**4. Bioconversion of silage to biogas**

In exploring the effect of pretreatment with wastes, many studies have explored the structure and hydrolysis degree of pretreated materials, which as an important index to evaluate the advantages and disadvantages of pretreatment [27]. Like physical and chemical pretreatment, biological pretreatment can also loosen the structure of lignocellulose (**Figure 4**), but it cannot be ignored that the change of lignocellulose structure is only a part of the indicators to evaluate the advantages and disadvantages. It is also an important part of the transformation ability of substrates and the type of metabolites. Various organic acids, especially lactic acid, produced by microbial metabolism during anaerobic pretreatment are important intermediates in

### **Figure 4.**

*Scanning electron micrographs of non-pretreated and pretreated dry corn stover.*

the AD process [28], which can improve the biogas yield and accelerate the process of biogas fermentation [29]. In addition, the organic acids can effectively neutralize the ammonia and other basic substances accumulated during AD to ensure that the pH value of the reaction system is maintained in a stable range [30]. This indicates that the bio-pretreatment method which can metabolize organic acids can effectively improve the biogas yield. However, not all biological pretreatments can convert LCB into organic acids in straw, or only have low conversion capacity, which may limit the significant improvement of subsequent gas production [31].

In the process of AD, raw materials are not only digestive substrates but also the source of nutrients for the survival of anaerobic microorganisms. The character of raw materials determines the time and the biogas yield of AD [32]. AD is a coordinated and mutually restricted metabolic process among starch, protein, fat, and lignocellulose. The types and quantities of raw material organic matter play a decisive role in the degradation process, raw material utilization efficiency, and biogas yield of AD. For example, AD with single LCB has the problem of imbalance between carbon and nitrogen. The C/N ratio of straw stalk is close to 70:1, whereas the best C/N ratio for AD is 20–30:1 [33]. Due to the low C/N ratio, ammonia nitrogen inhibition occurs in AD with single pig manure, which makes it difficult to form the optimal growth state required by biogas-producing microbiota [34]. Hydrolysis pretreatment is a limiting step, because the structure of the cell wall of the residual sludge can inhibit the hydrolysis of intracellular degradable substances. Gas production from conventional sludge digestion often requires a longer residence time [35].

To solve the above problems, mixed pretreatment with difficult and easy to decompose organic matter is one of the hotspots in the field of AD in recent years. Mixed AD can not only effectively regulate the nutritional balance of single raw materials but also improve the bioconversion rate of materials [36]. The author carried out anaerobic microbial ensilage pretreatment using dry corn stover mixed with pig manure and residual sludge, respectively, and then investigated the biogas yield on this basis. The gas production rate from total solids (TS) of untreated dry straw was only 296 mL/g, and the gas production index has been greatly improved through mixed microbial ensilage. The effect of the dry corn stover and pig manure microbial ensilage pretreatment was the best, whereby the gas production rate of TS reached 599 mL/g, and the average volumetric gas production rate was 0.86 L/(L·d). The AD of corn stover, excess sludge, and pig manure can be used to alleviate the nitrogen limitation when using silage as the main raw material. Broad bioconversion of such raw materials will play a decisive role in solving the problems of burning straw, residual sludge landfill, and non-point source pollution from livestock and poultry manure.

The combined AD of different silage materials has the following points:

