**3. Anaerobic microbial pretreatment**

Microbial ensilage under anaerobic conditions can be a good way to avoid the above problems of aerobic pretreatment, which become a reliable method for long-term storage of lignocellulosic biomass (LCB) [21]. Its basic principle is to use the anaerobic fermentation of lactic acid bacteria (LAB) in closed conditions to convert the soluble carbohydrates to organic acids, which inhibit the growth of detrimental microorganisms by a strong drop in pH to values between 3 and 4 [22]. After a year of microbial ensilage, the dry matter (DM) loss of raw materials is as low as 1–5%, while the digestibility is higher than that of dry storage [23], mainly because the degradation of non-structural carbohydrates reduces the biological resistance of LCB [24]. Microbial ensilage can solve the problems of straw collection, preservation, and pretreatment in large scale. Compared with the chemical methods, the technology does not need to consume a lot of energy and recover chemical reagents, nor produce harmful inhibitors in the reaction system. To date, the silage remains one of the main methods for the efficient utilization of LCB.

### **3.1 The steps involved in the anaerobic ensilage formation**

High-quality silage begins with harvesting at an appropriate stage of maturity to maximize the nutrient production. Next, the following management methods are essential for the successful fermentation and preservation of product (**Figure 1**).

### **Figure 1.**

*The pictorial presentation of the steps involved in the ensilage formation.*

### **Figure 2.**

*Large-scale storage in movable microbial ensilage membranes.*

The goal of good fermentation is to maximize the production of lactic acid, thereby reducing the pH value and establishing an environment that is not suitable for the growth of harmful organisms. Good silage is made with less exposure to the air, allowing the *Lactobacillus* to start fermenting quickly. For example, modern biogas engineering adopts the following pretreatment devices to complete the anaerobic fermentation process (**Figure 2**).

According to the main biochemical and microbiological transformations occurring during the process, ensiling can be divided into three phases as follows.

### 1.Aerobic fermentation process

Anaerobic microbial pretreatment is a straw pretreatment technology utilizing microbial fermentation under anaerobic conditions. However, in the process of straw closure, there is more or less oxygen in the straw raw materials, which makes aerobic microorganisms grow and propagate in the first few days of fermentation. Through the activity of these aerobic microbes, a small amount of sugar and oxygen in the straw can be converted into carbon dioxide and water.

Finally, the oxygen becomes less and less until the oxygen content drops to zero. Aerobic microorganisms do not survive.
