Structural insights, biocatalytic characteristics, and application prospects of lignin-modifying enzymes for sustainable biotechnology.

Lignin modifying enzymes (LMEs) have gained widespread recognition in depolymerization of lignin polymers by oxidative cleavage. LMEs are a robust class of biocatalysts that include lignin peroxidase (LiP), manganese peroxidase (MnP), versatile peroxidase (VP), laccase (LAC), and dye-decolorizing peroxidase (DyP). Members of the LMEs family act on phenolic, non-phenolic substrates and have been widely researched for valorization of lignin, oxidative cleavage of xenobiotics and phenolics. LMEs implementation in the biotechnological and industrial sectors has sparked significant attention, although its potential future applications remain underexploited. To understand the mechanism of LMEs in sustainable pollution mitigation, several studies have been undertaken to assess the feasibility of LMEs in correlating to diverse pollutants for binding and intermolecular interactions at the molecular level. However, further investigation is required to fully comprehend the underlying mechanism. In this review we presented the key structural and functional features of LMEs, including the computational aspects, as well as the advanced applications in biotechnology and industrial research. Furthermore, concluding remarks and a look ahead, the use of LMEs coupled with computational framework, built upon artificial intelligence (AI) and machine learning (ML), has been emphasized as a recent milestone in environmental research.

Techno-Economic Assessment of Whey Protein-Based Plastic Production from a Co-Polymerization Process.

Bio-based plastics, produced from natural and renewable sources, have been found to be good replacers to petroleum-based plastics. However, economic analyses have not been carried out for most of them, specifically those from whey. In this study, a techno-economic assessment of the industrial-scale production of plastics from whey protein is carried out considering two different scenarios: (1) low-cost dairy waste whey (DWP) and (2) purchased whey protein concentrate (WPC), as feedstocks, using SuperPro Designer software. Key economic indicators such as operating cost, capital investment, annual revenue, payback time, and return-on-investment (ROI), were analyzed. Sensitivity analyses of different parameters were performed to account for market fluctuations and other uncertainties, using Scenario 2 as the base case. Results showed that both scenarios have the capacity of producing over 3200 metric tons/year (t/yr) (or 5.5 t/batch) of plastic. With the unit selling price of plastic set at $7,000/t, both the scenarios showed profitable outcomes with the plant's payback time of 3.7 and 2.4 years, and ROI of 27.1% and 42.2%, for Scenario 1 and Scenario 2, respectively. Sensitivity analyses showed that the unit plastic selling price was the most sensitive parameter, followed by the amount of feedstock WPC, and the number of batches.

Preparation and Characterization of Whey Protein-Based Polymers Produced from Residual Dairy Streams.

The wide use of non-biodegradable, petroleum-based plastics raises important environmental concerns, which urges finding alternatives. In this study, an alternative way to produce polymers from a renewable source-milk proteins-was investigated with the aim of replacing polyethylene. Whey protein can be obtained from whey residual, which is a by-product in the cheese-making process. Two different sources of whey protein were tested: Whey protein isolate (WPI) containing 91% protein concentration and whey protein concentrate (WPC) containing 77% protein concentration. These were methacrylated, followed by free radical polymerization with co-polymer poly(ethylene glycol) methyl ether methacrylate (PEGMA) to obtain polymer sheets. Different protein concentrations in water (11-14 w/v%), at two protein/PEGMA mass-ratios, 20:80 and 30:70, were tested. The polymers made from WPI and WPC at a higher protein/PEGMA ratio of 30:70 had significantly better tensile strength than the one with lower protein content, by about 1-2 MPa (the best 30:70 sample exhibited 3.8 ± 0.2 MPa and the best 20:80 sample exhibited 1.9 ± 0.4 MPa). This indicates that the ratio between the hard (protein) and soft (copolymer PEGMA) domains induce significant changes to the tensile strengths of the polymer sheets. Thermally, the WPI-based polymer samples are stable up to 277.8 ± 6.2 °C and the WPC-based samples are stable up to 273.0 ± 3.4 °C.

Natural antibacterial agents from arid-region pretreated lignocellulosic biomasses and extracts for the control of lactic acid bacteria in yeast fermentation.

Bacterial contamination is one of the major challenges faced by yeast fermentation industries as the contaminating microorganisms produce lactic acid and acetic acid, which reduces the viability of yeast, and hence fermentation yields. The primary bacterial contaminants of yeast fermentations are lactic acid bacteria (LAB). This study aims to identify potential natural antibacterial fractions from raw and pretreated lignocellulosic biomasses found in Abu Dhabi, UAE, in terms of LAB inhibition capacity, allowing growth of the yeast. The analysis was carried out using plating technique. Pretreatment liquid of the mangrove stem Avicennia marina hydrothermally pretreated at 210 °C exhibited the widest inhibition zone with an average diameter of 14.5 mm, followed by the pretreatment liquid of mangrove leaf pretreated at 190 °C, Salicornia bigelovii pretreated at 202 °C and rachis of date palm Phoenix dactylifera pretreated at 200 °C. The compounds responsible for the antibacterial activity will be characterized in further study.

Effect of total solid content and pretreatment on the production of lactic acid from mixed culture dark fermentation of food waste.

Food waste landfilling causes environmental degradation, and this work assesses a sustainable food valorization technique. In this study, food waste is converted into lactic acid in a batch assembly by dark fermentation without pH control and without the addition of external inoculum at 37 °C. The effect of total solid (TS), enzymatic and aeration pretreatment was investigated on liquid products concentration and product yield. The maximum possible TS content was 34% of enzymatic pretreated waste, and showed the highest lactic acid concentration of 52 g/L, with a lactic acid selectivity of 0.6 glactic/gtotalacids. The results indicated that aeration pretreatment does not significantly improve product concentration or yield. Non-pretreated waste in a 29% TS system showed a lactic acid concentration of 31 g/L. The results showed that enzymatic pretreated waste at TS of 34% results in the highest production of lactic acid.

Process simulation and economic assessment of hydrothermal pretreatment and enzymatic hydrolysis of multi-feedstock lignocellulose - Separate vs combined processing.

Biorefinery based on multi-feedstock lignocellulose can be viable where a sustainable supply of a single substrate is limited, for example in arid regions. Processing of mixed feedstocks has been studied in lab scale, however, its economics are less studied. In this study, an economic comparison was made between separate and combined (mixed) processing approaches for multi-feedstock lignocellulose for the production of monomeric sugars. This modular approach of focusing on sugar platform makes the results applicable for many applications using the sugars as feedstock. Feedstock considered in this study were the green and woody lignocellulose residues: Bermuda grass, Jasmine hedges, and date palm fronds. Results showed that, at an identical total feed rate, combined processing was more advantageous as compared to separate processing. A further sensitivity analysis on mixed combined processing showed that the cellulase enzyme price and feed price are the two major factors affecting the production cost.

Factors affecting seawater-based pretreatment of lignocellulosic date palm residues.

Seawater-based pretreatment of lignocellulosic biomass is an innovative process at research stage. With respect to process optimization, factors affecting seawater-based pretreatment of lignocellulosic date palm residues were studied for the first time in this paper. Pretreatment temperature (180°C-210°C), salinity of seawater (0ppt-50ppt), and catalysts (H2SO4, Na2CO3, and NaOH) were investigated. The results showed that pretreatment temperature exerted the largest influence on seawater-based pretreatment in terms of the enzymatic digestibility and fermentability of pretreated solids, and the inhibition of pretreatment liquids to Saccharomyces cerevisiae. Salinity showed the least impact to seawater-based pretreatment, which widens the application spectrum of saline water sources such as brines discharged in desalination plant. Sulfuric acid was the most effective catalyst for seawater-based pretreatment compared with Na2CO3 and NaOH.

Exploring the selective lactic acid production from food waste in uncontrolled pH mixed culture fermentations using different reactor configurations.

Carboxylic acid production from food waste by mixed culture fermentation is an important future waste management option. Obstacles for its implementation are the need of pH control, and a broad fermentation product spectrum leading to increased product separation costs. To overcome these obstacles, the selective production of lactic acid (LA) from model food waste by uncontrolled pH fermentation was tested using different reactor configurations. Batch experiments, semi-continuously fed reactors and a percolation system reached LA concentrations of 32, 16 and 15gCODLA/L, respectively, with selectivities of 93%, 84% and 75% on COD base, respectively. The semi-continuous reactor was dominated by Lactobacillales. Our techno-economic analysis suggests that LA production from food waste can be economically feasible, with LA recovery and low yields remaining as major obstacles. To solve both problems, we successfully applied in-situ product extraction using activated carbon.

Hydrothermal pretreatment and enzymatic hydrolysis of mixed green and woody lignocellulosics from arid regions.

Utilization of multi-specie feedstocks is imperative for application of lignocellulosic biorefineries in arid regions. Different lignocellulosic residues vary in composition and anatomical features. Pretreatment and enzymatic hydrolysis are two processes at the front end of any lignocellulosics biorefinery applying biochemical pathway, and have to efficiently deal with the variance in the feedstock composition and properties. However, there is limited knowledge about effect of mixing different lignocellulosics on pretreatment and enzymatic hydrolysis yields. In this study effect of mixing on the yields from hydrothermal pretreatment and enzymatic hydrolysis was analyzed by mixing three different lignocellulosic residues - Bermuda grass, Jasmine hedges, and date palm fronds. Results showed that the individual and the mixed lignocellulosics gave same yields when treated under similar conditions of hydrothermal pretreatment and enzymatic hydrolysis. It indicates that this mixture can be a suitable feedstock for lignocellulosic biorefinery.

Waste biorefinery in arid/semi-arid regions.

The utilization of waste biorefineries in arid/semi-arid regions is advisable due to the reduced sustainable resources in arid/semi-arid regions, e.g. fresh water and biomass. This review focuses on biomass residues available in arid/semi-arid regions, palm trees residues, seawater biomass based residues (coastal arid/semi-arid regions), and the organic fraction of municipal solid waste. The present review aims to describe and discuss the availability of these waste biomasses, their conversion to value chemicals by waste biorefinery processes. For the case of seawater biomass based residues it was reviewed and advise the use of seawater in the biorefinery processes, in order to decrease the use of fresh water.

Recovery of carboxylic acids produced during dark fermentation of food waste by adsorption on Amberlite IRA-67 and activated carbon.

Amberlite IRA-67 and activated carbon were tested as promising candidates for carboxylic acid recovery by adsorption. Dark fermentation was performed without pH control and without addition of external inoculum at 37°C in batch mode. Lactic, acetic and butyric acids, were obtained, after 7days of fermentation. The maximum acid removal, 74%, from the Amberlite IRA-67 and 63% from activated carbon was obtained from clarified fermentation broth using 200gadsorbent/Lbroth at pH 3.3. The pH has significant effect and pH below the carboxylic acids pKa showed to be beneficial for both the adsorbents. The un-controlled pH fermentation creates acidic environment, aiding in adsorption by eliminating use of chemicals for efficient removal. This study proposes simple and easy valorization of waste to valuable chemicals.

Seawater as Alternative to Freshwater in Pretreatment of Date Palm Residues for Bioethanol Production in Coastal and/or Arid Areas.

The large water consumption (1.9-5.9 m(3) water per m(3) of biofuel) required by biomass processing plants has become an emerging concern, which is particularly critical in arid/semiarid regions. Seawater, as a widely available water source, could be an interesting option. This work was to study the technical feasibility of using seawater to replace freshwater in the pretreatment of date palm leaflets, a lignocellulosic biomass from arid regions, for bioethanol production. It was shown that leaflets pretreated with seawater exhibited lower cellulose crystallinity than those pretreated with freshwater. Pretreatment with seawater produced comparably digestible and fermentable solids to those obtained with freshwater. Moreover, no significant difference of inhibition to Saccharomyces cerevisiae was observed between liquids from pretreatment with seawater and freshwater. The results showed that seawater could be a promising alternative to freshwater for lignocellulose biorefineries in coastal and/or arid/semiarid areas.

Hydrothermal Pretreatment of Date Palm (Phoenix dactylifera L.) Leaflets and Rachis to Enhance Enzymatic Digestibility and Bioethanol Potential.

Date palm residues are one of the most promising lignocellulosic biomass for bioethanol production in the Middle East. In this study, leaflets and rachis were subjected to hydrothermal pretreatment to overcome the recalcitrance of the biomass for enzymatic conversion. Evident morphological, structural, and chemical changes were observed by scanning electron microscopy, X-ray diffraction, and infrared spectroscopy after pretreatment. High glucan (>90% for both leaflets and rachis) and xylan (>75% for leaflets and >79% for rachis) recovery were achieved. Under the optimal condition of hydrothermal pretreatment (210°C/10 min) highly digestible (glucan convertibility, 100% to leaflets, 78% to rachis) and fermentable (ethanol yield, 96% to leaflets, 80% to rachis) solid fractions were obtained. Fermentability test of the liquid fractions proved that no considerable inhibitors to Saccharomyces cerevisiae were produced in hydrothermal pretreatment. Given the high sugar recovery, enzymatic digestibility, and ethanol yield, production of bioethanol by hydrothermal pretreatment could be a promising way of valorization of date palm residues in this region.

Converting the organic fraction of solid waste from the city of Abu Dhabi to valuable products via dark fermentation--Economic and energy assessment.

Landfilling the organic fraction of municipal solid waste (OFMSW) leads to greenhouse gas emissions and loss of valuable resources. Sustainable and cost efficient solutions need to be developed to solve this problem. This study evaluates the feasibility of using dark fermentation (DF) to convert the OFMSW to volatile fatty acids (VFAs), fertilizer and H2. The VFAs in the DF effluent can be used directly as substrate for subsequent bioprocesses or purified from the effluent for industrial use. DF of the OFMSW in Abu Dhabi will be economically sustainable once VFA purification can be accomplished on large scale for less than 15USD/m(3)(effluent). With a VFA minimum selling price of 330 USD/tCOD, DF provides a competitive carbon source to sugar. Furthermore, DF is likely to use less energy than conventional processes that produce VFAs, fertilizer and H2. This makes DF of OFMSW a promising waste treatment technology and biorefinery platform.

Comparison of different pretreatment strategies for ethanol production of West African biomass.

Pretreating lignocellulosic biomass for cellulosic ethanol production in a West African setting requires smaller scale and less capital expenditure compared to current state of the art. In the present study, three low-tech methods applicable for West African conditions, namely Boiling Pretreatment (BP), Soaking in Aqueous Ammonia (SAA) and White Rot Fungi pretreatment (WRF), were compared to the high-tech solution of hydrothermal pretreatment (HTT). The pretreatment methods were tested on 11 West African biomasses, i.e. cassava stalks, plantain peelings, plantain trunks, plantain leaves, cocoa husks, cocoa pods, maize cobs, maize stalks, rice straw, groundnut straw and oil palm empty fruit bunches. It was found that four biomass' (plantain peelings, plantain trunks, maize cobs and maize stalks) were most promising for production of cellulosic ethanol with profitable enzymatic conversion of glucan (>30 g glucan per 100 g total solids (TS)). HTT did show better results in both enzymatic convertibility and fermentation, but evaluated on the overall ethanol yield the low-tech pretreatment methods are viable alternatives with similar levels to the HTT (13.4-15.2 g ethanol per 100 g TS raw material).

Effect of anaerobiosis on indigenous microorganisms in blackwater with fish offal as co-substrate.

The aim of this study was to compare the effect of mesophilic anaerobic digestion with aerobic storage on the survival of selected indigenous microorganisms and microbial groups in blackwater, including the effect of addition of Greenlandic Halibut and shrimp offal. The methane yield of the different substrate mixtures was determined in batch experiments to study possible correlation between methanogenic activity in the anaerobic digesters and reduction of indigenous microorganisms in the blackwater. By the end of the experiments a recovery study was conducted to determine possible injury of the microorganisms. In both anaerobic and aerobic samples, survival of Escherichia coli was better in the presence of Greenlandic Halibut offal when compared to samples containing blackwater only and blackwater and shrimp offal, possibly due to more available carbon in the samples containing Greenlandic Halibut offal. Reduction of faecal streptococci was large under both anaerobic and aerobic conditions, and the results indicated a complete removal of faecal streptococci in the anaerobic samples containing blackwater and a mixture of blackwater and shrimp offal after 17 and 31 days, respectively. Amoxicillin resistant bacteria were reduced in the anaerobic samples in the beginning of the study but increased towards the end of it. The opposite pattern was observed in the aerobic samples, with a growth in the beginning followed by a reduction. During the anaerobic digestion tetracycline resistant bacteria showed the least reduction in the mixture of blackwater and shrimp offal, which had the lowest methane yield while the highest reduction was observed in the mixture of blackwater and Greenlandic Halibut, where the highest methane yield was measured Reduction of coliphages was larger under anaerobic conditions. Addition of fish offal had no effect on survival of coliphages. The results of the recovery study indicated that a fraction of the E. coli in the aerobic blackwater sample and of the faecal streptococci in both the anaerobic and aerobic samples containing blackwater and Greenlandic Halibut were injured only, and thus able to resuscitate during recovery. The use of anaerobic digestion in the Arctic is limited to substrate types like those tested in this study because of absence of agriculture. The results indicate that anaerobic digestion of wastewater could benefit from the addition of fish offal, with respect to both microbial reduction and energy production.

Co-production of ethanol, biogas, protein fodder and natural fertilizer in organic farming--evaluation of a concept for a farm-scale biorefinery.

The addition of a biorefinery to an organic farm was investigated, where ethanol was produced from germinated rye grains and whey, and the effluent was separated into two streams: the protein-rich solid fraction, to be used as animal feed, and the liquid fraction, which can be co-digested with clover grass silage to produce biogas. A method for ethanol production from rye was applied by utilizing inherent amylase activity from germination of the seed. Biogas potential of ethanol fermentation effluent was measured through anaerobic digestion trials. The effluent from the trials was assumed to serve as natural fertilizer. A technoeconomic analysis was also performed; total capital investment was estimated to be approximately 4 M USD. Setting a methane selling price according to available incentives for "green electricity" (0.72 USD/m(3)) led to a minimum ethanol selling price of 1.89 USD/L (project lifetime 25 yr, at a discount rate 10%).

Ex-situ bioremediation of polycyclic aromatic hydrocarbons in sewage sludge.

Polycyclic aromatic hydrocarbons (PAH) are regarded as environmental pollutants. A promising approach to reduce PAH pollution is based on the implementation of the natural potential of some microorganisms to utilize hydrocarbons. In this study Proteiniphilum acetatigenes was used for bioaugmentation of sewage sludge to improve the PAH removal. Bioaugmentation experiments were performed in parallel semi-continuously fed reactors started up with digested primary and secondary sludge. Three bioaugmentation approaches were investigated: A1, addition of bacteria once during starting up; A2, addition of bacteria at the beginning and then every 2nd day and A3, addition of encapsulated bacteria once during starting up. Removal of PAH was found to be both biotic and abiotic. All three approaches had a positive effect of the biological removal of PAH. Highest biological removal of individual PAH (up to 80%) was observed using continuous addition (approach A2) of the bacteria to the reactors. In general, the effect of bioaugmentation was higher in the reactors fed with primary sludge compared to the reactors fed with mixed sludge. Bioaugmentation resulted in biological removal of low molecular weight PAH in the reactors fed with primary sludge using all three approaches while clear biological removal of the medium- and high molecular weight PAH only was observed if the bacteria were added continuously (approach A2).

Long term studies on the anaerobic biodegradability of MTBE and other gasoline ethers.

Anaerobic biodegradation of methyl tert-butyl ether (MTBE) using electron acceptors such as nitrate, Fe(III), sulfate and bicarbonate, may be more cost effective and feasible compared to aerobic treatment methods, for dealing with the MTBE problem. Currently, there are a few reports in the literature which have documented anaerobic biodegradation of MTBE in batch studies. However, some of the reports have been controversial, additionally many other studies have failed to document anaerobic biodegradation. Experiments were conducted over a long term period in both batch and continuous reactors to investigate the anaerobic biodegradability of MTBE and other gasoline ethers. Inoculums collected from various environments were used, along with different electron acceptors. Only one set of the batch experiments showed a 30-60% conversion of MTBE to tert-butyl alcohol under Fe(III)-reducing conditions, using complexed Fe(III). The use of complexed Fe(III) created an initial low pH of 1-2 in these batches due to its acidic nature, therefore, the removal may be due to acid hydrolysis rather than biological processes. Based on the findings obtained, caution should be applied in the interpretation of experimental data in which complexed Fe(III) is used for bioremediation of MTBE.

Model description and kinetic parameter analysis of MTBE biodegradation in a packed bed reactor.

A dynamic modeling approach was used to estimate in-situ model parameters, which describe the degradation of methyl tert-butyl ether (MTBE) in a laboratory packed bed reactor. The measured dynamic response of MTBE pulses injected at the reactor's inlet was analyzed by least squares and parameter response surface methodologies. Response surfaces were found to be statistically significant and thus suitable for estimating the global minimum as well as the 95% parameter uncertainty regions. The linear parameter uncertainty estimates for the half-saturation constant (K(S)) and the maximum growth rate (micro(max)) were: 0