Pursuing single or combined wheat straw based poly(butylene succinate) production routes: A life cycle approach of first- and second-generation feedstocks

The depletion of fossil resources and the climate change crisis call for an urgent shift to production pathways based on renewable and low-carbon sources. In addition, plastic pollution worldwide motivates the identification of new sources for their bio-based counterparts, which have an increasing demand. This research aims to evaluate the environmental feasibility of different cereal-based feedstocks for the production of poly(butylene succinate) (PBS), which is obtained from the polymerisation of succinic acid (SA) and 1,4 butanediol (BDO) monomers. The baseline scenario analysed corresponds to the use of wheat straw as a source of the fermentable sugars. Furthermore, five other cereal-based production routes combining first-generation (1G) feedstocks such as wheat and maize grain, and second-generation (2G) feedstocks, such as sorghum, barley straw, and maize stover, combined with wheat straw, were evaluated. The Life Cycle Assessment (LCA) methodology was used to identify the main hotspots of these valorisation routes at the early stage of the biorefinery design, considering all the burden categories provided by the ReCiPe impact method. The results showed that the straw-based PBS profile reached a Global Warming Potential of 3.43 kg CO2eq, whereas a range value from 2.34 to 7.27 kg CO2eq was estimated when wheat straw is combined with sorghum and barley straw, respectively. The pre-treatment stage represents a substantial impact on the strategy considered to produce fermentable sugars, particularly, for barley straw. Therefore, improvements are still required to reduce the energy demand and increase the sugar yieldThis research is supported by the project Enhancing diversity in Mediterranean cereal farming systems (CerealMed), funded by PRIMA Programme and FEDER/Ministry of Science and Innovation– Spanish National Research Agency (PCI2020-111978) and the project Transition to sustainable agri-food sector bundling life cycle assessment and ecosystem services approaches (ALISE), funded by the Spanish National Research Agency (TED2021-130309B-I00). R.R.L., M.T.M., S.G.G. belong to the Galician Competitive Research Group (GRC ED431C-2021/37) and to the Cross-disciplinary Research in Environmental Technologies (CRETUS Research Center, ED431E 2018/01)S

Synthesis and Characterization of Bacterial Cellulose from Citrus-Based Sustainable Resources

Citrus juices from whole oranges and grapefruits (discarded from open market) and aqueous extracts from citrus processing waste (mainly peels) were used for bacterial cellulose production by Komagataeibacter sucrofermentans DSM 15973. Grapefruit and orange juices yielded higher bacterial cellulose concentration (6.7 and 6.1 g/L, respectively) than lemon, grapefruit, and orange peels aqueous extracts (5.2, 5.0, and 2.9 g/L, respectively). Compared to the cellulosic fraction isolated from depectinated orange peel, bacterial cellulose produced from orange peel aqueous extract presented improved water-holding capacity (26.5 g water/g, 3-fold higher), degree of polymerization (up to 6-fold higher), and crystallinity index (35-86% depending on the method used). The presence of absorption bands at 3240 and 3270 cm-1 in the IR spectrum of bacterial cellulose indicated that the bacterial strain K. sucrofermentans synthesizes both Iα and Iβ cellulose types, whereas the signals in the 13C NMR spectrum demonstrated that Iα cellulose is the dominant type

Structure and function of tetrapyrrole biosynthesis enzymes: ALA synthase and cobalt chelatase

Modified tetrapyrroles are versatile compounds that are universally utilized by enzymes as co-factors in a myriad of enzyme cata lyzed reaction s. Examples include heme, the cofactor of hemoglobin and myoglobin, that carries and stores molecular oxygen in the blood and muscle respectively, and cobalamin a cofactor that is synthesized by prokaryotes and which is an important nutrient for humans. This work focuses on the enzymatic mechanism of metal chelation and in particular on the structural characterization of the cobalt-chelatases CbiK from Salmonella enterica and CbiX from Archaeoglobus /u/gidus that take part in the biosynthesis of cobalamin. Crystal structures of the enzyme-tetrapyrrole complex were obtained that reveal radically different modes of binding compared to the well characterized ferrochelatases. Furthermore protein structures reveal the evolutionary re lationships between cobaltochelatases from different organisms. The second part of this thesis is focusing on 5-aminolevulinic acid synthase (ALAS), an enzyme that catalyzes the synthesis of the first common precursor in t he biosynthesis of all tetrapyrroles, 5-aminolevulinic acid (ALA). Mutations in the C-terminal region of the human erythroid specific ALAS (ALAS2) lead to X-linked dominant protoporphyria a condition that is characterized by the accumulation of protoporphyrin in red blood cells leading to photosensitivity and liver disease. The erythroid specific ALAS2 and its disease - causing variants have been characterized using steady state kinetic methods to investigate the molecular basis of the disorder. Furthermore the "house-keeping" ALASl isoenzyme was characterized as well and the effect of mutations equivalent to the disease causing ALAS2 mutations have been tested on the enzyme. Chimeric bacterial enzymes that have been engineered to contain the human erythroid specific C-terminal extension were characterized as an alternative to understanding the role of the C-terminal extension in regulating the activity of the human ALASl and ALAS2EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Βιομηχανικές διεργασίες και σχεδιασμός της παραγωγής ηλεκτρικού οξέος μέσω συνεχών μικροβιακών ζυμώσεων με χρήση αποβλήτων που προέρχονται από βιομηχανίες παραγωγής χαρτοπολτού

The scope of this thesis is the evaluation of spent sulphite liquor (SSL) as the main fermentation carbon source for succinic acid (SA) production in continuous cultures. Subsequently, a techno-economic analysis of a process that produces SA from SSL under continuous and fed-batch fermentations strategies is presented. The purpose of the techno-economic analysis is to investigate the economic feasibility of the succinic acid that derived from the utilization of SSL and factors in which the succinic acid production cost has high dependence.SSL is the liquid by-product stream derived from the acidic sulphite process of mainly hardwoods. This waste stream contains solubilised lignin in the form of lignosulphonates and sugars which arise after hemicelluloses degradation. The type of the using wood and the processing conditions employed in the plant are the two main elements that define the sugars contained in SSL. In this study the waste stream that was used resulting from the pulp and paper industry of the Eucalyptus globulus wood.This work was initiated with the evaluation of SSL as substrate in continuous cultures using the wild-type rumen bacterial strains Actinobacillus succinogenes and Basfia succiniciproducens. These strains are considered as two of the most promising strains for industrial implementation. The cultures were initially carried out at constant dilution rate (0.04 h-1) and varying initial commercial xylose concentrations (23-55 g/L) or constant xylose concentration (40 g/L) and varying dilution rates (0.02-0.25 h-1) showing that dilution rates of 0.02-0.15 h-1 led to satisfactory succinic acid production by both microbial strains. In continuous cultures using nanofiltrated SSL, maximum yields of SA were achieved at dilution rate of 0.02 h-1 (0.48 g/g for A. succinogenes and 0.55 g/g for B. succiniciproducens) while maximum values of productivity were obtained at dilution rate of 0.04 h-1 in the case of A. succinogenes (0.67 g/L/h) and 0.1 h-1 (1.6 g/L/h) in the case of B. succiniciproducens. During the fermentation of both strains observed biofilm creation at the solid parts of the reactor. Due to this the biomass concentration was impossible to measure, so the biomass concentrations at different dilution rates for both strains was estimated using metabolic flux analysis. In the techno-economic analysis of SA, the design of the entire SA production process was based on data obtained from experimental results and the material and energy balances were carried out using spreadsheets and validated through UniSim software. Bacterial fermentations were conducted with Basfia succiniciproducens as the SA producer. The down stream design based on direct crystallization process using ion-exchange resins for acidification step, with final purified SA crystals equal to 99% and with a recovery yield of 97% which achieved by recirculation of the side sterams in down stream process. Process simulation was performed for a range of plant capacities, 5, 30, 100 kt of SA per year. The study indicated the feasibility of fermentative SA production from SSL and the minimum selling price (MSP) for 10% annual return on investment was calculated at 2.70 $/kg for continuous and 3.06$/kg for fed-batch strategy when commercial nitrogen sources were applied. The MSP was reduced to 2.16 $/kg and 2.76$/kg for continuous and fed-batch respectively assuming that commercial nitrogen sources are substituted with hydrolysates of corn steep liquor (CSL). The fermentation and evaporation units were identified as significant capital and operating cost contributors and therefore the optimization of bioreactor geometrical characteristic and alternative more cost-effective evaporation technics should be considered in future research.Στόχος της παρούσας διατριβής υπήρξε η αξιολόγηση του παράπλευρου ρεύματος που παράγεται από βιομηχανίες χαρτοπολτού (SSL), ως κύρια πρώτη ύλη για την παραγωγή ηλεκτρικού οξέος, μέσω μικροβιακών ζυμώσεων σε συνθήκες συνεχούς καλλιέργειας σε βιοαντιδραστήρα. Στα πλαίσια της επίτευξης οικονομικότερων διεργασιών, εκπονήθηκε τεχνοοικονομική ανάλυση των διεργασιών παραγωγής ηλεκτρικού οξέος, με χρήση του SSL ως πρώτη ύλη, σε συνεχείς και ημισυνεχείς ζυμώσεις. Απώτερος στόχος του εγχειρήματος αυτού ήταν η διερεύνηση της οικονομικής βιωσιμότητας της παραγωγής ηλεκτρικού οξέος αξιοποιώντας το SSL ως πρώτη ύλη, καθώς και η εύρεση στοιχείων που υποδεικνύουν τους οικονομικούς παράγοντες που σχετίζονται με την παραγωγή του προϊόντος. Το SSL αποτελεί το κύριο παράπλευρο ρεύμα που προκύπτει μετά την όξινη διαδικασία που πραγματοποιείται για την παραγωγή χαρτοπολτού κυρίως σε σκληρά ξύλα. Το υποπροϊόν αυτό περιέχει διαλύτη λιγνίνη υπό την μορφή λιγνοσουλφονικών αλάτων καθώς και σάκχαρα τα οποία προκύπτουν κατά την αποικοδόμηση της ημικυτταρίνης. Το είδος του χρησιμοποιούμενου ξύλου και οι συνθήκες επεξεργασίας αποτελούν τα δύο κύρια σημεία που καθορίζουν το είδος και την περιεκτικότητα των σακχάρων που περιέχονται στο SSL. Στη μελέτη αυτή, χρησιμοποιήθηκε το υποπροϊόν που προέκυψε από τη βιομηχανία επεξεργασίας του φυτού Eucalyptus globulus.Σαν πρώτο στάδιο της μελέτης, αξιολογήθηκε η ικανότητα χρήσης του SSL ως υπόστρωμα σε συνεχείς ζυμώσεις, χρησιμοποιώντας τα βακτηριακά στελέχη Actinobacillus succinogenes και Basfia succiniciproducens. Τα στελέχη αυτά θεωρούνται ως τα δύο πιο υποσχόμενα στελέχη για βιομηχανική εφαρμογή. Αρχικά, οι ζυμώσεις διεξήχθησαν με σταθερό ρυθμό αραίωσης ίσο με 0,04 h-1 και διαφορετικές αρχικές συγκεντρώσεις εμπορικής ξυλόζης (23-55 g/L) ή σταθερή συγκέντρωση ξυλόζης (40 g/L) και διαφορετικές τιμές ρυθμών αραίωσης (0,02-0,25 h-1). Τα αποτελέσματα αυτών των συνθηκών έδειξαν ότι και τα δύο στελέχη παρείχαν ικανοποιητική παραγωγή ηλεκτρικού οξέος σε ένα εύρος ρυθμών αραίωσης μεταξύ 0,02-0,15 h-1. Στις συνεχείς καλλιέργειες με χρήση SSL το οποίο είχε προεπεξεργαστεί με νανοδιήθηση, επετεύχθησαν μέγιστες αποδόσεις ηλεκτρικού οξέος σε ρυθμό αραίωσης 0,02 h-1. Συγκεκριμένα το στέλεχος A. succinogenes είχε απόδοση 0,48 g/g και το στέλεχος B. Succiniciproducens 0,55 g/g. Ωστόσο, η μέγιστη παραγωγικότητα του A. Succinogenes (0,67 g/L/h) παρουσιάστηκε με ρυθμό αραίωσης 0,04 h-1 ενώ στην περίπτωση του B. succiniciproducens η μέγιστη παραγωγικότητα ίση με 1,6 g/L/h επιτεύχθηκε σε ρυθμό αραίωσης 0,1 h-1. Κατά την πραγματοποίηση των ζυμώσεων και στα δύο στελέχη, παρατηρήθηκε η δημιουργία φυσικής ακινητοποίησης του μικροοργανισμού στα μέρη του βιοαντιδραστήρα. Εξαιτίας αυτού, η συγκέντρωση της βιομάζας καθίστανται αδύνατο να μετρηθεί επομένως οι συγκεντρώσεις της βιομάζας σε διαφορετικούς ρυθμούς αραίωσης και για τα δύο στελέχη προσδιορίστηκε με χρήση ανάλυσης μεταβολικής ροής. Για την τεχνοοικονομική ανάλυση του ηλεκτρικού οξέος, ο σχεδιασμός όλης της διεργασίας βασίστηκε στα πειραματικά αποτελέσματα ενώ τα ισοζύγια μάζας και ενέργειας πραγματοποιήθηκαν λύνοντας εξισώσεις που διέπουν οι νόμοι της θερμοδυναμικής και επαληθεύτηκαν μέσω του λογισμικού UniSim. Οι ζυμώσεις για την παραγωγή του ηλεκτρικού οξέος στα αποτελέσματα των οποίων βασίστηκαν οι υπολογισμοί, διενεργήθηκαν με τον μικροοργανισμό Basfia succiniciproducens. Ο πειραματικός σχεδιασμός για την παραλαβή του προϊόντος βασίστηκε στη διαδικασία της απευθείας κρυστάλλωσης του ηλεκτρικού οξέος όπου χρησιμοποιήθηκαν ρητίνες ιοντοανταλλαγής ως μέσω για την μετατροπή των οργανικών αλάτων σε οργανικά οξέα. Οι κρύσταλλοι ηλεκτρικού οξέος που προκύπτουν μέσω αυτής της διαδικασίας έχουν καθαρότητα ίση με 99% ενώ η ανάκτηση του προϊόντος τέθηκε ως 97%. Ο μεγάλος συντελεστής ανάκτησης του προϊόντος οφείλεται στην περαιτέρω επεξεργασία των παράπλευρων ρευμάτων που προκύπτουν από την συνολική διεργασία διαχωρισμού. Κατά την τεχνοοικονομική ανάλυση έγινε προσομοίωση της παραγωγής ηλεκτρικού οξέος για συνεχείς και ημισυνεχείς ζυμώσεις σε τρεις διαφορετικές δυναμικότητες παραγωγής (5, 30 και 100 kt) ηλεκτρικού οξέος ανά έτος. Σκοπός της μελέτης ήταν να αξιολογήσει την οικονομική βιωσιμότητα της βιοχημικής παραγωγής ηλεκτρικού οξέος με χρήση SSL ως πηγής άνθρακα. Από την μελέτη προέκυψε ως ελάχιστη τιμή πώλησης (MSP) ηλεκτρικού οξέος τα 2,70 $/kg στις συνεχείς ζυμώσεις και 3,06$/kg στις ημισυνεχείς με τη χρήση εμπορικών πηγών αζώτου. Περεταίρω μελέτη για την μείωση των ελάχιστων τιμών πώλησης έδειξε ότι η χρήση πηγών αζώτου προερχόμενων από υδρολύματα θα μείωνε το κόστος πώλησης στα 2,16 $/kg και 2,76$/kg, στις συνεχείς και ημισυνεχείς ζυμώσεις αντίστοιχα. Τα στάδια της ζύμωσης καθώς και η εξάτμιση αποδείχθηκαν σημαντικοί οικονομικοί παράγοντες όσον αφορά το λειτουργικό κόστος και το κόστος του κεφαλαίου. Επομένως, η βελτιστοποίηση των γεωμετρικών χαρακτηριστικών του βιοαντιδραστήρα καθώς και εναλλακτικές πιο οικονομικά προσιτές τεχνικές εξάτμισης, θα πρέπει να εξεταστούν σε μελλοντική έρευνα

Direct electrochemical extraction increases microbial succinic acid production from spent sulphite liquor

The fermentative production of succinic acid leads to increasing toxicity over time, and requires base addition to counteract acidification. Here we integrated a fed-batch Basfia succiniciproducens succinic acid fermentation with membrane electrolysis. This approach brings the broth in direct contact with an OH- and H-2 producing cathode, and enables in situ extraction of succinate towards the low volume H+ and O-2 producing anode compartment. In the latter, the succinate is acidified to succinic acid and precipitated. The key advantage of the cathodic process is the production of base maintaining fermentation pH with limited external base addition. The bacterial cells are recycled through the cathode compartment of the electrochemical cell, and exposed to H-2, creating biological reducing power. Fermentations were executed with glucose, xylose and ultrafiltered spent sulphite liquor (SSL), the side stream generated via acidic sulphite pulping of Eucalyptus globulus wood. The membrane was not permeable to cells, color and solids, which resulted in combined succinate extraction, clarification, acidification and concentration in a single unit operation. The succinic acid to by-product ratio increased, favoring the production of succinic acid over lactic, formic and acetic acids, with up to 15% higher total sugars to succinic acid conversion yield. The OH-production due to water reduction resulted in lower NaOH usage (up to 33% less) for maintaining the pH during fermentation. The maximum productivity also increased by 30% to a rate of 0.41 g L-1 h(-1) with the electrochemical system, with 1.65 kW h per kg succinic acid extracted in the case of SSL and 2.4 and 1.9 kW h per kg succinic acid extracted in the case of glucose and xylose, respectively. Considering the aforementioned advantages, this integrated system could evolve into a breakthrough technology in sustainable industrial succinic acid production from crude industrial side streams

Evaluation of 1,3-propanediol production by two Citrobacter freundii strains using crude glycerol and soybean cake hydrolysate

Biodiesel production processes using soybean as feedstock generates soybean cake and crude glycerol as by-products. These by-product streams were used as sole feedstocks for the production of 1,3-propanediol (PDO) using two bacterial strains of Citrobacter freundii. Soybean cake has been converted into a nutrient-rich hydrolysate by crude enzymes produced via solid state fermentation. The effect of initial glycerol and free amino nitrogen concentration on bacterial growth and PDO production has been evaluated in batch bioreactor cultures showing that C. freundii VK-19 is a more efficient PDO producer than C. freundii FMCC-8. The cultivation of C. freundii VK-19 in fed-batch bioreactor cultures using crude glycerol and soybean cake hydrolysates led to PDO concentration of 47.4 g/L with yield and productivity of 0.49 g/g and 1.01 g/L/h, respectively. The effect of PDO, metabolic by-products, and sodium and potassium salts on bacterial growth was evaluated showing that potassium salts initially enhance bacterial growth, whereas sodium salts cause significant inhibition to bacterial growth. Soybean cake hydrolysate and crude glycerol could be utilized for PDO production, but the fermentation efficiency is influenced by the catalyst used during biodiesel production

Bacterial Cellulose Production from Industrial Waste and by-Product Streams

The utilization of fermentation media derived from waste and by-product streams from biodiesel and confectionery industries could lead to highly efficient production of bacterial cellulose. Batch fermentations with the bacterial strain Komagataeibacter sucrofermentans DSM (Deutsche Sammlung von Mikroorganismen) 15973 were initially carried out in synthetic media using commercial sugars and crude glycerol. The highest bacterial cellulose concentration was achieved when crude glycerol (3.2 g/L) and commercial sucrose (4.9 g/L) were used. The combination of crude glycerol and sunflower meal hydrolysates as the sole fermentation media resulted in bacterial cellulose production of 13.3 g/L. Similar results (13 g/L) were obtained when flour-rich hydrolysates produced from confectionery industry waste streams were used. The properties of bacterial celluloses developed when different fermentation media were used showed water holding capacities of 102–138 g·water/g·dry bacterial cellulose, viscosities of 4.7–9.3 dL/g, degree of polymerization of 1889.1–2672.8, stress at break of 72.3–139.5 MPa and Young’s modulus of 0.97–1.64 GPa. This study demonstrated that by-product streams from the biodiesel industry and waste streams from confectionery industries could be used as the sole sources of nutrients for the production of bacterial cellulose with similar properties as those produced with commercial sources of nutrients

Several serum lipid metabolites are associated with relapse risk in pediatric-onset multiple sclerosis.

BackgroundThe circulating metabolome is altered in multiple sclerosis (MS), but its prognostic capabilities have not been extensively explored. Lipid metabolites might be of particular interest due to their multiple roles in the brain, as they can serve as structural components, energy sources, and bioactive molecules. Gaining a deeper understanding of the disease may be possible by examining the lipid metabolism in the periphery, which serves as the primary source of lipids for the brain.ObjectiveTo determine if altered serum lipid metabolites are associated with the risk of relapse and disability in children with MS.MethodsWe collected serum samples from 61 participants with pediatric-onset MS within 4 years of disease onset. Prospective longitudinal relapse data and cross-sectional disability measures (Expanded Disability Status Scale [EDSS]) were collected. Serum metabolomics was performed using untargeted liquid chromatography and mass spectrometry. Individual lipid metabolites were clustered into pre-defined pathways. The associations between clusters of metabolites and relapse rate and EDSS score were estimated utilizing negative binomial and linear regression models, respectively.ResultsWe found that serum acylcarnitines (relapse rate: normalized enrichment score [NES] = 2.1, q = 1.03E-04; EDSS: NES = 1.7, q = 0.02) and poly-unsaturated fatty acids (relapse rate: NES = 1.6, q = 0.047; EDSS: NES = 1.9, q = 0.005) were associated with higher relapse rates and EDSS, while serum phosphatidylethanolamines (relapse rate: NES = -2.3, q = 0.002; EDSS: NES = -2.1, q = 0.004), plasmalogens (relapse rate: NES = -2.5, q = 5.81E-04; EDSS: NES = -2.1, q = 0.004), and primary bile acid metabolites (relapse rate: NES = -2.0, q = 0.02; EDSS: NES = -1.9, q = 0.02) were associated with lower relapse rates and lower EDSS.ConclusionThis study supports the role of some lipid metabolites in pediatric MS relapses and disability

Evolution in a family of chelatases facilitated by the introduction of active site asymmetry and protein oligomerization

The class II chelatases associated with heme, siroheme, and cobalamin biosynthesis are structurally related enzymes that insert a specific metal ion (Fe2+ or Co2+) into the center of a modified tetrapyrrole (protoporphyrin or sirohydrochlorin). The structures of two related class II enzymes, CbiXS from Archaeoglobus fulgidus and CbiK from Salmonella enterica, that are responsible for the insertion of cobalt along the cobalamin biosynthesis pathway are presented in complex with their metallated product. A further structure of a CbiK from Desulfovibrio vulgaris Hildenborough reveals how cobalt is bound at the active site. The crystal structures show that the binding of sirohydrochlorin is distinctly different to porphyrin binding in the protoporphyrin ferrochelatases and provide a molecular overview of the mechanism of chelation. The structures also give insights into the evolution of chelatase form and function. Finally, the structure of a periplasmic form of Desulfovibrio vulgaris Hildenborough CbiK reveals a novel tetrameric arrangement of its subunits that are stabilized by the presence of a heme b cofactor. Whereas retaining colbaltochelatase activity, this protein has acquired a central cavity with the potential to chaperone or transport metals across the periplasmic space, thereby evolving a new use for an ancient protein subunit