Hepatitis C virus infection in a Brazilian population with sickle-cell anemia

Patients with sickle-cell anemia submitted to frequent blood transfusions are at risk of contamination with hepatitis C virus (HCV). Determination of HCV RNA and genotype characterization are parameters that are relevant for the treatment of the viral infection. The objective of the present study was to determine the frequency of HCV infection and the positivity for HCV RNA and to identify the HCV genotype in patients with sickle-cell anemia with a history of blood transfusion who had been treated at the Hospital of the HEMOPE Foundation. Sera from 291 patients were tested for anti-HCV antibodies by ELISA 3.0 and RIBA 3.0 Chiron and for the presence of HCV RNA by RT-PCR. HCV genotyping was performed in 19 serum samples. Forty-one of 291 patients (14.1%) were anti-HCV positive by ELISA and RIBA. Both univariate and multivariate analysis showed a greater risk of anti-HCV positivity in those who had started a transfusion regime before 1992 and received more than 10 units of blood. Thirty-four of the anti-HCV-positive patients (34/41, 82.9%) were also HCV RNA positive. Univariate analysis, used to compare HCV RNA-negative and -positive patients, did not indicate a higher risk of HCV RNA positivity for any of the variables evaluated. The genotypes identified were 1b (63%), 1a (21%) and 3a (16%). A high prevalence of HCV infection was observed in our patients with sickle-cell anemia (14.1%) compared to the population in general (3%). In the literature, the frequency of HCV infection in sickle-cell anemia ranges from 2 to 30%. The serological screening for anti-HCV at blood banks after 1992 has contributed to a better control of the dissemination of HCV infection. Because of the predominance of genotype 1, these patients belong to a group requiring special treatment, with a probable indication of new therapeutic options against HCV.

Hepatitis C virus seroprevalence and genotypes in patients with diffuse connective tissue diseases and spondyloarthropathies

Many extrahepatic manifestations, including rheumatic diseases, have been reported to be associated with hepatitis C virus (HCV) infection. In order to investigate the prevalence of HCV infection among patients with rheumatic diseases, in the present study we interviewed 367 patients and tested their blood samples for HCV antibodies (anti-HCV) by an enzyme-linked immunosorbent assay. Anti-HCV-reactive samples were retested for confirmation by a line immunoassay and also for HCV RNA detection by the polymerase chain reaction. HCV RNA-positive samples were genotyped by INNO-LIPA. An overall HCV infection prevalence of 1.9% (7/367) was found. Of the 7 HCV-infected patients, 4 had systemic lupus erythematosus and 3 rheumatoid arthritis, resulting in positivity rates of 2.3 and 3.4%, respectively. HCV RNA genotyping revealed the presence of subtypes 1a (57.1%), 1b (28.6%) and 3a (14.3%). The clinical course was favorable for all HCV-infected patients, except one, who died due to renal insufficiency related to lupus nephritis. These results demonstrate a low HCV infection prevalence among the population studied. In the few positive cases, we observed no adverse influence of this infection on the clinical evolution of the rheumatic disease.

Pattern of Wnt ligand expression during chick eye development

The dorsoventral axis of the eye is determined prior to optic cup invagination. A variety of signaling pathways have been implicated in the maintenance of the optic dorsoventral axis, including, but not limited to, bone morphogenetic protein 4, Sonic Hedgehog and retinoic acid. Here, we investigated the possible contribution of Wnt ligands to the establishment or maintenance of the optic axis by analyzing their expression pattern during early chick optic development. We performed in situ hybridization of Wnt-1, Wnt-3a, Wnt-4, and Wnt-5a during the optic vesicle, early optic cup and established optic cup stages and focused our analysis on the optic region. Our data showed that Wnt-5a, but none of the others, is expressed in the dorsal region of the eye starting from the Hamburger and Hamilton stage 14 (HH14). These results are supported by cryosections of the labeled optic region, which further reveal that Wnt-5a is expressed only in the dorsal retinal pigmented epithelium. Thus, we propose that Wnt-5a is a marker for dorsal retinal pigmented epithelium in chick embryos from HH14 to HH19.

Cytotoxic and DNA-topoisomerase effects of lapachol amine derivatives and interactions with DNA

The cytotoxic activity of amino (3a-e), aza-1-antraquinone (4a-e) lapachol derivatives against Ehrlich carcinoma and human K562 leukemia cells was investigated. Cell viability was determined using MTT assay, after 48 (Ehrlich) or 96 h (K562) of culture, and vincristine (for K562 leukemia) and quercetin (for Ehrlich carcinoma) were used as positive controls. The results showed dose-dependent growth-inhibiting activities and that the amino derivatives were active against the assayed cells, whereas the 4a-e derivatives were not. The allylamine derivative 3a was the most active against Ehrlich carcinoma, with IC50 = 16.94 ± 1.25 µM, and against K562 leukemia, with IC50 = 14.11 ± 1.39 µM. The analogous lawsone derivative, 5a, was also active against Ehrlich carcinoma (IC50 = 23.89 ± 2.3 µM), although the 5d and 5e derivatives showed lower activity. The interaction between 3a-d and calf thymus DNA was investigated by fluorimetric titration and the results showed a hyperchromic effect indicating binding to DNA as presented of ethidium bromide, used as positive control. The inhibitory action on DNA-topoisomerase II-a was also evaluated by a relaxation assay of supercoiled DNA plasmid, and the etoposide (200 µM) was used as positive control. Significant inhibitory activities were observed for 3a-d at 200 µM and a partial inhibitory action was observed for lapachol and methoxylapachol.

Genotyping hepatitis C virus from hemodialysis patients in Central Brazil by line probe assay and sequence analysis

The present study examined the distribution of hepatitis C virus (HCV) genotypes and subtypes in a hemodialysis population in Goiás State, Central Brazil, and evaluated the efficiency of two genotyping methods: line probe assay (LiPA) based on the 5' noncoding region and nucleotide sequencing of the nonstructural 5B (NS5B) region of the genome. A total of 1095 sera were tested for HCV RNA by RT-nested PCR of the 5' noncoding region. The LiPA assay was able to genotype all 131 HCV RNA-positive samples. Genotypes 1 (92.4%) and 3 (7.6%) were found. Subtype 1a (65.7%) was the most prevalent, followed by subtypes 1b (26.7%) and 3a (7.6%). Direct nucleotide sequencing of 340 bp from the NS5B region was performed in 106 samples. The phylogenetic tree showed that 98 sequences (92.4%) were classified as genotype 1, subtypes 1a (72.6%) and 1b (19.8%), and 8 sequences (7.6%) as subtype 3a. The two genotyping methods gave concordant results within HCV genotypes and subtypes in 100 and 96.2% of cases, respectively. Only four samples presented discrepant results, with LiPA not distinguishing subtypes 1a and 1b. Therefore, HCV genotype 1 (subtype 1a) is predominant in hemodialysis patients in Central Brazil. By using sequence analysis of the NS5B region as a reference standard method for HCV genotyping, we found that LiPA was efficient at the genotype level, although some discrepant results were observed at the subtype level (sensitivity of 96.1% for subtype 1a and 95.2% for subtype 1b). Thus, analysis of the NS5B region permitted better discrimination between HCV subtypes, as required in epidemiological investigations.

Chronicon episcoporum Finlandensium 24

Arkit: 1 arkintunnukseton lehti, 3A-3B4.

Lateral asymmetry of voluntary attention orienting

We recently demonstrated that automatic attention favors the right side of space and, in the present study, we investigated whether voluntary attention also favors this side. Six reaction time experiments were conducted. In each experiment, 12 new 18-25-year-old male right-handed individuals were tested. In Experiments 1, 2, 3 (a, b) and 4 (a, b), tasks with increasing attentional demands were used. In Experiments 1, 2, 3a, and 4a, attention was oriented to one or both sides by means of a central spatially informative visual cue. A left or right side visual target appeared 100, 300, or 500 ms later. Attentional effects were observed in the four experiments. In Experiments 2, 3a and 4a, these effects were greater when the cue indicated the right side than when it indicated the left side (respectively: 16 ± 10 and 44 ± 6 ms, P = 0.015, for stimulus onset asynchrony of 500 ms in Experiment 2; 38 ± 10 and 70 ± 7 ms, P = 0.011, for Experiment 3a, and 23 ± 11 and 61 ± 10 ms, P = 0.009, for Experiment 4a). In Experiments 3b and 4b, the central cue pointed to both sides and was said to be non-relevant for task performance. In these experiments right and left reaction times did not differ. The most conservative interpretation of the present findings is that voluntary attention orienting favors the right side of space, particularly when a difficult task has to be performed.

Evaluation of anti-Wnt/β-catenin signaling agents by pGL4-TOP transfected stable cells with a luciferase reporter system

Refractory and relapsed leukemia is a major problem during cancer therapy, which is due to the aberrant activation of Wnt/β-catenin signaling pathway. Activation of this pathway is promoted by wingless (Wnt) proteins and induces co-activator β-catenin binding to lymphoid enhancer factor (LEF)/T-cell factor protein (TCF). To provide a convenient system for the screening of anti-Wnt/β-catenin agents, we designed a bi-functional pGL4-TOP reporter plasmid that contained 3X β-catenin/LEF/TCF binding sites and a selectable marker. After transfection and hygromycin B selection, HEK 293-TOP and Jurkat-TOP stable clones were established. The luciferase activity in the stable clone was enhanced by the recombinant Wnt-3A (rWnt-3A; 100-400 ng/mL) and GSK3β inhibitor (2’Z,3’E)-6-bromoindirubin-3’-oxime (BIO; 5 µM) but was inhibited by aspirin (5 mM). Using this reporter model, we found that norcantharidin (NCTD; 100 µM) reduced 80% of rWnt-3A-induced luciferase activity. Furthermore, 50 µM NCTD inhibited 38% of BIO-induced luciferase activity in Jurkat-TOP stable cells. Employing ³H-thymidine uptake assay and Western blot analysis, we confirmed that NCTD (50 µM) significantly inhibited proliferation of Jurkat cells by 64%, which are the dominant β-catenin signaling cells and decreased β-catenin protein in a concentration-dependent manner. Thus, we established a stable HEK 293-TOP clone and successfully used it to identify the Wnt/β-catenin signaling inhibitor NCTD.

PRODUÇÃO DE LEITE DE CABRAS PURAS NO CURIMATAÚ PARAIBANO DURANTE A LACTAÇÃO

A presente pesquisa teve como objetivo determinar a produção de leite de cabras puras das raças Anglo Nubiana, Parda Alemã e British Alpine, durante 195 dias de lactação, na microrregião do Curimataú paraibano. Foram utilizados 6 animais puros de cada raça, de 2a e 3a ordem de parição. O controle leiteiro teve início 30 dias após a parição, com uma periodicidade de 15 dias, durante 195 dias, nas ordenhas matutina e vespertina, perfazendo um total de 72 coletas. A produção obtida para as raças Anglo Nubiana, Parda Alemã e British Alpine, correspondeu a 873,6, 1.162,0 e 2.069,0g, respectivamente. Foi evidenciada influência significativa (P<0,05) dos parâmetros raça, período de lactação e período de ordenha sobre a produção, não sendo observado para a raça British Alpine o efeito da lactação sobre a produção.

Montreal Cognitive Assessment (MoCA) no rastreio de comprometimento cognitivo leve (CCL) em pacientes com doença renal crônica (DRC) pré-dialítica

Resumo Introdução: Indivíduos com doença renal crônica (DRC) têm grande risco de desenvolver comprometimento cognitivo (CC), inicialmente leve (CCL), passível de identificação, mas ainda subdiagnosticado e subtratado. O Montreal Cognitive Assessment (MoCA) vem sendo indicado para rastreio de CCL na DRC. Objetivo: Avaliar o CCL em indivíduos com DRC pré-dialítica. Métodos: O estudo foi realizado em 72 indivíduos, não idosos, com DRC nos estágios pré-dialíticos. A avaliação neuropsicológica incluiu: o teste de cognição global MoCA; o teste do relógio (TDR); o Digit Span ordem direta (DOD) e inversa (DOI); o teste de fluência verbal (FV), fonêmica (FAS) e semântica (animais); o punho-borda-mão (PBM); e de memória 10 figuras. Resultados: A média de idade dos participantes foi de 56,74 ± 7,63 anos, com predominância de homens (55,6%), com escolaridade ≥ 4 anos (84,3%), a maioria com DRC 1, 2 e 3a e 3b (67,6%), hipertensa (93,1%) e diabética (52,1%). O CC (MoCA ≤ 24) foi observado em 73,6% dos usuários. Não encontramos associação das variáveis demográficas e clínicas com CC, mas tendência de associação com a idade (p = 0,07), com a escolaridade (p = 0,06) e com o DM (0,06). Os testes de função executiva, TDR, DOI e PBM, isoladamente, apresentaram boa sensibilidade e valor preditivo negativo comparados ao MoCA para a identificação de CC e, em conjunto, foram capazes de predizer o resultado do MoCA. Conclusão: O CCL é frequente em usuários não idosos com DRC pré-dialítica. O TDR, DOI e PBM associados são equivalentes ao MoCA na identificação do CC nessa população, sugerindo comprometimento de funções executivas.

Jacobus de Voragine. Ogerius Locediensis. Guillelmus de Malliaco.

F. 13-142v [Jacobus de Voragine, Sermones excerpti e collectione sermonum festivalium per anni circulum]; cf. Schneyer, Repert. lat. Serm., III, 246-268. [In Nativitate]; cf. ibid., n° 319, incomplet du début, et n° 324 (13-18v); — « In circumcissione [sic] Domini »; cf. ibid., n° 346 et 347 (18v-24); — « In Epiphania Domini »; cf. ibid., n° 351 et 354 (24-28v); — « In Purificatione sancte Marie »; cf. ibid., n° 374 et 376 (28v-34); — « In Adnunciatione beate Marie.; cf. ibid., n° 396 (34-36v) et n° 397, incomplet du début par lacune matérielle (38-39); — « De s. Johanne Baptista »; cf. ibid., n° 465 et 467 (39-44); — « De apostolis Petro et Paulo »; cf. ibid., n° 471 et 473 (44-49v); — « De b. Paulo apostolo »; cf. ibid., n° 474 (49v-53); — « De s. Maria Magdalena »; cf. ibid., n° 479 et 480 (53-59v); — « De b. Laurentio martyre »; cf. ibid., n° 497 et 499 (59v-65); — « In Assumpcione b. Marie »; cf. ibid., n° 504 et 509 (65-72v); — « De s. Augustino »; cf. ibid., n° 521 et 523 (72v-79v); — « In Nativitate b. Marie »; cf. ibid., n° 528 et 530, incomplet de la fin, en réclame « virginis » (79v-84v); — [De s. Michaele]; cf. ibid., n° 544, incomplet du début par lacune matérielle, et n° 550 (86-90v); — « In festo omnium sanctorum »; cf. ibid., n° 568 et 570 (90v-96v); — « De mortuis »; cf. ibid., n° 574-577 (96v-107v); — « De s. Martino »; cf. ibid., n° 582, incomplet de la fin (107v-109v) et n° 583 incomplet du début par lacune matérielle (111-113v); — « De b. Katerina »; cf. ibid., n° 591 et 592 (113v-119v); — « De dedicacione ecclesie »; cf. ibid., n° 594 et 595 (119v-127); — « De consecracione altaris »; cf. ibid., n° 597 (127-130); — « De vestibus sacris sacerdotis quid significant »; cf. ibid., n° 598 (130-134); — « De exposicione misse »; cf. ibid., n° 599 et 600 (134-142v). F. 142v-146 « Sermo ad religiosos. Quia existis indesertum... (Mt. XI, 7). Istam questionem quam fecit Dominus... ». F. 146-150v [Guillelmus de Malliaco, Sermo excerptus e collectione sermonum de Tempore dicta « Abjiciamus »]; cf. Schneyer, op. cit., II, 483-489. « De visitacione et officio visitacionis »; cf. ibid., n° 72. F. 150v-158 Sermones. « In concilio magnatum. Quoniam ecce reges terre... (Ps. XLVII, 5). Hic duo tanguntur scilicet magnorum conveniencia temporalis... » (150v-151v); — « In synodo clericorum. Pro patribus tui nati sunt tibi filii... (Ps. XLIV, 17). Adtendant ecclesiarum prelati tria... » (151v-155); — « Sermo ad religiosos. Deus qui habit are facit unanimes in domo secundum hebraicam veritatem et secundum Johannem. Deus qui habitare racit monachos... » (155-158). F. 158-160v [Jacobus de Voragine, Sermo de s. Mathia] « Sermo in electione », incomplet par lacune matérielle, en réclame « vir perfectus »; cr. Schneyer, loc. cit., n° 382, moins développé. F. 161-169v [Ogerius Locediensis] « Planctus b. Bernardi de dolore Marie virginis propter filium » (en titre-courant). « Quis dabit capiti meo aquam et oculis meis imbrem sicut presens dies demostrat [sic] cunctis aperte. Inclita regina celica rosa flos sine spina// ...memento mei »; extrait du De Laudibus b. Virginis, rédaction B; cf. H. Barré, dans Revue d'ascétique et de mystique, XXVIII (1952), 243-266, mss. et éditions. Le texte est incomplet par lacune matérielle, un f. ayant été coupé entre les fr. 163 et 164. F. 169v-184 Sermones. « Sermo in Assumptione b. Marie. Surrexit rex in occursum... (III Reg. II, 19). Quam multiplici figura Salomon ille... », incomplet de la fin (169v-171v); — sermon incomplet du début par lacune matérielle (174); — « Sermo in capite jejunii. Convertimini ad Dominum Deum vestrum... (Joel II, 13). Agreditur hodie Spiritus sanctus multitudinem peccatorum... » (174-175v); — « Sermo in Paraceve. Cum egressus fuero de urbe... (Ex. lX, 29). Verba ista sunt Moysi qui gerit typum Salvatoris... » (175v-180v); — « Alius sermo in Paraceve. O vos omnes qui transitis per viam... (Thren. I, 12). Consideranti michi piam et superpiam materiam... » (180v-183v); — « Domine, bonum est nos hic esse... (Lc. lX, 33). Ubi? Petro. Isti enim tria tanguntur in mentis sublimitate... » (183v-184). F. 184v Table des ff. 1 à 193. — Addition fin XIVe s. F. 185-193 Sermones. « Sermo in Nativitate. Sicut lux aurore oriente sole... (II Reg. XXIII, 4, 2). Verba sunt David cui Dominus... » (185-186); — « Sermo in Paraceve. Deducant oculi nostri lacrimas... (Jer. lX, 18). ...ut gloriosa Virgo septies flevisse.. » (186-191); — « Quod Corpus Christi vere sit in altari. Cenantibus autem eis, accepit Jesus panem... Item Joh. (VI, 51): Ego sum panis vivus... Credebant enim quod manducaretur sicut alie carnes. ..) (191-193). F. 193v Table des ff. 194 à 285. — Addition fin XIVe s. La suite de la table a été coupée. F. 194-335v Sermones de Tempore, excerpti praesertim e collectione « Abjiciamus » Guillelmi de Malliaco et e collectionibus De Tempore et De Sanctis et festis Jacobi de Voragine; cf. Schneyer, op. cit., II, 483-489 et III, 221-233 et 246-268. F. 194-208. [Guillelmus de Malliaco] « Dom. 1a in Adventu Domini »; cf. Schneyer, II, loc. cit., n° 1 et 2 (194-203); Dom. 2a »; cf. ibid., n° 3 (203-208). F. 208-213 [Jacobus de Voragine] « Dom. 2a in Adventu Domini »; cf. Schneyer, III, loc. cit., n° 5. F. 213-218 [Guillelmus de Malliaco] « Dom. 3a »; cf. Schneyer, II, n° 6. F. 218-222 [Jacobus de Voragine] « Dom. 3a »; cf. Schneyer, III, n° 8. F. 222v-228 [Guillelmus de Malliaco] « Dom. 4a »; cf. Schneyer, II, n° 7. F. 228-232 [Jacobus de Voragine] « Dom. eadem »; cf. Schneyer, III, n° 11. F. 232-236. « Feria 4a in capite jejunii. Cum jejunatis nolite fieri... (Mt. VI, 16). Hodie incipit tempus penitentie... ». F. 236-240 « De eadem feria. Convertimini ad me in toto corde... (Joel II, 12). Homo per peccatum tria mala incurrit... ». F. 240-246 [Guillelmus de Malliaco] « Dom. 1a in quadragesima »; cf. Schneyer, II, n° 25. F. 246-319 [Jacobus de Voragine] « Dom. eadem »; cf. Schneyer, 111, n° 41 (246-251v); — « Dom. 2a in quadragesima »; cf. ibid. n° 44 et 45 (251v-259v); — « Dom. 3a in quadragesima »; cf. ibid., n° 47 et 48 (259v-270); — « Dom. 4a in quadragesima »; cf. ibid., n° 50 et 51 (270-279v); — « Dom. de passione »; cf. ibid., n° 53 et 54 (279v-290); — « Dom. de Ramis »; cf. ibid., n° 56 et 57 (290-302); — « In Cena Domini »; cf. ibid., n° 401 et 402 (302-307v); — « In Parasceve »; cf. ibid., n° 405 et 411 (307v-312); — « In die Pasce »; cf. ibid., n° 414 et 415 (312-316v); — « Feria 2a post Pasca »; cf. ibid., n° 419 (316v-319). F. 319-325v « De eadem feria. Duo ex discipulis Jhesu ibant... (Lc. XXIV, 13). Introduxit nos Dominus in terram fluentem lac... » F. 326-334 [Jacobus de Voragine] « In adscensione Domini »; cf. ibid., n° 446 et 445 (326-329v); — [In Pentecoste]; cf. ibid., n° 452 et 457 (329v-334). F. 334-335v « De eodem. Ad Deum [sic pro: eum] veniemus et mansionem... (Job. XIV, 23). Super illo verbo dicit b. Eusebius... ». F. 335v-337v Extraits patristiques. « Incipiunt quedam problemata. Lingua mea calamus ego sum. » Sont cités s. Grégoire, s. Paul, s. Augustin, s. Isidore, Boèce, Bède, Josèphe, s. Hilaire, etc.

Intramolecular carbenoid insertions : the reactions of [alpha]-diazoketones derived from furanyl, thienyl, benzofuranyl and benzothienyl acetic acids with rhodium (II) acetate /

A number of synthetically useful ring systems can be prepared via the intramolecular insertion of a metal-stabilized carbenoid into a heteroaromatic systems. The chemical outcome of these reactions are dependent not only on the nature of the heteroatom but also on the length of the aliphatic tether linking the carbenoid moiety with the aromatic fragment. Our work with furanyl and thienyl systems containing a single methylene tether have allowed for some rather atypical chemistry. For example, treatment of l-diazo-3-(2-thienyl)-2-propanone (6) with catalytic rhodium (II) acetate yields 5,6- dihydro-4^-cyclopenta[Z>]thiophen-5-one (3) while, the isomeric l-diazo-3-(3-thienyl)-2- propanone(15) gives a spiro-disulphide (20). Novel chemistry was also exhibited in the analogous furanyl systems. While treatment of l-diazo-3-(3-furanyl)-2-propanone (52) with Rh2(OAc)4 resulted in the expected 2-(4-Oxo-2-cyclopentenyliden)acetaldehyde (54), isomeric l-diazo-3-(2- furanyl)-2-propanone (8) undergoes vinylogous Wolff rearrangement to give a mixture of 6a-methyl-2,3,3a,6a-tetrahydrofuro[2,i-^>]furan-2-one (44) and 2-(2-methyl-3-furyl)acetic acid (43). Rhodium acetate catalyzed decomposition of l-diazo-3-(3-benzofuranyl)-2- propanone (84) and l-diazo-3-(2-benzofuranyl)-2-propanone (69)also allows for vinylogous Wolff rearrangement, a chemistry unseen in benzofuranyl systems with longer tethers. A number of interesting products were isolated from the trapping of intermediate ketenes. Decomposition of l-diazo-3-(3-benzothienyl)-2-propanone (100) resulted in the formation of 2,3-dihydro-l//-benzo[^]cyclopenta[^thiophen-2-one (102). However, in addition to (102), a dimer was also generated from the decomposition of l-diazo-3-(2- benzothienyl)-2-propanone (109). The insight into the mechanistic underpinnings of the above reactions are provided by molecular modeling at a PM3 level.

Synthesis of fluorescent analogues of a-tocopherol as ligands for the human a-tocopherol transfer protein (a-TTP) /

To further understand in vivo localization and trafficking of a-tocopherol (a-Toe), the most biologically active form of vitamin E, between lipid environments, tocopherols are required that can be followed by teclu1iques such as confocal microscopy and fluorescence resonance energy transfer (FRET) assays. To this end, sixteen fluorescent analogues of a-tocopherol (la-d [(1)anthroy loxy -a-tocopherols, A O-a-Toes], 2a-d [w-nitro benzoxadiazole-a-tocopherols, NBD-aToes], 3a-d [w-dansyl-a-tocopherols, DAN-a-Toes], and 4a-d [w-N-methylanthranilamide-atocopherols, NMA-a-TocsD were prepared by substituting fluorescent labels at the terminus of w-functionalized alkyl chains extending from C-2 of the chroman ring while retaining key binding features of the natural ligand. These compounds were prepared starting from (S)-Trolox® acid VIa esterification, protection, and reduction producing the silyl-protected (S)-Trolox aldehyde that was coupled using Wittig chemistry to different w-hydroxyalkylphosphonium bromides. Reduction of the alkene generated the w-hydroxy functionalized 2-n-alkyl intermediates 9a-d having the necessary 2R stereochemistry. A series of functional group manipulations including mesylation, substitution with azide, and hydride reduction provided w-amino functionalized intermediates 12a-d as well. Coupling intermediates 9a-d and 12a-d with the selected fluorophores (9- anthracene carboxylic acid, 4-chloro-7-nitrobenz-2-oxa-l,3-diazole, 5- dimethylaminonapthalene-l-sulfonyl chloride, and I-methyl-2H-3,1-benzoxazine-2,4(1H)dione), followed by deprotection of the phenolic silyl group, gave the desired fluorescent ligands la-d, 2a-d, 3a-d and 4a-d in good yield. Assessment of their binding affinities with recombinant human a-tocopherol transfer protein (ha-TTP) utilizing fluorescent titration binding assays identified competent ligands for further use in protein studies. Compounds Id (C9-AO-a-Toc) and 2d (C9-NBD-a-Toc) both having nonyl alkyl chain extensions between the chromanol and fluorophore were shown to bind specifically to ha-TTP with dissociation constants (KdS) of approximately 280 nM and 55 nM respectively, as compared to 25 nM for the natural ligand 2R,4'R,^'R-a-tocophQxoL.

Intramolecular carbenoid insertion : reactions of [alpha] -diazoketones derived from benzothienyl, pyrolyl and indolyl alkanoic acids with rhodium (II) acetate

Recent studies have shown that the rhodium (II) acetate decomposition chemistry observed for a-diazoketones tethered to thienyl, furanyl, and benzofuranyl moieties is dependent not only on the nature of the heteroatom but also on the length of the aliphatic tether linking the diazoketone moiety with the aromatic fragment. The present thesis expands on these results and focuses on a-diazoketones tethered to benzothiophenes, pyrroles and indoles by a methylene linker. In the case of benzothiophenes, it was shown that the rhodium catalyst decomposition of I-diazo-4-(3-benzothienyl)-2-butanone (146) and 1-diazo-4-(3benzothienyl)- 2-butanone (152) allow for the isolation of 1,2,3a,3b-tetrahydro-3Hbenzo[ b]cyclopenta[1,3]cyclopropa- [1 ,2-d]thiophen-3-one (147) and 1,2,3a,3btetrahydro- 3H-benzo[b]cyclopenta[1,3]cyclopropa[1,2-d]thiophen-3-one (153). However treatment of 1-diazo-3-(3-Benzothienyl)-2-Propanone (165) with Rh(II) acetate results in the formation of 2,3-Dihydro-1H-benzo[b]cyclopenta[d]thiophen-2-one (159), while 1diazo- 3-(2-Benzothienyl)-2-Propanone with the same condition gives 5,5-bis( 1benzothiophen- 2-ylmethyl)-2(5H)-furanone (166) along with the tricycle 159. The chemistry of the pyrrolyl and the indolyl moieties linked to terminal adiazoketone systems was also investigated. The decomposition of I-diazo-(2-pyrrolyl)-2propanone (173) results in the formation of two products; the N-H insertion product IHpyrrolizin- 2(3H)-one (176) and the alkylation product 4,6-dihydrocyclopenta[b]pyrrol5( 1 H)-one (180). When 1-Diazo-3-(3-indoly)-3-propanone (194) is treated with catalytic amount of Rh (II) 3,4-dihydrocyclopenta[b]indol-2(1H)-one (193) is isolated quantitatively. The later reaction when monitored using IH NMR the intermediate 200 can be seen whose structure was confirmed by the comparison to series of model compounds. The mechanisms underlying these reactions as well as their synthetic utility is discussed.