Insect Control: Biological and Synthetic Agents - Index of

A large synthetic effort has gone into the modification
of the forosamine and rhamnose moieties, as
well as the core tetracycle (Crouse and Sparks, 1998;
Crouse et al., 2001; Anzeveno and Green, 2002; Kirst
et al., 2002b) (see Section 6.7). However, other
approaches to macrolide modification are now possible.
Sequencing the genes involved in spinosyn biosynthesis
(Waldron et al., 2000, 2001; Madduri et al.,
2001) has allowed the production of novel spinosyns
through biotransformation and modification of the
biosynthetic pathways. For example, a genetically
engineered strain of Saccharopolyspora erythraea
expressing the spnP glycosyltransferase gene was
used to produce biotransformed spinosyns, wherein
the b-d-forosamine moiety was replaced by a-lmycarose
at the C17 position (Gaisser et al., 2002).
Alternatively, modifications to the basic tetracycle
have been obtained via loading module swaps in the
PKS, leading to a variety of novel substitutions at
C21 and other positions (Burns et al., 2003; Martin,
2003). As one example, the ethyl group at C21 has
been replaced with an n-propyl group (Martin,
2003), resulting in a unique spinosyn as active as
spinosyn A (Tables 1 and 3).
6.2.4. Physical Properties of the Spinosyns
The spinosyns are high molecular weight (approximately
732 Da), relatively nonvolatile molecules
(Table 4). As with the biological activity, small
changes in structure can have far larger effects
than might otherwise be expected. Although spinosyns
A and D only differ by the presence of a methyl
group at C6 (Table 1), this has a surprising effect on
the physical properties, especially solvent solubilities.
For example, spinosyn A is soluble in water at
about 8.9 mg 100 ml 1 , while the water solubility of
spinosyn D is only 0.05 mg 100 ml 1 . This pattern
holds true for other solvents as well (Table 4).
6.3. Pharmacokinetics of Spinosad
6.3.1. Metabolism of the Spinosyns
6: The Spinosyns: Chemistry, Biochemistry, Mode of Action, and Resistance 213
6.3.1.1. Mammalian and avian spinosyn
metabolism Studies of 14 C-spinosyns A and D
metabolism in rats identified fecal excretion as the
major route of elimination for both spinosyns A and
D. In the fecal material, between 52% (males) and
28% (females) of the administered dose of spinosyn
A was excreted in the feces as parent or metabolites
within 0–24 h post-dosing. Of this material
only 6.4% (males) and 5.4% (females) was spinosyn
A (Domoradzki et al., 1996). For spinosyn D,
between 67.7% (males) and 73.3% (females) of
the applied dose was present in the feces, of which
34.5% (males) and 35.2% (females) was the parent
spinosyn D (Domoradzki et al., 1996). Thus, for
both spinosyns A and D a substantial amount of
metabolism had taken place in the first 24 h following
administration. The primary pathways for the
spinosyns in rats involved loss of a methyl group on
the forosamine nitrogen (N-demethylation), and/or
loss of one of the O-methyl groups on the rhamnose
(O-demethylation) (Figure 4), both followed by
conjugation. Conjugation of the parent was also
noted as an important pathway in rats (Domoradzki
et al., 1996).
In lactating goats, following 3 days of dosing with
either spinosyn A or D, eight metabolites of spinosyn
A and five metabolites of spinosyn D were
detected. As with rats, N-demethylation was an
important metabolic route. In addition, hydroxylation
of the macrolide ring was noted as a primary
metabolic pathway (Rainey et al., 1996) (Figure 4).
In studies with poultry, N-demethylation of
the forosamine nitrogen and O-demethylation of the
rhamnose moiety were also found to be the two primary
metabolic pathways (Magnussen et al., 1996).
In the latter case, the 2 0 /4 0 -O-demethyl metabolites
(i.e., spinosyns H and K, respectively, for spinosyn
A metabolism) predominated over the 3 0 -O-demethyl
metabolites. Another pathway, involving removal
of the forosamine sugar to form the C17 PSA, was
identified, but appears to be secondary to the N-and
O-demethylation (Magnussen et al., 1996) (Figure 4).
As demonstrated by these three studies, spinosyns
A and D are readily metabolized in vertebrate
and avian systems. Furthermore, N-demethylation
of the forosamine nitrogen is a metabolic route that
predominates in all three species. O-Demethylation
of the rhamnose sugar is also important in rats and
poultry. These particular pathways are both consistent
with oxidative metabolism via monooxygenases
and/or the action of glutathione transferases. In the
rat studies, glutathione conjugates were noted for
O-demethylated metabolites as well as for the parent
spinosyns (A and D). The presence of hydroxylated
macrolide metabolites in the goat provides
direct support for involvement of monooxygenases
in spinosyn metabolism.
6.3.1.2. Spinosyn metabolism in insects Spinosyns
A and D appear to have a number of potential sites
for metabolism, including N-demethylation of the
forosamine, O-demethylation of the rhamnose, epoxidation
of the 5,6 or 13,14 double bonds, opening
of the macrocyclic lactone, etc. As shown above (see
Section 6.3.1.1), some of these potential metabolites
have been observed. However, compared to more
conventional insect control agents, the spinosyns are