# Biodegradation of a Mixed Manure-Lignocellulosic System--A Possibility Study
## 3 Results and Discussion  
### Concentration of Acid-Insoluble Lignin  
For verifying the efficiency of biomass delignification with _Pleurotus ostreatus_, we determined the concentration of acid-insoluble lignin according to the well-known Klasson procedure. The results for mixtures with different mass ratios of chicken manure with sawdust and pre-treated wheat straw (80:20, 50:50) and the same with Miscanthus are presented in Table 1 (abbreviations: CMS:S--chicken manure with sawdust and ordinary wheat straw, CMS:_SP.o._--chicken manure with sawdust and wheat straw pre-treated with _Pleurotus ostreatus_ fungi and CMS:M--chicken manure with sawdust and ordinary Miscanthus; CMS:_MP.o._--chicken manure with sawdust and pre-treated Miscanthus with _Pleurotus ostreatus_ fungi).  
For the control, the concentrations of acid-insoluble lignin were determined in the untreated chicken manure with sawdust (16.42%), wheat straw (19.90%) and Miscanthus (18.43%). After an extensive literature review, it was clear that lignin concentration can vary significantly, depending on the maturity and origin of the crops. Data from the literature about lignin concentration are presented in continuation and compared to our values. Wheat straw contained 19.3, 23, 30 and 20% of lignin, according to [PERSON] et al. [39], [PERSON] et al. [40], [PERSON] et al. [41] and [PERSON] et al. [42], respectively. [PERSON] et al. determined the lignin content of different types of Miscanthus, where values varied from 8.58 to 14.44% [43]. Miscanthus contained 20, 25.2, 18.1 and 21.36% lignin, according to [PERSON] et al. [44], [PERSON] et al. [45], [PERSON] et al. [46] and [PERSON] et al. [47], respectively. [PERSON] et al. performed an additional study of fungal pretreatment of Miscanthus for enhanced enzymatic hydrolysis and obtained lignin concentrations of raw Miscanthus and fungal colonized Miscanthus of 20.9 and 17.1%, respectively [48]. In our test, we compared the values of ordinary Miscanthus to the values of mixtures after incubation with fungi, and we conclude that the lignin concentrations of mixtures are only 3 to 9% lower. Lower values can also be attributed to the lower lignin content of chicken manure and sawdust. [PERSON] et al. found that if the inoculum ratio is more than 30% and the moisture content between 60 to 75%, then, lignin degradation could be 25 to 35% after 28 d. However, they observed almost no lignin degradation with a 10 to 20% inoculum ratio [48].  
Table 1 shows that the values of lignin concentrations are dispersed and that there is no observable interdependence between the concentration of lignin and the mass ratio of the mixtures. Special care was taken in the preparation of mixtures; however, because of variations in the consistency of each component, it was difficult to prepare uniform mixtures. Chicken manure, among other things, contains sawdust and pebbles; pre-treated wheat straw and Miscanthus contain mycelium fungi; consequently, all ingredients are not evenly dispersed. The concentration of acid-insoluble lignin in all Miscanthus samples is 5-10% lower than that in the wheat straw ones. The average deviation in concentration of insoluble lignin in wheat straw and Miscanthus samples were 0.66 and 0.42%, respectively. We chose a time period of 30 d for performing our experiments, because we were focused on possible lignin degradation over a reasonable time span. However, lignin degradation was not evident during the 30 d of incubation.  
The mycelium growth was satisfactory on both substrates, but it was more pronounced on wheat straw samples compared to Miscanthus samples. [PERSON] et al. studied fungal treatment of lignocellulosic biomass and revealed that no mycelium growth of _Pleurotus ostretatus_ was observed on Miscanthus after 4 weeks. The mycelium growth was observed later, and after 8 weeks, the growth was about double [49]. In another study, _Trametes versicolor_ fungi caused a 46% mass loss of lignin in Miscanthus after 12 weeks of incubation at 20 \({}^{\circ}\)C [36].