Conversion of clionasterol into both fucosterol and isofucosterol by the insectTenebrio molitor

Summary [7,7-³H₂]Clionasterol was synthesized and fed, together with [4-¹⁴C]sitosterol, toTenebrio molitor larvae; fucosterol and isofucosterol, recovered from the sterol fraction were found to be doubly labeled, indicating that clionasterol is converted into both the ethylidenic compounds.     

Nest cell lining of the solitary beeHylaeus bisinuatus (Hymenoptera: Colletidae)

The nest cell lining ofHylaeus bisinuatus (Hymenoptera: Colletidae) was shown by high-resolution solidstate [¹³C]NMR to be composed of lipid polymer and protein. The lipid polymer was shown by reduction and subsequent GC/MS analysis to be comprised of -hydroxy fatty acids (C₂₀, C₂₂, C₂₄ and C₂₆) and fatty alcohols (C₁₆ to C₃₀). The protein portion of the lining had a silk-like amino acid composition.     

Aberrant,canavanyl protein formation and the ability to tolerate or utilize L-canavanine

Summary L-Canavanine, 2-amino-4-(guanidinooxy)butyric acid, and L-arginine incorporation into de novo synthesized proteins was compared in six organisms. Utilizing L-[guanidinooxy¹⁴C]canavanine and L-[guanidino¹⁴C]arginine at substrate saturation, the canavanine to arginine incorporation ratio was determined in de, novo synthesized proteins.Caryedes brasiliensis andSternechus tuberculatus, canavanine utilizing insects;Canavalia ensiformis, a canavanine storing plant; and to a lesser extentHeliothis virescens, a canavanine resistant insect, failed to accumulate significant canavanyl proteins. By contrast,Manduca sexta, a canavanine-sensitive insect, andGlycine max, a canavanine free plant, readily incorporated canavanine into newly synthesized proteins. This study supports the contention that the incorporation of canavanine into proteins in place of arginine contributes significantly to canavanine's antimetabolic properties.     

Insect tissues,not microorganisms,produce linoleic acid in the house cricket and the American cockroach

Summary Biosynthesis of linoleic acid, 182(n–6), was unambiguously demonstrated to occur in the cockroach,Periplaneta americana, and the cricket,Acheta domesticus. Axenic tissue from both of these insect species was demonstrated by radio-gas-liquid chromatography (radio-GLC) and radio-high-performance liquid chromatography (radio-HPLC) to incorporate [1-¹⁴C]acetate and [1-¹⁴C]oleate into this essential fatty acid.This work was supported by the National Science Foundation under grant DCB-8914417. We would like to thank Coby Schal for his generous gift of American cockroaches and Tania Kellermeyer for her excellent technical assistance.     

The role of glycine in the biosynthesis of steroids

Zusammenfassung Während der Biosynthese von Cholesterol mit homogenisierter Rattenleber wird [2-¹⁴C] des Glycins viel besser eingebaut als [1-¹⁴C].Saccharomyces cerevisiae produziert radioaktives Squelen (ausser Ergosterol mit Radioaktivität des Ringsystems) mit [2-¹⁴C] Glycin und mit [3-¹⁴C] Serin, aber nicht mit [1-¹⁴C] Glycin.

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Studies on Biosynthesis. Part VI. For Part V, seeA. K. Bose, K. S. Khanchandani andB. L. Hungund, Experientia,27, 1403 (1971). b) Presented at the 164th National Meeting of the American Chemical Society, New York, August, 1972.

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The support of this research by Stevens Institute of Technology and Sandoz Foundation is gratefully acknowledged. We wish to thank Drs.P. T. Funke, M. S. Manhas, P. K. Bhattacharyya, M. Anchel andH. Levey for valuable discussions and help with some of the experiments.     

Zur Biosynthese der C9–10-Einheit der Indolalkaloide

Summary After administration of various C₂-compounds as well as leucine-2-¹⁴C toCatharanthus roseus shoots and glycine-2-¹⁴C toStrychnos nux vomica plants no specific incorporation into the non-tryptophan C_9–10 moiety of indole alkaloids was observed. The results indicate that glycine-2-¹⁴C is transformed into serine and is incorporated via tryptophan into strychnine.     

Chemical defense in the three European species ofCrematogaster ants

The composition of the Dufour gland of the antC. scutellaris has been reinvestigated by gas chromatography/mass spectrometry. The major components of the gland are (2E,5E,12Z)-4-oxoheneicosa-2,5,12-trien-1-ol acetate (1a) its ¹⁴ and ¹⁶ double bond isomers (1b and1c), and the corresponding (Z,Z)-dienes5a and5b, all containing an acetylated C₂₁ chain. The previously proposed structures1d, 1e, and5c, which are based on an homologous acetylated C₂₃ chain, correspond to minor derivatives present in the gland. Traces of acetylated C₁₉ homologs, tentatively identified as1g-1i, have also been found. The Dufour gland contents of the two other EuropeanCrematogaster species have also been studied.C. auberti is very similar toC. scutellaris in producing mainly1a, 1b and1c, together with the same higher and lower homologs, but it lacks the dienic derivatives5, whereasC. sordidula contains essentially the acetylated C₁₉ compounds1g, 1h, and1i, accompanied by acetylated C₁₇ homologs.     

The role of glycine in the biosynthesis of a terpene

Zusammenfassung Mit doppelt markiertem Glycin konnte gezeigt werden, dass das Kohlenstoffatom der Methylgruppe viel besser als das Kohlenstoffatom der Karboxylgruppe in die Isoprengruppe des Ophiobolins B ausCochiobolus miyabeanus eingebaut wird. Die Radioaktivität wird in Ophiobolin B eingebaut, wenn die folgenden Verbindungen verwendet werden: [3-¹⁴C]-Serin, [¹⁴C-Methyl]-Sarcosin, [¹⁴C-Methyl]-Methionin, und [¹⁴C] Formate.

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a)Studies on Biosynthesis. Part V. for Part IV, seeM. Anchel, A. K. Bose, K. S. Khanchandani andP. T. Funke, Phytochem.9, 2135 (1970). b) Presented in part at the 3rd Natural Products Symposium, University of West Indies, Jamaica, January 1970.

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The support of this research by Stevens Institute of Technology is gratefully acknowledged. We wish to thank Dr.J. H. Davis and Prof.L. Z. Pollara for their interest and encouragement and Drs.H. Levy, P. T. Funke, M. S. Manhas, P. K. Bhattacharyya andM. Anchel for valuable discussions and help with some of the experiments. We are particularly thankful to Dr.L. Canonica for providing us with cultures ofCochiobolus miyabeanus that made our work on ophiobolin B possible.     

Metabolism of 1,3,7-trimethyldihydrouric acid in the rat: New metabolic pathway of caffeine

Summary [1-CH₃-¹⁴C] 1,3,7-trimethyldihydrouric acid which, in quantity, is the most important caffeine metabolite, was isolated and purified from the urine of rats fed with [1-CH₃-¹⁴C] caffeine. The oral administration of this metabolite to rats showed that 1,3,7-trimethyldihydrouric acid was excreted unchanged in urine and was therefore an end product of caffeine metabolism. This result implies a new metabolic pathway of caffeine.     

Isomériecis-trans des ménaquinones et oxydations phosphorylantes dans les extraits deMycobacterium phlei

Summary Incubation ofM. phlei washed cells with [¹⁴C³H₃]-l-methionine led to [2-¹⁴C³H₃] dihydromena-quinone-9 with an isotope ratio identical to that of methionine. Chromatography of the doubly labelled quinone indicated, despite a pronounced isotope effect, that bothcis andtrans isomers had the same isotope ratio. This result eliminates any possibility of hydrogen exchange in the 2-methyl group of menaquinones during oxydative phosphorylation, even in thecis isomer. Furthermore, it is confirmed that this compound is certainly formed from natural or synthetic menaquinones during isolation or incubation periods by the effect of daylight irradiation.     

The sterols of the khapra beetle,Trogoderma granarium everts

Summary The khapra beetle,Trogoderma granarium Everts, does not dealkylate and convert dietary C₂₈- or C₂₉-phytostorols to C _2T sterols such as cholesterol. There is, however, an increase in the concentration of cholesterol and campesterol in its tissues relative to the dietary concentrations of these sterols, presumably as a result of selective uptake.We thank Dr S. R. Dutky of the Insect Physiology Laboratory, Agricultural Research, SEA, USDA, Beltsville, Maryland for providing GC-MS analyses.     

A reinvestigation of the chemistry of the Dufours gland of the formicine ant,Anoplolepis custodiens

The components of individual Dufours glands excised fromAnoplolepis custodiens workers were analysed by GC-MS. In addition to then-alkanes andn-alkenes previously reported² in these glands, primary alcohols (C₁₉-C₂₂), secondary alcohols (C₂₀-C₂₃), 2-ketones (C₂₀-C₂₃) and possibly carboxylate ethyl esters (C₁₉ and C₂₁) were identified as components of these glands. It seems possible that these high-boiling compounds are used by the workers in laying trails on the hot sandy surfaces of their characteristic habitat and in lining of the inner walls of nests, but no standard compounds have been available to us for any behavioral studies.     

Metabolism in Porifera. IX. Studies on the biological conversion of cholesterol into 19-nor-cholestanol by the spongeAxinella polypoides

Summary The conversion of cholesterol into 19-nor-5-cholestan-3-ol by the spongeAxinella polypoides involves a partial loss (40%) of the 3-hydrogen atom; moreover administration to the sponge of [4-¹⁴C]cholesterol tritiated at C-4 and C-7 showed that the 4- and 7-hydrogen atoms are retained in this conversion. A competitive uptake experiment, [4-¹⁴C]cholesterol vs. [7-³H₂]5-cholestanol, showed that the sponge utilized exclusively cholesterol for the production of 19-nor-5-cholestan-3-ol.Part of this work has been presented at the Nato Conference on Marine Natural Products (Jersey, Great Britain, October, 1976).-This contribution is part of the Programma finalizzato Oceanografia e Fondi marini-sottoprogetto Risorse biologiche C.N.R. Italy.     

Inhibition of cholesterol synthesis ex vivo and in vivo by fluvastatin,a new inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase

The inhibitory effect of fluvastatin sodium (fluvastatin), a new type of 3-hydroxy-3-methylglutaryl (HMG) coenzyme A inhibitor, on de novo cholesterol synthesis was investigated and compared with that of pravastatin. Fluvastatin at a concentration of 12.5 mg/kg inhibited sterol synthesis ex vivo from [¹⁴C]acetate in rat liver and ileum by 97–99% with respect to the control, while the inhibition in kidney was 55%. The inhibition by fluvastatin in the liver and ileum persisted for approximately 9 h after administration. Significant differences between fluvastatin also had an inhibitory effect on cholesterol synthesis in vivo in various tissues of rats given [¹⁴C]acetate intraperitoneally. Sterol synthesis in the liver, ileum and kidney was inhibited by over 95% 3 h after administration of 6.25 mg/kg of fluvastatin. Significant differences between fluvastatin and pravastatin were found in the liver and ileum. Fluvastatin was more potent than pravastatin in inhibiting both ex vivo and in vivo sterol synthesis in the ileum (but not in kidney) and liver.     

Production of insulin-like growth factor I (IGF-I) and IGF-binding proteins by rat intestinal stromal cells in vitro

To determine if intestinal stromal cells secrete diffusible factors such as insulin-like growth factors (IGFs) capable of regulating epithelial cell growth in vitro, stromal cells were isolated by enzymatic digestion of rat intestine. Incorporation of [³H]thymidine into DNA and [¹⁴C]leucine into protein of IEC-6 cells, a model intestinal epithelial cell line, was significantly increased (two- to threefold) when the IEC-6 cells were co-cultured with stromal cells, relative to IEC-6 cells grown alone. Medium conditioned by stromal cells stimulated DNA synthesis of IEC-6 cells in a dose-dependent manner. Analysis of the conditioned medium revealed that intestinal stromal cells secreted IGF-I, but little IGF-II, in addition to an M _r 32,000 IGF-binding protein (IGFBP-2) and an IGFBP having M _r∼ 24,000. We conclude that rat intestinal stromal cells secrete one or more diffusible factors, which may include IGF-I and IGFBPs, capable of stimulating proliferation of IEC-6 cells in vitro. Received 25 August 1997; received after revision 7 November 1997; accepted 20 November 1997     

Autotrophic incorporation of C14O2 inCuscuta australis in relation to its parasitism

Riassunto L'incorporazione della C¹⁴O₂ inC. australis appare chiaramente di tipo fotosintetico in qualunque stadio di vita della pianta (plantula, fusto, infiorescenza). Fusti ed infiorescenze appaiono dotati di un metabolismo più veloce rispetto alle plantule.     

The C4-dicarboxylate transport system ofRhizobium meliloti and its role in nitrogen fixation during symbiosis with alfalfa (Medicago sativa)

TheRhizobium meliloti C₄-dicarboxylate transport (Dct) system is essential for an effective symbiosis with alfalfa plants. C₄-dicarboxylates are the major carbon source taken up by bacteroids. Genetic analysis of Dct^– mutant strains led to the isolation of thedct carrier genedctA and the regulatory genesdctB anddctD. The carrier genedctA is regulated in free-living cells by the alternative sigma factor RpoN and the two-component regulatory system DctB/D. In addition, DctA is involved in its own regulation, possibly by interacting with DctB. In bacteroids, besides the DctB/DctD system an additional symbiotic activator is thought to be involved indctA expression. Further regulation ofdctA in the free-living state is reflected by diauxic growth of rhizobia, with succinate being the preferred carbon source. The tight coupling of C₄-dicarboxylate transport and nitrogen fixation is revealed by a reduced level of C₄-dicarboxylate transport in nitrogenase negative bacteroids.     

Strong mutagenic action of a bipyridylium herbicide in a N2-fixing blue-green alga

Summary The herbicide paraquat (1,1-dimethyl-4,4-bipyridylium ion) has been found to exert a growth inhibitory effect on the N₂-fixing blue-green algaNostoc muscorum in nitrogen-free (N₂) and NO ₃ ^– media, without any apparent inhibitory or stimulatory effect on the heterocyst-forming capacity of the organism. With a dose of paraquat permitting about 20 and 50% survival of this alga a reverse mutation (fromhet ⁺ nif ^– auxotrophy tohet ⁺ nif ⁺ prototrophy), a forward mutation (for L-methionine-DL-sulfoximine [MSO]-resistance), and an auxotrophic mutation (for carbon-auxotrophy through methylamine [MA]-resistance) have been obtained. The toxic and mutagenic effects of this agrochemical have been compared with those of the well known mutagen MNNG (N-methyl-N-nitro-N-nitrosoguanidine) and found to be stronger than those of the latter in each case.The author wishes to thank Mr Girish Nath, Lakshami Chemicals Pvt. Ltd., Phatuha (Patna), India, for providing pure samples of the herbicide for the present investigation. The receipt of financial assistance for this work in the form of a Higher Research Associateship Award to the author from the University Grants Commission, Govt. of India, New Delhi-110002 (Grant No. F/16-51/83), is thankfully acknowledged.     

Dominance for enzyme activity inDrosophila melanogaster

Summary A regulatory element tightly linked to theGpdh locus inDrosophila melanogaster has been isolated from a natural population. Flies homozygous for second chromosomes bearing the element,H31, have half the GPDH activity of normal homozygotes. Heterozygotes betweenH31 andF orS alleles exhibit dominance in GPDH activity. Heterozygotes betweenH31, F orS andDf(2L) GdhA have half the diploid level. The contribution of theS allele to the activity inS/H31 heterozygotes is more than four times that ofH31. The regulatory element distinguishingH31 is tightly linked to theGpdh ⁺ locus.     

Serotonin-stimulated protein phosphorylation in aortic smooth muscle cells

Summary The effects of serotonin on the formation of inositol phosphates and protein phosphorylation were examined in cultured smooth muscle cells. Serotonin stimulated the formation of [³H]inositol monophosphate, [³H]inositol bisphosphate and [³H]inositol trisphosphate. This effect was prevented by 5-HT₂ specific antagonist, 6-methyl-1-(1-methylethyl)ergoline-8-carboxylic acid, 2-hydroxy-1-methylpropyl ester [Z]-2-butenedioate (LY53857). Serotonin stimulated the phosphorylation of many polypeptides, among which a 20 kDa polypeptide was the most prominent. The phosphorylation was also inhibited by LY53857. LY53857 alone produced no effects on protein phosphorylation. The 20 kDa polypeptides were also phosphorylated by the addition of 12-O-tetradecanoylphorbol-13-acetate. These results suggest that serotonin stimulates protein phosphorylation through 5-HT₂ receptors and possibly activates protein kinase C in intact vascular smooth muscle cells.Part of the data contained in this paper was presented at the 74th local meeting of the Japanese Society of Pharmacology at Kanagawa.