A novel method for the determination of ethanol in tequila based on the immobilized enzyme fluorescence capillary analysis (IE-EFCA) has been proposed. Alcohol dehydrogenase (ADH) was immobilized in inner surface of a capillary and an immobilized enzyme capillary bioreactor (IE-ECBR) was formed. After nicotinamide adenine dinucleotide (NAD(+)) as an oxidizer is mixed with alcohol sample solution, it was sucked into the IE-ECBR. The fluorescence intensity of the mixed solution in the IE-ECBR was detected at lambda(ex)=350 nm and lambda(em)=459 nm. The experimental conditions are as follows: The reaction time is 20 min; temperature is 40 degrees C; the concentrations of phosphate buffer solution (pH 7.5) and NAD(+) are 0.1 mol L(-1) and 5 mmol L(-1), respectively; immobilization concentration of ADH is 10 U L(-1). The determination range of ethanol is 2.0-15.0 g L(-1) (F=10.44C+6.6002, r>0.9958); its detection limit is 1.11 g L(-1); and relative standard deviation is 1.9%. IE-EFCA method is applicable for the determination of the samples containing alcohol in medicine, industry and environment.