In order to decrease the influence caused by the micellization and the interaction between Sodium Dodecyl Benzene Sulfonate (SDBS) and betaine on determination of SDBS or betaine in aqueous solution. Synchronous fluorescence spectra and spectrochemical method were utilized to analyze the concentrations of SDBS and betaine by adding 1∶1 (molar ratio) hydroxypropyl-β-cyclodextrin (HP-β-CD) according to the amount of SDBS in their complex aqueous solution.The results indicate that the Critical Micelle Concentration (CMC) values of SDBS could be greatly increased with increasing betaine concentration in SDBS and betaine complex aqueous solution. The CMC values of SDBS reduced from 1.256 mmol/L to 1.121, 0.989 and 0.684 mmol/L when the concentrations of betaine respectively increased from 0 to 1.0, 2.0 and 3.0 mmol/L in their complex aqueous solution. The synchronous fluorescence intensities of SDBS could also be easily affected by betaine even in low concentration SDBS solution (the concentration is lower than CMC). Meanwhile, effect of the interferences on the quantitative determination of betaine by spectrochemical method could be obviously increased in the presence of SDBS. Both the interferences of betaine on synchronous fluorescence spectrum of SDBS and the interferences of SDBS on the spectrochemical determination of betaine, could be greatly reduced by adding 1:1 HP-β-CD. The recovery rates of SDBS and betaine respectively changed from 104.2%~109.2% and 119.7%~159.6% to 101.5%~102.5% and 101.5%~103.0% in their binary complex aqueous solution.