TABLE 1.

Extraction efficacy and variance during sample processing

	γ-CEHC	α-CEHC	α-CMBHC	δTE-13′	δT-13′	γT	δT
Efficacy (%)	97	89	96	98	95	107	101
Intraday CV%	9	4	7	4	3	7	6
Interday CV%	3	11	6	10	5	11	8

CV, coefficient of variation. Tocopherols and metabolites were spiked into FBS to yield γ-CEHC (2 µM), α-CEHC (10 µM), α-CMBHC (2 µM), δTE-13′-COOH (1 µM), δT-13′-COOH (0.4 µM), γT (0.4 µM), and δT (0.4 µM). One hundred microliters of spiked FBS was extracted by 600 μl working methanol (with 0.2 mg/ml ascorbic acid) and 1.2 ml hexane via vortexing vigorously for 1 min. After centrifugation for 2 min at 10,000 rpm, the upper hexane layer was transferred into a tube with preadded 10 µl BHT (10 mg/ml). The methanol layer was transferred into a clean tube, and the pellet was extracted with 400 µl working methanol for the second time. The two methanol extractions were combined and dried under N₂. The dried methanol and hexane-extracted samples were resuspended in 100 µl working methanol before being analyzed by HPLC with fluorescent detection. Data are expressed as mean ± SD (based on >4 independent experiments).