Synthesis of tracer labelled [11,12‐3H]‐β‐carotene

The synthesis of tracer labelled [11,12‐³H]‐β‐carotene is described. The procedure uses Wittig condensation of tracer labelled ³H‐retinal (retinal spiked with [11,12‐³H]‐retinal) with retinyl triphenylphosphonium bromide. The preparation of tracer labelled[³H]‐β‐carotene is suitable for studies involving bioavailability and bioconversion of β‐carotene to vitamin A. Copyright © 2003 John Wiley & Sons, Ltd.     

Synthesis of tritium‐labelled BIBN4096, an experimental h‐CGRP‐antagonist

Synthesis of tritium labelled BIBN 4096 – 1‐piperidine‐³H‐carboxamide, N‐[2‐[[5‐amino‐1‐[[4‐(4‐pyridinyl)‐1‐piperazinyl]carbonyl]pentyl]amino]‐1‐[(3,5‐dibromo‐4‐hydroxyphenyl)methyl]‐2‐oxoethyl]‐4‐(1,4‐dihydro‐2‐oxo‐3(2 H)‐quinazolinyl)‐, [R‐(R*,S*)]‐, – a h‐CGRP‐antagonist for the treatment of migraine is described. Selective tritiation of a heterocyclic aromatic fragment in the presence of aromatic ring in a precursor of BIBN4096 was successfully carried out using the solid state catalytic exchange method. Subsequent completion of the synthesis sequence gave the final [³H]BIBN4096 with a specific activity of >170 Ci/mmol. Copyright © 2006 John Wiley & Sons, Ltd.     

A new [2H]‐labelled α‐trichloroimidate glucuronic ester for the synthesis of deuterated drug conjugates

A new reaction pathway for the synthesis of a [²H]‐labelled trichloroacetimidate precursor for the preparation of glucuronides is described. Therewith, stable isotope‐labelled drug glucuronides become accessible on a preparative scale, which can further be used as internal standards for quantitative analysis.     

Synthesis of [3′‐14C] coenzyme Q10

Radio‐labelled coenzyme Q₁₀, labelled at the 3′‐position with ¹⁴C, was synthesized starting from natural solanesol and ethyl [3‐¹⁴C] acetoacetate. The radiochemical yield was 8.0% from ethyl [3‐¹⁴C] acetoacetate. The specific radioactivity of the product was 44.8 μCi, 1.66 MBq/mg. The specific radioactivity and radiochemical purity are sufficiently high to enable us to use this labelled form of coenzyme Q₁₀ in metabolic studies. Copyright © 2002 John Wiley & Sons, Ltd.     

Synthesis of 14C‐labelled myosmine, [2′‐14C] ‐3‐(1‐pyrrolin‐2‐yl)pyridine

¹⁴C‐Labelled myosmine ([2′‐¹⁴C]‐3‐(1‐pyrrolin‐2‐yl)pyridine) was synthesized for autoradiography studies starting from [carboxyl‐¹⁴C]‐nicotinic acid by initial esterification of the latter in the presence of 1,1,1‐triethoxyethane. Without any purification the ethyl nicotinate formed was directly reacted with N‐vinyl‐2‐pyrrolidinone in the presence of sodium hydride, yielding ¹⁴C‐labelled myosmine. The product was purified by silica gel column chromatography. The radiochemical yield was 15% and the specific activity 55.2 mCi/mmol. Copyright © 2003 John Wiley & Sons, Ltd.     

Synthesis of 1‐(2′‐deoxy‐2′‐fluoro‐β‐D‐arabinofuranosyl)‐[Methyl‐11C]thymine ([11C]FMAU) via a Stille cross‐coupling reaction with [11C]methyl iodide

1‐(2′‐deoxy‐2′‐fluoro‐β‐D‐arabinofuranosyl)‐[methyl‐¹¹C]thymine ([¹¹C]FMAU) [¹¹C]‐ 1 was synthesised via a palladium‐mediated Stille coupling reaction of 1‐(2′‐deoxy‐2′‐fluoro‐β‐D‐arabinofuranosyl)‐5‐(trimethylstannyl)uracil 2 with [¹¹C]methyl iodide in a one‐pot procedure. The reaction conditions were optimized by screening various catalysts and solvents, and by altering concentrations and reaction temperatures. The highest yield was obtained using Pd₂(dba)₃ and P(o‐tolyl)₃ in DMF at 130°C for 5 min. Under these conditions the title compound [¹¹C]‐ 1 was obtained in 28±5% decay‐corrected radiochemical yield calculated from [¹¹C]methyl iodide (number of experiments=7). The radiochemical purity was >99% and the specific radioactivity was 0.1 GBq/μmol at 25 min after end of bombardment. In a typical experiment 700–800 MBq of [¹¹C]FMAU [¹¹C]‐ 1 was obtained starting from 6–7 GBq of [¹¹C]methyl iodide. A mixed ¹¹C/¹³C synthesis to yield [¹¹C]‐ 1 /(¹³C)‐ 1 followed by ¹³C‐NMR analysis was used to confirm the labelling position. The labelling procedure was found to be suitable for automation. Copyright © 2002 John Wiley & Sons, Ltd.     

Preparation of 3′‐deoxy‐3′‐[18F]fluorothymidine ([18F]FLT) in ionic liquid, [bmim][OTf]

Although 3′‐deoxy‐3′‐[¹⁸F]fluorothymidine ([¹⁸F]FLT) is a prospective radiopharmaceutical for the imaging of proliferating tumor cell, it is difficult to prepare large amount of [¹⁸F]FLT. We herein describe the preparation of [¹⁸F]FLT in an ionic liquid, [bmim][OTf] (1‐butyl‐3‐methyl‐imidazolium trifluoromethanesulfonate). At optimized condition, [¹⁸F]fluorinationin ionic liquid with 5 µl of 1 M KHCO₃ and 5 mg of the precursor yielded 61.5 ± 4.3% (n=10). Total elapsed time was about 70 min including HPLC purification. The rapid synthesis of [¹⁸F]FLT can be achieved by removing all evaporation steps. Overall radiochemical yield and radiochemical purity were 30 ± 5% and >95%, respectively. This method can use a small amount of a nitrobenzenesulfonate precursor and can be adapted for automated production. Copyright © 2006 John Wiley & Sons, Ltd.     

Synthesis of (E)‐2,3′,4,5′‐tetramethoxy[2‐11C]stilbene

In this paper, we describe the radiosynthesis of the compound (E)‐2,3′,4,5′‐tetramethoxy[2‐¹¹C]stilbene, a potential, universal tumour positron emission tomography imaging agent. The production of (E)‐2,3′,4,5′‐tetramethoxy[2‐¹¹C]stilbene was carried out via ¹¹C‐methylation of (E)‐2‐(hydroxy)‐3′,4,5′‐trimethoxystilbene by using [¹¹C]methyl trifluoromethanesulfonate ([¹¹C]methyl triflate). (E)‐2,3′,4,5′‐tetramethoxy[2‐¹¹C]stilbene was obtained with a radiochemical purity greater than 95% in a 20 ± 2% decay‐corrected radiochemical yield, based upon [¹¹C]carbon dioxide. Synthesis, purification and formulation were completed on an average of 30 min following the end of bombardment (EOB). The specific radioactivity obtained was 1.9 ± 0.6 GBq/µmol at EOB. Copyright © 2007 John Wiley & Sons, Ltd.     

Synthesis of all‐trans‐[10′‐3H]‐8′‐apo‐β‐carotenoic acid

The enzyme, 15,15′‐β‐carotene dioxygenase (BCDOX), facilitates the oxidation of β‐carotene to yield retinal. This is a remarkable process in which one of 11 double bonds in β‐carotene is selectively oxidized. To further probe the mechanistic aspects of BCDOX, the synthesis of all‐trans‐[10′‐³H]‐8′‐apo‐β‐carotenoic acid is reported. This compound will be used as a photoaffinity labeling reagent to probe the β‐carotene binding pocket within BCDOX. The synthesis outlines a simple and efficient route for the incorporation of tritium at the 10′ olefinic carbon of 8′‐apo‐β‐carotenoic acid. Copyright © 2002 John Wiley & Sons, Ltd.     

A convenient synthesis of 2′,3′‐dideoxyinosine‐13C5 and related 2′,3′‐dideoxyadenosine‐13C5

2′,3′‐Dideoxyinosine‐¹³C₅ (ddI‐¹³C₅) and the related 2′,3′‐dideoxyadenosine‐¹³C₅ (ddA‐¹³C₅) were prepared from (S)‐5‐[¹³C₅]2,3‐dideoxyribonolactone 1 . From a batch of this starting material ddI‐¹³C₅ was made in 27% overall yield in seven steps and ddA‐¹³C₅ in five steps and 14% overall yield. The known synthesis of ddI‐¹³C₅ from glucose‐¹³C₆ took 18‐steps_; therefore the present work is a substantial improvement. Copyright © 2010 John Wiley & Sons, Ltd.     

Contribution of a significant first‐pass effect of dimethyl‐4,4′‐dimethoxy‐5,6,5′,6′‐dimethylene dioxybiphenyl‐2,2′‐dicarboxylate in the liver to its poor bioavailability in rats

Objectives The objective of this study was to investigate the mechanism responsible for the poor oral bioavailability of dimethyl-4′,4′-dimethoxy-5,6,5′,6′-dimethylene dioxy-biphenyl-2,2′-dicarboxylate (DDB), a hepatoprotective agent, in rats. Methods DDB was intravenously administered to rats at doses of 0.2-1 mg/kg. To determine the hepatic first-pass effect in rats, DDB (1 mg/kg) was administered via the pyloric vein and the femoral vein. Direct measurement of intestinal permeability was attempted using Caco-2 cell monolayers and rat intestinal epithelium. Key findings A moment analysis indicated that the volume of distribution and clearance remained unchanged with the magnitude of the dose, indicating that DDB exhibited linear pharmacokinetics. When the area under the curve for DDB after administration to the pyloric vein was compared with that after femoral vein administration, the ratio (F_H) was found to be 0.294, indicating a significant first-pass effect for DDB. The permeability of DDB was high in the rat intestine (1.78 ± 0.229 × 10⁻⁵ cm/s) and in Caco-2 cell monolayers (6.8 ± 0.70 × 10⁻⁵ cm/s), suggesting that DDB, in soluble form, was readily permeable across the intestinal epithelium. Conclusions These observations indicated that despite the fact that DDB was readily permeable to the intestinal epithelium, a significant first-pass metabolism was associated with its pharmacokinetics in rats.     

Synthesis of deuterium‐labelled coenzyme Q10

Deuterium‐labelled coenzyme Q₁₀ ([2‐CD₃‐1′‐CD₂]coenzyme Q₁₀, coenzyme Q₁₀‐d₅ ) was synthesized by condensation of 2,3‐dimethoxy‐[5‐CD₃]methyl‐1, 4‐hydroquinone with [1‐CD₂]decaprenol. Five positions were selected for deuteration as replacement at these positions allowed examination of every step of the synthesis. This examination was carried out by a combination of ¹H‐ and ¹³C‐nuclear magnetic spectrometry and mass spectrometry. Further, these positions have been proved to be metabolically stable. This reagent makes simultaneous quantification of the source of coenzyme Q₁₀ (exogenously supplied or endogenously supplied) possible in biological samples by measurements on gas chromatography–mass spectrometry. Copyright © 2002 John Wiley & Sons, Ltd.     

Synthesis of (±) [5‐3H] N′‐nitrosoanatabine,a tobacco‐specific nitrosamine

Tobacco‐specific N′‐nitrosamines (TSNA) are a unique class of systemic organ‐specific carcinogens. The TSNA are formed by N‐nitrosation of nicotine and of the minor tobacco alkaloids after harvesting of tobacco and during smoking. The N‐nitrosation of anatabine leads to N′‐nitrosoanatabine (NAT; 1‐nitroso‐1,2,3,4‐tetrahydro‐2,3'‐bipyridyl) which requires in‐depth assays in laboratory animals other than the rat. Furthermore, delineation of its tissue distribution and metabolism is needed for structure:activity comparisons with other TSNA and for the assessment of potential human risk from this TSNA. We have, therefore, synthesized (±)[5‐³H]NAT. 5‐Bromo‐3‐pyridine‐carboxaldehyde was condensed with ethyl carbamate prior to Diels–Alder reaction with 1,4‐butadiene to give the racemic anatabine ring system. Hydrolysis, followed by reduction with LiAlT₄ and nitrosation, led to (±)[5‐³H]NAT (60% yield, specific activity 266 mCi/mmol, radiochemical purity of >99%). Copyright © 2002 John Wiley & Sons, Ltd.     

Transport properties of 3′-azido-3′-deoxythymidine and 2′,3′-dideoxyinosine in the rat choroid plexus

Transport properties of 3′-azido-3′-deoxythymidine (AZT) and 2′, 3′-dideoxyinosine (DDI) were characterized in the isolated rat choroid plexus. AZT and DDI competitively inhibited the active transport of [³H]benzylpenicillin, a prototypic organic anion, with K_i values of 85·4±13·1 and 155±22 μM, respectively. Accumulation of [³H]DDI was against an electrochemical potential via a saturable process (K_m=29·7±4·9 μM, V_max=13·5±2·4 pmol min⁻¹/μL tissue) that was inhibited by metabolic inhibitors (carbonylcyanide p -trifluoromethoxyphenylhydrazone, 10 μM, and rotenone, 30 μM) and sulphydryl reagents (p -chloromercuribenzoic acid, 100 μM, and p -chloromercuribenzenesulphonic acid, 100 μM), but did not require an inwardly directed Na⁺ gradient. Accumulation of [³H]DDI was inhibited by benzylpenicillin and AZT in a dose-dependent manner, with IC₅₀ values of 91·6±28·9 and 294±84 μM, respectively. In contrast, no significant accumulation of [³H]AZT was observed. These results suggest that DDI is transported, at least in part, by the transport system for organic anions located on the rat choroid plexus, whereas AZT is recognized, but not transported by this system. © 1997 John Wiley & Sons, Ltd.     

Synthesis of N‐[1‐13C]caproyl‐N′‐phenylthiourea

An optimal synthesis of N‐[1‐¹³C]caproyl‐N′‐phenylthiourea with isotopic enrichment 82% is described, starting from barium [¹³C]carbonate, using five synthetic steps. Yields were 95% relative to caproyl chloride and 46% relative to barium carbonate. Oxidation of the title compound with manganese dioxide yields the corresponding ureide. Structural similarities with anticonvulsants such as phenacemide make N‐caproyl‐N′‐phenylthiourea an interesting model compound. Copyright © 2004 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.     

Synthesis and Protective Effects of Kaempferol‐3′‐sulfonate on Hydrogen Peroxide‐induced injury in Vascular Smooth Muscle Cells

A novel water‐soluble sulfated derivative, kaempferol‐3′‐sulfonate acid sodium (KS) with the composition of [C₁₅H₉O₉SNa]·2.5H₂O, was synthesized and characterized by elemental analysis, IR, ¹H NMR, ¹³C NMR, and HRMS. Its protective effects on human vascular smooth muscle cells injured by hydrogen peroxide were evaluated by CCK‐8 method, flow cytometry, and Western blotting. The experimental results indicated that the KS can significantly increase cell viability and reduce apoptosis on H₂O₂‐injured VSMCs, as well as reverse the effects of H₂O₂ on Bcl‐2, Bad, and caspase‐3 expressions. In addition, LDH leakage, MDA levels, and SOD and GSH activities were also measured with spectrophotometry. The results indicated that the KS acted as antioxidant preventing LDH leakage and MDA production, while increasing intracellular SOD and GSH activities. These findings revealed that KS might potentially serve as an effective antioxidant agent for prevention and treatment of vascular disease caused by H₂O₂‐injured VSMCs.     

Synthesis of tritium‐ and deuterium‐labeled budesonide

Tritium‐labeled budesonide was prepared by the selective reduction of a double bond in the butenylenedioxy side chain using carrier‐free tritium and palladium on carbon as a catalyst in absolute ethanol. Although the reduction gave a mixture of the desired product and the expected byproducts resulting from over reduction of the other double bonds in ring A, the desired tritium‐labeled budesonide was easily isolated by reverse phase HPLC and with specific activity of 54 Ci/mmol. [D₈]‐budesonide was prepared from 16α‐hydroxyprednisolone and D₈‐butyraldehyde in 1,4‐dioxane in the presence of perchloric acid. The isotopic enrichment was found to be more than 99 atom% D. Copyright © 2008 John Wiley & Sons, Ltd.     

Synthesis of deoxyadenosine‐5′‐(35S)‐thiomonophosphate [dAMP(35S)] of high specific activity – a key intermediate for the synthesis of Sp‐deoxyadenosine‐5′‐(alpha;‐35S)‐thiotriphosphate [Sp‐dATP (α‐35S)]

Unprotected deoxyadenosine 1 was treated with an excess of phosphorus acid and stoichiometric proportions of N, N′‐di‐p‐tolylcarbodiimide in anhydrous pyridine to give deoxyadenosine‐5′‐monophosphite 2 . The latter was activated with trimethylsilyl chloride followed by sulphurisation with elemental ³⁵S (specific activity>1000 Ci/mmol) in toluene solution to give deoxyadenosine‐5′‐(³⁵S)‐thiomonophosphate [dAMP(³⁵S)] 3 . Enzymatic conversion of deoxyadenosine‐5′‐(³⁵S)‐thiomonophosphate to Sp‐deoxyadenosine‐5′‐(α‐³⁵S)‐thiotriphosphate [Sp‐dATP (α‐³⁵S)] 5 was carried out following a standard reaction protocol. Copyright © 2001 John Wiley & Sons, Ltd.     

Synthesis of tritium‐labeled (R,R)‐4‐methoxyfenoterol

The preparation of 2′,4′,6′‐[³H₃]‐(R,R)‐4‐methoxyfenoterol, a tritium‐labeled derivative of (R,R)‐4‐methoxyfenoterol was demonstrated on a 15 mCi scale providing material with a specific activity of 57 Ci/mmol. Copyright © 2009 John Wiley & Sons, Ltd.