A mixed substrate (MS) comprising oil palm empty fruit bunch (EFB), oil palm frond (OPF), and rice husk (RH) was evaluated for endoglucanase prodn. by Bacillus aerius S5.2. Effects of sulfuric acid, sodium hydroxide, N-methylmorpholine-N-oxide (NMMO), and hydrothermal pretreatments on endoglucanase prodn. were investigated. Endoglucanase prodn. by B. aerius on the untreated (0.677 U/mL) and pretreated MS (0.305 - 0.630 U/mL) was generally similar, except that the acid (0.305 U/mL) and hydrothermal (0.549 U/mL) pretreatments that were more severe consequently produced significantly lower titers. Alkali pretreatment supported the highest enzyme prodn. (0.630 U/mL) among all pretreatments that were studied. When endoglucanase prodn. on the alkali-pretreated MS and single substrates (SS) was compared, alkali-pretreated EFB produced a titer (0.655 U/mL) similar to the MS, and this was significantly higher than titers recorded on OPF (0.504 U/mL) and RH (0.525 U/mL). Lower enzyme prodn. was found to be consistent with higher pretreatment severity and greater removal of amorphous regions in all the pretreatments. Furthermore, combining the SS showed no adverse effects on endoglucanase prodn. [on SciFinder(R)]