In the present study, alginate nanoparticles were firstly prepared for paclitaxel (PTX) delivery with an average size of 200 ± 21 nm. To improve the stability and targeting effect, the chitosan (CS) and folate-chitosan (FA-CS) were introduced to form PTX-loaded CS/ALG NPs and FA-CS/ALG NPs by a new double emulsion cross-linking electrostatic attraction method. The optimization chitosan concentration was 0.5% obtained from the experiment results. The CS/ALG-PTX NPs and FA-CS/ALG-PTX NPs had the average particle size of 306.9 ± 12.9 nm and 283.6 ± 19.2 nm with the zeta potential of 31.1 ± 1.3 mV and –2.98 ± 0.7 mV, and had higher drug loading and entrapment efficiencies than ALG-PTX NPs. The in vitro drug release profile along with release kinetics and mechanism from PTX-loaded NPs were studied under two simulated physiological conditions. Further, the in vitro anti-cancer activity of nanoparticles and the cellular uptake of nanoparticles on HepG2 cells were investigated. The results demonstrated that alginate, CS/ALG and FA-CS/ALG can be used as nanoformulation drug carriers by our new method, and FA-CS/ALG was a promising vehicle for anticancer drug targeted delivery system.