BIOFILM REACTORS FOR INTENSIVE PRODUCTION OF BICARBOXYLIC ACIDS

The study carried out during the present Ph.D. program aimed at investigating succinic acid production by fermentation. The work was carried out at the Dipartimento di Ingegneria Chimica, dei Materiale e della Produzione Industriale of the Università degli Studi di Napoli ‘Federico II’. The commercial strain Actinobacillus succinogenes DSM 792 was selected for its ability to produce succinic acid at high concentration and yield from a broad variety of carbon source. The activities were articulated along the two paths described hereinafter. Characterization of the succinic acid fermentation process as regards the concentration and yields The study was aimed at the assessment of the fermentation performance - in terms of succinic acid concentration and yield – with reference to two classes of feedstocks: 1) sugars typically present in lignocellulosic hydrolysates (glucose, mannose, arabinose and xylose); 2) high sugar content beverages (HSCBs). The investigations were carried out with batch fermentation of free A. succinogenes cells at different sugar concentration. The conversion process was characterized in terms of metabolite production (succinic acid, formic acid and acetic acid), sugar conversion, pH and cell concentration. The study included tests with single sugar and sugar mixtures (synthetic and commercial): i) glucose, mannose, arabinose and xylose (GMAX) to mime the composition of a lignocellulosic hydrolysate; ii) sucrose, glucose and fructose (SGF), the sugars typically present in leftover beverages. All the investigated sugars were converted in succinic acid by A. succinogenes. Tests with mixtures pointed out that the fermentation took advantage of the co-presence of different sugars with respect to the tests carried out with single sugars. The synergy effect of the sugars led to an increase in final succinic acid concentration and SA selectivity. Development of a continuous process for the succinic acid production Continuous fermentation tests carried out in a CSTR were aimed to the systematic characterization of the kinetics of cell growth and succinic acid production. The continuous fermentation were carried out by feeding the CSTR with a glucose bearing medium at a wide interval of dilution rate. The kinetics of A. succinogenes growth on glucose was investigated by assessing the effects of glucose and acids - succinic, acetic, and formic - on cell growth. The experimental results were successfully correlated by a multiple product-inhibited interacted growth model. Experimental results pointed out the individual inhibitory effect of the acids on A. succinogenes growth. Under the investigated operating conditions, results pointed out that: no substrate inhibition was observed; acetic acid did not inhibit the cell growth and the succinic acid production. The activity aimed at the intensive succinic acid production regarded the design, set-up, operation and optimisation of a packed bed biofilm reactor (PBBR). The effects of the operating conditions - dilution rate (D) and medium compositions (glucose, GX, xylose, GAX) - on the PBBR performances were investigated. Succinic acid concentration, productivity and sugar(s) conversion generally decreased with D. A maximum succinic acid productivity of 35.0 g/L*h was achieved at a D= 1.9 h-1. The effect of HMF and furfural, the main inhibitors found in lignocellulosic hydrolysate, on succinic acid production was also investigated. HMF resulted to have a higher inhibition effect on succinic acid production compared to furfural

Bioreactors for succinic acid production processes

Succinic acid (SA) has been recognized as one of the most important bio-based building block chemicals due to its numerous potential applications. Fermentation SA production from renewable carbohydrate feedstocks can have the economic and sustainability potential to replace petroleum-based production in the future, not only for existing markets, but also for new larger volume markets. Design and operation of bio-reactors play a key role. During the last 20 years, many different fermentation strategies for SA production have been described in literature, including utilization of immobilized biocatalysts, integrated fermentation and separation systems and batch, fed-batch, and continuous operation modes. This review is an overview of different fermentation process design developed over the past decade and provides a perspective on remaining challenges for an economically feasible succinate production processes. The analysis stresses the idea of improving the efficiency of the fermentation stage by improving bioreactor design and by increasing bioreactor performance.</p

Continuous Succinic Acid Fermentation by Actinobacillus Succinogenes: Assessment of Growth and Succinic Acid Production Kinetics

Succinic acid is one of the most interesting platform chemicals that can be produced in a biorefinery approach. The paper reports the characterization of the growth kinetics of Actinobacillus succinogenes DSM 22257 using glucose as carbon source. Tests were carried out in a continuous bioreactor operated under controlled pH. Under steady-state conditions, the conversion process was characterized in terms of concentration of glucose, cells, acids, and pH. The effects of acid—succinic, acetic, and formic—concentration in the medium on fermentation performance were investigated. The fermentation was interpreted according to several models characterized by substrate and product inhibition. The selected kinetic model of biomass growth and of metabolite production described the microorganism growth rate under a broad interval of operating conditions. Under the investigated operating conditions, results pointed out that: no substrate inhibition was observed; acetic acid did not inhibit the cell growth and succinic acid productio

Biosuccinic Acid from Lignocellulosic-Based Hexoses and Pentoses by Actinobacillus succinogenes: Characterization of the Conversion Process

Succinic acid (SA) is a well-established chemical building block. Actinobacillus succinogenes fermentation is by far the most investigated route due to very promising high SA yield and titer on several sugars. This study contributes to include the SA production within the concept of biorefinery of lignocellulose biomass. The study was focused on the SA production by A. succinogenes DSM 22257 using sugars representative from lignocellulose hydrolysis— glucose, mannose, arabinose, and xylose—as carbon source. Single sugar batch fermentation tests and mixture sugar fermentation tests were carried out. All the sugars investigated were converted in succinic acid by A. succinogenes. The best fermentation performances were measured in tests with glucose as carbon source. The bacterial growth kinetics was characterized by glucose inhibition. No inhibition phenomena were observed with the other sugar investigated. The sugar mixture fermentation tests highlighted the synergic effects of the copresence of the four sugars. Under the operating conditions tested, the final concentration of succinic acid in the sugar mixture test was larger (27 g/L) than that expected (25.5 g/L) by combining the fermentation of the single sugar. Moreover, the concentration of acetic and formic acid was lower, consequently obtaining an increment in the succinic acid specificity

Succinic acid production from hexoses and pentoses by fermentation of actinobacillus succinogenes

Succinic acid is a precursor for many chemicals in the food, pharmaceutical, cosmetic, and biodegradable plastics industries. Nowadays, the biotechnological route to produce succinic acid is focused on processes based on the fermentation of natural materials characterized by high fraction of carbohydrates. Actinobacillus succinogenes is proposed in the literature as a promising strain for the production of succinic acid. Indeed, A. succinogenes may utilize a wide spectrum of carbohydrates as carbon sources. Moreover, the CO2 required for the fermentation allows to include the biotechnological route among the carbon capture and conversion processes. A key issue of fermentation processes aimed at producing succinic acid is the feedstock cost. Pre-requisites of the feedstock are: availability at high mass flow rate over all the year, and non-competitive with (human and animal) food. Lignocellulosic biomass is a potential feedstock because it fulfils the listed pre-requisites. This contribution reports the characterization of succinic acid fermentation by A. succinogenes 130Z adopting as carbon source sugars representative of the hydrolysis of lignocellulosic biomass: glucose, mannose, arabinose, and xylose. Batch fermentation tests were carried out using single sugar as carbon source for a systematic characterization of the conversion process of the investigated sugars. The conversion process was characterized in terms of concentration of biomass, sugars, and acids as well as of pH. The time-resolved data were processed to assess the sugar conversion, the succinic acid yield, and the productivity for each of the investigated sugars. A. succinogenes was able to convert all investigated sugars into succinic acid using MgCO3 as an indirect CO2 source. Glucose was the sugar characterized by the best performance when the initial concentration was set at 40 g/L. The performances did not depend on the investigated sugars if the initial concentration was quite low (<5 g/L)

Continuous succinic acid fermentation by Actinobacillus succinogenes in a packed-bed biofilm reactor

Background: Succinic acid is one of the most interesting platform chemicals that can be produced in a biorefinery approach. In this study, continuous succinic acid production by Actinobacillus succinogenes fermentation in a packed-bed biofilm reactor (PBBR) was investigated. Results: The effects of the operating conditions tested, dilution rate (D), and medium composition (mixture of glucose, xylose, and arabinose - that simulate the composition of a lignocellulosic hydrolysate) - on the PBBR performances were investigated. The maximum succinic acid productivity of 35.0 g L-1 h-1 and the maximum SA concentration were achieved at a D = 1.9 h-1. The effect of HMF and furfural on succinic acid production was also investigated. HMF resulted to reduce succinic acid production by 22.6%, while furfural caused a reduction of 16% in SA production at the same dilution rate. Conclusion: Succinic acid production by A. succinogenes fermentation in a packed-bed reactor (PBBR) was successfully carried out for more than 5 months. The optimal results were obtained at the dilution rate 0.5 h-1: 43.0 g L-1 of succinic acid were produced, glucose conversion was 88%; and the volumetric productivity was 22 g L-1 h-1.</p

Efficient succinic acid production from high-sugar-content beverages by Actinobacillus succinogenes

This study presents the production of succinic acid (SA) by Actinobacillus succinogenes using high-sugar-content beverages (HSCBs) as feedstock. The aim of this study was the valorization of a by-product stream from the beverage industry for the production of an important building block chemical, such as SA. Three types of commercial beverages were investigated: fruit juices (pineapple and ace), syrups (almond), and soft drinks (cola and lemon). They contained mainly glucose, fructose, and sucrose at high concentration—between 50 and 1,000 g/L. The batch fermentation tests highlighted that A. succinogenes was able to grow on HSCBs supplemented with yeast extract, but also on the unsupplemented fruit juices. Indeed, the bacteria did not grow on the unsupplemented syrup and soft drinks because of the lack of indispensable nutrients. About 30–40 g/L of SA were obtained, depending on the type of HSCB, with yield ranging between 0.75 and 1.00 gSA/gS. The prehydrolysis step improved the fermentation performance: SA production was improved by 6–24%, depending on the HSCB, and sugar conversion was improved of about 30–50%.</p

MOESM1 of Continuous succinic acid fermentation by Actinobacillus succinogenes in a packed-bed biofilm reactor

Additional file 1: Figure S1. Biofilm of A. succinogenes. a) at the end of the start-up phase; b) after 5 months of continuous operation. Figure S2. Time-course profiles of the fermentation results during the production phase from glucose. a Glucose (▼) and cell concentration (○) and dilution rate (dashed line); b succinic (∆), acetic (●) and formic (□) acid concentration and dilution rate. Figure S3. Time-course profiles of the fermentation results during the adaptation phase from glucose to xylose. a Glucose (▼), xylose (■) and cell concentration (○); b succinic (∆), acetic (●), and formic (□) acid concentration and xylose percentage (dashed line) in the medium. The dilution rate was set to 1.24 h−1. Figure S4. Time-course profiles of the fermentation results during the production phase from xylose. a Xylose (▼) and cell concentration (○) and dilution rate (dashed line); b succinic (∆), acetic (●), and formic (□) acid concentration and dilution rate. Figure S5. Time-course profiles of the fermentation results during the production phase from GAX medium. a Sugars [glucose (▼), xylose (■), and arabinose (∇)] and cell concentration (○) and dilution rate (dashed line); b succinic (∆), acetic (●), and formic (□) acid concentration and dilution rate