pipetted into each flask. After mixing, flasks
were placed in the dark for 2 hr for color
development at 20-25 C. Next, each solution
was transferred to a 250-m1 separatory funnel
and the flask rinsed with 10 ml of CHCI3. After
shaking for about I min, most of the color
had been transferred to the CHCI~ layer, which
was drawn off into a 50-ml volumetric flask.
The extraction procedure was repeated three
times with 10-ml volumes of CHC13. Finally,
10 ml of HC1-MeOH solution were added
which changed the color from yellow-orange
to purple. The flasks were made to mark with
CHCI~ and the colored solutions transferred
to matched 19- by 150-mm colorimeter tubes.
The absorbance was measured at 555 m~ against
a reagent blank using a Coleman 4 Model 14
spectrophotometer.