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Industrial utilization of lignin is of high interest since it represents around 30% of all nonfossil-based carbon sources worldwide. For various applications of lignosulfonates such as for dispersants or adhesives a larger molecular weight is essential. Here, we investigated laccase-catalyzed polymerization of lignosulfonate directly from the pulp and paper industry in the presence and absence of natural and synthetic mediators with and without oxygen supply. For example, laccase-mediated polymerization in the presence of a 2.5 mM TEMPO as mediator with a 10 cm 3 min −1 oxygen flow rate led to a 12-fold increase of the molecular weight, while without TEMPO a 13-fold increase was achieved. In contrast, without an external oxygen supply, only a 7-fold increase in molecular weight was achieved compared to a 4-fold increase for the TEMPO−laccase system. Fluorescence intensity, phenol content, and size exclusion chromatography measurements indicate that generally in the presence of high concentrations of mediators, such as TEMPO, vanillin, HBT, and 2,6-dimethoxyphenol, oxidation of other structural units in lignosulfonates may counteract desired polymerization reactions. In summary, for laccase-catalyzed polymerization of lignosulfonates, an external oxygen supply was found to be much more beneficial than the presence of laccase mediators.

A sensitive method for quantification of citric, fumaric, malic, malonic, oxalic, trans aconitic, and succinic acid in soil- and root-related samples is presented. The method is based on a novel, fast, and simple esterification procedure and subsequent analysis via liquid chromatography-mass spectrometry. Derivatization comprises in situ generation of HCl, which catalyzes the Fischer esterification with benzyl alcohol. As a key advance, the esterification with the aromate allows reversed-phase separation and improves electrospray ionization efficiency. The method provided procedural detection limits of 1 nM for citric, 47 nM for fumaric, 10 nM for malic, 10 nM for malonic, 16 nM for oxalic, 15 nM for succinic, and 2 nM for aconitic acid utilizing 500 μL of liquid sample. The working range was 3 nM to 10 μM for citric acid, 158 nM to 10 μM for fumaric acid, 34 nM to 10 μM for malic acid, 33 nM to 10 μM for malonic acid, 53 nM to 10 μM for oxalic acid, 48 nM to 10 μM for succinic acid, and 6 nM to 10 μM for aconitic acid. Quantification of the analytes in soil-related samples was performed via external calibration of the entire procedure utilizing (13)C-labeled oxalic and citric acid as internal standards. The robustness of the method was tested with soil extracts and samples from hydroponic experiments. The latter concerned the regulation of phosphorus solubilization via plant root exudation of citric, malic, and oxalic acid.

A bioactive O-carboxymethyl chitosan (CMCS) hydrogel crosslinked with natural phenolics with potential application in wound dressings was synthesized using a laccase from Myceliophthora thermophila (MTL). The highest degree of cross-linking (49.7%) was achieved with catechol. All the phenolic-CMCS hydrogels synthesized showed excellent anti-oxidant properties with a free radical scavenging activity up to 4-fold higher than in the absence of the phenolics, as quantified by the di(phenyl)-(2,4,6-trinitrophenyl)iminoazanium (DPPH) assay. In addition, the hydrogels produced showed an anti-inflammatory effect as evidenced by the inhibition of enzymes [myeloperoxidase (MPO), matrix-metalloproteinase-1 (MMP-1) and human neutrophil elastase (HNE)] over-expressed in chronic wounds. Sinapyl-CMCS hydrogels showed an MMP-1 inhibition of 37%. Further, the phenolic-CMCS hydrogels did not affect the viability of the NIH 3T3 mouse fibroblast cell line and were also able to slowly release human fibroblast growth factor 2, reaching 48.3% over a period of 28days. This study thus shows the possibility of synthesizing multifunctional bioactive chitosan based hydrogels with anti-oxidant and anti-inflammatory properties using natural occurring phenolics as crosslinkers.

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The aging population and accompanying diseases like diabetes resulted in an increased occurrence of chronic wounds. Topical wound treatment with antimicrobial agents to inhibit bacterial invasion and promote wound healing is often associated with difficulties. Here, we investigated the potential of succinyl chitosan (SC)-carboxymethyl cellulose (CMC) hydrogels which constantly release clinically relevant levels of hydrogen peroxide (HO). CMC hydrogel matrix was in situ converted by limited hydrolysis by a cellulase into substrates accepted by cellobiose dehydrogenase (CDH) for continuous production of HO (30 μM over 24 h). This dual-enzyme catalyzed in situ HO generation system proved its antimicrobial activity in a zone of inhibition (ZOI) assay best simulating the application as wound dressing and was found to be biocompatible toward mouse fibroblasts (95% viability). The hydrogels were thoroughly characterized regarding their rheological properties indicating fast gel formation (<3 min) and moderate cross-linking (1.5% strain, G' = 10 Pa). Cooling (fridge conditions) was found to be the simple on/off switch of the enzymatic machinery which is of great importance regarding storage and applicability of the bioactive hydrogel. This robust and bioactive antimicrobial hydrogel system overcomes dosing issues of common topical wound treatments and constitutes a promising wound healing approach for the future.

Enzymatic polymerization of lignin is an environmentally-friendly and sustainable method that is investigated for its potential in opening-up new applications of one of the most abundant biopolymers on our planet. In this work, the laccase from Myceliophthora thermophila was successfully immobilized onto Accurel MP1000 beads (67% of protein bound to the polymeric carrier) and the biocatalyzed oxidation of Kraft lignin (KL) and lignosulfonate (LS) were carried out. Fluorescence intensity determination, phenol content analysis and size exclusion chromatography were performed in order to elucidate the extent of the polymerization reaction. The collected results show an 8.5-fold decrease of the LS samples' fluorescence intensity after laccase-mediated oxidation and a 12-fold increase of the weight average molecular weight was obtained.

Biochar has a number of environmental applications, including soil amendment for agriculture, remediation of contaminated soils and sediments, and climate change mitigation. Dust formed during its production and field application may pose a health risk, but the cytotoxicity of biochar has, to the best of our knowledge, not previously been investigated. Therefore, we measured the concentration-dependent cytotoxicity of biochar on an NIH 3T3 mouse fibroblast cell line. We used a contaminant trap experiment to measure the total and nondesorbable polycyclic aromatic hydrocarbon (PAH) fractions of the biochar. PAH release was found to be negligible because of the biochar’s strong PAH sorption potential. The biochar was nevertheless observed to have a cytotoxic effect on the fibroblast cells; the EC10 values were 49.6 and 18.8 μg/mL after incubation for 24 and 48 h, respectively. This cytotoxic effect is likely to relate to the particulate nature and size distribution of the biochar; the biochar had particles similar in size to atmospheric particulate matter (PM2.5) that bound to the fibroblast cell surface. To minimize the risk of exposure, practitioners should wear respiratory protective equipment during biochar production. During field application, biochar should be applied in slurries and should always be mixed with a soil matrix to avoid secondary dust formation.