45
indicating competitive inhibition. The dissociation constant (Kp) for
enzyme*ligand complex could be graphically obtained. These results are
shown in Table I.
Quantitation of Matrix Bound Ligand
The immobilized 4-phenylbutylaraine was removed from the matrix by
solvolysis over a period of 5 months at 4C. The supernatant was removed
from the matrix by filtration followed by washing of the matrix with
methanolic tris*Ca++ buffer. The washings were combined and an absorption
spectra taken. Figure 12 shows a composite spectra of pure 4-phenylbutyl-
amine and the supernatant from the matrix. Figure 13 shows composite
spectra of matrices suspended in buffer. The break at 270 nm is where
the scan was stopped and the matrix resuspended by mixing. The results
show the successful solvolysis of ligand from matrix thus providing a
method for determining the degree of substitution of the matrix.