The enzymatic preparation and chromatographic purification of [4-3H]NADP and NADPH stereospecifically labeled with 3H on either the A or B faces at position 4 have been simplified. Commercially available [1-3H]glucose was used as a starting material for the sequential synthesis of [4B-3H]NADPH, [4-3H]NADP, and [4A-3H]NADPH. These products were rapidly purified by step elution of DEAE-cellulose minicolumns so that [4B-3H]NADPH was produced and purified from [1-3H]glucose in 2 h, [4-3H]NADP in 5 h, and [4A-3H]NADPH in 8 h. Yields of these products were 65 to 88% starting with [1-3H]glucose. Analysis of the products by high-performance liquid chromatography indicated radiochemical purities of 82-95% for these compounds and specific activities equivalent to that of the starting material (10-15 Ci/mmol).
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