Techno-economic optimization of a process superstructure for lignin valorization.

Lignin, the most abundant aromatic biopolymer on Earth, is often considered a biorefinery by-product, despite its potential to be valorized into high-added-value chemicals and fuels. In this work, an integrated superstructure-based optimization model was set up and optimized using mixed-integer non-linear programming for the conversion of technical lignin to three main biobased products: aromatic monomers, phenol-formaldehyde resins, and aromatic aldehydes/acids. Several alternative conversion pathways were simultaneously compared to assess the profitability of lignins-based processes by predicting the performance of technologies with different TRL. Upon employing key technologies such as hydrothermal liquefaction, dissolution in solvent, or high-temperature electrolysis, the technical lignins could have a market value of 200 €/t when the market price for aromatic monomers, resins, and vanillin is at least 2.0, 0.8, and 15.0 €/kg, respectively. When lower product selling prices were considered, the aromatic monomers and the resins were not profitable as target products.

Modelling of the Citric Acid Production from Crude Glycerol by Wild-Type Yarrowia lipolytica DSM 8218 Using Response Surface Methodology (RSM).

Crude glycerol is the main by-product of the biodiesel manufacturing industry (10% w/w). Its use as a substrate in microbial fermentations is a concrete strategy to efficiently address its market surplus. In this study, the conversion of crude glycerol to citric acid, a key biochemical in the emerging bioeconomy, by a wild-type yeast Yarrowia lipolytica DSM 8218 was modelled using the Response Surface Methodology. The model relates C/N mass ratio and crude glycerol concentration to maximize the citric acid yield in flask scale using two different N sources, yeast extract and ammonium sulphate. Under the optimal conditions (yeast extract, C/N 141, glycerol 33 g/L), the conversion yield was 0.249 g/g. The optimal conditions were used for up-scaling a fed-batch fermentation in a 2 L bioreactor highlighting a metabolic shift from mannitol to citric acid when high stirring rates were applied (800 rpm). In these conditions, a morphic transition from pseudo-mycelial form to round-shaped yeast-like cells was observed too.

From paper mill waste to single cell oil: Enzymatic hydrolysis to sugars and their fermentation into microbial oil by the yeast Lipomyces starkeyi.

Single cell oil (SCO) represents an outstanding alternative to both fossil sources and vegetable oils from food crops waste. In this work, an innovative two-step process for the conversion of cellulosic paper mill waste into SCO was proposed and optimised. Hydrolysates containing glucose and xylose were produced by enzymatic hydrolysis of the untreated waste. Under the optimised reaction conditions (Cellic® CTec2 25 FPU/g glucan, 48 h, biomass loading 20 g/L), glucose and xylose yields of 95 mol% were reached. The undetoxified hydrolysate was adopted as substrate for a batch-mode fermentation by the oleaginous yeast Lipomyces starkeyi. Lipid yield, content for single cell, production and maximum oil productivity were 20.2 wt%, 37 wt%, 3.7 g/L and 2.0 g/L/d respectively. This new generation oil, obtained from a negative value industrial waste, represents a promising platform chemical for the production of biodiesel, biosurfactants, animal feed and biobased plastics.

Optimisation of glucose and levulinic acid production from the cellulose fraction of giant reed (Arundo donax L.) performed in the presence of ferric chloride under microwave heating.

A two-step exploitation of the giant reed cellulose to glucose and levulinic acid, after the complete removal of the hemicellulose fraction, was investigated using FeCl3 as catalyst. In the first step, the microwave-assisted hydrolysis of cellulose to glucose was optimised by response surface methodology analysis, considering the effect of temperature, reaction time and catalyst amount. Under the optimised reaction conditions, the glucose yield was 39.9 mol%. The cellulose-rich residue was also converted by enzymatic hydrolysis, achieving the glucose yield of 39.8 mol%. The exhausted residue deriving from the chemical hydrolysis was further converted to levulinic acid by microwave treatment at harsher reaction conditions. The maximum levulinic acid yield was 64.3 mol%. On the whole, this cascade approach ensured an extensive and sustainable exploitation of the C6 carbohydrates to glucose and levulinic acid, corresponding to about 70 mol% of glucan converted to these valuable bioproducts in the two steps.

Single Cell Oil Production from Undetoxified Arundo donax L. hydrolysate by Cutaneotrichosporon curvatus.

The use of low-cost substrates represents one key issue to make single cell oil production sustainable. Among low-input crops, Arundo donax L. is a perennial herbaceous rhizomatous grass containing both C5 and C6 carbohydrates. The scope of the present work was to investigate and optimize the production of lipids by the oleaginous yeast Cutaneotrichosporon curvatus from undetoxified lignocellulosic hydrolysates of steam-pretreated A. donax. The growth of C. curvatus was first optimized in synthetic media, similar in terms of sugar concentration to hydrolysates, by applying the response surface methodology (RSM) analysis. Then the bioconversion of undetoxified hydrolysates was investigated. A fed-batch process for the fermentation of A. donax hydrolysates was finally implemented in a 2-L bioreactor. Under optimized conditions, the total lipid content was 64% of the dry cell weight and the lipid yield was 63% of the theoretical. The fatty acid profile of C. curvatus triglycerides contained 27% palmitic acid, 33% oleic acid and 32% linoleic acid. These results proved the potential of lipid production from A. donax, which is particularly important for their consideration as substitutes for vegetable oils in many applications such as biodiesel or bioplastics.

Hydrolysis of Corn Stover by Talaromyces cellulolyticus Enzymes: Evaluation of the Residual Enzymes Activities Through the Process.

The obtainment of sugars from lignocellulosic residues represents a sustainable and versatile platform for the production of a number of bio-based products. Cellulases are a family of enzymes which can effectively hydrolyze the biomass polysaccharides at mild conditions. Cellulolytic fungi belonging to the genera Trichoderma and Aspergillus are the most commonly source of commercial cellulases used so far. More recently, Talaromyces cellulolyticus was also scored as a promising cellulases producer. In comparison to the Trichoderma and Aspergillus systems, Talaromyces enzymes have been less investigated. The present research dealt with the conversion of steam-pretreated corn stover by commercial blend of T. cellulolyticus enzymes with respect to the common blends. The paper also investigated the stability of the enzyme preparation and tested the use of additives (namely Tween 80, Tween 20, and BSA) to improve the enzymes performances and the hydrolysis efficiency. The results indicated that, at the same process conditions, T. cellulolyticus cellulases were more effective and yielded 20% more sugars compared to control blends. Furthermore, the cellulase components displayed a synergistic interaction with hemicellulases. The results indicate that cellulases from T. cellulolyticus are less affected by the high dry matter consistency and the use of additives could increase the total activity by around 50% and ß-glucosidase capacity by 10-15%.

Yeast lipids from cardoon stalks, stranded driftwood and olive tree pruning residues as possible extra sources of oils for producing biofuels and biochemicals.

BACKGROUND: Some lignocellulosic biomass feedstocks occur in Mediterranean Countries. They are still largely unexploited and cause considerable problems due to the lack of cost-effective harvesting, storage and disposal technologies. Recent studies found that some basidiomycetous yeasts are able to accumulate high amount of intracellular lipids for biorefinery processes (i.e., biofuels and biochemicals). Accordingly, the above biomass feedstocks could be used as carbon sources (after their pre-treatment and hydrolysis) for lipid accumulation by oleaginous yeasts. RESULTS: Cardoon stalks, stranded driftwood and olive tree pruning residues were pre-treated with steam-explosion and enzymatic hydrolysis for releasing free mono- and oligosaccharides. Lipid accumulation tests were performed at two temperatures (20 and 25 °C) using Leucosporidium creatinivorum DBVPG 4794, Naganishia adeliensis DBVPG 5195 and Solicoccozyma terricola DBVPG 5870. S. terricola grown on cardoon stalks at 20 °C exhibited the highest lipid production (13.20 g/l), a lipid yield (28.95%) close to the maximum theoretical value and a lipid composition similar to that found in palm oil. On the contrary, N. adeliensis grown on stranded driftwood and olive tree pruning residues exhibited a lipid composition similar to those of olive and almonds oils. A predictive evaluation of the physical properties of the potential biodiesel obtainable by lipids produced by tested yeast strains has been reported and discussed. CONCLUSIONS: Lipids produced by some basidiomycetous yeasts grown on Mediterranean lignocellulosic biomass feedstocks could be used as supplementary sources of oils for producing biofuels and biochemicals.

Bioethanol production from steam-pretreated corn stover through an isomerase mediated process.

Agricultural by-products such as corn stover are considered strategic raw materials for the production of second-generation bioethanol from renewable and non-food sources. This paper describes the conversion of steam-pretreated corn stover to ethanol utilising a multi-step process including enzymatic hydrolysis, isomerisation, and fermentation of mixed hydrolysates with native Saccharomyces cerevisiae. An immobilised isomerase enzyme was used for the xylose isomerisation along with high concentrations of S. cerevisiae. The objective was to assess the extent of simultaneity of the various conversion steps, through a detailed analysis of process time courses, and to test this process scheme for the conversion of lignocellulosic hydrolysates containing several inhibitors of the isomerase enzyme (e.g. metal ions, xylitol and glycerol). The process was tested on two types of hydrolysate after acid-catalysed steam pretreatment: (a) the water soluble fraction (WSF) in which xylose was the largest carbon source and (b) the entire slurry, containing both cellulose and hemicellulose carbohydrates, in which glucose predominated. The results indicated that the ethanol concentration rose when the inoculum concentration was increased in the range 10-75 g/L. However, when xylose was the largest carbon source, the metabolic yields were higher than 0.51g(ethanol)/g(consumed) sugars probably due to the use of yeast internal cellular resources. This phenomenon was not observed in the fermentation of mixed hydrolysates obtained from the entire pretreated product and in which glucose was the largest carbon source. The ethanol yield from biomass suspensions with dry matter (DM) concentrations of 11-12% (w/v) was 70% based on total sugars (glucose, xylose, galactose). The results suggest that xylulose uptake was more effective in mixed hydrolysates containing glucose levels similar to, or higher than, xylose. Analysis of the factors that limit isomerase activity in lignocellulosic hydrolysates excluded any inhibition due to residual calcium ions after the detoxification of the hemicellulose hydrolysates with Ca(OH)2. By contrast, most of the enzyme activity ceased during the fermentation of the entire slurry after steam explosion, probably due to synergistic inhibition effects of various fermentation co-products.

Bioethanol production from mixed sugars by Scheffersomyces stipitis free and immobilized cells, and co-cultures with Saccharomyces cerevisiae.

Bioethanol can be produced from several biomasses including lignocellulosic materials. Besides 6-carbon sugars that represent the prevalent carbohydrates, some of these feedstocks contain significant amounts of 5-carbon sugars. One common limit of the major part of the xylose-fermenting yeasts is the diauxic shift between the uptake of glucose and xylose during the fermentation of mixed syrups. Thus, optimized fermentation strategies are required. In this paper the ability of Scheffersomyces stipitis strain NRRLY-11544 to ferment mixed syrups with a total sugar concentration in the range 40-80 g/L was investigated by using mono cultures, co-cultures with Saccharomyces cerevisiae strain Bakers Yeast Type II and single cultures immobilized in silica-hydrogel films. The experimental design for the fermentations with immobilized cells included the process analysis in function of two parameters: the fraction of the gel in the broth and the concentration of the cells loaded in the gel. Furthermore, for each total sugars level, the fermentative course of S. stipitis was analyzed at several glucose-to xylose ratios. The results indicated that the use of S. stipitis and S. cerevisiae in free co-cultures ensured faster processes than single cultures of S. stipitis either free or immobilized. However, the rapid production of ethanol by S. cerevisiae inhibited S. stipitis and caused a stuck of the process. Immobilization of S. stipitis in silica-hydrogel increased the relative consumption rate of xylose-to-glucose by 2-6 times depending on the composition of the fermentation medium. Furthermore the films performances appeared stable over three weeks of continuous operations. However, on the whole, the final process yields obtained with the immobilized cells were not meaningfully different from that of the free cells. This was probably due to concurrent fermentations operated by the cells released in the broth. Optimization of the carrier characteristics could improve the performances of the process with immobilized cells.

Ethanol production in immobilized-cell bioreactors from mixed sugar syrups and enzymatic hydrolysates of steam-exploded biomass.

We investigated ethanol production from mixed sugar syrups. Hydrolysates were prepared from enzymatic saccharification of steam-pretreated aspen chips. Syrups containing 45 g/L of glucose and 12 g/L of xylose were detoxified through two ion-exchange resins and then fermented with Pichia stipitis and Saccharomyces cerevisiae immobilized in Ca-alginate gel beads. Combinations of different gel fractions in the fermentation volume, amount of yeast cells, and ratios of P. stipitis vs S. cerevisiae within each bead were compared. In the best conditions, by using a total beads volume corresponding to 25% of the working volume, we obtained a yield of 0.39 g(ethanol)/g(initial sugars). This amount of gel entrapped an initial cell concentration of 6 x 1012cells/L with ratio of S. cerevisiae/P. stipitis of 0.25 g/g. Modified stirred-tank reactors were obtained either by adding marbles or by inserting a perforated metal cylinder, which reduced considerably the rupture of beads while visibly improving oxygenation of the medium.