Conjugation of diazinon with glutathione,and γ-glutamyl transferase activities in P. Americana (L)

1. The endogenous level of glutathione in the cockroaches from insecticide-treated localities (13.07 × 10⁻⁴ ±0.58 mg/mg protein) was significantly higher (P < 0.05) than those sampled from untreated localities (9.97 × 10⁻⁴ ± 2.84 mg/mg protein).2. A correspondingly higher level of the γ-glutamyi hydrolase activity was observed in cockroaches obtained from the treated area.3. Incubation of homogenates of cockroaches from these two localities with 1.5% diazinon showed no apparent difference in their conjugation of this insecticide with glutathione. However, the final residual amounts of GSH were higher in cockroaches regularly exposed to diazinon.     

Purification of β-glucosidase from the salivary glands of the green rice leafhopper, Nephotettix cincticeps (Uhler) (Hemiptera: Cicadellidae), and its detection in the salivary sheath

The green rice leafhopper, Nephotettix cincticeps (Uhler), is an insect pest of rice and discharges β-glucosidase (EC 3.2.1.21) from its salivary glands during feeding. To investigate the biological function of this enzyme, we purified it from the heads of 18,000 adult females by acetone precipitation and a series of chromatography steps: gel filtration, cation-exchange chromatography, metal-affinity chromatography and hydrophobic interaction chromatography. During cation-exchange chromatography, β-glucosidases were eluted in three peaks (isozymes). These β-glucosidases were monomeric proteins of 58 kDa as estimated by SDS-PAGE and 62 kDa based on gel filtration. All of the purified β-glucosidase isozymes exhibited maximum activity for p-nitrophenyl β-glucoside (NPGlc) and p-nitrophenyl β-galactopyranoside (NPGal) at pH 5.5 and 5.0, respectively. There was no significant difference in substrate specificity among the three isozymes. The Km values were estimated to be 0.13 μM for NPGlc and 0.9 μM for NPGal. Among the oligosaccharide substrates examined, laminaribiose (Glc β1-3 Glc) was the most extensively hydrolyzed, sophorose (Glc β1-2 Glc) and cellobiose (Glc β1-4 Glc) were comparatively well hydrolyzed, and gentiobiose (Glc β1-6 Glc), lactose (Gal β1-4 Glc), laminaritriose, cellotriose and cellotetraose were poorly hydrolyzed. Among the glycoside substrates examined, salicin was considerably well hydrolyzed. β-Glucosidase was detected in the salivary sheaths by activity staining with a fluorescent substrate. The salivary β-glucosidase of N. cincticeps may be involved in the hydrolysis of a phenol glucoside present in the saliva, which is a step in the solidification of gelling saliva to form salivary sheaths.