Natural epothilone B (EPOB) is currently in clinical trials for treatment of advanced cancers. In this study, two poly(ethylene glycol) (PEG)–EPOB conjugates were synthesized with carbodiimide chemistry with linear PEG Methoxy-PEG-Carboxymethy(mPEG-COOH) with different molecular weights (5 and 20 kDa). The products were confirmed by matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy and 1H-NMR, which showed that PEGylation only took place at the 7-OH site of EPOB. The solubilities of PEG5K–EPOB the conjugate of mPEG-COOH (MW 5,000) and epothilone B and PEG20K–EPOB the conjugate of mPEG-COOH (MW 20,000) and epothilone B were determined to be 4.93 × 10−2 and 1.58 × 10−2 mmol/mL; this showed improvements of 35 and 11 times, respectively, over that of free EPOB (1.4 × 10−3 mmol/mL). Moreover, the conjugates were more stable than that of free EPOB in plasma. The cytotoxicity of conjugates was evaluated on human breast cancer MCF-7 cells with an 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di- phenytetrazoliumromide(MTT) based assay. The half maximal inhibitory concentration of a substance(IC50) values of EPOB, PEG5K–EPOB, and PEG20K–EPOB were 6.0 × 10−4, 0.57, and 8.4 × 10−3 μM, respectively. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014, 131, 41123.