13 Carbon-labelling carboxylic acid synthesis

Kabalka, G. W., Delgado, M. C., Sastry, U., Sastry, K. A. R. 1982. Synthesis of carbon-13 labeled carboxylic acids via organoborane reactions. J. Chem. Soc. Chem. Commun. 21 1273-1274. 

Amino Acids and Peptides. - Wasserman s method of one-carbon homologation of carboxylic acids to give a-ketocarboxylates involves reaction with cyanomethylenetriphenyl-phosphorane followed by ozone (Scheme 24) and has been used as a key step in a chemo-enzymatic synthesis of isotopically labelled L-valine, L-isoleucine, and o/fo-isoleucine. Alkylation of the carbanion derived from the imino-substituted methylphosphonate diphenyl ester (186) with indol-3-ylmethyl bromide followed by appropriate deprotection has been used to prepare the phosphonate analogue (187) of tryptophan (Scheme 25). The deprotected analogue (188) and derived peptides show activity as inhibitors of chymotrypsin. Two approaches to solid phase Wadsworth-Enunons reactions which have applications in combinatorial chemistry have been reported. In one diethylphosphonoacetamide is bound to PEG-PAL resin via a peptide link, while... 

A biomimetic synthesis of the tobacco alkaloid anatabine (1,2,3,6-tetrahydro-2,3 -bipyridine) (69) has been reported starting from 1,2,3,6-tetrahydropyridine-2-carboxylic acid (68) labelled with carbon-13 and deuterium at C-2 (Scheme 15).9" ... 

According to the head-to-head condensation mechanism for the synthesis of C29 alkane the carboxyl carbon of Ci acid should be lost during the first step (Fig. 12). However, experiments with specifically and doubly labeled C], and Cig acids showed that the entire carbon chains of both acids were incorporated without decarboxylation into Q alkane in B. oleracea (Kolattukudy, 1966) and C31 alkane in P. sativum and Spinaceae oleracea (Kolattukudy, 1968c). 

Further evidence obtained from intact animals suggested that more than one pathway for fatty acid synthesis might exist. After injection of l-[ " C]acetate, palmitate is labeled throughout the molecule, but only the last carboxyl of stearate is labeled. These differences in labeling can now be explained the random labeling results from the repeated condensation of 2-carbon compounds, and the terminal labeling results from a process known to occur in mitochondria and referred to as elongation. ... 

A synthesis of papaverine-l- C in 11 % yield was performed starting from veratric acid labeled at the carboxyl group (CXII). The diazoketone prepared from the aeid chloride was eondensed with homoveratrylamine and the resulting amide (CXIII) cyclized to 3,4-dihydropapaverine (CXIV). Dehydrogenation of the latter with palladium-charcoal in tetralin afforded papaverine. The loealization of the labeled carbon atom, indicative of an Amdt-Eistert rearrangement, was confirmed because the base, on oxidation with neutral 5% potassium permanganate, afforded 6,7-dimethoxyisoquinoline-l-i C-l-carboxylic acid (CXV) (183). 

Roth et al. (196), using C -carboxyl labeled nicotinic acid or nicotinamide, have shown that the compounds are rapidly excreted. A total of about 3% of the carboxyl group was expired as carbon dioxide within 1 day after the injection. A few more per cent is given off as exhaled carbon dioxide several days later. It has been estimated that approximately 15 % of the radioactivity fixed in the tissue is exhaled as carbon dioxide. In later studies it was shown that carbon dioxide production from labeled nicotinic acid and nicotinamide also occurs in hamsters, rats, and dogs (221). Hamsters and rats excrete somewhat more by this route, and the dog very little. This is illustrated in Fig. 10. It is of importance to note that very little of the radioactivity appears in the feces. This may be significant with regard to intestinal synthesis of the vitamin. 

Figure 9 End-labeling strategies for the incorporation of bioactive epitopes, (a) Termination of the polymerization can be effected with aldehydes (molybdenum alkylidenes) or vinyl ethers (ruthenium carbenes) to install a single reporter group, (b) Termination of ruthenium carbenes with a vinyl carbonate or vinyl lactone leaves a terminal aldehyde or carboxylic acid that can be further derivatized. (c) Sacrificial synthesis An acetal orthioacetal block can be unmasked to reveal a single terminal alcohol or thiol that can be modified selectively.

These studies were confirmed by tracer experiments showing that nitrogen of nicotinic acid (formed by Neurospora) is derived from 3-hydroxyanthranilic acid (478). Experiments with doubly labeled tryptophan demonstrate that tryptophan is probably the only source of quinolinic acid in rat metabolism (645) and that carbon atom 3 of tryptophan, the precursor of the carboxyl carbon of 3-hydroxyanthranilic acid, becomes carboxyl carbon in nicotinic acid (310,340,341,373). In vitro studies of the enzymic oxidation of 3-hydroxyanthranilic acid confirm its relationship to quinolinic acid (498) and show that picolinic acid may also form from it (539,540) but nicotinic acid synthesis under... 

Diazomethane is a valuable reagent for one-carbon extension of acyl halides and anhydrides, as well as for ring expansion reactions of cyclic ketones " . It is also widely used in small-scale organic synthesis for the esterification of carboxylic acids and the etherification of phenols, enols and alcohols " . In these reactions, however, CH2N2 installs the label in positions that are potentially metabolically labile and usually unsuitable for use in metabolism smdies. For in vitro studies where metabolism is not an issue, tritiated diazomethane is usually preferred because of its higher specific activity. 

Desulfurization of [ C]thiourea can also be accomplished by reduction with Raney-Ni in the presence of NH4OAC, providing [ C]formamidine acetate in nearly quantitative yield . This was used in a reaction with 5-amino-2-fluoropyridine-4-carboxylic acid (71) to produce the carbon-14-labeled pyrido[3,4-d]pyrimidine 72, an intermediate in the synthesis of [ C]PD0205520 (73). an EGFR tyrosine kinase inhibitor. 

This substance has been shown by tracer studies to be an efficient precursor of terpenes and steroids. Mevalonic acid has six carbon atoms, whereas the isoprene unit has only five. Therefore, if mevalonic acid is the precursor of isoprene units, it must lose one carbon atom at some stage. Synthesis of mevalonic acid labeled at the carboxyl group with 14C, and use of this material as a starting material for production of cholesterol, gives unlabeled cholesterol. Therefore, the carboxyl carbon is the one that is lost ... 

The ability of the rat to use lactate, pyruvate, and D-glucose to equal extents in the synthesis of 1-naphthyl D-glucosiduronic acid was demonstrated by Packham and Butler.158 The lactate and pyruvate were labeled with C14 at the carboxyl carbon lactate labeled at Cl and C2 was also used and the D-glucose was uniformly labeled. D-Glucuronate-C14 was utilized for D-glucuronic conjugation on oral administration, but was twenty times as effective after intraperitoneal injection. 

This imidazoline-carboxylate synthesis involves the coupling of four separate cont5)onents (two imines, an acid chloride and carbon monoxide), and the generation of at least five separate bonds, all via a one-pot, palladium catalyzed process. From an analysis of the structure of the imidazoline carboxylate, the individual constituents can be seen (Figure 3). This stmcture might be considered to arise from the dipolar cycloaddition of an imine with a mesoionic l,3-oxazolium-5-oxide (5) intermediate, which itself could be generated from imine, acid chloride and carbon monoxide. Consistent with this potential formulation, performing the catalytic reaction with CO leads to the incorporation of the carbon-13 label into the carboxylate position of 4. 

The H CN (or CN, if the reaction is done under basic conditions) synthon has been mainly used to extend the carbon chain by one carbon. For example, cyanide ion has been used in the synthesis of amino acids labelled in the carboxylate group. This is accomplished using the high pressure-high temperature modification of the Bucherer-Strecker synthesis. In this reaction, bisulphite addition complex of an aldehyde reacts with cyanide ion in the presence of ammonium carbonate to form a hydantoin, which is then converted into the amino acid by basic hydrolysis (equation 61). 

During fatty acid biosynthesis, the carbon chain grows two carbons at a time by the condensation of an acyl-ACP with malonyl-ACP, with the malonyl-ACP becoming, in every case, the carboxyl end of the new acyl-ACP. Thus, the chain grows from methyl to carboxyl. Since " C-labeled malonyl CoA was added a short time before synthesis was stopped, the fatty acids whose synthesis was completed during this short period will be heavily labeled toward the carboxyl end (the last portion synthesized) and less heavily labeled, if at all, on the methyl end. 

A new synthesis of showdomycin (27), outlined in Scheme 3, utilizes the carboxylation of the ribofuranosyl ethyne (28). Showdomycin, labelled with carbon-14 at the /3-carbonyl group, has been prepared by Wittig condensation of the 3,6-anhydrohept-2-ulosonic acid derivative (29) with the phosphorane prepared from [l- " C]chloroacetamide (Scheme 4). ... 

As shown in Scheme 12.71, and as affirmed by labeling experiments, an amino acid (shown using phenylalanine [Phe, F] as an example in the scheme) to be added to the growing peptide (protein) chain is activated for reaction by attachment at the carboxylate to adenosine triphosphate (ATP) to make an anhydride of the amino acid with adenosine monophosphate (and the loss of inorganic phosphate). Then, in the next step, the activated amino acid is esterified by the C-2 or the C-3 hydroxyl of a ribosyl unit of AMP, which is attached via phosphate at the 5 carbon to the aminoacyl transfer end of tRNA. If attachment is to C-2, rearrangement to C-3 follows and the aminoacyl-tRNA is activated and ready to be added to the growing peptide chain at the synthesis site on the ribosome. 

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