In the present study, a new ultrasonic-assisted cloud point extraction (UA-CPE) approach combined with spectrophotometry was developed for quantification of 5-HMF in selected honey and beverage samples. The method is based on selective ion-association complex formation of 5-HMF with p-nitrophenylhydrazine (p-NPH) in presence of sodium dodecyl sulfate (SDS) using Triton X-45 as extracting agent at pH 4.0. The UA-CPE approach was successfully used for the separation and preconcentration of 5-HMF from matrix. Then, the method was applied successfully to determination of trace 5-HMF in the selected samples after separation and preconcentration with UA-CPE.