Nonparametric determination of reference intervals for plasma metanephrine and normetanephrine.

5-or 10CHOTHF) made up, on average, 30% (range, 0 – 69%) of the TF in erythrocytes of individuals with the T/T genotype. The preliminary results for individuals with the C/T genotype (n ϭ 5) indicated that erythrocytes contained only 5CH 3 THF polyglutamates. No THF was detected in the erythrocytes of individuals with either genotype. The discrepancies in findings between our work and that of Bagley and Selhub (12), i.e., we found small amounts of formylated folates in individuals with the C/C genotype and significant amounts of THF in individuals with the T/T genotype, might be explained by the difference in folate extraction. Bagley and Selhub (12) extracted frozen erythrocytes at high pH to prevent enzymatic deconjugation of folate polyglutamates. We used WB lysates generated at low pH (4.0) to obtain folate monoglutamates by the action of plasma pteroylpoly-␥-glutamate hydrolase. Although this is the generally recommended and clinically used procedure for WB folates, it is conceivable that folate metabolism takes place during the lysis process. As a result, some folate forms could be found in conventionally prepared WB lysates that might not be present in native erythrocytes. The THF we find is most likely the dissociation product of 5,10-methylenetet-rahydrofolic acid losing the C-1 group. We analyzed ϳ100 paired serum and WB lysate samples to assess what proportion of folates found in WB originates from serum. In general, we found no FA in WB lysates with the exception of those from two individuals who had serum FA concentrations Ͼ25 nmol/L, which spilled over into the WB lysate. Serum did not have quantifiable amounts of THF or 5,10CHϭTHF. On average , ϳ95% each of 5CH 3 THF and 5CHOTHF in WB lysates originated from erythrocytes, whereas the rest originated from serum. All of the THF and 5,10CHϭTHF found in WB lysates originated from erythrocytes. The presented method measures folate monogluta-mates only. If deconjugation of polyglutamates is incomplete , the TF concentration will be underestimated. Our findings on folate extraction from WB and the comparability of results obtained with different methods will be presented in a subsequent report. This is the first reported method for measuring intact folate monoglutamates in serum and conventionally prepared WB lysates by LC/MS/MS using automated high-throughput SPE. The method displays excellent sensitivity and satisfactory precision, accuracy, and stability of folates during the 5-h SPE procedure. The specificity of this method is a clear advantage when it comes to …