10 Aug

Effect of Vitamin E on Exhaled Ethane in Cigarette Smokers: Venous blood

Immediately after the ethane was collected and while the cassettes were still very cold, the charcoal from each trap was poured into a 13.27-mL test tube that was quickly sealed with an open screw top housing a 13-mm TFE/silicone septum (Alltech; Deerfield, IL). The two test tubes were heated for 3 min at 250°C and were agitated from time to time to liberate all of the collected ethane. Five milliliters of the headspace gas in each test tube was removed using a calibrated precision sampling syringe (Alltech) and injected into the column of a gas chromatograph (Hewlett-Packard 5890; Palo Alto, CA) connected in line to an integrator (Hewlett-Packard 3393A). The column was a 2-m long glass column packed with a carbon molecular sieve (Carbosphere 60/80; Alltech) maintained isothermal at 220°C. Helium flow rate was 20 mL/min and retention time under these conditions was 5.30 min. Calibration was performed with two known aliquots of an ethane/nitrogen standard (Scotts Specialty Gases; San Ber-nadino, CA) read allergies pills. Back calculation of the total ethane content in the headspace was then determined and the background ethane content of the inspirate was subtracted from the measured value. The mean of the duplicate samples was used for analysis. The total exhaled ethane was determined from the ethane content of the 6-L sample, the subject’s weight, the background ethane content of the inspirate, and expressed as picomoles per minute per kilogram of body weight.
Approximately 15 mL of venous blood was drawn into sodium heparin tubes (Vacutainer; St. Louis, MO) from each subject on each of the four visits. These tubes were put on ice in the absence of light for 20 min to be fully chilled. The whole blood was then spun down in a centrifuge (National Labnet Co; Woodbridge, NJ) at 2,000 RPM for 10 min to isolate the plasma. The plasma was then pipetted into 2-mL microcentrifuge tubes (Fisher Scientific; Los Angeles, CA) wrapped in foil to keep out any light, and stored at —70°C. Concentrations of vitamin E and P-caro-tene were measured as previously described.

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