Synthesis of deuterium-labeled 16 alpha,19-dihydroxy C19 steroids as internal standards for gas chromatography-mass spectrometry.

[2 beta,7,7,16 beta-2H4]16 alpha,19-Dihydroxyandrost-4-ene-3,17-dione (14) and [7,7,16 beta-2H3]3 beta,16 alpha,19-trihydroxyandrost-5-en-17-one (16), with high isotopic purity, respectively, were synthesized from unlabeled 3 beta-(tert-butyldimethylsiloxy)-androst-5-ene-17 beta-yl acetate (1). The deuterium introduction at C-7 was carried out by reductive deoxygenation of the 7-keto compound 3 with dichloroaluminum deuteride and that at C-2 beta and/or C-16 beta by controlled alkaline hydrolysis of 16-bromo-17-ketone 11 or 12 with NaOD in D2O and pyridine. [7,7-2H2]3 beta-Hydroxyandrost-5-en-17-one (6), obtained from compound 1 by a five-step sequence, was converted to compound 14 or 16 by an eight-step or seven-step sequence, respectively. The labeled steroids 14 and 16 are useful as internal standards for gas chromatography-mass spectrometry analysis of the endogenous levels.     

Stereochemistry of the cyclization-rearrangement of (+)-copalyl diphosphate to (-)-abietadiene catalyzed by recombinant abietadiene synthase from Abies grandis

[reaction: see text] Syntheses and enzymatic cyclizations of 8alpha-hydroxy-17-nor copalyl diphosphate (8a), (15R)-[15-2H1] 8b, and (15R,17E)-[15-3H1,17-2H1] copalyl diphosphate ([2H,3H] 2) catalyzed by recombinant abietadiene synthase (rAS) gave 17-nor manoyl oxide (9a), (16E)-[16-2H1] 9b, and (15S,16R)-[16-2H1,16-3H1] abietadiene ([2H1,3H1] 4), respectively. These and other results indicate that conversion of CPP (2) to abietadiene (4) occurs by anti S(N)' cyclization to a sandaracopimar-15-en-8-yl carbocation intermediate (13+, 13beta-methyl) followed by hydrogen transfer and methyl migration suprafacially on the si face of the vinyl group.     

Thiophene chemistry—XXII : Some reactions of benzo[b]thiophene-2(3H)one

Benzo[b]thiophene-2(3H)one has been prepared from 2-t-butoxybenzo[b]thiophene by dealkylation. Alkylation of sodium, thallium and tetrabutylammonium salts of benzo[b]thiophene-2(3H)one produces both C- and O-alkylation along with products due to ring-opening. At elevated temperatures benzo[b]thiophene-2(3H)one reacts with HMPA (hexamethylphosphorictriamide) to give 2-dimethylaminobenzo[b]thiophene. Other 2-aminobenzo[b]thiophenes are produced by refluxing benzo[b]thiophene-2(3H)one in HMPA in the presence of excess of the corresponding amine.     

Analysis of triterpenoids in <Emphasis Type="Italic">Ganoderma lucidum</Emphasis> using liquid chromatography coupled with electrospray ionization mass spectrometry

Triterpenoids extracted from Ganoderma lucidum (Leyss. ex Fr.) Karst were separated and characterized using optimized reversed-phase liquid chromatography with diode array detection and electrospray ion trap tandem mass spectrometry (HPLC-DAD-ESI-MS(n)). They could be classified into five types depending on the fragmentation behavior. All triterpenoids gave [M - H](-) and [2M - H](-) ions by electrospray ionization monitored in the negative ion mode; in addition, compounds of types III and IV gave prominent [M - H - H(2)O](-) ions and the unsaturated bond at C-20, 22 would reduce the abundance of [M - H - H(2)O](-) ion. The key fragmentation information was cleavage at C- and D-rings despite the predominant losses of H(2)O and CO(2). Compounds with hydroxyls at C-7 and C-15 would produce a list of b, b - 1, b - 2, and b - 16 ions attributed to cleavage of D-ring; if the second alcohol at C-15 were oxidized to ketone, the prominent cleavage would occur at C-ring and produce a group of ions of a; if C-7 were oxidized to ketone, transference of two hydrogen atoms would occur during the cleavage of rings and a list of ions about a + 2 and/or b + 2 would appear instead. The above fragmentations and regularities in fragmentation pathways were reported for the first time, and were implemented for the analysis of triterpenoids in G. lucidum. The chloroform extract was separated on a Zorbax SB-C(18) column, eluting with an acetonitrile-0.2% acetic acid gradient. A total of 32 triterpenoids, including six new ones, were identified or tentatively characterized based on the tandem mass spectra of the HPLC peaks.     

Synthesis and antimicrobial activity of some new pyrazole, fused pyrazolo[3,4-d]-pyrimidine and pyrazolo[4,3-e][1,2,4]-triazolo[1,5-c]pyrimidine derivatives

Hydrazonyl bromides 2a,b reacted with active methylene compounds (dibenzoylmethane, acetylacetone, ethyl acetoacetate, phenacyl cyanide, acetoacetanilide, ethyl cyanoacetate, cyanoacetamide and malononitrile) to afford the corresponding 1,3,4,5- tetrasubstituted pyrazole derivatives 5-12a,b. Reaction of 12a,b with formamide, formic acid and triethyl orthoformate give the pyrazolo[3,4-d]pyrimidine, pyrazolo[3,4- d]pyrimidin-4(3H)one and 5-ethoxymethylene-aminopyrazole-4-carbo-nitrile derivatives 13-15a,b, respectively. Compounds 15 a,b reacted with benzhydrazide and hydrazine hydrate to afford pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidine and [4-iminopyrazolo- [3,4-d]pyrimidin-5-yl]amine derivatives 16 a,b and 17 a,b. Reactions of compounds 17 a,b with triethyl orthoformate and carbon disulfide give the corresponding pyrazolo[4,3-e]- [1,2,4]triazolo[1,5-c]pyrimidine derivatives 18a,b and 19 a,b, respectively.     

1,3-Diphenylbenzo[e][1,2,4]triazin-7(1H)-one: selected chemistry at the C-6, C-7 and C-8 positions

1,3-Diphenylbenzo[e][1,2,4]triazin-7(1H)-one (6) reacts with tetracyanoethylene (TCNE) or tetracyanoethylene oxide (TCNEO) to give the deep green 2-[1,3-diphenylbenzo[e][1,2,4]triazin-7(1H)-ylidene]propanedinitrile (11) in 17 and 15% yields, respectively. Nucleophiles such as amines, alkoxides, thiols and Grignard reagents all reacted with the 1,3-diphenylbenzotriazinone 6 regioselectively at C-6, while halogenating agents reacted exclusively at C-8. Furthermore, 8-iodo-1,3-diphenylbenzo[e][1,2,4]triazin-7(1H)-one (32) undergoes palladium-catalysed Suzuki-Miyaura and Stille coupling reactions to give 8-aryl- or heteroaryl-substituted benzotriazinones. By combining both the C-6 and C-8 chemistries 1,3,6,8-tetraphenylbenzo[e][1,2,4]triazin-7(1H)-one (42) and 1,3-diphenyl-6,8-di(thien-2-yl)-benzo[e][1,2,4]triazin-7(1H)-one (43) can be prepared. All new compounds are fully characterized.     

Diazaborolyl-boryl push-pull systems with ethynylene-arylene bridges as 'turn-on' fluoride sensors

Two linear π-conjugated systems with 1,3-diethyl-1,3,2-benzodiazaborolyl [C(6)H(4)(NEt)(2)B-] as a donor group and dimesitylboryl (-BMes(2)) as acceptor were synthesised with -ethynylene-phenylene- (-C[triple bond, length as m-dash]C-1,4-C(6)H(4)-, 3) and -ethynylene-thiophene- (-C[triple bond, length as m-dash]C-2,5-C(4)H(2)S-12) bridges between the boron atoms. An assembly (20) consisting of two diazaborolyl-ethynylene-phenylene-boryl units, [C(6)H(4)(NCy)(N')B-C[triple bond, length as m-dash]C-1,4-C(6)H(4)-BMes(2)] joined via a 1,4-phenylene unit at the nitrogen atoms (N') of the diazaborolyl units was also synthesised. The three push-pull systems, 3, 12 and 20, form salts on fluoride addition with the BMes(2) groups converted into (BMes(2)F)(-) anions. The molecular structures of 3, 12 and (NBu(4))(12·F) were elucidated by X-ray diffraction analyses. The borylated systems 3, 12 and 20 show intense blue luminescence in cyclohexane with quantum yields (Φ(fl)) of 0.99, 0.44 and 0.94, respectively, but weak blue-green luminescence in tetrahydrofuran (Φ(fl) = 0.02-0.05). The charge transfer nature of these transitions is supported by TD-DFT computations with the CAM-B3LYP functional. Addition of tetrabutylammonium fluoride to tetrahydrofuran solutions of 3 and 20 resulted in strong violet-blue luminescence with emission intensities up to 46 times more than the emission intensities observed prior to fluoride addition. Compounds 3 and 20 are demonstrated here as remarkable 'turn-on' fluoride sensors in tetrahydrofuran solutions.     

Syntheses of 3-acetoacetylaminobenzo[b]furan derivatives having cysteinyl leukotriene 2 receptor antagonistic activity

Novel 3-acetoacetylaminobenzo[b]furan derivatives having a modified triene system at the 3-position were synthesized starting with 3-aminobenzo[b]furans. The enol isomers, 3-[(3-hydroxybut-2-enonyl)amino]benzo[b]furans (), of the 3-acetoacetylaminobenzo[b]furans were obtained as stable isomers owing to formation of a hydrogen bonding between the enol hydroxyl group and the amidocarbonyl group. The planarity of the C-2 substituent through the C-3 side chain suggested the existence of a modified conjugational triene system in the enol compound. Cysteinyl leukotriene 1 and 2 receptor antagonistic activities for these compounds were evaluated. 2-(4-Cyanobenzoyl or ethoxycarbonyl)-3-[(2-cyano-3-hydroxybut-2-enonyl)amino]benzo[b]furans (, ) were moderately active.     

Macropolyhedral boron-containing cluster chemistry. The reaction of B16H20 and B14H18 with [PtMe2(PMe2Ph)2] to give [(PMe2Ph)2PtB16H17Me] and [(PMe2Ph)2PtB14H16

Structurally characterised 17-vertex [(PMe2Ph)2PtB16H17Me] 3 is obtained, albeit in low yield, by platination of 16-vertex B16H20 1 using [PtMe2(PMe2Ph)2] under mild conditions. Platination has occurred on the {B10} subcluster of 1, interesting in that B16H20 itself deprotonates on the {B8} subcluster: the reference 16-vertex [B16H19]- anion 1a, prepared by deprotonation of 1 with 1,8-bis(dimethylamino)naphthalene, is also structurally characterised. [PtMe2(PMe2Ph)2] with 14-vertex B14H18 2 similarly gives a low yield of 15-vertex [(PMe2Ph)2PtB14H16] 5, of formulation and structure substantiated by DFT calculations.     

Synthesis of 3,4-Biheterylthieno[2,3-b]-thiophenes. Part I: Synthesis of 3,4-Bi(1′,3′,4′-thiadiazolyl-, s-triazolyl-,1′,3′,4′-thiadiazinyl-, 1′,3′,4′-triazinyl-, thiazolyl-, 1′,3′-thiazinyl- and primidinyl)-thieno[2,3-b]thiophenes

3,4-Diamino-2,5-dicarbethoxythieno[2,3-b]thiophene ( 1 ) was allowed to react with NaNO 2 and active methylenes to afford the corresponding azo compounds 2a-c . Hydrazonyl chloride 2a was treated with carbon disulfide, phenyl isothiocyanate, benzonitrile, benzyl cyanide, malononitrile, benzalaniline, ethyl mercaptoacetate, and ethyl glycinate to give 1,3,4-thiadiazolyl-, s-triazolyl-, 1,3,4-thiadiazinyl-, 1,3,4-triazinylthieno[2,3-b]thiophenes 3-6 respectively. The reaction of 2b,c with urea, thiourea, and guanidine afforded pyrimidinyl- and thiazinylazothieno [2,3-b]thiophenes 7-10 respectively. Bithiazolylthieno[2,3-b]thiophenes 11 and 13 were synthesized by treating compound 1 with CS 2 along with halo compounds. The addition of S,S-, N,S-, and N,O-acetals to the Schiff base 14 afforded compounds 15-17 respectively.     

Synthesis and cytotoxic activity of benzophenanthrolinone analogues of acronycine

Condensation of either 2-bromobenzoic acid (4) or 2-chloro-3-nitrobenzoic acid (5) with suitable aminoquinolines 6-8 afforded phenylquinolylamines 9-13. Acid mediated cyclization gave the corresponding 12H-benzo[b][1,7]phenanthrolin-7-ones 14 and 15, and 12H-benzo[b][1,10]phenanthrolin-7-ones 16-18. Compounds 14, 16, and 17 were subsequently N-methylated to 6-demethoxyacronycine and acronycine analogues 19-21, whereas reduction of the aromatic nitro group of 18 gave the amino derivative 22. Unsubstituted 12H-benzo[b][1,10]phenanthrolin-7-ones 16, 17, 20, and 21 were devoid of significant cytotoxic activity, whereas 18 and 22, bearing a nitrogen substituent at position 11, were significantly active. Unsubstituted 12H-benzo[b][1,7]phenanthrolin-7-ones 14 and 19, which include a pyridine nitrogen in the same 4-position as the pyran oxygen of acronycine exhibited cytotoxic activities within the same range of magnitude as acronycine itself.     

Stereochemistry of NaBH4 reduction of a 19-carbonyl group of 3-deoxy androgens. Synthesis of [19S-3H]- and [19R-3H]-labeled aromatase inhibitors having a 19-hydroxy group

To study the stereochemical aspects of the aromatase reaction of androst-4-en-17-one (1) and its 5-ene isomer 4, competitive inhibitors of aromatase, the [19S-(3)H]- and [19R-(3)H]-labeled 19-hydroxy derivatives 2 and 5, were synthesized through NaB(3)H(4) reduction of the corresponding 19-aldehydes 3 and 6 as a key reaction. The hitherto unknown stereochemistry of the NaB(3)H(4) reduction was established based on the deuterium-labeling experiments with NaB(2)H(4). A comparison of (1)H-NMR spectra of the NaB(2)H(4) reduction products of 19-als 3 and 6 with those of the respective authentic steroids revealed that the ratios of 19S-(2)H to 19R-(2)H were 90 : 10 for the 4-ene steroid 2 and 70 : 30 for the 5-ene isomer 5, respectively. Jones oxidation of the [19S-(2)H]19-ols, followed by the non-labeled NaBH(4) reduction, gave the corresponding [19R-(2)H]19-ols 2 and 5 (R-(2)H : S-(2)H=90 : 10 for steroid 2 and 70 : 30 for steroid 5). The stereoselectively (3)H-labeled compounds 2 and 5 were similarly obtained in these sequences.     

2]Catenane assembly from calix[4]arene crown ethers [In Process Citation

A variety of novel calix[4]arene-incorporating crown ethers with or without intramolecular hydrogen bonding have been prepared by two efficient methods and utilized as donor rings to assemble calix[4]arene [2]catenanes based on pi-stacking interaction between hydroquinone and bipyridinium units. Treatment of calix[4]arene crown ethers 4, 10a, or 10b, whose cone conformation was fixed by intramolecular hydrogen bonding within the calix[4]arene moiety, with dicationic salt 15 x 2PF6 and dibromide 16 afforded the corresponding [2]catenanes 17a x 4PF6, 17b x 4PF6, and 17c x 4PF6 in 20%, 53%, and 55% yields, respectively, whereas from the reactions of 15 x 2PF6 and dibromide 16 in the presence of conformationally flexible 11 or 12 with a cone conformation kept by two propyl groups, [2]catenanes 18 x 4PF6 and 19 x 4PF6 were obtained in 12% and 6% yields. [2]Catenanes 21a x 4Cl, 21b x 4Cl, and 21c x 4Cl, incorporating calix[4]arene in both the donor and acceptor rings, were also successfully assembled from 10a or 10b, 16, and dicationic salts 20a x 2PF6 or 20b x 2PF6. The dynamic 1H NMR and absorption spectra of the [2]catenanes have been investigated, which revealed a strongest donor-acceptor interaction in 17a x 4PF6 and that the cone [2]catenanes 17a-c x 4PF6 can isomerize to the partial cone isomer at high temperature. The difference of the dynamic properties of these catenanes was discussed. The results demonstrate that catenation is one new general method to change the conformational distributions of calix[4]arenes.     

Syntheses, structures, and reactivities of heteroleptic magnesium amide thiolates

The syntheses and characterizations of a family of novel heteroleptic magnesium amide thiolates are presented. The compounds are synthesized by ligand redistribution chemistry involving reactions of equimolar amounts of magnesium amides and magnesium thiolates. Utilization of the smaller thiolates [Mg(SPh)2]n and [Mg(S-2,4,6-iPr3C6H2)2]n results in the isolation of dimeric species, [Mg(THF)(N(SiMe3)2)(mu-SR)]2 (R = Ph (1), 2,4,6-iPr3C6H2 (2)), with four-coordinate metal centers and bridging thiolate functions. The sterically more encumbered thiolate S-2,4,6-tBu3C6H2 induces the formation of the four-coordinate, monomeric species Mg(THF)2(N(SiMe3)2)(S-2,4,6-tBu3C6H2) (3)). Careful choice of reaction conditions allows the successful syntheses of pure heteroleptic compounds; however, it remains difficult to obtain the compounds in high yields, since a tendency toward product symmetrization and ligand redistribution under re-formation of the starting materials is prevalent. One of these symmetrized products is also included in this report: the dimeric, four-coordinate magnesium thiolate [Mg-(THF)(S-2,4,6-tBu3C6H2)(mu-S-2,4,6-tBu3C6H2)]2 (4), isolated as the product of the reaction between [Mg-(N(SiMe3)2)2]2 and Mg(THF)2(S-2,4,6-tBu3C6H2)2. All compounds were characterized by NMR and IR spectroscopy, elemental analyses, and X-ray crystallography. Crystal data obtained with Mo K alpha (lambda = 0.710 73 A) radiation are as follows. 1: C16H31MgNOSSi2, a = 11.2100(1) A, b = 17.4512(3) A, c = 11.2999(2) A, beta = 97.952(1) degrees, V = 2189.32(6) A3, Z = 4, monoclinic, space group P2(1)/n, R1 (all data) = 0.0934. 2: C25H49MgNOSSi2, a = 11.1691(1) A, b = 11.0578(1) A, c = 26.0671(4) A, beta = 99.906(1) degrees, V = 3171.44(6) A3, Z = 4, monoclinic, space group P2(1)/c, R1 (all data) = 0.0557. 3: C36H71MgNO3SSi2, a = 42.8293(16) A, b = 10.9737(5) A, c = 16.8305(7) A, beta = 98.755(3) degrees, V = 7818.1(6) A3, Z = 8, monoclinic, space group C2/c, R1 (all data) = 0.1331. 4: C80H132Mg2O2S4, a = 18.8806(2) A, b = 19.3850(2) A, c = 27.3012(4) A, beta = 97.250(1) degrees, V = 9912.4(2) A3, Z = 4, monoclinic, space group P2(1)/n, R1 (all data) = 0.1023.     

Strained, stable 2-aza-1-phosphabicyclo[n.1.0]alkane and -alkene Fe(CO)4 complexes with dynamic phosphinidene behavior

The synthesis of highly strained bicyclic phosphirane and phosphirene iron-tetracarbonyl complexes, that is, complexes with 2-aza-1-phosphabicyclo[n.1.0]alkanes and -alkenes (n = 3-5), is explored by using intramolecular cycloaddition of an in situ generated electrophilic phosphinidene complex, [R(iPr)NP=Fe(CO)(4)], to its C=C- and C[triple chemical bond]C-containing R substituent. Saturated bicyclic complexes 7 a-c with n = 4-2 are remarkably stable, as illustrated by the X-ray crystal structure for 7 b (n=3), yet all readily undergo retroaddition to react with phenylacetylene. Shuttling of the phosphinidene iron complex between two equivalent C=C groups is demonstrated for a 1-butene-substituted 2-aza-1-phosphabicyclo[3.1.0]hexane by selective (1)H NMR magnetization transfer from the phosphirane protons to the olefinic protons. Even the more strained unsaturated bicycles 17 a,b (n = 4,3) are surprisingly stable as illustrated by the X-ray crystal structure for 17 a (n = 4), but the smaller phosphabicyclo[3.1.0]hex-5-ene (17 c, n = 2) dimerizes to tricyclic 19 with a unique ten-membered heterocyclic ring; an X-ray crystal structure is reported. Like their saturated analogues also the bicyclic phosphirenes readily undergo retroaddition as shown by the reaction of their phosphinidene iron moiety with phenylacetylene.     

Stereochemistry of the macrocyclization and elimination steps in taxadiene biosynthesis through deuterium labeling

Taxadiene synthase catalyzes the cyclization of (E,E,E)-geranylgeranyl diphosphate (GGPP) to taxa-4(5),11(12)-diene (Scheme 1, 5 --> 2) as the first committed step of Taxol biosynthesis. Deuterated GGPPs labeled stereospecifically at C-1, C-4, and C-16 were synthesized and incubated with recombinant taxadiene synthase from Taxus brevifolia to elucidate the stereochemistry of the cyclization reaction at these positions. The deuterium-labeled taxadienes obtained from (R)-[1-(2H1)]-, (S)-[1-(2H1)]-, and [16,16,16-(2H3)]GGPPs (9, 10, and 23b) were established to have deuterium in the 2alpha and 2beta CH2 and 16CH3 positions, respectively, by high-field 1H NMR spectroscopy (eqs 1-3). Incubation of (R)-[4-(2H1)]GGPP (17) with the recombinant enzyme gave a 10:10:80 mixture of [5beta-(2H1)]taxa-3(4),11(12)-diene, [5beta-(2H1)]taxa-4(20),11(12)-diene, and unlabeled taxa-4(5),11(12)-diene according to GC/MS analyses of the products (eq 4). It follows that C-1 of GGPP underwent inversion of configuration, that the A ring cyclization occurs on the si face of C15, and that the terminating proton abstraction removes H5beta from the final taxenyl carbocation intermediate. Thus, the C1-C14 and C15-C10 bonds are formed on the opposite faces of the 14,15 double bond of the substrate, i.e., overall anti electrophilic addition. The implications of these findings for the mechanism of the cyclization and rearrangement are discussed.     

Zinc(II), cadmium(II), mercury(II), and ethylmercury(II) complexes of phosphinothiol ligands

Neutral zinc, cadmium, mercury(II), and ethylmercury(II) complexes of a series of phosphinothiol ligands, PhnP(C6H3(SH-2)(R-3))3-n (n = 1, 2; R = H, SiMe3) have been synthesized and characterized by IR and NMR ((1)H, (13)C, and (31)P) spectroscopy, FAB mass spectrometry, and X-ray structural analysis. The compounds [Zn{PhP(C6H4S-2)2}] (1) and [Cd{Ph2PC6H4S-2}2] (2) have been synthesized by electrochemical oxidation of anodic metal (zinc or cadmium) in an acetonitrile solution of the appropriate ligand. The presence of pyridine in the electrolytic cell affords the mixed complexes [Zn{PhP(C6H4S-2)2}(py)] (3) and [Cd{PhP(C6H4S-2)2}(py)] (4). [Hg{Ph2PC6H4S-2}2] (5) and [Hg{Ph2PC6H3(S-2)(SiMe3-3)}2] (6) were obtained by the addition of the appropriate ligand to a solution of mercury(II) acetate in methanol in the presence of triethylamine. [EtHg{Ph2PC6H4S-2}] (7), [EtHg{Ph2P(O)C6H3(S-2)(SiMe3-3)}] (8), [{EtHg}2{PhP(C6H4S-2)2}] (9), and [{EtHg}2{PhP(C6H3(S-2)(SiMe3-3))2}] (10) were obtained by reaction of ethylmercury(II) chloride with the corresponding ligand in methanol. In addition, in the reactions of EtHgCl with Ph2PC6H4SH-2 and with the potentially tridentate ligand PhP(C6H3(SH-2)(SiMe3-3)) 2, cleavage of the Hg-C bond was observed with the formation of [Hg{Ph2PC6H4S-2}2] (5) and [Hg(EtHg) 2{PhP(O)(C6H3(S-2)(SiMe3-3))2}2] (11), respectively, and the corresponding hydrocarbon. The crystal structures of [Zn3{PhP(C6H4S-2)2}2{PhP(O)(C6H4S-2)2}] (1*), [Cd2{Ph2PC6H4S-2}3{Ph2P(O)C6H4S-2}] (2*), 3, 5, 6, [EtHg{Ph2P(O)C6H4S-2}] (7*), 8, 9, [{EtHg}2{PhP(O)(C6H3(S-2)(SiMe3-3))2}] (10*), and 11 are discussed. The molecular structures of 1, 2, 4, 7, and 10 have also been studied by means of density functional theory (DFT) calculations.     

Utility of isolated hepatocytes and radio-HPLC-MSn for the analysis of the metabolic fate of 19-nortestosterone laurate in cattle.

The metabolic fate of 19-nortestosterone laurate in cattle was investigated to evaluate target analyte(s) appropriate to surveillance for illicit use as a growth promoting agent. Bovine hepatocytes were incubated with either [3H]19-nortestosterone laurate (19-NTL; 4-estren-17 beta-laurate-3-one) or [3H]19-nortestosterone (19-NT; 4-estren-17 beta-ol-3-one; nandrolone). Hepatocyte medium was extracted with solid phase C18 media and analysed by narrow bore radio-HPLC-MSn (LCQ, Finnigan) to evaluate the structure of metabolites of 19-NTL and 19-NT. Radio-HPLC of hepatocyte medium extracts following incubation with [3H]19-NTL confirmed that the first step of biotransformation in liver was hydrolysis of the fatty acid ester to release [3H]19-NT, which, in turn, was converted into a range of metabolites of diverse polarity. Hydrolysis of hepatocyte medium extracts with beta-glucuronidase (Helix pomatia) indicated that some of these metabolites were glucuronide or sulfate conjugates. Structural analysis of unconjugated metabolities by positive-ion atmospheric pressure chemical ionisation MS2 and comparison with available reference preparations indicated biotransformation of 19-NT to 4-estren-17 alpha-ol-3-one, 4-estren-3, 17-dione (major metabolite after 1 h), n-hydroxy-4-estren-3, 17-dione, n-hydroxy-4-estren-17-ol-3-one, 5 beta-estran-3 alpha-ol-17-one (noretiocholanolone) and 5 beta-estran-3 alpha, 17 beta-ol (major metabolite after 4 h). Conjugated metabolites were analysed by electrospray ionization, which revealed the presence of glucuronide conjugates of alpha-(trace) and beta-epimers of 19-NT, n-hydroxy-4-estren-3, 17-dione, n-hydroxy-4-estren-17-ol-3-one and 5 beta-estran-3 alpha, 17 beta-diol. These studies provide a clear indication of the route of hepatic metabolism in the bovine, which may now be readily substantiated by reference to samples, such as urine or bile, derived from animals treated with unlabelled 19-NTL.     

Alpha-amino acids with metallocenyl side chains

A straightforward method for the synthesis of enantiomerically pure bis(valine)metallocenes is presented. Derivatives of lithium cyclopentadienylvaline 1a, b were obtained by addition of the (R)- or (S)-Sch?llkopf reagents to 6,6-dimethylfulvene as single enantiomers and gave with FeCl2 or [RuCl2(dmso)4] the chiral metallocenes [Fe[C5H4-CMe2-[C4H2N2(OMe)2iPr]]2] (2a, b) and [Ru[C5H4-CMe2-[C4H2N2(OMe)2iPr]]2] (3a, b). Complex 2b was hydrolyzed to the ferrocenylene-bis(valine-methylester) [[Fe[C5H4-CMe2-CH(NH3+)COOMe]2]2+(Cl-)2] (7) without racemization. Complex 7 could be used as ligand and was treated with [[Cp*IrCl2]2] to afford [Fe[C5H4-CMe2-CH(COOMe)(NH2-IrCp*Cl2)]2] (10). The reactions of 1 with CoCl2, [Re(CO)5Br], [[(cod)RhCl2]2] (cod= 1,5-cyclooctadiene) or [Cp*MCl3] (M= Ti, Zr) gave the cyclopentadienyl complexes [[Co[C5H4-CMe2-[C4H2N2(OMe)2iPr]]2]+ I-] (11) and [Re[C5H4-CMe2-[C4H2N2(OMe)2iPr]](CO)3] (13), [(C8H12)Rh[C5H4-CMe2-[C4H2N2(OMe)2(iPr)]]] (14). [[Rh[C5H4-CMe2-[C4H2N2(OMe)2(iPr)]]I]2(mu-I)2] (15), [Cp*Cl2Ti-[C5H4-CMe2-[C4H2N2(OMe)2(iPr)]]] (16), and [Cp*Cl2Zr[C5H4-CMe2-[C4H2N2(OMe)2(iPr)]]] (17), with chiral valine derivatives as substituents on the cyclopentadienyl ring and with excellent diastereoselectivities. Also the Seebach reagent (Boc-BMI) or O'Donnell reagent could be added to 6,6-dimethylfulvene to give the lithium cyclopentadienides Li[C5H4-CMe2-[C3H2(tBu)(N-Boc)(NMe)O]] (18) and Li[C5H4-CMe2-CH(NCPh2)(COOEt)] (21), which formed the ferrocene derivatives [Fe[C5H4-CMe2-[C3H2(tBu)(N-Boc)(NMe)O]]2] (19) and [Fe[C5H4-CMe2-CH(NCPh2)(COOEt)]2] (22). The stable cobaltocinium cation in 11 and the complex 19 could be hydrolyzed to the metallocenes 12 and [Fe(C5H4-CMe2-CH(NH3+)(COO-)]2] (20) with two valines in the 1,1'-position. The structures of 2a, b, 11, 15, and 16 were determined by X-ray diffraction and confirm the diastereomeric purity of the compounds.     

Biosynthetic studies on the cyclopentane ring formation of allosamizoline, an aminocyclitol component of the chitinase inhibitor allosamidin

Allosamizoline (1) is an aminocyclitol component of allosamidin, a Streptomyces metabolite, and has a cyclopentane ring originated from D-glucosamine. Biosynthesis of the cyclopentane ring was studied by feeding experiments with a variety of deuterium-labeled glucosamine and glucose. In the feeding experiments with [3-(2)H]- and [4-(2)H]-D-glucosamine and [1-(2)H]-D-glucose, deuterium was incorporated into C-3, C-4, and C-1 of 1, respectively. On the other hand, feeding experiments with [5-(2)H]- and [6,6-(2)H(2)]-D-glucosamine showed that deuterium on C-5 and one of the two deuterium atoms on C-6 of glucosamine were lost during the cyclopentane ring formation of 1. In the feeding experiments with (6R)- and (6S)-[6-(2)H(1)]-D-glucose, the (6R)-deuterium of glucose was incorporated into the proS position on C-6 of 1, but the (6S)-deuterium of glucose was not incorporated into 1. These results suggested that an intermediate with a 6-aldehyde group is involved in the biosynthesis of the cyclopentane ring moiety of 1 and overall inversion of stereochemistry of the C-6 methylene group occurred by stereospecific oxidation and reduction on C-6 during the formation of 1. The 6-aldehyde intermediate may play a key role in the biosynthetic step(s) of cyclization to form the cyclopentane ring and/or deoxygenation at C-5.