Silicon nanoparticle-assisted laser desorption/ionization (SPALDI) is shown to be a selective ionization method with low detection limits, high salt tolerance and reduced background in the low mass region. In order to study the internal energy deposition during SPALDI, five benzylpyridinium chloride salts are ionized by laser desorption/ionization (LDI) assisted by derivatized and untreated silicon nanoparticles. The fragmentation behavior of those labile benzylpyridinium salts in SPALDI is compared to that in regular MALDI with α-cyano-4-hydroxycinnamic acid (CHCA) as the matrix. Silicon nanopowder with a diameter of 30 nm is derivatized by fluorinated alkylchlorosilanes and used as a matrix in the LDI of the benzylpyridinium salts. The fragmentation behavior of benzylpyridinium salts during LDI is studied versus a series of laser fluence values. Both derivatized and untreated silicon nanopowders need less laser fluence than CHCA to produce benzylpyridinium ions. SPALDI with derivatized silicon powder as a matrix produces "cooler" ions than that with untreated silicon powder. The degree of fragmentation of the benzylpyridinium salts in SPALDI exhibits a dependency on the chain length of the surface derivatives. Fatty acids with different chain lengths and concentrations are also analyzed by SPALDI mass spectrometry to explore the practical uses of SPALDI in the negative ion mode. A chain-length effect in SPALDI MS is observed for fatty acid ionization. Derivatized silicon nanopowder provides detection limits of fatty acids down to ~300 pmol/μL. Compared to the derivatized powder, untreated silicon nanopowder gives much lower detection limits of fatty acids (~50 pmol/μL) but more background.