second step as it stayed active at pH 5.5 and converted (S)- and (R)-la at comparable rates. 

Poochikian and Cradock used acetonitrile - aqueous phosphate buffers in combination with octadecyl columns for the analysis of heroin and its hydrolysis products. The retention times were found to be shorter with increasing percentages of acetonitrile and with more strongly buffered mobile phases. Also, the peak sharpness increased under such conditions. Optimum pH was between 5 and 8.7. The order of elution was not effected by changes in pH, buffer strength, or percentage of acetonitrile in the mobile phase. As well as mobile phases consisting of acetonitrile and aqueous buffers, phases in which acetonitrile has been replaced by methanol have been used in combination with octadecyl... 

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