Simplified determination of the content and average degree of acetylation of chitin in crude black soldier fly larvae samples.

Insects are considered a promising alternative protein source for food and feed, but contain significant amounts of chitin, often undesirable due to indigestibility, disagreeable texture and negative effect on nutrients intake. Fractionation strategies are thus increasingly being applied to isolate and valorize chitin separately. The analysis of chitin generally requires an intensive pretreatment to remove impurities, and derivatization to generate sufficient detector response. In this work, a liquid chromatography method, without pretreatment nor derivatization, was developed for the simultaneous determination of chitin content and degree of acetylation in non-purified samples of black soldier fly (BSF) larvae. The method is found to be more suitable, compared to traditional methods, for assessing high degrees of acetylation. For the first time, the degree of acetylation of BSF chitin (81 ± 2%) is reported. Additionally, the chitin content of BSF soft tissues is estimated at approximately 20% of the total chitin content (8.5 ± 0.1%).

Continuous production of pectic oligosaccharides from sugar beet pulp in a cross flow continuous enzyme membrane reactor.

Sugar beet pulp pectin is an attractive source for the production of pectic oligosaccharides, an emerging class of potential prebiotics. The main aim of the present work was to investigate a new process allowing to produce pectic oligosaccharides in a continuous way by means of a cross flow enzyme membrane reactor while using a low-cost crude enzyme mixture (viscozyme). Preliminary experiments in batch and semi-continuous setups allowed to identify suitable enzyme concentrations and assessing filtration suitability. Then, in continuous experiments in the enzyme membrane reactor, residence time and substrate loading were further optimized. The composition of the obtained oligosaccharide mixtures was assessed at the molecular level for the most promising conditions and was shown to be dominated by condition-specific arabinans, rhamnogalacturonans, and galacturonans. A continuous and stable production was performed for 28.5 h at the optimized conditions, obtaining an average pectic oligosaccharide yield of 82.9 ± 9.9% (w/w), a volumetric productivity of 17.5 ± 2.1 g/L/h, and a specific productivity of 8.0 ± 1.0 g/g E/h. This work demonstrated for the first time the continuous and stable production of oligosaccharide mixtures from sugar beet pulp using enzyme membrane reactor technology in a setup suitable for upscaling.

Supercritical CO2 Extraction of Nannochloropsis sp.: A Lipidomic Study on the Influence of Pretreatment on Yield and Composition.

Algal lipids have gained wide interest in various applications ranging from biofuels to nutraceuticals. Given their complex nature composed of different lipid classes, a deep knowledge between extraction conditions and lipid characteristics is essential. In this paper, we investigated the influence of different pretreatments on lipid extraction with supercritical CO2 by a lipidomic approach. Pretreatment was found to double the total extraction yield, thereby reaching 23.1 wt.% comparable to the 26.9 wt.% obtained with chloroform/methanol. An increase in acylglycerides was concurrently observed, together with a nearly doubling of free fatty acids indicative of partial hydrolysis. Moreover, an alteration in the distribution of glyco- and phospholipids was noted, especially promoting digalactosyldiglycerides and phosphatidylcholine as compared to monogalactosyldiglycerides and phosphatidylglycerol. At optimized conditions, supercritical CO2 extraction provided a lipid extract richer in neutral lipids and poorer in phospholipids as compared to chloroform/methanol, though with a very similar fatty acid distribution within each lipid class.

Continuous production of pectic oligosaccharides from onion skins with an enzyme membrane reactor.

The aim of this research was to valorize onion skins, an under-utilized agricultural by-product, into pectic oligosaccharides (POS), compounds with potential health benefits. To achieve high hydrolysis performance with the multi-activity enzyme Viscozyme L, an innovative approach was investigated based on a cross-flow continuous membrane enzyme bioreactor (EMR). The influence of the various process conditions (residence time, enzyme concentration, substrate concentration) was investigated on productivity and yield. The composition of the POS mixtures in terms of mono- and oligosaccharides was assessed at the molecular level. At optimized conditions, a stable POS production with 22.0g/L/h volumetric productivity and 4.5g/g POS/monosaccharides was achieved. Compared to previous results obtained in batch for the enzyme Viscozyme L, EMR provided a 3-5× higher volumetric productivity for the smallest POS. Moreover, it gave competitive results even when compared to batch production with a pure endo-galacturonase enzyme, demonstrating its feasibility for efficient POS production.

Enzymatic production of pectic oligosaccharides from onion skins.

Onion skins are evaluated as a new raw material for the enzymatic production of pectic oligosaccharides (POS) with a targeted degree of polymerization (DP). The process is based on a two-stage process consisting of a chelator-based crude pectin extraction followed by a controlled enzymatic hydrolysis. Treatment of the extracted crude onion skin's pectin with various enzymes (EPG-M2, Viscozyme and Pectinase) shows that EPG-M2 is the most appropriate enzyme for tailored POS production. The experiments reveal that the highest amount of DP2 and DP3 is obtained at a time scale of 75-90min with an EPG-M2 concentration of 26IU/mL. At these conditions the production amounts 2.5-3.0% (w/w) d.m for DP2 and 5.5-5.6% (w/w) d.m for DP3 respectively. In contrast, maximum DP4 production of 5.2-5.5% (w/w) d.m. is obtained with 5.2IU/mL at a time scale of 15-30min. Detailed LC-MS analysis reveals the presence of more methylated oligomers compared to acetylated forms in the digests.

Enzymatic electrosynthesis of formate through CO2 sequestration/reduction in a bioelectrochemical system (BES).

Bioelectrochemical system (BES) was operated using the enzyme formate dehydrogenase as catalyst at cathode in its free form for the reduction of CO2 into formic acid. Electrosynthesis of formic acid was higher at an operational voltage of -1V vs. Ag/AgCl (9.37mgL(-1) CO2) compared to operation at -0.8V (4.73mgL(-1) CO2) which was strongly supported by the reduction catalytic current. Voltammograms also depicted a reversible redox peak throughout operation at -1V, indicating NAD(+) recycling for proton transfer from the source to CO2. Saturation of the product was observed after 45min of enzyme addition and then reversibility commenced, depicting a lower and stable formic acid concentration throughout the subsequent time of operation. Stability of the enzyme activity after immobilization on the electrode and product yield will be studied further.

Kinetics of dechlorination by Dehalococcoides mccartyi using different carbon sources.

Stimulated anaerobic dechlorination is generally considered a valuable step for the remediation of aquifers polluted with chlorinated ethenes (CEs). Correct simulation and prediction of this process in situ, however, require good knowledge of the associated biological reactions. The aim of this study was to evaluate the dechlorination reaction in an aquifer contaminated with trichloroethene (TCE) and its daughter products, discharging into the Zenne River. Different carbon sources were used in batch cultures and these were related to the dechlorination reaction, together with the monitored biomarkers. Appropriate kinetic formulations were assessed. Reductive dechlorination of TCE took place only when external carbon sources were added to microcosms, and occurred concomitant with a pronounced increase in the Dehalococcoides mccartyi cell count as determined by 16S rRNA gene-targeted qPCR. This indicates that native dechlorinating bacteria are present in the aquifer of the Zenne site and that the oligotrophic nature of the aquifer prevents a complete degradation to ethene. The type of carbon source, the cell number of D. mccartyi or the reductive dehalogenase genes, however, did not unequivocally explain the observed differences in degradation rates or the extent of dechlorination. Neither first-order, Michaelis-Menten nor Monod kinetics could perfectly simulate the dechlorination reactions in TCE spiked microcosms. A sensitivity analysis indicated that the inclusion of donor limitation would not significantly enhance the simulations without a clear process understanding. Results point to the role of the supporting microbial community but it remains to be verified how the complexity of the microbial (inter)actions should be represented in a model framework.

Temporal variations in natural attenuation of chlorinated aliphatic hydrocarbons in eutrophic river sediments impacted by a contaminated groundwater plume.

Chlorinated aliphatic hydrocarbons (CAHs) often discharge into rivers as contaminated groundwater baseflow. Biotransformation, sorption and dilution of CAHs in the impacted river sediments have been reported to reduce discharge, but the effect of temporal variations in environmental conditions on the occurrence and extent of those processes in river sediments is largely unknown. We monitored the reduction of CAH discharge into the Zenne River during a 21-month period. Despite a relatively stable influx of CAHs from the groundwater, the total reduction in CAH discharge from 120 to 20 cm depth in the river sediments, on average 74 ± 21%, showed moderate to large temporal variations, depending on the riverbed location. High organic carbon and anaerobic conditions in the river sediments allowed microbial reductive dechlorination of both chlorinated ethenes and chlorinated ethanes. δ(13)C values of the CAHs showed that this biotransformation was remarkably stable over time, despite fluctuating pore water temperatures. Daughter products of the CAHs, however, were not detected in stoichiometric amounts and suggested the co-occurrence of a physical process reducing the concentrations of CAHs in the riverbed. This process was the main process causing temporal variations in natural attenuation of the CAHs and was most likely dilution by surface water-mixing. However, higher spatial resolution monitoring of flow transients in the riverbed is required to prove dilution contributions due to dynamic surface water-groundwater flow exchanges. δ(13)C values and a site-specific isotope enrichment factor for reductive dechlorination of the main groundwater pollutant vinyl chloride (VC) allowed assessment of changes over time in the extent of both biotransformation and dilution of VC for different scenarios in which those processes either occurred consecutively or simultaneously between 120 and 20 cm depth in the riverbed. The extent of reductive dechlorination of VC ranged from 27 to 89% and differed spatially but was remarkably stable over time, whereas the extent of VC reduction by dilution ranged from 6 to 94%, showed large temporal variations, and was often the main process contributing to the reduction of VC discharge into the river.

Factors determining the attenuation of chlorinated aliphatic hydrocarbons in eutrohic river sediment impacted by discharging polluted groundwater.

This study explored the potential of eutrophic river sediment to attenuate the infiltration of chlorinated aliphatic hydrocarbon (CAH)-polluted groundwater discharging into the Zenne River near Brussels, Belgium. Active CAH biodegradation by reductive dechlorination in the sediment was suggested by a high dechlorination activity in microcosms containing sediment samples and the detection of dechlorination products in sediment pore water. A unique hydrogeochemical evaluation, including a delta2H and delta18O stable isotope approach, allowed to determine the contribution of different abiotic and biotic CAH attenuation processes and to delineate their spatial distribution inthe riverbed. Reductive dechlorination of the CAHs seemed to be the most widespread attenuation process, followed by dilution by unpolluted groundwater discharge and by surface water mixing. Although CAHs were never detected in the surface water, 26-28% of the investigated locations in the riverbed did not show CAH attenuation. We conclude that the riverbed sediments can attenuate infiltrating CAHs to a certain extent, but will probably not completely prevent CAHs to discharge from the contaminated groundwater into the Zenne River.

Batch-test study on the dechlorination of 1,1,1-trichloroethane in contaminated aquifer material by zero-valent iron.

Chlorinated aliphatic hydrocarbons are common groundwater contaminants. One possible remediation option is in-situ reductive dechlorination by zero-valent iron, either by direct injection or as reactive barriers. Chlorinated ethenes (tetrachloroethene: PCE; trichloroethene: TCE) have received extensive attention in this context. However, another common groundwater pollutant, 1,1,1-trichlorethane (TCA), has attracted much less attention. We studied TCA reduction by three types of granular zero-valent irons in a series of batch experiments using polluted groundwater, with and without added aquifer material. Two types of iron were able to reduce TCA completely with no daughter product concentration increases (1,1-dichloroethane: DCA; chloroethane: CA). One type of iron showed slower reduction, with intermediate rise of DCA and CA concentrations. When evaluating the formation of daughter products, the tests on the groundwater alone showed different results than the groundwater plus aquifer batches: DCA did not temporarily accumulate in the batches with added aquifer material, contrary to the batches without added aquifer material. 1,1-dichloroethene (DCE, also present in the groundwater as an abiotic degradation product of TCA) was also reduced slower in the batches without added aquifer material than in the batches with aquifer material. Redox potentials gradually decreased to low values in batches with aquifer material without iron, while the batches with groundwater alone maintained a constant higher redox potential. Either adsorption processes or microbiological activity in the samples could explain these phenomena. Polymerase Chain Reaction (PCR: a targeted gene probe technique) for chlorinated aliphatic compound (CAH)-degrading bacteria confirmed the presence of Dehalococcoides sp. (chloroethene-degraders) but was negative for Desulfobacterium autotrophicum (a known co-metabolic TCA degrader). DCA reduction was rate determining: first-order half-lives of 300-350 h were observed. TCA was fully removed within hours. CA is resistant to reduction by zero-valent iron but it is known to hydrolyze easily. Since CA did not accumulate in our batches, it may have disappeared by the latter mechanism or it may not have formed as a major daughter product.