Two new spliceostatin derivatives, designed as spliceostatin H (1) and spliceostatin I (2), and one known compound FR901464 (3), were isolated from the strain Pseudomonas sp. HS-NF-1408. Their structures were determined by the comprehensive spectroscopic data, including 1D, 2D NMR, MS spectral analysis and comparison with data from the literature. Compound 1 exhibited potent cytotoxicity activity against A549 and HepG2 with IC