inorganic chemistry

If the terbium(macrocycle) complex was covalently bound to
the stationary phase of a column, such as silica or alumina, a
dilute DPA solution could be readily concentrated. When applied
to environmental samples, this protocol would involve sample
collection, DPA release via physical (heating, pressure) or chem-
ical (germinant or lysozyme) means, and filtration to remove any
cell debris or other material. The dilute dipicolinate solution,
buffered to pH 7–10, could then be passed through the column
containing [Tb(DO2A)]þbound to the solid substrate. The high
binding affinity of the terbium(macrocycle) for DPA at this pH
would cause the dipicolinate to be retained on the column. After
saturation, the addition of a small aliquot of acidic solution
(pH2) would protonate the macrocyclic ligand and release [Tb
(DPA)]þ, which could then be quantified using fluorescence spec-
troscopy and correlated to the original filtrate volume to yield a
value of spores per milliliter of solution. The column could then
be treated with a TbCl 3 solution at neutral pH to reform the ter-
bium(macrocycle) in the solid phase for reuse. Current limits of
detection of bacterial spores in environmental samples for spec-
troscopic techniques are in the 10^3 – 104 spores mL^1 range
(81,99,158). The proposed technology could improve the current
LOD by several orders of magnitude.
We are currently working to modify the DO2A ligand to
append to solid substrates. Recent results indicate a negligible
change in binding affinity when one of the acetate moieties is
exchanged for an amide group with similar resilience to pH
variations ( 172 ).

V. Conclusions

We have summarized the common attributes of ancillary ligands
in improving detection techniques based on sensitized lanthanide
luminescence and also introduced a new model for enhanced bind-
ing affinity. We suggest that binding affinities can be increased by
an order of magnitude or more by ligand-induced anisotropy in

TABLE VI

PROTONATIONCONSTANTS OFRELEVANTLIGANDS.

Ligand pKa1 pKa2 pKa3 pKa4 References

DPA –1.05 2.22 5.22 – ( 159 )
DO2A 2.55 3.85 9.55 10.94 ( 96 )

38 MORGAN L. CABLEet al.