Chemical and enzymatic synthesis of tritium labelled coenzymes

Abstract

AbstractDetails of the synthesis of tritium labelled co enzymes‐nicotinamide adenine dinucleotide and coenzyme A by isotope exchange and enzymatic reactions are reported. It was established, that among the investigated chemical reactions, the most effective is solid state isotope exchange with gaseous tritium. This method was used to produce [3H] nicotinamide adenine dinucleotide (111 Ci/mmol), [3H] coenzyme A (3.9 Ci/mmol) and D‐[G‐3H] pantothenic acid (43 Ci/mmol). It was shown that most of the tritium in the labeled nicotinamide adenine dinucleotide and coenzyme A was localized in the nicotinamide (98%) and adenine (89%) sites, respectively. For synthesis of coenzymes labelled with tritium at other sites we developed enzymatic methods which used labelled precursors. Optimum conditions for enzymatic synthesis of [adenine‐3H] nicotinamide adenine dinucleotide from [2,8‐3H] ATP and [pantothenate‐3H] coenzyme A from D‐[G‐3H] pantothenic acid were determined. The tritium labelled acetyl coenzyme A was synthesized by acetylation of labelled coenzyme A with acetic anhydride. The methods chosen allow one to produce tritium labelled coenzymes at high specific activity. Copyright © 2003 John Wiley & Sons, Ltd.