A sensitive and simple column-switching high-performance liquid chromatographic (HPLC) method with fluorescence detection was developed for the determination of alendronate in human plasma. Alendronate and pamidronate (internal standard, IS) in plasma including $Na_2EDTA$ were precipitated with sodium hydroxide and calcium chloride after deproteinization using 10% trichloroacetic acid solution. The precipitated alendronate and IS were reconstituted by sodium citrate and citric acid and then derivatized with 9-fluorenylmethyl chloroformate. The resulting solution was injected onto an HPLC system consisting of a pretreatment column and an analytical column, which were connected with a six-port switching valve. The assay was linear in the concentration range of 2-100 ng/mL in 1 mL of plasma with high precision and accuracy, and the limit of detection was 0.5 ng/mL. It was successfully applied to evaluate the pharmacokinetic parameters of alendronate in human volunteers following single oral administration. The mean value of maximum alendronate plasma concentration ($C_{max}$) was 37.69 ng/mL, and the mean time to reach the $C_{max}$ ($T_{max}$) was 1.08 h. The area under the plasma concentration-time curve (AUC) and elimination half-life ($T_{1/2}$) were 106.48 ng/mL/h and 1.66 h, respectively.