The hydrolysates from pretreatment Steps through were collected

2.2.3. Step 3: microwave pretreatment (MP)
2.2.4. Step 4: microorganism decomposition (MD)
2.2.5. Step 5: ammonia immersion (AI)
The residual solids from Step 4 (Solid residue AN1 and AN2, Solid residue TR1 and TR2, and Solid residue PJ1 and PJ2) were immersed in a beaker containing an ammonia solution, with a mass ratio of 2:1 for ammonia to sugarcane. The beakers were subjected to autoclave at 90 °C for 30 min. Then the LY2886721 of the filtered hydrolysis solution was neutralized to 7.0 with 1% H2SO4 and retained for fermentation experiments. The residual solids (Solid residue AAN, ATR, and APJ) were dried at 65 °C and their solid loads were determined. These samples were then used in the next step of decomposition. After Solid residue AN2, TR2, and PJ2 were pretreated by Step 5, the solid–liquid mixtures were neutralized and kept as Solid–liquid mixture AAN, ATR, and APJ, respectively.
2.2.6. Step 6: enzymatic hydrolysis (EH)
2.3. Microorganism and inoculum preparation