摘要:

AbstractA synthetic coumarin, 7‐amino‐3‐phenyl coumarin (coumarin‐10), was used to study the uptake of ingested xenobiotics into hemolymph. Larvae were forcefed coumarin‐10 in peanut oil, and hemolymph was extracted and analyzed by fluorescence spectroscopy. Coumarin‐10 entered hemolymph within 5 min, reaching a steady state of concentration within 1 h. Assayed 2 h after feeding, hemolymph titers of 1–5 ng/μl were proportional to log dose between 10 and 100 ng/mg body weight; hemolymph did not reach saturation. Fluorescence spectra of hemolymph in saline revealed that energy was readily transferred from hydrophobic residues of hemolymph proteins to coumarin‐10. Ultracentrifugal density gradients revealed that 94% of absorbed coumarin‐10 was bound to sedimenting proteins while 6% bound to lipophorin. Native polyacrylamide gel electrophoresis (N‐PAGE) on minigels identified two major proteins responsible for binding. Though readily separated by native electrophoresis, these proteins were not fully separable by HPLC using a wide variety of columns. Gel permeation‐HPLC of the sedimenting proteins from hemolymph revealed a single major peak of 480,000 Mr. When upper and lower electrophoretic bands were isolated by preparative N‐PAGE, the upper band (band I) yielded subunits of 75,000 and 71,000 Mr, while the lower band (band II) yielded only one size subunit of 75,000 Mron denaturing (SDS) PAGE. The fluorescent products bound by sedimenting proteins were identified by thin‐layer chromatography and scanning fluorescence densitometry as coumarin‐10 (80% of total) and a polar metabolite (20%). In addition, lipophorin‐containing fractions contained an apolar metabolite (3% of total fluorescence). In vitro binding studies utilizing fluorescent energy transfer demonstrated saturat

ISSN:0739-4462    

DOI:10.1002/arch.940110202

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractIn unparasitized 4th and 5th‐instar larvae ofTrichoplusia niand in 4th‐instar larvae parasitized byChelonussp. 20‐hydroxyecdysone, 20,26‐dihydroxyec‐dysone, and 20‐hydroxyecdysonoic acid were the predominant metabolites formed 2 h after injection of [3H]ecdysone. Other unidentified metabolites were seen, but none seemed to be specific for either parasitized or unparasitized larvae. The major difference between parasitized and unparasitized larvae was seen with respect to the quantity of apolar (unidentified) and polar metabolites (20‐hydroxyecdysonoic acid and unidentified ones), which were produced to a greater extent in parasitized larvae. Ecdysone was rapidly converted into 20‐hydroxyecdysone and the other polar metabolites in all stages investigated, and the parasitoid seemed not to affect the conversion of ecdysone into 20‐hydroxyecdysone. When analyzing the fate of [3H]ecdysone in host and parasite separately, at a stage when the parasite drinks hemolymph of its host, we observed that 10–20% of the radioactivity was recovered from the parasitoid. Analysis of the parasitoid's ecdysteroids revealed that ecdysone and 20‐hydroxyecdysone represented only a small proportion of the recovered labeled ecdysteroids, the majority being apolar and polar metabolites. Our data suggest that the parasitoid takes up ecdysteroids from its host, converts them, and to some extent releases apolar meta

ISSN:0739-4462    

DOI:10.1002/arch.940110203

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractJuvenile hormone esterase (JHE) activity released by the corpora allata (CA) into incubation media (CA‐JHE) was titered daily during the course of the last (fifth [V]) larval stadium ofManduca sexta.This CA‐JHE activity was relatively low during the early last stadium up to the time of commitment (V4), then rose rapidly to a peak on V6. Activity declined sharply almost to precommitment levels by V8, before rising to a second peak on the first day of the pupal phase (P0). This pattern of activity is distinct from that of hemolymph JHE activity, which peaks just prior to wandering on V4 and again just prior to pupation (V9). Although the CA‐JHE and hemolymph‐JHE possess different temporal patterns of activity, isoelectric focusing, gel electrophoresis, and initial studies with selected inhibitors suggest that the enzymes responsible for the CA‐JHE and hemolymph‐JHE activities are similar, but not identical, in nature.Exposure of the V6 CA in vitro to JH II (0.1 μM) or fluoromevalonolactone (FMev; 0.1 mM) produced an approximate fivefold increase and 60% decrease in JH acid synthesis, respectively. Conversely, the same treatments resulted in an inhibition (JH II) and stimulation (FMev) of CA‐JHE activity. These observations suggest that JH may be involved in the direct positive feedback regulation of postwandering larval CA and that the CA‐JHE may also be integrally related to this positive fe

ISSN:0739-4462    

DOI:10.1002/arch.940110204

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractIn accordance with our earlier results, quinone methide formation was confirmed to be the major pathway for the oxidation of N‐acetyldopamine (NADA) by cuticle‐bound enzymes fromSarcophaga bullatalarvae. In addition, with the use of a newly developed HPLC separation condition and cuticle prepared by gentle procedures, it could be demonstrated that 1, 2‐dehydro‐NADA and its dimeric oxidation products are also generated in the reaction mixture containing a high concentration of NADA albeit at a much lower amount than the NADA quinone methide water adduct, viz., N‐acetylnorepinephrine (NANE). By using different buffers, it was also possible to establish the accumulation of NADA quinone in reaction mixtures containing NADA and cuticle. That the 1,2‐dehydro‐NADA formation is due to the action of a NADA desaturase system was established by pH and temperature studies and by differential inhibition of NANE production. Of the various cuticle examined, adult cuticle ofLocusta migratoria, presclerotized cuticle ofPeriplaneta americana, and white puparial cases ofDrosophila melanogasterexhibited more NADA desaturase activity than NANE generating activity, while the reverse was observed with the larval cuticle ofTenebrio molitorand pharate pupal cuticle ofManduca sexta.These studies indicate that both NADA quinone methide and 1, 2‐dehydro NADA are formed during enzymatic activation of NADA in insect cuticle. Based on these results, a unified mechanism for β‐sclerotization involving quinone methides as the reactive sp

ISSN:0739-4462    

DOI:10.1002/arch.940110205

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractN‐Acetylcysteine adducts ofo‐benzoquinones derived from catechol, 4‐methylcatechol, andN‐acetyldopamine were chemically synthesized and characterized by a combination of UV, IR, and NMR spectral studies. Oxidation of catechol, 4‐methylcatechol, andN‐acetyldopamine by cuticle‐bound phenoloxidase fromSarcophaga bullatain the presence ofN‐acetylcysteine resulted in the formation of covalent adducts between catecholic compounds andN‐acetylcysteine. Structural identities of these adducts were established by comparison of their HPLC retention time and UV spectra with those of synthetic adducts and by cochromatography with authentic samples. Although insect cuticle is known to contain only trace amounts of cysteine, the in vitro synthesis of quinone cysteine adducts mediated by cuticular phenoloxidase strongly indicates the occurrence of similar reactions in vivo as well and is in support of Pryor's quinone t

ISSN:0739-4462    

DOI:10.1002/arch.940110206

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

ISSN:0739-4462    

DOI:10.1002/arch.940110201

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY