The hexagonal NaYF4:Yb3+, Er3+ upconversion fluorescent materials with fibrous hollow structure were synthetized by one-pot hydrothermal method with water-soluble poly(ethylene imine) (PEI) as ligand. The as-prepared NaYF4:Yb3+, Er3+ upconversion materials were used as fillers which incorporated into chitosan/polyvinyl alcohol (CS/PVA) matrix to develop a novel luminescent composite film. The effects of PEI ligand amounts and pH values of reaction system on the crystal phase, morphology and fluorescent properties were investigated in detail. The doping concentration of NaYF4:Yb3+, Er3+ filler in CS/PVA fluorescent composite film was also optimized. It was found that the resulting hexagonal NaYF4:Yb3+, Er3+ upconversion materials possessed fibrous hollow structure and exhibited excellent upconversion luminescence under 980 nm laser excitation when the PEI contents was 0.3 g at pH=5. When the doping mass fraction of the fluorescent filler was 3.0%, the prepared NaYF4:Yb3+, Er3+/(CS/PVA) fluorescent composite film had the best transparency and up-conversion fluorescent properties.