Studies of hydrogen isotope scrambling during the dehalogenation of aromatic chloro-compounds with deuterium gas over palladium catalysts

Catalytic dehalogenation of aromatic halides using isotopic hydrogen gas is an important strategy for labelling pharmaceuticals, biochemicals, environmental agents and so forth. To extend, improve and further understand this process, studies have been carried out on the scrambling of deuterium isotope with protium during the catalytic deuterodehalogenation of model aryl chlorides using deuterium gas and a palladium on carbon catalyst in tetrahydrofuran solution. The degree of scrambling was greatest with electron-rich chloroarene rings. The tetrahydrofuran solvent and the triethylamine base were not the source of the undesired protium; instead, it arose, substantially, from the water content of the catalyst, though other sources of protium may also be present on the catalyst. Replacement of the Pd/C catalyst with one prepared in situ by reduction of palladium trifluoroacetate with deuterium gas and dispersed upon micronised polytetrafluoroethylene led to much reduced scrambling (typically 0–6% compared with up to 40% for palladium on carbon) and to high atom% abundance, regiospecific labelling. The improved catalytic system now enables efficient polydeuteration via the dehalogenation of polyhalogenated precursors, making the procedure viable for the preparation of MS internal standards and, potentially, for high specific activity tritium labelling.     

Benzylic deuteration of alkylnitroaromatics via amine-base catalysed exchange with deuterium oxide

This paper describes the deuterium-labelling of alkylnitroaromatics by base-catalysed exchange with deuterium oxide. As the alkyl protons alpha to the aromatic ring are the most acidic sites in the molecule, regioselective hydrogen isotope exchange at this benzylic location leads to a regiospecifically deuterated product. The exchange labelling takes place in good yields and with high atom% abundance in the presence of an appropriate nitrogen base. Alkylated 2,4-dinitrobenzenes deuterate at room temperature under catalysis by triethylamine, whilst alkylated 2-nitro- or 4-nitrobenzenes and related mono-nitroaromatics require higher temperatures and catalysis by 1,5-diazobicyclo[4.3.0]non-5-ene (DBN). The labelling reactions require an inert gas atmosphere, but otherwise are simple and high yielding with no obvious byproducts. Those compounds in which the benzylic protons are in an ortho-orientation with respect to the nitro group label somewhat more slowly than the analogues where there is a para relationship. In addition, higher alkyl homologues undergo benzylic deuteration at slower rates than methyl.     

Labelling of anilines,benzylamines and some N‐heterocyclics using cycloocta‐1,5‐dienyliridium(I)‐1,1,1,5,5,5‐hexafluoro‐pentan‐2,4‐dionate and isotopic hydrogen gas in DMF or DMA

A wide range of anilines, benzylamines and some N‐heterocyclics can be ortho‐deuterated at room temperature using deuterium gas and cycloocta‐1,5‐dienyliridium(I)‐1,1,1,5,5,5‐hexafluoropentan‐2,4‐dionate in DMF or DMA. The method is applicable to labelling with tritium. Copyright © 2004 John Wiley & Sons, Ltd.     

Convenient and efficient deuteration of functionalized aromatics with deuterium oxide: catalysis by cycloocta‐1,5‐dienyliridium(I) 1,3‐dionates

Aromatic compounds bearing an ortho‐directing substituent may be deuterated by exchange with deuterium oxide in the presence of a range of cycloocta‐1,5‐dienyliridium(I)1,3‐dionate catalysts. The exchange takes place in several dipolar aprotic solvents and is directly applicable to the deuteration of polar compounds. Isotope incorporation is efficient and regiospecific. The method is applicable to a wide range of ortho‐directing groups some of which are only weak directors for alternative ortho‐labelling approaches. In addition, the application of microwaves enables labelling within minutes even with sub‐stituents which are poor directors. Copyright © 2003 John Wiley & Sons, Ltd.     

Efficient tritiation of the translocator protein (18 kDa) selective ligand DPA‐714

DPA‐714 (N,N‐diethyl‐2‐(2‐(4‐(2‐fluoroethoxy)phenyl)‐5,7‐dimethylpyrazolo[1,5‐a]pyrimidin‐3‐yl)acetamide) is a recently discovered fluorinated ligand of the translocator protein 18 kDa (TSPO). Labelled with the short‐lived positron emitter fluorine‐18, this structure is today the radioligand of reference for in vivo imaging of microglia activation and neuroinflammatory processes with positron emission tomography. In the present work, an isotopically tritium‐labelled version was developed ([³H]DPA‐714), in order to access high resolution in vitro and ex vivo microscopic autoradiography studies, repeated and long‐lasting receptor binding studies and in vivo pharmacokinetic determination at late time points. Briefly, DPA‐714 as reference, and its 3,5‐dibrominated derivative as precursor for labelling, were both prepared from DPA‐713 in nonoptimized 32% (two steps) and 10% (three steps) yields, respectively. Reductive debromination using deuterium gas and Pd/C as catalyst in methanol, performed at the micromolar scale, confirmed the regioselective introduction of two deuterium atoms at the meta positions of the phenyl ring. Tritiodebromination was analogously performed using no‐carrier tritium gas. HPLC purification provided >96% radiochemically pure [³H]DPA‐714 (7 GBq) with a 2.1 TBq/mmol specific radioactivity. Interestingly, additional hydrogen‐for‐tritium exchanges were also observed at the 5‐methyl and 7‐methyl positions of the pyrazolo[1,5‐a]pyrimidine, opening novel perspectives in the labelling of compounds featuring this heterocyclic core.     

Recent advances in iridium(I) catalysis towards directed hydrogen isotope exchange

The initial discovery and establishment of a family of novel iridium catalysts possessing N-heterocyclic carbene units alongside bulky phosphine ligands allowed selected substrates to be labelled using deuterium or tritium gas at desirably low catalyst loadings via an ortho-directed C―H insertion process. Such a method has broad applicability and offers distinct advantages within the pharmaceutical industry, directly facilitating the ability to carefully monitor a potential drug molecule's biological fate. Over the past decade since these initial protocols were divulged, many additional advances have been made in terms of catalyst design and substrate scope. This review describes the broadened array of new iridium catalysts and associated protocols for direct and selective C―H activation and hydrogen isotope insertion within a number of new chemical entities of direct relevance to the pharmaceutical industry.     

A convenient method for palladium‐catalyzed reductive deuteration of organic substrates using deuterated hypophosphite in D2O

A convenient method for the deuteration of organic substrates using deuterated hypophosphite as the deuterium source was investigated. Transfer deuteration of organic substrates, such as aromatic halides, alkenes, alkynes, epoxides, and O‐benzyl derivatives, in the presence of palladium on carbon in deuterium oxide proceeded efficiently to give the corresponding deuterated products in excellent yields with high deuterium contents.     

Application of 1‐butyl‐3‐methylimidazolium hexafluorophosphate to Ir(I)‐catalyzed hydrogen isotope exchange labelling of substrates poorly soluble in dichloromethane

Substrate solubility remains a major limitation in Ir(I)‐catalyzed isotopic hydrogen exchange labelling. In the search for an alternative to the solvent dichloromethane, which is critical to the success of the reaction, we examined a series of ionic liquids for their suitability. Commercially available 1‐butyl‐3‐methylimidazolium hexafluorophosphate (abbreviated to [BMI][PF₆]) was found to support efficient deuterium and tritium exchange labelling of N‐(4‐methoxyphenyl)‐N‐methyl benzamide 1 under standard conditions. The solvent dissolves both polar hydroxyl and carboxylic acid substituted acetanilides, providing isotopomers in unprecedentedly high deuterium incorporation as compared to dichloromethane. We report the application of [BMI][PF₆] and its potential for extending the scope of Ir(I)‐catalyzed H/T exchange to more polar compounds. Copyright © 2003 John Wiley & Sons, Ltd.     

Catalyst stability as a factor in iridium‐mediated ortho‐deuteration

Crossover experiments show that iridium complexes such as Ir(cod)(Py)(PCy₃).PF₆ ( 1 ) and Ir(cod)(PPh₃)₂.BF₄ are inactivated after a comparatively short period during deuterium exchange reactions. This effect can be limited to some extent by optimizing the concentration at which exchange is performed, but altering physical parameters and adding labile ligands in an effort to stabilize reactive intermediates does not improve matters. Complexes of bidentate arsines and PN ligands and, in some cases, bidentate phosphines, exhibit better stability. Unexpectedly, following hydrogenolysis, 1 is able to mediate deuterium exchange from deuterium oxide into amides and N‐heterocycles at room temperature. Copyright © 2007 John Wiley & Sons, Ltd.     

Unusual base‐catalyzed exchange in the synthesis of deuterated PF‐2413873a

The preparation of deuterated PF‐2413873 (4‐[3‐cyclopropyl‐1‐(methanesulfonylmethyl)‐5‐methyl‐1H‐pyrazol‐4‐yl]oxy‐2,6‐dimethylbenzonitrile, 1) is described for use as a bioanalytical standard in clinical trials. Two strategies were investigated. The sulfone‐containing substituent was labelled by base‐catalyzed exchange, but unacceptable deuterium loss was noted under assay conditions. Alternatively, labelling 4‐cyano‐3,5‐dimethylphenol was achieved by heating with deuterium oxide over platinum oxide. After building up the pyrazole ring we discovered that, during the subsequent alkylation to attach the methylthiomethyl group, the base, potassium t‐butoxide, caused unwanted scrambling of deuteriums on the aromatic portion and the methylthiomethyl group. Thus, it was necessary to remove all base‐labile hydrogens to prevent their exchange. This was accomplished by alkylating the pyrazole with per‐deuterated chloromethyl methylsulfide, oxidation to the sulfone, and selective removal of its deuteriums by treatment with sodium hydroxide. The unusual sensitivity and selectivity of these base‐promoted exchange reactions are discussed. Thus, 4‐[3‐cyclopropyl‐1‐(methanesulfonylmethyl)‐5‐methyl‐1H‐pyrazol‐4‐yl]oxy‐[²H₆]2,6‐dimethyl‐[3,5‐²H]benzonitrile (17) was obtained, labelled with eight deuterium atoms and an acceptable D₀/D₈ ratio. Copyright © 2009 John Wiley & Sons, Ltd.     

Microscale synthesis of isotopically labeled 6R‐N5, N10 methylene‐5, 6, 7, 8‐tetrahydrofolate

A one‐pot chemo‐enzymatic microscale synthesis of isotopically labeled R‐[6‐^YH; 11‐^XH] N⁵, N¹⁰ methylene‐5, 6, 7, 8‐tetrahydrofolate (CH₂H₄folate) is presented, where Y=1 or 2 represents protium or deuterium, and X=1, 2 or 3 represents protium, deuterium or tritium, respectively. In this procedure, Thermoanaerobium brockii alcohol dehydrogenase (tbADH) and Escherichia coli dihydrofolate reductase (ecDHFR) were used simultaneously in the reaction mixture. First, tbADH stereospecifically catalyzes a hydride transfer from [2‐^YH] iPrOH to the re face of C‐4 NADP⁺. The ecDHFR then reduced 7, 8‐dihydrofolate (H₂folate) to form (6S)‐H₄folate. Finally, the enzymatic reactions were followed by chemical trapping with isotopically labeled formaldehyde ([^XH]‐HCHO) to form the final product. The preparation of deuterium‐ and tritium‐labeled formaldehyde is also presented. Two reverse phase HPLC methods were developed for analysis and purification of product R‐[6‐^YH; 11‐^XH] CH₂H₄folate. This isotopically labeled cofactor can be used to study 1° and 2° kinetic isotope effects (KIEs) with any CH₂H₄folate dependent enzyme as demonstrated by studies with E. coli thymidylate synthase (TS). Copyright © 2005 John Wiley & Sons, Ltd.     

s-cis,s-trans Isomerism about the Ala-Pro peptide bond in Nα-phospho- and Nα-phosphono-l-alanyl-l-prolines by 31P n.m.r.

Nα-(Phenethylphosphono)-l -alanyl-l -proline 1, a potent inhibitor of angiotensin converting enzyme, exhibits two ³¹P n.m.r. resonances (intensity ratio one to one), which exchange with a constant (k₁) of about 1 s^?1 and a free energy of activation ΔG*=～ 20kcal/mol at 23° in deuterated dimethylsulfoxide. Two resonances in exchange are also observed in deuterium oxide at pH 7.5. Thus the exchanging ³¹P resonances report the s-cis, s-trans conformational equilibrium about the alanyl-proline peptide bond. Similar results were observed with Nα-[(O-phenyl)-phenethylphosphono]-l -alanyl-l -proline 2. Nα -(O-phenylphospho)-l -alanyl-l -proline, 3 Nα-(O, O′-diphenylphospho)-l -alanyl-l -proline 4, and Nα-[2-(2-oxo-1, 3, 2-dioxaphosphiranyl)]-l -alanyl-l -proline 5 in deuterated dimethyl sulfoxide, deuterium oxide, and deuterochloroform. A ¹³C n.m.r. spectrum of 5 confirmed the presence of s-cis and s-trans resonances for the proline carbons in the same intensity ratio observed by ³¹P n.m.r.     

Manganese (IV) oxide‐catalyzed electrophilic diamination of electron‐deficient alkenes provides an easy synthesis of α,β‐diamino acid and ketone derivatives for peptidomimetic studies*

Abstract: Manganese (IV) oxide was found to catalyze the diamination reaction of α,β‐unsaturated esters and ketones with N,N‐dichloro‐p‐toluenesulfonamide and acetonitrile as the halogen and nitrogen sources. The reaction is convenient to be conducted by simply mixing three reactants in the presence of manganese dioxide catalyst and 4 Å molecular sieves, and provides an easy access to 1‐p‐toluenesulfonyl‐3‐trichloromethyl‐4,5‐imidazoline derivatives, which are useful building blocks for peptidomimetic studies.     

Syntheses of [2H3, 15N], [14C]Nexavar™ and its labeled metabolites

Nexavar?, Sorafenib tosylate (BAY 43‐9006 tosylate) is a potent small molecule Raf kinase inhibitor for the treatment of hyperproliferative disorders such as cancer. Both radiolabeled and stable isotope labeled compounds were required for drug absorption, distribution, metabolism and excretion (ADME) and quantitative mass spectrometry bio‐analytical studies. Nexavar? labeled with carbon‐14 in the carboxamide group was prepared in two steps in an overall radiochemical yield of 42% starting from 4‐chloro‐N‐methyl‐2‐pyridine‐[¹⁴C]carboxamide. The [²H₃,¹⁵N] version of Nexavar? was prepared in 75% yield based on 4‐chloro‐N‐[²H₃]methyl‐2‐pyridine‐[¹⁵N]carboxamide. The pyridine N‐oxide metabolite labeled with carbon‐14 as well as with deuterium and nitrogen‐15 and was synthesized by oxidation in yields of 59% and 87%, respectively. Starting from [²H₂, ¹³C]formaldehyde the N‐hydroxymethyl metabolite was labeled with carbon‐13 and deuterium in one step in a 45% overall yield. Copyright © 2006 John Wiley & Sons, Ltd.     

Evaluation of a P,N‐ligated iridium(I) catalyst in hydrogen isotope exchange reactions of aryl and heteroaryl compounds

We have developed a novel and efficient iridium‐catalyzed hydrogen isotope exchange reaction method with secondary and tertiary sulfonamides at ambient temperatures. Furthermore N‐oxides and phosphonamides have been successfully applied in hydrogen isotope exchange reactions with moderate to excellent deuterium introduction.     

New radiolabelling chemistry: synthesis of phosphorus–[18F]fluorine compounds

The feasibility of synthesizing compounds containing the P–¹⁸F bond has been demonstrated by labelling the pesticide, cholinesterase inhibitor Dimefox (N,N,N′N′‐tetramethylphosphorodiamidic fluoride) with F‐18. Radiolabelling was achieved in high radiochemical yield (96%) by nucleophilic substitution of the chloro group attached to phosphorus, in the oxidation state P(V), by ¹⁸F^? (activated with tetrabutylammonium carbonate in acetonitrile). Given the large number of important biological molecules possessing phosphorus such as oligonucleotides, phospholipids as well as phosphorylated proteins, sugars and steroids, this new labelling chemistry may provide an additional route to radiolabelling these biologically important compounds for use in PET. Copyright © 2005 John Wiley & Sons, Ltd.     

NMR analysis of t‐butyl‐catalyzed deuterium exchange at unactivated arene localities

Regioselective labelling of arene rings via electrophilic exchange is often dictated by the electronic environment caused by substituents present on the aromatic system. Previously, we observed the presence of a t‐butyl group, either covalently bond or added as an external reagent, could impart deuterium exchange to the unactivated, C1‐position of estrone. Here, we provide nuclear magnetic resonance analysis of this exchange in a solvent system composed of 50:50 trifluoroacetic acid and D₂O with either 2‐t‐butylestrone or estrone in the presence of t‐butyl alcohol has shed insights into the mechanism of this t‐butyl‐catalyzed exchange. Fast exchange of the t‐butyl group concurrent with the gradual reduction of the H1 proton signal in both systems suggest a mechanism involving ipso attack of the t‐butyl position by deuterium. The reversible addition/elimination of the t‐butyl group activates the H1 proton towards exchange by a mechanism of t‐butyl incorporation, H1 activation and exchange, followed by eventual t‐butyl elimination. Density functional calculations are consistent with the observation of fast t‐butyl exchange concurrent with slower H1 exchange. The σ‐complex resulting from ipso attack of deuterium at the t‐butyl carbon was 6.6 kcal/mol lower in energy than that of the σ‐complex resulting from deuterium attack at C1. A better understanding of the t‐butyl‐catalyzed exchange could help in the design of labelling recipes for other phenolic metabolites.     

Efficient,scalable and economical preparation of tris(deuterium)‐ and 13C‐labelled N‐methyl‐N‐nitroso‐p‐toluenesulfonamide (Diazald®) and their conversion to labelled diazomethane

A method for the preparation of multi‐gramme quantities of N‐methyl‐d₃‐N‐nitroso‐p‐toluenesulfonamide (Diazald‐d₃) and N‐methyl‐¹³C‐N‐nitroso‐p‐toluenesulfonamide (Diazald‐¹³C) and their conversion to diazomethane‐d₂ and diazomethane‐¹³C, respectively, is presented. This approach uses robust and reliable chemistry, and critically, employs readily commercially available and inexpensive methanol as the label source. Several reactions of labelled diazomethane are also reported, including alkene cyclopropanation, phenol methylation and α‐diazoketone formation, as well as deuterium scrambling in the preparation of diazomethane‐d₂ and subsequent methyl esterification of benzoic acid.     

Nitramine, 12. Mitt. Fluormethyl- und Azidomethyl-alkylnitramine

Nitramines, XII: N-Fluoromethyl- and N-Azidomethyl-N-alkylnitramines The N-chloromethyl-N-alkylnitramines 1 react with potassium fluoride and 18-crown-6 to give the unstable N-fluoromethyl-N-alkylnitramines 2 . With sodium azide the stable N-azidomethyl-N-alkylnitramines 3 are formed. With alkynes the latter cyclize to yield 1,2,3-triazoles, the structures of which were studied.     

Practical protocols for stepwise solid‐phase synthesis of cysteine‐containing peptides

Abstract: This study details a series of conditions that may be applied to ensure ‘safe’ incorporation of cysteine with minimal racemization during automated or manual solid‐phase peptide synthesis. Earlier studies from our laboratories [Han et al. (1997) J. Org. Chem. 62 , 4307–4312] showed that several common coupling methods, including those exploiting in situ activating agents such as N‐[(dimethylamino)‐1H‐1,2,3‐triazolo[4,5‐b]pyridin‐1‐ylmethylene]‐N‐methylmethanaminium hexafluorophosphate N‐oxide (HATU), N‐[1H‐benzotriazol‐1‐yl)‐(dimethylamino)methylene]‐N‐methylmethanaminium hexafluorophosphate N‐oxide (HBTU), and (benzotriazol‐1‐yl‐N‐oxy)tris(dimethylamino)phosphonium hexafluorophosphate (BOP) [all in the presence of N‐methylmorpholine (NMM) or N,N‐diisopropylethylamine (DIEA) as a tertiary amine base], give rise to unacceptable levels (i.e. 5–33%) of cysteine racemization. As demonstrated on the tripeptide model H‐Gly‐Cys‐Phe‐NH₂, and on the nonapeptide dihydrooxytocin, the following methods are recommended: O‐pentafluorophenyl (O‐Pfp) ester in DMF; O‐Pfp ester/1‐hydroxybenzotriazole (HOBt) in DMF; N,N′‐diisopropylcarbodiimide (DIPCDI)/HOBt in DMF; HBTU/HOBt/2,4,6‐trimethylpyridine (TMP) in DMF (preactivation time 3.5–7.0 min in all of these cases); and HBTU/HOBt/TMP in CH₂Cl₂/DMF (1:1) with no preactivation. In fact, several of the aforementioned methods are now used routinely in our laboratory during the automated synthesis of analogs of the 58‐residue protein bovine pancreatic trypsin inhibitor (BPTI). In addition, several highly hindered bases such as 2,6‐dimethylpyridine (lutidine), 2,3,5,6‐tetramethylpyridine (TEMP), octahydroacridine (OHA), and 2,6‐di‐tert‐butyl‐4‐(dimethylamino)pyridine (DB[DMAP]) may be used in place of the usual DIEA or NMM to minimize cysteine racemization even with the in situ coupling protocols.