Efficient syntheses of benzyl and n‐octyl [1,3‐13C2]acetoacetates,and their application to syntheses of 13C‐labelled pyrrole and 13C‐labelled hymecromone

Benzyl [1‐¹³C]acetate (2a) was prepared via esterification of sodium [1‐¹³C]acetate (1) with benzyl bromide in the presence of 18‐crown‐6‐ether in 97% yield. n‐Octyl [1‐¹³C]acetate (2b) was rapidly obtained by microwave irradiation of 1‐bromooctane and potassium [1‐¹³C]acetate (obtained by salt exchange of 1) absorbed on Al₂O₃ in 82% yield. Solvent‐free Claisen condensation of benzyl or n‐octyl [1‐¹³C]acetate (2a or 2b) in the presence of potassium tert‐butoxide efficiently gave benzyl or n‐octyl [1,3‐¹³C₂]acetoacetate (3a or 3b) in 51 or 68% yield, respectively. Dibenzyl 2,4‐dimethyl[2,4‐¹³C₂]pyrrole‐3,5‐di[¹³C]carboxylate (4) was synthesized from benzyl [1,3‐¹³C₂]acetoacetate (3a) in 54% yield. [2,4‐¹³C₂]Hymecromone (6) (7‐hydroxy‐4‐methyl[2,4‐¹³C₂]coumarin) was obtained from n‐octyl [1,3‐¹³C₂]acetoacetate (3b) and 1,3‐benzenediol (5) in 73% yield. Copyright © 2008 John Wiley & Sons, Ltd.     

Synthesis of C‐14 and C‐13, H‐2‐labeled IKK inhibitor: [14C] and [13C4,D3]‐N‐(6‐chloro‐7‐methoxy‐9H‐pyrido[3,4‐b]indol‐8‐yl)‐2‐methyl‐3‐pyridinecarboxamide

[¹⁴C]‐N‐(6‐Chloro‐7‐methoxy‐9H‐pyrido [3,4‐b]indol‐8‐yl)‐2‐methyl‐3‐pyridinecarboxamide (5B ), an IKK inhibitor, was synthesized from [¹⁴C]‐barium carbonate in two steps in an overall radiochemical yield of 41%. The intermediate, [carboxyl‐¹⁴C]‐2‐methylnicotinic acid, was prepared by the lithiation and carbonation of 3‐bromo‐2‐methylpyridine. [¹³C₄,D₃]‐N‐(6‐chloro‐7‐methoxy‐9H‐pyrido [3,4‐b]indol‐8‐yl)‐2‐methyl‐3‐pyridinecarboxamide (5C ) was synthesized from [1,2,3,4‐¹³C₄]‐ethyl acetoacetate and [D₄]‐methanol in six steps in an overall yield of 2%. [¹³C₄]‐2‐methylnicotic acid, was prepared by condensation of [¹³C₄]‐ethyl 3‐aminocrotonate and acrolein, followed by hydrolysis with lithium hydroxide. Copyright © 2006 John Wiley & Sons, Ltd.     

Synthesis of deuterium and C‐13‐labelled ethyl glycolate and their subsequent use in the synthesis of labelled analogues of the DNA adduct O6‐carboxymethyl‐2′‐deoxyguanosine

The adduct O⁶‐carboxymethyl‐2′‐deoxyguanosine (O⁶CMdG) is of importance as it has been previously linked to high red meat diet in humans, and as yet, a liquid chromatography‐mass spectrometry (LC‐MS) method has not been developed due to lack of appropriate standards. The synthesis of the deuterated and C‐13 analogues required the use of [²H₂]‐ and [¹³C₂]ethyl glycolate to label the carboxymethyl moiety of O⁶CMdG. [²H₂]Ethyl glycolate was synthesised via acid hydrolysis of ethyl diazoacetate using deuterated solvents (59% yield), whilst [¹³C₂]ethyl glycolate was synthesised from [¹³C₂]glycine in a three‐step procedure (35% yield). The labelled ethyl glycolates were then used to synthesise [²H₂]‐ and [¹³C₂]O⁶CMdG for future use as internal standards in the LC‐MS analysis of biological samples. Copyright © 2011 John Wiley & Sons, Ltd.     

An overview of radio or stable isotope‐labeled cis‐neonicotinoid analogs

To support the metabolism and toxicology study of cis‐neonicotinoids, radio or stable isotope was introduced into different sites of the key intermediate 2‐chloro‐5‐((2‐(nitromethylene)imidazolidin‐1‐yl)methyl)pyridine (6‐Cl‐PMNI). [³H₂]‐ and [¹⁴C]‐label were successively prepared from initial materials NaB³H₄ and [¹⁴C]‐nitromethane, respectively. Similarly, [D₂]‐6‐Cl‐PMNI was prepared from NaBD₄ in four steps, with 52.6% overall isotopic yield, and dual‐labeled [D₂, ¹³C]‐target was obtained from NaBD₄ and [¹³C]‐nitromethane, affording overall isotopic yield of 42.5%. Moreover, [¹⁴C₂] was introduced from [U‐¹⁴C]‐ethylenediamine dihydrochloride in three steps, with a 58.3% overall chemical yield. Finally, typical labeled cis‐neonicotinoids paichongding and cycloxaprid were prepared and characterized. The methods were proved to have good generality in the synthesis of other cis‐neonicotinoids, and all results would be useful in metabolism studies of new cis‐neonicotinoids. Copyright © 2012 John Wiley & Sons, Ltd.     

Synthesis of a highly potent leukocyte function‐associated antigen‐1 antagonist and its metabolite labeled with stable isotopes and carbon‐14, part 2

(S)‐2‐[(R)‐7‐(3,5‐Dichlorophenyl)‐5‐methyl‐6‐oxo‐5‐(4‐trifluoromethoxybenzyl)‐6,7‐dihydro‐5H‐imidazo[1,2‐a]imidazole‐3‐sulfonylamino]‐proprionamide (1), a potent lymphocyte function‐associated antigen‐1 antagonist and its sulfonamide metabolite (2) labeled with stable isotopes and carbon‐14 were prepared for Drug Metabolism and PharmacoKinetics and other studies. A long linear route was used to prepare [¹³C₂, ²H₃]‐(1) using [3,3,3‐²H]‐D‐alanine and [¹³C₂]‐glycine in 15 steps and 2.5% overall yield. With the availability of [¹³C₆]‐3,5‐dichloroaniline, the sulfonamide [¹³C₆]‐(2) was prepared in 12 steps and in 5.6% overall yield. For the carbon‐14 synthesis, a six‐step synthesis gave both compounds [¹⁴C]‐(1) and [¹⁴C]‐(2) from the common sulfonyl chloride intermediate [¹⁴C]‐(15) in 18% and 4% radiochemical yields and specific activities of 44 and 40.5 mCi/mmol, respectively. Copyright © 2011 John Wiley & Sons, Ltd.     

Synthesis of 13C and 15N labeled 2,4‐dinitroanisole

Syntheses of [¹³C₆]‐2,4‐dinitroanisole (ring‐¹³C₆) from [¹³C₆]‐anisole (ring‐¹³C₆) and [¹⁵N₂]‐2,4‐dinitroanisole from anisole using in situ generated acetyl nitrate and [¹⁵N]‐acetyl nitrate, respectively, are described. Treatment of [¹³C₆]‐anisole (ring‐¹³C₆) with acetyl nitrate generated in 100% HNO₃ gave [¹³C₆]‐2,4‐dinitroanisole (ring‐¹³C₆) in 83% yield. Treatment of anisole with [¹⁵N]‐acetyl nitrate generated in 10 N [¹⁵N]‐HNO₃ gave [¹⁵N₂]‐2,4‐dinitroanisole in 44% yield after two cycles of nitration. Byproducts in the latter reaction included [¹⁵N]‐2‐nitroanisole and [¹⁵N]‐4‐nitroanisole.     

Synthesis of benzoxazole‐[phenyl‐13C6] by directed ortho‐metalation chemistry

A new method for the preparation of benzoxazole‐[phenyl‐¹³C₆] ( 1 ) starting from aniline‐[¹³C₆] ( 4 ) has been developed involving directed ortho‐metalation (DoM) chemistry. The synthesis comprises four steps and an overall yield of 39% was obtained. Copyright © 2003 John Wiley & Sons, Ltd.     

Carbon‐14‐and carbon‐13‐labeled phosphoric acid [2‐4‐(4‐cyanophenyl)‐thiazol‐2‐yl]‐(2,4‐difluorophenyl)‐1‐[1,2,4]triazol‐4‐yl‐methylpropoxymethyl] monoester dilysine salt,a prodrug of ravuconazole

4‐Bromobenzoic acid [carboxyl‐¹⁴C] and 4‐(2‐bromoacetyl) [Ar‐¹³C₆]benzonitrile were transformed into the title compounds containing [ring¹⁴C‐thiazol‐4‐yl] and [Ar‐¹³C₆‐benzonitrile]. ¹⁴C‐Ravuconazole was prepared in 37% yield and Purity >99%. ¹³C₆‐Ravuconazole was made in 56% overall yield and Purity of >98%. Each labeled compound was converted by additional reaction steps to the corresponding labeled prodrug. Copyright © 2011 John Wiley & Sons, Ltd     

Synthesis of highly potent lymphocyte function‐associated antigen‐1 antagonists labeled with carbon‐14 and with stable isotopes,part 3

The drug candidates ( 2 ) and ( 3 ) are highly potent LFA‐1 inhibitors. They were efficiently prepared labeled with carbon‐14 using a palladium‐catalyzed carboxylation of an iodo‐precursor ( 5 ) and sodium formate‐¹⁴C to afford acid [¹⁴C]‐( 6 ), which was coupled via an amide bond to chiral amines ( 7 ) and ( 8 ) in 52% and 48% overall yield, respectively, and with specific activities higher than 56 mCi/mmol and radiochemical purities of 99%. For stable isotopes synthesis, the amine [²H₈]‐( 7 ) was synthesized in three steps from 2‐cyanopyridine‐²H₄ using Kulinkovich‐Szymonik aminocyclopropanation, followed by coupling to L ‐alanine‐2,3,3,3‐²H₄‐N‐t‐BOC, and then removal of the BOC‐protecting group. Amide bond formation with acid ( 6 ) gave [²H₈]‐( 2 ) in 36% overall yield. The amine [¹³C₄,¹⁵N]‐( 8 ) was obtained in two steps using L‐threonine‐¹⁴C₄,¹⁵N and then coupled to acid [¹³C]‐( 6 ) to give [¹³C₅,¹⁵N]‐( 3 ) in 56% overall yield.     

Synthesis of deleobuvir,a potent hepatitis C virus polymerase inhibitor,and its major metabolites labeled with carbon‐13 and carbon‐14

Deleobuvir, (2E)‐3‐(2‐{1‐[2‐(5‐bromopyrimidin‐2‐yl)‐3‐cyclopentyl‐1‐methyl‐1H‐indole‐6‐carboxamido]cyclobutyl}‐1‐methyl‐1H‐benzimidazol‐6‐yl)prop‐2‐enoic acid (1), is a non‐nucleoside, potent, and selective inhibitor of hepatitis C virus NS5B polymerase. Herein, we describe the detailed synthesis of this compound labeled with carbon‐13 and carbon‐14. The synthesis of its three major metabolites, namely, the reduced double bond metabolite (2) and the acyl glucuronide derivatives of (1) and (2), is also reported. Aniline‐¹³C₆ was the starting material to prepare butyl (E)‐3‐(3‐methylamino‐4‐nitrophenyl‐¹³C₆)acrylate [¹³C₆]‐(11) in six steps. This intermediate was then used to obtain [¹³C₆]‐(1) and [¹³C₆]‐(2) in five and four more steps, respectively. For the radioactive synthesis, potassium cyanide‐¹⁴C was used to prepare 1‐cylobutylaminoacid [¹⁴C]‐(23) via Buchrer–Bergs reaction. The carbonyl chloride of this acid was then used to access both [¹⁴C]‐(1) and [¹⁴C]‐(2) in four steps. The acyl glucuronide derivatives [¹³C₆]‐(3), [¹³C₆]‐(4) and [¹⁴C]‐(3) were synthesized in three steps from the acids [¹³C₆]‐(1), [¹³C₆]‐(2) and [¹⁴C]‐(1) using known procedures.     

Synthesis of two potent glucocorticoid receptor agonists labeled with carbon‐14 and stable isotopes

Two potent glucocorticoid receptor agonists were prepared labeled with carbon‐14 and with stable isotopes to perform drug metabolism, pharmacokinetics, and bioanalytical studies. Carbon‐14 labeled (1) was obtained from an enantiopure alkyne (5) via a Sonogashira coupling to a previously reported 5‐amino‐4‐iodo‐[2‐¹⁴C]pyrimidine [¹⁴C]‐(6), followed by a base‐mediated cyclization (1) in 72% overall radiochemical yield. Carbon‐14 labeled (2) was prepared in five steps employing a key benzoic acid intermediate [¹⁴C]‐(13), which was synthesized in one pot from enolization of trifluoromethylketone (12), followed by bromine–magnesium exchange and then electrophile trapping reaction with [¹⁴C]‐carbon dioxide. A chiral auxiliary (S)‐1‐(4‐methoxyphenyl)ethylamine was then coupled to this acid to give [¹⁴C]‐(15). Propargylation and separation of diastereoisomers by crystallizations gave the desired diastereomer [¹⁴C]‐(17) in 34% yield. Sonogashira coupling to iodopyridine (10) followed by cyclization to the azaindole [¹⁴C]‐(18) and finally removal of the chiral auxiliary gave [¹⁴C]‐(2) in 7% overall yield. For stable isotope syntheses, [¹³C₆]‐(1) was obtained in three steps using [¹³C₄]‐(6) and trimethylsilylacetylene‐[¹³C₂] in 26% yield, while [²H₅]‐(2) was obtained by first preparing the iodopyridine [²H₅]‐(10) in five steps. Then, Sonogashira coupling to chiral alkyne (24) and cyclization gave [²H₅]‐(2) in 42% overall yield.     

Potent and selective inhibitors of 11β‐hydroxysteroid dehydrogenase type 1 labeled with carbon‐13 and carbon‐14

(S )‐6‐(2‐Hydroxy‐2‐methylpropyl)‐3‐((S )‐1‐(4‐(1‐methyl‐2‐oxo‐1,2‐dihydropyridin‐4‐yl)phenyl)ethyl)‐6‐phenyl‐1,3‐oxazinan‐2‐one (1) and (4aR ,9aS )‐1‐(1H‐benzo[d]midazole‐5‐carbonyl)‐2,3,4,4a,9,9a‐hexahydro‐1‐H‐indeno[2,1‐b]pyridine‐6‐carbonitrile hydrochloride (2) are potent and selective inhibitor of 11β‐hydroxysteroid dehydrogenase type 1 enzyme. These 2 drug candidates developed for the treatment of type‐2 diabetes were prepared labeled with carbon‐13 and carbon‐14 to enable drug metabolism, pharmacokinetics, bioanalytical, and other studies. In the carbon‐13 synthesis, benzoic‐¹³C ₆ acid was converted in 7 steps and in 16% overall yield to [¹³C₆]‐(1). Aniline‐¹³C ₆ was converted in 7 steps to 1H‐benzimidazole‐1‐2,3,4,5,6‐¹³C₆‐5‐carboxylic acid and then coupled to a tricyclic chiral indenopiperidine to afford [¹³C₆]‐(2) in 19% overall yield. The carbon‐14 labeled (1) was prepared efficiently in 2 radioactive steps in 41% overall yield from an advanced intermediate using carbon‐14 labeled methyl magnesium iodide and Suzuki‐Miyaura cross coupling via in situ boronate formation. As for the synthesis of [¹⁴C]‐(2), 1H‐benzimidazole‐5‐carboxylic‐¹⁴C acid was first prepared in 4 steps using potassium cyanide‐¹⁴C , then coupled to the chiral indenopiperidine using amide bond formation conditions in 26% overall yield.     

Synthesis of isotope‐labeled HSP90 inhibitor: [13CD3] and [14C]‐TAK‐459

[¹³CD₃]‐TAK‐459 (1A), an HSP90 inhibitor, was synthesized from [¹³CD₃]‐sodium methoxide in three steps in an overall yield of 29%. The key intermediate [¹³CD₃]‐2‐methoxy‐6‐(4,4,5,5‐tetramethyl‐1,3,2‐dioxaborolan‐2‐yl)pyridine was synthesized in two steps from 2,6‐dibromopyridine and stable isotope‐labeled sodium methoxide. [¹⁴C]‐TAK‐459 (1B) was synthesized from [¹⁴C(U)]‐guanidine hydrochloride in five steps in an overall radiochemical yield of 5.4%. The key intermediate, [¹⁴C]‐(R)‐2‐amino‐7‐(2‐bromo‐4‐fluorophenyl)‐4‐methyl‐7,8‐dihydropyrido[4,3‐d]pyrimidin‐5(6H)‐one, was prepared by microwave‐assisted condensation.     

Synthesis of 13C6‐labeled Reyataz™ (BMS‐232632)

An efficient synthesis of ¹³C₆‐labeled Reyataz? (BMS‐232632) is described. ¹³C₆‐labeled Reyataz? was synthesized in eight steps from commercially available [ring‐¹³C₆]‐L ‐phenylalanine. The overall yield of the synthesis was 8% and the purity of the final product was greater than 98%. Copyright © 2005 John Wiley & Sons, Ltd.     

An improved synthesis of isotope labeled 6‐hydroxychlorzoxazone

[¹³C₆]6‐Hydroxychlorzoxazone, a major P450 metabolite of chlorzoxazone, was synthesized from [¹³C₆]benzene in an overall 18% yield. An improved procedure for converting 4‐chloro‐6‐nitrosoresorcinol to 6‐hydroxychlorzoxazone was developed, with a yield more than double that previously reported in the literature. Copyright © 2006 John Wiley & Sons, Ltd.     

Synthesis of C‐13‐labeled atrazine

Atrazine is a long‐lasting herbicide that has been shown to affect hormone levels in amphibians. Using the C‐13 labeled atrazine to detect its residue is effective and essential. This study presents three steps for the synthesis of [¹³C₃]atrazine, which starts from [¹³C]urea, and results in the incorporation of C‐13 atoms at the 1, 3 and 5 positions of the S‐triazine ring of atrazine. The method prepares the product in an overall yield of 57.6% and chemical purity of 98.6%, for use as an internal standard. Copyright © 2013 John Wiley & Sons, Ltd.     

Synthesis of [14C]‐ and [13C6]‐labeled potent HIV non‐nucleoside reverse transcriptase inhibitor

5,11‐Dihydro‐11‐ethyl‐5‐methyl‐8‐{2‐{(1‐oxido‐4‐quinolinyl)oxy}ethyl}‐6H‐dipyrido[3,2‐b:2′,3′‐e][1,4]diazepin‐6‐one, (1), labeled with carbon‐14 in the quinoline–benzene ring, in one of the pyridine rings of the dipyridodiazepinone tricyclic moiety, and in the side chain, was prepared in three different syntheses with specific activities ranging from 44 to 47 mCi/mmol (1.63–1.75 GBq/mmol). In the first synthesis, 5,11‐dihydro‐11‐ethyl‐8‐(2‐hydroxyethyl)‐5‐methyl‐6H‐dipyrido[3,2‐b:2′,3′‐e][1,4]diazepin‐6‐one (2) was coupled to 4‐hydroxyquinoline, [benzene‐¹⁴C(U)]‐, using Mitsunobu's reaction conditions, followed by the oxidation of the quinoline nitrogen with 3‐chloroperoxybenzoic acid to give ([¹⁴C]‐(1a)) in 43% radiochemical yield. Second, 3‐amino‐2‐chloropyridine, [2,6‐¹⁴C]‐, was used to prepare 8‐bromo‐5,11‐dihydro‐11‐ethyl‐5‐methyl‐6H‐dipyrido[3,2‐b:2′,3′‐e][1,4]diazepin‐6‐one (8), and then Stille coupled to allyl(tributyl)tin followed by ozonolysis of the terminal double bond and in situ reduction of the resulting aldehyde to alcohol (10). Mitsunobu etherification and oxidation as seen before gave ([¹⁴C]‐(1b)) in eight steps and in 11% radiochemical yield. Finally, carbon‐14 potassium cyanide was used to prepare isopropyl cyanoacetate (12), which was used to transform bromide (8) to labeled aryl acetic acid (13) under palladium catalysis. Trihydroborane reduction of the acid gave alcohol (14) labeled in the side chain, which was used as described above to prepare ([¹⁴C]‐(1c)) in 4.3% radiochemical yield. The radiochemical purities of these compounds were determined by radio‐HPLC and radio‐TLC to be more than 98%. To prepare [¹³C₆]‐(1), [¹³C₆]‐4‐hydroxyquinoline was prepared from [¹³C₆]‐aniline and then coupled to (2) and oxidized as seen before. Copyright © 2008 John Wiley & Sons, Ltd.     

A gram‐scale synthesis of [3,4‐13C2,1α,7‐2H2]cortisone

A gram‐scale synthesis of [3,4‐¹³C₂,1α,7‐²H₂]cortisone from prednisone was developed. The deuterium atom at the C‐1 position was introduced through a regioselective and stereoselective deuteration of the 1,2‐double bond of the 1,4‐diene‐3‐one using Wilkinson's catalyst. After the oxidative cleavage of the A‐ring, two carbon‐13 atoms were introduced via acetylation of an A‐ring enol lactone with [1,2‐¹³C₂]acetyl chloride. The steroidal A‐ring was then reconstructed to incorporate the carbon‐13 atoms into the C‐3 and C‐4 positions. The deuterium atom at C‐7 was introduced through a regioselective deuteration of the 6,7‐double bond of a 4,6‐diene‐3‐one intermediate using palladium on strontium carbonate. The M + 4 stable isotope labeled cortisone was thus prepared in ca. 4% overall yield. In addition, [3,4‐¹³C₂,1α,7‐²H₂]‐11‐dehydrocorticosterone, [3,4‐¹³C₂,1α,7‐²H₂]cortisol, and [3,4‐¹³C₂,1α,7‐²H₂]corticosterone were also prepared. Copyright © 2011 John Wiley & Sons, Ltd.     

The syntheses of [13C6] and [phenyl‐14C(U)]BMS‐816336, an inhibitor of 11β‐hydroxysteroid dehydrogenase type 1, for type 2 diabetes

Type 2 diabetes is a significant worldwide health problem. To support the development of BMS‐816336 as an inhibitor of 11β‐hydroxysteroid dehydrogenase type 1 for type 2 diabetes, the synthesis of carbon‐14 labeled material was required for use in metabolic profiling. [Phenyl‐¹⁴C(U)]BMS‐816336 was synthesized in 8 steps and 22% radiochemical yield from commercially available [¹⁴C(U)]bromobenzene. The radiochemical purity of [phenyl‐¹⁴C(U)]BMS‐816336 was 100% having a specific activity of 84.4 μCi/mg or 28.8 mCi/mmol for a total of 8.9 mCi. It was also necessary to synthesize [¹³C₆]BMS‐816336 for use as a liquid chromatography/mass spectrometry standard. [¹³C₆]BMS‐816336 was also prepared in 8 labeled steps in 26% yield from [¹³C₆]bromobenzene.     

Synthesis of stable isotope‐labeled metabolites of asenapine

The stable isotope‐labeled synthesis of four of the major metabolites of asenapine is described. The synthesis of [¹³CD₃]‐asenapine N‐oxide proceeded in two synthetic steps. Preparation of [¹³CD₃]‐asenapine 11‐hydroxysulfate and [¹³C₆]‐N‐desmethylasenapine paralleled established synthetic protocols with effective utilization of labeled precursors. The synthesis of [¹³CD₃]‐asenapine N⁺‐glucuronide was achieved in three chemical steps followed by purification.