Determination of adenosylhomocysteine in urine of immunodeficient children

A sensitive anion exchange separative procedure is described for the determination of adenosylhomocysteine in urine. Anionic components of the urine are first removed at a pH of 5 on a short anion exchange column, formate form. The column wash is then adjusted to pH 10 and applied to a longer anion exchange column, bicarbonate form. Elution is carried out with a sodium chloride gradient with the eluant solution containing 0.01 m sodium bicarbonate and 0.01 m sodium tetraborate. Adenosylhomocysteine forms a borate complex, giving an additional negative charge, and the separation of adenosylhomocysteine from adenine is enhanced. Adenine and adenosylhomocysteine in eluant fractions are detected after conversion to the fluorescent etheno derivatives. Using this assay procedure, we have analyzed urine specimens from seven children with severe combined immunodeficiency diseases and from three children with milder immunodeficiency disorders. Three of the children with severe combined immunodeficiency had excretion levels of adenosylhomocysteine (0.6-1.8 nmole/μmole creatinine) that were more than three standard deviations above the excretion levels noted for normal children (0.14 ± 0.07 nmole/μmole creatinine). The possible significance of these findings is discussed.