Porphyric Insecticides - Chlorophyll Biosynthesis and Technological Applications - page 415

Table 18.10 Response of isolated organs and tissues to incubation with ALA and Dpy or Oph a

Proto content after 5 h incubation in darkness the media
listed below (nmol/per 100 mg protein)
Buffer Insect Oph Tissue ALA Dpy ALA + Dpy Oph ALA+

T . ni

Midgut 0.6 0.6 4.4 3.0 8.6 6.5

- b

Integument + fat body 0.2 0.3 1.7 1.5 -

Integument 0.2 0.1 - - - -

Fat body 0.6 0.6 1.9 5.1 4.8 2.1

H . zea Midgut 1.8 0.7 15.9 20.5 23.2 29.2

Integument + fat body 0.5 0.7 1.9 2.1 - -

Integument 0.2 0.2 0.6 0.6 0.7 0.8

Fat body 0.1 0.8 10.3 11.2 8.6 8.3

B . germanica Male abdomen + gut 0.7 0.8 6.4 6.2 - -

Male gut 3.0 2.0 - - 4.8 2.3

Female gut 2.7 4.8 - - 6.1 6.9

Male abdomen - gut 0.2 0.3 - - 2.9 4.5

Female abdomen - gut 0.4 0.3 - - 1.5 1.3

A . grandis Abdomen + gut 0.5 0.8 1.9 1.7 - -

Adapted from Lee and Rebeiz ( 1995 )
a Incubation was in 5 mMALA, 1.5 mM Dpy orin ALA + Dpy or Oph at pH 5.5 for 5 h in darkness
b Not determined

and tissues to incubation with ALA + Dpy or ALA + Oph. In these experiments,
the following insects were used: Adult Blattella germanica (German cockroach),
Adult Anthonomus grandis (cotton boll weevil), fifth instar larvae of Heliothus zea
(corn earworm) and fifth instar larvae of T . ni (cabbage looper).
One week-old cockroaches, were starved overnight, and anesthesized with CO 2
prior to dissection. Abdomens of four cockroaches were removed with small
surgical scissors, rinsed in cold buffer (0.1 M potassium phosphate, pH 7.0), cut
into small pieces, and transferred to a small plastic petri dish (5 cm in diameter),
containing 3 ml of incubation buffer as well as ALA and a modulator at a specific
PH. Abdomens of 3 day old, adult cotton boll weevils were excised and processed
in a similar manner. For larvae of H . zea an T . ni , midguts were dissected and placed
in ice-chilled phosphate buffer pH 7.0, cleared of tracheal branches and Malpighian
tubules under a stereoscopic microscope, slit open and cleared of residual food
remains, and rinsed in fresh phosphate buffer. Fat bodies were also collected from
dissected larvae with a small spatula, washed twice in phosphate buffer and stored
in cold fresh buffer. Integuments were removed from the mid-section of the body,
cut into small pieces and placed in fresh cold phosphate buffer along with the fat
bodies and tracheal branches. Organs and tissues prepared as just described were
placed in incubation buffer containing ALA, a modulator or ALA + modulator
(Table 18.10 ), and were incubated in darkness for 5 h. After incubation the organ
and tissue pieces were homogenized in acetone: 0.1N NH 4 OH (9:1 v/v) and after
centrifugation the acetone extract was used for tetrapyrrole determination by