Experiments Towards the Synthesis of the Ergot Alkaloids and Related Structures. Part 8.17 The Total Synthesis of 4-Methyl-2,3,4,4a,5,6-hexahydro-benzo[f]quinoline-2-carbonitrile Hydrochloride Hemihydrate,an Immediate Precursor of the Despyrrole Analogues of Lysergic Acid and Its Amides

Exposure of the N -methoxycarbonyl-bicyclic-keto-acid 5 (improved preparation) to the Barnick β -keto-acid synthesis 1 yielded an aqueous solution of the sodium salts of the β -keto-acids 26 and 27 which on heating at 60-65°C furnished the N -methoxycarbonyl-tricyclic-ketone 9 (55%) plus the hydroxy-ketone 28 which on acid treatment raised the yield of 9 to 68%. Reduction (NaBH 4) of 9 yielded the alcohol 32 (94%) which was treated with thionyl chloride followed by copper (I) cyanide and sodium iodide in acetonitrile to give the tricyclic- N -methoxycarbonyl nitrile 35 whose relative configuration was obtained by X-ray analysis. Attempts to remove the N -methoxycarbonyl group from 35 were unsuccessful. Conversion of the alcohol 32 to its methoxypropyl ether 41 followed by reaction with ethereal MeLi-LiBr yielded the amino-alcohol 39 (75%) converted to the N -formyl-tricyclic alcohol 42 with formic-acetic anhydride (70%). The alcohol 42 was then converted into the N -formyl nitrile 44 via the chloride 43 as employed in the earlier synthesis of the nitrile 35. Removal of the N -formyl group from the nitrile 44 was achieved by refluxing methanolic hydrochloric acid to give the required amino-nitrile hydrochloride 46 (91%) whose structure was confirmed by X-ray analysis. Reaction of the free base with methyl iodide in ethyl acetate in the presence of calcium carbonate furnished the N -methyl base 48 isolated as its hydrochloride, hemihydrate 49 (59%). The overall yield of 49 via this eleven-step synthesis was 3.4%.     

Novel azido fatty acid ester derivatives from conjugated C(18) enynoate.

Methyl octadec-11Z-en-9-ynoate (1) was epoxidized to give methyl 11,12-Z-epoxy-octadec-9-ynoate (2, 81%). Acid catalyzed ring opening of the epoxy ring of compound 2 gave methyl 11,12-dihydroxy-octadec-9-ynoate (3, 80%). The latter was treated with mesyl chloride to yield methyl 11,12-dimesyloxy-octadec-9-ynoate (4, 76%). Reaction of compound 4 with sodium azide furnished methyl 11-azido-12-mesyloxy-octadec-9-ynoate (5a, 49%) and methyl 11-azido-octadec-11E-en-9-ynoate (5b, 24%). Compound 2 was semi-hydrogenated over Lindlar catalyst to give methyl 11,12-Z-epoxy-octadec-9Z-enoate (6, 90%). This allylic epoxy fatty ester (6) was reacted with sodium azide to give a mixture of methyl 11-azido-12-hydroxy-octadec-9Z-enoate (7a) and methyl 9-azido-12-hydroxy-octadec-9E-enoate (7b), which could not be separated into individual components by silica chromatography. Chromic acid oxidation of the mixture of compounds 7a and 7b furnished methyl 9-azido-12-oxo-octadec-10E-enoate (8, 42% based on amount of compound 6 used) and an intractable mixture of polar compounds. The various products were characterized by NMR spectroscopic and mass spectral analyses.     

Preparation of cyclic 2',3'-carbamate derivatives of erythromycin macrolide antibiotics

Tricarbonylation of clarithromycin has been effected in a one-pot reaction with phosgene. The 11,12-diol moiety was closed into a cyclic carbonate, while the dimethylamino alcohol of the desosamine sugar was cyclised with loss of a methyl group to form a cyclic 2',3'-carbamate. The 4' hydroxyl group in clarithromycin was converted into a chloroformate group and subsequently to an allyl carbonate which on Pd-catalysis furnished a novel N-demethylclarithromycin 2',3'-carbamate-11,12-carbonate. Hydrolytic removal of the cladinose sugar and a subsequent oxidation furnished the corresponding ketolide. The 11,12-cyclic carbonate moiety was cleaved by sodium azide to the 10,11-anhydro-9-ketone. 11-N-Arylated cyclic 11,12:2',3'-dicarbamate derivatives were prepared in a copper(I) chloride aided reaction between aryl isocyanates and 10,11-anhydro 9-ketones. The products are novel N-arylated-N'-demethylated 11,12:2',3'-dicarbamate ketolides derived from clarithromycin.     

The synthesis of [14C]SCH 40054/A-57219 hydrochloride

A two-step, no-carrier-added synthesis of 2-(4-amino-3,5-dichlorobenzyl)[¹⁴C)imidazoline, SCH 40054/A-57219 (1) was performed starting with [¹⁴C]KCN. The labelled cyanide ion was reacted with the appropriate benzyl chloride to give a nitrile. The nitrile was converted to an imidazoline by reaction with ethylene diamine. Purification of the product was complicated by its instability at neutral or alkaline pH.     

5,6-unsaturated hexofuranosyl glycosides and 5',6'-unsaturated hexofuranosyl nucleosides.

Methyl 5,6-dideoxy-2,3-O-isopropylidene-alpha-D-lyxo-hex-5-enofuranoside, prepared from methyl 2,3-O-isopropylidene-5,6-di-O-methylsulfonyl-alpha-D-mannofuranoside with sodium iodide in 2-butanone, was acetolyzed and the product coupled with 6-benzamidochloromercuripurine by the titanium tetrachloride method. Removal of the N-benzoyl group with pictic acid afforded 9-(2,3-di-O-acetyl-5,6-dideoxy-beta-D-xylo-hex-5-enofuranosyl)adenine. In a similar manner, methyl 5,6-dideoxy-2,3-O-isopropylidene-alpha-L-lyxo-hex-5-enofuranoside was prepared from L-mannose and converted into 9-(2,3-di-O-acetyl-5,6-dideoxy-beta-L-xylo-hex-5-enofuranosyl)adenine, further de-esterified to give the free nucleoside. 2,3:5,6-Di-O-isopropylidene-alpha-L-mannofuranosyl chloride, prepared from L-mannose, gave 9-(2,3-O-isopropylidene-alpha-L-mannofuranosyl)adenine, hydrolyzed into 9-alpha-L-mannofuranosyladenine. Treatment with methanesulfonyl chloride gave the 5',6'-dimethanesulfonate, which gave with sodium iodide in acetone the 5',6'-unsaturated nucleoside, further hydrolyzed into 9-(5,6-dideoxy-alpha-L-lyxo-hex-5-enofuranosyl)adenine.     

A concise synthesis of two isomeric pentasaccharides, the O repeat units from the lipopolysaccharides of P. syringae pv. porri NCPPB 3364T and NCPPB 3365

A concise synthesis of two isomeric pentasaccharides, alpha-L-Rhap-(1-->2)-alpha-L-Rhap-(1-->3)-alpha-L-Rhap-(1-->3)-[beta-D-GlcpNAc-(1-->2)]-alpha-L-Rhap (A) and alpha-L-Rhap-(1-->2)-alpha-L-Rhap-(1-->3)-[beta-D-GlcpNAc-(1-->2)]-alpha-L-Rhap-(1-->3)-alpha-L-Rhap (B), the O repeats from the lipopolysaccharides of Pseudonomonas syringae pv. porri NCPPB 3364T and 3365 was achieved via assembly of the building blocks, allyl 3,4-di-O-benzoyl-alpha-L-rhamnopyranoside (1), 2,3,4-tri-O-benzoyl-alpha-L-rhamnopyranosyl trichloroacetimidate (2), allyl 4-O-benzoyl-3-O-chloroacetyl-alpha-L-rhamnopyranoside (6), 3,4,6-tri-O-acetyl-2-deoxy-2-phthalimido-beta-D-glucopyranosyl trichloroacetimidate (7), and allyl 2,4-di-O-benzoyl-alpha-L-rhamnopyranoside (10). Coupling of 1 with 2 followed by deallylation and trichloroacetimidate formation gave the disaccharide donor 5, while condensation of 6 with 7, followed by dechloroacetylation, offered the disaccharide acceptor 9. Then, 5 was coupled with 10 to obtain the trisaccharide 11, and subsequent deallylation and trichloroacetimidate formation furnished the trisaccharide donor 13. Coupling of 9 with 13, followed by deprotection, afforded pentasaccharide 19, while condensation of 9 with 5, followed by deallylation and trichloroacetimidate formation, gave the tetrasaccharide donor 16, whose coupling with 10 and subsequent deprotection yielded another pentasaccharide 22.     

A convenient synthesis of (2R,8R)-8-methyl-2-decanol and its 2S epimer,the parent alcohols of rootworm pheromones

A highly stereospecific synthesis of the title compound (2R,8R)-8-methyl-2-decanol (I) has been devised via 16 simple steps. The required chirons (5 and 11) were prepared from easily accessible templates viz. (R)-methyl citronel-late and (S)-glutamic acid. This on Grignard coupling furnished the ketal (12) which was converted to the desired epoxide (14) and subsequently reduced to furnish the alcohol (2R,8R)-I. Its corresponding (2S)-epimer was prepared by total stereoselective inversion of its C-2 center. The title compounds are the parent alcohols of the pheromone components of the female rootworms.     

Growth factor-stimulated protein synthesis is inhibited by sodium orthovanadate.

The study of intracellular signaling pathways has been aided by the use of sodium orthovanadate, a cell-permeable inhibitor of tyrosine phosphatases. However, long-term addition of sodium orthovanadate is often cytotoxic. In this study we demonstrate that the growth factor-mediated increase in the rate of protein synthesis was inhibited by sodium orthovanadate. This effect of sodium orthovanadate was dose-dependent, with an IC50 of 40 microM and maximal inhibition obtained at 100 microM. As a consequence, the fetal bovine serum-mediated induction of the immediate-early genes, c-Fos and MKP-1, at the protein level was inhibited by orthovanadate. Orthovanadate's ability to attenuate protein synthesis was partially reversible, and was no longer evident when the agent was added 6 h after addition of growth factor to cells. Analysis of several elements of signaling pathways which are known to regulate protein synthesis in a positive manner (p42/p44 MAPK, AKT and p70 S6K stimulation, and hyperphosphorylation of PHAS-I) were not inhibited but rather were stimulated by orthovanadate. Thus, sodium orthovanadate is a potent inhibitor of growth factor-stimulated protein synthesis independent of p42/p44 MAPK or PI3K-p70 S6K activation.     

Nitrile hydratase and amidase from Rhodococcus rhodochrous hydrolyze acrylic fibers and granular polyacrylonitriles

Rhodococcus rhodochrous NCIMB 11216 produced nitrile hydratase (320 nkat mg of protein(-1)) and amidase activity (38.4 nkat mg of protein(-1)) when grown on a medium containing propionitrile. These enzymes were able to hydrolyze nitrile groups of both granular polyacrylonitriles (PAN) and acrylic fibers. Nitrile groups of PAN40 (molecular mass, 40 kDa) and PAN190 (molecular mass, 190 kDa) were converted into the corresponding carbonic acids to 1.8 and 1.0%, respectively. In contrast, surfacial nitrile groups of acrylic fibers were only converted to the corresponding amides. X-ray photoelectron spectroscopy analysis showed that 16% of the surfacial nitrile groups were hydrolyzed by the R. rhodochrous enzymes. Due to the enzymatic modification, the acrylic fibers became more hydrophilic and thus, adsorption of dyes was enhanced. This was indicated by a 15% increase in the staining level (K/S value) for C. I. Basic Blue 9.     

Acyl-CoA reductase specificity and synthesis of wax esters in mouse preputial gland tumors.

Long-chain alcohols are synthesized in the mouse preputial gland tumor (ESR-586) by NADPH:acyl-CoA oxidoreductase. In this study, a series of labeled acids was tested as substrates for the oxidoreductase in a cell-free system from the tumor, and the distribution of label into alcohols, waxes, and other products was determined. The system contained the labeled acid, an acyl-CoA-generating system, an NADPH-generating system, and tumor homogenate. The highest rates of alcohol synthesis were obtained with palmitic (16:0), heptadecanoic (17:0), stearic (18:0), myristic (14:0), elaidic (18:1 trans), and linoleic (18:2) acids, which yielded, respectively, 151, 124, 102, 76, 65, and 35 pmol alcohol/min per mg protein. Decanoic (10:0), lauric (12:0), oleic (18:1 cis), linolenic (18:3), arachidonic (20:4), and behenic (22:0) acids all gave lower activities. Acyl-CoA formation did not appear to be rate limiting with any of the substrates tested except behenic acid. In addition to the fatty alcohol product, a small amount of fatty aldehyde was formed in the system. Incorporation of the labeled fatty acids into wax esters was examined and the distribution of label between the alcohol and acid components of the waxes was determined. Incubation of [1-(14)C]palmitic acid yielded 3.4% free alcohol, 8.3% alcohol esterified in waxes, and 7.7% palmitoyl groups esterified into waxes, whereas, at the other extreme, [1-(14)C]linolenic acid yielded 0.8%, 0.6%, and 38%, respectively, into the homologous components.-Wykle, R. L., B. Malone, and F. Snyder. Acyl-CoA reductase specificity and synthesis of wax esters in mouse preputial gland tumors.     

Specific oligoribonucleotide synthesis using primer-independent polynucleotide phosphorylase.

Oligoribonucleotides of a predetermined base sequence beginning with adenylyl-3', 5'-adenosine at the 5' end have been synthesized in yields varying between 13% and 42%. The synthesis was carried out using primer-independent polynucleotide phosphorylase from E. coli in the presence of high concentrations of primer and of sodium chloride.     

Improved method for the synthesis of 1- or 3-acyl-sn-glycerols

Optically active 1- or 3-acyl-sn-glycerols were synthesized from 2,3- or 1,2-isopropylidene-sn-glycerols, respectively. The 2,3- or 1,2-isopropylidene-sn-glycerols were condensed with appropriate long saturated or unsaturated fatty acids and the resulting acyl isopropylidene compounds were treated with dimethylboronbromide at - 50 degrees C to give the title compounds. The ketal cleavage of acyl isopropylidene-sn-glycerols by dimethylboronbromide to produce the long 1- or 3-acyl-sn-glycerols was effective and gave good yields (70-90%). The reaction conditions were mild and there was no acyl migration, as shown by optical rotation of the monoacyl-sn-glycerols. The synthesis of 2,3-isopropylidene-sn-glycerol was improved to give an overall yield of 40% from L-arabinose. L-Arabinose was first converted to its 1,1'-diethylmercapto derivative and then condensed with 2-methoxypropene to yield 1,1'-diethyl-mercapto-4,5-isopropylidene-L-arabinose. Oxidation of this compound with sodium periodate followed by reduction with sodium borohydride under alkaline conditions yielded 2,3-isopropylidene-sn-glycerol [alpha]22D = -14.90 degrees, neat (Lit. 8 [alpha]22D = -14.5 degrees, neat; 14 [alpha]25D = -10.8 degrees; methanol C, 16.9). The optical purity of isopropylidene-sn-glycerols was determined as benzoyl derivatives on a high performance liquid chromatographic column packed with a chiral stationary phase.     

Synthesis of pyrrolo[2,1-f][1,2,4]triazine C-nucleosides. Isosteres of sangivamycin, tubercidin, and toyocamycin

Syntheses of pyrrolo[2,1-f][1,2,4]triazine C-nucleosides are reported. Treatment of pyranulose glycoside with aminoguanidine in acetic acid gave the corresponding semicarbazone in 96% yield. The ring transformation of the semicarbazone in dioxane afforded a 51% yield of 2-amino-7-(2,3,5-tri-O-benzoyl-beta-D-ribofuranosyl)pyrrolo[2,1-f]-[1,2,4]triazine. Vilsmeier formylation of the pyrrolotriazine gave the major product, 5-formylpyrrolo[2,1-f][1,2,4]triazine, in 69% yield. The aldehyde was treated with hydroxylamine hydrochloride in methanol to give aldoximes. Dehydration of aldoxime with trifluoromethanesulfonic anhydride and triethylamine in dichloromethane afforded 5-cyanopyrrolo[2,1-f][1,2,4]triazine in 44% yield. Conversion of the nitrile to the deprotected amide, 2-amino-7-(beta-D-ribofuranosyl)pyrrolo[2,1-f][1,2,4]triazine-5-carboxamide, was accomplished in 96% yield on treatment with 30% H2O2 in ethanol for 1 day at room temperature. Debenzoylation with sodium hydroxide solution produced deprotected C-nucleosides.     

Isolation, characterization and synthesis of alpha-foetoprotein from neonatal-rat skin.

Monospecific anti-[rat alpha-foetoprotein(alpha-FP)] immunoglobulin G was coupled to CNBr-activated Sepharose-4B (4.5 mg/ml packed volume of gel) to yield an adsorbent. The immunoaffinity column was used to isolate alpha-FP from neonatal-rat skin. Purified skin alpha-FP was found to be immunologically and electrophoretically similar to serum alpha-FP. It yielded a single band with mol.wt. 68000 on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. However, on polyacrylamide-gel electrophoresis under non-denaturing conditions, the alpha-FP displayed slow- and fast-moving variants similar to those observed in serum alpha-FP. A Scatchard plot of oestradiol binding to the alpha-FP yielded an association constant of 2.5 X 10(9)M-1 by dextran-coated-charcoal and 0.75 X 10(8)M-1 by Sephadex-gel-filtration procedures respectively. Skin explants from newborn rats were found to incorporate [14C]leucine into immunoprecipitable intracellular alpha-FP. Cycloheximide inhibited the synthesis of alpha-FP in skin explant culture. Our results indicate that newborn-rat skin contains alpha-FP that is similar to serum alpha-FP and which may arise in neonatal-rat skin as a result of synthesis in situ.     

A preparative synthesis of 3-amino-2,3,6-trideoxy-L-lyxo-hexose (daunosamine) hydrochloride from D-mannose.

A simple, preparative route in nine steps from methyl alpha-D-mannopyranoside (1) is described that affords, in 40% overall yield, the title amino sugar 11, the sugar constituent of the antitumor antibiotics adriamycin and daunorubicin. The 2,3:4,5-dibenzylidene acetal (2) of 1 is converted by butyllithium into the 2-deoxy-3-ketone 3, whose oxime 4 is reduced with high stereoselectivity to the D-ribo amine, isolated as its N-acetyl derivative 5 and converted by action of N-bromosuccinimide into the 4-O-benzoyl-6-bromide 7. Dehydrohalogenation of gives the 5,6-unsaturated glycoside 8, which, after O-debenzoylation to 9, undergoes stereospecific reduction by hydrogen with net C-5 inversion to give the crystalline, N-acetylated methyl beta-glycoside (10) of daunosamine, readily converted into daunosamine hydrochloride (11) and into the crystalline N-benzoyl (14) and N-acetyl(15) derivatives. No chromatographic procedures for isolation are required in any of the steps.     

Highly enantioselective synthesis, crystal structure, and circular dichroism spectroscopy of (R)-bambuterol hydrochloride

The present article describes the asymmetric synthesis of (R)-bambuterol hydrochloride based on 1-(3,5-dihydroxyphenyl)ethanone as starting material, which was esterified by dimethylcarbamic chloride, and brominated by copper (II) bromide. Then the carbonyl group was reduced efficiently using (-)-B-chlorodiisopinocamphenylborane [(-)-DIP-chloridetrade mark] as an asymmetrical reducing agent. Followed by epoxide ring closure with NaOH and ring expansion with tert-butylamine led to the desired product (R)-bambuterol with e.e. up to 99%. The optical properties and absolute configuration of (R)-bambuterol hydrochloride were further investigated using circular dichroism spectroscopy and X-ray single crystal analysis.     

Antimicrobial and antitumor activity of N-heteroimmine-1,2,3-dithiazoles and their transformation in triazolo-, imidazo-, and pyrazolopirimidines.

The reaction of Appel's salt with o-amino nitrile heterocycles 10-19 gave the corresponding 4-chloro-5-heteroimmine-1,2,3-dithiazoles 20-29 which were evaluated for their antibacterial, antifungal and antitumor activity. Although all these N-heteroimines were devoid of significant antibacterial activity, they showed significant antifungal activity. Moreover, the same derivatives represent highly versatile intermediates in heterocyclic synthesis, in fact the pyrazoleimino dithiazoles 20-26 can be converted in one step into 2-cyano derivatives of the corresponding 4-methoxy-pyrazolo[3,4-d]pyrimidines 30-35 by sodium methoxide in refluxing methanol. This provides a general and attractive route to 4-methoxy-6-cyano pyrazolo[3,4-d]pyrimidines from 1-substituted 5-amino pyrazoles 10-19 in two simple steps. Finally, the isosteric replacement of the pyrazole ring atoms to give the imidazole[3,4-d]pyrimidine and triazole [4,5-d] pyrimidine ring systems was examined.     

New useful reagents for peptide synthesis. Insoluble active esters of polystyrene-bound 1-hydroxybenzotriazole.

Insoluble 1-hydroxbenzotriazole-bound polystyrene was prepared through a series of chemical modifications of commercially available polystyrene. Reaction of 3-nitro-4-chlorobenzyl alcohol or of 3-nitro-4-chlorobenzyl bromide with polystyrene in the presence of aluminium trichloride yielded (3-nitro-4-chloro)benzylated polystyrene. Upon reaction with hydrazine it was converted to (3-nitro-4-hydrazine) benzylated polystyrene which was cyclized, by acidolysis, to yield 1-hydroxybenzotriazole-bound polystyrene. This was coupled, using N, N' -dicyclohexylcarbodiimide as the coupling agent, to many N-blocked amino acid derivatives, yielding polymeric polystyrene-bound active esters.Such derivatives are highly reactive and their efficacy in the synthesis of several peptides, including that of the tetrapeptide Boc-L-Leu-L-leu-L-Val-0bzl-L-Tyr-0Bzl and of thyrotropin-releasing hormone was demonstrated.     

Role of volume and prostaglandin synthesis in the suppression of renin by saline in the rat

To evaluate the contribution of plasma volume expansion per se on acute inhibition of renin release by sodium chloride infusion, renin responses to comparable plasma volume expansion with intravenous infusions of sodium chloride, sodium bicarbonate, or albumin were studied in separate groups of sodium chloride-depleted rats. In addition, urinary prostaglandin E2 (PGE2) excretion rate was compared in the saline- and sodium bicarbonate-infused animals to evaluate the relationship between acute changes in renin release and intrarenal PGE2 synthesis. All three groups were plasma volume-expanded by approximately 55%. Plasma renin activity (PRA) decreased in response to saline (12.3 +/- 1.0 to 6.7 +/- 0.7 ng AI/ml/hr; P less than 0.01) whereas PRA did not change with sodium bicarbonate (11.3 +/- 1.4 to 10.2 +/- 1.5) or albumin (9.9 +/- 0.7 to 8.2 +/- 1.0). The rate of PGE2 excretion was not changed by either saline (72.2 +/- 13.1 to 72.3 +/- 18.7 pg/min) or sodium bicarbonate infusion (70.7 +/- 8.8 to 64.9 +/- 7.0). These results support the hypothesis that acute suppression of PRA by infusion of saline is not dependent upon volume expansion per se. In confirmation of earlier observations, inhibition of renin release by sodium chloride was related to chloride. Finally, the results suggest that the renal tubular mechanism for inhibition of renin release by sodium chloride is not related to overall changes in renal PGE2 synthesis in the rat.     

Control of zinc-thionein synthesis in rat liver.

The rate of [35S]cystine incorporation into hepatic zinc-thionein (a metallothionein) was stimulated, with a maximum of 5-6h, after parenteral administration of 2mg of Zn2+ containing 65Zn. The binding of 65Zn to zinc-thionein was measurable by 2-1/2h and reached a plateau by 18h after the injection. A net increase in the hepatic 65Zn content was observed subsequent to the decrease in the rate of zinc-thionein synthesis. The incorporation of both 65Zn and [35S]cystine into zinc-thionein was inhibited by prior administration of either actinomycin D or cordycepin. A second injection of Zn2+, 20h after the initial injection, yielded a 4.9-fold greater increase in zinc-thionein synthesis compared with that after only one injection; however, this synthesis was also inhibitable by actinomycin D. These data support the concept that hepatic zinc-thionein synthesis responds quickly to changes in Zn2+ status and that Zn2+ is bound subsequent to synthesis of nascent thionein chains. The mechanism of control of zinc-thionein synthesis by Zn2+ appears to involve changes in the amounts of a short-lived, poly(A)-containing RNA whose translation can be derepressed by additional exposure to Zn2+.