Incorporation of [14C]glucosamine and [14C]leucine into mouse kidney β-glucuronidase induced by gonadotrophin

Male BALB/C mice were injected intraperitoneally with 2.5 i.u. of gonadotrophin. After the injection, increase of β-glucuronidase activity was first observed in the microsomal fraction. By 36h 45–50% of the total homogenate activity was found in the microsomal fraction compared with 20–25% in the control microsomal fraction. From 36 to 80h not only microsomal β-glucuronidase but also lysosomal β-glucuronidase increased progressively. After 69h stimulation with 2.5 i.u. of gonadotrophin, d-[1-¹⁴C]glucosamine or l-[U-¹⁴C]leucine was injected intraperitoneally. After a further 3h the kidneys were homogenized and five particulate fractions were prepared by differential centrifugation. The β-glucuronidase in the microsomal and lysosomal fractions was released respectively by ultrasonication and by freezing and thawing treatment. The enzyme was purified by organic-solvent precipitation and by sucrose-density-gradient centrifugation. The results demonstrated the incorporation of these two labels into the mouse renal β-glucuronidase. The microsomal β-glucuronidase was much more radioactive than the lysosomal enzyme and approx. 80% of the newly synthesized enzyme appeared in microsomes and approx. 20% of that was found in lysosomes at this period. These results suggest that the mouse renal β-glucuronidase is a glycoprotein and that the newly synthesized enzyme is transported from endoplasmic reticulum to lysosomes.     

Assay of Δ1-piperideine-2-carboxylate and synthesis of l-[14C]pipecolate from dl-[14C]pipecolate

Four assay methods were tested for the measurement of Δ¹-piperideine-2-carboxylate, a proposed alicyclic ketimino acid intermediate in the pathway of lysine metabolism to l-pipecolate, and the product of d-amino acid oxidase on d-pipecolate. The method using Δ¹-piperideine-2-carboxylate reductase from Pseudomonas putida was found to be most sensitive and specific. Measurement of Δ¹-piperideine-2-carboxylate by reduction with NaBH₄ and ninhydrin assay of the resultant pipecolate, by direct acidic ninhydrin assay, and by o-aminobenz-aldehyde assay were less desirable because of lower sensitivity and specificity. Two synthetic methods for preparing l-[¹⁴C]pipecolate from the racemic dl-[¹⁴C]pipecolate were investigated. Incubation of dl-[¹⁴C]pipecolate with a combination of d-amino acid oxidase and Δ¹-piperideine-2-carboxylate reductase or d-amino acid oxidase and NaBH₄ totally inverted the d-isomer to the l-isomer, with $\text{Δ}{}^1\text{-[}{}^{14}\text{C]piperideine-2-carboxylate}$ as an intermediate in each cycle of interconversion. No purification except desalting through a Dowex 50 (H⁺) column was necessary in order to recover l-[¹⁴C]pipecolate in pure form. The yield was 95–97% compared to <50% in the conventional method.     

A procedure for the synthesis of p-fluorosulfonyl[14C]-benzoyl-5′-adenosine with [14C] in the benzoyl moiety

Carboxy-p-fluorosulfonyl[¹⁴C]benzoyl-5′-adenosine has been synthesized with the radiolabel ultimately derived from carboxy-p-amino[¹⁴C]benzoic acid by a synthetic route employing four reaction steps. Starting with 1 mmol of p-amino[¹⁴C]benzoic acid, p-fluorosulfonyl[¹⁴C]benzoyl-5′-adenosine is obtained with an overall yield of 25–30%.     

双链核糖核酸免疫化学研究 Ⅰ.双链多聚[I]:多聚[C]的抗原性

用国产多聚肌苷酸和多聚胞嘧啶核苷酸(简称多聚[I]:多聚[C])制备成双链核糖核酸特异性抗血清,用环状沉淀和~3H标记的番茄花叶病毒双链核糖核酸测得抗血清效价分别为1:128和1:6400。用免疫琼脂双扩散、对流免疫电泳和放射免疫测定可测定出多聚[I]:多聚[C]的最低浓度分别为391ng、12.2/ng和10pg/ml抗血清和酵母、TMV、PVX 的单链核糖核酸、小牛胸腺DNA不反应。试验证明用国产多聚[I]:多聚[C]制备的抗血清,可有效地用干双链核糖核酸的免疫化学鉴定。由于在免疫双扩散和对流免疫电泳中抗血清能和微量双链核糖核酸反应并产生可见沉淀线,从而有可能利用这两种简便灵敏的方法测定病毒的双链RNA,单链RNA病毒的RF型RNA,以及双链RNA真菌病毒的筛选。     

Incorporation of label from d-β-hydroxy-[14C]butyrate and [3-14C]acetoacetate into amino acids in rat brain in vivo

The metabolism of ketone bodies by rat brain was studied in vivo. Rats starved for 48h were given either d-beta-hydroxy[3-(14)C]butyrate or [3-(14)C]acetoacetate by intravenous injection and killed after 3 or 10min. Total radioactivity in the acid-soluble material of the brain and the specific radioactivities of the brain amino acids glutamate, glutamine, aspartate and gamma-aminobutyrate were determined. A group of fed animals were also given d-beta-hydroxy[3-(14)C]butyrate. In the brains of all animals (14)C was present in the acid-soluble material and the specific radioactivity of glutamate was greater than that of glutamine.     

Interaction of α-cyano[14C]cinnamate with the mitochondrial pyruvate translocator

The binding of α-cyanocinnamate to rat-heart mitochondrial membrane was investigated using α-cyano[¹⁴C]cinnamate. The binding was correlated to the inhibition of pyruvate transport. The results obtained demonstrate that both these functions reach saturation at the same titre of the inhibitor. Quantitative parameters of α-cyano[¹⁴C]cinnamate binding have been determined. The binding can be prevented by pyruvate and other substrates of the carrier but not by acetate. Pyruvate decreases the affinity of α-cyanocinnamate binding, leaving the maximum number of binding unchanged. It is concluded that rat-heart mitochondria contain a specific site at which α-cyanocinnamate binds which is directly involved in the inhibition of pyruvate transport.     

Transport und Stoffwechsel von 2-[14C]Abscisinsäure in Wurzelsegmenten von Phaseolus coccineus L.

Summary Movement of 2-[¹⁴C]ABA through 1.5 cm and 5 cm long root segments of P. coccineus L. was acropetally polarised. The velocity of acropetal transport of [¹⁴C]ABA in 1.5 cm long segments was 4–5 mm·h^-1. Up to 11 h after the start of incubation [¹⁴C]ABA could be extracted unchanged. Beyond this time radioactivity became associated to an unidentified compound, which shows chromatographic qualities similar to those of Milborrow's Metabolite C (Biochemistry & Physiol. Plant Growth Substances, 1531, Runge Press Ottawa, 1968; and Chem. Comm. 966, 1969).

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Abkürzungen ABA Abscisinsäure - PPO 2,5-Diphenyloxazol     

Differential effects of veratridine and calcium on the release of [3H]noradrenaline and [14C]α-aminoisobutyrate from rat brain cortex slices

The efflux of [³H]noradrenaline (NA) and of the non transmitter, non metabolizable, amino acid [¹⁴C]α-aminoisobutyrate (AIB), was followed simultaneously from superfused rat brain cortex thin slices, that had been preloaded with those substances. Short (2 min) “pulses” of increasing veratridine concentrations were applied at 10 min intervals. When calcium in the superfusion fluid was 1 mM, [³H]NA efflux increased progressively with pulses of 1, 3, 10 and 30 μM veratridine, but further increase to 100 μM resulted in a decrease of the induced ³H-efflux. Veratridine-enhanced [³H]NA efflux decreased considerably in 0.1 mM calcium and was virtually suppressed when no calcium was added to the superfusion fluid. In 1 mM calcium, the efflux of [¹⁴C] AIB was increased progressively by pulses of 10, 30 and 100 μM veratridine, but no increase in efflux was seen with 1 or 3 μM drug. In 0.1 mM, or without added calcium, the induced efflux of [¹⁴C]AIB was markedly increased. Similar findings were seen when a long (10 min) pulse of 10 μM veratridine was given. After such long pulses there was a rapid return of AIB efflux to pre-veratridine levels if calcium was 1 mM, but in the absence of added calcium, the return to baseline levels of both [³H]NA and, especially, that of [¹⁴C]AIB efflux, was greatly impaired. The veratridine enhanced efflux of both NA and AIB was entirely blocked by 1 μM tetrodotoxin.     

Use of [4,6-Di-14C]-5′-DMT-thymidine Phosphoramidite Reagent for the Radiolabeling of Synthetic Oligonucleotides

Abstract

A novel synthesis of 5′-radiolabeled oligonucleotides is described. The labeling is carried out by the phosphoramidite method with the aid of building block 1. The feasibility of the method is demonstrated by preparation of 5′-radiolabeled 3′-phosphorylated dodecathymidylate phosphorothioate.     

Decarboxylative metabolism of [1′-14C]indole-3-acetic acid by tomato pericarp discs during ripening

The rate of decarboxylation of [1′-¹⁴C]indole-3-acetic acid (IAA) infiltrated into tomato (Lycopersicon esculentum Mill.) pericarp discs was much more rapid in green than in breaker and pink tissues. Studies were carried out in order to determine whether the decarboxylative catabolism occurring in the green pericarp discs was associated with ripening or was a consequence of wound-induced peroxidase activity and/or ethylene production. After a 2-h lag, the decarboxylative capacity of the green pericarp discs increased exponentially during a 24-h incubation period. This increase was accompanied by increases in IAA-oxidase activity in cell-free preparations from the intercellular space and cut surface of the discs. Although higher IAA-oxidase activity was detected in extracts from the tissue residue, which comprises mainly intracellular peroxidases, this activity did not increase during the 24-h incubation period. Analysis of the cell-free preparations by isoelectric focusing revealed the major component in all samples was a highly anionic peroxidase (pI=3.5) the levels of which did not increase during incubation. However, the intercellular and cut-surface preparations contained additional anionic and cationic peroxidases which increased in parallel with the increases in both the IAA-oxidase activity of the preparations and the decarboxylative capacity of the green pericarp discs from which they were derived. Treatment of green discs with the ethylene-biosynthesis inhibitors aminooxyacetic acid and CoCl₂, inhibited the development of an enhanced capacity to decarboxylate [1′-¹⁴C]IAA but the inhibition was not counteracted by exogenous ethylene. Another ethylene-biosynthesis inhibitor, aminoethoxyvinyl glycine, also reduced ethylene levels but did not affect IAA decarboxylation, indicating that the decarboxylation was not a consequence of wound-induced ethylene production. The data obtained thus demonstrate that the enhanced capacity to decarboxylate [1′-¹⁴C]IAA that develops in green tomato pericarp discs following excision is not associated with ripening but instead is attributable to a wound-induced increase in anionic and cationic peroxidase activity in the intercellular fluid and at the cut surface of the excised tissues.     

Virulence de l'entomophthoraleZoophthora radicans [zygomycetes] à l'égard des adultes deAedes aegypti [Dipt.: Culicidae]

Résumé Les auteurs ont mis au point un procédé d'estimation de la virulence des Entomophthorales à l'égard des adultes du moustiqueAedes aegypti. Ce paramètre est estimé en termes de CL 50 de conidies/mm². C'est la 1^re fois qu'une telle méthodologie est appliquée à ces champignons vis-à-vis de Diptères. L'activité de 2 souches deZoophthora radicans isolées le même jour dans la même localité a ainsi pu être comparée à des intervalles successifs après la mise en culture. Au début de l'expérience, les 2 souches se sont révélées capables d'entra?ner des taux de mortalité de 100% et leur virulence était comparable. La symptomatologie de la mycose était caractéristique, avec formation de conidiophores chez la quasitotalité des cadavres. Par la suite, une discordance a été notée dans le comportement respectif des 2 souches. Alors que la virulence de l'une est restée stable pendant les 6 mois qu'a duré l'expérience, celle de l'autre a diminué très rapidement, le taux maximum de mycose ne dépassant plus 20% après le 1^er essai. Le passage sur l'h?te expérimental n'a pas eu pour effet de modifier cette situation. Ces résultats, ainsi que leurs implications, sont discutés. Ils soulignent l'importante variabilité comportementale qui peut exister au sein d'une même espèce d'Entomophthorale. Avec la collaboration technique deMonique Arnault et deNelly Delatre.     

In Memorium: Herman P. Schwan [1915–2005]

Herman P. Schwan [1915–2005] was a distinguished scientist and engineer, and a founding father of the field of biomedical engineering. A man of integrity, Schwan influenced the lives of many, including his wife and children, and his many students and colleagues. Active in science until nearly the end of his life, he will be very much missed by his family and many colleagues.     

Les hôtes dePseudoperichaeta nigrolineata et deP. palesoidea [DIPT.: Tachinidae]

Résumé A partir des éléments publiés et de données inédites, les auteurs dressent la liste des Lépidoptères h?tes desTachinidae du genrePseudoperichaeta dans la région paléarctique. Aucun h?te deP. erebiae n'est connu. PourP. nigrolineata etP. palesoidea, 6 nouveaux h?tes sont signalés, respectivementHomœosoma nebulellum (Denis & Schiffermüller),Lobesia botrana (Denis & Schiffermüller) etCnephasia alticolana Herrich-Sch?ffer pour la première;Peribatodes rhomboidaria (Denis & Schiffermüller),Choristoneura murinana (Hübner) etTortrix viridana Linné pour la seconde. Quelques éléments biologiques et géographiques sur ces nouveaux h?tes sont mentionnés.      

Bioecologie deEubazus semirugosus,coeloïdes abdominalis etC. sordidator [Hym.: Braconidae] parasites dePissodes notatus [Col.: Curculionidae] dans le sud de la France

Résumé Cette étude porte sur différents aspects de la biologie deE. semirugosus, C. sordidator etC. abdominalis, se développant surP. notatus, ravageur du Pin maritime: comportement parasitaire à l'égard de ce charan?on, durée du développement, durée de la maturation sexuelle, espérance de vie des adultes, etc. Le cycle biologique de ces parasito?des est étudié ainsi que le problème de la co?ncidence temporelle entre ces braconides et leur h?te dans le cas d'une forêt du sud de la France.      

生物工程技术讲座[八]

应用微细胞检测基因产物的方法由于质粒和噬菌体等的DAN分子能在细胞内进行自我复制,以及DNA重组技术的普及,质粒DNA做为异源性DNA分子的载体,在基因工程中目前已被广泛应用。在克隆或分析真核生物或原核生物的多种不同基因时,常用检测基因表达来确定其存在。把含有某一目的基因的DNA片段和载体质粒的DNA进行重组,再通过重组DNA在受体细胞中的转录、翻译、     

Platystasius transversus [Hym.: Platygasteridae] parasitoïde oophage dePhoracantha semipunctata [Col.: Cerambycidae], au Maroc

Résumé Platystasius transversus (Thomson) parasite les œufs du CérambycidePhoracantha semipunctata (F.), près de Rabat. C'est la 1^re mention, pour l'aire géographique, d'un ennemi naturel de ce déprédateur d'eucalyptus, originaire d'Australie, et récemment signalé au Maroc.      

Incorporation of label from [1-14C]ethanol into the glutamate–glutamine pools of rat brain in vivo

Glutamate and glutamine were isolated from the brains of rats given [1-(14)C]ethanol by three different routes of injection with and without carrier ethanol or acetate. Without exception, the specific radioactivity of brain glutamine was 3- to 5-fold that of brain glutamate. The route of injection had no consistent effect on the results. These results indicate that small amounts of ethanol are oxidized by rat brain in the compartment responsible for the so-called ;small metabolic pool' of brain glutamate.     

Incorporation of [3H] uridine into the ribonucleic acid of rat uterus during pseudopregnancy and in the presence of I.C.I. 46474 [trans-1-(p-β-dimethylaminoethoxyphenyl)-1,2-diphenylbut-1 -ene]

1. The incorporation of [(3)H]uridine into RNA of rat uterine tissue has been measured during pseudopregnancy and in rats receiving different doses of an anti-implantation compound [trans-1-(p-beta-dimethylaminoethoxyphenyl)-1,2-diphenylbut- 1-ene, I.C.I. 46474]. 2. In the uterus of the pseudopregnant rat uridine incorporation into RNA increased markedly on day 3 of pseudopregnancy, remained high until day 5 and decreased sharply by day 6, rising again by day 9. 3. This pattern of change was similar to, but not identical with, that previously found in the pregnant rat. 4. Rats receiving I.C.I. 46474 at a dose concentration below that preventing implantation showed a pattern of RNA synthesis similar to that found in untreated control rats. 5. Rats treated with doses of I.C.I. 46474 sufficient to inhibit implantation revealed a totally different pattern of incorporation of [(3)H]uridine into uterine RNA. 6. The results are discussed in terms of previous findings with the normally pregnant rat. It is concluded that the increasing uterine RNA synthesis found on day 3 in the pregnant rat is an important requirement for the occurrence of the subsequent implantation.