Isolation and Screening of Microorganisms for the Effective Pretreatment of Lignocellulosic Agricultural Wastes

Lignocellulosic waste is the most abundant biorenewable biomass on earth, and its hydrolysis releases highly valued reducing sugars. However, the presence of lignin in the biopolymeric structure makes it highly resistant to solubilization thereby hindering the hydrolysis of cellulose and hemicellulose. Microorganisms are known for their potential complex enzymes that play a dominant role in lignocellulose conversion. Therefore, the current study was designed to isolate and screen potential microorganisms for their selective delignification ability for the pretreatment of lignocellulosic biomass. An extensive isolation and screening procedure yielded 36 desired isolates (22 bacteria, 7 basidiomycete fungi, and 7 filamentous fungi). Submerged cultivation of these desired microorganisms revealed 4 bacteria and 10 fungi with potent lignocellulolytic enzyme activities. The potent isolates were identified as Pleurotus, Trichoderma, Talaromyces, Bacillus, and Chryseobacterium spp. confirmed by morphological and molecular identification. The efficiency of these strains was determined through enzyme activities, and the degraded substrates were analyzed through scanning electron microscopy (SEM) and X-ray diffraction (XRD). Among all isolated microbes, Pleurotus spp. were found to have high laccase activity. The cellulose-decomposing and selective delignification strains were subjected to solid-state fermentation (SSF). SSF of field waste corn stalks as a single-carbon source provides Pleurotus spp. better condition for the secretion of ligninolytic enzymes. These isolated ligninolytic enzymes producing microorganisms may be used for the effective pretreatment of lignocellulosic agricultural wastes for the production of high value-added natural products by fermentation.

Alkaline post-incubation improves the saccharification of poplar after hydrogen peroxide–acetic acid pretreatment

Background Hydrogen peroxide–acetic acid (HPAA) is widely used in pretreatment of lignocellulose because it has a good capability in selective delignification. However, high concentration (more than 60%) of HPAA increases the cost of pretreatment and the risk of explosion. In this work, alkaline post-incubation was employed to decrease the HPAA loading and improve the saccharification of poplar. Results Pretreatment with 100% HPAA removed 91.0% lignin and retained 89.9% glucan in poplar. After poplar was pretreated by 100% HPAA at 60 °C for 2 h, the glucan conversion in enzymatic hydrolysis by cellulase increased to 90.1%. Alkaline incubation reduced the total lignin, surface lignin, and acetyl group of HPAA-pretreated poplar. More than 92% acetyl groups of HPAA-pretreated poplar were removed by alkaline incubation with 1.0% NaOH at 50 °C for 1 h. After incubation of 60% HPAA-pretreated poplar with 1.0% NaOH, the glucan conversion enhanced to 95.0%. About 40% HPAA loading in pretreatment was reduced by alkaline incubation without the decrease of glucose yield. Conclusions Alkaline post-incubation had strong ability on the deacetylation and delignification of HPAA-pretreated poplar, exhibiting a strong promotion on the enzymatic hydrolysis yield. This report represented alkaline incubation reduced the HPAA loading, improved pretreatment safety, exhibiting excellent potential application in saccharification of poplar.

Generation of highly amenable cellulose-Iβ via selective delignification of rice straw using a reusable cyclic ether-assisted deep eutectic solvent system

Cellulolytic enzymes can readily access the cellulosic component of lignocellulosic biomass after the removal of lignin during biomass pretreatment. The enzymatic hydrolysis of cellulose is necessary for generating monomeric sugars, which are then fermented into ethanol. In our study, a combination of a deep eutectic (DE) mixture (of 2-aminoethanol and tetra-n-butyl ammonium bromide) and a cyclic ether (tetrahydrofuran) was used for selective delignification of rice straw (RS) under mild conditions (100 °C). Pretreatment with DE-THF solvent system caused ~ 46% delignification whereas cellulose (~ 91%) and hemicellulose (~ 67%) recoveries remained higher. The new solvent system could be reused upto 10 subsequent cycles with the same effectivity. Interestingly, the DE-THF pretreated cellulose showed remarkable enzymatic hydrolysability, despite an increase in its crystallinity to 72.3%. Contrary to conventional pretreatments, we report for the first time that the enzymatic hydrolysis of pretreated cellulose is enhanced by the removal of lignin during DE-THF pretreatment, notwithstanding an increase in its crystallinity. The current study paves way for the development of newer strategies for biomass depolymerization with DES based solvents.

Polyoxometalate catalysts for biomass dissolution: understanding and design

The use of polyoxometalate catalysts for selective delignification of biomass presents a possible route toward using ionic liquids (ILs) to efficiently obtain high-molecular weight biopolymers from biomass. Rapid progress in this area will depend on recognizing and using the link with already well-developed inorganic chemistry in ILs pursued outside the field of biomass processing. Here, we use crystal structures determined from single crystal X-ray diffraction to better understand the behavior of [PV2Mo10O40]5-, a polyoxometalate catalyst known for its ability to promote selective delignification of biomass in the IL 1-ethyl-3-methylimidazolium acetate ([C2mim][OAc]). The crystal structure of [C2mim]5[PV2Mo10O40]·THF shows the formation of cationic shells around the anions which are likely representative of the interactions of this catalyst with [C2mim][OAc] itself. The reaction of NH4VO3 with [C2mim][OAc] is explored to better understand the chemistry of vanadium(V), which is critical to redox catalysis of [PV2Mo10O40]5-. This reaction gives crystals of [C2mim]4[V4O12], showing that this IL forms discrete metavanadates which are obtained from aqueous solutions in a specific pH range and indicating that the basicity of [OAc]- dominates the speciation of vanadium (V) in this IL.

Characterization of Schinopsis Haenkeana wood Decayed by Phellinus Chaquensis (Basidiomycota, Hymenochaetales)

Schinopsis haenkeana is a native tree to the Chaco Serrano Forests in Argentina. The white-rot fungus Phellinus chaquensis degrades its wood, causing a whiterot type of decay. The objective of this study was to investigate the structural alterations caused by P. chaquensis in S. hankeana decayed naturally and in vitro. Sound living branches with decay and basidiocarps of P. chaquensis were sampled from the field and in vitro decay tests were performed according to the ASTM D-2017-81 standard method. Naturally decayed branches exhibited an innermost discolored zone with white-rot decay and an outer yellowish-white portion of sound sapwood. Using LM and SEM, degraded tissue displayed diagnostic characters of selective delignification and simultaneous decay. Findings indicate that P. chaquensis causes a mottled pattern of decay (selective delignification plus simultaneous decay) in S. haenkeana wood. Other features such as accumulation of extractives, profuse deposition of crystals and tyloses, typical ofSchinopsis spp. heartwood, were additionally observed. In laboratory degraded material, signs of selective delignification and incipient stages of simultaneous decay were noticeable only microscopically. Chemical analysis revealed an oxidative alteration of aromatic moieties in naturally decayed samples which might be related to the accumulation of phenols as a response to fungal attack when compared to sound samples. Naturally degraded sapwood exhibits anatomical and chemical modifications that indicate the development of discolored wood derived from the host-pathogen interaction.

Effect of Physisporinus vitreus on wood properties of Norway spruce. Part 2: Aspects of microtensile strength and chemical changes

The biotechnological application of the white rot fungusPhysisporinus vitreusnamed “bioincising” is currently being investigated for permeability improvement of Norway spruce (Picea abies(L.) Karst.) wood. During short-term (<9 weeks) incubation, fungal activity induces degradation of pit membranes and a simultaneous alteration of the tracheid cell wall structure. In Part 1 of this article series, the occurrence of selective delignification and simultaneous degradation was shown by UV-microspectrophotometry (UMSP). Moreover, significant reduction of Brinell hardness was recorded after 7 and 9 weeks incubation. For a better understanding of the chemical alterations in the wood constituents and the corresponding changes of mechanical properties due to fungal activity, we applied microtensile tests on thin strips that were prepared from the surface of incubated Norway spruce wood. Indications for the occurrence of selective delignification and simultaneous degradation were evident. Determination of lignin content and carbohydrate analysis by borate anion exchange chromatography confirmed the results. The present study verifies the findings from Part 1 of this article series and from previously conducted microscopic investigations. Now, the degradation characteristics ofP. vitreusare established and the bioincising process can be further optimized with higher reliability.