Experiment with tritium-labeled

Reaction Mechanism Although the /Mysine 5,6-aminomutase requires cobalamin as cofactor instead of SAM its reaction mechanism seems to be similar to that of the lysine 2,3-aminomutase [73], Experiments with tritium-labeled lysine and B12 showed that Bn is directly involved in the hydrogen shift from position 5 in d-... 

The extraction procedure for both A -THC and A -THC, as revealed by experiment with tritium labelled... 

While in an experiment with tritium labeled triamcinolone in the dog Florini, et.al. identified 6P-hydroxytriamcinolone as the major metabolic product, no triamcinolone acetonide metabolic products have been reported thus far. 

Double-resonance experiments with tritium-labeled cyclobutane show that the principal ion-molecule reactions are hydride and tritide transfer and a condensation reaction ... 

Feeding experiments with doubly labeled 3-hydroxy-4-[14C]methoxy-A-meth-yl-(f -[3H]- and -(5)-[3H]A,-benzylamines in King Alfred daffodils produced oduline (186) with high (82-85%) tritium retention (139). This observation suggested that the incorporation of A-methylisovanillamine into 186 occurred by a nonstereospecific process in which hydrogen removal from the benzylic position was governed by a kinetic isotope effect. 

Attention has been turned to the cysteine-derived part of the penicillins. The results closely define (a) the fates of the hydrogen atoms present at C-2 and C-3 of cysteine (201) and (b) the overall stereochemistry involved in the incorporation of the amino-acid. L-[l/- C,2- H]Cystine was found to give penicillin G (199) with only slight loss of tritium. The tritium label was shown (in an experiment with singly labelled precursor) to be located exclusively at C-6, similar results being obtained with L-[2- H]cystine. These results, which confirm those obtained earlier, ... 

Feeding experiments with doubly labeled [1-3H2, l-14C]tryptamine showed that 50% of the tritium at C-l of tryptamine was lost during the formation of Cinchona alkaloids—a result which indicates that the cleavage between C-5 and N-4 in corynantheal is a stereospecific process. The ring expansion to quinoline must occur via the indolenine (178) and the recy-clization of 179 to furnish cinchonidinone (180), which was found by radiochemical dilution analysis to be a natural product in C. ledgeriana. The specific incorporation of [1 l-3H2]cinchonidinone in cinchonine and quinine and the incorporation of [ll-3H2]cinchonidine in cinchonidinone is best understood by the sequence 180 — 186 and its reversal 186 — 182, as shown in Scheme 7 (95). 

Experiments with tritium and C-labelled phenylalanine samples established that this amino-acid was incorporated intact as a Cg unit (with loss of C-1). Support for a pathway proceeding via cinnamic acid and hydroxylated derivatives and benzoylacetic acid derivatives was provided in further experiments with cinnamic acid, ferulic acid/isoferulic acid, and the (easily hydrolysed) ethyl esters of benzoylacetic acid and piperonylacetic acid. As far as they go the results suggest that a mixed pathway is in operation with introduction of hydroxy- and methylenedioxy-groups on more than one intermediate. 

There is an important experimental distinction between and kinetic isotope experiments. Deuterium isotope effects are usually measured by comparing reciprocal plots from separate experiments with deuterium-labeled reactants and with unlabeled reactants, which allows determination of the V isotope effect from the ratio of intercepts and the V/K one from the ratio of slopes. In contrast to that, tritium isotope effects are measured by the internal competition method, that is, by changes in specific activity in product or residual substrate, and thus provide only the isotope effect on V/K. The kinetic isotope effects on V with tritium can be measured only with the aid of carrier-free tritiated molecules. 

Evidence on the mechanism of fragmentation of the tyrosine/dopa aromatic ring comes from experiments with tyrosine labelled at C-3 and C-5 with tritium or deuterium. Conversion of tyrosine (7.57) into dopa (7.57) inevitably means loss of half of the label. Of three pathways leading from dopa to anthramycin 7.50) one (Scheme 7.4)... 

Haynes et al. (29) showed by feeding experiments on Croton linearis Jacq. with tritium-labeled compounds that d-( — )-coclaurine is... 

Binding assays for the saxitoxins were conducted with homogenized rabbit brain and saxitoxin exchange-labelled with tritium at C-11 (92, 93). If the various saxitoxins were available with suitably intense radiolabels, then the equilibrium dissociation constant, K, could be measured directly for each. Since only saxitoxin is currently available with the necessary label, the binding experiments instead measure the ability of a compound to compete with radiolabelled saxitoxin for the binding site. The value obtained, Kj, corresponds to the uilibrium dissociation constant, K, that would be observed for the compound if it were measured directly. Affinity is defined for this assay as the reciprocal of Kj. The affinities of several of the saxitoxins (94) are summarized in Figure 11, expressed relative to saxitoxin and plotted on a logarithmic scale. 

Reduction of aldonolactones and their derivatives with isotopically modified reducing agents leads to sugars labeled at the anomeric center. Glycosides substituted with deuterium or labeled with tritium are widely employed for kinetic isotope-effect measurements, mechanistic studies, isotope-tracing experiments, and so on. 

Laszlo Heredy We did make experiments with labeled anthracene, but the results were inconclusive as to whether any tritium exchange took place at any rate it is very small. It seems that the linear compound is simply less efficient both as solvent and reactant than an angular hydrocarbon like phenanthrene. 

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