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  • 1
    ISSN: 0951-4198
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Physics
    Notes: Metastable ions arising from pseudomolecular [M — H]- ions produced by fast-atom bombardment negative-ion mass spectrometry of a range of free ecdysteroids, ecdysteroid conjugates and polar metabolites were investigated by means of linked scanning at constant B/E. Free ecdysteroids displayed daughter-ion spectra which allow 20-hydroxyecdysteroids and ecdysteroids lacking C-20 hydroxylation to be readily distinguished. The ejection of acetic acid from acetylated ecdysteroids was also readily detectable. Characteristic metastable-ion decomposition of ecdysteroid acids was not observed, presumably as a result of charge localization. High-mass daughter ions were also lacking in the case of phosphate conjugates.
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Rapid Communications in Mass Spectrometry 7 (1993), S. 477-481 
    ISSN: 0951-4198
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Physics
    Notes: High-performance liquid chromatography/mass spectrometry (HPLC/MS) with thermospray ionization has been applied to the analysis of free ecdysteroids (polyhydroxylated steroids that serve as arthropod moulting hormones). Of a range of analytical conditions that were tested, optimum results were obtained by the use of re versed-phase HPLC using a methanol + water mixture containing 0.1 M ammonium acetate. The positive-ion spectra of ecdysteroids showed pseudo-molecular ions, [M + H]+, and fragment ions corresponding to the sequential losses of hydroxyl groups. The [M + H - H2O]+ and [M + H - H2O]+ ions were the most abundant for all the compounds tested. Ecdysteroids bearing a C-20 hydroxyl group (e.g. 20-hydroxyecdysone) showed ions characteristic of cleavage of the side-chain from the steroid nucleus. This mode of fragmentation was much less obvious in the case of ecdysteroids lacking C-20 hydroxylation (e.g. ecdysone). Full-scan mass spectra were readily obtained from 1 μg of free ecdysteroids, while a limit of detection of 5 ng (signal-to-noise ratio 3) was attained in the selected-ion monitoring mode. The application of HPLC/MS to the study of ecdysteroids in biological materials is demonstrated by the analysis of extracts of newly laid eggs of the desert locust (Schistocerca gregaria) and pupae of the cotton leafworm (Spodoptera litoralis).
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Archives of Insect Biochemistry and Physiology 2 (1985), S. 39-54 
    ISSN: 0739-4462
    Keywords: ecdysteroids ; fatty acyl esters ; ticks ; Ixodidae ; Boophilus microplus ; h.p.l.c. ; Chemistry ; Food Science, Agricultural, Medicinal and Pharmaceutical Chemistry
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The free ecdysteroid titre determined by radioimmunoassay in adult female Boophilus microplus showed a peak just prior to full engorgement and detachment of the ticks and decreased subsequently to a very low value. In contrast, the titre of polar ecdysteroid conjugates was very low. Ecdysone was the major ecdysteroid at peak titre and was accompanied by much lower levels of 20-hydroxyecdysone. In newly detached ticks, injected [3H]ecdysone was metabolized primarily (80%) into much less polar compounds, which could be resolved into at least three groups by reversed-phase h.p.l.c. These [3H] “apolar” metabolites were transferred to the newly laid eggs, where they accounted for the vast preponderance of ecdysteroids, the level of free hormone being low. Hydrolysis of the three groups of compounds with an esterase preparation from porcine liver yielding [3H]ecdysone, together with the release of [3H] ecdysteroid and fatty acids upon alkaline saponification of the compounds, suggests that they are of a fatty acyl ester nature. The chemical transformation of these “esters” into the corresponding acetonide derivatives indicates that the 2- and 3-hydroxyls of ecdysone remain unsubstituted in these compounds. Several tick tissues, including Malpighian tubules, ovaries, gut, and fat body, metabolized [3H]ecdysone in vitro forming the “apolar esters” as major products. The maternal ecdysteroid “esters” may function as storage forms of hormone (presumably hormonally inactive), which could be hydrolysed enzymically during embryogenesis releasing free ecdysteroids. Such enzymic hydrolysis of [3H]ecdysone “esters” by homogenates from developing eggs of B. microplus has been demonstrated.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
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  • 4
    ISSN: 0739-4462
    Keywords: 26-hydroxyecdysone 26-phosphate ; enzymatic hydrolysis ; ecdysteroid phosphate isolation ; ecdysteroid conjugate ; Manduca sexta ; Chemistry ; Food Science, Agricultural, Medicinal and Pharmaceutical Chemistry
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The major ecdysteroid conjugate present in eggs (48-64 h old) of the tobacco hornworm has been purified by XAD-2 chromatography, C18 SEP-PAK separations, and ion suppression reversed-phase high-performance liquid chromatography. Enzymatic hydrolysis of the conjugate with acid phosphatase from human seminal fluid gave 26-hydroxyecdysone. The conjugate was identified as 26-hydroxyecdysone 26-phosphate by nuclear magnetic resonance and fast atom bombardment mass spectrometry. This compound is also the major conjugate of newly laid eggs (0-1 h old) of the tobacco hornworm. The role for ecdysteroid conjugates is discussed.
    Additional Material: 3 Ill.
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  • 5
    ISSN: 0739-4462
    Keywords: [14C]cholesterol ; radiolabeled ecdysteroid conjugates ; 26-hydroxyecdysone 22-glucosidel ; 26-hydroxyecdysone 26-phosphate ; 3-epi-26-hydroxyecdysone ; 3-epi-20,26-dihydroxyecdysone ; enzymatic hydrolysis ; Chemistry ; Food Science, Agricultural, Medicinal and Pharmaceutical Chemistry
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Following injection into female Manduca sexta pupae, [14C]cholesterol is converted to a radiolabeled C21 nonecdysteroid conjugate as well as ecdysteroid conjugates, which in ovaries and newly-laid eggs consist mainly of labeled 26-hydroxyecdysone 26-phosphate. During embryogenesis, as the level of 26-hydroxyecdysone 26-phosphate decreases there is a concurrent increase in the amount of a new, labeled ecdysteroid conjugate. This conjugate, which is the major ecdysteroid conjugate (9.4 μg/g) in 0- to 1-hour-old larvae was identified as 26-hydroxyecdysone 22-glucoside by nuclear magnetic resonance and chemical ionization mass spectrometry. This is the first ecdysteroid glucoside to be identified from an insect. The disappearance of 26-hydroxyecdysone 26-phosphate in 0- to 1-hour-old larvae indicates that the 26-hydroxyecdysone 22-glucoside is derived from 26-hydroxyecdysone 26-phosphate. 3-Epi-26-hydroxyecdysone was the major free ecdysteroid isolated from these larvae and 3-epi-20,26-dihydroxyecdysone was the next most abundant ecdysteroid isolated. Interestingly, the 0- to 1-hour-old larvae contained the highest levels of 3α-ecdysteroids per gram of insect tissue (8.7 μg/g) to be isolated from an insect, yet there was a complete absence of the corresponding free 3β-epimers. The ecdysteroid conjugate profiles of ovaries and 0- to 1-hour-old larvae are discussed. Methodology is presented that permits the efficient separation of free and conjugated ecdysteroids and nonecdysteroid conjugates (C21-steroid conjugates).
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
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  • 6
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Archives of Insect Biochemistry and Physiology 4 (1987), S. 183-190 
    ISSN: 0739-4462
    Keywords: [14C]cholesterol ; radiolabeled ecdysteroid conjugates ; 26-hydroxyecdysone 2-phosphate ; enzymatic hydrolysis ; Chemistry ; Food Science, Agricultural, Medicinal and Pharmaceutical Chemistry
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Maturing eggs (48 to 64 h old) of the tobacco hornworm, Manduca sexta, contain at least three ecdysteroid conjugates, two of which have been previously identified as 26-hydroxyecdysone 26-phosphate (the major conjugate) and 26-hydroxyecdysone 22-glucoside. In this study we have isolated and identified the third conjugate as 26-hydroxyecdysone 2-phosphate by XAD-2 chromatography, C18 SEP-PAK separation, ion suppression reversed-phase high-performance liquid chromatography, nuclear magnetic resonance, and fast atom bombardment mass spectrometry. This compound is the second most abundant conjugate of ovaries from 4-day-old adult females. The possible role for this ecdysteroid conjugate is discussed.
    Additional Material: 2 Ill.
    Type of Medium: Electronic Resource
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  • 7
    ISSN: 0739-4462
    Keywords: tobacco hornworm ; epiecdysteroids ; ecdysteroid conjugates ; 3-epi-20-hydroxyecdysonoic acid ; mass spectrometry ; NMR spectroscopy ; Chemistry ; Food Science, Agricultural, Medicinal and Pharmaceutical Chemistry
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: [14C]Cholesterol was injected into fifth-instar larvae of Manduca sexta, and the metabolites were isolated and identified from 8-day-old male and female pupae. A major portion of the metabolized cholesterol was esterified either with a sulfate group or with fatty acids. The predominant ecdysteroid metabolites were 20-hydroxyecdysone, 20,26-dihydroxyecdysone, 20-hydroxyecdysonoic acid, and 3-epi-20-hydroxyecdysonoic acid. Smaller amounts of ecdysteroids were identified as conjugates of 26-hydroxyecdysone, 3-epi-20-hydroxyecdysone, 20,26-dihydroxyecdysone, and its 3α-epimer. The metabolic profiles were similar for both male and female pupae. The two ecdysteroid acids were identified by nuclear magnetic resonance spectroscopy and chemical ionization mass spectrometry and by mass spectral analyses of their methyl esters. Detection of 3-epi-20-hydroxyecdysonoic acid as a major metabolite is significant, as its occurrence has been scarcely reported. 3-Epiecdysteroid acid formation is discussed as a possible ecdysteroid-inactivating pathway that may be operating specifically in lepidopterous insects or in particular developmental stages such as eggs or pupae.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
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  • 8
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Archives of Insect Biochemistry and Physiology 10 (1989), S. 241-253 
    ISSN: 0739-4462
    Keywords: ecdysteroids ; hydroxylation ; 20-hydroxyecdysone ; Spodoptera littoralis ; protein kinase ; phosphoprotein phosphatase ; Chemistry ; Food Science, Agricultural, Medicinal and Pharmaceutical Chemistry
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Evidence is presented for the reversible activation-inactivation of the microsomal ecdysone 20-monooxygenase from fat body of the cotton leafworm, Spodoptera littoralis, in a manner commensurate with reversible changes in its phosphorylation state. The activity of the monooxygenase was higher following preincubation with fluoride (an inhibitor of phosphoprotein phosphatases) than in its absence. Preincubation with alkaline phosphatase or with cAMP-dependent protein kinase resulted in appreciable diminution or enhancement, respectively, in monooxygenase activity. Activation of ecdysone 20-monooxygenase activity could also be effected by incubation with a cytosolic fraction in the presence of cAMP, ATP, and fluoride; this activation was prevented by a cAMP-dependent protein kinase inhibitor. Similarly, inactivation of the monooxygenase was achieved by preincubation with cytosol, the effect being enhanced by Ca2+-calmodulin or by Mg2+ ions. The combined results provide indirect evidence that the microsomal ecdysone 20-monooxygenase exists in an active phosphorylated form and an inactive dephosphorylated form, interconvertible by a cAMP-dependent protein kinase and a phosphoprotein phosphatase.
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
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